key: cord- -jr yj o authors: forrest, d l; thompson, k; dorcas, v g; couban, s h; pierce, r title: low molecular weight heparin for the prevention of hepatic veno-occlusive disease (vod) after hematopoietic stem cell transplantation: a prospective phase ii study date: - - journal: bone marrow transplant doi: . /sj.bmt. sha: doc_id: cord_uid: jr yj o we evaluated patients undergoing high-dose chemo/radiotherapy (hdct) and hematopoietic stem cell transplantation (hsct) (allogeneic ( ), autologous ( )) to determine the safety and feasibility of administering low molecular weight heparin (lmwh) as hepatic veno-occlusive disease (vod) prophylaxis. patients received a once daily subcutaneous injection of dalteparin anti-xa i.u. commencing the day prior to starting hdct, and continuing until day + post hsct or hospital discharge, whichever came first. dosage adjustments were made for patients developing renal failure. all bleeding episodes were recorded and graded and vod was diagnosed and graded according to seattle criteria. at days of follow-up, the overall survival and probability of regimen-related mortality were and %, respectively. nine patients developed vod. the probability of developing vod post allogeneic and autologous hsct was % ( % ci, – ) and % ( % ci, – ), respectively. vod was graded as moderate (n= ) and severe (n= ). vod resolved in all cases except for one patient who died secondary to severe vod and multiorgan failure. clinically significant bleeding episodes occurred in three patients; patients developed minor bleeding not requiring specific therapy. all bleeding episodes resolved. these results suggest that lmwh for vod prophylaxis is safe with a low incidence of serious bleeding events. whether it is superior to unfractionated heparin, however, is unknown and should be addressed within the context of a randomized controlled trial. we evaluated patients undergoing high-dose chemo/ radiotherapy (hdct) and hematopoietic stem cell transplantation (hsct) (allogeneic ( ) , autologous ( ) ) to determine the safety and feasibility of administering low molecular weight heparin (lmwh) as hepatic veno-occlusive disease (vod) prophylaxis. patients received a once daily subcutaneous injection of dalteparin anti-xa i.u. commencing the day prior to starting hdct, and continuing until day + post hsct or hospital discharge, whichever came first. dosage adjustments were made for patients developing renal failure. all bleeding episodes were recorded and graded and vod was diagnosed and graded according to seattle criteria. at days of follow-up, the overall survival and probability of regimen-related mortality were and %, respectively. nine patients developed vod. the probability of developing vod post allogeneic and autologous hsct was % ( % ci, - ) and % ( % ci, - ), respectively. vod was graded as moderate (n ¼ ) and severe (n ¼ ). vod resolved in all cases except for one patient who died secondary to severe vod and multiorgan failure. clinically significant bleeding episodes occurred in three patients; patients developed minor bleeding not requiring specific therapy. all bleeding episodes resolved. these results suggest that lmwh for vod prophylaxis is safe with a low incidence of serious bleeding events. whether it is superior to unfractionated heparin, however, is unknown and should be addressed within the context of a randomized controlled trial. bone marrow transplantation ( ) , - . doi: . /sj.bmt. keywords: low molecular weight heparin; vod high-dose chemo/radiotherapy (hdct) and hematopoietic stem cell transplantation (hsct) are widely used to treat many malignant and nonmalignant conditions. regimen-related toxicity (rrt) remains a frequent and major complication of hsct. hepatic veno-occlusive disease (vod) is one of the most common life-threatening rrts reported to occur in - % of patients undergoing hsct. [ ] [ ] [ ] [ ] [ ] [ ] [ ] clinically, vod is characterized by jaundice, painful hepatomegaly, ascites and weight gain from fluid retention. in its most severe form, it often leads to multiorgan failure and death. [ ] [ ] [ ] hepatic vod is thought to result from injury to hepatocytes surrounding the central veins in zone of the liver acinus. early pathologic changes include edema of the subendothelium of hepatic venules, followed by thrombosis of the venular lumen. risk factors for vod have been reported to include a previous history of viral hepatitis, abnormal liver function tests at the time of hdct and hsct, the use of high-dose preparative regimens, mismatched or unrelated donor hscts, and the use of vancomycin or amphotericin b. [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] a number of prophylactic strategies have been explored in an attempt to prevent the injury and subsequent thrombosis of hepatic venules and therefore decrease the risk of vod. they include the administration of prostaglandin e , pentoxifylline, ursodeoxycholic acid and low-dose standard unfractionated heparin (uh). [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] the results of clinical trials using these prophylactic agents have been quite variable and thus their role in vod prophylaxis remains controversial. low-molecular weight heparin (lmwh) is formed by depolymerization of uh. in several large clinical trials, when administered as treatment or prophylaxis of deep venous thrombosis (dvt), it has been found to be as efficacious as uh with a low incidence of bleeding complications. [ ] [ ] [ ] there are, however, few published reports specifically examining the safety and efficacy of lmwh as vod prophylaxis. our centre therefore undertook a prospective pilot study of lmwh as vod prophylaxis for patients undergoing hdct and hsct. between october and november , patients underwent hdct and hsct (allogeneic ( ) , autologous ( ) ) at the queen elizabeth ii health sciences centre (qeii hsc), the quaternary referral centre for atlantic canada. patient characteristics are listed in table . eligibility criteria included: ( ) all patients undergoing hsct for malignant or nonmalignant disease excluding aplastic anemia; ( ) age x and p years; and ( ) satisfactory end-organ function. exclusion criteria were: ( ) significant renal dysfunction (measured h creatinine clearance o ml/min); ( ) previous history of heparininduced thrombocytopenia (hit); ( ) active bleeding or lesions at risk of bleeding at study enrollment where systemic anticoagulation was felt to be contraindicated; and ( ) the use of any other anticoagulant or antithrombotic drugs during the first days post-hsct except for locally administered anticoagulants or thrombolytic agents in central venous catheters to maintain patency. the study protocol was approved by the institutional review board at the qeii hsc and all patients provided written informed consent for study participation. the majority of patients received either busulfan or total body irradiation-based conditioning regimens. they included: busulfan mg/kg and cyclophosphamide mg/ kg (n ¼ ); etoposide mg/kg and melphalan mg/m (n ¼ ); melphalan mg/m and fractionated total body irradiation (ftbi) cgy  (n ¼ ); cyclophosphamide mg/kg and ftbi cgy  (n ¼ ); melphalan mg/m (n ¼ ); etoposide . g/m , cyclophosphamide mg/kg and ftbi cgy  (n ¼ ); and cyclophosphamide mg/kg and ftbi cgy  (n ¼ ). twenty two patients underwent an allogeneic hsct. the stem cell source for these patients was a histocompatible sibling in patients and an hla a, b, c and drmatched unrelated donor (using high-resolution molecular dna typing) in three patients. eighteen patients underwent autologous hsct. stem cells were collected in all autologous hsct recipients and from all sibling donors by apheresis of g-csf mobilized peripheral blood; stem cells were collected from unrelated donors by standard bone marrow harvest. the median absolute cd cell count infused to allogeneic recipients was .  /kg (range . - . ) and to autologous recipients was .  /kg (range . - . ). graft-versus-host disease (gvhd) prophylaxis gvhd prophylaxis for allogeneic hematopoietic stem cell recipients consisted of intravenous cyclosporine a (csa) and short-course methotrexate (mtx). treatment of acute gvhd consisted of high-dose corticosteroids, with the addition of antithymocyte globulin (atg) in those patients not responding to corticosteroids. all patients were nursed in high-efficiency particulate air (hepa) filtered, positive-pressure rooms until engraftment (absolute neutrophil count (anc) .  /l). tunneled triple lumen hickman central venous catheters were employed. irradiated blood products were administered to keep the morning hemoglobin (hgb) concentration g/l and morning platelet count x  /l. cmvnegative blood products were administered to cmv seronegative donor/recipient pairs. infection prophylaxis included low-dose acyclovir in all patients; fluconazole prophylaxis was administered only to those patients undergoing allogeneic hsct; antibacterial prophylaxis was not used, but intravenous antibiotics were initiated for febrile neutropenia. trimethaprim/sulphamethoxazole was commenced as pcp prophylaxis in allogeneic stem cell recipients from the point of engraftment until immunosuppression was discontinued. fluconazole or amphotericin b was used as required for documented or suspected fungal infection. the primary objective of the study was to determine the safety and feasibility of lmwh administration to hsct recipients. therefore, careful assessments of bleeding events, red cell and platelet transfusion requirements, time to white cell and platelet engraftment and adverse events were made. the secondary objective was to determine the incidence and severity of vod for patients undergoing hsct. once daily at h, patients received a single subcutaneous injection of dalteparin anti-xa i.u. (supplied as prefilled, single-dose syringes from pharmacia, canada) commencing the day prior to starting hdct and continuing until day + post hsct or hospital discharge, whichever came first. dosage adjustments were made in cases of renal failure as follows: patients developing moderate to severe renal insufficiency (calculated creatinine clearance p ml/min) received a % dose reduction of dalteparin (ie, anti-xa i.u.). in those patients requiring dialysis for renal failure, dalteparin was not administered. if renal function subsequently improved, then dalteparin administration was recommenced. patients were reevaluated at day + to determine hsct outcome and whether cases of documented vod had resolved. daily patient evaluation while receiving dalteparin included patient weight, oxygen saturation, liver size, presence of right upper quadrant (ruq) abdominal pain, ascites or pleural effusions, complete blood count (cbc), serum creatinine, assessment of bleeding and gvhd; adverse events, grading of rrt and the number of red cell and platelet transfusions were recorded; liver function tests (bilirubin, ast, alt, alkaline phosphatase), inr, ptt and fibrinogen were measured twice weekly. patient evaluation at day + included patient weight, liver size, presence of ruq abdominal pain, cbc, ptt, inr, fibrinogen, serum creatinine and liver function tests. vod was diagnosed according to seattle criteria requiring the presence of at least two of the following features before day + following stem cell reinfusion: ( ) hyperbilirubinemia (total serum bilirubin mmol/l) and ( ) the presence of hepatomegaly or ruq abdominal pain or ( ) unexplained weight gain ( % baseline body weight) or ascites in the absence of other identifiable causes. the severity of vod was graded as mild, moderate or severe according to modified seattle criteria ( ): ( ) mild ¼ total serum bilirubin mmol/l but o mmol/l and weight gain % over baseline but o % or hepatomegaly or ruq abdominal pain; ( ) moderate ¼ total serum bilirubin x mmol/l but o mmol/l or weight gain x % over baseline or documented ascites; ( ) severe ¼ total serum bilirubin x mmol/l or ascites compromising respiratory function or hepatic encephalopathy or death from vod. all bleeding episodes were recorded and graded according to the following criteria: grade ¼ no bleeding; grade ¼ minor mucosal bleeding or petechiae not requiring packed red blood cell (prbc) transfusion; grade ¼ any bleeding episode requiring transfusion of - u of prbcs/ episode in a -h period; grade ¼ any bleeding episode requiring transfusion of u of prbcs/episode, but o u in a -h period or retroperitoneal bleeding; grade ¼ any bleeding causing hemodynamic instability or requiring transfusion of x u of prbcs in a -h period or any cns bleeding. treatment was discontinued or interrupted for patients meeting the following criteria: ( ) days post-hsct limit; ( ) discharge from hospital; ( ) development of dialysisdependent renal failure; ( ) patient request; ( ) serious bleeding episode (xgrade ) at the discretion of the investigator; ( ) development of a thrombotic event necessitating treatment with another anticoagulant or higher dose of lmwh. estimates of overall survival, regimen-related mortality and the probability of vod were calculated using the methods of kaplan and meier. in all, patients were enrolled in the study. there were males and females. the median age at hsct was y (range - ) and the median time from diagnosis to hsct was months (range - ). diagnoses included: non-hodgkin's lymphoma ( ), multiple myeloma ( ), acute myeloid leukemia ( ), myelodysplastic syndrome ( ), myelofibrosis ( ), chronic myeloid leukemia ( ), hodgkin's disease ( ) and acute lymphoblastic leukemia ( ) . of these, patients underwent allogeneic hsct (related donor ( ) , unrelated donor ( )) and patients underwent autologous hsct. all patients had normal liver function tests (lfts) prior to commencing hdct except for two patients (chronic active hepatitis c infection (n ¼ ), lymphomatous involvement of the liver (n ¼ )); two other patients had a history of hepatic disease prior to hsct, but their lfts were normal at the time of hdct. the median duration of dalteparin administration was days (range - ); two patients required dalteparin dose reduction for renal impairment. all patients completed the study as per the protocol; early termination (before day + or hospital discharge) occurred in three patients because of dialysis-dependent renal failure (n ¼ ) or death (n ¼ ). at days of follow-up, the overall survival and the probability of regimen-related mortality was % ( % ci, - ) and % ( % ci, respectively. six patients have died (cardiac arrest ( ), multiorgan failure ( ), infection ( ), respiratory failure ( ) and demyelinating peripheral neuropathy ( )) on day + , + , + , + , + and + post hsct respectively. two patients (upn , ) had disease progression on or before day + . in total, patients engrafted with a median time to anc .  /l and sustained platelet  /l of days (range - ) and days (range - ), respectively; two patients (upn , ) died before engraftment on days + and + , respectively. rrt was xgrade iii in one or more organ systems in five patients ( . %). two patients (upn and ) developed lung toxicity requiring intubation and mechanical ventilation because of idiopathic interstitial pneumonitis; upn improved with high-dose corticosteroid therapy and was extubated, but he eventually died day + secondary to abdominal sepsis; upn continued to deteriorate despite treatment with high-dose corticosteroids and died of respiratory failure on day + . upn developed acute renal failure (arf) secondary to druginduced acute tubular necrosis necessitating a brief course of dialysis before renal function recovered. upn died of sudden cardiac arrest on day + . autopsy failed to reveal a specific cause of death. finally upn died on day + of arf, hepatic vod and acute respiratory distress syndrome (ards). all five patients who developed xgrade iii toxicity had undergone allogeneic hsct (sibling ( ), unrelated donor ( )). rrt was pgrade ii for all autologous hsct recipients. nine patients developed vod (autologous ( ), allogeneic ( )). the probability of developing vod post allogeneic and autologous hsct was % ( % ci, - ) and % ( % ci, - ), respectively ( figure ). patient characteristics and outcome are detailed in table . five of the nine patients had been heavily pre-treated with x lines of prior chemotherapy with or without concomitant radiotherapy. five patients had received tbi in the conditioning regimen, three of which underwent unrelated donor allogeneic hsct. one patient had chronic active hepatitis c infection, one patient had positive viral serology for hepatitis b antibody indicating prior hepatitis b infection, one patient had lymphomatous involvement of the liver at the time of hsct and one other patient had a history of drug-induced hepatitis that had resolved prior to commencing hdct. vod was graded as moderate in eight patients and severe in one patient (upn ). vod was diagnosed a median of days (range - ) post hsct and the median peak serum bilirubin was mol/l (range - ). vod resolved in all cases before day post hsct except for upn who died secondary to severe vod and subsequent multiorgan failure days following hsct. clinically significant bleeding episodes (xgrade ii) occurred in three patients (grade ii ( ), grade iv ( )), none of whom had concomitant vod. upn developed vaginal bleeding on day + following a sibling allogeneic hsct for relapsed non-hodgkin's lymphoma. cbc revealed a hgb of g/l and a platelet count of  /l. she was transfused with u of prbcs and u of random donor platelets. the bleeding resolved in h and no further bleeding occurred. upn developed melena stool day + following a sibling allogeneic hsct for myelofibrosis. the cbc at the time of bleeding showed a hgb of g/l and platelet count of  /l. he was transfused with u of prbcs and another u of prbcs h later. the bleeding subsequently resolved and no further gastrointestinal bleeding was noted. upn developed bleeding per rectum secondary to hemorrhoids on day + following a sibling allogeneic hsct for myelodysplastic syndrome. cbc at this time revealed a hgb of g/l and platelet count of  /l. she was transfused with u of prbcs and the bleeding resolved. a total of patients developed minor bleeding (grade ) consisting of petechiae, purpura or minor mucosal bleeding. these episodes did not require specific therapy and all resolved. platelet transfusion thresholds remained at  /l for all patients in the study, with the exception of the three patients, with clinically significant bleeding episodes. for these three patients, an attempt was made to keep the platelet count greater than  /l until the bleeding episode resolved, at which point the threshold was reduced to  /l. none of the bleeding episodes, including the three that were xgrade ii, were considered severe enough to contraindicate further anticoagulant prophylaxis for vod and dalteparin administration was continued in all cases. hepatic vod is one of the most common life-threatening complications of hdct and hsct. the incidence in the literature varies from - % depending on a number of patient characteristics such as disease status at the time of hsct, previous therapy, pre-existing liver disease etc as well as the type of transplant and prophylactic strategy utilized. [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] in its most severe form, it often leads to multiorgan failure and death . despite encouraging reports utilizing defibrotide for established severe vod following hsct, the mortality remains high and effective prophylactic strategies are needed. hepatic vod is believed to result from injury to the vascular endothelium of small hepatic venules, induced by the hdct conditioning regimen. , the endothelial injury is often followed by superimposed thrombosis and progressive fibrosis of the venular lumens. a number of prophylactic strategies have been explored in an attempt to prevent the endothelial damage and subsequent thrombosis including antioxidants, antiinflammatory agents, hydrophilic bile salts and various anticoagulants with varying success - . a recent randomized prospective trial has suggested that ursodeoxycholic acid administration postallogeneic hsct decreases the incidence of hepatic complications. this benefit, however, appears to be because of a reduction in the incidence of acute gvhd and nonrelapse mortality and not because of a reduction in the incidence of vod. therefore, its specific role in vod prophylaxis may be limited. the use of standard low-dose uh as vod prophylaxis has been the subject of several early studies, , - only one of which was a prospective randomized controlled trial. two of these uncontrolled trials showed an apparent benefit for uh, , however, the study by bearman et al using various doses of heparin resulted in a high incidence of bleeding or anticipated bleeding ( %) and a high incidence of vod ( %), bringing in question the efficacy of uh in vod prophylaxis. there has been only one prospective randomized controlled trial of uh in vod prophylaxis reported by attal et al. in that study, patients undergoing hdct and autologous or allogeneic hsct were randomly assigned to receive uh u/kg/day as a continuous intravenous infusion commencing day À until day + post hsct or no prophylaxis. uh administration appeared to be efficacious with a reduction in the incidence of vod from . % in the patients receiving no prophylaxis to . % in the patients receiving uh, without an increase in bleeding episodes. this trial has been criticized, however, for the small number of patients ( %) at high risk of vod before commencing hdct, none of whom developed vod irrespective of whether they received heparin or not. therefore, despite the apparent overall positive results of the attal trial, the role of uh remains controversial and the majority of subsequent studies examining the role of uh in vod prophylaxis have not shown a clear benefit for uh. , lmwh has a number of potential advantages when compared to uh including greater bioavailability and longer plasma half-life because of less plasma protein binding, a more predictable anticoagulant response, no need for laboratory monitoring, a low incidence of hit and in some studies, a lower incidence of bleeding. [ ] [ ] [ ] [ ] there have been a number of clinical trials comparing the efficacy and safety of lmwh to uh in the prophylaxis and treatment of dvt in medical and surgical patients. [ ] [ ] [ ] [ ] these studies have consistently shown that lmwh is both efficacious and safe with a low incidence of bleeding complications. all of these qualities potentially make lmwh an attractive anticoagulant agent in the hematopoietic stem cell transplant setting where the majority of patients experience a variable period of thrombocytopenia and increased risk of bleeding. there are few published studies, however, specifically examining the role of lmwh as vod prophylaxis for patients undergoing hdct and hsct. in a recent pilot study by or et al, patients undergoing hsct were randomized to receive lmwh (enoxaparin mg subcutaneous daily) or placebo. the duration of hyperbilirubinemia and the incidence of hepatomegaly were lower in the enoxaparin group compared to placebo. in another study by the ebmt examining the incidence and outcome of vod following hdct and hsct, of patients received lmwh as vod prophylaxis. the low number of patients receiving lmwh precluded an adequate evaluation of its prophylactic effect. our pilot study has shown that lmwh administration to patients undergoing hsct is safe with a low number of serious bleeding episodes. only three of ( . %) patients developed significant bleeding requiring transfusion of prbcs and ( %) patients developed minor mucosal bleeding or petechiae. no patients died of hemorrhage or suffered any permanent sequelae because of bleeding. these results are in keeping with the - % incidence of serious bleeding reported in the literature for patients undergoing hdct and hsct irrespective of whether they are receiving low doses of systemic anticoagulants. , , in the pilot study reported by bearman et al comparing the incidence of vod and severity of bleeding between a cohort of patients receiving low-dose uh and a control group of patients, the incidence of major bleeding in those patients receiving uh was % and all but one patient displayed evidence of minor bleeding. the site and severity of bleeding, however, was not statistically different between the uh and control groups. similarly, in the study reported by or et al where patients undergoing hsct were randomized between lmwh and placebo, the incidence and duration of bleeding was in fact lower in those patients receiving lmwh. this would support the fact that patients undergoing hdct and hsct are at an inherent risk of bleeding because of the development of obligate thrombocytopenia and damaged mucosal surfaces. the administration of low-dose uh or lmwh does not appear to further increase the risk of serious hemorrhage in this patient population and the need for maintaining higher platelet transfusion thresholds in those patients receiving low-dose systemic anticoagulants does not appear to be warranted. our pilot study was designed to primarily examine the safety and feasibility of lmwh (dalteparin iu daily) administration to patients undergoing hsct. the small number of patients, the lack of randomization and risk stratification made a determination of efficacy difficult. despite these limitations, the incidence of vod in our study was % and only one patient developed severe vod. these results are encouraging when compared to the - % reported incidence rates from large published studies where no specific vod prophylaxis was utilized. , , three of the patients ( %) undergoing autologous hsct is this study developed moderate vod. while this incidence is higher than what is commonly reported in the literature for this patient population, our results may have been affected by the small number of patients undergoing autologous hsct, in addition to specific patient characteristics, which may influence the incidence of vod within individual studies. finally, the diagnosis of mildmoderate vod, accounting for eight of the nine patients in our study, can sometimes be difficult to distinguish from other causes of mild hepatic dysfunction such as infection, gvhd and various medications, and this may lead to an overestimate of the true incidence of vod in this patient population. , in summary, this pilot study has demonstrated that the administration of lmwh (dalteparin anti-xa i.u. daily) to patients undergoing hdct and hsct is well tolerated and safe with a low incidence of serious bleeding events. the efficacy of lmwh compared to uh for vod prophylaxis, however, remains controversial and whether either preparation is superior to no prophylaxis is unknown. this should be addressed within the context of a prospective randomized placebo controlled trial. regimenrelated toxicity in patients undergoing bone marrow transplantation venocclusive disease of the liver after bone marrow transplantation: diagnosis, incidence, and predisposing factors venocclusive disease of the liver following bone marrow transplantation venocclusive disease of the liver after chemoradiotherapy and autologous bone marrow transplantation hepatic venocclusive disease in autologous bone marrow transplantation of solid tumors and 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european group for blood and marrow transplantation use of prostaglandin e for prevention of liver veno-occlusive disease in leukaemic patients treated by allogeneic bone marrow transplantation prevention of regimenrelated toxicities after bone marrow transplantation by pentoxifylline: a prospective, randomized trial a randomized controlled trial of pentoxifylline for the prevention of regimenrelated toxicities in patients undergoing allogeneic marrow transplantation ursodiol prophylaxis against hepatic complications of allogeneic bone marrow transplantation: a randomized, double-blind, placebocontrolled trial ursodeoxycholic acid for the prevention of hepatic complications in allogeneic stem cell transplantation preventive role of heparin for liver veno-occlusive disease (vod) a pilot study of continuous infusion heparin for the prevention of hepatic veno-occlusive disease after bone marrow transplantation prophylactic heparin does not prevent liver veno-occlusive disease following autologous bone marrow transplantation prevention of hepatic venoocclusive disease after bone marrow transplantation by continuous infusion of low-dose heparin: a prospective randomized trial prevention of venoocclusive disease of the liver after bone marrow transplantation: heparin or no heparin? low molecular weight heparin low molecular weight heparin given the evening before surgery compared with conventional low dose heparin in prevention of thrombosis a randomized double-blind study between a low molecular weight heparin kabi and standard heparin in the prevention of deep vein thrombosis in general surgery treatment of venous thrombosis with intravenous unfractionated heparin administered in the hospital as compared with subcutaneous low-molecular-weight heparin administered at home nonparametric estimation from incomplete observations multi-institutional use of defibrotide in patients after stem cell transplantation with severe veno-occlusive disease and multisystem organ failure: response without significant toxicity in a high-risk population and factors predictive of outcome heparin-induced thrombocytopenia in patients treated with low-molecularweight heparin or unfractionated heparin low molecular weight heparin for the prevention of veno-occlusive disease of the liver in bone marrow transplantation patients acute bleeding and thrombocytopenia after bone marrow transplantation we acknowledge the contribution of the medical and nursing staff of a/b ward and medical day unit at the qeii health sciences centre. key: cord- -duwfxeyt authors: dzieciątkowski, tomasz; przybylski, maciej; torosian, tigran; tomaszewska, agnieszka; Łuczak, mirosław title: prevalence of human herpesvirus antibodies and dna in allogeneic stem cell transplant patients: two-year single centre experience date: - - journal: arch immunol ther exp (warsz) doi: . /s - - - sha: doc_id: cord_uid: duwfxeyt introduction: human herpesvirus (hhv- ) has been recognized as a potentially significant pathogen in hemopoietic stem cell transplant (hsct) recipients. different clinical manifestations have been described, including fever, skin rash, bone marrow suppression, and encephalitis. materials and methods: a retrospective review of a group of adult recipients of allogeneic hscts was conducted. serum samples taken before transplant were examined for the presence of specific anti-hhv- igm and igg antibodies. after transplantation, quantitative real-time pcr was used to determine viral load in plasma samples from days – post-transplant. results: hhv- dna was detected in plasma samples in ( %) of the recipients between days and after transplantation. all of them developed fever of unknown origin and over % had graft-versus-host disease features. three individuals from this group died during detectable hhv- viremia. another two recipients showed a single positive pcr result at a later time. infection with hhv- was thus confirmed in ( . %) of the graft recipients. conclusions: there is a high frequency of detectable hhv- viral load in stem cell transplant recipients in poland. further investigation to monitor hhv- reactivation in graft recipients will be important to improve outcome for these patients. human herpesvirus type (hhv- ) belongs to the subfamily β-herpesvirinae and is closely related to hhv- and hhv- (formerly known as cytomegalovirus -cmv) [ ] . most people have had a primary hhv- infection in early childhood [ ] and the virus is widespread in the human population, as shown by the presence of specific antibodies in % of healthy adults [ ] . as a result of the primary infection, hhv- is assumed to establish a latent infection and viral dna can be detected in salivary glands [ ] , monocytes [ ] , and early bone marrow progenitor cells [ ] . hhv- is reported to be a causative agent of exanthema subitum [ ] and has been associated with a broad spectrum of diseases, including febrile convulsions [ ] , encephalopathy [ , ] , hepatitis [ ] , and lymphoproliferative disorders [ ] . an active hhv- infection can cause fatal disease in an immunocompromised host, including patients after stem cell transplantations (scts), but fatal outcome in immunocompetent persons due to hhv- infection is extremely rare [ , ] . hhv- presence in plasma or serum was documented in - % of hemopoietic stem cell transplant (hsct) recipients using molecular techniques [ ] . the peak viral load occurs early after transplantation and usually within the first four weeks after transplantation [ , ] . allogeneic sct, advanced hematological disease, young age, gender mismatch between the donor and recipient, and treatment with corticosteroids are commonly reported risk factors associated with hhv- infection after transplantation [ , ] . in a recent study we summarized retrospective results of the determination of hhv infection status in allogeneic stem cell transplant recipients. as infections with hhv- are rarely accompanied by clinical symptoms, our first aim was to compare hhv- infection status, measured as viral dna presence in the blood in the post-transplantation period and patients' anti-hhv- serological status by detection of igg and igm antibodies. this allowed us to determine both the presence of active infection and whether a recent infection was a result of primary contact with the virus or the reactivation of latent virus, which should be valuable information in terms of comparing hhv- infection status and clinical status of an infected patient. another virus belonging to the β-herpesvirinae subfamily, hhv- (cmv), is one from the most important viral pathogens causing clinical infections in patients receiving immunosuppressive treatment. as there is information in published data about a connection between hhv- and hhv- reactivation in graft recipients, our aim was to determine the appearance of simultaneous reactivation of both β-herpesviruses. we believe that this is probably the first report from poland involving this kind of examination in hsct recipients. this retrospective study involved patients who received an allogeneic hsct and were hospitalized at the hematology, oncology, and internal medicine clinics, medical university of warsaw. in the period under consideration (december to october ) there were patients receiving allogeneic stem cell transplants. the criteria for patient selection for the study included the appearance of at least one syndrome from those listed below within a period of days after hsct: pneumonia, appearance or intensification of graft-versus-host disease (gvhd), skin rash, or pyrexia of unknown origin. introduction of these criteria resulted in a group to adult receivers of hscts ( table ) . monitoring of clinical status of the patients and viral load in blood samples comprised a period of days after hsct. igg and igm antibodies against hhv- were measured from the panel of serum specimens, collected once before hsct, from the patients with different hematological disorders using a commercial enzyme immunoassay (panbio). the results obtained with both hhv- -specific igg and igm tests were expressed in panbio units (pbu). the pbu is calculated as the ratio of the sample absorbance to the cut-off absorbance and multiplied by (absorbance of sample/mean absorbance of cut-off × ). the collection of plasma samples from all patients for hhv- pcr investigations, according ebmt guidelines, began at a median of days after transplantation (range: - days) and lasted until a median of days (range: - days). routine collection of plasma samples was performed once a week until the th day after allogeneic hsct, thereafter once every two weeks, up to th day after hsct. the median number of blood samples per patient was (range: - ). a total of samples from the patients were collected. for the purpose of recent study, viral dna was extracted from µl of each plasma sample using a qiamp ® dna blood mini kit (qiagen) in accordance with the manufacturer's instructions and retrospectively used for detection of hhv- dna. for the detection of hhv- , a real-time pcr assay with fluorescent probes complementary to the sequence lying within the amplified product was used. the tests were run on a lightcycler . instrument (roche) with the commercial quantitative mutareal ® hhv- kit (alpco). all samples were examined according to the manufacturer's instructions. every tested sample was amplified with an internal control (positive control of the dna extraction and amplification process). each amplification reaction also included, besides the tested samples, positive hhv- -specific controls and a negative control of the dna extraction and amplification process. hhv- dna was detected using the commercial real-time test cmv quant kit ® (roche) developed for the lightcycler . instrument. the test uses internal scorpions™ fluorescent probes for the pcr amplification product. analogously to hhv- detection, for every sample an internal control was added and the amplification was performed in the presence of amplification-specific controls (positive, negative, and extraction process control). specific igm antibodies were present in the serum of ( %) patients, while the others were negative ( %). twenty of the persons ( %) had igg antibodies against hhv- before hsct and the other ( %) were negative. of the group of patients who had no detectable hhv- during the entire post-transplantation period, ( . % of the hhv- -negative patients) had a positive result only for the anti-hhv- igg test, but no anti-hhv- igm, in serum sample taken directly before transplantation, ( . %) were both igm and igg negative, and the remaining patient had anti-hhv- antibodies of both classes ( table ) . the serological status of the patients who had hhv- dna detected in one or more blood samples was very similar: patients ( %) had only igg, patients ( %) had no igg or igm antibodies, and one patient had hhv- -specific igg and igm. regarding the amplification results of dna isolated from plasma using hhv- -specific pcr, expressed as the presence of exponential increases in fluorescence, products were detected in samples ( %) taken from patients ( %). two of them ( %) had hhv- dna in only a single positive sample and another ( %) had positive results in two or more subsequent tests. in the patients with two or more plasma samples containing hhv- dna, viremia was observed between days and after transplantation (table ). in the two patients who had a single hhv- -positive blood sample, viral dna was detected at a later time. the quantitative results obtained by the mutareal ® hhv- test in all positive cases were at low levels, between - copies/ml. hhv- dna was detected in the plasma samples collected from four hhv- -negative patients. in all four cases, hhv- dnaemia was observed in the typical period of - days after transplantation and, moreover, a second hhv- viremia course occurred in two patients starting at days and after transplanta- abbreviations: all -acute lymphoblastic leukemia, aml -acute myeloid leukemia, cml -chronic myeloid leukemia, mdsmyelodysplastic syndrome, rpbsct -related peripheral blood stem cell transplantation, upbsct -unrelated peripheral blood stem cell transplantation, ucbsct -unrelated cord blood stem cell transplantation, rbmt -related bone marrow transplantation, ubmt -unrelated bone marrow transplantation, -: negative, +: positive in one plasma/serum sample, ++: positive in two or more plasma samples. abbreviations: ards -acute respiratory distress syndrome, cns -central nervous system, gvhd -graft-versus-host disease, n/a -not applicable, -: negative, +: positive in one plasma/serum sample, ++: positive in two or more plasma samples. tion. none of the hhv- -positive patients had hhv- dna in their plasma samples during the -day period. overall mortality in the entire group of hsct recipients during the first days after transplantation was . % ( of the patients) and the most frequent direct cause of death was acute respiratory distress syndrome ( patients), in some patients accompanied by sepsis ( patients), pneumonia ( patient), or central nervous system (cns) infection ( patients). other death causes included pneumonia ( patient), bleeding within the cns during infectious meningitis ( patient), and shock during subsequent hsct ( patient). seven of the patients died during the first days after hsct (table ). in the hhv- -positive patients, mortality was % ( out of patients). four of those patients died during the hhv- viremia period, which occurred within the first days after hsct. table summarizes the clinical features and the times and causes of death of the hsct recipients. herpesviruses persist throughout life after primary infection. viral proliferation occurs either spontaneously or under conditions of immunosuppression. reactivation can lead to illnesses that typically differ in their clinical presentation from the disease associated with the primary infection. after sct, reactivating members of the β-herpesvirinae subfamily (among them hhv- ) frequently cause serious, sometimes life--threatening disease [ , ] . hhv- infection or reactivation in these individuals has been associated with a delay or suppression of marrow engraftment [ ] , pneumonia [ ] , skin rash, and fever [ ] . although it is difficult to prove an etiologic association of the virus with these disease events, their propinquity with hhv- activity in the absence of other possible causes suggests that at the very least a subset of these events is due to hhv- activity. in the present study we found that % ( individuals) of the graft recipients developed hhv- infection ( table ) . eight patients ( %) had detectable viral dna levels in two or more samples in subsequent tests during the six weeks following sct and another two ( %) had a single sample positive at a later time. it is likely that the hhv- infection that occurred after transplantation was due to activation of the virus in the bodies of the recipients, since most of the recipients were immune to hhv- and no virus was found before sct. however, we do not know whether hhv- strains detected in the present study were derived from the donor or were transfered from other sources to the seronegative recipients. the virus probably remains latent in the body after primary infection, as do other human herpesviruses. if the hhv- was derived from the donor, it must have been latently infecting the donor's hemopoietic stem cells and was activated to replicate after transfer to the recipient. in other cases, the virus was probably reactivated from the recipient's own body by factors such as a profound immune dysfunction or an allogeneic reaction after transplantation. we did not find any correlation between viremia and the applied conditioning regimen or anti-gvhd prophylaxis. all of the hhv- -positive graft recipients had fever of unknown etiology during the six weeks after sct and half of them ( persons) had acute gvhd features. sixty-three percent of the described patients had pneumonia and % skin rash. no epstein-barr virus or cmv dna was found in all the plasma samples during hhv- onset ( table ) . the exact association between hhv- reactivation and mortality found in this study is not clear. three of the patients ( %) with detectable hhv- dna levels died during viremia shortly after transplantation, all due to coexisting pneumonia of unconfirmed etiology. acyclovir in typical doses was used as an antiviral prophylaxis during this period, but without any visible clinical success. studies in vitro have shown that hhv- dna replication is inhibited by ganciclovir, foscarnet, and cidofovir, but not by acyclovir [ ] . there is a high frequency of detectable hhv- viral load in sct recipients and it may lead to an increased risk of fatal symptomatic disease [ ] . the availability of quantitative real-time pcr means that results are available in a clinically helpful time-frame, which should assist with implementing timely therapeutic intervention and assessing response to treatment. further investigation to monitor hhv- reactivation on a larger group of sct recipients will be important in improving outcome for these patients. susceptibility of human herpesvirus to antivirals in vitro interstitial pneumonitis associated with human herpes-virus- infection after marrow transplantation human herpesvirus human herpesvirus- and human herpesvirus- infections in bone marrow transplant recipients virus taxonomy -classification and nomenclature of viruses. eighth report of ictv hepatitis due to human herpesvirus- human herpesvirus in salivary glands betaherpesviruses in transplant recipients meningoencephalitis associated with hhv- related exanthem subitum latent human herpesvirus infection of human monocytes/macrophages association of human herpesvirus infection of the central nervous system with recurrence of febrile convulsions high levels of human herpesvirus dna in peripheral blood leucocytes are correlated to platelet engraftment and disease in allogeneic stem cell transplant patients human herpesvirus latently infects early bone marrow progenitors in vivo inverse relationship between human herpesvirus- and - etection after allogeneic and autologous stem cell transplantation isolation of a new virus, hblv, in patients with lymphoproliferative disorders human herpesvirus- and - in pediatric stem cell transplantation antibody reactivity with hblv encephalitis caused by human herpesvirus- in transplant recipients: relevance of a novel neurotropic virus the spectrum of human herpesvirus infection: from roseola infantum to adult disease identification of human herpesvirus as causal agent for exanthem subitum human herpesvirus viremia in bone marrow transplant recipients: clinical features and risk factors clinical outcomes of human herpesvirus reactivation after hematopoietic stem cell transplantation key: cord- - bt x authors: crocchiolo, r.; bramanti, s.; vai, a.; sarina, b.; mineri, r.; casari, e.; tordato, f.; mauro, e.; timofeeva, i.; lugli, e.; mavilio, d.; carlo‐stella, c.; santoro, a.; castagna, l. title: infections after t‐replete haploidentical transplantation and high‐dose cyclophosphamide as graft‐versus‐host disease prophylaxis date: - - journal: transpl infect dis doi: . /tid. sha: doc_id: cord_uid: bt x background: recently, a platform of t‐cell replete haploidentical hematopoietic stem cell transplantation (haplo‐hsct) using post‐transplant cyclophosphamide (cy) has shown high reproducibility and acceptable safety profile. method: this prospective cohort analysis allowed us to collect data on infections among consecutive recipients of haplo‐hsct affected by various hematologic malignancies. results: after a median follow‐up of months, cumulative incidence of viral infections was % ( % confidence interval [ci] – ) at days and % ( % ci – ) at year; of patients at risk had cmv reactivation ( %) and the rate of polyomavirus‐virus‐associated cystitis was % ( / ). cumulative incidence of bacterial and fungal infections at year were % ( % ci – ) and % ( % ci – ), respectively. of note, only invasive fungal infection occurred beyond year after transplant (day + ). conclusion: in conclusion, despite a high rate of viral infections in the early period, present data suggest a satisfactory infectious profile after t‐cell replete haplo‐hsct using post‐transplant cy. these results may help clinicians to improve both prophylactic and therapeutic antimicrobial strategies in this emerging haploidentical setting. one of the major limitations of hematopoietic stem cell transplantation from haploidentical donor (haplo-hsct) is the impaired immune reconstitution owing to extensive immunosuppression necessary to overcome human leukocyte antigen disparity. despite important advances over the last decades, infections are still mostly responsible for toxicity and non-relapse mortality among transplanted patients, owing to prolonged immunosuppression related, or not, to chronic graft-versus-host disease (gvhd) ( , ) . a platform for t-cell replete haplo-hsct using posttransplant cyclophosphamide (cy) ( ) showed a low treatment-related mortality (trm) and a high feasibility with an acceptable safety profile. this type of haplo-hsct seems to compare favorably with t-cell depleted methods, in terms of infectious complications ( , ) . to further explore this topic, we report herein the incidence of infections in a single-center cohort of consecutive patients receiving t-cell replete haplo-hsct with post-transplant cy at our center, in order to provide useful information about the post-transplant period after this type of emerging transplant platform. data on patients with hematologic malignancies who underwent haplo-hsct between april and april at humanitas cancer center (milan, italy) were prospectively collected using electronic patients' charts. conditioning regimens were myeloablative, reducedintensity, or non-myeloablative. non-myeloablative conditioning consisted of cy . mg/kg/day intravenously (i.v.) on days À and À , fludarabine mg/ m /day i.v. on days À to À , and total body irradiation (tbi) cgy on day À . two reduced-intensity regimens were administered: (i) thiotepa mg/kg/ day i.v. on day À , fludarabine mg/m /day i.v. on days À to À , cy mg/kg/day i.v. on days À and À , and tbi cgy on day À ; or (ii) thiotepa mg/ kg/day i.v. on days À and À , fludarabine mg/m / day i.v. on days À to À , and busulfan . mg/kg/day i.v. on days À and À , as modified from sanz et al. ( ) . the myeloablative regimen consisted of thiotepa mg/ kg/day i.v. on days À and À , fludarabine mg/m / day i.v. on days À to À , and busulfan . mg/kg/day i.v. on days À to À . all patients received unmanipulated bone marrow or mobilized peripheral stem cells. gvhd prophylaxis was performed with cy mg/kg/day on days + and + ; tacrolimus mg/day i.v. from day + (to reach a concentration of - ng/ml), or cyclosporine mg/ kg/day i.v. from day + (to reach a concentration of - ng/ml), both up to day + and then tapered up to day + , unless gvhd occurred; and mycophenolate mofetil mg/kg a day orally from day + to + . infections were defined according to european society for blood and marrow transplantation (available at: http://www.ebmt.org/contents/about-ebmt/who-we-are/scientificcouncil/documents/idwpdefiniti ons.pdf), including microbiologically documented viral, bacterial, or fungal infections with or without laboratory and/or radiologic features consistent with organ involvement. cytomegalovirus (cmv) reactivation (or de novo infection) and disease were diagnosed as reported elsewhere ( ) , and invasive fungal infections (ifis) were classified according to the definitions of the eortc/msg consensus group ( ); only proven or probable ifis were recorded. data were recorded as of june , for all patients. the study was approved by the local institutional review board. antimicrobial prophylaxis was started during the conditioning regimen and consisted of acyclovir mg/ m² times in a day; levofloxacin mg/day; and cotrimoxazole tablets per day until day À , and then tablet every other day was resumed after hematologic reconstitution. antifungal prophylaxis was performed with an echinocandin (either caspofungin or micafungin mg/day) until day + , when itraconazole ( mg/day i.v.) was administered, unless contraindicated; otherwise the echinocandin was maintained. after september , the echinocandin was maintained for all patients. acyclovir, levofloxacin, and antifungal prophylaxis were administered until engraftment occurred. twice weekly blood polymerase chain reaction (pcr) cmv monitoring was started at day + until day + and weekly until day + , or when clinically indicated. weekly epstein-barr virus (ebv) monitoring by pcr was started at day + up to day + , or when clinically indicated. all other tests were performed whenever indicated. piperacillin-tazobactam alone or in combination with an aminoglycoside was administered as empirical therapy for febrile neutropenia, unless previous colonization for resistant bacteria was documented; in this case, an appropriate antibacterial agent was delivered. the same was true when a suspected bacterial infection occurred, with the exception of pneumonia for which linezolid was added, in combination with the abovecited antibacterial drug(s). first-line preemptive therapy for cmv infection/reactivation was with intravenous ganciclovir, whereas foscarnet was administered if the patient was in aplasia. ebv reactivation and polyomavirus-related hemorrhagic cystitis were treated by rituximab and cidofovir, respectively. threshold of cmv viremia for the initiation of therapy was copies/ml; threshold of ebv viremia was , copies/ml. analysis of circulating lymphocytes was performed at regular intervals whenever available, at days + , + , + , and + , and every month afterward. the following transplant infectious disease : : - monoclonal antibodies and combinations were used: cd , cd /cd , cd /cd , cd , cd /cd (beckman coulter, fullerton, california, usa), to quantify t, b, and nk cell compartments at the different time points studied. categorical variables were expressed as absolute numbers with respective percentage and continuous variables as the median with the respective range. cumulative incidence of viral, bacterial, or fungal infections was calculated using competing risk analysis ( ) starting from the day to the day of the first infection; death was considered as the competing event. infection incidence was also expressed as the events/ patient-days (pt-days) for each time period within year after transplant, with intervals defined as follows: days - , - , - , and - . owing to the paucity of late events, data beyond day + were collected singularly and classified according to pathogen group. neutrophil engraftment was defined as the first of consecutive days with a persistent count > . /l; platelet engraftment was defined as the first of consecutive days with a persistent count > /l (https://portal. ebmt.org/sites/clint /clint/documents/statguidelines_ oct .pdf). the kaplan-meier method was used to compute overall survival ( ); cumulative incidence of trm and acute and chronic gvhd were calculated using competing risk analysis ( ) . death because of documented infection was defined as infection-related death. logrank test was used to compare the incidence of infections with flow cytometry results. seventy consecutive adult patients undergoing haplo-hsct were identified. the main patient and transplant characteristics are shown in table . fifty-five patients ( %) were affected by lymphoma, and % of patients underwent haplo-hsct in partial or complete remission. only patient/donor pairs were cmv dÀ/rÀ. as concerns antifungal prophylaxis, patients did not receive itraconazole owing to moderate increase in pre-transplant liver function tests (n = ) or reduction in left ventricular ejection fraction (n = ); more patients did not receive itraconazole because haplo-hsct was performed after september . three patients received secondary antifungal prophylaxis with voriconazole because of previous pulmonary aspergillosis. median follow-up of living patients was months (range - ) from day of stem cell infusion. at last follow-up, a total of documented infectious events occurred among of patients, with a median of events/patient (range - ); % were of viral origin (n = ), % bacterial (n = ), % fungal (n = ). cumulative incidence of first viral infection was % ( % confidence interval [ci] - ) and % ( % ci - ) at day + and + , respectively; at day + , the incidence of bacterial infections was % ( % ci - ), and that of ifis was % ( % ci - ) (fig. ) . in % ( of patients at risk) at least cmv reactivation developed; of these, patients had cmv reactivation, and , , , and patients had a total of , , , and cmv reactivations, respectively. two non-fatal ( colitis, pneumonia) and fatal (pneumonia) cmv diseases occurred. no primary cmv infections occurred in the cmv dÀ/rÀ patient/donor pairs. polyomavirus-related hemorrhagic cystitis was observed in patients ( %): were caused by bk virus and by jc virus. importantly, no ebv-related lymphoproliferative disorders occurred so far. forty-five patients ( %) presented with at least documented bacterial infection: ( %), ( %), and ( %) patients had an infection by grampositive, gram-negative, or both types of bacteria, respectively. eleven ifis were detected in patients: n = probable invasive aspergillosis (pneumonia in patients and sinusitis in ), n = invasive candidiasis, all by non-albicans candida ( candidemias, colitis, and hepatosplenic candidiasis); median of occurrence was days from haplo-hsct (range - ). among the patients receiving secondary antifungal prophylaxis, only non-albicans candida colitis was observed at day + in patient. no ifis occurred under active gvhd. notably, only ifis occurred beyond day + : pulmonary aspergillosis at day + , and candidemia at day + , this latter in a patient who was under salvage treatment for post-transplant relapse. details of etiologies are reported in table . we did not observe significant differences of infectious events according to conditioning regimen administered (data not shown). when considering the timing of all episodes, bacterial infections occurred mostly between day and + , whereas viral infections/ reactivations between days + and + , with . bacterial events/ pt-days between day and + , and . viral events/ pt-days between days + and + (fig. ) . the overall incidence of viral events between day and day + was . events/ ptdays. a total of bacterial and viral infections were observed after year from transplant. engraftment rate was % ( of patients), with a median of days (range - ) and days (range - ) for neutrophil and platelet recovery, respectively. four patients died before engraftment (on days + , + , + , and + because of gram-negative sepsis, multiorgan failure, progressive disease, and bacterial pneumonia, respectively) and presented primary antibody-linked graft failure, and are alive at last follow-up, after autologous reconstitution (days + and + ). cumulative incidence of acute grade - and - gvhd was % ( % ci - ) and % ( % ci - ) respectively; chronic gvhd was % ( % ci - ). two-year overall survival was % ( % ci - ) and trm was % ( % ci - ); patients ( %) relapsed or progressed after haplo-hsct. infection-related deaths were % ( / , occurring between days + and + ; bacterial pneumonia = , gram-negative sepsis = , cmv pneumonia = , h n pneumonia = , and jc virus-related progressive multifocal leukoencephalopathy = ). the other, non-infectious, causes of trm were heart failure (n = ), secondary malignancy (n = myelodysplastic syndrome, n = esophageal cancer), multiorgan failure (n = ), thrombotic microangiopathy (n = ), and acute hepatitis (n = ). at last follow-up, patients are alive and of them are in complete remission. immunophenotypic analysis reveals a progressive increase in all lymphocyte subset counts from day + through later post-transplantation time points. we found a trend toward less viral infection incidence among those patients who have a total lymphocyte count > /mm at day + : hazard ratio . , p = . . no other associations were observed. lymphocyte subsets and number of patients analyzed are shown in figure . in the present analysis, we described infectious complications after unmanipulated, t-cell replete haplo-hsct using post-transplant cy in consecutive patients and found, aside from a high incidence of viral infections/reactivations, especially in the early posttransplant period, a quite low incidence of late bacterial infections, together with a very low incidence of ifis after day + ( events in the overall observed). present findings confirm that the infectious profile is better in t-cell replete vs. t-cell depleted haplo-transplantation ( ); the lower incidence of infections observed after day + may reflect a partial and quite effective restoration of antimicrobial immunity during the post-transplant period in this type of haplo-hsct. importantly, the low rate of chronic gvhd seen in our cohort is likely to contribute to this phenomenon, as chronic gvhd is known to be a major risk factor of late morbidity and mortality ( ) . nevertheless, we found an unexpected % trm incidence, higher than that originally reported with post-hsct cy ( ); this may be a result of the inclusion of patients with more advanced disease in the haplo-hsct program at our center. as concerns viral infections, our results are in line with previous publications in the setting of t-cell replete haploidentical transplants. ciurea et al. ( ) reported . events/ pt-days within the first months from transplant (vs. . events/ pt-days between day + and + in our hands), and raiola et al. ( ) found that % of patients presented with a viral infection in the first year. the % of cmv reactivations found here was comparable to the - % reported in similar haploidentical settings ( , , ); the slightly higher incidence that we found here may be explained by the longer follow-up in our series. the polyomavirus-associated cystitis rate of %, which is lower than that reported in the myeloablative setting ( ) , is likely because of the different conditioning regimens in our cohort, although a role played by the different gvhd prophylaxis cannot be excluded; indeed, bk virus nephropathy was found to be more frequently associated with tacrolimus than with cyclosporine in recipients of kidney allografts ( ) . here, a quarter of the patients ( / ) received cyclosporine as gvhd prophylaxis. interestingly, we confirm the lack of ebv-related lymphoproliferative disorders, as recently reported also by kanakry et al. ( ) . we found a % incidence of fungal infections, none of them fatal; it is important to note that fatal episodes of fungal infections are among the major limiting toxicities associated with t-cell depletion in haplo-hsct ( , ) ; we cannot exclude a role played by the use of anti-mold prophylaxis during marrow aplasia, although it is difficult to draw definitive conclusions owing to the lack of a true control arm in our study, and to the low number of ifis. of note, we observed only ifis months after transplant. concerning bacterial infections, we may explain the low prevalence of late bacterial infections (i.e., beyond year) by the surprising % incidence of chronic gvhd; in fact, the risk of bacterial events remained low in the absence of late immunosuppressive therapy ( ) . with a median follow-up of months, we observed late bacterial events in a total of patients having at least year of observation (last observation day is months after haplo-hsct). all this information argues in favor of the fact that giving a t-cell replete graft without deep in vivo t-depletion (i.e., with anti-thymocyte globulin or alemtuzumab) and with post-transplant cy allows a satisfactory infectious profile after transplant. the posttransplant high-dose cy permits naive and non-activated memory cells to reconstitute the immune system later on ( , ) , enabling patients to be protected from late infectious events. the same mechanism probably also explains the high viral reactivation incidence found in the first months, owing to the low number of adoptively transferred memory t cells in the early phase after transplantation (lugli e. et al., unpublished data). we acknowledge that potential selection bias may be present in the study, as we cannot exclude the possibility that some non-severe or very late infections were not captured because of incomplete reporting. however, all patients were followed at the same institution; therefore, it is unlike that clinically relevant infectious complications were missed; moreover, diagnostic procedures and prophylactic measures were similar for all patients, thus contributing to the accuracy of diagnosis of the infectious events. in conclusion, the present single-center data on consecutive patients receiving t-cell replete haplo-hsct with post-transplant cy confirm a high rate of viral infections before day + and a lower incidence of infections afterward, suggesting a satisfactory although non-optimal immune reconstitution after this type of transplantation. future comparisons with other haploidentical platforms and/or other alternative stem cell sources (i.e., cord blood), as well as investigations of novel strategies of transfer of immunity are warranted. furthermore, the present data may provide useful information in an attempt to improve control of infections by adequate prophylaxis and/or antimicrobial therapy in the early post-transplant period, after use of the emerging transplant platform of haplo-hsct. reduced mortality after allogeneic hematopoietic cell transplantation long-term survival and late deaths after allogeneic hematopoietic cell transplantation hla-haploidentical bone marrow transplantation for hematologic malignancies using nonmyeloablative conditioning and high-dose, posttransplantation cyclophosphamide outcomes of related donor hla-identical or hla-haploidentical allogeneic blood or marrow transplantation for peripheral t cell lymphoma cord blood transplantation from unrelated donors in adult with high-risk acute myeloid leukemia definitions of cytomegalovirus infection and disease in transplant recipients revised definitions of invasive fungal disease from the european organization for research and treatment of cancer/invasive fungal infections cooperative group and the national institute of allergy and infectious diseases mycoses study group (eortc/msg) consensus group estimation of failure probabilities in the presence of competing risks: new representations of old estimators nonparametric estimation from incomplete observations infectious complications in cord blood and t-cell depleted haploidentical stem cell transplantation risk factors for late infections after allogeneic hematopoietic stem cell transplantation from a matched related donor improved early outcomes using a t cell replete graft compared with t cell depleted haploidentical hematopoietic stem cell transplantation unmanipulated haploidentical bone marrow transplantation and posttransplantation cyclophosphamide for hematologic malignancies after myeloablative conditioning haploidentical transplantation using t cell replete peripheral blood stem cells and myeloablative conditioning in patients with high-risk hematologic malignancies who lack conventional donors is well tolerated and produces excellent relapse-free survival: results of a prospective phase ii trial ast infectious diseases community of practice. bk polyomavirus in solid organ transplantation absence of post-transplantation lymphoproliferative disorder after allogeneic blood or marrow transplantation using posttransplantation cyclophosphamide as graft-versus-host disease prophylaxis full haplotype-mismatched hematopoietic stem-cell transplantation: a phase ii study in patients with acute leukemia at high risk of relapse infusion of suicidegene-engineered donor lymphocytes after family haploidentical haemopoietic stem-cell transplantation for leukaemia (the tk trial): a non-randomised phase i-ii study high-dose cyclophosphamide for graft-versus-host disease prevention thymic t-cell development in allogeneic stem cell transplantation thanks: we thank all personnel working in the hematology and transplantation unit at humanitas cancer center for their remarkable contribution in patients' care and assistance to their families.author contributions: r.c. designed the study, performed data analysis, and wrote the manuscript; s.b., b.s., f.t., and e.m. provided clinical care; a.v. collected data and performed statistical analysis; r.m., e.c., and i.t. provided laboratory and microbiological data; e.l. and d.m. critically revised the manuscript; c.c-s., a.s., and l.c. provided clinical care and critically revised the manuscript.conflict of interest: all authors declare no financial conflict of interest. key: cord- - fznfp p authors: holtan, s g; hogan, w j; elliott, m a; ansell, s m; inwards, d j; porrata, l f; johnston, p b; micallef, i n; lacy, m q; gastineau, d a; litzow, m r title: cd (+) cell dose and establishment of full donor chimerism at day + are important factors for survival with reduced-intensity conditioning with fludarabine and melphalan before allogeneic hematopoietic sct for hematologic malignancies date: - - journal: bone marrow transplant doi: . /bmt. . sha: doc_id: cord_uid: fznfp p the combination of fludarabine and melphalan as a reduced-intensity conditioning (ric) regimen extends allogeneic hematopoietic sct (hsct) as a therapeutic option for elderly or frail patients with relapsed, refractory or other high-risk hematologic malignancies. whether any modifiable factors exist that could improve survival before or immediately after hsct is unknown. we reviewed the medical records of the first patients at our institution to undergo fludarabine/melphalan ric from september to september to determine factors associated with survival. a total of ( %) patients had undergone prior hsct and as such was a high-risk group of patients. on multivariate analysis, cd (+) cell dose greater than . × ( ) per kg (risk ratio (rr) . , % ci . – . , p= . ) and full donor chimerism at day + (rr . , % ci . – . , p= . ) remained independent prognostic factors. in our series, achievement of full donor chimerism at day + was associated with an approximately % -year survival, a favorable outcome in this high-risk group of patients. although the infused cd (+) cell dose is a modifiable variable, whether donor lymphocyte infusions or other immunologic interventions should be performed to promote the establishment of full chimerism early post transplant remains unknown. allogeneic hematopoietic sct (hsct) is a life-saving treatment for patients with high-risk or refractory hematologic malignancies. however, the potential severe toxicities of myeloablative conditioning preclude many patients from candidacy for this therapy. reduced-intensity conditioning (ric) regimens extend this modality to patients who would otherwise be deemed medically unsuitable to receive myeloablative conditioning. the combination of fludarabine and melphalan is a commonly used moderately myelosuppressive ric regimen that has antineoplastic activity in both myeloid malignancies including aml and myelodysplastic syndrome (mds) and lymphoid malignancies including all, hodgkin's and non-hodgkin's lymphomas , and multiple myeloma. its use results in the rapid establishment of donor chimerism, excellent rates of engraftment and a reduction in relapse risk compared with other ric regimens. [ ] [ ] [ ] fludarabine/melphalan ric also has proven tolerable in patients who have failed a previous autologous or allogeneic hsct, although a high rate of relapse as well as late complications of hsct may adversely affect outcome in this high-risk group of patients. because those who undergo ric allogeneic hsct have either a guarded prognosis from a disease or comorbidity standpoint or both, we sought to determine modifiable factors associated with survival to identify potential therapeutic targets to improve outcome in this group of patients. we reviewed the medical records of the first consecutive patients who underwent fludarabine/melphalan ric at our institution from september to september to determine factors associated with survival. the mayo clinic institutional review board approved the study. overall survival was defined as the time from day to date of death due to any cause. patients transplanted during - were given fludarabine mg/m on days À to À and melphalan mg/m on day À . those transplanted during and later received split dosing of the melphalan at mg/m on days À and À . factors analyzed for association with survival included demographic and disease factors (age at transplant, sex, lymphoid vs myeloid disease, karyotype and blast percentage in those with mds and leukemia, disease status at transplant, previous hsct, and previous chemotherapy for solid tumors), donor/graft factors (cd þ cell dose, graft source, related vs unrelated graft, degree of hla match, cmv serostatus and abo match), and post transplant factors (time to neutrophil, lymphocyte and platelet engraftment, day þ chimerism status, development of and severity of acute gvhd, development of and severity of chronic gvhd). full donor chimerism was defined as x % as previously described , and was monitored by a peripheral blood (or less commonly bm) method involving genomic dna extraction from unsorted mononuclear cells and subsequent pcr amplification of highly polymorphic short tandem repeats. occasionally, a fish method of labeling x and y chromosomes in opposite sex transplants was used. univariate survival analyses for nominal variables were performed according to the kaplan-meier method. the two-tailed log-rank test was applied to determine statistically significant differences. cox proportional hazards model was applied both for the univariate survival analysis with continuous variables and for the multivariate survival analysis. the following continuous data were also analyzed as binary variables after dichotomization: cd þ cell dose (above vs equal to or below the median of .  cells per kg), bm aspirate blast percentage (p % or %) and time to absolute lymphocyte count greater than  cells per liter ( days or less or greater than days) based on our institution's previous experience with evaluating post transplant immune reconstitution. univariate variables with pp . were included in the multivariate model, with the least significant one being eliminated in serial iterations until only statistically significant variables with a po . remained (backward selection). table lists baseline demographic data and univariate analysis. the median age at transplant was . years (range - years). most patients received hsct for myeloid malignancies (predominantly aml or mds, patients, %). approximately one-third patients were in cr at the time of transplant ( %). the majority also received stem cells from a sibling donor ( %). half ( patients) of the patients in this cohort had received previous hsct. seven patients had a prior allogeneic hsct for aml, and one had a prior allogeneic hsct for cll. one patient each had undergone prior autologous hsct for aml and breast cancer. the remaining patients who had prior transplants had undergone autologous hsct for lymphoid malignancies. the indication for ric allogeneic hsct for most ( patients) was a relapse of their primary malignancy. six patients were undergoing transplantation for a new primary malignancy (therapy-related mds with six patients received tacrolimus and mtx as gvhd prophylaxis, and the remaining one patient received no gvhd prophylaxis (second allogeneic transplant for relapsed leukemia o days before the first transplant from the same sibling donor). median time to neutrophil engraftment was days ( - days interquartile range), platelet engraftment was days ( - days interquartile range) and lymphocyte engraftment  per liter was days ( - . interquartile range). % of patients experienced grade - acute gvhd, and % experienced chronic gvhd ( % extensive). of , patients have died, from relapsed disease ( table ). only one of the deaths before day þ was due to relapsed disease, and mortality (primarily due to infections) at day þ was % ( patients). three deaths between day þ and year were due to relapse, and six were due to complications of the transplant. of those patients surviving more than year, one death was due to relapse, two deaths were due to gvhd, and one death was unexpected and of unknown cause. twenty-three patients ( %) are alive after a median of months of follow-up. we performed a multivariate analysis to determine factors independently associated with survival using those factors significant on univariate analysis (karyotype in myeloid diseases, presence of chronic gvhd, cd þ cell dose, day þ chimerism and time to neutrophil engraftment). cd þ cell dose greater than .  per kg (risk ratio (rr) . , % ci . - . , p ¼ . ) and full donor chimerism at day þ (rr . , % ci . - . , p ¼ . ) remained independent prognostic factors. figure shows kaplan-meier estimates for survival associated with cell dose and chimerism. achievement of full donor chimerism was not related to cd þ cell dose (p ¼ . ). our retrospective study has identified two potentially modifiable factors associated with improved survival in patients receiving fludarabine/melphalan ric before allogeneic hsct: cell dose and chimerism. cell dose has previously been associated with a reduced relapse risk, potentially because of improved post transplant lymphocyte recovery. cd þ cell dose is also associated with the development of chronic gvhd in the reduced-intensity setting, a factor strongly associated with survival with a similar ric regimen consisting of fludarabine and bu followed by hsct for high-risk aml and mds. an optimal cell dose for ric has been proposed at to  per kg cd þ . cell dose did not influence rates of full donor chimerism, the other independent prognostic factor we identified in this series. although mixed chimerism may not be detrimental to survival in allogeneic transplants for nonmalignant diseases, whether any degree of mixed chimerism is acceptable in allogeneic transplantations for hematologic malignancies is subject to debate. in our series, lineagespecific (t-cell vs myeloid) chimerism was not available on many patients and thus not formally evaluated for association with survival. in another series of fludarabinebased ric, delayed t-cell chimerism was clearly associated with poorer pfs in those with myeloid malignancies ( % relapse rate compared to % in those with mixed vs complete t-cell chimerism, respectively, p ¼ . ). whether lineage-specific chimerism should be routinely monitored or whether any immunologic therapies such as prophylactic donor lymphocyte infusions or cytokine therapies such as il- should be undertaken in an attempt to convert mixed chimerism to full donor chimerism in the early post transplant period is worthy of prospective study. our study has important limitations due to our limited number of patients from a single institution and retrospective analysis. chimerism from earlier time points (days þ and þ ) was performed on an insufficient number of patients to be included in analysis. in addition, lineagespecific chimerism and lymphocyte subset analysis was not performed on most patients. two other important and potentially modifiable factors that we did not have the statistical power to analyze in this cohort are the impact of donor source and various gvhd prophylaxis regimens on lymphocyte recovery and survival. these shortcomings could be addressed with a prospective study. finally, fludarabine and melphalan ric may not be suitable conditioning before unrelated cord blood transplants without additional immunomodulation owing to high rates of graft failure ( of reported patients), and consequently our study results may not be generalizable to those undergoing cord blood transplants. gvhd remains a significant cause of morbidity and mortality in ric allogeneic hsct. the addition of alemtuzumab to fludarabine and melphalan ric before allogeneic hsct for mds and aml has recently been associated with a sixfold reduction in rates of gvhd, although this was associated with a nonsignificant increase in disease recurrence rates compared with a historical cohort. gvhd was the cause of death in four patients ( %) in our series. however, of the patients who died of infections (two with bacterial sepsis and one with disseminated herpes simplex virus) were receiving cya and tapering doses of steroids for acute gvhd, bringing the total deaths attributable to gvhd or complications of its treatment to patients ( %). infections/multiorgan failure without gvhd and disease relapse were more frequently the cause of death (nine and five patients, respectively) in our series, arguing for the greater need for improved immunologic recovery rather than augmenting immunosuppression in our group. reduced-intensity conditioning regimens such as the combination of fludarabine and melphalan have extended allogeneic transplantation as a potentially life-saving modality for patients with high-risk or refractory hematologic malignancies who would otherwise be medically unsuitable for myeloablative conditioning. because the toxicities of conditioning were relatively tolerable even in our heavily pretreated high-risk population, developing methods of rapid and effective immune reconstitution to reduce the risk of infectious complications and disease relapse becomes even more important. in this study, we identified day þ full donor chimerism and cd þ cell dose greater than .  per kg as potentially modifiable prognostic factors in ric allogeneic transplantation. both frequent monitoring of chimerism status and early intervention with cytokines, donor lymphocyte infusions or other immunomodulatory treatments when o % donor chimerism is identified as well as optimizing cd þ cell dose may improve patient outcomes and could be studied prospectively. allogeneic hematopoietic stem cell transplantation for the treatment of high-risk acute myelogenous leukemia and myelodysplastic syndrome using reduced-intensity conditioning with fludarabine and melphalan reduced-intensity conditioning allogeneic stem cell transplantation is a potential therapeutic approach for adults with highrisk acute lymphoblastic leukemia in remission: results of a prospective phase study fludarabine-melphalan as a preparative regimen for reduced-intensity conditioning allogeneic stem cell transplantation in relapsed and refractory hodgkin's lymphoma: the updated m.d. anderson cancer center experience favorable long-term survival after reducedintensity allogeneic transplantation for multiple-relapse aggressive non-hodgkin's lymphoma fludarabine/melphalan conditioning for allogeneic transplantation in patients with multiple myeloma stable engraftment after a conditioning regimen with fludarabine and melphalan for bone marrow transplantation from an unrelated donor fludarabine phosphate and melphalan: a reduced intensity conditioning regimen suitable for allogeneic transplantation that maintains the graft versus malignancy effect comparison between two fludarabinebased reduced-intensity conditioning regimens before allogeneic hematopoietic stem-cell transplantation: fludarabine/ melphalan is associated with higher incidence of acute graftversus-host disease and 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on the outcome of patients undergoing reduced-intensity-conditioning allogeneic peripheral blood stem cell transplantation sustained remissions of high-risk acute myeloid leukemia and myelodysplastic syndrome after reduced-intensity conditioning allogeneic hematopoietic transplantation: chronic graft-versus-host disease is the strongest factor improving survival optimizing the cd + cell dose for reducedintensity allogeneic hematopoietic stem cell transplantation allogeneic hematopoietic sct in patients with non-malignant diseases, and importance of chimerism reduced intensity conditioning and prophylactic dli can cure patients with high-risk acute leukaemias if complete donor chimerism can be achieved in situ activation and expansion of host tregs: a new approach to enhance donor chimerism and stable engraftment in major histocompatibility complex-matched allogeneic hematopoietic cell transplantation high incidence of graft failure in unrelated cord blood transplantation using a reducedintensity preparative regimen consisting of fludarabine and melphalan fludarabine-melphalan conditioning for aml and mds: alemtuzumab reduces acute and chronic gvhd without affecting long-term outcomes the authors declare no conflict of interest. key: cord- -pc vayyl authors: ono, shintaro; okano, tsubasa; hoshino, akihiro; yanagimachi, masakatsu; hamamoto, kazuko; nakazawa, yozo; imamura, toshihiko; onuma, masaei; niizuma, hidetaka; sasahara, yoji; tsujimoto, hiroshi; wada, taizo; kunisaki, reiko; takagi, masatoshi; imai, kohsuke; morio, tomohiro; kanegane, hirokazu title: hematopoietic stem cell transplantation for xiap deficiency in japan date: - - journal: j clin immunol doi: . /s - - - sha: doc_id: cord_uid: pc vayyl background: x-linked inhibitor of apoptosis protein (xiap) deficiency is a rare immunodeficiency that is characterized by recurrent hemophagocytic lymphohistiocytosis (hlh) and splenomegaly and sometimes associated with refractory inflammatory bowel disease (ibd). although hematopoietic stem cell transplantation (hsct) is the only curative therapy, the outcomes of hsct for xiap deficiency remain unsatisfactory compared with those for slam-associated protein deficiency and familial hlh. aim: to investigate the outcomes and adverse events of hsct for patients with xiap deficiency, a national survey was conducted. methods: a spreadsheet questionnaire was sent to physicians who had provided hsct treatment for patients with xiap deficiency in japan. results: up to the end of september , patients with xiap deficiency had undergone hsct in japan, of whom ( %) had survived. all surviving patients had received a fludarabine-based reduced intensity conditioning (ric) regimen. although patients developed post-hsct hlh, of them survived after etoposide administration. in addition, the ibd associated with xiap deficiency improved remarkably after hsct in all affected cases. conclusion: the ric regimen and hlh control might be important factors for successful hsct outcomes, with improved ibd, in patients with xiap deficiency. electronic supplementary material: the online version of this article (doi: . /s - - - ) contains supplementary material, which is available to authorized users. introduction x-linked lymphoproliferative syndrome (xlp) is a rare inherited immunodeficiency characterized by an extreme vulnerability to epstein-barr virus (ebv) infection, frequently resulting in hemophagocytic lymphohistiocytosis (hlh) [ ] . major clinical phenotypes of xlp include fulminant infectious mononucleosis or ebv-associated hlh (∼ %), lymphoproliferative disorder (∼ %), and dysgammaglobulinemia (∼ %) [ , ] . the responsible gene was first identified as that encoding the sh d a or slam-associated protein (sap), located in the region of xq [ ] [ ] [ ] [ ] . in another cohort of patients with ebv-driven hlh, mutations in the x-linked inhibitor of apoptosis protein (xiap) gene (also known as the baculoviral iap repeat-containing protein (birc ) gene) were identified [ ] . although the sh d a and xiap genes are close together on xq , the molecular pathogenesis and clinical features of these diseases seem to be distinct [ , ] . xiap is a potent inhibitor of programmed cell apoptosis, blocking the activated forms of the effector caspases , , and via its bir and domains [ ] . patients with xiap deficiency are often affected with hlh, splenomegaly, inflammatory bowel disease (ibd), variable hypogammaglobulinemia, and autoinflammatory phenomena. monocytes from patients with xiap deficiency are impaired in their ability to secrete cytokines (including tnf-α, il- , il- , and mcp- ) in response to stimulation with nucleotide-binding oligomerization domain-containing protein (nod ) ligands. nod is the strongest genetic risk factor associated with crohn's disease [ ] [ ] [ ] . thus, one of the characteristic symptoms of xiap deficiency is ibd. hematopoietic stem cell transplantation (hsct) is the only curative treatment for xiap deficiency. however, hsct for xiap deficiency has been associated with a poor prognosis compared with that for sap deficiency [ , ] , as demonstrated by a retrospective international survey of xiap-deficient patients who were treated with the procedure [ ] . this study revealed that treatment with reduced intensity conditioning (ric) resulted in apparently better prognosis than treatment with myeloablative conditioning (mac), where of patients in the ric group survived, as opposed to only of patients in the mac group. the -year probabilities of survival were % for the mac group and % for the ric group. the major causes of death were hepatic veno-occlusive disease and pulmonary toxicity in the mac group and pneumonia, respiratory failure, and ongoing hlh in the ric group. to investigate the outcomes and adverse events of hsct for japanese patients with xiap deficiency, we conducted a national survey and identified a total of patients with the disorder from unrelated families, including previously reported patients [ ] [ ] [ ] . of the patients who underwent hsct, ( %) have survived. a spreadsheet questionnaire was sent to physicians who had provided treatment to patients with xiap deficiency in japan. patients and were previously reported [ , ] . all patients and families provided informed consent for genetic analyses in accordance with the declaration of helsinki, and the study protocol was approved by the ethics committee of tokyo medical and dental university. the day of hsct was defined as day . the first of the consecutive days with an absolute neutrophil count (anc) of . × /l or more was defined as the day of engraftment. primary graft failure was defined as failure to maintain an anc of . × /l for consecutive days after hsct, by day with bone marrow (bm) or peripheral blood stem cell grafts or by day with cord blood (cb) grafts [ ] . secondary graft failure was defined as initial engraftment followed by a decline of donor cells to < % [ ] . engraftment and chimerism of whole blood were measured using either xy fluorescence in situ hybridization for sexmismatched donors or variable number of tandem repeat analysis for same-sex donors. full and mixed chimerisms were described by detection of > and - % of donor hematopoietic stem cells in the recipient's bm or peripheral blood, respectively [ ] . conditioning regimens were classified as mac if they contained an alkylating agent (busulfan mg/ kg) or total body irradiation (tbi) at a dose that would not allow autologous bm recovery. conditioning regimens were classified as ric if they did not meet the definition of the mac regimen [ ] . if there was uncertainty regarding the intensity of the regimen (patient ), it was classified as an intermediate intensity regimen. acute graft-versus-host disease (gvhd) and chronic gvhd were graded according to the standard criteria [ , ] . the probability of survival was estimated through the kaplan-meier method. we used all patient data for determining the probability of survival, except for patient whose observation period was too short to evaluate long-term survival. infection or reactivation of viruses including ebv, cytomegalovirus (cmv), and adenovirus was periodically monitored by the antigenemia or quantitative pcr methods. twenty-nine patients with xiap deficiency were identified from families in japan. the characteristics of these patients are shown in supplementary table . twenty-three of the patients ( %) developed hlh. thirteen patients ( %) were affected by ibd by years of age, with the age at ibd onset ranging from months to years. the cumulative percentage of patients who experienced ibd is shown in supplementary figure . ten of the patients had undergone hsct by the end of september in japan. the median age at hsct was . years (range . - years). the indication for hsct was active hlh in patients, refractory ibd in patients, and both hlh and ibd in patient. the patient characteristics and xiap mutations are listed in table . the graft characteristics and conditioning regimens are shown in table . the graft source was unrelated bm in patients, related bm in patient, and cb in patients. four patients received fully matched related (n = ) and unrelated (n = ) grafts based on hla antigens (hla-a, hla-b, hla-c, and hla-drb ). five patients received singleallele mismatched unrelated grafts, and patient (patient ) received allele mismatched cb. nine patients received ric, and patient (patient ) received intermediate intensity conditioning including fludarabine, melphalan, cyclophosphamide, and tbi gy. fludarabine ( - mg/m ), melphalan ( - mg/m ), and a low dose ( - gy) of tbi or total lymphoid irradiation (tli) with or without anti-thymocyte globulin were , cyclophosphamide ( mg/kg), and tbi gy. etoposide (total - mg/ m ) was additionally administered to patients (patients , , , , and ) as a preconditioning regimen. the gvhd prophylaxis regimen was tacrolimus and methotrexate in patients, and tacrolimus only in patient. eight patients ( %) experienced hlh before hsct, and of them were not relieved until the conditioning regimen was started. six patients ( %) suffered from ibd, and all were refractory to conservative ibd treatment, which became the indication for hsct. all surviving patients were engrafted for a median of . days (range - days). eight patients ( %) maintained full donor type chimerism, and only patient developed mixed chimerism of % donor type. there were no primary or secondary graft failures in this cohort. the hsct-related adverse events are shown in table . five patients developed post-hsct hlh, of whom were treated with etoposide and dexamethasone palmitate (dp) or prednisolone. five patients (patients , , , , and ) were given etoposide as a conditioning regimen, and of them (patients and ) developed post-hsct hlh, whereas of patients without etoposide administration developed this condition. seven patients were complicated with acute gvhd, but of them showed only skin gvhd. only one patient (patient ) developed grade iii acute gvhd (gut , skin ). nine of the patients are currently alive and well at a median of . months (range - months) after hsct, with eastern cooperative oncology group performance status or ( table ) . patient , who received intermediate intensity conditioning, died on day post-hsct from complications due to engraftment syndrome, hlh, and acute respiratory distress syndrome [ ] . except for patient , who was followed for months only, we evaluated the probability of survival in the other patients to be % (fig. ) . although patient (a sibling of patient ) developed thrombotic microangiopathy and pulmonary artery hypertension, he improved after administration of phosphodiesterase inhibitors and plasma exchange. intriguingly, patients (patients , , , , , and ) were associated with ibd before hsct, all cases of which improved remarkably after hsct and maintained remission without any treatments. ibd was relieved during the conditioning regimen in all patients, at least by the point of engraftment. colonoscopic findings for patients , , and are shown tac i (skin ) atg antithymocyte globulin, gvhd graft versus host disease, cb cord blood, n/a not available, urbm unrelated bone marrow, rbm related bone marrow, flu fludarabine, mel melphalan, cy cyclophosphamide, tbi total body irradiation, eto etoposide, tli total lymphoid irradiation, mtx methotrexate, tac tacrolimus in fig. . colonoscopy before hsct revealed multiple ulcers and bleeding in the sigmoid colon, whereas that after hsct showed improvement to normal colon appearance. six patients had no virus infection or reactivation, but patients developed virus reactivation, including human herpesvirus , bk virus, jc virus, and cmv. they were successfully treated with antiviral drugs or improved without medication. xlp (sap deficiency and xiap deficiency) is a rare but lifethreatening disease. a large cohort study showed that most patients with xlp died by the age of years, and more than % of the patients had died before the age of years mainly as a result of fulminant infectious mononucelosis and hlh [ ] . although hsct is the only curative treatment for xiap deficiency, the result of a previous study revealed that table outcomes in the japanese patients with xiap deficiency post-hsct patient before and after hematopoietic stem cell transplantation (hsct). they revealed multiple ulcers before hsct, but showed a normal bowel mucosa after the procedure transplantation outcomes were apparently poorer than those for sap deficiency [ ] . an international survey reported that % of patients with xiap deficiency developed hlh, and % were affected by ibd [ ] . in a japanese survey, % of patients developed hlh (supplementary table ). although this hlh frequency is higher than the international average, it is equivalent to those in france ( / , %) and the usa ( / , %). twelve patients ( %) developed ibd up to the age of years, and this percentage is relatively higher than those in other countries ( - %) [ ] . the occurrence of ibd in xiap deficiency might be associated with ethnic background. in this study, of the patients underwent hsct, and of them survived. the transplantations were performed in different institutions, but all conditioning regimens were ric, except that for patient , who was given an intermediate intensity regimen, but died of hlh and acute respiratory distress syndrome on day post-hsct, in which virus infection or reactivation might not be involved. thus, the intensity of conditioning might be linked to survival. in the case of hsct for patients with xiap deficiency, ric was apparently superior to mac [ ] . successful hsct outcomes based on ric for xiap-deficient patients have been reported in several studies [ , , ] . patients , , and had the same r x mutation, and they developed fatal or severe regimen-related complications, indicating a possible association of gene mutation with the severity of disease. the probability of survival after hsct was %, and the outcome was better than that reported by a previous study, although the follow-up period was limited. it is possible that ric and hlh control might contribute to the better outcome of hsct for xiap deficiency. the high level of donor chimerism in all surviving patients was remarkable, although all of them underwent the ric regimen. a previous international survey reported that % of patients on a ric regimen of fludarabine, melphalan, and alemtuzumab developed mixed chimerism [ ] . in our study, the ric regimen consisted of fludarabine, melphalan, and low-dose tbi or tli, and only patient developed mixed chimerism. therefore, low-dose tbi or tli might contribute to a high level of donor chimerism. five of the patients ( %) developed hlh after hsct. hlh induced by uncontrolled macrophage activation is often a fatal complication after hsct and frequently leads to primary and secondary graft failures. thus, prophylaxis of post-hsct hlh might play an important role in successful hsct outcomes. since dp decreases the viability of primary human macrophages via glucocorticoid receptors in the cytoplasm, it is considered effective for the prevention of post-hsct hlh [ ] . in addition, etoposide can be considered as a preconditioning regimen to reduce post-hsct hlh [ ] . in our cohort, post-hsct hlh developed in of patients ( %) treated with etoposide, whereas of patients ( %) without etoposide treatment developed this condition. therefore, the use of dp and etoposide could reduce the risk of hlh related to hsct for xiap-deficient patients. alemtuzumab against cd was not used for the conditioning regimen in this study owing to its limited usage in japan. cd is highly expressed in monocytes and macrophages, and alemtuzumab administration might reduce the risk of post-hsct hlh and total doses of etoposide [ ] . ibd or hemorrhagic colitis is a characteristic finding in xiap deficiency. interestingly, ibd improved remarkably after hsct in all affected cases and maintained remission without any further treatments for the condition. ibd associated with primary immunodeficiency, including xiap deficiency or il- /il- r deficiency, was reported to be remarkably improved after hsct [ , ] . in xiap deficiency, monocytes are impaired in their ability to secrete cytokines mediated by nod stimulation, which may cause ibd. thus, it is supposed that ibd in xiap deficiency can be cured by hsct, based on the transplantation of normal monocytes. in conclusion, a national survey on xiap deficiency revealed that the probability of survival after hsct was prominent, and it is assumed that an ric regimen and hlh control might be important factors for successful outcomes. in addition, the ibd associated with xiap deficiency could be cured by hsct. correlation of mutations of the sh d a gene and epstein-barr virus infection with clinical phenotype and outcome in xlinked lymphoproliferative disease x-linked lymphoproliferative disease: twentyfive years after the discovery xlp: clinical features and molecular etiology due to mutations in sh d a encoding sap host response to ebv infection in x-linked lymphoproliferative disease results from mutations in an sh -domain encoding gene inactivating mutations in an sh domainencoding gene in x-linked lymphoproliferative syndrome the xlinked lymphoproliferative-disease gene product sap regulates signals induced through the co-receptor slam xiap deficiency in humans causes an x-linked lymphoproliferative syndrome human inhibitor of apoptosis proteins: why xiap is the black sheep of the family x-linked lymphoproliferative syndromes: brothers or distant cousins x-linked lymphoproliferative syndrome: a genetic condition typified by the triad of infection, immunodeficiency and lymphoma making a definitive diagnosis: successful clinical application of whole exome sequencing in a child with intractable inflammatory bowel disease association of nod leucine-rich repeat variants with susceptibility to crohn's disease host-microbe interactions have shaped the genetic architecture of inflammatory bowel disease reduced-intensity conditioning hematopoietic cell transplantation is an effective treatment for patients with slamassociated protein deficiency/x-linked lymphoproliferative disease type allogeneic hematopoietic cell transplantation for xiap deficiency: an international survey reveals poor outcomes clinical and genetic characteristics of xiap deficiency in japan sustained elevation of serum interleukin- and its association with hemophagocytic lymphohistiocytosis in xiap deficiency a female patient with incomplete hemophagocytic lymphohistiocytosis caused by a heterozygous xiap mutation associated with nonrandom x-chromosome inactivation skewed towards the wild-type xiap allele successful treatment of idiopathic colitis related to xiap deficiency with allo-hsct using reduced-intensity conditioning long-term followup of patients who experienced graft failure postallogeneic progenitor cell transplantation. results of a single institution analysis graft rejection after unrelated donor hematopoietic stem cell transplantation for thalassemia is associated with nonpermissive hla-dpb disparity in host-versus graft direction reduced-intensity conditioning hematopoietic sct for pediatric patients with lad- : clinical efficacy and importance of chimerism defining the intensity of conditioning regimens: working definitions clinical manifestations of graft-versus-host disease in human recipients of marrow from hla-matched sibling donors chronic cutaneous graft-versus-host disease in man x-linked inhibitor of apoptosis protein deficiency: more than an x-linked lymphoproliferative syndrome successful stem cell transplant with antibody-based conditioning for xiap deficiency with refractory hemophagocytic lymphohistiocytosis successful allogeneic hematopoietic stem cell transplantation in xiap deficiency using reduced-intensity conditioning dexamethasone palmitate successfully attenuates hemophagocytic syndrome after allogeneic stem cell transplantation: macrophage-targeted steroid therapy etoposide-containing conditioning regimen reduces the occurrence of hemophagocytic lymphohistiocytosis after sct cross-linking of the campath- antigen (cd ) triggers activation of normal human t lymphocytes clinical outcome in il- -and il- receptordeficient patients with or without hematopoietic stem cell transplantation acknowledgments this study was supported by grants from the ministry of education, culture, sports, science, and technology of japan (to h.k.) and the ministry of health, labour, and welfare of japan (to t.m). conflicts of interest the authors have no conflicts of interest to disclose. key: cord- -nzjm zyq authors: carreras, enric; cooke, kenneth r. title: noninfectious pulmonary complications date: - - journal: the ebmt handbook doi: . / - - - - _ sha: doc_id: cord_uid: nzjm zyq lung injury occurs frequently following hsct and significantly contributes to morbidity and mortality in the immediate post transplant period and in the months and years that follow. ideally, any respiratory/pulmonary complication observed after hsct must be evaluated following a predetermined institutional protocol (lucena et al. ) , which should include: . noninvasive tests: blood samples for culture and antigen determination, sputum culture, nasopharyngeal swabs testing cmv, respiratory syncytial virus (rsv), legionella, pneumocystis jirovecii (pj), parainfluenza virus (piv), adenovirus (adv), as well as urinary antigen tests and chest x-ray. . if negative → empirical treatment (variable behavior; some centers start empirical treatment before the bal, but many others start the treatment after bal). . if no response in a maximum of - days (or if galactomannan (gm) +) → (a) high-resolution chest-computed tomography (hrct despite not being included in most classifications of pulmonary complications after hsct, pulmonary edema (pe) as a consequence of a fluid overload (fo) is extremely frequent (rondón et al. characterized by development around day + after hsct of fever and nonproductive cough, dyspnea, tachypnea, hypoxemia, rales, and diffuse alveolar or interstitial infiltrates on x-rays or ct scans. all of the following must be present for accepting the ips diagnosis: the pathophysiology of ips is complex. data generated using experimental models support that ips is a process in which the lung is susceptible to two distinct but interrelated pathways of immune-mediated injury: a t-cell axis and an inflammatory cytokine axis. these distinct but related pathways of inflammation culminate in the recruitment of immune cells to the lung leading to tissue damage and dysfunction (cooke and yanik ) incidence -the strict methodology required to establish ips diagnosis and the increased use of ric have reduced its incidence of % to % observed years ago (at that time ips was called idiopathic pneumonia) -this reduction runs in parallel of the improvement in the diagnostic methodologies to detect infectious pathogens. however, the frequent absence of response to the specific treatment against a detected pathogen suggests that the true incidence of ips may be underestimated -nowadays: < % of allo-hsct ( % after mac; % after ric) timing -within first days after bmt, usually observed between days + and + ( years ago: around days + to + ) -late ips can be observed but they are exceptional (thompson et al. ) risk factors (from cooke and yanik ) older age / karnofsky index < / higher interval diagnosis-hsct mac or tbi (≥ gy) / hla disparity / gvhd prophylaxis with mtx acute gvhd/previous viral infection / other malignancies than leukemia supportive measures -supplemental o therapy -mechanical ventilation (invasive or not [high-flow nasal o , cpap]) -empiric broad-spectrum antimicrobials -strict control of fluids balance/hemofiltration specific treatment as mentioned, lung injury in ips can occur through two pathways, the tnf-alfa/lps dependent and il /il dependent (cooke and yanik ) ; consequently, treatment options are focused in these directions • methyl-pdn ≤ mg/kg/d; if not clear response, consider as soon as possible: • anti-tnfα: etanercept . mg/kg twice weekly (maximum of doses) + systemic steroids ( mg/ kg/d). the randomized study of etanercept + steroids vs. steroids + placebo was terminated prematurely due to slow accrual. in the limited number of patients examined, there were no differences in response rates (≈ %) at day + . these results do not necessarily imply that this agent is not effective (lack of evidence does not imply lack of effectiveness) (yanik et al. ) . in a phase ii trial in children, the cr rate was % and y survival was % . this combination has also been shown to be effective in exceptional cases of late ips with a % of cr and a y survival of % among responders (thompson et al. diffuse alveolar hemorrhage (dah) is a relevant cause of acute respiratory failure that occurs in - % of recipients, with similar incidence in both auto-and allo-hsct recipients (afessa et al. a) . dah is probably a consequence of damage to the alveolar capillary basement membrane (see chap. ). it is difficult to differentiate a true dah from the alveolar hemorrhage associated with an infection (majhail et al. ). usually observed within the first month after hsct (a median of days), often during the pre-engraftment phase; however, later onset is encountered in up to % of cases the clinical manifestations are those of all ips. hemoptysis is exceptional diagnosis based on bal: same criteria as ips plus a differential characteristic; the progressive bloodier return of bal fluid aliquots, in at least three segmental bronchi, indicating the presence of blood in the alveoli (or % hemosiderin-laden macrophage, although their absence does not exclude the diagnosis as it can take h to appear). note: dah can have infectious or noninfectious etiologies (majhail et al. ) risk factors -higher incidence after tbi and high-dose cy -similar incidence among mac and ric -there is no correlation with the platelet counts differential diagnosis with -classic ips: very difficult, only by means of bal. ips usually appears after the engraftment, predominates in allo-hsct, does not respond to steroids, and progresses to fibrosis in % of cases (only % on dah). note: noninfectious dah falls under the "diagnostic umbrella" of ips (panoskaltsis-mortari et al. ) -perds: almost impossible except for lba progressively bloodier -pulmonary hemorrhage: by fob, no blood is seen in dah -dah associated with infection: impossible without detection of the pathogen (majhail et al. ) treatment -although systematically treated with high doses of methyl-pdn ( - mg q h × days, followed by tapered dosage over - weeks) and aminocaproic acid (aca), the overall response to this treatment is disappointing (rathi et al. ) -a recent study seems to show that the best treatment is to use low steroid doses (≤ mg/d) ± aca (rathi et al. ) -factor viia addition does not appear to improve the results obtained with pdn (elinoff et al. ) -try to avoid mechanical ventilation by means of cpap prognosis -poor: overall mortality as high as % by day (rathi et al. ) -less than % of patients die as a direct consequence of dah, but the frequent evolution to mof increases mortality to > % ( % in auto and % in allo-hsct) (afessa et al. b ) -dah that appear early after allo-hsct ( % early vs. % late) or after auto-hsct have a better prognosis (afessa et al. b; majhail et al. ) the same as cgvhd but specifically involving the lung (cooke et al. ) . its course may be aggravated by respiratory infections, viral infections, and gastroesophageal reflux timing and incidence -average starting period: ( - ) months -incidence: % at years in the longest series (arora et al. ) ; % in a prospective study (bergeron et al. ) clinical manifestations -variable clinical course, usually insidious onset with progressive deterioration. sometimes can present as an acute, fulminating course -progressive breathlessness, nonproductive cough, and wheezing, although some asymptomatic cases are only detected by pfts. -it is necessary to carry out pft every m in the first year after hsct for an early detection a -in > % of the bos, there are chronic gvhd in other locations (continued) in addition to late-onset ips mentioned before and some other exceptional complications (thromboembolisms, pneumomediastinum), there are two forms of chronic pulmonary dysfunction commonly observed in patients surviving more than days after allo-hsct. one is an obstructive lung disease (bronchiolitis obliterans syndrome, bos) and the other a restrictive lung disease (cryptogenetic organizing pneumonia, cop). a recent prospective study showed that among patients included after day + , the cumulative incidence of lonipc is %, and that of bos is % at years among allo-hsct recipients (bergeron et al. ) . another study shows the impact of these complications on -year survival ( % with vs. % w/o lonipc) (nishio et al. ). pathogenesis, timing, incidence, clinical manifestations, diagnosis, and radiology of bos are shown in table . . treatment and prognosis of bos are included in table . . formerly called boop (bronchiolitis obliterans with organizational pneumonia). cop is a lonipc of that is associated with restrictive pulmonary dysfunction. reportedly, the incidence of cop among hsct recipients is increasing due to the use of transbronchial biopsies as diagnostic tool. the greatest diagnostic challenge is the differentiation of cop from bos (see table . ) (yoshihara et al. ; cooke et al. ). (chien et al. ; uhlving et al. ) -clinical manifestation (may be asymptomatic and only detected on pft) + -absence of active infection (demonstrated by bal) + -chronic gvhd in other locations b + -obstructive alteration with air entrapment (fev < % nv or > % decrease; ratio fev /fvc ratio < . ; residual volume > %) with nonsignificant bronchodilator test and a decreased dlco + -compatible radiology (see below) • definitive: histologic confirmation by thoracotomy, vats, or transbronchial biopsy c radiology -chest x-ray: normal or with signs of hyperinflation -ct scan: radiological pattern of constrictive bronchiolitis with aerial entrapment, attenuation in mosaic, bronchiectasis and bronchial wall thickening, characteristic air trapping at exhalation dlco transfer capacity of co, fev maximum expiratory volume in the first second, fvc forced vital capacity, vats video-assisted thoracoscopic surgery a some experts consider that a % decrease in the fev basal after hsct should make you suspect in bos diagnosis b if the lung is the only organ with cgvhd, a biopsy is needed to confirm the diagnosis (nih criteria) c rarely transbronchial biopsy is used (low sensitivity and low predictive value) to establish a diagnosis that is eminently clinical. if histology is available, the term bronchiolitis obliterans can be used; if not available, the process is referred to as bos anti-infectious prophylaxis if hypogammaglobulinemia: ivig treatment of gastroesophageal reflux respiratory physiotherapy specific treatment • prednisone: - . mg/kg/day, transient and unsatisfactory response in most cases. the addition of csa, azathioprine, atg, or photopheresis has few advantages • budesonide/inhaled formoterol has been shown to be transiently effective in % of the patients (bergeron et al. ) • etanercept/infliximab: effective in some cases (yanik et al. ) • fam combination therapy: effective in disease stabilization a : -fluticasone inhaled mcg c/ h (adult), mcg in children + -azithromycin mg/d (adults), mg/kg/d (children) b + -montelukast mg orally at night (adults), mg (children) two weeks before fam increase (or start) pdn to mg/kg/d, then decrease . mg/kg/d × week (williams et al. ) • in bos controlled but with a severe residual respiratory insufficiency, lung transplantation may be considered after a few years (cheng et al. ) prognosis -trm is very high; % ( - %) at years of hsct almost always get associated with progressive respiratory failure and opportunistic infections -srv around % ( %- %) at years a fluticasone theoretically decreases the inflammatory pulmonary component; azithromycin reduces il- levels and neutrophilia; and montelukast is an antagonist of the leukotriene receptors (bronchodilator) b however, the allozithro randomized trial has shown that early administration of azithromycin resulted in worse airflow decline-free survival than did placebo; the value of these findings is limited by early termination of the trial (bergeron et al. ) • lung injury occurs frequently following hsct and significantly contributes to morbidity and mortality in the immediate post transplant period and in the months and years that follow. it can be observed in - % of recipients. • noninfectious lung injury following hsct may be mediated by either immune or nonimmune mechanisms and could represent up to the % of noninfectious mortality after allo-hsct. • most relevant noninfectious early pulmonary complications are pulmonary edema by fluid overflow, idiopathic pneumonia syndrome, and diffuse alveolar hemorrhage, a vascular endothelial syndrome. • the most relevant late-onset noninfectious pulmonary complications are bronchiolitis obliterans and cryptogenetic organizing pneumonia. • all of them have specific diagnostic criteria, management, treatment, and prognosis. predictive value of bronchiolitis obliterans syndrome stage p in chronic graft-versus-host disease of the lung outcome of diffuse alveolar hemorrhage in hematopoietic stem cell transplant recipients diffuse alveolar hemorrhage in hematopoietic stem cell transplant recipients late acute and chronic graft-versus-host disease after allogeneic hematopoietic cell transplantation budesonide/formoterol for bronchiolitis obliterans after hematopoietic stem cell transplantation effect of azithromycin on airflow decline-free survival after allogeneic hematopoietic stem cell transplant: the allozithro randomized clinical trial noninfectious lung complications after allogeneic haematopoietic stem cell transplantation outcomes of lung transplantation after allogeneic hematopoietic stem cell transplantation bronchiolitis obliterans syndrome after allogeneic hematopoietic stem cell transplantation-an increasingly recognized manifestation of chronic graft versus-host disease idiopathic pneumonia syndrome after bone marrow transplantation lung injury following hematopoietic cell transplantation the biology of chronic graft-versus-host disease: a task force report from the national institutes of health consensus development project on criteria for clinical trials in chronic graft-versus-host disease recombinant human factor viia for alveolar hemorrhage following allogeneic stem cell transplantation pulmonary complications in hematopoietic sct: a prospective study diffuse alveolar hemorrhage and infection-associated alveolar hemorrhage following hematopoietic stem cell transplantation: related and high-risk clinical syndromes late-onset non-infectious pulmonary complications following allogeneic hematopoietic stem cell transplantation in children an official american thoracic society research statement: noninfectious lung injury after hematopoietic stem cell transplantation: idiopathic pneumonia syndrome low-, mediumand high-dose steroids with or without aminocaproic acid in adult hematopoietic sct patients with diffuse alveolar hemorrhage impact of fluid overload as new toxicity category on hematopoietic stem cell transplantation outcomes idiopathic pneumonia syndrome after hematopoietic cell transplantation: evidence of occult infectious etiologies utility of early versus late fiberoptic bronchoscopy in the evaluation of new pulmonary infiltrates following hematopoietic stem cell transplantation etanercept and corticosteroid therapy for the treatment of late-onset idiopathic pneumonia syndrome bronchiolitis obliterans after allo-sct: clinical criteria and treatment options lung parenchyma-derived il- promotes il- a-dependent acute lung injury after allogeneic stem cell transplantation azithromycin, and montelukast treatment for new-onset bronchiolitis obliterans syndrome after hematopoietic cell transplantation tnf-receptor inhibitor therapy for the treatment of children with idiopathic pneumonia syndrome. a joint pediatric consortium and children's oncology group study (asct ) randomized, double-blind, placebo-controlled trial of soluble tumor necrosis factor receptor: enbrel (etanercept) for the treatment of idiopathic pneumonia syndrome after allogeneic stem cell transplantation: blood and marrow soluble tumor necrosis factor receptor: enbrel (etanercept) for sub-acute pulmonary dysfunction following allogeneic stem cell transplantation bronchiolitis obliterans syndrome (bos), bronchiolitis obliterans organizing pneumonia (boop), and other late-onset noninfectious pulmonary complications following allogeneic hematopoietic stem cell transplantation open access this chapter is licensed under the terms of the creative commons attribution . international license (http://creativecommons.org/licenses/by/ . /), which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the creative commons license and indicate if changes were made.the images or other third party material in this chapter are included in the chapter's creative commons license, unless indicated otherwise in a credit line to the material. if material is not included in the chapter's creative commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. key: cord- - wwy m k authors: marty, francisco m.; baden, lindsey r. title: infection in the hematopoietic stem cell transplant recipient date: journal: hematopoietic stem cell transplantation doi: . / - - - - _ sha: doc_id: cord_uid: wwy m k nan there are three key elements of the hsct procedure that determine the type and timing of the infectious risk profile after transplantation [ , ] : , a. the duration of neutropenia and mucosal injury which is a function of the conditioning regimen selected (myeloablative or not) and the stem cells' procurement (cord or adult; peripheral or bone marrow acquisition among adult donors). b. the strategy chosen to prevent gvhd among allogeneic recipients. t cell depletion and other t cell manipulation procedures lead to delayed recovery of lymphocyte function and provide a specific immune deficiency profile. c. the occurrence and severity of acute and chronic gvhd and its treatment [ , ] . the temporal course of infection following hsct can be divided into three time periods (fig. - ) [ , , , ] : . conditioning to engraftment the duration of this period has become dynamic and depends on the conditioning regimen itself, the source and dose of stem cells infused and whether growth factors are used. it usually ranges from five (with nonmyeloablative transplants) to days (with bone marrow or umbilical cord blood transplants). the combination of profound granulocytopenia and mucositis with myeloablative conditioning makes the patient particularly vulnerable to bacterial and candidal infections. in addition, infection present in the transplant recipient pre-transplant may be amplified by the granulocytopenic state and deficiencies of t and b-cell numbers and function. thus, control of pre-transplant infection is needed before initiating the conditioning regimen. prior to engraftment (both with autologous and allogeneic transplants), approximately percent of patients will have fever of unknown origin, with bloodstream infection in ~ . percent and pneumonia in ∼ percent. the risk of an invasive mold infection is related to the duration of neutropenia and the environmental strategy used in a transplant center. . engraftment to post-transplant day during this time period viral infections, particularly cytomegalovirus (cmv) and the other herpes group viruses, are the major concerns. the occurrence, severity and treatment modalities selected for acute gvhd further modulates and increases the risk of herpesvirus infections, especially cmv and epstein-barr virus (ebv), and invasive mold infections [ ] [ ] [ ] [ ] . . more than days post-transplant in the absence of gvhd, the incidence of infection decreases significantly, with varicella zoster virus (vzv), pneumocystis jiroveci (formerly carinii) pneumonia (pcp) and pneumococcal infection being the primary problems of this time period. routine use of prophylaxis, such as with trimethoprim/sulfamethoxazole and acyclovir, significantly decreases the occurrence of pcp and herpesvirus infections, respectively. in addition, late or relapsing cmv infection may manifest during this time. if gvhd is present, it is typically treated with significant augmentation of immunosuppressive therapy such as with high-dose corticosteroids and monoclonal antibodies. patients in this last category (gvhd under treatment) are at particular risk for invasive mold infection, cmv reactivation, pcp and other common and opportunistic pathogens. there are four modes in which antimicrobial therapy can be administered to the hsct patient [ ] : . a therapeutic mode, in which antimicrobial therapy is prescribed for the treatment and eradication of identified microbes causing clinical illness. . a prophylactic mode, in which antimicrobial therapy is prescribed to an entire population before an event to prevent clinically important infection. for such a strategy to be successful, the infection(s) being targeted must be important enough to justify the intervention, and the antimicrobial therapy prescribed must be nontoxic and inexpensive enough to justify the intervention. by far the most effective antimicrobial prophylactic strategy is low-dose trimethoprim-sulfamethoxazole, which has virtually eliminated the occurrence of pneumocystis jiroveci, listeria monocytogenes, nocardia sp, and toxoplasma gondii in patients who adhere to the regimen. other prophylactic strategies commonly utilized in hsct patients include acyclovir to prevent herpes simplex virus (hsv) and vzv reactivation, fluoroquinolones [ ] to prevent gram-negative sepsis and fluconazole to prevent yeast infection. . an empiric mode, in which antimicrobial therapy is administered in response to a symptom complex. in this context, empiric antimicrobial therapy is initiated during the period of profound granulocytopenia in response to fever +/− rigors or subtle signs of sepsis (unexplained hypotension, tachypnea, an ongoing volume requirement, or acidosis). in the patient deemed not to be a therapeutic emergency, initial therapy is usually aimed at aerobic gramnegative bacilli (e.g., the enterobacteriaceae and pseudomonas aeruginosa). a variety of drugs have been utilized for this purpose, depending in part on the nature of particular problem organisms found at a given medical center. advanced spectrum beta-lactams (e.g., ceftazidime, piperacillin or imipenem), either alone or together with an aminoglycoside or a fluoroquinolone, are the mainstays of this approach. thus, empiric therapy is based on an algorithm rather than on microbiologic or other studies. . a preemptive mode, in which antimicrobial therapy is prescribed to a proportion of patients deemed to be at particularly high risk because of clinical/epidemiologic information or the isolation of microbial pathogens. examples of preemptive therapy in hsct are the molecular surveillance of cmv linked to deployment of ganciclovir or, more recently, the use of galactomannan monitoring for initiation of anti-aspergillus antifungal treatment [ ] . given the nature, duration and severity of host defense defects present in hsct patients, it is not surprising that bacterial infection is a regular feature of the post-transplant course. the most common involved sites include blood stream (often catheter-related), lung, gastrointestinal tract and skin/soft tissue. the greatest rate of bacterial infections occur during the period prior to engraftment; this rate is a product of granulocytopenia, mucositis that permits the translocation of bacteria and yeast from the oral cavity and gut into circulation, and the presence of vascular access devices that traverse the skin and serve as direct conduits into the systemic circulation. thus, the primary mucocutaneous barriers to infection are compromised, and the absence of granulocytes only amplifies the susceptibility of the patient [ , , ] . in an attempt to decrease bacterial infections during the neutropenic period, especially those due to gram-negative bacilli, strategies of prophylactic antimicrobial use have been studied, including the use of trimethoprimsulfamethoxazole and fluoroquinolones. some studies, most recently with levofloxacin, have demonstrated benefit in decreasing the occurrence of fever and microbiologically-confirmed bacterial infections [ ] [ ] [ ] . however, significant concerns regarding this approach have been raised given that no mortality benefit has been demonstrated, the emergence of resistant organisms, and the impairment this widespread antimicrobial approach has on the use of quinolones in future oral outpatient management. thus, in many transplant centers, an empiric antibacterial regimen targeting pseudomonas and other enterobacteriaceae in response to fever or other infectious syndromes remains a preferred approach. whereas gram-negative bacteremia was the major cause of blood stream infection to years ago, today gram-positive organisms are the most frequent cause of positive blood cultures. the possible reasons for this shift are many: the widespread use of fluoroquinolones, with their potent activity against gram-negative bacteria, as prophylaxis during this period; the presence of indwelling central venous catheters for prolonged periods; and the widespread use of systemic anti-gram-negative therapy all contribute to the gram-positive predominance. the bacteria isolated during the preengraftment period, then, include staphylococci (especially coagulase-negative staphylococcus), viridans streptococci, enterococci and corynebacteria, with fewer isolates of enterobacteriaceae or pseudomonas aeruginosa being identified. an increasing problem in the hsct population is antibiotic resistant organisms, particularly vancomycin-resistant enterococci, methicillin-resistant staphylococcus aureus, and resistant gram-negative bacilli (such as extended spectrum β-lactamase producing klebsiella and chromosomal inducible βlactamase producing enterobacter species) [ , , , , [ ] [ ] [ ] [ ] [ ] . the typical approach for the severely granulocytopenic patient at present is the initiation of empiric antibacterial therapy in response to an unexplained fever or other signs of sepsis. what remains controversial is what the regimen should be. since clinical deterioration can occur rapidly with untreated gram-negative sepsis in the granulocytopenic patient, anti-gramnegative therapy is always employed. the traditional approach of a β-lactam (e.g., piperacillin) plus an aminoglycoside is still favored by some experts, although nephrotoxicity from the aminoglycoside has led to the trial of other approaches, including the substitution of a fluoroquinolone for the aminoglycoside, or the prescription of a single advanced spectrum drug such as ceftazidime, cefepime, imipenem or meropenem. if fluoroquinolone prophylaxis has been utilized, then its use as a therapeutic agent may be diminished. empiric fluoroquinolone monotherapy is inferior to other regimens, and if pure aerobic gram-negative agents are utilized, (e.g., aztreonam, aminoglycosides) due to confirmed severe beta-lactam hypersensitivity, then the addition of empiric gram-positive coverage that targets aerobic and anaerobic streptococci of the gastrointestinal tract should be considered. use of extended interval (once-daily dosing) aminoglycoside administration may be safer and as effective. the second area of controversy is whether empiric gram-positive treatment should be initiated at the same time, given the preponderance of grampositive infection. as there is typically time to evaluate culture data and deploy targeted gram-positive antimicrobial therapy rather than empiricism, vancomycin should rarely be required empirically. furthermore, empiric gram-positive coverage is not associated with better outcomes [ ] . indications for the immediate initiation of vancomycin as part of the empiric therapy regimen include the following [ , , [ ] [ ] [ ] [ ] : catheter-related sepsis is likely because of evidence of infection at the insertion site (or within the tunnel), severe illness such as shock and/or respiratory distress are present, the patient is at particular risk for seeding of a prosthetic device (e.g., a prosthetic valve, a hip prosthesis, etc.), or the empiric gram-negative coverage exclusively covers aerobic gram-negative rods -such as the combination of aztreonam and gentamicin. vancomycin or other anti-staphylococcal agents should be started if cultures become positive for gram-positive cocci. in our experience, vancomycin can be discontinued safely in patients in whom vancomycin was started empirically, but in whom blood cultures remain negative after to hours and there is no specific syndrome, such as cellulitis, that requires treatment with vancomycin. on the other hand, empirical treatment against gram-negative organisms should be continued until resolution of neutropenia, whether fevers resolve or not [ ] . the emergence and persistence of multidrug-resistant organisms should guide local practice in a dynamic fashion. indwelling long-term catheters remain a feature of the early post-transplant period to provide chemotherapy, nutritional and blood product support until stable engraftment. routine anti-gram-positive antimicrobial therapy is not required just because a central catheter is in place for the prevention and management of catheter-related infections [ , ] . the use of antimicrobial-coated catheters should be studied in this population, especially when non-tunneled catheters need to be used. nonantimicrobial-based strategies to prevent bacterial infections during the neutropenic period include the systematic use of hand hygiene and the use of mask and gloves by health care personnel and family members. other nonantimicrobial strategies which may be beneficial in preventing infections, but have not been tested in hsct, include the use of palifermin to prevent mucositis [ ] in patients undergoing myeloablative conditioning. after engraftment, the risk of bacterial infections depend on the community exposures to common and opportunistic bacteria (e.g., nocardia, rhodococcus, listeria), the presence of acute and chronic gvhd, the degree of b-cell reconstitution and the use of trimethoprim-sulfamethoxazole prophylaxis. patients with chronic gvhd are at risk for invasive infection from encapsulated organisms, particularly streptococcus pneumoniae, haemophilus influenzae and neisseria meningitidis. it is postulated that the combination of b lymphocyte dysfunction secondary to the conditioning regimen and the effects of gvhd and its treatment have resulted in the loss and failure to develop an opsonizing antibody to these organisms, particularly streptococcus pneumoniae. in addition, for at least one to two years post-transplant, hsct patients have an inadequate response to pneumococcal vaccine. as igg levels are often low for some time after hsct, they should be routinely monitored, with replacement being considered when the igg level falls below mg/ml [ , ] . in addition, antimicrobial prophylaxis, such as with low-dose trimethoprimsulfamethoxazole (one single strength tablet daily for pcp prophylaxis), may afford further protection against this problem [ , ] . there are several classes of viral infection of particular importance in the hsct recipient: those due to herpesviruses (cmv, ebv, hsv, vzv and human herpesvirus- [hhv- ]); those due to hepatitis viruses (e.g., hepatitis b [hbv]); those due to respiratory viruses (e.g., influenza, rsv, parainfluenza, adenoviruses, and others), and those due to polyoma viruses. the human herpesviruses share a number of characteristics that make them particularly successful pathogens in hsct recipients [ , , ] : . latency once infected with a herpesvirus, one is infected for life, with a circulating antibody (seropositivity) in the absence of active viral replication being the classic marker for latent infection. reactivation from latency may be triggered by tumor necrosis factor (tnf), with the catecholamines epinephrine and norepinephrine and proinflammatory prostaglandins also playing a role. thus, the virus may be reactivated by such processes as sepsis, gvhd, allogeneic reactions, okt and antilymphocyte globulin. once a replicating virus is present, medications such as cyclosporine, tacrolimus and prednisone may significantly amplify the viral replication. . cell association these viruses are highly cell-associated, meaning that transmission occurs through intimate person-to-person contact, or transfusion or transplantation of latently or actively replicating cells from a seropositive donor. humoral immunity is, hence, less important than cell-mediated immunity. indeed, the key host defense is accomplished by major histocompatibility complex (mhc)-restricted, virus-specific, cytotoxic t cells, just that component of host defense most affected by gvhd and its treatment. . oncogenesis herpesviruses, such as ebv and hhv- , play a direct role in oncogenesis-causing post-transplant lymphoproliferative disease (ptld) and kaposi's sarcoma, respectively. herpesviruses may also play an indirect role in oncogenesis with symptomatic cmv disease, increasing the incidence of ebv-associated ptld severalfold. . indirect effects in addition to the direct causation of infectious disease syndromes, human herpesviruses, particularly cmv, have indirect effects that are clinically important. it is believed that cytokines, chemokines and growth factors produced in response to viral replication may be responsible for these effects. they include, in addition to the modulation of oncogenesis, increasing the net state of immunosuppression so that the risk of opportunistic infection is increased. this last point is particularly important, as a variety of experiments have shown that gvhd and infection are closely linked by the production of these mediators. that is, there is a bidirectional trafficking of mediators between these two processes. the clinically most important direct effects of cmv in the hsct recipient are pneumonia and gastrointestinal disease. before effective antiviral treatment became available, cmv pneumonia occurred in to percent of seropositive recipients and had an associated mortality around percent [ ] . cmv commonly causes fever in the absence of preemptive treatment, and end-organ disease (hepatitis, bone marrow dysfunction, retinitis, and encephalitis) may occur. among allogeneic hsct recipients, the risk of cmv reactivation ( - %) and end-organ disease is greatest in the seropositive recipient who receives a graft from a cmv seronegative donor (cmv d−/r+), likely due to the loss of native immunity during the transplant process and immune reconstitution with a cmv naïve allograft [ , ] . patients who are cmv d+/r+ have cmv reactivation ( - %) and disease risk that is similar to or slightly lower than that of the cmv d−/r+ patient. patients who are cmv d+/ r− have a lower risk of cmv infection ( - %) and disease, but higher than cmv d−/r− patients (< %). the risk of cmv infection in this latter group has been greatly decreased by use of leukoreduced blood products or by exclusive use of cmv negative products when available [ ] . the risks of cmv reactivation and disease among autologous hsct recipients is minimal (< %) [ , ] . another major risk factor for the development of cmv reactivation and disease is the occurrence, severity and treatment of acute gvhd [ , ] . other potential factors associated with an increased risk of cmv reactivation and disease are reception of t cell-depleted or cord blood allograft, whether the donor is unrelated or mismatched, or donated bone marrow (instead of peripheral stem cells), and whether the conditioning regimen was myeloablative [ , ] . the most widely used therapy for clinical cmv disease is ganciclovir, which can be administered either intravenously or orally in the form of a prodrug, valganciclovir, with an acceptable bioavailability profile (~ - %). typically, the parenteral form is administered until the patient is able to tolerate oral therapy. gastrointestinal absorption of valganciclovir, even in the setting of mild to moderate gi gvhd, has been demonstrated to be adequate [ , ] . duration of treatment depends on the clinical response and the nature of the recovery of native immune function. in the case of serious illness, particularly pneumonia, anti-cmv hyperimmune globulin can be considered as adjunctive therapy. despite these efforts, the mortality from cmv pneumonia remains high. the major toxicity of ganciclovir is myelosuppression, so that great effort is placed in monitoring these patients closely and adjusting doses appropriately [ , ] . occasionally, g-csf support may be required to preserve an acceptable neutrophil count and to allow adequate therapy of a serious cmv infection. while certain medications, such as atg and okt , are likely to induce cmv reactivation, others like sirolimus may inhibit this [ ] . current strategies are based on preventing cmv disease through prophylaxis or preemption. prophylaxis with ganciclovir from the time of engraftment until at least day post-transplant has been studied in randomized trials [ , ] . although cmv viremia and disease were prevented, there was no overall benefit of this strategy due to secondary bacterial and fungal infections related to ganciclovir-induced neutropenia. alternatively, a preemptive strategy is employed in which patients are monitored weekly for viremia through either a pcr assay for cmv dna or an antigenemia assay. positive results are linked to initiating ganciclovir or other antiviral drugs active against cmv. typically, these assays turn positive several days to weeks prior to the onset of clinical disease, permitting the use of effective preemptive therapy [ , , , , [ ] [ ] [ ] [ ] [ ] . a preemptive approach significantly decreases the amount of prophylactic medication used, thus minimizing medication-associated toxicity. in the pre-ganciclovir era, cmv disease typically occurred during the first three months post-transplant. increasingly, with the widespread use of a prophylactic or preemptive antiviral strategy, breakthrough occurs much later, typically one to three months after the cessation of the antiviral therapy. risk factors for late cmv disease include chronic gvhd, low cd -t cell counts, and cmv infection before day . relapse or the emergence of ganciclovirresistant virus also can occur, particularly in the face of high viral loads and inadequate courses or dosing of ganciclovir. foscarnet is the preferred drug in this setting or when further potential myelosuppression with ganciclovir is not advisable. the experience with cidofovir use in the hsct population is limited. both foscarnet and cidofovir are potentially nephrotoxic and should be administered with caution [ , ] . studies are examining the emerging strategies for the management of cmv infection and the use of cmv vaccines in donors and recipients, adoptive immunotherapy for patients with refractory or relapsing cmv infection and the use of maribavir for prophylaxis. the major recognizable clinical effect of ebv in the hsct patient is in the pathogenesis of ptld. following the recovery from primary ebv infection (> % of the adult population), ongoing lytic infection of b-cells occurs in the oropharynx, with latent infection of b-cells in the peripheral blood and lymphoid tissues. these latently infected cells can be transformed and immortalized, resulting in polyclonal proliferation. in the normal seropositive individual, these cells are kept in check by a specific cytotoxic t cell response. in the presence of immunosuppressive therapy, this surveillance system is inhibited in a dose-related fashion, thus permitting continued b-cell proliferation. such ongoing proliferation results in particular clones being favored and the potential for developing cytogenetic abnormalities, which leads to the development of a truly malignant process-ptld [ , , , ] . the spectrum of clinical disease seen with ptld is quite broad, ranging from a mononucleosis-like process or a polyclonal proliferation of lymphocytes that usually responds to decreasing immunosuppressive therapy, to a monoclonal, highly malignant b-cell lymphoma. the mononucleosis-like process is seen particularly in children with primary post-transplant ebv infection. the clinical presentation is one of fever, sore throat, cervical adenopathy and tonsillar hypertrophy and inflammation. unlike b-cell lymphoma in the normal host, in the transplant patient, particularly the adult, the process can be extranodal. thus, presentations may include central nervous system (cns) invasion (from involvement of the meninges to focal cerebral lesions), liver, lung and bone marrow diseases. not uncommonly, involvement of the gut (particularly the small bowel) may lead to recognition of the ptld, with a clinical presentation of small bowel obstruction, perforation, or occult gastrointestinal bleeding. disseminated, multi-organ disease is quite common in the hsct patient [ , , , ] . risk factors for developing ptld include: primary ebv infection in association with high-dose immunosuppression; interventions such as t cell depletion, umbilical cord blood transplant and the systemic administration of anti-thymocyte globulin increase the risk significantly; and intensive immunosuppression that results in suppression of the key host defense against ebv-transformed cells (mhc-restricted, ebv-specific, cytotoxic t cells) significantly increases the risk of ptld. in addition to the host characteristics mentioned, high ebv viral loads correlate with an increased risk of ptld. it has been suggested that ebv viral load surveillance in peripheral blood be carried out in high risk patients (those with primary ebv infection, anti-t cell antibody therapy for gvhd, hla-mismatched or t cell-depleted hsct recipients), with decreased immunosuppression +/− antiviral therapy (acyclovir or ganciclovir) carried out in the setting of high viral loads [ , , , ] . treatment of ptld remains controversial. all patients with diagnosed ptld should have a significant decrease in immunosuppressive medications. many centers also prescribe antiviral therapy. patients not responding to these measures are usually treated with an anti-b-cell monoclonal antibody (rituximab, an anti-cd monoclonal antibody) [ , ] . after that, therapies have ranged from anti-lymphoma chemotherapy to alpha-interferon and intravenous gamma globulin. hsv infection prior to the introduction of acyclovir was a major problem in the hsct recipient. occurring in the preengraftment period, hsv infection greatly exacerbated the severity of mucositis. not only were ulcers observed in the oral cavity and anogenital areas, ulcerations of the esophagus, stomach and intestine were also observed. hsv pneumonia was also noted, with rare cases of cutaneous dissemination and encephalitis. the current standard of care is to test all candidates for hsct for an antibody to hsv, with seropositive individuals then placed on antiviral prophylaxis, beginning prior to hsct. effective agents for hsv prophylaxis include acyclovir (intravenous or oral), valacyclovir or famciclovir. recurrence of hsv may occur later in the course, and should again be treated with an acyclovir regimen, with repeated episodes justifying long-term prophylaxis. acyclovir resistance is uncommon in this situation, but can occur, and requires treatment with foscarnet [ , ] . all patients and donors should have serologic testing for vzv prior to transplant. seronegative individuals post-transplant should avoid exposures to vzv, but if such an exposure occurs, valacyclovir or varicella hyperimmune globulin should be promptly initiated. before universal prophylaxis with acyclovir became standard, an estimated percent of hsct patients developed active vzv, with a median time of onset being five months post-transplant. the great majority of these patients had zoster, but approximately percent had a more generalized process resembling primary varicella. a significant concern was visceral involvement in the setting of disseminated disease as well as neurologic complications such as myelitis or encephalitis [ , [ ] [ ] [ ] [ ] . prophylaxis with acyclovir in the early period post-transplantation substantially decreases the occurrence of herpesvirus infections, including vzv, and is rarely, if ever, seen during acyclovir prophylaxis. prophylaxis is typically given for the first year post-allogeniec transplantation. vzv reactivation is often seen three months post-discontinuation of prophylaxis. as the vzv vaccine is a live attenuated viral vaccine, its use is contraindicated for at least two years posttransplantation, and unless a research study or close follow-up is involved, should be omitted. hhv- is a β-herpesvirus (as is cmv) whose role in post-transplant complications is being defined. in the great majority of instances, hhv- primary infection occurs by the third year of life, with a seroprevalence rate of percent at one year, and close to percent at three years [ , ] . the clinical effects associated with primary hhv- infection include exanthem subitum (roseola), and a form of encephalitis. in hsct patients, bone marrow suppression, especially delayed platelet engraftment, and encephalitis have been associated with hhv- type b. the encephalitis typically occurs one to two months after transplantation and is associated with profound memory loss, especially short-term memory, and mri changes in the mesial temporal lobes (limbic encephalitis) [ , ] . the highest risk patients for this complication are male, umbilical cord blood recipients for whom the attack rate may be as high as to percent. detecting hhv- dna in the blood of allogeneic hsct recipients is a common phenomenon occurring transiently in to percent of patients, yet encephalitis is a rather infrequent occurrence ( - %). as obtaining brain biopsies is not usually feasible early after transplantation, the diagnosis of hhv- encephalitis is currently achieved by developing an acute limbic encephalitis syndrome, confirmed with mri imaging of the brain and by the detection of hhv- in the csf [ ] . it remains unclear what the treatment of choice for this virus is. one approach that we currently favor is to use foscarnet. it is possible that anti-cmv preventative strategies with ganciclovir may have a beneficial effect on this virus as well [ , ] . hsct recipients are at significant risk for infection with respiratory viruses circulating in the community. these infections can occur at any time in the post-transplant course, and can be acquired in the community or during hospitalization from infected staff, family and friends. overall, an estimated to percent of hsct patients will become infected in the first year posttransplant, with the potential for this figure to rise significantly in the setting of a community-wide outbreak [ ] . the dilemma for the clinician is how to prevent these infections, as there is a far higher rate of progression to pneumonia (viral and/or bacterial or fungal superinfection), which carries a far higher morbidity and mortality than what is observed in the general population. in addition, antiviral therapy for these agents is in its infancy. it is important to attempt to make an etiologic diagnosis. avoiding exposure to infected individuals by systematic infection control measures in both family members and friends, but most importantly in health care workers, is the best preventative strategy available [ ] [ ] [ ] [ ] . although rsv can be acquired by inhaling an aerosol, direct contact with infected secretions is the usual mode of spread between individuals. in the hsct patient, both adult and pediatric, rsv is a cause of significant morbidity and mortality. the illness begins with the signs and symptoms of a viral upper respiratory tract infection (rhinorrhea, sinus congestion, sore throat and/or otitis media), that may progress to pneumonia, especially if the virus is acquired in the preengraftment phase. as with influenza, pneumonic syndromes can be due to rsv itself, but in our experience it is more frequently due to secondary bacterial and fungal infections. the advent of rapid rsv diagnosis by antigen detection in nasopharyngeal swabs has resulted in the recognition that rsv is a significant pathogen for both adults and children, particularly in immunosuppressed patients. optimal antiviral management, however, remains unclear. there are reports that aerosolized ribavirin +/− anti-rsv polyclonal or monoclonal antibody may have therapeutic benefit, but this remains unproven. there is also interest in prophylaxis with an anti-rsv antibody, although there have been no trials in hsct patients [ ] [ ] [ ] [ ] [ ] . as with rsv, the incidence of influenza infection in hsct patients reflects the level of influenza activity in the community. the impact of this virus on infected hsct recipients is demonstrated by the following statistics: ~ percent of the patients with influenza develop pneumonia and ~ percent of patients with influenza pneumonia die of progressive respiratory failure. when influenza is identified as a pathogen, use of a neuraminidase inhibitor (oseltamvir or zanamavir) or an amantadate (amantadine or rimanatine) should be considered. the neuraminidase inhibitors are attractive in this setting as they are effective against both influenza a and b and antiviral resistance occurs more slowly compared with amantadine use. annual influenza vaccination should be considered, but its benefit is attenuated; indeed, it is probably fair to say that maximal benefit from vaccination occurs when the vaccine is administered to health care workers, family, friends and other contacts of the patient. when an infection is diagnosed, early treatment should be considered [ , ] . there are more than serotypes of adenovirus and nearly all have been described to cause human disease. adenovirus disease post-transplantation is likely due to both a newly acquired virus and viral reactivation. the most common adenovirus-associated illness post-transplantation is hemorrhagic cystitis which has been described in a recent report to occur in up to percent of patients in the first year post-transplantation [ ] . the overwhelming majority of cases are asymptomatic and require no intervention [ ] . occasionally the severity of hemorrhage or bladder-associated pain is so great that intervention is required. other important adenovirus-associated syndromes include hepatitis and pneumonitis which may be fatal in the early post-transplant period. in the late post-transplant period adenovirus gastroenteritis may occur which is often a self-limited illness; however, severe disease has been described especially in patients requiring significant levels of immunosuppression for gvhd. therapeutic options for adenovirus are limited. the role of the antiviral cidofovir is controversial with mixed results having been reported [ ] . decreasing immunusuppression and attempting reconstitution of the native host immune response is critical. the role for other adjunctive therapies, such as ivig, is unproven, but can be considered in severe cases. avoiding exposure to new infection, as with all community-acquired pathogens, is central to optimal care. parainfluenza, rhinoviruses, metapneumovirus and coronaviruses are all capable of causing lower respiratory tract infection in hsct recipients. of these many viruses, parainfluenza virus type iii is especially associated with a high mortality [ , ] . again, specific therapy is not available, emphasizing infection control strategies in the hospital setting and avoiding individuals with respiratory tract complaints at home. when upper respiratory tract complaints occur in hsct patients, a diagnosis should be made, utilizing rapid diagnostic techniques (e.g., antigen detection assays or nucleic acid testing). preemptive therapy, when available, should be initiated, while immunosuppressive therapy diminished and isolation from other hsct patients should be accomplished. bk and jc viruses are the two important species in this family of viruses with a genitourinary and cns predilection, respectively. approximately to percent of adults have been infected with one or both of these viruses, typically in childhood. with immunosuppression, reactivation occurs which may lead to disease. bk virus is associated with hemorrhagic cystitis in the early post-transplant period. this virus is commonly found in the urine and rarely requires any therapeutic intervention. jc virus is the etiologic agent of progressive multifocal leukoencephalopathy (pml) which is a rare, but severe post-transplant complication. pml involves the white matter and presents with focal neurologic symptoms associated with the specific area of the cns where the lesion(s) occur. diagnosis requires correlating the clinical presentation, radiographic findings (typically by contrast-enhanced mri imaging) and csf pcr results for jc virus. control of jc virus is associated with an intact cell-mediated immune response. therapy for polyomavirus infection is quite limited with minimizing immunsuppression, when possible, being critical. the role of cidofovir is controversial with mixed results being reported. the use of quinolones for bk viruria is controversial at best and we do not recommend this practice [ ] . although the use of gatifloxacin was advocated by some authors, this drug is no longer available. leflunomide administration has been used by some for treatment of bk in renal transplant patients, but no randomized trial data exists to support or recommend its use in either kidney or hsct recipients at this time. hepatitis b and c viruses may cause chronic infection which often leads to eventual significant liver dysfunction. given the high global prevalence of these viruses, it is prudent to screen for past or current infection prior to transplantation. when ongoing infection is found, careful assessment of liver function and a pre-transplant liver biopsy should be considered to assess for occult cirrhosis, as this may influence peri-transplant management [ ] . hbv infects approximately million people worldwide chronically, and substantially more have had prior resolved infection. the use of the hbv vaccine as a routine childhood immunization will likely decrease the number of chronically infected individuals over the next several decades. the advent of nucleic acid detection technology has allowed a more precise mechanism to detect active hbv replication compared with antigen-(for surface and e) only methods. for patients with evidence of prior hbv exposure (hbv core antibody positive), it is important to consider hbv reactivation in the setting of post-transplant liver dysfunction and to differentiate this from other causes such as hepatic gvhd or medication toxicity, although reactivation initially occurs in the setting of normal liver tests. the best strategy for surveillance post-transplantation remains to be defined. some recommend routine surveillance for hbv reactivation post-transplantation, whereas others would suggest antiviral prophylaxis. it is important to be aware that old resolved infections, including those with hepatitis core and surface antibody, but without antigen or hbv dna detected, are at risk for reactivation (seroreversion) post-transplantation, especially in the setting of high levels of immunosuppression [ , ] . several therapeutic options have become available over the last several years and include lamivudine [ ] [ ] [ ] , adefovir [ ] , entecavir [ , ] and telbivudine. other agents such as tenofovir and emtricitabine also have excellent anti-hbv activity. use of these agents requires careful consideration to minimize the risk for the emergence of resistant virus, which may be as high as percent per year for lamivudine, but is less than percent for adefovir and entecavir. epidemiologic studies suggest that more than million people worldwide have been infected with hepatitis c virus (hcv) and the majority (approximately %) are chronically infected [ ] . over several decades, chronic hcv infection is associated with progressive hepatic fibrosis, liver failure, and hepatoma. this process is accelerated in certain immunocompromised patients including hsct recipients [ , ] . it is important to assess patients for seropositivity to hcv prior to transplantation and in those who are found seropositive, to assess the hcv viral load, genotype and liver pathology. the presence of elevated liver enzymes in the setting of hcv before allogeneic hsct has been associated with an increased incidence of vod [ ] . a more precise profiling of the hcv-infected patient, including liver biopsy, should be considered to better define the extent of the hcv-induced liver disease, and to optimize the conditioning regimen and frequency of surveillance posttransplantation. treatment of hcv is limited and typically requires use of an interferon and ribavirin which are likely to be poorly tolerated in the early post-transplant setting. in patients who are infected, it is prudent to counsel them to avoid hepatotoxins, receive the hepatitis a and b vaccines, and minimize the risk of transmission to close contacts. there are three categories of fungal pathogens that can infect the hsct patient: a) the classic opportunistic fungi, which cause > percent of the invasive fungal infections that occur in the hsct patient -candida, aspergillus and cryptococcus being the most important of these infections; b) the geographically restricted systemic mycoses caused by blastomyces dermatitidis, coccidioides immitis and histoplasma capsulatum; and c) invasive infection due to the so-called "newly emerging fungi" -fusarium, the zygomycetes and such dematiaceous fungi as scedosporium, scopulariopsis and dactylaria [ ] . candida is a major cause of fungal bloodstream infection during the preengraftment phase of hsct. although there is the possibility that the portal of entry can be vascular access catheters, it is believed that translocation of candida species across gut mucosa damaged by the pre-transplant conditioning regimen is the major route of access to the bloodstream in the granulocytopenic patient [ , ] . in the past, c. albicans and c. tropicalis accounted for virtually all of the candida bloodstream infections. the incidence of candidemia was ∼ to percent (with a median time to onset of two weeks post-transplant), resulting in a high rate of tissue invasion and an attributable mortality of nearly percent [ , ] . with the introduction of empirical antifungal therapy or fluconazole prophylaxis ( mg/day) during the preengraftment period, the incidence of candidemia has been significantly decreased, hepatosplenic candidiasis has become quite rare, and the attributable mortality has been significantly decreased. fluconazole-resistant candida sp, c. krusei and c. glabrata, have emerged as not uncommon causes of candidemia in hsct patients, as have the other non-albicans candida species [ , , [ ] [ ] [ ] [ ] [ ] [ ] [ ] . it is also important to recognize that other species of yeast (e.g., trichosporon sp, blastoschyzomyces capitatus, saccharomyces cereviseae and rhodotorula sp.) can cause clinical syndromes identical to those observed with invasive candidiasis (bloodstream infection, infection metastatic to the skin and subcutaneous tissues, as well as other sites, including hepatosplenic disease identical to that caused by candida species) [ ] . in an era of increased use of echinocandins for prophylaxis [ ] and empirical antifungal treatment [ ] , these organisms [ , ] and echinocandin-resistant candida sp, especially c. parapsilopsis, have become emerging causes of fungemia in the hsct units [ , , ] . invasive fungal disease has been most commonly caused by aspergillus sp, with a. fumigatus, a. flavus, a. terreus, a. niger and a. nidulans being the most common causes of invasive aspergillosis. the portal of entry for percent of cases of invasive aspergillosis is the lungs, with the nasal sinuses and the skin accounting for virtually all of the remaining cases. there are two major host defenses that are mobilized in response to inhalation of the aspergillus spores -granulocytes and cell-mediated immunity, specifically cytotoxic t cells. the importance of these mechanisms is demonstrated by the clustering of cases of invasive aspergillosis at two timepoints in the posttransplant course: preengraftment when profound granulocytopenia is present, with the incidence of invasive aspergillosis increasing steadily as the period of granulocytopenia is extended, and after the diagnosis of gvhd and the treatment of this adverse event. indeed, these late cases of invasive aspergillosis have become more common than the preengraftment cases. mortality rates have traditionally been high in patients who developed invasive aspergillosis in either time period [ , , , ] . the clinical syndromes caused by aspergillus invasion reflect the pathologic consequences of the vasculotropic nature of this mold. the three major consequences of the vascular invasion that characterizes aspergillus invasion include hemorrhage, infarction and metastatic disease. initial clinical complaints include persistent fever, chest pain, tachypnea, hypoxemia and hemoptysis, as well as symptoms related to metastases. before the availability of noninvasive fungal markers (galactomannan and β-glucan) and aggressive imaging with spiral chest computerized tomographic (ct) scanning, percent or more of patients experience disseminated infection at the time of first diagnosis, accounting for the high mortality observed in allogeneic hsct recipients. a particular problem is infection in the cns, where mortality historically has approached percent. metastases can present any site, but particularly important is the skin, as innocent appearing skin lesions can lead to early recognition of the disease, and should be aggressively biopsied [ ] . definitive diagnosis of invasive mold infections, including invasive aspergillosis, is usually accomplished by biopsying the site of abnormality. early diagnosis is the key to effective therapy [ ] . sputum or bronchoscopic samples rarely yield mold on culture. in recent years, considerable effort has been made to find other technology that will lead to an earlier and timely diagnosis. the ones that have been incorporated into practice are the systematic measurement of aspergillus antigens and serial chest ct imaging. monitoring the serum of hsct patients for galactomannan or β-glucan is now commercially available and has been incorporated into the current diagnosis guidelines [ , ] . the detection of circulating fungal dna in the blood by pcr [ ] remains experimental. findings on chest ct, in particular the halo sign ( fig. - ) , are associated in the neutropenic patient with invasive aspergillosis (although other pathogens can cause the same radiologic finding: fusarium and other vasculotrophic molds and nocardia asteroides being examples of this). european groups have been advocating protocol serial chest ct scans to find such pathology as a guide to early diagnosis [ ] . if prevention fails, then early diagnosis is the key to the patient's survival [ , , ] . given the limitations of current diagnostic techniques and the significant morbidity associated with invasive fungal infection, two strategies of antimicrobial use are commonly deployed in the hsct patient. the first is prophylactic fluconazole use during the initial transplant period, which has been shown to decrease fungal infections [ ] in one study, and overall mortality fig. - . computerized tomographic scan of the chest in a patient with a "halo sign" due to invasive aspergillosis. note that halo signs most commonly occur in granulocytopenic hsct recipients with invasive aspergillosis. however, it must be emphasized that a halo sign is occasionally seen in patients with nocardia, scedosporium, fusarium and other forms of pneumonia. the patient was treated with voriconazole monotherapy during neutropenia during consolidation chemotherapy for aml and his treatment was continued through allogeneic transplantation. [ ] in another, when started on day until engraftment [ ] or day + [ ] . it is important to note that a high background rate of candida infections was noted in both of these reports and may not represent the experience of other transplant centers. echinocandins may be an alternative to fluconazole prophylaxis during this risk period [ ] . the second common strategy is empiric antifungal therapy in neutropenic patients with persistent fever without a source, despite broad-spectrum antimicrobial therapy for > hours [ , ] . in this setting the primary concern is both candida and invasive mold infection, especially aspergillus [ ] . the traditional antifungal therapy utilized as empiric therapy is an amphotericin product [ , ] . caspofungin use in this setting has become common because of the favorable side effect profile of this class of agents, but at the expense of a more limited fungal spectrum [ ] . other echinocandins (micafungin, anidulafungin) are likely to be similarly effective, but no randomized comparisons with these latest drugs have been performed. the role of voriconazole in this setting is controversial [ ] . when treating invasive aspergillosis several approaches should be considered simultaneously: ) antifungal therapy, ) reverse or minimize the host immune defects (decrease corticosteroids, increase neutrophils), ) control permissive viral infections (e.g., cmv) and ) consider surgical excision, if possible. voriconazole has become a cornerstone of therapy for invasive aspergillus infections, though the management of potential side effects is substantial [ ] [ ] [ ] . whether the combination of therapeutic agents (polyenes, azoles and echinocandins) increases the therapeutic benefit has yet to be determined [ ] . increasing experience suggests that voriconazole alone is sufficient in most cases for a successful outcome in invasive aspergillosis and has decreased the morbidity and mortality of this infection [ , ] . another significant risk period for invasive fungal infections (ifi) is in the setting of significant gvhd, such as grade iii or iv, and its therapy [ ] . in this setting, posaconazole (versus fluconazole) prophylaxis has recently demonstrated some benefit in preventing ifi compared to fluconazole ( . % versus . %, p = . ) and in preventing probable or proven invasive aspergillosis ( . % versus . %, p = . -interestingly, these results were largely driven by results from galactomannan assay testing) [ ] [ ] [ ] . posaconazole has activity against the zygomycetes as well as aspergillus sp [ ] [ ] [ ] . when an azole is used in this patient population, careful assessment of drug interactions, both with the initiation and cessation of therapy, is critical. therapy for the emerging fungi fusarium and scedosporium should be guided by in vitro sensitivity testing done locally or at regional reference laboratories, but voriconazole use should be considered. when therapy for the endemic mycoses is indicated, initial treatment (induction therapy) with an amphotericin preparation should be considered, followed by a prolonged course of consolidative therapy with an oral azole. cryptococcal disease should be treated initially with an amphotericin preparation, cns involvement should be excluded by cerebrospinal fluid sampling, and the use of flucytosine should be considered if present. pneumocystis jiroveci, formerly carinii, is a ubiquitous environmental organism which is an important cause of pneumonia in patients who are immunosuppressed, such as those who have undergone an hsct, on chronic prednisone (typically > mg per day) or with advanced hiv infection. pcp infection typically presents as an interstitial pneumonitis with marked hypoxemia. severe infection can be life threatening. fortunately, universal prophylaxis of high risk patients during the high risk periods with trimethoprim-sulfamethoxazole has markedly decreased this complication. however, intolerance to prophylaxis, use of second line prophylaxis agents (e.g., dapsone, pentamidine, or atovaquone), poor medication compliance or failure to re-institute prophylaxis in the setting of augmented immunosuppression (e.g., treatment of gvhd) are common reasons why cases still occur. several important issues must be addressed after successful hsct to minimize infectious complications. first, it is important to avoid exposure to pathogens, especially when the immunosuppressive therapy to prevent gvhd is the highest. this includes avoiding gardening and soil exposures, mold exposures such as cleaning out damp basements or smoking marijuana, individuals with active respiratory infections, especially children, and avoiding enteric pathogens. second, optimal treatment or monitoring for latent infections such as herpesviruses, hepatitis viruses and prior granulomatous diseases (e.g., mycobacterium tuberculosis). those patients with a positive test for latent tuberculosis should receive secondary prophylaxis, which typically is begun within one month post-transplantation, after the acute regimen toxicities associated with transplantation have subsided, when screening and preventive treatment have not occurred previous to hsct. the first line therapy for secondary prophylaxis is isoniazid for nine months. however, in patients with significant hepatic dysfunction or peripheral neuropathy alternative regimens need to be considered. rifamycin-based regimens are difficult given the potential hepatotoxicity, as well as the significant drug interactions, especially with concomitant use of a calcineurin or an azole. a quinolone, such as levofloxacin, with ethambutol may be considered. when a mycobateriologically static regimen is chosen, the duration of therapy often must be extended with some using this combination for months as secondary prophylaxis (table - ) . third, optimizing vaccinations for routine pathogens such as diphtheria, tetanus, pertussis, influenza and pneumococcus (table - ) . this optimal timing of re-vaccination depends on the nature of the transplant, with earlier re-vaccination schedules being considered in the nonmyeloablative setting. fourth, prophylaxis for pcp, which is typically continued for approximately one year or until the immunosuppressive medications are tapered off. the optimal medication to use for pcp prophylaxis is trimethoprimsulfamethoxazole which offers some protection for a variety of other important pathogens including pneumococcus, hemophillus influenza, nocardia sp., toxoplasma, listeria, salmonella sp., and other enteric bacterial pathogens. if trimethoprim-sulfamethoxazole is not tolerated due to significant renal dysfunction or bone marrow suppression, then alternative agents for pcp prophylaxis include dapsone, atovaquone or aerosolized pentamidine; however, none of these second line agents afford the broad microbial protection which trimethoprim-sulfamethoxazole affords. and lastly, herpes group viral prevention which should include acyclovir to prevent hsv and vzv and systematic monitoring for cmv, in the allogeneic setting, with early use of a cmv active antiviral if evidence for cmv activation or disease is observed. prophylaxis must be re-assessed in the setting of persistent or augmented immunosuppression, such as in the setting of clinically significant gvhd, regardless of time since hsct. medication doses may need to be adjusted for renal dysfunction. * trimethoprim-sulfamethoxazole affords modest protection for a broad array of potential environmental and community pathogens including: nocardia sp, toxoplasmosis, pneumococci, h influenza, listeria, shigella, and slamonella sp. ** alternatively preemptive monitoring with serial viral antigen or viral load assays can be considered. *** for those with evidence of prior hbv infection (e.g., hepatitis b core antibody positive), consider monitoring hbv viral load for evidence of reactivation periodically. if reactivation is detected then consider treatment if persistent hbv viremia detected. specific hbv antiviral therapy is discussed in the text. **** pre-transplant secondary prophylaxis for mtb is preferred. ***** decision for systemic azole prophylaxis should be based on local epidemiology of invasive fungal infections. consider posaconazole prophylaxis in the setting of significant gvhd (e.g., grade or ) and its therapy. drug interactions must be carefully managed both with initiation and cessation of azole therapy. an important aspect of antimicrobial therapy in the hsct patient is the management of drug interactions, especially between antimicrobial agents (e.g., azoles, macrolides) and the immunosuppressive medications (e.g., calcineurin inhibitors, sirolimus) used to prevent and treat gvhd. there are three important categories of interaction to pay particular attention to, two of which are related to the major route of drug metabolism for the calcineurin inhibitors, hepatic cytochrome p enzymatic metabolism. these interactions are as follows: ) certain antimicrobial agents (most notably the macrolides [eryth romycin>clarithromycin>azithromycin] and the azoles [ketoconazole>itraco nazole>voriconazole>fluconazole]) will downregulate the metabolism of the calcineurin inhibitors, resulting in elevated blood levels of active drug, and an increased risk of nephrotoxicity, as well as over-immunosuppression and an increased incidence of opportunistic infection; ) certain antimicrobial agents (such as rifampin and rifabutin) upregulate metabolism of the calcineurin inhibitors, leading to a fall in blood levels and an increased risk of gvhd, and ) therapeutic blood levels of the calcineurin inhibitors, when combined with such drugs as amphotericin b, aminoglycosides and vancomycin, can cause significant renal toxicity. hsct has become one of the great success stories of modern medicine. it is the therapy of choice for an increasing number of conditions, including a variety of cancers, bone marrow failure states, congenital immunodeficiencies, metabolic disorders and even as a means for introducing new genes. the major hurdle in most of these attempts, however, remains infection. bacterial and fungal sepsis, as well as herpes group viral infection and community-acquired respiratory virus infection threaten the well-being of these patients. there are two phases of the post-transplant course when the patient is at particular risk: preengraftment with profound granulocytopenia and mucositis, and post-engraftment when gvhd and its therapy render the patient vulnerable to both fungal and viral infection. new preventative strategies are being formulated involving both prophylaxis and preemptive therapy. similarly, new non-culture diagnostic approaches that rely on antigen detection or pcr detection of microbial dna are being developed. new therapies, both antiviral and antifungal, have emerged. these should prompt much more effective prevention and therapeutic strategies. infection in hematopoietic stem cell 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patients receiving pcr-based preemptive therapy for cmv infection preemptive therapy in immunocompromised hosts epstein-barr virus and lymphoproliferatic disorders after transplantation cd monoclonal antibody (rituximab) for therapy of epstein-barr virus lymphoma after hemopoietic stem-cell transplantation anti-b-cell monoclonal antibodies in the treatment of severe b-cell lymphoproliferative syndrome following bone marrow and organ transplantation oral acyclovir for the long-term suppression of varcella zoster virus infection after marrow transplantation the effectiveness of the varicella vaccine in clinical practice postexposure prophylaxis of varicella in family contact by oral acyclovir varicella-zoster virus infection in adult patients after unrelated cord blood transplantation: a single institute experience in japan clinical outcomes of human herpesvirus reactivation after hematopoietic stem cell transplantation a population-based study of primary human herpesvirus infection mr 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infections after hematopoietic stem cell transplantation: risk factors, mortality, and the effect of antiviral therapy hemorrhagic cystitis in adults after unrelated cord blood transplantation: a single-institution experience in japan assessment of disseminated adenovirus infections using quantitative plasma pcr in adult allogeneic stem cell transplant recipients receiving reduced intensity or myeloablative conditioning cidofovir for adenovirus infections after allogeneic hematopoietic stem cell transplantation: a survey by the infectious diseases working party of the european group for blood and marrow transplantation parainfluenza virus infections after hematopoietic stem cell transplantation: risk factors, response to antiviral therapy, and effect on transplant outcome respiratory disease due to parainfluenza virus in adult bone marrow transplant recipients ciprofloxacin decreased polyoma bk virus load in patients who underwent allogeneic hematopoietic stem cell transplantation 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and marrow transplantation hepatitis c virus infection is a risk factor for liver failure from veno-occlusive disease after bone marrow transplantation guidelines for treatment of candidiasis should vascular catheters be removed from all patients with candidemia? an evidence-based review efficacy and safety of fluconazole prophylaxis for fungal infections after marrow transplantation-a prospective, randomized, double-blind study a controlled trial of fluconazole to prevent fungal infections in patients undergoing bone marrow transplantation infections in the neutropenic patient-new views of an old problem. hematology / the education program of the fungal infections in granulocytopenic patients: current approaches to classifications, diagnosis candidemia in immunocompromised patients infections due to resistant candida species in patients with cancer who are receiving chemotherapy prospective study of candida endophthalmitis in hospitalized patients with candidemia risk factors for hepatosplenic abscesses in patients with acute leukemia receiving empiric azole treatment practice guidelines for the treatment of candidiasis. infectious diseases society of america disseminated trichosporonosis caused by trichosporon loubieri micafungin versus fluconazole for prophylaxis against invasive fungal infections during neutropenia in patients undergoing hematopoietic stem cell transplantation caspofungin versus liposomal amphotericin b for empirical antifungal therapy in patients with persistent fever and neutropenia breakthrough trichosporonosis in patients with hematologic malignancies receiving micafungin epidemiology of aspergillus infections in a large cohort of patients undergoing bone marrow transplantation candidemia in allogeneic blood and marrow transplant recipients: evolution of risk factors after the adoption of prophylactic fluconazole prevention and early treatment of invasive fungal infection in patients with cancer and neutropenia and in stem cell transplant recipients in the era of newer broad-spectrum antifungal agents and diagnostic adjuncts defining opportunistic invasive fungal infections in immunocompromised patients with cancer and hematopoietic stem cell transplants: an international consensus should the consensus guidelines' specific criteria for the diagnosis of invasive fungal infection be changed? reactivity of ( -> )-beta-d-glucan assay with commonly used intravenous antimicrobials increasing volume and changing characteristics of invasive pulmonary aspergillosis on sequential thoracic computed tomography scans in patients with neutropenia empiric antibiotic and antifungal therapy for cancer patients with prolonged fever and granulocytopenia empiric antifungal therapy in febrile granulocytopenic patients. eortc international antimicrobial therapy cooperative group liposomal amphotericin b for empirical therapy in patients with persistent fever and neutropenia. national institute of allergy and infectious diseases 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conventional therapy: an externally controlled trial we would like to thank robert h rubin, m.d. for the many years of advice and teaching, and for critically reviewing this work. key: cord- -tylkbh h authors: chemaly, roy f.; rathod, dhanesh b.; couch, robert title: respiratory viruses date: - - journal: principles and practice of cancer infectious diseases doi: . / - - - - _ sha: doc_id: cord_uid: tylkbh h the respiratory viruses as a group are the most common cause of an acute infectious illness in developed societies. the immunocompromised state of many cancer patients constitutes the basis for the frequent failure of the host to promote a normal and rapid recovery from an acute respiratory viral infection and results in a more severe and prolonged infection that causes significant morbidity and mortality in these patients. those respiratory viruses that are most prevalent and most prone to produce lower respiratory illnesses and pneumonia in healthy hosts, rsv, influenza viruses, and parainfluenza viruses, are those most likely to cause severe illness and pneumonia leading to hospitalization in immunocompromised persons. however, viruses less prone to produce a lower respiratory illness but that are highly prevalent, such as rhinoviruses, may frequently be associated with severe illness. the limited availability of antivirals and vaccines for the acute respiratory viruses means that these infections will continue to be important for many years and dictate a need for utilizing infection control procedures as much as possible, particularly in hospitals and institutions, so as to minimize spread. efforts to develop specific vaccines are important as their use could prevent as well as reduce exposure of cancer patients to these viruses. development of specific antivirals is important for use in immunocompromised patients as normal recovery mechanisms may be seriously impaired. community respiratory virus infections were once primarily considered to be infections of children and generally nonserious. however, over the past two decades, it has become clear that these viruses can cause serious infections that require medical attention, particularly in infant, elderly, and immunocompromised patients. historically, the most common causes of respiratory infections in cancer patients were thought to be opportunistic bacteria and fungi, but newer diagnostic methods have revealed that respiratory viruses can cause serious morbidity and mortality in such patients, including leukemia patients and hematopoietic stem cell transplant (hsct) recipients. many viruses are known to cause respiratory tract infections, but the most common in hospitalized cancer patients are influenza viruses, respiratory syncytial virus (rsv), and parainfluenza viruses (piv) [ , ] . however, all respiratory viruses can cause respiratory infections in cancer patients including rhinoviruses, enteroviruses, coronaviruses, human metapneumoviruses (hmpvs), adenoviruses, as well as cytomegaloviruses, herpes simplex, and varicella zoster viruses. in this chapter, we discuss the common clinical presentations, diagnostic methods, treatments, and prevention measures for respiratory virus infections in general and in cancer patients in particular (table . of these cases, most were in hsct recipients ( %). the summer [ ] [ ] [ ] . rsv is a member of the paramyxoviridae family and the genus pneumovirus. it is an enveloped singlestranded rna virus approximately - nm in diameter that is contained in a helical nucleocapsid surrounded by a lipid envelope. this envelope bears two glycoproteins: the g protein, which attaches to the cell surface, and the f (fusion) protein, which enables the internal components to enter the cell after fusion of host and viral membranes and to initiate replication. both glycoproteins are integral to pathogenesis [ , , ] . there are two groups of rsv viruses, groups a and b [ ] [ ] [ ] [ ] . rsv infections can present with a wide array of upper or lower respiratory symptoms. the incubation period is - days in adults. in infants and young children, the primary infection starts in the upper respiratory tract, with rhinorrhea, low-grade fever, and cough; it may progress to lower respiratory infection (lri) (bronchiolitis or pneumonia), at which stage most patients seek medical attention [ , ] . it is also likely to involve the sinuses and the middle ear. rsv is known to cause apnea in infants, although the mechanism of this action remains unknown. older children and adults typically present with upper respiratory tract symptoms, but may also have constitutional symptoms such as fever and malaise [ , ] . rsv upper respiratory infections (uris) can also progress to the lower respiratory tract, particularly in institutionalized adults and those with severe combined immunodeficiency, leukemia, hsct, and in lung transplant recipients [ , , ] . these patients may present with wheezing and shortness of breath, with or without underlying comorbidities or known hyperactive airways. almost % of elderly patients with rsv infections report wheezing [ ] [ ] [ ] [ ] . patients with weakened immune systems because of malignancy, chemotherapy, steroid use, hsct, or solid organ transplantation are at high risk for developing severe illnesses. they also tend to have a longer duration of infection, with a varied presentation [ , ] . leukemia patients and solid organ and stem cell transplant recipients are particularly at risk, with the latter being at high risk for pneumonia and death prior to engraftment [ ] . rsv infections begin as uri, but progress to lri in - % of hsct recipients, leading to respiratory failure with significant morbidity and mortality; some early studies described a mortality rate of - % [ , ] . in hsct recipients, the risk factors for progression to lri [ ] [ ] [ ] [ ] . torres et al. [ ] found that a high apache ii score and not giving aerosolized ribavirin treatment were independent predictors of progression to pneumonia in leukemia patients; those with uri who were treated with aerosolized ribavirin were less likely than untreated patients to develop pneumonia ( vs. %, p < . ) and die with rsv infection ( vs. %, p = . ). the overall mortality rate in the study was % [ ] . solid organ transplantation patients with rsv infection may present with dyspnea, cough, fever, and wheezing which progress to pneumonia in more than % of patients [ ] . in lung transplant recipients, a higher frequency of rsv pneumonia has been reported, but the mortality rate is low; the mortality rate is also low in kidney transplant recipients [ ] . in pediatric liver transplant patients, pohl et al. reported a mortality rate of % for rsv pneumonia with early infection onset and preexisting lung disease as predictors of severe disease [ ] . in children, genetic polymorphisms in cytokine and chemokine-related genes (interleukin [il]- , il- , il- , il- , and ccr ) and genes related to potential virus-cell surface interactions or cell signaling (tlr- , cx cr , sp-a, and sp-d) have been associated with severe rsv infections [ ] . data on hiv patients with rsv infections are scarce; however, a cohort study by miller et al. [ ] performed in the winter of - found no evidence of influenza, rsv, parainfluenza, adenovirus, or enterovirus infections in the bronchoscopic alveolar lavage fluids of hiv- -positive patients. rsv can be diagnosed on the basis of the clinical presentation in infants with lri during an outbreak period [ ] . rsv infections in adults cannot be clinically differentiated from other viral infections that cause upper respiratory symptoms. for a specific diagnosis, rsv must be detected in respiratory secretions. nasal aspirates or washes are most likely to give a positive rsv test in young children. if a nasal aspirate or wash cannot be obtained, a nasopharyngeal swab or throat swab may be used. because immunocompromised or intubated patients are more likely to develop an lri due to rsv, tracheal aspiration or bronchoalveolar lavage should be performed [ , ] . the gold standard for diagnosing rsv remains the identification of virus causing typical syncytia in cultures of hep- cells [ ] . viral cultures may take days (shell vial cultures) and up to weeks (routine cultures) to become positive, making isolation less relevant in most clinical settings. other available methods include antigen detection by enzyme-linked immunosorbent assay (elisa), immunoflourescence assay, and polymerase chain reaction (pcr) tests. fan et al. [ ] reported a sensitivity of - % for rsv detection using rapid antigen testing with elisa. pcr tests have been shown to be more sensitive than direct antigen detection [ , ] . for most immunocompromised patients, rsv treatment is focused on reducing symptom severity and preventing progression to lris. nonsteroidal anti-inflammatory drugs and antihistamines have been used in patients with a uri. ribavirin, a nucleoside analog, has in vitro activity against rsv and has been approved by the united states food and drug administration for the treatment of rsv infections in children. two schedules of aerosolized ribavirin have been used in immunocompromised patients: continuous and intermittent. on the continuous schedule, a daily dose of g (concentration, mg/ml) is delivered over h via a small particle aerosol generator unit, administered via a face mask in a tent. on the intermittent schedule, a concentration of mg/ml is delivered over h every h. aerosolized ribavirin may be beneficial in certain adults and children with lris. its early use has been shown in some retrospective studies to reduce morbidity and mortality in hsct recipients [ , ] and patients with hematologic malignancies, particularly when the infection is treated early on [ ]. however, its effectiveness in solid organ transplantation patients remains unknown [ ] . an rsv-specific monoclonal antibody (palivizumab) did not demonstrate a therapeutic benefit in a major study conducted in children [ ] . although the combination of ribavirin and intravenous immunoglobulin (ivig) or palivizumab has not been evaluated in a randomized trial, it is sometimes used in severely ill patients with rsv pneumonia, especially hsct recipients, given that they have high mortality rates from this infection [ , , ] . in patients at risk for progression to lri, aerosolized ribavirin should be considered at an early stage. a recent trial demonstrated that both the intermittent and continuous schedules of aerosolized ribavirin were effective at preventing progression to lris in % and % of patients with rsv uris and hematologic malignancies (including hsct recipients), respectively [ ]. rsv infection may be acquired nosocomially, thus specific infection control measures should be implemented when dealing with patients with known or suspected infections. briefly, patients should be isolated in private rooms when possible, and appropriate personnel protective equipments should be used (i.e., disposable gloves, masks, and gowns) [ ] . no licensed vaccine is available for rsv; however, two agents, rsv-ivig and palivizumab, have been used to prevent rsv infection. rsv-ivig has been studied in children younger than months with severe lung disease and those who were born prematurely [ ], but it was removed from the market in . a randomized double-blind placebo-controlled study in children found that palivizumab recipients had a % relative reduction in rsv hospitalizations ( p = . ). twenty-one children ( . %) died in the palivizumab group vs. ( . %) in the placebo group with no deaths attributable to palivizumab [ ] . palivizumab is easier to administer than regular ivig, which must be given over - h, and does not interfere with other vaccinations. however, rsv-ivig and ivig may provide additional protection against other respiratory viruses as well [ , , ]. influenza viruses infect both the upper and lower respiratory tracts. outbreaks are common every winter, although the severity of the disease varies considerably. influenza viruses belong to the orthomyxoviridae family, with influenza types a, b, and c constituting one genus. these rna viruses are enveloped and measure about - nm in diameter. the designation of the viruses into types is based on the stable antigenic characteristics of the nucleoprotein and the matrix protein antigens. influenza a and b viruses have surface glycoproteins known as hemagglutinins (ha) and neuraminidase (na). three hemagglutinin subtypes (h , h , and h ) and two neuraminidase subtypes (n and n ) have been described for the influenza a viruses that infect humans. major antigenic changes in the glycoprotein (basis for new subtypes) are called antigenic shift and can cause pandemics; minor antigenic changes occur frequently, are called antigenic drift, and cause the annual epidemics. influenza b has only exhibited minor antigenic drift [ , ] . the pandemic influenza a (h n ) contains segments present in north american swine for years, but is a new reassortant virus that acquired m and na gene segments from a eurasian adamantane-resistant swine influenza virus [ ] . influenza typically has a short incubation period and an abrupt onset of symptoms, such as headache, fever, chills, myalgia, and malaise, along with respiratory symptoms of runny nose, cough, and sore throat. it can also present as a febrile uri or with constitutional manifestations only or with few-to-no respiratory symptoms. influenza can progress to pneumonia in otherwise healthy persons, but particularly in patients with comorbidities such as lung diseases, heart disease, diabetes mellitus, renal diseases, and hemoglobinopathies, in immunocompromised individuals, residents of nursing homes or chronic care facilities, and in individuals over years of age [ ] . primary pneumonia occurs when the influenza virus directly involves the lung and should be suspected when clinical symptoms progress to high fever, dyspnea, and hypoxemia [ ] . influenza virus infections also affect the epithelium of the tracheobronchial tree, leading to impaired defense and a secondary bacterial pneumonia. influenza infections can increase morbidity and mortality rates in cancer patients [ ] . in a study of leukemia patients with influenza infections, % of patients developed pneumonia [ ], with cough and dyspnea being the most common manifestations. half of the patients had lymphopenia [ ] . the incidence of influenza infection in hsct recipients is reported as . % [ ], with a mortality rate up to % [ ], mainly due to respiratory failure. in one study [ ] , % of patients who developed influenza after hsct presented with pneumonia; of those who presented with uris, % experienced progression to lri. risk factors associated with progression to lri were lymphopenia and days from transplantation (i.e., pneumonia developed more commonly among those infected earlier after transplantation); whereas use of systemic steroids and autologous stem cell transplantation appeared to be protective. x-ray findings include a diffuse interstitial pattern or focal pulmonary infiltrates [ ] . among patients with solid organ transplantation, lung transplant recipients are at the highest risk for influenza virus infection [ ] . the initial presentation in these patients may not always be in the respiratory tract, as illness can present with nonspecific gastrointestinal symptoms; however, patients who required hospitalization always presented with pneumonia [ ] . progression to bronchiolitis obliterans, which is a characteristic of chronic lung rejection after infection, was also reported. in patients postrenal transplantation, influenza pneumonia is often acute, with high fever, cough, dyspnea, cyanosis, leukopenia, and thrombocytopenia [ ] . influenza infection can only be diagnosed clinically in persons exhibiting the classic syndrome during an epidemic. however, because other viruses can produce the same syndrome and influenza infection can produce other respiratory syndromes, a confirmatory test detecting the virus or viral antigens in nasal washes, throat swabs, respiratory tract secretions, or bronchoalveolar lavage specimens is needed in sporadic cases and in immunocompromised patients. viral culture remains the gold standard for diagnosis, but can take up to h to yield results [ ] . sputum and nasal washes or nose swabs are superior to throat swabs for diagnosis [ ] . rapid antigen testing using immunoflourescence assays, enzyme immunoassays, and pcr-based testing are used frequently in clinical settings [ ] . the results of these tests can be obtained in hours and can have good sensitivity ( - %) and specificity ( - %) if obtained early from those with more severe illnesses [ ] . pcr-based assays are more sensitive than rapid antigen testing for diagnosing influenza a and b, but are not often used because of availability and cost [ ] . samples should be obtained within - h of the appearance of symptoms for the most accurate results. two classes of drugs are available to treat influenza infections [ ] : the neuraminidase inhibitors, zanamivir and oseltamivir, which are active against influenza a and b viruses; and the m inhibitors, amantadine and rimantadine, which are only active against influenza a viruses [ ] . amantadines are not effective against influenza b and resistance has been reported for novel h n virus; resistance to oseltamivir has increased for the seasonal h n virus (table . ) [ , ] . therapy should be initiated as early as possible, preferably within h after symptom onset [ ] . zanamivir and oseltamivir have been used extensively to treat both influenza a and b; both were shown to reduce the mean duration of symptoms by day when used within h of symptom onset [ ] . both neuraminidase inhibitors have been shown to significantly decrease the incidence of complications associated with influenza such as development of pneumonia when compared to placebo [ ] . the most common side effects of oseltamivir are nausea and vomiting, although other toxicities have been reported. central nervous system side effects such as anxiety, insomnia, impaired thinking, confusion, lightheadedness, and hallucinations have been reported with amantadine use. newer ni inhibitors including parenteral preparations are under development [ ] . the mainstay of influenza prophylaxis in the general population is the administration of influenza vaccine. annual vaccination has been recommended for many years for people at high risk for complications, including those older than years, residents of nursing homes or other facilities, adults and children with chronic pulmonary or cardiovascular conditions, adults and children hospitalized during the previous year, women in the second and the third trimesters of pregnancy, immunocompromised patients, healthcare workers, and family member of those at high risk prior to the onset of influenza season [ ] . the american committee on immunization practices has recently recommended vaccination for all persons for whom there is no contraindication [ ] . currently, there are two types of vaccines available, an intramuscular (inactivated virus vaccine) and an intranasal (attenuated virus vaccine). the intranasal form is to be used only in healthy individuals between the ages of and years; it should not be used to vaccinate immunocompromised patients [ ] . individuals with a significant allergy to eggs or those with acute febrile illness should not be given vaccine [ ] . the advisory committee on immunization practices recommends that immunocompromised individuals be vaccinated prior to influenza season and receive daily chemoprophylaxis with an antiviral medication during community outbreaks [ ] . influenza chemoprophylaxis may be used in patients at high risk for complications if the vaccine is contraindicated or not likely to be completely protective. antiviral drugs should be administered to patients within the first months of hsct, those with documented graft-versus-host disease, unvaccinated healthcare workers who care for immunocompromised individuals, and residents of long-term care institutions during outbreak periods [ ] . the american society for blood and marrow transplantation (asbmt) guidelines [ ] for prevention of infection in hsct recipients recommend annual inactivated influenza vaccine before the beginning of the season and before stem cell transplant. the vaccine may be given - months after hsct. they also recommended prophylaxis and preemptive treatment during community and nosocomial outbreaks of influenza a for hsct recipients regardless of the vaccination status in those who are within months of the transplant or in those with more than months posttransplant, but have gvhd and/or are on immunosuppression. the drug to be used for chemoprophylaxis depends on the susceptibility pattern of the outbreak virus. piv are enveloped, single-stranded rna viruses that belong to the paramyxoviridae family. the envelope contains two glycoproteins, one with both ha and neuraminidase activity and the other with fusion activity. there are five types that share certain antigens with other members of the paramyxoviridae family [ , ] . of the five, piv- is the most prevalent, with most adults demonstrating the presence of antibodies [ ] . piv- infections may be epidemic in the spring and summer, but may also occur through the year; piv- is associated with pneumonia and bronchiolitis in infants. piv- and are associated with croup in children which occurs as outbreaks every years in the fall; piv- a and b cause only mild illnesses [ , ] . piv infections have a wide spectrum of presentations, from a simple uri to a life-threatening complication such as pneumonia. the incubation period is usually - days. young children may present with coryza, sore throat, hoarseness, and cough with chest x-rays revealing interstitial infiltrates. in adults, most infections are mild, but in immunocompromised individuals, the infection can progress to a lower respiratory tract illness including pneumonia and cause prolonged illness and even death [ , , ] . immunocompromised individuals are at risk for a piv infection that can progress to pneumonia. a study at our institution found a higher rate of progression among leukemia patients than among hsct recipients [ ] . piv usually presents as an uri. in a large study in hsct recipients over several years, piv infections were documented in . % of cases, with % being communityacquired [ ] . patients who have undergone hsct are at particular risk for developing severe piv-associated pneumonia [ ] [ ] [ ] [ ] ; wendt et al. [ ] reported a mortality rate up to %. coinfections (aspergillus fumigatus being the most common) and mechanical ventilation were found to be significant risk factors for piv pneumonia-associated mortality in one study [ ] . other factors associated with progression include neutropenia within month prior to infection, an apache ii score higher than , and pulmonary coinfection; this study also found a mortality rate around %, with no difference between those treated and those not treated with aerosolized ribavirin [ ] . patients who have undergone solid organ transplantation do not appear to be at increased risk for developing severe piv illness, but only a few studies have been reported [ , ] . one of these studies in lung transplant patients found piv infections in % of patients, all of whom developed pneumonia, but the majority ( %) were treated with aerosolized ribavirin and all but one recovered [ ] . except for croup in young children, the clinical pattern of piv infection is similar to that of other respiratory viruses and cannot be distinguished on the basis of symptoms alone. during a community outbreak, a presumptive diagnosis can be made; however, confirmation in laboratory tests may be appropriate in immunocompromised individuals. the virus can be detected in respiratory tract secretions, nasal washes, nasal swabs, throat swabs, and bronchoalveolar lavage specimens. viral culture remains the gold standard for diagnosis, but can take days to yield a result [ , ] . rapid antigen detection by immunofluorescence or elisa is most commonly used and can have a sensitivity of as high as - % [ ] . recent pcr-based assays have sensitivities up to %, with high specificity [ , ] . management of piv infection is mostly supportive as no piv-directed antiviral therapy has been licensed by the u.s. food and drug administration. ribavirin has been shown to be active against the virus in vitro and in animal models and has been used occasionally to treat immunocompromised patients with severe piv infections [ ] . one case series reported a decrease in piv viral loads and clinical improvement after aerosolized ribavirin treatment in children with severe immunodeficiency [ ] . nichols et al. [ ] reported that aerosolized ribavirin did not reduce viral shedding or mortality rates in hsct recipients after the infection had progressed to the lower respiratory tract. our data also showed no apparent benefit of aerosolized ribavirin on the mortality rate in hsct recipients and leukemia patients [ ] . on the other hand, a combination of methylprednisolone and intravenous or oral ribavirin has, apparently, been used successfully to treat piv pneumonia in a hsct and a heart transplant recipient, respectively [ , ] . currently, no licensed vaccine is available for the prevention of piv infection. hence, infection control measures play an important role in containing the spread of the infection. patients with suspected or confirmed infections should be isolated, and personnel protective equipment should be used. human adenoviruses are dna viruses belonging to the mastadenovirus genus of the adenoviridae family which measure about - nm in diameter [ , ] . there are at least known human serotypes divided into subgenera a to f based on the dna genome and pattern of hemagglutination [ ] . adenovirus infections are reported most frequently in infants and children and can occur throughout the year; however, they may cause serious infections in immunocompromised patients. after a primary infection in childhood, adenoviruses establish latency in adenoidal tissues along with lifelong persistence of specific antibodies [ , ] . adenovirus infections are transmitted by either inhalation of aerosolized virus, inoculation of the virus into conjunctival sac, or through the fecal-oral route [ , ] . subgroup a, type and various types from subgroups b and c have been associated with pneumonia and hepatitis [ ] . serotypes , , , and are associated with outbreaks of acute respiratory disease in military recruits, mostly in winter and spring [ ] . types , , , and are most common in children and present as an acute upper respiratory tract illness which can progress to lower respiratory disease; types and are less common, but can cause severe disease. in adults, infections due to adenovirus are characterized by sore throat and gradual onset of fever. cough, coryza, and regional lymphadenopathy are commonly seen. the most common clinical symptoms besides respiratory symptoms are fever and diarrhea [ ] . adenovirus infections are common after hsct and can occur as a localized illness or as part of a disseminated disease. it has been associated with delayed engraftment and graft failure. infections due to adenovirus in hsct recipients have a reported incidence of . - % [ , ] and are more commonly reported in allogeneic hsct recipients than in autologous transplant recipients ( vs. . %) [ ] ; however, the mortality rate can be as high as % in both groups [ , ] . some reports also suggest that the incidence of adenovirus infection in patients after hsct may be rising due to transplantation practices [ , ] . disseminated infection may occur without respiratory tract symptoms and disease can develop in almost any organ causing gastrointestinal disease, hepatitis, nephritis, pneumonia, conjunctivitis, thrombotic thrombocytopenic purpura, pancreatitis, or hemorrhagic cystitis. viremia may be present, but is not detected in all cases of disseminated disease [ ] . adenovirus is known to be fatal even in the absence of any respiratory tract involvement; however, if pneumonia is present, the mortality has been reported to be higher ( vs. %) [ ] . coinfections with aspergillus spp. and bacteria such as nocardia, legionella spp., and mycobacterium tuberculous are frequently seen in this patient population [ , ] . risk factors for adenovirus infections include gvhd, unrelated donor, total body irradiation, t-cell depletion, younger age (< years old), chronic disease, and recent transplantation [ , , , ] ; the degree of t-cell depletion and posttransplant suppression of t-cell function are the most important ones [ ] . adenovirus types and are associated with severe infections in hsct recipients [ ] . there are a few reports of adenovirus infections in solid organ transplant patients in whom the virus involved the donor organ and led to pneumonia, hepatitis, hemorrhagic cystitis, nephritis, enterocolitis, or disseminated disease. in patients with previous liver transplantation, adenovirus pneumonia had a reported prevalence of . % with a mortality rate of % [ ] . serotype is known to be associated with hepatitis [ , ] , whereas serotypes and are more commonly associated with pneumonia. in lung transplant recipients, one study found adenovirus infection to be an early complication following surgery with a prevalence rate of . % [ ] . progressive adenovirus infections are known to be associated with graft loss, progression to bronchiolitis obliterans, or death [ ] . adenovirus infection should be suspected in cases of acute respiratory disease in military recruits or during outbreaks. in most cases, infection caused by the virus cannot be differentiated from those caused by other respiratory viruses from the clinical presentation alone [ ] . a definitive diagnosis can be established by viral culture or the detection of specific viral antigens. viral culture remains the gold standard for identification of adenovirus. nasopharyngeal aspirate or swab, throat swab, sputum samples, or bronchoalveolar lavage can be used, depending on the site of the infection. a cytopathic effect is seen in human cell lines such as hela (cervix), a (lung), hek (human embryonic kidney), and hep- (larynx) by strains of adenovirus except for types and . adenovirus and grow well in hek cells. adenovirus-specific enzyme immunoassay (elisa) or immunofluorescence assay can be used to detect the presence of virus in clinical samples. these rapid tests suffer from a sensitivity of only about %; pcr-based assays can detect adenovirus dna from a variety of clinical specimens and has better sensitivity [ , ] . viral load quantification is a useful tool to measure prognosis and monitor clinical response [ , ] . viral loads higher than × copies/ml have been associated with an increased likelihood of death [ , ] . there have been no randomized clinical trials for the treatment of adenovirus infection in immunocompromised patients. most patients are managed using symptom-based treatment and supportive therapy. cidofovir is currently being used in immunocompromised patients since it has been shown to decrease plasma viral loads in hsct recipients [ ] . although cidofovir is active against all strains of adenovirus in vitro, only retrospective data are available on the efficacy of cidofovir in hsct [ ] [ ] [ ] [ ] and solid organ transplant recipients [ ] . nephrotoxicity is commonly encountered with this drug and it should be used with caution. there are two accepted regimens: mg/kg every - weeks, or mg/kg times per week, with the latter being associated with less nephrotoxicity [ ] . orally active ether lipid-ester prodrugs of cidofovir (s)-hpmpa are under development with some promising results in in vitro experiments and phase i trials [ ] . when tested against five adenovirus serotypes, they were shown to be more active than the unmodified parent compounds [ ] . intravenous ribavirin has been used in a few reported cases, but results were conflicting [ ] [ ] [ ] . finally, another treatment option using adenovirus-specific donor t-cells infusion has been shown to be feasible and effective in protecting children from complications due to adenovirus infection and causing a significant decrease in viral loads [ ] . currently, there are no vaccines available for adenovirus infection other than the oral partially attenuated vaccines contained in enteric-coated capsules with use restricted to the military [ ] . only routine infection control practices are recommended for civilian populations where special precautions must be taken for contact and droplet exposure. in hsct recipients at high risk from adenovirus infection, weekly pcr surveillance of viremia and preemptive treatment with cidofovir can be used [ , ] . rhinoviruses are members of the picornaviridae family. they are nonenveloped, single-stranded rna viruses that measure about - nm. the capsid of the virus is icosahedral and contains copies of four polypeptides each. a canyon on the viral surface contains the attachment site for the host-cell receptor, with most rhinoviruses using this site to attach to the intercellular adhesion molecule- receptor expressed on the surface of host cells [ , ] . more than serotypes of the virus have been isolated, making a vaccine unlikely in the near future [ , , ] . rhinoviruses are proven to cause - % of common colds in adults [ ] . each year, adults experience or colds, whereas children may experience - [ , ] . children are the major reservoir for rhinoviruses with infection rates decreasing with age. although infections occur throughout the year, peaks may occur in the fall and spring. this infection is primarily due to the deposition of the virus on the nasal mucosa. this can occur via self-inoculation or contact with infected secretions such as small-and large-particle aerosols (respiratory droplets) [ , , ] . individuals with rhinovirus infections may be asymptomatic. when symptoms occur, they are typically those of the common cold: most commonly rhinorrhea and sneezing, which are associated with nasal congestion. infections due to rhinovirus have an incubation period of - days. adults characteristically experience sneezing, nasal obstruction and discharge along with cough, and a sore or scratchy throat. sinuses are commonly involved so that the illness is rhinosinusitis. symptoms may last for - days and usually resolve with no complications. fever is not usually associated with adult illness [ , ] . children, on the other hand, may experience fever, cough, and nasal discharge and obstruction. in addition, the duration of symptoms may be longer. although bronchitis, bronchiolitis, and bronchopneumonia have been reported in children, rhinovirus is not usually a major cause of lower respiratory illness [ , ] . however, they are an important cause of exacerbations of asthma and chronic obstructive pulmonary disease in children and adults and of lri in the elderly [ , ] . a retrospective study in adults with rhinovirus infections who had undergone hsct found that % of patients developed and eventually died of pneumonia [ ] . one patient was found to have a coinfection with aspergillus spp. on autopsy; most of the other patients had interstitial pneumonitis and/or acute respiratory distress syndrome [ ] . a study performed at another institution reported that % of hsct recipients with rhinovirus infections developed pneumonia and % died [ ] . in a study of community-acquired pneumonia in immunocompromised patients (after hsct or solid organ transplantation, with hiv infection, or receiving steroids or chemotherapy), rhinovirus was responsible for about % of cases, with a mortality rate of % [ ] . these findings suggest that rhinovirus may cause more severe complications in immunocompromised patients than previously thought. many viruses cause common cold symptoms, and a definitive diagnosis cannot be made only on the basis of the presenting symptoms but rhinoviruses are most commonly associated with colds. a definitive diagnosis can be made by isolating the virus from nasal washes or other nasal secretion specimens in tissue culture. newer diagnostic methods such as real-time reverse-transcription pcr (rt-pcr) have been used; however, these tests are not frequently performed given the self-limited nature of most infections. no specific antiviral treatment is available for rhinovirus infections. most cases are managed with supportive care using antihistamines, decongestants, and nonsteroidal antiinflammatory drugs. given the number of rhinovirus serotypes known to cause infection, an effective vaccine is unlikely to be developed in the near future. infection control measures such as hand washing and isolating patients who are known or suspected to have rhinovirus infections can help contain the spread of the virus. hmpv belongs to the subfamily pneumovirinae of the paramyxoviridae family. the virus was discovered in and is genetically similar to rsv. hmpv is an enveloped rna virus that is known to cause uris and lris in all age groups [ ] . although hmpv infections are more common in children, a recent study revealed an infection rate of . % in adults with acute upper respiratory illness; % of patients required hospitalization [ ] . hmpv usually causes a mild infection of short duration (about - days) and is self-limiting. the incubation period is - days [ ] . the most common presenting symptoms in adults are cough, nasal congestion, rhinorrhea, dyspnea, hoarseness, and wheezing [ ] . children often present with cough, rhinitis, fever, and wheezing [ , ] . a recent study of patients with hematologic malignancies revealed a % incidence of hmpv infection. nine of the patients who had undergone hsct had pneumonia; three of these patients died [ ] . another study in hsct recipients detected the presence of hmpv in % (in out of ) of bronchoalveolar lavage specimens. these patients presented with fever, cough, nasal congestion, and sore throat within the first days after transplantation. the infection progressed to respiratory failure, pulmonary hemorrhage, and culture-negative septic shock. the mortality rate was % ( of patients) [ ] . hmpv's growth in culture is slow and unreliable, which makes this method of diagnosis impractical. the presenting symptoms are similar to those of other infections that cause an acute respiratory illness, making clinical diagnosis impossible. pcr-based methods have been used to diagnose the infection in some centers. no specific antiviral therapy exists for hmpv infections. in vitro studies have demonstrated that ribavirin has activity against the virus [ ] ; however, no clinical studies have been reported. immune serum globulins may neutralize the virus as demonstrated in one in vitro study [ ] . anecdotally, a lung transplant recipient with respiratory failure secondary to hmpv pneumonia was treated successfully with aerosolized ribavirin [ ] . the use of general preventive measures for patients with known or suspected hmpv infections can help reduce the rate of transmission. no vaccine is currently available for this virus. coronaviruses are single-stranded rna viruses that measure about - nm in diameter. they are enveloped with clubshaped projections, giving them a crown-like appearancehence the name [ , ] . coronaviruses can cause diarrhea in infants and may play a role in demyelinating diseases of the central nervous system [ , ] . coronaviruses are difficult to grow in vitro; some strains can only be grown in human tracheal organ cultures. the major antigenic types that cause diseases in humans are e, oc , hk, and nl viruses which cause common colds and may also cause lower respiratory tract illnesses in young children, and sars-cov, which causes a severe acute respiratory syndrome [ , , ] . coronaviruses are found in all tropical, subtropical, and temperate climates. most infections occur in the late fall, winter, and early spring [ , ] . cyclical outbreaks of these infections may occur every - years. respiratory infections due to hcov- e and hcov-oc strains probably spread in a manner similar to rhinovirus, i.e., via direct contact with infected secretions or aerosol droplets [ ] . the clinical manifestations of coronavirus infections are similar to those of rhinovirus infections. the most common symptoms are rhinorrhea, throat congestion, and fever. the middle ear may also be affected, leading to effusions and acute otitis media, especially in children [ , ] . the incubation period is - days and the duration of the illness is shorter than that of rhinovirus infections, a mean of - days. the subtype sars-cov has a slightly longer incubation period of - days. in immunocompromised adults, coronaviruses can cause lower respiratory tract infections [ , ] . a recent case series found that the infection rate among immunocompromised patients was significantly higher when compared to immunocompetent patients ( . vs. . %) [ ] . no large studies have been conducted in cancer patients or other immunocompromised individuals, but a few cases have been reported. folz and elkordy [ ] reported a patient who had undergone autologous hsct and was diagnosed with a coronavirus lri. the patient developed a fever, sore throat, cough, and severe hypoxia and was treated successfully using supportive measures. kumar et al. [ ] described a patient who developed a fatal severe acute respiratory syndrome (sars) after liver transplantation. epidemiologically, these infections should be suspected during the late fall or winter or during an outbreak. however, no practical method is available to confirm the infection except pcr-based tests. these are used to diagnose coronavirus infections at some institutions [ , ] . similar to rhinovirus, no specific antiviral therapy is available for coronavirus. most patients respond well to supportive treatment with nonsteroidal anti-inflammatory drugs, decongestants and antihistamines. general preventive measures must be used around patients with coronavirus infections; hand washing, disposing of infected material carefully, and proper disinfection can help prevent the spread of the virus. no vaccine against the virus has been developed. enteroviruses are single-stranded rna viruses that can multiply in the gastrointestinal tract. they belong to the picornaviridae family, but are relatively stable at a low ph. the virus is surrounded by an icosahedral capsid comprising four viral proteins. it has no lipid envelope and is not susceptible to alcohol, ether, or detergents. poliovirus is the prototype of this group, but nonpolio enteroviruses have a wide spectrum of manifestations including acute respiratory illnesses [ , ] . enterovirus infections are more common in developing countries and socioeconomically depressed areas and are associated with poor hygiene and sanitation. they can occur throughout the year, but the peak occurs in the summer and fall [ , ] . they can be transmitted by direct contact with feces during activities such as cleaning and diaper handling. the clinical picture of enterovirus infections is similar to that of rhinovirus infections; no specific symptoms are associated with the virus. immunocompromised patients have been reported to develop central nervous system infections, chronic disseminated infections, and a dermatomyositis-like syndrome with enterovirus infection. in a study of respiratory tract infections in hematologic malignancy patients, % of patients had enterovirus infections; most ( %) were hsct recipients with % of these developing pneumonia [ ] . a study from spain reported on four hsct recipients who developed enterovirus infections, with a mortality rate of % ( of ) [ ] . these findings demonstrate that immunocompromised patients may be at risk for lower respiratory tract infections and death from an enterovirus infection [ , , ] . most enteroviruses can be isolated in cell cultures from nasopharyngeal or throat swabs. pcr-based testing is used to amplify the viral rna from throat swabs, cerebrospinal fluid, and tissues. for diagnosis, isolation of the virus from the throat is more clinically significant than from stool because this virus has a shorter duration of shedding from the throat. most enterovirus infections are self-limiting and do not require specific treatment. intensive care may be required for central nervous system, cardiac, and hepatic infections. ivigs have been used in some patients with severe infections, but no specific antiviral therapies are available [ , ] . general infection control measures must be undertaken around patients with enterovirus infections. special attention must be paid to hand hygiene. material in contact with or soiled by feces should be handled carefully and discarded with proper precautions. the respiratory viruses as a group are the most common cause of an acute infectious illness in developed societies. the variety of viruses that can cause infection and illness in all age groups and their presence in high frequencies throughout the year describe the risk of exposure of all persons at all times including immunocompromised persons residing in the community. the immunocompromised state of many cancer patients constitutes the basis for the frequent failure of the host to promote a normal and rapid recovery from an acute respiratory viral infection and results in a more severe and prolonged infection that causes significant morbidity and mortality in these patients. those respiratory viruses that are most prevalent and most prone to produce lower respiratory illnesses and pneumonia in healthy hosts, rsv, influenza viruses and piv, are those most likely to cause severe illness and pneumonia leading to hospitalization in immunocompromised persons. however, viruses less prone to produce a lower respiratory illness but that are highly prevalent, such as rhinoviruses, may frequently be associated with severe illness. although not generally considered respiratory viruses, the herpes viruses are known to produce respiratory infection and disease, sometimes severe, in immunocompromised patients (table . ). a historically important virus, measles virus, is now rarely encountered because of widespread vaccination. the limited availability of antivirals and vaccines for the acute respiratory viruses means that these infections will continue to be important for many years and dictate a need for utilizing infection control procedures as much as possible, particularly in hospitals and institutions, so as to minimize spread. efforts to develop specific vaccines are important as their use could prevent as well as reduce exposure of cancer patients to these viruses. development of specific antivirals is important for use in immunocompromised patients as normal recovery mechanisms may be seriously impaired. community respiratory virus infections in immunocompromised patients with cancer common community respiratory viruses in patients with cancer: more than just "common colds harrison's principles of internal medicine circulation patterns of genetically distinct group a and b strains of human respiratory syncytial virus in a community respiratory syncytial virus and parainfluenza virus risk of primary infection and reinfection with respiratory syncytial virus respiratory syncytial virus infections in previously healthy working adults respiratory syncytial and other virus infections in persons with chronic cardiopulmonary disease can respiratory syncytial virus and influenza a be distinguished clinically in institutionalized older persons? symptomatic respiratory syncytial virus infection in previously healthy young adults living in a crowded military environment respiratory syncytial virus and parainfluenza virus infections in the immunocompromised host respiratory virus infections in immunocompetent and immunocompromised persons respiratory syncytial virus infection in adults respiratory syncytial virusinduced acute lung injury in adult patients with bone marrow transplants combination therapy with aerosolized ribavirin and intravenous immunoglobulin for respiratory syncytial virus disease in adult bone marrow transplant recipients diagnosis and epidemiology of community-acquired respiratory virus infections in the immunocompromised host respiratory virus infections 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epidemiology of two consecutive outbreaks of parainfluenza in a bone marrow transplant unit paramyxovirus infection in lung transplant recipients influenza and parainfluenza respiratory viral infection requiring admission in adult lung transplant recipients comparison of different tissue cultures for isolation and quantitation of influenza and parainfluenza viruses efficiency of immunofluorescence for rapid detection of common respiratory viruses direct detection of respiratory syncytial virus, parainfluenza virus, and adenovirus in clinical respiratory specimens by a multiplex reverse transcription-pcr assay biochemistry and clinical applications of ribavirin treatment of respiratory viral infection in an immunodeficient infant with ribavirin aerosol treatment of parainfluenza virus pneumonia in a cardiac transplant recipient with intravenous ribavirin and methylprednisolone successful treatment of parainfluenza virus pneumonia with oral ribavirin and methylprednisolone in a bone 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severe clinical adenovirus infection, united states adenovirus infection in adult orthotopic liver transplant recipients: incidence and clinical significance adenovirus infection in pediatric liver transplant recipients adenovirus pneumonia in lung transplant recipients adenovirus infection in the lung results in graft failure after lung transplantation comparison of pcr, enzyme immunoassay and conventional culture for adenovirus detection in bone marrow transplant patients with hemorrhagic cystitis efficacy of pcr and other diagnostic methods for the detection of respiratory adenoviral infections effect of ribavirin on the plasma viral dna load in patients with disseminating adenovirus infection real-time blood plasma polymerase chain reaction for management of disseminated adenovirus infection internally controlled real-time pcr monitoring of adenovirus dna load in serum or plasma of transplant recipients high levels of adenovirus dna in serum correlate with fatal outcome of 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common respiratory viruses in asthmatic and nonasthmatic children typing of human enteroviruses by partial sequencing of vp enteroviral disease in the united states pulmonary enterovirus infections in stem cell transplant recipients successful treatment of echovirus meningoencephalitis and myositis-fasciitis with intravenous immune globulin therapy in a patient with x-linked agammaglobulinemia key: cord- -b w ob authors: nan title: th meeting of the austrian society of transplantation, transfusion, and genetics, october – , date: journal: eur surg doi: . /s - - - sha: doc_id: cord_uid: b w ob nan background. nowadays lung transplantation is an established standard procedure for the treatment of most end-stage respiratory disorders. in the last years, like for all solid-organ transplantations, the demand for donor lungs exceeded significantly the existing organ pool. thus, most specialised centers face a high mortality on the waiting list. this retrospective study compares the outcome of marginal versus ideal donor lungs. methods. we performed a retrospective analysis of consecutive primary lung transplantations from donors from / to / . recipients were divided in two groups (standard versus "extended") according to the donor lung acceptance criteria (age, > years; pao at fio /peep , < mmhg; positive tobacco anamnesis [> packages per year]; inhalation of noxious agents; presence of infiltration on chest x-ray or purulent secretions at bronchoscopy). results. twenty-three donors ( . %) were extended. twenty-six recipients ( . %) received organs from extended donors. according to our data, differences in intubation times, icu stay, and hospital stay were not statistically significant. furthermore, postoperative bleeding rates were comparable as well as bronchial anastomotic complications. we encountered no significant statistical difference in the -month (standard . % vs. extended . %) and -year (standard . % vs. extended . %) survival between the two groups. conclusions. our study suggests that the use of selected marginal donor lungs has no influence on the outcome after transplantation. background. female donor gender has been described to be an independent risk factor for primary graft failure. we performed this study to evaluate the impact of donor gender on outcome and complications after lung transplantation. methods. we retrospectively reviewed the impact of donor gender on outcome of primary lung transplant recipients ( recipients were male [ %], were female [ %]) from january to december . recipients were stratified whether they received a female-or male-donor organ. both groups were compared with regard to duration of intubation time, icu stay, postoperative complications and survival. both groups were comparable with regard to mean age, indications, and mean waiting list time. results. mean time until extubation was days in the group receiving organs from male donors and days in the group receiving organs from female donors (p = . ). mean icu stay of days for the male-donor recipient group was significantly shorter than that for the female-donor recipient group, days (p = . ). -month survival rates were comparable in both groups: . % (male-donor recipi-lunge th meeting of the austrian society of transplantation, transfusion and genetics background. nowadays lung transplantation is an established therapeutic option for most end-stage respiratory diseases and one of the fastest-growing solid-organ transplantation procedures in the world, reflected by the enormous demand for lungs. this retrospective review of the vienna lung transplantation group demonstrates data about waiting time and mortality rate for various end-stage respiratory diseases for the years and . methods. . different variables specific for pretransplant period such as probability of transplantation, death on the waiting list, and time till transplantation were analysed for most frequent end-stage respiratory diseases. results. we found no significant changes in the average mortality rate (constantly %) on the waiting list during the studied period. however, patients suffering from pph have the highest probability of dying while being listed for lung transplantation ( , n = [ %] ; , n = [ . %]), followed by patients with idiopathic fibrosis ( , n = [ %]; , n = [ . %] ). subsequently the next subgroup is represented by cystic fibrosis patients, who are characterized by a moderate mortality rate ( , n = [ . %] ; , n = [ %]). in contrast, patients who were suffering from copd showed the lowest probability of dying ( , n = [ %]; , n = [ . %]). the average waiting time for the observed years amounts to . days ( , ± [min, ; max, ] days; , ± [min, ; max, ] days). regarding the disease-specific waiting time, pph patients had to wait a longer period ( ± days) for their transplantation than patients with other diagnosis. conclusions. waiting time and mortality on the waiting list are showing remarkable differences within the disease-specific subgroups. v increased recipient vegf serum level is a risk factor for severe reperfusion edema after lung transplantation k. krenn, s. taghavi, w. klepetko, s. aharinejad laboratorium für kardiovaskuläre forschung, zentrum für anatomie und zellbiologie, medizinische universität wien, wien, Österreich background. primary graft dysfunction (pgd) due to ischemia-reperfusion injury is a severe complication in lung transplantation (ltx) . therapeutic strategies are limited and there exist no preoperative markers to predict the risk for reperfusion-induced edema. vascular endothelial growth factor (vegf) is a key regulator of vascular permeability. methods. preoperative vegf serum levels were measured by elisa for patients undergoing ltx. underlying diseases in ltx patients were copd (n = ), cystic fibrosis (n = ), idiopathic pulmonary fibrosis (n = ), emphysema (n = ), primary pulmonary hypertension (n = ), sarcoidosis (n = ), and others (n = ). the ischemia time of the grafts and the blood gas parameters in donors were comparable. reperfusion edema was diagnosed and scored by characteristic changes in chest radiographs and deteriorating blood gases according to the guidelines of the international society for heart and lung transplantation, grading pgd from to ( , none; , only radiographic evidence; , moderate, pao /fio ratio of - ; , severe, pao /fio ratio of < or ecmo support necessary). results. grade pgd occurred in %, grade in %, grade in %, and grade in % of the patients. the preoperative vegf serum levels were significantly higher in patients with pgd grade and versus those without clinically relevant pgd (grade and ) following ltx (p = . ). preoperative recipient vegf serum levels significantly predicted pgd in receiver operating characteristic analysis (p = . , auc = . , ci = . - . ). conclusions. preoperative serum vegf levels in patients awaiting ltx could identify those at risk for reperfusion-induced edema following transplantation. background. airway stenoses are well known after lung transplantation although most occur due to surgical problems of the bronchial sutures. we wanted to analyse the use of endobronchial stenting in stenoses not related to the bronchial anastomoses. methods. we performed a retrospective analysis in patients after bilateral lung transplantation with consecutive stent placement aside the bronchial anastomoses. the indication for stent implantation was central bronchial stenoses due to bronchomalacia, granulation tissue with bronchial wall destruction, and endoluminal stenosis with airflow limitation or occurrence of segmental or lobar atelectasis. we used boston ultraflex ( of ), rüsch polyflex ( of ), rüsch dynamic ( of ) stents. results. in lung allograft recipients, we implanted in total stents between and month after bilateral lung transplantation. the predominant locations were the right ( %) and left lower lobe bronchus ( %) and the bronchus intermedius ( %). the patient had granulation tissue proliferation due to ischemia and/or chronic bacterial or fungal bronchitis. only one patient had concomitant anastomoses dehiscence. the stents remained in situ from day to days after placement. in of stent placements no severe complications occurred. stent migration was observed in of ; severe granulation tissue that revealed further interventional treatment, in patients. one patient died because of necrotizing vasculitis and letal hemoptysis. all other stents are still in place ( of ), were in place at the time of death due to other reasons ( of ), or were explanted regularly ( of ). conclusions. endobronchial stent placement is an effective treatment for bronchial stenoses that are not related to the bronchial anastomoses. complications occurred in about % of all stents. v preoperative oral corticosteroids predict the risk of late postoperative bleeding and perioperative mortality in lung transplantation conclusions. patients listed for lung transplantation with high-dose preoperative oral cs intake have a significantly increased risk of late postoperative bleeding. the perioperative mortality and the probability of -year survival of recipients with late bleeding are severely affected compared with patients with no or early bleeding. background. the efficacy of induction therapy after lung transplantation remains controversial and data on its use are limited. we hypothesized that induction therapy would have an impact on incidence of early rejection after lung transplantation and may cause a higher rate of infectious complications during the first months post transplantation. methods. all patients who underwent lutx between jannuary and march and received induction therapy with rabbit antithymocyte globulin ( . mg/kg/d for - days) were analysed retrospectively. basic immunosuppression consisted of tacrolimus, mycophenolate mofetil, and prednisone. primary end point was patient survival after months. secondary end point was histologically proven grade i or higher grade of rejection within the first months, incidence of infectious episodes and bronchiolitis obliterans syndrome (bos). a total of adult patients who underwent single (n = ), heart-lung (n = ), or bilateral lung (n = ) transplantation entered the study. female, ( %); male, ( %). mean age was ± . years (range, . - . years). results. the -month survival using kaplan-meier analysis was %; patients are alive, two died and days after transplantation. one patient was retransplanted days after primary transplantation because of graft dysfunction. follow-up ranged from a minimum of days to days (mean, ± days). patients had one histologically proven rejection episode. rejections were graded ai for patients, grade ai-ii for , and grade aii for patients. rejection episodes were treated with iv methylprednisone, no recurrent or ongoing rejections were observed. incidence of bacterial infections requiring treatment was / patients days. cmv infections, non-cmv viral infection, and fungal infections ( candida, aspergillus) were diagnosed and treated. no patient developed signs of bos of grade ≥ . no lymphoma occurred. conclusions. this retrospective analysis suggests that induction therapy with atg in combination with tacrolimusbased triple immunosuppressive regimen results in excellent survival rates and a low rate of acute early rejections. however, this high immunosuppression efficacy was paralleled by a considerably high rate of bacterial infections during the first postoperative months. rin inhibitor based quadruple drug is. an extensive infectious monitoring was used in this cohort. results. one-year patient survival was . %, perioperative rejection rate %. infection incidence during first hospitalization was . % ( . episodes per transplant): pneumonia, %; sepsis, %; wound infection, %; hsv/vzv, %; uti, %. during follow-up, cmv-associated complications were observed in % of patients including cmv infection (n = ), cmv disease (n = ). there were nine patients with cmv syndrome, ten patients with cmv graft pneumonitis and two patients with cmv gastrointestinal disease. excluding the retransplants and the perioperative deaths, the incidence of aspergillosis was %, six patients with aspergillus tracheobronchitis and seven patients with invasive disease. a total of patients died during follow-up, from infectious complications. as part of our centers' microbiological monitoring, specimens ( /transplant) were collected. these specimens were taken on observation days. investigated specimens were sterile and in specimens microorganisms could be isolated ( normal flora, pathogens). a total of pathogens were identified: gram-positive cocci, gram-negative rods, pseudomonas/acinetobacter, candida. > % of staphylococcus aureus and % of coagulase-negative staphylococci were methicillin resistant. other multiresistant organisms were e. faecium (no vre), n = ; corynebacterium jk, n = ; stenotrophomonas maltophila/burkholderia cepacia, n = . conclusions. infection remains the most common complication and the most common cause of death following lung transplantation. further refinement of infectious prophylaxis is required to improve results. background. valgancyclovir is routinely used for prophylaxis against cytomegalovirus (cmv) reactivation in lung transplant recipients. problems may arise due to its myelotoxic properties. it is unclear whether cytotect ® , a human igg cmv antibody preparation, offers similar protection against cmv reactivation without causing neutropenia. methods. we report on a female patient ( years) who received a double lung transplant in . cmv status was d+r+. four months postoperatively, she developped cmv pneumonitis and gastroduodenitis and was treated with gancyclovir and cytotect. thereafter, valgancyclovir was instituted but had to be stopped repeatedly because of neutropenia; each withdrawal of valgancyclovir prompted further reactivation episodes as detected by pp monitoring. in july , she was started on cytotect, mg q.d. over five days, followed by weekly doses of mg for twelve weeks. results. cytotect was tolerated without side effects. of note, leucocyte counts remained within normal limits. since she has been started on cytotect, she has not experienced any further episodes of cmv reactivation until to date. conclusions. the observations made in this patient suggest that long-term therapy with cytotect ® has the potential to prevent cmv reactivation in selected patients who do not tolerate valgancyclovir due to its myelotoxic side effects. v die kombinationsprophylaxe verbessert die cmv bedingte morbidität und mortalität und reduziert das risiko der bronchiolitis obliterans (bos) nach lungentransplantation e. ruttmann, c. geltner, b. bucher, h. ulmer, d. höfer, h. b. hangler, s. semsroth, h. bonatti, r. margreiter, g. laufer, l. müller klinische abteilung für herzchirurgie, universitätsklinik für chirurgie, medizinische universität innsbruck, innsbruck, Österreich grundlagen. die opportunistische cmv-infektion stellt ein schwerwiegendes problem nach lungentransplantation dar. ziel dieser untersuchung war, den einfluss der cmv-kombinationsprophylaxe mittels ganciclovir und cmv-hyperimmunglobulinen (cmv-ig) bezüglich patientenüberleben, cmv-reaktivierung, klinischer cmv-erkrankung und entwicklung der bronchiolitis obliterans (bos) zu evaluieren. methodik. eine konsekutive serie von cmv-hochrisikopatienten (d+/r-, d+/r+) mit einem minimalen followup von mindestens jahr post-transplant wurden analysiert. dreißig patienten ( , %) erhielten eine alleinige ganciclovir-prophylaxe für monate (kontrollgruppe), transplantationsempfänger ( , %) erhielten eine zusätzliche prophylaxe mit cmv-ig (cytotect ® , biotest pharma) in dosen während des . postoperativen monats (studiengruppe). das mediane follow-up betrug , monate in der kontrollgruppe und , monate in der studiengruppe (p = , ). ergebnisse. insgesamt cmv-assoziierte todesfälle ( , %) ereigneten sich in der kontrollgruppe, jedoch keiner in der studiengruppe (p = , ). in der kontrollgruppe wurden fälle mit klinischer cmv-erkrankung beobachtet ( , %), in der studiengruppe patienten ( , %) (p = , ). zusätzlich zeigte sich ein signifikant verbessertes patientenüberleben in der studiengruppe (log-rank, p = , ). die -jahresfreiheit von cmv-reaktivierungen betrug , % in der kontrollgruppe und , % in der studiengruppe (logrank, p = , ). die -jahresfreiheit von bos war signifikant höher in der studiengruppe ( , % vs. %, log-rank, p = , ). schlussfolgerungen. eine zusätzliche cmv-hyperimmunglobulinprophylaxe senkt die cmv-assoziierte morbidität und mortalität. weiters kann das auftreten der bos mittels augmentierter cmv-prophylaxe reduziert werden. durch die dadurch reduzierte morbidität ist die kosteneffizienz gegeben. v extracorporeal photoimmune therapy (ecp) with uvadex in conjunction with standard therapy compared to standard therapy alone for the prevention of rejection in lung transplantation patients p. jaksch, r. knobler, b. schlechta, s. guth, w. klepetko klinische abteilung für herz-thoraxchirurgie, universitätsklinik für chirurgie, medizinische universität wien, wien, Österreich background. extracorporeal photopheresis has been shown to be beneficial in acute and chronic rejection in heart transplant patients and has also been used in lung transplant recipients with acute rejection or bronchiolitis obliterans. methods. we performed a prospective study to document the efficacy of photoimmune therapy in the prevention of acute rejections in the first months after lung transplantation. lung transplant recipients with copd were randomized in groups. group a ( pat) received in total ecp treatments (starting weeks after tx) and group b without ecp or other kind of induction therapy, both groups receiving standard triple immunosuppression with tacrolimus, mycophenolate, and steroids. surveillance bronchoscopies with biopsies were performed after , , , , , and weeks. primary objectives were acute biopsy-proven rejections of ishlt grade > , secondary objectives were number of infections (cmv, bacterial, fungal, viral non-cmv, tuberculosis, parasitic), patients and graft survival. results. demographics in both groups were similar (gender, age, underlying disease, cmv mismatch, and type of tx). fev % and mef % values after year were equal ( . ± . % vs. ± . and . ± . % vs. . ± . %, respectively) . the number of rejections in group a (with ecp) was lower than in group b ( . ± . vs. . ± . , p = . ), as well as the median rejection grade ( . ± . for group a vs. . ± . for group b, p = . ). after one year post tx, all patients are alive, none developed bo(s) within the first year post tx. conclusions. our preliminary data show a clear trend towards reducing the number and severity of acute rejections in lung transplant recipients. the number of infections was similar in both groups. adding ecp to a standard triple-drug immunosuppressive regimen seems to be a safe and efficient tool in reducing rejection rates without increasing the rate of bacterial, fungal, or viral infection. grundlagen. erhöhte natriumspiegel bei organspendern können mit der bildung von reperfusionsödemen und transplantatdysfunktion assoziiert sein. unklarheit besteht allerdings über die klinischen auswirkungen von erhöhtem spender-natrium nach herztransplantation (htx). in dieser studie wurde der einfluss von hohem spender-natrium auf die frühund -jahresmortalität nach htx in einem großen patientenkollektiv analysiert. methodik. es wurden herztransplantationen der eurotransplant-region aus dem zeitraum jänner bis dezember analysiert. entsprechend der spender-natrium-spiegel (sns) wurde das kollektiv in drei gruppen unterteilt: gruppe a, < mmol na + je liter n = ; gruppe b, - mmol na + je liter n = ; gruppe c, ≥ mmol na + je liter n = . eine kaplan-meier-Überlebensanalyse und eine multivariate analyse bezüglich des einflusses des sns wurden durchgeführt. endpunkte waren die mortalität ein monat und ein jahr nach htx. ergebnisse. der sns hatte in der univariaten analyse keinen einfluss auf die frühmortalität und einen grenzwertig signifikanten einfluss auf die -jahres-mortalität (p = , ). in der multivariaten analyse war dieser effekt nicht signifikant (p = , ). die kombination aus hohem spenderalter mit hohem sns hatte jedoch in der multivariaten analyse einen hochsignifikanten einfluss auf die früh-und -jahresmortalität (p = , ). schlussfolgerungen. diese daten zeigen, dass hohe sns mit einer erhöhten früh-und -jahresmortalität nach htx einhergehen. diese ergebnisse stehen im gegensatz zu früheren arbeiten mit geringeren patientenzahlen. vor allem die kombination aus höherem spenderalter und hohem sns zeigt einen deutlichen risikoanstieg. herzen von spendern mit einem natriumspiegel von > mmol/l sollten nicht elektiv transplantiert werden, bei gleichzeitig hohem spenderalter sollte das herz nicht verwendet werden. die organempfänger wurden entsprechend dem spenderalter in gruppen geteilt: gruppe , < a, n = ; gruppe , > a, n = . die gruppen waren hinsichtlich organischämiezeit, geschlechts-und cmv-mismatch sowie lipidstatus, nierenfunktion und medikamentöse therapie (immunsuppresion, ace-hemmer, statine, ca-antagonisten) jahr post-htx vergleichbar. pv war in gruppe tendenziell niedriger als in gruppe ( , ± , mm vs. ± mm ; p > , ). umgekehrt wies gruppe im verlauf des . jahres nach htx einen zuwachs an plaquevolumen auf, während in gruppe eine abnahme (∆pv, , ± , mm vs. , ± , mm ; p > , ) festgestellt wurde. von den oben angeführten risikoparametern zeigte lediglich der triglyzeridspiegel jahr post htx eine signifikante korrelation mit ∆pv (r = - , , p = , ). ∆pv und spenderalter waren nicht miteinander korreliert. schlussfolgerungen ergebnisse. die mortalität beträgt %. wegen gastrointestinaler beschwerden (Übelkeit, erbrechen) mussten patienten ( %) auf ein anderes immunsuppressives schema umgestellt werden. bei patienten (everolimus-talspiegel, > ng/ml) zeigte sich eine schwere pneumonie (pseudomonas), welche stationär behandelt wurde. es gab keine stationäre behandlung wegen cmv-infekten. die nierenfunktion war in allen patienten stabil (mittleres crea, , ± , ), außer in patienten, welche bereits vor der umstellung erhöhte kreatininwerte zeigten und in denen sich eine weitere erhöhung der kreatininwerte (+ , %) feststellen ließ. eine bei den meisten patienten auftretende hyperlipidämie konnte unter erhöhung der statintherapie eingestellt werden. in den routinemäßig durchgeführten endomyocard-biopsien fanden sich einen monat nach umstellung und danach keine akuten zellulären abstoßungen mit grad von > b nach dem ishlt-grading. schlussfolgerungen. certican erwies sich als sicher und verträglich, die umstellung auf das neue immunsuppressivum war in allen patienten komplikationslos. everolimus-talspiegel von ng/ml scheinen ausreichend, höhere spiegel könnten das infektionsrisiko erhöhen. bezüglich der nierenfunktion bleibt abzuwarten, wie sich ein cyclosporin-a-talspiegel von - ng/ml auswirkt. eine aussage bezüglich der cav steht zu diesem zeitpunkt noch aus. v jahre herztransplantation in wien (eine retrospektive über transplantationen) a. zuckermann empfänger-und spenderalter sind im laufe der jahre signifikant gestiegen (recipient age: , ± , vs. , ± , , p < , ; donor age, , ± , vs. , ± , , p < , ) . mehr patienten wurden präoperativ stationär aufgenommen ( % vs. %), jedoch hat sich die zahl der intensivpflichtigen patienten signifikant reduziert ( % vs. %). die zahl der patienten, die zur transplantation "gebridged" werden, ist ebenfalls massiv angestiegen ( vs. %, p < , ). innerhalb dieser gruppe hat die gruppe der patienten mit mechanischer herzunterstützung am stärksten zugenommen ( % vs. %, p < , ). pharmakologisches bridging ( % vs. %) und aicd ( % vs. %) blieben stabil. pharmakologisches bridging wird heutzutage vermehrt mit prostaglandinen und levusimendan als mit dopamin oder dobutamin durchgeführt. mehr patienten sind am herzen voroperiert ( % vs. %, p < , ), patienten warten länger auf die transplantation ( , ± , vs. , ± , , p < , ) . trotzdem hat sich die mortalität auf der warteliste stark verbessert ( , % vs. , %, p < , ), was ein klares zeichen der verbesserten Überbrückungsmaßnahmen ist. ischämiezeiten sind ebenso angestiegen ( , ± , vs. , ± , , p < , ) wie die liegezeiten auf der intensivstation ( , ± , vs. , ± , , p < , ). dies ist ein klares indiz dafür, dass heute ältere, kränkere und komplexere patienten transplantiert werden. diese veränderungen sind international bei allen zentren zu be-obachten. dies hat dazu geführt, dass mit der zunehmenden erfahrung und verbessertem Überleben die nachbeobachtungszeit der patienten stark gestiegen ist und damit die behandlung dieser patienten kostenintensiver geworden ist, was aber mit der guten lebensqualität der herztransplantierten patienten zu rechtfertigen ist. v neoplastic diseases after heart transplantation: a retrospective study d. kammerstätter, a. aliabadi, j. ankersmit, d. dunkler, g. wieselthaler, e. wolner, m. grimm, a. zuckermann klinische abteilung für herz-thoraxchirurgie, universitätsklinik für chirurgie, medizinische universität wien, wien, Österreich background. prolonged immunosuppression after solidorgan transplantation is associated with an increased risk for development of neoplasms. the purpose of this analysis was to investigate neoplasm incidence and outcome in patients with induction therapy. methods. cardiac recipients were included in this retrospective analysis. all patients received antibody induction therapy with either polyclonal-atg or monoclonal antibodies. neoplasms were devided into groups: ( ) skin cancer, ( ) ptld, ( ) other neoplasms. overall incidence of neoplasms, patient survival after diagnosis of neoplasms were analysed by the kaplan-meier method. results. a total of tumors were diagnosed at a mean follow-up of . ± . months after cardiac transplantation. freedom from neoplasms was . %, . %, and . % after -, -, and -year respectively. -year survival after diagnosis of tumor was . %. patients developed skin cancers after ± . months. -and -year survival after diagnosis was % and % respectively. there were tumor-related deaths in this group. patients developed ptld . ± . months after transplantation. -and -year survival was % and % with deaths associated with the neoplasm. in the third group, a total of patients were included. this group consisted of lung cancer (n = ), abdominal cancer (n = ), urogenital cancer (n = ), and other tumors (n = ). neoplasms were diagnosed at an average follow-up of . ± . months. -year and year survival was % and %. deaths were associated with tumor. conclusions. although all patients received antibody induction therapy, overall incidence of neoplasms was comparable to centers using no induction therapy. especially incidence of ptld was low. as long-term survival after cardiac transplantation increases steadily and the risk of cancer increases continuously, patients in long-term follow-up should be checked for malignant diseases on a routine basis. background. while the predictive value of n-terminal pro-b-type natriuretic peptide (nt pro-bnp) in nontransplant cardiac patients is well recognized, its value in heart transplantation (htx) is incompletely understood. certain graft factors (e.g., isolated diastolic dysfunction) may affect both, nt pro-bnp levels and peak exercise tolerance. we therefore hypothesized a relationship between these variables in long-term htx recipients. methods. we measured nt pro-bnp levels of htx patients before a symptom-limited upright bicycle exercise test was performed. graft function was stable in all patients and there were no signs of rejection. patients were divided according to a cut-off value of % exercise tolerance predicted normal into "low" and "normal" htx fitness groups. results. in patients ( m, f; ± years; . ± years posthtx; donor age, ± years; bmi, . ± . kg/m ; creatinine clearance, ± ml/min) peak exercise tolerance was "low" ( ± w), while in patients ( m, f; ± years; . ± . years posthtx; donor age, ± years; bmi, . ± . kg/m ; creatinine clearance, ± ml/min) it was "normal" ( ± w). in the "low" htx fitness group, nt pro-bnp levels were ± pg/ml, in the "normal" htx fitness group, ± pg/ml (p = . between groups). regression analysis of peak exercise tolerance, achieved percentage of exercise tolerance predicted "normal", and renal function with nt pro-bnp levels failed to demonstrate a significant relationship. conclusions. the findings confirm previous studies that nt pro-bnp levels are increased in asymptomatic long-term htx recipients. a larger sample size is warranted, however, to support the hypothesis that nt pro-bnp might be a useful indicator to predict physical fitness in these patients. calcineurin inhibitor therapy is an important cause of renal dysfunction in heart transplant patients. sirolimus (srl) is a novel immunosuppressive (is) drug without nephrotoxic side effects. however, cases of proteinuria associated with srl have been reported following renal transplantation. in cardiac transplantation the potential incidence of proteinuria is not known. long-term cardiac transplant patients (age, . ± . years) were switched from cyclosporine-based immunosuppression to a srl-based is . ± . years after transplantation. concomitant is consisted of mycophenolate-mofetil with or without steroids. two patients died and months post switch due to infection. both patients were dialysis dependent at time of death. one other patient was switched back to cni-based is due to interstitial nephritis. h collections of urine were performed in all patients before switch and at several time points post switch to measure proteinuria. proteinuria increased significantly from . ± . mg/dl (median, . ) pre switch to . ± . mg/dl (median, . ) post switch (p = . ). proteinuria of . - . mg/dl was seen in % of patients before switch and in % after switch. proteinuria of > . mg/dl was seen in % of patients before switch and in % after switch (n.s.). three patients developed severe proteinuria (> . mg/dl) after switch. one died on dialysis, one was switched back to cni and one still remains on srl. in conclusion, proteinuria may develop in cardiac transplant patients after switch to srl-based is. srl seems to have a potential adverse renal effect in these patients. srl should be used cautiously with close monitoring for proteinuria or increased renal dysfunction. v early growth-response factor- is involved in cellular injury of transplanted hearts background. we have shown a persistent mitochondrial pathology in patients with idiopathic dilative (dcm) but not ischemic (icm) cardiomyopathy following cardiac transplantation. early growth response factor (egr)- that is stimulated by cytokines and hypoxia is suggested to induce inflammation and tissue injury. whether egr- mediates the persistent cellular pathology in hearts transplanted to dcm patients is unknown. methods. egr- and hypoxia-inducible factor- (hif- ) gene expression was examined in left ventricular biopsies of explanted failing hearts in icm and dcm patients, as well as in donor grafts before reperfusion (control), at , , minutes after reperfusion, and at , , , , , posttransplant weeks, using real-time rt-pcr. hif- binding activity was examined using emsa. results. egr- myocardial gene expression was upregulated in dcm and icm (p < . ). hif- mrna levels were unchanged in both groups, whereas hif- binding activity was increased in icm only (p < . ) vs. controls. egr- and hif- myocardial expression increased during reperfusion in donor grafts (p < . ) vs. control hearts. in icm group, graft egr- and hif- expression returned to and remained at the baseline level of control hearts one week after transplantation. in contrast, in dcm group, egr- levels remained significantly upregulated during the follow-up period in transplanted hearts (p < . ), although hif- expression returned to the control baseline level one week after transplantation. conclusions. chronic hypoxia specifically triggers hif- binding activity in icm, and reperfusion upregulates egr- and hif- mrna expression in heart grafts. the persisting egr- overexpression in grafts transplanted to dcm patients could mediate mitochondrial impairment. targeting egr- overexpression that bypasses hif- might be beneficial to counteract acute reperfusion-induced injury, and the chronic cellular pathology in cardiac grafts transplanted to dcm patients. introduction. giant-cell myocarditits (gcm) is a rare and frequently fatal disorder of unknown origin that is defined histopathologically as diffuse myocardial necrosis with multinucleated giant cells in the absence of sarcoid-like granuloma. patients usually have ventricular arrhythmias or congestive heart failure. although a variety of systemic disorders have been seen in association with giant cell mycocarditis, most cases occur in previously healthy adults. conclusive diagnosis requires histologic analysis of myocardial tissue obtained by endomyocardial biopsy (emb). congestive heart failure (chf) is the most common cardiac presentation ( %), with sustained, refractory ventricular tachycardia. the case presented here is that of a -old-man suffering of gcm in whom an extracorporeal membrane oxygenation (ecmo) had to be implanted to overcome cardiogenic shock. antithymocyte gobuline (ratg, thymoglobuline, sangstat), respectively ciclosporine (inn: cyclosporine), mycophenolate and steroids were utilized to bridge the time to complete myocardial recovery. we are reporting the first successful implantation and bridging to myocardial regeneration in a patient suffering of gcm by means of ecmo and initiation of immunomodulating drugs including polyclonal ratg. clinical summary. a -year-old man was admitted to a public hospital because of thoracic pain, positive heart enzymes, and a highly pathologic electrocardiogramm. echocardiography demonstrated a pericardial effusion and reduced left ventricular function (lvf, ef %). performed angiography evidenced and a high-grade stenosis of the left anterior descending (lad) which led to stent implantation. despite stent placement, the clinical condition worsened leading to cardiogenic shock including incipient shock liver. in this clinical condition the patient was transferred to our institution. in addition, the patient developed malign ventricular tachycardia (lowen iv) and had to undergo repetitive defibrillation. in this clinical scenario it was decided to implant a femoral veno-arterial ecmo. the patient's metabolic data improved noticeably; however, due to repeated ventricular tachycardia, the patient had to be defibrillated multiple times (max. ×/d). to define exact cardiac pathology, we performed a heart biopsy with the pathology of gcm. immunosuppressive therapy with cyclosporine ( mg/d), mycophenolate and prednisolon ( mg/d) was initiated. in addition, rabbit atg (ratg, thymoglobuline, sangstat) at the dosage of mg/d was added to the therapy. this drug therapy was continued for days. of note is the fact that with initiation of ratg, cardiac fibrillation episodes abated immediately. routinely performed echo-cardiography (tee) revealed an improvement in ventricular function, and one week after ecmo support, we were able to wean the patient from extracorporeal support. moreover, a routine biopsy after days revealed complete remission of gcm in the heart tissue. an intracardial defibrillator (icd) was additionally implanted. three months after emergency admission to our department the patient was discharged. continuous medication of prednisolon mg/d, mycophenolat mofetil mg/d, and plavix mg/d was prescribed. month after initial event the patient describes nyha class i heart function and echocardiography reveals an moderate impairment of heart function (ef %). immunosuppressive with low-dose steroid and mycophenolate drug regimen is continued as the patient describes no side effects. discussion. this report adds to the available knowledge of giant-cell myocarditis by providing that (a) ecmo implantation is feasible if the patient is demonstrating acute haemodynamic deterioration because of biventricular dilation and medically intractable ventricular fibrillation and (b) after verified histological diagnosis of gcm immunemodulation with cyclosporine/mycophenolate with additional application of ratg is feasible with favourable outcome. in various studies, patients with gcm were treated with immunosuppression (cyclosporine, steroids, murine monoclonal antibodies [okt- ]) and even assist device as bridge to transplantation. heart transplantation has shown to be successful as method of treatment. autoimmune diseases and its mechanisms were suggested to be involved in the pathogenesis of gcm. most recently, a novel mechanism of action of immunoglobulin was proposed to be due to anti-inflammatory activities through the inhibitory fc receptos (fcrs). it has been suggested that t-cell-mediated autoimmune diseases is the result of inappropriate antigen presentation of either a self-antigen or an antigen with the capacity to mimic a self-antigen in the peripheral lymphoid tissues. relevant to this novel application of ratg, polyclonal suspensions like igm/g, ivig containing fc receptors and have been demonstrated to be beneficiary in autoimmune myocarditis. in an elegant study by shioji et al. ivig was highly effective in ameliorating experimental myocarditis. however it has to be mentioned that immunoglobulin treatment failed to ameliorate myocarditis. in respect to our patient suffering of rcm fc containing ratg provided remarkable clinical benefit. background. the criteria for liver donation have been widely extended due to the increasing waiting time and waiting list mortality. marginal donors provide an upcoming option to diminish the number of waiting patients. methods. the criteria for marginal were icu stay of > days, age of > years, bmi of > , alcohol or oral drug abuse, infection, hypernatremia (na, > mg/dl), high liver enzymes (ast, alt times the normal), liver parenchymal damage and metabolic diseases. from / until / our center reported donors, who fulfilled at least of the above criteria. results. all livers were transplanted either in standard or in piggyback technique with a cold ischemic period between minutes and hours. the mean recipient age was ( - ) years. livers were used for hu patients, for a retransplantation, were implanted in combination with kidneys, and organs were transplanted electively. primary diseases were cryptogenic cirrhosis, hepatocellular carcinoma, post-hepatitis cirrhosis, polycystic liver disease, scleroting bile ducts, hepatic artery thrombosis, and acute liver failure. month after transplantation, recipients were alive, died month after tx not transplant related. livers were implanted at our center in piggyback technique with retrograde reperfusion. patients were elective patients in good or moderate condition, patient was a hu patient suffering a hepatic artery thrombosis. the initial graft function was good (got, < mg/dl pod ) in , moderate (got, - mg/dl pod ) in , and delayed (got, > mg/dl pod ) in cases, all patients survived. conclusions. marginal livers are eligible for transplantation. delayed graft function has to be taken into account. hospital between january and december . we employed the local registry of the department of transplant surgery, where variables of all patients are routinely and prospectively recorded. primary outcome was initial graft function, secondary outcome was patient survival. results. cumulative number of marginal donor criteria was significantly and linearly associated with an increased rate of primary dysfunction (p = . ). in patients with more than three cumulative marginal donor criteria the rate of primary dysfunction was percent. patient survival was not influenced by the cumulative number of donor criteria (log-rank test, p = . ). independent marginal donor criteria to predict primary dysfunction were cold ischemia time of > hours (or, . ; % ci, . to . ) and donor peak serum sodiumof > meq/l (or, . ; % ci, . to . ), as assessed in a multivariate regression model. conclusions. the use of marginal liver donors with more than three marginal donor criteria shows deleterious effects on initial graft function. noteworthy, patient survival was not associated with marginal donor criteria, which may be explained by early and successful retransplantation of liver recipients with primary nonfunction. ergebnisse. alle transplantate zeigten eine gute initiale organfunktion, der transaminasenverlauf und das gesamtbilirubin, gemessen am . postoperativen tag, am . postoperativen tag und bei entlassung (mittelwerte mit standardabweichung) waren wie folgt: got u/l (± ), u/l (± ), u/l (± ); gpt u/l (± ), u/l (± ), u/l (± ); ggt u/l (± ), u/l (± ), u/l (± ); gesamtbilirubin , mg/dl (± , ), , mg/dl (± , ), , mg/dl (± , ) . background. meld score is a useful tool in predicting mortality in patients awaiting liver transplantation. its capacity to predict patient survival seems to be relatively poor and is still discussed controversially. the purpose of the study was to analyse the impact of alterations of the meld score during waiting time on the posttransplant survival rate. additionally, the impact of donor quality on posttransplant survival was investigated. methods. adult patients were transplanted between and because of end stage liver disease without malignancy. the meld scores at time of listing (meld on) and of transplantation (meld tx) were gathered. additionally the delta-meld was calculated from listing to transplantation. results. a high meld tx showed only a trend to poorer survival. patients who died within the st year after transplantation showed a significant increase in the meld score during waiting time (p < . ). patients with a delta-meld higher than during waiting time had a % -year mortality after transplantation, patients with a meld increase not higher than had only a . % -year mortality (p < . ). patients with a meld score higher than who received a marginal graft showed a trend to poorer posttransplant survival. conclusions. patients with a substantial increase of the meld score during waiting time had a significantly poorer year posttransplant survival. in contrast, the meld score at time of listing or transplantation had no impact on the posttransplant survival rate. the use of marginal grafts in patients with a higher meld score has to be evaluated carefully. there is no significant difference of serum sodium levels in ltx candidates with or without ascites crease in serum creatinine is a late event in patients with ascites and is not directly reflected within the meld formula. for this purpose we compared patients who died on the waiting list with patients who finally were transplanted. the impact of serum sodium and ascites on death on the waiting list was evaluated. methods. from to , adult patients with end-stage liver disease were listed for orthotopic liver transplantation (olt). only patients without hepatoma who died on the waiting list ( patients) or were finally transplanted ( patients) were evaluated. in addition to the meld score, the serum sodium and the ascites were investigated at time of listing and of coming off the list (transplantation or death). results. transplanted patients had a significantly lower meld on (p < . ) than the patients who died while on the waiting list. patients who died while on the waiting list had a significant increase in the meld score during waiting time (p < . ). patients who underwent transplantation showed a stable meld score during their waiting time. refractory ascites and spontaneous bacterial infection were evaluated as independent risk factors for death on the waiting list as well as the meld on. . % of the patients ( of patients) who died on the waiting list were suffering from ascites, in contrast to only . % of the transplanted patients ( of patients). there was no significant difference in the mean meld on between the patients who were suffering from ascites and those who were not (p = . ). nor was any significant difference found in the meld off (p = . ). the serum sodium of patients suffering from ascites showed no significant difference to those who showed no signs of ascites. conclusions. ascites was evaluated as independent risk factor for death on the waiting list. no significant difference in the serum sodium levels were found between patients suffering from ascites or not. therefore complications of portal hypertension should be treated adequately and rigorously, especially in patients with lower meld scores. renal failure is an established risk factor for impaired patient outcome after orthotopic liver transplantation (olt). as the endothelin pathway is known to be involved in the development of acute renal failure (arf), we designed a study to clarify its role in arf following olt. consecutive patients with intact kidney function scheduled for their first olt were prospectively studied. plasma big endothelin- (et- ) levels were measured before surgery, after graft reperfusion, and on the first and second postoperative days. according to postoperative gfr, patients were assigned to the acute renal dysfunction group (ardf) and the non-ardf group. each patient's gfr was estimated according to the four variable formula used in the modification of diet in renal disease before surgery, daily within the first postoperative week, and at , , and months after surgery. postoperative mean big et- lev-els correlated significantly with the maximum percent decrease of gfr within days after olt (p < . ). the proportion of patients who developed ardf was significantly correlated to mean postoperative big et- quartiles (p < . ). in the ardf group, the percent decrease of gfr within months was significantly higher (p < . ) as compared to the non-ardf group. in conclusion, patients who develop acute renal dysfunction immediately after olt do not fully recover to baseline regarding long-term kidney function. short-term gfr was significantly correlated with postoperative big et- plasma levels, suggesting renal dysfunction is mediated by the activated endothelin system. background. with improved survival of liver transplant recipients, chronic renal failure has become an important cause of morbidity and is associated with a high mortality. serum creatinine is widely used as marker for renal function, but it depends on various nonrenal factors and major changes will occur late in the course of progressive renal impairment. we evaluated cystatin c and urine microscopy for detection of renal dysfunction in patients after liver transplantation. methods. from november , liver transplant recipients at various intervals from liver transplantation were included to our follow-up. every three months we investigated serum creatinine, urea, renal creatinine clearance and cystatin c as marker for renal function. furthermore urinary sediment was examined by urinary test, automated urinary sediment analyser, and urine microscopy. in patients with reference to renal deterioration we tried to decrease immunosuppressive therapy, to optimize the blood pressure, and to discontinue nephrotoxic medication. infections were detected early by urine microscopy and treated, even when there was no clinical appearance and the urinary test was negative. results. the results of our study showed that concerning the renal function, cystatin c is more sensitive than creatinine and creatinine clearance. the microscopy of the urine sediment showed the highest sensitivity compared with the other methods. concerning damages of the kidney, urine microscopy offered the best possibility to identify the etiology. during the follow-up and after adequate and early therapy, fifteen patients ( . %) showed an amelioration of renal function after a few months. in patients ( . %) there was a marked deterioration. two of them had to receive a higher dose of immunosuppressive therapy and one had an infection which was treated with nephrotoxic medication. conclusions. the early identification of renal failure and its etiology are necessary in patients after liver transplantation. the results of our study confirmed cystatin c as early prognostic marker for patients with renal dysfunction. in combination with urine microscopy, renal dysfunction could be detected in time and renal function could be protected with an adequate therapy. background. hcv-infected patients and their grafts have short-term survival rates similiar to other indications. recent data on long-term outcome are contradictory, showing a trend towards poorer outcome in patients transplanted for hcv cirrhosis. in this study we present a retrospective analysis of our experience with patients who underwent liver transplantation (lt) due to hcv-associated end stage liver disease. methods. patients' charts were reviewed for survival, histologically defined hcv recurrence, genotype, presence of cirrhosis, donor and recipient age as well as type of immunosuppression (cyclosporine and tacrolimus; azathioprine and mycophenolate mofetil). survival was analysed by the kaplan-meier procedure, the influence of baseline variables was analysed by binary logistic regression. results. between and october , patients were transplanted for hcv-related cirrhosis. ten pts. received one and pts. two relts. in ( %) pts. recurrent hepatits c was the cause for relt, in vascular and/or biliary complications. hepatitis b coinfection was present in patients. median follow-up was months (range, . - ). the overall, -, -, -, -, and -year survival rates were %, %, %, %, and %, respectively, which were comparable to other indications. the probability of recurrent hcv was %, %, %, %, and % after , , , , and years, respectively, post lt. nine ( . %) pts. developed cirrhosis after a median of months ( - ). hcv recurrence did not negatively influence patient and graft survival. concerning genotype, cmv status, presence of hcc before lt, rejection episodes, immunosuppression, donor and recipient age only immunosuppression had a significant effect on survival. in cyclosporine-treated patients (lt after ) -, -, and -year survival rates were %, %, % compared to %, %, %, respectively, for tacrolimus-based regimens (p = . , log rank test). conclusions. on the basis of our data, the overall survival of hcv transplanted patients were similiar to other indications. recurrent hcv infection did not influence patient and graft survival. immunosuppression may have an impact on survival in hcv-positive recipients, but optimal regimens need to be better defined by prospective studies. the advent of highly sensitive molecular techniques has revealed the possible persistence of hepatitis b virus (hbv) genomes in hbsag-negative patients with or without serologic markers of previous infection, a state called occult hbv infection. the highest prevalence of such infection has been shown in patients infected with hepatitis c virus (hcv). some studies suggested that occult hbv infection might favor or accelerate the progression of hcv infection towards cirrhosis. hcv infection recurs almost in all patients after liver transplantation (lt). about - % of lt patients develop a fibrosing cholestatic hcv recurrence, which is associated with a very poor outcome. no specific risk factor for this pattern of recurrence has been described so far. the aim of this study was to determine the prevalence of occult hbv infection in patients presenting with fibrosing cholestatic hcv recurrence after lt. between and , hcv patients ( m; f) underwent lt at our institution. the mean follow-up was months (range, - months). the diagnosis of recurrence was based on biochemical and histologic parameters. genotype was predominant ( %), followed by ( %), ( %), and ( %). eleven patients ( . %; m, f) developed a fibrosing cholestatic pattern of recurrence characterized by highly elevated cholestatic parameters and typical histologic findings. also in this group, genotype was predominant (n = ), three had type , and one type . serum hbv dna was tested with the taqman test (roche, austria). five patients were hbsag negative, whereas six had serologic markers (antihbc positive). interestingly, hbv dna could not be detected in the sera of any of these patients with fibrosing cholestatic hcv recurrence. the actuarial -, -, -, and -year survival rates of all hcv patients were %, %, %, and %. hcv recurrence did not show a negative impact on patient and graft survival; however, the outcome of the patients with the fibrosing cholestatic pattern was less favorable. seven out of patients died due to hcv recurrence, one patient had to be retransplanted secondary to recurrent disease. only three patients are alive with a functioning first allograft. this study indicates that occult hbv infection is not associated with fibrosing cholestatic hepatitis c recurrence after lt. background. the use of grafts from hepatitis b core antibody (anti-hbcab)-positive donors for liver transplantation (lt) is associated with the risk of de novo hepatitis b. patients who test positive for anti-hbcab pretransplant are also theoretically at risk to develop graft hepatitis b. methods. the outcome of consecutive lts performed in individuals between and was retrospectively analyzed with regard to the presence of anti-hbcab in donors and recipients. patients with hepatitis b and recipients of known anti-hbcab-positive grafts received hbig/lamivudine prophylaxis. results. a total of recipients ( %) tested positive for anti-hbcab including patients ( %) with hbv-associated cirrhosis and three patients with fulminant hepatitis b. a total of allografts from anti-hbcab-positive donors were utilized, of those ( %) were allocated to anti-hbcab-positive recipients. anti-hbcab-positive recipients were significantly more likely to have concomitant hcv ( % vs. %, p < . ) and hepatocellular carcinoma ( % vs. %, p < . ). anti-hbcab-positive donors were more frequently non-caucasian ( vs. %, p < . ) and cmv seropositive ( % vs. %, p < . ). survival of anti-hbcab-positive individuals and recipients of allografts from anti-hbcab-positive donors did not differ from their anti-hbcab-negative counterparts. there were no reported cases of recurrent hepatitis b in anti-hbcab recipients or patients receiving lt for hbv-associated liver disease. three cases of de novo acute posttransplant hepatitis b were identified, one being acquired during unprotected intercourse and two being transmitted through the graft. the two with graft-transmitted disease were anti-hbcab negative, treated initially with lamivudine and were switched to adefovir due to the emergence of ymdd mutants. conclusions. the frequency of anti-hbcab-positive recipients in our series was higher than expected. these individuals seem at minimal risk for posttransplant hepatitis b. recurrence of hbv after lt in the setting of hbig/lamivudine prophylaxis is extremely rare with -year median follow-up. the risk of transmission of hbv through anti-hbcab-positive livers despite prophylaxis cannot be neglected and the emergence of an ymdd mutant is of particular concern. anti-hbcab-positive grafts should be preferably given to anti-hbcab-positive recipients. v the response to preoperative transarterial chemoembolisation predicts outcome of patients with hepatocellular carcinoma after liver transplantation division of gastroenterology and hepatology, department of internal medicine, medical unversity of innsbruck, innsbruck, austria liver transplantation (lt) is the only curative therapy for patients with early-stage hepatocellular carcinoma (hcc). the impact of prelt chemoembolisation (ce) on patient survival and incidence of hcc recurrence has been controversially discussed and remains undetermined. the aim of this study was to evaluate the efficacy of ce prior to lt in hcc patients with regard to tumor recurrence and patient outcome. between and , hcc patients ( m; f) underwent lt at our institution. the underlying liver disease was viral hepatitis in (hcv , hbv ), alcoholic liver cirrhosis in , and other diseases in patients. according to child-pugh classification, patients presented with child a, with stage b and with stage c. hccs were diagnosed according to the easl guidelines. on the basis of prelt radiology, patients were diagnosed with stage i, stage ii, stage iii, and stage iv according to the modified uicc criteria. ce was performed in patients prior to lt. in patients no treatment was performed prior to lt. patients underwent multiple cycles of ce (mean, . ce/patient). ce response prior to ce was assessed by ct scan. on explant histology, complete response with no evidence of vital tumor was found in of ( %) patients, ( %) patients showed a partial remission (tumor necrosis, > %), and ( %) patients showed a poor response or even progression. the intention-totreat analysis showed that patients with early-stage hcc showed an excellent survival with a -, -, and -year rate of %, % and %, respectively. ce prior to lt had no positive effect on overall patient outcome and rate of recurrence. however, patients with complete response to ce, on the basis of both pre lt and post lt histology, had a significantly bet-ter long-term survival ( -, -, and -year rates of %, %, and %) and rate of recurrence compared to those with partial or no response, but only in patients within milan and not san francisco criteria. the -, -, and -years survival rates of patients with advanced hcc were %, %, and %. hcc recurrence was found in patients, of presented with advanced stage (iii and iv). only had undergone ce prior to lt, and of those were nonresponders to ce. this study indicates that response to prelt ce may predict the outcome of hcc patients after lt. patients with early-stage hcc, who responded to ce pre lt, showed an excellent outcome with -and -year survival rates around %. patients with early-stage hcc and only partial or no response to ce had a higher risk of recurrence of hcc after lt, but outcome was still favorable compared with advanced-stage tumors regardless of ce response. the roles of the regenerative factors hepatocyte growth factor (hgf), transforming growth factor a (tgf-a), and of vascular endothelial growth factor (vegf) were described in the context of hypertrophy and regeneration after liver resection but not well known in the transplantation situation. in the recipients of consecutive liver transplantations with a graft survival of > weeks, the factors hgf, tgf-a and vegf were determined postoperatively (day , , , , , ) by an el-isa in the serum and correlated to graft survival (kaplan-meier). the median concentrations of hgf were constant in total during the observation period (day , pg/ml; day , pg/ml; day , pg/ml). an individual increase to levels above pg/ml in the middle of the observation period correlated to a significantly decreased one-year graft survival ( % vs. %). similar was the course of tgfa. an increase from the median concentration of pg/ml to levels above pg/ml was observed in the context of a decreased primary function. regarding vegf, an almost linear increase of the concentration from pg/ml via pg/ml to pg/ml (day , , and ) was observed. here it became obvious, that an extensive increase of the vegf concentration correlated to a good transplant function. under the premise that the systemically determined concentrations were in relevant correlation to the secretion and thus to the local concentration in the liver, it can be concluded that vascular regeneration induced by vegf substantially contributes to graft survival, whereas a temporal increase of hgf and tgfa rather has to be interpreted as an indicator of an injured graft with a decreased functional prognosis. in total there are about patients on dialysis in bh and percentage of transplanted is . . the aim of the paper is to analyse survival of grafts, patients, and grafts and patients over a -year period of living-related kidney transplantation history at university medical center tuzla. the results obtained were compared with the results of the centers with great number of transplants. methods. recipients and donors as family members were admitted after informative discussions and after the test results had been obtained from primary health care laboratories. the protocol was the standard one, in accordance with the recommendations of esot and eurotransplant. donors were tested first and then recipients. lumbotomy was done for nephrectomy. colins solution was used for kidney rinsing and perfusion, reconstruction of blood vessels was done as well as kidney biopsy. then, kidneys were stored at + °c and grafts implantations were done on the contralateral iliac fossa. terminolateral anastomosis between external iliac artery and vein was done. implantation of ureters was done by modified lich-gregore technic, with or without dj stent. all patients received basic immunosuppressive protocol. the peculiarity was introduction of basiliximab (simulect) in therapy on the first and fourth postoperative day. descriptive statistics was done using spss software for windows . . survival is presented by kaplan-meier curve. results are shown as means with standard deviations (sd). results. the first living-related kidney transplantation was done at university medical center tuzla on . . . . until . . . as many as transplants have been done. donors were related to recipients as follows (in parentheses, mean age [years] with sd): mother, n = ( . ± . ); father, n = ( . ± . ); sister, ( . ± . ); brother, n = ( . ± . ); others, n = ( . ± . ). table shows mean dtpa clearance rate in donors. average separate dtpa clearance rates were within normal limits. as a rule, left-side donor nephrectomy was done; and in five cases, right-side nephrectomy. three donors showed borderline dtpa clearance rates of about ml/min. bladder neck sclerosis was found in one patient and the air expansion of the bladder had to be done in the other one, at least to achieve its minimal capacity. two renal arteries were found in patients ( mothers, father, sister), two arteries were found with left kidney transplantation. termino-lateral anastomosis was done in cases. the average age of the donors was . ± . years. as many as donors were over years old in our transplants, which reflected in our results. the average donor age of males and females was . ± . years. the data on hypertension before transplants were not reliable. the average hemodialysis time was . ± . months. the most common recipients' diseases were chronic glomerulonephritis, pyelonephritis, interstitial nephritis, nondefined renal disease, diabetes mellitus, systemic lupus, vesico-ureteral reflux. there were only biopsies done before transplantation so that their histological background was known. the average serum creatinine after years is . ± . µmol/l. cumulative graft survival after years is shown by kaplan-meier curve: graft survival, . %; patients survival, . %; graft and patients survival, . %. conclusions. the results on year living-related kidney transplantation at university medical center tuzla, bosnia and herzegovina, are similar or identical to the results of the developed centers in the world. the existing program has got to be improved, especially with respect to the donor selection from the standpoint of biological and chronological age. the experience gained so far is the basis for the development of cadaveric kidney transplantation in bosnia and herzegovina. background. shortage of available organs has increased the demand for living kidney donation. whereas donation for relatives is well accepted, there remains some controversy in the setting of emotionally related donation. there might be a survival benefit for grafts donated by relatives due to better hla matching. a retrospective analysis of living donor kidney transplants which were performed at the university hospital of innsbruck was made. we divided the transplants in two groups according to the date of transplant: group ( - ), transplants; group ( - ) , transplants. the time period from until was analyzed in detail and data additionally compared to a cohort of cadaveric renal transplants. during these eleven years, five liv-ing donations were carried out in patients who were not eligible to receive a cadaveric graft within eurotransplant. results. overall there were lrt ( siblings, parents, off-springs) and ert ( spouses, others). mean donor age was years (range, - ); mean recipient age was (range, - ) years. in group only transplants were emotionally related ( %), whereas in group the proportion increased to %. graft survival of living donated kidneys was better when compared with cadaveric kidneys ( % versus % at one and % and % at five years), but the difference did not reach statistical significance (p = . ). lrts and erts produced equal outcome. overall graft loss rate after a median follow-up of six (range, . - . ) years was % (lrts) vs. % (erts). the rejection rate was slightly higher in the lrt group with % vs. % (p > . , n.s.). ten living donated grafts were lost and seven recipients of a living donated graft died. causes of death were cardiac ischemia (n = ), pulmonary embolism (n = ), fungal infections (n = ), suicide (n = ), and causes were not specified. conclusions. the frequency of living-related and unrelated kidney donation has increased during the past two decades at our institution. equal results for cadaveric grafts were achieved when compared to lrt and erl. background. the s proteasome plays an important role in the nonlysosomal pathway of intracellular protein degradation and apoptosis, thus being a possible marker for ischemic and reperfusion injuries. the aim of this study was to monitor the proteasome levels in patients receiving kidney transplants to detect a relationship with the return of renal function. methods. we examined patients with end stage renal disease, receiving kidney transplants: blood samples were collected intraoperatively and postoperatively on consecutive days and s proteasome levels were measured for each sample with a sandwich elisa as described by dutaud et al. anesthesia and immunosuppressive medications were standardized, creatinine clearance and urine output were assessed daily on a routine basis. results. patients who had no adequate urine output ( ± ml) after the th postoperative day had significantly higher proteasome levels intraoperatively than patients with high urine output ( ± ml; . ± . µg/ml low vs. . ± . µg/ml high urine output, means with standard deviations, p = . ). this difference in proteasome levels seemed to even out during the follow-up period. conclusions. patients with impaired renal function after kidney transplant had significantly higher proteasome levels intraoperatively. a higher plasma level of proteasomes intraoperatively may therefore be a negative prognostic marker for postoperative return of renal function of the transplanted kidney. background. dendritic cells (dcs) are the most potent antigen-presenting cells and are pivotal for initiating allograft immunity. recently, however, particular dc subsets have also been implicated in allogeneic tolerance induction. campath- h (alemtuzumab) is a novel t-cell-depleting antibody that is currently under investigation for the use in allogeneic organ transplantation and may confer tolerogenic properties. here we study the effect of alemtuzumab on peripheral dc subsets in kidney transplant patients under fk monotherapy in comparison to patients under conventional triple therapy. methods. patients receiving their first renal allograft were recruited within the tacam trial and randomly assigned to receive either alemtuzumab as induction agent followed by fk monotherapy (n = ) or to receive conventional immunosupression consisting of fk , mycophenolate mofetil and steroids (n = ). absolute numbers of peripheral blood dcs and their different subpopulations were assessed by four-colour, single-platform fluorocytometry at the day before and , , , and weeks after the transplant procedure, respectively. peripheral dcs were identified as hla-dr + and lineage-negative cell population. the respective dc subpopulations were cd c + dcs (myeloid dcs or dc ), cd + dcs (plasmacytoid dcs or dc ), cd c + and further, bdca + and bdca + dcs. results. induction with campath- h led to a strong and sustained reduction of the total number of peripheral dcs compared to controls. while the absolute number of peripheral dcs in control patients recovered within months after transplantation, campath- h-treated patients exhibited a profound reduction of their circulating dcs. interestingly, a prominent shift of the ratio of myeloid to plasmacytoid dc subsets (dc /dc ratio) was observed as early as one month after transplantation in campath- h-treated patients. conclusions. employment of campath- h as induction therapy in renal transplant patients is associated with a peculiar alteration of the peripheral dc repertoire. since plasmacytoid dcs have been linked to tolerance induction, the presented data suggest that the use of campath- h in solidorgan transplantation creates an opportunity to safely introduce novel strategies to achieve alloantigen-specific hyporesponsiveness. background. detection of c complement split product c d in peritubular capillaries represents a valuable diagnostic marker for antibody-mediated rejection (amr). numerous studies have demonstrated inferior allograft function and survival in c d-positive as compared with c d-negative kidney allograft recipients. however, anecdotal data implicate that in selected cases stable long-term function can be maintained despite detectable c d deposits. as recently dicussed in the literature, c d deposits could also reflect a state of graft acceptance (accommodation), rather than rejection. aim. the objective of this study was to investigate individual clinical outcomes in a large cohort of c d-positive kidney transplant recipients. our emphasis was thereby to identify and characterize a subpopulation of c d-positive recipients with only mild graft dysfunction and stable long-term graft function. methods. this retrospective analysis focused on out of adult kidney transplant recipients (transplantation between between and ) included on the basis of a positive c d result early after transplantation. results. at the time of c d-positive biopsy (median, days post-tx) median serum creatinine was mg/dl (range, . to mg/dl) with % of the patients dialysis-dependent. according to our local standard, patients with severe graft dysfunction (n = ) were subject to immunoadsorption treatment (ia). in another highly sensitized recipients, c d-positive graft dysfunction was diagnosed during or after pre-emptive peritransplant ia. furthermore, intense treatment included antilymphocyte antibody therapy ( %) and/or high-dose steroids in a high proportion of c d-positive recipients ( %). analyzing all c d-positive recipients, -month post-biopsy se-rum creatinine was . mg/dl ( % of the patients dialysis-dependent). -year graft and patient survival was % (serum creatinine, . mg/dl). in a subsequent subanalysis we focused on twelve patients (two were biopsied under ia/atg induction) with only mild to moderate graft dysfunction (serum creatinine, . to . mg/dl) at the time of c d-positive biopsy (banff borderline lesion in five, and banff i rejection in two biopsies). within this patient subgroup, we were able to identify five recipients in whom stable long-term graft function could be achieved following steroid bolus therapy only, without further therapeutic measures. in this particular subgroup excellent -year allograft function (serum creatinine levels between . to . mg/dl) was observed. conclusions. our results demonstrate that in the majority of patients peritubular capillary c d deposits are associated with severe graft dysfunction necessitating aggressive treatment. nevertheless, in a small subgroup of recipients stable graft function for a long period of time can be achieved without intense therapy despite capillary c d deposition in biopsy. background. combined kidney pancreas transplantation (ptx) evolved as excellent treatment for diabetic nephropathy with infections remaining common and serious complications. methods. consecutive enteric drained ptxs performed from to were retrospectively analyzed with regard to bloodstream infection. immunosuppression consisted of antithymocyteglobuline induction, tacrolimus, mycophenolic acid, and steroids for the majority of cases. standard perioperative antimicrobial prophylaxis consisted of pipercillin/tazobactam in combination with ciprofloxacin and fluconazole. results. one-year patient, pancreas and kidney graft survival were . %, . %, and . %, surgical complication rate was %, rejection rate %, and rate of infection %. in total, sepsis episodes were diagnosed in patients ( %) with a median onset on day (range, - ) post transplant. sepsis source was intra-abdominal infection (iai) (n = ), a contaminated central venous line (n = ), wound infection (n = ), urinary tract infection (n = ), and graft transmitted (n = ). nine patients ( %) experienced multiple sepsis episodes. overall, pathogens (iai sepsis, ; line sepsis, ; others, ) were isolated from blood. gram-positive cocci accounted for isolates ( %): coagulase negative staphylococci (n = [ %]) (nine multiresistant), staphylococcus aureus (n = [ %]) (four multiresistant), enterococci (n = [ %]) (one e. faecium). gram-negative rods were cultured in twelve cases ( %). patients with blood borne infection had a two-year pancreas graft survival of . % versus . % for those without sepsis (p = . ), patient survival was not affected. conclusions. sepsis remains a serious complication after ptx with significantly reduced graft but not patient survival. the most common source is iai. background. mobilized allogeneic peripheral blood stem cells (pbsc) are increasingly used as graft source instead of bone marrow. although the short-term safety profile of recombinant human granulocyte colony-stimulated factor (rhg-csf) seems acceptable, minimal data exist regarding long-term safety. methods. we reviewed data of allogeneic pbsc donors (siblings, n = ; unrelated donors, n = ) with respect to side effects of rhg-csf administration, adverse events of leukapheresis and late effects. written informed consent was signed before pbsc mobilization and collection. donors (m/f, / ) had a median age of years (range, - ). they received rhg-csf at a dose of times µg/kg of body weight a day for consecutive days starting on day . routinely, pbsc collection was started on day . on condition that donors' white cell count exceeded . /µl or cd -positive cells exceeded /µl, pbsc harvest was started on day . rhg-csf was administered until end of the apheresis period unless white blood cell count did not exceed . /µl. pbsc were collected with a continuous-flow blood cell separator processing - . times total blood volume. citrate was used as anticoagulation and in the majority of donors a continuous calcium infusion ( . mmol [ . mg] of calcium per h) was given. reinfusion of autologous platelet-rich plasma from pbsc collection was performed in donors with post-donation thrombocytopenia of < g/liter if further collection were necessary or in donors with platelet counts of < g/liter, respectively. pbsc harvest procedures were repeated until the predicted cd + cell yield of × /kg of body weight of recipient was collected. however, not more than consecutive collections were performed. flowcytometric analyses were performed using a becton-dickinson facs-scan or facs-calibur, respectively. for follow-up we assessed peripheral blood counts, electrolytes, lactic dehydrogenase (ldh), alkaline phosphatase (ap), total protein, albumin, on days , , , , and then yearly for years. donors received a questionnaire for evaluation of medical history and quality of life, results and evaluation are pending. results. the main adverse events related to rhg-csf administration were bone pain ( of , %), myalgia ( of , %), headache ( of , %), fatigue ( of , %), sleep disturbance ( of , %) and were rated as moderate to severe by % of the donors. due to continuous calcium infusion, incidence of citrate toxcicity was low ( of , %) and consisted only of mild paraesthesia. in of ( %) donors post donation platelet count decreased below threshold and required reinfusion of autologous rich plasma. eighty-eight of donors ( %) were lost for follow-up. eighty-two were sibling ( of , %) and ( of , %) were matched unrelated donors, respectively. from the remaining donors, unrelated donors ( of , %) had a median of check-ups (range, - ) during the first days (median; range, day to years) after donation, whereas siblings ( of , %) had a median of check-ups (range, - ) during the first days (median; range, day to years), respectively. two donors ( %), both siblings, were hospitalized within weeks after donation due to severe enteritis and subarachnoidal haemorrhage. the latter donor never had platelet counts below g/liter after pbsc donation and recovered without neurological deficit. follow-up of peripheral blood counts showed a loss of platelets during donation and early post-donation period and a decrease of white blood cells days after donation, both returning to normal within days after, respectively. ldh and ap showed a significant increase during pbsc mobilization, they returned to normal within week after donation, other blood parameters remained unaffected. conclusions. due to the fact that we observed hospitalization of donors within days after pbsc collection, a close monitoring of donors in the early post-donation period seems advisable. although reported anomalies in medical history of donors beyond days after pbsc harvest could not be associated with rhg-csf administration, a regular followup for at least years should be considered. with the particular focus on donor safety, a standardized approach to data collection of follow-ups to monitor short-and long-term effects in all centers should be established. background. donor-recipient sex mismatch is an established risk factor for poor outcome after allogeneic myeloablative hematopoietic stem cell transplantation (hsct). the risk of transplant-related mortality (trm) due to graft-versushost disease (gvhd) is higher in male recipients of female stem cells compared with female patients receiving a graft from a female donor. with longer follow-up, however, the graft-versus-leukemia (gvl) effect due to hy minor histocompatibility antigen mismatch may predominate. the contribution of donor-patient sex on outcome after nonmyeloablative hsct, however, has not been examined in detail yet. methods. we therefore analyzed a single-center cohort of high-risk patients transplanted with a related or unrelated stem cell graft after nonmyeloablative conditioning for outcome (acute and chronic gvhd, trm, relapse, and survival). results. of the patients, male patients received a graft from a female donor, males a graft from a male donor. sixteen female donors were transplanted with a male donor and with a female donor. around % of the patients with a sex-mismatched donor received stem cells with an hla mismatch compared to % of the patients without sex-mis-stammzellen matched donor, the other clinical differences were similar between all groups. the highest cumulative incidence for acute and chronic gvhd was detected in male patients receiving a stem cell graft from a male donor ( . % and . %, respectively). the highest relapse incidence ( . %) was detected in male patients transplanted with a female donor. this was borderline significant (p = . ) to female patients receiving a female graft ( % relapse incidence) and argues against an effective anti-hy response in this patient cohort. the mean cumulative incidence for trm was . %; however, female recipients receiving a graft from a female donor displayed an unexpectedly high incidence for trm ( . %) which could not be explained by clinical characteristics. the overall survival of % . years after transplant in this group, however, was not different from male patients receiving a female graft ( . % at . years after transplant). the overall survival from male patients with a male donor was slightly lower ( % at . years after transplant) compared with female patients transplanted with male stem cells ( . % at . years after transplant). conclusions. these data, analyzed in a small cohort of patients, show that a sex mismatch between patient and donor may have a negative impact also on outcome after nonmyeloablative hsct. however, studies with larger and homogeneous cohorts have to be performed to prove these findings. between and , patients with chronic lymphoid leukemia (cll), binet stage b or c (n = ) or a with risk factors (n = ) with a median age of (range, - ) years underwent autologous (n = ) or allogeneic (n = ) hematopoietic stem cell transplantation (hsct) at the medical university of vienna. the median time from diagnosis to hsct was (range, - ) months. eleven patients underwent autologous stem cell transplantation (asct) in partial remission (n = ) or complete remission (cr) (n = ) and received bcell-depleted peripheral blood stem cell (pbsc) grafts. nine patients with refractory disease (n = ) or chemosensitive relapse (n = ) underwent allogeneic hsct with unmanipulated bone marrow (n = ) or pbsc (n = ) from sibling (n = ) or unrelated (n = ) donors. in the majority, conditioning therapy consisted of total-body irradiation (tbi) of - . gy and cyclophosphamide. three patients underwent reduced-intensity conditioning (ric) with fludarabine and tbi of gy. graft-versus-host disease (gvhd) prophylaxis consisted mainly of cyclosporine (csa) and methotrexate for myeloablative and csa and mycophenolate mofetil for ric-hsct. complete clinical remission was attained in patients ( %) after asct and in ( %) after allogeneic hsct. molecular remission assessed by immunoglobulin heavy chain gene (igh) rearrangement pcr was attained in patients after asct and after allogeneic hsct. in seven patients we noticed persistence of the igh rearrangement, six of these patients died of disease progression or relapse - months after asct (n = ) or allogeneic hsct (n = ). after a median follow-up of (range, - ) months, nine autologous ( %) and four allogeneic ( %) graft recipients are alive and patients (asct, n = ; allogeneic hsct, n = ) are in clinical and molecular remission. the median time to clinical relapse was (range, - ) months. treatment-related death occurred only in one patient months after myeloablative hsct. probability of overall survival is % after asct and % after allogeneic hsct. in summary, all cll patients with long-term cr after asct and allogeneic hsct also attained sustained molecular remission of the igh rearrangement. . the incidences of the observed genotypes in this small group of patients and donors tested were in the range as published: il with % c/c, % c/a, % a/a; nod with % recipient or donor; % recipient and donor, and mpo with % g/g, % g/a, % a/a. since the patient population was very heterogeneous concerning diagnosis ( aml, all, nhl, cml, mds, saa, solid tumors, mm, omf, cll, and et), course of disease, and condition regimen, patients were divided into four groups for evaluation: with aml and identical high-dose induction therapy (bu/cy), with reduced condition regimen, with unrelated donors, and others. concerning all patients, of them ( %) died through trm ( aml, reduced, others), but none of them was at high risk determined by nod polymorphism (mutated donor and recipient) and only one determined by mpo a/a genotype (aml). four patients ( reduced, others) with nod mutations in donor and recipient dna did not die from trm. twenty-three patients developed severe (grade or ) acute gvhd ( aml, reduced, others), of them had il c/c genotype ( aml, reduced, others) and in three patients nod was mutated in donor and/or recipient dna ( aml, others). on the other hand, patients with il c/c genotype and patients with nod mutations in donor and/or recipient dna developed no or mild gvhd only. in conclusion, so far we were not able to find a correlation between gvhd/trm risks and polymorphisms of il , nod , and mpo. this might be due to the small number and the heterogeneity of the patients; however, a panel of additional snps may increase the accuracy of risk assessment prior to allogeneic sct. background. allogeneic stem cell transplantation is a curative therapy for patients with lymphoproliferative disorders as a result of the intensity of the conditioning regimen and the application of a graft-versus-lymphoma effect. however, conventional conditioning regimens have been associated with a - % risk of transplant-related mortality (trm) in advanced hodgkin's lymphoma (hl). in an attempt to reduce the high trm reported with allografting in lymphoma, reduced-intensity regimens have been investigated. methods. four patients between the age of and years underwent allogeneic peripheral blood stem cell (pbsc) transplantation (sct) from hla-identical sibling or unrelated donors at our institution. age, sex, manifestation of disease, and donor were as follows: patient - years, female, pulmonary bulk, sibling; patient - years, male, pulmonary bulk, unrelated donor; patient - years, male, pulmonary bulk, sibling; patient - years, male, abdominal bulk, sibling. all patients had received multiple courses of polychemotherapy (table ) and local radiotherapy prior to sct. patients and had undergone autologous stem cell transplantation. we administered a reduced-intensity conditioning consisting of beam (bcnu, etoposide, cytarabine and melphalan) over days. to permit durable engraftment in the allogeneic setting, patients received additional pretransplant immunosuppression with an anti-cd antibody (campath) at a total dose of mg over days and graft-versus-host disease (gvhd) prophylaxis of cyclosporine a. pbsc of donors were collected after g-csf stimulation at µg/kg of body weight given for days. results. patient rejected his unrelated-donor graft and received an autologous stem cell infusion days thereafter resulting in sustained hematologic recovery (anc, > . g/l on day ). all other patients engrafted uneventfully (anc, > . g/l on days - ; platelets, > g/l on days - ). none of the patients showed evidence of acute or chronic gvhd. two patients achieved complete donor chimerism in myeloid and lymphoid cell populations, another one had prolonged mixed chimerism and was given a donor leukocyte infusion of × cd -positive cells per kg. one patient experienced a cmv reactivation on day after sct and a sarcoidosis on day + . three months after sct, all patients showed marked regression of disease. after a follow-up of to months (median, months), patients experienced progression undergoing salvage chemotherapy. time to progres-sion was and months, respectively. two patients remained progression free for and months, respectively. overall survival is to months. conclusions. our data demonstrate that this regimen was well tolerated with a low risk of gvhd and transplant-associated morbidity. an increased dosage of campath could be considered to prevent rejection in unrelated-donor transplantation. longer follow-up and larger patient numbers are warranted for assessment of the efficacy of campath-beam with regards to durable remissions. background. as long-term survivors of hematopoietic cell transplantation (hct) become more numerous, studies addressing the issue of long-term follow-up are necessary. in this study, we report on the quality of life (qol) of hct patients who were alive at least at years after transplantation in comparison with an age-and sex-matched sample of healthy controls assessed in the same time period and the same geographical region. methods. the eortc-quality of life questionnaire (eortc-qlq c ) was sent by post to hct survivors. thirty-four patients answered the questionnaire. patients were compared with healthy controls from the same geographical region. patients and controls completed the eortc in the same time period. results. mann-whitney u tests identified significantly lower qol on the dimensions of physical and social functioning and on the financial impact symptom scale. conclusions. patients who had survived their hct for more than years did generally well in terms of global qol. this is consistent with kiss et al. ( ) who found that cml patients who were alive at least years after hct report lower physical functioning in comparison with healthy controls. problems in the areas of social functioning and financial difficulty can possibly be addressed by intensive rehabilitation processes integrating patients, family members, and significant others. interdisciplinary (medical, psychological, and social) treatment of patients should not come to an end after the acute phase of the illness but should continue during checkups following transplantation. background. bone marrow transplantation (bmt) together with costimulation blockade can reliably induce mixed chimerism and tolerance in certain experimental models. natural killer t cells play an important role in the induction of tolerance in several transplantation and autoimmunity models. it has been reported, for instance, that activation of nk t cells by α-galactosylceramide (α-gal) can prevent the onset and recurrence of autoimmune type i diabetes. in recent experiments we wanted to investigate the role of nk t cells in a model of tolerance induction through bmt with costimulation blockade. to delineate the role of donor and recipient nk t cells, we used nk-t-cell-deficient mice (jα -/-c bl/ and jα -/-balb/c) as recipient and/or donor strain. we also evaluated whether in vivo stimulation of nk t cells with α-gal has a beneficial effect. methods. c bl/ mice and c bl/ jα -/-received a total-body irradiation (tbi) of gy or gy (day - ), approximately × fully mismatched balb/c or balb/c jα -/-bone marrow cells (day ) and costimulation blockade consisting of anti-cd mab (mr , day ) and ctla ig (day + ). groups were additionally treated with α-gal or a vehicle ( µg on day - , + , + , + , and + ). multilinage chimerism and skin graft survival were followed for more than days. results. with our standard protocol, of mice developed long-term multilinage chimerism and permanent donor skin graft survival. when using recipients deficient in nkt cells, of became chimeric and showed long-term skin graft survival; using nkt knockouts as donors of and using nkt knockouts both as recipients and donors of became chimeric (p = n.s. for all comparisons). unexpectedly, stimulation of nk t cells by α-gal (using wild-type recipients and donors) prevented chimerism induction after gy tbi ( of vs. of and of in the vehicle group, p < . ) and did not promote engraftment after gy tbi ( of vs. of ). conclusions. nk t cells do not play a critical role in tolerance induction after bmt with costimulation blockade. their stimulation with α-gal even prevents induction of mixed chimerism and tolerance. background. phospholamban (plb) is a critical regulator of sarcoplasmic reticulum ca + -atpase activity and myocardial contractility. in this study we investigated the role of plb phosphorylation in ischemia and reperfusion following cardiac transplantation. methods. gene expression of plb was investigated in a syngeneic heterotopic cardiac transplant model in mice. grafts underwent h of cold ischemia or were tranplanted immediately after harvest. gene expression was analysed at various time points employing dna microarray and rt-pcr. for in vitro experiments, hl- cardiomyocytes were submitted to a protocol of global normothermic hypoxia for h and reoxygenation in the absence or presence of the ca + /calmodulin kinase ii inhibitor aip ( µm) or the beta-adrenergic blocker dl-propranolol ( µm) vs. beta-adrenergic stimulator isoproterenol ( µm). results. at h, gene expression of plb was diminished by . and . -fold in groups with and without ischemia, respectively. basal phosphorylation of plb at ser (protein kinase a site) and at thr (ca + /calmodulin kinase ii site) was present in cultured cardiomyocytes and heart lysates. in the mouse system, increase in plb phosphorylation is observed during early (up to min) reperfusion. thereafter, plb phosphorylation drops below that of control levels. addition of aip diminishes reperfusion-induced thr phosphorylation; propranolol significantly decreases ischemia-induced ser phosphorylation. in contrast, isoproterenol enhances plb-ser and thr phosphorylation. conclusions. ischemia regulates phospholamban by two different mechanisms, decrease in expression levels and alterations in the phosphorylation of critical regulatory sites. modulation by aip and dl-propranolol will help for investigation of the role of pbl phosphorylation in ischemia and reperfusion in cardiac transplantation in the future. background. the p mitogen-activated protein kinase (p -mapk) pathway plays a crucial role in pathological events like oxidative stress, inflammation, and abnormal cellular proliferation, resulting in activation of several signalling cascades, involving overexpression of tumor necrosis factor alpha (tnf-α). tnf-α is known to play an important role in chronic rejection. currently, pharmacological inhibitors of p -mapk are being tested clinically for the treatment of experimentelle transplantation chronic inflammatory diseases such as rheumatoid arthritis, morbus crohn and psoriasis as well as inhibition of vascular smooth muscle cell (vsmc) proliferation after balloon angioplasty. to date, there are no findings that address the role of p -mapk in the context of chronic allograft vasculopathy, which is characterized by vascular lesions in the graft that consist of concentric myointimal proliferation, resulting in deterioration of allograft function and organ loss. the purpose of this study was to understand the role of p -mapk in abnormal vsmc proliferation associated with chronic rejection and to investigate the potential therapeutic role of a specific inhibitor of p -mapk activation in chronic allograft vasculopathy. methods. in vivo, a mouse model of heterotopic aorta transplantation in an allogenic setting has been used. aortas were allografted into recipient mice by a carotid artery cuff technique, using c bl/ (h- b ) mice as donors and balb/c (h- b ) mice as recipients. four weeks after transplantation, the aortic segments were harvested and immunofluorescence was performed using anti-vsmc-α-actin and anti-phospho-p antibodies. tnf-α serum levels were measured by elisa. in vitro, vsmcs were isolated from c bl/ aortas. expression levels of total and phosphorylated forms of p -mapk as well as key cell cycle regulators were detected by western blot. immunocytochemistry was performed with primary antibodies directed against phospho-p -mapk. proliferation of vsmcs was measured by [ h]thymidine incorporation in the presence or absence of sb , a specific inhibitor of p -mapk activation. cell cycle progression was monitored by dna content analysis; apoptosis by the annexin v binding assay. cell lysates were probed with antibodies directed against cyclindependent kinase (cdk ) and yin yang (yy ). results. in vivo, weeks after the transplant aortic segments were significantly narrowed due to neointimal hyperplasia. the neointimal lesions mainly consisted of vsmcs and showed profound activation of p -mapk as demonstrated by immunofluorescence. further, serum tnf-α levels were significantly increased even weeks after allogeneic aortic transplantation, suggesting an important role of p activation in this model. in vitro, stimulation of vsmcs with % fcs resulted in a rapid increase of phosphorylated forms of p -mapk ( . ± . fold increase) when compared with the nonstimulated quiescent state. immunocytochemistry showed higher levels of phospho-p -mapk in the nuclei as well as in the cytosol after stimulation. sb in a dose-dependent manner significantly inhibited vsmc proliferation, which was due to inhibition of cell cycle progression at the g /g phase. we did not observe apoptosis in the sb -treated vsmcs at µm, the highest dose being tested. blockade of p -mapk activation decreased protein levels of cdk and the transcription factor yy , which plays an important role in vsmc dna replication and protein synthesis. conclusions. p mapk activation appears to play an important role in an in vivo allogeneic model of aorta transplantation as well as in vsmc proliferation in vitro, blocking of which prevents the cells from entering the s phase of the cell cycle, thus abrogating cell proliferation. targeting p -mapk might become a potent strategy in the treatment of vascular proliferative diseases like chronic allograft vasculopathy. background. ctla ig, a costimulation blocker which is currently under advanced clinical development, has been used for years as part of mixed chimerism protocols. recent data suggest that ctla ig also functions via modulation of tryptophan metabolism by upregulating indoleamine , -dioxygenase (ido) through b signals. we thus investigated the role of ido in our ctla ig-based mixed chimerism protocol. methods. c bl/ mice received a total body irradiation (tbi, d - ) of gy, approx. × fully allogeneic balb/c bone marrow cells (d ) and costimulation blockade consisting of anti-cd mab ( mg, d ) and ctla ig ( . mg, d ). different groups of mice were additionally implanted with -methyltryptophan pellets on d - ( -mt, which is a competitive inhibitor of ido, -day release at . mg/h) or on d - and d ( -day release) or placebo pellets. macrochimerism and deletion of donor-reactive cells were followed by flow cytometry. levels of tryptophan, kynurenine, and -mt were measured at several timepoints in serum by hplc. results. of mice which received bmt, tbi, mr plus ctla ig but only of mice without ctla ig developed lasting multilineage chimerism (p < . , measured at week post bmt). of mice with ctla ig treatment accepted donor skin grafts for more than days, whereas only of mice, which got no ctla ig injection accepted donor skin long-term (p < . ; representative data from two separate experiments), demonstrating that ctla ig is critical for our protocol. rd-party grafts were promptly rejected in all groups. long-term multilineage chimerism developed in of mice with -mt treatment, which is not significantly different from treatment with placebo pellets or standard protocol alone ( of , pooled data from both groups). -week treatment with -mt also did not lead to a significant difference in the rate of multilineage chimerism ( of with -mt vs. of without -mt). deletion of donor-reactive t cells was neither blocked nor enhanced by treatment with -mt. the kynurenine-to-tryptophan ratio in serum was similar in groups with ( . ± . ) and without ( . ± . ) ctla ig (p = n.s., measured on d ). substantial serum levels of -mt were detected in mice with -mt treatment but not in untreated mice, indicating that ido was indeed inhibited in the -mt groups. conclusions. ctla ig plays an essential role for tolerance induction in this model but its mechanisms of action does not critically depend on ido. background. induction of antigen-specific tolerance remains the ultimate goal in clinical organ and cell transplantation, as it would eliminate the need for continuous immunosuppression. one strategy leading to tolerance induction against a transplanted organ consists of infusing blood from the organ donor, an approach known as donor-specific transfusion (dst). although the mechanisms underlying tolerance induction by dst are not fully understood, clonal deletion of alloreactive t cells and generation of immunoregulatory cd + cd + t cells are important in the process. it is well established that expression of heme oxygenase- (ho- ) can promote the survival of transplanted organs. however, the mechanisms underlying this effect remain to be elucidated. we hypostesized that ho- is required for tolerance induction involving dsts and that ho- can magnify the tolerogenic effect of dsts. methods. allograft survival has been tested by using a well established model of costimulatory blockade (anti-cd l-ab) plus dst (day - ) in c bl/ (h- b ) heart allografted balb/c (h- d ) wt and ho- ko mice. further, ho- activity has been induced (by copp) or suppressed (by znpp) in b af (h- k/d,b ) mice receiving dba/ (h- d ) allografts plus dst on day or day - . donor-specific tolerance was tested by challenging the mice with a second dba/ or third-party fvb (h- q ) allograft. leukocytes (depleted or undepleted of cd + cd + t cells) from mice carrying allografts for > days were adoptively transferred to sublethally irradiated b af mice receiving dba heart allografts. rna levels of foxp , tgf-β, il- and ctla- have been assesed by using rt-pcr. cd + cd + t cells have been enumerated by flow cytomerty. results. anti-cd l-ab plus dst treatment resulted in balb/c recipients tolerizing fully mismatched c bl/ allografts (n = ). however, by using ho- -deficient recipients, this effect was abrogated, in that c bl/ allografts have been rejected in a similar manner as in untreated wt recipients (mst = . d, n = ). further, dst plus copp (in contrast to dst plus znpp) treatment resulted in donor-specific tolerance of dba/ allografts in b af recipients (n = ). tolerant animals showed significantly increased percentage of cd + cd + t cells and increased levels of foxp , tgf-β, il- , and ctla- mrna. adoptively transferred b af leukocytes retrieved from the lymph nodes and spleens transferred to sublethally irradiated b af recipients of dba allografts led to subsequent allograft loss (mst = . d, n = ); in contrast, transfer of leukocytes of tolerant (copp plus dst treated) mice led to indefinite allograft survival in this model. however, when those leukocytes were depleted of cd + cd + t cells, allografts were immediately rejected (mst = . d, n = ). conclusions. ho- in a graft recipient can be critical for long-term graft survival and for induction of tolerance. this mainly is mediated by generation of cd + cd + t cells (t regs). modulation of ho- expression and activity may be used therapeutically to promote graft tolerance. background. human immunoglobuline (ivig) has been advocated in the treatment of acute rejection in allograft transplantation and treatment of sepsis. mechanisms describing the immune modulatory activity are however scarce. we sought to investigate immune suppressive properties of pooled human immuneglobuline (ig), unspecific fc and fab fragments and their respective ligands by allogeneic blastogenesis assays. methods. human mixed lymphocyte reactions (mlr) were performed. in detail peripheral blood mononuclear cells (pbmcs) were purified from donors by ficoll density gradient centrifugation (amersham biosciences, buckinghamshire, england). , cells per well were stimulated with , radiated pbmc ( gy) and incubated for days at °c together with unspecific igg, igm (both sigma-alderich, st. louis, mo), anti-cd (lab vision, fremont, ca) and anti-cd (chemicon, temecula, ca) in a dose-dependent fashion. before harvesting cells were pulsed for h with [ h]thymidine ( . × bq/well) and the [ h]thymidine uptake was measured in a liquid scintillation counter. results. the addition of pooled human igg and igm to allogeneic stimulated cells both resulted in a significant and dose dependent decrease of proliferation, although the suppressive properties of igm was greater as compared to igg (both, p < . ). to investigate whether this effect was partly related to fc receptor involvement we blocked cd and cd on antigen presenting cells (apcs), known receptors in the activation of mlr. surprisingly, this assay demonstrated that sole fc blocking (high-and low affinity fc receptors) resulted in a significant downregulation of allogeneic response in vitro (both, p < . ). conclusions. our data evidence that the addition of unspecific immune globulines results in a reduction of proliferation in in vitro allogeneic blastogenesis assays and is partly fc receptor dependent. these results corroborate the clinical success of ivig and pooled igm in the treatment of solid organ allograft rejection and cautions the utilization of immune globulines in immune suppressed septic patients. die interaktion professioneller antigen-präsentierender zellen mit allogenen t-zellen resultiert in der ausbildung der sog. immunologischen synapse (is), welche für die effiziente aktivierung von t-zellen und damit letztlich für die transplantatabstoßung essentiell ist. im rahmen der is werden der t-zellrezeptor/cd -komplex, kostimulatorische wie adhäsionsmoleküle und komponenten des zytoskeletts in die kontaktzone des mhc-tcr/cd -komplexes der is rekrutiert. der einfluss von gängigen immunsuppressiva, die zur prävention der allogenen organabstoßung klinisch verwendet werden, auf die is-evolution ist bislang unbekannt. in dieser studie zeigen wir erstmals, dass kalzineurinhemmer wie csa oder fk , aber nicht mtor-inhibitoren, selektiv die rekrutierung des tcr/cd -komplexes in die is blockieren. ein ähnlicher effekt wurde für kortikosteroide, aber nicht mycophenolat beobachtet, was auf eine essentielle rolle des calcineurin-nf-κb-signalweges für die tcr/cd -regulation im rahmen der t-zellaktivierung hinweist. interessanterweise blockierte das neue immunsuppressivum fk nicht nur die tcr/cd -, sondern auch die lfa- -und f-aktin-rekrutierung in die is. die bedeutung dieser globalen interferenz mit der is-ausbildung für die spezifische immunantwort muss in weiterführenden studien geklärt werden. diese daten zeigen, dass klassiche immunsuppressiva nicht nur simple blocker der zytokinsynthese sind, sondern schon sehr früh die t-zell-apz-interaktion stören und damit einen weiteren immunologischen mechanismus besitzen, der ihre klinische potenz erklärt. background. ischemia (i) and reperfusion (r) trigger a series of events, which culminate in severe injuries to the affected organs in organ transplantation. cell death, metabolic alterations, and inflammation result in impairment of shortand long-term function. the group of mitogen-activated protein kinases (mapks) are central regulators of these events, they have been implicated through aberrant activation in many pathophysiological settings ranging from autoimmune diseases, cancer, to i/r-associated organ damage. methods. intracellular signaling pathways were analysed in vivo and in vitro employing a cardiomyocyte cell line and a murine heart transplant model. hl- cardiomyocytes are a cardiac muscle cell line derived from at- mouse atrial cardiomyocytes. syngeneic cardiac transplants were carried out us-ing male inbred balb/c mice, hearts were transplanted heterotopically into the neck of recipients. results. in summary, (i) reoxygenation was characterized by a dramatic increase in the activity of all mapks at the end of the observation period; (ii) growth factor abrogation together with hypoxia (h) and reoxygenation (r) had a substantial effect on the course of signaling events; (iii) signaling processes in response to ischemia and reperfusion in vivo are in line with observations made in cardiomyocytes. conclusions. data obtained so far in our study demonstrate the suitability of the chosen experimental approaches for investigation of i/r-associated alterations in intracellular signaling and cellular responses. preliminary data suggest that h/r treatment of hl- results in significant apoptotic cell death, the intracellular signaling pathways involved are therefore currently analyzed. background. ischemia and reperfusion (i/r) in cardiac transplantation results in inflammation and cell death. to gain further insight into the regulation of these processes, we investigated the role of lipocalin- ( p , ngal, uterocalin), a potential regulator of the acute inflammatory response and cellular apoptosis in vitro and in vivo. methods. c bl/ hearts were heterotopically transplanted to syngeneic recipients immediately and after hours of prolonged cold ischemia with the grafts harvested at various timepoints ( min, h, h, h, h, d) after transplantation. gene expression analysis on mrna extracts was performed employing cdna microarray and rt-pcr. protein synthesis was investigated by western blotting and apoptosis by using tunel assay. for the identification of the cellular source of lipocalin- and its function in i/r-associated cell death, the effect of recombinant lipocalin- was investigated in the hl- cardiomyocyte cell line. hl- cardiomyocytes undergoing ischemia/reoxygenation as well as purified mononuclear cells and granulocytes were analyzed for lipocalin- expression by rt-pcr. results. in the cardiac transplants, high levels of lipocalin- gene and protein expression were detected at h of reperfusion, whereby transcription was higher in groups without cold ischemia ( -versus . -fold). he staining demonstrated dense mononuclear infiltrates, cellular edema, and small focal necrosis in groups with and without prolonged cold ischemia. upregulation of gene transcription was confirmed by pcr. apoptotic cells were first detectable at day and peaked days after transplantation. expression of lipocalin- was also detected in hl- cells by rt-pcr and western blotting following i/r, demonstrating for the first time the presence of lipocalin- in this cell lineage. lipocalin- -transfected hl- cardiomyocytes showed a higher cell viability especially under ischemic condition. megalin, known as the lipocalin- receptor, was detected in hl- cells by rt-pcr. conclusions. this study demonstrates the time-dependent expression of lipocalin- in a cardiomyocyte culture as well as in transplanted hearts in response to ischemia and reperfusion/reoxygenation. lipocalin- is suggested to target cardiomyocytes with ameliorating effects on cell viability during ischemia. obvious alterations in lipocalin- expression at the protein level suggest a possible involvement of lipocalin- during i/r-induced cell death probably as a self-limiting process for inflammation. background. protease activation as well as inflammatory responses contribute to organ damage in response to ischemia and reperfusion. in this study we investigated the role of the protease inhibitor slpi in ischemia and cardiac transplantation. methods. hearts from slpi knockout mice (slpi-/-) were heterotopically transplanted to slpi-/-recipients. grafts underwent hours of cold ischemia (ci) prior to transplantation or were transplanted immediately. c bl/ wild-type isografts (wt) undergoing the same procedure served as controls. in selected groups, µg of recombinant slpi (rslpi) were added to the preservation solution or given i.v. after reperfusion. after evaluation of graft function, hearts were removed at min, h, h, and days. morphology was investigated by histology, slpi gene expression was analysed using quantitative rt-pcr. slpi protein expression was studied by immunohistochemistry (ihc). slpi, tnf-α, tgf-β, and nf-κb, cathepsin g, and elastase activity were analysed employing elisa and western blot. results. at min, recovery of graft function was normal in wt and slpi-/-mice transplanted without ci ( . ± . ). in contrast, slpi-/-hearts transplanted after h of ci showed no or marginal recovery of organ function ( . ± . ). at h, cardiac function in slpi-/-( . ± . ) was less when compared with wt ( . ± . ). single administration of rslpi i.v. had no effect; however, when slpi was added to the preservation solution, organ function comparable to wt mice was observed ( . ± . ). a mild mononuclear cell infiltrate and small focal necrosis where found in all groups at h. at days, postischemic inflammation as well as myocyte necrosis were significantly higher in the slpi-/group ( . ± . vs. . ± . and . ± . vs. . ± . ). myocyte vacuolisation as a sign of sublethal ischemic injury was present at high level in slpi-/-mice undergoing ci only. slpi gene expression was detected in wt mice at and h after reperfusion. gene transcription at h was significantly higher after prolonged ci ( . vs. . orders of magnitude) and was associated with significantly decreased nf-κb, tgfβ, and tnf-α activity. slpi protein was first observed at h, high levels of slpi protein were found at days. slpi-positive cells were mainly identified as macrophages (ihc). high intragraft levels of slpi activity were found early as well as days after application of recombinant protein. high slpi levels correlate with decreased cathepsin g early and decreased nf-κb, tgf-β, and elastase activity late after reperfusion. conclusion. herein we demonstrate that slpi has a substantial effect in prevention of inflammation and myocyte damage in response to ischemia and reperfusion of the heart via inhibition of nf-κb, tgf-β, and elastase. in addition, slpi seems to be crucial for recovery of organ function early after heart transplantation -inhibition of protease activity seems to be the underlying mechanism. perfusion with rslpi ex vivo represents a promising therapeutic option for modulating the destructive processes of postischemic inflammation while preserving its restorative nature. methodik. die studie wurde an hausschweinen durchgeführt. die tiere wurden in allgemeinnarkose versetzt und intubiert. alle hämodynamischen daten wurden invasiv gemessen. die parameter (abp, pap, zvd und lap) wurden wie in der klinischen praxis mit einem hp-patientenmonitor registriert und gespeichert. das herzzeitvolumen (co) wurde mit hilfe eines flowmeters der firma transsonic systems inc. gemessen. die hämodynamischen ausgangswerte von patienten vor ecmo wurden retrospektiv mit den experimentellen daten verglichen. ergebnisse. in der frühphase des tierversuchs nach erfolgter abklemmung blieb ein eindeutiger anstieg des pap bei von versuchstieren aus. auch ein rascher abfall des co war nicht zu beobachten. auch der pvr zeigte keine signifikante veränderung. die pap/ap-druckratio reagierte sehr rasch und in allen fällen mit einem anstieg auf das ereignis. von patienten, die mittels ecmo erfolgreich therapiert worden waren, hatten eine pap/ap-ratio über , , ein patient hatte eine pap/ap-ratio von , . die patienten mit pap/ap über , wurden mittels va-ecmo therapiert, der patient mit pap/ap von , mittels vv-ecmo. schlussfolgerungen. isolierte veränderungen der pulmonalen hämodynamik repräsentieren offenbar nur bedingt den schweregrad der vorhandenen beeinträchtigung. als wichtiger parameter für eine rasche einschätzung des grades der kompromittierung kann die druckratio zwischen pulmonalem und systemischem kreislauf angesehen werden. als leicht zu erhebender parameter könnte die pap/ap-ratio eine entscheidungsgrundlage für die zu wählende ecmo-konfiguration darstellen. nach den bisherigen erfahrungen wäre die indikationsgrenze bei einer pap/ap-ratio von etwa , zu ziehen. background. pancreas transplantation in the mouse is an extremely demanding procedure and severe technical problems have limited its widespread use. since the mouse, however, would be a good model for the study of various transplantation-related problems, such as ischemia-reperfusion injury or graft pancreatitis, we designed a new surgical strategy for cervical heterotopic vascularized pancreas transplantation using a cuff technique. methods. male syngeneic c bl (h- b ) mice (n = ) -to -week-old were used as size-matched donor and recipient pairs. recipients were intraperitoneally injected with . mg of streptozotocin per kg in order to become hyperglycemic (blood glucose, > mg/dl) and transplantation was performed days later. recipient operation: the right external jugular vein (ejv) and common carotid artery were dissected free. by using a polyethylen cuff (od, . mm) it became possible to evert the artery over the cuff body and finally fix the vessel with - silk ligatures. similarly, the ejv could be everted over a . mm cuff. donor operation: after a complete midline incision the pancreas was isolated using a no-touch technique on a segment of the aorta, including the celiac axis and the superior mesenteric artery. the venous outflow was provided by the portal vein. all grafts were flushed with °c saline solution. implantation: the graft was placed in the right cervical region and vascular anastomoses completed by pulling the pv over the ejv cuff and the donor aortic segment over the carotid cuff and held in place with a - silk ligature. after releasing venous and arterial clamps, all grafts immediately returned to their normal pink color with the arterial stump pulsating. results. out of recipients, surviving over days, animals died from haemorrhage (survival rate, %). donor operation lasted ± min and dissection of recipient vessels took ± min. implantation time was to min, resulting in a total pancreas ischemia time of ± min. no thromboembolic complications at the cuff side were observed. preoperative glucose levels were ± mg/dl and could all be normalized by po day ( ± mg/dl). histopathological examination on po day and showed almost normal islet cell and acinar architecture of all grafts. conclusions. for the first time a method of cervical heterotopic pancreas transplantation using a non-suture cuff technique in the mouse is described. major advantages are a short ischemia time, lack of arterial thrombosis or venous stenosis, and short operation time, and thus a very high survival rate. this model is especially applicable for investigating preservation, reperfusion injury, and graft pancreatitis. background. as human islet transplantation is limited by the lack of sufficient numbers of human donor organs, xenotransplantation with the use of porcine islet cells seems to be a promising therapeutic option to cure diabetes. in order to achieve sufficient numbers of viable islet cells, better protocols for organ preservation, isolation, and purification are needed. recent studies showed that the two-layer method (tlm) of pancreas preservation prior to isolation significantly improved islet yield. the tlm oxygenates pancreata and activates metabolism to generate atp and leads to resuscitation of ischemically damaged organs. another possibility to achieve a higher partial pressure of oxygen levels in fluids is the use of hyperbaric oxygenation (hbo). the aim of this study was to assess the influence of preoxygenated preservation solutions on the porcine pancreas. methods. university of wisconsin solution (uw), celsior, perfadex, custodiol, and a preservation solution especially designed for this study on the basis of ketoglutarate are oxygenated with % oxygen for minutes at . bar using a hyperbaric chamber. porcine pancreata are harvested at a local slaughter house and stored in preoxygenated and not oxygenated preservation solutions at °c for hours. tissue cuts are performed to assess the occurrence of apoptosis and to determine the oxidative stress. atp-to-adp ratio is measured and immunohistochemistry is performed. results. it is feasible to preoxygenate preservation solutions. the oxygen levels can be raised up to times in the preservation solutions using hbo and can be maintained for at least hours after hbo treatment. mda and carbonylated protein levels are not significantly elevated in organs stored in preoxygenated solutions. atp-to-adp ratio as a sign of viability is significantly higher in organs stored in preoxygenated solutions. conclusions. preoxygenation of preservation solutions using % oxygen and a hyperbaric chamber is feasible. hbo has a positive impact on porcine organ preservation. as ischemically damaged islet cells are likely to undergo cell death or lose functionality due to hypoxia, the use of preoxygenated preservation solutions is a promising method to achieve better yields after islet isolation and transplantation. alternative zur humanen pankreastransplantation und inselzelltransplantation stellt die xenotransplantation von porcinen inselzellen dar. um xenogene zellen erfolgreich transplantieren zu können, müssen sie vom immunsystem des empfängers abgeschirmt werden. die verwendung von mikrokapseln scheint eine vielversprechende methode dazu zu sein, wobei natrium zellulose sulfat (nacs) in graz verwendet wird. darüber hinaus muss eine ausreichende anzahl an vitalen zellen mit hoher reinheit aus dem porcinen pankreas isoliert werden. methodik. porcine pankreata werden von einem lokalen schlachthof erhalten. die organe werden kanüliert, und ein enzymgemisch aus neutraler protease und kollagenase nb wird infundiert. die digestion erfolgt in einer modifizierten ricordi-kammer mechanisch und enzymatisch, und die anschließende purifikation wird mit einem ficoll-dichtegradienten durchgeführt. vitalität der zellen wird mit fluorescein-diacetat/propidium jodid, mtt und der bestimmung der atp/adp-ratio überprüft. die reinheit wird mit einer dithizon-färbung festgestellt. insulinkonzentrationen werden mittels elisa detektiert, der dna-gehalt der inselzellen mit dem dneasy-kit festgestellt. inselzellen werden in kooperation mit der firma austrianova mit nacs mikroverkapselt. ergebnisse. die organe wurden durchschnittlich min digestiert, es wurde eine reinheit von % und eine zellzahl von durchschnittlich × zellen isoliert. beste resultate wurden mit dem lymphoprep tm -dichtegradienten erzielt. isolierte porcine inselzellen überleben bei °c im durchschnitt tage in vivo und produzieren glucose-abhängig insulin. die funktionalität der zellen bleibt über den gesamten zeitraum erhalten. mikroverkapselung mit nacs ist machbar. schlussfolgerungen. die isolation porciner inselzellen ist eine viel versprechende methode den mangel an humanen spenderorganen in der pankreastransplantation und humanen inselzelltransplantation zu umgehen. nacs als verkapselungsmaterial ist weniger immunogen und weitaus biokompatibler als alle bisher verwendeten materialien und die mikroverkapselung xenogener inselzellen mit nacs scheint eine innovative methode zur therapie des diabetes mellitus zu sein. background. to achieve its full potential, transplantation of pancreatic islets has to overcome a number of obstacles. one of the obstacles are the still lacking read-out parameters to assess the quality of human islets after the isolation procedure and prior to transplantation. being able to predict the functional potential of the pancreatic islets after isolation or even short-term culture would greatly enhance the success of islet transplantation. therefore one of the primary challenges in islet transplantation is to identify and understand the changes taking place in islets after isolation or culture. life confocal microscopy is a powerful tool to identify such changes in living islets not achievable by use of fixed-cell techniques. methods. islets were isolated according to the method of ricordi et al., using a continuous ficoll gradient. fluorescent dyes, dichlorodihydrofluorescein diacetate (dcf), tetramethylrhodamine methyl ester (tmrm), and fluorescent wheat germ agglutinin were used to assess either overall oxidative stress, time-dependent mitochondrial membrane potentials, or localisation of oligosaccharides. confocal microscopy was performed with an microlens-enhanced nipkow disk-based confocal system ultraview rs (perkin elmer, wellesey, ma, usa) mounted on an olympus ix- inverse microscope (olympus, nagano, japan). results. with the above described confocal system we were able to identify differences in the localisation and amount of oligosaccharides in endocrine vs. exocrine cells of freshly isolated pancreatic islets. the staining pattern changed during the course of culture, suggesting a remodeling of cell surface oligosaccharides. the study of the mitochondrial membrane potential proved to be very useful in order to early identify damaged or stressed cells and thereby gain insights into the vitality of the isolated islets. conclusions. this makes us believe that, especially in the light of the many other fluorescent dyes which can be used as subcellular markers, a combination of these with a powerful live confocal imaging system will be of great value for a better islet assessment after isolation and culture. background. therapeutic drug monitoring (tdm) of immunosuppressants is a well established concept supporting the work of clinicians from the historical onset of the use of these drugs. within the last years the combination of hplc (highperformance liquid chromatography) with tandem mass spectrometry (ms/ms) provides an alternative to antibody-based immunoassays. this new analytical method to quantify immunosuppressants affords a robust and rapid separation technique with high selectivity. up to now, the quantification of wholeblood levels of cyclosporin a, fk (tacrolimus), and -o-( -hydroxyethyl)-rapamycin (everolimus) at our institute have relied upon three different, indirect assays from different companies. however, these systems have drawbacks caused by cross reactions with active and inactive drug metabolites, fluctuations in assay performance, and comparatively high prices. in contrast, hplc-ms/ms platforms are now becoming frequently used to measure circulating drug levels and their metabolites with lower associated costs and higher specificity, accuracy, and precision. methods. the setup chosen at our laboratories consists of two independent mass spectrometers (one machine at the zimcl, a backup machine at biocrates) and allows a sample throughput of about samples per hour. all immunosup-varia pressants currently monitored in whole blood can be quantified within one analysis from a sample volume of less than µl. the sample workup (cell lysis and protein precipitation) is performed in a bar-code-supported parallel setup, minimizing the risk of sample mix-up. an online solid-phase extraction (spe) strategy has been chosen to reduce matrix interferences and ion suppression. daily calibrations and quality control samples performed with certified reference materials assure a high level of accuracy and precision. results. the hplc-ms/ms assay established for cyclosporin a, tacrolimus, everolimus, and sirolimus covers the analyte concentration range needed for tdm (e.g., up to mg/ml for cyclosporin a). intraday and interday repeats of the assay and data from quality control measurements were sufficiently accurate and precise (all cvs, < %; bias, < %). comparison of the quantitative results did show a linear relationship between antibody-based immunoassays and the hplc-ms/ms-derived data with bias values (ca. % for cyclosporin a and ca. % for tacrolimus on the basis of bland-altmann plots) in agreement with the literature. conclusions. the established hplc-ms/ms platform will allow replacement of the current antibody-based assays. the major advantage of this technique is the ability to simultaneously acquire absolute quantitative concentrations of each of the therapeutic drugs administered from one sample. therefore, the requirement for different sample preparation schemes or parallel measurements on different analytical instruments are no longer needed. the high sample throughput also assures timely tdm data report to the ward. a significant reduction of costs is now expected due to the lower consumable expenses in hplc-ms/ms assays. background. a clinical trial towards improved safety of the application of everolimus (certican) in heart transplantations (crad a , data collection started in august and is ongoing) allowed us to compare therapeutic drug monitoring (tdm) data (e and e levels) measured on two different platforms -an lc-ms/ms method in an external laboratory and a immunochemical method recently established at our institute. methods. lc-ms/ms measurements of everolimus were performed in a reference laboratory. the immunochemical measurements were performed with a fluorescence polarization immunoassay (innofluor certican immunoassay by seradyn) measured on a tdx instrument from abbott. method comparison included analysis of the time series of the patients (n = , observation pairs) and the quality control samples of the immunochemical assay gathered over a year as well as inter-and intra-assay data comparisons based on bland-altmann plots and regression data. results. comparison of the quantitative results obtained from the lc-ms/ms and the immunochemical assay showed good agreement between these methods. the bias between the methods (bland-altmann plot) was found to be rather small, with the immunochemical method measuring in average approximately % (± %) lower values as the lc-ms/ms method, which is in good agreement with current reports. the coefficient of variation (cv) (measured over several months) of the innofluor quality control measurements was < % for the medium (ca. ng/ml) and high (ca. ng/ml) quality control samples, whereas the lower quality control level (ca. . ng/ml) showed an increased cv (< %). however, especially during the phase of method establishment, higher cv values and bias deviations have been found conclusions. the method comparison did show that the immunochemical everolimus assay provided by seradyn is a good alternative to hplc-ms/ms measurements. the assay bias was found to be rather low and the assay uncertainty was within acceptable ranges. however, a stringent quality control network must be provided to assure stable assay performance over time. grundlagen. die inzidenz der endokarditis beträgt ca. / einwohner pro jahr in der gesamtbevölkerung. obwohl die immunsuppression das auftreten systemischer infektionen begünstigt, sind endokarditisstudien an transplantationsempfängern nicht verfügbar. ziel dieser epidemiologischen studie war, das auftreten der endokarditis und ihrer risikofaktoren nach organtransplantation zu evaluieren. methodik. insgesamt patienten, welche sich zwischen und einer soliden organtransplantation an unserem zentrum unterzogen, wurden untersucht. der mbds unserer klinik wurde zum patientenscreening herangezogen. ergebnisse. insgesamt wurden im beobachtungszeitraum endokarditisfälle beobachtet. neun endokarditisfälle ( , %) konnten erst post mortem mittels autopsie diagnostiziert werden, patienten ( , %) konnten durch alleinige antibiotische therapie geheilt werden. insgesamt transplantationsempfänger ( , %) mussten sich einem kardiochirurgischen eingriff unterziehen. die gesamtmortalität betrug , % ( patienten). staphylococcus aureus konnte in fällen ( , %) und pilze konnten in fällen als ursächliche keime isoliert werden. die inzidenz der endokarditis in transplantationsempfängern beträgt % ( % ci, . - . ) und zeigt ein -fach erhöhtes risiko verglichen mit der gesamtbevölkerung. schlussfolgerungen. die endokarditis stellt ein signifikantes problem nach organtransplantation dar und ist mit einer exzessiv hohen mortalität assoziiert. eine erhöhte aufmerksamkeit ist daher indiziert, da wir durch den einsatz der transösophagealen echokardiographie vermehrt derartige fälle in der zukunft diagnostizieren werden. ein -jähriger patient erhält eine herztransplantation aufgrund einer dilatativen kardiomyopathie bei altersentsprechender nierenfunktion. die immunsuppression besteht aus cyclosporin a und mycophenolat mofetil. innerhalb der ersten drei monate nach der operation steigt das kreatinin auf , mg/dl an, bleibt dann aber konstant. ab nimmt die nierenfunktion weiter ab, weshalb im august wegen des verdachtes auf eine chronische calcineurininhibitor-nephropathie auf sirolimus und mmf umgestellt wird. das serum-kreatinin beträgt zu diesem zeitpunkt , mg/dl. zwei monate später ist das serum-kreatinin auf , angestiegen, der inzwischen -jährige patient leidet unter einer renalen anämie, hyperphosphatämie und hypokalziämie. im harn finden sich leukozyten und erythrozyten, die allerdings nicht deformiert sind (keine dysmorphen erythrozyten und akanthozyten), sowie eine geringgradige proteinurie ( mg/ h). in der harn-polyacrylamidgel-elektrophorese zeigt sich vor allem alpha- -mikroglobulin als indikator einer tubulointerstitiellen schädigung. nach erneuter umstellung der immunsuppression von sirolimus/mmf auf niedrigdosis-cyclosporin/mmf kommt es zum raschen rückgang des serum-kreatinins, der harnbefund sowie die elektrolyte normalisieren sich, einzig die erythropoetin-substitution muss beibehalten werden, um den hämoglobin-zielwert von g/dl zu erreichen. acht wochen nach der zweiten therapieumstellung führen wir eine nierenbiopsie durch. es findet sich eine akute interstitielle nephritis ohne glomeruläre schädigung. unseres wissens ist dies der erste dokumentierte fall einer akuten interstitiellen nephritis auf sirolimus. background. group milleri streptococci (gms) are a heterogeneous group of streptococci including the species streptococcus intermedius, s. constellatus, and s. anginosus. due to their ability of producing toxins, they tend to cause chronic intra-abdominal and intrathoracic abscesses, which are difficult to treat, as gms are able to escape conventional antibiotic therapy. aim. evaluation of epidemiology, clinical significance, and impact on the outcome in all solid-organ recipients with gsm infections during a -year period. patients and methods. retrospective analysis comprising solid-organ recipients with gms. results. between and , solid organ recipients ( isolates) including liver, four kidney and pancreas, one kidney, two small bowel, three combined liver and kidney, and one combined kidney and small bowel transplant re-cipients developed infection with gms. in cases, gms caused intra-abdominal infection; in two cases, pleural empyema; and in one case, soft tissue infection. in only one case, gms were cultured from blood. in of the specimens ( %) which grew gms, also other pathogens could be isolated. gms frequently caused recurrent cholangitis (n = ) associated with anastomotic and anastomotic biliary strictures. these cases were managed by repeated stenting or surgical intervention and prolonged antibiotic therapy. no patient died directly related to gms infection. all responded to combined surgical and antibiotic treatment. one pancreas graft was lost due to erosion haemorrhage associated with an abscess. all isolated strains of gms were susceptible to penicillin g, carbapenems, and clindamycin, whereas cephalosporins and quinolones showed intermediate activity or resistance in some cases and gms in general were found resistant to aminoglycosides. conclusions. gms are frequent pathogens in transplant surgery and are capable of causing difficult to treat infections. their prevalence in transplant surgical site infection thus far might have been underestimated. therefore, we recommend empiric antibiotic treatment for sufficiently long time in combination with surgical intervention when necessary. background. by using intensified immunosuppressive protocols the incidence of immunological complications after solid-organ transplantation has constantly declined. however, the incidence of some infections, in particular complicated fungal infections, seems to increase. candida krusei (ck) is resistant to fluconazole and recently this pathogen has been more commonly isolated in severe infections, particularly in the immunocompromised host. methods. between . . and . . , a total of solid-organ transplants were performed at the innsbruck medical university. this included renal, liver, pancreas, intestinal, cardiac, lung, and islet transplants. prophylactic immunosuppression consisted of calcineurin inhibitor-based triple drug therapy for the majority of cases. antifungal prophylaxis was given to all pancreatic and intestinal recipients (fluconazole mg per day) and to all patients considered at high risk for filamentous-fungal infections (ambisome mg/kg). results. a total of five patients with ck infection were identified within this series ( %). five patients developed infection with candida tropicalis and three with candida glabrata. within this time period, another two patients with ck infection were identified, one had a pleural empyema following esophageal perforation and the other was treated with a ventricular assist device and developed pulmonary infection with ck. both patients were treated on the transplant intensive care unit. the transplant patients were three pancreas, one liver, and one lung recipient. all three pancreas recipients were diagnosed intra-abdominal infection, the liver recipient had an ischemic cholangiopathy with ck cholangitis and required retransplantation, and the lung recipient developed postoperative hemorrhage and subsequently ck pleural empyema. all patients presented also with other infectious complications. treatment of ck infection consisted in four cases of caspofungin or voriconazole or combination of the two, and in one case, ck was isolated from infected hematoma and did not require antifungal therapy after surgical removal. all infected collections were evacuated either surgically (n = ) or through pig-tail drainage (n = ). ck infection was successfully managed in all cases and patients are currently alive, only one pancreas graft was lost. conclusions. candida krusei infections now represent frequent severe complications in solid-organ recipients. however, rapid diagnosis and treatment with new antifungal agents such as voriconazole, caspofungin, or ambisome allow successful therapy of these infections. solid-organ recipients seem to be at increasing risk to acquire non-albicans candida infections. background. after solid-organ transplantation immunosuppression (is) and concomitant infection with cmv, ebv, hhv- , - , or papillomavirus put patients at risk for developing malignant diseases ( % cumulative risk at years; adami et al. ) . posttransplant lymphoproliferative disorders (ptld) and skin cancers are known to have the highest incidence in immunocompromised patients. reports on colorectal cancer after different types of organ transplantation are rare and the incidence was . % to . % and did not differ from that of the normal population. we analysed our database with regard to the incidence and course of colorectal malignancies following treatment and introduction of tor inhibitor-based is. methods. medical records of solid-organ transplants performed between and at our center were analysed retrospectively. in a total of patients heart, lung, kidney, liver, pancreas, and combined heart-lung transplantations were performed ( small bowel and hand transplantations not included). immunosuppressive therapy consisted of triple therapy comprising steroids, azathioprine/mmf, and cya/tac. some of them received induction therapy with antithymocyte globulin. results. a total of patients ( . %) developed malignancies. of them, patients ( female, male; median age at diagnosis, . years; kidney, heart, liver recipients) had colorectal malignancies ( . %) during a mean follow-up period of . years. on average, diagnosis was made . years after transplantation. four carcinomas were located in the rectum or at the rectosigmoid junction and five were colon cancers (five pt , one pt , and three pt stages). r resection was performed in all patients plus radio-and/or chemotherapy in all t stages. five patients ( %) died . years post transplant due to cardiovascular disease (n = ) and recurrent tumor disease (n = ). the -year survival rate was % for t and % for t rectal cancers, % for t and t each and % for the only t colon cancer. three anal neoplasms (one ain iii°, two anal cancers, pt and pt ; median age at diagnosis, years) developed on average . years after transplantation ( . % vs. . % in the general population) with a % -year survival rate. all patients were switched to rapamycin or everolimus after completion of primary therapy. conclusions. the incidence of anal but not of colorectal cancers in our transplant patient population differed from that of immunocompetent patients of corresponding age ( . % vs. . % and . % vs. . % tyrol tumor registry and . % seer). the -year survival rate was significantly decreased in the transplant group with t tumors ( % [rectum] and % [colon] vs. %). potential antineoplastic effects of rapamycin and overall less immunosuppression long-term may improve prognosis of colorectal malignancies following transplantation. background. solid-organ transplantation is the treatment of choice for terminal organ failure. due to the scarcity of organs, the mandate to utilize extended-criteria donors has dramatically increased over time and achievable results are good. this harbors the risk of transmission of infections from the donor to the recipient. aim. an overview on the magnitude of possible transmittable diseases that could accompany donor organs is given. possible preventive strategies are discussed. overview. the faith of an organism that is transmitted by a donor allograft depends on the virulence and the quantity of the transmitted pathogen and the site of infection and furthermore is impacted by prophylactic steps undertaken during donor procurement, organ perfusion, implantation, and post transplant course and lastly depends on the ability of the recipient to control the infection by the immune system. in the best case, the microorganism is cleared before it can do any harm; however, in the worst case scenario, transmitted pathogens can lead to fatality. there is a multitude of pathogens that can potentially be transmitted by an allograft including viruses, bacteria, fungi, and protozoa. there are microorganisms that can be transmitted by any type of graft and there are pathogens which due to a certain tropism can be transmitted by certain organs only. there are common pathogens such as cmv and ebv as opposed to extremely rare pathogens such as lyssa, west nile fever, or lymphochoriomenigitis virus. intracellular pathogens are preferably targeted by mhc restricted cytotoxic t-cell reaction. as in organ transplantation hla matching is in general not performed, donor-derived antigen-presenting cells cannot be recognised by recipient-specific cd + lymphocytes. therefore, either donor-specific cytotoxic t cells must function as counterparts or alternative tar-gets must be defined by the immune system. bacterial and/or fungal contamination must be considered in lung transplantation (bronchial stump), pancreas transplantation (duodenal segment), and small bowel allografts. cardiac, renal, and liver grafts can be considered sterile but can become contaminated during procurement. rare transmitted organisms are toxoplasma gondii, trypanosoma cruzii, and schistosoma mansoni. conclusions. when looking at allocation of latently virus-infected organs, they should be preferably given to recipients who have antibodies against the particular agent. in terms of bacterial or fungal contamination of graft and/or preservation solution, routine monitoring seems mandatory in order to assure quality. keeping blood and serum samples from donors should be carried out from an epidemiological and legal standpoint. bacterial and fungal prophylaxis should be used to prevent not only recipient-but also donor-derived pathogens. antiviral prophylaxis is standard for cmv and hbv, for all other viruses no final recommendations are available. methods. the aim of the current review was to investigate the impact of several risk factors on ptld. therefore, patients developing ptld after heart transplantation at our institution were screened. results. patients have been identified with ptld, all cases occurring during the last decade. mean age at ptld onset was . ± . years and time between transplant and development of ptld was . ± . months. there were ebv-associated ptlds, non-ebv-associated, cd -negative b-cell lymphomas, and t-cell lymphoma. immunosuppression at ptld onset was calcineurin inhibitor based (cyclosporine a, patients; tacrolimus, patients). initial immunosuppression included atg induction. six patients received perioperative antiviral prophylaxis with either valgancyclovir/gancyclovir (n = ) or acyclovir (n = ) in combination with anti-cmv hyperimmunoglobulin (n = ). two patients experienced a total of five episodes of acute rejection (ishlt ii°), all were treated with bolused steroids. four patients are still alive ( %), three of them in current remission of ptld, one patient is under therapy recently. median survival was months in survivors and . months in nonsurvivors. conclusions. these data show that ptld is associated with a high mortality rate. the majority of ptlds are ebvassociated. therefore, screening for ebv infection and prophylactic treatment may help to prevent a potentially fatal consequence of heart transplantation. background. late acute cellular rejection is associated with a decrease of survival and the development of cav. new immunosuppressice drugs have been introduced into clinical practice. everolimus, as one of these, has shown to be safe in cardiac transplantation. we report of our experience with everolimus in heart transplant recipients who developed late cardiac rejection. methods. patients with a history of previous rejection episodes who experienced cardiac rejection after at least months postoperative were switched to an everolimus, cyclosporine a, and corticosteroid-based immunosuppressive regime. all patients already received statins and antihypertensive medication before. everolimus, cyclosporine a trough levels and laboratory values were controlled monthly. drug administration was adapted to an everolimus trough level between and ng/ml, mean maintenance dosage was at . to . mg/day. death, safety, side effects, biopsy-proven acute rejection, laboratory values and blood levels were evaluated retrospectively. results. cardiac allograft recipients ( male, female), at a median of . days post orthotopic heart transplantation (ohtx) ( - ), received mg to . mg everolimus per day. in a follow-up period of at least month ( - ) mortality was %. the drug was well tolerated and no acute cellular rejection greater than grade a (ishlt grading) was observed after two month. in one patient raised cholesterol values and in two others elevated triglyceride levels were seen but were controlled with higher statine therapy. no obvious raised creatinine values were seen with certican ® . conclusions. in conclusion, conversion to an everolimus-based immunosuppressive regimen after late cardiac rejection is safe and effective. no major side effects were observed. p collagen iii in transplanted heartdonor or recipient derived m. pichler, b. tessaro, d. kniepeiss, r. kleinert, g. hoefler institute of pathology, medical university of graz, graz, austria background. transplantation of donor hearts is often associated with progressive development of interstitial myocardial fibrosis and alterations in composition and organisation of the extracellular matrix. changes in cardiac interstitial collagen network are thought to contribute to abnormal stiffness and loss of function of the myocardium. fibroblasts are the main producers of type i and type iii collagens, the major in-poster terstitial collagens found in the heart. in transplanted hearts, intragraft fibroblasts may consist of two cell populations. donor-derived fibroblasts preexist in donor organs, whereas hostderived fibroblasts may progressively immigrate as mesenchymal progenitors from the circulation to the allograft. purpose of the study. to determine the contribution of these two distinct fibroblast populations to progression of myocardial fibrosis, we studied endomyocardial biopsies over a time period of some years from a male patient who had received a heart from a female donor. methods. in these sex-mismatched patients two frequent genetic polymorphisms at the collagen iii locus were determined by polymerase chain reaction-based restriction enzyme digestion, both in donor allograft and in the corresponding explanted recipient heart. on the basis of differences of the collagen genotype in donor and recipient tissue, we selected endomyocardial biopsies by hematoxylin eosin staining covering an overall follow-up period of nine years since the transplantation event. immunohistochemistry, chromogene in situ hybridisation, and population-specific collagen expression using single nucleotide polymorphisms were used to determine the course of fibrosis. results. we developed an analytical system generally applicable to measure population-specific differences of collagen iii synthesis in transplanted organs. the amounts of interstitial collagen type i and type iii increased in a time-dependent manner within cardiac allograft. years after transplantation, a number of y-chromosome-positive recipient-derived cells consisted of noninflammatory spindle-shaped fibroblast-like cell types. conclusions. our data confirm the existence of a substantial number of fibrosis-mediating immigrated recipient-derived fibroblasts in cardiac allografts. furthermore, this suggests a potential, future therapeutic approach for reduction the cardiac fibrosis process. methods. between and a total of lung transplants ( patients) and six combined heart-lung transplants were performed at our center. immunosuppression consisted of atg induction, cyclosporine a, azathioprine, and steroids. all patients with rti underwent a meticulous micro-biological screening and underwent bronchoscopy with bronchiolo-alveolar lavage (bal) and transbronchial biopsies (tbb). rsv was detected from bal specimens by an immunoassay. results. a total of five lung recipients ( % of survivors) and one of the two alive heart-lung recipients developed rsv infection of the lung allograft. all cases were observed during an rsv epidemic in children in the area between october and may . the patients were - years old, all were females. one patient experienced two episodes. onset of rsv was median (range, - ) months post transplant. clinical symptoms included cough ( of ), rhinitis ( of ), and fever ( of ). deterioration of lung function occurred in six of the seven episodes with one deteriorating to respiratory failure requiring ventilator support. two individuals developed pulmonary infiltrates. in all patients, immunosuppression was significantly tapered, % required hospitalization and antibiotic therapy. the patient with rsv recurrence received inhalative ribavirin therapy during the second episode. all patients recovered and survival of this cohort was % after year and % at years after rsv infection. none of the patients developed bronchiolitis obliterans syndrome (bos) or rejection during follow-up. conclusions. rsv is a severe but benign complication following lung transplantation with a wide range of clinical presentations. routine use of antiviral treatment is not necessary; however, reduction in the level of immunosuppression is required. no long-term effects were observed in this cohort. a. stamatelopoulos , , s. guth , a. abrahim , g. m. marta , l. tsourelis , p. jaksch , c. konaris , s.taghavi , w. klepetko due to cni nephrotoxicity in a substantial number of ntx patients. we therefore studied the effects of a switch from a cni regimen (cni + mpa + p) to a dual regimen of sirolimus (srl) plus prednisolon (p) in pts with moderately impaired kidney function (s-crea, . + . mg%) due to either cni toxicity or clinical evidence for chronic allograft disease. pts received regimen consisting of srl mg dosage on day and from days to srl mg plus csa at half of maintenance dosage. from day on, pts received srl mg and csa was withdrawn. mpa or aza dose was continued halfed for to weeks. p was kept constant. target level for srl was - ng/ml. another pats received regimen consisting of srl mg and csa withdrawal on day and from days to srl mg was administered for a target level of to ng/ml. mpa or aza were continued halfed for to weeks, p was kept constant. with regimen there were dropouts due to adverse events, whereas with regimen only dropouts occurred. % of the patients showed a decrease of s-crea after -year observation period, % were unchanged and only % showed an increase. overall, there was a slight but significant increase of cholesterol and triglycerides, whereas other parameters were unchanged. conclusions. switch from cni containing immunosuppression to a dual regimen of sirolimus plus prednisolon results in an improved kidney function after years in the majority of pts. a regimen of sirolimus with target levels of to ng/ml and an overlap of immunosuppression shows a high rate of adverse events and is associated with a -fold dropout rate as compared to a regimen with a target level of - ng/ml. background. certain anatomical variations mainly concerning the portal system preclude living donor liver transplantation (ldlt). to the best of our knowledge, two left lateral segments with two arteries have never been transplanted so far. case report. a -month-old girl was diagnosed with endstage liver cirrhosis secondary to biliary atresia and therefore scheduled for ldlt. preoperative evaluation of the donor including ct with -d reconstruction revealed normal vascular supply of the liver with a left and a right hepatic artery. during donor operation, two tiny arteries with a diameter of mm for segments ii and iii were identified. they were found to arise from the left hepatic artery right behind the bifurcation of the proper hepatic artery, which made it impossible to preserve a common trunk. venous and portal venous reconstruction was performed in an end-to-end fashion. the left graft artery was directly anastomosed to the proper hepatic artery with / pds interrupted sutures using a microscope. the nd graft artery was revascularised with the help of saphenous vein from the recipient as interposition graft to the gastroduodenal artery but failed. therefore, the trunk of the inferior mesenteric vein was used for reconstruction with / pds with excellent outcome. under fk-based immunosuppression, postoperative course was uneventful with both arteries patent. conclusions. multiple arteries for the left lateral segments are not a contraindication for paediatric ldlt. the inferior mesenteric vein can be used as an interposition graft. combined renal-pancreas transplantation is an established treatment for patients with diabetic nephropathy, producing excellent results. we report on a patient who suffered from long-standing multiple sclerosis and underwent successful combined renal-pancreas transplantation. post transplant course was complicated by yeast intra-abdominal infection, which was treated with antifungal agents. using tacrolimus and sirolimus, both grafts are well functioning at two years and the patient is without any symptoms of disseminated encephalitis and does not require specific medication for multiple sclerosis. for diabetic patients suffering from multiple sclerosis, pancreas transplantation offers an excellent treatment option. whether normalisation of carbohydrate metabolism or chronic immunosuppression or both lead to complete response of multiple sclerosis is not clear. background. until recently, the peripheral blood stem cell (pbsc) donation procedure was used only infrequently among unrelated allogeneic donors. nowadays, both related and unrelated donors are expected to consider this alternative donation. to promote pbsc donation both safety and well-being of healthy unrelated volunteer donors must be protected and data are to be collected to establish the long-term safety of g-csf stimulation. methods. from to , pbsc aphereses on unrelated allogeneic donors have been carried out in our center. all pbsc donors were treated with µg of g-csf per kg twice a day from day - to day - . aphereses were performed using peripheral venous access on day and -if indicatedon day . since , all donors have annually received a questionnaire about their actual state of health and medication, as well as their physical and mental conditions. detailed questions concerned donors' anamnesis for epistaxis, bruises, thrombosis or embolia, as well as infections and fever, night sweat, and weight loss of unclear origin. results. male and female donors ( %) with an average age of . ( - ) years responded to the questionnaire. the observation periods were between and months (mean, . months) after g-csf stimulation. pbsc donors ( %, all male) reported that they had been severely ill during the observation period: one donor developed an exostosis of the th rib, one was operated on an umbilical hernia, one suffers from recurrent articular and muscle pain accompanied by night sweat and weight loss, one has a chronic compensated renal failure, one had diarrhoe and a common cold and suffered from fatigue, nausea, sleeping problems, and circulatory disorders. one female donor recognized dizziness and an increased tendency for bruises as well as paraesthesia in both arms. the disorders these donors reported occurred between and months after g-scf stimulation. all the other pbsc donors ( %) have never been severely ill or under medical observation. no donor had fever of unclear origin, phlebitis, thrombosis, or embolia, no donor recognized an increased tendency for epistaxis. three donors need medication they did not have before g-csf stimulation, which are to lower blood lipids and anti-inflammatory ones. one donor estimates that he is getting ill more easily than others, all the other donors feel themselves in best physical and mental condition. background. hematopoietic stem cell transplantation (hsct) has been successfully performed in patients with otherwise incurable malignant diseases. however, relapse after hsct is one of the main reasons for treatment failure and further therapeutic strategies with acceptable toxicity are warranted. since myeloablative (ma) conditioning after prior hsct has been associated with high treatment-related mortality (trm), reduced-intensity conditioning (ric) regimens have been developed as salvage therapies for these patients. so far, encouraging results have been achieved with ric; however, a direct comparison with standard conditioning has never been performed. therefore, we retrospectively analysed these two conditioning strategies in patients experiencing relapse after prior stem cell grafting. methods. we analyzed patients with relapsed disease (acute myeloid leukemia, n = ; indolent lymphoma, n = ; multiple myeloma, n = ; chronic myeloid leukemia, n = ; myelodysplastic syndrome, n = ; chronic lymphocytic leukemia, n = ; acute lymphocytic leukaemia, n = ; aggressive lymphoma, n = ; hodgkin's disease, n = ; ovarian cancer, n = ) after prior hsct who received either reduced-intensity or myeloablative conditioning for allogeneic hsct between and . ric consisted of fludarabine mg/m and total-body irradiation (tbi) of gy according to the seattle protocol (n = ) or alemtuzumab in combination with the beam regimen (n = ). myeloablative therapy consisted of cyclophosphamide (cy) and tbi of to gy (n = ), cy plus busulfan (bu) (n = ), c plus antithymocyte globulin plus bu (n = ), or bu alone (n = ). donors were syngeneic in , related in , and unrelated in patients. stem cell source was bone marrow in ( %) and peripheral blood in ( %) patients. for graft-versus-host disease (gvhd) prophylaxis, patients received cyclosporine a (csa) plus mycophenolate mofetil, csa plus methotrexate, mtx alone, and csa alone. results. all patients achieved complete hematopoietic engraftment by day after stem cell transplantation with complete donor chimerism. all patients conditioned with ric presented complete donor chimerism of t cells, myeloid progenitor cells, and nk cells to days after hsct. the in-cidence of acute gvhd was % and comparable in both groups consisting of grade i in patients, grade ii in , grade iii in , and grade iv in . fourteen patients died after ma conditioning of acute gvhd (n = ), infections (n = ), or severe toxicity (n = ), while only one patient died due to infection after ric. probability of transplant-related mortality (trm) at year after hsct was with % significantly (p = . ) higher in patients given myeloablative conditioning compared to % after ric. incidence of therapy requiring chronic gvhd was with % versus % significantly (p = . ) higher in patients who received ric. response rates were comparable between patients who received ric or ma conditioning ( % versus %). relapses occurred in % of patients after ric and % after ma conditioning. after a median follow-up of . (range, - ) months ( %) of patients of the ma group and ( %) of the ric group are currently alive. probability of survival at and years after hsct is with % versus % and % versus % significantly (p = . ) higher after ric than after myeloablative conditioning. conclusions. allogeneic stem cell transplantation is a highly effective treatment option in patients relapsing after prior hsct. durable hematologic engraftment and sustained complete remissions can be achieved in patients with otherwise poor prognosis. since transplant-related mortality of dose-reduced conditioning is in comparison to myeloablative hsct considerably lower, overall survival can be significantly improved with this new treatment modality. background. patients with comorbidities such as organ dysfunctions or preexisting infections experience a high treatment-related mortality which makes them ineligible for conventional conditioning therapy. for these patients, reduced-intensity conditioning (ric) is an option which offers the benefits of an allogeneic transplant with lower extramedullary toxicity. we report on pediatric and young adult patients who were considered for ric because of severe cumulative pretreatment or substantial comorbidities treated at our institution between and . methods. eight patients (median age, years; range, - years; m, ; f, ) diagnosed with aml (cr , n = ; cr , n = ), all (cr , n = ), alps (n = ), and relapsed ewing's sarcoma (n = ). of patients were planned to receive ric, in one patient the regime was changed during conditioning due to an acute viral infection. all patients received fludarabine (n = , mg/m ; n = , mg/m ) combined with melphalan (n = , - mg/m ), busulfan (n = , mg/kg), or total-body irradiation (n = , gy). patients received atg ( - mg/kg), two patients campath ( mg/m ). post-transplant immunosuppression consisted of cyclosporine a in all patients combined with mycophenolate mofetil (n = ) or methotrexate (n = ). of patients received hla identical grafts (sibling, n = ; hla identical mother, n = ; mud, n = ), one patient received a c-locus mismatched graft. stem cell sources were bone marrow in patients and peripheral blood stem cells in containing a median of . × cd + cells per kg of body weight of the recipient. results. all patients had primary engraftment; the median time to neutrophil recovery (n > . × /l) was days. complete donor chimerism as evidenced by vntr was achieved in median on day + (+ to + ). acute mild graftversus-host disease (gvhd) of the skin occurred in of patients and responded to prednisolone; one patient required additional immunosuppression with mmf. one patient progressed to extensive chronic gvhd; one patient developed chronic gvhd of the skin, another patient shows clinical evidence of chronic gvhd of liver and gut. the remaining patients showed no gvhd at all. of patients were positive for one or multiple viruses on routine viral pcr monitoring, requiring virostatic treatment. no treatment-related mortality occurred. the patient with ewing's sarcoma died months posttransplant from tumor progression; the patient with extensive chronic gvhd died of sepsis on day + . all patients with aml are in remission. the patient with alps is still positive for various autoantibodies. patients developed a posttransplant macrophage-activation syndrome (mas). all of them required a stem cell boost because of pancytopenia. conclusions. ric followed by allogeneic hsct was successful in terms of achieving primary engraftment and complete donor chimerism as well as avoiding transplant-related mortality in all patients; in our patients with myeloid malignancies it was also successful in terms of maintaining stable remission. as for posttransplant problems, we encountered acute gvhd of the skin in of patients, three of whom developed chronic gvhd; and viral infections in of patients, three of whom developed mas and eventually required a stem cell boost. background. due to an aggressive course mantle cell lymphoma is characterized by poor prognosis with a median survival of years and only - % long-time survivors. recent improvements in therapy have been made by combined immunochemotherapy and intensification with high-dose chemotherapy. methods und results. ten patients ( female, male) with a median age of ( - ) years were treated with rituximab plus chop for four or six cycles. furtheron, patients received or cycles of claeg-d ( days cladribine . mg/kg, ara-c . g/m , etoposid mg/m , and on day daunoxome mg/m ) including stem cell collection. as a result, out of newly diagnosed patients reached cr and pr. out of patients treated after first relapse, reached again cr and pr. autologous transplantation was performed at a median of ( - ) months after diagnosis. high-dose conditioning consisted of beam chemotherapy plus the addition of doses of rituximab (days - and - of the conditioning regimen and days + and + after transplantation). four patients could not receive the rituximab therapy at days + and + due to complications. a median number of . × cd + cells ( . - . ) per kg were reinfused. all patients had a short haematologic regeneration time (granulocytes, < . g/l day + [ - ]; platelets, day + [ - ]) and received a median number of erythrocyte ( - ) and platelet ( - ) concentrates. all patients suffered from mucositis grade i-ii, of had emesis grade i-ii. infectious complication of short duration appeared in patients ( fuo, pneumonia, and sepsis) early after transplantation. one patient developed late complications (hypothyreosis on day + and sarcoidosis on day + ). following transplantation, all patients reached clinical and molecular cr lasting for a median time of ( - ) months. two patients relapsed and months after transplantation. despite continuous salvage immunochemotherapy, one of these patients died months after transplantation, the other one is still in pr (+ months). median observation periode after diagnosis is ( - ) months. conclusions. treatment intensification with immunochemotherapy and high-dose consolidation is accompanied by acceptable toxicity and seems to be an effective treatment procedure for mantle cell lymphoma. background. bone marrow transplantation (bmt) together with costimulation blockade can reliably induce mixed chimerism and tolerance. in recent studies, we showed that regulation by cd + cd + t cells plays an important role in this model. stimulation of cd + cd + t regs by an anti-cd mab can inhibit and reverse the outbreak of certain autoimmune diseases, and the maintenance and function of t regs was demonstrated to critically depend on il- . we thus investigated if stimulation of regulatory cells by anti-cd or interleukin- facilitates bm engraftment and reduction of tbi in our model. methods. c bl/ mice received a total-body irradiation (day - ) of gy or . gy, approximately × fully mismatched balb/c bone marrow cells (day ) and costimulation blockade consisting of anti-cd mab (mr , day ) and ctla ig (day + ). additional groups received further treatment with (n = - per group): ( ) anti-cd mab ( µg, day to + ), ( ) il- ( ie/day, day - ), ( ) rapamycin ( . mg/kg/day, day - ), and ( ) rapamycin and il- (day - ). multilinage chimerism and skin graft survival were followed for more than days. results. with our standard protocol, of ( gy) and of ( . gy) mice developed long-term multilinage chimerism and accepted donor skin grafts permanently. while the majority of mice treated with anti-cd and gy ( of ), rapamycin plus . gy ( of ), and il- and rapamycin plus . gy ( of ) developed multilinage chimerism and longterm skin graft survival, groups treated with anti-cd plus . gy tbi ( of , p < . ) and il- plus . gy ( of , p = . ) showed markedly reduced rates of chimerism and tolerance. we investigated if these negative effects might be correlated with a cytokine shift caused by anti-cd treatment, but we did not observe such a shift towards th or th (as measured on day post-bmt). conclusions. unexpectedly neither anti-cd nor il- had a positive effect in this model, in some groups anti-cd treatment even showed a negative effect. thus, other strategies for augmenting the effect of regulatory cells need to be developed. the role of minor antigen disparities for the induction of mixed chimerism and tolerance through bmt plus costimulation blockade s. bigenzahn , i. pree , p. nierlich , e. selzer , f. mühlbacher , t. wekerle long-term donor skin grafts looked macroscopically intact in the b .d group but showed shrinking and scabs in the balb/c group. conclusions. minor antigen disparities pose a substantial barrier for the induction of mixed chimerism and tolerance. for increased clinical relevance, tolerance models should preferably use strain combinations including major plus minor mismatches. p immunohistochemical and confocal analysis of pancreatic tetranectin d. pirkebner , m. hermann , a. draxl , w. mark , r. margreiter , p. hengster background. islet transplantation is not yet widely used in part because of the shortage of human islet cells. gaining detailed knowledge of the physiological basis governing the processes of differentiation of pancreatic stem or progenitor cells that have the capacity to self-renewal and to generate differentiated beta cells is instrumental for the ambitious goal of engineering new pancreatic islets in order to cure type i diabetes. the aim of our study was to cultivate and characterize a pancreatic cell population expressing tetranectin (tn). the ability of tn to bind plasminogen indicates that it may have a role in regulating pericellular proteolysis and proteolytic activation of latent forms of metalloproteinases and growth factors. methods. islets were isolated according to the method of ricordi et al. ( ) using a continuous ficoll gradient. immunohistochemistry and immunofluorescence were performed with a monoclonal antibody against human tetranectin (amino acids - of human tetranectin monomer; antibodyshop, grusbakken, denmark). to determine in vitro cell proliferation, cells were labeled with brdu (roche, basel, switzerland). results. we describe the localization of tn-positive cells in the human pancreas and their growth in vitro. interestingly, individual positive cells are present within the exocrine acini. we were able to isolate human and murine islets and cultivate these tetranectin-positive cells under adherent and nonadherent conditions as shown by immunohistochemistry and confocal immunofluorescence. the possibility to culture these cells is a first step towards their better characterisation. conclusions. together with its above mentioned ability to bind plasminogen-like hepatocyte growth factor and tissuetype activator, tn may thereby play an important role in the survival of islets after islet transplantation. as we could show, tn positive cells can be isolated and maintained in culture after human islet isolation, thereby providing the possibility to further clarify their role and function in vivo as well as in the course of islet transplantation. p fty interferes with effector functions and downregulates protein expression of s p and s p in human dendritic cells tures with fty /fty -p-treated dc illustrated a cytokine production profile with a lower ifn-? ( % vs. % relative reduction) and a higher il- ( % vs. % relative increase), indicating a shift from th toward th differentiation as previously evinced for s p. dc yields, phenotypic differentiation into idc and mdc (besides a minor reduction in cd surface expression), as well as the investigated mechanisms of idc antigen uptake (bacterial phagocytosis, mannose receptor-mediated, and fluid-phase endocytosis), were not affected at therapeutically relevant concentrations. conclusions. we conclude that treatment of human dc with fty and fty -p interferes with dc effector functions that are essential for dc to serve their pivotal duty as professional antigen-presenting cells and that dc can therefore be added to the potential list of target cells of fty . impairment of dc migration and th -priming capacity due to downmodulation of dc-expressed s p and s p might represent a new aspect in the overall mechanism of action and hence contribute, in part, to fty -mediated immunosuppression. the ibal-fresenius: bioartificial liver innsbruck (ibal) utilizing fresenius standalone, rotating bioreactor m. wurm, v. lubei, p. hengster in order to establish a suitable environment for cultivation of hepatocytes serving as bioartificial liver (bal), we were testing and further developing fresenius standalone, rotating bioreactor. to create optimal conditions for cultivation of hepatocytes, a special environment of nearly gravity-free, low shear force and high mass transfer is needed. furthermore, basic parameters like stability of heating, gas exchange, and sufficiency of nutrition have to be evaluated allowing utilization of various breeding chambers. in summary, we have established a standalone bioreactor capable of quick mass transfer between small and big chambers and external media supply, in nearly gravity-free environment minimizing shear force thus allowing for cultivation of various cell types. background. laparoscopic donor nephrectomy is a less invasive alternative to open nephrectomy for living kidney donation. this study presents the results of the first laparoscopic donor nephrectomies in our center. methods. from june to may , patients underwent laparoscopic donor nephrectomy for living-related renal transplantation. patient demographic, intraoperative, and postoperative parameters and complications, as well as renal allograft outcome, were evaluated. results. patients ( female and male) donated their left kidney. the mean donor age was years ( - years). the mean surgical time was ± minutes. mean warm ischemia time was ± seconds. patients could be discharged from hospital after a mean time of . ± . days. in four patients ( . %) conversion to open surgery had to be performed. reasons for conversions were lack of operative progression in two cases, in one case venous bleeding, and in one case lesion of the renal artery. there were no reoperations required in the donors. in the recipients, ( . %) delayed graft functions and ( . %) primary nonfunction were observed. mean serum creatinine level in the recipients was . mg/dl months after transplantation. conclusions. laparoscopic live donor nephrectomy is safe for the donor and the transplant kidney. we believe that offering this technique for living renal donation can safely and effectively increase the pool of donor organs available to patients with end-stage renal disease. background. liver transplantation is the treatment of choice for end-stage acute and chronic liver failure. some liver diseases are associated with diseases of the intestinal tract such as primary sclerosing cholangitis and inflammatory bowel disease. moreover, post transplant immunosuppressive agents might cause colonic diseases and there is an abundance of opportunistic pathogens that can manifest in the large intestine. aim. we retrospectively analyzed the incidence and spectrum of colonic disorders in a cohort of liver recipients and determined the impact of these complications on survival. methods. a total of consecutive lts in individuals were performed between and at the mayo clinic, jacksonville, florida. standard immunosuppression consisted of tacrolimus, mycophenolic acid, and rapidly tapered steroids. results. there were patients transplanted for psc and were also diagnosed with inflammatory bowel disease (ulcerative colitis, n = ; crohn's disease, n = ), with eight having undergone colonic resection prior to lt. in four patients, colitis persisted post transplant. six patients had a history of colonic resection for malignant (n = ) or infectious diseases. five patients had pre-transplant endoscopic polypectomy. combined colon resection and transplantation were done in patients; one with peritonitis and multiple colonic perforations during retransplantation and the other for ischemic colitis leading to fulminant liver failure. in another case a preexisting transverse colostomy had to be reinforced. there were cases of clostridium difficile-associated enterocolitis. nine patients developed cmv gastrointestinal complications with three cases of colitis, one leading to perforation, intra-abdominal sepsis and death. two patients developed sigmoid diverticulitis and one appendicitis. colonic polyps were endoscopically removed in seven patients and three patients were diagnosed with colorectal cancer (one cecal, two rectal cancers), which all were surgically treated. chronic unexplained diarrhea was observed in fifteen patients, which led to withdrawal of mycophenolic acid. one patient developed a hemorrhage of the terminal ileum/cecal region in the course of intra-abdominal sepsis and was treated by endovascular embolization of the ileocolic artery. four patients had ongoing ulcerative colitis. one herpetic rectal ulcer and two perianal hsv-associated lesions were diagnosed. two patients developed hemorrhoids requiring surgical interventions, and two patients had perianal fistulas. conclusions. the frequency of colonic disorders in our series was higher than expected, with infections accounting for majority of cases. the high incidence of clostridial colitis warrants improvement in screening and preventive measurements. screening for polyps pre-transplant and annually post-transplant might be recommended. background. bartonella has been identified as causative agent of cat scratch disease but is also inflicted in other diseases in the immunocompromised host. case reports. we describe two cases of bartonella henselae-associated diseases in liver transplant recipients who both had contact with cats. the first recipient developed localized skin manifestation of bacillary angiomatosis in association with granulomatous hepatitis. he tested positive for igg antibodies against bartonella henselae. the second patient developed axillary lymphadenopathy, with biopsy showing necrotizing granulomatous inflammation and pcr studies were positive for bartonella henselae dna. her serology for bartonellosis showed a fourfold rise in antibody titers during her hospitalization. both patients responded to treatment with azithromycin in combination with doxycyclin. these were the only cases within a series of liver transplants in patients performed during a four-year period. conclusions. although bartonellosis is a rare infection in lt recipients, one should consider this disease in patients presenting with fever, cns symptoms, skin lesions, lymphadenopathy or hepatitis in particular if contact with cats is reported. background. new immunosuppressive protocols and advanced surgical technique resulted in a major improvement in the outcome of pancreatic transplantation. by reducing the incidence of immunological complications using intensified immunosuppressive protocols, the incidence of some infections, in particular complicated fungal infections, might increase. methods. enteric drained whole-pancreas transplants (ptx) performed at the innsbruck university hospital between march and october were retrospectively analysed. prophylactic immunosuppression consisted of atg induction, tacrolimus, mmf, and steroids for the majority of cases. perioperative antimicrobial prophylaxis consisted of amoxicillin/clavulanic ( ptx), pipercillin/tazobactam ( ptx), and others ( ptx). patients additionally received fluconazole. results. actuarial patient, pancreas and kidney graft survival at one year were . %, . % and . %, rejection rate was %. within this series, a total of patients developed invasive fungal infections. of those, four had aspergillosis, one zygomycosis, and the remaining ten were caused by yeast. two patients had aspergillosis and later pulmonary infection with candida albicans and candida glabrata. the zygomycosis was the only fungal infection that was diagnosed post mortem and this patient had received pretreatment with caspofungin for non-albicans candida wound infection. one patient died due to aspergillosis following his second pancreas retransplant. three cases of aspergillosis were successfully treated using liposomal amphotericinb in one and a combination of caspofungin and voriconazol in two cases. this combination was also used in a patient who developed intra-abdominal infection with candida krusei. the remaining infections were due to candida albicans including six cases of intra-abdominal infection, one urinary tract infection, and one mucocutaneous candidiasis. type ii diabetics were found at increased risk for fungal infection. conclusions. fungal infections represent frequent and life-threatening complications after ptx. they are amongst the most common causes of graft loss and death. non-albicans candida strains are increasingly isolated and the incidence of filamentous fungal infections has increased during the study period parallel to a decreasing rejection rate. c bl/ (h- b ) mice received a total-body irradiation (tbi, d- ) of gy and costimulation blockade consisting of anti-cd mab ( mg, d ) vß + cd -cells, p < . ). donor skin acceptance fty is the first agent in a new class of immunomodulators termed sphingosine- -phosphate (s p) human dc, which are known to express mrna for s p , s p , s p and s p , have not been described so far. methods. to elucidate for the first time the influence of s p receptor agonists on human monocyte-derived dc (mo-dc), we used therapeutically relevant concentrations ( - ng/ml) of fty and its phosphorylated metabolite fty -p and investigated their effects on dc surface marker expression (lineage markers, costimulatory and adhesion molecules, chemokine receptors), protein levels of s p receptors and dc effector functions: antigen uptake ccl /elc; or fty -p-treated mdc ( % vs. %; untreated dc, %) - datenkonvertierung und umbruch: manz crossmedia druckerei ferdinand berger & söhne gesellschaft m. b. h., horn, Österreich. -verlagsort: wien. -herstellungsort: horn. printed in austria p. b. b. / erscheinungsort: wien / verlagspostamt wien -fachkurzinformation rapamune mg bzw. mg überzogene tablette; rapamune mg/ml bzw. mg/ ml bzw. mg/ ml lösung zum einnehmen wirkstoff: sirolimus zusammensetzung: tablette enthält mg bzw. mg sirolimus. ml lösung enthält mg sirolimus. beutel zu ml bzw. ml enthält: mg bzw. mg sirolimus. sonstige bestandteile: tablettenkern: laktose-monohydrat, macrogol, magnesiumstearat, talkum, tablettenüberzug: macrogol engmaschige Überwachung der sirolimus-talspiegel im vollblut bei: leichter bis mittelgradiger leberfunktionsstörung; gleichzeitiger verabreichung starker cyp a -induktoren oder -inhibitoren sowie nach deren absetzen; bei absetzen oder deutlicher dosisreduktion von ciclosporin. begrenzte exposition gegenüber sonnen-und uv-strahlung bei patienten mit einem erhöhten risiko für hautkrebs. antimikrobielle prophylaxe gegen pneumocystis carinii pneumonie während der ersten monate nach der transplantation sowie eine zytomegalievirus (cmv)-prophylaxe über monate nach der transplantation ( insbesondere für patienten mit einem erhöhten risiko für eine cmv-erkrankung ) empfohlen. in kombination mit einem hmg-coa-reduktaseinhibitor oder fibrat Überwachung auf entwicklung einer rhabdomyolyse und anderen nebenwirkungen dieser präparate. bei kombinierter gabe mit ciclosporin nierenfunktion überwachen, ggf. bei erhöhten serumkreatininspiegeln eine angemessene dosisanpassung erwägen. vorsicht bei gleichzeitiger anwendung von anderen substanzen, die bekanntermaßen eine schädigende wirkung auf die nierenfunktion haben erhöhte laktat-dehydrogenase (ldh), arthralgie, akne, infektion des harntraktes gelegentlich: pankreatitis, lymphom/lymphoproliferative erkrankung nach transplantation, panzytopenie. die immunsuppression erhöht die anfälligkeit, lymphome oder andere bösartige neubildungen, vor allem der haut, zu entwickeln fachkurzinformation zu inserat von umschlagseite p liver transplantation for patients with hepatitis b-related liver disease: a single-center experience l. hinterhuber, i. w. graziadei background. liver transplantation (lt) is the only effective therapy for end-stage liver disease due to hepatitis b (hbv). before introduction of passive immunoprophylaxis with hepatitis b immunoglobulin (hbig) and new antiviral nucleoside analogues, hepatitis b recurrence was seen in the majority of the patients resulting in an inferior graft survival. in this study we analyzed the different clinical courses of hbv recurrence, the impact of hbv recurrence on patient survival, and potentially contributing factors for long-term outcome of hbv patients after lt.methods. between and , out of patients ( m, f; mean age, years) were transplanted secondary hbv cirrhosis at our center. the mean follow-up was months (range, - months). immunosuppression (is) consisted of cya/fk , prednisolone and/or azathioprine/mmf. fourteen patients received no hbig (prior to ), patients received hbig alone, and patients hbig in combination with lamivudine (lam).results. the actuarial overall -, -, -year survival rates were %, %, and %, comparable to those of other indications. patients with combined prophylaxis showed the best survival rates ( %, %, %), compared to patients treated with hbig ( %, %, %) and patients without treatment ( %, %, %). five patients required reltx, one patient two reltx. in total, patients ( . %) developed recurrent hbv infection after lt: % ( of ) in the non-hbig group, % ( of ) in the hbig mono, and % ( of ) in the combined prophylaxis group. four of the five patients in the hbig/lam group were hbv dna positive prior to lt, two presented with lam mutants. hbv recurrence, however, did not negatively impact patient outcome. all patients with recurrent disease were treated with antivirals (famcyclovir, lam, adefovir). forty-seven percent of patients responded to the treatment and remained hbv dna negative. only one patient was retransplantated due to hbv recurrence. no possible risk factors for overall survival were found to be significant.conclusions. our study showed that patients with combined hbig/lam prophylaxis had excellent long-term survival. recurrent hbv in the allograft could be effectively treated in the majority of patients and did not influence longterm survival. background. liver failure is associated with reduced synthesis of clotting factors, consumptive coagulopathy, and platelet dysfunction. the aim of the study was to evaluate the effects of liver support using the molecular adsorbent recirculating system (mars) on the coagulation system in patients at high risk of bleeding.methods. mars treatments in patients with acute liver failure (n = ), acute on chronic liver failure (n = ), sepsis (n = ), liver graft dysfunction (n = ), and cholestasis (n = ) have been studied. standard coagulation tests, standard thromboelastography (teg), heparinase-modified and abciximab-fab-modified teg were performed immediately before and minutes after commencement of mars and after the end of mars treatment. to all patients, prostaglandin i was administered extracorporeally. patients additionally received unfractioned heparin.results. three moderate bleeding complications in three patients, requiring - units of packed red blood cells, were observed. all were sufficiently managed without interrupting mars treatment. although there was a significant decrease in platelet counts (median, g/l; range, - to g/l) and fibrinogen concentration (median, mg/dl; range, - to mg/dl) with a consecutive increase in thrombin time, the platelet function, as assessed by abciximab-fab-modified teg, remained stable. mars did not enhance fibrinolysis.conclusions. mars treatment appears to be well tolerated in patients with marked coagulopathy due to liver failure. although mars leads to a further decrease in platelet count and fibrinogen concentration, platelet function, measured as contribution of the platelets to the clot firmness in teg, remains stable. according to teg-based results, mars does not enhance fibrinolysis. methodik. ein jahre alter patient war vor jahren aufgrund einer post-hepatitischen zirrhose (phcc) lebertransplantiert worden. nun war es im verlauf der letzten zeit zu einer deutlichen gewichtszunahme gekommen, sodass der pati- key: cord- -v iayczy authors: nan title: publication only date: - - journal: bone marrow transplant doi: . /bmt. . sha: doc_id: cord_uid: v iayczy nan introduction: natural killer (nk) cells are cytotoxic lymphocytes of the innate immune system capable of targeting virally transformed and tumor cells without prior sensitization. they have shown to be effective at targeting malignant cells without being direct effectors of graft-versus-host disease in both a transplantation setting and as an nk cell therapy alone. there have been numerous clinical trials using nk cells as an adoptive immunotherapy in both hematological and solid malignancies. these have shown nk cell immunotherapy to be safe however efficacy has been inconsistent. material (or patients) and methods: in order to strive towards an effective and robust clinical therapy we hypothesize that proliferation, persistence and activation of nk cells in vivo is fundamental for the development of a clinically relevant cellular product that will be able to target tumor cells efficiently in patients. our group has previously shown that frozen cd + cells isolated from umbilical cord blood (ucb) are the most favorable source of stem cells to differentiate functional nk cells in vitro. producing large cell numbers capable of secreting high levels of ifn-g and killing leukemic cell lines in vitro. we therefore tested whether the differentiated nk cells are capable of proliferating and getting activated following further cytokine stimulation and if they are able to persist in vivo to target and eliminate leukemic cells. we also assessed if the cells could be frozen and thawed whilst still maintaining functionality to allow for the production of a readily available off-the-shelf cellular product. results: we found that the differentiated nk cells could respond to interleukin- and proliferate in vitro to the same degree as adult peripheral blood nk cells. this suggests that the cells can further respond to cytokines and proliferate once injected into the patient. the function of the differentiated cells was unaffected by cryopreservation allowing for multiple infusions at different time points of the same nk product. the differentiated nk cells could kill leukemic cells and more importantly could persist for longer and in higher numbers in vivo over other sources of nk cells, indicative of improved survival post infusion. additionally we assessed pre-activation of the cells by priming the differentiated nk cells using ctv- lysate. in comparison to resting controls this consistently led to higher levels of killing of patient leukemic blasts in vitro, however this activation step was not required to observe killing of patient aml blasts in vivo. this indicates that the cells would not require prior activation before infusion resulting in a cellular therapy that was more economical and more easily translatable to the clinic. conclusion: in conclusion, we are able to generate and cryopreserve nk cells from ucb hematopoietic stem cells in high numbers allowing for multiple infusions that are highly cytotoxic towards tumor cells. these cells can further proliferate in response to additional cytokines and have a clear survival advantage in vivo over other sources of nk cells. this provides the potential for the development of a robust nk cell immunotherapy which can not only prevent infection post transplantation but also target malignancies in their own right to prevent relapse. disclosure of interest: none declared. long-term sustained graft-versus-lymphoma effect of donor lymphocyte infusion p. olioso ,* , s. santarone , a. natale , g. papalinetti , r. giancola , t. bonfini , p. accorsi , p. di bartolomeo hematology, bmt center, o spedale c ivile, pescara, italy introduction: allogeneic hematopoietic stem cell transplantation (sct) is a potentially curative option for patients with lymphoproliferative disorders such as hodgkin lymphoma (hl) and non-hodgkin lymphoma (nhl). for patients who relapse or have refractory disease after sct, the use of donor lymphocyte infusion (dli) represent an important part of the strategy to enhance graft-versus-lymphoma effect (gvl) . in this study we evaluate how dli is able to control the disease in patients with lymphoma who relapse or have persistent disease after sct. material (or patients) and methods: between february and july , patients ( males) with advanced lymphoma (high grade nhl n = , hl n = ) previously treated with sct received dli as treatment for progressive or relapsed disease. the median age was ( - ) years at time of sct. three of them underwent previous autologous sct. all patients were transplanted with pbsc, from hla identical donor and from an unrelated one. conditioning was myeloablative in and reduced intensity in . gvhd prophylaxis included cyclosporine and short course methotrexate in all patients. results: no patient developed dli-related aplasia or acute gvhd after dli. limited chronic gvhd was observed in only patients. three patients had no response to dli and died for progression of their lymphoma. four patients had sustained complete response with persistent and continued disappearance of the lymphoma. the cumulative response rates after dli to treat relapse or persistent disease after sct were / ( %) and / ( %), respectively. no correlation was observed between cd + cell dose infused and disease response. both patients who developed dli-related chronic gvhd had complete response. all four patients who achieved complete response after dli are still alive and disease-free after a median follow-up of ( - ) years. conclusion: these data support the existence of a clinically significant gvl effect in high-grade nhl and aggressive hl suggesting that dli is an effective treatment able to induce long-term remission without severe gvhd in patients with very poor prognosis. a dli program should always be considered for patients with lymphoma who show persistent disease or relapse after sct. disclosure of interest: none declared. human platelet lysate pathogen reduced through additive-free short-wave uv light irradiation retains its optimal efficacy for the expansion of human bone marrow mesenchymal stem cells s. viau , l. chabrand , s. eap , f. goudaliez , c. sumian , b. delorme ,* biotherapy division, macopharma, mouvaux, transfusion division, macopharma, tourcoing, france introduction: we recently developed and characterized a standardized and clinical grade human platelet lysate (hpl) that constitutes an advantageous substitute for fetal bovine serum for human mesenchymal stem cell (hmsc) expansion required in cell therapy procedures, avoiding xenogenic risks (virological and immunological) and ethical issue. because of the progressive use of pathogen reduced (pr) labile blood components, we evaluated the impact of the novel procedure theraflex uv-platelets for pathogen reduction on hpl quality (growth factors content) and efficacy (as a medium supplement for hmsc expansion). this technology is based on short-wave ultraviolet light (uv-c) and has the main advantage not to need the addition of any photosensitizing additive compounds (that might secondary interfere with hmscs). material (or patients) and methods: we applied theraflex uv-platelets procedure on fresh platelet concentrates (pcs) suspended in platelet additive solution and prepared hpl from these treated pcs. we compared the quality and efficacy of pr-hpl with the corresponding non-pr ones. results: we showed no impact on the content in cytokines tested (egf, bfgf, pdgf-ab, vegf and igf) and a significant decrease in tgf-b (- %, n = , p o . ). we performed large scale culture of hmscs during passages and showed that hpl or pr-hpl at % triggered comparable hmsc proliferation than fbs at % plus bfgf (n = ). moreover, after proliferation of hmscs in hpl or pr-hpl containing medium, their profile of membrane marker expression, their clonogenic potential and immunosuppressive properties were maintained, in comparison with hmscs cultured in fbs conditions. the potential to differentiate in adipogenic and osteogenic lineages of hmscs cultured in parallel in the conditions remained also identical. conclusion: in conclusion, we demonstrated the feasibility to use uv-c treatment to subsequently obtain pathogen reduced hpl, while preserving its optimal quality and efficacy for hmsc expansion for cell therapy applications. introduction: in patients with myeloma, it might be necessary to perform more than one autologous hematopoietic stem cell transplantations (asct). at our centre, we have been have asked to refreeze parts of autologous hematopoietic stem cells (hsc) units with high cell numbers, that already have been stored for several years. the idea is to use the refrozen part of the cells if there is a need for re-transplantation. this raises the question of the viability of the graft. material (or patients) and methods: the aim of the study was to analyze the effect of refreezing on hsc regarding the amount of cd + cells, cell viability, apoptosis and regeneration ability. hsc products from ten deceased patients, stored for more than years, were analyzed regarding the amount of cd + cells, cell viability measured with -aad and apoptosis measured with annexin v. we used flowcytometry for the analysis. the regeneration ability of hsc was analyzed with colony forming units. results: refreezing of hsc product, did not affect the amount of cd + cells significantly. however a significant decrease in cell viability, with a mean of %, was seen. the number of apoptotic cells was significantly increased with %. the regenerative ability, measured as colony forming units, decreased significantly with a mean of %. conclusion: it can be concluded, that refreezing of hsc product decreases the number of viable cd + cells with approximately %. disclosure of interest: none declared. introduction: umbilical cord blood (ucb) is largely employed as an alternative source of hematopoietic stem cells in the treatment of hematological diseases. one of the limitations to use ucb is the relative low content of hematopoietic stem cells. the main parameters used in umbilical cord blood banks include total nucleated cell (tnc) count, percentage of cd + cells and collection volume. the leuven cord blood bank (lnbb) collects units from a network of maternity wards since using the 'in utero' collection method both for vaginal deliveries and cesarean sections. since the lnbb has updated the ucb databank to register mode of delivery of every ucb collection. material (or patients) and methods: this analysis compares ucb obtained from vaginal deliveries and caesarean sections ( / - / ) . the volume of the collected cord blood was compared as well as quality parameters such as cell numbers (tnc) before processing and cd + cell content post-processing, by an independent samples t-test. the level of significance was set for a p-value o . . results: mean volume of ucb collected after cesarean section was significantly higher compared to ucb volumes collected after vaginal deliveries (po . ) (table ). no statistically significant difference between quality markers such as cell count and cd + count (table ) could be detected. conclusion: cord blood collection after cesarean route of delivery showed a larger collection volume than vaginal delivery. a possible explanation could be that, as the baby is placed above the placenta after the cesarean section, clamping the cord above the placenta possibly causes a downward flow of blood into the umbilical cord, which may result in a greater volume of ucb collected (nunes & zandavalli, ) . despite the difference in collected volume, ucb units after cesarean sections seem to contain similar progenitor content compared to vaginal deliveries as results are expressed as mean ± sd. s described in the literature (askari et al, ) . in utero ucb collections after cesarean sections are feasible, but further studies are needed to investigate the impact of the type of delivery on quality of the cord blood units. references: nunes r., zandavalli f. association between maternal and fetal factors and quality of cord blood as source of stem cells. rev bras hematol hemater. ; ( ) results: in this overview were discussed overall survival (os), disease free survival (dfs) in different groups, the factors for successful control of the underlying disease, the risks and therapy related complications. in the allogeneic group are considered causes of early death: transplant related mortality (n = ), death from relapse (n = ), death from gvhd ( n = ). the results showed an increase number of both autologous and allogeneic transplantsa global trend. the low trm proves that this high-tech procedure leads to increase os in some patients and increase their chances for a long dfs. disclosure of interest: none declared. pneumomediastinum case secondary to pneumocystis jirovecii infection as late complication in an allogeneic hematopoietic stem cell tranplant patient g. akyol ,* , e. ermiş turak , e. yıldızhan , k. demir , m. Çetin , l. kaynar hematology, erciyes univercity, kayseri, turkey introduction: pneumocystis jirovecii pneumonia (pjp) is important cause of mortality and morbidity in immunocompromised patients with especially acquired immunodeficiency syndrome (aids),and others like hematopoietic stem cell tranplantion or hematologic malignancies. even though pneumothorax is an expected complication during pjp, pneumomediastinum is quite rare. we report a case of spontaneous pneumomediastinum during treatment of late onset p.jirovecii pneumonia in an allogeneic hematopoietic stem cell tranplant recipient. material (or patients) and methods: case report -year-old male patient who underwent allogeneic hematopoietic stem cell tranplantation sixteen months ago, admitted to bone marrow transplantation clinic for shortness of breath. there was widespread rhonchi in bilateral lungs. patient had no fever with arterial blood gase oxygen level measured as sao %. computed tomography showed significant consolidation lung areas of in both upper lobe ( figure ). pneumocystis jirovecii pneumonia diagnosis was considered so that high tmp-smx doses intravenously with mg/kg methylprednisolone was started. piperacillin-tazobactam added to therapy as bacterial pneumonia can not be ruled out. in the fifth day of treatment, arterial blood gas sao was %. on control chest ct obtained for crepitus in the bilateral supraclavicular region and pain on anterior chest wall revealed gas density surrounding both hilar and mediastinal structures with the extension of the right supraclavicular region. this view was regarded as pneumomediastinum ten days after hospitalization ( figure ). there was no reason as a bronchoscopic procedure or trauma that can cause images in history. piperacillin-tazobactam therapy was discontinued. tmp-smx continued intravenously. control ct taken month later, pnönomediastinum appearance resolved completely ( figure ). conclusion: most frequent causes of pneumomediastinum are barotrauma, asthma which led to increased intrathoracic pressure increase, cavitation forming pneumonia or invasive s procedures etc. spontaneous pneumothorax is often reported to be % during the pjp, however pneumomediastinum is often mentioned in the literature as rare cases. and these cases usually resolve within - weeks. if infection is considered, antibiotherapy must be started immediately and the protection of patients barotrauma are crucial. disclosure of interest: none declared. anti-proliferative effect of natural killer subsets on hematopoietic stem and progenitor cells j. riewaldt , ,* , s. sayed , , t. tonn , , experimental transfusion medicine, german red cross blood donor service north east, transfusion medicine, medical faculty, technical university dresden, center for regenerative therapies dresden, dresden, germany introduction: human hematopoietic stem and progenitor cells (hspcs) reside in the bone marrow niche within a complex cellular network including mesenchymal stromal cells and various immune cells such as osteoblasts, t lymphocytes and megakaryocytes (boulais & frenette, ) . in addition, the innate immune system -with natural killer (nk) cells as one of its major componentsis present in the bone marrow niche, although it is currently not clear whether and how it may contribute to hspc homeostasis. here, we will present data that show negative regulation of hematopoiesis by nk cell subsets in vitro. material (or patients) and methods: human mononuclear cells (mncs) were isolated from either peripheral blood (for nk cell isolation), g-csf-mobilized peripheral blood or bone marrow (for hspc isolation) by density gradient centrifugation after obtaining informed consent of the donors. the study was approved by the local institutional review board. cd + hspcs were enriched from mncs using a magnetic activated cell sorting (macs) approach, while different subsets of nk cells were obtained by either macs technology or fluorescenceactivated cell sorting (facs) . hspcs and nk cell subsets were co-cultured in vitro in semi-solid medium to assess colony forming unit capacities of hspcs. results: co-cultivation of hspcs with purified cd + nk cells resulted in drastically diminished colony formation after two weeks, as compared to cultivation of hspcs alone. although both myeloid and erythroid progenitor numbers were found to be reduced by the presence of nk cells, erythroid colonies appeared to be stronger negatively affected than myeloid colonies. co-culturing of different nk subpopulations facspurified based on surface antigen expression appeared to have differential impact on hspc differentiation/ colony formation capacity. conclusion: based on our data we conclude that the innate immune system contributes to control of hspc fate and behavior by suppressing hspcs differentiation and growth in vitro through specific nk cell subsets. further studies to dissect specific interaction mechanisms are required. hematology/oncology/palliative care, institute for biostatistics and informatics in medicine and ageing research, university of rostock, seracell stammzelltechnologie gmbh, rostock, germany introduction: the outcome of allogeneic hematopoietic stem cell transplantation (hsct) depends on a variety of patient-, transplant-, and donor-related factors. our retrospective study aimed at the evaluation of the potential risk factor cd + stem cell dose on early mortality after allogeneic hsct. material (or patients) and methods: data sets of consecutive patients who underwent allogeneic hsct at a single center (university of rostock) from to / were analysed regarding their early mortality rates (d+ , d+ introduction: mobilization and collection of cd -positive stem cells for subsequent high-dose chemotherapy constitutes a standard approach in myeloma patients achieving a good remission after induction chemotherapy. there are several regimens for mobilization of stem cells including the application of different dose levels of cyclophosphamide and subsequent stimulation with g-csf. so far, the dosage of cyclophosphamide varies between different protocols and affects both the risk of chemotherapy-associated complications and the probability of successful stem cell harvest. material (or patients) and methods: we retrospectively analyzed the impact of . g/m ( pts.) versus . g/m ( pts.) cyclophosphamide in consecutively evaluable myeloma patients (median age years, range - years) who underwent stem cell mobilization in first remission. successful mobilization chemotherapy was defined as a stem cell harvest of at least million cd + cells per kg bodyweight. in addition, important clinical factors such as infectious complications and toxicity were analyzed for both subgroups in this study. results: successful stem cell mobilization was achieved in out of ( %) and out of ( %) patients, respectively. median time to apheresis ( vs. days) and median cd + content of stem cell harvest ( . vs. . million cd + cells per kg bodyweight) did not differ significantly between both groups. importantly, there was a significant difference of anc nadir in favor of the cyclophosphamide regimen with . g/m ( . vs. . /nl, p = . ) while neutropenic fever was more often observed in patients who received . g/m cyclophosphamide ( vs. %, p = . ). furthermore, after induction chemotherapy using bortezomib, cyclophosphamide, dexamethasone successful stem cell mobilization was achieved in of ( %). one mobilization failure ( . million cd +/ kg) in this subgroup was observed in a patient who had previously received adjuvant anthracyclinecontaining chemotherapy for breast cancer. conclusion: cyclophosphamide at a dosage of . g/m is safe and highly effective for stem cell mobilization in patients with newly diagnosed multiple myeloma in first remission after induction chemotherapy. introduction: allogeneic stem cell transplantation remains an effective curative strategy in the management of haematological malignancies. since alemtuzumab was introduced as a method for immundepletion of grafts at groote schuur hospital and allogeneic stem cell transplants for bone marrow failure syndromes or haematological malignancies were performed. this is a retrospective audit reviewing the outcome of transplants performed from with a minimum follow up of months. material (or patients) and methods: patients with haematological malignancies underwent transplantation in remission of the malignancy. the conditioning was with myeloablative dosed with busulfan ( kg/kg), melphalan ( mg/m ) and cyclophosphamide ( mg/kg). since , fludarabine ( mg/m ) was introduced instead. tbi and cyclophosphamide was prescribed in patients. stem cells from hla identical siblings were mobilised with filgrastim ug daily for days and ug on day of the day cycle. apheresis were performed with cobe spectra and optia instruments. infection prophylaxis was with an oral quinolone, azole antibiotics and valocyclovir. gvhd prophylaxis was by immunodepletion ex vivo with alemtuzumab mg for each mononuclear cells of the graft. patients received post-transplant cyclosporine for days. content of free alemtuzumab in the cell suspension was tested in grafts by elisa. results: over the study period patients were entered. the diagnoses were acute leukaemia: (adverse cytogenetics in ), advanced nhl: , mm: , mds: . median age was (range - ) years and were female. the median cd + cell number was . ( . - . ) x /kg and alemtuzumab dose "in the bag" was mg ( - ). alemtuzumab was not detectable in the supernatant of the graft in any of the donor grafts. all patients engrafted. twelve patients with consistently elevated ( . log value) cmv viral copies in the blood were treated with gancyclovir and none developed systemic disease. acute gvhd ( grade ) was seen in patients and it contributed to the death in . twenty patients ( %) died and at median follow up of days % survive in remission of their malignancies. the cause of death was infection during the neutropenic period, mainly from gram negative organisms (n = ) or candidaemia (n = ) and disease recurrence in . conclusion: for this younger population myeloablative conditioning followed by immunodepletion with alemtuzumab was well tolerated and associated with excellent engraftment, effective control of gvhd, low cmv reactivation and low recurrence rate of the malignancy. disclosure of interest: none declared. clinical utility of the hematopoietic precursor cells (hpc) count compared to the cd+ cell count in peripheral blood for prediction of apheresis performance introduction: hematopoietic stem cell (hsc) transplantation has become a standard therapy of hematological malignancies such as multiple myeloma (mm), hodgkin lymphoma (hl), non-hodgkin's lymphoma (nhl) and acute myeloid leukemia (aml). hsc are induced to peripheral blood as a subsequence of the mobilization regimens with chemotherapy and g-csf or plerixafor alone. the treatment permits the collection of stem cells during continuous flow apheresis. peripheral blood stem cells (pbsc) are stored either in - °c or in liquid n until reinfusion. the aim of this study was to establish the viability of the cd + cells after cryopreservation with application of propidium iodide (pi), annexin-v and -aminoactinomycin-d reagent ( -aad). moreover we evaluated if age, disease, mobilization scheme or cryopreservation period influence the hematopoietic recovery after autologous peripheral blood stem cell transplantation (apbsct). material (or patients) and methods: we evaluated twenty six peripheral blood stem cell products collected from patients ( males, females). the median age was year ( - ). the analyzed group consists of mm patients, hl, nhl and aml. different mobilization regimens were applied: hdctx, hdvep, hdarac or intensive chemotherapy schedules (edap, rdhap, beacopp) with g-csf or sometimes with plerixafor. apheresis procedures were performed with continuous flow cell separator com.tec (fresenius, germany). hsc were harvested when the pre-apheresis circulating cd + cell count exceeds cells/μl. apheresis products contained median , x /kg cd + cells. the product was commonly divided into three eva bags (ethinyl vinyl acetate). the % dmso was applied as a cryoprotective agent. the programmed freezer was used before freezing in - °c. viability of pbsc was analyzed immediately after thawing for infusion. viability was established according to cytometric assay protocol for propidium iodide, annexin-v (abcam) and -aad (bd stem cell enumeration kit) with bd facscalibur flow cytometer using bdfacsdiva software. results: pbsc product contained median , x /kg cd + cells ( , . the median time of storage was weeks ( - ). with the application of flow cytometry and apoptotic dyes we determined different phenotypes: + / aad + , + / aad -, annexin + /pi + , annexin + /pi -, annexin -/pi -. the results were expressed as a percentage of viable, early apoptotic, apoptotic cells and necrotic cells [table] . statistical analysis shows no difference in apoptosis and necrosis in cd + cells harvested from patients with mm, hl, nhl, aml. also there was no correlation in cell viability and applied mobilization scheme. slightly more viable cd + cells were present in pbsc cryopreserved for less than weeks (annexin -picells; p = . ). conclusion: flow cytometry has become a method of choice of evaluation in variety of cell systems. we conclude that flow cytometric analysis of pi, annexin-v and -aad binding in pbsc concentrated is a convenient markers for quality evaluation of viability status of pbsc post thaw material that undergo transplantation within apbsct. this study shows that cryopreservation time (over weeks) has a negative impact for a number of viable cells. disclosure of interest: none declared. introduction: cord blood (cb) is an alternative source for sct in patients with haematological diseases. criteria for cb banking lead to a huge discard of units. the italian cbb network, following the original idea of p.rebulla and coll., is promoting a multicenter study to produce cb derivedplatelet gel (cb-pg) from units not eligible for unrelated transplantation (essentially due to low tnc count) for application in diabetic ulcers repair. in future this strategy will rescue unsuitable cb to a new clinical use and reduce the discard rate favorably impacting on cbbs activity. for this reason units with tncs o x and plt x /l underwent a two step-procedure to prepare plt concentrates (cb-pcs) further activated to pgs by recalcification. first, a soft spin ( gx min) is used to obtain platelet-rich plasma (prp), then prp is centrifuged at gx min to pellet plts and adjust pcs concentration to - x plts/l for conservation at - °c. as cb units may be contaminated by a variety of environmental contaminants at time of collection, sterility testing is crucial for the safety of cb-pg products. the simplest approach is to test the final product (cb-pc), volume range - ml, despite very small specimens are available for inoculating culture vials. we previously demonstrated that ppp may represent an alternative specimen, even if a percentage of false negative results was noted especially for fungi. material (or patients) and methods: here we evaluated the mixture of the two discarded fractions (ppp and prbc) as alternative specimen for seeding bd bactec plus culture media, namely anaerobic/f for anaerobes and peds /f for aerobes (recommended seeded volume - and - ml, respectively). bd media were chosen because intended for testing aerobic and anaerobic bacteria and fungi and commonly available in standard laboratories. for this purpose, we inoculated fresh cbs with low bioburden ( cfu/ml) of the following microbial isolates tested in triplicates: e. coli and s. agalactiae (aerobic bacteria), b. fragilis and p. acnes (anaerobic bacteria), c. albicans and a. niger (fungi). then cbs were processed according to cb-pg protocol. for anerobes , and . ml of either ppp +prbc or pcs were inoculated into anaerobic/f vials, while for aerobes and . ml of either ppp+prbc or pcs were inoculated into peds/f vials; incubation time: days. results: we evaluated the adequacy of sample, inoculum volume and bd media to detect the growth of aerobic/ anaerobic microrganisms in our setting. a minimal inoculum of . ml cps was able to detect the growth of both aerobic and anaerobic bacteria except for propionibacterium (slow grower) whose growth was detectable by inoculating ppp +prbc (limit volume: ml). for the other strains ppp+prbc inoculations with limit volume of for aerobes and . ml for anaerobes demonstrated to be a suitable alternative sample. conclusion: as cb-pc is a small volume product, the volume destined to quality controls is necessarily minimal. here we demonstrated that in case of ppp+prbc both sample and volume are adequate to detect major contaminants. a very tiny volume of pcs ( . ml) is also validated and is a critical step as a close system is still unavailable. besides molecular methods able to minimize the seeding volume, our approach could be easily reproduced in contexts were such resources are not available. matched sibling donor, one with relapse post autologous sct and due to graft failure post matched unrelated transplantation). in / the donor was a parent, had an haploidentical sibling. conditioning regimen was tbi based in two, combination with fludarabine in four, busulfan based in and treosulfan based in the remainder . all but one had atg. the origin of the graft was peripheral blood in with graft manipulation (cd +selection) and bone marrow in with high-dose cyclophosphamide post bmt. the median cd cells was x /kgr. results: engraftment was achieved in all except for one, who had reinfusion of additional cells. neutrophil and platelet engraftment occurred at a median of + (range - d) and + days(range - d) respectively. all but two had donor lymphocyte infusions (dli). out of had full donor chimerism in three months post bmt. acute grade ii-iii gvhd occurred in patients. skin gvhd grade ii-iii appeared in patients post dli. severe infection complications were present in patients with ebv related ptld in and systemic aspergillosis in one patient. with a median follow up of . months, patients are alive and well, patients have died due to disease relapse and one of trm. conclusion: the implementation of conditioning regimens that reduce the risk of rejection, as well as the refinement of histocompatibility testing have allowed the wider performing of haploidentical hsct. in our limited cohort of patients, the majority achieved prompt engraftment and full donor chimerism. severe grade ii-iii gvhd occurred mainly post dli and even though the incidence of moderate to severe infections was high, those were manageable in the majority of patients. main cause of death was disease relapse or progression, as expected in this high risk group of multiply pretreated patients. evaluation in larger cohorts, is necessary to further assess the long term benefits against the risks and to attain some insight into the immune mechanism that underlay the overcoming of hla barrier. disclosure of interest: none declared. clonal t cell large granular lymphocyte expansion following haploidentical haematopoietic stem cell transplantation in a patient with hodgkin lymphoma -a case report e. chapchap ,* , l. introduction: t cell large granular lymphocytes (t-lgl) expansions have been described following autologous and allogeneic haematopoietic stem cell transplantation with a low incidence, an indolent course and a distinct behavior when compared to de novo t-lgl leukaemia. there is some evidence that, even clonal expansion, may be triggered by an ethiopathogenic process involving persistent immunologic stimulation, as previously observed associations with cytomegalovirus (cmv) infection and graft versus host disease (gvhd). to our knowledge clonal t-lgl associated with haploidentical bone marrow transplant (haplo-bmt) has not been described yet. herein we present probably the first case of clonal t-lgl following haplo-bmt in a patient with hodgkin lymphoma (hl). material (or patients) and methods: to describe and highlight the clinical features observed on a patient with clonal t-lgl expansion following haplo-bmt to relapsed hl. an eight colour flow cytometry technique was used to immunophenotype peripheral blood lymphocytes and the pcr for t cell receptor gama-delta (tcr-γδ) gene rearrangement to detect clonality. results: a twenty five years old, male patient post autologous relapsed classical hl (nodular sclerosis) on a third partial remission following six cycles of brentuximab vedotin rescue has undergone an haplo-bmt and post-transplant cyclophosphamide (cy), non myeloablative (flu/cy/tbi cgy), isogroup (o+) at july/ . the gvhd prophylaxis consisted of tacrolimus (fk), mophetil mycophenolate and cy. after a successful treatment of cmv reactivation at d+ (ganciclovir) and systemic adenovirus at d+ (cidofovir plus human immunoglobulin), this patient developed since d+ until d + (last follow-up) marked and persistent lymphocytosis (range . - . x /l) associated with mild to moderate neutropenia (range . - . x /l) without any signs of gvhd or lymphoid organ involvement or autoimmunity. patient was still using fk and donor chimerism was %. the peripheral blood immunophenotype demonstrated an expansion of . x /l ( % of total lymphocytes) cd +/cd + t lymphocyte. bone marrow lymphocyte immunophenotype revealed a similar pattern, corresponding to % of total lymphocytes. the cd /cd ratio was . (peripheral and bone marrow) and natural killer cell percentage was . % (peripheral) and % (bone marrow) of total lymphocytes. aml (n = ), all (n = ), cmml (n = ), nhl (n = ), chl (n = ); patients had received a previous sct: autologous sct, urd. all patients received a mac, tbf (thiotepa, blusulfan, fludarabine), tbi, fludarabine. at hsct, patients were in cr ( in rst cr, in nd or nd cr) while presented active disease. unmanipulated bone marrow was the stem cell source for all patients. gvhd prophylaxis was performed in all patients with pt-cy (d+ ,+ in patients; d+ ,+ in patients), cyclosporin from d , mycophenolate from d+ . results: pmn leukocyte engraftment was reached at a median of days (range - ). the cumulative incidence of grade ii-iii acute gvhd was . % and moderate chronic gvhd %. cumulative trm at d+ was % and the relapse rate was . %. at d+ , patients ( %) had full-donor chimerism. at d+ , a good immune reconstitution was present in % of evaluated patients. the actuarial month dsf rate was % for patients in remission at hsct and % for patients with active disease at hsct. causes of death were: relapse (n = ), infections (n = ), gvhd (n = ), mof (n = ). conclusion: haplo-hsct with pt-cy offers a useful treatment tool in selected patients with high-risk haematologic malignancies. in this multicentric study, we confirmed that this procedure offers long-lasting remission, with limited toxicity, a low-grade agvhd incidence and early immune reconstitution. new therapeutic strategies are needed to further reduce the relapse rate. a careful selection of alloreactive donors, hsct performed in the early stage of disease, and an adequate adoptive immunotherapy are the future challenges. disclosure of interest: none declared. introduction: hematopoietic cell transplantation (hct) from an haploidentical related donor is an alternative option for patients (pts) without a suitable hla-matched related or unrelated donor, or for patients at a high-risk of disease progression. in argentina the median time from the beginning of the search of an unrelated donor to the hct is . months. the haplo hct platform provides an opportunity for nearly all patients to benefit from hct. material (or patients) and methods: from january to june , pts with high-risk hematologic malignancies underwent t-cell replete haploidentical donor transplantation using posttransplant cy (pt-cy). median follow-up: . months (ci % . - . ), myeloablative (mac)/reduce intensity conditioning regimen (ric) / pts, median age . ( - ), female/male: / . patients underwent mac hct were older than ric hct: . vs . years-old (p = . ). . % of the entire group received a previous transplant (mac %, ric %; p = . ), , % were high-risk disease (≥ cr, pr,pd,ref), mac . %, ric %; and , % standard risk (cr , cr ), mac %, ric % (p = . ). hct-ci: - / ≥ : . %/ . %. mac were % bu-flu-cy, % flu-tbi, % bu-flu, % flu-cy-tbi, % others) and ric regimens ( % flu-cy-tbi, %bu-flu, %bu-flu-cy, % flu-mel, % others. in mac group % were all, % aml, . % cml in bc. in ric group % were aml, . % all, . % hd, . % nhl, . % mds, . % multiple myeloma. graft source in mac group was bone marrow (bm) %, peripheral blood (pb) . %, bm+pb . %; in ric group: bm . % pb . %, (p = . ). gvhd prophylaxis: pt-cy-fk-mmf ma/ric . % / . % and . % used cs-a instead of fk in both groups. results: non relapse mortality (nrm) cumulative incidence (ci) at and moths: . % and . %. % of nrm were attributed to infections. relapse ci at and months: . % and . %. nrm according to hct-ci: - = . % vs ≥ = . % (p = . ). one-year overall survival (os) was . %; according hct-ci at year: - = . % vs ≥ = . % (p = . ). results were not significantly different between mac and ric (table) . in the multivariate analysis, only hct-ci was associated with survival ( - vs ≥ : hr . ; % ci . - . ; p = . ). final graft contained a median of . x (range , x - x ) cd + cells/kg, . x (range , x - , x ) cd + cells/kg, . x (range . x - x ) cd + tcells/ kg in cd /cd depleted grafts; . x (range . x - . x ) cd + cells/kg; . x (range . x - , x ) tcrαβ tcells/kg; . x (range x - . x ) tcr γδ tcells/kg in tcr αβ/cd depleted grafts. several leukocyte subpopulations were analyzed in peripheral blood at days + ± , + ± , + ± after transplantation using multiparametric flow cytometry. results: engraftment was registered in / patients, median days (range - ) for neutrophil recovery, median days (range - ) for platelet recovery and all patients had full donor chimerism at time of engraftment. secondary graft failure developed in patients (mds and hlh). gvhd was not registered in % of cases. in fact, acute gvhd was registered only in patients, grade skin in patients, and grade in remaining cases. kaplan-meier estimate of overall survival is % at median follow up of -years. immune reconstitution was fast: leukocyte subpopulations levels were normalized in about days after transplantation. mean values of . x for cd + (+ days), . x for cd + + (+ days) and . x for cd + (+ days) were early reached. conclusion: graft manipulation by t-cell depletion is a highly effective method to prevent gvhd, improve outcome, and optimize immune reconstitution. use of haploidentical transplantation can become a first line treatment for children who lack sibling donor, considering his safety and efficacy. disclosure of interest: none declared. introduction: cd is an ectoenzyme with dipeptidyl peptidase (dpp ) activity expressed on a variety of cell types and is known to contribute to t cell activation with well-defined modulatory effects on immune response. evidence is mounting that cd /dpp expression and enzymatic activity contribute to the emergence of autoimmune disease, immune-mediated abortions and transplant rejection. we had shown previously that, in autologous stem cell transplantation (sct), the amount of cd ++ t cells contained in the autograft correlated with poor clinical outcome. in this study, cd ++ lymphocytes were investigated in patients who underwent first allogeneic haploidentical sct. material (or patients) and methods: to characterize cd ++ lymphocytes for future correlations with clinical outcome, especially chronic graft versus host disease (cgvhd). methods: between january and october , eligible patients who underwent first allogeneic haploidentical sct and their corresponding haploidentical donor were included in a prospective analysis. cd ++ lymphocytes were characterized by flow cytometry. flow cytometric analyses were performed at defined timepoints prior to and during haploidentical transplantation. in addition, the serum concentrations and the enzymatic activity of scd /dpp were measured with commercially available enzyme-linked immunsorbent assay (elisa) kits. results: cd ++ lymphocytes were identified to have a unique phenotype (cd + , cd + , cd + , cd ro + , ccr + ). the numbers of these cells showed substantial temporal heterogeneity after transplantation, and clinical data are currently collected to be matched with flow cytometric results and laboratory findings. conclusion: cd ++ lymphocytes define a unique set of t memory cells that have been postulated previously to possess chemoresistance and t-cell repopulating capacity. given that adenosine deaminase (ada) is anchored to t cells via cd , the high density of cd is a critical prerequisite for chemoresistance, longevity and protection of a t cell population with obvious pathophysiological importance. long-term follow up data are expected to allow the assessment of the clinical importance of this distinct subset in t-cell reconstitution, immune competence and chronic gvhd. disclosure of interest: none declared. outcomes of high grade gastrointestinal gvhd post-hsct in children material (or patients) and methods: this is a retrospective analysis of pediatric patients presented with a clinical diagnosis of stage and acute gvhd of the gis who were selected from allogeneic hematopoetic stem cell transplantation (hsct) performed. the demographics, the regimen used for conditioning and gvhd prophylaxis, clinical characteristics of gvhd including follow-up, laboratory parameters during gvhd, treatment modalities used for gvhd, response assessment in every week, complications of gvhd, and survival data were recorded. results: patient and transplant characteristics were summarized in table . conclusion: discussion overall survival at months after the onset of stage or gut gvhd was %. better outcome than the adult data might be related with more usage of bone marrow as a stem cell source or different characteristics of pediatric gastrointestinal system. in general, gvhd started days post-hsct, extended to gis gvhd in a week and progressed to high grade gis gvhd in days. the initial day of gvhd, days to extending to gis gvhd and progressing to high grade did not influence the mortality. melphalan and atg usage in conditioning regimen and the drugs used in gvhd prophylaxis were not associated with survival. low albumin level at any time of the severe gis gvhd was associated with high mortality possibly due to more inflammatory gis state of lost patients. because atg usage in the treatment of gis gvhd significantly increased the mortality in our study, it should not be used in a routine manner. better but not significant outcome with non-immunosuppressants seems a better treatment approach. today best approach to gis gvhd is to prevent and recognize it early. treatment without immunosuppressive therapy like photopheresis and mesenchymal stem cells seems a better approach which deserves further research with more patients. disclosure of interest: none declared. transfusion associated graft versus host disease in a patient treated for myeloma multiplexa case report introduction: transfusion-associated graft-vs-host-disease (ta-gvhd) is rare but usually fatal complication of transfused cellular blood components. the incidence of ta-gvhd is uncertain. diagnosis of ta-gvhd may be difficult. this disease is characterized by fever, rasch, abdominal pain, diarrhoea, pancytopenia and liver disfunction - days after transfusion. mortality rate is very high. death typically occur about days after the transfusion. we describe a fatal case of ta-gvhd that occurred after transfusion of blood in a patient who has been treated for multiple myeloma. material (or patients) and methods: case report: in a year old men patient (pt), diagnosis of myeloma multiplex was established in . pt received six cycles of chemotherapy (vad) prior to autologus sct. the maintenance therapy was applied after transplantation. one year after the transplantation relapse occured and secondary sct had been done in pt (in decembar .). posttransplanted autologous hematopoietic reconstitution was done timely. two weeks after transfusion in patients occur fever, nausea,vomiting and diarrhea with normal biochemistry. infection were excluded. twenty-three days after transfusion on physical examination noted skin rash and patechie all over the body. his respiratory, cardiovascular and central nervous system examination was normal. laboratory data revealed pancytopenia and liver disfunction. histology of bone marow was normal, while the histology of skin confirmed ta-gvhd. results: after the evidence ta-gvhd was treated with pulse steroids, then the triple immunosuppressive therapy (cyclosporin a, mycophenolate mofetil, steroids). during treatment leads to the development of infection. clinical application of treatment has led to a loss of fever, diarrhea and skin rash with recovery of renal function, of liver function and of blood count parameters. six months after transfusion leads to deterioration of general condition and development of severe infectious syndromes (reactivation of cytomegalovirus, blood culture grew pseudomonas aeriginosa), pancytopenia, heart and liver failure. the therapy is turned off csa, continued implementation mycophenolate mofetil and again implementation steroids. eight months after transfusion leads to worsening ta-gvhd and death. conclusion: the diagnosis was based on the clinical condition of the patient, fever, nausea, vomiting, diarrhea, skin rash and petechiae all over the body, kidney and liver weakness, but the diagnosis of ta-gvhd was confirmed by biopsy of the skin. the patient was treated with triple immunosuppressive therapy (cyclosporine a, mycophenolate mofetil, steroids), with the development of severe infectious syndrome as a complication. lethal outcome for ta-gvhd, but patient survival was four times longer than the average. due to its rarity, ta-gvhd requires further investigation in order to define uniform approach and develop treatment and transfusion guidelines. disclosure of interest: none declared. introduction: acute graft versus host disease (agvhd) remains a significant cause of morbidity and mortality following allogeneic haematopoietic stem-cell transplantation (allo-sct). treatment options for agvhd are limited, frequently sub-optimal and often associated with considerable sideeffects. corticosteroids (ccs) remain first-line therapy for agvhd, however even in those who respond (sr-gvhd) ( o %), therapy related morbidity including fungal infections or viral reactivation is high. patients refractory to ccs (nr-gvhd) have limited subsequent treatment options and there is no clear consensus for their use. moreover, responses are often inadequate and associated with significant complications. while the novel treatment strategies for agvhd are costly, the indirect costs of agvhd and the complications of current treatment strategies are less well delineated. material (or patients) and methods: in the current economic climate with finite resources, we sought to objectively quantify the economic burden of agvhd on our institution. we retrospectively analysed data for consecutive transplants over a -year period with respect to initial transplant bed stay and subsequent gvhd related re-admissions. we stratified groups by the presence or absence of agvhd, as well as further analysing differences between sr and nr-gvhd. the costs of these differences were calculated. results: between january and december patients underwent allo-sct within our institution from sibling (n = , %) or unrelated (n = , %) donors. the incidence of grade ii-iv agvhd was % (n = ). patients had sr-gvhd ( %) and ( %) had nr-gvhd. in nr-gvhd patients the mean number of subsequent agvhd treatments was [range [ ] [ ] [ ] [ ] . development of agvhd was associated with a significantly longer transplant admission, mean stay days in patients without gvhd vs days in those with sr-gvhd (p = . ) and s days in those with nr-agvhd (p = . ). those with agvhd patients had significantly more re-admissions in the first year post transplant (mean . without gvhd vs . (p = . ) and (p = . ) in sr and nr-gvhd respectively) resulting in longer overall inpatient days ( vs (p = . ) and days (p = . )). no differences were seen between sr and nr-gvhd, which is likely to reflect the higher death rates seen with nr-gvhd as a competing risk for admission. utilising current hospital tariffs the additional hospital stay associated with agvhd resulted in a mean excess cost of € , and € , for each sr-gvhd and nr-gvhd patient respectively culminating in an average annual institution cost of € , from bed stay alone. moreover, those with nr-gvhd had significantly more episodes of cmv reactivation ( . episodes) compared to no gvhd ( . ) (p = . ) or sr-gvhd ( . ) (p = . ). this is likely to further increase the economic burden of nr-gvhd and further cost analysis including antiviral, antifungal and diagnostic costs is underway. conclusion: the additional cost burden of agvhd is significant for the transplant community. strategies for steroid reduction to achieve reduced hospital inpatient stays are crucial for patient morbidity and to achieve institutional financial stability. additionally these data highlight the need for improved strategies in nr-gvhd. disclosure of interest: none declared. the role of prolonged ecp therapy İn bronchiolitis oblİterans after allo-hsct b. erer ,* , f. gulen , s. yilmaz pediatric bmt center, yeditepe university, istanbul, pediatric pulmonology and allergy, ege university, izmir, turkey introduction: chronic gvhd (chvhd) continues to be a major complication after allo sct. the lung involvement and development of bronchiolithis obliterans (bo) are particularly serious and life threathining conditions. despite the advanced medical and cellular therapy the mortality and morbity are still high. extracorporeal photopheresis treatment revealed a significant improvement of skin cgvhd, and several studies indicated steroid sparing effect on cgvhd in general. howeover the effect of ecp on lung function in patients with bo after hsc and lung transplantation is controversial. selective downregulation of t-cell-mediated immunity is thought to explain the effect of extracorporeal photopheresis in the treatment of gvhd. material (or patients) and methods: we report a case of extensive cgvhd (skin+lung) in years old girl affected by b-all who received unmanipulated bone marrow transplant in the nd remission of disesase from her hla = abo# brother. full donor engrafment was documented on day + . she developed grade ii-iii acute skin cgvhd responded to steroid therapy (metihylprednisolon mg/kg). she was discharged on day + . subsequently she has had several admittions to the hospital due to severe hemorrhagic cystitis. months after transplant she developed extensive sclerodermiform skin cgvhd. the patient refused the steroid therapy, thus the puva therapy was given for months. there was a significant response, except the permanence of lichenoid lesions localized on both ankles. the immunosupressive therapy (is) has been gradualy reduced and stopped. six months later she presented with dry cough and dispnea; wheezing and inspiratory squeaks were present on thoracic examination in absence of infection. based on hrct of the chest and pft the diagnosis of bo was made. the next months the symptoms, rft and radiologic findings (hrct) impaired despite the aggressive immunosuppressive therapy. long course of ecp program has been added to the is therapy. results: she has performed courses of ecp therapy. at the end of the second course of ecp therapy, cessation of disease progression was observed. she has completed months of ecp therapy. clinical symptoms have disappeared, pft and hrct of chest have drammaticaly improved. the effect of ecp in the treatment of cgvhd may be correlated to the duration of therapy. a prolonged ecp therapy should be considered as a therapeutic option in the treatment of bo after allojeneic hsct. disclosure of interest: none declared. comparison of two apheresis devices in mononuclear cell collection for extracorporeal photopheresis in refractory graft versus host disease e. gonzález , c. pascual ,* , l. solan , s. redondo , a. pérez-corral , m. bastos , , d. serrano , , j. gayoso , , m. kwon , , p. balsalobre , , j. anguita , , j. l. díez-martín , hematologia, hospital gregorio marañon, instituto de investigación sanitaria gregorio marañón, madrid, spain introduction: extracorporeal photopheresis (ecp) is an effective cell therapy in graft-versus-host disease (gvhd) after bone marrow transplantation, which requires an optimal mononuclear cells (mnc) collection through apheresis devices for further manipulation.the end of this study was to compare the semiautomatic cobe spectra (terumo bct ® ) and the new automated spectra optia (terumo bct ® ) in terms of efficacy and safety in ecp. material (or patients) and methods: all patients with refractory acute and chronic gvhd in treatment with ecp were retrospective studied, from june to may . patients were alternatively assigned with either apheresis device. peripheral blood sample was removed to the patients before and after each leukocyte apheresis (la) procedure and also had a sample of each product just before reinfused. we perform the cell blood count (cbc) in unicell dhx (beckman coulter ® ) in all samples. collection efficiency (ce) of white blood cell (wbc) and total mononuclear cells (tmnc) and platelet loss(pl)were calculated as: wbc ce% = [product wbc-ncx /ml x product volume ml/ preapheresis nc (x /ml)x(vol processed ml-ac volume ml)] x tmnc ce% = [product mncx /ml x product volume ml/ preapheresis mnc (x /ml)x (vol processed ml-acvolume ml)]x pl = [preapheresis count-postapheresis count/ preapheresis count]x *in all cases it was complete with saline solution to a total volume of ml. it were compared patient's cbc before and after the la, the cbc of the product and the procedure variables with both devices, using non-parametric methods (u mann-whitney). results: a total of procedures ( performed in cobe and in optia) were included, carried out in patients ( with chronic gvhd and with acute gvhd). central venous access was required in / ( . %) patients. cbc results before and after la were similar. in optia group, the processed volume, the volume of the product, the percentage of loss of platelets, the amount of acd used and the amount of acd in the product, were significantly lower. the hemoglobin and the hematocrit were higher in the optia group, and tmnc count was similar with both devices. notably, tmnc ce was greatest in the optia group.there were no differences in the duration of the procedure. details are shown in table . the procedures were well tolerated with both devices, and only mild episodes of hypocalcemia were recorded in four procedures (two in optia group, and two in cope group). conclusion: in our study, the spectra optia proved very effective and save in collecting mnc for ecp in gvhd.because of this, we could consider shortening the time of collection with this device. the lower pl is very important because these patients often have low platelet counts. it also provides the advantage of being automatic and lower use of acd what may reduce the hypocalcemia risk. the hemoglobin and the hematocrit were higher in the optia group, but in the established limits for proper irradiation. introduction: transplant-associated thrombotic microangiopathy (ta-tma) represents a challenging and potential lifethreatening complication after allohsct. this syndrome occurs approximately in % to % of patients after allohscts, and it is characterized by inflammation, thrombotic microangiopathy, hemolytic anemia, thrombocytopenia, and evidence of organ damage, particularly renal and neurological involvement. the mortality rate is approximately - %. the treatment is problematic and no generally effective therapies are available. therapeutic options consist in a wide range of treatment such as plasma exchange (pex) or immunosuppressive drugs, however lacking in high response rate due to the unclear ta-tma pathophysiology. recent data suggest that endothelial injury and complement activation could be involved in the pathogenesis of ta-tma. eculizumab, a monoclonal antibody against terminal complement, may be an innovative and effective therapeutic option for ta-tma. material (or patients) and methods: we report a case of ta-tma in a recipient of allohsct for acute lymphoblastic leukemia b ph negative. -year old male who underwent allohsct from unrelated donor / hla compatible with total nucleated cells . x /kg and cd + . x /kg. conditioning regimen consisted of tbi cgy and cyclophosphamide mg/kg on days - , - . anti thymocyte globulin, methotrexate and ciclosporine (csa) were given for graft versus host disease (gvhd) prophylaxis. results: at day + of reinfusion the patient experienced cutaneous acute gvhd (skin , global ) treated with prednisolone mg/kg. at day + , due to worsening of cutaneous rash and liver dysfunction (elevated transaminase and bilirubin levels), the patient started treatment with etanercept mg/sc for a total of infusions. meanwhile the ldh level was increased up to u/l and was associated to anemia, trombocytopenia and renal deterioration; schistocytes were detected at the peripheral blood smear ( hpf) and urinanalysis revealed a protein concentration of . mg/ h. csa was replaced with mycophenolate mofetil and the patient started treatment with plasma and pex daily, stopped after cycles due to trombocytopenia. few days later patient developed peripheral paresthesia and hypostenia; cranial mri was performed, with no pathological features. despite recovering renal status and hb level, c and c levels were inferior to normal range and patient started treatment with eculizumab mgs for a total of infusions. after the first dose of eculizumab there was a marked improvement of performance status, ldh, hb level and platelet count (fig. ) ; schistocytes in the peripheral blood decreased until disappearance. at + diarrhea gr appeared and a colonoscopy showed ulcerative mucositis and inflammatory infiltrates. patient received infliximab mg/mq ( doses). actually patient is at + and experience only mild cgvhd (nih scale: mouth , eyes ). conclusion: this report support the potential effective activity of eculizumab in the treatment of life-threatening ta-tma even when it is associated with refractory acute gvhd. introduction: we have been using at our institution phototherapy -puva and uvbnb as skin direct therapy associated to ecp to treat different manifestations of cutaneous gvhd for the last years. ecp is indicated to patients not responsive to steroids, those with is toxicity or those who need early withdrawal of is. material (or patients) and methods: all patients treated with phototherapy since were prospectively evaluated in a single center institution. puva and uvbnb was performed with standard machine and uva initial dose was based on skin type. ecp was performed with uvar ® photophresis system and liquid psoralen. puva and uvb-nb could also be combined with ecp for different clinical cutaneous manifestations or as ecp maintenance therapy. complete response (cr) was defined as more than % of clinical response and partial response (pr) between % and %. patients who had less than phototherapy sessions were considered early for evaluation (ea). results: puvatherapy was used in patients to treat lichen planus like and sclerodermic chronic gvhd with no visceral involvement. uvbnb was used in patients for vitiligo, limited lichen planus like chronic gvhd, follicular keratosis, patients with psoralen oral intolerance and children. ecp was reserved to treat patients with extensive sclerodermic, fasciitis cgvhd, systemic cgvhd and patients with acute gvhd corticosteroid-refractory. patients treated with ecp had more severe cgvhd (p o , ). ecp was combined with puva in six patients and in uvbnb in . conclusion: puva and uvbnb are topical direct therapies and should be considered in patients in whom long-term systemic is is antecipated. phototherapy as topical therapy on the skin associated with ecp helps in the treatment of sclerotic cgvhd extensive and vitiligo that are often seen in the same patient. this study shows encouraging results and suggests that phototherapy treatment results in more rapid reduction of immunosuppression. introduction: chronic graft versus host disease (cgvhd) is major predictor of long term survival after allogeneic stem cell transplantation (hsct). similarities with autoimmune diseases are due to b lymphocyte (b-ly) dysregulation and thus abnormal production of various auto antibodies (ab). using of chimeric monoclonal anti cd antibody (rituximab) is based on their "in vivo depletion" of b-ly and thus we estimate efficacy and safety of rituximab in the therapy of refractory severe cgvhd. material (or patients) and methods: seven patients, m/f / , have received rituximab after failure to standard treatment (methylprednisolone, cyclosporine a, mycophenolate mofetil, azathioprim, puva…). median age was years ( - ). all of them have received hla identical sibling transplant with stem cells from peripheral blood in and bone marrow in cases. despite standard prophylaxis, of them had acute gvhd s grade - and all of them had severe cgvhd with lichenoid, sclerodermic or pemphigoid skin feature, buccal mucosa involvement with sicca syndrome. extracutaneous manifestation included lung in , liver in and thrombocytopenia in cases. one patient had cmv infection prior to exacerbation of cgvhd. we applied rituximab in usually recommended regimen ( mg/m weekly for weeks). results: exact responses were measured with schirmer test, spirometry, liver function parameters and hematological findings. good effect was observed in patients ( , %) even after first application. best results were achieved concerning mucosa (significant reducing of sicca syndrome) and skin (improvement of turgor, lichenoid changes and pemphigoid reconstitution). lung and liver function were improved as well, but it should be mentioned that other immunosuppressant were also given (particularly mycophenolate mofetil and azathioprim). three patients had severe infectious complication, two of them have died from sepsis, and one of them have developed abscesses pulmonalis in fourth week of treatment. conclusion: results from our modest data suggests that monoclonal anti cd antibody in the treatment of severe refractory cgvhd is encouraging but needs further investigation. disclosure of interest: none declared. introduction: human herpesviruses (hhvs) are commonly found in general population but also in patients after allogeneic hematopoietic stem cell transplantation (hsct) where they may cause severe complications. particularly, cytomegalovirus (cmv) infection was significantly more frequent among adults and was associated with a higher risk of developing acute graft-versus-host disease (agvhd) ( ) ( ) ( ) . therefore, the aim of our study was to determine prevalence of particular hhvs in patients with oral chronic graft-versus-host disease (cgvhd) and possible role in these patients. material (or patients) and methods: this pilot study enrolled cgvhd patients who are part of a larger multidisciplinary cgvhd project at the university hospital center zagreb, croatia. patients were evaluated for clinical and cgvhd characteristics; severity of cgvhd and oral involvement were determined by using established nih consensus criteria. cytobrush swabs from oral mucosa were collected from all patients. specimens were analyzed by specific polymerase chain reaction (pcr) of several hhvs (cmv, epstein-barr virus -ebv, herpes simplex virus and , and varicella zoster virus -vzv). data were evaluated descriptively and chi-square test was used to calculate statistical significance. results: cgvhd patient were enrolled, of which men (average age . ) and women (average age . ).twenty patients had diagnostic signs of oral cgvhd ( . %), were ( . %) without, and ( . %) had distinctive signs of oral cgvhd. overall, in of patients ( %) no hhvs were identified, / ( . %) had cmv, / ( . %) had ebv, / ( . %) vzv and no hsv and were found, respectively. ebv was statistically significantly found more often than the second most common cmv (p = . ). there was no statistically significant difference in hhvs between patients with and those without oral cgvhd. conclusion: although proportion of patients with cgvhd, especially patients with oral cgvhd was ebv positive, indicating the possibility that ebv is more common in chronic gvhd, further studies with control groups are necessary to determine the role of ebv in cgvhd. our preliminary data also indicate that the use of molecular methods to determine the presence of hhvs is valuable method in the follow-up of cgvhd patient particularly by using oral cytobrush specimens for pcr analysis as less invasive alternative method for monitoring these patients. introduction: bleeding complications and graft-versus-host disease (gvhd) are severe complications in hematooncological patients and patients undergoing allogeneic transplantation (hsct). a systematic and prospective assessment of these complications is essential in order to evaluate therapeutic interventions, optimize patient care and thus improve patient outcome. different bleeding scores to be used in clinical practice but also for research purposes have been developed, including the who and bsms scores. , recently, the national institute of health (nih) updated their recommendations for diagnosis and staging of gvhd. for the prospective and systematic evaluation of bleeding complications and gvhd, we developed an electronic tool with the aim to improve patient care and for clinical research purposes. material (or patients) and methods: in collaboration with it-specialists we developed a browser based electronic tool for assessment of bleeding and gvhd according to the published guidelines. this tool (bleedhd) works on different devices and thus allows efficient bedside assessment. collected data include patient's and disease characteristics, type of intervention, type and site of bleeding as well as gvhd features, according to the published guidelines. results: bleedhd is a secured browser based, user-friendly tool and was improved over time and validated on a large number of patients. the current available tool allows easy collection of patient characteristics including names, date of birth, gender, age, diagnosis and interventions (type of transplantation and stem cell source). thereafter, the assessor can choose between assessment of bleeding (either who or bsms or both) or gvhd or both. according to the published nih guidelines, gvhd can be scored in case of first diagnosis, by every single organ (organ scoring), by activity assessment and finally by self-reporting by the patient himself. the user will be guided step-by-step through the different scoring systems. assessments can be interrupted at any time and resumed later. bleedhd automatically gives an overview over time and scores s of each patient. during assessment, explanations, additional information and pictures can be retrieved allowing correct assessment also for not experienced users. all the data are safely stored on an appropriate server. complete data can be exported as cvs file for further analysis and research purposes. in our experience bleedhd significantly improves the accuracy of bleeding and gvhd assessment in an easy, fast and straightforward approach. scoring patients became much more convenient and easy. additionally it allows a prospective bedside assessment and clear demonstration of clinical changes in case of therapeutic interventions. physicians and nurses from other institutions, which are involved in patient care can have access to bleedhd and also assess gvhd during follow-up. conclusion: bleedhd is an innovative and easy tool for an accurate, efficient and prospective assessment of bleeding complications and gvhd in hemato-oncological patients. this can improve patient care and outcome. additionally, it can be a useful tool for clinical research in the therapy of gvhd and to further investigate the efficacy of platelet transfusions or other interventions in thrombocytopenic patients. introduction: graft versus host disease (gvhd) is a lifethreatening complication of allogenic hematopoietic stem cell transplantation, which has a wide range of pleomorphic manifestations at multiple organs. extracorporeal photopheresis (ecp) is an immunomodulatory therapy that provides encouraging results in skin gvhd but also in multiorganic involvement (skin, gut and liver). the ecp reverses gvhd through the reduction in response of donor effector t cells and production of donor tregs. material (or patients) and methods: we retrospectively analyzed patients with refractory acute and chronic gvhd treated in our center with ecp from september to october . the median age was (range - ), the main primary disease was acute myeloid leukemia. there were patients with agvhd that were classified according to the degree of severity in grade ii and iii % and grade iv %, patients with cghvd patients were classified according the global scoring of nih consensus into moderate % and severe %. skin was the organ mostly involved ( %), while % of patients had multiorgan involvement. ecp was performed using the online system therakos, and the off line system using the spectra optia separator, and the uva device macopharma. patients with agvhd received - consecutive days of ecp at weekly intervals until response and then tapered to every to weeks, patients with cgvhd received consecutive sessions at week intervals. results: the overall response rate (orr) for agvhd and cgvhd was % and % respectively. the orr for patients with agvhd grade ii-iii was % and grade iv was %. in moderate and severe cgvhd the orr was and % respectively. we analyzed organ specific complete response (cr) rates in patients with agvhd and cgvhd (graphic ), in skin involvement cr was achieved in % and % (agvhd and cgvhd), in multiorganic involvement (skin, liver and gut) the cr was % and % respectively. seven patients died, the main cause of death was infectious disease. the procedure was well tolerated, only nci grade i adverse events were observed. conclusion: in our experience the use of ecp for agvhd, had favorable rates of response especially at early stages. there is tendency for higher responses in agvhd with cutaneous involvement. we observed significant rates of responses in moderate and severe cgvhd. we found that ecp has a considerable activity against multiorganic cgvhd. the early intervention with ecp improves the poor outcome of patients with steroid refractory gvhd. introduction: after autologous hematopoietic stem cell transplantation (hsct) it exists a period of time with an increased risk of infectious diseases because of immune dysfunction. for this reason, it is advisable to administer several vaccines beginning - months after transplantation. in our hospital, the haematologist responsible for posttransplant monitoring establishes the prescription of vaccination, both the dates and the number of doses, including vaccines against stp. pneumoniae, h. inluenzae type b, c. tetani, c. diphtheria, poliovirus and influenzavirus. after this, patients are sent to the primary care trust (pct) where the general practitioner is responsible for the correct completion of the set prescription, indicating the administration to the nursing unit. material (or patients) and methods: the aims of the current study are: a) to assess treatment adherence to the vaccination schedule elaborated in our unit and determine reasons for lacking of adherence, if this is the case; b) to identify administrative problems (lack of vaccines, absence of collaboration between the general practitioner and the haematologist responsible, etc.); c) to describe adverse reactions related to vaccines (fever, rash, etc.). methods: descriptive and retrospective study of patients who underwent autologous stem cell transplantation in our hospital from may to october , so it has been possible to complete the vacunal schedule. information was obtained by telephone interviews and reviewing medical histories. patients were questioned about: a) the administration of every prescribed vaccine; b) reasons why vaccines were not administered, if this occurred; c) administrative problems in obtaining them; d)administration of additional vaccines not prescribed by the haematologist; d) local and systemic adverse reactions after vaccine administration. results: transplanted patients ( male, female) with a median age of . years, diagnosed of multiple myeloma ( . %), hodgkin lymphoma ( . %), nhl-b ( %), nhl-t ( . %), acute leukaemia ( . %), plasmatic cell leukemia ( . %). of these, ( %) patients answered the questionnaire and ( %) were unable to be contacted due to exitus or change in hospital. of those who completed the questionnaire, ( . %) recieved all the prescribed vaccines by the haematologist, patients reported having problems with availability. from ( . %) patients who have not been administered any vaccines, of them were for haematologist indication (due to progression of disease), and obey to patient refusal. from all vaccinated patients, only ( . %) were given an additional vaccine, not included in the original calendar (influenza a). from those vaccinated, ( . %) reported any of the following adverse reactions: fever ( ), asthenia ( ), flu-like symptoms ( ), and local skin reaction ( ) . conclusion: in our hospital unit, even though there were a relatively small number of patients, the degree of adherence observed was quite high and without any relevant incidence. we consider that collaboration with the primary care trust in the administration of the post-transplant vaccination schedule has an important role when treating and caring for our patients. disclosure of interest: none declared. introduction: it has been reported that reactivation of hepatitis b virus (hbv) could be fatal in patients with malignancy following systemic immunosuppressive chemotherapy, especially combined with rituximab, or hematopoietic cell transplantation (hct). many guidelines regarding hbv reactivation following chemotherapy recommends that hb surface antigen (hbsag), antibody to hb core antibody (anti-hbc), and antibody to hb surface antibody (anti-hbs) should be tested before the chemotherapy is initiated. japanese guideline for hbv infection in whom received chemotherapy was published in ; however it remains a few cases reported who developed fulminant hepatitis from hbv reactivation following chemotherapy. this fact implies that the guideline is not properly observed. therefore we conducted this study to evaluate whether hbv screening tests before hct were employed according to the guideline in the hbv-endemic region of japan. material (or patients) and methods: we retrospectively analyzed patients who were treated with allo-or auto-hct between and at kansai medical university hospital. results: a total of patients were studied. patients underwent allo-hct and patients underwent auto-hct. all patients were examined on hbsag before the hct. patients ( . %) were proved to be hbsag positive carrier, who received nucleoside analog against hbv thereafter. only patients ( . %) were investigated neither on hbcab or hbsab. although patients ( . %) were seropositive either for hbcab or hbsab, patients ( %) among them were not examined on hbv-dna as follow-up. fortunately no fulminant hepatitis was experienced, patients developed late-onset reactivation. these were patients who underwent auto-hct. one was a hbv carrier who had received prophylactic nucleoside analog during the whole course of therapy, whose hbv-dna level resulted in undetected. she developed reactivation of hbv with long interval after auto-hct. the other had occult infection of hbv, who converted to late-onset reactivation after auto-hct. neither of them developed hepatitis after they were retreated with nucleoside analog against hbv. conclusion: improvement was achieved by the guideline on management against hbv reactivation following chemotherapy. while it did not pervade perfectly even in the hbvendemic region of japan. most hematologists would recognize how important to examine the hbv serostatus before the hct; however they are afraid to lose the following blood test regarding hbv. the guideline might be complicated to observe the follow-up study from the point of view. thus, the guideline is preferable to be simplified and the institutional surveillance should be established to encourage the follow-up study. if other medical staff composed of nurses, pharmacologists, and laboratory technicians as well as hematologists, is aware of hbv serostatus of the patient during the entire course of chemotherapy, it could be attributable to prevent from missing the follow-up study. introduction: citomegalovirus meningoencephalitis is a rare and fatal complication after allogeneic hematopoietic transplantation. we describe a paediatric patient who developed the infection one year after cord blood transplantation although preemptive treatment and good control of blood viremia. material (or patients) and methods: a -year-old male patient with ph+ acute lymphoblastic leukemia in third complete remission received an unrelated / hla matched single umbilical cord blood transplant. the patient and the donor (maternal igg serostatus) were cmv-seropositive. the conditioning regimen was based on fludarabine, thiotepa, busulfan and anti-thymocyte globulin. cyclosporine and a short course ( days) of prednisone were given as gvhd prophylaxis. because of acute grade ii gvhd (gut) patient received two courses of prednisone mg/kg/day and oral budesonide starting on day + and + , respectively. cyclosporine discontinuation started on day + . intravenous ig was administered monthly. the patient showed good engraftment and total donor chimerism although with a poor immune reconstitution (persistent low cd count) implying frequent cmv asymptomatic viremia. complete molecular remission was maintained. results: on day + the patient started with apathy, memory lost, language deficit and lethargy. brain magnetic resonance imaging showed signs compatible with meningitis. cytomegalovirus was detected by quantitative pcr technique in the cerebrospinal fluid (csf) ( . ui/ml) whereas it was negative in blood. an autoimmune cause was ruled out. at that moment the patient was receiving oral valganciclovir as preemptive treatment for a previous cmv viremia (maximum . ui/ml). he was admitted to the paediatric bmt unit to start double intravenous antiviral treatment based on ganciclovir and foscarnet. cyclosporine was discontinued in one week. a progressive neurological worsening was seen during the next two months although negativity of cmv in csf was achieved. the patient died on day + . conclusion: citomegalovirus meningoencephalitis is a severe complication that can occur late after hematopoietic transplantation. our patient experienced the fatal infection although effectiveness of preemptive antiviral treatment to clear blood viremia. disclosure of interest: none declared. a year-old male patient with saa was admitted with fever, periorbital swelling, periorbital pain, and bloody nasal discharge. the patient had been diagnosed with saa. computed tomography of the paranasal sinuses demonstrated a space-occupying lesion in the right maxillary and ethmoid sinus region. histopathologic examination revealed numerous pas-stained aspergillus hyphae without tissue invasion. tissue culture revealed aspergillus fumigatus. voriconazole administration was started. case . a -year-old male patient was admitted to our clinic due to pancytopenia. the patient was diagnosed with saa and allogeneic hsct was planned. however he had fever on day of admission. invasive pulmonary aspergillosis was suggested based on the presence of pneumonic consolidations on ct of the thorax and galactomannan positivity ( . index). liposomal amphotericin b was started in therapeutical dose. results: invasive fungal infections (ifi), particularly those caused by aspergillus species, have long been recognised as a major cause of death in saa. mortality rate is still high in saa patients with ifi despite antifungal therapy and even granulocyte infusion plus g-csf administration. a condition in which the response to immunosupressive therapy can take up to months to develop leading to a neutropenic period that is typically longer than that seen after hsct. hsct with active ifi places the patient at a higher risk for transplant related mortality. however, hsct without delay is only curative treatment approach in some patients. we performed hsct under antifungal therapy in both patients. both patients overcame the neutropenic period and they were discharged without severe morbidity. conclusion: we conclude that an immediate hsct may be life-saving and g-csf plus granulocyte transfusion may be used as a bridge to hsct in patients who do not satisfactorily respond to antifungal therapy although there are scarce data about the hsct in saa patients with active invasive fungal infection. r- results: this presentation focuses on the recently updated oral care guidance which is based on the most recent evidence, clinician feedback and expert opinion.the guidance continues to focus on the key principles of; an accurate assessment of the oral cavity, identification of risk factors, regular care, earlier intervention to prevent/reduce oral damage and the correct treatment interventions. conclusion: it is anticipated that this updated guidance will further assist health care professionals in planning and implementing oral care into everyday practice, thus reducing a significant health burden for the patient and reduce demands on limited health care resources. disclosure of interest: none declared. successful pregnancy and delivery after myeloablative cord blood allogeneic transplantation c. de miguel ,* , g. bautista , c. regidor , i. krsnik , j. r. cabrera hematology, hospital universitario puerta de hierro majadahonda, majadahonda, spain introduction: infertility is a frequent long-term side effect among young hematopoietic stem cell transplantation (sct) receptors . concern about fertility negatively affects the quality of life of the patients which frequently ask about the optimal time to address these issues and the ways to do it. a few cases of successful pregnancies after autologous or allogeneic transplants, have been recently reported but, to our knowledge, none after cord blood transplantation (cbt). we report the case of a young woman who successfully carried out a pregnancy with a donated oocyte, years after a tbi-including myeloablative cbt. material (or patients) and methods: our patient, a -year old woman, was diagnosed of pro-b acute lymphoblastic leukaemia in august . she was treated with a national high-risk all protocol (pethema hr-all ), with persistence of disease after induction. first complete remission was achieved after the fourth consolidation. an allogeneic cbt was performed in march . the cb unit was co-infused with haploidentical selected cd + sc from an unrelated donor. conditioning was myeloablative and included fractionated tbi (total dose cgy), fludarabine mg/m ( mg/m , days - to - ), cyclophosphamide mg/kg ( mg/kg, days - and - ) and thymoglobulin mg/kg ( mg/kg, days - and - ). after cbt, the patient has remained in complete remission with a normal performance status. the patient consulted because she had been trying unsuccessfully to conceive. she was amenorrheic. a complete hormonal evaluation was performed to confirm that thyroid function was preserved and prolactin levels were normal. after evaluation, a procedure of in vitro fertilization (ivf) of a donated oocyte using the patient's husband sperm was proposed. two embryos were transferred to the uterus of our patient. results: a single pregnancy ensued which was carried out till term with no complications. the baby, a healthy g girl, was born at week + after a cesarean section. ten months later, both mother and child remain in good health. conclusion: reproductive issues in young women who undergo myeloablative allogeneic sct are especially difficult to address and solve. physicians must improve their practices, provide information and educational materials and facilitate the physician-patient discussion on fertility preservation. guidelines with recommendations are available for health-care providers . these recommendations are based on best available evidence. successful pregnancies in this population have to be reported. to our knowledge, this is the first report showing a successful pregnancy and delivery after myeloablative cord blood allogeneic transplantation hc was also more frequent in males than females (p = . ). the median survival was . years ( % ci: . - . years) in the group of patients who presented hc and . years ( % ci: . - . years) in patients who did not (p , ). conclusion: the incidence of hc in our series of patients was similar to that found in other studies. in our cohort of patients being male and having acute gvhd was a risk factor to develop hc. disclosure of interest: none declared. frequency of human neutrophil antigens (hna) in recipients of allogeneic hematopoietic stem cell allogeneic transplantation (hsct) and correlation with procedure outcome f. pereira , j. introduction: the hematopoietic reconstruction with allogeneic bone marrow is an established method of treatment for a variety of haematological, oncologic and immunologic diseases. hsct is associated with considerable morbimortality due to factors like recurrence of underlying disease, hla compatibility between donor and recipient and type conditioning regime. this study investigated the association between human neutrophil alloantigens (hna) and engraftment, the occurrence of acute gvhd and transplant related mortality in patients who underwent allogeneic hematopoietic stem cell transplantation at our center. introduction: cardiac failure is a known complication after hematopoietic stem cell transplantation (hsct), but predominantly occurs in predisposed patients with low output fractions or valvular heart disease. hereby, we present a case of acute takotsubo cardiomyopathy in a patient without preexisting cardiac comorbidities resulting in cardiogenic shock and ventilator therapy on day + after allogeneic hsct. to our knowledge this is the first report of takotsubo cardiomyopathy after hsct. the year old female was diagnosed de novo aml m . after induction therapy (cytarabine, mitoxantrone), % blasts persisted in bone marrow on day . re-induction therapy with mitoxantrone, cytarabine and fludarabine lead to a clearance of blasts. bridging therapy to hsct with cycles of decitabine was performed with no signs of aml recurrence. apart from mild hypertension treated with beta blockers and at antagonists, the patient had no history of cardiopulmonary disease. after reduced-toxicity conditioning therapy with treosulfan, fludarabine and atg, the patient received , x e cd positive cells per kg bw from an hla / matched unrelated male donor without any acute distress during re-transfusion of stem cells. however, eleven hours after transplant, she complained of acute chest pain, hypotension and dyspnea with decreased oxygen saturation that was refractory to non-invasive ventilation and was immediately transferred to the icu in cardiogenic shock. results: the ecg showed a heart rate of /min and an r-peak reduction from v to v without st segment changes. high sensitive troponine, brain natriuretic peptide and creatinine kinase were elevated, but there were no signs of hemolysis. coronary angiography was performed immediately after intubation. on examination there were no signs of coronary artery disease, but ventriculography and subsequent echocardiography revealed severe hypokinesia of the apical and medial walls (output fraction %) consistent with takotsubo cardiomyopathy. the patient received intermittent vasopressor therapy and continuous dialysis with volume restriction. she was stabilized and extubated after days. echocardiographic follow up three weeks after transplant showed completely normalized ejection fraction. the patient was discharged with normal output fraction and is well and alive on day after hsct. conclusion: takotsubo cardiomyopathy is an infrequent cause of acute coronary syndrome and can be induced by physical and emotional triggers. in describing this case, we wish to remind physicans to take into account takotsubo cardiomyopathy as a rare differential diagnosis in hsct recipients with acute cardiac failure. disclosure of interest: none declared. efficacy of triple drug combination in preventing nausea and vomiting s after high-dose chemotherapy p. introduction: patients frequently cite nausea and vomiting as one of the most distressing and debilitating side effects of chemotherapy. chemotherapy-induced nausea and vomiting (cinv) can be divided into acute ( or fewer hours after chemotherapy), delayed (more than hours after chemotherapy). this distinction is made because acute cinv is believed to be mediated through serotonin receptor stimulation while delayed cinv is thought to involve multiple neurotransmitters, including opioid and neurokinin receptors. chemotherapeutic agents have variable emetogenic potential that is affected by dose and method of administration. high-dose chemotherapy (hdc), often combined with tbi or total nodal irradiation of a varied amount, and stem cell rescue is a treatment modality applied to a wide variety of medical conditions. the delivery of high-dose therapy is almost always associated with a great degree of nausea and vomiting. material (or patients) and methods: a clinical study of triple drugs combination (aprepitant+palonosetron+dexamethasone) was carried out to evaluate its efficacy in preventing both acute and delayed emesis after high-dose chemotherapy ( historical control groups of patients were treated with dexamethasone (dex) and palonosetron. the observation period started with the initiation of chemotherapy ( h) and continued for h after completion of the chemotherapy for acute phase, and during five days after finishing chemotherapy for delayed phase. the response rate of the study drugs was evaluated by the following -grade scale based on the condition of nausea and vomiting: highly, moderately or slightly effective and not effective. results: the best results were observed (triple drugs combination) in the early phase after beam conditioning and in delayed phase after mel high-dose chemotherapy. conclusion: triple drugs combination was more effective than palonosetron (+ dex) treatment in preventing cinv after high-dose chemotherapy before haematopoietic stem cell transplantation. cento hospitalar e universitario de coimbra, coimbra, portugal introduction: high-dose therapy with autologous stem cell transplantation (asct) has been considered a standard frontline treatment for younger multiple myeloma patients (mm) with adequate organ function; although several studies on safety are available, few reports focus on immediate complications following asct. our aim is to evaluate the early complications requiring hospital re-admission in the first days following asct in mm patients and its relationship with age and survival. material (or patients) and methods: we conducted a retrospective analysis of patients with mm who underwent asct between - - and - - , who were readmitted in the first days post-acst. survival was calculated according to kaplan-meier and log-rank methods. results: in these years, mm patients underwent asct, with a median age of . years (range - years) and a male/female ratio of . . high-dose melphalan conditioning regimen was used, followed by asct. half of the patients were discharged in the days after asct (range - days). median survival was not yet achieved in this population (median follow-up . years). in the first days post asct there were hospital admissions corresponding to patients ( , %). median hospital stay was days (range - days). the majority of those were due to infectious complications (n = ), renal insufficiency (n = ), thrombotic events (n = ), gastro-intestinal (gi) bleeding (n = ) and other causes (n = ). the main site of infection was gi tract (n = ), followed by respiratory (n = ) and urinary tract (n = ) infections. only in case (gi infection) was an infectious agent identified -e. faecium. there was death registered in this group of patients, attributed to disease progression. patients with hospital admissions in the first days post asct had shorter median survival ( , months) than those who were not admitted (p = , ). there was no statistical significant difference when comparing age (under or above years), the number of previous lines of treatment, melphalan dosage ( mg/m vs mg/m or the number of mobilizations required. these factors were also not able to predict survival in our population. conclusion: asct is widely used in mm patients, and is normally well tolerated, however a small proportion of patients require early hospital readmission within the first days. in our group of patients this was the only factor associated with reduced survival. disclosure of interest: none declared. "a pilot study on the efficacy of lactobacillus brevis cd lozenges in preventing oral mucositis by high-dose chemotherapy with autologous hematopoietic stem cell transplantation" s. giammarco ,* , a. di giovanni , e. metafuni , p. chiusolo , s. sica hematology institution, università cattolica del sacro cuore, rome, italy introduction: treatment-induced oral mucositis is one of the most debilitating side effects from the patient's perspective and profoundly influencing qol. om is a pathological process characterized by mucosal damage, ranging from mild inflammation to deep ulcerations and affecting one or more parts of the alimentary tract, from the mouth to the anus, as a consequence of chemotherapy and/or radiation therapy. ( ) om is associated with pain, bleeding, dysphagia, infections and food intake impairment, which can limit adequate nutritional intake. om is strongly associated with bacteremia and sepsis due to e. coli, p. aeruginosa, and candida. two major events lead to om; one direct injury caused by the ablative regimen, and local infections that develop on the damaged mucosal surface, which are exacerbated by neutropenia. treatment and prevention of om are palliative and the efficacy of most of the measures used have not been clearly established. prophylactic measures employed are use of: chlorhexidine, soda bicarbonate rinses, acyclovir and ice. for treatment of om and its associated pain local anesthetic, diphenhydramine, nystatin, or sucralfate are used alone or in combination as mouthwash. several studies showed the successful treatment of om with the strain of lactobacillus brevis, due to the process of microflora manipulation. ( ) the strain l. brevis produces high levels of the enzymes arginine deaminase and sphingomyelinase. arginine deaminase converts arginine to ammonia and citrulline, reducing the amount of available arginine to be converted to nitric oxide, a major mediator of inflammation. sphingomyelinase can hydrolyze platelet activating factor, a potent inflammatory cytokine, known to be associated with oral mucositis in radiation therapy. the aim is to assess if the probiotic l. brevis cd lozenges can reduce the incidence and severity of om in patients undergoing hsct. we enrolled patients (pts), affected by multiple myeloma and submitted to autologous hsct at our division between march and february . eight pts received l. brevis cd lozenges per day, from days prior chemotherapy and till resolution of om and reported all symptoms on a daily diary. introduction: safety of the discontinuation of ruxolitinib (ruxo), a jak / inhibitor, prior to allogeneic hematopoietic stem cell transplantation (allo-hsct) for myelofibrosis (mf) is still controversial. "jak allo study" reported serious adverse events in early phase during allo-hsct, indicating a rapid disease progression or withdrawal symptom by the discontinuation of ruxo, while such events were not described in several retrospective studies. furthermore no previous study evaluated the safety of the discontinuation of ruxo prior to allo-hsct in association with changes in serum levels of cytokines or growth factors. material (or patients) and methods: we conducted a pilot study in two patients with primary mf treated with ruxo prior to allo-hsct. ruxo was tapered off days before the initiation of reduced intensity conditioning regimen using fludarabine ( mg/m ), intravenous busulfan ( . mg/kg) and gy total body irradiation. primary endpoints were the following: ) engraftment and survival at weeks after allo-hsct, ) analysis of serum cytokines and growth factors before and after ruxo and during allo-hsct. this study was approved by the ethics board in our hospital and the study protocol is registered in university hospital medical information network clinical trials registry (umin ). results: both patients improved splenomegaly after the administration of ruxo and no disease progression or withdrawal symptom was developed after the discontinuation of ruxo (figure) , and achieved engraftment with complete donor chimerism by day after allo-hsct. serum levels of vegf, il- and sil- r were decreased after the administration of ruxo. however, those of il- , sil- r and mcp- were significantly increased after the discontinuation of ruxo. il- and sil- r persisted higher serum levels until day after allo-hsct, while those of mcp- was decreased to baseline on day . serum level of vegf was also slightly increased after the discontinuation of ruxo; however, the elevation was temporary and showed a stable transition during allo-hsct ( figure) . introduction: chylothorax is lymphatic fluid leakage into pleural space with high triglyceride content and milky appearance. etiology could be congenital, trauma, malignancy or granulomatous infections. this report describes multifactorial etiology of bilateral chylothorax in a child after stem cell transplantation (sct). material (or patients) and methods: a years old male with monosomy positive refractory aml had undergone sct from a matched related donor with total body irradiation based conditioning and secondary prophylaxis with voriconazole. results: at months posttransplant with % donor chimerism and monosomy negativity he had dysphagia, vomiting, anal pain and fever which prompted ampirical antibiotics. on the second day facial angioedemateous like swelling, resistant to steroids and antihistaminics was noticed with no sign of extramedullary relapse in paranasal and neck ct. on the th day he developed diarrhea. although rectal biopsy proved severe intestinal gvhd no therapy was given as diarrhea resolved. fifteen days later liver function tests increased and biopsy verified gvhd that lead to tacrolimus and steroid treatment. while performing liver biopsy milky appearing fluid was noticed. abdominal us showed perihepatic fluid ( mm). invasive aspergillosis history and newly developed cough directed a thorax ct which displayed bilateral pleural ( mm in left, mm in right) effusion and mm cavitary lesion in the left lung. thoracocentesis drained chylohemorrhagic fluid with a triglyceride content of mg/dl and galactomannan and aspergillus pcr positivity. central venous line was removed for leakage suspicion. as new lesions under voriconazole have emerged, liposomal amphotericin b (l-amb) and fatrestricted oral diet and ocreotide were started. in the meantime although morphologically in remission monosomy positivity was detected that encouraged tacrolimus withdrawal. at st month of ocreotide and l-amb treatment he developed fever with increased coughing. thorax ct showed ground glass appearance and enlarged effusion ( mm left, mm right). inflammatory fluid was drained and caspofungin was added to l-amb along with wide spectrum antibiotics. no microorganism was detected in further tests. besides ocreotide he had tpn nil by mouth for days which caused decrease in pleural effusions ( mm left, mm right). he received combined antifungal therapy for weeks with resolution of the lesions. repeated bone marrow examinations showed remission and monosomy negativity. he is now well at th months posttransplant with minimal pleural effusion receiving ocreotide and cyclosporine for upper gi gvhd. conclusion: chylothorax is an uncommon cause of pleural effusion in children. as treatment should be based on determining the underlying cause, it is of paramount importance to define the etiology. in the absence of apparent risk factors such as trauma this could be difficult. that is particularly challenging in the transplant setting since the etiology could be multifactorial as in our patient. although the beginning was chylous ascites the bilateral nature of chylothorax precluded us from considering it being the origin. we assume that the reason of bilateral chylothorax in our patient is iatrogenic trauma by tbi as well as noniatrogenic trauma by forceful emesis and coughing associated with gvhd and aspergillus infection where all could be common in the posttransplant setting. disclosure of interest: none declared. total body irradiation (tbi)-based versus chemotherapybased myeloablative conditioning regimen before allogeneic stem cell transplantation (allo-sct) for acute myeloid leukemia (aml and years (in bam one). there were no significant differences between the two groups regarding the stage of the disease. most of the patients were followed for nhl ( nhl and mdh in the bam group, nhl and mdh in the beam one). results: study of the short-term toxicities reveals that conditioning bam is more toxic considering platelets reconstitution with a median of days for the bam versus . days for beam (p = . ) for platelets above g/ l, a median number of transfusion . versus . mcp (p = . ), and a higher number of patients with severe mucositis (p = . ). nevertheless, there is no significant difference in hospitalization duration, aplasia length, and short-term infectious complications. long-term toxicities will be developed, available in march with a follow-up of year. conclusion: in conclusion, bam conditioning regimen induces longer thrombocytopenia, higher transfusion needs and increasing risk of bleeding and transfusion complications. the gastrointestinal toxicity is increased with more frequent and more severe mucositis, promoting undernutrition. a complement of this study will be presented regarding the long-term effectiveness of this type of procedure and its delayed toxicity. cd + : . % ( ) cd + cd + : . % ( ) cd + cd + : . % ( ) cd -cd + : . % cd + : % cd + igd + cd -: . % cd + igd -cd + : . % cd + igd + cd + : %. cd + cd + ra + : . % cd + cd -cd ra + : . % cd + cd ra -: . % the last ivig transfusion was on d+ and his igg is mg/ dl with normal igm levels. he was readmitted in the hospital due to cmv reactivation on day + which was treated by using ganciclovir, also he developed hypertension, hyperkalemia as side effects from the medications and local reaction at the site of bcg vaccine on d + that persist few days then resolved. the patient is now d + post-transplant doing well. introduction: allogeneic transplantation (allo-sct) in multiple myeloma is controversial mainly due to the relatively high non-relapse mortality (nrm). treosulfan is a prodrug of a bifunctional alkylating agent which has both myelotoxic and immunosuppressive properties ( ) . recently a regimen of moderate dose treosulfan has been used in combination with fludarabine and atg for treatment of malignant lymphoma and a small number of patients with multiple myeloma with low transplant related mortality ( , ) . the aim with this retrospective single-center study was to evaluate recent transplants particularly after conditioning with treosulfan plus fludarabine (treoflu). material (or patients) and methods: the study comprised patients (median age: years) that underwent allo-sct between and . twenty-five patients were in very good partial response or better (≥ vgpr) and in less than vgpr. results: the years overall survival (os) for the patients allografted after and before year was % vs % respectively (hr = . ( % ci . - . , p = . ) and for patients transplanted in ≥ vgpr and p = . introduction: prognosis of relapse of hematologic malignancy after allogeneic stem cell transplantation remains dismal. second allogeneic transplantation represents a valid clinical option in such patients. toxicity of the preparative regimen for second transplanation may sometimes limit a decision making process in offering the patients a second graft. goal was to evaluate the toxicity and tolerability of treosulfan, fludarabine and rabbit atg (thymoglobuline, r-tg) preparative regimen in the setting of the second allogeneic stem cell transplantation for disease relapse. material (or patients) and methods: we evaluated all patients who underwent second allogeneic transplantation for the relapse of their malignant disorder after the first allograft during years - at our department. preparative regimen in all patients consisted of treosulfan g/m for days, fludarabine mg/m for days and r-tg mg/kg. all patients received calcineurin inhibitor and mycophenolate mofetil as gvhd prophylaxis. day + and + mortality, platelet and neutrophil engraftment as well as infectious and acute non-infectious complications were recorded. results: we identified patients ( males and female), median age years ( - years) who underwent second allogeneic graft for relapsed disease after the st transplantation. patients had acute myeloid leukemia or myelodysplastic syndrome, patients had acute lymphoblastic leukemia, had chronic lymphocytic leukemia and primary myelofibrosis. patient were in subsequent cr of the disease and in relapse. patients received / matched unrelated peripheral blood stem cell graft and patients received / mismatched unrelated peripheral blood stem cell graft. all patients engrafted in platelets at median d+ ( - ) and in neutrophils at median d+ ( - ). median d+ donor chimerism was % ( - %), one patient experienced graft failure at d+ due to hhv- and subsequently died of hhv- encephalitis (confirmed). no other early deaths until d+ were observed. only patients had mucositis, only one being grade . all patients experienced episode of fever, of them diagnosed with g-bloodstream infection and of them with g + blodstream infection. patients had pneumonia. all patients recovered from their respective complications. overall d+ non-relapse mortality was . % ( / patients). conclusion: treosulfan, fludarabine and r-tg is feasible option in the conditioning of relapsed patients undergoing the second allograft. sunstained engraftment and acceptable early non-relapse mortality warrant its use in the setting of second transplant. infection remains the biggest challenge in this setting. disclosure of interest: none declared. splenic irradiation immediately before allogeneic stem cell transplantation in advanced phase myelofibrosis with massive splenomegaly m. parma ,* , c. julita , e. elli , r. renso , e. terruzzi , m. fedele , p. pioltelli hematology, radiotherapy, ospedale san gerardo, monza, italy introduction: allogeneic hematopoietic stem cell transplantation (hsct) in advanced phase myelofibrosis (mf) is a valid curative approach, but it is charged of an high risk of graft failure or rejection. this complications is though to be related to the particular condition of marrow micro-environment, but also spleen enlargement can play a role either in sequestering donor stem cells or in preserving autologous foci of stem cells. a proportion of mf patients (pts) can benefit from splenectomy before hsct but such procedures have been associated with substantial surgical morbility. considering these, we have tried an alternative approach with splenic irradiation immediately before hsct in a limited number of mf pts, and we evaluated the post transplant outcome. material (or patients) and methods: we selected pts with advanced chronic phase mf (according to cervantes scoring system pts intermediate- and pts high risk ipss) and one patient with acute evolution of mf; only one have been treated before with ruxolitinib, all of them have a great spleen enlargement ranging from to cm (ultrasonography measured). all pts received a schedule of fractioned splenic irradiation as a component of conditioning regimen ( gy divided in daily doses each of gy, between - to - days to transplant). conditioning regimen was busulfan-fludarabine for pts, reduced dose busulfan-fludarabine for patient and thiotepa-busulfan-fludarabine for patient. results: after fractioned splenic irradiation and fludarabinebased conditioning regimen, all pts showed remarcable spleen volume reduction within two months after hsct; / pts obtained successful donor cells engraftment, without severe complication related to splenic irradiation. median neuthrophil and platelet recovery ( /mm and . /mm ) was and days respectively. after months, chimerism status was % donor for pts and % donor for patient. three pts had a good outcome and are actually disease free with a complete chimeric status after , and months from hsct. one patient relapsed and rapidly presented a fatal leukaemic evolution two months after hsct, one patient relapsed with autologous haemopoiesis months after hsct. the only patient who did not engraft remained cytopenic surviving months. conclusion: splenic irradiation immediately before hsct could be considered a safe and alternative option to splenectomy for mf pts with massive splenomegaly, in order to obtain the improvement of donor engraftment. our limited experience shows that also these pts can have a favourable outcome. a largest number of cases would be required to discriminated the real advantage of this strategy. disclosure of interest: none declared. results of observational study on treatment patterns in adult patients with hematological malignancy (hm) undergoing hematopoietic stem cell transplantation introduction: available information on hsct comes mainly from us and european countries. the purpose of this multinational prospective observational study was to collect data on treatment patterns of hsct in hm in some non-us and non-eu countries. material (or patients) and methods: patients (pts) were planned. main eligibility criteria: adult pts with hm starting conditioning treatment in purpose of allogenic (allo) or autologous (auto) hsct, written informed consent. primary endpoint: to describe type of conditioning used in hsct. visit at and months after hsct were planned. results: eligible pts were recruited over months in countries, median age was years old (min-max: - ), ps was in . % pts and in . % pts. hm were multiple myeloma (mm) ( . %), acute myeloid leukemia ( . %), non-hodgkin lymphoma ( . %), hodgkin lymphoma (hd) ( . %), and acute lymphoblastic leukemia ( . %). myeloablative (ma) conditioning regimen was most frequently used in all tumor types (overall . % pts) and in allo and auto hsct ( . % and . %, respectively). source of hsc was peripheral blood in . % hsct. median day of engraftment was . graft failure was reported in pts ( . %) with allo hsct and pts ( . %) with auto hsct. in pts auto hsct from peripheral blood only, mobilization was chemotherapy (ct)+g-csf in . % pts, ct alone in . % pts and g/gmcsf alone in . % pts. cxcr antagonist was used in . % pts. median number of cd + cells prior first apharesis was /μl (min-max: . - ) and median number of apharesis was (min-max: - ). in pts allo hsct, . % had abo mismatch. gvhd prophylaxis was part of conditioning regimen in . % pts and was mainly rabbit atg ( . %). acute gvhd any grade was reported in . % pts and grade - in . % pts, and chronic gvhd any grade in . % pts and grade - in . % pts. majority of hsct were cmv d+/r+ ( . %). cmv prophylaxis at months was given in . % pts and median duration was days (min-max: - ). cmv reactivation was reported in pts and cmv infection in pts. febrile neutropenia, diarrhea and neutropenic bacterial infection were the most frequent events ( . %, . %, and . %, respectively). introduction: in europe rabbit anti-thimocyte-globuline (atg) is widely employed in patients submitted to matched unrelated donor (mud) hematopoietic stem cell transplantation (hsct) in order to reduce acute and chronic graft versus host disease (gvhd). atg seems to be very effective in reducing gvhd, but a significant increase of infective complications is often observed in these patients (pts). this fact and the reduction of "graft versus leukemia" effect make the use of atg in mud-hsct a matter of debate nowadays. we evaluated the immunologic recovery in pts treated with atg during the first year after mud-hsct and its correlation with the insurgence of opportunistic infection in the same period, excluding infection appeared before hematological uptake. material (or patients) and methods: we considered pts, affected by hematological malignancies, submitted to mud-hsct who received atg . mg/kg during the conditioning regimen. we discriminated two categories: pts who developed opportunistic infection ( pts) and pts without infection ( pts) in the first year after hsct. we analysed immunological recovery (lymphocyte sub population count) in these two series at two, four, six eight and twelve months after hsct. results: the more frequent infection is cytomegalovirus reactivation ( cases), the others are pneumocystis carinii pneumonia ( cases), ebv reactivation, other pneumonia, varicella-zoster reactivation. the majority ( / ) of pts who developed infections has been treated previously with atg. on the contrary, only a minority of pts ( / ) who didn't develop infections received atg. analysing data about immunological reconstitution, we have seen that pts who develop infections have a lower count of t cells and particularly of t , not only immediately after hsct but also during the entire observation time. b and nk cells and t /t ratio are lower in these pts for all the period, although the statistical significance is not reached due to the limited number of pts. conclusion: pts who receive atg have a higher susceptibility to infections respect to pts who do not. pts with infections have a slower immunological recovery than expected. we can conclude that pts who receive atg during the conditioning regimen should be strictly monitored for immunological recovery and for infection insurgence. the real role of atg in this prolonged immunological recovery remain to be defined. disclosure of interest: none declared. from mud ( , %) and from mmfd ( , %). stem cell sources included bone marrow (n = ), peripheral blood (n = ) and cord blood (n = introduction: as the survival of allo-hscts is constantly improving, more and more patients, especially the youngest, are susceptible to late complications. here, we present our institutional experience of late effects in the first years (y) following pediatric allo-hsct, comparing malignant versus non-malignant indications for hsct. material (or patients) and methods: all children allotransplanted between and were included in this single-center retrospective cohort. screening was carried out in accordance with international published guidelines. of transplanted patients, were included in the analysis; excluded due to short (o months) follow-up. results: median age at hsct was . y; the non-malignant group was younger. hsct indications were evenly divided between non-malignant (nmd) and malignant diseases (md) ( % of which were all). ten patients ( %) required a nd hsct. pre-transplant characteristics are listed in table . median follow-up was . y. twenty one patients ( %) had cgvhd; the rate of cgvhd was double in patients with md, and all severe cgvhd cases occurred in this group. the common affected organs were skin & joints, gi tract, lungs, [ab ] eyes & liver. late complications included hormonal dysfunction in patients ( %); % of gonadal insufficiency cases were in females. more than half ( %) had vitamin d deficiency. other involved systems included: cardiac ( %), pulmonary ( %), and skeletal (osteoporosis & avascular necrosis - %). the risk of late complications was not increased following nd hsct ( vs. %), although cgvhd incidence was higher ( vs. %). seven patients with md died during the follow-up period compared to case in the nmd group. see table for transplant related morbidity and mortality. conclusion: these data confirm that even in the first years following allo-hscts, pediatric patients are prone to a diverse array of complications, mainly endocrine. the underlying disease appears to affect morbidity: children with md have a two-fold risk of developing cgvhd and late effects (other than vitamin d deficiency), versus nmd. these differences could be attributed to higher cumulative induction/conditioning radio-chemotherapy dose in the md group. although the vast majority of deaths were in patients with md, most deaths were attributed to disease relapse; therefore no conclusion can be drawn regarding late transplant related mortality in md versus nmd. study limitations include the relatively small cohort, loss to follow-up and uncertainty regarding patient adherence to screening guidelines. as long-life follow-up is needed for this unique population, a continuing surveillance of the cohort study is planned [ab ] material (or patients) and methods: we aim to present the outcome of six pediatric patients with pid transplanted at the bone marrow transplant center in timisoara/ romania. three children with x-linked chronic granulomatous disease (cgd), one patient with griscelli syndrome type and hemophagocytic lymphohistiocytosis (hlh) and two other twins with il- receptor deficient t -b + nk + severe combined immunodeficiency (scid) were transplanted from matched related or unrelated donors at our center between - . all three patients with cgd received myeloablative regimens and the grafts were bone marrow in two and peripheral blood stem cells (pbsc) in one of the patients. the patient with griscelli received also a myeloablative regimen and pbsc.the two scid's had a non-myeloablative conditioning and were transplanted with bone marrow simultaneously. results: all of the patients engrafted, presenting complete donor chimerism early after the transplant. regarding transplant specific complications, two patients presented acute graft versus host disease and the child with hlh developed severe veno-occlusive disease early after the transplant. two out of the three patients with cgd are cured, the third died following severe pneumonia at months posttransplant with % donor chimerism and normal function of granulocytes and monocytes. one of the twins with scid is well with % donor chimerism and good immune reconstitution, the other one died of pulmonary complications, whereas the patient with hlh is also in remission, with a mixed chimerism. overall, four of six patients are alive and well with complete recovery of their severe immune deficiency. eleven patients developed acute gvhd (grade - ) and of them went on to develop chronic gvhd; all successfully managed with steroids and a combination of tacrolimus, cyclosporine and mycophenolate. interesting to note was that out of the patients with hlh/ lpd developed engraftment syndrome/ cytokine release syndrome. of note there was a high incidence of graft failures with primary and secondary graft failures. conclusion: in the absence of a matched sibling and matched unrelated donor, haploidentical parental stem cells are more readily utilized especially when the disease progression is rapid and a transplant is urgent. with improved techniques in stem cell manipulation haploidentical parental tranplatation offers a cure for these patients. however more work needs to be done in evaluating the engraftment and immune reconstituition in these patients. introduction: non-relapse mortality following allogeneic hematopoietic stem cell transplantation has rapidly decreased during the past years among pediatric cancer patients in developed countries, but little information about non-relapse or relapse-related mortality is available for pediatric cancer patients in developing countries. we aimed to estimate the cumulative incidence of non-relapse and relapse-related mortality following allogeneic hematopoietic stem cell transplantation among pediatric cancer patients in argentina. material (or patients) and methods: individuals eligible for our study were aged o years when diagnosed with cancer and underwent allogeneic hematopoietic stem cell transplantation between september , and may , at hospital de ninos sor marlia ludovica in la plata, argentina, which is a regional referral center for pediatric transplantation. our outcomes of interest were non-relapse and relapse-related mortality, where non-relapse mortality was defined as mortality from any cause other than cancer progression. these outcomes were prospectively documented following transplantation until july , . we used a competing-risks framework to estimate cumulative incidence and % confidence limits (cl) of nonrelapse mortality and relapse-related mortality. results: our study population comprised pediatric cancer patients, of whom % were aged o years at transplantation, % were male, % were diagnosed with leukemia, and % were transplanted with stem cells from related donors. we observed deaths ( relapse-related and nonrelapse) during person-years of follow-up (median = . years, interquartile range = . - . ). infections or infectionrelated consequences were the most common causes of nonrelapse mortality ( % of non-relapse deaths). the -year cumulative incidence of non-relapse mortality was % ( % cl: %, %) and the -year cumulative incidence of relapserelated mortality was % ( % cl: %, %). all non-relapse mortality in our population occurred within months of transplantation, whereas the latest relapse-related mortality occurred nearly years after transplantation. conclusion: our results suggest that relapse-related mortality is the predominant reason for mortality years post-allogeneic hematopoietic stem cell transplantation among pediatric cancer patients in argentina. nevertheless, non-relapse mortality is substantially higher in our population than posttransplant pediatric cancer patients in europe (cumulative incidence = . % ), and the predominant reason for mortality within the first year following transplantation. future research should aim to identify which modifiable factors can be targeted for intervention to help reduce non-relapse mortality following allogeneic hematopoietic stem cell transplantation in this population introduction: intra bone marrow transplantation (ibmt) of hematopoietic stem cells (hsc) might be a promising alternative to conventional intravenous application of hsc in order to improve the homing efficiency and to accelerate the early engraftment after transplantation. especially for transplantation of limited cell numbers the ibmt might prevent a delayed engraftment. here we investigated the impact of the graft infusion rate on the hematopoietic recovery as well as on the engraftment kinetics after allogeneic ibmt in a canine dla-identical littermate model following reduced intensity conditioning. material (or patients) and methods: dogs were conditioned with . gy total body irradiation. the graft volume was reduced by buffy coat centrifugation and dogs obtained x ml into the femur and humerus simultaneously. the infusion period of the graft was min in group (ibm , n = ) and min in group (ibm , n = ). cyclosporin a ( mg/kg, bid) was administered as immunosuppression from d- until d+ post transplantation. the hematopoietic recovery was monitored daily by blood counts and the development of the chimerism for the peripheral blood mononuclear cells (pbmc) and granulocytes was determined weekly. results: all animals of both groups engrafted. one dog died at d+ (infection) of the ibm group and was therefore not included. the median number of infused total nucleated cells was . * /kg in both groups (ibm : range . - . * /kg, ibm : range . - . * /kg; p = n.s.). the infused cd + cell numbers were median . * /kg (range: . - . * /kg; ibm ) and . * /kg (range: . - . * /kg; ibm ) (p = n. s.). time of leukocyte recovery was median d+ (range: - ) and d+ (range: - ) after and min infusion (p = n.s.). median leukocytes nadirs amounted to . * /l for ibm and . * /l for ibm (p = n.s.). the median duration of leukopenia ( o . * /l) lasted d for both groups (range: - d, ibm ; - d, ibm ) (p = n.s.). median platelet nadir was * /l for both cohorts (ibm : range . - . * /l, ibm : . - . * /l). the periods of thrombocytopenia ( ≤ . * /l) were d (ibm ) and d (ibm ; range - d, both; p = n.s. peripheral blood (pb) was the source of progenitor cells in patients, bone marrow (bm) in patients and cord blood in one patient. in allo hsct, patient transplanted / matched and patients / matched.gvhd prophylaxis regimen was cyclosporine+mtx. all patients engrafted. results: in allogeneic pbsct all patients' median time to absolute neutrophil count (anc) . × /l was days, and the median time to platelet count × was days vs and days in allo bm all patients. in allogeneic pbsct aml patients median time to anc . × /l was days, and the median time to platelet count × was days. e cd +/kg. ciclosporin a was given for gvhd prophylaxis, g-csf was instituted from day + ). results: hematopoietic recovery: for platelets /nl day ; /nl day ; for leukocytes . /nl day . pmns rose to . /nl on day , reached a maximum of . /nl on day , and then declined to - . /nl on day at which level they remained even under g-csf treatment. bone marrow cytology at this time showed exactly the same arrest of granulopoiesis at the level of promyelocytes, increased eosinophils and fully recovered erythro-and megakaryopoiesis as before sct. however, graft failure was ruled out as the bm cells expressed % donor chimerism. in search of an etiology, antibodies against granulocyte antigen hna- (cd ) were detected at titers of : , . plasmapheresis was started days post sct. the titers decreased to : , after the first, to : after the th, and to : or after the th (and final) plasmapheresis. peripheral pnm counts increased to . /nl after the th plasmapheresis, oscillated somewhat during weeks and remained within a range of . to . /nl since then. the patient successfully tackled a skin agvhd (stage ), cmv reactivation, adv infection and interstitial pneumonia, and is now alive, thriving and immunocompetent, at years post sct, enjoying stable pnm counts. conclusion: granulocyte transfusions -needed to surmount an appendicitis -apparently led to immunization against hna- in this child with scn, either because she was one of the % individuals who lack this glycoprotein, or because she has never expressed it since the very beginning of her embryonal granulopoiesis due to her underlying scn. the bone marrow cytologies of scn and of alloimmune neutropenia looked undistinguishably similar. the complete chimerism led us to correctly diagnose and treat this post sct enigma. disclosure of interest: none declared. results: the median mononuclear cell dose was . × /kg. the median time to reach absolute neutrophil count . × /l was days, and the median time to platelet count × was days. grade and grade mucositis was seen in % of our patients. ( ). we report the successful treatment of a patient with caebv (t-subtype). material (or patients) and methods: case report: a year old previously well female was referred with an ecog of and week history of pyrexia, weight loss and abdominal pain. this was secondary to hepatosplenomegaly, mild pancytopenia (hb . g/l, wcc . x /l, neutrophils . x /l and platelets of x /l), abnormal lfts (bilirubin mg/dl, alt iu/l, alk phos iu/l) and an ldh of iu/l. infection work up was negative apart from a positive ebv igg. her ebv dna viral load was . x . she was commenced on prednisolone and valganciclovir for four weeks for suspected infectious mononucleosis prior to referral with no improvement. a ct thorax/abdomen/pelvis confirmed hepatosplenomegaly and cm mesenteric/periportal adenopathy. a bone marrow aspirate and biopsy was performed which showed trilineage haematopoiesis with reactive features. the mesenteric node and liver were biopsied and were consistent with caebv, t-cell subtype with eber positive t-lymphocytes and clonality confirmed by tcr beta and gamma rearrangement. haemophagocytic lymphohistiocytosis was excluded as she had a ferritin o ng/ml, normal triglycerides and had no evidence of haemophagocytosis in the biopsies. results: she was treated with a hybrid protocol of gemcitabine, etoposide and dexamethasone for cycles because of her poor performance status and abnormal lfts and achieved a pr with improvement in lfts, blood count, scan and undetectable ebv dna. she was consolidated with a reduced intensity matched unrelated sct using fludarabine, busulphan and alemtuzumab conditioning resulting in engraftment at day . she had grade one skin graft versus host disease but no post-transplant ebv reactivation. she had lft normalization after six months and remains well eighteen months post-transplantation. conclusion: we have presented this case to raise awareness of a rare manifestation of ebv infection and effectiveness of allo-sct in its treatment ( ) . this case is unusual in that the patient had serological evidence of previous ebv infection with an unknown re-activation trigger, whereas it is usually triggered by a de novo infection. introduction: patients with relapse of high grade lymphoma after autologous transplantation or with primary refractory disease have a dismal prognosis. allogeneic hematopoietic cell transplantation (allo-hct) is a potentially curative salvage treatment but data on outcome with this approach are limited. material (or patients) and methods: we retrospectively analyzed all adult patients with high-grade lymphoma who underwent allo-hct at our center between and . results: a total of adult patients received allo-hct during this period. histologies were diffuse large b cell lymphoma (dlbcl, n = ), grey zone lymphoma (n = ), and transformation from indolent lymphoma (fl, n = ; cll, n = ; marginal zone lymphoma, n = ). the median age at time of allo-hct was (range, - ) years. % (n = ) of patients had received an autologous transplantation prior to allo-hct. if allo-hct was used as primary transplantation strategy, refractory disease or inadequate mobilization of hematopoietic stem cells was present. prior to allo-hct patients were treated with a median of chemotherapeutic regimens (range, - ) and % (n = ) of patients received rituximab prior to allo-sct. at start of conditioning therapy, % of patients had refractory disease and % were in complete remission. myeloablative conditioning regimens (mac) were used in % of patients and % were treated with reduced intensity conditioning (ric). there were no differences in median hct-ci score between ric and mac group (range - ). the median age for patients receiving mac and ric was years and years, respectively. peripheral blood stem cells were exclusively used as graft source. donors were matched related (mrd, n = ), matched unrelated (mud, n = ), mismatched unrelated (mmud, n = ), or haploidentical (n = ). median follow-up of patients alive was months with a kaplan meier estimated overall survival (os) for all patients of %, % and % and a progression-free survival (pfs) of %, %, and % at , , and years. transformation in high grade lymphoma did not result in inferior os (p = . ). the use of ric regimen resulted in significantly better median survival rates irrespective of disease risk index (mac months vs ric months, p = . ). the non-relapse mortality (nrm) at day , year and years was %, % and %. nrm after year was higher after mac with % compared to % after ric. the cumulative incidence of acute gvhd ≥ ii°was % and of chronic gvhd % (limited %, extensive %). after ric patients had increased rates of limited chronic gvhd with % vs. only % in the mac group. rates for extensive chronic gvhd were similar with % vs. %. rates for limited chronic gvhd were %, % and % in the mrd, mud and mmud group. extensive chronic gvhd was observed in % and % in the mrd and mud group. conclusion: allogeneic hct is a potentially curative treatment option even in advanced and refractory patients with a long term os of % and pfs of % regardless of conditioning regimen with ric resulting in significantly better os. disclosure of interest: none declared. early metabolic remission post autologous hematopoietic transplantation in hodgkin lymphoma conditioned with either beam or busulfan-based regimen is correlated with significantly superior disease free survival d. mallouri ,* , v. constantinou , m. iskas , i. batsis , s. papadimitriou , a. k. panteliadou , v. kalaitzidou , a. bouinta , s. papaemmanouil , p. palladas , i. datseris , a. anagnostopoulos , i. sakellari haematology department -bmt unit, pathology department, radiology department, george papanicolaou hospital, thessaloniki, nuclear medicine and pet/ct department, evangelismos general hospital, athens, greece introduction: fluorine- -deoxyglucose positron emission tomography scan (pet) is a valuable diagnostic tool in hodgkin lymphoma (hl). in the autologous haematopoietic cell transplantation (ahct) setting, the prognostic value of pre-ahct pet has been established by current studies, but the significance of early post-ahct pet negativity has not yet been fully investigated. material (or patients) and methods: in this retrospective, single center analysis, we evaluated the role of early post-ahct (+ to + months) metabolic remission in consecutive hl patients (pts), aged ( - ), who underwent ahct during - , for primary refractory ( ), or relapsed disease ( ) post ( - ) lines of treatment. disease status was chemosensitive to salvage treatment in pts. the conditioning regimen was beam ( ) or busulfan-based (busulfan . mg/kg, etoposide mg/m and melphalan mg/m , buem) ( ). results: forty eight pts were evaluated by pet at + to + months (m) while the rest had disease progression by ct assessment ( received additional involved field irradiation post-ahct, as part of the treatment plan). thirteen pts had positive and negative pet. the overall early-response rate by petevaluation was % ( / evaluated pts). in univariate analysis, overall survival (os) and disease free survival (dfs) were superior in pts achieving early post-ahct negativity ( y-os % vs %; p = . , y-dfs % vs %; p = . ). patients' characteristics according to the conditioning regimen were similar in terms of disease phase and chemosensitivity (x test). with a median follow up of ( - ) m (beam , buem m), the estimated year dfs and time to progression (ttp) were similar in both conditionings. at last follow-up / ( %) pts were alive and ( %) in cr. treatment related mortality was low with both regimens ( %). overall survival was high and similar in both conditioning groups, with no significant difference in dfs. in multivariate analysis disease phase, chemosensitivity, type of conditioning and early-pet evaluation were incorporated. early pet-negativity was the only significant factor correlated with superior os (p = . ). in terms of dfs, disease chemosensitivity identified as significant favorable factor (p = . ), while early metabolic remission was of borderline significance (p = . ). conclusion: with the limitations of this retrospective analysis, early metabolic complete remission, along with disease chemosensitivity to salvage treatment, seems to be the most significant predictor in hl pts for improved overall survival and time to disease progression. both conditioning regimens proved to be similarly efficacious and feasible, remaining the key players for a successful outcome of ahct in lymphomas. additionally, early pet can optimize the outcome in hl patients regarding the need for post ahct consolidation in the era of novel targeted agents. introduction: high-dose chemotherapy and either autologous or allogeneic hematopoietic stem cell transplantation (hsct) is one of the considered treatment options for patients with relapsed or refractory non-hodgkin's lymphoma (nhl) but there are limited data about this issue in the pediatric setting. material (or patients) and methods: we examined the role of hsct for patients aged less than or equal to years at the time of transplantation with relapsed or refractory lymphoblastic (n = ), burkitt ( ), diffuse large b cell ( ) and anaplastic large cell lymphoma (n = ), receiving autologous (n = ) or allogeneic (n = ) (n = matched sibling/family, n = unrelated and n = mismatched related donor) registered at the turkish pediatric bone marrow registry. risk factors affecting survival were evaluated using stratified cox regression. fifty-seven patients ( %) had chemo-sensitive disease at the time of transplantation. results: overall survival (os) and event-free survival (efs) were % and % with a median follow-up of months; os and efs according to histopathological classification were % and % for lymphoblastic, % and % for burkitt, % and % for anaplastic large cell and, % and % for diffuse large cell lymphoma, respectively. the outcomes were similar in regard of both os and efs in patients receiving autologous and allogeneic hsct ( % vs % and % vs %, respectively). both os and efs among patients with chemosensitive disease at the time of hsct were significantly higher than those in patients with chemo-resistant disease ( % vs %, p = . ; % vs %, p o . , respectively). multivariate analysis showed that chemoresistant disease at the time of transplantation was the only factor predicting limited both os (hazard ratio = . ) and dfs (hazard ratio = . ). conclusion: intensive chemotherapy followed by either allogeneic or autologous hsct is an effective strategy for children with relapsed or refractory nhl and offers durable disease free survival for a significant group of pediatric patients, especially for patients with chemosensitive disease at the time of transplantation. material (or patients) and methods: newly diagnosed patients with aggressive nhl were included from march to december in our center. before proceeding transplantation, cases of patients in cr are , cases of cr , cases of pr and cases of nr. all patients were been treated by chemotherapy of three times (ctod or moed) plus highdose mtx ( - g/m ) before transplantation. ceac is considered as the preparative regimen that including additional local supplementary radical radiotherapy for patients with large mediastinal masses. the total four weeks biological treatment including five days of il- and two days of rest, that was applied for the patients with hematopoietic recovery. the patients who showed cr before and after transplantation adopted consolidation treatment every three months, of which total courses are to and to . patients who showed pr following transplantation accepted autologous transplantation in months after transplantation. patients who presented nr or re accepted allogeneic transplantation. results: all patients acquired hematopoietic reconstitution, in which the average reconstituted time of anc was + . d and plt was + . d respectively. the main complications are tolerable, including gastrointestinal reactions ( . % of incidence rate), oral mucositis ( . % of incidence rate) and mild liver damage ( . % of incidence rate) which does not affect the progress of sequential treatment. the median follow-up time was months (range - months). among cr survivors, patients are disease-free ( . %) and survivors with disease( . %) and no death; cr survivors are disease-free ( . %), cr survivors with disease ( . %) and deaths ( . %); pr survivors are disease-free ( . %), pr survivors in disease ( . %) and deaths ( . %); nr survivors are disease-free ( %), nr survivors with disease ( %) and deaths ( %). the overall survival and progression-free survival of years for all patients are . % ( % ci . - . %) and . % ( % ci . - . %), respectively. conclusion: sequential asct showed its efficacy in treatment for aggressive non-hodgkin's lymphoma by increasing the cure rate of disease. disclosure of interest: none declared. hematology, hospital ampang, selangor, malaysia introduction: high-dose chemotherapy with autologous stem cell transplantation (asct) remains the standard of care for patients with multiple myeloma (mm) in malaysia despite the promising development of novel agents. here we present the treatment outcomes of our mm patients who have undergone high-dose chemotherapy with asct for the last years with different induction regimens. material (or patients) and methods: we retrospectively analyzed a total of patients who received high-dose melphalan (hd-mel) followed by asct from august till october in our centre. results: the majority of the patients were from the malay ethnicity, comprising of . % of the cohort. most of our patients were above years old with . % in the - years age-group and . % aged above years respectively. the induction strategy prior to asct has evolved from vad regimen accounting for . % to thalidomide(t)based or velcade (bortezomib) (v)-based regimens totaling . % with time. about . % of the patients had dual-novel agents (both thalidomide and velcade) as induction treatment prior to asct. there were no significant age differences among the groups. the velcade-based and dual-novel agent groups had more male patients, accounting for % and % respectively. a majority had igg subtype mm (vad %, thalidomide-based: %, velcade-based: % and dual-novel agent: %). at least / of patients in all groups achieved vgpr after induction with the highest cr/ncr among patients receiving dual-novel agents at induction. more patients in the thalidomide-based, velcade-based group and dual-novel group attained cr/ncr ( %, % and % respectively) as compared to vad regimen ( %) prior to asct. all patients successfully underwent pbsc mobilization and proceed with asct. the median day of neutrophils engraftment was day earlier for thalidomide-and velcade-based as well as dualnovel group as compared to vad for which median engraftment was day+ . the incidence of trm was %. the overall survival (os) of all mm patients from the time of diagnosis was % at months. the median survival post transplant was months from the time of transplant. conclusion: asct remains a relevant treatment option to deepen the response post induction and to prolong survival of mm patients in resource-limited settings such as malaysia as compared to maintenance with new novel agents which are not as easily accessible as compared to asct. disclosure of interest: none declared. comparison of double transplantation-tandem transplantation or transplantation in relapsein patients with multiple myeloma a. jungova ,* , s. vokurka , m. schutzova , k. steinerova , l. mohammadova , m. karas , d. lysak , p. jindra hemato-oncology department, charles university hospital pilsen, pilsen, czech republic introduction: multiple myeloma (mm) is malignant haematological disease affecting up to people a year in the czech republic. treatment options are constantly improved but autologous transplantation (tx) remains an integral part of treatment protocols despite the development of modern drugs. profit in using two autologous tx versus single tx is documented but without a recommendation of time interval between these two tx. we compared groups of patients with tx in our studyin the first group ( patients) was performed second tx calculatedly on reaching pr and worse after induction in median of . months ( - ) . in the second group ( patients) was performed second tx in relapse without previous re-induction in median months ( - ). subanalysis divided this group in subgroups according value of paraprotein before the second tx (cutt of for …low level" of paraprotein was g/l,event. mg/l in mm with light chains), other findings were taken as "high level" including patients with extramedullary disease. other parameters were comparable (age, type of induction treatment, cytogenetic risk) without statistic difference in both groups. material (or patients) and methods: retrospective analysis of patients with mm in order to compare time to initiation other treatment after last autologous tx (tfs), overall survival (os) in two different groups and baseline level of paraprotein by second tx in patients transplanted in relapse. results: the analysed group consisted of patients. group was divided in two parts. in patients in the first part autologous tx was performed as tandem tx (n = ), in the second part was until the time of relapse (n = ). median follow-up . months ( - ) vs. months ( - ) -p = . . in october lived / patients ( %), died patients ( %). median age in both groups is years (p = . ), representation of gendersthe ratio of men/ women / vs. / (p = . ), stage according iss / ( %) vs / ( %) (p = . ). cytogenetic favourable aberrations in / ( %) vs. / ( %), p = . and unfavourable findings in / ( %) vs. / ( %) (p = . ). median tfs was months vs. months (p = . ) and median os was not yet reached vs. months (p = . ). at the dividing of second group in two subgroups -"low level of ab introduction: angiogenesis plays an important role in the growth and metastasis of colorectal carcinoma, which is currently one of the targets of cancer therapy. it has been reported that the cd (endoglin) involved in angiogenesis and is a powerful marker for angiogenesis in colorectal carcinoma. level quantitative angiogenesis in peritumor and intratumor area is important to know because it is closely related to the micro environmental factors that influence the occurrence of cancer angiogenesis. the goal of this study analyze the distribution pattern of angiogenesis in colorectal cancer by comparing the distribution of angiogenesis in peritumor and intratumor areas between well, moderate and poorly differentiated colorectal carcinoma, and between carcinoma with metastasis and nonmetastasis. material (or patients) and methods: this study analysed fifty samples of resected colorectal adenocarcinoma. angiogenesis was assessed by immunohistochemical method using a primary monoclonal antibody cd . positive expression of cd was assessed through the cd protein expression in vascular endothelial cells, while the distribution pattern of angiogenesis assessed by counting the positive expression of cd protein in hot spots by using the mvd (microvessel density) in the peritumor and intratumor areas and then performed statistical analysis. results: there is a significant difference between quantitative level of angiogenesis in peritumor and intaratumor areas of well (p o . ), moderate (po . ) and poorly ( o . ) differentiated adenocarcinoma. significant difference between the quantitative levels of angiogenesis in peritumor and intratumor areas of non-metastatic colorectal cancer (p o . ) and lymph node metastases (o . ) was found, but not in colorectal cancer with liver metastatis. conclusion: the results of this study provide a fundamental data regarding the distribution patterns of angiogenesis in colorectal carcinoma that may be beneficial in development strategies of antiangiogenesis. disclosure of interest: none declared. disclosure of interest: none declared. references: . therapy-related acute myeloid leukemia with t( ; )(q ;q ) after treatment for osteosarcoma second malignant neoplasm in patients with osteosarcoma of the extremities treated with adjuvant and neoadjuvant chemotherapy disclosure of interest: none declared. ab outcome of the pediatric patients with normal karyotype acute myeloid leukemia in yeungnam region, south korea: multi-center retrospective analysis disclosure of interest: none declared. ab salvage therapy with allogeneic hematopoietic cell transplantation for relapsed/refractory aggressive b-cell lymphomas: a hematopoietic stem cell transplantation for refractory or recurrent non-hodgkin lymphoma in children and adolescents allogeneic haematopoietic transplantation (allo-hsct) may be a therapeutic option because of the inability of current therapies to eradicate myeloma cells. material (or patients) and methods: our study retrospectively evaluated a distinct group of patients with relapsed/refractory multiple myeloma who underwent an allo-hsct after - lines of treatment addressed to reduce the burden disease results: overall, pts ( %) achieved a cr; pts ( %) had a pr and pts ( %) showed a persistent refractory disease. pts in cr ( %) relapsed after the transplantation. the cumulative incidence of acute gvhd (agvhd) grade ii-iv was %. about half pts developed chronic graft versus host disease (cgvhd), with an equal frequency as limited or extensive and a cumulative incidence of %. trm before day th regarded cases due to agvhd grade iv. nrm was %. pts in cr with extensive cgvhd died because of obliterans bronchiolitis at + and + months respectively. interestingly, additional pts in cr with oral cgvhd incurred a fatal tongue neoplasia and a non-fatal intestinal neoplasia. the relapse and/or progression of disease were observed in pts ( %) and resulted the cause of death in pts ( %). a total of ( %) patients are still alive and ( %) pts are in a complete remission state. the median time of follow-up was months. trm at and years was respectively % and %. os at five years was %. pfs at years was % most of the patients ( / ) received high dose melphalan and autologous stem cell transplantation before allo-sct. tumor status at allo-sct: complett remission (cr) n = ( %), very good partial remission (vgpr) n = ( %), partial remission (pr) n = ( %), progressive disease (pd) n = ( %) trm in the acut gvhd group was %. chronic gvhd was %. the incidence of cgvhd among the three-year survivors was %. two patients was given dli during of progression or disease relapse without effectivity. there was no significant difference neither in os nor in pfs depending from the donor type, conditioning therapy, time from the diagnosis to allo-sct, age (under/above years), cytogenetic abnormalities and number of therapies before allo-sct. conclusion: trm is very frequent in pr and pd groups. in cr group trm is zero so it seems to be important to treat the patients before allo-sct until cr or vgpr. allo-sct is useless for the patients with pd. chronic limited gvhd is necessary for long survival. dli treatment could be given for patients with residual disease after allo-sct routinely conclusion: hematopoietic stem cell transplantation can lead to durable remissions in children and adolescents with leukemia and increase in survival of children. pbsct in childhood all was consistent with significant faster anc and platelet recovery in allogeneic pbsct, hospitalization was shorter. longer follow-up is required to evaluate fully efficacy and long-term results. disclosure of interest: none declared. acute myeloid leukemia in a year old patient previously treated for osteosarcoma -case report c. dragan ,* , m. murat , d. georgescu , m. tevet , f. mihai hematology, colentina clinical hospital bucharest, bucharest, romania introduction: as patients with osteosarcoma become longterm survivors, increasing attention has turned to the burden of late effects. recent studies showed an increase in the incidence of secondary malignant neoplasms in patients with osteosarcoma compared with the general population. the risk of developing leukemia was reported to be in an increasing rate in the last decade. different molecular and pathogenic agents may be implicated in the development of second malignant neoplasms in long-term survivors of osteosarcomas. some authors believe that the treatment with topoisomerase ii inhibitors may have a role in the development of saml,while others talk about cisplatin-associated saml. [ , , ] material (or patients) and methods: case report. in this report we present the case of a year old patient diagnosed in the hematology department of colentina clinical hospital in february with acute myeloid leukemia following the treatment (chemo-and radiotherapy) for an osteogenic osteosarcoma performed years earlier.results: in this report we present the case of a year old patient diagnosed in the hematology department of colentina clinical hospital in february with acute myeloid leukemia following the treatment (chemo-and radiotherapy) for an osteogenic osteosarcoma performed years earlier. the cytogenetic examination revealed an abnormal karyotype with chromosomes (xxy). the molecular biology exam did not find mutations in the flt itd, flt d and npm a genes. after the first induction course of chemotherapy the patient achieved complete remission, which was consolidated by more courses of chemotherapy and an allogeneic stem cell transplant. conclusion: considering the prior treatment for the osteosarcoma consisting of radiotherapy and chemotherapy we considered this to be a secondary acute myeloid leukemia, this representing a major negative prognostic factor, along with the cytogenetic abnormalities found. these factors strongly indicate that the first complete remission should be consolidated by allogeneic stem cell transplant. the particularities of this case are the association of the two malignancies in a young patient, the presence of the abnormal karyotype (hyperploid), and also the good response of the patient to the induction therapy, achieving complete remission after the first induction course, and of course, sustaining that response. tandem auto transplants of peripheral blood stem cells for lymphoblastic lymphoma: a single center retrospective study j.-l. li , q. wen , l. zhu , x. xiang , l. gao , l. gao , d. zeng , c. zhang , p. kong p , h. liu , x. zhang ,* department of hemaotlogy, xinqiao hospital,third military medical university, chongqing, china introduction: in order to evaluate the effecacy and safety of tandem autologous peripheral blood stem cell transplants (apsct) for lymphoblastic lymphoma. material (or patients) and methods: the clinic data of cases with lymphoblastic lymphoma in xinqiao hospital were analyzed. the patients were classified by ann arbor staging system, including stage iii cases, stage iv cases. before the first transplantation, cases were achieved complete remission and cases achieved partial remission. mobilizetion regimen was mit+vds+vp +dex+mtx combined with g-csf ug/(kg.d). the conditioning regimens for the tandem transplantation were ccnu+vp- +ara-c+ctx and ida+ara-c +ctx respectively. the interval of the tandem transplantation was to months. in tandem transplantation, the cells of mnc transfused were . × /kg ( . ~ . ) × /kg and . ( . ~ . ) × /kg; cd + cells were . ( . ~ . ) × /kg and . ( . ~ . ) × /kg respectively (p . ). results: all patients obtained prompt and sustained hematopoietic reconstitution. the interval of anc ≥ . × /l was + d(+ ~+ d ) days and + d(+ ~+ d)days; platelet ≥ × /l was + d(+ ~+ d)days and + d(+ ~+ d) days, respectively (p . ). after the tandem transplants, cases relapsed, among case died of recurrence, case died of disease progression, case died of transplantation related mortality, cases were still in disease-free survival, remained stable after median followup of ( ~ ) months. the -years disease-free survival (dfs) was . %. the -years overall survival (os) was . %. the main complication of transplantation were fever ( . %) and gastrointestinal symptoms (nausea ( . %) and vomiting ( . %)). there was no transplantation related mortality. conclusion: the tandam apsct was safe and effective for lymphoblastic lymphoma. disclosure of interest: none declared. centro hospitalar e universitário de coimbra, coimbra, portugal, centro hospitalar e universitário de coimbra, centro hospitalar e universitário de coimbra, coimbra, portugal introduction: at the present time multiple myeloma (mm) is a consensual indication for high dose chemotherapy (hdc) followed by autologous stem cell transplantation (asct). however, most of the larger trials evaluated the role of asct in mm patients with years of age (y) or younger. once mm is frequently diagnosed in the elderly, it is crucial to understand if there is a role for asct in an older group of patients. in this study we aimed to compare the immediate toxicity and long term results of mm patients submitted to asct according to their age. material (or patients) and methods: we retrospectively evaluated all mm patients eligible to hdc and asct in our centre between january/ and december/ . analyzed data: age at the time of asct, number of hospitalization days (hd), number of days with neutropenia o , x /l, number of days with thrombocytopenia o x /l, gastro-intestinal toxicity, oral mucositis, need for endovenous antibiotherapy, number of packed red blood cells (rbc) and platelet's concentrate transfusions, need for parenteral nutrition, treatment related mortality (trm), progression free survival (pfs) and overall survival (os). for the purpose of toxicity analysis, all the transplants were considered (n = ) whereas for the outcome investigation, only first transplants were analyzed (n = ). results: the male:female ratio was . . at the time of the asct the median age was y . patients were o y and ≥ y. the elderly group had higher number of hd (mean days vs days; p = . ). the number of days with thrombocytopenia o x /l ( vs days) and with neutropenia ( vs days) and the number of rbc units transfused ( . vs . ) were superior in the older group as well (p = . , . and po . , respectively). there were no significant differences regarding other toxicity related parameters and the trm was null in both groups. regarding long term outcomes, pfs was similar in both groups (median pfs was . y in the o y group and . in the ≥ y, p = . ). we did not find statistically significant differences in os as well (median os not reached in both groups, p = . ). conclusion: despite the higher hd and hematological toxicity in the ≥ y group, there was no major impact in patients' prognosis and also no increase in trm. on the other hand, the outcome was comparable in both groups. even aware of the limitations regarding the small sample size and the retrospective analysis, our experience indicates that acst is safe and effective in mm patients older than y as in the younger group. disclosure of interest: none declared. are timing and quality of response predictors of asct outcome in multiple myeloma patients?a single centre experience a. f. baptista ,* , c. geraldes , r. introduction: autologous stem cell transplantation (asct) remains a standard of care for multiple myeloma (mm) patients who are eligible to receive high-dose therapy. factors that can predict success of this approach are still object of various studies. we aimed to investigate if early asct or better response to this therapy were predictive of treatment success. material (or patients) and methods: we retrospectively reviewed the outcome of first asct in mm patients from - . all tandem transplants were excluded. groups analyzed: ) patients receiving asct less than year (y) after diagnosis (o y) vs the ones who received asct ≥ y after and ) patients who reached very good partial response or better (≥ vgpr) after asct vs the ones with inferior responses (ovgpr). responses were evaluated days after asct according to the international myeloma working group response criteria. results: there were patients with a median age at the time of the transplant of y [ - ] and male:female ratio of . . in cases asct took place o y and in ≥ y. three patients were excluded from this analysis due to lack of clear information. we found that reached very good parcial response or better (≥ vgpr) and inferior responses ( ovgpr). median progression free survival (pfs) was superior in the early asct group ( . y vs . y; p = . ). overall survival (os) was also superior in the o y asct (not reached vs y, p = . ). median pfs was higher in the ≥ vgpr group ( . y vs . y, p = . ). os was slightly better in the ≥ vgpr group but there was no statistically significant difference (median os not reached in any group; p = . ). conclusion: both timing of transplantation and quality of treatment responses seemed to be good predictors of clinical outcomes. early transplantation associated with better pfs and os. moreover, the better response group showed higher pfs and os, even though only superiority of pfs was proven statistically significant. although being a small and retrospective study with considerable limitations, our experience favors the effort to proceed to asct as soon as possible and to achieve better responses in order to obtain a more effective control of the disease. disclosure of interest: none declared. treatment of bence-jones multiple myeloma with chemotherapy with/without stem cell transplantation-treatment of bence-jones multiple myeloma with chemotherapy with/without stem cell transplantation-single centre experience a. zivanovic-ivic , d. introduction: bence-jones myeloma multiplex is a progressive disease characterized by excesssive numbers of abnormal plasma cells in the bone marrow and overproduction of incomplete immunoglobulins, containing only the light chain portion of the immunoglobulini. this type of myeloma occurs - %. the median overall survival is approximately years. pts outcome in bencejones myeloma has been remarkably s improved due to the use of combination therapies including chemotherapy and stem cell transplantation. material (or patients) and methods: a retrospective analysis of outcome of treatment bence-jones myeloma. since . until . we treated pts ( men and female), average age , years.iss pts, iss pts, iss pts.renal insufficiency was present in pts. high risik pts was defined by the presence of following factors:b m , mg/l, albumineo , gr/dl, crp , high lactat dehydrogenase, renal failure,stage iii.pts treated with induction,consolidation and maintenance therapy.conventional induction treatments were applied as following regimens: vad( ), mp( ), ctd ( ), pad ( ),tad( ). results: clinical response was achieved in pts ( %),while in pts ( %) established disease was resistant. sct had been done with pts ( %), while pts ( %) were treated with conventional chemotherapy adjusted to the vital age and comorbiditity. in the group of pts with transplantation done tandem had been carried out with pts and secondary sct had been done in relapsed pts. with pts with tandem sct allogenic (singen) sct had been done. trm is %. maintenance therapy had been done in pts.impact of high risk factors on outcome/pfs/os was of no significance, except for elevated high lactat dehydrogenase values that are associated with rapid relapse. the presence of bone lesions associated with short pfs and with no difference in os. in the group of transplanted no difference in os with/without a response to primary therapy, but in the nontransplanted pts os was significance shorter in patients who did not achieve a response.the transplanted patients had significantly longer pfs (mediana months vs months, p o , ) and longer os (mediana months vs months, po , ). pts ( %) of treated pts are living, while pts ( %) died. univariate log. regres. analysis showed that non-transplant patients are , times more likely to terminate lethal compared to transplant patients (rr , ( %c.i. , ) , p o , ). conclusion: our study showed asct is a more effective method of treatment of patients with bencejones myeloma compared to the conventional chemotherapy, but the results are still unsatisfactory. impact of high risk factors on outcome/ pfs/os was of no significance, except for elevated high lactat dehydrogenase values that are associated with rapid relapse. one of the major efforts to improve the results of intensive therapy and asct involves the integration of novel agents (proteasome inhibitors and immunomodulatory drugs) into the transplantation sequence. disclosure of interest: none declared. the outcome of bortezomib-based regimen for the first time induction chemotherapy in patients with multiple myeloma after auto sct l. ni ,* , y. luo , y. tan introduction: the outcome of multiple myeloma (mm) has been significantly changed by novel agents. bortezomib, as the representative, has been recommended as first-line treatment in guideline for many years. but whether bortezomib-based regimen as first induction chemotherapy has different outcome remains unknown. material (or patients) and methods: we retrospectively analyzed patients with mm undergoing autologous stemcell transplantation to compare different outcomes in patients treated with bortezomib-based regimen (n = ) for the first time induction chemotherapy to those without (n = , including cases administrating bortezomib later because of ineffectiveness of original plans or relapse) after newly diagnosed between and . results: before transplantation, all patients achieved at least partial response (pr) according to international myeloma working group uniform response criteria. the median (range) numbers of peripheral blood mononuclear cells and cd + cells in each group were . and . × /kg (p = . ), . and . × /kg (p = . ), respectively. the median (range) time to reach a neutrophil count . × /l was ( . - ) and ( - ) days (p = . ), respectively. meanwhile, platelets ( × /l without transfusion lasts for days) recovered in average time of . ( . - . ) and ( - ) days (p = . ), respectively. the median overall survival (os) after transplantation had not been reached in bortezomib-based group, and months in group without bortezomib. -year os and -year os showed no difference ( % vs %; % vs %, p = . , respectively). median time to disease progression in two groups was both months. conclusion: bortezomib-based regimen as the first induction chemotherapy may have no significant benefit in terms of os and ttp. but further prospective randomized trials and multicenter researches are still required to verify whether this regimen make sense or not. disclosure of interest: none declared. conclusion: the role of allo-hsct in multiple myeloma is strongly discussed and frequently neglected. due to of small numbers, heterogeneity of patients and follow-up from periods with different therapeutic protocols, there is no conclusive consensus of allo-hsct implementation. our retrospective analysis suggests that in the setting of primary refractory disease allo-hsct is probably ineffective. however, in the contest of responsive therapy disease allo-hsct could determine cr and possibly offer permanent cures.the current use of new agents can determine a more incisive reduction of burden disease before allo-hsct trough increasing its efficacy. disclosure of interest: none declared. the relation between initial f-fdg pet/ct findings and baseline clinical parameters in multiple myeloma t. tuglular ,* , f. pepedil tanrikulu introduction: in a recent study, a positive correlation was reported between baseline fluorodeoxyglucose (fdg) positron emission tomography (pet) findings and clinical parameters in patients with multiple myeloma (mm). fdg-pet findings were associated with disease outcomes. material (or patients) and methods: we aimed to investigate the correlation between baseline fdg-pet findings and clinical parameters in our mm patients. we conducted a retrospective analysis in a total of patients that were followed up between january and december . forty-eight chemotherapy naïve patients were found to have baseline fdg-pet evaluation. we compared the initial clinical characteristics of these patients with their pet findings: the number of focal bone lesions (fls) and the maximum standardized uptake value (suvmax). results: twenty-three out of patients were male. the median age was years. among patients with secretory disease ( . %), iga, igg, and light chain only disease were evident in . %, . %, and . %, respectively. the median percentage of bone marrow plasma cells was % (range - ). median serum beta- microglobulin was . mg/l (range: . - . ) and % of patients had international staging system (iss) stage . durie-salmon (ds) stage iii disease was detected in . %. forty-one ( . %) had bone lesions on fdg-pet imaging. of those, ( . %) had more than three fls. the median suvmax for those fls was . (range: . - ). the number of fls on pet correlated only with ds stage iii disease. conclusion: a statistically significant positive correlation was found between baseline fdg-pet findings and ds stage in mm patients. disclosure of interest: none declared. outcome of myeloablative allogenic hematopoietic cell transplantations in multiple myeloma z. csukly ,* , a. barta , a. batai , l. gopcsa , l. lengyel , p. remenyi , m. reti , e. torbagyi , t. masszi dept. of haematology and stem cell transplantation, st. laszlo hospital, budapest, hungary introduction: the purpose of this study was to assess prognostic factors which determine overall survival (os) and progression free survival (pfs) of patients with multiple myeloma who received myeloablative conditioning and key: cord- - gwbc z authors: zecha, judith a. e. m.; raber-durlacher, judith e.; laheij, alexa m. g. a.; westermann, anneke m.; epstein, joel b.; de lange, jan; smeele, ludi e. title: the impact of the oral cavity in febrile neutropenia and infectious complications in patients treated with myelosuppressive chemotherapy date: - - journal: support care cancer doi: . /s - - - sha: doc_id: cord_uid: gwbc z febrile neutropenia (fn) is an inflammatory response causing fever that may develop during cancer therapy-induced neutropenia. fn may herald life-threatening infectious complications and should therefore be considered a medical emergency. patients presenting with fn are routinely subjected to careful history taking and physical examination including x-rays and microbiological evaluations. nevertheless, an infection is documented clinically in only – % of cases, whereas a causative microbial pathogen is not identified in over % of fn cases. the oral cavity is generally only visually inspected. although it is recognized that ulcerative oral mucositis may be involved in the development of fn, the contribution of infections of the periodontium, the dentition, and salivary glands may be underestimated. these infections can be easily overlooked, as symptoms and signs of inflammation may be limited or absent during neutropenia. this narrative review is aimed to inform the clinician on the potential role of the oral cavity as a potential source in the development of fn. areas for future research directed to advancing optimal management strategies are discussed. fever is a manifestation of multiple pathways including the release of pro-inflammatory cytokines (including interleukin (il)- , il- , and tumor necrosis factor (tnf)-alpha) as a consequence of infection or inflammation [ ] . fever that develops during cytotoxic cancer therapy-induced neutropenia, or "febrile neutropenia" (fn), is a medical emergency. fn may be an early and only indication of a serious infection, since profoundly neutropenic patients are unable to mount a robust inflammatory response, and signs and symptoms of inflammation are typically attenuated or absent. in these patients, infections may rapidly progress into life-threatening complications (e.g., systemic inflammatory response syndrome (sirs), (severe) sepsis, and septic shock). it is critical to early recognize fn (oral temperature > . °c or two consecutive readings of > . °c for h and an absolute neutrophil count < μ/l or expected to fall below this threshold), and depending on the estimated risk to develop life-threatening complications, initiate empiric systemic broad-spectrum antibacterial therapy and other supportive care measures promptly [ ] . moreover, fn is associated with higher morbidity, unplanned or prolonged hospitalization, potential intensive care admission, and can necessitate chemotherapy (ct) dose reductions and/or treatment delays, which may lead to poorer clinical outcome and survival [ , ] . if a patient presents with fn while on myelosuppressive ct, one should assume that this is due to infection, particularly if there is no evidence for an alternative explanation (e.g., transfusion reactions, medication allergies and toxicities, vasculitis or other inflammatory conditions, and tumor(lysis)-related fever) [ ] . nevertheless, patients must be carefully evaluated for potential infection. these patients are routinely subjected to careful history taking, physical examination, chest xray, and microbiological evaluations. however, infections can be documented clinically in only - % of febrile episodes, whereas a causative microbial pathogen cannot be identified in the majority (> %) of cases [ ] [ ] [ ] [ ] [ ] [ ] , and the cause of the fever may remain unidentified (fever of unknown origin; fuo). neutropenia risk depends on patient-, tumor-, and treatment-related factors. although ct dose intensity and myelotoxicity represent significant risk factors, neutropenia and its complications may occur after treatment with almost any type of ct [ ] . however, not all neutropenic patients develop fever, and not all episodes of fn progress into lifethreatening sepsis. ct regimens can be classified as high, intermediate, and low risk for fn [ ] . those with > % fn risk are considered high risk. this includes, but is not limited to, patients treated with high-dose ct with or without hematopoietic stem cell transplantation (hsct). most ct regimens used for the treatment of adult solid tumors and lymphoma are rated as intermediate (between and %) risk for fn, whereas low-risk cytotoxic regimens are typically considered to be associated with fn risk < % [ , ] . tools aimed to identify patients with fn at low risk of serious complications have been developed [ ] . however, identification of high-risk patients remains a challenge [ ] . it is well recognized that oral mucositis (om) as well as gastrointestinal mucositis may cause fever [ , ] and predisposes to systemic translocation of microorganisms [ , ] . if a patient presents with fn, the physician routinely performs an oral inspection to assess whether ulcerative om or mucosal infections are present. however, fever may be also caused by inflammation and infection at other oral sites, including the periodontium, the dentition, and salivary glands. most frequently, this involves asymptomatic chronic infections that were present before the initiation of ct. these infections may exacerbate and present with redness, swelling, and pain [ ] , but they may also be asymptomatic or manifest with only minimal signs of inflammation. nevertheless, these predominantly asymptomatic infections (particularly those of the periodontium) may contribute to systemic inflammation [ ] and risk of bacteremia and are associated with fn [ ] [ ] [ ] . unfortunately, these infections likely remain undiagnosed by simple oral inspection [ ] . in order to ascertain diagnosis and management of these "hidden" oral infections, the national institute of health [ ] stated that all patients should receive a comprehensive oral examination by a dental professional and potential foci of infection should be eliminated prior to the start of ct. in most centers, however, this is may be only routinely performed in patients considered at high risk for complications: i.e., patients scheduled for hsct and those undergoing (chemo)radiation therapy to the head and neck, whereas patients treated with other myelosuppressive ct regimens are not typically referred for pre-treatment oral evaluation. it is thus conceivable that particularly in these patients, fn may have originated from an oral source that remained undiagnosed. this narrative review is aimed to inform the oncologist of current evidence of the contribution of oral infections and non-infectious conditions (e.g., om) to the development of fn and infectious complications associated with myelosuppressive ct. areas for future research directed to developing optimal management strategies are presented. oral homeostasis is maintained due to complex and finely tuned interactions between the host and resident microorganisms. having cancer, myelosuppressive cancer treatment regimens and concurrent medications, including antibiotics, disturb the equilibrium of the oral ecosystem via direct and indirect mechanisms, and contribute to the development of oral complications and infectious sequelae. the mouth houses a diverse microbial community, harboring over species of bacteria that colonize the hard surfaces of teeth and soft tissues as biofilms [ ] . residing microbes prevent colonization and overgrowth of opportunistic and pathogenic microorganisms. studies suggest that treatment with myelosuppressive ct induces alterations of the oral microbiome ( table ) . as a result of cytotoxic ct and other concurrent medications, patients may develop hyposalivation (reduced salivary flow) [ ] . saliva has a key role in maintaining a stable oral ecosystem as it contains peptides and proteins that have many different qualities. these include antibacterial, antifungal, and antiviral functions as well as lubricant, buffering, remineralization, and digestion properties [ ] . a lack of these salivary defense mechanisms may contribute to oral dysbiosis contributing to an increase of the incidence and severity of complications, including om, oral mucosal infections, and dental caries [ , ] . the oral cavity is the primary gateway to the human body as it is contiguous with the tonsils, pharynx, esophagus, eustachian tube, middle ear, trachea, lungs, nasal passages, and sinuses. microorganisms colonizing the oral cavity may migrate via contiguous epithelial surfaces to these anatomical sites [ ] . in addition, oral microorganisms and inflammatory products may translocate into the circulation through ulcerated periodontal pockets and breached epithelial lining of mucous membranes and disseminate systemically [ , ] . this may contribute to fever and infectious complications in neutropenic or otherwise immunocompromised patients [ , , ] and has been reported to contribute to increased risk to early non-tumorrelated mortality in hsct recipients [ ] [ ] [ ] . om is an inflammatory condition of the oral mucosa induced by ct and/or radiation therapy. most often, it manifests at the buccal and labial surfaces, ventral surface of the tongue, floor of the mouth, and the soft palate [ ] . mucosal changes can range from erythema to extensive ulcerations and hemorrhage. patients experience om as a painful condition associated with significant suffering and a negative impact on the quality of life [ , ] . ulcerative om may not only act as a portal of entry for microorganisms and inflammatory products, but as it often compromises food intake, oral medication use, and oral hygiene, it may also contribute indirectly to infection risk [ , ] . the majority of patients treated with high-dose ct regimens (with or without hsct) develop om (all grades), whereas its incidence in less intensive forms of ct is estimated to vary between and % [ , ] (summarized in table ). there is a temporal relationship between having ulcerative mucositis, fever, and neutropenia [ , [ ] [ ] [ ] . however, the role of neutrophils in the pathogenesis and resolvent of om has not yet been fully elucidated, and studies on granulocyte stimulating growth factors for the prevention and treatment of om show conflicting results [ ] . sonis suggested that cells rendered apoptotic or necrotic by ct, release ct-associated molecular patterns ("cramps"), which play an important role in initiating inflammation [ ] . bacteremia with cons may originate from oral mucosal membranes hsct [ ] immunocompromised patients including cancer patients [ ] hsct [ ] soga et al. [ ] mrsa was detected in the oral cavity following hsct, especially during om peak severity hsct ct chemotherapy, cons coagulase-negative staphylococci, hsct hematopoietic stem cell transplantation, mrsa methicillin-resistant staphylococcus spp., om oral mucositis when pattern recognition receptors (prrs) expressed by cells of the innate immune system within the mucosa (e.g., macrophages, neutrophils, and dendritic cells) are exposed to cramps, this results in the release of cytokines that act locally, and also may have distant effects by activating intracellular and intercellular signaling loops [ ] . these include the hypothalamic-pituitary-adrenal axis resulting in fever, and effects on the liver resulting in the generation of acute phase proteins. all of which can ultimately result in sirs and sepsis. indeed, blijlevens and colleagues reported that mucositis induces a systemic inflammatory response characterized by fever in neutropenic hsct recipients, even in the absence of bacteremia [ , ] . although om is not an infectious process, studies suggest that poor oral hygiene and pre-existing oral infections aggravate the severity and duration of toxicity induced mucosal reaction and thereby contribute to the risk to develop fn and infectious complications [ , [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] (table ) . in addition to fn associated with om, oral mucosal ulcerations with a non-mucositis etiology may induce fn. however, from a clinical perspective, it can be difficult to discriminate between such lesions as they may develop simultaneously. herpes simplex virus (hsv) is a major pathogen causing mucosal ulceration that may be confused with om or may aggravate this condition [ ] . most hsv infections occur as a result of reactivation of latent virus in hsv seropositive patients. upregulation of nf-κb resulting in loss of mucosal barrier in om may also induce signaling pathways that stimulate virus reactivation [ ] . hsv reactivation can be controlled with antiviral prophylaxis or treatment in most patients, although reactivation is still possible despite prophylaxis [ ] . herpetic lesions in patients receiving ct can be painful and may be associated with hemorrhage and necrosis, and bacterial and fungal superinfections. in myelosuppressed patients, hsv infection has atypical clinical manifestations, mainly manifesting as intraoral ulcerations with or without labial involvement. hsv infection may develop at sites, which are usually not affected by om (e.g., the dorsum of the tongue, gingiva, and the hard palate). herpetic lesions in compromised hosts may persist for extended periods until recovery from myelosuppression or appropriately treated. in addition, other herpesviruses, epstein-barr virus (ebv) in particular, may predispose for developing oral mucosal ulcerations during neutropenia and thus may directly or indirectly play a role in fn originating from the oral cavity [ ] . periodontal pockets [ ] and salivary glands seem important reservoirs of cytomegalovirus (cmv) [ ] . the virus can be shed (via gingival crevicular fluid and salivary glands) in whole saliva forming a main mechanism for transmitting cmv. cmv induces local production of pro-inflammatory cytokines and remains a major cause of morbidity in hsct recipients [ ] . oral candidiasis is typically caused by opportunistic overgrowth of candida albicans, c. krusei, c. tropicalis, c. dubliniensis, and other commensal oral yeasts. neutropenia, mucosal damage, depressed cell-mediated immunity, hyposalivation, immunosuppressive medications, and use of antibiotics are risk factors. the most common forms are pseudomembranous and erythematous candidiasis [ ] . during oral infection with candida spp., a large amount of inflammatory cytokines is generated in the oral mucosa, potentially inducing fever [ , ] . current prophylactic strategies have reduced systemic candidiasis, although oral and oropharyngeal candidiasis still may have serious consequences. good oral hygiene is important in addition to antifungal therapy. rarely, molds, such as zygomycetes, coccidiomycosis, histoplasmosis, and aspergillus spp., cause oral infections. typical lesions include ulcers in the oral cavity, tongue, or gingival regions that may progress into invasion of deep tissues, including bone. early recognition (and confirmation by biopsy), aggressive debridement, and antifungal therapy with extended spectrum azoles are paramount [ ] . pre-existing dental pathologies that may cause fn and infectious complications during myelosuppressive ct include periodontal disease, profound dental caries, and periapical pathology due to infection of the root canal, (partially) impacted teeth, and retained roots [ ] . gingivitis is a localized or generalized inflammation of the gingiva without loss of periodontal attachment, whereas periodontitis is a chronic inflammation of the gingiva, affecting the deeper parts of the periodontium. as dental plaque migrates in apical direction, periodontal attachment degrades, leading to ulcerative periodontal pockets, bone resorption, and tooth loss. about - % of adults has chronic gingivitis, around % suffer from moderate periodontitis, whereas the prevalence of severe periodontitis is estimated to range from epstein and schubert [ ] treatment of dental pathologies and reducing the oral microbial load resulted in reduction of incidence and severity of om hsct [ ] hematologic malignancies [ ] kashiwazaki et al. [ ] maximal crp levels in peripheral blood and incidence of fn is significantly lower in the group who performed intensive oral hygiene hsct santos et al. [ ] treatment of dental pathologies and reducing oral microbial load resulted in shorter duration of om hsct khan and wingard [ ] presence of oropharyngeal mucositis is an independent risk factor for bacteremia in neutropenic patients hsct, non-hodgkin lymphoma, leukemia laine et al. [ , ] association between the presence of oral mucosal ulcers and episodes of fever and sepsis lymphoma sonis et al. [ ] schuurhuis et al. [ ] om is associated with higher fn risk as compared with patients without om ruescher et al. [ ] elting et al. [ ] bochud et al. [ ] marron et al. [ ] om was most likely origin of bacteremia with oral viridans streptococci, frequently resulting in fever and potentially leading to ards and septic shock hsct [ ] patients treated with ct [ , ] hematologic malignancies [ ] ebinuma et al. [ ] maintenance of good oral hygiene may reduce the gene mediating methicillin resistance. the oral cavity may act as a reservoir for harboring this gene hsct fn febrile neutropenia, om oral mucositis, hsct hematopoietic stem cell transplantation, crp c-reactive protein, ards acute respiratory distress syndrome, ct chemotherapy to % in western populations [ ] . periodontitis risk increases with age (peaks between and years) and is more prevalent in men than in women. the severity of the periodontal disease depends on environmental and host factors (e.g., genetic susceptibility, comorbidities, oral hygiene, and smoking) [ ] . inflamed and infected periodontal tissues can expose up to - cm of connective inflamed tissue and may serve as a reservoir of endotoxin (lipopolysaccharide), pro-inflammatory cytokines, and proteolytic enzymes that may spread systemically via ulcerated pocket epithelium and contribute to endothelial dysfunction, a pro-coagulant state, and low-level inflammation [ , ] . periodontal infection has been associated with bacteremic seeding of heart valves and prosthetic devices [ ] , fuo, and a wide range of systemic conditions, including ischemic cardiovascular disease, metabolic syndrome and type diabetes, rheumatoid arthritis, and respiratory diseases, including aspiration pneumonia [ ] [ ] [ ] . table summarizes the literature on gingivitis and periodontitis and their association with bacteremia, fn and infectious complications. ulcerated periodontal pocket epithelium can allow translocation of microorganisms into the bloodstream [ ] . bacteremia may be caused by facultative or strictly anaerobic bacteria likely originating from the periodontium, including capnocytophaga spp. and fusobacterium nucleatum [ ] [ ] [ ] [ ] [ ] . however, bacteremia with these microorganisms is relatively uncommon, whereas bacteremias with viridans streptococci and cons occur frequently in profoundly myelosuppressed patients. increases of gingival viridans streptococci and cons have been documented during highdose ct [ ] , and one study reported that periodontitis may contribute to the risk of bacteremia with viridans streptococci and cons during the neutropenic phase of hsct [ ] . periodontal disease may be occult without symptoms and clinical signs that may only be detected upon comprehensive periodontal evaluation and not identified based on general oral examination. a number of studies on the potential role of odontogenic infections in the development of fn do not distinguish between periodontal disease and other infections related to the dentition (table ). there is evidence suggesting that infectious and inflammatory processes of the oral cavity may contribute to fn, particularly in patients treated with high-dose ct with or without hsct. however, most studies are retrospective and include only a small number of patients. well-powered, prospective observational studies in homogenous groups of patients, including those treated with myelosuppressive ct for solid tumors, are needed to obtain a better understanding of the contribution of om and oral infections to fn and its potential sequelae. particularly, om [ , ] and periodontal disease [ , , , , , , , [ ] [ ] [ ] seem important in the development of fn, whereas the literature on other potential oral sources of fn (such as mucosal infections, periapical pathologies, and pericoronitis) is scarce. some studies report raber-durlacher et al. [ ] presence of periodontitis is associated with the development of bacteremia. hsct laine et al. [ ] in % of the febrile episodes, mild to moderate gingivitis was present in % of the febrile episodes, severe gingivitis was present lymphoma hong et al. [ ] . % of patients had severe gingivitis pediatric malignancies peterson and overholser [ ] greenberg et al. [ ] overholser et al. [ ] bergmann et al. [ ] laine et al. [ ] meurman et al. [ ] soga et al. [ ] gingivitis and periodontitis were found to be associated with bacteremia, fever, and sepsis in neutropenic patients russi et al. [ ] fn may be induced by periodontal inflammation and infection head and neck cancer fn febrile neutropenia, hsct hematopoietic stem cell transplantation infrequent conversion of chronic dental disease to an acute state during ct and conclude that there is no need for the treatment of "asymptomatic" chronic dental infections prior to ct or conditioning therapy for hsct [ , ] . however, studies should not focus solely on these acute exacerbations, associated with swelling and pain, since the inflammatory response may be muted or absent due to myelosuppression. it is essential that endpoints of future studies should also include the presence of "asymptomatic" disease, particularly chronic periodontitis which requires comprehensive examination including periodontal probing and appropriate dental imaging. there is convincing evidence from the periodontal literature that gingivitis as well as periodontitis induces frequent episodes of bacteremia during daily activities such as chewing and tooth brushing [ , ] . in non-myelosuppressed individuals, these episodes of bacteremia are typically transient. in myelosuppressed and otherwise immunocompromised cancer patients, however, bacteremia risk is likely increased as in addition to breached epithalial barriers, these patients are less capable of clearance of microorganisms from the blood stream, and infectious complications may develop. in addition to hematogenous spread of oral microorganisms and inflammatory mediators, there is evidence that poor oral and dental conditions may predispose for aspiration pneumonia [ , ] . systemic dissemination of oral microorganisms should be investigated in more detail in cancer patients treated with myelosuppressive ct. future investigations, using new open-end next-generation sequencing techniques, should aim to further characterize the role of the oral/periodontal microbiome in the pathobiology of mucositis as well as in systemic infectious complications. among other factors, the use of prophylactic antibiotics has changed the spectrum of isolated pathogens from the bloodstream as the most common causes of bacteremia in myelosuppressed cancer patients are coagulasenegative staphylococci, viridans group streptococci, and enterococci [ ] , which may all originate from the periodontium and oral mucosa [ , , , [ ] [ ] [ ] [ ] [ ] . particularly, the contribution of staphylococcus epidermidis originating from the oral cavity should be studied. an intriguing hypothesis that needs further exploration is the concept that inflammatory complications induced by cytotoxic cancer therapies may have a common pathobiological background and share genetically determined risk factors [ ] . inflammation, particularly an imbalance between upregulated greenberg et al. [ ] patients who did not receive dental treatment prior to ct: -in % of patients, sepsis occurred. - % of patients developed acute exacerbations of oral bacterial infections. patients in which dental infections were treated before ct: -in % of the patients, sepsis occurred. -no acute exacerbations of oral bacterial infections occurred. laine et al. [ ] patient with febrile episodes had more severe dental pathologies than those without fever ( . % vs . %) toljanic et al. [ ] incidence of acute dental problems during febrile episodes of % solid and hematologic malignancies akintoye et al. [ ] out of positive blood cultures with microorganisms were likely to originate from the oral cavity; of these bacteremias the periodontium was the most likely origin. hsct cullen et al. [ ] in % of febrile episodes an oral infection was thought to be the focus of infection hong et al. [ ] weighted prevalence of dental infections of . % patients treated with cytotoxic ct hong et al. [ ] weighted prevalence of dental infections was . %; pericoronitis was present in . % of patients akashi et al. [ ] development of septic shock in patients of which it was assumed that this was associated with a chronic odontogenic infection. no bacterial evidence was found from the blood cultures. fn febrile neutropenia, ct chemotherapy, hsct hematopoietic stem cell transplantation pro-inflammatory cytokine pathways and suppressed antiinflammatory cytokines, may be a major common driver of these complications [ ] . in addition, macrophages have been identified to play a critical role in the potential bidirectional links between periodontal and systemic inflammatory diseases [ ] . it has been proposed that the presence of inflammation (anywhere in the body) primes for a dysregulated and exaggerated inflammatory response following a subsequent inflammatory stimulus [ ] . this "two-hit model" has been hypothesized to underpin a potential association between periodontitis and om [ ] . moreover, it has been suggested that patients with periodontitis are more prone to developing fever and septic shock syndrome, since endotoxins derived from periodontal pockets may lead to precipitating conditions [ ] . these intriguing findings and hypotheses deserve further exploration. an important clinical question concerns the management of potential oral foci of fn. there is a consensus that patients planned to be treated with high-dose ct with or without hsct, or those who will be treated with curative (chemo)radiation for head and neck cancers should be referred to a dental professional for a comprehensive oral and periodontal pre-treatment evaluation with appropriate treatment. in patients with head and neck cancers, rigorous elimination of potential foci of infection (i.e., in most cases by extraction) is mandatory in patients with at-risk conditions in the radiation volume [ ] , whereas in other patient categories, protocols may be less rigid. there is substantial evidence suggesting that intensive oral care performed before and during cancer treatment results in a reduction of infectious complications [ , , , , [ ] [ ] [ ] . elad and coworkers [ ] developed a predictive model suggesting that no dental treatment increased the probability of dying due to a dental/periodontal infection in hsct recipients. in selected cases, treatment of periodontal infections by debridement and reducing the oral bacterial load, and endodontic treatments may be preferred over extractions. in addition, approaches aimed at resolving periodontal inflammation hold significant promise [ ] . these interventions may also positively affect om. when fn (likely) originating from the oral cavity develops during ct treatment, antimicrobial therapy should be preferably based on microbiological evaluation of swabs and/or oral rinsing samples. in conclusion, current recommendations for the prevention of complications include dental evaluation and work up of high-risk patients as an integrated part of the patient workup and management. nevertheless, large longitudinal studies are needed to 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retrospective evaluation. radiotherapy and oncology : journal of the champlin r ( ) oral mucositis in patients undergoing bone marrow transplantation a prospective study to evaluate a new dental management protocol before hematopoietic stem cell transplantation progress of oral care and reduction of oral mucositis-a pilot study in a hematopoietic stem cell transplantation ward a decision analysis: the dental management of patients prior to hematology cytotoxic therapy or hematopoietic stem cell transplantation periodontitis: a host-mediated disruption of microbial homeostasis publisher's note springer nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations acknowledgements we kindly acknowledge support from the eklund foundation. conflict of interest the authors declare that they have no conflict of interest. key: cord- - nynqtq authors: moulton, bart; barker, alan f. title: pulmonary complications date: - - journal: blood and marrow transplant handbook doi: . / - - - - _ sha: doc_id: cord_uid: nynqtq after hematopoietic stem cell transplant (hsct), up to % of patients develop pulmonary complications. in spite of antibacterial, antiviral, and antifungal prophylaxis, reduced host defenses render the hsct patient vulnerable to pulmonary and other infections in the early weeks and even months post-transplantation. this chapter suggests an integrative approach followed by a description of the most common pulmonary syndromes seen in hsct patients, including diffuse alveolar hemorrhage (dah), idiopathic pneumonia syndrome (ips), bronchiolitis obliterans syndrome (bos), and cryptogenic organizing pneumonia (cop). the high risk of developing pulmonary complications after hsct necessitates a pre-transplant pulmonary workup. pulmonary function tests (pfts) are done prior to hsct and include spirometry and diffusion capacity of carbon monoxide (dlco). low dlco and alveolar–arterial oxygen gradient on pfts carry increased mortality post-hsct. decreased dlco and forced expiratory volume in one second (fev ) < % are indicators of developing respiratory failure post-hsct. investigating new pulmonary complaints is challenging. all patients should undergo an extensive workup of new pulmonary findings, including dyspnea, cough, fever, and hypoxia. in the first – weeks post-hsct, immunocompromised patients can develop bacterial pneumonia. pathogens include gram-negative rods (pseudomonas or klebsiella), staphylococcus aureus, and nocardia. while chest x-rays could show typical lobar or multilobar opacities, computed tomography (ct) scan of the chest (noncontrast ct scans are adequate for workup of infectious processes) may yield additional characteristic findings (nodules, ground glass opacities, etc.). fungal pneumonias, primarily aspergillus, can also develop in this early period. there is a very strong association between invasive aspergillus pneumonia and neutropenia lasting more than days. viral pneumonia may develop as well in this patient population; however, it tends to occur later. cytomegalovirus (cmv) is the most common viral pathogen, but with monitoring and preemptive therapy, the incidence has declined. other viruses have emerged as pathogens, including respiratory syncytial virus (rsv), influenza, parainfluenza, and human metapneumovirus (see chap. ). . spirometry is used to aid in the diagnosis of obstructive versus restrictive lung disease. two-year mortality after hematopoietic stem cell transplant (hsct) has been estimated using a pre-transplantation assessment of mortality (pam) score which incorporates spirometry and diffusing capacity variables in combination with the presence of renal and hepatic dysfunction, conditioning regimen, and disease risk. a. obstructive lung disease is diagnosed with a forced expiratory volume in one second (fev )/forced vital capacity (fvc) ratio < % and fev < %. if plethysmography (measurement of changes in lung volumes) is performed, increased residual volume (rv) indicates air trapping as seen in bronchiolitis obliterans syndrome (bos). b. low fvc with normal fev /fvc ratio indicates restriction. lung volumes will help confirm restriction as seen in idiopathic pneumonia syndrome (ips) or usual interstitial pneumonia. all lung volumes, including rv, will be reduced with restriction. . dlco a. diffusion capacity of carbon monoxide (dlco) corrected for hemoglobin should be used (dlcoadj) b. > % normal, - % mild, - % moderate, < % severe impairment . bronchoalveolar lavage (bal) via bronchoscopy should be pursued once pneumonia is considered. . pre-procedure stabilization with supplemental oxygen is key. a. depressed mental status may increase the risk of the procedure. b. the presence of severe hypoxia and depressed mental status may require endotracheal intubation to safely perform the procedure. c. conscious sedation with fentanyl and/or midazolam is often used for comfort and amnesia. . unless there is active bleeding, correction of coagulopathy is not required, and there is no absolute platelet level required for safety with bal alone. a. if transbronchial biopsy will be attempted, a pre-procedure platelet count of ≥ , /mm and international normalized ratio (inr) of < . is recommended. . complications of bronchoscopy include worsening hypoxemia, airway hemorrhage, and respiratory failure. . the risks with transbronchial biopsy are much higher, including pneumothorax, respiratory failure, and difficult to control airway bleeding. . electromagnetic navigational bronchoscopy with bal and biopsy may improve rates of diagnosis. . appropriately stained bal smears may suggest a pathogen in a matter of hours while cytology, culture, and genetic results are pending. bal fluid should routinely be sent for: a. cytology, including stains for organisms (fungi, pneumocystis jiroveci pneumonia (pcp)) and hemosiderin-laden macrophages b. bacterial cultures (including nocardia) and sensitivity c. fungal smear and culture d. mycobacterium smear and culture e. cell count and differential f. galactomannan antigen (aspergillus) g. polymerase chain reaction (pcr) for respiratory viral panels h. pcr for legionella i. direct fluorescent antibody (dfa) staining for pcp diffuse alveolar hemorrhage (dah) is a subset of pulmonary hemorrhage that can develop in up to % of all post-hsct recipients with mortality rates ranging between and % based on the two largest case series. about % of the cases develop in the first weeks post-hsct. chest x-ray often shows bilateral diffuse alveolar opacities which could be confirmed by ct scan imaging ( fig. . ) as ground glass opacities. these findings are not specific and may be seen in many other conditions. b. pulmonary function tests (pfts) show increased dlco; however, often these patients cannot participate in such testing. c. bronchoscopy with bal is the confirmatory diagnostic method. bal shows progressive bloody return. cytology with prussian blue staining should show > % hemosiderin-laden macrophages. this test is limited if alveolar hemorrhage occurred < - h before the procedure, as duration of time may be too short for red blood cells (rbc) phagocytosis by pulmonary macrophages. there is no clear etiology for dah post-hsct. the development of dah around the engraftment period suggests an inflammatory cascade involving the alveoli. pre-hsct conditioning regimens (including total body irradiation (tbi)) may initiate the inflammatory process. patients with suspected dah should be transferred to the medical intensive care unit, given that respiratory failure may develop rapidly. some patients require high-flow oxygen and subsequent mechanical ventilation for acute respiratory distress syndrome (ards). supportive management and high-dose systemic steroids are the key elements of dah treatment. a. mechanical ventilation should be tailored to each individual, reflecting the ards mechanical ventilation protocol/low tidal volume for management of acute lung injury. this practice has not been validated in dah, but the pathological pattern of dah is similar to acute lung injury/ards. similarly, prone positioning may be of benefit in refractory cases. b. immunosuppressive therapy with high-dose corticosteroids is the mainstay of therapy based on case reports and retrospective series. doses of up to g of methylprednisolone daily divided into - doses should be given daily for - days, followed by a slow taper over - months. alternate dosing schedules have been suggested, beginning at mg/kg daily in divided doses, tapering over a -month period. c. correction of underlying coagulopathy by maintaining platelet count > , / mm and inr < . d. bal to rule out a concomitant infectious pathogen. e. recombinant factor viia (novoseven ® ) has been used; however, no benefit has been demonstrated. f. aminocaproic acid (amicar®) has been used less frequently with limited supporting data. ips is severe lung injury that develops after allogeneic hsct with no evidence of an infectious process. the incidence ranges between and % with mortality rates ranging from to %. more recent studies report a lower incidence likely reflecting improved diagnosis of viral infections with newer pcr tests. if mechanical ventilation becomes necessary, mortality approaches %. ips typically occurs within the first months post-hsct. however, delayed onset has been reported. delayed pulmonary toxicity syndrome (dpts) is considered distinct from ips per the american thoracic society official statement due to its relationship with a specific conditioning regimen. dpts occurs in up to % in patients who receive a conditioning regimen containing bis-chloroethylnitrosourea (bcnu), cyclophosphamide, and cisplatin. . risk factors a. grade - agvhd b. donor cytomegalovirus (cmv) positivity c. conditioning regimens containing tbi d. older age e. certain malignancies (acute leukemia, myelodysplastic syndrome) f. drug toxicity has been implicated; however, there is no method to discriminate between drug-induced lung damage and ips. findings are indistinguishable from pneumonia which include fever, cough usually productive of scant or no phlegm, shortness of breath, and hypoxia. all patients with suspected ips should undergo chest imaging and bronchoscopy with bal to rule out infection. occasionally, chest x-ray does not show obvious infiltrates and ct scan of the chest is warranted. the criteria for diagnosis of ips proposed by the national heart lung and blood institute in include: a. radiologic imaging evidence of multilobar diffuse alveolar infiltrates. b. hypoxia or elevated alveolar-arterial gradient. c. negative bal for blood and cultures for bacterial, fungal, and viral pathogens. d. negative infectious studies from the blood, specifically for cmv. evaluation of bal fluid from ips patients shows elevated inflammatory cytokine markers compared to negative or healthy controls. ips is likely a complex cytotoxic and immune-mediated attack of the lung. a. corticosteroids should be started early in the disease course. historically, patients who developed ips around engraftment responded better to steroids. a reasonable starting dose is mg/kg daily of methylprednisolone (or equivalent) for the first week followed by a slow taper over the course of - months. b. pcp and fungal prophylaxis are recommended. c. etanercept (enbrel ® ) mg sq twice weekly for weeks has been used in conjunction with corticosteroids; however in small case series, no additional benefit was seen when compared with placebo. the most common late pulmonary complication following allogeneic hsct is bos. the reported incidence varies from to % with estimate as high as %. however, recent studies suggest the incidence is more prevalent than previously reported. the median time to onset is year post-hsct. however, the onset varies from months to > years post-hsct. bos is rarely reported after autologous hsct or umbilical cord blood hsct. most investigators consider bos to be gvhd of the lung. it is also important to recognize bos as a separate clinical entity from cryptogenic organizing pneumonia (cop). the national institutes of health (nih) diagnosis and staging working group prepared a consensus definition for bos to provide uniform inclusion criteria for future studies. to make the diagnosis of bos, these four criteria must be present along with active chronic gvhd in at least one organ other than the lung: a. fev < % of predicted normal b. evidence of airway obstruction with a ratio of fev /fvc < . c. expiratory high-resolution chest ct that reveals air trapping, small airway thickening, or bronchiectasis or rv > % of predicted normal d. absence of active infection or pathologic confirmation i. lung biopsy typically shows cicatricial bronchial obliterans (i.e., obliteration of airways by dense fibrous scar tissues) insidious course manifested by nonproductive cough, wheezing, and dyspnea. early in bos, pulmonary exam may be normal; however, later stages are manifested by wheezing, prolonged expiratory phase, and inspiratory crackles. a. chest imaging should be carried out in all patients undergoing workup for bos. chest x-rays may be normal early in bos. as the disease progresses, hyperinflation may be present. b. high-resolution ct (hrct) of the chest is more specific (see fig. . ) . inspiratory and expiratory phases should be included to evaluate for air trapping or "mosaic lung appearance" which indicates regional airflow obstruction during the expiratory phase. c. pfts are obtained as part of every patient's pre-hsct baseline evaluation. i. the definition of airflow obstruction includes fev < %, fev /fvc < . , or a decline in fev > % in year. ii. also noted is air trapping or increased rv and rv to total lung capacity (rv:tlc) ratio. iii. dlco is not expected to be reduced but is often low pre-transplant and/or after induction chemotherapy. d. bronchoscopy is not routinely performed during the workup of bos unless imaging is suspicious for an infectious process. e. transbronchial biopsy is often nondiagnostic as the disease process is patchy. f. surgical lung biopsy has higher chance of demonstrating constrictive bronchiolitis, the pathology seen in bos. i. with the introduction of hrct, surgical lung biopsy is often not required to confirm a diagnosis of bos. . pathogenesis of bos bos may be a manifestation of primarily chronic gvhd with the etiology related to recognition of disparate antigens present in the context of hla class i and class ii major histocompatibility complex (mhc) molecules. it begins with fig. . a inspiratory ct scan of the chest. b expiratory phase ct scan chest in a patient with bos. ct computed tomography, bos bronchiolitis obliterans syndrome fibroproliferative disease of the small airways, which results in inflammation, epithelial metaplasia, and denudification. submucosal/mucosal fibrosis then develops, resulting in obliteration of the airways. allogenic inflammatory conditions such as viral infections may also contribute to the development of bos. management of bos mainly involves intensifying immunosuppressive therapy and supportive care. there are no specific recommendations associated with treatment of bos. the management of bos mimics that of chronic gvhd. a. response to bronchodilators is often minimal but nevertheless should be considered because of presence of airflow obstruction. b. corticosteroids - . mg/kg prednisone per day for - weeks, then tapered over - months if there is a response. this regimen is based on case series and expert opinions. c. other immunosuppressive medications maybe effective as steroid-sparing agents, including calcineurin inhibitors. tacrolimus the management of bos is complicated and requires a multispecialty approach (bone marrow, pulmonary, and radiology specialists). prognosis of progressive bos (> % fev decline per year) is poor. two-year overall survival has been reported at % with a -year survival rate of only %. the majority of patients die of respiratory failure triggered by infection. attention to dyspnea and early and frequent pfts may allow for earlier identification of bos before permanent (fibrotic) airway changes, respiratory insufficiency, and pneumonia occur. cop, previously known as bronchiolitis obliterans organizing pneumonia (boop), is a disease process of unknown etiology that differs from bos in clinical findings, response to treatment, and prognosis. one case series of open-lung biopsies done in patients who underwent hsct found that cop was the most common inflammatory pathology ( %). . risk factors a. no risk factors have been identified. however, a correlation has been demonstrated between the development of cop and viruses, radiation exposure, connective tissue disease, inhalational drugs (cocaine), amiodarone, and inflammatory bowel disease. the presentation of cop is similar to many respiratory disorders; most commonly, dyspnea is accompanied by nonproductive cough and fever. physical exam is primarily notable for the presence of crackles and the absence of wheezing. . diagnostic tests a. chest x-ray may show patchy consolidation with ground glass or nodular infiltrates. b. ct scan of the chest is typically required to demonstrate areas of bilateral organizing pneumonia and consolidation in subpleural or peribronchial distribution associated with areas of ground glass opacities. migratory opacities on ct scan chest have been described in % of patients with cop. c. pfts typically show a restrictive pattern with decreased fvc, fev /fvc > %, and decreased dlco; airflow obstruction (decreased fev /fvc) is generally absent. d. bronchoscopy with bal may be helpful in determining the diagnosis. bal fluid demonstrates lymphocytes with a decreased cd /cd ratio. e. lung biopsy, either by transbronchial biopsy or by video-assisted thoracic surgery (vats), is occasionally required to confirm the diagnosis. typical pathology shows granulation tissue plugs in the bronchioles and alveolar ducts associated with surrounding chronic interstitial inflammation. a. prognosis of cop is favorable; % of patients can be expected to recover. b. bronchoscopy with bal is often required to rule out infectious processes. c. corticosteroids have been used with great efficacy. however, relapses may occur if steroids are tapered too rapidly. diffuse alveolar hemorrhage in hematopoietic stem cell transplant recipients bronchiolitis obliterans syndrome epidemiology after allogeneic hematopoietic stem cell transplantation nhlbi workshop summary. idiopathic pneumonia syndrome after bone marrow transplantation risks and outcomes of idiopathic pneumonia syndrome after nonmyeloablative and conventional conditioning regimens for allogeneic hematopoietic stem cell transplantation pulmonary complications of solid organ and hematopoietic stem cell transplantation recommended screening and preventive practices for long-term survivors after hematopoietic stem cell transplantation an official american thoracic society research statement: noninfectious lung injury after hematopoietic stem cell transplantation: idiopathic pneumonia syndrome a risk score for mortality after allogeneic hematopoietic stem cell transplantation clinical and radiologic distinctions between secondary bronchiolitis obliterans organizing pneumonia and cryptogenic organizing pneumonia bronchiolitis obliterans after allogeneic hematopoietic stem cell transplantation soluble tumor necrosis factor receptor: enbrel (etanercept) for subacute pulmonary dysfunction following allogeneic stem cell transplantation a randomized double-blind, placebo-controlled trial of soluble tumor necrosis factor receptor: enbrel (etanercept) for the treatment of idiopathic pneumonia syndrome following allogeneic stem cell transplantation. a blood and marrow transplant clinical trials network (bmt ctn) protocol bronchiolitis obliterans syndrome (bos), bronchiolitis obliterans organizing pneumonia (boop), and other late-onset noninfectious pulmonary complications following allogeneic hematopoietic stem cell transplantation key: cord- -uprv a k authors: brodszki, nicholas; turkiewicz, dominik; toporski, jacek; truedsson, lennart; dykes, josefina title: novel treatment of severe combined immunodeficiency utilizing ex-vivo t-cell depleted haploidentical hematopoietic stem cell transplantation and cd ra+ depleted donor lymphocyte infusions date: - - journal: orphanet j rare dis doi: . /s - - - sha: doc_id: cord_uid: uprv a k background: allogeneic hematopoietic stem cell transplantation (hsct) is the only curative treatment available for severe combined immunodeficiency (scid); although, there is a high incidence of severe infections and an increased risk of graft-versus host-disease (gvhd) with hsct. early intervention is a crucial prognostic factor and a hla-haploidentical parental donor is often available. haploidentical hsct protocols utilizing extensively ex vivo t-cell depleted grafts (clinimacs system) have proven efficient in preventing gvhd, but cause a delay in early t-cell recovery that increases the risk of viral infections. here, we present a novel approach for treating scid that combines selective depletion of gvhd-inducing alpha/beta (α/β) t-cells from the haploidentical hsct graft with a subsequent donor lymphocyte infusion (dli) enriched for cd ro+ memory t-cells. results: our patient was diagnosed with scid (t-b + nk+ phenotype). at months of age, he received a t cell receptor(tcr)α/β-cell depleted graft from his haploidentical mother, following a reduced intensity conditioning regimen with no additional gvhd prophylaxis. engraftment was rapid with complete donor chimerism and no signs of gvhd. however, at weeks post hsct, the patient was still t-cell lymphopenic with clinical symptoms of multiple severe viral infections. consequently, therapeutic dlis were initiated for enhanced anti-viral immunity. the patient was treated with cd ra+ depleted haploidentical maternal donor lymphocytes enriched from unmobilized whole blood, and a total t-cell dose of no more than x ( ) cd + cells/kg with > . % purity of cd + cd ro+ memory t-cells was transferred. following the dli, a prompt increase in cd + cd + and cd + cd + counts was observed with a subsequent clearance of viral infections. no acute or chronic gvhd was observed. conclusions: automated depletion of cd ra+ naïve t-cells from unmobilized whole blood is a simple and rapid strategy to provide unmanipulated dlis, with a potentially broad repertoire of pathogen specific memory t-cells. in the haploidentical setting, cd ra+ depleted dlis can be safely administered at low t-cell doses for efficient enhancement of viral immunity and limited risk of gvhd. we demonstrate the successful use of this approach following tcr-α/β-cell depleted hsct for the treatment of scid. severe combined immunodeficiency (scid) is a rare disorder defined by a profound developmental or functional defect of t lymphocytes. if unrecognized, it can cause death during the first years of life, due to the life-threatening increased susceptibility to infections [ ] therefore early detection and treatment of scid, before infections become overwhelming, optimizes survival [ ] [ ] [ ] . today, most forms of scid are treated with hematopoietic stem cell transplantation (hsct) and if hsct is not successful and genetic treatment is not an option, the alternatives are few [ ] . for patients lacking a human leukocyte antigen (hla)matched sibling or unrelated stem cell donor or when there is an urgent need for a hsct, an hla-haploidentical parental donor is often readily available [ ] . given the major hla donor-recipient disparities in the haploidentical setting, measures are required to prevent the occurrence of alloreactive responses, i.e. graft rejection or severe graftversus host-disease (gvhd). the extensive ex-vivo depletion of donor t-cells (clinimacs system) has proven efficient in preventing gvhd [ , ] , but is inevitably coupled to a delay in early t-cell recovery and, thus, to an increased risk of viral infections [ ] [ ] [ ] [ ] . recent efforts to balance the risk of gvhd against that of delayed immune reconstitution include the selective depletion of gvhdinducing alpha/beta (α/β) t-cells while retaining potentially beneficial gamma/delta (γ/δ) t-cells in the haploidentical graft [ , ] . unselected donor lymphocyte infusions (dlis), which are frequently used as a "tool" to boost anti-viral immunity post-transplant, harbor a significant risk of inducing severe gvhd in the haploidentical setting [ , ] . gvhdinducing cells reside mainly in the cd ra+ naïve t-cell population whereas viral-specific memory t-cells are predominantly cd ra-negative [ ] . thus, the depletion of cd ra+ t cells from dlis may provide a potentially broad repertoire of donor-derived viral-immunity with a limited risk of gvhd [ , ] . herein, we report on the successful treatment of scid by combining t cell receptor (tcr)-α/β-cell depleted haploidentical hsct with cd ra+ depleted dli for an antiviral boost. the male patient was born at full term, after a normal pregnancy, as the third child of afghanistani related parents. both parents and the siblings are healthy. he had his first upper respiratory tract infection (urti) at the age of weeks and at the age of month, he was admitted for the first time to a pediatric ward for weeks with coughing, low grade fever and hoarseness. in the following months, he was followed regularly due to coughing, hoarseness and failure to thrive. at the age of months, he was infected with chicken-pox. three weeks after the onset of the infection, new blisters were still forming and varicella keratitis developed. the unusually severe course of the infection and the failure to thrive warranted investigation for primary immunodeficiency disorder (pid), in conformity with our protocol [ ] . routine blood investigations during his first months of life showed normal leukocyte and neutrophil counts on several occasions. total lymphocytes were measured during the first admission at month of age, but the low value of . (ref: - . × /l) was attributed to an upper respiratory tract infection (urti) and was followed up only at the beginning of the chicken-pox infection when the level was . x /l. other laboratory tests, including electrolytes, alt, ast, gt, pancreatic amylase, bilirubin, organic acids, amino acids, and the thyroid hormone status, were normal. at the time point of pid investigation, he was anemic (hemoglobin: g/l) with normal platelets and a total lymphocyte count of . x /l. the immunological investigation revealed very low t-cell counts, confirming a t-b + nk+ scid phenotype. the cultures taken at this point showed multiple infections: positive staphylococcus aureus blood culture, cytomegalovirus (cmv), and varicella virus dna positivity in blood, varicella dna in the cerebrospinal fluid, pneumocystis jiroveci and coronavirus nl on throat swabs, rotavirus in feces, and high serum beta-d-glucan. genomic dna was prepared from blood collected in edta, fragmented to an average length of bp, and exome enriched using the sureselect xt human all exon v technology (agilent). sequencing was carried out to an average coverage of -fold using pe x bp sequencing (illumina hiseq ). the bioinformatic analysis was carried out using the mutation identification pipeline (mip) [ ] . results were presented in an interactive browser-based visualization tool (scout), developed in-house (science for life laboratory, solna, sweden). the clinical interpretation of the results was restricted to a predefined set of genes known to be involved in primary immunodeficiencies. sequence changes in exons and exon-intron boundaries were analyzed. only rare variants (allele frequency < . ) were considered. for recessive disease genes, at least two variants were required. the analysis was not designed to detect structural rearrangements or copy number alterations. the exon sequencing, restricted to the predefined set of immunodeficiency genes, did not identify any low-frequency variants that could account for the patient's clinical presentation. a homozygous missense variant (val phe) in the cd g gene, was identified, but it received a low rank score due to its reported high frequency of . in the genomes database and was, therefore, considered a natural variant. thus, no known genetic defect was identified. considering the low numbers of lymphocytes and the life-threatening infections, the decision was made to perform a hsct. the parents provided written informed consent for this procedure. no hla matched siblings were available and the probability of finding a matched unrelated donor was considered low given the patient´s ethnical origin. consequently, the choice was made to select a hla-haploidentical parental donor. the patient and his parents were investigated according to our institutional protocol for haploidentical hsct, including high resolution allelic typing for hla-a*, -b*, -c,* drb *, dqb *, bpb * (pcr-sequence specific primers [ssp], olerup ssp ab), hla-donor specific antibodies (dsa) (labscreen singel antigen, luminex platform, one lambda, thermo fisher), abo-blood grouping and cmv-serology. the mother was selected as the donor based on the outcome of the pre-transplant investigation, and this selection was further supported by the reported higher survival rate in haploidentical hsct patients grafted from the mother compared with recipients of paternal grafts [ ] . the hla-typing confirmed patient and donor haploidentity with a drb *, dqb *, and dpb * allelic match in the host-versus-graft direction. no dsa were detected in the patient´s serum. the patient and the donor were matched for abo-blood group and were both cmv positive. the patient was conditioned with a myeloablative regimen, according to guidelines [ ] regarding hsct for primary immunodeficiencies with a haploidentical family donor in patients older than months. however, due to his very poor health status, we decided to use a reduced intensity regimen conditioning. this consisted of i.v. anti-thymoglobuline (atg fresenius ®) given from day - to - (first day mg/kg bodyweight and then mg/kg bodyweight), fludarabine ( mg/m /day, from day - to - ), thiotepa ( mg/kg twice a day; day - ), and melfalan ( mg/m day - to - ). the sole form of gvhd prevention was the immunomagnetic depletion of tcr-α/β + cells from the graft. prophylaxis against posttransplant epstein-barr virus-associated lymphoproliferative disease (ebv-ptld) consisted of one dose of rituximab ( mg/m ) given at day + . no posttransplantation pharmacological immunosuppression was used. peripheral blood progenitor cells (pbpcs) of the haploidentical maternal donor were mobilized by granulocytecolony-stimulating factor (g-csf, μg/kg/day) and collected by leukapheresis (cobe spectra, terumo bct) on day of g-csf administration. collected pbpcs were processed under clean room conditions with the clini-macs system (miltenyi biotech), according to the manu-facturer´s protocol. the cells were sequentially labelled with tcr-α/β-biotin reagent and clinimacs anti-biotin reagent, and processed with the clinimacs device using the d . program with the depletion tubing set (dts). the graft cell contents were determined by flow cytometric analysis (facs, canto ii, becton dickinson [bd]), using antibodies against cd , cd , cd , cd , cd , cd , tcr-α/β and tcr-γ/δ with -aad as a viability marker (bd). according to our institutional protocol for haploidentical hscts, the target cell dose of the tcr-α/β-cell depleted graft was set to > × cd + cells/kg (minimum × cd +/kg) and < × tcr-α/β + cells/kg (maximum × tcr -α/β+ /kg). a bag of unmobilized whole blood was drawn from the donor. utilizing a closed blood bag system, whole blood was separated by density centrifugation to obtain a leukocyte enriched cell fraction for further processing with the clinimacs system. according to the manufac-turer´s protocol, cells were labelled with cd ra reagent and processed under clean room conditions with the clinimacs device using the d . program with the dts. the cell contents of the cd ra-depleted target fraction was assessed by facs (canto ii, bd), using antibodies against cd , cd , cd , cd ro, and cd ra with -aad as a viability marker (bd). engraftment following hsct was monitored by daily counts of leukocytes, neutrophils, and platelets. immune recovery was assessed post hsct and after dli by facs (canto ii, bd) of peripheral blood lymphocytes, using antibodies against cd , cd , cd , cd , cd , tcr-α/β, tcr-γ/δ, cd ra and cd ro (bd). donor myeloid and t-cell chimerism was monitored by dna genotyping of short tandem repeat polymorphisms (pcr-str, dx genetic analyzer, applied biosystems). the patient was evaluated for signs of acute and chronic gvhd. all procedures followed were in accordance with the ethical standards of the helsinki declaration of , as revised in . the regional ethical review board in lund, sweden advised us that signed consent from the patient's parents was sufficient for publication of this case and the consent was obtained. a single leukapheresis procedure provided × cd + cells/kg recipient weight for subsequent tcrα/β-cell depletion with the clinimacs system. the log tcr-α/β-cell depletion rate was - . , corresponding to . % tcr-α/β + cells in the target fraction. the recovery of cd + cells and tcr-γ/δ + cells was %. cell viability post-sorting was %. the tcr-α/ β-depleted graft contained per kg of recipient weight . × cd + cells, . × tcr-α/β + cells, . × tcr-γ/δ + cells, . × cd + b-cells and . × cd + nk-cells and was transplanted in total to the patient. the donor had antibodies against cmv, vzv ebv and herpes simplex. hematological recovery post hsct was prompt with a white blood cell count > × /l at day + , neutrophils > . × /l at day + , and thrombocytes > × /l at day + . complete myeloid and t-cell donor chimerism was reached at day + . as expected ( ), nk-cells co-infused with the graft expanded rapidly, with normal peripheral counts from day + (ref: . - . × /l). however, despite the prompt three-linear hematological recovery, the patient remained profoundly t-cell lymphopenic (fig. ) . atg clearance vary between individuals and the efficacy of atg varies between batches [ , ] ; therefore an explanation of the lymphopenia could be that since we used atg in a relatively high dose, given timewise quite close to the grafting, this affected the graft, hence could be considered as additional in vivo t-cell depletion. due to the persistent effect of rituximab, b-cell recovery was also markedly delayed. a single unit of donor whole blood provided a surplus of cd + cells ( . × /kg recipient weight) for subsequent cd ra+ depletion using the clinimacs system. the log cd + cd ra+ cell depletion rate was - . , corresponding to . % residual cd + cd ra+ cells in the target fraction. the recovery of cd + cd ro+ cells was %. cell viability post-sorting was %. the target fraction contained . × cd + cd ro+ cells and . × cd + cd ro+ cells per kg of recipient weight, with an increased cd +/cd + ratio compared with the starting product ( . versus . ). aliquots of cd ra+ depleted products containing × cd + cells/kg of recipient weight were prepared for fresh dli and cryopreservation. the patient received a fresh cd ra+ depleted dli of × cd + cells/kg from the original hsct donor at day + post hsct. at the time of dli (day + post hsct), peripheral blood cd + cd + and cd + cd + levels were < × /l, but reached × /l and × /l, respectively, at days following dli. at days after dli, all tlymphocyte subset counts were normalized (fig. ), corresponding to a continuously complete donor t-cell chimerism. the circulating t-cells were predominantly cd + cd ra+ ( × /l) and cd + cd ra+ ( × /l). the patient did not experience any acute gvhd after hsct or dli. the early post-transplant period was complicated at day + by acute mechanical ileus with unknown etiology. this was resolved by mechanical manipulation during the exploratory laparotomy. the blood tests showed continued vzv positivity and the numbers of cmv copies were increasing. his condition deteriorated further and he suffered an enterococcus faecium septicemia at day + , followed by severe diarrhea and gastrointestinal tract bleeding, signs of probable colitis. the stool analysis revealed cmv, in addition to rotavirus. two weeks after dli, the test results showed that the cmv was resistant to ganciclovir; therefore, the antiviral treatment was changed to foscarnet. following dli, the time to clearance of vzv and rotavirus was days. the cmv levels decreased rapidly (fig. ) and the patient's condition improved considerably. the stool analysis for cmv was negative at days following dli and cleared from peripheral blood at days. supplementation with immunoglobulins was discontinued on day + post hsct. he was discharged approximately months post hsct. no chronic gvhd was observed after the hsct or dli. he developed blindness and one-sided microphthalmia due to the varicella keratitis. at months after hsct, the patient had a normal lymphocyte count with normal lymphocyte subsets and neither new nor were exacerbations of preexisting infections observed. allogeneic hsct is the only curative treatment available for scid, although it is associated with a high incidence of severe infections and an increased risk of gvhd [ ] [ ] [ ] . early intervention is a crucial prognostic factor and the prompt availability of a parental donor may favor the hlahaploidentical donor alternative. haploidentical hsct protocols utilizing extensively ex vivo t-cell depleted grafts have proven efficient in preventing gvhd [ , ] . the remaining challenge of enhancing post-transplant immunereconstitution has been addressed by several investigators by utilizing partially t-cell depleted grafts [ , , [ ] [ ] [ ] or adoptive transfer of donor immune cells [ ] [ ] [ ] . here, we present a novel approach for treating scid, combining tcr-α/β-cell depleted haploidentical hsct with cd ra+ depleted dli for a therapeutic antiviral boost. our patient received a tcr-α/β-cell depleted graft from his haploidentical mother, following a reduced intensity conditioning regimen with a short course of atg, a single dose of rituximab to prevent ebv-ptld, and no additional gvhd prophylaxis. engraftment was rapid with complete donor chimerism and no signs of gvhd. however, at weeks post hsct, the patient was still t-cell lymphopenic with clinical symptoms of multiple severe viral infections. consequently, the decision was made to initiate therapeutic dlis for enhanced anti-viral immunity. in the haploidentical setting, unselected dlis with a tcell add-back of only × cd + cells/kg may cause severe gvhd [ , ] . in contrast, enriched cd + cd ro + memory t-cells may provide viral immunity with markedly reduced alloreactivity [ , ] . notwithstanding, cross-reactivity of memory viral-specific t-cells with mismatched hla has been described and is, thus, a potential source of gvhd in haploidentical hsct [ ] . based on these observations, the patient was treated with cd ra+ depleted haploidentical donor lymphocytes, transferring a total t-cell dose of no more than × cd + cells/kg with > . % purity of cd + cd ro+ memory t-cells. following dli, a prompt increase in cd + cd + and cd + cd + counts was observed, with a concomitant reduction in viral load and a subsequent clearance of viral infections. no acute or chronic gvhd was observed. as previously demonstrated [ ] , tcr-α/β cells could be efficiently depleted (log - . ) with the clinimacs instrument, preserving high numbers of cd + cells and potentially beneficial tcr-gd + cells and nk-cells in the graft. with the current approach of transferring a very low dose of cd ra+ depleted donor t-cells ( × /kg cd + cells), a sample of donor peripheral blood will provide sufficient numbers of lymphocytes. to ensure that the manufacturing of dlis was in compliance with good manufacturing process (gmp) conditions, a closed blood bag system was used for the collection and leukocyte enrichment of donor whole blood and the clinimacs instrument was used for subsequent cd ra+ depletion. naïve t-cells (cd ra+) were efficiently depleted (log - . ) providing sufficient numbers of cd + cd ro+ cells for the fresh dli and for cryopreservation. a benefit of the current protocol is the time and cost effectiveness of using a whole blood donation rather than a leukapheresis product as starting material. moreover, this approach is also less strenuous for the donor. a long-standing approach to control refractory viral infections following hsct is the generation of antigenspecific t-cells against single pathogens such as cmv, adenovirus, or ebv [ ] . new strategies have been developed for ex vivo selection of pathogen-specific donor t-cells, using antigen induced stimulation and isolation by cytokine secretion assay [ ] or hla-peptide streptamers [ ] . however, this approach is time consuming and technically much more demanding than the simple method of cell depletion based on surface cd ra+ antigen expression. also, the generation of antigen stimulated t-cells for clinical application is subjected to a more complex regulatory framework than that of cd ra+ depleted, unmanipulated lymphocytes. recently, the often limited therapeutic effect of using antigen-specific t-cells has been attributed to the limited in vivo persistence of transferred donor t-cells [ ] , suggesting that adoptive transfer should include a variety of donor t-cell populations to favor long-lasting clinical responses. very recently, three different groups [ , , ] investigated the use of cd ra+ depletion as part of the haploidentical or hla-mismatched donor graft processing. using different conditioning regimens with different stem cell sources and graft compositions, with or without serotherapy, two of the groups [ , ] reported limited or no gvhd despite high numbers of transferred cd + cd ro+ t-cells ( - x /kg) while the third group reported a . % incidence of grade iii/iv acute gvhd. further studies are required to determine the optimal timing, t-cell dose, and source of cd ra+ depleted donor lymphocytes -as part of the graft or as prophylactic, preemptive, or therapeutic dlis. automated depletion of cd ra+ naïve t-cells from unmobilized whole blood donations is a simple and rapid strategy to provide unmanipulated dlis, with a potentially broad repertoire of pathogen specific memory t-cells. in the haploidentical setting, therapeutic cd ra+ depleted dlis can be safely administered at low t-cell doses for efficient enhancement of viral immunity and limited risk of gvhd. we demonstrate the success of this approach following tcr-α/β-cell depleted hsct for the treatment of scid. educational paper. the expanding clinical and immunological spectrum of severe combined immunodeficiency neonatal diagnosis of severe combined immunodeficiency leads to significantly improved survival outcome: the case for newborn screening the natural history of children with severe combined immunodeficiency: baseline features of the first fifty patients of the primary immune deficiency treatment consortium prospective study transplantation outcomes for severe combined immunodeficiency how i treat severe combined immunodeficiency megadose transplantation of purified peripheral blood cd (+) progenitor cells from hla-mismatched parental donors in children haploidentical transplantation for acute lymphoblastic leukemia in childhood long-term outcome after haploidentical stem cell transplantation in children transplantation of cd /cd depleted allografts from haploidentical family donors in paediatric leukaemia hlahaploidentical stem cell transplantation after removal of alphabeta + t and b cells in children with nonmalignant disorders improved immune recovery after transplantation of tcralphabeta/ cd -depleted allografts from haploidentical donors in pediatric patients the who's who of t-cell differentiation: human memory t-cell subsets engineering human peripheral blood stem cell grafts that are depleted of naive t cells and retain functional pathogen-specific memory t cells depletion of naive t cells using clinical grade magnetic cd ra beads: a new approach for gvhd prophylaxis primary immunodeficiency in infection-prone children in southern sweden: occurrence, clinical characteristics and immunological findings rapid pulsed whole genome sequencing for comprehensive acute diagnostics of inborn errors of metabolism survival after t cell-depleted haploidentical stem cell transplantation is improved using the mother as donor ebmt/esid guidelines for haematopoietic stem cell transplantation for primary immunodeficiencies immune reconstitution and strategies for rebuilding the immune system after haploidentical stem cell transplantation high rabbit-antihuman thymocyte globulin levels are associated with low likelihood of graft-vs-host disease and high likelihood of posttransplant lymphoproliferative disorder haploidentical stem cell transplantation augmented by cd ra negative lymphocytes provides rapid engraftment and excellent tolerability rapid memory t-cell reconstitution recapitulating cd ra-depleted haploidentical transplant graft content in patients with hematologic malignancies cd ra depletion in hla-mismatched allogeneic hematopoietic stem cell transplantation for primary combined immunodeficiency: a preliminary study adoptive transfer of pp -specific t cells for the treatment of chemorefractory cytomegalovirus disease or reactivation after haploidentical and matched unrelated stem cell transplantation adoptive transfer and selective reconstitution of streptamer-selected cytomegalovirus-specific cd + t cells leads to virus clearance in patients after allogeneic peripheral blood stem cell transplantation hlahaploidentical transplantation with regulatory and conventional t-cell adoptive immunotherapy prevents acute leukemia relapse alloreactivity across hla barriers is mediated by both naive and antigen-experienced t cells effective tcr α/β+ cell depletion using the clinimacs® system to produce peripheral blood progenitor cell products for haploidentical transplantation immunotherapy for viral and fungal infections the design of this project did not need any special funding and the authors would like to thank the medical faculty of lund university and skåne university hospital for the opportunity to carry out this project. the authors would like to thank the patient and his family for their kind collaboration in this project, as well as their doctors. the authors declare that they have no competing interests.authors' contributions nb: conception and planning, collection and compiling of the clinical and laboratory data, and writing the manuscript. dt, jt, and lt: interpretation of the results and supervising the writing of the manuscript. jd: collection and compiling of the laboratory data and supervising the writing of the manuscript. all authors read and approved the final manuscript.• we accept pre-submission inquiries • our selector tool helps you to find the most relevant journal submit your next manuscript to biomed central and we will help you at every step: key: cord- -xez zso authors: stephens, r. scott title: icu complications of hematopoietic stem cell transplant, including graft vs host disease date: - - journal: evidence-based critical care doi: . / - - - - _ sha: doc_id: cord_uid: xez zso hematopoietic stem cell transplant (hsct) is an essential treatment modality for many malignant and non-malignant hematologic diseases. advances in hsct techniques have dramatically decreased peri-transplant morbidity and mortality, but it remains a high-risk procedure, and a significant number of patients will require critical care during the transplant process. complications of hsct are both infectious and non-infectious, and the intensivist must be familiar with common infections, the management of neutropenic sepsis and septic shock, the management of respiratory failure in the immunocompromised host, and a plethora of hsct-specific complications. survival from critical illness after hsct is improving, but the mortality rate remains unacceptably high. continued research and optimization of critical care provision in this population should continue to improve outcomes. non-infectious complications of hsct. respiratory failure is the most frequent cause of intensive care unit (icu) admission after hsct, most frequently from an infectious cause. the patient was placed on high-flow nasal cannula (hfnc) for oxygenation support and started on a norepinephrine infusion for hemodynamic support. antibacterial coverage was changed to meropenem, levofloxacin, and vancomycin, and voriconazole was added for antifungal coverage. her central line was removed. her respiratory status continued to decline, and endotracheal intubation and mechanical ventilation were required h after admission. she was placed on volume assist-control with a tidal volume of ml/kg predicted body weight, and bronchoscopy with bronchoalveolar lavage (bal) was performed. the bal fluid was initially bloody, but cleared with sequential aliquots. microbiologic studies of the bal fluid were positive for respiratory syncytial virus and pseudomonas aeruginosa. she was maintained on low tidal-volume ventilation and vasopressors were weaned off. her white blood cell count slowly began to recover, and her respiratory status began to improve. she was extubated on hospital day and was discharged from the icu on hospital day . hematopoietic stem cell transplant (hsct) has become an essential therapeutic modality in the treatment of malignant and non-malignant hematologic disease. in , more than , hscts were performed in the united states, including approximately , autologous hscts and more than allogeneic transplants [ ] . allogeneic transplants are associated with more morbidity and mortality than autologous transplants, and are further categorized based on conditioning regimen (myeloablative [ma] vs non-myeloablative [nma]), donor-recipient relation (related vs unrelated), hla matching (full match vs haploidentical vs mismatched), and stem cell source (bone marrow, peripheral blood, umbilical cord blood). in general, nma regimens are associated with less peri-transplant morbidity and mortality than fully ablative transplants. in both ma and nma transplants, the cytotoxic conditioning regimen required in hsct rapidly induces neutropenia by injuring hematopoietic precursor cells within the bone marrow [ , ] . neutropenia persists until donor cell engraftment or bone marrow recovery. the period of aplasia and neutropenia places the hsct patient at high risk for infectious complications. in addition to the lack of immune cells, the mucosal barrier of the intestinal tract is disrupted by chemotherapy, creating portals through which enteric pathogens can enter the bloodstream [ ] [ ] [ ] . the respiratory system is also more susceptible to infection, with qualitative and quantitative dysfunction of alveolar macrophages, lymphocytes, and neutrophils [ ] [ ] [ ] . even after the marrow and mucosal surfaces have recovered, the immunological consequences of hsct can cause further complications requiring critical care. refinement of transplant techniques over the last decades has dramatically decreased transplant-related mortality, but approximately % of hsct patients require critical care [ ] and earlier icu admission has been associated with improved survival rates [ , ] . still, icu mortality in allogeneic hsct patients remains approximately % [ ] . the early complications of hsct (day - ) are predominantly infectious in nature, and patients typically present to the icu with septic shock or respiratory failure. the latter is the most common reason for icu admission after hsct [ ] . non-infectious complications also occur, and can involve nearly any organ system. neutropenic fever, defined as any fever higher than . °c or a sustained fever greater than . °c for more than h with an absolute neutrophil count (anc) less than cells/ mm , occurs in more than % of patients undergoing hsct [ , ] . no organism is identified in about % of neutropenic fevers [ , ] . bacteremia is documented in up to % of patients. gram-positive bacteria are most commonly isolated [ , ] (table . ), while gram-negative infections confer a higher mortality risk [ ] . fungal infections, particularly candida and aspergillus species, are frequent, especially in prolonged or profound neutropenia [ ] . approximately % of allogeneic hsct patients will develop severe sepsis during the engraftment period [ ] , and mortality is approximately % in those who go on to develop septic shock [ , , ] . mortality predictors include concomitant graft-vshost disease (gvhd), respiratory failure, positive blood cultures, and multi-organ failure [ , ] . neutropenic fever and sepsis are medical emergencies, and appropriate empiric antibiotics must be started without delay: ideally within min of presentation [ , , ] and potentially within min [ ] . empiric antibiotics must cover common organisms and should be tailored to patientspecific culture data and institutional epidemiology [ , ] . appropriate empiric antibiotics include an anti-pseudomonal penicillin or cephalosporin (e.g. piperacillin/tazobactam or cefepime, respectively), or a carbapenem [ , ] . vancomycin is not routinely indicated but should be added in the presence of a suspected catheter-related infection, soft tissue infection, oral mucositis, pneumonia, known colonization with resistant gram-positive organisms, or hemodynamic instability [ , , ] . aminoglycosides should not be added to an anti-pseudomonal beta-lactam unless required by allergies, resistant organisms, or refractory hemodynamic instability [ , , [ ] [ ] [ ] . fluoroquinolones, which are frequently used as prophylaxis in hsct patients, should not be used as empiric monotherapy due to the likelihood of resistance. in hemodynamically unstable patients, anti-pseudomonal beta-lactams should be escalated to a carbapenem and consideration should be given to the addition of an aminoglycoside or aztreonam [ , , , ] . vancomycin should be added if not already part of the regimen, and anti-fungals with activity against yeasts and molds (e.g. liposomal amphotericin, caspofungin, or voriconazole) should be strongly considered in all unstable patients [ , , , ] . identification of infectious organisms and control of infectious sources are essential to optimize outcomes but the infectious workup should not delay antibiotic administration. blood cultures and respiratory cultures should be obtained and sinus, head, chest, and abdominal imaging performed as indicated [ , ] . abdominal pain or diarrhea associated with fever suggests neutropenic enterocolitis (typhlitis) which can lead to intestinal necrosis [ , ] . in the hemodynamically unstable patient with a central venous catheter, early catheter removal is associated with improved survival [ ] ; infected or potentially infected catheters should be removed without delay. acute respiratory failure and acute respiratory distress syndrome (ards) are major problems after hsct [ , ] . data from the s indicated that - % of patients undergoing hsct experience a respiratory complication [ ] . more recent data suggest that more than % of patients undergoing allogeneic transplant develop ards with a mortality rate of - % [ , ] . most cases of respiratory failure and ards after hsct are related to infection, either a primary pulmonary infection or sepsis [ ] . common pulmonary infections and associated risk factors are shown in table . hsct [ , ] , and is associated with significant mortality, especially with progression to lower respiratory tract infection [ ] [ ] [ ] . in some cases, antiviral therapy with agents such as oseltamivir (influenza) or ribavirin (respiratory syncytial virus) is indicated [ ] . bacterial pneumonias are also common and may occur as a co-infection or secondary infection with a respiratory virus. fungal and other opportunistic infections such as pneumocystis jirovecii must also be considered [ ] . as in immunocompetent patients, treatment of ards centers on treatment of the underlying cause while providing supportive care with low tidal volume mechanical ventilation. neuromuscular blockade and prone positioning should be considered in patients with an arterial po : fio ≤ mmhg [ ] [ ] [ ] . non-invasive ventilation (niv) is frequently used as firstline respiratory support in hsct patients [ ] . however, early studies which showed a mortality benefit in immunosuppressed patients with using niv compared to invasive mechanical ventilation were limited by relatively few numbers of hsct patients and extremely high mortality in the control groups [ , ] . it is nearly impossible to control delivered tidal volume with niv and high delivered noninvasive tidal volumes are linked to higher rates of niv failure [ ] . more recent data suggest that niv may not be beneficial in hsct patients and heated humidified high-flow oxygen may be a better option [ , [ ] [ ] [ ] [ ] [ ] . chest computed tomography (ct) scanning should be performed in all patients with respiratory symptoms [ ] . the presence of respiratory failure, respiratory symptoms, or abnormalities on chest imaging should prompt evaluation for a respiratory infection. in many cases a non-invasive evaluation is appropriate, but bronchoscopy may be indicated in some patients [ , , ] , and bronchoscopic findings that lead to a change in management are associated with improved outcomes [ ] . two specific forms of respiratory failure after hsct warrant special mention: diffuse alveolar hemorrhage and idiopathic pneumonia syndrome. diffuse alveolar hemorrhage (dah) occurs in up to % of patients and is associate with poor outcomes [ , ] . diagnosis is most commonly made by observation of progressively bloody aliquots of bronchoalveolar lavage. steroids are the mainstay of treatment of dah, with some evidence that efficacy is greatest at doses < mg/day of methylprednisolone equivalent [ ] . idiopathic pneumonia syndrome (ips) is a form of noninfectious lung injury after hsct and is clinically defined by diffuse alveolar injury when infection, cardiac dysfunction, renal failure, and volume overload have been excluded [ ] . ips can have many manifestations, including ards, pulmonary capillary leak, dah, or cryptogenic organizing pneumonia. ips is thought to affect up to % of patients after myeloablative allogeneic hsct, and only ~ % of patients after non-myeloablative hsct. median time of onset of ips [ , ] is days after hsct and mortality ranges from - % in all patients, with nearly % mortality if mechanical ventilation is required [ ] . though the pathophysiology of ips is incompletely understood, research indicating a pathogenic role for tnf-α has led to the use of the anti-tnf-α antibody etanercept to treat ips, with mixed clinical results [ ] [ ] [ ] [ ] . the single randomized placebo-controlled trial in adults included only patients and showed no benefit to etanercept when added to steroids (methylprednisolone mg/kg/ day) [ ] . neurologic complications are frequently encountered after hsct [ ] . intracerebral hemorrhages are a constant threat in thrombocytopenic patients. infections of the central nervous system (cns), including viral, bacterial, and fungal, can occur, and may require modification of antibiotic regimens to ensure cns penetration. seizures and generalized encephalopathy can occur, often with cryptic causes. posterior reversible encephalopathy syndrome (pres) is increasingly recognized, especially in patients receiving tacrolimus-based gvhd prophylaxis. any of these complications may be life-threatening, and close collaboration with neurology and neurocritical care specialists may be required. acute kidney injury (aki) is common after hsct and affects up to % of patients, with higher incidence after allogeneic transplant than autologous transplant [ , ] . in addition to the usual icu causes of aki such as septic shock, there are many specific contributors to the risk of aki in hsct, including preparative chemotherapeutic regimens, nephrotoxins (e.g. tacrolimus, cyclosporine, antimicrobials), elevated cytokine levels, gvhd, and hepatic sinusoidal obstruction [ ] . hemorrhagic cystitis arising from chemotherapy toxicity or viral infection can cause significant blood loss and obstructive nephropathy due to blood clots. management of aki primarily consists of limiting exposure to nephrotoxins (if able) and maintaining adequate hemodynamics. if hemorrhagic cystitis is present, continuous bladder irrigation with a three-way catheter should be considered. the requirement for renal replacement therapy is ominous and portends a high mortality rate [ ] . hepatic veno-occlusive disease (vod), also known as sinusoidal obstruction syndrome, primarily occurs after myeloablative hsct, but can occur after a non-myeloablative transplant [ ] . vod is thought to be caused by damage to the hepatic endothelium and leads to obliteration of hepatic sinusoids and hepatocyte necrosis. incidence of vod is thought to be approximately %, though estimates vary. diagnosis is based on clinical findings (table) including hepatomegaly, elevated bilirubin ascites, and weight gain [ ] . there are limited therapeutic options for vod, and the prognosis is poor. a major complication of allogeneic hsct is graft-versushost disease (gvhd), which is divided into acute and chronic forms. acute gvhd is a major contributor to peritransplant morbidity and mortality, and is caused by donororigin t-cells recognizing recipient tissues as foreign and instigating an immune response against the transplant recipient [ ] . acute gvhd generally occurs within the first days after transplant and can affect the skin, mucosa, intestinal tract, and liver. grading is based on severity of clinical manifestations, which include skin erythema or maculopapular rash; nausea, emesis, or diarrhea, and elevated bilirubin levels ( fig. . , table . ) [ ] [ ] [ ] [ ] . acute gvhd can progress to frank epidermal desquamation, massive diarrhea and hematochezia, and fulminant liver failure, respectively. severe skin acute gvhd behaves much like a burn injury, and the expertise of a burn center may be required. corticosteroids are the mainstay of treatment for acute gvhd, and the prognosis of steroid-refractory disease is poor [ , ] . prophylaxis against gvhd is an essential part of allogeneic transplant regimens and includes a variety of modalities, including calcinuerin inhibitors, anti-metabolites, and post-transplant cyclophosphamide [ ] . as gvhd prophylaxis has improved, more patients are presenting with grade ii or grade iii acute gvhd (and fewer are presenting with grade iv acute gvhd), the incidence of hepatic acute gvhd is decreasing, and overall mortality from acute gvhd is decreasing in patients treated with tacrolimusbased gvhd prophylaxis [ ] . chronic gvhd is the major cause of non-relapse-related mortality after hsct [ ] . despite its name, chronic gvhd is defined clinically, rather than by time after transplant, and can present at any time during the transplant course. by years after transplant, up to - % of patients will have some manifestation of chronic gvhd [ ] [ ] [ ] . though the biology of chronic gvhd is complex and incompletely understood, clinically it mimics autoimmune disease [ ] . while diagnostic criteria include effects on the skin, oral mucosa, eyes, liver, gi tract, joints, genitals, and lungs, chronic gvhd can affect almost any organ system and is staged according to severity of organ involvement [ ] . selected manifestations and diagnostic criteria are in table . . of the manifestations of chronic gvhd, the most relevant to the icu physician is pulmonary chronic gvhd. the only recognized manifestation of chronic pulmonary gvhd is bronchiolitis obliterans syndrome (bos), which is diagnosed by documentation of the new onset of an obstructive ventilator defect (fev : fvc < . and fev < % predicted) and air trapping (documented by expiratory ct scan or pulmonary function tests) in the absence of an explanatory pulmonary infection [ ] . bos results from a b c d peribronchiolar fibrosis and obliteration of small airways resulting in the characteristic obstructive physiology [ ] . interstitial and subpleural fibrosis may also occur, resulting in concomitant restrictive physiology. bos occurs in approximately - % of all patients after allogeneic hsct, and % of those with chronic gvhd, but is likely underdiagnosed [ , , ] . inhaled corticosteroids appear efficacious in improving fev in established bos [ ] . systemic steroids are also commonly used to treat bos and most patients are maintained on anti-gvhd immunosuppression with tacrolimus, sirolimus, or a calcineurin inhibitor [ ] . the combination of inhaled fluticasone, azithromycin, and montelukast (fam) appears to slow the decline in lung function with bos [ ] but has not yet been proven in a randomized controlled trial. while fam is standard therapy for established bos, recent data argue strongly against using azithromycin as prophylaxis against bos due to decreased survival due to a higher rate of hematologic relapse [ ] . though fam has been shown to decrease the progression of bos, mortality due to progressive lung disease remains high, and patients typically present to the icu with respiratory failure. unfortunately, with end-stage bos, there are no effective therapeutic options. a select few patients may be eligible for consideration for lung transplantation, but this is unusual, hsct is increasing in volume and importance as a therapeutic modality, and the volumes of hsct patients requiring critical care is accordingly continuing to increase. there is good reason to think that the pathogenesis of critical illness is substantially different in the immunosuppressed hsct patient. yet our understanding of critical illness in this population is limited, and many practices are extrapolated from the general critical care population without direct evidence in the hsct population. in response to this, research agendas for critically ill hematology and oncology patients have been proposed [ ] . neutropenic sepsis is typically thought of as an uncontrolled variant of non-neutropenic sepsis. however, the real picture is likely much more complicated, and neutropenic sepsis and respiratory failure may be very different from their nonneutropenic counterparts. even the phrase "neutropenic sepsis" is a misnomer, as the hsct myelopreparative regimens also result in pancytopenia. leukopenia, including neutropenia, lymphopenia, and monocytopenia, dramatically changes not only the acute response to infection, but the regulation of the adaptive immune response and the resolution and repair of injury [ , , , [ ] [ ] [ ] [ ] [ ] [ ] [ ] . platelets are increasingly recognized to play a vital role in the defense against bacterial, viral, and fungal infections, and like leukocytes, are integral to the development and resolution of organ failure [ ] [ ] [ ] [ ] [ ] . not surprisingly, thrombocytopenia is associated with poor outcomes in critical illness [ ] . taken together, these data strongly support the notion that common critical care conditions, such as sepsis and respiratory failure, may differ dramatically in hsct patients compared to "normal" patients. encouragingly, survival in neutropenic sepsis appears to be improving, but still lags that of non-neutropenic patients [ , , ] , and more research is needed. drug-resistant and multi-drug resistant (mdr) organisms are an increasing problem in hsct patients, and the situation shows no sign of improving [ , , ] . vancomycinresistant enterococcus (vre) bacteremia affects up to % of patients after hsct and is associated with poor outcomes [ ] [ ] [ ] . similarly, mdr gram negative infections, particularly carbapenem-resistant enterobacteriaceae (cre), are associated with high mortality rates in allogeneic hsct patients [ ] . successful treatment cre infections is challenging, and requires early use of multi-drug antibiotic regimens, typically including aminoglycosides, carbepenems, and polymyxins. however, none of the available regimens are particularly effective, and new antimicrobials are desperately needed. an increasingly recognized complication of hsct is hsctassociated thrombotic microangiopathy (hsct-tma) [ ] , which has some features in common with betterknown microangiopathic processes such as thrombotic patients can present to the icu with acute kidney injury and neurologic changes in addition to hemolytic anemia and thrombocytopenia [ , ] . management is predominantly supportive, with blood pressure control, cessation of any possible pharmacologic instigators (tacrolimus or cyclosporine), and renal replacement therapy playing major roles. recent case reports have suggested a possible role in some patients for the anti-cd antibody rituximab or the anti-complement antibody eculizumab, though neither of these agents has been definitively proven effective [ ] . as noted above, advances in transplant techniques have allowed the increased use of alternative donors, including related haploidentical donors [ ] [ ] [ ] . similarly, peripheral blood stem cells (pbsc) are increasingly used for transplant instead of bone marrow stem cells [ ] . however, the use of peripheral blood results in a larger number of donor t-cells included in the transplanted stem cells. this higher t-cell dose can result in a profound syndrome of fevers, vascular permeability, hemodynamic instability, acute kidney injury, and respiratory failure. this constellation of findings is associated with elevated levels of inflammatory cytokines and has accordingly been labeled as cytokine release syndrome (crs). while most associated with chimeric antigen receptor (car) t-cell therapy [ ] , crs is increasing recognized after pbsc transplant and is associated with poor outcomes [ ] . emerging data suggest that anti-il- therapy with tocilizumab may improve outcomes, but more research is needed [ ] . hsct continues to grow as a therapeutic modality and the pool of both potential donors and recipients continues to increase. as hsct volumes increase and the complexity and potential toxicity of hsct regimens expands, the number of critically ill hsct patients will increase [ ] . in response to this growing icu volume, some centers continue to admit hsct patients to general icus, but many high-volume transplant centers have developed specialty hsct icus. while specialty hsct icus have a number of potential benefits, there are few data to support (or dissuade) their development. development of best practices for the provision of critical care to hsct patients, including the 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critically ill adult chimeric antigen receptor-t cell therapy patient: a critical care perspective severe cytokine-release syndrome after t cell-replete peripheral blood haploidentical donor transplantation is associated with poor survival and anti-il- therapy is safe and well tolerated key: cord- -jr mbh s authors: calore, elisabetta; marson, piero; pillon, marta; tumino, manuela; tison, tiziana; mainardi, chiara; de silvestro, giustina; rossin, sara; franceschetto, genny; carraro, elisa; pescarin, matilde; varotto, stefania; destro, roberta; gazzola, maria vittoria; basso, giuseppe; messina, chiara title: treatment of acute graft-versus-host disease in childhood with extracorporeal photochemotherapy/photopheresis: the padova experience date: - - journal: biol blood marrow transplant doi: . /j.bbmt. . . sha: doc_id: cord_uid: jr mbh s acute graft-versus-host disease (agvhd) is the major cause of morbidity and mortality after allogeneic hematopoietic stem cell transplantation. systemic steroid treatment represents the first-line therapy for agvhd and is associated with a response rate of % to %. steroid-resistant patients have a poor prognosis with high transplantation-related mortality (trm). several second-line therapies have been proposed for the management of unresponsive agvhd, without proven beneficial effects on patients' outcome or overall long-term survival. for these reasons, extracorporeal photochemotherapy/photopheresis (ecp), a cell-based approach to control gvhd that spares generalized immunosuppression, seems to be promising. in this study, we report the outcome of consecutive pediatric patients treated with ecp between and for agvhd. among them, patients had steroid-resistant agvhd, had steroid-dependent agvhd, and did not receive steroid as first-line therapy because of clinical contraindications. a complete response was obtained in % of patients, a partial response was observed in %, and there was no response in % of patients. at day + , trm was % in the whole cohort; trm was % and % among responders and nonresponders to ecp, respectively (p < . ). the -year overall survival was %, showing a difference between responders and nonresponders of % and %, respectively (p = . ). the -year time to progression of primary disease was %, without any significant difference between the groups. moreover, the -year progression-free survival of primary disease was %, with a significant difference (p = . ) between responders ( %) and nonresponders ( %) to ecp. in conclusion, this study demonstrates that ecp is highly effective in agvhd without a negative impact on primary disease. allogeneic (allo) hematopoietic stem cell transplantation (hsct) is increasingly used as a therapeutic approach for hematological and nonhematological diseases. in the last decade, improvements in infection monitoring and prophylaxis, immunosuppressive (is) strategies, high-resolution hla typing, and supportive care measures led to better outcomes after this procedure. despite these advancements, acute graft-versus-host disease (agvhd) remains the major cause of morbidity and mortality after allo-hsct [ , ] . to standardize diagnosis and management of agvhd, a british guideline was published by a joint working group of the british committee for standards in haematology and the british society for bone marrow transplantation. this document included recommendations for diagnosis and management of agvhd as well as primary treatment options for patients with steroid-refractory (sr) disease [ ] . standard management of agvhd included steroids at different doses depending on agvhd grade. if no improvement of agvhd after days was noted or progression within hours were observed, then the addition of secondline agents should be considered. second-line options are mycophenolate mofetil (mmf), anti-tnf antibodies, mammalian target of rapamycin inhibitors, il- receptor antibodies, and extracorporeal photochemotherapy/photopheresis (ecp) [ ] . unfortunately, despite multiple clinical trials, no single agent improving overall survival (os) for patients who failed steroid treatment has been identified [ , ] . moreover, the current survival at years in patients who respond to steroids is about % versus % in nonresponders (nr) [ ] and it has been shown that transplantation-related mortality (trm) is higher in steroidresistant patients [ , ] . in our study, we focused on ecp, one of the most promising treatments for agvhd. briefly, ecp consists of procedures: collection of peripheral leukocytes cells, irradiation of cells by ultraviolet a light in presence of methoxypsoralen , and reinfusion of treated cells in the patient. the underlying mechanism of action of ecp in gvhd is not completely understood [ , ] . within hours, this process induces apoptosis of all treated cells (including t cells) and subsequent phagocytosis by antigen-presenting cells; as a result, this might regulate immune homoeostasis, modulate the cytokine production, and induce tolerance of antigen-presenting cells [ ] [ ] [ ] [ ] . ecp has been demonstrated to be effective in treating both steroid-resistant and steroid-dependent patients with agvhd [ ] [ ] [ ] [ ] . in the pediatric setting in particular, the reported response rate ranges from % to %, according to the organs involved. in agvhd steroid-resistant patients, - year os is significantly increased in complete responders to ecp, % compared with % for nr [ ] . in , the italian society of haemapheresis and cell manipulation and the italian group of bone marrow transplantation elaborated best practice recommendations for ecp, which reflected the common clinical practice in most italian transplantation centers where ecp is performed with a total of procedures per year [ ] . despite this large use of ecp, most of the published reports deal with retrospective data that are difficult to compare, as patients' selection criteria, treatment schedules, patients' monitoring, and patients' assessment protocols differ among institutions. moreover, no randomized studies have been conducted in patients with agvhd. here, we report our single-center experience on ecp treatment in pediatric patients with agvhd. the response to ecp, trm, os, progression-free survival (pfs) of primary disease, and time to progression (ttp) of primary disease of patients treated with ecp were analyzed. from january to june , consecutive pediatric patients ( males, females) affected by agvhd were treated with ecp at the hsct unit of university hospital of padova. fifteen of these patients have been previously reported [ ] . the clinical characteristics of patients are shown in table . the median age at ecp was . years (range, . to . years) and the median body weight was kg (range, to kg). fifteen children weighted less than kg. the last follow-up was fixed on june . in detail, patients were treated with ecp for agvhd refractory to steroids, which was defined as a progression or no improvement in agvhd after at least days or days on methylprednisolone (mp) ! mg/kg body weight, respectively (sr group); patients for steroid-dependant agvhd, defined as a flare-up of agvhd during the tapering of mp (sd group); and children with agvhd who required a reduction of pharmacological is or contraindications to is therapy for viral reactivations, systemic mycoses, or intolerable side effects (group with infectious complications [ic]). in particular, of patients in the ic group (ic-a group) underwent ecp without steroids as a first-line therapy because of contraindications: for tcr ab and cd edepleted haploidentical transplantation with probable invasive pulmonary aspergillosis (ipa) and adenovirus (adv), for proven ipa, for concomitant proved ipa and cytomegalovirus (cmv) reactivation, for probable ipa and cmv and bk virus (bkv) reactivations, for proven ipa and multiple viral reactivations, including adv, cmv, and epstein-barr virus (ebv), for cmv reactivation, for cmv and ebv reactivations, and for cmv, ebv, adv, and bkv reactivations. the other patients (ic-b group) had sd agvhd and cyclosporin a (csa)erelated renal insufficiency in only patient; sd agvhd and concomitant infections in the remaining patients: cmv reactivations, ; cmv and ebv reactivations, ; cmv and bkv reactivations, ; cmv, adv, and bkv reactivations, ; ebv reactivations, ; ebv and adv reactivations, ; ebv reactivations and probable aspergillosis, ; hepatitis b virus, ; hepatitis b virus and ebv reactivations, ; cmv, ebv, and bkv reactivations, ; adv, human herpesvirus- , bkv, and coronavirus, ; hepatitis c virus, cmv, and proven ipa, n ¼ . in our practice, surveillance for viral and fungal infections in blood is routinely performed during the first days after hsct in all patients and includes ebv-dna, cmv-dna, adv-dna, human herpesvirus- dna, bkv-dna, jc-dna, and galactomannan antigen search. this schedule is performed once each week in allo-hsct recipients from hla-identical sibling and twice each week in allo-hsct recipients from unrelated or haploidentical donors. monitoring viral infections in urine comprises cmv-dna, bkv-dna, jcv-dna in a weekly search. blood, urine, and stool cultures; nasal and throat swabs; and nasopharyngeal aspirates were weekly performed. search for other viruses or microbiological agents was performed upon clinical symptoms. viral reactivations were detected by pcr positivity for ebv-dna (cut-off: copies/ml), cmv-dna (cut-off: copies/ ml), and adv-dna in qualitative test. clinical systemic fungal infections were defined proved or probable according to european organization for research and treatment of cancer criteria [ ] . the cut-off for the galactomannan index was set at . (enzyme immuno assay (e.i.a.) method). the algorithm for agvhd treatment used in our center is shown in figure . csa was administered for and months in children who received hsct from an hla-identical sibling or unrelated donor, respectively. in unrelated hsct, short-term methotrexate and rabbit antithymocyte globulin (atg) were given. in unrelated cord blood hsct, prophylaxis included csa and atg. in haploidentical setting, ex vivo elimination of ab t cells and cd b cells was done and atg was administered before the cells were infused; no other is therapy was given after hsct. informed consent was obtained from patients' parents, as well as from the patients themselves when possible, and the use of ecp was approved by the ethical committee of the hospital of padova. the clinical organ involvement was graded and then combined to obtain an overall grade, according to glucksberg criteria for agvhd [ ] . histological confirmation was obtained whenever clinically indicated to confirm gvhd diagnosis. eligibility criteria for ecp treatment were as follows: children with sr agvhd (n ¼ ); children with sd agvhd (n ¼ ); patients with agvhd in whom is therapy was contraindicated or who required a rapid decrease of is therapy for increasing ebv viral load, cmv reactivation in subsequent samples, systemic fungal infections, intolerable side effects (n ¼ ). all children must present in complete hematological remission and full donor chimerism; white blood cell (wbc) count >  /l, and no concomitant treatment with either atg or monoclonal antibodies. ecp was performed using different techniques: "in-line" treatment (uvar photopheresis instrument, therakos, exton, pa) was used in of patients and the "off-line" technique (cobe spectra, bct terumo, lakewood, co) was used in of children. technical descriptions have already been published [ ] . the "off-line" technique was introduced in to treat lowweight children. for patients weighing less than kg, priming of the leukapheresis circuit with irradiated and leuko-reduced red blood cells (regardless of baseline hemoglobin level) was performed, as recommended in the italian society of haemapheresis and cell manipulation-italian group of bone marrow transplantation indications [ , ] . pre-ecp hemoglobin levels were maintained between g/dl and g/dl. the cell product was treated with -mop and diluted to a final concentration of mg/ ml to mg/ ml, according to specific procedures (in-line technique, mg/ ml; off-line technique, mg/ ml). in all patients, a to french hickman double-lumen central catheter was systematically used for the procedure. to provide adequate flow rates, ie, to ml/kg/minute, anticoagulation with urokinase , u for hours as lock-therapy was performed on the day of the procedure. the leukapheresis product contained a median of wbc of .  /ul (range, . to .  ), a median of mononuclear cells (mnc) of . % (range, % to %). the median number of wbc reinfused to the patients was  (range, to ,  ), whereas the median number of mnc reinfused to the patients was  (range, . to .  ). patients were treated with ecp twice each week for the first month, every weeks during the second and third months, and then monthly for at least more months, for a total of procedures. progressive tapering and discontinuation of ecp were decided upon evaluation of individual response. any concomitant is therapy was initially maintained, then modified or discontinued according to the clinical response. criteria for defining response to ecp were previously reported [ ] . all patients enrolled for ecp before day þ were included in this group and response to ecp was evaluated at day þ , day þ , and at the end of ecp treatment. complete response (cr) was defined as the resolution of all signs of agvhd and partial response (pr) as at least a % improvement in the clinical signs. in the latter case, given the complexity of assessing response, we defined pr for each organ as follows: for the skin, at least a % reduction in the body surface area affected; for the gi tract and liver a % reduction in the volume of diarrhea or value of bilirubin. any worsening of organ involvement, as well as the appearance of new signs or symptoms of gvhd, was defined as progressive disease (pd). patients with stable or pd were considered nr. seventy-two consecutive patients with agvhd received ecp at a median time of days (range, to ) after hsct and days (range, to ) from the diagnosis of agvhd. sixty-four patients had skin involvement (grade iv, n ¼ ; grade iii, n ¼ ; grade ii, n ¼ ; grade i, n ¼ ). fifty-five patients had gastrointestinal (gi) agvhd (grade iv, n ¼ ; grade iii, n ¼ ; grade ii, n ¼ ; grade i, n ¼ ). twelve patients had liver involvement (grade iii, n ¼ ; grade ii, n ¼ ; grade i, n ¼ ). regarding the number of organs involved: in patients skin was affected and presented gi involvement, whereas patients had combined skin and gi agvhd, patient had combined gi and liver agvhd, and patients had combined skin, gi, and liver agvhd. ic-a group is those with infectious complications and no steroid before ecp; the ic-b group is those with infectious complications and steroid before ecp. * hla match considered / . the overall clinical grading of agvhd was as follows: grade i, n ¼ ; grade ii, n ¼ ; grade iii, n ¼ ; and grade iv, n ¼ ; details of different grades in the patients' subgroups can be found in table clinical evaluation of the patients was conducted at every ecp procedure. sixty-three of patients with agvhd grades i to iv received mg/kg/ day of mp as first-line therapy. the median dose of steroid at the beginning of ecp was mg/kg/day. in detail, the is therapies before ecp were csa, n ¼ ; csa plus steroid ( mg/kg), n ¼ ; tacrolimus plus steroid ( mg/kg), n ¼ ; and csa or tacrolimus plus mmf plus steroid, n ¼ . ecp was used as first-line therapy in of patients, as second line therapy in of patients (among them, haploidentical hsct was treated only with csa), as third-line in of patients, and in of patients as fourth-line therapy. patients' characteristics were compared using the chi-squared or fisher's exact test (as appropriate) in case of discrete variables, or the mann-whitney test in case of continuous variables. trm was calculated from the date of hsct to day þ and to the last follow-up, considering as event any nonrelapse cause of death. os was calculated from the date of hsct to the date of death from any cause, or to the last follow-up. pfs was calculated from the date of hsct to the date of relapse of underlying primary disease or death for any cause or to the last follow-up. ttp was calculated from the date of hsct to the date of relapse of primary disease or to the last follow up. cumulative incidences (ci) of relapse of underlying disease were estimated in the competing risk model, considering death from any cause or cgvhd as the competing events. survival analysis was performed using kaplan-meier method with % confidence interval. standard error (se) for each survival and incidence rate is given. differences between groups were compared using the log-rank test and the gray's test. all reported p values were -sided, and statistical significance was set at a ¼ . (sas institute, cary, nc; release . ) [ ] . response to ecp treatment, evaluated according to the overall grading of agvhd and to the organ involvement at day þ , day þ , and at the end of ecp, is summarized in table . at the end of treatment with ecp, of ( %) patients had a cr, of ( %) had a pr, and of ( %) were nr. among the patients showing a cr, patients had agvhd grade i, patients had grade ii, had grade iii, and had grade iv. in particular, the cr rate for patients with agvhd grades i and ii and grades iii and iv were % and %, respectively (p ¼ . ), whereas the pr rate for patients with agvhd grades i and ii and grades ii to iv were % and %, respectively (p ¼ . ). no significant statistical difference in cr rate was observed according to the subgroups analyzed (sr, %; sd, %; ic groups, %) (p ¼ . ). at ecp discontinuation, cr of agvhd manifestations of skin, gut, and liver was observed in %, %, and % of patients, respectively. maximal response to ecp was observed after weeks of treatment ( procedures). as a result of ecp, at the end of treatment, it was possible to discontinue is therapy in patients ( %) and reduce it in patients ( %), of them who received allo-hsct from an unrelated donor. regarding the steroid tapering, in patients treated with mg/kg/day before ecp, the steroid dose was reduced by % after month of ecp treatment, % after months, and % after months of ecp treatment. the median lansky/karnofsky performance score improved from % before ecp to % after completing the treatment. no association was found between responders and nr to ecp and the major clinical risk factors affecting agvhd (table ) . cgvhd twenty-three of patients ( %) presented clinic manifestations of cgvhd (table ). in detail, patients ( %) had progressive cgvhd ( nr and pr to ecp) and patients ( %) had quiescent cgvhd onset after a median of months (range, days to months) from the end of ecp. overall grading of cgvhd, based on the national institutes of health consensus [ ] , was mild for patients, moderate for patients, and severe for patients. among the patients with progressive cgvhd, ecp was used with other is therapies in of patients, obtaining cr in only of them. overall, of patients were alive at the last follow-up: of had no cgvhd and discontinued is therapy, whereas only patient presenting with cgvhd was still in treatment. all the patients with quiescent cgvhd were alive at the last follow-up: patients were free from cgvhd and without is therapy and the other patients had cgvhd and were still on treatment with is therapy plus ecp. no association was found between responders and nr to ecp and the onset of quiescent cgvhd. at day þ , the overall trm was % (se, %). trm was % (se, %) and % (se, %) for responders and nr to ecp, respectively (p < . ). at last follow-up, the overall trm was % (se, %), whereas trm stratified between responders and nr was % (se, %) and % (se, %), respectively (p < . ) (figure a,b) . the -year os was % (se, %) with a statistically significant difference between responders and nr ( %; se, % versus %; se, %, respectively; p ¼ . ) ( figure a,b) . the -year pfs of primary disease for all the group was % (se, %), with a significant difference (p ¼ . ) between responders ( %; se, %) and nr ( %; se, %) ( figure a,b) . overall, the -year ttp of primary disease was % (se, %), without any significant difference between the groups (responders: %; se % versus nr: %; se, %; p ¼ . ) ( figure a,b) . we compared patients' survival rates on ecp treatment used as first, second, or third/fourth-line therapy. no difference was observed at -years between responders and nr in term of os (p ¼ . ), pfs (p ¼ . ), and ttp (p ¼ . ). the overall -year ci of relapse of the underlying disease was % (se, %); in particular, it was % (se, %) and % (se, %) for responders and nr to ecp, respectively ( figure a,b) . overall, at the last follow-up (median time from hsct of years; range, . to . years), patients were alive ( %); of them ( %) were without gvhd and without any is therapy. twenty-one patients ( %) died: from relapse of primary disease and from nrm. among this last group, patient with agvhd died at day þ from hsct because of sepsis; patients with cgvhd died from cmv pneumonia ( case), acute hepatitis from hcv infection ( case), encephalopathy ( case), and multiorgan failure ( cases); and patient died from cmv pneumonia at day þ from hsct, without evidence of cgvhd. side effects observed during ecp were generally mild and more frequent in low-weight children. ecp caused mild hypotension in patients associated with abdominal pain in all cases ( episodes out of apheresis sessions). these adverse effects did warrant suspending the procedure. a transient reduction in hemoglobin, platelet, and/or wbc count during the ecp treatment, likely independent from the post-transplantation course and putatively ecp-related, was observed in , , and patients, respectively. one patient with grade iv agvhd on high-dose steroid therapy ( mg/kg/ day) experienced acute gi bleeding after the second course of ecp: gi endoscopy showed multiple ulcers in the stomach. a girl with pre-existing cardiac impairment showed acute heart failure for fluid overload after the procedure that quickly responded to adequate therapy. one girl, after ecp procedures, had anaphylaxis (cough, vomiting, abdominal pain, hypotension, and palpebral edema) a few minutes after the end of -mop irradiated bag infusion. she responded to antihistamine and steroid therapy, but ecp treatment was then stopped. the aim of this retrospective study was to analyze the role of ecp for the treatment of agvhd. the efficacy of ecp is well established for treatment of cgvhd [ , ] , whereas in agvhd, no prospective randomized studies have been published. however, the use of ecp is suggested as second-line therapy, together with mammalian target of rapamycin inhibitors, mmf, il- receptor antibodies, and anti-tnf antibodies [ ] . the largest phase prospective study exploring feasibility and efficacy of ecp in treatment of agvhd in adults, involving patients, was performed by greinix et al. [ ] and reported a cr rates of % for skin and % for liver and gi agvhd. so far, data on pediatric patients treated with ecp for agvhd have been reported, showing an overall cr rate ranging from % to % and a survival rate ranging from % to % [ , , [ ] [ ] [ ] [ ] [ ] [ ] [ ] . to date, our is the largest pediatric case series treated in a single center. in our sample size, we found a higher overall response rate to ecp compared with a multicenter retrospective study of the italian association for pediatric hematology/oncology (aieop) ( % versus %) [ ] . we attempted to determine the factors that may have influenced our observed higher response. in the last years, many changes have been introduced in hsct, such as highresolution hla typing, new agents in the conditioning regimen, more use of atg, monoclonal antibodies, and new antifungal drugs. it is difficult to determine which modification may have influenced the outcome. we could also hypothesize that specific expertise in pediatric hsct and earlier treatment with ecp ( days in our series versus days in aeiop study) may have improved the overall outcome. further studies are needed to address this topic. no association was found between responders and nr setting to ecp and major risk factors for agvhd. in addition, in our series, no difference was found according to the grade of gvhd (grade i and ii, %; versus grade iii and iv, %; p ¼ . ) and to the subgroups of patients analyzed (sr, %; sd, %; ic, %; p ¼ . ). our results showed better response rate than those reported in literature for advanced stages of disease, where higher grades and poorer response to is therapy correlate with a worst outcome [ ] . nevertheless, higher cr rates were observed in grade ii gvhd, suggesting that an early start of ecp sessions may be beneficial, even if in our study the timing to start ecp (< versus > days) did not influence the response. in our group, ecp seemed to be effective in all the involved organs. as previously reported, our results support that ecp is a steroid-sparing treatment; in fact, the steroid dose was reduced by %, %, and % after , , and months, respectively from the onset of ecp. we performed ecp as front-line therapy in patients with fungal infection and viral reactivation and agvhd, with complete response in of them. to our knowledge, this is the first report of ecp as first-line treatment. is therapy was either discontinued or reduced in % of responding patients. it is well known from the literature that is therapy increases the risk of infectious complications and relapse of underlying disease after allo-hsct [ , , , , , ] . in children, who may be particularly vulnerable to the consequence of gvhd itself and prolonged treatment with is agents, the use of ecp is particularly appealing. the efficacy of ecp in controlling gvhd did not affect the preservation of graft-versus-leukemia effect; in fact, the low incidence of relapse of underlying disease was recorded by us and others [ , , , ] . many concerns has been raised related to the technical aspects of apheresis in the pediatric setting. in children with low body weight, the caregivers should carefully monitor patients for signs and symptoms of hypovolemia. in our experience, ecp was well tolerated, with few and mild adverse effects, the most frequent of which were mild hypotension, abdominal pain, and headache. curiously, these symptoms were recorded more often during ecp compared with other apheretic procedures [ , ] . the majority of side effects were observed in the earliest period in which ecp was performed in our center. all these observations support the idea that there has been a learning process for the management of technical elements and side effects. in our experience, ecp was feasible even in very young children with low body weight (< kg). technically, we performed priming of the circuit with irradiated and leukodepleted red blood cells (regardless of baseline hemoglobin level). some authors recently reported that saline infusion or albumin boluses may be an alternative priming approach in patients with body weight ranging from to kg [ ] . however, it should be proven that this approach could be safely transferred to population weighting less than kg. currently used ecp techniques include the "off-line" and the "in-line" devices [ ] . in our center, both techniques were used in different time periods with no difference in response rate observed. the number of wbc collected and mnc reinfused did not affect clinical outcome. notably, all patients underwent the procedure with the bilumen central venous line already in place (hickman-broviac bard access systems, salt lake city, ut, usa), which is different from the majority of reports, in which a larger central venous line (for instance, quinton) is placed. it would be interesting to extend our experience to determine if urokinase anticoagulation allows the proper flow rate of pre-existing central venous line. further, because of the experience of the staff in completing the procedure, no patient required sedation. the ci of cgvhd in pediatric population ranged from % to % according to the source of stem cells (hla-identical sibling cord blood versus matched unrelated donor peripheral blood) [ , ] , whereas in the aieop experience, the ci of cgvhd was reported to be % [ ] . in our small series, the incidence of cgvhd was %. the majority of our children presented progressive cgvhd ( %) and few had quiescent cgvhd ( %). for this reason, it is hard to determine if patients previously treated with ecp for agvhd could benefit from retreatment. our data are consistent with literature and the results encourage us in exploiting this promising approach for agvhd. in conclusion, a standardized approach to ecp treatment is needed for pediatric patients. from this perspective, sharing single-center experience is of great value in building experience; however, it is time to propose randomized prospective trials. 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rattray, rogan m.; press, richard d. title: the clinical impact of coronavirus infection in patients with hematologic malignancies and hematopoietic stem cell transplant recipients date: - - journal: j clin virol doi: . /j.jcv. . . sha: doc_id: cord_uid: uf ao x background: compared to other respiratory viruses, relatively little is known about the clinical impact of coronavirus (cov) infection after hematopoietic stem cell transplant (hsct) or in patients with hematologic malignancies. objectives: to characterize the role of cov in respiratory tract infections among hsct and hematologic malignancy patients. study design: we conducted a retrospective review of all cases of cov infection documented by polymerase chain reaction, (pcr)-based testing on nasopharyngeal and bronchoalveolar lavage fluid samples between june and . cases of cov infection occurring in hsct and hematologic malignancy patients were identified and the clinical characteristics of these cases were compared to other respiratory viruses. results: cov was identified in . % (n = ) of all samples analyzed (n = ) and in . % of all samples testing positive for a respiratory virus (n = ). of ( . %) of patients in whom cov was identified were hsct and hematologic malignancy patients. among these patients, cov was detected in . % of unique infection episodes, with only rhinovirus/enterovirus (rhv/env) infection being more common. group i cov subtypes accounted for . % of cases, and % of infections were diagnosed between december and march. cov infection was associated with upper respiratory tract symptoms in most patients, similar to other respiratory viruses. possible and proven lower respiratory tract disease was less common compared to other respiratory viruses except rhv/env. conclusions: cov is frequently detected in hsct and hematologic malignancy patients in whom suspicion for a respiratory viral infection exists, but is less likely to progress to lower respiratory tract disease than most other respiratory viruses. the clinical significance of respiratory viruses such as influenza, respiratory syncytial virus (rsv), parainfluenza viruses (pivs), human metapneumovirus (hmpv), rhinovirus (rhv), and adenovirus (adv) in patients with hematologic malignancies or recipients of autologous or allogeneic hematopoietic stem cell transplant (hsct) is well described [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] . far less data have been published that specifically address the impact of coronavirus (cov) infection in these patients [ ] . lower respiratory tract disease (lrtd) due to cov has been described on a case-report level [ ] [ ] [ ] [ ] [ ] , but a retrospective analysis of bronchoalveolar lavage (bal) samples obtained from hsct recipients with any acute pulmonary process did not identify cov in any sample [ ] . later, a prospective surveillance study in allogeneic hsct recipients performed at a single center over the course of one year found a relatively high cumulative incidence of cov infection ( %) in the first days following allogeneic hsct, but only one case in which cov was detected in a lower respiratory tract sample [ ] . other series studying the impact of respiratory viral infections have not specifically or directly assessed the clinical impact of cov in patients with hematologic malignancies and hsct recipients [ ] [ ] [ ] [ ] [ ] [ ] [ ] . with an apparently high frequency of infection but relative paucity of information pertaining to the impact of cov in patients with hematologic malignancies and hsct recipients, we conducted a retrospective review of all cov infections at our institution over a three year period in order to further characterize the role of cov in respiratory tract infections in these patients. all nasopharyngeal (np) and bal samples submitted from outpatient clinics and inpatient wards for respiratory virus testing between june , and july , at oregon health and science university (ohsu) were included in this study. demographic and clinical information pertaining to each patient testing positive for cov or other respiratory viruses assayed as part of a multiplex pcr panel (described below), were obtained by medical chart review. a patient was considered to have a hematologic malignancy if they were diagnosed with, and were receiving treatment for, any type of leukemia, lymphoma, multiple myeloma, aplastic anemia, mastocytosis, myelodysplastic syndrome, or amyloidosis. all patients who underwent allogeneic or autologous hsct, or cord blood transplant (cbt), were included and classified as such regardless of the underlying disease necessitating transplant. testing was performed using a multiplex respiratory virus pcr panel assay (xtag rvp; luminex) [ ] per manufacturer's instructions. since this assay does not distinguish between rhv and enterovirus (env), these results are grouped together in this analysis. the cov pcr reagents that are included in the rvp testing kit were further validated for clinical testing (under college of american pathologists (cap)-and clinical laboratory improvement amendments (clia)-regulatory conditions) by the ohsu molecular diagnostics lab. a unique episode of infection was counted at the initial identification of a respiratory virus in an np or bal sample. when the same respiratory virus was identified in multiple consecutive samples from the same patient without interim negative test results, this was considered as representative of prolonged shedding, and was therefore counted as a single unique episode of infection. mortality was attributed to respiratory virus infection if death was due to respiratory failure without identification of a cause other than a respiratory virus [ ] . respiratory virus inpatient-acquired infection was defined as onset of new symptoms, days or more after admission to an inpatient ward. criteria for possible and proven lrtd were identification of a respiratory virus in an np sample (possible lrtd) or bal sample (proven lrtd), along with new pulmonary infiltrates on thoracic imaging [ ] . comparison of categorical variables between viral groups was performed using two-tailed fisher's exact test. a total of np and bal samples obtained from the general hospital inpatient and outpatient populations -inclusive of, but not limited, to hsct recipients and hematologic malignancy patientswere analyzed. of these, samples ( %) submitted during unique episodes of infection tested positive for a respiratory virus (table ) . cov was the third most-common virus identified after rhv/env and piv , accounting for ( . %) positive samples. three bal specimens were positive for cov (see below), representing % of all cov-positive samples. of the unique episodes of infection, ( . %) were in the hematologic malignancy and hsct patient populations (table ) . cov was identified in ( . %) episodes among all patients, and ( . %) of episodes among hematologic malignancy and hsct patients. ( . %) of the episodes of cov infection in the hsct and hematologic malignancy population were diagnosed during the winter months of december-march ( fig. ). samples from the hematologic malignancy and hsct patients tested positive for cov, with the majority ( . %) being group i subtypes (nl (n = ) and e (n = )), as compared to group ii subtypes (oc (n = ) and hku (n = )). of the ( . %) cov positive samples were positive only for cov, and the other samples were co-infected with rhv [ ] , rsv a [ ] , hmpv [ ] , and both rsv a and adv [ ] . in order to better define the role of cov in respiratory tract syndromes among patients with hematologic malignancies and hsct recipients, we limited our clinical analysis to patients infected only with cov, excluding those co-infected with other respiratory viruses ( table ). patients were infected only with cov, and of those ( . %) had an underlying hematologic malignancy (n = ) or had undergone allogeneic (n = ) or autologous (n = ) hsct. the clinical characteristics of cov infection in these patients were examined and compared to episodes of mono-infection with other respiratory viruses (piv - , hmpv, rsv, and rhv/env) in the same patient population. the median age of those infected with cov was similar to the other respiratory viruses analyzed. surprisingly, cov was identified significantly more often in females ( %) than males compared to hmpv, piv - , and rhv/env, with a trend toward the same finding for rsv. five episodes of cov infection ( . %) were hospital-acquired. four of these cases, all subtype nl , occurred on the same inpatient hsct/oncology ward within a day period between january and march , . while indicative of an epidemiological link, we were unable to conclusively prove this. according to surveillance performed as part of ohsu's infection control program, during this time period, only two other cases of nosocomial acquisition of a respiratory virus occurred on this ward -one rhv/env and one influenza a (personal communication, dr. lynne strasfeld). of cases acquired as an outpatient, ( . %) required admission to the hospital, typically for evaluation of fevers or new pulmonary infiltrates. the requirement for hospitalization during cov infection was not significantly different from those during infection with other respiratory viruses. similar to most other respiratory viruses, the majority of patients ( . %) in whom cov was detected had uri symptoms (rhinorrhea, congestion, or sore throat). however, evidence of lrtd such as cough and new radiographic abnormalities was observed less frequently during cov infection, with a trend toward less hypoxemia as well. three patients ( . %), all allogeneic hsct recipients, met criteria for possible cov lrtd (tables and ) based on the identification of cov from an np specimen and new radiographic abnormalities [ ] , significantly less than during infection with the other respiratory viruses except rhv/env. none of these patients underwent bronchoscopy. two patients, both allogeneic hsct recipients, met criteria for proven lrtd based on identification of cov in a bal sample along with new radiographic abnormalities (tables and ) [ ] . however, one of these patients also had rsv, adv, and methicillin-resistant staphylococcus aureus identified in the bal specimen, making the contribution of cov to lrtd unclear. the combined incidence of possible and proven lrtd was significantly lower during cov infection compared to all of the other respiratory viruses analyzed except rhv/env. four of the five cases of possible or proven lrtd occurred after day + following hsct, and all five were associated with acute or chronic graft-versus-host disease (gvhd). a third episode of proven lrtd, accounting for the third cov-positive bal (table ) , occurred in a patient who had not undergone hsct and did not have an underlying hematologic malignancy; characteristics of that case were therefore not further analyzed. there were no cases in which mortality was directly attributable to cov infection. the results presented here complement and expand upon the few existing studies evaluating the role of cov infection in the immunocompromised host [ ] [ ] [ ] ] . cov, primarily group i subtypes, was frequently identified in the hsct and hematologic malignancy populations when suspicion for a respiratory viral infection prompted diagnostic testing. in this population, cov infection was associated with similar rates of uri symptoms, nosocomial acquisition, and requirement for hospital admission compared to most other respiratory viruses. however, cov infection resulted in less possible and proven lrtd than other respiratory viruses with the exception of rhv/env. the frequency of detection of cov among all samples tested in this study is consistent with previous reports of cov detection in patients hospitalized with respiratory syndromes [ ] [ ] [ ] . however, our results may not reflect the true prevalence of cov infection in our patients as we do not routinely screen asymptomatic patients; doing so would most likely increase the prevalence of infection due to detection in asymptomatic shedders [ ] . it is also important to note that the true rate of influenza a and b infection is under-represented in our results since the initial test of choice for these pathogens at our institution is an influenza-specific pcr assay, not the respiratory virus panel used in this study. accounting for this would reduce the relative frequency of cov as well as that of all other non-influenza respiratory viruses. we found that the hematologic malignancy and hsct populations accounted for the majority of patients testing positive for cov and the other respiratory viruses. this finding almost certainly reflects testing bias in that these patients are more likely to be tested for respiratory viruses when symptomatic given the importance placed on these pathogens [ ] . interestingly, we also found a female-predominant gender distribution among those infected with cov, which to the best of our knowledge, has not been reported before [ ] [ ] [ ] , ] . while the frequency of respiratory illnesses in the general population has been described to be higher among females, perhaps due to increased exposure [ ] , if this was responsible for the higher burden of cov infection among women compared to men, then a similar pattern might be expected for the other respiratory viruses as well. this finding requires confirmation by additional studies. the majority ( . %) of cov identified in the hematologic malignancy and hsct patients in this study belongs to group i. similarly, group i covs have been associated with infection in the immunocompromised host more than group ii covs in several series and case reports [ , , , , ] . in contrast, studies involving general or pediatric patient populations have reported a majority of cov belonging to group ii [ , , , , [ ] [ ] [ ] [ ] . group i and ii covs differ in the mechanism of virus entry into the host cell and the degree of genetic variability, but whether these differences affect pathogenicity is not known [ ] . indeed, reports associating cov subtypes with disease manifestations are conflicting [ , , ] . further studies into the relative incidence of cov subtypes among various patient populations are necessary and may provide insight into fundamental biological differences between group i and ii subtypes. similar to previous reports [ , ] , the rate of cov-proven lrtd was low but was not significantly different from other respiratory viruses. however, these results must be interpreted with caution since bronchoscopy, which we used as a criteria for defining "proven lrtd", is no longer done in many cases when a respiratory virus is identified in an upper airway sample at this institution or others [ ] . analyzing "possible lrtd" instead, cov was indeed associated with significantly less disease compared to all the other respiratory viruses analyzed except rhv/env, with the rate of possible lrtd due to the other respiratory viruses in this report being generally consistent with the published literature [ , ] . however, not all patients had radiography performed at the time of diagnosis of respiratory viral infection, which may bias the results toward those more likely to have evidence of lower respiratory tract illness. interestingly, all cases of possible and proven cov-associated lrtd occurred in hsct recipients, whereas the cases of possible and proven lrtd during infection with the other respiratory viruses were divided more evenly among hsct recipients and hematologic malignancy patients. additionally, all cases occurred in the setting of acute or chronic gvhd. taken together, these findings suggest that cov may be inherently less virulent compared to other respiratory viruses, and therefore, requires augmented immune suppression after hsct in order to progress to lrtd. the low number of cov-associated lrtd in this study precludes meaningful risk factor assessment but additional studies may make this more feasible. aside from lrtd, cov infection was associated with other clinically-relevant consequences. namely, % of outpatients were admitted to the hospital based on manifestations of cov infection, and hospital acquisition occurred in cases, with a cluster of nosocomially-acquired cases occurring within a month span. additionally, % of cov-infected patients received an empiric course of antibacterial therapy (data not shown). notably, while cov is detected as part of the respiratory virus panel used at our institution during the study period, the cov results were not reported in the medical record since the assay was not approved by the food and drug administration for this purpose. in theory, a "negative" respiratory virus panel result may lead to more frequent admission to the hospital for further medical evaluation, antibacterial use, or nosocomial transmission. however, we found the rates of these events during cov infection to be similar to those during infection with other respiratory viruses ( table and data not shown), highlighting the overall impact of respiratory viral infections in these patients in terms of utilization of medical resources. in conclusion, we found that cov is detected frequently in patients with hematologic malignancies and hsct recipients in whom suspicion for a respiratory viral infection exists, but is associated with less lrtd than other respiratory viruses except rhv/env. the retrospective nature of this work mandates confirmation of our findings by additional, prospective studies. human metapneumovirus infection in hematopoietic stem cell transplant recipients rhinovirus infections in myelosuppressed adult blood and marrow transplant recipients rhinovirus infections in hematopoietic stem cell transplant recipients with pneumonia clinical characterization of human metapneumovirus infection among patients with cancer adenovirus infections in adult 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malignancies epidemiology of viral respiratory tract infections in an outpatient haematology facility viral respiratory infections diagnosed by multiplex pcr after allogeneic hematopoietic stem cell transplantation: long-term incidence and outcome xtag rvp assay: analytical and clinical performance parainfluenza virus lower respiratory tract disease after hematopoietic cell transplant: viral detection in the lung predicts outcome clinical disease in children associated with newly described coronavirus subtypes epidemiology and clinical presentations of the four human coronaviruses e, hku , nl , and oc detected over years using a novel multiplex real-time pcr method epidemiology of viral respiratory infections a prospective molecular surveillance study evaluating the clinical impact of community-acquired respiratory viruses in lung transplant recipients human coronavirus infections in rural thailand: a comprehensive study using real-time reverse-transcription polymerase chain reaction assays human (non-severe acute respiratory syndrome) coronavirus infections in hospitalised children in france coronavirus hku and other coronavirus infections in hong kong new vaccine surveillance network, human coronavirus in young children hospitalized for acute respiratory illness and asymptomatic controls fields virology the authors wish to thank dr. lynne m. strasfeld and kevin langstaff for providing data pertaining to nosocomially-acquired respiratory viral infections at ohsu. no conflicts of interest. none. none declared. approval from the institutional review board of ohsu was obtained prior to beginning this study. key: cord- -tyrlpl authors: dsouza, kevin; pywell, cameron; thannickal, victor j. title: late noninfectious pulmonary complications in hematopoietic stem cell transplantation date: - - journal: oncologic critical care doi: . / - - - - _ sha: doc_id: cord_uid: tyrlpl hematopoietic stem cell transplantation (hsct) is an established therapeutic modality for a number of malignant and nonmalignant conditions. pulmonary complications following hsct are associated with increased mortality and morbidity. these complications may be classified into infectious versus noninfectious, and early versus late based on the time of occurrence post-transplant. thus, exclusion of infectious etiologies is the first step in the diagnoses of pulmonary complications. late onset noninfectious pulmonary complications typically occur months post-transplant. bronchiolitis obliterans is the major contributor to late-onset pulmonary complications, and its clinical presentation, pathogenesis, and current therapeutic approaches are discussed. idiopathic pneumonia syndrome is another important complication which usually occurs early, although its onset may be delayed. organizing pneumonia is important to recognize due to its responsiveness to corticosteroids. other late onset noninfectious pulmonary complications discussed here include pulmonary venoocclusive disease, pulmonary cytolytic thrombi, pleuroparenchymal fibroelastosis, thoracic air leak syndrome, and posttransplant lymphoproliferative disorders. hematopoietic stem cell transplantation (hsct) is an established therapeutic modality for a number of malignant and nonmalignant conditions. pulmonary complications following hsct are associated with increased mortality and morbidity. these complications may be classified into infectious versus noninfectious, and early versus late based on the time of occurrence post-transplant. thus, exclusion of infectious etiologies is the first step in the diagnoses of pulmonary complications. late onset noninfectious pulmonary complications typically occur months posttransplant. bronchiolitis obliterans is the major contributor to late-onset pulmonary complications, and its clinical presentation, pathogenesis, and current therapeutic approaches are discussed. idiopathic pneumonia syndrome is another important complication which usually occurs early, although its onset may be delayed. organizing pneumonia is important to recognize due to its responsiveness to corticosteroids. other late onset noninfectious pulmonary complications discussed here include pulmonary venoocclusive disease, pulmonary cytolytic thrombi, pleuroparenchymal fibroelastosis, thoracic air leak syndrome, and posttransplant lymphoproliferative disorders. hematopoietic stem cell transplantation (hsct) is an established form of therapy for a number of malignant as well as nonmalignant conditions. more than , hscts were conducted in the united states in (https://www.cibmtr.org). the two main types of hscts are autologous and allogenic. autologous hsct involves collection of stem cells from the patient that are then infused back after chemotherapy. allogenic hsct involves infusion of stem cells from a donor. the morbidity and mortality from hsctrelated complications have been on the decline over the past several years. these complications are broadly categorized based on etiology, namely, whether they are infectious or noninfectious, and the time of onset of such complications, early vs. late. pulmonary complications are the most common life-threatening complications post hsct occurring in - % of patients [ ] . we define late onset complications are occurring months after post-hsct (fig. ) . in this chapter, we discuss late noninfectious pulmonary complications in hsct, and current concepts on their pathogenesis, diagnosis, and management. the primary focus will be on bronchiolitis obliterans (bo) which is the most common and carries the highest mortality of the late onset noninfectious complications [ ] . the other late onset noninfectious complications that will be discussed include idiopathic pneumonia syndrome, organizing pneumonia, pulmonary venoocclusive disease, pulmonary cytolytic thrombi, pleuroparenchymal fibroelastosis, thoracic air leak syndrome, and posttransplant lymphoproliferative disorders. bo typically occurs between months to several years post hsct and is inclusive of the spectrum of chronic graft versus host disease (cgvhd) [ , ] . bo is characterized by progressive, irreversible airway narrowing due to circumferential small airway fibrosis. there is limited understanding of bo pathogenesis. bo is a pathological diagnosis requiring invasive surgical lung biopsy, which is uncommonly performed in the clinical setting. bo syndrome (bos) is a more useful clinical diagnosis that is made based on irreversible airflow limitation on pulmonary function testing (pft) without the need for lung biopsy. bo is the most common late onset noninfectious complication of hsct. the reported incidence of bo/bos after allogeneic hsct varies based on the diagnostic criteria used, ranging from % to % in retrospective studies [ ] . a recent prospective study to evaluate the epidemiology of late non onset noninfectious complications after allogenic stem cell transplant reported a cumulative incidence of bos months posttransplant at . % [ ] . bo/bos following autologous hsct is rare but has been reported [ , ] . clinical features of bo/bos are nonspecific. in early stages of the disease, patients are asymptomatic and are identified by airflow limitation on pfts. nonproductive cough, dyspnea on exertion, and decreased exercise tolerance are common [ ] . physical examination may be normal or reveal signs of airflow obstruction such as wheezing, hyperinflation, or diffuse crackles. other causes of such presentations, in particular, respiratory infections, should be ruled out. the chest radiograph may be normal or reveal hyperinflation. high resolution chest tomography (hrct) reveals small airway involvement with features of air trapping evidenced by mosaic attenuation on expiratory views (fig. ) . histopathology, when available, shows narrowing or complete occlusion of the bronchiolar lumen due to subepithelial inflammatory fibrosis is a hallmark of bo (fig. ) [ ] . transbronchial biopsies are insufficient to yield a diagnosis and surgical lung biopsies often prohibitively expose patients to procedural risks. in most cases, bos can be diagnosed without a histopathological diagnosis using pfts in the appropriate clinical setting. the updated national institutes of health (nih) guidelines for diagnosing bos are based on the following criteria [ ] : (b) the fifth percentile of predicted is the lower limit of the % confidence interval. (c) for pediatric or elderly patients, use the lower limits of normal, defined according to national health and nutrition examination survey iii calculations [ ] . . fev < % of predicted with % decline over less than years. fev should not correct to > % of predicted with albuterol, and the absolute decline for the corrected values should still remain at % over years. . absence of infection in the respiratory tract, documented with investigations directed by clinical symptoms, such as chest radiographs, computed tomographic (ct) scans, or microbiologic cultures (sinus aspiration, upper respiratory tract viral screen, sputum culture, bronchoalveolar lavage). . one of the two supporting features of bos: (a) evidence of air trapping by expiratory ct or small airway thickening or bronchiectasis by high-resolution chest ct or (b) evidence of air trapping by pfts: residual volume > % of predicted or residual volume/total lung capacity elevated outside the % confidence interval it is important to recognize that a significant number of bronchioles must be involved to manifest airflow limitation on pfts or to develop clinical symptoms. hence, early stages of bo may be missed. several risk factors for the development of bos have been identified, and most of these are closely associated with the occurrence of cgvhd [ ] . increasing age of the donor/recipient, development of acute gvhd [ ] , abo incompatibility [ ] , the presence of extra thoracic gvhd, low circulating igg, and non-caucasian race [ ] have been identified in several retrospective studies. additional risk factors include the type of transplant procedure such a peripheral blood stem cell transplant [ ] and busulfan-based conditioning regimens [ ] . use of antithymocyte globulin (atg) as part of the pretransplant conditioning regimen is associated with a decreased incidence of bos [ ] . prior to meeting established criteria for bos, a decrease in fev from pretransplant levels has also been identified to be a risk factor for the subsequent development of bos [ ] . more recently, a % decline in fev from pretransplant to day posttransplant has shown to be predictive for the development of bos [ ] . the precise pathogenetic mechanisms involved in the development of bo are unclear. the pathogenesis appears to be multifactorial and may involve diverse etiologies [ ] . airway epithelial injury is the inciting factor secondary to gastroesophageal reflux disease, respiratory infections, and chemotherapeutic drugs. this is typically followed by a dysregulated immune response that leads to the development of fibrotic changes in small airways. t cells and humoral mechanisms have been implicated [ , ] . genetic polymorphisms in nod /card have been linked to susceptibility to bos [ ] . there are currently no effective treatments for bos complicating hsct. most treatment protocols are based on combinations of immunosuppressive drugs and, until recently, were largely based on expert opinion. traditionally, immunosuppression in the form of systemic steroids for extended periods has been used; alternatively, calcineurin inhibitors and azathioprine, agents that impair lymphocyte activation and proliferation, have been employed. long-term, high-intensity immunosuppression is no longer recommended due to the increased risk of infections [ ] . most clinical studies of treatments for bos are difficult to interpret due to small sample sizes, their retrospective nature, and confounding effects of treatment for cgvhd. these studies have included systemic corticosteroids [ , , ] , rituximab [ , ] , imatinib [ ] , etanercept [ , ] , as well as extracorporeal photopheresis [ ] . a recent retrospective matched cohort study recently showed that extracorporeal photopheresis improves survival in hsct patients with bos [ ] . treatment with inhaled budesonide/formoterol led to significant fev improvements in patients with mild/severe bos after allogeneic hsct [ ] . another trial showed fev stabilization using a combination of fluticasone, azithromycin, and montelukast along with pulse dosed systemic steroids [ ] . a randomized, double-blinded, placebo-controlled trial of azithromycin alone did not reveal any change in fev in late bos post hsct [ ] . prophylactic azithromycin has been shown to prevent the onset of lung transplant-related bos, as well as stabilize fevi in post-lung transplant bos [ , ] . however, prophylactic azithromycin has not shown to be effective in bos post-hsct in retrospective studies [ ] . a prospective study on the prophylactic use of azithromycin to prevent airflow decline resulted in early termination secondary to hematological relapses [ ] ; an fda warning was issued in august until further review (https://www.fda.gov/ drugs/drugsafety/ucm .htm). prospective trials of azithromycin, bortezomib, inhaled cyclosporine, and neutrophil elastase inhibitors for prophylaxis and/or treatment of bos are underway (www.clinicaltrials.gov). current treatment strategies include high-dose inhaled glucocorticoid with or without a long acting inhaled beta agonist based on symptoms, with close monitoring of lung function with pfts. if progressive decline in fev occurs with this regimen, initiation of a combination of fluticasone, azithromycin, and montelukast (fam) therapy can be considered [ ] . occasionally, patients with chronic gvhd and refractory bos may respond to increased immunosuppression. in patients who progress despite medical therapies, lung transplantation may be the only option [ , ] . novel therapeutic approaches for bo/bos are currently being investigated. the pleiotropic small molecule p mak inhibitor, pirfenidone, has been shown to ameliorate bo in a murine model of cgvhd [ ] . an early phase clinical study evaluating the safety and tolerability of pirfenidone in bos is currently underway (www.clinicaltrials.gov; nct ). the clinical course of bos is variable, with some patients experiencing rapid declines in lung function, while others stabilize or improve. mortality from bos is most commonly due to progressive respiratory failure. bos confers a . fold increased risk of death after diagnosis [ ] . early onset of bos and lower fvc, especially within a year of transplant, is associated with a worse prognosis [ , ] . recent estimates indicate a - year survival of - %, and a -year survival of - %; this is an improvement in overall survival from a decade ago, when -year survival was %, and -year survival was %. early recognition, newer treatment strategies, and better supportive care likely account for this improved survival [ ] . idiopathic pneumonia syndrome (ips) is a severe noninfectious complication of hsct with an incidence of % when it was first described in the s; more recently, incidence is cited at - % [ , ] . ips is more common as an early complication of hsct but can occur after months. median time of onset is days posttransplant, with a range from to days [ ] . ips is an acute lung injury process characterized by diffuse alveolar damage in the absence of an identifiable lower respiratory tract infection. although the exact cause of ips remains unknown, immune involvement has been invoked; murine models involving immune mismatches between donor and recipient support this concept [ , ] . elevated levels of lipopolysaccharide (lps) and tumor necrosis factor-alpha (tnf-α) have been observed in bal samples of murine ips models [ ] . tnf-α may contribute to pathogenesis by direct toxicity, upregulation of major histocompatibility complex (mhc), increased leukocyte recruitment, and cellmediated apoptosis [ ] . donor t lymphocytes secrete chemokines which further amplify the inflammatory cascade [ ] . decreased level of pulmonary surfactant has also been associated with ips development [ , , ] . risk factors for ips include full-intensity conditioning regimens such as total body irradiation and busulfan, acute gvhd, advanced age, and underlying acute leukemia and myelodysplastic syndrome (mds) [ , , ] . reduced-intensity conditioning regimens have decreased the incidence of ips [ ] . in children, risk factors for ips include the underlying disease and busulfan-based conditioning regimens [ ] . interestingly, risk of ips increases with the number of platelet transfusions received, though the transfusion requirement could be a marker of disease severity [ , ] . the definition of ips, updated in the american thoracic society guidelines, is based on the following criteria [ radiographic findings can be nonspecific, but hrct findings include ground glass opacities that are bilateral, central, symmetric, with consolidation seen in more severe cases [ ] . recent advances in diagnostic capabilities have increased detection of occult infections which help separate ips from infectious hsct complications; the distinction is critical due to their vastly distinct treatment approaches. many patients diagnosed with ips were later discovered to have detectable pathogens, most commonly hhv- , human rhinovirus, and aspergillus, when their bal samples were re-examined [ ] . historically, treatment of ips has been largely supportive. once infections have been ruled out, systemic corticosteroids are the mainstay of treatment; ips in association with diffuse alveolar hemorrhage may require higher doses [ ] . the results of other immunotherapeutic agents such as the soluble tnf-α inhibitor, etanercept, has been mixed. a retrospective single-center study over two distinct timeperiods comparing steroids alone (earlier timeperiod) versus combined steroids and etanercept showed significant improvement in survival to hospital discharge [ ] . however, the more recent use of reduced-intensity conditioning regimens and improved supportive care may have accounted for this improvement. patients with late-onset ips who responded to etanercept have greatly improved short-and long-term mortality [ ] . results in children have been similarly promising, with complete response in % of patients [ ] . a randomized, doubleblind, placebo-controlled trial comparing corticosteroids with placebo to corticosteroids with etanercept was inconclusive due to slow patient accrual and early termination [ ] . other potential therapies are under investigation. blockade of nf-κb, a transcription factor downstream of the tnf-α receptor, has shown promise in murine models [ ] . pulmonary surfactant replacement is also being studied as an intervention to treat ips [ ] . outcomes for ips remain poor, with mortality rates between % and %. rapid clinical deterioration can occur and > % of patients requiring mechanical ventilation succumb to the disease [ , , ] . veno-venous extracorporeal membrane oxygenation (ecmo) as a rescue therapy or bridge-to-recovery has met with mixed results [ , ] . short-term survival has improved with treatment advances, but -year survival remains low. based on a small study, a more rapid peak and decline in severity of infiltrates on hrct has been linked to a more favorable prognosis [ ] . biomarkers to predict patients who at risk for ips and who respond to biologic therapy are being studied [ ] . organizing pneumonia (op), formerly known as bronchiolitis organizing pneumonia (boop), is a complication of hsct. it can occur as a part of the ips spectrum or as a stand-alone late onset pulmonary complication of hsct [ ] . op as a complication of hsct was first described by thirman et al. in [ ] . although it has been described as a complication of both autologous and allogenic hsct, it is more common following the latter. the incidence of op post-allogenic hsct ranges from . % to . % with a median time of onset post-hsct of days [ ] . among patients, who underwent allogenic hsct, cases of histological boop/op was reported, an incidence of . % [ ] . op presents with fever, nonproductive cough, and dyspnea and can be precipitated by a recent taper of immunosuppressive regimen. pfts commonly reveal a restrictive pattern, but could be normal, obstructive, or mixed with a decreased diffusion capacity of carbon monoxide (dlco) [ , ] . high resolution chest tomographic scans in patients with op are notable for airspace consolidations, ground glass opacities, and nodular opacities (fig. ) [ ] . in a study of patients with biopsy-proven op post-hsct, ground glass opacities were noted to be the most common radiological feature [ ] . bronchoalveolar lavage is useful to exclude infections. surgical lung biopsy is the gold standard for diagnosis of op, as transbronchial biopsies have lower yield; however, the latter approach may be useful in certain clinical settings [ , ] . histopathological features on biopsy include presence of the buds of granulation tissue which contain myofibroblasts and a loose connective tissue (fig. ) . these buds are intra-alveolar and can extend into the bronchioles causing obstruction with associated mild interstitial inflammation [ ] . a prior history of acute or chronic gvhd was found to be a strong risk factor for op [ ] and suggests a common pathogenesis. in a study of patients of post-allogenic hcst recipients, hla disparity, female-to-male hsct, and peripheral blood stem cell transplant (pbsct) were associated with an increased risk of developing op. in contrast, busulfan-based or fludarabine-based reduced-intensity conditioning compared to total body irradiation and t cell depletion regimens were associated with a lower risk [ ] . the precise pathogenetic mechanisms of post-hsct op are unclear. the association with gvhd and increased incidence post-allogeneic hsct suggests alloimmune-mediated lung injury. murine models of op after respiratory reovirus infections have demonstrated the role of t cells and cytokines such as interferon-α in the development of the process [ ] . hsct-associated op, as in most other cases of op, is primarily treated with high-dose systemic corticosteroids. there are no specific guidelines on the dose and duration of steroid therapy. current recommendations are based on expert opinion, and clinical judgment should be exercised. prednisone in doses ranging from . - . mg/kg/ day have been used for - months with a slow taper over - months [ ] . further studies are needed to establish ideal doses and duration of corticosteroid therapy. erythromycin in combination with corticosteroids has been reported with favorable outcomes [ ] , although the role of macrolides for treatment of op is unclear. about % of patients with hsct-associated op have a favorable prognosis with resolution or stabilization after corticosteroid therapy [ ] . clinical improvement is seen usually within a week of starting therapy followed by radiological improvement. failure to respond to treatment may result in progressive respiratory failure and death [ ] . pulmonary venoocclusive disease (pvod) resulting in pulmonary hypertension is a rare late onset complication of both autologous and allogeneic hsct [ ] . in reported cases, it was noted in patients less than years old and presented several weeks to months posttransplant [ ] . presenting complaints are usually nonspecific, primarily fatigue and exertional dyspnea. physical examination findings are similar to those of pulmonary hypertension which may be normal in early stages and become more apparent in later stages: elevated jvd, peripheral edema, and hepatomegaly. elevated second heart sound, parasternal lift and palpable second heart sounds in the second left intercostal space can be recognized in some patients along with tricuspid regurgitation murmur. computed tomographic (ct) scans of the chest may show septal thickening, diffuse or mosaic ground glass opacities, small nodules, and areas of consolidation. right heart catheterization reveals an increased pulmonary artery pressure and normal pulmonary capillary pressure. the triad of severe pulmonary hypertension in the setting of normal pulmonary artery occlusion pressure and radiographic evidence of pulmonary edema could be suggestive of pvod but is not diagnostic. pvod can be definitively diagnosed only by surgical biopsy and is characterized by the progressive intimal proliferation, fibrosis, and occlusion of the pulmonary venules as well as small veins [ ] . the pathogenesis of pvod post hsct is unclear and has been attributed to toxic endothelial injury secondary to chemotherapeutic conditioning regimens and/or viral infections [ ] but does not seem to be associated with cgvhd [ ] . there are no effective treatments for pvod and prognosis is poor. conventional arterial vasodilator therapy for pulmonary hypertension could worsen pulmonary edema in pvod and if initiated should be done with close monitoring [ , ] . reported cases of pvod post hsct have shown some favorable response to steroid therapy [ , , ] . pulmonary cytolytic thrombi (pct) is a complication of allogenic hsct and primarily occurs in children with gvhd [ ] . the incidence is between . % and % with a median onset of months post hsct [ ] . patients typically present with fever, cough, and dyspnea. ct scans of the chest shows peripheral pulmonary and pleural nodules [ ] . bronchoalveolar lavage is indicated to exclude infectious etiology. the diagnosis of pct is based on surgical biopsy of the lung nodules which are characterized by vascular occlusive and hemorrhagic infarcts secondary to thrombi containing intensely basophilic amorphous material as well as entrapped leucocytes [ ] . the prognosis of pct is favorable as it responds to treatment with systemic corticosteroids. there has not been any mortality attributed to pct in the reported literature. pleuroparenchymal fibroelastosis (ppfe) is a rare complication of hsct, grouped under rare interstitial pneumonias with a prevalence of around . % in hsct recipients [ ] . it is characterized by progressive sub pleural fibrosis predominantly in the upper lobes. ppfe has been reported post allogeneic and autologous hsct. the etiology of ppfe post hsct is unclear. chemotherapeutic drugs, radiation therapy and a possible association with cgvhd have been hypothesized [ ] to be predisposing factors. patients can present with dry cough, exertional dyspnea, and chest pain secondary to spontaneous pneumothorax [ ] . pfts reveal a restrictive or mixed picture of obstruction and restriction. ct scan is characteristic of pleural thickening, fibrosis, subpleural reticulations, and traction bronchiectasis predominantly in the upper lobes [ ] . histopathological exam reveals alveolar collapse, subpleural fibrosis, and extensive elastic deposition [ ] . the disease is progressive with worsening symptoms and poor prognosis. currently no therapeutic options are available except for lung transplantation [ ] . thoracic air leak syndrome (tals) consists of a spectrum of conditions that includes spontaneous pneumothorax, subcutaneous emphysema, pneumomediastinum, interstitial emphysema, and pneumopericardium. it occurs as a late onset complication of allogenic hsct and is associated with cgvhd. according to reported literature, prevalence of tals post allogenic hsct is reported to be . - . % with a median time range of - days post hsct [ ] . the mechanism of air leak in bos is secondary to increased intra-alveolar pressure leading to alveolar rupture into the pulmonary interstitium with subsequent retrograde dissection of air into the mediastinum and subcutaneous tissue causing pneumomediastinum and subcutaneous emphysema. similarly, rupture into the pleural space causes pneumothorax [ ] . ct scan of the chest is the imaging modality of choice and can detect air in the pleural space, mediastinum, and subcutaneous tissue. infections, cough, emesis, and other causes of a similar presentation should be ruled out. prognosis is poor with -year and -year survival of % and %, respectively [ ] . post-transplant lymphoproliferative disorder (ptld) is a rare complication following allogeneic hsct and has a cumulative incidence of % [ ] . in a study of , hsct patients' risk factors such as unrelated or hla mismatched donors, use of atg or monoclonal antibodies against t cells for gvhd prophylaxis or treatment, t cell depleted donor marrow, age > years and second hsct were identified. the incidence of ptld ranged from . % in patients with no risk factors to more than % with more than risk factors [ ] . the pathogenesis of post hsct ptld is attributed to proliferation of donor epstein-barr virus (ebv) infected b cells in the setting of weakened t-cell immunity. though post hsct is common in the lymph nodes, spleen, and liver, pulmonary involvement occurs in about % of the cases. median onset is around - months post hsct [ ] . clinical presentation varies and could range from asymptomatic to fulminant tumor lysis syndrome. ct scans of the chest are notable for multiple pulmonary nodules with basal and peripheral predominance, mediastinal and hilar lymphadenopathy, patchy consolidation, pleural effusion, and chest wall or pleural-based masses [ ] . diagnosis may require a biopsy or could be made via noninvasive methods in the appropriate clinical setting. a probable diagnosis is made by increased ebv dna levels in the setting of lymphadenopathy or hepatosplenomegaly and when other causes have been ruled out, proven disease requires a biopsy. treatment strategies include rituximab, reduction of immunosuppression, donor lymphocyte infusion, chemotherapy, and ebv-specific cytotoxic t lymphocyte infusions [ ] . the prognosis of ptld post hsct is poor compared to that occurring after solid organ transplantation [ ] . late onset pulmonary complications following hsct are a major cause of mortality and morbidity. advances in transplant techniques, earlier diagnosis, prevention, and management of infectious complications have led to better outcomes as well as shifted the focus to late noninfectious pulmonary complications in hsct. these complications are myriad and require further studies to develop more effective screening, 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transplant consortium and children's oncology group study (asct ) bronchiolitis obliterans syndrome (bos), bronchiolitis obliterans organizing pneumonia (boop), and other late-onset noninfectious pulmonary complications following allogeneic hematopoietic stem cell transplantation key: cord- - t zix authors: wallhult, elisabeth; quinn, barry title: early and acute complications and the principles of hsct nursing care date: - - journal: the european blood and marrow transplantation textbook for nurses doi: . / - - - - _ sha: doc_id: cord_uid: t zix haematopoietic stem cell transplantation (hsct) generally includes preparative or conditioning regimes containing chemotherapy and/or radiotherapy in high doses. these regimens, as well as other treatments before and after hsct such as immunosuppressive drugs to prevent graft versus host disease (gvhd) (see chap. ), may affect the patient’s organs and tissues and may cause both acute and long-term complications. in the evolving field of stem cell therapies, some complications that traditionally have been regarded as early complications are now, due to changes in preparative regimens and choice of stem cell source, sometimes seen later in the post-transplant out-patient setting. the complications covered in this chapter generally occur within days post hsct and are thus classified as early complications. two of the most common early complications are oral complications/mucositis and sepsis. some other relatively rare complications are also covered here: haemorrhagic cystitis (hc), endothelial damage (ed) syndromes including engraftment syndrome (es), idiopathic pneumonia syndrome (ips), diffuse alveolar haemorrhage (dah), transplant-associated microangiopathy (tam) and sinusoidal obstruction syndrome/veno-occlusive disease (sos/vod). for all complications, recommendations for prevention and principles for nursing care are presented since careful nursing monitoring, prompt intervention and care may have an influence on patients’ morbidity and mortality. oral care in transplantation mindful of the many developments in the field of hsct aimed at improving survival and quality of life, the correct and consistent approach to managing oral care problems still remains a challenge in many transplant settings across europe. there is much evidence to show that rather than taking a proactive approach to this aspect of care, many clinicians simply react to oral complications once they occur with a sometimes inconsistent and anecdotal approach. oral problems and damage may be temporary or permanent resulting in a significant health burden for the individual while making substantial demands on limited healthcare resources. however, oral complications are not always inevitable, and much can be done to reduce or minimise the severity of symptoms by taking a more proactive approach to this aspect of care. working as a multidisciplinary team with the patient at the centre of care and treatment plan, the early detection of potential and actual problems and treatment can help to reduce oral problems and prevent interruptions to treatment while maximising patient safety and comfort (national cancer institute ). oral mucositis (om) has been defined by rubenstein et al. ( ) , al-dasoogi et al. ( ) and others as the inflammation of the mucosal membrane, characterised by ulceration, which may result in pain, swallowing difficulties and impairment of the ability to talk. the mucosal injury caused by om provides an opportunity for infection to flourish, and in particular putting the severely immunocompromised patient in the hsct setting at risk of sepsis and septicaemia. om and oral problems in the hsct setting (table . ) can be expected to occur in as many as % of patients undergoing autologous hsct and % of patients undergoing allogeneic hsct (filicko et al. ; bhatt et al. ). with the increasing use of targeted drug therapies and approaches in the cancer and haematology setting, problems in the oral cavity will increase and become even more of a challenge . all treatment strategies aimed at improving mouth care are dependent on four key principles: accurate assessment of the oral cavity, individualised plan of care, initiating timely preventative measures and correct treatment (quinn et al. ). the assessment process should begin prior to hsct by identifying all the patient risks most likely to increase oral damage. each patient needs to be assessed in relation to the following risk factors that may put them at higher risk of oral complications during treatment: patients about to commence hsct should undergo dental assessment by a specialist (elad et al. ) . this is to establish general oral health status and identify and manage existing and/or potential source of infection, trauma or injury. any identified dental problems should be corrected before starting treatment regimen. some patients will need regular dental follow-up following treatment. a further baseline assessment of the oral mucosa should be taken as close to the administration of the first treatment dose as possible. the oral cavity should be assessed by trained healthcare professionals using a recognised grading system to ensure accurate monitoring and record keeping. the tool chosen should contain both objective and subjective elements. the assessment should include changes to the oral mucosa, the presence or absence of pain and the patient's nutritional status (quinn et al. ) . assessments should be completed daily during hsct and at regular intervals posttreatment to monitor for complications. patients can be encouraged to assess their own mouth using a patient-reported tool and to report any changes they notice or experience to the transplant team. • clinical tools: good light source, gloves, tongue depressor and dry gauze • patient in convenient and comfortable position • use valid and reliable assessment instrument which is easy to interpret • oral sites to be evaluated (cheeks, lips, soft palate, floor of mouth, tongue) care of the oral cavity is central to helping to prevent and/or reduce oral complications during and after treatment. the oral care team in the hsct setting includes dental professionals, dietician, nurse, doctor and pharmacist. the support provided by the team along with good communication and the patient at the centre of all care plans is central to maintaining patient's oral health. all patients should be provided with clear instructions and encouraged to maintain good oral hygiene. education should also include potential oral complications to enable patients to identify and report these early (clarkson et al. ; quinn et al. ) . all patients should receive written information, as well as verbal instruction, about oral care as part of the prevention and treatment of oral changes. good nutrition is vital in helping to fight infection, maintain mucosal integrity, enhance mucosal tissue repair and reduce exacerbation of existing mucositis. issues that may affect nutrition such as loss of appetite, taste changes and dysphagia should be addressed. there are certain foods that can damage the oral mucosa; this may include rough, sharp and hard foods and should be avoided. spicy, very salty and acidic foods may cause mucosal irritation but may be preferred or tolerated by some patients. brushing of teeth, gums and tongue should be performed two to four times a day preferably after meals and before going to bed (peterson et al. ) . soft-bristled toothbrush (manual or electric) is recommended to prevent injury to the oral mucosa and must be rinsed thoroughly with water after each use. if the mouth is painful or patients cannot open their mouths fully, soft oral sponges may be used. to prevent infections, the toothbrush should be stored with the brush head upwards and not soaked in disinfectant solution. these should also be monitored for evidence of fungal/bacterial colonisation. in order to protect the enamel, nonabrasive toothpaste containing mild fluoride ( - ppm) should be used. daily interdental cleaning with brushes may reduce plaque formation between the teeth (sambunjak et al. ). however, the use of interdental cleaners should be used with caution for patients with thrombocytopenia or clotting disorders. after each meal, dentures must be rinsed. thorough cleaning by brushing with soap and water should be performed at least twice a day. dentures should be cleaned, dried and stored in a closed container overnight (duyck et al. ) . the goal of using mouthwashes may include oral hygiene, preventing/treating infection, moistening the oral cavity or providing pain relief. as a minimum to keep the mouth clean, bland gargles and rinses with water, normal saline ( . % nacl) and saltwater are recommended at least four times a day (lalla et al. ; quinn et al. ) . some patients will require assistance; it may be necessary for healthcare professionals to perform/support oral care through rinsing with normal saline ( . % nacl) (elad et al. ) , with or without suction. lubricants, lip balm or lip cream may be used to moisten the lips. patients should maintain adequate hydration and drink water frequently to keep the mouth moist. several factors could contribute to dryness such as oxygen therapy and supportive care medications (e.g. antidepressants, antihistamines, sedatives and opioids). to keep the oral mucosa moist, regular sipping or spraying water may help. use of saline sprays and mouthwashes as well as use of saliva substitutes may be used. there is anecdotal evidence that fresh pineapple chunks may also help stimulate saliva but should be used with caution as their acidity could irritate the oral mucosa and affect the teeth (lalla et al. ). the choice of prevention regimens should be guided by evidence-based or expert opinion interventions, working with the individual patient and the potential risk of oral mucositis which may include the following (adapted quinn et al. while good oral hygiene is fundamental, antifungal and antiviral treatments will be prescribed to reduce infections in patients in the transplant setting. patients should receive an antifungal agent given orally or intravenously. antiviral prophylaxis should also be given. the choice of drug will be dependent on local policies/ guidance. all treatment plans should be based upon the grading of oral damage and patient reports; these may include the following. • once oral damage develops, patients should be supported to continue oral care. • frequency of oral rinsing may be increased. the aim is to keep the oral surfaces clean and moist (elad et al. ). • check for oral infections, swab and treat appropriately. review of antifungal treatment, local or systemic, should be administered if required (watson et al. ). • dexamethasone containing gels may be used for aphthous lesions. • consider mucosal protectants ). • dietary requirements should be assessed and foods causing discomfort avoided. • swallowing problems, malnutrition and weight loss should be monitored and patients given support/advice. adjustments to food consistency, methods of intake, food fortification and methods of intake should be assessed, support and education offered to patients. use of supplement drinks, peg, rig or nasogastric feeding should be considered ). • fluid intake should be assessed and route of administration of pain relief continually monitored. general health problems should also be assessed (swallowing of tablets, decreased blood sugar levels and decreased blood pressure, decreased renal function leading to overdosing of substances). • patients will need adequate pain medication including topical and systemic analgesia such as paracetamol, codeine, morphine rinses, benzydamine mouthwash, trimecaine and lidocaine. patients should be offered education on use and possible side effects including numbness of the oral mucosa. • increase pain medication following patient needs • increase nutritional support • increase oral rinses and care when oral damage progresses, closer monitoring and support for patients is required. an important aspect of care is to control the pain thereby helping the patient to continue food and fluid intake, communication and sleep. for topical treatment the use of topical analgesics can be intensified. there is insufficient evidence that many products reduce the severity of mucositis but comfort can be provided for the patient by some of these oral care products. institutions can offer a range of mouthwashes selecting the most appropriate for the clinical situation and the patients trying out which one works best for them. generally spoken, topical antibacterial substances are not recommended. the use of oral rinses, topical gels or films can be individually considered. any with sufficient safety and positive experiences can be used: caphosol ® , mugard ® , oralife ® , gelclair ® and episil ® are just a few of them. for systemic pain medication, it is useful to follow a step-by-step increase, with the aim of the patient becoming pain-free within h. it can be helpful to monitor the efficacy of pain medication with pain assessment tools. institutions should follow a standardised pattern of pain medication following the who recommendations where applicable. in severe mucositis, the use of opiates with the optimal application route should be considered. the best route of application depends on many individual and setting factors and may be oral, subcutaneous, intravenous or transdermal with patches. patients may require a combination of slow-release and fast-acting drugs. patient controlled analgesia should be considered. careful monitoring should include pain relief and any potential side effects, and including family members may prove helpful to obtain a wider view of how well the patient copes outside the treatment unit. bleeding from om continue mouth gargling. tranexamic acid has been widely used in oral surgery, and gargling/ swishing with tranexamic acid ( mg) as a mouthwash may be worth considering (watson et al. ) . as this may be due to or increased by concurrent mediation, a review of the patient's medications is needed and if possible adjustments made. patients should be encouraged to increase sip-ping of fluids. artificial saliva, viscous solutions and gels to protect and moisten the mucosa should be considered; patients should be counselled on correct application. in chronic radiotherapy-related xerostomia, pilocarpine should be used. this is a common side effect during and post high-dose radiotherapy. patients should be given helpful exercises, and the team may consider mechanical devices to help alleviate the problem. oral damage may be a hallmark of graft versus host disease (gvhd) in patients following allogeneic stem cell transplantation, and the presence of lichenoid hyperkeratotic plaques (diagnostic sign), gingivitis, mucositis, erythema, pain, xerostomia and ulcers may indicate gvhd. kuten-shorrer et al. ( ) suggest that solutions of dexamethasone or other steroids are used as firstline treatment; second-line may include solutions of steroids in combination with other immunosuppressant drugs. oral damage in the hsct will require several weeks/months to heal, and patients need continuing support and care during this period. advice and support by suitably qualified health professional should continue during this period. support to manage side effects including pain and the gradual reduction of analgesia is extremely important. chronic side effects may include dental decay, trismus, fibrosis, lymphedema, chronic xerostomia and chronic pain and will require careful management. all patients should be individually assessed and appropriate care and treatment given. follow-up care should be planned and supervised to address longer-term and late complications. the principles presented here are intended as a support and in no way should replace clinical decision-making related to the particular patient and clinical situation. depending on the severity of oral complications and the impact on the patient, the team will need to review the plan of care. sepsis and principles of care the increased risk of infections in patients undergoing haematopoietic stem cell transplantation (hsct) is well known, and infection is a leading cause of morbidity and mortality. hsct patients are particularly at risk, especially during the neutropenic period following the conditioning treatment. in hsct patients, signs and symptoms of sepsis may be subtle and difficult to recognise due to neutropenia or other complications of the transplant procedure. preventive measures should be applied, but vigilance and close monitoring of the patient, strong team collaboration and immediate action will allow for prompt and appropriate management of septic patients. (singer et al. ) , sepsis can be defined as: a life-threatening condition caused by aberrant and dysregulated host response to infection. the pathobiology is still not completely known but the divergent infection response injures the body's own tissues and organs and causes organ dysfunction. that is also what differentiates sepsis from infection in general. septic shock is a subset of sepsis in which particularly profound circulatory, cellular and metabolic abnormalities are associated with a greater risk of mortality than with sepsis alone. the table below has limited clinical diagnostic relevance but is a schematic description of the evolution from systemic inflammatory response system (sirs) to septic shock (table . ). it is worth noting that the symptoms of sirs will not delineate between sirs and sepsis itself, and since many of the signs and symptoms may be present in hsct patients, an individual assessment, including other examinations as well, needs to be performed for the diagnosis of sepsis. the consequence of the inflammatory response and evolution of sepsis is called the sepsis cascade and is illustrated in fig. . . in the early phase of hsct, i.e. day to day + , the main risk factors for infections are (rovira et al. ): a longer period of neutropenia can be expected in allogeneic than in autologous transplant. the stem cell source also affects the length of the neutropenic period where peripheral blood (pbsc) has an expected neutropenic phase of about weeks, bone marrow (bm) weeks and cord blood (cb) weeks. myeloablative conditioning (mac) treatment will cause a longer neutropenic phase than reduced intensity conditioning (ric). all kinds of barrier breakdown will increase the infection risk and mucositis occur in almost all transplant patients.skin breakdown can be caused by, e.g. drugs and acute graft versus host disease (agvhd). indwelling catheters such as peripheral cannulas, central lines, urinary catheters and pyelostomy catheters are a potential port of entry for microorganisms into the blood stream. allogeneic transplant is always followed by long-lasting immunodeficiency. conditioning treatment may include t-cell depleting agents and even non-myeloablative regimens cause lymphodepletion with prolonged periods of immune incompetence. donor type and degree of histocompatibility (human leukocyte antigen (hla) match) are other factors that influence the time to immune reconstitution. immunosuppression for gvhd prophylaxis is necessary in allogeneic hsct and will delay immune reconstitution (toubert ) . need for immunosuppressive treatment will increase the risk for infections. the sepsis cascade starts with an inflammatory response that will cause microvascular injury, vasodilation and tissue hypoxia. the microvascular injury will lead to capillary leak resulting in oedema, decreased urinary output, tachycardia, with an initially bounding pulse which will then become weaker, and an increased respiratory rate. hypotension is another symptom caused by both microvascular injury and vasodilation. the vasodilation will also cause decreased renal blood flow. the hypovolemia will cause poor tissue perfusion causing tissue hypoxia with anaerobic metabolism. in this process oxygen and lactate are released for metabolism and thus causing metabolic acidosis (e-learning package sepsis and sepsis six http://sonet.nottingham.ac.uk/, ) mucosal or skin barrier breakdown further increases the risk. if the patient is not in remission at hsct, there is a greater risk for infection and sepsis. comorbidity, such as diabetes, is another risk factor. the most important action to prevent infections acquired by exogenous organisms is good hand hygiene performed correctly. all clinical staff should also wear a uniform that is clean and short sleeved. protective isolation during the neutropenic phase is recommended, and the patient should not be in contact with any staff or visitors with symptoms of infection. for prevention of endogenous infections, oral hygiene and skin care to maintain the mucosal and skin barrier and use of prophylactic antibiotics are the most important actions. correct handling of any indwelling catheters is also a key nursing responsibility in infection control. other areas where infections can be prevented are air and water quality, food hygiene and the environmental cleaning. environmental cleaning includes medical equipment as well. for more detailed guidance on infection control, see chap. . routine surveillance screening for infection by bacterial and/or fungal cultures, i.e. blood, urine, faeces, swabs from nasopharynx and central line insertion site and serum galactomannan blood test, may allow for earlier identification and implementation of therapy, although the benefit of such routines can be discussed (nesher et al. ) . regular monitoring of blood tests such as full blood count, electrolytes, urea and/or creatinine and c-reactive protein (crp) may assist in detecting any changes that could indicate infection. prophylactic antibiotics, e.g. fluoroquinolones, antifungal and antiviral medication, will be used in most hsct patients, at least during the neutropenic phase. early recognition and treatment is vital for a successful outcome of sepsis. temperature, pulse, blood pressure, respirations and saturation (vital signs) should be frequently monitored. signs of infection are not always obvious, but if the patient has a temperature ≥ . °c, cultures should be taken, i.v. antibiotics and i.v. fluids started or increased and oxygen therapy initiated. the goal is always to start antibiotic treatment within h from detection of fever (swedish "pro sepsis" programme group sepsis ) . this is sometimes referred to as "the golden hour" (or "door to needle time" for patients admitted from outside the hospital) and is the most critical period in the patient's survival from sepsis. recognising sepsis can be a challenge in hsct patients during the immediate posttransplant period where often a plethora of symptoms are present but also after discharge, in the outpatient setting, since some symptoms are rather unspecific. other than fever, chills or rigouring, feeling unwell or different (without clear explanation), changes in behaviour or mental changes, feeling faint or changes in skin tone can indicate sepsis. an increased respiratory rate can be seen even if saturation is normal. an increased pulse and lowered blood pressure may be noted. some patients may not develop fever, and hypothermia, i.e. < °c, can also be a sign of sepsis. if an outpatient with symptoms that could be sepsisrelated reports a normal body temperature, it should be checked again in the clinic with a reliable thermometer and correct method. diarrhoea and vomiting are frequently seen in sepsis but can easily be mistaken for gastroenteritis, mucositis or acute graft versus host disease (agvhd). diffuse or local pain, e.g. in the abdomen, is common. falls are often secondary to sepsis particularly in elderly patients. any of these indices need prompt and thorough assessment. the concept of the sepsis six has been developed as a guide to prioritise interventions in patients where sepsis is suspected (daniels et al. when sepsis is suspected, all cultures should be taken prior to commencing antimicrobials, if possible (rhodes et al. ) . cultures should be taken from central lines, wounds, nasopharynx, urine and faeces. it is also sensible to consider peripheral i.v. cannulae as a possible source of infection. despite conventional practice to collect blood cultures at a fever spike in order to increase the chances of detecting bacteraemia, there is so far no data to support this principle (kee et al. ) . testing could include polymerase chain reaction (pcr) virology (e.g. for cytomegalovirus (cmv) or epstein-barr virus (ebv)) and screening for fungus (e.g. oral swab), depending on symptoms and suspected microbial agent. for the procedures for diagnosis of central line-associated bloodstream infections (clabsi), please see chap. . laboratory tests should be taken to monitor electrolyte status, organ function, blood count and signs of infection. a site of infection may not always be identified. if a source of infection is confirmed, or strongly suspected, applicable actions should be taken, e.g. wound care or removal of peripheral i.v. needle with signs of thrombophlebitis (schorr et al. ) . upon initiation of antimicrobial treatment, a broad-spectrum antibiotic is usually used. depending on the results of the cultures performed, the chosen drug may need to be changed later. fever and infection will affect the blood count and frequently cause platelet consumption why transfusions may be necessary. early recognition and intervention are achieved by frequent monitoring of the patient's vital signs and general condition and paying attention to subtle changes that should be promptly reported. as described above, immediate action is required at the first indication of sepsis. when treatment has been initiated, the patient must be continually monitored to determine the effect of treatment or worsening of the condition. this includes vital signs, fluid balance including weight and assessment of identified and/or potential infection sites (mouth, skin, any indwelling or tunnelled catheter, urine, stools, etc.), mental status, signs of bleeding, pain and general appearance and well-being. antibiotics should be delivered with strict adherence to the prescribed time schedule. antipyretics should be avoided since they may mask fever but may under certain circumstances be used to alleviate patient discomfort and pain. laboratory tests results will guide the need for electrolyte replacement and blood product transfusion that may be ordered prophylactically or in case of bleeding. cultures may need to be repeated to confirm infection and/or response to treatment. oxygen should be administered as needed to ensure adequate saturation (i.e. ≥ %, or - % for patients with chronic obstructive pulmonary disease (copd) (royal college of physicians )). if the patient's condition worsens and organ support such as assisted ventilation or haemodialysis is required, the patient may need to be prepared for transfer to the intensive care unit (icu). extra psychological support is important for both the patient and family. educating the patient and the carer about the condition and actions taken or planned will prevent unnecessary worrying and enable them to alert the staff about symptoms or changes. information and education may also facilitate mental preparedness if the condition worsens and a higher level of care, icu, is needed. patients with sepsis are likely to need additional nursing care such as assistance with oral care and personal hygiene. it is important to ensure that the patient's and caregivers' information, education and support needs are met. on discharge from the hospital, we need to ensure that the patient and their caregiver are aware of when, why and how to contact the clinic or hospital that they have a fever thermometer at home, know when to take their temperature and are aware of the level that constitutes a fever. haemorrhagic cystitis haemorrhagic cystitis (hc) is sometimes seen in haematopoietic stem cell transplantation (hsct) patients and can on its own or by subsequent complications cause significant morbidity and even death. according to nci dictionary of cancer terms, it is defined as "a condition in which the lining of the bladder becomes inflamed and starts to bleed. the blood can be seen in the urine. symptoms include pain and a burning feeling while urinating, feeling a need to urinate often, and being unable to control the flow of urine. haemorrhagic cystitis may be caused by anticancer drugs, radiation therapy, infection, or being exposed to chemicals, such as dyes or insecticides" (nci dictionary of cancer terms https://www.cancer.gov/publications/ dictionaries/cancer-terms?cdrid= ). haematuria can be symptomatic or asymptomatic. it can be described as microscopic (not visible to the eye but detected on a dipstick and in the microscope) or macroscopic (red urine or visible blood or clots) (table . ). normally about million erythrocytes are excreted daily in the urine. this is equal to one to three erythrocytes per highpower field (magnification × ) under the microscope. haematuria is defined as abnormal presence of blood in the urine, i.e. more than three erythrocytes per high-power field in the micro-scope. to be confirmed as microscopic haematuria, two positive samples on consecutive days are needed. the haematuria can be visually detected (macroscopic) as red urine at levels as low as ml blood per litre urine. the visible blood does however not necessarily correspond to the degree of blood loss through the urine. red urine may also have other causes which will not be described here. cystitis is the term used to describe inflammation of the bladder. the inflammation can be caused by an infection or as a reaction to certain drugs or radiation therapy. the following symptoms may be seen in all types of cystitis: • urinary urgency and frequency • burning or stinging with urination or right after • pain, dysuria (painful urination), lower abdominal or supra-pubic pain • nocturia, when sleep is disturbed twice or more at night due to a need to urinate • urinary incontinence • general feeling of illness reported incidences of hc after hsct range between % and % depending on risk factors and use of preventive measures or not, but most materials describe an incidence between % and %. the pathogenesis leading to hc is not completely known but is likely to be multifactorial. the onset is seen either early, within the two first weeks after start of conditioning treatment, or late, more than weeks after hsct. conditioning treatment with chemotherapy, irradiation, cytopenia, viral infections due to immunosuppression and alloimmune reactions (immunisation by development of antibodies in response to an antigen, i.e. a protein from a donor, e.g. by receiving hsct or transfusion) may all contribute to hc in the posttransplant period. higher incidence of late-onset hc in hsct with unrelated donors, older patients, and in patients with graft versus host disease (gvhd) and thrombocytopenia does support the conclusion that the pathogenesis is multifactorial (de padua silva ). early-onset hc is usually a direct and immediate effect of the conditioning treatment. conditioning therapy for hsct often contains one or more alkylating agent. cyclophosphamide, ifosfamide, busulfan, melphalan and thiotepa are among the most commonly used drugs in conditioning regimens and the major drug-related cause of hc. use of other alkylating agents and etoposide may also increase the risk of hc. when cyclophosphamide or ifosfamide is metabolised in the body, it produces a metabolite called acrolein. acrolein will cause direct toxicity to the inner lining of the urinary tract, the urothelium. the degree of damage is dose dependent, and the toxicity may increase with previous or concomitant radiation therapy and if busulfan is included in the conditioning regimen together with cyclophosphamide. the time of duration that acrolein is exposed to the bladder also contributes to the degree of damage. for cyclophosphamide, the maximal concentration of active metabolites is reached after - h of oral or intravenous administration. most of the cyclophosphamide, - % of the dose, is excreted in the urine as metabolites, and up to % is excreted as intact drug (hassan and ljungman ) . in patients with decreased renal function, particularly in severe cases, decreased renal excretion may result in increased plasma levels of cyclophosphamide and its metabolites. this can cause increased toxicity. the following substances may also increase the concentration of toxic metabolites, possibly through inhibited breakdown or decreased renal excretion: allopurinol, cimetidine, hydrochlorothiazide and hiv protease inhibitors. when hc occurs more than weeks after hsct, a common cause in the immunocompromised host can be viral infection. viral particles are frequently identified from the urine of hsct recipients. of these, reactivation of the polyoma bk virus (bkv) is the commonest and most consistent risk factor for hc following hsct, as the virus is almost invariably present in the urine of patients with hc (leung et al. ) . the damaged urothelial cells provide a milieu for viral replication. immunosuppression leads to viral reactivation and causes viruria. however, the exact pathogenetic link between bkv and hc remains enigmatic. other viral agents such as adenovirus, cytomegalovirus (cmv) and other polyomaviruses similar to bkv may also but less often cause hc. alloimmunity after engraftment by attack from donor lymphoid cells against infected urothelial cells has not been confirmed as causing hc but may be an additional potential factor for development of this complication. the diagnosis of hc is confirmed by the presence of haematuria and symptoms of cystitis taking into account risk factors such as: in most cases of chemotherapy-induced hc with pre-engraftment onset and in polyomaviruria, the condition is self-limiting and the prognosis is good. if the viruria is caused by adenovirus, the prognosis is worse with the risk of progression to systemic adenovirus infection. in these cases early pharmacological intervention with antiviral drugs, e.g. cidofovir, is recommended. hyperhydration with forced diuresis, i.e. l/ m / h with the goal of a diuresis of > ml/h, during and until the day after administration of an alkylating agent is the most important preventive action. if the diuresis is insufficient, diuretics should be administered. the forced diuresis will not just dilute the urine but shorten the time of duration for acrolein exposure to the bladder and thus prevent the toxic effects. during the days of hyperhydration, the patient shall be closely monitored for fluid balance, including weight, at regular intervals. an electrocardiogram (ecg) should be taken, and approved, prior to start of treatment, and vital signs (blood pressure, pulse, oxygen saturation and respiratory rate) should be checked throughout the day in order to ascertain circulatory stability. electrolytes and renal function should be monitored by blood samples and electrolyte substitution given where required. a need for potassium substitution is not uncommon. the patient should also be assessed for any urinary or low abdominal pain or discom-fort. all assessments mentioned above should be performed at least every h. informing the patient about the treatment and treatment goals as well as the importance of reporting any symptoms of hc will help ensure that appropriate actions and early intervention can be applied without delay. for patients receiving cyclophosphamideor ifosfamide-based regimens, the drug mesna (sodium -mercaptoethanesulfonate) can be used as pharmacological prophylaxis, although the additional benefit in the hsct setting has not been scientifically proven in comparison with hyperhydration and forced diuresis. mesna binds to the toxic metabolite acrolein and forms a non-toxic compound. by additional actions mesna also reduces the forming of acrolein in the urine. the drug itself has low toxicity (mesna summary of product characteristics (spc) [in swedish]). in hsct conditioning with cyclophosphamide, the recommended dose of mesna according to the summary of product characteristics (spc) is % of the cyclophosphamide dose and the first mesna dose should be administered immediately prior to the cyclophosphamide. subsequent doses will then be given at , , and h after administration of cyclophosphamide (totalling % of the cyclophosphamide dose). it is important to adhere to the timing of mesna doses in order to ensure efficacy of the treatment. mesna treatment should be continued during the cyclophosphamide treatment period plus the time predicted for the metabolites to reach non-toxic levels. this will usually occur between and h after completed cyclophosphamide administration. this treatment schedule for mesna may however vary according to conditioning regimen and doses as well as to patient individual factors. an example of a checklist to be used during high-dose cyclophosphamide treatment is enclosed. bk virus-induced hc may be prevented by the administration of quinolones (e.g. ciprofloxacin) (dropulic and jones ) . although quinolones are not strictly antiviral, fluoroqui nolones are capable of inhibiting the helicase activity of bkv large t antigen (tag) protein, which seems to be crucial for separation of the double-stranded dna genome during replication of the virus (umbro et al. ) . there is currently no consensus regarding this prophylaxis because many patients with bkv do not develop hc. the fact that there is a general increase of multidrug-resistant microorganisms makes the use of this prophylaxis a matter for careful consideration. the first intervention will be hyperhydration with forced diuresis to prevent clot formation. hc is usually painful and analgesia should be administered. if the patient is thrombocytopenic, a higher threshold level for platelet transfusion and intensive platelet support should be applied, in particular in haematuria grades iii-iv. catheterisation and bladder irrigation with . % sodium chloride (normal saline) may be necessary to prevent clot obstruction. catheter insertion should be performed so that the risk of additional injury to the urothelium is minimised. treatment with bladder instillation of various compounds such as formalin, alum, silver nitrate, sodium hyalonurate, prostaglandins, gm-csf, fibrin glue, cidofovir, ciprofloxacin or ribavirin has been reported as effective, but experiences are still limited (carreras ) . if obstruction occurs, cystoscopy can be performed. selective embolisation of bladder arteries and catheterisation of both ureters to rest the bladder are actions that can be taken in severe cases. cystectomy remains the last resort if all other treatment attempts fail. in addition to actions mentioned above, treatment with systemic administration of palifermin, oral oestrogens and recombinant fviia may be used (carreras ) . systemic antimicrobial drugs, e.g. cidofovir, ciprofloxacin and ribavirin, can be started, if the hc is confirmed or likely attributable to adeno-or bk virus. decreased immunosuppression could be considered in particular in cases of relapsing viral cystitis. note that anticoagulants such as tranexamic acid and aminocaproic acid are contraindicated in hc due to risk of clot formation and retention! another type of treatment that has proven effective is hyperbaric oxygen (savva-bordalo et al. ) . the patient then receives % oxygen in a hyperbaric chamber but limited access to hyperbaric chambers, and the likely need and inability for the patient to move to another treatment unit often makes this intervention less of an option. during treatment with hyperhydration, the same need for close monitoring and assessments as in the prophylactic setting applies (see above). assess the need for platelet transfusion prior to catheterisation as well as after. blood transfusions may also be necessary with significant blood loss. standard monitoring for signs of infection, injury, pain, clot formation and other potential complications from the urinary catheter is important. in cases of bladder irrigation keeping the fluids for irrigation at ambient temperature may alleviate discomfort. complications of the irrigation can be prevented or minimised by close monitoring and recording of fluid balance. it is also important to maintain patient comfort by adequate pain management and general nursing interventions such as comfortable positioning and assistance with personal hygiene. the need for information and psychological support should be observed for both patient and family. since in particular viral hc may occur after discharge from the hospital, careful assessment of any signs and symptoms related to the urinary tract and that may indicate viral infection is just as important in the outpatient setting. assessments for treatment with high dose g/m or g/m cyclophosphamide (cy) sinusoidal obstruction syndrome/veno-occlusive disease sinusoidal obstruction syndrome (sos) is also known as veno-occlusive disease (vod) and is referred to as sos/vod hereafter. of the early complications that are considered to be of vascular endothelial origin, this is the most described. there are diagnosis and severity criteria (mcdonald et al. (mcdonald et al. , jones et al. ; deleve et al. ; mohty et al. ) , although the ebmt criteria proposed in (mohty et al. ) is expected to be further validated, and there is approved treatment available. careful monitoring of hsct patients allows early detection of sos/vod. treatment can then be started without delay, ultimately improving patient outcomes. from pre-transplant assessment to medical management and overall care of the patient, nurses thus have an essential role to play as part of a multidisciplinary team (wallhult et al. ). there are specific differences between the clinical presentation of sos/vod in adults versus in children which has not been reflected in the older diagnosis and severity criteria. for this reason, ebmt has also developed a classification for diagnosis and severity criteria for sos/vod in paediatric patients (corbacioglu et al. ). the information presented below is related to adults. for the paediatric population, please see original article and/or the vod learning programme on the ebmt website. when drugs used in haematopoietic stem cell transplant (hsct) conditioning regimens are metabolised in the liver, it results in toxic metabolites being produced by the hepatocytes. the metabolites trigger the activation, damage and inflammation of the endothelial cells lining the sinusoids (sinusoids being small capillary-like blood vessels in the liver). this trigger mechanism can start as soon as the conditioning treatment is administered. the activated sinusoidal endothelial cells release inflammatory cytokines, chemokines and the enzyme heparanase which break down the extracellular matrix that supports the structure of the sinusoids. the endothelial cells are then forced to round up, and gaps form between the cells. the gaps allow for red blood cells, white blood cells and other cellular debris to exit through these gaps in the sinusoid walls into the space of disse. (the space of disse is the perisinusoidal space that is located between the endothelium and the hepatocytes.) when cells and debris accumulate in this space, the sinusoids become narrower. due to the sinusoidal damage, endothelial cells can dissect off and embolise further downstream thus contribute to the narrowing. the damage also leads to an increase in the expression of tissue factor (tf) and plasminogen activator inhibitor- (pai- ). this coagulopathy causes an increase in clot formation and a decrease in the breakdown of clots. the deposition of fibrin and the clot formation will contribute to the narrowing of the sinusoids and may ultimately lead to hepatic sinusoidal obstruction. the result is sos/vod which is characterised by obstruction of the sinusoids, portal vein hypotension and reduced hepatic venous outflow. severe cases can progress to multi-organ dysfunction (mod)/multiorgan failure (mof) and death. sos/vod usually develops before day + after hsct with a peak incidence around day , but about - % of the sos/vod cases have a late onset, after day + . although relatively rare, sos/vod is one of the main causes of non-relapse, transplant-related mortality. a mean incidence of % (coppell et al. ) has been reported, but it varies with the diagnostic criteria, depending whether the seattle (mcdonald et al. (mcdonald et al. , or the slightly stricter baltimore criteria (jones et al. ) have been used. it will also depend on risk factors including intensity of conditioning regimen and type of transplant. after allo-hsct with myeloablative conditioning (mac), the incidence is approximately - %, but if reduced intensity conditioning (ric) is used, the incidence is < %. this is the same incidence as for auto-hsct. early-stage sos/vod, mild sos/vod, may not be particularly well-recognised since the symptoms are subtle, may not require treatment and spontaneously resolve within a few weeks. unrecognised sos/vod may however progress, sometimes very rapidly, into moderate or severe. severe sos/vod is associated with mod/mof and a mortality rate of %. the risk factors for sos/vod can be divided into patient-and disease-related and transplantrelated risk factors (mohty et al. ) . as mentioned above, the risk factors, as well as the clinical presentation of sos/vod, differ between the adult and the paediatric population, and the risk factors presented here are related to adults. the patient-and disease-related risk factors are: despite the fact that diagnostic criteria were developed in the s and have been used in clinical practice and research studies, it is often hard to identify early or mild cases of sos/vod before it progresses to a more severe form. some reasons are lack of sensitivity and specificity of the criteria, the dynamic manifestations that makes definition of the condition hard and that early signs and symptoms often are subtle and makes differentiation from other transplant complications difficult. given the poor prognosis of severe sos/vod, it is however vital to identify mild cases before they progress to moderate, with signs of hepatic injury and requiring more aggressive intervention, or further progress to severe sos/vod with mod/ mof. the most recent diagnostic criteria proposed by ebmt (mohty et al. ) are the same as the baltimore criteria (jones et al. ) for classical sos/vod with onset within the first weeks after hsct, but if sos/vod develops after day + , elevated serum bilirubin level is not always seen, why a modified version of the criteria can be used for diagnosis of late sos/vod (mohty et al. ) (table . ). the ebmt criteria will also better capture the dynamic manifestations of the disease and thus facilitate an early diagnosis as well as a more accurate assessment of severity. treatment can then be started at a stage with greater chance for treatment response. differential diagnoses will need to be excluded by assessing risk factors, symptoms and lab tests since liver dysfunction can also be seen in sepsis, viral infection, graft versus host disease (gvhd) and iron overload and as a side effect from many of the drugs used in the hsct setting. in addition to the signs and symptoms required for diagnosis haemorrhagic complications, thrombocytopenia with platelet refractoriness, pulmonary dysfunction, renal dysfunction and encephalopathy are "late" signs that can be seen in more severe cases of sos/vod. further it is worth noting that all symptoms are also observed in other conditions and that many other complications may coexist with sos/vod. examples of differential diagnosis for classical symptoms of sos/vod are listed in table . . when sos/vod is diagnosed, it is important to classify the severity grade in order to intensify the monitoring and identify patients that will need therapeutic intervention. the ebmt severity grading criteria (mohty et al. ) stress the importance of taking the time since the appearance of the symptoms into account. a rapid progression of symptoms, and in particular bilirubin kinetics (the rate of increase) with a doubling time of h, should be classified as a more severe grade than if symptoms develop more slowly over several days (table . ). the first strategy for prevention is to be aware of pre-existing risk factors and try and eliminate them as far as possible and potentially establish supportive or treatment measures prior to transplant. the patient-and disease-related risk factors, including hepatic, are often difficult or impossible to change, but the transplant-related risk factors should be carefully considered in the pre-transplant setting. no proven medical prophylaxis exists but sodium heparin, prostaglandin e , ursodeoxycholic acid and low molecular weight heparin have jones et al. ( ) been tried, although data about effectiveness remains inconclusive (carreras (carreras , . defibrotide, approved for treatment of severe sos/vod, has also been used as prophylaxis (dignan et al. ) , and one randomised study in children has shown a reduction in sos/vod incidence (corbacioglu et al. ). as soon as sos/vod is suspected, supportive therapy should be initiated. in mild cases of sos/ vod, close monitoring to detect progression and supportive management is often sufficient. the monitoring should include: for more details about nursing interventions see below. the only curative treatment for sos/vod is the drug defibrotide. defibrotide protects the endothelial cells, reduces inflammation and restores thrombo-fibrinolytic balance (richardson et al. ) . the recommended dose is . mg/kg body weight administered as a -h, i.v. infusion every h (to a total dose of mg/kg/day). recommendation for treatment duration is at least days but should continue until the symptoms and signs of severe vod resolve. defibrotide is generally well tolerated (keating ) but should not be used with products that affect platelet aggregation, e.g. nonsteroid anti-inflammatory drugs (nsaids), anticoagulant therapy or other products that increase the risk of bleeding. it is important to perform a risk assessment considering the risk factors mentioned above and to take baseline measurements including defining a threshold of > % for weight gain or what level and pattern of weight gain that represents a clinical concern. most baseline measurements will be standard for hsct patients, but in patients at high risk for sos/vod, assessments of abdominal girth, right upper quadrant (ruq) pain and inspection of sclera should be added. standard daily monitoring should include temperature, pulse, blood pressure, respiration rate and saturation. one of the most important daily monitoring aspects is an accurate fluid balance including intake, output and weight since fluid imbalance is one of the earliest signs of sos/vod. a fluid retention which does not respond to diuretics represents an early sign of endothelial damage. when performing abdominal girth measurement, it is advised to use a marked line for placement of the measuring tape and to choose one position (i.e. sitting/standing/lying) for the patient, to be used consequently. abdominal discomfort, tenderness, pain (in particular ruq pain) and inspection for collateral circulation and/or spiders should always be included in abdominal assessment. for nurses trained in palpation and percussion for ascites, bulkiness, liver margins and size these assessments should also be performed. sclera and skin should be assessed for bleeding/bruising and discoloration (jaundice). knowledge of the relevant reference ranges of daily laboratory values, particularly liver enzymes, serum bilirubin, blood count, electrolytes, urea and serum creatinine will enable early detection of significant change or trend in values since nurses are likely to take blood samples and see the results first and can alert medical colleagues. all findings should be precisely documented and any changes promptly reported. this is especially important in patients identified as high risk as early detection of sos/vod may affect the overall outcome. if sos/vod is suspected, the monitoring should be intensified and adequate vascular access established. in addition to standard lab tests, coagulation parameters should be performed daily. if possible, hepatotoxic drugs should be avoided and diuretics and pain medication administered as needed. electrolyte replacement may be necessary, and in case of thrombocytopenia or bleeding, blood products will be administered. if fluid restriction is enforced, it is important to know the smallest volumes that can be safely delivered. the patient may also need assistance to be comfortably positioned. when sos/vod has been diagnosed, the supportive care and monitoring will be further intensified including assessing for failure in respiratory, cardiac and renal function. defibrotide treatment will most likely be started, and patients in need for ventilatory support should be prepared for transfer to the intensive care unit (icu). patients should be informed and educated to notify the staff of any signs and symptoms that may need closer monitoring or intervention. in case sos/vod is diagnosed, both patient and family will need reassurance and support. other early complications of endothelial origin a number of early complications to haematopoietic stem cell transplantation (hsct) seem to be initiated by damage to the vascular endothelium. the most well defined and well described of these complications is sinusoidal obstruction syndrome (sos)/veno-occlusive disease (vod) described in a separate section of this chapter. other syndromes in this group have been named engraftment syndrome (es), diffuse alveolar haemorrhage (dah), idiopathic pneumonia syndrome (ips) and transplant-associated microangiopathy (tma). the similarities in their clinical manifestations and the lack of established diagnostic criteria often make determination of incidence and differential diagnosis difficult (soubani and pandya ; afessa et al. ) . although many times mild and with spontaneous recovery, these complications also share the risk for progression to multi-organ failure (mof)/multi-organ damage (mod) resulting in a poor outcome. ongoing research and efforts for better characterisation and treatment indicate that there will be future changes in terminology and diagnostic criteria, as well as interventions, for the early hsct complications mentioned here. several factors in the hsct setting activate the endothelial cells that line the blood vessels. contributing factors are the conditioning treatment and use of other drugs such as granulocyte colony-stimulating factor (g-csf) and calcineurin inhibitors (cni), e.g. cyclosporine-a, and microbial products translocated through mucosal barriers. the result is that fluid and proteins leak out of tiny blood vessels and flow into surrounding tissues. if unrecognised, this may lead to dangerously low blood pressure and subsequently mof and shock. the symptoms often appear around the time of neutrophil recovery, i.e. when the absolute neutrophil count (anc) increases to ≥ . × /l, which is why the complex process of engraftment may also play a role in activation of endothelial cell damage. the activation of the endothelial cells leads to further damage and inflammation by the release of pro-inflammatory cytokines. since the incidence of vascular endothelial syndromes is higher after allogeneic transplantation, alloreactivity (the immune response to non-self cells) is considered to play a role in activation and damage of endothelial cells. engraftment syndrome (es) es usually occurs after auto-hsct although described in allo-hsct as well, in particular when reduced intensity conditioning (ric) and cord blood (cb) have been used. due to lack of diagnostic criteria, the term es has been used as synonymous with capillary leak syndrome (cls), auto-aggression syndrome, peri-engraftment respiratory distress syndrome (perds), aseptic shock syndrome and autologous graft versus host disease (agvhd). although there are differences, their common denominator is that they share some or all symptoms that have been attributed to es. engraftment is defined as when the number of neutrophils in the patient's blood rises to an absolute neutrophil count (anc) of ≥ . × /l. peri-engraftment can be defined as the period within days of neutrophil engraftment. due to the diagnosis difficulties, there is no reliable incidence figure and numbers between % and % have been reported. there is also a lack of survival data. most cases are mild and respond well to corticosteroid therapy, but es may progress and lead to transplant-related mortality and decrease in overall survival. patients who require mechanical ventilation has a poor prognosis. a number of potential risk factors related to patient characteristics, disease, previous treatment, conditioning treatment, stem cell source and supportive drug treatment have been reported, but there is a lack of consensus which can in part be contributed to the lack of diagnostic criteria. changes in hsct practices with new drugs and alternate stem cell sources may impact the risk factors in the future. among the risk factors described are: • female gender • advanced age • no or little prior chemotherapy • previous use of bortezomib and lenalidomide in multiple myeloma patients • cord blood transplantation • cd + cell number and engraftment rate • g-csf treatment • amphotericin treatment • cyclosporine (cya) treatment • auto-hsct for amyloidosis, multiple myeloma, poems (polyneuropathy organomegaly endocrinopathy monoclonal protein and skin abnormalities) syndrome and autoimmune diseases there are two tools to aid diagnosis of es; the spitzer ( ) and the maiolino et al. ( ) diagnostic criteria. the clinical manifestations are divided into major or minor clinical criteria (table . ), but maiolino only has one major criteria, non-infectious fever. the timing of symptoms relative engraftment also differs between the two, where maiolino has a stricter timeframe from h before to any time after neutrophil recovery compared to spitzer's h after (table . ). however, in some patients others have described onset of symptoms from days before (for patients with poems) to days after engraftment, and in cases with more severe symptoms, the early symptoms may have been overlooked, why the clinical criteria sometimes could be used regardless of appearance of symptoms in relation to time for engraftment (chang et al. ). c-reactive protein (crp) is not used for diagnosis in either criteria, but a sudden and significant increase in the crp level has been found to support the diagnosis. early recognition of signs and symptoms is the most important aspect since there is no standard prophylaxis for es, although there is evidence that corticosteroids may prevent this complication. before treatment is initiated, other diagnoses such as infection, drug rash, diarrhoea associated with infection or medication and intravenous (i.v.)-related fluid overload should be excluded. broad-spectrum antibiotics should be used until infection is ruled out (cornell et al. ) . if cultures are negative, symptoms remain after - h of antibiotic treatment and other etiologies can be excluded, corticosteroid treatment can be initiated. methylprednisolone in doses of - mg/kg/ day i.v. are recommended until symptoms begin to subside. response to treatment is usually seen within - days. corticosteroids could then be switched to oral administration and should be slowly tapered. early intervention with steroids prevents progression to more severe manifestations, and in the vast majority ( %) of patients, there is then complete resolution in less than days. in cases with no response to steroid treatment after h, biopsies of affected organs may be necessary. if biopsies are performed for evaluation of diarrhoea, the findings may not be able to distinguish from gvhd. this does however not exclude es since overlap and coexistence with gvhd is possible. if a biopsy supports the es diagnosis treatment with additional immune suppressants should be started and continued until response. if the result of the biopsy is an alternative diagnosis, the patient should be treated accordingly. in addition to pharmacological treatment supportive care with i.v. fluids, with electrolyte supplement as needed, and oxygen therapy may be necessary depending on the symptoms. in cases of encephalopathy or severe es with mof, plasma exchange may be considered (yeoung-hau and syed ). daily nursing assessments are critical in early detection and diagnosis of all complications to hsct. the patient's general well-being should be assessed, and listed below are the nursing assessments that should be carried out frequently, the findings that could indicate es and actions that can be taken in order to detect or rule out the es diagnosis (table . ). all findings should be monitor frequently, and if symptoms of pulmonary dysfunction, e.g. dyspnoea, tachypnoea, change in breathing pattern, chest pain or cough, are present, a chest x-ray or pulmonary ct scan may be performed. in order to ensure adequate oxygenation, administration of oxygen therapy may be necessary weight and fluid balance assess the patient's weight daily and perform calculation of fluid balance at least once daily to note any trends. if oedema, ascites or other symptoms of fluid retention occurs diuretics should be administered as ordered skin perform assessment at least daily and note any rashes. if a rash is detected, review the patient's medication chart for medication that may cause drug rash jaundice and yellow sclera are signs of liver dysfunction and bilirubin levels should be checked stools monitor frequency and consistency and obtain cultures and test for clostridium difficile in cases of diarrhoea in order to rule out infection. pale stools are a sign of liver dysfunction and bilirubin levels should be checked (continued) documented and any abnormalities promptly reported to the treating physician. if steroid treatment is started, the patient should be assessed for possible side effects such as hyperglycaemia and insomnia. blood glucose should be monitored daily. idiopathic pneumonia syndrome pulmonary complications (pcs) are the leading cause of patients' admission to intensive care unit (icu) after hsct. pc can be divided into infectious or non-infectious. one of the non-infectious pcs is idiopathic pneumonia syndrome (ips). for the purpose of this chapter, ips will be defined and described according to the definition by the american thoracic society (panotskaltsis-mortari et al. ) as "an idiopathic syndrome of pneumopathy after hsct, with evidence of widespread alveolar injury and in which an infectious etiology and cardiac dysfunction, acute renal failure or iatrogenic fluid overload have been excluded". the alveolar injury is a result from the release of proinflammatory cytokines during engraftment increasing alveolar permeability and causing diffuse alveolar or interstitial infiltrates. ips also includes a subset of diagnoses of primary lung injuries classified according to the anatomical sites of inflammation. they can either be related to the pulmonary parenchyma (e.g. acute interstitial pneumonitis and acute respiratory distress syndrome (ards)), the airway endothelium (e.g. bronchiolitis obliterans syndrome (bo)), the vascular endothelium (e.g. different forms of es (perds, cls)) or be unclassifiable. other less frequent non-infectious pcs have also been identified. none of these entities will be described here. pcs are common in hsct recipients and a major cause of morbidity and mortality. ips is more often seen in patients undergoing allogeneic hsct with a mean estimated incidence of - % ( % in auto-hsct) (chi et al. ). the overall outcome is different between auto-and allo-hsct recipients, and where ips in patients that have undergone auto-hsct usually has a favourable prognosis, the mortality is - % in the allo-setting (carreras ) . ips has a progressive nature, and patients with progression to respiratory failure and need for mechanical ventilation have a very poor prognosis with % mortality. for ips the following risk factors have been identified (diab et al. ): • older age • low performance status (karnofsky score) • high-intensity conditioning regimen • total body irradiation (tbi) • allo-hsct • acute graft versus host disease (agvhd) • malignant disease pre-transplant pulmonary function abnormalities have also been associated with early respiratory failure and mortality (chien et al. ) . educate the patient about signs and symptoms of es and explain why it is important to report any symptoms without delay. explain actions taken in diagnosis and management of es and provide emotional support to both patient and family thoele ( ) the most common signs and symptoms are fever, non-productive cough, rales, dyspnoea, tachypnoea and low saturation with an increasing need for oxygen support. the diagnosis will be based on alveolar injury confirmed clinically, radiologically and/or functionally. x-ray will reveal diffuse pulmonary infiltrates. infection must have been ruled out by negative cultures and tests in bronchoalveolar lavage (bal) or lung biopsies (zhu et al. ) , and there should be no evidence of cardiac dysfunction, acute renal failure or treatment-related fluid overload. it is however considered possible that some cases of ips may be caused by an unidentified underlying infection since infections may lack typical signs and symptoms in the neutropenic patient. the ips diagnosis can thus be supported by lack of improvement despite broad-spectrum antibiotics and other antimicrobial drugs. the typical onset will be around day + , but ips may also present later after hsct, why it is important to be alert for this complication also after discharge from the hospital, in the outpatient setting. there are no standard guidelines for diagnosis and evaluation of pc after hsct, but the course of illness should be considered when differential diagnoses are to be excluded. when symptoms occur, ips may rapidly progress to pulmonary dysfunction requiring mechanical ventilation. for patients at risk for ips, careful consideration of treatment options pre-and posttransplant such as avoiding conditioning with tbi or highintensity regimens and choice of gvhd prophylaxis may be beneficial. monitoring of pulmonary function and symptoms after transplantation will enable prompt intervention. in patients with decreased lung function prior to hsct and suspected lung injury in the posttransplant setting, close collaboration with pulmonary specialist or the intensive care team may prevent progression of pulmonary dysfunction (elbahlawan et al. ). beyond supportive care, there is no proven treatment for ips. in auto-hsct patients, corticosteroids can be effective, but this is usually not the case for allo-transplanted patients, irrespective of steroid dose. studies with etanercept, a tnf-αbinding protein, given in combination with corticosteroids have reported improved pulmonary function in patients with ips following allogeneic hsct and may be considered (carreras ) although a small but later study (yanik et al. ) could not confirm the benefit of this treatment. the close monitoring and daily nursing assessments that apply for all hsct patients should be employed. depending on risk factors, extra attention may be needed to early and subtle symptoms of pulmonary dysfunction, such as decrease in saturation, shortness of breath and cough. monitoring of daily weight and fluid balance, with administration of diuretics if necessary, will prevent and rule out fluid overload. several different tests and examinations may be performed to establish or rule out the diagnosis of ips. sputum cultures and laboratory tests, such as polymerase chain reaction (pcr) for mycoplasma, and serum galactomannan for aspergillus may need to be obtained, and chest x-ray or computed tomography (ct) scan performed to rule out infection. in case a bal, with or without transbronchial biopsy, will be performed, information to the patient and preparation prior to the procedure as well as support both before and after and post procedure monitoring is important. the bal may add substantial discomfort, in particular to an already seriously ill patient. other lung function tests may also be repeated, for comparison with pretransplant results. when corticosteroids are administered, the blood glucose levels should be followed daily and the patient should be informed of and assessed for other side effects, e.g. insomnia. oxygen therapy may need to be administered and noninvasive positive pressure ventilation necessary. respiratory difficulties generate anxiety, and the patient should be offered psychological support as well as assistance with positioning and breathing techniques and exercises. medication for anxiety may be necessary. referral to a physiotherapist, respiratory therapist or other staff with expertise in pulmonary diseases should be made for advice on tools and exercises that may help the patient to maintain pulmonary function and prevent worsening of the condition. if the condition shows no signs of improving, the patient should be prepared for transfer to the icu. identification of patients at risk, prompt intervention to signs and symptoms of pulmonary dysfunction and close collaboration within the team will increase the chances of a positive outcome. diffuse alveolar haemorrhage diffuse alveolar haemorrhage (dah) is a noninfectious pulmonary complication associated with haematopoietic stem cell transplant (hsct) and other causes (park ) . it is differentiated from idiopathic pneumonia syndrome (ips) through confirmation of pulmonary haemorrhage by bronchoscopy and bronchoalveolar lavage (bal). the bleeding can be either insidious, causing a gradual pulmonary dysfunction, or a more acute bleeding into the alveolar space. damage to the alveolar-capillary barrier from conditioning treatment and the engraftment process with recovery of neutrophils leads to entry of blood into the alveolar space. an approximate incidence of around % up to %, with a mortality rate ranging between % and %, has been reported for dah in hsct recipients (afessa et al. ; majhail et al. ; carreras ) . the incidence is similar between auto-and allo-hsct. the implication of prognostic factors has not been well studied, but early-onset dah (within the first days after transplant) in patients undergoing auto-hsct has a favourable prognosis. risk factors for the development of dah in hsct recipients include: • older age • total body irradiation (tbi) • myeloablative conditioning (mac) regimens • acute graft versus host (agvhd) disease dyspnoea, dry cough and fever are the most common complaints. haemoptysis is rarely observed in hsct recipients. hypoxemia may be present and diffuse or focal interstitial or alveolar infiltrates can be found on chest x-ray or computed tomography (ct) scan. with such findings, bronchoscopy with bal and transbronchial biopsy is indicated although performing these invasive tests in patients with severe illness, and unstable respiratory status is a challenge. the diagnosis is based on bal findings which become progressively more blood stained, indicating blood in the alveoli. other causes, such as heart failure and fluid overload, should be excluded. infection needs to be ruled out by obtaining relevant cultures. presence of hemosiderin-laden macrophages in bal fluid is not diagnostic for dah but may support the diagnosis. it is often very difficult to differentiate dah from ips and the es form of respiratory distress (perds). ips is more common in allo-hsct, after engraftment, and does not respond to corticosteroids and has a more progressive nature. in perds the majority of patients do not have bal findings becoming progressively bloodier. the mean onset of dah has been reported on day after transplant and days after absolute neutrophil count (anc) recovery. reversal of some risk factors, e.g. choice of conditioning treatment, may be possible, but otherwise no prophylaxis exists. high-dose corticosteroids, using methyl prednisolone in doses of - mg every h for - days followed by slow tapering, is considered first-line treatment even if efficacy can be questioned. with early diagnosis and treatment with steroid therapy, respiratory failure can often be prevented. noninvasive ventilation may decrease mortality although the majority of patients with dah require mechanical ventilation, and sepsis and mof/mod will cause death in a large proportion of patients (rabe et al. ) . other pharmacological therapies, as well as plasma exchange, have been tried for treatment of dah. recombinant factor viia (rfviia) has been administered and achieved temporary control of bleeding. tranexamic acid or the tnfα-inhibitor etanercept have been used in addition to corticosteroids but have not proved to be effective. transfusion of platelets and red blood cells (rbc) may be necessary. patients need frequent monitoring for early detection of any pulmonary symptoms. respiration rate and saturation should be assessed together with temperature and other standard assessments. if cough is noted, this should be reported to the team and the treating physician. cultures and blood tests may be necessary to rule out infection. cultures should be performed according to signs and symptoms, but screening cultures can be collected to possibly enable detection of occult infection. the patient's circulatory status and fluid balance should be controlled by monitoring pulse, blood pressure, weight and input and output. the patient should be instructed to report all symptoms, and if bal and lung biopsy will be performed, patient information and support throughout the whole procedure is vital. administration of transfusions, oxygen therapy and non-invasive ventilation should be performed as ordered and since dyspnoea and other breathing difficulties are associated with a great deal of anxiety patient support, sometimes with pharmacological treatment, is crucial. proper positioning together with breathing exercises using appropriate breathing technique may alleviate some discomfort. during high-dose corticosteroid treatment, blood glucose should be monitored, and it is important to be alert to steroid-related changes in the patient's mental status. transplant-associated microangiopathy (tam) transplant-associated microangiopathy (tam) is also known as haematopoietic stem cell transplantation (hsct)-associated thrombotic microangiopathy (ta-tma). in this text, the term tam is being used. tam is characterised by microangiopathic haemolytic anaemia with schistocytes (fragmented red blood cells) and thrombocytopenia from platelet consumption. these symptoms are due to endothelial dysfunction causing small vessels thrombosis in the microcirculation. tam is a multi-visceral disease most often affecting the kidneys, but pulmonary, gastrointestinal and central nervous system (cns) involvement can also be seen. complement system dysregulation plays an important role in the severity of tam. defects in the complement system lead to formation of the lytic complex c b- . this complex can be detected in blood, and an increased level will support the tam diagnosis. the incidence will vary with the criteria used to diagnose tam. in retrospective data, the incidence is approximately % in auto-hsct, % has been reported in allo-hsct (carreras ) , whereas one prospective study has shown an incidence close to % (jodele et al. ) . conditioning intensity, myeloablative (mac) versus reduced (ric), has not shown any difference in incidence in allo-hsct. as with many early complications in hsct prompt recognition of early signs and symptoms with early diagnosis and intervention will increase the chances of a positive outcome. cases of mild tam where calcineurin inhibitor (cni), e.g. cyclosporine, tacrolimus and sirolimus, is the cause generally have a good prognosis if cni can be discontinued. if tam is not related to cni treatment, the prognosis is worse due to lack of effective treatment options. exact figures for mortality rate are difficult to establish, but in patients with tam and multi-organ involvement, the mortality is as high as > %. patients surviving tam are as a consequence at greater risk for chronic kidney disease (ckd) and hypertension later on. use of total body irradiation (tbi) in conditioning treatment, cni, graft versus host disease (gvhd), infections (e.g. cytomegalovirus (cmv) and fungal infections) and unrelated donor transplant (in particular if mismatched) are all considered risk factors or triggers for tam, although reported data is conflicting (nadir and brenner ; rosenthal ). tam usually has an onset between and months after hsct but can be seen both earlier and later. several slightly different criteria for diagnosis of tam are being used (sahin et al. ) . see adapted table . . the diagnosis is difficult but can be confirmed with a biopsy tissue sample although this invasive test may not always be an option for the seriously ill hsct recipient. tam has clinical similarities with idiopathic thrombotic thrombocytopenic purpura (ttp), and laboratory testing for the von willebrand factor regulator adamts can be performed to support the diagnosis. in classical ttp, there is a severe deficiency, while no significant decrease of adamts is seen in tam (graf and stern ) . renal tam should be suspected if the patient requires higher doses of antihypertensives than would be expected considering the situation and concomitant and/or nephrotoxic medication. example of a differential diagnosis is virusrelated nephropathy. symptoms such as tachycardia, chest pain and hypoxemia should lead to suspicion of lung involvement and pulmonary hypertension. the diagnosis can be supported by findings of cardiomegaly on chest x-ray, pericardial effusion on transthoracic echocardiography and blood tests. intestinal tam presents with the same symptoms as acute gvhd (agvhd), abdominal pain, diarrhoea, vomiting and gastrointestinal bleeding. the symptoms can also be mistaken for infectious colitis, but in tam the cause of the bleeding is ischemia in the bowels due to the microangiopathy. in addition to the general diagnostic criteria, specific criteria for gastrointestinal tam have been proposed. besides the clinical symptoms, x-ray findings with signs of ileus and thick mucosal wall and endoscopy with mucosal erosions and haemorrhages are included in the gastrointestinal tam diagnostic criteria, but the only definite diagnostic test is a biopsy tissue sample. as a result of generalised vascular injury in tam, polyserositis with pericardial and pleural effusion and ascites can occur. it can easily be mistaken for gvhd, but where gvhd more seldom is associated with microangiopathic anaemia, proteinuria and hypertension, these symptoms are common in tam. no specific prophylaxis exists, so vigilant monitoring of clinical signs and symptoms is neces-sary. cni concentration in blood, lactate dehydrogenase (ld or ldh) and serum creatinine should be closely followed, i.e. two to three times/week, with laboratory testing. additional blood tests with peripheral blood smear, haptoglobin and direct and indirect antiglobulin tests (dat and iat) should be performed if an increase is seen in cni, ld and creatinine levels. there is currently no established treatment for tma but supportive measures should always be taken. traditionally the first step is to discontinue cni, despite paucity of evidence for this action. it is also important to treat infections, gvhd and jodele et al. ( ) hypertension. changing to other gvhd prophylaxis and use of antimicrobial drugs should be based on a risk-benefit assessment where, for example, nephrotoxicity is considered. administration of diuretics may be necessary to treat fluid and sodium retention due to steroid treatment. vasodilators and renin-angiotensin antagonists may also be used to treat hypertension. it is recommended to restrict platelet transfusion in microangiopathic disease, but this is often impossible due to the need to prevent bleeding complications. a potential treatment for tma is eculizumab. eculizumab stops the complement-activating cascade preventing formation of c b- . this leads to hampering of the intravascular haemolysis. eculizumab has shown effect when started early after diagnosis (jodele et al. ) . monitoring for effect by following serum concentration levels is important, and dose adjustments may be necessary to reach and maintain the desired therapeutic levels and effect. in a small number of cases, successful treatment with rituximab and other monoclonal antibodies has been reported. treatment attempts have also been made with defibrotide at the same dosing as approved for treatment of severe sinusoidal obstruction syndrome/veno-occlusive disease (sos/vod) but with variable results. total plasma exchange (tpe) has been tried due to the clinical similarities between tam and ttp, but where ttp can be successfully treated with tpe, it is not recommended for tam due to poor response rates. careful assessments will facilitate early diagnosis of, or ruling out, tma and thus improves the outcome. close monitoring of vital signs and being alert to any changes or trends is standard. keeping track of fluid balance and weight is equally important. blood pressure should be kept below / in adult patients (jodele et al. ) . the patient's urine should be monitored for proteinuria and the patient instructed about what abnormal findings and symptoms to look for and to notify staff of any discomfort including signs of gastrointestinal bleeding. if invasive tests such as biopsies are to be performed, proper preparation and support is vital. if pharmacological treatment with eculizumab is started, serum level concentration needs to be followed. treatment with rituximab and defibrotide should be administered as ordered, and the patient should be monitored accordingly for effect and side effects. since the onset of tam can occur after discharge from the transplant unit, it is important to be observant to symptoms and consider this diagnosis even in the outpatient setting. open access this chapter is licensed under the terms of the creative commons attribution . international license (http://creativecommons.org/licenses/by/ . /), which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the creative commons license and indicate if changes were made. the images or other third party material in this chapter are included in the chapter's creative commons license, unless indicated otherwise in a credit line to the material. if material is not included in the chapter's creative commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. outcome of diffuse alveolar hemorrhage in hematopoietic stem cell transplant recipients risk factors and outcome of pulmonary complications after autologous hematopoietic stem cell transplant emerging evidence of the pathobiology of mucositis implementation of a standardised protocol for prevention and management on oral mucositis in patients undergoing haematopoitiec stem cell transplant ebmt-esh handbook on haematopoietic stem cell transplantation how i manage sinusoidal obstruction syndrome after haematopoietic cell transplantation et al ngraftment syndrome after 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allogeneic hematopoietic stem cell transplant recipients with indwelling central venous catheters at a comprehensive cancer center american thoracic society committee on idiopathic pneumonia syndrome. an official american thoracic society research statement: noninfectious lung injury after hematopoietic stem cell transplantation: idiopathic pneumonia syndrome diffuse alveolar hemorrhage on behalf of the esmo guidelines committee. management of oral and gastrointestinal mucosal injury: esmo clinical practice guidelines for diagnosis, treatment, and follow-up united kingdom oral care in cancer guidance: second edition. www.ukomic.co guidelines for the assessment of oral mucositis in adult chemotherapy, radiotherapy and haematopoietic stem cell transplant patients severe respiratory failure due to diffuse alveolar hemorrhage: clinical characteristics and outcome of intensive care drug safety evaluation of defibrotide surviving sepsis campaign: international guidelines for management of sepsis and septic shock: hematopoietic cell transplantationassociated thrombotic microangiopathy: a review of pathophysiology, diagnosis, and treatment chapter : infections after hsct. in: ebmt-esh handbook royal college of physicians. national early warning score (news) standardising the assessment of acuteillness severity in the nhs clinical practice guidelines for the prevention and treatment of cancer therapy -induced oral and gastrointestinal mucositis diagnostic criteria for hematopoietic stem cell transplant-associated microangiopathy: results of a consensus process by an international working group an overview of hematopoietic stem cell transplantation related thrombotic complications flossing for the management of periodontal diseases and dental caries in adults clinical effectiveness of hyperbaric oxygen therapy for bk-virus-associated hemorrhagic cystitis after allogeneic bone marrow transplantation severe sepsis and septic shock management and performance improvement the third international consensus definitions for sepsis and septic shock (sepsis- ) the spectrum of non-infectious pulmonary complications following hematopoietic stem cell transplantation engraftment syndrome following autologous hematopoietic stem cell transplantation programme group sepsis. care program severe sepsis and septic shock -early identification and initial intervention [för svenska infektionsläkarföreningen "pro sepsis" programgrupp sepsis. vårdprogram svår sepsis och septisk chocktidig identifiering och initial handläggning engraftment syndrome in hematopoietic stem cell transplantations immune reconstitution after allogeneic hsct. in: ebmt-esh handbook on haematopoietic stem cell transplantation possible antiviral effect of ciprofloxacin treatment on polyomavirus bk replication and analysis of non-coding control region sequences veno occlusive disease (vod) learning programme management of veno-occlusive disease: the multidisciplinary approach to care palliative adult network guidelines double-blind placebocontrolled trial of soluble tumor necrosis factor receptor: enbrel (etanercept) for the treatment of idiopathic pneumonia syndrome after allogeneic stem cell transplantation: blood and marrow transplant clinical trials network protocol brain teaser: encephalopathy after stem cell transplantation incidence, risks, and outcome of idiopathic pneumonia syndrome early after allogeneic hematopoietic stem cell transplantation acknowledgement european oral care in cancer group (eocc). key: cord- -lmly rd authors: renaud, christian; englund, janet title: respiratory syncytial virus and human metapneumovirus infection in transplant recipients date: - - journal: transplant infections doi: . / - - - - _ sha: doc_id: cord_uid: lmly rd respiratory viral infections due to respiratory syncytial virus (rsv) and human metapneumovirus (hmpv) cause infections in immunocompromised transplant patients ranging from mild upper respiratory infections to severe lower respiratory tract disease with respiratory failure. these viruses are more readily diagnosed due to improvements in sensitive molecular diagnostic methods. the epidemiology of rsv and hmpv is similarly becoming more readily appreciated in hematopoietic stem cell transplant (hsct) patients of all ages as well as solid organ transplant (sot) patients, with lung transplant recipients having evidence of more frequent and severe complications related to these viruses. rsv and hmpv infection typically but not always present with upper respiratory signs and symptoms that progress to lower respiratory tract disease. treatment options for rsv are limited, with aerosolized, intravenous, and oral ribavirin all studied in hsct and lung transplant patients. no antiviral therapy for the treatment of hmpv is available, although ribavirin has shown some effectiveness in vitro. new antiviral agents including rsv fusion inhibitors and nucleoside analogs are being developed, with some under clinical evaluation. respiratory viruses may cause serious morbidity and mortality in the immunocompromised host, and the transplant recipient appears particularly vulnerable. the impact of infection with respiratory viruses and the subsequent development of severe lower respiratory tract disease has been increasingly appreciated as respiratory viruses become more readily detectable. respiratory syncytial virus (rsv) and human metapneumovirus (hmpv) are among the best-documented respiratory viruses causing a wide range of respiratory disease in transplant recipients, ranging from asymptomatic shedding to fatal respiratory failure. understanding the epidemiology and clinical characteristics of rsv and hmpv permits clinicians to intervene pretransplant, provide appropriate infection control and prevention measures, potentially treat patients, and manage immunosuppressive therapy. severe respiratory disease in the seriously immunocompromised host in the mid-twentieth century was originally attributed to infection with opportunistic pathogens such as gram-negative bacteria, fungi, pneumocystis jiroveci , and mycobacteria, as well as cytomegalovirus (cmv) and adenovirus. in the later twentieth century, episodes of acute upper respiratory infection (uri) and lrti without an identifi ed etiology were often considered to be "idiopathic" pneumonia or attributed to regimen-related toxicity or acute respiratory distress syndrome (ards). in the classic study of non-bacterial, non-fungal pneumonias in allogeneic hematopoietic stem cell transplant (hsct) recipients, % of the episodes of pneumonia remained undiagnosed. the overall mortality rate associated with idiopathic pneumonia in these hsct recipients was %, a strikingly high fi gure [ ] . the potential morbidity of rsv infections was fi rst recognized in immunocompromised children in the s, more than a decade earlier than in immunocompromised adults [ - ] . the recognition of respiratory viruses as an important clinical problem likely refl ects the increasing number of severely immunodefi cient patients, more aggressive attempts to identify the cause of respiratory illness in high-risk patients, and the increasing ability of clinical virology and pathology laboratories to identify respiratory viruses in clinical specimens. although uncommon or atypical pathogens may be responsible for respiratory disease in the transplant recipient, the same viruses that cause typically mild but acute respiratory illness in the general population are responsible for hospitalizations in persons of all ages with underlying medical conditions [ ] . these same viruses are also a common cause of respiratory disease in transplant recipients [ - ] . with the widespread availability of sensitive and reliable molecular diagnostic methods, rsv and hmpv have been detected worldwide in transplant recipients and shown to be common causes of respiratory disease in the immunocompromised host [ , ] . in both the general population as well as in transplant recipients, rsv and hmpv may produce a wide constellation of clinical syndromes ranging from the common cold to bronchiolitis to severe pneumonia, but in contrast to the general population, rsv and hmpv may signifi cantly impact the morbidity and mortality of the transplant recipient. rsv was fi rst identifi ed in and became appreciated as a major cause of epidemic bronchiolitis and pneumonia in young children in the s [ , ] . hmpv was fi rst identifi ed in by molecular techniques in symptomatic children by van den hoogen et al. as a paramyxovirus causing bronchiolitis and uri in children [ ] . both viruses are classifi ed within the pneumovirinae subfamily of the paramyxoviridae family of non-segmented, negative-strand, enveloped rna viruses [ ] . hmpv belongs to the metapneumovirus genus whereas rsv is a member of the pneumovirus genus. both viruses are highly pleomorphic and their sizes vary from to nm. the rsv and hmpv genomes are approximately - kb in length and closely resemble each other, excluding a few differences in the order of the genes and the absence of the non-structural genes (ns and ns ) from hmpv genome. the remaining eight genes code for nine proteins present in both viruses: the nucleoprotein (n protein), the phosphoprotein (p protein), the matrix protein (m protein), the fusion glycoprotein (f protein), the putative transcription factor (m - protein), the rna synthesis regulatory factor (the m - protein), the small hydrophobic glycoprotein (sh protein), the attachment glycoprotein (g protein) and the viral polymerase (l protein). the rna core of the virion is associated with p, n, l, m - , m - proteins, surrounded by m protein and covered by a lipid envelope. f is the most highly conserved of the envelope glycoproteins within each virus and between rsv and hmpv. the fusion glycoprotein is essential in promoting attachment and fusion of the virus with the cell membrane during viral entry. the fusion protein is the target of many vaccines under development as well as that of monoclonal antibodies such as palivizumab, which is used to prevent rsv disease in preterm infants. by contrast, the g gene is the most variable. whole genome analysis of both rsv and hmpv has shown the existence of two genotypes, a and b. in hmpv, those two major genetic groups are further divided into subgroups a , a , b , b based upon the sequence variability of the g and f genes. subgroup a is again divided into a a and a b. rsv and hmpv infections produce both humoral and cellular immune responses. humoral immunity protects against reinfection while cellular immunity controls established infection and terminates viral shedding. protective immunity in immunocompetent hosts is thought to be relatively shortlived. both viruses interfere with the host's innate immune system resulting into incomplete clearance and partial immunity. prompt and accurate identifi cation of respiratory viral pathogen is critically important in the transplant recipient because it enables specifi c infection control precautions to be instituted, the initiation of specifi c antiviral therapy, impacts the use of immunosuppressive therapy, and potentially affects whether transplantation should proceed [ - ] . furthermore, identifi cation of a respiratory viral pathogen can assist in avoiding unnecessary therapy, procedures, and surgical procedures (such as open lung biopsy), as well as assist in the identifi cation of a potential cluster or epidemic of infections within the medical unit, hospital, or community. in hospitalized adults (both immunocompromised and immunocompetent), rapid viral diagnosis has been shown to reduce mortality and decrease the length of hospital stay and total cost [ , ] . laboratory diagnosis of respiratory viruses including rsv and hmpv has evolved considerably; adequate specimen collection is still essential for the successful identifi cation of viruses in clinical samples. newer types of nasopharyngeal swabs have shown improved viral diagnostic sensitivity compared to previous swabs, with similar sensitivity to nasal washes when using sensitive molecular methods in patients [ , ] . for example, nylon fl ocked swabs and foam swabs increase cell capture within the swab and then release into the transport media, increasing viral recovery [ , ] . nasal wash or aspiration methods are superior for isolation of viruses by culture and increase the sensitivity of culture, antigenic assays and quantitative molecular assays [ ] . nasal washes are well tolerated in cooperative adults and offer the advantage of visualizing the quality of the specimen. bronchoalveolar lavage remains the specimen of choice to diagnose lower respiratory tract infections because of the ability to simultaneously test for potential co-pathogens such as fungi, pneumocystis jiroveci , and bacteria, as well as to document viral infection in the lower airways. discordance in viral detection between upper and lower respiratory tract samples have been described with both viruses; negative upper tract and positive lower tract specimens in immunocompromised patients are possible but more discordance has been noted for hmpv compared with rsv. molecular diagnosis of rsv and hmpv is faster and more sensitive than viral culture or antigen detection, and most laboratories currently use commercial or in-house molecular assays to detect rsv and hmpv (table - ) . many genes have been targeted to detect rsv, including the n, f and l genes, with similar genes also targeted to detect hmpv. many rapid assays have been approved including some highly multiplexed respiratory panels allowing detection of rsv and hmpv as well as many other respiratory viruses and bacteria. several different primer sets may be utilized simultaneously in the reaction mix, and the virus identifi ed by the size of the amplicon or following hybridization with a virus-specifi c probe. some commercial assays are very rapid and require minimal technical expertise, with only - h of turn around time [ , ] . some laboratories have developed quantitative assays using hydrolysis probe technology with standard curves to help understand the signifi cance of positive results and to follow viral loads under therapeutic management [ - ] . no quantitative commercial assays are yet available. molecular assays have also been used to detect viral rna from blood/serum as a prognostic marker [ ] . unlike molecular methods, isolation of virus by culture confi rms the presence of a complete infectious unit capable of further multiplication. positive culture results may be obtained with as little as a single infectious virion, below the threshold of detection for most other detection methods, including some nucleic acid amplifi cation test (naat) methods . another advantage of viral culture is that multiple viruses may be identifi ed from a single sample and viruses can grow independently of point mutations that could potentially create false negative results by naat. the major limitations of viral isolation include the time, expense, and expertise required for virus isolation. a variety of cell lines can be used to grow rsv (hep- , a , rhmk) or hmpv (llc-mk , vero), and detection by cell culture can also be accomplished using several types of cells together, such as the r-mix cells (mixture of mink lung cells and a cells) [ ] (diagnostic hybrids, athens, oh). centrifugation combined with viral antigen detection methods permits more rapid diagnosis [ ] . rsv-and hmpv-specifi c monoclonal antibodies have been used for immunofl uorescence (ifa) techniques either directly on respiratory specimens or in cell culture [ , ] . the sensitivity of ifa is lower than that of naat for detection of rsv and hmpv. however, results can be reasonably fast, the method is relatively inexpensive, and importantly, this method also confi rms that an appropriate specimen has been properly obtained by looking at ciliated epithelial cells. sensitivity of ifa when performed by an experienced laboratory is as high as - % of the samples positive by pcr-at least in children [ ] . ifa can detect viruses that would be missed by naat because of point mutations. ifa positivity also has good clinical correlation while low grade naat positivity can be detected for longer periods of time with unclear signifi cance and transmissibility. enzyme immunoassays (eias) and rapid antigenic diagnostic tests ( radts) are commercially available for rsv and to a lesser extent for hmpv. these assays lack sensitivity and/or specifi city and are not recommended in transplant populations. more than one diagnostic method should be used, since no method is perfect. molecular assays have the greatest sensitivity-although perhaps at times can perhaps be too sensitive. paradoxically, rare point mutations causing mismatches have been described causing false negative results of naat in transplant units. viral cell culture requires time, is becoming less available in laboratories, and is more expensive and less sensitive, although it can catch those strains with mismatches and provide information on viral replicative nature while receiving treatment. further characterization of rsv and hmpv strains obtained from culture or directly from the clinical specimen is frequently desirable. antigenic differences among virus strains isolated from different geographic locations or at different times may also be examined. pools of monoclonal antibodies and "rna fi ngerprinting" have been used in the analysis of rsv strains in nosocomial outbreaks [ , ] but direct sequencing of the f and g glycoproteins is more commonly utilized [ , ] . next-generation sequencing is a very promising tool to characterize rsv or hmpv strains during severe infection. this could provide information on phylogenicity to identify outbreaks and also detect mutations that could be associated with antiviral or monoclonal resistance as well as increased virulence. some human gene alleles in the human genome may increase rsv severity in infants, but little is known yet on these genomic variations in transplant recipients. next-generation sequencing has the potential to provide information on the virus and the human genomes simultaneously. rsv is well known to cause annual winter outbreaks in the community ( figure - ). surveillance studies of respiratory viruses from transplant centers have established the high frequency and the signifi cant clinical impact of respiratory viral infections in hsct recipients overall [ - , , ] as well as the relative importance of rsv in terms of morbidity and mortality (table - ). a retrospective review conducted at the children's hospital of philadelphia revealed a [ ] . one of the earliest studies demonstrated fatal rsv pneumonia in four of immunocompromised adult hsct and solid organ transplant recipients with rsv infection [ ] . most studies conducted in the twentieth century utilized classical virological methods that were relatively insensitive for the detection of rsv and/or did not detect recently described respiratory viruses such as hmpv. thus, these studies likely underestimated the true frequency of rsv overall. nonetheless, early studies from large transplant centers reported serious sequelae in small numbers of patients who developed pneumonia where rsv was detected [ , , ] . at the fred hutchinson cancer research center (hutch), a prospective surveillance study conducted in documented respiratory viral infections in ( %) of immunocompromised patients who were followed until hospital discharge [ ] ; fi ve ( %) patients developed pneumonia and two ( %) patients died (one with rsv, the other with adenovirus). a subsequent prospective surveillance study described viral infections and revealed an overall frequency of viral infections of approximately % [ ] ; % of rsv isolates were from bal. this study demonstrated the relatively high numbers of patients with rsv lower respiratory tract disease, which was higher than rates of lrti caused by piv ( %) infl uenza ( %), or rhinovirus ( %). results of viral surveillance in transplant units vary depending on the type of surveillance protocols utilized, time of year surveillance was conducted, type of clinical samples evaluated, and laboratory tests utilized. at the huddinge university hospital, stockholm, a prospective surveillance study conducted among hsct recipients between and detected ( . %) respiratory viral infections in patients, including rsv in % [ ] . at m.d. anderson cancer center (mdacc), prospective surveillance conducted using culture techniques among hospitalized adult hsct recipients during two -month winter periods detected ( %) respiratory viral infections in hospitalized patients with acute respiratory symptom. nearly half of these were due to rsv ( patients, %). the impact of these illnesses was considerable: ( %) patients had rsv infection progressing to pneumonia, and patients with rsv pneumonia died. during the winter period when community viruses were frequent and nosocomial transmission was high, the frequency and mortality of respiratory viral-associated pneumonias was more than four times as high as cmv-associated pneumonia. recent prospective studies conducted in transplant centers worldwide continue to demonstrate the importance of rsv and report incidence of infections and risk factors for fatal disease (table - ). a study from barcelona, spain, described the -year incidence of symptomatic respiratory viral infections in over patients followed for up to several years posttransplant. altogether, % of allogeneic hsct recipients and % of autologous hsct recipients had respiratory viruses detected, with patients ( . %) receiving either autologous or allogeneic transplants having symptomatic rsv disease [ ] . risk factors associated with having a respiratory virus identifi ed included close household contacts with children under the age of years and chronic graft versus host disease. lymphopenia was identifi ed as a major risk for uri progression. similar rates have been demonstrated in other centers in the usa [ ] , south america [ ] , and europe [ ] . less data is available documenting the impact of rsv in sot recipients (table - ), rsv has been shown to cause lower respiratory tract disease and has been associated with other complications, such as organ rejection and bronchiolitis obliterans. lung transplant recipients have the highest rates of rsv lower respiratory tract disease in diverse types of organ transplant. rsv infections in lung transplant recipients have also been associated with organ rejection and progressive bronchiolitis obliterans, both of which have been observed to be seasonally related [ , ] . adult renal transplant recipients have also been reported to develop rsvassociated lower respiratory tract disease. the mortality has been low, and most patients have recovered without specifi c antiviral therapy [ , ] . rsv infections in pediatric liver transplant recipients have been associated with signifi cant morbidity and some but relatively low mortality [ ] . among pediatric liver transplant recipients cared for at the university of pittsburgh between and , ( . %) children developed rsv infections, three-quarters of which were nosocomially acquired. the majority of the children had lower respiratory tract involvement, and two ( %) children died. specifi c antiviral therapy was not administered. the risk factors for more severe disease included onset of infection early after transplant, preexisting lung pathology, augmented immunosuppression prompted by rejection, and younger age. infections occurring late after transplantation in the absence of rejection were usually not severe. an intensive prospective approach to determining the incidence and risk factors for respiratory viruses was carried out at the hutch among hsct recipients, who were pro-spectively enrolled between - and tested weekly through days posttransplant using both culture and rt-pcr detection methods [ ] . the cumulative incidence estimates of hmpv and rsv at day were similar, at . % and . %, respectively. multivariable analysis demonstrated that only recipient cmv seropositivity was associated with increased risk for acquisition of a respiratory virus (hazard ratio = . , ci . - . , p = . ). the frequency of rsv infections and associated morbidity and mortality differs substantially, potentially accounting for the variability reported by different institutions. these differences refl ect the intensity of viral surveillances, the time of surveillance, the viruses prevalent in the community, the degree of immunosuppression of the patients, infection control policies, the inclusion of potential as well as actual transplant recipients, surveillance in outpatients as well as inpatients, the types of laboratory assays utilized, and the case defi nition utilized (i.e., both clinical and laboratory defi nitions). hmpv has been detected worldwide with a seasonal distribution. community outbreaks occur yearly mainly in winter and spring (january to may in the northern hemisphere; june to july in the southern hemisphere) (figure - ). often hmpv outbreaks will be concomitant with or subsequent to rsv outbreaks. hmpv most commonly affects young children less than years old and is second only to rsv as a cause of bronchiolitis. seroprevalence studies have shown a high percentage of children have contracted the virus by age - years. however, reinfection can occur later secondary to insuffi cient immunity or infection with different genotypes. predominant hmpv strains can vary from location to location and from year to year. vicente et al. have reported higher virulence by genotype a [ ] , while papenburg et al. reported higher virulence by genotype b [ ] . however, the interaction or impact of hmpv with other viruses or bacteria remains unclear, particularly in immunocompromised patients. the importance of hmpv in transplant recipients has not been as well studied as rsv (table - ). it was fi rst reported shortly after the detection of hmpv by boivin et al. [ ] . an early prospective longitudinal study from spain documented hmpv in both autologous and allogeneic hsct recipients, with the incidence and clinical impact of hmpv and rsv disease documented to be quite similar [ ] . one early prospective study documented hmpv infections in adults with hematologic malignancies that progressed from upper respiratory infection to pneumonia, with a case-fatality rate close to % [ ] . lower respiratory tract disease and pneumonia due to hmpv infection in hsct recipients has been reported to have an overall incidence of - % [ , , , ] . a single case series described hmpv-positive nasal aspirate samples in % of adults following hsct, many of whom were asymptomatic [ ] . this study demonstrated very high rates of genetically similar viruses and differs from most other studies due to high rates of genetically identical viruses. these authors suggested that these hmpv infections may have originated in the hospital nosocomially; nonetheless, this study is an outlier compared to other reports of hmpv in transplant centers. pneumonia rates following hmpv infection have been reported at - % with mortality rates of - % [ - ] . among hsct recipients who underwent bal for investigation of lower respiratory tract disease with pulmonary infi ltrates by radiographic imaging, hmpv was detected in bal samples from of ( %) patients; four of these fi ve died with acute respiratory failure highlighting the potential severity of hmpv pneumonia [ ] . a retrospective cohort study at the hutch described a high mortality rate of % among patients with hmpv pneumonia, a rate similar to rsv pneumonia mortality [ ] . studies from other transplant units continue to document the potential of hmpv to cause severe lower respiratory tract disease, with clinical presentations and outcomes generally similar to rsv [ , ] . the signifi cance of hmpv infection in sot recipients remains less well defi ned, with the exception of hmpv disease in lung transplant recipients [ ] . case reports of severe disease have been described following liver and renal transplantation [ , ] . rates of hmpv infection in lung transplant recipients have been reported to be similar to those seen in studies of hsct recipients, varying from - % [ , ] . in most of these patients, hmpv appears to frequently be the sole pathogen detected. detection of hmpv in lung transplant recipients may not necessarily signify disease, as was noted in a study of lung transplant recipients undergoing bal mainly for surveillance purposes; four cases of hmpv was detected in asymptomatic patients [ ] . hmpv infection has been found in - % of lung transplant recipients, but prevalence may be higher during nosocomial outbreaks [ , ] . one study in the setting of a community outbreak identifi ed hmpv in bal samples from of ( %) patients; their clinical presentation varied from asymptomatic infection to severe disease [ ] . acute allograft rejection was more frequent in the hmpvinfected group than in the non-hmpv-infected group ( % vs. %, respectively; p = . ); and overall mortality was also higher ( % vs. %, respectively; p < . ) [ ] . another prospective study found hmpv infection as frequent as rsv after lung transplantation, and to cause as much pneumonia and acute allograft dysfunction ( % vs. %, respectively), but only rsv was associated with chronic allograft dysfunction at months [ ] . in another study, % of hmpv infections in lung transplant recipients were associated with acute allograft dysfunction compared with % for rsv [ ] . a meta-analysis of hmpv respiratory infections and allograft rejection, among lung transplantation recipients indicated that detection of hmpv from airway secretions may be a signifi cant posttransplantation occurrence. a total of samples from lung transplant recipients, were analyzed for virus detection; samples had viruses identifi ed and ( %) were positive for hmpv. twenty of these ( %) cases of hmpv had acute rejection within months of viral detection. there were fi ve ( . %) cases of chronic rejection in association with hmpv. all studies included in the meta-analysis, with the exception of one, identifi ed rejection within months. another study has also described cases of chronic rejection within months [ ] . disease manifestations of rsv are dependent on many factors including the immunity and immune competence of the host, the time of infection related to transplant, the type of transplant, the age and underlying health of the patient, and the degree and duration of immunodefi ciency. rsv infections in hsct recipients typically follow the same clinical sequence as rsv infections in previously healthy children: signs and symptoms of a uri such as rhinorrhea, sinus congestion, sore throat, or otitis media frequently precede signs of lower respiratory tract disease including cough, wheezing, hypoxia, and pneumonia [ , , ] (tables - and - ). the presence of wheezing with respiratory symptoms during the respiratory virus season may provide the clue that rsv may be present. progression of uri to lri has been associated with patients who are early (< month) posttransplant, risk factors for the progression of rsv upper respiratory disease to lower tract disease or pneumonia and factors relating to fatal disease have been evaluated. the most common risk factor described in multiple centers using different methods of case ascertainment and viral detection remains lymphopenia. in a prospective multicenter study carried out by the european group for blood and marrow transplantation, lymphopenia but not neutropenia significantly increased the risk for lower respiratory tract disease [ ] . older age and donor status are also signifi cant risk factors in some studies, whereas cmv serostatus, acute graft versus host disease, time relative to engraftment, and preemptive aerosolized ribavirin at a low dose of h daily were not signifi cant [ ] . investigators from mdacc have demonstrated that season of year, relapse of malignancy, presence of graft versus host disease, increasing age, and lack of engraftment are inpatient risk factors for the development of rsv pneumonia [ , , , ] . recently, large retrospective studies have identifi ed graft source including cord or marrow (adjusted hazard ratio (hr), . , % ci . - . ) and oxygen requirement (adjusted hr . , % ci, - . ) to be independently associated with death due to respiratory failure in hsct recipients [ ] . smoking history, conditioning with high-dose total body irradiation, and an absolute lymphocyte count < /mm at the time of uri onset are also signifi cantly associated with disease progression [ ] . since initiation of therapy hinges on prompt diagnosis, the possibility of false negative laboratory tests must be considered in individual patients and the diagnosis should be aggressively pursued by other means, such as bal. regardless of the therapeutic intervention, high rates of mortality due to rsv pneumonia are documented in seriously immunocompromised patients when therapy has been initiated after the development of respiratory failure (figure - a ) . survival rates remain low for severely immunosuppressed patients who are intubated due to progressive rsv pneumonia unrelated to super-imposed pulmonary hemorrhage, pulmonary edema, or bacterial superinfection [ , , , , ]. hmpv may cause upper or lower respiratory tract infections in hsct recipients. asymptomatic shedding from upper respiratory tract has been reported, indicating that not all hmpv infections result in severe lower tract disease [ , , ] . hsct recipients with hmpv disease in the immediately posttransplant period typically present with respiratory symptoms including nasal congestion, sore throat, cough, or fever. once lower respiratory tract disease develops, rapidly progressive pulmonary infi ltrates frequently accompanied by hypotension, septic shock, or both may be present [ ] ( figure - b and c ) . in a prospective viral surveillance in hsct recipients where samples for respiratory viruses were obtained weekly, regardless of the presence of respiratory symptoms, a cumulative incidence estimate of hmpv of . % ( % ci, . - . ) over year was determined; all cases of hmpv had clinical respiratory symptoms identifi ed ranging from mild to more severe disease with single or multiple symptoms [ ] (table - ) . among hsct recipients with hmpv detected by rt-pcr in bal, ( %) had positive hmpv rt-pcr in nasal wash sampled within days prior to or following the bal [ ] . viral rna was detected in the serum of one of these severely immunocompromised hsct recipients at two time points, days apart, with a viral load of ~ log copies/ml in each sample. this patient died of severe respiratory disease. in assessing risk factors associated with overall mortality at day posttransplant, the use of bone marrow as the stem cell source, steroid treatment and oxygen use have been associated with overall mortality [ ] . radiographic fi ndings associated with hmpv infection in the hsct recipient may consist of centrilobular nodules, ground glass opacities, tree-in-bud to diffuse bilateral alveolar infi ltrates [ , ] (figure - b-f ). centrilobular nodules have been associated with less mechanical ventilation while ground glass opacities tended to be associated with increased rates of hypoxemia [ ] . alveolar consolidation corresponds to more extensive damage on histological examination. histological evaluation has also shown hyaline membrane formation, foci of bronchiolitis obliterans and organizing pneumonia and diffuse alveolar hemorrhage. variable disease severity has been reported following hmpv infection in sot recipients. in one study of lung transplant recipients, hmpv was detected in four symptomatic and one asymptomatic patients ( . %), but viral infection was not persistent and resolved without major complications [ ] . in another study, nine lung transplant recipients with hmpv were compared with transplant recipients without hmpv infection; hmpv infection was associated with signs of acute graft rejection and increase overall mortality (three of nine with hmpv-infected patients died and none died in the hmpv-negative group) [ ] . outbreaks of respiratory viruses occur annually in the community, with potential for patients, families, or health care workers to become infected following exposure outside the hospital. however, nosocomial transmission within the hospital setting becomes a serious concern because of the high rates of morbidity and mortality in immunocompromised patients documented in nosocomial outbreaks. hospitalbased oubreaks of rsv infection in hsct recipients can occur through introduction of circulating community strains as well as transmission of identical viral strains among patients [ , ] . outbreaks of rsv have been associated with high mortality rates ranging up to % of infected patients [ , ] . the transmission of identical strains of rsv within the outpatient setting into the hospital setting has also been shown [ ] , demonstrating the importance of infection control measures in both the inpatient and outpatient setting. nosocomial transmission of hmpv can also occur, with outbreaks possible in both inpatient and outpatient units. in one study, patients were diagnosed with hmpv within weeks in a tertiary care cancer unit [ ] . molecular subtyping revealed infection with genotype a a virus, implicating nosocomial transmission. four patients ( . %) died from hmpv-associated pneumonia and consequent multi-organ failure. current options for the antiviral therapy of rsv disease in immunocompromised hosts are limited. large, controlled, therapeutic trials for rsv pneumonia or lower tract disease in immunocompromised patients have not been conducted. aerosolized ribavirin, licensed in the usa for the therapy of rsv bronchiolitis and pneumonia in infants and young children in , is the antiviral agent currently utilized for the treatment of rsv disease in immunocompromised patients. in general, ribavirin is given as an inhaled aerosolized solution via a face mask in a protective environment, such as a "scavenging tent," to protect health care workers from potential drug contamination. ribavirin was initially licensed for use when given for - h/day, but the use of intermittent aerosolized ribavirin given over h, three times daily, was found to have similar effectiveness to - h/day continuous infusion ribavirin in healthy children [ - ] and has been utilized in adults because of ease of administration and enhanced tolerability. a randomized trial in hsct patients at risk for lrti evaluated intermittent dosing of ribavirin given over h three times daily versus continuous ribavirin administration using an adaptive randomized trial design in hsct patients, with the authors concluding that the intermittent schedule was preferable because of ease of administration and evidence of higher effi cacy [ ] . only one randomized, controlled, multicenter clinical trial of aerosolized ribavirin for the prevention of rsv disease progression to lri has been conducted in hsct recipients early posttransplant, and despite an enrollment period of several years, only patients were enrolled [ ] . none of the ten patients randomized to high dose, short duration aerosolized ribavirin (administered as g/ ml water given over h three times daily) had disease progression compared to / control patients, a trend that was not statistically significant ( p = . ). viral loads appeared to be reduced during the ribavirin treatment, but did rebound after cessation of therapy. data demonstrating effectiveness of ribavirin is mainly retrospective. in an open trial in adult hsct recipients with "rsv-induced acute lung injury," monotherapy with aerosolized ribavirin was reported to be of benefi t if initiated prior to the development of radiographic infi ltrates [ ] . in another open trial in adult hsct recipients with radiographically proven rsv pneumonia, combination therapy with aerosolized ribavirin and high rsv-titered ivig was reported to be of benefi t only if initiated prior to the onset of respiratory failure [ , ] . a retrospective mdacc study of confi rmed rsv infections in allogeneic hsct recipients from to utilized multivariable logistic regression to demonstrate that lack of ribavirin aerosol therapy at the upper respiratory tract disease stage was an important risk factor associated with rsv lrti and all-cause mortality [ ] . in a retrospective study of hsct recipients with confi rmed lower respiratory tract rsv infection based on analysis of bronchoalveolar lavage fl uid at the hutch, viral rna detection in the blood was detected in % of patients at a median of days following diagnosis of lower respiratory tract disease [ ] . neutropenia, thrombocytopenia, and mechanical ventilation increased the risk of rsv rna detection in the plasma or serum but lymphopenia and steroid use did not. the detection of rsv rna in the serum or plasma increased the risk of overall mortality with an adjusted hazard ratio (ahr) of . ( p = . ). data in solid organ transplant recipients is even more limited. favorable responses have been reported in an open trial of lung transplant recipients with lower respiratory tract disease who received monotherapy with aerosolized ribavirin [ ] , as well as open trials with oral ribavirin [ - ] , although controlled studies have not been performed. oral ribavirin was found to be well-tolerated, result in less hospitalization, and be less expensive than intravenous or inhaled ribavirin in a retrospective study of lung transplant recipients [ ] . the treatment of rsv disease with a combination of antiviral therapy and passively administered immunoglobulin has been investigated in animal models and in children [ - ] . therapy with ivig containing high levels of rsvspecifi c antibodies alone does not seem to be effective in placebo-controlled trials in children who were not immunocompromised [ , ] . in small open trials at mdacc, combination therapy with aerosolized ribavirin ( h/day) and high rsv-titered ivig ( . g/kg every other day) was associated with a favorable response in adult hsct recipients and patients undergoing induction chemotherapy for leukemia who had rsv lower respiratory tract disease in whom therapy was initiated prior to respiratory failure [ , ] . at the dana farber cancer institute, combination therapy with aerosolized ribavirin ( h/day) and rsv-ivig ( . g/kg for one dose) was similarly associated with a favorable response in hsct recipients with clinically severe rsv pneumonia occurring early following transplant [ ] . in subsequent years, mdacc has utilized a combined regimen with similar response, although standard ivig in frequent and large doses ( mg/kg qod) has been substituted for high-titered ivig. other therapeutic options for the treatment of rsv include the use of oral ribavirin, which has been studied in hsct recipients and found to be safe and less expensive than intravenous or aerosolized ribavirin [ , , ] , and is recommended as an option in addition to intravenous and inhaled ribavirin by the european conference on infections in leukemia [ ] . other options include iv ribavirin (an investigational drug, icn) and topical immunoglobulins administered with aerosolized or intravenous ribavirin [ , , ] . the relative ease of administration of iv ribavirin is attractive, but high rates of mortality ( %) and signifi cant cases of hemolytic anemia ( %) make this option currently problematic. although the european experience with combination aerosolized/intravenous ribavirin has been favorable [ ] and intravenous ribavirin is relatively simple to administer, the high rates of mortality and signifi cant cases of hemolytic anemia make this approach controversial. monotherapy with iv ribavirin may be more toxic in these patients than has been previously reported in patients with hemorrhagic fevers [ , ] . the decision to initiate therapy with aerosolized ribavirin with or without immunotherapy for a rsv-uri must take into consideration many factors, including the patient's risk of developing serious lower respiratory tract disease (and specifi cally, the degree of anticipated lymphopenia), the potential exposure of health care workers to the medication, the psychological and physical discomfort to the patients of aerosol therapy, the adverse effects of aerosolized ribavirin such as bronchospasm, the high cost of these drugs as well as the intensive respiratory therapy needed to safely administer aerosolized ribavirin, and the need for hospitalization with more frequent or prolonged ribavirin dosing regimens. in patients who have already undergone conditioning therapy and stem cell infusion but have not yet engrafted, the initiation of antiviral therapy at the uri stage may be beneficial. early studies conducted in the s were small and uncontrolled. one study conducted at fhcrc treated hsct recipients with upper tract rsv disease with low dose aerosolized ribavirin administered at a high concentration ( mg/ml) for h each day (total: g/day [ ] ). unfortunately, / patients developed pneumonia, and seven of these died. another study evaluated combination therapy with aerosolized ribavirin and mg/kg ivig every other day in patients, two of whom developed pneumonia and died [ , ] . this "preemptive" strategy, similar to that used in the prevention of cmv disease and cmv pneumonia, is used at some transplant centers for those patients at highest risk of rsv disease progression, such as pre-engrafted patients with rsv detected in the fi rst weeks following transplantation. other options for preemptive therapy include immunotherapy with ivig or rsv-specifi c monoclonal antibodies, although little data on effi cacy of immunotherapy is available. no antivirals for the therapy of hmpv are currently available or routinely utilized. ribavirin is active in vitro and in vivo against hmpv, although there are no controlled studies or evidence from large retrospective reviews for the treatment of hmpv pneumonia in humans and no drug has yet demonstrated clinical effectiveness in humans [ , ] . intravenous, oral or inhaled ribavirin alone or in combination with ivig has been reported as potentially successful therapeutic options. a retrospective analysis compared the outcome between immunocompromised patients with hmpv pneumonia treated with ribavirin ± ivig and ten untreated patients. ribavirin treatment was associated with more hypoxemia and similar mortality, possibly related to late initiation of therapy [ ] . a seattle study describing hmpv lower respiratory tract disease in immunocompromised children, including nine undergoing hsct and eight sot recipients, demonstrated that hsct recipients had more evidence of severe disease [ ] . five of eight hsct recipients but no sot recipients had lower tract disease and were treated with aerosolized ribavirin; three had been diagnosed with hmpv pretransplant and during the posttransplant period received both ribavirin and ivig. two additional children received aerosolized ribavirin only. ribavirin was generally administered at a dose of g given three times daily for - days. two of the three patients diagnosed with hmpv pretransplant who received ribavirin and ivig died [ ] . an aggressive infection control strategy can be effective in reducing the nosocomial acquisition of rsv by transplant recipients [ , ] . infection control strategies play a crucial role in the prevention of respiratory viral infection [ , - ] . an effective strategy is based on understanding the potential seriousness of these infections in transplant recipients, knowledge of the viruses circulating in the community, and ongoing surveillance in high-risk patients. continuing education of patients, family members, visitors, and staff regarding the potential seriousness of these infections must be repeatedly emphasized. frequent and routine clinical screening of high-risk patients for acute upper and/or lower respiratory tract illness or fl u-like illness must be conducted, with sampling of respiratory secretions from symptomatic high-risk patients routinely performed both pretransplant and posttransplant. each health care-acquired infection should be viewed as a sentinel event warranting an investigation and reaffi rmation or modifi cation of the preventative strategy. infection control strategies should be designed to prevent spread by multiple modes of transmission [ , ] . multiple respiratory viruses may circulate in the community concurrently and can be spread by different means. infection control measures may need to be intensifi ed during community or hospital outbreaks, and the intensity and duration of infection control measures should be tailored to the risk of serious disease in different subsets of transplant recipients, and to what works in the "real world." guidelines for the prevention of opportunistic infections among hsct recipients have been issued by the centers for disease control and prevention (cdc), the infectious diseases society of america, and european and american societies for blood and marrow transplantation. the guidelines clearly present evidencebased recommendations rated by the strength of the recommendation and the quality of the supporting evidence, similar to guidelines previously issued for the prevention of opportunistic infections in those with human immunodefi ciency virus [ ] . preventive strategies for hsct recipients, their household contacts and other close contacts, and health care workers are clearly outlined in this document. the prevention of nosocomial acquisition of respiratory viral infections in hsct recipients has been demonstrated in one prospective study comparing rates of infection in patients cared for in a "protected environment" with patients cared for on a transplant unit where infection control measures were strongly encouraged, but not rigidly enforced [ ] . the effectiveness of infection control interventions has also been demonstrated by the dramatic decline in the frequency of nosocomial crv infections among hsct recipients cared for in this transplant unit after the implementation of an aggressive, multifaceted infection control strategy [ ] . although this intensive multifaceted approach has been effective, modifi ed versions of this strategy have also been effective. for instance, the seattle cancer care alliance (scca ) adult inpatient transplant unit uses a similar strategy with the exception that health care workers and other visitors do not wear masks when entering the patient room [ ] . however, these workers are intensively screened for signs and symptoms of respiratory illnesses prior to entering the unit, and restricted from entering the unit if they are symptomatic. similarly, hsct recipients with respiratory symptoms are not transferred to other units, but are cared for on the transplant unit using modifi ed droplet precautions with all persons entering the room wear gloves, gown, and mask, and the door to the room is kept closed. it may not be feasible to so intensively protect patients for the duration of increased susceptibility to respiratory viral diseases. protective strategies are costly and cumbersome, and pose unpleasant restrictions on the freedom and quality of life of the patient and their families. this problem is further compounded by the growing trend to discharge patients early from the hospital and to perform outpatient hsct or posttransplant care. transplant recipients residing in the community and followed frequently in the outpatient setting are another group in which infection control practices must become priority [ ] . the prevention of exposure to respiratory viruses is particularly challenging among high-risk transplant recipients living in the community because respiratory infections are so prevalent and so contagious. examples of protective measures for outpatients include washing hands frequently and thoroughly, avoiding close contact with individuals suffering from respiratory illnesses, and encouraging close contacts to vigorously practice respiratory hygiene. in many cases, such as individuals living with children, such efforts may be nearly impossible. consideration of removing day care exposures for young children or decreasing exposure to transplant recipients to children (including siblings), can and should be discussed with families. the rigor and duration of prophylactic measures need to be individualized based on the immunologic status of the patient and the risk for serious disease, the needs of the patient, and quality-of-life issues. passive immunization with immunoglobulin, immunoglobulin products, and humanized monoclonal antibodies have been actively studied in the pediatric infant population. palivizumab , a humanized monoclonal antibody directed against the rsv f protein, is licensed for the prevention of rsv disease in premature infants and infants with congenital heart disease, and is administered as a monthly injection during the - months of rsv season. the cost of this therapy has led to new guidelines for use in the pediatric population [ ] . similar interventions have been utilized to attempt to decrease the morbidity of serious rsv disease in hsct recipients but direct proof of effectiveness has not yet been demonstrated [ ] . for example, passive immunoprophylaxis in immunocompromised patients has been evaluated in an open trial conducted [ ] using high-titered human rsvig. in this adult study, signifi cant antibody titer increases to other respiratory viruses were extremely variable, although the subset of patients with the lowest titers appeared to receive the greatest increase in viral-specifi c antibody. the cost of this potential therapy remains quite high. the monoclonal rsv antibody palivizumab (synagis) has been studied in an open label study in adult hsct recipients [ ] . immunoprophylaxis with monoclonal rsv antibodies would be prohibitively expensive in older children and adults, but may have the potential to protect against rsv infection, based on pediatric studies. new antibody products, including long-lasting monoclonal antibodies with enhanced activity against rsv, are under development and if available at a less expensive price, could potentially provide protection against rsv for patients in the pretransplant and immediate posttransplant phase. newer monoclonal antibodies that have the potential to neutralize against both rsv and hmpv have also been described, offering hope for newer preventive modalities that may protect against both these viruses [ ] . screening of transplant recipients for respiratory viruses prior to transplant is not routinely recommended based on current us and international transplant guidelines [ , ] . however, the assessment of viral shedding or infection in symptomatic transplant candidates prior to transplant is recommended. delay of transplant based on detection of rsv or hmpv may be warranted depending on the evaluation of the risk and benefi ts of continuing on with transplantation. consequences of postponing transplantation should be considered, including progression of underlying malignancy, logistical issues regarding the donor availability, and accessibility of services for the patient. an early study of rsv diagnosed prior to hsct demonstrated that delaying transplant reduced the risk of pneumonia following transplant [ ] . more recently, a large prospective study conducted in patients at the hutch demonstrated that nearly % of subjects had respiratory viruses detected pretransplant [ ] . overall, patients with a virus detected prior to transplant had fewer days alive and lower survival at day (ahr . ; % ci . - . ) compared with patients who did not have viruses. in hsct recipients who had respiratory symptoms and a virus detected prior to transplant (mainly adults), an increased overall mortality was seen compared with patients without respiratory viruses detected. higher rates of pretransplant infection and sequelae of infection were seen in pediatric patients. detection of respiratory viruses in asymptomatic patients was not associated with increased mortality. this data strengthens current guidelines that recommend patients with respiratory symptoms prior to transplant should be tested for respiratory viruses and transplant delayed, when feasible. however, this study was performed chiefl y in adults. the higher rates of respiratory viruses documented in children pretransplant make routine screening of pediatric patients worthy of further investigation. experimental approaches to the therapy of rsv antiviral therapy include novel fusion inhibitors [ ] , nucleoside agents, small rna inhibitory molecules [ ] , and high-titered monoclonal antibody preparations. two promising rsv antivirals that have shown effi cacy in challenge studies in healthy adults include the alios compound al (alios biopharma, south san francisco, ca), an oral anti-rsv nucleoside designed to inhibit rsv replication by acting on the viral polymerase, and the gilead sciences compound gs (gilead sciences, foster city, ca), an orally bioavailable rsv fusion inhibitor [ , ] . clinical trials of these agents in healthy young children with rsv have been proposed [ ] . an international multicenter, placebo-controlled clinical trial of gs was initiated in july , and remains ongoing in adult hsct recipients with documented rsv infections (clinica ltrials.gov identifi er nct ). other antiviral agents are under development. no rsv or hmpv vaccine is currently available. in general, active immunization of transplant recipients will be unlikely to prevent severe disease seen in the fi rst several months posttransplant. however, prevention of rsv infection in families, staff, and nosocomial spread of virus by the use of vaccines holds true promise to benefi t transplant recipients themselves. promising advances in new vaccines directed against both rsv and hmpv have been reported over the past decade, with progress evident in both rsv fusionprotein based vaccines and live attenuated vaccines. advances in the understanding of the pre-and post-fusion nature of the rsv f protein [ ] has led to increased work in developing protein-based vaccines appropriate for older children, adults, and pregnant women. advances in technology and better molecular understanding of rsv and hmpv have resulted in new potential rsv candidate vaccines [ ] . at least rsv vaccines are in clinical studies in phase or clinical studies, with one rsv f vaccine under study in pregnant women. examples of live rsv vaccine candidates under study include live-attenuated vaccines relying on genetic manipulation of the rsv genome [ ] , vectored virus vaccines utilizing the chimpanzee adenovirus or vaccinia virus ankara [ ] , or chimeric viruses containing a backbone of attenuated parainfl uenza with the f gene of rsv added [ , ] . a chimeric hmpv vaccine containing a backbone 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against human metapneumovirus key: cord- -v crgj w authors: pastores, stephen m.; dulu, alina o.; desouza, shilpa a. title: what has been learned from postmortem studies? date: - - journal: pulmonary involvement in patients with hematological malignancies doi: . / - - - - _ sha: doc_id: cord_uid: v crgj w infectious and noninfectious pulmonary diseases are commonly found on postmortem autopsy studies in patients with hematological malignancy. despite the technological advances in diagnostic testing and imaging modalities, obtaining an accurate clinical diagnosis remains difficult and often not possible until autopsy. major diagnostic discrepancies between clinical premortem diagnoses and postmortem autopsy findings have been reported in these patients. the most common missed diagnoses are due to opportunistic infections and cardiopulmonary complications. these findings underscore the importance of enhanced surveillance, monitoring and treatment of infections and cardiopulmonary disorders in these patients. autopsies remain important in determining an accurate cause of death and for improved understanding of diagnostic deficiencies, as well as for medical education and quality assurance. infectious and noninfectious pulmonary complications occur in - % of patients with hematological malignancy and recipients of hematopoietic stem cell transplantation (hsct) and are associated with signifi cant morbidity and mortality [ ] . in allogeneic hsct patients who develop respiratory failure requiring mechanical ventilation, the intensive care unit (icu) and hospital mortality rates often exceed % and %, respectively [ ] [ ] [ ] [ ] . the factors that contribute to the development of pulmonary complications in these patients include immunologic defects due to the underlying disease and its treatment, conditioning regimens, development of graft-versus-host disease (gvhd), and the type of hsct [ , , ] . the spectrum of pulmonary complications in patients with hematological malignancy and hsct recipients is changing because of recent advances in antineoplastic therapies, such as the use of monoclonal antibodies and other targeted agents, increased application of hsct for older patients, widespread use of prophylactic antibiotics and novel antimicrobial agents, and advances in supportive care [ ] . prompt investigation and diagnosis of pulmonary complications in patients with hematological malignancies are essential to improving patient survival. unfortunately, despite the technological advances in diagnostic testing and imaging modalities, obtaining an accurate clinical diagnosis in these patients remains difficult and at times is made only at the time of the postmortem autopsy examination. autopsy rates in cancer patients are much lower ( - %) as compared to general medical and surgical patients ( - %) [ ] [ ] [ ] . this probably reflects the unwillingness on the part of physicians and family members to subject the cancer patient to the same level of scrutiny applied to other major illnesses in determining the primary and contributory causes of death [ ] . additional concerns include legal issues regarding exposition of physicians' errors and non-reimbursement for postmortem examinations [ ] . this chapter will review the infectious and noninfectious pulmonary findings that have been described at autopsy in patients with hematological malignancies, including blood and bone marrow transplant recipients. in addition, we discuss the frequently noted diagnostic discrepancies between premortem clinical diagnoses and postmortem autopsy findings in these patients. finally, we highlight the difficulties in diagnosing many of these conditions antemortem and emphasize the important role of the postmortem examination in accurately establishing the cause of death. table . lists the infectious and non-infectious pulmonary disorders reported in autopsy studies of patients with hematologic malignancy, including hsct recipients. the incidence of invasive fungal infections in patients with hematologic malignancies has increased steadily over the past three decades [ ] . this has been attributed to improvements in the prevention or preemptive treatment of bacterial infections and other opportunistic pathogens, particularly candida and cytomegalovirus; increased administration of chemotherapies with profound and prolonged immunosuppressive effects on t-cell function (e.g., purine nucleoside analogs, anti-t-cell immunoglobulin, and monoclonal antibodies); and the growing number of allogeneic and nonmyeloablative transplantation procedures that carry a higher risk for chronic gvhd [ ] . patients with hematologic malignancies, especially in the neutropenic state after aggressive chemotherapy or hsct and hsct recipients with gvhd, are particularly susceptible to invasive fungal infections [ ] . approximately - % of patients with hematological malignancies and hsct recipients have evidence of invasive fungal infections at autopsy [ ] . these include invasive aspergillosis and fungal infections due to candida spp., zygomycetes spp., trichosporon spp., fusarium spp., and various chaetomium species. in neutropenic patients with acute leukemia, the histopathological pattern of invasive pulmonary aspergillosis (ipa) is characterized by scant inflammation, hyphal angioinvasion with a high fungal burden, and extensive coagulative necrosis [ , ] . in contrast, among hsct recipients with gvhd, the histopathological findings consist of severe lung inflammation and less abundant aspergillus burden. several autopsy [ , [ ] [ ] [ ] [ ] [ ] [ ] [ ] . in recent years, treatment of ipa with voriconazole has led to better responses, improved survival rates, and fewer side effects than amphotericin b [ ] . autopsy studies have assisted in describing and confirming the immune reconstitution inflammatory syndrome (iris) in patients with ipa who are recovering from the neutropenia [ ] . when clinical and radiologic worsening coincides with neutrophil recovery, it is usually assumed that this deterioration is related to progressive aspergillosis, prompting changes in patient man agement. however, its temporal relation with neutrophil recovery suggests that it may be caused by iris. the patients who died during the first month had no evidence of aspergillosis at autopsy. finally, autopsy studies have proved helpful in documenting the efficacy of systemic antifungal therapy and surgery for ipa [ ] . since the early s, the incidence of invasive candidiasis (candidemia and/or hepatosplenic candidiasis) has continued to decrease due to effective antifungal prophylaxis and empirical treatment of high-risk patients with echinocandins and voriconazole [ ] . mucosal damage is a risk factor for invasive candidiasis among patients receiving antineoplastic therapy. hsct recipients who received conditioning regimens with total body irradiation and patients treated with chemotherapy regimens containing high-dose cytarabine or an anthracycline have an increased risk of developing invasive disease. in recent years, there has been an increase in bloodstream infections caused by non-albicans candida species such as candida glabrata and c. krusei. the diagnosis of invasive candidiasis is difficult to prove due to the lack of specific clinical features and the low sensitivity of blood cultures to isolate candida, especially in patients receiving fluconazole prophylaxis [ ] . difficult-to-treat opportunistic molds, such as zygomycetes, trichosporon spp., and various chae tomium spp., including c. atrobrunneum, c. strumar ium, c. globosum, c. perlucidum, and c. cinereus are being described with increasing frequency on autopsy studies in patients with hematologic malignancies [ , [ ] [ ] [ ] [ ] [ ] [ ] . pulmonary involvement with these mold infections is characterized by tissue necrosis from angioinvasion and subsequent thrombosis. as with many fungal infections, diagnosis of these infections is often not possible until autopsy. treatment modalities usually involve lipid-based amphotericin b formulations and surgical debulking or debridement in selected cases [ ] . pneumocystis jiroveci pneumonia (pcp) remains a serious infection in patients with acute and chronic leukemias, myelodysplastic syndrome, and hsct recipients [ ] . however, diagnosis of pcp is frequently obtained by bronchoalveolar lavage with or without lung biopsy; thus, the diagnosis is made on autopsy in only a minority of cases. bacterial pneumonias caused by pseudomonas aerugi nosa, streptococcus spp., staphylococcus aureus, serratia spp., and legionella pneumophila have been described on autopsy studies in patients with hematologic malignancy and hsct recipients [ ] . as the majority of these patients commonly receive empiric antimicrobial therapy during the initial diagnostic workup for infection, very few autopsy studies report unusual bacterial infections. a single center autopsy series of patients with hematological malignancies found multidrug-resistant strains such as enterococcus faecium to be very prevalent [ ] . two coagulasenegative staphylococcus epidermidis strains were also noted. a few autopsy case reports have also described lethal pulmonary hemorrhage due to stenotrophomonas maltophilia [ ] . the incidence of autopsy-proven cytomegalovirus (cmv) pneumonia in patients with hematologic malignancy and hsct recipients has been decreasing in recent years as a result of improvements in early diagnosis and treatment and more effective preventive strategies [ ] . however, it is also possible that the declining rate of autopsies may account for the decrease in the number of reported cmv pneumonias. other etiologies of viral pneumonias that have been described in autopsy reports include infection due to herpes simplex virus, respiratory syncytial virus (rsv) [ ] , and measles virus [ ] . reactivation of latent toxoplasmosis is a rare but wellrecognized opportunistic infection in immunocompromised patients. besides encephalitis, the other common presentation with toxoplasma gondii infection is interstitial pneumonitis. because of its non-specific clinical and radiological presentation and its lethal outcome, toxoplasmosis is often misdiagnosed and only revealed at autopsy [ ] . toxoplasmic pneumonitis follows the same pathogenetic mechanism, but occurs less frequently than either toxoplasmic encephalitis or other opportunistic pneumonias, such as pcp. diagnosis is based upon a high degree of clinical suspicion and demonstration of t. gondii in bal fluid and/or lung biopsy specimens. widely disseminated necrotic areas with numerous cysts of toxoplasma gondii are commonly reported in autopsy cases. noninfectious pulmonary complications account for up to % of autopsy findings in patients with hematologic malignancies, particularly in hsct recipients. the most common complications are diffuse alveolar damage (dad) and diffuse alveolar hemorrhage (dah) [ ] . diffuse alveolar damage (dad) is a nonspecific finding at autopsy often in association with various infectious and noninfectious etiologies, such as shock, aspiration, alveolar hemorrhage, peri-engraftment respiratory distress syndrome, drug toxicity, and radiation therapy. it is characterized by the presence of alveolar injury and the absence of active lower respiratory tract infection. dad has been reported at autopsy in . % of patients with treated leukemia and lymphoma and close to % among hsct patients [ , ] . infections as the cause of dad are identified on autopsy in only a third of hsct patients, while approximately % have dah. in over % of patients with dad, no etiology is determined, and these patients are considered as having idiopathic pneumonia syndrome (ips). it is possible that empirically treated previous infections could have caused the histological changes noted in patients classified as having ips. only one third of the cases of dad are diagnosed antemortem [ ] . given that almost half of the cases of dad may be secondary to ips, the role of corticosteroids may need to be furthered studied. diffuse alveolar hemorrhage (dah) is a clinical syndrome characterized by the acute onset of alveolar infiltrates, bloody bronchoalveolar lavage, and hypoxemia in the absence of infection [ , ] . the incidence of dah ranges from - % to - % in autologous and allogeneic hsct recipients, respectively [ ] [ ] [ ] . a few case reports have been described in patients with acute leukemia more commonly in association with chemotherapy [ ] [ ] [ ] . dah has also been described in patients undergoing an umbilical cord hsct [ , ] . the majority of patients with dah develop severe respiratory failure with high mortality rates. vascular damage and inflammation from chemotherapy and radiation therapy used in the con ditioning regimen and immune-mediated events including gvhd have been implicated in the pathogenesis of dah [ , , ] . wojno et al. reported that % of allogeneic hsct patients who underwent autopsies had extensive pulmonary hemorrhage, which was thought to have led to severe respiratory failure and death [ ] . these patients were subdivided into those with significant acute gvhd and those without. of the patients with acute gvhd, % died of acute respiratory failure secondary to dah compared with % of those without gvhd. pulmonary hemorrhage was also independently found to be associated with pre-transplant total body irradiation. although pulmonary and systemic infections cause alveolar damage through similar mechanisms [ ] . infection-associated alveolar damage has traditionally been excluded from analyses of dah. autopsy studies have shown that pulmonary infections are frequently underdiagnosed in hsct recipients; thus, patients with alveolar hemorrhage and underlying undetected infections can be misclassified as having dah [ , , ] . because inflammation is thought to play a role in the pathogenesis of post hsct-dah, high-dose steroids have been used for its treatment, based on anecdotal case reports and small retrospective series [ , ] . other treatment modalities that have been used for dah in hsct recipients include epsilon aminocaproic acid and recombinant factor viia [ , ] . unfortunately, the mortality from dah in hsct recipients remains high because of misdiagnosis and lack of effective treatments. primary pulmonary lymphomas are very uncommon, especially those arising from bronchus-associated lymphoid tissue (balt) and have a low mortality. they represent % of the extranodal non-hodgkin's lymphomas (nhls) and only . % of all primary pulmonary malignant neoplasms and less than % of lymphomas. eighty percent of the cases are low-grade b-cell lymphomas, which are slow growing and respond well to therapy. autopsy case reports suggest that pulmonary balt lymphoma can remain restrictive to the thorax for long periods before dissemination, but tend to relapse frequently. lymphomatous involvement of the lung is common and occurs in % and %, respectively, of patients with hodgkin's lymphomas (hl) and nhl [ ] . typical findings in the lung include peribronchialperivascular, nodular, alveolar, interstitial, and pleural involvement. peripheral t-cell lymphomas also involve the lung frequently, % at diagnosis and a further % during the course of the disease. the nodular pattern is a characteristic of lung infiltration in hl, but no differences could be detected in the subtypes. pulmonary parenchymal involvement by multiple myeloma cells is very rare and described on autopsy in few case reports. the antemortem diagnosis of lung involvement by myeloma is difficult to make as infections and alveolar hemorrhage can have the same radiologic features. leukemic infiltration of the lungs may occur in - % of hyperleucocytic patients with acute myeloid leukemia (aml). pulmonary infiltrates are usually microscopic and invariably associated with hyperleukocytosis. there are autopsy case reports of pulmonary leukemia as a cause of pulmonary infiltrates, even in non-hyperleukocytosis aml patients with low blast counts. radiographically, patients present with an airspace disease with a diffuse interstitial reticular pattern in cases of hyperleukocytosis similar to cases of infectious pneumonia. pulmonary leukostasis syndrome involves the occlusion of small blood vessels in the lungs and typically occurs with a wbc count of greater than , per ml. the increased number of wbcs causes blood viscosity to rise due to the decreased deformability of the abnormal leukocytes, resulting in cell clumping and stasis in the microvasculature, leading to severe hypoxemic respiratory failure. autopsy studies reveal extensive infiltration by leukemic cells in the pulmonary vasculature; pulmonary infarction with hemorrhage is also noted. [ ] rarely, lung involvement by intravascular large b-cell lymphoma (ivlbcl) is noted on autopsy [ ] . early diagnosis is difficult as neither computed tomography nor -gallium scintigraphy can detect lung involvement. however, -fluro-deoxyglucose positron tomography (fdg-pet) may be a powerful tool for the early diagnosis of ivlbcl with pulmonary involvement [ ] . autopsy studies show that pulmonary thromboembolism (pte) infrequently complicates the course of patients with acute leukemia and severe thrombocytopenia, and hsct recipients with an incidence rate ranging from % to . % [ ] . [ ] patients with acute leukemias commonly have clinically silent haemostatic abnormalities, but some may show clinical manifestations, including venous thromboembolism, pulmonary embolism, disseminated intravascular coagulation, and life-threatening thrombohemorrhagic syndrome. the pathogenesis of pte is complex and multifactorial and may involve tumor cell-derived procoagulant, fibrinolytic or proteolytic factors, and inflammatory cytokines, which affect clotting activation. chemotherapy and anti-angiogenic drugs also increase the thrombotic risk in patients with lymphoma, acute leukemia, and multiple myeloma. infectious complications are another important factor: endotoxins from gram-negative bacteria induce the release of tissue factor (tf), tumor necrosis factor (tnf), and interleukin- (il- ), and gram-positive organisms can release bacterial mucopolysaccharides that directly activate factor xii. needleman et al. reviewed consecutive autopsies in leukemia patients and found three patients with previously undiagnosed pte, all of whom had been severely thrombocytopenic. however, candida forms were abundant in the thromboemboli in all three patients, with some containing septate hyphal forms consistent with mucor or aspergillosis. no vessel wall invasion or necrosis was noted, and fungus was not shown to be present in pulmonary vessels in segments of the lung not involved with thromboembolism [ ] . leukemic patients may also be affected by other prothrombotic factors, including hyperleukocytosis, increased tf expression and activation, and the prothrombotic properties of therapeutic agents, such as all-trans retinoic acid and l-asparaginase, which can induce thrombosis involving multiple organs. a higher index of suspicion may lead to the diagnosis, but the signs and symptoms of pte in patients with hematologic malignancy are variable and nonspecific as with pte in other populations. the majority of patients with hematological malignancies who develop organizing pneumonia (boop) have been exposed to various chemotherapeutic agents, including cytarabine and anthracyclines as well as radiation therapy. [ , ] in one autopsy series, sharma et al. reported on patients who had undergone hsct, of whom % had bo and % had boop [ ] . unusual histological variants have also been described, including a case report of acute fibrinous and organizing pneumonia following hsct in a patient with aml [ ] characterized by prominent intraalveolar fibrin deposition and organizing pneumonia. the radiographic presentation revealed patchy consolidation in the lower lobes and a diffuse miliary pattern. clinically, these cases can have subacute presentations similar to cryptogenic organizing pneumonia or have more rapid progression with clinical features similar to ards. boop and bo are rare autopsy findings, which may be because infections are being treated aggressively, and often patients not responding to antibiotics and with no clinical evidence of infections are given a trial of corticosteroids. yokoi et al. reported bronchiolitis obliterans (bo) on autopsy in of patients who underwent allogeneic bmt with aml or all. all patients received conditioning regimens with total body irradiation and cyclophosphamide with or without busulfan or cytosine arabinoside. immunosuppressive therapies were administered to all patients after bmt, including methotrexate with or without cyclosporine. the onset of respiratory symptoms was - days after bmt, and the symptoms were non-productive cough, dyspnea, fever, chest pain, and pneumothorax. seven patients died of progressive respiratory failure and one of relapsed leukemia. coexistent infections included cmv, varicella zoster, mycobacterium tuberculosis, and aspergillus [ ] . paz pulmonary veno-occlusive disease (pvod) is a rare cause of pulmonary hypertension that is characterized histopathologically by intimal proliferation and fibrosis of the pulmonary venules and small veins leading to progressive vascular obstruction [ ] . the etiology of pvod remains unclear. it has been reported as an unusual complication of both myeloablative allogeneic, autologous and cord hsct, suggesting that it might be regimen-related toxicity [ ] [ ] [ ] [ ] [ ] [ ] . surgical lung biopsy provides definitive diagnosis [ ] . pvod carries a poor prognosis, with most reported patients experiencing progressive disease and death within years of diagnosis [ ] . autopsy findings reveal intimal fibrosis of most of the pulmonary veins, with no significant intraluminal thrombi or arterial changes [ ] . treatment recommendations are anecdotal. steroids and heparin have been reported to possibly improve outcomes [ ] . autopsy studies in patients with hematologic malignancy (particularly with myelodysplastic syndrome) and following hsct have revealed rare cases of secondary pulmonary alveolar proteinosis (pap) characterized by intra-alveolar accumulation of surfactant components and cellular debris, with minimal interstitial inflammation or fibrosis [ ] [ ] [ ] . secondary pap is frequently noted among patients with hematologic malignancies who develop fungal infection, especially pulmonary aspergillosis. kl- protein is produced by type ii alveolar pneumocytes and can be helpful in establishing an early diagnosis of pap [ , ] . the standard therapy for pap with hematological malignancy has not yet been firmly established. the administration of gm-csf has been suggested to activate the alveolar macrophages and increase the rate of surfactant clearance [ ] . case reports indicate that the prognosis of pap with leukemia is very poor because of the high frequency of superinfections in the affected alveoli [ , ] . several autopsy series have reported diagnostic discrepancies between premortem clinical diagnosis and postmortem autopsy findings ranging from % to % in patients with hematologic malignancy and hsct recipients (table . ) [ , [ ] [ ] [ ] [ ] [ ] [ ] [ ] . the goldman criteria are commonly used to categorize discrepancies between clinical and pathological diagnoses or causes of death [ ] . class discrepancies are defined as missed major diagnoses with potential adverse impact on survival and would have changed management. class ii discrepancies are missed major diagnoses with no potential impact on survival and that would have not changed therapy. class iii discrepancies are defined as missed minor diagnoses related to terminal disease, but not related to the cause of death, and class iv are other missed minor diagnoses. gerain et al. reported a % major discrepancy rate in cancer patients who were admitted to a medical oncological icu over an -month period. [ ] the majority of major discrepancies were due to complications of the cancer itself or its treatment (such as non-cardiogenic pulmonary edema, acute hemorrhage, and pulmonary embolism) rather than infection. pastores et al. reported a major missed diagnosis rate of % in autopsies performed on cancer patients who died in a medical-surgical icu [ ] . of the patients, ( %) had undergone hsct, ( %) had either leukemias or lymphomas, ( %) had solid tumors, and ( %) were surgical cancer patients. among the patients with discrepancies % had class i discrepancies, % had class ii discrepancies, and % had both class i and class ii discrepancies. of the discordant cases, had hematological malignancies and underwent hsct. opportunistic infections due to viral, fungal, bacterial, and parasitic organisms and cardiac complications were the most common class i discrepancies. the majority of class ii discrepancies were accounted for by cardiopulmonary complications due to pulmonary embolism and thrombotic endocarditis. the study was limited by the retrospective study design and selection bias that may have occurred as physicians and family members of patients with premortem diagnostic uncertainty would have been more likely to pursue an autopsy. xavier et al. reported a class i discrepancy rate of . % in autopsies of patients with hematological malignancies or severe aplastic anemia [ ] . the most common diagnoses causing these discrepancies were hematological disease, pneumonia, and gastrointestinal hemorrhage. class i discrepancies were more common in elderly patients (> years) and in patients who had not received previous specific treatment for the hematological malignancy, had not been treated with bone marrow or peripheral blood hsct, or had not been treated in a specialized hematology unit. seftel et al. reported a discrepancy rate of % ( % major, % minor) in autopsies of patients who underwent hsct (blood and bone marrow) [ ] . infectious complications, including pulmonary aspe rgillosis, candidiasis, and infective endocarditis, accounted for the majority of the major discrepancies. hoffmeister et al. found a % discrepancy rate ( % major, % minor) in autopsies of patients who had undergone hsct [ ] . in contrast, al saidi et al. found a pneumonia - significant concordance between the clinical and postmortem diagnoses in critically ill hsct patients [ ] . ten ( %) of the twenty eight patients had discrepancies uncovered on autopsy; only two discrepancies would have influenced patient management, and none would have altered patient outcome. most of the unexpected diagnoses were infections, and the rest included non-infective endocarditis, gvhd, and gastrointestinal and neurologic diagnoses. the authors concluded that clinical diagnosis alone might be appropriate for withdrawal of care decision-making in these patients. infectious and noninfectious pulmonary diseases are commonly found on postmortem autopsy studies in patients with hematological malignancy and hsct recipients. despite the technological advances in diagnostic testing and imaging modalities, obtaining an accurate clinical diagnosis remains difficult and is often not possible until autopsy. major diagnostic discrepancies between clinical premortem diagnoses and postmortem autopsy findings have been reported in patients with hematologic malignancy. the most common missed diagnoses are due to opportunistic infections and cardiopulmonary complications. these findings underscore the importance of enhanced surveillance, monitoring, and treatment of infections and cardiopulmonary disorders in these patients. autopsies remain important in determining an accurate cause of death and for improved understanding of diagnostic deficiencies, as well as for medical education and quality assurance. the goldman classification of diagnostic discrepancies was applied. major discrepancies included two categories (class i referred to missed diagnoses that would have led to a change in management with possible increased survival or cure; class ii referred to major missed diagnoses that would not have impacted survival because no treatment was available, treatment was given even though the diagnoses was unknown, or the patient refused further treatment). minor discrepancies included two categories (class iii referred to missed minor diagnoses related to the terminal disease process but not directly related to the cause of death, and class iv referred to missed minor diagnoses that did not contribute to the cause of death but ultimately may have been significant had the patient survived the major illness) congestive heart failure - acute pancreatitis - pulmonary embolism - invasive pulmonary aspergillosis - pulmonary edema - class ii ( . %) pneumonia - hematological disease - invasive pulmonary aspergillosis - pulmonary hemorrhage - other forms of aspergillosis - pulmonary candidiasis - class iii ( . %) hemosiderosis - secondary malignant neoplasm of kidney and renal pelvis - pleural effusion in conditions classified elsewhere - secondary malignant neoplasm of other unspecified digestive organ - pulmonary hemorrhage - class iv ( . %) pulmonary hemorrhage - pulmonary edema - gastrointestinal hemorrhage - nontraumatic intracerebral hemorrhage - pleural effusion in conditions classified elsewhere - secondary malignant lung neoplasm - chronic tubulo-interstitial nephritis - outcome of critically ill allogeneic hematopoietic stem-cell transplantation recipients: a reappraisal of indications for organ failure supports identification of poor prognostic features among patients requiring mechanical ventilation after hematopoietic stem cell transplantation pulmonary complications in lymphoma patients treated with high-dose therapy autologous bone marrow transplantation pulmonary complications after bone marrow transplantation: an autopsy study from a large transplantation center pulmonary complications in adult blood and marrow transplant recipients: autopsy findings causes of deaths in an oncology unit pathologists request autopsy revival trend reversal in the frequency of mycoses in hematological neoplasias: autopsy results from to invasive fungal infections in patients with hematologic malignancies in a tertiary care cancer center: an autopsy study over a -year period epidemiology and outcome of mould infections in hematopoietic stem cell transplant recipients pulmonary aspergillosis in an unselected autopsy series relationship between premortem and postmortem diagnosis in critically ill bone marrow transplantation patients fungal infections in cancer patients: an international autopsy survey auto psyidentified infections among bone marrow transplant recipients: a clinico-pathologic study of patients. bone marrow transplantation team trends in the postmortem epidemiology of invasive fungal infections at a university hospital clinical and radiologic predictors of invasive pulmonary aspergillosis in cancer patients: should the european organization for research and treatment of cancer/mycosis study group (eortc/msg) criteria be revised? clinical applicability of the new eortc/msg classification for invasive pulmonary aspergillosis in patients with hematological malignancies and autopsy-confirmed invasive aspergillosis bronchobronchiolitis obliterans as a complication of bone marrow transplantation: a clinicopathological study of eight autopsy cases voriconazole versus amphotericin b for primary therapy of invasive aspergillosis pneumocystis carinii pneumonia in patients with malignant haematological diseases: years' experience of infection in gimema centres successful treatment of invasive mould infection affecting lung and brain in an adult suffering from acute leukaemia effect of fluconazole prophylaxis on fungal blood cultures: an autopsy-based study involving patients with haematological malignancy invasive mycotic infections caused by chaetomium perlucidum, a new agent of cerebral phaeohyphomycosis mucormycoses in patients with hematologic malignancies: an emerging fungal infection autopsy case of disseminated trichosporon inkin infection identified with molecular biological and biochemical methods fatal pulmonary infection in a leukaemic patient caused by hormographiella aspergillata mucormycosis in hematologic malignancies: an emerging fungal infection successful management of cerebral and pulmonary mucormycosis with liposomal amphotericin b in a -year-old woman with acute lymphoblastic leukemia patients at high risk of invasive fungal infections: when and how to treat pulmonary complications in patients with haematological malignancies treated at a respiratory icu frequent detection of multidrug-resistant pneumonia-causing bacteria in the pneumonia lung tissues of patients with hematological malignancies lethal pulmonary hemorrhage caused by a fulminant stenotrophomonas maltophilia respiratory infection in an acute myeloid leukemia patient fatal cytomegalovirus pneumonia in patients with haematological malignancies: an autopsy-based case-control study cytologic manifestations of respiratory syncytial virus pneumonia in bronchoalveolar lavage fluid: a case report diagnosis of measles viral pneumonia in a patient with hodgkin's disease by reverse transcription-polymerase chain reaction of serum toxoplasmosis with hemophagocytic syndrome after bone marrow transplantation: diagnosis at autopsy diffuse alveolar hemorrhage syndromes diffuse alveolar hemorrhage in hematopoietic stem cell transplant recipients increasing incidence of diffuse alveolar hemorrhage following allogeneic bone marrow transplantation: cryptic etiology and uncertain therapy diffuse alveolar hemorrhage: an evolving problem pathogenesis of massive pulmonary hemorrhage in acute leukemia diffuse alveolar damage and hemorrhage in acute myelogenous leukemia treated with corticosteroids diffuse alveolar hemorrhage as a complication in patients with hematologic malignancies after chemotherapy: report of two cases and literature review diffuse alveolar hemorrhage after unrelated cord blood transplantation autopsy findings in umbilical cord blood transplant recipients pulmonary hemorrhage as a cause of death in allogeneic bone marrow recipients with severe acute graft-versus-host disease inflammatory response to infectious pulmonary injury diffuse alveolar hemorrhage in allogeneic bone marrow transplantation. a postmortem study corticosteroid therapy for diffuse alveolar hemorrhage in autologous bone marrow transplant recipients corticosteroids as adjunctive therapy for diffuse alveolar hemorrhage associated with bone marrow transplantation diffuse alveolar hemorrhage after allogeneic hematopoietic stemcell transplantation: treatment with recombinant factor viia diffuse alveolar hemorrhage: retrospective review of clinical outcome in allogeneic transplant recipients treated with aminocaproic acid histologic patterns of lung infiltration of b-cell, t-cell, and hodgkin lymphomas pulmonary leukostasis mimicking pulmonary embolism intravascular large b-cell lymphoma (ivlbcl): a clinicopathologic study of cases with special reference to the immunophenotypic heterogeneity of cd intravascular large b-cell lymphoma with fdg accumulation in the lung lacking ct/( )gallium scintigraphy abnormality pulmonary embolism in patients with acute leukemia and severe thrombocytopenia organizing pneumonia in patients with hematologic malignancies: a steroidresponsive lesion bronchiolitis obliterans organizing pneumonia in cancer: a case series acute fibrinous and organizing pneumonia following hematopoietic stem cell transplantation bronchiolitis obliterans after autologous bone marrow transplantation pulmo nary veno-occlusive disease following reduced-intensity cord blood transplantation pulmonary venoocclusive disease following bone marrow transplantation pulmonary venoocclusive disease accompanied by microangiopathic hemolytic anemia year after a second bone marrow transplantation for acute lymphoblastic leukemia pulmonary veno-occlusive disease following allogeneic peripheral blood stem cell transplantation for chronic myeloid leukaemia pulmonary veno-occlusive disease after bone marrow transplantation pulmonary veno-occlusive disease in an adult following bone marrow transplantation. case report and review of the literature pulmonary venoocclusive disease pulmonary veno-occlusive disease following hematopoietic stem cell transplantation: a rare model of endothelial dysfunction secondary alveolar proteinosis is a reversible cause of respiratory failure in leukemic patients a case of myelodysplastic syndrome complicated by pulmonary alveolar proteinosis with a high serum kl- level pulmonary alveolar proteinosis and disseminated atypical mycobacteriosis in a patient with busulfan lung difference in sero-diagnostic values among kl- -associated mucins classified as cluster causes of deaths in an oncologic intensive care unit: a clinical and pathological study of autopsies premortem clinical diagnoses and postmortem autopsy findings: discrepancies in critically ill cancer patients missed diagnosis in hematological patients -an autopsy study autopsy diagnoses of malignant neoplasms. how often are clinical diagnoses incorrect hodgkin's disease: correlation between causes of death at autopsy and clinical diagnosis high rate of discordance between clinical and autopsy diagnoses in blood and marrow transplantation clinical utility of autopsy after hematopoietic stem cell transplantation the value of the autopsy in three medical eras key: cord- -wydk p authors: even-or, ehud; nasereddin, adeeb; dinur schejter, yael; shadur, bella; zaidman, irina; stepensky, polina title: haploidentical stem cell transplantation with post-transplant cyclophosphamide for osteopetrosis and other nonmalignant diseases date: - - journal: bone marrow transplant doi: . /s - - - sha: doc_id: cord_uid: wydk p allogeneic hematopoietic stem cell transplantation (hsct) is curative for a variety of nonmalignant disorders including osteopetrosis, bone marrow failures, and immune deficiencies. haploidentical hsct is a readily available option in the absence of a matched donor, but engraftment failure and other post-transplant complications are a concern. post-transplant cyclophosphamide (pt-cy) regimens are gaining popularity and recent reports show promising results. we report our experience with nine pediatric patients with nonmalignant diseases who were transplanted from a haploidentical donor with pt-cy. from to , nine children with nonmalignant diseases underwent haploidentical hsct with pt-cy, two as a second transplant and seven as primary grafts after upfront serotherapy and busulfan-based myeloablative conditioning. patient’s diseases included osteopetrosis (n = ), congenital amegakaryocytic thrombocytopenia (n = ), hemophagocytic lymphohistiocytosis (n = ), and wiskott aldrich syndrome (n = ). two patients failed to engraft following upfront pt-cy transplants, one was salvaged with a second pt-cy transplant, and the other with a cd + selected graft. none of the patients suffered from graft-versus-host disease. three patients died from early posttransplant infectious complications and six patients are alive and well. in conclusion, haploidentical hsct with pt-cy is a feasible option for pediatric patients with nonmalignant diseases lacking a matched donor. allogeneic hematopoietic stem cell transplantation (hsct) is a curative treatment option for children with a variety of genetic nonmalignant disorders including osteopetrosis, bone marrow failure, and immune deficiencies [ ] [ ] [ ] . however, only - % of the patients have an available matched sibling donor, and the odds of finding a matched unrelated donor in international registries varies from % for white patients of european descent to as low as % in some ethnic groups [ ] . haploidentical hsct is a readily available alternative option for patients who do not have a matched donor, but engraftment failure, high rates of graft-versus-host disease (gvhd), delayed immune reconstitution and other posttransplant complications are of significant concern. the use of post-transplant cyclophosphamide (pt-cy) for in vivo tcell depletion has emerged as a promising strategy in the setting of haploidentical hsct. the availability, safety profile, effectiveness in reducing gvhd, and low cost of the procedure have contributed to the increasing popularity of this modality all over the world [ ] . for pediatric nonmalignant diseases, regimens with pt-cy for haploidentical hsct are increasingly adapted as well, and recent reports of haploidentical hsct with pt-cy for nonmalignant diseases show feasibility of this approach and promising results [ ] [ ] [ ] [ ] [ ] . our center introduced pt-cy regimens for pediatric haploidentical hsct in . in this case series, we report our experience with unmanipulated haploidentical hsct with pt-cy for nine pediatric patients with nonmalignant diseases, including five patients with osteopetrosis, two patients with congenital amegakaryocytic thrombocytopenia (camt), one patient with hemophagocytic lymphohistiocytosis (hlh), and one patient with wiskott aldrich syndrome (was). this retrospective study was performed on pediatric patients who underwent a haploidentical hsct with pt-cy for nonmalignant diseases from february to august at hadassah medical center. the study was approved by the institutional review board and informed consent was obtained from the families according to the declaration of helsinki. the collected data included patient age at hsct, gender, conditioning regimen, gvhd prophylaxis, time to neutrophil and platelet engraftment, signs and symptoms of gvhd, infectious complications, last chimerism status, and survival. neutrophil engraftment was defined as the first day of an absolute neutrophil count > /μl of consecutive days. platelet engraftment was defined as a platelet count > , /μl for at least days without transfusion support. the diagnosis of the patients was confirmed by gene mutation analysis with whole exome sequencing and confirmed in all cases by sanger sequencing. the diagnosis of osteopetrosis was supported by radiological studies. all patients in the cohort received myeloablative regimens (table ) . busulfan was given every h over days and the dose was adjusted according to blood levels to target an area under the curve (auc) of µm min/l. fludarabine was given at a dose of mg/m /day for days. for three of the patients thiotepa was added to the conditioning regimen in two doses of mg/kg on day − pre-hsct, and for one patient a low dose of cyclophosphamide ( . mg/kg) was added on days − and − . serotherapy was given to all patients. for the first two patients, thymoglobulin was given at a dose of . mg/kg/day for days (day − to − ) and for the rest of the patients alemtuzumab was given for days (day − to − ) at a dose of . mg/kg/day. for five of the patients rituximab was given at day − pre-hsct at a dose of mg/m . in two cases, we used the pt-cy modality for a salvage transplant and in two cases engraftment failure followed an upfront pt-cy transplant. for the salvage transplants, a treosulfan-based regimen was used in three of the cases, and in one case, due to the clinical status of the patient, a reduced intensity conditioning (ric) with fludarabine and cyclophosphamide was used (table ) . for the pt-cy transplants, all patients received an unmanipulated bone marrow graft and pt-cy was employed at a dose of mg/kg/day on days + and + . cyclosporine a and mycophenolate mofetil were started on day + for additional gvhd prophylaxis. all patients were treated with prophylactic trimethoprim/ sulfamethoxazole, acyclovir, and fluconazole according to our institutional guidelines. ursodeoxycholic acid was given for prevention of veno-occlusive disease (vod). viral loads of cytomegalovirus (cmv), epstein-barr virus, and adenovirus were followed weekly by blood pcr tests. cmv viremia was treated preemptively with ganciclovir or foscarnet. antiemetic treatment and pain control were given as necessary. donor chimerism was assessed weekly after engraftment by molecular testing of peripheral blood short tandem repeats (str) at the tissue laboratory. table . the underlying diseases of the patients included osteopetrosis (n = ), camt (n = ), hlh (n = ), and was (n = ). the median age of the patients at the time of hsct was . months (range . months to years). all grafts used for the pt-cy haplo-transplants were from bone marrow source and were collected from the donor at the operating room under general anesthesia. the median total nucleated cell count of the collected grafts was × cells/kg (range . - . × cells/kg) and the median cd + cell count was . × cells/kg (range . - . × cells/kg). two of the hsct in the cohort were transplanted with upfront cd + selected haploidentical grafts that after engraftment failure were salvaged with a pt-cy transplant. one additional cd + selected graft was used as a salvage transplant after engraftment failure of a pt-cy transplant. the grafts used for all three cd + selected haploidentical transplants were pbsc after mobilization with gcsf. the cell counts of the cd + depleted grafts are depicted in table . the first two pt-cy transplants done at our center, both for osteopetrosis patients, were second transplants following primary engraftment failure after cd + selected haploidentical transplants. the pt-cy modality was chosen for these two salvage transplants with the hope to achieve engraftment using a different approach. one of the patients survived and has complete donor chimerism and the other died days post second transplant from sepsis. another two patients failed to engraft following upfront haploidentical hsct with pt-cy. one, a patient with camt, successfully engrafted after a second transplant from the other parent, which was performed on day + after the first procedure. in this case, considering the primary and complete failure to engraft following the first transplant we opted to use the other parent for the second transplant while repeating the pt-cy modality. at the time of publication, almost years post his second hsct, he is well, with complete donor chimerism and cured from his disease. the other, a patient with osteopetrosis, initially engrafted after his first transplant but soon after developed a secondary graft failure with chimerism rapidly dropping down to % months post hsct despite rapid weaning from immunosuppressive treatment. he underwent a second transplant days after the first hsct with a peripheral stem cell collected graft after cd + selection from the same donor and successfully engrafted. in this case, we opted to use the same donor for the second transplant since there were no signs of rejection of the graft, rather a process of secondary graft failure, and in order to enhance the engraftment process we decided to change the transplant modality and give a robust amount of stem cells using a cd + selected pbsc graft. at the time of publication, months post his second transplant he is alive and well and has mixed chimerism with % donor cells on his latest str test. five patients in the cohort underwent a single haploidentical transplant with pt-cy, three of them survived and have complete donor chimerism, and two died from early post-transplant infectious complications. a total of five patients had cmv reactivations, but only one progressed to a clinically significant disease. three patients had bacterial infections during the post-transplant neutropenia, two of which progressed to severe sepsis and death. none of the patients suffered from any form of gvhd. the six survivors are all cured of their disease, alive and well, with a median follow-up time of months (range - months). of the three surviving patients with osteopetrosis, one was already blind prior to hsct (patient # ), and on latest follow-up he is years old, developing well and attending a school for children with special needs. the second patient (patient # ) has severe optic damage with blindness in one eye and decreased vision in the other eye on his latest ophthalmologic assessment. otherwise, his neurological examination is normal. the third patient (patient # ) has preserved vision and no nystagmus despite significant bilateral optic nerve atrophy. he was years old on his latest follow-up and achieved all developmental milestones. all three patients have no known hearing deficits. three patients died, all from early post-transplant infectious complications ( table ). two of the deceased patients had osteopetrosis, both died from overwhelming klebsiella pneumonia sepsis shortly after transplant; one on day + post his second transplant following engraftment failure and the other on day + post hsct. the third deceased patient, a patient with hlh, suffered from vod and severe cmv pneumonitis after transplant and eventually died on day + post hsct from pulmonary bleeding and respiratory failure. in this paper, we report our experience with haploidentical hsct with upfront serotherapy, busulfan-based myeloablative conditioning and pt-cy in nine pediatric patients with nonmalignant diseases. the modality of pt-cy for in vivo t-cell depletion without need for an additional graft manipulation increased the availability and popularity of the haploidentical option over the world [ ] . furthermore, the advantages of this cost-effective modality have set alight the debate of preference of an haploidentical family donor over an acceptably matched unrelated donor for various hsct indications [ ] . recent reports focusing on haploidentical hsct with pt-cy in pediatric patients with nonmalignant diseases offer different promising approaches using a variety of conditioning regimens [ ] [ ] [ ] [ ] [ ] . klein et al. reported hsct of children with nonmalignant disorders showing very good outcomes with limited gvhd, excellent engraftment outcomes and no treatment related mortality using a ric regimen with fludarabine, melphalan and upfront serotherapy with alemtuzumab [ ] . another recent report by mallhi et al. of pediatric nonmalignant patients also used a ric regimen including low dose cyclophosphamide ( mg/kg), fludarabine and a low dose of total body irradiation ( - gy) and showed favorable outcomes but relatively high rates of gvhd [ ] . kurzay et al. used a treosulfanbased myeloablative conditioning regimen with upfront serotherapy and an addition of thiotepa and low dose cyclophosphamide in a recent report of pediatric patients with nonmalignant diseases showing good overall survival, low rate of gvhd and few infectious complications [ ] . finally, neven et al. recently reported haploidentical hsct with pt-cy for pediatric nonmalignant diseases using a busulfan-based myeloablative conditioning regimen with upfront alemtuzumab showing a high engraftment rate ( of patients engrafted) and a years overall survival rate of . % [ ] . since successful engraftment is a major concern in haploidentical transplants, especially in osteopetrosis patients with a damaged bone marrow, we elected to adopt a similar approach to neven et al. using a busulfan-based myeloablative regimen rather than a ric or a treosulfanbased regimen. in view of the relatively high treatment related mortality in our cohort ( %), a less aggressive conditioning regimen may be considered for this patient population. as our patient population is very young, we opted not to expose them to irradiation. rather, we gave serotherapy (thymoglobulin or alemtuzumab) for additional immune suppression (table ) . during , we switched from thymoglobulin to alemtuzumab following personal communications with other european centers with an aim to establish a standard practice for haploidentical transplants for pediatric inborn errors. the highly immunosuppressive properties of alemtuzumab are particularly appealing in the setting of nonmalignant hsct, where gvhd provides no clinical benefit and relapse of malignancy is not applicable. indeed, none of our patients developed any signs of acute or chronic gvhd as compared to the relatively high rates of acute and chronic gvhd reported by mallhi et al. using a ric conditioning without serotherapy [ ] . commonly used pediatric transplantation regimens include alemtuzumab doses of . - mg/kg given over - days [ ] . we gave a relatively low dose of alemtuzumab ( . mg/kg), and distal to the transplant day (days − to − ) in order not to compromise t-cell immunity and allow a swift immune reconstitution. however, considering the high rate of engraftment failure in our cohort, a higher dose may be considered for augmenting immune ablation. our cohort included a variety of nonmalignant diseases; five patients with infantile osteopetrosis, two patients with camt, one patient with hlh, and one patient with was. patients with infantile osteopetrosis usually presents in early infancy with a rapidly progressing disease, which in most cases may be curable by hsct [ , ] . we have recently published our experience with hsct for osteopetrosis patients with hla matched or one-locus mismatched donors, showing favorable outcomes [ ] . however, not all patients have an available matched donor or an acceptable mismatched donor. for these patients, haploidentical hsct may be a readily available option. historically, outcomes of haploidentical hsct for osteopetrosis patients were significantly worse than hsct from matched donors [ ] . improved outcomes were reported using mega-doses of cd + selected haploidentical grafts with an intensive, busulfan-based myeloablative conditioning, but post-transplant complications such as vod and respiratory failure were common [ ] . recent reports of successful haploidentical transplantations with alpha-beta t-cell depletion for osteopetrosis are promising [ ] , but this modality is not readily available in many centers due to its high cost, especially in developing countries with limited resources [ ] . reports of haploidentical hsct with pt-cy for osteopetrosis are scarce. in one report of three osteopetrosis patients by bahr et al. only one of the three had a successful transplant course and the other two failed to engraft [ ] . a recent publication by neven et al. of haploidentical hsct with pt-cy included five patients with osteopetrosis, all of which engrafted successfully and are alive and well [ ] . in our cohort, three out of the five osteopetrosis patients failed to engraft after the first haploidentical transplant. two failed to engraft after a cd + selected graft and the third failed after a pt-cy transplant. the high engraftment failure rate may be attributed to the damaged and sclerotic bone marrow niches of these patients. it is still too early to draw conclusions from these scarce number of patients, but engraftment failure seems to be a grave concern in these transplants. despite the positive experience we have with reduced toxicity treosulfan-based conditioning in matched donor transplants for osteopetrosis, a myeloablative busulfan-based regimen was chosen for the haplotransplants because of the concern of engraftment. however, weighed against the relatively high treatment related mortality in our cohort ( %), a less aggressive conditioning regimen may be considered for this patient population. furthermore, until optimal timing and dosing of alemtuzumab are determined in prospective trials, a higher dose should be considered to augment engraftment, especially for patients with osteopetrosis. finally, in view of these results, our current institutional policy is to prefer an unrelated donor if available, even with a one-locus mismatch, over a haploidentical donor for these patients. two of the patients in our cohort were transplanted for camt, an inherited bone marrow failure syndrome characterized by the absence of megakaryocytes in the bone marrow [ , ] . literature reporting hsct with alternative donors for camt is very scarce. a report of five patients with camt who underwent umbilical cord blood transplantations showed promising results [ ] . to our knowledge there are no reports of haploidentical transplants with pt-cy for this indication yet. our two camt patients are both cured of their disease, alive and well. the first engrafted well with no complications but the second required a second transplant due to engraftment failure. of note, a second successful transplant was performed using pt-cy from another donor. it is difficult to draw conclusions from only two cases but haploidentical hsct with pt-cy seems a feasible option for camt patients. furthermore, this case demonstrated the feasibility of re-using the same modality of in vivo t-cell depletion using another haploidentical donor for a salvage, second transplant. one patient in our cohort was transplanted for familial hlh, a life-threatening systemic hyperinflammatory syndrome [ ] . hsct for hlh patients is challenging and associated with high rates of toxicities and mortality, likely related to the underlying inflammatory state of these patients [ ] . our patient did not survive and died days post hsct from vod and cmv pneumonitis. however, there are some recent reports with promising results of successful haploidentical transplants with pt-cy for patients with hlh [ , ] . one successful transplant in our cohort was of a patient with was, a rare x-linked disorder characterized by a triad of immunodeficiency, eczema, and thrombocytopenia. hsct is a curative treatment for was, with excellent results for patients with hla-matched family or unrelated donors [ ] . a recent report by yue et al. showed promising results in a case series of five patients who underwent haploidentical hsct with pt-cy for was [ ] . in conclusion, in our experience, haploidentical hsct with pt-cy is a feasible option for pediatric patients with nonmalignant diseases, but with significant risks of posttransplant infectious complications and engraftment failure. hematopoietic stem cell transplantation for osteopetrosis inherited bone marrow failure syndromes: considerations pre-and posttransplant hematopoietic stem cell transplantation for primary immunodeficiencies hla match likelihoods for hematopoietic stem-cell grafts in the u.s. registry post-transplant cyclophosphamide, a promising anti-graft versus host disease prophylaxis: where do we stand? haploidentical hematopoietic stem cell transplantation with post-transplant cyclophosphamide for primary immunodeficiencies and inherited disorders in children t-cell replete haploidentical bone marrow transplantation and post-transplant cyclophosphamide for patients with inborn errors haploidentical stem cell transplantation with post-transplant cyclophosphamide for primary immune deficiency disorders in children: challenges and outcome from a tertiary care center in south india alternative-donor hematopoietic stem cell transplantation with post-transplantation cyclophosphamide for nonmalignant disorders hla-haploidentical hematopoietic cell transplantation for treatment of non-malignant diseases using nonmyeloablative conditioning and post-transplant cyclophosphamide outcomes of haploidentical transplant compared with matched donor allogeneic stem cell transplant principles of alemtuzumab immunoablation in hematopoietic cell transplantation for nonmalignant diseases in children: a review one disease, many genes: implications for the treatment of osteopetroses successful hematopoietic stem cell transplantation for osteopetrosis using reduced intensity conditioning hematopoietic stem cell transplantation for infantile osteopetrosis hla-haploidentical blood progenitor cell transplantation in osteopetrosis transplantation of haploidentical tcrab-depleted hematopoietic cells allows for optimal timing and sustained correction of the metabolic defect in children with infantile osteopetrosis transplantation of hematopoietic stem cells for primary immunodeficiencies in brazil: challenges in treating rare diseases in developing countries haploidentical transplantation with post-transplant cyclophosphamide following reduced-intensity conditioning for osteopetrosis: outcomes in three children congenital amegakaryocytic thrombocytopenia: clinical presentation, diagnosis, and treatment allogeneic hematopoietic stem cell transplantation for inherited bone marrow failure syndromes durable engraftment and correction of hematological abnormalities in children with congenital amegakaryocytic thrombocytopenia following myeloablative umbilical cord blood transplantation pediatric hemophagocytic lymphohistiocytosis (hlh) reduced-intensity conditioning for hematopoietic cell transplant for hlh and primary immune deficiencies successful haploidentical stem cell transplant with posttransplant cyclophosphamide for hemophagocytic lymphohistiocytosis outcomes after allogeneic transplant in patients with wiskott-aldrich syndrome posttransplant cyclophosphamide for haploidentical stem cell transplantation in children with wiskott-aldrich syndrome acknowledgements we thank our patients and their families for putting their trust in our care and allowing us to publish this paper. we thank our departmental nursing and administrative staff for their devoted work. we also thank professor zeev rotstein, director of hadassah medical center, for his support of our department. shadur b's position is supported by the australian government research training program scholarship and hadassah, australia. this work was supported by the deutsche forschungsgemeinschaft (discovery and evaluation of new combined immunodeficiency disease entities (decide); grant dfg wa / - ) and the era-net erare consortium euro-cid. conflict of interest the authors declare that they have no conflict of interest.publisher's note springer nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations. key: cord- - avisuge authors: versluys, a.birgitta; rossen, john w.a.; van ewijk, bart; schuurman, rob; bierings, marc b.; boelens, jaap j. title: strong association between respiratory viral infection early after hematopoietic stem cell transplantation and the development of life-threatening acute and chronic alloimmune lung syndromes date: - - journal: biol blood marrow transplant doi: . /j.bbmt. . . sha: doc_id: cord_uid: avisuge alloimmune lung syndromes (allo-ls), including idiopathic pneumonia syndrome, bronchiolitis obliterans syndrome, and bronchiolitis obliterans organizing pneumonia, are severe complications after hematopoietic stem cell transplantation (hsct). in our cohort of pediatric patients, had allo-ls ( . %), with ideopathic pneumonia syndrome and with bronchiolitis obliterans syndrome. multivariate analysis showed that respiratory viral infection early after hsct is an important predictor for the development of allo-ls (p <. ). this was true for all viruses tested. in multivariate analysis, allo-ls was the only predictor for higher mortality (p = . ). paradoxically, prolonged administration of immunosuppressive agents because of acute graft-versus-host disease had a protective effect on the development of allo-ls (p = . ). we hypothesize that early infection of the respiratory tract with a common cold virus makes the lungs a target for alloimmunity. pulmonary complications are common after allogeneic hematopoietic stem cell transplantation (hsct). between % and % of adult hsct recipients reportedly experience pulmonary complications, representing a major cause of mortality [ ] [ ] [ ] [ ] . in children undergoing hsct, the incidence of pulmonary complications varies from % to %, and onset is a poor prognostic event carrying a significantly increased risk of mortality [ , ] . at one time, most pulmonary complications were directly related to infection; today, however, noninfectious pulmonary complications, such as idiopathic pneumonia syndrome (ips) and bronchiolitis obliterans syndrome (bos), are seen more frequently [ , , ] . respiratory virus (rv) infections occur in %- % of hsct recipients. most previous studies have examined the progression from upper respiratory tract infection (urti) to lower respiratory tract infection (lrti) and described the risk factors for this progression [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] . some have reported an association of early rv infection with late, obstructive lung injury [ ] . an association between rv infection and alloimmunity in lung transplant recipients was recently reported [ ] . lung transplant recipients develop more acute and chronic graft rejection after common rv infection early (\ days) after transplantation [ ] . isolated alloimmune lung disease (ie, bos or ips) after hsct suggests a specific trigger making the lung a target organ for alloreactivity. this is in line with the -step process reflecting the current view of the development of alloreactivity: ( ) tissue damage, resulting in ( ) release of inflammatory cytokines, resulting in ( ) activation and influx of t lymphocytes [ ] . we speculated that the presence of a common rv might trigger alloimmune lung syndrome (allo-ls) in hsct. we prospectively studied the influence of these rvs on the development of allo-ls and overall survival (os) in a cohort of pediatric hsct recipients. all patients who underwent allogeneic hsct between january and may at the pediatric hematology and immunology department of the wilhelmina children's hospital/university medical center were included in this prospective study. patients were enrolled in the hsct protocol after providing written informed consent for the hsct and the research protocol. all patients received antiemetic drugs. prophylactic anticonvulsive therapy (clonazepam) was given to those patients receiving busulfan. antibiotic prophylaxis involved daily ciprofloxacillin and fluconazole from the start of conditioning until the resolution of neutropenia ( days of . , neutrophils/ml). additional prophylaxis against streptococcus viridans in the mucositis phase was given with cefazoline. starting month after transplantation, cotrimoxazole times a week was given as pneumocystis carinii pneumonia prophylaxis. only in cases of positive serology for herpes simplex virus was prophylaxis (with acyclovir) administered. no prophylaxis for other viruses was given. igg levels were checked every weeks; intravenous immunoglobulin was given only to those patients with an igg level \ g/l. graft-versus-host disease (gvhd) prophylaxis consisted of cyclosporine (aiming for a trough level of - mg/l, based on national protocol guidelines), supplemented with methylprednisolone (mp; mg/ kg/day for days) in patients receiving a cord blood (cb) transplant, or methotrexate (short course, mg/ m on days , , and ) in patients receiving an unrelated bone marrow (bm) or peripheral blood stem cell (pbsc) transplant. in patients receiving an unrelated donor graft (cb, bm, or pbsc), antithymocyte globulin (atg) serotherapy was administered until day - , with atg-fresenius for patients with acute lymphoblastic leukemia and thymoglobulin for all other indications. to monitor bacterial colonization, nose/throat swabs and stools were cultured weekly and processed in accordance with standard microbiological procedures. up to june , we tested for galactomannan (platelia aspergillus enzyme immunoassay; bio-rad, hercules, ca) in cases of suspected aspergillus infection, based on such clinical symptoms as prolonged fever during systemic broad antibiotic therapy and radiologic findings. after june , we routinely monitored galactomannan twice weekly. plasma was tested weekly for epstein-barr virus (ebv), cytomegalovirus (cmv), human herpes virus (hhv ), and adenovirus dna positivity by real-time polymerase chain reaction (pcr) (see next section). in patients deemed positive (viral load . cp/ml), this test was done twice a week. adenovirus (viral load . cp/ml) was treated preemptively with cidofovir. cmv (viral load . cp/ml) was treated preemptively with foscavir or ganciclovir. depending on the viral load, the immunosuppressive regimen, and signs of posttransplantation lymphoproliferative disease, ebv was treated preemptively with anti-cd (rituximab). before august , nasal pharyngeal aspirate (npa) samples were obtained for pcr only in the presence of symptoms of a urti or lrti, and then only up to day posttransplantation. from august onward, we performed surveillance studies on npa samples of all patients admitted to our hsct unit. reversetranscriptase (rt)-pcr was done for all common rvs (see later). we repeated the npa weekly in patients negative for rv and twice weekly in patients positive for rv. nucleic acids were extracted using the total nucleic acid protocol with the magna pure lc nucleic acid isolation system (roche diagnostics, basel, switzerland). for detection of rna viruses, cdna was synthesized using multiscribe rt and random hexamers (applied biosystems, foster city, ca). detection of viral and atypical pathogens was performed in parallel, using real-time pcr assays specific for the following viruses: cmv; ebv; hhv- ; respiratory syncytial virus a and b; influenzavirus a and b; parainfluenzavirus - ; rhinoviruses; adenoviruses; human coronaviruses oc , nl , and e; human metapneumovirus; mycoplasma pneumoniae; and chlamydia pneumoniae. real-time pcr procedures were performed as described previously [ ] . in brief, samples were assayed in duplicate in a -ml reaction mixture containing ml of cdna, . ml of taqman universal pcr master mix (applied biosystems), - nmol/l of the forward and reverse primers, and - nmol/l of each probe. all samples had been spiked before extraction with an internal control virus (murine encephalomyocarditis virus [rna virus] and porcine herpesvirus [dna virus]) to monitor for efficient extraction and amplification, essentially as described previously [ ] . the cycle of threshold (ct) gives an impression of the quantity of the viral load (ie, a semiquantitative value). all patients were observed for signs of respiratory disease early and late after transplantation. all clinical symptoms were recorded. in patients with urti symptoms, npa samples were obtained and tested for rv infection by pcr (see earlier). in patients with signs of lrti, chest x-rays were obtained. other tests, performed as indicated, included bronchoalveolar lavage (bal) for broad infectious screening with bacterial/fungal cultures, viral pcr, and galactomannan, as well as high-resolution computed tomography (hrct) scans. in cases of suspected allo-ls, hrct and bal were always performed (see definitions of disease shortly). because performing pulmonary function tests (pfts) is difficult in young children, in our cohort routine pfts before transplantation were performed only in children aged $ years. pfts also were performed in all children with such respiratory symptoms as shortness of breath, dry cough, and tachypnea after discharge. in patients with a diagnosis of allo-ls, pfts were repeated at least monthly until the disorder resolved. patients aged $ years underwent spirometry testing and, when technically possible, body plethysmography and co diffusion testing according to european respiratory society guidelines [ ] . total lung capacity (tlc) and forced expiratory volume in second (fev ) were expressed as percentage of the predicted normal value, using published equations for children and adults [ ] , giving tlc % predicted and fev % predicted. a tlc % predicted of \ % was designated a ''restrictive'' pattern; an fev % predicted of \ % and fev /forced vital capacity (fvc) of\ %, an ''obstructive'' pattern; a tlc % predicted of \ % and fev /fvc of \ %, a ''mixed'' pattern; and co diffusion of \ %, ''impaired diffusion.'' urti was defined as rhinorrhea and/or dry cough only. lrti/pneumonia was defined as cough and/or fever and pulmonary infiltrates on chest x-ray, with elevated c-reactive protein and/or positive microbiological cultures from sputum, bal fluid, or blood. ips was defined as the presence of acute bilateral pulmonary infiltrates with cough, dyspnea, and hypoxemia in the absence of infection (excluding an rv) or heart failure. by this definition, ips included such entities as diffuse alveolar bleeding and periengraftment syndrome [ ] . bos was defined as typical hrct changes, such as bronchial wall thickening, air trapping, and mosaic parenchymal attenuation [ ] , in the absence of signs of infection and, whenever pulmonary function testing could be done, abnormal pulmonary function test results (ie, decrease in fev of . % or in fev /fvc of \ %). bronchiolitis obliterans organizing pneumonia (boop) was defined as restrictive pft (if pft were done) and consolidation on chest x-ray [ ] . allo-ls was defined as ips, bos, and boop, subdivided into acute (ips) and chronic (bos/boop) forms. in general, urti was not treated; only in the patients with influenza a was a neuraminidase inhibitor administered. lrti/pneumonia was treated with empiric antibiotic therapy (vancomycin and ceftazidime). whenever a bacterial pathogen was found, therapy was adjusted according to antibiotic resistance. in patients with probable or proven aspergillus spp, voriconazole was administered; if no response to voriconazole was noted (progressive clinical or radiologic findings), granulocyte transfusions were given. allo-ls was treated with mp mg/kg/day i.v. for days and mg/kg/day thereafter, tapering by % per week to . mg/kg/day. the mp pulses were repeated every weeks until recovery, up to a maximum of courses. recovery was defined as normalization of pfts and/or resolved symptoms, with no extra oxygen requirement. in between the subsequent courses of mp, prednisone . mg/kg/day was given. other immunosuppressive agents (usually cyclosporine) were continued. in addition, azythromycin was given, because of its suggested immunomodulatory effect [ ] . along with immunosuppressive therapy, supportive care was provided, with extra oxygen and mechanical ventilation when necessary. igg level was maintained above g/l. the primary endpoint of this study was the development of acute and chronic allo-ls. the secondary endpoint was os. differences between the rv-positive and rvnegative groups were tested using pearson's c test. results with a p value \. were considered statistically significant. the duration of follow-up was the time to the endpoints, the development of an allo-ls and death, or the last assessment for survivors. to analyze risk factors for outcomes, we considered variables associated with the recipient (age at transplantation, sex, cmv serology, rv positivity, single/multiple viruses), the disease (malignant vs nonmalignant), the donor/transplantation technique (cell source, hla disparity, donor relationship, conditioning regimen), hsct complications (allo-ls, acute gvhd [agvhd], cmv and adenovirus plasma dna positivity, venoocclusive disease), and relapse. to examine the influence of the various viruses on the primary endpoint, rhinovirus was compared with the other viruses, and multiple viral infection was compared with single viral infection. in the analyses, we tested for allo-ls as a group (ips bos/boop) based on the hypothesis that early viral infection might be a trigger for both acute and chronic allo-ls. in addition, we tested both syndromes separately (ie, bos/boop excluding ips from the analyses, and ips excluding bos/boop). associations between variables (including recipient, disease, and hsct technique) and the primary endpoint were evaluated using cox proportional hazard models. dichotomous outcomes (eg, allo-ls: yes/no) were used as dependent variables, and predictors were used as independent variables. univariate predictors of outcome with a p value \. were used for multivariate analysis. results are expressed as hazard ratios (hrs) and corresponding % confidence interval (cis). cis not including were considered statistically significant. analyses for the association between hsct complications and the primary endpoint (allo-ls) as well as the secondary endpoint (os) were done using logistic regression. dichotomous outcomes (eg, allo-ls or survival: yes/no) were used as dependent variables, and predictors were used as independent variables. univariate predictors of outcome with a p value \. were used for multivariate logistic regression analysis. results are expressed as odds ratio (ors) and corresponding % cis. cis not including were considered statistically significant. probabilities of allo-ls and os were calculated using the kaplan-meier estimate; the -sided log-rank test was used for comparisons. all statistical analyses were performed using spss . (spss inc, chicago, il). a total of patients were included in the study, from january to august (before routine npa testing), and after from august to may . six patients who underwent transplantation during this period were excluded from the study because they experienced autologous recovery (n ), early graft rejection (within month after transplantation; n ), or early death (before engraftment; n ) and thus were considered not prone to alloreactive disease. the median age at transplantation was . years (range, months to years), and body weight ranged between and kg. baseline characteristics of the rv-positive and rv-negative groups are shown in table . no significant differences between the groups were evident, although there were slightly more matched donor transplants in the rv-negative group and more cb donors in the rv-positive group. in this cohort of patients, ( %) had an rv infection. the median day of onset was day posttransplantation (range, day to day ). symptoms were usually mild. the majority of patients with rv infection (n ) had urti symptoms only. eleven patients required extra oxygen, and patient needed ventilator support (associated with a bacterial infection). two patients, both with influenza a infection, were treated with a neuraminidase inhibitor; all other patients experienced spontaneous clinical recovery within - days. although symptoms disappeared, virus was detected in npa samples for weeks to months afterward, with high viral loads (pcr ct values of - ; see patients and methods). thirty-eight patients had a single rv, and patients had multiple viruses. in patients, no rv was detected, but the clinical picture was typical for rv infection. these patients had mild respiratory symptoms (rhinorrhea) with no other cause, and all recovered spontaneously. the distribution of the various viruses is shown in table . thirty patients were diagnosed with allo-ls ( . %), with bos ( . %) and with ips ( . %). no patient developed boop. one patient presented with pulmonary hypertension with histologically proven vasculopathy, with lymphocyte infiltration that responded to immunosuppressive agents. we considered this patient to have ips. for the patients with allo-ls, the median time of onset was weeks (range, - weeks) after transplantation. ips occurred earlier, with a median time of onset of weeks (range, - weeks); bos developed later, after a median of weeks (range, - weeks). in univariate analysis, rv positivity, cb stem cell graft, and a chemotherapy-based conditioning regimen were predictors for the development of allo-ls (table ) the median duration from rv positivity and the development of allo-ls was weeks (range, . - weeks) ( figure a ). the timing of development of rv positivity seems to be important as well. patients who were rv-positive early after transplantation (before the median of day ) had a slightly greater likelihood of developing allo-ls than those who became rv-positive after day (hr, . ; % ci, . - . p . ). univariate analysis of the influence of hsctassociated complications on allo-ls showed that adenovirus reactivation (or, . ; % ci, . - . ; p . ) was predictive of allo-ls. agvhd grade ii-iv in other organs appeared to be a negative predictor (or, . ; % ci, . - . ; p . ). in multivariate analysis, only agvhd remained a strong predictor for preventing allo-ls (or, . ; % ci, . - . ; p . ). the influence of agvhd on the development of allo-ls for the whole group and for the rv-positive patients is shown in figure b and c. in this study, agvhd clearly developed before the onset of allo-ls. the mean time to onset was weeks for gvhd (range, - weeks) and weeks for allo-ls (range, - weeks). in particular, in the rv-positive group, agvhd grade ii-iv in another organ was strongly protective against the development of allo-ls. we found no influence of the different individual viral species, or of the presence of a single virus or multiple viruses, on the development of allo-ls (data not shown). all patients who developed allo-ls were treated according to the protocol with mp pulse therapy, as discussed earlier. all patients demonstrated prompt initial improvement of clinical symptoms. os was % ( / ) after a median follow up of weeks (range, - weeks). cause of death was relapse in patients ( . %) and nonrelapse mortality in patients ( . %). fourteen of the patients with allo-ls died ( %), all from transplantation-related causes: from refractory agvhd, from invasive fungal infection, from adenovirus disease, from sudden cardiac death of unknown cause, and from ongoing lung disease. univariate analysis identified adenovirus reactivation (or, . ; % ci, . - . ; p . ) and the development of allo-ls (or, . ; % ci, . - . ; p . ) as predictors for lower survival. relapse had no significant influence on os (hr, . ; % ci, . - . ; p . ). in multivariate analysis, only allo-ls remained a predictor in this cohort (or, . ; % ci, . - . ; p . ). the impact of allo-ls on os is depicted in figure d . our cohort of patients had a high incidence ( %) of early rv infection, occurring at a median of days after hsct. rhinovirus infection was the most common rv detected. the rv infections usually had a mild clinical course, and most patients experienced spontaneous recovery within weeks. rv infection occurring during the first days after hsct appeared to be the sole predictor for the development of acute and chronic allo-ls, which was found in . % of the patients. all patients had recovered from their initial urti symptoms before a new episode of respiratory symptoms occurred, leading to the diagnosis of allo-ls. the presence of a single rv or multiple rvs, or the presence of rhinovirus and other (nonrhinovirus) rvs, was not associated with the development of allo-ls. paradoxically, agvhd had a protective effect against the development of allo-ls, likely resulting from the prolonged immunosuppressive therapy in the patients with agvhd. all of the patients with allo-ls initially exhibited good clinical response to mp pulse therapy. the development of allo-ls was associated with high mortality, however. a possible weakness of our study is that we changed our policy on testing for rv during the study period. in the early phase of the study, we tested npa samples for rv only in those patients exhibiting symptoms. later in the study period, once the significance of rv was recognized, weekly surveillance assays were done in all patients. these surveillance assays identified rv-positive patients without symptoms at the time of sampling; however, all of these patients developed urti symptoms within days after positive sampling, and so these rvs would ultimately have been detected regardless of the testing policy. thus, we believe that we did not miss any rv-positive patients in the presurveillance period, and that the change in testing policy had no major impact on our results (data not shown), the only difference being that the median time to rv positivity would have been shorter had we monitored the whole group routinely. three patients with symptoms and a clinical course typical of viral urti were considered rv-positive despite negative rv-pcr results. in these patients, symptoms might have resulted from a virus not detectable by our pcr panel. it would be interesting to test these for more recently identified viruses, such boca and wu/ki. because of the obvious symptoms and the fact that other study groups have included similar patients in their analyses, we decided to do so as well [ ] . had we considered these patients rv-negative in our analysis, the results would have been the same (only of the patients developed allo-ls). we realize that we may well have missed some mild urti symptoms in the period between discharge and day and so do not have full data on rv positivity after discharge from the hospital. no patient who was rv-pcr-negative on discharge developed allo-ls, however. acquisition of rv very early after hsct appears to be important for the development of allo-ls; the patients who were rv-positive within days of hsct tended to be more susceptible to allo-ls. a remark about the definition of ips is warranted. in the consensus definition of ips (established by a national institutes of health workshop), all infectious agents, including rvs, should be excluded. in our patients, we observed prolonged shedding of rv for months, and thus we could not formally diagnose ips. in all patients, the initial urti symptoms disappeared spontaneously within - weeks, however. subsequently, after a period of at least days without significant respiratory problems, symptoms of hypoxia and/or airway obstruction recurred. we believe that this represents not a direct progression of viral infection, but rather a combination of several factors in which alloreactivity (triggered by tissue damage because of persistent viral infection) plays a pivotal role. therefore, we chose to define ips as discussed earlier, not taking into account the presence of an rv identified by pcr as was done in this study. moreover, the formal definition of ips was promulgated in an era when molecular diagnosis of rv was not yet available. we hypothesize that rvs may contribute to the pathogenesis of any allo-ls as follows. the rv damages the respiratory epithelium, causing an inflammatory response at the time of immune recovery. normally ips and bos/boop are viewed as distinct clinical entities, and so we first studied them separately. because we noted the same strong associations among rv positivity, gvhd, and os in the groups, we combined both ips and bos/boop in subsequent analyses. we also combined bm and pbsc sources in our analysis. it would be interesting to evaluate patients receiving pbscs as a separate group, because of their apparent higher risk of chronic gvhd (cgvhd). the small number of patients our cohort who received pbscs (n ) precludes meaningful analysis, however. numerous studies have explored the incidence and outcome of both nosocomial and community-acquired rv infections after hsct [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] . reported incidence varies from % to %. this wide range of incidence can be explained by differences among studies in the definition of rv infection, the period of monitoring for rv infection (eg, inpatients/outpatients, seasonal influence), and sensitivity of the analysis methods used. in our cohort, the high incidence of rv urti ( %) might be attributed to our close monitoring for respiratory symptoms and performance of rt-pcr surveillance assays. in addition, nosocomial rv infections were frequently observed, and genotyping studies suggested that these had spread throughout the ward during the study period, also contributing to the relatively high incidence (data not shown). literature data on the morbidity associated with rv infection after hsct are conflicting. some groups found no progression of viral urti to lrti in patients after hsct [ , ] , whereas others reported progression in up to % of patients [ , , , , ] . in our cohort, we found no direct progression to lrti. almost all patients had mild urti symptoms and recovered spontaneously. the median day of onset of rv infection was only days in our cohort, compared with at least days in previous studies. progression to lrti might be expected in patients with rv infection early after transplantation, because of poor immune status, but this was not seen. in our opinion, it is more likely that the moment of immune recovery defines the onset of symptoms. what we define as ips in this study (ie, symptoms after a period of quiescent rv infection) might have been reported in other studies as progressive viral pneumonia, with symptoms occurring at the onset of rv infection later after hsct, when some immune recovery has already occurred. the reported incidence of ips after hsct ranges from % to % of patients [ , , , ] , and that of bos ranges from % to % [ , , , ] . most reported data are from adult studies. the combined incidence of allo-ls in our cohort of . % is comparable to that reported in the literature. our incidence of ips is relatively high, most likely because our cohort included a high number of early rv-positive patients. we found a strong association between rv infection and the development of allo-ls. to the best of our knowledge, this is the first report of such a strong association between rv infection and life-threatening allo-ls. earlier, erard et al. [ ] described a relationship between rv infection during the first days after hsct and a decline in airflow leading to increased overall mortality. the decline in airflow was detected immediately after infection and did not return to baseline values, suggesting sustained airway inflammation leading to permanent loss of lung function. pulmonary disease was much more severe in our cohort compared with the cohort of erard et al. [ ] . this discrepancy can be explained by the early moment of rv infection, when immune recovery has not yet occurred and the persistent rv likely causes more tissue damage, ultimately resulting in a stronger inflammatory response. the absence of immunity at the time of primary rv infection also might explain the absence of an immediate decline in clinical lung function. pulmonary function deteriorated only after at least weeks after rv infection, when the first signs of immune recovery were evident. it would be interesting to routinely perform pfts earlier after hsct, but for reasons of hygiene and patient comfort, we decided not to do this in the present study. most previous studies have found an association between the presence of gvhd and the development of allo-ls [ , , , ] . in those studies, agvhd was associated with ips, but other risk factors, including conditioning regimen and infection, might have been involved as well. cgvhd is considered an important risk factor for bos in adults. alloreactive t cells, in the context of agvhd or cgvhd, play an important role in bos [ ] . bos is also seen in the absence of cgvhd in %- % of patients, however [ ] . this percentage may be different in children, who are less susceptible to cgvhd [ ] . our finding that agvhd grade ii-iv in other organs has a protective effect on the development of allo-ls does not necessarily contradict these results. like agvhd, allo-ls is a manifestation of alloimmunity. we speculate that the apparent protective effect of agvhd reflects the influence on alloreactive t cells of immunosuppressive agents used to treat agvhd grade ii-iv. none of our patients had lung involvement at the onset of agvhd. this may be because the lungs are less susceptible to acute alloreactivity than the classic target organs of gut, liver, and skin. this is also reflected by the fact that the median time to allo-ls was longer ( weeks) than the median time to agvhd ( weeks). all patients who developed allo-ls did so during tapering of immunosuppressive therapy or after this therapy had been stopped. all of the patients with agvhd where on prolonged immunosuppressive therapy (including steroids) and thus likely had less chance to develop alloimmunity in the lungs. in other words, the rv-positive patients who did not develop allo-ls had significantly greater immune suppression than the rv-positive patients who did develop allo-ls (data not shown). this effect of immunosuppressive therapy on the development of allo-ls is in line with previous findings [ ] . regarding cgvhd, in contrast to other studies, in our cohort, only of patients ( %) with bos had signs of cgvhd in other organs. our finding of no association between cgvhd and bos/boop might be because of the generally low incidence of cgvhd in the pediatric hsct population. this, together with the fact that viral infections are more frequent in childhood, might make the respiratory epithelium a preferential target for chronic alloreactivity, at least in this pediatric cohort. no protective effect of cgvhd on the development of allo-ls was noted, most likely because cgvhd (mainly the limited form) was not treated with systemic immunosuppressive agents (eg, steroids). a similar association between rv infection and allo-ls has been reported after lung transplantation. a recent prospective cohort study in lung transplant recipients clearly showed had significantly more acute or chronic rejection episodes in patients with an rv infection occurring within days posttransplantation [ ] . bos occurring after lung transplantation is considered a manifestation of allograft rejection [ , ] because of an alloimmune process. some animal models of lung transplantation have demonstrated an association between the presence of rv and the development of bos exclusively in the allogeneic transplantation setting [ ] . these results are in line with the association between rv infection and allo-ls seen in our hsct population, and strongly support the hypothesis that airway damage from an rv infection alone does not lead to severe problems, but triggers alloimmunity. because of this strong association, the current practice in lung transplantation is to increase immunosuppression by adding steroids in the presence of an rv infection posttransplantation, to avoid rejection (personal communication lung transplantation programm umc groningen and umc utrecht, ). this practice has led to a decreased graft rejection rate. increasing immunosuppression solely because of the presence of a rv may sound paradoxical, possibly predisposing the patient to other potentially lifethreatening complications, but it is supported by our observation that patients with rv infection early after hsct were less vulnerable to allo-ls when receiving immunosuppression for agvhd. at present, we cannot predict which rv-positive patients are actually at risk for developing allo-ls. early recognition of the disease by the detection of biomarkers associated with lung damage and the development of allo-ls, or the identification of certain risk groups by studying the genetic polymorphisms of innate immunity in these patients, might be of additional value to fine-tune the initiation or adjustment of immunosuppressive therapy in the hsct setting. in conclusion, we have shown a clear relation between early rv infection and the development of allo-ls in pediatric hsct recipients. tissue damage because of the persistence of rv in the lung might be a trigger for the development of allo-ls. agvhd, but more likely greater immunosuppression because of the agvhd, appears to protect for allo-ls. these findings suggest that prevention of rv infections early after hsct is of utmost importance. in addition, prolonged immune suppression in transplant recipients with an rv infection might prevent development of allo-ls, in analogy with current practice in lung transplantation. a. birgitta versluys designed the study, provided clinical data, analyzed data, and wrote the paper. john rossen developed and performed the viral pcr assays. rob schuurman developed and performed the viral pcr assays. bart van ewijk performed pulmonary function tests. marc bierings designed the study and provided clinical data. jaap jan boelens designed the study, provided clinical data, performed statistical analysis, and wrote the paper. financial disclosure: the authors have no conflicts of interest to disclose. chronic lung disease after hematopoietic stem cell transplantation acute lung injury after hematopoietic stem cell transplantation pulmonary complications in adult blood and marrow transplant recipients: autopsy findings pulmonary complications of bone marrow transplantation natural history of pulmonary complications in children after bone marrow transplantation pulmonary complications after bone marrow transplantation in children: twentyfour years of experience in a single pediatric center parainfluenza virus infections after hematopoietic stem cell transplantation: risk factors, response to antiviral therapy, and effect on transplant outcome prospective study of the incidence, clinical features, and outcome of symptomatic upper and lower respiratory tract infections by respiratory viruses in adult recipients of hematopoietic stem cell transplants for hematologic malignancies respiratory virus infections after stem cell transplantation: a prospective study from the infectious diseases working party of the european group for blood and marrow transplantation frequent detection of respiratory viruses in adult recipients of stem cell transplants with the use of real-time polymerase chain reaction, compared with viral culture community respiratory virus infections among hospitalized adult bone marrow transplant recipients rhinovirus infections in myelosuppressed adult blood and marrow transplant recipients respiratory viral infections in adults with hematologic malignancies and human stem cell transplantation 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measurements. official statement of the european respiratory society lung function in children and adolescents: methods and reference values immunomodulatory activity and effectiveness of macrolides in chronic airway disease acute lung injury after allogeneic stem cell transplantation: is the lung a target of acute graft-versus-host disease? incidence and outcome of idiopathic pneumonia syndrome in pediatric stem cell transplant recipients bronchiolitis obliterans following haematopoietic stem cell transplantation bronchiolitis obliterans and other late onset noninfectious pulmonary complications in hematopoietic stem cell transplantation association between acute and chronic graft-versus-host disease and bronchiolitis obliterans organizing pneumonia in recipients of hematopoietic stem cell transplants t-cell depletion prevents from bronchiolitis obliterans and bronchiolitis obliterans with organizing pneumonia after allogeneic hematopoietic stem cell transplantation with related donors incidence, outcome, and risk factors of late-onset noninfectious pulmonary complications after unrelated donor stem cell transplantation risk factors for bronchiolitis obliterans: a systematic review of recent publications infectious etiology of bronchiolitis obliterans. the respiratory viruses connection: myth or reality? respiratory viral infections aggravate airway damage caused by chronic rejection in rat lung allografts key: cord- -cx eh ff authors: donadieu, j; michel, g; merlin, e; bordigoni, p; monteux, b; beaupain, b; leverger, g; laporte, j p; hermine, o; buzyn, a; bertrand, y; casanova, j l; leblanc, t; gluckman, e; fischer, a; stephan, j l title: hematopoietic stem cell transplantation for shwachman-diamond syndrome: experience of the french neutropenia registry date: - - journal: bone marrow transplant doi: . /sj.bmt. sha: doc_id: cord_uid: cx eh ff our objective was to study the outcome of allogeneic hematopoietic stem cell transplantation (hsct) for shwachman-diamond syndrome (sds). among sds patients included in the french severe chronic neutropenia registry, received hsct between and in five institutions. the indications were bone marrow failure in five cases, and myelodysplastic syndrome (mds) or leukemia in five cases. the median follow-up of patients who survived without relapse is . years ( . – . years). the conditioning regimen consisted of a busulfan–cyclophosphamide combination (n= ) or total body irradiation plus chemotherapy (n= ). six patients received stem cells from unrelated donors and four from identical siblings. engraftment was complete in eight patients and unassessable in two patients. these latter two patients died of infections and days after hsct, with grade iv graft-versus-host disease and multiorgan dysfunction. a third patient died from an acute respiratory distress syndrome months after hsct with progressive granulocytic sarcoma. one patient had an mds relapse months after hsct and died months later. the overall -year event-free survival rate is ± %. we conclude that hsct is feasible for patients with sds who develop bone marrow failure or malignant transformation. our objective was to study the outcome of allogeneic hematopoietic stem cell transplantation (hsct) for shwachman-diamond syndrome (sds). among sds patients included in the french severe chronic neutropenia registry, received hsct between and in five institutions. the indications were bone marrow failure in five cases, and myelodysplastic syndrome (mds) or leukemia in five cases. the median follow-up of patients who survived without relapse is . years ( . - . years). the conditioning regimen consisted of a busulfan-cyclophosphamide combination (n ¼ ) or total body irradiation plus chemotherapy (n ¼ ). six patients received stem cells from unrelated donors and four from identical siblings. engraftment was complete in eight patients and unassessable in two patients. these latter two patients died of infections and days after hsct, with grade iv graft-versus-host disease and multiorgan dysfunction. a third patient died from an acute respiratory distress syndrome months after hsct with progressive granulocytic sarcoma. one patient had an mds relapse months after hsct and died months later. the overall -year event-free survival rate is %. we conclude that hsct is feasible for patients with sds who develop bone marrow failure or malignant transformation. shwachman-diamond syndrome (sds) (omin ) is a multisystem autosomal recessive disorder characterized by exocrine pancreatic dysfunction, bony metaphyseal dysostosis, mild intellectual retardation, and variable neutropenia. almost all patients have a mutation in the sbds gene located on chromosome . bone marrow failure and myelodysplasia/acute leukemia are the main life-threatening complications. hematopoietic stem cell transplantation (hsct) is currently the only potentially curative treatment for these latter patients. only hsct procedures have been described in this setting, in separate publications. [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] here we report the outcomes of sds patients included in the french severe chronic neutropenia registry who underwent hsct during the last years. the french severe chronic neutropenia registry was created in , with approval from the french computer watchdog commission (cnil certificate no ). the patients' files are monitored by clinical research associates who visited each center at least once a year. all patients with sds are included in the registry, even if they are not profoundly neutropenic. sds was diagnosed on the basis of neutropenia associated with exocrine pancreatic deficiency and skeletal, skin or liver abnormalities, after pearson's syndrome had been ruled out. the patients or next of kin were required to give their informed consent to participation to the registry. the french scn database was crossed with the french transplant database (etablissement franc¸ais des greffes). a total of patients were included in the sds database, and a complete report of the survey, analyzed at the cut-off date of march , has been published elsewhere. in all, patients developed bone marrow failure, which was transient in five cases and persistent ( months) in six cases. among these latter six patients, the patient who did not receive hsct died. eight patients developed mds/leukemia, of whom three patients who were not transplanted died. sbds gene mutations were evaluated as described elsewhere. acute leukemia was defined by who criteria, that is, at least % of blast cells on bone marrow smears. as dysplastic cytological abnormalities were nearly always present in these patients, myelodysplastic syndrome (mds) was diagnosed if cytological abnormalities in addition to refractory anemia or thrombocytopenia requiring blood transfusion, as well as clonal cytogenetic abnormalities were present. bone marrow failure was diagnosed in case of refractory anemia or thrombocytopenia requiring blood transfusion, if no cytogenetic clonal abnormalities were found. the time of bone marrow failure was recorded as the date of the first transfusion, and the time of mds diagnosis was the date when the first cytogenetic abnormalities were detected. stata s software version was used for all statistical analyses. the end point for survival analyses was relapse (if mds/acute leukemia (al)) or death. the period taken into account was the interval between hsct and either death or mds/al relapse or the last examination when no event occurred. the kaplan-meier method was used to construct survival rates. the cutoff date for this analysis was june , . the median follow-up for the six disease-free surviving patients is . years ( . - . years). description of the patients prior to hsct (table ) the patient group consisted of six males and four females. median age at transplantation was . years ( . - . years). four patients were screened for sbds mutations and all were positive. patients who had bone marrow failure tended to be younger than those who developed mds/al (median age . vs . year; p ¼ . ) and the interval between diagnosis of sds and the hematological event was lower in bone marrow failure than in mds/al (median age . vs . year; p ¼ . ). the median interval between the hematological event and hsct was . years, and was longer in patients with bone marrow failure than in those with mds/al ( . vs . year, p ¼ . ). polychemotherapy with an aml-like regimen, including high-dose cytarabine, mitoxantrone, vp and amsacrine, was administered to one of the five patients with mds/ leukemia; it resulted in partial disease control and disappearance of the cytogenetic clone, but cytological bone marrow abnormalities persisted (about % blasts). the other four patients with mds did not receive chemotherapy prior to hsct. the clinical and hematological characteristics of the patients are shown in table . four patients received marrow from a matched sibling transplant, while six patients received hsct from unrelated donors (three / loci-matched unrelated donors and three single-antigen-mismatch donors). bone marrow was used in all cases and was t cell-depleted in two cases. all the patients received preparatory myeloablation. six patients received a combination of busulfan ( mg/kg in four cases and mg/kg in one case) and cyclophosphamide ( mg/kg), plus antithymocyte globulin in three cases. four patients received total body irradiation ( grays), combined with cyclophosphamide ( mg/kg) in three cases and with melphalan ( mg/m ) in one case. the total number of infused nucleated cells ranged from .  - .  /kg (median  /kg). graft-versus-host disease (gvhd) prophylaxis consisted of methotrexate and cyclosporine a (csa) in five cases, methotrexate, csa and steroids in two cases, csa alone in two cases and steroids alone in one case. chimerism analysis was performed by means of cytogenetics or polymerase chain reaction (pcr) amplification of microsatellites. results (table ) hematological recovery after hsct and engraftment engraftment could not be evaluated in two cases because of early death, while full hematological recovery occurred in the other eight cases. the median time required for the neutrophil count to reach .  /l was days (range - days). the platelet count reached  /l after - days (median . days), without further transfusions. all the patients who are alive and disease-free are currently free of erythrocyte and platelet transfusions. neutrophil counts normalized in every case. complete chimerism was found in all eight assessable patients. three patients developed grade iv acute gvhd (skin/gut in two and skin/liver in one) after receiving a mud transplant in two cases and a matched sibling transplant in one case. two patients with grade iv gvhd died. three patients had grade ii gvhd. two patients developed chronic gvhd, one case with a sicca syndrome and one case with chronic cutaneous gvhd, with persistent livedo. four patients died. the overall -year event-free survival rate was %. two early deaths occurred. one patient (upn ) developed acute grade iv gvhd, thrombocytopenic microangiopathy with multiorgan failure and stenotrophomonas maltophilia sepsis, and died on day of coronavirus e pneumonia. the second patient (upn ) died days after hsct from acute grade iv gvhd, acute respiratory distress syndrome, and hhv infection. the third patient (upn ) developed granulocytic sarcoma of the knee, ischium and shoulder, year after hsct. monosomy was detected in tumor cells by fish analysis. the granulocytic sarcoma was treated with local radiotherapy. at the time of death (due to an undocumented acute respiratory distress syndrome), the marrow morphology was normal and no cytogenetic aberrations were detected. the disease course of this patient was remarkable. on day after bone marrow transplantation (bmt), blast cells were observed on a blood smear, concomitantly with partial chimerism ( % of donor cells). a donor lymphocyte infusion was given on day , resulting in the disappearance of blast cells and in full donor chimerism. a fourth patient (upn ) with mds/al relapsed months after transplantation, and died months later from a hemorrhagic syndrome. these four patients had received hsct for mds/al. although the number of patients is small, survival was significantly poorer in patients who received hsct for malignancy than for bone marrow failure, four of the five deaths involving patients with mds/al and the other death involving a patient with bmf (log rank test, p ¼ . ). one patient had long-term complications, with aseptic osteonecrosis, cardiac hypokinesia, chronic keratitis and amenorrhea. despite the correction of hematological abnormalities, other clinical characteristics of sds such as pancreatic insufficiency, short stature and impaired cognitive performance are still present in all survivors, albeit with no apparent modifications related to hsct. we report the outcome of hsct in patients with sds and severe hematological complications (bone marrow failure in five cases and myelodysplasia/al in five cases). bone marrow aplasia and myelodysplasia/al appear to be the most serious complications of sds, and are always fatal despite conventional management with transfusions and chemotherapy. in sds, al is diagnosed on the basis of cytologic criteria. it can, however, be difficult to distinguish between bone marrow aplasia and myelodysplasia on the sole basis of cytologic bone marrow studies. indeed, cellular dysplasia is present in both instances and the cytologic aspects do not correspond to the fab classification of myelodysplasias. it is therefore crucial to follow cytologic bone marrow studies by cytogenetic examination in order to distinguish between simple bone marrow depletion and clonal progression ('myelodysplasia'). in addition, the presence of a cytogenetic anomaly of iso chromosome , or a deletion of chromosome , as found in one of our patients, has also been observed in patients without transfusion requirements and with nonfatal outcome, , , complicating the decision to proceed with hsct. as shown here, marrow transplantation with both sibling donors and matched unrelated donors can correct the stem cell disorder encountered in sds. full and sustained engraftment was observed in eight patients, arguing against a major stromal defect in sds. despite the small number of patients, we found a significant difference in event-free survival between patients receiving hsct for bone marrow aplasia and those undergoing the procedure for leukemic transformation. such a difference, albeit nonsignificant, was also observed among the published cases of hsct for sds for which this information is mentioned ( deaths or relapses among patients with mds/al; and one death among six patients with bone marrow failure, p ¼ . ). [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] several factors may explain the poor results of hsct for myelodysplasia/al in sds. hsct failed to correct the leukemia in one of our patients and in three of the patients described in the literature. in addition, patients with sds appear to be more susceptible to the toxicity of the hsct procedure than are patients with other myelodysplastic disorders associated with monosomy , like the kostmann syndrome and juvenile myelomonocytic leukemia. this susceptibility may be related to older age or to disease characteristics. indeed, there is a marked age difference between patients receiving hsct for bone marrow aplasia and for leukemic transformation, both in our series and in the literature (median age . and years, respectively). several risk factors could progress with age in this setting, including the nutritional consequences of exocrine pancreatic insufficiency, liver disease, repeated infections and cardiac disorders. a risk of cardiomyopathy has been reported in patients with sds in the absence of hsct, fibrosis and necrotic lesions being found in % of sds patients at necropsy; this was also the case of patient upn , whose sibling died of cardiomyopathy. it is therefore necessary to carefully assess nutritional, cardiac and hepatic status prior to transplantation. all the patients received a myeloablative-conditioning regimen prior to hsct, usually consisting of the busulfancyclophosphamide combination. in patients with bone marrow aplasia, who are generally under years of age, this conditioning appears effective and well tolerated. stability of the engraftment over several years suggests that more intensive conditioning is not warranted. in contrast, the poor results in patients with leukemic transformation suggest that milder conditioning would be inappropriate and that very careful attention must be paid to nutritional status. in conclusion, allogeneic hsct can be envisaged for patients with sds who become transfusion-dependent, with or without clonal cytogenetic features, and for those who develop overt leukemia. our results indicate that bmt is associated with significant morbidity, potentially owing to pre-existing poor nutritional status, immune deficiency and/or the underlying disease. these patients should be closely monitored for infections after hsct, and their nutritional status should be optimized before the procedure. shwachman-diamond syndrome mutations in sbds are associated with shwachman-diamond syndrome allogeneic bone marrow transplatation in a patient with shwachman-diamond syndrome bone marrow transplant in shwachman diamond syndrome related donor liver transplant for veno-occlusive disease following t-depleted unrelated donor bone marrow transplantation successful unrelated bone marrow transplantation for shwachman-diamond syndrome does isochromosome q mandate bone marrow transplant in children with shwachman-diamond syndrome? unrelated donor bone marrow transplantation for children and adolescents with aplastic anaemia or myelodysplasia adult onset of acute myeloid leukaemia (m ) in patients with shwachman-diamond syndrome shwachman-diamond syndrome: early bone marrow transplantation in a high risk patient and new clues to pathogenesis successful allogeneic hematopoietic stem cell transplantation (hsct) for shwachman-diamond syndrome bone marrow transplantation in shwachman-diamond syndrome unrelated donor marrow transplantation for inborn errors successful unrelated donor bone marrow transplantation for shwachman-diamond syndrome with leukemia bone marrow transplantation in shwachman-diamond syndrome: report of two cases and review of the literature allogeneic bone marrow transplantation in shwachman-diamond syndrome with malignant myeloid transformation. a case report liver failure complicating non-alcoholic steatohepatitis following allogeneic bone marrow transplantation for shwachman-diamond syndrome acute leukemia complicating bone marrow hypoplasia in an adult with shwachman's syndrome shwachman-diamond syndrome and matched unrelated donor bmt fatal cyclophosphamide-induced congestive heart failure in a -year-old boy with shwachman-diamond syndrome and severe bone marrow failure treated with allogeneic bone marrow transplantation analysis of risk factors for myelodysplasias, leukemias and death from infection among patients with congenital neutropenia. experience of the french severe chronic neutropenia study group coronavirus e-related pneumonia in immunocompromised patients malignant myeloid transformation with isochromosome q in shwachman-diamond syndrome clonal evolution in marrows of patients with shwachman-diamond syndrome: a prospective -year follow-up study shwachman-diamond syndrome: an inherited preleukemic bone marrow failure disorder with aberrant hematopoietic progenitors and faulty marrow microenvironment hematopoietic stem cell transplantation in severe congenital neutropenia: experience of the french scn register treatment of juvenile chronic myelomonocytic leukemia by allogeneic bone marrow transplantation this work was supported by grant raf from the inserm afm network for rare diseases, socie´te´d'he´matologie et immunologie pe´diatrique, amgen sa, and chugai aventis. we thank florence mesnil (etablissement franc¸ais des greffes) for her participation in patient screening and david young for editorial assistance. key: cord- -hyvu nuq authors: salman, huda; cooke, kenneth r.; lazarus, hillard m. title: fibrosing alveolitis in hematologic malignancy patients undergoing hematopoietic cell transplantation date: - - journal: pulmonary involvement in patients with hematological malignancies doi: . / - - - - _ sha: doc_id: cord_uid: hyvu nuq although advances in antineoplastic therapy have considerably improved the survival of patients with hematological malignancies, current treatment modalities increase the risk of late complications. several forms of chronic pulmonary dysfunction due to infectious or noninfectious causes commonly occur in the months to years after chemo-radiotherapy and can be fatal or result in long-term morbidity. the judicious use of prophylactic antimicrobial agents has tipped the balance toward noninfectious etiologies. hence, while opportunistic infections still contribute to chronic lung disease, late sequelae resulting from antineoplastic therapy have been identified and reported. patients who proceed to receive hematopoietic cell transplantation (hsct) are particularly prone to developing lung complications. pulmonary dysfunction occurring after hsct may manifest with obstructive or restrictive pulmonary mechanics and may range in severity from subtle, subclinical functional changes to frank respiratory failure. insights generated using animal models suggest that the immunologic mechanisms contributing to lung inflammation after hsct may be similar to those responsible for graft-versus host disease. in sum, chronic fibrotic pulmonary dysfunction is a frequent and significant complication facing survivors of hematologic malignancies and their practitioners. the high incidence and suboptimal response to current support care and immunosuppressive therapy underscore the need for heightened awareness and continued research in this area. a -year-old man patient with de novo acute myelogenous leukemia (aml) was induced into complete remission with chemotherapy consisting of idarubicin and cytarabine. after consolidation with high-dose cytarabine, he later received conditioning with cyclophosphamide mg/kg/day intravenously for days and fractionated total body irradiation (tbi) , cgy followed by allogeneic hematopoietic cell transplant (hsct) using a hla-identical sibling donor. graftversus-host disease (gvhd) prophylaxis consisted of tacrolimus and short-course methotrexate. his clinical course was uncomplicated, but after withdrawal of immunosuppression he developed extensive chronic gvhd involving skin and liver. this complication was controlled with the re-institution of tacrolimus. surveillance pulmonary function testing completed days after hsct showed evidence of mild reductions in forced expiratory volume in s (fev ) with preservation of forced vital capacity (fvc). follow-up study revealed significant and rapid worsening of obstructive lung disease (old) despite resolution of hepatic and skin gvhd and continued prophylaxis against viral, fungal and pneumocystis infections using acyclovir, fluconazole and trimethoprimsulfamethoxazole, respectively. reductions in pulmonary function ultimately were associated with shortness of breath and dyspnea with exertion. subsequent workup revealed ground-glass opacities with air trapping on chest computed tomography (ct) scan and evidence of progressive afo on pulmonary function testing based on reduction of fev ( % of predicted normal), a ratio of fev to fvc of . and a residual volume of . l ( % of predicted normal). bronchoalveolar lavage was negative for infection. video-assisted thoracoscopic biopsy of the lungs revealed changes consistent with bronchiolitis obliterans with early fibrosis. the patient continued to receive tacrolimus, and ultimately a course of oral prednisone ( mg/kg/day) and etanercept mg subcutaneous once weekly was initiated. clinical symptoms resolved and pulmonary function improved. he remains in complete remission regarding the aml. fibrosing alveolitis (fa) is a progressive and often fatal disorder characterized by sequential acute lung injury with subsequent scarring and end-stage lung disease. historically, idiopathic pulmonary fibrosis (ipf) encompassed a heterogeneous group of histologic and clinical entities arising in an idiopathic setting [ ] . patients with hematologic malignancies treated with chemotherapy, radiation or hsct, such as the patient described above, commonly develop a wide variety of late and chronic pulmonary dysfunction states [ ] . these complications share many of the clinical and pathologic features described in typical idiopathic fa. this spectrum of pulmonary toxicity observed during fa can be simplified by considering the time of diagnosis in relation to institution of therapy, whether the radiographic abnormalities are focal or diffuse, and by underlying histopathology. in addition, there are individual patient factors that should be considered when formulating a differential diagnosis. these include: radiotherapy delivered to the chest wall or as part • of total body irradiation (tbi) exposure to pulmonary-or cardio-toxic chemother-• apeutic agents current or prior immunosuppressant therapy • history of high-dose chemotherapy exposure prior • to autologous or allogeneic hsct history of opportunistic pulmonary infection (fun-• gal or otherwise) in the case described herein, the patient was exposed to radiation therapy in preparation for hsct and received an allogeneic graft from his hla-matched sibling. while his early posttransplant course was uncomplicated, he developed chronic gvhd of the skin and liver lung after immunosuppression was tapered. the widespread and appropriate use of prophylactic antibiotics has shifted the spectrum of pulmonary dysfunction in hsct recipients from infectious to noninfectious etiologies. this chapter will address the chronic lung complications that lead to pulmonary fibrosis and persistent organ dysfunction in each context with specific focus on hematologic malignancy patients treated using hsct. in patients with hematologic malignancies, severe lung infections frequently lead to the development of acute respiratory distress syndrome (ards). bacterial infections predominate (see table . ) and arise because of severe immune suppression inherent to these disorders and their treatments. the pathology of ards involves severe alveolar epithelial cell damage, hyaline membrane formation, and festinate myofibroblast proliferation and fibrosis in the intra-alveolar spaces. affected hsct recipients who deteriorate and require intubation and mechanical ventilation for ards experience a high mortality. in one series, overall intensive care unit (icu) mortality was % [ ] . in recent years, advancements in supportive care have resulted in significant improvement in survival [ ] . however, longterm survivors continue to have residual lung dysfunction that may progress over time. in one series, autopsy evaluation revealed pulmonary fibrosis in % of such patients, underscoring the importance of dysregulated reparative mechanisms in the lung after an acute insult [ ] . factors influencing progression to the fibro-proliferative phase of ards versus resolution and reconstitution of the normal parenchymal architecture are poorly understood. abnormal repair and remodeling may be profoundly affected by both environmental and genetic factors. in this context, mechanical ventilation may affect the macromolecules that constitute the extracellular matrix (collagen, elastin, fibronectin, laminin, proteoglycan and glycosaminoglycans) and impact the biomechanical balance within the lung parenchyma. fungal infections also may follow a chronic course of prolonged inflammation with focal or diffuse scarring ultimately resulting in significant pulmonary dysfunction. invasive aspergillosis (ia) occurs frequently in hematologic malignancy patients, particularly after an allogeneic hsct, presenting classically as angio-invasive or airway-invasive disease. angioinvasive ia is characterized histologically by invasion and occlusion of small to medium-sized pulmonary arteries by fungal hyphae. this effect leads to the formation of necrotic hemorrhagic nodules or pleuralbased, wedge-shaped hemorrhagic infarcts. the "halo sign" (nodules surrounded by areas of ground-glass attenuation) on chest ct scan strongly suggests a diagnosis of ia [ ] . airway-invasive aspergillosis is characterized by the presence of organisms in the basement membrane of the bronchioles and within the airway lumen. positive yield from respiratory samples such as sputa examination or broncho-alveolar lavage (bal) is more likely in this subtype of ia than in the angio-invasive variety. clinical manifestations of acute airway-invasive aspergillosis include: acute tracheobronchitis, exudative bronchiolitis and bronchopneumonia. using high-resolution ct, the associated bronchiolitis is characterized by the presence of peri-bronchial consolidation, centri-lobular micro-nodules, and branching linear or nodular areas of ground-glass attenuation having a "tree-in-bud" appearance [ ] . this form of airway-invasive aspergillosis can be associated with pseudo-membranous necrotizing tracheal involvement that can cause pneumo-mediastinum and has a high [ ] . airway-invasive aspergillosis can also follow a chronic course known as chronic necrotizing aspergillosis. this condition is characterized by an indolent, granulomatous cavitary infection that may mimic reactivation of tuberculosis radiographically [ ] . mortality is lower compared with the other forms of ia and often is related to the underlying disease of the patient. hematologic malignancy patients treated with chemotherapy or chest wall radiation therapy, or those who proceed to receive a hsct may develop a wide variety inflammatory noninfectious lung disorders that ultimately may lead to pulmonary fibrosis. radiation-induced lung injury first was described in , soon after the development of roentgenograms [ ] . in the distinction between two separate types of radiation-induced lung injury, radiation pneumonitis and radiation fibrosis, was made [ ] . an entire chapter from drs. gallego and rello in this book is dedicated to radiation-related lung injury. radiation-induced lung injury results from the combination of direct cytotoxicity upon normal lung tissue and, perhaps more importantly, the development of fibrosis triggered by radiation-induced cellular signal transduction. the cytotoxic effect is largely a consequence of dna damage and death in normal lung epithelial cells. the development of fibrosis that can compromise lung function is mediated by a number of different cytokines. clinically, the most extensively studied radiation-induced cytokine is transforming growth factor beta (tgf-b), which can induce fibroblast collagen deposition. a normal plasma tgf-b concentration at the conclusion of a clinical course of radiotherapy has been observed to be a predictor for the risk of pneumonitis [ ] . other proinflammatory cytokines, including, but not limited to, interleukin il- , tumor necrosis factor-alpha tnfa and il- , are upregulated immediately after irradiation. increased il- plasma concentrations correlate with an increased risk of radiation-induced lung injury [ , ] . platelet-derived growth factor (pdgf) and basic fibroblast growth factor (bfgf) are upregulated in animal models of lung irradiation injury and antedate the development of fibrosis [ ] . factors affecting the development of radiation-induced lung disease are numerous and are included in table . [ ] [ ] [ ] [ ] ; all have been reported to raise the risk of radiation pneumonitis. radiographic and bronchoscopic findings are nonspecific, and the diffusion capacity for carbon monoxide (dl) typically is depressed in patients with radiation-induced lung damage. long-term glucocorticoids may be effective in the treatment of radiationassociated lung injury in which cop is the leading pulmonary involvement; however, symptoms and radiographic abnormalities, as well as immunologically mediated lymphocytic alveolitis frequently recur with discontinuation of therapy [ , ] . early studies suggested that pentoxifylline may have a role •• method of irradiation such as conformal radiation therapy or specialized techniques including intensity-modulated radiation therapy and stereotactic body radiation therapy [ ] [ ] [ ] [ ] in the treatment of radiation-induced fibrosis involving the skin and subcutaneous tissues as this agent also inhibits experimental bleomycin-induced pulmonary fibrosis in rats, likely via its anti-tnfa effects [ ] . pentoxifylline showed a significant protective effect for both early and late lung radiotoxicity. amifostine is a pro-drug that is de-phosphorylated by alkaline phosphatase in tissues to a pharmacologically active free thiol metabolite. this drug can reduce the toxic effects of chemotherapy by acting as a scavenger of free radicals generated in tissues exposed to radiation. early evidence suggests that amifostine may decrease radiation-induced pulmonary injury without diminishing the therapeutic effect [ , ] . captopril and other ace inhibitors also have been shown to reduce radiation-induced lung fibrosis in rats [ ] , but there are no published data in humans. improvements in the perfusion and ventilation of radiation-injured lung tissue may be expected from to months after radiation therapy. beyond months, however, further significant improvement appears unusual [ , ] . patients with hematologic diseases are exposed to a host of traditional and newer chemotherapeutic agents that can cause lung injury at an incidence that ranges from less than % to as high as % [ , ] . the increased complexity of multi-modality treatments and high-dose protocols designed to augment antineoplastic efficacy, particularly in the context of hsct, has increased the incidence of pulmonary complications. the diagnosis of drug-induced respiratory disease often is complex because: ( ) patients may be exposed to several pneumo-toxic drugs concurrently or in sequence due to earlier treatment failure; ( ) time to onset of pulmonary toxicity may be delayed, making it difficult to ascertain which agent is responsible for the pulmonary reaction; ( ) the combination of drugs to treat malignant hematologic conditions may lead to unexpected drug interactions, producing enhanced toxicity compared with the toxicity of each agent considered separately; and ( ) radiation therapy to the chest or tbi. other factors that play a role in the development of pulmonary toxicity include advanced age, current smoking, abrupt withdrawal of corticosteroids and the use of hsct (allogeneic vs autologous). changes in blood neutrophil counts, thrombocytopenia, coagulation deficits, volume overload or left ventricular dysfunction also can influence the spectrum and severity of pulmonary drug toxicity. in addition to overt pulmonary toxicity, subclinical drug-induced lung dysfunction often occurs in the form of reduced dlco and lung volumes or changes in cell populations in bal fluid. upon cessation of exposure to the agent, most of these changes reverse slowly in a few weeks or months. drug-induced lung injury can manifest in several patterns ( the majority of cases) , azathioprine, chlorambucil, cyclophosphamide, procarbazine and, rarely, vinca alkaloids. the onset of this condition is unpredictable; symptoms may develop a few days to years after exposure. the clinical picture includes increasing dyspnea, dry cough, high fevers and rash. the severity of illness can vary from mild to progressive respiratory failure, and associated radiographic findings may range from bilateral (usually symmetrical) interstitial or alveolar opacities to extensive consolidation with air bronchograms and volume loss [ ] [ ] [ ] . pleural effusions and mediastinal lymph node enlargement have been reported in patients with methotrexate-induced lung injury [ , ] . bal fluid usually shows lymphocyte predominance. a low ratio of cd to cd lymphocytes is suggestive, but not specific, for drug-induced lung disease. other bal findings include neutrophilia or a combined pattern of lymphocytosis with neutrophilia or eosinophilia [ ] . appropriate stains, cultures and molecular techniques in bal fluid should be performed to exclude opportunistic infections. a lung biopsy may be required in selected cases. histopathologic features include interstitial inflammation and pulmonary granulomas. fibrosis can be present, but is generally not the dominant histopathologic feature. alveolar edema or hemorrhage may be found as a manifestation of severe methotrexate pneumonitis [ ] . high-dose corticosteroids may be indicated with more advanced disease, as drug-induced nsip can lead to mortality if it is not treated promptly, but in milder cases, symptoms can subside after simple drug withdrawal [ ] . although rechallenge with the drug may be safe, it is not generally recommended [ ] . eosinophilic pneumonia (ep) is an unusual and unpredictable pattern of response to chemotherapeutic agents as opposed to that described following the use of some antibiotics. ep in patients with hematologic malignancies can result from treatment with fludarabine and, rarely, interferons, inhaled or parenteral pentamidine, and radiographic contrast media [ ] . although methotrexate and procarbazine pneumonitis can often be associated with peripheral eosinophilia, bal and histopathologic features are not those of eps. typically, the syndrome of ep develops during or shortly after termination of treatment. a history of an allergic disorder, or repeated courses of treatment with the specific drug, may predict for a higher risk. the diseases could manifest as acute pneumonia and progress to respiratory failure [ , ] . radiograghic findings of ep include alveolar infiltrates and the classic pattern of "photographic negative" pulmonary edema [ ] . it also could cause faint ground-glass opacities, or kerley's "b" lines (dense and diffuse). ep is diagnosed by the presence of increased percentages or numbers of eosinophils in blood, bal, or lung tissue. a lung biopsy is rarely required, but discontinuance of the offending drug is essential. corticosteroid drug therapy is suggested in cases with severe involvement. the prognosis for this condition usually is good. chemotherapy-induced organizing pneumonia (op) may manifest with chest pain, dyspnea and diffuse radiographic abnormalities with [ ] or without acute respiratory failure [ ] , or may be discovered incidentally on chest imaging [ ] . nodular op typically is seen in patients exposed to chemotherapy who develop round-shaped foci that localize mainly in lung bases, may abut the pleura and simulate metastatic nodules [ ] [ ] [ ] . nonspecific findings are retrieved from bal, such as increases in the percentage of lymphocytes, neutrophils or eosinophils. open lung biopsy guided by the results of ct scan is the procedure of choice. the nodules correspond to sterile aggregates of mononuclear cells. histology reveals interstitial inflammation, superimposed on the dominant background of alveolar and ductal fibrosis. lung nodules with the histopathologic features of cryptogenic organizing pneumonia or of localized fibrosis can be observed after treatment with bleomycin, cyclophosphamide, vinblastine and, rarely, fludarabine [ ] [ ] [ ] [ ] . drug discontinuation and, if required, corticosteroid therapy usually are followed by improvement in most cases. organizing pneumonia (formerly boop) can be seen following hsct and is described in detail later in the chapter. diffuse alveolar damage (dad) is a serious form of pulmonary pathology that may develop in the context of drug-related lung injury. single chemotherapeutic agents (e.g., bcnu or other nitrosoureas, bleomycin, busulfan, chlorambucil, cyclophosphamide, melphalan, procarbazine, vinblastine) or multiagent cytostatic chemotherapy have been reported to cause this lung toxicity [ ] . some regimens may be associated with a greater likelihood of dad than others even if they differ in one agent only. for instance, in patients with de novo-treated hodgkin's lymphoma, the substitution of gemcitabine for dacarbazine, e.g., abvg rather than abvd (doxorubicin, bleomycin, vinblastine and gemcitabine instead of dacarbazine), was associated with a % incidence rate of pulmonary toxicity [ ] . likewise, the substitution of gemcitabine for etoposide in the dose-escalated beacopp regimen (bleomycin, doxorubicin, cyclophosphamide, vincristine, procarbazine, prednisone and gemcitabine rather than etoposide) significantly escalated the likelihood of pulmonary toxicity [ ] [ ] [ ] . concurrent administration of radiation therapy to the chest or use of tbi, supplemental oxygen and possibly colonystimulating factors (csfs) may increase the risk of dad. time to onset of dad can vary from shortly after the first administration of the offending drug to much later into the treatment course [ ] . restrictive lung function patterns and hypoxemia are typical. dlco abnormalities often precede clinical symptoms. the clinical evolution of drug-induced dad varies from an isolated decrease in dlco [ , ] or evidence of fibrosis in trans-bronchial or pulmonary biopsies [ , ] as the only manifestation of toxicity to bilateral, interstitial and alveolar infiltrates [ , ] . severe cases progress to an ards picture and death [ ] . high-resolution ct scanning may show groundglass opacities and intra-lobular septal thickening, and the extent of changes correlates with clinical severity [ ] . dysplastic pneumocytes may be retrieved by bal [ , ] . a lung biopsy is reserved for patients with an atypical presentation or for those who do not improve with empirical antibiotic and corticosteroid treatment [ ] . the main histopathologic feature of dad is consistent with hyaline membranes and fibrin deposits lining the alveolar border, dysplasia of type ii cells, free alveolar fibrin, cells and debris in alveolar spaces and various stages of interstitial edema, inflammation and organization [ ] . dad may be reversible after discontinuation of drugs or after the addition of corticosteroids, or both [ ] . the usual doses of oral corticosteroids may not prevent the condition from developing, but higher doses are reported to reduce the incidence [ ] . the high incidence, severity and unpredictability of dad associated with chemotherapy suggest that it is reasonable to discontinue such treatment once the dlco has decreased % compared with pre-therapy values. although smaller decrements in dlco do not equate to toxicity and should not lead to withdrawal of chemotherapy, a precipitous decrease in the dlco indicates impending toxicity [ ] . when radiation therapy is planned after the administration of chemotherapeutic agents, it is advisable to wait for any chemotherapy-induced decrease in the dlco to stabilize or show a trend toward improvement before starting radiation. finally, drug-induced pulmonary fibrosis may develop in patients receiving cytotoxic agents, such as bleomycin, busulfan, bcnu, lomustine, ccnu chlorambucil, cyclophosphamide, melphalan, vinca alkaloids, radiation therapy and tbi [ ] . this entity more often is diagnosed months or years after termination of treatment. early signs of this disease are basilar or diffuse streaky opacities and volume loss. this condition can progress to honeycombing and fibrotic changes; reversal of this toxicity and the response to corticosteroids are unpredictable and often unsatisfying. histologic exam can demonstrate the characteristic dysplasia of type ii pneumocytes that reflects exposure to alkylating agents and radiation therapy. in a few patients, especially children treated for hematologic malignancies, pleural or pulmonary fibrosis may develop [ ] . this process results in thoracic deformity, encasement of the lungs and severely restricts lung physiology. an accelerated variant of pulmonary fibrosis, acute interstitial pneumonia (formerly termed the hamman and rich syndrome), has been described after treatment with chlorambucil and methotrexate [ ] [ ] [ ] . the prognosis of this condition is poor despite drug withdrawal and institution of high-dose corticosteroids. as seen in the patient description at the start of this chapter, a decline in lung function long has been identified as a significant complication in the months to years that follow allogeneic hsct. a clinical pearl from dr. bergeron in this book also very nicely describes this type of pulmonary involvement. noninfectious conditions now represent the major pulmonary causes of morbidity and mortality after hsct. idiopathic pneumonia syndrome (ips), discussed in another chapter in this book, remains one of the more common and serious pulmonary complications occurring within months after hsct. although graft-versus-host reactions may play an etiologic role, the major contributing factor is conditioning-related toxicity. among lung conditions that are more closely associated with gvhd, both bronchiolitis obliterans (brob) (onset months to years after hsct) and bronchiolitis obliterans with organizing pneumonia (cop) may lead to fa. the term cop should not be used interchangeably with bronchiolitis obliterans (bo) to describe a patient with chronic lung dysfunction after hsct, although such usage unfortunately is widespread. the two disorders differ with respect to histopathology, pulmonary function characteristics and, most importantly, response to therapy. brob is an inexorably progressive condition, whereas cop behaves similarly to idiopathic cop seen in other populations. cop after hsct usually is quite responsive to corticosteroids and in other settings may resolve spontaneously, whereas brob is not [ , ] . organizing pneumonia also is associated with restrictive (rather than obstructive) changes on pulmonary function testing (table . ). in allogeneic hsct recipients, the disparity in match between the donor graft and the recipient for the human leukocyte antigens (hlas) mediate both gvhd and graft rejection (host-versus-graft reaction). the presence of alloreactive injury to the lung attributed to gvhd is poorly defined and remains debated. in the skin, liver and intestine, gvhd produces a characteristic t-lymphocyte-mediated epithelial destruction. there are few data to support such a defined lesion with the exception of lymphocytic pneumonitis [ ] . a variety of pulmonary complications have been described as manifestations of gvhd, but these associations are based primarily on the simultaneous occurrence of pulmonary abnormalities, the absence of an infectious agent and nonspecific histopathologic lesions in the setting of established gvhd in other organs. nevertheless, both acute and late-onset lung injury syndromes have shown a clinical association with gvhd, including ips, engraftment syndrome, diffuses alveolar hemorrhage, brob and cop [ ] . several murine models also demonstrate pathologic lung changes in the setting of gvhd, thus supporting a mechanistic relationship between gvhd and lung injury. old and chronic afo are the most common noninfectious late pulmonary complications of allogeneic hsct. these entities are manifested on pulmonary function testing by a diminished fev or fev /fvc. the incidence of these syndromes ranges from % to %, depending upon the definition of afo applied in each study [ , ] . typically, the presentation occurs beyond the third month after hsct [ ] . among patients who develop chronic gvhd, new-onset afo may develop in up to one third of the patients. in a study of cases the underlying process accounting for afo was brob in % [ ] . histologically, this process demonstrates fibrous obliteration of the lumen of respiratory and membranous bronchioles. in the absence of histopathologic evidence, new onset afo after allogeneic hsct often is referred to as "bronchiolitis obliterans syndrome" (bos). in addition to chronic gvhd, risk factors for the development of afo include increasing recipient age, pre-transplant reduction in the ratio fev /fvc, low serum immunoglobulin levels, use of methotrexate and a history of respiratory viral infection within the first days [ ] . the onset typically is insidious with presenting symptoms including dry cough ( - %), dyspnea ( - %) and wheezing ( %), but fever is uncommon [ , , ] . the chest radiograph is usually normal, but high-resolution ct scans often demonstrate evidence of expiratory air trapping, hypo-attenuation and bronchial dilation [ , [ ] [ ] [ ] . demonstrating persistent afo using pulmonary function testing and exclusion of other causes of afo such as asthma, tobacco-related emphysema, and viral or bacterial respiratory infection establish the diagnosis. except for its utility in excluding an infectious etiology, bal is usually nonspecific [ ] , and transbronchial biopsies typically are nondiagnostic due to the patchy nature of this small airway process and the limited size of samples obtained. surgical lung biopsy is rarely indicated. the etiology of new onset afo after hsct is unknown. those recognized causes in otherwise normal hosts rarely include recurrent aspiration, viral infection (influenza, adenovirus, measles) and bacterial infection (mycoplasma sp.) [ ] . immunologic mechanisms inducing bronchial epithelial injury are suggested by the strong association between chronic gvhd and new onset afo. indeed, the lung epithelium may be the target of immune-mediated injury induced by donor cytotoxic t cells in chronic gvhd [ ] . thus, brob after hsct may represent a manifestation of gvhd in the lung. disease progression is variable; however, the syndrome is associated with significantly increased mortality rates, and improvement in lung function is uncommon. many patients develop a progressive decline in lung function resulting in respiratory failure [ , ] . there are no prospective studies of the treatment of new onset afo after hsct. old in the presence of chronic gvhd is managed primarily by controlling gvhd. various immunosuppressive agents have been reported to result in stabilization of lung function in - %, but improvement in only - % [ , ] . in the hope that early recognition and treatment may improve outcome, routine spirometry among patients with chronic gvhd is encouraged to detect the insidious onset of this process. restrictive lung disease (rld) is defined by reductions in fvc, total lung capacity (tlc) and dlco as measured by standard pulmonary function tests (pfts). in rld, the ratio fev /fvc is maintained near % [ , ] . rld is common after hsct. significant decreases in fvc or tlc have been reported in as many as - % of allogeneic hsct recipients by day . a decline in tlc or fvc after hsct (even if the absolute values for each measurement remained within the normal range) has been associated with an increase in nonrelapse mortality. tbi-containing conditioning regimens and the presence of acute gvhd have been associated with rld, in addition to obstructive lung disease [ ] [ ] [ ] ; however, the impact of age on the development of rld is less clear. early reports suggested that the incidence of rld is lower in children compared to adults and that the incidence increases with advancing recipient age [ ] . more recent studies have revealed significant rld in children receiving hsct [ ] . organizing pneumonia after hsct falls under the rld pattern on liver function tests and recently was shown to be associated with prior acute and chronic gvhd. organizing pneumonia has been described in case reports as occurring after both allogeneic and syngeneic hsct; these data suggest an association of the lung lesion with chronic gvhd and intestinal ulcerations. in addition, corticosteroid therapy appeared beneficial in the resolution of the lesion. in a recent case control study, freudenberger et al. reviewed cases of histologic cop [ ] . the clinical features of cop in this population were similar to idiopathic and other etiologies with an association between acute and chronic gvhd and the subsequent development of cop. affected patients were more likely to have skin involvement with acute gvhd and chronic gvhd affecting the gut and oral mucosa. the causes of cop following hsct remain enigmatic, but possible etiologies include direct allo-immunologic reactions, atypical infection or atypical manifestations of ips. regardless, the clinical presentations and responses of cop are similar to other cases of idiopathic cop. the published literature contains a paucity of therapeutic trials for chronic lung injury after hsct. a study by payne and colleagues showed that the use of cyclosporine and methotrexate as gvhd prophylaxis prevented the development of old when compared to historic controls receiving prednisone and methotrexate [ ] , but results of prospective, randomized trials in this setting are not available. three recently published case series have exploited the antiinflammatory effects of azithromycin to treat old in both allogeneic hsct and lung allograft recipients. each study suggested a beneficial effect of this drug on pulmonary function when administered for or more weeks [ ] [ ] [ ] . the potential role for tnfa in the pathogenesis of both old and rld suggests that agents such as etanercept may have promise, and several studies have demonstrated a potential benefit of this drug in some hsct patients with chronic lung injury [ , ] . the immunologic mechanisms responsible for chronic, fibrotic pulmonary dysfunction after hsct remain poorly defined, in large part because of the lack of correlative data obtained from afflicted hsct recipients and the paucity of suitable sct animal models for either rld or old. chronic pulmonary disease following allogeneic hsct likely involves an initial insult to lung parenchyma followed by an ongoing inflammatory process involving the interplay between recruited donor-derived immune cells and the resident cells of the pulmonary vascular endothelium and interstitium. mechanistic insights into old following hsct have been derived from studies of lung allograft rejection. data generated from both humans and mice support the hypothesis that the development of brob in this scenario involves the secretion of inflammatory cytokines and chemokines, along with interactions between apcs and activated lymphocytes [ , ] a tri-phasic model of chronic noninfectious lung injury after hsct has been proposed [ ] . in phase i, an acute pneumonitis develops as a consequence of an allogeneic immune response, resulting in the sequential influx of lymphocytes, macrophages and neutrophils into an inflamed pulmonary parenchyma. in phase ii, a persistent inflammatory signal, in the setting of dysregulated repair mechanisms, promotes the transition from acute to chronic injury. if the inciting injurious stimulus predominantly involves bronchiolar epithelial cells, phase ii is associated with the concentric infiltration of lymphocytes and collagen deposition in the peri-bronchiolar areas resulting in the development of chronic bronchiolitis. if, however, the alveolar epithelium is the primary target, leukocyte recruitment and matrix deposition are confined primarily to the interstitial space. as chronic inflammation proceeds to phase iii, lung fibroblasts increase dramatically in number and contribute to the enhanced deposition of collagen and granulation tissue in and around bronchial structures, ultimately resulting in complete obliteration of small airways and fixed old. by contrast, fibroblast proliferation and intra-septal collagen deposition during phase iii ultimately result in interstitial thickening, septal fibrosis, significant volume loss and severe rld. clinical and experimental data suggest that the progression to a chronic, pro-fibrotic form of pulmonary toxicity involves the secretion of cytokines and chemokines [ ] [ ] [ ] , and in this context, tnfa may be a central factor in the proposed tri-phasic model of disease. evidence for a role of tnfa in the transition from acute to chronic lung injury comes from studies using targeted over-expression of tnfa in the lungs of rodents [ ] . in these models, early lung histopathology includes a lymphocytic infiltrate similar to that seen in experimental ips models [ , ] , whereas the histologic changes associated with more prolonged exposure to tnfa show both interstitial and peribronchial inflammation that closely resemble changes seen at later time points after hsct [ , ] . fa is characterized by sequential acute lung injury that can culminate in scarring and end-stage lung disease. despite the high success rate in treating hematologic malignancies with or without using hsct, this sequence of events continues to be a significant contributor to nonrelapse morbidity and mortality in patients with hematologic malignancies because of either the disease itself or as a result of treatment modalities employed. the pathophysiologic mechanisms contributing to the initiation and progression of disease remain poorly defined. to this end, current treatment options remain suboptimal and primarily limited to supportive care measures and antiinflammatory agents, such as corticosteroids or other immunosuppressant therapy. 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disease in bone marrow transplant recipients key: cord- -nkaow h authors: sim, starling a.; leung, vivian k.y.; ritchie, david; slavin, monica a.; sullivan, sheena g.; teh, benjamin w. title: viral respiratory tract infections in allogeneic hematopoietic stem cell transplantation recipients in the era of molecular testing date: - - journal: biol blood marrow transplant doi: . /j.bbmt. . . sha: doc_id: cord_uid: nkaow h viral respiratory tract infection (vrti) is a significant cause of morbidity and mortality in patients undergoing allogeneic hematopoietic stem cell transplantation (allo-hsct). this study aimed to assess the epidemiologic characteristics, risk factors, and outcomes of vrti occurring in the period from conditioning to days after allo-hsct in the era of molecular testing. this study was a retrospective record review of patients who underwent allo-hsct at royal melbourne hospital between january and december . symptomatic patients were tested using respiratory multiplex polymerase chain reaction (pcr). logistic regression and kaplan-meier analysis were used to identify risk factors for vrti and the risk of death or intensive care unit (icu) admission, respectively. a total of patients were reviewed, and episodes of vrti were identified in patients ( . %). rhinovirus accounted for the majority of infections ( . %). the majority of episodes presented initially with upper respiratory tract infection ( . %), with . % of them progressing to lower respiratory tract infection. eleven episodes ( . %) were associated with icu admission. there were no deaths directly due to vrti. previous autologous hsct was associated with an increased risk of vrti (odds ratio, . ; % confidence interval, . to . ). the risks of death (p = . ) or icu admission (p = . ) were not significantly different by vrti status. vrti is common in the first days after allo-hsct and is associated with icu admission. respiratory viruses (rvs), including influenza virus, parainfluenza virus, respiratory syncytial virus (rsv), rhinovirus, coronavirus, adenovirus, and human metapneumovirus (hmpv), are common causes of viral respiratory tract infection (vrti) [ ] [ ] [ ] . previous studies have shown that rvs cause significant morbidity and mortality in patients undergoing hematopoietic stem cell transplantation (hsct), in particular allogeneic hsct (allo-hsct) [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] . the diagnosis of rvs, especially in older studies, has been dependent mainly on nonmolecular diagnostic methods, including direct antigen detection or cell culture, which are limited by poor sensitivity [ ] . molecular testing using polymerase chain reaction (pcr) allows rapid detection of rvs with high sensitivity and specificity. moreover, multiplex assays enable the detection of multiple rvs in a single test, and these are now the current standard of care in many clinical settings. the purpose of this study was to assess the epidemiologic characteristics, risk factors, and outcomes of vrti in allo-hsct recipients in the era of molecular testing. melbourne hospital (rmh) between january and december . only the first allo-hsct was considered in patients who underwent multiple transplantations during the study period. patient records for the first days following allo-hsct were reviewed. demographic and clinical data were collected from hospital clinical records using a case report form and included age, sex, underlying disease, previous therapy, stem cell source, conditioning therapy, graft-versus-host disease (gvhd), and outcomes (ie intensive care unit [icu] admission and death). baseline measurements of lymphocyte and neutrophil levels and cytomegalovirus (cmv) seropositivity were collected. data on respiratory viral pcr testing and results from symptomatic patients were extracted from the rmh pathology database. for patients with rv infection, the number of vrtis, type of rvs, clinical presentation, antiviral therapy, and outcomes (ie, icu admission, death, use of mechanical ventilation, and progression to lower respiratory tract infection [lrti]) were also obtained during the -day period. during the review period, treatment of rsv with i.v. ribavirin was recommended in the setting of radiologic changes in patients within days of allo-hsct. all specimens, including nasal and throat swabs, nasopharyngeal aspirates, and sputum or bronchoalveolar lavage specimens, were examined by real-time pcr at the melbourne health shared pathology service according to previously published protocols [ , ] . rvs included in the multiplex pcr panel were rsv, influenza type a and b, and rhinovirus. the testing of copathogens was not part of routine assessment and was performed at the discretion of treating physician. these tests included conventional culture for bacteria and fungi; pcr for legionella, pneumocystis jirovecii, and aspergillus [ ] ; viral pcr for herpes simplex virus (hsv)- , hsv- , cmv, varicella zoster virus, human herpesvirus (hhv)- , hhv- , hmpv, and adenovirus; and bacterial pcr for mycoplasma pneumoniae, chlamydophila pneumoniae, and chlamydia psittaci. baseline measurements of lymphocytes and neutrophils were defined as measurements obtained on the day of admission ± days. lymphopenia was defined as an absolute lymphocyte count below . × /l, and neutropenia was defined as absolute neutrophil count below . × /l. conditioning intensity was defined in accordance with center for international blood and marrow transplant research guidelines [ ] . upper respiratory tract infection (urti) was defined as an rv detected in an upper respiratory tract fluid specimen together with symptoms and/or signs, with the exclusion of other possible causes. lrti was defined as detection of rv in respiratory secretions, preferentially in samples obtained from sites of involvement together with pathological sputum production, hypoxia, or pulmonary infiltrates [ ] . progression to lrti was defined as the onset of lrti in patients with a previous urti. an episode of infection was defined as evidence of rv, urti, or lrti detected during the conditioning period and for up to days following allo-hsct. an infection was considered subsequent if the rv detected was nonidentical or if it was identified at least days following the previous episode with the identical rv [ ] . the presence of a copathogen was defined as a bacterial, fungal, or nonrespiratory virus pathogen isolated from a respiratory tract sample during an episode of infection. overall mortality was defined as death due to any cause within days of vrti diagnosis or allo-hsct. death attributable to vrti was defined as death resulting from respiratory failure with other causes excluded. the risk of vrti was calculated as the number of patients testing positive for any rv among all allo-hsct recipients. patients' baseline characteristics were compared by vrti status using the chi-square test or fisher exact test for categorical variables and the student t test or wilcoxon rank-sum test for continuous variables. only the first episode of vrti was considered for risk factor analysis. univariable and multivariable analyses of risk factors for vrti were performed using logistic regression models. variables with a p value <. were included in the multivariable testing, along with variables previously identified as important risk factors for vrti, including lymphopenia, gvhd, and donor relation [ , , ] . patients with vrti had their records reviewed for days following infection. in patients without vrti, the review was censored to the date of death/icu admission or days following transplantation, whichever was sooner. survival (to death or icu admission) within days following transplantation was assessed using kaplan-meier plots, stratified by vrti status. the log-rank test was used to assess differences in survival among groups. all statistical analyses were performed using r version . . , with statistical significance defined at α = . . approval to conduct the study was granted by the melbourne health human research ethics committee (reference qa ). a total of patients were identified during the review period (january to december ). medical records could not be obtained for patients, including patient with vrti ( figure ). the patients reviewed included slightly more males (n = ; . %) than females (n = ; . %), and the median patient age was years (interquartile range [iqr], to years). the median age of patients with vrti was significantly lower than patients without (p = . ). other patient demographic data and characteristics were not significantly different by vrti status ( table ) . out of patients, were tested for vrti ( . %) within days following allo-hsct and episodes of vrti were identified in patients ( . %). eight patients had multiple episodes, with episodes and with episodes. the median time to onset of the first infection from the date of allo-hsct was days (iqr, . to . days). among the patients identified with vrti, their first episode was identified during the first days following transplantation in ( . %). the clinical characteristics of vrti episodes are summarized in table . rhinovirus accounted for the majority of infection episodes ( of ; . %), followed by rsv ( of ; . %), and there was episode of coinfection with these pathogens. most of the cases initially presented with urti ( of ; . %), and this was most pronounced for rsv ( of ; . %). progression to lrti was observed in . % ( of ) of all patients with an initial presentation of urti and was most common for rhinovirus ( of ; . %). the most common symptom observed was cough (n = ; . %), followed by fever ( ; . %), coryza ( ; . %), and sputum production ( ; . %). the rates of cough ( % versus . %), sputum production ( . % versus %), and fever ( . % versus . %) were significantly higher in the patients with lrti compared with those with urti (all p < . ). antiviral therapy was administered in cases ( . %). among the patients with rsv, ( . %; urti and lrti) received ribavirin, and ( . %; urti) received either placebo or gs- as part of a clinical trial. all patients who had either influenza a or b received oseltamivir, and patient with an lrti with rhinovirus infection (n = / , . %) received i.v. immunoglobulin. antiviral therapy was initiated within a median of . days (iqr, . to . days) after the detection of rv. the median duration of antiviral therapy was . days (iqr, . to . days). sixteen copathogens were identified from vrti episodes ( . %). one episode had copathogens ( bacterial and fungal), and episodes had copathogens ( episodes with fungal pathogens and episode with a bacterial and a fungal pathogen). bacterial copathogens were found in out of episodes ( . %) and accounted for one-half the copathogens detected ( of ; . %). these included staphylococcus spp. (n = ), enterococcus faecium (n = ), haemophilus influenzae (n = ), chlamydophila pneumoniae (n = ), and pseudomonas aeruginosa (n = ). of note, fungal and nonrespiratory virus copathogens were only found concomitantly with rhinovirus infection. fungal copathogens identified include aspergillus (n = ), candida albicans (n = ), cladosporium sp. (n = ), saccharomyces cervisiae (n = ), and p. jirovecii (n = ). no episodes met the european organization for research and treatment of cancer/mycoses study group criteria for invasive fungal disease. the nonrespiratory viruses isolated were cmv and hhv- . in univariate and multivariable analyses, only previous autologous hsct was associated with increased risk of vrti (odds ratio, . ; % confidence interval, . to . ). other variables, including age, sex, underlying disease, cmv seropositivity, donor relation, stem cell source, conditioning regimen and intensity, acute gvhd, lymphopenia, and neutropenia were not statistically significant (table ). in addition, no variables were significantly associated with the risk for vrti for fixed periods following hsct (< days, to days, or to days following transplantation). there were a total of episodes of vrti in our study cohort. during the study period, episodes ( . %) were associated with icu admission within days of vrti, with episodes ( . %) necessitating the use of mechanical ventilation. of the episodes, episodes of vrti ( rhinovirus, rsv, and influenza a) had lrti as the initial presentation and respiratory symptoms that precipitated the icu admission. patients were admitted to icu within a median of . days (iqr, to . days) after vrti and for a median duration of days (iqr, . to . days). the risk of icu admission within days following transplantation between patients with vrti ( of ; . %) and those without vrti ( of ; . %) was not significantly different (p = . ) ( figure ). five patients died within days of vrti, all of whom had rhinovirus infection and prior icu admission. three patients were admitted to the icu day before the detection of rhinovirus, whereas the other patients had icu admission on day and day following the detection of rhinovirus. of the patients who died, ( . %) had lrti as the initial site of infection and ( . %) had copathogens detected. however, none of these deaths was directly attributable to vrti. causes of death included multiorgan failure (n = ), pneumonitis (n = ), acute pulmonary edema (n = ), and aspiration pneumonia, gvhd, and fluid overload (n = ). the risk of mortality within days following transplantation between patients with vrti ( of ; . %) and those without vrti ( of ; . %) was not significantly different (p = . ) ( figure ). all vrti patients who died within days following transplantation developed vrti during the first days following transplantation. in this study, vrti was prevalent among allo-hsct recipients during the first days following transplantation. the frequency of vrti among allo-hsct recipients was . %, which is consistent with the rates of . % to % reported elsewhere [ , , , ] . however, differences between our study and previous studies include a shorter duration of review ( days), inclusion of symptomatic patients only, and the range of pathogens tested [ , , , , ]. outcomes (icu admission or death) days following transplant. rhinovirus accounted for the majority of infections, as has been reported previously, albeit among symptomatic and asymptomatic patients [ , ] . copathogens were identified in % of the episodes and were most common in patients with rhinovirus infection. ison et al. [ ] hypothesized that rhinovirus might predispose patients to additional infection. respiratory colonization with potentially pathogenic bacteria also may increase the risk of subsequent vrti [ ] . however, we note that in both that study and our present study, the number of patients was small, and further investigation of a potential link is warranted. in our cohort, patients who had undergone previous auto-hsct were at increased risk of developing vrti independent of gvhd, donor relation, and lymphopenia. auto-hsct has been identified as an important risk factor for progression to lrti [ ] . previous auto-hsct may reflect advanced disease status and thus the susceptibility of patients to vrti. in addition, cumulative immune suppression from multiple lines of previous therapy may increase the risk of vrti, as has been seen in patients with multiple myeloma [ ] . delays in immune recovery following auto-hsct also could explain the increased risk of vrti in this patient population [ , ] . a key findings of the present study was the low number of deaths in patients with vrti, with no mortality directly attributable to vrti. this finding was consistent with rates reported in previous studies ( % to . %) [ , , , ] . we were unable to directly assess factors contributing to reduced mortality, but postulate that it may be attributable to high standards of supportive care. in addition, antiviral therapy was initiated in some rsv cases and in all episodes of influenza type a and b infection early in the course of infection. among the patients with vrti, all deaths occurred in patients who acquired the infection within days of transplantation. this high risk and poor prognosis subgroup supports the need for strict infection prevention measures for staff and patients in the early days after transplantation [ , [ ] [ ] [ ] [ ] [ ] , along with careful assessment of patients presenting for allo-hsct with respiratory symptoms. our study demonstrates that vrti is an important cause of morbidity in allo-hsct recipients. up to % of vrti episodes necessitated icu admission and % required the use of mechanical ventilation, percentages consistent with previous reports [ ] [ ] [ ] . overall, close to % of patients had lrti on initial presentation. progression to lrti occurred in % of infections, most commonly involving rhinovirus. progression to lrti was also observed in % of rsv urti episodes, a higher rate than reported previously [ ] . both the innate and adaptive arms of the immune system, particularly neutrophils and cytotoxic t cells, play integral roles in the defense against vrti [ ] . th -dominant responses, * includes episode in which both rsv and rhinovirus were detected concurrently. † percentages do not sum to because episodes of rhinovirus infection were identified as neither urti nor lrti, because there was insufficient information available for classification. ‡ nonattributable death was calculated using the number of patients. mediated by ifn-γ, appear to have a protective role in rsv infections, whereas th responses may be associated with more severe disease manifestations [ ] . the profound immune depletion and impaired immune responses seen in the acute period following transplantation could account for the rates of initial lrti and progression seen even with respiratory viruses considered less pathogenic, such as rhinoviruses. in addition, our cohort had a lower rate of antiviral therapy for rsv. because we did not evaluate pulmonary function test results, the risk for vrti and later complications, such as airway disease, as shown previously by erard et al. [ ] , could not be assessed. a limitation of this study was the number of rvs included in the multiplex pcr panel. other rvs, such as parainfluenza virus, adenovirus, coronavirus, and hmpv, were not routinely tested but might be important causes of morbidity in these patients. as such, rhinovirus as a cause of vrti and its burden in the study may be overestimated. furthermore, because in this study we examined only prevalence, risk factors, and outcomes of vrti within days following transplantation our results might be not translatable beyond this time frame. however, because this is a known high-risk period [ , ] , our study focused on this time frame to highlight the importance of early preventive measures and infection control measures among patients as well as healthcare workers following transplantation. in conclusion, vrti was prevalent among our allo-hsct recipients. although its impact on mortality appears limited, it led to a number of icu admissions and necessitated the use of mechanical ventilation in several cases. our findings indicate that an association between previous auto-hsct and an increased risk of vrti. outcomes were poorest for patients diagnosed within days following transplantation, a group of patients at high risk and with a poor prognosis for whom early preventive and infection control measures should be targeted. the authors thank ms. jenny collins for her assistance with the transplantation-related data and the staff of melbourne health shared pathology service for the pathology information. aspergillus pcr testing was performed by the department of microbiology at westmead hospital. financial disclosure: the world health organization collaborating centre for reference and research on influenza is supported by the australian government department of conflict of interest statement: there are no conflicts of interest to report. respiratory viral infections in adults with hematologic malignancies and human stem cell transplantation 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and outcome of h n influenza infection in allo-sct patients: a british society of blood and marrow transplantation study outcome of influenza infections in outpatients after allogeneic hematopoietic stem cell transplantation infections with the h n influenza virus among hematopoietic sct recipients: a single center experience respiratory syncytial virus infection in recipients of allogeneic stem-cell transplantation: a retrospective study of the incidence, clinical features, and outcome the human immune response to respiratory syncytial virus infection airflow decline after myeloablative allogeneic hematopoietic cell transplantation: the role of community respiratory viruses rhinovirus infections in myelosuppressed adult blood and marrow transplant recipients key: cord- - bnv t f authors: piñana, josé luis; pérez, ariadna; montoro, juan; giménez, estela; gómez, maría dolores; lorenzo, ignacio; madrid, silvia; gonzález, eva maría; vinuesa, víctor; hernández-boluda, juan carlos; salavert, miguel; sanz, guillermo; solano, carlos; sanz, jaime; navarro, david title: clinical effectiveness of influenza vaccination after allogeneic hematopoietic stem cell transplantation: a cross-sectional, prospective, observational study date: - - journal: clin infect dis doi: . /cid/ciy sha: doc_id: cord_uid: bnv t f background: vaccination is the primary method for preventing influenza respiratory virus infection (rvi). although the influenza vaccine is able to achieve serological responses in some allogeneic hematopoietic stem cell transplantation (allo-hsct) recipients, its clinical benefits are still uncertain. methods: in this prospective, cross-sectional study, we retrospectively analyzed the effect of inactivated trivalent influenza vaccination on the prevalence of influenza rvi in a consecutive cohort of allo-hsct adult recipients who developed rvi over flu seasons (from to ). respiratory viruses in upper– and/or lower–respiratory tract specimens were tested using multiplex polymerase chain reaction panel assays. results: overall, we diagnosed episodes ( %) of influenza rvi in allo-hsct recipients. influenza rvi occurred in % of the non-vaccinated compared to % of the vaccinated recipients (p = . ). a multivariate analysis showed that influenza vaccination was associated with a lower prevalence of influenza rvi (odds ratio [or] . , p = . ). a multivariate risk factor analysis of lower–respiratory tract disease (lrtd) identified conditions associated with the probability of influenza rvi progression: influenza vaccination (or . , % confidence interval [ci] . – , p = . ) and a high-risk immunodeficiency score (or , % ci . – , p = . ). influenza vaccination was also associated with a lower likelihood of an influenza-related hospital admission ( % vs %, p = . ). conclusions: this study shows that influenza vaccination may have a clinical benefit in allo-hsct recipients with virologically-confirmed rvi, in terms of a lower influenza rvi prevalence, slower lrtd progression, and lower likelihood of hospital admission. the influenza virus has a significant impact on morbidity and mortality in allogeneic hematopoietic stem cell transplantation patients (allo-hsct), leading to complications ranging from self-limited upper-respiratory tract infections to life-threatening or fatal pneumonias [ ] [ ] [ ] [ ] . the particular threat influenza poses for allo-hsct recipients was well documented during the influenza a/h n pandemic, as well as during consecutive seasonal influenza epidemics. it showed increased risks of subsequent bacterial pneumonia, hospital admissions, mechanical ventilation requirements, and mortality [ , ] . vaccination is the primary method for preventing influenza infections, but it is less effective in immunocompromised patients than in healthy individuals [ ] [ ] [ ] [ ] . based on studies analyzing serological responses, the inactivated influenza vaccine is strongly recommended annually for patients beyond months post-transplant [ ] [ ] [ ] [ ] ; beyond months after allo-hsct if without graft-versus-host disease (gvhd) or immunosuppression [ ] ; or during community outbreaks [ , ] . however, allo-hsct recipients respond poorly to vaccinations: only onethird of allo-hsct recipients will achieve protective influenza antibody titers [ , , ] , which obviously limits the efficacy of such a vaccine, challenging reports on its clinical benefits. in fact, clinical evidence of influenza-inactivated vaccine effectiveness after allo-hsct is very limited [ ] . strategies based on an influenza vaccine with a higher antigen dose [ ] , multiple vaccine doses [ ] , or adjuvanted vaccines [ ] might improve immune responses, although no controlled trials have been performed. we conducted a prospective, cross-sectional, observational epidemiological study of community-acquired respiratory virus (carv) respiratory tract disease (rtd) in allo-hsct recipients who developed upper rtd (urtd) and/or lower rtd (lrtd) symptoms after transplant. patients' influenza vaccination status was prospectively recorded at the time of their respiratory virus screening. in this study, we report the prevalence of influenza rtd according to the vaccination status over consecutive influenza seasons in a consecutive series of allo-hsct recipients with virologically-documented respiratory virus infections (rvis). this was a prospective, cross-sectional study of rvis in adult (> years) allo-hsct recipients that was conducted at spanish transplant centers. the whole cohort comprised consecutive allo-hsct recipients with respiratory symptoms who were prospectively screened for respiratory viruses at the hospital clinic universitari of valencia (hcuv) between december and may and at the hospital universitari i politècnic la fe in valencia (hlf) from june to may . in december at the hcuv and in may at the hlf, we implemented the medical information/education for recipients and caregivers, explaining in detail the risks of having rvis in the context of immunosuppression. the specific information provided included a description of the respiratory symptoms that merit urgent notification to the transplant team; recommendations concerning screening for respiratory viruses; details of available therapies; and infectious prevention control measures for patients and caregivers. a telephone number (on-call h) for emergent conditions was also provided [ ] . the local ethics committee approved the study protocol. we prospectively recorded participants' clinical and biological characteristics at the time of carv polymerase chain reaction (pcr) test screening, including the month and year of vaccination and the influenza vaccination status at each flu season. variables included the immunodeficiency scoring index (isi) [ ] and basel immunodeficiency grading [ , ] results; hospital admissions; the use of immunosuppressant drugs; the presence of signs or symptoms of acute or chronic gvhd; and transplant characteristics. with the aim of retrospectively comparing the influenza rtd prevalence between vaccinated and non-vaccinated participants, we included recipient/rv episode/vaccination status in each flu season, following the selection algorithm described in figure . we designed the case selection as follows: the first inclusion criterion was to retrospectively select all rvi episodes (irrespective of the carvs detected) with known vaccination statuses. the next step was to divide the carv episodes into groups according to the vaccination status at each rvi episode (vaccinated or unvaccinated, in each corresponding flu season). following recommendations from the world health organization's guidelines [ ] , we excluded the carv episodes taking place outside the flu seasons (between june and november) from both groups. we considered december to may as recruitment periods, according to our local and national epidemiological data, since influenza viruses were circulating in our community. the third step was to exclude episodes that occurred before day after a stem cell infusion from the unvaccinated group, since the vaccine was not given to any recipients during that period and the first recorded influenza vaccine was given at day after transplant. for both cohorts, we also excluded rvi episodes when baseline disease relapses occurred before the carv episode. for recipients with more than rvi episode in the same flu season, we applied the following criteria: recipients with more than episode of carv rvi other than influenza during the same flu season were computed only once and classed as not having an influenza infection in that season. recipients with more than episode of rvi and whose vaccination status changed between consecutive carv rvi episodes in the same season were computed twice: once in the non-vaccinated group and once (after vaccination) in the vaccinated group. if a recipient developed rvi episodes caused by different influenza viruses (ie, influenza a h /n and b) at different time intervals during the same flu season, both episodes were computed in the corresponding group. when a recipient developed influenza virus rvi episodes in the same season and the same influenza virus was detected, only the first episode was computed. finally, when a recipient developed rvi episodes, of them caused by the influenza virus, we only computed the influenza episode in that season. annual influenza vaccination was recommended to all patients at both transplant centers after the third month following allo-hsct. recipients received a seasonal inactivate trivalent influenza vaccine according to the national health vaccination program that ran from november until february in each flu season. for patients with moderate to severe gvhd at the time of the vaccination program who had received gammaglobulin, anti-thymocytic globuline, or rituximab within the months before the flu vaccine period, vaccine administration remained at the physician's discretion. urtd was defined as a combination of upper-respiratory tract symptoms (rhinorrhea, sinusitis, otitis, or pharyngitis), as well as the positive identification of a carv by a pcr test and the absence of lower-respiratory tract infection symptoms and/ or any indication of pulmonary infiltrates in chest x-ray or computed tomography (ct) scan radiology results. we classified lrtd as possible, probable, or confirmed, as previously described [ ] . there were no probable episodes, because we did not perform bronchoscopies in patients without the radiological proof of pulmonary involvement. we defined episodes as an urtd or lrtd according to european conference on infections in leukaemia- recommendations [ ] . acute gvhd was diagnosed and graded according to the standard criteria [ ] . patients with urtd symptoms underwent nasopharyngeal aspiration, nasopharyngeal swabs, or an induced sputum test, while bronchoalveolar lavage (bal) was performed in patients with a lrtd whenever possible. carv testing in bal samples were performed with real-time pcr multiplex platforms, as described elsewhere in detail [ ] . at the hcuv, samples were tested by real-time pcr using the luminex xtag rvp fast v assay (luminex molecular diagnostics, toronto, canada), while at hlf the clart pneumovir dna array assay (genomica, coslada, spain) was performed and interpreted following the manufacturer's recommendations. the clart pneumovir dna array assay differs from the luminex xtag rvp fast assay in that it detects influenza c virus, but not alphacoronavirus nl virus or betacoronaviruses hku and oc . the clart pneumovir identifies the new influenza a/h n v. the primary objective of the study was to compare the prevalence of influenza urtds and/or lrtds and clinical characteristics among vaccinated and unvaccinated allo-hsct recipients. secondary endpoints included identifying the risk factors for both influenza rtd and progression of the influenza virus from an urtd to a lrtd. frequencies were compared using the χ test (fisher exact test) for categorical variables. differences between medians were compared using the mann-whitney u test. univariate and multivariate analyses of the association of clinical and biological risk factors with the progression of influenza lrtd were calculated using logistic regression models. for multivariate analyses, only variables with parameter estimates showing a p value ≤ . in the univariate analysis were finally included. we reported -sided exact p values, and p values ≤ . were considered statistically significant. the data were analyzed with the spss (version . ) statistical package. overall, allo-hsct recipients developed episodes of upper-and/or lower-respiratory tract symptoms and were screened for rvis. in total, allo-hsct recipients ( %) developed at least episode of a virologically-documented rvi, accounting for episodes ( %) of proven rvis. according to the algorithm selection (figure ), we finally included allo-hsct recipients with virologically-documented rvi episodes over flu seasons. out of recipients, were computed twice since they changed their vaccination status during the course of the study: during the same season and within consecutive seasons. thus, we finally included the characteristics of recipient cases according to the subjects' vaccination statuses (table ). there were seasonally-non-vaccinated allo-hsct recipients with rvi episodes and seasonally-vaccinated recipients with rvi episodes who accomplished the selection criteria for comparison purposes. of the participants, ( %) had computable rvi episodes in multiple flu seasons: specifically, had computable rvi episode in each of consecutive seasons, whereas recipients had computable episode in each of consecutive seasons. patients' clinical and biological characteristics at the time of their rvi episodes are summarized in table according to their vaccination statuses. as expected, the non-vaccinated group had significantly higher rates of conditions related to poor serological influenza vaccine responses. active gvhd, steroid therapy, ongoing immunosuppression therapy, and hypogammaglobulinemia were significantly over-represented in the non-vaccinated group as compared to the vaccinated group ( % vs %, % vs %, % vs %, and % vs %, respectively; p < . for all comparisons). of note, the rate of low total lymphocyte counts at the time of rvis at different cut-offs (< , < , and < ) were not statistically different among groups. we accounted for proven influenza rvi episodes over flu seasons in recipients, with a median time of onset of days after stem cell infusion (range - ). we observed a higher number of influenza rvi episodes in seasons- - (n = ) and - (n = )-as shown in table and figure . together, these seasons represent % of all influenza episodes over the flu seasons. there were episodes limited to urtd, whereas ( %) involved the lower respiratory tract ( possible and proven lrtds). the rate of hospitalization directly attributable to the influenza virus was % ( out of episodes). the most common influenza virus subtype was a, in episodes ( %; h n , h n , and the remainder not subtypable), and b, in episodes ( %). in cases, influenza a and b were detected in the same episode/sample and were classed as co-infections. the observed -season prevalence of influenza rvis was significantly higher in the non-vaccinated ( %) compared to the vaccinated group ( %; p = . ). this statistical difference was even higher regarding the influenza lrtd prevalence ( % in non-vaccinated vs % in the vaccinated group, p = . ). the progression rates of urtds to lrtds in recipients with influenza rvis were % ( out of ) in the non-vaccinated group compared to % in the vaccinated group ( out of ; p < . ). influenza-related hospital admissions were more common in the non-vaccinated group compared to the vaccinated group ( % vs %, p < . ). logistic regression univariate and multivariate analyses of risk factors for influenza virus respiratory infections and progressions to lrtds are shown in table . in order to identify the conditions associated with influenza virus infections, we studied the evaluable recipient/episode pairs. a multivariate analysis identified the flu vaccine as the main factor associated with a reduced risk of influenza virus infection (odds ratio [or] . , % confidence interval [ci] . - . , p = . ). to analyze the risk factors for lrtd progression of an influenza virus infection, we focused the analysis on a multivariate analysis identified independent conditions associated with lrtd progression. again, the flu vaccine was associated with a lower probability of lrtd progression (or . , % ci . - , p = . ). in contrast, a high-risk isi score predicted a higher probability of the influenza virus lrtd progression (or , % ci . - , p = . ). this study shows that, irrespective of the flu season, a trivalent influenza inactivated vaccine given after allo-hsct was the most important factor associated with the lower prevalence of influenza rvis among recipients with proven carv rvis. in allo-hsct recipients with influenza rvis, a multivariate analysis showed that influenza vaccination was associated with a lower probability of the influenza virus progressing to lrtd. we also observed that a high-risk isi score was highly predictive of influenza lrtd progression in our cohort of patients. although this study was not designed to assess the vaccination rate in our population, only % of the recipients with proven carv rvis received the influenza vaccine over flu seasons. this agrees with prior epidemiological studies and enquiries, where the vaccination rate has ranged from - % after allo-hsct [ ] [ ] [ ] . however, some factors merit consideration when interpreting the reported vaccination rate. we did not detect all vaccinated recipients in our prospective respiratory virus survey, since those who were vaccinated may have had lower incidences of carv rvis and a lower propensity to seek medical attention or testing. however, the lack of consensus across current guidelines [ ] [ ] [ ] [ ] regarding the timing and conditions in which the influenza vaccine should be administered, in particular when immunosuppressant therapy is required to treat active/uncontrolled moderate-to-severe gvhd, is likely the most important contributor to this apparently low vaccination rate. in these scenarios, physicians may decide to defer vaccination. thus, based on our findings, we currently recommend flu vaccination at months post-transplant to all of our recipients, irrespective of their immunosuppression status. in our selected cohort, half ( %) of the unvaccinated recipients developed influenza rvis, with moderate to severe clinical consequences, as reflected in higher hospital admission and lrtd progression rates. to address the clinical efficacy of influenza vaccination in recipients with gvhd, corticosteroid therapy, hypogammaglobulinemia, or with ongoing immunosuppressant use, we compared the influenza rvi prevalence in vaccinated recipients according to the presence or absence of such conditions. although the number of cases was limited, we did not find statistical differences regarding vaccinated recipients with or without gvhd ( % vs %, respectively, p = . ), corticosteroids ( % vs %, p = . ), immunosuppressants ( % vs %, p = . ), or hypogammaglubulinemia ( % vs %, p = . ). in fact, our multivariate analysis revealed that vaccination status was the main condition associated with a lower influenza rvi prevalence in recipients with at least episode of a carv rvi. this is an important finding, since we provide clinical evidence that seasonal influenza vaccines could be clinically beneficial in allo-hsct recipients, even when a significant number of vaccinated recipients had gvhd ( %), used immunosuppressants ( %), or used corticosteroids ( %). although the serological vaccination response in such recipients is poor, we can speculate that vaccination may also exert a cellular-mediated response. influenza vaccination was able to mediate peripheral blood t-cell responses, characterized by production of the th cytokine ifn-gamma by cd + cells, both in patients vaccinated more than months after transplantation and those vaccinated earlier, after stem cell transpantation [ ] . these data suggest that, even in cases where the expected serological response could be suboptimal (allo-hsct < mo, gvhd, immunosuppressant use, or corticosteroid use), the influenza vaccine could elicit a clinical benefit through a cellularly-mediated effect, reducing the influenza infection prevalence and/or its severity. although the influenza rvi prevalence in vaccinated recipients was still substantial ( %), it should be noted that the risk factors for influenza lrtd progression identified vaccination as a condition associated with a reduced rate of lrtd progression. furthermore, vaccination was also associated with a lower hospital admission rate. given these findings, we speculate that vaccination could mitigate the severity of influenza rvis, even in the presence of conditions related to decreased serological responses. another relevant finding was the usefulness of the isi score in stratifying the lrtd progression risk of influenza rvis, even when we analyzed episodes occurring after day after stem cell infusion. isi was developed by investigators from the md anderson cancer center to predict outcomes in allo-hsct recipients with respiratory syncytial viruses [ ] . the same investigators demonstrated its value in predicting lrtd progression in the setting of influenza rvis [ ] . therefore, the use of the isi could be applied to assess the need for prophylactic/therapeutic intervention with several doses of the influenza vaccine and/or with high doses of antiviral drugs in prospective studies. abbreviations: alc, absolute lymphocyte count; allo-hsct, allogeneic hematopoietic stem cell transplantation; anc, absolute neutrophil count; atg, anti-thymocytic globuline; basel ig, basel immunodefciency grading; ci, confidence interval; gvhd, graft-versus-host disease; hla, human leucocyte antigen; igg, immunoglobuline g; is, immunosuppressants; isi, immunodeficiency score index; log. regr., logistical regression model; lrtd, lower-respiratory tract disease; ns, not significant; nt, not tested; or, odds ratio; rtd, respiratory tract disease; rvi, respiratory virus infection. a these variables were not included in the final multivariate models, since they were included in the isi score. regarding influenza virus epidemiological data from the current study, most influenza rvi cases occurred in the - and - flu seasons. these data are in accordance with spanish epidemiological data, where the influenza prevalence was significantly higher in - ( cases/ hab) and - ( cases/ hab) as compared to the - , - , and - flu seasons ( , , and cases/ hab, respectively) [ , ] . these observations confirm that influenza rvi prevalence in our allo-hsct recipients mimicked influenza virus prevalence in the general population. finally, we acknowledge that this study has some important limitations, such as the retrospective nature of the analyses, the small sample size, the somewhat heterogeneous vaccination policy, and the cohort selection method. in addition, the use of different pcr methods, which differed (minimally) in their analytical performance, can be viewed as a limitation. in spite of this, our study has strengths that merit consideration. we used molecular testing. our prospective carv survey mirrored national epidemiological data in influenza rvis in each flu season. we provided data encompassing complete flu seasons, limiting the distortion likely introduced by the vaccination coverage variability between seasons. in conclusion, we provide clinical evidence that influenza vaccination after allo-hsct is associated with a lower prevalence of influenza rvi and a lower severity of the disease. 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diseases society of america. idsa clinical practice guideline for vaccination of the immunocompromised host th european conference on infections in leukaemia guidelines for vaccination of patients with hematological malignancies and hsct recipients european guidelines for prevention and management of influenza in hematopoietic stem cell transplantation and leukemia patients: summary of ecil- the benefit of influenza vaccination after bone marrow transplantation a double-blind, randomized trial of highdose vs standard-dose influenza vaccine in adult solid-organ transplant recipients repeated vaccination is required to optimize seroprotection against h n in the immunocompromised host immunogenicity of virosomal adjuvanted trivalent influenza vaccination in allogeneic stem cell transplant recipients epidemiologic and clinical characteristics of coronavirus and bocavirus respiratory infections after allogeneic stem cell transplantation: a prospective single-center study immunodeficiency scoring index to predict poor outcomes in hematopoietic cell transplant recipients with rsv infections community-acquired respiratory paramyxovirus infection after allogeneic hematopoietic cell transplantation: a single-center experience outcome of influenza infections in outpatients after allogeneic hematopoietic stem cell transplantation evaluation of influenza vaccine effectiveness: a guide to the design and interpretation of observational studies world health organization parainfluenza virus lower respiratory tract disease after hematopoietic cell transplant: viral detection in the lung predicts outcome ecil- ): guidelines for diagnosis and treatment of human respiratory syncytial virus, parainfluenza virus, metapneumovirus, rhinovirus, and coronavirus clinical manifestations of graft-versushost disease in human recipients of marrow from hl-a-matched sibling donors comparison of the performance of commercial multiplex pcr platforms for detection of respiratory viruses in upper and lower tract respiratory specimens toronto invasive bacterial diseases network. invasive pneumococcal disease in adult hematopoietic stem cell transplant recipients: a decade of prospective population-based surveillance vaccination adherence in hematopoietic stem cell transplant patients: a pilot study on the impact of vaccination cards and reminder telephone calls vaccination guidelines after hematopoietic stem cell transplantation: practitioners' knowledge, attitudes, and gap between guidelines and clinical practice outcomes of influenza infections in hematopoietic cell transplant recipients: application of an immunodeficiency scoring index informe semanal de vigilancia de la gripe en españa key: cord- - o dssj authors: waggoner, jesse j.; soda, elizabeth a.; deresinski, stan title: rare and emerging viral infections in transplant recipients date: - - journal: clin infect dis doi: . /cid/cit sha: doc_id: cord_uid: o dssj emerging viral pathogens include newly discovered viruses as well as previously known viruses that are either increasing, or threatening to increase in incidence. while often first identified in the general population, they may affect transplant recipients, in whom their manifestations may be atypical or more severe. enhanced molecular methods have increased the rate of viral discovery but have not overcome the problem of demonstrating pathogenicity. at the same time, improved clinical diagnostic methods have increased the detection of reemerging viruses in immunocompromised patients. in this review, we first discuss viral diagnostics and the developing field of viral discovery and then focus on rare and emerging viruses in the transplant population: human t-cell leukemia virus type ; hepatitis e virus; bocavirus; ki and wu polyomaviruses; coronaviruses hku and nl ; influenza, h n ; measles; dengue; rabies; and lymphocytic choriomeningitis virus. detection and reporting of such rare pathogens in transplant recipients is critical to patient care and improving our understanding of posttransplant infections. emerging infectious diseases are caused by pathogens that are newly recognized or whose incidence has either increased in the preceding decades or threatens to increase. viral diseases account for a large proportion of such infections. in the context of transplant recipients, important emerging viruses can be considered to be of types: ( ) novel viruses; ( ) known viruses increasing in incidence in the general population and, potentially, in transplant recipients; and ( ) previously known viruses that cause disease of increased severity in the immunocompromised host. in this review, we begin by discussing viral diagnostics and the evolving field of viral discovery, which has increased the speed of virus identification but has created new challenges. our focus then shifts to specific emerging and reemerging viral pathogens in the transplant community. viruses described in case series or multiple case reports are listed in table . viruses described only in single case reports are listed in table . the potential risks of viral transmission as the result of xenotransplantation will not be addressed [ ] . viral discovery has typically relied on the ability to detect new viruses in cell culture. although clinical virology laboratories affiliated with transplant centers routinely perform viral culture, many pathogens do not grow well or do not grow at all, and viral detection using culture is further limited by the number of cell lines a laboratory can realistically maintain. pathogen detection in the clinical laboratory is also limited by the available tests, which often target conserved sequences (polymerase chain reaction [pcr] or real-time [rt]-pcr) or specific antigens or antibodies to detect known viruses. multiplex testing for clinical syndromes, particularly for respiratory-tract infections, allows for a less biased approach to viral diagnosis but still faces limitations in identifying emerging pathogens [ ] . in rare situations, an unusual virus may be detected by testing for known pathogens, as in the case of a woman who presented with usutu viremia, which gave a low-positive result by west nile virus rt-pcr [ ] . a number of more rapid molecular methods are now being employed in viral discovery, categorized as sequence dependent (such as the pan-viral microarray) or sequence-independent techniques [ ] . the pan-viral microarray is an array spotted with oligonucleotide sequences representing known viral pathogens. novel viruses can be identified if sufficient similarity exists between sequences in the new virus and those on the array. amplicons can then be recovered from the array, then cloned and sequenced [ ] . this technology was used in the identification of sars coronavirus from a cultured patient isolate [ ] . pcr based on conserved sequences generally has limited applicability in viral diagnostics, as viruses do not contain highly conserved sequences analogous to s ribosomal rna sequences utilized in bacterial identification [ ] . the sequence-independent amplification and sequencing of viral nucleic acids in biological samples has been termed viral metagenomics [ , ] . sequence-independent approaches include subtractive hybridization or representation difference analysis, sequence-independent single-primer amplification, and rolling circle amplification. these techniques have been used to identify agents such as human herpes virus , torque teno viruses (ttv), hepatitis e virus (hev), norwalk virus, parvovirus , and human bocavirus (hbov) [ , ] . viral metagenomics has been aided by the development of a number of new sequencing platforms. termed next-generation sequencing (ngs, or deepsequencing), such technologies allow for the rapid and parallel generation of to over sequences per run. most current technologies rely on nonspecific amplification of viral dna or rna from samples treated to remove host nucleic acids. amplification is followed by sequencing by synthesis using different technologies to detect base incorporation [ ] [ ] [ ] . ngs has been utilized to identify novel viruses in patient samples and in studies of fevers of unknown origin [ , ] . ngs has a great ability to detect both known and previously unknown (divergent) viruses, but mere detection does not demonstrate causation. for many of these viruses, classical koch's postulates cannot be applied, and as demonstrated with ttv and hbov, establishing a causative role for these agents can be difficult [ , , ] . mokili and colleagues [ ] proposed "metagenomic koch's postulates," but whether they are sufficient remains moot. at this time, ngs is largely a tool for research purposes. sequencing reactions take a good deal of time to set up and perform [ ] . these runs generate massive amounts of data that must be filtered prior to analysis using various alignment programs designed to handle the large numbers of short reads [ , ] . finally, results must be interpreted carefully. contaminants from the laboratory and even from commercial reagents are often identified (eg, xenotropic murine leukemia virus-related virus), and confirming the presence of a virus identified with small numbers of reads may not be possible [ , ] . human t-cell leukemia virus type human t-cell leukemia virus type (htlv- ) seroprevalence rates range from % to % in endemic areas, to < % in western countries [ ] . chronic htlv- infection is associated with adult t-cell leukemia (atl) and htlv- -associated myelopathy (ham) in % or fewer of those infected, but there is concern that immunosuppression in htlv- -positive transplant recipients may trigger progression to these complications [ ] [ ] [ ] . yoshizumi et al [ ] identified htlv- -positive, living donor liver transplant recipients. atl developed in patients at - days post transplantation; all patients died, including from atl. overall survival rates did not differ between htlv- -positive recipients and htlv- -negative liver transplant recipients from the same institution [ ] . case reports of atl following renal transplantation in htlv- positive patients have been documented, though in case series of renal transplant recipients (totaling patients with - years of follow-up), no cases of atl or ham developed [ ] [ ] [ ] [ ] . ham has been reported in htlv- d+/r+ living-related liver transplant recipients [ ] . atl responds poorly to conventional chemotherapy, with the highest median survival rates reported in clinical trials being approximately months [ ] . as a consequence, hematopoietic stem cell transplantation (hsct) has been evaluated for the treatment of atl in htlv- -positive patients. (hsct will be used to describe the transplantation of multipotent stem cells from bone marrow, peripheral, or cord blood.) the largest study of hsct for atl involved the retrospective analysis of patients with atl who had undergone an allogeneic hsct at centers in japan [ ] . their -year survival rate was %; atl recurred in % of patients who survived to days post transplant. those who received transplants from a related htlv- seropositive donor had a higher risk of disease-associated mortality relative to those whose related donor was htlv- negative. hsct recipients in complete remission at the time of transplantation had a higher rate of survival compared to patients not in complete remission ( % vs %) [ ] . the transmission of htlv- through transplantation or transfusion has been documented. in spain, htlv- -negative recipients of organs from a single htlv- -positive donor ( liver and kidney transplants) developed ham within years of transplantation [ ] . two case reports document the occurrence of ham in a heart transplant recipient and an hsct recipient who acquired htlv- through blood transfusions [ , ] . in low-prevalence areas, however, universal donor screening with enzyme-linked immunosorbent assay followed by western blotting resulted in many false positives, and the practice is no longer recommended by the united states organ procurement and transplantation network [ ] . hev is a common cause of acute liver disease in the developing world, primarily from fecal-oral spread through contaminated drinking water. infections in developed nations are well described, but typically result from the consumption of undercooked pork products. in immunocompetent hosts, hev acute infection is self-limited with rare progression to fulminant liver failure. however, in the immunosuppressed host, chronic infection marked by persistent viremia and abnormal liver function with eventual progression to cirrhosis can occur [ ] [ ] [ ] . cases have been described in recipients of a variety of transplants, including kidney, liver, heart, and lung [ , ] . in a multicenter review of cases of acute hev infection, . % of the solid organ transplant (sot) recipients developed chronic hepatitis of whom . % developed cirrhosis. the use of tacrolimus compared with cyclosporine a was an independent predictor of chronic infection [ ] . rare cases of encephalitis and polyradiculopathy with hev rna detection in the cerebrospinal fluid (csf) have also been described [ ] . the majority of hev infections following sot result from de novo infections and are unlikely to represent virus reactivation [ , ] . rare instances of transmission through blood transfusion or the donated graft have been reported [ ] . determining the overall incidence of hev-related disease following transplantation is hampered by the available diagnostic tests, none of which are food and drug administration (fda) approved. commercial serological assays have variable test characteristics, and tests for hev rna detection in serum or stool samples are not routinely available [ ] . reports of hev infection following hsct are limited. while an individual case of hev reactivation following hsct has been reported, a review of anti-hev immunoglobin gpositive patients prior to hsct showed no evidence of disease reactivation [ ] . treatment of prolonged hev viremia often involves reducing immunosuppression. pegylated interferon administration has been shown to induce a sustained virologic response in a limited group of patients [ ] . both approaches to viral control may increase the risk of graft rejection. ribavirin monotherapy has induced sustained virologic responses without the risk of rejection and may represent the first-line agent for treatment [ ] . there are clinically relevant non-sars human coronaviruses (hcov): oc ; e; hku and nl , both identified in the last decade; and the middle east respiratory syndrome hcov (mers-cov), identified in . a prospective study found that % of hcov infections were asymptomatic and none of infected allogeneic hsct recipients developed lower-respiratorytract infection, although prolonged viral excretion is frequent [ ] . nonetheless, there have been reports of fatal hcov infection following hsct. there is evidence among sot recipients that hcov can cause severe lower-respiratory-tract infections and increase the risk of graft rejection [ , ] . hcov-hku and nl do not appear to be more virulent than the previously discovered hcov-oc and e; however, the recently identified mers-cov has been associated with severe pneumonia and a high mortality rate. cases have not yet involved immunocompromised hosts. efforts to identify novel respiratory pathogens have led to the discovery of hbov and ki and wu polyomaviruses (kipyv and wupyv) [ , [ ] [ ] [ ] . while these viruses have been detected in patients with respiratory symptoms, evidence to support a causative role for these agents in severe disease is lacking [ , ] . studies evaluating hbov as a respiratory pathogen in immunocompromised adults have detected the virus infrequently and have not documented an effect on patient outcomes [ , ] . the establishment of hbov as a respiratory pathogen has also been complicated by high rates of copathogen detection and hbov detection in asymptomatic patients [ , ] . viral dissemination in transplant recipients occurs, with hbov detected in blood and stool. however, patients often had hbov detected after weeks of hospitalization, and other pathogens were also detected during these episodes [ , ] . some commercial platforms for multiplex detection of respiratory pathogens include hbov. no specific antiviral treatment is available [ ] . kipyv and wupyv have been detected in nasopharyngeal and bronchoalveolar lavage samples from sot and hsct recipients [ ] [ ] [ ] . detection of these viruses has been associated with sputum production and wheezing following hsct [ ] . however, similar to hbov, these viruses are often codetected with other pathogens, and they have not been associated with severe respiratory tract disease or mortality [ , ] . immunocompromised hosts are more susceptible to complications of influenza; however, it is not clear that emerging strains will necessarily cause more severe disease. one case series of sot patients with h n influenza showed that % required icu admission and % died [ ] . a series among hsct recipients showed similar findings [ ] . in a comparison of outcomes in kidney transplants and immunocompetent patients with h n , there were no differences in morbidity or mortality [ ] . to date, no cases of h n or h n influenza have been reported in transplant recipients. the recent rise in measles incidence brings it into consideration here. the most significant manifestation may be subacute measles encephalitis (sme), though severe cases of pneumonia have been documented [ ] . sme has developed in renal transplant recipients and a single hsct recipient. patients may present with a measlescompatible illness, which improves. they develop altered mental status and seizures - weeks later; fever is uncommon. the first imaging changes are seen by magnetic resonance imaging, and diagnosis is confirmed by immunoglobin m (igm) seroconversion or rt-pcr. the clinical course is one of deteriorating mental status and treatment-refractory seizures [ ] . four of transplant cases of sme have died. the survivors both had significant neurological deficits [ ] . the incidence of measles in transplant recipients, as well as the proportion with severe disease, is unclear. two series identified cases of interstitial pneumonia ( fatal) among hsct recipients diagnosed with measles, though methodological limitations existed in both studies [ ] . dengue virus (denv) is the most common vector-borne viral disease worldwide and has been detected in an increasing number of countries over the last years. in case series involving renal transplant recipients, only a single case of severe dengue developed, with no fatalities or loss of graft function [ , ] . severe cases of dengue, including deaths, have been reported in renal transplant recipients along with fatal cases in a liver transplant recipient and an hsct recipient [ ] . in patients who died, disease typically developed within the first month post transplant. human-to-human transmission of denv as a result of sot or hsct has been postulated, and transfusion-related denv infections have been reported [ ] . fda-approved diagnostics include tests for igm detection and a centers for disease control and prevention-developed rt-pcr; management consists of supportive care. seventeen cases of rabies have been reported in transplant recipients, and to date, all have been transmitted through the transplanted tissue or organ [ , ] . nine cases followed corneal transplantation, including deaths [ ] . the sole survivor, reported in , began postexposure prophylaxis (pep) on postoperative day [ ] . two clusters (texas, , and germany, ) , totaling rabies cases, have occurred following sot [ , ] . these cases followed the transplantation of liver, lung, kidney, kidney-pancreas, and iliac artery grafts. patients typically developed encephalitis between and days post transplant, and all symptomatic patients died [ ] . patients in germany received pep and antiviral treatment, though not until postoperative day [ ] . the liver recipient in this cluster had been previously vaccinated and never developed disease [ ] . both donors were later determined to have rabies exposures (bat and dog bites, respectively) [ , ] . a recent report (maryland) documented a fatal case of rabies developing a year after kidney transplant. transmission of raccoon-variant rabies through the donated graft was confirmed. three other graft recipients from the same donor are alive, though full details are not available [ ] . the management of rabies focuses on prevention with vaccination in high-risk patients or pep. transplant recipients who receive pep can mount adequate responses (antibody titers of . international units/ml), though titers are lower than in immunocompetent patients [ ] . based on the experience of the german liver transplant recipient, rabies vaccination may remain effective even after transplantation. cases of lymphocytic choriomeningitis virus (lcmv) transmitted through organ transplantation ( clusters, including cases and deaths) document the ability of this pathogen to cause severe disease in the immunocompromised host [ , [ ] [ ] [ ] . another cluster involved the transmission of a related arenavirus in australia, with similar outcomes ( liver and kidney recipients; deaths) [ ] . as with rabies infections post transplant, all cases resulted from transmission through organ transplantation [ , [ ] [ ] [ ] . at this time, cases have not been described in the hsct population. the case clusters of lcmv infection occurred in the united states and involved kidney, liver, and lung transplants [ ] [ ] [ ] . symptoms developed between and days post transplant and included fever, abdominal pain, nausea, diarrhea, and altered mental status. patients often developed a peri-incisional rash and tenderness. csf findings included elevated protein (often marked), normal to low glucose, and a mild pleocytosis [ ] [ ] [ ] [ ] . three patients survived lcmv infection following sot, kidney recipients and a liver recipient. ribavirin has been employed in some cases, though the benefit remains unclear [ ] . three corneal transplant recipients were potentially exposed to lcmv, though none of them developed symptoms or seroconverted [ ] . contact investigation revealed exposure to rodents or positive testing for lcmv in donors [ ] [ ] [ ] . investigation into the fourth donor revealed no exposure, and all testing performed on remaining tissues was negative [ ] . it has been advised that immunocompromised patients avoid contact with rodents, including pets, though this was not the mode of lcmv acquisition in these outbreaks [ ] . for the majority of viral infections discussed here, data are insufficient to determine the true incidence of disease in transplant recipients. measles, mumps, and yellow fever are vaccinepreventable illnesses, though these vaccines are live-attenuated and not recommended following transplantation. also, antibody response to vaccines is less than in immunocompetent patients. donor-transmitted rabies carries a dire prognosis, and though limited data exist, the use of pep in transplant recipients appears safe. given their apparent rarity, screening for many of these diseases in organ donors is not recommended. the examples of htlv- (discussed earlier) and lcmv are illustrative of some of the difficulties involved with donor screening. in the outbreak investigations for lcmv, only of donors had detectable antibodies. indeed, rt-pcr from multiple samples failed to detect lcmv from donor, and yielded a positive result in only a single lymph node in another [ ] [ ] [ ] . it seems prudent to obtain a comprehensive history of potential organ donors, though it remains unclear how certain findings, such as rodent ownership, should affect one's donor status. reporting rare infections in transplant recipients will help to identify agents for which more research is needed and screening may be warranted. however, it is likely that these infections are underdiagnosed as symptoms may be attributed to more common, and potentially coincident, posttransplant infections. xenotransplantation-associated infectious risk: a who consultation rare and emerging viral infection in the transplant population usutu virus infection in a patient who underwent orthotropic liver transplantation viral metagenomics viral discovery and sequence recovery using dna microarrays metagenomics and future perspectives in virus discovery from orphan virus to pathogen: the path to the clinical lab next-generation sequencing and its applications in molecular diagnostics 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analysis dengue has a benign presentation in renal transplant patients: a case series non vector-borne transmission modes of dengue centers for disease control and prevention. cdc confirms rabies death in organ transplant recipient prevention of inter-human rabies transmission after corneal graft management and outcomes after multiple corneal and solid organ transplantations from a donor infected with rabies virus transmission of rabies virus from an organ donor to four transplant recipients lymphocytic choriomeningitis virus transmitted through solid organ transplantation -massachusetts transmission of lymphocytic choriomeningitis virus by organ transplantation solid organ transplant-associated lymphocytic choriomeningitis, united states lymphocytic choriomeningitis virus infection in organ transplant recipients -massachusetts update: interim guidance for minimizing risk for human lymphocytic choriomeningitis virus infection associated with pet rodents acknowledgments. the authors thank dr benjamin a. pinsky for his comments on this manuscript. we also thank dr jose g. montoya for his support during the preparation of this review.potential conflicts of interest. all authors: no reported conflicts. all authors have submitted the icmje form for disclosure of potential conflicts of interest. conflicts that the editors consider relevant to the content of the manuscript have been disclosed. key: cord- - lripsf authors: piñana, josé luis; gómez, maría dolores; montoro, juan; lorenzo, ignacio; pérez, ariadna; giménez, estela; gonzález‐barberá, eva maría; carretero, carlos; guerreiro, manuel; salavert, miguel; sanz, guillermo; hernández‐boluda, juan carlos; borrás, rafael; sanz, jaime; solano, carlos; navarro, david title: incidence, risk factors, and outcome of pulmonary invasive fungal disease after respiratory virus infection in allogeneic hematopoietic stem cell transplantation recipients date: - - journal: transpl infect dis doi: . /tid. sha: doc_id: cord_uid: lripsf background: there is growing evidence that community‐acquired respiratory virus (carv) increases the risk of pulmonary invasive fungal disease (ifd) in the allogeneic hematopoietic stem cell transplantation (allo‐hsct) setting. to date, there is a lack of knowledge regarding the risk factors (rfs), as well as the most critical period for subsequent onset of ifd after carv infections in allo‐hsct recipients. methods: in this prospective longitudinal observational carv survey, we analyzed the effect of carv on subsequent ifd development in adult allo‐hsct recipients diagnosed with carv episodes from december to december . multiplex pcr panel assays were used to test carvs in respiratory specimens. findings: twenty‐nine out of allo‐hsct recipients ( %) developed ifd after a carv episode. the median time of ifd onset was days (range, ‐ days) from day of the first carv detection. generalized estimating equation model identified risk factors for ifd: atg‐based conditioning regimen [odds ratio (or) . , % confidence interval (ci) . ‐ . , p = . ], carv lower respiratory tract disease (or . , % ci . ‐ . , p < . ), carv infection during the first year after transplant (or . , % ci . ‐ . , p = . ), and corticosteroids during carv (or . , % ci . ‐ . , p = . ). conclusion: allo‐hsct recipients conditioned with atg and under corticosteroid therapy at the time of carv lrtd during the first year after transplant may require close monitoring for subsequent ifd. pulmonary invasive fungal disease (ifd) is a relevant cause of morbidity and mortality in the allogeneic hematopoietic stem cell transplantation (allo-hsct) setting. ifds in allo-hsct [mainly for invasive aspergillosis (ia)] include factors associated with a profound immunosuppressed status, such as duration of neutropenia before engraftment, use and duration of glucocorticosteroid therapy, type of donor, baseline disease, recipient's age, and graft-versus-host disease (gvhd). [ ] [ ] [ ] [ ] [ ] [ ] [ ] however, there is growing awareness and evidence that community-acquired respiratory virus (carv) also increases the risk of ia in immunocompromised and healthy patients, primarily but not limited to influenza infections. , [ ] [ ] [ ] [ ] [ ] [ ] [ ] allo-hsct recipients suffer annual and lifelong exposure to carv infections; in fact, these infections after allo-hsct are as common as around %. , obviously, not every allo-hsct recipient with carv infection has the same risk of developing ifd. to date, there are limited data regarding the true rate of ifd, the rfs for specific respiratory virus infection, and the most critical period for ifd development after carv infections in allo-hsct recipients. the current study comes from a -year prospective longitudinal observational epidemiologic survey of carv infections and complications in allo-hsct recipients with upper (urtd) and/or lower respiratory tract disease (lrtd) symptoms after allo-hsct. we report herein the incidence rate, characteristics, and rfs of ifd in a series of consecutive allo-hsct recipients after a molecular-proven carv infection. we conducted a prospective longitudinal study of carv infections in adult (> years) allo-hsct recipients at two transplant centers in the carv survey methodology has previously been described in detail elsewhere. , briefly, at both centers, we developed a prospective carv survey in allo-hsct recipients with symptoms suggesting a respiratory virus infection. irrespective of their transplant date, all recipients with urtd and/or lrtd symptoms were encouraged to undergo a detailed respiratory virus screening through molecular testing. we do not monitor viral shedding negativity after a first detection, except for recipients with active and persistent respiratory symptoms for longer than one month. in such cases, we repeated the pcr test to rule out the acquisition of a new carv. if the same subtype of carv was found, we considered as a long-lasting carv episode. clinical and biological variables at the time of carv pcr testing were prospectively recorded. variables included transplant characteristics, conditions included in the immunodeficiency scoring index (isi), hospital admission, immunosuppressant drugs, presence of acute or chronic gvhd, use of antifungal prophylaxis at the time of carv infection, and subsequent onset of ifd. all allo-hsct recipients were followed on a regular basis at each transplant center for lifelong. the study protocol was approved by the ethics committee of the hospital universitari i politècnic la fe with reference code number / . at hcuv, from january to february , antifungal prophylaxis was based on voriconazole mg twice daily (bid) from day + to day + and thereafter in cases with active gvhd requiring steroid therapy. from march , antifungal prophylaxis consisted of micafungin ( mg/day iv) from the start of the conditioning regimen until neutrophil recovery and switched to posaconazole mg daily in tablets until day + or while on steroids to treat moderate-to-severe gvhd. at hlf, from january to december antifungal prophylaxis was based on fluconazole ( mg/day) from the start of conditioning until the day of stem cell infusion (day ) and voriconazole mg/bid a day thereafter until day + or while on steroids. in january , antifungal prophylaxis was changed and consisted of fluconazole ( mg/day) during conditioning until day + and posaconazole ( mg/day) thereafter until day + or while on steroids. urtd was considered in recipients with upper respiratory symptoms (rhinorrhea, sore through, sinusitis, otitis, or pharyngitis) and a positive identification of carv by pcr test in respiratory samples and the lack of lrtd symptoms without any sign of pulmonary infiltrates in chest x-ray or ct scan. we classified lrtd as possible or confirmed according to previous definitions. there were no probable episodes since we do not perform bronchoscopies in patients without radiological evidence of pulmonary involvement. we defined carv urtd and/or lrtd episodes following ecil- guidelines. according to the ecil- recommendations, an episode was considered to be resolved when complete remission of respiratory symptoms was observed. in cases with a new onset of respiratory symptoms in which we detected the same virus from the prior carv episode along with another newly emerging carv, we considered it as a new co-viral infectious disease episode. in contrast, cases in which we detected a carv and the recipient had long-lasting respiratory symptoms that were re-screened during follow-up testing positive for a new emerging carv along with the initial carv were considered as a unique co-viral carv infectious disease episode. as per protocol, we started oseltamivir in allo-hsct recipients with influenza infection provided the respiratory symptoms have not been resolved at the time of microbiological results. annual influenza vaccination was also recommended to all patients at both transplant centers after the third month following allo-hsct. for patients with moderate-to-severe gvhd at the time of vaccination program who had received gammaglobulin, antithymocytic globulin (atg), or rituximab within the three months before the flu vaccine period, vaccine administration remained at physician discretion. respiratory syncytial virus (rsv) and human parainfluenza virus (hpiv) were managed according to our interventional protocol. briefly, oral ribavirin was given at a loading dose (maximum daily dose of mg/kg) until resolution of respiratory symptoms in recipients with lrtd caused by rsv or hpiv, whereas patients with urtd must had or more isi points and/or or more risk factor according to the ecil guidelines, and/or presenting one or more co-infective virus(es) before starting ribavirin. routine prophylactic intravenous immunoglobulins ( . g/ kg) were given when igg serum levels were below mg/dl. definitions of the european organization for research and treatment of cancer and mycoses study group (eortc/msg). possible ifd episodes were considered, but finally, they were not included in the analyses. the ifd date was taken as the date the test yielded the diagnosis. ifd was considered to be directly related to carv infection when it occurred in the months following either the date of carv detection or date of most recent carv detection (in cases of longlasting active respiratory symptoms and viral shedding with several consecutive pcr tests detecting the same carv). recipients with probable-proven ifd prior to the development of a carv episode were not considered as a carv-related ifd, and they were not included in the analyses. acute graft-versus-host disease (agvhd) was defined according to standard criteria. carv testing was performed with three rt-pcr multiplex platforms, two of which have been described in detail elsewhere. , briefly, the luminex xtag rvp fast v assay (luminex molecular diagnostics) and the clart ® pneumovir dna array assay (genomica) were used at the hcuv and at hlf, respectively. at hlf from july onwards, carv screening in allo-hsct recipients was performed by biofire filmarray ® respiratory panel week from day of stem cell infusion until hospital discharge and on a weekly basis thereafter until day + and at each outpatient visit while receiving steroids to treat gvhd afterward. gm was also screened in bronchoalveolar lavage fluid. the primary endpoint of the current study was to determine the ifd incidence and its rfs in allo-hsct recipients with prior documented carv, urtd, and/or lrtd. secondary endpoints included the analysis of carv-related ifd characteristics and its effect on overall survival (os). categorical variable frequencies were compared using the χ test (fisher's exact test), whereas the mann-whitney u test was used to compare differences between medians. generalized estimating equation (gee) was used to assess the association of clinical and biological rfs with carv-related ifd. this method is used when analyzing clustered data, as ours, where episodes can be grouped within the same patient. gee fits marginal mean models with asymptotically normal and consistent estimators if there is a correct specification of marginal means. two different models were carried out in the current study: one to evaluate rfs for ifd after overall carv ( ) and those limited to the lower respiratory where response variable is ifd (y = , ) and factors are as follows: atg as a part of conditioning (x = , ), carv lrtd (x = , ), allo- cmv status (x = , ). moreover, bj is the random effect associated with each individual j and sij the experimental error, both are considered random and independent. the method for best model selection was "quasi-likelihood under the independence model criterion" (qic). this method was one hundred and forty-two ( %) out of recipients had carv episode, recipients ( %) had carv episodes, ( %) had three-hundred and ninety-four carv episodes ( %) were limited to the urtd, whereas ( %) had lrtd involvement ( possible lrtd and proven lrtd). antiviral therapy with oseltamivir or ribavirin was given in ( %) out of carv episodes. table summarizes the most common carv types and rates of urtd and lrtd. out of allo-hsct recipients, ( %) developed ifd within two months after a carv episode at a median of days (range - days) from the day of carv diagnosis. in five out of cases ( %), ifd was diagnosed at median of days (range to days) after the initial carv diagnosis. however, these cases had long-lasting respiratory symptoms and repeated pcr test detected the same carv subtype in the two months before the diagnosis of ifd. all the ifds involved the lungs, and in ( %) cases, the diagnostic was ia, meeting criteria of probable (n = ) or proven (n = ). probable ia cases had gm index in blood (n = ) and/or bal (n = ) above the threshold values. gee models of rfs for ifd after overall carv episodes and those with lrtd are shown in table . in order to identify transplant and carv circumstances related to ifd, we included the evaluable recipient/episode pairs. the developing ifd after carv was associated with worse os at three months after carv diagnosis. os was % for carv episodes not developing ifd vs % for those with ifd (p = . ). however, the os in allo-hsct recipients with lrtd developing ifd was similar to those without ifd ( % vs %, respectively, p = . ; figure a ,b). this study shows an overall ifd rate of % ( out of allo-hsct recipients), whereas the rate was % according to overall carv infectious episodes ( out of carv episodes). gee analysis identified atg, carv lrtd, having a carv episode within the first year carv without ifd (n = ) immunodeficiency scoring index, n (%) a anc < . × /l ( pts) ( ) ( ) . alc< . × /l ( pts) ( ) recent or pre-engraftment allo-hsct ( pt) ( ) ( ) . low risk ( - ) ( ) ( ) moderate risk ( - ) ( ) ( ) . high risk ( - ) ( ) ( ) other characteristics a . day + ( ) ( ) . day + ( ) ( ) . abbreviations: alc, absolute lymphocyte count; allo-hsct, allogeneic hematopoietic stem cell transplantation; anc, absolute neutrophil count; atg, antithymocyte globulin; carv, communityacquired respiratory virus; gvhd, graft-versus-host disease; ifd, invasive pulmonary infectious fungal disease; is, immunosuppressants; lrtd, lower respiratory tract disease. a all variables were captured at the time of carv diagnosis. ta b l e clinical and biological characteristics of carv episodes in allo-hsct recipients according to the development of a later invasive pulmonary infectious disease after transplant and corticosteroids during carv as the main rfs for ifd. these rfs led to the stratification of risk groups with statistically significant differences in ifd rates. remarkably, patients at low risk (none of the rfs) had a very low ifd rate ( . %). although the ifd rate was still low ( %) in recipients with or rfs, the recipients with or rfs showed a higher rate of ifd ( %). additionally, we found atg, active gvhd, and a carv episode during the first year of transplant as the main rfs for ifd in recipients with carv lrtd. although the relationship between carvs and invasive mold infections and/or ia has been consistently established in the allo-hsct setting, , , , the true rate of later-occurring ifd in recipients developing carv infections after allo-hsct is not known. data from retrospective series reported rates of ifd after carv in allo-hsct recipients as ranging from % to %. , however, our ifd rates vary according to carv location (urtd or lrtd), time of carv diagnosis, and presence of concurrent rfs. in this sense, the ifd rate in recipients with urtd was %, whereas it was % in those with lrtd. the ifd rate was even higher ( %) in recipients developing a carv lrtd during the first year after stem cell infusion. our data show that ifd rates were lower than previously carv involvement in alterations to the bronchial mucosa or pulmonary defensive mechanisms. , the fact that ifd was a later event in our series [ ( %) of recipients developed ifd after day + ] might explain why neutropenia did not enter as a potential rf in the gee model. in contrast, the use of atg was an important rf for ifd. this finding is not surprising, since atg produces lymphopenia and a significant deferral in t-cell immune reconstitution for longer than a year after stem cell infusion. , atg has been previously identified as a rf for ia. in fact, aspergillus-specific cd + t-cell activity has been recognized as significant in regulating active pulmonary inflammation and likely adding antifungal effector activity. [ ] [ ] [ ] another potential explanation could be that atg in our series was mainly used in the context of single cbu transplantation ( % of overall atg use) and this procedure has been consistently associated with higher incidence of ifd than other sources of hematopoietic stem cells. as per protocol, in the current series, the recipients on corticosteroids therapy received triazole antifungal prophylaxis. however, corticosteroids use was still associated with increased risk of ifd during carv infections in our series. this finding is not completely unexpected since it is well established that corticosteroids increased the risk of ifd in allo-hsct recipients. finally, the development of invasive pulmonary ifd complicating carv infections after allo-hsct had a negative effect on outcome. the dismal prognosis of ifd after allo-hsct is well characterized. in the current series, three-month all-cause mortality was significantly higher in recipients with ifd. this fact highlights the need for effective diagnostic and prevention strategies for detecting and reducing ifd in these severely immunosuppressed patients. we acknowledge that this study has some limitations, such as the use of different antifungal prophylaxis strategies over time and the use of three different pcr methods differing (minimally) in analytical performance. in spite of this, our study has strengths that merit consideration, notably our use of molecular testing and our data collection from a prospective carv survey. in conclusion, we provide evidence that ifd after carv infection is significant, showing a negative association on outcomes. allo-hsct recipients with or rfs may require close monitoring for the future development of ifd. we would like to thank juan a. carbonell-asins from the invasive aspergillosis following hematopoietic cell transplantation: outcomes and prognostic factors associated with mortality incidence and outcome of invasive fungal diseases after allogeneic hematopoietic stem 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hl-a-matched sibling donors longitudinal data analysis using generalized linear models akaike's information criterion in generalized estimating equations r: a language and environment for statistical computing the r package geepack for generalized estimating equations package 'mumin'. version, : statistical methods for the analysis and presentation of the results of bone marrow transplants. part i: unadjusted analysis statistical methods for the analysis and presentation of the results of bone marrow transplants. part : regression modeling lower respiratory tract respiratory virus infections increase the risk of invasive aspergillosis after a reduced-intensity allogeneic hematopoietic sct immunity and pathophysiology of respiratory tract infections the challenge of respiratory virus infections in hematopoietic cell transplant recipients analysis of engraftment, graft-versus-host disease, and immune recovery following unrelated donor cord blood transplantation graft-versus-host disease following allogeneic transplantation from hla-identical sibling with antithymocyte globulin-based reduced-intensity preparative regimen distinct cd + t cell responses to live and heat-inactivated aspergillus fumigatus conidia immune responses to aspergillus fumigatus infections generation of highly purified and functionally active human th cells against aspergillus fumigatus invasive aspergillosis before allogeneic hematopoietic stem cell transplantation: -year experience at a single transplant center key: cord- - nx f ss authors: paulsen, grant c.; danziger-isakov, lara title: respiratory viral infections in solid organ and hematopoietic stem cell transplantation date: - - journal: clinics in chest medicine doi: . /j.ccm. . . sha: doc_id: cord_uid: nx f ss respiratory viruses are common in solid organ and hematopoietic stem cell transplant recipients and a recognized cause of significant morbidity and mortality. epidemiology, risk factors, and attributable mortality in both populations are reviewed. in addition, virus-specific prevention and treatment options, including emerging investigational therapies, are discussed for respiratory syncytial virus, influenza, adenovirus, parainfluenza, and other respiratory viruses. common respiratory viral infections (rvis) are an important cause of morbidity and mortality following solid organ transplant (sot) and hematopoietic stem cell transplant (hsct). , rvis are typically caused by respiratory syncytial virus (rsv), influenza, parainfluenza, rhinovirus, adenovirus, and human metapneumovirus (hmpv). there is also increasing recognition of human coronavirus and human bocavirus in these populations. in addition, in sot and hsct patients, respiratory infections can be caused by viruses less commonly associated with the respiratory tract, such as cytomegalovirus (cmv), human herpesviruses (herpes simplex virus [hsv] , hsv ), and varicella zoster virus (vzv). this article focuses on the epidemiology, outcomes, and specific prevention and treatment options for rvis in sot and hsct patients. rvis are a well-recognized cause of morbidity and mortality following sot, especially within the thoracic transplant population. recent prospective surveillance of lung transplant recipients in spain found an overall rate of respiratory viruses, asymptomatic and symptomatic, of . per patient-year and a significantly higher rate of . rvis per patient-year in symptomatic patients. nasopharyngeal swabs collected from asymptomatic patients were positive . % of the time compared with . % positive in symptomatic patients. the most frequently detected rvis in symptomatic patients were picornaviruses, such as rhinovirus and enterovirus, at %, followed by coronavirus ( . %) and influenza ( . %). symptomatic rvi detection progressed to lower respiratory tract infection (lrti) in % of patients. a prospective swiss study reported similar results with an rvi incidence of . per patient-year, detection of respiratory viruses in % of those screened and % of symptomatic patients, and rhinovirus/enterovirus as the most common rvi. human bocavirus, identified in , has been reported to cause upper respiratory infection (uri), fevers, wheezing, lrti, and diarrhea in normal hosts, and plays an unclear role in sot recipients. parainfluenza, rsv, hmpv, and influenza were the most frequently found viruses in lrti and were associated with higher rates of hospitalization. analyses of rvi in lung transplant recipients have reported a rate of infection from . % to %, with detection times more frequent if symptoms were present (table ) . , reported risks for rvi in lung transplant patients include increased calcineurin inhibitor levels, age less than years, and underlying cause for transplant other than cystic fibrosis. , as noted for sot patients, rvis in hsct recipients have been well-characterized causes of significant morbidity and mortality. reported incidences of rvi in hsct recipients vary between % in earlier reports using antigen detection and culture and % to % using polymerase chain reaction (pcr) testing (see table ). , - bocavirus, as mentioned earlier, also plays an unclear role in the hsct population, with report of disseminated bocavirus in a pediatric hsct recipient. risk factors for progression to lrti include age greater than years, lymphopenia, neutropenia, alternative/nongenoidentical sibling donor, and chronic graft-versus-host disease (gvhd). , several publications have attempted to delineate morbidity and mortality following rvi in thoracic transplant recipients, specifically with respect to acute rejection and chronic lung allograft dysfunction (clad)/bronchiolitis obliterans syndrome (bos). in the study from spain mentioned earlier, lrti was associated with significant change in lung function (forced expiratory volume in second [fev ]) at and months following infection and nested case-control analysis reported a significant association between rvi within months and acute rejection (hazard ratio [hr], . ; confidence interval [ci], . - . ; p . ). alternatively, the swiss study with similar incidence rates found no such association. clad compromises long-term survival following lung transplant and although an association with previous viral infection has been explored in published literature, a definitive link remains unclear. pooled incidence rates for clad in the meta-analysis mentioned earlier for virus-positive cases were % ( out of cases) compared with . % ( out of ) in virusnegative cases, but because of limited number of overall events a link could not be confirmed. however, there are published epidemiologic links between rvi and clad as well as data on biologically plausible mechanisms underlying a causal relationship. , - in addition, a recent large retrospective cohort (n ) of lung transplant recipients found an independent association between rvi and development of clad within the next months (hr, . ; ci, . - . ; p<. ). paulsen & danziger-isakov published attributable mortality caused by lrti in hsct from a respiratory virus varies, up to % to % in some reports, most commonly involving influenza, rsv, adenovirus, and hmpv. , important risk factors for mortality in the hsct population include lymphopenia, corticosteroids (> mg/kg), and viral or bacterial coinfection. bos following allogeneic stem cell transplant is also associated with significant morbidity and mortality. the most recognized risk factor for bos following hsct is gvhd, but there is some evidence that rvis may play a role as well. , the array of diagnostic tools for rvis in immunocompromised patients has greatly increased over the last few years, providing increased sensitivity as well as decreased processing times. in general, testing for rvis has moved away from viral antigen and culture methods and now relies heavily on molecular methods. real-time quantitative and qualitative reverse transcription pcr testing is a well-established method for identifying viral infection and is now incorporated into many of the guidelines in use. multiplex platforms that test for multiple viruses simultaneously from a single sample are now common. as the sensitivity of molecular diagnostic tools has improved, the issues of asymptomatic viral shedding and sampling methods have become more relevant than ever. as noted earlier, asymptomatic viral shedding is seen frequently, including reports of asymptomatic rsv shedding in hsct recipients for to days and persistent rhinovirus detection for to months in lung transplant recipients. , , , the ability of current molecular testing methods to detect virus in samples with few viral copies combined with prolonged viral shedding in sot and hsct recipients can create a challenge for clinicians trying to determine whether a positive molecular test indicates a true pathogen. the source or sampling method used is an important consideration. respiratory samples are routinely obtained from aspirates/washes or swabs from the nasopharynx (np) or oropharynx (op) as well as more invasive collection methods such as bronchoalveolar lavage (bal). however, the choice of upper respiratory sample collection methodology for optimal viral recovery is uncertain. in children hospitalized because of respiratory infections, sensitivity of rvi detection with np aspirates of % to % have been reported versus nasal swab sensitivity of % to %. a similar comparison of np swabs with or without op swabs in immunocompetent children found a higher sensitivity of np swabs for rvi of % to %, which was greater than or equal to op swabs ( %- %), with combined testing increasing detection by % to %, depending on the virus. investigation in adults reported higher sensitivity with np washes ( %) than np swabs ( %) or op swabs ( %), and noted that maximal sensitivity was achieved through a combination of all methods. rvi prevention is a key component to minimize infections and subsequent complications. interventions can be patient specific, such as antiviral prophylaxis following transplant and immunization both before and following transplant. for the most common rvis, immunization is only available for influenza, as discussed later. vaccines for many of the other rvis are still under development and not commercially available. in addition, interventions at the level of the health care system decrease the incidence of rvi, including the appropriate use of respiratory or contact precautions, screening of visitors, and immunization of health care staff. , these interventions are included in the general us centers for disease control and prevention (cdc) recommendations to reduce health care-associated pneumonia. prevention of rvi in hsct recipients is also addressed in the joint cdc, infectious diseases society of american (idsa), and american society for blood and marrow transplantation guidelines as well (http://asheducationbook.hematologylibrary. org/content/ / / .long). other interventions, such as universal mask use for all health care staff and visitors on an hsct unit, strict isolation of all patients, mandatory hand washing, and visitor restriction for children less than years of age, have been shown to significantly reduce the incidence of rvis and are used at some centers. [ ] [ ] [ ] virus-specific outcomes, prevention, and treatment respiratory syncytial virus epidemiology, risk, and attributable mortality rsv has long been recognized as a concerning pathogen in immunocompromised hosts, with increased mortality if the infection involves the lower respiratory tract. in the solid organ population, lung transplant recipients are at increased risk for rsv-related mortality and morbidity compared with the other organs. , incidence of rsv in lung transplant recipients is variable and accounts for roughly % to % of rvi infections. risk factors for morbidity and mortality are not clearly defined, with reports of young age (< years), recent transplant, preexisting lung disorder, recent rejection, and multivisceral transplant as risk factors. , rsv infection following hsct depends on several factors, including patient age and type of transplant. incidence of rsv infection following hsct has been reported at % to % for allogeneic and . % for autologous, with pediatric patients at the greatest risk of rsv. , several risk factors have been reported for rsv as well as progression to lrti or severe infection. in hsct, risk factors for rsv include less than month posttransplant (preengraftment), both younger (< years) and older (> years) age, gvhd, relapsed disease, and smoking. , risks for presenting with lrti or progression to lrti include lack of rsv-directed therapy, high-dose total body irradiation, respiratory coinfection, absolute neutrophil count less than cells/mm , and an absolute lymphocyte count (alc) less than to cells/mm . , in contrast, an alc greater than cells/mm was protective against progression to lrti. attributable mortality in rsv infection has been reported in several publications; with mortality approaching % in untreated hsct patients and decreased to % to % with prompt supportive care and/or treatment. more recent assessment of adult hospitalized patients reported % to % mortality in hsct and % to % in sot recipients in the setting of ribavirin treatment. , , age greater than years and lymphopenia were risk factors for mortality and compared with nontransplant patients. a recent report in pediatric hsct recipients also showed better outcomes than historical reports, with only % progression to lrti and no mortality, with a primary treatment modality of intravenous immunoglobulin (ivig) supplementation. although transplant patients are likely at increased risk for morbidity and mortality, especially those with lrti, pediatric patients presenting in clinic with upper respiratory tract infection (urti) symptoms have also been managed with good outcomes in the outpatient setting. therefore, although rsv infection in sot and hsct populations is associated with increased morbidity and mortality, precise determination of its impact remains elusive, but overall outcomes seem to be improving over time. in addition to the general preventive measures reviewed earlier, the only us food and drug administration (fda)-approved agent for the prevention of severe rsv infection is palivizumab. palivizumab is a humanized monoclonal antibody targeting the f glycoprotein of rsv, and is approved for prevention of rsv in high-risk patients less than years of age. recommendations were published in by the american academy of pediatrics for palivizumab prophylaxis for infants and children less than months of age with specific predisposing conditions. with respect to immunocompromised children, no specific recommendations were made, aside from considering prophylaxis in profoundly immunocompromised patients younger than months of age. a more recent multidisciplinary consensus conference in italy, referencing the lack of adequate clinical trials and statistical power, recommended against palivizumab for children with primary or acquired immunodeficiencies. in practice, the use of palivizumab for rsv prophylaxis in immunocompromised patients varies widely. approximately % of surveyed pediatric sot centers used palivizumab for prophylaxis, in both candidates and recipients, with % of those targeting infants to months old and % extending use to to months. only % provided palivizumab for patients to years old and % gave palivizumab for patients more than years of age. this variability in practice is likely caused by the limited data on the efficacy of palivizumab prophylaxis in immunocompromised patients. in a retrospective pediatric hsct cohort from memorial sloan kettering cancer center (n ), nearly half of the high-risk patients received intravenous (iv) palivizumab. in the palivizumab treatment group, % developed rsv compared with only % of those who did not receive prophylaxis. similarly, an approach limiting palivizumab only to those hsct pediatric patients younger than months with either a chronic oxygen requirement or severe combined immunodeficiency, pretransplant to days posttransplant, found no increase in the incidence of rsv or patient outcomes compared with historical controls with wider use of palivizumab. if time allows, delaying hsct has also been reported to be an effective strategy to prevent serious rsv infection in hsct candidates with rsv urti before conditioning. , one reported strategy delayed transplant until symptom resolution and negative repeat rsv testing, resulting in a significant reduction in rsv pneumonia following transplant and improved mortality compared with those patients whose transplants were not delayed. careful thought must be given to the underlying disease process, risk of progression, as well as type of transplant before making the decision to delay hsct. several management options for rsv have been considered and reported in the literature, and although there are reports of improved outcomes, no placebo-controlled trial has clearly delineated the indication for and efficacy of treatment. the only randomized controlled trial accrued hsct patients over years and reported that aerosolized ribavirin decreased rsv viral load compared with supportive care but did not significantly improve outcomes. ribavirin is a broadspectrum nucleoside analogue with activity against dna and rna viruses. reported toxicities of inhaled ribavirin include bronchospasm, cough, nausea, rash, and decreased pulmonary function. iv ribavirin adverse effects include hemolysis, hyperbilirubinemia, and leukopenia, whereas oral ribavirin can cause anemia and rash. at present, aerosolized ribavirin remains the only fdaapproved drug for the treatment of severe rsv infection, and it is only approved for use in children. immunomodulators that have also been investigated include ivig and anti-rsv monoclonal antibody (palivizumab). because of the lack of clear evidence of efficacy, wide variation in management of rsv exists. recommendations and guidelines have been published for hsct and sot patients, and although based on the best available data, they are not strong recommendations in many cases. for example, the sot recommendations for rsv lrti are for consideration of aerosolized ribavirin in combination with rsv ivig or palivizumab. because of the increased mortality in hsct patients, recommendations for treatment within this population are stronger. based on published reports as well as selfreported treatment strategies in surveys from sot centers, lung and heart-lung recipients are often treated for urti or lrti with rsv; lrtis may be treated in non-lung transplant sot recipients, although this is inconsistent. retrospective and prospective studies report improved outcomes in symptomatic lung transplant patients treated with iv ribavirin plus corticosteroids ; oral ribavirin plus corticosteroids ; oral or iv ribavirin , ; and inhaled ribavirin plus corticosteroids, ivig, and palivizumab, highlighting the lack of consensus on treatment strategies in this population. recommendations for hsct patients from the infectious diseases working party of the german society for haematology and medical oncology recommend ivig in general and ribavirin in particular for rsv in patients with cancer, largely based on data in hsct recipients. guidelines from the united kingdom recommend inhaled ribavirin and ivig for allogeneic hsct recipients with either lrti or urti and risk factors for progression to lrti; they also suggest oral ribavirin if the inhaled form is not available. outside of guidelines and recommendations, several prospective and retrospective studies have been published on the treatment of rsv in hsct patients, and, despite the available literature, there is no commonly accepted approach. reports of improved outcomes in the treatment of rsv in hsct patients can be found for inhaled, oral, or iv ribavirin, , , as well as combinations of ribavirin with ivig, palivizumab, and/or rsv-specific ivig ; most support treatment at the urti stage, before progression to lrti. a systematic review of the available retrospective studies in reported that any form of ribavirin, alone or in combination with an immunomodulatory agent, was effective in preventing progression of urti to lrti, with a trend toward better outcomes with inhaled ribavirin plus an immunomodulator. although negative studies are potentially less likely to achieve publication, there are data available suggesting that adjunctive corticosteroid use and palivizumab alone do not improve outcomes. , complicating the assessment of rsv treatments in sot and hsct patients are recent reports of good outcomes with minimal intervention, such as the immunocompromised pediatric patients diagnosed with symptomatic rsv without any mortality despite only ( %) receiving directed therapy. another analysis of pediatric hsct recipients with rsv reported no attributable mortality with no ribavirin therapy; all patients were managed either with supportive care alone or immunoglobulin therapy. although it is unclear which, if any, rsv-specific treatment is the most effective intervention, there are potentially effective investigational drugs being developed. treatment with aerosolized aln-rsv (alnylam pharmaceuticals, cambridge, ma), a small interfering rna that targets the rsv nucleocapsid messenger rna, has shown some early promise in potentially preventing bos in lung transplant recipients with rsv. another agent, ri- (adma biologics, inc, ramsey, nj) contains standardized levels of high-titer anti-rsv neutralizing antibody. in a report of compassionate use in patients with rsv lrti already receiving treatment with ribavirin, ri- was well tolerated and showed at least a -fold increase in geometric mean titer of rsv antibodies. ri- (adma biologics, inc, ramsey, nj) is a new immunoglobulin formulation that was developed using plasma collected from individuals tested to have high-titer anti-rsv antibodies, and in early trials showed a significant increase in anti-rsv neutralizing antibodies when administered to primary immunodeficient patients. in addition, there are several other small molecule therapies in various stages of development, including early clinical trials. one such molecule, gs- , an oral rsv entry inhibitor, was reported to have significantly lower viral load, total mucus weight, and total symptom score versus placebo in a healthy adult challenge. phase b trials with the same novel small molecule in lung transplant recipients and bone marrow transplant (bmt) patients have completed enrollment, with results pending (nct , nct ). epidemiology, risk, and attributable mortality the impact of influenza infection in sot patients can be particularly severe, especially in lung transplant recipients. rates of severe influenza in lung transplant patients have been reported in % to % of those infected, with an attributable mortality of % to %; even higher mortalities of % caused by the h n infection in lung transplant recipients with preexisting bos grade were reproted. [ ] [ ] [ ] a report of australian lung transplant recipients with h n influenza noted a % fev decline at presentation and % of patients had prolonged allograft dysfunction. reports of the most severe infections are largely based on outcomes following the h n pandemic; however, a recent retrospective cohort study of brazilian renal transplant recipients with influenza a between and reported a % incidence of both intensive care unit (icu) admission and mortality, which is higher than expected. similar to solid organ recipients, influenza infection in hsct patients causes significant morbidity and mortality. progression from urti to lrti varies widely depending on the report, but has been found in % to % of adult hsct patients or patients with hematologic malignancy, with overall mortality of % to %. , , risk factors for either presentation with lrti or progression to lrti include an alc less than to cells/ml, lack of influenza-directed therapy, increased creatinine level, and delay in seeking care. there is a significant survival benefit if treated with influenza-directed therapy, with % mortality in treated patients versus % mortality in untreated patients. , , prevention vaccination remains the primary focus and most strongly recommended method of influenza prevention. seasonal influenza vaccines cover either or strains of influenza based on antigenic characterization of the previous year's circulating strains. in general, quadrivalent vaccines cover an additional influenza b strain compared with the trivalent vaccines without interfering with vaccine response. influenza vaccines are available in inactivated (intramuscular or intradermal administration) and live, attenuated (intranasal) formulations. the live, attenuated vaccine is not recommended for immunocompromised recipients, and the inactivated vaccine is preferred for household contacts. , overall influenza vaccine response in sot and hsct patients remains variable based on the population. the optimal timing of vaccination following sot has not been precisely determined, with guidelines from the american society of transplantation infectious diseases community of practice and the idsa recommending vaccination between and months after transplant. , reports of immunogenic response to influenza vaccine in sot show a wide variation depending on the organ transplanted and the year assessed, with historical ranges between % and % protection. seroprotective responses have been reported between % and % in lung transplant recipients, [ ] [ ] [ ] % and % in adult renal transplant recipients, and greater than % in pediatric liver recipients. various adjuvants and dosing strategies have been evaluated in an effort to increase immunogenicity, with variable results. high-dose influenza vaccine contains times the antigen dose of the standard influenza vaccine, and in early trials has been reported to increase the percentage of pediatric sot patients who achieved an increase in protective titers. there are several likely explanations behind the reported variability in response, including the organ transplanted, duration of time from transplant, and degree of immunosuppression. the impact of immunosuppression on immunogenicity shows mycophenolate mofetil as most consistently associated with a decrease in response, whereas sirolimus has been associated with an increase in vaccine responsiveness. , because of concerns about the potential development of human leukocyte antigen alloantibodies following the adjuvanted pandemic h n influenza vaccine, evaluations of acute rejection following influenza immunization have been conducted. [ ] [ ] [ ] a subsequent postauthorization safety study following the / ph n vaccine found no increased risk of acute rejection associated with vaccination. further, seasonal influenza immunization from to also found no increased risk of acute rejection. similar to sot patients, influenza immunization is recommended for hsct recipients and is less effective compared with healthy controls. again, response rates to influenza in hsct vary based on time from transplant, influenza season/year studied, and degree of immunosuppression. rates of seroconversion of % to % are typical. , in addition, there are encouraging reports of increased immunogenicity with high-dose inactivated influenza vaccine compared with standard dose. although inactivated influenza vaccination is strongly recommended for sot and hsct recipients, other approaches have been suggested to prevent infection. in addition to use for treatment of influenza, oseltamivir and zanamivir are also approved for influenza chemoprophylaxis. a prospective study, predominantly in adult sot recipients, found that daily oseltamivir for weeks during seasonal influenza circulation significantly reduced the incidence of laboratory-confirmed influenza as determined by reverse transcription pcr. oseltamivir, given for to days, has also been reported to be effective in prevention of influenza infection in a hematology/hsct inpatient unit during a nosocomial h n outbreak. treatment early treatment with antiviral drugs has been shown to improve outcomes and reduce hospital admissions and mechanical ventilation use. , , , the mainstay of treatment of influenza are the neuraminidase inhibitors (nais), oseltamivir, peramivir, and zanamivir. the influenza a m protein inhibitors amantadine and rimantadine are active against influenza a only, but are no longer recommended because of significant resistance in circulating influenza strains. literature on the treatment of influenza in sot and hsct are reliably reproducible with maximum benefit seen the earlier in the course that virus-specific therapy is initiated. during the pandemic h n season, sot recipients treated with antiviral agents within hours of symptom onset were significantly less likely ( % vs %) to require icu admission. although treatment within to hours is optimal, benefit has been shown even with delayed treatment. most experts therefore endorse influenza-specific antiviral treatment of sot and hsct patients with influenza at any point in their illness. oseltamivir is an oral nai indicated for the treatment of influenza a and b in adults and children more than weeks of age. recommended duration of treatment is days in immunocompetent children and adults. in general, treatment recommendations and practice in sot patients with influenza are for days of oseltamivir as well. , although days of oseltamivir is the typical treatment duration, there are reports of treatment for days or longer in patients with persistent symptoms. a clinical trial also investigated conventional-dose versus doubledose oseltamivir for days in immunocompromised patients, with no results yet released (clinicaltrials.gov, nct ). zanamivir is an inhaled nai approved for treatment of influenza a and b in adults and children years of age and older. zanamivir is used less frequently than oral oseltamivir, likely because of the delivery route and rare reports of inhaled zanamivir failure. peramivir is active against influenza a and b and currently is the only iv nai approved for clinical use in patients aged years and older. iv formulations of both zanamivir and oseltamivir are under investigation in clinical trials. , there is generally less experience with peramivir compared with oral oseltamivir, but published reports of clinical effectiveness and reduction in viral load are encouraging. peramivir is a viable treatment option, especially in those patients in whom oral or inhaled antivirals are not the optimal route. , the pandemic h n influenza strain was also notable for an increased frequency of nai resistance in up to % of strains. however, nai resistance is currently uncommon, with an overall incidence of . % to . % of isolates, but does remain an area of growing concern. the most frequent neuraminidase mutation is the h y substitution, which results in highlevel oseltamivir resistance, reduced peramivir susceptibility, and generally preserved zanamivir activity. , options for oseltamivir-resistant influenza are limited. inhaled zanamivir may have activity in many, but not all, cases. peramivir has been reported to be effective in patients with oseltamivir-resistant influenza and has shown encouraging results in preventing lethality in mouse models of nai-resistant h n /avian influenza models. , das (ansun biopharma, san diego, ca) is a recombinant fusion protein that removes the sialic acid receptor for influenza binding and entry into the cell, potentially inhibiting influenza and parainfluenza infection. das has shown promising in vitro results of activity against oseltamivir-resistant influenza strains, and additional testing versus several nai-resistant strains is ongoing. , aside from advances in supportive care, no specific adjunctive therapies are routinely recommended. corticosteroids have been shown to decrease the need for mechanical ventilation and progression to lrti but at the cost of prolonged viral shedding. therefore, although corticosteroids are not routinely recommended, if corticosteroids are indicated for another reason, such as rejection or gvhd, worsening infection with influenza is not clear reason to withhold steroids. epidemiology, risk, and attributable mortality adenovirus is a double-stranded dna virus made up of immunologically distinct types, with serotypes and most commonly associated with pneumonia. in nonimmunocompromised patients, adenovirus typically causes self-limited disease, such as uri, conjunctivitis, and/or gastroenteritis. adenovirus infection in immunocompromised patients can range from asymptomatic viremia to significant localized or disseminated disease. in contrast with many of the other community-acquired respiratory viruses, adenoviral infection can occur from primary acquisition or through reactivation of virus. in study, viral reactivation was linked to the timing of immune reconstitution and cd t-cell counts. depending on patient factors, clinical disease in immunocompromised patients can include pneumonia, hepatitis, colitis, hemorrhagic cystitis, and encephalitis. in a to retrospective cohort of rvis in pediatric sot, hsct, and oncology patients, adenoviral infection was associated with the greatest length of stay, and was the only specific virus that increased the risk of morbidity and mortality related to rvi (odds ratio, . ; ci, . - . ; p . ). adenoviral infections have been reported in renal, liver, small bowel, lung, and heart transplant recipients, and although most are asymptomatic, they can be associated with severe disease. [ ] [ ] [ ] [ ] [ ] [ ] although adenoviral infection in hsct is more likely to cause severe disease than in sot, there are reports of graft infection and rejection with adenovirus in essentially all sot populations. , , infection in lung recipients is common (up to % in series) and there are multiple reports of severe infection, including graft loss, progression to bos, and death. , [ ] [ ] [ ] [ ] infection in hsct recipients can be severe and associated with significant mortality. in adult hsct recipients with adenovirus viremia, most of whom were treated with antivirals, almost % developed invasive adenoviral disease and % died of the infection. adenoviral lrti and disseminated disease generally carry the greatest mortality in pediatric and hsct patients (up to % in some reports). [ ] [ ] [ ] treatment treatment options for adenoviral infection or disease are limited because there are currently no approved antiviral agents for treatment. reports of recovery with reduced immune suppression alone make the need for therapy and optimal timing for intervention uncertain. as for most viral infections in sot and hsct patients, immunosuppression reduction is recommended if possible. cidofovir, approved for treatment of cmv retinitis in patients with acquired immunodeficiency syndrome, has been the most common antiviral used for treatment. cidofovir is a nucleotide analogue that inhibits viral dna polymerase with broad antiviral activity against dna viruses, such as herpesviruses and adenovirus. although cidofovir has in vitro activity against adenovirus and is generally accepted as the standard of care, cidofovir treatment efficacy is controversial. , , dosing of cidofovir is generally mg/kg iv once every days, or mg/kg iv times per week, often in conjunction with probenecid and hydration. cidofovir nephrotoxicity, which is not dose dependent, is the most common reason for discontinuation of therapy. alternative therapies, including ribavirin and combination cidofovir plus ivig, have been reported with limited data, and these are not routinely recommended for use. , , investigational adenovirus therapies because of new antiviral development and alternative treatment modalities, there may be additional options in the future. brincidofovir is an orally bioavailable lipid conjugate of cidofovir that has potent in vitro activity against adenovirus, and has shown promising results for both treatment of serious invasive adenovirus infections and asymptomatic viremia. , recognizing the vital role of t-cell immunity in control of viral infections and the loss of this immunity during hsct and sot, the use of adoptive t-cell immunity is promising as well. adoptive t-cell immunity uses donor virus-specific t-cells to treat infection, and has been reported to be safe and effective when performed early in the course of the infection. , adoptive t-cell transfer has generally been limited to a few centers and predominantly in hsct recipients, because of both the time and the expertise needed for cell preparation, but more recent methods may allow shorter generation time and more access to this therapy. , parainfluenza epidemiology, risk factors, and attributable mortality parainfluenza virus (piv) is a single-stranded, enveloped rna virus with distinct serotypes (types - ). serotypes vary in seasonality and disease, with piv associated with pneumonia and bronchiolitis, and year-round activity that peaks in spring and summer. piv and piv are commons causes of pediatric laryngotracheobronchitis (croup) and typically peak in fall and winter. piv infection is rarely associated with disease. a retrospective analysis of rvi in pediatric immunocompromised patients reported that % of rvis in sot patients were caused by piv. parainfluenza infections presented more commonly as urti with or without lrti symptoms, and less frequently as lrti alone. although there are previous reports of significant mortality in sot recipients caused by piv (up to % in series ), more recent publications in lung and other organ recipients report decreased mortality. , retrospective reports place the incidence of piv infection following hsct between % and %, with greater incidence in children than in adults. [ ] [ ] [ ] lrti after hsct is associated with high morbidity and mortality. risk factors for lrti caused by piv are lymphopenia (< cells/mm ), neutropenia (< cells/ml), apache (acute physiology and chronic health evaluation) ii score greater than , myeloablative conditioning, high-dose corticosteroids for gvhd, and coinfection. , reported risk factors for mortality in hsct include lrti, early infection, mismatched related donor, apache ii score greater than , new oxygen requirement at diagnosis, low monocyte counts (< cell/ml), and high-dose steroid use (> mg/kg/d). , , in a series of hsct patients with piv lrti, mechanical ventilation was necessary in % and attributable mortality was %. other recent publications report % to % mortality in hsct recipients with probable or proven parainfluenza lrti. , , treatment there are no currently fda-approved antiviral treatments for parainfluenza disease. treatment is supportive and includes reduction in immunosuppression. ribavirin and/or ivig have been used, off label, in parainfluenza infections with variable results and no definitive evidence of efficacy. , [ ] [ ] [ ] [ ] [ ] [ ] [ ] investigational parainfluenza virus therapies das , as discussed previously for nai-resistant influenza, can potentially inhibit piv binding to respiratory epithelial cells. das is an inhaled treatment typically administered via a dry powder inhaler for to days, and has been used under compassionate use and clinical trial protocols in hsct and sot recipients, including lung transplant patients. in published reports, das has shown encouraging results, including reduction in piv quantitative viral load and overall outcomes. [ ] [ ] [ ] [ ] in addition, there are intriguing studies examining the impact of cholesterol reducing agents such as gemfibrozil and lovastatin on disrupting viral assembly in piv, rsv, and influenza. reports of parainfluenza-specific t-cell generation from healthy donors may also ultimately lead to effective adoptive t-cell therapy for piv. epidemiology, risk factors, and attributable mortality human metapneumovirus (hmpv) is an rna virus identified in , in the same paramyxovirus family as rsv and parainfluenza, that typically causes a self-limited urti in immunocompetent persons. hmpv is found worldwide and occurs predominantly in the late winter and spring months, often following the rsv season. as with many of the other community-acquired rvis in sot and hsct patients, progression from the upper respiratory tract to the lower respiratory tract is associated with increased morbidity and mortality. hmpv has been a well-documented cause of symptomatic and asymptomatic infection in lung transplant recipients. in a retrospective population of symptomatic adult lung transplant recipients, % were hmpv positive via nasopharyngeal aspirate or bal. as with most rvis, up to % can be asymptomatic, whereas others develop severe pneumonia or acute graft dysfunction. rhinorrhea, cough, and sputum production are the most frequently reported symptoms, with mortality caused by acute respiratory distress syndrome and graft rejection also reported. , identification of replicating hmpv in respiratory samples has been seen with simultaneous biopsy-proven graft rejection, suggesting a potential association between hmpv and acute rejection. infection in hsct patients is variable as well, with recent reviews reporting an overall incidence of % to %. , the same analysis found that lrti occurred in % of hmpv infections in hsct recipients, with a mortality of %, greatest in those with lrti. mortalities caused by hmpv infection have been reported to be as high as %. progression from urti to lrti is seen in up to % of hsct recipients and has been associated with steroid use (> mg/kg), low lymphocyte count (< cells/mm ), and onset of infection less than days from hsct. there are no currently approved antivirals for the prevention or treatment of hmpv. as with rsv, ribavirin has reported in vitro activity against hmpv. the efficacy of ribavirin for hmpv infection has not been reliably shown. there are reports in lung transplant recipients of oral ribavirin resulting in quicker return to baseline and decreased incidence of subsequent bos, and case reports of survival with iv and inhaled ribavirin. , however, most reports are from small single-centered studies and do not include a control population. in addition, other studies have not been able identify a similar beneficial effect of ribavirin for hmpv treatment. , investigational human metapneumovirus therapies adoptive t-cell transfer has not yet been achieved, but hmpv-specific t cells have been generated, with further work ongoing. there are also reports in vitro and in mouse models of a human monoclonal antibody for prophylactic and therapeutic hmpv infections. cytomegalovirus epidemiology, risk factors, and attributable mortality cytomegalovirus (cmv) is a b-herpesvirus well recognized as a significant pathogen in immunocompromised patients. cmv has been reported to affect from % to % of heart and lung transplant recipients and % of hsct patients. [ ] [ ] [ ] although cmv can cause a wide array of infections, from asymptomatic to tissue-invasive disease, cmv pneumonitis/pneumonia is of particular concern for thoracic transplant recipients. the diagnosis of proven cmv pneumonitis is based on compatible clinical signs and/or symptoms and documented cmv in lung tissue. traditionally, tissue-invasive cmv is based on histopathologic or immunohistochemical (ihc) findings consistent with tissue invasion on biopsy. , cmv cultures or quantitative nucleic acid amplification testing of tissue samples are difficult to interpret because a positive finding could indicate tissue-invasive disease, shedding in the setting of active viremia, or both. recent updates to the definition of cmv pneumonia now include proven, probable, and possible cmv pneumonia. proven disease still relies on identification of viral antigens or inclusion bodies via immunohistochemistry in biopsy material, but probable cmv pneumonia is defined as compatible symptoms plus cmv detection via viral isolation, culture, or quantitation of cmv dna in bal fluid. there is no definitive cutoff for cmv dna load in the setting of cmv pneumonia; however, some reports suggest greater than to iu/ml as a possible cutoff. , possible cmv pneumonia has been suggested based on positive quantitative pcr performed on biopsy tissue. risk factors for cmv disease have been reported as advanced age and reduced-dose valganciclovir. delayed-onset cmv pneumonitis (> days posttransplant), donor with positive cmv serology, asymptomatic cmv infection, and cmv disease at any time have been associated with increased mortality in lung transplant recipients. [ ] [ ] [ ] cmv in hsct patients can also cause a wide array of clinical manifestations, with cmv pneumonia being the most serious, resulting in a mortality of approximately %. , cmv reactivation alone is associated with lower overall mortality following hsct. incidence of cmv pneumonia is unclear, largely because of the difficulties with definitive diagnosis, but an autopsy study of patients with cancer and hsct reported an incidence of cmv pneumonia of %. risk factors associated with cmvattributable mortality include female sex, lymphopenia, and mechanical ventilation at onset. , diagnostic classification of cmv pneumonia/ pneumonitis is the same as discussed earlier for sot. iv ganciclovir (gcv) and oral valganciclovir, with or without cmv immunoglobulin (cmvig), are the agents recommended for prophylaxis of cmv infection in sot recipients. the most common strategies used are universal prophylaxis versus preemptive therapy, with consensus recommendations favoring universal prophylaxis for highrisk heart and lung recipients. risk stratification in organ transplant recipients relies heavily on donor and recipient cmv serologic results before transplant. donors who are cmv immunoglobulin (ig) g positive (d ) paired with recipients who are igg negative (rÀ) are considered the highest risk group. recipients who are cmv dÀ/rÀ are generally considered low risk, although community-acquired infection can occur posttransplant, and r patients are variably classified as intermediate-risk or high-risk, often center dependent. recommended duration of prophylaxis varies based on organ system and risk stratification. for d /rÀ heart transplant recipients, the recommended minimum duration of cmv prophylaxis is between and months. for d /rÀ lung transplant recipients the minimum recommended duration is between and months, with some advocates for longer, even indefinite, prophylaxis. centers using indefinite prophylaxis for lung transplant recipients report low incidence of cmv infection, which must be balanced against reports of association between gcv-resistant cmv and prolonged cmv prophylaxis, with an incidence of % to % gcv-resistant infection in some cases. , , for r recipients the minimum duration in lung is months, and months in heart recipients. in dÀ/rÀ populations the routine use of cmv prophylaxis is not generally recommended. there are several published reports of potential benefit to the addition or sole use of cmvig for prophylaxis, but, because of the limited data to support routine use, the addition of cmvig is not routinely recommended. , the most recent consensus guidelines note that some experts add cmvig for intermediate and higher-risk recipients, but there are no randomized studies indicating that cmvig is any better than gcv or valganciclovir alone. as opposed to sot recipients, hsct patients with positive cmv serology (r ) before transplant are at higher risk for reactivation and non-relapserelated mortality. preventive strategies following hsct are similar and generally consist of either universal prophylaxis for at least days following hsct or a preemptive approach, with the latter as the more commonly reported practice. , there are also reports that suggest immunosuppression choice alters cmv infection risk. lower incidences of cmv infection are reported in patients treated with regimens including a mammalian target of rapamycin (mtor) inhibitor. [ ] [ ] [ ] [ ] although these results are not consistently reproducible, some experts recommend considering the use of mtor inhibitors in the presence of clinically relevant, recurrent, or gcv-resistant cmv infection. [ ] [ ] [ ] further, cmv vaccination is under evaluation with several candidate vaccines assessed in clinical trials, including live attenuated; recombinant glycoprotein b (gb); dna plasmid; and virus-like particle systems. investigations in immunocompromised patients have reported on the safety of a cmv dna vaccine candidate (asp ) and a chimeric peptide vaccine (pf ) in hsct, with some early evidence of potential clinical benefit as well. , similarly, a phase trial of an adjuvanted gb vaccine versus placebo in pretransplant liver and kidney patients showed reduced cmv viremia and days of gcv therapy following transplant, with the greatest effect seen in cmv seronegative patients. therefore, although the current cmv vaccine candidates are not yet ready for clinical use, the available data plus the number of clinical trials either planned or ongoing in immunocompromised patients are very encouraging. iv gcv and oral valganciclovir are the most commonly used treatments for cmv infection or disease. , foscarnet and cidofovir are active agents as well, but are generally reserved for gcv-resistant infection or those hsct recipients with concern for potential bone marrow suppression caused by of gcv, such as during the preengraftment period. , cmvig has been reported to be effective in several publications and may have a role in certain settings. , cmvig is often used for cmv pneumonia in hsct recipients, although a recent analysis failed to find significant improvement with adjunctive ivig or cmvig administration in that population. the consensus guidelines on cmv treatment in sot recipients recommend consideration of adjunctive ivig or cmvig for recurrent cmv disease in thoracic organ recipients as adjunctive therapy in cases of hypogammaglobulinemia. investigational cytomegalovirus therapies investigational drugs and therapies for cmv prevention and treatment are currently being developed and/or tested in clinical trials. letermovir interacts with ul , a component of viral dna cleavage and packaging, and has been reported to be effective in reducing cmv infection in hsct recipients and reducing viral load in sot patients. , maribavir is a competitive inhibitor of ul that failed to show noninferiority of cmv prevention in liver transplant patients and hsct recipients, but is currently under investigation for use in cmv infections that are refractory to gcv, valganciclovir, foscarnet, or cidofovir (clinicaltrials.gov, nct ). , brincidofovir, referenced earlier for adenovirus, has been shown to reduce cmv events compared with placebo in hsct recipients. in addition, adoptive t-cell therapy for cmv disease has been reported to be effective in a lung transplant recipient and as well as in several hsct recipients. [ ] [ ] [ ] herpes simplex virus and varicella zoster virus hsv and hsv are a-herpesviruses and common causes of infection in immunocompetent and immunocompromised persons. in immunocompromised patients, hsv can cause a wide variety of clinical infection from asymptomatic oropharyngeal shedding to mucocutaneous and disseminated disease, including pneumonitis. hsv pneumonitis is uncommon in the era of antiviral prophylaxis, but thoracic transplant and hsct recipients are reportedly at greatest risk. [ ] [ ] [ ] similar to cmv pneumonitis, the recommended diagnostic test for hsv pneumonitis is tissue histopathology with ihc for hsv. positive bal pcr testing for hsv may represent contamination or oropharyngeal shedding. iv acyclovir is the treatment of choice for severe, disseminated hsv, including pneumonitis. it is recommended to continue iv acyclovir until resolution, or days, at which time oral medication may be given. vzv, another herpesvirus, can also cause significant disease in sot and hsct recipients. the most common manifestations of vzv are primary varicella, or chickenpox, in susceptible seronegative patients or herpes zoster, or shingles, in those with previous vzv infection or vaccination. visceral dissemination is more common in hsct and less common in sot. disseminated disease includes infection of lung tissue resulting in vzv pneumonia with significant morbidity and mortality. , risk of dissemination is increased in acute and chronic gvhd, as well as increased 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recipients cmv immunoglobulins for the treatment of cmv infections in thoracic transplant recipients letermovir for cytomegalovirus prophylaxis in hematopoietic-cell transplantation preemptive treatment of cytomegalovirus infection in kidney transplant recipients with letermovir: results of a phase a study maribavir prophylaxis for prevention of cytomegalovirus disease in recipients of allogeneic stem-cell transplants: a phase , double-blind, placebo-controlled, randomised trial efficacy and safety of maribavir dosed at mg orally twice daily for the prevention of cytomegalovirus disease in liver transplant recipients: a randomized, double-blind, multicenter controlled trial cmx to prevent cytomegalovirus disease in hematopoieticcell transplantation adoptive t-cell therapy of a lung transplanted patient with severe cmv disease and resistance to antiviral therapy transfer of minimally manipulated cmv-specific t cells from stem cell or third-party donors to treat cmv infection after 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patients with malignancies and transplant recipients are at high risk of infection-related morbidity and mortality. children at the highest risk for infections are those with acute myeloid leukemia (aml), relapsed acute lymphoblastic leukemia (all), and hematopoietic stem cell transplant recipients (hsct). these patients are at high risk for life-threatening bacterial, viral, and fungal infections which are associated with prolonged hospital stay, poor quality of life, and increased healthcare cost and death. recognition of risk factors which predisposes them to infections, early identification of signs and symptoms of infections, prompt diagnosis, and empiric/definitive treatment are the mainstay in reducing infection-related morbidity and mortality. infection control and prevention programs also play a crucial role in preventing hospital-acquired infections in these immunosuppressed hosts. there are various factors which contribute to the increased susceptibility to infections in pediatric hematology/oncology (pho) and hsct patients, most prominent of them being disruption of cutaneous and mucosal barriers (oral, gastrointestinal, etc.), microbial gastrointestinal translocation, defects in cell-mediated immunity, and insufficient quantities and inadequate function of phagocytes. goals of infection control and prevention in this population are based on mitigating the risk inherent with the underlying malignancy and associated treatments (i.e., chemotherapy, radiation). this chapter discusses infection control and prevention measures specifically in patients with hematological malignancies as well as hsct recipients. hand hygiene and standard precautions during the care of pho and hsct patients are key components in reducing the risk of infections. additional isolation precautions may also be undertaken depending on the pathogen isolated and/or symptoms that the patient is experiencing (e.g., contact precautions would be appropriate in patients experiencing diarrhea). further information on general infection prevention measures can be found in chap. . minimizing injury to mucosal surfaces and decreasing heavy colonization of the skin reduce the likelihood of microbial invasion through these sites. thus, the importance of meticulous skin care and daily inspection in pho and hsct patients is paramount and provides opportunities to identify areas of inflammation or breakdown early. skin inspection should be done routinely, with special attention to highrisk areas like intravascular catheter insertion sites and the perineum. rectal thermometers, digital rectal examinations, and suppositories should be avoided to prevent mucosal breakdown. as part of an effort to reduce colonization of cutaneous surfaces, daily chlorhexidine baths have been shown to reduce hais and transmission of multidrug-resistant organisms (mdro) in oncology patients [ , ] . chlorhexidine gluconate (chg) is a cationic bisbiguanide that serves as a topical antiseptic. chg binds to negatively charged bacterial cell wall proteins altering the bacterial cell wall equilibrium and helps in reducing bacterial colonization of the skin [ ] . education of patients, families, and staff on the importance of these practices is key to compliance with this preventative strategy and should be made a priority. many experts recommend a complete periodontal examination be performed prior to initiation of chemotherapy with reevaluations throughout the treatment course and after completion [ , ] . oral mucositis, which can be considered an acute inflammation and/or ulceration of the oral/oropharyngeal mucus membranes, is a common adverse effect of chemotherapeutic agents. it can cause oral pain/discomfort as well as difficulties in eating, swallowing, and speech. mucositis is most commonly caused by chemotherapeutic agents which prevent dna synthesis such as methotrexate, -fluorouracil, and cytarabine, particularly in hsct recipients. oral rinses with normal saline or chg-containing products are recommended - times per day to prevent oral mucositis [ , ] . patients with painful mucositis might not comply with oral care regimens, however, putting them at increased risk for infections from oral flora such as bacteremia due to viridans streptococci. mouth rinses containing alcohol should be avoided because they can aggravate mucositis. neutropenic patients should also be instructed to brush their teeth carefully in order to prevent gingival injury [ ] . a regular soft toothbrush or an electric brush can be used to minimize trauma [ ] . any elective dental procedure should be ideally performed prior to starting chemotherapy and after discussion with the primary medical team. the absolute neutrophil count, platelet count, and stage of treatment should be considered before performing any dental procedures in this vulnerable population [ , ] . the presence of central venous catheters (cvc) in this population puts them at risk for central line-associated bloodstream infection (clabsi) and its related complications. clabsi is the most commonly reported hai in most pediatric series. among all the pediatric hai reported to national healthcare surveillance network (nhsn), % were from oncology units; streptococcus viridans ( %) and klebsiella pneumoniae/oxytoca ( %) were the two most common pathogens in this study [ ] . in the nhsn report, antibiotic resistance was noted to be high in oncology units, including ampicillin and/or vancomycin resistance for enterococcus faecium and fluoroquinolone resistance for escherichia coli [ ] . although less than % of enterobacteriaceae were reported to have carbapenem resistance, the emergence of such organisms in this population is of significant concern [ ]. among candida infections in this population, fluconazole resistance among non-c. albicans and non-c. parapsilosis isolates was up to %, whereas fluconazole resistance in c. albicans and c. parapsilosis was < % [ ] . mucosal barrier injury (mbi)-associated laboratory-confirmed bloodstream infections (mbi-lcbi) have gained attention in recent years [ , ] . these are clabsis related primarily to mucosal barrier injury (i.e., mucositis) and not due to the direct presence of the cvc per se. in the nhsn definition, a positive blood culture would qualify as a mbi-lcbi if it results from one or more groups of selected commensal organisms of the oral cavity or gastrointestinal tract and occurred in the presence of signs and symptoms consistent with mucosal barrier injury (mbi) in pho or hsct patients [ ] . eligible organisms for mbi-lcbi include candida species, enterococcus, enterobacteriaceae, viridans group streptococci, other streptococcus species, and anaerobes [ ] . specific guidelines for central line insertion and maintenance bundles have been proposed by the centers for disease control and prevention (cdc) and the infectious diseases society of america (idsa) to reduce the clabsi rates and healthcare costs [ , ] . several studies have demonstrated that a multifaceted approach reduces clabsi rates in this population [ , ] and includes standardizing cvc insertion practices and maintenance bundles, tracking cvc infections using standardized definitions, and using dedicated nursing staff or "cvc champions" specifically trained in cvc maintenance and tracking in conjunction with other infection control methods (including oral and hand hygiene, optimizing nurse/patient ratio, etc.). clabsi is discussed in greater detail in chap. . the american society for blood and marrow transplantation recommends a low microbial diet for hsct recipients [ ] . there is little evidence, however, to suggest that this helps in pho patients. routine safety in handling and preparing food should be practiced by patients and parents. in general, eating unpasteurized milk/ cheese, undercooked meat, and raw fruits and vegetables is discouraged during periods of neutropenia to reduce incidence of infection. the need to minimize risk of infection, however, should be balanced with the nutritional needs and quality of life of the patient [ , ] . pets can be a great source of companionship and comfort to children; however, there are several diseases that can be transmitted by pets to these immunosuppressed hosts [ ] [ ] [ ] . certain animals like reptiles, birds, rodents, or other exotic animals that cannot be immunized and could carry unusual human pathogens should not be kept as pets in households with pho or hsct patients. immunosuppressed patients should avoid petting zoos due to the risk of diseases secondary to enteric pathogens (such as salmonella or campylobacter) [ ] [ ] [ ] [ ] . dogs and cats, preferably more than year old, are generally considered safe for pho and hsct patients. they should be routinely evaluated by veterinarians for diseases and their immunizations kept up-to-date. extreme care should be taken to maintain hand hygiene during and after handling the pets [ ] [ ] [ ] [ ] . further information regarding pet therapy is available in chap. . studies performed in adult oncology patients have consistently shown the benefit of using prophylactic antibiotics in reducing the incidence of bacterial infections [ ] . levofloxacin prophylaxis in adults has been shown to reduce the incidence of fever, bacterial infection, hospitalization rates, and all-cause mortality [ , ] . based upon such data in adults, the idsa guidelines for the use of antimicrobial agents in neutropenic patients with cancer state that fluoroquinolone prophylaxis should be considered for high-risk patients with prolonged severe neutropenia [ ] . pediatric studies on antibiotic prophylaxis are limited. a pediatric pilot study on the use of ciprofloxacin prophylaxis for pediatric patients receiving delayed intensification therapy for acute lymphoblastic leukemia (all) showed a reduction in hospitalization, intensive care admission, and bacteremia when compared to controls [ ] . in another study, levofloxacin prophylaxis in patients with all reduced the odds of febrile neutropenia, possible bacterial infection, and confirmed bloodstream infection by ≥ %. it also reduced the use of other broad-spectrum antibiotics and the incidence of c. difficile infections [ ] . in other studies, however, ciprofloxacin prophylaxis did not decrease the incidence of overall bacteremia or duration of fever or mortality in pediatric acute myelogenous leukemia (aml) patients [ ] . furthermore, increasing quinolone resistance among gram-negative organisms is a concern recently observed in the nhsn database of pediatric oncology patients with clabsi [ ] . in addition, the use of antimicrobial prophylaxis in pho could increase the possibility of developing other mdros, invasive fungal infections, or drug-related toxicities. though some authors suggest that antibiotic prophylaxis should be considered in children undergoing induction chemotherapy for all, there is currently insufficient data to inform definitive guidelines for antibiotic prophylaxis to prevent bacterial infections in pediatric oncology patients [ ] [ ] [ ] . notably, an open-label randomized clinical trial was recently conducted of levofloxacin prophylaxis vs. no prophylaxis in children with aml, relapsed all, and hsct recipients. among patients with aml and relapsed all, prophylaxis was associated with a reduction in rates of bacteremia; there was a numeric reduction in bacteremia in the hsct recipients, but this did not achieve statistical significance. it is unclear at this time how these new findings will influence practice and future guidelines [ ] . infections with common respiratory and gastrointestinal viruses can result in significant morbidity and mortality in pho and hsct patients. the most common respiratory viruses encountered include rhinovirus, coronavirus, adenovirus, rsv, parainfluenza, human metapneumovirus, and influenza. common gastrointestinal viruses affecting both healthy and immunocompromised children include norovirus, rotavirus, enteric adenoviruses, and enteroviruses among others. infection prevention strategies should include education provided to the patient and the family about hand hygiene, prevention techniques, avoidance of ill visitors, disease surveillance in the community and hospital, vaccination against influenza and prompt identification, and testing and treatment (if possible) of any respiratory viral illness. implementation of routine infection control prevention policies on oncology wards should reduce transmission of common respiratory and gastrointestinal viruses. all visitors should be screened for any signs and symptoms of acute viral illness and restricted from visitation on the unit or contact with any immunocompromised hosts. chapter outlines infection control guidance for hospital visitors in greater detail. immunization of healthcare workers and household contacts needs special consideration in settings with pho and hsct patients. given the immunosuppressed status of children with malignancy and/or hsct, immunization of those closest to them at home and those caring for them in the hospital is critically important in preventing infections. live attenuated vaccines contain a theoretical risk of being transmitted to an immunocompromised host. live oral polio vaccine, which is no longer administered in the united states, is an absolute contraindication for people taking care of this high-risk population. however, data suggests that measles, mumps, and rubella (mmr), varicella zoster, and herpes zoster vaccines can be safely provided to healthcare workers and household contacts [ ] . if healthcare personnel develop a rash that cannot be covered within the first days following receipt of the varicella vaccine, they should avoid any contact with immunocompromised patients until all rash has crusted to avoid the potential risk of transmitting vaccine strain varicella to patients [ ] . infants living in households with persons who are immunocompromised including pho and hsct patients may be safely immunized against rotavirus; it is recommended, however, that immunocompromised persons avoid contact with the infant's diapers/stool for weeks following vaccination to minimize risk of acquiring vaccine strain rotavirus infection [ ] . an inactivated influenza vaccine is preferred for personnel taking care of immunocompromised children as opposed to live attenuated influenza vaccine [ ] . vaccination against other non-viral pathogens (such as pneumococcus or pertussis) by family members is another important method to minimize the risk of serious infection in pho patients. hospital environments are designed to minimize the potential for fungal disease in the highest-risk patients. high efficiency particulate air (hepa) filters have been shown to reduce nosocomial infection in hsct patients, and the cdc recommends hepa filters in hsct recipient's rooms. the rooms should also have directed airflow and positive air pressure and be properly ventilated (≥ air changes per hour) [ ] . avoidance of carpets and upholstery is also recommended. since outbreaks secondary to aspergillus have been reported during hospital renovation or construction, appropriate containment should be in place, and strict precautions should be taken to prevent exposure to patients during such periods [ ] . infection control and prevention departments should be involved in risk assessment, planning, and approval of all construction or renovation projects in healthcare facilities including inpatient units, clinics, and infusion centers caring for these patients [ ] . cytotoxic chemotherapies and radiation therapy used in the treatment of malignancies are myelosuppressive and result in variable duration and severity of neutropenia. in addition, certain malignancies that originate from bone marrow precursors (i.e., leukemia) or metastasize to the bone marrow (e.g., lymphoma, neuroblastoma, and sarcomas) can result in a decreased number of normal blood cell precursors and consequent neutropenia. hence, pediatric cancer and hsct patients are frequently immunosuppressed and at risk for a wide range of pathogens. febrile neutropenia is a common condition in the pho/hsct population. with regard to this entity, fever is defined as a single temperature > . °c ( °f) or a temperature ≥ . °c ( . °f) on two occasions hour apart. neutropenia is classified as mild (absolute neutrophil count [anc] > - /mm ), moderate (anc ≥ - /mm ), or severe (anc < /mm ). febrile neutropenia (also known as fever and neutropenia) is the combination of these two events in the patient with malignancy or hsct and is a common complication of cancer treatment. it has been estimated that - % of patients with solid tumors and up to % of patients with hematologic malignancies will develop fever during at least one chemotherapy cycle associated with neutropenia [ ] . moreover, fever may be the only indication of a severe underlying infection as other signs and symptoms are often absent or minimized due to an inadequate inflammatory response. therefore, physicians must be particularly aware of the infection risks, diagnostic methods, and antimicrobial therapies required for the management of febrile neutropenia in cancer patients. in the majority of febrile episodes, a pathogen is not identified, with a clinically documented infection occurring in only - % of cases. of these patients, bacteremia occurs in - %, with most episodes seen in the setting of prolonged and/or profound neutropenia (anc < neutrophils/mm ) [ , ] . on the other hand, the most common sites of focal infection include the gastrointestinal tract, lung, and skin [ ] . over the past five decades, the rates, antibiotic resistance, and epidemiologic spectrum of bloodstream pathogens isolated from febrile neutropenic patients have changed substantially under the selective pressure of broad-spectrum antimicrobial therapy and/or prophylaxis [ , ] . early in the development of cytotoxic chemotherapies, during the s and s, gram-negative pathogens predominated in febrile neutropenia. subsequently, during the s and s, gram-positive organisms became more common as use of indwelling plastic venous catheters became more prevalent, which can allow for colonization and subsequent infection by gram-positive skin flora [ , ] . gram-positive bacteria currently account for - % of culture-positive infections in pediatric cancer patients [ ] . importantly, a recent systematic review of the epidemiology and antibiotic resistance of pathogens causing bacteremia in cancer patients since showed a recent shift from gram-positive to gram-negative organisms [ ] . the main causes for this new trend are to be determined, but the use and duration of antibiotic prophylaxis are an important factor to consider as the incidence of gram-negative bacteria was significantly higher in groups who did not receive antibiotic prophylaxis. the use of antibiotic prophylaxis, however, may conceivably select for resistant organisms; increasing rates of antibiotic resistance in both gram-negative and gram-positive bacteria have been reported in the global community as well as the cancer population and are of significant concern [ , , ] . overall, the most common blood isolate in the setting of febrile neutropenia is coagulase-negative staphylococci. other less common blood isolates include enterobacteriaceae, non-fermenting gram-negative bacteria (such as pseudomonas), s. aureus, and streptococci (see table . ). providers should review the local data at their institution for prevalent blood isolates and antimicrobial susceptibility profiles. management of febrile neutropenia continues to evolve given the awareness that interventions previously considered standard of care (such as inpatient treatment with intravenous broad-spectrum antibiotics) may not be necessary nor appropriate for all patients [ ] . it has become increasingly important to identify patients at high risk of infectious complications requiring more aggressive management and monitoring (i.e., inpatient setting with intravenous antibiotics). in addition, clinicians may be able to identify low-risk patient populations who may be managed in a less aggressive and more cost-effective manner (i.e., outpatient setting and/or with oral antibiotics). in order to address these issues, algorithmic approaches to neutropenic fever, infection prophylaxis, diagnosis, and treatment have been developed [ , [ ] [ ] [ ] . it is well established that stratification of patients to determine the risk for complications of severe infection should be undertaken at presentation of fever [ , ] . this determines the type of empiric antibiotic therapy (oral vs. intravenous), venue of treatment (inpatient vs. outpatient), and duration of antibiotic therapy. generally, the risk for serious infection is directly related to the degree and duration of neutropenia. pediatric patients with mild (anc ≥ ) and brief periods of neutropenia (< days) are less likely to have infectious complications than those [ , , ] a. dutta and r. flores with moderate to severe neutropenia (anc ≤ ) lasting more than days. similarly, the risk for bacteremia and septicemia increases dramatically when the anc is < . infectious complications that are more common with severe and prolonged neutropenia include bacteremia, pneumonitis, cellulitis, and abscess formation. it is important to consider individual patient risk incorporating the latest recommendations for the management of neutropenic fever in children with cancer and hsct [ , ] . patients are generally stratified as either high or low risk as follows: . high-risk patients -anticipated prolonged (> days duration) and profound neutropenia (anc < cells/mm following cytotoxic chemotherapy) and/or significant medical comorbid conditions, including hypotension, pneumonia, new-onset abdominal pain, or neurologic changes [ ] . low-risk patients -anticipated brief (< days duration) neutropenic periods in those with no or few comorbidities [ ] in addition, risk classification may be based on the multinational association for supportive care in cancer (mascc) score (table . ) [ ] . a mascc risk score of ≥ is recommended as the threshold for definition of low risk, with % of such patients developing serious medical complications compared to % of those scoring < [ ] . however the mascc score was developed and validated in adults and has not been validated in a pediatric population. the consensus in the field is for all patients considered to be at high risk by mascc or by clinical criteria to be treated as inpatients with empiric iv antibiotic therapy. carefully selected low-risk patients may be candidates for oral and/or outpatient empiric antibiotic therapy. table . summarizes the recommendation for the management of febrile neutropenia based on recommendations of the idsa and the international pediatric fever and neutropenia guideline panel. importantly, in neutropenic febrile patients with an obvious source of infection on clinical exam, management should be tailored to that source. of note, adequate antibiotic stewardship is of utmost importance during the treatment of neutropenic patients in order to decrease the incidence of antibiotic-related adverse drug events, prevalence of antibiotic resistance, and decrease treatment costs. blood cultures must be closely monitored, and once a microorganism has been identified, an appropriate plan for antibiotic de-escalation and/or treatment duration should be promptly instituted. invasive fungal diseases (ifd) are one of the leading causes of morbidity and mortality in pho and hsct patients and present many diagnostic and therapeutic challenges. one of the principal risk factors contributing to the development of ifd relates to the patient's oncologic diagnosis. patients with aml and high-risk and relapsed all, recipients of allogenic hsct, and those with chronic or severe acute graft-versushost disease (gvhd) are at the highest risk of ifd [ , ] . often a combination of other risk factors is present in these patients which may include prolonged neutropenia, high-dose corticosteroid use, immunosuppressive therapy, parenteral nutrition, presence of a cvc, preceding antibiotic therapy, presence of bacterial coinfection, oral mucositis, and admission to an intensive care unit [ , ] . the highest risk of ifd is during periods of profound neutropenia which for hsct recipients occurs during the first days posttransplant and during neutrophil engraftment [ ] ; for pho patients, the highest risk period is during induction chemotherapy [ ] . high-risk patients use monotherapy with an antipseudomonal β-lactam, a fourth generation cephalosporin, or a carbapenem as empirical therapy in pediatric high-risk fn depending on the local prevalence of multidrug-resistant gram-negative rods strong recommendation high-quality evidence reserve addition of a second gram-negative agent or a glycopeptide for patients who are clinically unstable, when a resistant infection is suspected, or for centers with a high rate of resistant pathogens in an era of growing prophylactic antifungal use, children receiving mold-active agents have been shown to be at higher risk of non-aspergillus species fungal infection [ ] . voriconazole prophylaxis in adults has been shown to be an independent risk factor for mucormycoses [ ] . likewise, breakthrough trichosporonosis has also been reported in patients receiving micafungin as prophylaxis [ ] . these phenomena are likely in part related to the selection of fungi with reduced intrinsic susceptibility to the prophylactic agent. the most common ifd are invasive aspergillosis (ia) and invasive candidiasis (ic), with a recent upward trend seen in non-aspergillus mold infections [ ] [ ] [ ] . among aspergillus species, a. fumigatus is the most common, followed by a. flavus and a.niger [ ] . among non-aspergillus molds, mucormycoses (rhizopus, mucor, absidia) are most frequently reported followed by a number of other species (e.g., fusarium, scedosporium, curvularia, exserohilum, etc.) [ ] . among ic, c. albicans is the single most common candida species, but nonalbicans candida species (especially c. parapsilosis and c. tropicalis) have been increasingly reported among this population [ ] . ifd should be suspected in patients with fever and neutropenia lasting for more than days without any identifiable cause [ ] . ic can present as septic shock or may have more non-specific findings such as fever, cough, nausea/vomiting, abdominal pain, and cutaneous lesions depending on the site of involvement. in children, the most common sites of ic are the lungs, liver, and spleen, but dissemination can occur to the other organs including the heart, eyes, or brain. disseminated disease is an independent risk factor for death in children with ic [ ] . the primary sites of ia are the lungs, skin, and sinuses [ ] . the clinical presentation of fungal rhinosinusitis may include fever, rhinorrhea, nasal congestion, and facial pain; many cases, however, may not present with any symptoms and may be diagnosed based on imaging performed in a persistently febrile patient with profound and prolonged neutropenia. cutaneous lesions can present as macules, papules, or nodular ulcerative lesions with or without surrounding erythema and tenderness. clinical presentation secondary to other molds, such as fusarium or scedosporium, is indistinguishable from ia. mucormycoses deserve special mention since dissemination and death are higher due to ifd caused by these species when compared to ia [ ] . early recognition and prompt treatment of ifd are crucial for optimal management. diagnostic tests should include blood cultures (though often with low sensitivity), cultures of appropriate sterile sites (such as urine or csf), and diagnostic biopsies of involved sites for culture and histopathology. fungal biomarkers can be used as both a screening test during high-risk periods and adjunct diagnostic test in patients with suspected ifd, especially during the periods of prolonged fever and neutropenia. galactomannan (gm) is a cell wall component released by aspergillus species which can be detected in blood, bronchoalveolar lavage fluid, and cerebrospinal fluid. a cutoff value of a gm optical index of ≥ . in blood and a bronchoalveolar lavage fluid level of ≥ is considered a positive test, though an optimum cutoff value is not well defined in children [ , ] . invasive fungal disease due to fungi other than aspergillus species may have negative galactomannan tests. β-d-glucan is a cell wall component found in many (but not all) species of fungi, and an elevated serum β-d-glucan assay can be caused by ic, ia, and other molds [ , ] . the optimum cutoff value of β-dglucan for a positive test is unknown in children, but ≥ pg/ml is used in most studies [ ] . both gm and β-d-glucan assays have variable sensitivity and specificity among children and should be interpreted with caution. the sensitivity of gm has been reported to range from to % in children with malignancy and ia [ , ] ; by contrast the β-d-glucan assay has high sensitivity for ifd (~ %) but suffers from poor specificity [ ] . false-positive β-d-glucan can be due to systemic bacterial or viral coinfection, receipt of antibiotics (such as piperacillin-tazobactam or amoxicillinclavulanate), hemodialysis, receipt of albumin or intravenous immunoglobulin, material containing glucan, oral mucositis, and other gi mucosal breakdowns [ ] . other pcr-based fungal diagnostic tests are under investigation but have low sensitivity and specificity. gm and β-d-glucan monitoring twice weekly is suggested to evaluate treatment response in those with confirmed/probable disease and as a screening tool in patients at high risk for ifd [ , ] . all pho and hsct patients with febrile neutropenia that persists beyond days and/or those with suspected ifd should undergo computed tomography of the chest, abdomen, and pelvis and of other areas if indicated [ ] . the most common findings on imaging suggestive of ifd are pulmonary nodules, especially those with a halo sign, air crescent sign, or cavitations. hepatosplenic and renal nodules should also raise suspicion of ifd. other studies to consider include an echocardiogram and dilated retinal examination, especially in patients with disseminated candidiasis. if symptoms of sinusitis or new lesions on the palate are present, a prompt nasal endoscopic examination and ct of sinuses are warranted. there are three main classes of antifungals used in patients with ifd: ( ) polyenes, which include amphotericin b (amb) and its lipid formulations (liposomal amb is most commonly used in pho and hsct patients); ( ) triazoles (fluconazole, itraconazole, voriconazole, and posaconazole); and ( ) echinocandins (caspofungin, micafungin, anidulafungin). antifungal prophylaxis should be considered in patients who are at high risk for ifd including hsct recipients and those undergoing intensive remission-induction therapy or salvage-induction therapy [ , ] . a high incidence of ifd has been reported in children with aml (newly diagnosed and relapsed) [ ] and patients with relapsed all [ ] , and such patients may be considered candidates for prophylaxis. among hsct recipients, those with an unrelated donor or a partially matched donor are at higher risk of ifd [ ] . recent studies show that children with aml receiving antifungal prophylaxis have reduced rates of induction mortality and resource utilization compared to those who did not receive prophylaxis [ ] . posaconazole was found to be superior to fluconazole or itraconazole in reducing incidence of ifd in children [ ] . echinocandins have been shown to be as or more effective for ifd prophylaxis than triazoles, especially in hsct recipients, with less adverse effects and can be an alternative option for prophylaxis [ ] . the idsa and the european conference on infections in leukemia (ecil- ) recommend using posaconazole, voriconazole, or micafungin during prolonged neutropenia to prevent ifd [ , ] . posaconazole is recommended for prophylaxis in patients with gvhd who are at high risk of ia [ ] . variable absorption of oral azoles in children should be taken into consideration when choosing oral antifungals. for patients with prolonged fever and neutropenia without an alternative explanation, consideration must be given to the possibility of an active fungal infection. empiric antifungal therapy should be considered for neutropenic patients with persistent or recurrent fevers after - days of antibiotic therapy and whose overall duration of neutropenia is expected to be > days [ ] . in low-risk patients, routine use of empiric antifungals is not recommended [ ] . liposomal amphotericin b or an echinocandin, both of which are fungicidal, are the first-line therapy for empiric antifungal treatment [ ] . there is insufficient data to provide specific guidance for patients with concern for a new fungal infection who are already receiving moldactive (i.e., anti-aspergillus) prophylaxis; however, some experts suggest switching to a different mold-active antifungal [ ] . surgical debridement of any fungal lesions or abscesses and prompt removal of cvc in the event of fungemia are crucial to reduce the progression of ifd. therapeutic drug monitoring (tdm) should be performed for patients receiving voriconazole, itraconazole, and posaconazole. there is extreme variability in triazole serum levels among pediatric patients owing to diversity in bioavailability in this population. for voriconazole tdm, a serum trough level between and mcg/dl has been considered safe and effective in preventing breakthrough ifd in children [ ] . for posaconazole, a trough level of . mg/l- mg/l has been shown to be effective [ ] . due to increased toxicity associated with vinca alkaloids, high doses of cyclophosphamide, and anthracyclines, azoles should not be co-administered with these agents. the antifungal agents most commonly used in children with pho and hsct and their indications are noted below (table . ). although combination antifungals are not well studied in children, they are used frequently in this population. pediatric data are variable regarding the benefit of combination antifungal therapy but overall report an increase in adverse events [ ] ; the risk of systemic toxicity must therefore be taken into account when considering the use of antifungal combinations. combination therapy could be considered in patients with refractory disease or as salvage therapy. granulocyte transfusions for profound or persistent neutropenia, adjunctive cytokines (e.g., granulocyte colony-stimulating factor [gcsf]), and reduction of immunosuppression and tapering of steroids are recommended as an adjunct to antifungal agents in the treatment of ifd [ ] . in summary, children and adolescents with malignancy have additional risk factors for healthcare-associated infections. meticulous attention to personal and oral hygiene, diet, environmental safety, and appropriate immunizations should be practiced in this high-risk population. the use of antimicrobial prophylaxis should be considered in periods of severe neutropenia to prevent bacterial and fungal infections as necessary. prompt diagnosis and management strategies to prevent infectious complications are key to preventing morbidity and mortality in these immunocompromised hosts. daily bathing with chlorhexidine and its effects on nosocomial infection rates in pediatric oncology patients infection prevention in the cancer center oral and dental considerations in pediatric leukemic patient guideline on dental management of pediatric patients receiving chemotherapy, hematopoietic cell transplantation, and/or radiation pathogen distribution and antimicrobial resistance among pediatric healthcareassociated infections reported to the national healthcare safety network antibiotic use during infectious episodes in the first months of anticancer treatment-a swedish cohort study of children aged - years mucosal barrier injury laboratory-confirmed bloodstream infection: results from a field test of a new national healthcare safety network definition the centers for disease control and prevention definition of mucosal barrier injury-associated bloodstream infection improves accurate detection of preventable bacteremia rates at a pediatric cancer center in a low-to middle-income country strategies to prevent central line-associated bloodstream infections in acute care hospitals: update guidelines for the prevention of intravascular catheter-related infections preventing clabsis among pediatric hematology/oncology inpatients: national collaborative results rapid cycle development of a multifactorial intervention achieved sustained reductions in central line-associated bloodstream infections in haematology oncology units at a children's hospital: a time series analysis guidelines for preventing infectious complications among hematopoietic cell transplantation recipients: a global perspective high rates of potentially infectious exposures between immunocompromised patients and their companion animals: an unmet need for education pet ownership in immunocompromised children--a review of the literature and survey of existing guidelines should immunocompromised patients have pets? antibiotic prophylaxis for patients with acute leukemia levofloxacin to prevent bacterial infection in patients with cancer and neutropenia antibacterial prophylaxis after chemotherapy for solid tumors and lymphomas clinical practice guideline for the use of antimicrobial agents in neutropenic patients with cancer: update by the infectious diseases society of america a pilot study of prophylactic ciprofloxacin during delayed intensification in children with acute lymphoblastic leukemia levofloxacin prophylaxis during induction therapy for pediatric acute lymphoblastic leukemia clinical and microbiologic outcomes of quinolone prophylaxis in children with acute myeloid leukemia effect of levofloxacin prophylaxis on bacteremia in children with acute leukemia or undergoing hematopoietic stem cell transplantation: a randomized clincial trial updated recommendations of the advisory committee on immunization practices for healthcare personnel vaccination: a necessary foundation for the essential work that remains to build successful programs redbook: report of the committee on infectious diseases guidelines for environmental infection control in health-care facilities. recommendations of cdc and the healthcare infection control practices advisory committee (hicpac) management of fever in neutropenic patients with different risks of complications repeated blood cultures in pediatric febrile neutropenia: would following the guidelines alter the outcome? pediatr blood cancer etiology and clinical course of febrile neutropenia in children with cancer changes in the etiology of bacteremia in febrile neutropenic patients and the susceptibilities of the currently isolated pathogens contemporary antimicrobial susceptibility patterns of bacterial pathogens commonly associated with febrile patients with neutropenia emergence of carbapenem resistant gram negative and vancomycin resistant gram positive organisms in bacteremic isolates of febrile neutropenic patients: a descriptive study changing epidemiology of infections in patients with neutropenia and cancer: emphasis on gram-positive and resistant bacteria recent changes in bacteremia in patients with cancer: a systematic review of epidemiology and antibiotic resistance management of febrile neutropenia in malignancy using the mascc score and other factors: feasibility and safety in routine clinical practice guideline for the management of fever and neutropenia in children with cancer and hematopoietic stem-cell transplantation recipients: update guidelines for the use of antimicrobial agents in neutropenic patients with cancer outpatient management of fever and neutropenia in adults treated for malignancy: american society of clinical oncology and infectious diseases society of america clinical practice guideline update summary the multinational association for supportive care in cancer risk index: a multinational scoring system for identifying low-risk febrile neutropenic cancer patients clinical practice guideline for the use of antimicrobial agents in neutropenic patients with cancer: update by the infectious diseases society of america invasive mycoses in children receiving hemopoietic sct a prospective, international cohort study of invasive mold infections in children epidemiology and outcomes of invasive fungal infections in allogeneic haematopoietic stem cell transplant recipients in the era of antifungal prophylaxis: a singlecentre study with focus on emerging pathogens invasive mold infections in pediatric cancer patients reflect heterogeneity in etiology, presentation, and outcome: a -year, single-institution, retrospective study antifungal prophylaxis in pediatric hematology/oncology: new choices & new data. pediatr blood cancer breakthrough zygomycosis after voriconazole treatment in recipients of hematopoietic stem-cell transplants trichosporonosis in pediatric patients with a hematologic disorder results from a prospective, international, epidemiologic study of invasive candidiasis in children and neonates risk factors for mortality in children with candidemia invasive mucormycosis in children: an epidemiologic study in european and non-european countries based on two registries practice guidelines for the diagnosis and management of aspergillosis: update by the infectious diseases society of america ecil- ): guidelines for diagnosis, prevention, and treatment of invasive fungal diseases in paediatric patients with cancer or allogeneic haemopoietic stem-cell transplantation clinical practice guideline for the management of candidiasis: update by the infectious diseases society of america threshold of galactomannan antigenemia positivity for early diagnosis of invasive aspergillosis in neutropenic children galactomannan antigenemia in pediatric oncology patients with invasive aspergillosis beta-d-glucan screening for detection of invasive fungal disease in children undergoing allogeneic hematopoietic stem cell transplantation guideline for primary antifungal prophylaxis for pediatric patients with cancer or hematopoietic stem cell transplant recipients antifungal prophylaxis associated with decreased induction mortality rates and resources utilized in children with new-onset acute myeloid leukemia antifungal prophylaxis with posaconazole vs. fluconazole or itraconazole in pediatric patients with neutropenia key: cord- -vwgai k authors: nan title: publication only date: - - journal: bone marrow transplant doi: . /bmt. . sha: doc_id: cord_uid: vwgai k nan introduction & objectives: literature states that human postnatal dental pulp stem cells (hdpscs) have the ability to differentiate to osteoblastic cells. the purpose of this paper is to present the results obtained in the differentiation of hdpscs with three different media and to compare their osteogenic ability. materials & methods: human dental pulp was extracted from teeth of healthy adult subjects aged to years. the pulp was gently removed and immersed in a digestive solution for h at cº. after digestion, cells were cultured and adherent cells were isolated. after the second pass the cells were placed in three different fl asks with three classes of differentiation media. medium : osteodiff (miltenyi®); medium : alpha-mem supplemented with % fetal bovine serum (fbs), u/ml penicillin, . mg/ml streptomycn, and . mg/ml amphotericin b; medium : alpha-mem medium, supplemented with % fbs, mm p-ascorbic acid, mm l-glutamine, u/ml penicillin, . mg/ml streptomycin, and . mg/ml amphotericin b. flasks were incubated at ºc in a % co and the medium changed twice a week for days. to quantify the different amount of mineralized nodules the absorbance rate was used. results & discussion: hdpscs were obtained at a good rate and differentiated with any of the three media into osteoblastic cells that developed mineralization nodules (clusters), as revealed by alizarin red staining. this staining was signifi cantly more intense with medium than medium and medium (absorbance values . , . and . respectively). conclusions: this study demonstrates the ability of hdpscs to differentiate into osteoblasts. the medium (osteodiff medium, miltenyi®) , was the best to differentiate these cells to the osteogenic lineage. long-term haematopoietic reconstitution and clinical evaluation of autologous peripheral blood stem cell transplantation after cryopreservation of cells at - °c in a mechanical freezer for longer than months l. calvet, a. cabrespine-faugeras, n. boiret-dupre , e. merlin, c. paillard, m. berger, j.-o. bay, o. tournilhac, p. halle chu (clermont-ferrand, fr) controlled-rate freezing in or % of dmso and storage in the nitrogen is the standard technique for cryopreservation of hematopoietic progenitor cells (phs). the main inconveniences are its high cost and dmso toxicity. many teams try to reduce dmso infused by phs concentration before cryopreservation or wash before infusion. however, labor intensive increases the cost and not free of cell loss. we developed an easier and cheaper technique, the cryopreservation of the phs at - °c, an uncontrolled rate freezing with . % hes, % albumin and only . % of dmso allowing infusion without wash. this technique preserves the functional capacities of phs, can produce successful engraftment and reduces toxicity during infusion. does the cryopreservation of the phs at - °c allow a long-term hematopoietic reconstitution and clinical course even if storage is greater than months? patients who had undergone autografts ( adults, children) were studied. the median storage time of the phs cryopreserved was . months [ . with . % ( / ) preserved more than months (median , [ - ] ). the median recovery of nucleated cells and cd + cells were similar, for the preserved phs <or > months ( % versus % , p= . ) and ( % versus % , p= . ), respectively. only mild infusion-related toxicity was observed in . % (nauseas/vomiting . %, shivers . %). median time to reach . x /l granulocytes (pn), and x /l platelets (pl) were , and days respectively. delay to reach hematopoietic reconstitution was similar between phs preserved < or > months except for pl > x /l. this delay was signifi cantly longer for phs kept > months versus [ - ] (n= . ) with a correlation between cd + cells dose and the number of days need to reach x /l pl. in order to assess long term hematopoietic reconstitution, only patients without other treatment (n= ) were studied at , and months. median values were , and x /l for the platelets and , , , and , x /l for the pn at , and months respectively. mortality at post-autograft days was of . %. median overall survival was months and years survival rate was of %. the long term hematopoietic reconstitution was satisfactory. this easier and cheaper cryopreservation method leads to successful engraftment even if phs had been cryopreserved more than months. improve mobilization in these patients have been described. another exciting option for these patients is the new cytokine, amd . this agent is an inhibitor of sdf binding to cxcr and appears to promote mobilization of cd + cells into the circulation. the use of this amd in combination with g-csf in patients unable to collect adequate cd + cells with g-csf alone was recently reported in patients with lymphoma and multiple myeloma (mm) . in this study g-csf was given at a dose of mcg/kg per day and amd was started at mcg/kg on day of mobilization. in contrast, clinical studies showed that aml, cll and pcl cells may also be mobilized by amd via cxcr inhibition. due to these concerns, aml, cll and pcl patients are excluded from amd trials. we here report patients ( female/ male) with non hodgkins lymphoma (n= ), mm (n= ) and germ cell cancer (n= ) who failed stem cell mobilization after chemotherapy and g-csf administration (patient characteristics table ). patients received x µg/ kg daily of g-csf for days followed by µg/kg of amd given subcutaneously - hrs before collection on day . our aim was to assess the effect of amd on the mobilization of cd + cells. administration of g-csf and amd were continued daily until end of collection cycle. adequate collection of cd + cells ( . and . x cd + cells/kg) were achieved in patients. in patients additional bone marrow collection were performed, patient failed mobilization with amd . until now patients underwent autologous transplantation with . , . , . and . x cd + cells/kg respectively and achieved sustained leukocyte and platelet engraftment. in conclusion, amd in combination with g-csf was generally safe and offers a new treatment to collect cd + cells for autologous transplant from poor mobilizers. due to the reported mobilization of leukemic cells, amd should be restricted to patients with lymphomas, mm and solid tumors. evaluating the effect of substance p on expansion of human umbilical cord blood cd + haematopoietic stem cells in a serum-free media s. shahrokhi ( ) , m. ebtekar ( ) , k. alimoghaddam ( ) , m. kheirandish ( ) , a. pourfathollah ( ) , a.r. ardjmand ( ), a. ghavamzadeh ( ) ( )tarbiat modares university (tehran, ir); ( ) hematology, oncology and bone marrow transplantation research center (tehran, ir) ex vivo expansion of cord blood hematopoietic stem cells has been progressively interested as alternative sources for stem cell transplantation. using different combination of growth factors especially cytokines has been investigated in most reports, but there are little evidence about regulatory roles of other factors including neuropeptides in this way, then we choose substance p (sp) to evaluate its effect on expansion. material and methods: cd + purifi ed from umbilical cord blood by macs, were cultured in a serum-free liquid culture system. different concentration of sp used in combination with cytokine cocktail of scf, fl, tpo, il and il . phenotypic and functional analysis of the cells produced in culture, was performed by fl owcytometry. count and percentage of cd + cells were compared in different groups of treated cells. results: ex vivo expansion cultures of cd + cells of ucb were signifi cantly increased, in cells cultivated in "sp + cytokine cocktails" group compared cytokine groups alone. conclusion: consideration of the role of other growth factor such as sp along with cytokines, may enable us to overcome the diffi culties before us in ex vivo expansion of cord blood cells. our studies indicate that sp could act as a superior supplement for expansion of ucb-hsc cytokine cocktails. additional studies are needed to establish the functional activity of expanded ucb-hsc as well as the effects of substance p. standard protocols for cryopreservation of peripheral blood progenitor cells (pbpc) use rate-controlled freezing and storage in liquid nitrogen, which are both time-consuming and expensive. in the last years we used a simplifi ed method (galmes et al ) consisting of storage in a mechanical freezer at - ºc, with dmso as the sole cryoprotectant. this study evaluates the safety of this approach, in terms of infusion-related toxicity and hematopoietic reconstitution, in consecutive autologous transplantations performed from / to / in patients (median age ; underlying disease: lymphoma in , myeloma in , acute leukaemia in , breast cancer in ). after mobilization with g-csf ± chemotherapy (usually cyclophosphamide . g/m²) pbpc were collected in a cs + separator (fenwall), mixed in autologous plasma and dmso (to a fi nal concentration of %) and frozen in plastic bags (cryocyte, fenwall) at - ºc. median cd + count was . x /kg and median storage duration was days ( - ). infusion-related toxicity was frequent ( %) and generally mild (transient hypoxemia, broncospasm, hypertension or arrhythmia, and abdominal pain, nausea or diarrhea) but there were cases of acute congestive heart failure and anaphylactic shock (probably related to dmso). engraftment to neutrophils and , platelets/ul occurred on days + and + (median). bacteremia occured in % transplantations, and grade or toxicity in %. median hospitalisation duration was days. mortality at day + and + was . and . % respectively. an engraftment delay beyond d+ was seen in cases. there were no secondary graft failures. with a median follow up of months, % patients are alive. these results confi rm the feasibility and safety of this simpler and cheaper cryopreservation methodology. belarus y. isaikina, n. minakovskaya, o. aleinikova belarusian center for ped oncohematology (minsk, by) introduction: recent studies suggest that cotransplantation of mesenchymal stem cells (mscs) can improve the engraftment of allogeneic hematopoietic stem cells and prevent graft-versus-host disease (gvhd) due to their immunomodulatory properties. we analyzed the clinical effect of msc infusion on day + after hsct for prophylaxis of gvhd and applying of mscs for treatment of severe steroid-resistant gvhd. patients and methods: eight pts after allogeneic hematopoetic stem cell transplantation (hsct) underwent mscs infusions (median age of pts was years, male/female: / ) between and . diagnoses included:all- , aml- , aa- , mds- .gvhd prophylaxis for pts with all, mds consist of csa and mtx mg/m² (n= ); for pts with aa -csa+mmf; for pts with aml -csa and mtx mg/m² (n= ). for the treatment of gvhd all pts received metylprednisolon - mg/kg. mscs were prepared applying technique of expansion in vitro from bone marrow of hla-identical siblings, haplo-identical and haplo-nonidentical family donors and unrelated donors. four pts received mscs once and four -twice. for three pts mscs was used for prophylaxis of gvhd on day + after hsct and the median dose was , ( , - , )x /kg and fi ve pts received mscs for treatment of steroid-resistant gvhd with medium time of mscs infusion after hsct ( - ) days and the dose was , ( , - , )x /kg. results: there was no evidence of early and late side effect of msc infusion. one patient died from pulmonary gvhd month after cotransplantation mscs and seven pts-alive. all pts (n= ), who received mscs on day + for prophylaxis gvhd developed grades ii-iv gvhd and needed the secondary mscs infusion and the median time between mscs infusions were ( - ) days. four pts out of fi ve with steroid-resistant gvhd showed signifi cant improvement of clinical sign of gvhd that allowed reducing immunosuppressive therapy and stopping the steroids. conclusion: our experience demonstrates the absence of positive gvhd prophylactic effi cacy when infusion of mscs was done on day + . however, we observed decreasing of gvhd grades from iii-iv to -ii, when mscs were used as treatment of steroid-resistant gvhd. clinical characteristics of early-onset acute graft-versushost disease after allogeneic haematopoietic stem cell transplantation t. yamashita, y. najima, t. kikuchi, h. muto, c. sakurai, w. munakata, m. yamamoto, k. ohashi, h. sakamaki, h. akiyama tokyo metropolitan komagome hospital (tokyo, jp) acute graft-versus-host disease (gvhd) is one of the major factors that have infl uence on the outcomes of allogeneic hematopoietic stem cell transplantation (hsct). traditionally, acute gvhd has been defi ned as a syndrome after neutrophil engraftment within the fi rst days following hsct. but in our practice, we sometimes encounter acute gvhd that may occur s both early, even before engraftment, and late, beyond day . the latter has been defi ned as "late-onset acute gvhd", but the former may not be clearly identifi ed yet. in this retrospective study, we evaluated the incidence, clinical manifestations and outcomes of "early-onset acute gvhd", defi ned as that occurring before engraftment after transplantation, among consecutive myeloablative allogeneic hscts at our hospital. of patients, the median age was years. ninety-three percent of patients received allogeneic hsct for hematologic malignancies. thirty-eight percent of patients received an hlamatched related donor transplant, % received hla-matched unrelated donor grafts and % received hla-mismatched unrelated donor grafts. the stem cell source was bone marrow in % of patients and peripheral blood in %. the conditioning regimen was tbi-based for % of patients and % received busulfan-based conditioning. forty-three percent (n= ) of the cases developed grade ii-iv acute gvhd. of these, ( %) cases were described as early-onset acute gvhd (group e). other cases of acute gvhd occurred after engraftment (group c). the median onset date of acute gvhd is day in group e and day in group c. grade iii-iv acute gvhd was seen in % of group e and in % of group c (p= . ). the frequency and severity of each involvement site were comparable in both groups. major primary therapy for acute gvhd was mpsl - . mg/kg/day, but % cases in group e were refractory for this primary therapy and % in group c (p= . ). three-years overall survival (oas) was % in group e and % in group c (p= . ). in group c, oas of cases without gi symptoms was %, whereas oas of cases with gi involvement was % (p= . ). in group c, oas was not affected by with or without gi-gvhd (p= . ). in conclusion, early-onset acute gvhd accounts for a substantial proportion of acute gvhd after allogeneic hsct. patients with early-onset acute gvhd tend to be refractory to steroid therapy and will have poor prognosis if gi involvement exists. contrast enhanced ultrasound sonography in intestinal acute graft-versus-host disease e. benedetti ( ) a year old female with high risk acute b cell leukemia received a fully ablative peripheral blood stem cell transplant from a allele (at the b locus) mismatched unrelated donor. conditioning consisted of cy/tbi and gvhd prophylaxis of cyclosporine (csa) and short course mtx. on day + she developed steroid refractory (biopsy proven) acute skin gvhd. photopheresis was started with major skin improvement. on day + she developed nausea, vomiting and profuse diarrhea. standard endoscopy with gastric biopsies showed gvhd. infections were ruled out. a trans-abdominal sonography (ta-us) revealed mucosal oedema and thickening of the terminal ileum ( . mm) and the ascending colon. moreover, pillcam capsule endoscopy showed mucosal oedema, erosions and lymphagectasies. infl iximab at mg/kg was added and, after doses, despite a major clinical improvement, her terminal ileum was still thickened. to investigate if this thickening was associated with residual active gvhd she underwent a contrast enhanced ultrasound sonography (ceus) using a linear phased-array . -mhz transducer. a sulphur hexafl uoridebased with a phospholipid shell microbubble contrast agent (sonovue®, bracco) was injected i.v. as a bolus ( . ml) followed by ml saline fl ush. sonovue® is a blood pool second generation contrast agent. ceus showed an intense and sustained enhancement in the arterial phase involving the whole ileum wall with a late phase wash out. such enhancement pattern has been previously described in active crohn disease. given the clinical improvement, infl iximab was discontinued to reduce the risk of infections. however, as ceus revealed active gvhd she continued on budesonide, beclometasone, csa and prednisone. forty days later her abdominal symptoms had completely resolved and a ta-us showed a normal terminal ileum. four months later her intestinal gvhd (confi rmed by colon biopsies) fl ared. ceus was performed on descending colon (most involved intestinal tract by standard ultrasonography) and showed intense arterial phase enhancement with late phase wash out. rituxan and mmf were added with slow resolution of symptoms and normalisation of us features. in conclusion ceus showed residual gvhd activity despite the improved clinical symptoms. moreover, good concordance with clinical symptoms and standard colonoscopy when gvhd fl ared was also shown. further prospective studies are needed to evaluate its usefulness in monitoring intestinal gvhd. extensive chronic graft-versus-host disease is a frequent complication after peripheral blood stem cell transplantation -results of long-term follow-up d. stamatovic, l. tukic, b. balint, o. tarabar, m. elez, g. ostojic, b. todoric zivanovic, z. tatomirovic, o. tasic, b. cikota, m. malesevic, s. marjanovic military medical academy (belgrade, rs) introduction: many studies have compared effi cacy of allogeneic stem cell transplantation (sct) from peripheral blood (pb) with bone marrow (bm), but fi nal conclusion concerning this treatment modality is still not well defi ned. aim: to compare effi cacy of pbsct with bmt in the treatment of hematological malignancies with respect to engraftment, transfusion need, frequency and severity of acute and late complications and overall survival (os). methods: we have analyzed patients (pts), median age years ( - ), m/f / , with various hematological diseases (saa- , cml- , aml- , all- , mds- , mm- , mh- , granulocytic sarcoma- ) in whom we perfomed allogeneic sct from till . in pts we perfomed secondary allogeneic sct in due to graft rejection ( ) or relapses ( pts). pts were divided into two groups concerning sc origin- pts in bm group and pts in pb group. all pts had hla-dr sibling transplant ( singeneic, fully matched, mismatched and haploidentical). sc were collected from bm up to standard method and from pb with one apheresis after fi ve days aplication of granulocytic growth factor. all pts have received unmanipulated suspension of sc. conditioning were adjusted to primary diseases and gvhd prophylaxis was mostly combination of cyclospirine a and metothrexate. prevention of infections were standard. results: pts with sc originate from pb have received signifi cantly more mononuclear cells ( , ± , vs , ± , , p< , ) in comparisson with bm. engraftment was more rapid (p< , ) in the pb group approximately for days. transfusion requirements were much higher in bm group (p< , ). those pts had more frequent oropharingeal mukositis grade - ( , % vs , %, p< , ). there were no difference in the incidence of acute ( , % vs , %, ns) or chronic gvhd ( , % vs , %, ns). pts with pbsct had signifi cantly more frequent extensive cgvhd ( , % vs , %, p< , ). there were no difference considering trm ( , % vs , %, ns) or relapses ( , % vs , %, ns). pts with bmt had better overall survival but with no statistical signifi cancy. conclusion: results of this analysis mostly corresponds with other studies showing that pbsct have rapid engraftment and less acute complications. pbsct is connected with more frequent extensive chronic gvhd that is potentialy fatal, making results of this particular treatment option less better. future will bring defi nite estimation of pbsct effi cacy. a preliminary study of human natural killer t-cell recovery post allogeneic stem cell transplantation b. rees ( ) , r. morse ( ) , s. robinson ( ) , j. hows ( ) , c. donaldson ( ) ( )centre for research in biomedicine, university of the west of england (bristol, uk); ( )university hospitals bristol nhs foundation trust (bristol, uk) natural killer t cells (nkt), defi ned by their cell surface immunophenotype cd +, v alpha +, v beta + and their specifi c activation pathway by the glycolipid alpha-galactosyl ceramide are a unique and small ( . - . %) subset of lymphocytes. these cells may play a key role in the cure of leukaemia after stem cell transplantation (sct) through activation of the graft versus leukaemia (gvl) effect. they have the ability to stimulate both innate and adaptive immune responses through cytokine production and the activation of 'classical' t, b and natural killer (nk) cells. campath, a complement fi xing monoclonal antibody targets the cd antigen expressed by t, b and nk cells and may be used in vitro and/or in vivo for donor lymphocyte depletion during stem cell transplantation. our previous work has shown that cd +, v alpha +, v beta + nkt cells also express the cd antigen and so are also susceptible to damage by campath. twelve patients (median age . years, range - ) on the bmt unit, university hospitals bristol were recruited. diagnoses were aml ( ), all ( ), anll ( ), cml ( ), mds ( ), nhl ( ) , and hd ( ) . seven received reduced intensity conditioning, tbi and / received campath. all patients received adult stem cells, from matched siblings, from unrelated donors. nine survived more than year, including the patient with hd who relapsed months post autologous sct and is alive months post matched unrelated sct. the normal range for nkt cell numbers in adult blood was established, mean . x /l (sd . ) (n= ). cells stained with cd -pecy , v alpha -fitc and v beta -pe were analysed using the becton dickinson facs vantage se cell sorter with cell quest software. recovery of nkt cells was studied up to - months post transplant, with mean levels of . ± . x /l. all individual values were below those in the normal adult population. recovery of other lymphocyte subsets was comparable with those reported in previous studies. nk cells recovered to within their normal range to months post sct, cd t cells numbers were within the normal range by approximately months and cd t cells only attained values in their normal reference range by months. the slow recovery of nkt-cells has not been previously reported and this may contribute to a reduced gvl effect. n. nakano, a. kubota, m. tokunaga, y. takatsuka, s. takeuchi, t. itoyama, a. utsunomiya imamura bun-in hospital (kagoshima, jp) background: adult t-cell leukemia/lymphoma (atll) has a poor prognosis because of its chemo-resistance. many chemotherapeutic regimens have been created but none of them have shown suffi cient results. we proposed allogeneic stem cell transplantation (allo-sct) for atll patients and showed an improved survival rate. however, relapse or progression of atll is one of the major limiting factors of survival in post sct patients. objectives: in order to establish a better treatment strategy for poor responders after sct for atll, we analyzed the outcome of relapse or progression cases after allo-sct. we paid special attention to the graft versus atll (gvatll) effect. methods: there were atll patients in which allo-sct was performed in imamura bun-in hospital (ibh) from june to november . twenty seven cases survived over days after sct. sixteen of the patients relapsed. using data in medical records of ibh, we analyzed transplant characteristics and the outcome of these patients retrospectively. results: disease status at sct was cr in pts, pr, sd, and pd. eight patients received conventional stem cell transplantation (cst) and the other eight patients received reducedintensity stem cell transplantation (rist). fourteen patients in obtained remission ( cr and pr), but the remaining did not ( sd and pd) after sct. the sites of relapse or progression in were skin in patients, lymph node, peripheral blood, central nervous system, and bone. all patients discontinued immunosuppressants after relapse or progression. eleven patients obtained remission. especially, in out of patients, remission was obtained only by discontinuation of immunosuppressants, and the time to remission after discontinuation of immunosuppressants was between to days. twelve patients were complicated with acute gvhd (grade i-iv). twelve patients died after sct. the causes of death were disease progression of atll in patients, acute gvhd, infectious complications, and interstitial pneumonia. four patients who were complicated with acute gvhd survived over months. conclusions: a certain number of patients obtained remission only by the discontinuation of immunosuppressants. four patients survived more than years with their complication of acute gvhd. these results suggest that the gvatll effect after sct exists and plays an important role in longer survival for poor responders of post allo-sct in atll patients. adoptive immune transfer in paediatric and young adult patients with refractory malignancies p. sovinz, w. schwinger, h. lackner, m. benesch, a. moser, c. urban medical university graz (graz, at) background: patients with metastatic malignancies refractory to or relapsing after conventional ± high-dose chemotherapy have a poor prognosis. graft-versus-tumor (gvt) effects have been reported in small numbers of patients for various solid tumors. patients and methods: eight pediatric and young adult patients (male: female = : ; age . to years) underwent allogeneic hematopoietic stem cell transplantations (allohsct). diagnoses were relapsed/ refractory neuroblastoma (n= ), second relapse of hodgkin's disease, refractory mediastinal large-b-cell-lymphoma, metastatic ewing sarcoma/ osteosarcoma /wilms tumor, respectively. five patients had received high-dose chemotherapy with autologous stem cell rescue. conditioning regimens consisted of fl udarabine (n= ) combined with melphalan ±atg (n= ) or melphalan/thiotepa/okt (n= ) or treosulfan/thiotepa/okt (n= ); and treosulfan/melphalan (n= ). haploidentical donors (parents, n= ) underwent aphereses: one product was cd / depleted, the other cd selected; grafts from matched donors (siblings:n= , unrelated: n= ) were not manipulated. median cd -number was . x /kg; median cd -number in haploidentical grafts was . x /kg. in the absence of graft-versus-host disease (gvh) immunosuppression was stopped median on day + . to date, a median of donor lymphocyte infusions (dli; - ; dose range: . x to x ) were given to / patients, starting on median day + . results: neutrophil engraftment (> . x /l) was achieved median on day + . acute gvh of the skin (i-ii) developed in patients, of skin+liver (iii) in one; chronic gvh occurred in patients (skin:n= , gut:n= ) there was no transplant-related mortality; / patients survive for a median of days (range: - ) in complete (cr; n= ) or partial remission (pr; n= ) with ongoing regression (disease status not yet evaluated: n= ). two patients who were transplanted in disease progression showed partial response after allohsct but eventually died of progressive disease on day + (mediastinal large-b-cell-lymphoma) and + (neuroblastoma, after the second allohsct). conclusions: eight heavily pretreated pediatric and young adult patients with poor-prognosis metastatic malignancies tolerated the conditioning regimens well. all patients showed at least transient partial response to allohsct ±dli; six patients in partial remission or better before allohsct survive in cr or pr with evidence of further tumor regression. cmv infection in seropositive patients with haematologic malignancies after allogeneic peripheral blood stem cell transplantation t.-d. tan koo foundation sun yat-sen cancer center (taipei, tw) objective: to investigate the incidence and outcomes of cmv infection in our seropositive population patients after allotransplant as compared with other western patients. we also investigate the impact of post-transplant occurrence of acute graft-vs-host disease and the use of anti-thymocyte globulin upon the outcome of our patients. methods: cmv seropositive patients of various hematologic malignancies underwent allogeneic peripheral blood stem cell transplantation at our institute between march and november . we used weekly cmv pcr to monitor cmv infection following neutrophil engraftment until day + or when any infectious complication occurred. when two consecutive pcrs were positive with > copies present or cmv was found histopathologically, we treated patients with intravenous ganciclovir mg/kg q h for to days. results: patients (median age . , ~ ) of various hematologic malignancies including aml (n= ), cml (n= ), all (n= ), nhl (n= ), hl (n= ), myeloma (n= ), myelodysplastic syndrome (n= ), underwent myeloablative or non-myeloablative allotransplant ( vs ). the source of stem cells includes related ( patients), unrelated ( patients), and umbilical cord blood stem cell ( patients). cmv infection or reactivation rate was . % ( in ) with median date of occurrence ranges + to + days with the median of + days and the immediate cmv-related mortality rate was . % ( in ). the incidence of cmv infection in patients with grade ~i vs ii~iv acute gvhd are . % vs . %, respectively, with risk ratio (p= . ). the occurrence of cmv infection in patients with or without the use of anti-thymocyte globulin use was . % vs . %, respectively, with risk ratio . (p= . ). the -year event-free survival and overall survival of our patients with or without cmv infection are . % vs . %(p= . ), and . % vs . %(p= . ), respectively. conclusions: our cmv seropositive patients do not have higher incidence of cmv infection or reactivation than other lower seropositive patients reported in the western world. there is an increased incidence of cmv infection in the patients who suffer from grade ii~iv acute gvhd, and there are signifi cant differences in efs and os between patients with or without cmv infection. on the contrary, the impact of atg use in our patients is not clear. objectives: patients after hematopoietic stem cells transplantation (sct) have markedly increased susceptibility to moulds infections. according to recent data, the moulds of fusarium spp are emerging as human pathogens associated with significant morbidity and mortality in immunocompromised patients. in current report we are describing disseminated invasive fungal infections caused by fusarium incarnatum in three recipients of allogeneic hematopoietic stem cells, a pathogen not earlier reported for such patients. methods: blood samples were analyzed using automatic bact/ alert system. the culture and identifi cation were performed according to conventional microbiological procedures. the sabouraud agar was used for strain's isolation and the samples were incubated in °c for days. the cream to nut-brown mould's colonies were suggestive for fusarium incarnatum. also the microscopic analysis of direct samples revealed microand macroconidias typical for fusarium genus. results: the -years-old male and a -years-old female patients, with relapsed and refractory acute myelogenous leukemia (aml) have been treated by allogeneic sct from matched unrelated donors after myeloablative conditioning. the third patient, a -years-old woman with hodgkin's lymphoma relapsed after autologous sct was transplanted from hla-matched sibling donor after reduced intensity conditioning. all patients suffered from neutropenic fever which did not respond to broad-spectrum antibiotics and fl uconasole. the appearance of nodular, painful skin lesions with characteristic dark red colour and central necrotic area in later stadium suggested skin microembolism caused by infectious microorganism. the mycological analysis confi rmed fusarium incarnatum as a pathogen. i.v. voriconazole in standard doses was started as soon as invasive fungal infection was suspected. the two female patients responded well to voriconazole with gradual resolution of fever and skin lesions. this corresponded with neutrophil engraftment. the male patient with aml died of disseminated fusariosis (autopsy confi rmed) before achieving engraftment. conclusions: we identifi ed fusarium incarnatum as a new mould pathogen which can cause disseminated fatal infections in immunocompromised patients and sct recipients. although the voriconazole was proven to be an effective agent to treat these patients, the hematological recovery seems to be a prerequisite factor needed to survive the disseminated fusariosis. background: infections are the most common complications of stem cells transplantation and chemotherapy induced neutropenia. bacterial infections predominate during the early stage after transplantation. during this phase deep neutropenia and central venous catheter are the most important risk factors. because of high rate of mortality due to gram-negative bacteria, prophylaxis against this microorganisms is mandatory, but this strategy offer gram-positive predomination in all sites of isolation. despite low rate of mortality due to gram-positive bacteria, infections caused by streptococcus today became a real problem. material and methods: during a years period we have performed stem cells transplantation in patients with different hematological malignancies(aml: ; all: ; cml: ; cll: , nhl: ; hodgkin diseases: ; multiple myelomas: ; aplastic anaemia: ;myelofi brosis: ewing sarcoma: ; male: female . median age: years ( - ). in order to monitoring local micro-fl ora we perform in all patient two times a week: blood-culture, sputum, urine-culture, and simples from central venous catheters. cultures were performed using standard microbiological tools. patients were treated in sterile room conditioned with hepa fi lters, gram-negative prophylaxis with ciprofl oxacine , gr. per day, low bacterial diet. results: gram-positive cocci were predominantly isolated microorganisms ( %), then gram-negative bacteria ( %) and fungi ( %). the most frequent isolated bacteria was staphylococcus coagulaza negative, from central venous catheter, while streptococcus pneumonia was the most common bacteria isolated after day + , predominantly from sputum. meticillin resistant staphylococcus aureus (mrsa) was isolated in % from all gram positive bacteria. we have no vancomicyn-resistant enterococcus isolation. conclusion: the epidemiological pattern of bacterial infection continues to evolve globally and locally at the institutional level, as do patterns of susceptibility and resistance. these trends are often associated with local treatment practices and have a signifi cant effect on the nature of empirical antibiotic prophylaxis and therapy. in our center gram positive bacteria were isolated predominantly. gram-positive prophylaxis is doctrinary used in some centers, but there is a problem with gram-positive resistance. heptavalent pneumococcal vaccination may be reasonable choice. background: invasive fungal infections (ifi) are an important life-threatening complication after allogeneic hematopoietic stem-cell transplant (ahsct). risk factors that further increase the risk of ifi in these patients include prolonged neutropenia, graft failure, immunosupression and graft-versus-host-disease (gvhd). aim: to evaluate the effi cacy and safety profi le of posaconazole as prophylaxis of invasive fungal infection after ahsct. material and methods: in patients at high risk who received posaconazole for prophylaxis we analyzed the incidence of ifi during the treatment period. demographic, clinical, laboratorial and radiologic variables of all patients were studied including age, gender, underlying disease and it´s status at allogeneic transplantation, presence of gvhd, treatment with steroids, adverse events, galactomannan antigen in plasma and high resolution computed tomography (ct-scan). adverse events were also analyzed. results: from a total of patients received posaconazol patients were included in the study, among them received ahsct. during the treatment period there were no proven ifi reported. probable ifi were reported in patient. no serious adverse events related to treatment were reported. during the observational period the overall mortality was % ( patients) and none of them died due to ifi. patients ( , %) were receiving steroids during the treatment period and none of them developed ifi. the incidence of global gvhd was %. acute gvhd incidence was %. patients had galactomannan positive and ct-scan were performed in all of them without found ifi in any case. conclusions: posaconazole prophylaxis is a useful and safe approach in order to prevent ifi avoiding systemic antifungal treatment in patients who had undergone ahsct. mucormycosis are an emerging form of invasive fungal infections (ifi) with high mortality rate ( %). early treatment contributes to improve prognosis. posaconazole is a broad spectrum azole that prevents ifi in patients with aml and in patients receiving an immunosuppressive treatment for gvhd. we describe two cases of mucormycosis (cunninghamella bertholletiae) in patients receiving posaconazole prophylaxis. the fi rst received allogeneic haematopoietic stem cell transplantation with reduced-intensity conditioning for myeloma in relapse. because of grade ii cutaneous gvhd, corticosteroids s were added to ciclosporine months later associated with posaconazole prophylaxis. however, the patient developed a digestive gvhd. at this date, cunninghammella bertholletiae was found in bronchioalveolar lavage cultures. amphotericin b was added. the patient died with disseminated infection. autopsy confi rmed multiple pulmonary lesions of mucormycosis. the second patient was hospitalised with aml for induction therapy. posaconazole was introduced on the fi rst day. ten days after, a febrile episode occurred without documentation. liposomial amphotericin b was substituted. five days later, mucormycosis was identifi ed in skin biopsy. despite anti-fungal treatment associating amphotericin b and posaconazole, he died months later with disseminated infection. residual concentrations of posaconazole were assessed retrospectively by hplc, using sera conserved at a temperature of °c (therapeutic residual plasma concentration: . and mg/l). for the fi rst patient, the serum concentration was below detection threshold (< . mg/l). for the second patient, two sera were collected at prophylaxis and curative treatments ( . and . mg/l, respectively). in both cases, the pathogens were susceptible to posaconazole (in vitro minimal inhibitory concentrations values). our second patient had probably been imunocompromised for several months (long-lasting neutropenia preceding the onset of aml, and history of diabetes). our fi rst patient had an intestinal gvhd with major diarrhoea, which was likely responsible for the very low (undetectable) levels measured when mucormycosis was diagnosed. in conclusion, our report stresses out the necessity to closely evaluate the use of broad spectrum prophylactic antifungal therapy. the prophylaxis in patients with gvhd and/or diarrhea must be used with caution. we recommend to systematically monitor posaconazole levels at least in these cases. inhalation of mold spores can lead in immunocompromised patients to an invasive disease and pneumonia. invasive fungal infection (ifi) has still a high mortality rate. mold-dna can be detected by a polymerase chain reaction (pcr) based method. using it for the bronchio-alveolar lavage (bal) can help to detect an ifi in an early stage. the pcr can discriminate between different mold species and directs the treatment. in our study on patients, a mold pcr from bal was conducted in addition to routine diagnostics. the pcr with primers specifi c for mitochondrial aspergillus-dna and ribosomal s dna for zygomycetes. our results show that mold pcr is more sensitive than standard fungal diagnostics. based on these pcr results, an intensifi ed therapy was undertaken successfully. hence, mold pcr from bal is a useful additon of the microbiological investigations. the mould pcr allows the proof of a zygomycosis at an early stage and thereby ensures successful treatment. further investigations are to show if computer-tomography of the lung combined with mold pcr are suffi cient to diagnose for sure a pulmonal mold infection. introduction: cartilage hair hypoplasia (chh)is a rare autosomal recessive disorder caused by mutations in the ribonuclease rna-processing rmrp complex. hsct has resulted in immune restoration, yet fails to correct the chondrodysplasia. we describe a patient with chh and combined immune deficiency who developed granulomatous infl ammation. treatment with anti-tnf-alpha monoclonal antibodies (moab) caused reactivation of jc virus with ensuing progressive multifocal leukoencephalopathy (pml). case report: at age y a female chh patient ( c>t and a-g mutation in rmrp) with combined immune defi ciency developed painful non-caseating granulomas. no infectious agent was identifi ed and antibiotic therapies failed. finally at age y anti-tnf-á moab(infl iximab) was started with partial response. after the rd administration she developed a debilitating intentional tremor of the right hand. mri t and flair showed demyelination in the right cerebellum. jc virus pcr was (+)in blood and in cerebrospinal fl uid (csf) and (pml) was diagnosed. weekly administrations of cidofovir, followed by two-weekly administrations for month resulted in a partial response. cidofovir was continued two-weekly. months after diagnosis of pml, hsct with a / unrelated donor was performed with reduced intensity conditioning according to ebmt-esid guidelines. there was neutrophil engraftment at d+ and stable donor chimerism of > % at d+ . at d+ , the patient complained of dizziness, with evidence of a cerebellar syndrome. mri and csf polyoma virus copies were stable. at d+ , she presented with hypertensive encephalopathy including convulsions reminiscent of posterior reversible encephalopathy. discontinuation of ciclosporine led to resolution of the encephalopathy. however, pml progressed despite restoration of t cell function, with increasing cerebellar and brain stem symptoms including ataxia, dysarthria, aphasia, n. facialis and n. glossopharyngeus paralysis with corresponding mri imaging and increase in jc virus pcr copies in the csf. despite intensifi cation of cidofovir treatment, trials of steroids, fl uoroquinolones, mirtazapine, lefl unomide as well as high dose ivig and cytarabine iv, the neurodegeneration was progressive and the patient died of respiratory failure at d+ . conclusion: we describe the fatal course of pml due to jc virus reactivation in a patient with chh, despite successful hsct in terms of myeloid engraftment and restoration of t cell function. a. tomaszewska ( ), b. nasilowska-adamska ( ), t. dzieciatkowski ( ), b. marianska ( ) ( introduction: viral infections still are a serious diagnostic and therapeutic problem in patients undergoing alternative donor transplants. betaherpesviruses (hhv , hhv , hhv ) are recognized pathogens in this group of patients. we report a case of hhv encephalitis complicated by guillain-barré syndrome (gbs) in a hematopoietic stem cell transplant (hsct) recipient with preceding reactivation of cmv infection. methods: a year-old-man with a history of chronic myeloid leukemia underwent hsct from a matched unrelated female donor in october . sero-status for cmv was igg positive in the recipient and igg negative in his donor. on the day + patient developed acute graft-versus-host disease successfully treated with iv methylprednisolone. in march he was admitted to our unit due to cmv infection reactivation. he started pre-emptive therapy with iv gancyclovir. after weeks of treatment he revealed high fever, uroschesis, paraparesis, impaired consciousness and generalized epileptic seizure. computed tomography of his brain was normal. a lumbar puncture revealed pleocytosis ( /µl) and elevated level of protein ( . mg/dl). investigation of cerebrospinal fl uid (csf) by pcr for infective causes of patient's neurological decline including hsv t. / , vzv, adenovirus, cmv and dna candida and aspergillus were negative as well as csf culture, real-time pcr revealed in his csf presence of hhv dna. according to these fi ndings and neurological status of our patient we made a diagnosis of an hhv encephalitis complicated by gbs. the therapy with foscarnet (all symptoms revealed during pre-emptive therapy with gancyclovir) and ivig was started. due to gbs diagnosis we performed procedures of plasmapheresis. we observed gradual improvement in neurological status. after discharging home the therapy was continued with cidofovir given once a week during four weeks. at present, . year after this episode, the patient remains in a good condition without cmv and hhv reactivation, with slight neurological defi ciency. conclusions: betaherpesviruses are emerging pathogens in the hsct setting and may cause central nervous system disease. gbs is a very rare complication among stem cell transplant recipients and usually has been attributed to infection. our successfully diagnosed and treated case of hhv neuroinfection complicated by gbs suggests that hsct recipients with cns signs and symptoms should have their csf investigated for hhv as well as other pathogens. zygomycosis is a rapidly growing systemic fungal infection, commonly fatal, despite intensive antifungal treatment. it almost always occurs among patients with an immunosuppressive background, diabetes mellitus, prolonged neutropenia, recent chemotherapy and an excessive iron overload. iron is essential for the growth, development and virulence of many fungi, and particularly of the zygomycetes, which are incapable to grow under iron-deprived conditions. we report on a -year old male patient, who at the age of was diagnosed with cd + b-cell acute lymphoblastic leukemia and achieved a cr following chemotherapy of hyper-cvad type. the patient remained relapse-free for almost years, but when he relapsed, he was treated with the g-mall protocol and a second cr was obtained after cycles of treatment. at that point a fully matched related pbsc allograft, obtained from his -year old sister was offered. he engrafted on day+ , and the post-transplant period was complicated by cmv reactivation and mild chronic gvhd. the patient relapsed on day+ and he was treated with high dose cytosine arabinoside days - and -h infusional mitoxandrone on days + and + . during the aplastic phase he was complicated by histologically proven, extensive left rhinocerebral and pulmonary zygomycosis, with left facial nerve paresis. at that time point he had a transferring saturation of % and ferritin ng/ml. the patient was refractory to initial treatment was surgical debridement and a combination of liposomal amphotericin-b and posaconazole. since no signifi cant improvement was obtained despite a second surgical intervention, deferasirox mg/kg of body weight was added to his antifungal regimen. following weeks of treatment with the triple combination fever was rapidly subsided, as did both, nasal and facial symptoms and lesions. the pulmonary lesions were clearly improved. transferrin saturation decreased to % and ferritin to ng/ml. unfortunately, chemotherapy produced a minor response and months later leukemia reappeared. the patient fi nally succumbed from pulmonary hemorrhage, following salvage treatment with clofarabine and cyclophosphamide, without any sign or symptom of recurrence of his previous zygomycosis. introduction: despite the relatively high transplant-related mortality (trm), the management of the end-life care is poorly understood issue and the problems of providing palliative care to patients submitted to stem cell transplantation (sct) may be underestimated. in this regard, the use of palliative sedation therapy (pst) in the sct setting remains a major concern. patients (pts) and methods: in order to address this issue, a retrospective study on the use of pst in our tertiary sct unit was performed. search criteria were: death and previous sct. data regarding symptoms, symptoms control and use of pst were collected. we identifi ed dead pts. last line of therapy before death was sct and a salvage treatment given for a post sct relapse in and patients respectively. near the death, / patients experienced a total of refractory symptoms and in cases more than one of them was present. intractable symptoms were: excruciating dyspnoea in ( %), agitated delirium in ( %), severe pain in ( %) and massive bleeding in ( %). results: pst was started in all patients, at a median of ( - ) days before death. the most used sedative drug was midazolam, that was administered to / pts as single agent and in cases in association with promazine; pt received the latter agent alone. at the start of pst, pts with pain were receiving parenteral morphine. symptoms control was adequate in cases (complete and partial symptoms control in and respectively) and not adequate in . conclusion: pst is a controversial issue in palliative medicine, although it has been clearly claimed that when it has the intent to provide symptom relief, pst should be considered a proportionate intervention. sct failure represents a so strongly discouraging event to determine diffi culties to recognize end life status. as a consequence, the risk of an inadequate symptoms assessment and of an inappropriate palliation should be considered. in our experience, in a patient closed to the death, when other treatments failed to relieve the intolerable suffering from refractory and otherwise intractable symptoms, pst represented a valid palliative care option by a reduction in patient consciousness, using appropriate drugs carefully titrated to the patient's comfort. adequate symtom control was obtained in more than % ( / ) of pts. an internal operative protocol is under construction to improve those results. donor lymphocyte infusion as therapy for persistent pure red cell aplasia following major abo-incompatible stem cell transplantation a. lübking, i. winqvist, s. lenhoff lund university (lund, se) pure red cell aplasia (prca) after abo-mismatched allogeneic stem cell transplantation (sct) is not uncommon. however, spontaneous remissions within months are frequent. we here report a case of long-lasting prca refractory to multiple therapies that eventually responded to donor lymphocyte infusion (dli). a year old woman received peripheral blood cells from an unrelated hla-identical donor following myeloablative conditioning six months after diagnosis of aml. there was a major abo incompatibility between recipient ( +) and donor (b+). engraftment of granulocytes (> , x /l) and platelets (> x /l) was noted on day and respectively. due to the absence of reticulocytes, bone marrow analysis was performed on day showing the total absence of erythroid precursors. initial treatment with steroids, erythropoetin and withdrawal of immunosuppressive therapy was not successful. four doses of rituximab were given from day without any effect. starting on day immunoabsorbtion on three consecutive days was performed followed by methylprednisolone, cyclophosphamide and immunoglobulin infusions. although the igg and igm antidonor isoagglutinins were reduced from : to : and : respectively, the prca persisted. from day she received doses of dli within months in escalating doses ( , , and million cd /kg). three months after last dli she developed signs of a mucosal gvhd accompanied by moderate eosinophilia. concomitantly, stable reticulocytosis occurred from day and she became transfusion independent. since residual recipient b-and plasma cells are presumed to be responsible for production of anti-donor-isoagglutinins causing prca, inducing gvhd by withdrawal of immunosuppressive therapy or dli might be a reasonable option. there are previously published cases of successful dli treatment for prca, but in many cases dli was given relatively shortly after transplantation, i.e. when spontaneous remission still was possible and the time between dli and reappearance of reticulocytes varied. in our case stable reappearance of reticulocytes occurred concomitant with signs of gvhd. we therefore fi nd our case highly suggestive of that inducing gvhd with dli can overcome post-sct prca refractory to almost all other therapy options. cystatin c level as a marker of renal function in haematopoietic stem cell transplantation h. muto, k. ohashi, m. ando, r. hanajiri, t. kikuchi, w. munakata, c. sakurai, m. yamamoto, t. kobayashi, t. yamashita, h. akiyama, h. sakamaki tokyo metropolitan komagome hospital (tokyo, jp) hematopoietic stem cell transplantation (hsct) recipients have an increased risk of acute kidney injury (aki) or chronic kidney disease (ckd). however, serum creatinine level may underestimate the prevalence of these renal complications because of decreased lean body mass or concurrent liver disease, which was frequently observed in a hsct setting. cystatin c measurement may be more sensitive for detecting impaired kidney function. we retrospectively reviewed the medical records of hsct ( allogeneic and autologous) recipients who had at least one chance to monitor serum cystatin c level during last years in our institution, and evaluated cystatin c as a possible new marker which can predict subsequent renal dysfunction. the occurrence of aki was defi ned by the rifle classifi cation and ckd staging was based on kdoqi criteria. of transplant recipients, patients developed aki after median days (range - days) after hsct, while worsening ckd stage was observed in patients during observational periods. cystatin c level was not infl uenced by autologous transplant (p= . ), but signifi cantly elevated after allogeneic transplantation (p< . ). pretransplant advanced disease status also had an infl uence on cystatin c level before transplantation (p= . ) multivariate analysis disclosed that the use of calcineurin inhibitor was a major cause of cystatin c elevation (odds ratio . , p= . ). there was also a strong inverse correlation between cystatin c and estimated gfr (r=- . , p< . ). proportional hazard modeling analysis revealed that the episode of aki after transplantation were a great risk for substantially worsening ckd stage (hazard ratio . , p< . ). cystatin c measurement could be a useful clinical tool to identify hsct recipient at increased risk for ckd. control of severe bleeding from acute gvhd by treatment with tranexamic acid j. hasenkamp ( ) acute graft-versus-host disease (agvhd) is a major cause of morbidity and mortality after allogeneic hematopoietic stem cell transplantation. % of the cases with intestinal agvhd are refractory to standard treatment regimen. these patients suffer frequently from severe agvhd grades to including massive gastro-intestinal bleedings. we report from clinical courses of two cases treated with tranexamic acid for diffuse, life-threatening gastro-intestinal bleedings caused by steroid-refractory agvhd. the agvhd was confi rmed by biopsy and histopathology. immunosuppression consisted of tacrolimus, mycophenolate mofetil, prednisolone and second line treatment with alemtuzumab. one patient received additionally extra-corporal photopheresis and mesenchymal stem cells. global coagulation and factor xiii plasma levels were kept in normal ranges by substitution. thrombocytopenias were compensated by adequate transfusion of cell separated thrombocytes concentrates. bloody stool volumes of and kg in h lead to dropped hemoglobin levels despite massive transfusion of erythrocyte concentrates. because of this persistent, diffuse gastro-intestinal bleedings, both patients were treated additionally with mg tranexamic acid i.v. every h. after three infusions of tranexamic acid the bleedings in both patients stopped. treatment with tranexamic acid was discontinued without reoccurrence of the bleedings. there were no adverse events of tranexamic acid observed. local hyperfi brinolysis in the gastro-intestinum may contribute to bleedings from tissue damage caused by agvhd. tranexamic acid is indicated for prophylaxis and treatment of bleedings by systemic and local hyperfi brinolysis after e.g. surgery or plasminogen activator treatment. abortion of hyperfi brinolysis can contribute to stabilization of coagulation. prophylaxis or control of severe agvhd is preferred for prevention of hemorrhage. however, tranexamic acid is a treatment option in otherwise unmanageable gastro-intestinal bleeding caused by agvhd. further studies are desired to charge the signifi cance of tranexamic acid in this indication. a low or high body mass index is not predictive for outcome following allogeneic haematopoietic stem cell transplantation j. auberger, j. clausen, b. kircher, g. gastl, d. nachbaur innsbruck medical university (innsbruck, at) objectives: recently it was hypothesized that a low (< ) bodymass index (bmi) is signifi cantly correlated with an increased transplant-related mortality, decreased survival and relapsefree survival after allogeneic sct (k le blanc, haematologica ; : ). patients: patients receiving a fi rst allogeneic transplant were studied. underlying diagnoses were acute myeloid leukemia (aml) (n= ), acute lymphoblastic leukemia (all) (n= ), lymphoma (n= ), and other diseases (n= ). median patient age at time of transplant was (range, - ) years. patients were grafted from an hla-identical sibling donor and patients received grafts from volunteer unrelated donors. conditioning was myeloablative in and of reduced intensity in the remaining patients. results: overall survival for the entire cohort was % ( %- %, % confi dence interval, ci). there was a trend for a poorer outcome in patients with < % and > % percentile bmi (i.e. bmi ≤ and ≥ ) (os % vs %, p= . log rank test) due to a higher non-relapse mortality in this patient cohort ( % vs. %). these differences were observed in both, the myeloablative as well as reduced intensity transplant cohorts. the bmi had no infl uence on relapse incidence in either patient cohort. conclusion: by deviding patients into percentiles bmi had no signifi cant impact on outcome and non relapse mortality neither following myeloablative nor following reduced intensity allogeneic stem cell transplantation. autoimmune thyroiditis after haplo-identical stem cell transplantation for severe combined immunodefi ciency f. dogu ( ) introduction: thyroid dysfunction is a well known complication in survivors of hematopoietic stem cell transplantation, and is reported after tbi as well as radiation-free conditioning. the most common disorders after radiation free conditioning are euthyroid sick syndrome(ets) and compansated hypothyroidism. autoimmune thyroiditis is rarely reported after hsct in children and it has never been described after hsct for scid. here we report an autoimmune thyroiditis developed months after the third haploidentical stem cell transplantation for scid. case: a -months-old girl was referred to clinic with the diagnosis of t-b-nk+ scid. as she didn't have a fully matched sibling donor and her clinical condition was unstable she received peripheral blood stem cell transplantation(pbsct) from his haploidentical father after cd + cell selection without conditioning. engraftment wasn't achieved on day + and she received second haploidentical cd + selected pbsct from her mother. third transplantation was performed months after the second one, due to graft failure and this time she received bu/cyclo for conditioning and csa for gvhd prophylaxis. myeloid and platelet engraftments were achieved on day+ and + respectively. grade i acute gvhd developed on + and treated with corticosteroid for ten days. she was discharged on day+ with full donor chimerism. thyroid hormone levels which were normal before hsct revealed compansated hypothyroidism at posttransplant months in a routine follow-up visit. elevated antithyroid proxidase ( iu/ml) and anti-thyroglobulin ( iu/ml) titers were all consistent with the diagnosis of autoimmune thyroiditis(hashimoto). levothyroxin treatment was started. since the thyroid hormone levels were normal and antithyroid antibodies were negative in her mother, the transfer of autoimmune disorder was excluded. conclusion: regular screening of thyroid functions is important and necessary to detect and treat thyroid illness, especially in young children following hsct. once-daily intravenous busulfan as myeloablative reduced-toxicity conditioning regimen in haematopoietic stem cell transplantation s. santarone, e. di bartolomeo, p. bavaro, p. di carlo, p. olioso, g. papalinetti, p. di bartolomeo bmt center (pescara, it) postulating favorable antileukemic effect with reduced toxicity and improved safety, we used i.v. busulfan (bu) associated with either cyclophosphamide (cy) or fl udarabine (flu) as conditioning therapy for hematopoietic stem cell transplantation (hsct) in patients affected by aml (n= ), mds (n= ), all (n= ) and thalassemia major (n= ) between may and june . patient age was - (median ) years. five of them were older than years. nine patients received flu at a dose of mg/m /day for days (from - to - ) immediately followed by bu given in single i.v. administration over hours at a dose of , mg/kg day for days (total dose , mg/kg). five patients received the same dosage of bu from day - to day - followed by cy mg/kg/day from - to - . donors were hlaidentical (n= ) or antigen mismatched siblings (n= ) and were matched unrelated (mud). the graft-versus-host disease (gvhd) prophylaxis included cyclosporine and short course methotrexate for all patients with the addition of antithymocyte globulin for the mud transplants. eleven patients received bone marrow cells (median dose of nucleated cells , x / kg, range , - , ) and were given peripheral blood stem cells (median dose , x /kg cd + cells, range , - , ). all patients achieved primary engraftment. the median time to . x /l neutrophils and x /l platelets was (range, - ) and days (range, - ) respectively. chimerism studies revealed that of were complete chimeras ( % donor) at year post-hsct. acute gvhd was observed in patients (grade i in , grade ii in , grade iii in ). two patients had mild to moderate chronic gvhd. there was no death due to the transplant procedure. the transplant-related complications were limited. grade iii who hepatic toxicity occurred in patients, hemorrhagic cystitis in , moderate oral mucositis in and a single episode of seizures in . six patients developed cmv reactivation between day and post-transplant (median, day ). two patients relapsed and died. as of december , patients ( %) are alive and disease-free after a median follow-up of days (range, - ) . although the small number of patients does not permit any fi nal conclusion, our hsct protocol treatment confi rms that i.v. bu, associated either with flu or cy, is a well tolerated reduced-toxicity myeloablative conditioning regimen and deserves further study with more patients and longer follow-up. background: down's syndrome (ds) is associated with higher incidence of both haematological and non haematological neoplastic diseases, if compared with general population. reduced susceptibility to chemo-and radiotherapy and the frequent comorbidities limit the use of high dose treatments, especially required in adult patients. case report: a year old male with ds developed an acute myelogenous leukaemia, fab m , aml /eto rearranged, in september . he received standard induction treatment obtaining complete remission (cr), consolidation therapy and, in february , autologuos transplantation with bu-mel conditioning regimen using mobilized peripheral blood stem cells. patient relapsed in february , at the age of , was treated with mec schedule, obtaining a second cr. hla typing showed the presence of an identical sibling. a full clinical evaluation revealed mild reduction of the ejection fraction due to corrected congenital fallot tetralogy (ef= %) and a pulmonary hypertension. a reduced intensity conditioning regimen was proposed, consisting of thiotepa ( mg/kg iv /d x dd) and fludarabine ( mg/kg iv /d x dd) followed by allogeneic stem cell reinfusion in june . standard gvdh prophylaxis was given; engraftment was achieved at day + for anc > , x /l and + for plt > x /l; grade who was recorded for liver toxicity and grade for mucosal toxicity. full donor chimerism was documented at day + . the patient developed stage agvhd; however, months after transplantation relapse was diagnosed with immunological features of all. immunosoppression was suspended, although blast percentage increased rapidly to % and a salvage therapy with all active drugs was started. discussion and conclusion: few data are reported on allogeneic stem cell transplantation in adult patients with ds. this is the fi rst case of ric allosct in a ds adult. those sporadic data do not allow conclusions about outcome on ds adult. a retrospective analysis on large database and a prospective study would be useful to address this issue, helping physicians on treating adult ds pts, when immunogenic effect of allosct play a crucial role to prevent relapse. severe immune hemolysis and pure red cell aplasia after haploidentical non-myeloablative allogeneic stem cell transplantation g. nair, a. mischo, g. stüssi, u. schanz university hospital (zurich, ch) haploidentical stem cell transplantation (sct) offers potential cure to patients without hla-identical donor. recently nonmyeloablative conditioning regimens with in-vivo t-cell depletion have been introduced. severe immune hemolysis rarely occurs after hla-identical sct, but little is known about the occurrence after haploidentical sct. here, we describe patients receiving haploidentical sct for high-risk or relapsed aml ( ), cml after blast crises ( ), and as a rescue therapy in a patient with all after primary graft failure following hla-identical mud sct. all patients were in morphological remission at the time of sct. conditioning regimen included fl udarabine ( mg/qm x days), cyclophosphamide ( mg/qm x days), and alemtuzumab ( mg x days) for in-vivo t-cell depletion. gvhd prophylaxis comprised mycophenolate mofetil (day - ) and cyclosporine (day - - ) and all patients received prophylactic antibiotic treatment. g-csf was administered until hematologic recovery. peripheral blood sct was performed over - days with a median number of . x ( . - . ) cd positive cells. the early posttransplant course was uneventful, the median time of aplasia was ( - ) days. acute gvhd occurred in / patients (i: ; ii: ; iv: ). four patients experienced posttransplant infectious complications ( cmv, bk, fungal infections, one pulmonary infection). two patients experienced severe immune hemolytic anemia and concomitant pure red cell aplasia in the bone marrow and months after sct. in both patients relapse was diagnosed shortly before or after the onset of hemolysis. the direct antiglobulin test was positive for igg and c d. the serum of both patients reacted with all cells in a cell antibody search panel without evidence for cold-reacting antibodies and no antibody specifi city could be evaluated. one patient was treated with steroids, ivig, rituximab, and high-dose cyclophosphamide, but eventually died due to fatal hemolysis. the second patient is currently being treated with steroids, ivig and high dose cyclophospamide with a marked reduction of hemolytic activity. the remaining three patients are currently in complete remission without evidence of hemolysis. in conclusion, nonmyeloablative conditioning regimens in haploidentical sct offer new possibilities for patients without a hla-identical donor. however, physicians should be aware of the potentially fatal complication of severe immune hemolysis. one of the major side effects poorly tolerated, especially in children, is represented by emesis post-chemotherapy. the use of antiemetic during chemotherapy (three to four doses for day) is necessary to reduce this complication. in this work was evaluated using a single dose of palonosetron intravenous for the prevention of nausea and vomiting secondary to chemotherapies. methods: since we have used the palonosetron in pediatric patients of which males and females, undergoing bone marrow transplantation, allogeneic (both sibling that mud) and autologous. the median age is years (range - ) and the median weight is kg (range - kg). the diseases in young patients are reported in table , the conditioning transplantation are listed in table . the dosage used, including scientifi c literature data, was mcg /kg body weight. the palonosetron was considered effective when the emesis was not more than episodes in hours and nausea no more than nd grade. results: it was encouraging, having achieved a good control of nausea and/or vomiting induced by chemotherapy, in fact, only seven patients ( %) was necessary to resort to a second dose of antiemetic, in four of seven ( % of total) was repeated the success with palonosetron a distance of four days after the fi rst dose using the same dosage. in patients ( %) has not been no emetic episode while in the remaining group ( %) episodes were occasional and not have needed any treatment. in all patients was not noted any adverse event or side effect. conclusions: our experience, although on a small sample, it suggests that palonosetron can be considered an effective drug in preventing the nausea induced by chemotherapy, is also a drug that not have adverse events, so well-tolerated and easily manageable, it is necessary a single dose within hours before the start of chemotherapy, not least the assessment of the reduction in costs compared to conventional antiemetic. allogeneic or autologous haematopoietic stem cell transplantation (hsct) is an established mode of treatment of different diseases. loss of protective immunity to pathogens has been consistently demonstrated in patients referred to hsct. impairment of humoral and cell-mediated immunity is commonly seen after transplantation. the degree of immunodefi ciency is determined by many factors, particularly by the type of disease and transplant, the presence of graft-versus-host disease (gvhd) or ongoing immunosuppressive treatment. the aim of the study was to evaluate ) immunogenicity of a revaccination schedule in pediatric hsct recipients ) quality of recipient immune reconstitution and protection against ordinary pathogens. patients and methods: twenty one patients (pts) . - (average . ) years old, boys and girls after autologous ( , %) and allogeneic ( , %) hsct were included in revaccination program. indications to hsct were: solid tumors - , hematological malignancies - , immunodefi cency states - and aplastic anemia pts. time interval between hsct and begining of vaccination protocol was . - (av. . ) years. vaccines used in protocol were as follows: diphtheria and tetanus toxoids, pertussis (for patients < years old), hbv, vzv, haemophilus infl uenzae type b conjugate, -valent pneumococcal polysaccharide, inactivated infl uenza, inactivated polio and attenuated measles-mumps-rubella vaccines. plasma samples to determine specifi c antibodies by elisa tests were collected before and after vaccinations. results: with the exception of one patients presented with repeated fevers, lymph nodes enlargement, muscles and joints pain, no important side effects of vaccinations were observed. a meningococcial meningitis developed in one patient who refused vaccinations. plasma antibody concentrations before and after vaccinations were as follows: antidiphteria ( - , mean . ; - , mean ), antitetanous ( - , mean ; - , mean ) and antihbv ( - , mean ; - , mean ) iu/ml. conclusions: ) systemic immunization is necessary at appropriate time intervals following transplantation to re-establish immunity. ) a signifi cant increase of antibodies titer after hbv, diphtheria and tetanus toxoids was detected. ) vaccinations in patients after hsct are effi cient and well tolerated. ) a delay in begining of vaccination can result in life threatening complications. ministry of science rp, grant number /g/ . according to the world bank data, released in the report, romania has an upper-middle-income economy. the hematopoietic stem cell transplantation (hsct) program started in romania in and more than transplants (auto and allo) were performed. we analyzed the outcome for patients who underwent an allogeneic hematopoietic stem cell transplantation from matched related donor for acute leukemia ( patients) and aplastic anemia ( patients). for of the patients the procedure was performed in romania and for patients abroad. for both categories the follow-up after transplant was done in hematology units in romania. the overall survival was . months, with the longest survival of months and respectively shortest outcome for less than one month. on the st november , there were patients alive, between and months from the procedure, with a median survival of months. sixteen patients died, the median survival being months after transplant. four out of patients died during the fi rst month after transplant, and a total of patients died during the fi rst months after transplant. the transplant related mortality was . %, . % died due to relapsed disease and . % died of graft failure. for these results, there could be incriminated the irregular and inadequate drugs and reagents supplies in the romanian health system, an ineffi cient follow-up system and registry and home-care facilities defi ciencies in romania. in conclusion, the gross national income (gni) per capita and the human development index (hdi) are very important factors for the outcome of recipients of hematopoietic stem cell. background: umbilical cord blood stem cell transplantation has many advantages over bone marrow transplantation or peripheral blood stem cell transplantation. but, there are some problems to be solved in order to be applied to adults. the main problem is limitation of volume, which can be collected from one placenta was only between ml and ml. to overcome this problem, the ex vivo expansion of cryopreserved umbilical cord blood stem cells is needed. the object of this study was to evaluate the effect of cryopreservation on ex vivo expansion potential and viability of umbilical cord blood stem cells. methods: after normal delivery, cord blood was drawn from umbilical cord vein and was used to evaluate the mononuclear cell count, the cell viability and clonogenic capacity of cord blood stem cells before and after cryopreservation. results: before cryopreservation, the mononuclear cell count of umbilical cord blood was . ± . x /ml, cell viability was ± . %, total colony count was . ± . and percentages of cfu-gm, cfu-gemm, bfu-e were . ± . %, . ± . % . ± . %, respectively. the mononuclear cell count of umbilical cord blood cryopreserved for days was . ± . x /ml and cell viability was ± . %. total colony count of umbilical cord blood cryopreserved for days was . ± . and percentages of cfu-gm, cfu-gemm, bfu-e were . ± . %, . ± . %, . ± . %. but, there were few colony count which could be observed after cryopreserving for days. conclusion: there was no difference of clonogenic capacity of umbilical cord blood stem cells before and after cryopreservation. the cell viability of umbilical cord blood stem cells was decreased after cryopreservation but there was no difference between umbilical cord blood cryopreserved for days and days. therefore, it is possible that suffi cient umbilical cord blood stem cells could be obtained by ex vivo expansion of cryopreserved umbilical cord blood in order to be used for adult patient. objective and methods: combined hematopoietic stem cell transplants (hsct) plus solid organ transplants (sot) have been rarely reported. the majority of patients with a previous history of liver transplants were children that underwent hsct for aplastic anemia after viral hepatitis. here we report an adult patient who received a cord blood hsct after a preceding liver transplantation. results: in a year old man required orthotopic liver transplantation for cirrhosis after b viral hepatitis. in april acute myeloid leukaemia m citotype , normal karyotype, flt-itd positive was diagnosed and a fi rst complete remission was reached after induction and consolidation cycles. at that time the patient was not considered eligible for a transplant program due the previous history of sot. in february the patient relapsed and came to our centre: he was treated with high-dose cytosine-arabinoside chemotherapy, that was complicated by a pulmonary aspergillosis , but reached a second complete remission. we decided to start a cord blood donor search, since siblings were not available and he could not wait for an unrelated donor search. a cord blood with hla locus a allelic mismatch and locus c antigenic mismatch was identifi ed. patient's comorbidity index according sorror at transplant was . in may a preparative regimen containing treosulfan, fludarabine and atg fresenius was administered and , x /kg cd + cells were reinfused. grade i mucositis and grade ii hepatoxicity were observed. a bacterial pneumonia and cmv reactivation occurred at day and at day respectively and both rapidly resolved. a neutrophil count > x /l was reached at day and platelet counts > and > x /l platelet count were reached at day and day respectively. no acute and chronic gvhd were observed. a % donor chimerism has been reached in whole peripheral blood and in cd + cells since days onwards. no minimal residual disease has been detected by marrow immunophenotyping and by wt- gene expression until last follow-up, at day . conclusion: to our knowledge this is the fi rst report of a successful cord blood allogeneic hsct in an adult patient with a history of liver transplantation. this case might encourage physicians to propose allogeneic hsct by any stem cell source to patients with high-risk haematological diseases, who had previous liver or other sot's. double unit cord blood transplantation(cbt) has been established as an alternative source of donor cells for allogeneic haematopoietic stem cell transplantation (hsct). we reported here an interesting case of long-term mixed full donor chimerism during the regular follow-ups of one year after hsct. a year-old woman with acute lymphoblastic leukaemia in second complete remission received two units of cord blood after a myelo-ablative conditioning regimen. the cord blood units were hla / identical with the recipient ( b mismatches and a+ b mismatches). total nuclear cell doses infused were respectively . x and x , whereas the cd +cells number was identical in the cbus and the cd +cells number was higher (x ) in the fi rst one (table ). neutrophil recovery was observed at day and platelets engraftment at day after cbt. only one event of acute graft versus host disease (gvhd)grade i was reported at day . currently the patient does not have chronic gvhd and is disease free. analysis of chimerism was performed by str-pcr or rq-pcr on whole blood and specifi c lineage cells (cd +, cd + and cd +). follow-up was done at , , and months post transplant. full donor chimerism (fd) was achieved on day . each of the two units contributed at different levels to the donor chimerism in specifi c lineage cells: whole blood and cd were about % cbu /cbu , cd cells were preferentially from cbu origin ( %), and cd cells were preferentially from cbu origin ( %). this mixed origin of donor cells was detected early and was constant during regular follow-ups. usually, recipients of double unit cbt were engrafted predominantly with one of the units after months. the mechanism of a long-term mixed full donor chimerism is still unknown for our patient. kir ligand analysis showed an absence of mismatch in gvh direction between recipient (c -c ) and each cb unit (c -c and c -c ) while there was a mismatch between the units. in this case, a state of full tolerance settled down between the various lineages, either immune mediated interaction between host/graft or between graft/graft could explain s this chimerism pattern, but it will have to be clarifi ed: a specifi c study of treg cells is in progress. optimising cd yields in pbsch: a comparative analysis of mobilisation regimens c. black, t. elston, m. streetly, m. kazmi guy's hospital (london, uk) cyclo/g-csf (cyclophosphamide/ granulocyte-colony stimulating factor) has been the mobilisation regime of choice when collecting peripheral blood stem cells (pbscs) for transplantation yet pbsc harvests post chemotherapy produce effi cacious yields. this data seeks to compare and inform current mobilisation strategies in this centre. dhap (p= . , and ara-c (p= . , t-test) therapy yielded signifi cantly better cd results compared to cyclophosphamide. mm patients mean cd for cyclo mobilisation (n= ) were . x /kg (range: . - . ), g-csf only (n= ) . x /kg (range: . - . ), and ara-c ( n= ) . x /kg ( . - . ). myeloma patients post ara-c yielded signifi cantly more cd + cells (p= . ) compared to cyclo than those mobilised with g-csf only. nhl patients mean cd harvest results for cyclo mobilisation (n= ) were . x /kg (range: . - . ), g-csf only (n= ) . x /kg (range: . - . ), and dhap (n= ) . x /kg (range: . - . ). cd yield of nhl patients mobilised with cyclo compared with those harvested post dhap, a signifi cantly higher harvest result was noted (p= . ) than those mobilised with g-csf only (p= . ). paired mm data (n= ) compared patients fi rst mobilised with cyclo-gsf and post ara-c, (p= . , paired t-test). this study suggests that harvesting of patients post ara-c, or post dhap is valuable, giving greater cd yields than traditional agents and should be considered. paired data also indicates that ara-c could be used for effective second mobilisation. can type of delivery infl uence cord blood units' quality? g. pucci, a. pontari , d. marcuccio, i. bova, r. monteleone, d. princi, g. gallo, a. dattola, e. spiniello, c. garreffa, t. moscato, p. iacopino ao bianchi melacrino morelli (reggio calabria, it) the cord blood banks use the total nucleated cell (tnc) number as principle to proceed or not to cryopreservation of the cord blood (cb) units. we know that tnc and cd -positive cells infused on unrelated umbilical cord blood transplantation in haematological disease are fundamental for the engraftment of haematopoietic stem cells (hsc) background: immunomagnetic cd + selection is a procedure used both for autologous grafts to perform cellular purging and for allogenic transplant. in aploidentic transplant the purpose of cd + is to reduce the quantity of cd + and cd + cells so as to reduce the incidence rate of the graft versus the host disease (gvhd). aims: in this study we have valued the purity and the cellular recovery after immunomagnetic selection performed with clini-macs automatic system (miltenyi biotec, germany); a group of concentrates has been selected after incubation manually performed, while another group has been submitted to incubation and to the subsequent washings using an automated system (cytomate (baxter oncology, chicago il). methods: in our study we subjected peripheral blood stem cells (pbsc) concentrates taken from donors with microcythemia to immunomagnetic cd + selection, in order to perform aploidentic grafts on children affected by beta-thalassemia major. concentrates have been submitted to washings pre and postincubation and to incubation using the automatic system cytomate, while concentrates have manually been worked. cell count of nucleated cells (nc) was performed using an electronic cell counter while cd +, cd +, and cd + were quantifi ed using fl ow cytometry. results: the following table shows the results. conclusion: immunomagnetic selection in microcythemic donors determines according to our experience, a less recovery in comparison to the data reported in literature, nevertheless in our study results evident, even though casuistry is not very ample, that the use of an automatic system for the washing and the incubation of the cellular concentrates has determined a greater recovery and a greater purity in comparison to the procedures manually performed. allogeneic transplantation from hla identical family donor is a common therapeutic approach in patients with intermediate risk aml in fi rst cr. we present an unusual onset of acute leukemia in a female patient that was a healthy donor of pbsc (previously mobilized with g-csf mcg/kg) for her hla identical sister with diagnosis aml (fab-m ). the transplantation was preformed in january with myeloablative bu-cy conditioning and conventional cy+mtx gvhd prophylaxis. at day + acute gvhd gr i/ii was observed and resolved with addition of median dose of corticosteroids. in a period of years after this occasion, acute leukemia (aml-m no cytogenetic abnormalities) was diagnosed in the donor and treatment with chemotherapy was started. the induction chemotherapy was provided with dae regimen. with consecutive cycles cr was achieved. the patient followed consolidation treatment with hd-arac and anthracikline. further treatment with allogeneic transplantation was on schedule and the source of stem cells would be taken under consideration. can a person with aml and years surviving in a complete remission become a donor of its own donor (hla dna identical) with the same diagnosis is a question that has to be resolved in a higher number of patients. clinical outcome and characteristics of donor graft failure in patients with haematopoietic disease given donor cell boost, second allogeneic transplantation or no treatment r. ahmed nacer, m. benakli, f. mehdid, r. belhadj, n. rahmoune, m. baazizi, a. talbi, f. kaci, r.m. hamladji pierre and marie curie center (algiers, dz) introduction: donor graft failure (gf) is a life-threatening complication of allogeneic hematopoietc stem cell transplantation(hsct), determined when anc had not reached , /l by day (primary gf) or when anc decreased irreversibly after engrafment (secondary gf). frequency of gf is variable in function to hematopoietic disease. material and methods: from may to december , patients (pts) underwent allogeneic hsct from hla-identical sibling donor. pts are appraisable for this study and gf was diagnosed in pts ( , %) : primary gf: pts, secondary gf: pts with median time engrafment to ; median age at transplant years ( - ); sex ratio (m/f) , ; hematologic non malignant disease (hnmd; n: ): aplastic anemia: / ( , %), major athalassemia: / ( %) and hematologic malignant disease (hmd): / ( , %); pts had received more than transfusions before allograft; abo incompatibility between donor/recipient was seen in d/r pair; median interval from diagnosis to transplant months ; pts was multiparous; pts received myeloablative conditioning regimen (mcr ) and one pt reduced intensity conditioning (ric); pts received peripheral blood stem cell (pbsc) and pts bone marrow transplant (bmt). the chimerism testing was performed in cases: predominant host population (host population: - %) in pts and mixed (host population < %) in pts. pts were given donor cell boost with no additional conditioning with median time gf to treatment days ( - ) and fi ve pts a second hsct within one a third hsct (mcr: ; ric: ) with median time gf to treatment days ( - ). pts had not been treated. at november maximal follow up is months and minimal months. results: pts are alive ( %) within pts (donor cell boost: pts; second hsct: pt) with success engraftment(donor population: %) after median follow up months ( - ) and pts (second hsct: pt; no treatment: pts) with autologous reconstitution (donor population: %). pts died ( %) within pts after given donor cell boost, pts after second hsct and pts before given donor cell boost. os at years is %. conclusion: gf is rare but serious and concern hnmd more than hmd. better outcome can be obtained after chimerism testing study to choice treatment : predominant donor population will have donor cell boost and predominant host population second hsct with another donor. mixed haematopoietic chimerism: how the initial dynamics of mixed chimerism correlate with later chimerism status k assing ( ), c. heilmann ( ) ( )herlev university hospital (herlev, dk); ( )university hospital, rigshospitalet (copenhagen, dk) background: the natural history of mixed (hematopoietic) chimerism (mhc) has been extensively studied in hematopoietic stem cell recipients, in order to fi nd determinants for relapse or graft rejection. methods enabling quantitative prediction of later mhc status have not been devised. methods: recipients, receiving hematopoietic stem cells due to non-clonal disorders and displaying at least % donor chimerism at minimum one time point, were serially tested for whole blood chimerism over a median period of . years (range: . - . years). relative changes in the host fraction (termed alfa ) between the median time points: . and . weeks post-transplantation were correlated with later mhc status. the predictivity of alfa values for later mhc outcome was assessed in a linear regression model. findings: all recipients engrafted. subsequently, . % became mixed chimera and . % achieved complete donor chimerism. weekly chimerism fl uctuations prior to six months posttransplantation ( . % points; . - . % points) exceeded those after six months time ( . % points; . - . % points, p< . ). at seventeen weeks, alfa values correlated with endpoint mhc levels at . years (r = . , p< . ). negative alfa values predicted ( % confi dence intervals) the presence of less than % host cells, while alfa values between: . - . , were predictive of mhc with ≤ % host cells. the only recipient experiencing rejection ( . %) displayed the largest alfa value and had a predicted mhc outcome of . % host cells ( % ci: . - . %). interpretation: we have devised a simple mathematical method enabling us, early post-transplantation, to predict later mhc status and thus determine at an early time point, where intervention is needed in order to prevent rejection or poor graft function. feasibility of out-patient autologous stem cell transplantation for malignant haematologic disorders a. ghavamzadeh, a. allahyari, k. alimoghaddam, a. karimi, r. aboulhasani, a. manookian, m. asadi, a.r. shamshiri hematology-oncology and sct research center (tehran, ir) introduction: high-dose chemotherapy with autologous stem cell support is utilized for the treatment of a variety of malignancies including non-hodgkin's lymphoma, hodgkin's lymphoma, and acute leukemias. the aim of this study was to explore the feasibility and safety of performing autologous stem cell transplantation (asct) on an out-patient basis. material and methods: total of patients affected by malignant hematologic disorders( cases of hl, cases of nhl, cases of aml) with median age of y (range : - y) and in complete remission and without medical problem were selected. they received conditioning regimen (ceam for nhl and hl, busulfan and etoposide for aml) and stem cell infusion in hospital. the day after sct, patients were discharged and followed by outpatient sct team; include a general physician, staff nurse and care giver during their neutropenic period, and to be rehospitalized in the case of febrile neutropenia, after sepsis workup and performing chest x-ray, they were received the fi rst dose of antibiotic in hospital and treatment continued in their home. results: median time for wbc recovery was days (range: - days), median time for plt recovery was days (range: - days), median number of transfused single donor plt was . units (range: - unit). mucusitis grade was seen in patients, median duration of neutropenic fever was days (range: - days), patients was rehospitalized because of the neutropenic fever, median duration of rehospitalization in these patients was days, median follow up of patients was days (range: - days), all patients were alive and in complete remission. conclusion: results show that out-patient autolgous sct in malignant hematologic disorders (hl, nhl, and aml) is feasible and its complication is manageable. haplo-identical sct as a salvage therapy in haematological malignancies: a single-centre experience o. paina, y. stankevich, i. kazantsev, n. stancheva, a. golovacheva, e. babenko, a. alyanskiy, n. ivanova, e. semenova, p. krugliakov, d. polyntzev, l. zoubarovskaya, b. afanasyev spb state i. pavlov medical university (st. petersburg, ru) background: allogeneic hematopoietic stem cell transplantation (allo-hsct) is the one of curative option for patients (pts) with acute leukemias, though its usage is often limited by lack of matched related donor or the time required for search of unrelated one. usage of haploidentical donors allows to avoid these problems and to perform allo-hsct in time. patients and methods: very high risk pts underwent haploidentical sct: all - ( %) pts, aml ( %) pts, jmml - pt, cml- pt, and resistant neuroblastoma- pt. the total number of resistant/in progression pts was ( %), ( %) pts were in remission. ( %) pts were children (age - ), ( %) were adults (age - ). in all cases reduced intensity conditioning regi-mens (ric) were used: fl udarabine and atg with addition of different alkylating agents (busulphan, melphalan or thiotepa). sources of hsc -peripheral blood stem cells (pbsc) and bone marrow. for pbsc cd + positive selection clinimacs was used. the mean cd + count was , x /kg ( , ) . in pts agvhd prophylaxis consisted of csa and short course of mtx with or without mmf. in pts tacrolimus and mmf were used. in pts at d- used mesenchymal stem cells (msc) from third -party donors were used prevention of agvhd, in pts, msc were used for treatment of acute gvhd. results: the incidence and severity of agvhd weren't higher, than in other types of allohsct: ( %) pts had grade iii-iv agvhd with skin and gut involvement, one pt died. when mcs using in conditioning regimen agvhd, i stage was observed. treatment of agvhd with msc was successful: in pts in cr. the toxicity of the conditioning regimen was acceptable, ( %) developed grade ii-iii organ toxicity. ( %) pts had invasive aspergillosis and ( %) pts of them had cmv reactivation. the -year os is %, with mediam observation terms of , months ( to months). pts died in relapse and in cr (infection - pt, another failed to engraft and acute gvhd of the gut). conclusions: haploidentical hsct with ric is characterized by acceptable toxicity and agvhd control, stable engraftment. it proved to be a good option for the group of pts with poor prognosis. randomized clinical trials are necessary for estimation of therapeutic effect of mscs in haploidentical hsct pts. results: there were donation requests involving donors ( females, males). one donor was contacted but declined to donate dli. one donor donated twice for the same recipient. for / donors no details of donation are available. the median age at time of donation was . years (range - years). there were no failed collection procedures. / donors experienced mild citrate toxicity. / donors had a vasovagal episode, but both recovered rapidly and collection was able to be completed. donor required central access for dl collection: she had also previously required central access for pbsc donation. a median . donor blood volumes was processed (range . - . ). no late donor complications were reported. in total, donors had dl collected. among the prospective recipients ( female; male), indications for dl were: mixed chimerism(n= ); residual disease(n= ); molecular relapse(n= ); clinical relapse(n= ); ebv reactivation(n= ); pancytopenia of uncertain cause(n= ); no data(n= ). of patients for whom data were available ( %) actually received dl infusions. for the remaining , reasons for not proceeding were: spontaneous improvement in blood counts; death from ebv; death from relapsed disease; development of gvhd prior to dli; and spontaneous resolution of mixed chimerism. an escalating-dose regimen was used at -monthly intervals depending on response: the median number of doses reinfused was (range - ). patients ( %) developed gvhd s following dli. the dli was successful in treating the stated indication in patients ( %). there were recipient deaths: relapsed disease(n= ), infection(n= ), gvhd(n= ) and progressive ebv(n= ). only the two gvhd deaths were considered dl-related. conclusions: our single-centre experience confi rms that dl are frequently an effective treatment for mixed chimerism or early relapse post-hsc transplant, and that donor experiences are generally good. although requirement for dl is itself an adverse prognostic factor following hsc transplant, % of recipients had a successful outcome. nowadays, haematopoietic stem cell transplantation (hsct) remains the single curative approach to the treatment patients (pts) with the resistant primary and secondary aml. these pts have extremely poor prognosis with the level of relapse at least % and the risk of trm - %. as known, high level of blasts to the moment of transplantation infl uences on dfs and os. patients and methods: at the russian children research hospital between october and june hsct were made in refractory aml pts ( m/ f). the median age was ( - ) years. fab-type: m - pts, m - pts, m - pt, m - pts, m - pts, m - pts, mx - pts. primary refractory aml was diagnosed in pts and secondary refractory -in pts. kids were transplanted from msd, pts -from mud ( mmud) and pts -from mmfd ( haplo-pbsc) with the usage of cd /cd -depletion ( pts) or cd -selection ( pt) of the graft. the median level of blasts in bone marrow prior to hsct was % ( % - %). the myeloablative conditioning regimens were used in hsct and non-myeloablative regimen -in second hsct. pts received double-phase conditioning regimens. a median dose of cd ±cells was ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) x /kg. pts received dli on median day ( - ); pts received prophylactic dlis and pts received treated dlis due to increasing of mrd level or the mixed chimerism. results: the engraftment level was % with a median time to neutrophil recovery days ( - ) and to platelets recovery - days ( - ). % pts achieved cr to day , pts had the progression of disease, one pt died before engraftment, and the rejection was documented in pt. acute gvhd developed in pts ( %), chronic gvhd -in ( %) from pts who were alive to day after hsct. pts had the liver toxicity grade - and two pts had the pulmonary toxicity grade - (who-classifi cation). trm level was %. relapse was diagnosed in pts (dfs %). at this time pts are alive in cr, pts died ( -from relapse, -from gvhd and sepsis and one pt -from dag). os for all pts was % with a median time months ( - ) . but after double-phase conditioning regimen os and dfs were % both and trm was %. overall rfs depended on the presence of gvhd, the type of donor, the using of dli. conclusion: our results show that even for very high risk aml pts hsct may be performed successfully without the significant increasing of trm particularly with prophylactic dlis. a.a. hamidieh, a. ghavamzadeh, m. jahani, k. alimoghaddam, a. mosavi, m. iravani, b. bahar, a. khodabandeh, m. jalili hematology-oncology and sct research center (tehran, ir) objective: hematopoietic stem cell transplantation (hsct) has been extensively used in the treatment of pediatric leukemia. this is follow-up report of pediatric patients (< years) with acute myeloid leukemia (aml) and acute lymphoblastic leukemia (all) whom transplanted in hematology-oncology and stem cell transplantation research center, tehran, iran. methods: pediatric patients boys and girls (median age= years) with acute leukemia ( patients with aml and patients with all) received hsct between and . the most common conditioning regimen was cyclophosphamide + busulfan. they have received allogeneic ( aml/ all) or autologous ( aml/ all) hsct from bone marrow ( aml/ all), peripheral blood ( aml/ all) or cord blood ( aml). donor type for allogeneic transplantation in patients( aml/ all) were stem cell from hla matched siblings, patients ( aml/ all) received from other related (hla-matched confi rms with high resolution method) and patients(aml) from other related with one or more than one antigen mismatch. prophylaxis regimen for graft-versushost disease (gvhd) was cyclosporine a and methotrexate or cyclosporine a alone. results: ( . %) patients are alive, ( . %) patients died ( aml/ all). the most common cause of death was relapse of disease in . %. among patients who received allogeneic transplantation acute gvhd occurred in % and chronic gvhd in . %. two years overall survival and disease free survival of aml patients were % and % respectively. two years overall survival and disease free survival of all patients were % and % respectively. no statistical difference between aml group and all. conclusions: our results of overall survival and disease free survival are compatible with literatures. autologous haematopoietic stem cell transplantation with busulfan and etoposide as conditioning regimen for acute myelogenous leukaemia patients s. mousavi, k. alimoghaddam, f. khatami, m. jahani, m. iravani, b. bahar, a. khodabandeh, a. jalali, a. ghavamzadeh hematology-oncology and stem cell transplantation research center (tehran, ir) introduction: acute myelogenous leukemia(aml) is a potentially lethal disease. hematopoietic stem cell transplantation(hsct) has increased disease free survival (dfs) and overall survival (os) of patients more than conventional treatment. the result of autologous hsct in patients without suitable donor is near to allogeneic transplantation. we performed autologous transplantation with busulfan and etoposide as conditioning regimen for patients who didn't have suitable donor. methods: since january until oct , patients received autologous transplantation. we included all children and adult with aml in fi rst or second complete remission without suitable donor and end organ failure who can tolerate high dose chemotherapy. mobilization regimen was cyclophosphamide g/m² for one day and g-csf µg/kg for days. we have done stem cell harvesting when patient's white blood cell (wbc) count raised to /µl then the patients received oral busulfan mg/kg(from - to - ) and etoposide mg/kgiv (from - to - ) as conditioning regimen. after that patients have transplanted with their peripheral blood stem cells. results: median age at time of transplantation was . years (age range: - ), male/female: / . patients were in fi rst complete remission and patients were in second complete remission. median infused mononuclear cells was . * /kg despite of the progress in the treatment of acute myeloid leukemia allogeneic hematopoietic stem cell transplantation (hsct) remains the single curative approach to the treatment of resistant aml. these patients (pts) have extremely poor prognosis with the level of relapse - % and the risk of trm - %. high level blasts to the moment of transplantation infl uences on dfs and os. the usage double-phase conditioning regimens with the aim to reduce the level of blasts maximally to the transplantation date can improve dfs and os without the elevation of trm. patients and methods: we reviewed the records of refractory aml pts ( m/ f) who underwent hsct at the russian children research hospital between october and june . the median age was ( - ) years. fab-type: m - pts, m - pts, m - pts, m - pts, mx - pt and secondary aml - pt. primary refractory aml was diagnosed in pts and secondary refractory -in pts. kids were transplanted from msd, pts -from mud, pt -from mmud and pts -from mmfd with cd /cd -depletion of graft. the median level of blasts in bone marrow prior to hsct was % ( - ). first phase included flam ( pts), flag/go ( pts), ham ( pts), flae ( pt), go ( pt) or dacogen ( pt). mylotarg doses were - mg/m². the interval between -st and -nd phase of conditioning protracted - days. second phase of conditioning was treosulfan-( pts) or bu-based ( pts) . a median dose of cd ±cells was , x /kg. seven patients received dli on median day ( - ). results: engraftment was documented in all patients with a median time to neutrophil recovery days ( - ) and to platelets recovery - days . patients achieved cr to day and one patients had pr. acute gvhd developed in patients, chronic gvhd -in from patients who were alive to day . patients had the liver toxicity grade - , and one patient had the pulmonary toxicity grade - . trm level was , %. relapses were diagnosed in patients (dfs %). nowadays pts are alive in cr, pt has bm-relapse, pts died ( -of the relapse, pt -of viral infection, pt -of vod). os amounted % with a median time months. rfs depended on the presence of gvhd and type of donor. conclusion: our results show that the usage of double-phase conditioning regimen generally doesn't increase the level of toxicity and trm and allows to achieve the long-term survival in pts with very high risk aml. allogeneic haematopoietic stem cell transplantation for acute myelogenous leukaemia in other than fi rst complete remission status k. alimoghaddam, f. khatami, a. jalali, m. jahani, s. mousavi, m. iravani, a. khodabandeh, b. bahar, a. alimohammadi, n. bahar, a. ghavamzadeh hematology-oncology and stem cell transplantation research center (tehran, ir) introduction: for patients with aml in whom an initial remission cannot be achieved or for those who have a relapse after chemotherapy, stem cell transplantation from an hla-identical sibling offers the best chance for cure. this study reports the result of years allogeneic hsct for aml in other than fi rst complete remission status. patients and methods: from march until november , aml patients, male and female with median age of years old (range: - yrs) received allogeneic hsct. the status of them before transplantation was other than cr (including second cr, third or more cr, relapse and primary induction failure). source of hematopoietic stem cells was peripheral blood, bone marrow, and cord blood. result: median time to absolute neutrophil count ≥ . * l was days post hsct and median time to platelet count ≥ * l was days post hsct. forty-two recipients developed acute graft versus host disease (gvhd) and twenty-four developed chronic gvhd. at present ( %) patients are alive. the most common cause of death was relapse. median follow up period was month (range: - month). six month disease free survival (dfs) and overall survival (os) were % (se= %) and % (se= %), respectively. year dfs and os were % & % (se= %). conclusion: allogeneic hsct for aml in other than fi rst complete remission could be advice and can improve the result of treatment in these high-risk patients. outcome of haematopoietic stem cell transplantation for patients with acquired aplastic anaemia at a cancer center, amman, jordan: experience of a young hsct program in a developing country f. abdel-rahman, i. al-sadi, a. badeeb, h. el taani, a. ahmed, r. rihani, a. al zaben, m. sarhan king hussein cancer center (amman, jo) purpose: to evaluate the outcome of hsct in patients with acquired aplastic anemia at khcc. patients and methods: between ( / - / ) , patients had allogenic hsct for aplastic anemia. there were adults ( %), and children ( %), with a median age of years (range: - years). there were patients ( %) with severe aplastic anemia and patients ( %) with very severe disease. the source of stem cells was bone marrow in patients ( %), and peripheral blood in patients ( %). the median time from diagnosis to transplantation was days. among the group, patients had a full hla matched-related donor, one had / matched related donor and had / donor. the conditioning regimens were cyclophosphamide +antithymocyte globulin (atg) in patients, and different conditioning in the other patients. results: the main end points of the study are overall survival for the whole group, and overall survival according to the age, severity of the disease, occurrence of graft versus host disease, and degree of hla match. the median duration of follow up was . months ( . - . months) . the median time for the wbc engraftment was engrafted the wbc or the platelets, and patients never engrafted the platelets. the median survival for the whole group was . months. from the patient, patients are still alive. from the deaths, patients died from sepsis and one from massive gi-bleeding secondary to gut gvhd. from the adult patients are alive ( %), while from the pediatric patients are alive ( %) from the patients with severe aplastic anemia are alive while from the patients with very severe disease are alive. from the patients who had transplant from / hla matched related donor, are alive ( %). the three other patients who received mismatched graft died. acute gvhd was associated with increased mortality. six of nine patients who develop gvhd died while only out of patients who did not develop gvhd died. four patients had second transplant, two of them are still alive. conclusion: the important predictors of the outcome are: -degree of hla match: survival % in / hla match, versus % for the mismatch transplant. -occurrence of gvhd: survival is % in patients without gvhd, and % in patients with gvhd. therefore, the most important factor for predicting survival is the degree of hla match. our plan is not to transplant aa from mismatched donors except according to an international study protocol. ( ) diabetes type is caused by immune destruction of insulinproducing b cells of pancreas. it has recently been shown that immunoablation combined with transplantation of autologous hematopoietic stem cells may alter the course of the disease and alleviate exogenous insulin requirement [voltarelli et al. jama, ; : - . we report a year old patient with an early diabetes type (typical clinical course, presence anti-gad antibodies, diagnosis weeks prior to study inclusion) with sustained presence of c-peptide in the blood, in good clinical condition without other serious comorbidities who has been chosen for treatment after signing informed consent for study protocol earlier accepted by local bioethics committee. treatment consisted of plasmapheresis followed by mobilization with cyclophospamide ( g/m²) and granulocyte colony stimulating factor (g-csf) analogues at µg/kg from day + . three x cd + cells/ kg were obtained by leukapheresis and were later used for transplantation without further selection. conditioning regiment consisted of cyclophosphamide ( mg/kg for days - to - each) with atg (thymoglobuline , g /kg over days - to - ). and was followed by transfusion of collected peripheral blood cells on day . results: the transplantation was performed on the . . . patient engrafted on day + . during the cytopenic period no major complications were observed. the patient insulin requirement was: , iu/kg -before moblization, , iu/kg on the transplantation day, , iu/kg on engraftment. insulin was discontinued shortly after the regeneration (+ ). glucose monitoring showed normal glucose levels without the need for insulin injections from that day on. hba c levels at diagnosis were , %, , % after months from transplantation and , % months after the transplantation. continuous glucose monitoring was performed around months after the transplant and showed normal values (glucose - mg/dl) -fi gure . intravenous glucose tolerance test showed normal values of glucose levels after minutes, however the st phase of insulin secretion was not present . conclusion: this case support the notion that immunoablation followed by autologous stem cell transplantation in patients with early diabetes type may at least temporary alleviate insulin requirement with excellent control of glycemia. introduction: the allogeneic stem cell transplantation (hsct) represent an effective curative treatment in cml but treatmentrelated morbidity and mortality can be substantial. with the era of bcr-abl kinase inhibitor the place of sct is in discuss for children. we report the results of myeloablative allogeneic hsct underwent in patients (pts) under years of age. material and methods: from december to april ( months period) pts under years of age with cml (chronic phase: , accelerated phase: ) underwent myeloablative allogeneic hsct from hla-identical sibling donors; median age at transplant years ( - ); sex ratio m/f , ; median interval from diagnosis to transplant months ( to ). all patients received chemotherapy based conditioning regimen: tutshka (n: ), tutshka with additional vp (n : ) and santos (n: ). pts received peripheral blood stem cell with median cd + cell , /kg body weight (bw), pts bone marrow transplant with median nuclear cell ( nc ) , /kg bw, one pt blood cord transplant with , nc/ kg bw. graft-versus-host disease (gvhd) prophylaxis consisted of association ciclosporin and methotrexate. the molecular bcr-abl transcripts diagnosis concerned pts (m(b a ): , m(b a ): and double transcripts m (b a ; b a ): ). molecular monitoring of disease using real-time quantitative polymerase chain reaction (rq-pcr) concerned pts. at july maximal follow-up is months and minimal months. results: the median time of aplasia was days ( - ). eighteen pts ( %) are alive in complete hematological remission (complete molecular remission: ; major molecular remission: ; no evaluated: ) after median follow-up time months . acute gvhd occurred in pts ( %) with grade ii-iv, and chronic gvhd in pts ( , %) with extensive. disease relapse occurred in pts ( %) within are in complete remission with imatinib. seven pts died ( %): acute gvhd grade iv (n : , trm: %) ; relapse (n: ; %). the os and event free survival (efs) at years are respectively % and %. conclusion: our results confi rm that trm is low in young pts and the mayor problem is still relapse disease. the relapse after graft can be treated with bcr-abl kinase inhibitor ( pts in our study). the question about using allogeneic hsct or bcr-abl kinase inhibitor in children with cml is still open. reduced-intensity conditioning allogeneic stem cell transplantation in advanced chronic lymphocytic leukaemia. the impact of conditioning regimen on the non-relapse mortality. a single-centre experience j. el-cheikh, c. oudin, l. wang, c. faucher, s. furst, d. blaise institut paoli calmettes (marseille, fr) purpose: a unicentric retrospective study to determine the transplant related toxicity in patients with advanced chronic lymphocytic leukemia (cll) after reduced intensity conditioning hematopoietic stem-cell transplantation (hsct) including or not antithymoglobuline (atg). patients and methods: we studied patients with progressive or relapsing chronic lymphocytic leukemia (cll) treated with hematopoietic stem cell transplantation (hsct) in our cancer centre of marseille. males and females, (median age: years). all patients received a reduced intensity conditioning regimen. we compared patients ( %) receiving a non myeloabaltive conditioning including atg with fl udarabine, busulfan (atg group) to patients ( %) receiving fl udarabine, total body irradiation (tbi gy) and anti cd without atg (non atg group). patients ( %) had a matched related and patients ( %) a matched unrelated donors. graft-versus-host disease (gvhd) prophylaxis consisted of cyclosporine alone in the atg group or a combination with mycophenolate mofetil (mmf) in the non atg group. results: after a median follow-up of months, patients ( %) still alive and in complete remission (to date). mrd was monitored in those patients with cr; all patients achieved a molecular cr. patients had acute and/or chronic gvhd, ( % in atg group vs % in non-atg group). at the last follow up patients died ( %), and the cause of death in all of them was the treatment related complications (infections and/ or gvhd); the trm at days was %; % at one year and % at three years of transplantation. ( % in atg group vs % in the non atg group); overall survival (os) at three years was % in the atg group vs % in the non atg group. the os at one and three years was % and % respectively. fig . in conclusion: despite the small effective, we can conclude that hsct after reduced conditioning is effective and has the capacity to induce a long term complete remissions, the real impact of atg should be revaluated on further large multicentric studies. dasatinib: optimal bridge to stem cell transplant in chronic myeloid leukaemia blast crisis a. gozzini, s. guidi, c. nozzoli, b. bartolozzi, r. saccardi, b. scappini, a. bosi bmt unit (florence, it) pts presenting cml-bc have a survival of - months and scarce response to imatinib. dasatinib (bms- ) is an oral, multi-targeted kinase inhibitor, currently being used in pts with imatinib-resistant advanced cml or relapsed/refractory ph+ all. most of these pts will be evaluated for sct, even though for them this curative therapy showed higher incidence of gvhd, vod and trm. we report here fi ve pts affected from cml-lb who received dasatinib prior to allosct. donors were matched siblings ( ), matched unrelated ( ) or blood cord unit ( ) . were male and female with a median age of , ( - ) years. first line therapies included chemotherapy (vcr) plus high dose imatinib. all pts after - months from diagnosis received dasatinib mg bid. t i mutation occurred in patients, y and e k in patients, and a non codifi ed mutation in patient. dasatinib induced complete hematological response (chr) in pts, and complete (n= ) and partial cytogenetic response (pcyr) (n= ) prior to sct. patients did not achieved a complete haematological response presenting % marrow blasts and % respectively prior to sct. all pts were conditioned with myeloablative protocol. gvhd prophylaxis consisted of csa and mtx (n= ) or micofenolate association until + (n= ). pts received a mobilized peripheral blood stem cell graft with , - , x cd + cells/kg (n= ) and cord blood unit with , x cd + cells/kg (n= ). dasatinib was stopped days before transplant procedure. / pts successfully engrafted reaching anc > . x /l on day + ( - ) and plt > x /l on day + ( - ). dasatinib was introduced again in patients days after sct. one of them stopped therapy because of haematological toxicity after weeks. / patients presented chimerism was - %. transplant related toxicities were grade i/ii. no pts developed hyperbilirubinemia or vod. hyperacute extensive gvhd (gr iii) was observed in only pts at + . five patients are alive, all of them in complete molecular response with a median follow-up of . ( - ) months, died of agvhd. we may conclude that in pts undergoing sct following dasatinib there is no evidence of adverse effect on sct outcome, organ toxicities. larger studies and longer follow-up are obviously indicated to confi rm our preliminary results. both t i positive patients are alive in chr. dasatinib represents an effi cient bridge to transplant to improve the outcome of this subset of patients. inmatinib combined myeloablative allogenetic haematopoietic stem cell transplantation for advanced chronic myeloid leukaemia y. luo, y. tan, j. shi, x. han, g. zheng, x. zhu, x. lai, h. huang zhejiang uninversity school of medicine (hangzhou, cn) improved strategies are needed to treat patients with advanced chronic myeloid leukemia (cml) in order to reduce the need for lifelong therapy. we treated patients with advanced cml ( in ap, in bc) with myeloablative allogeneic stem cell transplantation (allo-sct) combined with pre-transplantation imatinib. the donors included hla-matched and -locus mismatched unrelated volunteers (n= ), and hla-matched siblings (n= ). graft-versus-host disease (gvhd) prophylaxis consisted of cyclosporine, mycophenolate mofetil and short-term methotrexate. out of ( . %) evaluable patientsdeveloped ii-iv agvhd, ( . %) patients suffered from agvhd grade iii-iv. two patients suffered from intensive chronic gvhd. after a median follow-up of months (range ¨c months), the overall survival was ( . %) / . the ten patients were all in molecular remission. imatinib combined with allo sct could provide a safe, well-tolerated therapeutic option for patients with advanced cml. this conclusion needs to be tested in prospective randomized clinical trials. p-beam group. both groups did not differ in terms of time of hospital stay, days of iv antibiotics, mucositis and infections. with the median follow-up of ( - ) years, the probability of overall survival at years equaled % for p-beam and % for chopp-cbv group (p= . ). the probability of progressionfree survival was % and %, respectively (p= . ). conclusions: p-beam and chopp-cbv protocols followed by autopbsct are effective and well-tolerated salvage therapies for patients with advanced hl. prolonged administration of the therapy seems to be appropriate for this group of patients. towards safer autotransplants in patients with non-hodgkin's lymphoma: cardiac pre-evaluation, angiotensin-converting enzyme inhibition in patients with decreased left ventricular function, antimicrobial prophylaxis and vigilant supportive care e. jantunen, s. hämäläinen, t. kuittinen, k. penttilä, m. pyörälä, a. juutilainen, i. koivula, t. nousiainen kuopio university hospital (kuopio, fi) autologous stem cell transplantation (asct) for nhl is associated with an early non-relapse mortality rate of - % most commonly due to sepsis. during - nhl patients received asct at our department. seventeen patients ( %) experienced severe sepsis and nine ( . %) died due to septic shock. severe sepsis was caused by gram-negative bacteria including pseudomonas in a signifi cant proportion of the patients (hämäläinen et al. scand j infect dis ). subclinical anthracycline cardiomyopathy may be important in regard to the development of severe sepsis in some nhl patients. since january we have applied prospectively cardiac pre-evaluation (radiocardiography), angiotensin converting enzyme inhibition in patients with decreased left ventricular ejection fraction (lvef) (< %), ciprofl oxacin prophylaxis and start with ceftazidime plus tobramycin in patients with neutropenic fever in nhl patients undergoing asct. febrile patients are observed closely with measurements of pro-brain type natriuretic peptide (bnp) and c-reactive protein (crp) for three days. also blood pressure, blood oxygen saturation, hydration and diuresis are monitored. until nov , altogether patients with nhl ( m, f) with a median age of years (range - ) have received beam followed by pb infusion according to this protocol. lvef was < % in six patients ( %) pre-transplant and they received enalapril during the peritransplant period. neutropenic fever was observed in patients ( %). no cases with gram-negative bactereamia or severe sepsis have been observed. the median peak crp value was mg/l ( - ) and was reached in a median of two days after rise of fever. serum bnp values were above normal limit in / patients with fever on day . elevated bnp values were observed in / patients on day , in / patients on day , and in / patients on day , respectively. whether severe sepsis or early deaths could be prevented with this approach remains to be seen in upcoming years with larger number of patients. outcome of refractory/relapsed patients affected by hodgkin's lymphoma treated with or without peripheral blood stem cells autografting: a single-centre experience f. angrilli, s. falorio, f. fioritoni, s. santarone civic hospital (pescara, it) introduction: despite a high curability rate, to % of patients (pts) affected by hodgkin lymphoma (hl) fail to respond or relapse after front-line treatment with polychemotherapy alone or combined with radiotherapy. the treatment of choice for refractory or early relapsed pts is high-dose chemotherapy (hdc) followed by peripheral blood stem cells autografting (pbsca), while late relapsed pts may be treated with either conventional therapy or hdc plus pbsca. methods: from to december , untreated pts with hl have been admitted in our institution. after front-line therapy, ( %) pts obtained a complete remission (cr) and pts ( %) were refractory to standard treatment. overall, pts relapsed within months after diagnosis of hl, while pts experienced late relapse. the aim of this retrospective study is to evaluate the outcome of the our refractory/relapsed pts according to the type of salvage treatment. twenty-six pts received as salvage treatment - courses of igev (iphosphamide, gemcytabine, vinorelbine), patient courses of coppebvcad (cyclophosphamide, carmustine, melphalan, epirubicin, vinvristine, vinblastine, prednisone) and patient courses of abvd (doxorubicin, bleomycin, vincristine, dacarbazine). today, pts completed salvage chemotherapy. of them, ( with refractory hl and with relapsed disease) have been submitted to pbsca. conditioning regimen consisted of beam in all cases. results: pts were male and female (m/f ratio , ). median age was years (range - ). overall, pts obtained a cr ( %) and pts had progressive disease ( %). in particular the cr were ( %) in the group of the pts receiving pbsca and ( %) in the other pts (p < . ). one patient died in cr of beam toxicity prior pbsca and pts died of progressive hl. after a medium follow-up of months, overall survival was % for the pts who received pbsca and % for those who received conventional treatment (p= . ). conclusions: our data confi rm the benefi t of hdc plus pbsca both in relapsed and in refractory patients with hl. nevertheless, a portion of refractory or early relapsed pts fail to respond to pbsca and died of hl. for these pts tandem pbsca or allogeneic stem cell transplantation should be proposed, especially if they are not in cr prior to pbsca. t cell lymphoma is a heterogeneous group of aggressive lymphomas associated with poor prognosis with standard chemotherapy and autologous hematopoietic progenitor cells transplantation (hpct) is offered as consolidation in fi rst remission or at relapse. in this study we conducted a retrospective analisis of patients underwent hpct from december to august . seven patients had diagnosis of peripheral t-cell lymphoma, four patients of systemic anaplastic large cell, and fi ve patients of linfoblastic lymphoma. five patients were transplanted in fi rst complete or partial response, ten patients in second or beyond complete or partial response and one patients in second refractory disease. median age was , years; seventy-fi ve percent preesented advanced (iii-iv) ann arbor stage, % had b symptoms, % had high lactate dehydrogenase. with a median follow-up of months from diagnosis and , months from transplantation, the -year progression-free survival (pfs) and overall survival (os) were , % and , % respectively. based on these preliminary results the hpct as consolidation therapy may offer a durable survival benefi t. the chimeric anti-cd monoclonal antibody rituximab offers new therapeutic options in the treatment of b-cell nhl (non-hodgkin's lymphoma). the addition of rituximab to chop (cyclophosphamide, doxorubicin, vincristine, and prednisolone) or cvp (cyclophosphamide, vincristine, and prednisolone) regimen was found to signifi cantly improve the response rates and survival in patients with untreated diffuse large b-cell lymphoma (dlbcl) and is now considered as the standard therapy option. rituximab also has been shown to improve response rates when combined with salvage chemotherapy. there are few studies regarding the effects of rituximab on mobilization. we compared the effi cacy of rituximab plus eshap (etoposide, metil prednisolone, cytosine arabinoside, cisplatin) with eshap alone as mobilization regimen in ( %) hodgkin's and ( %) non-hodgkin's lymphoma patients. ( %) patients were dlbcl. ( %) relapsed and ( %) refractory patients were involved. ( %) patients were treated with r-eshap and ( %) patients with eshap regimen. aphaeresis were evaluated. median number of aphaeresis was . days for r-eshap patients and . days for eshap patients. median number of mononuclear cell aphaeresis was . * per kg (kilogram) and . * per kg respectively. total number of cd + cells was . * per kg in the r-eshap group and . * per kg in the eshap group. toxicities were similar in both groups. there were no engraftment delays in the r-eshap group. so we conclude that r-eshap is effective and feasible as eshap regimen for mobilization. total number of cd + cell aphaeresis was slightly lower in the r-eshap group but did not have an effect on engraftment. prospective randomized studies are needed to evaluate whether rituximab really decreases mobilization adequacy or not. no benefi t of autologous stem cell transplantation as consolidation for high and high-intermediate risk diffuse large b-cell lymphoma in .cr after r-chop therapy -a single-centre experience m. karas, k. steinerová, p. jindra, d. lysák, s. vokurka, v. vozobulová, m. schützová, l. mohammadová, v. koza charles university hospital pilsen (pilsen, cz) objectives: the role of high-dose therapy (hdt) and autologous stem cell transplantation (asct) for patients (pts) with high and high-intermediate (h/hi) risk diffuse large b-cell lymphoma (dlbcl) in .cr was not clearly defi ned especially after addition of rituximab (r) to fi rst line chemotherapy (cht) and the use of rituximab also as maintenance therapy. therefore, we retrospectively analysed outcome of pts treated in our transplant centre with hdt and asct for h/hi risk dlbcl in .cr after - cycles of r-chop- chemotherapy and we compared their outcome with a control group of pts with h/hi risk dlbcl in .cr treated only with chemoimmunotherapy. patients and methods: between and (median follow-up months, range - months) consecutive pts with median of age years (range - years) with h/hi risk dlbcl in .cr after - cycles of r-chop- underwent hdt (beam) and asct. the median of time from diagnosis to asct was months (range - months). source of stem cells was peripheral blood and median of infused cd + cells was , x /kg (range , - , x /kg). the control group consisted of consecutive pts with h/hi risk dlbcl in .cr treated only with chemoimmunotherapy ( - cycles of r-chop- , % maintenance therapy with rituximab). the control group except for the older age did not differ in any prognostic parameters. results: in the transplanted group pts ( %) are alive in cr. pts ( %) relapsed and died. no patient died due to transplant-related mortality (trm). the estimated probabilities of -years disease-free survival (dfs) and overall survival (os) were % and %. in the chemoimmunotherapy treated group pts ( %) are alive in cr. patient ( %) relapsed and died. the estimated probabilies of -years dfs and os were % and %. we did not observe between both groups any significant difference in cumulative relapse incidence (p= , ), dfs (p= , ) and os (p= , ). conclusion: our data suggest that hdt with asct in pts with h/hi risk dlbcl in .cr after r-chop chemotherapy was well-tolerated with no trm death but in comparison with pts treated only with chemoimmunotherapy we did not observe any improvement of outcome among transplanted pts. of course relatively lower number of evaluated pts and retrospective type of analysis could infl uence our results and only prospective randomized studies can fi nally defi ne the role of frontline hdt with asct for h/hi risk dlbcl in .cr after chemoimmunotherapy. kyrcz-krzemien medical university of silesia (katowice, pl) background and aims: autologous peripheral blood stem cell transplantation (autopbsct) is widely used for the treatment of poor-risk patients with hodgkin's lymphoma (hl), however, the optimal preparative regimen has not been established. we assumed that patients with advanced hl may benefi t from receiving intensive pre-transplant therapy with prolonged administration of cytostatics and the addition of oral drugs, such as procarbazine or chlorambucil. therefore, we modifi ed the commonly used beam and cbv protocols by incorporating oral agents and prolonging the distribution of the total doses to and days, respectively. the goal of this pilot study was to evaluate safety and effi cacy of those regimens. patients and methods: patients ( males and females, median age years, range - ) with relapsed hl were included in this study. previous therapy consisted of median ( - ) lines of treatment and ( - ) chemotherapy cycles results: / patients died due to septic complications in chopp-cbv group, whereas no procedure related mortality was observed among patients treated with p-beam. all remaining patients engrafted. time to achievement anc > . g/l was signifi cantly shorter in p-beam vs. chopp-cbv group: ( - ) days vs zevalin-beam conditioning in transformed follicular lymphoma; acceptable toxicity and possible therapeutic benefi t a hdt-sct) using beam conditioning has become standard therapy for relapsed fl, however recurrent disease especially in transformed follicular lymphoma (t-fl) remains the commonest cause of death. the addition of zevalin (ibritumomab tiuxetan), a cd targeted radiolabelled antibody to beam is safe and may improve the effi cacy of hdt we analysed patients aged to with advanced stage t-fl who had received a median of (range - ) lines of therapy prior to zevalin-beam sct. the median time from diagnosis to hdt-sct was months (range - . ) and all patients had chemosensitive disease in partial remission bcnu mg/m , etoposide mg/m , cytarabine mg/m , melphalan mg/m ) from day - to - . the median stem cell dose was the remaining patients remain stable at a median of months (range - ) post-sct conclusion: the zevalin-beam protocol is as well tolerated as standard beam conditioning. the disease free survival in this small cohort of high risk patients with t-fl is encouraging but needs longer follow-up rituximab or not? a historical comparison of eshap and r-eshap as mobilisation regimen in patients non-myeloablative allogeneic stem cell transplantation in patients with high-risk lymphoma: a multicentre experience g. console ( ), g. irrera ( ), m. martino ( ) a. meliadò ( ), c. rigolino ( ), t. del vecchio ( ), o. iacopino ( ), m.c. cannatà ( ), p. scaramozzino ( ), i. bova ( ), d. marcuccio ( ), c. stelitano ( ), s. molica ( ), r. cantaffa ( ), l. nocilli ( ), a. mele ( ) pugliese-ciaccio" (catanzaro, it); ( )osp patients (pts) ( females and males), median age , years (range - ) underwent nst for high risk hodgkin disease (hd, cases) and non hodgkin lymphoma (nhl, cases) conditioning regimens consisted of fludarabine, thiotepa and cyclophosfamide in cases, tli and atg in cases, fludarabine and cyclophosfamide in cases, fludarabine and thiotepa in case, fludarabine, melphalan, thiotepa and atg in case, campath- , fludarabine, melphalan and tbi were employed in cases. in case tbi and fludarabine. cyclosporine-a (cya) and methotrexate (mtx) were used as gvhd prophilaxys in cases, in campath- and moftil micofenolate (mmf) were combined and in cases cya and mmf were used. a mean number of . x /kg cd + cells (range , - , ) were infused. pts received a mean of . (range - ) packed red blood cells after a median follow up of , months, (range - ), pts are still alive ( in cr, pr, in relapse), experienced cgvhd ( cutaneos w.h.o. grade - , pneumonial w.h.o. grade , liver w.h.o ). pts died for liver agvhd, patient died for cerebral vasculitis at months to the transplant, patient died for ards at months from transplant. pts for acute bacterial pneumonia. pts for mof respectively at and months from transplant, for aptt, at months from transplant. patient for interstitial pnemoniae at months to the transplant, pts died for disease recurrence at , and months post-transplant respectively here we report on a single centre, retrospective analysis evaluating the outcome of patients (pts.) with dlcl treated with high-dose chemotherapy and autologous (auto) or autologous-allogeneic (auto-allo) hsct. patients and methods: in total, ( , %) male pts. and ( , %) female pts. with dlcl underwent auto ( pts.) or autoallo hsct ( pts) between . . and . . . pts. received auto hsct as part of fi rst-line therapy (group ). in pts auto ( pts, group ) or auto-allo ( pts., group ) hsct was performed as second-line therapy. the median patient age was years : / / , group : / / , group : / / . patients who received auto hsct as fi rst-line therapy (group ) tended to have a better median os ( vs. d, p= . ), rfs ( vs. d, p= . ) and -ysr-os ( % vs. %, p= . ) compared to pts of group . furthermore patients of group had a signifi cant better os and -yrs-os in the auto-allo group of pts died ( pts died from severe infection with multiorgan failure and patient from relapse of disease). in contrast, none patient died from trm after second auto hsct, but died from progressive disease and pt from relapse. conclusion: the survival of patients with relapse of dlcl could not be improved by using the therapeutic approach of auto-allo hsct compared to an auto hsct based regime, due to the high trm in the auto-allo group. however, for interpretation of these results some facts have to taken into account since year , patients with primary myelofi brosis ( females and males age - y median . ) received allo hsct ( sib and unrelated donors matched at allele level). according to the dupriez prognostic system: , , patients were in high, intermediate an low risk of the disease. the diagnosis was proved by trephine biopsy, which revealed that all patients were at advanced stage of fi brosis, all patients had splenomegaly and abnormal blood smear with the presence of erythroblasts. the length of the disease duration was from to months (median ). six patients were transfusion dependent because of anaemia and thrombocytopenia, three patients were on steroids and six on hydroxycarbamide. splenectomy prior to transplantation was performed in two patients. two patients received myeloablative conditioning (busulfan mg/ kg cyclophosphamide mg/kg) and eight reduced intensity conditioning (busulfan mg/kg, fludarabine - mg/m² or melphalan - mg/m² and low dose atg). all patients were transplanted with pbpc with cd dose from . to . x /kg (median . x /kg). two patients died due to transplant toxicity (one with additional ebv reactivation and sepsis and one with vod symptomatology). in other patients toxicity was mild and there was no agvh exceeding grade i. two patients transplanted with major blood group incompatibility developed prca. plasmapheresis and erythropetin were successfully employed in those patients. finally all surviving patients reconstituted haematologically. a trephine biopsy performed months post transplant documented the process of bone marrow remodeling with a normal picture six months post transplant. all patients except one had full chimerism. eight out of ten patients are alive and with normal hematopoesis during observation period from to months (median ). the post transplantation course was similar in patients having and lacking jak mutation. in conclusion haematopoetic stem cell transplantation in primary osteomyelofi brosis is associated with rather low risk and results in sustained haematological recovery. nutritional assessment of children undergoing haematopoietic stem cell transplantation for primary immunodefi ciency or severe autoimmune disease m. slatter, c. ferguson, e. rogerson, a. yurasova, p. askew, t. flood, m. abinun, a. cant, s. bunn, j a major challenge post hsct is adequate nutrition, as poor nutritional status adversely affects outcome. patients undergoing hsct for pid often fail to thrive pre-hsct due to underlying disease. we aimed to document nutritional intake of pid children undergoing hsct at our centre. children who underwent hsct for pid or severe autoimmune (ai) disease from april -january were evaluated. the following prospective data was collected: diagnosis, age, donor, conditioning, presence of infection and growth. nutritional intake, biochemical indices, use of antiemetics and complications were documented on admission, after weeks, then monthly until day , + , + , + , + , + then monthly until discharge home. patient characteristics: patients had scid, had other pid. had severe ai disease. age at transplant ranged from months to . years (median months). were ≤ yr. had unrelated ( cords), matched family, matched sibling and haploidentical donor. all had chemotherapy conditioning - bu/cy, flu/melph, treo/flu, treo/cy. results: all received supplementary feeding via nasogastric ( ) or percutaneous jejunal tube ( ) . only required total parenteral nutrition, with severe ai disease and with persistent norovirus enteritis. all received at least anti-emetic. had viral enteritis - norovirus, adenovirus. in patients for whom adequate data was available, all had a reduction in calorie and protein intake in the - weeks following hsct, because of fl uid restriction. had grade ii skin gvhd, none developed gut gvhd. had mucositis requiring morphine. only patient lost weight overall from time of admission to discharge, one had static weight, but gained weight, by time of discharge. further evaluation of nutritional indices is required. the time around hsct is the most challenging to support adequate nutrition. careful nutritional assessment of patients undergoing hsct is critical and should direct nutritional support. patients should be optimised nutritionally prior to hsct, as the high metabolic demands around the time of hsct are unlikely to be met over the immediate transplant period. thalidomide+dexamethasone and partly b, b+dexamethasone, b+adriamycin+ dexamethasone (pad). the b group had a post-tx maintenance therapy with b weeks: , mg iv doses weekly + dexamethasone mg days. results: length of survival times (os) without and with b were signifi cantly different. further analysis of the curves in complete remission indicated % survival probability and % disease free survival (dfs) in patients in a -month period. in the very good partial remission (vgpr) group ( pts) the os was %, however, the dfs was only %. the survival curves were signifi cantly worse when tx was made in partial remission (os: %, dfs: % by pts). conclusions: . the author›s data support the fi nding that lasting survival can be expected when tx is performed in cr or vgpr. . in the interest of this, in cases of a more aggressive disease, the fi rst line pad protocol before tx is the best therapy. after the tx a consolidation therapy with b+dexamethasone is very useful. . in a slightly less aggressive disease or with accompanying diseases a thalidomide+dexamethasone fi rst therapy may also be possible. . tx performed in partial remission maybe dangerous. at this time needed put in "the therapy arsenal". acute renal failure in myeloma patients during mobilisation procedures for autologous transplantation a. pivkova ( ) during the last years blood cell separation, generally referred to aphaeresis, has established a central role in both blood donor programmes and therapeutics. the technological advances in aphaeresis equipment have made procedures safer, faster and more effective. we present cases ( males and female) with multiple myeloma treated at our department during until . initial chemotherapy treatment was provided with thalidomide based regimens (c-thal dex cycles or thaldex in cycles) or cycles of vad in one patient. all patients before diagnosis and during initial treatment had normal and stable renal function. after completing remission in all, mobilisation of pbsc was preformed with g-csf mcg/kg in duration of days. the number of wbc count prior collection was median x /l ( - ) with median lymphomonocyte percent , ( - ). aphaeresis was preformed at day with cobe spectra cell separator and large volume aphaeresis. in all patients after fi nishing the fi rst procedure we registered increase of renal degradation products in the serum during the fi rst hours post aphaeresis and complete anuria which revealed in acute renal failure (renal type) treated with haemodialysis in several consecutive occasions. one month after resolving the renal impairment the patients continued with second mobilisation procedure with the same regimen and obtained a minimal mnc count of , x /kg. autologous transplantation followed by melphalan reduced dose conditioning mg/m². engraftment was registered for ne> , x /l and plt > x /l on median day + ( to ). the patients had no need for blood transfusions. all are in cr med mths ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) after transplant. in one patient months after, a double transplant was preformed. concerning the small group of patients, we can evaluate the possible impact of large volume aphaeresis in the renal impairment in these patients or the infl uence of cytokine mobilised cells on renal tubules. key: cord- -zml lbve authors: cuvelier, geoffrey d.e.; rubin, tamar s.; junker, anne; sinha, roona; rosenberg, alan m.; wall, donna a.; schroeder, marlis l. title: clinical presentation, immunologic features, and hematopoietic stem cell transplant outcomes for ikbkb immune deficiency date: - - journal: clin immunol doi: . /j.clim. . . sha: doc_id: cord_uid: zml lbve ikbkb immune deficiency is a rare but life-threatening primary immunodeficiency disorder, involving activation defects in adaptive and innate immunity. we present sixteen cases of a homozygous ikbkb mutation (c. dupg) in infants characterized by early-onset bacterial, viral, fungal and mycobacterial infections. in most cases, t- and b-cells were quantitatively normal, but phenotypically naïve, with severe hypogammaglobulinemia. t-cell receptor excision circles were normal, meaning newborn screening by trec analysis would miss ikbkb cases. although ikbkb immune deficiency does not meet traditional laboratory based definitions for scid, this combined immune deficiency appears to be at least as profound. urgent hsct, performed in eight patients, remains the only known curative therapy, although only three patients are survivors. ongoing infections after transplant remain a concern, and may be due to combinations of poor social determinants of health, secondary graft failure, and failure of hsct to replace non-hematopoietic cells important in immune function and dependent upon ikk/nf-κb pathways. in the canadian province of manitoba, our group has periodically managed young infants of northern cree first nations (aboriginal) descent presenting with early-onset and life-threatening viral, bacterial, mycobacterial, and fungal infections, clinically resembling severe combined immune deficiency (scid). infants were initially identified in the s from two small northern manitoba communities. in later years, infants from the neighboring province of saskatchewan were also identified. in , our group and collaborators identified the genetic defect in four patients as being due to a homozygous duplication mutation (c. dupg) in exon of ikbkb, the gene encoding inhibitor of nuclear factor kappa b kinase subunit beta (ikkβ, also known as ikk ) [ ] . the homozygous ikbkb duplication mutation seen in our population results in complete loss of ikkβ expression, a critical component of the canonical ikk-nuclear factor kappa b (nf-κb) pathway [ ] . normally, signaling through tumor necrosis family receptors, toll-like receptors, and antigen receptors on t-and b-cells induces activation of the ikk complex, which consists of the kinases ikkα and ikkβ, as well as the regulatory protein nuclear factor kappa-b essential modifier (nemo) [ ] . the active ikk complex, and specifically ikkβ, phosphorylates the inhibitory protein iκbα, resulting in release of cytoplasmic nf-κb. free from the influence of iκbα, nf-κb translocates to the nucleus to induce the transcription of a multitude of immune and inflammatory genes. given the central role of the ikk/nf-κb pathway in regulating both innate and adaptive immune responses, it is not surprising that patients with mutations in genes from this pathway manifest with primary immunodeficiency disorders [ ] . males with hypomorphic hemizygous mutations in ikbkg, which encodes for the nemo protein and results in reduced nemo expression, present with a combined immune deficiency that includes enormous allelic, immunologic and clinical heterogeneity [ ] [ ] [ ] [ ] [ ] [ ] . patients with hypermorphic gain of function mutations in nfkbia [ ] [ ] [ ] , resulting in reduced degradation of the iκbα protein, have also been shown to exhibit significant combined immune deficiencies. furthermore, the ikk/nf-κb pathway operates in both hematopoietic and non-hematopoietic cells, leading to the extra-immune findings seen in patients with defects in nemo and iκbα, such as ectodermal dysplasia and colitis. similar to patients with ikbkg and nfkbia mutations, the first four patients reported by our group with deficiency of ikkβ due to the c. dupg ikbkb mutation also presented with immune deficiency [ ] . since then, six additional patients from various ethnic backgrounds (including turkey, qatar, and the arabian peninsula) with different nonsense ikbkb mutations have been reported, with all patients presenting with early onset and profound combined immune deficiencies, but typically without findings of ectodermal dysplasia [ ] [ ] [ ] . in our initial report, patients had normal to near-normal numbers of t-and bcells of an almost exclusively naïve phenotype, severe hypogammaglobulinemia, and activation defects to a variety of stimuli in a number of immune cells, both innate and adaptive, more suggestive of a global functional defect as opposed to a quantitative lymphocyte deficiency [ ] . herein we describe the clinical presentation, immunologic features, and hsct outcomes for the largest cohort of infants with ikbkb immune deficiency resulting from complete loss of ikkβ expression published to date. the study protocol was approved by the health research ethics board of the university of manitoba. the authors identified all known (genetically confirmed) or highly suspected infants with the ikbkb mutation managed through the department of pediatrics, winnipeg children's hospital, cancercare manitoba and the manitoba blood and marrow transplant program dating to the s. data collected by retrospective chart review included clinical presentation, immunologic features, and hsct approaches and outcomes. although data was complete for patients diagnosed in a contemporary era (after ), data availability was variable for some patients before this . young infants of northern cree descent from the known affected communities presenting with a clinical picture consistent with a severe immune deficiency but without genetic confirmation of an ikbkb mutation were considered highly suspected if they also had a close familial relationship to a genetically confirmed infant with the mutation (e.g. sibling, or second-degree relative where there was known consanguinity within the family). we chose to report the decade of diagnosis (instead of year) to provide the reader with perspective on the immunologic investigations and treatments available at the time, while protecting against the potential for inadvertently identifying individuals, given that patients and their families reside in small communities. detailed family trees documenting consanguineous relationships within families have not been included for the same reason. patient laboratory values, including absolute lymphocyte count, cd , cd and cd t-cell counts, cd b-cell counts, cd nk-cell counts, and immunoglobulin levels were compared against published, age-adjusted variables [ , ] . laboratory parameters are described as low, normal, or high with respect to these ranges. maternal engraftment of t-cells was assessed by analysis of short tandem repeats using the ampf str® profiler plus™ pcr amplification kit from life technologies. chimerism analysis after hsct was performed on t-cells, b-cells, and myeloid cells labeled with monoclonal antibodies to cd , cd , and cd / b and separated using an automated immunomagnetic cell separation method with the robosep instrument from stemcell technologies. due to a potential paucity of cells with each chimerism test, purity of the various lymphocyte and myeloid subsets was not evaluated with each assay. from validation experiments, however, the purity of separated subsets is > % with a ± % error around lineage specific chimerism results. t-cell proliferative responses using pha were performed by hthymidine incorporation assay of peripheral blood mononuclear cells after stimulation with pha at : - : following a -day culture. percentage of normal pha response was measured by the stimulation index of the patient relative to the average stimulation index of two concurrently run healthy adult control samples. trec analysis, when performed at the time of diagnosis (n = ), was either done locally in our laboratory (n = ) [ ] or at a clinical laboratory (mayo clinical laboratories) (n = ). units of measure for trecs (copies/μl whole blood, copies/million pbmcs, copies/million cd cells) changed over time. sanger sequencing for the c. dupg in exon of ikbkb was performed locally on a genetic analyzer™ as per the method developed by pannicke [ ] . samples containing homozygous g insertions within the region of interest were classified as affected, and confirmed by sanger sequencing at an independent laboratory. sixteen patients with genetically confirmed (n = ) or highly suspected (n = ) ikbkb mutations were included (table ) . genetic confirmation for the five infants with highly suspected ikbkb immune deficiency was not possible since all had died, and dna was not available (all were born in years before the ikbkb mutation was known). one patient was diagnosed immediately after birth by direct ikbkb mutational analysis of a guthrie card, as part of a targeted newborn screening project started in november in two of the heavily affected communities. this patient's younger sibling was later diagnosed in-utero, due to the known family history, with confirmation of the mutation after birth. both siblings were placed in protective isolation and proceeded to hsct by - months of age. the other fourteen patients presented to health care facilities with severe infections at a median of . months of age (range: weeks - months). twelve patients came from the province of manitoba and four from the province of saskatchewan. four patients were previously reported in less detail in our initial report of ikbkb immune deficiency [ ] . we included these patients in order to describe additional clinical and immunologic characteristics and provide updates on long-term outcomes. details on the number and type of infections present at the time of initial presentation (before hsct) were complete for twelve and incomplete for two (patient who died of disseminated mycobacterium bovis-bacillus calmette-guerin (bcg) and patient who died of unknown sepsis). the two siblings identified by newborn screening and family history had no infections at the time of diagnosis of ikbkb immune deficiency and proceeded to urgent hsct without infections. for patients not identified by newborn screening, clinical presentation consisted of a multitude of severe bacterial, fungal, mycobacterial, and viral infections beginning at an early age (< months), usually in association with failure to thrive. mucocutaneous infection of the oropharynx and perineum with candida was common (n = ) with evidence of disseminated candida infection table clinical presentation and outcome of ikbkb immune deficiency. no. scid was suspected but since ikbkb mutation had not yet been described diagnosis was never confirmed. no. multi-organ failure present. not hct candidate. died after hct in the blood (n = ), urine (n = ), and cerebrospinal fluid (n = ) in three. severe, recurrent gram-negative and gram-positive bacterial infections were frequent and included bacteremia / sepsis episodes with escherichia coli (n = ), pseudomonas aeruginosa (n = ), klebsiella pneumoniae (n = ), enterococcus species (n = ), staphylococcus aureus (n = ), stenotrophomonas maltophilia (n = ), serratia marcescens (n = ), streptococcus pneumoniae (n = ), staphylococcus hominis (n = ), listeria monocytogenes (n = ), morganella morganni (n = ), leuconostoc lactis (n = ) and enterobacter cloacae (n = ). intracranial infections were present in six infants and included a combined staphylococcus aureus and candida albicans meningitis in one, and listeria monocytogenes meningitis, enterococcus meningitis, klebsiella pneumoniae meningitis, serratia marcescens intracerebral abscesses, and mycobacterium bovis meningitis with intracerebral abscesses in the other five. disseminated viral infections occurred with cytomegalovirus (n = ), adenovirus (n = ), varicella (n = ), and herpes simplex virus (n = ). severe pneumonia syndromes were documented in eleven as result of unknown pathogens (n = ), cytomegalovirus (n = ), respiratory syncytial virus (n = ), polymicrobial gram negative bacteria (n = ), adenovirus (n = ), rhinovirus (n = ), and parainfluenza (n = ). consistent with standard practice in the two affected communities at the time, four infants received the bcg vaccination after birth, all of whom developed disseminated bcg and died. biopsies (n = ) and autopsies (n = ) revealed mycobacterium bovis in multiple organs including the lungs, liver, spleen, lymph nodes, thymus, bone marrow, adrenal glands, kidneys, eyes, skin, brain and gastrointestinal tract. following these deaths, bcg vaccination in the two affected communities was discontinued. more recently, with the advent of targeted newborn screening for the homozygous ikbkb mutation in the two affected communities, bcg vaccination after birth is held until results of the newborn screen confirm that an infant is not affected. one patient met clinical and laboratory criteria for hemophagocytic lymphohistiocytosis (hlh), although it is possible that additional patients would have met hlh criteria but were unrecognized. despite the connection of ikbkb to the ikk/nemo complex, none of our patients exhibited ectodermal dysplasia. only one patient had documented organ abnormalities (dysplastic kidney at time of autopsy). of the sixteen ikbkb immune deficiency patients, eight died of overwhelming infection before hsct could be performed, and eight proceeded to hsct. detailed autopsy reports were available for three who died prior to hsct. aside from evidence of infection, all exhibited severe thymic hypoplasia with scant lymphoid cell populations, minimal to no lymph node and tonsillar tissue, and absence of germinal follicles in the spleen. a range of immunologic tests were performed over time, with the extent and type of testing dependent upon the year of presentation and initial suspicion of an immunodeficiency diagnosis. extensive immunologic investigations were more likely to occur in the contemporary era (after ). median total lymphocyte count at the time of diagnosis was . × /l (range: . - . × /l), with / patients exhibiting normal (n = ) to elevated (n = ) total lymphocyte counts for age. the three lymphopenic infants all had severe infections (disseminated bcg in two and klebsiella sepsis in one) at the time their initial blood work was performed, along with various lymphocyte subset and hematologic cytopenias (anemia, thrombocytopenia), more consistent with bone marrow suppression from infection. median cd t-cell count ( patients) was cells/μl (range: - cells/ μl), with only one patient having a cd t-cell count < cells/μl t-cell proliferative responses were available for patients (fig. ). relative to controls, three had pha responses < %, two had pha responses between and %, and eight had pha responses above % (range: % - %). no relation was evident between pha response and total cd t-cell count, although two of the patients with severe cd t-cell lymphopenia and disseminated bcg had no pha response. the two infants identified at birth without infections both had normal pha responses ( % and %). trecs were performed in six infants. all had normal trec expression levels ( copies/ pbmcs; , copies/ cd ; , copies/ cd ; copies/μl; copies/μl; copies/ μl). more extensive t-cell immune phenotyping was performed in three patients. all exhibited a preponderance of naïve cd t-cells (cd + cd ra + cd l + cd +) between %- % of the cd t-cell population, and naïve cd t-cells (cd + cd ra + cd l + cd +) between and % of the cd t-cell population. minimal to no central and effector memory t-cells ( - %) were present in any of the cases. one patient had tcr v-beta repertoire performed, revealing a normal polyclonal t-cell population. eight patients underwent hsct, using either bone marrow from a human leukocyte antigen matched parent or sibling (n = ) or unrelated umbilical cord blood (n = ) as the graft source (table ) . two patients not initially receiving a conditioning regimen (patients and ) due to critical illness from infections both failed to engraft, with a second transplant performed in one using myeloablative conditioning, resulting in subsequent engraftment. conditioning regimens were initially used in the other six. two patients ( and ), one of whom received a reduced intensity conditioning regimen followed by a cord blood transplant, initially engrafted but experienced secondary graft failure over two years post-hsct. four patients received a busulfanbased myeloablative conditioning regimen (with busulfan pharmacokinetic monitoring after ) with either full or mixed donor chimerism, and generally more favorable myeloid engraftment. three patients receiving transplant remain alive -months, -years and -years post-hsct. two patients died soon after hsct (days + and + ) from complications related to infections present at the time of transplant. three others died of new infections acquired post-hsct, including two (patients and ) who died of disseminated mycobacterium avium-intracellulaire infection in association with secondary graft failure at -and -years post-hsct. the third (patient ) died of streptococcus pneumoniae bacteremia and meningitis despite full donor chimerism at -months post-hsct. new clinically relevant infections, including viral, bacterial, fungal, and mycobacterial infections, were common in patients who survived at least -months after transplant, and were still occurring years after transplant despite full and mixed lymphoid and myeloid donor chimerism and normal neutrophil and lymphocyte counts. such infections included streptococcus pneumoniae bacteremia (n = ), meningitis (n = ), osteomyelitis (n = ), and septic arthritis (n = ); staphylococcus aureus bacteremia, cellulitis, and osteomyelitis (n = ); salmonella bacteremia and enterocolitis (n = ); and pseudomonas aeruginosa bacteremia (n = ). viral infections (e.g. influenza, parainfluenza, varicella, adenovirus) also occurred, with resolution in all patients. one patient developed mycobacterial tuberculosis pneumonia at -year post-hsct, was successfully treated with anti-tuberculosis medications, and remains a long-term survivor. all patients achieved normal numbers of cd +, cd +, cd +, cd +, and cd + cells post-hsct. immunoglobulin replacement was variable between patients and generally continued between -and -months post-hsct. pha responses were normal (above %) for all patients. despite these findings, specific antibody production to posttransplant vaccinations once immunoglobulin was discontinued was poor. for instance, patients and did not make anti-pneumococcal fig. . phytohemagglutinin responses at the time of ikbkb diagnosis by patient number. percent of control is defined as the stimulation index of the patient relative to the average stimulation index of two healthy controls. corresponding cd t-cell count (cells/μl) at the time of diagnosis is under each patient number. patients and both had disseminated bcg with bone marrow involvement and severe cytopenias, including lymphopenia, which may have impacted pha response. hematopoietic stem cell transplant outcomes for ikbkb immune deficiency. patient: detailed autopsy reports were available for patients after hsct. aside from evidence of infection, both exhibited severe thymic hypoplasia with scant lymphoid cell populations, minimal to no lymph node and tonsillar tissue, and absence of germinal follicles in the spleen. patient , who experienced secondary graft failure by -years post-hsct, developed disseminated mycobacterium avium-intracellulaire and was managed for -year with anti-tuberculous medications and splenectomy before death. the spleen was entirely replaced by atypical mycobacteria. at autopsy, performed -years after hsct, the thymus was small and atrophic, weighing only g, and lymph nodes were not readily apparent. this series presents the clinical features, immunologic presentation, and hsct outcomes for the largest cohort of ikbkb immune deficiency patients published to date. a number of important conclusions can be made. first, the clinical presentations in our case series indicate that ikbkb immune deficiency, as result of profound defects in innate and adaptive immunity, manifest as exceptional susceptibility to a variety of different but severe infections, including bacteria, viruses, fungi, and mycobacteria. we believe that ikbkb immune deficiency, although not meeting conventional laboratory-based diagnostic criteria for scid [ ] (due to higher cd t-cell counts, moderately decreased to normal pha responses, normal trecs, high proportions of naïve cd ra+ t-cells, and low cd ro+ t-cells in ikbkb immune deficiency), is as severe an immune deficiency as scid at a clinical level. we would suggest therefore, that inclusion of ikbkb deficiency under the descriptive category of "combined immune deficiencies generally less profound than scid" in the most recent international union immunologic societies classification [ ] , likely underestimates the severity of the immune deficiency and should be modified in future iterations. supporting a more profound immune deficiency are the other six reported patients with ikbkb mutations, who also presented with severe bacterial, fungal and viral infections as young infants. mycobacterial infection was also prominent, with disseminated bcg noted in four of the six [ ] [ ] [ ] . similar to our cohort, previously reported ikbkb immune deficient patients exhibited normal to elevated absolute t-cell and b-cell numbers, low cd ro+ memory t-cells, and absent or low class-switched memory b-cells. immunoglobulins were universally low or undetectable, but proliferative responses to pha were present and typically normal. only one patient was noted to have features of ectodermal dysplasia at an older age ( months) [ ] , a feature which we have not seen in any of our three post-hsct survivors. two of the other reported patients died from overwhelming infection prior to definitive therapy, and four received hsct, with only one long-term survivor. secondly, this data further expands the understanding of primary immune deficiencies involving aberrant ikk/nf-κb signaling. at least patients with germline gain of function mutations in nfkbia have been reported, with a severe but more variable clinical and immune phenotype that appears dependent upon the genotype (with missense mutations resulting in a more severe clinical phenotype) [ ] . like ikbkb immune deficiency, patients with hypermorphic nfkbia mutations that result in reduced degradation of iκbα, present with multiple and severe bacterial, fungal and viral infections starting at an early age, typically before -months. infections reported include invasive bacteria such as klebsiella, pseudomonas, haemophilus influenzae, and staphylococcus aureus, mycobacteria (including vaccine-strain bcg), as well as chronic mucocutaneous candidiasis, pneumocystis pneumonitis, and severe viral infections (rotavirus, norovirus, parainfluenza, rsv, cmv). similarly, patients with nfkbia mutations present with normal to elevated t-cell counts, an excess of naïve cd and cd cells, and normal mitogen proliferative response to pha in most. dysgammaglobulinemia (usually with one or more quantitatively abnormal immunoglobulin isotypes), impaired response to vaccine antigens, and when tested, absent or reduced memory b-cells and class switched memory b-cells have also been reported. unlike patients with ikbkb immune deficiency, however, almost all patients with iκbα defects show some degree of ectodermal dysplasia. by comparison, individuals with hemizygous hypomorphic ikbkg mutations with reduced nemo expression manifest a more variable clinical and immunologic phenotype when compared to patients with ikbkb immune deficiency and hypermorphic nfkbia mutations [ ] . in addition to infectious predisposition, patients with nemo deficiency may present with autoimmunity ( %) and inflammatory conditions (most notably inflammatory bowel disease in %), features not described in either our cohort of ikbkb immune deficient patients nor in those with hypermorphic iκbα defects. as in patients with ikbkb immune deficiency, patients with nemo deficiency commonly present with infections due to pyogenic bacteria ( %), mycobacteria ( %), viruses ( %) and fungal and opportunistic pathogens ( %) [ ] the median age at hsct for a large cohort of nemo deficient patients was . years old (range: . months - . years), an age that is older compared to our cohort, suggesting nemo deficiency may be a less severe immune deficiency relative to ikbkb immune deficiency [ ] . nemo deficiency also has a more variable immunologic phenotype, with hypogammaglobulinemia in only / ( %) and defects in specific antibody production in / ( %). similar to ikbkb immune deficiency, most patients with nemo deficiency have normal to elevated cd and cd counts, and % have normal mitogen induced proliferations [ ] . a third important finding of this study is that scid newborn screening by trec assays will not detect ikbkb immune deficiency. given the severe clinical phenotype of ikbkb immune deficiency, a pilot project of targeted newborn screening by direct genetic mutation analysis of guthrie cards for the ikbkb mutation was developed in select northern cree communities in manitoba. this investigation identified a carrier frequency of in . newborns, with a predicted mutation homozygosity of / births [ ] . over -years ( - ), newborns from two communities were screened, with one infant identified at birth (patient ) and a subsequent sibling (patient ) identified in-utero. both were placed in protective isolation after birth and received hsct at an early age, before the onset of serious infections, important variables associated with excellent outcomes in scid hsct [ , ] . in other populations with founder mutations or high rates of consanguinity, a similar genetic-based newborn screening strategy such as ours might be applicable. unfortunately, patient later died of invasive streptococcus pneumoniae infection in their home community, -months following an otherwise successful transplant. this illustrates a complex interplay between what we suspect are ongoing defects in immunity post-hsct, combined with poor social determinants of health. for familial and cultural reasons, our patients often have strong desires to return home soon after hsct, and the tertiary care hsct center is not easily accessible from their remote communities. with the death of patient , and with the recent transplant of patient , we have instituted a program of housing the family close to the tertiary care center for at least . years post-hsct, as well as regular immunoglobulin replacement and indefinite daily anti-pneumococcal antibiotic prophylaxis. much remains unknown about the immune abnormalities in patients with ikbkb immune deficiency, both before and after hsct. from our previous investigations, the broad immune defects relate to abnormal ikk/nf-κb signaling in t-cells, b-cells and innate immune cells, resulting in generalized activation defects to a variety of stimuli [ ] . with the exception of regulatory t-cells and gamma-delta t-cells, which are absent in ikbkb immune deficiency, other lymphocyte subsets appear to develop normally. despite thymic hypoplasia in five of our patients who died either prior to or after hsct, six other patients had normal trec levels at the time of diagnosis, suggesting no intrinsic defect in t-cell development and maturation within the thymus. unfortunately, none of the patients who had autopsies performed had detailed trec analysis, making direct correlation with the underdevelopment of the thymus and trec levels in a single patient impossible. detailed t-cell studies were not available for most of our patients pre-hsct, but when previously studied in one patient (and in one additional patient in this series), the t-cell receptor beta-chain variable region repertoire was polyclonal [ ] . we are unable to resolve this apparent contradiction in the autopsy and immunologic findings, and believe it warrants further study. likewise, when studied in two patients, kappa-deleting recombination excision circles (krecs) in dried blood spots from newborn guthrie cards of two patients were found to be normal, suggesting normal b cell development. in a mouse model involving a hypermorphic mutation of iκbα, nf-κb signaling in non-immune "architectural" cells were shown to be severely disrupted [ ] . although such cells are not derived from hematopoietic stem cells, they are nonetheless integral to lymphoid organogenesis, such as in the development of lymph nodes, peyer's patches, splenic marginal zones, follicular dendritic cells, and the formation of germinal centers in the spleen, which were absent. further, the immune defect in this model was not fully corrected by hsct. it is reasonable to suspect, therefore, that similar architectural cell dysfunction with resultant disruption of the immunologic niche could also play a role in ikbkb immune deficiency. failure to correct non-immune cells with hsct could explain the ongoing infectious complications seen years after an otherwise apparently successful, lymphoid engrafted transplant in ikbkb immune deficiency. the absence of splenic germinal centers and lack of tonsils and lymphoid tissues in the autopsies of patients who died post-hsct, and absent specific vaccine responses despite excellent donor t-and b-cell chimerism, supports that hsct does not fully correct all aspects of the immune deficiency in ikbkb. in years past, and in patients who were profoundly ill at presentation, full immune evaluations were not always available to further elucidate the nature of the immune deficiency in this population. at the moment, there are only three survivors in our cohort, for whom detailed t cell studies pre-and post-hsct (including serial measurements of t cell markers such as recent thymic emigrants, markers of naivety and memory cells and markers of activation or exhaustion) have been limited to date. moving forward, and with genetic-based newborn screening introduced for this condition in our province, we will now have this opportunity. despite these challenges, we believe that ikbkb immune deficiency is still a disorder that requires hsct. to our knowledge, no individual has been a long-term survivor of ikbkb immune deficiency without hsct. we are unaware of any reports of hypomorphic ikbkb mutations producing a milder clinical phenotype, allowing clinicians to take a wait and see approach. hsct for other disorders involving the ikk/nf-κb pathway have been previously described, including for hemizygous hypomorphic ikbkg mutations [ ] and hypermorphic nfkbia mutations [ ] . in the cohort of fourteen iκbα gain of function patients reported by boisson et al., eleven received hsct, with only five survivors. one patient died without receiving hsct, and two patients survived without receiving hsct. since most centers will not have genetically based newborn screening for ikbkb immune deficiency, clinicians would likely be faced with an infant presenting with severe life-threatening infections, given that trec based newborn screening will not detect the disorder. commercially available next-generation sequencing gene panels for immune deficiency disorders (including many scid panels) increasingly include ikbkb, which should facilitate diagnosis, although not always in a timely manner before hsct. in our series, two of the three patients surviving hsct had serious life-threating infections before transplant, including a case of multiple serratia marcescens intracerebral abscesses, where the long-term neurocognitive outcome was initially unclear. we have tended to be aggressive with antimicrobials and other supportive care measures, with the goal of reaching hsct as quickly as possible. this strategy has resulted in long-term survival of / infants receiving hsct with active infections. that being said, we suspect that survival will be significantly improved with current newborn screening strategies, in the absence of pre-hsct infection. unfortunately, and based on this limited dataset, we believe myeloablative conditioning may be required for engraftment in ikbkb immune deficiency. the largest and most recent study of hsct in nemo patients, however, did not find this to be necessarily true [ ] . in their cohort of fifteen patients receiving classical myeloablative conditioning and thirteen patients receiving reduced intensity conditioning, the global engraftment rate was %, with similar survival rates and secondary graft failure rates, independent of conditioning regimen intensity. our center currently uses once daily intravenous dosing of busulfan for days (aiming for an area under the curve approximating μmol/l*min), fludarabine . mg/kg/dose for days, and rabbit anti-thymocyte globulin . mg/kg/dose for days. we acknowledge other conditioning regimens might also be efficacious, although caution that late secondary graft-failure appears to be a concern in ikbkb immune deficiency. finally, and importantly, clinicians should be mindful that despite apparently reasonable basic laboratory immune reconstitution data for ikbkb immune deficiency patients post-hsct (e.g. t-cell counts, chimerism studies), late infections, including bacterial and mycobacterial infections in particular, appeared to be common. of interest, one of our long-term survivors who was fully engrafted developed primary mycobacterium tuberculosis pulmonary disease at -months post-hsct. with appropriate therapy, the infection resolved, suggesting hsct corrected enough of the immune deficiency. in contrast to our post-hsct patients with ikbkb immune deficiency, miot et al. found that bacterial infections only recurred at > -months post-hsct for nemo deficiency in / patients [ ] . we hypothesize that failure of hsct to improve all aspects of innate and adaptive immune function after transplant, as shown by the number of ongoing serious bacterial and mycobacterial infections and lack of protective specific antibody responses to vaccinations, could be because hsct does not correct cells of non-hematopoietic origin important in the immune response that are also dependent upon ikk/nf-κb signaling. a similar pattern of incomplete immune correction and ongoing bacterial and viral infections after hsct has been reported in a patient with the related autosomal dominant anhidrotic ectodermal dysplasia with immune deficiency due to hypermorphic iκbα defect [ ] . in summary, we present a relatively large cohort of a rare primary immunodeficiency disorder, ikbkb immune deficiency, characterized by the presence of naïve t-cells, evidence of a functional activation defect, and amenable to hsct, although with ongoing long-term infectious challenges. deficiency of innate and acquired immunity caused by an ikbkb mutation regulation and function of nf-kappab transcription factors in the immune system rare mendelian primary immunodeficiency diseases associated with impaired nf-kappab signaling x-linked susceptibility to mycobacteria is caused by mutations in nemo impairing cd -dependent il- production nemo mutations in unrelated boys with severe infections and conical teeth irak and nemo mutations in otherwise healthy children with recurrent invasive pneumococcal disease infectious diseases in patients with irak- , myd , nemo, or ikappabalpha deficiency a novel x-linked disorder of immune deficiency and hypohidrotic ectodermal dysplasia is allelic to incontinentia pigmenti and due to mutations in ikk-gamma (nemo) xlinked anhidrotic ectodermal dysplasia with immunodeficiency is caused by impaired nf-kappab signaling a hypermorphic ikappabalpha mutation is associated with autosomal dominant anhidrotic ectodermal dysplasia and t cell immunodeficiency a novel mutation in nfkbia/ikba results in a degradation-resistant n-truncated protein and is associated with ectodermal dysplasia with immunodeficiency the same ikappabalpha mutation in two related individuals leads to completely different clinical syndromes immunodeficiency and disseminated mycobacterial infection associated with homozygous nonsense mutation of ikkbeta immunodeficiency associated with a nonsense mutation of ikbkb a nonsense mutation in ikbkb causes combined immunodeficiency lymphocyte subsets in healthy children from birth through years of age: the pediatric aids clinical trials group p study retrospective trec testing of newborns with severe combined immunodeficiency and other primary immunodeficiency diseases establishing diagnostic criteria for severe combined immunodeficiency disease (scid), leaky scid, and omenn syndrome: the primary immune deficiency treatment consortium experience international union of immunological societies: primary immunodeficiency diseases committee report on inborn errors of immunity human ikappabalpha gain of function: a severe and syndromic immunodeficiency hypomorphic nuclear factor-kappab essential modulator mutation database and reconstitution system identifies phenotypic and immunologic diversity hematopoietic stem cell transplantation in patients hemizygous for hypomorphic ikbkg/nemo mutations newborn screening for ikbkb deficiency in manitoba using genetic analysis immune reconstitution and survival of scid patients post-hematopoietic cell transplant: a pidtc natural history study transplantation outcomes for severe combined immunodeficiency defective lymphoid organogenesis underlies the immune deficiency caused by a heterozygous s i mutation in ikappabalpha successful allogeneic hemopoietic stem cell transplantation in a child who had anhidrotic ectodermal dysplasia with immunodeficiency we would like to thank all of the patients and their families; the nursing staff (debbie hanson, jennie pitura, shannon cisneros) who have taken care of the patients; drs. ulrich pannicke, stephan ehl, and klaus schwarz who were instrumental in the discovery of the ikbkb gene; drs. cheryl greenberg and paul van caeseele for their contribution to the newborn screening project for ikbkb deficiency; and luvinia kwan and agapito gentile for performing the ikbkb mutational analysis.this research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors. key: cord- - u e q authors: nan title: selected abstracts from the th j project meeting, antalya, turkey, march - , date: - - journal: j clin immunol doi: . /s - - - sha: doc_id: cord_uid: u e q nan hans d. ochs the identification of single gene defects involving genes that play crucial roles in adaptive or innate immunity is not only important for confirming a pid diagnosis, but may contribute to optimal therapy, and contribute to genetic counseling, carrier identification and pre-natal diagnosis. to accomplish this, the diagnostician has to consider the inheritance of these disorders: x-linked, autosomal recessive, autosomal dominant and the type of mutation: loss of function, hypomorphic, dominant negative or gain of function. conventional techniques to screen for single gene mutations include flow cytometry to measure disease-specific expression of proteins (cell surface, cytoplasmic or nuclear) or to analyze relevant signaling pathways (e.g. stat b phosphorylation via the il- r; pstat via the il- receptor); and sanger sequencing of mrna or genomic dna using dye-terminator sequencing. next generation sequencing ("by synthesis") has been refined, and is being used increasingly to study families with multiple affected members with an atypical pid phenotype, or to explore consanguineous families with one member affected. whole exome sequencing requires less data analysis, compared with whole genome sequencing, but may miss intronic or regulatory elements. the challenge of whole exome/genome sequencing is to confirm that the multiple variants identified by these techniques are causative for the clinical phenotype of the study patient. however, with increasing experience, next generation sequencing will become a standard procedure for the identification of genetic defects responsible for inherited diseases, including pid. stephen jolles immunodeficiency centre for wales, university hospital of wales, cardiff, uk. immunoglobulin (ig)-replacement therapy represents the mainstay of treatment for patients with primary antibody deficiency and is administered either intravenously (ivig) or subcutaneously (scig). recent developments using a high-purity recombinant human hyaluronidase have allowed the longer term repeated use of this enzyme to facilitate the delivery of immunoglobulin and other molecules including antibiotics, local anesthetics, insulin, morphine and fluid replacement into the subcutaneous space. hyaluronidase facilitated scig (fscig) has helped overcome the limitations on the volume which can be delivered into the subcutaneous tissues by enabling dispersion of scig and its absorption into lymphatics. the rate of facilitated scig infusion is equivalent to that of ivig, and the volume administered at a single site can be greater than ml, an enormous increase over conventional scig, at - ml. the use of fscig avoids many of the systemic side effects of ivig, and has higher bioavailability than scig. over three years of safety data are now available for this approach though longer term safety data and information on anti-hylauronidase antibodies and their relevance will be required. fscig could aid several areas of patient management in both primary antibody deficiency and immunomodulatory indications. key factors influencing how it will be used in future are long-term safety data and cost-benefit analysis. date after the cd deficiency was firstly described in a turkish girl in . the patients with cd deficiency had normal b-cell differentiation in bone marrow, normal absolute number of b cells in peripheral blood and normal bcr repertoire. also, the patients had normal stimulation via the bcr, normal proliferation response upon antigen stimulation but reduced memory-b-cell compartment in peripheral blood. the cd deficiency leads to hypogammaglobulinemia and impaired antigen-specific humoral immune responses after vaccination. we had described thirty carriers in relatives of our two patients with cd deficieny and also showed that the mfi value of cd and cd expressions were lower in the carriers than in controls. during the last five years, it was also showed that the mutations of the other coreceptors of b cells such as cd and cd caused antibody deficiency. in conclusion, the description of cd deficiency reminds the importance of the molecules on b cells and contribute to identify new genetic defects (cd , cd ), and it was showed that coreceptors could affect the expressions and the functions of each other. ege university faculty of medicine, dept of pediatric immunology, izmir, turkey ig class switch recombination deficiencies are rare pids ( : , births) with normal or elevated serum igm and low igg, iga and ige levels, defective or normal somatic hypermutation, defective t/b cooperation ( %), intrinsic b cell defect ( %), susceptibility to bacterial infections begining from the first year of age (impaired b cell immunity) and lack of germinal centres in secondary lymphoid organs. we present a cd l defective case with clinical findings such as recurrent otitis media, recurrent upper and lower respiratory tract infections, sinusitis, arthritis, relapsing polychondritis , ebv-associated cervical lymphoproliferation, cmv infection, bronchiectasis, liver and spleen enlargement, multiple nodules in the liver, chronic diarrhea due to persistent cryptosporidium parvum, fungal pneumoniae, osteoporosis, and schwannoma. this case is remarkable with low igm levels and normal cd l ezpression on activated t cells although he had a novel mutation in cd l gene (a novel missense mutation in cd lg (c. c t), leading to an a. a. change from histidine to tyrosine at position (h y) at the start of the extracellular domain). in addition, we present two cases with cd deficiency with normal cd expression on b cells.both of these cases had homozygous-cd -mutation leading to a longer protein due to deletion of stop-codon. in conclusion; cd molecules although non-functional in b cells, may be normally expressed on cell surface. these cd molecules are unable to trigger signal, because cd l + il activation leads to complete lack of proliferation. evaluation of cd or cd l expression by flow cytometry may lead false results. study of cd l + cytokine (or cd + cytokine)induced b cell proliferation appears as a useful tool for these diagnosis. institute for immunology and physiology (ub ras). yekaterinburg, russia regional children clinical hospital № , yekaterinburg, russia the ural regional center of clinical immunology, which based on children clinical hospital number one (№ ) in yekaterinburg, observe patients from different territories of ural region and neighboring areas. it consists of laboratory department, consultative department, vaccination and treatment rooms, beds and boxes in special departments in the regional children hospital. the close collaboration with j-project started in . this is an example of such collaboration. in patient a. an international consilium was diagnosed a progressive neurodegenerative disease as a manifestation of primary immunodeficiency: x-linked agammaglobulinemia with b-cell deficiency. mri results: unspecified leukodystrophya rapidly progressive multifocal brain lesions with demyelinating, generalized cerebral atrophy, iii degree, signs of periventricular leukomalacia in the anterior horns of the lateral ventricles. the brain biopsy was recommended in order to clarify the nature of the defeat of the pathological process and define the role of the immune mechanisms of its development (held in the neurosurgical department with subsequent histological and immunohistochemical studies). histological and immunohistochemical study of the brain tissue of the right frontal lobe: a signs of productive meningoencephalitis in brain tissue with vasculitis, perivascular and focally moderate diffuse infiltration of mononuclear (accumulation of mononuclear cd rb+, vimentin+), most of which are cd + lymphocytes with granules of granzymeb. around -dystrophy and necrobiosis neurons, intracellular edema, small focuses of gliosis -there are isolated myeloid cells (myeloperoxidase +) and plasma cells with cytoplasmic expression of immunoglobulin light chains lambda and kappa; cells and the extracellular matrix of brain tissue expressing cd antigen and s protein. virological and bacteriological studies of brain tissue and liquor: connection of progressive degenerative changes and infectious process weren't obtained verified acknowledgments of an infectious or autoimmune process has not been received. search for a genesis of cytotoxic process in the brain continues. center for chronic immunodeficiency, university medical center freiburg and university of freiburg, germany the essential role for igg replacement therapy (iggrt) for common variable immunodeficiency (cvid) has been demonstrated in many studies and metaanalyses. while patients with "infection only" reach a nearly normal life expectancy -though still not quality -under iggrt, cvid patients with additional manifestations like inflammatory lung, bowel or liver disease, lymphoproliferative and/or autoimmune disease often require additional immunosuppressive treatment. there is little consensus on the form of immunosuppressive regimen, once steroids have failed, with possibly the one exception of rituximab treatment for autoimmune cytopenia. additional studies are essential to guide therapeutic algorithms. some of these patients suffer from late onset combined immunodeficiency (locid). as in classic forms of combined immunodeficiency, iggrt can be only a part of the treatment strategy, which needs to additionally address the cellular immunodeficiency of the patients. therefore a retrospective survey was performed on patients diagnosed with cvid who underwent hematopoietic stem cell transplantation. the results of this study are currently in revision. in summary, iggrt is the baseline therapy for cvid but does not address sufficiently the immune dysregulation in a subgroup of patients. better predictive markers have to be identified for the selection of patients for additional, potentially even definite forms of treatment in order to prevent the morbidity and mortality associated with these secondary manifestations of cvid. the first department of primary immunodeficiencies in russia was established on the basis of the institute of immunology in , when patients with pid were registered. currently, patients with pid are followed in the department of immunopathology in adults. % of pid adult patients have pid with immunoglobulin deficiency. analysis of this group of adult patients showed that the diagnosis of pid, on average has a delay of - years from the first symptoms. in % of cases, there is an infectious clinical phenotype, % -combined infectiouslymphoproliferative phenotype, % -infectious and enteropathy. the study of immunophenotyping of b-lymphocytes for the degree of maturation in this group of patients was begun. patients are currently included in this study. patients showed complete absence of b-lymphocytes, -the reduction of b-cells, patients of those have a normal amount switched memory b cells (mbc), people -a decrease amount of switched mbc. persons of the group with decreased amount of switched mbc had an expansion of transitional mbc. at present, a clear link of immunophenotypes with specific clinical phenotypes in not found, but this may be due to small sample of patients at the moment, the investigation continues. for the treatment of this category of patients only intravenous immunoglobulins are available in russia. we use drugs in various concentration of russian and foreign production. the availability of immunoglobulins for the adult patients unfortunately is not sufficient in russia, so the recommended pretransfusion level of igg is not achieved in about % of patients. our work presents the experiences of our center with the subcutaneous form of immunoglobulin therapy (scig). we have patients on such therapy. the youngest child is months old. the largest group consists of cvid patients, next-xla patients. we also substitute children diagnosed with the dgs and accompanying hypogammaglobulinemia and some children with subclasses deficiency as well as secondary hypogammaglobulinemia. in most cases we start therapy with intravenous preparates, but there have been some children to whom we proposed the subcutaneous form at the initial stage of the therapy. the main factors which made us change the mode of the drug application were adverse reactions to ivig, poor vein access and the parents`wish. the administration of scig is very rarely complicated by severe adverse reactions (the risk of their incidence amounts to about . %). even patients with serious side effects to previous immunoglobulin therapy and/or blood transfusion can be safely treated with scig. the most common side effects are local reactions but their incidence decreases during following substitutions. we can observe swelling, redness, induration, soreness. but we should remember that more severe side effects are also possible, for example: the first cvid patient presented with fever, weakness, difficulties in breathing during the following infusions. changing the brand of the drug turned out to be a sufficient method of getting rid of side effects. the second patient, also with cvid, suffered from nausea, headache, meningismus. we changed the drug brand, slowed down the infusion rate and introduced premedication with an antihistaminic drug. the third patienta girl with dgs and hypogammaglobulinemia, after having been operated on for hypoplastic left heart syndrome, responded to infusions with high fever, muscle and joint pain, skin changes (erythrodermia). we introduced premedication, changed the drug brand and slowed the infusion rate, yet without any positive effects. in two patients we observed adverse reactions after preparates at a concentration of twenty percent. there were: weakness, chills, fever, headache, and very intense pain in the place of injection. during the subcutaneous treatment of xla patients, we observe significant reduction in the number of infections and days of school absence. despite that, all our patients with xla suffer from chronic sinusitis. similar results occurred in cvid patients, but the severity of infections was the same. the use of scig results in more stable and higher igg through levels especially in xla patients. in our practice, we had only a few cases in which iv form appeared to be better than sc one. in the case of two boys with higm syndrome, we observed recurrent enthesitis of the first patient and progression of lung fibrosis of the other. ivig was better to control platelets levels in the girl with cvid and thrombocytopenia. it has also occurred that parents refuse to allow us to start subcutaneous therapy, giving two main reasons: they feel safer under frequent doctor`s control and they are afraid of making mistakes in procedures. as for the youngest children (below )their fear of needle is independent of its size, which is the third reason. in conclusion, we would like to emphasize that education programmes implemented by doctors and nurses are essential to make this form of therapy easier, safer and more satisfying for patients and their parents. despite intensive investigation into the nature of cvid, the exact molecular defect(s) and pathogenesis of disease remain unknown. our aim was to evaluate the role of t cells in the mechanisms of cvid development. additionally the impact of some innate and adaptive immunity related genes (hla, cytokine gene polymorphism, mbl genes) was investigated. based on previously observed by us constellation of shared immunogenetic profiles a comparison of t-cell phenotype of cvid patients, and elderly/young healthy individuals was performed. ten patients with cvid were enrolled ( male, female; average age - , years) presented mainly with pulmonary infections, followed by bronchoectasis and splenomegaly. our study demonstrated multiple t-and b-cell abnormalities in cvid patients such as: decreased cd +, increased cd + t cells and low cd /cd ratio, loss of naïve and early differentiated t cells, expansion of terminal effectors (cd + cd ra + cd l-) t cells, memory/effectors (cd + cd -cd -) and terminally differentiated (cd + cd +) t cells. excessive t-cell activation reflecting the prevalence of activated t cell phenotype was also detected, due perhaps to an antigen-driven process. the very low numbers of circulating mature (cd + cd +) and class-switched memory (igm-igd-cd +) b cells were pathognomic for our patients and could be used as an additional diagnostic criteria in the national guidelines. furthermore high level of nonclass switched (igm + igd + cd +) b memory cells and suppressed nk cell count was observed. decreased responsiveness to polyclonal stimuli via cd and cd pathway correlated with the loss of cd expression which was more pronounced in the treatmentnaïve cvid patients. these findings were further discussed in the context of the similarities that exist along with markers for immune senescence (lack of cd or expression of cd ). increased frequency of ifn-γ polymorphisms associated with low expression level found could indicate genetically predisposition to high activation of th lymphocytes in cvid and consequently support the concept of impaired th -type responses. in conclusion our study provided new insight into the pathogenesis of cvid. this work was partially granted by medical university sofia, grant# bcg vaccination at birth is the constant element of vaccination programmes in poland. high reactogenic bcg danish vaccine has been replaced in , by bcg moreau vaccine. frequency of disseminated bcg infection, in children with primary immunodeficiencies after bcg moreau vaccine manufactured by biomed, poland were estimated. one thousand five hundred sixty three cases of primary immunodeficiencies were diagnosed in the department of immunology, children's memorial health institute in warsaw between - . among patients with t cell predominant deficiency, group high risk of bcg infection, scid was recognized in children. mendelian susceptibility to mycobactarial diseases (msmd) was detected in four patients: ifgr deficiency and il deficiency -equally in two patients, and nemo -in one. in the group of primary immunodeficiencies regarded to be less prone to mycobacterium infections, cgd was diagnosed in , hies in patients, and xl-higm in patients. disseminated bcg infection was recognized in scid patients, of them died, because of bcg diseases. all patients with msmd developed bcg infection, one with il- deficiency died. during nearly -year-follow-up, no case of tuberculosis or disseminated bcg infection have been diagnosed among cgd , hies and xl-higm patients. early anti -tb drug prophylaxis and usage of wide range of antibiotics in therapy is crucial for cleaning of bcg infection. peter Čižnár ; julia horáková ; peter Švec ; ivana boďová ; sabina Šufliarska ; linda libai veghová ;, marieta hricová st pediatric department, comenius university medical faculty, children`s university hospital, bratislava, slovakia. transplantation unit, department of pediatric haematology and oncology, children`s university hospital, bratislava, slovakia. objectives: severe combined immunodeficiency (scid) is a group of disorders due to more than genetic defects, characterized by increased susceptibility to severe infections and early life death. the diagnosis is supported by the demonstration of low absolute t lymphocyte count variably associated with numerical defects of b and nk cells. patients are very heterogeneous regarding clinical course, immune parameters and clinical outcome. bcg (bacillus calmette-guerin) vaccine, a life attenuated vaccine was the part of slovak immunization program, administered at birth until . a comparison of clinical course of bcg exposed (bcg+) and non-exposed (bcg-) scid patients in slovakia in period of past years are given. results: incidence rate of diagnosed and treated scid in slovakia was calculated to : . , meaning , cases per year. in total cases represent ada patients, il rg deficiencies, case of complete del q and in cases genetic defect was not found by analysis of rag / , il rg, artemis, il ra, jak and ada genes. all patients were confirmed absent trec (t-cell receptor excision circles) copies in a retrospective neonatal guthrie card analysis. seven out of these patients underwent hsct, in the hsc source was a mud. favorite outcome was achieved in of them. half of our patients have been exposed to a live bcg vaccine during neonatal period. patients vaccinated with bcg faced severe complications and organ damage due to generalized skin and organ abscess formation, requiring prolonged (up to months) hospital care and complex antibiotic therapy with more than four types of anti-mycobacterium drugs, for more than years. average length of hospital care for bcg exposed patient was , months vs. , months in non-exposed group (p < , ). no statistical difference was found between the time of recognized first symptoms, and time of diagnosis in bcg + and bcg-group. the clinical presentation of non-bcg vaccinated patient differs in the initial symptoms when failure to thrive and pneumonia at months was the most common finding. post-transplantational recovery in bcg-group was less complicated. conclusions: two major improvements for the outcome for scid patients in slovakia have occurred in past years. early life vaccination for tuberculosis has been retreated and improvements in diagnostics for severe t cell defects have been made, including flow cytometry phenotyping and genetic testing within the middle european countries cooperation, the j-project. bcg and late diagnosis prolongs time for hospital care, immune reconstitution and carries severe complications, consequently it increase the costs of health care and decrease the quality of patients' life. the perspective of newborn screening for scid would be the next major step in improving the outcome of scid patients. ahmed aziz bousfiha ; leïla jeddane ; nahla erwa ; monika esser ; shereen m. reda in africa, primary immunodeficiencies are still largely undiagnosed, with no cases reported in of countries. though the african society for immunodeficiencies (asid) already organized international meetings and training schools, their impact outside the hosting country is still insufficient. at this time, only a few pid patients are reported in africa (less than patients), the majority of whom are in north africa and south africa. so, asid propose the a-project, a training program based on the j-project. some issues prevent effective training for pid in africa: diversity of languages, only a few are initiated to pid, lack of resources for travel expenses, difficulty to access to care and shading by the hiv pandemic. a-project is designed as a one-day training by an african pid expert in a small group of motivated caregivers. this project is adapted to the african context, as it only requests minimum funding and can reach more people. each a-project will be co-organized by asid and a local committee, and shall lead to some commitments to be realized by locals, in particular the establishment of a registry, a network between physicians and scientists and creation of a patient association. moreover, each a-project will be done in the medical language used in the country (english, french or portuguese). the first a-project was already done in benin, and five more are already planned for . our goal is to reach all countries where no patient were reported in years. this clinical program will raise pid awareness in africa and can potentially discover new aspects of the immunity. till very recently primary immunodeficiency diseases (pids) were not being frequently recognized in india. however, the scenario has changed over the last years or so. the indian society for primary immune deficiency (ispid) was founded in - . over the last years the ispid has organized international conferences (at new delhi and mumbai), national conferences (at chandigarh and varanasi) and continuing medical education programmes (at new delhi and lucknow). these meetings have served to act as catalysts for the cause of pids and have resulted in increasing the awareness about these conditions amongst paediatricians and physicians in our country. several centres now have the clinical skills and the technical wherewithal to perform laboratory investigations for these patients. the repertoire of tests includes nephelometery, elisa based tests and flow cytometry. facilities for molecular diagnosis of pids are also being developed at some of these centres. a lot more, however, needs to be done. the clinical phenotype of several pids in india is likely to be different from that in the west . further, the type of infections in these patients is also likely to be different. this is because of the differences in the micro-and macro-environment to which these patients exposed in developing countries. these differences have been well brought in the recent publication on chronic granulomatous disease from our centre . further, the genetic background of the indian population is diverse and several new mutations are likely to be identified amongst these patients. the indian council of medical research has taken up the lead in this regard and is proposing to set up centres for advanced research (cars) in pids - each at the post graduate institute of medical education and research, chandigarh and the institute of immunohematology, mumbai. the foundation for primary immunodeficiency (fpid), usa has also been closely involved in these efforts and has helped facilitate the development of these cars. the field of pid research in india is wide open and we are likely to witness new and exciting scientific developments in the coming years. references: . gupta s, madkaikar m, singh s, sehgal s. primary immunodeficiencies in india: a perspective. annals of the new york academy of sciences ; : - . the prevention of pid's complications, the improvement of the health care of patients with pid, the creation of the registry of the patients with pid, the implementation of the finance regulations for detection and treatment of patients with pid diseases in public health facilities, the training and professional development of medical professionals in this field. objective: to improve the detection and diagnosis of pid diseases and the life quality of patients with pid in the republic of kazakhstan. undertaken activities for realization of the project: . professor. l. marodi visited kazakhstan in october and the meeting was held in ministry of health of the republic of kazakhstan, presentations were given at the international conference held in national research center for maternal and child health, the negotiations with heads of the national medical holding' clinics and scientific center of pediatrics and pediatric surgery were held. international islamic university, kuantan pahang, malaysia. two hundred and sixty three ( ) suspected pid (primary immunodeficiencies) cases were referred to clinical immunologist led clinics in malaysia from - . patients referred were from all states of malaysia and seen at the institute of pediatric, hospital kuala lumpur and university associated hospitals in north and central peninsular malaysia the initial pid patients were seen by - pediatric immunologist between - followed by another , beginning in followed by the other in . there were ( . %) patients with at least abnormality on the immune prameter recorded and regarded as probable pid. however ( . %) were recorded as pid based on existing criteria. (who scientific committee , iuis scientific committee, primary immunodeficiency disease. ) our population were mainly children, % below years and % below year. only were above years. pid were classified as; predominant antibody deficiencies %, combined immunodeficiencies . %, other cellular immunodeficiencies . %, phagogocytic defect . %, immunodeficiency associated lymphoproliferative disorders %. our data differed from most classification where predominant antibody deficiency is most frequent as high as %, (steihm, ) . of the specific pid recorded, x linked a gammaglobulinemia (xla) , hyper igm syndrome (higm) , common variable immunodeficiencis (cvid ), selective iga deficiencies severe combined immunodeficiencies (scid) , di george syndrome (dgs) ,chronic granlomatous disease (cgd) , hyper immunoglobulin e syndrome (hige ) , primary cd deficiencies , ataxia telangiectassi % malaysia comprises of multi ethnic groups with a population of . million in . pid amongst them showed, malays at . %, chinese %, indians . % and others . % whilst the male predominate over female at a ratio of . : . family history of affected sibling or in first degree relative, or early death with suspected infant dying of infection was positive in . % which is higher than in most reports eg egypt . % (reeda ). this could be due to high consanguinity in the population. alternatively the symptomatic sibling of affected patients is more likely to be referred to the clinical immunologist. scid records the most varied organism from the positive microbiological isolate viz bacteria , fungus , virus , parasite . chromobacterium violaceum was seen in cgd patients in which deteriorated with eventual death. as in many national registries diagnostic delays remains prominent. in our series the mean diagnostic delay was o . ± . years. in comparison thailand stands at . ± . years, while france, a median of . years. malaysia remains committed to provide better diagnostic services and improved care of the pid patients through research collaboration with foreign partners with a drive for creating subspecialty training. patient groups aligned to ipopi is now closer to its formation with the creation of its protem committee ensuring that patients' interest will always be guarded aknowledgement. the authors thank all who had contributed directly or indirectly, especially medical officers, nurses, consultant pediatricians especially dr kamarul azhar, institut of pediatrics hkl, laboratory scientists especially dr shanaz murad of the imr kuala lumpur university of manchester, uk although the concept of grouping mendelian disorders associated with an upregulation of type i interferon (ifn) has not been previously recognised in the medical literature, our past and current work argues that this concept has scientific validity and clinical utility. i will discuss the possibility that such conditions can usefully be considered to represent a novel set of inborn errors of immunity, and that the recognition of diseases as type i interferonopathies will have significance for the development of targeted therapies, as well as informing our understanding of viral and retroelement biology, and the pathogenesis of some forms of autoimmunity. classic kaposi sarcoma (ks) is exceedingly rare in children from the mediterranean basin, despite the high prevalence of hhv- infection in this region. we hypothesized that rare single-gene inborn errors of immunity to hhv- might underlie classic ks in childhood. we report here autosomal recessive ox deficiency in an otherwise healthy adult with childhood-onset classic ks. ox is a costimulatory receptor expressed on activated t cells. its ligand is expressed on various cell types, including endothelial cells. the mutant ox protein was poorly expressed on the cell surface and failed to bind ox ligand, resulting in complete functional ox deficiency. the ox -deficient patient had a low proportion of effector memory cd + t cells in the peripheral blood, consistent with impaired cd + t-cell responses to recall antigens in vitro. the proportion of effector memory cd + t cells was less diminished. the proportion of circulating memory b cells was low, but the antibody response in vivo was intact, including to a vaccine boost. together, these findings suggest that human ox is important for cd + t-cell memory, but redundant for immunity to most common pathogens, with the notable and surprising exception of hhv- . the chronic mucocutaneous candidiasis disease (cmcd) is characterized by persistent or recurrent infection of skin, nails, oral, or genital mucosae with candida albicans. il- -mediated immunity has been concerned in host defense against candida on body surfaces. we have investigated nine patients with chronic mucocutaneous candidiasis disease (cmcd) and signal transducer and activators of transcription (stat ) mutations. the novel c. c > a (n k) and c. a > g (q r) mutations in the coiled-coil domain (ccd) and the c. c > t (t m) mutation in the dna-binding domain (dbd) of stat are gain-of-function (gof) for γ-activated factor (gaf)-dependent cellular responses to stat . low proportion of il- a-and il- -producing t cells, lower levels of intracellular il- a and il- by t cells and impaired candidainduced secretion of il- a and il- by leukocytes from cmc patients compared to that in healthy controls were found. the c. c > t (r w) mutation affecting the ccd and the c. c > t (t m) mutation affecting the dbd of stat and resulted in gain-ofphosphorylation and gof. these mutant alleles enhanced the cellular responses to cytokines via stat signalling pathway. these data provide further insight into the mechanism of host defense against candida. heterozygous gain-of-function stat mutation is known as a major etiology of chronic mucocutaneus candidiasis. gof mutation affecting the stat coiled-coil domen (d g) was initially discovered in -year boy with cmc. gof mutation of dna-binding domen (t m) was found in the second our patient with cmc. both patients have early manifestation of recurrent or persistent infections of the skin, mucous membranes, and nails with candida albicans. they also have skin infections with dermatophytes. patient presented from the first months of age with severe recurrent sinopulmonary infections. recurrent pneumonia and chronic bronchitis complicated by bronchiectasis, which resulted in cor pulmonale and congestive heart failure. patient suffered from recurrent hsv infection, recurrent aphthous stomatitis, has several episodes of bacterial skin infections. he also has chronic bronchitis and several episodes of pneumonia, but does not have bronchiectasis. both boys developed esophageal stricture, patient necessitating nissen fundoplication in the age of years. the patients have mild autoimmune features: uveitis (p ) and alopecia (p ). immunological investigation revealed different impairment of immune system: more severe, similar to combined immunodeficiency in p , which declines with age. the p does not have changes in lymphocyte number and immunoglobulin's, but impaired antibody production to pneumococcal antigens. western blotting performed with nuclear extracts of lymphocytes of both patients showed stronger stat phosphorylation after stimulation with cytokines ifnγ, ifnα, il- . mononuclear blood cells from both patients released much smaller amounts of il- a and il- than candida-exposed cells from healthy control. stat activation triggers transcription of interleukin (il)- which is crucial for mounting protective immune responses against fungi. several mutations affecting the stat /il- pathway have been reported, resulting in selective susceptibility to fungal (candida) infection, a hallmark of chronic mucocutaneous candidiasis (cmc). in patients with autosomal-dominant (ad)-cmc we previously reported defective th responses and identified an underlying gain-of-function (gof) stat mutation leading to hyperphosphorylation of stat . how this affects stat or leads to decreased il- remains to be determined. in patients with ad-cmc, we assessed how gof-stat mutations affect stat activation, dna-binding, gene expression, cytokine production and the effect of epigenetic modification. we show that stimulation of stat in the presence of gof-stat mutations leads to significantly reduced transcription of stat -inducible genes (rorc/il- / il- /il- /c-fos/socs /c-myc). this was not due to impaired stat phosphorylation, altered nuclear translocation nor sequestration of stat into stat /stat heterodimers. dna binding to a stat-consensus binding site construct (hsie) was intact but binding to an endogenous stat dna target was impaired. the reduced stat -dependent gene transcription could be normalized by inhibiting stat activation by fludarabine or enhancing acetylation with histone deacetylase (hdac) inhibitors trichostatin a or itf . silencing hdac , hda and hdac indicated an important role for hdac . impaired stat -dependent gene transcription likely underlies decreased th- cytokine production, susceptibility to fungal infections and other pathology seen in ad-cmc patients and could be a new target for defining novel therapeutic approaches for this potentially lethal disease. autoimmune features have been long thought as association with immunodeficiency disorders, but are now viewed as a crucial component of some diseases attributed to the breakdown of self tolerance or defects of immune regulators. it had been previously established that a single gene defect of the foxp gene (foxp in humans) caused widespread autoimmunity in both humans and mice. the clinical syndromes observed in both scurfy mice and humans suffering from ipex are similar to those observed in experimental models in which treg are selectively depleted. in , three groups demonstrated that these diseases were indeed the result of a regulatory cell deficiency. around one third of the patients with clinical manifestation closely resembling ipex syndrome, foxp is not mutated, these patients are referred to as ipex like. here we present a case; a female patient years with multiple autoimmune manifestations; dm, coeliac disease and ulcerative colitis with marked decrease in the percent of cd + cd + foxp + cells. as she has a siblings suffering from dm; the whole family was investigated. the father and the mother had % cd + cd + foxp + cells. background: hids is an autosomal recessive disease, first recognized as a separated entity at . in patients with hids, the activity of mevalonate kinase is reduced to - % of normal levels. hids is caused by mutations in the mevalonate kinase gene (mvk), located on the long arm of chromosome ( q ). it is manifested by cyclic attacks of fever initiated usually during first year of age. the frequency and severity of attacks tend to decrease later in life. materials and method: a retrospective analysis of medical history , clinical course and laboratory findings of two albanian children with periodic fever , diagnosed with hyper igd syndrome. results: case presentation : an -year-old boy admitted to the hospital because of periodic fever spikes, which occurred every - weeks and lasted - days, presented since the first year of life and coincided with the beginning of immunization. he had a tonsillectomy and adenoidectomy at the age of . the fever attacks were associated with chills, malaise, and abdominal pain without gastrointestinal signs. between attacks the patient was free of symptoms. from his family history, recurrent febrile episodes during childhood were reported to his father. physical examination showed normal findings, except for a cervical lymphadenopathy. laboratory: marked increase of erythrocyte sedimentation rate and crp. wbc-ranged from . to /mm . , high asto. serum igd was repeated several times and was always elevated (mean value: serum igd iu/ml). the mutation v i is found from the genetic examination done for gene mutations in chromosome ( q ). he repeated attacks after initial treatment with corticosteroid ,than is suggested. the second case was a four-year-old girl hospitalized five times because of prolonged fever, and diagnosed as pneumonia, tonsillitis, acute otitis media and sinusitis, treated by antibiotics. her laboratory findings were not remarkable except for increased acute inflammatory responses. serum amyloid a (saa) μg/l ( μg/l) and igd was extremely high . iu/ml. genetic examination for two mutations were negative, but reduced mevalonate kinase activity in white blood cells was demonstrated in more thorough investigations. treatment regime: colchicine conclusions: auto inflammatory syndromes always pose diagnostic and therapeutic challenges to the clinicians. the clinical description of the diversity of periodic fever syndromes is helpful in the assessment and management of these patients. although hids is predominantly identified in populations from northern european areas, it has to be considered in children with periodic fever. anastasiia bondarenko ; liudmyla chernyshova ; iryna sychova shupik national medical academy of postgraduate education, kiev, ukraine. dniepropetrovsk regional children's hospital, dniepropetrovsk, ukraine. background. aspergillus is an actual pathogen in chronic granulomatous disease responsible for about % of all infections. in - % lungs are involved and in % -cns. case. we report a case of combined loci in -years old female patient with ar cgd. the child was born from iii pregnancy, ii delivery on th week of gestation with body mass g. she received bcg vaccination at -th day of birth. at months the local inflammation in site of bcg with regional lymphadenitis developed which was treated with isoniazid for months. then bilateral purulent cervical lymphadenitis developed at , and months treated with wide spectrum antibiotics. culture from pus was negative. pcr for mycobacterium tuberculosis complex was negative. at months systemic infection without loci occurred with fever, lymphadenopathy, hepatosplenomegaly, loss of weight, progressive anemia, inflammatory changes in blood for almost months. bacteriological cultures were negative. treatment with wide spectrum antibiotics was insufficient for months. disseminated bcg infection was suspected and -compound amb treatment was started ex juvantibus with positive effect: the fever has stopped, the sizes of lymph nodes, liver and spleen have decreased, the weight of a body normalized. the child suffered from recurrent pyogenic infections, underwent disseminated salmonellosis. at the age of years blood samples were tested at the laboratory of human genetics of infectious diseases, inserm (paris, france). an absence of p phox protein expression detected by western blot conferring a complete defect in cyba due to compound of geterozigous mutations in q . at years primary tuberculosis complex of right upper lobe (mbt -) was diagnosed. at the age of during the unexplained fever multiple formations were identified in the liver, biopsy showed caseosis suspected the mycobacterial nature of lesions but mbt (-). at the age of years because of shade in the left upper lobe and ineffective standard antibiotic treatment the tuberculosis again was suspected. due to ineffective antimycobacterial treatment for months multi drug resistant tubercullosis was considered. anti-tb drugs ii line was appointed without clinical response. fever persisted. mri of brain revealed mass lesion in the left parietal lobe. because of suspected tumor the brain biopsy was done and the pus was obtained. microbiological studies revealed aspergillus fumigatus. at the same time subcutaneous tumor-like infiltrate х mm appeared on chest in a proection of lung lesions. the pus was obtained during thoracentesis. result of microbiological studies: aspergillus fumigatus. drainage of abscesses and intravenous voriconazolum led to dramatic clinical improvement and normalization of blood parameters. conclusion. features of our case is spread lesions of aspergillosis with relatively slow progression of infection. high incidence of tuberculosis in ukraine leads to a high suspicion regarding this infection. diagnosis of tb is mainly based on instrumental studies. radiological and histological differential diagnosis between tuberculosis and other infections with granulomas in cgd is difficult. high suspicion of tuberculosis led to late diagnosis of aspergillosis. great north children's hospital, newcastle upon tyne hospitals nhs foundation trust, and primary immunodeficiency group, institute of cellular medicine, newcastle university, newcastle upon tyne, uk even following the introduction of biologic disease modifying antirheumatic drugs (dmards), a small number of children suffering from severe, refractory autoimmune (ai), rheumatic and/or autoinflammatory disorders will not get into clinical remission (cr) and will potentially further suffer from multiple side-effects of combined and long-term immunosuppressive and anti-inflammatory therapies, in particular severe infections (marodi l, casanova jl. jaci ; abinun m. ped health ). whilst autologous t cell depleted hsct following the immunosuppressive conditioning regimen achieved complete clinical remission in majority of children with severe juvenile idiopathic arthritis (jia) (de kleer im et al. ann rheum dis ) , infection-related mortality remains significant (abinun m et al. mol immunol ) . therefore, following the success of allogeneic hsct in treating children with immunodysregulation, polyendocrinopathy, enteropathy, x-linked (ipex) syndrome (nademi z et al. bmt ) , we treated further children with different severe ai (alps, autoimmune lymphoproliferative syndrome (n = ); complex ai disorder (n = )), rheumatological (jsle, juvenile systemic lupus erythematosus (n = ); jia (n = )) and autoinflammatory disorders (mkd, mevalonic kinase deficiency/ traps, tnf-receptor associated periodic fever syndrome (n = ); eoc, early onset colitis (n = )). overall, of the children are alive (follow up - years), in complete and (complex ai disorder) in partial cr, original disease (alps) relapsed in , and children died ( each with alps and eoc). children had significant, but transient acute (grade - ) and chronic (limited) graft vs. host disease (gvhd), experienced multiple virus reactivation(s), and remarkably we saw significant secondary ai diseases post-hsct (transient nephritic syndrome (n = ) and cytopaenias (n = ); psoriasis, n = ; and thyroid disorders (grave's thyrotoxicosis and hypothyroidism), n = ). our data add to the positive experience and evidence acquired over the last - years (daikeler t et al. bmt ; snowden ja et al. bjh ) to propose the allogeneic hsct as a viable treatment option for the small group of children suffering from severe autoimmune disorders. the wiskott-aldrich syndrome (was) is an x-linked primary immune deficiency disorder characterized by thrombocytopenia, microthrombocytopenia, recurrent, mostly respiratory tract infections, eczema and increased risk of autoimmune disorders and malignancies. was is caused by mutations in the wasp gene which encodes wasp, a -amino acid protein. wasp plays a critical role in actin cytoskeleton organization, signalling and different functions of immune cells. we present here the results of genetic analysis of patients with was from eleven eastern and central european (ece) countries, turkey, iran and azerbaijan. clinical and laboratory information of affected males and carrier females from was families were collected. the wasp gene was sequenced from genomic dna of patients with was, as well as their family members to identify carriers. in this large cohort, we identified unique mutations including novel sequence variants. the mutations were scattered throughout the wasp gene and included single base pair changes ( missense and nonsense mutations), small insertions, deletions, and splice site defects. this study was financially supported by the tÜbİtak project s and bap project tda- - . background: chronic granulomatous disease (cgd) is a rare primary immunodeficiency disorder of phagocytes. resulting in impaired killing of bacteria and fungi. a mutation in one of the four genes encoding the components p phox , p phox , p phox and p phox of the leukocyte nadph oxidase leads to autosomal recessive (ar)-cgd. a mutation in the cybb gene encoding gp phox leads to x-linked recessive cgd. methods: we report here the results of genetically and functionally characterized patients with cgd from turkish families in turkey. results: most of the families ( %) have an ar genotype (% p phox , % p phox and % p phox ) and % have an x-linked genotype. patients with a , a and x phenotypes with oxidase null activity (dhr stimulation index of ≤ . ) were found in patients. however, in p phox deficient cases and in other ar cases with high residual oxidase activity (dhr stimulation index ≥ ) were found in patients. conclusions: residual oxidase activity is similarly lack in the x , a and a phenotype except ar cases with missense mutation. in our cohort, the percentage of ar-cgd was different from european and usa registries (in comparison with % , % and % of p phox , p phox and p phox deficient ar-cgd cases, respectively) with the higher percentage of patients with p phox ( %) and p phox -deficent ( %) phenotypes, and the lower percentage of patients with p phox -deficient ( %) phenotype. the basic difference in our results from those reported is the higher percentage of patients with ar-cgd (% ), which was lower than in the european and usa registries, probably because of the higher prevalence of consanguineous marriage in turkey. introduction: schnitzler syndrome is an autoinflammatory disorder of unknown etiology. at least some of its clinical presentation is mediated through an activation of inflammasome and release of il- , as was repeatedly demonstrated by a prominent therapeutic effect of il- blockade. recent reports bring an evidence of an important role of mitochondria in inflammasome activation and in a pathogenesis of autoinflammatory diseases. we have therefore investigated mitochondrial function and structure in patients with schnitzler syndrome. materials and methods: activity and amount of oxidative phosphorylation complexes (oxphos) were analysed by spectrophotometry, histochemistry and imunoelectrophoretic methods in fibroblast cell lines derived from skin biopsies of three adult male patients with schnitzler syndrome. ultrastructure of mitochondria, mitochondrial network and reactive oxygen species (ros) were analysed by fluorescent and electron microscopy. results: the activities and amount of oxphos complexes i, iii and iv were decreased in patients with schnitzler syndrome. interindividual differences in the degree of impairment (from severe to moderate) in analyzed mitochondrial parameters were found. content of ros, previously suggested as main inducers of inflammasome, were not significantly increased in cells with schnitzler syndrome. we, however, did find consistent and prominent changes in mitochondrial structure of all three patients. disturbed mitochondrial network and mainly abnormal, partially swelling mitochondria with unusual and sparse cristae were characteristic for all patients. we did further notice marked accumulation of neutral lipids in all tested fibroblasts. conclusion: severe structural damage of mitochondria associated with milder functional changes represented a consistent feature found in all tested schnitzler syndrome patients. along with progress in basic and clinical immunology worldwide, the knowledge and activities in the field of primary immunodeficiencies (pids) have developed during last two decades. in , a group of junior doctors and students joined seniors in this filed to establish iranian primary immunodeficiency registry (ipidr). several national and international research projects have been done so far which led to lots of publications, while improving the diagnosis of patients with pids, construction of iranian primary immunodeficiency association (ipia), and establishment of research center for immunodeficiencies were other activities which lead to better management of the patients. organizing annual meeting on clinical immunology and immunodeficiencies, celebrating pi week annually and active participation in the international congresses were all helped in to increase knowledge of physicians in the country. the overall activities in the field of pids led to an increased trend in recognition of more patients in the recent years, which was associated with decreased delay in diagnosis. based on recent report of the registry, published recently in the j clin immunol, more than new patients with pid, in addition to previously reported patients, were presented. predominantly antibody deficiencies were the most common form of disease, followed by combined immunodeficiencies, congenital defects of phagocytes, and other well-defined syndromes with immunodeficiency. the rapid progress in identification and registration of the patients with pids is important not only as of epidemiological aspect, but also as of timely diagnosis and appropriate treatment of the patients. the j project physician education and clinical research collaboration program was launched in in eastern and central europe (ece). in less than years, it has achieved remarkable success. this project aims to increase knowledge in the field of primary immunodeficiency disorders (pid), and to improve the diagnosis and treatment of patients worldwide, particularly in countries with limited economic resources, which currently report fewer such patients than expected. in most ece countries, gene sequencing, which can provide a definitive diagnosis of pid, still remains unavailable. by contrast, such technology is used elsewhere to detect the more than pid-causing genes that have been discovered in the last three decades. thus, pid awareness programs like the j project remain critically important, to improve diagnostic facilities and treatment and to promote clinical research collaboration. this paper highlights the achievements of the j project and the spread of its concepts and spirit to the countries of western asia. primary immunodeficiencis (pid ) are rare genetically determined diseases , occurring with an incidence of per inhabitants. it is heterogeneous group of disorders, from quite commonly found and usually asymptomatic iga deficiency ( in ), to a very rare diseases such as chediak -higashi ( in inhabitants). in , within the framework of a government research project no. pbz-kbn- /p / -" development , improvement and implementation of highly specialized diagnostic procedures for immune-mediated diseases", a network of cooperating national centers for diagnosis and treatment of pid, named polish working group on primary immunodeficiencies (pgr pno) was founded. as a result of joint efforts of the group as well as an implementation of three eu grants [euro-pid -nas qlrt - - ( - , euro-policy -pid sp -ct- - , euro-gene -scan ( , )] the number of centers actively working in the diagnosis and treatment of primary immunodeficiencies increased. up todate pgr pno includes pediatric centers , and since - centers for adults. development and dissemination of new diagnostic and therapeutic standards contributed significantly to the increase in detection pid. with early diagnosis of the disease -the implementation of appropriate treatment, including gamma globulin replacement therapy together with quality of life has improved. in spite of all efforts recognition of pid in poland is very rare and currently is . to , what is almost times smaller than in europe (www.esid.org). at the moment, a nationwide registry of children and adults with pid consists of patients. in september pgr -pno summarized in the annual report the current status of the substitution therapy with intravenous and subcutaneous immunoglobulin therapy in children and adults with pid in poland. on the basis of available information, half of all patients ( children and adults) were diagnosed to have antibody deficiency. these data are similar to the register of a european database esid , where the percentage of patients with pid with a predominance of antibody deficiency is more than % (www.esid.org) development and dissemination of new diagnostic and therapeutic standards as well as a national cooperation contribute significantly to the increased detection of pid. early diagnosis of the disease is followed by earlier implementation of appropriate treatment, including gammaglobulin replacement therapy together with improvement of quality of life. mutations of was were identified in was boys and in heterozygote mothers. frequently genetic damages occur in , , , exones and , intrones: deletions ( ), splice-site mutation ( ), missens ( ), insertions ( ). deletions and insertions lead to stop-codon in cases. nbs patients were homozygous for del mutation and oneheterozygote for del had delt x-cgd: deletions ( ) and nonsens ( ) mutations in different exons of cybb were observed most often. in families was performed prenatal diagnosis: x-scid- , higm - , xla- , was- , nbs- , a-t- , cgd- , xlp- , dnalig - . healthy children - . recurrent severe complicated infections developed in % of pid patients. antibody deficiencies: bacterial infections - %, enteroviral - %, tuberculosis cases, atypical mycobacteriosis - case. combined pid: bacterial infections - %, fungal - %, viral - %, opportunistic (pneumocystis jiroveci) - %, mycobacterial - % ( -complication of bcg vaccination, -tuberculosis or atypical mycobacteriosis). cgd: bacterial - % (staph.aureus, e.coli, b.cepatia, salmonella spp., klebsiella spp.), mycobacterial - % ( % bcg origin, % -tuberculosis), aspergilosis - % immune disregulation syndromes (xlp [ ] and alps [ ]): bacterial infections - %, viral - % (ebv). autoimmune violations were observed in % of all pid cases: % of combined pid, % of antibody deficiencies, % of other well defined syndromes, % of autoimmune syndromes. cytopenias developed in %, vasculitisin %, ulcer colitisin %, arthritisin %. oncology diseases developed in % of patients: mainly in nbs, a-t, was and cvid: t-and b-leukemia's, lymphomas, solid cancer. the study of primary immunodeficiency is a unique model for studying the molecular basis of immunity. question about intravital pid verification is still relevant. a regional center of clinical immunology was established in at the regional children's clinical hospital № , yekaterinburg. the regional clinical hospital № joined the center in . institute of immunology and physiology, ural branch of the russian academy of sciences carries scientific management of the center. the centre works closely with foreign counterparts in the international project j project and it has been one of the centers of jmf since . over years we formed a regional register of patients with primary immunodeficiency, comprising patients: children and adults. analysis of the regional register allows to investigate the causes of deaths in patients with pid. infectious syndrome mortality -sepsis, generalized mycobacterial infection - patients, proliferative process - patients, dominate the mortality structure. creation of specialized immunological centers permit to raise the educational level of the medical staff, precisely identify nosological forms of pid among different groups of patients, to prevent the birth of children with pid, increase the length and quality of life such patients and take part in the collaboration with international experience in the pid. introduction: the aim of this retrospective study is to determine the frequency, and demographic, clinical and laboratory features of adult cvid patients referred to our clinic. materials and methods: we retrospectively evaluated adult patients ( female (% , ), male (% , ); aged to years: median years) who were diagnosed as cvid according to esid and pagid criteria during a year period (january -march ). results: the median current age of patients was , and the median cvid diagnosis age was , years. the diagnostic delay in patients with cvid was , years (median). cvid patients presented lower levels of igm ( patients, , %), iga ( patients, , %) and igg ( patients, , %). according to lymphocyte lmunophenotypes of cvid patients, cd ( patients, , %), cd ( patients, %) and cd /cd ( patients, %) values were observed the most lower ones. discussion: we found that both of the patients with bronchiectasis showed lower levels of immünoglobulins and lower imunophenotypes of b cell than the others that do not have bronchiectasis. in our patients cd , cd and cd /cd values have got enough priority to be mentioned about an immun deficiency. in conclusion, despite recent improvements in diagnostic tools, the diagnosis of mild or moderate cvid is often delayed. however, it seems that the diagnosis of cvid is delayed especially in adulthood on account of the fact that the lack of awareness of these illnesses among the medical professionals all over the world. primary immunodeficiency disease is important in turkey because of the high rate of consanguineous marriage. the lack of awereness about immunodeficiency can cause late-diagnosis and severe complications. the objective of this study was to assess pid awereness before and after clinical immunogy education among medical students. one hundred and thirty-two questionnaires with items ( ) were distributed to seventh somestre medical students and ( %) completed questionnaires were evaluated before (first) and after (second) their education about clinical immunology courses for hours. questionnaire scores (qs) were detected as total correct answers. the mean of the first qs was . ± . and second qs was ± . (p < . ). there was no statistically difference in gender ( m and f). of questions, there were related with pid directly. the correct responses rate less than % before education were of questions. all participants corrected their responses after education. the best improvement was detected in the responses of the clinical signs related with pid. it was remarkable that the participants have known the family history related with pid excellent before education. the majority of the participants ( %) believed that a lymphocyte count of /mm was related to immunodeficiency. nbt and ch test were not found to be related with pid before education. it is also important to increase the awareness of pid among the physicians during their education in medical school and more comprehensive education in pid appears to be useful for medical students. chronic granulomatous disease (cgd) is a rare primary immunodeficiency with mutations in nadph oxidase enzyme complex which causes failure of phagocytic cells to produce superoxide and subsequent intracellular killing of microorganisms. we retrospectively analysed medical records of patients diagnosed with cgd in the last years from immunological diagnostic centres from central and eastern european countries (estonia, poland, belarus, ukraine, czech republic, slovakia, hungary, serbia and slovenia) and russia. genetic sequencing from patients' dna was performed in genetic centres in ljubljana, belarus and netherlands for mutations in known genes involved in cgd pathogenesis: cybb, cyba, ncf , ncf . we included patients with cgd in our cohort, were female. the mean age at presentation of the disease was months and at diagnosis , years. lymphadenitis ( %), dermatitis ( %), enteritis ( %), pulmonary infections ( %), liver abscesses ( %) and septicaemia ( %) were the most common clinical presentation. complications of bcg vaccination ( %) were the most common presenting infection. in total , years of followup in our cohort, the patients suffered different severe infectious episodes ( . per year). respiratory ( %), lymph node ( %) and gastrointestinal tract ( %) infections represented the most prevalent severe infections. we identified different mutations out of genes tested. in patients we identified different mutations in cybb gene, unrelated patients had the same mutation in cyba gene and in patients had typical deletion in ncf gene. in our cohort we observed high incidence of bcg infections as a presenting symptom. apart from high bcg infections patients included in our study had similar frequencies of infections and infecting microorganisms as patients described in previous series. objectives: the aim of this study is to determine the carrier rate in healthy controls from central european and balkan region. methods: we screened more than healthy subjects from countries in the region. exon and was pcr amplified and subsequently sequenced with abi prism genetic analyzer. results: heterozygous mutations were found in % of apparently healthy hungarians, % of slovenians, % of bosnians, % of serbians and in % of apparently healthy macedonians. mutations found in hungarian population were as follows: v a ( ), k r ( ). mutations found in slovenian population were: v a ( ), k r ( ) and e q ( ). mutations found in bosnian population were: v a ( ), k r ( ) and f c ( ). mutations found in serbian population were: e q ( ), k r ( ). mutations found in macedonian population were as follows: e q ( ), k r ( ) and m v ( ). conclusion: we found higher than expected carrier rate in screened populations, from % to %. it is interesting to note that more than half ( %) of detected carriers in all analyzed populations has k r mutation. progress in the field of primary immunodeficiencies (pids) is reflected in national pid registries. data from slovenian pid registry were analyzed. patients' data were collected retrospectively before and prospectively afterward. patients were classified according to international classification and updated regularly. data of patients with different pids were analyzed. interestingly, complement deficiencies are the most common, accounting for % of all entries. second most common are antibody deficiencies with %, followed by well-defined syndromes ( %), immune dysregulation ( %), neutrophil defects ( %), combined deficiencies ( %), autoinflammatory disorders ( %) and defects of innate immunity ( %). prevalence of diagnosed pids in slovenia has changed in the last years; less complement deficiencies and more antibody deficiencies were diagnosed in comparison to previous decades. the number of new pid cases has been gradually increasing, a more prominent increase has been noted in the last years. the prevalence increased most for combined immunodeficiencies, cvid and autoinflammatory disorders. the spectrum of pid entities has also widened in the last decade. three patients with scid were diagnosed and successfully treated in the last three years (incidence - : . births). high prevalence of complement deficiencies reflects early implementation of good complement diagnostic facilities and awareness among infectologists. this group of patients was prospectively collected from . combined immunodeficiencies, cvid and autoinflammatory syndromes were all probably underdiagnosed before due to lack of awareness among physicians. distribution of pid groups is more consistent with esid registry in the last five years. identification and successful treatment of scid patients in the last years is an important quality marker. bulgarian association for clinical immunology was set up in aiming to get together all specialists working in the field of clinical immunology. one of the important objectives of the association was to raise the public awareness and attract attention of specialists, national health system, government and other related societies in order to improve the diagnosis and access to treatment for children and adults with pid. efforts of immunologists led to the following results: . consensus on the diagnosis and treatment of the basic pid groups was created by the pid national working group that was established in , and specific guidelines were disseminated as well. register for pid patients has been set up in bulgaria that allowed the collection of data on the incidence and prevalence of pid and the negative effect of these conditions on the population. . educational program to improve the qualification of the physicians and provide available resources to general practitioners and raise the public awareness were introduced. collaboration with patient's organizations was developed. . treatment of pid patients has been fully covered by the public health system since march . all these steps made it possible to advance the diagnosis and management of pid in our country. pediatric pid patients care -single center experience g. petrova; p. perenovska; s. mihailova; e. naumova umhat "alexandrovska", sofia, bulgaria j-project in bulgaria started in and up to now we have elaborated programme with diagnostic criteria, well equipped laboratory, established some mutual connections with foreign colleagues, held regional meetings, conferences; created clinical standards for treatment and ensured an immunoglobulin treatment and replacement therapy. here are examples of some of the problems we face: . seven-years old boy with hypogamaglobulinaemia (normal number b-ly with abnormal function). ivig had some initial effect, but lately we noted very fast deceleration in the overall health status with possible need of lung transplantation. the case is posing a question what more could we do, could we have prevented this rapid worsening. . ten-month old girl with scid with severe bcg infection after first vaccination, referred relatively late to our center, but successfully transplanted. the case is posing a question about timing of bcg vaccination and of referral to specialized center. . nine-years old girl with unidentified immune deficiency, normal immunological follow up but clinical course as an immune deficiency with very favorable effect of ivig according the parents. the case is posing the question should we stop or should we continue ivig, despite failing to find immunological defect, based on the good clinical response. unfortunately pids are not very well recognized and sometimes the patients are referred late. sometimes poverty and lack of knowledge of patients leads to miscalculation and neglecting of their conditions by themselves, or refusal for specific tests for clarifying the diagnosis. background: although rare, primary immune deficiencies (pid) are manifested with high rate infections as well as with autoimmune and malignant disorders that are treated hardly and inefficiently. pid are not only immunological problem; they require close collaboration between immunologists, pediatricians, ent, lung and gut specialists, dermatologists, hematologists, oncologists, patients and administration. the aim of this study was to summarize the activity on registration and replacement therapy of pid patients in plovdiv region at the university hospital "st. george"-plovdiv for year ( . - . ). methodology: children with pid of humoral immunity hospitalized at the clinic of pediatrics, and adults with hereditary angioedema (hae) were included in the study using immunological and other lab tests, clinical follow up and treatment: iv and sc ig for children with pid and c esterase inhibitor (ruconest, berinert) for hae patients. results: three national workshops and a national conference on pid were hosted and organized in plovdiv since . well established university hospital immunological laboratory, detecting serum immunogloblins, blood lymphocyte populations and subpopulations and complement proteins; clinic of pediatric and genetic diseases and an information center for rare diseases and orphan drugs function in plovdiv. an expert center for diagnosis and treatment of pid was created at the university hospital. it is a team of two competent pediatricians, an immunologist and an allergist. the targets are hospitalized pid children, outpatient pid children and hae adults. the center introduced regular replacement therapy with iv and sc iv and c inhibitor, reimbursed by the national insurance. together with icrdod the experts provide education for patients and parents how to perform sc ig application as well as consultations of patients and relatives about pid.since march indicated reimbursed replacement therapy with iv ig -octagam, started regularly in hospitalized pid children with bruton hypogammaglobulinemia, cvid, omenn syndrome or igg id. these patients, aged form to years, had to hospitalizations for one year. four outpatient children with omenn syndrome or igg subclass deficiency were subjected to sc ig -gammanorm, and hae type outpatients had good response for replacement c inhibitor therapy as follows: conestat alfa (ruconest)in hae adults ( iu/ml weekly), and berinert ( u/kg b.w. weekley)in hae adult patients. conclusions: the expert pid center in plovdiv university hospital provides competent diagnosis, therapy, education and consultations for pediatric and adult pid patients from plovdiv region. the recent introduction of reimbursed replacement therapy for pid patients (hospitalized or outpatients) allows regular immunological and clinical follow up of the diseases. background: intravenous immunoglobulins (ivigs) are scarce biological products used in a broad variety of disorders. tolerance to infusions is usually good but adverse events, including some serious ones, have been reported. methodology: a cohort study aimed for detection of adverse events that occur during and following intravenous immunoglobulin (ivig) infusions at cairo university children hospitals [patients were recruited at neonatal intensive care units (nicu), pediatric intensive care units (picu), general and specialized inpatient wards ] over a time period of six months, from april through september, . the study included transfusions for different disease conditions in patients.three maltose-stabilized intravenous immunoglobulin products were administered to patients. assessments were done before, during and after the infusions. results: there were symptoms and laboratory changes of adverse events during ivig transfusions, with some patients experiencing more than one adverse reaction. adverse events were noted to occur most frequently within to h from onset of ivig infusion (n = , . %). first hour after infusion onset was the most common timing for symptoms of adverse reactions (n = , . %). patient characteristics of those with adverse reactions: adverse reactions occurred in . % of the infusions (n = ) with the majority belonging to the - years old age group (n = , . %), with variable diagnostic categories. ten patients observed during infusions ( . %) had one or more risk factors for complications, while patients observed during infusions ( . %) had no risk factors. the commonest risk factor was administration of nephrotoxic drugs (n = , . %), followed by presence of a suspected autoimmune disorder (n = , . %) and preexisting renal insufficiency (n = , . %). using regression analysis, the predicting variables for each complication were noted .for example ,fever and chills were related to infusion rate and dose whereas the predicting variables for pallor were infusion rate and presence of existing risk factors. conclusions: clinicians should be aware of the high need for special monitoring while infusing ivig to patients with primary immunodeficiency disorders, autoimmune hematological disorders and sepsis. certain diagnosis of a primary immunodeficiency disorders (pid) is most confirmedly performed by investigation of a gene defect, allowing genetic counseling and screening. molecular diagnosis helps both parents and index pid patient by carrier detection and pre-implantation testing for selecting appropriate reproductive decisions. furthermore, for confirmation of diagnosis and establishment of the inheritance pattern genetic analysis is necessary. this survey in all pid cohorts should be considered for long-term planning such as bone marrow transplantation of a pid infant at birth. the result of this testing also is important for screening of newborns and for those in specific family or ethnic groups. the prevalence of pids has been estimated to be more than / worldwide. based on the total population of iran reported in ( , , ), the expected prevalence of pids in iran would be more than , individuals. however, because of the high rate of consanguineous marriages in iran and an increased risk for development of disorders with an autosomal recessive pattern of inheritance, this prediction is likely to be an underestimation. to date, clinically diagnosed patients of pids have been reported in iran, and a definite diagnosis, defined by mutation analysis, was made in individuals. as a result, . % of the expected pid patients have been identified, and among these, . % have been diagnosed at a molecular level. the proportion of genetically definite diagnosis varied between . and % in the different disease categories. this wide spectrum might be due to unknown underlying genetic defects or modifying genes, especially in patients with predominantly antibody deficiency. on the other hand, the latter patients also had the lowest percentage of clinically diagnosed cases. croatia is a small country with inhabitants . according to expected prevalence of pids we expected approximately four hundred patients with pid. for the past few years university hospital center zagreb is the reporting center for esid registry. it is also the national center for diagnosis and treatment of pids, including haematopoietic stem cell transplantation. current pid database is running manually, with the exception of patients reported to esid registry. most of them are of pediatric age, reffered to the hospital from all over the country. the pids patients are classified according to international classification ( iuis-international union of immunological societies ). there are pids patients included in the database at the moment. the majority of them have antibody deficiency. combined immunodeficiency and well defined syndromes appear in equal distribution. other types of pids are reported in small numbers. although no consanguinity was reported, we noticed the geografic distribution of severe combined immunodeficiency patients (scid) mostly in regions, istra and podravina. it can be explained with genetic isolation. the establishment of national online pid registry is in process. the aim is to improve the diagnosis od pid specially in adult patients, who are not included in present database, with exception of the patients diagnosed in childhood. primary immunodeficiency disorders are underdiagnosed in croatia, specially in adults. establishing national pid registry will improve the physicians awareness of pids, which is particularly important for adult patients. we expected the number of diagnosed pid patients will rise. this will give the opportunity to make progress in diagnosis and treatment, and the opportunity for further epidemiological and clinical studies. another patients have well defined immunodeficiency syndromes from which: are with di george anomaly; with wiskott-aldrich syndrome; with ataxia-teleangiectasia; with hyper ige syndrome; with schimke syndrome and are with nijmegen breakage syndrome. in early nineties, two ( ) of our patients had chronic mucocutaneus candidiasis. patients are registered as x-linked scid and with xlinked lymphoproliferative syndrome (xlp). patient has chronic granulomatous disease; have severe congenital neutropenia and have hereditary angioedema. patients are with whim syndrome. patients are with anti-inflammatory disorders: has hyper ig d syndrome (hids) and have familial mediterranean fever. the j-project realization in the republic of kazakhstan allows to update the pid problem, to raise the availability of early diagnosis of pid, to improve the life quality of patients with pid by providing substitution therapy and as a result, the infant mortality and disability rate has been reduced. common variable immunodeficiency (cvid) is the most common primary immunodeficiency (pid) characterised by impaired immunglobulin production and immune dysregulation. chronic and recurrent infections and its results are typical manifestation of this disease. in addition there is a higher risk of autoimmune disorders, lymphoproliferative or granulomatous diseases and malignancies. successful management of cvid patients is based on prevention and consistent therapy of infections with sufficient immunoglobulin replacement and/or antibiotics, prevention and active screening of cvid related complications. in our study we analysed data of patients gained from medical records. these data were also input into czech pid registry and pid registry organized by esid (european organisation for immunodeficiency). we aimed at the period before diagnosis-onset of the symptoms and their characters and the course of the disease-effect of therapy, occurence of the related complications. finally, we compared our data with similar performed studies. chronic and recurrent upper and lower respiratory infections were the most frequent first manifestation of our cvid patients, but developed chronic lung disease or autoimmune disorder as well. in all patients the intravenous or subcutaneous imunoglobulin replacement therapy, eventually combined therapy with antibiotic prophylaxis, was initiated. beside chronic lung disease the most common complications were autoimmunity disorders, especially autoimmune thyroiditis, evans syndrome, trombocytopenia (itp), autoimmune hemolytic anemia (aiha). on the contrary we revealed patients with insuline dependent (type ) diabetes mellitus and cvid. only few case reports have been published with such association. successful management of cvid patients is based on a prevention and a consistent treatment of infections with sufficient immunoglobulin replcement and/or antibiotic therapy, a prevention and an active screening of cvid related complications. such approach can significantly improve the prognosis of cvid patients and the quality of their life. laura zilinskaite ; ieva bajoriuniene , ; raimundas sakalauskas ; brigita sitkauskiene , department of pulmonology and immunology, lithuanian university of health sciences, kaunas, lithuania. background. primary immunodeficiency (pid) is considered to be a rare disease. despite that it is thought that six million people may be living with a pid worldwide. in the hospital of lithuanian university of health sciences (hluhs) patients with suspected immune disorders have been diagnosed and treated since . we aimed to review the structure of pid diagnosed and treated in hluhs during the last five years. methods. data about patients with pid consulted in the hluhs was collected from the department of medical statistics. case histories of these patients were revised and patients' data was collected: onset of symptoms, type and duration of disorder, type of treatment. all patients with pid were divided into several groups according to the classification of international union of immunological societies primary immunodeficiency diseases classification committee ( ). results. there were patients with pid diagnosed in the centre. antibody deficiency was diagnosed for patients: -bruton's disease, common variable immunodeficiency (cvid) and selective immunoglobulin (ig) a deficiency. complement deficiencies were diagnosed for patients: -c esterase inhibitor deficiency and -c deficiency. another well-defined syndrome with immunodeficiency was found in six patients. the most prevalent symptom in patients with predominant igg deficiency was recurrent pneumonias which occurred at the age . ± . yrs. the mean time between the onset of symptoms and confirmation of the diagnosis was . ± . yrs. thirteen patients are on the replacement therapy with intravenous immunoglobulin. these patients had - infections/year before the treatment initiation and only - infections/year during the treatment. •pid center has successful sideproject as regional charitable public organization of invalids "society of patients with primary immunodeficiency diseases in st. petersburg" solovushka (nightingale) "". web site for patients (www.opidspb.ru) was opened due to mutual efforts of pid center staff and patients. pid center laboratory is based on spb pasteur institute central clinical diagnostics laboratory and laboratory of molecular immunology and seroepidemiology. main groups of tests perform for pid patients are: general clinical assays; flow cytometry ( -color assay on facs canto ii); humoral factors assays (ig levels, igg subclasses, post-vaccination igg levels, etc.); burst-test on flow cytometer; genetic analysis of btk, rag , rag ,was, cybb genes. despite of relatively short story of spb pid center it has a variety of completely diagnosted and successfully cured cases of pid including agammaglobulinemia with b-cell in identical siblings, wiskott-aldrich syndrome, chronic granulomatous disease (cgd), etc. introduction: the primary immunodeficiency diseases are a group of disorders caused by basic defects in immune function that are intrinsic to, or inherent in, the cells and proteins of the immune system. there are more than primary immunodeficiency diseases.. laboratory studies are necessary to determine the presence of a primary immunodeficiency diseases. the standard screening tests for antibody deficiency starts with measurement of immunoglobulin levels in the blood serum. these consist of igg, iga and igm levels. the results must be compared to agematched controls. additional studies used to evaluate patients with antibody deficiencies include measuring the different types of lymphocytes in the blood by marking those cells with molecules that can identify the different types. a commonly used test is called flow cytometry that can identify b-cells and t cells present in the circulation. methods: in in our laboratory of immunology is created the sector of examination for the pid and we measure the immunoglobulin levels with a beckman coulter immage immunochemistry system fully automated rate turbidometry and rate nephelometry method. we have determine the aged-matched levels of immunoglobulins in our laboratory. we have install also a new flow cytometer of beckman coulter company and we can determine the b cells by cd marker and t cell with cd , cd and cd marker. this examinations has been of great help in diagnosis of primary immunodeficiencies. results: in years - from the examination of children aged from - years old reported in our laboratory for immunological examination from the pediatric department of uhc mother teresa in tirana we have selected cases with disorders regarding the primary immune deficiencies. we have cases ( cases reported in - and new cases ) with nul b cell in cytoflorometry (b cd cell = %). the level of immunoglobulins was indetectable for igg , iga and igm in the moment of diagnosis. they were boys and the age at diagnosis was and years and actually they are and years treated with ivig. we have classified them as bruton (xla). we have case with low cd b-cell in circulation but only in of them we have done the immunoglobulin level. the low level of cd b cell is accompanied with different kind of hypoglobulinemies: common variable immunodeficiency cvid, with isolated iga deficiency, with igg deficiency and with both igg + igm deficiency. the immunoglobulins disorders : we have classified them according the antibody deficiencies (tab ) and we noted that the most frequent one is the deficiency of iga ( / or . % -from which iga isolated, accompanied with low igm and with low igg ) with average age at diagnosis . years old. in children in the first years of live aged from . to years old we found case with transient hypogammaglobulinemia of infancy. according this survey we can report our cases of primary immunodefiecies in uhc mother teresa of tirana with different classification conclusion: the finding are to be completed with other cases in albania and it is necessary to do the national register for pid in order to estimate exactly the prevalence of this disorders in our country. nyíregyháza-debrecen, hungary the stat mutation was confirmed in as the cause of autosomal dominant hyper-ige syndrome (ad-hies). the disease, which also mentioned in the literature as job's syndrome, is a rare primary immunodeficiency. this disease can be characterized by the following classic triad: recurrent purulent skin infections, cold abscessus which are formed in the ground of chronic ekcematoid dermatitis, pneumatocele, formation causing pneumonia and extremely high ige level. dental training interdependency as well as bone and connective tissue disorders frequently occur in the nonimmunological symptoms as multi-systemic disease. the stat protein has an important role in the area of wound healing, immunity, tumor and neovascularization. we would like to show this in case of -years old kitti whom the perinatal medical history was eventless. there is a frequent hospitalization in kitti's case history since her infancy. due to the age of three months because of serious exsiccotoxicosis, months old bilateral bronchopneumonia and pleurisy required icu care. bronchoscopy was made because of recurrent pneumonia, which excluded the bronchial malformation. lately rather otitis, mastoiditis and airway obstructive symptoms dominated the clinical picture. from serum immunoglobulins-igg is very low, whereas high levels of igewas indicated. on this basis, the diagnosis of job's syndrome was up, which is a negative stat mutations as the molecular genetic test performed demonstrated. the disease has a great clinical importance, since the risk of emergence of serious and often life-threatining complications are high. because of the prevention of disseminated infections, the early detection and the appropriate treatment are essential. in default of causal therapy, the treatment primarily concern to prevention of infections, or their aggressive antibiotic therapy. calcium and vitamin d and as well as histamine on the case of itchy skin symptoms are reccurrended to use. laboratory of molecular immunogenetics, human genetics institute, cnrs and university montpellier , montpellier, france. we performed clinical, immunological and genetic studies of hyper-ige syndrome (hies) patients from hungarian, lebanese, one russian, one polish, and one swedish families with autosomal dominant (ad) or sporadic forms of the disease to reveal cross-ethnicity of recurrent and novel mutations in the signal transducer and activator of transcription- gene (stat ). four patients from hungarian families, and one russian, and one swedish patient carried the heterozygous r w germline mutation at the dna-binding site of stat . the recurrent v m mutation affecting the src homology (sh ) domain was detected in one lebanese and one polish family, and the v del deletion located in the dna-binding domain was unveiled in another lebanese family. a novel h y mutation affecting the dna-binding site of stat in three hungarian patients from a gypsy family was also found. the segregation of this mutation with hies, restriction fragment length polymorphism analysis of stat from patients and controls and the negligible production upon il- stimulation of monocyte chemotactic protein- by the patient's blood mononuclear cells suggested that the h y mutation was disease-causing. these data suggest, that dominant negative mutations of the dna-binding and sh domains of stat cause ad and sporadic cases of hies in different ethnic groups with r w as the predominant mutation found in of the families. functional and genetic data support that the novel h y mutation may result in the loss of function of stat and leads to the hies phenotype. published in molecular immunology. : - . males with an expressed mutation in the sap (signaling lymphocyte activating molecule [slam]-associed protein) gene have an x-linked syndrome characterized by an increased vulnerability to infection with epstein-barr virus (ebv). we evaluated two related male patients with fatal infectious mononucleosis (fim) and mutation in the sap gene. sequence analysis revealed hemizygous g to a transition at nucleotide position in exon in one of the patients, and heterozygosity for this mutation in the genomic dna from his mother and maternal grandmother. this mutation resulted in asparagine instead of glycine in the sequence of the sap protein at amino acid position . to analyse the effect of this missense mutation on protein function cdna was generated by site-directed mutagenesis and cloned in pcmv-flag vector. we found that the mutant sap (sap/g d) protein was defective in protein folding as manifested by the reduced half-life compared to that of wild type sap. furthermore, the sap/g d protein was defective in binding to its philological ligands slam and b . these results suggest that defects in protein folding and ligand binding collectively contribute to the loss of function of the sap protein in patients carrying g d mutation. dedicator of cytokinesis (dock ) deficiency is an innate error of adaptive immunity characterized by recurrent viral, bacterial and fungal infections, very high serum ige concentration and a progressive deterioration of t-and b cell-mediated immunity. traditional sanger sequencing may fail to identify mutations in dock , due to overlapping large deletions in heterozygous patients. we studied the genetic and immunological features of two sisters ( and years of age) born to healthy hungarian parents. mutational analysis of genomic dna and cdna from the patients and parents by a combination of pcr and bidirectional targeted sequencing failed to identify the mutation. however, a multiple ligation-dependent probe amplification (mlpa) assay revealed two previously unknown large deletions, del - exons and del - exons, of dock in both patients. the children's mother was heterozygous for the del - exons mutation, whereas the father carried the del - exons deletion. immunoblot analysis showed an absence of dock protein from the peripheral blood lymphocytes of both patients. these data suggest that the new compound heterozygous del - exons and del - exons mutations result in a loss of dock protein function and a typical dock deficiency phenotype. our findings suggest that traditional sequencing technology may give misleading results in such cases and that mlpa may be indispensible for the definition of the large deletions frequently observed in patients with dock deficiency. we describe here a patient with invasive cryptococcus laurentii infection and the x-linked form of hyper-igm syndrome (x-higm). c.laurentii is an extremely rare human pathogen. this fungus was previously considered saprophytic and non-pathogenic to humans, but it has been isolated as the etiologic agent of skin infection, keratitis, endophthalmitis, lung abscess, peritonitis, meningitis, and fungemia. most affected individuals had a compromised immune system because of leukemia, cancer, diabetes mellitus, aids, or prematurity. repeated isolation of c.laurentii from the oropharynx of an immunocompromised patient has also been docu- the wiskott-aldrich syndrome (was) is an x-linked recessive immune deficiency disorder characterized by thrombocytopenia, small platelet size, eczema, recurrent infections, and increased risk of autoimmune disorders and malignancies. was is caused by mutations in the wasp gene which encodes wasp, a -amino acid protein. wasp plays a critical role in actin cytoskeleton organization and signalling, and functions of immune cells. we present here the results of genetic analysis of patients with was from eleven eastern and central european (ece) countries and turkey. clinical and haematological information of affected males and carrier females from was families were collected. the wasp gene was sequenced from genomic dna of patients with was, as well as their family members to identify carriers. in this large cohort, we identified unique mutations including novel sequence variants. the mutations were scattered throughout the wasp gene and included single base pair changes ( missense and nonsense mutations), small insertions, deletions, and splice site defects. genetic counselling and prenatal diagnosis were applied in four affected families. introduction: mhc-class ii deficiency is an autosomal recessively inherited combined immunodeficiency disorder characterized by less than % expression of hla-dr on b cells or monocytes. it is caused by mutation of genes (ciita, rfxank, rfx , rfxap) regulating transcription factors which controls expression of mhc-class-ii molecules on cell surface. mhc-class ii molecules are expressed on thymic epithelial cells, antigen presenting cells (b lymphocytes, dendritic cells, monocytes and macrophages) and activated t cells. these molecules are of critical importance to immunity, cd + t cell development, antibody production, tolerance induction and inlammatory response. consequently, patients present with clinical findings related to combined immunodeficiency during infancy. material and methods: medical records of thirteen patients with mhc-class ii deficiency, followed-up in ankara university, medical school, division of pediatric immunology/allergy from to , were evaluated retrospectively. findings: during study period, male and female patients were diagnosed with mhc-class ii deficiency. age of diagnosis were between months and years of age. consanguinity were present in eleven out of thirteen patients. most frequent clinical findings during initial diagnosis were failure to thrive, pneumonia and oral moniliasis. lymphopenia was absent in all of the patients, however, low serum igg level was present in all of them. except for the yo female patient with a positive family history and whose hla-dr expression was , %, hla-dr expression was % in the rest of the patients. eight patients underwent hematopoietic stem cell transplantation (hsct). two patients were lost soon after hsct due to complications and three patients died of opportunistic infections. four patients died of severe opportunistic infections without underwent hsct. results: mhc-class ii deficiency is a combined immunodeficiency and not considered a rare disease in our country. to date, only known treatment is hsct. since it has poor prognosis, hsct should be performed before development of chronic viral infections and sequelae related to infections in patients who have hla-matched sibling. fatih celmeli ; giancarlo la marca ; ines santisteban ; michael s. hershfield purine nucleoside phosphorylase deficiency (pnp deficiency) is a rare autosomal recessively inherited type of immunodeficiency. pnp deficiency constitutes about to percent of all combined immunodeficiencies. it is characterized by progressive combined immunodeficiency and neurologic findings which includes ataxia, developmental delay, and spasticity. the immunodeficiency is progressive, with normal immune functions at birth, but severe t cell deficiency with variable b cell functions presented by the age of years . the only curative treatment is the hematopoietic stem cell transplantation (hsct). here, we present a year-old girl with recurrent respiratory tract infections, short stature and spastic paraplegia. immunological, biochemically and genetics investigation revealed pnp deficiency with a t mutation in pnp gene. case report: a years-old girl was referred to our pediatric immunology clinic for recurrent sinopulmonary infections since year of age. she was full term neonate and her parents were not consanguineous. she had a history of prolonged and resistant bronchopneumonia, and an attack of generalized chickenpox complicated with pneumonia. she had also severe zona infection resolved with ulceration in cornea, one year ago. she has been suffering from frequent infections like chronic sinusitis, oral moniliasis, recurrent pneumonia and sclerosing cholangitis. she has also nonprogressive cerebral palsy, spastic paraplegia, behavioral problems and limited motor and mental retardation. there was no family history of recurrent infections or immunological disorders. on her physical examination, there was failure to thrive, (her weight and height < rd p) oral moniliasis, bilateral crepitus ralls, and splenomegaly. she has also marked spasticity with brisk reflexes in lower extremities. laboratory tests revealed lymphopenia: hemoglobin g/dl and platelets /mm wbc /mm , absolute lymphocyte /mm . the laboratory results are shown in the table. lymphocyte proliferation is lower than normal limits in response to pha stimulation (wst assay). ppd response and hiv was negative. antihbs, and antihav were negative despite vaccination. uric acid level , mg/dl. direct coombs was negative, thyroid autoantibodies were within normal limits. genetic analysis revealed homozygous missense mutation (c. g > a), which causes the a t amino acid substitution in pnp gene, in exon , which has previously been reported. additionally, a homozygous g/a polymorphic site in ivs has been detected (c. + g (ivs + g)). discussion: pnp deficiency is caused by mutations in the pnp gene at q . . this gene encodes the protein purine nucleoside phosphorylase, one of the enzymes involved in the purine salvage pathway. adenosine deaminase (ada) deaminates adenosine to yield inosine, which is then converted to hypoxanthine by pnp. pnp also converts guanosine to guanine. a number of metabolites are elevated in the plasma and urine in pnp deficiency, including deoxyguanosine and deoxyinosine. there is an intracellular accumulation of their deoxy triphosphate compounds, particularly deoxyguanosine triphosphate (dgtp). the latter is toxic to t cells, a property similar to deoxyadenosine triphosphate in adenosine deaminase deficiency. in this report, we demonstrate the clinical characteristic of the patient with late diagnosis of pnp deficiency. pnp mutations likely lead to an intense alteration of the enzyme activity which in turn, cause severe and early onset of the clinical findings. however, in our case, the clinical onset of the disease is quite late (after years) which can be explained by the residual activity of the pnp. in conclusion patients with pnp deficiency can be late onset. additionally, late diagnosis of this patient can cause severe comorbidity which limits the chance of bone marrow transplantation. di george syndrome, a disorder caused by a defect in chromosome ( q . deletion), results in the poor development of several body systems. clinical features include congenital heart defects, hypoparathyroidism and thymic hypoplasia or aplasia leading to t-cell immunodeficiency. the aim of our study is to screen and determine the incidence of di george syndrome within only one tube of blood in children with congenital heart anomalies in our population. children who were found to have a cardiac defect during routine visits in pediatric cardiology and neonatalogy departments were included into the study. cases with known genetic syndromes and newborns younger than gestational weeks of age and small for gestational age (birth weight < gr) were excluded. a total of patients were included. there were ( %) males and ( %) females. age ranged between - months ( . ± . months). parental consanguinity was % (n = ) in the study group. the majority of patients diagnosed after murmur was heard during the routine physical examination (n = , %). five patients ( %) diagnosed antenatally. remaining clinical signs on admission were as follows; respiratory distress (n = , %), tachycardia (n = , %) and central cyanosis (n = , %). echocardiagorafic examinations revealed ventricular septal defect (vsd) (n = ), tetralogy of fallot (tof) (n = ), vsd-asd (n = ), aortic coarctation (n = ), double outlet right venticle (dorv) (n = ), transposition of the great arteries (n = ), truncus arteriosus (n = ) and pulmonary atresia (n = ). pulmonary stenosis, endocardial cushion defects, total pulmonary venous return anomaly and hypoplastic left heart were the other defects. q . deletion was ascertained in ( . %) patients; these patients were diagnosed to have tof ( . %), truncus arteriosus ( . %), dorv ( . %), vsd ( . %) and vsd-asd ( . %). preliminary results of the study showed that the frequency of q . deletion is % in patients with known cardiac defects. single tube of blood is enough for flow cytometric and genetic analyses. further studies involving higher number of patients is mandatory to give sufficient information about the exact incidence of the disease. di george syndrome -where do we stand now? małgorzata pac; małgorzata skomska; ewa bernatowska department of immunology, the children's memorial health institute, warsaw, poland di george syndrome (dgs) classically comprises t-cell deficiency (due to thymic hypoplasia), hypoparathyroidism, cardiac malformations,and facial abnormalities. deletions of the long arm of chromosome at position q. are most commonly associated with dgs. syndrome is also found associated with other genetic abnormalities ( p deletions, char ge), certain teratogenic influences (retinoid acid, foetal alcoholic syndrome, maternal diabetes). the dgs phenotype is very heterogenous with variable expression of the different features including the immunodeficiency. the initial treatment emphasis is to control the hypoparathyroidism. correction of congenital heart defects (if present) is usually needed. the best treatment of the immune defects of dgs is still controversial. both hsct and transplant with fetal thymus are the option for complete dgs (cdgs). long term survival after hsct has been reported, though at a lower rate ( - %) compared to survival after hsct for scid. survival in the subgroup receiving matched sibling donor transplants was better at over %. the use of post natal human thymus was pioneered by markert at duke university and has become established as the treatment of choice for cdgs, with the result of out of treated patients survived ( %). more recently this approach has also been used in london, at gosh. . under care of cmhi there are patients fulfilling esid criteria, girls ( . %) and boys ( . %), age / - y.o. in % of them q deletion in locus d s was found. the vast majority children were diagnosed as partial dgs. none of them had significant hypogammaglobulinemia and no regular ivig therapy or antibiotic prophylaxis were required.. the mean number and percentage of cd , cd and cd lymphocytes as well as lymphoproliferative answer to pha and cd in dgs patients were slightly diminished. in many improvement of cellular immunity was observed with age. about % presented with congenital heart disease, requiring surgery, while almost % had the symptoms of hypocalcemia and hypoparathyroidism, next % -speech and learning difficulties. one child was diagnosed as cdgs. scid is a rare, inherited condition, is caused by numerous molecular defects that lead to severe compromise in the number and function of t cells, b cells, and occasionally natural killer cells. seventeen patents with scid were registered during the period from till in the children's clinical university hospital. medical charts of these patients have been reviewed. there were boys and girls. positive family history was in families. mean age at the onset of symptoms and scid diagnosis was . ± . and . ± . months, respectively. pneumonia ( %), candidacies ( %), bcg infection ( %), diarrhea ( %) were the most important infections. anemia and relative lymphopenia were in % cases , growth retardation, hypotrophy had % children . pathogens such as candida albicans ( ), mycobacterium tuberculosis complex ( ), cmv ( ) and others have been identified. totally patients died.two girls are alive ( and months post-transplant). autopsy was done in patients. we saw different changes in thymus and lymphatic nodes. artemis deficiency (n = ), t-b-scid (n = ), t-b + scid (n = ), γc deficiency (n = mutation r w in γ-chain of receptor for il- ), unspecified scid (n = ) were detected. conclusion: generalized bcg infection had % of our scid patients. (incidence of tb is still high . / population in latvia and newborns obligatory are vaccinated on the second to fifth day of life). due to possibility of absence for pid routine genetic identification in only few scid forms were identified precisely. children's hospital, university of freiburg, freiburg, germany. purpose: ipex (immunodysregulation, polyendocrinopathy, enteropathy, x-linked) is a rare x-linked recessive life-threatening disorder characterized by autoimmunity and early death. pulmonary complication related with ipex has not elucidated exactly. here, we report i.e. patients, of which died from severe pulmonary disease. methods: clinical data and laboratory findings included autoantibodies, immunoglobulin levels as well as number of t, b and nk cells were evaluated. foxp expression was performed by flow cytometry. genomic dna was isolated and all exons and exon-intron boundaries of the foxp gene were sequenced by sanger sequencing. results: patient i (pi) presented with nephrotic syndrome at years of age and then developed autoimmune hepatitis without eczema, enteropathy or high ige and died at years of age due to acute respiratory distress syndrome (ards). two cousins of pi had the same hypomorphic splice site mutation leading to normal foxp protein expression and suppressive capacity. however, they exhibited typical symptoms such as eczema, diabetes and enteropathy with eosinophilia at early age (pii, piii) and were transplanted in infancy. one of them had severe respiratory distress right after birth (piii). patient iv from another family presented with chronic diarrhea without autoimmune manifestations and died due to ards. conclusion: lung disease related to ipex syndrome has not been reported before and this entity could be a critical factor in disease outcome. severe combined immunodeficiencies (scids) are a group of primary immunodeficiencies that comprise a number of monogenic disorders characterized by a block in t cell differentiation with or without impairment of b cell and natural killer (nk) cells. without early diagnosis and treatment most children die in the first year of life. a lack or very low number of t-cell receptor excision circle (trec) detected by realtime quantitative polymerase chain reaction assay (qpcr) is consistent with t-cell lymphopenia and has repeatedly demonstrated clinical validity in population based newborn screening for scid. however, the impact of population screening will be less in communities with high consanguinity and family history of scid, in which targeted screening may be more appropriate. here, we screened high risk neonates and infants (with one or more of the following: clinical presentation and/or family history suggestive of pid, failure to thrive otherwise unexplained, lymphopenia) at cairo university hospitals. their full history and clinical examination were recorded. immunoglobulin profile and immunophenotyping of peripheral lymphocytes were performed as confirmatory tests. sixteen classical scid cases were detected, as well as another scid variants (omenn syndrome and major histocompatibility complex class ii deficiency) (totally . % of all subjects screened). the rate of consanguinity in this group was . %. secondary causes of low trecs, other than scid, in our series included: bacterial septicemia ( preterm, full-term), prematurity ( cases), one preterm with omphalocele and facial dysmorphism, one preterm with congenital adrenal hyperplasia, one full term with microvillus inclusion disease, and one full term with idiopathic tcell lymphopenia. this demonstrates that in populations with high consanguinity rates, as in egypt, targeted (non-population based) trecs assay may provide a more efficient screening strategy. case : a is a years old female, st kid of non consanguineous marriage presented with fever for months. one week after the onset of the fever red patches appeared on the face, hands, abdomen, ll. mother sought medical advice and received antibiotics, antipyretics and oral steroids for about one month with no signs of improvements regarding fever. one week later the mother noticed pallor and sought medical advice and the baby was admitted to local hospital and received one bag of blood. and then she was referred to our hospital (zagazig university hospital) where she developed acute pallor again which needed recurrent blood transfusions. dark colored urine occurred in frequent attacks with abdominal enlargement and pain, and interestingly upto this time, no improvement regarding fever, bone pain or pallor. on examination she was underbuilt (all parameters are under rd centile), pallor, tinge of jaundice, generalized skin pigmentation, generalized lymphadenopathy, nd degree clubbing and hsm investigations: cbc (pancytopenia with reticulocytopenia) -lft (increased ast with indirect hyperbilirubinemia)-kft (normal)-ldh (highly elevated u/l)-esr ( ) -fibrinogen ( . gm/ l)serum triglycerides ( . mg/dl)-ebv-vcm igm positive and igg negative -c ( . normal) -rf and ana(-ve) -serum ferritin ( ng/ml) -cd is low -bone marrow biobsy revealed dysplastic changes -l.n. biopsy revealed non specific inflammatory changes. treatment: hlh protocol with no sct prognosis: patient passed initial phase with complete resolution and waiting for bmt case : m is months old boy, nd kid of non consanguineous marriage (the st is healthy years old female) presented with fever and difficult breathing since the age of months. the fever was of gradual onset stationary course and not responding to treatment with antipyretics and antibiotics. one week later the mother noticed abdominal enlargement with red rash over the abdomen that was associated with pallor but there was no evidence of bleeding from any site, no change in the color of urine, no jaundice, no ecchymosis, no joint swelling then he is referred to our hospital (zagazig university hospital). this boy is delivered by nvd at term with no evidence of any problem either during pregnancy or delivery, he is exclusively breast fed and vaccinated as scheduled. examination: his weight was kg, length cm, and head circumference cm all are average for age, he was pale with no jaundice or cyanosis he had hsm and other systemic examination was quite well. children with pid usually are admitting to the hospitals and intensive care units of infants' pathology regional children clinical hospital № , but at a later date, in serious condition, after the development of clinical manifestations in the form of severe generalized infectious disease, or various complications including hematological. the screening technology approves children to be diagnosed in newborn period and to be observing by immunologist and hematologist for the next preventative therapy and bone marrow transplantation or hematopoietic stem cell transplantation. costs for the differential diagnosis and verification of the diagnosis before the manifestation and complications development of pid - usd. costs for the differential diagnosis and verification of diagnosis after manifestation and complications development of pid would be usd in months. economic loss prevention in pid - usd. economic loss prevention in t -lymphopenia - usd. we also should include into account the contribution to the state's economy, which will later be obtained, due to the presence of a healthy member of society. at is an autosomal recessive multisystem disorder characterized by progressive cerebellar ataxia, telangiectasia and increased susceptibility to infections and malignancies, particularly lymphomas and leukemia. laboratory immune investigations typically show decreased peripheral tcells, particularly of naïve t cells, with abnormal in vitro response to mitogens. most at patients have decreased serum iga and igg subclass concentrations. while about % of patients with at show raised serum igm concentrations during the course of the disease, it is unusual to find a high level of igm at onset. as cerebellar ataxia and oculocutaneous telangiectasia are not present at very young age, these patients are often erroneously diagnosed as hyper igm syndrome (higm). to prevent mistaking a-t patients for higm it is proposed to add dna repair disorders as a possible cause of higm. . diagnosis of at, suggested by elevated alfa-feto-protein and increased sensitivity of patients' cells to irradiation, can be confirmed by identifying a mutation in the atm gene.we report female case of at that with diagnosis of hyper igm received ivig but later they had manifestation of ataxia in the course of their disease and then had telangiectasia of conjunctiva. first case was a yrs girl that was suffered from itp and granulomatosis lesion of skin associated with hyper igm before at diagnosis. second case was a years old girl with microcephaly, sever ftt , neurodevelopmental delay , abnormal faces and hypo and hyperpigmentation lesions and anemia. with respect to these manifestation and increased afp ,nijmegan breakage syndrome was suggested. third case was suffered from hyperigm before a t diagnosis for years. we present a patient with a dock deficiency. mutations in the dedicator of cytokinesis gene (dock ) cause a combined primary immunodeficiency syndrome that is characterized by elevated serum ige levels, depressed igm levels, eosinophilia, sinopulmonary infections, cutaneous viral infections, and lymphopenia. onset of the disease was observed at -month of age with severe eczema and recurrent respiratory infections (pneumonia, bronchitis, otitis). at years of age neuroblastoma was diagnosed. from the age of years he started with severe skin infection , subsequently recurrent mucocutaneous aspergillosis was established based on skin biopsy and bacteriological studies. immunological investigations revealed persistent leukocytosis, hypereosinophilia, low level of igm and increased ige up to iu/ml. so hyper ige syndrome was diagnosed and genetically confirmed when a large deletion of the dock -gene was identified. stem cell transplantation was performed in when the patient was y.o. one year later progressive multifocal leukoencephalopathy secondary to infection by polyomavirus jc was diagnosed. but after immunosuppressive therapy with cyclosporine awas suspended our patient's condition improved: the load of polyomavirus jc on plasma showed a decrease; mri brain was essentially stable ; immunological tests showed an initial improvement of subpopulations and proliferative response to mitogens; donor chimerism % stable. Özdemir Öner ; bozdoğan sıla department of pediatrics, division of allergy/immunology, sakarya university, medical faculty, adapazarı, türkiye. background: bronchiolitis and infantile asthma are the most frequent causes for typical wheezing signs in infants. however, when a physician comes across patients with recurrent wheezing are resistant to β -agonist and anti-cholinergic therapy, known as atypical wheezing cases; he should investigate for hypogammaglobulinemia in these patients. aim: here, three cases are reported to make pediatricians aware of hypogammaglobulinemia, which is one of the reasons causing recurrent and persistent wheezing attacks during infancy and beyond. case presentations: case : month-old girl presented to us with complaining of coughing and persistent wheezing. she has been having wheezing and breathing difficulty for the last months after she got upper respiratory tract infection. her symptoms persisted even though she was using religiously nebulized salbutamol + budesonid therapy. before this episode, she had had other wheezing attacks in her past medical history beginning from months of age. in her family history, her father has asthma. physical examination revealed her breathing difficulty. ronchi as well as rales were heard on the auscultation of her lungs. at the fourth day of admission, she was given ivig mg/kg/dose. later, her symptoms did improve and not recur for the last months. laboratory findings showed normal routine biochemistry, complete blood count and sedimentation rate. chest x-ray showed normal findings. echography was normal. ph-metry for reflux investigation was normal. sweat test was normal. in the serological evaluation: low igg level for his age ( mg/dl) was detected at two different times. igg subgroups, igm ( mg/ dl), iga ( mg/dl) and ige ( ) levels were within normal. case : -month-old girl came to our outpatient clinic with complaints of coughing and wheezing. at months of age, she had urinary and upper respiratory tract infections. despite antibiotic therapy, wheezing persisted for months and wheezing severity increased and it did not respond to β -agonist therapy. thereafter, she was admitted to the hospital for days and symptoms resolved. however, she came back to hospital due to recurrence of her symptoms in days. in her family history, grandmother and her cousins have asthma. physical examination showed breathing difficulty. ronchi as well as rales were heard on her lungs. although salbutamol, ipratropium, antibiotherapy (clarithromycin) and anti-reflux therapies were given, her symptoms did not improve for weeks. at the th day of admission, she was given ivig mg/kg/dose. later, her respiratory system symptoms did not recur for the last several months. once she was evaluated for persistent wheezing attacks during admission, biochemistry, cbc, esr were normal. chest x-ray and echography were normal. in serological evaluation: low igg level for his age ( mg/dl) was detected at two different times. igg subgroups, igm ( mg/dl), iga ( mg/dl) and ige (< ) levels were normal. case : month-old boy was brought to us complaints of having frequent lower respiratory tract infections (bronchiolitis). he was experiencing recurrent wheezing attacks almost every other week for the last months. in past medical history, he was diagnosed with trisomy and hypothyroidism at the months of age. he went thru an operation for atrio-venticular septal defect. physical exam revealed dyspnea, tachypnea and wheezing. crackles were heard on the chest auscultation. abdominal, cardio-vascular and the rest of the examination were normal. when he was evaluated for frequent wheezing attacks in our outpatient clinic, routine biochemistry, cbc and esr were normal. chest x-ray showed normal findings. in serological evaluation: low igg level for his age ( mg/dl) was detected twice. igg subgroups, igm, iga and ige levels were within normal. he was given ivig mg/kg/dose. for the last three months, he did not have any lower respiratory tract infection. conclusion: the awareness of immunodeficiency among pediatricians has been greatly improved. recurrent respiratory tract infections are major infections in these patients. thi is a relatively common condition associated with infant hypogammaglobulinemia. in patients with recurrent and/or persistent wheezing symptoms during infancy and beyond, especially resistant to therapy, hypogammaglobulinemia should be excluded from possible diagnoses. background: the acquisition of new food allergy after transplantation or transplant-acquired food allergy (tafa) is usually reported in adults and rarely in children. tafa is described mainly after liver, but also after small bowel/intestinal, lung and heart transplantations. in different studies, the male/female ratio is equal. literature data suggest that children with tafa typically present within the first year after surgery and they are typically allergic to multiple foods. aim: tafa is generally characterized with allergy to multiple foods and increased level of total and/or spesific ige. here, a patient although who had normal total. ige and specific ige test results, he developed reaction to skin prick test for cow's milk is presented and his clinical presentation will be discussed. case presentation: month-old-boy came to our allergy clinic with complaints of vomiting after drinking cow's milk and skin rush on the area where contact ed with chocolate. in his past medical histroy, left lateral segment of liver (donor was his mother) was transplanted to him when he was at months. the liver donor was not recorded as having a history of allergic disease. methylprednisolone and tacrolimus immunosuppression were used after the transplantation, and tacrolimus therapy was continued for prophylaxis of chronic rejection. when he was at months, family fed the patient with cow's milk but hours later he began to vomit. he vomited five times in two hours. then, he developed constipation. rectal irrigation was used. then oral intake stopped for two days. he was thought to be having food protein induced enterocolitis. his vomiting complaints repeated after intake of formula and baby food which include grain. so he fed with special formula including short-chain peptides and free aminoacids and his symptoms improved. past medical history: extrahepatic biliary atresia was diagnosed at weeks age with conjugated hyperbilirubinemia (according to scintigraphy and biopsy results). family history: his father has penicillin allergy and his aunt has asthma. at our outpatient clinic: height and weight were within normal percentiles. physical examination reevaled normal examination findings. laboratory findings: wbc was . /mm , with % neutrophils, % eosinophils and % lymphocytes. his hemoglobin was . g/ dl , platelet count was . /mm . total ige : < and immunocap specific ige against milk, grain and other classsic foods was < . . skin prick test results: saline: x mm, histamine x mm, fresh cow's milk: x mm, other food allergens (peanut, egg, fish, soybean, wheat): x mm. conclusion: our patient seemed to have cow's milk allergy related to liver transplantation. laboratory investigations and clinical presentation of the patient did not look like typical ige-mediated food allergy, which is expected in tafa. patient history: years old male patient, the first pneumonia at the age of followed by times pneumonia attacks/year, otitis media and sinusitis. hospitalization due to respiratory insufficiency caused by bronchiolitis obliterans and diagnosed with hypogammaglobulinemia at the age of . no consanguinity. patient findings at diagnosis ( / ) had hepatospenomegaly, bilaterally cervical, supraclavicular axillar lap, osteoporosis, bronchiolitis obliterans organizing pneumonia, hyper igm, lower igg, iga, ige, low b-cell. no response to tetanus toxoid. İsohemagglutinin antib was ¼. cd ,cd l, aid, taci, baffr, icos gene mutation were negative. protein electrophoresis revealed polyclonal igm increase, immunofixation was no clonality. lymph node biopsy result was available paracortical expantion . cd (+) t and cd were positive. b lymphocyte distribution were normal. malignancy ruled out. no giant germinal centers. evaluation of bone marrow aspiration/ biopsy were abnormal localization of megakaryocytes, dismegakaryopoiesis, blasts in normal range. lymphocyte ratio % mostly consisting of cd + cells, cd + cells were rare, plasma cell ratio was %, amyloid negative. progression of patient, igm level and spnenomegaly were increased and patient had respiratory failure. splenectomy could not be done due to respiratory failure. than patient was treated with rituximab ( mg/kg/week). after rituximab therapy, lymph nodes and splenomegaly were ( cm), regressed and igm level decreased (from mg/dl to mg/dl), increased effort capacity. after month after rituximab therapy splenomegaly and igm level were progressed. splenoctomy was performed. pathological evaluation of the spleen malignancy ruled out. current igm level is mg/dl. conclusion: the most convinient scenerio for this patient would be a csr defect of unknown etiology presented as cvid. the recent litarature revealed genetic defects of some molecules operating in dna repair pathways such as msh , msh , msh , msh , mlh , rad , rad , nbs leading to csr abnormality and impaired antibody maturation. cvid is characterized by hypogammaglobulinemia, recurrent respiratory and gastrointestinal bacterial infections. good's syndrome(gs) is a thymoma-related immunodeficiency and characterized by hypogammaglobulinemia, decreased b cell and variable deficiencies in cell-mediated immunity. case: a -year old male patient presented with a palpable anterior chest wall mass. in , he was diagnosed with rheumatoid arthritis. laboratory showed hypogammaglobulinemia and all autoantibodies were negative. cd + , cd + , cd + cells were < %. bone marrow(bm) examination demonstrated low cd + b-lymphocyte and increase in cd + t cells. tuberculin skin test was positive. t-cell proliferative response was normal. immunizations with h.influenzae type-b and tetanus toxoid revealed no response. anti-b titer was low. taci, btk and icos mutattions were negative. ultrasonography showed hepatosplenomegaly. mild edema, mononuclear cell infiltration (suggested the early stages of extrahepatic biliary obstruction) were detected on liver biopsy. in the medical history, he had reported chronic sinusitis, otitis and bronchitis dating back to rd decade of life. at age , he underwent surgery for thymoma. in follow-up, the computed tomography showed soft tissue mass on the anterior chest wall and pathology was thymoma. discussion: frequent respiratory infections with encapsulated pathogens beginning at the age of , lack of opportunistic pathogen infections, presence of hepatosplenomegaly and rheumatoid arthritis, bm examination findings and successful management of infections with ivig therapy all indicated a diagnosis of cvid. the coexistence of cvid and thymoma has been reported in the literature. introduction: common variable immunodeficiency (cvid) is a primary immunodeficiency disorder characterized by impaired b cell differentiation with defective immunoglobulin production. it has heterogeneous clinical manifestations including recurrent infections, chronic lung disease, autoimmune disorders, gastrointestinal disease, and susceptibility to lymphoma. patients with this disorder have evidence of immune dysregulation leading to autoimmunity. autoimmune cytopenias are a more common presenting disorder in children and may be the initial manifestation of the disease. we want to present a patient presenting with autoimmunue hemolytic anemia and finally diagnosed as cvid. case: a years old, previously healthy female patient applied to emergency clinic with complaint of paleness, light headedness and yellow discoloration of her scleras. her history was not compatible with blood loss. she denied having melena, hemotochezia or hematuria. her mensturation history was also normal. her hemoglobin level was . gr/dl, reticulocyte count was % . , complete blood count was otherwise normal. direct coombs was (+++). for the differential diagnosis of immune hemolytic anemia, viral serology, ana and anti-dsdna were studied, but the results were normal. igg, igm and iga levels were lower than normal normal for her age. other causes of hypogammaglobulinemia were excluded. blood group was arh (+), blood isohemagglutinin were anti a(-) and anti b (-/ +). lymphocyte subsets were also studided. as the patient has reduced immunoglobulin levels with normal lymphocyte subset analysis, she presented after puberty and other defined immunodeficiency states were excluded, she was diagnosed as cvid and monthly immunoglobulin replacement therapy was planned. she had aseptic meningitis after her first ivig transfusion. the ivig preparation was changed with another trade and she did not have any problem during the following treatments. it has been one year since she was diagnosed and she did not have any other medical problems. conclusion: the diagnosis of cvid requires decreased igg, igm and iga levels are also reduced but are less valuable for diagnosis. ige level is checked to exclude other disorders. igg subclass determinations are indicated if antibody titers are decreased but immunoglobulin levels are near normal. hypogammaglobulinemia secondary to other disorders should be excluded. autoimmune conditions can be the presenting signs/symptoms in cvid. autoimmune hematologic disorders may precede, present at the time of diagnosis or develope during the course of cvid in approximately one-half of the patients with autoimmune problems. selective immunoglobuline deficiency is an uncommon dysgamaglobulinemia, in which immunoglobuline levels except igm level are normal. it can be primary or secondary to cancer, autoimmune diseases, gastrointestinal system diseases and immunosuppressive therapy. patients can be asymptomatic or have recurrent infections, asthma, angioedema, autoimmune diseases, celiac disease and bronchiectasis. allergic diatheses are the second commonest presentation of selective igm deficiency. in this presentation, we report a case with asthma and angioedema who has selective immunoglobuline m deficiency. a year old male patient who has been diagnosed with asthma for years with a well controlled asthma for years presented with labial angioedema. he had labial angioedema daily without antihistamines. he did not have any suspected food or drug allergy. he did not have family history of angioedema. physical examination was normal under antihistamine therapy. laboratory evaluation revealed a . % percentage of eosinophils. absolute lymphocyte count was , absolute neutrophil count was cells/mm . immunoglobuline e value was ng/ml, levels of immunoglobuline g and a were within normal limits for age. immunoglobulin m value was . mg/dl ( - ). anti a was / , anti b was ½ positive, antihbs was above mlu/ml. lymphocyte subsets were normal. because of the continuous usage of antihistamines, prick tests could not be done. levels of d was . , d was . ku a /l. thyroid functions, antitpo, c , c and c esterase inhibitor values were normal. antinuclear antibodies and antitransglutaminase lga was negative. immunoglobuline m value of his father was normal, immunoglobulin m value of his brother was . mg/dl ( - ). selective immunoglobuline deficiency is a rarely seen dysgamaglobulinemia. it was reported in children with asthma, but it was not reported in children with angioedema. it can have value in the clinical evaluation of patients with angioedema. cukurova university, department of pediatric allergy and immunology, adana, turkey a patient who is at age of years at present and who has been followed up at the our clinic with xla diagnosis presented with pain in his ankles and wrists and swelling of his right knee. he also was suffering from skin tightening of the lower extremities. his physical examination revealed arthitis of the right knee and sclerotic changes were detected in the skin. skin biopsy was performed and it revealed morphea (localized scleroderma). after the diagnosis of morphea and arthritis, ivig gm/kg and nsaid were applied. following the treatment skin findings and arthritis resolved, however approximately two months later liver enzymes were detected to be high in his routine control. liver biopsy performed to clarify the aetiology of elevated liver enzymes was reported as autoimmune hepatitis. in addition to ivig gm/kg, budenofalk mg/day was started. after months of treatment, his liver enzymes normalized. currently he is being treated with ivig monthly and ursofalk daily. patients with xla typically present with recurrent bacterial infections and it might be associated with some autoimmune diseases. there are not any reports indicating an associaton of autoimmune hepatitis and scleroderma with xla. bruton agammaglobulinemia is an inherited immunodeficiency disease caused by mutations in the gene coding for bruton tyrosine kinase (btk). the median age at the diagnosis of the antibody deficiency is about . years in turkey. here, we report a case of bruton disease presenting with recurrent cervical abscess at two months old infant. a -months-old boy was firstly hospitalized for the treatment of the right cervical abscess at days old. after the recurrence of swelling on the cervical area, the patient was referred to the admitted to hospital secondly. there was no consanguinity between parents and, his family history was unremarkable except four of the mother's cousins, died because of unknown etiology in infancy. on his physical examination, his weight, height andhead circumference were normal range by age. there were no visible tonsils. there was a palpable, mobile x cm mass on right upper cervical area. the molecular analysis of the causal gene for bruton's tyrosine kinase (btk gene) revealed the mutation in exon . this mutation g. delg (c. + del) leads to the changes of amino acid order in the protein with the subsequent changes in activity of btk (at the level of dna: substitution of glutamic acid (p.glu *) causes non-sense mutation leading to the formation of stop codon with premature end of dna transcription to cause stop codon. after initiating the intravenous immunoglobulin with antibiotics, the cervical mass was getting smaller in a short period, and had not observed again. neutropenia was improved within the months. this case is an important example to diagnose bruton disease in early life. it demonstrates that maintaining a high level of clinical suspicion is essential for the diagnosis of bruton disease in a child with recurrent cervical masses. elif azarsiz; neslihan edeer karaca; guzide aksu; necil kutukculer ege university faculty of medicine, department of pediatric immunology, izmir, turkey transient hypogammaglobulinemia of infancy (thi) is characterized by recurrent infections and reduced serum immunoglobulin levels. typically, thi patients recover spontaneously, mostly within - months of age, but sometimes recovery may be delayed until - years. the use of intravenous immunoglobulin (ivig) as an alternative to antibiotic prophylaxis remains contraversial also in symptomatic patients. some authors believe that ivig therapy may cause a delay in the maturation of the humoral immune system because of the interference from passively transfered antibodies. the aim of this study was to investigate the effect of ivig replacement on recovery from immunodeficiency in these patients. patients ( %) received ivig therapy while patients ( . %) showed spontaneous normalization without ivig. the percentages of patients who had more than six times the number of febrile infections in a year decreased from % to % in the group receiving ivig treatment. at admission, before being recruited to ivig therapy, serum immunoglobulin g (igg) levels and anti-hemophilus b (hib) antibody titers were found to be significantly low in cases who were selected for replacement. the percentages of patients who did not have protective levels of anti-hib, anti-rubella or anti-rubeola-igg were also significantly high in ivig cases. there was no statistically significant difference in the age at which igg levels normalized between both groups. patients in the ivig group and non-ivig group reached normal igg levels at the age of . ± . and . ± . months, respectively. in conclusion, ivig infusions do not cause a delay in the maturation of the immune system in thi patients. the very low and non-protective specific antibody responses against previously applied vaccines are important factors to consider when selecting patients for ivig therapy. zoltán ellenes-jakabffy ; ibolya kovács ; mihaela bătăneanţ ; maria cucuruz ; margit Şerban ; lászló maródi department of pediatrics, clinical city hospital oradea, romania. objective: to study the amplitude of the chronic inflammatory phenomena: atopic and autoimmune diseases, as well as their associations in pediatric sigad patients and to study the sigad patients' family history ( st degree relatives) for pids (primary immunodeficiencies). methods: retrospective analysis of the clinical and laboratory records of pediatric sigad patients diagnosed between and at the departments for pediatric immunology of the medical universities of debrecen (hungary), oradea and timişoara (romania). results: out of patients, we found out ( %) with atopic diseases, mostly with respiratory localizations (asthma and allergic rhinitis), ( , %) patients with autoimmune diseases (jia, psoriasis, celiac disease, thyroiditis etc.) and other patients without clinical symptoms of autoimmunity but constantly elevated autoantibody levels. there were patients with coexistent atopic and autoimmune diseases. regarding the family history, we identified families with multiple cases of pids : with multiple sigad, with sigad and cvid, with sigad and higms (hyper igm syndrome). conclusions: the chronic inflammatory phenomena are present in the majority of the studied sigad patients: symptomatic atopic diseases in %, symptomatic autoimmune diseases in , %, that means a cumulative %. there are comorbid associations within the atopic and autoimmune disease groups and also between the two groups. sigad is the most common primary immunodeficiency. it's prevalence is / - / . aim was to assess the prevalence of co-morbidity in patients with sigad in latvian pediatric population and the analysis of some immunological abnormalities in these patients. the study included patients - years old. medical charts have been reviewed. into account were taken the data, which were made at time of diagnosis. patients were divided into groups: st -patients with allergic disease, nd -patients with autoimmune diseases, rd -patients with infectious diseases, th -asymptomatic patients or patients with sigad unrelated diseases. patients with multiple co-morbidities of various disease groups were not placed in any of these groups. each patient group was divided by age: - years, - years - years. results. . % were boys and . % girls. sigad in . % of the cases were diagnosed before years of age (inclusive). % of the patients had co-morbidities: allergic ( %), autoimmune ( . %) or infectious diseases ( . %). patients - years old: children with infectious and autoimmune diseases have . times greater igg than healthy children or children with allergic diseases (p < , ); children with autoimmune diseases has . times more cd cells than children with allergic diseases (p < , ).; children with infectious diseases have . times lower absolute number of cd cells than children with autoimmune diseases (p < , ); patients - years old: children with infectious diseases have . times more absolute number cd cells than children with allergic diseases (p < , ). patients - years old: children with autoimmune disease have . times higher cd /cd index than children with infectious diseases (p < , ) karakina m. l. , , , tuzankina i. a. , , vlasova e. v. introduction: antiepileptic drugs are known to cause immunosupression in some cases. levetiracetam is an anticonvulsant medication used to treat epilepsy in the posttraumatic seizures. we report a rare case of hypogammaglobulinemia and b cell aplasia associated with levetiracetam treatment. case report: a -year-old female was operated for pituitary tumor with transnasal surgery and required second operation for postoperative rhinorrhea. after operation, menengitis developed and antibiotic treatment was administrated. however, there was a poor response to this treatment after one month and craniotomy was performed due to the diagnosis of "shimic menengitis". her seizures occurred as a postoperative complication and levetiracetam was initiated. after the -month follow-up, the findings of menengitis could not be controlled with antibiotherapy. she was referred to our immunology department for chronic menengitis with fever, headache and high cerebrospinal white blood cell count. results: in her clinical evaluation, it was learned that she had previously healthy. laboratory examination showed that decreased levels of igg mg/dl (normal: - ) and iga mg/dl (n: - ). peripheral blood flow cytometric analysis revealed the absence of b cells (cd + b cells; < %). t cell subsets and natural killer cell numbers were normal. neutrophil function, chemotaxis, phagocytosis and oxidative burst activity were found to be normal. isohemaglutinin titer, levels of pneumococal and tetanus specific igg antibodies were also normal. antiepileptic drug was discontinued after epileptic seizure was controlled. b cells gradually increased three weeks later and returned to normal within two months (cd + b cells: . %). conclusion: patients requiring levetiracetam should have serum immunoglobulins measured and lymphocyte subsets analysis performed if they experience recurrent or persistent infections. mustafa gulec ; fevzi demirel ; ugur musabak ; ozgur kartal ; sait yesillik ; abdullah baysan ; ergun ucar ; osman sener gulhane medical school, division of immunology allergic diseases, ankara, turkey. gulhane medical school, department of chest diseases, ankara, turkey. introduction: common variable immune deficiency (cvid) may present with several clinical manifestations involved in different organs and tissues in adults. we present a case with a history of chronic cough for more than twenty years and further diagnosed as cvid. case: a -year-old male who works in a chemistry lab admitted to our clinic with a history of frequent upper respiratory tract infections for more than years. he also had intermittent diarrhea symptoms and his respiratory symptoms have been worsened since . he had been hospitalized due to pneumonia and empyema several times. he had undergone left lower lobectomy due to bronchiectasis in . he had been admitted to intensive care unit due to worsening of his medical condition. he was further diagnosed with cvid and ivig treatment was initiated. he is currently under remission with monthly ivig treatment and without any respiratory or gastrointestinal symptoms. discussion and conclusion: cvid is a clinical disorder in which the humoral part of the immune system is affected. most frequent presenting symptoms belong to respiratory, gastrointestinal systems and skin. however, due to the organ specific physical examination and lack of awareness, the diagnosis is frequently overlooked. in our case, frequent upper respiratory infection, loss of weight, diarrhea, bronchiectasis and empyema with unknown etiology are the most informative clinical signs. medical history is the most important part of patient evaluation. immunoglobulin replacement therapy is a basic treatment in primary immunoglobulin deficiency disorders. immunoglobulin substitution can be given intravenously (ivig) or subcutaneously (scig) for patients with antibody deficiency. both of these treatments are effective in prevention and cure of infections, although differences in advers events profile and patients' quality of life can be seen. the authors describe here their experiences in switching patients from ivig treatment to scig and a few years observation of scig therapy of patients with antibody deficiencies. sirje velbri , mirja varik tallinn children's hospital, tallinn, estonia. north-estonian regional hospital, tallinn, estonia. j clin immunol ( ) : - antibody deficiencies are the most common group of primary immunodeficiencies. the main hallmark of antibody deficiencies are recurrent infections but the patients have also higher risk of autoimmune and allergic diseases. we analysed retrospectively the frequency and character of autoimmune diseases in patients ( children and adult patients) with primary antibody deficiencies. there were ana-lysed patients with xla, patients with cvid, patients with selective iga deficiency, pa-tients with iga/igg subclass deficiency and patients with isolated igg subclass deficiency. autoimmune diseases were found in patients ( , %) besides in children ( %) and in adult patients ( %). in two boys with xla there was not found autoimmune diseases but in patients with other forms of antibody deficiencies in - % of cases. autoiimmune diseases were found more often in iga/igg subclass defi-ciency ( %) than in other forms of antibody deficiencies ( - %). the spectrum of auto-immune diseases differed in adults and child-ren and in different forms of antibody de-ficiencies. immune thrombocytopenia was found in adult patients with cvid or igg subclass deficien-cy, autoimmune connective tissue disorders in iga and iga/igg subclass deficiency. in children there was found mainly thyroiditis, diabetes i type and juvenile arthritis. rostov state medical university. research institute of clinical immunology, rostov-on-don, russia. primary immunodeficiency (pi) is currently one of most important genetically determined immunological clinical pathology which is hard to manage . among different types of pis almost % of cases occurs due to a deficiency in antibodies production. injections of immunoglobulin are current standard in the management of pi. however this treatment is expensive, often is hard for patients, and frequently has limited effectiveness. substitution of immune proteins frequently is inefficient for treatment of severe infectious conditions in pi patients. we investigated maturation, activation and differentiation of immune t-cells in the dynamics of ivig replacement therapy. we observed patients with with cvid ( ) and xla ( people) over a year of regular replacement therapy. we have found that recovery of the humoral component does not affect the maturation and the differentiation of t-cells, but can reestablish activation and regulatory properties. these changes are more evident among patients with cvid, which immuneregulatory and functional potential reestablish faster. obviously, the effects tlymphocytes increase the effectiveness of replacement therapy in patients with xla, cvid common variable immunodeficiency (cvid) is one of the most frequent symptomatic primary immunodeficiencies. the diagnosis is based on significantly decreased levels of immunoglobulins, with poor or absent response to vaccines and by excluding other defined causes of hypogammaglobulinaemia. as suboptimal antibody production is mainly due to b cell defects, therefore, we aimed to study lymphocyte subgroups of cvid patients and to compare the patterns with the clinical presentation in these patients. six adult patients with cvid diagnosis were studied. lymphocyte subpopulations were determined by flow cytometry. for b-cells subgroups cd , cd , igm and igd reagents w e r e u s e d . a l l l y m p h o c y t e s w e r e g a t e d f o r f i n d i n g cd + cd + memory b cells and from this population switched (cd + cd + igd -igm -) and non-switched (cd + cd + igd + igm + ) memory b cells were counted. all patients had normal levels of total lymphocyte count and absolute counts of cd + , cd + and cd + /cd + cells were also normal in all patients. only one patient showed low levels of cd + cells levels. according to paris classification scheme the patients could divided into two subgroups: mb and mb . although almost all our cvid patients had normal number of total b cells, most of them showed reduced number of memory b cells and/or switched memory b cells. all of our patients had very low or absent level of class-switched memory b cells, therefore can be possible associated with a higher risk of granulomatous disease and splenomegaly. detailed investigation of b-cell phenotypes can better characterise cvid patients and can provide more information about possible clinical outcome. background: common variable immunodeficiency (cvid) is one of the most frequent symptomatic primary immunodeficiencies, often related to spectrum of infectious and autoimmune diseases. in cvid patients wide spectrum of gastrointestinal disorders, including infections, are frequently seen. inflammatory bowel disease, helicobacter pylori infection, giardia lamblia infection, campylobacter or salmonella infection have been reported. however, abnormal liver function test and liver disease are found in approximately % of cvid patients. case history: a -year old female patient was admitted to infectious disease department due to recurrent pneumonia and purulent rhinosinusitis. blood analysis confirmed panhypogammaglobulinaemia with impaired responses to vaccinations, elevated liver function tests and anti-hcv positivity, interpreted as old and passed infection. due to cvid intravenous immunoglobulin substitution was started. however, liver function tests remained elevated and with hcv-rna analysis high hepatitis c viral load was detected. chronic hepatitis c virus infection was diagnosed and treatment with peginterferon α- a and ribavirin was started. conclusion: our case emphasizes the need for hcv-rna and hbv-dna analysis in patients with hypogammaglobulinaemia, as the serological detection is impaired and prognosis for chronic hepatitis in immunodeficiency patients is poor. outpaitent clinic of clinical immunology and allergology, east tallinn central hospital, tallinn, estonia background: primary antibody deficiencies are the most frequent primary immunodeficiencies. recurrent respiratory tract infections may result in permanent lung damage in - % of patients, most commonly presenting with the development of bronchiectasis. we aimed to evaluate the lung function and radiographic pulmonary changes in our patients with primary antibody deficiency. material and methods: we reviewed the records of adult patients with a confirmed diagnosis of primary antibody deficiency at our clinic. patients were included in this analysis in whom ct scan was performed during the last months, comprising patients with cvid and two with igg subclass deficiency (median age years; range - years; % females). all patients were on regular immunoglobulin replacement and one of the patients was on prophylactic antibiotic at the time of analysis. mean trough levels of igg were calculated based on the results measured during the last months prior to ct scan. the spirometry was performed according to published protocols. results: all patients demonstrated normal spirometry data based on fev and fvc. two patients had slightly lower mmef rates, however, the changes were not associated with higher rate of infection nor changes in ct scan. none of our patients had bronchiectasis or atelectasis. among parenchymal changes fibrotic lines were most frequently detected. in two patients ground glass due to fibrosis was noted. mean immunoglobulin trough levels in our patients were between . - . g/l, with the median of mean trough levels of all our patients . g/l. when comparing the trough levels to lung function and ct scan results, no significant associations were seen. conclusion: no remarkable changes in lung function or chest ct scan in our patients with primary antibody deficiency were noted. as regular immunoglobulin replacement therapy could have prevented the development of permanent lung damage. rationale: patients with primary immunodeficiencies (pi) (n = ) were treated subcutaneously (sc) with immunoglobulin g (ig) preceded by recombinant human hyaluronidase (ighy) at or week intervals based on their previous intravenous ig (igiv) dose. we report data for a subset of patients aged ≥ years from the final efficacy, safety and tolerability data of a pivotal phase trial of ighy. methods: patients received igiv for months at prestudy doses and frequencies. subsequently, ig % was administered sc, at % of the weekly equivalent of the iv dose, following rhuph infused through the same sc needle at a dose of u/g igg. after a ramp up from a -to a -or -week dose interval, patients received ighy every weeks for months. the primary efficacy endpoint was the mean rate of validated acute serious bacterial infections (sbis) per patient-year during the efficacy period. results: fifty-nine patients received ighy infusions; . % were completed without administration changes due to tolerability concerns or adverse events (aes). median infusion sites/month was . . the temporally associated systemic ae rate was . /infusion (ighy) vs. . /infusion (igiv). the local adverse drug reaction rate was . /infusion. the annual sbi rate was . and . /patientyear for all infections. conclusion: in adults with pi, ighy was effective in preventing infections. the majority of patients received full -to -weekly doses of ig using a single sc site with good local and systemic tolerability. rationale: in a pivotal phase trial of facilitated-subcutaneous (sc) infusion of human immunoglobulin g (igg), %, and recombinant human hyaluronidase (rhuph ) (ighy) in patients with pi, rhuph permitted most patients to have a single-site infusion (every - -week igg dosing) with bioavailability and infusion rates comparable to intravenously administered igg (igiv). we report the final analysis of the long-term extension of the initial phase study, with a duration of up to years of treatment with ighy plus additional follow-up. methods: sixty-six patients who completed the initial phase study enrolled in the extension study. patients continued their pre-study ighy dose/frequency every - weeks. after months, some patients switched to -week dosing to evaluate effects of shorter ighy interval on trough igg levels. from the final analysis, tolerability and safety after up to years of treatment were evaluated. the ighy part of the extension study was followed by a - week observation period during which patients received igg % administered iv, or sc weekly without rhuph . results: in the extension study, patients were treated with ighy and discontinued prior to safety follow-up. following discontinuation of rhuph , patients switched to follow-up. no patients withdrew due to ighy-related reactions (adrs). no serious adrs related to ighy were reported. the maximum ighy exposure for the initial and extension studies combined was years (total exposure = . patient-years; n = ighy infusions); during this time, there were no clinically observable long-term changes in the skin or sc tissue. the rate of temporally related systemic adverse events (aes), excluding infections, was . / infusion. the rate of all local aes was . /infusion. of the ighy infusions administered in the extension study, . % had no administration changes (rate reduction, interruption or discontinuation) due to tolerability concerns or adverse events. the annual rate of all infections under ighy treatment was . /patient-year. reducing the dosing interval from to weeks (same monthly dose) resulted in a % increase in trough igg levels. thirteen patients had at least non-neutralizing anti-rhuph antibody titer of ≥ : with no associated aes; no patients had neutralizing anti-rhuph antibodies. conclusions: in the extension study, ighy was well tolerated and effective, with no serious adrs for treatment periods up to years. over a maximum -year ighy exposure (for an individual patient) in the initial phase and extension studies combined, no long-term changes in skin or sc tissue were observed. the rates of infections and adverse reactions were stable or decreased over the course of the two studies, suggesting no increased risk with continued exposure to ighy. rationale: we report interim analysis of safety, tolerability and pharmacokinetics (pk) of igsc % in patients with primary immunodeficiencies (pi) aged ≥ years in europe. methods: epoch : igsc % or intravenous ig % (igiv) administered at pre-study doses every months. epoch : igsc % administered time per week for months at epoch doses. serum igg trough levels are maintained at > g/l. the primary endpoint is validated acute serious bacterial infection (sbi) rate. results: at the interim analysis in october , patients started the study. during igsc % treatment (n = ), acute sbi episode (pneumonia, moderate in severity) was reported. the infection rate per patientyear was . (igsc %). there were no serious adverse events considered related to any treatment. the rate of local adverse drug reactions (adrs) was . /infusion and all were mild in severity; no severe systemic adrs were reported with igsc %. of igsc % infusions, only . % required slowing or interrupting the administration rate. mean serum igg trough levels were . g/l (igsc %, n = ), . g/l (igiv -week interval, n = ) and . g/l (ivig -week interval, n = ). conclusion: igsc % provided an effective and well-tolerated therapy, with no dose adjustments needed from pre-study ig dose. this study is ongoing to confirm results over months. it is well known, that intravenous immunoglobulin (ivig) is the main therapeutic modality in b-cell primary immunodeficiencies (pid), it decreases mortality and morbidity in these patients dramatically. yet, it is also well known that all ivig products are very expensive, especially considering life-time use and almost normal life expectancy in these patients, if treated correctly. irregular treatment and problems with insurance/state coverage of ivig in some countries stems from this. goal: the goal of our study was to compare medical and other costs, related to the disease in patients with humoral pids with or without ivig treatment. patients: patient with b-cell deficiencies ( % x-linked agammaglobulinemia, all genetically confirmed, % common variable immunodeficiency). the age of patients varied from to years. methods: we analyzed medical and other disease-related costs during years preceding the diagnosis (without ivig therapy) retrospectively and during years on regular ivig therapy. the costs included those incurred by the state (hospitalization, home visits, emergency calls) and by the parents (costs of drugs, private consults, etc). the state costs caused by parents missing work were also considered. for standardization purpose for calculation we used the prices for the end of . results: the patients analyzed fell into different categories: . the st group -( % of patients studied) -patients with several severe infections before therapy, some chronic conditions as a result of those. age of diagnosis varied from years to years (with average time to diagnosis years). in this group costs of ivig treatment were , times higher than before the diagnosis (fig. ). nd group( % of all). patients with late diagnosis (average time to diagnosis years), who had multiple severe infections before the ivig treatment and acquired serious chronic lung complications due to it. in this group the costs before the treatment were higher, than on ivig treatment. . very early diagnosis -within the first year of life (mostly because of preceding family history) ( %). the comparative analysis was not possible, but it was noted that these patients had no history of serious infections before of while on ivig therapy. the only additional costs, besides ivig, were related to bad venous access, requiring occasionally day in-patient hospitalization for ivig infusion. as expected, in all groups, the number of infectious episodes, the number of hospitalizations (fig. ) and doctor visits (fig. ) after beginning of regular ivig treatment dropped dramatically. we also followed patients with xla, who did not receive ivig therapy because of social aspects. both patients died from severe infections. we evaluated this fact in economical prospective: in russia one worker, who works continuously and retires at years of age brings about mln roubles into the state budget (when costs for schooling and routine medical care are subtracted). if one supposes that an xla patient have been diagnosed very early in life, did not form complications prior to therapy and was on regular ivig therapy for life, this sum equals to years on ivig. conclusions: regular ivig therapy not only leads to reduction of infectious episodes, hospitalizations, and as a result improved quality of life. in some cases it even brings down the disease-related costs, incurred by the medical system and the family, and is economically advantageous for the state. introduction: replacement of immunoglobulins is a standard therapy for patients with primary immunodeficiency disease (pidd) characterized by primary antibody deficiency (pad). this poster represents our clinical experiences of initiation of home-based treatment with subcutaneous immunoglobulin (scig) with the patients diagnosed with primary variable immunodeficiency (cvid). case report: the patient (age ) has been treated at our immuno-allergy outpatient clinic since with the diagnosis of hypogammaglobulinemia (igg, iga, igm) with normal b cell count, withsusp. cvid. with the repeated administration of intramuscular and intravenous immunoglobulins (ivig, imig) repeatedly occurred serious adverse reactions, which resulted in discontinuation of the replacement therapy. in february the health condition of the patient worsened due to recurrent bacterial respiratory infections. there was a progressive decrease of serum concentrations of immunoglobulins (igg , g/l, igm , g/l, iga , g/l). the patient was admitted to the intensive care unit of the st internal department, university hospital bratislava, for a subcutaneous immunoglobulin replacement trial. despite serious adverse reactions with previous administration of several types of immunoglobulins, there have not occurred any clinically relevant side effects. conclusion: compared with im or iv formulations and administration, for selected patients, scig is better tolerated, clinically efficacious, safe, and appreciated by the patients. background: common variable immunodeficiency (cvid) is primary immunodeficiency (pid) classically viewed as antibody deficit. although, cvid is considerd to be a humoral immunodeficiency, approximately % of cvid patients have low t-cell counts or abnormal tcell function. despite adequate immunoglobulin replacement patient morbidity and mortality is variable and a number of complications are not those typically seen in pure antibody pid e.g. xla. so, t-cell rather than b-cell phenotype could determine outcome in patients with cvid. many patients with cvid have clinical history suggestive of allergic respiratory disease, but prevalence of asthma and role of atopy have not been well established. apart from recurrent infections and their sequelae, cvid patients suffer from other disease-related complications in up to % of the cases. about % have onset before the age of years. aims: ) to present one more case of tadolescent with cvid and allergic asthma, ) emphasise ultimate need of collaborative network of primary immunodeficiency centers. case report: parents of , y boy were sure that "something was wrong" with their son and were seaking for problem solution for many years. since age , child had frequent respiratory infections. adenoidectomy and tonsillectomy were performed at age . sinuitis was diagnosed several times. boy was complaining of fatique for a long time. last several years, his main problems were fever (max c) usually lasting days till weeks, accompanying running nose, coughing, conjunctival problems; intermittently headache (lasting for a few hours till all day). oral aphtae were present almost every two weeks. he was incompletely vaccinated (bcg, and once diteper). morbilli and varicellae infections passed without complications. in jan he was diagnosed as allergic asthma in sarajevo (allergy to pollen, soya, nuts and antibiotics ("ceclor" and "pancef"). in february was admitted at children's hospital sarajevo for suspected primary mmunodeficiency. he had slightly lower levels of igg and iga (twice measured), normal ige and decreased number of t helper ly. due to suspected pid, boy was checked up in two nearest regional pid centers : hypogammaglobulinemia was confirmed (igg , , iga , , igm , , as well as deficiency of igg , igg and igg ). flow cytometry showed slightly raised concentration of lymphocites b (cd ), slightly raised number of nondifferentiated b cells. cvid was suspected but not proved. in july child visited center for primary immunodeficiencies in munchen, germany, were he was diagnosed ad probable cvid on the basis of hypogammaglobulinemia, lower levels od switch memory b cells, normal number of t-cells, positive antibodies to vaccinations and overcome infection (morbilli, varicellae). allergic asthma was additionaly confirmed in specialised pediatric pulmology hospital in germany (abnormal spirometry, normal ige, positive skin prick test, abnormal fractional exaled nitric oxid test, incipient brochiectasies due to asthma confirmed by high -resolution lung ct scan). low human immunoglobulin replacement was started ( mg/kgbw) as well as antiasthmatic therapy (inhalatory steroids, antihistaminics). excellent therapeutical response were achieved : after one , y follow up, we can confirm patient has excellent general condition, no subjective symptoms, no tiredness, no severe infections. conclusion: diagnosing cvid is challenging task and quite often could be "per aspera ad astra". there is ultimate need of collaborative work of primary immunodeficiency network aimed of diagnosing patients on time. cvid patients with history suggestive of allergic asthma, are negativne on traditional tests, additional test designed to identify allergic asthma might be conducted. common variable immune deficiency (cvid) is a heterogeneous syndrome characterized by hypogammaglobulinemia, recurrent infections, immune disregularity (autoimmunity, autoinflamation) and propensity to malignancies. in the us report, . % of cvid patients had a lymphoid malignancy, and cancers of other sorts developed in % of patients. it is not clear why cvid patients have higher risk of malignancy but chronic antigenic stimulation, chronic inflammation and increased chromosomal radiosensitivity may be the cause. cvid patients with higher igm level, reduced or absent b cell numbers, cd t cells lower than and pli phenotype have higher prevalence of malignancy. allergy and clinical immunology department of rasool e akram hospital has registered cvid patients. mean age of the onset of cvid symptoms was . years. mean diagnosis age was . years with mean diagnostic delay of years. mean follow up time was years (minimum . -maximum y). malignancy occurred in the follow up of patients ( %). one patient had two different malignancies (breast cancer and gi adenocarcinoma). malignancy risk per case was %. hodgkin's lymphoma was the most common type ( % of cancers common variable immune deficiency is a heterogeneous syndrome characterized by hypogammaglobulinemia , recurrent infections , auto immunity and auto-inflammation . more than % of cvid patients have auto immune complications and among them, auto immune cytopenia is the most common. cvid patients with higher igm levelhigher low b cellslower t reg levelslower cd /cd ratio and lower class switched memory b cells have higher prevalence of autoimmunity. allergy and clinical immunology department of this hospital has registered cvid patients. mean age of onset of cvid symptoms was . years. mean diagnosis age was . years with mean diagnostic delay of years. mean follow up time was years (minimum . maximum y). autoimmunity was detected in cases ( %) and cases ( %) had more than one autoimmunity. autoimmunity was the first symptom of cvid in percent of cases. autoimmune disorders should be considered in the follow up of cvid patients. igg is major immunoglobulin and classified subgroups as igg , igg , igg and igg . igg and igg subclasses are rich in antibodies aganist proteins such as the toxins produced by the diphtheria and tetanus, as well as antibodies aganist viral proteins. recurrent ear infections, sinusitis, bronchitis and pneumonia are common in ig g subclass deficiency. ig g is the major subclass of ig g. igg subclass deficiency is very rare. chronic eosinophilic pneumonia is one of the eosinophilic lung disease and is seen rarely. in the presence of peripheral eosinophilia and radiological pulmonary infiltrates diseases suspected. when increase in the number of eosinophils in bronchoalveolar lavage fluids and/or presence of eosinophils in lung tissue diagnosis is confirmed. according to different recording systems chronic eosinophilic pneumonia is % - of the interstitial lung disase. there is not any criteria for diagnosis but also diagnoses is confirmed with suspected findings. symptoms inludes that: )in the presence of respiratory symptoms for two weeks long )eosinophilia at alveolar lavage and\or peripheral blood ( bal fluid cytological examination > % , blood > /mm ) )radiological imaging of the lung peripheral infiltration )exclusion of the other causes of eosinophlic lung disease there is eosinophilia over /mm nearly all patients. one of third or half of patients have diagnosed as asthma. disease begin with systemic symptoms such as night sweats, weight loss, anorexia and pulmoner symptoms such as cough, shortness of breath, wheezing. patients have restrictive or obstructive findings in pft. one of third patients, especially with history of asthma, have obstruction in pft. İn the pathologic biopsy findings include; thickening of alveolar walls and accumolation of eosinophils and lymphocytes. long time used corticosteroids treatment is recomended. relaps is common when treatment is discontinued. we present the patient who has ig g deficiency, chronic eosinophlia and % eosinophils in bronchoalveolar lavage fluid. the patient improved long time used oral steroid then inhaled steroids. this was presented in terms of clinical association. case: a years old female patient who were followed due to asthma in the other center for two years, although use of combination inhaled fluticasone and salmeterol, patient was admitted with cough and sputum production. in thorax ct there were, bronchiectasis at right lower lobe, pneumonic consolidation in the right lower lobe and ground glass opacities. we detected as igg mg/dl ( - mg/dl), iga mg/dl ( - mg/dl), igm mg/dl ( - mg/dl), ige . mg/dl, igg mg/dl ( - mg/dl), igg mg/dl ( - mg/dl), igg mg/dl( - mg/dl), igg mg/dl ( - mg/dl). because eosinophilia ( cells) and symptoms continued, bronchoscopy was performed. left main bronchus was normal, right bronchus were seen dilated. purulent secretion was aspirated on rigt bronchus with flexible bronchoscopy ..in cytological examination % eosinophils was detected in bal. bronchoalveolar lavage cultures was negative. the patient was diagnosed chronic eosinophilic pneumonia and mg/ kg oral steroid was began. ivig was given up to patient; because of frequently recurrent sinopulmonary infection and patient had igg subclass deficiency. in the third month of oral steroid therapy physical examination findings and pft were improved and started inhaled steroid. conclusion: immunodeficiencies often can be seen alone. although, the pathogenesis of immundeficiencies with lung disease is not well understood and associated with interstitial lung disease. therefore, investigations must be include bronchoscopy and bronchoalveolar lavage. scid is a congenital heterogeneous group of diseases characterized by severe impairment of t,b, nk cell development and function. the hematopoietic stem cell transplanted patients with scid in the time period from to were evaluated. male/female ratio is / . median follow-up time is years ( months- years). parental consanguinity ratio was , %. mean age of onset of the symptoms was ± , months. most common clinical findings on admission were; pneumonia ( , %), moniliasis ( , %), diarrhea ( , %), dermatitis ( . %). classification according to t, b, and nk cell counts; (figure ). molecular genetic defects were determined in patients (figure ) are given. there is no difference between age groups according to occurrence of acute and chronic gvhd. death ratio increases with the increasing age. acute and chronic gvhd and number of deaths were significantly higher in peripheral stem cell transplanted patients. there is no statistically significant difference in occurrence of acute/chronic gvhd between b-and b + scid, who had been hla idantically transplanted from family donor. both groups have similar ratios of ivig treatment need after hsct. there is no statistically significant difference in occurrence of acute/ chronic gvhd between nk-and nk + scid, who had been hla idantically transplanted from family donor. both groups have similar ratios of ivig treatment need after hsct. death ratio was similar in groups. bcg dissemination in bcg vaccinated patients is significantly higher in the nk + group. the rate of complications due to severe infections is high and increases with age in patients with scid. hsct is curative, should be considered as early as possible. trec analysis for neonatal screening will give chance for early diagnosis and treatment of the patients. the hyper-ige syndromes are rare pids and are characterized by atopic dermatitis, skin abscesses recurrent pneumonias, elevated serum ige levels and sometimes mucocutaneous candidiasis. we have been evaluating and following up a group of patients with features suggestive for autosomal recessive hyper ige syndrome (ar-hies). homozygous dock mutations were identified in several of these patients. however, two siblings from a consanguineous family in this group of patients showed homozygous block in chromosome p . in homozygosity mapping which includes stk gene. sanger sequencing was performed for stk deficiency and showed a novel c. - delgata mutation in stk gene causing a premature stop codon. clinical manifestations of stk deficiency, also known as a macrophage stimulating (mst ) deficiency, stk deficiency comprise recurrent and severe viral skin infections including molluscum contagiosum and warts, fungal and bacterial infections and autoimmunity which are also the features of ar-hies. our patients's main features include autoimmune cytopenias (aiha and itp), cutaneous viral (molluscum contagiosum and mild perioral herpetic lesion), mild atopic dermatitis, seborrheic dermatitis, lymphopenia (particularly cd lymphopenia), and intermittent mild neutropenia. serum ige level was mildly high in these patients. our results indicates that patients that show clinical phenotype of ar-hies needs to be also evaluated for stk deficiency. determination of the underlying defect and reporting the patients are required for the description of the phenotypic spectrum of pids and the frequency of these diseases as well as for genetic counselling. mendelian susceptibility to mycobacterial disease (msmd) is a rare entity. patients with msmd have susceptibility to salmonella and some other intracellular microorganisms in addition to weakly pathogenic mycobacterial species. il- rβ deficiency, most common form of msmd, is caused by mutations in the il rb gene. patients who have symptoms suggestive for msmd or history of sibling death due to bcgosis were evaluated. the expression of the il- rb was detected in patients and family members by monoclonal antibodies on the lymphocyte surface by flow cytometry after the lymphocytes were stimulated in vitro with pha. mutation analyses was done by sanger sequencing. all index cases were presented either with bcg or salmonella infection. two patients, though they were bcg vaccinated had no clinical symptom, six presented with the symptoms of salmonella infections, two developed leukocytoclastic vasculitis, candidiasis was the accompanying feature in seven. recurrent leishmaniasis that necessitated subcutaneous interferon-γ and prophylactic amphotericine b therapy was present in a patient. in all patients the percentage of lymphocytes with il rβ expression was found to be less than % . prophylactic antimycrobial treatment and in severe and resistant infectious episodes if-γ therapy, should be given. many patients have associated mucocutaneous candidiasis. the prognosis is good unless the patient admits at the later stages of bcg infection. the results of our patients showed that the analysis of the surface expression of il rβ on activated lymphocytes is an effective diagnostic method which can also be used in screening of the patients with probable msmd. ataxia-telangiectasia (a-t) results from the loss of ataxia-telangiectasia mutated gene (atm) function and is a heterogeneous, multisystemic disease and characterized by accelerated telomere loss, genomic instability, progressive neurological degeneration, immunodeficiency, premature ageing and increased neoplasia incidence. even in classic a-t with ataxia and telangiectasia, the onset of clinical symptoms and the rate of progression are variable. here, we report two siblings was diagnosed as a-t with severe and early hypogamaglobulinemia, decreased cd , increased cd ro, no ataxia and telangiectasia, purulent otitis and hemophagocytosis that harbor a rare frameshift mutation of atm gene. Özdemir Öner ; bozdoğan sıla department of pediatrics, division of allergy/immunology, sakarya university, medical faculty, adapazarı, türkiye. department of pediatrics, sakarya university, medical faculty, adapazarı, türkiye. background: the acquisition of new food allergy after transplantation or transplant-acquired food allergy (tafa) is usually reported in adults and rarely in children. tafa is described mainly after liver, but also after small bowel/intestinal, lung and heart transplantations. in different studies, the male/female ratio is equal. literature data suggest that children with tafa typically present within the first year after surgery and they are typically allergic to multiple foods. aim: tafa is generally characterized with allergy to multiple foods and increased level of total and/or spesific ige. here, a patient although who had normal total. ige and specific ige test results, he developed reaction to skin prick test for cow's milk is presented and his clinical presentation will be discussed. case presentation: month-old-boy came to our allergy clinic with complaints of vomiting after drinking cow's milk and skin rush on the area where contact ed with chocolate. in his past medical histroy, left lateral segment of liver (donor was his mother) was transplanted to him when he was at months. the liver donor was not recorded as having a history of allergic disease. methylprednisolone and tacrolimus immunosuppression were used after the transplantation, and tacrolimus therapy was continued for prophylaxis of chronic rejection. when he was at months, family fed the patient with cow's milk but hours later he began to vomit. he vomited five times in two hours. then, he developed constipation. rectal irrigation was used. then oral intake stopped for two days. he was thought to be having food protein induced enterocolitis. his vomiting complaints repeated after intake of formula and baby food which include grain. so he fed with special formula including short-chain peptides and free aminoacids and his symptoms improved. past medical history: extrahepatic biliary atresia was diagnosed at weeks age with conjugated hyperbilirubinemia (according to scintigraphy and biopsy results). family history: his father has penicillin allergy and his aunt has asthma. at our outpatient clinic: height and weight were within normal percentiles. physical examination reevaled normal examination findings. laboratory findings: wbc was . /mm , with % neutrophils, % eosinophils and % lymphocytes. his hemoglobin was . g/ dl, platelet count was . /mm . background: except for antibody deficiency and complement defects, hematopoietic stem cell transplantation (hsct) is the single best curative treatment defined for primary immunodeficiency (pid) so far. in the current study, we aimed to assess the role of pid type, donor type and clinical status on hsct success rates. materials and methods: we retrospectively reviewed the records of a total of hscts procedures performed in patients diagnosed with pid between and , in ankara university pediatric allergy and immunology department. results: of the patients, had severe combined immunodeficiency (scid) and had non-scid. the survival rates following hsct, in both scid and non-scid patients were %. when classified according to the source of donor, patients who had a hla-matched sibling donor (msd) in the scid group had . % survival rate post transplantation, those who had a matched related donor (mrd) had . % and those who received a haploidentical donor had . % survival rates. in the non-scid group there were patients with haploidentical transplants ( omenn syndrome and mhc class ii deficiency) and all patients died. we assessed several potential risk factors associated with survival in scid patients. patients diagnosed over months of age with a pre-existing pulmonary infection, requiring intensive care and/or mechanical ventilation had significantly lower survival rates. conclusion: hsct is the best curative treatment for pid. our results demonstrated that hsct performed from matched family donors or even haploidentical parents is a lifesaving treatment in various types of pid's, especially in scid. introduction: in case of donor availability allogeneic hematopoietic stem cell transplantation (hsct) can be regarded as a definitive therapy for a variety of primary immunodeficiency syndromes (pids), including severe combined immunodeficiency (scid) and non-scid pids. study period: we retrospectively reviewed the hospital records of consecutive children with pid, who had allogeneic hsct in the last years, between january and january . our median follow-up time is , years ( months - , years) patients and methods: the median age of children at hsct was , months ( , months- years). boys/ girls diagnoses were based on anamnestic data, clinical findings, and immunological and genetic analysis. conditioning regimens included busulphan + cyclophosphamide, busulphan + cyclophosphamide + atg, fludarabine + melphalan or fludarabine + anti-cd in b-, t-, nk-scid cases conditioning was not used. indications for transplantation: patients' diagnoses were severe combined immunodeficiency (n = ), wiskott-aldrich syndrome (was, n = ), chronic granulomatous disease (cgd, n = ), xlinked lymphoproliferative disease (xlp, n = ), whim-syndrome (n = ), hyper igm syndrome (cd ligand deficiency, n = ), leukocyte adhesion deficiency (lad, n = ), dock mutation (n = ) transplantations: hscts for children were performed. patients were retransplanted ( pt once, pt twice, pt times), because of rejections. at the first hscts in / cases sibling bone marrow, / sibling peripheral blood, / sibling cord blood, / unrelated cord blood, / unrelated bone marrow, / unrelated peripheral blood, / haploidentical donors were used median cd count was , x /kg ( , x - , x /kg) patients engrafted on median ± day (anc > , g/l) acute gvhd occured in / cases ( pathomorphological findes: distortion of the structure of lymph nodes due to enlargement of paracortical zones and follicules was found in all patients. presumed atypical cells (makrolymphoblastes and berezovsky-sternberg-reed cells), were detected in patients overwise during follow-up for - years in these patients revealed no specific infectious and malignansy. accumulation of proliferating dn cells (a lot of mitoses) was characteristic fiture in lymph nodes and spleen of alps patients. setting of plasma cells in follicular zones in the lymph nodes was reveald in cases, and eosinophilesin , pronounced immunoblast proliferation in , sinus histiocytosisin . multiple hyperplastic follicles, extended sinuses with many phagocytizing macrophages, lymphocytic infiltration was revealed by histological examination of the spleen in all cases. immunohistochemistry findes: t-and b-cells proliferation (ki- expression), atypical location of lymphocyte populations, settings of plasma cells in all lymph node zones. in paracortical zones was found cd +, cd +, cd + cd -, cd + cd -cells in patients, cd +, cd + cells in patient; and in restricted follicular zones -cd +, cd +, hla-dr + cells in patients, cd +, cd + and cd + in patients. the presentation of a case of kimura disease in a patient with was. the was was diagnosed in year months, based on infectious syndrome, atopic dermatitis, hematological syndrome -skin hemorrhages and thrombocytopenia - * /l. after he performed often sars. regularly received ivig at mg/kg, antibiotic therapy with a positive effect -infectious syndrome stopped, controlled hemorrhagic syndrome. since there was a herpes virus infection with frequent exacerbations on the face. patient received acyclovir, valacyclovir, famvir with a temporary effect. in . on the left side of the face -periorbital region, eyelids, malar region appeared the site of soft tissue hyperproliferation - cm in diameter with a thickness of - cm, with moderate moist, painful on palpation, the left eye was closed, the lid margin ciliated dramatically thickened, deformed. in october-november was admitted in diagnostic department, massive antibiotic, antimycotic, antiviral therapy, ivig therapy. the histological study showed the angiolimphoid hyperplasia with eosinophilia without immunomorphological signs of malignant tumor growth. based on histological, immunohistochemical studies of the skin, these clinical and laboratory findings, diagnosis: mass lesion of the left face -kimura disease. patient helded courses of chemotherapy with positive dynamics: mass lesion decreased, erosive surfaces disappeared and now receives romiplostimum, acyclovir, biseptolum, ivig . g/kg time per month. under the observation there is a family k., which is unique in the deletion variants of the same region q. . and the presence of an adult patient with the di george syndrome. clinical manifestations of the syndrome in family members is differ. methods: clinical, laboratory, instrumental and genetic ( dna was isolated using a kit «qiaamp dna mini kit» and dna from dried blood spots was isolated using a commercial kit "dna-sorb-b». mlpaanalysis set salsa mlpa probemix p -b digeorge, genetic analyzer applied biosystems ). mother ( years), had few incidents of pneumonia before years. she was diagnosed ullrich-turner syndrome at age years. all childbirths by caesarean section. she has not any laboratory and instrumental findings of immunodeficiency, endocrinopathy, cardiac and thyroid abnormalities. she has deletion in the starting area of the di george region (lcr -a), including genes cltcl , hira, cdc , cldn , gp bb, tbx , txnrd , dgcr . father ( years), healthy, current smoker. the dna microstructural violations in di george region haven't been identified. son was from i pregnancy, heart defect was set prenatally, marked growth retardation. childbirth at weeks. he had unstable reduced cell parameters and hypogammaglobulinemia, the size reduction of the thymus and congenital heart disease: truncus arteriosus, dc b stage by lang. after days of life operated for congenital heart disease. during the surgical intervention the thymus wasn't found in a typical place. the postoperative period was complicated by sepsis, heart and respiratory failure. received ivig, antibiotic and antifungal therapy. he suffered from sars, severe course, the degree of respiratory failure ( months) -death. in mlpa-dna defined microdeletion disorders starting region di george (lcr -a) -from the -year-old dry blood sample (postmortem study a year old boy with bleeding, eczema and recurrent pyogenic infection was admitted to our department. the child who received treatment in hematology accidentally a few times, but has not had the effect of treatment . laboratory parameters -cd -cells- % ( ), cd -cells - % ( ), cd -cells - % ( ), cd -cells - % ( ), cd / -cells - % ( ), cd / cd - . , hla-dr- , nbt- . igg- . q /l, iga- . q / l, igm - . q / l, ige- u / ml. hb- g / l, erythrocytes- . x / l, leukosytes - . x / l, neutrophils- %, lymphocytes - % ( ), eosinophils- %, monocytes- %, metamielosytes:- %, plateletsrare was observed.normal bone marrow cells, the separation of platelets from meqakariosyt has become weak. the advantage of erythroid unordered have been observed. bone marrow puncture the mielokariosytes - . x / l, meqakariosytes - . x / l, blasts- . %, myelosytes - . %, metamyelosytes - . %, seqments - . % neutrophils-% . , eosinophils- . %, lymphocytes- . %, monocytes- . % erythroblasts- . %, pronormoblasts- . %, normoblasts- . % meqakariosytes - . %, plasmatic cells- . %.his younger brother with eczema, pyogenic infections (furunculosis) was admitted to our department. cd - % ( ) ataxia telangiectasia (a-t), is a genetic disorder caused by the homozygous mutation of the atm gene and, frequently associates with variable degrees of cellular and humoral immunodeficiency. however, the immune defects in patients with a-t are not well characterized. to our knowledge, there is no work on major lymphocyte subpopulations and recent thymic emigrants of a-t patients comparing to age-matched healthy controls. according to esid criteria, patients diagnosed as a-t and agematched healthy children were assigned to the study. both patients and healthy controls were grouped as - , - , - years and older than years. using flow cytometer, major lymphocyte subpopulations and cd + cd ra + cd + recent thymic emigrants (rte) were determined as per cent and absolute cell numbers and compared. no significant differences regarding all lymphocyte subpopulations were observed between age groups of a-t. comparing to the healthy controls, there were a decrease (in t cells, effector memory t cells, b cells, naïve b cells, switched b cells and rte) and there were an increase (in active t cells, naïve t cells and nonswitched b cells) in the absolute number and percent of some cell populations in the a-t group. findings of this study showed effector functions in some cell lymphocyte populations were decreased and could be thought that bone marrow of patients should be tried to increase the cells numbers. however, the study has an important limitation about patient and healthy population. ataxia-telangiectasia (a-t) results from the loss of ataxia-telangiectasia mutated gene (atm) function and is a heterogeneous, multisystemic disease and characterized by accelerated telomere loss, genomic instability, progressive neurological degeneration, immunodeficiency, premature ageing and increased neoplasia incidence. even in classic a-t with ataxia and telangiectasia, the onset of clinical symptoms and the rate of progression are variable. here, we report two siblings was diagnosed as a-t with severe and early hypogamaglobulinemia, decreased cd , increased cd ro, no ataxia and telangiectasia, purulent otitis and hemophagocytosis that harbor a rare frameshift mutation of atm gene. nijmegen breakage syndrome(nbs) is a rare autosomal recessive disease usually presenting at birth with microcephaly. here we present a case of nbs diagnosed at the age of twenty seven who admitted to our outpatient clinic with malaise, loss of appetite, weight loss and dyspnea on effort . she was hospitalized in another centre years before because of pneumonia, pancytopenia, generalized lymphadenopathy, hepatosplenomegaly and hypogammaglobulinemia (very low igg and iga) where she couldn't have any definite diagnosis. her past medical history was remarkable for primary amenorrhea and basal cell carcinoma excision from preaericular region when she was . she had no severe infection history before except frequent upper respiratory tract infections. her parents were consanginous and she had a brother died at months of age. microcephaly together with short stature, multiple palpable lymphadenopathies , splenomegaly and absent secondary sexual characteristics were prominent features in physical examination. direct fluorescence sequencing of the nbn gene showed a homozygous mutation in exon (c. _ delacaaa) which confirmed the diagnosis of nbs in our patient. nbs is mostly diagnosed in childhood. the delay in diagnosis was partly due to the lack of severe infections in her past medical history until she was . another factor that lead to the delay is that it is an unknown disease among physicians in turkey. the longest known survival is years in a patient who had no clinical features of nbs other than primary amenorrhea. as a result, our patient is one of the oldest patients reported in the literature presenting nearly all of the classical features of the disease and carrying the most common pathologic mutation. wiscott-aldrich syndrome (was) is a rare x-linked recessive immunodeficiency disorder characterized by thrombocytopenia, small platelets, eczama, recurrent infections and an increased risk of autoimunity and malignancy. the gene responsible for was is located in chromosome xp . -p - , which consists of exons, and encodes a -amino acid protein. in this study, we aimed to screen was gene mutations and analyze the effects of determined mutations in boys with non-classical was phenotype. ivs + g > a gene alteration in intron of was gene was identified in case , previously. so, rna isolation and then cdna synthesized was carried out. in case , after amplification of exons of was gene by pcr, the amplicons were sequenced. in this patient, g > c alteration was detected in the first base of intron . afterwards, cdna synthesized for detecting the splicing effect. based on the gel image results, cdna found to be base pairs smaller than the normal in case . in case , g > c alteration was detected in the first base of intron and then we determined multiple splicing products. two different splicing mutations (ivs + g > a and ivs + g > c) were detected in two cases with non-classical phenotype. ivs + g > a splicing mutation was stated in the literature, previously, but ivs + g > c mutation was first time identified in this study. whim is rare (< / ), heterozygous, autosomal dominantly inherited pid, caused by mutations in the gene encoding for the chemokine receptor cxcr , mapped on q locus. the altered cxcr / cxcl interaction impairs cellular homeostasis and trafficking, resulting in immunological dysfunctions with abnormal retention of mature neutrophils in the bone marrow (myelokathexis) and consecutive severe neutropenia, variable degree of lymphopenia and hypogammaglobulinemia. whim patients suffer from recurrent bacterial infections since early childhood and later on manifest a specific susceptibility to hpv infections with developing widespread warts. because of rarity of the disease, heterogeneity in clinical presentation and usually incomplete phenotype, the diagnosis is often delayed and whim syndrome is not suspected. a year old boy who is suffering from recurrent bacterial infections, often complicated with bronchopneumonia, with severe neutropenia, but also, with lower level of lymphocytes and still, normal serum immunoglobulin level is presented. he has no developed warts; neither his parents nor relatives have warts. at the age of years was unveiled the diseasecausing mutation in the cxcr gene (c. c > t; p.r x; heterozygote). severe congenital neutropenia accompanied with lymphopenia and findings of mature neutrophils in bone marrow, might be an easy approach for getting closer to the clinical diagnosis of whim syndrome. early identification is important for clinical and therapeutic management, allowing a more comprehensive follow-up and administration of appropriate therapy. we report two cases of congenital heart disease (tetralogia fallot and interruption aortic arch) with confirmed microdeletion chromosome q . by karyotype and fluorescence in situ hybridization analysis (fish). children underwent surgical correction of congenital heart defects with good postoperative outcome, although were complex. the phenotype of these patients can be extremely variable, frequently leading to clinical confusion, diagnostic delay, excess morbidity, early mortality. identification of these patients is essential for their adequate management and genetic counseling. a multidisciplinary approach is fundamental to ensure that the patient will be able to attain his or her maximal potential. the underlying cause of the juvenile periodentitis is not well understood but is now thought to be related to an abnormal immune system and to invading bacteria in the cementum of the teeth. instead painful fissures and recurrent pyogenic infections of the skin seem to be the most common medical complications. however, a number of pls patients with abscesses or pseudotumors of the liver have been described. there have been reports of pls patients with other stigmata such as growth retardation, non-symptomatic intracranial calcifications and mental retardation furthermore, coinheritance of pls and albinism type has been reported. case presentation: a yr old girl admitted to our hospital with chief compliant of skin lesions since early months of birth. in the past medical history; she had skin abscess and failure to thrive. on admission she had erythematous, shiny skin with generalized dry scaly predominantly palmoplantar hyperkeratosis and loss of teeth except four or five molar teeth. she informed that she had malformed teeth since childhood which fell off one by one. she had also poor oral hygiene non-pitting edema on lower extremities. no hepatosplenomegaly detected. family history was negative. she investigated for probable immune-deficiency with regard to skin lesions, history of skin abscess and ftt the lab finding are as following: cbc diff = normal ,crp = neg, esr = , ast = , alt = , alph = , alb = . , total pr = . , urea = , cr = . igg = , igm = , iga = , ige = . , cd = %, cd = %, cd = %, cd = %, cd = %, cd = % conclusion: according to nearly normal lab values and presenting signs such as: generalized pyogenic periodentitis, palmoplantar hyper keratosis and negative family history were attributed to a very rare autosomal recessive disorder with ectodermal dysplasia known as papillon-lefevresyndrome manifesting with palmoplantar hyperkeratosis and severe early onset of destructive periodontal leading to pre-mature loss of both primary and permanent dentitions. nijmegen breakage syndrome (nbs) is a rare autosomal recessive syndrome of chromosomal instability mainly characterized by microcephaly at birth, combined immunodeficiency and predisposition to malignancies. the disease is caused by mutations in the nbs gene, which encodes nibrin, a component of the hmre -rad -p complex involved in cellular response to dna doublestrand breaks. the aim of the present case report was to discuss two siblings with immunologically and clinically different phenotypes of disease presentation and to make an attempt to explain possible genotype-phenotype relation. the two patients and their non-consanguineous parents were clinically, laboratory and genetically investigated. for mother and father we observed no clinical presentation and no immunological abnormalities. sibling no. ( years old boy) had no history of recurrent infections and no deviation in immunological tests. sibling no. . ( month old girl) had recurrent infections since birth and iga, igg and igg deficiency as well as t-and b-cells deficiency. cytogenetic analysis revealed variable percent of spontaneous chromosomal instability which was more severe (in % of chromosomes analyzed) in sibling no. . additionally we sequenced bi-directionally ( amplicons) the dna samples from all family members to survey the germline genetic variation in the nbs gene. the del (exon ) was detected in both siblings in homozygous and in both parents in heterozygous feature. in order to explain different clinical and immunological presentation of two siblings the rest of the nbs exones were analyzed for genetic heterogeneity. no additional changes were observed. in conclusion patients with the same nbs genotype may show different phenotypes. other gene/epigenetic factors seem to play a role in phenotype modulation. omenn syndrome [mendelian inheritance (omim )] is an autosomal recessive form characterized by the presence of fatal generalized severe erythroderma, lymphoadenopathy, eosinophilia and profound immunodeficiency. objective: we studied clinical and immunological presentation of the disease manifestation and frequency c. - delaa (p.k vfs ) in rag gene among eastern slavs population. results: we collected clinical and immunological data of patients ( from belarus, -ukraine, -russia) females, males. age of omenn syndrome manifestation varied from st day of life to yr month. age of diagnosis - days to year months. in patients had classical immunological phenotype t(+/-)b-nk+, pt had tlowb + nk + with cd + tcrgd + expansion. in pts had mutation in rag gene, in had c. - delaa (p.k vfs ) in one or two alleles. at present moment in pts are alive, were transplanted, pt is prepared to bmt. conclusion: this study demonstrates that the most popular genetic abnormalities in eastern slavs children with omenn syndrome is c. - delaa (p.k vfs ) in rag gene. this information may be useful for rapid diagnostic of omenn syndrome in laboratories used sscp (single strand conformation polymorphism) before sequencing. under examination the patient particularly bright phenotype attracted attention: microcephaly, "birdlike" facial features (sloping forehead, nape, hypoplasia of brow ridges, broad nasal bridge and protruding midface, hypoplasia of the mandible). in addition, besides specific anomaly of the facial bones we noted: big ears, sparse hair and clinodactyly of the fifth fingers. clinical and immunological characteristics: the feature of the case is pancytopenia syndrome we have diagnosed at the early stages of observation and which is continued throughout the period of observation. erc - . - . x /l hb - - g/l leuk - . - . x /l neu - - % ( - cells/mcl) plt - - x /l data of immunological examination: iga - , g / l igm - , g / l igg - , g / l (other results of immunological examination are without features) the deep insufficiency of antibody production in our patient was the cause of serious, recurrent, and subsequently chronic bacterial sinopulmonary infections after years old. the results of clinical laboratory and immunological examination without significant features: erc - , - , x /l hb - - g/l leuk - , - , x /l neu - - % ( - cells/ mcl) plt - x /l iga - , g/l - . g / l igm - , g/l - . g / l igg - , g/l - . g / l (other results of immunological examination are without features) the x-linked chronic granulomatous disease (cgd) is a primary phagocytic cell deficiency characterized by severe bacterial and fungal infections of various organs. we report of a years of a male patient with xlinked cgd who presented with recurrent hepatic abscesses as the sole manifestation of the disease. phagocytic and bactericidal activities of granulocytes were studied by using microbiological assays. generation of superoxide anion by blood granulocytes was measured by the ferricytochrome c reduction test. cgd is an immunodeficiency caused by mutations in genes encoding subunits of the nadph oxidase complex. normally, assembly of the nadph oxidase complex in phagosomes of phagocytic cells leads to a "respiratory burst" essential for the clearance of microorganisms. cgd patients lack this mechanism, which results in life-threatening infections and granuloma formations. the leading cause of death are pneumonia and pulmonary abscess, septicemia and brain abscess. in neurogical manifestations various pathogens have been involved including aspergillus spp., s. prolificans, a. infectoria, salmonella and staphylococcus spp. there are only some several reports on fungal brain and spinal cord infection, aspergillus abscess resembling brain tumor, meningitis due to streptococcus and candida spp. in the past years we treated children with cgd. we present the infectological challenge of an x-linked cgd patient with brain abscess. in spite of our effort we were unable to identify its causative pathogen. empiric therapy sometimes resembles polypragmasia in cgd. to decrease mortality and morbidity from fungal infections in cgd the prophylactic use of itraconazole or voriconazole is widely recommended. a relatively new azole, posaconazole is active in pulmonary and cerebral fungal manifestations , indeed may be effective against fungi with inherent resistance to ampb or voriconazole. there was no etiological diagnosis in our case that did not respond to conventional antifungal and antibacterial treatment. based upon the findings and literature data we presume the causative agent might be some kind of moulds. we suppose the use of echinocandin and posaconazole as salvage ("prophylactic") therapy has resulted significant regression of the brain abscess. the diagnosis of chronic granulomatous disease (cgd) was verified in children during the past few years in the department of clinical immunology of regional children's hospital of chelyabinsk (russia). in our opinionin casesthe diagnosis was establishedearly enough. michael w. transferred to the neonatal pathology unit of our clinic at the age of days with vesiculopustules. take into consideration our own experience of observing children with this form of primary immunodeficiency in previous years, the child was conducted immunological examination, in particular, the test of restoration nitroblue tetrazolium by superoxide anionformed when oxygen explodes in leukocyte (nitro blau tetrasolium -nbt-test). this decision is caused by the fact that previously observed children with chronic granulomatous disease, had vesiculopustules in % at birth. the survey has revealed a complete lack of production of reactive oxygen species by neutrophils in the evaluation of nbt-test, which allowed us to suggest the diagnosis of cgd. the baby was banned vaccination against tuberculosis, and after reliever vesiculopustules the boy was discharged home. however, at the age of month the baby suffered from glandular abscess and right-segmental pneumonia. in cbc there is expressed anemia (hb g/l, erythrocytes - . x /l), leukocytosis ( x /l), accelerated esr ( - mm/h). at the age of months in the university of debrecen, medical and health science center debrecen, hungary, molecular genetic studywas conducted by prof. dr. laszlo marodi. it was revealed a mutation in c. g > a in exon of gene cybb (encoding subunit gp -phox), after that the diagnosis of cgd was finally verified. it was assigned a basic preventive antimicrobial therapy by trimethoprimsulfamethoxazole, and itraconazoleto prevent fungal infections. despite this since months the child has repeatedly and consistently suffered from bilateral groin lymphadenitis during the year, acute hematogenous osteomyelitis of the left ulna, pneumonias, purulent mesadenitis, endoperitonitis. the fact that the child aged months was diagnosed sepsisis evidence of the severity of infectious complications. clinical and biochemical blood tests were distinguished by consistently high levels of esr and the presence of leukocytosis, and also severe anemia. now the child is years months, he is undergoing treatment at the department of clinical immunology, russian children's clinical hospital (moscow) after bone marrow transplantation. case history № , christina n. from the early history we know that the girl's newborn period was uneventful. she was vaccinated against tuberculosis in the nursing home (no reaction). it was noted the formation of abscesses after vaccination against whooping cough, diphtheria and tetanus in and . months. in years months she suffered from mezootit. for the first time the girl came under our observation in the children's hospital in chelyabinsk (russia) at the age of years with right segmental pneumonia, complicated by the destruction. after further examination it was diagnosed tuberculosis of intrathoracic lymph nodes to the right with upper lobe bronchopulmonary defeat. to take into consideration given above, it was conducted immunological test that revealed a complete lack of production of reactive oxygen species by neutrophils in the evaluation of nbt-test. on the basis of the history, clinical manifestations and results of immunological examination chronic granulomatous disease was diagnosed. after fourmonth period treatment a recovery came from tuberculosis and child was transferred to outpatient monitoring. in years in this family a second daughter was born -arina n. taking into account revealed immunodeficiency of her sister, the child was not vaccinated against tuberculosis on our recommendations. a six-month-old child was conducted immunological examination, which, like her older sister, also shows a complete lack of production of reactive oxygen species by neutrophils in the evaluation of nbt-test. during the molecular genetic studies of both sisters it was identified identical mutation c. - del gt gene ncf (encoding subunit p phox). now the older girl is and her sister is years old. observing them in the dynamics neither of them do not show any life-threatening infections. thus, the recovery test of nitrobluetetrazolium by superoxide anion formed when oxygen explodes in leukocyte (nitro blau tetrasolium -nbt-test) is a fairly reliable method of early diagnosis of chronic granulomatous disease. chronic granolomatous disease (cgd) is due to defective phagocyte superoxide production leading to impaired microbial killing. it is comprised of a group of five genotypes with a common phenotype, chracterized by recurrent severe bacterial and fungal infections and tissue granuloma formation. patients with cgd often present with pneumonia, liver abscess, skin infections, lymphadenitis, osteomyelitis. a five month old boy was referred to pediatric infection unit by lymphadenitis. on the medical history, he had taken antibiotic therapy for lymphadenopathy when he was months old. the abnormal eye movement was noticed by the family and on the eye examination peripheric chorioretinal hypopigmented lesions were determined when he was months old. İn his labratory examination, viral serology were negative. his parents were not consanguineous. his mother's brother had died at years old. he had history of skin infections and osteomyelitis. his grandmother had been diagnosed as having tuberculosis lymphadenitis one year ago. on his physical examination, his growth was normal, patological findings were horizontal nistagmus, / degree systolic murmur, fistulized lymph tissue on the left submandibuler region. on the laboratory findings; he had anemia and neutropenia. immunglobulin levels and lymphocyte subtypes were normal. his respiratory burst activity was very low. chronic granulamatous disease was thougth in the patient by the clinical and laboratory findings. İt was detected gp phox mutation in the genetical analysis. he was diagnosed as having x-linked cgd. antibiotic prophylaxis (tmpsmx, flucanazol) and interferon gamma were started. he is months old now and he is on the list of match unrelated donor screening. patiens who has history of lympadenitis, skin infections and chorioretinal findings should be evaulated for the x-cgd. leucocyte adhesion deficiencies (lads) are rare autosomal recessive inherited disorders. three different forms of lads have been described so far. in lad-i, the most common leucocyte adhesion deficiency, the function of β integrin cd is lost. . while one of the first signs of the disease consist in delayed separation of the umblical cord, severe infections already start early during infancy. another feature of lad-i includes impaired wound healing. therefore, mortality during infancy is high. a fifty day old boy was referred to the hospital due to diarrhea and leukocytosis. the patient was delivered following an uncomplicated full term pregnancy. the parents were first degree cousins. father's brother and mother's uncle had died during infancy period. his umblical cord had separated on the th day. patient had applied because of diarrhea by starting in the first days of life and leukocytosis was detected ( . / mm ). on the physical examination, his body weight was gr ( th to th percentile) and his height was cm ( th percentile). there was a granuloma on the umbliculus. other systems were normal. in the laboratory examination, leukocyte count was high, immunoglobulins, respiratory burst activity, gaita analysing and culture were normal. in the lymphocyte subtype analysing, cd + b cells ratio was mildly low. cd , cd a, cd b, cd c levels were found to be very low. in the genetic analysing, it was detected two deleterious mutations in the itgb gene. patient had been diagnosed as lad- . he treated by antibiotics and then started prophylactic antibiotherapy. family screened for tissue match. his older sister was found full matched. he was referred to transplantation unit. it was applied bone marrow transplantation when he was months old. presence of delayed umblical cord separation and leucocytosis should be considered in the diagnosis of lad but these patients might have different symptoms such as diarrea. mendelian susceptibility to mycobacterial disease (msmd) is a rare entity. patients with msmd have susceptibility to salmonella and some other intracellular microorganisms in addition to weakly pathogenic mycobacterial species. il- rβ deficiency, most common form of msmd, is caused by mutations in the il rb gene. patients who have symptoms suggestive for msmd or history of sibling death due to bcgosis were evaluated. the expression of the il- rb was detected in patients and family members by monoclonal antibodies on the lymphocyte surface by flow cytometry after the lymphocytes were stimulated in vitro with pha. mutation analyses was done by sanger sequencing. all index cases were presented either with bcg or salmonella infection. two patients, though they were bcg vaccinated had no clinical symptom, six presented with the symptoms of salmonella infections, two developed leukocytoclastic vasculitis, candidiasis was the accompanying feature in seven. recurrent leishmaniasis that necessitated subcutaneous interferon-γ and prophylactic amphotericine b therapy was present in a patient. in all patients the percentage of lymphocytes with il rβ expression was found to be less than % . prophylactic antimycrobial treatment and in severe and resistant infectious episodes if-γ therapy, should be given. many patients have associated mucocutaneous candidiasis. the prognosis is good unless the patient admits at the later stages of bcg infection. the results of our patients showed that the analysis of the surface expression of il rβ on activated lymphocytes is an effective diagnostic method which can also be used in screening of the patients with probable msmd. introduction: mendelian susceptibility to mycobacterial disease (msmd, online mendelian inheritance inman ) is a rare immunodeficiency characterized by predisposition to infections caused by weakly virulent mycobacteria, such as mycobacterium bovis bacille calmette-gue´rin (bcg), environmental nontuberculous mycobacteria (ntm), and salmonella strains in otherwise healthy individuals. il- rb deficiency is the most common form of msmd and is characterized by childhoodonset mycobacteriosis with frequent recurrence. it has been found that patients with il- rb deficiencies are also prone to developing infections with nontyphoidal salmonella species with bacteremia and lymphadenopathy. here we present a girl with recurrent cutaneous leukocytoclastic vasculitis (clv) with salmonella enteritidis due to il- rb deficiency. case report: a four year old girl that had been diagnosed serologically with recurrent salmonella infections, associated with lymphadenopathy and skin eruption was admitted as having henoch-schönlein purpura. she had been vaccinated with bcg and developed left axillary lymphadenitis which spontaneously drained and had recurrent oral monilia plaque. edema and purpuric eruptions were present on the upper and lower extremities and the abdomen.multiple mobile, painful,enlarged submandibular lymph nodes of about x cm in diameter were palpable. skin biopsy showed a dense inflammatory site with eosinophils, neutrophils and fibrin in the upper dermis and dermal vessel wall,compatible with leukocytoclastic vasculitis. serological studies to assess diagnostic markers for vasculitis and infectious agents were all negative. immune work-up were unremarkable other than hypergammaglobulinemia. salmonella enteritidis was identified in blood culture. she responded dramatically to ceftriaxone treatment within a few days and lesions cleared completely. extended immunological and molecular genetic examination of the patient was carried out for il- /ifn-γ pathway defects. on the facs analysis of t cells for cell surface expression of the cytokine receptor chains, she did not express any il- rβ . discussion: in thepresent report, we describe a child with clv with salmonella enteritidis due to il- rb defi-ciency.in a large cohort of patients with il- rb deficiency, ntm, m. tuberculosis, disseminated bcg infection after inoculation with the vaccine, and salmonella infection have been described. sporadic cases with other infectious agents have also been reported. salmonella infections reported in these patients were due to extraintestinal, or septicemic, recurring infections caused by nontyphoidal salmonella species. only two il- rb -deficient patients have been identified with vasculitis due to salmonella strains; both came from turkey, where consanguineous marriages are common. kutukculer and colleagues reported the first case of s. enteritidis-associated clv. sanal and colleagues reported a clv case associated with group d salmonella infection. leukocytoclastic vasculitis is an immune complex mediated disease predominantly involving small vessels of the skin and can be associated with drugs or can be found as a component of other disease, such as infections, connective tissue disorders, and malignancies. infectious agents can cause vasculitis directly or clinically mimic primary vasculitis .multiple infectious agents have been suspected as triggering or contributory factors in the vasculitic process . several factors contribute to the primary vasculitis related to infections: a type or immune-complex reaction, cell-mediated hypersensitivity, abnormal immune regulation, and direct endothelial cell invasion by infectious agents. in our case, extensive evaluation was performed to determine the underlying vasculitis process. clinical and laboratory examinations revealed no association between vasculitis and other infections or an underlying connective tissue disease or medication. she responded well to ceftriaxone treatment, and clinical manifestations gradually resolved within a few days, providing strong evidence that improvement of the vasculitic lesions was due to elimination of the salmonella with antibiotics conclusion: our patient has one of the exceptional forms of il- rb deficiency, with recurrent clv due to salmonella enteritidis. although common presentations for salmonella infection in individuals with il- rb deficiency are lymphadenopathy and bacteremia, it can be present clinically as clv. some infections such as salmonella may be responsible for different types of vasculitis even though they are not common . in this respect, clinicians should be aware of possible infectious causes of vasculitis, and children presenting with unusual recurrent infections caused by non typhoidal salmonella, bcg, or ntm should be investigated for ifn-γ ⁄ il- pathway defects. gülez n.; genel f. dr. behcet uz children hospital allergy-immunology department, izmir, turkiye the complement system is an important part of the innate immune defense and also plays a major role in shaping the adaptive immune response. these functions are required for a good defense against infections, especially bacteria. the c deficiency is a rare disease that is associated with recurrent neisserial infections, especially meningits caused by n. meningitidis. the patient, a seven years old girl was admitted to hospital with high fever and diffuse, purple-coloured skin lesions. her symptoms gave the diagnosise meningococcal meningitis. she had also earlier been diagnosed with the same disease when she was years old. a sister to the patient had died from meningitis at years of age. she has also one older and one younger sister. there is no consanguinity between her parents. the laboratory analyses of the classical pathway measured as complement hemolytic activity (ch ) and c concentration revealed no activity and absence of c , respectively. analysis of serum from her younger sister showed the same results, while her older sister's ch and c levels were found normal. thus, our patient and her younger sister were diagnosed with hereditary c deficiency. the genetic analyses have not been completed yet. we here report the third and fourth cases of c deficiency in turkish patients. interferon-g receptor- (ifngr ) deficiency is caused by mutations in ifn-γ receptor- gene and is characterized mainly by susceptibility to mycobacterial disease. we report a boy with complete recessive ifngr deficiency, afflicted by recurrent mycobacterial diseases with m. bovis, m tuberculosis, m. avium intracellulare and m.fortuitum. genetic analysis showed a homozygous mutation ( inst) in ifngr gene leading to complete ifngr deficiency. in addition, he had atypical mycobacterial skin lesions caused by m.avium intracellulare and he developed scrotal and lower limb lymphedema secondary to compression of large and fixed inguinal lymphadenopathies. to our knowledge, the patient is the first case with interleukin- /interferon − γ pathway defect and severe lymphedema. defects in the il- / ifn − γ pathway must be considered in patients with disseminated or recurrent mycobacterial infections and in patients with severe viral infections, especially in countries where bcg vaccination is part of the national health programme. it must be kept in mind that these patients may develop granuloma-like skin lesions and severe lymphedema. hsct must be applied at the earliest time before developing organ damages. t lymphocyte/nk cells. restricted defective molecules in the circuit and recently discovered cybb responsible for autophagocytic vacuole and proteolysis have been identified in around % of patients with the mendelian susceptibility to the mycobacterial disease (msmd) phenotype. primary defects in oxidase activity in chronic granulomatous disease (cgd) lead to severe, life-threatening infections. the role of phagocytic respiratory burst in host defense against mycobacterium tuberculosis was controversial. previous studied showed that the critical role at reactive oxidants is to serve as intracellular signals for activation of microbicidal enzymes, rather than excretions a microbicidal effect perse.the role of phagocytic respiratory burst in host defense against m. tb is further supported by recent studies discovered immunological defects secondarily affecting phagocyte respiratory burst function and resulting in primary immunodeficiencies with varied phenotypes, including susceptibilities to pyogenic or mycobacterial infections. the patients with severe pid's like scid have broader diverse infections susceptibility and mycobacterial infections as well, however, common variable immunodeficiency (cvid) mostly characterized by a deficiency of immunoglobulins and recurrent sinopulmonary infections. method: we overview the clinical rate of mycobacterial disease in our pid cases and evaluate the complex cases. results: two hundred pid cases were evaluate between - in our clinic, among % of them which diagnosed as msmd nearly all presented with mycobacterial infection. % diagnosed as cgd and interestingly % of them have been experienced mycobacterial disease sometimes in their life, as disseminated bcg or late onset complications of bcg including osteomyelitis or mtb once or more than one episode through their life. also we have presented a cvid patient with disseminated tb and granulomatouse hepatitis, tb arthritis , peritonitis and a patient with lad and nontubercolouse mycobacterial infectiouse abcesses of her skin. conclusion: pid cases like cgd, msmd or cvid which are living in area's with high prevalence of mycobacterial infection could have quiet different presentations and the study of these complex cases has provided essential insights into the functioning of the immune system. despite the conventional view we have confirmed that the generation of rois by phagocytic respiratory burst may play a role in the defense of the host against m. tuberculosis by clinical evidence. goran ristic, srdjan pasic, bojana slavkovic division of clinical immunology, mother and child health care institute of serbia, belgrade chronic granulomatous disease (cgd) is a rare disease caused by mutation in any of the five components of the nicotinamide adenine dinucleotide phosphate (nadph) oxidase in phagocytes, resulting in recurrent, life-threatening bacterial and fungal infections of the affected individuals. our proband male patient presented at age of years with bilateral pneumonia and positive serology for aspergillus sp. the phorbol myristate acetate (pma) stimulated nitroblue tetrazolium (nbt) test showed no reduction ( %) in our patient and partial reduction ( %) in his mother. analysis of the cybb gene showed a deletion of nucleotide g (c. delg) exon causing frame shift mutation and early termination of translation (p.val serfs). this mutation was not previously described. at the moment of diagnosis, his mother was already pregnant, th week of gestation. fetal ultrasound showed that she was carrying a male fetus. the fetal blood sample, obtained by the percutaneous umbilical cord blood sampling, showed male karyotype and the pma stimulated nbt test showed % reduction. there were no complication during pregnancy or delivery and a healthy boy was born. the proband patient underwent allogenic hsct, his sister was the identical sibling donor. in the cases where the family-specific mutations are unknown, partial or complete gene deletions can be recognized by multiplex ligase-dependent probe amplification (mlpa) or array comparative genomic hybridization ( features of the patients with ar-hies dock- deficient and non-dock- deficient group are given in the table . all patients with dock- deficiency presented with cutaneous viral infections or early onset and severe atopic dermatitis. many have also food allergy and/or asthma. neurological complications and malignancy were seen in % and % respectively. sixty seven percent of patients had low t; %, low cd levels; %, high ige. the latter features were shared between patients with or without dock- deficiency, except atopic dermatitis which was mild when present in patients without dock- deficiency and ige levels were only mildly high or normal. we identified stk and coronin a deficiency in two siblings each among the patients who showed overlapping features with ar-hies and do not have dock- deficiency. our results showed that patients with dock deficiency have early onset and more remarkable eczema, food allergy, asthma, more marked eosinophilia, higher ige, low igm levels and development of malignancy. these features may be helpful in differentiation dock patients from patients without dock- deficiency. it seems routine lymphocyte subset study results are not helpful for this differentiation. alişan yıldıran , stephan borte murat elli , tunç fışgın ondokuz mayıs university, school of medicine, samsun-turkiye immunodeficiency center leipzig (idcl) at klinikum st. georg ggmbh, leipzig-germany hsct might be curative for some pids. our immunology and transplantation center was newly established. we retrospectively reviewed all children with pid who diagnosed and received hsct at ondokuz mayıs university or somewhere between june and december . twenty-two patients were identified. four of them were referred to us for hsct from other centers. the median age was months ( month- yr) at hsct. patients' diagnoses were scid (n = ), chs (n = ), leukocyte adhesion deficiency (n= ), mhc class ii deficiency (n= ), chronic granulomatous syndrome (n = ), hlh (n = ), was (n = ) and omenn's syndrome (n = ). seven patients received hla-matched related hsct; twelve haploidentical hsct and two matched unrelated hsct. one scid patient died just after her diagnosis. two patients developed bcgosis secondary to reactivation of pretransplant vaccination. one of them died due to hemophagocytic bone marrow aplasia, and the other has recovered. five patients had graft failure; two of them received no conditioning regimens because of general health status and the other because of cmv infection. at a median follow up of months (range - ), patients are alive, with overall survival of %. we conclude that; our clinic undertakes an important duty in our region for pid patients. also, different pid's could be seen in our region. nerological complication patient (epilepsy, brain infarct) outcome died (one with post -bmt comp., with malignancy all alive department of pediatrics and adolescent medicine department of allergology, rheumatology and clinical immunology, university children's hospital, university medical centre, ljubljana, slovenia. developed.there is no severe life-threatening complications development. we describe here a patient with invasive cryptococcus laurentii infection and the x-linked form of hyper-igm syndrome (x-higm). c.laurentii is an extremely rare human pathogen. this fungus was previously considered saprophytic and non-pathogenic to humans, but it has been isolated as the etiologic agent of skin infection, keratitis, endophthalmitis, lung abscess, peritonitis, meningitis, and fungemia. most affected individuals had a compromised immune system because of leukemia, cancer, diabetes mellitus, aids, or prematurity. repeated isolation of c.laurentii from the oropharynx of an immunocompromised patient has also been documented. invasive c.laurentii infection has not been reported in patients with any form of primary immunodeficiency disorder emphasizing the true rarity of disease due to this fungus. two groups have recently reported that dectin- deficiency due to the mutation of clec a or premature termination of the dectin- signal transduction molecule card may predispose patients to chronic mucocutaneous candidiasis (cmc). we studied the frequency of clec a mutation in healthy individuals, patients with the hyper-ige syndrome (hies), and patients with autoimmune polyendocrine syndrome type (aps- ), all aged between to years. genomic dna was isolated from peripheral blood. monocytes and monocyte-derived dendritic cells (mddcs) were used to study the phenotypic expression of dectin- . clec a gene was sequenced with the big dye terminator cycle sequencing kit. mononuclear cells (mcs) were isolated from heparin-treated venous blood. mdccs were obtained by culturing monocytes isolated by immunomagnetic cell separation assay. receptor expression was assessed by flow cytometry. secretion of il- a by mcs stimulated with killed c. albicans blastoconidia was assessed by elisa. we report here on healthy individuals with homozygous (one year-old man) or heterozygous ( men and women) tyr x mutation in the dectin- gene but no signs of cmc. dectin- levels on monocytes and mddcs were negligible in the homozygous man and the heterozygous individuals displayed intermediate levels of dectin- , between those of the homozygous man and the wild-type controls. markedly lower levels of il- a production were observed in the cells of the man with the homozygous mutation than in the control cells. levels of production of this cytokine were intermediate in heterozygotes. the frequency of tyr x heterozygous individuals was % among healthy donors and % in patients with hies and % in patients with aps . importantly, the year-old hies girl with heterozygous tyr x mutations has never had mucocutaneous candidiasis. we suggest that dectin- receptor-mediated immunity is redundant for host defense against cmc, possibly due to the involvement of multiple lectin receptors in the recognition and uptake of candida. chronic mucocutaneous candidiasis (cmc) is a heterogenous group with recurrent chronic candida infections spesifically involving nails, skin and oropharynx. several immunodeficiencies as dock- deficiency, severe combined immune deficiency, autoimmune polyglandular syndrome type (apeced), il- rβ and il- p deficiencies, card , stat- and stat- can cause cmc. we report here a years old boy, born to consaginous parents with onicomycosis, moniliasis, recurrent pneumonia, recurrent herpetic lesions, autoimmune thyroiditis and trombocytopenia. last admission was due to generalized tonic-clonic convulsion and mycotic aneurism on middle cerebral artery was detected. flow cytometry revealed cd lymphopenia, immunoglobulin values were in normal range. polymorphism on exon for aire gene (t/c heterozygot g g) and heterozygot stat mutation (c. > a; q h) were detected. various stat gof mutations (affecting the coiled-coin domain or the dna-binding domain) have been systematically associated with susceptibility to cmc. autoimmune manifestations associated with stat- mutations have been attributed to increased type interferon. the aneurism formation is not elucidated whether it's due to candida infection or vascular damage directly affected by stat- mutation. purpose: chronic granulomatous disease (cgd) is a rare genetic disease of phagocytic system. affected patients commonly present with bacterial infections associated with pneumonia, abscesses and lymphadenitis. in this study, we investigated the clinical and laboratory findings of our cgd patients. materials and methods: the demographic data (age at diagnosis, initial presenting symptoms, family history, follow-up period), mutation analysis, therapy options, complications, radiological findings and prognosis were evaluated retrospectively. results: among cgd patients, autosomal recessive form was detected in of them. the age at onset was statistically lower in x' linked cgd patients than ar form ( . ± . mo vs . ± . mo; p = . ).respiratory tract infections (sinusitis, otitis, pneumonia) and recurrent abscesses were more commonly seen at onset. microbiological culture revealed a. fumigates from lung biopsy in one patient and s.marcescens from blood specimen in other ones. bcgitis was observed in one patient and five patients received anti-tb therapy. non-infectious complications were granulomatous uveitis, recurrent pericardial effusion, skin granuloma, noduler formation in lung and brain area. conclusion: due to high rate of consanguinity, autosomal recessive inheritance was observed highly in our patient cohort. since, patients with cgd are susceptible to tuberculosis and bcg complications; initiation of tuberculosis prophylaxis is advisable in countries where bcg is still administrated at birth. key words: chronic granulomatous disease, consanguinity, bcg. acquired immunodeficiency research center, isfahan university of medical sciences, isfahan, iranbackground: defects of the immune system in primary immunodeficient diseases (pids) predispose individuals to recurrent infections. complex genetic components for susceptibility to mycobacterial disease have been suggested. natural human immunity to the mycobacteria group, including mycobacterium tuberculosis(mtb), bacille calmette-guérin (bcg) or nontuberculous mycobacteria (ntm) relies on the functional il- / -ifn-γ integrity of macrophages (monocyte/dendritic cell) connecting to key: cord- - nplyyai authors: hawkinson, dana; hinthorn, daniel; danziger-isakov, lara title: novel antiviral agents for respiratory viral infection in immunocompromised adults date: - - journal: curr infect dis rep doi: . /s - - - sha: doc_id: cord_uid: nplyyai respiratory viruses cause significant morbidity and mortality in immunocompromised populations such as stem cell transplant and solid organ transplant patients. few viruses causing respiratory tract infection have an approved therapy, and many of the viruses have no therapeutic options at all. in this article, we describe novel agents under development for treatment options against several respiratory viruses. respiratory viruses cause a wide range of upper and lower respiratory tract infections (urtis and lrtis) in the general population. in the immunocompromised populations, particularly those undergoing chemotherapy for malignancy and solid organ and hematopoeitic stem cell transplant patients, viral respiratory infections can cause high rates of morbidity and mortality. few antiviral therapies exist that provide benefit against these pathogens, but new therapies are under investigation [ , ] . however, there remain limited data about the efficacy of many of these new agents in immunocompromised populations. this review describes the currently published literature regarding novel antiviral agents for use against respiratory viruses in the immunocompromised host. influenza, a single-strand, segmented rna virus, remains one of the more challenging respiratory viral infections for the immunocompromised population. the mainstay of treatment for seasonal influenza, pandemic h n , and influenza b continues to be the neuraminidase inhibitors (nais) oseltamivir and zanamivir; although the m inhibitors amantadine and rimantadine are approved for influenza a, widespread resistance at present precludes use of the latter class of agents [ ] . one limitation to the use of nais is the emergence of resistance, which occurs more frequently in immunocompromised populations [ ] . peramivir, an intravenous (iv) nai, has been studied for the treatment of more severe influenza infections. peramivir use emerged under the emergency investigational new drug (eind) program in to treat serious illness caused by pandemic influenza a h n . in one review of patients from different states, only of ( %) patients treated under eind during the early phase of the pandemic were transplant patients: patients with solid organ transplants and with hematopoietic stem cell transplant (hsct) [ • ]. an additional patients in the eind cohort were on systemic corticosteroids. overall, the authors classified patients who received peramivir as immunocompromised, of which were children. all of the patients in the study had severe illness with respiratory failure. two adult patients died, while recovered. one child recovered, and died with peramivir therapy. in a review of peramivir use under the emergency use authorization in california, the authors identified immunosuppressed patients ( % of the total), defined as "immunosuppressive drugs, cancer, chemotherapy, leukemia, systemic lupus erythematosus, transplant, or other immunosuppressive conditions" [ ] . five of patients lived; baseline characteristics did not predict mortality in this cohort. finally, several case reports describe treatment with peramivir. in one case, a pediatric renal transplant patient with severe oseltamivir-resistant influenza a h n recovered following peramivir therapy; however, this patient also had "aggressive critical care support" and reduction in his immunosuppression [ ] . in hsct patient, peramivir was associated with recovery from severe pandemic influenza a h n , but peramivir use was concurrent with lymphocyte recovery and oseltamivir use and was preceded by triple combination antiviral therapy with oseltamivir, oral ribavirin, and oral amantadine, as well as inhaled ribavirin and intravenous immunoglobulin [ ] . there were no detected oseltamivir h y resistance mutations. additionally, the role of peramivir in the management of patients with documented resistance to oseltamivir has been called into question. in allogeniec hsct patient with oseltamivirresistant virus due to neuraminidase mutation h y, virus replication continued despite peramivir therapy [ ] . overall, the limited data do not allow conclusions about the efficacy of peramivir in immunocompromised populations [ ] . inhaled zanamivir is another nai approved for influenza therapy. there are limited data on inhaled zanamivir in transplant populations, but the tolerability and outcomes suggest that it is a reasonable option, although further studies are needed [ ] . investigational iv zanamivir has been available on a compassionate-use basis in the setting of failure of other antivirals, with contraindication to inhaled zanamivir, or as part of a clinical trial. failure to absorb oseltamivir and the development of oseltamivirresistant (specifically, h y) mutations were reported as indications for usage [ ] [ ] [ ] . in a retrospective study of iv zanamivir in critically ill patients with pandemic influenza a h n who had already received at least days of oseltamivir prior to iv zanamivir, of ( %) were immunocompromised [ ] . seven of the patients developed acute respiratory distress syndrome, and patients ( %) died. several case reports of pediatric immunocompromised hosts, including those with hsct, leukemia, and liver transplantation, appear in the literature, predominantly with oseltamivir-resistant pandemic influenza h n viral isolates. while survival results are mixed for these patients, most report a decrease in the viral load as evaluated by polymerase-chain reaction (pcr) threshold cycle values. most of the remaining case reports relate to pediatric transplant patients, who had mixed survival results [ ] [ ] [ ] [ ] . data on iv zanamivir in the immunocompromised population remain scant, but there may be benefit for its use in oseltamivir-resistant influenza infections. laninamivir octanoate (cs- ) is a long-acting, inhaled nai that was approved in japan in october . given its persistence in the airway, a single dose has generally been used in most studies to date. laninamivir octanoate has shown efficacy in seasonal influenza, oseltamivir-resistant influenza a (h n ), and influenza h n viruses in immunocompetent subjects [ ] . in a postmarketing study of laninamivir octanoate in , patients, only ( . %) were classified as having "diseases accompanied by a reduction in immune function" [ •] . of that group, ( %) were categorized as responding to treatment, with fever resolution and relief from influenza symptoms. at present, there is little data on the efficacy of laninamivir octanoate in immunocompromised persons. the sialidase fusion protein das is another novel agent that has activity against both influenza and parainfluenza viruses [ , ] . the mechanism of action of das is to cleave the terminal sialic acid linkages on the host cells to reduce binding and infectivity of the virus [ , ] . in vitro and in vivo studies suggest that das is active against oseltamivir-resistant strains of influenza. currently, it has been studied in animal models and one phase ii trial [ , •, - ] . in the phase ii study, the patients were infected with seasonal influenza a h n , pandemic influenza a h n , or influenza b [ ] . das was associated with a statistically significant reduction in viral load from days - in these otherwise generally healthy patients. one death occurred in a female patient with previously undiagnosed hiv infection, who expired from respiratory failure, with severe pneumonia and h. influenzae and s. aureus isolated from sputum cultures. overall, further evaluation of das and influenza infection is necessary, particularly in immunocompromised populations (see below for data on the management of parainfluenza virus). respiratory syncytial virus (rsv), an enveloped nonsegmented rna virus in the paramyxovirus family, causes urtis and lrtis in immunocompromised hosts. rsv infection has been linked to significant morbidity and mortality in this population. progression to lrti is associated with increased mortality after hsct, while rsv has been epidemiologically associated with the development of bronchiolitis obliterans syndrome (bos) in adult lung transplant recipients. various therapies have been assessed for an impact on disease progression, morbidity, and mortality, and several emerging therapeutics are under investigation. ribavirin is a guanosine analog active against rna and dna viruses that is available for oral, intravenous, or aerosolized administration. it was discovered more than years ago, and the majority of research and clinical use centers on inhaled and aerosolized forms of the drug; there is emerging data on the use of oral ribavirin for the management of rsv. ribavirin orally is~ %- % absorbed without food. the half-life is days when administered over time, and there is extensive accumulation in all tissues [ ] . it has activity against influenza, measles, west nile virus, rsv, and other viruses. when oral ribavirin became available, pelaez described its use in conjunction with steroids in adult lung transplant recipients, finding oral therapy less costly than aerosolization; this intervention was not directly compared with placebo or other formulations of ribavirin [ ] . additionally, a comparison between outcomes for lung transplant recipients receiving oral (n = ) and inhaled (n = ) ribavirin in conjunction with steroids and ivig was performed at a single center. there were no significant differences reported for survival or bos development/progression at months postinfection in this cohort [ ] . studies assessing the concentration of oral ribavirin in lung fluid in transplant patients have not been conducted. despite limited data on its efficacy in clearing rsv infection in solid organ transplant (sot), ribavirin, especially in the aerosolized form, has additionally been evaluated in conjunction with other therapies, including steroids, ivig, and palivizumab (see below), and remains a standard part of many local treatment protocols. palivizumab, motavizumab, and medi- are all antibodies against rsv fusion protein. currently, only palivizumab is approved for prophylaxis of rsv infection in a select group of premature infants and children < years of age with uncorrected cyanotic congenital heart disease or chronic lung disease. motavizumab is times more potent than palivizumab. in a randomized study, motavizumab exhibited increased skin reaction adverse events but similar clinical outcomes as palivizumab [ ] . medi- has a longer halflife, which may allow for less frequent dosing. neither motavizumab nor medi- has been studied in immunocompromised hosts to date. palivizumab has been assessed in both hsct and lung transplant recipients. the largest study of palivizumab evaluated two groups of hsct recipients [ ••] . in the first group, hsct patients without rsv received palivizumab, noting a serum half-life of . days. an additional hsct patients with rsv (urti= , lrti= ) received palivizumab in addition to aerosolized ribavirin. palivizumab was cleared more rapidly in the rsv-infected group with a half-life of . days. all patients with urti recovered without progression to lrti, while patients with lrti survived past the -day study period. in addition, palivizumab given to hsct patients along with infection prevention measures, including patient cohorting, visitor screening, and hand hygiene, was successful when administered prophylactically during a nosocomial outbreak of rsv [ ] . finally, palivizumab was used as part of a multidrug regimen that included inhaled ribavirin, steroid, and ivig in lung transplant recipients with rsv [ ] . forced expiratory volumes in one second (fev ) after treatment were similar to baseline. however, progression of bos was not impacted by rsv therapy; patients with highergrade bos at infection had more rapid declines in fev over the follow-up period compared to those without bos at infection. aln-rsv is a small interfering rna that can enter the cell to target rsv-specific viral mrna and interferes with viral replication. two randomized placebo-controlled trials of this aerosolized agent have been performed in lung transplant recipients, while no studies have been performed in hsct to date. zamora and colleagues reported on lung transplant recipients with confirmed rsv who received aln-rsv (n = ) or placebo (n = ) for days [ •]. both cumulative total of the daily symptom score (p =. ) and new or progressive bos rates at day (p =. ) were lower in the aln-rvs recipients. simon and colleagues studied an additional lung transplant recipients with rsv who received days of aln-rsv or placebo [ •] . with a longer follow-up period of days, the aln-rsv -treated patients had lower rates of new or progressive bos, as compared with placebo ( . % vs. . %, p =. ). the treatment effect was seen regardless of ribavirin coadministration. novel rsv-specific antivirals rsv (previously a- ) is reported to have novel anti-rsv activity against both subtypes a and b. after a phase i safety and tolerability study in healthy controls, studies were planned in hsct recipients with rsv [clinicaltrials.gov (nct )], but further information and results are not yet available in the literature. two fusion inhibitors that act by blocking an rsv fusion protein are under investigation. tmc has been evaluated in mouse and monkey models showing in vivo anti-rsv activity. mdt- , which is given via a dry powder aerosol, was successfully tested in - in phase i studies for safety and tolerability in healthy and asthmatic adults. neither agent has been evaluated for use in immunocompromised hosts to date. alx- nanobodies also bind to rsv fusion protein f, allowing for the attachment of the virus to the cell, but not fusion with the cell precluding infection [ ] . a novel antibody that arose from a discovery in dromedaries, alx- can be delivered directly to the lung via aerosolization. alx- has been studied in healthy male volunteers in a phase i randomized placebo-controlled protocol to evaluate dosing, safety, tolerability, and pharmacokinetics of the agent but has not been tested in immunocompromised hosts. parainfluenza virus (piv), a single-stranded, enveloped rna virus, has been described as causing severe infections, leading to increased morbidity and mortality in immunocompromised populations. previously published reports described infection in up to % of stem cell transplants and up to % of lung transplant patients [ ] . there have been several published case reports describing treatment of piv type (piv ) lrti in the immunocompromised population with das , whose mechanism has been described [ •, •, •] . two patients with unilateral lung transplants with severe lower respiratory tract piv disease received days of therapy with das . symptoms improved in both patients soon after treatment started. reduction of viral load occurred in both cases, and both patients were discharged after recovery from the illness. side effects included mild elevations in alkaline phosphatase, which resolved after completion of therapy. in hsct, two cases have been described [ •, • ]. an autologous hsct recipient with a history of multiple myeloma and an allogeneic hsct recipient recovered after and days of therapy with das , respectively. they also had variable reductions in their viral load during and after therapy. however, patient experienced relapse of both piv infection and primary disease, at which point further therapy for piv was not pursued [ •] . the human rhinovirus (hrhv), a member of the picornavirus genera, has > serotypes in three different species: a, b, and the more recently characterized c [ ] . the spectrum of disease in immunocompromised populations also ranges from mild to severe [ ] [ ] [ ] . treatment for hrhvs remains supportive, since no active agents are currently commercially available [ ] . a new oral drug vapendavir or bta (biota pharmaceuticals, inc., melbourne, australia), which binds to the viral capsid to inhibit/interfere with receptor binding and subsequently prevent further infection, is under development. a phase randomized, double-blind, placebo-controlled study in asthmatics with hrhv infection showed greater mean reduction in asthma score and higher evening peak expiratory flow in the vapendavir group, versus placebo [ ] . additional investigation in immunocompromised hosts is needed. adenovirus (adv) is a double-stranded, nonenveloped dna virus. there have been more than serotypes and seven species (a-g) identified [ , ] . advs cause both disseminated and localized disease, such as pneumonia in both immunocompetent and immunosuppressed populations [ , [ ] [ ] [ ] . nucleoside analogs ribavirin, ganciclovir, and cidofovir have shown in vitro activity to adv; however, only cidofovir has been recommended as treatment for adv infection [ ] . most recently, a lipid-conjugate, oral form of cidofovir, cmx (chimerix, durham, nc), has been studied in both hsct and sot. first, under an eind program to treat severe adv infections [ ••] , children and adults were enrolled, % ( ) with hsct and % ( ) after sot. while % survived to study completion, deaths ( with disseminated infection and with localized infection) were attributed to adv disease, while others expired due to other causes. the survival of those with localized lung disease was not reported. florescu reported on patients who received cmx as salvage therapy for disseminated adv, of which were available for analysis ( hsct, sot, severe combined immunodeficiency cid) [ ••] . respiratory disease was diagnosed in patients, of which patient expired secondary to adv pneumonia. nine patients experienced a > % decrease in plasma viral load by the end of treatment or follow-up period. in patients who responded to cmx therapy with decreased viral load, survival was improved from median days versus . days (p =. ), as compared with nonresponders. cmx has been well-tolerated and should be explored further as a therapeutic option for adv infections in immunocompromised populations. the enveloped rna virus human coronavirus (hcov) of the family coronaviridae generally causes infections during the winter respiratory viral season. usually mild in immunocompetent hosts, hcov infection in immunosuppressed populations can lead to significant lower respiratory tract disease [ , ] . furthermore, emerging hcovs such as severe acute respiratory syndrome-associated hcov (sars-cov) in - and the more recently identified novel coronavirus (ncov, now mers) in - have prompted further concern for developing therapeutics against this family of viruses. while effective antiviral agents are currently lacking [ ] , discovery and in vitro evaluation of novel hcov therapeutics is underway, with investigation of entry inhibitors, human monoclonal antibodies, proteosome inhibitors, and even a scorpion venom peptide [ ] [ ] [ ] [ ] . for now, hcov treatment remains supportive. the current antiviral armamentarium against respiratory viral infections aside from influenza therapies is limited, resulting in few options to impact the associated morbidity and mortality from these illnesses in immunocompromised hosts. new drugs are currently in development that could have a significant impact on treating these infections. some of the drugs, such as laninamivir octanoate, are available in certain markets, while others can be accessed through the eind mechanism in the united states. still other antivirals are in early clinical and preclinical investigation stage. additional investigation is needed, particularly in immunocompromised hosts, to expand the options for effective therapy for respiratory viral infections. newer influenza antivirals, 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adults: effect on asthma control in a phase clinical trial molecular evolution of human adenoviruses lethal hemorrhagic alveolitis after adenovirus pneumonia in a lung transplant recipient adenoviral infections in adult allogeneic hematopoietic sct recipients: a single center experience severe community-acquired pneumonia caused by adenovirus type in immunocompetent adults in beijing european guidelines for diagnosis and treatment of adenovirus infection in leukemia and stem cell transplantation: summary of ecil- cmx is a potential treatment for adenovirus infection: preliminary antiviral activity results from an open-label, expanded access study of cmx for the treatment of serious or lifethreatening diseases caused by doublestranded dna viruses. paper presented at: european group for blood and marrow transplantation annual meeting . important data regarding effectiveness of cmx as salvage therapy safety and efficacy of cmx as salvage therapy for severe adenovirus infections in immunocompromised patients human coronaviruses: insights into environmental resistance and its influence on the development of new antiseptic strategies novel inhibitors of sars-cov entry acting by three distinct mechanisms human monoclonal antibodies against highly conserved hr and hr domains of the sars-cov spike protein are more broadly neutralizing severe acute respiratory syndrome coronavirus replication is severely impaired by mg due to proteasome-independent inhibition of m-calpain virucidal activity of a scorpion venom peptide variant mucroporin-m against measles, sars-cov and influenza h n viruses acknowledgments the authors would like to thank chimerix and biota pharmaceuticals for providing their data, including posters and oral presentations. conflict of interest dana hawkinson, daniel hinthorn, and lara danziger-isakov declare that they have no conflict of interest human and animal rights and informed consent this article does not contain any studies with human or animal subjects performed by any of the authors. key: cord- - jm c lj authors: abandeh, foad i.; lustberg, mark; devine, steven; elder, pat; andritsos, leslie; martin, stanley i. title: outcomes of hematopoietic stem cell transplant recipients with rhinovirus infection: a matched, case-control study date: - - journal: bone marrow transplantation doi: . /bmt. . sha: doc_id: cord_uid: jm c lj the impact of rhinovirus in hematopoietic stem cell transplant (hsct) recipients is not well defined. a retrospective, matched, case-control study of hsct recipients with rhinovirus was conducted between and . controls were matched for timing relative to transplant, malignancy, and stem cell source. there were cases and controls. cases and controls did not differ with respect to age, gender, ethnicity, donor source, malignancy, conditioning regimen, immunosuppression, antimicrobial prophylaxis, or significant comorbidities. there were no differences in need for icu care, day mortality, hospice discharge, relapse of disease, gvhd or development of disease or infection due to cmv or ebv. other infectious complications after rhinovirus diagnosis were also equal. however, there was an increased number of recurrent hospitalizations from any cause among cases ( . % vs. . %, p= . ). recurrent hospitalizations due to any infection were also more common in cases ( % vs. . %, p= . ). for patients who were diagnosed with rhinovirus pre-transplant (n= ), there was no difference in outcomes compared to matched controls. hsct recipients with rhinovirus have an increased risk of hospital readmission. however, there was no difference in outcomes compared to matched controls. transplantation in patients with active rhinovirus infection appears to be safe. rhinovirus is a ubiquitous rna virus known to cause the 'common cold.' infections occur throughout the year and are generally transmitted by aerosols or direct contact. rhinovirus preferentially infects the upper airways and typically does not cause specific pathological changes. in symptomatic patients, the most common clinical manifestations include rhinorrhea, nasal obstruction, sneezing, sore throat, headache, malaise and fevers. diagnosis of rhinovirus infectious disease is usually made on clinical grounds; however, viral culture, ag detection, pcr and serology may be used for diagnosis. treatment of rhinovirus infectious disease is mainly supportive due to the lack of agents targeting this virus. viral respiratory infections in hematopoietic sct (hsct) recipients are a frequently encountered problem. several studies have shown that rhinovirus can account for - % of cases. , although rhinovirus is known to cause mild upper respiratory tract infectious disease in immunocompetent hosts, it has been linked to fatal respiratory failure in immunosuppressed patients. , these reports are limited, however, and the exact role or pathogenesis of rhinovirus in lower respiratory tract infectious disease has not been fully elucidated. in addition, around % of hsct patients with rhinovirus infection may be completely asymptomatic and prolonged shedding of the virus in respiratory secretions is common. because of these concerns, some medical centers might postpone the hsct procedure and implement infection control measures in infected patients. the aim of the present study is to evaluate the impact and clinical features of rhinovirus on this special patient population. we conducted a retrospective, matched case-control study ( : ) ratio at the ohio state university wexner medical center (osuwmc), a bed tertiary medical center located in central ohio. using microbiological and demographic data from the hsct program database, we identified all cases of rhinovirus isolated through pcr of the upper or lower respiratory tracts in patients who underwent hsct from st october to st october . if a patient had more than one hsct or more than one positive sample for rhinovirus during the study period, then only the first transplant and first rhinovirus diagnosis that fit the inclusion criteria were used. the study was approved by the institutional review board, protocol number h . the cases were defined as patients who underwent hsct in the study period and had rhinovirus infection or disease anywhere from days before transplant to any time after transplant. the controls were defined as patients who underwent hsct in the same study period, and who were never diagnosed with rhinovirus, whether they had pcr testing of the upper or lower respiratory tracts or not. they were then matched according to year of transplant, timing relative to transplant, underlying malignancy, donor source (allogeneic vs autologous) and for allogeneic hsct recipients, and source of stem cells (peripheral blood vs umbilical cord blood). patients under years of age, prisoners and those who did not have documented follow-up after transplant were excluded from the study. all cases were followed for days after rhinovirus diagnosis or days after transplant, whichever was later. in each of the controls, the time of follow-up was determined according to the time of rhinovirus diagnosis in the corresponding case. the following data were collected after reviewing the patients' medical records: age, gender, ethnicity, underlying malignancy, type of hsct, medical comorbidities, conditioning regimen, immunosuppression and anti-infective prophylaxis at the time of rhinovirus diagnosis, laboratory findings at the time of rhinovirus diagnosis ( ± days), other infections after rhinovirus diagnosis, cmv and ebv infection or disease after transplant, in addition to outcomes at the end of the follow-up period including mortality, hospitalizations and gvhd. for the cases, clinical features of rhinovirus infectious disease, timing of diagnosis and radiographic findings were also evaluated. gvhd was diagnosed clinically and confirmed histologically whenever possible by the hsct team and graded according to the consensus criteria. recurrent hospitalizations were defined as hospitalizations for any reason after rhinovirus diagnosis in the follow-up period. asymptomatic patients undergoing hsct were not routinely screened for respiratory viral infections, and as all the patients with rhinovirus in our study had symptoms suggestive of upper or lower respiratory tract disease, we will use the term rhinovirus infectious disease to describe these patients. the term infection by itself should be limited to the detection of such viruses in the absence of signs or symptoms. diagnosis of all rhinovirus cases from upper or lower respiratory samples was done by the luminex xtag respiratory viral panel pcr assay. this assay was universally implemented at osuwmc at the beginning of the study period in october of and is widely used by our clinicians in patients who are undergoing or underwent hsct and have upper or lower respiratory tract infection symptoms to guide approach to therapy. in addition to rhinovirus, this assay can be used to detect influenza, parainfluenza, adenovirus and respiratory syncytial virus. other infections were diagnosed on clinical grounds by the caring physicians by use of routine microbial cultures for bacterial and fungal infection as well as agand pcr-based assays for other viruses and fungal disease. patients' characteristics and outcomes were compared between cases and controls using fisher's exact test or a two sample t-test as appropriate unless otherwise specified. for multi-level categorical variables, comparisons were made using the w -test. in all analyses, a two-tailed p-value of o . was considered to be statistically significant. all data analysis was performed using stata . . there were patients who underwent hsct at the osuwmc during the study period. fifty-five patients were diagnosed with rhinovirus infectious disease. of those, cases were included in the study. eight cases were excluded for the following reasons: prisoner (n ¼ ), lost to follow-up (n ¼ ), could not be matched to controls (n ¼ ) and diagnosis of rhinovirus days before hsct (n ¼ ). there were no significant baseline differences in demographics between cases and controls (table ) . medical comorbidities, conditioning regimen, immunosuppressive and anti-infective agents were also similar. table lists the clinical features of rhinovirus infectious disease among the hsct recipients. the average number of days for diagnosing rhinovirus infectious disease was . days post transplant (median days) and in the few cases before transplant (n ¼ ), . days pre-transplant (median days). all patients were symptomatic at the time of diagnosis. most of the cases were initially diagnosed through a nasopharyngeal swab ( . %) while three patients were diagnosed through a bronchoalveolar lavage (bal) sample (one had upper and lower respiratory tract samples obtained simultaneously). two of the three patients who underwent bal died. there were no significant outcome differences in need for intensive care unit (icu) care, day mortality or hospice discharge, relapse of disease, gvhd or development of disease or infection due to cmv or ebv (table ) . other infectious complications after rhinovirus diagnosis, including bacteremia, c. difficile enterocolitis, febrile neutropenia, pneumonia or other respiratory infections, were also equivalent. however, overall there was a significantly increased number of recurrent hospitalizations from any cause in rhinovirus cases ( . % vs . %, p ¼ . ). recurrent hospitalizations due to an infectious cause were also significantly more common in the rhinovirus cases ( % vs . %, p ¼ . ). of these, other upper and lower respiratory tract infectious diseases were the cause of recurrent hospitalizations in seven cases and six controls (p ¼ . ). for patients who were diagnosed with rhinovirus infectious disease pre-transplant (n ¼ , mean of . days prior), there was no difference in outcomes compared with equivalent pre-transplant matched controls. there was one case of respiratory syncytial virus infectious disease after rhinovirus diagnosis among the cases, while there were seven parainfluenza virus infectious diseases among the controls. one of the control patients was admitted to an outside hospital and died the same day of admission. information about that admission was not available for analysis. the role for rhinovirus in lower respiratory tract disease is not well established. there are several reports linking rhinovirus infection with croup, bronchiolitis and chronic obstructive airway disease exacerbation in addition to bronchial asthma. some reports suggest that rhinovirus might also be associated with lower respiratory tract disease in immunosuppressed hsct recipients and be associated with poor outcome. in one study, patients who underwent conditioning chemotherapy in preparation for hsct were identified as having an acute respiratory illness. ninety-three patients ( %) had community respiratory viruses isolated, of which were identified to be rhinoviruses. seven of these cases were complicated by pneumonia, in which rhinovirus was also isolated from a bal specimen or an endotracheal aspirate in six. two of these patients had an autopsy confirmed rhinovirusassociated interstitial pneumonia or ards. in another study, bal samples from hsct recipients with acute pulmonary infiltrates were tested for rhinovirus and coronavirus by pcr. rhinovirus was detected in six patients. all but one of the patients died; however, a co-pathogen was detected in addition to rhinovirus in all of these patients and their mortality did not differ from patients who tested negative for rhinovirus in bal samples. two other cases of fatal lower respiratory tract infection in sct recipients attributable to rhinovirus have been described. both patients had symptomatic shedding of rhinovirus before their respiratory deterioration. a retrospective study of patients with upper or lower respiratory infections by rhinovirus and enterovirus in adult patients with hematological malignancies has also been described. lower respiratory tract infection was present in patients ( enterovirus infections and rhinovirus infections). three patients with lower respiratory infections died. however, pulmonary co-pathogens were involved in all cases as well. this is the first study comparing the outcomes of rhinovirus infectious disease on adult hsct recipients with non-infected controls. we did not see any difference in terms of mortality, icu care, or other associated infections. the significant difference was mostly observed in the number of recurrent hospitalizations with rhinovirus cases admitted to the hospital more frequently after the rhinovirus diagnosis. this was true whether the readmission was for all cause hospitalization or due to a specific infectious event. whether the rhinovirus itself increases susceptibility to infections, especially of the respiratory tract and thus admission rates, is difficult to determine. the other possible explanation of this finding is that hsct patients with rhinovirus have more symptoms than non-infected controls and are therefore admitted more frequently. there are several limitations to our study. it was retrospective and conducted at a single facility with a relatively small number of subjects. in addition, the pcr assay used may not always reliably distinguish between other picornaviruses (such as enteroviruses) and rhinoviruses. this means that it is at least possible that some of the cases included in the study as rhinovirus infections were actually secondary to enteroviruses or other closely related viruses. given the ubiquity of rhinoviruses and their being the most common respiratory virus worldwide, it is safe to assume that the vast majority, if not all of our cases, were true rhinovirus infections. additional studies are needed to determine if transplanting patients with rhinovirus needs to be postponed. current recommendations suggest that caregivers should consider deferring conditioning or chemotherapy for hsct or leukemic patients with respiratory virus infections. although deferring such therapies may be better established in other respiratory virus infectious diseases, it is unclear if such an approach would be necessary in the case of rhinovirus infection. distinguishing between actual infectious disease and infection may have a role in this determination. in this study, patients were diagnosed with rhinovirus pre-transplant. when comparing these patients to matched controls, there was no significant difference in mortality, icu care or recurrent hospitalizations (p ¼ . , . and . , respectively). transplanting these patients appeared to be safe, but the small number of patients who were diagnosed pretransplant has made these comparisons small. in conclusion, this study demonstrates that hsct patients with rhinovirus infectious disease do not have worse outcomes compared with matched controls. larger prospective studies are needed to study the impact of rhinovirus infection and disease on this unique population, including specific determination of rhinovirus subtypes associated with severe disease, the incidence and effects of prolonged shedding, or association with other lower respiratory tract infections. until then, in line with current recommendations regarding the prevention of these infections, continued infection control measures in hsct patients with suspected respiratory viral infections should continue both in and out of the hospital setting. principles and practice of infectious diseases, th edn ecil- ): guidelines for diagnosis and treatment of human respiratory syncytial virus, parainfluenza virus, metapneumovirus, rhinovirus, and coronavirus respiratory virus infections after marrow transplant: the fred hutchinson cancer research center experience respiratory viral infections after bone marrow/peripheral stem-cell transplantation: the christie hospital experience rhinovirus infections in myelosuppressed adult blood and marrow transplant recipients rhinovirus as a cause of fatal lower respiratory tract infection in adult stem cell transplantation patients: a report of two cases consensus conference on acute gvhd grading rhinovirus and the lower respiratory tract rhinovirus infections in hematopoietic stem cell transplant recipients with pneumonia upper and lower respiratory tract infections by human enterovirus and rhinovirus in adult patients with hematological malignancies dr lustberg's effort on this study was in part supported through a training grant, t ca . the authors declare no conflict of interest. key: cord- -e ylkonb authors: mo, xiao-dong; zhang, xiao-hui; xu, lan-ping; wang, yu; yan, chen-hua; chen, huan; chen, yu-hong; han, wei; wang, feng-rong; wang, jing-zhi; liu, kai-yan; huang, xiao-jun title: treatment of late-onset hemorrhagic cystitis after allogeneic hematopoietic stem cell transplantation: the role of corticosteroids date: - - journal: ann hematol doi: . /s - - - sha: doc_id: cord_uid: e ylkonb we aimed to evaluate the treatments, particularly the role of corticosteroids, in patients with late-onset hemorrhagic cystitis (lohc) after allogeneic hematopoietic stem cell transplantation (allo-hsct). one hundred and sixty-three consecutive patients who underwent non-t-cell-depleted allo-hsct and met the criterion of lohc after allo-hsct were enrolled in this study. the median time from allo-hsct to the occurrence of lohc was (range, – ) days. pathogens identified in blood and/or urine samples from patients were mostly viruses. all of the patients with lohc received intravenous fluid hydration, alkalization, and forced diuresis, of which patients achieved complete remission (cr) after these treatments. the remaining patients received anti-infection therapies and achieved cr after the therapies. corticosteroids were additionally applied to out of patients who did not achieve cr after anti-infection therapies, and . % (n = ) of them showed a grade to lohc at the beginning of corticosteroid therapy. thirty-five patients showed an immediate response (cr or downgraded at least one grade) within week after the beginning of the corticosteroid therapy. sixty-four patients ( . %) achieved cr after corticosteroid therapy, and the median period from the beginning of corticosteroid therapy to cr was days. thus, we observed that viruses were the most common pathogens in lohc after allo-hsct and that anti-infection therapies were critical. for patients not showing a satisfactory response to anti-infection therapies, additional corticosteroid therapy may help to achieve cr. allogeneic hematopoietic stem cell transplantation (allo-hsct) is an effective treatment for hematological malignancies. although transplantation techniques have progressed significantly, late-onset hemorrhagic cystitis (lohc) is still one of the major complications, with an incidence rate varying from . to % [ , ] . it can cause mortality and lead to a low survival rate [ , ] . there are several therapeutic methods for lohc, including ensuring appropriate hydration, hematological homeostasis (maintaining high platelet counts, appropriate red cell counts, and levels of clotting factors), pain relief, catheterization for cystoscopic clot extraction, continuous bladder irrigation with normal saline for prevention of clots and bladder tamponade, anti-infection (particularly antiviral), hyperbaric oxygen, estrogen, clotting factors, and keratinocyte growth factor therapies [ ] . although lohc tends to be selflimiting after or weeks, some patients, particularly the patients who received alternative donor allo-hsct, can show refractory lohc which can persist for several months [ ] . refractory lohc may lead to poor quality of life, even mortality, after allo-hsct. given the assumption that alloimmune injury might also be involved in the pathogenesis of lohc [ ] , huang et al. [ ] used corticosteroids to treat patients with refractory lohc, and all of them achieved complete remission (cr). however, the small sample size of the patients (n = ) in this pilot study limited the ability to further identify the efficacy of corticosteroids in the treatment of lohc. thus, in the present retrospective study, we aimed to further evaluate the treatments, particularly the use of corticosteroids, in patients with lohc after allo-hsct. patients a total of consecutive patients underwent non-t-celldepleted allo-hsct at the peking university institute of hematology (puih) from january , , to december , : , , and had undergone human leukocyte antigen (hla)-unrelated donor (urd) hsct, hla-identical sibling donor (isd) hsct, and hla-haploidentical related donor (haplo-rd) hsct, respectively. one hundred and sixty-three patients who met the criterion of lohc after allo-hsct were enrolled in this study : , , and had undergone urd hsct, isd hsct, and haplo-rd hsct, respectively ( table ). the final follow-up visits for the endpoint analysis were conducted on july , . informed consent was obtained from all patients or their guardians, and the study was conducted in accordance with the declaration of helsinki. the study protocol was approved by the ethics committee of peking university people's hospital. preconditioning consisted of cytarabine, busulfan ( . mg kg − day − , administered intravenously on days − to − ; day being the first day of donor cell infusion), cyclophosphamide ( . g m − day − , days − to − ), and simustine ( mg m − , day − ). cytarabine was administered at g m − day − (days − to − ) to the haplo-rd group, at g m − day − (days − to − ) to the urd group, and at g m − day − (day − ) to the isd group. rabbit antithymocyte globulin (thymoglobulin, . mg kg − day − , days − to − ; sanofi, france) was administered to the haplo-rd and urd groups [ , ] . granulocyte colony-stimulating factor (g-csf)-mobilized, fresh, and unmanipulated bone marrow (bm) and peripheral blood harvests were infused into the recipients on the day of collection. in addition, the patients received cyclosporine a (csa), mycophenolate mofetil (mmf), and short-term methotrexate (mtx) as graft-versushost disease (gvhd) prophylaxis [ ] . donor selection, hla typing, and stem cell harvesting were performed as described previously [ , ] . minimal residual disease (mrd)-directed immunotherapy (e.g., donor lymphocyte infusion and interferon-α) was given before hematological relapse, as a preemptive intervention therapy, months post-hsct [ , ] . comorbidities in hsct recipients were assessed by the hematopoietic cell transplantation-specific comorbidity index (hct-ci) [ ] . all of the patients were hospitalized in rooms with highefficiency particulate-arresting (hepa) air filters for - weeks, from day − until the time at which neutrophil recovery was achieved. all of the patients received oral antibiotics (e.g., fluoroquinolone) for gastrointestinal decontamination before and during the period of neutropenia. the patients also received trimethoprim-sulfamethoxazole to prevent pneumocystis carinii infection from days − to + , fluconazole or itraconazole capsules for invasive fungal infection from days − to + , and acyclovir ( - mg) for herpes simplex virus and varicella-zoster virus, administered orally, twice daily, from day + until the time of csa discontinuation. ganciclovir ( mg kg − ) was administered intravenously, twice daily, from days − to − , for prophylaxis against cytomegalovirus (cmv) infection. the infection surveillance and treatment protocols used at our institute have been described in detail elsewhere [ ] . [ ] . for positive cmv and ebv samples, the pathogen load was determined by quantitative pcr. additionally, urine samples were subjected to gram, fungal, and acid-fast bacilli staining, and to bacterial and fungal cultures. these staining and cultures were also performed on blood samples from patients who presented with fever. blood and urine samples were tested repeatedly weekly. diagnosis of hemorrhagic cystitis (hc) required the presence of sustained hematuria along with dysuria and/or lower abdominal pain. diagnosis of lohc required that the symptoms of hc occurred beyond days post-transplantation. the severity of hc was graded according to the published criteria: grade , microscopic hematuria on more than two consecutive days; grade , macroscopic hematuria; grade , macroscopic hematuria with clots; grade , macroscopic hematuria with clots and impaired renal function secondary to urinary tract obstruction [ ] . severe lohc was defined as grade to lohc, and non-severe lohc was defined as grade to lohc. as prophylaxis for hc, all of the patients were given l m − day − of intravenous fluid, from h before to h after the administration of cyclophosphamide. sodium mercaptoethanesulfonate was given intravenously, at a dose of mg kg − , prior to cyclophosphamide administration and every h thereafter, over h, until the last dose of cyclophosphamide [ ] . with the diagnosis of lohc, along with the supportive therapies (ensuring appropriate hydration, alkalization, forced diuresis, hematological homeostasis, and pain relief), the patients without gvhd were subjected to tapering of immunosuppressant. the anti-infection therapies included antiviral and antimicrobial therapies. the patients showing cmvnegative blood and urine samples could receive empirical antiviral therapies with mg kg − day − of intravenous ganciclovir or - mg kg − day − of intravenous foscarnet sodium [ , ] . however, the patients showing cmv-positive blood and/or urine samples should receive anti-cmv therapies, i.e., mg kg − day − of intravenous ganciclovir or mg kg − day − of intravenous foscarnet sodium. the patients with refractory cmv infection, that is, cmv dnaemia lasting for > weeks despite the administration of a full dose of antiviral drug therapy, received the combination of foscarnet and ganciclovir. antiviral therapies were given for - days or until week after the virus tests became negative. the patients could also receive empirical antimicrobial therapies, such as carbapenem, cephalosporins containing the β lactamase inhibitor, fourth-or third-generation cephalosporin, or quinolone, for at least days. for the patients who did not achieve cr after anti-infection therapy, an additional corticosteroid therapy was applied. the dose was prednisone mg kg − day − , if the patient's weight was < kg, or prednisone mg day − for patients whose weight was > kg. prednisone could be changed by equivalent doses of dexamethasone or methylprednisolone. the patients' response was evaluated - days later. for the patients responding to corticosteroids, the dose of corticosteroids was reduced, weekly, to two thirds of the dose of the previous week [ ] . however, for the patients having other transplant-related complications that also needed corticosteroid therapy during lohc (e.g., gvhd), the initial time, dosage, and duration of corticosteroid therapy were applied according to these transplanted complications instead of lohc. how to manage patients with concurrent active gvhd, cmv infection, and lohc was critical in the treatment of refractory lohc in the present study. for the patients having concurrent cmv infection and refractory lohc, we suggested that systemic corticosteroid therapy could only be considered when the virus was cleared but the loch was not cr. however, for the patients having concurrent grade ii to iv acute gvhd and refractory lohc, systemic corticosteroid therapy should be added immediately with the use of empirical antiviral therapies or anti-cmv therapy (fig. ) . the criteria of therapeutic efficiency for lohc were as follows: ( ) cr, hc symptoms were relieved and microscopic hematuria disappeared, and there was no recurrence in the week after achieving cr; ( ) partial remission (pr), hc symptoms had significantly improved (such as > % reduction in the frequency of urination or relief from dysuria and burning) and/or gross hematuria was downgraded at least one grade, but cr was not achieved; and ( ) non-remission (nr), hc symptoms and hematuria did not improve or showed significant deterioration (e.g., gross hematuria upgraded at least one grade). the patients were classified as bhigh risk^if they ( ) had acute leukemia in its third complete remission (cr ) or greater; ( ) had acute leukemia in pr, in nr, or in a state of relapse before hsct; and ( ) had chronic myeloid leukemia after the first chronic phase. all the other patients were stratified into standard-risk categories. neutrophil engraftment was defined as the first day when the absolute neutrophil count was ≥ . × l − for three consecutive days, and platelet engraftment was defined as the first day when the platelet count was ≥ × l − for seven consecutive days without transfusion. gvhd was diagnosed in accordance with the accepted international criteria [ , ] . relapse was defined as morphologic evidence of disease in samples from the peripheral blood, bone marrow, or extramedullary sites, or by the recurrence and sustained presence of pre-transplantation chromosomal abnormalities. patients exhibiting mrd were not classified as showing relapse. non-relapse mortality (nrm) was defined as death by any cause in the first days post-hsct or death without evidence of disease recurrence beyond day . overall survival (os) events were defined as death from any cause. disease-free survival (dfs) was defined as the survival period with continuous cr. data were censored at the time of death, relapse, or the last available follow-up. continuous variables were compared using the mann-whitney u test; categorical variables were compared using the χ test and fisher's exact test. the kaplan-meier method was used to estimate the probability of survival. competing risk analyses were used to calculate the cumulative incidence of lohc, using gray's test to evaluate differences between the groups [ ] . nrm and relapse were the competing events. the level of significance was set at p < . . all reported p values were based on two-sided tests. data analyses were primarily conducted with spss software (spss inc., chicago, il, usa), while the r software package (version . . ; http://www.r-project.org) was used for competing risk analysis. table summarizes the characteristics of the patients with lohc in this study. fifteen were children ( . %) and the other were adults ( . %). all the patients achieved neutrophil engraftment within days after hsct, with a median time to neutrophil engraftment of (range, - ) days. during the follow-up period, patients exhibited platelet engraftment, with a median time to platelet engraftment of (range, - ) days. six patients showed relapse and died of nrm after hsct. the -year probability of dfs and os after hsct was . and . %, respectively. one hundred fig. the management of concurrent active gvhd, cmv infection, and lohc. for the patients with refractory lohc only, the dose of systemic corticosteroid therapy was prednisone mg kg − day − , for patients whose weight was < kg, or prednisone mg day − for patients whose weight was > kg. for the patients with grade ii to iv acute gvhd, the dose of systemic corticosteroid therapy was methylprednisolone to mg kg − day − and four patients ( . %) showed acute gvhd, and patients ( . %) showed grade iii to iv acute gvhd. the median time from transplantation to the occurrence of lohc was ( - ) days. all the patients exhibited frequent and urgent urination and odynuria. thirty-four patients exhibited a fever, and the median temperature at lohc diagnosis was . ( . - . )°c. the median count of white blood cells, hemoglobin, and platelets on lohc diagnosis was . (range, . - . ) × cells l − , (range, - ) g l − , and (range, - ) × cells l − , respectively. sixteen ( . %), ( . %), and ( . %) patients showed grade , , and lohc, respectively, at diagnosis. ninety-six patients showed progression from the initial grade to a more advanced grade, and ( . %), ( . %), ( . %), and ( . %) patients showed grade , , , and lohc, respectively, at peak. the cumulative incidence of lohc at days after hsct was . % for the total population and . , . , and . % for isd hsct, haplo-hsct, and urd hsct recipients (p < . ; fig. a ). the cumulative incidence of grade to lohc at days after hsct was . % for the total population and . , . , and . % for isd hsct, haplo-hsct, and urd hsct recipients, respectively (p = . ; fig. b ). pathogens were identified in blood and/or urine samples from patients: had positive blood samples, had positive urine samples, and had simultaneous positive blood and urine samples. forty-three and patients had infections caused by single and multiple viruses (≥ types of viruses), respectively. in blood samples, cmv was the most common virus ( . %), followed by bk virus ( . %), ebv ( . %), hsv ( . %), jc virus ( . %), hhv- ( . %), adv ( . %), and piv ( . %). in urine samples, the most common virus was bk virus ( . %), followed by jc virus ( . %), cmv ( . %), and adv ( . %). the median plasma and urine viral loads of cmv were . × /copies ( . × /copies- . × /copies) and . × /copies ( . × /copies- . × /copies), respectively, and it was identified simultaneously in plasma and urine samples from patients. the median plasma loads of ebv were . × /copies ( . × /copies- . × /copies). bk virus was identified simultaneously in plasma and urine samples from patients, and jc virus was identified simultaneously in plasma and urine samples from patients. the most common bacterium identified in urine samples was escherichia coli (n = ; . %), followed by enterococcus faecalis (n = ; . %), proteus mirabilis (n = ; . %), and acinetobacter junii (n = ; . %). all patients with lohc received supportive therapies, and achieved cr after these treatments (grade : n = ; grade : n = ; no pathogens were detected in blood and urine samples). the remaining patients received anti-infection therapies. all of them received antiviral therapy ( . % of patients received anti-cmv therapies, and the other . % of patients received empirical antiviral therapies). a total of patients received antibacterial therapies. seventy-one out of patients ( . %) achieved cr after anti-infection therapies, and the period from the beginning of anti-infection therapies to cr was (range, - ) days. the remaining patients ( . %) did not achieve cr (nr: n = ; pr: n = ). the rate of cr after anti-infection therapies was fig. cumulative incidence of late-onset hemorrhagic cystitis. the cumulative incidence of total (a) and grade to (b) late-onset hemorrhagic cystitis according to different donors significantly lower in the severe lohc group compared to that of the non-severe loch group ( . vs. . %, p < . ). for the patients that did not achieve cr after anti-infection therapies, of them were not subjected to corticosteroid therapy (severe lohc: n = ; non-severe lohc: n = ), and none of them achieved cr until the last available follow-up. the other patients received the additional corticosteroid therapy, and . % (n = ) of them showed a severe lohc at the beginning of corticosteroid therapy. nine patients showed virus positivity when corticosteroids were administered (cmv: n = ; ebv: n = ; bk virus: n = ). thirty-three of the patients received corticosteroid therapy because of other transplant-related complications (group a), including gvhd (n = ), engraftment syndrome (n = ), poor graft function (n = ), and diffuse alveolar hemorrhage (n = ). the other patients that received corticosteroid therapy did not have additional transplant-related complications (group b). the median period from the beginning of anti-infection therapies to the beginning of corticosteroid therapy was (range, - ) days, which was significantly shorter in group a than that in group b ( vs. days, p < . ). sixty-nine out of patients ( . %) responded to corticosteroid therapy (cr: n = ; downgraded at least one grade, although not achieving cr: n = ). thirty-five out of ( . %) patients showed an immediate response within week after the beginning of the therapy; particularly, achieved cr within week. for the patients that downgraded at least one grade after therapy, finally achieved cr. thus, a total of patients ( . %) achieved cr after corticosteroid therapy. the median period from the beginning of corticosteroid therapy to cr was (range, - ) days, which was comparable between groups a and b ( . vs. days, p = . ). in addition, among the other five patients who downgraded at least one grade after corticosteroid therapy, all the symptoms of lohc were relieved persistently; nevertheless, continuous microscopic hematuria was observed until the last available follow-up. among the patients that did not achieve cr after antiinfection therapies, the cumulative incidence of cr for lohc was significantly higher in the corticosteroid-treated group than that in the corticosteroid non-treated group ( . vs. . %, p = . ; fig. ). for the patients that did not show any response to additional corticosteroid therapy, relapsed and did not receive further therapies for lohc, and died of nrm (infection: n = ; gvhd: n = ; thrombotic microangiopathy: n = ). for the other patients, were subjected to electrocoagulation hemostasis with cystoscope, and all of them achieved cr after cystoscopy. for the remaining patients, corticosteroid therapy was discontinued, and the patients were subjected to intravenous fluid hydration, alkalization, forced diuresis, continuous bladder irrigation with normal saline, and antiinfection therapies. cr was achieved at the last available follow-up. the treatments of lohc are summarized in fig. . although transplantation techniques have progressed significantly, lohc after allo-hsct continues to seriously influence patients' quality of life. in the present study, we observed that viruses were the most common pathogens in lohc after allo-hsct. for patients not showing a satisfactory response to supportive and anti-infection therapies, additional corticosteroid therapy may help to achieve cr. the present study was the largest study identifying the efficacy of corticosteroid therapy in the treatment of lohc after allo-hsct. in the present study, cmv was the most common virus in patients with lohc. cmv-associated lohc has been described, as case reports, in some centers [ , ] . xu et al. [ ] showed that cmv viremia is a risk factor for lohc (rr = . ; % ci, . - . ; p = . ). han et al. [ ] showed that the cumulative incidence of lohc at day in patients with and without cmv viremia (prior to or at the onset of lohc) was . and . % (p = . ), respectively, and that cmv viremia (hr = . ; % ci, . - . ; p = . ) was an independent risk factor for the development of lohc. thus, decreasing the occurrence of cmv viremia is important to decrease the risk of lohc after allo-hsct. however, we observed that cmv was identified in urine in only patients, and cmv loads were lower in urine than in plasma. thus, the role of cmv in lohc pathogenesis should be further elucidated. bk virus was another important virus for lohc, which was the second most common virus in blood samples and the most common virus in urine samples in the present study. several studies showed that lohc is strongly associated with the presence of bk virus, although the role of bk virus in lohc pathogenesis has not yet been fully elucidated [ , ] . however, we did not detect copies of bk virus, and we could not further identify the association between the burden of bk virus and lohc. we observed that pathogens were identified in most patients ( / ) and viruses were the most common pathogens. in addition, many patients ( / ) showed infection with multiple viruses. previous studies also reported that virus infection was the most important pathogenesis for lohc [ , , , , ] . thus, we hypothesized that anti-infection therapies should be the basis for the treatment of lohc after allo-hsct. in the present study, more than % of the patients with grade to lohc achieved cr after anti-infection therapies. although infection may be the most important cause of lohc in allo-hsct recipients [ ] , some patients showed unsatisfactory response to anti-infection treatments, particularly for those with severe lohc. in the present study, only . % of patients with grade to lohc could achieve cr after anti-infection therapies alone. thus, other pathogeneses, such as immune injury, may also contribute to the occurrence of lohc after allo-hsct. ost et al. [ ] suggested that post-engraftment lohc represent uroepithelial gvhd. several studies also reported the association between acute gvhd and lohc after allo-hsct [ , ] . in the present study, we observed that more than % of the patients showed acute gvhd. thus, the association of lohc with gvhd regarding timing, incidence, and severity suggests that its pathogenesis may involve a local inflammatory environment, cellular immune responses, effector mechanisms, and hla as well as non-hla genetics [ ] . in addition, several infectious agents can trigger autoimmunity via different mechanisms [ ] , and several studies have observed that infections, particularly viral infections, can trigger a graft-versus-host reaction [ ] [ ] [ ] . in the model of leung et al. [ ] for lohc, infected uroepithelial cells are attacked by donor lymphoid cells, leading to tissue destruction. thus, the role of additional corticosteroid therapy after anti-infection and supportive therapies is worth considering in the treatment of patients with lohc, particularly those showing unsatisfactory responses to anti-infection therapies. huang et al. [ ] reported that patients with refractory lohc received a low dose of corticosteroids, and all of them achieved cr. in the present study, although . % of the patients who showed an unsatisfactory response to supportive and anti-infection therapies and received additional corticosteroid therapies had severe lohc, more than half of them showed an immediate response to corticosteroid and . % finally achieved cr. thus, the additional corticosteroid therapies may be able to significantly shorten and change the course of lohc and it may be a potential therapy for lohc patients showing unsatisfactory response to antiinfection therapies. the present study had several limitations. first, this was a retrospective study, which might have influenced the accuracy fig. treatments for patients with late-onset hemorrhagic cystitis. cr, complete remission; nr, non-remission; nrm, nonrelapse mortality; pr, partial remission of our findings. second, we might have underestimated the occurrence of virus-associated lohc in this study, because the number of viruses that could be tested was relatively small. third, the several, varied treatments for lohc introduced heterogeneity into our study results and reduced the reliability of our conclusion about the advantage of corticosteroid therapy for clinical outcomes in patients with lohc. future prospective and multicenter studies will provide more information about pathogens and treatments of lohc after allo-hsct. in summary, we observed that viruses were the most common pathogens in lohc allo-hsct and that anti-infection therapies were critical for these patients. for patients showing unsatisfactory response to anti-infection therapies, additional corticosteroid therapy may help to achieve cr, particularly for those with severe lohc. hemorrhagic cystitis following high-dose chemotherapy and bone marrow transplantation in children with malignancies: incidence, clinical course, and outcome haemorrhagic cystitis in paediatric bone 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autoimmune disease viral infection as a trigger in flares of acute graft-versus-host disease chronic graft-versus-host disease following varicella-zoster virus infection in allogeneic stem cell transplant recipients the complex relationship between human herpesvirus and acute graft-versus-host disease acknowledgments the authors thank editage for their assistance in editing this manuscript and dr. yu-jia chi, wei guo, and li-jia ma for their assistance in collecting the data. informed consent was obtained from all patients or their guardians, and the study was conducted in accordance with the declaration of helsinki. the study protocol was approved by the ethics committee of peking university people's hospital. the authors declare that they have no conflict of interest. key: cord- -bej xbf authors: lum, lawrence g.; bollard, catherine m. title: specific adoptive t-cell therapy for viral and fungal infections date: - - journal: management of infections in the immunocompromised host doi: . / - - - - _ sha: doc_id: cord_uid: bej xbf despite advances in anti-infective agents, viral and fungal infections after hematopoietic stem cell transplantation (hsct) continue to cause life-threatening complications that limit the success of hsct. early adoptive t-cell immunotherapy studies showed that administration of allogeneic virus-specific cytotoxic t lymphocytes (vctl) can prevent and control viral infections and reconstitute antiviral immunity to cytomegalovirus (cmv) and epstein-barr virus (ebv). advances in immunobiology, in vitro culture technology, and current good manufacturing practice (cgmp) have provided opportunities for advancing adoptive cell therapy for viral infections: ( ) t cells have been expanded targeting multiple pathogens; ( ) vctl production no longer requires viral infection or viral vector transduction of antigen-presenting cells (apcs); ( ) the source of lymphocytes is no longer restricted to donors who are immune to the pathogens; ( ) naive t cells have been redirected with chimeric antigen receptor t cells (carts) or armed with bispecific antibody-armed t cells (bats) to mediate vctl activity; ( ) these technologies could be combined to targeted multiple viral or fungal pathogens; and ( ) pathogen-specific t-cell products manufactured from third parties and banked for “off-the-shelf” use post-hsct may soon become a reality. infections remain the leading cause of mortality and morbidity during the first months after hematopoietic stem cell transplantation (hsct) [ ] [ ] [ ] [ ] . despite advances in prophylactic viral and fungal therapy to minimize the viral and fungal burden early after hsct, breakthrough viral and fungal infections remain life-threatening, and for some viral and fungal infections, there are no effective therapies [ ] [ ] [ ] [ ] [ ] . vaccine strategies to induce immunity to cmv began in the s but have been limited in their success [ ] [ ] [ ] . the conditioning regimens for hsct that vary from non-myeloablative to myeloablative create an immunodeficiency that leaves the allogeneic hsct recipient susceptible to viral and fungal infections while immune reconstitution occurs during the first - months after hsct. immune reconstitution is further abrogated by intensive immunosuppression used to prevent and/or control gvhd. it is clearly established that the kinetics and rate of t-cell reconstitution are critical to controlling viral infections. factors that speed t-cell recovery will decrease the risk of viral infection during the first months after hsct [ , , ] . early studies showed that donor lymphocyte infusions (dli) given before t-cell reconstitution from the stem cell donor were effective for treating viral infections in hsct recipients but were associated with a high risk of gvhd [ ] . since the early s, investigators began to develop virus-specific cytotoxic t lymphocyte (vctl) for adoptive immunotherapy against specific targets early during immune reconstitution after hsct [ , ] . advances in vctl therapy have benefited from ( ) advances in understanding of immune responses to conserved t-cell epitopes for various pathogens [ ] [ ] [ ] , ( ) technological advances in ex vivo expansion of t cells and advances in the preparation of antigen-presenting cells [ ] [ ] [ ] , and ( ) assays that evaluate vctl activity and the mhc restriction of vctl [ , ] . in this chapter, we review the following areas of how: ( ) t cells have been expanded to target multiple pathogens; ( ) vctl production no longer requires viral infection or viral vector transduction of antigen-presenting cells (apcs); ( ) the source of lymphocytes is no longer restricted to donors who are immune to the pathogens; ( ) naive t cells have been redirected with chimeric antigen receptor t cells (carts) to target pathogen-infected cells; ( ) bispecific antibody (biab)-armed t cells (bats) can mediate vctl activity; and ( ) pathogen-specific t-cell products can be manufactured by third parties and banked for "off-the-shelf" use post-hsct. we summarized the methodological approaches, clinical trials using vctl, promising preclinical studies, and early clinical trials of anti-pathogen ctls that have promise. these advances provide the rationale and impetus for future vctl adoptive immunotherapy. production of vctl as a guiding principle to decrease the risk of gvhd in allogeneic hsct recipients, strategies excluded alloreactive t cells by selecting virus-specific t cells. four major approaches were used: ( ) stimulation with viral antigen(s) during ex vivo culture of donor t cells from peripheral blood mononuclear cells (pbmc), ( ) direct selection of donor cells, ( ) genetic modification of t cells to confer specific recognition of pathogen or pathogen-infected cells, or ( ) arming of ex vivo expanded t cells with bispecific antibody to target the viral antigen ( fig. . ). numerous ex vivo culture approaches have been used to produce cytomegalovirus (cmv)-specific ctl or epstein-barr virus (ebv)-specific ctl [ , , [ ] [ ] [ ] [ ] [ ] [ ] . cmv viral-or peptide-specific stimulation in vitro expands single or multiple pathogenspecific vctl. the advantages of culture over cell selection are the generation and expansion of polyclonal vctl to clinically useful quantities of vctl from small amounts of blood [ ] . however, the major disadvantages of this strategy is the daunting task of culturing and processing after stimulation to expand the vctl (up to more than month) and the hlahistocompatibility requirement of finding a closely matched donor. during these longer-term cultures, the vctl may lose their capacity to self-renew and to persist in vivo, particularly after prolonged ex vivo culture [ ] . it should be noted that clinical trials infusing ex vivo expanded vctl post-hsct showed prolonged persistence [ ] and that ex vivo expansion using pathogen-specific stimuli decreased alloreactivity [ ] . this may be due to selection of virus-specific clones and deselection of alloreactive clones. one study showed that residual alloreactivity seen in vctl is clinically insignificant [ ] . the initial trials of vctl therapy required cmv lysates on apc, cmv-infected fibroblasts, or ebv-lymphoblastoid cells lines as a stimulant for expansion of donor-derived memory t cells [ , , ] . the discovery of dominant and highly conserved antigens such as cmv-pp and adenovirus hexon and penton led to replacement of live viral stimulation with either -mer peptide pools spanning viral proteins or dna plasmidtransduced antigen-presenting cells [ , ] . the newer approaches to rapidly expand and manipulate apcs enabled use of a less restricted population of donors and the targeting of an increased number of pathogens in a single culture [ , ] . in a recent rapid vctl protocol, the addition of il- and il- leads to production of cd + t cells with a th phenotype, whereas il- and il- tended to favor in vitro natural killer (nk) cell expansion [ ] . the ideal population to adoptively transfer may be ex vivo expanded central memory t cells with a cd l and cd ra phenotype as these cells have a superior ability to persist in vivo after adoptive transfer [ , ] . selection via cell capture sorting direct selection relies on cell sorting of immune donor pbmcs, usually after pulsing them with the antigen(s) of interest, to drive expansion of virus-specific t-cell clones [ ] . this approach would not be viable for obtaining immune ctls from pathogen-naive donors. multimer selection is achieved by binding of hla-peptide complexes to t-cell receptors (tcrs) of known antigen specificity, followed by purification of bound cells, e.g., by magnetic column separation. alternatively, antiviral t cells expressing interferon-γ (ifn-γ) can be isolated using the gamma capture assay. direct selection methods have the advantage of rapid manufacturing time. unfortunately, these approaches require apheresis of donors in order to collect sufficient cells for sorting and processing for clinical appli-cations and pre-existing and detectable pathogenspecific t cells in the blood. multimer selection is major histocompatibility (mhc)-restricted and selects only cd + t cells of a limited specificity. this could possibly allow pathogen evasion and impair persistence of vctl in vivo [ ] . earlier studies suggested that persistent binding of multimers to the tcr may impair t-cell function [ ] . recent reversible streptamer technology for direct selection may overcome the problem of impaired function [ ] . ifn-γ positive selection captures polyclonal antigen-specific cd + and cd + t cells and selects for a wider range of antigen-specific cells. combining direct selec-tion, culture expansion methods, and cytokine cocktails can optimize the selection of central memory t cells in vctl products and improve yields on targeted cellular phenotypes [ , ] . tcr or car gene modifications t cells can be modified to redirect their specificity with retroviral and lentiviral vectors to introduce the transgenes for high-affinity tcrs or chimeric antigen receptors (cars) consisting of a single-chain variable fragments (scfvs). high-affinity tcr genes can be cloned and transduced into polyclonal t cells to generate a large population of ( ) blood is obtained from donors (autologous, allogeneic, or umbilical cord blood) or is drawn or apheresis is performed to obtain a larger quantity of blood; ( ) pbmcs are processed via: (a) cell selection panel using multimers with a pathogen-derived peptide associated with a type-i hla molecule or column selection after in vitro stimulation of t cells with antigens followed by binding of ifnɣ or cd -expressing t cells with antibody-coated immu-nomagnetic beads; (b) cell expansion by stimulating the pbmc with apcs produced by antigenic peptide pools, viral transduction, or nucleofection; (c) genetic modification that involves the transfer of high-affinity pathogenspecific tcrs or cars to redirect the specificity of the t cells; and (d) polyclonal expansion of t cells for - days and arming with biabs directed at the pathogen of interest on one hand and the tcr on the other hand; ( ) quality control and release testing; and ( ) infusion into patients tcr pathogen-specific ctls [ ] . a similar strategy was used to produce tumor-specific t cells after tcr gene transfer [ ] . in contrast, cars have an extracellular region that consists of a scfv that binds to antigen, with an intracellular signaling complex composed of tcr zeta chain for first-generation cars, the tcr zeta chain and the cd for second-generation cars, and tcr zeta and cd or bb for third-generation cars [ ] [ ] [ ] . the high-affinity tcr-transduced ctls have been used to target cmv-infected cells [ ] , hpv-infected cells [ ] , hepatitis b-infected cells [ ] , hepatitis c-infected cells [ ] , tuberculosis-infected cells [ ] , sarsinfected cells [ ] , chlamydia-infected cells [ ] , and hiv-infected cells [ ] . car t cells were used to target cd in hiv-infected cells [ ] [ ] [ ] [ ] and for recognition of β-glucans in fungi [ ] . ex vivo ctl expansion is the most common method for producing clinical ctls for most clinical trials ( [ ] . cmv has been the primary focus of the first virus targeted therapy trials and remains a primary focus in subsequent studies (table . ). the first clinical report in which cd + cmvspecific ctls were isolated via tetramer selection [ ] generated complete or partial clinical responses in nine patients, but there was limited data on long-term persistence of the infused cmv-specific ctls. ifn-γ column selection (gamma capture, miltenyi) to produce cmv-ctls was associated with partial and complete responses in of patients who were given one dose of cmv-ctls [ ] . ifn-γ selection after stimulation with recombinant pp or an overlapping peptide pool of -mers covering the pp protein was used to produce cmv-ctl [ ] . infusions of cmv-ctls administered prophylactically after stem cell transplantation successfully protected seven patients from the development of viral reactivation and disease. further, in vivo expansion of cmv-ctls was detected in patients [ ] . cmv-ctls from hsct donors using reversible streptamers with mhc-restricted pp peptides were used to successfully treat two patients with cmv reactivation after hct [ ] . the strategy for using bispecific antibodies (biabs) to target cancer was nearly abandoned due to cytokine storm reactions. however, the last years has seen a resurrection of interest particularly for targeting t cells to various cancer antigens. studies using retargeted t cells have been reported for her in breast and prostate cancer using anti-cd x anti-her biab atc [ , ] ; egfr in colorectal, pancreatic, and lung cancer using anti-cd x anti-egfr biab atc [ ] ; and cd in non-hodgkin's lymphoma using anti-cd x anti-cd biab atc [ ] [ ] [ ] . since chemical or molecularly engineered constructs could be used to target the tcr on one hand and tumor-associated antigen (taa) on the other hand, we reasoned that cmv could be targeted by chemically heteroconjugating okt (anti-cd , anti-tcr) with cytogam® (polyclonal donor-derived anti-cmv igg, designated cmvbi) to kill cmv-infected fibroblasts [ ] . in this strategy shown in fig. . , anti-cd monoclonal antibody-activated t cells (atc) which expanded in low-dose il- were the t effector cells. atc alone do not kill cmvinfected targets. arming doses of cmvbi ranging from as low as . ng/ atc to ng/ atc exhibited high levels of specific anti-cmv cytotoxicity in targets infected with cmv at multiplicities of cmv infection (moi) ranging from . to . the polyclonal nature of the cytogam may provide multiple antibody clones directed at multiple cmv epitopes on the cmv-infected targets leading to the increased potency at a low arming dose of cmvbi. cytotoxicity was evident at effector-to-target ratios (e:t) of : , : , : , and : compared to unarmed atc alone. at an moi of . , the mean % specific anti-cmv-specific cytotoxicities at e:t of , , and were %, %, and %, respectively, whereas unarmed atc at the same e:ts killed < %. unarmed atc, cytogam®, or cmvbi alone did not exhibit significant killing of uninfected or cmv-infected fibroblasts. furthermore, cultures of cmvbiarmed atc with cmv-infected targets induced cytokine and chemokine release from cmvbiarmed atc. this simple targeting strategy bypasses mhc-restricted cytotoxicity for treating viral disease in organ transplant and hsct recipients. it was shown that cmvbi atc do not react to alloantigens in vitro in a mixed lymphocyte culture, and they can be frozen and reinfused at different time points as an "off-the-shelf" drug. although promising, it is not clear from these data whether targeting cmv or other disease agents using this approach will be clinically effective. ebv-ctls have been used for prevention and treatment of post-hsct lymphoproliferative disease (ptld) as well as ebv + lymphoma. irradiated ebv-lymphoblastoid cells (ebv-lcl) were used to generate ebv-specific ctls in vitro for prophylaxis or treatment for ebv-ptld in patients [ , ] . remarkably, the first patients received gene-marked ctls, and followup studies showed the gene-marked cells persisted up to months after hsct (table . ). hla-a -specific pentamers and ifn-γ selection procedures were used to produce ebv-ctls. hla-a specific pentamers were used to produce ebv-ctls from the haploidentical mother of a patient with ebv-ptld who had received a cord blood transplantation [ ] . a complete clinical response was obtained following two infusions of ebv-ctls. three of six patients with early ebv-induced ptld treated with ebv-ctls produced by ifn-γ selection achieved complete responses whereas three patients with advanced, multiorgan disease did not respond [ ] . the latest strategy is to target ebv with multiviral ctl products (below) or third-party-derived ebv-ctls. most studies targeting adenovirus (adv) use multiviral ctls [ , , , ] . a few exclusively target adv by selection technology. adv-ctls produced by ifn-γ selection was used for treatment of nine patients with drug-refractory adv infections [ ] . there was in vivo ctl expansion in five of six patients and four patients cleared their disease. in all studies using cell selection, clinical benefit was observed in spite of very low doses of vctls infused (< × cells/kg in most studies) [ , ] . recent antiviral ctl therapy trials target multiple viruses (cmv, ebv, and adv as primary targets). cmv, ebv, and adv are the three leading causes of viral-associated mortality after allogeneic hsct. clinical-grade adv vector ad f pp contains the immunodominant cmv antigen pp , providing a unique opportunity to transduce donor-derived dendritic cells or ebv-lcl to serve as apcs for the ctl cultures. triviral (cmv, ebv, and adv-specific) ctls were tested in a dose-escalation trial involving patients [ ] . there were no adverse effects at doses ranging from × to × cells/m , and all patients were effectively protected against cmv, ebv, and adv. interestingly, although ebv-and cmv-specific ctls were detected by ifn-γ elispots, adv-ctls were not detectable except during infection. in a follow-up trial using ad f -transduced ebv-lcl to produce ebvand adv-ctls, patients received prophylaxis or treatment for ebv and adv infections after hsct [ ] . although the ctls provided protection in vivo, the adv-ctls could not be detected except in the setting of adv infection; these data suggest that levels of specific vctls below the limits of detection by ifn-γ elispots provide protection and infection induces clonal expansion. similarly, ad f pp transduced dendritic cells (dc) used to produce cmv-and adv-ctls were clinically effective in patients after allogeneic hsct [ ] . there were a few cases of cmv reactivation in the setting of lowdose prednisone. this approach was applied to patients after allogeneic hsct with triviral (cmv, ebv, adv-specific) ctls using two methods: were produced by pulsing donor dcs with the hla-a restricted cmv peptide nlvpmvatv and were produced using ad f pp -transduced donor dcs [ ] . only of patients had cmv reactivation after ctl infusions and only of patients required antiviral drug therapy after steroid treatment for acute gvhd. advances in processing protocols have validated -mer peptide pools that include immunodominant viral antigens that replace viral transduction of apc thereby removing safety and regulatory barriers associated use of viral vectors [ ] . the use of gas-permeable rapid-expansion (g-rex) bioreactors has simplified ctl culture [ ] . these advances in technology led to the development of a rapid manufacturing protocol for expanding virus-specific t-cell products (vsts) that yield clinically relevant numbers of vsts in - days. further, vst products targeting multiple viral antigens have been shown to provide effective antiviral protection (against cnv, ebv and ad) in ten patients after hsct [ ] . this rapid manufacturing protocol was subsequently adapted to produce five virus-specific ctls targeting ebv, cmv, adv, hhv , and bk virus infections in a single t-cell product for patients following allogeneic hsct [ ] . fourteen of vst products manufactured from hsct donors recognized all viral components while ( %) recognized or more by ifn-γ elispots. unexpectedly of the donors were cmv seronegative and vsts produced predictably lacked cmv specificity. these vsts were used to treat patients after hsct. the patients treated prophylactically remained free of viral infections and patients with viral reactivations received vsts, with all experiencing partial or complete responses in their cmv, ebv, adv, or hhv infections. although there was intense interest in the use of ctl therapy for hiv, there was only limited success to date [ ] . attempts to expand and reinfuse autologous hiv-specific ctls resulted in only transient improvements in viral load [ ] . a larger number of clinical trials focused on genetically modified ctl to target hiv using transduction of a modified tcr or cars. these trials established safety, but exhibited limited antiviral efficacy [ , ] . a major challenge for this approach is the outgrowth of escape mutants expressing alterations of the target epitope so the infected cell can no longer be targeted by the effector cells. a more successful approach has been inserting genes that would provide hiv resistance. this approach was clinically tested when antisense gene complementary to hiv env was transduced into t cells from patients using lentiviral vectors [ ] . the ctls persisted for weeks, homed to gut-associated lymphoid tissue, and were well-tolerated with clinical toxicities. infusions of ctls in two of eight patients who underwent antiviral treatment interruption keep the viral load u ndetectable for and weeks. when ccr -delta mutations were introduced to cd enriched t cells through the use of a zinc-finger nuclease [ ] , the ccr -edited t cells were subsequently infused in patients, and engineered t cells were detectable in the peripheral blood for up to months post infusion. in six patients who underwent antiviral treatment interruption, the absolute number of gene modified cd + t cells decreased at a lower rate than non-modified t cells. recent studies showed that dual gene editing of cxcr and ccr via zinc-finger nucleases was successful in a t-cell line, and preclinical studies show that the t cells were highly resistant to hiv infection [ ] . it is not clear whether this approach could prevent primary infection or have a clinical impact as an hiv cure strategy. there are a few studies that target other viruses with adoptive immunotherapy. the john cunningham virus (jcv) is a ubiquitous polyoma virus which can cause progressive multifocal leukoencephalopathy (pml), which occurs in immunocompromised individuals such as acquired immunodeficiency syndrome (aids), recipients of hsct or solid organ transplants, or primary immunodeficiency disorders. donorderived jcv-specific ctls were used in a -year-old patient with pml after prolonged steroid treatment for gvhd following hsct. cells were manufactured using -mer peptide pools that included jc antigens vp and lt and infused twice leading to clearance of jv-dna from the cerebrospinal fluid with improvements in neurologic status [ ] . human papillomavirus (hpv) disease can be a late complication of hsct. peptide pools spanning the hpv e and e proteins were used to generate hpv-specific ctls from patients with oropharyngeal or cervical cancer that arise after hpv infection [ ] . the ctls exhibited specific activity directed at hpv e and e and antitumor activity against the hpv cervical cancer cell line caski. adverse events after infusions for patients on protocols by the groups at baylor college of medicine were reported [ ] . side effects were limited to mild adverse events observed within h of infusion; nausea and vomiting were most common with nonserious adverse events (fever, chills, nausea) that occurred within h. no significant gvhd was attributed to ctl infusions. the only significant complications were rare reports of systemic inflammatory responses in patients with bulky ebv+ lymphomas following ebv-ctl therapy. seven cases of acute gvhd occurred in patients who had a greater degree of hla mismatch than controls after infusions of ebv-ctl. some of the cases of gvhd were attributable to reducing the corticosteroid dose prior to the cmv-ctl infusions [ ] . for years, the selection or culture of anti-pathogen ctls was dependent on the presence of pathogen-specific memory t cells in the blood of donors, and, therefore, the approach could not help allograft recipients of pathogen-naive hematopoietic cell products after hsct. one strategy to address this problem is to provide "off-theshelf" pathogen-specific ctls derived from third-party donors. this strategy was first validated in a phase i trial involving patients who received partially matched ebv-ctls for ptld that developed after solid organ transplantation [ , ] and confirmed in a cohort of patients in a phase ii trial [ ] . the latter trial showed a response rate of % at weeks and % at months; the outcomes correlated with the degree of hla matching between the ctl donor and recipient. in the hsct patients, two patients with refractory ebv-ptld after cord blood transplantation (cbt) with third-party ebv-specific ctls [ ] . a bank of ctl lines with characterized activity against ebv, cmv, and adv were used to match for patients with refractory viral infections. this strategy resulted in partial or complete antiviral responses in %, %, and % of those with cmv, adv, and ebv, respectively [ ] . this is a marked improvement from standard therapy response rate of % in eight patients for whom a matched line could not be identified. despite partial hla matching at one to four loci, there were only two patients who developed grade i gvhd. clones that are responsible for gvhd have been selected against in the expansion culture and may exist at such low precursor frequencies after culture that they do not expand enough to cause clinically significant gvhd. the lower rate of response against ebv relative to cmv and adv may reflect selective expansion of t cells against immunodominant epitopes of the latter two viruses, thereby complicating the selection of an ideal third-party pathogen-specific line that fulfills the requirements of antiviral activity and mhc-restriction against multiple pathogens. the methods for producing third-party-virus-specific ctl include pentamer selection for ad, cmv, ebv [ ] , and ifn-γ selection for adv-ctl [ ] . a few studies reported transducing ctls with a virus-specific tcr [ , , ] . a trial of transgenic ctls using a retroviral vector that expresses a cmv-specific tcr is ongoing in the united kingdom (morris e. et al. mrc# g ). alternatively, kumaresan et al. transduced t cells with the β-glucan receptor dectin [ ] . since the carbohydrate β-glucan is found in the cell wall of most fungi [ ] , investigators used its natural receptor, dectin- , as a recognition receptor coupled to a cd (a key co-stimulatory molecule) and cd -zeta transgene to initiate signaling and killing in t cells. the same group showed that the antifungal carts could mediate damage to hyphae in vitro and in vivo [ ] . these novel approaches would allow creation of specific ctls from pathogennaive donors; however, they are subject to the regulatory challenges in gene transfer technology. furthermore, use of a single antifungal tcr allows for antigenic escape. a major advance in adoptive viral ctl therapy was development of virus-specific ctls from virus-naive donors. ctl could be produced from a % fraction from cord blood using donorderived dcs and ebv-lymphoblastoid cell line (lcl) as apc and ad f pp transduction as a source of cmv and adv antigens [ ] . the resulting viral ctls exhibited specific anti-cmv, ebv, and adv ifn-γ elispots responses as well as specific cr cytotoxicity with no alloreactivity. epitope mapping showed that the immunodominant epitopes recognized by cord blood-derived ctls were different from the immunodominant epitopes recognized by the cmv and ebv seropositive adult donors. the hla-a -restricted epitope nlvpmvatv was notably absent in the cord blood-derived lines. ctls derived from cord blood were successfully infused in cbt recipients in the ongoing act-cat trial (safety, toxicity and mtd of one intravenous iv injection of donor ctls specific for cmv and adenovirus, # nct ). recently, multiviral ctls were produced from cmv-naive adult donors using columnselected cd ra+ naive t cells stimulated by donor dcs pulsed with cmv -mer peptide pools [ ] . preclinical studies suggest that multiviral ctls will exhibit similar anti-cmv activity to dcs pulsed with cmv -mer peptide pools. ebv-ptld is a significant long-term risk in solid organ transplant recipients. rituximab can be effective, but treatment often requires reduction of immunosuppression which can lead to graft rejection. autologous ebv-ctls have been used in this setting [ ] . several prophylactic infusions of autologous ebv-ctls reduced the ebv viral load without adverse reactions despite ongoing treatment with calcineurin inhibitors [ ] . a heart transplant recipient who developed hodgkin's lymphoma-type ptld years after transplant had remission after being treated with autologous ebv-ctls in combination with chemotherapy without alterations in his immunosuppression [ ] . this observation supports the prior observations that calcineurin inhibitors block proliferation, but do not impair ctl activity. fungal infections are a major cause of morbidity and mortality in allogeneic hsct recipients, with gvhd being the major risk factor. candidal infections can range from mucocutaneous colonization of the skin and mouth to life-threatening systemic infections. aspergillus species are ubiquitous molds that cause invasive pulmonary infections as well as widespread infection including central nervous system dissemination in highly immunocompromised patients [ ] . patients with inherited immunodeficiencies (e.g., chronic granulomatous disease), patients with prolonged neutropenia after repeated rounds of chemotherapy (e.g., for acute leukemia), and those receiving immunosuppression after lung transplant or allogeneic hsct are at the highest risk for mycoses [ ] . the importance of t-cell immunity in defense against invasive aspergillosis and other filamentous fungi is not clear, since patients with these invasive fungal diseases usually have severe deficiencies in multiple components of the immune system. in patients with advanced aids, invasive aspergillosis is an uncommon complication and generally occurs when other forms of immune impairment (e.g., neutropenia and use of corticosteroids) are present. despite these unknowns, it may be clinically useful to target fungal infections with fungusspecific t cells after hsct. the adaptive immune response against invasive aspergillosis is believed to be orchestrated by cd + t cells. table . summarizes preclinical studies that developed fungal-specific ctls against candida, aspergillus, and rhizopus (a member of the mucorales group) species. aspergillus-specific ctls were produced by stimulation of pbmc with antigens from aspergillus extracts, selection with ifn-γ secretion, and culture [ ] . the ctls were predominantly cd +, cd r + memory cells that secrete ifn-ɣ in response to aspergillus and penicillium. the fungal-specific ctl enhanced hyphal damage by neutrophils and apcs. ifn-ɣ selection and stimulation with candida albicans, aspergillus fumigatus, and rhizopus oryzae extracts were used to produce multifungalspecific ctl lines, which were also nearly all cd + cd ro+ hla-dr+ that exhibited activation markers of ifn-ɣ, cd , and tnfα and enhanced oxidative activity of neutrophils when co-incubated with antigen and apcs [ ] . several studies target the candida mp and aspergillus crf antigens. to produce multipathogen-specific t cells that secrete ifn-ɣ, proliferate, and kill cmv, ebv, adv, candida, and aspergillus, donor pbmcs were incubated with peptide libraries from cmv-pp , ebv-lmp , adv-hexon, candida mp , and a -mer peptide from aspergillus crf [ ] . however, it remains unclear what the significance of mp and crf is in antifungal immunity [ ] [ ] . expanded memory/effector th cells following stimulation with rhizopus extracts were used to generate memory/effector th cells for mucormycosis, and the product exhibited specificity to the original rhizopus oryzae extract as well as other mucorales species [ ] . candida-specific t cells generated with cellular extracts of candida albicans released cytokines that caused hyphal damage and increased neutrophil activity against hyphae [ ] . ctls produced by stimulation with inactivated conidia (spores) from aspergillus fumigatus resulted in clonal cd + ctls with anti-aspergillus activity by ifn-ɣ elispots [ ] . these donor t-cell clones specific for aspergillus antigens were then infused in patients following haploidentical hsct. of patients who developed invasive aspergillosis, patients received anti-aspergillus ctls, while patients did not. nine of treated patients cleared their infections whereas only of untreated patients cleared their infections. aspergillus-specific ctls were detected in high frequencies in patients who received immunotherapy while they were barely detectable in untreated patients [ ] . despite notable advances in antifungal ctls, a better understanding of the immunodominant t-cell targets that should be selected for various fungal species is needed, and standardized clinical-grade cgmp fungal antigen sources are needed to provide consistency between trials. although there have been major advances in producing pathogen-specific ctls, important questions remain regarding methods that affect potency and efficacy of the t-cell products. it is unclear whether manufacturing ctls to include more pathogens in a single culture will affect potency and specificity in the ctl cultures. although the proportions of virus-specific ctls for each virus decrease as the number of antigens increases, these effects have not seemed to impact clinical trials. ctls specific for viruses (cmv, ebv, adv, bk, hhv , rsv, and influenza) pro-duced using peptide pools for antigens exhibited specific activity against all targeted viruses [ ] . the question remains as to whether adding additional viral targets will skew specific cytotoxicity, alter potency for each target, induce alloreactive t cells, or compromise in vivo responses. a major challenge is achieving consistent and optimal culture conditions for generating the most effective ctl product. although multiple rounds of stimulation with antigen select and expand the specific antiviral clones, prolonged culture may lead to t-cell exhaustion. some groups have decreased production time using newer bioreactors [ ] . identification of the "correct" subset of t cells for clinical use (however selected) will require well-designed randomized phase ii trials using a specific ctl product made by the same group or a common standard operating procedure (sop) in a homogeneous group of hsct patients. assays for measuring ifn-ɣ elispots and cytotoxicity need to be standardized and the timing of the studies needs to be the same. recently, a new population of healthy donors jolink [ ] transgenic tcr-transduced cells tuberculosis murine cells feng [ ] mhc-streptamer-enriched antigen-specific t cells listeria murine cells stemberger [ ] (proof of principle) -targeting of ova-expressing parasites murine cells polley [ ] transgenic trc-transduced cells chlamydia murine cells roan [ ] "stem cell memory t cells" has been putatively identified -which possess characteristics ideal for use in adoptive immunotherapy. unfortunately, there are no randomized phase ii trials to date to support continued development and commercialization of clinically effective ctls. the presence of immunosuppression remains a barrier for optimal immunotherapy after allogeneic hsct and solid organ transplantation since most agents also suppress ctl functions. nearly all protocols require recipients to be receiving less than . mg/kg/day of prednisone and wait at least days after anti-t-cell serotherapy to be eligible to receive ctl therapy. virtually all of the calcineurin inhibitors (cyclosporin a, tacrolimus, or sirolimus) at therapeutic doses impair ctl activity. ebv-specific ctls can be made resistant to tacrolimus by knockdown of fkbp via a retrovirally transduced specific sirna and exhibit anti-ebv lymphoma activity in the presence of tacrolimus [ ] . similarly, ebv-specific ctls can be made resistant to both cyclosporine a and tacrolimus by mutating calcineurin [ ] . the mutation does not alter the phenotype or antiviral activity of the ctls and mutated cells have a growth advantage in calcineurin inhibitors. although they have not been applied clinically, they have great potential for treating hsct and solid organ transplant recipients. there is one preclinical report of t cells used to target bacterial and parasitic infections [ ] , but there are no clinical trials evaluating t-cell immunotherapy for bacterial and parasitic infections. despite numerous studies evaluating in vitro t-cell responses, there is no consensus on the role of t cells in defense against aspergillosis. infusions of anti-pathogen ctls in several hundred patients over the past several decades have been established as a safe and highly effective therapy following allogeneic hct. identifying preserved viral t-cell epitopes, probing the antigen limits in ctl monoculture, testing the clinical efficacy of immunosuppressive-resistant ctls, and improving conditions for rapid and specific expansion will further broaden the usefulness of this treatment strategy. as advances in protocols and methods for manufacture achieve acceptable clinical standards that can be supported commercially, ctl therapy may become an integral component of care offered to allogeneic hsct or immunodeficiency patients. late cytomegalovirus disease and mortality in recipients of allogeneic hematopoietic stem cell transplants: importance of viral load and t-cell immunity marked increased risk of epstein-barr virus-related complications with the addition of antithymocyte globulin to a nonmyeloablative conditioning prior to unrelated umbilical cord blood transplantation adenovirus infection rates in pediatric recipients of alternate donor allogeneic bone marrow transplants receiving either antithymocyte globulin (atg) or alemtuzumab (campath) epidemiology and outcome of invasive fungal infection in adult hematopoietic stem cell transplant recipients: analysis of multicenter prospective antifungal therapy (path) alliance registry update in management of ganciclovirresistant cytomegalovirus infection antiviral drugs for cytomegalovirus diseases rising pp antigenemia during preemptive anticytomegalovirus therapy after allogeneic hematopoietic stem cell transplantation: risk factors, correlation with dna load, and outcomes cidofovir for cytomegalovirus infection and disease in allogeneic stem cell transplant recipients. the infectious diseases working party of the european group for blood and marrow transplantation cd monoclonal antibody (rituximab) for therapy of epstein-barr virus lymphoma after hemopoietic stem-cell transplantation the history of vaccination against cytomegalovirus infusion of cytomegalovirus (cmv)-specific t cells for the treatment of cmv infection not responding to antiviral chemotherapy clinical aspects of cmv infection after stem cell transplantation cytomegalovirus and varicellazoster virus vaccines in hematopoietic stem cell transplantation infusions of donor leukocytes to treat epstein-barr virus-associated lymphoproliferative disorders after allogeneic bone marrow transplantation the use of anti-cd and anti-cd monoclonal antibodies to clone and expand human antigen-specific t cells restoration of viral immunity in immunodeficient humans by the adoptive transfer of t cell clones cmv pp and ie- t cell epitopes recognized by healthy subjects identification of hexon-specific cd and cd t-cell epitopes for vaccine and immunotherapy t-cell therapy in the treatment of post-transplant lymphoproliferative disease functionally active virus-specific t cells that target cmv, adenovirus, and ebv can be expanded from naive t-cell populations in cord blood and will target a range of viral epitopes cytotoxic t lymphocyte therapy with donor t cells prevents and treats adenovirus and epstein-barr virus infections after haploidentical and matched unrelated stem cell transplantation large-scale expansion of dendritic cell-primed polyclonal human cytotoxic t-lymphocyte lines using lymphoblastoid cell lines for adoptive immunotherapy analysis of cd t cell reactivity to cytomegalovirus using protein-spanning pools of overlapping pentadecapeptides expansion of t cells targeting multiple antigens of cytomegalovirus, epstein-barr virus and adenovirus to provide broad antiviral specificity after stem cell transplantation reconstitution of cellular immunity against cytomegalovirus in recipients of allogeneic bone marrow by transfer of t-cell clones from the donor a phase i-ii trial to examine the toxicity of cmv-and ebv-specific cytotoxic t lymphocytes when used for prophylaxis against ebv and cmv disease in recipients of cd -selected/t cell-depleted stem cell transplants infusion of cytotoxic t cells for the prevention and treatment of epstein-barr virus-induced lymphoma in allogeneic transplant recipients administration of neomycin-resistance-genemarked ebv-specific cytotoxic t lymphocytes to recipients of mismatched-related or phenotypically similar unrelated donor marrow grafts immunotherapy targeting ebv-expressing lymphoproliferative diseases autologous epstein-barr virus (ebv)-specific cytotoxic t cells for the treatment of persistent active ebv infection adoptive immunotherapy for posttransplantation viral infections acquisition of full effector function in vitro paradoxically impairs the in vivo antitumor efficacy of adoptively transferred cd + t cells long-term outcome of ebv-specific t-cell infusions to prevent or treat ebv-related lymphoproliferative disease in transplant recipients allogeneic virus-specific t cells with hla alloreactivity do not produce gvhd in human subjects adoptive cellular therapy for early cytomegalovirus infection after allogeneic stem-cell transplantation with virusspecific t-cell lines generation of cmv-specific t lymphocytes using protein-spanning pools of pp -derived overlapping pentadecapeptides for adoptive immunotherapy rapidly generated multivirus-specific cytotoxic t lymphocytes for the prophylaxis and treatment of viral infections naïve t-cell-derived ctl recognize atypical epitopes of cmvpp with higher avidity than cmvseropositive donor-derived ctl -a basis for treatment of post-transplant viral infection by adoptive transfer of t-cells from virus-naïve donors adoptive transfer of effector cd + t cells derived from central memory cells establishes persistent t cell memory in primates molecular signatures distinguish human central memory from effector memory cd t cell subsets the role of virus-specific adoptive t-cell therapy in hematopoietic transplantation evasion of cd + t cells is critical for superinfection by cytomegalovirus reversible hla multimers (streptamers) for the isolation of human cytotoxic t lymphocytes functionally active against tumor-and virus-derived antigens adoptive transfer and selective reconstitution of streptamerselected cytomegalovirus-specific cd + t cells leads to virus clearance in patients after allogeneic peripheral blood stem cell transplantation immunobiology : the immune system in health and disease adoptive cellular therapy: a race to the finish line the promise and potential pitfalls of chimeric antigen receptors tumor-specific t-bodies: towards clinical application cmv-specific tcr-transgenic t cells for immunotherapy generating hpv specific t helper cells for the treatment of hpv induced malignancies using tcr gene transfer engineering virus-specific t cells that target hbv infected hepatocytes and hepatocellular carcinoma cell lines transduction of human t cells with a novel t-cell receptor confers anti-hcv reactivity development of genetically engineered cd + and cd + t cells expressing tcrs specific for a m. tuberculosis -kda antigen engineering t cells specific for a dominant severe acute respiratory syndrome coronavirus cd t cell epitope antigen-specific cd + t cells respond to chlamydia trachomatis in the genital mucosa reconstitution of anti-hiv effector functions of primary human cd t lymphocytes by transfer of hiv-specific alphabeta tcr genes t-cell engineering by a chimeric t-cell receptor with antibody-type specificity for the hiv- gp anti-hiv designer t cells progressively eradicate a latently infected cell line by sequentially inducing hiv reactivation then killing the newly gp -positive cells characterization of t cell-expressed chimeric receptors with antibodytype specificity for the cd binding site of hiv- gp lentiviral vectors encoding human immunodeficiency virus type (hiv- )-specific t-cell receptor genes efficiently convert peripheral blood cd t lymphocytes into cytotoxic t lymphocytes with potent in vitro and in vivo hiv- -specific inhibitory activity bioengineering t cells to target carbohydrate to treat opportunistic fungal infection adoptive transfer of cytomegalovirus-specific ctl to stem cell transplant patients after selection by hlapeptide tetramers adoptive transfer of pp -specific t cells for the treatment of chemorefractory cytomegalovirus disease or reactivation after haploidentical and matched unrelated stem cell transplantation directly selected cytomegalovirus-reactive donor t cells confer rapid and safe systemic reconstitution of virus-specific immunity following stem cell transplantation reconstitution of ebv-specific t cell immunity in solid organ transplant recipients complete regression of posttransplant lymphoproliferative disease using partially hla-matched epstein barr virus-specific cytotoxic t cells treatment of epstein-barr-virus-positive post-transplantation lymphoproliferative disease with partly hlamatched allogeneic cytotoxic t cells allogeneic cytotoxic t-cell therapy for ebv-positive posttransplantation lymphoproliferative disease: results of a phase multicenter clinical trial a novel haploidentical adoptive ctl therapy as a treatment for ebv-associated lymphoma after stem cell transplantation effective and long-term control of ebv ptld after transfer of peptide-selected t cells successful treatment of ebv-associated posttransplantation lymphoma after cord blood transplantation using third-party ebv-specific cytotoxic t lymphocytes successful treatment of a classic hodgkin lymphoma-type posttransplant lymphoproliferative disorder with tailored chemotherapy and epstein-barr virus-specific cytotoxic t lymphocytes in a pediatric heart transplant recipient thirdparty virus-specific t cells eradicate adenoviraemia but trigger bystander graft-versus-host disease safe adoptive transfer of virus-specific t-cell immunity for the treatment of systemic adenovirus infection after allogeneic stem cell transplantation safety of autologous, ex vivo-expanded human immunodeficiency virus (hiv)-specific cytotoxic t-lymphocyte infusion in hiv-infected patients prolonged survival and tissue trafficking following adoptive transfer of cd zeta gene-modified autologous cd (+) and cd (+) t cells in human immunodeficiency virus-infected subjects a phase ii randomized study of hiv-specific t-cell gene therapy in subjects with undetectable plasma viremia on combination antiretroviral therapy antiviral effects of autologous cd t cells genetically modified with a conditionally replicating lentiviral vector expressing long antisense to hiv gene editing of ccr in autologous cd t cells of persons infected with hiv polyomavirus jc-targeted t-cell therapy for progressive multiple leukoencephalopathy in a hematopoietic cell transplantation recipient rapid salvage treatment with virus-specific t cells for therapyresistant disease transferring functional immune responses to pathogens after haploidentical hematopoietic transplantation monoculturederived t lymphocytes specific for multiple viruses expand and produce clinically relevant effects in immunocompromised individuals prophylactic infusion of cytomegalovirusspecific cytotoxic t lymphocytes stimulated with ad f pp gene-modified dendritic cells after allogeneic hemopoietic stem cell transplantation multicenter study of banked third-party virusspecific t cells to treat severe viral infections after hematopoietic stem cell transplantation safety and clinical efficacy of rapidly-generated trivirus-directed t cells as treatment for adenovirus, ebv, and cmv infections after allogeneic hematopoietic stem cell transplant donor-derived cmv-specific t cells reduce the requirement for cmv-directed pharmacotherapy after allogeneic stem cell transplantation activity of broad-spectrum t cells as treatment for adv, ebv, cmv, bkv, and hhv infections after hsct targeted t-cell therapy in stage iv breast cancer: a phase i clinical trial phase i study of anti-cd x anti-her bispecific antibody in metastatic castrate resistance prostate cancer patients anti-cd x anti-egfr bispecific antibody redirects t cell cytolytic activity to egfr-positive cancers in vitro and in an animal model t cells armed with anti-cd x anti-cd bispecific antibody enhance killing of cd + malignant b-cells and bypass complement-mediated rituximab-resistance in vitro cd -targeted t cells after stem cell transplantation for high risk and refractory non-hodgkin's lymphoma multiple infusions of cd -targeted t cells and low-dose il- after sct for high-risk non-hodgkin's lymphoma: a pilot study targeting cytomegalovirus-infected cells using t cells armed with anti-cd × anti-cmv bispecific antibody accelerated production of antigen-specific t cells for preclinical and clinical applications using gas-permeable rapid expansion cultureware (g-rex) t-cell therapies for hiv simultaneous zinc-finger nuclease editing of the hiv coreceptors ccr and cxcr protects cd + t cells from hiv- infection human papillomavirus type e /e -specific cytotoxic t lymphocytes for adoptive immunotherapy of hpv-associated malignancies adverse events following infusion of t cells for adoptive immunotherapy: a -year experience human mhc class i-restricted high avidity cd t cells generated by co-transfer of tcr and cd mediate efficient tumor rejection in vivo cord blood t cells retain early differentiation phenotype suitable for immunotherapy after tcr gene transfer to confer ebv specificity beta-glucan recognition by the innate immune system immunity to fungal infections current challenges in the diagnosis and management of invasive fungal infections: report from the th international symposium on infections in the immunocompromised host generation of highly purified and functionally active human th cells against aspergillus fumigatus generation of a multipathogen-specific t-cell product for adoptive immunotherapy based on activation-dependent expression of cd clinicalscale generation of multi-specific anti-fungal t cells targeting candida, aspergillus and mucormycetes clinical-scale generation of human anti-aspergillus t cells for adoptive immunotherapy robust polyfunctional t-helper responses to multiple fungal antigens from a cell population generated using an environmental strain of aspergillus fumigatus generation and characterization of anti-candida t cells as potential immunotherapy in patients with candida infection after allogeneic hematopoietic stem-cell transplant cross-protective th immunity against aspergillus fumigatus and candida albicans characterization of the t-cell-mediated immune response against the aspergillus fumigatus proteins crf and catalase in healthy individuals cd + and cd + t cells mediate adoptive immunity to aerosol infection of mycobacterium bovis bacillus calmette-guerin lowest numbers of primary cd (+) t cells can reconstitute protective immunity upon adoptive immunotherapy adoptive immunotherapy against experimental visceral leishmaniasis with cd + t cells requires the presence of cognate antigen biochemical and immunological characterization of mp , a major mannoprotein antigen of the opportunistic human pathogen candida albicans characterization of the cellular immune responses to rhizopus oryzae with potential impact on immunotherapeutic strategies in hematopoietic stem cell transplantation generation of epstein-barr virus-specific cytotoxic t lymphocytes resistant to the immunosuppressive drug tacrolimus (fk ) generation of ebv-specific cytotoxic t cells that are resistant to calcineurin inhibitors for the treatment of posttransplantation lymphoproliferative disease acknowledgments special thanks to the clinical coordinators for dedicating their efforts to serve the immunotherapy patients. we thank manley huang for his thoughtful reading of the chapter. the studies were supported in part by r ca (lgl) and r ca (lgl), translational grants # - (lgl) and # - (lgl) from the leukemia & lymphoma society, and uva cancer center support grant nci p ca - .lgl is a founder of transtarget, inc. cmb is supported in part by the nichd k -hd- award to mdk and cprit r rp and nci p ca - awards to cmb. key: cord- -dh j lyl authors: haskologlu, sule; kostel bal, sevgi; islamoglu, candan; aytekin, caner; guner, sukru; sevinc, selin; keles, sevgi; kendirli, tanil; ceylaner, serdar; dogu, figen; ikinciogullari, aydan title: clinical, immunological features and follow up of patients with dedicator of cytokinesis (dock ) deficiency date: - - journal: pediatr allergy immunol doi: . /pai. sha: doc_id: cord_uid: dh j lyl biallelic mutations in the dedicator of cytokinesis gene (dock ) cause a progressive combined immunodeficiency (cid) characterized by susceptibility to severe viral skin infections, atopic diseases, recurrent respiratory infections, and malignancy. hematopoietic stem cell transplantation (hsct) is only curative treatment for the disease. however, there is limited information about long‐term outcome of hsct and its effect to protect against cancer development in dock ‐deficient patients. in this study, we retrospectively evaluated clinical and immunologic characteristics of dock ‐deficient patients and outcome of patients who underwent hsct. we aimed to report the experience of our center and the result of the largest transplantation series of dock deficiency in our country. median follow‐up time is months (min‐max: ‐ ) in all patients and months (min‐max: ‐ ) in transplanted patients. atopic dermatitis ( / ), recurrent respiratory tract infections ( / ), and food allergy ( / ) were the most frequent clinical manifestations. failure to thrive ( / ), liver problems ( / ), bronchiectasis ( / ), chronic diarrhea ( / ), and autism spectrum disorders ( / ) were remarkable findings in our series. elevated ige level ( / ) and eosinophilia ( / ), low igm level ( / ), and decreased cd + t ( / ) and cd + t ( / ) cell count were prominent laboratory findings. hsct was performed in patients. all patients achieved adequate engraftment and showed improvement in their clinical and immunologic findings. atopic dermatitis and food allergies improved in all patients, and their dietary restriction was stopped except one patient who was transplanted recently. the frequency of infections was decreased. the overall survival is % in hsct‐received patients and % in all. hsct at the earliest possible period with most suitable donor‐ and patient‐specific appropriate conditioning regimen and gvhd prophylaxis is lifesaving for dock deficiency cases. and its effect to protect against cancer development in dock -deficient patients. in this study, we retrospectively evaluated clinical and immunologic characteristics of dock -deficient patients and outcome of patients who underwent hsct. we aimed to report the experience of our center and the result of the largest transplantation series of dock deficiency in our country. median follow-up time is months (min-max: in all patients and months (min-max: in transplanted patients. atopic dermatitis ( / ) , recurrent respiratory tract infections ( / ) , and food allergy ( / ) were the most frequent clinical manifestations. failure to thrive ( / ) , liver problems ( / ) , bronchiectasis ( / ) , chronic diarrhea ( / ), and autism spectrum disorders ( / ) were remarkable findings in our series. elevated ige level ( / ) and eosinophilia ( / ) , low igm level ( / ) , and decreased cd + t ( / ) and cd + t ( / ) cell count were prominent laboratory findings. hsct was performed in patients. all patients achieved adequate engraftment and showed improvement in their clinical and immunologic findings. atopic dermatitis and food allergies improved in all patients, and their dietary restriction was stopped except one patient who was transplanted recently. the frequency of infections was decreased. the overall survival is % in hsct-received patients and % in all. hsct at the earliest possible period with most suitable donor-and dock deficiency is an autosomal recessive cid syndrome characterized by atopy (dermatitis, food allergies, and asthma), recurrent sinopulmonary and severe/persistent cutaneous viral infections, mucocutaneous candidiasis, early-onset malignancy, and elevated ige levels and eosinophilia. - dock protein is a member of the dock -related family, which is involved in the cytoskeletal rearrangements. dock is required for t lymphocyte, natural killer (nk) cell function and survival, dendritic cell migration, b-cell proliferation, immunoglobulin production, and antiviral cytokine production. [ ] [ ] [ ] [ ] atopic dermatitis (ad), food allergy (fa), and wheezing are usually the initial symptoms of dock deficiency during the early infancy period. however, chronic or severe infections, autoimmunity, and cancer development may affect the course of the disease and survival. , hsct is the lifesaving therapy in dock -deficient patients; it should be performed as early as possible, before the occurrence of malignancies and fatal infections. hsct has been shown to cure nearly all clinical and laboratory manifestations of dock deficiency by reconstituting the normal function of the immune system. [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] since the parental consanguinity rate is high ( . %) in turkey, dock deficiency has an important place among cids. here, we retrospectively evaluated the clinical and immunologic features and treatment modalities of patients with dock deficiency and follow-up results of hematopoietic stem cell transplant (hsct) in patients among them as a single-center experience. twenty patients ( girls and boys) from families diagnosed and followed up with dock deficiency from to in ankara university school of medicine, department of pediatric immunology-allergy of children's hospital, were evaluated retrospectively. dock protein expression was evaluated by flow cytometry as previously described. next-generation sequencing (ngs) and then sanger's sequencing methods were used to detect and validate the dock mutations, respectively. the study was approved by the local ethics committee of ankara university faculty of medicine and performed in accordance with the principles of the declaration of helsinki. the written informed consent was obtained from all the patients and/or their parents. eleven of / patients (six girls and five boys) are transplanted so far. the median time from diagnosis to hsct was months ( - months). myeloablative conditioning (mac) regimen consisting of iv fludarabine mg/m /day and iv busulfan mg/kg/day on days − , − , − , and − was administered to two patients (p and p ), while all others received treosulfan-based reduced-intensity conditioning (ric). treosulfan was given at a dose of or g/m (< year of age) in three divided doses on − , − , and − days. csa was given alone in six patients transplanted from mrd and in combination with mycophenolate mofetil (mmf) or methotrexate (mtx) in two patients where peripheral blood-derived stem cells (pbscs) were used. one patient received tacrolimus and mtx. antithymocyte globulin (atg) was added to the conditioning regimen in a mud ( of hla matched) and a haploidentical (in conjunction with post-transplantation cyclophosphamide (pt/cy) for gvhd prophylaxis) donor setting. acute and chronic gvhds were graded according to modified glucksberg and nih consensus criteria, respectively. , immunosuppression was stopped at months after hsct if there was no evidence of cgvhd. myeloid engraftment was defined as the absolute neutrophil count being higher than . × /l and platelet counts exceeding × /l without transfusion for at least three consecutive days. immune reconstitution was evaluated by measuring the levels of cd +, cd +, cd +, cd +, cd +, cd + +, cd + cd ra+, and cd ra+ cd patient-specific appropriate conditioning regimen and gvhd prophylaxis is lifesaving for dock deficiency cases. clinic, dock deficiency, follow-up, hematopoietic stem cell transplantation, immunological features in our patients with dock deficiency, we obtained some different findings with classical findings. in addition, we present the results of the largest transplantation series of dock deficiency in our country. (recent thymic emigrant, rte) cells with flow cytometry (beckman coulter, navios software, kaluza, miami, usa). dock protein expression was analyzed with flow cytometry before and after months following hsct as described previously (figure ). myeloid and t-cell donor chimerisms were measured by using pcrbased amplification of short tandem repeat sequences in dna of the cells following the separation of peripheral blood samples at + st, nd, rd, th, th, th, and th months following hsct by automated magnetic cell sorting (miltenyi biotec). full donor chimerism was defined when > % of the cells originated from the donor. all patients were nursed in an isolated room, and they all received weekly prophylactic, antimicrobial, and ivig therapies. the clinical characteristics, genetic features, and treatment modalities of the patients are presented in table . the immunologic data are given in figure . all patients were born to consanguineous parents. the median age at diagnosis and at hsct was years in addition to previously reported data about a patient (p ), all patients received ig replacement therapy, trimethoprim-sulfamethoxazole, acyclovir, and fluconazole prophylaxis. severe oral herpes infection of p was treated successfully with interferon α b, while in p , interferon α b treatment was ceased due to side effects (fever and myalgia) early after its initiation. a total of three patients died: two (p and p ) because of intracranial nhl and one (p ) because of cholestatic liver failure before the definitive dock diagnosis. donor screening is still being processed in six patients. table . table . here, we report clinical and immunologic findings of dock deficient patients while reviewing the outcomes of transplanted cases among them. our study is the largest single-center cohort reported from turkey. furthermore, six novel mutations were reported in this cohort. our patients had unique features that turkish patients participating in previous studies did not have. , in addition to classical clinical features, gastrointestinal problems (liver and inflammatory bowel disease) were more common, while viral skin infections were less common in our series compared with the literature. , bcg vaccine-related complications were seen in five patients for the first time. the median age of our patients ( years, month- years) was lower than the previous studies, and mild clinical findings were found in patients diagnosed during infancy period. molluscum contagiosum or herpes zoster infections were not detected in our series. hpv was detected in only one patient. mucosal herpes simplex virus (hsv) infections were observed at similar rates with the literature ( %). gastrointestinal problems (liver and inflammatory bowel disease) in this study were more common than in previous studies. , these abovementioned differences might be due to patients' young age, the characteristics of the genetic mutations, and epigenetic factors influencing the genotype. there is limited information about the speech and developmental problems in dock deficiency. three of our patients have speech problems and pervasive developmental problems. because all these patients' parents are consanguineous and two of the patients are siblings, it suggests that other genetic etiologies may be responsible for the pdd. more than % of the non-transplanted patients are reported to die due to malignancy or severe infections before the age of years. in our series, three patients died before the detection of dock mutations due to central nervous nhl (p and p ) and liver failure (p ). thus, hsct is suggested to all patients with dock deficiency; even patients with significant comorbidities should undergo hsct as early as possible after aggressive treatments of these conditions to minimize complications and to achieve engraftment. [ ] [ ] [ ] [ ] [ ] recently, hsct outcomes of transplanted patients with - in our study, cgvhd rate is higher than the previous studies. [ ] [ ] [ ] [ ] it could be related to severe organ damage and inflammatory morbidities due to delayed diagnosis and hsct in our cohort. all patients who developed cgvhd were older than years of age. severe cgvhd was the most important complication we have seen after hsct in dock -deficient patients. previous series report persistence of food allergies after hsct. , however, in our series food allergies were improved in all patients except for one patient who was transplanted recently. we did not observe allergic reactions in the patients after stopping dietary restrictions. although the underlying cause of development of autoimmunity is not clear in dock deficiency, autoimmune diseases-even celiac disease-disappeared in our patients after transplantation. eight of patients undergoing hsct had failure to thrive. only two patients were able to achieve normal growth after is lifesaving for dock deficiency. cgvhd is the most important complication after hsct. in our opinion, gvhd prophylaxis is very important and gvhd findings should be followed very closely and treated aggressively. long-term follow-up is necessary to see how immunologic features progress and how effects on the prevention of malignancy and survival in transplanted patients will be. despite most of the mutations found in dock -deficient patients are loss-of-function mutations that abolish dock protein expression, very low but detectable levels of dock protein can be expressed in some patients. the residual dock protein contributes to the variable disease phenotype. in our cohort, four patients with mild phenotype (p , p , p , and p ) diagnosed in early infancy demonstrated some residual dock protein expression by flow cytometric analysis. therefore, examination of dock protein expression by flow cytometry in early infancy in suspected patients is a rapid and reliable diagnostic approach. we suggest that all patients suspected to have dock deficiency be followed up closely, and dock expressions be measured by flow cytometry and confirmed by genetic study and performing hsct as soon as possible. the authors declare that they have no conflicts of interest. sule haskologlu https://orcid.org/ - - - autosomal recessive hyperimmunoglobulin e syndrome: a distinct disease entity large deletions and point mutations involving the dedicator of cytokinesis (dock ) in the autosomal-recessive form of hyper-ige syndrome su hc combined immunodeficiency associated with dock mutations dedicator of cytokinesis (dock ) deficiency dock is a cdc activator critical for interstitial dendritic cell migration during immune responses dock functions as an adaptor that links tlrmyd signaling to b cell activation dock regulates signal transduction events to control immunity dock deficiency: clinical and immunological phenotype and treatment options-a review of patients the extended clinical phenotype of patients with dedicator of cytokinesis deficiency inborn errors working party of the european group for blood and marrow transplantation and the european society for primary immunodeficiencies. hematopoietic stem cell transplantation as treatment for patients with dock deficiency matched related and unrelated donor hematopoietic stem cell transplantation for dock deficiency hematopoietic stem cell transplantation outcomes for patients with dedicator of cytokinesis (dock ) deficiency haploidentical related donor hematopoietic stem cell transplantation for dedicator of cytokinesis deficiency using post transplantation cyclophosphamide curative treatment of autosomal-recessive hyper-ige syndrome by hematopoietic cell transplantation successful long-term correction of autosomal recessive hyper-ige syndrome due to dock deficiency by hematopoietic stem cell transplantation successful engraftment of donor marrow after allogeneic hematopoietic cell transplantation in autosomal-recessive hyper-ige syndrome caused by dedicator of cytokinesis deficiency successful allogeneic hematopoietic stem cell transplantation for dock deficiency successful bone marrow transplantation for dock deficient hyper ige syndrome clinical and immunological correction of dock deficiency by allogeneic hematopoietic stem cell transplantation following a reduced toxicity conditioning regimen in dock deficiency donor cell engraftment post-genoidentical hematopoietic stem cell transplantation is possible without conditioning successful hematopoietic stem cell transplantation after myeloablative conditioning in three patients with dedicator of cytokinesis deficiency (dock ) related hyper ige syndrome hematopoietic stem cell transplantation from unrelated donors in children with dock deficiency flow cytometry diagnosis of dedicator of cytokinesis (dock ) deficiency consensus conference on acute gvhd grading national institutes of health consensus development project on criteria for clinical trials in chronic graft-versus-host disease: i. the. diagnosis and staging working group report plasmacytoid dendritic cell depletion in dock deficiency: rescue of severe herpetic infections with ifn-α b therapy insights into immunity from clinical and basic science studies of dock immunodeficiency syndrome gvhd. c p 's immunosuppressive treatment was discontinued mo ago, and p was still receiving immunosuppressive treatment. key: cord- -vac r authors: nan title: physicians abstracts: ebmt date: - - journal: bone marrow transplant doi: . /bmt. . sha: doc_id: cord_uid: vac r nan b cells have been demonstrated to present antigen to t cells in vivo. cd -activation dramatically improves antigen presentation by normal and malignant b cells and has therefore been studied as an approach to generate autologous "non-artifi cial" antigen presenting cells for active immunotherapy. furthermore, cd -b cells have recently been shown to expand tumorantigen and viral specifi c ctl as well as regulatory t cells and are therefore of great interest for post-transplant immunotherapy. human b cells when activated via cd -l/il- can be expanded from small amounts of peripheral blood in - days. cd -activated b cells can prime naïve cd + and cd + t cells, expand memory t cells and express important surface homing molecules. nevertheless, it remains unclear whether cd -activated b cells migrate to secondary lymphoid organs (slo) in vivo to attract and interact with t cells. to address this question we established a methodology to generate murine cd -activated b cells. at day of culture, these cells are > % cd + and cd / /mhci/mhciihi. murine cd -activated b cells present a 'homing phenotype'; migrate toward slo chemokines such as ccl , ccl and cxcl ; and induce t-cell chemotaxis in vitro. upon cd l activation, b cells up-regulate ccr while down-regulating cxcr expression which suggests direction of activated b cells toward the b-zone/t-zone boundary. we compared the homing of gfp + cd -activated b cells to resting gfp + b cells and show for the fi rst time that cd -activated b cells home to slo significantly more effi ciently than resting b cells. furthermore, cd activated b cells localize in the b-cell areas, and a signifi cant fraction move to the b-t boundary close to the t-cell zone over time. to dissect t-cell-apc interactions on a single cell we analyzed three-dimensional migration in collagen matrix using time-lapse videomicroscopy. interestingly, antigen-loaded cd -activated b cells differ from immature and mature dc by displaying a rapid migratory pattern undergoing highly dynamic, short-lived ( . min) and sequential interactions with cognate t cells. taken together, these data revealed that cd -activated b cells can home to secondary lymphoid organs and interact dynamically with t cells thus underlining their potential as cellular adjuvant for cancer immunotherapy. long-term follow-up of upfront tandem autologous -ric (reduced intensity conditioning) allogeneic transplantation versus autologous transplantation (nmam ) in multiple myeloma g. gahrton, b. björkstrand, s. iacobelli, u. hegenbart, a. gruber, h. greinix, l. volin, f. narni, p. musto, m. beksac, a. bosi, g. milone, p. corradini, h. goldschmidt, t. de witte, c. objectives: treatment of multiple myeloma with allogeneic hematopoietic stem cell transplantation is controversial. the nmam study compares tandem autologous (asct)/ reduced intensity conditioning (ric) allogeneic transplantation (ricallo) to only asct or asct in tandem (asct ) in a prospective study based on genetic randomization. patients and methods: out of myeloma patients accrued during - from ebmt centres with an hlaidentical sibling donor were allocated to tandem asct/ricallo and without to asct only. previously untreated patients with at least stable disease after vad (vincristine, doxorubicine, dexamethasone)-like induction treatment were included at the time of conditioning for asct. single (n = ) or tandem (n = ) asct was optional in the asct arm. conditioning for asct was melphalan mg/m , and for ricallo fl udarabine mg/m x plus tbi gy. the two treatment groups were well matched for standard prognostic parameters and response status at asct. results: on an intention to treat basis the cumulative and months non-relapse-mortality (nrm) was % and % with asct/ricallo and % and % with asct respectively (p = . (gray test)). the cr rate within months was % with asct/ricallo and % with asct (p = . : improved in time (fine/gray model)). at months after asct relapse/progression rate was % and % (p = . : reduction in time fine/gray model)), pfs % and % (p = . reduction of risk in time (cox)) and os % and % (p = . reduction in time (cox)) (at months % and %) for asct/ricallo and asct respectively. a comparison between those patients who received a second allo (n = ) versus a second auto (n = ) relapse/progression rate at months from second transplant was % and % and pfs % and % in asct/ricallo and auto respectively. information about the chromosome deletion (del( q )) was present in patients. in patients with the deletion os at months was % and %, and pfs % and % for asct/ricallo and asct respectively. gvhd in ricallo was as expected (agvhd grade - : %; grade - : %; cgvhd limited: %; extensive: %). conclusion: the risk of myeloma relapse is lower with tandem asct/ricallo as compared to asct or tandem asct, both in an intention to treat analysis and in patients that received the correct treatment. nrm is higher, but considered acceptable in view of the improved pfs and os with tandem asct/ricallo. a fi ve biomarker panel predicts acute graft-versus-host disease s. paczesny ( ) , d. bickley ( ) , s. choi ( ) , j. crawford ( ) , t. braun ( ) , s. pitteri ( ) , j. hogan ( ) , p. reddy ( ) , s. hanash ( ) , j. ferrara* ( ) , j. levine* ( ) ( )university of michigan (ann arbor, us); ( )fred hutchinson cancer research center (seattle, us) * have contributed equally to this work current evidence suggests that a composite panel of four biomarker proteins (il ra, tnfr , il , hgf) is diagnostic and prognostic for acute graft-versus-host disease (gvhd). elafi n is a biomarker that has also been found to be diagnostic for gvhd of the skin, as well as prognostic for overall survival (os). we sought to evaluate whether these fi ve biomarkers are able to predict future occurrence of gvhd, non-relapse mortality (nrm) and os. we measured by sequential elisa, levels of these fi ve proteins in plasma collected prospectively from allogeneic hct patients randomly divided into training ( gvhd-, gvhd + ) and validation ( gvhd-, gvhd + ) sets. we obtained samples - days before onset of gvhd (median day ) and at equivalent time points in patients without gvhd. there were no signifi cant differences between sets in age, conditioning intensity, donor source, hla match or gvhd grade between training and validation sets. the median day of sample acquisition was day and day , respectively. logistic regression determined that a linear combination of the fi ve proteins levels predict the occurrence of acute gvhd in the training set. the receiver operating characteristic curves of each of the fi ve individual biomarkers are shown in figure with an area under the curve (auc) for the composite panel of . ( %ci: . - . ). when this model was applied to samples of the validation set, the corresponding auc was . ( %ci: . - . ). this -biomarker panel therefore discriminated between patients who later developed gvhd and those who did not. proportional odds logistic regression models determined that the panel gave prognostic information regarding the eventual maximum grade of gvhd (p< . in training set, p = . in validation set). given this correlation, we next divided the patients into high and low risk groups based on their predicted probability of developing gvhd (high risk ≥ . and low risk < . ) and analyzed these groups for differences in nrm, relapse mortality and os. the differences in year nrm and os between groups were signifi cant in both the training and validation sets (table ). when adjusted for other known risk factors (age, conditioning intensity, donor source, and hla match), the difference in os remains signifi cant (p = . ) in both sets. in conclusion, a -biomarker panel can predict gvhd before any clinical manifestation and provides prognostic information including long term survival. s their interaction with dendritic cells. recent clinical data has indicated that the content of plasmacytoid dendritic cells (pdc) in an allograft is associated with the risk of relapse following allogeneic bone marrow transplantation (bmt). we have previously shown that the addition of donor pdc to a graft comprised of purifi ed hsc and t-cells led to enhanced th activation of donor t-cells with improved gvl activity without increasing gvhd in multiple murine bmt model systems (li and waller ji ). here we studied the mechanism that donor pdc augment gvl without increasing gvhd, and present a novel model for regulation of post-transplant immunity. methods: donor hematopoietic stem cells (hsc), t-cells and pdc in the allograft were rigorously purifi ed using immuno-magnetic selection and high-speed fl uorescent-activated cytometry (facs) in the h b b ->h k b .br transplant model. donor cell subsets purifi ed from wild type (wt) or interferon gamma knock-out(ko),(ifn-gko), interferon gamma receptor ko (ifn-grko), and indolamine , dioxygenase (ido) ko (ido-ko) were combined to determine the role of the ifn-g and ido in the separation of gvhd from gvl in murine bmt. results: in the b ->b .br bmt model, the addition of , donor pdc to a graft comprised of , hsc and , t-cells lead to th immune polarization and enhanced proliferation of donor t-cells with higher levels of donor t-cell chimerism and improved gvl activity without increasing gvhd. the absence of allo-reactivity was dependent upon donor t-cell synthesis of ifn-g and the presence of ifn-gr on donor pdc. co-culture of t-cells with syngenic pdc in one-way mlr lead to up-regulation of ido in donor pdc. increased gvhd after transplanting pdc from ido ko donors in combination with wt t-cells and hsc demonstrated the central role of ido in the initiation of counter-regulatory effects that limit gvhd. wt t-cells and pdc lead to the generation of donor-derived t-reg in bmt recipients that limited gvhd. pdc from ido-ko donors had markedly reduced numbers of donor t-reg. conclusions: donor pdc regulate the initial activation and gvl activity of donor t-cells and subsequently generate t-reg that limit gvhd. the dynamic regulation of the activation and alloreactivity of donor t-cells by donor pdc suggest novel clinical approaches to enhance gvl activity without gvhd. j. peccatori ( ) , d. clerici ( ) , a. forcina ( ) , m. bernardi ( ) , r. crocchiolo ( ) , c. messina ( ) , m. noviello ( ) , s. mastaglio ( ) , f. giglio ( ) , s. malato ( ) , m.t. lupo stanghelllini ( ), s. marktel ( ) , a. assanelli ( ) , m. battaglia ( ) , a. ferraro ( ) , s. rossini ( ) , m.e. bernardo ( ) , a. bondanza ( ) , m. g. roncarolo ( ) , c. bonini ( ) , f. locatelli ( ) , f. ciceri ( ) ( background and aim: rapamycin, in contrast to calcineurin inhibitors, allows t regulatory cell (t-regs) proliferation while inhibits effector t cell expansion. we investigated the safety of infusion of t-cell repleted unmanipulated pbsc from haploidentical donor with a combination of rapamycin, mycophenolate and atg as gvhd prophylaxis, to preserve early t-regs function (trramm study, eudract - - ) . patients and methods: since , fi fty-nine patients (pts) underwent sct for aml ( pts), all ( ), mds ( ), cml-bc ( ), nhl ( ) or hd ( ) . median age was years (range - ). at transplantation pts were in early phase, and in advanced phase. median comorbidity index (ci) score was ( - ). the conditioning regimen included treosulfan ( g/m for days), fludarabine ( mg/m for days) and an invivo t and b-cell depletion by atg-fresenius ( mg/kg for times) and rituximab (a single mg dose). all pts received allogeneic unmanipulated pbsc from an hla-haploidentical related donor. gvhd prophylaxis consisted of rapamycin (target level - ng/ml, till day + ) and mmf ( mg/kg tid till day + ). results: all patients engrafted and all but eight were in disease remission at fi rst marrow evaluation on day + . cumulative incidence of grade - and grade - agvhd were and % respectively. only patients developed cgvhd. cumulative incidence of trm and relapse incidence were % and % respectively. after a median follow-up of months, projected os at year is %. immunoreconstitution was fast and sustained with a median circulating cd + cells/μl (range - ) from day . the immune-reconstitution was polarized towards central memory (cd ra-cd l + cells . % ± . ), il- producing cells (il- + cells . % ± . ). we detected high levels of cd + cd + cd -foxp + t regulatory cells (up to % of circulating cd + t lymphocytes) starting from day . these cells were able to suppress in vitro proliferation of autologous effector cells demonstrating to be regulatory t cells. conclusions: rapamycin-mycophenolate-atg are effective as gvhd prevention in t-cell replete unmanipulated haploidentical peripheral sct and are associated with an early t-cell immunoreconstitution characterized by the in-vivo expansion of earlydifferentiated t cells and t-regs. further studies are warranted to gain insight on the role of rapamycin as platform for exploitation of t-regs in allogeneic hsct from mismatched donors. direct interaction of human nk cells with aspergillus fumigatus induces a th immune response and provokes signifi cant fungal killing but not via their usual cytotoxic pathways m. bouzani, m. ok, o. kurzai, h. einsele, j. loeffl er wurzburg university (wurzburg, de) objectives: invasive aspergillosis (ia), caused mainly by aspergillus fumigatus (af), is a highly devastating disease for immunosuppressed subjects. host's defence is principally confi ned to innate effector cells like alveolar macrophages, neutrophils and dendritic cells. in our study, we questioned the possible interaction of af with another potent component of the innate immunity, the natural killer (nk) cells. methods: human nk cells were isolated after magnetic depletion of the peripheral blood of volunteers and they were used after h priming with u/ml recombinant human interleukin , rhil . interferon gamma (ifn-g) and tumor necrosis factoralpha (tnf-a) regulation were assessed after nk-af coculture. fungal damage was investigated through plate killing assays. to investigate the infl uence of rhil on nk cells, plate killing experiments were performed with resting and primed nk cells. transwell permeable membranes, nk cell granule depletion (treatment with strontium chloride), surface expression of degranulation markers cd a/b and neutralization of nk death ligands (tnf-related apoptosis-inducing ligand [trail] and fasl) by blocking antibodies were used to evaluate the means of interaction. results: fungal germlings induced towards nk cells a th immune response with upregulation of ifn-g and tnf-a (p< . ). nk cells displayed strong fungicidal effect against germlings (p< . ), but they were inactive against conidia. priming with rhil (p< . ) and direct effector-pathogen contact (p< . ) were required for their interaction. the cytotoxic effect was not attributed neither to the release of perforingranzyme, nor to the engagement of nk cell death ligands. conclusion: human nk cells are stimulated in vitro by af, which triggers a th immune response and causes important fungal killing. this interaction requires priming of nk cells with rhil and conditions of direct contact between nk cells and fungus. interestingly, nk cells do not mediate their cytotoxic effect via perforin -granzyme pathway, neither through the engagement of death ligands, suggesting that another pathway is involved in nk cell -af interaction. our study attributes to nk cells anti-aspergillus properties, suggesting them as a future potential immunotherapeutic tool against ia. objective: to conduct a survey on indications, effi cacy and safety of growth hormone (gh) treatment in children and ado-lescents after haematopoietic stem cell transplantation (hsct) within the ebmt. methods: in this retrospective survey using a two step approach, we asked ( st questionnaire) for information on endocrine follow-up, the use of gh, reasons for not using gh, indications for gh treatment, number of patients treated with gh and diagnostic tests to diagnose gh defi ciency (ghd); and ( nd questionnaire, to follow) data on growth, diagnosis of ghd, interval between hsct and gh treatment, dose schedules, other endocrine defi ciencies. results: st questionnaire: centres replied until . . : endocrine follow-up (n = ) was performed by the hsct-clinic and an endocrinologist in %, the hsct-clinic in %, an endocrinologist in %, a specialist in another clinic in % and the hsct-and another clinic in %. % ( / ) centres treated patients after hsct with gh, whereas % ( / ) did not. of the centres reporting, / centres from italy used gh, whereas / did not. in contrast, / centres from france used gh whereas / did not. german centres reported the use in / centres. all centres from finland ( ), great britain ( ), spain ( ) and switzerland ( ) reported the use of gh. indications for gh treatment were growth failure in % ( / ), growth failure or ghd in % ( / ) and ghd only in % ( / ). reasons for not using gh were: no indication in % ( / ), risk of side effects in % ( / ) and lack of fi nancial support in % ( / ) . the numbers of patients treated in centres was less than patients in % ( / ), to patients in % ( / ), to patients in % ( / ) and more than patients in % ( / ). diagnostic tests for ghd used were the insulin-test (n = ), arginin-test (n = ), spontaneous gh-secretion (n = ), clonidin-test (n = ) and ghrh-test (n = ). conclusions: in the majority of centres the endocrine follow-up was performed by an endocrinologist. gh treatment was regularly used for the treatment of growth failure after hsct. about % of the centres used gh treatment for growth failure due to ghd only. in centres not using gh the main reason was that they saw no indication for gh treatment. only a minority did not use gh due to the risk of side effects. the insulin-test, arginintest and spontaneous gh-secretion were the major tests used for the diagnosis of ghd. a. rovo, m.t. van lint, m. aljurf, n. salooja, g. sucak, a. hunter, m within nonmalignant complication after hematopoietic stem cell transplantation (hsct) gonadal dysfunction with absence of sperm production leading to defi nitive infertility is a common long-term sequela. young age at hsct and longer interval time since hsct has been associated as a favourable factor for spermatogenesis recovery. the total body irradiation (tbi) used as part of the conditioning regimen plays a central role in posttransplantation infertility. we assessed in a retrospective multicenter study on a large cohort of male survivors, the probability of sperm recovery after hsct, and evaluated associated factors for fertility recovery. male patients in which at least one seminal fl uid analysis was performed after hsct being in complete remission and in whom the results were available were the target population. ninety centers reporting to the ebmt were asked to participate, accepted and so far centers contributed with patients. the median age at hsct and at time of st sperm fl uid analysis (sfa) was and ( - ) years respectively. the median time interval between hsct and sfa analysis was months . ( %) received an allogeneic hsct, ( %) had bone marrow as source of stem cells, ( %) received a myeloablative conditioning and ( %) received tbi (with a median doses of gy ( . - . ) as part of the conditioning. during the follow-up recipients ( %) had any grade of acute gvhd, ( %) any type of chronic gvhd (cgvhd); ( %) from them had ongoing cgvhd at sfa time. presence of at least one spermatozoa in sfa was considered as recovery of spermatogenesis. spermatozoa were detected in the semen in / ( %) patients; ( %) of them showed normozoospermia. in the univariate analysis following associated factors were signifi cant (table ) : younger age at hsct, longer time interval between hsct and sfa, conditioning without tbi, patients without ongoing cgvhd. in the multivariate analysis, absence of tbi (p< . ), longer time interval (p = . ), absence of ongoing cgvhd at sfa (p = . ) were signifi cant. in conclusion, this is the largest population of male survivors evaluating spermatogenesis after hsct. in this study we confi rm the established factors which infl uence recovery of spermatogenesis such as age (univariate analysis), time interval between hsct to sfa and tbi. here for the fi rst time evidence for graft versus testis effect can be demonstrated. introduction: development of leukemia or myelodysplasia derived from donor cells termed donor cell leukemia (dcl) is a rare but severe complication following allogeneic hematopoietic transplantation. the estimated incidence is low ranging from cases of dcl per , transplants (mostly myeloablative conditioning, retrospective analysis) to , per , transplants (non-myeloablative, single institution experience). although about cases have been reported in the literature, very little is known about pathogenesis and clinical management of donor cell leukemia. factors proposed to be involved in malignant transformation and development of dcl include immunoregulatory dysfunction, immunosuppression, host environment triggering malignant processes, replicative stress and epigenetic reprogramming as well as antigenic or viral stimulation. due to the increasing number of case reports throughout the last few years the late effects working party decided to analyze and evaluate the experiences with donor cell leukemia within the ebmt centers. methods: a fi rst, short questionnaire will be sent to all ebmt centers asking for observed, proven or suspected cases of dcl. centers reporting one or several cases will be asked to complete a second, more detailed questionnaire. the aim of the study is to identify and analyze a suffi cient number of cases to answer the following questions: how is the incidence of dcl evolving? is development of dcl associated with viral status before and viral reactivation/infection during as well as after transplantation? is there any infl uence of graft source and donor type (bone marrow vs. peripheral blood stem cells vs. cord blood and related vs. unrelated)? is the risk of malignant transformation of grafted cells increasing with donor age? is there any association with the form and extent of pretreatment (radiotherapy and chemotherapy) implicating a role of damaged microenvironment in the development of dcl? and fi nally, is dcl also observed as a very late complication of hematopoietic stem cell transplantation? conclusion: the answers to these questions will help to further characterize donor cell derived leukemia and provide new insights into leukemogenesis in general. a. crotta, a. tichelli, a. ruggeri, i. ionescu, a.l. herr, k. boudjedir, e. gluckman, v unrelated cord blood transplant (ucbt) is associated with a reduced incidence of chronic gvhd when compared to other sources of stem cells, however risk factors analysis for incidence is scarce in the literature. we retrospectively analyzed patients (pts), children (age≤ ) and adults, receiving fi rst single (n = ) or double ucbt (n = ) in ebmt centers, between and , for malignant and non-malignant diseases, who survived at least days from transplantation with neutrophils recovery and without relapse or autologous reconstitution. median age was years ( . , most common disease was acute leukemia ( %). % of units were hla-identical (antigen level for hla-a and b, allelic for drb ), while % and % had or - mismatches, respectively. median tnc infused was . x /kg. conditioning regimen was myeloablative (mac) in % of cases (tbi> gy in %), ric in %. busulphan-based conditioning was used in % and atg was added in % of cases. csa + steroid was the most common gvhd prophylaxis ( %); in children prednisone was used in % of pts, mycophenolate mofetil was used in combination in % of adults. median follow-up was months ( - ). incidence of cgvhd at y was ± % and ± % in children and adults respectively (p< . ). due to statistical difference between children and adults, risk factors analyses were performed separately. gvhd was extensive in % of children and in % of adults. out of pts who developed cgvhd, had previously agvhd ( out of children and out of adults). in univariate analysis in children, factors associated with increased incidence of gvhd were: age (> y), advanced status of disease at transplant (>cr ) and number of hla disparities; cgvhd was ± % and ± % (p = . ) in - / cord blood unit and - / respectively. in multivariate analysis only hla disparities (hr = . , p< . ) and status of disease at transplant (hr = . , p = . ) were associated with increased incidence of gvhd. in adults, higher incidence of cgvhd was associated with male sex, advanced disease at ucbt and use of mac regimen. in multivariate analysis only the status at transplant was independently associated with cgvhd (hr = . , p = . ). in conclusion, incidence of cgvhd was lower in children than in adults. status of disease at transplant was associated with increased incidence of gvhd in children and adults. number of hla disparities was associated with increased cgvhd only in children. in ms, progression free survival has been reported to range from to % at years after hsct. the most frequent conditioning regimen adopted in europe is the association of beam and atg, an intermediate-intensity scheme able to completely suppress mri activity for at least years in most of patients. mortality has dropped in the last years from a relevant , % to . %, due to a better transplant management and patients selection. in rapidly progressive ssc -years mortality is estimated to be - % [ ] . the most commonly used conditioning regimen in europe is cyclophosphamide ( mg/kg) and atg. in these patients, progression free survival after hsct has been showed to be above % at years. furthermore, the use of high intensity conditioning regimens does not seem to offer special advantages over lower intensity schedules for this disease. the distribution of transplants per year shows a drop of activity in sle and especially in infl ammatory arthritis, following the introduction of new biological agents after . the emergence of pharmacological resistance in long-term treatments may suggest a possible renewal of the activity also in this subset of patients. in chronic infl ammatory bowel diseases (ibd) an increasing activity has also been shown in the last years. hsct is able to provide a high rate of long-term, immunosuppression-free remissions in ad patients resistant to conventional therapies. appropriate selection of patients is crucial for providing the best risk-benefi t equipoise. evidence of effectiveness and careful assessment of toxicity is expected from ongoing and future prospective clinical trials. data reported to the ebmt registry at december european activity in multiple sclerosis r. martin ( ) , g. mancardi ( ) intensive immunosuppression followed by autologous transplantation of hematopoietic stem cells (ahsct) has been investigated as a possible strategy for the treatment of severe autoimmune disorders, including multiple sclerosis (ms) . it has been demonstrated that ahsct leads to the elimination of self reactive t cells and complete or almost complete renewal of the immune repertoire. immune system renewal has been demonstrated at the level of the t cell receptor repertoire in cd + and cd + t cells and in subsets of nk-and t cell population. according to the database of the european blood and transplantation group (ebmt) registry, more than ms cases have been treated with ahsct. the retrospective survey, carried out in , of all the patients recorded in the registry, with a median follow up of more than years, showed that improvement or stabilization of neurological conditions occurred in % of cases. transplant related mortality was very high ( , %) in the - period, but subsequently dropped to . % in the - years. the results appear to be particularly impressive in rapidly evolving severe ms cases unresponsive to conventional therapies. in these cases, defi ned also as malignant forms of ms, numerous european groups convincingly demonstrated the capacity of ahsct to suppress the progression of the disease, with an unexpected relevant improvement of the neurological conditions lasting for a long period of time. the establishment of ahsct as a rescue therapy in highly active ms cases unresponsive to other conventional therapies represents a very important step in ms clinical management, and the above mechanistic studies strongly support the rationale of this approach. despite these advances, the acceptance of ahsct as a rational treatment escalation remains low among neurologists, and this is probably one of the main reasons why the phase study (astims), comparing ahsct vs. mitoxantrone with a mri primary endpoint, had to be terminated due to recruitment diffi culties. based on this experience, a new phase iib/iii study is currently being designed. this study will compare ahsct with best available and approved therapy in the group of highly active relapsing-remitting ms patients, who failed fi rst-line therapy and one escalation step. the main goals of this trial will be to establish effi cacy in a controlled trial and to address important mechanistic questions. the status of the current discussion toward the trial design as well as strategies to recruit interested centers with ms-specialized neurologists and experienced transplant physicians at one location and potential mechanisms of funding will be presented. systemic sclerosis is a potentially life-threatening chronic autoimmune disease characterized by skin thickening, vasculopathy, and visceral involvement, mainly of lungs, gut, heart and kidneys. the clinical manifestations are diverse and refl ect a spectrum of subsets, ranging from limited to diffuse disease. diffuse cutaneous systemic sclerosis (dcssc) generally runs a more aggressive disease course, requiring intensive immunosuppressive therapy. haematopoietic stem cell transplantation (hsct) has resulted in long-term improvements of skin thickening, stabilization of organ involvement, but at the expense of mainly cardiopulmonary toxicity which has resulted in death in - % of cases according to a recent analysis by the ebmt working party autoimmune diseases. to assess whether the benefi ts outweigh the risks, a prospective randomized controlled trial is nearing completion comparing hsct with iv pulse cyclophosphamide in which dcssc patients have been enrolled in centres. the primary endpoint is event-free survival, defi ned as survival until death or development of major organ failure during years follow-up. formal analyses of safety and effi cacy will be done in . interim safety analyses have lead to changes in the transplant protocol and eligibility criteria to also target patients with very early disease. the rationale of the trial is that effective intervention in very early disease is needed to induce long-term remissions. all patients are followed-up for at least years. in anticipation of the results from the astis trial, a new smallscale clinical trial will be done to comprehensively investigate the mechanism of action of hsct in ssc focusing on its effects on biomarkers of fi brosis, autoimmunity, infl ammation and vasculopathy. it is hoped that the results of the astis trial and those of the north american scot trial will provide solid data to decide on the design of a subsequent randomized trial. if astis and scot indeed prove that hsct is superior over conventional chemotherapy, then one option would be to modify the trans-plant regimen to reduce the risk of cardiopulmonary toxicity and/or enhance the intensity of the control arm. crohn's disease (cd) is characterized by chronic infl ammation in segments of the digestive tract leading to tissue damages. uncontrolled infl ammation is associated with excessive immune responses towards the microbiota. progress has been recently made in the management of cd, with increased use of immunosuppression and biologic therapies. early optimized use of these treatments in patients with poor prognosis may provide a satisfactory control of the disease in the majority of patients. however, a fraction of cd patients experiences severe disease refractory to medical management, which may result in progressive tissue damages, need for surgery and chronic disability. the fi rst evidence of effectiveness of hematopoietic stem cell transplantation (hsct) was reported with the observation that patients with cd, who underwent allogenic or autologous hsct for a haematological or solid malignancy, experienced long-lasting remission of their ibd. although few cases regarding autologous hsct in concomitant cd and malignant diseases have been reported, collected data have shown similar results to allogeneic transplantation in terms of remission. beyond case reports, autologous hsct as primary treatment for cd has been investigated in two single-centre phase ii studies. despite the low number of patients and their limited follow-up, autologous hsct was shown to have the capacity to induce complete clinical and endoscopic remission, with impressive results. the astic trial, designed to assess its effi cacy and tolerability, is ongoing. included patients undergo mobilization of stem cells, and are then randomized to transplantation after one month or after one year. autologous hsct may represent a therapeutic option in cd patients with refractory disease, and could change the natural history of the disease, inducing in some cases long term remission. haematopoietic cell transplantation for autoimmune diseases: ebmt/cibmtr collaboration m. pasquini (milwaukee, us), r. saccardi (florence, it) high dose immunosuppressive therapy with autologous haematopoietic cell rescue has the potential to halt the autoimmune process and result in a medication-free period. since , more than patients in europe and patients in north and south america who underwent transplantation for autoimmune disease (aid) have been registered in the ebmt and cibmtr databases. despite this activity, questions related to the timing of transplantation and what aid benefi ts more from transplantation remain unanswered. the development of a collaborative strategy has the potential to answer some of these questions and assist the development of prospective studies that can advance the fi eld. following this strategy the cibmtr and ebmt autoimmune disease committees joined forces in to expand the study portfolio and establish an infrastructure to facilitate future studies. the most common aid indication for transplantation is multiple sclerosis (ms). data collection from both registries was harmonized into a single disease-specifi c form developed by a group of investigators affi liated to both committees. collaborative project to focus on long term outcomes after transplant for ms was developed, especially to determine factors associated with prolonged disease-free intervals. two in person discussions on the topic leveraged the development of an international phase iii clinical trial with the objective to compare transplant with best medical treatment in a population with rapidly progressive ms. the next collaborative initiative is to focus on the second most common indication, systemic sclerosis (ssc). we plan to start harmonizing disease-specifi c forms and a collaborative study focused in this disease. lastly, as the majority of patients present to transplantation with advanced aid, studying patients with coexistent aid who present for allogeneic transplantation, represent an opportunity to understand the graft-versus-autoimmunity effect. thus, the cibmtr is planning to identify these patients at time of registration and subsequently enrolled them on a prospect cohort study to evaluate the impact of graft-versus-host disease on aid. outcomes database on transplant for aid are currently underutilized due to the heteroneity of diseases, scarcity of comprehensive level disease-specifi c information and loose integration with non transplant specialist. bringing the ebmt and cibmtr together in the last years, helped engage investigators, including neurologists and rheumatologist to develop collaborative studies. an update on autologous stem cell transplantation in severe systemic lupus erythematosus and in early diabetes type d. farge, r. cervera, d. jayne, a. voskuyl, j.m. van laar, a. doria, m. mosca, d. boumpas, r. saccardi, e. snarski, j.f objectives: to present the experimental and clinical rational for the ongoing ebmt or nih trials for ahsct in severe sle and in early diabetes type i. background: since , autologous hematopoietic stem cell transplantation (ahsct) has been used successfully in severe systemic lupus erythematosus (sle) and early insulin type i dependent diabetes (t d) to induce durable remission. in sle bilag a, the nih or eurolupus cyclophosphamide (cy) protocols and mycophenolate mofetil (mmf) for induction are associated with % failure, % relapse and - % death at yrs. among sle out of ebmt pts with fi rst ahsct in , years pfs was % ( % ci: %- %) with % ( % ci: %- %) overall survival (farge d haemetologica ). higher remission rates and also some relapses, were shown in the north american experience (burt r, jama, ) with the possibility of resetting the autoimmune response and inducing tolerance in sle after hsct and then in new onset type i diabetes (voltarelli and burt et al, jama, ) . in susceptible strains of mice, allogeneic hsct prevents insulitis and development of type dm. in , follow-up of the ahsct for t d with in brasil, showed that / remained insulin free (and normal hba c) with longest insulin free follow-up beyond yrs and no correlation with pre-hsct c-peptide, but ahsct must be performed within months of diagnosis. eight patients with t d (< weeks from diagnosis, c-peptide positive, anti gad -antibodies positive) treated in poland using mg/cy conditioning and ahsct plus antithymocyte globulin (atg genzyme) had no major complications after ( - ) mths median follow-up and / pts became independent from exogenous insulin within ( + -+ ) days after ahsct. acarbose may have positive effects on the duration of remission of t d after ahct. planned studies: these were the basis for ) the multicenter, ebmt approved astil phase ii study, to analyse the effi cacy of ahsct followed by mmf as maintenance for severe sle yrs since diagnosis with sustained or relapsed active bilag a sle after at least months of best standard local therapy among patients. after cy g/m mobilisation, unselected ahsct will use cy ( mg/kg body wt in daily doses) plus rabbit atg and maintenance by mmf ( g/day). the primary effi cacy endpoint will be the proportion of patients alive who achieve clinical success at months and maintenance of this status until months after inclusion; ) the nih approved phase i/ii study of ahsct in newly-diagnosed t d pts within months of diagnosis, and a positive antibody to an islet cell autoantigen and fasting c-peptide > . nmol/l. to compare cy g/ m mobilization, unselected ahsct using cy ( mg/kg body wt in daily introduction: graft versus leukemia (gvl) effect of hematopoietic stem cell transplantation (hsct) is mostly based on donor t cell-mediated alloreactivity. this is particularly relevant in the context of hsct from family haploidentical and matched unrelated donors (mud), in which mismatched hla molecules are potent targets for donor t cells. still, upon in vivo selective pressure by donor t cells, leukemia can undergo genomic rearrangements which result in loss of the patient-specifi c hla, a mechanism which our group recently demonstrated to be frequently responsible for leukemia relapse after haploidentical hsct (vago et al., nejm, ). methods: consecutive patients who underwent a partially hla-matched transplantation for myeloid malignancies from to present, were included in our analysis. for patients the donor was family haploidentical, and for was unrelated and mismatched for a median of / hla alleles. all patients received donor t cells as part of the hsct protocol. post-transplantation follow-up comprised monthly bone marrow examination, with short tandem repeat (str) chimerism analysis and hla typing performed in parallel. in cases of relapse with suspected loss of the mismatched hla, str chimerism and hla typing were performed also on purifi ed leukemic blasts. results: disease relapse occurred in and patients after haploidentical and mud transplantation, respectively. after haploidentical transplantation, / relapses ( . %) were due to mutated leukemic blasts which had lost the patientspecifi c hla haplotype. interestingly, of the relapses after mud transplantation occurred through the same mechanism, in a patient who had received two subsequent transplants from hla-c-mismatched donors. upon detection of the mutated leukemic blasts, of these patients were enrolled to receive a subsequent hsct from a different donor, mismatched for the remaining hla haplotype. conclusions: our data consolidate the clinical relevance of this escape mechanism from the gvl effect mediated by alloreactive donor t cells. screening for these mutants should be encouraged in patients who relapse after partially hla-mismatched hsct, to guide therapeutic strategies, avoid predictably ineffi cacious donor t cell add-backs, and quickly enroll the patients to a salvage transplant. to improve this approach, novel diagnostic and therapeutic tools are warranted, to provide earlier molecular detection and specifi c targeting of these mutants. impact of allogeneic stem cell transplantation on prognosis of patients with high-risk aml: results of the aml shg and / trials k. wagner, g. heil, m. zucknick, d. hoelzer, o.g. ottmann, h. martin, m. lübbert, j. finke, w. heit, w. fiedler, l. kanz, g. schlimok, h. kirchner, a. raghavachar, w. brugger, a. ganser, j objective: patients with high risk acute myeloid leukemia (aml) have a dismal prognosis after chemotherapy. allogeneic stem cell transplantation (allosct) in fi rst complete remission (cr) is widely recommended for these patients but its impact on outcome is uncertain. methods: in the prospective multicenter trials aml and / , we treated adult patients with aml (except apl) up to years. all patients received intensive double induction and early consolidation chemotherapy. high risk patients were defi ned by either bad response to induction i (persistent bone marrow blasts on day after start of therapy) or high risk karyotype (all karyotypes other than normal or cbf-leukemias). all high risk patients with a matched related donor (mrd) were scheduled for an allosct as late consolidation. patients without a family donor in the trial should receive an autologous stem cell transplantation, whereas in the / trial an allosct from matched unrelated donors (mud) was recommended. results: median follow up of the patients is months. of the patients fulfi lled the high risk criteria. median overall survival (os) of these patients is months and the six-year os is %. in multivariate analysis, a complex or monosomal karyotype (hr . ; % ci . - . ) and age above the median of years (hr . ; % ci . - . ) were identifi ed as adverse prognostic factors for os. of the high risk patients ( . %) achieved a cr. median relapse-free survival (rfs) is . months and six-year rfs is %. leukocytes above the median (hr . ; % ci . - . ) and a complex/ monosomal karyotype (hr . ; % ci . - . ) were identifi ed as independent prognostic factors for rfs. of the patients who achieved cr, received an allosct from mrd (n = ) or mud (n = ) in fi rst cr. median os of the transplanted patients is not reached and six-year os is %. median rfs after allosct is months and six-year rfs is %. outcome did not differ between mrd and mud allosct. the impact of allosct on prognosis was analyzed in a multivariate cox-model with allosct included as a time-dependent covariable. in this analysis, allosct in fi rst cr signifi cantly improved os (hr . ; % ci . - . ) and rfs (hr . ; % ci . - . ). conclusion: allosct from matched related or unrelated donors in fi rst cr improves overall and relapse-free survival of patients with high risk aml. cord blood transplantation for adults with acute lymphoblastic leukaemia -a survey of outcomes conducted by eurocord and the acute leukaemia working party of the ebmt d. purtill ( ) cord blood transplantation (cbt) is considered an option for adults with acute lymphoblastic leukaemia (all) for whom stem cell transplantation is indicated. however, little information is available regarding outcomes of this procedure in this group of patients. we conducted a retrospective review of the eurocord database in order to describe outcomes of adults treated with single or double cbt for all at ebmt centres from until . two-hundred and thirty six patients with a median age of years ( - years) were included. most were transplanted in fi rst complete remission (cr ) (n = ) or cr (n = ). median white cell count at diagnosis was . x /l ( . - ) and % (n = ) of patients in cr had t( ; ) or t ( ; ). double cord blood transplantation was performed for patients ( %). overall median total nucleated cell dose at freezing was . x /kg ( . - . ) and it was . ( . - . ) and . x /kg ( . - . ) respectively for double and single transplants. most patients received cord units with one (n = ) or two (n = ) hla disparities. overall cumulative incidence (ci) of neutrophil recovery was ± % and acute graft-vs-host disease was ± %. transplant-related mortality (trm) was ± % and ± % for patients in cr and cr respectively, and ± % for the remaining patients with more advanced disease (cr , relapsed or refractory disease). ci of relapse were ± %, ± % and ± % respectively for cr , cr and advanced disease. at the median follow-up of years, leukaemia-free survival (lfs) was ± % for the whole population and ± % for cr , ± % for cr and ± % for advanced disease. lfs for patients in cr with t( ; ) or t( ; ) was ± % and for double cbt it was ± %. on multivariate analysis, being in cr or cr at transplant was the only factor associated with improved lfs (hr . , %ci . - . , p< . ). when this group was analysed separately, the use of a reduced intensity conditioning (ric) regimen was associated with improved lfs ( ± % vs. ± %, hr = . , %ci . - . , p = . ). in particular, the patients who received the combination of cyclophosphamide, fl udarabine and tbi ( - gy) had an lfs of ± %. cord blood is an alternative source of stem cells for allogeneic transplantation for adults with high risk all. results with ric and double cbt are encouraging and should be further investigated in a larger series of patients. superior long-term survival with a high rate of allogeneic stem cell transplantation in aml (non-apl) patients below years of age g. juliusson ( ) introdution: allogeneic stem cell transplantation (allosct) prevents relapse in aml, but the indication in intermediate risk is unclear and depends on the transplant-related mortality. the swedish acute leukemia registry (blood ; : ) includes patients (pts) with acute leukemia diagnosed - covering % of all cases in sweden, with million inhabitants. using this population-based registry, we evaluated the sct rates in aml, and the long-term outcome. allo-sct rates in different swedish regions were also evaluated. transplantation data was individually validated in , and survival was updated through the national population registry. patients and results: of adult aml (non-apl) pts < yrs (median yrs), ( %) had an allosct ( % with secondary aml) in cr (n = ; %), or at later stages (n = ; %), with decreasing rates by age ( figure ). donors were unrelated in %. median follow-up was . yrs, and ( %) of allografted pts were alive. treatment-related mortality (trm, i.e., death in cr), and aml deaths according to age, donor, and status at allosct is shown in figure . estimated -yr and -yr os was % and %, respectively. these results could be compared to recently published large aml studies from ecog, eortc-gimema, uk mrc and gamlcg, with allosct rates of - % and -yr os of - %. the health care region se, which had the highest allosct rate ( % of aml pts < yrs) had a -yr os of % for the total aml population, as compared to the other swedish regions with a mean allosct rate of % and a -yr os of % (log rank, p = . ). the difference in allosct rate was mainly due to more allografts among older pts: in the age cohort - yrs, % of cr pts underwent allosct yrs in region se as compared to % in the rest of sweden, associated with a more pronounced survival benefi t (p = . ). conclusions: this is a real world population-based study on allosct rates, treatment-related mortality and long-term survival in aml pts. our results indicate that allosct is performed in sweden with relatively low total non-relapse mortality, and with a moderate relapse rate. the association found between a high allosct rate in pts - yrs and a better survival is intriguing, is suggestive of a positive impact of a more frequent transplantation practice in this age cohort, and supports the need for prospective studies. role of the graft-versus-leukaemia effect after reducedintensity conditioning allogeneic stem cell transplantation as treatment for acute myeloid leukaemia: a survey from the acute leukaemia working party of the ebmt f. baron, m. labopin, m. mohty, n. basara, d. niederwieser, n. milpied, j.j. cornelissen, c. malm, l. vindelov, d. blaise, j.j.w.m. janssen, e. petersen, g. socie, v previous studies have observed a lower risk of relapse in patients (pts) who experienced chronic gvhd after ric allo-sct versus in those who did not. the objective of the current study was to further investigate the association between chronic gvhd and relapse in a large cohort of pts given ric allo-sct as treatment for aml. data from aml pts in fi rst or second cr transplanted between and following a ric regimen at ebmt affi liated centers were analyzed. patients were given pbsc from hla-identical sibling (mrd, n = ), or from hla-matched unrelated donors (mud, n = ). median pt age at transplantation was yrs in pts given grafts from mrd, versus yrs in those given grafts from mud. the impact of chronic gvhd on relapse risk, non-relapse mortality (nrm) and leukemia-free survival (lfs) was assessed by time-dependent multivariate cox models and in a landmark analysis. three-yr incidences of relapse, nrm and lfs were ± %, ± %, and ± %, respectively, while -yr incidence of chronic gvhd was ± %. in a landmark analysis at months after allo-sct, -year relapse rates were ± % versus ± % for patients with or without chronic gvhd (p = . ), respectively. in multivariate cox models, cr versus cr (p = . ), pt age > yrs (p = . ), alemtuzumab use in the ric (p = . ), tbi-based ric (p = . ), high-risk cytogenetics (p = . ), and absence of chronic gvhd (p = . ) were each associated with higher risk of relapse. factors associated with high nrm were mud versus mrd (p = . ), grade ii-iv acute gvhd (p<. ), and chronic gvhd (p = . ). factors associated with lower lfs were cr versus cr (p = . ), pt age > yrs (p = . ), alemtuzumab use in the ric (p = . ), and high-risk cytogenetics (p = . ). in conclusion, in this cohort of aml patients transplanted in remission, chronic gvhd was associated with a lower risk of relapse while profound in-vivo t cell depletion with alemtuzumab was associated with higher relapse rate suggesting that gvl effects play a role in preventing aml relapse in patients given ric allo-sct. in spite of the presence of gvl effect, chronic chvd was associated with higher nrm and therefore there was no net impact on lfs. strategies to decrease nrm related to chronic gvhd should be further investigated in order to keep the benefi ts from gvl effect and improve lfs. finally, the impact of chronic gvhd duration and severity on lfs need to be studied. allogeneic stem cell transplantation in acute myeloid leukaemia with normal karyotype: a risk factor analysis in patients, based on molecular markers and stage at transplantation t. pfeiffer ( ) , m. schleuning ( ) cytogenetically normal acute myeloid leukaemia (cn-aml) represents a heterogenous disease, that can be subdivided by molecular analysis. only limited data is available on sct in different molecular subgroups, particularly in advanced stage. therefore, we retrospectively analyzed data on pts. with cn-aml, who uniformly had received the flamsa-ric regimen for sct in european centres. pts. suffered from de novo aml ( %), saml/mds ( %), and taml ( %). median age was y ( - ). donors were matched or mm family, and matched or mm unrelated donors in %, %, % and %, respectively. sct was performed in untreated disease ( %), primary induction failure (pif, %), st complete remission (cr , %), and >cr ( %). median follow-up of survivors was mo. overall survival (os) and leukaemia-free-survival (lfs) at y from sct was % and %. the stage at sct was the most important factor (p = . for os, <. for lfs). results were promising after sct in cr ( y os/lfs %), and pif ( y os/lfs %), but were inferior after sct in untreated disease ( y os/lfs %), or >cr ( y os %, lfs %). information on molecular markers was available in pts. ( %). as suggested (schlenk, nejm ) , analysis was based on subgroups: pts. with isolated npm mutation (npm mut), and pts. with other genotypes (flt -itd, n = ; or wildtype flt /wildtype nmp [flt wt/npm wt], n = ). pts. with npm mut had a y os/lfs of / % with identical outcome, when transplanted in pif, cr , or >cr . pts. with other genotypes showed an os/lfs of %/ % at y and of %/ % at y, without differences among pts. with flt-itd and flt wt/npm wt. however, in this subgroup, outcome was highly dependent on the stage at sct, with excellent results after sct in cr ( y os/lfs: %) or pif ( y os/lfs: %/ %), but inferior outcome after sct >cr ( y os/lfs %/ %; p = . for os, . for lfs). conclusion: allosct following flamsa-ric produces excellent survival in pts. with cn-aml, particularly when performed in cr . encouraging results in pif support an early sct, regardless of molecular subgroup, when cr is not reached after double induction. in npm mut, sct in advanced disease achieved identical results as in early stage, supporting the delay of sct until relapse has occurred. in contrast, pts. with flt -itd or flt wt/npm wt achieved signifi cantly worse results when transplanted >cr , arguing in favour of sct in cr for this molecular subgroup. comparison of outcomes after allogeneic sct for adult patients with aml in remission using either i.v. busulfan plus cyclophosphamide or tbi plus cy in the myeloablative conditioning regimen: an-alwp-ebmt survey a. nagler ( ) , m. labopin ( ) , a. shimoni ( ) tbi/cy and oral bu/cy are the most common myeloablative regimens for adults with aml with comparable survival and relapse probabilities. intravenous (i.v.) bu in contrast to oral bu has more predictable pharmacokinetics and a more favorable toxicity profi le. outcomes with i.v. bu/cy, thus, may be superior to those achieved with tbi/cy. in order to address these issues, the alwp of the ebmt performed a survey comparing i.v. bu/cy to tbi/cy as conditioning regimen for adult pts. with aml undergoing allosct. overall, allosct were analyzed: bu/cy - , tbi/cy - . median age was (range, - ) and (range, - ) years and median transplant year was ( - ) and ( - ) for the bu/cy and tbi/cy groups, respectively (p< . ). disease status at allosct was cr - % vs. % and cr - % vs. % for the bu/cy and tbi/cy groups, respectively (ns). wbc count at diagnosis was x /l in both groups. cytogenetic: good - % and %, intermediate - % and % and poor risk - % and %, respectively (na - %- % of pts). % and % of both groups underwent allosct from sibling donors, while % and % from mud, respectively. follow up was ( . - ) and mo. for both groups, respectively. % and % of the bu/cy and tbi/cy groups received pbsc, while % and % received bm grafts, respectively (p< . ). engraftment was similar in the bu/cy vs. the tbi/cy groups, % and %, respectively. cumulative incidence of nrm at y was + % and ± %, respectively (p = . ). acute gvhd both >ii-iv and iii-iv were signifi cantly lower in the bu/cy vs. the tbi/ cy groups: % and %and . % and . %, respectively (p< . ). while, vod was signifi cantly higher in the bu/cy vs. the tbi/cy groups, . % vs. . %, respectively (p< . ). two year relapse incidence was comparable, + % and + % for bu/cy vs. tbi/cy, respectively (p = . ). lfs was comparable, as well, + % and + %, respectively (p = . ). in multivariate analysis poor prognostic factors for lfs were: poor cytogenetics (p≤ - ), disease status (cr vs. cr ) (p≤ - ), age > y (p = . ), mud vs. sib. donor (p = . ) and pb vs. bm grafts (p = . ). in conclusion, in this retrospective ebmt survey, outcomes of allosct including engraftment, trm, rr and lfs was comparable, while acute gvhd probability was signifi cantly lower and vod probability signifi cantly higher in adult pts. with aml in cr using myeloablative i.v bu/cy vs. tbi/cy conditioning regiments, respectively. higher incidence of relapse with higher doses of cd + cells from leukapheresis products infused in adults with acute myelocytic leukaemia autografted during the fi rst remission n.c. gorin, m.m. labopin, d. blaise, f. witz, t. de witte, g. meloni, m. attal, d. carreras, v. rocha on behalf of the alwp purpose: the cell source for autologous stem cell transplantation has shifted from bone marrow (bm) to peripheral blood (pb). for patients with aml in cr , we previously showed that the risk of relapse was greater with pb than bm and a poorer outcome was associated with a shorter interval from cr to pb transplantation (≤ days) (jco in press). leukemic and normal progenitors bear the cd + antigen and can be mobilized together; we questioned whether there was a relation linking the doses of cd + cells infused to the outcome. methods: out of patients autografted with pb more than days post cr and reported to the ebmt registry using medb form, the dose of cd + cells infused was available in . results: the cd + cell doses were categorized by percentiles to divide the whole group into fi ve categories containing the same number of patients. we identifi ed the fi fth percentile (> . × /kg) as the cut off point for relapse incidence (ri) and leukemia-free survival (lfs). patients receiving the highest dose had a higher ri ( ± %, vs. ± %; p = . ) and a lower lfs ( ± % vs. ± %; p = . ). in a multivariate analysis adjusted for differences, ri was higher in patients receiving the highest cd + cell dose ((hazard ratio [hr], . ; % ci, . - . ; p = . ).and the lfs was worse (hr, . ; % ci, . - . ; p = . ). conclusion: for patients with aml in cr autografted with pb, risk of relapse is greater and lfs is lower in those receiving the highest doses of cd + cells. oral session : early side effects o dna transfer from donor to host cells as a mechanism for epithelial chimerism and genomic alterations after allogeneic haematopoietic cell transplantation m. waterhouse ( ) , m. themeli ( ) , h. bertz ( ) , n. zoumbos ( ) , j. finke ( ) , a. spyridonidis ( ) ( )albert ludwigs university of freiburg (freiburg, de) ; ( )university hospital patras (patras, gr) research in the fi eld of allogeneic hematopoietic cell transplantation (allo-hct) revealed hidden consequences of the co-existence of two genetically distinct populations in the transplant recipient. first, epithelial cells with donor-derived genotype emerge and second, epithelial tissues of the host acquire genomic alterations. we asked whether horizontal transfer of donor-dna to host epithelium is operating in (and linking) both phenomena. buccal samples were obtained from allotransplanted patients and analyzed with microsatellite markers for the presence of donor-dna and microsatellite instability (msi). the presence of graft-derived hematopoietic cells in the samples was excluded by immunocytochemistry. the results were associated with clinical data. genomic instability induction and dna transfer in epithelial cells by allogeneic lymphocytes was assessed in vitro. we found a high contribution of donor-dna (mean . %) in buccal samples in out of evaluable patients. in addition, % of the samples were positive for msi (msi + ). the extent of donor-dna was signifi cantly correlated with the occurrence of genomic alterations (p< . ). the age of the patient and a female-to-male transplantation were signifi cantly correlated with msi. there was a trend for increasing risk of msi for patients who experienced severe graft-versushost disease. during follow up secondary malignancy was diagnosed in patients ( %) who exhibited msi but only in ( %) with no msi. by applying a time-dependent statistical analysis we found that the probability of secondary tumor development was signifi cant higher in the msi + as compared to the msipatients (p = . , hazard ratio . , % . in an in vitro model, we demonstrated that apoptotic lymphocytes may not only induce genomic alterations but also transfer dna through a phagocytotic process to co-cultured epithelial cells. the ingested lymphocytic dna was also incorporated into the nucleus and integrated into the genome of the recipient epithelial cells, resulting in the creation of hybrid chromosomes. our results indicate that continuous charge of the transplant recipient with apoptotic donor-dna and its illegitimate integration into host epithelium may come in light as epithelial cells with donorderived genome or genomic instability events and may provide a new model for elucidating protean clinical manifestations after allo-hct, such as secondary malignancy. the haematopoietic cell transplantation-specifi c co-morbidity index and non-relapse mortality after reduced intensity conditioning: fi nal analysis from the alwp of ebmt in aml patients in fi rst complete remission m. mohty, m. labopin, n. basara, j.j. cornelissen, r. tabrizi, c. malm, j. perez-simon, a. nagler, n. kröger, b. rio, r. martino, m. eder, k. bilger, d. bunjes, g. socié, d. blaise, e. polge, v the current study was designed to test the performance of the adapted charlson comorbidities index (ci; so-called "sorror index") and its association with outcomes among a cohort of patients (i) aged > y.; (ii) diagnosed with a single disease entity, aml in cr , and (iii) who underwent a ric allo-sct reported to the ebmt registry between and . in this series, the median year of allo-sct was , and the median age was y. (range, - ) . % of patients needed more than one induction course to achieve cr . a fl udarabinebased ric regimen was used in % of patients, while % of patients received low-dose tbi as part of their ric, and % received other non-specifi ed ric regimens. % of the patients received allo-sct from an hla-matched sibling donor. based on score calculated with hazard ratios (hr) estimated on the population studied, patients ( %) had a ci score of , patients ( %) had a score of , and ( %) had a ci score of . the remaining patients ( %) had ci scores of or more. in this cohort, years overall and leukemia-free survival rates were ± % and ± % and the years relapse and non-relapse mortality (nrm) cumulative incidences were ± %, and ± % respectively. the years nrm incidences according to comorbidities score , , and + were ± %, ± %, ± % and ± %, respectively. in multivariate models (adjusted for recipient age, donor type, use of tbi or not, and cytogenetics risk group) comorbidities such as moderate active liver disease, obesity, prior history of renal dysfunction, and prior history of severe liver disease were associated with the highest hrs for years nrm (varying from . to . ) and years cumulative incidences of nrm varying from % to %, whereas previous solid tumor, diabetes, rheumatologic abnormalities, moderate pulmonary diseases, cardiac abnormalities (other than arrhythmia and valve disease) were associated with the lowest hrs (varying from . to . ) with years cumulative incidences of nrm varying from to %. results from this large study performed in a single disease entity and homogeneous allo-sct setting, suggest that the hematopoietic cell transplantation-specifi c ci is a simple, informative and useful tool for capturing pre-transplant comorbidities, and for predicting nrm after ric allo-sct for aml in cr in patients aged > y. such index may be used for clinical trials and patient counselling before ric allo-sct. fibrin glue directly applied on damaged bladder mucosa during cystoscopy is highly effective to treat severe, refractory, haemorrhagic cystitis after allogeneic transplant m.c. tirindelli ( ) ( ) background: hemorrhagic cystitis (hc) occurring after hematopoietic stem cell transplant (hsct) signifi cantly affects quality life of patients, prolongs hospitalization and in some cases can become a life-threatening complication. its management has not been established. fibrin glue (fg) is a hemostatic agent derived from human plasma with proven effi cacy in repairing damaged tissues. study design and methods: this study included patients who met the following criteria: grade ≥ hc not responding to hyperhydration, bladder irrigation, antiviral treatments and transfusion support. fg was obtained using vivostat system, an automatic method for processing and applying fg. during conventional cystoscopy and maintaining bladder distension by a co insuffl ator, fg was accurately sprayed through a specifi c applicator on bleeding mucosa. fg polymerized on contact and set over several days. the response to the treatment was defi ned complete (cr) for disappearance of hematuria, partial (pr) for at least one grade regression of hc and no response (nr). results: from jan to oct , patients undergoing an autologous (n = ) or allogeneic (n = ) hsct were registered at the rtn. no autologous hsct recipients developed hc of severe grade, whereas of patients ( %) undergoing an allogeneic hsct met the criteria to enter the study. these patients ( m, f) with a median age of . years (range, - ) had been submitted to a hsct from hla identical sib. (n = ), unrelated cb (n = ), mud (n = ) or related haploidentical donor (n = ) for different hematological malignancies. all patients, deeply immunosuppressed with positive bkv viruria > x copies/ml, developed a very severe hc, refractory to all current treatments including antiviral therapy. at time of fg application, hc persisted for a median of days (range, - ) and was grade and in and patients, respectively. the number of fg applications was in patients, in and in patient for a median of ml (range, . - . ) of glue. the treatment was successful in out of patients ( %). all patients with grade hc responded and the response was complete in ( %) and partial in ( %), while of the patients with grade hc: achieved cr, pr and nr. no patient died of hc. conclusions: fg therapy is a feasible, safe, easy repeatable, not invasive, small time consuming, lightly expensive and highly effective procedure in treating severe, refractory posttransplant hc. glutamine-enriched intravenous total parenteral nutrition doesn't improve mucositis and clinical outcome after stem cell transplantation for childhood malignancies. a prospective double-blind controlled study on behalf of aieop group c. uderzo ( ) , e. marrocco ( ) , p. rebora ( ) , f. cichello ( ) , m. bonetti ( ) , s. cesaro ( ) , s. varotto ( ) , p. verlato ( ) introduction: high dose chemo-radiotherapy followed by stem cell transplantation (sct) constitutes the principal cause of post-transplant severe mucositis and related complications. intravenous glutamine-enriched solution (ges) has been advocated as one of the support treatment capable to improve marked body protein wasting, oxidative stress which result in a mucosal damage, often "primum movens" of infectious diseases beginning from g.i.tract damage. patients and methods: patients ( males, median age at sct . years) with haematological malignancies have been treated from june to june with high dose chemotherapy and/or tbi followed by allogeneic sct ( match related, match unrelated, mismatch.patients were randomly and double-blinded assigned to undergo either for total parenteral nutrition (tpn) or for ges tpn at the dose of , g/kg/day from day of sct until the end of tpn. the principal endpoint of the study was the mucositis assessment according to who criteria.the evaluation of the clinical,haematological and laboratory parameters served to perform the evaluation of secondary end-points.statistical analysis was set up to demonstrate a % difference in terms of mucositis incidence and severity with a power of % (alfa = . ; two-sided tests). results: both study groups were comparable for age, gender, diagnosis and type of sct. the ges patients ( ) were clinically similar to the controls at the entry; however, they didn't experienced a reduction of rate and grade of mucositis, with an odd ratio of . ( . - . ) adjusted for the type of sct. neither the type of analgesic treatment nor the duration of opiod treatment, were statistically different. days of both different tpn, length of hospital stay, trm at day + ,acute or chronic gvhd, incidence of severe infectious diseases, immunological recovery, progression of malignancies were similar in both groups. nutritional status at the beginning and at the end of both types of tpn didn't show any difference at univariate analysis. no toxicity of ges was observed. conclusions: the current study is one of the few designed to demonstrate the utility of ges on mucositis in the setting of children undergoing sct for malignancies. ges failed to infl uence incidence and severity of mucositis after sct and didn't offer any advantage on clinical outcome, as claimed by other retrospective studies often performed in adult transplanted patients. clinical and genetic risk assessment for overall survival in haematopoietic stem cell transplantation a. dickinson ( ) objectives: non-hla gene polymorphisms and clinical risk factors impact on outcome after hsct. age, stage of the disease, time from diagnosis to transplant, histocompatibility and donor and patient gender combination are key transplant risk factors for hsct. we have assessed the impact of non hla gene polymorphisms on the ebmt risk score for overall survival in a eurobank cohort of hla identical sibling and matched unrelated donor (mud) transplants. methods: hsct patients with acute leukaemia (aml + all (al)) (n = ), cml (n = ) plasma cell neoplasia (n = ) or lymphoma (n = ) and their donors from transplant centres were genotyped for non-hla polymorphisms within the tumour necrosis factor receptor ii (tnfrsf b), estrogen receptor (esr ), vitamin d receptor (vdr), interleukin (il) (il ), il- receptor antagonist (ilrn), interferon gamma (ifng) and il (il ) genes. genetic factors were assessed using the log rank test (p value < . ). candidate factors were included in addition to the ebmt risk score in a stepwise cox regression procedure to select fi nal genes. predictive value of the model for overall survival was assessed through the concordance (c) index and prediction error curves. results: in the whole cohort the presence of allele c in the donor il- genotype (snp il - ) was associated with lower survival time (especially in the cml sub group p = . ) and improved the prediction of the ebmt risk score. in the al subgroup the absence of patient il (any t) and presence of il rn (any c) (high risk group) were associated with lower survival time (compared to the remaining patients (low risk group)), and taken together also improved the predictive value of the ebmt risk score. figures a and b show the lower probability of survival (and increased trm) in the high risk versus low risk group. conclusions: this study confi rms the importance of non-hla genotyping for risk assessment in allogeneic hsct. improvement of fi t of the ebmt risk score presents a powerful novel tool to assess this impact of gene polymorphisms in a complex heterogeneous hsct population. in a confi rmatory study the ebmt risk score was associated with survival for an al patient group. assessment of patient il (any t) and il rn (any c) in this second cohort is in progress. background: cognitive impairments have been found among patients receiving chemotherapy and hsct. however, studies with representative samples are rare, but needed to clarify the short and long term impact of different conditioning regimens and hsct. in the present multicenter trial, we assessed the prevalence and course of cognitive dysfunctions in patients with hematological diseases undergoing allogeneic hsct. moreover, the role of neurotoxic intensity of conditioning regimens was investigated. methods: patients ( % male) with mixed hematological cancers ( % acute myeloid leukemia) at an average of years of age were assessed before (t ), days after (t ) and one year following (t ) allogeneic hsct. % of the participants received intense neurotoxic conditioning regimens. a battery of neuropsychological tests using computer-and paperpencil-based methods was used covering the domains of attention, memory, executive and psycho-motor function. measures assessing self-perceived cognitive impairments and psychological distress were additionally applied. results: compared to published test norms at each assessment time point, signifi cant impairments in several neuropsychological test parameters across all cognitive domains were found. at baseline, patients had impaired cognitive functions in % of the test parameters. no signifi cant change in the prevalence of cognitive impairments was observed over time except for a mild increase in psycho-motor dysfunction at t . patients who were classifi ed as having intense neurotoxic conditioning were more likely to show impairments in the domain attention at t (p< . , d = . ) and showed a different time course of performance in attentional tasks compared to patients with mild neurotoxic conditioning (p< . , eta² = . ). conclusions: a remarkable amount of patients was classifi ed as having cognitive impairments prior to hsct. possible explanations include the impact of the hematological cancer disease, invasive treatments applied prior to conditioning and hsct, and treatment related distress. a decline in cognitive functioning was primarily limited to psycho-motor functions. however, subgroups of patients and in particular those with intense neurotoxic conditioning regimens might be at greater risk for developing short term cognitive impairments particularly in the domain attention. acknowledgement of funding: this study was supported by a research grant from the german josé carreras leukemia foundation. clinical outcomes of second allogeneic haematopoietic stem cell transplantation for acute leukaemia and myelodysplastic syndrome relapsing after fi rst allogeneic transplantation t. yamashita, t. kikuchi, i. kamiya, r. hanajiri, y. nagata, s. wakabayashi, k. taoka, t. kobayashi, k. ohashi, h. sakamaki, h. akiyama tokyo metropolitan komagome hospital (tokyo, jp) background: allogeneic hematopoietic stem cell transplantation (hct) is an effective therapy in acute leukemia and myelodysplastic syndrome (mds). but even after hct, these diseases recur in some cases and prognosis of these patients is very poor. second allogeneic hct (hct ) may be one of the most effective therapies in some patients who relapsed after fi rst allogeneic hct (hct ). because of the heterogeneity of hct , it is often diffi cult to fi nd useful prognostic factors and appropriate strategies. in our single-center study, we retrospectively analyze the clinical data of patients with acute myeloid leukemia (aml), acute lymphoblastic leukemia (all) and mds who received hct in our hospital to investigate the factors that affect clinical outcomes of hct . patients and methods: we included patients with aml, all and mds who received hct between and in our hospital. probability of overall survival (oas) and event-free survival (efs) were calculated using the kaplan-meier method. cumulative incidence rates were calculated using standard methods for hematopoietic recovery, acute and chronic gvhd, relapse and non-relapse mortality (nrm). multivariate analysis was performed with variables that can affect the clinical outcomes using cox proportional-hazards regression models. results: a total of patients were eligible for this analysis. five years oas of hct is . %. years cumulative incidence of relapse (ri) . % and patients relapsed after hct . relapsed patients group was signifi cantly lower incidence of chronic gvhd (p = . ). in this patients group, hct was performed for patients with aml (n = ), all (n = ), mds (n = ). myeloablative conditioning was used in cases. five years oas of hct was . %, and oas and efs of myeloabative hct was . % and . %, respectively. five years ri was , % and trm was . %. multivariate analysis showed that ) relapse to hct over days, ) age at hct under years, ) cr at hct , ) related donors are signifi cant favorable prognostic factors for oas of hct . conclusion: our study shows that myeloablative hct brought sustained remission in a proportion of patients with aml, all and mds who relapsed after hct . age and disease status at hct are important factors that infl uence the clinical outcomes. this study also suggests that the duration from relapse after hct and hct have impact on regimen-related toxicity of hct . hepatic dysfunction is frequent and diverse before and after allo-ric and has a major impact on the transplant outcomes p. barba ( ) introduction: hepatic dysfunction is one of the most frequent and less studied organ impairments in the setting of allogeneic stem cell transplantation (allo). to analyze the impact of pretransplant liver function on the outcome of allo-reduced intensity conditioning (allo-ric) and to determine the incidence, characteristics and risk factors of hepatic injury after allo-ric we conducted a retrospective study in two spanish centers. patients and methods: we analyzed adult patients receiving an allo-ric. the median follow-up for survivors was years (rg . - ). ric consisted in fl udarabine mg/m plus melfalan - mg/m or busulfan - mg/kg. pretransplant indicators of liver injury analyzed were aspartate aminotransferase (ast), alanine aminotransaminase (alt), gammaglutamyl transpeptidase (ggt), alkaline phosphatase (ap), total bilirubin (bil), prothrombin time (pt) and the category of severe hepatic disease according to hct-comorbidity index (hct-ci_hep). posttransplant severe hepatic injury (shi) was defi ned as maximal total serum bilirubin (bil) level > mg/dl or encephalopathy of hepatic origin (according to hogan, blood, ) . results: pretransplant liver laboratory tests abnormalities were found in ( %) patients. we observed high levels of ast, alt, ggt, ap, bil, pt and hct-ci_hep in ( %), ( %), ( %), ( %), ( %), ( %) and ( %) patients, respectively. among the pretrasplant indicators of liver injury analyzed, hct_ci hep showed to be the best predictor of transplant outcomes. in multivariate analysis (mva), hct-ci_hep high risk patients showed higher -days and -years nrm (hr . [ %ci . - . ] p = . and hr . [ %ci - . ] p = . , respectively), lower overall survival (os) ( % vs. %, p = . ), higher grade - agvhd p< . ]), higher grade - agvhd and a trend to higher cgvhd ( . [ %ci . - . ] p = . ). after allo-ric, a total of patients ( %) developed grade iii-iv liver toxicities. a total of ( %), ( %), ( %) and ( %) patients developed grade iii-iv levels of ast, alt, ggt, and bil at a median time of (rg - ), (rg - ), (rg - ) and days (rg - ), respectively. sixty-seven patients ( %) developed shi at years with a cumulative incidence of % ( %ci - ). main causes of shi were gvhd (n = , %) and pharmacological toxicity (n = , %). in mva, risk factors for shi were unrelated donors (hr . a hallmark of acute graft-versus-host disease (agvhd) is the selective attack of certain tissues, namely the gastro-intestinal tract, liver and skin but not others such as the pancreas and kidneys. imaging techniques such as whole body in vivo bioluminescence imaging, dynamic multiphoton-laser-scanningmicroscopy and ultramicroscopy utilized by others and our own group help to elucidate the elusive mechanisms underlying alloreactive t cell traffi cking. here we dissected the molecular mechanism that direct alloreactive t cells via vascular endothelial cells to target tissues in a murine mhc major mismatch hematopoietic cell transplantation mouse model. as a strategy we employed endothelial ligand blocking antibodies during the agvhd effector phase after alloreactive t cells leave secondary lymphoid organs. immunohistochemical analysis of target and non-target tissues revealed organ dependent upregulation of infl ammation induced endothelial ligands. the identifi ed surface molecules subsequently served as therapeutic targets. we learned that during the agvhd effector phase multiple endothelial ligands need to be blocked simultaneously to avert agvhd target manifestation. blocking of single molecules such as madcam- , vcam- , e-selectin, p-selectin alone could not prevent alloreactive t cell traffi cking to the intestinal tract, liver and skin. when we treated transplanted mice with the combination of two or even three antibodies we observed a reduction of alloreactive t cells. however, only when we blocked four targets simultaneously, namely madcam- , vcam- , e-selectin, p-selectin in combination we could prevent gvdh target manifestation in the skin, in the liver and the small intestines. from these data we conclude that alloreactive t cell homing depends on the recruitment by infl ammation induced endothelial ligands of target tissues. after clonal expansion of alloreactive t cells simultaneous blocking of several endothelial ligands was required to effi ciently abrogate agvhd in target tissues. our data indicate that t cell homing remains highly attractive for therapeutic interventions. our results also point out that several pathways should be targeted simultaneously in order to prevent agvhd. distinct temporal and spatial roles for host conventional and langerhans cells in the development of graft-versushost injury f. fallah-arani, h. goold, b.r. flutter, j. sivakumaran, c.l bennett, r. chakraverty royal free and ucms (london, uk) background: following bone marrow transplantation, donor t cell reactivity can be confi ned to the lympho-haematopoietic system or, in the presence of infl ammation, can additionally extend to peripheral tissues leading to graft-versus-host disease (gvhd). in this study, we have explored the role of individual host dendritic cell (dc) subsets in regulating these processes using models involving inducible depletion following donor t cell transfer to allogeneic chimeras. methods: balb/c thy . + t cells were transferred to established allogeneic chimeras generated following reconstitution of lethally irradiated b mice with a mix of balb/c and b .cd c-dtr bone marrow. cd c.dtr mice express a primate diptheria toxin receptor (dtr) under the control of the cd c promoter. in the resulting chimeras, host cd c + conventional dc (cdc) were depleted at the time of donor t cell transfer by injections of diphtheria toxin (dt). in experiments to test the role of host langerhans cells (lc), balb/c thy . + splenocytes were transferred to balb/c + b > b .langerin. dtr, where co-treatment with dt leads to depletion of host, epidermal lc. in each of the above settings, infl ammation was induced in some chimeras by systemic or local application of a toll-like receptor (tlr) agonist at the time of t cell transfer. results: absence of host cdc at the time of delayed t cell transfer abrogated accumulation of donor cd cells in recipient spleens and reduced in vivo cytotoxicity against host target b cells. this was associated with a lack of conversion to full donor chimerism, demonstrating a requirement for host cdc in priming the lympho-haematopoietic graft-versus-host response in the steady state. we then explored the role of host cdc and lc in a model of cutaneous gvhd, where local gvhd is induced following application of a tlr agonist (imiquimod). maximal imprinting of e-selectin ligand expression upon donor cd cells within the draining lymph node required the presence of host cdc. however, even in their absence, donor t cells were able to access the epithelium and cause injury. selective depletion of host lc had no effect on imprinting or tissue infi ltration of donor t cells. however, the capacity of donor t cells to induce gvhd was severely impaired in this case. conclusions: these data show that host cdc are required for gvh reactivity in the steady state. in contrast, in a model of skin gvhd, host cdc are dispensable whereas lc have a unique post-priming role. failure to imprint donor cd memory and exhaustion may explain loss of gvt responses following donor leukocyte infusions b.r. flutter ( ) , f. fallah-arani ( ), s. sivakumaran ( ) , c.l. bennett ( ) , s. ghorashian ( ) , g. freeman ( ) , m. sykes ( ) ( )university college london (london, uk); ( )harvard medical school (boston, us) background: donor t cell alloreactivity can be co-opted to deliver graft-versus-tumour (gvt) responses following donor leukocyte infusions (dli) given after bone marrow transplantation (bmt). however, the major reason for treatment failure following dli is relapse, suggesting a long-term failure of antitumour immunity. a distinctive element to the gvt response is that the antigens (ag) to which the response is directed are cleared from the haematopoietic system but continue to be expressed by non-haematopoietic cells. in this study, we have explored the infl uence of non-haematopoietic ag upon the antihost cd t cell response in a model of delayed dli. methods: b cd . t cells were given after weeks to allogeneic chimeras where ag was ubiquitous (b + bdf >bdf ) or restricted to the haematopoietic compartment (b + bdf >b ). in each setting, direct priming of the initial t cell response by bdf ag-presenting cells resulted in the eradication of bdf haematopoietic cells. however, in b + bdf >bdf chimeras, ag was still present in peripheral tissues, whereas in b + bdf >b chimeras, ag was cleared completely. the donor t cell response was tracked at timed intervals following transfer in both sets of chimeras. results: the continued presence of non-haematopoietic ag led to a failure to sustain donor cd cytotoxic and effector cytokine responses by d following transfer and this was associated with a failure to establish a central memory (t-cm) population. in contrast, where ag was absent in peripheral tissues, functional t-cm populations were preserved. the failure to generate donor t-cm in chimeras with ubiquitous ag expression occurred at two distinct levels. firstly, residual dli-derived cd t cells demonstrated a pd- hi/cd lo/ ifn-g neg 'exhausted' signature. consistent with this, cd functions were partially restored by in vivo antibody blockade of the pd- pathway. secondly, during the initial phase (< d) of the response, we observed a lack of memory precursor formation when ag was ubiquitous. thus, when ag was ubiquitous, the capacity of activated, post-mitotic, cd t cells to establish recall immunity following transfer to ag-free hosts was much reduced. conclusions: these data demonstrate that alloantigen within non-haematopoietic tissues is not ignored, but rather blocks memory imprinting and drives eventual cd t cell exhaustion. while these effects may lessen the risk of gvhd, they might also impair the durability of the gvt response. background: adoptive transfer (at) of tcr-gene transduced t cells has become a promising therapeutic tool, however, limited in vivo survival and the development of an unresponsive state often termed 'exhaustion' has hampered reproducible clinical success. here we demonstrate that a) upregulation of pd- on donor-derived tcr-engineered t cells is associated with a loss of gvl effects after late at and b) pd-l blockade after hematopoietic stem cell transplantation restores robust gvl-effects. methods: we chose a mhc-mismatched allogeneic bmt model (b .a mice (h- a) into c bl/ (h- b) mice) and a retrovirally encoded tcr (ot- anti-ovalbumin) that was introduced into b .a-derived donor t cells for at after mhc-mismatched hct. after hct, leukemia-bearing (c -ova) mice (syngeneic hct-recipients served as reference) received at and were monitored for gvhd, gvl, and in vivo t cell persistence/ functionality. for pdl- blocking experiments pdl- antibodies (clone f. g ) were used. results: ) up to x tcr-transduced cd + t cells were generated in vitro within days using a retroviral vector system linking the genes for the á-and â-chain via a a sequence. % of cd + t cells coexpressed the respective tcr chains vá and vâ . ) the introduced tcr mediated comparable specifi c cytotoxicity against c -ova in vitro being functional on autologous and allogeneic t cells. ) after early at transduced t cells rescued up to % of mice in a dose dependent manner. (p = . - . versus mock-transduced controls). ) after late adoptive transfer (day ) autologous t cells peaked in numbers by day after at and provided strong gvl-effects. in contrast, numbers of allogeneic tcr-modifi ed t cells declined rapidly within the peripheral blood and increasingly expressed pd- . lower frequencies of allogeneic t cells in the periphery translated into nearly abolished gvl effi cacy upon leukemia challenge days after at (p = . ). ) pd-l blockade in vivo after allogeneic hct restored gvl-effi cacy of adoptively transferred t cells raising leukemia free survival from % to % (p = . compared to isotype controls) without increasing gvhd rates. conclusion: at with tcr-engineered t cells after allogeneic hct can decrease the risk of gvhd and provide potent gvl effects. however, increased expression of pd- on adoptively transferred t cells is associated with decreased gvl effi cacy and can effectively be reverted by pd-l blockade. allogeneic hematopoietic stem cell transplantation (allo-hsct) is a curative treatment for many hematologic malignancies or hematopoietic dysfunction syndromes in adult patients, but the application is still limited due to major complications, such as severe graft versus host disease (gvhd). diagnosis of agvhd is based on clinical features and biopsies. we have shown earlier that agvhd can be diagnosed using a proteomic pattern established by screening with capillary electrophoresis (ce) and mass spectrometry (ms). the agvhd-specifi c proteomic pattern has been evaluated prospectively and blinded on more than samples collected from more than patients undergoing allo-hsct at hannover medical school (mhh) and additional clinics. since a new diagnostic proteomic pattern for agvhd was developed, using a higher resolution ms. thus, patients from mhh and patients from collaborating clinics were subjected to double screening for development and prospective evaluation of the ms- -agvhd-specifi c pattern. the majority of the patients included was transplanted for hematological malignancies (n = ), for hematopoietic failure syndromes ( pnh, saa) . conditioning regimens included reduced intensity conditioning regimens (ric) for about % of the patients of mhh, as well as standard conditioning regimens (mainly tbi + cy or busulfan + cy). gvhd-prophylaxis was cyclosporine a (csa) and mycophenolate (mmf) or csa metothrexate (mtx), as appropriate. in addition, about % percent of the patients received atg (antithymocyte globulin) prior to hsct. the new agvhd-specifi c pattern, ms_ , consisting of differentially excreted proteins and peptides was developed and prospectively evaluated in parallel for the last years. prospective and blinded evaluation of the patients included in this analysis for early recognition of patients at risk for agvhd development revealed the correct classifi cation of patients developing agvhd about days (range: - days) prior to the development of clinical symptoms for agvhd with a sensitivity % and specifi city of about %. pre-emptive therapy ad ministered upon positivity of the protemic patterns reduced the incidence and severity of agvhd (p = . ). thus, a multicenter study has been initiated in germany to test the effi cacy and safety of pre-emptive therapy. hhv and acute gvhd c. pichereau, k. desseaux, a. janin, r. peffault de latour, m. robin, p. ribaud, s. chevret, g. socié hôpital saint louis (paris, fr) background: previous studies have suggested that hhv infection is correlated with acute gvhd following allogeneic hsct, but whether hhv triggers, is associated with, or is a differential diagnosis of gvhd is unknown. methods: patients received an allogeneic hsct at the saint louis hospital (paris) between january and december . whenever acute gvhd was suspected, hhv rq pcr and organ biopsy was performed. cumulative incidence of hhv reactivation was estimated using competing risks approaches. predictive factors of increased risk of reactivation or acute gvhd were assessed using cause-specifi c hazard cox models. effect of reactivation on further occurrence of acute gvhd was tested by introducing a time-dependent variable. results: pts were tested for hhv : ( %) were male, median aged years . ( %) had malignancies and suffered from non malignant disorders (aplastic anemia, sickle cell disease, thalassemia). pts were grafted with related donors ( %). the conditioning regimen was myeloablative in cases ( %). the source of hsct was bone marrow in cases, peripheral blood in cases and cord blood in cases. pts ( %) developed acute gvhd. hhv was detected in pts, of whom developed gvhd. hhv reactivation was signifi cantly associated with cord blood graft ( % versus %, p< . ) and unrelated transplant ( % versus %, p = . ). gvhd was signifi cantly associated with previous hhv reactivation (hazard ratio, . ; % confi dence interval, . - . ; p = . ). tissue biopsies were available for of the hhv infected pts ( skin and gut samples). there were no signifi cant difference between pts who had a biopsy and pts who hadn't, except for the incidence of acute gvhd ( % versus %, p = . ). pts who had no or poor pathological evidence of gvh (grade or ), later on developed severe clinical acute gvhd (grade iii and iv), suggesting the role of hhv as a trigger of severe gvhd. beside this, pts who didn't have clinical and histopathological gvhd (grade ) showed a significant lymphoid infi ltrate on their biopsy. immunohistochemical analysis are ongoing, that could provide the evidence of "pure" hhv -related skin rash after hsct. conclusion: this study suggests the role of hhv as a trigger of acute gvhd, particularly in its severe manifestations. beside this, it confi rms that hhv infection is a differential diagnosis of acute gvhd in a signifi cant proportion of patients. prochymal® improves response rates in patients with steroid-refractory acute graft-versus-host disease involving the liver and gut: results of a randomized, placebo-controlled, multicentre phase iii trial in gvhd p.j. martin ( ) , j.p. uberti ( ) background and methods: steroid-refractory acute gvhd (sr-gvhd) remains a signifi cant and life-threatening complication of allogeneic hematopoietic cell transplantation (hct). prochy-mal® (mesenchymal stem cells, msc, derived from unrelated volunteer adult donors) was evaluated in addition to standard of care, including institutionally selected second line treatment, in a randomized ( : ) trial in patients with sr-gvhd (protocol ). patients received infusions of x msc/kg over weeks (or volume equivalent for placebo), with an additional infusions administered weekly after day in patients who had a partial response, defi ned as improvement in at least one organ without progression in others. the primary endpoint was durable complete response (dcr) for ≥ days. additional prospectively defi ned outcomes included responses in patients by organ involvement. patients and results: patients with sr-gvhd (skin involvement n = , gastrointestinal involvement n = , liver involvement n = ) were enrolled and treated on a : basis: prochymal (n = ), placebo (n = ). there were no signifi cant differences in age, pre-transplant conditioning, graft source, hla-matching or second-line therapy between treatment arms. for the prochymal and placebo arms, respectively, the grades of gvhd at entry were b ( % vs. %), c ( % vs. %), and d ( % vs. %). the respective dcr rates were % vs. % (p = . ) in the intent-to-treat population and % vs. % (p = . ) in the per protocol population. results for secondary endpoints are shown in table . patients with gvhd affecting all organs had overall complete or partial responses rate of % vs. % (n = , p< . , fisher's exact test) at day . patients treated with prochymal had less progression of liver gvhd at weeks and respectively ( % vs. %, p = . ; and % vs. %, p = . ). the incidence of infections was not different between arms. incidence rates were % vs. % for recurrent malignancy, . % vs. . % for infusional toxicity, and . % vs. . % for ae-related discontinuation in the prochymal and placebo arms, respectively. conclusion: gvhd with liver or gut involvement is a life-threatening complication of hct. these results suggest that the addition of prochymal produced signifi cant improvement without additive toxicity in patients with sr-gvhd involving visceral organs. s o treatment of steroid-refractory acute gvhd with mesenchymal stem cells improves outcomes in paediatric patients. results of the paediatric subset in a phase iii randomized, placebo-controlled study p. szabolcs ( ) , g. visani ( ) background: successful treatment of steroid-refractory acute graft-versus-host disease (sr-gvhd) following allogeneic hematopoietic cell transplantation remains a signifi cant challenge. because of their immunomodulatory properties and safety profi le, adult mesenchymal stem cells (mscs) have been proposed as a treatment for sr-gvhd. intravenous allogeneic msc therapy (prochymal) for sr-gvhd was independently evaluated in the pediatric subset of a double-blind, placebocontrolled study. methods: pediatric patients (< years old) with grade b-d sr-gvhd were randomized to receive either prochymal or placebo in addition to standard of care, including institutionally selected second line agent. patients received infusions of × cells/kg for weeks (or volume equivalent for placebo), with more infusions weekly in the case of a partial response (pr). the primary endpoint was durable complete response (cr days); secondary endpoints included incidence of cr, pr and progression through days, survival, and safety. results: twenty-eight children were randomized to prochymal ( % male, % caucasian) or placebo ( % male, % caucasian), with a median age of yrs (range - ) and years (range - ), respectively. the dominant transplant graft source was cord blood ( % prochymal, % placebo), with mostly unrelated donors ( % vs. %, respectively). the median duration of agvhd prior to enrollment was days for prochymal and days for placebo (p< . ). at baseline, agvhd grades b:c:d were : : for both arms. for prochymal, organ involvement was % skin, % gi, and % liver. for placebo patients, organ involvement was % skin, % gi, and % liver. durable cr was % for prochymal and % for placebo (p = . ). prochymal improved rates of cr and or (table) . the median time to cr was days vs. days. the safety data showed no infusional toxicity and no evidence of prochymal leading to ectopic tissue. there were no aes leading to discontinuation of therapy. conclusion: in a sr-gvhd population in which % of patients had grade c/d disease, the addition of prochymal to standard of care resulted into a faster and better cr. in view of their increased response rates and a well-tolerated safety profi le, mscs appear to be a safe and effective therapy in the treatment of pediatric patients with sr-gvhd. aim: response rate after induction and after asct. patients and methods: td consisted of thalidomide mg/d and dexamethasone mg on days - and - at -week intervals for cycles. vtd was identical to td plus velcade . mg/m² on days , , , of each cycle. combination chemotherapy plus velcade consisted of cycles of vbmcp/vbad followed by cycles of velcade. results: as of december , , patients (median age: yrs, m , f: ; igg , iga , light chain , others ) entered the study. of ( %) had high-risk cytogenetics (t( ; ), t( ; ) and/or p deletion). all patients (td: , vtd: and vbmcp/vbad/velcade: ) were evaluable for response and toxicity to induction therapy. the cr plus vgpr rate was signifi cantly higher with vtd than with vbmcp/vbad/velcade ( vs. %, p = . ) and td ( vs. %, p< . ). the progressive disease (pd) rate was higher with td than with vtd ( vs. %, p = . ). in patiens with high-risk cytogenetics vtd was associated with a higher cr rate than td and vbmcp/vbad/velcade ( vs. %, p = . and vs. %, p = . , respectively) as well as with a lower pd rate ( vs. %, p = . and vs. %, p = . , respectively). the incidence of > grade thrombotic events was higher with td than with vtd ( % vs. %, p = . ) and grade > peripheral neuropathy was higher with vtd than with td or vbmcp/ vbad/velcade ( vs. % vs. %, p< . ). patients were evaluable for response after asct. the cr rate was higher with vtd ( %) than with vbmcp/vbad/velcade ( %) and td ( %) although the differences did not reach statistical signifi cance. the estimated os at yrs. was % with no significant differences among the arms. ttp and pfs were shorter with td (p = . and p = . , respectively). summary: ) induction with vtd produced a higher cr + vgpr rate, ) in patients with high-risk cytogenetics, td resulted in a signifi cantly lower cr and higher pd rate than bortezomibcontaining regimens, ) the pfs was shorter with td and ) asct increased the cr rate in %, % and % in the td, vtd and vbmcp/vbad/velcade arms, respectively. interim analysis of a phase iii, prospective randomized trial of melphalan stem cell source was pbsc in % pts. % pts received myeloablative allogeneic sct as part of a planned tandem strategy, whereas % received a non myeloablative conditioning. overall response at allo sct was % and included complete remission (cr %), very good partial remission (vgpr %), partial remission (pr %). results: all pts except had sustained donor engraftment. among pts, %, % and % achieved cr, vgpr and pr respectively at day . median follow-up from allo sct was months (range - ). event-free survival (efs) was % ( - ) at years. cumulative incidence of transplant related mortality (trm) at days was % and was mostly related to pneumonia. a single line of treatment before tandem auto/allo sct was associated with improved -year-efs (p = . ). % pts experienced grade i to ii acute graft-versus-host-disease (gvhd) and % grade iii to iv. % pts had limited and % extensive chronic gvhd. whereas grade i to ii acute gvhd was associated to better -year-efs (p< . ), extensive chronic gvhd had no statistically signifi cant impact on efs. moreover the absence of deletion was not associated with a better efs. conclusion: our results suggest that the number of treatment lines received before tandem auto/allo sct is an important issue, with an improved efs for pts treated by a single line before tandem. whereas better control of acute gvhd might further improve survival, impact of the deletion on outcome seems limited. purpose: the outcome of mm patients treated with low dose tbi and allogeneic related sct following autologous sct has been shown to be superior to that from two autologous sct (gahrton et al.; bruno et al.) . since related donors are available for approximately % of patients only, the outcome of allogeneic unrelated sct in patients without a related donor was compared to that of patients with a related donor in an intention to treat analysis. patients and methods: mm patients (n = ) with a median age of (range - ) years were treated at the university of leipzig with autologous sct followed by allogeneic sct. autologous stem cells were mobilised with cyclophosphamide and transplanted after melphalan mg/m on day - . allogeneic sct was performed at a median of (range - ) months after autologous sct using fludarabine ( mg/m /d from days - to - ) and tbi ( gy on day ) followed by cyclosporine ( . mg/kg twice daily from day - ) and mycophenolate mofetil ( mg/kg daily from day ). patients received an unrelated (n = ) or related (n = ) stem cell graft. results: patients with unrelated donors (n = ) showed hematological toxicity comparable to those of related donors. all patients had stable hematopoietic engraftment with t-cell chimerism of % - months after sct. with a median follow up of (range - ) months overall survival (os) at years was ± % and ± % (p = , ) in patients with unrelated and related donors, respectively. progression free survival (pfs) was ± % at years for patients with an unrelated compared to ± % for patients with a related donor. this difference was due to a decreased relapse incidence (ri) in patients with unrelated donors ( ± %) compared to related donors ( ± %). non relapse mortality was ± % in the whole population with no difference between the two groups. of the patients having received allogeneic unrelated sct, ( %) are currently in cr, ( %) in vgpr, ( %) in pr and ( %) has stable/progressive disease, with no difference between related and unrelated sct. conclusion: our data confi rm the feasibility of autologous sct followed by low-dose tbi conditioning regimen and unrelated sct. pfs seems to be higher and ri lower in unrelated compared to related sct. these feasibility data provide the basis for a phase iii study comparing auto-allo unrelated to auto-auto sct. background: poems syndrome is a rare paraneoplastic syndrome resulting from a clonal plasma cell proliferation producing a small monoclonal protein usually lambda type restricted. this syndrome is a multisystemic disease, its major clinical feature being a progressive peripheral neuropathy. single osteosclerotic lesions should be treated with radiation therapy only. in widespread disease, high-dose therapy followed by autologous hematopoietic stem cell rescue (asct) has shown to be an effective therapeutic option in short series, but with significant morbidity. patients and methods: between december and september , patients with poems syndrome ( female/ male) were treated with melphalan- ( patients) or melphalan- ( patient) followed by asct at spanish institutions. median age was years (range: - ). all of them presented a m protein lambda type (iga: ; igg ), peripheral polyneuropathy ( ), osteosclerotic lesions ( ), organomegaly ( ), endocrinopathy ( ), skin lesions ( ), extravascular volume overload ( ), papilledema ( ), polycythemia ( ) and thrombocytosis ( ). four patients had pulmonary hypertension, two portal hypertension, and three castleman disease. four patients were previously untreated. the median number of prior therapies was (range, - ). median time from diagnosis to asct was months (range, - ). results: no transplant-related-mortality (trm) was observed. after a median follow-up of months (range, - ), one patient has died of progression months post-asct. five patients presented an engraftment syndrome and two a primary graft failure resolved, one with a back-up infusion on day + and the other presented a delayed engraftment. sixteen patients were evaluable for response, achieved a complete hematologic response (cr) (if-), a near-cr (negative electrophoresis but if positive) and one died of progression months post-asct. clinical improvement was observed at - months post-transplantation. all patients had a signifi cant organic improvement. the four patients with severe pulmonary hypertension and the two with portal hypertension improved of both pressures. conclusions: in this series, asct proved to be a highly effective therapy for patients with widespread poems syndrome. despite no trm was observed, these patients may have a delayed hematopoietic recovery and may develop an engraftment syndrome that must be diagnosed/treated promptly. a salvage treatment containing novel agents consolidated by allogeneic stem cell transplantation with reducedintensity conditioning improves outcome of multiple myeloma patients failing autologous transplantation f. patriarca ( ), h. einsele ( ) objectives: allogeneic stem cell transplantation (allo-sct) employing reduced intensity conditioning (ric) is a feasible procedure in selected patients with relapsed multiple myeloma (mm), but its effi cacy is still a matter of debate. we investigated the role of ric allo-sct in mm pts who relapsed after auto-sct and were then treated with a salvage therapy based on novel agents. our study was structured similarly to an intention to treat analysis and included only those pts undergoing a hlatyping immediately after the failure of auto-sct. the cohort of pts having a donor (donor group) was compared with the one not having a suitable donor (no donor group). patients and methods: one hundred thirty-six consecutive pts were retrospectively evaluated. fifty-seven found a donor and ( %) underwent an allo-sct: identical sibling ( %), mud ( %). conditioning regimens were fl udarabine, melphalan±thiotepa in patients ( %), fl udarabine + gy tbi in cases ( %), fl udarabine and cyclophosphamide in patients ( %) and fl udarabine and treosulfan in the remaining cases ( %). seven pts having a donor did not receive allo-sct for progressive disease or severe comorbidities. median age of donor group was signifi cantly younger than no donor group ( versus years, p = , ). the groups were balanced with regard of time between auto-sct and relapse, treatment of fi rst relapse, duration of salvage treatment, and quality of response after salvage treatment. results: the median follow-up for all patients was months ( - ) after relapse. the median time to progression-freesurvival (pfs) and overall survival (os) for all patients were and months, respectively. two-year pfs was % in the donor-group and % in the no-donor group (p = . ). twoyear os was % in the donor-group and % in the no-donorgroup (p = . ). in multivariate analysis the availability of a donor was statistically signifi cant for pfs (p = . ); moreover a signifi cant difference in outcome was observed comparing patients who achieved cr + vgpr versus pr versus sd/pd after salvage treatment (p = . , p = . , p = . for pfs, p = . , p = . , p = . for os). conclusions: this study comparing salvage treatment with novel agents consolidated by ric allosct versus salvage treatment alone after a failed auto-sct provides evidence for a signifi cant pfs benefi t and a trend for a prolonged os in mm patients having a donor. grouping patients based on d and non-d kir ds allelic status revealed individuals homozygous for kir sd (d) had significantly shorter duration of fever, compared to those heterozygous or homozygous for kir ds (non-d), (p = . ). this held true on multivariate analysis. no signifi cant association was seen between kir ds genotype and neutrophil engraftment time or microbial isolates. of note, there was a signifi cant association between kir dl positivity and gram-ve bacteremia (p = . ). these data support a role for kir genotype and infectious complications post-asct. kir genotyping may aid risk assessment of infectious complications post-asct and optimization of anti-infectious prophylaxis, surveillance and treatment in those deemed at risk of sepsis. these results suggest a functional role for the deleted kir sd variant, previously believed non-functional in the innate immune response. in the era of novel therapeutic agents, high-dose chemotherapy and autologous stem cell transplantation (asct) remains an integral part of treatment for multiple myeloma (mm). therefore, the choice of new drug combinations for induction therapies must take into consideration the requirement to collect a suffi cient number of haematopoietic stem cells (hscs) for one or more courses of asct. lenalidomide and melphalan have been shown to impair hsc mobilisation, but induction therapies containing either thalidomide or cyclophosphamide do not have a relevant impact on the hsc collection yield. we considered the possibility that the combination of cyclophosphamide and thalidomide could have an additive impact on the hsc compartment and carried out a retrospective analysis of the outcome of peripheral blood hsc mobilisations performed in mm patients. patients who had received induction therapy with ctd (oral cyclophosphamide, thalidomide, dexamethasone; n = ) were compared with a control group of patients (n = ) who had received vad (n = ) or z-dex (idarubicin, dexamethasone; n = ) during the same -year period. all mobilisations were performed with cyclophosphamide and g-csf. our standard collection target was × cd + cells/ kg, with a minimal target of × being accepted if patients failed the standard target. the total number of cd + cells harvested was signifi cantly lower in the ctd group ( . vs. . × /kg, p = . ). the number of cd + cells harvested on the fi rst day of apheresis and per apheresis procedure were also lower in the ctd group ( . vs. . × /kg, p = . ; . vs. . × /kg, p = . ). more patients in the ctd group failed to achieve both the standard ( . % vs. . %, p = . ) and minimal ( . % vs. . %, p = . ) stem cell harvest target, despite a higher number of patients in the ctd group undergoing two or more apheresis procedures ( . % vs. . %, p = . ). the failure rate on the fi rst day of apheresis was also higher in the ctd group both for the standard ( . % vs. . %, p = . ) and the minimal target ( . % vs. . %, p = . ). age or number of induction therapy cycles did not have an impact on mobilisation failure in the entire cohort or the ctd group alone. these observations provide novel evidence that drugs with no previously demonstrated effect on hsc mobilisation can have a considerable negative impact when used in combination, which can result in a high rate of hsc collection failures. a. nagler ( ) background: allogeneic transplantation of hematopoietic stem cells (allo-sct) from an hla-matched related (mrd) or unrelated donor (urd) is a curative option for patients (pts) with high-risk hematological disease (hrhd). in the absence of a mrd, pts have been offered investigational transplant strategies such as umbilical cord blood (ucb) or family haploidentical sct (haplo-sct). in our institution, all patients with hrhd are typed at entry; if a suitable mrd donor is missing a urd search is promptly activated. our policy is to offer an haplo-sct to adult pts lacking an mrd or urd in order to adequately treat hrhd in the ideal appropriate time according to clinical indications to allo-sct agreed in ongoing protocols for primary disease. methods: here we report the retrospective intention-to-treat (itt) analysis of alternative donor transplantation at our institution. data were obtained from institutional database. results: between jan- and nov- , pts ( % of the following itt analysis; median age y) received indication to allo-sct according to ebmt recommendations. eightytwo pts ( %) received a transplant from a mrd; pts ( %) activated an urd search in the ibmdr registry; pts ( % of total pts, % of urd searching) received a urd transplant; pts ( . %- , %) received an haplo-sct due to lacking of a suitable urd in the appropriate timing according to disease status, or absence of suitable criteria to engage an urd donor; pts ( , %- , %) received a ucb because lacking a suitable haplo donor. overall, pts received an haplo-sct ( %): after ibmdr research, up-front. nineteen pts died before receiving a transplant ( %), ( %) are searching for a suitable donor. if we consider only pts with acute leukemia ( pts, median age y, range - ; over y pts) pts ( %) received a transplant from a mrd; pts ( %) activated an urd search; pts received a urd transplant ( %), pts ( %) a ucb, pts an haplo-sct ( %). eight pts died before receiving a transplant ( %), ( %) are searching for a suitable donor. allo-sct was performed in complete remission in pts ( % alive), in persistence of disease in pts ( % alive). conclusion: in itt analysis, % of overall pts candidate received an allo-sct: % from an alternative donor, % from a mrd. the highly committed policy performed in the alternative-sct setting and the implementation of an alternative donor option are essential prerequisite to obtain these results. there was a marked variation in total numbers of transplants performed between countries ranging from to . the median age of hsct programs was yrs (range - ). the total number of hsct performed per year has continued to increase and is yet to plateau. a greater proportion of transplants was allogeneic hsct (allo-hsct) compared to autologous hsct (asct) ( % vs. %). acute leukemia constituted the main indication for allo-hsct( %). there was a relatively high rate of hsct for bone marrow failure (n = , %) and hemoglobinopathies (n = , %) when compared to data reported to ebmt and ibmtr. cml continued to be an indication for . % of allo-hsct in . the rate of unrelated donor hsct remained low, with only non-umbilical cord unrelated donor transplants over the surveyed period. the use of peripheral blood stem cells varied between countries though increasingly constituted the main source of stem cells in allo-hsct. ric was used in . % of allo-hsct in the the survey. asct rates continue to increase; while acute leukemia was the main indication for asct in the s, overall the main indications for asct in the survey were lymphoma ( %) followed by myeloma ( %). conclusion: we present the fi rst survey of hsct activity in the em region over years which refl ects the unique health conditions of this region, accounting for notable differences in transplant practices from europe and north america. annual hsct rates continue to rise. economic, logistic and other factors are likely to be responsible for disparities in activity within the region. this survey may be valuable in providing a basis for healthcare planning in the fi eld of hsct in the region. background: related haploidentical donors, as cord blood, can be alternative donor sources in stem cell transplantation (sct). severe graft-versus-host disease (gvhd), however, has interfered the progress of haploidentical stem cell transplantation (haplosct). to deal with this strong gvhd, t cell depletion has usually been used in european countries. on the other hand, based on the difference of ethnicity prone to less severe gvhd in japan, we have performed unmanipulated haplosct using myeloablative or reduced intensity preconditioning regimen for ten years. in this meeting, we will summarize our experience of ten years. , and patients over years old or with comorbidities or repetitive sct (including second to fi fth sct) underwent reduced intensity preconditioning regimen consisting of flu/(ca)/bu/atg or flu/(ca)/mel/atg (haplomini, n = ). high dose ara-c (ca) was optional to reduce tumor burden. atg (fresenius) dose was mg/kg/day for four days. gvhd prophylaxis consisted of taclolimus (tac), methylprednisolone (mpsl) mg/kg/day, short term mtx, and mycophenolate mofetil (mmf) mg/kg/day in haplo-full, and tac, mpsl mg/kg/day in haplo-mini, respectively. for elderly patients over years old in haplo-mini, mmf was added. results: hematopoietic engraftment in haplosct was as rapid as that in hla-identical sct, except eight cases of graft rejection. acute gvhd (grade ii-iv) was observed in %. overall survival in fi ve years is % in haplo-full and % in haplo-mini, respectively. if limited to cr cases, overall survival reached over % in haplo-mini. there is no difference in survival rate among patients' diseases. discussion: unmanipulated haplosct is feasible and effective for refractory diseases. atg dose used in haplo-mini is rather low compared with that of european cases reported so far. although it should be too early to refer long term outcome, unmanipulated haplosct could be considered as an option to fi ght against refractory diseases. background: while chemotherapy + g-csf (g) is effective for autologous stem cell mobilization (scm) it exposes patients (pts) to risks and side effects. plerixafor (p) + g provides a safer alternative for mobilization of hematopoietic stem cells in pts with mm or nhl. to better understand the comparative effectiveness of standard and newer approaches for scm, we conducted a study aimed to: ( ) determine the clinical outcomes and cost of scm with cyclophosphamide (cy) plus g ( ) compare outcomes of cy + g to a clinical trial of p + g. methods: a retrospective study was conducted in all pts undergoing fi rst scm from / to / (n = ) with cy ( gm/m ) and g ( mcg/kg started day after cy for days with planned fi rst day of collection on day ). apheresis was initiated when peripheral blood had > cd + cells/μl. positive clinical outcome (pco) was defi ned as > × cd + cells/kg collected on planned day of collection and within or apheresis without a prior negative event that led to additional clinic/inpatient evaluation. the cost of drugs, apheresis, product processing, and clinical events were reported based on medicare part b physician, laboratory, and ancillary fee schedule. data on pts undergoing scm with p + g were obtained from published clinical trials in pts with myeloma (mm) or lymphoma (l) undergoing fi rst scm using g ( mcg/kg without dose escalation) and a maximum of days of p. results: among cy + g pts, ( %) were males; ( %) had mm, ( %) had l ( -other diagnoses); and ( %) pts. had prior radiation. pco was seen in ( %) pts.; % of mm and . % of l pts. median total cost of cy + g scm was $ , (range, - ). pco was associated with a lower cost than non-pco in the overall group, (mean, $ , vs. $ , , p = . ), in mm pts. (mean, $ , vs. $ , , p = . ), and in l pts (mean, $ , vs. $ , , p = . ). assuming a similar distribution of pco in pts. with mm and l, the projected per pt cost of scm would be $ , and $ , (mean, $ , ) with cy + g. projected costs of scm using p + g for pts. with mm and l were $ , and $ (mean, $ , ). conclusion: scm with cy + g was associated with a lower rate of pco and higher cost than p + g, supporting front-line use of p + g for scm. future prospective studies should investigate whether scm with p + g translates into decreased resource utilization and improved quality of life for pts undergoing scm. . x /l, platelet count (pt) > × /l, and hemoglobin level (hb) > g/l], good partial response (gpr) [anc > × /l, pt > × /l, and a hb level > g/l] and poor partial response (ppr) [anc > . × /l, pt > × /l, hb level > g/l and transfusion independency]. no response (nr) was defi ned as failing criteria for at least ppr. non-responding patients received a second-line therapy (hsct or androgens), a second course of ist with lg ( mg/kg/day/ × days) or tg ( . mg/kg/day/ × days); or no treatment. subgroup analyses were conducted and differences in response were tested using the chi-square statistic test. immunoablation followed by immune reconstitution by transplantation of autologous peripheral blood stem cells is a promising approach to treat refractory or otherwise incurable s autoimmune diseases. it has recently been suggested that this treatment could lead to remission in some cases of the early diabetes type if administered prior complete destruction of beta cells by autoimmune mechanisms. the objective of this study was to verify these fi ndings on an independent group of patients. methods: eight patients (age - ) with early diabetes type (no more than weeks from diagnosis, c-peptide positive, anti gad -antibodies positive) were qualifi ed for the treatment. the patients were subjected to - plasmaphereses to remove circulating immune complexes and then mobilized with cyclophosphamide and g-csf. leucaphereses were continued until > , x cd + cells/kg were collected. the conditioning consisted of cyclophosphamide ( mg/kg/day on days - , - , - , - prior to transplant) and antithymocyte globulin (atg genzyme -of . mg/kg/day given on day - , . mg/kg/day given on days - , - , - , - and - ). results: median follow up for the patients as of december is months (range - months). no major complications were observed during the transplantation and in the posttransplantation period. all patients ( / ) became independent from exogenous insulin after the transplantation. median day of insulin withdrawal was + (range + to + ). one patient resumed insulin months post transplant. six out of patients were given acarbose to reduce the potentially toxic infl uence of observed hyperglycemias. after the transplantation patients exhibited good control of glycemia -the average hba c concentrations were . % at the diagnosis, and . %; . %; . % - , , and months after the transplantation respectively. this was correlated with increased levels of c-peptide after the transplantation. conclusion: this report independently confi rms that independence of exogenous insulin could be reached in diabetes type patients following immunoablation and reconstitution of the immune system with autologous peripheral blood stem cells providing that the procedure is performed prior total destruction of beta cells in the pancreas by the autoimmune mechanisms. the use of acarbose may have potentially positive effect on the duration of remission of diabetes type in the patients after the transplantation. new perspectives on the use of autologous haematopoietic stem cell transplantation in multiple sclerosis: three strategies of high-dose immunosuppressive therapy with autologous haematopoietic stem cell transplantation during the last decade high-dose immunosuppressive therapy with autologous hematopoietic stem cell transplantation (hdit + asct) has been used with increasing frequency as a therapeutic option for ms patients. we aimed to study clinical outcomes in multiple sclerosis (ms) patients after early, conventional, and salvage of hdit + asct. ms patients (secondary progressive- , primary progressive- , progressive-relapsing- , relapsing-remitting- ) were included in this study (mean age- . ; male/female - / ). beam or mini-beam conditioning regimens were used. patients did not have any supportive treatment after hdit + asct; patients with risk factors were administered mitoxantrone during a year after asct as a consolidation therapy. patients underwent early transplantation (edss . - . ), -conventional (edss . - . ), -salvage transplantation (edss . - . ). median edss at base-line - . . the mean follow-up duration - months (range - ). neurological evaluation was performed at baseline, at discharge, at , , , months, and every months post transplant. transplantation procedure was well tolerated with no transplant-related deaths. the effi cacy analysis was performed in the group of patients who did not have consolidation treatment. out of patients with the follow-up at least months the following distribution of patients according to clinical response at months post transplant was observed: patients ( %) achieved an objective improvement of neurological symptoms; patients ( %) had disease stabilization; patients ( %) progression. in the patients who underwent early transplantation (n = ) ( %) patients stabilized and ( %) improved. after conventional transplantation (n = ) ( %) patients stabilized, ( %)-improved, and ( %)-progressed. after salvage transplantation (n = ) patients were stable, and improved. at long-term follow-up (median- . months) out of patients improvement was registered in ( %) patients; stabilization -in ( %) patients, and progression in patients ( patient after early; -after conventional transplantation). no active, new or enlarging lesions were registered in patients without disease progression. thus, hdit + asct appears to be a safe and effective treatment for ms. further studies should be done to establish the best timing for transplantation and to evaluate consolidation therapy in patients receiving early, conventional, and salvage transplantation. in the last decade, the so-called nonmyeloablative or reduced intensity conditioning (ric) regimens for allogeneic stem cell transplantation (allo-sct) have emerged as an attractive modality to decrease allo-sct-related toxicities and nonrelapse mortality. indeed, ric allo-sct represents an attempt to harness the immune graft-versus-tumor effect while attempting to control or overcome toxicity. the work of different pioneering groups rapidly proved that this approach is feasible in several disease settings or patients' categories, and had the added benefi t of expanding the transplant option to patients who are ineligible for standard myeloablative allo-sct. currently, the use of ric allo-sct has challenged the need for high-dose conditioning regimens. unfortunately, and despite several thousands of patients receiving ric allo-sct, the true value of ric allo-sct in the management of hematological and non-hematological malignancies, especially aml is, as yet, diffi cult to delineate. currently, there are only very few, if any, prospective, randomized, or controlled trials that addressed the specifi c role of ric allo-sct versus other treatment strategies in aml. based on a large number of registry-based analyses, the ric subcommittee of the alwp has attempted over the last few years to better defi ne the role of ric allo-sct in aml patients through fi ne analysis of leukemia-free survival and overall survival balanced against treatment-related toxicity, complications, and death. a major advance was the identifi cation of the most important comorbidities that are likely to impact outcome after ric allo-sct for aml. another major achievement is the launch in of the multicenter/multinational ebmt randomized phase study evaluating the role of ric allo-sct in elderly patients with aml. at present, the use of ric prior to allo-sct appears to be on the cutting-edge. the ric approach proved to be much more complex than originally thought. since the fi rst reports published in the late s', the ric allo-sct literature has exponentially expanded. the complexity of ric allo-sct practice is progressively deciphered and the optimism to regard ric allo-sct as a potential and promising treatment modality for many aml patients remains very high among investigators, warranting continuous and renewed clinical and therapeutic research in this area. transfusion of donor lymphocytes (dlt) for treatment of leukaemia relapse after allogeneic stem cell transplantation (sct) has been a milestone in the fi eld of immunotherapy against malignant disease. it can be regarded as the proof of principle for the graft-versus-leukemia effect. during recent years, the immunotherapy subcommittee of alwp has initiated a variety of retrospective studies to evaluate in detail the role of unmanipulated dlt in acute leukaemia. in contrast to cml and early stages of mds, dlt was less effective in the treatment of post sct relapse in highly proliferative diseases as acute leukaemia. nevertheless, when given as maintenance therapy after effective cytoreduction, dlt could induce long term remissions in selected patients in aml, both after standard and reduced intensity conditioning transplants. strategies that included dlt were more effective than treatment without the use of donor cells. in contrast, no such effects could be shown in all. these data have prompted the strategy to use donor cells already before the occurrence of overt haematological relapse, i.e in minimal residual disease, mixed or falling donor chimerism, or even prophylactically. this strategy is now widely used across european transplant centres and has shown promising results. future strategies include the combination of targeted therapies and donor cell based strategies, as well as the use of specifi c subsets of donor cells in order to augment the antileukemic effi cacy and control for side effects of dlt, such as gvhd. allogeneic stem cell transplantation (allosct) with reduced intensity conditioning (ric) is being increasingly used for acute myelogenous leukemia (aml) patients with high comorbidities not eligible for standard myeloablative conditioning. new compounds and formulations including intravenous busulfan, treosulfan and recently clofarabine have been introduced into the pre transplant conditioning regimens in an attempt to reduce transplant related toxicities while keeping or increasing the anti leukemic effect, overcoming the increased relapse rate usually associated with ric allosct. similarly, replacing bm by peripheral blood stem cell (pbsc) grafts may increase the gvl, albeit with the price of increasing toxicities, gvhd and trm. thus, the optimal conditioning regimen for adults with aml is yet unknown. in order to address these issues, the alwp of the ebmt recently performed several surveys analyzing the use of iv bu for allosct, comparing iv bu/cy to tbi/cy, comparing pbsc and bm, and is about to compare treosulfan to iv bu as conditioning regimens for adult patients with aml. in the iv bu/cy vs. tbi/cy survey that included patients, the fi ndings indicated that outcomes of allosct including engraftment, trm, rr and lfs were comparable while gvhd probability was signifi cantly lower and vod probability signifi cantly higher in adults patients with aml using iv bu/cy vs. tbi/cy conditioning, respectively. in a separate survey analyzing the use of iv bu allosct for aml the year cumulative incidence of vod was found to be . + %. the factors associated with vod were mismatched unrelated donors and being not in remission. the pbsc vs. bm survey included patients. we observed signifi cantly higher incidence of gvhd and nrm and lower incidence of relapse rate with the use of pbsc, resulting in equivalent lfs. the treosulfan vs. iv bu survey is ongoing. in house comparison revealed similar lfs in aml patients in remission, while results were better using d iv bu combinations in patients with active disease. for mds the flu/treo conditioning regimen was the best. in conclusion, it is conceivable that introducing new compounds in the pre allosct conditioning regimens in combination with pre-and post-allosct targeted therapy will enable us to tailor the conditioning regimen to the specifi c disease category, reducing toxicity while improving the anti tumor activity. allogeneic hsct (allohsct) is generally considered the best consolidation option for younger patients diagnosed with an aml with high-risk cytogenetics. nonetheless, the cytogenetic high-risk category comprises different aml subtypes, and the specifi c results of allohsct for most of these cytogenetic entities are mostly unknown. moreover, several molecular lesions, such as mutations of flt , mll or wt genes, identify subsets of patients with a poor-prognosis disease despite harboring a non-adverse-risk cytogenetic abnormality. therefore, the precise role of allohsct for the management of high-risk aml should be determined after a comprehensive analysis of the outcome of transplant in all these biological aml varieties with an adverse prognosis. in this context, the molecular markers sc is conducting a project to analyze the results of allohsct for diverse high-risk aml subsets, such as aml with t( ; ), aml with q abnormalities, and normal karyotype aml with internal tandem duplication of flt gene (flt -itd three-year lfs was % and . % for patients in cr and pif, respectively. finally, the prognostic impact of flt -itd on the outcome of allohsct has been analyzed in a cohort of patients (median age: , - ) with a normal karyotype aml. donor was an hla-identical sibling in % of cases, whereas conditioning regimen was a myeloablative regimen in all transplants. all patients received transplant in cr . of note, -year lfs, ri, and nrm was . %, . %, and . %, respectively, without a signifi cant difference between transplants from related and unrelated donors. in summary, the outcome of allohsct in patients with two high-risk aml subtypes such as aml with t( ; ) and flt -itd aml showed a relatively favourable outcome, which compares favourably with reports from non-transplanted patients. on the contrary, the outcome of patients with aml and q rearrangement seems inferior, even in patients transplanted in an early phase of the disease. therefore, in order to elucidate the current role of allohsct for high-risk aml, transplant analyses should be focused on specifi c biological aml entities, since the results might differ largely among aml subtypes. finally, a careful collection of biological features at diagnosis is essential for addressing studies based on biological categories. hsct is a well-recognized option for the treatment of acute leukemia with the number of transplantations continuously growing over the last decades. however, the hsct rates vary strongly between countries. in particular, the activity in central and eastern (c&e) europe is markedly lower compared to the western part of the continent, suggesting the role of various socio-economic and geographic factors. using the human development index (hdi) as a surrogate marker we demonstrated that the socio-economic status (ses) of a country strongly correlates with the total number of hsct per population, as well as separately with sibling-hsct, unrelated-hsct and autologous-hsct in europe. we further speculated that in line with the transplant activity the results of hsct may also vary between countries or regions. direct comparison of hsct performed from sibling donors for aml in cr showed superimposable results obtained in c&e and western europe, however, the transplant characteristics differed with younger recipient age, longer interval from diagnosis to hsct and less frequent use of peripheral blood, tbi, t-depletion and reduced-intensity conditioning in c&e countries. subsequent analysis demonstrated that the outcome of sibling-hsct for acute leukemias in c&e europe improved over time. in particular the cumulative incidence of nrm decreased from % for patients treated between - to % for hsct performed between - , despite increasing recipient age. in another analysis we demonsatrated the infl uence of the ses on outcome in europe, which, however, could not be explained by regional differences. best outcome (increased lfs and reduced ri) after myeloablative hsct from either related or unrelated hla-matched donor was observed in countries with the highest hdi, while no differences could be demonstrated for the remaining analyzed countries. interestingly, transplantations in countries with the highest hdi were characterized by increased recipient age and shorter interval from diagnosis to hsct. finally, we speculated that the center experience may infl uence results of hsct. using transplants with reduced-intensity conditioning (ric-hsct) as a model we analyzed results according to the team experience as defi ned by: ) time from the fi rst allohsct and from the fi rst ric-hsct, ) the total number of allohsct and the number of ric-hsct performed in a study period ( ) ( ) ( ) ( ) ( ) ( ) ( ) . we found that results in centers performing few ric-hsct (< in years) were inferior compared to the remaining centers in terms of all, lfs, ri and nrm, which has been confi rmed in a multivariate model. altogether, in a series of retrospective analyses based on the ebmt alwp registry we demonstrated that both rates and outcomes of hsct for acute leukemia vary among countries and centers. both socio-economic factors and team experience infl uence results. we previously reported the ebmt experience with salvage high-dose chemotherapy (hdct) in pediatric patients with extragonadal germ-cell tumour (gct) (de giorgi u et al. -bjc ) . we analyzed a total of children with extragonadal gct, median age years (range - ), treated with salvage hdct with haematopoietic progenitor cell support. the gct primary location was intracranial site in nine cases, sacrococcyx in eight, retroperitoneum in four, and mediastinum in two. twenty-two patients had a nongerminomatous gct and one germinoma. nine patients received hdct in fi rst-and in second-or third-relapse situation. no toxic deaths occurred. overall, of patients ( %) achieved a complete remission. with a median follow-up of months (range - months), ( %) have been continuously disease-free. of six patients who had a disease recurrence after hdct, one achieved a diseasefree status with surgical resection followed by chemotherapy and radiotherapy. in total, patients ( %) were diseasefree at the time of analysis. eight of patients ( %) with extracranial primary and three of nine patients ( %) with intracranial primary gct were disease-free at the time of analysis. hdct induced impressive durable remissions as salvage treatment in children with extragonadal extracranial gcts. after years from this fi rst analysis (done in published in ), we decide to analyze the long-term results of this experience integrating these data with other from other cases from the ebmt registry previously not included, and with data from additional patients from italian registries (gitmo and aieop). high-dose chemotherapy and autologous stem cell transplantation in relapsed germ cell tumours: do we need a randomized study? p. pedrazzoli ( ) nasopharyngeal carcinoma (npc) is an epstein-barr virus (ebv)-related malignancy expressing a restricted set of viral antigens. the outcome of patients with npc failing conventional radio-chemotherapy is poor, the median overall survival of patients receiving second line treatments being less than two years. hence, the need for alternative therapies capable of improving disease-free survival and associated with reduced toxicity. since , we have implemented a t-cell therapy program for patients with npc failing conventional treatment. so far, we have treated patients with disease resistant/relapsing after > lines of radio-chemotherapy in two sequential trials of cell therapy with autologous ebv-specifi c cytotoxic t-lymphocytes (ctl). in detail, ebv ctl ( escalating doses to a maximum of × ctl/dose in the fi rst trial, or doses of - × ctl in the second trial) were administered in patients without or with preceding lymphoablative chemotherapy and recombinant human interleukin- (rhil- ). the proportion of patients achieving response (partial, pr, complete, cr) or stable disease (sd) lasting at least months (recist criteria), as well as immunologic correlates of effective treatment, were evaluated. overall, the objective control of disease was %, with / patients showing complete (n = ), partial (n = ) and minimal (n = ) responses, and / disease stabilization (median duration months). no severe adverse events were observed after cell therapy; patients showed an infl ammatory reaction at the tumor site. the use of preparative chemotherapy and increased ctl dose did not infl uence outcome. importantly, patients showing response to cell therapy showed the emergence of ebv lmp antigen-specifi c t-cells in their peripheral blood. ebv-specifi c ctl therapy is safe and associated with clinical benefi t in patients with advanced npc refractory to standard therapies. the use of ctl with higher specifi city for the ebv subdominant antigens expressed by the tumor, such as lmp , at an earlier stage of disease, could further implement the strategy and ameliorate the outcome of patients with relapsing/refractory npc. this study was supported by a grant from the italian association for cancer research (airc). , and csa given orally ( mg/kg/d). patients randomized to receive g-csf were given glycosylated g-csf from day to ( microg/m /d, sc). os at years was %, it was % for patients with severe aa and % for patients with very severe aa (p = . ). survival decreased with increasing age from % (< years) and % ( - years) to % ( - years) and % (> years) (p < . ). there was no difference, overall and when stratifi ed according to age and aa severity in os (p = . ) and efs, defi ned by death, need for transplantation, relapse and non response as events (p = . ) between the study arms at years ( table ). the median neutrophil count was signifi cantly higher between day and , in the g-csf group; this difference did not persist to day , when g-csf was stopped. there were fewer infections ( % no g-csf; % with g-csf; p = . ), and less days of hospitalization during the fi rst days (p = . ) in the g-csf group. patients died, mostly from infection ( %).there was no difference in death rates, cause of death and response rates between both groups. overall % of patients with and % without g-csf did respond to is (p = . ). patients did not respond to fi rst-line therapy, patients relapsed during the fi rst year of treatment (no difference between both groups). in conclusion, g-csf added to standard is increases neutrophil counts, and decreases rate of infections and days of hospitalization but has no impact on os, efs, remission, death and relapse rates. this has to be weighed against possibly higher risks of mds/aml, as suggested by previous studies. an hla-identical sibling ( %), pts received a myeloablative regimen ( %) and bone marrow ( %) with gvhd prophylaxis consisted in csa ± mtx in pts ( %). during evolution, pts did not engraft ( %). acute gvhd of grade >i developed in patients ( %). chronic gvhd developed in pts ( extensive). after a median ±se follow-up time of ± months, pts died ( infections, gvhd). the -year os rate was ± %. none of the common variables examined for association with os were statistically signifi cant, except for sct indications with an increasing risk from rsh, saa and thr ( figure , p = . ). based on the pairs obtained through the matching process, the -year os estimate after thr ( figure ) was ± % for pts with sct and ± % for pts without sct [hr . ( %ci . to . ); p = . ]. of note, the os among non-sct pts was signifi cantly worse (p = . ) in the non-matched than in the matched group, raising the question of pts over selection due to the matching process. concerning pts with saa complication, after exclusion of pts with thr and of non-sct pts with a follow-up time < months after saa, the -year os was ± % for pts without sct and ± % for pts with sct, but the groups differ signifi cantly for age at saa and year of saa. conclusions: further matching processes are necessary to conclude on this large cohort of pnh pts in order to defi ne the exact place of sct in pnh, especially in the era of the eculizumab. introduction: currently peripheral blood (pb) is more commonly used as stem cell source than bone marrow (bm) in both autologous and allogeneic hematopoietic stem cell transplantation (hsct) (gratwohl, bmt, ) . however, pb is associated with an increased risk of chronic graft-versus-host disease (gvhd), which is a disadvantage in non-malignant diseases. a recent study of ebmt/cibmtr showed that incidence of chronic gvhd and overall mortality were higher after hsct with pb than after hsct with bm in young patients with severe aplastic anemia (saa) (schrezenmeier h, blood, ) . the ebmt transplant activity survey shows that the number of pbsct has increased rapidly since early 's and exceeded the one of bmt in in family donor and in in unrelated donor (ud)-hscts. the number of pbsct has also increased in saa. we were therefore interested to review the current status of stem cell source selection in bone marrow failure ( in conclusion, pb has been increasingly used as a stem cell source in bmf despite of its higher risk of chronic gvhd. there were major differences in stem cell source distribution, regarding donor type, the global region as well as countries in regions. it may refl ect the differences in infrastructure in each center/country, donor and physicians preferences and policy of marrow donor program. clearly, recommendation for stem cell source is warranted in bmf. we conclude that q-syndrome can be cured by allogeneic transplantation, but additional abnormalities reduce the curability. the role of lenalidomide given before transplant will also be analysed. patients. an interim toxicity analysis was performed when the fi rst hundred patients had been included. the safety committee agreed to resume the trial. we will report the feasibility and the toxicity in both arms. an interim analysis is currently ongoing. other preliminary studies demonstrate that vtd is a highly active and relatively well tolerated regimen. the combination is used in the relapse setting, as well as fi rst line, consolidation and maintenance. in this protocol, the starting doses of velcade and thalidomide are relatively high and the duration of treatment is long. we will assess the superiority of vtd over td in the relapse setting as well as its toxicity. objectives: chronic lymphocytic leukemia (cll) is susceptible to well characterized graft-versus-leukemia effects. the leukemic clone is eradicated at the molecular level in > % of patients. yet, up to % of patients have persisting disease or relapse after allogeneic hematopoietic cell transplantation (hct). the application of donor lymphocyte infusions (dli) appears to be attractive in this situation. the aim of this retrospective analysis is to study the long-term effects of dli in patients with cll. methods: data from patients with cll who received allogeneic hct between and and had received at least one dli were included. the outcome of dli was analysed in two situations. ) "pre-emptive" dli prior to frank relapse or disease progression ) "therapeutic" dli given after documented relapse. baseline and follow-up data were downloaded from the ebmt database. results: patients fulfi lled the inclusion criteria. major characteristics were median age years, a median of three prior chemotherapy regimens, % matched sibling donor hct, % reduced intensity conditioning, % received peripheral blood stem cells. atg or alemtuzumab was used in %, ex vivo t-cell depletion (tcd) in %, no tcd in % of the patients. the median follow-up after the fi rst dli was months. in the cohort of patients who received dli prior to relapse, patients received more than one dli, patients more than two dli, patients more than three dli. grades ii to iv acute gvhd was reported in out of patients ( %) with informative data. pr or cr was documented in out of patients ( %). overall survival and progression-free survival years after the fi rst dli was % ( % ci, % to %) and % ( % ci, % to %) respectively in this group of patients. patients received dli as treatment of relapse. among these, patients received more than one dli, patients more than two dli, patients more than three dli. the overall response rate was % ( out of informative patients). nine out of patients ( %) experienced acute gvhd grades ii to iv. in this group the -year overall survival after the fi rst dli was % ( % ci, % to %). only patients had a follow up of more than years. conclusion: in patients with cll donor lymphocyte infusions appear to have only moderate long-lasting activity. further investigation to delineate factors associated with improved outcome is warranted. , centres %, centres . %, centres %, centres %. centres using % dmso washed cells prior to infusion although another centres also washed cells in occasional patients mostly less than % of their patients. centres added additional agents to the freezing mixture, mostly albumin ( ), hydroxy ethyl starch ( ), heparin ( ) and tissue culture medium ( ). the median amount of dmso given per centre per patient was g, although the upper limit set by the centre was often considerably higher. % of centres did not use any delay between bags of stem cells and the median duration of infusion was minutes. side effects were defi ned using nci criteria (version . ) and initially results were analysed by each centre. patients in centres who used washed cells or % dmso experienced less nausea and vomiting and 'severe other' effects, although hypotension and hypertension did not seem to be affected (p = . , . , . and . respectively). on a centre basis, the use of an upper limit to the amount of dmso which could be given ( g or the amount given ≤ g versus > g) did not result in any reduction of any of the groups of side effects. similarly when the amount of dmso given and the duration of dmso infusion were compared on a centre-basis, no signifi cant differences were found. further analysis will be undertaken using individual data where appropriate and the results presented. introduction: cmv infection and disease still remain serious and frequent complications after hsct. both morbidity and mortality have been reduced by prophylactic and pre emptive strategies based on biological tests and on more or less effective anti-viral drugs. however, in absence of comparative studies, there is no consensus for diagnosis, prophylaxis and treatment. then we conduct a survey to assess the current cmv management for patients less than years in ebmt centres. materials and methods: in december , a -item questionnaire about diagnostic tests, monitoring schedules, prophylactic and pre emptive strategies and treatment modalities was send to centres. defi nition of treatment failure, recourse to drug resistant cmv mutant research, practice of cell therapy and combined anti-cmv therapy were also tested. results: by the st december ' , we have received responses from countries. a second shipment will send in order to increase the number of responding centres. as expected, whatever the considered aspect i.e. monitoring, prophylactic or pre-emptive applied strategy, used drug in front or second and more line therapy etc. none centre report same procedure than another. all results will be presented and discussed during pdwp meeting in vienna. then, some proposals will be made regarding studies comparing results in different centres with "opposite" procedures. the fi nal goal of this work is to collect enough data to build consensual procedure. here we describe the results of the fi rst patients. all cases were confi rmed by pcr in nasopharyngeal or bronchoalveolar lavage samples. the fi rst case was diagnosed on july- . patients' characteristics are shown in table . eight cases were considered nosocomial infections. in sct patients, the median time from transplant to infl uenza diagnosis was days ( - ). s-oiv characteristics are given in table . the most frequent symptoms were: fever ( %), cough ( %), rhinorrhea ( %), odynophagia ( %), myalgia ( %), headache ( %) and dyspnea ( %). diarrhea was rare ( %). of those who presented with upper respiratory tract infection alone, ( %) had progression to pneumonia. % had pneumonia, with no difference between sct patients ( %) and non-sct patients ( %). fifteen out pneumonias in sct patients occurred in patients during the fi rst postransplant year or later but under immunosuppressive treatment. five patients were transferred to the intensive care unit (icu). four out of of the most severe cases (admitted to icu or fatal course) had lymphopenia (< / mm ) compared with % of those with less severe forms (p . ). compared with adults, children had more pneumonia ( % vs. %) although the difference was not statistically signifi cant (p . ). only one of the vaccinated patients against seasonal infl uenza were diagnosed with pneumonia compared with % who were not vaccinated (p . ). all patients except one were treated with oseltamivir for a median of days ( - ). zanamavir was added (in combination or sequentially) in cases. no major adverse events relating to anti-infl uenza treatment were reported. summary: the novel s-oiv is a serious disease in stc and oncohematological patients with a high incidence of pneumonia ( %) and signifi cant mortality ( %). diarrhea was not a frequent symptom. lymphopenia was linked to the most severe forms. vaccination for seasonal infl uenza might protect against the development of pneumonia. nevertheless, the severity of new pandemic infl uenza seems to be similar to seasonal infl uenza in this patient population. background: ebv-ptld (epstein-barr virus-related post-transplant lymphoproliferative disorder) is a rare but serious complication after hsct and number of patients at risk is increasing over time. available data do not refl ect general practice of diagnosis and treatment of this complication. objective: assessment of the current strategy of diagnosis and preemptive use of rituximab for ebv-ptld in ebmt transplant centers (tc). methods: in survey, data regarding ebv strategy from participating tc were registered on specifi c forms and centrally analyzed. responses from centers were excluded due to lack of specifi c strategy. results: regular monitoring for ebv after hsct is done by / ( %) tc, and in / ( %) tc is related to clinical situation. the monitoring is performed in all allohsct patients in / ( %) tc, while in remaining tc in following subgroups: mud ( / ), t-depletion in vitro ( / ), t-depletion in vivo ( / ), family mismatched ( / ), cord blood transplants ( / ), and in single centers in patients with saa, ebv mismatch, or after autohsct for autoimmune disorders. quantitative ebv-dna by pcr is performed in / ( %) centers. the assay is performed in: whole blood - / tc ( %), plasma - / ( %), lymphocytes - / ( %), serum - center. the test is repeated twice a week in / tc ( . %), once a week in / ( . %), once per weeks in / ( . %), once a month in / ( . %) and adjusted to risk in / ( . %) tc. the monitoring for ebv reactivation after hsct is performed for a period of: less than months in / ( %), months in / ( %), months in / ( %), months in / ( %) and adjusted to risk in / ( %). rituximab as a pre-emptive therapy for ebv-ptld is routinely administered in / ( %). the number of ebv-dna copies as an indicator for preemptive therapy with rituximab was given by tc, but varied between the centers, and were based also on symptoms and signs (table) . conclusions: ebv-ptld strategy exists in most of the responding centers. differential approach regarding indications for preemptive therapy is seen between centers: rituximab is administered as preemptive therapy in % of participating transplant centers. objectives: the aim of this study was to analyze the clinical risk factors, donor and recipient cytokine gene polymorphisms associated with cytomegalovirus (cmv) infection within days after allogeneic hematopoietic stem cell transplantation (allo-hsct). methods: we studied in a total of pairs of recipients and their donors, who underwent allo-hsct at our center. we analyzed single nucleotide polymorphisms (snps) in pro-and anti-infl ammatory cytokines genes, tumor necrosis factor ( . %of patients developed cmv positive antigenemia without disease, only patients developed cmv disease ( patients with pneumonia and patients with enteritis). there was a higher incidence of early cmv infection in transplantation with unrelated donors ( . % vs. . %, p = . ) and in recipients who were cmv seropositive before transplantation( . % vs. . %, p = . ). the recipient's tgf-beta - and + genotypes were signifi cantly associated with the incidence of early cmv infection in both the unrelated transplantation cohort and sibling transplantation cohort, but the infl uence of the donor's tgf-beta - and + genotypes was signifi cant only in the sibling transplantation cohort. multivariate analysis identifi ed two risk factors for early cmv infection: cmv seropositive recipients (rr: . , %ci: . - . , p = . ), recipients with tgf-beta + c allele (rr: . , %ci: . - . , p = . ). donors with tgf-beta - t allele was found to be less signifi cant factor (p = . ). conclusion: although cmv disease has been reduced in the era of antiviral prophylaxis and preemptive therapy, our fi ndings suggest the incidence of cmv infection remains high and provide the fi rst report of relationship between genetic background of donor and recipient to the risk of cmv infection in a chinese population. hsct. toll-like receptors (tlr) are essential components of the innate immune system. c h/hej mice that lack functional tlr did not develop cystitis after intravesical instillation of e. coli. individuals with the asp gly polymorphism of the tlr gene showed a diminished infl ammatory responsiveness to endotoxin. because of the requirement of tlr in infl ammatory response we hypothesized that tlr asp gly reduces the risk of bk virus-induced hc after hsct. children (median age, years) who underwent allogeneic bone marrow (n = ) or peripheral blood stem cell transplantation (n = ) in a single center and their respective donors were genotyped of tlr for rs (a g, asp gly) using taqman real-time polymerase chain reaction. the donor was hla-matched unrelated in % of transplants or hla-identical related in % of transplants. conditioning regimen was myeloablative in all cases. two forms of post-transplant immunosuppression predominated, cyclosporine a and methotrexate in % of transplants and cyclosporine a alone in % of transplants. the asp gly polymorphism was present in of the patients ( . %) and in of the donors ( . %). interestingly, we found a signifi cantly reduced incidence of bk virus-induced hc in patients with the asp gly polymorphism ( % vs. . %, p = . ). in addition, we observed a significantly reduced incidence of bk virus-induced hc in patients who were transplanted from a donor with this specifi c polymorphism ( . % vs. . %; p = . ). in ten of the donor-patient pairs the polymorphism was present in both individuals and no hc was observed. the occurrence of the tlr asp gly polymorphism, in either recipients or donors, had no signifi cant impact on acute and chromic gvhd, relapse rate, bacterial and fungal infectious complications, transplant related mortality, and overall survival. in conclusion, asp gly polymorphism of the tlr gene in the recipient and/or the donor is associated with a signifi cant decrease of bk virus-induced hc after hsct in childhood. this study provides the fi rst evidence that the innate immune system through tlr signaling pathway seems to play an important role in the pathogenesis of bk virus-associated hc after hsct. design and methods: one hundred-seventeen patients, median age ( - ) years, with various haematological malignancies transplanted between and entered the study. eighty-seven recipients negative for hbv surface antigen (hbsag), anti-hepatitis b core antigen antibodies (anti-hbc) and hbv-dna with hbsag negative donors were defi ned as at no risk of hbv reactivation whereas all the remaining patients were defi ned as at risk. in accordance with the italian national guidelines for immunocompromised hosts, patients at risk transplanted after received lamivudine to prevent hbv reactivation from conditioning up to - months after discontinuation of all immunosuppressive drugs whereas before no prophylaxis was given. results: patients at no risk did not experience hbv reactivation/ hepatitis. among patients at risk, hbsag negative recipients from hbsag positive donors ( / ), hbsag positive recipients from negative donors ( / ) and / anti-hbc positive were treated with lamivudine. none developed hepatitis b after a median follow-up of months ( - ). hepatitis developed in anti-hbc positive untreated patients conditioned with a reduced intensity regimen up to months after discontinuation of immunosuppression and in none of those on prophylaxis. conclusions: we observed: no risk of hepatitis b in recipients serologically negative for hbv, transplanted from hbsag neg donors; effi cacy of lamivudine in controlling hbv reactivation in both hbsag positive recipients and negative recipients from s hbsag positive donors; a signifi cant risk of hbv reactivation in hbsag negative/antihbc positive recipients and effi cacy of prophylactic lamivudine in this setting and the importance of prolonged prophylaxis even after the discontinuation of immunosuppressive drugs. republic -a single-centre experience p. hubacek ( , ) , d. boutolleau ( , ) , c. deback ( , ) , p. keslova ( ) despite the improvement of infection monitoring and antiviral treatments, cytomegalovirus (cmv) infections remain an important cause of morbidity and mortality in allogeneic hematopoietic stem cell transplant recipients (allohsct). aim was to investigate cmv resistance in paediatric and adult allohsct recipients and to clarify the cmv load in whole peripheral blood and target tissue in situation of cmv disease. between i/ and xii/ , we tested whole blood samples from adults (median /patient) and children (median /patient) after allohsct. additional biological non blood samples were tested too.samples were tested for cmv and albumin gene quantity by rq-pcr and results were normalized to human genome equivalents. ganciclovir as fi rst-line therapy was initiated when cmv load exceeded normalized viral copies (nvcs) and switched to foscarnet or cidofovir in case of none response.clinical resistance was suspected after weeks of unsuccessful well-conducted treatment.resistance was studied using ul and ul gene sequencing.in the patients deceased in consequence of cmv infection,dna quantity in the tissue was tested too. virostatic treatment was started in adults ( %) and ( %) children.clinical resistance was observed in adults and children.known genotypic cmv resistance was proved in of them ( . % of treated) detecting mutation c g, a v, a v, l s, l f, g s and del - in ul and del - , n k, v m, p s in ul ). natural polymorphisms and unknown phenotype changing were also detected in both genes. despite the virostatic treatment, symptomatic cmv disease was observed in adults and children. ten patients deceased in consequence of cmv infection,mainly combined with gvhd. typical pathological signs of cmv infection and antigen detection in the tissue were found in patient only.despite this median of cmv quantity in mainly affected lung tissue was nvcs (range - ) while median in the whole blood was only nvcs (range - ). quantifying of cmv dna is very useful in the treatment of cmv infection. unfortunately our observation suggests the limits in case of cmv disease. genotypic evidence of cmv resistance proved to be very useful in the improvement of therapeutic management of patients. further studies are required to ascertain the true nature of the novel mutations described within ul and ul same as compartmentation of cmv infection. supported by mz fnm , mstm . cmv reactivation often occurs in patients after allogeneic hsct. recently, pdgfr-alpha was suggested as the cellular receptor for human cmv (soroceanu et al., nature, ) . pdgfr-alpha binding and activation of a downstream pi kinase signaling pathway was essential for cmv propagation. many tkis, such as imatinib, effi ciently inhibit pdgfr-alpha at concentrations readily achievable in patients. the aim of this study was to retrospectively assess a possible link between cmv reactivation and tki therapy. . also, at years, the relapse incidence was not signifi cantly associated with the type of ric regimen: ± %, ± % in the tbi-based ric and chemobased ric groups respectively (p = . ). finally, nrm was also comparable between both groups ( ± % and ± % in the tbi-based ric and chemo-based ric groups respectively; p = . ). despite its retrospective nature, results from this large study suggest that ric allo-sct is a valid option for aml patients not eligible for standard allo-sct. the overall outcomes (lfs, nrm and relapse) appear not to be signifi cantly different between aml patients in cr receiving a low dose tbi-based ric allo-sct versus those receiving a chemotherapy-based ric allo-sct using an hla-identical sibling donor. patients with all who relapse after allogeneic hct have a grim prognosis and the optimal salvage therapy remains unknown. aim of this retrospective analysis was to report the outcome of relapsed all after allogeneic hct, to identify factors which have a prognostic signifi cance and to evaluate whether any relapse treatment strategy was associated with better outcome. for this study the ebmt database was searched for: ) adult patients with all aged > y at hct; ) transplanted in remission; ) with hematological relapse after an allogeneic hct; ) hla identical or mud hct performed from to ; ) suffi cient med-c data. adult pts (median age y, - ) met these eligibility criteria. diagnosis was % b-lineage all, % t-all, % ph + all and previous allograft ( % were hla identical and % mud) was performed in remission (cr %, cr %, cr %). the median interval from transplant to relapse was months ( . - ). post-relapse interventions were: no further treatment in % pts, % pts received chemotherapy at a median of d after relapse , % pts received dli at a median of d post-relapse ( - ) and % pts underwent a second transplant at a median of d after relapse ( - ). % of ph + all pts in addition received a tki inhibitor. overall, % pts reached a subsequent cr. after a median follow up of months ( - ), the estimated -, -and -year overall survival (os) was %, % and %. the table below shows the statistically signifi cant parameters associated with -year os. in the multivariate analysis, os was associated with disease status at hct (cr + vs. cr p = . , hr: . ), time to relapse after hct (>vs< median months, p< . , hr: . ) and number of peripheral blood blasts at relapse (>vs< %, p< . , hr: . ), but it was not signifi cantly infl uenced by cytogenetics (ph + vs. other), donor type (sibling vs. unrelated), stem cell source (bm vs. pb), type of conditioning (tbi vs. chemo) or type of treatment for relapse (dli or hct-based therapy vs. therapy without cell infusion). this study highlights the extremely poor outcome of all pts who relapse after allogeneic hct. better outcomes were found in patients transplanted in cr , with late relapse after hct (> months) and lower tumor burden at relapse (< % of pb blasts). these factors should be taken into consideration when discussing further therapeutic options for these patients. introduction: prognosis and survival in aml patients is associated primarily with cytogenetic abnormalities but also with molecular markers. the internal tandem duplication (itd) of the flt gene is one such marker associated with poor prognosis. we report here an analysis of the impact of allogeneic sct on clinical outcome of cytogenetically normal aml patients according to their flt -itd status. patients and methods: flt -itd mutation was analyzed by capillary electrophoresis after pcr amplifi cation of material frozen at the time of diagnosis from a total of / patients with normal karyotype entered between and into two osho studies (aml and aml ). sct was performed after conditioning with cytoxan and cgy tbi followed by gvhd prophylaxis with cyclosporine and methotrexate. related and unrelated donor search was initiated as soon as possible and patients allocated to groups with or without a donor. a total of patients were allocated to the donor group and patients to the no donor group. results were analyzed as intention to treat. results: event free survival (efs) of all patients was % after years. in patients (n = ) allocated to receive hct from an allogeneic donor, efs was % compared to % in those allocated to the non-transplant treatment (n = ). as previously described, detection of a flt -itd mutation had a negative impact on efs at years, which was % in flt -itd positive and % in flt -itd negative patients (p = . ). subsequently, efs was analyzed according to flt status and post-remission treatment. in the flt -itd positive group, efs was inferior in patients without donor ( % at years) compared to those with a donor ( % at years). this difference was due to a decreased relapse incidence in the donor group ( %) compared to the no-donor group ( %). however, a difference in efs was also observed in flt -itd negative patients in the donor ( % at years) compared to the nodonor group ( % at years). this difference was also associated with a reduced relapse incidence in the donor group ( %) versus the no-donor group ( %; p = . ). conclusion: in conclusion, allogeneic hct improves efs in aml patients with a normal karyotype by reducing relapse incidence irrespective of the flt -itd mutation status. the median follow-up of living patients was months (range, to months). at years the overall survival (os) was % and progression-free survival (pfs) was %, while the incidence of non-relapse mortality (nrm) and relapse were % and %. we performed multivariate cox regression modelling for os and pfs while competitive event statistics were applied for nrm. in order to adjust for a potential selection bias the hct-specifi c comorbidity score, karnofsky performance status, delay between diagnosis and hct and age were forced into the multivariate model. we performed a complete case analysis based on patients. the hazard ratios (hr) of a matched or a mismatched ud compared to a sibling donor were . ( % ci, . to . ) and . ( % ci, . to . ) for os and . ( % ci, . to . ) and . ( % ci, . to . ) for pfs. in interaction analyses we did not identify signifi cantly differing effects according to age and disease stage. of note, having a matched ud did not signifi cantly affect the risk of nrm (hr= . ; % ci, . to . ). our results confi rm the current practice in high risk aml to deliver allogeneic hct from matched ud and sib. an updated analysis will be presented at the meeting. ( ) ( this phase-iii randomized ebmt-intergroup trial studied the impact of a consolidating haematopoietic stem cell transplantation (autohsct) vs. wait and watch for patients with cll in binet stage a progressive, b or c, in cr, nodular pr or vgpr after fi rst or second line therapy. the primary objective was to show that autohsct increase the -year progression-free survival (pfs) by %. here we present a fi rst analysis based on % of expected follow-up forms. results: between november and july , patients were enrolled (sfgm-tc/fcllg n = , mrc n = , gcllsg n = , sakk n = , other ebmt centers n = ). there were % males and % females. binet stages were progressive a %, b %, c %; % were in cr, and % in very good or nodular pr. of note, sfgm-tc/fcllg included only patients in cr. eighty three percent of the patients were enrolled in st, and % in nd line treatment. patients were randomized between group (autohsct n = ) and group (observation n = ) after an induction treatment which was left at the discretion of the investigators. median pfs was . months ( . - ) in the observation group and . months ( - . ) in the autohsct group; the -year pfs was % and %, respectively (p< . ). accordingly, the -year relapse incidence was % vs. %; p< . . the cox modeling for randomization arm, binet stage, disease status, line of treatment, contributing group (country), and the interaction between randomization arm and contributing group confi rmed that autohsct signifi cantly improved pfs (hr . [ . - . ] p< . ). the benefi cial effect of autohsct was stable over all contributing groups. at years, the probability of os was % and % for autohsct and observation, respectively; p = . . signifi cant differences in terms of non-relapse death were not observed. in conclusion, in patients with cll in fi rst or second remission, consolidating autohsct reduces the risk of progression (pfs) by more than %, but has no effect on overall survival. further analyses on variables affecting the outcome are underway and results from a quality of life study on both groups are awaited. improved long-term outcome after highly purifi ed peripheral blood cd + cell transplantation from here, we report about the long-term follow-up results of a prospective phase ii study in patients after transplantation of highly purifi ed peripheral blood cd + stem cells from hla-identical sibling donors for cml in . cp. a total of pts with a median pretransplant ebmt risk score of (range - ) have been included in this study. all patients received a myeloablative conditioning regimen with tbi, cyclophosphamid with/or without thiotepa and atg, but without any prophylactic immunosuppression post transplantation. one patient received an unmanipulated graft due to poor cd + donor cell mobilization, while two patients were successfully retransplanted with an unmanipulated graft from the primary donor after secondary graft failure. of the pts, were eligible for the application of dli, but pts did not receive dli due to sustained molecular remission and complete chimerism. thirty-two pts ( %) received dli because of increasing bcr-abl transcript levels or hematologic relapse, and pts ( %) as programmed t-cell add-back. the median starting dose was . ( . - ) x cd + cells per kg with a median maximum dose . ( . - ) × cd + cells per kg. dli alone induced a lasting reduction of median bcr-abl transcript levels (bcr-abl/gapdh ratio) of more than log and the estimate of being in a complete molecular remission at years is % ± %. pts. ( %) did not respond to dli alone, but of these pts. attained a cytogenetic and molecular response by imatinib (n = ), dasatinib (n = ) and/or interferon treatment. four patients were retransplanted with an alternative donor. the cumulative risk of grades ii-iv acute gvhd is % ± % for all study pts, and the risk of chronic gvhd is % ± %, respectively. after a median follow-up period of months (range - ) for all patients, the cumulative -year survival estimate is , % ± %. including all therapies for molecular relapse after transplant, of the included patients ( %) were in molecular remission at the last time point of observation. causes of death (n = ) were disease progression, secondary malignancy, liver failure, septicemia, and systemic capillary leak syndrome in one patient each. in conclusion, the concept of highly purifi ed pb cd + cell transplantation in conjunction with adoptive dli is associated with a particularly low risk of non-relapse mortality and allows induction of lasting molecular disease control in the majority of . cp cml. allogeneic stem cell transplantation (asct) after reducedintensity conditioning (ric) has become a reasonable treatment option for patients with advanced myelofi brosis. the role of characteristic molecular genetic abnormalities such as jak v f on outcome of asct is not yet elucidated. in out of myelofi brosis patients with known jak v f mutation status who received asct after ric the impact of jak genotype, jak v f allele burden and clearance of mutation after asct was evaluated. a non-signifi cant higher treatment-related mortality ( % vs. %, p = . ) and relapse incidence ( % vs. %, p = . ) was noted in patients harbouring jak wild-type, resulting in a decreased overall survival in a multivariate analysis (hr . , p = . ). no signifi cant infl uence on outcome was noted for the mutated allele burden analyzed either as continous variable or after dividing in quartiles. achievement of jak v f negativity after asct was signifi cantly associated with a decreased incidence of relapse (hr . , p = . ). in a landmark analysis, patients who cleared jak mutation level in peripheral blood months after asct had a signifi cant lower risk of relapse ( % vs. %, p = . ). we conclude that jak v f mutated status but not allele frequency resulted in an improved survival and rapid clearance after allografting reduced the risk of relapse. early autologous stem cell transplantation in poor-risk chronic lymphocytic leukaemia. long-term results from the gcllsg cll trial focusing on incidence and type of secondary malignancies f. mcclanahan ( ) ( ), p. dreger ( ) ( introduction: as previously reported, early autosct as conducted in the cll protocol is a feasible therapy option for younger patients (pts). purpose of the present analysis was to study the impact of fish karyotype and ighv mutational status on long-term pfs and os and the incidence and type of secondary malignancies occurring in this trial. trial design and patients: the design of the protocol has been extensively described previously. from to , pts were registered. as cases had to be excluded due to screening failure (n = ), withdrawn consent (n = ) or other reasons (n = ), pts were eligible for the current analysis. male to female ratio was : and the median age at diagnosis was yrs (range - ). results: sct was performed in pts ( %) at a median time of months (range - ) after initial diagnosis, whereas pts did not proceed to sct (mobilization failure n = , disease progression n = , early death n = , pts preference n = , unknown reasons n = ). at a median follow-up of months (range - ), median os of all pts was . yrs ( . yrs after sct, . yrs without sct, hr . , % ci . - . , p<. ). median pfs was . yrs ( . yrs with sct and . yrs without, hr . , % ci . - . , p = . ). diagnostic samples for assessment of ighv mutational status were available for ( %). an unfavorable ighv rearrangement was present in pts ( %), and was associated with signifi cantly worse pfs (p = . ) and os (p = . ). fish was possible in pts. whereas pfs (p <. ) and os (p <. ) were considerably reduced in pts with del p-, there were no signifi cant differences between the other subsets. altogether, secondary malignancies were observed, with cases of t-mds/ t-aml, translating into a -year incidence of % ( % ci - %), with no signifi cant difference among pts treated with and without sct (p = . ). however, all cases of t-mds/ t-aml occurred after sct, yielding a -year incidence rate of % ( - %). overall survival after secondary neoplasms was months , with no difference between t-mds/aml and other malignancies. conclusions: unmutated ighv remains an adverse prognostic factor. while del p-is associated with a very poor outcome, autosct may eliminate the unfavorable impact of del q-seen with conventional therapy. secondary neoplasms are a serious problem after early autosct, but do not appear to occur more frequently than reported after sct for other diseases. reduced intensity conditioning (ric) has reduced non-relapse mortality (nrm) associated with allogeneic hsct in hodgkin lymphoma and relapse is now the commonest cause for treatment failure. however, there is little published evidence to guide management of relapse. we performed ric allografts for hl ( matched related, unrelated donor) incorporating t cell depletion with alemtuzumab. ( %) were primary or salvage-refractory, ( %) had failed a prior autograft, and the median number of prior treatment lines was . patients were monitored by pet-ct and relapse defi ned by recurrence or progression of fdg-avid lesions in sites of prior disease. if fdg-avidity occurred only in new sites, relapse was confi rmed by biopsy if accessible (n = ), otherwise an interval scan was performed at - weeks. following cyclosporine withdrawal and in the absence of gvhd, patients received dose-escalating dli for mixed chimerism (mc) or progression. the year ci of nrm was % and of relapse was %. patients received a total of dlis (median , range - ). treatment for mc alone (n = ) resulted in conversion to full donor in / evaluable patients, continuing improvement in , and remained stable. only of these relapsed (ci % at years from initial dli), receiving further dlis following chemotherapy. relapse was treated in patients with dli either alone (n = ) or following debulking chemotherapy (n = ). median maximal doses were × in unrelated and × cd + t cells/kg in related donor transplants. complete responses were seen in and partial responses in of evaluable patients (response rate %), and did not differ signifi cantly according to donor source ( / mrd vs. / ud). the majority of responders developed gvhd ( gd iii-iv acute, extensive chronic). cr are maintained at a median of . years from last dli ( without prior chemotherapy), died of gvhd-related complications, and progressed (at . and . years). / with pr progressed. the projected year os and pfs from relapse are % and % in this group. the os and current pfs for the entire cohort of patients are % and % at years. in conclusion, the data demonstrate favourable long term survival in this heavily pretreated hl cohort, a strikingly low relapse incidence in those receiving dli for mc, and the ability of dli guided by pet-ct to induce high response rates and durable salvage in the setting of relapse post t cell depleted transplantation. a recent update of consecutive prospective trials with high-dose therapy and autograft, without or with rituximab, as primary treatment for advanced-stage follicular lymphoma shows a sizeable group of patients surviving in continuous complete remission c. tarella, m. ladetto, f. benedetti, u. vitolo, a. pulsoni, c. patti, v. callea, a. rambaldi, a. piccin, l. devizzi, m. musso, e. iannitto, p. spedini, a.m. liberati, f. ciceri, a. gallamini, f. rodeghiero, g. gini, a. de crescenzo, f. di raimondo, a. levis, t. chisesi, t. perrone, d. rota scalabrini, g. rossi, a.m. carella, g. parvis, i. majolino, r. passera, m. ruella, a. pileri, a.m ) or in the proportion of patients with systemic symptoms or with elevated serum lactate dehydrogenase. the median time from the diagnosis to asct was also comparable as well as the number of previous treatment lines and the use of rituximab before asct ( % vs. %). the elderly patients were less often in fi rst complete remission at the time of asct ( % vs. %, p = . ). the most common conditioning regimen used was beam ( % in the elderly vs. %) followed by cyclophosphamide plus total body irradiation ( % vs. %). non-relapse mortality (nrm) was comparable at months ( . % in the elderly vs. . % in younger patients) and at year ( . % vs. . %) but was higher in the elderly patients at years ( . % vs. . %, p = . ). the most common causes of nrm were infections ( % vs. %). with a median follow-up of months for the elderly patients and months for the younger patients, the risk of relapse was % and % (n.s), respectively. progression-free survival was also comparable ( % vs. % at years) without any plateau. overall survival was worse in elderly patients ( % vs. % at years, p = . ). autologous stem cell transplantation is feasible in selected elderly patients with mcl. early nrm is low and comparable to younger patients. risk of relapse and progression-free survival are also comparable but overall survival is worse in the elderly patients. continuously increasing relapse risk indicates the need for improvements in pre-and posttransplant strategies in order to improve long-term outcome in this lymphoma type. phase ii study of radioimmunotherapy with yttrium- ( ) allogeneic hematopoietic cell transplantation (hct) using reduced intensity conditioning (ric) offers a potential curative therapy to patients with advanced indolent nhl. combined use of radioimmunotherapy (rit) with ric may increase anti-lymphoma activity of ric while avoiding additional toxicities. forty patients have been enrolled in a multicenter phase ii study of allogeneic hct using rit with yttrium- -ibritumomab tiuxetan (y -cd , zevalin) with . mci ( mbq)/kg on day - combined with ric using fl udarabine ( mg/m² day - to - ) and gy tbi (day ) followed by allogeneic hct from matched related or unrelated donors. gvhd-prophylaxis consisted of cyclosporine and mycophenolate mofetil. diagnoses were follicular lymphoma (fl, n = ), chronic lymphocytic leukemia (cll, n = ), mantle cell lymphoma (mcl, n = ), marginal zone lymphoma (n = ) and immunocytoma (n = ). median age was (range, - ) years. pbsc grafts were either from matched related (n = ) or matched unrelated donors (n = ). all patients were "high risk" with refractory disease or relapse after preceding autologous hct. engraftment was rapid and sustained with no graft rejections. median time to > granulocytes/μl was (range, - ) days, and to > platelets/μl (range, - ) days. in patients platelets were never < /μl and in patients granulocytes never < /μl, illustrating the nonmyeloablative intensity of the conditioning regimen. trm in the fi rst days was % (n = ) and overall % (n = ). no additional toxicity due to rit compared to our previous experience with the same ric as single modality was observed. incidence of grade ii-iv gvhd was % (ii = , iii = , iv = ). to date, chronic gvhd occurred in patients ( %, limited = , extensive = ). deaths occurred due to infections = , gvhd = , relapse = and cerebral bleeding = . / ( %) of all patients are alive with a median follow up of (range, - ) days, resulting in a kaplan-meier estimate year survival of % for all, and % in fl, % in cll and % for mcl patients. risk factors for decreased overall survival and trm in multivariate cox regression analysis were non-fl histology (p = . and p = . ) and agvhd >grade (p = . ). in conclusion, a combination of rit with ric is feasible with no additional toxicity due to rit and with stable engraftment in all patients. disease response and overall survival seems promising even in this elderly and heavily pretreated cohort of patients. allogeneic stem cell transplantation after autologous haematopoietic stem cell transplantation relapse in aggressive lymphoma patients: fi nal report of a retrospective gitmo study l. rigacci, a. bosi, b. puccini, p. corradini, l. castagna, n. cascavilla, g. milone, a. bacigalupo, r. scimé, g. specchia, a. rambaldi, p. leoni, f. ciceri, a. levis, s. guidi, b. bruno, r. oneto, r. fanin on behalf of gitmo autologous hematopoietic stem cell transplantation (ahsct) has been shown to be an effective therapy for patients (pts) with aggressive lymphoma. pts who relapse after an ahsct have a very poor prognosis. allogenic hematopoietic stem cell trasplantation (allohsct) has shown to be effective in the rescue of indolent lymphoma pts relapsed after conventional therapies. according to this data we have retrospectively analized all pts with diagnosis of aggressive lymphoma in the gitmo data-base who have performed an allohsct after an ahsct relapse. from to , pts were selected from the gitmo data-base. one of the principal objective in this fi nal report was the completeness of the data. data missing were: acute gvh was evaluable in % of pts, chronic gvh in % of pts, response after allogeneic transplantation in %, condition at the moment of allotransplant in %, therapy pre-allo-hsct in % of pts. the other characteristics were evaluable in all patients. one-hundred and one were male ( %), presented a diagnosis of dlbcl. the stem cell donor was related in pts, the stem cell source was peripheral blood in pts and bone marrow in . the conditioning regimen was conventional in pts and reduced intensity (ric) in pts. the median time between ahsct and allo-hsct was months. ninethy-seven pts ( %) obtained at least a partial remission before allo-hsct, ( %) were treated with active disease and in cases the data was not available. after allo pts ( %) obtained a cr and a pr with an orr of %, pts ( %) showed a rapid progression of disease. ninethy-seven pts died, due to disease and to treatment related mortality (trm). acute graft versus host disease was recorded in pts and a chronic one in pts. with a median follow-up period of months ( - ) from allo the os was % and after a median period of months ( - ) from allo the pfs was %. in multivariate analysis the only factors which affected pfs were the status at allo and a cr after allo transplant. this conclusive retrospective analysis confi rms that the only one parameter affecting either os or pfs was the response status at the moment of all-hsct and does not confi rm that ric could permit to obtain better results in aggressive lymphoma. probably a myeloablative conditioning should be reconsidered in pts with aggressive disease because of the slow-acting graft versus lymphoma effect is overriden by the rapid growth of the tumor. patients ( %), ii in ( %), iii in ( %), and iv in ( %). forty-nine percent of the patients had b symptoms, % bulky disease, and % extranodal involvement. karnofsky performance status < was reported in patients. treatment following relapse consisted on conventional chemotherapy and/ or radiotherapy in patients ( %), second asct in ( %), and allogeneic stem cell transplantation in ( %). a significant higher proportion of patients, age < years, received a second transplantation in comparison to older patients ( % vs. %, p =. ). after a median follow-up of months (range - ), the os from relapse after asct was of % at years and % at years. in multivariate analysis, independent risk factors for os were early relapse (< months), stage iv, bulky disease, poor performance status, and age > at relapse. in conclusion, according to the ebmt database, most hl patients relapsing after asct were treated with chemo-radiotherapy approaches and some of them with a second transplantation. those patients in a good performance status presenting with a localized late relapse seem to be the ones showing the best prognosis. hematopoietic cell transplantation (hct) is the therapy of choice for a variety of malignancies. hct provides disease benefi t through both the high-dose conditioning regimen and an allogeneic graft versus tumor effect (gvt). however graft-versus-host disease (gvhd) still remains a major obstacle. it has been proposed that host conditioning may not only be crucial in the activation of alloreactive t cells but also determine acute gvhd organ manifestation. we wanted to investigate how the host-conditioning regimen affects the host target tissues in terms of infl ammatory cytokines and their role in donor t cell recruitment. utilizing a non invasive bioluminescence imaging (bli), and fl ow cytometry in a murine allogeneic hematopoietic cell transplantation (allo-hct) model, we compared lethally irradiated ( gy) vs. non-irradiated balb/c wild type, and balb/c rag-/-cgc-/-(dko) (h- d) mice that received allogeneic luciferase + fvb/n t cells (h- q). we observed that the proliferation (bli, cfse), acquisition of activation markers (cd , cd , cd ) and homing receptors (a b , aeb , p-selectin ligand, e-selectin ligand) by alloreactive t cells occurred independently whether allogeneic recipients were conditioned or not conditioned before allo-hct. as t cell recruitment may have occurred as a result of alterations of the milieu infl ammatory cytokines in gvhd target tissues, we compared the cytokine profi le in conditioned vs. non-conditioned hosts. at days and after allo-hct, host tissues were analyzed for a th /th /th a cytokines from the target tissues; liver, large bowel, small bowel, peripheral blood and a non target tissue: kidney. at day , high levels of inf-g, tnf and il- were detected in the balb/c wt conditioned compared to the non-conditioned host in all target tissues and most markedly in blood and the large bowel. more importantly both the balb/c rag-/-cgc-/-(dko) conditioned and non-conditioned host displayed very high levels of the infl ammatory cytokines, with balb/c wt and balb/c rag-/-cgc-/-(dko) conditioned hosts displaying higher levels. similar results with a reduced expression of the cytokines were observed at day indicating that the cytokine storm peak was maybe at day . in summary host conditioning is not a requirement for alloreactive t cell activation rather induced infl ammatory cytokines such as tnf and inf-g are the determinant factors for effector t cell recruitment to gvhd target tissues. acute graft-versus-host disease (gvhd) after gender mismatched stem cell transplantation may be caused by female donor t cells recognizing hy minor histocompatibility antigens expressed by male recipients. hy-specifi c cd positive cytotoxic t lymphocytes (ctl) have been detected in peripheral blood samples collected at the onset of acute gvhd. however, it is still unclear whether these ctl reach acute gvhdaffected tissues. we validated the sensitivity and specifi city of fl uorescently labeled multimeric hla-a molecules containing hy peptide, i.e. hla-a /hy dextramers, to stain cryosections prepared from skin biopsies obtained from healthy male hla-a positive volunteer donors. these skin explant tissues were cultured with female hy-specifi c ctl or control ha- specifi c ctl before cryopreservation. unlike conventional hla-a /hy tetramers, hla-a /hy dextramers stained hy-specifi c ctl, but not ha- specifi c ctl, when applied to already cryopreserved skin explant tissue. control staining of serial sections with hla-a dextramers containing infl uenza peptide showed no positive signal. next, the presence of hla-a /hy dextramer positive cells was analyzed in cryopreserved skin biopsies derived from male hla-a positive pediatric patients who developed acute gvhd of the skin after receiving a hematopoietic stem cell graft from a hla matched unrelated donor. while a few cd positive hla-a /hy dextramer negative cells were detected in the skin of boys who received male hematopoietic stem cells, significantly higher numbers of cd positive cells were detected in the skin of boys who received a female graft; - % of these cd positive cells were specifi c for hy. skin-infi ltrating hyspecifi c t cells were visualized as early as weeks after sct when total peripheral blood lymphocyte counts were still low. our results underline the usefulness of this multimeric staining reagent for in situ characterization of donor-derived t cells that infi ltrate acute gvhd affected tissues. ( ) granulocyte-colony stimulating factor (g-csf) is increasingly described as an immuno-modulatory agent for a diverse range of diseases and although the cytokine is usually associated with the regulation of immune responses, clear evidence exists that it can also exacerbate pathology in some settings. the clinical shift toward utilizing g-csf mobilized stem cell grafts has resulted in enhanced hematopoietic reconstitution, reduced relapse rates in advanced disease, similar levels of acute gvhd but a striking increase in chronic gvhd. the mechanisms by which stem cell transplantation (sct) promotes chronic gvhd are unclear. comparison of cytokine expression following tcr activation of splenocytes from naïve and g-csf treated b or balb/c donors (low and high responders respectively) showed little effect of g-csf on th (ifn-gamma and tnf) or th (il- , il- and il- ) cytokine production. in contrast, il- production was dramatically enhanced in response to g-csf treatment in both strains but was highest in balb/c donors. the amplifi cation of il- production by g-csf occurred in both cd and cd conventional t cells and by using relevant knock-out mice or blocking reagents we demonstrated that this was independent of il- , tgf-beta or il- signalling. however, the induction of il- by g-csf was completely lost in the absence of il- signalling. g-csf induced the production of il- in cd t cells, and this occurred independently of il- production itself. we next utilized multiple models of gvhd using g-csf mobilized b or balb/c wild-type or il- deficient donors, in both mhc matched and mismatched transplant settings. surprisingly, il- was critical for the induction of sclerodermatous chronic gvhd occurring late after transplant using either donor strain. importantly, il- controlled the dramatic sequestration of macrophages into skin that coincided with the fi brogenic response. this study provides a logical explanation for the propensity of allogeneic stem cell transplantation to invoke sclerodermatous gvhd and suggests a therapeutic strategy for intervention. non-hla gene polymorphisms contribute to the immune response leading to graft-versus-host disease (gvhd). we applied a systematic approach using microsattelite (ms) marker typing for a large number of immune response genes on pooled dna of japanese donors and recipients of haematopoietic stem cell transplants (hsct) to identify recipient and donor risk loci for gvhd. ms, due to their multiple alleles, are more informative than single nucleotide polymorphisms (snp). we selected , ms markers, tagging , target genes (representing the 'immunogenome') at close proximity (< kb). we selected unrelated hsct donor/recipient pairs from the japan marrow donor programme (jmdp) registry, based on clinical homogeneity (acute leukaemia, age - years, myeloablative conditioning, bone marrow source). . % of pairs had a / hla match. the population was split into discovery and confi rmation cohorts with / pairs each. eight dna pools, four for each of the two independent screening steps were created using a highly accurate dna pooling method. while , ms were typed on the four pools of the st screening step, only markers positive here were typed on the nd screening pools. fisher's exact test for x (each ms allele) and xm chisquare tests were performed, comparing allele frequencies of recipients with gvhd grade - with gvhd grade - (donors accordingly). markers positive after both independent screening steps (p-value < . , same associated allele, consistent odd's ratio (or)) were genotyped for confi rmation on individual samples of all pairs. the independent, -step pooled dna screening process has effectively reduced false-positive associations. in the fi nal analysis, (recipient) and (donor) ms loci remain associated with risk or protection from gvhd. ( ), g. afram ( ) we have evaluated the impact of nih score system in the outcome of patients and studied additional prognostic factors among patients who develop cgvhd at european centers. furthermore, we tried to determine the prognostic impact of the different organs involved in order to defi ne which ones must always be evaluated and which ones could be avoided in a routine examination. patients' characteristics: patients transplanted from to at different institutions were analyzed; % received stem cells from a related donor and % from alternative donors. results: median follow up was days (range: to ). % and % developed overall and extensive cgvhd, respectively. according to nih classifi cation %, % and % of the patients were categorized as mild, moderate and severe cgvhd, respectively. type of onset was de novo in %, quiescent in % and progressive in %. cumulative incidence of delayed acute gvhd was % while the respective value for overlapping syndrome was %. among patients with cgvhd, the extent of cgvhd infl uenced on survival ( % vs. % for extensive vs. limited cgvhd, p< . ), as well as the development of overlapping syndrome (os % vs. % for patients who did or did not have overlapping syndrome, p = . ), of severe cgvhd ( %, % vs. % for patients with mild, moderate vs. severe cgvhd, p< . ), and the type of onset ( % vs. % vs. % for de novo, quiescent and progressive cgvhd, p< . ), while delayed acute gvhd did not infl uence on outcome. in multivariate analysis patients with severe cgvhd displayed a dismal outcome [hr = . , %ci ( . - . ); p = , ]. the type of onset allowed to identify prognostic subgroups even among patients with severe cgvhd, so that the combination of both variables discriminate different patients subpopulations in terms of outcome [hr = . , %ci ( . - . ); p = , ]. among targeted organs performance score at the time of cgvhd had the most signifi cant effect on survival [hr = . , %ci ( . - . ), p< . ]. also liver, gut and lung involvement adversely infl uenced on survival while eyes or mouth involvement had no infl uence on outcome. conclusion: nih scoring system plus cgvhd type of onset allows to defi ne patients subgroups in terms of survival. patients with overlapping syndrome have a poor outcome. among targeted organs, performance score was an independent prognostic factor while eyes and mouth involvement did not infl uence on outcome. chronic gvhd (cgvhd) remains the leading cause for late morbidity and mortality after allogeneic hematopoietic stem cell transplantation (hsct). the consensus conference summarized the current evidence on treatment of cgvhd and developed guidance for clinical practice. the consensus process included hsct centers within germany, austria, and switzerland participating in four meetings accompanied by two surveys on treatment of cgvhd send to all centers within the german and austrian working party on bone marrow and blood stem cell transplantation and the basel transplant center (switzerland). evidence was graded into: i (based on randomized trials), ii (based on case controlled or well designed phase ii trials), iii- (several case series), iii- (one case series), iii- (case reports) according to all available publications. recommendations were based on effi cacy and safety profi le: a (should always be applied), b (should generally be applied), c- (may be applied st-line), c- (may be applied nd-line), c (may be applied advanced line), c- (may only be applied in special circumstances). for st line treatment of cgvhd the following agents have been proposed: prednisone a i, calcineurin inhibitors (cni) c- ii, mmf c- iii. for nd-line treatment the following treatment options have been proposed: prednisone b iii- , cni c- iii- , mtor inhibitors c- iii- , mmf c- iii- , photopheresis b ii. a pulse of steroids c- iii- and rituximab c- iii- should be generally applied after nd-line treatment but may be used earlier in special circumstances. advanced line treatment options being proposed are: mtx c- iii- , hydroxychloroquine c- iii- , clofazimine c- iii- , pentostatine c- ii, thoracoabdominal irradiation c- iii- , and imatinib c- iii- . additional agents in advanced line treatment are retinoids c- iii- , azathioprine c- iii- , and thalidomide c- ii. other treatment options have been rated as experimental and may only be applied in clinical trials or special circumstances: alemtuzumab c- iii- , etanercept c- iii- , alefacept c- iii- . the low level of evidence of the proposed treatment options resulting in a low level of recommendation indicates the urgent need for further evaluation in clinical trials. moreover, the lack of indicators for response to certain agents requires a "trial and error" approach in choosing a treatment option and clinical trials for evaluation of biomarker for response are urgently needed. recently, we reported an elevation of immature cd + cd -b lymphocytes in patients with active cgvhd. here, we investigated b lymphocyte subsets in cgvhd cohorts with abnormal serum immunoglobulin levels. patients and methods: seventy-fi ve patients were enrolled into our study a median of months after hct. they consisted of groups: with cgvhd and high serum igg (> mg/dl), with cgvhd and low igg (< mg/dl) and patients with resolved cgvhd and normal igg (control group). severe cgvhd was present in % and % with more than organs involved in % and % in the high igg and low igg group. signifi cantly more patients in the high igg group than in the low one had autoantibodies ( % vs. %, p = . ). peripheral blood cells were analyzed by multiparameter fl ow cytometry after staining for cd , cd , cd , cd , cd , cd , and igd. results: while the high igg group had equal cd + b cell numbers, the low igg group had signifi cantly diminished ones compared to the control ( vs. vs. × /l p< . ). numbers of cd high most immature b cells in the high igg, low igg and control group were , , and × /l. immature cd + cd -b cell proportions were . %, . %, and . % in the high, low igg and control group (p = . ). in the high igg group transitional b cells were signifi cantly increased ( vs. × /l, p = . ) compared to the control. in the high igg group naive and class-switched memory b cells were equal to the control whereas in the low igg group both naïve ( vs. × /l p = . ), class-switched ( vs. × /l, p = . ) and non-class-switched b cells ( vs. × /l, p< . ) were signifi cantly lower compared to the control. cd -cd low cd high b cell proportions were the same in the high igg and control group, while they were signifi cantly elevated ( . % vs. . % p< . ) in the low igg group. conclusions: cgvhd patients with hypergammaglobulinemia have normal b cell numbers with elevated immature and transitional b cells but no signifi cant impairment of b cell maturation. in contrast, patients with hypogammaglobulinemia have both a signifi cant b cell defi ciency and a distortion of b cell homeostasis in the circulation. our data indicate different pathogenetic mechanisms of cgvhd leading to different clinical presentations. preliminary results of a phase ii trial of montelukast for the treatment of bronchiolitis obliterans syndrome after hsct k. williams ( ) bronchiolitis obliterans syndrome (bos) after allogeneic hsct is a serious manifestation of cgvhd. current treatments yield poor and transient responses. although the pathogenesis of bos after hsct is unknown, a similar disease, bos after lung transplant is associated with elevated leukotriene levels. we present preliminary results from an irb-approved prospective, open label, phase ii trial to test the effi cacy of montelukast, a leukotriene inhibitor, for the treatment of bos after hsct. bos diagnostic criteria included: fev < %, fev /vc < . or air trapping on ct and rv> % or rv/tlc> % in the absence of infection and presence of another cgvhd manifestation. subjects had stable or declining fev on stable or decreasing immunosuppression. eleven patients have enrolled. one withdrew prior to medication and / patients have reached the primary endpoint ( months) and have continued on study medication ( mg montelukast po nightly). study participants ranged from - years, / female, with baseline fev from to % predicted, and / patients required supplemental oxygen. fev increased - % predicted in patients, remained stable in , and declined less than % in . comparison of patient pre-study fev decline to on-study fev values was generated using the slope of fev volume vs. days post-transplant. the difference in pre-and primary endpoint slope revealed: / improvement and / decline. three patients reduced immunosuppression on study with complete cessation of tacrolimus in , cessation of steroids in , and decreased tacrolimus in (including with stable fev and with increase in fev ); patient had a steroid increase less than mg/kg/day. two patients had worsening of other cgvhd manifestations, including skin fl are that resolved with local therapy ( ) and gastrointestinal cgvhd fl are that improved with increased steroids and local therapy ( ) . notably, after months, patient demonstrated fev increase of % predicted (from baseline) on tacrolimus taper after steroid cessation and patient no longer required oxygen supplementation for exercise and sleep. montelukast was well-tolerated with only one grade ii attributable adverse event (insomnia) during the six-month collection period. using nih consensus criteria, improvements were also noted in / with buccal mucosa cgvhd and in / with evidence of liver disease. these fi ndings suggest that montelukast is a promising therapy for bos after allogeneic hsct. oral session : paediatric issues o objective: allogeneic hematopoietic stem cell transplantation (hsct) remains the main treatment option for children with advanced primary or secondary myelodysplastic syndrome (mds). relapse rate following hsct for advanced mds ranges between %- %. this analysis was performed to asses the outcome of patients treated with a second hsct (hsct ) for disease recurrence after hsct . patients and transplant: within studies ewog-mds and , patients with advanced mds relapsed after fi rst hsct (hsct ); a second allograft was performed in of these patients. for the patients with hsct , diagnosis prior to hsct was primary advanced mds (n = ), therapy-related mds (n = ) and secondary mds after bone marrow failure disorder (n = ). preparative regimen of hsct consisted of busulfan, cyclophosphamide and melphalan in / patients. median time to relapse was . months ( . - . ) after hsct , time from relapse to hsct . months ( . - . ), and median age at hsct . years ( . - . ). twelve patients were transplanted from a matched sibling donor, from a matched unrelated and from an alternative family donor. in cases hsct was performed using the same donor than in hsct . stem cell source was peripheral blood (n = ) or bone marrow (n = ). conditioning regimen and gvhd prophylaxis varied widely according to the centers preferences, a tbi-based regimen was given to / patients. outcome: median follow up was . years ( . - . ) after hsct . all but patients engrafted. acute gvhd grade ii -iv was seen in patients, chronic gvhd in of patients at risk. transplant related mortality occurred in patients; of these died from gvhd, from sudden cardiac arrest . years post hsct . a second relapse was noted in patients, all of whom succumbed to their disease. there was no signifi cant difference in relapse rate according to the choice of donor (identical or different donor than in hsct ) or the occurrence of acute/ chronic gvhd. for the whole cohort of patients, the probability of event free survival at years was . ( . - . ). conclusion: we conclude that hsct after mds relapse is feasible and may rescue a considerable proportion of patients. reduced-intensity conditioning for children with refractory cytopenia: results of the ewog-mds study b. strahm ( ) , p. bader ( ) objective: refractory cytopenia (rc) is the most common subtype of childhood myelodysplastic syndrome (mds). hematopoietic stem cell transplantation (hsct) in rc following a myeloablative preparative regimen has resulted in an eventfree survival of % with a very low probability of relapse and a probability of therapy related mortality of %. this analysis was performed to asses the outcome of children transplanted for rc following a reduced intensity conditioning. patients and transplant procedure: fifty-six patients ( male/ female) were diagnosed with rc at a median age of . years ( . - . ). none of the patients had an abnormal karyotype. the median time to hsct was . years ( . - . ). patients were grafted from a matched sibling donor (msd) (n = ), an alternative family donor (n = ) or an unrelated donor (ud) (n = ). ud were matched / or / based on hla-a, -b, -c, -drb and -dqb molecular typing. stem cell source was bone marrow (n = ) or peripheral blood (n = ). all patients were prepared with thiotepa ( × mg/kg) and fl udarabine ( × mg/m ). prophylaxis for graft-versus-host-disease (gvhd) was csa ± mtx, ± mmf for msd, and csa, mtx and anti-thymocyte globuline for patients transplanted from an ud. results: two patients each experienced primary or secondary graft failure. fifty-four patients reached neutrophil engraftment at a median time of days ( - ). platelet engraftment was demonstrated in patients at a median time of days ( - ) ; in patients an additional stem cell boost was necessary. out of the four patients with graft failure three patients are alive after second hsct. the incidence of acute gvhd grade ii-iv and grade iii-iv was % and %, respectively. ten of ( %) patients at risk developed chronic gvhd which was extensive in . one patient died of from veno-occlusive-disease with multi-organ-failure. viral infections were the most prevalent complication. fifty-four patients are alive with a median follow-up of . ( . - . ) years resulting in a probability of overall survival of . and . for patients transplanted from mud and msd, respectively. conclusion: in summary the conditioning regimen with thiotepa and fl udarabine offered an excellent survival for patients rc. chronic gvhd and viral infections were the main complications. long term follow is needed to assess the expected reduction in long term sequelae. allogeneic stem cell transplantation (sct) is indicated in approximately - % of children with acute lymphoblastic leukemia (all). unrelated umbilical cord blood (ucb) is an established stem cell source for sct. we retrospectively analyzed children with all in complete remission (cr) (n = ), cr (n = ) and cr or advanced disease (n = ) who received ucbt as fi rst transplant. patients were transplanted in ebmt centers from - . median age was . years (y) ( patients less than y). the most common immunophenotype was b-cell precursor all and patients had biphenotypic all. of patients transplanted in cr , % had poor risk cytogenetics (t ; or t ; ). grafts consisted of one (n = ) or two (n = ) units; % had - hla mismatches with recipients while % had - mismatches (antigen level for hla-a and b, allelic level for drb ). median tnc cell dose at freezing and infusion was . × /kg and × /kg, respectively. conditioning regimen was myeloablative in % of cases and tbi > gy was used in %. other regimens included busulphan with cyclophosphamide ± thiotepa or melphalan. atg was added in % of cases and gvhd prophylaxis was csa ± steroids in %. median follow-up was . months ( - ). cumulative incidence (ci) of neutrophil recovery, acute gvhd and trm were ± %, ± % and ± % respectively. in multivariate analysis tnc infused > × /kg (p = . ) and remission status at ucbt (p = . ) were associated with improved neutrophil recovery. ci of y relapse was ± % ( % for cr , % for cr , % for advanced). disease status at ucbt (hr = . , p . ) and use of tbi> gy (hr = . , p = . ) were independently associated with lower ci of relapse. y probability of leukemia-free-survival (lfs) was ± % ( % for cr , % for cr , % for advanced; p = . ). in multivariate analysis, disease status at ucbt was the only factor associated with improved lfs (hr = . , p = . ). causes of death were infections or other transplantrelated events (n = ) or disease progression (n = ). in conclusion, in the absence of an hla identical donor, ucbt remains a valuable alternative option for children with high risk all. disease status at transplant and cell dose are the most important risk factors for outcomes. use of tbi in the conditioning regimen was associated with decreased incidence of relapse but was not associated with improved lfs. the role of minimal residual disease as a predictor of outcomes of ucbt is under investigation. treosulfan-based conditioning in children: retrospective analysis of the german and austrian experience r. beier ( ) background: treosulfan (treo) is an alkylating agent with good myeloablative activity and a favourable toxicity profi le. it has been widely used in hematopoietic stem cell transplantations (hct) of high-risk adult patients. here we report on our retrospective analysis of children treated in germany and austria. patients and methods: all pediatric transplant centers in germany and austria were asked to submit data on treo conditioning. nine centers reported a total of transplantations, autologous and allogeneic. results: the median age at transplantation was , (range , - , ) years (n = ). underlying diseases were malignancies (n = : aml, all, mds, ewing sarcoma, neuroblastoma, rhabdomyosarcoma, b-all, lgl, biphenotypic leukemia, nasopharynx carcinoma, alcl, nephroblastoma), immunodefi ciencies (n = : scid, cgd, unclassifi ed, hyper-igm, interleukin- receptor defi ciency, hlh, was, cvid, mhc class ii defi ciency), hematologic diseases (n = : thalassemia, camt, osteopetrosis, sds, sickle cell disease, saa, dkc, scn), and metabolic disorders (n = : mld, mannosidosis). donors were matched sibling or matched related (n = ), matched unrelated (n = ), autologous (n = ), mismatched unrelated (n = ) or haploidentical family donors (n = ). two patients received a combination of a cord and a haploidentical graft. treosulfan was given at a total dose of g/m (n = ), g/m (n = ), and g/m (n = ). additional conditioning drugs included fl udarabine (n = ), thiotepa (n = ), melphalan (n = ), cyclophophamide (n = ), or tbi (n = , gy). atg (n = ), alemtuzumab (n = ), okt- (n = ), and a combination of atg and okt (n = ) were used together with csa ± mtx or mmf as gvhd-prophylaxis. eight patients experienced non relapse mortality, six of them had heavily pretreated malignancies. one engraftment failure (thalassemiahaplo) and three rejections ( immunodefi ciency, thalassemia, scn) were reported. all other patients engrafted rapidly. toxicity and gvhd grades in these patients were mostly i and ii. event free survival after years (data available for patients) was % for non-malignant and % for malignant diseases. conclusion: treo based conditioning regimens in children were effective in terms of engraftment and survival. optimal treo dosing for children is still not well defi ned. pharmakokinetic studies in children are needed. objectives: hepatic veno-occlusive disease (vod) is a severe complication following hematopoietic stem cell transplantation (hsct), with a mortality rate of %- %. the diagnosis of vod is based on clinical criteria. however, the late onset of clinical signs may delay the due treatment. studies in adults have addressed to plasminogen activator inhibitor- (pai- ) a possible role as marker of vod. our aim was to prospectively evaluate the role of fi brinolytic parameters to discriminate vod from other liver disorders occurring after hsct in a pediatric population. methods: we studied children (males , females , mean age , ± , years) who underwent hsct performed at the pediatric hematology oncology of padua university. the prevalence of vod was recorded. in hsct the levels of alanine amino-transferase (alt), total bilirubin, pai- antigen (pai- :ag) and activity (pai- :act), t-pa antigen (t-pa:ag), d-dimer, pt, aptt, antithrombin, fi brinogen and platelet count were assayed before and weekly for month after hsct. results: the prevalence of vod was . % ( / hsct), and it was signifi cantly higher in patients with pre-existing risk factors for vod as compared with those without ( . % vs. . %; p< . ). all but one patient who developed vod showed an early increase in pai- :ag, pai- :act, t-pa:ag and d-dimer levels, even before the clinical diagnosis of vod. the increase in fi brinolytic parameters, and in particular pai- :ag, was statistically signifi cant in comparison with that observed both in patients without complications (pai- :ag p< , ) and in those with non-vod liver disorders (pai- :ag p< . ). similar fi ndings were also seen when vod patients were compared with subgroups of children with infections and/or different types of hepatic diseases complicating hsct. conclusions: our study demonstrates a role of fi brinolytic tests in the diagnosis of vod after hsct in the pediatric population. in particular, the early rise in pai- antigen levels may suggest an incoming vod, even in the absence of clinical signs. in addition, pai- antigen increase may help to discriminate vod from other hepatic complications after hsct. late onset non infectious pulmonary complications (lonipc) after allogeneic hematopoietic stem cell transplantation (hsct) such as chronic graft-versus-host disease (cgvhd) of the lung, bronchiolitis obliterans (bo), bronchiolitis obliterans organizing pneumonia (boop), sinu-bronchial syndrome and restrictive lung disease are well characterized. immunodefi ciency, with or without cgvhd, and recurrent respiratory tract infections may be a vicious circle which requires optimal supportive therapy. mucociliary dysfunction is described in chronic pediatric lung diseases like cystic fi brosis (cf) and primary ciliary diskinesia (pcd). treatment with dornase alpha as a mucolytic drug with anti-infl ammatory effect improves morbidity and mortality rate in children with various chronic lung diseases. we hypothesized that outcome of our patients with lonipc measured by mortality, lung function, inhalative therapy and systemic immunosuppressive treatment could be infl uenced by dornase alpha inhalations i.e. once daily. we analyzed children with lonipc who have received an allogeneic hsct between to for malignant (n = ) and non malignant (n = ) diseases. we compared children (study group) obtaining dornase alpha inhalations with children without dornase alpha. characteristics of both groups are shown in table . additional supportive care was similar in both groups. dornase alpha inhalations were well tolerated with no side effects and high compliance. children treated with dornase alpha had a lower mortality rate ( %) than in the control group ( %) and showed in the long-term follow-up an improved lung function, predominantly of the peripheral airways (mean expiratory fl ow (mef) after %, % and % of forced ventilation capacity), in the forced expiratory value after second (fev ), in resistance (r eff) and in the intrathoracic gas volume (itgv) compared to the control group. treatment with dornase alpha reduced signifi cantly the inhalation frequency of salbutamol, fl uticason or tiotropium after , and months ( ± times per day vs. ± , p < . ; vs. ± , p < . ; vs. ± , p < . ) compared to the control group. immunosuppressive medication was similar in both groups. our data showed that inhalation with dornase alpha in children with lonipc is a well tolerated treatment option which may improve mortality rate and lung function. additional inhalative therapy but not systemic immunosuppression could be reduced. in order to fi nd hallmark of protection we studied the functional composition and magnitude of cmv-specifi c t cell response in cmv-seropositive (cmvpos) and cmv-seronegative (cmvneg) healthy donors and compared it to children after hct. polychromatic fl ow cytometry was used for detection of cmvspecifi c immune responses. the ability of cd + and cd + tcells to produce cytokines: interferon-ã (ifng) and interleukin- (il- ) and to express activation marker cd l and/or to mobilize the degranulation marker cd a in response to cmv antigens was evaluated by intracellular cytokine staining method after in vitro stimulation. peripheral blood samples were collected from patients who underwent allogeneic sct and healthy donors. monitoring of viral load was performed weekly from day + using quantitative rt-pcr. monitoring of cmvspecifi c t-cells begun on day + and was performed weekly during the time of hospitalization and on day + , + , + and + after sct. we have compared functional signatures in patients controlling their pcr documented reactivations (controllers) and patients non-controlling reactivations (non-controllers). as a reference we have included healthy cmvpos and cmvneg donors. among cd + t-cells we found three functional signatures (ifng + , ifng + /il- + and ifng + /il- + /cd l + ) that signifi cantly differed between controllers and non-controllers. whereas single ifng + cd + t-cells were the most frequent of cmv-specifi c t-cells, they were not restricted to cmv controlling individuals. we found signifi cant correlation between number of weeks with low viral load reactivations and frequency of ifng + cd + tcells (r = . , p< . ). dual ifng + /il + cd + t-cells were present in majority of controllers, but they were virtually absent in non-controllers. triple ifng + /il- + /cd l + positive cd + t-cells represented a rare subset of novel cd + helper phenotype. no cd + t-cell functional signature resolved controllers and non-controllers. we ( ) ( aberrant dna methylation contributes to the malignant phenotype in cancer including myeloproliferative neoplasms and myeloid leukemia. we studied aberrant dna methylation in candidate loci in blood and bone marrow samples of children with juvenile myelomonocytic leukemia (jmml) and asked whether it is associated with clinical, hematologic or prognostic features of the disease. the pattern of hypermethylation allowed the categorization of jmml cases into several groups. high methylation was strongly associated with higher age and increased hemoglobin f level at diagnosis. importantly, hypermethylation at diagnosis characterized cases with signifi cantly increased probability of leukemia relapse after hsct: the year relapse incidence was . ( . - . ) in the no-methylation group but . ( . - . ) in high-methylation cases (p< . ). the predictive power of high methylation for outcome following hsct was also refl ected in a multivariate cox model which included age at diagnosis, sex, platelet count and mutational category (ras, ptpn , cbl mutation or clinical diagnosis of nf ); here methylation was the only signifi cant prognostic factor (p = . ). future guidelines for intensity of graft-versushost prophylaxis in jmml will have to take the methylation status at diagnosis into account. ecfcs have recently been described as vascular progenitor cells with robust proliferative potential and vessel-forming capacity. vascular integrity depends on endothelial and pericyte functions. this study was performed to establish ideal conditions for the generation of stable vessels by ecfc/msccotransplantation. mscs were propagated as previously described. ecfcs were isolated with a novel recovery strategy and propagated under animal protein-free culture conditions with pooled human platelet lysate (phpl) replacing fetal bovine serum (fbs). ecfc long-term proliferation potential was monitored and phenotype was analyzed by fl ow-cytometry and immune-cytochemistry. genomic stability was assayed with chromosome g-banding and array-comparative genomic hybridization (array-cgh). additionally we compared telomere-length and telomerase-activity of ecfcs at different time points during culture with fl ow-fl uorescence in situ hybridization (flow-fish) and telomere repeat amplifi cation protocol-assay (trap). functionality was studied during vascular network assembly in vitro and in human vessel formation in immune-defi cient mice in vivo. a mean of four ecfc colonies/ml of peripheral blood could be recovered. the progeny of these cultures could be expanded to mean . ± . × ecfcs within - days. consecutive analysis confi rmed ecfc purity, immune phenotype and sustained proliferation potential for > population doublings with preserved progenitor hierarchy. genomic stability was confi rmed by karyotyping and array-cgh. telomere-length analysis revealed longer telomeres in cord blood compared to peripheral blood-derived ecfcs and a constant shorting of chromosomal ends through passaging, which could be further confi rmed by a lack of telomerase activity. large-scale expanded ecfcs functioned to form complex vascular networks in vitro and assembled stable cd + /vimentin + /von willebrand factor + human vessels when transplanted together with msc in vivo under defi ned conditions. these human vessels were connected to the mouse circulation as indicated by a rich content of ter + murine erythrocytes. this demonstrates that functional and stable human vessels can be generated in vivo as result of ecfc/msc-cotransplantation. this procedure represents a promising tool to develop innovative experimental, diagnostic and therapeutic strategies. hematopoietic progenitor cells (hpcs) which circulate in the blood are increasingly recognized as co-regulators of the development of tumor solid tumors through their ability to support neovessel formation and the transduction of the hpcs with suicide genes offers a new modality targeted antitumor cellular therapy. further to clarify mechanisms of their tumor homing, we investigated the role of the small gtpases rac and expressed in hematopoietic cells during tumor development in mice. mice with an inducible deletion of the rac gene in hematopoetic cells and/or constitutive deletion in rac were used. rac deletion was induced by polyic treatment in rac- fl ox/fl ox mice harbouring a hematopoietic-specifi c inducible cre recombinase. mice had previously been inoculated with lewis lung carcinoma by s.c. injection. we found that in mice with hematopoietic specifi c deletion rac genes, llc tumors grew more slowly in the absence of rac genes and compared to wild type (wt) controls. at the same time, numbers of mononuclear cells and hpcs increased in the blood. mobilization was further increased in tumor bearing mice. in contrast, the content of cd + cells in tumors of mice with an induced defi ciency of rac and in hpcs was greatly diminished as shown by immunohistochemistry. we then performed competitive homing experiments by co-injecting green and red fl uorescence-stained wt and rac / -/-hpcs in tumor bearing mice. this revealed a - % decrease in numbers of rac / -/-hpcs homing to tumors compared with controls. next, we studied in vitro adhesion in parallel plate fl ow chambers to delineate specifi c adhesion defi ciencies of the rac deleted hpcs. we found decreased arrest and persitstent adhesion of rac / -/-hpcs on endothelial cells on sdf- a precoated endothelial cells. this correlated with a decreased ability of these cells to migrate through gradients fo sdf- a in transwells. moreover, analysis of fi rm adhesion on both, vascular cell adhesion molecule- and intercellular cell adhesion molecule- , either alone or in the presence of sdf- a revealed a critical role of rac and genes in adhesion strengthening of hpcs. in conclusion, rac and negatively regulate mobilization of hpcs in tumor bearing mice, but are critically involved in entry of hpc into tumor tissue in a process involving sdf- a. modulation of rac activation in progenitor cells may thus be a tool to modify tumor growth and to target suicide gene delivery to tumor sites. the however, hoxa , hoxb , hoxc and hoxd were defi ned as good candidate markers to discriminate cb-msc and bm-msc from ussc. thus, our data suggest that the "biological fi ngerprint" based on the hox code can be used to distinguish functionally distinct stem cell populations from bone marrow and cord blood. this work was supported by a grant from the dfg ko / - . combined action of endothelial and mesenchymal niche cells to amplify haematopoietic progenitor expansion in a humanized system d. thaler ( ) the hematopoietic stem/progenitor cell (hspc) niche is an anatomically confi ned space governing hspc proliferation, differentiation and self renewal. recent research identifi ed distinct compartments described as stromal and vascular niches. this study was initiated to directly compare mesenchymal stromal cell (msc) and endothelial colony-forming progenitor cell (ecfc) contribution to niche functions in a humanized co-culture system. mscs and ecfcs were propagated under animal protein-free conditions. autologous msc-ecfc pairs were established to avoid donor variation. purifi ed cd + hspcs were used as responders in a cytokine-driven (il- , ng/ml; flt- -l, ng/ml; scf, ng/ml) niche cell-regulated co-culture system. different standard media were employed to compare serum-free conditions with humanized cultures supplemented with pooled human platelet lysate (phpl). primary expansion culture of hspcs with and without niche cell support was followed by colony forming cell (cfc) assays to determine maintenance of clonogenicity. liquid cultures supplemented with % phpl were more efficient than serum-free cultures resulting in a mean - -fold increase in nucleated cell progeny within days. mscs or ecfcs further amplifi ed hspc proliferation. interestingly, the supportive effect of mscs or ecfcs was constantly more pronounced in humanized phpl-supplemented compared to serum-free cultures, indicating an important role of the natural platelet-derived growth factors. the combination of mscs + ecfcs resulted in hspc proliferation with up to -fold nc increase. the hspc progeny was mainly comprised of differentiating myeloid cells. cfc assays revealed that phpl-supplemented liquid cultures were more effi cient than serum-free cultures resulting in a . - . fold cfc expansion. both msc and ecfc initiated a more than -fold increase of cfcs, whereas combined action of mscs + ecfcs resulted in a maximum of . -fold cfc increase in phpl-supplemented cultures. the impact of human ecfc as compared to msc cotransplantation on human hspc engraftment and reconstitution in immune defi cient mice is currently under investigation. our data indicate that mscs and ecfcs are effi cient in supporting hspc proliferation and cfc amplifi cation in a humanized co-culture system. the combination of both niche cell types had no further additive effect. the humanized co-culture thus provides a novel model system for analyses of hspc-niche cell interactions. a. reinisch ( ), n.a. hofmann ( ) , a. ortner ( ) , n. etchart ( ), c. , c. dullin ( ) , c. diwoky ( ) osteosarcoma (os) and ewing's family tumors (eft) are the most frequent bone sarcomas in young adults. unresectable or metastatic presentations are currently characterized by an extremely severe prognosis, with a consequent great need for new therapeutic approaches. we conducted a preclinical study to investigate the potential effi cacy of cytokine-induced killer (cik) cells as adoptive immunotherapy for bone sarcomas. cik cells are a heterogeneous subset of ex-vivo expanded t lymphocytes presenting a mixed t-nk phenotype and endowed with a mhc-unrestricted antitumor activity. cik cells can be used both as autologous adoptive immunotherapy or even infused from a donor after allogeneic hct. we successfully generated cik cells from patients with metastatic bone sarcomas ( os; eft). ciks were generated from pbmc, cultured for - weeks with the timed addition of ifn-a; ab anti-cd and il . at the end of the culture we obtained a heterogeneous t cell population with a median of cd + cd + cells of % ( - ), % ( - ) were cd + and presented a high expression (> %) of nkg d, the receptor mediating most of ciks' antitumor activity. the median ex-vivo expansion, calculated on the cd + cd + fraction, was fold ( - ). cytotoxicity experiments (n = ) demonstrated that cik cells effi ciently killed different os cell lines ( %, %, % and % of specifi c killing at a : , : , : and : effector/target ratio respectively). more striking, in selected experiments (n = ), we could confi rm the potent killing activity of cik cells against the autologous os tumor (figure ). the test was performed by staining target cells with cfse and co-culturing them with ciks at different ratios; after hours data were analyzed by fl owcytometry. we confi rmed that both cell lines and autologous os cells presented a high expression of mic a/b, main ligands for the nkg d receptor of cik cells. the tumor-specifi c cytotoxicity of cik cells was confi rmed by the absence of any signifi cant killing against non-tumoral mhc-mismatched targets. our data are the fi rst report of antitumor activity of cik cells against autologous os cells. our fi ndings are encouraging and support the designing of adoptive immunotherapy clinical trials with autologous cik cells for os patients. the observed safety across major hla-barriers suggests that also donor-derived cik cells might be considered in peculiar experimental clinical settings exploring allogeneic hct for solid tumors. m. nonn, j. knapstein, a. brunk, d. tomsitz, s.a. khan, e. distler, m. theobald, w. herr, u.f. hartwig johannes gutenberg university (mainz, de) donor lymphocyte graft engineering to separate graft-versushost (gvh) and graft-versus-leukemia (gvl) immunity is of central interest in allogeneic hematopoietic stem cell transplantation. therefore, we established a xenograft-transplantation model using nod/scid/il r gamma chain null (nsg) mice to evaluate engraftment, residual gvh-reactivity and gvl-immunity of human leukemia reactive cd + t cell lines in vivo. previous studies demonstrated engraftment of resting or polyclonally stimulated human cd + t lymphocytes to high levels in spleen and bone marrow following adoptive transfer into nsg mice and induction of xenogeneic gvhd (x-gvhd) within - weeks depending on the applied t cell dose. the engrafted human t cells contained both cd and cd subsets in the same ratio as analyzed prior to transfer. further transfer experiments using isolated human cd + and cd + t cell subpopulations revealed long-term engraftment of cd + t cells and induction of xenoreactivity. cd + t cells isolated ex vivo from spleen from these mice demonstrated an activated and memory phenotype suggesting a xenoantigen driven response in vivo. in contrast, adoptively transferred cd + t cells did neither engraft nor induce x-gvhd in nsg recipients. similar results were obtained upon transfer of several leukemia-reactive cd + t cell lines generated by short-term mixed leukemia lymphocyte coculture of cd + donor lymphocytes and acute myeloid leukemia (aml) blasts. investigating different cd + t cell growth and differentiation promoting cytokines we found that administration of human interleukin (il)- or il- failed to improve cd + t cell survival and expansion in vivo, whereas co-transfer of % autologous, naïve cd + t helper cells or repetitive injections of human il- resulted in robust cd + t cell engraftment in nsg recipients. in addition to these data, fi rst results of studies obtained in a therapy model to investigate gvl-responses of hla-mismatched aml-reactive cd + cytotoxic t cells upon transfer into primary aml engrafted nsg mice in the presence of autologous t helper cells or il- will be reported. in summary, we have shown that cd + t cell engraftment and homeostatic proliferation in nsg mice is dependent on cd + t cell-mediated or, alternatively, il- -driven signals. we conclude that our nsg transplantation model provided with these signals may be a valuable tool for evaluating gvh-and gvlreactivity of ex vivo modifi ed human donor t cell grafts in vivo. oral session : reduced-intensity conditioning o ric allo-hsct for haematological malignancies up to years is feasible without extratoxicity: a retrospective study of patients on behalf of the sfgm-tc p. chevallier, d. blaise, n. milpied, m. michallet, j.p. vernant, n. fegueux, m. renaud, b. rio, n. gratecos, j.y. cahn, g. socie, i. yakoub-agha, a. huynh, s. francois, j.o. bay, c. cordonnier, a. buzyn, n. contentin, e. deconinck, m the aim of this report was to assess the outcome of patients aged ≥ years who received a ric allo-sct, with a special emphasis on the comparison of the outcome of patients aged - years (y.) and patients aged > y. between and , patients aged ≥ years with different haematological diseases were treated with a ric allo-sct, and reported to the sfgm-tc registry. patients twice allografted were excluded from this study. this series included males ( %) and females ( %). the median age for the whole cohort was (range, - ) y. patients ( %) were diagnosed with a myeloid malignancy. patients ( %) had high risk disease features. patients ( %) received a ric allo-sct from an hla-matched related donor. pbsc were used in % of the patients (n = ). the conditioning regimen consisted of fludarabine and busulfan in cases ( %), fludarabine and low dose tbi in cases ( %). the remaining patients ( %) received other socalled ric protocols. atg was used in half of cases. with a median follow-up of (range, - ) months after allo-sct, engraftment was %, grade ii-iv and grade iii-iv acute gvhd occurred at a median of days after allo-sct in % (n = ) and % (n = ) of patients, respectively. chronic gvhd was observed in out of evaluable patients ( %). nrm was % (n = ). -year os was % ( %ci, - %). in order to assess the applicability of ric allo-sct to the older age group, we next compared the outcome of patients aged from to y. (n = ; median age y.) and those aged > y. (n = ; median y.). except for age, in univariate analysis, these groups were not statistically different in terms of demographic, disease or transplant characteristics. overall, the median time to anc> /μl was (range, - ) days in each group. the grade ii-iv acute gvhd ( % vs. %), the overall nrm ( % vs. %), the relapse ( % vs. %) and the deaths ( % vs. %) incidences, as well as the -year os ( . % ( %ci, - %) vs. % ( %ci, - %)) were comparable between the younger population vs. the older one (p = ns). in a cox multivariate analysis accounting for all relevant factors, age > y. was not found to be a statistically signifi cant factor associated with worsened survival. despite its retrospective nature and the inherent selection biases, this data support the use of ric-allo-sct in patients up to years. final results of uk multi-centre, phase ii study of campath- h dose de-escalation prior to non-myeloablative hla-identical sibling stem cell transplantation g. orti, k. peggs, m. collin, r. clark, d. milligan, h. roddie, e. liakopoulou, c. crawley, a. clark, r. pettengel, n. chowdhry, m. roughton, j. snowden, e. morris, k. thomson, p. kottaridis, a. fielding, s. mackinnon, r. chakraverty on behalf of the uk collaborative group background: inclusion of mg campath- h (cam) as part of a fludarabine/melphalan conditioning regimen is effective at preventing gvhd and reducing nrm following allogeneic sct. however, these benefi ts are offset by high rates of infection and potentially a loss of graft-versus-tumour effects. we reasoned that reductions in the dose of cam would permit improved immune reconstitution post-transplant. methods: we performed a multi-centre trial in which the total dose of cam was reduced step-wise in cohorts from mg to mg prior to hla-identical sibling transplantation (n = ). eligibility criteria were patients with haematological malignancies aged - , life expectancy > months and unsuitable for myeloablative conditioning. primary endpoints were nrm, incidence of gvhd or infection. the study received irb approval and all patients gave informed consent. the cam dose was reduced over cohorts: mg (group a, n = ); mg (group b, n = ); mg (group c, n = ) and mg (group d, n = ). results: median follow up is . years. median age was yrs (range - ). -yr os was % with no differences observed in between the groups. -yr nrm was % overall ( . % in group a, . % in b, . % in c and . % in d, with no significant differences between the groups). cumulative incidences of grade ii-iv acute gvhd was . % in group a, . % in b, . % in c and . % in d. -yr cumulative incidences of extensive chronic gvhd were . %, . %, % and . % in groups a, b, c and d respectively. the incidence of infection and kinetics of cd , cd , total lymphocyte recovery did not differ between the groups. the lowest dose cohort ( mg, group d) compared unfavourably to the other cohorts combined ( - mg, groups a-c), in terms of cumulative incidence of severe grade iii-iv acute or chronic, extensive gvhd (hr . , % ci . - . ) and a trend for greater number of non-relapse deaths due to infection (hr . , % ci . - ). patients died of grade iii-iv acute gvhd in group d with no deaths related to this complication in the other groups. conclusions: signifi cant dose de-escalation of cam prior to hla-identical sibling transplantation is feasible without increasing nrm, although reductions below mg are associated with a higher risk of severe acute and extensive chronic gvhd. in future studies, we will determine whether use of cam at a dose of mg for hla-identical sibling transplantation will translate into improvements in progression-free survival. comparison of bu-cy-based standard myeloablative conditioning vs. fl udarabine and busulfan (flu-bu)-based reduced-intensity conditioning m. mohty, m. labopin, g. socié, n. milpied, m. attal, d. blaise, o. ringden, p. brown, b. lorentz, m. brune, r.m. hamladji, r.f. duarte, a. nagler, v the aim of this analysis was to compare outcomes (lfs, nrm, and relapse incidence) between patients receiving the classical bu-cy myeloablative conditioning (mac) regimen vs. patients receiving a fl udarabine and busulfan (flu-bu) reduced-intensity conditioning (ric) regimen prior to allo-sct. since the use of ric in young patients is still limited, the analysis was restricted to patients aged > years, who were transplanted using an hla identical sibling donor for aml in cr . in summary, in this specifi c setting of aml in cr , lfs is not statistically different when using mac or ric allo-sct for patients older than years. however, for younger patients ( - y. age group), the use of ric is associated with a higher relapse rate. prospective trials addressing the use of ric in patients younger than years are needed. indeed, reducing toxicity without compromising disease control could be of signifi cant benefi t to many patients, but "more intensive" regimens, despite the hazard of increased toxicity, may be necessary in others. thus, the trade-off between dose intensity, toxicity, and disease control will remain to be assessed for each individual patient. long-term results of reduced-intensity conditioning with busulfan and fl udarabine ± atg prior to allogeneic hct: -year follow-up k. sockel, m. bornhäuser, n. kröger, d. beelen, a. fauser, r. schwerdtfeger, w. siegert, l. kraut, a. cook, t. zabelina, c. theuser, g. ehninger, j reduced intensity conditioning (ric) prior to allogeneic stem cell transplantation (hct) is being used for years now. the long-term outcome of ric conditioning with respect to late relapse and late non-relapse mortality (nrm) is still subject to ongoing research. therefore, we sought to determine factors predictive for long-term nrm and overall survival (os). methods: we retrospectively analyzed data from patients who received ric based on busulfan and fl udarabine prior to allogeneic hct between march and july in six major german transplant centers. overall survival was analysed using multivariate cox regression models, while competitive event statistics were applied for nrm. results: we identifi ed patients with a median age of years (range, to ). the underlying hematologic diseases were: aml/mds ( %), cml ( %), cll ( %), lymphoma ( %) or other hematologic malignancies ( %). donors were matched siblings in %, other matched relatives in %, matched unrelated in % and unrelated with single mismatches in % of the patients. the incidence of acute gvhd grades ii-iv was %. the cumulative incidence of any episode of extensive chronic gvhd at years after hct was %. patients were alive with a median follow up of years (range, to ). the -year os and relapse-free survival was % ( % ci, % to %) and % ( % ci, % to %). non-relapse mortality at years was % ( % ci, % to % that, these regimens produce similar year os (primary endpoint). however, fba is associated with better early pfs and efs and socially acceptable cost-effectiveness ratio but worse early qol. fba is also associated with better long term disease control, whereas ftbi tends to produce lower trm and higher rejection rates. clinical data might help designing individual and optimal strategies for each candidate patient, while economical data may help hospitals to tailor their transplant program, depending on the patient population that they care for. allogeneic-related stem cell transplantation with reduced-intensity conditioning versus best standard of care in older patients with acute myeloid leukaemia in fi rst complete remission. interim analysis of a prospective multi center phase iii trial t.l. kiss ( ) introduction: allogeneic transplants after reduced intensity conditioning (rict) are increasingly used in patients (pts) with acute myeloid leukemia (aml). there is however a lack of prospective data supporting this practice. we present an interim analysis focusing on safety of an ongoing phase iii multi center trial aiming at determining whether rict from an hla matched sibling donor (msd) leads to improved overall survival (os) compared to standard of care in elderly pts with aml in fi rst complete remission (cr ). patients and methods: elderly pts with de novo or secondary aml in cr with non favourable risk cytogenetics were enrolled if they had a potential sibling donor and assigned to the rict group or control group depending whether a msd was identifi ed. both groups were followed prospectively. in the rict group % of pts were conditioned with fl udarabine ( mg/m and busulphan . mg/kg iv or mg/kg po). graft versus host disease (gvhd) prophylaxis consisted of ciclosporine with mycophenolate mofetil or methotrexate. the study is supported by the canadian blood and marrow transplant group. results: between and june pts ( males, females), median age ( - ) yrs were enrolled and allocated to the rict (n = ) or control (n = ) group. median follow up for the rict and control group was and months, respectively. after a median of . months ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) pts in the rict group ( %) underwent allografting. pts did not reach transplant due to non-relapse death (n = ); relapse-related death (n = ) or comorbidities (n = ; infections, cardiac disease). six pts ( %) died after transplant without previous relapse, due to gvhd (n = ), infection (n = ) and vod (n = ). relapse rate for patients assigned to the rict group was % (n = ), for patients actually transplanted % (n = ). in the control group there were ( %) non-relapse deaths and ( %) relapses. kaplan-meier estimates of year os and pfs in the whole study group was % and % respectively, with no signifi cant difference between rict and control arms. conclusion: this early analysis indicates that rict can be performed with relatively low transplant related mortality and with a moderate relapse rate. the full potential of rict can only be assessed after inclusion of more patients, longer follow-up, and by reducing the rate of pts not reaching transplant due to early relapse. in this interim analysis, no conclusions can be drawn on the putative positive effect of rict in elderly patients with aml. t-cell depleted reduced-intensity transplantation followed by donor leukocyte infusions to promote graft-versuslymphoma activity results in excellent long-term survival in patients with multiply relapsed follicular lymphoma k. thomson ( ) follicular lymphoma is an indolent disorder, which is treatable but considered incurable with chemotherapy alone. the curative potential of allogeneic transplantation using conventional myeloablative conditioning has been demonstrated, but this approach is precluded in the majority of patients with fl, because of excessive toxicity. reduced intensity conditioning regimens are therefore being explored. this study reports the outcome of consecutive patients with fl transplanted using fl udarabine, melphalan and alemtuzumab. patients were heavily pretreated, having received a median of lines of prior therapy, and % had failed previous autologous transplantation. median patient age was years and % received stem cells from unrelated donors. with a median follow-up of years, the non-relapse mortality was % at years ( % for sibling, % for unrelated donor transplants), acute gvhd grade ii-iii occurred in %, and the incidence of extensive chronic gvhd was only %. relapse risk was %, and this was signifi cantly reduced where mixed chimerism had been converted to full donor chimerism by the use of dli ( % vs. %; p = . ). in addition, / ( %) given dli for relapse post-transplant remitted, with of these responses sustained and one responding to further dli. in these patients, cr is currently ongoing at a median of months ( - ) following last dli. current pfs at years was % for the whole cohort: % for those with sibling donors and % for those with unrelated donors. the excellent long-term survival with associated low rates of gvhd and frequency and durability of dli responses makes this an extremely encouraging strategy for the treatment and potential cure of fl. we analyzed the incidence and the risk of secondary solid tumors for , adult recipients ( , related bone marrow s (bm) recipients, , related peripheral blood (pb) recipients, , unrelated bm recipients, , unrelated cord blood (cb) recipients objectives: allogeneic hematopoietic stem cell transplantation (hsct) can severely compromise patients' health related quality of life (hrqol), but there is lack of evidencebased data in this area. this is the first report to provide a longitudinal assessment of hrqol in transplanted thalassemia children and their families also using a parent proxy-report evaluation. hrqol scores in children and parents were prospectively evaluated as well as parent healthcare satisfaction. methods: fifty-eight thalassemia patients from middle eastern countries were evaluated at baseline. twenty-eight children (median age . years) were transplanted from an hlamatched donor ( sibling, unrelated and haploidentical) in italian hsct centers. the pedsql . generic core scales were administered to patients and their parents both before and after ( , , months) transplantation. parent healthcare satisfaction was assessed using the pedsql healthcare satisfaction generic module. these questionnaires were also administered to conventionally treated thalassemia patients and their parents, all coming from the same geographical area. change from baseline at , and months was assessed using the wilcox signed rank test. results: two-year overall survival, thalassemia-free survival, mortality and rejection were %, %, % and %, respectively. the cumulative incidence of acute and chronic gvhd was % and %, respectively. eighteen months after transplantation, total pedsqol scores were signifi cantly higher than baseline ratings, both in child ( + . , p = . ) and parent reports ( + . , p< . ) (figure ). while physical, emotional and social functioning scores generally tended to decrease after months in comparison with baseline values, they all returned to higher levels at months after transplantation. final pedsqol scores were also signifi cantly higher than those obtained for conventionally treated patients (p< . ). conclusion: this is the fi rst longitudinal study showing a trend towards a better hrqol in thalassemia children at months after transplantation compared to pretreatment levels and conventionally treated patients. also parent satisfaction with cure and doctor-family communication was high. these data could help to relieve some of the uncertainty surrounding the diffi cult choice of transplantation in a chronic disease like thalassemia. allogeneic stem cell transplantation (sct) is potentially curative therapy for patients (pts) with acute leukemia and mds. sct is associated with substantial mortality during the fi rst years after sct due to relapse and non-relapse mortality (nrm) whereas after years survival curves often reach a plateau. the pattern of late events was reported in myeloablative conditioning (mac) but is not well defi ned in the reduced intensity (ric) setting. to explore late outcomes we analyzed sct results in a cohort of pts with aml/ mds and all. pts meeting standard eligibility criteria were given mac (bucy or cy/tbi)). pts considered at excessive risk for nrm were given reduced toxicity conditioning (rtc) such as fl udarabine (f) with high-dose busulfan (bu) or treosulfan or a ric regimen of f and reduced bu or melphalan. the -year overall survival (os) was % ( ci, - ) and was similar with the various regimens. we identifi ed pts with aml/mds (n = ) or all (n = ) who were alive and leukemia-free years after sct from related (n = ) or unrelated donors (n = ), using ric (n = ), rtc (n = ) or mac (n = ). median age was ( - ). at the -year time-point, pts had a history of acute gvhd and had active chronic gvhd which still required immune suppressive therapy (ist) in . the probability of remaining alive for the next years was % ( ci, - ). it was %, %, and %, after ric, rtc and mac, respectively (p = ns). there were deaths beyond years; due to relapse and due to nrm. nrm included deaths due to solid cancers. there were additional secondary cancers in pts currently alive. two pts died of myocardial infarction and two of chronic gvhd. in all, the cumulative incidence of late nrm and relapse mortality was % ( - ) and % ( - ), respectively. advanced age (> ) was the most signifi cant predicting factor for shortened os; % and % in the older and younger groups, respectively (p = . results: recipients with mbl levels < ng/ml were at increased risk to suffer from one or more major infections (p = . ) during entire follow-up. infectious susceptibility was increased after neutrophil recovery, particularly until months (hazard ratio (hr) . ) with sustained effects afterwards (hr . ) (figure ). in multivariate analysis mbl serum concentrations < ng/ml were independently associated with major infections after neutrophil recovery (p = . ) when corrected for recipient age at transplantation, sex, diagnosis, history of total body irradiation, history of splenic irradiation or splenectomy, acute or chronic graftversus-host disease, and duration of immunosuppression. in subgroup analysis occurrence of severe herpes virus infections in particular was associated with signifi cantly lower mbl levels (p = . ). conclusion: our fi ndings indicate that low mbl levels may predict markedly increased susceptibility to severe infections with sustained effects even late after allogeneic hsct. determinations of mbl status should therefore be included into pretransplantation risk assessment. introduction: aim of this study was to evaluate frequency and risk factors of both secondary myelodysplastic syndromes/acute leukemias (smds/al) and solid tumors in lymphoma patients treated with the high-dose (hd) squential (hds) chemotherapy with peripheral blood progenitor cell (pbpc) autograft. patients and methods: data have been collected on , lymphoma patients treated at centers associated to gitil ( figure ). patients received either the original or modifi ed hds regimens, as shown in figure . pbpc were collected after hdcyclophosphamide (cy), and, in a subgroup, after a nd round of mobilization with hd-ara-c. to reduce the incidence of acute and chronic graft versus host disease (gvhd) anti-t-cell globulins (atg) have been incorporated into the preparative regimen for allogeneic stem cell transplantation (sct) from alternate donors by many centers. different atg preparations are available and little is known about the optimal dosing. therefore we conducted this retrospective registry study utilizing specifi c questionnaires to participating centers. chronic myelogenous leukemia (cml) in chronic phase has been selected as underlying disease in order to have a rather homogenous patient population. a total of patients (pts) have been analyzed. pts had received no atg, atg-fresenius, thymoglobuline® (genzyme), other in vivo t-cell depletion, mainly campath, and had received in-and ex-vivo t-cell depletion, utilizing thymoglobuline® for in-vivo depletion. a cumulative dose of less than mg/kg atg-fresenius or less than mg/kg thymoglobuline® has been defi ned as low-dose. the median follow-up for surviving pts is months (range: with no statistically difference in the different pts groups. only the use of atg-fresenius and thymoglobuline® proved to be an independent positive prognostic factor for overall survival in multivariate analysis incorporating the ebmt risk score. this was due to decreased treatment related mortality. however, any of the analyzed t-cell depletion strategies increased the risk of relapse, which did not translate into overall survival, since relapse after allo sct is manageable in cml pts. when also analyzing the dosing of atg, the use of high dose atg-fresenius was associated with the best long-term overall survival of about %. when comparing high dose fresenius versus all others the use of high dose fresenius had the same impact on overall survival as the ebmt risk score, indicating that the use of high dose fresenius is an independent positive prognostic factor. similar effects were not seen with high-dose thymoglobuline®. interestingly, the positive effects of atg only became obvious after months after transplant suggesting no protection against acute gvhd but protection against mortality from chronic gvhd. although unrelated allogeneic sct in chronic phase cml is nowadays a rather rare indication these data nevertheless prove benefi cial effects of in vivo t-cell depletion and also emphasize, that the different preparations are not interchangeable and that the dosing is of great importance. skewed t-cell receptor repertoires in very long-term survivors after allogeneic haematopoietic stem cell transplants a. rovó ( ), c. arber ( ), p. fisch ( ), u. siegler ( ) disturbed immune reconstitution and late auto-immune like phenomena have been observed after hsct and described as "late altered immunity" in very long-term survivors. in a previous cross-sectional study on long-term survivors after allogeneic hsct (> years) and their respective donor, we demonstrated that recipients with cgvhd and a female donor had signifi cant telomere shortening of the hematopoietic cells, and subtle signs of altered late organ function as compared to their respective donor (baerlocher et al., blood, ). we were therefore interested to learn more about these differences and analyzed in the same cohort the t cell receptor beta (tcrb) repertoire diversity by spectratyping of peripheral blood t lymphocytes. we compared by visual analysis the gaussian like distribution of the peaks from vb subfamilies. pattern of each vb subfamily was classifi ed as normal or skewed. normal refers to > vb subfamilies with a gaussian like distribution, abnormal to > vb subfamilies with a non gaussian pattern. the overall complexity of the tcrb repertoire (total number of peaks of all vb subfamilies) from the recipients was compared with the respective donor. the median age at hsct and at time of the study was . ( - ) and ( - ) years for the recipients; . ( - ) and ( - ) for the donors. the median time interval between hsct and tcrb repertoire analysis was years ( - ). during follow-up / analyzed recipients had cgvhd. the median number of skewed vb subfamilies was . ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) for recipients, and ( - ) for donors (p = . ). the median overall complexity was ( - ) for recipients and ( - ) for donors (p = . ). / analyzed recipients showed an abnormal pattern of the tcrb repertoire. all but two donors had a normal repertoire (p = . ); one of them suffered of neoplasms after donation one patient with an abnormal tcrb repertoire ( figure , upn ) had still extensive, severe cgvhd and showed the shortest telomere on the leukocytes and t-lymphocytes: . and . kb respectively, versus median . ( . - . ) and . kb ( . - . ) respectively. these data indicate that t cell repertoire diversity in long term survivors after allogeneic hsct is abnormal and differs from the repertoire of their respective donors. the cause of this skewed t cell repertoire remains open but gives a formal basis for the clinically observed late altered immunity. the clinical consequences thereof need to be evaluated in the future. a second allogeneic sct was given to patients (median time from sct mo, from relapse . mo), using sic ( %) or ric ( % background: center specifi c outcome data are being discussed as useful tools for patients, the insurance industry, and politicians to assess quality of specifi c centers and to guide decision making processes. hematopoietic stem cell transplantation (hsct) requires signifi cant infrastructure and is costly. hsct therefore qualifi es as a target for such an exercise. methods: we made use of the comprehensive database of the swiss blood stem cell transplant group (sbst) to evaluate center specifi c mortality rates. nine centers reported a total of hsct, allogeneic ( . %), autologous ( . %) in patients between and to treat acute or chronic leukemia ( %), lymphoid neoplasia ( %), non malignant disorders ( %) or solid tumors ( %). because of mandatory reporting by legal requirement these include data on all hsct performed in switzerland over this period. data were analyzed separately for recipients of allogeneic and autologous hsct for survival and transplant related mortality (trm) at day and at years. results: overall survival at years was + % after allogeneic and + % after autologous transplantation. there were signifi cant differences among centers in unadjusted analyses of survival and trm. these differences became absent or marginal, when results were adjusted for disease, year of transplant and the ebmt risk score (incorporating patient age, disease stage, time interval between diagnosis and transplantation, and for allogeneic transplants donor type, and donor recipient gender combination) in multivariate analysis. conclusions: these data indicate comparable quality among centers in switzerland. they furthermore demonstrate that comparison of crude center specifi c outcome data without adjustment for patient risk factors may be highly misleading. mandatory data collection and outcome analysis including all cases within a comprehensive quality management system may serve as a model for other cost intensive therapies to ascertain quality. . contrary, the more relevant negative aspect is the increase of workload ( %). the major modifi cation produced by the qms is to recognize the critical situations ( %) and to report in proper forms the nca is useful also to resolve them ( %). also, % think that internal audits are helpful to check the activities. the last question aimed to understand the relevance of qms: % agree that qms is part of daily activity and only for % "there is too much to do". the collected data show that the quality culture is, today, part of the work background and this result is recognized by % of lab staff, % of nurses and % of physicians. in general, the perception of qms is good but there are still areas to be improved, particularly regarding the management and resolution of critical situations. it is clear that to study the perception of qms is a tool to verify his effi cacy and continuously improve it, to make clear the diffi culties and fi nd appropriate corrective actions. so, qt plans to repeat the survey at least once at year in order to build a complete evaluation's system for work environment' quality and staff satisfaction. background: the liver is the current site for pancreatic islet transplantation (tx) but has many drawbacks due to immunological and non-immunological factors as well as important technical limitations. we asked whether pancreatic islets could be engrafted in the bone marrow (bm), an easily accessible and widely distributed transplant site that may lack the limitations seen in the liver. methods: pancreatic islets were implanted into the bm of diabetic c bl/ mice. islet survival, function and morphology were evaluated in comparison with the liver site. moreover a phase i/ii open label pilot study to assess the feasibility and safety of bm as alternative site for islet tx was started in patients with type diabetes mellitus. results: syngeneic islets engrafted effi ciently in the bm of c bl/ mice rendered diabetic by streptozocin (stz) treatment. for over a year post-tx these animals showed parameters of glucose metabolism that were similar to those of non-diabetic mice. islets in bm had a higher probability to reach euglycemia than islets in liver ( . fold increase, p = . ), showed a compact morphology with a conserved ratio between alpha and beta cells, and affected bone structure only very marginally. islets in bm did not compromise hematopoietic activity, even when it was strongly induced in response to a bm aplasia-inducing infection with lymphocytic choriomeningitis virus. in humans, diabetic patients received an intra bm islet infusion at the level of iliac crest without any adverse effects. in all recipients islets engrafted as demonstrated by the presence insulin producing cell in addition, preferential pairing facilitated by introduction of an extra disulfi de bond in the constant regions of the tcr chains can increase cell surface expression of the transferred tcr and decrease formation of mixed tcr dimers. another strategy based on the fact that tcrs differ in their capacity to compete for cell surface expression, is to select recipient t cells with weak competitor phenotypes. both weak and strong competitor phenotype virus-specifi c t cells were used to assess improvement of ha- -tcr cell surface expression using the different strategies. the most marked improvement in ha- -tcr expression and functionality was observed after tcr transfer of a cysteine modifi ed and codon optimized ha- -tcr resulting in % and % of ha- tetramer positive weak and strong competitor t cells, respectively. this resulted in effi cient recognition of primary leukemic cells that endogenously process and present ha- , independent of whether the recipient t cells were strong or weak competitor t cells. furthermore, results demonstrate that next to increased ha- -specifi c reactivity, neoreactivity after ha- -tcr gene transfer was dramatically reduced by cysteine modifi cation of the ha -tcr. based on these results, cysteine modifi ed and codon optimized ha- -tcrs will be used for the planned phase i/ii clinical trial to treat patients with relapsed hematological malignancies. reactivation of latent cytomegalovirus (cmv) infection is a frequent complication in cmv-seropositive patients after allogeneic hematopoietic stem cell transplantation (hsct). the s cmv-related morbidity and mortality are particularly high, if the donor is cmv-seronegative and thus, no cmv-specifi c memory t cells are being transferred from donor to recipient during transplantation. grafting non-reactive t cells of cmvseronegative donors by virus-antigen specifi c t cell receptors (tcr) may be an effi cient means to transfer cmv-specifi c t cell function into allogeneic hsct recipients. in this study, we have reprogrammed t cells of cmv-seronegative donors with human tcr recognizing the immunodominant hla-a* -binding cmvpp peptide epitope - . to overcome the limitations of retroviral tcr gene transduction that hamper clinical translation, we used in vitro transcribed rna encoding cmv-specifi c tcr for electroporation of non-reactive t cells. this procedure resulted in transient surface expression of the introduced tcr for at least days as demonstrated on both cd + and cd + t cells by specifi c tcrvb chain and hla tetramer staining analyses. the cmvpp tcr rna-transfected t cells showed hla-a* -restricted ifn-g secretion and cytolysis against cmvpp - peptide-pulsed target cells and against human fi broblasts upon cmv infection. we also observed that tcr rna transfection of cd + t cells turned them into potent cmvpp /hla-a* -specifi c t helper cells. this was demonstrated by co-incubating them with immature dendritic cells (dc), which resulted in maturation of dc only in the presence of the cmvpp epitope. furthermore, we transfected pure naïve and memory cd + t cell subsets isolated from peripheral blood of cmv-seronegative donors. although % of naïve cd + t cells were cmvpp /hla-a* tetramer positive after electroporation, they mediated only marginal lysis toward cmv-infected fi broblasts. in contrast, memory cd + t cells showed strong cytotoxicity against cmv-infected fi broblasts for at least days upon tcr rna transfection. in summary, our data demonstrate that non-reactive human t cells can be easily redirected with cmvpp tcr rna, thereby gaining cmv-specifi c t cell effector function for a considerable time period. we therefore believe that cmvpp tcr rna has the potential to be further developed as a therapeutic "off-the-shelf" reagent for cmv-seropositive patients who undergo allogeneic hsct from cmv-seronegative donors. functional cytotoxic t lymphocytes emerge in patients with ph + acute lymphoblastic leukemia under imatinib therapy and may be exploited for adoptive t-cell therapy c. quadrelli ( ) an effective specifi c immune response against adult all has so far been reported only in the allogeneic stem cell transplant setting. we have recently demonstrated that bm-resident, ifng-producing, p bcr-abl specifi c autologous t cells may occur in ph + all patients upon treatment with high dose im, and that the presence of these cells correlates with lower minimal residual disease (mrd) values and better clinical outcome. on the basis of these preliminary data, we proceeded to assess for the presence of cytotoxic t-cells among the bm resident, bcr-abl-specifi c t-cells, and characterized the cytotoxic t-cell populations identifi ed, with the aim of optimizing a protocol for the ex-vivo expansion of p bcr-abl-specifi c cytotoxic t lymphocytes (ctls) to be employed in protocols of t-cell therapy to control minimal residual disease after chemotherapy or hsct. p bcr-abl-specifi c ctls were identifi ed by co-colturing bm mononuclear cells (bmmc) of ph + lla patients with bcr-abl peptide-pulsed autologous dendritic cells; the peptides employed derived from the complete spanning of p bcr-abl fusion region. the p bcr-abl-stimulated t cell lines generated with this protocol were mainly cd + /cd + or cd + /cd + cells with effector memory phenotype, that exerted a specifi c lytic activity against bcr-abl fusion protein. in detail, we observed a p bcr-abl specifi c lytic activity > lu / in of the patients tested (median lysis against p -derived peptidepulsed targets: lu / ). in of the patients, for whom autologous leukaemia blasts were available, we could demonstrate a strong leukemia-directed lytic activity (median lysis: lu / ). the lysis directed towards autologous blasts was mainly mediated by cd + t-cells, and was hla class irestricted. lytic activity towards autologous, mock-pulsed pha blasts, or allogenic ph-blasts was low/absent. our fi ndings demonstrated that p bcr-abl-specifi c t cell responses may emerge in patients with ph + all under im treatment, and may be stimulated and expanded ex-vivo. whether these p bcr-abl-specifi c t cells may have a possible therapeutic role in ph + all patients, and may be expanded from healthy donors to be employed in protocols of adoptive cell therapy after hsct remains to be addressed in future studies. background: a positive selection of leukemia associated antigen (laa)-specifi c t cells would be highly desirable to amplify the gvl effect and to decrease the risk of gvhd. wilms tumor gene (wt ) and the receptor for hyaluronic acid mediated motility (rhamm) are laas recognized by cd + t lymphocytes. streptamers constitute a novel multimer technology to select specifi c cd + t cells, available at good manufacturing production (gmp) level. material and methods: both tetramers and streptamers were used to detect the frequency of hla-a restricted cd + t cells in the naïve peripheral blood (pb) from both healthy donors (hds) and aml patients. rhamm-and wt -specifi c cd + t cells were further characterized for the expression of cd , cd , cd ra and ccr . in the next step, laa-specifi c cells were positive selected by macs columns after labeling with streptamers and thereafter immunophenotyped. moreover, mixed lymphocyte peptide cultures (mlpcs) were performed to enrich wt specifi c t cells derived from the pb of hds. rhamm and wt specifi c cells were subjected to carboxylfl uorescein succimidyl ester (cfse) based proliferation and cytotoxicity assays. results: of hds showed naïve laa specifi c t cell frequencies of . to . % of all cd + t cells. in aml patients in complete remission signifi cantly higher frequencies of laaspecifi c t cells than in patients at diagnosis could be detected, up to . % of all cd + t cells. these cells revealed to be cd + cd -cd + cd ra + ccr -effector t cells in fl ow cytometry. after positive selection by macs columns a purity of - % could be achieved for laa-specifi c t cells. after a maximum of three rounds of mlpc, only a frequency of - % could be achieved, thus demonstrating the power of the streptamer technology. both tetramer-and streptamer-based selections yielded similar amounts of laa-specifi c t cells. after positive selection, these t cells preserved an effector t cell phenotype and showed active proliferation in cfse staining. streptamerselected t cells showed a trend towards higher cd expression thus indicating a more activated t cell status. conclusion: in summary, the streptamer technology allows to select highly pure fractions of rhamm-and wt -specifi c effector t cells with proliferative and cytotoxic properties. in analogy to dlis specifi c for viral antigens such as cmvpp , production of leukemia specifi c dlis is feasible on a gmp level. further leukemia antigens are currently evaluated by our group. delicate balance between regulation and stimulation at the antigen presenting cell site determines the likelihood of successful priming and survival of antigen-specifi c t-cells from the naïve donor repertoire i. jedema, t.s. lam, m. van de meent, c. hoogstraten, j.h.f. falkenburg leiden university medical center (leiden, nl) although the in-vitro induction of tumor-and pathogen-specifi c t cells from the naïve donor repertoire has been shown to be feasible, the robustness of the procedure remains limited, hampering large scale clinical application. in this study, we investigated the role of individual parameters like the frequency of antigen-specifi c precursor t cells (tprec) and regulatory t cells (treg), the number of antigen presenting cells (apc) used as stimulator cells, and the number of targeted antigens (ag) on the ability to prime, enrich and expand ag-specifi c t cells from primary immune responses in-vitro. therefore, we developed an in-vitro model system allowing the monitoring of ag-specifi c activation and proliferation of individual naïve donor t cells in the fi rst days of the immune response using pkh labeling, cd counterstaining and quantitative fl owcytometric analysis. in this model we exposed naïve tprec to allogeneic apc in different responder/stimulator (r/s) ratios in the presence of different numbers of innocent bystander cells. optimal t cell activation was seen at specifi c tprec/s ratios between / and / , irrespective of the number of innocent bystander cells. lowering the number of stimulator cells per tprec resulted in incomplete activation and proliferation, but more importantly, exposure to an excess of stimulator cells resulted in induction of activation-induced cell death (aicd) of the antigen-specifi c tprec. next, we investigated the role of treg in the induction phase of the immune response. treg were like tprec attracted to the site of the apc and their activation further increased their inhibitory potential. especially when they were at a numeric advantage, treg were capable of impairing antigen-specifi c tprec priming. increasing the number of apc in the induction phase of the immune response could overcome this negative regulation since the number of treg per apc is diminished. however, at low tprec frequencies, in case of single peptide responses, this will lead to aicd of the antigen-specifi c tprec. this may be prevented by increasing the number of antigenspecifi c tprec by simultaneous targeting of multiple antigens. in conclusion, the in-vitro generation of antigen-specifi c primary immune responses can only be successfully and reproducibly performed by creating an optimal balance at the priming site of the immune response (e.g. the apc) between the number of negative regulators (treg) and responding cells (tprec). fistulas are an invalidating, often diffi cult to treat, complication of patients with crohn's disease (cd), a disabling, chronic, relapsing infl ammatory enteropathy caused by dysregulation of the immune tolerance towards intestinal bacteria in genetically susceptible individuals. mesenchymal stromal cells (mscs) represent a promising tool in approaches of regenerative medicine. we investigated the feasibility, safety and effi cacy of local injection of autologous bone marrow (bm)-derived mscs for refractory cd fi stulas. mscs were isolated and expanded ex vivo in the presence of platelet lysate from bm of patients ( males, median age yrs, range - ). patients received intrafi stular injection of mscs scheduled every weeks (median infusions) and were monitored at time of each injection, and , , , months after the last treatment. the cytokine profi le of mscs and their ability to infl uence apoptosis of mucosal t cells obtained from involved and uninvolved colonic areas were also analyzed. msc expansion was successful in all patients and no adverse event was recorded during and up to months after treatment. intrafi stular injection of mscs was effective in inducing sustained closure of fi stulas, with the appearance of regenerative tissue along the tracks. in particular, patients ( %) benefi ted from complete and sustained healing of fi stula tracks, while three had partial response. all patients showed a signifi cant reduction of both cd activity index (pre-and posttreatment median values: sd and sd at months after the last infusion; p < . ) and perianal disease activity index (pre-and post-treatment median values: . sd . and . sd . at months after the last infusion; p < . ) reaching disease remission usually after the second procedure. the immunephenotype of circulating t lymphocytes showed progressive increase of the number of cd + cd bright foxp + cells, which became signifi cant (p< . ) after the second procedure and remained stable up to months after the last infusion. no modifi cation of serum cytokines was observed at any time point. mscs caused a sort of block of the rates of both apoptotic and living cells when incubated with t lymphocytes from diseased mucosa, whilst critically increased the apoptotic rate when incubated with t lymphocytes from apparently healthy mucosa. local injection of autologous bm-derived mscs appeared feasible, safe and successful in treating fi stulas associated with cd. we recently showed that nk cells require two signals to initiate lysis; the fi rst primes (s ) and a second triggers (s ). we found nk resistant cell lines which express s but lack s and that resting nk cells stimulated with these cells retain the primed state even after cryopreservation. these are termed tumour activated nk cells (tank). we are conducting a clinical trial in patients with aml in cr or pr with less than % blasts who have exhausted conventional treatment options. tank are generated from a haploidentical family donor by overnight incubation of nk cells with lysed ctv- cells. tank are then purifi ed from the lysate and released at a single dose of nk/kg with a t cell contamination of < /kg and cryopreserved. pre-infusion conditioning -fludarabine ( mg/m /day) for days plus gy tbi on day . eleven patients have been enrolled to date, of whom have been treated. patient characteristics: pt - yr female; previous autologous transplant, treated in cr . pt - yr male; in pr after -azacytidine. pt - yr male; previous allogeneic transplant, treated in cr . pt - yr male; treated during relapsing disease and circulating blasts. pt - yr female treated in cr following one course of induction chemotherapy. pt - yr male; treated in cr . results: no infusional toxicity. all patients suffered a degree of bone marrow suppression needing in-patient supportive care. aplasia ranged from weeks up to days. pt remains in cr at month + ; pt achieved and remained in cr until month + , relapsed and treated with second tank infusion; pt prolonged and severe pancytopaenia requiring cd selected stem cell rescue, achieved and remained in cr until month + . ; currently undergoing re-induction chemotherapy. pt cleared peripheral blasts for months; now with progressive disease. pt in cr, months post infusion; pt is non-evaluable at present. extremely prolonged nk chimerism (up to + months) has been seen in all patients in the absence most cord blood transplants (cbt) are performed using cord blood units mismatched with the recipient at one or two hla loci. it is not known whether the locus at which mismatches occur infl uences outcomes. we examined the effect of locusspecifi c mismatches on outcomes of hla-mismatched (one mismatch, n = , two mismatches, n = ) single cbt performed for haematological malignancies at ebmt centres from to . two digit typing for hla-a and hla-b and digit typing for hla-drb was used. patients were transplanted for all (n = ), aml (n = ), mds (n = ), chronic leukaemia (n = ), lymphoma (n = ) or myeloma (n = ). among the patients who received a cord with one hla mismatch, the mismatch occurred at hla-a (n = ), hla-b (n = ) or hla-drb (n = ). transplant-related mortality (trm) for this group at years was ± % and was ± %, ± % and ± % for hla-a, -b and -drb mismatches respectively. on multivariate analysis the mismatched locus had no infl uence on trm when adjusted for age, year of transplant, stage of disease, cmv status and cell dose. there was a trend for less relapse for drb mismatches ( ± % vs. ± %, p = . ) but this did not lead to a difference in disease-free survival (dfs) between groups ( ± % vs. ± %, p = ns). neutrophil engraftment and acute graft-versus-host disease (gvhd) were unaffected by locus of hla mismatch. for the patients who received a cord with two hla mismatches, mismatch combinations included a + b (n = ), b + drb (n = ), a + drb (n = ) or two mismatches each at either a (n = ), b (n = ) or drb (n = ). trm for this group was ± % at years and was ± % (a + b), ± % (b + drb ), ± % (a + drb ), ± % (a + a), ± % (b + b) and ± % (drb + drb ) by mismatch group. the loci at which hla mismatches occurred did not infl uence trm on multivariate analysis, nor when grouped according to mhc class (class i only vs. class ii±i). however, double drb mismatch was associated with more grade ii-iv acute gvhd ( ± % vs. ± %, rr = . , p = . ). in addition, on multivariate analysis a + b mismatches were associated with improved probability of dfs ( ± % vs. ± %, hr = . , p = . ). a cell dose cut-point could not be defi ned in either population. in summary, for cbt with one hla mismatch, the locus at which mismatch occurs does not infl uence outcomes. however, for cbt with two hla mismatches, it seems that two mismatches at class i (hla-a and -b) are preferable to other mismatch combinations. matched unrelated donor is associated with lower relapse than matched related donor in reduced-intensity conditioning transplantation: implications for graft-versus-malignancy effect v. ho, h. kim, j. aldridge, g. kao, d. liney, j. koreth, p. armand, c. cutler, j. antin, r. soiffer, e. alyea dana-farber cancer institute (boston, us) as success of reduced intensity conditioning (ric) stem cell transplantation (sct) relies on graft-vs-malignancy (gvm) effect, the extent of minor hla antigen disparity in matched related donors (mrd) vs. matched unrelated donors (mud) could impact clinical outcomes. while convention dictates that mrd is preferred over mud, does this wisdom hold in ric sct? we conducted a retrospective review of ( mud, mrd) ric sct performed at our institution from / through / . all received uniform ric conditioning with busulfex ( . - . mg/kg) and fludarabine ( mg/m ). gvhd prophylaxis was mostly tacrolimus/mini-methotrexate ± sirolimus ( %). sc source was pbsc ( %), marrow( %). all donors were / allele matched at hla a, b, c, drb , dqb . the mud and mrd cohorts were balanced in demographics, disease (mds/aml % vs. %), high disease risk ( % vs. %), prior sct ( % vs. %), and gvhd prophylaxis. median cd + dose was higher in mud ( . vs. . × cd + /kg, p = . ). cumulative incidence of grade ii-iv acute gvhd and -yr chronic gvhd were similar, % vs. % (p = . ), and % vs. % (p = . ) in mud and mrd, respectively. there was no difference in transplant related mortality (trm), time to neutrophil or platelet engraftment, or % achieving over % donor chimerism at days + and + . cumulative incidence of relapse was % for mud, % for mrd, p = . . with a median follow-up of and months, -yr progression free survival (pfs) was % for mud and % for mrd, p = . ( figure ). -yr overall survival (os) was % for mud, % for mrd (p = . ). improvement in pfs did not translate to os benefi t because many relapses were salvaged with second sct or other therapy. in competing risks regression, mrd was associated with increased risk for relapse compared to mud (hr . , p = . ), but no difference for trm. in cox regression, mrd, prior sct, high disease risk, and mds/ aml diagnosis were associated with inferior pfs (table ) . donor age, cd + dose, and year of sct were not associated with worse survival. in ric sct, mud is associated with signifi cant improvement in relapse and pfs compared to mrd, without increased gvhd or trm. this improvement is not related to younger donor age or higher cd + count, suggesting that hla minor antigen disparity in mud sct may mediate stronger gvm. while further investigation is warranted, these results could have great implications for the future selection of donors for ric sct. tolerance of g-csf-administration was different in the two groups: / ( , %) sib donors stated, that they only poorly had tolerated g-csf administration, / ( , %) mud donors said that their tolerance to g-csf had been poor. / ( , %) sibs indicated, that the infl uence of pbsc donation to their health status had been negative, while / ( , %) unrelated donors said that there had been a negative infl uence of pbsc-donation on their health status. / ( , %) sibs described their current health status as "bad", while none of the muds did so. the most serious adverse effects that occurred after donation were one case of hodgkin's disease in one of the related donors and one case of acute lymphatic leukaemia in an unrelated donor. furthermore, one case of squamous cell lung carcinoma occurred in a related donor and one case of mamma carcinoma in an unrelated donor. in addition to these malignancies the following health problems could be noted in the donors: subdural haematoma a few days after donation: sib, mud. sarcoidosis: sib, mud. cardiac disorder: sibs; mud. essential hypertension: sibs, muds. discus hernia: sib, muds. hypothyreosis: sib, mud. depression: sib, mud. tinnitus: sib, muds. the presented data show that serious health problems after donation may occur and therefore global collection of these data is necessary in order to calculate the actual risks of pbsc donation in advance. evaluation, consent and care of related donors and recipients at haematopoietic stem cell transplant centres in the united states: practice patterns and potential for confl ict of interest p. anderlini ( ), t. pedersen ( ), d. confer ( ) background: a real or perceived confl ict-of-interest may arise in the situation where the same physician is responsible for the care of two individuals whose care is interdependent. in hematopoietic stem cell transplantation (hsct) from unrelated donors facilitated by the national marrow donor program, donors and recipients are under the care of separate physicians in order to provide unbiased care and eliminate the potential for a confl ictof-interest. however, the practice patterns of evaluation and care for related donors and recipients at centers performing hsct are unknown. material and methods: a practice pattern survey of transplant centers in the united states reporting to the cibmtr was conducted by the donor health and safety committee between december and july . the survey was administered as an online survey tool sent to the center medical directors. the survey focused primarily on determining the type of provider involved in medical clearance, informed consent and medical management of the donor and the providers relationship to the recipient. results: of centers surveyed, evaluable responses were received ( %). the median number of related donor transplants per year at responding institutions was (range: - ); the median total number of transplants per year was ( - ). as shown in the table, transplant physicians in greater than % of centers were involved in overlapping care of the donor and the recipient during the donor evaluation, clearance and collection phases. these patterns were similar between transplant teams caring for adult or pediatric donors and recipients. conclusion: among responding centers, it appears that medical management of recipients and their related donors by the same provider is common, and may not be viewed as a potential confl ict-of-interest. whether this potential confl ict-of-interest affects donor care is unclear, and deserves further investigation. long-term follow-up and risk factors for outcomes after related hla-identical cord blood transplantation for patients with malignancies. an analysis on behalf of eurocord-ebmt a.l. herr ( ) , n. kabbara ( ), p. teira ( ), c. bonfi rm ( ) outcomes after related hla identical bone marrow or cord blood transplantation (cbt) in children have been reported to be comparable (nejm ) . in order to analyze risk factors for outcomes, we studied patients with malignancies who had a fi rst single unit related hla identical unmanipulated cbt and were reported to eurocord-ebmt. cbt were performed since , in countries and centers. median fu was . years (range months to years). nearly all patients were children, median age being years. acute leukemia (al) was the main diagnosis (n = ) followed by myelodysplasia (n = ), chronic myeloid leukemia (n = ), solid tumors (n = ), lymphomas (n = ) and hemophagocytic lymphohistiocytosis (n = ). according to ibmtr criteria, patients had low, intermediate and high risk diseases. all cb donors were siblings except : one nd degree relative and one child. cb units contained a median of . × /kg total nucleated cells (tnc) at collection and . × /kg tnc after thawing. the conditioning regimen was myeloablative for patients, % including tbi. the most frequent gvhd prophylaxis regimen was cyclosporine alone. methotrexate (mtx) was used for patients. the cumulative incidence (ci) of neutrophil recovery was % at d + with patients achieving a neutrophil count of . × /l after a median time of days. infused tnc ≥ . × /kg (p = . ) and the lack of mtx in gvhd prophylaxis (p = . ) were independently associated with improved neutrophil recovery. ci of acute and chronic gvhd was % at d + and % at years, respectively. ci of non-relapse mortality was % and of relapse was % at years. remission status (p = . ) and the use of tbi-containing regimens (p = . ) were independently associated with a lower relapse rate in al patients. for cbt performed in year < vs. ≥ , disease-free survival at years was % vs. % (p = . ), and overall survival (os) at years was % vs. % (p = . ). transplant year ≥ (p = . ), low or intermediate risk disease status at cbt (p = . ) and infused tnc ≥ . × /kg (p = . ) were independently associated with improved os. five-year os in al patients was associated with remission status (p = . ): % for patients in cr , % in cr , % in cr and % in advanced phase of the disease. improving results over time and the low toxicity of related hla-identical cbt in malignancies support banking and use of family cb units with suffi cient cell dose. in the past, the outcome of adolescents with acute lymphoblastic leukaemia (all) in nd complete remission (cr) given hsct s prophylaxis was carried out with atg or okt . primary engraftment occurred in %. after reconditioning, fi nal engraftment was achieved in %. gvhd grade - occurred in %. % had grade ii, % had grade iii. chronic gvhd occurred in patients. no gvhd related mortality was observed. median follow up was . years. efs at year was % (cr ), % (cr ) and % (≥cr ); efs at years was % (cr ), % (cr ) and % (≥cr ). median survival of patients with active disease was . years. relapse rates at year were . (cr ), . (cr ) and . (≥cr ); relapse rates at years were . (cr ), . (cr ) and . (≥cr ). trm at year was %, causes of death remained viral and fungal infections, no ebv lpd occurred. conclusions: positive selection of progenitor cells or depletion of t and b cells can minimize acute and chronic gvhd in both matched unrelated and mismatched related transplantations and may prevent gvhd related mortality. lethal infections occurred in % of the patients probably due to the delayed recovery of t cells. despite profound depletion of donor t cells, relapse rates were acceptable and remained on a stable level after the fi rst year in patients with complete remission. thus, absence of gvhd may not necessarily be associated with high relapse rates in childhood all. apart from t cells, other effector cells like nk cells are likely to exert gvl effects and may contribute to the favorable efs of our patients. fanconi anemia (fa) is a rare disorder characterized by progressive bone marrow failure, congenital abnormalities and a predisposition to cancer. bmt is the only treatment able to restore normal hematopoiesis in this disease. here, we update the brazilian experience using only cyclophosphamide mg/ kg (cy ) as a preparative regimen for the treatment of pts with fa. objective: analyze the outcome of pts with fa submitted to a related bmt using cy mg/kg. material and methods: period: / - / ; gender: f/ m age: - ys (m: , ). type of donors: matched siblings donors (msd): pts; other related donors (ord): pts. pts were in aplastic phase while had mds. all pts received a preparative regimen with cy mg/kg and gvhd prophylaxis with cyclosporine and methotrexate. statistical analysis was performed using spss. overall survival (os) was estimated using the kaplan-meier method. multivariate analysis was performed by cox method. results: pts are alive between - days (m: ) after bmt. os: , % at ys. pts transplanted below the age of ( pts) had an os of , % at ys. there was no statistical difference between transplants performed in curitiba( pts) and other bmt centers ( , % × , %). pts died before d + and were not evaluable for engraftment. only one pt had primary graft failure and he is alive and well ys after the rd transplant. late rejection occurred in pts ( pts with mds, pts with ord, pts with msd) at a median of days after bmt. / pts are alive and well after a nd bmt. cumulative incidence of rejection at y was %. pts receiving > transfusions had a lower survival when compared to those less transfused ( % × % p = , ). mucositis grade ii-iii occurred in the majority of pts; / pts developed acute-gvhd grade ii-iv has been reported to be worse than that of children. in order to evaluate whether in recent years the outcome of adolescents has become more similar to that of children, we studied paediatric patients (age < years at transplantation) reported to the aieop-hsct group registry who received allogeneic hsct for all in nd cr in a paediatric institution between / and / . a total of patients ( % males and % females) were anallyzed. patients were children (< years old), whereas the remaining patients where adolescents ( - years old). median age at hsct was . and . years, for children and adolescents, respectively. a mfd was employed in % of children and in % of adolescents; the remaining patients having received a mud hsct. conditioning regimen included tbi in % of patients, in most cases associated with thiotepa and cyclophosphamide. cyclosporine (csa) alone was used for gvhd prophylaxis in % of patients transplanted from a mfd, whereas the combination of csa, mtx and anti-thymocyte globulin (atg, . mg/kg from day - to day - ) was employed in % of patients given mud hsct. no difference between children and adolescents was observed for patient-and transplant-related variables, with the exception of the infused cell dose which was greater in children. as of october , the probability of developing grade ii-iv, grade iii-iv acute gvhd or chronic gvhd was %, % and % for children, and %, % and % for adolescents (p = ns). eight-year probability of efs,trm and ri were %, % and % for children and %, % and % for adolescents, respectively (p = ns). univariate analysis showed that s + s bfm classes at relapse (p< . ), recipient male gender (p = . ), mud hsct (p = . ), grade or iv acute gvhd (p< . ) were poor-risk factors for efs. only bfm class at relapse and acute gvhd (grade or iv) maintained their prognostic signifi cance in multivariate analysis. adolescents and children < years of age, treated in paediatric institutions, had the same outcome. multivariate analysis confi rmed the gvl favourable effect in patients with grade i-iii acute gvhd.results obtained with mud hsct were comparable to those achieved with mfd hsct. adolescents should be referred for transplantation to treatment teams that have experience in the management of childhood all. long-term survival and relapse rate after transplantation of highly t and b cell-depleted stem cells from alternative donors in paediatric patients with acute lymphatic leukaemia p. lang ( ) we present long term results in children with all who received highly t and b cell depleted stem cells from matched unrelated (n = ) or full haplotype mismatched related donors (n = ) at our institution. our aim was to minimize gvhd and to avoid ebv lpd in both matched and mismatched transplantations. remission status was: cr , n = ; cr , n = ; ≥cr , n = ; non remission or second transplant, n = . graft manipulation was carried out with microbeads and the clinimacs tm device (indirect depletion of t and b cells with cd + positive selection (n = ); or direct depletion with anti-cd /anti-cd coated microbeads (n = )). t and b cell counts were reduced for - log with median numbers of residual t cells of /kg bw (cd + selection) and /kg bw (cd / depletion). no pharmacological immune suppression was given after positive selection, whereas patients with cd / depletion received mmf. the conditioning regimens were either tbi or bu based (n = ) or a toxicity reduced protocol (flud, tt, mel) was used (n = ). rejection more than candidate donors were evaluated at our institution (median age y, r - y; donors ≥ y; male ). donors were evaluated with marrow smear and cytogenetic test, / were eligible for donation ( bone marrow harvest, peripheral blood gcsf mobilization), proceeded to stem cell donation without marrow evaluation. out of marrow were morphologically normal ( , %), ( , % -median age y, donors > y) presented abnormalities, ( , %) were not evaluable. we registered cases of plasma cells hyperplasia ( not eligible to donation) ( , %, donors > y) presented abnormalities, was not evaluable. we registered cases of chromosome y deletion ( not eligible to donation), of pericentric inversion of chromosome (p ;q additional follow-up will give us more important information on safety of hsc donation procedure. o impact of kir-ligand mismatches and kir genotypes on the outcome of patients receiving unrelated donor stem cell transplantation for lymphoid malignancies m. morelli ( ), a. bermema ( ) all pts received rabbit anti-thymocyte globulin for in-vivo t cell depletion. nineteen donor/recipient pairs were matched at molecular level for the hla-a, b, c, drb , dqb loci and were mismatched ( at class i, at class ii). the majority of pts had chemosensitive disease (n = , %). the median follow-up was months (range - ). we studied kir genotypes using the kir-typing kit (miltenyi biotec) and hla-cw kir ligand using high-resolution molecular techniques. results: twenty-one pts were c /c heterozygous, were c /c and were c /c homozygous. in the combination patient c absent/donor kir dl /kir dl present (n = , %), we observed no trm and only cases of acute gvhd (agvhd) as compared to other combinations (n = ) in which cases of trm and cases of agvhd were observed. this combination was associated with a trend of better cumulative incidence of trm, os, and pfs os: % vs %) or in the recipient (n = , %) showed no signifi cant association with a better os or pfs. sibling donors (n = ), haploidentical cd /cd depleted grafts (n = ), cd + selected graft with okt ( ) hlh ( ), leukocyte adhesion defi ciency ( ), chronic granulomatous disease ( ), severe immune dysregulation ( ), other pid ( ) mfd ( ) hla matched - / ). of were cords ( - / hla matched). follow up is - months (median months) children died: / in the treosulfan/fl udarabine group, / in the treosulfan/cyclophosphamide group -hlh disease day- , graft rejection and cmv, infection ( ), gvhd, gvhd +infection, gvhd + cerebral infarcts, cerebral haemorrhage, pneumonitis, vod and mds/ aml. skin toxicity was common. vod occurred in children in combination with cyclophosphamide and both had enterovirus in the gut. patients rejected (mhc ii defi ciency successfully retransplanted autoimmunity after bmt in primary immunodefi ciency diseases: single-centre report of children had aml ( in cr at high risk, in ≥cr and in relapse), had all ( in cr ; in relapse) and had high grade nhl in relapse. conditioning was: gy single fraction tbi, thiotepa ( mg/kg × ), fl udarabine ( mg/m² × ), cyclophosphamide ( mg/kg × ) no gvhd developed in / patients, developed ≥ grade ii gvhd. ten patients died ( vod, fungal pneumonia, bacterial sepsis, cns aspergillosis, systemic toxoplasmosis, adenoviral infection, mof). cd and cd counts reached, respectively, /μl medianly on days (range - days) and (range - )of cmv reactivation were signifi cantly fewer than after our "standard haplo" transplants. in kir ligand-mismatched transplants, speed of nk cell reconstitution/maturation and size of donor vs. recipient alloreactive nk cell repertoires were preserved. in conclusion, in the setting of haploidentical transplantation infusion of tregs makes administration of a high dose of t cells feasible for the fi rst time donor nkt cells and outcome following hla-identical peripheral blood stem cell transplantation d. nachbaur oral session : myelodysplasia / regulatory issues and quality management o monosomal karyotype, higher blast count and ex vivo t-cell depletion predict poor outcome after allogeneic stem cell transplantation for mds/saml patients with chromosome abnormalities m. van gelder, j. schetelig, l. volin, j. maertens, g. socié, e. petersen, h. thomssen, a. biezen, r. brand, t. de witte, n background: high-risk mds/saml patients with a chromosome abnormality have a poor prognosis with conventional therapies. these patients usually proceed to allogeneic sct (allo-sct). data on the effect of this approach are scarce. objective. description of outcome and identifi cation of risk factors of allosct in mds/saml patients with a chromosome abnormality. methods: from the ebmt database patients with mds/saml having any chromosome abnormalities (median age years) who underwent fi rst allosct between and november were identifi ed. stem cells from related donors were transplanted in patients ( hla-identical) while received unrelated grafts. bone marrow was used as stem cell source in patients. standard conditioning was applied in patients. sixty-three patients fulfi lled the criterion for complex cytogenetic abnormalities (ca). a monosomal karyotype (mk), defi ned as at least one autosomal monosomy and at least one other chromosomal abnormality (breems da et al., jco ; : ) , was present in patients, of whom did not have ca. results: median follow-up (fu) of patients alive at last fu was years (range - years). estimate -year overall (os), and relapse-free (rfs) survival was ± % and ± % respectively. the relapse rate at , and months was ± %, ± % and ± % resp. non-relapse mortality at , and months was ± %, ± % and ± % resp. in multivariate models including age, mk, ca, % blasts at allosct, donor type, sex match, conditioning regimen, t-cell depletion and year of allosct three factors remained statistically signifi cant predictors for poor outcome: mk, ≥ % blasts at allosct and ex vivo tcd (adjusted hr for os resp. . [ . - . %ci], . [ . - . %ci] and . [ . - . % ci] resp.). when patients with ex vivo tcd were excluded, -year os in the remaining mds/saml patients with any chromosome abnormality and < % blasts at allosct with or without mk was ± % and ± % resp. and for the patients with ≥ % blasts -year os was ± % and ± % resp (see figure) . conclusion: this large study on outcome of allosct for patients with mds/saml and any chromosome abnormality shows that long-term survival can be achieved in patients without the poor risk factors mk, ≥ % blasts at allosct and ex vivo tcd. for patients with mk new approaches that tackle the high and early relapse rate are warranted. bone marrow fi brosis on outcomes of patients with mds/saml undergoing allogeneic stem cell transplantation n. kröger, t. zabelina, a. van biezen, r. brand, t. bone marrow fi brosis has an important impact on the prognosis of patients with mds. we evaluate the impact of bone marrow fi brosis in patients who underwent allogeneic hematopoietic stem cell transplantation (hsct) for mds/aml and were reported to the ebmt. no fi brosis was noted in pts, mild or moderate fi brosis in pts and severe fi brosis in pts. diagnosis in the none, mild/moderate and severe fi brosis group were ra/rars ( %, % and %), raeb ( %, % and %) and raeb-t ( %, % and %). stem cell source were from related ( %, % and %) or unrelated ( %, % and %) donors. leukocyte engraftment (> . × /l) was observed after a median of , and , days for the days for the none, mild/moderate and severe fi brosis group, respectively (p = . ). there was a trend for more graft failure in severe fi brosis group ( %, p = . ). in a multivariate analysis bone marrow fi brosis did not infl uence non-relapse mortality, but severe fi brosis signifi cantly infl uenced risk of relapse (hr . , p = . ), resulting in a reduced disease-free (hr . , p = . ) and overall survival (hr . , p = . ). other factors for reduced survival were advanced disease at transplant (hr . , p = . ), non-cr before transplant ( . , p = . ) and hla-mismatch (hr . , p = . ). in summary, apart of the disease status, no complete remission at transplant, and hla mismatch transplantation, severe bone marrow fi brosis is an independent prognostic factor for disease-free and overall survival for mds patients undergoing allogeneic stem cell transplantation. management and outcome of myelodysplastic syndrome (mds) and secondary acute myeloid leukaemia relapsing after allogeneic stem cell transplantation -a retrospective analysis on patients by the mds subcommittee of the ebmt chronic leukaemia working pa c. schmid, a. van biezen, r. brand, j. finke, l. volin, a. vitek, d. selleslag, d. heim, p. van dem borne, a. ganser, m. mohty, m. boogaerts, t. de witte, n the aim of this study was to obtain reliable data on management and outcome of relapse after allogeneic sct for mds and saml in adults. median age of patients was years. patients (pts.) had been transplanted for ra/rars ( %), raeb ( %), raeb-t ( %) and saml ( %), either previously untreated ( %), in cr ( %), relapse/progression ( %) or primarily refractory ( %). standard (sic) and reduced intensity (ric) transplants were performed in % and %, donors were id siblings ( %), mud or mm relatives ( %), and syngeneic twins. median time from sct to relapse was mo . median follow up from relapse of survivors was . mo. overall survival (os) at , and years was %, % and %. in a cox regression model, the mds subtype at time of transplant (ra/rars vs. raeb [hr , , ci , , ci , vs. saml hr = , , ci , , p< . ) , and the interval from sct to relapse ( st vs. nd [hr = , , ci , ] vs. rd [hr = , , ci , ] vs. th quartile [hr = , , ci: , - , ], p< . ), were associated with survival. in pts., data on the management of relapse were available. pts had received dli. among them, median time from sct to relapse was . mo, median time from relapse to dli was mo. , and patients received , , and dli. median os from relapse was . mo. again, remission duration after sct (p<. ) and less advanced disease at time of sct (p = . ) were associated with better os. constitutional variability in genes involved in innate immunity (irf- , hamp, ptx ) and in cell proliferation (atbf and nfat ) infl uences disease free survival after allogeneic stem cell transplantation b. martín-antonio, i. alvarez, f. márquez-malaver, p. trujillo, m. carmona, j. falantes, i. espigado, Á. urbano-ispizua hospital universitario virgen del rocío (seville, es) genes involved in innate immunity and in regulation of cell proliferation may play an important role in infections and modulating the intensity of the infl ammatory response after allogeneic stem cell transplantation (allo-sct). thus, genetic variability in donor and recipient in these genes might be an important factor infl uencing the outcome of allo-sct. we have studied the potential infl uence of single nucleotide polymorphisms (snps) in donor and recipient in genes of innate immunity (irf- , hamp, ptx , hbd ) and regulation of cell proliferation (atbf , nfat , akt , nm , cd , tcirg , sh kbp ) on clinical outcomes after allo-sct, specifi cally on the incidence of acute gvhd, transplant related mortality (trm), relapse, and disease free survival (dfs). study population consisted of donor-patient pairs undergoing hla identical sibling allo-sct in a single institution. patient median age was years (range, - ). % of the patients were in advanced phase of disease. cumulative incidence for acute gvhd, trm, relapse and dfs was computed with the cmprsk package and with kaplan-meier. patient irf rs aa, donor atbf rs aa and patient akt rs cc dominant genotypes, were associated with a higher incidence of relapse (p = . , p = . and p = . , respectively) and lower dfs (p = . , p = . and p = . , respectively). all of them retained signifi cance at multivariate analysis. variant rs aa in atbf showed the same prognostic values when present in donor or in patient (relapse: % vs. % and dfs: % vs. %). when it was present both in donor and patient, the differences were more prominent (relapse: % vs. %, p = . and dfs: % vs. %, p = . ). donor hamp rs ag genotype and donor nfat rs aa dominant genotype were associated with a lower and a higher incidence of trm (p = . and p = . , respectively) infl uencing in a higher and lower dfs (p = . , p = . , respectively). they retained its signifi cance at multivariate analysis. nfat is necessary for optimal t cell development and rs aa variant showed the highest mrna expression. interestingly, none of the patients with a donor carrying ptx rs aa recessive genotype had trm but showed a higher tendency in the incidence of relapse ( %). in conclusion, genetic variability in innate immunity and in cell proliferation has a strong infl uence on the clinical outcome of allo-sct, which might be important when choosing allo-sct protocols. the factors to predict adverse events occurred within days of post-peripheral blood stem cell donation at family donors y. kodera, s. kim, k. nagafuji, m. hino, k. miyamura, r. suzuki, a. tamakoshi, m . fukushima on behalf of the japan society for hematopoietic cell transplantationbackground: it has become obvious that certain adverse events might occurre at peripheral blood stem cell (pbsc) donors during or after the donation process. it is therefor crucial that certain factors which can predict the occurrence of such adverse events at normal donor are cralifi ed to prevent the adverse events. the jshct has continued the nation-wide consecutive follow up of pbsc family donors and in this project, the day report of post donation was included. this time, we present the outcome of the day report analysis. materials and methods: among , pbsc family donors who were consecutively pre-registered to jshct donor center between april and march , , day reports were obtained and subjected for analysis. the relationship between family donors' ) gender, ) age, ) body-weight, ) daily and ) total dose of granulocyte-colony stimulating factor (g-csf), ) current and ) past health conditions and the occurrence of ) thrombocytopenia, ) prolongation of hospitalized period, ) any adverse events (bone pain, fatigue, head ache, insomunia, anorexia, nausea, vomiting, splenomegary) as well as the mobilized cd + cell numbers were statistically examined. results: risk factors for thrombocytopenia were higher total dose of g-csf and older age. risk factors for prolongation of hospitalized period were higher total dose of g-csf, any present illness, older age and low body-weight. the risk factor for bone pain was higher body-weight. the risk factors for fatigue were female and lower body-weight. the risk factors for insomnia were older age and female and the risk factors for anorexia were female and low body-weight. predictive factors for lower cd + cell mobilization/donor's body-weight were older age and higher total g-csf administration, and predictive factors for higher cd + cell mobilization were male and younger age. discussion: it was revealed that certain adverse events which occurred within days of post-donation and cd + cell numbers to be mobilized could be predicted by utilizing the basic information of donors and of pbsc mobilization/harvest process. to predict them, to notify them to donors and to prepare for possible events will contribute to keep pbsc donor's safety and trust. background and aim: allogeneic hematopoietic stem cell transplantation (hsct) has become an established therapy and the numbers of such procedures increase year by year. the risk for while / pts developed chronic-gvhd (extensive: pts). in the multivariate analysis only acute-gvhd grade ii-iv and extensive chronic-gvhd were associated with a lower os. pts developed oral squamous cell carcinoma (scc) between - ys after bmt (all had c-gvhd) and pt died. pts died between - days after bmt. major causes of death were related to rejection or gvhd. trm at days: % and at y: %. conclusions: this regimen was well tolerated and had an excellent survival especially for pts below the age of . long term follow up is still needed to determine the incidence of cancer in this group of pts. survey of outcomes of unrelated cord blood transplant in patients with haemoglobinopathies: a retrospective study on behalf of cibmtr, nycb and eurocord a. ruggeri ( ) allogeneic hematopoietic cell transplantation (hct) from hla-identical donors is curative in patients with hemoglobinopathies. in order to extend the availability of hct, unrelated cord blood transplantation (ucbt) has been investigated as an alternative. between and , patients receiving a single ucbt for hemoglobinopathy were reported to the eurocord and cibmtr registries. thirty-four had thalassemia major (thal) and had sickle cell disease (scd). all thal patients were transfusion-dependent with a median time from diagnosis to ucbt of months and of patients with scd had a history of stroke. median age at ucbt was years and median follow-up was years. cord units were matched at of ( %), of ( %) and of ( %) hla loci (antigen-level for class , allele-level for class ). median infused cell dose was . × /kg ( . - ). thirty-nine patients received a myeloablative conditioning regimen with busulfan (bu) and cyclophosphamide (n = ) or bu with other agents (n = ). reduced intensity conditioning regimens (ric) (n = ) were fl udarabine-based with bu < mg/kg or melphalan < mg/m². cumulative incidence (ci) of neutrophil recovery at d was ± %, with of patients reaching neutrophil recovery at a median time of days. higher cell dose (> . × /kg) ( % vs. %, p = . ) and ucbt performed after ( % vs. %, p = . ) were associated with improved neutrophil recovery. day- chimerism analysis was available for patients: patients had complete donor chimerism, mixed chimerism and autologous recovery. among patients who did not achieve neutrophil recovery, had available data on subsequent treatment. two had an autologous back-up and one received a second ucbt which engrafted. ci of grade ii-iv acute-graft versus-host disease (gvhd) was % and patients had chronic gvhd. the -year probability of overall survival was %. thirty-eight patients ( thal, scd) are alive, with full donor chimerism ( thal, scd). of the deaths, occurred prior to day- , mainly due to infections. no deaths due to gvhd were reported. ten of patients who received ric are alive, with donor engraftment. despite the small number of subjects, these results show a particularly high risk of graft failure after ucbt for hemoglobinopathy. tnc dose plays a key role in engraftment. novel approaches modifying the conditioning regimen and/or increasing cell dose in prospective clinical trials are needed. graft rejection and second haematopoietic cell transplantation in patients with thalassaemia major s. santarone, e. di bartolomeo, p. bavaro, p. di carlo, p. olioso, g. papalinetti, p. di bartolomeo bmt center (pescara, it) graft rejection for patients with thalassemia major (tm) receiving haematopoietic cell transplantation (hct) includes early graft failure (gf) (no evidence of engraftment), late gf (lost of engraftment), and recurrence of tm. we examined our series of consecutive patients aged to years (median years) with tm who were transplanted either from hla-identical related (n = ) or unrelated (n = ) donor. conditioning regimen consisted of busulfan (bu) and cyclophosphamide (cy). graftversus-host disease (gvhd) prophylaxis included cyclosporine (csa) and methotrexate (mtx). all patients received bone marrow (bm) cells. the -years overall cumulative incidence of graft rejection was % + . . the impact of pre-transplant patient risk factors (age, gender, number of transfusions, ferritin, splenectomy, ast, alt, hepatomegaly, hbv and hcv infection, liver fi brosis, cmv serology), donor characteristics (age, heterozygous state) and hct regimens (dose of bu, gvhd prophylaxis, bm dose) was studied in univariate analysis. no factor was univariately associated with signifi cantly increased probability of graft rejection. eleven events occurred: early gf, late gf, and recurrences of tm. patients with either early or late gf underwent an urgent second hct, whereas of patients with recurrence of tm choose to be submitted to second hct. at all, patients received a second hct from the same donor. their median age was years ( - ). conditioning regimen for second hct was immunosuppressive for patients (atg in addition to fl udarabine (flu) or cy) and myeloablative for patients (bucy in and thiotepa (th), treosulphan (treo) and flu in ). gvhd prophylaxis consisted of csa alone for patients and csa /mtx for patients. six patients were given bm cells and received peripheral blood stem cells. results: patients died from hct related causes (heart failure at day and acute gvhd at day , respectively). seven patients ( %) are living. one patient had no sign of engraftment and is now living with recurrence of tm following a third hct with double unrelated cord blood cells. tm recurred in an other patient at months post-second hct. this patient is now living with transfusion therapy. five patients ( %), including the patients transplanted with th-treo-flu regimen, are cured with a median follow up of years ( , - ). this study shows that the incidence of graft rejection following hct for patients with tm is low and second transplant is successful in an high proportion of patients. treosulfan is a bi-functional alkylating agent which causes less vod than busulphan and does not require anticonvulsant prophylaxis and drug monitoring. we report the use of treosulfan in children undergoing hct for primary immunodeficiency (pid) at two supraregional transplant centres in the uk. children received g/m² or g/m² treosulfan in combination with either cyclophosphamide mg/kg ( ) or fl udarabine autoimmunity is a complication after haematopoietic stem cell transplantation in patients with primary immunodefi ciencies (pid) that sometime occurs when bmt is performed from alternative donors with extensive stem cell manipulation. the fact that only purifi ed stem cells (cd positive cells) are injected could lead to a slower and incomplete post-transplant immunological reconstitution. since the majority of children treated in our unit received haploidentical cd positively selected grafts, we retrospectively analysed the population of pid patients grafted in our institution between and . of the affected by severe combined immunodefi ciency (scid), received an haploidentical transplant, while the others received a matched unrelated or a hla-identical transplant. patients were affected by inborn errors; received an haploidentical transplant, the others received a matched unrelated or a hla-identical transplant.we analysed the incidence of autoimmunity in these two groups of patients. out of children, experienced autoimmune symptoms after hsct. among this group, had a diagnosis of scid, of leaky scid, of omenn's syndrome, one of chediak-higashi and one of wiskott-aldrich syndrome. autoimmune symptoms included autoimmune haemolytic anaemia ( ), autoimmune hypotiroidism ( patients), cutaneous autoimmunity ( ), vasculitis ( ) and chronic bronchoreactivity. all patients except for received a myeloablative conditioning therapy. of them received an haploidentical bmt and a mud or a hla-identical bmt. no correlation was found analysing onset of autoimmunity and split chimerism in all groups of patients nor we could fi nd a correlation with immunsuppressive therapy. in conclusion we found a low incidence of autoimmunity despite the fact that the majority of our patients received a cd positive selected graft with an even distribution of the complication independently from the type of donor or stem cell manipulation procedure. introduction: there is still a discussion about the best bm harvesting method to obtain enough progenitor cells during the bone marrow (bm) harvesting procedure. the technique used in most bmt centers is the multiple aspiration of a small ( ml) amount of bm from the iliac crest, sternum or tibia. this is proposed to minimize the dilution with peripheral blood. others are using few large amount aspirations. this procedure was not evaluated in children so far. to fi nd out possible differences in graft composition between this two methods and to evaluate the feasibility in children we initiated the following prospective study. patients and methods: bm harvestings were done in patients (median age , , ( , - , y), f , m ). all patients were transplanted in our bm transplantation unit. there were bm harvests for autologous bmt and for allogeneic bmt. the autologous donors were median age , ( , - y), f , m ), the allogeneic donors were median age , ( , - y), f , m ). the amount of harvested bm was median ml ( - ml) and median ml/kg bw of the recipient ( - ml/kg). the method a was defi ned as an aspiration of ml bone marrow at most before the position of the needle was changed. the method b was defi ned as an aspiration of ml at least before the location of the needle was changed. the bm was collected and analyzed among others for mnc, cd positive progenitor cells, cfu, and t-cells. two operators carried out the bm harvest. one began with method a and changed in the second part to method b. the second operateur carried out the procedure in the opposite way. both operators collected the same amounts for each harvesting method so long that the amounts of bm harvested in one method was identically with the second. results: we found no statistically signifi cant difference between method a and b regarding the content of mnc, cd + t-cells, and cfu (mnc/ml a versus b wilcoxon test p = . ; mnc/kg . e a versus . e , p = . ; cd /ml a versus , p = . ; cfu/ e mnc a versus b, p = . ), but a clinically not relevant but statistically signifi cant difference between cd positive cells (cd /kg . a versus . b, p = . ). all transplanted patients showed a regular engraftment. conclusion: bm harvest by large amount few punctures method (b) is as suffi cient as the common used small amount frequent punctures method (a), and could be therefore used equally. haploidentical transplantation, with extensive t cell depletion to prevent gvhd, is associated with a high incidence of infectionrelated deaths. the key challenge is to improve immune recovery with allogeneic donor t cells without triggering gvhd. as t regulatory cells (tregs) controlled gvhd in preclinical studies, background: natural killer t (nkt) cells represent a small but signifi cant lymphocyte population which have been demonstrated to play a regulatory role in autoimmune disease as well as in gvhd/gvl effects. methods: the number of cd + cd + nkt cells in the graft was retrospectively correlated with clinical outcome of fi fty-eight patients receiving a fi rst allogeneic peripheral blood stem cell transplant (myeloablative conditioning, n = ; reduced-intensity conditioning, n = ) from their hla-identical sibling donors between dec and jul . results: a median number of . (range, . - . ) × /kg cd + cd + nkt cells was transplanted with the graft. nkt cells within the graft signifi cantly correlated with the number of cd + t cells, cd + nk cells, and mnc (p< . ). overall survival for the entire cohort at two years was % ( %- %, % ci) with no difference between pts. receiving a high (> . × /kg) or low (< . × /kg) nkt cells. the cumulative relapse incidence for the entire cohort was %, with a trend for a lower relapse incidence in pts. receiving a low vs. high nkt cell dose ( % vs. %). the nonrelapse mortality was % showing a trend for a lower nrm in patients receiving a high nkt cell dose ( % vs. %). the cumulative incidence of agvhd ii-iv was % for the entire cohort. the incidence of agvhd ii-iv was lower in pts. receiving a high nkt cell dose ( % vs. %). the cumulative incidence of cgvhd in pts. at risk was %, with no difference between pts. receiving low or high nkt cell numbers. by multivariate cox analysis including cd , nk, nkt, treg, cd cell number, recipient age, and time interval between diagnosis and sct as numerical variables, and risk category by the underlying disease (standard risk vs. high risk), sex mismatch (male recipient/female donor vs. other combinations), and myeloablative vs. reducedintensity conditioning as categorical variables the most powerful predictive parameters for survival were standard vs. high-risk disease (p = . ), a high nk (p = . ) and a low nkt cell dose (p = . ). the most powerful predictive parameters for relapse were high risk disease (p = . ), a low nk (p = . ) and a high nkt cell dose (p = . ). discussion: these data indicate that in the setting of peripheral blood stem cell transplantation from hla-identical sibling donors a higher number of nkt cells in the graft might increase the risk of relapse by lowering the incidence of acute gvhd. objectives: adoptive transfer of nk cells with or without previous allogeneic progenitor cell transplantation may represent a promising therapeutic option in patients with hematological or oncological diseases. different nk cell isolation strategies have been pursued. here, two different methods were compared with respect to recovery, immunophenotype and cytotoxic capacity of purifi ed nk cells. methods: nk cells of healthy donors were isolated from a lrs chamber of a platelet apheresis. cells were enriched by (i) cd positive selection of t cell (cd ) depleted lymphocyte fractions (method i, mi) or (ii) depletion of non-nk cells (untouched protocol, method ii, mii) by magnet-activated cell sorting (macs). recovery of nk cells obtained by m i or ii were compared. nk cells derived by mi were activated by stimulation with interleukin (il)- ( u/ml) or il- ( ng/ml) overnight and analyzed for cytotoxic activity. for expansion of nk cells cd /cd antibody coated macsibeads (miltenyi) were used as artifi cial stimulators in il- supplemented media ( u/ml) in a two weeks culture. finally, immunophenotype of effector cells and killing of target cells were assessed. results: median nk cell recovery was . % (n = ) and independent of strategy of enrichment (mi versus mii). overnight activation of nk cells led to a -fold enhanced killing of k cells. nk cell expansion led to a -fold increase in numbers of functional active nk cells. expanded nk cells showed characteristics of activated nk cells as indicated by the induced expression of nkp (cd ). surface expression of tac-tile (cd ), a marker for certain activated immune cells, was found with cd antibody th- elevated on bead-expanded nk cells in comparison to nk cells cultured only in il- conditioned media, which may enhance reactivity of expanded nk cells against the ligand of cd , namely polio virus receptor expressing tumours. conclusion: t cell depletion and cd positive selection led to an acceptable recovery of enriched nk cells that could be successfully activated by il- or il- . the described nk cell isolation and activation protocol can easily been transferred into a gmp laboratory. for clinical application, further expansion of isolated nk cells under gmp conditions may be preferable. nk cells enriched, expanded and activated by these means may be an adoptive cellular therapeutic option that in addition may be improved by combination with tumour antigen specifi c antibodies. s o a randomized trial comparing cd enriched grafts versus cd /cd depleted grafts in partial t-cell depleted allogeneic stem cell transplantation n. schaap, d. eissens, f. preijers, a. van der meer, a. schattenberg, h. dolstra, w. van der velden, t. de witte, m. smits, n.m.a. blijlevens radboud university medical centre (nijmegen, nl) we initiated a randomized trial in aml, all and mds patients (pts) treated with partial t cell depleted allogeneic sct using immunomagnetic cd selection versus cd /cd depletion. a benefi cial effect was hypothesized on long term engraftment and dfs using cd /cd depletion. residual cell populations in the graft after cd /cd depletion may play a protective role during early phase after sct and generate an increased gvl effect. from may until may pts with aml, all and mds were stratifi ed for diagnosis and randomized. median age of the pts was and yrs, respectively. stem cells were obtained after g-csf ( ug/kg). t cell add back to a fi xed dose of × e cd + t cells/kg bw was standard in all pts. pts without significant (> grade ) acute and/or chronic gvhd were eligible for prophylactic dli. in this interim analysis we determined immune reconstitution of t cells, b cells and nk cells using fl ow-cytometry and functional essays and evaluated clinical outcome. reconstitution: nk cells are completely maintained in cd / cd depleted transplants. b cell depletion is equally effective. cd + cells are signifi cantly higher in the cd /cd depleted grafts (median . versus . × e /kg, p< . ). all pts engrafted and the time to engraftment was not different. in the fi rst months after sct, the immune reconstitution of lymphocytes, especially cd t cells, nk and nk-t cells was faster in the cd /cd group. fourteen days after sct the cytotoxic nk and regulatory nk subsets were already present in the cd /cd group whereas the cd group needed months to full recovery. the cytolytic response against a leukemia target was stronger in the cd /cd group (p = . ). nk receptor expression of nkg a (inhibitory) was strongly decreased whereas expression of nkg c (activating) was enhanced. clinical outcome; acute gvhd > grade was % in both groups. extensive chronic gvhd was % (cd ) vs. % (cd /cd . one year trm, rr, lfs and survival using cd selection and cd .cd depletion were % vs. % (p = . ), % vs. % (p = . ), % vs. % (p = . ), vs. % (p = . ), respectively. conclusion: compared to cd selection, cd /cd depletion resulted in a faster reconstitution of cd cells,nk-t and nk cells. however survival was signifi cantly better in pts transplanted with cd selected grafts. due to a loss of power in favor of our hypothesis ( % increase in overall survival) using statistical simulation models, this trial was stopped. ( ) introduction: in haploidentical hematopoietic stem cell transplantation (hsct), the infusion of donor lymphocytes transduced to express the herpes simplex virus thymidine kinase (hsv-tk) suicide gene allows to control gvhd, to mediate gvl, and to rapidly provide an effective and polyclonal anti-infective t cell repertoire (ciceri and bonini et al., lancet oncology, ). even though their engraftment is necessary to achieve these effects, hsv-tk + cells represent the minority of lymphocytes circulating in treated patients. therefore, we investigated the putative role of hsv-tk + cells in promoting thymic activity and t cell development from graft progenitors. methods: thymic function was assessed in adult patient who underwent haploidentical hsct and infusion of suicide genemodifi ed donor t cells for hematologic malignancies, after validating the methods in healthy pediatric and adult controls. single joint t cell receptor excision circles (sjtrec) were quantifi ed by qpcr in peripheral blood mononuclear cells (pbmcs) and purifi ed t cells, and the proportion of cd + recent thymic emigrants (rtes) in cd + naïve t cells was measured with immunophenotype analysis in pbmcs. thymic output was correlated with thymic volume, assessed by computed tomography (ct) scans. t cell receptor repertoire was assessed by vbeta spectratyping. the relative contribution of hsv-tk + and hsv-tk-donor t cells to post-transplantation anti-host alloreactivity was studied by mixed lymphocyte cultures. results: at the moment of t cell immune reconstitution (defi ned as cd + cells > /μl peripheral blood), the cd + naïve t cell subset was almost entirely comprised by cd + rtes, and this percentage remained high, as compared to age, also in subsequent months. in informative patients, rte frequency before hsct and before hsv-tk + cell infusion was in line with agerelated healthy controls, suggesting a direct role of the infused cells in mediating the effect. accordingly, in treated patients ct scans documented an increase in thymic volume following hsv-tk + cell add-backs. finally, low absolute sjtrec counts could be detected, in line with the documentation of extensive peripheral proliferation and low rte absolute numbers. conclusions: these data show that after the infusion of suicide gene-modifi ed t cells a renewal of thymic activity takes place, which contributes to the recovery of the peripheral t cell repertoire. elevated pretransplant serum ferritin levels have been associated with an increased susceptibility for opportunistic infections and increased incidence of morbidity and mortality after allogeneic hct. we studied in patients who underwent myeloablative allogeneic hct for acute myeloid leukemia pre-and posttransplant serum ferritin levels and correlated the serum ferritin levels with the tlr expression and the cellular immune reconstitution and months post transplant. further, we studied in vitro-experiments in kasumi cells the tlr , tlr , tlr , tlr , tlr , tlr and tlr expression after overwhelming iron exposure. the average pretransplant serum ferritin level was μg/ml (mean) in all aml-patients (mean μg/l for patients with aml in .cr and mean μg/l for patients with aml > .cr). post transplant serum ferritin level increased up to μg/ml (mean) for all aml patients (mean mg/l for aml in .cr and mean μg/l for patients with aml > .cr). the application of ng iron to acute leukaemia cell lines sd , and kasumi- cells increased signifi cantly tlr ,tlr , tlr , tlr , tlr and tlr expression in relation to the housekeeping gene abl measured by real-time rt-pcr. in kasumi- cells tlr increased up to , % (p = . ) tlr . % (p = . ), tlr , % (p = . ), tlr . % (p = . ), tlr . % (p = . ), tlr . % (p = . ) and tlr , % (p = . ) compared to untreated kasumi cells. further, patients with elevated post transplant ferritin level > μg/l had an increased tlr expression in mononuclear cells (tlr /abl quotient versus ; p< . ) months post transplant. the number of cytotoxic t cells cd + cd + in patients with elevated serum ferritin level was signifi cantly elevated after transplant (mean versus cells/μl months post transplant, p = . ), whereas no differences were found in the number of cd + cd + t helper cells, b + cells, nk cells. these results indicate that elevated ferritin levels might activate the innate immune system by increasing tlr expression. this might be of importance since we recently showed that increased tlr expression was associated with adverse impact on nonrelapse mortality in the transplant setting. further exaggerated tlr expression has been discussed to induce overwhelming immune responses as sirs or ards. more studies are definitely necessary to evaluate the role of elevated iron overload on the innate immune system. clinical scale generation of functional human natural killer cells from umbilical cord blood cd -positive cells for immunotherapy j. spanholtz, m. tordoir, c. trilsbeek, j. paardekooper, t. de witte, n. schaap, f. preijers, h. dolstra umc st.radboud (nijmegen, nl) immunotherapy based on natural killer (nk) cell infusions is a potential adjuvant treatment for many cancers. we established an extremely effi cient cytokine-based culture system for ex vivo expansion of nk cells from hematopoietic stem and progenitor cells from umbilical cord blood (ucb). systematic refi nement of this two-step system using a novel clinical grade medium resulted in a therapeutically applicable cell culture protocol. the use of gbgm culture media in a clinical applicable protocol allows the ex vivo expansion and differentiation of cd + cells to more than -logs into cd + cd -nk cells with very high purity. these ex vivo-generated cd + cell products contain nk cell subsets expressing cd /nkg a and kir as well express high levels of activating nkg d and ncrs. functional analysis showed that cd + cell products containing alloreactive nk cells effi ciently kill myeloid leukemia and melanoma tumor cells as well as primary acute myeloid leukemia (aml) cells. we have currently translated the protocol to clinical scale production using a closed cell culture bioprocess ( figure ). cd + selection from cryopreserved ucb samples (n = ) using the clinimacs device was optimized and the selection resulted in a cd + enrichment with a mean number of , ± , × cells and a purity of ± % cd + cells. validation experiments using these cells showed, that the generation of suffi cient numbers of nk cells without contaminating t-cells or bcells under a closed system process is feasible. the cell cultures using bags show a mean expansion of ± fold (range - fold, which generated , × - , × nk cells from cord blood-derived cd + cells within maximal weeks of culture. the mean purity of the nk cell product was ± % (range - %) of the total cells in the bag cultures. in order to improve the purity of the product, we include bioreactors during the differentiation culture process. using this modifi cation we were able to synchronize the differentiation process in small (plate) and large scale (bag) experiments, which allows the generation of large scale nk scale products with a purity of more than % cd + cells devoid of t-and b cells (figure a and b) . these nk cell products will be used for immunotherapy in elderly aml patients. this study is a phase i dose escalation study in a series of aml patients who have successfully achieved cr (< % blasts in the bone marrow) after standard intensive chemotherapy. key: cord- -z ehfrg authors: barton, todd d.; blumberg, emily a. title: viral pneumonias other than cytomegalovirus in transplant recipients date: - - journal: clin chest med doi: . /j.ccm. . . sha: doc_id: cord_uid: z ehfrg community-acquired respiratory viruses (carvs) are frequent causes of upper and lower respiratory tract infections in tranplant recipients. in most series, respiratory syncytial virus and parainfluenza are the most common carvs, followed by influenza and adenovirus. significant morbidity and mortality are associated with these infections, particularly when they progress to pneumonia or when they are associated with bacterial or fungal coinfections. outcomes are also poor with adenovirus, frequently reflecting disseminated infection. efforts to prevent morbidity and mortality from carv infection should focus on prevention, because treatment options are limited with inconclusive data to support their efficacy. community-acquired respiratory viruses (carvs) are frequent causes of upper respiratory infection (uri) in adult and pediatric populations, usually occurring in seasonal outbreaks. in healthy outpatients, the morbidity caused by these infections is minimal, because progression to lower respiratory tract infection (lrti) is rare, and most infections are selflimited in duration. although case reports of viral pneumonia complicating hematopoietic stem cell transplantation (hsct) or solid organ transplantation (sot) have been described for decades, it is only in recent years that larger case series and therapeutic trials have been conducted and reported, providing greater insight into the impact of carv on these immunosuppressed hosts. after some general observations about carv infections, this article focuses on this important recent literature and specifically on the four most common pathogens, respiratory syncytial virus (rsv), influenza virus, parainfluenza virus (piv), and adenovirus. it concludes by briefly touching on several less commonly reported causes of viral pneumonia, including some potentially important emerging pathogens. although dozens of published studies have described the epidemiology of some or all of the carvs, their findings are often widely disparate. this differences in part result from the nature of the diseases, because both their seasonality and relative frequencies may vary depending on the climate of the reporting institutions. similarly, studies that track only a single year's incidence of the carvs may over-or underestimate the general relative frequency of the pathogens based on a particularly widespread epidemic of a single viral pathogen, as might be seen in a year with an especially widespread influenza epidemic. table reviews the relative frequencies of the carvs in several recent reports. depending on the center and the year, rsv, piv, or influenza has been the most common pathogen, whereas adenovirus generally accounts for fewer than % of carv infections [ - ] . finally, studies that include children may report higher rates of carv infections than those focusing on adult populations, probably reflecting in part the higher carriage of carvs in children. most commonly, investigators have employed two major strategies to gain an understanding of the general epidemiology of respiratory virus infections. in the first, consecutive tranplant recipients have been screened at regular intervals, usually in the first to months after transplantation, regardless of symptomatology. results of such studies have shown overall incidence rates of carv infection in hsct recipients ranging from % to % [ - , ] . in contrast, a recent prospective surveillance study in sot recipients showed only a % incidence of carv in adult liver transplant recipients during the first weeks after transplantation, although interpretation of this study is limited by the investigators' use of throat swabs alone to detect carv infection [ ] . more frequently published are large retrospective case series of carv infections. because these series do not include patients who had asymptomatic infection, overall reported rates of carv infection are predictably lower, ranging from % to % in hsct recipients [ - , ] to % to % in lung transplant recipients [ , , ] . although these larger reported series represent the best estimates of the carv disease burden in the general transplant population, it is important to remember that the reports are biased by the seasonal occurrence of carv in both nosocomial and community settings and the potentially devastating impact of these infections on hsct and sot patients [ ] . a major limiting factor in the understanding of carv infection has been the limited sensitivity of what currently are the most widely used diagnostic tests. in three patient series involving more than episodes of symptomatic uri in hsct recipients [ , , ] , fewer than half the patients had a virus isolated from clinical specimens. most studies (and clinical centers) use a combination of direct (dfa) or indirect (ifa) fluorescent antibody testing and viral culture. results of fluorescent antibody testing are typically available in about hours, but viral culture may not be positive for to days. in children, these tests are often performed on samples from nasopharyngeal lavage. in adults, swabs from the nasopharynx or throat often are substituted. whereas the dfa or ifa tests may have a sensitivity of up to % for carv infection in immunocompetent hosts, two studies comparing ifa and viral culture in hsct recipients have shown a composite ifa sensitivity of % [ - , , ] . palmer and colleagues [ ] reported a dfa sensitivity of % in their series of lung tranplant recipients who had carv infections. both fluorescent antibody testing and viral culture probably have higher yields when the sample is obtained from the lower respiratory tract. in one recent study, two thirds of carv diagnoses were made from bronchoalveolar lavage (bal) samples [ ] . recently, two series have used real-time polymerase chain reaction (pcr) assays to test for carv infections in hsct recipients [ , ] . in one series of adult hsct recipients being monitored with routine nasal and throat swabs over a -month period, real-time pcr was positive in patients, whereas viral culture was positive in only [ ] . many of the additional positive tests were asymptomatic patients who had rhinovirus infection. bredius and colleagues [ ] tested children with symptomatic respiratory tract infection; ifa was positive in ; viral culture was positive in ; and real-time pcr was positive in . viruses, perhaps especially piv, are often copathogens with other bacterial or fungal infections [ , ] . infection with both a carv and cytomegalovirus, aspergillus species, or pneumocystis jiroveci have been described in up to % of carv pneumonias [ , ] . conversely, because providers may halt a diagnostic work-up after isolation of a single bacterial or fungal pathogen, it is not known how often infection with these more traditionally oppor- tunistic pathogens is complicated by coinfection with a carv. the overall reported mortality from carv infection has varied widely in published series. probably no mortality is associated directly with carv infection limited to the upper respiratory tract. in older case series of predominantly hospitalized hsct or sot recipients who had lrti, mortality was frequently greater than % [ ] . it can be difficult to compare some series based on varying definitions of lrti: in some series, a positive chest radiograph is required to define lrti or pneumonia, whereas in others series physical examination findings (eg, rales, hypoxia) consistent with lower tract disease have sufficed. most recent series in hsct recipients have included more outpatients who have uri and have shown much lower mortality, ranging from % to % [ , , , ] . raboni and colleagues [ ] in brazil reported % mortality from carv infection in a cohort of patients. many of these patients had received allogeneic bone marrow transplants within the previous year. fewer case series exist in sot recipients. palmer and colleagues [ ] reported % mortality in their series of carv in lung transplant recipients , but case series in renal and liver transplant recipients showed no mortality from the infection [ , ] . of particular recent interest has been the possible link between carv infection and the development of chronic rejection in lung transplant recipients. this link is supported by a mouse model in which piv infection aggravated chronic rejection of lung allografts [ ] . epidemiologically, the possibility that carv infection contributes to chronic lung transplant rejection manifesting as bronchiolitis obliterans syndrome (bos) has been supported by the observation that bos may have a seasonal pattern of onset [ ] . this observation has not been true in all studies, however [ ] . although bos occurs in up to % to % of lung transplant recipients after years [ ] , the overall incidence rates for each of the four most commonly reported carvs generally have not exceeded % in lung transplant cohorts [ , , - ] . as noted previously, this percentage may reflect a limitation in current diagnostics rather than a lower incidence of these infections. carv infection may produce wheezing and bronchospasm in immunocompetent adults, so it is not surprising that case series of carv infections in lung transplant recipients report a decline in perfor-mance on pulmonary function tests during the acute illness [ , ] . in two reports that have followed serial pulmonary function tests for several months after carv infection, there have been no significant changes in the pulmonary function testing after a few months' follow-up [ , ] , although one investigator reported impairment of pulmonary function persisting beyond days in % of patients [ ] . three reports demonstrated marked variability in rates of acute rejection at the time of carv infection. vilchez and colleagues [ ] reported allograft rejection in ( %) of patients who had piv infection, whereas two other series reported acute rejection in ( %) of and ( %) of patients, respectively, at the time of carv infection [ , ] . further study is needed to clarify this important disparity. two larger series specifically designed to investigate the link between any carv infection and bos bear closer analysis. khalifah and colleagues [ ] followed adult lung transplant recipients prospectively and identified carv infections. nearly all were lower respiratory infections found on bronchoscopy performed either for surveillance or in response to a new clinical syndrome. patients who had any history of carv infection in this cohort were more likely to develop severe bos ( % versus %), to die from bos ( % versus %), or to die from any cause ( % versus %) than were patients who had no history of carv infection [ ] . additionally, the authors note that of eight patients who had rsv infection, three who were treated with antiviral agents did not develop bos, whereas four of five patients not treated with antiviral agents did develop box. billings and colleagues [ ] followed adult lung transplant recipients and found carv infections in patients over an -year follow-up period. again, the majority of identified carv infections in this series were lrti, and many were identified from bronchoscopy specimens when bronchoscopy was performed for surveillance or follow-up of treatment for rejection. in this series, carv lrti was found to be predictive of severe (grade ) bos, but not of moderate (grade ) bos. the authors noted that bos was clearly a risk factor for carv [ ] . it is possible that the chronic rejection facilitates colonization, infection, or progression to lrti by carvs. it is also probable; however, that patients who have bos undergo more frequent bronchoscopy, making it more likely that carv infection will be identified. taken together, the reports from khalifah [ ] and billings [ ] strongly suggest an association between carv infection and bos. the significance and directionality of this association remain to be determined. epidemiology rsv occurs annually in late autumn or winter outbreaks in the general population, with a low level of persistent year-round activity. these same seasonal patterns have been found in hsct recipients [ , ] . like most carvs, rsv affects children more than adults, and this observation has been confirmed in single-center analyses where adult and pediatric hsct programs coexist [ ] . the epidemic nature of rsv infections must be stressed, because these outbreaks have been responsible for significant morbidity and mortality [ ] . several important factors may contribute to rsv outbreaks among hsct or sot recipients. on inpatient units, hsct or sot recipients tend to be housed on dedicated wards, thus exposing them to each other and also to a common group of hospital staff and care providers. in fall or winter seasons, % to % of these providers may shed rsv asymptomatically, and that number may increase to % during community outbreaks [ ] . additionally, hsct or sot recipients are more likely than immunocompetent patients to have prolonged shedding of rsv, thus introducing the potential for a single index case to infect many other patients [ , , ] . together, these factors may explain why multiple reported series of rsv infection note that more than % of cases were nosocomially acquired [ , , ] . among hsct and sot recipients, rsv is the most commonly reported carv in most series (see table ). large series of rsv infection have been reported in recipients of allogeneic hsct [ , - ] , autologous hsct [ , ] , liver transplants [ ] , and lung transplants [ , ] , in addition to case reports or small series from nearly all other transplant types. it is difficult, however, to compare the incidences reported in these trials directly, because study designs include longitudinal studies, single-year surveys, and prospective surveillance data. mccarthy and colleagues [ ] reported a cumulative rsv incidence of . % in allogeneic hsct recipients over a -year period. small and colleagues [ ] reviewed consecutive hsct recipients over a -year period and demonstrated an incidence of . % in allogeneic hsct recipients as compared with . % in autologous hsct recipients. other reports have confirmed this higher rate in allogeneic hsct recipients, including a multicenter european study that showed a single-year incidence of symptomatic rsv of . % in recipients of allogeneic hsct and of . % in autologous hsct recipients [ ] . longitudinal stud-ies of pediatric liver transplant recipients and adult lung transplant recipients showed incidence rates similar to the longitudinal studies, at . % and % to %, respectively [ , ] . rsv infection begins in the upper respiratory tract, with cough present in % to % of immunocompromised patients [ ] . most patients also report rhinorrhea or sinus congestion, and nearly half report subjective wheezing [ ] . although fever may be present in most immunocompromised patients, the prevalence of fever in hsct and sot recipients has not been well clarified; in one series, only % of patients who had rsv lrti were febrile [ ] . morbidity and mortality from rsv are directly attributable to progression of the infection to pneumonia. although bacterial infection may occur coincidentally with other carv infections, it is not seen frequently with rsv in hsct or sot recipients [ ] . many recent series have documented that about % of patients who have rsv infection develop pneumonia; some present with pneumonia, and others develop pneumonia after initial presentation with uri (table ) . nearly all patients who present with pneumonia give a history of several days of antecedent uri symptoms. once the infection progresses to pneumonia, mortality rates are high. the results of several recent reports are presented in table and show that ( %) of hsct recipients who had rsv pneumonia died [ , - , , , , , , , ] . diagnosis of rsv infection at most centers is done primarily by fluorescent antibody testing because of the wide availability of the tests (ifa or dfa) and the rapid turnaround time. in immunocompetent hosts, dfa or ifa for rsv may be up to % sensitive, whereas culture is only % to % sensitive [ ] . englund and colleagues [ ] , however, reported concerning data in immunocompromised adults who had hematologic malignancies, in whom dfa testing on specimens from throat swabs and nasopharyngeal washing had a sensitivity of only %. when applied to specimens from bal, the dfa was % to % sensitive [ ] . these results parallel those reported by billings and colleagues [ ] , in which % of rsv diagnoses were made from bal samples. the poor yield of dfa on easily obtained samples highlights the need for newer diagnostic tests or strategies. preliminary data suggest that real-time pcr may have a sensitivity of greater than % for rsv infection [ ] . there are no large series employing this diagnostic modality, however. as with other pcr testing, there may be issues with false-positive tests (in this case, defined as a positive pcr for rsv when rsv is not the true cause of the patient's symptomatology), and the definition of an appropriate criterion standard test may prove difficult in future studies. one recent series used real-time pcr on all samples, and found four cases of rsv, three of which were missed by viral culture and were found only by real-time pcr [ ] . serology has a reported sensitivity of up to % but is not commercially available [ ] . ribavirin, a synthetic guanosine analogue, is an antiviral agent with activity against rsv. its exact role in the treatment of hsct or sot recipients who have rsv infection remains unclear, however. dozens of studies using ribavirin in various formsintravenous, oral, or aerosolized-and at different points in the illness now have been reported. although generalization of so many studies is difficult, the preponderance of data suggests a benefit to ribavirin therapy, which may be augmented by the addition of nonspecific or rsv-specific intravenous immune globulin. this benefit has been demonstrated best in trials using aerosolized ribavirin, in contrast with decidedly mixed evidence for oral or intravenous ribavirin. this combination therapy is now endorsed in several sets of consensus guidelines [ , ] , although most of the data on which this recommendation is made are from experience in treatment of hsct recipients. the reader is referred to the excellent review by englund and colleagues [ ] for a detailed summary of years of ribavirin trials for rsv infection through . despite the recommendations in favor of ribavirin therapy for rsv, the patient population that might benefit most from the therapy has not been identified definitively. as noted previously, morbidity and mortality from rsv are linked to progression of infection to the lower respiratory tract. several investigators, therefore, have sought to use ribavirin-based therapy for the treatment of documented rsv uri. recent series by ghosh and colleagues [ ] and small and colleagues [ ] have demonstrated low rates of progression to pneumonia in hsct recipients who had rsv uri and who received early therapy with ribavirin and intravenous immune globulin. antiviral treatment of rsv uri is specifically not recommended by the american society of transplantation, however [ ] . in the ghosh [ ] and small [ ] series, those patients progressing to pneumonia while receiving therapy also had low mortality rates in comparison with historical controls. interestingly, the benefit of therapy for established pneumonia, which is recommended in the guidelines, has not been as clear in the literature [ ] . mortality rates in sot or hsct recipients who had respiratory failure caused by rsv pneumonia have been estimated at %, however [ ] , and at least one consensus group does not favor treatment at that late stage [ ] . it is important to highlight the potential difficulties of administering aerosolized ribavirin, which requires a small-particle nebulizer machine that may not be present in all hospitals. in addition, ribavirin is tera- togenic, so pregnant women may not enter the room of a patient receiving the therapy. those who do enter the room (including the patient) may develop a number of bothersome side effects from exposure to the drug, including headache, rash, and conjunctivitis [ ] . these potential barriers to drug delivery are best addressed in advance if a provider group wishes to ensure timely administration of the drug when a patient is identified. epidemiology piv and rsv are both members of the paramyxovirus family, but, unlike rsv, there are four major serotypes of piv that cause disease in humans. piv- and piv- cause annual winter outbreaks in a pattern similar to rsv or influenza, whereas piv- circulates in low levels year-round, with epidemic spread frequently seen in the spring or summer [ ] . piv- and piv- are the classic causes of childhood croup, whereas piv- is more associated with adult disease and with lrti or pneumonia. the epidemiology of piv- is not clearly defined, because it is the least commonly isolated serotype. as in the case of rsv, epidemics of piv are frequently reported both in the community and in dedicated hsct units and have been the cause of significant morbidity and mortality [ , ] . data on factors contributing to epidemics in these settings are limited; one series reported piv shedding for months in two hsct recipients [ ] . piv infections account for % to % of carv infections in recent case series of hsct and sot recipients (see table ) [ - ] . two excellent longitudinal surveys of piv infection in hsct recipients were published in [ , ] . nichols and col-leagues [ ] at the fred hutchinson cancer research center reviewed hsct recipients who received transplants between and and found ( . %) piv infections. of these, piv- accounted for %, piv- for %, and piv- for %. elizaga and colleagues [ ] in london similarly reviewed hsct recipients from to and found ( . %) with piv- infection. unlike most reports of rsv in hsct recipients, piv was found with similar frequency in recipients of allogeneic ( . %) and autologous ( . %) hsct [ ] . among sot recipients, case series of piv have been reported in renal transplant [ ] and lung transplant recipients [ - ] . between . % and . % of lung transplant recipients may develop piv infection, although some may be asymptomatic infections detected during frequent bronchoscopies [ ] . in lung transplant recipients, piv- accounted for % of piv isolates, piv- for %, and piv- for % [ ] . cough is the hallmark symptom of piv infection, but other uri symptoms (eg, rhinorrhea) may be absent. fever is uncommon: in lung transplant recipients who have piv lrti, only % to % are febrile [ , ] . the frequency of lrti in recent series of piv infection in hsct recipients is reviewed in table [ , , , - , , , ] . overall, lrti (as either the presenting syndrome or as progression from uri) is reported in one third of patients, and half of those with lrti die, but considerable variability exists among series. of patients in series with at least cases of piv infection, rates of lrti vary from % to %, and mortality among patients who have lrti varies from % to %. importantly, both large reviews of piv infection in hsct recipients demon- [ , ] . the diagnosis of piv infection usually is made with ifa or dfa testing of respiratory secretions, along with viral culture. it is notable that most widely available fluorescent antibody tests for piv do not test for piv- , which may explain in part why there has been only one reported case in an hsct recipient [ ] . although real-time pcr testing is being developed, there are not yet sufficient data to permit comment on its relative usefulness in comparison with conventional testing. ribavirin has activity against piv, and a number of smaller recent reports in which ribavirin was given to all hsct recipients who had piv have demonstrated lrti and mortality rates lower than historical controls [ , , , , ] . the two largest series, however, demonstrated no benefit of ribavirin given alone or in combination with intravenous immune globulin [ , ] . furthermore, nichols and colleagues [ ] demonstrated a failure of ribavirin therapy to shorten duration of shedding time. a recent consensus statement from the infectious disease community of practice of the american society of transplantation recommends for piv lrti that providers ''consider aerosolized ribavirin as no other options exist but experience to date provides little evidence for efficacy'' [ ] . influenza, an orthomyxovirus, is one of the most common community-acquired respiratory viruses and is a significant cause of morbidity in transplant recipients. the actual incidence of influenza in transplant recipients is unknown; many cases are likely undiagnosed, and case reports of influenza illness probably overestimate the severity of illness in this population. although most cases of influenza are acquired in community settings, nosocomial acquisition has been noted in both sot and hsct units. because nosocomial acquisition often is associated with earlier acquisition after transplantation, these cases are more likely to result in worse outcomes. influenza occurs on a seasonal basis, with the vast majority of cases occurring during winter months. both influenza a and b have been described in transplant recipients; the distribution of these infections mirrors community patterns of infection. influenza virus has been reported to be a significant pathogen in both hsct and sot recipients; among sot recipients, lung transplant recipients may be at special risk for infection [ , ] . transplant recipients have been documented to have persistent influenza viral shedding, serving as a potentially significant reservoir of virus that can spread to others in both community and institutional settings [ , ] . the timing of influenza infection with respect to transplantation significantly affects the outcome of infection, with more severe infection occurring in the earlier posttransplantation period. in most cases, upper respiratory tract symptoms predominate. lower respiratory tract involvement is uncommon. other complications of influenza include bacterial superinfection, central nervous system involvement, myocarditis, and transplant rejection [ , ] . influenza mortality remains low. influenza should be suspected in any individual presenting with fever, rhinorrhea, coryza, myalgia, and headache during the winter months. diagnosis of influenza typically relies on isolation of the virus, either by fluorescent antibody techniques (dfa or ifa) or by viral culture of nasal and oropharyngeal epithelial cell samples obtained by nasal lavage (in pediatric patients) or swab sampling [ , ] . alternatively, bal specimens can be assayed. serologic diagnoses are retrospective and may be limited by impairment of humoral responses in recent hsct recipients or sot recipients, especially those receiving mycophenolate mofetil. there are several antiviral agents with demonstrated efficacy against influenza, most of which have been used to varying degrees in transplant recipients [ , ] . amantadine and rimantadine have antiviral activity limited to influenza a; oseltamivir and zanamivir have activity against both influenza a and b. to date, no significant drug interactions have been reported with any of these medications and immunosuppressive therapies including calcineurin inhibitors. the recommended dosages and duration of treatment are the same as those for the normal host. antiviral resistance has been reported rarely in transplant recipients [ ] , but there are currently no specific recommendations for altering antiviral therapy. there are several fundamental differences between adenovirus infection and infection with the other common carvs. adenovirus may be acquired by person-to-person transmission as a primary respiratory tract infection, as is the norm for rsv or piv. most adenovirus disease in immunocompromised patients, however, is probably reactivation of latent infection. in addition, adenovirus infection can produce a wide variety of clinical syndromesgastroenteritis, hepatitis, and hemorrhagic cystitisin addition to respiratory tract illness. these patterns of illness may vary by host and by adenovirus serotypes [ ] . adenovirus infections account for % to % of carv infections in recent large case series (see table ) [ - ] . in general, adenovirus is more common in children, in whom infection with a new serotype may be primary infection and more likely to produce true clinical disease. although adenovirus may be the least commonly reported of the four main carv infections in these series, it is important to recall that most of these series test patients who had symptomatic uri; the true incidence of adenovirus infection in these patient populations probably would be higher if other clinical manifestations were included. in general, three patterns of adenovirus infection are described in hsct and sot recipients who have positive sputum testing for adenovirus infection: ( ) asymptomatic; ( ) symptomatic respiratory tract infection; and ( ) disseminated disease, with or without respiratory tract involvement. mortality from adenovirus clearly is tied to dissemination of the infection, but dissemination does not require progressive respiratory tract infection. therefore, unlike rsv or piv infections, many cases of fatal adenovirus infection are reported in patients who have adenovirus isolated from the upper respiratory tract only and without radiographic evidence of pneumonia or positive testing from lower respiratory tract samples [ , ] . outcomes from adenovirus respiratory tract infection after hsct have been poor. four recent series describe mortality rates ranging from % to %, with a cumulative mortality of % ( of cases) [ , , , ] . although mortality from adenovirus pneumonia is high, it is noteworthy that one third of these deaths occurred in patients who had adenovirus uri without evidence of pneumonia. fewer cases of adenovirus infection have been reported in sot recipients, but details presented in some reports shed important light on the nature of the disease in this population. mcgrath and colleagues [ ] performed the largest review of adenovirus infection in adult liver transplant recipients and showed an overall incidence rate of . %. four ( %) of their patients who had positive cultures were asymptomatic. of the seven who had clinical disease, three had pneumonia, but all had evidence of disseminated disease, making it unclear if the pneumonia was a primary event or the result of dissemination of uncontrolled infection. therefore, it is unclear if any patients in this series had a true, newly communityacquired respiratory tract infection with adenovirus. several papers have reviewed the potential significance of adenovirus infection in lung transplant recipients. approximately % to % of lung transplant recipients may develop adenovirus infection in longitudinal studies [ , , , ] . in many of these patients, adenovirus has been tied closely to graft failure and to acute and chronic rejection, but the numbers of patients involved prevent rigorous statistical analysis or the ability to draw a firm conclusion. in the two largest recent series examining the potential link between bos and carv infection, for example, adenovirus accounted for only of isolates [ , ] . adenovirus infection may be diagnosed using the widely available immunofluorescent antibody kits. this test is insensitive for adenovirus in sputum, however, with a reported sensitivity of perhaps % in immunocompetent hosts. given that dfa and ifa are the most commonly used assays, this lack of sensitivity may explain, in part, the relative infrequency of adenovirus in some surveys of carv infection. the virus can be cultured as well and is identified readily by characteristic smudge cells on histopathology. because adenovirus frequently may be a reactivation disease, more sensitive assays, perhaps pcr based, may detect the infection more frequently. further study is needed to determine whether patients who have more indolent adenovirus replication actually have clinical disease, or whether another pathogen is responsible for the clinical presentation. the widely reported high mortality rates from adenoviral infection, particularly in the early posttransplantation period, have prompted many clini-cians to push for early and aggressive treatment of documented adenovirus infections in hsct and sot recipients. unfortunately, there are limited data to support the efficacy of available therapeutics. recently cidofivir has been shown to improve outcomes in small studies of children after hsct and has had anecdotal reports of success in adults. because of the significant risk of nephrotoxicity associated with cidofivir, this agent should be used with caution in transplant recipients who may be at increased risk for renal impairment. although both have been tried in the past, neither ribavirin nor ganciclovir has demonstrated significant efficacy; consequently, neither agent is recommended [ ] . two recent articles have questioned the need for treatment in all patients who have documented adenovirus infections. walls and colleagues [ ] recently reported the results of retrospective testing for adenovirus in consecutive pediatric hsct recipients. in this series, children had at least one positive test for adenovirus, but of the spontaneously cleared the infection without antiviral therapy. two children died of disseminated disease, and each first tested positive in the first weeks after hsct. a recent prospective monitoring study by van kraaij and colleagues [ ] in hsct recipients also demonstrated several cases of early but asymptomatic adenovirus infection. the rhinoviruses (comprised of more than serotypes) are among the most common causes of the common cold in immunocompetent adults and children. only a few studies have addressed their potential role as respiratory pathogens in sot and hsct recipients, but the available data suggest that rhinovirus infection may be underappreciated. four studies of prospective active surveillance for carv infection-one in adult lung transplant recipients [ ] , one in pediatric hsct recipients [ ] , and two in adult hsct recipients [ , ] -noted rhinovirus infection in their cohorts; in two studies, rhinovirus was the most common isolate. in these groups, most rhinovirus infections were asymptomatic, but, taken together, three other longitudinal studies of hsct recipients who had symptomatic respiratory tract infection identified rhinovirus in a total of ( %) of isolates [ , ] . of these reported cases of symptomatic rhinovirus respiratory tract infection, ( %) progressed to lrti, and two ( % of lrti) deaths were attributed to rhinovirus pneumonia. several of these series used real-time pcr for viral surveillance; as this newer diagnostic technology becomes more widely used, the true epidemiology and impact of rhinovirus infections in hsct or sot recipients will become clearer. at present, there is no specific antiviral therapy available for the treatment of rhinovirus infections. coronaviruses, like rhinoviruses, are frequent causes of benign uri occurring in annual wintertime community outbreaks. laboratory isolation of these agents is difficult, so no systematic study of their possible role in lrti in hsct or sot recipients has been undertaken [ ] . the recent experience with the newly identified causative agent of severe acute respiratory syndrome (sars), sars coronavirus, bears mention. kumar and colleagues [ ] in toronto reported a liver transplant recipient who died from sars. a study of tissue obtained at autopsy revealed that a dramatically higher concentration of the sars coronavirus was present in this patient's tissues than in those of other case patients, suggesting both a reason for the fatal course and the possible role of immunosuppressed patients as 'super-spreaders' of the epidemic. nearly all viruses in the herpes virus family have been reported as occasional causes of pneumonia in hsct and sot recipients. herpes simplex virus type (hsv- ) will reactivate almost universally after hsct or sot in the % to % of adults with latent infection, and several reports have documented pneumonia, occasionally fatal, from this pathogen [ , - ] . hsv- is suppressed effectively by agents used for cytomegalovirus prophylaxis, however, and reports of hsv- pneumonia have greatly decreased in the era of universal prophylaxis. at most centers, prophylactic acyclovir is given to suppress reactivation of herpes simplex disease even when the recipient and the donor are cytomegalovirus negative. similarly, acyclovir and ganciclovir are active against varicella-zoster virus (vzv), which has been reported as a rare cause of pneumonia in sot or hsct recipients [ , , ] . vzv may reactivate at any point in the posttransplantation course, and shingles is a common disease in all immunosuppressed populations. because most cases of vzv pneumonia in hsct or sot recipients are preceded by the characteristic vesicular skin rash [ ] , and because vzv pneumonia is rare, it is assumed that prompt antiviral therapy with acyclovir can abort the progression of reactivation disease to pneumonia. vzv pneumonia may be more of a concern in children or adult patients who have no innate immunity to vzv from either natural exposure or previous vaccination, because primary vzv infection is more likely to cause pneumonia. finally, the roles of human herpesvirus and human herpesvirus (hhv- ) as pulmonary pathogens remain poorly understood. active replication with both viruses can be detected, probably as reactivation of latent infection, in % to % of hsct or sot recipients [ ] . several authors have reported isolation of human herpesvirus from sputum or lung tissue in patients who have otherwise idiopathic pneumonia after hsct [ - ] . the overall data, however, conflict as to whether a causative role can be established [ , ] . ross and colleagues [ ] have reported hhv- in seven ( %) of seven lung transplant recipients who had early bronchiolitis obliterans with organizing pneumonia and in three ( %) of four patients who had diffuse alveolar damage. in this series, hhv- was detected in ( %) of lung transplant recipients who had no pathology on biopsy and in ( %) of patients who had acute or chronic rejection (ie, bos). these findings are thought-provoking and confirm the need for further research into the potential role of hhv- as a single or copathogen for certain patterns of pulmonary disease. given that carv infections in hsct or sot recipients occur frequently and are associated with poor outcomes, attention must be given to the prevention of carv infection. recommendations for a multifaceted approach to the prevention of morbidity and mortality from these infections are presented in box . any approach to the prevention of carv infection must start with appropriate hand hygiene. with the possible exception of influenza [ ] , which may be transmitted in part by aerosolized droplets, carv infections are transmitted by larger droplet particles that are introduced to the host oropharynx from the hands. regular hand washing with attention to hand washing before food preparation or meals significantly reduces the incidence of carv infection. in the inpatient setting, the widespread availability of alcohol-based hand-washing products has reduced the transmission of most hospitalacquired infections. additional infection control measures are recommended to prevent the spread of carv infections on inpatient units. the centers for disease control and prevention recommend contact isolation for all patients who have carv infection [ ] . in addition, droplet precautions should be taken in the rooms of patients who have adenovirus or influenza respiratory tract infections. importantly, patients should be placed under special precautions when the infections are first suspected, not when they are first confirmed, to limit the exposure of other patients and staff to infectious droplets [ ] . these measures have been used successfully on a number of occasions to limit nosocomial spread of carv infections. one group reported an % reduction in rsv cases on a hsct unit with institution of droplet precautions and with cohorting case patients [ ] . many authorities, including the centers for disease control and prevention, advocate strongly for restricting visitors during the winter months as well [ , ] . infection with many carvs produces a measurable, type-specific antibody response. this response is neither long lasting nor protective [ ] . unfortunately, trials of active vaccination for rsv or piv have been disappointing, including trials of subunit vaccines and live attenuated viruses [ ] . cortez and colleagues [ ] recently reported on the use of passive vaccination (pavilizumab) in allogeneic bone marrow transplant recipients. although titers of rsv-specific immune globulins were increased, no difference in the rates of rsv infection were observed. prevention of influenza in transplant recipients has been focused on vaccination, and transplant recipients are among the immunosuppressed hosts targeted for influenza vaccination [ ] . the standard inactivated vaccine is composed of two influenza a and one influenza b strain. vaccine composition varies annually based on predicted antigenic drifts and shifts in circulating virus; consequently, annual reimmunization is recommended for optimal protection. live attenuated intranasal influenza vaccine is not recommended for immunosuppressed hosts, including transplant recipients, or for family or health care providers who are in close contact with the patients. numerous studies over several decades have examined vaccine responses in transplant recipients and have demonstrated conflicting results. in general, both humoral and cellular vaccine responses seem to be suboptimal when compared with healthy controls and cannot be reliably predicted based on the level of immunosuppression [ - ] . anecdotal reports have suggested a potential linkage between vaccination and organ rejection; however, this association has not been supported by the majority of studies examining the immunogenicity of vaccine in sot recipients. current recommendations support annual vaccination of all sot recipients, although it is likely that those with more recent transplants may be less likely to respond to vaccine. hsct recipients may be especially poor vaccine responders within the first years after transplantation [ ] . prophylactic administration of licensed antiviral agents (including oseltamavir and zanamavir) may serve as an alternative preventive measure for individuals who are unable to receive influenza vaccination (eg, those with egg allergy) or who are anticipated to be especially box . approaches to reducing morbidity and mortality from community-acquired respiratory virus infections in hematopoietic stem cell transplant or solid organ transplant recipients careful hand hygiene, especially during fall and winter months vaccination of patients and close contacts (particularly for influenza) avoidance of contact with patients who have symptomatic uri patient education (eg, how carv infection is spread, how to avoid sick contacts, how to perform appropriate hand hygiene) combination nasal/throat swab fluorescent antibody testing in early uri consideration of bronchoscopy for bal sample if symptoms progress or in any hsct or sot patient with an unexplained lrti or pneumonia routine testing for carv infection in all sot or hsct recipients presenting with pneumonia, including those patients who have an already identified bacterial or fungal pathogen expanded use of newer diagnostic tools (eg, real-time pcr) awareness of seasonal patterns of carv infection and of circulating viruses in the local community patient education (eg, regarding need to contact a physician when uri symptoms occur) consideration of pre-emptive antiviral therapy for rsv or piv uri in the early posttransplantation period specific antiviral therapy for influenza specific antiviral therapy for influenza aerosolized ribavirin therapy for rsv, possibly in combination with intravenous immune globulin appropriate therapy for bacterial or fungal coinfections strict adherence to infection-control guidelines in hospitals, including attention to hand hygiene and contact, droplet, or aerosol isolation as dictated by accepted guidelines consideration of cohorting patients on inpatient units active surveillance or case finding by infection control personnel careful monitoring of staff and visitors for symptoms of uri, particularly during times of heightened carv prevalence in the community separation of sick from healthy patients in outpatient waiting areas unlikely to respond to vaccine [ ] . although not specifically studied in transplant recipients, these antiviral agents have been demonstrated to be effective in preventing the acquisition of influenza when administered to immunocompetent individuals during periods of peak influenza activity [ ] . because transplant recipients may be suboptimal vaccine responders and are at increased risk for adverse outcomes from influenza, consideration should be given to immunization of household contacts before the influenza season. carvs are frequent causes of both uri and lrti in hsct or sot recipients. in most series, rsv and piv are the most common carvs. significant morbidity and mortality are associated with these infections, particularly when they progress to lrti. outcomes are also poor with adenovirus, frequently reflecting disseminated infection. efforts to prevent morbidity and mortality from carv infection should focus on prevention, because treatment options are limited, with inconclusive data to support their efficacy. respiratory virus infections in bone marrow recipients: the european perspective respiratory tract viral infections in bone marrow transplant patients respiratory viral infections after bone marrow / peripheral stemcell transplantation: the christie hospital experience low mortality rates related to respiratory virus 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prophylaxis during the influenza season in a paediatric cancer centre: prospective observational study key: cord- - m h m authors: jarmoliński, tomasz; matkowska‐kocjan, agnieszka; rosa, monika; olejnik, igor; gorczyńska, ewa; kałwak, krzysztof; ussowicz, marek title: sars‐cov‐ viral clearance during bone marrow aplasia after allogeneic hematopoietic stem cell transplantation – a case report date: - - journal: pediatr transplant doi: . /petr. sha: doc_id: cord_uid: m h m background: respiratory viral infections are known causes of mortality after allogeneic hematopoietic stem cell transplantation (hsct). here, we report a unique case of a child with viral pneumonia caused by coinfection with human metapneumovirus (mpv), respiratory syncytial virus (rsv), and sars‐cov‐ after hsct. case report: a ‐year‐old girl with acute lymphoblastic leukemia underwent allogeneic hsct from a matched, unrelated donor. during the posttransplant period, in profound leukopenia (below leukocytes/µl), she was diagnosed with sars‐cov‐ , mpv and rsv pneumonia and was treated with ribavirin and chloroquine. before leukocyte recovery, the girl became asymptomatic, and sars‐cov‐ and rsv clearance was achieved. the shedding of sars‐cov‐ stopped before immune system recovery, and one may hypothesize that the lack of an inflammatory response might have been a contributing factor to the mild clinical course. conclusions: posttransplant care in hsct recipients with covid‐ infection is feasible in regular transplant units, provided the patient does not present with respiratory failure. early and repeated testing for sars‐cov‐ in posttransplant patients with concomitant infection mitigation strategies should be considered in children after hsct who develop fever, respiratory symptoms and perhaps gastrointestinal symptoms to control the spread of covid‐ both in patients and healthcare workers in hospital environments. training of staff and the availability of personal protective equipment are crucial for containing sars‐cov‐ infection. case report a -year-old girl was admitted to the department of bone marrow transplantation in march for hsct. at the age of years, she was diagnosed with pre-b common acute lymphoblastic leukemia. due to early bone marrow relapse, the child continued chemotherapy with bortezomib and was enrolled in one course of blinatumomab followed by hsct from a / hla matched unrelated donor. on admission, the patient was in good condition with normal vital signs, mild cough, a slightly sore throat and normal auscultatory findings. this article is protected by copyright. all rights reserved. pneumoniae, and moraxella catarrhalis) revealed mpv rna. however, testing for sars-cov- was not available at the time of admission. despite mpv replication, the decision was made to proceed with transplantation due to her good condition because the timing for hsct was optimal and any delay would have increased the risk of leukemia relapse. pretransplant conditioning consisted of fractionated total-body irradiation (tbi) at a total dose of gy, etoposide (at a dose of mg/kg bw) and antithymocyte globulin (atg, grafalon, neovii, total dose of mg/kg bw). cyclosporine a (csa) from pretransplant day - and methotrexate on posttransplant days + , + and + were administered as graft-versus-host disease prophylaxis. on the first day of tbi, the patient developed a fever of . °c, and empiric antibiotic therapy with piperacillin/tazobactam and amikacin was started. on pretransplant day - , the repeated pcr multitest revealed mpv with concomitant rsv infection. fenoterol with ipratropium in nebulization and oral ribavirin (copegus, roche; mg bid) were initiated. the early post-hsct period was uneventful, with persistent respiratory symptoms and signs and blood oxygen saturation > % on room air. as of posttransplant day + , mucositis was observed, which is a standard complication after hsct. her respiratory symptoms progressed, and she exhibited diffuse wheezes and rhonchi with crepitant rales limited to basal areas of the lungs. due to posttransplant patient isolation, no imaging studies were performed. on posttransplant day + , the patient complained about weakness, chills, dysuria and mucositis-related oropharyngeal pain, with a temperature of . °c, an increase in c-reactive protein (crp) to over mg/l (normal range < mg/l) and an increase in procalcitonin to . ng/ml (normal range < . ng/ml) (suppl. figure ). because of swallowing difficulties, ribavirin was stopped after six days. on the + posttransplant day, information about covid- was revealed in the radiotherapy center where the patient underwent the tbi procedure. a pharyngeal swab was positive for sars-cov- using a vitassay qpcr test, which is based on qualitative assessment of two viral this article is protected by copyright. all rights reserved. genes: orf ab and n. the covid- diagnosis was made on the + posttransplant day, and treatment with chloroquine ( mg/kg bw/ h; bid for days) and azithromycin ( mg/kg bw as the first dose followed by two daily doses of mg/kg bw) was initiated on posttransplant day + after covid- confirmation (as recommended for polish pediatric patients in march ); ribavirin was restarted on posttransplant day + . the identification of covid- was followed by a decision to allow the patient to stay in the area of the pediatric transplantation unit. a part of the transplant unit was isolated for the patient, and she was isolated together with her mother in a single room with separate bathrooms and anteroom. the air conditioning in the patient's room was turned off to avoid spreading of infectious material due to positive pressure. procedures typical for covid- were initiated (continuous remote vital sign monitoring, call contact, dedicated nursing staff, examination without auscultation). a broad and repeated screening for sars-cov- among the health care personnel was started, and new regulations on personal protective equipment (ppe) were issued. visitations were limited to physicians and nurses trained to use class n /ffp ppe and to enter the isolation area, and no sars-cov- infections were diagnosed among the dedicated staff. of the remaining personnel, cases of covid- were diagnosed within days, which might have been transmitted by a patient. the patient continued csa, acyclovir, cefepime and micafungin, and filgrastim was started at a dose of µg/kg/day. the girl's condition gradually improved, and her cough diminished. bone marrow reconstitution was achieved on posttransplant day + . a repeated sars-cov- test on posttransplant days + , + , and + did not reveal virus replication and secretion. at the time of this report, the patient was alive and well, with full donor engraftment. we were not able to identify the source of infection, but the patient's mother and other members of the family from imminent proximity repeatedly tested negative for sars-cov- by pcr, and the mother was negative this article is protected by copyright. all rights reserved. this article is protected by copyright. all rights reserved. this article is protected by copyright. all rights reserved. fourth european conference on infections in leukaemia (ecil- ): guidelines for diagnosis and treatment of human respiratory syncytial virus, parainfluenza virus, metapneumovirus, rhinovirus, and coronavirus covid- coronavirus pandemic effectiveness and safety of antiviral or antibody treatments for coronavirus covid- virus and children: what do we know? arch pédiatrie characteristics of and important lessons from the coronavirus disease (covid- ) outbreak in china case report of covid- in a kidney transplant recipient: does immunosuppression alter the clinical presentation? first cases of covid- in heart transplantation from china. j hear lung transplant co-infection with respiratory pathogens among covid- cases precautions are needed for covid- patients with coinfection of common respiratory pathogens accepted article this article is protected by copyright. all rights reserved sars-cov- and influenza virus co-infection the first pediatric patients with coronavirus disease (covid- ) in japan; the risk of coinfection with other respiratory viruses the epidemiology and clinical characteristics of co-infection of sars-cov- and influenza viruses in patients during covid- outbreak single and multiple respiratory virus infections and severity of respiratory disease: a systematic review multiple versus single virus respiratory infections: viral load and clinical disease severity in hospitalized children etiology and impact of coinfections in children hospitalized with community-acquired pneumonia a multicenter consortium to define the epidemiology and outcomes of inpatient respiratory viral infections in pediatric hematopoietic stem cell transplant recipients significant transplantation-related accepted article this article is protected by copyright mortality from respiratory virus infections within the first one hundred days in children after hematopoietic stem cell transplantation respiratory syncytial virus and human metapneumovirus after allogeneic hematopoietic stem cell transplantation: impact of the immunodeficiency scoring index, viral load, and ribavirin treatment on the outcomes hydroxychloroquine with or without azithromycin in mild-to-moderate covid- china national health commission. chinese clinical guidance for covid- pneumonia diagnosis and treatment triple combination of interferon beta- b, lopinavir-ritonavir, and ribavirin in the treatment of patients admitted to hospital with covid- : an open-label, randomised, phase trial remdesivir and chloroquine effectively inhibit the recently emerged novel coronavirus ( -ncov) in vitro clinical features of patients infected with novel coronavirus in wuhan remdesivir for the treatment of covid- this article is protected by copyright. all rights reserved. this article is protected by copyright. all rights reserved. key: cord- -wgodcsav authors: cant, andrew; cole, theresa title: infections in the immunocompromised date: - - journal: hot topics in infection and immunity in children vi doi: . / - - - - _ sha: doc_id: cord_uid: wgodcsav infections in the immunocompromised differ significantly from those in the immunocompetent. they can be more serious, more often life threatening, more difficult to diagnose and are caused by more unusual organisms. children can be immunocompromised for a variety of reasons and the numbers, worldwide, are growing. cryptosporidium and aspergillus species. b lymphocyte defects can result in infections by bacteria such as streptococcus pneumoniae, haemophilus influenzae and staphylococcus aureus, as well as echo virus and protozoa such as giardia, whereas phagocytic defects will result in infections by staphylococcus, pseudomonas and aspergillus species. when considering infections in those undergoing hsct, it is also important to note that there is a recognized sequence of risk for different infections at different times after hsct, which equates with different aspects of the immune system compromise at these times (fig. ) . within the first days, when the patient is neutropenic, there is a significant risk of infection by both gram-negative and grampositive bacteria, along with herpes-simplex virus. between and days after transplant, when t-cell immunity is still limited, there is a rise in the numbers of fungal and cmv infections. later infections are more commonly caused by varicella zoster virus (vzv) or s. pneumoniae. whatever the cause of the immunocompromise, possible infection requires a different approach to investigation and management from those in an immunocompetent child. although an understanding of the type of immunocompromise is helpful to predict the likely organism, infection in the immunocompromised also needs to be considered by the system affected. infections can occur in any system of the body, but the respiratory system and gastrointestinal tract are especially vulnerable, as they have large surface areas and their barrier defences are, of necessity, compromised by the need to transport oxygen and nutrients, respectively. disseminated viral and fungal infections are another important risk, whilst central venous catheter (cvc) infections also constitute a frequent problem in the immunocompromised. each of these will be discussed in turn. the respiratory tract can be exposed to a wide variety of different organisms. pneumocystis jiroveci pneumonia (pcp), cmv and aspergillus are particularly important and well recognized sources of infection in the immunocompromised host; however, other significant pathogens have more recently been identified. these include respiratory syncytial virus (rsv); influenza; parainfluenza; adenovirus; picornaviruses; measles; human metapneumovirus; cocavirus; coronaviruses nl , and hku and polyomaviruses wu and k . pneumonitis and bronchiolitis are the most common presentations of respiratory infection, but lobar pneumonia may also occur. a defective immune/inflammatory response means that patients may have few respiratory symptoms, so there should be a low threshold for investigation. in one study where broncho-alveolar lavage (bal) was performed in children with immunodeficiency pre-hsct, pathogens were isolated in of these, six of whom were asymptomatic. pcp and bacteria were the most commonly identified organisms, followed by parainfluenza virus, cmv, rsv, influenza b and human herpes virus- (hhv ) (slatter et al., ) . accurate diagnosis depends on collecting the right samples and using appropriate diagnostic techniques. these include throat swabs, nasopharyngeal aspirates (npa), bal fluid and even lung biopsy, as deemed appropriate. samples must be sent to look specifically for bacteria, fungi and viruses. some respiratory pathogens will not be isolated from the upper respiratory tract; for example, pcp will not be identified on npa, whilst other organisms found on npa may not be found in the lower respiratory tract. this highlights the importance of bal as a diagnostic procedure. lung biopsy may be particularly important in the diagnosis of fungal infection, especially when there is a negative bal in patients with persistent signs, symptoms or chest x-ray changes. diagnosis may require culture of organisms (bacteria, mycobacteria or viruses), immunofluorescence (viruses), polymerase chain reaction (bacteria, viruses and fungi) or antigen testing (e.g. galactomannan for aspergillus). serological testing is often ineffective, as immunodeficient children may not mount an antibody response or may be receiving intravenous immunoglobulin (ivig), which will make results impossible to interpret. it is important to know what tests are available in your local laboratory. discussion with the local microbiologist or virologist is essential to ensure the right samples are sent for appropriate investigations, so as not to miss a serious infection. high resolution computerized tomography (hrct) of the chest is more sensitive than chest x-ray, aiming to classify a disease as interstitial, airway or involving airspace, which may aid diagnosis. pcp has historically been associated with hiv but is also a significant cause of morbidity in other groups of immunocompromised patients, particularly those with haematological malignancies, brain tumours requiring prolonged courses of steroids, prolonged neutropaenia or lymphopaenia, and those undergoing hsct. therefore pcp prophylaxis is important, as recommended by a recent cochrane review (green et al., ) . this treatment is generally in the form of cotrimoxazole given three times per week. in children that cannot tolerate cotrimoxazole, either dapsone or aerosolized pentamidine can be used. p. jiroveci infection commonly presents with tachypnoea, non-productive cough and fever, but the severity can vary. there is usually a sub-acute diffuse pneumonitis and chest x-ray changes can be subtle. these often take the form of bilateral diffuse interstitial changes, although lobar, miliary or nodular changes can be seen. hrct may show ground glass attenuation, consolidation, nodules, thickening of interlobular septa and thin walled cysts. mortality ranges between and %, if treated, but can reach nearly % if left untreated. identification of pcp can be difficult. definitive diagnosis depends on identifying the organism in respiratory tract secretions or lung tissue, usually from tracheal secretions, bronchial secretions or from lung biopsy. more recently, pcr technology has been developed for identifying pcp from secretions. in a review of children diagnosed with severe combined immune deficiency (scid) treated at a supra-regional center, out of were identified as having pcp. one was diagnosed on bal prior to transfer to the supra-regional center, one was diagnosed on nasopharyngeal secretions and bal, seven were diagnosed on bal alone, and in one diagnosis was not made until lung biopsy was performed (berrington et al., ) . recommended first line pcp treatment is high dose cotrimoxazole. this can, however cause a number of adverse effects, for example, neutropenia, anaemia, renal dysfunction, rash, vomiting and diarrhea. those that cannot tolerate cotrimoxazole or those that have not improved after - days of treatment should be changed to a different agent. choices include pentamidine, atovaquone, clindamycin/primaquine or dapsone, but experience with these agents in children is limited. corticosteroids should be given as an adjunctive therapy in moderate and severe pcp. a number of studies have shown a reduction in acute respiratory failure, decreased need for ventilation and decreased mortality (sleasman et al., ; bye et al., ; mclaughlin et al., ) . a recent cochrane review supports the use of corticosteroids in hiv-infected patients with pcp, especially in those with substantial hypoxaemia (briel et al., ) . a wide variety of respiratory viruses will also cause significant morbidity and mortality in the immunocompromised. measles is an important respiratory pathogen in the immunocompromised host and it must be remembered that the typical rash may not develop. mortality can be high, especially amongst patients with leukaemia and those undergoing hsct. a prospective multi-center review of patients undergoing hsct found direct rsv-associated mortality to be . %, and mortality directly attributable to influenza a to be . % (ljungman et al., ) . respiratory viruses often present with non-specific symptoms but progress to a significant lower respiratory tract infection. chest x-ray will often show a pneumonitis picture with diffuse interstitial changes. hrct may show peri-bronchial thickening and ground glass attenuation without consolidation in a lobular distribution. diagnosis requires identification of the organism from respiratory secretions. this may be possible on nasopharyngeal secretions or throat swab but may require more invasive testing, such as bronchoscopy and bal. laboratory techniques include immunofluorescence, pcr and viral culture. treatment is mainly supportive, but specific treatment options are evolving, making rapid and accurate diagnosis increasingly important. appropriate isolation and infection-control measures are essential to prevent transmission between immunocompromised patients, as these viruses can be easily spread. one uk study in a hsct unit identified cases of rsv over one winter season, and eight of the nine rsv strains that could be tested by molecular methods were found to be identical (taylor et al., ) . specific treatments for rsv infection include ribavirin and rsv monoclonal antibody (palivizumab). ribavirin can be given orally, intravenously or via inhalation; however, the aerosolized route has been used most frequently for rsv infection. historically, pooled hyperimmune rsv immunoglobulin has been proposed as an additional treatment, but this has been superseded by the anti-rsv monoclonal antibody, palivizumab. combinations of inhaled ribavirin and intravenous palivizumab have shown encouraging results. palivizumab has been shown to be safe and well tolerated in patients undergoing hsct, with a suggestion of better outcome (improved survival) when compared to ribavirin alone (boeckh et al., ; chavez-bueno et al., ) . there are two groups of drugs available for the treatment of influenza -namely the adamantanes (effective against influenza a, e.g. rimantadine) and the neuraminidase inhibitors (effective against both influenza a & b, e.g. oseltamivir). in recent years, there has developed increasing resistance to adamantanes. the neuraminidase inhibitors have been shown to reduce the duration of illness by one day when given to an immunocompetent host within h of onset of symptoms. although there are few data on the benefit of treating influenza in an immunocompromised patient with a neuraminidase inhibitor, their use appears sensible and safe. there is, thus far, no specific treatment available for rhinovirus, coronavirus or human metapneumovirus. ribavirin has been proposed as a treatment for parainfluenza virus infection but evidence, so far, of benefit is disappointing. although there is little clinical data on the use of ribavirin for measles pneumonitis, it does have in vitro activity and therefore, due to the high level of mortality with this condition, should be considered. a review of respiratory viral infection in children with primary immune deficiencies in a hsct unit found of patients admitted for hsct had respiratory viral infection. of these, had paramyxoviruses (rsv or parainfluenza i-iv), and were treated with aerosolized ribavirin and ivig. five of these patients also received nebulized immunoglobulin and corticosteroid. three of these five survived, compared to two out of the six who did not receive nebulized treatment. it was concluded that the nebulized treatment was well tolerated and could be a useful adjunctive therapy (crooks et al., ) . in children who have undergone hsct, infection and inflammation can become inextricably interwoven to generate pneumonitis. in this case, in addition to the need for anti-infective agents, immunomodulation will be required through agents such as steroids, ivig and anti-tumour necrosis factor monoclonal antibodies. the gastrointestinal tract is also exposed to a wide variety of organisms and viruses which are of particular concern in the immunocompromised child, notably enteroviruses, adenovirus, rotavirus, caliciviruses, but also protozoa, mainly cryptosporidium and giardia. presentation is most commonly with diarrhea and vomiting, which may protracted. cryptosporidium can also be responsible for ascending cholangitis and liver disease. in some cases, identification of the causative organism can be difficult. culture may be required to identify some viruses. pcr can be useful, for example, for adenovirus and is more sensitive than microscopy alone in detecting cryptosporidium. prevention of transmission between immunocompromised patients is essential. there must be strict adherence to infection-control policies to prevent hospital wards from becoming sources of infection. one study looking at the extent of gastroenteric virus contamination in a pediatric-primary immunodeficiency ward and a general pediatric ward found viruses on and % of environmental swabs, respectively. interestingly, these were contaminating objects used by parents rather than stafffor example the parents' room television, the parents' toilet tap and the microwave used by parents on the pediatric-primary immunodeficiency ward (gallimore et al., ) . this highlights the importance of ensuring that parents and visitors, as well as staff, comply with hand washing and infection control measures. rotavirus infection, which is usually relatively mild and self-limiting in the immunocompetent, can lead to persistent vomiting and diarrhea and, if untreated, severe malnutrition, in the immunocompromised. it can be identified in stool by using enzyme immunoassay and may also be identified on electron microscopy. there is no specific treatment. fluid and electrolyte management is important. orally administered immunoglobulin has been used in some cases. caliciviruses, namely noroviruses and sapoviruses can also cause significant problems in the immunocompromised. symptomatic infection and virus shedding can be prolonged; for example, one case report of a child undergoing hsct for cartilage hair hypoplasia demonstrated norovirus shedding for days following transplant, during the period of immune reconstitution. the child was symptomatic throughout this time (gallimore et al., ) . again, there is no specific treatment but meticulous management of fluids, electrolytes and nutritional support is essential, allowing time for immune reconstitution and consequent viral clearance. adenovirus will be discussed in more detail in the section on disseminated infection in the immunocompromised. cryptosporidium species are oocyst-forming protozoa that cause watery diarrhea which can result in severe dehydration and even death, if not treated. disease is normally confined to the gastrointestinal tract, but there is a risk of biliary tree, pulmonary or even disseminated disease in the immunocompromised. infection may be diagnosed on identification of oocysts by microscopy. enzyme immunoassays have also been used and pcr, too, can be helpful. treatment of cryptosporidium infection can be difficult and a number of agents have been proposed, including nitazoxanide, paromomycin, rifabutin and the macrolides. evidence is limited but a recent review has indicated that nitazoxanide may reduce parasite load and therefore be useful (abubakar et al., ) . in the authors' experience, azithromycin and nitazoxanide are safer options in post-hsct patients, as paromomycin has been associated with significant hearing loss, particularly when given with ciclosporin. supportive care remains essential. in those with hiv, anti-retroviral therapy, with its associated improvement in cd count, can result in improvement in the cryptosporidium infection. giardia intestinalis is a flagellate protozoan that exists in trophozoite or cyst forms. the cysts are the infective form. children with humoral immunodeficiencies are particularly at risk of chronic symptomatic infection, with foul-smelling stool, abdominal distension and anorexia. cysts may be identified on stool microscopy or by using immunofluorescent antibody testing. treatment is with metronidazole, tinidazole or nitazoxanide. it may be necessary to use combination therapy in the immunocompromised if they have failed to respond to single-agent treatment. disseminated viral infection in the immunocompromised is of particular concern. the most significant culprits are adenovirus and members of the human herpes virus: cmv, ebv, hhv , hsv and vzv. these can affect the lungs, gastrointestinal tract and brain, resulting in a variety of symptoms. reactivation of latent herpes viral infection is more common than primary infection after sot or hsct. investigation using pcr techniques allows early diagnosis and quantification of viral load, and is now possible for adenovirus, cmv, ebv and hhv . prophylaxis to prevent cmv and hsv reactivation is used for children undergoing hsct and many sots. surveillance in high-risk patients enables pre-emptive treatment to be given before damaging disease occurs. treatment will depend on the causative virus. adenovirus is usually responsible for relatively minor upper respiratory tract or gastrointestinal infection but can result in life-threatening pneumonia, meningitis, encephalitis and disseminated disease in the immunocompromised. those most at risk are patients who receive allogeneic bone marrow transplant, those with active graft versus host disease and those who receive total body irradiation. there are a number of different species of adenovirus, and these are divided into serotypes, some of which are primarily associated with the respiratory tract, while others have a predilection for the gastrointestinal tract. young children are particularly vulnerable, as they often carry adenovirus in their gastrointestinal tract, predisposing them to reactivation and dissemination when they become immunocompromised. in view of this, screening can be important in the immunocompromised and adenovirus is usually identified in urine, stool, or sometimes respiratory secretions prior to being identified in blood. a study of patients undergoing hsct were screened for adenovirus in stool, urine, on throat swab and in peripheral blood during the post transplant period. % had a positive adenoviral pcr on at least one screening test, but this was not associated with clinical signs unless it was detected in peripheral blood and, even then, there was a median delay of weeks from first detection of adenovirus until the patient demonstrated clinical signs. in one study, mortality was as high as % in those with adenovirus detected on peripheral blood. this highlights the importance of early recognition and consideration of pre-emptive use of antivirals (lion et al., ) . successful treatment of adenovirus infection has so far been limited. the most widely used agents are cidofovir or ribavirin, which may be given together with ivig. although cidofovir has potent nephrotoxic effects, these can be greatly reduced by the concurrent use of intravenous hyperhydration and probenecid. cidofovir has been shown to be more effective in adenovirus and is now considered the best first-line treatment. data on the clinical effectiveness of ribavirin in adenoviral infections are more conflicting. in vitro data suggest that ribavirin alone has activity against subgenus c serotypes. in a post-hsct patient with adenoviral infection, immune suppression should be reduced as much as possible, as t-cell immune reconstitution is very important for viral elimination. cmv infection is often asymptomatic in the immunocompetent; however, in the immunocompromised it can lead to pneumonia, colitis and retinitis. cmv persists in a latent form after primary infection and can result in reactivation in someone who later becomes immunosuppressed -for example, when undergoing hsct. cmv can be identified from respiratory secretions, urine and blood. as with adenovirus, pcr screening may be useful in identifying the virus before a child becomes symptomatic, especially in cases where reactivation is likely with immunosuppression. treatment is usually with intravenous ganciclovir, with foscarnet or cidofovir as second-line treatment. oral valganciclovir is very well absorbed and is also now an option for treatment. foscarnet has also been used in cases of children undergoing hsct to avoid the myelosuppressive effects of ganciclovir. ivig should be used alongside antiviral therapy. there has been one case report of ganciclovir-and foscarnet-resistant cmv being successfully treated with artesunate (shapira et al., ) . there is also interest in the new antiviral agent maribavir for resistant cmv. ebv is associated with lymphoproliferative disorders in the immunocompromised. replication of ebv in b cells is usually inhibited by natural killer cells, antibodydependent cell cytotoxicity and t-cell cytotoxic responses. therefore, children with cellular immune deficiencies are at risk of uncontrolled lymphoproliferation. those at particular risk are children who are transplant recipients, both sot or hsct, and those with hiv. ebv can be detected in blood by pcr and viral load can be monitored. alongside monitoring of the virus, it is important to monitor for signs of lymphoproliferation, both clinically and biochemically. biopsy of suspicious lesions is often needed to make a diagnosis. ebv infection requires treatment if it causes b lymphoproliferation or posttransplant lymphoproliferative disease (ptld). this may take the form of the anti-cd monoclonal antibody rituximab, chemotherapy or radiotherapy. decreasing immunosuppression whenever possible in a post-transplant patient is very important. more recently there have been encouraging results from work with cytotoxic t-cell therapy in ptld. this involves the infusion of ebv-specific cytotoxic t lymphocytes (ctls) generated from ebv sero-positive blood donors. in one recent multi-center study, patients who had failed conventional therapy were recruited and monitored for response: patients achieved complete remission while three showed a partial response (haque et al., ) . primary hhv infection in the immunocompetent host leads to the typical clinical picture of roseola or a non-specific febrile illness. the virus remains latent after primary infection and therefore, similar to cmv, can reactivate in immunocompromised states. the importance of hhv as a pathogen in the immunocompromised is probably underestimated, and many labs do not screen for infection; thus, many infections may not be recognized. hhv can cause fever, rash, hepatitis, pneumonia and encephalitis, as well as bone marrow suppression. hhv also appears to have synergistic effects and interactions with other infectious agents, such as cmv, adenovirus and fungi. it can be identified and quantified on blood samples by pcr. treatment, where necessary, is with intravenous ganciclovir or foscarnet. primary varicella infection results in chickenpox, a common and generally selflimiting childhood illness. in the immunocompromised, there is a significant risk of both primary or reactivated disease becoming disseminated. this is particularly associated with t lymphocyte defects. vzv is the second most common cause of viral pneumonitis in children with aids. it should be remembered that fatal vzv infection has been reported in cases where the only immunosuppressant medication has been corticosteroids at a dose of mg/kg/day of prednisolone for weeks. the virus can be identified from vesicular fluid. treatment is usually in the form of intravenous aciclovir, but, oral valaciclovir is a useful alternative in older children. an important area to consider in relation to vzv infection is that of postexposure prophylaxis. although long-term prophylaxis for vzv is not usually recommended, post-exposure prophylaxis in non-immune immunocompromised children is important. two options are available. the most widely used is varicella zoster immunoglobulin (vzig). however, due to a shortage of vzig a few years ago, oral aciclovir was reconsidered and has been shown to be effective. it must be remembered, however, that aciclovir has low bioavailability when given orally and requires multiple daily dosing. it may be more appropriate to consider the oral pro-drug valaciclovir, which has been shown to be effective and well tolerated (nadal et al., ) . further work to clarify the best prophylactic and pre-emptive treatment regimens is needed. fungal infections must be considered in specific circumstances; for example, in those who are neutropenic (where risk increases exponentially with duration of neutropenia), those on steroids and those with graft versus host disease. candida and aspergillus are of particular interest in children who have undergone hsct. symptoms that should raise the suspicion of fungal infection are persistent fevers unresponsive to antibiotics, skin nodules, chest pain and radiological evidence of infection crossing tissue planes. candida is most commonly associated with cvc infection but can also cause disseminated disease. aspergillus infection can have an insidious onset, frequently affecting the respiratory tract but then spreading to involve other areas such as the spine and intracranial cavity. investigation and diagnosis remain difficult and may require antigen testing, pcr, cross-sectional imaging and biopsy of suspicious lesions/areas. persistent mucocutaneous candidiasis is seen in patients with defects in t-cell function and may be a presenting feature for hiv infection or primary immune deficiency. disseminated infection can involve almost any organ or any anatomical site and can be rapidly fatal. it is a particular concern in patients with cvc, especially those receiving multiple infusions and/or parenteral nutrition. there are a number of different candida species that can result in disseminated infection. candia albicans is the most common but c. parapsilosis, c. glabrata, c. tropicalis and c. krusei are increasingly common (fig. ) . diagnosis may be difficult, as blood cultures are not always positive. however, identification can be made by microscopy of biopsy specimens. suspicious lesions, which are often found in organs such as the liver, kidney, spleen and brain, are best identified by cross-section imaging. pcr techniques have been developed, as well as detection of antigen from the fungal cell wall (mannan). however, these techniques are not as yet wholly reliable. there are a number of agents available for treatment, including amphotericin b, caspofugin or an azole, such as voriconazole. prolonged treatment is usually required and if there is a cvc invasive aspergillus infection in the immunocompromised usually involves lungs, sinuses, brain or skin and commonly crosses tissue planes. less commonly, it can cause endocarditis, osteomyelitis, meningitis and infection around the eye or orbit. it can cause angio-invasion, resulting in thrombosis and, occasionally, erosion of the blood vessel wall, often with catastrophic hemorrhage as a consequence. there are a number of aspergillus species that cause invasive disease. most commonly it is due to aspergillus fumigatus, but a. flavus, a. terreus, a. nidulans and a. niger are also responsible for invasive infection. diagnosis can be challenging. crosssectional imaging is very important in identifying suspicious lesions. aspergillus is infrequently identified from blood and is most commonly indicated from biopsy specimens. galactomannan, a complex sugar molecule found in the cell wall of the aspergillus species, may also be identified from blood and can be useful in aiding diagnosis. treatment is usually with amphotericin b, voriconazole or caspofungin and requires a prolonged course. surgical excision of fungal lesions may be required, especially if there are significant areas of necrotic tissue into which antifungal agents will not penetrate effectively. there is also an important association between aspergillus infection and building work on a hospital site. one study in an italian hematology unit found three cases of proven aspergillosis in patients with acute leukemia that coincided with renovation work on the hospital site and high levels of a. fumigatus in the corridors (pini et al., ) . this highlights the importance for high-risk patients (e.g. after hsct) of sterile isolation in cubicles maintained at positive pressure with highly purified air. extra attention must be paid to reducing exposure of immunocompromised patients when there is building work on any hospital site. many immunocompromised children will have indwelling cvc for treatment, be this an external broviac or hickman line, or an internal portacath. although very beneficial they, unfortunately, provide a site for infection. catheter-related blood stream infections can be serious and in some cases life-threatening. clinical features of catheter-related blood stream infection can be very non-specific. diagnosis is often made on identification of organisms from blood culture along with lack of focal infective symptoms/signs. organisms causing cvc infection are often those that would be non-virulent normal flora in an immunocompetent host; for example, coagulase negative staphylococci, enterococci and viridans streptococci. however, mycobacterial cvc infections also occur (hawkins et al., ) , as do candida cvc infections. prevention has to be the priority. lines should be inserted under strict aseptic technique and, once in place, access should be by fully trained staff using aseptic technique. local policies should be followed for accessing and flushing cvcs. historically, cvcs were often removed when infection was identified; however, many patients were left in the difficult situation of poor venous access and in need of a further general anaesthetic to replace the line. many catheter-related blood stream infections can be treated with antibiotics, without requiring cvc removal. if there is clinical suspicion of catheter-related blood stream infection, antibiotics for both coagulase negative staphylococcus and gram negative organisms should be introduced. once organisms are identified from blood culture, antibiotics can be tailored appropriately. antibiotic "locks" can be used alongside systemic antibiotics to reduce colonization within the cvc. antibiotic "locking" involves instilling - ml of concentrated antibiotic solution in to the cvc and leaving it for a pre-determined time before removal. antibiotics used in studies to treat cvc colonization have included vancomycin, amikacin and minocycline. there is also limited evidence on the use of amphotericin locks. studies have attempted to look at whether using locks alone or in combination with systemic antibiotics has benefits. the results are variable and, at this stage it must be concluded that locks are a useful adjunct to systemic treatment. there is not enough evidence to suggest they can be used alone in immunocompromised children with cvcs (berrington and gould ) . in an attempt to present cvc infection, antibiotic-impregnated cvcs have also been developed. a recent systematic review found significant reductions in catheterrelated blood stream infections in heparin-coated or antibiotic-impregnated cvcs, when compared to standard cvcs, as well as those coated with chlorhexidine, silver sulphadiazine, or silver-impregnated. there were, however, some concerns about the development of antibiotic resistance and further study is required before recommendations can be made about the most appropriate cvc to be used (gilbert and harden, ) . it must be remembered that catheter-related blood stream infection can be life threatening and there should be a low threshold for removal of the cvc if there are signs of clinical deterioration on treatment or if blood cultures drawn from cvcs are repeatedly positive, despite ongoing appropriate antibiotic treatment. there is increased mortality associated with delayed catheter removal in s. aureus and fungal infections, and so removal must be considered urgent if these organs are isolated. the benefits of removing the cvc if gram-negative organisms are identified is slightly more difficult to assess due to scarcity of data; however, it is likely that immediate removal does contribute to increased survival. in all infections the risk/benefit ratio of removing or retaining cvcs should be carefully considered. in children receiving treatment for malignancy, febrile neutropenia is a significant cause of morbidity and mortality. over time, outcome has improved dramatically but it still remains a frequent reason for hospitalization. it has been shown that (schmipff et al., ) ; hence, empiric antibiotics have become a standard part of treatment for children and adults with febrile neutropenia. fever with neutropenia in any immunocompromised child should be acted on promptly. however, exactly how this is defined and what is appropriate management varies widely. this was highlighted by a recent review of febrile neutropenia management in the united kingdom children's cancer study group centers (phillips et al., ) . there was wide variation in the definition of fever (from persistent temperature higher than . • c to a single reading of • c) and neutropenia (absolute neutrophil count < × , < . × or < . × ). empirical antibiotic regimes also varied greatly, including aminoglycosides plus a second agent (piperacillin based, cephalosporin or carbapenem), carbapenem alone or, in two cases, cefuroxime plus flucloxacillin and ciprofloxacin plus ceftazidime. timing of the anti-fungal therapy was even more variable, in terms of when to start and the duration of empirical treatment. some of this variation can be explained by variations in organisms isolated and antibiotic sensitivity from unit to unit, but this does not seem to account for all the differences in practice. therefore, although local findings should influence presenting patterns, further work is required to devise a framework within which local policies that target specific patient populations and microbiological flora are implemented. a specimen protocol is shown in fig. infections in immunocompromised children offer a variety of challenges in both diagnosis and management. organisms that result in mild, self-limiting illness in an immunocompetent host can have catastrophic effects on an immunocompromised child. signs and symptoms are often less specific and finding a causative organism can be more difficult. it is important to have a low threshold for thinking about infections and looking for them. negative tests should not be taken to be reassuring if there is clinical suspicion and it may be necessary to look further and more closely. it is important to develop a good relationship with local microbiology and virology laboratories to aid this process. once an infection is identified, it must be acted upon quickly as delay may be disastrous. treatment of any infection in an immunocompromised child is likely to be more intense and prolonged than in a child with a fully functioning immune system. it is also important to consider prophylaxis for specific patient groups in specific situations (e.g. post hsct) and each unit should have defined policies and guidelines to follow for these patients. in summary, when dealing with an immunocompromised child, for whatever reason, when there is suspicion about infection, think 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cord- -n mlxe p authors: nan title: cis annual meeting: immune deficiency & dysregulation north american conference date: - - journal: j clin immunol doi: . /s - - - sha: doc_id: cord_uid: n mlxe p nan a y.o. female was referred to our clinic with a history of multilineage cytopenias/evans syndrome, a history of idiopathic thrombocytopenic purpura, hemolytic anemia, chronic neutropenia, lymphopenia, and hypogammaglobulinemia treated with ivig. our patient was healthy until she was years old; at that time, she developed joint pain, rash, and bruising. she was found to have evans syndrome with idiopathic thrombocytopenic purpura (itp), neutropenia, and lymphopenia. she was initially diagnosed with lupus and was given steroids. her bone marrow biopsy did not conclude myelokathesis. when she was years old, she remained thrombopenic and was started on high dose of immunoglobulin replacement therapy. in ( years old), she developed polyarthritis in her upper and lower extremities. in ( years old), she had a severe nosebleed, for which she was admitted and treated with amicar twice; her platelets were found to be , k/ul. she received rituximab weekly for weeks resulting in an increase of platelet count to - k/ul. she recently (march ) had a splenectomy to remove her large spleen, and since then, her platelets have rebounded to - k/ul. in , she was placed on long-term immunoglobulin replacement therapy after being hospitalized for bilateral pneumonia for nights requiring iv antibiotics for treatment. in , she developed and was treated for another pneumonia. her family history is characterized by multiple members with autoimmune multilineage cytopenia as well as autoimmune diseases such as multiple sclerosis (mother), thyroiditis and enteropathy. on physical examination, she did not present with any warts and the remainder of her physical examination being unremarkable, except for her scar from her splenectomy and a cervical lymphadenopathy. immunologic evaluations showed igg mg/dl, iga < mg/dl, and igm mg/dl. cbc with differential and lymphocyte screen were as follows (cell/mm ): wbc . x , hemoglobin . g/dl, platelets x ; % neutrophils (anc: ), % lymphocytes, % monocytes, % eosinophils; absolute total t-cell number was ( - cells/mcl), cd + t-cells ( - cells/mcl), cd + t-cells ( - cells/mcl), natural killer cells ( - cells/ mcl), and absolute number of b cells was ( - cells/ mcl). she came to our clinic with her sister, who also had multilineage cytopenia and hypogammaglobulinemia, treated with monthly ivig; and her nephew whom had neutropenia. based on this family presentation all three underwent whole exome sequencing (wes). the patient, the patients sister and the patients nephew were all found to have a variant on cxcr (frameshift mutation on chromosome , p.val fs; refnt: tca; altnt: t). as an important note, the patient had a bone marrow biopsy, which did not conclude myelokathesis. in summary, our patient with trilineage cytopenia and hypogammaglobulinemia, without any warts or myelokathexis, had whim syndrome (warts, hypogammaglobulinemia, immunodeficiency, and myelokathexis), which was discovered by studying her wes. with the identification of her specific diagnosis, this allowed us to discuss the potential future indication of plerifaxor (antagonist of the alpha chemokine receptor cxcr ). and equally important, we discussed family planning and future pregnancies given that the mutation is autosomal-dominant. ( ) submission id# taha al-shaikhly, mbchb , kathleen mohan, arnp , matthew basiaga, do, msce introduction: complement component- (c ) is shared by the classical, lectin and alternative complement activation pathways. c , a major opsonin, facilitates phagocytosis of encapsulated microorganisms. inherited c deficiency is rare and is associated with increased risk of bacterial infections. subjects with connective tissue diseases (ctd) and c nephritic factors can have low and occasionally undetectable c levels, yet they are at an underappreciated infectious risk. we hypothesize that excessive c consumption in secondary complement deficiency disorders (scd) is associated with higher risk of bacterial infections similar to primary complement deficiency disorders (pcd). objectives: to compare the rate of bacterial infections between pcd and scd patients and evaluate the association between c level and bacterial infection risk. methods: we performed a retrospective cohort study. subjects with an undetectable complement activity (ch ) or any of the complement components measured at seattle childrens hospital from - were included in our study. we recorded the number of infections, observation periods, diagnosis (pcd, scd and its underlying etiology), lowest complement component levels, and the immunosuppressive agents used. the date of birth, and date of lowest c level were considered as start points to calculate the observation periods for pcd and scd subjects respectively. infections requiring hospitalization or parenteral antibiotics were categorized as serious bacterial infections (sbis). descriptive analyses were performed to determine medians and ranges for continuous variables. differences in rates of bacterial infection were assessed using the chi-square and kruskal-wallis tests when appropriate. among subjects with ctds, we treated every c measurement as a single observation (n= , ) and studied the association between c concentration and the -day odds of having a sbi. multivariable logistic regression was performed to determine infection risk based on c level while controlling for contributing factors. results: we identified subjects with pcd, and subjects with scd. scd consisted of three subgroups (ctd-related (n= ), nephritic factor-related (n= ), and infection-related (n= )). collectively, ctd subjects had a lower median rate of sbi compared to pcd subjects (p = . ). subjects with ctd and c level < have higher rate of bacterial infection (of any severity) (p = . ) and of sbi (p = . ) when compared to ctd subjects with c >= at the beginning of observation period ( figure ). while controlling for immunosuppression level pediatric resident, baystate medical center faculty advisor, baystate medical center introduction: zap codes for a -amino acid enzyme, zap , a member of the syk-protein tyrosine kinase family that plays an important role in t cell development and activation. zap is phosphorylated at tyrosine kinase residues upon t cell receptor (tcr) stimulation resulting in tcr-mediated signal transduction with src family kinases. zap deficiency results in a rare t+b+ nk+ severe combined immunodeficiency (scid). we report a novel compound heterozygous mutation in zap leading to presumed absent zap function in an infant with a normal trec newborn screen and scid. case description: the patient is a term, fully immunized female, born to non-consanguineous parents who was hospitalized for rsv bronchiolitis at mo. at mo she developed an erythematous, papular rash on her face and extremities, nonresponsive to topical antifungal therapy. at mo she was re-hospitalized with rsv bronchiolitis and subsequently treated with multiple courses of antibiotics for presumed bacterial pneumonia followed by albuterol and oral steroids for possible reactive airways disease. during this course of treatment, her rash resolved. at mo she presented with failure to thrive (wt < . % for age), multifocal pneumonia and respiratory failure requiring intubation. bronchial alveolar lavage confirmed pneumocystis jiroveci pneumonia prompting an immune evaluation. total immunoglobulins were normal for age, however antibody titers to tetanus, diphtheria and streptococcus pneumoniae were absent. lymphocyte enumeration revealed elevated cd t cells and markedly diminished cd t cells, normal b and nk cells. t cell proliferation to mitogens (pha, pwm) and antigens (candida, tetanus) was absent, however t cells proliferated normally to stimulation with pma and ionomycin. trec number was normal by newborn screening, but was std deviations below the mean and would have resulted in a positive screen upon repeat. invitae gene scid panel revealed two variants of unknown significance, c. c>g (p.arg gly) leading to substitution of arg with gly and c. _ dupgcat (p.ile metfs* ) resulting in a premature translational stop signal expected to disrupt the last amino acids of zap protein. parental sequencing revealed these variants to be on opposite chromosomes. the patient was successfully treated for pjp pneumonia and has since successfully engrafted a / matched unrelated donor stem cell transplant. discussion: we report a novel compound heterozygous mutation in zap which we presume led to t+ b+ nk+ scid. our patients clinical presentation of failure to thrive, recurrent lower respiratory tract infections, dermatologic findings and pjp pneumonia are consistent with previously reported cases of zap scid. her paucity of cd t cells, abundance of cd t cells and absent proliferation to mitogens are also consistent with previously described cases of zap . normal proliferation of t cells when bypassing the tcr by stimulating cells with ionomycin and pma confirms a defect in the tcr. we believe this is the second documented case of missed scid by newborn screen in ma since the implementation of trec screening in . pediatric resident (pgy iii), goryeb children's hospital attending physician, pediatric and adult asthma, allergy and immunology, llc introduction: acute otitis media (aom) is one of the most common reasons for antibiotic use in early childhood. we explored the challenges when aom fails traditional therapies and immunologic evaluation does not identify a commonly described immunodeficiency. case description: an eighteen-month-old male presented with episodes of aom and recurrent purulent otorrhea requiring intravenous antibiotics. laboratory evaluation revealed a normal cbc, normal immunoglobulins (igg , iga , igm , ige ) and igg subclasses. lymphocyte subset panel was normal. initial responses to dtap and prevnar boosters were normal, however, there was rapid decline to tetanus and pneumococcal antibody titers. a sub optimal response to haemophilus influenza type b vaccine was noted. although vaccinated twice for mmr, he never mounted mumps specific igg. mitogen response to pha was normal with decreased responses to cona and pokeweed and no detectable tetanus nor candida responses. further investigation revealed decreased non-class and class switched memory b-cells. the patient was recently vaccinated to pcv and at the present time has protective titers. discussion: it has been previously suggested that decreased memory b cells may contribute to decreased antibody responses to select vaccine antigens resulting in recurrent aom in children. our case supports the need to investigate beyond typical immunologic screening for immunodeficiencies. introduction: dna mismatch repair (mmr) system corrects replication errors in newly synthesized dna, and prevent recombination between dna sequences when they were not identical ( ) . msh is a part of mmr genes, ( ) ( ) ( ) . case: a ten-year-old girl presented with fever, brown spots on her skin, hair loss, recurrent pulmonary infections, arthritis on the left hand and right ankle. she has also been followed up with nf ( figure ). there was a first-degree cousin marriage between her parents. physical examination revealed findings of pneumonia and nf. anti-nuclear antibody, anti-ndna, anti-dsdna, anti-histone, anti ro and anti-nucleosome antibodies were positive. in her immunologic assessment showed low igg and iga levels associated with high igm level ( table ). the coexistence of nf, hyper igm syndrome, sle, were considered in the patient. intravenous ig ( mg/kg, every weeks) treatment was started due to hypogammaglobinemia. the frame shift mutation in exon of the msh gene was detected in the boztug's laboratory. in the follow up period, she admitted at years old with back pain. a mass in the left paravertebral area, related to the spinal canal and neural foramina, was detected at the l -l levels in spinal mri. the lymphadenopathy around the liver and hilum and the left parietal bone lesions were developed within two months despite surgical excision of primary mass ( figure ). as a result of pet examination; suvmax was found to be around . in the mass lesion in the paravertebral region and suvmax values did not exceed . in other lymphadenopathy and masses. atypical cellular infiltration suggesting neoplastic events, which were including small-medium size atypical pleomorphic mononuclear cells and t cells. since all these formations did not indicate definite cancer, chemotherapy was not started. interestingly, although chemotherapy was not given, progression stopped, and partial spontaneous regression was observed. discussion: the effect of msh mutations on patients may significantly vary with the inheritance pattern ( ) . leukemias or lymphomas are not common in heterozygote mmr gene defects ( , ) . however, homozygote mutations in mmr genes show a different pattern. wimmer and etzler proposed the new term constitutional mismatch repair-deficiency syndrome (cmmr-d) for patients who have a homozygous mutation in mmr ( ) . cmmr-d characterized by development of childhood cancers, mainly hematological malignancies and/or brain tumors, as well as early-onset colorectal cancers, and neurofibromatosis type ( ) . bi-allelic germline mutations in any of the mmr genes in which msh is involved increases hematological malignancies by % ( , ) . msh mutation has been associated with many cancers since its identification. leukemia, lymphoma, colorectal cancer, endometrial cancer, brain tumors are some of these cancer types ( ) ( ) ( ) ) . msh deficiency is an important disease that can affect different systems at the same time. there is a high risk of malignancy in the cases and therefore they must be closely monitored. this case has also shown that atypical lymphoproliferation may occur in msh homozygous mutant cases. (normal rage: - ) background: advances in inborn errors of human immunity have supported the discovery of new syndromes that are marked by striking features of autoimmunity and immune dysregulation often associated with cytopenias, lymphoproliferation, and a predisposition to reticuloendothelial malignancies leading to evaluation with hematologists/oncologists. moreover, hematologists/oncologists have also seen an increasing use of effector cell-based therapies, checkpoint inhibitors, immunomodulatory and targeted therapies resulting in autoimmunity and hyperinflammatory complications. a working knowledge of clinical immunology could help practicing hematologists/oncologists in the identification and management of these conditions. objectives: to support the advancement of aspho members and the field by facilitating education regarding the best practices in diagnosis and management of immunological disorders. to create a platform for the development of collaborative clinical research in patients with hematological/oncological manifestations of immunological disorders or those requiring hematopoietic stem cell transplantation for a underlying immunological disorder. design/methods the aspho clinical immunology sig was initiated based on collaboration with the clinical immunology society (cis). aspho members who are pediatric hematology/ oncology clinicians, clinical researchers, and trainees are eligible to participate. we have established a steering committee with representatives from across the united states and canada with diverse clinical and research expertise. through regular teleconferences and annual in-person meetings, we have developed a platform to provide our members with a network of immunology resources to ensure a strong foundation of knowledge and tools to conduct clinical care and research pertaining to the diagnosis, evaluation, and treatment of patients with immunological disorders. results we currently support over members within our online community. several educational initiatives have been successfully launched. we have submitted an invited review to pediatric blood and cancer which provides a case-based review of primary immune regulatory disorders. we hosted the first immunology for hematology oncology practice (i-hop) cased-based webinar series. this series features case-based discussions of patients with primary immunodeficiency disorders presented by fellow trainees and mentored by senior clinicians. we will also be hosting an aspho webinar focusing on the laboratory evaluation of primary immunodeficiencies and immune dysregulation syndromes. we have also begun the process of laying the groundwork for clinical research initiatives. conclusion: the aspho clinical immunology sig seeks to serve as a collaborative resource for pediatric hematology/oncology clinicians and researchers. through the development of educational and research initiatives, we envision improving the care of patients with immunological disorders that are often managed by pediatric hematologists/oncologists. moreover, we hope to broaden our understanding and application of clinical immunology within pediatric hematology/oncology. we hope that this successful initiative will serve as a blueprint for the development of future collaborations with other specialty societies and patient groups. autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy (apeced) is a rare autosomal recessive disease caused by aire gene mutations. clinical diagnosis is established by the presence of at least two components of the classic triad of chronic mucocutaneous candidiasis, hypoparathyroidism, and addisons disease. in europe, the classic presentation is widely recognized and nonendocrine autoimmune manifestations are rarely reported. a recent study of american apeced patients demonstrated a more heterologous presentation, with many nonendocrine manifestations including urticarial eruption, hepatitis, gastritis, intestinal dysfunction, pneumonitis and sjogrens-like syndrome, all uncommon in european reports. within the american cohort, % of patients developed a mean of three non-triad manifestations before reaching the classic triad. finding of aire mutations and high-titer antiifn-autoantibodies is seen in both european and american cohorts. we present the case of two siblings, who demonstrate an apeced-like phenotype with both classical and atypical features. they share the same heterozygous c + _ + delinsct aire mutation. the older, an eight-year-old boy, with history of prematurity, bronchopulmonary dysplasia and onychomadesis in infancy, came to medical attention at months of age due to failure to thrive (ftt), in addition to fevers and urticarial rash lasting months after his mmr vaccine. the fevers resolved with anakinra, which was discontinued two years later due to pneumonia. from age - he developed an alps negative lymphadenopathy which self-resolved. lung issues include chronic cough, initially treated as asthma but with poor bronchodilator response, and frequent lung infections, including - pneumonias per year. at age five evaluation for ftt revealed growth hormone deficiency. two years later he was diagnosed with primary addisons disease. chronic abdominal discomfort, bloating, cyclical constipation/diarrhea, recurrent rashes, dystrophic nails, and sicca symptoms are also present. his sister, age five, shows ftt, but no growth hormone deficiency. at age one, she too developed a fever and rash syndrome lasting months. severe gerd and constipation started in infancy and are ongoing. at age three she developed a transaminitis, initially diagnosed as ebv, but later thought to be autoimmune hepatitis. she has frequent viral respiratory infections, and pneumonia at age two. she has had a chronic cough, with poor bronchodilator response, for most of her life. evaluation of seizure at age three showed normal brain activity. brain mri revealed partial agenesis of the corpus callosum and microgyria. her brother has similar mri findings. both children have had developmental motor delay and poor tone. brain dysgenesis and neurodevelopmental delay has not previously been described in apeced. although there were both typical and atypical symptoms, the history in combination with genetic findings led to further investigation of an apeced-like syndrome. autoantibody testing confirmed high-titer antiifn-autoantibody typical of apeced in both children and hightiter bpifb autoantibodies found almost exclusively in apeced pneumonitis in the brother. whole exome sequencing and copy number variation analyses are underway to further evaluate the patients condition. this case demonstrates the importance of clinical presentation in the evaluation of genetic results and in the guidance of therapeutic management. ( ) submission id# rationale: infants with low t cell receptor excision circles (trec) born in queens, nassau, and suffolk counties are referred to our center for further evaluation. this study elucidates the demographic and laboratory characteristics of referred infants with transient or persistent idiopathic t cell lymphopenia (tcl) without clearly identified genetic or acquired etiology. methods: a retrospective analysis was performed from september (when trec screening started) through the end of december . descriptive statistics were calculated for demographic and laboratory characteristics. t-test or mann-whitney tests were used to compare laboratory variables. pearson or spearman tests were used to determine correlation between initial trec levels and t cell counts. by definition, the cd +, cd +, and cd + populations of transient tcl patients normalize by age year. results: eighteen infants with transient and with persistent tcl were identified. males comprised . % of the transient and . % of the persistent tcl cohorts. whites comprised . % of the transient and . % of the persistent tcl cohorts. the mean initial trec levels did not differ between the transient and persistent cohorts ( . vs. . trecs/l of blood, p = . ). mean initial absolute counts of cd + ( vs. cells/l, p < . ), cd + ( vs. cells/l, p < . ), and median initial absolute counts of cd + ( vs. cells/l, p = . ), were higher for transient vs persistent cohorts. initial trec level did not correlate with initial cd +, cd +, or cd + absolute counts. the median age of resolution for the transient cohort was . days (range - ). the absolute cd +, cd +, or cd + counts rarely exceeded the reported median values for age, and remained closer or below the th percentile for age up to days of life. the majority of both transient and persistent tcl patients demonstrated unremarkable lymphocyte proliferation to mitogens. conclusion: our centers transient tcl cohort appears to be predominantly male and non-white, whereas the persistent tcl cohort is more evenly distributed by sex but still predominantly non-white. the transient cohort had lower initial trec levels, but higher initial t cell counts. both cohorts appear to have relatively intact in vitro function. introduction: primary immune deficiency disease (pidd) is typically considered a pediatric illness, although advances in treatment and diagnosis are changing this paradigm. currently, data on pidd in older patients are very limited. objectives: to characterize the prevalence of pidd among older individuals using a patient database maintained by the consortium of independent immunology clinics (ciic), comprised of specialty immunology outpatient practices in the us. methods: patients with pidd were identified in the ciic database using icd- codes d , d. . , d . , d . , d . , d . , d . , d . , d . , and d . . a total of records from geographically-diverse clinics were identified and characterized by age, gender, and pidd diagnosis. results: of the pidd patients in the ciic registry, ( %) were between - years of age (see figure) . within this age group, most patients were female (n= , %). the most common diagnoses among patients > years of age included common variable immunodeficiency with predominant abnormalities of b-cell numbers and function (d . ; n= , %) and antibody deficiency with near normal immunoglobulins (d . ; n= , %) . in comparison, the registry included ( %) patients aged - years; this age group was predominantly male (n= ; %). the most common icd- codes within the younger cohort were relatively evenly distributed between hereditary hypogammaglobulinemia (d . ), antibody deficiency with near normal immunoglobulins (d . ) , and common variable immunodeficiency with predominant abnormalities of b-cell numbers and function (d . ). conclusions: our data suggest that pidd in patients over age may be more prevalent than previously reported. additional research is needed to corroborate these findings, further characterize the nature of pidd in this population, and determine whether there are unique diagnostic and treatment considerations within this demographic. introduction/background: increased susceptibility to invasive infections with neisseria has been well documented in patients with deficiency of terminal complement proteins. the molecular attack complex is constructed with complement components c to c . a deficiency in complement c has been described previously in both african american and south african populations. complement c deficiency is inherited in a co-dominant pattern, with multiple known mutations. we present a case of a -year-old, previously healthy male, who presented with invasive n. meningitides infection. he was found to have a novel mutation noted on genetic sequencing of the complement c gene. objective: we present the case of a -year-old, previously healthy male, who presented with invasive n. meningitides infection. on genetic sequencing, he was found to have three mutations of the complement c gene. two of which have been described previously, and a third novel mutation. methods: a -year-old male with no known history presented to us with a -hour history of emesis. he was found to be febrile, and quickly decompensated, developing septic shock. blood cultures were drawn, and within hours grew n. meningitides. he was treated with broad spectrum antibiotics upon arrival, and subsequently narrowed to ceftriaxone. his hospital course was complicated by disseminated intravascular coagulation, as well as acute tubular necrosis, leading to endstage renal disease for which he is listed for kidney transplant. results: on immunodeficiency evaluation, he was noted to have an undetectable ch (< , reference range - ). complement levels returned with c of . (reference range - ) and c r of . % (reference range - %). complement c function screen returned at % (reference range . - %). all other complement levels were within normal limits. genetic sequencing showed the patient to be compound heterozygous for two of known four variants which have been reported to recur in african patients with complement c deficiency. this included c. del and c. del, which are predicted to result in frameshift and premature protein termination. he was also found to be heterozygous for sequence c g>a, which results in amino acid substitution p.arg lys. this variant is rare, with one large database reporting it in of alleles, and not in a homozygous state. it has not been reported in a case of c complement deficiency previously. conclusions: we present the case of a previously healthy -year-old male with invasive meningococcal disease. he is compound heterozygous for two mutations that have been associated with total complement c deficiency; however, he was found to have subtotal c deficiency. furthermore, he has a third novel mutation of the complement c gene. further investigation is warranted on the significance of this finding and impact on relevance to possible kidney transplant. background: measuring the function of the classical pathway of complement activation is useful in several disease states, including complement deficiency, autoimmune conditions such as systemic lupus erythematosus and certain forms of nephritis. the original method for assessing classical pathway activity was the haemolytic ch method, but this assay can be time consuming and has reagent stability issues due to the use of sheep red blood cells. there can also be high lab-to-lab variability due to differences in the protocols used. here we report the assay characteristics of an automated, commercial, liposome-based assay to measure ch activity. we also compare the results obtained using the traditional haemolytic method with the automated, liposome-based method used on the spaplus turbidimetric analyser. methods: a linearity study was performed based on clsi guideline ep -a. the linear range of the spaplus ch liposome assay was established by analysis of a series of sample dilutions and evaluation of results against pre-defined goals for recovery and %cv. precision was assessed based on clsi guideline ep -a over days. samples with different ch activities ( . - . u/ml) were run in duplicate, with two runs per day using reagent lots and different analysers. interference analysis was performed by spiking haemoglobin, bilirubin, chyle, ascorbic acid or saline (as a control) into samples before measuring the ch activity. for the assay comparison study, sera from routine patient samples were used. samples were collected from chulalongkorn hospital, faculty of medicine, chulalongkorn university, thailand. ch classical pathway activity was assessed using a haemolytic method and also using the liposome based ch assay for use on the spaplus turbidimetric analyser (the binding site ltd., birmingham, uk). c protein concentrations were also available for of these samples. results: the liposome ch assay gives a linear response over the range . - . u/ml, covering the measuring range of the assay ( . - . u/ml) at the standard analyser dilution (neat). the within run, between run and between day %cvs were all . %. the total %cv was . % in all samples. minimal interference was observed with the four common interferents tested. a significant correlation was observed between the two ch methods (p< . , r= . , y= . x± . ), with . % agreement between the methods in determining whether patients were above or below the lower limit of the assay normal range. the individuals in disagreement had normal ch results using the haemolytic method, and low ch values in the liposome assay. of these, c values were available for / , and had c concentrations below the lower limit of the assay normal range. conclusion: the liposome ch assay for use on the spaplus analyser has passed assay development guidelines based on those set out by the clsi for linearity, precision and interference, and there is a strong correlation between this automated assay and the haemolytic ch method used here. five additional patients with low c concentrations were defined as having a low ch using the spaplus liposome method compared to the haemolytic method. ( ) submission id# background/aims: rotavirus vaccine is a live viral vaccine that is part of the routine u.s. childhood immunization schedule. live viral vaccines administered to infants of mothers who received biologic medications during pregnancy can potentially cause vaccine-associated disease. infant death from disseminated mycobacterial infection after vaccination with bacille calmette-guerin (bcg) in infants whose mothers received infliximab during pregnancy has been reported. it is currently recommended that infants born to women who received biologic therapy during pregnancy not receive live viral vaccines, however there is a paucity of information regarding adverse events from live viral vaccines. we report two infants, born to mothers receiving infliximab during pregnancy, who tolerated the complete series of rotavirus vaccine. methods: two infants who received rotavirus vaccine and whose mothers received infliximab (monoclonal antibody against tumor necrosis factor alpha which blocks the inflammatory response) during pregnancy were identified and their charts were reviewed. each mothers chart was assessed for timing of the biologic doses during pregnancy and concurrent immunosuppressant therapy. results: the mother of the first infant had crohn's disease and received infliximab every weeks throughout her pregnancy (final infusion at approximately weeks estimated gestational age [ega] ). she did not take additional immunosuppressive drugs throughout her pregnancy. the infant was born at weeks ega. the infant received rotavirus vaccine at , , and months of age. the infant did not have coexisting medical conditions or recorded hospitalizations during the first year of life. there were no side effects from rotavirus vaccine documented during well child examinations. the childs growth was normal during the first year of life. the mother of the second infant also had crohn's disease and received infliximab infusions every six weeks during pregnancy until weeks ega. additionally, she took mesalamine (anti-inflammatory) daily. the infant was born at weeks ega. the baby had a brief and uncomplicated neonatal intensive care unit stay. she did not have medical conditions diagnosed at the time of birth, or in the first year of life. the child received rotavirus vaccination at , , and months of chronological age, and the infant did not experience documented adverse reactions. the child presented to the emergency department twice in the first year of life: once for thrush at months of age and once for viral gastroenteritis at months of age. the childs growth curve was unremarkable. conclusions: we report two infants, whose mothers received infliximab during pregnancy, who safely tolerated the -dose series of rotavirus vaccination. neither infant in this case series suffered from minor or severe adverse events as a direct consequence of receiving rotavirus vaccine. this suggests that administration of rotavirus vaccine may be safe in infants whose mothers received biologic therapy. introduction: combined immunodeficiencies (cids) can arise from partial loss of function variants in recognized scid genes, which can lead to relative lymphopenia with poorly functioning and oligoclonal t cells. cids have been most commonly associated with variants of the rag genes, but other genes are also implicated. clinical symptoms may be less severe, and the onset generally is delayed, compared to typical scid presentations. case report: a -year-old female presented with a history of recurrent and progressively worsening infections involving multiple microorganisms and organs, starting in infancy and requiring frequent hospitalizations. bacterial or viral infections included rhinosinusitis, otitis media, herpetic stomatitis, dental abscesses, pneumonias, pulmonary mycobacterial abscesses, cmv hepatitis, urinary tract infections, dermal abscesses, and groin hidradenitis. fungal and yeast infections included cryptococcal meningitis, oral thrush, dermatophytosis of the face, osteomyelitis of a finger, and onychomycosis. laboratory tests in showed: mildly low t cell counts ( /ul) with a reversed ratio of cd /cd t cells ( . ); almost absent b cells ( /ul) ; and low nk cell counts ( /ul). cd + t cells were mostly of the memory phenotype ( %). t cell development showed low counts of th cells. t-cell stimulation tests demonstrated poor proliferation responses (< %) to concanavalin a, tetanus toxoid, and candida albicans, with near-normal responses to pokeweed (> %) and pha (> %). she had low ig levels (iga , igm , ige < ), except for igg ( mg /ml; due to replacement since early childhood). limited genetic evaluation at age showed a heterozygous variant in the rag gene (g. t>c, c. t>c, p.met thr; nm_ . ). discussion: loss of function variants in rag or rag genes are known to cause a t-b-nk+ type scid. more than missense variants have been reported for rag , with disease-associated variants predominantly in zinc binding regions. the rag missense variant in our patient also lies within the zinc binding region (amino acids - ). the variant is rare (mean allele frequency . in gnomad) and has been identified in at least one other individual with scid (t-, b cell-, nk+). although classified as a variant of unknown significance, occurrence in at least two individuals with deficiencies of t and b cells-within a functionally important rag domainsupports an interpretation that the variant may be pathogenic. most patients with cid with rag variants are either homozygous for a poorly functional allele or have one nonunfucitonal and a second, poorly functional allele. we detected only a single potentially pathogenic allele. our patient has decreased nk cells in addition to t and b cell defects. further genetic studies including whole exome sequencing, are planned to identify further variants in rag or other relevant genes. rationale: infants with low t cell receptor excision circles (trec) born in queens, nassau, and suffolk counties in new york were referred to northwell health for further evaluation after abnormal newborn screens. the demographic and immune parameters of infants with transient t cell lymphopenia (ttcl) without clearly identified genetic or acquired etiology are described. tcl is considered transient if the lymphopenia resolves by months of age. similar data from the following infants with low lymphocytes (fill) program of the united states immunodeficiency network (usidnet) are presented. methods: a retrospective analysis of two separate patient cohorts with ttcl are described. cohorts include patients referred to a single center, northwell health, in ny from september to december and at usidnet using data tracked by fill from june to july . results: out of , referrals at northwell, infants with ttcl were identified. infants were predominantly male ( . %) and non-caucasian ( . %). out of fill participants, infants with ttcl were identified. infants were predominantly male ( . %) and non-caucasian ( . %). initial laboratory parameters for the northwell versus fill cohorts are summarized: a) median trec levels: . vs. . trec/l of blood; b) median absolute cd + count: vs. cells/l; c) median cd + count: vs. . cells/l; d) median absolute cd + count: . vs. . cells/l. initial naïve cd + t cell information was available for northwell and fill infants (median %). mitogen proliferation studies were performed in ( . %) northwell and ( . %) fill infants with % of these northwell and % of these fill infants demonstrating normal proliferation. genetic testing, such as targeted genetic panels or chromosomal microarrays (cma), was performed in northwell and fill infants. no genetic or chromosomal aberrations were identified. whole exome sequencing (wes) was not performed in either cohort. of ( . %) northwell and of ( . %) fill infants did not receive the initial rotavirus vaccine. no fill infants were vaccinated but no adverse effects were reported in of ( . %) northwell infants who received the first rotavirus dose. of these, of ( . %) had normal mitogen proliferation while ( . %) had decreased proliferation to phytohemagglutinin. conclusions: identifying biomarkers for ttcl and developing evidencebased guidelines for the diagnosis and management of ttcl are important knowledge gaps. this descriptive study is limited by small sample size and the constraints of registry-based research. although there appear to be differences between these cohorts, our findings suggest that ttcl may disproportionately affect different segments of the population. ttcl infants with normal mitogen proliferation may be able to tolerate rotavirus vaccination. thus, routinely checking proliferation studies in all ttcl infants may help risk stratify these patients and minimize vaccinerelated adverse events. currently, there is insufficient evidence to recommend more extensive genetic testing such as genetic panels, cma, or wes. systematically collecting information about patient characteristics and outcomes, as well as encouraging increased participation in registries such as fill, may help address these shortcomings. background: systemic lupus erythematosus (sle) is a chronic, inflammatory disease that affects multiple organs. the measurement of anti-dsdna antibodies (abs) is a gold standard serological test used in the diagnosis and monitoring of sle, with higher serum levels associated with worse prognosis. however, not all anti-dsdna abs are pathogenic, and some patients have consistently high levels with low disease activity. one mechanism suggested for the pathogenicity of these antibodies is complement activation. here we describe an assay to measure the c q binding activities of anti-dsdna abs in sle patients. materials & methods: the concentration of anti-dsdna abs was determined using the quantalite dsdna elisa kit (inova) as per the manufacturers instructions. in order to determine the c q binding capacity of bound abs, samples were added to the pre-coated plate and incubated. bound anti-dsdna ab/c q complexes were then detected using a biotinylated anti-c q antibody ( ng/ml) and streptavidin peroxidase ( mg/ml). normal reference ranges were developed in serum samples from healthy controls, and upper limits of these normal ranges were used as cut-offs. the dsdna abs and c q binding capacity of bound abs was then assessed in sle patients, and compared to other markers and the sle disease activity index (sledai) score. results are displayed as absorbance at nm (au). results and conclusions: the th percentile ranges for anti-dsdna abs ( . - . au) and c q binding activities ( . - . au) were developed from the measurements generated in healthy serum samples. sle patients with an increased anti dsdna ab concentration (> . au) were then separated into those with low (< . au) and high (> . au) c q binding activities. patients whose dsdna abs had high c q binding activity were found to have significantly higher sledai scores (mean . vs . ) . serum c q concentration, serum dsdna abs (measured by another method) and serum c and c concentrations were not significantly different between the two groups. this assay suggests that dsdna abs from sle patients differ in their ability to bind complement, and that high complement binding activity of these antibodies may be linked to a more active form of disease. x-linked lymphoproliferative (xlp) is a primary immunodeficiency, caused by signaling lymphocyte activation molecule (slam)-associated protein (sap) deficiency. patients with xlp have severe immune dysregulation, usually triggered by ebv infection, leading to fulminant infectious mononucleosis, dysgammaglobulinemia and lymphoproliferation. without hematopoietic stem cell transplant (hsct) fatality is reportedly % by age . we report the natural history of xlp in a patient, and describe the lessons learned. our patient was healthy and developed normally until -years of age, when he developed progressive respiratory symptoms. lung biopsy revealed mature lymphoplasmacytic infiltrate in the alveolar septa, consistent with lymphoid interstitial pneumonia (lip). he received corticosteroids and cyclophosphamide with significant improvement. at age , he developed severe infectious mononucleosis (fever, hepatosplenomegaly, lymphadenopathy, lymphocytosis). he had a protracted clinical course, but eventually recovered and seroconverted to a typical convalescent pattern. he subsequently developed hypogammaglobulinemia, and was started on intravenous immunoglobulin (ivig). during the same year, his -year-old brother developed lip, and subsequently hemophagocytic lymphohistocytosis (hlh) and died within months from overwhelming candidiasis. unfortunately, his youngest brother (age ) then developed lip and died months later from a massive gastrointestinal bleed. both siblings were treated with corticosteroids and cyclophosphamide; they did not have detectable ebv infection. at age years, our patient experienced recurrent strokes and was found to have biopsy-proven cns vasculitis. he was treated with interferon-and recovered with residual left sided weakness, but was lost to follow-up. he continued on ivig, with no other immunomodulatory agents for several decades. he had progressive lung disease and recurrent seizures controlled with anti-epileptics. at age , he developed sudden vision change, headache and right-sided weakness, followed by a seizure. mri of the brain revealed small bilateral areas of acute infarction suggestive of a central embolic event, however, no primary thrombus was identified. he did not receive any immunosuppression but was anti-coagulated. eventually he was discharged home with resolution of weakness to his baseline. the patient was referred to our clinic after discharge and we re-evaluated him after years. immune profiles at the time showed therapeutic igg troughs, low/undetectable igm/a/e, normal t/b/nk-cell counts, normal spontaneous, but decreased antibody-dependent nk cytotoxicity, % sap protein expression (on cd +cd +, cd -cd + and cd + cd + cells), and deletion on the x chromosome encompassing the sh d a gene which encodes sap. his mother was a carrier of the same deletion. his functional status excluded the option of hsct. a year later, he had rapid deterioration with recurrent lung infections, liver failure, and thrombocytopenia. bone marrow biopsy revealed hodgkins lymphoma. he declined chemotherapy and died few days after diagnosis. our case represents a rare patient with xlp surviving to the fifth decade without hsct, particularly having experienced mononucleosis and non-ebv related cns vasculitis. our patient survived decades longer than his brothers (who most likely shared the same genetic defect) without evidence of somatic reversion ( % sap expression in cd +cd +) to explain his milder clinical phenotype. this case may help in understanding the natural history of xlp, and confirms that prognosis remains poor without hsct. haematology and oncology, chu de québec ctla- is a major negative regulator of immune responses, and ctla- haploinsufficiency has been identified as a monogenic cause of primary immunodeficiency in patients presenting with a common variable immunodeficiency (cvid) phenotype with autoimmunity. here we present the case of pb, a -year-old man who had been followed by the immunology service of our center for years. a diagnosis of cvid had first been made when the patient presented with atypical transverse myelitis, low immunoglobulin levels, and lymphopenia. over the years, his clinical picture was dominated by various forms of autoimmunity, namely inflammatory demyelinating disorder of the central nervous system, autoimmune haemolytic anemia, immune thrombocytopenia, cryptogenic organizing pneumonia, rheumatoidlike polyarthritis, chronic liver transaminitis with biopsy-proven moderate fibrosis, and lymphocytic colitis with malabsorption. immunoglobulin replacement therapy was started at diagnosis, and autoimmunity was sequentially treated with methotrexate, interferon beta -a, cyclophosphamide, mycophenolate mofetil, rituximab, and finally a combination of low-dose prednisone and sirolimus, with stabilization of his neurological condition, the most debilitating complication of his immune dysregulation syndrome. bone marrow transplant had been offered, but declined by the patient due to perceived good quality of life compared to transplant-associated risks. the patient was later referred to our hematology ward in july of for septic shock complicating febrile neutropenia, which was part of a twomonth, gradual-onset pancytopenia. the diagnosis of immune-mediated aplastic anemia soon became apparent, as demonstrated by a bone marrow biopsy performed in a peripheral center two days prior to admission. the underlying pneumonia and thereafter biopsy-induced staphylococcus aureus iliac osteomyelitis and soft-tissue abscess were treated with broad-spectrum antibiotics as well as multiple surgical interventions. the patient was started on eltrombopag, high-dose corticosteroids and cyclosporin a, the latter promptly switched to tacrolimus due to liver enzymes disturbances, all of which resulted in no significant hematologic response despite over seven weeks of treatment (with concurrent treatment of complicating infection, upper gastrointestinal bleeding, and intensive-care-unite myopathy). during that time, genetic confirmation of ctla- haploinsufficiency was received, and the patient was thereafter started on abatacept on day of current hospitalization. administration of equine anti-thymocyte was initially foregone because of perceived infectious risk in the setting of poor iliac wound healing and superimposed adenovirus viremia; however, given the lack of response, it was given on days through of hospitalization. haematologic response began on day of hospitalization with a steady rise in alllineage myelopoiesis up to a complete neutrophil response, platelet near-complete response as well as resolution of transfusion needs by day . while waiting for a well-matched bone marrow donor, isolated platelet decrease was observed and attributed to multiple factors, including low-grade thrombotic microangiopathy, inflammatory consumption and drug-related thrombocytopenia, but the patient remained well. to our knowledge, our patients presentation is one of the most severe manifestation of ctla- haploinsufficiency to have responded to targeted therapy with abatacept, as a bridge to hematopoietic stem cell transplantation, with resolution of both immune and infectious complications, showing that genetic diagnosis is helpful in optimizing the management of presumed cvid patients. hospital de octubre health research institute (i+ ), madrid, spain, dept. of immunology, university hospital octubre. madrid. spain background: xlf/cernnunos deficiency is a rare primary immunodeficiency classified within the dna repair defects. these patients present severe growth retardation, microcephaly, lymphopenia and increased cellular sensitivity to ionizing radiation. here, we describe two unrelated cases with the same nonsense mutation in the nhej gene showing significant differences in clinical presentation and immunological profile but a similar dna repair defect. methods: missense nhej mutation was identified by targeted next-generation sequencing with an in-house designed panel of genes. for foci experiments, primary skin fibroblasts were irradiated with ionizing irradiation ( cs) or treated with mm etoposide for hour. after irradiation, the cells were seeded at a density of x cells/ml in t flasks in triplicate. to evaluate cell sensitivity to gamma-ir ( and gy),adherent cells were trypsinized and counted days later. pbmcs from patient and healthy controls were irradiated with gy, fixed and stained for cd , cd and phospho-histone h ax. mean fluorescence intensities (mfi) of gamma-h ax were evaluated on gated cd + lymphocytes. results:we report two patients harboring the same homozygous mutation in cernunnos/xlf/nhej gene. strikingly, their clinical phenotype ranges from severe combined immunodeficiency to isolated thrombocytopenia followed until escolar age (table ) . they harbour the same c. c>t mutation in nhej gene but different immunologic features (table ) . p presented with mild t lymphopenia, hypersensitivity and nhej repair defect, typical for patients with xlf/nhej defects. on the other hand, p presented a more severe phenotype (t-b-) , however hypersensitivity and nhej repair defect was similar to p .of note, p has survived into the first decade of live. both patients are alive and well after hsct. discussion: usually the repair defect in these disorders is assessed by immunofluorescence assays of irradiation-induced gamma-h ax foci using skin fibroblasts. a high throughput, sensitive and reliable assay to quantify gamma-h ax foci in pbmcs isolated from blood samples would be a valuable tool to diagnose these patients and perform hsct early. flow cytometry (fc) can be applied as a rapid diagnostic tool for dna repair disorders. patients with the same homozygous mutation (p.r x) in nhej gene have been previously reported. two patients died at . and years while another of the patients is already years old and is alive (without hsct). however,none of these patients presented severe t lymphopenia as it has been observed in our first patient. conclusions: the assignment of a timely and accurate diagnosis is of paramount importance in the management of patients with defects in dna repair. in the era of nbs an abnormal trec assay should be followed by ngs approach as cernunnos deficiency may present early in life as scid,as other rs-scid defects. since genetic diagnosis takes time,functional radiosensitivity assays in peripheral blood may lead to the correct diagnosis and avoid exposure to alkylating agents during the conditioning regimen prior to genetic diagnosis. it would also be helpful in cancer patients to individualize and to guide the dosing of ionizing radiation (ir) and/or genotoxic agents to avoid accumulation of cells with genomic instability that could accelerate cancer development. figure ). her skin lesions also significantly improved after starting the medication ( figure ). her hospitalizations were complicated by fluid overload and hypertension. both fluid overload and hypertension resolved prior to discharge. she remains on mg prednisone daily, cetirizine, ranitidine, cromolyn and benadryl and hydroxyzine prn. to our knowledge, this is the youngest patient successfully treated with midostaurin and she is doing very well on therapy with no apparent side effects. she has had resolution of many of her systemic mastocytosis symptoms including skin lesions, axillary mass and improvement in her diarrhea and growth as well as objective improvements in her tryptase levels. case report: a two-year-old male presented to the hospital with a painful, non-pruritic facial and groin rash. the rash started one week prior to presentation. he had no associated fevers. his history was remarkable for failure to thrive (ftt) and chronic bilateral leg pain with antalgic gait. over the preceding months, he had been diagnosed with hand-foot-mouth disease and varicella. he had also had recurrent cervical lymphadenopathy (lad) for greater than one year requiring incision and drainage. gram stain and gomori methenamine-silver nitrate stain (gms) were negative and pathology showed only acute and chronic inflammation with areas of necrosis. his family history was negative for autoimmune disease or immunodeficiency. infectious exposure history was significant for an incarcerated father with unknown tuberculosis status and history of living in a shelter. on physical examination, the patient was well appearing with multiple erythematous papules, with superficial erosions and scabbing on the face (figure ), lower abdomen, genital area, buttocks and proximal lower extremities. he had large, firm, non-tender submandibular lymph nodes. he also had small palpable axillary and inguinal lymph nodes bilaterally. his laboratory workup revealed normal white blood cell and platelet counts, but microcytic anemia, an erythrocyte sedimentation rate of mm/hr, and c-reactive protein of . mg/dl. full body magnetic resonance imaging (mri) revealed bilateral cervical, supraclavicular, right hilar and inguinal lymphadenopathy and a patchy right upper lobe consolidation with at least one small area of cavitation ( figure ) and an adjacent smaller area of ring enhancement. it also revealed three small nonspecific hypodense foci within the right lobe of the liver and borderline splenomegaly. given these findings, there was concern for granulomatous diseases. the patient underwent a liver biopsy ( figure ) which showed non-specific evidence of necrotizing granulomatous disease. microbiological cultures and stains for bacteria, acid-fast bacilli and fungi were negative. his infectious work-up was negative for hsv, tuberculosis, hiv, syphilis, histoplasmosis, and toxoplasmosis. superficial bacterial cultures from the face and groin grew mixed gram positive and negative organisms, including methicillin-susceptible staphylococcus aureus (mssa). his immunologic workup revealed borderline elevated iga and igg with normal igm, normal t,b, nk-cell counts and pneumococcal and tetanus titers. a dihydrorhodamine (dhr) flow cytometric test was positive, consistent with a diagnosis of chronic granulomatous disease (cgd). genetic testing confirmed x-linked disease. he was treated with acyclovir and ceftriaxone with resolution of his rash. conclusion: we present a case of a two-year-old male with newly diagnosed x-linked cgd. though he had been seen by multiple healthcare providers for recurrent lymphadenopathy over the preceding year, he had no other history of recurrent viral or bacterial infections or significant family history that might implicate a primary immunodeficiency. at time of presentation, he had diffuse rash which could have caused his palpable lymphadenopathy on exam. a high index of suspicion for cgd in the setting of recurrent lad and ftt prompted sending the dhr, which led to the diagnosis. chronic granulomatous disease (cgd) is an inherited primary immunodeficiency (pid) which results in both inflammatory response dysregulation and an increase in susceptibility to certain bacterial and fungal infections. without curative treatment such as a bone marrow transplant, it remains a chronic disease with daily medication management, intermittent treatment and life-long surveillance. in general, chronic disease involves physical, psychological and social effects which can affect the patients quality of life. although some research has been done on how pid affects quality of life, there is little research in the united states about how cgd affects patients quality of life. to examine the effect of cgd on patients quality of life, as a part of a voluntary research protocol examining the natural history of immune deficiencies, we administered the who qol-bref instrument to adult cgd patients enrolled on a nih irb approved protocol and seen in the infectious disease clinic at the national institutes of health (nih) over a five-month period. the who qol-bref is comprised of items, which measure the following broad domains: physical health, psychological health, social relationships and environment. each item is rated on point likert scale. it has been validated cross culturally and has been widely field tested. the survey was interview administered to patients ( males, females) with genetically confirmed cgd. the age range was - years old (mean age . years) with a distribution of % x-linked cgd and % autosomal recessive cgd. results have been obtained and will be presented. rationale: common variable immunodeficiency (cvid) is the most common primary immunodeficiency with an estimated prevalence of : , . we aimed to analyze the clinical presentations and their associated comorbidities amongst cvid patients in usa. methods: data on , cvid patients reported in the united states immunodeficiency network (usidnet) from to were analyzed based on clinical, immunological and genetic factors. univariate analysis with spearman rank coefficients was done to analyze correlations between disease outcomes. observed survival was estimated using the kaplan-meier method. results: among the patients, ( . %) were female and ( . %) were male. median age at diagnosis was years [mean (sd), . ( . ); range, - ; iqr, - ] with median age of onset of years (mean (sd), . ( . ) ; range, - ; iqr, . females showed a longer delay in diagnosis ( . vs. . years, p= . ). higher body mass index (bmi) linearly correlated with the age of diagnosis (r= . ). in survival analysis, a -year delay in age at diagnosis increased the risk of death by . % (hr: . , % ci: . - . , p= . ). conclusions: our study suggests a longer delay in diagnosis in female subjects and a strong association with diagnosis of cvid in patients with higher bmi. females may have a longer period without symptoms leading to a diagnostic delay. gender-based and disparities-based inquiry into these trends may need additional study. the physical well-being of those with primary immunodeficiency (pi) and the physical maladies of those with pi are well-documented. since the s, advances in identification and treatment of pi has for many led to lives where the physical infections of these groups of diseases are manageable. however, not as well understood are the emotional and mental health aspects of living with pi. as part of a larger survey project the idf national patient survey, this study aims to quantify any potential mental health issues or challenges faced by adults with pi. our hypothesis-those with pi, suffer from statistically higher rates of depression when compared to the u.s. general population. the idf national patient survey was a nationally distributed, unincentivized, mail-based survey of , persons in the idf patient database identified as being either adults with pi or the parent/caretaker of a child with pi. the questionnaire comprised approximately main questions about pi as well as the validated sf- v , brief fatigue inventory and the patient health questionnaire- (phq- ) instruments. additional questions asked about current use of prescription medications for anxiety, depression, stress and pain. for the purpose of this study, only adult respondents with pi are included as the basis for analysis. the two-item patient health questionnaire (phq- ) meets the criteria for general screening of depression suggested by the u.s. preventive services task force. scored on a scale of - , a score of three or higher is suggested as the cut-point for depressive screening. according to a ahrq study that utilized meps data, , of the , ( %) respondents scored three or greater. in our survey of the ( %) adults scored three or greater ( <. .) overall, those in our survey scored lower on the sf- v mcs scale when compared to the u.s. population ( . v. . , p<. ) . further, adults with pi who scored three or higher on the phq- had an average mcs of . . those who met the phq threshold in our survey were also more likely to report moderate to severe limitations in normal activities as a result of emotional problems than those that fell below the threshold ( % versus %, p <. ). not surprisingly, those that met the phq threshold reported much higher use of prescription medications for anxiety, depression, stress ( % versus % below threshold, p <. ) as well as a higher reported use of prescription pain medications ( % versus % below threshold, p <. ). though moderate to severe fatigue was reported by % of those below threshold, % of those with phq scores at threshold reported experiencing moderate to severe fatigue (p <. ). health care providers should consider including the phq- in the overall health assessments of their patients with pi. those scoring three or higher should be referred to the appropriate professional for further evaluation. (lek et al., ) . the w l is a semi-conservative amino acid substitution, which may impact secondary protein structure. in-silico analyses supported a deleterious effect, located within the sh domain, which is a critical functional domain (chandesris et al., ; koskela et al., ) . it was thus determined that this variant is likely pathogenic. the patients prophylactic treatment was optimized with tmp-smx ( mg- mg) twice daily for prevention of infections. she was also started on hibiclens (chlorhexidine) baths once per week. she was referred to pulmonology for optimization of pulmonary health in the setting of bronchiectasis and mild decline in dlco. she was advised to followup on a yearly basis to the primary immunodeficiency clinic to assess for recurrent infections and for changes in pulmonary health. finally, targeted testing and clinical evaluation of both of the patients parents was recommended to determine if w l was inherited or arose de novo. the pathogenic role of the w l missense change would be further supported if it had occurred de novo or if it segregates with the disease in the family. uploaded file(s) uploads pulmonary function testing results.pdf j clin immunol ( ) (suppl ):s -s s introduction: lipopolysaccharide-responsive and beige-like anchor protein (lrba) deficiency is a rare autosomal recessive disease of the immune systems characterized by hypogammaglobulinemia and decreased ctla expression on t regulatory cell (t regs) due to defective intracellular trafficking of ctla . previous in vitro study has shown a significant increase of ctla expression on lrba deficient t cells after overnight culture with chloroquine, an older anti-malarial agent. this effect is likely due to increasing lysosomal ph. however, there is no evidence of such effect in human subjects after administration of weight appropriate doses anti-malarial agents. we are presenting a set of siblings with lrba deficiency who had ctla expression measured before and four weeks after starting hydroxychloroquine. case reports: case is a -year-old east-indian boy with autoimmune thyroiditis, type diabetes mellitus (dm), short stature, autoimmune cytopenias, and lymphadenopathy. he was referred to immunology clinic at years of age for suspicion of autoimmune lymphoproliferative disorder. primary immunodeficiency genetic panel was sent which revealed a homozygous mutation in lrba gene (c. _ del). this novel variant resulted in a frameshift and created a premature stop codon amino acids downstream from this location which may lead to absent or abnormal protein. lung ct scan showed interstitial lung disease. lung biopsy showed interstitial nodular and diffuse lymphoid proliferation. this diagnosis led to the testing of his sister (case ) given her history of autoimmune illnesses and the family history of consanguinity. case is a now -year-old girl with type dm, autoimmune thyroiditis, lymphadenopathy, psoriatic arthritis, and seizures. her lung imaging showed pulmonary nodules without interstitial lung disease. both cases received hydroxychloroquine while waiting for insurance approval of abatacept. ctla expression on tregs was measured prior to and four weeks after starting hydroxychloroquine treatment. at baseline, . % of case s cd cells were treg (foxp +ve, cd hi) and . % of them expressed ctla- (in contrast to . % tregs in the healthy control) with mean fluorescence intensity (mfi) of . this ratio and mfi did not change after weeks of hydroxychloroquine treatment ( mg/kg/day). soluble interleukin- receptor levels were measured: case had a baseline level of pg/ml, which decreased to pg/ml after weeks of hydroxychloroquine treatment. for case : . % of her cd + t cells were found to be foxp +cd hi and . % of these tregs expressed ctla- . this ratio increased by % after one month of hydroxychloroquine. increase in mfi was also noted from to . case had a drop in soluble interleukin- receptor level from pg/ml to pg/ml after treatment. conclusion: in contrast to the previous in vitro assays, we did not find a significant increase in ctla expression on t regulatory cells in vivo after weeks of mg/kg/day hydroxychloroquine. interestingly, soluble il- receptor levels improved dramatically with hydroxychloroquine. ( ) submission id# human nf-kappab defect results in defective intrinsic b-cell differentiation, function and class switching introduction/background: autosomal dominant heterozygous mutations in nfkb (encoding for the protein nf-kb ) have been identified in the etiology of a form of primary immunodeficiency disorder that presents with hypogammaglobulinemia, defects in b-cell maturation, endocrinopathy, and autoimmune manifestations. in humans, the effects of altered nf-kb and mechanisms of immune system impairment have not been fully delineated. objectives: to understand the mechanism of the antibody deficiency in patients with hypomorphic mutations in nfkb (c. dela; p.lys serfs* ) by evaluating b-lymphocyte proliferation, differentiation, function, and gene expression. methods: immunophenotyping of primary b-cells from subjects with mutant nfkb was completed by flow cytometry. proliferation of b-cells was assessed by cfse stimulation of primary cd + b-cells from healthy and nfkb mutant subjects. differentiation of healthy and affected naïve b-cells (cd -cd -) into plasmablasts (cd +cd +) following stimulation was assessed by flow cytometry. the supernatant from these cells were assayed for iga, igg and igm production by elisa. to study the defect in class-switch recombination, naïve b-cells and ebvtransformed b-cells from affected and healthy individuals were stimulated and expression of the aicda gene was quantified by qpcr. in parallel experiments, ebv b-cells from wildtype and nfnb mutant individuals were stimulated and aid (activationinduced cytidine deaminase) protein levels were determined by western blot. results: patients with hypomorphic mutations in nfkb (c. dela) had low memory b-cell (cd + cd + igd-igm+) and class-switched memory b-cell (cd + cd + igd-igm-) numbers. in vitro, primary bcells from these patients demonstrated a % reduction in proliferation and cell division in response to cd l and il- (p = . ). compared to healthy naïve b-cells, mutant naïve b-cells had a significant reduction in plasmablast differentiation (p = . ) and secreted significantly lower levels of immunoglobulins in response to cd l and il- stimulation. mutant naïve b-cells and mutant ebv b-cells failed to increase aicda expression and aid protein levels in response to cd l and il- stimulation. conclusions: our studies demonstrate that a hypomorphic nfkb mutation in humans affects intrinsic b-cell proliferation and differentiation. the mutation impairs transcription of the aicda gene that encodes aid, a key protein involved in b-cell class-switch recombination. the nfkb gene defect also impairs immunoglobulin production, as seen in common variable immunodeficiency-like cases. these studies provide unique translational insights into physiological activities of nf-kb in downstream immunologic outputs in humans, expanding those suggested by experimental observations in mice. background: few studies have evaluated the quality of life (qol) and patient reported outcomes of primary immunodeficiency disease (pidd) patients, and no studies have assessed medical provider perceptions of their pidd patients qol, neurocognition, physical well-being and psychosocial health. understanding provider beliefs regarding patient reported outcomes is essential to improving clinical management of pidds. here we report our pidd medical provider survey results. methods: providers were contacted via email with the assistance of the clinical immunology society. participants completed adult and/or pediatric-based likert scale survey questions via a secure online survey service. in addition to demographic information, survey questions assessed provider perceptions of patients overall qol and their impression of the impact of disease or its associated treatment on mental health, physical well-being, neurocognition, social relationships and school/work performance. clinicians were expected to make their assessments based on their pidd patient cohort as a whole rather than on specific diagnoses or patients. given the small sample size, a p-value < . was considered statistically significant; repeated measures anova and paired t-test analyses were used. results: study participants (n= ) were primarily from the united states ( %), born between - ( %) , and trained in allergy/ immunology ( %). % of survey takers practiced within an academic center, % were female and % cared for children with % of providers concurrently caring for adults. there was a statistically significant difference (p= . ) in the perceived overall qol of pediatric versus adult pidd patients with % of providers feeling as though their pediatric patients had a good qol while only % believed their adult patients had a good qol. clinicians believed adult pidd individuals had more difficulties related to associated co-morbidities rather than their actual pidd compared to pediatric pidd patients (p= . ). providers felt that the neurocognition and school performance of children were more often negatively affected by a pidd than the neurocognition and work performance of immunodeficient adults (p= . ). clinicians believe children with pidd more frequently had difficulties related to their concentration than memory (p< . ). % of those who care for pidd adults believe their patients work performance or daily mental functioning is at times negatively impacted. anxiety symptoms and social relationships were viewed as being more negatively impacted by a pidd diagnosis or treatment than anger or depressive symptoms in both children and adults (p< . ). % of pediatric clinicians feel their pidd patients experience anxiety symptoms often or almost always. of physical health parameters, energy, rather than mobility or pain, was deemed to be more deleteriously influenced by an immunodeficiency in adult and pediatric patients (p< . ). conclusions: our results show that medical providers perceive the overall qol of pediatric pidd patients to be superior to that of adults with pidd, but most clinicians feel a diagnosis or associated treatment regimen for pidd can negatively impact the physical well-being, psychosocial health, school/work performance and neurocognition of both children and adults. [cbm] complex is a critical signalling adaptor that regulates lymphocyte activation, proliferation, survival, and metabolism. primary immunodeficiencies affecting each component (termed cbmopathies) result in broad clinical manifestations ranging from severe combined immunodeficiency (scid) to lymphoproliferation. we present the laboratory and clinical findings of two canadian first nations patients found to be homozygous for the same novel card mutation (c. c>t; p.r *). results: we have identified an -month-old boy who presented with a severe case of entero/rhinovirus bronchiolitis with interstitial lung disease and a -year-old boy with a history of severe pulmonary infections (including pjp), chronic sinusitis, candidiasis, invasive bacteremia, and severe ileo-colitis and oral ulceration requiring total colectomy. both patients possessed absent tregs, absent memory b cells, and hypogammaglobulinemia. however, only the -month-old had poor t cell proliferation to pha, cona, and cd . both patients were found to be homozygous for the same novel variant of card (c. c>t; p.r *). the mutation rendered card protein expression unstable and it was undetectable by immunoblot. to confirm card deficiency, we stimulated patient b cells with phorbol -myristate acetate (pma) and ionomycin across a time-course and immunoblotted for various signalling proteins in both the nf-b (ikk/, ib, p ) and mapk (mek / , mkk , jnk / , erk / ) pathways as well as various cleavage substrates of the malt paracaspase (relb, cyld, bcl , hoil ). nf-b and jnk activation were completely absent and malt paracapase activity was lost, but surprisingly, mkk (which acts upstream of jnk) was intact. furthermore, co-immunoprecipitation experiments revealed that card was required for optimal malt association with bcl in response to stimulation. conclusions: these two cases highlight the crucial role of card in regulating lymphocyte development, function, and humoral responses. in addition, we have identified the oldest known living individual with card deficiency and he presented uniquely with inflammatory gastrointestinal disease in addition to scid, further adding to the spectrum of phenotypes associated with card -related primary immunodeficiencies. abstract: the usidnet registry began in with an niaid contract with the immune deficiency foundation, which continues today. it aims to provide a resource for clinical and lab research through enrollment of known immunodeficiency patients into a national registry, the usidnet. nih is a major national and international referral center for clinical trials on inborn errors of immunity, or primary immunodeficiency diseases. it is a mechanism for depositing nih data into usidnet. a registry of patient information may help us understand how many people have each disease. the information may improve how we diagnose and treat these conditions. the patient registry is designed to obtain longitudinal data on a large number of patients with primary immunodeficiency diseases who come to nih to participate in research. the data is collected from the nih electronic medical record system, cris and is deposited into a secure registry with restricted and monitored access. all medical information is anonymized for patient privacy. department of biochemistry, emory university, atlanta, ga oas is an intracellular sensor for dsrna that generates the second messenger '- '-oligoadenylate to activate rnase-l as a means of antiviral defense. we describe four patients with a complex early-onset autoinflammatory and immunodeficiency disease caused by heterozygous de novo oas mutations. patients presented early in life with lung inflammation including pulmonary alveolar proteinosis and interstitial lung disease. they had febrile flares with dermatitis specifically with macular, pustule and bullous features often progressing to ulceration. infants had episodes of bloody diarrhea in patients (assoc. with villous blunting and cryptitis in two patients and oesophagitis in one patient). immunoglobulin igm, igg, and iga levels were low while t cell, b cell, and nk cell numbers were generally in the normal range. exome sequencing identified de novo heterozygous oas missense mutations in all patients. one patient had a heterozygous de novo oas mutation p.ala val, with mutant oas protein being expressed in ex vivo generated t cell blasts. in sorted primary patient monocytes and b cells, oas p.ala val was associated with spontaneous rna degradation and apoptosis as determined by rna chip technology and flow cytometry, respectively, while t cells were not affected. monocytes displayed disturbed terminal differentiation and functioning as indicated by reduced gm-csf-r expression and signaling. b-cells display reduced class-switch-recombination. proliferation of allogeneic t-cells was reduced in response to sorted oas mutated monocytes and b-cells. activation of interferon response genes in pbmcs was detected. two further unrelated patients had a heterozygous de novo oas mutation p.cys tyr, which appeared to compromise protein stability in transformed patient fibroblasts and when transfected. cells transfected with this mutant protein had reduced - oligoadenylate synthesis compared to wild type transfected cells. immortalized fibroblast lines demonstrated higher levels of inflammatory cytokines and spontaneous cleavage of rnas. a th patient with the clinical phenotype had a heterozygous de novo oas variant p.val gly, but has yet to have formal validation of the variant. three patients underwent hematopoietic stem cell transplants in an effort to control their diarrhea and skin inflammation. one patient died with ongoing chronic graft versus host disease, while the two others (p.ala val, cys tyr) are alive and reasonably well with a followup of . - years. the untransplanted patient died as a result of respiratory failure. in summary, patients with de novo heterozygous oas mutations have chronic ongoing inflammation of multiple organs. this is at least in part due to spontaneous rna cleavage, apoptosis and production of inflammatory cytokines and type i interferons. this defines a new category of autoinflammatory disorder. introduction: increased susceptibility to infections is the most common complication of chronic granulomatous disease (cgd). hemophagocytic lymphohistiocytosis (hlh) is a severe disorder resulting from hyperinflammation and hypercytokinemia that can lead to multi-organ system dysfunction ( ) characterized by certain criteria: fever, splenomegaly, cytopenias, hypofibrinogenemia or hypertriglyceridemia, hyperferritinemia, increased soluble cd /il- ra, evidence of hemophagocytosis, or decreased/absent nk cell cytotoxicity ( ) . secondary hlh occurs infrequently but often is preceded by smoldering infection in cgd ( , , ) . we present a case of hlh in a -day old male, the youngest reported case with cgd. case: a -day old male with previously diagnosed x-linked cgd, due to known family history, presented with fevers. initial evaluation was unrevealing including chest x-ray, urinalysis, and blood and csf cultures. he was admitted and treated empirically with cefepime. ct demonstrated multiple multifocal nodules of the lungs and spleen. after lung nodule biopsy was performed, antimicrobial therapy was broadened to iv meropenem, voriconazole, and micafungin. despite this, he continued to have fever and developed new onset tachycardia, respiratory distress, and lactic acidosis. further decompensation with vasoactive refractory shock was treated with vasopressors and stress dose hydrocortisone. additional laboratory evaluation revealed rising liver enzymes (ast u/l, alt u/l), cytopenias (hemoglobin g/dl, anc /ul, platelets , /ul), and coagulopathy (fibrinogen - mg/dl). splenomegaly was present on abdominal ultrasound. a diagnosis of evolving hlh was considered and dexamethasone was administered. within hours of clinical decompensation, the patient died of multiorgan failure. subsequent blood cultures returned with gram-negative rods (and ultimately burkholderia cepacia). autopsy confirmed hemophagocytosis within the bone marrow. no mutations were found in genes associated with primary hlh. discussion: patients with cgd are susceptible to infectious complications and auto-inflammation most commonly involving the lungs, gi, and gu systems ( , ) . patients with cgd can be at increased risk of hyperinflammatory syndromes secondary to infections and chronic inflammation. as shown in the included case, hlh can present in infancy and can be deadly. early consideration and directed treatment of hlh is imperative, even in the setting of sepsis malignant proliferation of gamma-delta t cells include hepatosplenic t-cell lymphoma (hstl), primary cutaneous t-cell lymphoma and t-cell large granular lymphocytic leukemia (t-lgl). the former two have often been associated with splenomegaly and cytopenias. however, reactive proliferation of gamma-delta t cells in spleen mimicking malignancy has only been reported once and has a significant risk of misdiagnosis. a -year-old female presented with two years of unintentional weight loss, persistent leukopenia and thrombocytopenia, with leucocytes around - x ^ /l and platelets around x ^ / l. she also had associated macrocytic anemia (hemoglobin= - g/dl) with laboratory evidence of dat (direct anti-globin test) negative hemolysis. physical examination and computed tomography (ct) imaging showed splenomegaly. there was no hepatomegaly or lymphadenopathy. serum liver function test, auto-immune studies, hemolysis and hereditary diseases workup, viral and bacterial serologies were all normal or negative, except for mild hyperbilirubinemia and ldh elevation. bone marrow examination performed four months prior to the splenectomy revealed mildly hypocellular marrow ( %) with trilineage hematopoiesis. flow cytometric analysis and cytogenetics of the bone marrow aspirate and peripheral blood were normal except for small population of large granular lymphocyte and mild low absolute b cell counts in peripheral blood. a laparoscopic splenectomy was performed for diagnostic and therapeutic purposes due to patients worsening luq pain. there was no other treatment given prior to surgery. hours postsplenectomy her leucocytes increased to . and platelets to . her three-month post-splenectomy wbc count and platelet count was . and , respectively. hemoglobin also improved to . . pathology showed red pulp expansion by small lymphocytes (fig. ) and subsequent ihc (immunohistochemistry) was positive for cd ( fig. ) , cd , cd , tia- and negative for cd , cd and cd . cd was difficult to interpret. eber was negative. flow cytometry ( fig. ) showed increased gamma-delta t-cell population ( %) with positive cd , cd and cd and negative cd , cd and cd . molecular studies by pcr didnt reveal any t-cell receptor gamma or beta gene rearrangement. cytogenetics was negative for isochromosome q or any other abnormalities. she was symptom free at months from her splenectomy. the morphology and immuno-phenotype of these gamma-delta t cells show significant overlap with the malignant cells seen in hstl and t-lgl, such as loss or downregulation of cd , cd and cd . awareness of this reactive condition is necessary to prevent making a wrong diagnosis of a malignant disease with a potentially benign, spontaneously resolving disease. additional studies of similar cases is needed in order to establish more definitive criterion to separate benign from malignant processes and delineate the role of gamma-delta t cells. uploaded file(s) uploads fig . flow cytomtery.pptx background: sex steroids in the human thymic environment influence aire expression as well as interactions with its partners, i.e. genes coding for aire interactors. here we investigated the effects of sex steroids on these interactions during minipuberty the surge of sex hormones that occur along the first six months of life -and up to months of life. we employed a network-based approach for investigating aire-interactors gene-gene relationships and how abundantly co-expressed thymic mirnas covariate with those genes. aire-interactors networks allowed the measuring of gender-related differences in gene-gene expression correlation disclosing relevant differences between minipuberty groups. methods: total rna was extracted from thymic surgical explants obtained from male (m) and female (f) infants -aged - months (groups mm and mf, for minipuberty) and - months (group nm and nf, for nonpuberty) and used in dna microarray assays. gene coexpression network (gcn) analyses were performed for aire and its interactors and for mirna-gene coexpression analysis. the set of genes coding for the aire-targeted proteins was previously identified in tecs by abramson et al. (cell : - , ) . aire-interactors networks were obtained for all groups (link strength cut-off for gene-gene > | . | and for mirnagene < - . ). aire expression in mtecs was quantified by immunohistochemistry. these methodologies are described in moreira-filho et al. (sci rep : , ) . results: the mm x mf networks comparison showed that abundantly expressed mirnas are interacting with the different aire interactor genes in both networks. it is interesting to note that network topology were more similar between nm and nf groups, although aire interacts with only one distinct mirna in each network (mir- - p in the nm group or mir- in the nf group). conversely, in the non-puberty networks the sets of mirnas and their interacting genes are distinct for each network. immunohistochemistry analysis revealed a higher percentage of mtec aire positive cells in the minipuberty groups: i.e. there is a significant difference between mm x nm (p = . ) and between mf x nf (p = . ). conclusions minipuberty and genomic mechanisms shape thymic sexual dimorphism along the first months of life. this process does not involve changes in aire expression between genders, but differences in the interactions of aire with its partners that persist throughout the non-puberty period, probably regulated by mirnas and also by genetic and epigenetic factors. introduction: neutrophils are presumed to defend against aspergillus species by releasing reactive oxygen species (ros) and neutrophil extracellular traps (nets) to degrade fungal hyphae. triazole antifungals synergistically enhance neutrophil mediated hyphal degradation. patients with cgd are particularly susceptible to aspergillus species likely due to their inability to create ros and nets, and in severe cases may not be amenable to antifungal therapy alone. objective: we present a case of severe disseminated aspergillosis in a patient with cgd in whom gt served as an important adjunct to antifungal therapy and bridge to transplant. results: a -year-old boy with known cgd, lost to follow up and nonadherent to prophylaxis, presented acutely with right-sided hemiparesis. neuroimaging revealed an embolic left middle cerebral artery infarction and cardiac magnetic resonance imaging showed extensive vegetations involving both right and left ventricles and atria, with an ejection fraction of %. the patient was admitted to intensive care, started on liposomal amphotericin b, meropenem and vancomycin, and underwent debulking of the intracardiac masses on post admission day (pad) . operative findings showed severe constrictive pericarditis with multiple abscesses and intracardiac vegetations. thorough debridement of the vegetations was undertaken, however some deep seated abscesses in the myocardium were not amenable. operative cultures were positive for aspergillus fumigatus. clinical status remained precarious, with ongoing requirement for inotropic and ventilator support. antimicrobial therapy was refined to voriconazole, with amphotericin b remaining on board until therapeutic levels of voriconazole were achieved. as effective neutrophils are integral in the immune response against aspergillus, the decision was made to start granulocyte transfusions to aid in clinical stabilization prior to hsct. interferon gamma infusions were not administered because of the risks of adverse effects and potentially increasing transplant rejection. gts were started on pad , at a dose of approximately x ^ granulocytes, three times a week. the patient tolerated the infusions well, with no allergic or inflammatory response. neutrophil oxidative burst measured one hour post infusion showed . % mean fluorescent intensity, compared to a baseline of % ( figure ). clinical improvement was seen, with inotrope cessation on pad and extubation to bipap on pad . human leukocyte antigen (hla) allosensitizaton was tested on pad , days after the first gt, with no evidence of hla antibodies. a total of gts were given over months, prior to proceeding to a / hla matched related donor transplant (pad ), with two transfusions given before neutrophil engraftment (anc ) on day + . the patient is now stable months post transplant, with no evidence of graft rejection. he remains on chronic suppressive antifungal therapy, to continue until full lymphoid reconstitution. conclusion: gt may be a useful adjunct to antifungal therapy in patients with impaired neutrophil function with severe invasive aspergillosis, and potentially provide a life sustaining bridge to hsct. methods: subjects were enrolled in irb protocol for rvt- . rvt- was implanted into the quadriceps with immunosuppression. results: subject was normal at q . but had hypocalcemia, an asd, pda, and abnormal ears. the subject received a cord blood transplant mismatched at hla-b and hla-c alleles at age months. subsequently mild graft-versus-host disease (gvhd) developed and was treated with antithymocyte globulin, steroids and cyclosporine. donor t cells developed in low numbers. twelve years later, the subject developed epstein barr virus lymphoma and suffered two relapses. while in remission, subject received unmatched rvt- . two weeks after rvt- implantation, the subject developed an adenovirus infection resulting in skin and gut gvhd, presumably from activation of the cord blood t cells. subject was treated with corticosteroids, cyclosporine, cidofovir and infliximab. four years post rvt- , subject is healthy with genetically recipient t cells/mm and % naïve cd t cells. subject was normal at q . but had an asd, pda, hypoparathyroidism, and no t cells at birth. his genetic defect is unknown. subject was treated with a ric myeloablative, allogenic, unrelated, / cord blood transplant, and a subsequent myeloablative, unrelated / cord blood transplant. hematopoietic chimerism was established without t cell development. rvt- expressed the one allele in the recipient that was not expressed by the second cord donor. the post-thymic transplant course included immune thrombocytopenia requiring rituximab and splenectomy and generalized adenopathy for years but no gvhd. he failed weaning of immunoglobulin replacement. three years post rvt- , he has cd , cd , and cd t cells/mm . he is active in school. subject had absent trecs on newborn screening with cd + t cells/ mm. a single mutation in foxn was identified; she has sparse scalp hair. subject received a / matched unrelated umbilical cord transplant. the post-transplant course was complicated by significant morbidity, and no naïve t cell development. rvt- expressed the one allele in the recipient that was not in the cord blood donor. the subject did not develop gvhd, is healthy and at months has naïve cd + t cells. she had resolution of longstanding norovirus and sapovirus gastroenteritis. conclusion: rvt- can improve t cell immunity after poor or failed correction with allogeneic hematopoietic transplants. in subject , gvhd post rvt- was related to an acute viral infection; cord t cells attacked hla mismatches in the recipient. subjects and were given rvt- matched to recipient alleles that were not expressed in the hematopoietic donor. we hypothesize that thymocytes developing in rvt- , if strongly reactive to the recipient-mismatched allele, are deleted by the bonemarrow-donor dendritic cells (that acquire recipient mhc from the recipient-allele-matched thymic epithelial cells) thereby preventing gvhd. rationale: ctla haploinsufficiency is an autosomal dominant immune dysregulation syndrome characterized by variable phenotypes. here we present a young woman diagnosed with evans syndrome and lymphoproliferation as a child, found to have a novel ctla variant as a young adult, and who developed hypogammaglobulinemia and a bacterial endocarditis while stabilized on ctla- replacement therapy. methods: sequencing of genes, including ctla , in primary immunodeficiency panel. results: our patient was diagnosed with evans syndrome at age with manifestations of anemia and thrombocytopenia recalcitrant to treatment over many years with steroids, cyclosporine, and vincristine. bone marrow biopsy reportedly showed normal trilineage maturation and her symptoms responded for a short time to splenectomy at age . symptoms recurred at age when she was also found to have pulmonary reticular opacities, prominent lymph nodes, and elevated b cells. repeat bone marrow and lymph node biopsies at that time were unrevealing. minor responses to treatment with ivig, rituximab, mycophenolate mofetil and gcsf were noted. at age , she developed varicella-related encephalitis shortly after vaccination. with a strong suspicion of an immune dysregulation syndrome, immune evaluation revealed normal immunoglobulins with good vaccine responses, elevated b cell numbers, normal t cell numbers, and normal mitogen proliferation. ctla sequencing revealed a mutation in exon [c. c>a, p.tyr *] causing a premature translational stop signal, which was consistent with previously reported cases of ctla haploinsufficiency. she was started on rapamycin initially for her cytopenias but was then transitioned successfully to abatacept with almost complete resolution of her anemia, neutropenia, and pulmonary opacities. after months of stable control, she developed a precipitous drop in her platelets and was eventually diagnosed with streptococcus viridans endocarditis of her native mitral valve. this responded to antimicrobial therapy, but eventually needed surgical intervention due to ongoing insufficiency. around this time, she was also found to be newly hypogammaglobulinemic, necessitating ongoing igg supplementation therapy. during successful replacement of her mitral valve with a biosynthetic prosthesis, it was noted that her aortic valve also had evidence of previous disease, implicating a prior endocarditis as part of her clinical syndrome as well. conclusions: in this patient, the presentation of recalcitrant cytopenias, lymphadenopathy, elevated b cells, vaccine-induced viral infections and lung findings precipitated concern for immune dysregulation syndromes and allowed for identification of a novel deleterious ctla mutation. in addition to previously reported clinical findings, our patient presents with the first reported case of repeated endocarditis in the setting of ctla insufficiency disease. given the finding in this patient of prior (unrecognized) disease, regularly screening patients with ctla insufficiency for evidence of cardiac affectation may be prudent. clinical research nurse, johns hopkins university background: the relationship between elevated serum alpha fetoprotein (afp) concentration and age, mortality, genotype and neurologic outcome in ataxia telangiectasia (a-t) patients has remained inconclusive over the past decades, leaving afp as a useful marker for disease diagnosis without further clinical significance. objective: to examine the relationship between afp levels and age, mortality, genotype and neurologic outcome using a data set larger than any prior study. methods: we retrospectively collected data on a-t patients at johns hopkins medical center ( - years of age) with both classical (predicted protein null) and variant a-t. this included serum afp measurements ( serial levels in a-t patients, max observations per patient). mixed model compound symmetry covariance was used for statistical analysis to examine the effect of age at visit on afp levels. subgroup analysis by mutation type, mortality, feeding/swallowing scores as a surrogate for neurologic function, x-ray induced in vitro chromosomal breakage and serum transaminase levels were similarly analyzed. results: significant association between age and afp level was found such that for every year increase in age, afp level increases ng/ml (p< . ). subgroup analysis by mutation type found that the patients with missense mutations showed a negative linear relationship be-tween log afp levels and age (r= - . , p= . ). we found greater afp levels in patients who subsequently died, after controlling for age (least square mean afp level in log scale . greater in deceased patients versus living patients, p= . ). we found a significant decline in feeding score by . units (score range - ) per ng/ml afp increase (p= . ) after adjusting for age. there was no significant relationship between afp levels and serum transaminase levels. conclusion: afp increases with age in a-t patients, though this may not apply to patients with missense mutations. there is a statistically significant increase in mortality and worsened swallowing scores with increasing afp levels, but this remains to be proven clinically significant. here we present a pediatric hae patient who had recurrent abdominal attacks in which constipation, secondary to the adhd medication dexmethylphenidate (focalin), appears to be a trigger. of importance, this is the first pediatric patient with hae to be described as having safely undergone a capsule endoscopy for direct visualization of the gastrointestinal tract. this was done to decrease the risks associated with the more invasive procedure of traditional endoscopy and colonoscopy. case presentation: the patient was an -year-old male with hereditary angioedema who presented with day history of diffuse abdominal pain and nausea. in the ed, patient was in no acute distress. abdominal ultrasound showed severe circumferential thickening of the wall of multiple bowel loops and a large amount of simple ascites. x-ray revealed stool in the colon. he was admitted for pain control and hydration. in the next year, he visited the ed five more times for exacerbations of angioedema of his hand, penis, and bowel. each time, he presented he had underlying abdominal pain and constipation. he was seen by gastroenterology and had a workup that was negative for helicobacter pylori, parasites, and other gastrointestinal infections. to further evaluate his abdominal pain, capsule endoscopy was performed and well tolerated. during an admission in january he received a full inpatient bowel cleanout, after which, his angioedema finally improved. of note, he was diagnosed with adhd and started on dexmethylphenidate (focalin) just prior to this period of recurrent angioedema attacks, and he did not have attacks during the summer months when he was off the medication. discussion: abdominal pain is a common complaint in pediatric hospitals, and further workup consists of endoscopy and colonoscopy. this may be easily accomplished in the general population, however, in patients with hae, these procedures carry greater risk and may be avoided, leading to delayed diagnosis and treatment ( , ) . a newer and less commonly used alternative for direct visualization of the gastrointestinal tract is capsule endoscopy. some benefits are that it does not require sedation, is less invasive, and is less likely to be irritating to the mucosa ( ). additionally, since psychological stress may be a trigger for angioedema attacks, the decreased stress associated with a noninvasive procedure such as capsule endoscopy, makes it safer to use ( ) . limitations of capsule endoscopy include dependence on battery life and its inability to biopsy or administer therapy if needed ( ) . hereditary angioedema treatment consists primarily of avoiding triggers and managing acute episodes. in this first case of hae in a pediatric patient where capsule endoscopy was used, the procedure was well tolerated without any complications. recognizing constipation as a trigger and capsule endoscopy as a safe method of direct visualization of the gastrointestinal tract will help others to control and decrease the severity of their hae attacks as well. a year old male with past medical history of common variable immune deficiency (cvid) and related autoimmune complications, including granulomatous-lymphocytic interstitial lung disease (glild), hepatosplenomegaly, leukopenia, and thrombocytopenia tolerated monthly subcutaneous immunoglobulin replacement as outpatient for several years with infrequent infectious complications. four months ago, he was found to have elevated liver enzymes on routine chemistry. a liver biopsy two months later showed pathology consistent with nodular regenerative hyperplasia (nrh) without overt cirrhosis. a hepatic venous pressure gradient (hvpg) of mmhg was found, consistent with portal hypertension. his hepatitis viral markers were negative, he did not drink, and portal venogram was negative for thrombosis. in early october, the patient was admitted to the hospital with anasarca and tense ascites. he underwent a diagnostic and therapeutic large volume paracentesis and was also found to have spontaneous bacterial peritonitis (sbp) and bacteremia with group b streptococcus. the patients course was complicated by polymicrobial peritonitis, vre bacteremia, fungemia, variceal hemorrhage, hepatic encephalopathy, and hepatorenal syndrome. his hepatic complications from portal hypertension were out of proportion to his liver parenchymal disease. transjugular intrahepatic portosystemic shunt (tips) was considered to alleviate portal hypertension but was not feasible due to his degree of encephalopathy. immunosuppressants such as high dose steroids were given while in the hospital with plans to start rituximab to treat patients glild after he had recovered from the acute infections. unfortunately, after two months in the hospital, the patient succumbed to sepsis and progressive liver failure. this case emphasizes the importance of systematic screening and continued vigilance for hepatic complications in patients of cvid as studies have shown that nrh of the liver is present in more than % of cvid patients who undergo a liver biopsy (pmid: ). a cross-sectional study of patients with primary hypogammaglobulinemia and hepatic dysfunction found that histological findings of nrh were present in % of cvid patients and was associated with portal hypertension in % of cases (pmid: ). another study estimated the minimal prevalence of nrh in cvid patients as % (pmid: ), stating that this was likely a gross underestimate as nrh may also be present in patients with normal liver function tests that are not routinely biopsied. therefore, liver enzyme levels may not anticipate the severity of liver involvement. there is currently no treatment for cvid-related liver disease. other causes of non-cirrhotic portal hypertension, including hepatic veno-occlusive disease and budd-chiari syndrome should be ruled out or treated in cvid patients presenting with hepatic disease. in the case of hepatic nrh in cvid patients, early detection could lead to earlier interventions (such as tips prior to hepatic encephalopathy), to mitigate complications. we describe the application of epigenetic quantification of t regulatory (treg) cells in addition to cd +, cd +, cd + t cells, b cells, nk cells, monocytes and neutrophils from as little as μl of fresh, frozen or dried blood. the method yields identical results to flow cytometry from fresh blood samples of a healthy donor cohort, with the advantage of being more sensitive and precise with limited amount of blood and minimal sample preparation (sci transl med ). we have used this method ) to immunophenotype patients with early onset immune regulatory disorders (pird) and primary immune deficiency (pid), and ) to evaluate cell subsets reconstitution early after hematopoietic stem cell transplantation (hsct). patients with immune dysregulation, polyendocrinopathy, enteropathy, x-linked (ipex) and ipex-like pird were evaluated by analyzing the treg-specific demethylated region (tsdr) of the foxp locus in the total of cd + t-cells. despite the dysfunctional foxp mutated protein, ipex patients exhibited elevated treg/cd + cell ratios which seemed to correlate with disease severity. in contrast, most of the patients with ipex-like symptoms without foxp mutations exhibited decreased treg/cd + cell ratios -in line with the possible central pathogenic role of treg function and number in pird. using epigenetic quantification of cd +/b-and nk cells, out of confirmed scid and xla cases were correctly identified within a cohort of newborn dried blood spot (dbs) samples ( % sensitivity, % specificity). the method identified one delayed onset scid as well as a xla case that were missed by combined trec/krec testing. epigenetic immune cell quantification missed one scid case with maternal engraftment that was identified by combined trec/krec testing. abnormally elevated treg/cd + ratio was also detected in a dbs from a newborn who was subsequently confirmed to be affected with ipex syndrome. when applied to serial blood samples during engraftment and reconstitution post-hsct, the epigenetic method allowed identification of the different blood cell subsets, including treg cells, at earlier time points than flow cytometry according to current clinical practice. this opens the way to a better understanding of the correlation between early immune reconstitution events and graft vs. host disease or viral reactivation, earlier than with the current methods, in different types of hsct. these studies underscore the suitability of epigenetic immune cell quantification for accurately measuring multiple immune cell types from limited blood sample sources. we propose this method as uniquely suitable for novel molecular diagnostic applications in settings with limited fresh blood sample or limited cell number, at the point of care as well as for newborn screening. we evaluated a -year-old male with hyperpyrexia, hypertrichosis, conical hypodontia, and a history of illnesses concerning for nemodeficiency syndrome. starting at six months of age, he suffered recurrent episodes of acute otitis media (non-typeable hib and actinobacter iwolffli), pneumonia, and rsv bronchiolitis. whole exome sequencing demonstrated a de novo heterozygous c. g>a (p.r q) mutation in the eda-receptor (edar) gene not present in the parental dna. his physical exam findings and mutation were consistent with hypohidrotic ectodermal dysplasia (hed), a rare genetic condition characterized by abnormal development of skin, teeth, hair, and sweat glands. hed is caused by defects in the ectodysplasin-a (eda)-nfkb signaling pathway but is not typically associated with immune deficiency. consistent with this, immunophenotyping showed normal sub-populations of t-, b-, and nk-cells. immunoglobulin and complement levels were quantitatively appropriate. he had normal mitogen-induced lymphocyte proliferation and normal antibody response to pneumococcal vaccination. nk-cell studies demonstrated robust cytotoxicity. however, nasal mucosa biopsy showed diffuse squamous metaplasia and the absence of ciliated epithelial cells. we hypothesize that recurrent infections in our patient arose from impaired mucociliary clearance due to a ciliary defect. this case raises the possible association between edar variants and ciliary dysfunction. it also underscores the importance of evaluating the immune status of hed patients with recurrent infections which could mimic nemo-deficiency and have broad implications about clinical management. the rapid pace of new gene discovery and phenotype expansion for primary immunodeficiency diseases (pidds) creates challenges for genetic testing and variant interpretation. whereas well-described clinical case reports in published literature have traditionally served as the source of phenotypic data used for variant interpretation, for pidds the causal variants are often private to the patients family and thus the sole source of phenotypic information for a novel genetic variant is frequently the history provided by the clinician on the test requisition form. taking into account such heterogeneous information during variant interpretation requires establishing objective criteria for its inclusion as part of the variant interpretation process. to this end, we adapted our laboratorys preexisting, evidence-based variant classification framework, called sherloc, by developing point-based criteria for the inclusion of clinical information such as a patients phenotype, familial segregation patterns, and whether the variant is inherited or de novo in the patient. as part of this process, we defined clinical criteria for pidd genes. here, we illustrate the application of this method and the importance of integrating clinical information into variant interpretation. between april and october , our commercial diagnostic laboratory performed immunological genetic tests, and information about the patients clinical history was provided in ( %) of these orders. restricting our analysis to just the genes for which case report information is currently used in variant interpretation, these tests revealed variants, ( %) of which were classified as pathogenic or likely pathogenic (p/lp). information from case report descriptions, segregation patterns, and de novo status were applied for %, % and % of p/lp variants, respectively. in ( %) cases, the clinical information provided by the clinician on the test requisition form was used as evidence in the classification of the patients variant as p/lp. ten variants were initially classified as being of uncertain significance and reclassified following receipt of further clinical information or testing of additional relatives. in addition, suspicious variants of uncertain significance were identified in which one or two additional patient case reports would allow for reclassification from uncertain significance to p/lp. these data illustrate the importance of providing good quality clinical information to the genetic testing laboratory both at the time of sample submission and following the receipt of genetic test results. background: cartilage-hair hypoplasia (chh) is a skeletal dysplasia with combined immunodeficiency, variable clinical course and increased risk of malignancy, mostly non-hodgkin lymphoma and basal cell carcinoma. there is a paucity of long-term follow-up data, as well as knowledge on prognostic factors in chh. objective: we conducted a prospective cohort study in finnish patients with chh to describe clinical course and analyze risk factors for adverse outcomes. methods: we recruited finnish patients with chh in - and performed clinical follow-up in - . we obtained health information from finnish national medical databases (covering time period of - ), the finnish cancer registry and the cause-of-death registry of the statistics finland and analyzed all patients' health records. standardized mortality ratios (smrs) were calculated based on the population data. primary outcomes included immunodeficiencyrelated death (from infections, respiratory diseases or malignancies), the development of lymphoma and the development of skin cancer. results: the study cohort included males and females. median age at recruitment was . yrs (range weeks - . yrs) and median duration of follow-up for the surviving patients was . yrs (range . - . yrs). half of the patients ( / , %) had no symptoms of immunodeficiency, while ( %) and ( %) patients manifested symptoms of humoral or combined immunodeficiency respectively, including six cases of late-onset immunodeficiency. in a significant proportion of patients ( / , %), clinical features of immunodeficiency progressed over time. of the patients with non-skin cancer, eight had no preceding symptoms of immunodeficiency. altogether patients had deceased (smr= . , % confidence interval (ci)= . [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] including deaths due to pneumonia (n= ), malignancy (n= , smr= , %ci= . - ) and lung disease (n= , smr= , %ci= . . malignancy was diagnosed in / ( %) patients, mostly lymphoma (n= ) and skin cancer (n= ). severe short stature at birth (compared to normal, smr/smr ratio= . , , symptoms of combined immunodeficiency (compared to asymptomatic, smr= ( %ci= . - ) vs smr= . ( %ci= . - . ), hirschsprung disease (odds ratio (or) . , %ci= . - ), pneumonia in the first year of life or recurrently in adulthood (or= . / , , and autoimmunity (or= , %ci= . - ) in adulthood associated with early mortality. in addition, recurrent pneumonia in childhood was associated with the development of lymphoma, while warts and actinic keratosis were associated with the development of skin cancer. birth length standard deviation score correlated significantly with the age at the diagnosis of first malignancy (p= . ), lymphoma (p= . ) and skin cancer (p= . ), demonstrating that patients with shorter birth length developed malignancies at an earlier age. conclusions: patients with chh have high mortality due to infections and malignancies, but also from lung disease. some subjects present with late-onset immunodeficiency or malignancy without preceding symptoms of immune defect, warranting careful follow-up and screening for cancer even in asymptomatic patients. we provide clinicians with the risk factors for adverse outcomes to assist in management decisions. autoimmune lymphoproliferative syndromes (alps and related disorders) are characterized by insufficient apoptosis due to defects in the fas apoptosis pathway. fadd deficiency (omim ) is an autosomal recessive disorder resulting from a mutation in fas-associated protein with death domain (fadd), the adaptor protein involved in fas signaling to caspases and . we present a case of fadd deficiency identified by whole exome sequencing with a novel genetic mutation we describe two brothers with recurrent febrile episodes accompanied by seizures and respiratory compromise. the older sibling initially presented with status epilepticus following the measles mumps rubella vaccination later experiencing similar episodes until his demise at months of age. the younger sibling, who is unvaccinated, presented at months with fever, rash, vomiting, and diarrhea. he developed status epilepticus with respiratory depression that required intubation. he also had enlarged cervical lymph nodes that regressed with antibiotics and steroids. he recovered from that episode but subsequently had a series of similar illnesses with fevers, altered mental status and seizures. with the exception of elevated hhv igg, extensive infectious workup up in all instances was negative. previously described fadd deficiency patients demonstrate an alps like phenotype with increased circulating double negative t cells, lymphocyte apoptosis defects, elevated fas ligand and il , encephalopathy, functional asplenism but no splenomegaly or lymphadenopathy. our patients clinical and laboratory findings were similar. he had normal igg and iga, decreased igm, and lack of isohemagglutinins. absolute cd + count is elevated, with elevated percent of cd + tcr+ cd -cd -. normal mitogen and antigen t lymphocyte stimulation, but with defect in pokeweed induced b cell proliferation. fas ligand and il level are increased (see table ). no hepatosplenomegaly, but howell jolly bodies were detected in peripheral blood indicating functional hyposplenism. whole-exome sequencing revealed two different genetic alterations in the fadd gene: a maternally inherited nonsense mutation predicted to severely truncate the protein and a paternally inherited missense mutation in codon . although this paternal mutation has not been described as pathogenic, a different variant in same nucleotide of fadd has been associated with fadd deficiency (reference ). there are very few cases in the literature of fadd deficiency patients and the overall prognosis is poor compared to classical alps patients, as these patients are at significant risk of deadly sepsis from encapsulated organisms or death from neurologic complications. of the fadd deficiency patients described in the literature, several died prior to years old. while pneumococcal prophylaxis may reduce the risk of sepsis, hematopoietic stem cell transplant has been reported for patients with fadd deficiency (reference ), and is being considered for our patient. rationale: hcuvp is a patient product-introduction program that provides cuvitru® (immune globulin subcutaneous [human], % solution [ig gly]) free of charge for the first infusions to eligible patients with primary immunodeficiency disease (pid). using patient data from this ongoing program, our analysis described the clinical characteristics and infusion parameters of pediatric and adolescent patients who were initiated on ig gly through hcuvp. methods: hcuvp eligibility criteria were: patients aged years old, with a primary icd- -cm code verifying diagnosis of pid, and no current or prior use of ig gly at program initiation. data from patients who received the first ig gly infusion between january , , and september , were included. data from patients receiving infusions after october , were censored. descriptive statistics were calculated for patients demographic and clinical characteristics and prescribed and actual infusion characteristics by age group (< years and years). results: in total, patients who completed all infusions were included in the analysis, of whom were aged < years. among those who previously received immunoglobulin (ig) therapy, a greater percentage of patients aged < years were treated with intravenous ig therapy (n= ; %) compared with adult patients (n= ; %) before initiating ig gly. nine patients aged < years were treatment naïve. the mean infusion volume per site was lower among patients aged < years ( years: . ml; years: . ml; and years: . ml) than among patients aged years ( years: . ml and years: . ml). however, the mean infusion rate per site was similar between patients aged < years ( xmen disease (x-linked immunodeficency with magnesium defect, epstein-barr virus infection and neoplasia) is a primary immune deficiency caused by mutations in magt and characterized by chronic infection with epstein-barr virus (ebv), ebv-driven lymphoma, cd t-cell lymphopenia, and dysgammaglobulinemia. magt gene codifies to magt protein, a mg +-selective transporter, expressed in the human immune system, specifically in the spleen and the thymus. functional studies have established the key role of magt in t cells and natural killer (nk) cell activation. upon cd + t-cell receptor stimulation, magt mediates a transient mg + influx that is necessary for phospholipase c gamma (plcy ) activation, which drives ca + rise and downstream signaling. this mg + influx also regulates cytotoxic functions of nk and cd t cells through nkgd , reason why these patients have impaired cytolytic responses against ebv. eleven male xmen patients have been described. we present the case of a -year old hispanic infant with a pathogenic variant in magt gene that clinically manifested with early pneumocystis jirovecii and cytomegalovirus (cmv) interstitial pneumonia, and ebv chronic infection with good response to intravenous immunoglobulins supplementation without hematopoietic stem cell transplantation or gene therapy. laboratory study highlights low levels of nkg d ligands. the objective of this case report is to broaden the spectrum of clinical presentation of xmen disease, that manifests initially as a combined immune deficiency (cid) and evolved with a favorable course of the disease with intravenous immunoglobulins supplementation therapy and chemoprophylaxis with trimethoprim-sulfamethoxazole. introduction: lysinuric protein intolerance (lpi) is a recessively inherited disorder of the cationic amino acids transporter subunit y+lat caused by variants in the slc a gene. the disease is characterized by protein-rich food intolerance has a heterogeneous presentation. the clinical findings are a result of depletion of lysine, ornithine, and arginine. symptoms can include hyperammonemia, failure to thrive, protein aversion, neurologic disease, and lung disease. there is also evidence that inflammatory manifestations are mediated through upregulation of nfb, il , and tnf that occur independent of intracellular arginine levels and can lead to lifethreatening episodes of hemophagocytic lymphohistiocytosis (hlh). case presentation: a -year-old male presented with history of anxiety, depression, eating disorder, delayed puberty and complex partial seizures. due to poor nutrition and failure to thrive, a gastrostomy tube was placed. following commencement of enteral feeds, he presented with altered mental status, bilateral mydriasis, hyperreflexia, and agitation which lead to a picu admission. ammonia peaked as high as μmol/l and episodes ceased with cessation of enteral feedings. prior to enteral feeds, he had been self-restricting protein in his diet. biochemical testing was consistent with lpi and illumina next-generation sequencing revealed compound heterozygous variants in slc a (p.s lfs* and p.e dfs* ). hyperammonemia resolved quickly with cessation of protein intake and high rate dextrose infusion without the need for ammonia scavenging agents. he was subsequently started on proteinrestricted enteral feeds. at diagnosis he did not have any respiratory symptoms, ct scan of chest showed patchy areas of groundglass opacification that was suggestive of early pulmonary alveolar proteinosis (pap). bronchoalveolar lavage demonstrated foamy, cloudy pink fluid and elevated bronchioalveolar macrophages on cell differential. his clinical course and slc a genotype led to suspicion for smoldering hlh. the findings of elevated ferritin, hypertriglyceridemia, decreased fibrinogen, splenomegaly, elevated il- receptor, decreased nk cell function, along with hemophagocytosis on bone marrow biopsy confirmed the diagnosis. because of his pap and hlh, in addition to dietary modifications, a trial of il- beta inhibition (anakinra) at mg/kg/day was initiated. follow up ct scan of chest months after initiation of anakinra showed complete resolution of pulmonary groundglass opacifications and pap. bone marrow evaluation showed continued hemophagocytosis in spite of the normalization in ferritin, soluble il- receptor, nk function, and triglycerides levels. overall, he is significantly improved on daily anakinra and no longer meets criteria for hlh or pap. discussion: recent data has shown in y+lat models that thp- macrophages and a airway epithelial cells upregulate il and tnf regardless of intracellular arginine content. this suggests that inflammatory manifestations may continue independent of dietary modifications. we present a year old patient with newly diagnosed lpi who was treated dietary modification and anti-il therapy resulting in resolution of hlh and pap. more research is needed to see if long-term il blockade that can consistently control both the immunologic and pulmonary manifestations of lpi and positively impact morbidity and mortality. learning objective: recognize that symptoms of bartonella endocarditis and associated complications can share features of certain immunocompromising conditions. case description: an -year-old caucasian boy with history of repaired pulmonary atresia and aortic root dilation was diagnosed with pancytopenia and splenomegaly during a brief hospitalization for atypical pneumonia. pancytopenia persisted, splenomegaly worsened, and five months after presentation, he developed hypertension and renal insufficiency. he was diagnosed with hypocomplementemic, diffuse sclerosing and crescentic glomerulonephritis and was started on mycophenolate mofetil with improvement in kidney function and stabilization of cytopenias. as part of a comprehensive immune work-up, alps (autoimmune lymphoproliferative syndrome) panel was sent and demonstrated elevated double-negative t (dnt) cells with out of positive immunologic criteria for alps. neither targeted sequencing for alps and alpslike disorders nor whole exome sequencing revealed pathogenic mutations. by age , the patient remained on mycophenolate, but developed failure to thrive, with weight dropping from th percentile to less than rd percentile. he was hospitalized again for low-grade fever, increased work of breathing, left shoulder pain and fatigue and was found to have right lower lobe pneumonia. pancytopenia worsened, and he was started on cefepime and azithromycin without improvement in symptoms. echocardiogram revealed vegetations in his pulmonary conduit and bilateral branch pulmonary arteries, but multiple blood cultures were negative. upon further history, the patient reported contact with kittens. bartonella henselae titers and polymerase chain reaction (pcr) from blood were sent and were both positive. he completed a -week course of gentamicin, -month course of ceftriaxone, and was transitioned to doxycycline and rifabutin. after initiating antimicrobial therapy, his weight and energy significantly improved, his blood bartonella pcr became negative, and his splenomegaly resolved. approximately one year later, the patient underwent pulmonary artery conduit replacement and bartonella pcr testing of the tissue specimen was positive. he has had sustained weight increase, resolution of hypocomplementemia and splenomegaly, decrease in dnt cell frequency from > % to . %, and improvement though not resolution of cytopenias. he currently remains on doxycycline and rifabutin and continues treatment with mycophenolate. discussion: alps is characterized by defective lymphocyte apoptosis and clinical features such as lymphadenopathy, splenomegaly, hepatomegaly, cytopenias, and glomerulonephritis. the hallmark laboratory finding is expansion of dnts. our patient met criteria for a probable alps diagnosis based on the presence of both required criteria (chronic splenomegaly and elevated dnt cells) and secondary additional criteria (typical immunologic findings noted on alps panel). pediatric cases of bartonella henselae endocarditis have been associated with splenomegaly, cytopenias, and glomerulonephritis which mimic many features of monogenic immune dysregulatory disorders. the diagnosis of bartonella endocarditis in our patient therefore raises the question of whether his immunosuppression predisposed him to infection or if his entire clinical presentation can be explained by bartonella endocarditis. physicians taking care of patients with immune dysregulatory disorders should consider bartonella endocarditis in the differential diagnosis of onset or exacerbations of immune dysregulation. rationale: while fever is considered a sign of infection, many individuals with primary immunodeficiency (pi) anecdotally report a lower than normal average body temperature. on immune deficiency foundation (idf) friends and idf pi connect research forum online, pi patients report a diminished fever response even when other signs of infection are present. there is limited knowledge about the average body temperature in persons with pi. however, the implications of missing an infection in those with pi is well established. methods: study investigators partnered with patient investigators to design a prospective cohort study to determine whether body temperature differed between persons living with and without pi. three hundred fifty adults with pi were recruited from idf and one adult household member without pi was also recruited. mckesson digital oral thermometers (model - bgm) were provided and used to record temperatures in all participants three times a day for five consecutive days. descriptive statistics were calculated. median body temperatures were compared between the two cohorts at each time point using mann-whitney test. results: data from households were used for analysis ( . % participation rate). the pi population was largely female ( . %) with a median age of years and largely caucasian population ( . %). the non-pi population was largely male ( . %) with a median age of years and largely caucasian population ( . %). pi diagnoses included cvid ( . %), hypogammaglobulinemia ( . %), igg subclass deficiency ( . %), selective iga deficiency ( . %), specific antibody deficiency ( . %), agammaglobulinemia ( . %), chronic granulomatous disease ( . %), combined immunodeficiency ( . %), and complement deficiency ( . %). a total of individuals with pi ( . %) reported a lower than normal non-sick body temperature, while individuals with pi ( . %) reported a normal (between °f - °f) non-sick body temperature. a total of individuals with pi ( . %) reported absence of fever with infection, while individuals ( . %) reported a normal fever response with infection. the median body temperature was significantly higher for the pi patients in the morning, but not evening or bedtime, reading in of the days (monday: pi = . °f vs. non-pi = . °f, p = . ; tuesday: pi = . °f vs. non-pi = . °f, p = . ; wednesday: pi = . °f vs. non-pi = . °f, p = . ; thursday: pi = . °f vs. non-pi = . °f, p = . ; friday: pi = . °f vs. non-pi = . °f, p= . ). conclusions: despite the limitations of this non-clinical study, individuals with pi are knowledgeable about their conditions and can offer unique insights and direction to researchers. this study demonstrates that collaboration with patient advocacy groups may facilitate patient-centered and patient-driven research with high participation among the target population. introduction: familial mediterranean fever (fmf) is a hereditary condition characterized by recurrent episodes of painful inflammation caused by mutations in the pyrin (mefv) gene. alterations in the mefv gene affect pyrin production leading to recurrent fevers and painful inflammation in the peritoneum, synovium, and pleura. amyloidosis may also develop as a complication. arabic, turkish, armenian, and sephardic jewish populations are most commonly affected. homozygosity for mefv mutations are associated with a more severe course. there is a paucity of information regarding pediatric fmf in the literature. case: we present a case of a -year-old male with minor speech delay diagnosed with compound heterozygous fmf. patient was initially referred due to recurrent fevers and infections. at months of age, he was hospitalized with septic shock requiring intubation secondary to adenovirus. at months of age, the patient began to have recurrent fevers every to weeks, leading to multiple blood draws and courses of antibiotics prior to referral. at , , and months of age, he developed three separate episodes of febrile seizures. a total of - lifetime episodes of acute otitis media occurred prior to bilateral myringotomy tube placement. four episodes of streptococcus pyogenes pharyngitis confirmed by throat culture preceded tonsillectomy. no oral ulcers, joint pain, or abdominal pain were reported. no other infections such as pneumonia, sinusitis, uti, non-viral gastroenteritis, fungal infections, or skin infections were reported. both parents are ashkenazi jewish and a maternal history of early miscarriage was noted. family history was negative for immunodeficiency, malignancy, and autoimmunity. the patients vital signs and physical exam were unremarkable. serology indicated leukocytosis of . k/l with elevated monocytes of cells/l, elevated eosinophils of cells/l, and slightly elevated cd t cell count of cells/l. neutrophil, cd t cell, b cell, nk cell enumeration, and immunoglobulin panel were normal for age. tetanus, diphtheria, rubella, streptococcus pneumoniae, and haemophilus influenzae b titers were protective. genetic analysis identified that the patient was compound heterozygous for the e q and v mutations in the mefv gene. family was instructed to keep a fever diary. colchicine . mg once a day was given initially, then increased to . mg once a day for inadequate response. loose stools were observed while patient was maintaining a lactose free diet so he was switched to colchicine . mg bid with resolution of loose stools. apart from two occasions when his colchicine dose was missed, the patient remained afebrile at his follow up visits. conclusion: we present a pediatric case of compound heterozygous fmf (e q and v mefv mutations) in an otherwise healthy -year-old male of ashkenazi jewish background, initially symptomatic at months of age. individuals who are compound heterozygous for the e q and a second mevf mutation are generally symptomatic, although severity cannot be predicted. additional pediatric research on symptomatic heterozygous and compound heterozygous fmf is recommended. natural killer (nk) cells are innate lymphocytes that play a key role in defense against virally-infected cells and in tumor surveillance. nk cells can be divided in two subsets. the majority of nk cells in peripheral blood expressed intermediate levels of cd and are referred to as cd (dim). these nk cells are responsible for nk cell cytotoxicity. a minor population of nk cells express very high expression of cd and are referred to as cd (bright). these nk cells are responsible for cytokine production and are precursors to cd (dim) nk cells. a few immunodeficiencies have been described in which there are abnormal nk cell subsets, such as autosomal dominant gata deficiency where cd (bright) nk cells are absent and irf where there is a paucity of cd (dim) nk cells and relative expansion of cd (bright) nk cells. here we present a patient with an absence in cd (bright) nk cells secondary to cd deficiency. our patient is a -year-old african american female born to non-consanguineous parents. the patients past medical history is significant for chronic lung disease secondary to prematurity, recurrent acute otitis media, failure to thrive and congenital hypothyroidism. family history is significant for an older sister that presented at age with ebv-associated hodgkin lymphoma whose treatment was complicated by chronic activated ebv infection and who ultimately underwent hematopoietic stem cell transplantation (hsct). our patient presented with pancytopenia, fever, lymphadenopathy and splenomegaly. she was found to have ebv viremia with greater than , copies in whole blood by pcr. she was treated with two doses of rituximab followed by etoposide and dexamethasone as a bridge to hsct. whole exome sequencing demonstrated a homozygous mutation in cd . cd is a member of the tumor necrosis factor receptor family and influences the function of t cells, b cells and nk cells. in nk cells, cd is primarily expressed in cd (bright) nk cells. cd deficiency is an autosomal recessive disorder associated with persistent symptomatic ebv viremia, including ebv-driven hemophagocytosis and lymphoma, hypogammaglobulonemia and specific antibody deficiency. our patients immune evaluation prior to initiation of chemotherapy and immunosuppression was notable for very elevated igg, iga and igm. despite hypergammaglobulonemia patient had only out of protective titers against streptococcus pneumoniae. the patient had pan-lymphopenia with appropriate percentages of lymphocyte subsets. assessment of her b cell subsets showed a slight increase in the percentage of transitional b cells/plasmablast and a nearly complete absence of cd -expressing b cells. her nk cell phenotyping demonstrated a complete loss of cd (bright) nk cells with reduced nk cell cytotoxicity, comparable to what has been previously reported in patients with gata deficiency. previous reports of patients with cd deficiency denote normal nk cell numbers with normal to moderately reduced nk cell cytotoxicity, however, cd deficiency causing a specific loss of the cd (bright) nk cell subset has not been previously reported. cd deficiency should be consider in patients with ebv driven disease and abnormal nk cell studies. introduction/background: the transcription factor ikaros is encoded by the ikzf gene and plays a crucial role in lymphopoiesis. somatic, and more recently also germline mutations of ikzf are associated with a hematologic malignancies, most notably b-cell precursor acute lymphoblastic leukemia. germline mutation in ikzf was first reported as a monogenic cause of human disease characterized by marrow failure and immune deficiency in a single neonate in . subsequently, mutations leading to haploinsufficiency were discovered to underlie a proportion of patients with cvid and low b cell numbers, and dominant-negative mutations have been observed to cause more severe combined immune deficiency phenotypes. at this time, there is very little known regarding allogeneic hematopoietic cell transplantation (hct) outcomes for patients with severe dominant-negative ikzf mutations. concerningly, ikaros deficiency has been observed to have a negative impact on graft versus host disease in mouse models. objective: to describe allogeneic stem cell transplant outcomes in patients with the dominant-negative ikaros mutation. methods: we collected transplant data from patients who underwent allogeneic hct at transplant centers around the world. results: patients underwent allogeneic hct using a variety of conditioning regimens. patients received bone marrow (n= ) or cord blood (n= ) grafts from an hla-matched sibling donor (n= ) or single allele hlamismatched unrelated donor (n= ). neutrophil engraftment occurred between day + and + post-transplant. platelet engraftment occurred between day + and + except in one patient who did not have return of normal platelet counts due to underlying liver dysfunction. all patients were documented to have greater than % whole blood donor chimerism at a median of days (range - days) following transplant and maintained > % donor chimerism until last follow-up. only one patient developed grade ii acute gvhd. no patients developed chronic gvhd. one patient died approximately year post transplant related to cryptosporidium cholangitis which existed prior to hct. at the most recent follow up of the surviving patients (range: . - . y), ivig had been discontinued, antimicrobial prophylaxis had been stopped, and patients had received routine vaccinations. they all had excellent performance status. conclusions: allogeneic hct may be a safe option to consider for patients with dominant-negative ikaros mutation as there does not appear to be an increased risk of death or gvhd. moreover, -out-of- of the transplanted patients are alive and well and show no features of the disease. however, because of the limited number of patients evaluated and the retrospective nature of this analysis, our data do not allow firm conclusions to be made, and further studies will be needed to evaluate outcomes in larger cohorts. introduction: when evaluating patients with t-cell lymphopenia, we often are concerned about defects in lymphocyte production and function, especially in the setting of frequent infections. here we outline a case demonstrating t-cell lymphopenia due to increased loss, which should be considered in the differential diagnosis. case report: we report a -year-old male who initially presented with recurrent, right-sided pneumonias requiring frequent hospital admissions including severe episodes necessitating intensive care unit admission. his work up for the pneumonias included a bronchoscopy revealing normal anatomy with minimal inflammation, and a chest ct with mild peribronchial wall thickening. as his pulmonary disease progressed, he developed a persistent, productive cough with expectorated mucous plugs that were plastic-like in appearance. while his pulmonary symptoms responded to steroids, his mucous plug production persisted. sputum cultures were intermittently positive, isolating cryptococcus neoformans and aspergillus niger. he underwent vats and wedge biopsy, concerning for recurrent aspiration. an immunologic evaluation initially demonstrated normal t-and b-cell counts, but serial evaluation of his lymphocyte population demonstrated low cd + cells (ranging - cells/cumm), and low normal cd cells (ranging - cells/cumm) with normal b-and nk-cell numbers. further t-cell evaluation revealed normal ratios of naive and memory p o p u l a t i o n s ( c d c d r a + % , c d c d r o + % , cd cd ra+ %, cd cd ro %), normal trec ( copies per ^ cd cells) and normal thymic emigrants (cd cd cd ra+ : , normal - ), indicative of sufficient thymopoiesis. mitogen and antigen stimulation assays demonstrated normal responses to phytohemagglutin, concanavalin a, and pokeweed mitogen, with a low lymphocyte response to candida. he had normal quantitative immunologlobulins, normal diphtheria, tetanus and streptococcus pneumonia titers. his dihydrorhodamine flow cytometry and fish for chromosome q . deletion were negative. given normal function and thymic output, his immunologic profile was concerning for t-cell loss. our patient was registered with the undiagnosed disease network, and had a second review of his lung biopsy, concerning for plastic bronchitis. subsequent lymphatic imaging demonstrated abnormal lymphatics within the bilateral clavicular space, right greater than left, with questionable partial thoracic duct, explaining his unilateral symptoms. he was diagnosed with plastic bronchitis secondary to abnormal lymphatic drainage, with lymphatic fluid filling his airways and secondary t-cell loss. discussion: plastic bronchitis is a rare and potentially fatal disorder, seen commonly after the fontan procedure for congenital heart disease. this process has resulted in t-cell loss into the airway and subsequent t-cell lymphopenia. in patients with fontan-related protein losing enteropathy, multiple immune abnormalities have been described including reduced immunoglobulins, lymphopenia, and selective cd lymphocyte deficiency. similar findings have been reported in patients with lymphatic malformations. although the impact of t-cell loss on adaptive immunity is not entirely known, there is no indication of increased risk for atypical infections. given his normal mitogen assay, our patient did not start prophylactic antibiotics. he continues to have symptomatic episodes with lymphopenia, but has had no opportunistic infections, and remains stable with an aggressive pulmonary regimen. we conclude by reiterating the importance of considering t-cell loss in patients presenting with lymphopenia, particularly with evidence of normal thymopoeisis and t-cell function. introduction: granulomatous disease (gd) has been described with a variable incidence ( . - . %) in patients with common variable immunodeficiency (cvid). an increase in malignancies has been reported in cvid patient cohorts, particularly for lymphoma, reported in . - . % of the cvid patients depending on the cohorts. prior analysis of a cohort of cvid patients included patients with gd (gd+). in these, there was a suggestion of more cases of lymphoma ( . %) when compared to cases without (gd-) ( . %) although the difference was not statistically significant (p=. ). objectives: compare the frequency of lymphoma in gd+ and gd-patients in the cvid patient cohort from the usidnet registry. methods: we submitted a query to the usidnet registry requesting deidentified data for patients with the diagnosis of cvid, through august . statistical analysis was performed on spss, with comparisons done with pearson chi-square or fisher's exact test, depending on the sample sizes, using an alpha level of . . results: a cohort of cvid patients from the usidnet registry was analyzed. ninety-one patients ( . %) were gd+. overall, patients ( . %) had a malignancy diagnosis, of these ( . %) with lymphoma. lymphoma was present in / gd+ patients ( . %) versus / gdpatients ( . %) (p=. ). overall malignancy was present in / gd+ ( . %) versus / ( . %) (p=. ). discussion: in the cohort of cvid patients from the usidnet registry, we found a frequency of lymphoma of . %, which is in the range of previously described cohorts. the frequency of lymphoma was . % in patients with gd, higher than the . % frequency for gd-patients, but these differences were not statistically significant. our identified frequency of lymphoma in gd+ patients was lower than the one previously identified in the cvid patient cohort, but with similar proportional differences between gd+ and gd-patients. despite no statistical significance, the frequency of lymphoma, as shown here and elsewhere, was higher in cvid patients gd+ than gd-in both studies, with no full understanding of this increased risk of lymphoma. expanding this analysis to larger groups of cvid patients may help to confirm, or deny a more robust association, which may have a meaningful impact in the outcomes of this particular population. introduction: patients with refractory pericarditis have been treated with intravenous immunoglobulin (ivig) or interleukin receptor antagonist (anakinra) with limited and transient benefit. separate or combined therapy with subcutaneous immunoglobulin (scig) and interleukin (il) inhibitor (rilonacept) for refractory pericarditis in a cohort of patients has not been previously described. case descriptions: patients were referred for recurrent pericarditis refractory to traditional therapies at ages ranging from to years. they all had multiple serious sequelae of their pericarditis and abnormal immune parameters including hypogammaglobulinemia, poor responses to vaccines, poor mitogen induced lymphocyte proliferation, and/or b cell lymphopenia. the patients had varied past medical histories and associated conditions. patients were started on ig, with some initiated on ivig, though all were transitioned to hyaluronidase-facilitated scig (hyqvia). patients were then started on either anakinra or rilonacept with patients continuing on rilonacept and remaining on anakinra. all patients had complete or near complete resolution of their pericarditis on dual therapy for greater than year. the markedly elevated il prior to therapy seen in all of the patients normalized post-therapy. some patients had elevated il prior to therapy that also improved post-therapy. patient who has also been diagnosed with familial mediterranean fever (fmf) has stopped both therapies for greater than year with no further episodes of her pericarditis. discussion: patients with recurrent refractory pericarditis and signs of immunodeficiency and autoinflammatory disease on laboratory testing responded to dual therapy with hyqvia and rilonacept or anakinra resulting in resolution of pericarditis. inflammasome and immune abnormalities may be implicated or associated with recurrent pericarditis and may respond to targeted therapies. chief, laboratory of clinical immunology and microbiology, idgs, dir, niaid, nih, bethesda, md, usa hypomorphic recombination activating gene (rag ) mutations result in residual t-and b-cell development in both humans and mice and have been found in patients presenting with delayed-onset combined immune deficiency with granulomas and/or autoimmunity (cid-g/ai). recent studies have shed light on how hypomorphic rag mutations alter the primary repertoire of t and b cells, but less is known about their effect on immune dysregulation in targeted organs. in order to investigate the role of these mutations in determining intestinal disease, we set out to evaluate gut immunity and microbiota interplay in rag mutant hypomorphic mice. we evaluated two mouse models carrying homozygous rag mutations (r q and r w), corresponding to human mutations (r q and r w, respectively) described in patients with cid-g/ai. both mutations fall in the coding flanksensitive region of the rag c-terminal domain. on the basis of aminoacid properties and in vitro studies, the r q mutation has demonstrated a moderate effect on rag protein stability while the r w mutation resulted highly disruptive. analysis of intestinal pathology in rag mutant mice (niaid animal protocol lcim e) revealed different degrees of spontaneous colitis, with the most severe inflammatory infiltrate observed in mice carrying the most disruptive mutation, r w. colonic inflammation was characterized by crypt elongation, epithelial hyperplasia, and an abundant inflammatory infiltrate extending to the colonic lamina propria, with occasional crypt abscesses. a significant increase in activated cd hicd lcd + t cells expressing the gut homing receptor was observed in mesenteric lymph nodes (mlns) of both mutant strains, and was especially prominent in r w mutant mice. additionally, the proportion of mln cd + t regulatory (treg) cells was increased in both mouse models. finally, mln of mutant mice contained a high number of myeloid cells (cd b+ ) along with a decreased number of b + b cells, and these abnormalities were also more prominent in r w than in r q mice. in summary, we have shown that rag mutant hypomorphic mice present with different degrees of inflammatory bowel disease, with the mouse model carrying the most disruptive mutation presenting with the most severe phenotype. we are currently performing studies to evaluate the impact of rag mutations on microbiome composition and diversity in these mouse models of cid-g/ai. background: hypogammaglobulinemia or low serum immunoglobulin g (igg) levels either inherited (primary) or acquired (secondary) is associa t e d w i t h i n c r e a s e d i n f e c t i o n r a t e s . p r i m a r y ( °) hypogammaglobluinemia can be caused by many primary immune deficiencies (pid) including combined variable immune deficiency (cvid), while secondary ( °) hypogammaglobluinemia can be caused by many acquired conditions such as lymphomas, leukemias, or chemotherapies and other immunosuppressive drugs. immunoglobulin replacement therapy (irt) has been the mainstay of treatment in patients with hypogammaglobulinemia by reducing infection through replenishing the quantitative igg. there are other applications of ig therapy such as in autoimmune diseases, where the mechanism of action is thought to be ig mediated immunomodulation. innate immune cells have shown to be involved in such mechanism, but whether irt modulates adaptive immune cells in patients with hypogammaglobulinemia is not well known. hypothesis: irt has an immunomodulatory effect on t-cell function and proliferation in patients with hypogammaglobulinemia. methods: blood from thirty patients with °(n= ) or °(n= ) hypogammaglobulinemia recruited from the immunodeficiency clinic at the ottawa hospital was drawn for peripheral blood mononuclear cell (pbmc) isolation, before starting irt and minimum weeks after starting irt. data regarding igg level, number and type of infections after receiving irt was collected. pbmcs were analyzed using flow cytometry for quantitation of t-cell subset. cultured and anti-cd /cd stimulated pbmc were also analyzed for extracellular and intracellular cytokine production, measured by e l i s a a n d f l o w c y t o m e t r y, r e s p e c t i v e l y. c o m b i n e d cytomegalovirus, epstein-barr virus and influenza virus (cef) peptides were used to study specific t-cell responses. anti-cd / cd stimulated pbmc were used for celltrace t-cell proliferation a s s a y s . d a t a w a s g r o u p e d b a s e d o n n a t u r e o f hypogammaglobulinemia i.e. °or °. results were compared between before and after irt using wilcoxon matched-pairs signed rank test. results: irt was not found to significantly alter proportion of treg, cd +, or cd + t-cell populations or activation state as measured by cd ra/r expression. however, irt was found to significantly increase expression of intracellular ifn-y in cd + and cd + t-cells post-cd /cd stimulation in °(p = . ), but not in °h ypogammaglobulinemia patients. there was no change in extracellular il- and il- cytokine production in both groups. in contrast, cd + tcells in °hypogammaglobulinemia patients showed significantly higher expression of intracellular ifn-y and tnf-a post-cef viral peptide stimulation (p = . ). cd + and cd + t-cell proliferation after cd /cd stimulation was found to be decreased after irt for both groups (p = . & p = . ). conclusions: our results suggest that irt can alter cd + and cd + t-cell function with differential effect in patients with °o r °hypogammaglobulinemia in addition to replenishing serum igg level. more experiments assessing cytotoxicity of t-cells will be conducted to further study t-cell subset function as well as bcell function. these laboratory results will be analyzed for association with clinical outcomes. uploaded file(s) uploads background: severe congenital neutropenia (scn) is a rare immunodeficiency disorder characterised by the extremely low absolute neutrophils count (anc) less than . x /l. the clinical feature of scn is recurrent bacterial infections and the patients the risk of leukemia development. the incidence of scn is estimated to be in individuals. mutations in more than genes have been described causing scn and it is either recessive, dominant or x-linked inheritance. case presentation: we described an years old malaysian girl who presented with recurrent abscesses over the whole part of the body, recurrent oral candidiasis, growth failure and recurrent pneumonias since months old. she also had history of a few episodes of acute tonsillitis, chronic suppurative otitis media and herpes zoster infections. throughout her age, she had persistent neutropenia less than . x /l but in few occasions, her anc elevated up to more than . x /l . she was treated as autoimmune neutropenia, respectively due to few positive results of autoimmunity workout such as antinuclear antibodies (ana) and double stranded dna (dsdna) but eventually later to be negative. later at the age years old, whole exome sequencing was performed and confirmed by sanger s e q u e n c i n g , f o u n d a h e t e r o z y g o u s v a r i a n t i n e l a n e gene(c. g>t; p.gly ter), an autosomal dominant which was described to cause scn. both parents do not carry this mutation, hence, it is a de novo mutation. currently, she had few on and off recurrent infections. despite that, she is relatively well and on prophylaxis antibiotic. conclusion: to our knowledge, we report for the first time a malaysian girl with scn, with confirmed mutational analysis of the elane gene. the delayed diagnosis might be due to the insufficient awareness of the phenotypic presentation of this rare disease. moreover, the genetic analysis is not available in malaysia and need to be done outside of the country. this case demonstrates the importance of the genetic analysis which may help in improving the diagnosis and management of the patient. ( ) submission id# professor of paediatrics and immunology, university college london; great ormond street hospital nhs trust; orchard therapeutics, london, uk background: ada-scid is a rare genetic disorder which causes severe combined immunodeficiency. historically, ada-scid has been treated using enzyme replacement therapy (ert) followed by allogeneic hematopoietic stem cell (hsc) transplant (hsct) from a matched related donor (mrd) or, if none is identified, a non-mrd (matched/mismatched unrelated or mismatched related donor). we developed a self-inactivating lentiviral vector (lv), in which a codon optimized human ada cdna is driven by the short form of the elongation factor- alpha (efs) promoter (efs-ada lv). the drug product (otl- ), composed of autologous hscs transduced ex vivo with the efs-ada lv, was evaluated in a prospective, historically-controlled phase i/ii clinical trial in ada-scid pediatric subjects. we report safety and efficacy at months in ada-scid subjects treated with lentiviral gene therapy (gt) compared to a historical cohort of ada-scid patients treated with hsct. methods: twenty subjects ( male, female; mo . yrs) were treated with gt. autologous cd + hscs were isolated from bone marrow and pre-stimulated with cytokines before transduction with efs-ada lv. busulfan was administered at a single dose ( mg/kg) prior to infusion of otl- . the control group included patients ( . mo . yrs) treated with allogeneic hsct (mrds n= , non-mrds n= ) at great ormond street hospital, uk (n= ) or duke university childrens hospital, usa (n= ) between . results: at months, overall survival (os) and event-free survival (evfs), defined as survival in the absence of ert reinstitution or rescue allogeneic hsct) were statistically significantly higher in the gt group compared with the hsct group (table) . successful engraftment of genetically modified hsc was observed in all gt subjects at months, which persisted over months, based on vector gene marking in granulocytes (median . copies/cell [range . - . ] at months) and peripheral blood mononuclear cells (median . copies/cell [range . - . ] at months), and was associated with increased red blood cell ada enzyme activity and metabolic detoxification from deoxyadenosine nucleotides. over months, none of the gt subjects required peg-ada ert reinstitution and % were able to stop receiving immunoglobin replacement therapy (igrt), whereas % hsct patients required rescue hsct or reinstitution of peg-ada ert, and % were able to stop receiving igrt (table) . nine subjects in the gt group experienced a serious adverse event (sae), most frequently infections and gastrointestinal events; only one was considered treatment-related. in the gt group, there were no events of autoimmunity during the study. due to the autologous nature of the product, there was no incidence of graft vs host disease (gvhd) in the gt group; whereas patients in the hsct group experienced acute gvhd and experienced chronic gvhd events, one of whom died. conclusions: treatment with lentiviral gt for ada-scid is well tolerated and has a favorable benefit-risk profile at months based on sustained gene correction and restoration of immune function, as well as improved os and evfs compared with hsct (mrd or non-mrd) at months. background: ada-scid is a rare genetic disorder that causes severe combined immunodeficiency, with minimal or absent b cell function. prior to, and often after, treatment with allogeneic hematopoietic stem cell (hsc) transplant (hsct) or autologous ex vivo hsc gene therapy (gt), patients are managed with enzyme replacement therapy (ert) and immunoglobulin (ig) replacement therapy (igrt). we evaluated a gt treatment with autologous hscs transduced ex vivo with a self-inactivating lentiviral vector (lv), in which a codon optimized human ada cdna is driven by an internal short form of the elongation factor- alpha (efs) promoter ("efs-ada lv"). at months follow-up, pediatric ada-scid subjects treated with gt were compared to a historical cohort of ada-scid patients treated with hsct. here, we report on b cell reconstitution in these cohorts. methods: twenty subjects ( male, female) aged mo - . yrs received gt. autologous cd + hscs were isolated from bone marrow and pre-stimulated with cytokines before transduction with efs-ada lv. genetically modified cells were administered after conditioning with single dose busulfan ( mg/ kg). the control group included patients aged . mo to . yrs treated with hsct at great ormond street hospital (uk) (n= ) or duke university children's hospital (us) (n= ) between - . the hsct patients received an allogeneic transplant from matched related donors (mrds) (n= ) or non-mrds (n= ). subjects continued to receive igrt post-gt until a clinical decision was made to stop, factoring in b cell reconstitution, general medical condition and seasonal infections. results: by month , in the gt group, % had stopped treatment with igrt compared to % in the hsct group overall. by months and , higher proportions of gt-treated subjects had stopped igrt ( % and %, respectively) compared with mrd hsct patients ( % and %, respectively) and non-mrd hsct patients ( % at both timepoints) (table) . in the gt group, vector gene marking was detectable in peripheral blood mononuclear cells within months and persisted at months post-infusion (median . copies/cell [range . - . ]), suggesting successful gene modification. as evidence of b cell reconstitution, iga and igm levels in peripheral blood sera more than doubled by months, from . mg/dl (range to ) to . mg/dl (range to ) and . mg/dl (range to ) to . mg/dl (range to ), respectively. additionally, antibody response following tetanus vaccination, was evaluated in subjects. all subjects mounted a protective response to the vaccine (median antibody response . iu/ml [range . to . ]), based on a normal threshold of . iu/ml (hammarlund clin infect dis ) and a laboratory reference range ( . to . iu/ml). conclusions: gt with autologous hscs transduced ex vivo with efs-ada lv resulted in b cell reconstitution, as evidenced by doubled iga and igm production at months, cessation of igrt in % of patients by months, and protective specific antibody responses to tetanus vaccine in patients that were evaluated. background: x-linked chronic granulomatous disease (xcgd) results from mutations in cybb encoding the gp phox subunit of phagocyte nadph-oxidase. attempts to treat xcgd with gene therapy (gt) using transduced autologous hematopoietic stem cells (hsc) transduced ex vivo with a gammaretroviral vector have met with limited efficacy due to transient engraftment of gene corrected hscs, gene silencing, and vector insertion-mediated activation of oncogenes leading to myelodysplasia. we developed a novel self-inactivating (sin) lentiviral vector (g xcgd lv) with a chimeric cathepsin g/cfes myeloid-specific promoter driving gp phox expression from a codon optimized cdna. following transplant of g xcgd lv ex vivo transduced autologous hscs into busulfanconditioned xcgd patients, there was long-term restoration of oxidase activity in peripheral blood polymorphonuclear neutrophils (pmn) at months in of severely affected xcgd patients without evidence of genotoxicity. here we present data about the multiple assays used to assess quality and quantity of restoration of pmn oxidase activity. methods: similar trials of gt with g xcgd lv were initiated in the uk (n= , plus compassionate use patient) and usa (n= ). all patients had histories of inflammatory disease and severe, persistent infections (some non-responsive to conventional therapy at time of gt). g-csf plus plerixafor-mobilized cd + hscs were transduced with ex vivo g xcgdlv. subjects received myeloablative conditioning with singleagent busulfan, targeted to net area-under-the-curve of , ng/ml*hr. freshly prepared or cryopreserved quality-tested genetically-modified hsc, manufactured on-site, were administered intravenously. pmn oxidase activity post-gt was assessed by p-nitroblue tetrazolium (nbt) reduction, dihydrorhodamine (dhr) flow cytometry assay, and quantitative ferricytochrome c assay (ferric) measurement of superoxide generation. results: we report results for patients (aged - years) with - . years of follow-up; two additional patients were treated but died within three months of gt from complications deemed related to pre-existing diseaserelated co-morbidities (severe pulmonary disease and anti-platelet antibodies). within month post-gt, oxidase (+) pmn were present in peripheral blood based on nbt testing and dhr flow cytometry. expression of the corrective transgene was confirmed by flow cytometry using antibody detection of gp phox. quantitative biochemical measurements of oxidase activity were also confirmed in some samples using the ferric assay, demonstrating quantitative levels of superoxide production per corrected cell that were within the normal range. functional testing of oxidase burst activity using dhr fluorescent assays was applied serially to follow levels of corrected pmn where oxidase activity per corrected cell also were in the normal range. all patients had > % pmn dhr+ within one month, which remained stable for most patients over the follow-up period ( figure) . follow-up demonstrated sustained stable persistence of - % oxidase burst positive neutrophils in of surviving subjects at months, with restoration to clinically beneficial levels (defined as % of pmn being dhr+) in these patients as of december . conclusion: these results demonstrate corrected pmn function within month in x-cgd patients treated with autologous gt. pmn oxidase activity was sustained at levels which restore biochemical function and provide clinically beneficial levels of immunity for months in / patients. the formulation for igsc % was developed based on the knowledge acquired from the formulation of grifols currently licensed % immune globulin (human), gamunex®-c; however, the protein concentration was increased from % to % to facilitate efficient subcutaneous administration. gamunex-c has an extensive record of safety and tolerability when administered intravenously and subcutaneously for greater than years in diverse patient populations. the igsc % manufacturing process employs the same purification steps as gamunex-c and was demonstrated to be robust and to provide an igg product with the required potency, purity, and quality. the formulation excipient characteristics and compatibility with the drug product have been well established. glycine has been an excipient of intramuscular immune globulin (human) for fifty years and intravenous immune globulin (igiv) for over twenty years. the igsc % formulation has low buffering capacity, and a low ph was selected to achieve a product with low aggregates, low fragments and viscosity suitable for subcutaneous administration. to improve visual clarity, the igsc % formulation contains a small amount of polysorbate (ps ), which is widely used in biopharmaceutical products. subcutaneous administration of the igsc % formulation has been well tolerated in clinical studies. objectives: the goal was to provide the pid population with a new % immunoglobulin liquid product for subcutaneous administration (igsc %). methods: igsc % is manufactured using the current manufacturing process for gamunex-c, followed by an additional concentration step so that the product can be formulated at a higher protein concentration. igsc % and gamunex-c batches were produced at full industrial scale and then subjected to a series of analytical testing including assessment of purity, composition and neutralizing activity. results: the igsc % and gamunex-c manufacturing processes and formulations have preserved the igg integrity, molecular characteristics and potency. the manufacturing processes have eliminated lipids, alcohols, and acetate and coagulation factor impurities, including fxia, which were undetectable by either specific or global methods. the igsc % and gamunex-c batches were % gamma globulin by agarose membrane electrophoresis, and have a subclass distribution similar to normal plasma and acceptable specific antibody content. igsc % was shown to be primarily monomer plus dimer igg ( ± %) with minimal aggregate or fragment, which confirms that appropriately gentle processing conditions were used during the concentration of % igg solutions to % igg. conclusions: igsc % is a highly concentrated igg solution with characteristics comparable to gamunex-c, but with twice the igg concentration in order to facilitate subcutaneous administration with reduced volumes and shorter infusion times. analytical testing demonstrates suitable potency, purity, and neutralizing activity for a number of specific antigens. funding: this study was funded and conducted by grifols, a manufacturer of % immunoglobulin for subcutaneous administration. disclosure: all authors are employees of grifols. frequent respiratory tract infections and seizures cause recurrent hospitalizations in these children and are typically considered a result of neurological impairment and poor airway clearance. evaluation of these patients for immunodeficiency is not a common clinical practice. here we report combined immune deficiency in patients with mds and recurrent respiratory tract infections. case presentation case : a boy with mds was initially referred at age months for an abnormal newborn screen with low t cell receptor excision circles (trec) for severe combined immunodeficiency (scid). initial evaluation revealed moderate cd + and cd + t cell lymphopenia (figure ). initial immunoglobulins levels were normal. he was placed on antiseizure medications. he later developed recurrent and severe respiratory tract infections starting in infancy. at months of age, he developed hypogammaglobulinemia ( figure ). in addition, t cell counts progressively decreased and stayed around cells/ul. immunoglobulin replacement therapy started at months of age. hospitalizations due to respiratory tract infections significantly decreased. case : a -year-old boy with mds had recurrent bacterial and viral respiratory infections which required numerous hospitalizations including intensive care unit stays. newborn screening for scid was negative. he had been on anti-seizure medications. immunologic evaluation at years of age revealed low total cd + cells and cd + t cells (cd +: cells/ul[normal range - cells/ul], cd +: cells/ ul[normal range - cells/ul]), hypogammaglobinemia (igg: mg/dl[normal range - mg/dl]), and non-protective igg levels to tetanus, varicella and pneumococcus serotypes. immunoglobulin replacement therapy started at years of age which resulted in reduced frequency and severity of respiratory infections, and improved quality of life. discussions: t cell lymphopenia and hypogammaglobulinemia were seen in both our cases of miller-dieker syndrome. to our knowledge, immune deficiency has never been reported in mds. one of our cases suggests that low t cell counts may start as early as at birth and may be detected by newborn screening. hypogammaglobulinemia may be primary or secondary due to antiepileptics. both children had reduced frequency and severity of respiratory infections and improved quality of life after immunoglobulin replacement highlighting the importance of screening and early management of immunodeficiency. conclusion: miller-dieker syndrome is likely another syndromic primary immune deficiency disorder. a high index of suspicion with early screening and management of immunodeficiency may be beneficial for children with miller-dieker syndrome. uploaded file(s) uploads this prospective, multi-center, open-label study assessed the pharmacokinetic (pk), safety, and tolerability of immune globulin subcutaneous (human), % caprylate/chromatography purified (igsc %) in subjects with primary immunodeficiency (pi). the objectives were to determine a weekly subcutaneous (sc) dose of igsc % that is noninferior to the intravenous (iv) dose of immune globulin injection (human), % caprylate/chromatography purified (igiv-c %) and to determine the steady state trough igg levels after igsc % and igiv-c % infusions. there were possible phases. if not on a qualifying igg regimen at enrollment, subjects (n= ) were required to enter the run-in phase, receiving igiv-c % to achieve steady-state before entering the iv phase to determine steady-state area-under-the-curve (auc) of iv infusions. subjects with a qualifying igiv-c % regimen ( - mg/kg) (n= ) directly entered the iv phase for steady-state iv pk assessments. upon completion of the iv pk assessments subjects entered the sc phase, receiving weekly doses of igsc % for up to weeks, with steady-state auc determined at the th dose. igsc % was not associated with any reports of serious local infusion site reactions (isrs). the majority of local isrs were mild-to-moderate. igsc % (at a dose conversion factor of . ) provided equivalent exposure to igiv-c % as assessed by steady-state auc - days, with % higher mean igg trough values, lower fluctuations in igg concentrations and the flexibility of at home administration. igsc % was well tolerated with a safety profile comparable to igiv-c %. clinicaltrials.gov identifier: nct disclosure: kecia courtney, elsa mondou, and jiang lin are employees of grifols, a manufacturer of igsc %. grifols is the sponsor of this study. background: in two reports described the deficiency of adenosine deaminase (dada ) as early-onset lacunar strokes, intermittent fevers, livedoid rash, and early onset polyarteritis nodosa (pan). since these first reports, the clinical spectrum has dramatically expanded to include antibody deficiency, liver disease, vasculopathy, pure red cell aplasia, cytopenias, and lymphoproliferative disease. methods: forty-two patients were enrolled in an irb approved study at the nih. sequencing of ada , the gene encoding adenosine deaminase (ada ), was performed in all patients. information was obtained by chart review of all clinical, serologic, and radiographic testing. results: all patients had germline biallelic loss of function mutations in ada , leading to absent or significantly decreased protein expression and function of ada . the cohort comprises females ( %) and males ( %). there were sibling pairs and families with affected individuals. twenty-seven patients had a history of at least one ischemic stroke and experienced a hemorrhagic stroke. the average age at the time of first stroke is . years (range months - years), and the average number of strokes is (range - ). no new strokes have occurred in patients on anti-tnf therapy. skin manifestations occurred in % of patients and include livedo ( %), cutaneous vasculitis resembling pan ( %), and raynauds ( %). hepatomegaly ( %) and splenomegaly ( %) were also notable. portal hypertension was observed in ( %) patients, with patient requiring a spleno-renal shunt for a massive variceal bleed. abdominal mra revealed arteritis and aneurysm in / patients evaluated; patients developed bowel necrosis. peripheral vasculopathy was seen in patients, with one requiring amputation of gangrenous digits. the most common immune abnormality seen in this cohort is hypogammaglobulinemia ( %); patients have low igg, patients have low igm, and patients have low iga. ten of these patients are on immunoglobulin replacement. specific antibody responses to vaccines were inadequate in / patients challenged. lymphocyte phenotyping revealed decreased class-switched memory b cells in / patients ( %) tested. however, there was no relationship between absolute number of class switched memory b cells and hypogammaglobulinemia or infection frequency. hematologic abnormalities include transfusion depended anemia ( %), neutropenia ( %), lymphopenia ( %), and thrombocytopenia ( %). seven patients developed pancytopenia, presented with pure red cell aplasia, and developed aplastic anemia. three patients have undergone bone marrow transplant, with two of those patients requiring a second transplant for graft failure. conclusions: the spectrum of dada has expanded from strokes, intermittent fever, and cutaneous manifestations to include portal and systemic hypertension, immune deficiency, cytopenias, vascular abnormalities, and bone marrow failure. while initiation of anti-tnf therapy improves inflammatory markers, and no new strokes have occurred while on therapy, cytopenias do not seem to improve. bone marrow transplantation should be considered in patients with findings of bone marrow failure, although transplant of our patients has been complicated by immune mediated neutropenia. disease manifestations are heterogenous, making a comprehensive evaluation critical to our understanding of this disease. given the increase in neonatal diagnosis of athymia, clinical care is provided by the referring medical centers prior to rvt- implantation and patients return to the referring centers earlier after rvt- . this creates the need for clear, concise guidelines for the care of these patients. primary goals of pre-transplantation clinical care are ( ) management of pre-existing medical needs such as feeding difficulties, airway obstruction, congenital cardiac defects and developmental disabilities; ( ) management of symptoms related to oligoclonal recipient t cell expansion (autologous gvhd/atypical complete digeorge anomaly) and ( ) prevention of infections. most deaths in the pre and early post-transplantation period are secondary to pre-existing infections. necessary surgical and medical procedures (ie cardiac surgery, hearing aids) should not be delayed. for the first to months after rvt , patients have profoundly low naïve t cell numbers and may require immunosuppression to prevent rejection of rvt- by oligoclonal recipient t cells. immunosuppression needs to be closely monitored and titrated for desired effect while minimizing side effects such as renal toxicity, electrolyte abnormalities and hypertension. t cell counts should be performed every months and are used to guide weaning of immunosuppression. most patients with successful transplants develop greater than /mm naïve t cells by months post rvt- . infection prevention, clinical stability and optimal nutrition are critical for lasting engraftment. clinical guidelines have been developed to address immunosuppression, management of autologous gvhd symptoms (gut, skin and liver), preservation of renal function, and developmental considerations. after the development of naïve t cells, patients should continue to be monitored regularly by an immunologist. patients may develop autoimmune complications such as thyroid disease and transient cytopenias. while risk of complications related to viral infections is greatly decreased after development of naïve t cells, patients with comorbidities (central venous access device dependence, tracheostomy, chronic lung disease) continue to require complex care from multidisciplinary teams. medical conditions associated with athymia but not alleviated by thymus transplantation, such as hypoparathyroidism or cardiac defects, may require lifelong medical care. lastly, patients must be evaluated for readiness for killed and live vaccines. transplant outcomes are influenced by the clinical condition at the time of rvt- implantation and optimization of immunosuppression, nutrition and clinical stability in the first months following rvt- . clinical care that maintains a well-nourished, clinically stable, infection free patient yields the best chance for successful t cell development. guidance documents supporting these goals ensure patients are best prepared to receive rvt- and develop long lasting thymic function. hemophagocytic lymphohistiocytosis (hlh) is a life-threatening disease of immune dysregulation characterized by unchecked inflammatory responses leading to end-organ dysfunction. primary hlh results from inherited mutations that impair capacity for immune regulation whereas secondary hlh arises from inappropriate response to an immune stimulus such as infection, malignancy or autoimmunity. we report a -monthold male who presented with symptoms of hlh as an initial manifestation of congenital disorder of glycosylation (cdg) due to mutations in the gene component of oligomeric golgi complex (cog ) resulting in cog -cdg (cdg-iij). a -month-old male with history of mild motor delay presented with days of fever, vomiting, and diarrhea. initial evaluation identified highly elevated ferritin and triglycerides, transaminitis, coagulopathy, and hyperammonemia. he subsequently developed generalized seizures. liver and bone marrow biopsies demonstrated erythrophagocytosis consistent with hlh. immunologic evaluation was notable for mild hypogammaglobulinemia, neutropenia, thrombocytopenia, and anemia. serum cd levels and nk functional studies were later found to be normal. the patient was initially treated with ammonia-scavenger therapy and fresh frozen plasma (ffp) for coagulopathy with subsequent intravenous immunoglobulin and dexamethasone several days later. within hours after starting ffp, the patients ferritin level declined sharply. hyperammonemia and transaminitis also resolved, and his fever curve improved. additional immunosuppression was considered, but not initiated due to the patients ongoing clinical improvement. over the next months, the patient experienced two further acute episodes of fever, liver dysfunction, coagulopathy, and sepsis physiology. the second episode was successfully treated with ffp, though no clear infectious trigger was identified. the third episode occurred days after routine vaccinations. the patient had prolonged hypotension requiring ionotropic support that resolved after receiving daily ffp, and hypoxia with pleural effusions that resolved after a single treatment with protein c concentrate. as the patient had met / clinical diagnostic criteria for hlh, but also had a history of hyperammonemia, he underwent concurrent biochemical and genetic evaluation for both primary hlh and inborn errors of metabolism. whole exome sequencing identified compound heterozygous mutations in cog , part of an oligomeric protein complex involved in golgi apparatus structure and function. cog mutations have previously been reported in two patients with autosomal recessive cog -cdg (cdg-iij), who were described to have similar clinical symptoms of hypotonia, seizures, coagulopathy, and liver dysfunction, as well as recurrent infections. subsequent immune phenotyping while the patient was healthy was notable for slightly low numbers of nk cells, but normal cd a mobilization and perforin/granzyme b expression in vitro. our patient represents a novel presentation of cdg due to cog defect with associated immune dysfunction manifesting as recurrent episodes of inflammatory crisis with features of hlh. cdg and inborn errors of metabolism should be considered during diagnostic evaluation for patients with hlh symptoms, as cdg patients may develop acute episodes of severe inflammation, in the absence of cellular regulatory defects, for which ffp and protein c concentrate may have therapeutic value. of the deaths with identifiable causes, ( %) were related to infections. the rate of death per person-year was . . the most common autoimmunity-related complication was sweets syndrome, seen in patients ( %) with anti-ifn-g autoantibodies. sixteen of those patients ( %) had recurring sweets syndrome. additionally, patients ( %) developed lymphatic obstruction, which continued to recur in patients ( %). seven patients ( %) in this study did not have anti-ifn-g autoantibodies. the median [iqr] age of autoantibody-negative patients was [ , ] years and patients ( %) were female. none of the autoantibody-negative patients developed new infections during follow-up. at the end of the follow-up period, none of the patients had active/progressive disease and patients ( %) had died. conclusions: ninety-one percent of hiv uninfected thai patients with disseminated ntm infection with or without other opportunistic infections had detectable anti-ifn-g autoantibodies. about one third of patients with autoantibodies to ifn-g had recurrent infections during follow-up. after approximately years of follow-up, % of patients with anti-ifn-g autoantibodies had inactive disease following multi-drug antibiotic therapy while % had active/progressive disease and % had died. patients with anti-ifn-g autoantibodies are at risk for recurrent infections and autoimmunity-related complications. therefore, longterm follow-up is recommended. life-long secondary antibiotic prophylaxis may be required to prevent recurrence of infection in the setting of persistent anti-ifn-g autoantibodies. the study of early t cell development in patients with severe t cell immunodeficiencies is challenging because of the rarity of these diseases, the difficulty to obtain hematopoietic stem cells (hscs), and limitations in the assays to assess in vitro differentiation of hscs to mature t cells. we recently developed a serum-free system that allows faithful analysis of sequential steps of t cell differentiation. in this system, artificial thymic organoids (atos) are generated, based on the d aggregation and culture of a delta-like canonical notch ligand (dll )-expressing stromal cell line (ms -dll ) with cd + cells isolated from bone marrow (bm) samples of normal donors (nd). in this project, we set out to evaluate the possibility of using the ato system to study t cell differentiation in patients carrying t cell defects, in order to define the exact steps of t cell development affected by different genetic defects. using the ato system, we studied in vitro t cell differentiation from cd + cells obtained from patients carrying defects that are intrinsic to hematopoietic cells (rag , rag , ak , il rg) or that affect thymus development (digeorge syndrome, dgs). the ak -deficient patient showed a markedly decreased viability in cd + cells and a very early defect in t cell development, already at the pro-t cell stage. this defect was very similar to that observed in a patient carrying a null il rg mutation who was reported to show autologous reconstitution after unconditioned haploidentical hsc transplantation. in contrast, cd + cells from a patient carrying a missense il rg mutation and with a leaky scid phenotype were capable of differentiating into mature t cells in vitro, although with -fold decreased efficiency as compared to normal donors (nd). interestingly, in the patient carrying the null il rg mutation, we noticed very few cells that could reach full maturation, with an absolute number of cd + tcrab+ cells around -times less than in nd. at variance with pro-t cells (that failed to express the gc protein), these mature t cells did express normal levels of gc, suggesting that they may have derived from residual cd + cells from the bm donor. in addition, cd + cells from the patients carrying rag and rag hypomorphic mutations were able to differentiate to cd +cd + double positive cells, but not to cd +tcrab+ cells. finally, the dgs patient showed a completely normal in vitro t cell differentiation, confirming that t cell deficiency reflected thymic abnormalities. in summary, our data show that the ato system could be extremely useful in determining whether the lack of t cells in patients with unknown gene defects reflect hematopoietic or thymic intrinsic problems, and may therefore provide critical evidence in deciding whether hsc or thymus transplantation is warranted, even without knowing the actual gene defect. introduction: ataxia-telangiectasia (at) is an autosomal recessive disorder caused by mutations in the ataxia telangiectasia mutated (atm) gene, which aids in detection and repair of dna damage. at is characterized by progressive cerebellar ataxia, oculomotor apraxia, choreoathetosis, conjunctival telangiectasias, variable degrees of t-cell lymphopenia (tcl) and immune compromise. patients are at an increased risk for malignancy, particularly leukemia and lymphoma, and are unusually sensitive to ionizing radiation. with the advent of trecbased newborn screening (nbs) for scid, at patients are being recognized with asymptomatic tcl in early infancy. objectives: we present an older child with at and chronic granulomatous lesions and discuss how this may be avoided in individuals with at diagnosed following abnormal nbs. case report: a y/o male was born at term following an uncomplicated twin pregnancy and delivery, prior to institution of scid nbs. he demonstrated mild gross motor and speech delay as an infant and was diagnosed with at at age . he had received all routine immunizations, including live vaccinations. he developed granulomatous skin lesions at age , initially small papules on his cheeks and ears, which subsequently formed large disfiguring plaques on sun-exposed areascheeks, arms and hands (fig ) . following an extensive workup, his lesions were found to be secondary to a mutated vaccine-strain rubella (ra / ) based on bp genotyping, previously described in other immunocompromised individuals [perelygina/sullivan et al. jaci ] . his lesions have been refractory to multiple treatments including nitazoxanide. he is currently on daily oral and topical steroids, tmp/smx and ivig. retrieval of his nbs for trec determination revealed that he would have screened positive [mallot/puck et al. j clin immunol ] . when first measured at age , cd t-cells were low, /ul, with cd /ul and cd /ul. b and nk cell numbers were normal. since april , cases of at were seen at ucsf in infants with non-scid tcl on nbs. these males and female were all born at term and discharged from well-infant nurseries. at was diagnosed at - months of age. their initial trecs ranged from - /ul (normal with perkinelmer enlite kit > ), and all had low t-cells on initial flow cytometry ( - cd /ul, ref range> ) with decreased cd ( - /ul) and cd ( - /ul) t-cells; however naïve t-cells were present, ruling out typical scid and raising concern for non-scid tcl. three infants also demonstrated low b-cells (< - /ul), while nk cells were normal in all. two are currently receiving ivig, one of whom is also on tmp/smx. all have avoided not only rotavirus but also mmr and varicella live vaccinations. conclusions: at is now often diagnosed in infants with low trecs on scid nbs, prior to neurologic manifestations. benefits of early diagnosis include avoidance of live vaccines, including mmr, which led to the debilitating granulomas in our older patient. additionally, patients receive prompt immunologic monitoring and treatment, avoidance of unnecessary radiation, specialty referrals and family genetic counseling. while there is no cure for at, ongoing research may bring neuroprotective treatments in the future. introduction: subcutaneous immune globulin %, ig gly, was well tolerated in the phase / north american study in patients with primary immunodeficiency diseases (pidd). here we assess comorbidities, use of concomitant medications, infusion parameters, and tolerability in advanced age patients ( y) treated with ig gly in the north american study. methods: patients aged years with pidd received weekly ig gly infusions at volumes ml/site and rates ml/h/site for~ . years in the north american study (nct ). the medical history at baseline, medical conditions that were ongoing (defined as comorbid events), use of concomitant medications, adverse events (aes), tolerability, and infusion parameters were assessed by age: in advanced age patients ( y; n= ), adult ( < y; n= ), and pediatric/adolescent patients (< y; n= ). results: the mean number of medical history events at baseline was higher in advanced age patients ( . events/patient; events in patients) versus adult ( . events/patient; events in patients), and pediatric/adolescent patients ( . events/patient; events in patients). of these, the medical conditions that were ongoing at baseline (comorbid events) were also higher in the advanced age patients ( . events/patient; events in patients) versus adult ( . events/ patient; events in patients), and pediatric/adolescent patients ( . events/patient; events in patients). in the advanced age patients, neurological comorbidities ( events) were the most common, followed by those related to eyes, ears, nose, and throat ( events), gastrointestinal ( events), and musculoskeletal comorbidities ( events). concomitant medications were given to treat a preexisting condition in all patients in the advanced age group ( medications in patients). despite the higher mean number of comorbid conditions, infusion parameters in the advanced age patients were comparable to those in the adult age group. median maximum infusion rates and infusion volumes/site were comparable in the advanced age patients ( ml/h/site; . ml/site) and adults ( ml/h/site; ml/site); lower infusion rates and volumes/site were reported in the pediatric/adolescent patients ( . larger infusion volumes and faster infusion rates were not associated with increases in causally related local aes in the advanced age group, consistent with the trends seen in the pediatric/ adolescent and adult patients. conclusions: despite the higher mean number of comorbidities in advanced age patients with pidd, ig gly was infused at relatively high rates and volumes and was well tolerated. introduction: hyqvia (ighy; immunoglobulin infusion % with recombinant human hyaluronidase [rhuph ]) is an immunoglobulin (ig) replacement therapy approved for patients with primary immunodeficiency diseases (pidd) that allows larger infusion volumes, up to ml/site, and has improved ig bioavailability compared with conventional subcutaneous ig products. a post-authorization safety study is being conducted in the united states to acquire long-term safety data on ighy and to assess prescribed administration regimens in routine clinical practice. infusion characteristics and treatment-related adverse events from an interim analysis are reported here. methods: patients aged years with pidd receiving ighy were included in this ongoing, prospective, non-interventional, open-label, uncontrolled, multicenter study. as a part of routine clinical practice, patients are treated with ighy according to standard medical care and their treatment regimen is at the discretion of the treating physician. adverse events (aes) are collected from enrollment to study completion/discontinuation using a subject diary and assessed at every study visit (every months or standard practice). aes are assessed based on seriousness, severity, and causal relatedness to ighy. the presence of anti-rhuph antibody is evaluated on a voluntary basis. treatment preferences for various attributes of ig therapy were assessed annually using a treatment preference questionnaire. results: a total of patients were enrolled at us study sites (data cutoff date: august , ). infusions were self-administered at home ( %) or at the clinical site ( %) most commonly using -week infusion intervals ( . %). the mean maximum ig infusion rate was . ml/h and the mean ig dose was mg/kg bodyweight/ weeks. the mean number of infusion sites used for administration was . and mean infusion duration was . hours. most infusions ( . %) were administered without a rate reduction, interruption, or discontinuation due to aes. there were no serious aes (saes) related to ighy. sixteen patients experienced a causally related non-serious local ae ( . %; . events/patient-year, . events per infusion) and patients experienced a causally related non-serious systemic ae ( . %, . events/patient year, . events per infusion). seven of patients who were tested for anti-rhuph antibody had positive binding antibody test to rhuph (titer : ; maximum titer : at enrollment, : during the study); no neutralizing rhuph antibodies were detected. of the patients who responded to the treatment preference questionnaire at the end of year , the majority ( / [ . %]) preferred to receive their ig therapy at home; . % ( / ) preferred the doctors office; patients preferred treatment at the hospital, had no preference, or indicated other. almost all patients ( / [ . %]) indicated a preference to continue treatment with ighy. conclusion: this interim analysis of patients with pidd treated with ighy in routine clinical practice supports previous observations that ighy is a well-tolerated and preferred therapy with no reports of treatment-related saes or neutralizing anti-rhuph antibodies. background: cartilage hair hypoplasia (chh) is an autosomal recessive chondrodysplasia associated with variable immunodeficiency. pathogenic defects in rmrp, encoding the untranslated rna subunit of ribonucleoprotein endoribonuclease complex (rmrp), result in reduced mrna and rrna cleavage. rmrp c. a>g is the most common variant, increased in finnish and amish populations. while cellular immunodeficiency is associated with increased morbidity and mortality, there is no established correlation between clinical and immunological phenotype. lymphocyte radiosensitivity has not been described. case: a full-term amish female infant had low trec copies on newborn scid screen. flow cytometry at months-old demonstrated severe t and b cell lymphopenia (cd +t-cells cells/mcl, range: , - , cells/mcl; cd +b-cells cells/mcl, range: - , cells/mcl) with normal nk quantitation (cd / + cells/mcl, range: - , cells/mcl) and cd + memory t-cell expansion ( . %) relative to the naïve subset ( . %). t-cell functional mitogen responses were normal. she was diagnosed with chh with homozygous rmrp c. a>g mutation. lymphocyte subset (t, b and nk cells) radiosensitivity was evaluated by flow cytometric analysis of phosphorylated (p) atm, smc and gamma-h ax after low-dose ( gy) irradiation. an increase in gamma-h ax level was observed in a subset of non-irradiated t cells ( . % v. . % gamma-h ax+) and nk cells ( . % v. . % gamma-h ax+) in the patient, suggestive of a constitutive defect in dna repair. the relative distribution of t, b and nk cells expressing patm, psmc and gamma-h ax at hour postirradiation (ir) was not significantly different from the experimental healthy control (ehc) or pediatric reference range (prr). however, the kinetics of dephosphorylation at hours post-ir was altered with residual gamma-h ax expression in a subset of the patients t cells (delta . %, mode ratio mean fluorescence intensity (mfi)= . ; ehc: delta . %, mode ratio mfi= . ; prr: delta . %, mode ratio mfi= . ). a similar finding was observed in a subset of patient b-cells for gamma-h ax (delta . %, mode ratio mfi= . ; ehc: delta . %, mode ratio mfi= . ; prr: delta . %, mode ratio mfi= . ). the frequency of the patient's lymphocytes with residual gamma-h ax persistence at h post-ir was prominent, with . % t-cells demonstrating persistence of gamma-h ax (compared to . % in the ehc, and . % in the prr), and . % b-cells gamma-h ax+ (compared to . % in the ehc, and . % in the prr). there has been lack of follow-up, but verbal report suggests no significant immunological or infectious concerns at year of age. discussion: lymphocyte radiosensitivity is a novel finding in chh with t and b cell lymphopenia. the ability of rmrp to associate with telomerase reverse transcriptase (tert) and function as an rna-dependent rna polymerase, yielding distinct silencing rna sequences, may underlie radiosensitivity in rmrp mutants. systematic characterization of lymphocyte radiosensitivity and immunological phenotype could provide useful information on whether this could serve as a biomarker for the magnitude or complexity of immunodeficiency. assessment of radiosensitivity has implications in conditioning regimen selection for patients requiring allogeneic hematopoietic cell transplantation. we recommend lymphocyte radiosensitivity assessment in chh infants identified by nbs scid and chh patients with significant immunodeficiency and/or malignancy. novel primary immunodeficiency with lymphoproliferative disease due to biallelic defects in nckap l background: three children from non-consanguineous families and different ethnic backgrounds developed lymphoproliferative disease by years of age. they also had recurrent infections, including pneumonia and bronchiectasis, otitis media, and skin pustules. immune phenotyping revealed low cd + t cell percentages, an accumulation of memory-like cd + t cells, impaired t cell proliferation, and low total nk cell numbers. methods: the affected individuals, unaffected parents, and other unaffected family members underwent exome sequencing. results: all affected cases had rare and bioinformatically damaging biallelic variants, with appropriate familial segregation, in nckap l, which encodes hem . hem is an essential component of the wave regulatory complex (wrc). immunoblotting confirmed destabilization of the wrc in all patients. immunofluorescence microscopy demonstrated defective f-actin and wave localization to immune synapses in nk cells. significant abnormalities were identified in patient lymphocyte and neutrophil migration and morphology, consistent with altered wrc-mediated cytoskeletal dynamics. all patients exhibited impaired inside-out integrin activation. knockdown of hem produced deficient proliferative responses and mtorc -mediated akt activation in control t cells. conclusions: the immunologic and clinical phenotype in the affected individuals recapitulates the phenotype observed in hem -deficient mice. biallelic defects in nckap l therefore result in a novel human primary immunodeficiency disease characterized by lymphoproliferation and susceptibility to infections. background: concurrent existence/significance of immunodeficiency with new onset lymphoproliferative disease remains understudied. just two studies to date have evaluated the prevalence of hypogammaglobulinemia in chronic lymphocytic leukemia (cll) and neither studied prevalence and impact of ige deficiency on outcomes in cll [ , ] . therefore, the objective of this study was to examine the prevalence of hypogammaglobulinemia, examining all isotypes, in newly diagnosed cll patients and to test the hypothesis that patients with hypogammaglobulinemia have a distinct clinical profile and outcome. methods: using the banked sera of newly diagnosed, treatmentnaïve, cll adult patients from the lymphoma molecular epidemiology resource (l-mer), ig (igg, iga, igm and ige) levels were measured. the l-mer was initiated as an observational cohort study of prospectively enrolled newly diagnosed lymphoma patients evaluated at the mayo clinic (rochester, mn) and the university of iowa (iowa city, ia) [ ] . igg/a/m levels were measured using immunoturbidimetric assay whereas the ige level was determined using electrochemiluminescence immunoassay. the associations between ig deficiencies and clinical factors were evaluated with wilcoxon rank sum and chi-squared (fishers exact, where appropriate) tests. cox regression models were used to assess the effects of clinical variables on overall survival (os). time was calculated from biopsy to death due to any cause; patients still alive were censored at last contact. all tests were two-sided and assessed for significance at the % level using sas v . (sas institute, cary, nc). results: the mean age (sd) of the selected cll cohort was . ( . ) years with a male predominance ( . %). . % of the patients were white. with a median follow-up of five years, there were deaths. hypogammaglobulinemia in newly diagnosed, treatmentnaïve cll was common in our cohort with ( . %) patients having a measurable isotype deficiency. the most common ig deficiency was igm ( . %, % ci . - . %), followed by igg ( . %, % ci . - . %), ige ( . %, % ci . - . %) and iga ( . %, % ci . - . %). multiple deficiencies in the same patient were common ( figure ). iga and ige deficiency were associated with higher rai stages (grading system for cll) at presentation (p< . and . respectively) as well as with higher white blood cell counts at presentation (p= . and . respectively). a higher proportion of iga deficient patients needed second treatment during follow-up ( % compared to %, p= . ). when comparing predictors of overall survival, higher rai stage [ - vs , hazard ratio (hr) . , % ci . - . , p= . ] and age (hr . , % ci . - . , p< . ) correlated with worse overall survival. individual immunoglobulin deficiencies did not correlate with overall survival. conclusions: a significant proportion of treatment-naïve patients with cll have underlying ig deficiencies-both in isolation and a combination of different isotypes. a deficiency of iga or ige was associated with severe disease at presentation. the underlying relationship between these two immunologic disorders deserves further study. background: patients with primary immunodeficiency (pid) have an increased risk of developing autoimmune diseases, including rheumatoid arthritis (ra). management of these patients is challenging as immunomodulators can further increase their risk for infections. additionally, patients with ra that undergo therapy with drug modifying antirheumatic drugs (dmards) may develop a secondary immunodeficiency. there are few studies reviewing the characteristics of patients with a pid who later develop ra, and no studies have been reported comparing these patients to those who develop an immunodeficiency after starting dmard therapy for ra. methods: patients were identified as having inflammatory arthritis and a concomitant immunodeficiency (id) at our institution between / / - / / using icd- and codes. manual chart review was performed to confirm and identify the timing of diagnosis of these disorders. patients were excluded if either there was no definitive diagnosis of id or ra (clinically diagnosed by a practicing allergist/immunologist and meeting acr criteria for ra with a score of or higher, respectively), or rituximab was administered prior to diagnosis of id . clinical symptoms, treatment, and laboratory data were extracted. fishers exact test was used to compare the categorical variables between the groups; ttest was used to compare the continuous variables. results: patients met the inclusion criteria. patients were diagnosed with an id and developed ra later in life (group ), and patients were diagnosed with ra and subsequently developed a clinically significant id (group ). the mean ages of diagnosis of id and ra in group patients were . years (sd ± . ) and . years (sd ± . ), respectively. in group , the mean age of diagnosis of ra was . (sd ± . ), compared to . years (sd ± . ) for the diagnosis of id. most patients in both groups were female ( % in group and % in group ). all patients in both groups had a humoral id, including common variable immunodeficiency (cvid) ( % of group patients), specific antibody deficiency (sad) ( % of group and % of group patients), and hypogammaglobulinemia ( % of group and % of group patients). all patients in group were seropositive for rheumatoid factor (rf) or anti-cyclic citrullinated peptide (anti-ccp), whereas only % of patients in group were positive for rf or anti-ccp (table ). most patients in both groups were treated with immunoglobulin replacement therapy. treatment of ra in both groups was similar, but combination dmard therapy was not used in group patients in contrast to group patients. conclusions: our study indicates that even though clinical characteristics and management are similar in patients with coexisting id and ra, rf and anti-ccp are usually negative in patients who develop ra after id, possibly due to impaired antibody production in immunodeficient patients. assistant professor of allergy and immunology, arkansas children's hospital, university of arkansas medical sciences introduction/background: complement deficiencies are relatively rare, comprising less than % of primary immunodeficiencies. they are associated with increased risk for infections with encapsulated organisms and autoimmunity. of all complement deficiencies, the rarest are defects in the alternative complement pathway. properdin deficiency is the most commonly described alternative pathway deficiency, with factor b and factor d deficiency more rarely described. fewer than patients with factor d deficiency have been reported with all reported cases being children of consanguineous parents who succumbed to meningococcal sepsis. objectives: to describe a case of factor d deficiency associated with recurrent respiratory infections with streptococcus pneumoniae pneumonia with associated lung abscess and empyema. methods: retrospective chart review was conducted. laboratory investigations included lymphocyte immunophenotyping by flow cytometry, lymphocyte proliferation to mitogen, quantitative serum immunoglobulins, vaccine titers, complement assays and functional evaluation, and genetic evaluation by next generation sequencing. results: a year old marshallese male was transferred from an outside hospital to our facility for further evaluation of worsening pneumonia and was found to have right-sided pleural effusion and pulmonary abscess in the right lower lobe. the abscess was drained and was found to be positive for streptococcus pneumoniae via polymerase chain reaction. he improved after chest tube placement and treatment with intravenous antibiotics. his medical history was significant for recurrent acute otitis media and prior hospitalization out-of-state for pneumonia with empyema secondary to streptococcus pneumoniae, which required chest tube placement and admission to the pediatric intensive care unit at months of age. immunologic work up revealed age-appropriate lymphocyte subpopulations, lymphocyte proliferative responses to mitogens, quantitative immunoglobulin levels, pneumococcal/tetanus/diphtheria titers, and ch complement assay. ah complement assay was decreased to units/ml. complement testing was repeated -with normal ch and ah of units/ml. further evaluation revealed normal levels of factors b, h, i and properdin. factor d level was . mcg/ml, and factor d function was decreased to units/ml, indicating a diagnosis of factor d deficiency. sequencing of the cfd gene revealed a previously undescribed homozygous deletion (c. _ del and p.lys del). the parents were not agreeable to personally undergoing genetic evaluation to determine if this was a de novo mutation. the patient was managed with pneumococcal and meningococcal immunizations, prophylactic amoxicillin and intravenous gamma globulin (ivig) without any further infections. unfortunately, after two ivig infusions, he was lost to follow up. conclusion: factor d deficiency is an extremely rare alternative complement pathway deficiency, described in less than patients. all infections described thus far have been secondary to neisseria meningitidis. this case represents not only a novel mutation in the cfd gene leading to factor d deficiency, but also the first description of a patient with factor d deficiency developing invasive infection secondary to streptococcus pneumoniae. background: viral infections are a significant cause of morbidity and mortality in patients with primary immunodeficiency disorders and following hematopoietic stem cell transplantation. adoptive immunotherapy using virus specific t-cells (vsts) has been shown to prevent and treat viral infections in immunocompromised hosts. human parainfluenza virus- (hpiv ) is a common cause of severe respiratory illness in immunocompromised patients and has no approved antiviral therapies and has not previously been used as a target for t cell therapeutics. introduction: we previously reported that fatigue is increased in common variable immunodeficiency (cvid). however, in previous studies, fatigue was not defined using validated tools. our aim from this study is to identify the prevalence of patient-reported fatigue, using validated questionnaires, and determine the factors predisposing to fatigue in cvid methods: data from cvid who responded to the idf patient national survey a were analyzed. fatigue was measured using the brief fatigue inventory (bfi) questionnaire, which includes seven items to identify fatigue, and measure fatigue severity. a total of patients with cvid and responses to bfi were enrolled. demographics, co-morbidities, immunoglobulin replacement therapy (iggrt) route and dose, co-morbidities, infections, depression, quality of life (qol) (using the sf- v ) and disability were compared between fatigued and non-fatigued. logistic regression was used to identify the significant variables. ebv reactivation without ptld, treated with rituximab. alive and well. j clin immunol ( ) (suppl ):s -s s granulomas are the most significant day-to-day problem for cvid patient management. currently, there are limited options for their treatment and the optimal therapy is unknown. in case reports and small series, infliximab has been reported effective while others found it useless. we here describe a yo white male referred for monthly ivig in august . at age , he developed large areas of erythematous polymorphic plaques in his cheeks, arms and legs. a skin biopsy showed tuberculoid granulomas negative for bacteria, baar and fungi, with infiltrating cd + lymphocytes. a prolonged course of steroids did not improve his skin. he also had multiple pneumonias and bronchiectasis, and oral candidiasis. he received all vaccines, including bcg with no complications. with low immunoglobulins and a poor response to pneumococcal polysaccharides and tetanus toxoid he was diagnosed as cvid and placed on ivig at yo with excellent infectious control since then. at age , his skin lesions persisted and deepened to the bone on his left leg. broad spectrum antibiotics for months were unsuccessful. at yo to yo, skin grafts were performed on his arms, legs and both cheeks. two ulcers persisted on his left leg until august that increased in size, deepened and became erythematous and extremely painful (fig. ) . in september, two new ulcers appeared on his right cheek and right gluteus, respectively. one week later a third ulcer was found on his left calf. on september th, infliximab mg/kg ( mg) was administered. on the second infliximab dose, october th, the pain was completely gone and all ulcers were shrinking, and those ones in the cheek, gluteus and calf almost completely resolved. by the third dose, on november rd the ulcers in his right leg were almost closed (fig. ) . infliximab mg treatment continues every weeks. lab test remained unchanged from till , when his wounds got worsened. (table ) granulomatous disease in cvid is a challenge. both b and t cell directed therapies are encouraged. we add a new case of an infliximab responsive patient to others already reported. over genes have been reported to cause monogenic cvid. a year old girl presented with recurrent pneumonias and a diagnosis of cvid. the parents sought a second opinion. born at weeks gestational age, she was "always smaller and sicker than her friends," and in the prior months she had episodes of pneumonia with fever to f requiring emergency department treatment. two of these were associated with rsv and metapneumovirus, respectively. laboratory evaluation confirmed low levels of igg ( mg/dl) iga ( ) and igm ( ) congenital tuberculosis (ctb) is a rare disease most often associated with maternal genitourinary (gu) tuberculosis (tb) or disseminated tb. due to infertility caused by gu tb, ctb is rarely reported even in endemic countries. infants can acquire tb hematogenously via the placenta or umbilical vein or by fetal aspiration of infected amniotic fluid. presenting symptoms include respiratory distress, fever, hepatosplenomegaly, poor feeding, lethargy, and low birth weight. we report a premature female infant conceived via in vitro fertilization (ivf), who was born to indian immigrant parents at weeks of gestation due to preterm premature rupture of membranes. maternal history was significant for pulmonary tb at years of age. she denied abdominal or gu symptoms. infants nicu course was complicated by opacifications in the right lung and leukocytosis with neutrophil predominance, identified during evaluation of frequent apnea and bradycardia episodes at month of age. clinical improvement was noted after treatment with vancomycin, amikacin and piperacillin-tazobactam; however, leukocytosis of unknown etiology persisted. at . months of age she was discharged to inpatient rehabilitation. at months of age, she was readmitted for fever and respiratory distress. during this admission, an immune evaluation was undertaken due to persistence of symptoms along with unresolved leukocytosis with a peak of , cells/l with neutrophilia to , cells/l, and chest ct evidence of progressive multifocal lung disease worse in the right upper lobe despite empiric treatment with broadspectrum antibiotics. infectious work-up was negative, including acid-fast bacilli testing from bronchoalveolar lavage. due to the pronounced and persistent leukocytosis and neutrophilia, a primary immune defect was suspected. immune evaluation included: normal immunoglobulins (ig) g, a, and e, elevated igm, vaccine-specific antibody titers protective to diphtheria and of streptococcus pneumonia strains, mildly elevated t and b cells, a normal flow cytometry for dihydrorhodamine, myeloperoxidase stain and glucose- -phosphate dehydrogenase level, as well as a peripheral smear with no giant azurophilic granules. her primary immunodeficiency genetic panel was unrevealing. she underwent lung biopsy via video-assisted thoracoscopic surgery, which showed noncaseating granulomas and eventual growth of multi-drug-resistant mycobacterium tuberculosis (mtb). upon treatment with an appropriately adjusted anti-tuberculosis regimen, she showed rapid clinical and laboratory improvement. endometrial samples obtained from mother showed gu tb, confirming the diagnosis of ctb. the slow-growing nature of mtb that resulted in delayed diagnosis, along with the presence of non-caseating granulomas and persistent neutrophilia, prompted an immune work up that was completely normal. this case demonstrates the importance of considering ctb in the differential diagnosis of an infant presenting with severe lung infection, persistent neutrophilia, suboptimal response to broad-spectrum antibiotics and relevant epidemiologic risk factors. furthermore, in the setting of appropriate parental exposures and infertility prompting the use of ivf, maintaining a high level of suspicion of ctb can aid in earlier diagnosis of affected neonates. -year-old caucasian male who initially presented with recurrent otitis media, persistent hsm, lad, and hypogammaglobinemia (igg < mg/dl) at years of age. he was diagnosed with common variable immunodeficiency (cvid) and chronic arthritis when he was and years of age, respectively. subsequently, he developed hepatitis and recurrent pneumonia with mycobacterium avium complex (mac). his arthritis partially responded to anti-tumor necrosis factor (tnf) agents and tofacitinib, but did not respond to anti-interleukin- treatment. a combination of anti-tnf inhibitor, tofacitinib, and low dose prednisone was required to control his arthritis. hypogammaglobulinemia (igg < mg/dl), recurrent otitis media, pneumonia, crohn's disease, celiac disease, lad and failure to thrive at years of age with more recent development of hsm. he required only immunoglobulin replacement therapy. case# is a -year-old caucasian male, the half-brother of case# , who initially presented with recurrent pleural effusion and bilateral pulmonary infiltrates, hsm, lad, abdominal distension and ascites at years of age. a transbronchial lung biopsy revealed chronic eosinophilic pneumonitis. liver biopsy showed increased eosinophils in the sinusoids with diffuse enlargement of hepatocytes, but without hepatitis. colon biopsy revealed minimal colonic eo-sinophilia. his pulmonary infiltrates and pleural effusion responded to prednisone, and he has not required additional treatment for past . years. conclusions: the clinical manifestations of the same genetic variant may be variable and unpredictable even in the same family. stat gof syndrome should be considered in children with multisystem autoimmune diseases, lad, hsm and low switched memory b cells regardless of presence of hypogammaglobulinemia or history of recurrent infections. background: patients with primary immune deficiencies characterized by severe t lymphopenia and/or poor t cell function and patients posthematopoietic cell transplantation are at high risk of severe viral infections. antiviral medications are expensive, not always effective and associated with significant toxicity and/or long-term side effects. as such, there has been increasing interest in the use of donor-derived or thirdparty virus-specific t cells (vsts), and several studies have demonstrated efficacy of vsts generated using various manufacture strategies. however, in depth immunologic and metabolic characterization of vsts has not been reported, limiting correlative investigations into efficacy. methods: ebv-vsts were generated from apheresis t cells collected from healthy donors using three methods: ( ) stimulation and expansion with hla-matched ebv-lymphoblastoid cell lines (lcls) purchased from astarte biologics or sigma-aldrich over a period of weeks, ( ) stimulation with ebv peptivator from miltenyi followed by expansion over - days with different cytokines, and ( ) stimulation with ebv peptivator followed by isolation of activated cells using the ifn-gamma capture system from miltenyi. immunophenotyping by flow cytometry was performed using the miltneyi macsquant analyzer. the nanostring ncounter system was used to measure gene expression for metabolic pathway analysis, and the agilent seahorse xf cell mito stress test system was used to measure mitochondrial respiration. results: ebv-vsts generated using lcls or peptivator plus il- both resulted in a high percentage of cd t cells skewed to the effector memory and terminal effector memory phenotype with high expression of the exhaustion markers pd- , tim- , and lag- . conversely, ebv-vsts generated using peptivator plus il- and il- and the ifn-gamma capture system resulted in a mixed cd and cd t cell population with a high number of central memory t cells and lower percentage of cells positive for pd- , tim- , and lag- . stimulation with peptivator followed by expansion with il- resulted in an intermediate immunophenotype. nanostring results demonstrated upregulation of the glycolytic pathway in ebv-vsts stimulated with peptivator followed by expansion with il- or il- compared to ebv-vsts generated using the other manufacture approaches. the seahorse mito stress test demonstrated that the peptivator plus il- ebv-vsts had a significantly lower spare respiratory capacity than other ebv-vsts and a low extracellular acidification rate despite upregulation of the glycolytic pathway. the peptivator plus il- and il- ebv-vsts had the highest basal oxygen consumption rate, atp-linked respiration, and extracellular acidification rate. conclusions: manufacture of ebv-vsts using the various approaches currently employed clinically results in t cell pools with different immunophenotypes and different metabolic profiles. ebv-vsts stimulated with peptivator followed by expansion in il- and il- and ebv-vsts isolated using the ifn-gamma capture system have immunophenotypes and metabolic phenotypes suggestive of potential greater in vivo persistence, whereas ebv-vsts expanded in il- and il- have characteristics correlated with increased effector function. however, these vsts are more likely to be short-lived and to have impaired metabolic fitness. these phenotypes will enable better correlation with clinical results and suggest combinatorial approaches depending on clinical indication. introduction: majority of patients with primary immunodeficiencies (pid) require life-long replacement therapy with immunoglobulins (ig) to prevent severe infections and irreversible complications. in addition to safety and efficacy, tolerability and convenience of administration of ig products are essential factors for patients. a new . % ig preparation octanorm (octapharma, lachen; tradename cutaquig® in north america) has been developed for subcutaneous administration (scig) derived from the established manufacturing process of octapharmas intravenous ig (ivig) brand octagam®. objectives: primary outcome was assessment of the efficacy of octanorm in preventing serious bacterial infections. main secondary endpoints included (among others) evaluation of tolerability and safety of octanorm, the number and rate of other infections, number of days missed at work, and use of antibiotics. methods: a prospective, open-label, non-controlled, single-arm phase study involving adult patients with pid was conducted at russian centers. patients treated with at least infusions of ivig prior to enrollment and with igg trough levels . g/l underwent an -week wash-in/wash-out period followed by a week efficacy period. during the study, patients received weekly administrations of octanorm at the same monthly dose as during previous ivig treatment (monthly ivig dose divided by for weekly dose). in total, each patient received scig infusions. results: twenty-four patients completed the study. one patient terminated early (after infusion , during wash-in/wash-out phase; personal reasons). mean age was . years (range - years). fifteen patients ( %) were female and patients ( %) male. no serious bacterial infections were recorded. during the efficacy period a total of non-serious infections was observed in patients. seventeen infections in patients were of mild and infections in patients of moderate intensity. the infection rate per person-year was . . in total patients received infusions of study drug. the average dose of cutaquig® was . g/kg/week. during the entire study, systemic adverse events were reported (including infections). three of these systemic adverse events were rated as related to study drug, all were non-serious. there was no serious or significant adverse event nor was there an adverse event leading to withdrawal. infusion site reactions were reported for % of infusions. serum igg trough levels were nearly constant during the efficacy period. median igg trough levels were . g/l at screening, . g/l at the end of wash-in/wash-out period and . g/l at the termination visit. one patient had a trough level g/l at visits during the efficacy period and the dosing was subsequently adjusted for this patient. during the primary treatment period patients ( . %) used antibiotics in treatment episodes (total of treatment days; range - days) and patients had absences from work or school due to infections (total of days of absence). conclusion: this study demonstrated that the new subcutaneous human normal immunoglobulin . % is well tolerated, safe and effective in adult patients with pid. background: children with chronic granulomatous disease (cgd) are at high risk for fungal infections (especially with aspergillus species) and these infections usually have contiguous site involvement. most patients have pulmonary presentation. infective endocarditis and fungal osteomyelitis of skull are distinctly unusual. we report one such case. case: a -year-old boy, born out of a non-consanguineous marriage, presented with soft tissue swellings of skull for months. his past history was significant with an episode of pneumonia at year and recurrent soft tissue swellings all over the body since ½ years of age. on examination he was wasted, had signs of micronutrient deficiency, rickets, pallor, cervical lymphadenopathy and two abscesses, x cm on right temporo-parietal region and x cm over left frontal region. he was also found to have hyperdynamic precordium with an ejection systolic murmur. investigations revealed hemoglobin g/l; platelet count . x /l; total leukocyte count x /l(n /l /m /e ); elevated c-reactive protein( mg/l) and a raised erythrocyte sedimentation rate( mm sthr). chest x ray revealed cardiomegaly (cardiothoracic ratio %) and d echocardiography showed vegetation of x mm over the anterior mitral leaflet suggestive of infective endocarditis. blood and urine cultures were sterile. culture from pus over the temporo-parietal abscess showed growth of aspergillus fumigatus. human immunodeficiency virus serology was non-reactive. immunoglobulin profile revealed elevated igg . g/l ( . - . g/l) and iga . g/l( . - . g/l); igm was . g/l( . - . g/l). in view of strong suspicion of cgd, nitroblue tetrazolium dye reduction test (nbt) was carried out-it revealed no reduction and dihydrorhodamine (dhr) assay showed a low stimulation index ( . ). flow cytometry for gp phox and gp phox was normal and dhr of mother did not reveal x linked carrier state. contrast enhanced computerized tomography (cect) of head showed osteomyelitis of the calvarial bones. contrast enhanced magnetic resonance imaging (cemri) brain showed heterogeneously enchancing soft tissue lesion in the scalp at right fronto-parietal region and left frontal region with underlying bony destruction suggestive of osteomyelitis. he was given intravenous antimicrobials (ceftriaxone, gentamycin, cloxacillin, voriconazole). after weeks of therapy, he showed resolution of findings on mri brain and a repeat d echocardiography showed significant decrease in size of mitral leaflet vegetation. conclusion: this case highlights a rare presentation of cgd with infective endocarditis and skull osteomyelitis due to aspergillus fumigatus. to the best of our knowledge, this has not been reported previously. background: genetic defect in il r affect cellular immunity, underlie mendelian susceptibility to mycobacterial disease (msmd) and inflammatory bowel disease (ibd) through different pathways. we present for the first time a patient with il- r deficiency from a consanguine family with two different phenotypes. initially diagnosed as crohn's disease prior to the msmd diagnosis. method and material:patient was referred to the clinical immunology and allergy clinic at the at alzahra university hospital for immunological and genetic evaluation . blood samples from patient, his family and healthy donor controls were collected upon informed consent. in this study, we investigated effect of il r mutation in il- /ifnaxis by evaluation of patients whole blood cell response to il- and ifn-, il- r expression in pbmcs and t cell blasts. also wholeexome sequencing has been performed. result and discussion: a years old male from consanguine family , with history of right sub-axillary bcg lymphadenitis, recurrent mouth ulcers , chronic diarrhea in childhood and appendectomy at age of was investigated. based on his clinical presentation abdominal pain, significant weight loss, chronic and bloody diarrhea , endoscopic and pathological findings treatment for crohn's disease (cd) was started at the age of seven . unfortunately, protracted patient's symptoms ends up to resection of his colon and colostomy two years later. he was presented with multi focal osteomyelitis at the age of . although no bacteria was detected in pcr and tissue culture of the bone biopsy and the patient was not responded to antibacterials , he had a dramatic response to empirical anti mycobacterial treatment and his severe bone pain and lesions were healed. even though the bone manifestations were completely controlled, he continuously was under treatment for his gastrointestinal symptoms. genetic analysis was confirmed segregation of homozygous mutation in splice site of exon in il- r . expression of gene was completely abolished in pbmcs of patient and the surface expression of il rb was not detectable in t cell derived pbmcs of the patient compared to healthy control. furthermore, did not response to il stimulation since we could not detect increase of inf-after stimulation with il and bcg. our patient received bcg vaccination at birth and had bcg lymphadenitis as an infant, cd and mycobacterial multifocal osteomyelitis as a child. furthermore there are some evidences which indicate the role of atypical mycobacterial infections as a trigger for cd. conclusion: we reported for the first time contemporary msmd and ibd in years old patient, who had impaired il- signaling and abolished il r expression in pbmcs and t cell blast. however, mycobacterial osteomyelitis is a typical phenotype of msmd patients with deficiency in ifn-r or stat, there were no mycobacterial osteomyelitis reported in il- r deficient patients. background: advanced genetic studies help explain the occurrence of many undiagnosed, rare conditions. recently, nbas variants were identified as a causative basis of recurrent liver failure in infants (infantile liver failure syndrome , ilfs ). the nbas (neuroblastoma amplified sequence) gene encodes a protein involved in golgi to er retrograde transport. nbas functions seem to be broad and loss of function variants in nbas have been associated with multisystem manifestations. case report: a y m old chilean male presented to the er with a three day history of vomiting, diarrhea and one day of fever ( . °f). on examination he was pale, lethargic, and tachycardic. a chemistry profile revealed markedly elevated liver enzymes, increased bilirubin, and coagulopathy, consistent with the acute hepatic failure (alt , ast > , total bilirubin . ( . db), ggt , and inr of . ). he was hospitalized, given vitamin k, and kept on intravenous fluids, ursodiol, and antipyretics. his liver function improved significantly within days of admission (alt was down to , ast , total bilirubin . ). work-up of possible etiologies including autoimmunity and infectious hepatitis was negative. liver sonogram was normal, but liver biopsy was consistent with acute hepatitis with some necrosis. urine organic acid and plasma amino acid screens were not consistent with any inherited metabolic disorders. his parents recalled two previous episodes of liver failure at ages and years. both were preceded with a mild febrile illness and non-specific symptoms including fever, coughing, vomiting, diarrhea, lethargy, and decreased po intake. these subsequently were followed by jaundice and marked elevation of liver enzymes. flu a and adenovirus were identified as causes of febrile illnesses of the two previous episodes. for this admission, adenovirus was found in the respiratory secretions and a mild ebv viremia was also detected. genetic evaluation in chile was reportedly normal. after a literature review we obtained sequencing of nbas which revealed two variants: c. g>t,p.glu * and nbas c. t>g, p.iie ser. both variants have been previously reported in patients with an infantile onset, recurrent liver failure syndrome. his other clinical features include developmental and speech delays, failure to thrive, and facial dysmorphism. he also has a history of recurrent ear infections and has had sets of tympanostomy tubes. further testing was limited due to the lack of insurance coverage. conclusion: nbas deficiency is a newly described syndrome of recurrent acute liver failure that occurs early in life. once individuals have survived to adulthood they do not seem to develop liver failure with illness. typically, liver crisis is triggered by a common childhood febrile illness. the mechanism of disease is thought to be thermal instability of hepatocytes which improves over time in most cases. however, although spontaneous recovery can occur following the crises, each episode can be fatal or result in permanent liver damage required liver transplantation. increased awareness of this disease will lead to the early establishment of the diagnosis. appropriate and timely management of fever at the onset of illness can significantly improve outcome in this potentially fatal disease. associate prof., infectious diseases and tropical medicine research center, isfahan university of medical sciences, isfahan, iran background: pre-eclampsia, a pregnancy-specific complication, has been shown to be associated with cytomegalovirus (cmv) infection. cmv specific t-cell response plays the major role in cmv infection or disease .we explored whether a change in cmv-specific cell-mediated immunity (cmi) is related to the development of preeclampsia. method: cmv-specific cmi was assessed using cmv-quantiferon (qf-cmv) assay in serum from women with pre-eclampsia as well as normal pregnancy controls retrospectively. participants were matched for gestational age individually. proportion of reactive results, mean value of interferon-level produced in mitogen and antigen tubes were compared between the cases and controls via chi-square, wilcoxon rank-sum tests, respectively. odds ratio (or) and confidence interval (ci) were calculated as well. result: no significant differences observed between demographic characteristics of the case and control groups. the qf-cmv assay turned reactive (qf-cmv [+]) in of of patients ( %) vs. of controls ( . %) (p = . ). women with pre-eclampsia had lower mean ifn-levels in antigen tube ( . ± . ) compared with normal pregnancy controls ( . ± . ) (p = . ). there was no statistically significant differences in this value of mitogen tube between cases ( . ± . ) and controls ( . ± . ) (p = . ). women with suppressed cmv-cmi were . times more likely to manifest pre-eclampsia (or= . , % ci: . - . ). this result even strengthened after adjustment for age, gestational age and gravidity (or = . , % ci: . - . ). conclusion: our finding support an association between suppressed cmv specific cmi and pre-eclampsia. introduction: the triad of susceptibility to infections, auto-inflammation, and cancer in a patients personal and family history are always suggestive of an underlying primary immunodeficiency; however, in some cases the diagnosis might be delayed for years. furthermore, the results of immunological and inflammatory evaluation can also be affected by ongoing immunomodulatory therapy initiated by different specialists upon clinical diagnosis. objective: to describe a unique presentation of auto-inflammatory disease with combined immunodeficiency in an adult patient. case presentation: we report here the case of a year old male, who had a long history of infections including recurrent sino-pulmonary bacterial infections starting during childhood, osteomyelitis at years of age, recurrent tonsillitis requiring tonsillectomy at years of age, recurrent cellulitis, an episode of prostatitis with septicaemia, as well as recurrent varicella zoster and warts. the patient was also diagnosed with sclerosing mesentheritis, and reynauds phenomenon, recurrent oral ulcers, arthritis, uveitis, autoimmune thyroiditis, lung fibrosis and suffered repeated episodes of abdominal pain. furthermore, there is a family history of early childhood death, multiple soft tissue cancers, crohns disease, and autoimmune thyroiditis. upon physical examination, the patient had multiple telangiectasia, baseline erythroderma, and flushing. immunological evaluation showed lymphopenia with significant reduction in both circulating b and t cells, however, assessment of humoral immunity revealed low igg and decreased igm with normal iga levels. at the time of the evaluation he had been on low dose daily prednisone ( . mg), colchicine, and methotrexate as immuno-modifying therapy. genetic evaluation revealed a heterozygous mutation in nod as well as compound heterozygous mutations in the mefv gene. discussion: mutations in nod have been described in association with blau syndrome a multisystem auto-inflammatory syndrome which may explain many of the features experienced by our patient. to our surprise next generation sequencing revealed a second aberration in the mefv gene which causes familiar mediterranean fever, another multisystem auto-inflammatory disease, which might lead to the phenotype observed in the patient. conclusion: this is the first report of genetic lesions in two different genes leading to a severe course of auto inflammation. monogenic autoinflammatory syndromes (mais) are a diverse group of disorders characterized by primary over-activation of the innate immune system. induction of the inflammasome complex by innate immune sensors and increased production of il- b are implicated in the pathogenesis of mais. macrophage activation syndrome (mas) is a life-threatening illness defined by acute hyper-inflammation and unopposed cytokine release. it is considered an acquired condition secondary to infection, rheumatoid disease or malignancy. the early therapeutic use of il- b inhibition has profoundly improved the prognosis mas. it has recently been shown that increased free il- levels in the blood are causatively linked to the development of mas. significant overlap in clinical presentation and laboratory markers between patients with mais and mas led us to explore the role of free il- and therapeutic use of il- b inhibition in a patient with cdc mutation. here, we report the case of an months-old female who presented with hydrops fetalis in utero, and later developed failure-to-thrive, splenomegaly, anemia, thrombocytopenia, arthralgias, rashes, frequent febrile episodes and mild facial dysmorphism along with massive increase in crp, esr and ferritin. whole exome sequencing (wes) identified a heterogenous likely pathogenic de novo variant in cell division control protein homolog (cdc ) c. g>a (p.c y). cdc encodes a small rho family gtpase that regulates multiple signaling pathways controlling cell polarity, migration, endocytosis and cell cycle progression. single allele mutations in the cdc gene were recently reported to cause takenouchi-kosaki syndrome manifesting with growth retardation, developmental delay, facial dysmorphism, and thrombocytopenia however systemic autoinflammation has not been described. cdc closely interacts with the wiskott-aldrich syndrome protein but little is known about the mechanism underlying immune abnormalities associated with cdc mutations. our patient had an inflammamosopathy-like syndrome. because of significant clinical overlap to mas, we measured il- , il- , free il- and il- binding protein, all of which were significantly increased. this increase in free il- heightened her risk of developing mas. her il b level was normal, but an increase in il- b is hardly ever detectable in the serum despite playing a critical role in this type of inflammation. indeed, chronic il- b excess in the tissues promotes systemic inflammation and is associated with chronically elevated crp and esr. with this rationale we started the il- receptor antagonist anakinra. within hours from starting anakinra, the parents observed an increase in appetite, resolution of arthralgias and improved mobility. over the course of the following weeks, fever, anemia, thrombocytopenia and rash disappeared, the spleen massively decreased in size and the patient started to meet developmental milestones. crp, esr eventually normalized while ferritin and free il- are still trending down. conclusions: significant increase in free il- and extremely encouraging clinical response to therapy with anakinra in a patient with novel cdc mutation suggests a link between mas and defects in cdc . elucidating the mechanism of inflammasome activation and the drivers of il- increase in mas and mais more broadly may shed light on novel therapeutic targets like the use of human recombinant il- binding protein. j clin immunol ( ) (suppl ):s -s s maintenance; smarcal is enriched in cells that maintain telomeres via the alternative lengthening of telomeres pathway and smarcal decifient cells demonstrate telomere instability with replication fork collapse and increased telomere-associated dna damage. [ , ] telomere analysis of siod patients, including one patient who received a hematopoietic stem cell transplant (hsct) years prior, as well as heterozygous family members revealed significantly shorter telomeres in siod patients compared to heterozygous family members and compared to agematched, healthy controls. methods: peripheral blood mononuclear cells were isolated using a ficoll-hypaque density gradient, cryopreserved, then sent to repeat diagnostics in north vancouver, bc. telomere length measurements were performed at a single-cell level using flow-fluorescence in situ hybridization as previously described. [ ] telomere length was measured in total lymphocytes, naive and memory enriched t cells, b cells, and nk cells and compared to reference samples from age-matched, healthy individuals. results: compared to age-matched healthy controls, three siod individuals had mean telomere lengths (mtls) less than the st percentile for age across all lymphocyte subsets (total lymphocytes, b cells, nk cells, naïve and memory t cells). in comparison, three unaffected family members had normal mtls ( th percentile< x < th percentile) across all subsets, and two unaffected family members had low mtls ( st< x < th percentile) in some subsets. the siod individual who received a matched-sibling hsct years prior, had normal mtl in nk cells ( th < x < th percentile) but low mtls ( st< x < th percentile) for all other subsets. conclusions: these data show that siod patients have significantly impaired telomere lengths across multiple lymphocyte lineages and support a limiting role for smarcal deficiency in telomere maintenance. in comparison, unaffected family members, heterozygous for smarcal mutations, have mean telomere lengths that are normal or slightly low for age. this suggests that abnormally short telomeres are seen in individuals with homozygous but not heterozygous smarcal mutations. for the individual who received a hsct, we do not have pre and post-hsct telomere data, but these results support obtaining pre and post-hsct telomere length analysis in future cases. abnormally short telomeres have been linked to widespread perturbation of gene expression. [ ] we hypothesize that smarcal deficiency, by the effect of stalled forks and shortened telomeres, leads to perturbation in the transcriptome of affected tissues. shortened telomeres may explain the reduced hematopoietic bone marrow production in siod, as bone marrow failure is a cardinal feature of dyskeratosis congenita, a disorder of impaired telomere maintenance. future studies to investigate the role of telomere maintenance in siod include measurement of telomerase activity in polyclonally activated t cells and transcriptome analysis using rna-seq background: yellow fever is a potentially fatal disease for which only supportive treatment is available. vaccination is the primary strategy for prevention of this disease and the vaccine is extremely effective, but there are a few specific populations where it is contraindicated. regarding iga deficiency (the most frequent primary immunodeficiency), current recommendations in the literature are controversial. there are no specific studies in this disease, so case series addressing the safety or possible adverse events after vaccination are essential for decisionmaking during epidemic scenarios, as experienced in brazil in the last years. in this context, this study aimed to describe adverse events after the use of the yellow fever vaccine in iga deficient patients. method: a retrospective cross-sectional study was conducted including iga deficient patients followed at a specialized pediatric outpatient clinic between and . all patients had at least one year of follow-up. immunoglobulin levels, antibody response to vaccines and lymphocyte subset count were evaluated to exclude other immunodeficiencies or the presence of abnormalities that could contraindicate vaccination. demographic data, the presence of infections and comorbidities, use of immunosuppressive medication and adverse events after vaccine administration of the vaccine were described. results: thirty-eight patients with iga deficiency were included in the study and received the vaccine. vaccinated patients had a mean age at the time of the study of . years (sd ± . y). six out of the presented comorbidities: thyroiditis (n= ), type diabetes mellitus (n= ), celiac disease (n= ) and juvenile rheumatoid arthritis (n= ). all patients were atopic and only one had recurrent infections in the last year despite the use of antibiotic prophylaxis. all patients had normal igg and igm levels for their age, positive vaccine responses for measles, rubella and mumps, and age-appropriate lymphocyte subset count. after months of observation, no immediate or late adverse events were reported. among the non-vaccinated patients, only one had a formal contraindication (systemic erythematosus lupus using immunosuppressive therapy). five out of the non-vaccinated patients reported being afraid of receiving the vaccine, still intended to receive it and for other patients data regarding vaccination was unavailable. conclusion: despite the small number of patients, the absence of adverse events in this case series suggests that immunization with yellow fever vaccine may be safe in iga deficient patients, excluded other contraindications. more studies are essential to confirm the safety and help the decision-making process regarding the vaccine administration for iga deficient patients, especially in this yellow fever outbreak scenario. introduction/backround: immunodeficiency, centromeric instability, and facial anomalies syndrome (icf) is a rare group of autosomal recessive disorders involving the triad of hypogammaglobulinemia, centromeric instability, and facial anomalies. the majority of patients have hypo-or agammaglobulinemia, but t cell defects have also been reported. we present the case of a child with icf- who presented with nk deficiency and ultimately developed an ebv-driven malignancy and was successfully treated with bone marrow transplant. methods: whole exome sequencing and nk cell function via -cr cytotoxicity assay and phenotyping via flow cytometry were performed at baylor college of medicine and texas childrens hospital. centromeric banding studies were performed at university of pittsburgh medical center. results: the female patient presented at months of age with cmv pneumonitis and persistent cmv viremia requiring treatment followed by prophylaxis with valgancyclovir. she initially had hypogammaglobulinemia and low t, b, and nk cells; she had normal trecs, lymphocyte mitogen proliferation responses and zap , mhci and mhcii expression. the hypogammaglobulinemia and t-and b-cell lymphopenia resolved within months after initial presentation as she clinically improved from her cmv infection. she was found to have nk cell deficiency on three separate commercially tested samples. whole exome sequencing revealed a homozygous variant in zbtb indicative of icf- syndrome that was confirmed with sanger sequencing (c. _ del, p.q vfs). repeat nk cell studies confirmed impaired function, and phenotyping showed an increase in cd -bright and a decrease in cd -positive cells, suggesting either impaired transition from immature to mature nk cells or impaired survival of mature cells. her karyotype and centromeric banding studies were normal, as were centromeric instability studies. she later developed a memory b-cell defect and presented at months of age with persistent fever, respiratory distress, loss of vaccine titers, hypogammaglobulinemia and low b and t cells. she was found to have ebv viremia and an eber-positive diffuse large b-cell lymphoma in her right lung. due to tenuous clinical status, she received rituximab for treatment of ebv prior to definitive lymphoma diagnosis. she was treated with chemotherapy per protocol anhl , group b (pre-phase with cop, courses and with copadm, and courses and with cym) and her course was complicated by seizures attributed to methotrexate toxicity. she ultimately underwent reduced intensity conditioning with hydroxyurea, alemtuzumab, fludarabine, mephalan, and thiotepa followed by a cd- selected, hla-matched, unrelated donor peripheral blood stem cell transplant. her early post-transplant course was complicated by adeno-, ebv, and cmv viremia, all successfully treated with antivirals and a donor lymphocyte infusion. she is now greater than months posttransplant, off immunosuppression with % donor engraftment, no evidence of organ toxicity or gvhd, and with excellent immune reconstitution. conclusions: this is the first reported case of impaired nk cell function and phenotype and ebv-driven malignancy in a patient with icf- . this case expands the phenotype of icf- and suggests that early bone marrow transplant should be considered in these children. it also demonstrates a novel requirement for zbtb in human nk cell maturation and function. rationale: common variable immunodeficiency (cvid) is a disorder that affects the production of immunoglobulins and is associated with development of autoimmunity. multiple mutations have been described that are associated with cvid, but plcg mutations have only been described in patients with phospholipase c gamma (plc ) associated antibody deficiency and immune dysregulation (plaid) and autoinflammatory plc associated antibody deficiency and immune dysregulation (aplaid). we present a case of a y/o male cvid patient with recurrent upper respiratory tract infections, steroid-dependent autoimmune thrombocytopenia, low b cell count, hepatosplenomegaly, and restrictive lung disease. he was found with a variant of unknown significance at the plcg gene. in contrast to plaid our patient does not exhibit cold urticaria. method: case presentation of a cvid patient followed in our clinics. patients chart and previous laboratories were reviewed. sequence analysis and deletion/duplication cvid panel testing was performed using invitae© discussion: genetic testing has revolutionized the diagnosis of immune deficiencies, but variants of unknown significance are being increasingly reported. in this case, a variant of uncertain significance was identified which replaces threonine for alanine at codon of the plcg protein. this codon is located at the sh domain, which is part of a region that provides auto-inhibitory enzymatic functions. plaid mutations have been identified in sh domain, but it has been known that both sh and sh domains facilitate plcg association with other proteins. studies with deletion of plcg gene have shown functional abnormalities in b cells, natural killer cells and mast cells. to our knowledge, there has not been any previous report of a cvid patient with a variant mutation at the sh domain of the plcg gene without being diagnosed as plaid or aplaid. our patient has immunodeficiency, recurrent upper respiratory tract infections, steroid-dependent recurrent autoimmune thrombocytopenia, rheumatoid arthritis, hepatosplenomegaly, early-osteoporosis and restrictive lung disease. he does not have cold urticaria as seen in plaid, but exhibits autoimmunity not observed in aplaid. conclusion: conclusion: plcg is an important protein in the pathway of b cell development. a novel mutation in the sh domain of the plcg gene may be associated with the cvid phenotype of low b cells and autoimmunity. this could lead to a gain-of-function mutation as seen in plaid but without early-onset cold urticaria. functional studies are required to confirm the significance of this mutation. primary (or familial) hemophagocytic lymphohistiocytosis (hlh) is a rare, life-threatening hyper-inflammatory disease affecting mainly young children. it is caused by mutations in genes involved in the granule-dependent cytotoxic pathway, and is characterized by extreme inflammation and massive tissue infiltration by activated t cells and macrophages. to this day, hematopoietic stem cell transplantation is the only available curative treatment with a transplantrelated mortality of %. thus, the development of new, more efficient anti-inflammatory treatments would be a significant advancement in the treatment of hlh. here, we hypothesize that combination therapies targeting both jak-dependent and independent cytokines will be more effective than either one alone to reduce the lifethreatening symptoms induced by this pathology. using a perforin-deficient mouse model of hlh, we first compared the effect of targeting individual cytokines with blocking antibodies on the progression of the disease. we show that blocking ifng and il- , but not il- , significantly reduces the severity of hlh. targeting the jak-stat signalling pathway with ruxolitinib, a specific inhibitor of jak and jak , downstream of ifng and il- , but not il- , is similarly beneficial. more importantly, combination therapies using ruxolitinib and blocking antibodies to either ifng or il- show synergistic effects, further mitigating the progression of the disease. these results suggest that jak-dependent and independent cytokines drive the pathogenicity of hlh in perforin-deficient mice. it further supports that ruxolitinib, although effective in reducing the symptoms of hlh, should be used in combination with anti-ifng and/or anti-il- antibodies to prevent hlh progression. this is particular relevant since the former were recently approved for the treatment of hlh while the latter (il- binding proteins) are in clinical trials for il- -dependent macrophage activation syndromes. despite the increased risk of opportunistic lung infection in patients with severe t cell dysfunction (e.g. cd l deficiency) and/or severe cd t cell lymphopenia, we are not aware of any reports of disseminated pneumocystis jiroveci infection in non-human immunodeficiency virus (hiv) patients with primary immunodeficiency (pid). we report the first case, to our knowledge, of disseminated pjp in a patient with cvid like/ctla haploinsufficiency. he had been diagnosed with common variable immunodeficiency (cvid) in , approximately eight years prior to being referred to us, and was on intravenous immunoglobulin (ivig). he was also diagnosed with multilineage evans syndrome in . his medical history was also significant for potential granulomatous lymphocytic interstitial lung disease (glild) (lung biopsy in the remote past with interstitial disease), significant splenomegaly ( . cm), severe portal hypertension, nodular liver disease (likely nodular regenerative hyperplasia) complicated by anasarca, history of chronic diarrhea (potential enteropathy), lymphadenopathy s/p biopsy with nodular lymphoid hyperplasia, and a history of multiple pneumonias. in , he had developed disseminated pjp with lung, liver, and bone involvement. the t vertebra pjp invasion was confirmed with a bone biopsy; gomori methenamine silver staining and pcr were performed and concluded pjp. he was treated with trimethoprim sulfamethoxazole (tmp-smx) and steroids, then was continued on tmp-smx prophylaxis. due to his liver damage and his chronic neutropenia, tmp-smx was replaced by atovaquone as a secondary prophylaxis for pjp. his laboratory studies were significant for an absolute neutrophil count of . k/ul, absolute lymphocyte count of . k/ul, hemoglobin of . g/dl, platelets of k/ul, total bilirubin of . t-cell receptor beta chain repertoire analysis showed an oligoclonal distribution. severe combined immunodeficiency panel through ambry genetic testing was negative as was genetic testing for cd l deficiency. given his complex clinical history, whole exome sequencing was obtained and detected an autosomal dominant heterozygous missense mutation (c. g>a) implicated in ctla- haploinsufficiency and previously reported by schwab et al. our patient is currently undergoing therapy with abatacept (ctla- fusion protein), which has been reported to improve glild, splenomegaly and enteropathy in patients with ctla- haploinsufficiency. he is improving on this regimen. he has met with the stem cell transplant team, but at this point of time, due to his abnormal lung function, his liver damage and his significant splenomegaly, he is not a good candidate. defects in the nf-b signaling pathway are implicated in the pathogenesis of several primary immune deficiencies in humans. the clinical features of these conditions vary significantly, reflecting the complexity of the pathway, and its broad role in innate and adaptive immune responses, and the development and differentiation of lymphoid organs. here we report the first case of a human pid caused by a homozygous mutation in nfkbid in a year-old male. he was the second child of consanguineous parents, and was diagnosed with possible cvid at the age of , after recurrent episodes of pneumococcal pneumonia. however the clinical features have evolved over time; he developed severe ebv infection at age , causing hepatitis and pancreatitis. at the age of , he presented with an anca-negative systemic vasculitis, manifesting as pulmonary haemorrhage, and acute necrotizing pauci-immune glomerulonephritis. pulsed methylprednisolone and cyclophosphamide induced an initial remission, however, relapse a year later led to end-stage renal failure. he is now dialysis-dependent, and due to the underlying pid, and chronic cmv viraemia, is not a candidate for renal transplantation. genomic dna was subjected to whole-exome sequencing. variants were filtered using a model of autosomal-recessive inheritance and functional analysis of primary cells was performed. we identified a novel, homozygous, single-base deletion resulting in a frame-shift, and premature stop in nfkbid. nfkbid encodes ibns, a non-classical inhibitor of nf-b signaling. at diagnosis the patient had reduced levels of igg , iga and igm, elevated ige, with absent humoral immune responses to pneumococcal polysaccharide vaccine, and an intact response to tetanus. lymphocyte numbers were initially within normal reference ranges, albeit with an increased proportion of cd +:cd + t cells. however, over time there has been a significant reduction in b cells and cd + t cells. cd + t cells demonstrated a skewing towards a central memory phenotype (cd ro+/ccr +), and cd t cell proliferative responses to pha were comparable to a healthy control. functional analysis of primary cells from the proband revealed a complete absence of bns protein expression, dysregulated nf-b signaling, and elevated pro-inflammatory cytokine production. the patient is currently receiving a trial of targeted therapy to modulate the aberrant immune responses. this novel pid highlights the importance of regulation of nf-b signalling, in orchestrating an appropriate immune response, maintenance of self-tolerance, and protection against viral pathogens. primary immunodeficiency diseases (pid) are a heterogeneous group of conditions with variable clinical features that are frequently associated with significant diagnostic delay. accurate diagnosis has significant therapeutic benefit and may lead to personalized therapies. we established the immunology flagship of melbourne genomics health alliance in australia to determine the clinical utility of genomic sequencing for diagnosis and management of individuals with suspected and confirmed cases of pid. adults and children with suspected or confirmed pid (n= ), autoinflammatory disease (n= ) and hereditary angioedema (hae, n= ) were recruited to the melbourne genomics immunology flagship. whole-exome sequencing (wes) was performed, with targeted gene analysis. variant curation and reporting was performed according to the american council of medical genetics guidelines. overall, wes was diagnostic in % ( / ), confirming a preexisting diagnosis in % ( / ), and offering a new or more specific diagnosis in % ( / ). variants of uncertain significance were identified in a further patients ( %) in genes known to be associated with their clinical diagnosis, that warrant further functional validation. in the hae group, diagnosis was confirmed in only patients ( %), suggesting that wes may not be the appropriate technique for genetic diagnosis in this condition. a higher diagnostic rate was observed for autoinflammatory disorders ( %; / ) compared to pid ( %; / ). of those who received a diagnosis, immediate changes to patient management and treatment occurred for / patients ( %), including hsct for and specific targeted therapy for ( %) individuals. we have demonstrated the utility of wes for accurate diagnosis of complex immune diseases, with the potential to change diagnoses, guide therapeutic intervention and provide opportunities for genetic counseling. further longitudinal analysis will determine clinical outcomes and health economic implications of genomic sequencing for diagnosis and management of immunological conditions in australia. at birth he had neonatal asphyxia and cerebral palsy. at years old he had presented involuntary movements, left paresis, bilateral horizontal nystagmus. at years of age, he had a right nasal obstruction. it was resected by otorhinolist and informed by biopsy: inflammatory polyp and chronic sinusitis. he has had pneumonias, sinusitis and diarrhea. at the age of years, the ataxia telangiectasia was confirmed by sequencing with pcr ( exons, bp) of the atm gene: transition g> a, nucleotide position , codon , affecting splicing. alpha fetoprotein - . u/ml. brain mri, say cerebellar atrophy. he had igg mg / dl - mg / dl, iga . mg / dl, < mg / dl, igm mg / dl - mg / dl, ige . -< iu / ml. subclasses of igg: igg : . g / dl, igg : . gr/dl, low. igg anti hepatitis b , . no seroconversion. hiv negative tcd + lymphocytes: , %, = cells / mm , ltcd +: , % = , cel / mm , ltcd +: , % = , cells / mm , cd / cd : . . for all of the above, common variable immunodeficiency was diagnosed. he receives human immunoglobulin. at , i arrived at this hospital due to fever, respiratory distress and lymphadenopathy in the neck. ct showed ganglionic conglomerate on right side neck. lymph node biopsy: strong tumors with cd and bcl , weak and moderate diffuse pax- ; negativity with cd , cd and cd , and a cell proliferation index with ki of %, diagnosis: diffuse large b cell lymphoma. treated with rituximab and chemotherapy. lymphoma completely remitted. conclusion: the association ataxia telangiectasia and lymphoma is frequent. by contrast, cvid and ataxia telangiectasia are extraordinarily rare. introduction: chronic granulomatous disease (cgd) is a primary immunodeficiency wherein affected patients are susceptible recurrent infections caused by specific bacteria and fungi as a result of defective nadph activity. additionally, inflammatory complications involving the bowel and lungs can cause significant morbidity. currently the only proven permanent cure to cgd remains hematopoietic stem cell transplant. case: a -year-old patient was diagnosed in infancy with x-linked cgd. at age yrs he received a nonmyeloablative peripheral blood stem cell transplant from his / non-carrier sister as previously reported (nejm : , ) . conditioning was cyclophosphamide ( mg/kg) on d- and d- ; daily fludarabine ( mg/m ) on d- through d- ; antithymocyte globulin at mg/kg on d- through d- . posttransplant immunosuppression consisted of cyclosporine on d- through d+ . he received . x cd + peripheral blood stem cells which were t-cell depleted with x add back of cd + cells on day . after days of neutropenia (anc < ) there were signs of engraftment. per protocol, he received donor peripheral-blood lymphocytes containing . x cd + cells/kg on d+ after transplantation. since donor t cells constituted less than percent of his circulating cd + t cells and he had no graft versus-host disease, he received . ¬ cd + cells/kg on d+ . after the discontinuation of cyclosporine, he received a total of three donor-lymphocyte infusions ( . ¬ cd + cells/kg) at -day intervals achieving % t cell and myeloid engraftment at months post-transplant with no acute nor chronic gvhd. at last follow-up years post-transplant ( ) he had % and % lymphoid and myeloid peripheral chimerisms, respectively. the patient and family declined further periodic followup. then, in october he presented with malaise, cough and fevers. he eventually was found to have a large consolidation and a bal grew burkholderia cepacia. his dhr showed % activity and peripheral blood myeloid and lymphoid chimerisms were % and %, respectively. discussion: this late graft failure following peripheral blood transplant occurred following a conditioning regimen which is not the current standard for transplant in cgd. in the case series in which this patients transplant is reported (nejm ), another patients myeloid chimerism fell to % by years post-transplant, remaining stable at that level of chimerism without any serious infections over regular periodic follow up to the present time. current regimens typically include busulfan to enhance engraftment and prevent graft failure. this case reinforces the need for prolonged monitoring of primary immune deficiency patients after transplantation. introduction: with the introduction of severe combined immunodeficiency (scid) newborn screen (nbs) in the state of kansas in , a case of complete digeorge syndrome (dgs) was discovered in an infant born to a diabetic mother with atypical features. this is the first dgs case diagnosed after adding the scid nbs, which emphasizes the need to establish scid nbs in all states. case presentation: the female infant was born via spontaneous vaginal delivery at / weeks to a year old g now p mother. maternal history was significant for chronic hypertension, obesity, insulin dependent type diabetes, anxiety, depression, and scoliosis. the infant was noted to have a left sided abdominal wall defect and hernia, imaging identifying left renal agenesis, and was initially suspicious for vater syndrome. fortunately, the infant's scid nbs revealed low t cell receptor excision circles (trecs). her initial white blood cell count was . with an absolute lymphocyte count of . k/ul. ebv pcr, cmv pcr, and hiv studies were negative. chest imaging discovered absent thymus, abnormal vertebrae with only ribs on the right and ribs on the left, and abnormally formed thoracic vertebrae (t ). echocardiogram detected an atrial septal defect measuring . cm, possible pfo versus secundum asd. endocrinology was consulted for management of labile calcium and phosphorus levels. fish was negative for q . deletion. microarray r evealed a variant of unknown signif icance arr[grch ] p . ( _ )x . sequence analysis of combined and severe immune deficiency genes showed a variant of uncertain significance c. c>a (p.leu met). management and outcome: additional evaluation included: cd ul ( - ul), cd ul ( - ul), cd ul ( - ul), cd ra cells/ul ( - cells/ul), normal cd , and cd / , normal immunoglobulin g level, and normal dihydrorhodamine assay. skeletal survey, ct abdomen and chest, and hla typing were performed in preparation for thymic transplant. discussion: patients with complete dgs, a form of scid found in less than percent of patients with qds, have absent thymus and a t cell count < standard deviations below normal for age (typically < naïve cd + t cells/mm ). in a large series of patients with complete dgs, only percent had an identifiable q . deletion [ ] . infants of a diabetic mother have various genetic and syndromic associations including diabetic embryopathy. [ ] despite the importance of immunological aspects in pregnancy, few studies have reported on the cellular immune modifications of diabetic embryopathy. diabetes during pregnancy may affect the development of the thymus and thus maturation of the immune system in the offspring. [ ] the recent addition of a trec assay to newborn screening can identify such a subset of infants with atypical presentations. scid nbs uses an assay for trecs, a biomarker of t cell development. [ ] [ ] [ ] this initial presentation now places the immunologist in the role of "first responder" with regard to diagnosis and management of these patients, who may present with atypical features. newer genetic and molecular techniques now allow for earlier identification of immune defects in such disorders with life-long clinical concerns. [ ] references: introduction/background: goods syndrome is a rare cause of combined b-and t-cell immunodeficiency occurring in association with a thymoma. affected patents are susceptible to bacterial, fungal, viral, and opportunistic infections. an association with autoimmunity has also been reported. current knowledge of goods syndrome is primarily limited to case reports and small series. objectives: to examine the spectrum of clinical and laboratory features of a major cohort of goods syndrome patients in the us. methods: we conducted a retrospective analysis of patients with goods syndrome in the usidnet registry and the mount sinai hospital (msh) cohort. r e s u l t s : we i d e n t i f i e d p a t i e n t s w i t h t h y m o m a a n d hypogammaglobulinemia (usidnet, n= ; msh, n= ; median age: years; female: %), representing data from patient years. the median age at diagnosis of thymoma and hypogammaglobulinemia were years (range - ), and . years (range - ), respectively. two patients were deceased (at age and years, cause unspecified). all patients had low igg (median mg/dl, range - ). iga and igm were reduced in % and % of patients, respectively. low cd + b cells (median . /mm^ , range - ) were reported in all available records. the absence of cd + b cells was observed up to years postthymectomy. a wide range of additional laboratory abnormalities were identified: low cd + t cells (n= ), low cd + t cells (n= ), low cd / cd ratio (n= ), low nk cells (n= ), and absent peripheral eosinophils (n= ). the most common sites of infections were lower respiratory ( %), upper respiratory ( %), and gastrointestinal ( %). in addition, sepsis ( %), meningoencephalitis ( %), osteomyelitis ( %), and urinary tract infection ( %) were also observed. identifiable infectious agents included: bacteria ( %), virus ( %), fungus ( %), parasites ( %), and protozoa ( %), with opportunistic infections recorded in % of patients. opportunistic infections were significantly associated with absolute cd lymphopenia (p= . , fishers exact test). enterovirus was identified as a previously unreported cause of meningoencephalitis in this population. autoimmune manifestations were reported in % of patients, with a higher prevalence of inflammatory colitis ( %) than previously reported. hashimoto thyroiditis, fibromyositis, and bronchiolitis obliterans organizing pneumonia (n= each) were identified as previously unreported autoimmune/inflammatory conditions in this population. a case of alopecia areata was also observed. additionally, bronchiectasis was recorded in % of patients. all patients were initiated on immunoglobulin replacement, with antibiotics prophylaxis in %, and immunosuppressive medications employed in % of patients post diagnosis of immunodeficiency. conclusion: goods syndrome is a combined immunodeficiency, with a wide range of autoimmunity in a subset of patients. we expanded upon the spectrum of associated infectious and inflammatory complications through a major us cohort. persistent immune dysregulation was observed up to decades post-thymectomy. introduction: primary immunodeficiencies (pids) constitute a large group of rare disorders that affect the immune systems function. some pid patients develop autoimmunity in addition to having increased susceptibility to infections due to their impaired immunity [ ] . ( ) case presentation/ management: a year old caucasian female with history of bipolar disorder, factor v leiden deficiency, anti thrombin deficiency, pulmonary embolism, endometriosis, and seasonal allergies was evaluated for chronic granulomatous disease (cgd) in . the main symptoms were inflammatory breast lesions necessitating surgeries on the right breast, and back, facial, genital, ocular, mouth, and scalp sores. biopsy with cultures of the wounds was positive for corynebacterium, coagulase-negative staphylococcus, enterococcus, bacteroides species, and provatella. neutrophil oxidative burst was ordered by the infectious disease specialist and showed normal and abnormal neutrophil populations, a finding consistent with cgd carrier. patient was started on interferon gamma- b after failing multiple courses of antibiotics. her symptoms were well controlled on interferon gamma- b mcg/ . ml sq every other day, trimethoprim mg tab ( tabs in am and tab in pm), cefixime mg once daily, and topical mupirocin as needed except for her recurrent genital ulcers. cgd can be rarely associated with oral ulcers however there is a limited literature describing associated genital ulcers. according to the international study group diagnostic criteria published in ( ), the patient was diagnosed by a rheumatologist as having behcets disease (bd). there are no pathognomonic laboratory tests in bd; as a result, the diagnosis is made clinically. patient failed a trial of colchicine and was later started on cyclosporine, which resulted in decrease of her mouth and genital ulcers. discussion: bd is a rare disease mostly seen along the silk road. the prevalence has been reported as . (usa) to (in a single village, northern turkey) for inhabitants. ( ) cgd is a primary immunodeficiency caused by defects in any of the five subunits of the nadph oxidase complex responsible for the respiratory burst in phagocytic leukocytes. patients with cgd are at increased risk of life-threatening infections with catalase-positive bacteria and fungi, and inflammatory complications such as cgd colitis. ( ) reports of cgd female carriers with discoid lupus erythematosus, photosensitivity rashes, and other autoimmune phenomena have been published [ , ] ( ) . to the best of our knowledge, this is the first case to report bd in an affected cgd carrier. the treatment of inflammatory disease in patients with cgd poses a difficult balance between therapeutic immunosuppression and the increased risk of severe infection. ( ) . high dose intravenous immunoglobulin, and targeted therapies such as ctla -ig for t cell mediated pathologies, rituximab for b-cell mediated pathologies, and anti-tnf for ibd, may be preferable over the broad immunosuppressive activity of glucocorticoids. in addition, emerging evidence suggests that hematopoietic stem cell transplantation has indication for cases that have been difficult to control using immunosuppression. ( ) given all that, our case emphasizes the need to maintain suspicion for autoimmune disorders / immune dysregulation in patients with pid. introduction: cd -ligand deficiency is an x-linked combined immunodeficiency, characterized by susceptibility to infection, often with associated neutropenia, malignancy, and autoimmunity. central nervous system (cns) manifestations are less commonly reported than respiratory or gastrointestinal complications, but are most often attributed to infection. herein we describe a challenging case of gradual onset episodic memory loss, confusion, and unilateral hemiplegia in a young male with cd ligand deficiency. case presentation: the patient is a -year-old male with cd -ligand deficiency on immunoglobulin replacement therapy presenting with recurrent, episodic altered mental status (ams) and gradual neurocognitive decline. initial neurologic symptoms began at age years, and included fever, nausea, and eyelid fluttering. initial comprehensive infectious workup at this time, including blood and urine cultures, lyme antibody, serum pcr for hsv, cmv, ebv, respiratory viral pcr including atypical viruses, csf studies including culture, lyme eia, pcrs for enterov i r u s , v z v, e b v, c m v, h s v / w e r e u n r e v e a l i n g . electroencephalogram (eeg) and mri displayed generalized slowing and global atrophy, respectively. definitive diagnosis was not made. the patient continued to decline with worsening developmental delay and memory loss. one year later, at age years, he had a recurrent episode of ams with repeat negative infectious workup including blood and urine cultures, respiratory virus pcr including atypical viruses, csf culture including acid fast bacillus and fungi, cryptococcal antigen, viral encephalitis panel by pcr, and serum pcr for ebv and hhv- . eeg at this time showed left hemispheric epileptogenic potential, consistent with seizure activity. his presentation, at age years, was notable for right-sided hemiplegia with facial numbness, dysarthria, nausea, and fever. he was found to have anello virus on pcr of csf, abnormal left temporal region on eeg, and global atrophy with stable, diffuse generalized volume loss on mri. he was diagnosed with occult anello virus-induced encephalitis with hemiplegic migraine and discharged on valproate. discussion: here we present the first reported case of anello virus detected by pcr in a cd -ligand deficient male with neurocognitive manifestations, attributed primarily to hemiplegic migraine. given the anello virus prevalence and relatively avirulent character, it is presumed to be unlikely culprit for encephalitis; however, the significance of this finding is as yet unknown. this case highlights diagnostic challenges in immunodeficiency: infection may go undetected by standard diagnostic techniques; however, the significance of infections identified with advanced techniques may not yet be understood. background: henoch-shönlein purpura (hsp) is an iga-mediated small vessel vasculitis that presents with a tetrad of abdominal pain, arthritis, glomerulonephritis, and purpura. hsp is typically a selflimiting disease of childhood following a viral illness. there is no universal treatment for patients with chronic or recurrent hsp. we report a chronic refractory case of hsp that was successfully treated with a tumor necrosis factor inhibitor (tnfi), etanercept. etanercept functions as recombinant protein that consists of a tnf-alpha receptor ligand-binding region that links to the fc portion of human igg. it is currently approved for use in diseases: juvenile rheumatoid arthritis, rheumatoid arthritis, ankylosing spondylitis, plaque psoriasis, psoriatic arthritis. tnfi are categorized into two broad categories, recombinant receptors (etanercept) and neutralizing antibodies (ex. infliximab and adalimumab). there have been prior case reports of hsp associated with tnfi agents during the treatment of other autoimmune conditions in the adult population. to our knowledge, there have been prior etanercept related hsp reports, one report associated with adalimumab, and one with infliximab. however, there has been no prior report of etanercept use successfully treating chronic refractory hsp. case presentation: a -year-old native american male with year history of chronic hsp, hla-b positive, and enthesitis related arthritis who was initially treated with steroids, sulfasalazine and methotrexate for symptoms of joint pain and purpura. his iga level was mg/dl prior to therapy. despite treatment for one month of steroids, eight months of sulfasalazine exclusively and eight months of methotrexate and sulfasalazine, he continued to have persistent purpura on bilateral extremities without improvement. he was subsequently initiated on etanercept mg weekly and methotrexate was discontinued. approximately one month later, his rash significantly improved. his rash and joint pain recurs when he misses a dose of etanercept. punch biopsies were taken months after initiation of etanercept. the biopsies of a lesion from his left arm showed early leukocytoclastic vasculitis and from his left leg showed weak granular deposition of iga, igm and c within vessel walls. there is controversy whether this is a true iga vasculitis. however, we believe that his clinical presentation and the deposition of iga and c within blood vessel walls seen on biopsy correlates with chronic henoch-shönlein purpura. conclusion: there is no standard treatment of chronic hsp, but there are reports of benefit with nsaid and corticosteroids. per our literature review, there are no prior reports of etanercept use in the treatment of chronic hsp. tnf inhibitor, etanercept should be considered as a treatment for chronic refractory hsp in the pediatric population as it has showed rapid resolution of purpura in this case report. further studies of etanercept in the treatment of chronic hsp should be conducted given the controversial literature of anti-tnf ab induced hsp during the treatment of other autoimmune diseases. although clinical manifestations of iron overload appear to be quite uncommon in patients who are heterozygous carriers of hfa mutation, we present cases that appear to suggest an increased risk non allergic rhino-sinusitis. case report: we present a year old gentleman with perennial colored rhinorrhea, with facial pressure and tenderness, constant post nasal drip, dry cough and bilateral congestion that had been going on for the past several years. he also had a frequent urge to clear his throat and had frequent episodes of sore throat despite having no history of gerd or lpr. he reported to have multiple sinus infections every year that would progress to pneumonia and eventually require long courses of oral antibiotics. all started in his s intensified in the recent past. he had other siblings; one died in his s due to liver complications of hh and had a carrier sister and brother with a hx of sino nasal problems exactly similar to the patients. his exam was remarkable for bilateral narrowed nasal passages and moderate edema of the mucosa. his rhinolaryngoscopy showed significant edema and purulent drainage, most notably from bilateral middle meati. his skin test was negative. his cbc showed a wbc count of . /ml with % eosinophils and his immunoglobulin panel showed an iga of mg/dl, igg of mg/dl and ige of mg/dl. patient was placed on alkalol sinus rinses and azelastine nasal spray, which he reported to work pretty well. he left for costa rica and is expected to return back with his siblings to a&i clinic in the coming months. discussion: hh is one of the most common inherited disorders in people of northern european descent with an incidence of : and carrier rate of : .. most affected hh patients are homozygous for the mutation designated c y at the hfe gene located at the th chromosome. unlike hereditary hemochromatosis, clinical manifestations of iron overload appear to be quite uncommon in patients who are heterozygous carriers. hh patients are at risk for a number of infections with bacteria whose virulence is increased in the presence of excess tissue iron. hh is also a risk factor for acute fulminant frs . here the mechanism is postulated to be due to quantitative or qualitative neutrophil defects as this condition is mostly seen in patients with dm, aplastic anemia, and can happen in patients undergoing antineoplastic chemotherapy. no known increased susceptibility for infections through either mechanism is postulated for patients with the heterozygous carrier state. here we present hh carrier patients who present with recurrent rhinosinusitis with no allergen sensitizations and normal ige levels. since most fungal immunity is at the tissue level and is cytokine driven, it can be speculated that increased tissue levels of iron might interfere with mechanisms of innate immunity. chief, human immunological diseases section, laboratory of clinical immunology and microbiology, niaid, nih, bethesda, md background: dedicator of cytokinesis (dock ) mutations are associated with a combined immunodeficiency disorder marked by atopic features, infectious susceptibility with a striking preponderance of cutaneous viral disease, and a risk for the development of malignancy including lymphoma. almost all cases can be diagnosed by documentation of the loss of dock protein expression. methods: we describe a -year-old male with a diagnosis of pre-b cell acute lymphoblastic leukemia (all) followed by epstein-barr virus (ebv) associated diffuse large b cell lymphoma (dlbcl). compound heterozygous mutations in dock were documented following the completion of whole exome sequencing (wes). the pathogenicity of the variants was assessed. flow cytometric quantification of intracellular dock protein was completed. dock protein function was assessed by evaluating the morphology of patient lymphocytes when migrating in a d collagen matrix. results: a concern for a primary immunodeficiency was raised due to a history of recurrent otitis media which began at months of age. by years of age, numerous warts were noted on his fingers; however, they were transient for a duration of only years. no atopic features were appreciated. at years of age, a diagnosis of pre-b cell all was made. during all therapy, infectious complications were severe including an intestinal perforation, osteomyelitis, and sepsis. at years of age, still in an ongoing remission from his all, an incidental finding of a lung nodule led to a diagnosis of ebv-associated dlbcl. during therapy, however, infectious complications were again severe including a soft tissue infection and sepsis. wes was performed and compound heterozygous mutations in dock (c. _ del and c. - g>c) were documented. flow cytometric quantification of intracellular dock protein was normal when compared to a normal control. nevertheless, additional functional assessment of dock protein was completed. when migrating through a d collagen matrix, % of the patient lymphocytes studied demonstrated abnormal elongation (stretch ratio > defined by length/width) compared with % of lymphocytes from a normal control. he is being evaluated for hematopoietic stem cell transplant. conclusion: autosomal recessive mutations in dock are a rare cause of a combined immunodeficiency marked by atopic features, infectious susceptibility with a striking preponderance of cutaneous viral disease, and a risk for the development of malignancy including lymphoma. here, pre-b cell all followed by the development of a subsequent malignant neoplasm (ebv-associated dlbcl) led to the discovery of dock deficiency. hence, as our case underscores, for rare instances of high clinical suspicion despite normal dock protein expression, additional functional testing is crucial to make a definitive diagnosis and plan treatment. understanding the spectrum of dock mutants and their phenotypes will improve our understanding of dock deficiency. background: autosomal dominant hyperimmunoglobulin e syndrome (ad-hies) is a rare primary immunodeficiency caused by heterozygous loss-of-function mutations in the signal transducer and activator of transcription (stat ) gene. ad-hies classically characterized by recurrent cold staphylococcal abscesses, pneumonia, eczema, and an elevation of ige level. other additional clinical manifestations of hies have been recognized including skeletal dysplasia (scoliosis, pathologic fractures, delayed dental deciduation), pneumatoceles, coronary-artery aneurysms, brain lesions, and chiari malformations. objective: to describe a unique case of abdominal abscesses in a patient with ad-hies. method: a -year-old female with known ad-hies (c. c>t (p.arg trp)) and a complicated history of early pneumococcal pneumonia and meningococcemia resulting in bilateral amputation below the knees along with loss of several digits, presented for evaluation of skin infection. she had a history of recurrent staphylococcal skin abscesses and presented with inability to use her prostheses due to pain from inflammation around her amputation sites. she underwent imaging and was found to have bilateral extremity abscesses with an associated osteomyelitis of her l tibia (which was found to be mrsa after incision and drainage). while receiving intravenous antibiotics for her osteomyelitis, she developed intractable abdominal pain. imaging showed a thick-walled, multi-septated, paranephric abscess as well as several smaller abscesses scattered throughout her abdomen. she underwent multiple drain placements and drainage of retroperitoneal fluid collections via interventional radiology (ir). purulent fluid from the abdominal abscess drainage grew mrsa. the patient continued to have re-accumulation of abscesses despite multiple drainages. repeat imaging noted increased paranephric abscesses which were not communicating with drains. given lack of response to several ir-placed abdominal drains and to weeks of intravenous antibiotics, she had an open surgical washout with minimal improvement. hospital course was further complicated by development of a left lower lung lobe consolidation and sub-segmental pulmonary embolism necessitating treatment with heparin. finally, after several weeks of escalating antimicrobial therapy and with additional drain placements, the retroperitoneal abscesses started to recede. repeat abdominal imaging several months later while asymptomatic revealed slow but continuing resolution of the abscesses. conclusion: the present case raises awareness of an unusual location for infection in a patient with ad-hies. although the majority of complications of ad-hies are sinopulmonary and skin infections, recalcitrant intra-abdominal abscesses should be considered in the differential of infections in hies. introduction/background: the recent epidemiologic studies have revealed that primary immunodeficiencies (pids) are more common than previously thought. however, there are very few data on epidemiology of pids in korea. objectives: we attempted to estimate the pid epidemiology and disease burden in korea and provide the background information for pid registry for future. methods: to review the previously reported scientific studies, pubmed, koreanmed, google scholar were searched. any studies on pids reported in scientific journal (korean or international) from january to november were searched. both korean and english reports were searched. diagnosis for pid was categorized from group i to group xi according to iuis phenotypic classification. study period was divided into two periods: period from to and period from to , because there was a multicenter study to estimate pid epidemiology from to . in addition, the number of pid patients and the cost for care were estimated among patients who requested reimbursement to health insurance review and assessment service (hira) korea for one year in . results: a total of pid patients were identified in reports. one hundred and ninety-nine patients ( reports) and patients ( reports) were found in period and period , respectively. the pids were reported in patients for immunodeficiencies affecting cellular and humoral immunity, patients for combined immunodeficiency with associated or syndromic features, patients for predominantly antibody deficiencies, patients for diseases of immune dysregulation, patients for congenital defects of phagocyte, patient for defects in intrinsic and innate immunity, patients for auto-inflammatory disorders, patients for complement deficiencies, and none for phenocopies of pid. from hira reimbursement data, the number of pid patients were for combined immunodeficiency, for predominantly antibody deficiency, for common variable immunodeficiency, for functional defect of neutrophils, for immunodeficiency associated with other major defects, for other immunodeficiencies. a total of , pid patients were treated for , days and $ , , was reimbursed in . conclusions: we performed a systematic review on published studies for pid in medical journals and national open data system of hira to estimate the pid disease burden for the first time in korea. to obtain more information on true pid epidemiology and disease burden in korea, a national multicenter study for pid registry is required in the future. micro-thrombocytopenia is one of the most serious challenges for wiskott-aldrich syndrome (was) and x-linked thrombocytopenia (xlt) patients. thrombocytopenia leads to severe, potentially life-threatening, bleeding episodes, which require frequent transfusions and account for % of deaths in patients experiencing was mutations. the gold standard treatment for was patients is hematopoietic stem cell transplantation (hsct) from an hla-identical donor but more recently a number of gene therapy (gt) trials in europe and usa showed promising results. in particular, it has been shown that was patients receiving lentiviral mediated gt, consisting of autologous cd + cells transduced with lentiviral vector encoding the human was gene under the control of the endogenous promoter, in combination with a reduced intensity conditioning regimen, have a significant increase in platelet (plt) counts. even though plt counts do not reach normal levels, treated patients decreased the severity and frequency of bleedings. here, in a cohort of xlt and was patients, fifteen treated with gt, the plt phenotype and function were analyzed by electron microscopy, flow cytometry and proteomic profile. the aim of the project is to assess the presence of plt defects in was untreated patients and the impact of gt treatment on the correction of plt behavior. we demonstrate that plts of untreated was patients have reduced size and abnormal ultrastructure along with hyperactivated phenotype at steady state, showing increased expression of cd p, activated iib integrin and cd l; conversely, activation response to agonist and aggregation capacity are both decreased. analyzing plt samples isolated from treated patients, we found that gt restores plt size and ultrastructure very early after treatment and reduces the hyperactivated phenotype proportionally to was protein (wasp) expression and follow-up length. plts isolated from gt treated patients showed a normal activation response to agonists and restored aggregation capacity in out of analysed patients. by proteomics, various protein pathways were found downregulated in untreated plt samples, mainly involving cytoskeletal-rearrangement proteins, integrins, signal transduction molecules, vesicles-transport proteins; additionally, decreased metabolic capacity were observed. these results are in line with the functional defects observed in plts in terms of activation and aggregation. conversely, the expression of protein-pathways found downregulated in untreated patients is comparable to healthy controls in gt-treated plt samples, reflecting the amelioration of plt phenotype and function. overall, our study highlights the coexistence of multiple defects in the activation and aggregation responses occurring in was patient plts in absence of wasp. gt was able to normalize the plt proteomic profile followed by consequent restoration of plt ultrastructure and phenotype, suggesting gt is responsible for the observed reduction of bleeding episodes in treated patients. introduction: pik cd is an autosomal dominant genetic disorder of the immune system that results in persistent activation of pi k. signaling through pi k is essential for immune cell regulation of metabolism, migration, proliferation and differentiation, leading patient to present with lymphadenopathy, immunodeficiency and senescent t cells. the mutated protein causes t cells to over activate and mature too quickly leading to their death, this over activation also blocks the maturation of b cells. case presentation: a -year-old female with a childhood history of failure to thrive, asthma, chronic rhinitis and common variable immunodeficiency on intravenous immunoglobulin replacement, was seen in immunology clinic to establish care. she reported frequent episodes of pneumonia and bronchitis in her childhood. her family history was significant for family members with leukopenia, but no diagnosed immunodeficiency. patient had son who did not report symptoms concerning for immunodeficiency. physical exam was within normal limits with no lymphadenopathy. laboratory examinations exhibited normal iga ( mg/dl), igg ( mg/dl), and igm ( mg/dl). while flow cytometry showed normal absolute cd ( - cells/ul), cd ( cells/ul), nk cells ( cells/ul), cd ( cells/ul), cd ra ( cells/ul), cd ro ( cells/ul), cd ( cells/ul), and hla-dr ( cells/ul), nonswitched memory cells ( cell/ul) and class-switched memory cells: ( cells/ul). ( - cells/ul). vaccine response was not pursued as patient had been on ivig. genetic testing was pursued, and revealed a mutation in pik cd gene, specifically a mutation in the c. g>a; p.val met variant (rs ). this mutation though seen in databases, is not currently reported in medical literature as associated with this condition. based on these, ct chest was ordered to screen for bronchiectasis, adenopathy and lymphoma. ct showed no cardiopulmonary disease or adenopathy, but did show an incidental adrenal mass which is now being worked up. while the pattern of inheritance of this mutation is autosomal dominant, her son is asymptomatic and testing of her son has not been pursued, though it was advised for her cousins given history of leukopenia. patient has continued on igg replacement therapy. conclusion: recent publication by the clinical immunology society suggests consideration for next generation sequencing when it can affect future family planning or it has treatment and prognostic implications. this case highlights all aspects of the importance of genetic testing as part of the diagnosis of cvid, since it can affect progeny, it offers the possibility of treatment with immune modulating agents and has implications on screening, since patients are at increased risk for malignancies. background: abnormal v(d) j recombination activity in patients with mutations in the recombination-activating genes and (rag / ) results in markedly reduced usage of distal vand j genes at the t cell receptor alpha (tra) locus. mucosa-associated invariant t (mait) cells express a semi-invariant t cell receptor containing the distal trav - gene. mait cells can be identified by flow cytometry using a mab directed against valpha . , which recognizes the product of the trav - gene. by performing high throughput sequencing (hts) of tra rearrangements and flow cytometry, we have confirmed lack of t cells using distal valpha genes in patients with known rag mutations. we now report that flow cytometry with mab against valpha . successfully identified rag deficiency in two patients with an atypical presentation. methods: tra rearrangements were analyzed by hts using gdna from sorted t cell subsets from rag-mutated patients and healthy donors. distal valpha usage was measured in whole blood by flow cytometric analysis with an anti-valpha . antibody. rag mutations were detected by sanger sequencing. patients were enrolled in niaid protocol -i- . results: hts of tra rearrangements revealed lack of distal trav and traj gene usage in patients with rag / mutations. the presence of circulating mait cells in controls and patients with known rag / mutations and various clinical phenotypes was analyzed by flow cytometry using mab against valpha . . we found a virtual lack of valpha . expression in rag mutated patients (< . %) compared to controls ( - %) . we used the valpha . assay to test two patients with unknown immunodeficiency manifesting as skin granulomas and autoimmune cytopenia, and found nearly absent expression ( . % and . %). targeted sequencing of rag / revealed that both patients were compound heterozygous for rag mutations: p.r h/p.c y and p.r w/p.r q, respectively. conclusions: patients with mutations in rag / demonstrate a skewing of their tcralpha repertoire. the reduction in recombinase activity in these patients does not allow for rearrangements of the most distal valpha segments. rapid identification of patients lacking valpha . + t cells by flow cytometry may prompt sanger sequencing and identification of rag / mutations in a matter of days. this assay represents a simple but powerful tool to reduce the cost and time associated with other analysis methods. acknowledgements: supported by dir/niaid/nih. director, centro de inmunología clínica dra.bezrodnik y equipo introduction: the fate of effector t cells is strongly dependent on the expression of bcl- or blimp- , which are inhibited reciprocally through a complex signaling pathway. several studies have shown that bcl- is a key transcription factor for differentiation towards the follicular helper t cells (tfh) lineage able to collaborate with b lymphocytes (bl). on the contrary, the transcription factor blimp- is highly expressed in t lymphocytes th , th and treg, thus regulating the differentiation towards tfh. materials and methods: whole fresh blood and peripheral mononuclear cells from a patient with homozygous mutation in stat b were analysed by flow cytometry. analysis of ctfh (cd +cd ra-cxcr +), ctfh (cxcr +), ctfh (ccr +), ctfh (cxcr -ccr -), naïve bl (lb igm+igd+cd -), memory (mbl) (lb igm+ igd-cd +), switched (mbl-sw) (igd-igm-) and plasmablast (pbc) (cd +cd ++) cells was performed. immunoglobulins were measured in serum. results: the patient with stat b deficiency showed increased values of ctfh ( %) (healthy donors p -p : , - , %) that presented an activated phenotype (icos+ and pd- +) with a skewed to a th profile (ccr +), consistent with her hipergammaglobulinemia and the marked and sustained increase in the switched mbl and pbc subpopulations in peripheral blood over the years. discusion: this immunological phenotype described in the patient with stat b deficiency could explain in part the pathophysiology of the autoimmune disorders. this patient (as well as the other two patients with mutations in stat b previously described by our group), have had chronic hypergammaglobulinemia, autoantibodies and consequently autoimmune processes (psoriasis, hypothyroidism, eczema, alopecia and celiac disease, among others). we believe that the link between this clinical symptomatology and the molecular defect relies in the fact that the absence of stat b promotes a greater expression of bcl- , which generates a bias towards the production of ctfh cells, that give rise to a greater activation of lb, generation of lbm and plasma cells (dysregulation in the cg), events that manifest as hypergammaglobulinemia and autoimmunity. in summary, we provide promising evidence of the mechanisms that lead to autoimmunity in this type of patients that could also be a consequence of the defect in the regulation of gc, highlighting the crucial role of stat b in the humoral immune response and maintenance of the tolerance of the immune system. background/introduction: the term primary immunodeficiencies (pid) encompasses a phenotypically and genetically diverse group of conditions. genetic testing for these conditions can guide treatment, reduce morbidity and mortality, allow for genetic counseling, and identification of additional at-risk family members. however, this testing can be complicated by a number of factors, including pseudogenes, high homology, methodology limitations, and the heterogeneous nature of pids. methods: mayo clinic laboratories launched their first set of nine pid next generation sequencing (ngs) tests approximately one year ago. these tests include one single gene assay for gata deficiency and eight targeted next generation sequencing panels for: atypical hemolytic uremic syndrome (ahus), autoinflammatory disorders, b-cell disorders, monogenic irritable bowel disease (ibd), phagocytic defects, severe combined immunodeficiencies (scid), and severe or cyclic neutropenia. herein we summarize our first year of experience with these ngs tests, with a focus on the eight targeted panel tests. results: from march through november we performed testing for cases. our highest volume of tests was for the ahus panel ( / cases, %). a variant was reported in / cases ( . %). these variants included variants of uncertain significance, likely pathogenic variants and pathogenic variants. the indication with the highest percentage of cases where a variant was reported was scid ( / cases, . %). the number of cases that were considered solved, where the genotype likely explains the patients phenotype, varied widely by indication. twenty cases were found to have a pathogenic or likely pathogenic variant or variants; however / cases were heterozygotes for an autosomal recessive condition and were not considered solved cases. the panel with the highest percentage of solved cases is our scid panel ( / cases, . %). conversely, we have yet to solve an autoinflammatory, irritable bowel disease, or telomere defects case; however % of cases in each of those three panels have had a variant of uncertain significance reported. we hypothesize that one of the reasons for the low detection rate for these three panels is inappropriate test orders. we are also actively looking for ways to update all panels to increase detection rates and clinical utility, for example expanding the gene list of our ibd panel, including large deletion/duplication detection, and including ncf , a difficult gene to capture by ngs, on the phagocytic panel. finally, we present the molecular findings from a number of interesting cases that were solved using our targeted ngs panels. conclusions: the launch of our pid ngs tests in march of has allowed us to aid patients by confirming diagnoses and providing molecular diagnoses that will enable more accurate genetic counseling and risk assessment. we have also uncovered areas for improvement, both on the clinical side: provider education is important to enable better identification of patients who can benefit from molecular genetic testing for pids, and on the laboratory side: introduction of more expanded panels and additional methodologies. the progressive decrease of red blood cells, platelets or neutrophils via a self-directed immune process is jointly termed as autoimmune cytopenias. while autoimmune cytopenias, including autoimmune hemolytic anemia (aiha), immune thrombocytopenic purpura (itp), and autoimmune neutropenia (an), are a common presentation of autoimmunity in the general population, they are particularly frequent and can appear as the first sign in patients with primary immunodeficiencies (pids). possible causes of cytopenia in pids comprise mainly immune dysregulation, bone marrow failure (bmf) and myelodysplasia. our goal is to investigate possible immune mediated mechanisms underlying chronic cytopenia in children in order to achieve an early diagnosis and consequently offer timely and appropriate therapy. we selected patients affected by chronic cytopenia, evaluated with immunophenotyping by flow-cytometry; data were subjected to multivariate analysis by principal component analysis (pca). next generation sequencing (ngs) analysis of genes frequently implicated in pids was performed. among the patients, were affected by bone marrow failure, of which were diagnosed with fanconi anemia and severe congenital neutropenia; were affected by immune-mediated cytopenia and by idiopathic cytopenia. the immunephenotyping showed a typical pattern of cd t cell subpopulations expression in patients compared with healthy donors with an increase of naïve t cells and a reduction of central memory (cm) and effector memory (em) t cells levels. we observed a decrease in total b cells, b switched and b memory cells and an increase in cd low cells. pca showed an overlap between groups, however it revealed a peculiar trend of some single patient, suggesting the pathway involved in immune defect. preliminary results from ngs studies revealed genetic variations in genes previously associated with pids in out of patients investigated. in particular we identify one patient with a mutation in fas, one with a mutation in aire and one with a mutation in ikaros. concerning the remaining patients further studies are ongoing to validate the pathogenicity of the genetic variations. pca is a very effective tool to analyze several parameters at the same time, highlighting patients whose phenotype shows the main peculiarities. the presence of specific lymphocyte subpopulation patterns can be important indicators of immune-mediated cytopenias and helpful signs of specific pids that should promptly be investigated with genetic analysis. the rapid of discovery of novel, monogenic primary immunodeficiencies has been made possible by the broad availability of clinical whole exome sequencing (wes). however, clinical wes has major shortcomings that should be understood by practicing immunologists. focusing on the iuis list of~ monogenic primary immunodeficiency genes, we show here limitations in coverage that could significantly impact clinical interpretation. on the agilent whole exome capture kit, the most common wes platform, there are a number of genes with exons that are poorly covered. specifically, there are at least genes with less than % exonic coverage, with less than % coverage and with less than % coverage (e.g. ikbkb, ncf , taci, unc b and tbx ). beyond this challenging technical issue, there are more subtle issues as well. these include the presence of pseudogenes in at least of our genes (e.g. ak , c qbp, cd , cftr, cr , msn, ncf , ncstn, ikbkg, nhp , pms , pten, rnaseh c, rps, sbds and was), which can make accurate sequencing very challenging. finally, there are many known causative intronic (e.g. btk, ctla- , wasp) and copy number variant mutations (e.g. rag and xiap) as well as large deletions (e.g. dock ) that we cannot expect to be optimally covered using wes. this list of genes requires consideration even with a negative exome and may require additional approaches including whole genome sequencing, sanger sequencing, cnv arrays and/or long-read ngs sequencing. wes is a powerful genomic diagnostic tool, but to avoid missing key diagnostic insights using these alternative approaches may be critical when certain genes are in the differential diagnosis. going forward, as pid phenotypes continue to broaden, these issues remain fundamentally important even if these genes are not obviously implicated in a given clinical phenotype. more physicians are utilizing targeted genetic panels to reach a definitive diagnosis for their patients with immunodeficiency. however, this increase in testing also has led to the discovery of many more variants of uncertain significance (vus) in the genes tested. these findings can often leave the patient and the physician with more questions than answers. we present a patient with recurrent infections found to have multiple variants of uncertain significance in several genes associated with primary immunodeficiency. a -year-old female who was diagnosed with crohns disease at age after intestinal perforation and jejunal resection experienced two discrete episodes of epstein barr virus (ebv) meningoencephalitis and septic shock. the first episode was diagnosed when patient had fever and altered mental status and occurred prior to her crohns disease diagnosis and the second episode was complicated with altered mental status, disseminated intravascular coagulation (dic) and hypotension requiring picu admission. aside from these two major infections, the family denied any other infections requiring antibiotics in the last years and reported a remote history of repeated streptococcal pharyngitis that have not recurred. immunology was consulted at the time of the second episode of meningoencephalitis and work up was mainly unremarkable with normal immunoglobulins, adequate vaccine response to hib, tetanus, diphtheria, rubella, measles and pneumococcus ( out of protective titers). she had normal t cell numbers with slightly decreased natural killer numbers for age. neutrophil studies showed normal dihydrorhodamine (dhr) analysis, glucose- -phosphate dehydrogenase levels and myeloperoxidase (mpo) stain. commercial testing of her toll like receptors ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) showed normal function. invitae primary immunodeficiency panel demonstrated a heterozygous variant in nod (c . c>t; p.arg trp) as well as heterozygous variants of uncertain significance in il r (c. g>t; p.ser ile) and tlr (c. c>g; p.leu val). the patients nod variant is known to be associated with an increased risk for crohns disease. even with our patients presentation with recurrent severe viral infections and ibd, it is not immediately clear how these genetic results explain the pathology. innate immune defects probably contribute to her presentation and it is currently unclear if and how the combination of multiple genetic variants has left her immunologically vulnerable. we use this case to demonstrate that even when genetic testing does not elucidate a clearcut diagnosis of primary immunodeficiency, it can still provide helpful insight into a patients underlying immune phenotype. introduction: xiap deficiency is a rare primary immune deficiency characterized by hemophagocytic lymphohistiocytosis, recurrent fever and inflammatory syndromes, inflammatory bowel disease, hypogammaglobulinemia, recurrent infections, and other manifestations. loss of xiap results in abnormal tnf receptor signaling and nlrp inflammasome actvity which leads to dysregulated production of il- beta and il- . we hypothesized that suppressing the nlrp inflammasome with either targeted deletion or pharmacologic inhibition would suppress abnormal production and secretion of inflammatory il- beta and il- . methods: bone marrow derived macrophages (bmdms) from control, xiap-deficient, and xiap and nlrp double knock-out mice were derived with week of culture in l -cell conditioned media. bmdms were stimulated with a variety of tlr agonists or tnf-alpha, with or without a variety of inhibitors including the nlrp inhibitor mcc , the cathepsin b inhibitor ca- , and quercetin, which is a natural flavonoid (antioxidant) found in many fruits and vegetables, and available as a nutritional supplement. il- beta, il- , and tnf-alpha were measured in supernatants by elisa, and cell death was evaluated by flow cytometry using pi exclusion. results: as expected, bmdms from xiap deficient mice had markedly increased tlr-agonist-or tnf-alpha-induced il- beta production compared to normal bmdms. genetic deletion of nlrp and the pretreatment of cells with the nlrp inhibitor mcc greatly reduced abnormal il- beta production; residual production of il- beta could be inhibited by caspase- inhibition. pre-treatment of cells with the cathepsin b inhibitor ca- also decreased cytokine production but was toxic at higher concentrations. quercetin reliably abrogated il- beta, and also il- . quercetin was found to inhibit priming of the nlrp inflammasome (decreased upregulation of pro-il beta and nlrp ) and also decreased tnf-alpha secretion following tlr agonist stimulation. conclusion: quercetin suppresses the nlrp inflammasome and may be a promising therapeutic option for patients with xiap deficiency. it prevents il- beta and il- secretion. it is a particularly appealing option given that it is a naturally occurring antioxidant, has a great safety profile, and is readily available as a nutritional supplement. human studies are needed. recently, single cell rna sequencing (scrnaseq) analysis in mice has disclosed an unexpected complexity of thymic stromal cells, and medullary thymic epithelial cells (mtecs) in particular. however, the developmental origin, hierarchy, and function of these subpopulations remain ill-defined. moreover, although cortical tecs (ctecs) are thought to represent a more homogeneous population, their characterization has been largely restricted to the adult thymus. we have previously shown that impaired lymphostromal cross-talk in the thymus of patients with combined immunodeficiency (and of corresponding mouse models) is associated with abnormalities of thymic architecture and tec maturation. here, we sought to compare tec distribution and gene expression in wild-type (wt) and in mice carrying rag hypomorphic mutations observed in patients with combined immune deficiency and immune dysregulation. methods: multi-color flow cytometry and scrnaseq were used to analyze composition and distribution of ctec and mtec subpopulations in wt and rag mutant mice at various weeks of age (niaid animal protocol: lcim- e). results: we observed that rag mutant mice have an excess of ctecs, and that their mtec compartment is predominantly represented by cells with high levels of mhc class ii (mhc-ii) expression, recapitulating the phenotype of neonatal wt thymi. while mhc-iihi mtecs are thought to represent a minor fraction of mtecs in adult wt mice and include mature aire+ cells, a relative abundance of mhc-iihi mtecs is observed also at neonatal age, where they are thought to represent immature mtecs. to define more precisely tec maturation, we performed scrnaseq on sorted cd -epcam+ cells, and identified and distinct clusters of tecs in wt and rag mutant mice, respectively. a large proportion of cells in rag mutant mice could be ascribed to the ctec compartment, confirming our previous flow cytometry and histopathology results. furthermore, scrnaseq analysis also disclosed a different distribution of mtec subsets in wt and rag mutant mice. to address the hypothesis that this difference in ctec and mtec abundance and subset distribution may reflect different maturation stages in tec development in wt and rag mutant mice, we will perform lineage tracing and transplantation experiments, and we will also extend tec scrnaseq analysis to wt and mutant mice of embryonic and neonatal age. in parallel, to evaluate the contribution of thymocyte maturation in shaping the stromal populations, scrnaseq will be performed on thymocytes. conclusions: we have further refined the complexity of tecs, and shown that impaired development of t cells in combined immune deficiency (as exemplified by rag mutant mice) has profound effects on the composition and maturation of tecs and may thus contribute to abnormalities of immune tolerance that are often associated with these conditions. the advent of next-generation sequencing (ngs), with the development of whole-exome sequencing (wes) in particular, has allowed the identification of unknown genetic lesions for many diseases and the implementation of specific therapeutic strategies. primary immunodeficiencies (pids) are a group of rare diseases which have benefited from ngs, with the discovery and molecular characterization of previously genetically undefined diseases and the identification of novel molecules involved in the regulation of the immune system. pids are often associated with autoimmune disease due to the dysregulation of the immune system as a whole. the clinical phenotypes are heterogeneous and often overlapping. while a monogenic cause of disease has been identified in a most subsets of patients, the recent application of whole-genome sequencing has found that a polygenic cause is likely. our aim is to investigate the genetic background of patients with immunedysregulations and autoimmunity and to evaluate the possible pathogenicity of the identified gene variants through extensive functional studies. we select patients with sign of immunedysregulation and autoimmunity, extended immunophenotyping and next-generation sequencing (ngs) analysis of genes frequently implicated in pids was performed. in six of them we identify a single gene as responsible of the clinical feature. in particular, we identify two patients with gain of function mutation in stat , one patient with a mutation in ctla , one patient with an activating pik cd mutation, one with a rag mutation and one with a fas mutation. in most of them variants in multiple genes have been detected. interestingly, we find that some genes are recurrently mutated in more then one patient such as was, dock , casp , casp , nfatc and fcgr a. further studies are ongoing to validate the effect of the variations identified. our results strongly suggest that the old hypothesis, based on a single gene mutation as a cause of illness, should be revised in favor of the concept that "is the sum that causes the effect" and that a different point of view on pids now seems inevitable. physician, omni allergy, immunology, and asthma introduction/background: immunoglobulin replacement therapy (igrt) may be optimized to reduce the severity and incidence of infections and potentially delay or abrogate the development of pulmonary complications of primary immune deficiencies. pulmonary complications including bronchiectasis are common in common variable immune deficiency (cvid) and contribute significantly to morbidity and mortality in these patients. it remains unclear whether continued obstructive bronchial changes are a result of repeated respiratory infections, associated inflammation and immune dysregulation, or simply lung-damage that is irreversible by the time therapy is initiated. it has also been suggested that under-treatment in addition to the diagnostic delay may contribute to the development of bronchiectasis in patients with pid. lower serum igg levels with any given dose of immunoglobulin replacement therapy have been demonstrated in patients with bronchiectasis compared to those pid patients without this complication. in addition, earlier studies have shown that greater doses of ig ( mg/kg/ month) may reduce the frequency and duration of infections and help prevent or slow progression of chronic lung disease. objective: to evaluate the prevalence of bronchiectasis in a cohort of patients with a diagnosis of cvid and identify associated ig dosing patterns and clinical outcomes. methods: data were analyzed from the ideal (immunoglobulin, diagnosis, evaluation, and key learnings) patient registry. this is a prospective, longitudinal registry study of patients receiving ig replacement therapy in the home or ambulatory infusion suite with one national home infusion provider. nursing and pharmacy standard of care forms were collected, and dose, infection rate, and prevalence of bronchiectasis were evaluated in patients with a diagnosis of cvid (icd- codes: d . , d . ) results: there were patients in the registry with cvid, ( . %) of which bronchiectasis was also observed. seventy-nine percent (n= ) of the study population was female, and % (n= ) of the cases of bronchiectasis were observed in females. the mean age of the patients with concurrent bronchiectasis was ± . at start of care compared to ± . in those without this observed bronchial obstruction. most bronchiectasis patients (n= ) received igrt subcutaneously every week with a mean dose of . ± . mg/kg/wk. the mean dose of ig in the remaining patients receiving ig intravenously was . ± . mg/kg/month. the average annual rate of infection in ivig and scig patients with bronchiectasis was . ± . and . ± . , respectively, however many were serious bacterial infections. at time of analysis, of the bronchiectasis patients remained active in the registry and had withdrawn. reasons for withdrawal included stopping igrt due to the following: patient decision (n= ), physician decision (n= ) insurance change (n= ), and patient expired (n= ). conclusions: there were documented cases of bronchiectasis in our cohort of cvid registry patients, and dosing patterns aligned with standard doses despite the presence of bronchial obstruction. further studies are necessary to assess evolution of lung damage with respect to ig dosing in patients with cvid. background: activated phosphoinositide -kinase syndrome type (apds ) is a combined immunodeficiency resulting from gain-offunction (gof) mutations in pik cd, the gene encoding the catalytic subunit of phosphoinositide -kinase (pi k). this form of pid is characterized by recurrent respiratory tract infections, susceptibility to herpes virus infections, impaired antibody responses, lymphoproliferation and autoimmunity. previous studies showed that patients with apds have b cell defects that contribute to the clinical phenotype. furthermore, these patients display t cell abnormalities, including increased numbers of memory t cells and t follicular helper cells (tfh), reduction of naïve t cells and impaired t regulatory cell (treg) function. whether these t cell abnormalities are also associated with perturbations of t cell repertoire in unknown. objective: we aimed to investigate the effects of increased pi k signaling on the t-cell repertoire of patients with apds. methods: high throughput sequencing was used to study composition and diversity of t-cell receptor (tra) and t-cell receptor (trb) repertoire in sorted treg, tfh, conventional cd + (tconv), and cd + t cells from patients with pik cd gof mutations and healthy controls. results: treg cells of patients with apds show restriction of tra and trb repertoire diversity, and increased clonality. no repertoire restriction was detected in tfh, tconv, and cd + t cells from the same patients. however, the trb repertoire of treg and cd + cells was enriched for the presence of hydrophobic amino acids in position and of the cdr , a biomarker of self-reactivity. conclusion: these data demonstrate that the t-cell repertoire of patients with apds is characterized by a molecular signature that may contribute to the increased rate of autoimmunity associated with this condition. furthermore, our result support the notion that the pi k pathway is a key regulator of treg cell development and homeostasis in humans. j clin immunol ( ) (suppl ):s -s s ( ), iii. predominantly antibody deficiencies ( ), i. immunodeficiencies affecting cellular and humoral immunity ( ), vii. auto-inflammatory disorders ( ), ix. phenocopies of pid ( ) . two non related cases of ataxia-telangiectasia and one case of schimke syndrome (smarcal compound heterozygous mutation) were diagnosed in the last year. we observed a wide range of age (we evaluate adult and pediatric population) with a male:female ratio close to : immunodeficiency, immune dysregulation, and systemic autoimmunity. clinical diagnosis of these disorders is complicated by overlapping phenotypes. in april , a -gene next generation sequencing (ngs) panel inclusive of copy number variation analysis was launched by a commercial laboratory to facilitate clinical diagnosis of primary immunodeficiency (pid), monogenic autoimmunity and autoinflammatory disorders. we assessed the outcomes of genetic testing utilizing this panel on a cohort of pediatric patients with immunohematologic phenotypes evaluated at our tertiary care center during an -month period ( / / - / / ). eligible subjects were evaluated by at least two of three providers from a multidisciplinary pediatric hematology-immunology team, including a hematology physician, immunology physician and a geneticist or genetic counselor. twenty-three patients met inclusion criteria; ( %) were caucasian, ( %) were male with an average age of . years. the two most common phenotypic diagnoses included cytopenias, single-or multilineage (leukopenia, neutropenia, anemia, thrombocytopenia) primarily attributed to autoimmune causes or hypogammaglobulinemia. five ( %) were given a definitive genetic diagnosis as a result of panel testing, though in two of these cases, the causative mutations were listed as variants of uncertain significance (vus). diagnoses included common variable immunodeficiency due to a pathogenic variant in nfkb , stat multiorgan autoimmunity due to gain-of-function mutation, and familial cold autoinflammatory syndrome due to a pathogenic mutation in nlrp . biallelic dnmt b vus were found in a patient whose phenotype and further laboratory studies (including karyotype) were consistent with immunodeficiency-centromeric instability, facial anomalies syndrome. further, a stat vus was identified in a patient with multiorgan autoimmunity and his father with hypothyroidism; studies from an outside research laboratory were consistent with gain-of-function with this variant (private communication). an additional three patients had vus identified that were suspected to be related to their phenotype, prompting eligibility for research studies. four ( %) patients had increased risk alleles in nod , conferring an increased risk of crohns disease. three ( %) patients had pathogenic or likely pathogenic carrier findings warranting genetic counseling. in addition, vus (an average of per patient) thought to be unrelated to phenotype were identified, necessitating further investigation and counseling. the use of an ngs panel in a cohort of pediatric patients with immunohematologic disorders led to a definitive diagnosis in % of previously undiagnosed patients and prompted further research investigation in several more. genetic testing also led to the identification of clinically significant carrier findings, risk alleles and vus unrelated to phenotype, necessitating genetic counseling. our experience illustrates the value of genetic testing for diagnosis of immunohematologic disorders, and the importance of multidisciplinary care, including genetic counseling, for the proper evaluation and management of these patients. background: allogeneic hematopoietic cell transplantation (allohct) is curative for primary immune deficiencies (pid). however, many patients lack a fully-matched unaffected sibling, or may have an unknown underlying genetic defect, rendering it undesirable to use related donors. many pid patients have significant comorbidities at the time they are referred to allohct, precluding the use of myeloablative conditioning. the use of alternative donors with reduced-intensity conditioning (ric) has historically led to increased rates of graft failure, graft-versus-host disease (gvhd), and transplant-related mortality (trm). posttransplantation cyclophosphamide (ptcy) as gvhd prophylaxis immunomodulates the graft through the preferential sparing of regulatory t cells and hematopoietic stem cells from its cytotoxic effects, thus allowing for robust donor engraftment that overcomes the hla barrier while effectively preventing severe acute and chronic gvhd. we report the outcomes of two institutions using a ric allohct regimen with alternative donors and ptcy in patients with pid. design: we transplanted pid patients (table ) using alternative donors and ric, either serotherapy-free (n= ) or alemtuzumab-based (n= ). all patients received ptcy for gvhd prophylaxis on days + and + , either alone (n= ), or combined with mycophenolate mofetil and either sirolimus (n= ) or tacrolimus (n= ). donors included haploidentical family members (n= ), matched unrelated (n= ), and mismatched unrelated (n= ). stem cell source was t cell-replete bone marrow (n= ) or peripheral blood stem cells (n= ). results: the median follow-up is months (range . - years). at months, overall survival is %, and event-free survival (defined as alive without graft failure) is %. the median days of neutrophil and platelet engraftment are (range - ) and (range - ), respectively. there were patients who developed acute gvhd, grade (n= ) or grade (n= ), and there were no cases of grade or agvhd. seven of eight patients treated with systemic corticosteroids responded, and one was corticosteroid-dependent, then responded to second-line therapy. one patient developed skin-only chronic gvhd, which responded to corticosteroids and puva light therapy. five patients developed graft failure, either primary (n= ) or secondary (n= ), and four were successfully re-transplanted and remain engrafted. one patient with secondary graft failure had autologous recovery and has not required a second allohct given some durable infection control gained during initial engraftment. there were three deaths prior to day due to infection, and one death at . years secondary to presumed overdose. in ongoing follow-up of engrafted survivors (n= ), evidence of phenotype reversal has been demonstrated in all patients, with complete or ongoing resolution of some or all of their underlying disease manifestations, including infection, transfusion-dependence, autoimmunity, malignancy, and/or immune dysregulation. discussion: we have observed high rates of engraftment, low rates and severity of acute and chronic gvhd, and low trm in patients with pid transplanted using alternative donors, ric, and ptcy-based gvhd prophylaxis. ric allohct with ptcy shows promise for curing pid, and its use minimizes toxicity and widely expands the donor pool, thus allowing us to offer this curative therapy to many more patients with pid. chronic granulomatous disease (cgd) is a primary immune disorder that involves mutations in the nicotinamide adenine dinucleotides (nadph) oxidase complex (deffert, cachat, & krause, ) . two-third of cgd cases are caused by loss-of-function mutations in the cybb gene that encodes the gp pox subunit of the nadph. the increased in patients' life expectancy thanks to progress in diagnosis and management has underlined the burden of inflammatory manifestations occurring independently of infectious agents (dunogue et al., ; marciano et al., ) . cgd patients develop inflammatory granulomatous disorders, notably colitis, as a consequence of a dysregulated inflammasome activation. the treatment of inflammatory manifestations remains challenging, as it can be associated with an increased risk of infections. thus, understanding the pathophysiological mechanism of auto-inflammation in cgd could help improve the therapeutic arsenal for the management of these manifestations. to reveal the precise pathophysiological mechanism of auto-inflammation in cgd, we have developed a cellular model that reproduces the cgd phenotype in phagocytic cell. through crispr-cas gene-editing we generated a thp- c e l l l i n e h a r b o r i n g t h e p r e v i o u s l y d e s c r i b e d mu t a t i o n c. _ delccginsggt (p.tyr ter) in the cybb gene responsible for gp phox knock-out by early termination of translation. this cell line recapitulates the phenotype of cgd phagocytes: (i) decreased h o production (ii) and enhanced inflammatory responses after pma stimulation as evidenced by increased il- , il- and tnfa secretion levels (kuijpers & lutter, ) . these features were rescued by complementation through lentiviral transduction of a wild type cybb gene. this new model will help us to investigate the auto-inflammation reported in cgd patients and also to propose new therapeutic targets of inflammatory manifestations in this disorder. interleukin- (il- ) driven responses. children with irak- deficiency are predisposed to recurrent and invasive infections secondary to streptococcus pneumoniae, staphylococcus aureus and other pyogenic bacteria with high mortality rates in early childhood. the frequency and severity of infections is thought to decrease with age due to the acquisition of humoral immunity and immunologic memory, however due to the rarity of the disease, the natural history of this condition beyond early childhood is not well described. objectives: we present three unrelated irak- deficient patients with persistent chronic rhinosinusitis with nasal polyposis that developed in childhood. cases: patient is a y/o male with compound heterozygous mutations in irak (p.g afs* /c. - g>t) with a history of recurrent s. pneumoniae osteomyelitis (left hip at age and left knee at age ) and c. septicum sepsis at age following acute bowel perforation. additionally, he experienced recurrent aom during infancy and recurrent uti since age . despite prophylactic antibiotics and ivig, he has had recurrent polymicrobial (mrsa, s. pneumoniae, h. influenzae, p. aeruginosa, a. fumigatus) rhinosinusitis with nasal polyposis since age refractory to medical management requiring surgical intervention and prolonged courses of iv antibiotics. patient is an y/o female with homozygous deletions (exons - ) in irak with a history of ruptured appendicitis complicated by pseudomonas abscess and bacteremia at age , culturenegative sepsis with septic arthritis and osteomyelitis of the right leg at age , and septic shock secondary to mssa bacteremia complicated by rhabdomyolysis and dic at age . she has a history of chronic rhinosinusitis, and despite ivig and prophylactic antibiotics, she developed polymicrobial (h. influenzae, b. fragilis) rhinosinusitis with associated nasal polyposis pending surgical management. patient is a y/o female with homozygous mutations in irak (q x/q x on exon ) with a history of s. pneumoniae meningitis at months, m. catarrhalis epiglottitis and neck cellulitis at months, rsv bronchiolitis at months, enterococcus bacteremia at months, s. pneumoniae sepsis at age and streptococcus lymphadenitis at age . despite ivig and prophylactic antibiotics, she developed recurrent polymicrobial (h. influenzae, b. fragilis, mssa, v. cholera, p. aeruginosa, a. fumigatus) rhinosinusitis refractory to medical management requiring surgical intervention and iv antibiotics. conclusions: in our centers experience, irak- deficient patients continue to suffer from infectious complications, most prominently recurrent polymicrobial sinus infections beyond early childhood. the consistent presence of sinonasal polyps in these children is unusual, as it is not typically found in uncomplicated pediatric chronic rhinosinusitis. these infections have occurred despite antimicrobial prophylaxis and ivig, highlighting the role of irak- in sinopulmonary epithelium. additionally, the infectious organisms identified in our patient cohort are not commonly associated with irak- deficiency. further study of chronic rhinosinusitis and nasal polyposis in a larger cohort of irak- deficient patients and other innate immunodeficiencies may help identify pathways for targeted treatment of these patients. introduction: chronic granulomatous disease (cgd) is an inherited phagocytic defect associated with inability to clear catalase positive organisms. infections in patients with cgd are severe and recalcitrant. commonest infections are pulmonary followed by soft tissue infections and suppurative lymphadenitis. osteomyelitis is an uncommon infection in patients with cgd. it poses several diagnostic and therapeutic challenge. we herein report our experience of osteomyelitis in cgd over the last years. material and methods: review of records was carried out to describe the profile of osteomyelitis in cohort of patients with cgd at pediatric immunodeficiency clinic, advanced pediatrics centre, postgraduate institute of medical education and research, chandigarh, india. the diagnosis of cgd was based on nitroblue tetrazolium dye reduction test (nbt) and dihydrorhodamine reduction (dhr) assay. results: of the patients with cgd, ( . %) had osteomyelitis ( males and females; age range - years). most patients had their first episode of serious infection in early childhood (mean age: . years). stimulation index (si) of dhr assay ranged from to . . mutational analysis was done in / patients ( x-linked; autosomal recessive). site of involvement was variable ribs- ; vertebrae- ; radius- ; skull- ; tibia- . aspergillus fumigatus was the most common isolate ( %; / ); others had aspergillus flavus, aspergillus terreus and serratia marcescens each. all patients with rib osteomyelitis had concurrent pneumonia, and fungus was isolated in all of them (aspergillus fumigatus- , aspergillus flavus- , zygomyces spp.- ). antifungals (intravenous amphotericin b) were given for a duration of - weeks and were followed by oral voriconazole in therapeutic doses for to months in majority of them. debridement and resection of ribs was required in one patient, while other patients were managed conservatively. out of patients, ( %) succumbed to pneumonia and respiratory failure. conclusion: osteomyelitis in the context of cgd is usually caused by aspergillus spp. involvement of ribs and vertebra usually occurs with the contiguous spread of infection from the lungs. therapy often requires prolonged duration of anti-microbials, and may require surgical debridement in addition to it. a -year-old woman with history of hypogammaglobulinemia and acute liver failure a -year-old woman with a -month history of nausea, vomiting, and abdominal pain was admitted to an outside hospital with new onset of jaundice and anasarca. liver biopsy was thought most consistent with alcoholic steatohepatitis, and she was discharged with counseling on alcohol cessation and medical management of liver disease. she presented to our facility for a second opinion. over the following days, she developed further rise in direct hyperbilirubinemia up to . mg/dl, new coagulopathy with an inr . and hypoalbuminemia to . mg/ dl in the absence of ongoing alcohol consumption. liver sonography revealed course echotexture and patent vessels. pcrs directed against multiple hepatotropic viruses were negative and copper studies were normal. due to a history of moderate alcohol consumption, she was started on high-dose corticosteroids due to a presumptive diagnosis of alcoholic hepatitis. additional history raised concern for a possible primary immunodeficiency, including idiopathic thrombocytopenic purpura at years of age, multiple episodes of sinusitis treated with antibiotics and sinus surgery, one episode of suspected bacterial pneumonia, and one hospitalization for influenza a during which she developed neutropenia. in her s, she developed refractory genital warts, prompting infectious diseases evaluation. initial immune evaluation had revealed low immunoglobulins (iga < mg/dl, igg mg/dl, igm mg/dl) with very low responses to tetanus and diphtheria, despite a recent booster dose, and b and t cell lymphopenia (cd + cells/μl, cd + cells/μl, cd + v, cd + cells/μl, cd / + cells/μl); antigen and mitogen proliferation were not assessed. intravenous immunoglobulin replacement was initiated but discontinued by the patient due to infusion-related adverse effects, and she was lost to follow up until she presented with liver failure. both parents were deceased from cardiovascular disease in their s and she had no siblings. she had limited knowledge of family history but no known immune diseases. due to suspicion for genetic etiology of immune disorder and liver disease, we performed next-generation sequencing of a panel of over genes implicated in primary immune deficiencies. patient was heterozygous for a nucleotide substation (c. + g>a) within a splice site at the exon /intron boundary of the nfkb gene. during the hospitalization, immunoglobulin replacement and trimethoprim-sulfamethoxazole prophylaxis were initiated. an attempt was made to refer the patient for additional immunological evaluation and transplantation evaluation but unfortunately, she developed worsening liver failure and multiple complications, including extended-spectrum beta-lactamase (esbl)-producing e. coli bacteremia, hypotension requiring vasopressors and extensive bowel ischemia, and died in the hospital. in summary, this case highlights both the risk of diagnostic delay in adult patients presenting with a primary immune deficiency and potential for genetic testing to clarify the diagnosis. while the particular genetic change has not been described, other splice site and predicted loss-offunction mutations have been reported as pathogenic in this gene, which have been implicated in autosomal dominant common variable immunodeficiency. this case further expands on the genetic causes and spectrum of disease associated with changes in the nfkb gene. introduction: malnutrition and micronutrient deficiency are underrecognized causes of acquired immunodeficiency in adults, and may occur even in patients with high body mass index (bmi). methods: a -year-old woman with a medical history significant for one remote urinary tract infection presented to the emergency department after sudden onset of severe right flank pain. the pain was accompanied by urinary frequency and not relieved by ibuprofen; she denied fevers or chills. she was diagnosed with pyelonephritis and discharged on ciprofloxacin, which was later changed to trimethoprim-sulfamethoxazole after her culture grew resistant e. coli. her pain continued despite treatment, prompting her to return to the hospital three days later. upon presentation, she was afebrile with blood pressure of / mmhg and heart rate of bpm. her body mass index was . kg/m^ . her physical exam was otherwise notable for right costovertebral angle tenderness. laboratory studies revealed a leukocyte count of , /ul with % neutrophils; alkaline phosphatase of units/l and albumin of . g/dl, but otherwise normal liver function tests; normal lactic acid; and urinalysis with , wbc/hpf, rbc/hpf, moderate bacteria, and the presence of wbc clumps. ct scan of the abdomen and pelvis demonstrated an obstructing mm right renal stone with hydronephrosis and a right renal abscess contiguous with a right-sided hepatic abscess measuring . x . x . cm. she was treated with ceftriaxone and metronidazole, and underwent imaging-guided drainage of the abscesses. abscess cultures again grew resistant e. coli. she was discharged from the hospital with drains in place and a plan to continue trimethoprim-sulfamethoxazole until definitive management of her nephrolithiasis with ureteroscopy and lithotripsy. discussion: there remained the question of how an ostensibly immunocompetent patient had developed such severe intraabdominal infection with little systemic inflammatory response (e.g. no fever and only mild leukocytosis). a hiv antibody screen was negative. on further interview, she described a lb intentional weight loss over the preceding years, accomplished by dietary restriction to less than calories per day. nutritional assays revealed prealbumin, vitamin c, and vitamin b levels below the threshold of detection. she had low-normal b and b . out of concern for an acquired immunodeficiency resulting from malnutrition with micronutrient deficiency, balanced nutrition was discussed with the patient who agreed to liberalize her diet. background: the past decade has brought dozens of new mendelian disorders of immunity. yet, the genetic contribution(s) to diverse disorders of the immune system remain largely unelucidated. the majority of research participants referred to the national institute of allergy and infectious diseases (niaid) for what may be a mendelian disorder evade molecular diagnosis. making progress in this area requires a coordinated, systematic, and transparent approach to clinical genomics research which leverages the unique environment at the national institutes of health clinical center (nih cc). methods/design: this study is designed to systematically apply exome sequencing and related technologies with clinical grade interpretation and reporting to niaid research participants at the nih cc under a single protocol in order to facilitate research and clinical genetics care across niaid. we are recruiting approximately participants per year from approximately intramural clinical investigators. we generate genomic data, collect standardized phenotyping and report clinical interpretation in the medical record, all while providing linked genetic counseling. results: to date, we consented participants, we sent out samples for exome sequencing and samples underwent copy number variant analysis. we have completed analysis for families ( individuals) and finalized and resulted cases. here we present a case series illustrating some of our findings. case : a year-old female was referred to niaid for neonatal onset multisystem inflammatory disease (nomid). developmental delay and mild intellectual disability were appreciated on clinical evaluation. exome sequencing detected a mosaic novel likely pathogenic variant in nlrp . chromosomal microarray analysis (cma) showed ã mb interstitial deletion of chromosome previously associated with developmental delay and intellectual disability. case : a year-old ukrainian male was referred to niaid for the clinical diagnosis of autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy (apeced). exome sequencing and cma did not detect pathogenic variants in aire, but did find a de novo variant in fam b. defects in fam b are associated with poikiloderma with tendon contractures, myopathy, and pulmonary fibrosis (poiktmp). the clinical features of the patient were consistent with poikmp. case : a -year-old man had a history of brain, liver and kidney nocardiosis, disseminated mac infection, prostate cancer and lymphoma. family history was significant for prostate cancer. exome sequencing showed a heterozygous pathogenic variant in brca , associated with susceptibility to breast-ovarian, male breast, pancreatic and prostate cancer. conclusion: this case series illustrates that multiple diagnoses, unexpected diagnoses, secondary genomic findings, and data sharing helped identify variants in candidate genes. process standardization supports data integrity and efficiency while accommodating the need for investigator flexibility and providing tailored patient care. rationale: activated pi kinase delta syndrome (apds) is a primary immunodeficiency caused by dominant mutations that increase activity of phosphoinositide- -kinase (pi k). the catalytic subunit p is mainly expressed in cells of the hematopoietic system, primarily lymphocytes and myeloid cells, and mutations affect both b-and t-cells. we sought to further evaluate the role of the t-cell receptor (tcr) repertoire in immune dysregulation and the pathogenesis of autoimmunity and lymphoproliferation in patients with apds. methods: we evaluated the tcr repertoire in the peripheral blood in patients with pik cd mutations and compared these to the peripheral tcr repertoire in patients with common variable immunodeficiency (cvid) and healthy controls to investigate the role of the tcr in disease. the tcr repertoire in affected tissue of patients with pik cd mutations was also evaluated (tissue included lymph nodes for both patients, in addition to gastrointestinal tract and lung tissue in one patient). a fixed number of tcrs were subsampled ( , for blood and , for tissue) and diversity was calculated using the gini and shannon indexes. results: using the shannon and gini diversity indexes, the tcr repertoire in patients with pik cd mutations had less diversity/ increased clonality as compared to healthy controls and those with cvid ( figure ). for the two apds patients with biopsy tissue available for analysis, the diversity of the tcrs in tissue was increased as compared to the peripheral blood tcr repertoire ( figure ). conclusions: pi k plays an important role in the development and function of both b-and t-cells. patients with apds were found to have decreased tcr repertoire diversity in the circulating t-cell compartment compared to healthy controls and other cvid patients. the increased tcr diversity in the affected tissues compared to peripheral blood implicates the pi k/akt signaling pathway with t-cell trafficking and tissue immune homeostasis, and suggests this pathway may play a role in the development of inflammatory and lymphoproliferative complications in these patients. gain-of-function mutations in pi kd result in a human primary immunodeficiency, named apds (activated pi k-delta syndrome), characterized by lymphopenia, lymphoproliferation, respiratory infections and inefficient responses to vaccination. however, what promotes these immune disturbances at the cellular and molecular level remains unknown. we have recently published a mouse model that recapitulates major features of this disease and used this model and patient samples to probe how hyperactive pi kd fosters aberrant humoral immunity. we found that mutant pi kd alters the intrinsic function of t and b cells, leading to icos-independent increases in t follicular helper (tfh) and germinal center (gc) b cells, disorganized gcs, and poor class-switched antigen-specific responses to immunization. these phenotypes were associated with increased phosphorylation of akt and s in t and b cells, and lower threshold of activation, with altered regulation of foxo and bcl family members. moreover, b cells showed enhanced responsiveness and proliferation to both antigens and innate stimuli, accompanied by reduced cell death. strikingly, aberrant responses were accompanied by increased reactivity to gut bacteria, and a broad increase in autoantibodies that were dependent on commensal microbial stimulation, as demonstrated by striking reduction of self-reactivity upon antibiotic treatment in mutant mice. we now have further examined b cell function in these mice and demonstrate that altered foxo plays a major role in disruption of both b and t cell function. we further provide evidence for altered activation of metabolic pathways in b cells, compared to wt cells, that may contribute to the dysregulated b cell reactivity. our findings suggest that proper pi kd regulation is critical for ensuring optimal host-protective humoral immunity despite tonic stimulation from the commensal microbiome. this research was supported in part by the intramural research program of the nih, nhgri and niaid. autoimmune cytopenias are seen in a significant proportion of patients with immunodeficiencies affecting antibody production. previous b-cell maturation studies using fluorescence-activated cell sorting (facs) have associated various phenotypes of primary immunodeficiency diseases affecting antibody production with differing levels of b-cell differentiation. in this study we analyzed the peripheral b-cell compartment of patients with a hypogammaglobulinemia and > % b-cells with and without a history of autoimmune cytopenias. b-cells were isolated from peripheral blood using monoclonal anti-cd and these cells were gated to identify the proportion of memory b cell (cd +cd + ), igm+ memory b (cd +igm+), marginal zone b-cells (igm+ igd+), isotype-switched memory b-cells (cd +igm-igd-) and transitional cells (igmhicd hi). pid patients with a history of aic had decreased proportions of total cd + b-cell ( . % vs . %; p= . ) and igm memory b cells ( . % vs . %; p = . ). conversely, the proportion of marginal zone b-cells was increased in this group ( . % vs . %; p = . ). consistent with previous reporting, the proportion of isotype-switched memory b-cells was significantly lower in the aic group ( . % vs . %; p = . ). statistically significant inter-group difference was not seen within the transitional b-cell subset. our data suggest that maturation arrest of marginal zone (cd +igm+ igd+) b-cells may be implicated in the development of autoimmune cytopenias in humoral immunodeficiency. ( ) submission id# taissa de matos. kasahara , sudhir gupta, md phd student, state university of rio de janeiro and university of californis irvine professor, university of california at irvine, irvine, ca, usa introduction/background: common variable immunodeficiency (cvid) is the most frequent form of primary hypogammaglobulinemia with decreased serum igg and iga levels and variable levels of igm in adults. in addition to decreased serum immunoglobulins, - % of cvid patients present autoimmune manifestations. the mechanisms that lead to a breakdown of selftolerance in cvid are not completely understood. however some differences in b and t cells subsets and autoreactive b and t cells can be detected. elevated expression of surface igd and downregulation of igm receptor are hallmarks of anergic naïve b cells that contain autoreactive receptors in human peripheral blood. moreover, memory b cells that have class switched to igd and present an igd+igm-phenotype are also highly reactive to self-antigens in healthy individuals. the role of these autoreactive naïve and memory b cells in the immunopathogenesis of cvid has not been evaluated. here we investigated the frequency of cd -and cd + b cells expressing igd and igm in peripheral blood of cvid patients. methods: peripheral blood mononuclear cells (pbmc) from cvid patients (n= ) and health subjects (n= ) were separated by ficollhypaque and incubated with anti-human cd -percp, cd -fitc, igd-bv and igm-apc to identify different subsets of b cells by flow cytometry. cd +cd -igd+igm-and cd +cd -igd+ igm+ b cells were sorted, loaded with cfse and cultured with cpg and ant-cd for days to evaluate the proliferation. results: among the compartment of cd -b cells, cvid patients showed an increased frequency of igd+igm+ cells and a lower frequency of igd-igm-cells as compared to control group. no differences were observed in the frequency of igd+igm-cells in cd -b cells between cvid patients and controls. in contrast, in the compartment of cd + b cells, cvid patients showed an increased frequency of igd+igm-, igd+ igm+ and igd-igm+ cells and a lower frequency of igd-igm-cells when compared to health subjects. when the patients were divided in two groups based on autoimmune manifestations, the group with autoimmune disease showed an increased frequency of igd+igm+ and igd-igm+ cells in cd -b cells when compared to the control groups. both patient groups showed an increased frequency of igd+igm-, igd+igm+ and igd-igm+ cells and a lower frequency of igd-igm-cells when compared to health subjects. regarding the proliferation, naïve b cells from cvid patients showed a reduced proliferative capacity in response to in vitro stimulation as compared with naïve b cells from health subjects. conclusion: our results suggest that the increase of cd +igd+igm-b cells can be related to the susceptibility of autoimmunity in cvid patients. introduction: immunoglobulin g -related disease (igg -rd) is a group of immune-mediated conditions where tissues are affected with dense lymphoplasmacytic infiltrations with a predominance of igg -positive plasma cells and storiform fibrosis, usually in the setting of elevated serum concentrations of igg . common presentations include autoimmune pancreatitis, sclerosing cholangitis, retroperitoneal fibrosis, salivary gland disease, and orbital disease, among others. symptoms of asthma or allergy are present in approximately percent of patients and they typically exhibit a good initial therapeutic response to glucocorticoids. case presentation: a -year-old female with a history of gastroparesis, cutaneous lupus erythematosus and suspected autoimmune pancreatitis was referred to allergy/immunology clinic for evaluation of elevated igg . she reported a -year history of recurrent abdominal pain attributed to recurrent pancreatitis based on previous mild lipase elevations. prior endoscopic ultrasound (eus) of the pancreas revealed edema. there was concern for gallstone pancreatitis but ercp followed by cholecystectomy, biliary and pancreatic sphincterotomy had no change in her symptoms. in , she was noted to have a positive ana and high serum igg , per patient (values from osh records could not be obtained). symptoms improved with a course of steroids, hence suspicion for autoimmune pancreatitis. in she developed a rash on her arms and face. biopsies of the affected areas revealed cutaneous lupus erythematosus on the arms and a basal cell carcinoma on the face, which was excised. ana was only : at that time. at the visit, she complained of severe allergic rhinitis, joint pains, as well as a malar rash, which responded to intermittent courses of prednisone by prior providers. laboratories obtained at initial visit were significant for thrombocytopenia ( thou/cu mm), positive lupus anticoagulant ( sec) and elevated igg ( mg/dl; normal range - mg/dl). c , c , c q, ana, anti-double stranded dna, anti-smith antibodies, antiphospholipid panel, upep and spep were all unremarkable. ct chest and abdomen were also normal. given the patient's history of cutaneous lupus erythematosus, plaquenil was started as a steroid sparing agent. eus of the pancreas with possible biopsy was ordered in an attempt to obtain a histopathologic diagnosis of igg -rd. conclusion: this case exhibits the association between elevated igg , pancreatitis of unknown origin, allergic rhinitis, and cutaneous lupus erythematosus, highlighting the value of identifying a pathologic connection between seemingly unrelated disorders in patients with elevated igg , as they may be manifestations of igg -rd. in order to make the diagnosis, histopathologic findings showcasing lymphoplasmacytic tissue infiltration consisting mainly of igg -positive plasma cells and small lymphocytes is essential. the majority of patients respond to glucocorticoids, and while the duration of response is variable, most patients flare during or after glucocorticoids are tapered, as noted in this patient. rituximab has been shown to be effective in some patients and will be considered in this patient if symptoms persist. ( ) submission id# rationale: pnp deficiency is an autosomal recessive disorder due to defective purine metabolism leading to severe combined immunodeficiency (scid) and neurological deterioration. newborn screening utilizing t-cell receptor excision circle (trec) assay can detect affected patients before complications arise. herein, we describe an infant initially identified by newborn screening with pnp deficiency and congenital cmv, a previously unreported presentation. methods: cmv quantitative pcr (qpcr) was performed by nebraska medicine, pnp enzyme activity by duke and genetic sequencing by invitae. results: a small for gestational age (sga) male infant was reported to have an abnormal trec assay on day of life (dol) . he was hospitalized for further evaluation. initial studies revealed profound lymphopenia, normal lymphocyte proliferation to mitogens and no evidence of maternal engraftment. additionally on dol , he had cmv viremia and viruria; thus with sga, failed unilateral hearing screen and head ultrasound with bilateral parenchymal calcifications, congenital cmv was suspected. pnp enzyme activity was abnormal. cmv treatment was initiated with ganciclovir on dol . foscarnet was added on dol . cmv qpcr levels decreased below the limit of detection by dol . genetic testing found a pathogenic homozygous mutation in pnp (c. - g>a). the infant has a / hla-matched, unaffected, cmv positive sibling and will proceed to hematopoietic stem cell transplantation. conclusions: to our knowledge, this is the first reported case of pnp deficiency identified through newborn screening. this novel case of congenital cmv and pnp deficiency highlights the importance of cmv screening and need for treatment strategies for congenital cmv in scid. despite a dramatic increase in the use of next generation sequencing over the last decade, the majority of the more than million identified human genomic variants do not have well-established clinical implications. progress is being made on this complex challenge through multiple approaches, including data sharing. to maximize our understanding of genomic data, platforms that enable effective and responsible data-sharing are essential. this means that genotypic and phenotypic data must be findable, accessible, interoperable, and reusable under conditions that are ethical and transparent. to highlight innovations in data-sharing and their potential to advance discovery, we present three data-sharing mechanisms. for each platform, we will present a case highlighting its key functionality and discuss opportunities and challenges that may arise as each platform is scaled up. ( .) genomic research integration system (gris) is a collaborationengendering web application that facilitates the identification of genetic variants associated with rare immunological disorders. users can access integrated and standardized phenotypic and genomic data that is analyzable within the platform. gris enables systematic and automated capturing, and links patient data from disconnected systems and paperbased records. standardized annotations allow for the comparison of data from different clinical studies. the main goal of this tool is discoverability of other affected individuals enrolled in separate protocols within the niaid intramural research program. this internal database was used to find a second family with a rare variant in a candidate gene. ( .) the genomic ascertainment cohort (tgac) is a resource that aims to improve our understanding of the phenotypic consequence of genetic variation by providing access to aggregate, de-identified genomic data from large nih intramural and related cohorts. participants have provided informed consent to be re-contacted for additional phenotyping in the future. the main goal of this tool is to enable further study of the clinical consequence of variants in a large, unbiased cohort of patients ascertained for many indications. this database was used to investigate findings in participants with previously published pathogenic variants in genes associated with primary immune deficiency based on medical record review. ( .) clingen is dedicated to building an authoritative central resource that defines the clinical relevance of genes and variants for precision medicine and research. through the sharing of genetic and health data, clingen seeks to answer whether a given gene is associated with a disease (clinical validity)?; whether a given variant is causative (pathogenicity)?; and whether the information is actionable (clinical utility)? this resource is meant to convene disease-and gene-specific expert groups to curate the medical literature on mendelian disease to better define gene-disease and variant-disease relationships using many lines of evidence. this resource was used to clarify clinical validity of disease-gene assertions. together these efforts help create a clinical research ecosystem that maximizes the value of clinical research data and ultimately improves patient care. this research was supported by the intramural research program of the nih, niaid. introduction: according to the population reference bureau, the number of elderly americans, defined as age and older, is projected to more than double from million to million by , rising from % to % of the total population. the impact of immunodeficiency in this important segment of the population remains understudied. methods: the usidnet registry was queried to obtain demographic, clinical data of elderly patients defined as age and older. descriptive analyses were performed on the data. results: participants ( . %) were eligible out of total registry participants. the median age of the cohort was years and predominantly female ( . %) and white ( . %) with a median bmi of . ± . .the majority ( . %) of subjects were living. humoral deficiencies comprised the majority of diagnoses ( . %), with common variable immune deficiency being the most frequent ( . %). of the remaining non-humoral diagnoses, immune dysregulation ( . %) and immunodeficiency with myelodysplasia ( . %) were the most frequent. the majority ( . %) of subjects reported having received immunoglobulin replacement therapy (igrt) at some point, with . % reporting via iv route. of the infections that occurred in this cohort, sinopulmonary infections were the most commonly reported, specifically sinusitis ( . %), pneumonia ( . %), upper respiratory infection ( . %), and otitis media ( . %). in this cohort, autoimmune, cardiovascular, and granulomatous complications were reported . the number of patients with malignancy was , with some patients diagnosed with multiple malignant disorders. of the reported malignancies, the majority ( . %) were solid tumors. conclusions: compared to the age-matched non-immunodeficiency united states population, this cohort had more females . % (usidnet) versus . % (us population) and fewer whites . % (usidnet) vs . % (us population. humoral immunodeficiencies, specifically cvid, were most common diagnoses, similar to other age groups of immunodeficiency patients. majority of these patients have received igrt, with approximately half via iv route. this cohort reported living with a variety of non-infectious complications, including autoimmunity and malignancies. more research which specifically focuses on elderly patients with immunodeficiency is needed. clinical microbiologist and infectious disease physician, university of calgary x-linked agammaglobulinemia (xla) is a primary immunodeficiency caused by mutations in the bruton tyrosine kinase gene which leads to b cell maturation failure and defective antibody production. this puts patients at risk of recurrent sinopulmonary infections, gastrointestinal infections, and recurrent skin infections including infections caused by helicobacter sp. helicobacter sp are gram negative bacilli commonly found in the gastrointestinal tract of various animals. helicobacter sp. have been linked with gastritis most notably helicobacter pylori causing gastric ulcers in humans. helicobacter sp. has been found in rare cases to cause disseminated infections including pyodermic gangrenosum and cellulitis notably in patients with agammaglobulinemia. infections caused by helicobacter bilis are challenging to diagnosis due to difficulties with culturing the pathogen as well as poor guidelines for antimicrobial management. case report: the patient was diagnosed with x-linked agammaglobulinemia at the age of months with a history of recurrent sinusitis and was started on ivig q weeks. despite regular ivig, he developed bronchiectasis. at years of age in , he developed a chronic rash around his left knee resembling erythema nodosum. by , he had developed a left knee effusion associated with left sided calf pain. his knee pain was found to improve during courses of ciprofloxacin to treat recurrent lung infections. given case report data of h. pylori causing erythema nodosum in patients with agammaglobulinemia, he was treated empirically for an h. pylori infections with no improvement. in he was found to have progressive cellulitis with pyomyositis of the left leg. a skin biopsy of a calf nodule was found to be culture negative but s pcr was positive for h. bilis. he was started on treatment with ertapenem and levofloxacin with subsequent resolution of his rash. his left ankle pain progressed and by late and was found to have possible osteomyelitis of the left ankle on mri. in he was found to be bacteremic with h bilis. due to progressive symptoms with significant impact on function and rising inflammatory markers despite months of antimicrobial treatment, doxycycline and flagyl were added leading to clinical improvement and normalization of his inflammatory markers. he was continued on oral doxycycline and flagyl for months for a chronic osteomyelitis. discussion: h. bilis is a slow growing pathogen which is challenging to culture in the laboratory often requiring special agar plates and prolonged incubation. in patients with agammaglobulinemia and associated chronic skin infections or erythema nodosuma, h bilis should be suspected as a possible pathogen. due to challenges with culturing, s pcr or amplification of the s ribosomal subunit should be considered to try to identify the pathogen. there are poorly delineated clinical antimicrobial breakpoints to help guide therapy with minimal evidence. case reports suggest prolonged therapy with aminoglycosides and penicillin. other studies have successfully treated patients with a carbapenem, azithromycin and levofloxacin. in the absence of sensitivity data, prolonged treatment ( months) should be considered with a combination of antimicrobials. patients should be followed closely as recurrent infections are not uncommon. chief, human immunological diseases section, laboratory of clinical immunology and microbiology, niaid, nih, bethesda, md introduction: dock deficiency is a combined immunodeficiency characterized by eczema, recurrent sinopulmonary infections, viral skin infections, malignancy and early mortality. in recent years, liver disease and vasculopathy have been increasingly recognized as a complication of dock deficiency. we clinically characterized our cohort of dock deficient patients, with a specific focus on these newly identified areas of disease involvement. methods: chart reviews were performed on patients seen at nih with genetic and clinical diagnosis of dock deficiency. patients were all enrolled on irb approved niaid protocols. results: we identified patients from families with dock deficiency in our nih cohort, ranging in age from - years. of the families, had homozygous mutations. of the patients, food allergy was diagnosed in ( %), eczema in ( %), and asthma in ( %). chronic or recurrent viral skin infections were seen in / ( %). chronic ebv viremia by pcr positivity was seen in / patients ( %); only patients were known to be ebv immune without viremia. cmv viremia was infrequent. sinopulmonary infections were common, with bronchiectasis occurring in / ( %) with available imaging. liver disease was diagnosed in ( %), with having biliary tract abnormalities on imaging and stool positive for cryptosporidia; most patients with cryptosporidia were without diarrhea. the incidence of cryptosporidia is likely under-represented due to more recent availability of sensitive assays for cryptosporidia detection. other liver abnormalities included fatty liver, metastatic disease from malignancy and medication related hepatitis. vasculopathy, predominantly of the aorta and cerebral arteries, was diagnosed in , with patients in the last years being prospectively imaged. autoimmunity was rare ( %) including autoimmune cytopenias and hypothyroidism. of with follow-up are alive ( %) with age range - years. of the living patients, ( %) have had a hsct. causes of deaths include malignancy ( ), infection ( ) , and hsct complications ( ) . long-term follow-up of patients with hsct (up to years) has revealed resolution of the infection susceptibility and eczema, no new cancers, and stabilization of vasculopathy. conclusions: in addition to the well described manifestations of dock deficiency including eczema, allergy, recurrent sinopulmonary infections, skin viral infections and malignancy, our cohort revealed a relatively high incidence of liver disease, frequently associated with stool positivity for cryptosporidia, as well as vasculopathy. both of these clinical manifestations should be considered during preparation for hsct as they may affect management through transplant. autoimmunity has likely been over-estimated in prior descriptions of dock deficiency. long-term follow-up after hsct is needed to determine the prognosis from the vasculopathy, liver disease, and malignancy risk. ( ) submission id# yasuhiro yamazaki , stefano volpi , luigi d. notarangelo introduction/background: extl (exostosin like glycosyltransferase ) is an exostosin family member which initiates heparan sulfate (hs) chain biosynthesis and elongation. we have reported homozygous extl hypomorphic mutation (r w) as a cause of immunoosseous-dysplasia syndrome. fourteen patients who have extl homozygous mutation were reported so far. eight of them manifested t cell lymphopenia, and presented with severe combined immunodeficiency (scid) or omenn syndrome. using patient-derived induced pluripotent stem cells (ipscs) as a model, we have previously reported that extl mutations affect differentiation to thymic epithelial progenitor cells as well as expansion of hematopoietic progenitor cells. consistent with the latter, previous studies have suggested that mutations in other genes involved in hs biosynthesis affect hematopoietic stem cell (hsc) differentiation. however, the exact mechanisms by which extl mutations affect hematopoiesis are not known. objectives: we tried to clarify gene expression difference in hscs derived from wild-type, extl hypomorphic and extl knock-out (ko) human ipscs. methods: the control bj ipsc line was engineered by crispr/cas gene targeting. extl ko ipscs were obtained which carried compound heterozygous extl mutations (c. _ inst; c. _ insgatattt). hsc differentiation was induced using the stemdiff hematopoietic kit (stemcell technologies). bulk rna from each ips cells and each differentiated cd +cd +cd + was analyzed by rna sequencing. results: as compared to control ipscs, patient-derived cells showed slightly lower capacity to generate cd +cd +cd + cells. on the other hand, extl ko cells showed no differentiation into cd + cd +cd + cells. gene set enrichment analysis showed enriched expression of genes involved in hematopoietic progenitor cell differentiation, regulation of hemopoiesis, and positive regulation of hemopoiesis in both control and patient-derived cd +cd +cd + cells compared to parental ipscs. moreover, these gene sets were more abundantly enriched in control than in patient-derived cd +cd +cd + cells. the gene set of response to type i interferon was significantly enriched in control versus patient-derived cd +cd +cd + cells. conclusions: these results confirm that extl plays an important role for hsc homeostasis in human cells. because type interferons play a role in hsc proliferation, the decreased type i interferon signature may account for the reduced number of hscs that we have previously reported upon in vitro differentiation of extl -mutated versus control-derived ipscs. this study was supported by the division of intramural research, niaid, nih, under protocol -i-n . a case of autoinflammatory syndrome with osteoporosis and specific antibody deficiency autoinflammatory syndromes are inherited disorders with an exaggerated inflammatory response with no specific trigger. the clinical phenotypes of variants of autoinflammatory syndromes may overlap. we report a case of a year old male with prior diagnosis of specific antibody deficiency, periodic fever, aphthous stomatitis, pharyngitis, cervical adenitis (pfapa) syndrome, arthralgia and moderate atopic dermatitis. he was diagnosed at years of age with specific antibody deficiency based on persistently low pneumococcal titers against repeat immunizations. due to recurrent infections, he was placed on immunoglobulin replacement therapy (igrt) at years of age. igrt was discontinued at years of age due to full resolution in infections and patient demonstrated robust response to immunizations. patient had lifelong history of recurrent fevers (every weeks) associated with pharyngitis and aphthous ulcers consistent with diagnosis of pfapa. as he became older these episodes became less frequent. last episode of fever was over a year ago. the father had similar symptoms of recurrent fevers and oral ulcers as a child but currently remains asymptomatic. paternal grandfather died of kidney disease. patient has been generally in good health until recent year with intermittent abdominal pain, arthralgia and several long bone fractures with no history of prior trauma. a bone density scan revealed osteopenia and osteoporosis with a z score of - . of lumbar spine, - . of left femoral neck, - . of left hip. given history of familial autoinflammatory disease, and antibody deficiency genetic testing was obtained which identified a pathogenic heterozygous variant of taci and mefv c. g>a (p.met lle). taci mutation has been linked to antibody deficiency syndromes. genetic study for family members is pending. the mefv gene is associated with autosomal recessive familial mediterranean fever (fmf) and has been reported in autosomal dominant fmf as well. fmf is characterized by recurrent episodes of fever associated with serositis, arthralgia, and arthritis. patients with fmf have elevation in acute phase reactants during attacks with most returning to normal levels during the episode-free periods. multiple studies have shown that patient with fmf have lower bone mineral density and zscores than the general population. inflammation in fmf is thought to be mediated by several different cytokines (il- , il- , il- , il- , il- , il- , il- and tnf-). these same cytokines play a role in osteoclast activity and bone resorption. it has been suggested chronic inflammation during acute attacks and subclinical inflammation during the disease-free period lead to bone loss and osteoporosis. regular use of colchicine, the main treatment for fmf, may slow down osteoporosis. beside careful monitoring of clinical and laboratory phenotype, genetic evaluation is an important step in distinguishing between overlapping entities and can prevent complication and promote targeted intervention. a year old previously healthy boy was referred for periodic fever/ pfapa and mosquito bite hypersensitivity. eight weeks earlier he developed fever to f, mouth sores and exudative tonsillitis; a rapid strep screen was negative. one week later he developed moderate cervical lymphadenopathy and had a positive ebv early antigen antibody.. one month later he had several severe local reactions to mosquito bites. each manifested - cm of erythema and induration with a + cm bullae which left an ulcer after rupture and healed with a hypopigmented scar. the bites were accompanied by fever to f for days. one febrile episode was treated with low dose prednisolone for presumed pfapa, and the fever resolved within hours. his past history was positive for nasal allergy and mild asthma. his parents are not related: mom is of european-indonesian and dad european-african (creole ancestry. testing prior to this visit showed normal igg, iga and igm, elevated ige ( , u/l) and normal cbc. lymphocyte subsets revealed cd + % ( /mcl), cd + % ( /mcl), cd + % ( / mcl), cd + % ( /mcl), nk cells % ( /mcl). on examination he appeared well with height at th%ile and weight at th%ile. there was no lymphadenopathy, hepatosplenomegaly or inflammed skin lesions; there was a cm round scar on the right plantar surface at the site of a prior mosquito bite. laboratory studies confirmed nk lymphocytosis % ( /mcl) and elevated ige ( , u/l). lymphoproliferation to mitogens, cd /cd , cmvand hsv were normal, but absent to tetanus and candida antigens. ebv antibodies reflected past infection (vca-igg+, vca-igm-, ebna+); quantitative ebv pcr was > , , copies/ml whole blood. nk cytotoxicity and cd a expression were decreased. bone marrow nk analysis suggested conality. the patient was diagnosed with "hypersensitivity to mosquito bites with ebv-associated t-/ nk lymphoproliferation." this disorder represents a subset of chronic active ebv (caebv) that is rarely seen outside of east asia. the lack of organomegaly or lymphadenopathy with hyper-ige and nk lymphocytosis and decreased nk function support the likelihood that nk cells are the target of ebv infection in this patient. this diagnosis may be a precursor to hemophagocytosis, liver necrosis or lymphoma/leukemia, and the only curative treatment is bone marrow transplantation. the patient's sister is a / hla match. she is seropositive for past ebv infection, and she has no history of extreme reactions to mosquito bites. genetic mutations that cause familial hemophagocytic lymphohistiocytosis have not been reported in caebv, and to the best of our knowledge familial cases of this disorder have not been identified. the response to bmt in this patient is pending. introduction/background: a number of case reports have described symptomatic hypogammaglobulinemia following administration of anti-epileptic drugs (aeds), specifically lamotrigine, carbamazepine, and levetiracetam. the mechanism by which symptomatic hypogammaglobulinemia develops is unclear. we evaluated the prevalence and the clinical significance of hypogammaglobulinemia associated with use of these aeds. objectives: our aim was to characterize the prevalence of aed-induced hypogammaglobulinemia, identify specific aeds associated with hypogammaglobulinemia, and characterize the timeline to development of hypogammaglobulinemia after initiation of therapy. methods: a retrospective, multicenter, electronic medical record review spanning years identified patients with hypogammaglobulinemia who were on aed therapy (lamotrigine, carbamazepine, or levetiracetam). patients were excluded if they had a pre-existing primary immunodeficiency (pid), malignancy, protein-losing enteropathy, or significant proteinuria. patients on chronic immunosuppressive therapy, those without laboratory criteria for hypogammaglobulinemia, or those on one of the aeds for less than one month were also excluded. results: of the cases reviewed, patients met our inclusion criteria. the median age was ; % were adults, % were female, and % were white. lamotrigine was implicated in / of the cases, carbamazepine in / , and levetiracetam in / . tetanus and pneumococcal titers were available for / patients. of those patients, / had protective titers to both per report with responses to > % of the serotypes. only one patient reported severe, recurrent infections while the remaining four had little to no symptoms. interestingly, the patient with severe infections did have protective titers. of the five laboratory proven hypogammaglobulinemia patients, one died of an infection, two have continued on the medication due to refractory seizures responsive only to these medications, and two are currently being tapered off of their aed. conclusion: while it appears that aed-induced hypogammaglobulinemia is quite rare, it should be considered in a patient without other secondary causes of hypogammaglobulinemia on aed therapy. many antiepileptics downregulate nfkb signaling suggestive that patients who develop symptomatic hypogammaglobulinemia may have hypomorphic mutations in the nfkb signaling pathway. ( ) submission id# autoimmune lymphoproliferative syndrome (alps) results from defective apoptosis of lymphocytes mediated through the fas/fas ligand (fasl) pathway. the hallmark lab finding is an expansion of t cells that express the alpha/beta t cell receptor, but lack both cd and cd (double negative t cells) in the setting of normal or elevated lymphocyte counts. patients present with chronic, nonmalignant, noninfectious lymphadenopathy or splenomegaly. for definitive diagnosis, patients need to have ( ) a pathogenic mutation in fas, fas ligand or caspase or ( ) a defective fas-induced lymphocyte apoptosis. we describe a probable case of alps with heterozygous mutation in fas c. a>g(p.his arg), a variant that has not been previously reported (his lymphocyte apoptosis assay is pending). unique to this case is the patients castleman disease-like features on pathology. a year-old male referred from hematology clinic presented with an year history of chronic lymphadenopathy, splenomegaly, anemia, and no underlying diagnosis. malignancy had previously been excluded by bone marrow aspirate and biopsy years prior. however, he had a right sided lymph node that had increased in size for the past months. he was otherwise asymptomatic. a lymph node biopsy years prior was reportedly normal. his exam demonstrated significant bilateral lymphadenopathy, greater on right, with an approximately x cm mobile right neck mass. he had splenomegaly palpated cm down and across to midline. he was therefore admitted for excisional lymph node biopsy to evaluate for possible malignancy and labs were sent to evaluate for alps. labs were supportive of alps. he had elevated t cell receptor alpha beta double negative t cells (tcr a/b dntcs) in blood ( . %). b level was elevated (> pg/ml). plasma soluble fasl level was elevated ( pg/ml). interleukin- (il- ) and il- levels were elevated ( and pg/ml respectively). he had multilineage cytopenias: anemia with hgb of . g/dl and neutropenia (absolute neutrophil count of k/ul). he had hypergammaglobulinemia with an igg level of mg/dl. broad infectious work-up was negative, including hiv, quantiferon, cocci, bartonella, toxoplasma, coxiella burnetii, ebv pcr and, cmv igm. lymph node biopsy showed no evidence of malignancy. immunostains and flow cytometry showed the presence of expanded tcr a/b dntcs in the lymph node, consistent with alps. interestingly, lymph node histology showed morphologic features typical of plasma cell variant castleman disease. numerous castlemanlike follicles showed typical regressive changes with onion-skinning morphology. paracortical hyperplasia with sheets of plasma cells was noted. there was negative staining for hhv (a well-known cause of plasma cell variant castleman disease). the diagnosis of idiopathic multicentric hhv -negative castleman disease was excluded by definition in the setting of alps, per evidence-based consensus criteria published in . in addition, our patient did not show any symptoms typically associated with it, such as fever, night sweats, weight loss, weakness or fatigue. should his fas-induced lymphocyte apoptosis be defective (in separate assays), this would confirm his alps-fas diagnosis and we would start the patient on sirolimus. head of immunology unit, children' s hospital ricardo gutierrez introduction: slc a gene encodes the proto-couple folate transporter (pcft), which supports intestinal folate uptake, and participates in folate transport into the central nervous system. slc a mutations cause pcft defects, resulting in low folate levels in serum and cerebrospinal fluid. hereditary folate malabsorption (hfm) is a rare, autosomal recessive disorder with pcft deficiency resulting in cerebral folate deficiency. most of the patients present megaloblastic anaemia, moderate pancytopenia in the first few months of life, failure to thrive, diarrhoea and/or later onset neurological symptoms including seizures and developmental delay. i m m u n o d e f i c i e n c y i n h f m c a n m a n i f e s t i t s e l f w i t h hypogammaglobulinemia with normal t-cell function. b-cell precursor compartment seems to be particularly vulnerable to folate deficiency in some hfm patients. this immunodeficiency can be restored with specific treatment with folic acid. aim: to describe a female patient with a homozygous pathological variation in the slc a gene. results: a months old girl, born of non-consanguineous parents. she started at months old with diarrhoea due to rotavirus, low weight and bicytopenia with normal bone marrow aspiration. she presented low levels of folic acid . ng/ml (nv . - . ng/ml) at first thought due to secondary to malnutrition. treatment with folic acid supplementation was administrated, improving platelets counts. at months old she presented steatorrhea with severe perianal panniculitis which required surgical treatment. no germs were rescued after a skin biopsy. moreover, she suffered from a respiratory infection due to picornavirus with two episodes of pneumothorax which required intensive care. at that moment ivig treatment was administered due to hypogammaglobulinemia and clinical severity. chronic diarrhoea worsened with bloody depositions. three rectal ulcers were found in the gut biopsy. bowel inflammatory disease was suspected and mesalazine administration was started with weight improvement. furthermore, at months old she presented status epilepticus, with pathological eeg and normal mri; one of them related to a cmv infection, successfully treated. in the immunological evaluation igg and iga were low with normal igm and igd. the protein-antibody response was not evaluated. she presented normal lymphocyte and t cells extended populations, t cells proliferation assay, dhr, treg cells, complement, cd a expression, alpha-fetoprotein, without autoantibodies a molecular panel testing was done by ngs and a homozygous variant in slc a gene was found, causing impaired intestinal folate absorption. conclusion: hfm should be considered in the diagnosis of patients with cytopenias and hypogammaglobulinemia in order to provide specific treatment. hfm has wide clinical manifestations, not only with megaloblastic anaemia and neurological impairment but also with gastrointestinal and skin manifestations. with folate treatment, clinical and immunological defects can be normalized. introduction: multifocal epithelial hyperplasia (meh), or hecks disease, is a rare, benign infection of the mucosa caused by human papilloma virus (hpv). clinically, meh manifests as numerous painless, soft, sessile papules or plaques, and typically occurs in the labial, lingual, and buccal mucosa. meh lesions are usually associated with hpv types and , and seen more commonly in patients of caribbean or central/south american descent. prior studies in adults have shown that tumor necrosis factor alpha (tnf) promotes hpv, and may influence duration of hpv infection. case: we present a five-year-old full term male of haitian descent referred for assessment of multiple flesh colored, papular lesions on the buccal and labial mucosa that had persisted and quantitatively increased over one year, although some lesions regressed. he had no pain or difficulty eating. medical history significant for one seizure; negative for infection. no family history of infection, immunodeficiency, consanguinity, or miscarriage. head and neck examination failed to reveal cervical lymphadenopathy, masses, or hypertrophy in the salivary glands. intraoral examination revealed multiple papular nodules, mostly flat although some were corrugated. the greatest concentration was noted on the lower left labial surface extending to the mucosal vermillion interface, not involving the vermillion or commissure region. lesions extended into the mandibular vestibule and the left buccal mucosa. no other lesions were noted on extremities, genitalia, or any other visualized mucosal surface. based on history and exam, he was diagnosed with meh. white blood cell count, neutrophils, lymphocytes, cd and cd t cell, b cell, nk cell enumeration, and immunoglobulin panel were normal for age. tetanus and streptococcus pneumoniae titers were protective. cytomegalovirus igg and igm were negative. epstein-barr virus igg was positive, igm and early antigen ab negative. serology was significant for elevated tnf ( pg/ml; reference range < pg/ml) while interferon gamma and interleukins , , , , , , , , , and were normal, as was il- receptor cd . one month after the initial visit, lesions were stable and unchanged. nine-valent hpv vaccination was considered, but not administered. conclusions: meh is a rare but benign disease caused by hpv. awareness of the disease and its course is important to prevent unnecessary expanded immunodeficiency work-up and possible procedures to eliminate lesions. although mucosal immunity can be site specific, especially with hpv, our understanding of t-cell cytokine and chemokine responses to hpv in cervical and laryngeal lesions may be instructive. the mechanism which allows hpv persistence in meh is not characterized, but it likely is due to increased viral persistence and an inability for the host immune response to successfully induce viral latency and successful containment. elevated tnf levels, with normal levels of il- , il- , il- , il- , may correlate with decreased clearance of hpv and prolonged duration of meh. it remains unclear if viral persistence is the cause of, or the sequela of, increased tnf. longitudinal monitoring of cytokine (tnf, il- , il- , il- , il- ) and chemokine (ccl , ccl , ccl , ccl , ccl , and ccl ) serum concentrations may be useful biomarkers for disease resolution. introduction: autosomal dominant hyper ige (jobs) syndrome is a rare primary immunodeficiency characterized by eczema and sinopulmonary infections as well as musculoskeletal and vascular complications. as in all chronic illnesses, patient education is an ongoing need. in the rare disease population, patient education is especially important as patients must be able to explain their unique healthcare concerns in a variety of medical settings. we focused on ad-hies, due to our relatively large cohort of patients, the frequent lack of classic signs of illness often impairing diagnosis of severe infection, and the diverse nonimmunologic clinical features of this disease. objectives: we aimed to increase understanding of the clinical manifestations of ad-hies to promote earlier recognition of symptoms and to increase self-efficacy for symptom management in the adult hies population. methods: adult patients were asked to participate in a patient education project. demographic information was collected from participants. they also completed a -item multiple choice test about symptom recognition in ad-hies and promis self-efficacy for managing symptoms, an item validated survey. then, patient education handouts that focused on pulmonary symptoms, eczema, bone health, and cardiovascular complications were reviewed with the participant. six weeks later, participants were asked to repeat the -item test and the self-efficacy survey. the demographic information, test, and self-efficacy were collected anonymously. results: participants provided demographic information, completed the test and the self-efficacy survey. of the participants, were male and were female. participants ranged in age from to years. / ( %) reported looking for information about ad-hies using search engines and most patients ( %) report that they have been given information about ad-hies from a doctor. / ( %) participants identified pulmonary symptoms as the symptom that concerns them most and / ( %) participants identified more than one symptom of concern. participants returned the second test and second survey. the mean test score increased from . to . with / participants achieving a score of / or higher. the self-efficacy scores were unchanged with a mean score of . before reviewing the patient education handouts and . after. conclusions: participant feedback to this project was generally positive. ad-hies patients are seeking information and an educational intervention can improve their understanding of disease. self-efficacy results were mixed and unchanged overall, but suggest that ad-hies patients manage symptoms as well as other patients with chronic illnesses. patient education should continue at each encounter. this project can be expanded to include more topics, pediatric patients, and other rare disease populations. funded by the nci contract no. introduction: bcl b plays an important role in the development and maintenance of the immune system and the central nervous system. expression of bcl b represses nk and myeloid factors while inducing t cell lineage genes in thymocytes at the dn stage. conditional loss of bcl b expression in murine thymocytes leads to t cell deficiency while complete knockout of bcl b was fatal within a few days of birth. recently, specific heterozygous bcl b mutations have been reported in individuals with global development delay. however, only of these cases, both carrying heterozygous missense variants, had low trec values with other cases having frequent infections. little is known regarding the impact of bcl b on human nk and t cell function. methods: we identified a novel heterozygous truncating mutation in bcl b in an infant who was first detected by trec newborn screening. she subsequently developed severe autoimmune hemolytic anemia at the age of months. we used standard immunoblotting and flow cytometry methods to assess protein expression and the impact of this bcl b mutant on t cell and nk cell development and function. results: the patient has a novel single base-pair deletion in the bcl b gene, which is predicted to produce a truncated protein with the loss of of zinc finger domains in bcl b. immunoblotting of t cell blast lysates revealed a reduced bcl b expression in the patient consistent with the heterozygous defect in bcl b but also generated a novel band with a smaller molecular weight that we postulate represents the truncated protein product. while mitogen responses to cona and pha were normal, both cd + and cd + t cell counts were decreased, especially cd + naïve and cd +cd + naïve t cells, suggesting reduced thymic output. the function of th cells was skewed with reduced il- production but increased ifn levels after pma and ionomycin stimulation. moreover, t regulatory cell counts were below normal range. nk cell counts were normal but these were mostly cd bright nk cells. of the few cd dim nk cells that presented, approximately half did not express cd , the fc receptor for adcc. perforin was only present in cd expressing nk cells. as such, anti-cd stimulation understandably led to low but not defective nk cell degranulation. function after stimulation with k cells was normal when controlled for nk cell counts. conclusion: we report a novel bcl b truncating mutation with a leaky scid phenotype that manifested with t-cell lymphopenia and autoimmunity. lowered thymic-derived naïve t and regulatory t cells, skewed th cytokine response, and incomplete nk cell development suggests that bcl b is important for the development and differentiation of multiple lymphocyte lineages. introduction: chronic diarrhea is one of the most common gastrointestinal complaints in patients with common variable immune deficiency (cvid) and can lead to life-threatening complications such as malabsorption and malnutrition. chronic diarrhea in cvid could be caused by infections, an inflammatory bowel disease-like picture, as well as malignancy. giardia lamblia is one of the most common parasites causing diarrhea in cvid (up to %), and can be refractory in these patients, leading to villous atrophy, weight loss, and failure to thrive. case report: a -year-old female with a history of cvid presents with chronic diarrhea and significant weight loss. her cvid was diagnosed by hypogammaglobulinemia (low levels of igg, igm, and iga), inadequate responses to protein and polysaccharide-based vaccines, decreased memory b cells (cd +cd + . %), and recurrent sinopulmonary infections. she was started on immune globulin replacement therapy and had significant improvement in her rate of infections. four years before her presentation to our center, she developed chronic, severe diarrhea. work up revealed giardia lamblia infection on endoscopy and colonoscopy. biopsy showed intraepithelial lymphocytes, villous blunting, and atrophic gastritis with rare plasma cells concerning for non-infectious enteropathy related to her cvid, in addition to the high burden of giardia organisms. she was initially treated with metronidazole for several weeks. however, her diarrhea did not improve, and she developed significant peripheral neuropathy leading to lower extremity weakness and limited mobility. her diarrhea persisted and was associated with approximately a -pound weight loss. repeat endoscopy and colonoscopy two years later showed persistent high burden giardiasis of the small intestine, as well as reactive lymphocytic infiltrates and atrophic gastritis. she was treated with nitazoxanide but continued to have diarrhea, and her stool continued to show trophozoites. given the significant inflammation and the lack of response to multiple antiparasitic agents, she was referred to our center for further evaluation. she was started on oral budesonide ( mg daily) and oral immune globulin ( grams weekly for weeks). with this regimen, she had significant improvement in her diarrhea with a -pound weight gain. repeat colonoscopy showed considerable improvement in inflammation and resolution of her giardia infection, though her stool antigen continues to be positive. conclusions: persistent diarrhea in our patient is most likely due to a combination of cvid enteropathy and giardiasis. a prolonged course of metronidazole and later nitazoxanide did not control her diarrhea and led to significant side effects. switching to an immunomodulatory approach significantly decreased the inflammation in her bowel and may even have helped to reduce the burden of giardia in the gut. targeting both underlying bowel inflammation as well as active infection in cvid patients with chronic diarrhea might be needed to control symptoms. introduction: sphingosine- -phosphate (s p) is a lipid chemoattractant that is critical for lymphocyte egress from lymphoid organs. following a s p concentration gradient maintained by s p lyase ubiquitously expressed in tissues, lymphocytes within lymphoid organs are drawn to efferent lymph and blood unless their s p receptor is internalized or downregulated. owing to diminished degradation of not only s p, but also other sphingoid bases, deleterious mutations in sgpl (encoding s p lyase) perturb sphingolipid catabolism in numerous tissues. correspondingly, human s p lyase deficiency results in multiorgan dysfunction including kidney, skin, endocrine gland, and neurologic impairment alongside expected lymphopenia. although severe t cell lymphopenia (< cells/microliter) rivaling that of severe combined immunodeficiency (scid) can be seen in patients with s p lyase deficiency, no such patients have been identified by newborn screening of t cell receptor excision circle (trec) counts, which are a surrogate measure of effective t cell production. herein, we describe an infant boy with an undetectable trec count at birth who was found to have two novel, biallelic sgpl mutations resulting in s p lyase deficiency. case description: a -day-old boy with a preceding history of fetal hydrops is born at a gestational age of weeks and presents with renal failure, anasarca, and respiratory failure. trec analysis of a dried blood spot obtained at hours of life reveals zero copies/microliter. subsequent peripheral blood studies show profound lymphopenia, with diminished cd + t ( /microliter; cd +, cd +), cd + b ( /microliter), and cd / + natural killer ( /microliter) cell counts. recent thymic emigrants are reduced ( . % of cd + t cells are cd ra+cd +), as is the ratio of naïve-to-memory cd + t cells ( % cd ra+, % cd ro+). expedited whole genome sequencing identifies two novel variants in sgpl a paternally inherited splice site variant (c. + t>c) predicted to impact a canonical splice donor site, and a maternally inherited missense change (c. g>a; p.cys tyr) located in a well-established functional domain of s p. in addition to nephrotic syndrome and lymphopenia, the patient displays evidence of adrenal insufficiency and has increased plasma levels of sphingoid bases and ceramides. before further analyses could be pursued, the infant dies at days of age due to ongoing complications of renal failure and eventual cardiorespiratory failure. summary: we report the first case of s p lyase deficiency identified by newborn trec screening for scid. as sgpl is not included in most commercially-available, scid-tailored gene panels, s p lyase deficiency would be missed by conventional genetic testing. therefore, analysis for variants in sgpl should be considered in neonates with low-to-undetectable trec counts, nephrotic syndrome, and other suggestive sequelae. w a r t s , hypogammaglobulinemia, recurrent infections, and myelokathexis) is a rare autosomal dominant primary immunodeficiency. it is caused by a defect in the gene encoding the chemokine receptor cxcr . this receptor, along with the associated ligand cxcl , regulates leukocyte migration. we present the case of a -year-old female, who presented after she self-identified the signature signs of whim syndrome in herself and multiple family members. objectives: we present the case of a -year-old female who presented with a history of recurrent warts, leukopenia of unknown cause, and recurrent infections as a child. as a child, she experienced multiple ear and sinus infections, along recurrent warts on her upper and lower extremities that have persisted to this day. furthermore, during a routine examination when she was -years-old, she had a complete blood count drawn significant for leukopenia. no further workup was undertaken at that time. when continued leukopenia was noted at the age of , referral to a hematologist and a bone marrow biopsy was completed. bone marrow was significant for myelokathexis with borderline hypercellular marrow for patient age ( % cellularity), and normal cell line quantity. a trial of neupoegen was undertaken, without significant improvement. her family history is significant for father and brother with both leukopenia and recurrent warts. results: genetic analysis showed a heterozygous pathogenic variant in the cxcr gene, c. _ dup (p.ser phe fs* ). recent complete blood count was significant for a total wbc count of . k/ul, with a differential consisting of % neutrophils and % lymphocytes. lymphocyte subsets were significant for quantitatively low cd +, cd + and cd + subsets, with normal numbers of cd + and nk cells. immunoglobulin levels revealed an igg of mg/dl, iga of mg/ dl, and igm of mg/dl; igg anti-diphtheria and tetanus titers were protective, however, none of the s. pneumoniae serotype titers were > . ug/ml. mitogen (pha, cona and pwm) and antigen (candida and tetanus) stimulation of lymphocytes were normal for all stimuli. conclusions: we present the case of a -year-old female with a history of recurrent infections, warts, and myelokathexis. on genetic analysis, she is noted to have a pathogenic mutation of the cxcr gene. the substitution of a phenylalanine for a serine decreases one of the seven serine phosphorylation sites in the carboxy tail of the molecule that occurs upon binding to its ligand, cxcl (sdf ). additionally, the variation generates a premature stop condon terminating the remainder of the carboxy terminal amino acids including ser - , known to have a role in carboxy terminial beta-arrestin binding. failure to generate adequate beta-arrestin binding sites leads to prolonged cxcr cxcl interaction resulting in myelokathexis. background: lacking protective antibodies, patients with primary antibody deficiencies (pad) suffer from frequent respiratory infections leading to chronic pulmonary damage. macrolides prophylaxis has been proven effective to successfully manage chronic lung diseases as cystic fibrosis, bronchiectasis, copd. we conducted a trial to evaluate the efficacy and safety of orally low-dose azithromycin prophylaxis when added to the usual care in pad patients. methods: a -year, phase ii, prospective, multicenter, randomized, double-blind, placebo-controlled trial on pad patients (age - years) with chronic infection-related pulmonary disease. patients received azithromycin mg or placebo once daily three-times a week for months. the primary outcome was the decrease of annual episodes of respiratory exacerbations. secondary endpoints included: time to the first exacerbation, number of hospitalizations, additional doses of antibiotics, health related quality of life measures, and safety. results: forty-four patients received azithromycin and patients received placebo. the mean number of exacerbations was · per patientyear ( %ci · - · ) in the azithromycin arm, and · ( %ci · - · ) in the placebo arm (p= · ). in the azithromycin group the hr for having an acute exacerbation was · ( %ci , - · , p= , ) and the hr for hospitalization was . ( %ci , - · ) (p= · ). the rate of additional antibiotic treatment per patient-year was · ( %ci · - · ) in the intervention and · ( %ci · - · ) in placebo groups (p= · ). improvement in hrqofl was observed in intervention group. azithromycins safety prole was comparable with placebo. conclusion: in pad with respiratory exacerbation, azithromycin prophylaxis led to reduction of exacerbation episodes, of additional courses of antibiotics, and of risk of hospitalization. given the deleterious effects of respiratory diseases adding azithromycin to pad treatment should be considered as a valuable option. background: the autosomal-dominant hyper-ige syndrome (hies), is a primary immunodeficiency caused by mutations in signal transducer and activator of transcription (stat ) that leads to defective th immunity. adverse reactions following -valent pneumococcal polysaccharide vaccine (ppsv ) have been reported in % of stat -hies patients, including severe local reactions that appear to be specific to this vaccine. case report: we present the case of a six-year-old girl, second child of nonconsanguineous healthy parents, that developed an extensive inflammatory skin reaction at the vaccination site following a single dose of ppsv . the vaccine was prescribed due to history of recurrent respiratory tract infections and an incomplete vaccine calendar with no previously administered pneumococcal vaccines. the reaction began after hours with local erythema and edema at vaccination site, expanding in hours to a phlyctenular lesion with no well-defined borders. within the first weeks, it progressively evolved to a deep necrotic lesion that required surgical debridement. the subsequent skin defect required surgical repair with a split-thickness skin graft from her right thigh as the donor site. the complete wound healing process took about months, leaving a large scar ( figure) . the patient had a longstanding history of recurrent infections with multiple hospitalizations including severe neonatal pneumonia that required respiratory support, a colon perforation with secondary peritonitis and septic shock that required a hemicolectomy at months of age, recurrent oral candidiasis, recurrent pneumonias of different lobes, recurrent acute otitis media, a cervical phlegmon, three episodes of dental abscess and multiple kidney abscesses due to gram-negative bacteria treated with intravenous antibiotics and surgical drainage. family history is notable for an older sibling that died due to sudden infant death syndrome. the patients mother has large and wide nose suggestive of stat -hies phenotype, but no history of infections. immunological work up showed mild eosinophilia ( cells/ mm ), elevated ige ( mg/dl), normal igg, iga, igm and lymphocyte subsets (cd , cd , cd , cd , cd ). peripheral th cells were markedly decreased ( . % vs. . % of normal control). specific pneumococcal antibodies evaluated month after psv revealed / serotypes in protective levels. high resolution thorax ct showed multilobar bronchiectasis. echocardiogram and total spine x-rays were normal. stat -hies was suspected with a national institutes of health score of . a novel heterozygous missense variant in stat affecting the src homology (sh ) domain (p.lys glu) was found by next-generation panel sequencing. a variant in the same position (p.lys met) has been previously reported in a hies patient (clinvar). currently, she is on monthly ivig and prophylactic antibiotics (cotrimoxazole, azithromycin and fluconazole). conclusions: the case presented raises awareness on the risk of severe local adverse reactions to ppsv in stat -hies patients. the etiology of such reactions is unclear and warrants further study. the benefits and risks of immunizing stat -hies patients with ppsv should be weighed carefully by medical providers. abstract (max words) introduction: dock deficiency is a rare primary immunodeficiency characterized by susceptibility to viral infections, atopic eczema, defective t-cell activation and th differentiation, impaired eosinophil homeostasis and dysregulation of ige. to date, there are no reported cases from malaysia. objective: we aimed to describe the clinical, immunological profile and mutational analysis of three siblings of consanguineous parents, presented with hyper-ige and lymphopenia between the years and , which were solved by mutational analysis of the second and third siblings. methods: clinical data and investigation results were collated from the medical record. scoring of the symptoms and physical examination findings using nih score was performed. t, b, nk lymphocyte subsets and serum igg, iga, igm, total ige quantification, lymphocyte proliferation test and pneumococcal specific antibody response were performed. mutational analyses were performed in freiburg, germany. result: three siblings presented at different time points over a -year span with raised ige levels, recurrent infections, eczema, hypereosinophilia and bronchiectasis. the nih scores for hyper-ige syndrome (hies) ranged from . we also documented two serious infections in the siblings, which were disseminated cryptococcus neoformans and salmonella sp. immunological results showed t-cell lymphopenia, defective t-cell proliferation, decreased igm, raised ige, hyper-eosinophilia and defective pneumococcal antibody responses present but not in all siblings. we identified a large deletion in dock starting from exon - in of the siblings from mutational analysis performed. we will proceed with next generation sequencing and dock protein assay in malaysia to further characterize the defect. conclusion: our on-going study is the first description of dock in a family from malaysia. the diagnosis of dock should be suspected in cases with raised ige levels, recurrent infections and lymphopenia, despite no warts infection in the history. this study emphasized the importance of international research collaboration and networking in solving complicated cases. the index patient presented at the age of years with increased susceptibility to lower airway and gastrointestinal infections (hospital admissions x/year until puberty). she suffered from mumps and varicella disease despite immunization, as well as from recurrent local, partially destructive hsv infections. she was diagnosed with common variable immunodeficiency (cvid) at age and started on immunoglobulin replacement therapy. following a hypoglycemic seizure at age , the patient was diagnosed with isolated acth insufficiency with secondary adrenal insufficiency requiring hormone substitution. during and following her first pregnancy at age , she suffered from recurrent bronchopneumonias including pneumocystis jirovecii infection, resulting in bronchiectases documented on chest ct at age . currently, chronic lung disease is severely limiting her quality of life (table ) . her daughter was noticed to be hypogammaglobulinemic soon after birth and failed to develop antibody responses to inactivated vaccines. she was started on immunoglobulin replacement therapy. she has not suffered from severe lower airway infections, but developed alopecia totalis at age and nail dystrophy. w h o l e e x o m e s e q u e n c i n g r e v e a l e d a h e t e r o z y g o u s c. _ insacccgag (p.lys profster , nm_ ) mutation in exon of nfkb in both mother and daughter. this monoallelic loss-of function frameshift mutation was not found in gnomad, gvs washington or clinvar databases. as previously published, a monoallelic mutation in this c-terminal domain leads to impaired phosphorylation and subsequent reduced nuclear translocation of the nfkb /p active form. pediatricians and internal specialists need to be aware of the combination of hypogammaglobulinemia, acth deficiency, immune dysregulation and ectodermal dysplasia which is unusual for cvid -possibly indicating nfkb deficiency. this clinical syndrome may overlap with symptoms and signs found in both apeced/ aire (ar) and eda-id/nfkbia (ad) deficiencies. besides ig and hormone replacement therapy, curative treatment with hematopoietic stem cell transplantation is a therapeutic option for patients with nfkb deficiency, although the experience is limited. table introduction: the modes of immunoglobulin (ig) administration for primary immunodeficiency diseases (pidd) differ in pharmacokinetics, infusion parameters, and tolerability. during consecutive clinical studies, a cohort of patients with pidd experienced all modes of administration with the same ig % product in sequence from intravenous (iv) to subcutaneous (sc), then to hyaluronidase-facilitated sc (ighy), providing a unique opportunity to assess each administration modality within the same patient cohort treated and observed at the same sites. here we report the rates of infections stratified by igg trough levels, and the rates of adverse events (aes) with the modes of ig administration (ivig, scig, ighy) within this patient cohort. design and methods: this analysis included patients with pidd aged years who participated in clinical studies: in study (nct ) patients received ivig % every weeks followed by weekly scig %; in study (nct ), patients were treated with ighy every weeks; in study (nct ; extension of study ), patients continued with the same ighy dose. to assess a potential association between the administration route at comparable igg trough levels and the infection rate, igg trough levels were categorized as < mg/ dl, < mg/dl, < mg/dl, < mg/dl, < mg/dl and mg/dl. periods where patients had trough levels within these strata were assessed, and the infection frequency was calculated. the time periods for this analysis were months for ivig and months each for ighy and scig % ( . years) treatments. in order to account for differences in the frequency of administration, rates of systemic and local aes were assessed as aes/patient-year for each mode of therapy. results: for igg trough levels of < mg/dl, the associated annual infection rates were lower or similar for ighy than scig ( the treatment involves the control of infections and immune dysregulation with chemotherapeutic regimens followed by definitive treatment with hematopoietic stem cell transplant (hsct). aim: to describe a female patient with a pathogenic variation in stx with normal cd a expression. results: she was a years old female, the th daughter of nonconsanguineous parents, without relevant personal or family records. she was admitted due to a prolonged febrile syndrome, lymphoproliferation, pancytopenia and hepatitis, with hhv rescued in bone marrow and blood. gancyclovir treatment started with good response. she was admitted one month later with similar clinical symptoms with relapsed hhv infection. furthermore, hemophagocytosis was found in the bone marrow and evaluation of nk cell cytotoxicity demonstrated slightly reduced cytotoxic activity. functional studies for primary fhl were performed: perforin expression and cd a surface expression were normal. she fulfilled criteria of fhl, and treatment with gancyclovir and steroids was administered. despite this treatment, she persisted with activated macrophagic parameters, and started with hlh treatment protocol. she improved the clinical symptoms and laboratory parameters, but persisted with hhv low viremia. three months later, when immunosupression was decreased, she was readmitted with similar clinical manifestations and added neurological symptoms (facial paralysis, abnormal movements and sleep tendency). cerebral spinal fluid was pathological with hhv positive rescue. immunosupresive treatment was adjusted, but hhv copies in blood increased markedly. foscarnet treatment was administered and immunosupression was suspended for days in order to control viral infection. unfortunately the patient died days later. although specific functional tests were normal, sequencing of stx gene by ngs revealed a homozygous variation in c. _ deltgcc, which is a previously reported mutation responsible for fhl. conclusion: despite the fact that cd a was normal, the strong clinical and laboratory results must keep the fhl diagnosis in mind and intensive treatment should be early administered; in order to give the patient the opportunity to achieve the curative treatment. objectives: to report and characterize the clinical course of a patient with apeced and specific antibody deficiency. methods: retrospective chart review was performed. the patient was enrolled in niaid irb-approved protocol -i- . results: the patient is a year-old-girl with apeced caused by homozygous aire c. _ del , who manifested cmc, hypoparathyroidism, adrenal insufficiency, sjogrens-like syndrome, autoimmune hepatitis, intestinal dysfunction and autoimmune pneumonitis. she suffered from recurrent sinusitis and severe pneumonias requiring hospitalization and administration of intravenous antibiotics several times per year. at age , she presented to our institution with fever and cough, a computed tomography (ct) of the chest revealed bilateral pulmonary infiltrates and bronchiectasis. bronchoscopy showed mucopurulent secretions in the bilateral lower lobes with culture of the bronchoalveolar lavage fluid growing streptococcus pneumoniae. further evaluation for an underlying disorder such as primary ciliary dyskinesia and cystic fibrosis including exome sequencing and sweat chloride testing was unrevealing. quantitative immunoglobulins were normal. despite prior vaccination, specific antibody testing showed negative rubeola igg and protective levels (> . mcg/ml) to only of pneumococcal serotypes. lymphocyte enumeration showed normal b cell subsets. as approximately % of apeced patients may experience asplenia, splenic ultrasound was performed confirming the presence of a cm spleen and peripheral blood smear did not reveal howell-jolly bodies. serotyping of the s. pneumoniae isolate confirmed serotype f, which is part of the -valent vaccine. follow up vaccine challenge with the valent pneumococcal polysaccharide vaccine showed an inadequate response. hence, she was started on monthly immunoglobulin replacement and over the following years she has experienced a single methicillin sensitive staphylococcus aureus pneumonia. she has missed very few school days and other parameters including linear growth have improved, she is now along the fifth percentile for height and along the tenth percentile for weight. although she continues to experience intermittent cough she remains active participating in sports without limitation. conclusions: we report the evaluation, treatment and outcome of a patient with apeced complicated by autoimmune pneumonitis and specific antibody deficiency. as infectious susceptibility of apeced classically pertains to the signature infectious disease, cmc, patients with invasive or recurrent infections should be evaluated for underlying immune deficiency. investigation should include assessment for asplenia, quantitative immunoglobulins and specific antibodies with response to antigens. in patients with predominate respiratory symptoms, autoimmune pneumonitis should be evaluated given the near % prevalence of pneumonitis observed in american apeced patients. acknowledgements: supported by dir/niaid/nih introduction: autoinflammatory diseases are genetically heterogeneous disorders of innate immunity characterized by recurrent fever, rash, and/ or serositis, which generally are considered distinct from autoimmune diseases. we report a case of a patient with lupus-like disease and a mutation of nucleotide-binding oligomerization domain-containing protein (nod r w, yao syndrome) suggestive of an overlap between autoinflammatory and autoimmunity processes. case presentation: a -year-old man was evaluated for recurrent pleural effusions, morning stiffness, erythematous rashes, and fever up to °c. history was notable for hashimotos thyroiditis and multiple admissions for presumed pneumonia with recurrent bilateral lung infiltrates and pleural effusions. transbronchial biopsy showed nonspecific pneumonitis and organizing pneumonia. antinuclear and anti-dsdna antibodies were positive. he received prednisone for presumed lupus pneumonitis leading to improvement. prednisone was tapered and hydroxychloroquine was started, but his fevers, pleuritic pain and pleural effusion reoccurred. genetic testing revealed a nod sequent variant (r w) associated with autoinflammatory disease. hydroxychloroquine was stopped and colchicine was added to his regimen, allowing prednisone to be tapered without recurrence of symptoms. further immunological testing revealed increased signaling through the type i interferon receptor (interferon signature). conclusion: although this patient had several clinical (serositis, arthralgia) and immunological (antinuclear and anti-dsdna antibodies, interferon signature) manifestations of lupus, his clinical presentation also was consistent with yao syndrome. in retrospect, he had been having recurrent inflammatory symptoms for many years. recent studies in both mice and humans suggest that inflammasome activation and il- production are involved in the pathogenesis of lupus. this case provides further support for the idea that lupus and hashimotos thyroiditis, prototypical autoimmune diseases, may have overlapping autoinflammatory features. background: the implementation of severe combined immunodeficiency (scid) newborn screening by trec assay has played a pivotal role in identifying these patients early in life. the screen has also led to the identification of infants with other immunologic abnormalities, of which the clinical implications have been unclear and there are limited data on their outcomes. objective: to review immunologic and genetic outcomes of infants referred to an immunology service of a tertiary care center with abnormal newborn scid screens. methods: we retrospectively reviewed charts of infants with positive scid screen from july to november . we excluded patients who had positive screen at < weeks corrected gestational age. we classified outcomes into groups including scid, non-scid t-cell lymphopenia (nscid-tcl) and normal t-cell count. idiopathic t-cell lymphopenia was defined as nscid-tcl (cd + < , cells/mcl) with negative chromosome microarray and negative whole exome sequencing/or genetic panel (either genedx® scid panel or invitae® primary immunodeficiency panel). results: of infants, % were male, % were caucasian, and % were african-american. fifty-four % and % of infants were identified by illinois and missouri screens, respectively. the mean age at initial evaluation was days ( - days). % of infants had a normal tcell count (n= ) or normal repeat newborn screen (n= ), % had nscid-tcl, including mild (cd + , - , cells/mcl, n= ) and moderate (cd + - , cells/mcl, n= ) tcl, and % had scid (n= ), leaky scid (n= ) or complete digeorge (n= ). genetic etiologies of nscid-tcl included q deletion (n= ), trisomy (n= ), and mutations of tbx (n= ), foxn (n= ), and cd e (n= ). three of these infants had novel variants at the time of diagnosis. secondary causes of tcl were identified in infant (thoracic infantile fibrosarcoma). one infant had idiopathic tcl. eighteen infants with nscid-tcl were followed clinically without complete genetic testing performed. for scid, mutations were found in jak (n= ), ada (n= ), il rg (n= ), and rag (n= ). the patient with leaky scid had negative whole exome sequencing. all patients with scid and leaky scid underwent hematopoietic stem cell transplantation at a median age of weeks ( weeks - months), with successful engraftment in all but patient. of idiopathic and nscid-tcl cases followed clinically, had at least one follow-up visit at median age months ( . months . years) and the majority had improved or stable lymphocyte count without serious infections requiring intravenous antibiotics, though had a hospitalization for rsv infection. the mysm patient died after cord blood transplant from unclear etiology. our study had limitations. half of infants with nscid-tcl did not have a complete genetic workup, and only a fifth of patients with nscid-tcl were inpatients, potentially explaining the relatively low number of infants with secondary lymphopenia. conclusions: in our cohort, one-fourth of infants with abnormal scid screen had nscid-tcl. although the majority of nscid-tcl did well, approximately one-third of them had underlying genetic abnormalities associated with their t-cell lymphopenia. ( ) submission id# introduction: accumulation of intracellular adenosine and deoxyadenosine nucleotides (daxp) due to adenosine deaminase deficiency results in profound lymphopenia and severe combined immunodeficiency. left untreated this form of scid is uniformly fatal. while allogeneic hematopoietic cell transplant (hct) and autologous gene corrected stem cell therapy (gt) are potential cures for ada-scid , initiating enzyme replacement therapy (ert) immediately upon diagnosis regardless of definitive treatment is standard of care. hct and gt are not therapeutic options for all ada-scid patients and ert offers immediate therapeutic intervention for these patients leading to partial immune reconstitution, and durable survival in most patients treated. adagen (pegademase), approved by the fda in in the usa, is a pegylated bovine ada (nada) with the enzyme harvested from bovine intestines. this unsustainable production process led to the development of a recombinant enzyme source based on the bovine protein sequence and an improved pegylated linker by using succinimidyl carbamate (revcovitm-(elapegademase-lvlr). methods: a phase ii/iii clinical trial was performed at us sites under institutional irb approval. eligible ada-scid subjects were stable on adagen and without complicating underlying conditions. demographics, medical history, lymphocyte counts, immunoglobulin levels, trough plasma ada activity and rbc daxp measurements were collected. patients were treated with adagen as a single, weekly im dose adjusted to achieve a trough plasma ada activity of > mmol/hr/l and rbc daxp < . mmol/l (protocol target levels). once patients had achieved this level ( - weeks), a seven-day pk on adagen was done and the patients were transitioned to revcovi based on the formula for enzyme equivalent activity of mg revcovi = units adagen. after weeks on revcovi, trough ada and daxp were assessed and a seven-day pharmacokinetic study was conducted at week . patients were assessed periodically for clinical and laboratory values and evaluation of the study endpoints was done at week . subjects subsequently continued on revcovi and were assessed periodically. results: six patients, ages - entered the trial with initial adagen dosing at . - . u/kg/wk (see table ). adagen dosing was adjusted to target endpoints of ada trough activity (> mmol/hr/l) and rbc daxp (< . mml/l). patients transitioned to weekly revcovi using the aforementioned conversion formula at doses of . - . mg/kg/wk. the spectrum of clinical manifestations range from infections to autoimmunity and inflammation among patients with hypomorphic recombination gene and (rag / ) pathogenic variants. auto-antibodies targeting cytokines ifn-alpha, ifn-omega and il- were reported in a large proportion of these patients and their occurrence often coincides with viral infections. we report the time of emergence and relative frequency of anti-cytokine antibodies in children and adults, and their persistence among patients with hypomorphic rag deficiency. antibodies were measured from plasma samples of patients by enzyme linked immunoassay (elisa). our rag cohort includes patients with rag (n= , %) and rag deficiency (n= , %). antibodies targeting ifn-alpha ( %) were most common followed by il- and ifn-omega ( % each). two asymptomatic patients who were detected by newborn screening for scid and received hematopoietic stem cell transplantation had no detectable anti-cytokine antibodies. in the cohort of young children (ages mo- years, n= ), all patients had detectable antibodies to ifn-alpha, prior history of severe viral infection and subsequently developed autoimmune cytopenias. other anti-cytokine antibodies were less common (ifn-omega %, il- %). similarly, children between - yo age (n= ) also had high fraction of anti-ifn-alpha antibodies ( %) with prior history of infections ( %) and continued to have other anticytokine antibodies less commonly (ifn-omega %, il- %). in the adult cohort (n= , ages - years) the frequency of anti-ifnalpha anti-cytokine antibodies were lower ( %,) and il- and ifnomega ( % each) continued to persist. three adult patients had anticytokine (ifn-alpha, ifn-omega and il- ) antibodies tested at multiple timepoints and elevated titers persisted up to years. our data demonstrates that anti-cytokine antibodies, especially those targeting ifn are frequent and emerge early in life in association with viral infections in patients with rag deficiency. a lower fraction of adult patients have detectable anti-cytokine antibodies, and maintain these over several years. anti-ifn-alpha may serve as a useful biomarker for identifying partial rag deficiency among young and adult patients with history of viral infections and autoimmune cytopenias. the role of these antibodies to cytokines is yet to be determined but a specific signature of these antibodies may help to identify an underlying immunodeficiency and initiate early definitive treatment with bone marrow transplantation. anti-cytokine antibodies appear to be a novel tool in evaluation of autoimmune diseases including rag deficiency. introduction: norovirus is one of the most common pathogens causing gastroenteritis in immunocompromised patients, often leading to chronic infection, causing villous atrophy, malabsorption, weight loss, organ failure, need for parenteral nutrition, and death. norovirus treatment in immunocompromised patients is challenging. oral immunoglobulin (poig) has been used to treat norovirus gastroenteritis with variable success. our aim in this study was to determine the outcomes of treating norovirus gastroenteritis in immunocompromised patients methods: electronic medical records were reviewed for patients with norovirus infection confirmed by rt-pcr since january . our initial cohort was focused on patients with primary immunodeficiency (pid), lung, and liver transplant. data on demographics, immunological phenotype, treatment with poig, the number of bowel movements (bm), and virus clearance were collected. descriptive statistical methods were used to describe treatment outcomes. further analysis of patients immunophenotype, immunosuppression medications, and co-morbid illnesses is underway. results: twenty-six immunocompromised patients ( norovirus infection episodes, as one patient had reinfection) were analyzed twelve females, age range months- years. twelve patients had pid diagnosis ( common variable immunodeficiency, severe combined immunodeficiency, x-linked agammaglobulinemia, wiskott-aldrich syndrome, digeorge syndrome, hyper-igm, stat gain-of-function, nemo and lymphopenia in a patient with trisomy ), patients were status-post liver transplant, and two patients were status-post lung transplant. of patients were on ig replacement therapy at the time of the norovirus infection. the average number of bm/day in all patients was . (range - ) . eight patients received poig ( - mg/kg) weekly for a duration from - weeks. three of those received additional nitazoxanide and received ribavirin. / patients in the poig group were receiving total parenteral nutrition (tpn), and / on no treatment group received tpn. the average number of bm/day in poig before treatment was . (range - ), and . (range - ) in those who did not receive any treatment. of ( %) on poig vs. of ( %) in the no treatment group cleared the virus. the average number of weeks to return to baseline bm was . (range - ) in the poig group vs. . (range days- weeks) in the no treatment group. of on poig continued to have chronic diarrhea that is still ongoing. conclusion: despite anecdotal reports suggesting successful use of poig in immunocompromised patients, our data did not show a significant decrease in stool output in patients treated with poig, compared to no treatment. however, poig led to a higher rate of virus clearance. a study with larger sample size might be warranted to identify the patients who benefit from poig in the context of norovirus infection and ensure the appropriate use of ig products, especially given the concerns for the national shortage of ig products. chief medical officer, novimmune sa primary hemophagocytic lymphohistiocytosis (phlh) is a life-threatening, immune regulatory disorder characterized by immune hyperactivation that is driven by high production of interferon (ifn)-. patients with hlh typically develop fever, splenomegaly, cytopenias and coagulopathy. until recently, there have been no fda approved treatments for hlh, and standard dexamethasone/etoposide-based treatment has not evolved significantly in + years. emapalumab-lzsg (ni- ) is a fully human, anti-ifn-monoclonal antibody that neutralizes ifn-and which was recently approved (november ) by the fda for the treatment of adult and pediatric (newborn and older) patients with phlh with refractory, recurrent, or progressive disease or intolerance with conventional hlh therapy. results of the pivotal trial supporting this approval are presented herein. methods: this open-label pivotal study (nct ) includes patients years with a diagnosis of phlh and active disease. data presented were from patients, of whom had failed conventional hlh therapy prior to study entry. the initial emapalumab-lzsg dose was mg/kg given intravenously every - days. subsequent doses could be increased up to mg/kg based on the evolution of response parameters. dexamethasone was administered concomitantly at to mg/m /day and could be tapered during the study. treatment duration was weeks, with possible shortening to a minimum of weeks, or extension up to the time of allogeneic hematopoietic stem cell transplantation (hsct). the primary efficacy endpoint was the overall response rate (orr) at end of treatment, assessed by pre-defined objective parameters, including normalization or at least % improvement from baseline of fever, splenomegaly, cytopenias, hyperferritinemia, fibrinogen, d-dimer, central nervous system (cns) abnormalities, and with no sustained worsening of scd serum levels. the primary analysis used an exact binomial test to evaluate the null hypothesis that orr be % at a one-sided . significance level. patients were eligible to enter an extension phase for follow-up after completing the main study (nct ). the data cut-off applied is july . results: patient characteristics are summarized in table and efficacy is summarized in table . disease at study entry was consistent with the broad spectrum of phlh abnormalities. over % of patients had signs and/or symptoms of cns disease. orr was significantly higher than the pre-specified null hypothesis of %, meeting the primary endpoint. the response rate based on investigators clinical judgement was . %. emapalumab-lzsg infusions were in general well tolerated, with mild to moderate infusion-related reactions reported in % of patients. the observed safety events (pre-hsct conditioning) mostly included hlh manifestations, infections or toxicities due to other administered drugs. infections caused by pathogens potentially favored by ifn-neutralization occurred in patient during emapalumab-lzsg treatment (disseminated histoplasmosis), and resolved with appropriate treatment. no off-target effects were observed. conclusions: treatment with emapalumab-lzsg was able to control hlh activity with a favorable safety and tolerability profile in a very fragile population. the majority of patients proceeded to hsct with favorable outcomes. our results indicate that emapalumab-lzsg should be considered as a new therapeutic option in phlh thanks to its targeted mode of action. results: a total of genes were differentially expressed between t cells of qds patients (n= ) and healthy controls (n= ) (log fold change range (- . , . )).when these genes were tested for pathway enrichment, the top pathways in t lymphocytes based on their p value included communication between innate and adaptive immune cells, cross talk between dendritic cells and natural killer cells, allograft rejection signaling, dendritic cell maturation, and b cell receptor signaling. the top biological processes with differential expression included immune response, inflammatory response, apoptotic process, interferon gamma mediated signaling pathway, nucleosome assembly, defense response to virus, lipopolysaccharide mediated signaling pathway, positive regulation of nf-kappa b import into nucleus, type i interferon signaling pathway, and neutrophil chemotaxis genes. we compared gene expression between qds participants with low t cell counts (n= ) and qds participants with normal t cell counts (n= ) and found genes that were differentially expressed (q< . ) (log fold change range (- . , . ) patient began experiencing recurrent high fevers and developed splenomegaly. elevated transaminases and concern for lymphoproliferative disease prompted a splenectomy and liver biopsy. both the spleen and liver biopsy were positive for ebv but were negative for malignancy. bone marrow biopsy was unrevealing. genetic testing identified a pathogenic variant in xiap/ birc ( c>t), and the patient was treated with high dose oral steroids resulting in an improvement in symptoms. subsequently, therapy with anakinra was started and steroids were tapered. during the steroid taper, he noticed a change in the vision of his left eye consistent with naion, as well as worsening of his colitis. there was loss of the inferior visual field and fundoscopic exam was significant for left optic disc swelling. oct noted superior retinal nerve fiber layer thinning. oral steroids were restarted with improvement in optic disc swelling, but without improvement or change in vision. as of his most recent exam, the patient has persistent bilateral inferior visual field defects with segmental optic nerve atrophy typical of naion. he has continued therapy with anakinra, and subsequently tapered off of prednisone; though he remains on a physiologic dose of hydrocortisone. conclusions: this case demonstrates an unreported ocular manifestation in a patient with xiap deficiency, which clinically appeared sensitive to immunomodulation. our patient is an unusual candidate for naion due to his young age, the average age of onset being the mid to late s, and lack of vascular risk factors. we hypothesize that his hyper-inflammatory condition contributed to irreversible vascular damage in the optic nerve head, resulting in naion. therefore, it may be useful to consider the involvement of systemic inflammatory and immune dysregulatory conditions when treating patients with atypical naion. additionally, naion should be considered in patients with xiap deficiency and sudden unilateral vision loss. the importance of de novo mutations in causing severe sporadic immune disease is well described, yet significance of such a variation in less severe and later onset of immune disease is poorly investigated. whole exome sequencing has been a powerful tool to resolve and explain the genetic basis of novel syndromes in immune related diseases. however, proving causation can be complicated due to low number of the affected individuals. we performed whole exome sequencing in a cohort of patients with noncongenital immune defects, along with detailed cellular biochemical phenotyping. we report and describe a novel non-congenital combined immune deficiency arising from a de novo gain-offunction mutation in ikbkb(c. g>a). this gene encodes ikk , and activates canonical nfkb signalling. cellular and biochemical studies of the proband revealed that ikk v i results in enhanced nf-kb signaling, as well as t and b cell functional defects. ikk v is a highly-conserved residue, and to prove causation, we generated a crispr/cas mouse model that carry the precise orthologous missense mutation. we show that mice and humans carrying this missense mutation exhibits remarkably similar cellular and biochemical phenotypes. dysregulation in patients. total rna isolated from cryopreserved peripheral blood mononuclear cells was reverse transcribed to generate cdna. we selected four known gata transcriptional targets, gata , gata , tal and zfpm (encoding fog ) and used droplet digital pcr to quantify transcript levels normalized to the low-expressing gene tbp . we used samples from individuals with wild-type gata (wt), known gata mutation patients (mut) and two individuals suspected of gata deficiency but without identified mutation or allelic imbalance (unk , unk ). transcript analysis revealed significantly decreased transcript levels of gata , gata and tal in mut pbmcs compared to wt. most wt samples had higher zfpm transcripts than gata mutated patients however it did not reach statistical significance. strikingly, we were able to use this analysis for two individuals suspected of gata deficiency. in the first case (unk ) a yr old female with primary lymphedema, hypogammaglobulinemia, recurrent infections and possible family history of leukemia was referred for gata testing. no mutation was identified however it was noted that she was homozygous across the gene preventing allelic evaluation. the second patient (unk ), a yr old female, had erethemya nodosa on legs, mycobacteria kansasii and cytopenias. in each of the targets analyzed, transcript levels from unk were lower than the wt samples and in a similar range as the gata mutation samples while unk had a profile consistent with the wt samples. we propose the use of gata targets as surrogate markers in cases where a mutation is not identified and allelic expression analysis is uninformative. are often under-reported and under-recognized. we sought to further understand and evaluate the prevalence, type, and association with serum immunoglobulin e (ige) for cvid patients with atopic manifestations. methods: we performed a retrospective analysis of cvid patients with atopic manifestations in the partners healthcare cvid cohort. we evaluated baseline patient characteristics, atopic diagnoses, and serum ige levels. results: in the partners cvid cohort, the average age was years old (± ) and % female. / ( . %) of patients had a diagnosis of asthma, with the majority of these diagnosed by an allergist ( %) or pulmonologist ( %). eczema/atopic dermatitis was diagnosed in / patients ( %), by either an allergist ( %) or a dermatologist ( %). allergic rhinitis was diagnosed in / ( . %) with positive skin prick testing in % of these patients. food allergy was diagnosed in patients ( . %). the median cohort serum ige was . iu/ml. the median serum ige was higher in patients with or more atopic complications compared to those with one or less atopic condition ( vs. iu/ml), which was statistically significant (p= . ). conclusions: we report higher rates of atopy than previously described in other cvid cohorts. consistent with previous reports, we find a low median cohort serum ige level in cvid patients compared to the general population. however, we identify a subset of patients with a predisposition towards atopy and higher ige levels within the broader characterization of cvid, and these patients may have a more specific molecular diagnosis that leads to elevated ige and atopic conditions. whole exome sequencing is underway to further evaluate this hypothesis. whim (warts, hypogammaglobulinemia, infections, and myelokathexis) syndrome is a primary immunodeficiency with autosomal dominant inheritance. in most patients, the genetic cause of the disease is a gain-offunction variant in c-x-c chemokine receptor type (cxcr ) that results in arrest of neutrophil migration from the bone marrow. most patients develop hypogammaglobulinemia and early waning of antibody response with vaccination. however, the exact origin of aberrant humoral immunity in whim syndrome patients is yet to be clarified. here we describe a -year-old iraqi female with a heterozygous cxcr p.ser ter variant, which is presented with haemophilus influenzae meningitis, history of tetralogy of fallot, early onset intermittent neutropenia, lymphopenia, recurrent bacterial and viral infections. immunologic evaluation revealed hypogammaglobulinemia, elevated igm level and a lack of protective vaccine titers after tetanus and prevnar vaccinations. a bone marrow biopsy was consistent with myelokathexis. immune phenotyping, functional studies and apoptosis assays were performed on peripheral blood cells by flow cytometry in our whim patient and controls. although we found that all lymphocyte compartments were reduced, naïve cd t helper cells and switched memory b cells were predominantly affected. spontaneous apoptosis was most pronounced in b rather than t cell compartments in whim patients. in addition, naïve b cells easily activated and died upon activation in vitro. cxcl , a ligand of cxcr , induced elevated t helper cell migration and increased actin polymerization in p.ser ter mutant cells. we conclude that intrinsic b cell abnormalities, such as increased rate of apoptosis and altered activation, might be responsible for defective antibody response in whim patients. although most individuals effectively control herpesvirus infections, some suffer from unusually severe and/or recurrent infections requiring anti-viral prophylaxis. a subset of these patients possesses defects in nk cells, innate lymphocytes which recognize and lyse herpesvirus-infected cells; however, the exact genetic etiologies are rarely diagnosed. plcg encodes a signaling protein in nk cell and b cell receptor-mediated signaling. dominant-negative or gain-of-function mutations in plcg cause cold urticaria, antibody deficiency, or autoinflammation. however, loss-of-function mutations and plcg haploinsufficiency have never been reported in human disease. we examined families with autosomal dominant nk cell immunodeficiency with mass cytometry and whole-exome sequencing to identify the cause of disease. we identified two novel heterozygous loss-of-function mutations inplcg that impaired nk cell function, including calcium flux, granule movement, and target killing. although expression of mutant plcg protein in vitro was normal, phosphorylation of both mutants was diminished. in contrast to plaid and aplaid, b cell function remained intact. plcg +/-mice, as well as targeted crispr knock-in mice, also displayed impaired nk cell function with preserved b cell function, phenocopying human plcg haploinsufficiency. we report the first known cases of plcg haploinsufficiency, a clinically and mechanistically distinct syndrome from previously reported mutations. therefore, these families represent a novel disease, highlighting a role for plcg haploinsufficiency in herpesvirus-susceptible patients and expanding the spectrum of plcg -related disease. we pursued genetic diagnosis, which identified bi-allelic frameshift mutations in the rag gene which had not been previously described: c. delg (p.v sfsx ) and c. _ del insaaaagagtg (p.v kfsx ). taken together, his presentation suggested significant immune dysfunction had evolved since transplant leading to extensive pulmonary nontuberculous mycobacterial infection and possible bronchiolitis obliterans. he therefore will undergo a subsequent unconditioned cd + stem cell boost from his sister, the original donor, once he completes mycobacterium abscessus treatment. this case highlights the potential long-term immune dysfunction which may evolve after unconditioned allogeneic stem cell transplant for scid, in which full engraftment in all myeloid and lymphoid compartments is not expected. it also highlights the importance of guideline-driven follow-up of these patients to monitor for said dysfunction, to prevent serious infection and long-term sequelae. somatic hypermutation (shm) in the b cell receptor (bcr) heavy (igh) and light chain genes promotes affinity maturation and also mutation away from self-reactivity, therefore serves as an important peripheral tolerance checkpoint. as an example, unmutated bcr ighv - genes give rise to antibodies that bind to i/i antigen on red blood cells (rbc) and may elicit cold agglutinin disease (cad), a variant of autoimmune hemolytic anemia (aiha). in case of healthy individuals, frequent shms in the i/i binding site of bcr ighv - genes decrease rbc reactivity and cad. patients with primary immunodeficiencies (pid) paradoxically develop autoimmune diseases, including autoimmune cytopenias, especially aiha. it is unclear if impaired shm of bcr, in particular mutation away from i/i binding, is relevant in the development of rbc reactivity and consequently aiha in a pid background. our studies focus on pid patients with hypomorphic recombination activating gene (rag and ), combined immunodeficiency phenotype and history of autoimmunity, in particular aiha (rag cid/ai). we detected increased frequency of unmutated ighv - bcr in memory b cell repertoires of rag-cid/ai patients as well as elevated titer of unmutated ighv - antibodies in the patients' plasma. lower level of shm likely reflect abnormal germinal center (gc) reaction. as rag and heterotetramer primarily shapes the pre-immune t and b cell repertoire, we studied the interaction of follicular helper t cells (tfh) and naive b cells via in vitro co-culture experiment. interestingly, tfh cells from rag cid/ai patients exhibited highly activated phenotype with increased expression of cd l and il- compared to healthy controls and were able to initiate exaggerated response (class switching and shm) of healthy donor naive b cells. on the contrary, in vitro activated naive b cells from rag cid/ai patients showed impaired proliferation, class switching and decreased level of shm with diminished induction of genes involved t cell co-stimulation (cd , il- r) and shm (aicda, repair enzymes) compared to healthy donor naive b cells indicating intrinsic defect in patient b cells. furthermore, b cells from rag cid/ai patients also showed increased apoptosis and accumulation of gamma-h ax foci at steady state indicating reduced cellular fitness. these findings suggest that the development of aiha is a multifactorial process in partial rag deficiency. our studies highlight that impaired germinal center reaction is an important tolerance checkpoint with the inability of patient's b cells to respond to hyperactive tfh cells and introduce proper level of shm. hence, we propose that b cell fitness is compromised which impairs proper gc interaction, shm, including mutation away from self and sustains rbc reactivity in hypomorphic rag deficiency. introduction/background: the forkhead box n (foxn ) transcription factor is an essential regulator of t cell development, affecting the differentiation and expansion of thymic epithelial cells (tecs). autosomal recessive mutations in foxn cause a t-b+nk+ lymphocyte phenotype due to a thymic aplasia in conjunction with alopecia universalis and nail plate dystrophy resulting from keratinocyte dysregulation. this is a classic nude/scid (omim # ) phenotype. we report on the identification of two independent patients, identified through newborn screening with absent trecs and with a t-nk+b+ scid phenotype who presented with a t cell lymphopenia who had compound heterozygous mutations in foxn . notably, these individuals had normal hair and nail beds. objectives: to determine whether distinct compound heterozygous mutations in foxn cause a novel t-nk+b+ phenotype in the absence of a classic nude presentation. neutralizing autoantibodies (autoabs) against cytokines increase the susceptibility for selected infections (e.g. anti-ifn-autoabs for nontuberculous mycobacteria and non-typhoid salmonella, anti-il- -autoabs for mucocutaneous candidiasis and anti-gm-csf-auotabs for infections by cryptococcus, nocardiae and aspergillus spp). however, the role of anti-il- -autoabs is less clear. il- is a key mediator of the acute-phase response and released early in bacterial infections. patients with impaired signaling or affected production of il- are at increased risk for severe bacterial infections. only three patients with high-titer and neutralizing anti-il- -autoabs who suffered from severe infections caused by s. aureus, s. intermedius and e. coli have been described so far. to investigate the prevalence of anti-il- -autoabs in patients with bacterial infections, we investigated a cohort of patients and identified three further patients, all previously healthy, with neutralizing auotabs against il- who hardly developed an acute-phase response. the first patient suffered from life-threatening pneumonia caused by s. pneumonia, the second patient developed a submandibular abscess and septic arthritis caused by s. pyogenes and the third patient suffered from life-threatening pneumonia caused by s. aureus. we also discovered neutralizing anti-il- -autoabs in two adults among a cohort of patients with autoimmune diseases (n = ), in one adolescent among a cohort of obese individuals (n = ) as well as in three mothers of neonates with impaired il- signaling. so far none of the later individuals developed a severe bacterial infection. this suggests that naturally occurring and neutralizing anti-il- -autoabs are a risk factor for severe bacterial infections yet with incomplete penetrance. ( ) submission id# persistent transaminitis in copa syndrome researcher, immunodeficiencies research unit, national institute of pediatrics, mexico city social service intern, immunodeficiencies research unit, national institute of pediatrics pediatrics resident, pediatrics hospital, st century national medical center, mexican institute of social security researcher, data science department, mexican autonomous institute of technology researcher, department of research methodology, national institute of pediatrics background: inborn errors of immunity constitute a heterogeneous group of over individually rare congenital diseases that involve genes coding for proteins of the immune system, and which result in increased susceptibility to infection, inflammation, autoimmunity, allergy and cancer. the complexity of the diagnostic task, and the intrinsic biases and limitations of the human mind, can be aided by computational tools. among the available machine learning approaches, decision tree algorithms select the best node to split based on entropy and information gain; random forests build hundreds or thousands of decision trees randomly (bootstrapping), to improve accuracy and reduce overfitting. aim: to implement a machine learning-assisted clinical decision support system for the diagnosis of inborn errors of immunity (iei). methods: with a local database of patients with suspected iei, we built a decision tree using c . dtc, and a random forest on python (jupyter notebook, scikit, mathplotlib, pandas, numpy). the database was obtained by conducting an electronic search on medsys of patients with the term immunodeficiency in their electronic medical records, and then hand-picking cases in which an iei had been confirmed or ruled out. it consisted of patients, of which had been diagnosed with iei. we first split the dataset randomly into training ( %) and testing ( %) sets. the decision tree was tasked with classifying correctly pid or not. after running the algorithm in the training set, we evaluated in the testing set. the random forest classified all cases by majority vote into nine groups ( to ), according to the iuis pid group. next, we repeated the process on a larger scale with a dataset of , patients from usidnet. accuracy was assessed by out-of-bag (oob) error estimates. results: accuracy was greater than % for the local dataset (pid/ not, groups), and for the usidnet dataset ( groups). we provide a list of decision nodes and a diagnostic route with those questions that achieved a greater information gain and less entropy. this might help clinicians direct their interrogation and diagnostic approach of suspected iei patients. discussion: we built two classification models. decision trees lend themselves more easily to learning and deriving rules of thumb from their sequences. random forests are more robust and better suited for categoric (as opposed to binary) classification. we next want to develop a chatbot that will ask relevant questions in optimal sequence, and extract undiagnosed patients with suspected iei, based on statistical red flags. researcher, immunodeficiencies research unit, national institute of pediatrics, mexico city dna repair defects are inborn errors of immunity that result in increased apoptosis and oncogenesis. dna ligase -deficient patients suffer from a wide range of clinical manifestations since early in life, including: microcephaly, dysmorphic facial features, growth failure, developmental delay, mental retardation; hip dysplasia, and other skeletal malformations; as well as a severe combined immunodeficiency, radiosensitivity and progressive bone marrow failure; or, they may present later in life with hematological neoplasias that respond catastrophically to chemo-and radiotherapy; or, they could be asymptomatic. we describe the clinical, laboratory and genetic features of five mexican patients with lig deficiency, together with a review of other patients available in pubmed medline. four out of five of our patients are dead from lymphoma or bone marrow failure, with severe infection and massive bleeding; the fifth patient is asymptomatic despite a persistent cd + lymphopenia. most patients reported in the literature are microcephalic females with growth failure, sinopulmonary infections, hypogammaglobulinemia, very low b-cells, and radiosensitivity; while bone marrow failure and malignancy may develop at a later age. dysmorphic facial features, congenital hip dysplasia, chronic liver disease, gradual pancytopenia, lymphoma or leukemia, thrombocytopenia and gastrointestinal bleeding have been reported as well. most mutations are compound heterozygous, and all of them are hypomorphic, with two common truncating mutations accounting for the majority of patients. stem-cell transplantation after reduced intensity conditioning regimes may be curative. department of laboratory medicine, clinical centre immunology, allergy and rheumatology division, department of pediatrics, baylor college of medicine, texas children's hospital, houston,texas, usa laboratory of clinical immunology and microbiology, fungal pathogenesis section, national institute of allergy and infectious diseases, department of intramural research, national institute of allergy and infectious diseases (niaid), national institute of health, bethesda maryland, usa card deficiency is an autosomal recessive primary immunodeficiency known to underlay increased fungal infection susceptibility mostly presenting as invasive cns candida infections (in infancy or adulthood) and dermatophyte infections. more recently, a rare card variant (c. + g>c, leading to exon skipping, card del ) showed a significant protective association towards inflammatory bowel disease (ibd) when present in heterozygosity. at the nih we studied an -year-old male patient (p ) born to a non-consanguineous marriage who presented as an infant with recurrent/severe thrush, candida esophagitis, and an episode of tinea pedis; p also has mild hypogammaglobinemia (igg mg/dl at age y). p s gdna was tested by whole exome sequencing and showed a card c. + g>c mutation in homozygous state. segregation analysis and sanger confirmation determined that both parents and p s elder brother carried the same variant in heterozygosity, while his asymptomatic younger brother (p ) was also homozygous. as previously described, this variant caused card exon deletion as determined in p and p s pbmcs by cdna sequencing and by a lower molecular weight card protein by immunoblot evaluation. p and p s pbmcs, as well as the heterozygous parents cells, showed a defective cytokine generation (tnf-, il- , il- and gm-csf) in response to heat killed candida (hkc), but not to lps. while patients pbmcs failed to induce phospho-erk and phospho-p- upon hkc-stimulation but presented an intact response to pma+ionomycin; the parents cells responded normally to both stimuli. moreover, t-cell activation and proliferation was affected in response to hkc but not to pha in both patients, whereas the parents exhibited normal results under the same conditions. when hek cells were transiently transfected with wt or card del vectors together with a trim plasmid (e -ubiquitin ligase, naturally associated to card ), we confirmed that card del failed to bind trim by immunoprecipitation. furthermore, malt , bcl and trim were only co-precipitated by wt card , but no by card del , strongly suggesting trim is an integral part of the card /bcl /malt -cbm-complex. in summary, herein we demonstrate that the card del allele fails to bind trim , and in turn is unable to conform a complete/functional cbm complex. our data also show that card del acts in a dominant negative fashion in terms of cytokine generation (previously reported), but one wt allele seems sufficient to generate normal levels of hkcinduced p-erk and p-p- , as well as t-cell proliferation. while decreased cytokine generation associated with card del in heterozygosity has been described to be sufficient to protect towards ibd, other defective pathways are affected in homozygosity and likely necessary to confer increased susceptibility to fungal infections. altogether these results suggest that card del acts through a gene dosage mechanism that can dissect pathways that associate ibd protection and fungal infection susceptibility. further work is warranted to explore card del role, if any, in b-cell and t-cell biology. professor, endocrinology, university of michigan medical school background: acquired generalized lipodystrophy (agl) syndromes are a heterogeneous group of diseases characterized by selective dysfunction and loss of adipose tissue after birth. this causes ectopic lipid deposition and deficiency of the adipokine leptin, which promotes metabolic dysfunction through impaired glucose handling resulting in insulin-resistant diabetes mellitus, dyslipidemia and steatohepatitis. while the metabolic effects of altered adipokine secretion are known, the molecular mechanism is less clear. many agl cases are suspected to have an autoimmune etiology. effector and regulatory t cells, dendritic cells and macrophages reside in normal adipose tissue. t cells within adipose tissue highly express pd- and regulatory t cells express ctla , which limits immune activation in the adipose tissue under normal circumstances. thus, inhibition of these immune checkpoints may hypothetically cause immune activation, leading to adipocyte dysfunction and autoimmune destruction. we have encountered two cases that raise clinical concern for this process. patient cases: patient is a -year-old female who presented with failure to thrive at months. she was diagnosed with insulin-resistant type diabetes and hypertriglyceridemia at ages and years with progressive subcutaneous fat loss and low leptin levels culminating in a diagnosis of agl. her childhood clinical course was complicated by hypertrophic cardiomyopathy, hepatomegaly, autoimmune hemolytic anemia with massive splenomegaly and severe chronic diarrhea secondary to autoimmune enteropathy. she presented at years with acute liver failure, thrombotic microangiopathy, nephrotic syndrome and progressive kidney insufficiency. evaluation for her multi-faceted autoimmune presentation identified a familial heterozygous pathogenic variant in the ctla gene (c. _ insgttgg,p.ala glyfster ). despite aggressive immune therapies, including ctla -ig (abatacept), her kidney disease and enteropathy have progressed. patient is a -year-old male diagnosed with localized malignant melanoma of the right neck in july . he underwent excisional biopsy and regional lymph node dissection with negative margins. he relapsed in november and underwent a modified radical neck dissection with lymph node positive for disease and received external beam radiation from january-february . additionally, he was started on anti-pd- therapy with the humanized antibody drug pembrolizumab in april but discontinued the drug in february in the setting of toxicities including hypothyroidism. subsequently, he developed up to . % weight loss with progressive loss of subcutaneous fat first in his face, then generalized to the rest of his body. in the ensuing months, imaging with pet-ct demonstrated loss of subcutaneous fat concurrent with elevations in alt and triglyceride levels plus a low leptin level consistent with agl. conclusion: these cases raise concern that inhibition of the immune checkpoints ctla and pd- may facilitate the development of agl. we hypothesize that these defects significantly increase t cell autoimmune activity in the adipose tissue and/or alter t cell metabolism resulting in agl. disorders of immune dysregulation should be considered in the etiology of agl. similarly, patients with either genetic or pharmacologic inhibition of immune checkpoints should be monitored for the development of agl with careful physical exam and periodic monitoring of glucose and triglyceride levels. background: rosai-dorfman disease (rdd; also known as sinus histiocytosis with massive lymphadenopathy) is a rare non-langerhans cell histiocytosis. it is characterized by proliferation and accumulation of activated histiocytes in affected tissues. classically, rdd presents with bilateral, non-tender, and often markedly enlarged cervical lymphadenopathy. case presentation: a -year-old female presented with a -week history of asymptomatic, persistent and bilaterally enlarged cervical lymph nodes. she was otherwise healthy with no significant past medical history. operative excision biopsy of the largest lymph node confirmed the diagnosis of rdd. three months following diagnosis, routine bloodwork revealed that she had developed lymphopenia (lymphocyte count . x /l). between -year and -and-a-half-years post-diagnosis, the patient was hospitalized and treated with intravenous antibiotics for presumed episodes of osteomyelitis and presumed episodes of lymphadenitis. given the recurrent presumed infections and persistent lymphopenia, the patient was referred to immunology for evaluation. she received a full immunologic work-up. lymphocyte immunophenotyping revealed low cd ( cells/mm ) and low cd ( cells/mm ) counts. the rest of her immunologic work-up was within normal limits. approximately -and-a-half-years post-diagnosis, the decision was made to initiate treatment for rdd. she was started on a -week tapering course of prednisone therapy. within -weeks of starting corticosteroid therapy, the lymphadenopathy had diminished, and by -weeks, the lymphopenia completely resolved. at her most recent clinic visit, she had been free of serious infections for more than -years, and her lymphocyte counts had remained stable and within normal limits for over one year. discussion: in the literature, immune system dysfunction has been reported in rdd, with both auto-antibodies and cellular immunodeficiency implicated. in this patient, the persistent lymphopenia and recurrent episodes of presumed infections appeared consistent with an immunodeficiency. given the known association of rdd with immunologic dysfunction, this was certainly a reasonable assumption; however, when these issues resolved following corticosteroid therapy, we questioned whether her clinical presentation could instead represent a manifestation of her underlying rdd. this case highlights the diagnostic challenge of differentiating between an infection and an rdd exacerbation. the episodes of presumed infections were considered probable but not confirmed with microbiologic or histopathologic specimens. the mechanism underlying lymphopenia in rdd is not clear but may involve decreased production, increased destruction, or sequestration of lymphocytes. to our knowledge, this has not been specifically studied in rdd in the past, however lymphopenia has been linked to lymphocyte maldistribution in other diseases. for example, studies have shown that experimentally altering either the surface of the lymphocyte or the environment through which the lymphocyte travels through can cause sequestration of lymphocytes in various lymphoid organs including lymph nodes. conclusion: we describe the case patient with rdd that developed persistent lymphopenia, and multiple episodes of presumed infections resulting in hospitalization and intravenous antibiotic therapy. her lymphopenia resolved and she had sustained remission of rdd following treatment with corticosteroids. we hypothesize that lymphocyte sequestration in enlarged lymph nodes may have resulted in lymphopenia. this, combined with recurrent rdd exacerbations that clinically resemble infections created a presentation that mimicked an immunodeficiency. background: there is an expanding spectrum of immunodeficiency phenotypes linked to dna repair defects, and some patients may not be diagnosed until adulthood. the most well recognized genetic defect linked to dna repair is in the gene, ataxia telangiectasia mutated (atm), which causes ataxia telangiectasia, characterized by combined immunodeficiency, neurodegeneration, radiation sensitivity, and ocular telangiectasias. however, there are several other dna repair defects associated with immunodeficiency, including some syndromic and severe combined immunodeficiency (scid) disorders. objective: we present the case of an adult patient with prolonged history of recurrent infections, facial abnormalities, and autoimmunity who was found to have radiosensitivity suggestive of a dna repair defect. methods: retrospective chart review, immunodeficiency evaluation, flow-based radiosensitivity assay, gene sequencing. results: a -year-old female was referred to our clinic due to a complex history of recurrent infections and immune dysregulation. the patient had a lifelong history of sinopulmonary infections and panhypogammaglobulinemia with low vaccine responses, leading to a diagnosis of common variable immunodeficiency (cvid), necessitating treatment with immunoglobulin replacement. clinical features were also notable for congenital dysmorphia (strabismus, thin and angular face, high arched palate, nasal septal defect, small mouth, missing dentition, clinodactyly, severe equinovarus, and scoliosis). she was subsequently diagnosed with autoimmune features of vasculitides requiring trial of cyclophosphamide, azathioprine, rituximab and belimumab, which was later discontinued due to neutropenia and worsening sinopulmonary and skin infections despite immunoglobulin replacement. in the course of our evaluation she was revealed to have severe b cell lymphopenia ( %), cd naïve t cell lymphopenia, persistent iga and igm deficiency one-year post rituximab therapy, and elevated alpha fetoprotein (afp). radiosensitivity assay revealed decreased atm phosphorylation and elevated levels of h ax -hours after low-dose ( gy) radiation in her lymphocyte subsets (t, b and nk cells) . due to the evidence of radiosensitivity and elevated afp levels, there was concern for an atm or other genetic defects in a dna repair pathway. therefore, a targeted (primary immunodeficiency genes) panel was pursued for genetic testing ( genes, invitae, san francisco). the evaluation did not identify a variant in the atm gene but rather a variant of uncertain significance was identified in the chd gene, in exon , c. g>a (p.gly arg), which may be mosaic. this variant has not been reported in population databases. chd is typically associated with charge syndrome, and while this patient has some dysmorphic features, she is not typical for charge syndrome. currently, studies on copy number variation (cnv) and deep intronic variants in atm are pending. conclusion: dna repair defects may occur in adult patients with a primary diagnosis of cvid. our patient exhibits some phenotypic features of both a chd variant, and atm leading to possible abnormal dna damage responses (ddr). the exact cause of the immune deficiency in our case remains presently unsolved. this case highlights the relevance of both functional studies and genetic evaluation of complex cases of immune dysregulation, for improving our understanding of the phenotypic variability in these immunological disorders. background: womens health issues in patients with immunodeficiency are largely underrepresented in the literature. there are no studies assessing for fertility issues in patients with antibody deficiencies, and there are few sizable studies examining pregnancy and outcomes on progeny in the same cohort. the two largest studies of pregnancy in antibody deficiency, an idf survey and a study of the czech population, provide conflicting data about the safety of pregnancy for these patients. immunoglobulin replacement has been shown to be safe and beneficial in pregnancy for patients with cvid, however, dosing strategies are unguided. we sought to further understand these and other issues associated with fertility and pregnancy in a large cohort of patients with antibody deficiencies. methods: we performed a retrospective chart review of over patients with icd and/or icd codes of cvid or another antibody deficiency from january to december . inclusion criteria also comprised of having reached at least years of age, the beginning of child bearing years. data collected included disease characteristics, comorbidities, laboratory values, and outcomes. this was followed by a phone survey to elucidate data regarding fertility, pregnancy, delivery complications, and outcomes of children. this study was irb approved. results: the current age of women included ranged from to years of age, currently being in childbearing years to being post-menopausal. forty percent of the women had been pregnant, delivering an average of babies per woman who had been pregnant. fertility issues were not a prominent factor for women who never became pregnant. a majority of women who had babies ( %) did not receive a diagnosis of antibody deficiency until after their child bearing years. recurrent upper respiratory tract infections, bacterial sinusitis, and urinary tract infections during pregnancy were common even in those not yet diagnosed with antibody deficiency. immunoglobulin levels and dosing of intravenous and/or subcutaneous replacement were recorded for a subset of patients with recent pregnancies. the data re-enforced that increases in dosing are needed in the third trimester. cord blood igg levels were also recorded for baby and were the same or higher than the mothers most recent igg prior to delivery. it was rare for children of our patients to be diagnosed with antibody deficiency or a related condition, although cvid, hypogammaglobulinemia, combined immunodeficiency, lymphoma, rheumatoid arthritis, and other diagnoses were found. conclusion: this is the largest report of outcomes before, during, and after pregnancy for patients with antibody deficiencies in the united states. this report highlights the importance of closely monitoring women during pregnancy for recurrent infections regardless of whether a diagnosis of antibody deficiency is present. it also highlights that close monitoring of igg levels during pregnancy is necessary for women with antibody deficiencies. backgrounds: autoinflammatory diseases (aids) are a group of disorders with an inborn error of innate immunity, characterized by recurrent episodes of fever and inflammatory attacks. the spectrum of aids is expanding, but no data on clinical presentation and symptom variability exist for the iranian population for timely precise diagnosis. this study aims at establishing the first autoinflammatory registry of an iranian population focusing on the clinical and laboratory features that may help clinicians toward a better understanding and diagnosis of these disorders. methods: clinical and laboratory characteristics of patients who clinically and or genetically diagnosed with aids collected. we used the updated version of classification criteria from the eurofever registry for the clinical diagnosis. results: in our retrospective study, clinical and laboratory characteristics of the participants collected. mean age of disease onset, disease course manifestation, the mean duration of episodes, atypical symptoms, laboratory and imaging studies as well as complications, and response to treatment also reviewed. data resulted in patients of whom were male. their age ranged from to years. out of were genetically diagnosed. familial mediterranean fever (fmf) was the most common clinically and genetically approved diagnosis. there were also patients suspected of nlrp and nod mutations. age at disease onset differed variably and ranged from the neonatal period to adulthood. fever was present in all the participants and the duration of episodes was - days. the frequency of attacks was between to more than per year. some of the common clinical manifestations were as follows: myalgia or fatigue ( %), arthralgia and arthritis ( %), abdominal pain ( %), aphthous stomatitis ( %), chest pain ( %), chronic gastrointestinal symptoms ( %), skin lesion ranging from urticarial rash and severe nodular acne to pyoderma gangrenosum ( %), exudative and or erythematous pharyngitis ( %), consanguineous parents ( %), symptoms of a type of allergy ( %), lymphadenopathy ( %), splenomegaly ( %), increased acute phase reactant ( %), elevated liver function test ( %) . out of of the individuals reported positive family history and in one of the cases, a patient carrying the homozygous mutation in the mefv gene has shown no clinical manifestation. conclusion: this study highlights the most common manifestations of aids in the population of iranian origin and can be used as evidencebased clinical criteria for their diagnosis. background: the term benign ethnic neutropenia (ben) is used to describe patients of african/arabic descent with absolute neutrophil counts (ancs) less than cells/ul in the absence of other causes. historically, race has been used to support the diagnosis of ben, but self-reported race is notoriously imprecise. the duffy null phenotype (fya -/fyb-) is a known molecular cause of ben and may be a more reliable marker of ben than self-reported race. in addition, although the anc is known to be lower in patients with ben, the lower limit of ancs is poorly described. it is important to differentiate patients with ben from primary immunodeficiency diseases (pidd) and to recognize their expected anc values. methods: eligible subjects included patients less than years seen at the university of michigan between january -july . duffy null (fya -/fyb-) patients were identified using electronic medical record search engine (emerse) software and search terms duffy and fyab. potential subjects were identified; patients met inclusion criteria including duffy null status and the absence of other conditions or medications, potentially impacting ancs. unique healthy anc values were recorded from the duffy null patients. age and sex matched controls were identified using emerse software with search terms tonsillectomy, department of anesthesiology and absolute neutrophil count. subjects with conditions or medications that might impact the anc or of african/arabic descent were excluded from the control group. asian and caucasian patients included as controls were presumed to be duffy null given that < % of these populations are expected to be duffy null. control subjects were identified; met inclusion and exclusion criteria. statistical analysis was performed using two-sided two-sample t-test, anova and onesample t-test. results: the median age of the duffy null cases was . years (iqr: . - . ) with . % (n= ) male and all of african or arabic descent. mean anc for duffy null patients was cells/ul (n= , sd= ) while mean anc for controls was cells/ul (n= ; sd= ) with a mean difference between controls and duffy null cases of cells/ul ( % ci: - ; p= . ). the anc levels between duffy null individuals and controls were evaluated by five age categories (p= . for all age categories). however, there was no difference in anc levels between duffy null cases at different age categories (anova, p= . ). ( . %) duffy null cbcs had anc levels in the nonneutropenic range (> cells/ul), ( . %) cbcs had mild neutropenia ( - cells/ul), ( . %) cbcs had moderate neutropenia ( - cells/ul), and ( . %) cbcs had severe neutropenia (< cells/ul). conclusions: although neutropenia can be associated with pidds and is often a sign of a compromised immune system, duffy null patients have a wide range of values that are often much lower than previously appreciated. the degree of neutropenia related to duffy null phenotype appears to persists throughout childhood and young adulthood. in the context of patients of african/arabic descent presenting with asymptomatic neutropenia, duffy null status should be assessed, and ben should be considered in the differential. complications, hypogammaglobulinemia and a unique characteristic of decreased susceptibility to enveloped viral infections. objective: to investigate the role of impaired host n-linked glycosylation on viral susceptibility to ebola virus. methods: to mimic the condition observed on cdg-iib patients, we tested in vitro three proprietary iminosugars (emergentbiosolutions©), uv b, uv , and uv , which act as competitive inhibitors of -glucosidase i and ii. their ability to inhibit the trimming of n-glycans was compared to known n-glycans modifiers as castanospermine, tunicamycin, as well as the bacterial enzyme peptide-nglycosidase f (pngase-f). ebola virus envelope protein gp was chosen as a prototype glycoprotein, as it is heavily glycosylated with nglycosylation sites. hek t cells were seeded at x ^ cells/well in well plate. after h, cells were transfected with pflag-ebolavirus gp by coupling with effectene®. after h, cells were treated with the inhibitors and harvested h after treatment. trimming of n-glycans was evaluated via molecular weight assessment by western-blot. results: all three inhibitors had comparable effectiveness in inhibiting trimming of nglycans from ebola gp glycoprotein compared to castanospermine. a greater molecular weight shift was seen with tunicamycin and pngase f as expected. conclusions: chemical inhibition of the n-linked glycosylation pathway was successfully achieved using three new mogs inhibitors. this approach merits further investigation on potential applications on antiviral therapies. investigator, laboratory of human genetics of infectious diseases, necker branch, inserm u , necker enfants malades hospital, paris, france head, immunodeficiencies research unit, national institute of pediatrics stat gof mutations are associated with infections, autoimmunity and inflammatory manifestations; the rosacea is one of the manifestations described in this disease, however, the etiology rosacea is not clearly established. the characteristics of rosacea are not described in stat gof in the different clinical series. we describe the different characteristics of rosacea in a family with affected members with stat gof. a family with eight members with stat gof mutation were diagnosed through a first affected member affected with tuberculosis and onychomycosis. seven members more had a clinical history of mycobacterial, viral and fungus infections and autoimmunity disease, in all the seven, was documented the same mutation stat gof. in six of these adults patients, we documented rosacea, it started after adolescence, it was localized in the face and/or eyes, was progressive and not ameliorated with medical treatment and caused nose deformity. rosacea has been described previously as a unique manifestation, and the etiology is not clear, an autoimmune hypothesis has been proposed. the fact that is present in patients with stat gof could suggest that have effectively an autoimmune component. physicians face the patients with rosacea must look for other manifestation presents in stat gof mutations. genetic studies in rosacea patients could evidence an new gene defect. introduction: homozygous mutations causing loss of function of the transcription factor forkhead-box n (foxn ) underlie autosomal recessive severe combined immunodeficiency with congenital alopecia and nail dystrophy (nude scid). affected humans, like the scid mouse, have small or absent thymus, absent or severely diminished t cells, alopecia, and nail dystrophy. infants with nude scid have had neonatal lymphopenia and severe, life-threatening infections. studies of heterozygous carriers of foxn mutations are limited, some having been reported with no phenotype or mild disease manifestations, such as nail dystrophy without lymphopenia or recurrent infections. objective: we describe six infants, including two brothers, with t-cell lymphopenia (tcl) following abnormal california newborn screens (nbs) for scid. each had a single heterozygous variant in foxn . case reports: six infants ( female, male) were referred for evaluation after abnormal california nbs for scid (table ) , with t-cell receptor excision circle (trec) counts from undetectable to (normal > ). all infants were well at the time of initial evaluation. five infants with absolute cd t cell counts > cells/ul and cd t cell counts > cells/ul began evaluation as outpatients on home isolation. patient , with undetectable trecs, cd t cell count , and cd t cell count was urgently admitted for inpatient evaluation and management and immediately started on antimicrobial prophylaxis. patient further evaluation was significant for lymphocyte proliferation to mitogens that was initially normal but waned with time, prompting treatment with a paternal haploidentical hematopoietic cell transplant at months of age. patients and developed neutropenia within weeks of birth treated with granulocyte colony stimulating factor (gcsf). patient remains well on gcsf but has had persistent growth failure under continued evaluation. patients , , and remain stable off antimicrobial prophylaxis, but with persistent moderate tcl. as part of an immune evaluation, patients and - had gene panel testing revealing heterozygous variants in foxn . only the variant of patient (presumed shared by patient , his brother) was predicted to be pathogenic; patient had dystrophic nails and sparse hair most evident after years of age, features shared by his mother and his brother, patient . the other patients lack the clinical features of the previously described phenotype of nude scid. their heterozygous foxn variants are of unknown significance; the functional role of these variants in the patients clinical phenotype is unknown. conclusion: six infants with abnormal nbs for scid had lymphopenia and heterozygous variants in foxn . for these infants, variation exists in level of tcl and presence of hair and nail findings. heterozygous variants of unknown significance in foxn have been uncovered in others, including infants with abnormal nbs for scid, highlighting the need for functional studies to address the possible role of each heterozygous foxn variant in congenital lymphopenia and neutropenia. more work is needed before attributing tcl to a novel foxn variant of unknown significance in the absence of family history, abnormal hair or nails, or functional evidence. remains poorly understood. we characterized the intestinal microbiome and metabolome in patients with cgd to determine if intestinal microbiome and metabolomic signatures could distinguish subpopulations of patients with cgd while using the metabolome to add a functional dimension to observed microbiome signatures. methods: clinical metadata and fecal samples were collected crosssectionally from healthy volunteers (hv; n= ) and patients with cgd (n= ). metabolomic profiling and s rrna (v ) sequencing was performed on fecal samples (total samples: ; reads/sample: , to , ; median: , ) . results: samples from patients with cgd had distinct intestinal microbiome signatures and metabolomic profiles depending on genotype, presence of cgd-ibd and specific interventions (e.g. treatment with an elemental diet). notably, samples from patients with active cgd-ibd (compared to samples from patients without a history of cgd-ibd) had significantly different alpha-and betadiversities, and were enriched for enterococcus spp. signal transducer and activator of transcription gain of function (stat -gof) is a primary immunodeysregulatory disease in which a subset of patients have features of autoimmunity and autoinflammation. enteropathy with growth failure and nutrient wasting is a more common feature of immunodysregulation. ruxolitinib is a janus kinase-stat inhibitor that has been shown effective for the treatment of immunodysregulatory features in stat -gof. our patient is a year old male with stat -gof (c. a>g p.h r) with severe total parenteral dependent enteropathy that led to growth failure (weight . kg). treatment with ruxolitinib led to resolution of diarrhea, return of normal diet, and catch up growth. a dose of . mg twice daily was initially started but was decreased to . mg every morning and mg every evening due to elevated transaminases and thrombocytopenia. over the following year the patient thrived gaining . kg with normal every other day stools. despite weight gain, he remained stable on the same dose of ruxolitinib. as he outgrew his dose, he developed an increased frequency of upper respiratory infections (parainfluenza, coronavirus, rhinovirus). one year after initiation of ruxolitinib, he again developed profuse watery diarrhea that was norovirus positive (weight kg, bsa . ). he was placed on bowel rest and ruxolitinib was dose escalated with a goal of mg/m /day. when he reached mg twice daily, enteropathy completely resolved but liver function tests began to rise. he gained weight and began thriving after weeks of therapy. six months later, enteropathy is controlled, and transaminases have remained elevated (alt iu/l, ast iu/ml) but stable. the appropriate dose and pharmacokinetics for ruxolitinib for the treatment of immunodysregulatory symptoms in pediatric patients has not been thoroughly studied. the dose used was extrapolated from data on the use of ruxolitinib in pediatric myelofibrosis. a dose of mg/m /day appears to provide the most benefit with tolerable adverse effects. this dose should be maintained in order to prevent recurrence of disease related manifestations. abstract clathrin-mediated endocytosis (cme) is the major endocytic pathway by which eukaryotic cells internalize cell-surface cargo proteins and extracellular molecules, thereby allowing for a broad range of biological processes, including cell signaling, nutrient and growth factor uptake, and cell fate and differentiation . the fbar domain only proteins and (fcho /fcho ) are involved in the initiation of clathrin coat pit formation. whether fcho and fcho are functionally redundant or have distinct functions is unclear. we report here the first cases of a severe immunodeficiency due to a genetic defect affecting cme. by using whole exome sequencing and genomic analysis of a targeted pid gene panel, we have identified biallelic loss-of-function fcho mutations in five patients from unrelated families of italian (p ), turkish (p , p , and p ) and algerian (p ) origin with severe t cell lymphopenia manifesting as recurrent and severe infections of bacterial, mycobacterial, viral and fungal origin. p developed ebv-associated diffuse large b cell lymphoma. three patients (p -p ) died in childhood, whereas p and p are alive with full donor chimerism at and . years after allogeneic hematopoietic stem cell transplantation, respectively and have cleared pre-transplant infections. patients p , p , and p carried homozygous frameshift mutations predicted to cause premature termination. western-blotting analysis of ha-or flag-tagged fcho constructs showed expression of truncated products in p and p , whereas no protein was detected in p , presumably due to mrna decay. p and p carried homozygous splice-site mutations at the invariant - and + positions, respectively, leading to skipping of exon in p 's fcho cdna. qpcr analysis demonstrated differential expression of the fcho and fcho genes, with the former being predominantly expressed in lymphoid cells, whereas fcho was more abundantly expressed in fibroblasts and k cells. analysis of t cell activation in p (the only patient for whom pre-transplant pbmc were available) revealed reduced t cell proliferation. while tcr internalization in response to cd cross-linking was normal (consistent with recent evidence that tcr internalization occurs through a clathrin-independent pathway), chase experiments demonstrated that transferrin internalization was abolished in activated t cells from p . we had previously reported that a missense mutation in tfrc, encoding transferrin receptor , impairs transferrin internalization and intracellular iron delivery, causing a combined immunodeficiency with defective t cell proliferation. our data identify the first form of severe immunodeficiency due to defects of clathrin-mediated endocytosis, and provide additional evidence in support of the critical role played by iron cellular metabolism in t cell function and homeostasis. natural history of anti-interferon-gamma autoantibody-associated immunodeficiency syndrome in thailand submission id# centralized sequencing initiative at niaid: year therefore, we set out to investigate the pneumococcal-specific responses of igg, igg , iga and igm to prevnar ® in igg subclass deficient (iggscd) patients in this study. pneumococcal responses were measured using the vacczyme pneumococcal capsular polysaccharide igg, igg , iga and igm elisas (the binding site group, birmingham, uk) in control (n= , median age years, range - ) and iggscd patients (n= , median age years, range - ) recruited from the immunodeficiency unit at the karolinska university hospital iga and igm antibodies in response to pcv vaccination was observed weeks post vaccination in iggscd patients (median, . th and . th percentile these median concentrations were lower than those observed in control patients (median, . th and pcv igg mg/l, - however, percentage changes between pre to post vaccination concentrations of igg, igg and iga in response to pcv in iggscd patients were not significantly different to the control patients u/ml vs . u/ml, respectively) iga u/ml and pcv igm u/ml) responders and non-responders of pcv igg iga and igm in response to pcv in iggscd patients were generally lower compared to the control population. these results support the fact that in addition to igg and igg , measurement of iga and igm could also provide useful information for the clinician gain-of-function ikbkb mutation causes human combined immune deficiency submission id# neutralizing anti-il- -autoantibodies are a risk factor for pyogenic bacterial infections national institutes of health, national institutes of allergy and infectious diseases service of immunology and rheumatology, garrahan national pediatric hospital copa mutations impair er-golgi transport and cause hereditary autoimmune-mediated lung disease and arthritis copa syndrome: a novel autosomal dominant immune dysregulatory disease analysis of pulmonary features and treatment approaches in the copa syndrome expanding the phenotype of copa syndrome: a kindred with typical and atypical features the forest and the trees: machine learning to classify cases of suspected inborn errors of immunity using decision tree and random forest algorithms submission id# card Δ gene dosage: from mono-allelic protection to ibd, to bi-allelic increased fungal infection susceptibility yamanaka d , walkiewicz m , lionakis m and rosenzweig s stim mutation associated with a syndrome of immunodeficiency and autoimmunity a novel hypomorphic mutation in stim results in a late-onset immunodeficiency clinical, histological and genetic characterisation of patients with tubular aggregate myopathy caused by mutations in stim gain-of-function mutation in stim (p.r w) is associated with stormorken syndrome gain-of-function mutations in stim and orai causing tubular aggregate myopathy and stormorken syndrome stormorken syndrome caused by a p.r w stim mutation: the first italian patient and a review of the literature by studying ecs-pre and ecs-post patients we were able to describe the bona-fide effect of gcs on the immune system in general, and t lymphocytes in particular. decreased lymphocyte/thymic output, as well as increased apoptotic tcell death underlies lymphopenia in ecs/chronic gcs-exposed patients. under such conditions, il- was significantly decreased in plasma and our in-vitro studies showed that il- replenishment was able to increase bcl (anti-apoptotic molecule) and bcl expression, and efficiently counteract the apoptotic effects of gcs. recombinant il- has been explored as a co-adjuvant treatment for multiple human cancers and may offer a treatment option for lymphopenia and its genetic counselor, co-director of personalized medicine, division of hematology/oncology/bmt and the institute for genomic medicine, nationwide childrens hospital genetic counselor, division of hematology/oncology/bmt, nationwide children's hospital acknowledgments. genetic sequencing was kindly provided by drs. raif geha and janet chou at the division of immunology, allergy, rheumatology and dermatology, boston children's hospital, harvard medical school. the following grants are acknowledged: . rui . /cippt/ (usm) . bmbf eo (freiburg) the authors would like to thank the director general of health of malaysia for permission to publish this scientific presentation. while severe viral infections may also be an initial presentation of primary immunodeficiency, an immune evaluation is not always obtained in this scenario. patients with xla have an increased susceptibility to severe enterovirus infections, manifesting as chronic meningoencephalitis, which can be fatal. the following case describes a patient with newly diagnosed xla presenting as suspected coxsackievirus and confirmed hhv- meningitis, pseudomonas meningitis and bacteremia. this may be the first reported new diagnosis of xla presenting with both severe bacterial and viral coinfection. case description: a year old, partially vaccinated, hispanic male with a history of febrile seizures presented to the emergency room with fever, oliguria, watery diarrhea, lethargy, meningismus, ecthyma gangrenosum and lower abdominal pain. eight days prior to presentation, he was seen by his pediatrician for facial rash and low grade temperature, and was diagnosed with hand-foot-and mouth disease. he worsened on empiric antibiotics. he had no history of sinopulmonary infections. he did not attend daycare. his vaccines were delayed due to parental choice, and he had not received live vaccines (rotavirus, mmr or vzv). full sepsis evaluation was performed. csf demonstrated pleocytosis, and he was started on empiric antibiotics and transferred to picu. due to worsening abdominal pain, ct of the abdomen was performed, which was consistent with ruptured appendicitis and septic emboli at the lung bases. csf pcr panel was positive for hhv- and he was started on gancyclovir. csf and blood cultures subsequently grew pseudomonas aeruginosa. immune evaluation was performed. serum immunoglobulins were undetectable. in addition to iv antibiotics, he received mg/kg ivig and lymphocyte subsets revealed profound b cell lymphopenia ( . %, cells/ul). btk protein analysis revealed hemizygous btk pathogenic variant confirming the diagnosis of x-linked agammaglobulinemia. the hospital course was further complicated by brain abscesses and pyoventriculitis. he was treated with additional doses of mg/kg ivig and iv antibiotics. repeat mri of the brain nearly weeks after admission demonstrated significant improvement. there was significant clinical recovery. he was discharged home at baseline neurological status. his igg level upon discharge home was mg/dl with the plan to increase dose to mg/kg per month with close monitoring. conclusion: both severe opportunistic bacterial infections and severe viral infections as the initial presentation of xla have been well reported in the literature. this case describes the first reported severe pseudomonas aeruginosa and hhv- co-infection in a newly diagnosed xla patient. this case further highlights the necessity for an increased index of suspicion of primary immunodeficiency in a patient who presents with a severe first infection, despite lack of recurrent infections. we present two patients with dock deficiency due to compound heterozygous variants including a copy number loss at chromosome band p . spanning approximately . mb with partial deletion of the dock gene and a novel c. c>t (p.ser leu) missense variant [chr : (grch ) nm_ ] in dock . functional data is presented to support the pathogenicity of the missense change, along with a review of the literature on dock variants. the proband is a -year-old female with elevated serum ige, severe atopic dermatitis, mild persistent asthma, food allergies, and seasonal allergic rhinitis. she is currently healthy following haploidentical bone marrow transplant in june . she has a -year-old brother with dock deficiency with the same compound heterozygous variants. the brother had later onset of symptoms and a milder presentation of intermittent asthma and seasonal allergic rhinitis. each of the parents is heterozygous for one of the two variants. we evaluated the pathogenicity of the c. c>t missense variant with western blots of dock protein expression, intracellular flow cytometry, and dock stretch assays. flow cytometry showed decreased dock protein expression and stretch assays revealed t cells that were stretched in collagen gels. notably, dock is a large gene containing exons spanning kb and it is relatively common to be a carrier of a rare missense change. in fact, gnomad has approximately individuals with rare (< . frequency) missense alleles in dock . therefore, it is important to demonstrate the potential pathogenicity of any given rare missense change, since few pathogenic missense variants in dock have been reported. of the published dock variants listed in the human gene mutation database (hgmd) only are missense. the majority are gross deletions, of which were reported in hgmd. the remaining reported dock variants include nonsense, splicing, small deletions (all frameshifting), small insertions (all frameshifting), small indels, and gross insertions/duplications. this case demonstrates the relatively infrequent but important contribution of missense changes to pathogenic dock alleles. functional validation of missense alleles is critical in the complex evaluation of dock deficiency. background: hsct is the only known curative option currently for cd l deficiency, an x-linked disorder. in cd l deficiency and other x-linked immune deficiencies, there is an ongoing debate regarding the use of a carrier female sibling or mother as hsct donor. skewed lyonization despite complete donor chimerism has raised concerns for incomplete disease control post-hsct. no data exist regarding the efficacy of related female carrier as hsct donor for cd l deficiency. we herein report outcomes of three patients with cd l deficiency who underwent hsct using a related female carrier donor. method: retrospective review of patients who received hsct from carrier female related donor at three separate institutions. results: three patients with cd l deficiency underwent hsct between - . patient had recurrent episodes of pneumocystis jiroveci pneumonia (pjp) despite being on bactrim and immunoglobulin replacement. patient presented with pjp and severe neutropenia. patient presented with acute respiratory failure from severe respiratory viral infections, cmvand had severe neutropenia requiring g-csf treatment. age at the time of hsct ranged from . - yrs. all three underwent reduced toxicity hsct with busulfan and fludarabine-based preparatory regimens. two of them received matched sibling bone marrow hsct and one received tcr and cd depleted mobilized maternal pbsc haploidentical hsct. donor cd l expression varied from % - % on activated cd cells. immunoglobulin profile and lymphocyte subset were done in two of donors, they were within normal range for age, and none had significant infection history. no history of intermittent neutropenia or oral ulcers noted in donor and the absolute neutrophil count of the donor varied between /l. donor age ranged from . yrs years. cd dose ranged from . x - . x cells/kg and cd dose ranged from x . x cd + cells/kg. gvhd prophylaxis consisted of csa/mmf (n= ) and tcr-a/b depletion and no csa (n= ). neutrophil engraftment ranged from - days and platelet engraftment ranged from days. none of the patients developed acute or chronic gvhd. all three patients maintain full donor myeloid chimerism at the latest testing ( months months); t cell chimerism was % in one and mixed in two patients ( % at nine months, % at months). all three patients had excellent t cell immune reconstitution; two patients came off immunoglobulin replacement - months post hsct, whereas the rd patient is ivig dependent, though iga level was mg/dl at nine months post-transplantation. latest evaluation, months post-hsct, revealed % - % cd l expressing activated cd t cells, which correlated with donor cd l expression and t-cell chimerism. conclusion: our data suggest that hsct utilizing x-linked carrier appears to be safe and results in durable engraftment with excellent humoral and cellular immune reconstitution in patients with cd l deficiency. longer follow-up and data from a larger cohort is needed to make a definitive determination of safety and efficacy of utilizing female carrier as hsct donors in this disease. chief, immunology service, department of laboratory medicine, nih clinical center, bethesda, md, usa background: ikaros belongs to a hematopoietic-specific zinc-finger (zf) family of transcription factors. after dimerizing and dna binding to pericentric-heterochromatin (pc-hc) regions, ikaros is described as a central regulator of lymphocyte differentiation. somatic mutations/ deletions affecting ikaros n-terminal zf have been identified in b-acute lymphoblastic leukemia (all) patients, and germline n-terminal mutations were reported in cvid patients with progressive lack of b cells, hypogammaglobinemia, autoimmune diseases and b-all. methods: we performed targeted sequencing panel for known inborn errors of immunity disease-causing genes in a previously healthy male pediatric patient with burkitt lymphoma, followed by benign lymphoproliferation, thrombocytopenia and neutropenia. b-cells and immunoglobulin levels were normal. ikaros dna-binding, nuclear localization and protein binding were evaluated by emsa, fluorescence microscopy and immunoprecipitation. protein modeling was also performed. results: a novel heterozygous germline mutation in ikaros c-terminal zf dimerization domain (p.r l) was detected in this patient. this mutant showed normal pc-hc localization but dna-binding was markedly reduced in terms of ikaros dimerization and multimerization. moreover, reduced wt-mutant binding was also detected. mutant/wt cotransfection experiments suggest a haploinsufficient defect. geometry based docking of wildtype ikaros predicted that r is within the homodimer interface and may abolish cation-pi interactions and destabilize the ikaros-zf dimerization domain. conclusion: a novel germline ikaros c-terminal mutation affecting homodimerization/multimerization and resulting in reduced dna binding to its dna consensus site was detected in a patient with burkitt lymphoma, benign lymphoproliferation and cytopenias. further studies are warranted to formally establish the casual connection between this genotype and phenotype.( ) submission id# patricia pichilingue-reto, md , prithvi raj, phd , igor dozmorov, phd , quan-zhen li, md, phd , edward wakeland, phd , nancy kelly, md , maria teresa de la morena, md , nicolai s. van oers, phd methods: mice were generated by crispr/cas technology to genocopy the foxn compound heterozygous mutations identified in one of the human patients. thymopoiesis and hair follicle extrusion was analyzed in the various heterozygous and homozygous mutant mice. gene expression analyses of the hypoplastic and normal-sized thymii and the developing skin were performed. in addition, a structure-function analysis was performed with luciferase reporter assays using distinct and previously unreported foxn mutations uncovered in patients who presented with low trecs. results: mice harboring compound heterozygous mutations in foxn that match the human patient phenocopy the t-b+nk+ scid phenotype with normal hair and nails. a functional characterization of the diverse foxn mutations suggests that the severity of the block in thymopoiesis depends on whether the mutations affect the dna binding or transactivation domains of foxn . a -amino acid segment at the end of the dna binding domain appears to be essential for tec development. however, this segment is not required for normal keratinocyte functions in the skin and nail plate. gene expression comparisons are revealing key targets of foxn that suggest a dichotomy in its function in the thymus versus the skin. conclusions: novel compound heterozygous mutations in foxn are causal to a t-nk+b+ phenotype with normal hair shaft extrusion and nail plate extension. this differs from the classic nude/scid (omim # ) reported for individuals with autosomal recessive mutations in foxn . assistant professor of medicine and pediatrics, department of allergy and immunology, uva introduction: copa syndrome is a recently described monogenic immunodysregulatory syndrome. the cop protein, encoded for by the copa gene, is expressed in all cell types and is involved in trafficking from the golgi complex to the endoplasmic reticulum ( ) . the most common clinical features of copa syndrome are interstitial lung disease, pulmonary cysts or follicular bronchiolitis, pulmonary hemorrhage, arthritis, glomerular disease, and autoantibody development ( , ) . atypical features of copa syndrome identified thus far include: extrapulmonary cysts in the liver and kidney, renal and neuroendocrine malignancies, autoimmune neurological disorders such as neuromyelitis optica, and infections, such as meningitis ( ) . clinical case: we present a case of a year-old male with copa syndrome (de novo heterozygous mutation in exon , c. g>c; p.ala pro) manifesting as lymphocytic interstitial pneumonitis, peripheral blood b-cell lymphocytosis, mediastinal lymphadenopathy and persistent transaminitis (alt and ast - u/l, nl ast< u/l, alt < u/l) with normal bilirubin, alkaline phosphatase and pt/inr. the transaminitis was noted prior to diagnosis of copa syndrome, and has persisted despite seven months of therapy with pulse dose steroids, two cycles of rituximab and maintenance therapy with hydroxychloroquine and prednisone. he has had a normal ck and aldolase excluding muscle injury as a source of his transaminitis. a congenital cholestasis panel was normal. markers of autoimmune liver disease including ana, anti-liver kidney microsomal antibody and anti-smooth muscle were negative. serum ceruloplasmin and alpha- -antitrypsin level were normal and celiac serologies, were negative. liver ultrasound was normal. a liver biopsy did not demonstrate inflammatory changes, hepatocyte necrosis, mononuclear cell infiltrates or fibrosis. nonspecific biopsy findings included occasional intraparenchymal neutrophils. it is unclear if these scattered neutrophils and the transaminitis are due to an early as yet unidentified autoimmune process, perhaps in response to hepatocellular stress exacerbated by the copa mutation. discussion: liver involvement has not been reported in copa syndrome. we describe a child with copa syndrome who has had chronic transaminitis with no clear alternative cause. if the phenotypic spectrum of copa syndrome involves the liver, it may limit immunomodulatory options for the treatment of this disease. background: in humans, biallelic stat lost-of-function (lof) mutations lead to a very low or complete absence of the wild-type (wt) protein. whereas, heterozygous mutations can lead to partial loss of function. these patients are susceptible to mycobacteria and herpes virus infections. on other hand, heterozygous gain-of-function (gof) mutations in the stat gene result in a hyperphosphorylated state where patients develop recurrent or persistent chronic mucocutaneous candidiasis (cmc), other cutaneous mycosis, bacterial infections, disseminated dimorphic fungal infections, viral infections and autoimmune disease. methods: in this study, we evaluated novel stat mutations, three gof and one lof. in vitro, pbmcs from these patients were stimulated with ifn-and ifn-for , , and minutes and levels of phospho-stat were measured by flow cytometry. the stat phosphorylation and activity (firefly and renilla luciferase activities) were evaluated in u a-stat deficient cells transfected with a reporter plasmid (for luciferase), wt or mutant-stat plasmids. results: we observed higher levels of stat phosphorylation after two hours of stimulation from three gof mutations compared to wt. however, a lof mutation showed absent stat activation at baseline and in response to ifn-and ifn-. luciferase reporter assay confirmed gain of function and loss of function stat activity observed by flow cytometry. conclusions: using flow cytometry followed by a luciferase assay, we confirmed four novel stat mutations. measuring phosphorylation of stat by flow cytometry is sufficient to determine whether the stat mutation is disease causing. this assay can be translated to a clinically accessible test for stat related disease. background: variants in recombination-activating genes (rag) are common genetic causes of autosomal recessive forms combined immunodeficiencies (cid) ranging from severe combined immunodeficiency (scid), omenn syndrome (os), atypical scid (as) and cid with granulomas and/or autoimmunity (cid-g/ai). the clinical and immunological presentation is broad, ranging from severe infections secondary to near absence of t and b lymphocytes and hypogammaglobulinemia to the occurrence of autoimmunity with late manifestations with partly preserved immune subsets and near normal immunoglobulin levels and broad spectrum of autoantibodies. objective: we aim to estimate the incidence, clinical presentation, genetic variability and treatment outcome with geographic distribution of patients with the rag defects in populations inhabiting south, west and east slavic countries. due to shared ancestry, we also investigated our cohort for founder variants in rag and rag genes. methods: demographic, clinical and laboratory data were collected from rag deficient patients of slavic origin via chart review, retrospectively. results. based on the clinical and immunologic phenotype, our cohort of patients from families represented a wide spectrum of rag deficiencies, including scid (n= ), os (n= ), as (n= ) and cid-g/ai (n= ). sixty-six ( . %) patients carried rag and patients ( . %) carried rag biallelic variants. we estimate that the minimal annual incidence of rag deficiency in slavic countries varies between in , , live birth and it may vary secondary to health care disparities in these regions. in our cohort, % of the patients carried rag p.k vfs* (c. _ delaa), either in homozygous (n= , %) or compound heterozygous (n= , %) form. the majority ( %) of patients with homozygous rag p.k vfs* originated from vistula watershed area in central and eastern poland, and compound heterozygote cases distributed among all slavic countries except bulgaria. clinical and immunological presentation of homozygous rag p.k vfs* cases was highly diverse suggestive of strong influence of other genetic and/or epigenetic factors in shaping the final phenotype. survival of rag deficient patients without hematopoietic stem cell transplant (hsct) (n= , . %) is poor and dramatically improved in the last decade with access to hsct and tailored conditioning regimens. conclusion: we propose that rag p.k vfs* is a founder variant originating from the vistula watershed region in poland, which may explain a high proportion of homozygous cases from central and eastern poland and the presence of the variant in all slavs. our studies in cases with rag founder variants confirm that clinical and immunological phenotype only partially depend on the underlying genetic defect. hsct is becoming available for rag deficient patients in eastern europe with improving outcome. clinical immunologist, centre hospitalier universitaire de montréal (chum) background: acute gvhd following solid organ transplantation is a rare complication. intestinal and liver transplantation have the greatest risk of gvhd among solid organs due to high number of donor lymphocytes in these organs. prevalence of acute gvhd after liver transplantation is estimated to be around , - % and has a poor prognosis ( ) . chronic neurological gvhd is a rare form of gvhd with three subtypes described: cerebral vasculitis, demyelinating disease and immune mediated encephalitis. acute neurological gvhd has no clear definition and is still considered a controversial entity. case presentation: a year-old male underwent cadaveric liver transplantation for alcoholic cirrhosis and hepatocellular carcinoma. the donor was a year-old man who died from anoxic brain injury. the receiver was induced with basiliximab and then put on prednisone, azathioprine and tacrolimus. he was readmitted weeks later for myalgia, headache, fever and neutropenia. clinical state initially improved with empiric antibiotics. he then developed a skin eruption, colitis and dic. the latter was thought to be tacrolimus-induced. he was switched to cyclosporine. skin and rectosigmoid biopsies were compatible with acute gvhd. he received basiliximab and ivig and developed a refractory convulsive state. csf analysis showed elevated proteins and slight pleocytosis. cerebral mri showed non-specific white matter lesions and conventional angiography was normal. chimerism on peripheral blood was % but was % donor on csf. with the presence of chimerism on csf, evidence of cutaneous and digestive gvhd and no infectious cause, neurological gvhd was considered the most likely diagnosis. brain biopsy showed non specific change including neuropil spongiosis, microglial activation and reactive gliosis; but no signs of vasculitis or demyelinating disease. he was treated with atg, highdose systemic corticosteroids, cyclosporine, ivig and intrathecal methotrexate and corticosteroids. csf pleocytosis, proteins and chimerism improved with treatment ( % to % donor). no improvement was noted regarding his neurological state and he developed pancytopenia. he was then transfer to palliative care and died shortly after ( month and a half after liver transplant). discussion: to our knowledge, there is only one prior case published of neurological gvhd following liver transplantation ( ) . both patients were old, had hepatocellular carcinoma and had at least one hla match. age > year, hepatocellular carcinoma and shared hla antigen are known risk factors for gvhd following liver transplantation ( ). our patient had only one hla match with the donor. this case is intriguing as there was a great discrepancy between blood and csf chimerism. acute neurological gvhd following transplantation is a real complication. it must be taken into consideration in patients with neurological involvement after transplant, even solid organ transplantations. introduction: hyper-igm syndrome are rare. although no data are available on the frequency of activation-induced cytidine deaminase (aid) deficiency, this disorder is estimated to affect less than : , , individuals. by the year , cases worldwide ( ) with such mutation have been described. we describe a patient with hyper igm by mutation in the aicda gene. case report: mvv, -year-old boy, born to consanguineous parents, was referred with recurrent pneumonia, which started shortly after discontinuation of breastfeeding at months old. repetitive otitis evolved with bilateral tympanic and partial hearing loss. he was submitted to adenoidectomy without improvement. immunological evaluation showed normal numbers of b and t cells with cd + ( /mm , %), cd + ( /mm , %), and cd + ( /mm , %). immunoglobulin concentrations were: igg = mg/dl (p ). treatment with intravenous immunoglobulin and prophylactic antibiotic was initiated and he had no infections during the follow up except for one episode of sinusitis. at years of age, molecular evaluation was performed and a mutation in homozygosity in the aicda gene (omim * ) at position chr : . . was found, confirming the clinical suspicion. conclusion: the role of aid in the immunoglobulin class-switch recombination (csr) and somatic hypermutation (shm) have not been fully elucidated. summarizing within the shm and csr processes, aicda mutation can induce dna lesions in directed sequences in the s and v regions required for dna cleavage. recurrent infections and consanguinity raised the suspicion of inborn errors of immunity in this patient. the literature described late diagnosis as in the second or even the third decade of life. it was suggested that high levels of igm antibodies may provide effective defense, at least, against some infectious agents. it is important to emphasize that the impossibility to obtain genetic diagnosis did not prevent to introduce therapy. * aicda: activation induced cytidine deaminase gene patients with chronic granulomatous disease (cgd) are at risk for recurring infections and non-infectious inflammation, reduced quality of life and life expectancy. conventional treatment with life-long anti-bacterial and antifungal prophylaxis prolongs lifespan but does not eliminate the lifelong risk of infection and inflammation. allogenic stem cell transplantation is currently the only curative option for this disease. although sct with reduced intensity conditioning has improved treatment-related mortality and efficacy, it remains a matter of debate whether all patients with cgd benefit from sct, whether pre-existing infections and non-infectious inflammation are risk factors and at what age sct should be performed. we compared patients with cgd on conventional treatment with those after stem cell transplantation for their prognosis and evaluated potential risk factors for stem cell transplantation outcome followed up in six european centers. frequency of infections, inflammatory complications, hospitalizations, operations and immunomodulative/immunosuppressive therapy, height and weight were compared in patients on conventional treatment /before stem cell transplantation versus patients after sct. correlation between transplantation outcome and patient characteristics or medical history was tested. patients were recruited, on ct, after stem cell transplantation. before/without transplantation % of patients suffered from at least one infection, , % from inflammatory complications. patients on conventional treatment developed infection/inflammation/ hospitalization/surgery at a median of , (range [ , - , ] , iqr , ) per year, versus (range , iqr , ) in the first year after stem cell transplantation but (range [ - ], iqr , ) after the first year post stem cell transplantation. there was a significant decrease of all complications after stem cell transplantation (p < . ). growth improved significantly after stem cell transplantation (z-score weight - , versus - , (p. ), z-score height - , versus - , (p. )). nevertheless, complications post stem cell transplantation are frequent: % of patients had at least one infection, % had severe acute gvhd, % chronic gvhd, % had graft rejection, % died. preexisting active mold infection increased the risk for complications after stem cell transplantation. in summary infections and non-infectious inflammation are common in patients with cgd on conventional treatment, their growth is significantly impaired. stem cell transplantation, if successful, significantly reduces the risk for infections and non-infectious inflammation. however, treatment related mortality of stem cell transplantation in patients with cgd remains considerable. introduction: development of a diverse t cell repertoire is essential for full immune recovery following definitive treatment for severe combined immunodeficiency (scid), whether by allogeneic hematopoietic cell transplantation (hct); autologous gene therapy (gt); or, in the case of adenosine deaminase deficiency, enzyme replacement therapy (ert). however, the time course and depth of diversity of t cell receptor rearrangements have been difficult to measure directly, necessitating estimates from total and naïve t cell counts and from spectratyping, in which t cell receptor (tcr) beta chain diversity is estimated by the length distributions of cdna amplicons between a series of tcr beta chain variable (v-beta) segments that have productively recombined with the tcr beta-chain constant region. analysis of the actual sequences of rearranged tcrs could indicate more precisely the status of the t cell compartment of these patients, and might reveal oligoclonal expansion of dysregulated t cells, t cell insufficiency, or t cell exhaustion. objectives: we wished to ascertain whether deep sequencing of individual tcr v-beta rearrangements in peripheral blood could be performed sequentially following diagnosis and treatment of scid to differentiate satisfactory immune reconstitution from incomplete or skewed repertoire development that might require further cellular therapies. methods: equal amounts of total rna were obtained from peripheral blood of controls and scid patients pre-hct and at d, and mo, and yearly post-treatment(s). cdna was used as template to semi-quantitatively amplify rearrangements at the tcr-beta locus (trb). raw sequences were filtered to remove pcr errors, and resulting fastq files were converted into fasta format (seqtk software, github, inc), filtered for productive rearrangement, and analyzed for v, d, and j gene composition and length (imgt highv-quest software). the vdj statistics file (past program) was used to calculate a shannon entropy (h) index to measure repertoire diversity, taking into account both abundance and richness of the overall repertoire; and a gini-simpson index of unevenness, measuring inequality in the relative representation of species in a given sample. graphical representations of repertoire diversity were generated by hierarchical tree maps of the trb repertoires (irepertoire software): each dot represents a unique sequence and the dot size corresponds to frequency of that sequence in the total sample. results: tcr v-beta sequence analysis of scid patients (image) showed (top) baseline poor diversity due to pre-treatment ada deficiency followed by improvement to normal complexity (shannon h > . ) after receiving peg-ada and autologous lentivirus gene therapy at age m; (middle) increasing diversity in xscid after maternal t-depleted unconditioned hct, although b cells did not recover; and (bottom) failure of initial unconditioned maternal t-depleted hct in another xscid patient at m, followed by autologous lentivirus gene therapy with subsequent improvement (shannon h increasing from . to ) months later. conclusions: tcr v-beta diversity sequence analysis provided a detailed assessment of repertoire diversity in response to cellular therapies for scid. this method could become a useful predictive tool to measure successful t cell immune reconstitution, both as early as d and in the years following treatment. background: the stim (stromal interaction molecule ) protein, encoded by the stim gene, is involved in calcium regulation in the endoplasmic and sarcoplasmic reticulum. pathogenic variants in this gene are associated with three different disorders. homozygous loss-of-function (lof) pathogenic variants in stim have been reported to cause autoimmune cytopenias, lymphoproliferation, enamel defects, anhydrosis, and iris hypoplasia. the first described cases had frequent mortality in early childhood due to recurrent life-threatening infections and development of kaposi sarcoma ( ), while recently discovered cases have had more prolonged survival, though still with recurrent serious infections ( ) . heterozygous gain-of-function (gof) pathogenic variants in stim have been associated with both tubular aggregate myopathy (tam) and stormorken syndrome. tam is a clinically heterogeneous progressive muscle disorder with a variable age of onset. muscle biopsy characteristically demonstrates tubular aggregates, with type ii muscle fiber atrophy ( ) . stormorken syndrome has a phenotype that includes miosis, thrombocytopenia, intellectual disability, mild hypocalcemia, muscle fatigue, asplenia, and ichthyosis ( ) . the thrombocytopenia has not been reported to be immune-mediated; rather it is due to abnormal platelet calcium regulation ( ). we report a patient with stim pathogenic variant presenting with tam and immune-mediated thrombocytopenia, along with lymphoproliferative features, arthritis, and a mild immune deficiency. case: the patient is a -year-old with a history of congenital thrombocytopenia (platelets ranging , - , ) who presented with acute arthritis of bilateral hand joints after exposure to cold temperatures, which resolved with naproxen. he had back pain without muscle weakness, and preceding sore throat and general fatigue. labs were significant for leukocytosis and elevations in his inflammatory markers and creatine kinase. mri of his lower extremities was negative for inflammatory myositis, but did demonstrate bilateral hip and knee effusions, and significant inguinal lymphadenopathy and hyperintense linear signal changes in the mid-and distal femurs with patchy red marrow signal. abdominal ultrasound could not identify a definite spleen. bone marrow biopsy was negative for malignancy but significant for toxic granulation of neutrophils, evident of inflammation. alpha-beta double negative t cells were not elevated. interferon-gamma was mildly elevated. flow cytometry demonstrated normal t, b, and nk cell absolute counts. circulating antibodies against platelets (both igg and iga) were detected. on lymphocyte antigen and mitogen proliferation testing, he did not exhibit any proliferation when stimulated with tetanus toxoid even though he had been fully vaccinated against tetanus. muscle biopsy demonstrated large vacuoles consistent with tam on both light and electron microscopies. invitaes primary immunodeficiency panel identified a pathogenic variant in stim (c. c>t; p.arg trp), consistent with a diagnosis of autosomal dominant stim -related conditions, including stormorken syndrome ( ) . conclusion: this patient expands the phenotypic spectrum of stim related disease. based on previous evidence, gof pathogenic variants in stim are associated with tam and stormorken syndrome, while lof pathogenic variants in stim are associated with immune deficiency. however, our patient with a stim gof pathogenic variant has features of lymphoproliferation and immune dysregulation in addition to tam. stim gof pathogenic variants should be considered in the differential of patients with immune thrombocytopenia and lymphoproliferation. references: introduction / background: card is critical for protein binding upstream of nf-kb (nuclear factor kappa b) and mtorc (mammalian target of rapamycin complex ) the signaling pathway involved in t-cell activation and inflammatory response. prior testing of card mutations demonstrated variable t-cell dysfunction. in vitro studies have demonstrated reduced interferon gamma cytokine production, interference of t-cell receptor (tcr) signaling, and th phenotype skew in t-cells with card defects. while homozygous mutation causes severe combined immunodeficiency deficiency, heterozygous card defect is associated with atopy by way of inappropriate th skewing. heterozygote atopy is characterized by eosinophilia, elevated ige, and severe dermatitis. despite multiple studies demonstrating in vivo consequences of card on t-cell function, little is known of the clinical significance. moreover, few studies have demonstrated the impact of card mutations on b-cell maturation and development, despite the recognized tcr and interleukin signaling deficits. objectives: this case demonstrates a card defect that evolved from atopy to combined immunodeficiency requiring intravenous immunoglobulin therapy. it highlights the poorly understood effect of card mutation on t-cell function, and the downstream impact on b-cell quality. methods: -year-old male, with past medical history of t-cell lymphoma and no evidence of disease status post autologous stem cell transplant, was found to have card e d missense mutation by genetic testing. consistent with previous literature regarding heterozygous card defects, the patient suffered from frequent asthma exacerbations, aeroallergen sensitivity, and eczema. lab work was consistently positive for elevated ige and eosinophilia. family history was positive for a son born with congenital molluscum, and multiple other children with recurrent infections. one child was also identified with card mutation. the patient had flow cytometry demonstrating % of circulating cells with atypical immunophenotyped cd + t-cells, and positive gene rearrangement studies. his qualitative immunoglobulin levels were significant for consistently low igm, but normal quantity igg. in the patients adulthood, he had recurrent bronchitis and pneumonia requiring hospitalization and intravenous antibiotics. given his recurrent infections, the patient underwent immunodeficiency evaluation. despite previous infection with herpes zoster, the patient did not have protective titers. additionally, the patient had received the pneumococcal conjugate vaccine once, and the pneumococcal polysaccharide vaccine four times. the most recent vaccination was one year prior to evaluation. despite repeated vaccinations, titers were unprotective. consequently, the patient was diagnosed with combined immunodeficiency, and initiated on intravenous immunoglobulin therapy. results: in summary, card defect is a cause of atopy, observed to become less severe with age. studies of card heterozygote mutations have demonstrated in vitro deficiencies in t-cell activation, likely secondary to skewed or decreased inflammatory cytokine production and tcr activation. our patient demonstrates that the variable t-cell dysfunction seen in vitro can have significant clinical implications evidenced by his inadequate vaccine response, and recurrent infections. his combined immunodeficiency poses a connection between card defects and, not only t-cell, but also b-cell function. conclusions: further studies are needed to determine deficits in t-cell and b-cell function in the setting of card defect, as this case suggests the clinical implications span further than atopy. genetic variants in the scaffold gene card cause disorders of the immune system. the clinical course and treatment depends on whether the card variant causes gain-or loss-of-function. however, lymphocyte immunophenotyping and proliferation assays in cells expressing card variants don't easily distinguish between gain-and loss-of-function. to address this challenge in variant interpretation, we used multiplexed genome editing in a lymphoma b cell line (tmd ) to generate cell populations expressing all possible singlenucleotide variants in the n-terminal amino acids of card . to assess function in each variant, we tracked its relative abundance over multiple conditions using dna sequencing. since card is required for survival of tmd lymphoma b-cells, cells expressing clinically identified gain-of-function variants grew faster relative to cells expressing other variants, even in the presence of upstream pathway inhibitors. upon evaluation of the relative abundance of each variant in genomic dna and mrna, we found that clinically identified loss-of-function variants were depleted in mrna, which could be attributed to alterations in splicing or to nonsensemediated decay. to address the impact of splicing, we modeled a newly-identified splice donor mutation (c. + g>a) found in two patients from one family diagnosed with combined immune deficiency, autoimmunity and atopy that was also observed in our screen. we show that the variant causes deletion of exon four and that card missing exon four exerts a dominant-negative effect leading to decreased nf-kb signaling and cell growth. these experiments demonstrate the utility of multiplexed functional assays for determining variant effect in clinically-relevant genes, which will improve diagnosis and treatment in patients. mutations in the rag and rag genes in humans cause a wide spectrum of phenotypes, ranging from severe combined immunodeficiency (scid) with lack of t and b cells to omenn syndrome (os), atypical scid (as) and combined immunodeficiency with granulomas and/or autoimmunity (cid-g/ai). here, we sought to investigate the molecular basis for phenotypic diversity presented in patients with various rag mutations. methods: we have recently described a novel flow-cytometrybased assay in which mouse rag -/-pro-b cells containing an inverted gfp cassette flanked by recombination signal sequences (rss) are transduced with a retroviral vector expressing either wild-type or mutant human rag (hrag ). the green fluorescent protein expression directly relates to the activity of rag proteins, representing a quick and powerful tool to correlate between defective activity of hrag mutant and severity of the clinical phenotype. the genetic variants of hrag analyzed in this study were affecting the various domains of the protein: ring, zinc finger ring type domain (amino acids - ); nbr (amino acids - ); hbr (amino acids - ) and the core domain (amino acids - ). using this sensitive assay, we tested the recombination activity of human rag variants that have been reported in patients. results: we have demonstrated correlation between the recombination activity of the mutants and the in vivo clinical phenotype of patients. in particular, similarly low levels of recombination activity were observed in patients with scid and os, whereas patients with as and especially those with cid-g/ai carried mutations that retained significant residual levels of activity. conclusions: these data provide a framework to better understand the phenotypic heterogeneity of rag deficiency. here we report a case of a child with b. cepacia lymphadenitis, ultimately diagnosed with takayasu arteritis. takayasu arteritis is a large vessel vasculitis which may have a nonspecific clinical presentation in childhood possibly leading to difficulty in diagnosis. case: a -month-old female presented with two weeks of fever, respiratory distress, and lymphadenopathy, and was treated with ivig for presumed atypical kawasaki disease. imaging studies performed due to worsening respiratory distress revealed retropharyngeal abscess with bilateral cervical lymphadenopathy, culture-positive for prevotella oralis and melaninogenica, with improvement following incision and drainage and antibiotic therapy. recurrence of fever and respiratory distress prompted ct imaging of her neck significant for worsening lymphadenopathy. cultures from lymph node biopsy grew b. cepacia. following treatment, she was readmitted with respiratory distress requiring chronic steroid treatment and found to have candida albicans on bronchoalveloar lavage and necrotizing granulomatous inflammation on lung biopsy. an immunologic evaluation was notable for two normal dhr assays. cgd genetic panel was negative for pathogenic variants in cybb (p ), ncf (p ), cyba (p ), ncf (p ). testing was also notably negative for hiv pcr, bartonella pcr, cryptococcal antigen, histoplasma antigen, bal afb stain and mycobacterial cultures, cmv pcr, ebv pcr, anca, serial blood cultures, and sweat test. lymphocyte subsets were normal for age. mitogen stimulation test, myeloperoxidase antibody igg, serine protease igg, c level, lad panel, and cytokine panel were normal. autoimmune lymphoproliferative disorders (alps) panel was negative. whole exome sequencing demonstrated heterozygous mutations in cfi and jak , not considered to be clinically relevant given the patients clinical picture and laboratory evaluation. the patient was then lost to follow-up for over a year. at the age of years, the patient presented with fever and back pain. imaging revealed severe large vessel vasculitis involving the aorta and subclavian, vertebral, mesenteric, and renal arteries. she also had evidence of cardio-embolic strokes on brain mri. she had had no significant interval infections, and her immunologic evaluation remained unrevealing. in the context of her new vasculitis, evaluation for deficiency of ada (dada ) was negative. she was ultimately diagnosed with takayasu arteritis and has begun therapy with systemic corticosteroids, aspirin, and etanercept. conclusions: we describe a case of b. cepacia infection in a child without identified immunodeficiency, ultimately diagnosed with a large vessel vasculitis. the presence of b. cepacia infection warrants a thorough investigation. burkholderia has been previously associated with giant cell arteritis, another type of large vessel vasculitis, though causation has not been established. to our knowledge b. cepacia infection has not been associated with takayasu arteritis. christopher santaralas, valentine jadoul, jacqueline squire, john cannon, jessica trotter, susan aja, neil goldenberg, david graham, jennifer leiding background: chronic granulomatous disease (cgd) is a primary phagocytic immunodeficiency secondary to mutations in any of the components of nadph oxidase. in addition to infection susceptibility, patients with cgd can develop auto-inflammatory disease that is difficult to manage. metabolomics is the systematic study of small molecule biomarkers of the clinical phenotype of disease. we sought to investigate plasma metabolic profiles in cgd as we hypothesized that unique signatures may differentiate patients with cgd. methods: plasma collected from subjects with cgd ( x-linked, p phox-deficient, p phox-deficient) and x-linked cgd carriers was analyzed using a targeted multiplex assay by liquid chromatography mass spectrometry (lc-ms) and simultaneously a profiling assay by lcms. sufficient signal was present for metabolites. x-linked cgd and p phox-deficient groups were sufficiently sized for multivariate and univariate analyses in metaboanalyst. twelve patients had a single time point of plasma metabolomics analysis and three had multiple time points, including one in whom both pre-and post-hematopoetic cell transplantation time points were assessed. post-hoc comparisons were also performed for those with, versus without, clinical comorbidities of autoinflammation. results: plasma from patients with x-linked and p phox deficient cgd had a differential metabolomic signature at baseline. many metabolites as measured by ion intensity were present at high levels, particularly homocysteine, kyneurine, tryptophan, citric acid, carnitine, methionine, and adenosine. increased values of metabolites reduced to that of normal (compared to post hct). homocysteine levels were elevated among patients with (mean . x ), versus without (mean . x ), clinical comorbidities of auto-inflammation (i.e., colitis, lupus). baseline samples showed elevated kynurenine among all cgd patients, relative to historical normal controls (unmatched, separate analysis). patients with colitis had elevated citric acid levels that were higher among patients with (mean . x ), versus without (mean . x ), colitis irrespective of genotype. conclusions: preliminary data with a small patient subset suggest that patients with cgd have metabolomic signature distinguishable by phenotype. citric acid cycle metabolites are elevated in crohns disease and ulcerative colitis. based on our data, citric acid may too act as a biomarker for inflammatory bowel disease in cgd. analyzing a larger number of samples, across time points, will likely describe a metabolomics profile for cgd and identify biomarkers for auto-inflammation in cgd. no significant medical history in mother; paternal history is unknown and unavailable.no significant medical history in mother or father. rationale: ataxia telangiectasia is a disorder with variable phenotypes characterized by cerebellar degeneration, immunodeficiency, chromosomal instability, radiosensitivity, and cancer predisposition which may correspond to the degree of atm protein expression and/or radiosensitivity. we used in vitro cytometric assessment of atm, smc and h ax phosphorylation to assess dna damage in response to radiation and found that two siblings with the same copy number gain in atm have variable clinical neurologic and immunologic phenotypes. methods: chart review and radiosensitivity assays using cytometric assessment of patm, psmc , and h ax expression after irradiation with gy. results: patient a is a month old male identified after having low trecs on newborn screening, then found to have lymphopenia and elevated igm. he has diffuse café au lait macules and no neurologic symptoms. his year old sister, patient b, was being followed by neurology for several years for ataxia. she has selective iga deficiency, normal lymphocyte counts, lymphocyte proliferative responses, gammaglobulins, and vaccine specific antibodies. both patients have a copy number gains in atm (exons - ). mother and father both have copy number gains in atm and are healthy without neurologic symptoms or recurrent infections. both patient a and b have normal atm protein expression. phosphorylated atm, smc , and h ax was assessed in lymphocyte subsets (t, b, and nk cells) after low-dose irradiation to induce dna double-stranded breaks (dsbs). these parameters were assessed at hour post-irradiation when they are expected to be maximal and at hour post-irradiation, when under conditions of normal and effective dna repair, the phosphorylation state returns to baseline. patient a had abnormal patm and psmc but normal h ax expression hour and hours after irradiation of t, b, and nk cells. patient b had normal patm, psmc , and h ax expression in t cells but abnormal patm and psmc expression in b and nk cells hour after irradiation. patient b, however, had abnormal atm phosphorylation at hours after irradiation of t, b, and nk cells.conclusions: our results indicate that a unique copy number gain in atm within a family can correspond to different clinical and immunologic phenotypes as well as variable degree of radiosensitivity. the persistence of h ax at hours post-irradiation and impaired phosphorylation of atm and smc at hour post-irradiation demonstrates defects in dna dsb repair, and this is variably altered in different lymphocyte subsets. correlation between atm phosphorylation in lymphocytes with outcomes may be an area for future studies and particularly important in counseling patients regarding outcomes. antibodies have been implicated in both protection and pathology of dengue virus infections. however, much of this data is gathered from serum/plasma responses that is a cumulative of historical and ongoing infection. to precisely understand the role of antibodies with respect to the ongoing dengue virus infection, we employed the cutting edge approach of generating of human monoclonal antibodies from individual plasmablasts from peripheral blood of dengue patients that allows us to probe for answers at a single cell level. this method involves ex vivo single cell sorting of plasmablasts from peripheral blood of well-characterized dengue infected patient followed by single cell molecular cloning of immunoglobulin heavy-and light-variable regions into expression vectors containing the defined constant region followed by transient cotransfection of hek a cells with the heavy and light chain expression vectors made from genes arising from the same cell. thus far, using this powerful technology, for the first time in india, we have made number of human monoclonals, of which are specific to dengue and neutralize dengue virus at various concentrations. all the neutralizing antibodies are dengue-envelope specific and bind the highly conserved fusion loop of the dengue virus envelope. together, with the ongoing comprehensive analysis of the b cell repertoire and somatic hypermutations, these studies provide a detailed understanding of the dengue-specific plasmablast cell response at a single cell level and create a platform for testing these antibodies for basic research, diagnostic, prophylatic and as well as therapeutic applications. surviving. six of the ( . %) surviving patients remain dependent on ig replacement despite robust donor chimerism of - % and no active gvhd. all but two received rituximab pre-hsct. of the patients who are independent of ig replacement, only one ( . %) received rituximab post-hsct, whereas / of the ig dependent patients received rituximab post-hsct. t cell immune profiling revealed that the absolute numbers of lymphocyte subsets, cd + naïve t cells, and cd + recent thymic emigrants were not statistically different between ig independent and dependent patients ( figure ). however, there was a marked decrease in the number of total b cells, the percentage of memory b cells (cd + b cells), and classswitched memory b cells (cd + igd-igm-cells) in ig dependent patients ( figure ). t follicular helper (tfh) cell populations (cd +cd ra-cxcr +pd +) were evaluated in four patients and the frequency was similar to healthy controls ( . +/- . vs. . +/- . %). the ability of the patients naïve b cells to class-switch was assessed following exposure to il- , anti-cd antibody, and anti-human igm, and revealed normal b cell class-switching and differentiation to plasmablasts ( figure ) . additionally, t cell ability to provide b cell help was assessed by coincubating naïve b cells with activated cd + t cells. this revealed comparable b cell class switching to that of healthy controls. conclusion: the high incidence of poor long-term functional b cell reconstitution following allogeneic hsct for xlp- could be related to the use of rituximab in the post-hsct setting rather than pre-hsct. normal tfh numbers and function, and ability of b-cells to class-switch in-vitro suggest that persistent hypogammaglobulinemia is these patients is unlikely from a b or t-cell intrinsic defect. the possibility of rituximab induced acquired lymph nodal stromal defect in these patients is being explored. further studies are needed to understand the biology of persistent hypogammaglobulinemia in xlp- . additionally, due to the high incidence of persistent hypogammaglobulinemia, exposure of rituximab should be limited post-hsct. background: tandem mass spectrometry (ms/ms) has emerged as a primary platform for many clinical and newborn screening laboratories. the application of ms/ms mainly focuses on the quantification of accumulated small metabolites in plasma resulting from various metabolic defects. however, many disorders do not yield such metabolic markers and would benefit from the direct quantification of intracellular target proteins. unfortunately, the extremely low (e.g., pmol/l range) protein concentrations in blood cells limit their detection via ms/ms. in recent years, peptide immunoaffinity enrichment coupled to selected reaction monitoring (immuno-srm) has emerged as a promising technique for the quantification of low abundance proteins in complex matrices, including dried blood spots (dbs). our lab has demonstrated that immuno-srm methods are able to reliably distinguish affected patients from the normal controls for wilson disease (wd), wiskott-aldrich syndrome (was), severe combined immunodeficiency (scid), and x-linked agammaglobulinemia (xla) (j. proteome res., and front. immunol., in press). these results demonstrate the utilization of immuno-srm as a sensitive platform for multiplexed quantification of signature peptides in the low pmol/l range. methods: several candidate peptides for each protein were selected based on uniqueness using in silico blast tools and lc-ms/ms response. monoclonal antibodies (mabs) were then generated for peptide enrichment from dbs. blood from normal controls, wd, xla, scid, and was patients was spotted onto filter paper, dried, and stored at - °c until use. proteins were extracted from dbs, digested with trypsin, and enriched using mabs bound to magnetic beads. the enriched peptides were then eluted and analyzed using srm mode with a waters xevo tq-xs. results/conclusions: to date, immuno-srm methods have been generated for wd, was, scid, xla, and cystinosis. preliminary data shows immuno-srm methods are able to reliably quantify target proteins using signature peptides and accurately distinguish affected patients from normal controls. analysis of signature peptides found statistically significant reduction or absence of peptide levels in affected patients compared to control groups in each case (was and btk: p = . , scid: p = . ). intra and inter-assay precision ranged from - % and - %, respectively, and the multiplexed assay showed a broad linear range ( . fmol peptide) . in a blinded sample set of pidd patients and normal controls, immuno-srm-predicted diagnoses showed excellent agreement with clinical or genetic diagnoses. every molecularly-confirmed case of was and btk was also diagnosed by immuno-srm analysis. in addition, randomly selected samples provided by the nbs laboratory of washington state were tested and peptide concentrations were found to be within normal ranges. efforts are underway to validate and incorporate peptide biomarkers for adenosine deaminase deficiency, dock deficiency, and ataxia telangiectasia, as well as general markers for nk cells and platelets into a single multiplexed assay. in addition, scid, was and xla samples continue to be run while we focus on reducing assay costs, time, and necessary sample input. our data herein provides proof of concept for the immuno-srm workflow to be extended to various other genetic diseases as potential multiplexed newborn screening methods.( ) submission id# the background: the long-term effects of glucocorticoids (gcs) on the immune system have been extensively studied in patients with different underlying conditions (e.g, malignancies or autoimmune conditions), as well as in healthy volunteers receiving short-term courses of these drugs. although these approaches provided highly relevant data, neither of them answered the unbiased/bona-fide effect of long-term gcs use on the immune system. endogenous cushing syndrome (ecs) may be caused by pituitary or ectopic acth-producing adenomas, or by tumors or hyperplasia of the adrenal cortex. patients with ecs present with different gcsdependent manifestations, including those affecting the immune system as neutrophilia and lymphopenia. when tumors are removed, most of the effects of gcs tend to progressively regress. methods: paired samples from patients with ecs due to acth-producing adenomas (age range - y, females) were studied before (ecs-pre) and - months after tumor removal (ecs-post). extended lymphocyte phenotypes and apoptosis in different cell subsets were evaluated by flow cytometry. cytokine production (elisa) and responses, as well as their effects on cell proliferation and viability, were evaluated using cell trace violet and annexin-v staining. results: among multiple immunophenotypic changes, ecs-pre patients showed significantly reduced naïve t cells and recent thymic emigrants (rte) as well as increased apoptosis in t cells when compared to themselves (ecs-post) or age matched healthy controls. moreover, significantly increased exhausted cd t cells were observed in ecs-pre patients. interestingly, ecs-post patients showed full cellularity recovery of t cells and rte with increased proliferation and reduced apoptosis, in addition to correction of most of the other changes evidenced. significantly lower il- plasma levels were also detected in ecs-pre when compared to ecspost patients. to determine the role of il- in an ecs-resembling condition, healthy control pbmcs were treated with gcs in-vitro and the effect of il- and other cytokines was tested. a significant reduction in apoptosis was observed in the il- -treated cells that almost completely countered the pro-apoptotic effects of gcs; il- was also significantly more efficient than il- , il- , ifn-alpha and ifn-gamma in rescuing cells from apoptosis. il- -specific upregulation of bcl and bcl expression was evidenced in these cells.publisher's note springer nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations. key: cord- -t tysvr authors: cho, sung-yeon; lee, hyeon-jeong; lee, dong-gun title: infectious complications after hematopoietic stem cell transplantation: current status and future perspectives in korea date: - - journal: korean j intern med doi: . /kjim. . sha: doc_id: cord_uid: t tysvr hematopoietic stem cell transplantation (hsct) is a treatment for hematologic malignancies, immune deficiencies, or genetic diseases, ect. recently, the number of hscts performed in korea has increased and the outcomes have improved. however, infectious complications account for most of the morbidity and mortality after hsct. post-hsct infectious complications are usually classified according to the time after hsct: pre-engraftment, immediate post-engraftment, and late post-engraftment period. in addition, the types and risk factors of infectious complications differ according to the stem cell source, donor type, conditioning intensity, region, prophylaxis strategy, and comorbidities, such as graft-versushost disease and invasive fungal infection. in this review, we summarize infectious complications after hsct, focusing on the korean perspectives. hematopoietic stem cell transplantation (hsct), known previously as bone marrow transplantation, is performed to treat two broad categories of diseases. the first category consists of functional failure of bone marrow or marrow-derived cells, including aplastic anemia, myelodysplastic syndrome, immunodeficiency syndromes (severe combined immune deficiency or chronic granulomatous disease), genetic diseases (mucopolysaccharidosis, glycogen storage diseases, etc.), or hemoglobinopathies (thalassemia, sickle cell anemia, etc.). in such cases, hsct is applied to replace the de-fective, non-functional bone marrow tissues. diseases in the second category-which includes hematologic malignancies such as acute or chronic leukemia, multiple myeloma, lymphomas, and myeloproliferative neoplasms-are more common indications for hsct, the performance of which aims ( ) to restore the myelosuppressive or myeloablative effect of cytotoxic treatment (such as intensive chemotherapy and/or total body irradiation [tbi]) to eliminate malignant cells, and ( ) to induce a graft-versus-leukemic effect by providing anti-neoplastic immune cells expressing tumor-specific or -associated antigens [ , ] . until the s, hscts were simply classified as au- tologous or allogeneic according to their donor status, and survival rates were very low. since the introduction of the human leukocyte antigen (hla) concept, the incidence of graft failure and/or graft-versus-host disease (gvhd) has decreased. in recent years, allogeneic hsct has involved various sources of hematopoietic stem cells (bone marrow, peripheral stem cell, cord blood, and mesenchymal cells), donors (sibling, unrelated, haploidentical), and conditioning regimens (standard [myeloablative], reduced intensity). supportive therapies such as transfusion, colony stimulating factors, and antimicrobial agents have also been developed. although various early diagnosis and therapeutic techniques have been developed to improve transplant performance, infectious diseases still affect the prognosis of hsct recipients [ ] [ ] [ ] . in korea, hsct was first performed in [ ] , and the number of transplants has rapidly increased over the past years. in addition to being susceptible to infectious diseases due to neutropenia immediately after hsct, delayed immune recovery is evident over a long period of time after engraftment, depending on the type of hsct, immunosuppressants, and acute or chronic gvhd. because the prevalent infectious diseases vary geographically and over time, state-of-the-art knowledge is needed [ ] [ ] [ ] . in this manuscript, infectious complications after hsct, particularly those common in korea, are reviewed, as well as the epidemiology, diagnosis, preventive or therapeutic strategies, and novel drugs for improving outcomes of patients. the post-hsct period is usually divided into the ( ) pre-engraftment period (day to days , ( ) immediate post-engraftment period (engraftment to day ), and ( ) late post-engraftment period (days to ). in general, the neutrophil count recovers to weeks after hsct. however, the functional recovery of various cell types is also important. immunological recovery takes to months for nk cells, to months for b-cells and cd t-cells, and to years for cd t-cells [ ] . the use of t-cell-depleting agents (anti-thymocyte globulin [atg] , alemtuzumab, etc.) to reduce the risk of gvhd and graft failure could improve the outcomes of transplantation but may delay immune recovery [ ] . recipients of t-cell-depleted transplantation may be vulnerable to viral infections-such as cytomegalovirus (cmv), epstein-barr virus (ebv), or adenovirus-and adoptive immunotherapy may be applied [ ] . during the pre-engraftment period, the risk of opportunistic infection varies depending on the type of anticancer drugs used, conditioning intensity (myeloablative or reduced intensity), and presence of acute gvhd. autologous hsct leads to more rapid recovery of immune function than allogeneic hsct. in the post-engraftment period, the immune system is reconstituted and recovered in autologous hsct recipients. however, allogeneic hsct recipients undergoing long-term immunosuppressive therapy for chronic gvhd remain at risk of infection. table shows the risk factors for, and infectious diseases commonly encountered after, hsct. as shown in table , the infectious diseases that occur before the engraftment are similar to those that develop during the neutropenic phase after chemotherapy [ , ] . neutrophils are important in innate immunity against microorganisms. a decrease in the number of neutrophils increases the susceptibility to infection. in addition, patients with neutropenia have a reduced number of leukocytes. therefore, inflammatory findings, which are common in patients with normal leukocyte counts, are not often seen, with the exception of fever, which is difficult to diagnose, so the appropriate time to start treatment may be missed [ ] . the main sources of bacterial infections in the pre-engraftment phase are the normal gastrointestinal flora and indwelling vascular catheters. gram-negative bacilli (gnb) are common pathogens in the former, while gram-positive cocci (gpc) are more common in the latter [ ] . in general, microbiologically defined infections (mdis) account for only % to % of cases of neutropenic fever [ ] . the distribution of causative organisms in febrile neutropenic patients varies geographically. in the united states and europe, gpc accounted for % to % of mdi until the early s, while the proportion of the korean journal of internal medicine vol. , no. , march gnb increased after the mid- s. in a multicenter epidemiological survey, the proportions of gram-positive and -negative cases were % (range, % to %) and % (range, % to %), respectively [ ] [ ] [ ] . enterobacteriaceae was the most common causative organism, accounting for % (range, % to %). the second most common organism was coagulase-negative staphylococci ( % [range, % to %]). in korean studies, gnb were also more common until the mid- s, which is consistent with other reports from the asia-pacific region [ ] . intestinal bacteria such as escherichia coli, klebsiella pneumoniae, and pseudomonas aeruginosa are frequently reported gnb, and enterococcus, streptococcus, and staphylococcus species are common among gpc [ ] [ ] [ ] [ ] [ ] [ ] . because the epidemiology of bacterial pathogens and the resistance profiles varies, early empirical antimicrobial agents must be selected according to the distributions and susceptibilities of the frequently detected bacterial taxa in that center [ , , ] . in the late post-engraftment phase after autolo-gous hsct, the immune system is reconstituted and restored, which is consistent with the time at which chronic gvhd typically develops after allogeneic hsct. therefore, autologous transplant recipients rarely experience opportunistic infections at this time, but allogeneic hsct recipients (who take chronic immunosuppressive drugs for several months) are at risk of infection. in this period, infectious diseases may continue to develop. in addition, the immune deficiency resulting from chronic gvhd is associated with infections by encapsulated bacteria such as streptococcus pneumoniae, haemophilus influenzae, and neisseria meningitidis [ , ] . deficiencies in immunoglobulin g (igg ) and igg have been reported in patients with chronic and severe gvhd, and are associated with severe pneumonia, meningitis, and sepsis due to s. pneumoniae [ ] . early diagnosis and treatment are essential, and vaccination should be emphasized in this high-risk group of patients. immunization after hsct will be described in a separate section of this review. over the past decade, the frequency of detection of resistant bacteria has increased worldwide, including in patients with neutropenic fever and hematologic malignancies [ ] . among extended-spectrum β-lactamase (esbl)-producing enterobacteriaceae, k. pneumoniae comprises around two-thirds, and e. coli about one-thirds (range, % to %), depending on the region [ , [ ] [ ] [ ] . in the late s, % of e. coli and k. pneumoniae bacteremia isolates from neutropenic patients were es-bl-producing strains in a single-center study [ ] . the incidence of infections caused by carbapenem-resistant enterobacteriaceae (cre), along with esbl-producing enterobacteriaceae, is increasing in hematologic malignancy patients [ , [ ] [ ] [ ] [ ] . k. pneumoniae, e. coli, p. aeruginosa, and acinetobacter baumannii are clinically important strains associated with the acquisition of carbapenem resistance. a study of prior colonization by cre as a risk factor for cre bloodstream infections found that % of neutropenic patients with carbapenem-resistant k. pneumoniae rectal colonization experienced bloodstream infection by an identical strain [ ] . there is no multicenter study of cre in hsct recipients. however, a single-center, retrospective cohort study performed in korea reported that the incidence of carbapenem-resistant a. baumannii bacteremia was . cases per , patient-days; post-engraftment infection by this organism can be fatal [ ] . the rate of methicillin-resistant coagulase-negative staphylococci is reportedly higher than % in most centers, while the incidence rate of staphylococcus aureus in hsct patients is low with a high methicillin-resistance rate of median % (range, % to %) [ ] . in the late s, coagulase-negative staphylococci were reported to have a methicillin resistance rate of more than % and s. aureus a rate of more than % [ ] . the incidence of enterococcus bloodstream infections was . cases per , patient-days, with vancomycin-resistant enterococcus (vre) accounting for . % [ , ] . most vre bloodstream infections are caused by e. faecium and are associated with long-term hospitalization and an underlying medical condition, but the resistance itself has not been associated with mortality [ ] . the epidemiology of drug-resistant pathogens is important for establishing strategies for prophylaxis and initial empirical antimicrobial therapy. because domes-tic data are limited to single-institution and retrospective studies, multicenter nationwide studies of the epidemiology of bacterial infections in korea are required. prophylaxis plays an important role during the pre-engraftment phase ( table ). oral fluoroquinolone prophylaxis can reduce febrile neutropenic episodes and related mortality in allogeneic hsct recipients [ , ] . according to the guidelines from the infectious diseases society of america and infectious diseases working party of the german society of hematology and oncology, ciprofloxacin and levofloxacin are equally effective, but levofloxacin may be more effective in terms of broader coverage of viridans streptococci when oral mucositis is present [ , ] . although fluoroquinolone prophylaxis has led to an increase in the frequency of drug-resistant (including quinolones) pathogens, the benefit reportedly outweighs the risk [ ] . fluoroquinolone prophylaxis decreased the rate of identification of p. aeruginosa [ ] . however, fluoroquinolone prophylaxis is reportedly correlated with resistance development and clostridium difficile-associated diarrhea [ ] [ ] [ ] [ ] [ ] [ ] . moreover, fluoroquinolone prophylaxis may reduce the rate of bloodstream infections, but not the overall mortality rate [ ] . therefore, the benefit of routine fluoroquinolone prophylaxis should be weighed against its toxicity and effect on the local epidemiology. in neutropenic patients during the pre-engraftment phase, infections can progress rapidly. in addition, as it is difficult to distinguish between bacterial infection and non-infectious fever in the early stages, empirical antimicrobial therapy is recommended immediately in all patients with febrile neutropenic episodes. except for the presence of definite infection foci, the broad-spectrum β-lactam antibiotics ceftazidime, cefepime, piperacillin/tazobactam, and carbapenem are recommended as they have activity against gram-positive and -negative bacteria, including p. aeruginosa [ , , , ] . step-down therapy can be considered after an initial regimen comprising two or more antimicrobial agents, depending on the local epidemiology and resistance profiles. the de-escalation approach is generally recommended when high incidence of resistant pathogens in neu- the korean journal of internal medicine vol. , no. , march tropenic fever, or high colonization rates of multiple drug resistance bacteria, or if the patient has a history of infection with drug-resistant pathogens [ , , ] . however, the data on cut-offs for resistance are insufficient to formulate such a strategy. in addition, no randomized controlled study has compared the therapeutic efficacy of the escalation and de-escalation approaches in hsct recipients. selection of the appropriate strategy should take into account the patient's condition and the prevalence of resistant organisms in the center. aspergillus species are the most common causes of invasive fungal infections (ifis) in patients with hematologic diseases, followed by candida spp. and rare fungi [ ] . the causative fungi are not different after hsct [ , ] . the characteristics of, and risk factors for, fungal infections differ according to the phase after transplantation. during the pre-engraftment period, neutropenia and mucosal damage are risk factors for invasive candidiasis [ ] . antifungal prophylaxis regimens used during and/or after hsct are listed in table [ , ] . recent advances in the use of prophylactic antifungal agents during hsct have led to changes in the epidemiology of candidiasis [ ] . the korean epidemiologic data show that the incidence of candida albicans infection is markedly reduced by antifungal prophylaxis [ ] . nonetheless, the incidence of c. albicans has been decreased by the increasing prophylactic use of fluconazole or echinocandins, and attention should be paid to the selection of antifungal agents for empirical or targeted therapy for invasive candidiasis [ , ] . post-engraftment, the risk of candida infection decreases with the recovery of neutropenia and mucosal skin loss, but the risk of aspergillosis remains [ ] . invasive aspergillosis (ia) that develops during the neutropenic period typically involves angioinvasion due to host immunosuppression. in contrast, ia that occurs in non-neutropenic patients, usually due to long-term use of steroids or immunosuppressive agents for gvhd, is more likely to be caused by local inflammation (i.e., airway invasiveness) than by vascular invasion [ ] . in the seifem b- study, the most frequent etiological agents of ifi were aspergillus ( . %) and candida ( . %) species. the ifi-attributable mortality rate was . % ( . % for allogeneic hsct recipients and . % for autologous hsct recipients) [ ] . in a prospective observational study performed in the asia-pacific region, table examples of therapy antimicrobial prophylaxis intermediate autologus hsct anticipated neutropenia less than - days bacteria: consider fluoroquinolone prophylaxis during neutropenia a fungus: consider prophylaxis during neutropenia and for anticipated mucositis, consider pcp prophylaxis virus: during neutropenia or longer depending on risks high allogenenic hsct including unrelated or family mismatched donor anticipated neutropenia more than days prolonged neutropenia secondary neutropenia after engraftment status of malignancy not in remission ghvd with significant steroids treatment (> mg/day of prednisolone equivalents) use of secondary immunosuppressive agents due to refractory gvhd (e.g., tnf-α inhibitor) bacteria: consider fluoroquinolone a fungus: consider prophylaxis during neutropenia, consider pcp prophylaxis virus: during neutropenia or longer depending on risks hsct, hematopoietic stem cell transplantation; pcp, pneumocystis jirovecii pneumonia; gvhd, graft-versus-host disease; tnf-α, tumor necrosis factor-α. a recent data concern the correlation with fluoroquinolone prophylaxis and development of resistance or clostridium difficile associated diarrhea. ia accounted for . % of ifi, followed by . % of invasive candidiasis. interestingly, aspergillus spp. was not the major fungal pathogen in centers in thailand and vietnam, probably because galactomannan testing was unavailable. the heterogeneity of diagnostic, prophylactic, and therapeutic approaches for ifi necessitates local epidemiological data [ ] . the incidence of ifi is higher in allogeneic than in autologous hsct, and the established risk factors include chronic gvhd and steroid use [ ] . according to a recent multicenter study performed in korea (risk study), the cumulative incidence of ifi after transplantation is [ ] . therefore, active anti-mold prophylaxis should be considered in patients with those risk factors. currently, voriconazole is available in korea for secondary prophylaxis of ia after hsct, and po-saconazole for patients on significant immunosuppressive agents for gvhd. the diagnosis of ifi after hsct is not markedly different from that before engraftment. however, the computed tomography (ct) findings of invasive pulmonary aspergillosis in patients with hematologic diseases may differ depending on the time of neutropenia and the time of neutrophil recovery after hsct, suggesting that the diagnostic criteria may need to be redefined [ ] . to diagnose ifis, radiologic examinations such as chest x-ray and ct, and microbiological studies including fungal culture, and galactomannan and β-d-glucan assays should be performed periodically. for accurate diagnosis, bronchoscopy and bronchoalveolar lavage (bal) fluid examination, lung biopsy, and culture are required. voriconazole is the drug of choice for treating ia. voriconazole therapeutic drug monitoring can be used in many centers, and the target trough level is to . mg/l. the cyp c polymorphism affects the pharmacokinetics of voriconazole. among korean hematologic patients, % are extensive metabolizers (ems), % are heterozygous ems, and % are poor metabolizers. while subtherapeutic initial trough levels were common in ems, there was no significant relationship between cyp c genotype and the clinical outcomes of ia or the toxicity of voriconazole [ ] . isavuconazole is non-inferior to voriconazole for the primary treatment of suspected invasive mold disease, with fewer drug-related adverse events [ , ] . salvage therapy includes liposomal amphotericin b, caspofungin, posaconazole, and itraconazole, or a combination of them [ ] . treatment duration is at least to weeks, which should be individualized according to the changes in radiologic, microbiologic, and immunologic parameters. ifis can occur during treatment with anti-mold agents. patients with gvhd under long-term immunosuppressive therapy should be on antifungal agents to prevent ifis (table ). in particular, there is a risk of ifis other than aspergillosis, such as mucormycosis or other rare molds [ , , ] . a retrospective single-center study reported that the prevalence and incidence of voriconazole-breakthrough ifis were . % and . cases per year, respectively. the overall mortality rate was . % [ ] . the possible causes of breakthrough ifi during voriconazole treatment were persistent immunodeficiency, neutropenia, low voriconazole concentration, or poor vascular supply (i.e., abscess or necrotic tissue) [ ] . in voriconazole-refractory ia, clinicians should consider the following: misdiagnosis or coinfection with another mold, inadequate blood voriconazole concentration, inadequate tissue drug concentration, immune reconstitution inflammatory syndrome, or infection with voriconazole-resistant aspergillus [ ] . proven/probable ia patients reportedly have a low culture-positive rate ( . %) [ ] . to determine the azole-resistance rate of aspergillus clinical isolates in korea, culture-positive cases should undergo susceptibility testing. the major symptoms of pneumocystis jirovecii pneumonia (pcp) are fever, difficulty breathing, and dry cough rather than purulent sputum. typical radiologic findings are bilateral diffuse infiltrates originating from the periphery of both lungs, but may be normal at the beginning. pcp can present as segmental consolidation in the upper lobe, or subtle ground glass opacities and pneumothorax. it usually occurs within months after transplantation but also after months of immunosup-pressive therapy for chronic gvhd [ , ] . the known risk factors for pcp in hematologic patients are acute lymphoblastic leukemia, allogeneic hsct recipients, alemtuzumab, fludarabine, cyclophosphamide, rituximab, and steroid use (> mg/day prednisone for weeks) [ ] . if pcp is clinically suspected, ct, bronchoscopy, and bal should be performed as early as possible. in a retrospective study of bronchial washings or bal fluid from non-human immunodeficiency virus (hiv)-infected patients suspected of respiratory infection, among the polymerase chain reaction (pcr)-positive patients, . % were classified as pcp and . % as non-pcp. the majority of patients ( %) in the non-pcp group recovered without treatment for pcp [ ] . in two recent meta-analyses, the sensitivity and specificity of quantitative pcr (qpcr) assays were superior to those of non-qpcr assays [ , ] . the serum β-d-glucan test result has a high negative-predictive value but is not useful for the follow-up of pcp [ ] . the prognosis is worse in the presence of concomitant infectious diseases, particularly cmv, aspergillus, and so on. steroids can be used as an adjunctive therapy in patients with hypoxia (pao ≤ mmhg or pao -pao ≥ mmhg). the incidence of pcp has been significantly reduced by trimethoprim/sulfamethoxazole prophylaxis in highrisk patients. high-risk patients who do not take prophylaxis or who show poor compliance with the drug regimen are the main populations at risk of pcp. in a systematic review and meta-analysis of non-hiv immunocompromised hosts (patients with acute leukemia and recipients of hsct and solid organ transplant), the incidence of pcp was reduced by % (relative risk [rr], . ) in trimethoprim/sulfamethoxazole prophylaxis group compared with placebo [ , ] . in addition, pcp-related mortality was significantly reduced by % (rr, . ). however, trimethoprim/sulfamethoxazole prophylaxis did not markedly reduce all-cause mortality in a hematology population [ ] . mortality rates remain very high in hematology patients ( % to %), particularly in hsct recipients ( % to %), compared with % to % in patients with hiv infection [ ] . nevertheless, given the more severe course of pcp and the higher pcp-related mortality rates, trimethoprim/sulfamethoxazole prophylaxis can likely save lives in other www.kjim.org https://doi.org/ . /kjim. . immunocompromised groups as well. pcp should be prevented for at least months or until the immunosuppressant is discontinued in allogeneic hsct recipients [ , , ] . although the optimum duration of pcp prophylaxis is controversial, it is suggested to be continued for a period of time after the immunosuppressant is discontinued. in the setting of corticosteroid-containing regimens, prophylaxis should be continued while steroids are being weaned and/or for weeks after their cessation [ ] . with some chemotherapy regimens (i.e., alemtuzumab) consideration should be given to extended pcp prophylaxis for up to months because of the high rate of late-onset pcp [ ] . table summarizes the methods of preventing pcp after hsct. cmv infection can occur after hsct and can be fatal. cmv can be reactivated or the patient can be reinfected after transplantation. the spectrum of cmv infection is extensive, from cmv reactivation without organ involvement (presenting mainly as asymptomatic antigen-emia or dnaemia) to cmv disease such as esophagitis, gastritis, colitis, hepatitis, pneumonia, retinitis, and encephalitis. in addition to direct organ involvement, cmv reactivation can exert indirect effects-such as graft failure or immunosuppression-that may result in the development of concurrent bacterial and/or fungal infections [ ] . cmv disease, especially cmv pneumonia or encephalitis, can be fatal despite aggressive anti-cmv therapy [ , ] . a -year retrospective single-center study performed in korea reported that cmv disease occurred in . % of allogeneic hsct recipients and . % of autologous hsct recipients. pneumonia ( . %), retinitis ( . %), and enteritis ( . %) frequently developed in patients with cmv infection. the average time of onset of cmv disease was days after transplantation, ranging from to days [ ] . another study in the same institution reported that cmv dnaemia developed in about % of hsct recipients [ ] . cmv reactivation can be associated with a higher non-relapse mortality rate (rr, . to . ) [ ] . in another report, % of hsct recipients had experienced any level of cmv antigenemia, of whom % had received ganciclovir therapy for significant cmv reactivation and % had cmv diseases. however, this incidence could be underestimated because disease prevalence was not evaluated in all patients undergoing pre-emptive therapy [ ] . because the diagnosis of cmv disease is based on the pathology findings, the diagnosis might be limited depending on the patient's condition. particularly for the diagnosis of cmv pneumonia, obtaining lung tissue from critically ill patients by transbronchial lung biopsy is problematic. one study aimed to measure the cmv level in bronchial washing fluid and suggest a cut-off for pneumonia diagnosis. cmv dnaemia of > , copies/ml ( , iu/ml) may be associated with cmv pneumonia in post-hsct patients, as determined by the receiver operating characteristics curve [ ] . however, further data are needed because it is difficult to distinguish whether the viral dna in alveolar hemorrhage reflects viremia rather than lung tissue involvement, or whether cmv is detected as a bystander in bronchial washing fluid. management of cmv is categorized into prevention, pre-emptive treatment, and definitive treatment. pre-emptive therapy is anti-cmv treatment even in the absence of clinical symptoms in cases with cmv infection (reinfection or reactivation). most transplantation centers introduce pre-emptive therapy rather than routine universal prevention because of insurance coverage, cost-benefit ratio, and adverse drug reactions. studies of monitoring strategies and early detection have resulted in the use of cmv pp antigenemia testing and real-time qpcr for the surveillance and identification of patients suitable for pre-emptive therapy [ , ] . however, in hsct, the relationship between cmv viral load and cmv disease is different from that in solid organ transplantation. cmv gastrointestinal disease can develop without preceding cmv antigenemia or dnaemia, while > , copies/ml of cmv dnaemia is reportedly associated with an increased risk of cmv retinitis after hsct [ , ] . however, the correlations were moderate, and antigenemia or dnaemia does not necessarily precede or accompany cmv disease. therefore, it is important to identify the at-risk groups and clinical features of the various cmv diseases to facilitate early diagnosis and treatment. a recent prospective double-blind trial evaluated letermovir prophylaxis for preventing cmv in hsct recipients. a total of patients underwent randomization and received letermovir or placebo through week after transplantation. the incidence of clinically significant cmv infection was lower in the letermovir group than in the placebo group by week after transplantation ( of patients [ . %] vs. of patients [ . %], p < . ). letermovir prophylaxis resulted in a significantly lower risk of clinically significant cmv infection than placebo. adverse events with letermovir were mainly of low grade [ ] . as a result, the u.s. food and drug administration has approved letermovir for the prevention of cmv infection and diseases in adult cmv-seropositive patients undergoing allogeneic hsct. in korea, which has a high cmv-seropositive rate ( % to %) in adults, most donors and recipients are seropositive (d+/r+) and the frequency of cmv infection after transplantation is up to % [ ] . in addition, owing to active cmv monitoring and pre-emptive therapy, there has been a decrease in the incidence of cmv infection immediately after transplantation. however, late cmv infection (> months after transplantation) is increasing, especially if cmv-specific t-cell function has not been restored due to chronic gvhd. therefore, immune monitoring is required in patients with high-risk of cmv disease. cmv-specific immune recovery has been studied in kidney transplantation (kt) and hsct recipients [ ] [ ] [ ] [ ] [ ] [ ] . cmv-specific cytotoxic t lymphocytes (ctls) play an important role in the reconstitution of cmv-specific immunity in immunocompromised patients. the methods used to evaluate cmv-specific immunity include tetramer assay, intracellular cytokine analysis by flow cytometry, measurement of interferon-γ secretion by cmv-specific cd t-cells, and enzyme-linked immunospot assay using ie- and pp peptide pools [ ] . quantification of cmv-specific t-cell immunity after hsct facilitates the identification of patients at risk of cmv-related complications [ , ] . currently available anti-cmv agents in korea are ganciclovir, valganciclovir, foscarnet, and cidofovir. since february , the use of valganciclovir in hsct patients with cmv infection has increased due to changes in the reimbursement rules. regarding adverse events, attention should be paid to bone marrow suppression, and in patients with renal insufficiency. gvhd or the use of www.kjim.org https://doi.org/ . /kjim. . monoclonal antibodies (i.e., alemtuzumab) can increase the incidence of cmv infections. such infections may not be distinguishable from gvhd, or may coexist with gvhd, and may not respond to antiviral drugs if diagnosis is delayed [ ] . if there is persistent infection despite anti-cmv therapy, cmv refractoriness can be considered if a > log decrease in cmv dna level in blood or plasma is not achieved after ≥ weeks of treatment. in cmv-refractory cases, resistance should also be suspected. although cmv resistance remains uncommon in hsct recipients from hla-matched donors ( % to . %), in high-risk patients, the incidence of cmv resistance is up to . % [ ] [ ] [ ] . the gold standard of cmv resistance testing is an increase in the ic (half maximal inhibitory concentration) value by plaque reduction assay. however, cmv resistance can be defined when one or more genetic mutations associated with ganciclovir, valganciclovir, foscarnet, and cidofocir are identified with clinical refractoriness. therefore, in recent years, mutations in ul and ul can be tested. predisposing factors for cmv resistance include prolonged use of anti-cmv agents, recurrent cmv infections, inadequate antiviral absorption, subtherapeutic antiviral level, haploidentical transplantation, or t-cell depletion [ ] . if resistance is confirmed, a drug to which resistance has not been identified, or combination of drugs, may be used, but treatment is currently limited. clinician's experience is important in such complicated cases. herpes zoster is caused by the reactivation of virus latent in the posterior ganglia after primary infection with varicella zoster virus (vzv). in korea, vzv is transmitted by natural infection in most cases and its seroprevalence in adults is > %; previous vzv infection is a prerequisite for herpes zoster [ ] [ ] [ ] . the clinical features of herpes zoster include abnormal sensation or pain through skin segments to days before skin lesions develop, erythematous spots with irritation, and rapid formation of blisters. the blisters burst and form ulcers, which scar and become dry. pain and postherpetic neuralgia are major problems. in most cases, one or two dermatomes are involved unilaterally, but dissemination to several other dermatomes or systemically can occur and is associated with visceral or central nervous system involvement [ ] . treatment consists of antiviral administration and adjuvant therapy to reduce the acute pain and postherpetic neuralgia. it is not clear whether antiviral treatment is effective hours after the onset of rash. however, immunocompromised patients with persistent or new vesicular rash, eye involvement, and/or neurologic complications should receive antiviral treatment even > hours after the onset [ ] . the risk of varicella is highest in the first months after hcst, or during immunosuppressive therapy for gvhd [ ] . in a korean transplantation center, . % of patients who underwent allogeneic hsct in to experienced herpes zoster. the cumulative incidence was % at year post-transplant, and . %, . %, and . % at , , and years (fig. ) . chest dermatomes were most commonly involved ( . %), and . % of cases were disseminated. there was no herpes-zoster-related mortality. a live attenuated zoster vaccine is available but cannot be used for hsct recipients. acyclovir prophylaxis can reduce the incidence of herpes simplex virus (hsv) and vzv infections not only before engraftment but also in the long term until the immunosuppressant is stopped [ ] . it is recommended that acyclovir be maintained for at least year after allogeneic hsct and for to months after autologous hsct [ , ] . antiviral prophy- ebv reactivation can occur to months after transplantation, typically in patients with chronic gvhd. however, progression to disease is relatively rare. fever and neutropenia may occur as a result of ebv symptoms, which are similar to those of infectious mononucleosis. most ebv reactivations are subclinical and require no therapy [ ] . in addition, aplastic anemia, oral hairy leukoplakia, and post-transplant lymphoproliferative disease (ptld) can occur. ptld occurs less frequently after hsct than after transplantation of other solid organs. ebv-related ptld occurs in cases of unrelated donor transplantation, t-cell-depleted transplantation, gvhd, and use of an anti-lymphocyte antibody to prevent gvhd [ ] . the diagnosis of ebv-associated ptld can be established by tissue biopsy for histopathology and detection of ebv [ ] . among other herpesviruses, human herpes virus (hhv ) may be responsible for meningitis and hemorrhagic cystitis [ ] . hemorrhagic cystitis in hsct recipients can be classified according to the onset time (before vs. after engraftment), and cases occurring within days after transplantation are usually non-infectious. non-infectious causes include radiotherapy and chemotherapy (e.g., cyclophosphamide, ifosfamide, busulfan, and etoposide), while post-engraftment hemorrhagic cystitis can be caused by virus. viral reactivation can be accompanied by bladder urothelial damage due to the conditioning regimen. viral hemorrhagic cystitis can be caused by polyomaviruses (bk and jc viruses), adenovirus, cmv, hsv, and hhv [ , ] . of these viruses, bk virus (bkv) is the most common; % to % of adults are bkv seropositive [ ] [ ] [ ] . bkv-associated hemorrhagic cystitis is predominant in allogeneic hsct patients, and bkv-associated nephropathy in kt patients. why bkv reactivation manifests as these two major forms in kt and hsct patients is unclear [ ] . the diagnostic criteria defined by the european conference on infections in leukemia (ecil)- comprise the following triad: viral replication (urine bkv > copies/ml), symptoms of cystitis, and hematuria of grade ii or higher. a blood bkv dna level of > - copies/ml is reportedly associated with significant viruria. however, a negative plasma viral load does not rule out bkv-associated hemorrhagic cystitis [ ] [ ] [ ] . the risk factors for hemorrhagic cystitis are myeloablative conditioning regimen, atg, and gvhd. the incidence is higher in allogeneic than in autologous hsct patients, and in adults than in pediatric cases. treatment for virus-associated hemorrhagic cystitis should be initiated by minimizing immunosuppressants. cidofovir might be effective for adenovirus, cmv, and bkv, and can be considered if there is ≥ grade iii hematuria (gross hematuria with blood clots) and no response to conservative treatment [ ] . in eight retrospective and two prospective studies, the use of cidofovir with or without probenecid (either to mg/kg with probenecid or . to . mg/kg without probenecid) was investigated. a reduction in the urine and blood bkv load reportedly occurs in % to % and % to % of patients, respectively [ ] [ ] [ ] [ ] [ ] [ ] [ ] . a retrospective study did not recommend levofloxacin because of insufficient data and the low level of evidence [ ] . in addition, retrospective data are available for vidarabine, leflunomide, hyaluronic acid, and mesenchymal cells, but these are not recommended by the guidelines [ ] . brincidofovir markedly reduces bkv replication in vitro [ ] . in cases of persistent hematuria, renal function can be impaired by blood clots and related post-renal obstructions. conservative care such as intravenous hydration and removal of obstructions is also important. tients. crv infections can also occur during neutropenia and show significant morbidity and mortality. the epidemiology of crv infection in hsct patients is likely to reflect that in the community, with seasonal variations. influenza and respiratory syncytial virus (rsv) infections usually occur in winter, parainfluenza virus (piv) infections in summer, and rhinovirus throughout the year. in hsct recipients, crv infection is not limited to urtis and is more likely to progress to lrtis [ , ] . respiratory virus multiplex pcr enables rapid diagnosis of crv infections in clinical practice [ ] . there are little data on crv infection in hsct patients in korea. in a -year retrospective study in the authors' hsct center from to , of , hsct patients ( . %) had cases of crv-lrti. rsv ( . %) was the most common pathogen of crv-lrti, followed by piv ( . %), influenza virus ( . %), and rhinovirus ( . %) [ ] . the overall mortality rate at day after crv-lrti was . %, and high-dose steroid usage (> mg/kg/day), severe immunodeficiency, and lymphopenia (absolute lymphocyte count < cells/mm ) were significantly associated with mortality [ ] . these findings are similar to the prevalence, mortality rate, and mortality-related risk factors of crv-lrti in europe and the united states. the average mortality rate of rsv-lrti is % (range, % to %) in international studies, and the major risk factors for progression to lrti are lymphopenia, old age, mismatched/unrelated donor, and neutropenia [ , ] . in cases of piv-lrti, the overall mortality rate is % to %, and high-level corticosteroid exposure, neutropenia, lymphopenia, and early onset after hsct are the major risk factors for lrti [ ] [ ] [ ] . the rate of progression of influenza to lrti is % to % and the overall mortality rate is % to % [ ] [ ] [ ] . the risk factors for progression to influenza-lrti are early onset after hsct, lymphopenia, old age, neutropenia, and delayed antiviral administration [ ] [ ] [ ] . in cases of rhinovirus, most infections are asymptomatic, less than % of patients progress to definite pneumonia, and the mortality rate is < % [ , ] . in addition, human metapneumovirus, adenovirus, coronavirus, and bocavirus can cause urtis and lrtis in hsct patients [ , ] . management of crv infection in hsct patients can be classified into four categories: prevention (infection control), selective therapy for urti, antiviral therapy, and supportive care. currently, a commercialized vaccine is available only for influenza, and so preventive strategies to stop the spread of crv infections-such as hand hygiene, wearing gloves and mask, and isolation of symptomatic patients-have been emphasized [ , , , ] . it is recommended that healthcare workers and family members of patients be vaccinated against influenza, as well as undergo post-exposure prophylaxis (in the korea influenza guidelines, oseltamivir can be administered prophylactically for days if an immunocompromised patient comes into contact with a patient with confirmed influenza or influenza-like illness) [ , [ ] [ ] [ ] . because some crv-urtis do not progress to lrti and therapeutic agents are limited, it is important to identify crv-urti patients at increased risk of lrti [ , , ] . treatment of rsv-urti with aerosolized ribavirin significantly reduces the frequency of progression to lrti ( % vs. %) [ ] . in addition, the following immunodeficiency scoring index has been proposed: neutropenia (< cells/mm , points), lymphopenia (< cells/mm , points), age ≥ years ( points), myeloablative conditioning ( point), gvhd (acute/chronic, point), corticosteroids ( point), and pre-engraftment or within days of transplant ( point). a score of > is associated with a significantly increased risk of progression to lrti, suggesting the necessity of criteria for identifying high-risk groups [ ] . palivizumab, a humanized anti-rsv monoclonal antibody, does not reduce the progression to lrti in hsct patients and is currently recommended only for prophylactic usage in high-risk children [ ] . the risk factors for progression to lrti have been discussed for piv, human metapneumovirus, and rhinovirus urtis, but treatment is not recommended because of a lack of evidence [ , , ] . oseltamivir, zanamivir, and peramivir can be used to treat influenza urtis and lrtis and should be administered within hours of symptom onset [ , ] . delayed administration is also beneficial, so these agents should be administered even after hours [ , ] . although there is no definitive evidence, most guidelines recommend that antiviral treatment be extended the korean journal of internal medicine vol. , no. , march beyond the usual duration in cases of severe influenza lrti [ , , ] . aerosolized ribavirin and intravenous immunoglobulin (ivig) combination treatment significantly reduces rsv-lrti-related mortality and is thus recommended by most guidelines [ , , , ] . although evidence is lacking, if aerosolized ribavirin cannot be used, oral ribavirin or intravenous ribavirin may be considered [ , , , ] . a recent double-blind, placebo-controlled study of gs- (presatovir), an oral rsv-entry inhibitor, involved rsv-infected healthy adult volunteers who received rsv challenge intranasally. gs- reduced the viral load and the severity of clinical disease [ ] . proven piv-lrti should be treated with aerosolized ribavirin and ivig in combination [ , , ] . unfortunately, aerosolized ribavirin is costly and available only through the korea orphan & essential drug center in korea, so its use is limited. no antiviral is available to treat crv, rhinovirus, human metapneumovirus, and bocavirus lrtis, so supportive care is the mainstay. hsct patients with crv infection are often susceptible to other pathogens, and so work-up for co-existent pathogens is required [ , , ] . s. aureus and s. pneumoniae infections are commonly followed by influenza infection. therefore, patients with influenza lrtis should be administered antibiotics to which these bacteria are susceptible [ ] . as it is not possible to distinguish these bacteria from other co-pathogens (i.e., pcp, cmv, or fungi, etc.) using only imaging tests and clinical symptoms, examinations such as bronchoscopy are required [ , , ] . tuberculosis (tb) is an important opportunistic infection among hsct recipients, in whom its incidence is -to -fold higher than that in the general population; however, this is lower than in solid-organ-transplant patients [ ] [ ] [ ] . hsct recipients are immunosuppressed as a result of their hematologic disease, chemotherapy, radiotherapy, immunosuppressive agents, and gvhd [ ] . several risk factors for the development of tb after hsct have been reported, including acute myeloid leukemia, chronic myeloid leukemia, myelodysplastic syndrome, busulfan, cyclophosphamide, tbi, corticosteroid therapy, hla-mismatched transplant, gvhd, or a history of tb infection [ ] . of hsct patients who develop tb, % and % had undergone allogeneic and autologous transplant, respectively. a single-center study performed in korea reported that . % of allogeneic hsct recipients were diagnosed with tb after hsct. the median time to development of tb was days after transplantation (range, to ). the standardized incidence ratio of tb, compared with that in the general population, was . ( % ci, . to . ). extensive chronic gvhd was associated with the development of tb (p = . ) (fig. ) [ ] . another center in korea reported that hsct patients with tbi-based conditioning are likely to have tb disease [ ] . extrapulmonary tb comprises % of post-hsct tb cases, which is higher than that in the general population ( % to %) [ ] . owing to the risk of reactivation or new infection, prophylaxis is recommended for hsct recipients with a positive tb-specific interferon-γ release assay result [ ] . isoniazid is well tolerated in the post-hsct period. however, concurrent itraconazole is not recommended due to drug interactions, and the impact of voriconazole or posaconazole is unclear. isoniazid should be contin- ued for at least months until immunosuppression is reduced. rifampin may undergo drug-drug interactions with immunosuppressants, which makes this option not practical. to determine the optimum timing and duration of isoniazide prophylaxis in hsct patients, further studies are needed. post-hsct vaccination is recommended regardless of the type of transplantation and source of stem cells. guidelines recommend that both allogeneic and autologous hsct recipients should be immunized as scheduled even during immunosuppressive therapy for chronic gvhd [ , ] . however, if > . mg/kg prednisolone is administered for gvhd, it may be temporarily delayed to increase the immune response until the dosage of immunosuppressants become decreased. the recommended schedules in korea are presented in table [ ] . hsct is performed to treat various hematologic malignancies and other disorders. although survival rates have improved due to developments in transplantation techniques, infectious complications remain major causes of morbidity and mortality after hsct. the severity of infectious complications varies according to the phase post-transplant, type of transplantation, gvhd, and the degree of immunosuppression or reconstitution. further studies are required to improve 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autologous haematopoietic stem cell transplantation in paediatric patients date: - - journal: respirology doi: . /resp. sha: doc_id: cord_uid: eq vkm i background and objective: high‐dose chemotherapy (hdct) followed by autologous haematopoietic stem cell transplantation (hsct) is widely used in paediatric cancer patients, but few data about noninfectious interstitial lung disease (ild) following this treatment are available. therefore, we aimed to evaluate the incidence, clinical features and risk factors of noninfectious ild after hdct in paediatric patients. methods: this was a retrospective cohort study of paediatric solid tumour patients who underwent hdct and autologous hsct between and . ild was diagnosed using clinical symptoms and radiography after excluding cardiac, renal and infectious causes. risk factors were analysed using a cox proportional hazard regression model. results: three hundred and forty patients were enrolled, and the median age was years (interquartile range – ). eight patients ( . %) were diagnosed with noninfectious ild. the median duration of symptom onset was months (range – ). six ( %) of eight ild patients died during the study period, even though steroids were administered for treatment. high‐dose cyclophosphamide use (hazard ratio = . , % confidence interval = . – . , p = . ) and sex (hazard ratio = . , % confidence interval = . – . , p = . ) were associated with late‐onset, noninfectious ild upon multivariate analysis. conclusion: the incidence of noninfectious ild after hdct and autologous hsct was not negligible, and the clinical features of ild showed late onset and a poor prognosis. female gender and high‐dose cyclophosphamide treatment may be risk factors for noninfectious ild, but further studies with a larger number of ild patients are suggested. high-dose chemotherapy (hdct) followed by autologous haematopoietic stem cell transplantation (hsct) is widely used as a treatment for advanced or refractory paediatric solid tumours. [ ] [ ] [ ] [ ] many chemotherapeutic agents, such as alkylating agents, anthracyclines, anti-metabolites and vinca alkaloids, produce steep dose-response curves against different tumour cell types. in hdct, the dose of chemotherapeutic agents is escalated at least - times. as this treatment may produce bone marrow failure, patients receive autologous hsct to recover bone marrow function. however, concerns still exist regarding the toxic effects of hdct on other organs. toxic effects to the lung after hsct are common, and they contribute significantly to patient morbidity and mortality. as the prophylaxis and treatment of infectious complications continue to advance, noninfectious pulmonary complications have become more important. toxicity to the lung after hdct and autologous hsct has been reported, and the implicated chemotherapeutic agents include melphalan, carmustine (bcnu) and busulfan. [ ] [ ] [ ] noninfectious interstitial lung disease (ild) is one of the most recognized lung injuries that can occur in adult patients with allogenic hsct. , even though many chemotherapeutic agents (bleomycin, busulfan, chlorambucil, cyclophosphamide, methotrexate, mitomycin and nitrosoureas) are known to cause ild, few studies have investigated noninfectious ild after autologous hsct, especially in paediatric patients. therefore, we intended to determine the incidence of noninfectious ild, the clinical characteristics of patients and the risk factors associated with noninfectious ild after hdct and autologous hsct in paediatric patients. this was a retrospective cohort study conducted at a single tertiary institute. the medical records of consecutive patients (under years of age) who had a solid tumour and were treated with hdct chemotherapy and autologous hsct from april to september were reviewed. patients who had undergone additional allogenic hsct after autologous hsct were excluded (fig. ). all clinical research protocols were approved by the institutional review board of samsung medical centre (irb no. - - - ). patients received hdct according to their underlying pathology and risk group. the included conditioning regimens were carboplatin, etoposide and melphalan (cem, first hdct) or carboplatin, etoposide and cyclophosphamide (cecy, first hdct) and carboplatin, thiotepa and melphalan (ctm, second hdct) or thiotepa and melphalan (tm, second hdct) for neuroblastoma patients. for brain tumour patients, carboplatin, thiotepa and etoposide (cte, first hdct) and cyclophosphamide and melphalan (cym, second hdct) were used. the cem regimen consists of carboplatin ( mg/m for days), etoposide ( mg/m for days) and melphalan ( mg/m for day and mg/m for day). the cecy regimen includes carboplatin ( mg/m for days), etoposide ( mg/m for days) and cyclophosphamide ( mg/m for days with -mercaptoethane sulfonate sodium). carboplatin ( mg/m for days), thiotepa ( mg/m for days) and melphalan ( mg/m for day) make up the ctm regimen. the tm regimen includes thiotepa ( - mg/m for days) and melphalan ( mg/m ) with or without iodine- -meta-iodobenzylguanidine (mibg) treatment. therapeutic mibg ( or mci/kg) was administered between the first and second hdct sessions. the cte regimen entails carboplatin ( - mg/m for days), thiotepa ( - mg/m for days) and etoposide ( - mg/m for days). finally, the cym regimen consists of cyclophosphamide ( - mg/m for or days) and melphalan ( - mg/m for or days). the dosages of other hdct regimens are similar to the dosage of chemotherapeutic agents stated earlier, except for some modifications. tbi was delivered in fractioned doses ( cgy/fraction, fractions). all stem cells were collected from peripheral blood during the recovery phase of chemotherapy or during the steady state with g-csf alone. a noninfectious ild was defined when all the following were present: (i) clinical symptoms or signs of cough, dyspnea or desaturation; (ii) diffuse parenchymal infiltration on chest x-ray or computed tomography (ct); (iii) lesions not caused by pneumonia, which can be diagnosed by clinical resolution of symptoms after antimicrobial or supportive treatment and/or detection of pathogenic microorganisms in respiratory respiratory secretions were tested to identify infectious pathogens. respiratory sampling included endotracheal aspirate, bronchoalveolar lavage (bal) and nasopharyngeal aspirates, between and virus culture was performed to detect adenovirus, respiratory syncytial virus, influenza virus and parainfluenza virus, whilst multiplex polymerase chain reaction was used to detect adenovirus, respiratory syncytial virus, influenza virus, parainfluenza, coronavirus and rhinovirus after . gram stain and culture was also performed on tracheal or bronchial secretions. fungi were cultured in separate media using tracheal suction or bal fluid, and galactomannan enzyme-linked immunosorbent assay for aspergillus species was tested on serum or bal fluid. for mycobacterium, acid-fast bacillus stain, culture and dna polymerase chain reaction were performed. mycoplasma pneumonia was studied by antibody titer in blood and dna detection in respiratory secretions. pneumocystis jirovecii was evaluated by fluorescent staining of secretion smears. all lung biopsies were conducted using video-assisted thoracoscopic surgery at the acute or subacute stage of symptoms if the patients could tolerate the operation. the biopsy specimens were tested for adenovirus, pneumocystis jirovecci and cytomegalovirus by immunohistochemistry. twenty-five parameters were analysed as potential risk factors for noninfectious ild, including age at hdct; gender; underlying tumour diagnosis; use of each chemotherapy agent (cisplatin, etoposide, cyclophosphamide, carboplatin, vincristine, ifosfamide, doxorubicin, methotrexate, actinomycin d and bleomycin); use of each hdct agent (carboplatin, thiotepa, melphalan, cyclophosphamide, etoposide, busulfan and ifosfamide); mibg treatment; interleukin- usage; number of hdct treatments; total body irradiation; and thoracic field radiation therapy. the association between the development of noninfectious ild and each potential risk factor was assessed using univariate and multivariate cox proportional hazard regression models with stepwise selection. all tests were two-tailed, and p-values < . were considered statistically significant. the sas program (ver. . , sas institute, cary, nc, usa) was used for all statistical analyses. this study included a total of patients. one hundred six patients had symptoms and radiology suggestive of ild, patients of these were diagnosed as clinical pneumonia (fig. ) . in patients ( . %) out of those , pathogenic microorganisms were detected, which included respiratory syncytial virus ( ) , influenza virus ( ), parainfluenza virus ( ), gram-positive/gram-negative bacteria ( ), adenovirus ( ), mycoplasma pneumoniae ( ), coronavirus ( ), rhinovirus ( ), fungus ( ) and pneumocystis ( ) . the other patients ( . %) were diagnosed with pneumonia according to the clinical course of the disease. the median age of participants was years (interquartile range (iqr): - ). the most common primary diagnosis was neuroblastoma ( . %), followed by medulloblastoma ( . %). the number of hdct and autologous hsct treatments was in . % of patients and in . % of patients. the most common hdct regimens were cyclophosphamide with melphalan ( . %), followed by carboplatin and etoposide with cyclophosphamide ( . %), and carboplatin and thiotepa with etoposide ( . %). some patients received total body irradiation ( . %), thoracic area radiation therapy ( . %), mibg treatment ( . %) and interleukin- therapy ( . %) ( table ) . eight ( . %) out of patients were diagnosed as having noninfectious ild (fig. ) . they had no history of serious respiratory infection before diagnosing ild. the median age of these eight patients was years (range - ). the median duration of symptom onset after hdct and autologous hsct was months (range - ). all of these patients received systemic steroids as part of their treatment after ild diagnosis, and five patients (subjects , , , and ) were administered methylprednisolone pulse therapy for days. six of the eight died during the study period. ct findings were subpleural thickening or consolidation, interstitial thickening and ground glass opacity. lung biopsy showed interstitial fibrosis and/ or thickening with inflammatory cell infiltration and pleural thickening. all eight patients received highdose cyclophosphamide as a conditioning regimen before autologous hsct (table ). based upon univariate analysis, risk factors for noninfectious ild were female gender (p = . ), the use of cyclophosphamide for the hdct regimen (p = . ) and mibg treatment (p = . ) ( table ). the usual dose of chemotherapy, which includes cyclophosphamide (p = . ), cisplatin (p = . ), carboplatin (p = . ), etoposide (p = . ), vincristine (p = . ), ifosfamide (p = . ), doxorubicin (p = . ), methotrexate (p = . ), actinomycin (p = . ) and bleomycin (p = . ), was not significantly associated with the development of noninfectious ild. the multivariate analysis revealed that high-dose cyclophosphamide (p = . , hazard rate = . , % ci: . - . ) and male gender (p = . , hazard rate = . , % ci: . - . ) were associated with noninfectious ild (table ). in this study, the incidence of noninfectious ild following hdct and autologous hsct in paediatric interstitial lung disease and transplant solid tumour patients was . %. cyclophosphamide as an hdct regimen and female gender were significant predictors of the development of noninfectious ild. our patients showed a delayed onset of symptoms, and six of eight ild patients ( . %) died during the study period, even though all patients were administered steroid treatment for ild. the incidence of noninfectious ild following autologous hsct is not well known. in an adult study, the incidence of noninfectious ild following allogenic bone marrow transplant was . - . %. , , in a paediatric autologous hsct study, two of patients ( . %) showed abnormal chest x-ray, respiratory symptoms and restrictive pulmonary function tests, but infectious causes were not excluded. the incidence of noninfectious ild after autologous hsct may be different in different study groups, depending on patients' age, underlying disease or its treatment. the clinical features of our patients do not fit well with the previously reported syndromes of noninfectious pulmonary complications after hsct. idiopathic pneumonia syndrome (ips) is one common pulmonary complication after hsct. , the american thoracic society defines ips as follows: (i) widespread alveolar injury; (ii) the absence of infection; and (iii) the absence of cardiac, renal or iatrogenic aetiology, which is similar to our definition of noninfectious ild except that it limits the injury site to the alveoli instead of the entire lung parenchyma. the mortality of ips patients is - %, which is similar to that of our patients. [ ] [ ] [ ] because pulmonary complications follow a predictable timeline after hsct, an onset time of a median of - days (range: - days) after hsct is one major difference, in that all of our patients presented with symptoms after more than days, and six of our patients developed symptoms after more than months. , , delayed pulmonary toxicity syndrome is another form of mild-to-moderate pulmonary injury that can occur after hdct and autologous hsct in breast cancer patients. [ ] [ ] [ ] patients with delayed pulmonary toxicity syndrome develop symptoms with a mean onset of weeks after hdct containing cyclophosphamide, cisplatin and bcnu. unlike our patients, they show good responses to steroid treatment and good prognosis. many drugs are known to cause ild, and % of all ild can be attributed to drug-induced ild. the incidence of drug-induced ild has not been well known, but the incidence density of . per million person-years was reported in a study over a -year period ( - ) , and the time trend in incidence was not statistically significant. cyclophosphamide is known to cause drug-induced ild, and the clinical and pathologic patterns of ild are subacute ild/ non-specific interstitial pneumonia, pulmonary fibrosis, acute respiratory distress syndrome and organizing pneumonia. drug-induced ild, including that produced by cyclophosphamide, may not only develop during the administration of the medication but it can also present several years after treatment. late-onset pulmonary fibrosis was identified in two children treated with cyclophosphamide and has been reported up to years later in children who received bcnu. because the diagnosis of druginduced ild is made by temporal eligibility and exclusion of other causes, it is difficult to diagnose patients who develop symptoms after finishing the medication. in this study, cyclophosphamide was associated with an . -times increased risk of ild, and it raises a high suspicion for cyclophosphamide having a causative role in late-onset drug-induced ild. depletion of reduced glutathione and impaired antioxidant defences might be suggested as possible mechanisms. according to the univariate analysis conducted in this study, mibg treatment and female gender were ild risk factors. mibg shares the same uptake and release mechanism as norepinephrine, and it is used for the detection of neuroendocrine tumours or targeted radiotherapy for advanced neuroblastoma. , i-mibg is also taken up by the lung through energyrequiring, sodium-dependent transport mechanism, so mibg scintigraphy is used to assess pulmonary endothelial cell injury. in i mibg mega-therapy groups, transient interstitial pneumonia and fatal pneumonia were reported, but the risk of pulmonary complications warrants further investigation. even though gender was a risk factor in this study, the specific mechanism for this trend is not known. gender-based differences in bleomycin-induced pulmonary fibrosis have been reported in an animal study. male gender was also determined to be a risk factor for ild in nonsmall-cell lung cancer patients who received gefitinib treatment. the gender effect of the development of ild after autologous hsct needs to be studied. total body irradiation (tbi, > cgy) among patients older than years is also known to produce a high risk for ips. however, tbi was not associated with ild in this study, which may have been due to the young age of our patients or the lower dose of tbi ( cgy/fraction, fractions) that was used. bleomycin and methotrexate, which are both drugs that are known to cause ild, were used at conventional chemotherapy doses in this study group, but they had no significant association with noninfectious ild. our large study number of autologous hsct (n = ) patients allowed us to analyse the risk factors associated with noninfectious ild after autologous hsct, which has previously only been anecdotally reported or investigated only in regard to one individual therapeutic agent or individual disease cases. long follow-up periods (median . years) allow for the detection of late-onset noninfectious ild. there were some limitations in this study. we could not obtain pulmonary function tests as screening tests because most patients were too young to perform them. additionally, approximately % of ild patients may not present with abnormal chest x-rays. therefore, some ild patients with minimal symptoms or with a normal chest x-ray may not have been detected using this study definition, so the incidence could be higher than . %. for this reason, caution should be used when interpreting any negative results for suspected risk factors in this population. conclusively, this study presents the incidence of late-onset noninfectious ild after autologous hsct in paediatric solid tumour patients. female gender and high-dose cyclophosphamide treatment might be risk factors for noninfectious ild, but further studies with a larger number of noninfectious ild patients are suggested. high-dose sequential chemotherapy and autologous stem cell reinfusion in advanced pediatric solid tumors high-dose melphalan, etoposide, total-body irradiation, and autologous stem-cell reconstitution as consolidation therapy for high-risk ewing's sarcoma does not improve prognosis high dose melphalan in the treatment of advanced neuroblastoma: results of a randomised trial (ensg- ) by the european neuroblastoma study group myeloablative megatherapy with autologous stem-cell rescue versus oral maintenance chemotherapy as consolidation treatment in patients with high-risk neuroblastoma: a randomised controlled trial dose: a critical factor in cancer chemotherapy high-dose therapy with stem cell rescue for pediatric solid tumors: rationale and results pulmonary complications of bone marrow transplantation the spectrum of noninfectious pulmonary complications following hematopoietic stem cell transplantation melphalan-associated pulmonary toxicity following high-dose therapy with autologous hematopoietic stem cell transplantation pulmonary toxicity syndrome in breast cancer patients undergoing bcnucontaining high-dose chemotherapy and autologous hematopoietic cell transplantation high-dose busulfan and the risk of pulmonary mortality after autologous stem cell transplant late-onset noninfectious interstitial lung disease after allogeneic hematopoietic stem cell transplantation incidence, outcome, and risk factors of late-onset noninfectious pulmonary complications after unrelated donor stem cell transplantation interstitial lung disease induced by drugs and radiation risk factors and outcome of pulmonary complications after autologous hematopoietic stem cell transplant long-term pulmonary sequelae after autologous bone marrow transplantation in children without total body irradiation noninfectious pulmonary complications after hematopoietic stem cell transplantation: practical approach to imaging diagnosis idiopathic pneumonia syndrome after bone marrow transplantation an official american thoracic society research statement: noninfectious lung injury after hematopoietic stem cell transplantation: idiopathic pneumonia syndrome clinical course of idiopathic pneumonia after bone marrow transplantation idiopathic pneumonia syndrome: changing spectrum of lung injury after marrow transplantation risks and outcomes of idiopathic pneumonia syndrome after nonmyeloablative and conventional conditioning regimens for allogeneic hematopoietic stem cell transplantation incidence and outcome of idiopathic pneumonia syndrome in pediatric stem cell transplant recipients delayed pulmonary toxicity syndrome following high-dose chemotherapy and bone marrow transplantation for breast cancer pulmonary drug toxicity in patients with primary breast cancer treated with high-dose combination chemotherapy and autologous bone marrow transplantation pulmonary toxicity of induction chemotherapy prior to standard or high-dose chemotherapy with autologous hematopoietic support comparison of registries of interstitial lung diseases in three european countries drug-/ radiation-induced interstitial lung disease in the united kingdom general population: incidence, all-cause mortality and characteristics at diagnosis lung toxicity associated with cyclophosphamide use. two distinct patterns late-onset pulmonary fibrosis and chest deformity in two children treated with cyclophosphamide active lung fibrosis up to years after chemotherapy with carmustine (bcnu) in childhood metaiodobenzylguanidine as an index of the adrenergic nervous system integrity and function megatherapy combining i( ) metaiodobenzylguanidine and high-dose chemotherapy with haematopoietic progenitor cell rescue for neuroblastoma involvement of pulmonary endothelial cell injury in the pathogenesis of pulmonary fibrosis: clinical assessment by i-mibg lung scintigraphy age and sex dimorphisms contribute to the severity of bleomycin-induced lung injury and fibrosis predictive factors for interstitial lung disease, antitumor response, and survival in non-small-cell lung cancer patients treated with gefitinib normal chest roentgenograms in chronic diffuse infiltrative lung disease key: cord- - zaoxn authors: nan title: publication only date: - - journal: bone marrow transplant doi: . /sj.bmt. sha: doc_id: cord_uid: zaoxn nan ischemic myocardial damage is an increasing cause of heart failure in the western world and has long been considered irreversible because adult cardiomyocytes are terminally differentiated and do not proliferate. stem cells are undifferentiated cells capable of self-renewal, proliferation, and differentiation into multiple lineages permitting tissue regeneration. a number of types of stem cells are now recognised, as well as partially differentiated progenitor cells that are capable of proliferation and differentiation to multiple lineages. reversal of heart failure would require myocardial revascularization, remodelling of the left ventricle and replacement of damaged myocyte. we investigated the safety of transplanting un-manipulated autologous bone marrow into infracted myocardium in two patients. these patients underwent coronary bypass using the pi-circuit technique and external reshaping of left ventricle in off-pump surgery. we evaluated the efficacy of this combined technique in the improvement of cardiac function. autologous bone marrow ( ml) was obtained by bilateral posterior iliac bone aspiration at the time of surgery. bone marrow mononuclear cells were isolated by means of a density ficoll-paque gradient. then the cells were exhaustively washed and resuspended in a normal saline solution containing % human serum albumin. cell count, viability and cultures were appropriately performed. following the operation the bone marrow mononuclear cells ( ml) were injected directly to the myocardium of the left ventricle. no significant complications were observed. the left ventricular ejection fraction at rest was improved significantly in both patients from and % to and % respectively, three months following the operation. furthermore, we observed significant reduction of the end diastolic volume of the left ventricle and improvement in the inferior-posterior wall motion, this area was not revascularized, in comparison to the previous one before the operation. these findings suggest that transplantation of unmanipulated autologous bone marrow into scar tissue of the human heart is safe and enhances cardiac function, when used in combination with myocardial revascularization and remodelling of the left ventricle. this benefit can be seen after months of the bone marrow transplant and is maintained after months of follow-up. a. symeonidis, m. tiniakou, a. spyridonidis, m. karakantza, e. triantafyllou, a. kouraklis-symeonidis, v. pesli, p. matsouka, n. zoumbos univers of patras med school (patras,gr) the haematopoietic sct unit in patras university was established in , aiming to cover the requirements of the area of peloponese and south-western greece. since then autologous and allogeneic stem-cell transplants have been performed in patients. auto-transplanted patients were male and female (median age , range - years), diagnosed as non-hodgkin's lymphoma (nhl= ), multiple myeloma (mm= ), hodgkin's lymphoma (hl= ), acute myelogenous leukemia (aml= ) chronic myelogenous leukemia (cml= ) and chronic lymphocytic leukemia ( ) . nine mm patients received tandem auto-transplants, while auto-and allo-were second transplants. at transplantation patients were in cr, in pr and had refractory disease. bone marrow (bm) grafts were used in cases and peripheral stem-cell grafts (pbsc) in . patients were conditioned with melphalan ( ), cbv ( ), beam ( ), busulfan-based regimens ( ) and thio-tepa-based regimen ( ). engraftment was achieved in / cases. three patients died early post-transplant. after a median follow-up of months, patients are alive ( %), of them disease-free, and have died, due either to transplant-related complications ( ), or after an early (< year post-transplant) (nhl= , aml= , mm= ), or a late relapse (> years post-transplant) (mm= , hl= , nhl= ). for all allo-transplants the donor was a matched sibling. patients were male and female (median age , range - years), diagnosed with aml ( ), acute lymphoblastic leukemia (all= ), myelodysplastic syndrome (mds= ), aplastic anemia ( ), cml ( ) and myelofibrosis (mf= ). excluding patients with aa, disease status at transplantation was st cr for patients, nd cr for , and active residual disease for four. stem-cell source was bm in and pbsc in . a standard conditioning was used in cases and a reduced intensity one in . engraftment was achieved in all cases. one patient died early, due to uncontrollable vod, and (all= , aml= ) due to disease progression, which was early (< months) in cases, and late (at and months post-transplant) in . five patients developed acute gvhd grade i-ii, and one grade iv. after a median follow-up of months, twelve patients are alive and disease free (aml= , mds= , aa= , cml= , mf= ) but of them have manifested chronic gvhd, (extensive in ). in one patient chronic gvhd emerged after dli, administered for smoldering relapse. activation of plts by collagen, measurement of hypotonic shock response (hsr) and extent of shape change (esc) were tested in addition. in os-pc and c-pc plt yield was . ± . and . ± . x /unit (recovery . % and . %, p= . ), residual erythrocytes were < . x /unit, residual leucocytes < . x /unit in all pc. on d values for po in os-pc and c-pc were . ± . and . ± . mmhg, pco . ± . and . ± . mmhg (p< . ), hco . ± . and . ± . mmol/l (p< . ), respectively. on d values for po in os-pc and c-pc were . ± . and . ± . mmhg, pco . ± . and . ± . mmhg (p< . ), hco . ± . and . ± . mmol/l, respectively. results (mean±sd) of metabolic and activation markers and morphologic features on d and d are shown in the table. in both groups functional parameters revealed a sufficient capacity for aggregation during storage. compared to c-pc os-pc were significantly more efficient in recovery of plts, whereas cd p and cd were significantly higher due to a higher extent of plt activation in the automatic system. a possible difference in clinical outcome of transfusion of os-pc compared to c-pc has to be investigated in further studies. objectives: recently the investigation of differentiation potential of bone marrow stromal cells becomes important because of their implementation in transplantation. but the structure of the stromal cells pool is still not fully known. the aim of the study -to investigate the differentiation ability of bone marrow clonogenic fibroblasts and to elaborate the standart assays for clonogenic proliferation and adipogenic and osteogenic differentiation (ad and od) of bone marrow fibroblasts in children. methods: the material -bone marrow aspirates from healthy donors, patients with acute lymphoblastic leukemia (all) and patients with acute myeloblastic leukemia (aml). bone marrow stromal fibroblasts were cultured accoding a.fridenstain in our modification. od was induced by: b-glycerophosphat x - m; dexametasone x - m; ascorbic acid x - m. for ad were used: dexametasone x - m and insulin x - m. results: optimal conditions for maximum cloning efficiency were following: isolation bone marrow mononuclear cells; density by explantation - x /ml cells; culture medium: medium with % human serum of any blood group. osteogenic induction increased the proportion of fibroblasts colonies from normal bone marrow with alkaline phosphatase activity from , % to , %; the adipogenic inductors increased the proportion of colonies with lipid-rich vacuoles (detected by sudan) from , % to , %. the cloning efficiency of stromal precursors (the number of colonies per x explanted cells) in the patients with aml did not differ from normal donors ( , and , ) and was much lower in all patients ( , ). stromal cells from leukemic patients showed decreased (in comparison with normal donors) potency for od in the presence of inductors (from , % to , % for aml; from , to , for all patients). the ability of stromal cells to ad did not differ in patients with acute leukemias and normal donors. conclusion: standardization of stromal fibroblasts clonogenic cultivation assay is necessary for the evaluation of bone marrow stroma state. the using of osteogenic and adipogenic inductors demonstrate the possibility of in vitro differentiation of stromal progenitors and showed the differences between normal and leukemic stromal cells. initiation of leukapheresis for peripheral blood stem cell collection at a low level of circulating cd + cells l.k. tan, t.g. soh, b. tan, j. mah, t.c. liu, c.s. chen, p. law national university hospital (singapore, sgp) objectives: most patients or donors undergoing leukapheresis (lp) for autologous or allogeneic peripheral blood stem cell (pbsc) collection require multiple lp to achieve a sufficient cd + cell dose (e.g., ≥ . x /kg). lp is initiated when peripheral blood (pb) cd + reached a certain level (e.g., ≥ per microliter). the aim of this retrospective analysis is to summarize our institutional experience of initiating lp at a lower pb cd + cells level of per microliter and to investigate the merits of this practice. methods: all patients or donors underwent lp (using cobe spectra or baxter amicus) processing times the blood volume. a total of procedures ( autologous and allogeneic) was performed in patients or donors between jan and oct . autologous patients were mobilized with chemotherapy and g-csf ( mcg/kg) while allogeneic donors with g-csf ( mcg/kg) alone. a "good" lp is defined as having ≥ x cd + cells/kg in the collection so that a minimum dose of x /kg can be achieved in sessions. cd + cells were measured by sequential gating (staining with antibodies against cd and cd together with forward and side-scattering). results: each lp contained . x wbc/kg (median, range: . - . ) and . x cd + cells/kg (median, range: . - . ). cd + cells/kg in lp were correlated to pb cd + cell counts (r = . ). as shown in table , initiating lp at higher levels of pb cd + cell (≥ per microliter) increased the proportion of "good lp", whether "all" collections or only the first collections were considered. however, a substantial number of "good lp" (≥ %) would be missed if lp was initiated at or cd + cells per microliter in pb (table ), but almost none at cd + cells per microliter. conclusion: the result demonstrated that initiating lp at pb cd + cells per microliter is helpful to some patients /donors. additional criteria may need to be adopted to determine whether lp should be discontinued if mobilization is adequate to minimize resource utilization. g-csf mobilised peripheral blood progenitor cells (pbpc) grafts are widely used in haematopoietic transplantation, but the effect of this cytokine on the cellular content of the graft is not fully understood. apart from the cd +cells, t lymphocytes, nk cells and dendritic cells present in pbpc grafts play a crucial role in the haematopoietic and immune reconstitution following allogeneic transplantation. the aim of this study was to evaluate and compare cell populations present on g-csf mobilised pbpc collected on the th day of mobilisation and on peripheral blood (pb) prior to mobilisation of healthy adult donors with a median age of (range - ). the cell populations studied were t lymphocytes (cd and cd ), nk cells (bright and dim) and dendritic cells (myeloid and lymphoid). total nucleated cells (tnc) were determined by haematological counters, cell phenotypes were evaluated by flow cytometry using colour staining, and each cell subset was determined using specific markers. t lymphocytes were divided in cd +cd + or cd +cd +; nk cells into nkdim (cd -cd +) and nkbright (cd cd ++); dendritic cell were considered lin-and hla-dr++ and then divided into lymphoid (ldc) cd ++ (high) and myeloid (mdc) cd c+. in both pb and pbpc collections t lymphocytes were the main cell population present. with the mobilisation procedure there was a variable fold increase of cell populations (table ), namely fold increase for tnc, while t lymphocyte, dc and nk populations showed , and fold respectively. despite the differences in number, the t lymphocyte, dc and nk sub-populations kept the same ratio. there was a significant correlation between the pb tnc concentration and the fold increase of tnc, cd + cells and nk cells in the pbpc graft. there was no correlation with the increase in dc and any pb parameter evaluated. in summary, g-csf mobilisation while increasing the tnc number (up to fold) does not affect nk, dc and t lymphocytes sub-populations ratio in the grafts from healthy adult donors. further studies are required to determine if the development, maturation and functional activity, namely cytotoxic and immune capacity of these cells are affected by the procedure. toxicity and efficacy of donor lymphocyte infusion after haematopoietic stem cell transplantation s. roncon, m. bini-antunes, f. campilho, i.l. barbosa, a. avila, s. ferreira, h. leal, c. pinho vaz, a. campos, r.b. ferreira, p. pimentel, a. carvalhais instituto portugues de oncologia -crop (porto, p) relapse remains one of the main complications after allogeneic haematopoietic stem cell transplantation (hsct). donor lymphocyte infusion (dli) is a therapeutic approach that is able to mediate antitumour effects and restore prolonged remissions. this antitumour effect has been well established in chronic myeloid leukaemia (cml) and less in other malignant diseases. we retrospectively analysed patients ( f/ m) who relapsed or were at high risk of relapse after hsct between december and september . patients median age was years ( - ). they had the following diagnosis: cml , acute myeloid leukaemia (aml) , acute lymphoid leukaemia (all) , multiple myeloma (mm) , hodgkin disease (hd) , aplastic anemia (aa) , non-hodgkin lymphoma (nhl) , myelodisplastic syndrome (mds) . graft source was g-csf mobilized peripheral blood in patients and bone marrow in . conditioning was myeloablative in patients (in of them graft was t cell depleted in vitro) and of reduced intensity (ric) in the remaining patients. eight patients underwent a nd allogeneic transplantation and of them later received dli. the indications for dli were relapse/disease progression in patients and pre-emptive in . the median interval between transplantation and the first dli was months . the number of dli was one for patients, two for and three for patients. the median dose of cd + lymphocytes infused was x /kg ( , - x /kg). a complete response was observed in % of patients ( / acute leukaemia, / mm, / lmc) and a partial response in ( mm and smd). there was no remission in % of patients ( / hd, / aa, / cml, / la, / mm). in two patients there was no available information. all the five patients relapsed after prophylactic dli. eight patients developed graft versus host disease (gvhd) "de novo" after dli. seven patients with cml did not achieve molecular remission after dli but did it when imatinib mesylate was associated. at the last follow up, patients remained alive and of them established full donor chimerism. in summary, complete remissions induced by dli were inferior to reported in literature. gvhd developed just in an acceptable number of cases and was controlled. further studies need to be performed to evaluate which patients really benefit from dli. report of stem cell transplantation in solid tumours in iran k. alimoghaddam, n. mahdavi, s. samiee, m. jahani, a. mousavi, a. ghavamzadeh horc (tehran, ir) introduction: the prognosis for many pediatric and young adult patients with solid tumors that have metastasized at the time of diagnosis or have relapsed after therapy remains very poor the lack of efficacy of chemotherapeutics, radiotherapy and cytokine-based immunotherapy for many patients with metastatic cancers has catalyzed, at least in part, enthusiasm for exploring allogeneic-based immunotherapy against solid tumors. while there is little doubt about the potential for its application in oncology, extensive use of nonmyeloablative hematopoietic cell transplantation (nmhct) in the treatment of solid tumors will likely remain limited until both the safety and efficacy of the approach are improved. materials and method: we collect the data of patients who had undergone stem cell transplantation by reviewing their records from the date of their transplantation up to the date of last contact. analysis of the data has done via using spss software. results: of all the patients ( . %) were male, and ( . %) were female. the main diagnoses before transplant are brought in the following table. (table- ) the median age was years old; ( - y/o) respectively. ( . %) patients received autologous and ones ( %) received allogeneic stem cell transplantation. ones ( . %) had peripheral blood and ( %) ones had bone marrow as graft type. the median duration of hospitalization for autologous transplanted patients was days which was days for allogeneic transplanted ones. transplant mortality rate in the first days was %. the median follow-up duration was days, with minimum and maximum of and days respectively and during this period the overall survival (os) is . % and the disease free survival rate (dfs) was . %. palifermin for oral mucositis prophylaxis in autologous transplantation n. miranda ( ) , p. santos ( ), t. mendonça ( ) , i. ferreira ( ) , a. guimarães ( ) , j.l. , m. abecasis ( ) ( )instituto português de oncologia (lisbon, p); ( )hospital egas moniz (lisbon, p) objectives: oral mucositis is a near universal complication of intensive chemotherapy. the patient discomfort often require i.v. narcotics and is the main cause for total parenteral nutrition. some authors have associated the duration and the severity of mucositis with transplant outcomes. until very recently no effective treatment or prophylaxis was available. palifermin is the recombinant human keratinocyte growth factor and a previous study have shown its ability to decrease the length and severity of mucositis in conditioning regimens including total body irradiation (tbi). from august to november we have used palifermin for the prophylaxis of mucositis in adult patients (pts) under autologous bmt with conditioning regimens without tbi. methods: nine patients ( females and males) with a median age of . (range - ) have been included in an open label study included in an extended access program. the predominant diagnosis was non hodgkin's lymphoma ( ). the conditioning regimen was beam in , vp + melphalan in and carboplatinum + vp + thyotepa in . palifermin was given according to the manufacturer instructions micrograms/kg body weight on days before chemotherapy and on days starting on day after stem cell infusion. we analysed severity and length of mucositis, narcotic need, haematological reconstitution and side effects of palifermin. we have compared the results with a historical cohort matched for diagnosis, age, conditioning regimen and number of cd + cells transplanted. results: there was only one case of discontinuation of treatment due to toxicity (generalized skin oedema with severe hypotension). severe pruritus was present in pts and generalized rash in all but one patient. a statistical significant increase in weight was observed in pts treated with palifermin on day + (median + kgs versus - kgs without palifermin). the number of days under iv narcotics was lower on palifermin group ( . vs . ) and more patients did not require narcotics at all ( out of nine versus in ). the mean of the number of days without any oral nutrition was lower on palifermin group ( . versus . ). all this differences did not reach statistical significance. the haematological recovery, antibiotic need, bacteriological isolates and length of stay on hospital were equivalent in both groups. conclusion: in our series palifermin decrease the severity and duration of mucositis and was associated with significant skin toxicity. clinical experiences with the new keratinocyte growth factor palifermin in patients treated with high-dose chemotherapy followed by autologous stem cell transplantation m. fillitz, e. koller, e. schlögl, e. pittermann-höcker hanusch-hospital (vienna, a) oral mucositis is a common side effect of chemotherapeutic and/or radiotherapeutic treatment in malignant diseases resulting in pain, diarrhoea, malnutrition, gastrointestinal bleeding complications, local and systemic infection leading to prologation of hospitalization duration and increased medication and treatment costs. this side-effect of cancer treatment has to be expected in up to % of all treated cancer patients and the percentage is even higher in the hematopoietic stem cell transplantation setting. as the functional role of different cytokines such as tumor necrosis factor-alpha and different interleukines and probable protective factors like secretory iga in the pathological pathway leading to mucositis is not yet well understood, neither a widely accepted prophylaxis nor a therapy is available. we report our data of prophylactic application of the recombinant human keratinocyte growth factor palifermin in autologous stem cell transplantation recipients(female ; male ). in cases diagnosis was relapsed non-hodgkin´s lymphoma, patients were suffering from multiple myeloma. the patients age ranged from to years, karnofsky status was % in all. palifermin was given as intravenous bolus injection on consecutive days , third dose at least hours before administration of high dose chemotherapy. second cycle consisted of another doses that were given on consecutive days starting with the day of reinfusion of the stem cell harvest product. the single injection doses ranged from . to . mg according to the producer´s mcg/kg/d recommendation. we experienced only one case of who-grade -mucositis which could not be distinguished from mucosal hyperproliferation due to the medication, all other patients showed no sign of oral mucositis. diarrhoea was maximum who-grade . treatment related side effects consisted of mucosal oedema (vagina, tongue, palate, lids), erythema of the face and upper body,dysphagia and disturbances of taste and sensibility. in one patient excessive proliferation of the oral mucosa lead to detachment from palate and tongue, in another patient mild temporary dyspnoea occurred. altogether those findings were well manageable and resolved without additional measure within days after last injection. we observed no septic event. the hospitalization duration was slightly reduced to former comparable patients even though the first cycle of palifermin led to a days prologation of the pre-transplant phase. prevention of oral mucositis in patients undergoing radiochemotherapy and allogeneic stem cell transplantation with recombinant human keratinocyte growth factor (palifermin): a single-centre experience w. rabitsch, p. kalhs, w. köstler, s. wöhrer, a. schulenburg, v. supper, m. mitterbauer, h. greinix bone marrow transplantation (vienna, a) objectives: oral mucositis is one of several common adverse effects of myeloablative therapy. it is particulary frequent in patients receiving high-dose chemotherapy with or without total-body irradiation for hematopoietic stem cell transplantation (hsct). palifermin is the first agent to be approved for the prevention of oral mucositis induced by myelotoxic therapy. so far, data on ist use in allogeneic hsct are rare. patients and methods: we analyzed the efficacy of palifermin on consecutive patients (median age: , range - years) undergoing allogeneic peripheral blood stem cell transplantation (sibling donor; n= ; unrelated donor, n= ) at our institution. all patients received a conditioning therapy with cyclophosphamide and total body irradiation. palifermin was administered intravenously in a dosage of µg/kg/day days before myeloablative therapy and days after stem cell infusion. mucositis was assessed daily and graded according to the who-scale. results: one patient died on day + after transplantation due to gram-negative sepsis and multi-organ failure. the other patients were eligible for assessment of mucositis. the eligible patients experienced only who grade mucositis. the drug was generally well tolerated without severe side effects. one patient developed erythema on the face, which resolved -hours after palifermin administration without the need of medication. in the other patients no side effects were observed. only patient experience acute graft-versushost disease (gvhd) of the skin grade ii. currently, of patients are alive, , and months after hsct. conclusion: palifermin represents the first drug approved by the us fda for reduction of incidence and duration of oral mucositis in a specific patient cohort. in our cohort of patients we observed no serious side effects and only low grade mucositis. the effect of palifermin on gut epithelia or gvhd incidence and severity has to be assessed in larger patient cohorts. pegfilgrastim in comparison with filgrastim after allogeneic stem cell transplantation c. lutz, g. massenkeil, i. tamm, t. terwey, s. neuburger, b. doerken, r. arnold university hospital charité (berlin, d) purpose: after autologous and allogeneic peripheral blood stem cell transplantation (pbsct) filgrastim (g-csf) is given daily to enhance neutrophil recovery and prevent risk of infection. peg-filgrastim (polyethylen-glykol-g-csf) has been approved in . since the clearance of peg-filgrastim is mediated by neurophils, in neutropenia activity and serum concentration is prolonged and elevated. the aim of this study was to evaluate for the first time the efficacy of a single fixed dose ( mg) in patients after allogeneic pbsct from matched unrelated donors. methods: on day + after allogeneic pbsct five patients ( aml, all, cml; median age y) received mg peg-filgrastim s.c. the neutrophil and platelet recovery was compared to patients (n= ; all, aml, mds, cml, mm; median age , y) treated with daily filgrastim ( µg/kg i.v. over h). definition of neutrophil recovery was ≥ , /nl, platelet recovery ≥ /nl on three consecutive days and without transfusions respectively. g-csf serum levels were measured by elisa (r&d systems). results: peg-filgrastim was well tolerated. in patients treated with peg-filgrastim the neutrophil engraftment was days versus days in the filgrastim group. the platelet recovery in the peg-filgrastim group was days and days in the filgrastim group. these differences were not significant. g-csf levels were measured after the first administration of filgrastim or peg-filgrastim respectively. the results showed a neutrophil mediated clearance of peg-filgrastim with a peak serum level on day + that dropped in parallel to neutrophil recovery. the peak level of peg-filgrastim was about ten fold higher than the peak level of filgrastim ( ng/ml vs ng/ml). conclusions: our data suggest that a fixed dose of mg peg-filgrastim used after allogeneic pbsct is effective and gives comparable results to daily administration of filgrastim. larger prospective studies are needed. use of pegfilgrastim after high-dose melphalan and autologous peripheral blood stem cell transplant in multiple myeloma patients ( ) , p. iacopino ( ) ( )azienda ospedaliera bmm (reggio calabria, i); ( )policlinico universitario (messina, i) single dose of pegfilgrastim is equivalent to daily filgrastim after standard dose chemotherapy in decreasing the duration of neutropenia. daily filgrastim started within - days after autologous peripheral blood stem cell transplant (apbsct) leads to decrease in time to neutrophil engraftment. we undertook a study of pegfilgrastim after high-dose melphalan (hdm) and apbsct. in all, patients with multiple myeloma (stage iii durie-salmon classification), eligible to undergo hdm and apbsct, were enrolled. patients were conditioned with hdm at a dose of mg/m² intravenously on day - .the day of apbsct was termed day . the median stem cell dose infused was x /kg (range - ). patients received a single dose of mg pegfilgrastim subcutaneously h after apbsct. there were no adverse events secondary to pegfilgrastim. neutrophil engraftment was defined as first of consecutive days of an anc equal or greater than . x /l after a previous nadir. platelet engraftment was defined as platelet count of equal or greater than x /l. all patients engrafted neutrophils and platelets with a median of days (range, - ) and days (range, - ), respectively. the incidence of febrile neutropenia was . % ( / ). the median duration of febrile neutropenia was days (range, - ). the mean number of platelet and packed red blood cell transfusions were + . u and . + . u. it's interesting to note that / patients didn't required packed red blood cell transfusion. one patient died for cerebral bleeding after engraftment, on day + ; patients experienced persistent reversible neurophilia. neutrophil and platelet engraftment were compared with a cohort of patients (same diagnoses, method of stem cell collection, conditioning regimen and stem cell dose) treated at our institute, who received filgrastim at mg/kg subcutaneously daily, starting at day + or day + and no statistical difference were shown (p= ns). in conclusion, pegfilgrastim given as a single fixed dose of mg appears to be safe after hdm and apbsct. pegfilgrastim may be convenient to use in outpatient transplant units. the concept of treating chemosensitive leukemia, lymphoma and myeloma patients with high dose chemotherapy followed by reinfusion of peripheral stem cell has been implemented in the set-up of bone marrow transplantation, and often considered as transplantation procedure. with the use of growth factors to stimulate bone marrow and allow the harvest of peripheral stem cells with an apheresis machine haemoneticsr mcs p and cryopreservation, the procedure of autologous peripheral stem cell transplant (apst) became feasible in hematology departments, and not necessarily in bone marrow transplant units. for this reason, we decided eight years ago to set up such a system in our institute. our team included three md hematologists, one nurse and one apheresis technician. we equipped our institute with the necessary separator machine and after a period of ten months trial of investigation of the harvest quality, we started to treat our patients in our institute, instead of referring them to a transplant unit. our institute is a part of a -bed university hospital and covers a population of inhabitants. in this abstract we would like to present our results. using two haemonetics, approximately harvests have been performed with achievement of an excellent yield of cd stem cells (median, . x /kg). the harvest was performed at the day care unit and then cryopreserved in liquid nitrogen. in most patients myeloablative chemotherapy was also given ambulatory. following the reinfusion of the harvest, patients were hospitalized in isolation rooms. the median time of recovery (neutrophils > . x /l) was days. at the end of the year (eight years activity), auto-transplantations were performed, including tandem. the diagnoses were: multiple myeloma in , non-hodgkin's lymphoma , hodgkin's disease , aml , all , amyloidosis and cll cases. treatment related mortality was . %. we believe that such endeavor should be encouraged and advised for more hematology centers. hematologists in training and senior hematologists have the benefit of keeping their patients under close supervision with the challenge of further therapies, increasing the clinical level, the motivation and the interest in the field of hematooncology. h. dimitriou, e. linardakis, g. martimianaki, e. stiakaki, a. fillipidi, m. kalmanti university of crete (heraklion, gr) mesenchymal stem cells (mscs) are multipotent progenitor cells within the bone marrow (bm) capable of differentiating into various tissue specific cells. mscs form an integral part of the bm stroma, have immunomodulatory functions and play an important role in the support of hematopoiesis. their multipotentiality and ease of ex vivo expansion has raised great interest in the clinical use of mscs for tissue repair and gene therapy. in order to evaluate if malignant and non malignant hematological diseases quantitatively and qualitatively affect bm derived mscs, bone marrow from children with acute lymphoblastic leukemia (all diagnosis n= , different phases of treatment n= , end of therapy n= ), idiopathic thrombocytopenic purpura (n= ), autoimmune neutropenia (n= ) and control patients (solid tumors without bm involvement, n= ) was harvested and the mononuclear cell (mnc) fraction isolated. mscs were expanded in amem supplemented with % selected fcs, characterized and compared in terms of their phenotypic characteristics, clonogenicity and ability to differentiate into adipo-(a), osteo-(o) and chondrocytes(c). mncs at day expressed high levels of cd , cd , cd and cd , and very low levels of cd , cd and cd . expression of hematopoietic markers on cells at passage (p ) and thereafter progressively diminished while expression of cd , cd , cd and cd increased approaching %. cell doubling time ranged from to days at all passages. high clonogenicity was observed in all samples at all passages as shown by the presence of cfu-f colonies (> cells) with the exception of all samples at diagnosis which showed impaired proliferation and clonogenicity that returned to normal since remission was achieved at the following phases of treatment till the end of therapy. at p or p , mscs were differentiated towards the a, o, and c lineages by using specific induction media. differentiation was assessed by histochemistry and rt-pcr (lpl and ap for a, osteoprotegerin, osteocalcin and alp for o, aggrecan and col ii for c). p or p mscs from all groups exhibited bi-or trilineage differentiation. preliminary cloning experiments showed that msc population is composed of cells with differing proliferation potential and clonogenicity. these results indicate that blood diseases of childhood do not affect the characteristics of mscs which could have clinical applications particularly in hematopoietic reconstitution following transplantation. pegfilgrastim after autologous stem cell transplantation c.r. rinaldi, c. becchimanzi, a.m. risitano, n. marra, b. rotoli, g. de rosa university federico ii (naples, i) recombinant hemopoietic growth factors are not added routinely after autologous stem cell transplantation (asct) in our institution. in myeloma patients, a multicentric protocol to which we partecipate indicates granulocyte growth factor after re-infusion, at the dose of ug/kg, from day + to hematological recovery. because of equivalence between a single peg-filgrastim (peg) dose and daily filgrastim doses in decreasing the duration of neutropenia after standard dose chemotherapy, we used peg after asct in patients affected by myeloma or lymphoma. from february to november , we enrolled patients, suffering from myeloma ( undergoing single-asct and tandem-asct) and suffering from lymphoma ( non-hodgkin, hodgkin), conditioned by mel and beam, respectively. all patients received a single dose of mg pegfilgrastim subcutaneously h after autologous stem cells infusion. all patients engrafted neutrophils and platelets with a median time of and days, respectively, (regardless the underlying disease and type of conditioning). the incidence of febrile neutropenia was % ( / ) with a median duration of hours. we observed no adverse events secondary to peg injection. no patient had clinically significant mucositis. we compared this cohort of myeloma patients with an historical group of autotransplanted myeloma patients treated by standard daily doses of filgrastim and of lymphoma patients transplanted without g-csf administration. time to neutrophil and platelet recovery was identical in both groups of myeloma patients, and appeared sensibly reduced in lymphoma patients treated with peg as compared to no-g-csf patients (days and in nhls, and in hls) . we conclude that a single dose of pegfilgrastim after asct is safe, well tolerated and accelerates neutrophil recovery, thus decreasing time of hospitalization it seems equivalent to daily dose of filgrastim. we also documented no differences between our cohort of patients and historical groups in order of febrile neutropenia and proved infections. since peg disappearance from the circulation is not due to renal or hepatic clearance, but only to uptake on granulocytic cells and their precursors, this drug can be given even early after stem cells infusion, and will be utilized as soon as the engraftment occurs. mesenchymal stem cells are able to stimulate alloreactive immune cells l. fang, c. lange, m. engel, a.r. zander, b. fehse university medical center eppendorf (hamburg, d) bone marrow-derived mesenchymal stem cells (msc) have been suggested to be "immune-privileged" while exerting a strong immune-modulatory function as "third-party" cells in an hla-independent manner. therefore msc are interesting candidates for cell and gene therapeutic applications. however, a better understanding of the mechanisms underlying their immune-modulatory potential would be very important for msc application in clinical settings. we investigated the interaction of msc with allogeneic immune cells by co-culturing them with un-matched pbmcs and using them as third party in mixed lymphocyte cultures. we present data demonstrating that the immune-privileged state of msc results from an interplay of stimulating and suppressing factors. the directly stimulating activity leads to both active lymphocyte proliferation and secretion of pro-inflammatory cytokines by allo-reactive lymphocytes. stimulation is, however, dominant only at low msc:effector cell (< . under our experimental conditions), but outweighed with higher msc numbers by the suppressing activity. allogeneic mixed lymphocyte cultures as well as msc-mediated stimulatory effects are efficiently suppressed by the addition of mscconditioned medium underlining the important role soluble factors play in msc-mediated immune modulation. in conclusion, based on our data we suggest that the "immuneprivileged status" of msc reflects a sensitive balance of mhcmediated immune activation and the suppression of immunological reactivity largely conferred by soluble factors. circulating endothelial progenitor cells in children with non-malignant diseases following allogeneic bone marrow transplantation m. massa, v. rosti, r. campanelli, e. bonetti, v. meli, d. poddighe, t. mina, d. pagliara, d. lisini, f. locatelli irccs policlinico san matteo (pavia, i) bone marrow-derived endothelial progenitor cells (epcs) circulate in the peripheral blood (pb) of healthy subjects (hs). epcs seem to play an important role in maintaining vessel wall homeostasis, in the neo-angiogenetic processes and in the re-endothelization of the wall of injured vessels. the aim of the study is to assess the number and origin of circulating epcs in children with non-malignant diseases who received allogeneic bmt from an hla-identical sibling or a matched unrelated donor. we studied patients with thalassemia major (n= ), fanconi anemia (n= ), sickle cell disease (n= ), mucopolysaccharidosis (n= ), diskeratosis congenita (n= ), acquired aplastic anemia (n= ), or chediak higashi syndrome (n= ). we evaluated pb samples at , , , , and days after transplant. the number of epcs was evaluated as cd +vegfr- + or cd +cd +vegfr- + cells by cytofluorimetric analysis, and by in vitro culture. the analysis of pb samples from age matched donors (hs) was included in the study. donor or recipient origin of epcs was assessed on at least individually picked endothelial colonies by micro-satellite analysis. in patients tested days after transplant the percentage of circulating cd +vegfr- + cells (median . %, - . ) and the percentage of cd + co-expressing the cd and vegfr- antigens, representing a restricted subset of immature epcs (median . %, - . ), were comparable to those found in hs (median . %, . - . ; median . %, . - . , respectively). the number of epc derived colonies was also comparable in patients tested at days after transplantation (median / mononuclear cells, - ) and in hs (median . / mononuclear cells, - ). neither the percentage of circulating cell subsets, nor the number of epc derived colonies, showed significant modifications during day follow up (by anova test). microsatellite analysis was performed on the epc derived colonies of patients, tested at time points ranging from to months. in patients, all the analysed colonies were of donor origin; in the third patient all the analysed colonies were of patient origin (hematopoietic engraftment donor/recipient %/ %). circulating epcs are detectable in patients given allogeneic bmt from days up to months after transplantation. further studies are needed to definitively conclude their origin and to assess whether their recovery can be correlated to the clinical outcome of the transplanted patients. a. spiropoulos, e. goussetis, m. theodosaki, k. stefanaki, i. peristeri, v. kitra, e. petrakou, s. graphakos "aghia sophia" children's hospital (athens, gr) bone marrow and peripheral blood of adults contain a subset of progenitor cells, which are able to differentiate into mature endothelial cells, thus contributing to re-endothelialization and neo-vascularization. the number of these cells in healthy subjects is rather low; almost . % of total mononuclear cells and a variety of factors may further influence their number. to investigate how allogeneic stem cell transplantation (sct) influence circulating endothelial progenitor cells (cepcs), we obtained peripheral blood samples from transplant recipients at different time points (ranging from months to years after transplantation). peripheral blood mononuclear cells were separated by ficoll density-gradient centrifiguration and were seeded to fibronectin-coated well dishes containing endocult medium (stem cell technologies). in order to remove monocytes and mature endothelial cells, non-adherent cells, at day two of culture, were harvested and further cultured for an additional three days to allow formation of endothelial colonies. the phenotype of the cells that emerged in culture was characterized by immunohistochemistry, and their origin was determined using a polymerase chain reaction (pcr)-based assay for polymorphic short tandem repeats (strs). all samples gave rise to epcs colonies in days. the mean number of epcs colonies/ cells was ± . (range; - ) and it didn't seem to correlate with the post-transplant time. the cultured cells expressed typical endothelial markers such as cd and vwf. for each patient and at all time points, str-pcr analysis showed that cultured cells came exclusively from the donor. these results demonstrate that cepcs are detectable after sct and that their number is independent of post-transplant time. early cd + cells recirculate after autologous peripheral blood stem cell transplant and peg-filgrastim administration for haematological malignancies s. de matteis, n. piccirillo, s. de vita, f. sorà, m. tarnani, l. laurenti, p. chiusolo, g. reddiconto, s. sica, g. leone università cattolica del sacro cuore (rome, i) cd protein is widely accepted as reliable marker for identifying hemopoietic stem or progenitor cells in bone marrow and in peripheral blood. cd + cells represent a heterogeneous cell population consisting of primitive uncommitted and pluripotent progenitors as well as committed stem cell. previous studies showed that these progenitors, mainly myeloid-committed subsets, are detectable in the early phase following infusion of autologous/allogeneic stem cell at day + (albo et al, haematologica ) . based on these findings, we investigated the kinetics of appearance of cd + cells after autologous peripheral blood stem cell transplant (apbsct) and administration of pegfilgrastim mg at day + , and its correlation with haematological engraftment. we studied in consecutive patients (pts), affected by haematological malignancy and treated with apbsct (table ) , the percentage of cd + cell in peripheral blood every other day from day + until patient discharge. these cells were detectable starting from day + after transplantation. the peak of the cd + cells was at day + (range - ), at this day wbc were /mmc (range - /mmc) and the number of cd +/mmc were . (range . - ). there was no correlation between total number of cd +/kg infused and day, absolute number and percentage of cd + peak. statistical analysis demonstrated a significant negative correlation (r- . , p= . ) between age of pts and peak of cd +, while a significant positive correlation (r . , p= . ) between age of pts and day of cd + peak. when haematological reconstitution after apbsct we observed a significant positive correlation between day of cd + peak and time to absolute lymphocyte> . x /l(alc) (r . , p= . ), pmn> . x /l (r . , p= . ), plt> x ³/mmc (r . , p= . ), length of hospitalization (r . , p= . ). this data seems to link up the appearance of cd + cells with the bone marrow reserve: younger pts release higher number of cd + in peripheral blood after apbsct and these cells are detectable sooner than in the older pts. furthermore, time to alc> . x /l and pmn> . x /l recovery, plt> x ³/mmc and length of hospitalization are longer in pts that release later cd + cells, i.e. the older pts. early appearance of cd + cells after apbsct and peg-filgrastim might be considered as surrogate marker of bone marrow reserve. further analysis of cd + subset are ongoing to confirm these data and to clarify their significance. imbalance of osteoprotegerin/receptor activator of nuclear factor-kb ligand in bone marrow plasma and microenvironment after allogeneic stem cell transplantation c. selleri, l. tauchmanovà, p. ricci, a.m. risitano, m.c. martorelli, g. cerciello, i. imperatore, a. casale, t. musella, g. lombardi, a. colao, b. rotoli federico ii university (naples, i) persistent decrease in bone mineral density (bmd) is a known complication after allogeneic stem cell transplantation (allo-sct) due to the transplant procedure, gonadal failure, immunosuppression and deficit of osteoblast precursors in the marrow microenvironment. in addition, transplanted patients may develop chronic endocrine and immunological disorders, including chronic graft versus host disease, that can further affect bone turnover. osteoprotegerin (opg) plays a pivotal role in bone remodelling, by neutralizing the effect of rankl on differentiation and activation of osteoclasts. we investigated the relationships between densitometric values, circulating opg and interferon-gamma (ifn-gamma) levels; moreover, opg and rankl levels were measured in marrow plasma and in conditioned medium of long-term cultures (ltc) of marrow mesenchymal-derived osteogenic cells. thirty six allo-sct patients ( females; age: ± yrs) were enrolled and compared to controls matched for age, gender and body mass index. bmd was measured at lumbar spine and femoral neck by the dexa technique. lumbar and femoral bmd were lower in patients than in controls ( . ± . vs . ± . , p< . ). serum opg and ifn-gamma were significantly (p< . ) higher in patients than controls. patients' serum ifn-gamma correlated with opg levels (r= . ; p= . ). by contrast, opg levels were lower in patients than in controls in marrow plasma (p< . ) and in conditioned media after one (p= . ) and months (p= . ) of ltc of marrow mesenchymal-derived osteogenic cells. rankl values were similar between patients and controls, being lower in conditioned medium than in marrow plasma (p< . , both groups). the rankl/opg ratio in ltc was significantly higher in patients than in controls, although this ratio progressively increased with the time of ltc in both groups (p< . ). there was no correlation between serum, in situ opg levels and densitometric values. in conclusion, our findings suggest that after allo-sct: ) the paradoxically high serum opg levels, which correlate with ifn-gamma, are likely due to deranged immune system; ) low marrow and microenvironment opg levels confirm persistent quantitative and qualitative deficit of osteoblastic precursors; ) the increased microenvironment rankl/opg ratio may negatively affect bone remodelling. mesenchymal stem cells (mscs) are endowed with multilineage potential and immunomodulatory ability, these properties rendering them attractive for tissue engineering and immunotherapy. however, it is still a matter of debate whether donor mscs have a sustained engraftment in the host bone marrow (bm) after allogeneic hematopoietic stem cell transplantation (hsct). in particular, studies on the fate of mscs transplanted with cord blood (cb) are lacking. the aim of this study was to analyse the donor/recipient origin of mscs in pediatric patients receiving an allogeneic hsct. thirty-six patients undergoing allogeneic hsct for either malignant ( cases) or non-malignant disorders ( cases) were enrolled in the study; patients received cb transplantation (cbt, from a related and from an unrelated donor) and patients bm transplantation (bmt, from a related and from unrelated donor). results were also compared with those obtained in adults given hsct for either malignant ( cases) or non-malignant ( cases) disorders. mscs were grown from bm aspirates taken - months after hsct. msc samples at the third-fourth passage were phenotipically characterized and resulted to be positive for cd , cd , cd , cd , cd (> %) and negative for cd , cd , cd (< %). donor/recipient origin of mscs was assessed by amelogenin assay (in case of male recipient/female donor) and microsatellite analysis. mscs were grown from pediatric patients; in samples ( after bmt and after cbt) a confluent layer of cells did not grow, leading to an insufficient quantity of mscs for chimerism analysis. molecular analysis on mscs demonstrated a full recipient chimerism in / and in / of the assessable pediatric patients given bmt and cbt, respectively. a mixed msc chimerism with donor cells was observed in patients transplanted with bm cells and in children given cbt. chimerism analysis performed on peripheral blood mononuclear cells (pbmcs) of the same patients, showed a full donor chimerism in all children given bmt but one, while a mixed chimerism was detected in out of children given cbt. a full recipient msc chimerism was observed in all adult patients, who also displayed a full donor pbmc chimerism. these data suggest that bm soil of pediatric patients might be more favourable than that of adults for the engraftment of transplanted mscs and that mscs able to engraft in the host can also be transferred with cb. graft engineering r cryopreservation of peripheral blood progenitors for autologous transplantation in haematological malignancies with different concentration of cryoprotectant -five-year single-centre experience a. pivkova, l. cevreska, n. siljanovski, z. stojanoski, o. karanfilski, s. genadieva stavrik, i. panovska, s. krstevska balkanov, s. trajkova, b. georgievski clinical center (skopje,mk) in this study we present our five year center experience with cryopreservation of pbsc and autologous transplantation in patients with hematological malignancies treated in a period - at department of hematology, skopje. material and methods: diagnosis of patients were ( aml, nhl, mm, hd) and median age at transplant was years ( - ). mobilization of pbsc was provided with etoposide (vp- ) + g-csf mcg/kg in aml patients, and high dose cyclophosphamide - gr/msq+g-csf mcg/kg or alone g-csf mcg/kg in patients with limphoproliferative diseases. collected pbsc were cryopreserved in solutions with % dmso in patients and % dmso in patients, computer programmed until - c, and stored different period in liquide nitrogen on - c. autologous transplant was preformed wid conditioning consisted of myeloablative highdose chemotherapy, bucy in aml patients, high dose mel in mm patients, beam or hd ice in nhl patients and beam in hd patients. cell viability was assessed by fluorescence microscopy using acridine orange dye exclusion. results: a total of pbsc cryopreservation procedures were preformed in our group of patients with median ( - ) apheresis procedures. median period from storage of cryopreserved pbsc grafts until thawing was days ( - ). total number of infused cd +cells was between , - x /kg and median number of mononuclear cells was , x /kg ( , ) . the amount of infused dmso solution ranged between - ml (median ml) with dmso concentration ranging ml- ml (median ml) in a group preserved with %dmso and - ml (median ml) in % dmso cryopreserved grafts. the viability of the fresh harvests before storage vas median % (range , - , %). the poorer viability was associated with harvest cell count. bellow x /l the median viability was % and only / cases had < % viable cells. harvests count above x /l the median viability was % ( , %- %). in a group of patients that received pbsc grafts preserved with % dmso, also revealed signs of mild dmso infusion related toxicity ( %vs %). hematopoietic recovery was similar in both groups, sor ne> , x /l on day + ( - ), plt > x /l on day + ( - ). our results confirm that the infusion of cryopreserved autologous pbsc in hematological malignancies revealed successful engraftment in all patients and good cell viability. we did not registered "hard to mobilize" patients and graft failure. thermogenesis axp™ and bioarchive™ systems for automated cord blood banking l. dobrila ( ), s. jiang ( ), j. chapman ( ), d. marr ( ), k. kryston ( ), p. rubinstein ( ) ( )national cord blood program at new york blood center (new york, usa); ( )thermogenesis corp (rancho cordova, usa) background: good tissue practices (cgtp) in cord blood banking require product uniformity and reproducible mononuclear cell recovery and viability, suggesting that automation could be critical to facilitating cgtp-compliance for cord blood banks. processes that lend themselves to automation are cord blood volume reduction, controlled rate freezing, storage and retrieval that avoids unnecessary transient warming events. we have evaluated the autoxpresssystem (axp),that allows for automated volume reduction in a closed system. the autoxpress consists of a microprocessor-controlled device and a disposable closed blood bag set that provides for the separation of cb into a freezing bag, an erythrocyte bag and an excess plasma bag, the mononuclear cell product is concentrated into a uniform volume in the freezing bag, ready to be cryoprotected and fully compatible with the bioarchive system,a system that allows the controlled-rate freezing, liquid nitrogen storage and retrieval of , cbus. study design: the efficiency with which cord blood hematopoietic progenitor cells can be concentrated into the freezing bag of the autoxpress bag-set was determined using the cd cell marker and colony-forming unit (cfu) counts as principal indices. the product was cryoprotected with % dmso, frozen in the bioarchive system, stored for - weeks under the liquid n level and then retrieved and thawed using the standard clinical protocol. twenty-three consecutive cord blood units were evaluated for cell recovery by measuring the collection and product volumes, the hematocrit and the counts of total nucleated cells (tnc), mononuclear cells (mnc), cd + cells and colony-forming units (cfu),before and after axp processing. we also determined these indices after freezing, storage in the bioarchive system and thawing. results: results are presented as the mean s.d. (n= ) for all values. the axpprocess achieved mnc fraction volumes of . . ml with a final average hematocrit of . . %. the post-processing recovery of cd + cells was . . % and those of cfu . . %, of mnc . . % and of tnc . . %. less than % of tnc were lost into the excess plasma bags. granulocytes, accounting for % of tnc and less than . % of the cd + cells were lost into the red cell bags. post-thaw the recoveries of cfu and viable cd + cells were . % and . %, respectively. conclusions: the axpefficiently and reproducibly separates cord blood mononuclear and cd + cells into a consistent, uniform volume. these cells retained their viability post bioarchive freezing, storage and retrieval (> %). thus, axpcoupled with the bioarchive system supports a very high quality standard for automated cord blood processing. the influence of cryopreservation and the duration of frozen storage k. nowak, s. giebel, m. krawczyk-kulis, j. wojnar, j. holowiecki silesian medical university (katowice, pl) among the factors which enable successful transplantation, the ability to store and subsequently recover sufficient viable hematopoietic stem cells to reestablish hematopoiesis is critical. the goal of this study was to evaluate the impact of freezing procedures and cryopreservation in liquid nitrogen into - °c on cell viability, wbc, cd + cells recovery and clonogenic capacity. in the retrospective study samples derived from patients with hematological malignancies (n= ) and healthy donors (n= ) were analysed. median duration of the product storage was . years ( day - . years). the viability (%), wbc (g/l), and cd + (%) all decreased significantly after cryopreservation (p< . ) with median relative changes of - . %, - . %, and - . %, respectively. after thawing the viability equaled . % ( - %), wbc . g/l ( . - . g/l), and cd + cells . % ( . - . %). in multivariate analysis the following factors were associated with poor recovery: ) viability: presence of malignant disease (p= . ), use of cyclophosphamide (ctx) for mobilisation (p= . ), ) wbc: presence of malignant disease (p= . ), older age (p= . ), ) cd + cells: presence of malignant disease (p= . ), storage duration (p= . ). after thawing the median number of clones/ cells was as follows: ) cfu-gm on day : ( - . ), ) cfu-gm on day : . ( - . ), ) bfu-e: . ( . - ), ) cfu-gemm: . ( - . ). the clonogenity was negatively influenced by: ) cfu-gm on day : presence of malignant disease (p= . ), chemotherapy-containing mobilisation regimen (p= . ), ctx for mobilisation (p= . ), ) cfu-gm on day : older age (p= . ), presence of malignant disease (p= . ), chemotherapy-containing mobilisation regimen (p< . ), ) bfu-e on day : presence of malignant disease (p= . ), chemotherapy-containing mobilisation regimen (p< . ), cfu-gemm on day : diagnosis other than cml (p= . ). we conclude that both diagnosis and mobilisation regimen have impact on recovery and clonogenity of cryopreserved hematopoietic stem cells. on the other hand, even long-term storage enables preservation of vial cells with high clonogenic potential, which may be used for transplantation. immune reconstitution after hla-haploidentical transplantation using unmodified marrow and cd depleted blood stem cell: not different from hla-identical transplantation p. rujirojindakul ( ) immune reconstitution (ir) is a key role of allogeneic hematopoietic stem cell transplantation (hsct) not only because persistent immune defects is related to posttransplant infectious morbidity, but also because it may influence the risk of relapse and the development of secondary malignancies after hsct. many factors have an impact on ir, especially the degree of genetic differences between donor and recipient. several studies have shown extreme slow ir in patients receiving t-cell depleted graft from human leukocyte antigen (hla)-haploidentical (hap) donor. differently, we have used unmodified marrow on day and cd -depleted mobilized blood cells (mbc) on day for haploidentical transplantation. cd -depleted mbc are devoid of cd -positive cells, they contain cd -positive cells, nkcells and a minority of cd -positive cells. to compare the ir during the first year post transplant between hap and hlaidentical (id) recipients, we have carried out a prospective, longitudinal analysis of ir in hsct patients in each group. we analysed reconstitution of naïve, memory t cells, b cells, natural killer cells ,dendritic cells and monitored thymic output by using tcr rearrangement excision circles (trecs). surprisingly, reconstitution did not differ between the groups. this study reveals that ir in hap transplantation using unmodified marrow and cd -depleted mbc is not worse than id group. background: autologous graft versus host disease (gvhd) is a novel self-limited autoaggression syndrome, encountered in the autologous pbsct (peripheral blood stem cell transplantation) setting with spontaneous occurrence in patients receiving cd + enriched autografts. aim: to present the case report, concerning the occurrence of a rather rare form of gvhd in a patient with cd + enriched auto-pbsct for ewing sarcoma. case report: we present the case of a years old female patient with ewing sarcoma of the scull and right hip who underwent autologous pbsct with a cd + enriched graft. the preparative regimen consisted of busulfan and melphalan. engraftment occurred on day + for neutrophils and on day + for platelets. at day + signs of acute gvhd involving only the skin occurred initially in the axillae, flexion sites of the elbows and popliteal region and shortly afterwards involving the face, neck, trunk and abdomen. beside maculopapular exanthema, bullae formation and large epidermal descuamation were observed, an image corresponding to stage iv of acute gvhd. the patient remained feverless, without pruritus or other signs or symptoms. at the time of onset the investigation of immune reconstitution presented marked lymphopenia (abslolute values per microliter):cd + = ,cd + = ,cd +/cd + = ).no other modified biological parameters could be found. skin biopsy revealed lymphoid, predominantly cd + infiltration. in days the lesions progressively disappeared, with the maintenance of residual hyperpigmentation and nail keratosis. conclusion: autologous gvhd, confined to the skin without any clinical-biological evidence of internal organ disease is a possible self-limited complication of hct cd +cell enrichment of the graft could be in our case a predisposing factor.the impact on the evolution will remain an issue to be assessed. why will an amotosalen-based protocol for extracorporeal photopheresis be valuable? j. e. hearst ( ) , f. heshmati ( ), s. talib ( ) ( )university of california (berkeley, usa); ( )hopital cochin (paris, f) bone marrow transplantation has the potential of providing a complete cure of the disease symptoms of hematologic malignancies and, ultimately, with an appropriate safety profile, the symptoms of hemoglobinopathies as well. even in the case of related fully matched bone marrow transplantation, there is morbidity and mortality associated with graft-versus-host-disease (gvhd). successful application of mismatched (related-haploidentical and unrelated) bone marrow transplantation (bmt) in patients with leukemia or lymphoma requires that improved overall outcomes of bmt be obtained, coupled with the avoidance or successful treatment for the gvhd now experienced in existing mismatched bmt protocols. these two goals may require use of reducedintensity conditioning (ric) in support of the transplantation. extracorporeal photopheresis (ecp) has proven to be effective as a treatment modality for gvhd following transplantation and is gaining popularity as a treatment protocol. yet, there remain significant technical challenges in the application of ecp. as a solution to many of these problems, we propose the development of a more simple and less expensive process for the psoralen photochemical treatment of the transplant patient's autologous leukocytes. this process will define the therapeutic dose of photochemically treated cells required for clinical responses equivalent to those which have been observed to be most effective using presently approved ecp instrumentation and protocols. however, this new process uses amotosalen, a more photochemically efficient psoralen, and it uses lymphocytes collected by the conventional methods of blood banks. the greater photoefficiency of the psoralen compound results in far shorter exposures of the target leukocytes to uva light, leading to less non-specific damage of the cells, leaving them metabolically active, although unable to divide. it is our premise that these are the ideal properties that photochemically treated lymphocytes must have in order to s maximize the positive immunological responses associated with a successful photopheresis treatment. uva phototherapy as a treatment for sclerodermic chronic graft-versus-host disease refractory to immunosuppressive therapy g. marotta, m. pellegrino, s. sammassimo, m. tozzi, c. miracco, m. fimiani, f. lauria azienda ospedaliera universitaria senese (siena, i) chronic graft versus host disease (cgvhd) is the most common late complication affecting long-term survivors of allogeneic hsct. several organs are targets of cgvhd but the skin is the tissue mainly affected. two subtypes of involvement, cutaneous lichenoid and sclerodermoid, have been described, based on clinical and histopathological examinations. chronic gvhd is usually treated with immunosoppressive drugs which, however, are not effective in about - % of patients. in this setting of refractoty/resistent patients, extracorporeal photochemotherapy and ultraviolet a (uva) radiation often show a positive clinical outcome, but the results are limited by the low number of treated cases. in our institute, uva phototherapy has been used for treating patients with advanced cutaneous cgvhd (generalized sclerodermoid skin involvement) resistant to conventional immunosuppressive therapy. patients enrolled gave fully informed consent and they underwent skin biopsies before and after uva phototherapy. moreover, the cutaneous elasticity has been evaluated by means of an elastometer. uva radiation ( - nm) was emitted by a gp- h irradiation unit with a dose of j/cm². irradiance was measured with a spectroradiometer and found to be mj/cm² at skin level. patients were treated times per week for a total of weeks (total j/cm²) and the cutaneous lesions were carefully inspected and palpated before starting every treatment. patients were conditioned with a reduced intensity regimen and received unmodified g-csf mobilized pbsc from matched related donors. gvhd prophylaxis consisted of csa and mtx. all patients had failed to respond to at least three lines of immunosuppressive therapy. at the end of sessions the clinical response was assessed subjectively and objectively, and it was graded as good (obvious softening), moderate (mild softening) and poor (no change). two patients had a good response and one a moderate response reporting remarkable softening of skin lesions after uva phototherapy without side effects. clinical responses were associated to an improvement of histopathological findings. the evaluation of skin elasticity showed a significant increase in resilience, hysteresis, and distensibility. our results demonstrate the efficacy of uva phototherapy. it appear, in the setting of cutaneous cgvhd, as a procedure well tolerated and effective particularly for patients who do not respond to standard immunosuppressive treatments. rituximab in chronic graft-versus-host disease of the lung i. von olshausen, a. spyridonidis, h.c. bremer, w. windisch, h. bertz, j. finke university hospital freiburg (freiburg, d) we report about a year old male patient in whom severe obstructive chronic graft-versus-host-disease (cgvhd) of the lung improved with rituximab, an anti-cd antibody. the patient had undergone hla-matched unrelated allogeneic peripheral stem cell transplant for his acute myeloid leukaemia in august . with discontinuation of cyclosporine after day + mild cgvhd of the skin and liver occurred, but improved spontaneously within a few weeks. in january the patient presented with severe dyspnoea associated with an afebrile upper respiratory tract infection. the work-up revealed severe obstruction on lung function tests, nodular infiltrates in computed tomografy scan (ct), acute bronchitis in bronchoscopy, but no infectious agent in bronchial lavage. initial and long term treatment with various antibiotics, antimycotics and several immunosuppressants (prednisone, rapamycin, mycophenolatmofetil, cyclosporine a, cyclophosphamide) was unsuccessful. after a single dose of rituximab ( mg/m²) dyspnoea improved for a few days, but symptoms reappeared. we started a regular weekly rituximab application in july resulting in marked clinical improvement, slight improvement on lung function (peak exspiratory flow, pef) and regression of the nodular infiltrates in ct scan despite tapering off other immunosuppression and intermittent respiratory infections. the patient was able to participate in household thresholds again. rituximab therapy was discontinued in november because of patient's admission with spontaneous pneumothorax. the patient is now being evaluated for lung transplantation. like in our case, monoclonal anti-cd antibody has recently shown efficacy in cgvhd and several autoimmune diseases, probably due to elimination of b cells which may act as antigen presenting cells for t cells and as a source of autoantibody production. conclusion: we propose regular weekly rituximab application as a treatment option in cgvhd of the lung. are peak level measurements of cyclosporin a useful in allogeneic stem cell transplantation? c. scheid, c. heinz, u. holtick, k. hübel, v. göde, t. zander, a. engert, m. weihrauch, m. hallek university of cologne (cologne, d) peak level plasma levels of cyclosporin a (csa) or hours after drug administration (c or c ) correlate with the degree of calcineurin inhibition and with the auc much better than trough levels (c ). in solid organ transplantation dosing according to c levels rather than c led to reduced rejection rates. in allogeneic stem cell transplantation csa doses are still adjusted according to c levels. to investigate whether c and c levels would be useful in csa dosing after stem cell transplantation these measurements were performed in consecutive patients in addition to c levels. if intravenous csa was given, peak levels were assessed after the end of a hour infusion. so far serial measurements were undertaken in transplant patients ( aml, all, saa, myeloma, cll). there was only a weak correlation between trough and peak levels of csa and between dose and plasma levels irrespective whether the drug was administered orally or intravenously. although all trough levels were within the target range of - ng/ml peak levels were all below ng/ml. there was no evidence of so-called "late absorbers" as all c levels were lower than the c levels. two patients showed evidence of microangiopathic hemolytic anaemia, however peak and trough levels of csa were not different from other patients. all but two patients showed signs of acute gvhd. in view of the csa doses given and the trough levels, the peak levels were considerably lower than expected from results in liver or renal transplantation. this was even more surprising as most patients received itraconazole, a drug known to increase csa levels by interfering with csa metabolism in the liver. in summary c monitoring is feasible and demonstrates lower peak levels than expected from solid organ transplantation. these levels should lead to a lesser degree of calcineurin inhibition. whether this translates in increased rates of gvhd remains to be seen with a larger patient number. if this holds true csa dosing with higher c target levels might decrease acute gvhd after stem cell transplantation as previously shown for rejection rates after liver transplantation. clinical and histopathological features of oral chronic graft-versus-host disease following allogeneic reducedintensity conditioning haematopoietic stem cell transplantation: a single-centre blind study f. demarosi ( ) introduction: oral involvement of chronic graft-versus-host disease (cgvhd) occurs in % of patients suffering from cgvhd and the oral cavity may be the primary or even the only site of cgvhd involvement. lichen planus-like lesions are the most distinctive oral changes of cgvhd. the histopathologic changes of oral cghvd include epithelial atrophy, apoptotic bodies, hydropic degeneration of the basal cells, and a mononuclear subepithelial cell infiltrate with lymphocyte invasion. aim: the aim of this blind study was to compare clinical and histological features of oral mucosa in patients who underwent allogeneic reduced intensity conditioning hematopoietic stem cell transplantation (ric hsct). patients and methods: this study enrolled adult patients who consecutively underwent allogeneic ric hsct for hematological malignancies between october and october . all patients were assessed for the presence of oral cgvhd by oral examination performed on the day + at the unit of oral pathology and medicine, university of milan. clinical lichenoid changes of the oral mucosa were regarded as positive for cgvhd. following informed consent, an incisional biopsy was taken from oral mucosa of all patients, either with or without oral cgvhd lesions. biopsies were examined by a pathologist who was unaware of clinical aspect of the mucosa (blind). results: biopsies were taken from patients with clinical evidence of oral cgvhd and patients with apparently normal oral mucosa. biopsies were performed at a time point ranging from to days (median . days) after transplantation. sites of biopsies were buccal mucosa and gingiva. histological cgvhd changes were detected in all the patients having also clinical evidence of oral cgvhd, and in out of patients with apparently healthy mucosa. conclusions: while histological changes of oral mucosa without corresponding clinical changes are not sufficient to make a definite diagnosis of oral cgvhd, their detection might be of considerable help to predict the onset of the disease following ric hsct. a longer follow up of patients showing histological changes with no clinical counterpart, will possibly elucidate whether such changes are indeed predictive of the occurrence of clinically evident lesions. evaluation of safety and tolerability of extracorporeal photochemotherapy in paediatric patients affected with graft-versus-host disease c. del fante, c. perotti, gl viarengo, p. bergamaschi, d. romano, l. salvaneschi policlinico s. matteo (pavia, i) objectives: extracorporeal photochemoterapy (ecp) is a therapeutic option for treatment of acute and chronic graft versus host disease (agvhd and cgvhd) resistant to standard drug therapy. in the pediatric context, ecp procedure has some technical limitations when compared to adult subset. low body weight, venous accesses, hypersensitivity to hypocalcemia, fluid balance and transfusion demand may represent a limitation for ecp treatment in children. we report our experience in very low body weight children (≤ kg) affected with agvhd and cgvhd in terms of safety and tolerability of ecp. methods: ecp consists of distinct steps: ) collection of mononuclear cells by spectra cobe (version . ) cell separator device processing blood volumes; ) ex-vivo dilution with saline and addition of -mop to the bag, transfer of the buffy coat in a uv-a permeable bag and irradiation at °c; ) reinfusion of the cells to the patient after hours to avoid hypotermia. patients, median age . years (range: - ) median weight . kg (range: - ) were treated. a central venous catheter was positioned in all patients. our treatment protocol consisted in procedures per week in agvhd and procedures per week in cgvhd, both followed by procedures every weeks for - weeks then by procedures per month. patients were affected with agvhd (grade ii-iv with skin, liver and gut involvement); patients had extensive cgvhd (skin, mucosal, liver, and lung).the cell separator device was primed every time with filtered and irradiated rbc. calcium gluconate was continously administered by pump during the procedure. the acd/whole blood ratio was always setted at : . all patients were monitored for blood pressure and heart rate during the entire procedure. results: the total number of ecp procedures performed was , with a median of (range: - ) procedures/patient. / ( . %) patients experienced mild hypotension, / ( %) moderate hypotermia and / ( . %) hypocalcemia (nausea and vomiting); no procedure was discontinued. the transfusion demand did not augment during all the course of treatment; no life-treathening infections were recorded. conclusion: we demonstrated that ecp is feasible and safe even in very low body weight patients on condition that ecp is performed adopting some simple precautions. our experience broadens the cohort of patients who can benefit of this therapeutic option. silesian centre for cellular transplantation, poland k. suchnicki, a. lange lower silesian center for cellular transplantation (wroclaw, pl) haematological patients -malignancies (cml: n= ; aml:n= ; all:n= ) and saa (n= ); children (n= ) and adults (n= ) were transplanted with bm (n= ) and pbpc (n= ) from hla-identical sibling donors in years to . all leukemia patients received myeloablative conditioning (bucy or bucyvp);saa patients were conditioned with cy or cyatg. chimerism was tested and proved to be complete in all but cases (death before day + ). during years of follow up: patients died of transplant related causes (regimen related toxicity:n= , rrt/agvhd:n= , infection: n= , agvhd/multi organ failure/inf:n= , cgvhd/mof/inf:n= ) or relapse (n= ). agvhd was seen in , % cases (grade = , % of patients with hematological recovery, grade = %, grade> = , %). severe agvhd (grade> ) was seen only in leukemia patients, who received toxic conditioning, but not in saa conditioned only with cy+-atg. agvhd was more frequent and severe in pts with a high grade of toxicity (among patients with low rrt there were % cases of agvhd= and no cases of agvhd> ; among patients with high rrt there were % agvhd= and % agvhd> ). in addition to toxicity infections and inflammation seemed to aggravate agvhd as shown by the presence of an elevation of serum crp at the advent of severe agvhd. apparently the presence of toxicity and agvhd had an additive negative effect on survival. the incidence of cgvhd among all patients living>day+ was % and increased with :i) diagnosis: saa( , % of saa cases),aml ( , %),all( , %),cml( , %),ii)previous agvhd: all surviving patients with agvhd> developed cgvhd. cumulative survival is better in following :i)saa vs aml/all/cml (p= , ),ii)all, cml early stage vs advanced (p= , ),iii) conditioning bucy vs bucyvp (p= , ), iii) age: children vs adults (p= , ),iv) gender: female recipient from male donor better than all other pairs (p= , ),v) rrt: toxicity grade - (sum of who degrees for toxicity of skin, mucous, liver, gastro-intestinal tract)vs grade - vs grade - (p= , ), vi) crp: serum level of - vs level> (p= , ),vii) agvhd grade - - vs grade - (p= , ), viii) cgvhd grade lim better than no cgvhd better than extcgvhd (p= , ) which reflects gvl surveillance of gvh cells (less relapses) without overt, life threatening reaction. this observation documented that the outcome of transplantation in sibling setting depends largely on toxicity, infections and agvhd which in turns aggravate themselves. the role of atg in reducing acute gvhd in related and unrelated donor transplants y. zalyalov, a. ganapiev, s. alexeev, a. smirnova, n. stancheva, n. ivanova, a. alyanskiy, b. afanasyev pavlov's state medical university (st. petersburg, rus) background: acute graft versus host disease (gvhd) remains the major complication of allogeneic haemopoietic stem cell transplantation (allohsct) with an incidence of - % and mortality of up to - %. atg is commonly used in the conditioning regimens for donor transplants to reduce the risk of rejection and gvhd. patients and methods: since up to date we have performed allohsct ( ( %) with atg and ( %)without atg) for the following patients -aml (n= ), all (n= ), cml (n= ), non-hodgkin's lymphoma (n= ), aa (n= ), hodgkin's lymphoma (n= ), mds (n= ), kostmann's syndrome (n= ), hypereosinophilic syndrome (n= ). the average age was years . dose of atg was mg/kg.b.w. ( - ). myeloablative conditioning regimen consisted mainly of bu-cy ( pts ( %), nonmyeloablative conditioning treatment -flu-cy ( pts ( %). the gvhd prophylaxis was short term mtx and csa. results: rate of gvhd grade -ii in pts with atg was % ( pts), gvhd grade iii-iv - % ( pts). rate of gvhd -ii in group pts without atg was %( pts) and gvhd iii-iv - % ( pts). tmr in group with atg was % ( pts), without atg - %( pts). os in pts with atg was %, without atg - %. conclusions: the use of atg in conditioning regimen reduces the incidence of acute gvhd grade iii-iv and increases overall survival in patients after allohsct (p= . ). intracellular markers of eosinophils and mast cells in patients with lymphoproliferative and myeloproliferative disorders associated with high-grade eosinophilia l. komarova, y. zueva, n. mikhaylova, b. afanasyev, a. totolian pavlov state medical university (st. petersburg, rus) objectives: distinct eosinophilia rarely occurs in chronic graftversus-host disease (gvhd) after allogeneic hematopoetic stem cell transplantation (hsct), but is often seen in hematological malignancies, including lymphoproliferative diseases. a hypothesis exists that marked eosinophilia in chronic gvhd is mainly of non-clonal origin and, in general, is associated with favorable prognosis. the aim of this study was to investigate the serum levels of secretable eosinophil and mast cells proteins in the patients with chronic gvhd and other hematological malignancies. patients and methods: six post-transplant patients with chronic gvhd with eosinophilia were followed up after allo-hsct. thirty hematological malignancies in patients with marked eosinophilia (> . x /l) were also observed, including cases of lymphoproliferative (n= ) and myeloproliferative (n= ) diseases. eighteen patients with bronchial asthma (ba) comprised a reference group for polyclonal eosinophilia. high content of peripheral blood eosinophils served as the major criterion of patients' selection and was confirmed by flow cytometry. eosinophil cationic protein (ecp) and tryptase were measured in serum by fluoroimmunoenzyme assay (pharmacia, sweden). results: the percentages of circulating eosinophils in patients with lymphoproliferative (median: . %; range, - %) and myeloproliferative diseases (median . %; range, - %), and in cases of chronic gvhd after allo-hsct (median . %; range, - %) were significantly higher than in ba patients (median . %; range, - %); Ð= . . the levels of total ecp were markedly increased in the patients with lymphoproliferative diseases (median: . ng/ml; range, - ng/ml) compared to ba (median: . ng/ml; range, . - ng/ml); p= . . however, the serum levels of tryptase and ecp in total group of hematological patients (median: . ng/ml; range, to . ng/ml) were not increased, as compared with asthma cases (median: . ng/ml; range, . to . ng/ml); p= . . likewise, in chronic gvhd with eosinophilia, the serum contents of both ecp and tryptase did not differ from those in ba. conclusion: the significant differences in ecp levels in blood serum in the patients with lymphoproliferative diseases and absence of differences in ecp and tryptase levels in blood serum of chronic gvhd and ba patients suggests intact functions of mature eosinophils in gvhd, thus reflecting nonclonal genesis of this feature in both states. natural killer cell activity in the early phase after allogeneic stem cell transplantation: impact of conditioning strategies l. fischer, o. penack, c. gentilini, e. thiel, l. uharek charité campus benjamin franklin (berlin, d) background: research into the role of natural killer (nk) cells in allogeneic stem cell transplantation (sct) has been greatly expanded recently. nk cells may contribute to the gvl reaction possibly without exerting a gvhd effect. using a novel flow cytometric assay, which detects the lytic granule membrane protein cd a as a marker for nk cell degranulation, we investigated the effect of in vivo t cell depletion and the type of conditioning on nk cell function in the early phase after allogeneic sct. methods: peripheral blood mononuclear cells were collected at day + and + after allogeneic sct and incubated with the nk sensitive cell line hl ( hours at °c, e:t ratio : ). pe-cy conjugated anti-cd a antibody was added prior to incubation, monensin ( . mm) was added after hour of incubation. finally cells were further stained against cd , cd and cd and the proportion of cd a positive nk cells was measured by flow cytometry and the absolute number of degranulating nk cells was calculated. results were compared to values from healthy controls. results: twenty two patients were investigated of whom had been treated with a conventional dose conditioning regimen and had received a reduced dose regimen. at day + , the proportion of nk cells with cytotoxic activity (cd a+/cd +) was significantly reduced as compared to normal donors ( . % vs. . %, p< . ). at day + the percentage of degranulating nk cells in five evaluated patients was within normal range (mean . %). the predominant proportion of degranulating cells was in the cd dim/cd -subpopulation (mean . %). at day + the percentage of cd a+ cells averaged . % after conventional conditioning compared to . % after reduced intensity conditioning (p= . ). the absolute number of degranulating nk cells was significantly reduced after conventional conditioning ( . /µl vs. . /µl, p= . ). neither the percentage ( . % vs. . %, p= . ) nor the absolute number ( . /µl vs. . /µl, p= . ) of cd a+ nk cells differed significantly in patients with and without atg induced t cell depletion at day + . conclusion: according to our data cytotoxic activity of nk cells is reduced after allogeneic sct. the absolute number of nk cells with cytotoxic activity is significantly higher after reduced intensity conditioning which may impact on the outcome of this strategy. we saw no significant influence of antibody mediated in vivo t cell depletion by atg on nk cell activity during the first two months post sct. it is known that nk alloreactivity is involved in the control of neoplastic cells in the setting of aploidentical bone marrow transplantation (bmt) in patients with acute myeloid leukemia (aml). the role of nk alloreactivity in the setting of marrow unrelated transplant (mud) in lymphoid neoplasia is still controversial. in a series of patients ( acute lymphoid leukemia, non-hodgkin's lymphoma l, severe aplastic anaemia), we investigated whether nk alloreactivity is involved in the control of lymphoid neoplastic cells. in addition, using monoclonal antibodies (moabs), we evaluated the expression of killer immunoglobulin-like receptors (kirs), killer lectine-like receptors (klrs), and natural cytotoxicity receptors (ncrs) in patients under study. according to hla-cw alloreactivity, patients were separated into two groups: a group of patients with potential nk alloreactivity (group a: all, nhl, saa), a second group of patients lacking nk alloreactivity (group b: all, nhl). the mean follow up was months ± months. % of group a patients were alive, whereas % of group b were still in remission, indicating a role for alloreactivity in lymphoid neoplasia. concerning expression of nk receptors, group a was characterized by the high expression of kirs, potentially involved in alloreactivity in out of patients. klr was represented by cd /nkg a in out of and in one case by cd /nkg c. this last case showed a recovery of cd /nkg a phenotype after months. ncrs and nkg d were usually expressed, with exception of nkp . group b was characterized by an heterogeneous pattern of expression of kir, whereas cd /nkg a was expressed in out of cases and cases expressed nkg c. interestingly, one of these two cases relapsed during follow-up. ncrs were usually expressed, with the only exception of nkp . our data indicate that nk alloreactivity might be an advantage for survival in patients affected by acute lymphoid neoplasia. no correlation could be demonstrated with expression of natural killer receptors (nkrs) and clinical behaviour, suggesting that analysis at clonal level is mandatory to get insights into the mechanism involved. this study was supported by a grant from fondazione città della speranza. a rare complication after allogeneic stem cell transplantation: acute axonal neuropathy a.d. moicean, d.n. colita, v.n. mirea, a.m. dumitrescu, t. puscariu, z. varady, a. tanase, i. constantinescu fundeni university institute (bucharest, ro) despite considerable progress in the management of allo-hsct, infection remains an important cause of morbidity and mortality after transplant. cmv is frequently involved in the infectious pathology after hsc transplantation. we present a -year old women with high-risk acute lymphoblastic leukemia who underwent matched sibling pbsc transplantation. she received standard conditioning regimen (endoxan mg/kgc and tbi gy). gvhd prophylaxis received was with csa and methotrexate. she developed grade ii gvhd (cutaneous and intestinal) on day + with resolution under corticotherapy. starting with day + she had weakness, osteoarticulary and muscular pain with functional impairment. neurological examen and the electrophysiological study showed an acute axonal neuropathy. she had more than cmv copies in the blood pcr. she received mg bid i.v ganciclovir for days as induction therapy and than mg/d for more weeks with complete remission of the neurological symptoms a year old female patient was diagnosed with cml in chronic phase and stem cell transplantation (sct) was planned from her one antigen mismatched sister. pre-transplant routine investigations revealed a high antibody titer against toxoplasma gondii (ift : , kbr : , igm negative) indicating the history of a previous infection. cerebral magnetic resonance imaging (mri) ten days before transplantation showed no abnormalities. the patient was conditioned with busulfan ( mg/kg) and cyclophosphamide ( mg/kg). because of the hla-a mismatch rabbit atg (fresenius, mg/kg) was added. graftversus-host disease (gvhd) prophylaxis consisted of cyclosporine a and mtx. pbscs containing . * /kg cd pos stem cells were transfused. engraftment occured rapidly on day + . on day + the patient developed severe headache and hours later generalized seizures. computered tomography showed multiple patchy hypointense lesions in the subcortical white areas with corresponding nodular signal enhancements in t weighed mri analysis suggesting an infectious process. eeg revealed pronounced focal abnormalities in the left hemisphere extending from frontal to the temporal regions. liquor analysis revealed no causative infectious agent. assuming reactivation of toxoplasmosis we initiated a specific quatruple therapy consisting of pyrimethamin orally, sulfadiazin orally, clindamycin i.v. and tmp-smx i.v. clinically the patient improved rapidly. two weeks later cerebral mri showed regression of the temporoparietal white matter lesions with further improvement during the following weeks resulting finally in complete resolution. antibody titers against toxoplasmosis slowly regressed during the months following transplantation under continuation of oral specific therapy. the patient is now more than one year after stem cell transplantation alive and well without signs of toxoplasmosis and without neurological sequela. toxoplasmosis is a rare but dreaded complication usually developing between and months after stem cell transplantation with a high mortality rate. cml patients seem to be at increased risk for reactivation of toxoplasmosis after stem cell transplantation. the course of our patient, however, is unusual because of the very early manifestation after stem cell transplantation, the rapid course and prompt response to treatment. mucositis is a common side effect of chemotherapy and radiotherapy with no effective treatment. it occurs when cancer treatment destroys the rapidly dividing epithelial cells, particularly in the oral cavity, leaving the mucosal tissue open to ulceration and infection. the aim of this study was to assess the state of oral mucosa in a patient after allo-pbsct who has received palifermin, a recombinant human keratinocyte growth factor. materials and methods: a -year-old male was treated in the department of haematology of the medical university in warsaw due to the aml. conditional chemotherapy was applied, according to the bucy + atg regimen and allogeneic haematopoietic cells transplantation from an unrelated donor. he was receiving palifermin ( microg/kg/d) intravenously for consecutive days immediately before the initiation of conditioning therapy (on days - , - , - ) and after allo-pbsct (on days + , + , + ). on day + the oral mucous membrane was pale and swollen, with linea alba visible on cheeks. superficial glossitis and viral pharyngitis were noted. beginning with day + /+ proliferative gingivitis was observed. on day + gingival contour was altered and the gingiva covered nearly completely tooth crowns of all teeth. the gingiva were whitened, as if covered by thick epithelium. slight gingival hyperplasia was still observed on day + . during the forming of gingival hyperplasia the patient had a subjective "membrane growing" sensation with tingling and itching. he reported an oral cavity pain score of in the -point pain scale. since day + /+ skin rash coexisted, spreading over hairy head skin, face, dorsum and chest. disseminated papulopustular (acne-like) lesions were observed. some of them were related to the hair follicles. skin changes were present till day + . neutropenic fever was noted on day + (absolute leucocytosis . g/l). concomitant medications: orungal x mg p.o., heviran (aciclovir) x mg p.o., tazocin x . g i.v. on days + and + , maxipime x . g i.v. on days + till + , vancomycin x g on days + till + , metronidazole x mg i.v. on days + till + , neoral x mg i.v. since day + and x mg since day + , zyrtec tablet/d since day + . conclusions: palifermin is an efficient pharmaceutical in mucositis prevention in patients after allogeneic pbsc transplantation. transient complication of hyperplastic gingivitis with a concomitant skin eczema of a papulopustular nature arose. progressive fatal respiratory failure due to pseudomembranous aspergillus tracheobronchitis following allogeneic stem cell transplantation p. sedlacek, p. hubacek, k. zdrahalova, p. pohunek, o. nyc, p. pavlicek, j. stary university hospital motol (prague, cz) invasive fungal infections are frequent and often fatal in patients following allogeneic hematopoietic stem cell transplant (hsct). long-term immunosuppressive therapy for graft versus host disease (gvhd) seems to be the most predisposing factor. we present a year old girl with paroxysmal nocturnal hemoglobinuria (pnh). due to unfavourable course (severe attacks of hemolysis requiring hemodialysis, budd-chiari syndrome) she was indicated for hsct. she consequently failed to engraft two unmanipulated grafts from hla mismatched (b, cw alleles) unrelated donor. first time bone marrow was infused with cd x / /kg, conditioning consisted of treosulfan, cyclophosphamide and atg. second time peripheral blood stem cells (pbsc) were used with cd x / /kg after fludarabine, cyclophosphamide and atg. after months lasting aplasia third allogeneic graft (pbsc -cd , x / /kg) was given with no conditioning surprisingly followed by rapid and full trilineage engraftment. she than developed severe acute and chronic extensive gvhd, treated with cyclosporine a, steroids, tacrolimus, mycophenolate, sirolimus and antihymocyte globuline. during post-transplant course she suffered from bkv hemorrhagic cystitis, repeated cmv reactivations and colitis, drug induced nephropathy and steroid diabetes. she had a long-term and lasting preemptive voriconazole prophylaxis. for acute hemodynamic instability due to severe gastrointestinal bleeding months after hsct she was transferred to icu and electively intubated. at that time she was also heparinized for acute vein thrombosis. early on she started to desaturate, chest x-ray showed unilateral atelectasis. bronchoscopy revealed whitish membranes, plugs and casts causing extensive obstruction of both lungs. cultures grew aspergillus fumigatus. therapy including caspofungin, nebulized amphotericin b and repeated mechanical removal of obturating membranes failed to stop progression and patient died due to isolated respiratory failure days upon arrival to icu. autopsy confirmed fungal involvement of trachea, larynx and bronchi. pseudomembranous mycotic tracheobronchitis may be a rapidly progressing complication in heavily immunocompromised patients. complex therapy including even combination of potent antifungals may fail to overt fatal course. even in patients on antifungals this complication must be actively looked for with early use of bronchoscopy and exact identification of pathogen. systemic candidiasis is a rare but life threatening complication in immunosuppressed patients undergoing allogeneic sct. combination of new antifungal agents might improve outcome in these patients. here, triple anti-mycotic therapy is described in an all patient in urgent need of allogeneic bone marrow transplantation. the patient with t-cell acute lymphoblastic leukemia of thymic differentiation achieved remission after treatment according to the gmall / protocol. two months after the consolidation therapy relapse resistant to treatment regimens containing fludarabin, cytarabin, etoposide and amsacrine occured. even after claeg (cladribine, etoposide, cytarabine and campath- h) the patient still had % leukemic bone marrow blasts requiring high dose chemotherapy with allogeneic stem cell transplantation. one day after start of the conditioning regimen the patient showed mycotic skin manifestations and blood cultures became positive for candida cruzei despite fluconazol prophylaxis. because of the limited sensibility of fluconazol resistant candida species to liposomal amphotericin b and the high mortality rate in patients with systemic candidiasis voriconazol was added immediately to liposomal amphotericin. subsequently, the patients body temperature increased and caspofungin was added. since the mycotic skin manifestation responded to this triple anti-mycotic combination allogeneic peripheral blood stem cell transplantation from an unrelated donor could be performed. altough the fever resolved later the patient showed signs of a septic shock requiring intravenous administration of dopamine. with the unchanged triple antifungal therapy the patient became afebrile, skin manifestations showed complete resolution and cultures became negative. three months after the onset of systemic candidiasis the patient was fully active with no signs of fungal infection and in haematological and molecular remission. this case shows, that the entire intense conditioning chemotherapy could be administered and allogeneic stem cell transplantation is feasable in patients with systemic candidiasis when such combined antifungal treatment is given. i. vrelust ( ), a. gadisseur ( ), e. steel ( ), w. schroyens ( ), a. van de velde ( ), z.n. berneman ( ) ( neutropenic patients, especially following aggressive chemotherapy, are at high risk for infectious complications. these are an important contributary factor to the treatment related morbidity and mortality (trm). because neutrophils represent the first line of host defense, granulocyte transfusion therapy is a tempting therapeutic approach. although such therapy has been employed sporadically for several decades, clinical benefit has been compromised by technical problems and low granulocyte yields resulting from inadequate donor stimulation. the discovery of granulocyte colony-stimulating factor (g-csf) as a means to elevate blood neutrophil counts in healthy donors has rekindled interest in granulocyte transfusion therapy. we describe a -year old female patient with an acute myeloid leukaemia (aml) who underwent remission induction chemotherapy. in absence of repopulation of the peripheral blood the bone marrow showed persistent aml. despite antifungal prophylaxis the patient developed an invasive pulmonary aspergillosis. voriconazole, g-csf and caspofungin were added, and granulocyte infusions were started because of the continuing neutropenia. the granulocytes were collected from donors with a compatible blood group through leucaferesis after stimulation with g-csf and dexamethasone ( mg), and then irradiated. the transfusions were given three times a week. conditioning for a reduced intensity (ric) allogeneic peripheral blood stem cell transplant (pbsct) with a hla-identical brother was started. in total granulocyte infusions were given; of which after transplantation, the last on day + after pbsct. they were discontinued when the anc count reached . x /l. all granulocyte infusions were tolerated very well. four months after transplantation she is in a complete hematologic remission without signs of graft-versus-host-disease (gvhd), and without signs of active pulmonary infection. the precise role of donor granulocyte infusions remains to be delineated, partly because of the lack of defined clinical trials. we conclude that granulocyte transfusion therapy may be useful for neutropenia-related fungal infections in patients with hematologic malignancies. the use of granulocyte infusions during a ric-pbsct procedure does not seem to lead to an increased risk of gvhd or hamper engrafment itself. a. kerguelen, m. canales, a. lopez, m. martin, t. cobo, d. hernandez, j. g-bustos, f. hernandez-navarro university hospital la paz (madrid, e) introduction: patients with haematological diseases previously diagnosed with invasive fungal infection (ifi) are considered to be at high risk of suffering reactivation of the infection during subsequent intensive chemotherapy. methods: in the last years patients with haematological diseases ( aml and acquired aplastic anaemia) and previous invasive aspergilosis have undergone allogeneic haematopoietic stem cell transplant in our centre. all patients received as primary antifungal therapy combination of liposomal amphotericin b (ambisome) and caspofungin displaying complete or good partial radiological resolution of the infection. itraconazole and voriconazole was continued as secondary prophylaxis. conditioning regimen consisted of busulfan and cyclophosphamide in patients with aml and atg and cyclophosphamide in the aplastic anaemia patient. cyclosporine in combination or not with methotrexate was the regimen administered to prevent ghvd. results: in a patient with aml no clinical or radiological signs of reactivation of fungal infection were observed through the transplant procedure. in the other patient with aml itraconazole was changed by liposomal amphotericin because of unresponsive fever. in the patient with aplastic anaemia combined therapy with amphotericin and caspofungin was initiated because of radiological worsening, but galactomannan antigen was negative in all analysis performed. no patient has died because of infectious complication during or after transplant. conclusion: availability of new antifungal agents does allow pre-transplant therapy of previous ifi, aiming to achieve a clinically undetectable state of infection, and an adequate antifungal treatment during transplant to diminish risk of reactivation of fungal infection in allografted patients. however, the optimal use of antifungal agents or their combinations remains to be determined and more studies are necessary to confirm our experience. the frequency of invasive fungal infections, in particular infections due to aspergillus and other moulds, has increased over the past two decades. whereas invasive aspergillosis mainly involves the respiratory tract, lung or sinus, the lower gastrointestinal tract is rarely affected, most often in the frame of secondary dissemination. primary invasive aspergillosis of the gut is a rare event associated with high mortality and has not been reported to date in patients after autologous stem cell transplantation (sct). we report on a -year-old boy who developed isolated intestinal aspergillosis soon after autologous stem cell transplantation for pnet of the central nervous system. the boy received broad-spectrum antibiotics because of neutropenic fever, and antimicrobial prophylaxis included trimethoprim-sulfamethoxazole, metronidazole, acyclovir, fluconazole and topical amphotericin b. antimycotic therapy was started because of persistent fever and abdominal pain, and rising serum levels of galactomannan and the isolation of aspergillus fumigatus from the stool suggested invasive aspergillosis. the boy underwent enterostomy on day + , and diagnosis of intestinal aspergillosis was pathohistologically confirmed. no other site of invasive aspergillosis was evident. the patient was treated with antimycotic combination therapy consisting of liposomal amphotericin b, voriconazole and caspofungin. the clinical condition slowly improved over the next months. enterostomy was removed on day + and antimycotic treatment has been stopped soon after. currently, on day + , the boy is at home without major gastrointestinal complaints. we conclude from this case that primary intestinal invasive aspergillosis can occur in patients undergoing autologous stem cell transplantation, and therefore, this diagnosis has to be considered in this setting in patients suffering from fever and abdominal pain. in case of positive galactomannan antigenemia, an extensive search for invasive aspergillosis should be performed and, at the same time, early antifungal therapy should be started. mycobacterial disease is a rare and difficult diagnosis in hematopoietic transplant recipients. we report the case of a twenty-year-old woman who underwent a second matched unrelated donor (murd) hsct for a null all in second complete remission. conditioning regimen consisted on cyclophosphamide mg/msq; busulfan mg/kg and thiotepa mg/msq. on day + , while receiving acyclovir prophylaxis, and a previous cotrimoxazole prophylaxis, she developed an acute ascending paraparesis with anaesthesia and reflex loss and loss of sphincter control. mri imaging showed gadolinium contrast enhancement on cauda equina roots. csf analysis showed elevated cell count ( /mm³, normal range nr: - /mm³), elevated csf protein concentration ( mg/dl: nr: - mg/dl), and no hypoglucorraquia ( mg/dl with simultaneous plasma concentration of mg/dl). flow cytometry analysis showed % t lymphocytes, . % non clonal b lymphocites, , % monocytes and , % neutrophils. bacterial, fungal and mycobacterium presence was ruled out and empirical antiviral treatment consisting on foscarnet ( mg /kg /day), immunoglobulin ( . g/kg/ hours) was started. metilprednisolone g per day for tree days flowed by mg per day for three consecutive weeks. no clinical response was observed and neurotrophic viral infection was excluded by pcr technique. a second lumbar puncture was performed days later showing a lower number of lymphoid cells ( /mm³) but with an increase in protein concentration ( mg/dl) without hypoglucorraquia; moreover auramine positive rods were present. antituberculous treatment with rifampicin ( mg/d), isoniazide ( mg/d), ethambutol ( . g/d) and pyrazinamide ( g/d) were started, with no clinical improve but no worsening. hepatic toxicity was developed and isoniazide an rifampicin were suspended after a month of complete treatment and ofloxacin ( mg/ h, ethambutol and pyrazinamide was started as a second-line therapy. this is to our knowledge the first reported case of transplantationrelated tuberculous arachnoiditis. pentastomiasis of the liver in a patient following unrelated stem cell transplantation with estimated hepatic acute graft-versus-host disease j. dahlke ( ) , r. kobbe ( ), a.a. oyekunle ( ) , f. ayuk ( ) , n. kröger ( ) we report a case of human hepatic pentastomiasis (armillifer armillatus) in a -year old man with aml who immigrated to germany from togo in and died as a result of relapse, days post hla-matched unrelated stem cell transplantation. at day + post transplantation, diarrhoea occurred and gastroscopy with excisional biopsy revealed an acute graft versus host disease of the stomach and upper small bowel. for treatment we started with a tapering schedule of methylprednisolone ( mg/kg body weight/day). three days later we observed a hyperbilirubinemia and slightly increased liver enzymes. the abdominal ultrasound showed an increased size of the liver and an increased portal perfusion. the findings were interpreted as a hepatic involvement of the acute graft versus host disease (agvhd) and a salvage treatment with mycophenolat mofetil was initiated. the patient died as a consequence of intracerebral bleeding in a cerebral infiltrate of the leukaemia. at autopsy, in addition to the cerebral findings, we found multiple pentastomides (documented by photographs) up to cm length (spec. armillifer armillatus) subcapsular, in the parenchymatous tissue and in the portal veins. no agvhdlike inflammatory infiltrate in the liver was observed. late effects and quality of life r trilineage hypoplasia after initial neutrofil engraftment in patients with acute myeloid leukaemia undergoing autologous transplantation with bu/cy conditioning: frequency, outcomes and prognostic significance (case report) a. pivkova, l. cevreska, n. siljanovski, z. stojanoski, s. genadieva stavrik, i. panovska, s. krstevska balkanov, s. trajkova, b. georgievski clinical center (skopje, mk) myeloablative conditioning with autologous stem cell transplantation (asct) is a treatment option for aml patients with lack of sibling donor. hematopioetic engraftment may be prolonged depending on the type of myeloablative regimen, mobilization regimen, dose of mnc and patients age. in a year period ( ) ( ) ( ) ( ) ( ) ( ) we realized a total of autologous transplanations with cryopreserved peripheral blood cells (pbsc) and busulfan and cyclophosphamide conditioning. we present two cases ( %) of aml (standard risk) patients (two females and years at transplant) that underwent autologous transplantation at department of hematology, skopje. mobilisation of pbsc was preformed with vp- mg/msq + g-csf µgr/kg in one patient and hd-arac gr/mscx , idarubicine mg/msqx +g-csf µgr/kg in the other patient. a minimum of , x /kg mnc and , x /kgmnc respectively were collected and preserved in % dmso solution. we registered engraftment for ne> , x /l on day + and + and for plt> x /l on day + and + . patient were followed up in outpatients and pancytopenia was registered one month after transplant with plt< x /l with mild haemorrhagic complications plt transfusions dependable, hb < g/dl and red cell transfusion dependence (on two week interval), wbc < , x /l with cytokine treatment (twice a week g-csf µgr/kg). bone marrow biopsy revealed trilineage hypoplasia, no signs of organomegaly, no microbiological and viral findings. one patient has recovered completely months after transplantation and the other is still in good physical condition but present pancytopenia months posttransplant. we conclude that prolonged pancytopenia is due to bu/cy conditioning, the age could be a significant factor for starting myeloablative conditioning prior autologous transplant as well as minimum mnc dose could prolonge immune reconstitution. neurological long-term follow-up after allogeneic bone marrow transplantation p. sostak, c. padovan, m. holtmannspötter, g. ledderose, h. kolb, a. straube klinikum großhadern (munich, d) to improve our knowledge about the neurological outcome after allogeneic bone marrow transplantation (bmt), we have started a prospective study already years ago. so far we could show that within the first year after transplantation a significant proportion of patients ( %) had developed neurological sequelae. besides well-defined neurological complications more than half of the study population suffered from new neurological abnormalities of unknown origin predominantely affecting the peripheral nervous system. in a small subgroup of patients already central nervous signs, cognitive deficits and white matter lesions could be detected and this was in relation to chronic graft-versus-host disease (gvhd)/immunosuppression. to determine whether central nervous system (cns) involvement during gvhd might manifest at a later time in more bone marrow recipients, the follow-up period now was extended and patients again received a neurological examination, underwent a neuropsychological test battery and standard mri sequences. long-term follow-up results will increase the insight onto the spectrum, incidence and etiology of neurological sequelae after allogeneic bmt. they will be discussed in relation to retrospective and experimental data, which suggest involvement of the cns during gvhd. keratoconjunctivitis sicca with recurrent calcium deposition in the cornea and severe visual loss due to gvhd p. keslova ( ) we present severe keratoconjuctivitis sicca with recurrent calcium deposition in the cornea after keratoplasty in a patient with extensive chronic graft versus host disease (gvhd). a -years-old girl with myelodysplastic syndrome (raeb) underwent stem cell transplantation (sct) using peripheral blood stem cells of two alleles mismatched unrelated donor (b, cw). conditioning regimen consisted of busulfan, cyclophosphamide and melphalan, for gvhd prophylaxis combination of rabbit antithymocyte globulin (atg fresenius) and cyclosporine a (csa) with short term methotrexate were used. due to renal toxicity csa was early switched to tacrolimus and corticosteroids were started at day+ . week after she developed steroid resistant acute gvhd grade iv with skin and gut involvement. for that she was successfully treated with combination of steroids, mycophenolate mofetil and sirolimus (day+ through day+ ). during period of acute gvhd severe keratoconjuctivitis sicca with deep corneal defects has been developing and visual acuity had deteriorated. there were increasing pancorneal calcium deposits (calcareous degeneration). serum calcium and phosphate levels were normal at several times points. longterm immunosuppression improved symptoms of gvhd but healing of corneal defects was not reached despite local therapy of antibiotics and corticosteroids. at day+ patient underwent removal of calcium deposits and both eyes were covered by amniotic membrane. at day+ keratoplasty in the left eye was performed with transient improvement of visual acuity. two weeks after keratoplasty calcified plaques have recurred in the transplanted tissue and fast visual loss reappeared. we suggest that ectopic corneal calcifications are probably associated with persistent epithelial and stromal defects and keratoconjunctivitis sicca as a symptom of persistent active chronic gvhd. combined immunosuppressive therapy should therefore continue. we plan to repeat surgery at the later time under better control of gvhd. patient is now one year after sct with no signs of leukaemia relapse with full donor hematopoiesis, but prognosis of vision still remains at this moment uncertain. supported by vz fnm mzo thyroid dysfunction is an important problem in patients receiving bone marrow transplantation. however there was no case of hashimoto encephalitis in a patient after stem cell transplant reported. we describe the case of year old female patient who underwent allogeneic bone marrow transplantation and who developed hashimoto encephalitis. aml patient was subjected to allogeneic stem cell transplant with reduced intensity conditioning (flag-ida) in october . early posttransplant period was complicated by reactivation of cmv infection and prolonged peripheral cytopenia. months after transplant the patient chimerism analysis revealed graft rejection without aml relapse. months after transplantation the patient developed fatigue, loss of appetite, vomiting but also psychiatric symptoms: hallucinations and paranoid ideations. ct scans does not revealed anything specific. typical psychiatric treatment was not efficient. later patient experienced episodes of epilepsy and developed cerebellar ataxia and progressive unilateral paresis with impaired consciousness. in accessory investigations -in csf elevated level of proteins and eeg abnormalities were noted, blood and csf cultures were negative, thyroid hormones level were slightly decreased, tsh and atpo antibodies titers were elevated. results of mri confirmed disseminated pathologic changes in white matter. hashimoto disease with encephalitis was diagnosed and the patient was treated with high dose methylprednisolone i.v. for days with rapid improvement noticed within first h of the treatment. the neurological state normalize within one week. maintenance treatment with decreasing dose of oral prednisone is carried out. occurrence of hashimoto encephalitis in described patients seems to be connected with chronic graft rejection process and impaired prolonged regeneration of immunity after transplant. the role of different infections mainly viral should also be investigated. bmt: care about donors a. tellier, c. bauchetet, z. marjanovic, j.-p. marie, b. rio hotel-dieu paris (paris, f) purpose: bmt improvement must take in consideration ethical issues regarding donors. based on donors refusal cases report, our analysis underlines the significance of identifying «unwilling donors». moreover, we are concerned about the process of obtaining informed consent of family membres to undergo hla histocompatibility tests in order to prevent psychological consequences of peripheral blood and bone marrow donation. background: studies about psychological issues of bone marrow donation show that donors may be worried about their health status (switzer et al., ; molassiotis et holroyd, ) . switzer et al. correlate donors difficulties to their hesitation during donation decision process and consider that screening donors motivations is useful to a psychoprophylaxis approach. other authors note that families donors are even more exposed to psychological problems (chang et al, ) . directly concerned by recipient reactions to bmt, family donors (wollcott et al, ) may feel unconscious guilt in cases of unsuccessful or complicated bmt (futterman and wellisch, ) . deeper psychodynamics of bmt process indicates that donor can confuse biological (hla) and psychological identity and be anxious about being ill. (alby, (alby, , ascher, ascher, , . topall-rabanes et al. ( ) notice that about % of studied family donors are classed as «reluctant» : for these subjects, donation decision is not considered as a real choice. they suffer from health problems and feelings of regret even longtime after donation. methods and results: we reviewed the records of donor refusal cases representing % of allografts performed in our twenty years bmt practice. in-depth analysis of patients psychosocial situation, quality of social support, family members relationships, patient-family-staff communications modalities, in particular regarding to donation information process. we will present these cases and retrospective analysis to introduce ethical questions on hla histocompatibility testing. conclusion: regarding to our experience and to our data, it seems that information process of family membres about bone marrow donation may lead to an impossible choice. appointed by hla testing as donor, family member is moved to suffer of biological determination and to become «unwilling donor». protecting these persons is a medical responsability. more research is necessary on this subject. there are patients with paranoiac reactions (persecution, sensitive, litigious, invention) were possible to allocate among the mental disorders at our patients. results: paranoiac reactions with ideas of persecution (n = ) are the most often in our sample. in such cases patients suspect those around them, especially medical personnel, in preconceived attitude to them and, possibly, to conspire for damnification of patients. such ideas have a systematic character and are resistant for treatment. sensitive paranoiac reactions (n = ) are joined with perception of physical handicap and characterized by sensation of slighting attitude and mockery those around patient and by confidence in dissemination of detractive rumours. litigious paranoiac reactions (n = ) are characterized by lowsystematized and insufficiently well-founded requests and multiple joined with hematological malignancies such as requests to compensate the prejudice applied by disease or laying claims to medical personnel are at fault, in opinion of patient, in the prejudices joined with disease. invention paranoiac reactions (n = ) in our sample are characterized by elaboration of self-treatment methods of his/her disease. treatment of examined states was significantly resistant and included antipsychotic and anxiolitic medications. conclusions: pr at the patients with hemato-oncological disorders after bone marrow transplantation are the separate problem demanding the special attention both the hematologist and the psychiatrist, especially relative to its treatment. b.v. afanasiev ( ), n.v. stancheva ( ), l.s. zubarovskaya ( ), e.v. semenova ( ), m.a. ovsyannikova ( ), t.i. ionova ( ), t.p. nikitina ( ), a.a. novik ( ) ( )st. petersburg state medical university (st. petersburg, background: quality of life (qol) is increasingly used as a treatment outcome along with traditional clinical outcomes in children with cancer undergoing bone marrow transplantation (bmt)/stem cell transplantation (sct). the aim of our study was to estimate qol parameters and symptoms in survivors of childhood blood cancer after bmt/sct. patients and methods: fifteen survivors were evaluated - years (median, years) after allogeneic bmt/sct for acute leukemia ( ), chronic leukemia ( ) and myelodysplastic syndrome ( ) . median age at transplantation - yrs (range . - . ), girls/boys- / . pedsql™ generic core scales and sf- questionnaires were used for qol assessment in the group younger than yrs at the time of the survey and in the group yrs and older, respectively. nj children cancer symptom inventory and md anderson symptom inventory were used for symptom assessment in the younger and older groups, respectively. for comparison healthy controls ( for younger group; -for older group) matched to survivors by age and gender were included in the study. results: no significant differences in qol parameters (physical, psychological and social functioning) between survivors and control group were revealed. only school functioning for children younger than yrs was lower in the group of survivors ( vs , p< . ). seven survivors experienced moderate or severe symptoms ( to scores on - scale). four of them had pronounced psychological symptoms. other pronounced symptoms were chronic pain, fatigue, lack of appetite, shortness of breath, drowsiness and nausea. six survivors had at least two moderate or severe symptoms. qol parameters, namely, physical, psychological and social functioning in long-term survivors of childhood blood cancer post allogeneic bmt/sct is comparable to healthy people. however, nearly half of survivors experience different symptoms in long-term period after transplantation. this confirms the importance to monitor and control late related symptoms in order to preserve qol of long-term survivors of childhood blood cancer. differential diagnosis of t-cell lymphoproliferative disease after allogeneic haematopoietic progenitor cell transplant s. koschmieder, m. stelljes, a. schmidt, g. silling, t. spieker, g. köhler, c. renne, w.e. berdel, j. kienast university of muenster (muenster, d) two cases are presented of a rapidly developing cervical lymphadenopathy days after allogeneic peripheral blood stem cell transplant for acute myeloid leukemia. patient # , a year-old female, developed painful bilateral cervical adenopathy (up to cm) overnight, fever ( . °c), and a fine maculopapulous rash. no further enlarged lymph nodes were found by computed tomography, and the right cervical mass was removed. histology showed infiltration of the lymph node by atypical lymphoid cells that expressed cd and cd . molecular analysis revealed monoclonally rearranged tcr gamma chains. histology of the skin showed a leukocytoclastic vasculitis without any signs of acute gvhd. ebv pcr was consistently negative in peripheral blood and lymph node. the diagnosis of t-ptld was made, and the patient received a -day course of methylprednisolone. with this regimen, the patient´s condition rapidly improved, and the fever, lymphadenopathy, and rash resolved completely within a few days. patient # , a year-old female, developed painful acral bullous erythematous lesions on her arms, legs, and in her mouth on day post transplant which progressed to generalized maculopapulous rash. histology showed acute graft versus host disease (gvhd) of the skin. on day + , the patient presented with painful bilateral cervical adenopathy (up to . cm), and abdominal ultrasound and mri showed no other regions of lymphadenopathy. histology of one removed lymph node demonstrated a highly proliferating, diffuse infiltrate of cd positive t cells with % of t cells expressing cd and % expressing cd , respectively, as well as scattered cd positive cells. molecular analysis detected a polyclonal pattern of tcr rearrangement and an oligoclonal immunoglobulin receptor rearrangement. ebv pcr was negative in peripheral blood and lymph node. the clinical scenario was interpreted as concomitant lymphadenopathy associated with acute gvhd, and the patient received methylprednisolone for days and completely recovered. this report highlights the necessity to remove suspiciously enlarged lymph nodes, developing after allogeneic transplant, in order to distinguish between t-ptld and lymphadenopathy accompanying acute gvhd. the evaluation of t-cell receptor gamma gene expression for prognosis of relapse development after stem cells transplantation a. sipol, d. butlitskiy , u. vorobjeva , a. aljanskiy , m. zarayskiy , l. zubarovskaya, b. afanasyev st. petersburg state med pavlov university (st. petersburg, rus) background: the hematopoetic stem cells transplantation (hsct) in patients with hematological disorders is the most radical method of the therapy. the early diagnostics of relapse in post-hsct period could improve significantly the therapy effectiveness. the purpose of this study was to develop the minimal residual tumor cells detection method by studying the mediated mechanisms of organism's reaction to tumor clone. materials and methods: nineteen patients with different hematological malignances, who were undergone the hsct, have been included in the research. total mrna was extracted from peripheral blood leukocytes, sampling in the time of conditioning regime completion, just prior to stem cells transfusion. we performed the rt-pcr with primers specific to v-j-gamma junctions (tcr-gamma gene). specific signal was detected in % agarose gel. results: absence of tcr-gamma gene expression at day- were significantly more often in group of the patients who were staying in remission after hsct (p= . ). in group of the patients with myeloablative pre-transplant conditioning the significantly differences in tcr-gamma gene expression depending on the fact of relapse has not been revealed. in group of the patients with reduce intensity conditioning regime the positive correlation between the presence of tcr-gamma gene expression at day- and relapse in post-hsct period was observed (p= . ). conclusion: the definition of tcr-gamma gene expression before hematopoietic stem cells infusions is the method of a tumor process activity estimation during the therapy, especially in case of using the reduce intensity conditioning regime. we suggest this criterion as the early prognostic factor for the relapse developing in post-hsct. mrd-directed adoptive immunotherapy following allogeneic stem cell transplantation fails to permanently revert disease progression in childhood acute lymphoblastic leukaemia between viii/ and vii/ we performed allogeneic haematopoietic stem cell transplantation (hsct) in consecutively children with acute lymphoblastic leukaemia (all). we analysed prae-and post-transplant minimal residual disease (mrd) levels using quantitative rq-pcr targeted to immunoglobulin and/or t-cell receptor rearrangements in of them with available targets (with adequate sensitivity and specificity according to esg-mrd-all criteria). seven of patients with detectable mrd prior hsct (n= ) relapsed after transplant and one died in ccr before day + due to transplant related complications. in the group of prae-hsct mrd negative patients (n= ), only one relapse appeared. in a total of patients, there was a time-frame for an attempt to avert the disease progression detected by rq-pcr. in one bcr/abl+ patient, imatinib mesylate dose was increased and doses of dli ( . x /kg cd + cell) were administered. despite this, patient relapsed + days after hsct. second bcr/abl+ patient developed molecular relapse despite chronic gvhd. therefore, immunosuppression was quickly discontinued and imatinib mesylate was administered. he achieved temporary molecular remission but months later died of cns disease progression. third patient developed mrd positivity + days after hsct and therefore immunosuppression was quickly tapered. gvhd reactivation required steroid and csa treatment. after gvhd resolution immunosuppression was ceased and three escalating doses of dli ( x , x , x cd +/kg) were given. nevertheless, this did not avert haematological relapse. this patient after high-dose chemotherapy achieved second complete remission (cr) with mrd negativity prior to second hsct and now is months after hsct in continuous cr and mrd-negative. the fourth patient (bcr/abl+) developed relapse despite three dlis and imatinib mesylate treatment given for positive mrd + after hsct. adoptive immunotherapy (rapid cessation of immunosuppression, infusion of dli in to weeks interval and/or use of imatinib mesylate in bcr/abl+ all) after the transplantation was not successful in our cohort in relapse prevention and might only postpone the manifestation of relapse and facilitate further efficacious chemotherapy and retransplantation. we are confident that all effort should be aimed to a better control of the pre-transplant mrd levels. grant support: fnm , mz - , gauk / , mz and msm . the dynamics of chimerism evolution determines the differential outcome of various transplant settings i. buño, p. balsalobre, g. iglesias, d. barroso, c. manzano, r. carrión, d. serrano, a. gómez-pineda, j.l. díez-martín hosp. g.u. gregorio marañon (madrid, e) background: several factors such as the intensity of the conditioning regimen, the t-cell content of the graft or the gvhd prophylaxis, influence the degree of chimerism after sct. objective: to evaluate the dynamics of chimerism after different sct settings (ablative, reduced intensity conditioning -ric-and t-cell depleted -tcd-) and its influence in the success of the procedure. patients and methods: sct: ablative (including from mud), ric and tcd (including from haploidentical donors and from mud with hla disparity). chimerism analysis was performed by fish or str-pcr (sensitivity %). samples: bone marrow (bm) and peripheral blood (pb) on days + , + , + , + and once a year thereafter, as well as pb and leukocyte lineages (t lymphocytes cd +, b lymphocytes cd + and myeloid cells cd + isolated (purity > %) by immunomagnetic means, automacs, miltenyi biotec), every weeks, starting on day + and until complete chimerism (cc) was achieved. results of chimerism follow-up were censored once the diagnosis of relapse or rejection was established. results: incidence of mixed chimerism (mc) on day + (ablative: bm %, pb %; ric: bm %, pb %, cd %; tcd: bm %, pb % , cd %), as well as its dynamics (mc on day + : ablative bm %, pb %; ric bm %, pb %, cd %; tcd bm %, pb % , cd %) were different in the three sct settings. moreover, the percentage of recipient cells (%r) was significantly higher after ric and tcd than after ablative sct, as well as in t lymphocytes than in bm or pb ( / cases with simultaneous studies showed mc in t lymphocytes and cc in pb). all ric sct evolved to cc by day + while tcd sct showed persistent mc ( patients with stable mc after one year). the incidence of rejection was greater after ric ( / ) and tcd ( / ) than after ablative sct ( / ). all these patients showed mc, mainly in t lymphocytes, which allowed early diagnosis and successful treatment with immunosuppression withdrawal and donor leukocyte infusion. patients with cc in pb/t lymphocytes on day + had a higher incidence of gvhd>i than those with mc. in the present series, however, a relationship between chimerism and relapse, disease free survival or overall survival, was not observed. conclusions: sct with greater incidence of mc (ric and tcd) favor immune tolerance between donor and recipient which reduces the risk/severity of gvhd at the expense of a higher incidence of graft rejection. reliable quantification of haematopoietic chimerism after allogeneic stem cell transplantation by real-time quantitative pcr analysis a. picardi ( ) introduction: increasing mixed chimerism (mc) represents a poor prognostic factor after allogeneic stem cell transplantation (sct). moreover, to define the best timing of immune-suppression withdrawal and donor lymphocytes infusion, a strict monitoring of donor hemopoiesis is needed. methods: we evaluated donor/recipient pairs using a quantitative real-time pcr (qrt-pcr) with the aims ) to evaluate the informativeness of this chimerism assay and ) to compare qrt-pcr analysis with standard methods such as fluorescence in situ hybridization (fish) for mismatched sex pairs or variable nucleotide tandem repeats (vntr) for matched sex pairs. qrt-pcr (lightcycler . , roche) was performed on bone marrow and peripheral blood samples collected monthly, using eleven biallelic dna genetic system located on chromosomes , , , , , , , x and y. glyceraldeyde phosphate dehydrogenase (gapdh) gene was used as active reference. before quantification, donor and recipient dnas were genotyped using primers and probes specific for all genetic markers. patients had a median age of . years (range - ) and were affected by acute leukemia (n= ), or lymphproliferative disorders (n= ). standard regimen was used in cases, reduced intensity conditioning in , while patients underwent an unrelated sct. median follow-up of the patients was . months (range . - . ). results: both qrt-pcr and fish detected donor/recipient differences in % of pairs, while vntr was not informative in % of sex matched pairs. mixed chimerism was observed in / patients ( . %) using qrt-pcr and in of the patients ( . %) evaluable with standard methods. overall, / patients ( . %) relapsed; before relapse, mixed chimerism was observed in all patients by qrt-pcr and in / by fish/vntr. qrt-pcr detected mixed chimerism days (range - ) earlier than standard methods. in cases in which vntr was either not informative or not predictive for relapse, the interval between detection of mixed chimerism by qrt-pcr and relapse was and days, respectively. conclusions: chimerism determination using qrt-pcr is more informative than standard methods and may represent an useful tool for the follow up of allogeneic sct. reduced intensity transplant programme. a single-centre experience a. bonini, a. tieghi, f. merli, l. gugliotta asmn (reggio emilia, i) from we introduced a reduced-intensity conditioning regimen for allogeneic transplants(allo). this regimen is tailored for old patients (pts) or for those who have previously received an allo or autologous bmt. at now we have transplanted pts; of them received previously an autologous and an allo bmt. they were males and females and the donors were males and females. the mean age of the pts was yrs (range - yrs).the diagnoses were: hd, nhl, mds, renal cancer, sarcoma, myeloma and cml; the solid tumors were all metastatic, pts were in pr, in ii cr, in iii cr and the one with mds at the onset. the abo compatibility was:complete for , minor for and major for . the cmv status was: donor/pt positive for and positive donor/negative pt for . the conditioning regimen consisted of thiotepa, fludarabine and cyclophosphamide except for the pt with cml who received busulphan and fludarabine. the source of stem cells was peripheral blood in and bone marrow in and the stem cells were cryopreserved after the harvest. the gvhd prophylaxis consisted of csa and short course mtx. the pts who received peripheral stem cells the mean cell dose was x /kg cd + cells, for the one who received bone marrow was . x /kg mnc. the mean time to reach pmns > . x /l and plts > x /l was respectively days(range - days)and days (range - days). the mean number of transfused rbc and plt units was respectively (range - )and (range - ). the mean grade of mucositis (according to the who classification) was . the major complications during neutropenia were fuo, gram + bacteremias and cerebral aspergillosis. no cases of vod of the liver were observed. pts had agvhd( grade and grade ) and limited cgvhd. two pts developed cmv reactivation after allo; the complication was cured with gancyclovir. nine over pts died: for disease (the solid tumors but for the with renal cancer a response even if transient was observed and nhl), for acute gvhd and for cerebral aspergillosis. the chimerism was complete for all the evaluable pts at days. none needed dli. six patients are in cr with a median follow up of year ( were in partial remission and in ii chronic phase at transplant) and relapsed. this pilot study demonstrated an acceptable regimen-related toxicity(according to the age and previous chemotherapy including transplants), the possibility to reach a very early full-donor chimerism and to cure high risk pts. introduction: autologous stem cell transplantation (autosct) is usually preferred to allosct, due to its widespread availability, lack of the immunological problems intrinsic to the development of graft-versus-host disease (gvhd), and the infrequent bone marrow involvement present in hodgkin's disease(hd), for patients undergoing high-dose chemotherapy/radiotherapy. allosct has been associated with a high transplant-related mortality (trm) in patients with hd due to a high incidence of gvhd and of fatal infectious events after transplantation. the poor outcome of these patients after allosct may reflect in part the advanced status of the disease at transplantation and the poor performance status of the patient population allografted. furthermore, the high trm present in the conventional allosct setting has never allowed a proper evaluation of a possible graft-versus-hodgkin's effect. in an effort to reduce the trm associated with allosct, low-intensity regimens have been developed; the curative potential of these protocols would rely on the graft-versus-leukemia effect of the allogeneic infusion more than in the conditioning regimen per se. case: in our hospital a total of patients with relapsed hodgkin's disease underwent reduced-intensity conditioning (ric) allogeneic stem cell transplantation (allo-sct) from an hla-identical sibling. we explored reduced-intensity allografts using fludarabine-melphalan conditioning and early withdrawal of immunosuppression as an alternative to palliative chemotherapy. graft-versus-host disease (gvhd) prophylaxis was mini-methotrexate and ciclosporine with weaning at day . the age of patients was , and years. the time from initial diagnosis was , and months and from autograft was , and months. the patients were in refractory relapse. time to neutrophil recovery (absolute neutrophil count ≥ /microl) was days for patients and days for the other. time to platelet recovery ( platelet count ≥ /microl) was , and days.the -day mortality was . none of them developed acute gvhd. one patient developed mild chronic gvhd. as of november patients remain in complete remission. conclusion: although the number of hd patients allografted with reduced-intensity protocols is low and the follow-up still short, it seems that the reduced-intensity allogeneic stem cell transplantation is effective in relapsed and refractory hodgkin's disease where autografts have failed. remarkable reduction of acute gvhd and infectious complications after reduced-intensity conditioning and low-dose ( mg) campath- h p. reményi, Á. bátai, b. kapás, a. sipos, s. lueff, i. bodó, m. réti, v. goda, t. masszi national medical centre (budapest,hun) during the four month-period from may, to september, we performed reduced intensity (ric) allogeneic transplantation for five patients with their hla indentical sibling donors. the median age of the female and male patients was . years ( . - . ). two patients had chronic lymphocytic leukemia (cll), one hodgkin's disease (hd), one follicular nhl grade i (fl) and one myelodysplastic syndrome (mds). the hd patient relapsed after previous autologous stem cell transplant. the fl patient was in complete remission, the others were in partial remission before transplant with low tumour burden. the conditioning regimen consisted of x mg campath- h and fludarabine x mg/m² for all patients, adding melphalan ( mg/m²) for the lymphoid malignacies, and busulphan ( mg/kg) for the mds patient. for gvhd prophylaxis mg/kg cyclosporin a (continuously) + mg/m² methotrexate was given on days , , . all patients engrafted. one patient developed grade i acute gvhd. two patients had febrile neutropenia, one developed central venous line infection. no one had cmv reactivation or disease. after a median of day-follow up ( - days) all patients developed full donor chimera tested by vntr pcr. the two cll patients are in remission proven by flow cytometry (less than % cd /cd + cells). the fl patient is in cr and received four courses of mabhtera ( mg/m²) as maintenance therapy. the mds patient is in cr according to bm histology, but still thrombopenic ( g/l). the hodgkin's patient has active disease with minimal tumor burden with mixed chimerism at day , now waiting for dli. conclusion: low dose ( mg) campath- h+ csa/mtx gvhd prophylaxis is a well balanced regimen regarding the incidence and severity of acute gvhd, infectious complications and gvl effect after ric conditioning. these preliminary results -especially concerning the late infections complications -compare much better to those we observed in our previous series of ric transplants with higher doses ( or mg) of campath- h, or that we read in the literature. with the publication of these preliminary results we would like to underline the message that less is better regarding campath- h in reduced intensity conditioning. conditioning regimens consisted of fludarabine mg/m²/day x days plus melphalan mg/m²/day x day (n= ) or plus oral busulphan mg/kg/day x days (n= ) or plus cyclophosphamide mg/kg/day x days and globuline antithymocyte . mg/kg/day x days (n= ). the patients were grafted with bone marrow (n= ), cord blood (n= ) or pbsc either unmanipulated (n= ) or cd + selection (n= ) or cd /cd depletion (n= ). gvhd prophylaxis was performed with csa+ mtx (n= ), csa only (n= ) and csa + steroids (n= ). donors were either related (n= ) or unrelated (n= ). the median number of cd + cells infused was . x /kg recipient bw (range . - . ). results: there were a rapid recovery of neutrophils (median days; range - ) and platelets (median days; range - ). the median length of hospital stay was days ( range - ). with a median follow-up of months (range - ) the incidence of agvhd and cgvhd were ± % and ± % respectively. the probability of trm was ± %. patients grafted with manipulated pbsc had the lowest trm ( ± %). the relapse incidence was ± %. high number of infused cd + cells (p= . ) and cgvhd (p= . ) was associated with a lesser ri. the event-free survival was ± %. nine patients died of relapse or progressive disease (n= ), agvhd (n= ), cgvhd (n= ) and other (n= ). conclusion: fludarabine-based ric provide a good alternative to myeloablative conditioning for allogeneic transplantation either malignant or non-malignant disease in children. induction: treatment related mortality is the trade off of allogeneic transplants. although the probability at two years has been reduced to - % following non-myeloablative (reduced intensity conditioning) allogeneic transplant it remains a barrier to administer these transplants. we desired to identify mechanisms that compromise safety or would enhance safety. methods: the different outcome parameters in transplantation were defined as safety or efficacy parameter and per safety and efficacy parameter the literature was searched for investigational method, prophylactic, supportive or therapeutic measurement. furthermore the toxicity scales were reviewed and known side effects were listed and include in the search. results: outcome parameters defined as safety parameter included engraftment, disease recurrence; side effects of major concern, acute and chronic graft versus host disease, infections, death as well as life threatening side effects defined by the safety scales. more that first complete remission was considered and adverse safety parameter. ) engraftment failure was noted after inadequate cytoreductive treatment such as cgy tbi and cyclophosphamide or underdosing of fludarabine (< mg/m²); adequate dosing of cytoreductive and immune suppressive treatment appeared critical for rapid trilineage engraftment. ) tbi appeared associated with higher risk of graft versus host disease and conditioning with drugs only. ) in vivo t-cell depletion by f.e anti-thymocyte globulin was associated with high risk of cytomegali virus reactivation. ) t-cell depletion of the graft is associated with increases risk of relapse ) b-cell depletion of the graft seems to reduce the risk of chronic graft versus host disease ) cmv positivity is not a prerequisite for cmv reactivation ) a combination of immune suppressive agent reduces the risk of acute graft versus host disease discussion: based on these and less stringent criteria we defined a practice guideline and make recommendations for reduction of side effects and treatment related mortality. the method developed model will be presented. monitoring of mixed chimerism by single nucleotide polymorphism analysis in two children supported with granulocyte transfusions after allogeneic stem cell transplantation w. schwinger, p. sovinz, h. lackner, h. dornbusch, m. benesch, a. moser, g. lanzer, c. urban medical university graz (graz, a) objectives: neutropenia is a major risk factor for early transplant related mortality in children undergoing haematopoietic stem cell transplantation (hsct). in case of infection, refractory to antibiotic therapy the combination with allogeneic granulocyte transfusions is a logical approach to manage this problem. these patients are chimeras of at least three different cell populations (recipient -stem cell donorgranulocyte donor). two patients where followed closely by single nucleotide polymorphism (snp) analysis to reveal the duration and percentage of leukocytes derived from granulocyte donors. patients and methods: one patient suffering from evans syndrome and was transplanted with cord blood of a mmud after myeloablative conditioning with busulfan, thiotepa, etoposide and atg. the child received one neutrophil transfusion at day + . the second patient was diagnosed with aml (fab m ) and was transplanted in persistent bm-aplasia after induction chemotherapy with bm of her hla-identical mother after reduced intensity conditioning (ric) with fludarabine, melphalan and atg. this child was transfused with eight granulocyte transfusions of three different allogeneic donors starting at day - until day + with two three days intervals. snp analysis was performed the day after each granulocyte transfusion and once weekly until stable allogeneic engraftment was achieved. results: both patients showed allogeneic engraftment of > % donor chimerism at day and respectively. third party leukocyte dna could be detected from day + until day + in the first patient and from day - until day + in the second patient. in the second child up to five genetically different leucocyte dna populations where detectable and could be quantified. conclusions: granulocyte transfusions of third party donors after allogeneic stem cell transplantation are increasingly used. snp analysis allows precise monitoring of donor chimeras even in case of two or more donor cell populations after transplantation. this could be important especially in case of ric transplantations in which precise follow up of engraftment kinetics is very important. patients are alive and without disease. one year overall survival and disease free survival (dfs) is % ( - %) and % ( - %), respectively. transplant related mortality (trm) was the cause of death in patients, months and one year cumulative incidence (ci) of trm were % ( - %) and % ( - %), respectively. eight patients relapsed (median time: days ( - ); one year ci % ( - %). twelve patients developed acute graft versus host disease (agvhd); ci % ( - %) fourteen of evaluable patients developed chronic gvhd; ci % ( - ). the development of cgvhd was associated with better dfs: % ( - %) vs. % ( - %) p= , . conclusion: ric allo-sct is a curative option in patients with advanced age diagnosed of amd/mds with an acceptable trm. age should not be an exclusion criteria in patients with aml/mds for allo-sct. cgvhd is associated with better dfs. finaced in part by g / , /xt . allogeneic stem cell transplant (allo hsct)is a curative approach for patients with hematologic malignancies but it is associated with high treatment related morbidity and mortality. because transplant related mortality increases with advanced age, advanced disease and unrelated donors, patients older than - years may be excluded from this procedure. reduced intensity conditioning and new preparative regimens are therefore explored to allow hsct to a wider patient population. the aim of this study was to evaluate efficacy and toxicity of the combination treosulfan (water soluble alkylating agent, busulphan derivative) and fludarabine as preparative regimen for allo-hsct in patients receiving match sibling or unrelated donors for advanced heavily pretreated hematologic malignancies. since july to november patients ( all, aml, cml, mm ) entered this study. mean age was years (range - ). conditioning consisted of treosulfan gr/m² for days, fludarabine mg/m² for days, cyclosporine plus short mtx and anti-thymocyte globulin (thymoglobulin) at a total dose of mg /kg. all patients engrafted; mean time to neutrophil recovery > x /l was (range - ) days , to platelets > x /l was ( range - ) days. no conditioning regimen related deaths was observed. three ( ) patients experienced gi toxicity ( grade , grade ), patients had grade liver toxicity. no acute gvhd was observed. all patients are alive with a follow-up ranging from to days. despite the short follow-up, in this preliminary report we underline that treosulfan-fludarabine -atg conditioning is characterized by reduced toxicity; long term follow-up is necessary to evaluate os, dfs and relapse in these patients. background: prognosis of patients with aml relapsing within a year of allografting is poor. management options are limited and response to donor lymphocyte infusion(dli) is poor due to disease kinetics. second allografting using conventional conditioning is unpopular due to high transplant related mrotality(trm).there is paucity of data on second allografting with reduced intensity conditioning(ric) as a salvage for relapses post-allografting. herein, we report such patients who were successfully salvaged and achieved a durable complete remission(cr) with ric and second allografting. case : a year old male with normo-cytogenetics aml received an allograft from a hla identical sibling after myeloablative conditioning. he relapsed months after transplant with loss of donor chimerism. he was salvaged with flag regime followed by serial dlis and achieved cr which lasted months before relapsing again. he was reinduced with flag regime followed by pbsc infusion from the original donor. he received no post-transplant immunosuppression with the development of grade graft versus host disease(gvhd). he has since achieved % donor chimerism and remains in cr for more than months. case : a year old male with myelofibrosis transforming into aml associated with del( q) received his first allograft from a hla identical sibling after non-myeloablative conditioning. donor chimerism declined after months and blast counts continued to rise despite serial dlis. he was reinduced with idarubicin and cytarabine, followed by a second allografting from the original donor with ric (fludarabine mg/m² d- to - , melphalan mg/m² d- to- ). full donor chimerism ensued and he remains in cr for more than months. immune suppression was tapered with development of grade ii gvhd. conclusion: remission durations after the second allografting exceeded those after the first. a ric regime with second allografting is a more effective modality than dli in the treatment of relapsed aml. a ric regime lessens the trm associated with second transplants, while allowing for engraftment of infused pbscs with execution of gvl effect. immune modulation by aggressive manipulation of posttransplant immune suppression also appears to be a key element in successful second allografting. this salvage approach offers a practical, well-tolerated and potentially curative treatment for patients who in most circumstances, would have been precluded from further active management. reduced intensity conditioning (ric) allogeneic stem cell transplantation (allo-sct) can reduce the frequency of transplant-related toxicities, at least in the early period after allo-sct. here, we analyzed the profile of platelets recovery and transfusion requirements in the first months after ric in a single institution series of consecutive patients receiving allo-sct from an hla-identical sibling. patients and graft characteristics were: age y. (range, - ), diagnoses: myeloid malignancies ( %), lymphoid malignancies ( %), and metastatic solid tumors ( %). pts ( %) received a fludarabine, busulfan and atg-based ric, while pts ( %) received a low dose irradiation-based ric. all patients received a pbsc graft. pts ( %) developed grade - acute gvhd. platelets recovery (> . /l) was observed at a median of days (range, - ). the kinetics profile of platelets recovery is shown in the figure below. in the whole study population, the nadir was observed around day + after allo-sct, and a plateau was reached by the end of the first month. in this series, patients needed a median of unit (range, - ) of filtered and irradiated donor apheresis platelets. of note, pts ( %) did not require any platelets transfusion during the follow-up period (median follow-up: days). among the patients ( %; %ci, - %) who received at least one transfusion of platelets, were not transfused beyond day + after allo-sct. when comparing these patients, to the group of patients who were never transfused, platelets count prior to ric allo-sct (median count . /l vs. . /l; p= . ) and the occurrence of severe acute gvhd (p= . ; % of patients with grade - acute gvhd were transfused) were the parameters significantly associated with platelets transfusion needs. in this cohort, pts could be assessed for platelets recovery at day + : among them, ( %) had a platelet count > . /l. at day + after allo-sct, a diagnosis of myeloid malignancy (aml, cml or mds) was associated with a delayed platelet recovery (p= . ). overall, these observations show a significantly lower rate of platelets transfusions and a quicker kinetics of platelets recovery after ric allo-sct. in addition, the low level of myeloablation observed after ric, may offer a window of opportunity for testing of megakaryocytic growth factors, towards further improving the safety and outcome of ric or nonmyeloablative allo-sct. fungal infections have become the major cause of infectious morbidity and mortality in patients undergoing bone marrow transplantation (bmt). in many cases invasive aspergillosis infections create a major therapeutic dilemma and contraindication to marrow transplantation. we report on a years old boy with secondary acute myeloid leukemia, who underwent unrelated donor peripheral blood stem cell transplantation (pbsct) with previously diagnosed pulmonary aspergillosis and successfully recovered from the infection. probable invasive pulmonary aspergillosis (ipa) was diagnosed in the patient acc. to eortc-criteria. a large diffuse wedge-shaped infiltration was observed in thorax ct scans two months before pbsct and throughout the early posttransplant period. liposomal amphotericin b (l amb) mg/kg/d i.v. and voriconazole p.o. x - mg/kg/d were administered for the whole peritransplant period. after conditioning regimen incl. treosulfan x g/m², melphalan mg/m², atg fresenius x mg/kg the patient received pbsc ( . x cells cd +/kg recipient bw) from the unrelated donor, who was mismatched at a*-allel. csa, mtx and atg were used as gvhd prophylaxis. a rapid and sustained allogeneic engraftment (neutrophils > . g/l on day + , thrombocytes > g/l on day + ) was observed. the posttransplant period was uneventful except for weeks long lasting exhausting morning cough and subtle breathing difficulties requiring passive oxygen therapy. three weeks after pbsct, bronchoscopy with broncho-alveolar lavage revealed no pathogens. nevertheless it was decided to continue treatment with l amb mg/kg every second day. in control thorax ct the infiltration was considerably smaller and no indication was found to perform an open lung biopsy. the patient received further treatment with voriconazole p.o. for months. regularly performed thorax cts revealed a continuous regression of pulmonary changes. the patient remains alive and well in cr months from transplant without any pulmonary abnormalities, except for a slight thickening of the interlobar groove and is given itraconazole p.o. this report demonstrates that administration of full-dose antifungal therapy and shortening the neutropenia period due to peripheral blood stem cell transplantation allow the successful outcome, even in high-risk patients with previous aspergillosis. post-transplant pulmonary complications are not rare and their mortality is still very high. we present -old year girl with high-risk acute lymphoblasic leukemia after allogenic hematopoetic cell transplantation (hct) from matched related sibling donor. conditioning regimen consisted of fractioned total body irradiation ( gy) and high-dose etoposid ( mg/kg). as graft versus host disease (gvhd) prophylaxis cyclosporine a was used. graft contained , x /kg cd + cells. on day + persistent fever occurred, antibacterial and antifungal treatment were administrated. on day + ug/kg g-csf was added. we observed symptoms of respiratory failure. on day + wbc was /ul, cutaneous gvhd grade iii appeared and steroids were administrated. on day + she has all symptoms of pulmonary oedema, was intubated and mechanically ventilated. simultaneously rapid hematopoesis reconstitution was observed: leukocytes> . g/l and granulocytes> . g/l on day + , and platelets> g/l on day + . after few days of the gradual improvement, her status suddenly deteriorated. chest x-ray showed fluid in the alveolar space in both lungs. pentamidine was added to treatment. severe but stabile status (opportunity to controlled ventilation, fio above %) lasted about months. she was treated with wide-spectrum antibiotics and antifungal drugs (liposomal amphotericin, voriconazole) although her blood cultures were still negative. in this time we also treated her with anti-tnf-alfa antibodies. because of probable invasive aspergillosis and candidiasis we introduced treatment with caspofungin. gradual improvement was observed: significant decrease of the requirement for oxygen, possibility to reduction of controlled pressures and respiratory rates and simultaneous improvement of radiologic changes. after few days of continuous positive airway pressure ventilation right pneumothorax appeared and continuous suction drainage had been used for three weeks. simultaneously she was detoxicated from thiopental and morfine using fenobarbital and methadone. on day + mechanical ventilation was discontinued. requirement for passive oxygen therapy gradually decreased. nowadays chest tomography shows mild pulmonary fibrosis and bronchiectasia. the patients is alive, in good general status, under intensive physical and s pulmonary rehabilitation with stable full donor chimerism without immunosupression. nevertheless regular long-term pulmonary follow-up is still required. cns symptoms and their severity strongly correlate with outcome of patients with familial haemophagocytic lymphohistiocytosis (fhl). here we describe a case of a patient without primary cns involvement related to fhl who developed progressive cns damage of unknown aetiology after allogeneic stem cell transplantation (sct). the patient was diagnosed at the age of . years with fhl, mutation in perforin gene was not proved, no signs of reduced perforin expression were observed. he was treated according to protocol hlh and then transplanted in clinical and haematological remission of fhl from his haploidentical father (peripheral blood stem cell -cd positive selection; clinimacs) because no matched donor was available. myeloablative conditioning consisted of busulfan, cyclophosphamide and ratg. soon after engraftment he suffered from cmv reactivation and he consequently experienced acute graft rejection. following okt and steroids further t cell depleted graft sct was infused days after st sct. early after second engraftment he developed severe acute encephalopathy and syndrome of inadequate adh secretion (hyponatremia, hypothermia, neurologic seizures,…), hhv variant b was at that time detected in cerebrospinal fluid (csf) and blood. this was early followed by marked ebv and b cell proliferation treated with rituximab. after engraftment, complete donor haematopoiesis was confirmed with exception of almost full autologous recovery of t lymhocytes (split chimaerism). at day+ all t lymphocytes ( , . /l) were activated (expressing hla dr) and were of host origin. eleven months after sct hhv was repeatedly detected in peripheral blood and later frequent pharmacologically almost uncontrolled epileptic seizures started initiating progressive mental retardation. throughout this whole period repeated mri scans and examinations of csf, blood and marrow failed to document involvement of cns due to uncontrolled fhl. one year after sct cytototoxic assays showed significantly decreased activity of t cells. despite the number of donor origin t cells started to predominate the host ones only years after sct, there were no clear clinical signs of fhl. unfortunately the patient developed irreversible cns damage. we speculate that multiple herpetic infections were responsible for proliferation of activated autologous t lymphocytes that contributed to severe cns damage in this patient. supported by grants of mh cr no. , and consecutive twenty seven patients with inborn errors: four inborn metabolism: three metachromatic leukodystrophy, and one mucopolysacaridosis type : hurler´s syndrome; also one osteopetrosis, two mayor thalassemia, two fanconi anemia, and eighteen inmunodeficiencies: twelve severe-combined immunodeficiency (scid), two wiskott-aldrich syndrome and four hemophagocytic lymphohistiocytosis, were transplanted in our centre during the last nine years. we report two cases of scid: scid type jak deficiency (t-, nk-, b+) and major histocompatibility complex (mhc) ii deficiency which were treated with double haploidentical parental donor positively selected hla-mistmached cd + progenitor cells were isolated from peripheral stem cells, after mobilization with granulocyte colony-stimulating factor (g-csf) at standard dose, by the isolex (baxter)and clinimacs(miltenyi)systems selection devices respectively. the first case was a male, . months years old with jak deficiency (t-, nk-, b+) who was undergone to transplant without conditioning regimen; value of peripheral cd + cells infused was . x /kg and the mean cd + cells number was . x /kg; and . log t-cell depletion. he had graft rejection at day + and therefore was undergone to second haploidentical from the same family donor: father, this time with no-myeloablative conditioning with fludarabine (f) + melphalan + anti-thymocyte globuline (atg) and prophylaxis for graft-versus-host disease (gvhd) with cyclosporine (cya), he died ten days after the second transplant by heart insufficiency and metabolic failure. the second patient was a female, months years old with mhc class ii deficiency, she was undergone to haploidentical transplant with myeloablative conditioning based f+ busulfan and ciclophosphamide and atg. the value of peripheral cd + cells infused was . x /kg and the mean cd + cells number was . x /kg, and . log t cell depletion; she had graft rejection at day + and mixed chimerism therefore was underwent to second haploidentical transplant from the same parental donor: mother, the conditioning regimen was based okt- and dexametasone, she died five days after the second transplant by lung bleeding. despite poor prognosis at diagnosis of these cases, and the engraftment failure regardless mega-doses of cd +; this approach could be a feasible therapeutic option for patients lacking a suitable donor. introduction: mobilized peripheral blood stem cells (pbsc) represent the most important source for autologous stem cell transplantation, even in children with malignancy. the current practice is administration of g-csf alone or in combination with chemotherapy. recently, a polyethylene glycol (peg)conjugated form of g-csf (pegfilgrastim) has been licensed. preliminary data indicate it has the same effects of filgrastim in terms of elevation of absolute neutrophil count, mobilization of pbsc, and reduction of duration of chemotherapy-induced neutropenia, with the obvious advantage that these effects could be sustained for several days from a single injection without added toxicity. in a recent experience the efficacy of a single dose ( microgr) of pegfilgrastim, in combination with salvage chemotherapy, was tested in an open-label phase ii study of pretreated patients. the authors concluded that pegfilgrastim as an adjunct to chemotherapy is a predictable and highly effective mobilization regimen in pretreated lymphoma patients (isidorins ). very limited experience is available on the use of pegfilgastrim in children. in the only two available report it was used to shorten the duration of severe neutropenia after cytotoxic chemotherapy in five children with ewing sarcoma (te poele em ) and in seven pediatric cancer patients (wendelin g ). we are not aware of any report on the use of pegfilgrastim for mobilization in children. patients: we treated children, males, aged , and years, affected of rhabdomyosarcoma, ewing sarcoma, and b-nhl. all received chemotherapy with vp and edx and than a single subcutaneous dose ( microgr) of pegfilgrastim. two patients had a good response, with the peak cd + count ( and ) on day + and + , while the failed to mobilize and to collect a sufficient number of cd + cells even after conventional g-csf and bone marrow harvests. the only patient who so far has completed the treatment program underwent, according to the treatment protocol, autologous transplant repeated three times and engrafted (pmn > /microl) on day + , + , and + . conclusion: mobilization with peg-filgastrim was safe and efficient in our patients; failure to mobilize was observed only in an heavily pre-treated patient with ewing sarcoma. stem cell transplantation (sct) is the treatment of choice for patients suffering from severe aplastic anaemia (saa) who do not respond to immunosuppressive treatment (ist). patients transplanted from an hla-identical sibling have an excellent prognosis compared to patients who were grafted from alternative donors. t-cell depletion by cd positive selection of peripheral stem cells reduces the risk of gvhd considerable. this, however is followed by an increased risk of graft rejection. most recently, it could be shown, that changing the graft processing from cd positive enrichment to depletion of cd and cd positive peripheral cells does facilitate and improve engraftment in children transplanted from hla-haploidentical parents. consequently, also patients with saa who are at highest risk for graft rejection might benefit from such a new graft processing technique. a -year-old girl who failed to respond to ist received an allograft from mud. the conditioning regimen consisted of fludarabin ( x mg/m²), thiotepa ( x mg/kg), melphalan ( x mg/m²) and okt ( x . mg/kg). g-csf mobilized peripheral blood stem cells (pbsc) were purified using anti-cd /cd microbeads (miltenyi). recovery of cd + progenitor, cd + nk cells and cd + monocytes was more than % each. residual t-and b-cells were < . and . %, respectively. in total, . x cd + cells; . x cd + cells; . x cd + cells and . x cd + cells per kg were administered. without g-csf, engraftment occurred on day + for leucocytes and both neutrophils and platelets on day + . acute toxicity was mild (grade i). post transplant immunosuppression was performed using mmf. gvhd grad i was observed, which responded to prednisolone. noteworthy, although t-cells were severely depleted, t-cell regeneration was fast with more than /µl cd and cd positive cells detectable already on day + . chimerism analysis showed complete donor chimerism. at day , good immune recovery with /µl cd +, /µl cd +, /µl cd + and /µl cd + cells was detected. in conclusion, cd /cd depleted peripheral stem cells from a mud in combination with a reduced intensity conditioning regimen could be a promising option in the treatment of patients with aplastic anaemia not responding to immunosuppressive treatment and lacking a matched sibling donor. idiopathic myelofibrosis (imf) comprises myelofibrosis, extramedullary haematopoiesis, hepatosplenomegaly and pancytopenia. in adults imf represents a poor prognosis, progressive fibrosis and leukaemic transformation are frequent. allogeneic hematopoietic stem cell transplantation (hsct) is a treatment option but is connected with high risk of graft failure and toxicity. in children the disease is rare and variable, stable course or spontaneous remission has been reported. we describe two cases of imf in children. in a girl mild anaemia and thrombocytopenia were first documented at the age of years. at years pancytopenia was found, trephine bone marrow (bm) biopsy revealed normocellular haematopoiesis with myelofibrosis. she remained in a good clinical state with a stable blood count but further bm biopsies showed decreased cellularity with myelofibrosis. . years after the diagnosis her blood count dropped off, hepatosplenomegaly was noted and bm biopsy revealed marked myelofibrosis. months later at the age of years hsct was therefore performed (matched unrelated donor, flu+mel+ratg). anc engrafted on day , platelets on day , complete donor chimaerism achieved on day . corticosteroids started on day for mild extensive gvhd. bm biopsy at day remained hypocellular with myelofibrosis, however blood count was normal. -year-old male presented with pallor, quickly developed pancytopenia with blasts. bm biopsy showed myelofibrosis, increased blasts, trisomy + . hsct from a hla identical sibling was performed months later (flu+mel+ratg), anc engrafted on day , platelets on day . complete donor chimaerism was achieved on day , no gvhd. at day bm biopsy did not display myelofibrosis, peripheral blood count was stable. clinical deterioration started months post-hsct with fever, weight loss, hepatosplenomegaly, mixed chimaerism, thrombocytopenia, blasts and extramedullary infiltration of breast. trephine and breast lump biopsy confirmed relapse of imf in transformation to aml-m with trisomy + and trisomy + . second hsct from the same donor was carried out months after the first hsct (bu+cy+mel). gvhd grade ii treated with steroids manifested on day . anc engrafted on day , platelets not engrafted. despite artificial ventilation diffuse alveolar haemorrhage resulted in death on day . reduced intensity conditioning composed of fludarabine and melphalan is preparative regimen of choice for hsct in children with imf. there are no studies on the connection between graft versus host disease (gvhd) and angiogenesis. however, chen et al have shown in nat med ( ) that the vascular endothelial growth factor-c (vegf-c) -vascular endothelial growth factor receptor- (vegfr- ) -axis has an effect on alloimmunity, and that the blockade of vegfr- signaling is immunosuppressive. both vegf-c and angiopoietin (ang ) are important in angiogenesis and lymphangiogenesis. in this study we measured the levels of these two factors in children after urd-sct (unrelated allogeneic stem cell transplantation). patients and methods: nine patients aged - yrs were included, six of whom developed significant acute gvhd (agvhd, gr ≥ ) while three did not. the diagnoses of the agvhd patients were all (acute lymphoblastic leukemia) (n= ) and saa (severe aplastic anemia) (n= ). the non-gvhd patients also had all (n= ) and saa (n= ). gvhd prophylaxis consisted of cyclosporin and short methotrexate. the serum concentrations of vegf-c and ang were analyzed by elisa at - days posttransplant, - samples/patient. results: the vegf-c and ang concentrations (median, range) were ( - ) pg/ml and ( - ) pg/ml, respectively. the presence or absence of agvhd did not make any difference in the levels. neither did we find correlation between hemoglobin, white blood cell count, bilirubin, crp or sedimentation rate and these two angiogenic factors. the vegf-c levels were significantly lower than in our previous study on all patients at diagnosis. the individual maximal ang levels correlated with survival (≥ mo follow-up, p= . ). both absolute lymphocyte count and platelet count correlated with vegf-c levels, probably because these cells are known to produce vegf-c. conclusion: our results did not support the hypothesis about correlation of the levels of angiogenic factors with gvhd. instead, there was a novel finding about low concentration of ang being predictive of a good outcome. our findings pose important questions on the emerging role of angiogenic factors in the evaluation of the pathogenesis of gvhd. background: hematopoetic stem cell transplantation (hct) does appear to be a therapeutic option for children and adolescents suffering from shwachman-diamond syndrome with severe cytopenias and/or myelodysplastic syndrome. the paucity of experience with children undergoing hct for sds has been the major obstacle for recommendations regarding time point, transplant regimen, and patient subgroup benefiting most from hct. most but three reported patients received a preparative regimen either consisting of bu/cy or bu/tbi. however, severe early toxicity with cardiomegaly, myocardial fibrosis, and cyclophosphamide associated cardiomyopathy have been described. therefore, we tested the feasibility of a cyclophosphamide free protocol using fludarabine, treosulfan, and melphalan as a conditioning regimen. methods: between and two children with sds were enrolled. age at transplantation was and years. both patients received conditioning with fludarabine ( mg/m²/day x ), treosulfan ( g/m²/d x ), melphalan ( mg/m²/d x ), and campath- h. all children received a non manipulated fresh bone marrow graft. the first patient from a hla-identical sibling, the second from a / locus matched unrelated donor. mean cell doses transplanted were . x nucleated cells/kg bw ( . x /kg and . x /kg). results: both patients achieved donor derived engraftment, no gvhd exceeding grade ii was observed, and both maintained donor chimerism at %. all patients developed grade iii mucositis. on patient experienced a cerebral seizure early after transplant most likely caused by csa toxicity. gvhd prophylaxis was switched to mmf and the patient fully recovered from this single event. apart from this, no rrt > grade ii was seen. all patients are alive after a follow up of and month. conclusion: a fludarabine based, cyclophosphamide free conditioning regimen seems to be a feasible approach for matched sibling and matched unrelated hct in children and adolescents with sds. larger numbers and a longer follow-up are needed to make these results more comparable to traditionally used preparative regimens. a prospective comparison of immune reconstitution after autologous haematopoietic stem cell transplantation in children v. wiegering, b. winkler, m. eyrich, p.-g. schlegel university of wuerzburg (wuerzburg, d) introduction: autologous haematopoietic stem cell transplantation (auto hsct) has become an established therapy for numerous advanced paediatric solid tumours. after haematopoietic stem cell transplantation all recipients experience a period of immunodeficiency. regeneration of adequate t-cell numbers and repertoire diversity are key elements in the recovery of immune competence. patients: immune reconstitution was studied in children ( transplants; median , years; range m- y; female, male). blood samples were drawn before auto hsct, on days (take), , , , and and > months. methods: we analyzed lymphocyte subpopulations using flow cytometry. intracellular cytokines (ifngamma, il , tnfalpha, il , il , and il ) were determined by facs after in vitro stimulation with pma, ionomycin and brefeldin for h. additionally, we measured il , il and tgfbeta in unstimulated sera by elisa. additional we measured trec´s and spectatypes. results: as to lymphocyte subpopulations after auto hsct, nk-cells were the first to regenerate. in t-cells, an inverted cd -cd -ratio could be detected during the first year. in cd + t-cells the memory phenotype (cd ro+) predominated. as to cytokine levels in unstimulated sera, we saw high levels of il shortly after transplantation, levels decreased to pretransplant values during one year. tgfbeta levels increased during the first year and decreased thereafter. ifngamma levels remained stable. in stimulated t-cells, ifngamma and tnfalpha increased in the first year and went down afterwards. interestingly, high ifngamma levels after transplantation correlated significantly with a better survival. non-irradiation containing conditioning regimen for children with fanconi's anaemia m. jarrar, m. sarhan, m. milhem, f. abdel-rahman, r. rihani, s. sharma, i. na'em king hussein cancer center (al-jubeiha, jor) fanconi anemia is an inherited disorder that leads to progressive bone marrow failure. the only curative treatment of the severe aplastic anemia that ultimately develops in these patients is allogeneic stem cell transplantation. patients with fanconi anemia have increased chromosomal fragility. as a result they are prone to both short and long term complications when conditioning regimens containing radiation are used. we report on patients ( males and females) with fanconi anemia who were transplanted from matched siblings without using radiation as a part of conditioning. median age at time of transplant was years ( - years). none of the patients had mds changes or leukemia prior to transplant. conditioning regimen consisted of fludarabine mg/kg, cyclophosphamide mg/kg and rabbit atg mg/kg. peripheral blood was the source of stem cells in patients, while bone marrow was the source in patients. gvhd prophylaxis consisted only of cyclosporine. at a median follow up time of days ( - days), patients are alive with normal hematopoesis. one patient failed to engraft. he was transplanted again with a different conditioning; however he had late rejection and died of sepsis days after second transplant. median cd + cell/kg infused was . million ( . - . million). median time to neutrophil and platelet engraftment was days and days, respectively. two patients had fever with atg; one patient had bacteremia during the neutropenic period. two patients developed fungal infections after engraftment. two patients had mild vod. three patients received vod prophylaxis consisting of urisidiol and spironolactone. cmv reactivation occurred in patients and was treated with gancyclovir. grade - skin and liver acute gvhd occurred in patients. limited chronic gvhd occurred in patients. one patient developed extensive chronic gvhd. conditioning without radiation is well tolerated in fanconi anemia patients and results in prompt engraftment. infectious complications appear to be high. autologous peripheral blood stem cell transplantation in low weight paediatric patients: methods and clinical outcome a. galmes, m. canaro, m. guibelalde, m. morey, l. bibiloni, a. vila, a. gutierrez, j. besalduch son dureta hospital (palma de mallorca, e) peripheral blood stem cell (pbsc) collection may be difficult in low weight body pediatric patients, with technical and clinical problems related to vascular access, low total blood volume, citrate toxicity, high extracorporeal volume, and patient's tolerance. methods: we present our experience with consecutive children weighing ≤ kg, diagnosed with acute leukaemia ( ) and solid tumors ( ), which were collected and transplanted between september and february at our centre. patients mean body weight was kg (range - ); median age was years (range, - years) ( table ).harvesting of pbsc was started after days of cytokine alone (g-csf mcg/kg/ hs. s.c.). procedures were performed using a baxter cs- plus separator primed with a mixture of irradiated and white cell-depleted red cells resuspended in % albumin and diluted with saline to match the patient's haematocrit. heparin and acd-a was used for anticoagulation (heparin ui in ml acd-a) in patients weighing < kg. the median number of leukapheresis was (range - ), processing . volemia in each session. the platelet count decreased significantly after each procedure without requirement of platelet transfusions. special monitoring of toxicity was done. the children were no sedated and showed no serious side-effects. all pbsc were cryopreserved with dmso % and stored at - ºc in mechanical freezer. results: the median time from cryopreservation to transplantation was . ( - ) days, and the median number of infused mononuclear cells and cd + cells were . ( . - . ) x /kg and . ( . - . ) x /kg, respectively. the median number of infused post-thawing cfu-gm was . ( . - . ) x /kg. all patients showed a safe and sustained engraftment. median time to reach and neutrophil per microl was ( - ) and ( - ) days. median time to and platelets level per litter was ( - ) and ( - ) days (table ) . long-term hematopoietic recovery at , and months was achieved in all cases (table ) . conclusion: our experience shows that our pbsc collection and cryopreservation method is a safe and efficient procedure in children weighing less than kg., with sustained haematopoietic reconstitution. an approach to retrospective validation and performance monitoring of pbsc collection and other white cell procedures using the cobe spectra cell separator: "cells collected" as a function of "cells processed" by different methods k.w. douglas, j. sinclair, m. mcgarvey, a. mcphelim, s. taylor, m. drummond s.n.b.t.s. clinical apheresis unit (glasgow, uk) validation of cell separator machines for a specific apheresis procedure poses a number of challenges. in particular, for white cell collection procedures it is not feasible for individual centres to carry out a prospective validation process prior to introduction of the procedure, because there is no way of performing a "dummy run" of a white cell procedure without actually putting a donor on the machine. we therefore attempted retrospective validation of our cobe spectra cell separator machines for pbsc collection using the mononuclear cell (mnc) procedure by estimating efficiency of the mnc procedure for each of the individual machines, in terms of the total mononuclear cell dose achieved in the apheresis product as a function of the number of mononuclear cells processed by the machine. there should be a direct correlation between total mononuclear cells processed by the machine and the final mnc dose in the product. the difficulty is in estimating "total mononuclear cells processed by the machine", which will only ever be an approximation. we estimated "cells processed" using different methods: method : the donor's peripheral mononuclear cell count (lymphocytes plus monocytes) multiplied by the run time; method : the number of total blood volumes processed multiplied by the donor's peripheral mononuclear count. retrospective audit was performed on pbsc collections carried out on five spectra machines over an -month period. total mononuclear cell dose in the apheresis product was graphed as a function of "cells processed" as calculated by both methods above. both methods showed a clear, statistically significant linear correlation, but there was less scatter and a lower p value using method (r= . , p< . ). it was noted that the gradient of the trend line is a measurement of the efficiency of pbsc collection. the data was therefore subdivided depending on which of the spectra machines had been used for collection, and individual data analysis was performed for each machine. this showed that the five machines all performed the mnc procedure with very similar efficiency (gradients of trendlines . to . ). the same process can easily be applied to ongoing performance monitoring of the five machines, and our aim will be to do this annually from now on. m. miorin, e. brunetta, e. scquizzato, s. varotto, c. messina, s. cesaro, g. binotto, i. baesso, e. calore, m. facco, e. ave, m.v. gazzola, r. destro, g. semenzato, r. zambello, l. trentin university of padua (padua, i) allogeneic bone marrow transplantation (abmt) is one of the powerful therapies for several hematological malignancies. multiple mechanisms contribute to the graft success, such as the entity of primary disease, donor bone marrow availability, minimal residual disease (mrd) and complications including infections and acute graft-versus-host disease (gvhd). gvhd represents a major complication of abmt and is the main cause of morbidity and mortality. the present study takes into account the recovery of the t cell-compartment before and after abmt in pediatric patients affected by different hematological malignancies. to evaluate the pattern of accumulation of t cells, we investigated the usage of t cell receptor (tcr-beta) chain variable regions (tcrbv) and the complementarity-determining region (cdr ) up to year follow-up after abmt. increased expression of some tcrvb families were observed in patients during the months after abmt. in the following months after abmt, we confirmed the presence of the same t cell clones and, sometimes, we showed the appearance of a new tcrbv family subset. we observed a random distribution of overexpressed tcrbv families and we did not show a preferential expression of a peculiar tcrbv. in order to clarify whether cells expressing a tcrbv region were clonally expanded, we performed cdr spectratyping and sequencing analysis. a predominant polyclonal pattern was observed in donors and patients before transplantation while at and months after bmt some clonal subsets were identified. in the majority of patients the presence of these subsets persists until month after abmt. a skewed tcrbv repertoire and oligoclonal/monoclonal subsets we observed may explain the post-bmt immunodeficency detectable after transplantation and may reflect responses to pathogens or alloantigens in correlation with clinical gvdh. this study was supported by a grant from fondazione città della speranza. s. ganepola, k. rieger, c. loddenkemper, j. maul, a. muessig, e. berg, h. stein, e. thiel, r. duchmann, l. uharek university medicine berlin, charite (berlin, d) introduction: tregs are involved in the control of immune responses to foreign antigens and play an important role in the pathophysiology of gvh-reactions. they are characterized by expression of cd +, cd + and the transcription factor foxp . although tregs are of emerging interest in allogeneic cell therapy, their precise enumeration in heterogeneous cell products has been extremely difficult. using monoclonal antibodies (moabs) against foxp and immunenzymatic labelling at the single-cell level in paraffin-embedded tissues, we have investigated different techniques to identify tregs in cellular products and tissue biopsies. methods: µl of the anti-human pe conjugated foxp antibody (clone pch , ebioscience) were used for intracellular labelling of x cells of cd depleted, macs sorted cd +/cd + cell fractions of peripheral blood mononuclear cells. the abcam goat polyclonal foxp antibody was used for double immunoenzymatic labelling of foxp /cd and foxp /cd of pbmc cytospins, of macssorted cd +cd + selected cell fractions and of paraffinembedded tissues. results: nuclear staining of the foxp and measurement by flow cytometry showed bright results in macs-sorted peripheral blood cells as well as in cytospins. gating on cd + cells of the cd depleted, cd enriched cell fraction, % of the cd + cells are positive for the foxp marker. the frequency of foxp + treg in the peripheral blood lymphocytes of healthy individuals ranged between - % of total lymphocytes, like previously described. in double labelling cytospin analyses of foxp and cd , we confirmed co-expression of cd on all foxp positive cells. counting high-power fields, % of the cd +/cd + cells are foxp +. in healthy individuals pbmc-cytospins we found % cd +/cd +/ foxp + cells. in double labelling analyses of foxp and cd in paraffin embedded tissues, we confirmed co-expression of cd on all foxp positive cells. only a few weakly cd -stained cells were negative for foxp staining, indicating a preferential staining of the cd high cell population. conclusion: direct staining of cellular products with moabs against nuclear foxp and subsequent flow cytometry can give similar results as nuclear foxp -staining in cytospin preparations or the assessment of cd +cd + cells by flow cytometry. these new techniques allow straightforward identification and quantification even of very low numbers of treg in peripheral blood subsets or other tissues. patient age and granulocytic contamination of apheretic harvests are important factors for adverse events after infusion of cryopreserved hsc g. milone, a. strano, s. mercurio, s. coppoletta, k. battiato, s. leotta, m. poidomani, r. giustolisi ospedale ferrarotto (catania, i) adverse events (ae) after cryopreserved cellular infusions are frequent and seldom can be life-threatening . dmso is considered important in their pathogenesis, however other factors could well play a role. we have prospectively collected data on ae presenting after hsc infusions following high dose chemotherapy performed in our centre during a y. period in patients affected with haematological neoplasm. stem cell source was pbsc in cases while bone marrow in . in all cases an endotoxin-free dmso was used. one or more ae was registered in / infusions ( . %). gastrointestinal complains were reported in % of all infusions, skin rashes in . % of cases, shaking in %, cough in . % , fever in . %, shortness of breath in . %, dizziness in %, headaches in . %. in univariate analysis patient age over was significantly associated to a higher incidence of ae, in fact incidence of ae was % below years of age, % in - decade and % over years (p= . ). frequency of ae was higher after pbsc than after bm ( % versus . %,chi-test: p= . ). in univariate logistic regression factors found important for ae in pbsc group were total number of cells infused /kg ( p= . ) and volume/kg of dmso infused (p= . ). adverse events were more frequent also when total number of granulocytic cells/ present in pbsc harvest was > . x /kg in respect to infusions containing a total number of granulocytic cells below this threshold ( % versus %, chi test: p= . ). all aforementioned factors were evaluated in multivariate logistic regression and age of patient (p= . ) and granulocytic contamination over . x /kg (p= . ) were still significant while the total volume of infused dmso loose importance (p= . ). no cardiovascular events were recorded during infusions, however we registered a statistically decrease of blood pressure and a statistically decrease of cardiac frequency. a significant correlation existed between reduction of cardiac frequency and volume/kg of dmso infused (r: . , p= . ). in conclusion while non cardiovascular ae are dependent from patient age and from granulocytic contamination of apheretic harvests, cardiovascular changes are influenced only by volume/kg of dmso infused. as far as non cardiovascular adverse events are concerned, particular attention should be paid in infusions of hsc in patients over the age of years and when the grafts have a granulocytic contamination over to . x /kg. haematopoietic stem cell transplantation for haematological malignancies: a -year single-centre experience b. georgievski, l. cevreska, n. siljanovski, a. stojanovic, o. karanfilski, z. stojanoski, s. genadieva stavrik, i. panovska, s. krstevska balkanov, a. pivkova clinical center (skopje,mk) hematopoietic stem cell transplantation (hsct) is widely used therapeutic method in the treatment of patients with hematological malignancies. in this study we present our results in years experience in transplantation for hematological malignancies. in a period - a total of transplants have been realized, allogeneic sibling and autologous transplantations. in the group of patients treated with allogeneic sct, % of patients were in active disease prior transplantation, with diagnosis ( aml; cml; aa; nhl; cll; all); ratio males:females = : , median age years ( - ). bone marrow (bm) as source of hsc was used in patients and were preformed with peripheral blood stem cells (pbsc) with donor sex ratio : ( / ). conditioning was provided with bu/cy ( pts), bu/cy+mel ( pts), beam ( pts), hdice( pts), nonmyeloablative flag/ida( pts), flu/mel( pt). the amount of infused fresh bone marrow was ml( - ml) with mnc . x /kg( . - , ) and pbsc , x /kg ( , - . ). median number of transfused blood products was for er median doses ( - ) and plt doses ( - ). engraftment for ne> . x /l and plt > x /l was recognized on day + ( - ). acute graft versus host disease (gvhd) gr iii/iv was noticed in patients and chronic extensive gvhd in patients. in the second group of autologous recipients, patients with diagnosis ( aml; nhl; hd; all) received fresh bm as source of hsc and the other group of patients ( aml; all; hd; nhl; mm) received pbsc previously mobilized with g-csf+chemotherapy. median age was years ( - ), males:females= : . % of patients with limfoproliferative diseases were with refractory/relapsed disease and other patients were in complete remission before sct. conditioning regimens consisted of high-dose chemotherapy mainly bu-cy, beam, ice high-dose, melphalan. the amount of infused fresh bm was ml( - ml) with mnc , x /kg( . - . ) and pbsc . x /kg ( . - . ). median number of transfused blood products was for er median doses ( - ) and plt doses ( - ). engraftment for ne> . x /l and plt > x /l was recognized on + ( - ). median follow up for both groups was months ( - ), trm in allogeneic recipients was patients ( , %) with survival of patients transplanted in cr of , %, and in the autologous group trm was patients ( , %) with survival of , %. s. genadieva-stavrik, l. cevreska, a. pivkova, z. stojanoski, b. georgievski clinical center (skopje, mk) introduction: the administration of a combination of chemotherapy and cytokines g-csf is associated with a significantly increased efficacy of stem cell mobilization compared with either modality alone. method: the aim of this study was to evaluate the efficacy of g-csf preceded by chemotherapy (cyclophosphamide g/m sq for dose) for hematopoetic progenitor cell mobilization for lymphoma and myeloma patients. we started g-csf as a fixed dose mu sq every day as soon as the leukocyte counts began to rise after chemotherapy induced myelosupression. leukapheresis was commenced at the time when leukocyte count rose up to /ul, and repeated for - consecutive days until target number of cd + cell, at least x /kg was collected. results: thirty-five patients (male to female, : , age range - , lymphoma , myeloma ) underwent a total of courses of leukapheresis for hematopoetic progenitor cell collection prior to autologous transplantation from april through may . the target amount of marrow was harvest in all patients. all the patients achieved good engraftment after autologous transplantation. the mean days required for wbc count to be over , /ul was - days. patient's age, sex, underlying malignancy, exposure to chemotherapy before mobilization did not show any statistically significant correlation. conclusion: we can conclude that chemotherapy followed by g-csf administration is an effective way for mobilization of hematopoetic progenitor cell and verified itself as a good mobilization method. cyclophosphamide + gcsf as mobilising schedule in multiple myeloma and malignant lymphoma patients: factors associated with mobilisation efficiency r. carrion, j. anguita, m. calderon, d. serrano, p. balsalobre, i. buño, a. gomez-pineda, jl. diez-martin hospital gregorio marañon (madrid, e) backgroung: cyclophosphamide (cy) at dose of . grs/m² and gcsf is commonly used to mobilize blood stem cells to support high dose therapy in patients (pts) with multiple myeloma (mm) or malignant lymphoma (ml). however, - % of pts do not achieve the minimum stem cells dose needed for a rapid hematopoietic engraftment and risk factors for poor mobilization are not well known. methods: pts diagnosed of mm or ml and candidates to autologous stem cells transplant (asct) between october and march were treated with cy . grs/m² (d ) followed by gcsf mcg/kg/d from d+ . a first apheresis procedure was planned on d + . all pts were treated as outpatients and all of them were naïve to mobilization procedures. we analyzed some clinical factors, in addition to cd + cells count, wbc and platelet count on the first day of apheresis. we defined failure as: preapheresis cd count less than /microl, or cd yield ( st day) lower than . x /kg, or cd yield ( st round, three days) lower than . x /kg. results: male/female / ; median age (min , max ); mm/ml / ( , hodgkin disease). according to diagnosis (mm vs ml) mm pts had: a higher proportion of women ( % vs %, p= . ), older (median vs , p= . ), with a lower number of previous courses of chemotherapy ( vs , p= . ). asct did not performed in pts: due to poor mobilization ( pts by disease progression, by graft tumoral contamination, and by a second neoplasm), mm pts and ml pts. twelve pts ( / , %, mm pts and ml pts) had to undergo another mobilization program (most of them with gcsf alone) to achieve sufficient stem cells. as a whole, pts ( %) would be considered as a failure to get an adequate stem cells collection. a median of days was necessary to get a minimal preapheresis cd count. this interval was not different between mm and ml pts(p= . ). significant correlations were seen between preapheresis cd + counts, cd yield ( st day) and total ( st round) (p= . ). some covariates were selected to explain failure of mobilization (bone marrow involvement in ml pts, number of chemotherapy courses, and preapheresis wbc and platelet). in contrast with other studies none factor associated with mobilization failure was found by logistic regression analysis. conclusions: ) a combination of cy + gcsf is a safe, predictable and effective for the most of mm or ml pts ( %). s ) we have not identified any premobilization factors predicting poor mobilizer pts. k. ali moghaddam, s. samiee, n. mahdavi, l. nedaeifard, m. jahani, a. mousavi, m. iravani, b. bahar, a. ghavamzadeh horc (tehran, ir) introduction: umbilical cord blood (ucb) may be an alternative source for allogeneic transplantation for the treatment of hematological malignancies and genetic disorders in patients without suitable donors particularly in ethnic minorities. early experience with umbilical cord blood transplantation (cbt) demonstrated a lower incidence of graftversus-host disease (gvhd) even though the procedure was performed with hla-disparate grafts. the major drawbacks of cbt are slow hematopoietic recovery and a high incidence of graft failure, as a result of a lower number of progenitors infused. materials and method: we collect the data of patients who had undergone cord blood transplantation by reviewing their records from the date of their transplantation up to the date of last contact. analysis of the data has done via using spss software. results: patients received cbt which in our center during the past years, consist of talassemia, severe combined immune deficiency (scid), aml-m , mps- (hurler syndrome). of all the patients ( . %) were male, and ( . %) were female. the median age was years old; ( months - ys) respectively. donors were hlaidentical siblings in ( . %) patients and unrelated in ( . %) patients. the conditioning regimen for . % of patients was busulphan, and cyclophosphamide. patients had not gvhd, ( of them died in first days), had only gi gvhd, had gi and skin gvhd, and had gi, skin and also liver gvhd. the median duration of hospitalization for transplanted patients was days. transplant mortality rate in the first days was %. the median follow-up duration was days, with minimum and maximum of and days respectively and during this period the overall survival (os) is . % and the disease free survival rate (dfs) was . % conclusion: cord blood is a considerable alternative source for hematopoietic stem cells for allogeneic transplantation for malignant or nonmalignant hematopoietic disorders. as our cases were few we can not conclude definitely about the advantages or disadvantages, but in our study cord blood recipient from hla-identical siblings had lower gvhd and mortality than unrelated donors. peripheral blood stem cell collection: from outsourcing to in-service process. an opportunity to optimise the procedure. results after one year at eio -milan d. laszlo, a. agazzi, a. alietti, l. orlando, m. cassatella, l. roveda, g. vasaturo, m. venturino, a. lettiero, c. massaro, c. rabascio, a. cocquio, g. martinelli european institute of oncology (milan, i) peripheral blood stem cell (pbsc) collection by leukapheresis (laf) represents the standard method to obtain blood stem progenitors. this procedure is usually referred to transfusion centers joined with transplant units. a possible pitfall of this kind of management could be the impossibility to perform "ad hoc" the procedure in any time. from a transplant program with pbsc collection is ongoing at the eio; until june the laf has been performed by the transfusionist team outsourcing and then governed by the medical-nurse staff of our division. aim of the study was to evaluate the outcome of this shift in management. methods: firstly we considered what kind of laf-related variable could be examined to optimise the service for the patients in terms of harvest quality and assistance and for the institute in terms of reduction of time and cost of the procedure. secondly we organized a special training with the aim to perform the laf in-service using our know-how and resources. successively a comparative analysis of the data after one year of our management has been performed and compared to the data collected during one year of the previous management. in particular all the following variables of the procedure have been analyzed by a specific data-base: a)quality of the stem cell product; b)comfort for the patient; c)time related to procedure; d)costs. results: from july to july we performed consecutive laf. the staff was operative hours every day saturday and sunday included. the two populations of patients were matched for age, sex, diagnosis, stage of disease and previous treatment. the collection target were> . x /kg and > . x /kg cd + for donors and patients respectively. the table shows the collection results obtained. the best ratio optimal collections/patients obtained with our management ( %vs %) could be attributed to the optimisation of the procedure timing and to the excellent cooperation between medical doctors and the nurses in the in-service experience. concerning the time for an optimal collection, no difference was observed into the two managements (only % of the patients needed three or more procedures). the activity of the internal staff has permitted to cut down significantly on expenses in charge of the institute for the transfusionist's performance. . conclusions: thanks to the constitution of an internal dedicated team, we were able to perform the pbsc collection with similar results and lower cost in comparison with the outsource management. ( ) ( )leningrad regional clinical hospital (st. petersburg, rus) background: mobilized peripheral blood stem cells (pbsc) have become the main source for autologous transplantation in patients with haemathological malignancies or solid tumours. the aim of the study was to establish the influence of diagnosis, sex, age, number of previous courses of ct, mobilization regimen, bone marrow involvement on the outcome of pbsc mobilization. patients and methods: patients with haemathological malignancies and solid tumors were included in the study (hodgkin's lymphoma, hl (n= ), non-hodgkin lymphomas, nhl (n= ), multiple myeloma, mm (n= ), acute leukaemias, al (n= ), solid tumors, st (n= )). ( %) were females, ( %)males. ( %) patients were > years of age, ( %) patients were an age of less than years. patients ( %) were mobilized with g-csf mcg/kg for days, a combination of ct and g-csf ( mcg/kg) has been used in patients ( %). patients ( %) received more than six courses of ct and ( %) less than six respectively. bone marrow (bm) involvement was diagnosed in ( %) patients. apheresis was performed on 'spectra'v. . (gambro). the following criteria for the mobilization outcome were used: non-mobilizable patients < ± cd +/kg; poorly mobilizable patients > ± xcd +/kg; mobilizable patients > ± xcd +/kg (s.fu et al., ) . results: patients ( %) were non-mobilizable; patients ( %)-poorly mobilizable; ( %)-mobilizable patients. according to the diagnosis we observe the following results: hl- . ± . x cd +/kg, nhl . ± . x cd +/kg, mm- . ± . x cd +/kg, st- . ± . x cd +/kg, al- . ± . x cd +/kg. comparing all groups between each other we found no significant differences. there were also no influence of age and sex on stem cell mobilization (p= . , . respectively). bm involvement does not seem to be an independent factor with significant adverse influence on pbsc mobilization (p= . ). in patients received less than six courses of ct stem cell yield was significantly higher ( . ± . x cd +/kg against . ± . x cd +/kg (p= . )). better outcome was seen in patients mobilized with ct plus g-csf than g-csf alone ( . ± . x cd +/kg against . ± . x cd +/kg (p= . )). conclusions: better stem cell yield was seen in patients received less than courses of ct and in patients mobilized with g-csf combined with ct than g-csf alone. diagnosis, age, sex, bm involvement doesn't influence outcome of pbsc mobilization. inroduction: the phenomenon of nonspecific cell aggregation (cell clumping) can be observed in nucleated cell preparations obtained from bone marrow, peripheral blood and umbilical cord blood (ucb) following thawing. such preparations containing populations of both mature as well as immature hematopoietic progenitor cells are obtained by processing whole cord blood and freezing. different techniques may be applied for the thawing of such preparations. object: to evaluate the phenomenon of cell clumping, the influence on it was examined at increasing densities of nucleated cell suspensions which had been extracted from cord blood and then frozen. the two selected techniques of thawing were also evaluated and their influence on cell clumping. methods: the fraction of nucleated cells from the ucb was obtained by sedimentation. samples containing the suspensions of these cells ( , , , mln/ml)were cryopreserved. in vitro cultures of the colony forming cells (cfc) were performed before freezing and after thawing. results: the intensity of the cell clumping increased simultaneously with the increasing density of the cell suspension. it increased from approx. % in the and mln/ml groups to approx. % in the mln/ml group, when thawed accordingly to "the classic" technique. if the solution containing dextran (rubinstein's technique) was applied post thaw to dilute the cell suspension, the clumping phnenomenon was markedly inhibited. it didn't exceed approx. % in any density group. independently to the intensity of the aggregation process, the number of the cfc among the whole pool that remained suspended after thawing (per cells), maintained a quite stable level of approx. % of the pre-freezing value. conclusions: the intensity of the cell clumping phenomenon is directly influenced by the probability of the cell contact. neverthless, the substances like dextran may markedly inhibit this process. this phenomenon is not selective process and affects both early hematopoietic cells (cfc) as well as mature cells, independly of the initial density of the frozen suspension. it seems in order to protect thawed nucleated cell suspensions from clumping and preventing the associated looses, future studies will have to concentrate on the composition of adequate freezing mediums containing clumping inhibitors. centre of slovenia, ljubljana d. domanovic, z. dovc, a. nunar-perko, p. mali, m. cukjati blood transfusion centre of slovenia (ljubljana, svn) collection of peripheral blood stem cells (pbsc) by aphaeresis is safe and reliable procedure that is generally well tolerated. beside the adverse reactions associated with g-csf or gm-csf mobilization, some adverse effects related to the aphaeresis procedure can occur. in the period from january to the end of october we retrospectively analyzed pbsc collections in autologous and allogeneic donors. pbsc collections were performed in the blood transfusion centre of slovenia, ljubljana, by standard volume aphaeresis procedures (two blood volumes processed) on the amicus blood cell separator (baxter). the targeted number of collected cd + cells was > x /kg of recipient's body weight (bw) and > x / kg bw in plasmocytoma donors. peripheral antecubital vein access was established in ( %), femoral catheter in and subclavia catheter in collections were placed without documented side effects. we have identified ( . %) adverse effects and ( . %) instrument troubleshooting. in ( . %) collections, the adverse effects occurred during the establishment of venous access (repeated phlebotomy %, unsuccessful phlebotomy %). during the collection we documented ( . %) adverse reactions. the citrate reaction in collections ( . %), hematoma in collection ( . %), fatigue in collections ( . %) and heart arrhythmia in collections ( . %). citrate reactions were mostly present as significant paresthesias and cured by oral calcium tablets and aphaeresis continued. there were no significant post donation decrease of platelet count and platelet transfusions were not necessary. the one troubleshooting was due to the unrecoverable stop ( . %) an four were during the disposable set instalation ( . %). the data confirmed the presence of mild but relatively common adverse effects in the collection of pbscs by apheresis especially in the autologous donors. the serious adverse reactions were not documented. predictive factors that affect the mobilisation of cd + cells in healthy donors treated with recombinant granulocyte colony-stimulating factor m. martino, i. callea, a. pontari, g. praticò, t. moscato, e. massara, g. console, e. quartarone, g. irrera, g. pucci, a. condemi, a. dattola, p. iacopino azienda ospedaliera "bianchi-melacrino-morelli (reggio calabria, i) no specific characteristics have been identified as predictors of peripheral blood stem cells (pbsc) mobilization in healthy donors. in this study, clinical characteristics and laboratory data for healthy donors who underwent apheresis on day of treatment with recombinant granulocyte colony-stimulating factor (g-csf) were retrospectively analyzed for correlations with cd + cell mobilization. the variables that were analyzed included age, sex, body weight, basal complete blood count and maximum white blood count (wbc) before apheresis, g-csf type and dosage. median age and body weight were . years (range - ) and . kg (range - ), respectively. by univariate analysis, male sex (p= . ), body weight (< vs. > kg, p= . ) and donors age (< vs. > years; p= . ) were correlated with the number of cd + cells mobilized. by multivariate analysis, donor's age and male sex were the only two variables that significantly predicted a high cd + cell level. in conclusion, our data suggest that male sex and younger age are the only factors that significantly affect cd + mobilization in healthy donors. hla-a is a "broad" specificity, which includes a few serologically defined antigens (spits): a , a , a , a , a , a . the hla-a occurs at a rather high frequency in most human populations, and the a splits are found at differing frequencies in different ethnic groups and populations. the distribution of a splits is very important for transplantation medicine, especially for bone marrow transplantation. incorrect assignment of a splits is one of a major problems in using serology for typing hla-a antigens. sometimes serology hardly discriminates a splits and the low resolution dna typing is required. the data of a split distribution in russians are few and ambiguous. the objective of our study was to determine the distribution of a splits in russian donors typed in research center for hematology (moscow) in . methods: donors were typed. serological typing was performed using commercial sera. in case of hla-a or homozygosity in hla-a locus revealed by serology the samples of donor blood were retyped using pcr-ssp ("protrans" or kit of research center for hematology, moscow). results: by serological typing hla-a was found in donors ( . %). the a splits were defined in . the hla-a* (by pcr-ssp) was detected in donors ( %). the most frequent among hla-a splits was a* (n= ; . %). a* and * were found with the equal frequency (n= ; . %). a* was observed only in cases ( . %) and a* in cases ( . %). in our donors we didn't reveal a* . in one case serologically defined a was not confirmed by pcr-ssp. conclusion: these results confirm the studies that dna typing for hla class i improves the typing quality. serological typing is insufficient for hla-a split identification. objectives: the availability of unrelated donor hematopoietic stem cell transplantation(hsct) could be increased with the use of allele mismatch donors if the status of hla mismatching were better understood. with the use of different transplant and immunosuppression regimens according to the patient-risk grouping will allow donor selection criteria to be refined to meet the needs best of the individual patient. methods: we conducted a hla matched or mismatched unrelated donor hsct based on the hla high-resolution typing for consecutive cases( male, female) of adult acute myeloid leukemia(aml) patients. the median age was (range, - ) and the median follow-up duration was months(range, . compared to the group of patients received hsct from hla perfect matched( / ) unrelated donor(cohort ), twentyfive( % of total population as cohort ) mismatched unrelated donor at the level of hla-cw antigen(n= ) or - alleles(n= ) at the hla-a, b, and dr loci were considered. two cases were mismatched at the hla-cw combined with allele level. fortyeight patients were in st cr and were in impending relapse status and most of them had a very poor cytogenetic profiles. the majority of patients(n= ) had intermediate or unfavourable cytogenetic features. the main conditioning regimen consisted in cyclophosphamide plus tbi(n= ) with our standard gvhd prophylaxis containing fk- plus short course methotrexate. some of cohort patients(n= ) received additional mycophenolate mofetil from the day posttransplant. the majority of patients received bone marrow as a stem cell source. the two groups had similar pre-transplant characteristics. results: all, except (cohort ), transplanted patients were engrafted. the incidence of acute gvhd( % vs %, cohort vs cohort ) was significantly different in the two cohort, but not in the chronic gvhd( % vs %). five( %) patients were relapsed in the cohort . the -year non-relapse trm was % vs %. the estimated probability of dfs and efs at -year was % vs % and % vs %, respectively. conclusion: these data suggest that allele and antigen mismatches can elicit more detrimental gvhd together with even a chance of graft failure that lead to increased trm. analysis of large hsct populations with a diversity of mismatches is necessary to define a tolerable mismatched unrelated donor so that we can enlarge the available allogeneic unrelated hsct donor pool. the majority of haematopoietic transplantation are currently performed with peripheral blood progenitor cells (pbpc). in donors, it is very important to optimize pbpc harvesting to obtain the target cd + cell dose required for transplant (> . x /kg body weight of the recipient) with a reduced number of apheresis. in the present study we retrospectively analysed data from pbpc collections of healthy adult donors (n= , m / f) with a median age of years (range - ) performed between august and october . donors were mobilized with daily administration of g-csf ( - mg/kg) for days, with generally mild to moderate side effects. total nucleated cells (tnc) and cd + cells present on pbpc grafts were evaluated with haematologic counters and flow cytometry respectively. on the collection day, the peripheral blood from most adult donors contained > cd + cells/µl, previously established as a marker of a good mobilisation. healthy donors had standard apheresis collection using the cobe spectra, with few adverse consequences, mainly related either to vascular access or to metabolic or hemodynamic changes. the majority of donors (n= %) underwent one apheresis, whilst the remaining % and % of donors had and apheresis respectively. for adult donors the pbpc collected yielded a median . tnc x /kg ( . - . ) and . x cd + cells/kg ( . - . ) body weight of the recipient and only in collections the target number of cd + cells was not reached. we found a correlation between age of the donor and the number of cd + cells collected per kg bw of the recipient, being in the older donors more difficult to achieve the required cd + cell dose. according to gender, male donors underwent one apheresis, had significantly higher pb cd + cell count prior to aphaeresis and cd + cell/kg bw recipient collected in comparison to the female donors who had a median of apheresis and had less cells (pb cd +cells: vs and cd + cell/kg bw recipient . vs . for male and female respectively). there was no correlation between the g-csf dose used in the mobilization and pb cd + cell count prior to apheresis or the total cd + cell collected. our results show that g-csf mobilisation and pbpc collection in adult healthy donors is feasible and safe for the harvesting of the graft for allogeneic haematopoietic transplantation and the main factors affecting collection are age and gender of the donor. thrombophilic screening in healthy donors treated with recombinant-human granulocyte-colony stimulating factor for mobilisation of peripheral blood stem cells m. martino, t. moscato, g. console, g. irrera, g. messina, e. massara, g. praticò, e. quartarone, c. mammì, f. luise, a. piromalli, p. iacopino azienda ospedaliera bmm (reggio calabria, i) the granulocyte-colony stimulating factor (g-csf) induces an activation state of endothelial cells and coagulation system increasing thrombotic risk. laboratory testing for the identification of heritable trombophilia in high-risk subject groups have become common practice. the objective of this study was to evaluate the effectiveness of thrombophilia screening in healthy donors prior to use g-csf to mobilize peripheral blood stem cells (pbsc). thrombophilia screening comprised of testing for factor v leiden (fvl) g a, prothrombin (ptm) g a, thermolabile variant (c t) of the methylene tetrahydrofolate reductase (mthfr) gene, protein c (pc), protein s (ps), factor viii (fviii) and homocysteine (hcy) plasmatic levels, antithrombin iii (atiii) activity, and acquired activated protein c resistance (apcr). we investigated prospectively healthy donors, men and women, with a median age of years (range - ). five donors ( , %) were heterozygous carriers of fvl g a; two healthy donors had the heterozygous ptm g a gene mutation; c t mutation in the mthfr gene was present in ( , %) donors in heterozygous and in donors ( , %) in homozygous.apcr was revealed in donors of the study ( . %). the pc plasmatic level was decreased in donors ( . %); the ps level was decreased in donors ( . %). an elevated fviii dosage has been showed in donors ( . %) and hyperhomocysteinemia in donors ( . %). concentration of atiii was in the normal range for all study group donors. the fvl mutation was combined with the heterozygous ptm g a in two cases and with ps deficiency in one case; two healthy donors presented an associated deficiency of pc and ps. the basal screening of thrombofilia permitted to identify healthy donors with a higher risk of thrombotic events. a careful monitoring should be considered in these cases before administer g-csf. during g-csf therapy, we administered low-molecular-weight heparin in all donors and folic acid, vitamins b and b in healthy donors with c t mutation in the mthfr gene and a hyperhomocysteinemia plasmatic level. our strategy of prophylaxis was correlated with the absence of short-and long-terms thrombotic and hemorrhagic side effects. no complications known to be related to the anticoagulant occurred in this cohort of healthy donors. differential levels of chimeric tolerance in a single patient. how much change in quantitative chimerism values is enough for clinical significance? d. kristt, j. stein, r. narinski, h. or, t. klein, d. kristt rabin mc (petach tikvah, il) chimerism monitoring based on strs is frequently used following sct. it is best implemented in terms of long-term tracking since engraftment is a dynamic process. when viewed this way, trends and fluctuations in the chimerism values can be observed. one fundamental question raised by such observations is what is the minimal change in chimeric status that has biological and/or clinical significance. as an indirect approach to this question, we present a case of thalassemia major which we were fortunate to follow over a year multi-sample course with two scts each with its own stable % chimerism level. following the first sct the patient developed a stable chimeric tolerance of approximately % [ - %]. however, he remained transfusion dependent, necessitating a second transplant two years ago. once again his chimerism stabilized but at a higher plateau of approximately % [ - %]. he is currently healthy, and no longer requires transfusions. in conclusion, this interesting case afforded us the opportunity to compare two different, but stable levels of chimeric tolerance, differing by approximately - %. since over the long-term, str-platform error is approximately - %, the patient is likely to have sustained a stable elevation of approximately % chimerism that freed him from transfusion dependency. the case raises a number of important questions regarding the biological implications of chimerism values, such as: ) is % a relative or absolute figure; ) is there a minimum/threshold chimerism value for functionality of the graft in these metabolic diseases; ) do all patients have the same % chimerism interval/differential and threshold? novel aspect of long-term chimeric tolerance in a patient with severe combined immune deficiency: in utero stem cell transplant suppressed by subsequent paternal iatrogenic transplant t. klein, i. yaniv, b. garti, d. kristt rabin mc (petach tikvah,il) severe combined immune deficiency (scid) is often treated with stem cell transplantation (sct). the successfully treated patient usually lives in a state of mixed graft-host chimeric tolerance. this relationship, however, may not be entirely static. we report here a case of a child in which a transfusion of maternal stem cells occurred in utero. this was documented by demonstrating a mixed chimerism in the child based on an evaluation of the child's blood using hla tissue typing and molecular dna methods. at age mo the patient manifested scid, which necessitated an exogenous sct from the father. following the paternal sct, three dna sources were demonstrated, with four hla haplotypes. however, in the ensuing period progressive loss of the maternal component of the patient's tolerance state was observed. now, after yr post-paternal sct, the patient is healthy, but there is no significant maternal dna signal demonstrated in the child, although he does have a stable chimeric level of - %. in conclusion, the three-way mixed chimerism was not tolerated, and the in utero maternal stem cell component was ultimately suppressed by the subsequent paternal sct. this case suggests that even a tri-valent tolerogenic state may be regulated by the dynamic interactions between the host and graft, enabling one set of graft-host interactions to become dominant. successful treatment of graft rejection with immunosuppression withdrawal and/or donor leukocyte infusion after early diagnosis based on t-cell mixed chimerism i. buño, p. balsalobre, g. iglesias, d. barroso, c. manzano, d. serrano, r. carrión, a. gómez-pineda, j.l. díez-martín hosp. g.u. gregorio marañon (madrid, e) background: graft rejection is a serious and difficult to manage complication after sct. objective: to evaluate the usefulness of chimerism monitoring for the early diagnosis of graft rejection in different sct settings, as well as for the follow up after treatment with immunosuppression withdrawal (isw) and/or dli. patients and methods: sct ( ablative, ric, tcdincluding haploidentical-). chimerism analysis was performed by fish for the sex chromosomes or str-pcr (sensitivity %). chimerism was analyzed in pb and leukocyte lineages (t lymphocytes cd +, b lymphocytes cd + and myeloid cells cd + isolated (purity > %) by immunomagnetic means, automacs, miltenyi biotec), every weeks, starting on day + (except in ablative), and until complete chimerism (cc) was achieved. results: after initial engraftment in all patients, graft rejection was diagnosed in ( ( %) ablative, ( %) ric, ( %) tcd), either established (severe pancytopenia and bm aplasia) or incipient (progressive decrease in pb and bm cell counts) a median of . days (range - ) after sct. all patients showed mixed chimerism (mc) in bm and pb with higher percentages of recipient cells (%r) in pb. in / patients studied, t cells showed persistent mc with high %r (> % in / ; < % and > % in / ). b cells showed mc in / patients studied, with lower %r (< % in / , > % in / ). only patients showed mc, transient in one of them, in myeloid cells. patients were not treated due to concurrent multiorgan failure and subsequently died. reduction of is in patients obtained response (normal pb and bm counts, and cc). the other patients underwent isw but no further response was obtained. one of them received a second sct while the other were treated with dli, and all of them responded. the last patient (transplanted from a haploidentical family donor) who was not receiving is, responded to treatment with dli. time from therapeutic intervention to response was variable with a median of months (range - ). patients developed gvhd>i, which was the cause of death in one and was controlled in the other three. one patient died from sepsis in complete remission (cr) months after the transplant. patients are alive in cr a median of months after sct (range - ). conclusions: the observation of mc, mainly in t lymphocytes, together with a decrease in pb and bm cell counts, allowed early diagnosis and successful treatment ( / patients) of graft rejection. immunosuppressive effects of nucleic acids -or how to learn from artefacts? t. yang, h.j. kolb, i. steinmann, m. svihla, r. buhmann gsf (munich, d) defibrotide, a single-stranded nucleic acid (ssna), was already shown to mediate immunosuppressive effects. in the current survey we investigated whether randomly chemically synthesized ssnas of different length and composition could provide similar effects. for this purpose, purified t-cells were stimulated in the presence of dntp´s or ssna with allogeneic, irradiated pbmc´s, pha or anti-cd /cd dynabeads. cellular proliferation was assessed by incorporation of tritiumlabelled thymidine (³h) thymidine), respectively (³h)damp or by staining with cfda (carboxyfluorescein diacetate, succinimidyl ester). after h or h of incubation, the incorporation of (³h)thymidine, or (³h)damp as well as the cfda distribution was assessed. cell viability was measured by trypan blue exclusion. t-cell activation was measured after h by quantifying the number of cd + t-cells expressing the activation markers cd and cd . cellular uptake of cy labelled nas was detected by fluorescence microscopy. moreover, the interference of different nas or singular nucleotides with nucleoside analogues on t cell proliferation was tested by cfda-assays. na of different length, composition or concentrations (up to mm) did not cause cytotoxic effects to lymphocytes. but the incorporation of (³h)thymidine or (³h)damp was competed by na. these effects were found to be dependent on length, concentration and base-composition of the na. the proliferative capacity of the t-cells, as assessed by cfda-staining, seemed to be unaffected. moreover it could be shown, that nas interfere with nucleoside analogues and antagonized the antiproliferative and cytotoxic effects of these drugs. the standard approach to detect cellular proliferation by incorporation of tritium-labelled nucleotides or derivatives is not useful to assess changes in cellular metabolism or proliferation in context with nas. even more important, treatment approaches using nucleoside analogues like fludarabine, cytarabine e.g. in context with nas might be critical and diminish the efficacy of these drugs. e. elli, f. colnaghi, a. colombo, m. parma, v. rossi, e. terruzzi, l. verga, a. biondi, e. pogliani ospedale s. gerardo (monza, i) introduction: chimerism status (cs) analysis is useful for evaluation of donor (d) cells engraftment after allogeneic haematopoietic stem cell transplantation (hsct). adverse events are often described as associated to a loss of chimerism, but a strict correlation between cs and residual disease is still controversial. pcr-based assays analyzing polymorphic short tandem repeat (str) markers are actually the more employed methods in cs monitoring, even if a standardized specific panel is not still available. objective: we tested a semi-quantitative method, based on multiplex pcr amplification of str markers using a commercial kit (powerplex system promega), in order to evaluate its informativity in cs monitoring and the correlation between cs and some clinical variables. methods: the informativity of the assay was tested on peripheral blood samples from allografted patients (pts) and their related sibling donors; perspective evaluation of cs was performed on pts at , , , , , , and months after hsct. pts who showed no evidence of recipient (r) cells were considered to have a complete chimerism (cc), pts who presented each d and r cells were defined as mixed chimerism (mc). results: the multiplex assay gave at least one high informative marker (range: - , median ) in all the pts. we analyzed blood samples from pts, ( , %) presented cc, ( , %) mc and none autologous reconstitution. some pts were defined as having an increasing mc (imc) when they shifted from cc to mc or when in a mc setting the r amount increased in two or more consecutive controls. we evaluated if there was a correlation between cs and some clinical variables: r/d gender, diagnosis, r/d sex mismatch, abo system incompatibility, conditioning regimen, cd + and cd + cell dose infused, disease status at hsct, stem cells source, acute and chronic graft versus host disease (gvhd), marrow relapse. chi-square analysis demonstrated a significant correlation between icm and a brief time marrow relapse, moreover a low incidence of chronic gvhd, male d and diagnosis of acute leukemia seem to be associated with increasing level of r dna. conclusion: pcr amplification of a panel of str loci is an informative method to evaluate cs in pts after hsct. imc seems to be useful to predict marrow relapse; some clinical conditions such as male donor and acute leukemia diagnosis seem to limit a complete d engraftment; chronic gvhd is favorable for a stable cs. non-haematopoietic tissue repair r gmp production of autologous cd + cells for intracoronary administration after acute myocardial infarction r. giordano ( ) subjects affected by acute myocardial infarction (ami) with absent angiographic myocardial blush (mb) and lack of st segment elevation resolution after primary angioplasty, have short-and long-term poor clinical prognosis. we recently started a phase i/ii randomized controlled study based on the hypothesis that, in this target population, after primary angioplasty and stenting, intracoronary injection of cd + cells from bone marrow (bm, group a) or mobilized peripheral blood (mpb, group b) could enhance endothelial regeneration and improve heart function compared to controls treated with standard pharmacological therapy alone (group c). the study started in june and it is expected to enroll patients ( per each randomization group). up to november , patients have been included. in group a (n= ), bm was processed within hours of collection. in group b (n= ), the administration of g-csf ( µg/kg/day for - days), started from day - after ami and leukapheresis was performed following standard procedures. an automated cd + stem cell selection was performed with the clinimacs® plus instrument (miltenyi biotec) in our class b -iso facility. the mean (±sd) number of cd + cells after immunoselection was . x (± . ) in bm samples and . x (± . ) in mpb samples respectively, with a purity of % (± ) in the group a and % (± ) in group b. the percentages of viable cells (propidium iodide) in the post-selection samples were (± ) in group a and (± ) in group b, respectively. the sterility tests for bacteria and fungi on the purified samples were negative. purified cd + cells were injected in the culprit vessel using a well-sized over-the-wire angioplasty balloon within three minutes of occlusion. no adverse events have been reported during and immediately after the cell administration. this results show that cd + selection is feasible and safe also in ami patients. the short and long term efficacy of this cell therapy approach in preserving the myocardial viability and function after ami is currently under investigation using pet-based techniques and echocardiography. defibrotide (df) is a polydisperse mixture of % singlestranded polydeoxyribonucleotides with anti-thrombotic, profibrinolytic and anti-apoptotic functions. df is already successfully used in the treatment of hepatic veno-occlusive disease in allogeneic stem cell transplantation (sct). our observation that df can also protect endothelial cells (ec) from conditioning (fludarabine)-mediated apoptosis ( ) prompted us to apply it prophylactically to patients (pts) at risk for endothelial complications. pending on the magnitude of risk, pts received - mg every h in h-infusions, usually from day (d) - until d+ post sct. circulating ec (cec) as a marker of conditioning-mediated endothelial toxicity ( ) were detected by magnetic bead separation of cd + cells from edta blood of sct pts ( df, non-df) and co-staining with ulex europaeus antigen lectin . cec maxima until d+ post sct were compared between the two groups. df pts showed significantly lower maxima of cec than untreated pts ( [± ] in the df treatment group vs. [± ] cec/ml in non-df pts, respectively, p= . ). similarly, when cec maxima were compared in the time period of df prophylaxis, again, df pts had less cell counts ( [± ] vs. [± ] cec/ml in control pts, respectively, p= . ). in an overlapping cohort of pts ( non-df, df) serum was assayed for its induction of apoptosis in a human microvascular endothelial cell line (hmec), a monitoring approach that had been found to correlate with episodes of gvhd and severe microangiopathy ( ). apoptotis was determined by flow cytometric analysis of the cellular granularity of propidium-iodide-negative indicator hmec. similar to the cec measurements apoptosis inducing maxima until d+ were compared between df and non-df pts and turned out to be significantly different (apoptosis by pts´ sera normalized to untreated control hmec: . [± . ] in df pts vs. . [± . ] in non-df pts, p= . ). these preliminary analyses suggest the protective efficacy of df prophylaxis in the course of sct. the final proof of principle is to be validated in long-term clinical follow-ups. .g. eissner et al., blood , - ( ). .a. woywodt et al., blood , - ( ). .a. ganster et al., bone marrow transplant. , - ( . treosulfan, cyclophosphamide and anti-thymocyte globulin for allogeneic haematopoietic stem cell transplantation in severe aplastic anaemia s. giebel, j. wojnar, m. krawczyk-kulis, m. markiewicz, i. wylezol, t. kruzel, m. kopera, j. holowiecki silesian medical university (katowice, pl) graft rejection is the major cause of failure after allohsct in severe aplastic anemia (saa), when cyclophosphamide is used as a single agent for conditioning. to reduce the risk of this complication we decided to intensify the preparative regimen by adding reduced dose of treosulfan -an alkylating agent possesing both immuno-and myeloablative properties. between between - years) were treated in a single institution with allohsct from either hlaidentical sibling (n= ) or an unrelated volunteer (n= ). conditioning regimen consisted of treosulfan g/m²/d on days - , - , cyclophosphamide mg/kg/d on d. - , - , - , - , and anti-thymocyte globulin (thymoglobulin, genzyme) mg/kg/d on d. - , - , - . each bone marrow and peripheral blood was used as a source of stem cells in cases. all patients engrafted with the median time to neutrophil > . x /l and platelet > x /l recovery of ( - ) days and . ( - ) days, respectively. complete donor chimerism was achieved on day + in all cases. none of the patients developed grade iii-iv acute gvhd, one patient experienced grade ii acute gvhd. at one year the cumulative incidence of extensive chronic gvhd equaled %, overall cgvhd - %. at the median follow-up of . ( - ) months all patients remain alive and disease-free with complete donor chimerism. at one year the karnofsky index equaled % in patients, % -in one case. we conclude that treosulfan + cyclphosphamide + antithymocyte globulin is a well-tolerated preparative regimen and allows stable engraftment in saa patients. the use of treosulfan allows intensification of the conditioning without providing an additional non-hematological toxicity. clinical outcome in adults with severe aplastic anaemia. a retrospective single-centre analysis s. buchholz, e. dammann, c. koenecke, e. mischak-weissinger, m. stadler, m. eder, j. krauter, b. hertenstein, a. ganser hannover medical school (hannover, d) introduction: allogeneic bone marrow transplantation (bmt) is the treatment of choice in young patients suffering from severe aplastic anaemia (saa). due to improved, less toxic conditioning regimens and advances in prophylaxis against graft-versus-host-disease (gvhd) survival has improved steadily. nonetheless, long-term side effects, such as chronic gvhd, occur in up to % of patients requiring treatment and leading to an increased mortality. here we present the analysis of for patients with saa comparing. patients and methods: between and , twenty one patients ( male, female) with saa and one patient (female) with paroxysmal nocturnal haemoglobinuria (pnh) were transplanted in our centre. the median age at transplantation was . years (range - ). fifteen patients were transplanted with stem cells from their hlaidentical related donor, patients from hla-identical matched unrelated donors and patients were transplanted from a syngeneic donor. in cases stem cell source was bone marrow (bm), patient received bm and peripheral stem cells (pbsc) and were transplanted with pbsc. conditioning regimen consisted of cyclophosphamid (cy) alone (n= ), or in combination with either total nodal irradiation (n= ), or with fludarabin (flu, n= ). one patient was treated with cy and total body irradiation (tbi) while patient received flu, cy and tbi. gvhd-prophylaxis was cyclosporin (csa) and methotrexate (mtx) in all but the patients transplanted from syngeneic donors. all patients engrafted. acute gvhd developed in patients ( %) and was readily controlled by immunosuppression. chronic gvhd occurred in patients ( %; limited, extensive) within a median follow-up of years (range . - . years) . twenty one out of patients are alive and free of haematological disease, one patient died because of toxoplasmosis before day + . conclusion: the incidence and severity of acute and chronic gvhd is similar other studies. our data suggest that bmt is a favourable therapy for young patients with aplastic anaemia, showing good engraftment, controllable complications and a good clinical outcome. stem cell transplantation and immunosupressive treatment of severe aplastic anaemia: single-centre experience l. tukic, d. stamatovic, o. tarabar, v. glavicic, m. elez, l. simic, s. marjanovic, m. malesevic military medical academy (belgrade, cs) background: stem cell transplantation (sct) from an hlaidentical fully mached sibling donor (msd) is the best treatment option for severe aplastic anaemia (saa). patients without suitable msd should be treated with immunosuppressive therapy (ist). patients and methods: between / and / patients with newly diagnosed saa were treated either with sct from msd ( patients) or with ist ( patients). there were performed allogeneic sct in patients. all donors were hla-identical sibling ( donor was identical twin). source of stem cells was bone marrow in (one with second transplant) and peripheral stem cell in scts. conditioning regimens were based on cyclophosphamide (cy) with atg in and fludarabine with cy and atg in scts. patients received combined ist with antithymocyte or antilymphocyte globuline (atg/alg), cyclosporine a and steroids and patients atg with steroids. the median interval from diagnosis to ish was days (range to ). results: engraftment was documented in patients with allogeneic sct ( patients died without engraftment). one patient developed acute gvhd grade - and died after days, and the other had pneumonitis interstitialis (cmv+) and died after days. till november of patients ( %) are alive with sustained engraftment. median survival from sct is months (range to ). concidering ist, of patients ( . %) achieved response ( had two or tree cycles of ist). one patient relapsed year after ist. two patients from ist group died, major causes of death were infection and hemorrhage. overall survival in the ist group is . % ( / patients) after a median follow up of . months (range to ). conclusion: our results confirm significant improvement in outcome of saa patients during last decades in due to modern induction front line therapy including allogeneic autoimmune diseases r successful treatment of autoimmune thrombocytopenic purpura after bone marrow transplantation with anti-cd antibody: a case report b. giannini, c. bosi, m. stanzani, g. bandini, f. bonifazi seragnoli (bologna, i) we describe a case of persistent, severe autoimmune thrombocytopenia, refractory to prednisolone but responsive to chimeric monoclonal antibody anti-cd ( rituximab). a patient transplantated for hodgking disease (upn ), developed autoimmune thrombocytopenia with severe bleeding, days after unrelated bone marrow transplantation: at the same time there were no signs of graftversus-host-disease, cytomegalovirus infection, sepsis or microangiopathic process. high titer of antibody against platelet antigens was found. during treatment with prednisolone mg/kg, for two days, the platelet count remain below /µl. in spite of increasing the dose of cyclosporine and methilprednisolone ( mg/kg/daily) with addition of intravenous immunoglobulin for five days, only a transitory partial response ( platelet /µl) was observed. after four days from last dose of immunoglobulin, the platelet count fell again below /µl; antiplatelet antibodies still highly positive. after two weeks of therapy with steroid, we started with anti-cd antibody with first dose of mg/m² followed by dose of mg/m² once weekly for weeks. complete response was achieved after weeks from therapy initation, with a complete normalitation of platelet count and with disappearance of antiplatelet antibody in peripheral blood. no apparent toxicity, or side effects that could be attributed to rituximab were observed. the patient is in complete response mounths after therapy with anti-cd . rituximab induced complete response in approximately % of the patient with immune thrombocytopenic purpura refractory to prednisone or splenectomy; it's also effective in patients with secondary itp. to our knowlege, only few cases of itp after bone marrow transplantation have been successful treated with rituximab. we suggest for an early use of anti-cd in similar cases. treatment of a malignant form of multiple sclerosis with immune ablation and autologous stem cell transplantation v. kimiskidis, i. sakellari, c. smias, k. kapinas, a. anagnostopoulos, a. kazis, a. fassas g.papanicolaou (thessaloniki, gr) malignant forms of multiple sclerosis (ms) are rare cases characterized by very aggressive demyelination and rapid progress to disability leading to death within years from onset despite treatment, which fails to control the disease. we report a case of a young male patient of years old with aggressive ms who was treated with a high-dose immunosuppressive regimen (hdis) using myeloablation followed by autologous blood stem cell transplantation (asct) that has induced a dramatic and long-lasting remission of the disease. the patient was diagnosed with ms in june . at that time he had minor disability, his edss score being was . , and active disease on mri showing gadoliniumenhacing (gd+) lesions. he was put on steroids and interferon-beta which, however, had no effect, while disability was rapidly accumulating. by february , i.e. within months, the edss score rose to . and the patient was unable to walk unaided for more than metres. on mri, gd+ lesions increased to , as did their volume. in march it was decided to treat him with hdis and, in order to mobilize blood stem cells, he received cyclophosphamide (cy) g/m² plus gcsf ug/kg. six days after cy infusion he had a disease flare with worsening of edss score to . , and further increase in the number and volume of gd+ lesions. the patient was treated with steroid pulses for days and showed some improvement which allowed the continuation of g-csf and subsequent stem cell collection. two months after cy infusion, he underwent asct with busulfan mg/kg over days plus antithymocyte globulin . mg/kg for conditioning. one month post asct the edss score dropped to . and no gd+ lesions were detected. the patient continued to improve over the following years. the last assessment at months after asct showed nearly absent disability (edss score: . ) and, again, no gd+ lesions on mri. the spectacular responses of this case and also of the three similar cases reported in the literature support the role of asct in rapidly evolving, so-called malignant, forms of ms. an obvious amelioration in a case of scleromyxedema after successful double autologous peripheral blood stem cell transplantation followed by thalidomide and bortezomib consolidation s. ataergin, f. arpaci, a. ozet, m. demiriz, s. komurcu, b. ozturk, o. kuzhan gata (gulhane) objective: scleromyxedema is characterized by cutaneous deposition of mucin, dermal fibrosis and monoclonal gammapathy. the response to treatment has been variable after several treatment modalities including high-dose treatment (hdt) and autologous stem cell transplantation (asct). methods: we report a case of scleromyxedema who achieved amelioration after double hdt and asct, followed by thalidomide and bortezomib consolidation. a -year-old male patient was admitted with persistent pruritis. he was treated with anxiolytic and antidepressant drugs with diagnosis of neurodermatitis; however, the skin was thickened and papular lesions appeared. a skin biopsy revealed scleromyxedema. he was treated with topical corticosteroids and retinoic acid preparations. however, no amelioration were noted. topical cyclophosphamide, systemic corticosteroids were used for three months, then the patient has discontinued all the medications. low-dose interferon-a treatment ( . mu) was initiated for three times weekly; however, the treatment was stopped due to an acute and severe rhabdomyolisis after the third administration. on his admittance in our department, he had papular lesions of . mm without pain, erythema or desquamation all over his body. he had also some nodular formations of . x . cm, on his face and neck. the whole blood count and biochemical analysis were normal. erythrocyte sedimentation rate, crp were within normal limits and all viral markers were negative for hepatitis, ebv, cmv, toxoplasma, hiv as well as rheumatologic markers. serum immunoglobulin (ig) levels were within normal ranges except for ig g and ig light-chain lambda. the peripheral blood smear and bone marrow aspiration and biopsy revealed no pathology. he then underwent an asct after conditioning with melphalan ( mg/m²) and a second transplant was done four months later using the same conditioning regimen ( mg/m²). results: after the transplantation, the immunoglobulin levels have partially regressed. physical appearance has been ameliorated and the skin biopsy revealed a regression in mucin deposition in dermis. consolidation treatment was initiated with thalidomide followed by bortezomib. he is still on his follow-up without any progression for three years. conclusion: hdt and asct may be an alternative treatment in the amelioration of lesions related to scleromyxedema. ( ) the depletion of autoreactive lymphocyte populations in the graft is mandatory for the success of autologous transplantation in patients with severe autoimmune disease. clinical grade cd + selection can be used to obtain lymphodepletion in autologous leukapheresis products. we report on the implementation of a gmp production process of autologous purified cd + cells from mobilized peripheral blood of patients affected by systemic sclerosis (ss) in our institution. for cd + cell mobilization, patients ( male, females, aged years ± ) with ss refractory to standard immunosuppressive therapy, received cyclophosphamide (cy) g/mq and g-csf (filgastrim, ug/kg/day s.c.) starting days after the last cy administration until stem cell collection. leukapheresis was performed when wbc and cd + cell count were at least x /l and /ul respectively, using a fresenius hemocare as (c y) instrument. the automated selection (clinimacs, miltenyi) was performed in hours of leukapheresis collection. the selection procedure and the preparation of the final product for the cryopreservation were performed in our class b -iso facility. the cells, resuspended in normal saline solution and human serum albumin, were cryopreserved in % final dmso. for quality control of the cryopreserved product, the cells from a satellite segment of the cryopreserved bag were thawed and their viability evaluated by trypan blue exclusion. the mean (±sd) wbc, cd + and cd + cell content of the pre-selection products were , x (± , ), x e (± ) and , x (± , ) respectively. after immunoselection we obtained a mean (±sd) cd + and cd + cell number of x (± ) and . x (± . ), with a purity of % (± ). the mean percentage of viable cells (propidium iodide) in the post-selection samples was (± ). the sterilty tests for bacteria and fungi on the purified samples were negative. the viability of the post-thawed samples was % (± ). the conditioning regimen consisted of cy mg/kg/day for days i.v. and rabbit anti-thymocyte globulin . mg/kg/day for days. neutrophil engraftment was reached at day + (± ), and all patients are alive, with stable or improved clinical conditions after months (± ) after transplantation. these results demonstrate that gmp production of cd + cells for autologous transplantation is feasible, safe and could efficiently support a transplantation program for patients affected by ss. high-dose immune immunoablative therapy and autologous stem cell transplantation in severe resistant crohn's disease: profound response for months followed by treatable relapse y. sorour, k. robinson, p. hurlstone, k. el-ghariani, a. lobo, j.a. snowden sheffield teaching hospitals (sheffield, uk) we report the case of a year old female with severe resistant crohn's disease (cd) treated with high-dose immune immunoablative therapy (hdit) and autologous stem cell transplantation (sct). diagnosed with cd at age , initial disease control had been achieved with azathioprine and steroids, but, at age , colectomy and ileostomy were performed for a severe flare. from age , increased disease activity was unsuccessfully controlled with azathioprine, steroids, infliximab, methotrexate, combination rifabutin/metronidazole/clarithromycin, thalidomide and tacrolimus. from nov -jan , surgical episodes had resulted in resection of . m small bowel. dietary modifications had included an elemental diet, but from the patient was dependent on home total parenteral nutrition. severity of symptoms had resulted in recurrent and prolonged inability to work and to warrant care under a palliative medicine specialist. based on poor quality of life, risk of life threatening complications, and inability to control the disease effectively, the option of autologous transplant was pursued after examination of the case and proposed treatment protocol by the local research ethics committee, review by two independent gastroenterologists and one transplant haematologist, and obtaining informed written consent. treatment commenced in sept with mobilisation using cyclophosphamide (cy) g/m² and g-csf. in nov the patient was treated with cy mg/kg, rabbit atg mg/kg and methylprednisolone gx followed by . x /kg isolex enriched cd + cells. treatment was complicated by neutropenic sepsis, oropharangeal and stomal mucositis. engraftment time was within normal limits. discharge was on day+ . pre-and months post-treatment data are summarised in the table. cd remained inactive until march with the development of increased stoma output and abdominal pain. relapse was confirmed by ileal biopsy. in contrast to pre-sct, disease control has been achieved with immunosuppressants and surgery, permitting ileal reanastomosis, pouch formation and reversal of ileostomy in sept . the patient remains stable as of nov . in conclusion, hdit and autologous sct may be an effective therapy for medium term control of severe, treatment resistant cd. it remains to be seen whether post-sct relapse is easier to control than cd activity before sct, as suggested by data in other autoimmune diseases. in addition to randomised trials, future studies could look at means of prolonging responses, such as maintenance treatments. hyperkalemia is one of the side effects of cyclosporin (csa). the mechanism of hyperkalemia is unclear. apparently many factors in pathogenesis of hyperkalemia may be involved. nephrotoxicity of csa that significantly impairs renal perfusion, glomerular filtration reduced k+ excretion and secondary hypoaldosteronism seemed to be the reasons for csa-associated hyperkalemia. there are publications about hyperkalemia in patients after renal transplantation but only few reports devote this phenomen in bone marrow transplantation patients. we report about cases of hyperkalemia with bradyrhythmia during csa administration in patients with cml in cp, male and years old undergoing bmt from hla -identical siblings. hyperkalemia ( , - , mmol/l) and bradyrhythmia ( - /min) in - days after transplantation was observed. at that time a serious worsening in renal function (increase serum creatinine and serum urea, decrease filtration rate) and in serum csa level rise was found in both. after discontinuation of csa treatment all these symptoms disappeared. these observations suggest that csa may be a cause of hyperkalemia associated with bradyrhythmia in bone marrow transplantation patients. careful monitoring of csa level in blood and renal function may be important in prevention these complications. objectives: allogeneic haematopoietic stem cell transplantation (allo-hsct) is the only curative therapy for patients with chronic lymphocytic leukaemia (cll). however, transplant-related mortality remains relatively high and relapse is still a major problem. there are only few anecdotic reports of the use of thalidomide, an immunomodulating agent, in such patients. case report: -year old patient with resistant cll is presented. he was treated unsuccessfully with several cycles of different agent combinations, started with chlorambucilmethylprednisolone then fludarabine-cyclophosphamide and also with monoclonal antibody rituximab. as he has hla matched sister we proposed allo-hsct. on admission before transplantation patient was presented with massive lymphadenopathy, anemia (hb g/l), wbc . x /l with absolute lymphocytosis (lymphocytes %) and thrombocytopenia x /l. pre-transplant conditioning consisted of high-dose cyclophosphamide and total body irradiation. combination of monoclonal antibody alemtuzumab (campath- h), cyclosporine a and short methotrexate were given for the prevention of acute graft versus host disease (gvhd). the patient received . x /kg stem cells. there were no serious complications in post transplant period. lympadenopathy completely disappeared. patient was discharged weeks after transplantation, without gvhd and with normal complete blood counts (cbc). on bone marrow examination there was still residual leukaemic infiltration ( %). two months later acute gvhd developed, with skin involvement only. at that time lymph nodes massively enlarged again and high wbc with absolute lymphocytosis reappeared. cyclosporine immunosuppressive therapy was stopped and thalidomyde mg/day was introduced combined with methylprednisolone mg three times per week. during period of months treatment he was readmitted once for lung aspergillosis which responded well to voriconazole. improvement appeared slowly, with regression of lymphadenoapthy. after months he still has residual leukaemic marrow infiltration -about %, but normal cbc without absolute lymphocytosis. conclusion: our patient is new evidence that thalidomide may have significant antileukaemic effect in refractory cll. due to this and anti gvhd effect perhaps in the future it could be incorporated as a first line gvhd prophylaxis regimen in patients transplanted for cll. intensive combination therapy and autologous pbpct for blast crisis cml revisited d. heim ( ) imatinib is the most active therapy for chronic myeloid leukaemia (cml) in all phases of the disease. overall survival of patients with blastic phase cml treated with imatinib monotherapy however is months only. the reason for imatinib resistance in advanced phase cml is mostly due to bcr-abl independent mechanisms. therefore a combination therapy of imatinib with conventional high dose chemotherapy is often used for remission induction and allogeneic stem cell transplantation is performed if a suitable donor is available. autologous stem cell transplantation has drawn new attention in the treatment of cml since sufficient numbers of peripheral stem cells (pbpc) can be mobilized under concomitant treatment with imatinib and collection of bcr-abl negative autologous peripheral stem cell transplants has been reported. we tested in a pilot trial of patients the feasibility of a treatment consisting of imatinib mg qd combined with cycles of cytarabine mg/m² x d in patients with cml in blast crisis (bc) who do not have a hla-matched stem cell donor. pbpc were mobilized after the th cycles of cytarabine with g-csf (filgrastim) ug per day. pbpc were cryopreserved and reinfused after a conditioning therapy with either cyclophosphamide mg/kg and busulfan mg/kg (bucy)(table : patient + ) or cyclophosphamide mg/kg plus gy tbi (cy/tbi)(table : patient ) the stem cell products of all patients contained sufficient numbers of cd + cells after mobilization with g-csf after the th cycle of high dose cytarabine and imatinib given through. the autologous pbpc graft of one of the patients was bcr-abl negative, a second graft had detectable bcr-abl at low level in the q-rt-pcr (table ). no data on the presence of bcr-abl is available for the third graft. engraftments of the transplants were in the expected range. no excessive toxicity was recorded. treatment of cml in bc with imatinib combined with high dose cytarabine and autologous pbpct after high dose chemo-/radiotherapy is feasible and may result in sustained complete molecular remission. alemtuzumab and autologous sct in cll, experience in a small centre e. zappone, e. ortu la barbera, u. coppetelli, c. ciabatta, f. ciccone, s. nardelli, a. centra, g. potente, a. de blasio ospedale s.m. goretti (latina, i) alemtuzumab (al) as a single agent or in combination with chemotherapy is an effective treatment for chronic lymphocytic leukemia (cll) in refractory or relapsing patients (pts) and has been shown to induce complete molecular responses. autologous stem cell transplantation (sct) induce prolonged and durable remission in many hematological malignancies but its role in the treatment of cll is controversial. we included al in the treatment of refractory, relapsed or high risk cll pts prior to stem cell collection and high dose chemotherapy consolidation of the response obtained. we treated pts with binet stage iii cll in partial remission (pr) or stable disease (sd) after to lines of chemotherapy, including fludarabine and pt with advanced b-lymphocytic lymphoma in pr. chemotherapy debulking prior al was necessary in pts with high disease burden and large nodal involvement. disease status before al treatment was complete remission (cr) , pr , and sd . al was given subcutaneously, treatment duration was - weeks and total dose was - mg. results after al were cr and pr , all pts had below normal lymphocytic counts, and lymphoid marrow infiltration below %; out of cll pts had cd /cd expression in the marrow below %. after al out of pts were mobilized: with cyclophosphamide - gr/mq and with additional chemotherapy followed by aracytin; mobilization is planned in the last pt. stem cell collection was adequate: - cd x /kg, in - apheresis procedures. two pts were not mobilized after being treated for cytomegalovirus (cmv) infection at the end of al treatment, they maintain cr without further treatment and months after al. four pts undergo sct: engraftment and clinical course were normal, pt progressed before transplant. of the transplanted pts are in ccr at , and months post sct, pt progressed months post sct and was retreated with al with minor response. cmv reactivation occurred in out these pts and antiviral therapy was necessary in . al was effective in inducing significant clinical response in these high risk pts, stem cell collection was feasible and autologous sct could be performed without significant early or late complications. cmv reactivation occurs in the majority of al treated pts and must be carefully monitored. the results in this very small group of patients are encouraging but we can not draw any conclusion about the general application of this program in cll. at the time of imatinib era in cml treatment hsct became a disputable issue. in this study we compared the outcome of unrelated donor transplantation to that in sibling donor setting in cml patients receiving reduced toxicity conditioning. patients received hsct from matched sibling donors, patients from unrelated donors. reduced toxicity consisted of: : busulfan mg/kg, fludarabine mg/m² (allo sib) or mg/m² (mud) and atg. donors for unrelated transplant were matched for specificities at following resolution levels : loci a, b, c at intermediate or high resolution and dr at high resolution level only. patients were stratified into groups : i) allo-sib ( hla matched ; n= ) , ii) mud fully matched ( / match at intermediate or high resolution level for hla class i and high resolution for class ii ; n= ) , iii) mud mismatched ( at least allele mismatched ; n= ). cumulative proportion survival was: i) % at the end of y follow up period in sib hsct , ii) % in patients transplanted from mud donors fully matched in specificities and iii) % in patients transplanted from donors mismatched in at least one allele. (figure) our results document, that optimal matching in five loci benefit the outcome of transplantation, which in unrelated donor transplantation can be similar to that obtained with sibling donors providing / matched donor at high resolution level. refractory graft-versus-host disease after stem cell transplantation in thalassaemia: pentostatin is safe and effective treatment g. leopardi, g. visani, c. giardini, g. sparaventi, f. d'adamo, g. nicolini, b. guiducci, s. barulli, m. lucesole, l. malerba, a. isidori san salvatore hospital (pesaro, i) toxicity, graft rejection with return of the thalassemia and graft-versus-host disease (gvhd) are the main causes of failure after stem cell transplantation (sct) in thalassemic patients, particularly in those at poor prognosis (i.e. class and ). steroid refractory gvhd is associated with a not negligible non-relapse mortality. few data are actually avalaible on the use of pentostatin for refractory gvhd in thalassemic transplanted patients. we analyzed four children, females and males, aged - (median . years), transplanted for beta-thalassemia major ( class , class , class ). two patients (class and class ) received unrelated sct ( bone marrow, pbsc). the two other children (both class ) were transplanted from hla identical sibling after previous transplantation procedures, having rejected twice and once, respectively. three patients ( unrelated and hla identical sct) developed refractory acute gvhd grade iii-iv with multiple organ involvement at + , + and + days, respectively. one patient had chronic extensive gvhd. in the patient receiving the third allogeneic hla identical sct both acute and chronic refractory gvhd occurred. patients affected by acute severe gvhd failed to respond to primary treatment with cyclosporine a and methylprednisolone at doses varying from to mg/kg/day and thus received salvage therapy with pentostatin . mg/sqm for consecutive days. one presenting with acute multiorgan gvhd (skin, liver and gastrointestinal tract) had short term response. two patients responded: one developed a secundary, chronic extensive, gvhd. this child as well as the other with chronic severe extensive gvhd (involving skin, liver, eyes and oral mucosa in one case and skin and mouth in the other) starting at + and + days from transplant were treated with pentostatin ( mg/sqm i.v. every weeks) for and months. they are alive with significant improvement of skin and mouth symptoms and tapered concurrent immunosuppressive treatment. no toxicity or impairing chimerism due to pentostatin were observed. pentostatin thus appears as safe and effective treatment for acute and chronic severe gvhd after sct for thalassemia. supported in part by ail pesaro onlus pure red cell aplasia after mud stem cell transplant in thalassaemia major: successful treatment with rituximab m. lucesole, g. visani, c. giardini, b. guiducci, g. sparaventi, g. nicolini, f. d'adamo, g. leopardi, s. barulli, l. malerba, a. isidori hematology and transplant center (pesaro, i) pure red cell aplasia (prca) is a not infrequently observed complication of allogeneic sct performed across the abo complex and often refractory to standard strategies. the peculiar recovery of erythropoiesis after sct in thalassemia major could be a possible factor confounding for a correct diagnosis, as well as for the therapeutic choice. a young male patient was transplanted from abo incompatible (donor b rh+; recipient rh+) hla-identical unrelated donor for beta thalassemia major. after standard myeloablative conditioning regimen (bu-cy-thio), he received cyclosporine a and mtx as gvhd prevention prophylaxis. the post-transplantation course was characterized by an incomplete haematological recovery. a poor graft function was observed at day + . the bone marrow was hypoplastic with the apparent absence of erythroid precursors, and a possible rejection of the graft was suspected; nevertheless, fish analysis of chromosome y for the engraftment showed % of donor cells, with a normal count of the beta chains ( %); no evidence of haemolysis was recorded. all dna virus were negative. the diagnosis was prca. epo . u/every other day was started by day + ; additionally we submitted the patient to plasmapheresis ( procedures, last at day + ) without hematologic response ( - units rbc/weeks, - platelets for week). at day + a first dose of rituximab (rtx) ( mg/sm) was administered; we observed a progressive increase in reticulocytes and platelet counts (respectively /mm³ and /mm³) days after the infusion of rtx. insorgence of cystitis and pielonephritis (p.aeruginosa) delayed the administration of a following dose of rtx with a progressive, increasing transfusion dependency. after the resolution of the infective complications, a second dose of rtx was administered on day , and a third dose on day + . two weeks after the last dose of rtx a response was observed: reticulocytes count increased, and the level of hgb slowly normalized .the patient is now in complete remission months after transplantation, and with a complete hematologic take. rituximab could thus be a promising agent for treatment of not infrequent cases of prca, in transplanted thalassemia patients, refractory to standard therapies. a successful t-lymphocyte engraftment achieved by megadose cd +selected peripheral blood stem cell s transplantation in a t-b-nk+ scid case without using conditioning regimen a. ikinciogullari ( ), c. aytekin ( ), f. dogu ( ) , m. yuksek ( ), a. yildiran ( ) severe combined imunodeficiency (scid) is a genetic disorder characterized by profoundly defective or absent t and b cells functions. allogenic stem cell transplantation (sct) is to date the only curative therapeutic option for scid. here we report a t-b-nk+scid patient who was transplanted by megadose peripheral blood stem cells (pbsc) collected from his father without conditioning regimen. case: a months old boy referred to us with the diagnosis of t-b-nk+scid. his physical examination showed disseminated hyperceratotic papular skin lesions. immunohistochemical investigation of the skin biopsy revealed hsv infection. he was treated with acyclovir and foscarnet combination, but skin lesions didn't resolve. thus he received a megadose cd + selected (clinimacs, miltenyi biotec) pbsc ( x cd +cells/kg) transplantation from his father without conditioning regimen. he received cyclosporine for gvhd. he engrafted at posttransplant day . detection of chimerism performed by str (short tandem repeat) pcr analysis and whole blood samples showed mixed chimerism with % donor t cells. acute gvhd grade i developd at day rapidly resolved with systemic corticosteroid treatment. his chronic hyperceratotic papular herpetic lesions completely recovered at second month after sct. he is doing well with successful immunoreconstitution five months after sct. to our knowledge he is the first successfully engrafted scid case with t-b-nk+ phenotype following uncoditioned haploidentical pbsc transplantation. myeloid-related proteins (mrp ) and (mrp ), both s proteins, are the major calcium-binding proteins expressed in phagocytes during specific stages of differentiation. they form stable complexes and are present in circulating neutrophils and monocytes, representing the first cells invading inflammatory lesions. the protein complex is found in inflammatory fluids in distinct inflammatory conditions, including rheumatoid arthritis, allograft rejection, inflammatory bowel disease, and lung disease. prerequisite for its secretion is the contact of phagocytes with extra-cellular matrix proteins or inflamed endothelium, resulting in elevated intracellular calcium levels and activated protein kinase c. mrp /mrp is thereby released specifically at inflammatory sites and leads to increased serum levels in correlation with the degree of inflammation, indicating an extra-cellular role of these molecules in inflammatory processes. we report a year-old girl with: a) severe anemia, b) neutropenia, c) inflammation and d) severe growth failure. bone marrow examination showed moderate dyserythropoiesis. we did not detect hemolysis, iron deficiency, hemoglobinopathies, immunological diseases or any autoantibody. serum levels of copper and ceruloplasmin were within normal range, although serum zn concentration was markedly increased ( µg/dl). urinary zn excretion and erythrocyte zn concentrations were within normal range. family studies demonstrated normal zn and cu plasma levels. patient's plasma calprotectin concentration showed a -fold increase ( mg/l) compared to normal values. calprotectin concentration is known to be elevated in many inflammatory conditions but is generally below mg/l and thus far below the levels reported in this patient. we describe this case as an inborn error of zinc metabolism caused by dysregulation of calprotectin metabolism, which mainly presented with the features of chronic microcytic anemia and inflammation. we suggest that bone marrow transplantation could be the best clinical intervention for this new disease. sequential autologous peripheral blood stem cell transplantation with beam conditioning as salvage treatment for refractory high-grade lymphoma e. van hul, a. gadisseur, e. steel, w. schroyens, a. van de velde, z.n. berneman antwerp university hospital (edegem-antwerp, b) t( ; ) mature b-cell (burkitt's) lymphoma/leukaemia (bl) is classified as one entity in the world health organisation (who) classification. bl is a poor-risk, aggressive non-hodgkin lymphoma. despite significant improvements in the treatment of bl, outcomes of adults are generally inferior to those of children. further intensification of the chemotherapy regimens and the inclusion of up-front, high-dose therapy and autologous peripheral blood stem cell transplantation (asct) has significantly improved the duration of response and survival. strategies to improve survival in these poor-risk patients also include sequential asct, and asct followed by non-myeloablative allogeneic transplantation. we present a -year old male who was diagnosed with bl with a double translocation t( , ) and t( , ). he was in remission after cycle of the hovon study protocol (prednisone, vincristin, daunorubicin, asparaginase) but relapsed after the second cycle (ara-c, mitoxantrone). he was then treated according to the hoelzer protocol but after initial good response proved progressive after the second block, with increasing abdominal mass, acute renal failure and metabolic encephalopathy. salvage therapy was initiated with an autologous peripheral blood stem cell transplantation (pbsct) with beam (carmustine, cytarabine, etoposide, melphalan) conditioning resulting in a very good partial remission, which was then consolidated at day + by a second autologous pbsct after beam conditioning. he was planned for an haploidentical allogeneic pbsct according to the perugia protocol on day + after the second pbsct but died unexpectedly of complications of an acute gastrointestinal bleeding before the conditioning (tbi, thiotepa, fludarabin, atg) could be started. we present what appears to be the first reported case of tandem asct with beam conditioning in an adult patient with burkitt's lymphoma/leukaemia . this intensive therapy proved feasible and relatively well tolerated, certainly in view of the bad condition of the patient at the start of the first conditioning. the use of palifermin (keratinocyte growth factor) in the future could further increase the tolerability of this regimen. beam has been proved to be a highly effective treatment in lymphoma but provokes a severe mucositis. nevertheless, tandem beam and autologous pbsct should be further developed as a salvage regimen in the treatment of patients with poor-risk aggressive lymphomas. an association between anbioimmunoblastic t-cell lymphoma and hcv infection: therapeutic difficulties g. mihailov, p. ganeva, n. vasileva national center of haematology and transplantation (sofia, bg) angioimmunoblastic t cell lymphoma (ail) is a rare lymphoproliferative disorder characterized by systemic lymphadenopathy, hepatosplenomegaly, fever, liability to s infections, skin eruption, polyclonal hypergammaglobulinaemia, hemolytic anaemia. clinicaly the disease runs a fatal course in the majority of patients even after multiagent chemotherapy, interpheron á, cyclosporine a, corticosteroids, danazol and recently purine analogues. fewer than % of patients survive years after diagnosis. highdose chemotherapy (hdct) followed by autologous bone marrow transplantation represents a promising new treatment modality for patients with this type of lymphoma. we present a case of year-old woman with association of ail and hcv+ infection with high replication of virus and interesting clinical course of her disease (cns infiltration and complication with bacterial meningitis). it is widely thought, but not yet explained that there might be a pathogenetic link between the infection of hepatitis c virus and onset of b non hodgkin's lymphoma (nhl). in our case we have association with t cell lymphoma. we discuss our treatment difficulties ( courses imvp- , course for brain type lymphoma), following by fludarabine. there is an evidence that ail is susceptible to hdt and autologous stem cell transplantationshuold be considered to the patient. recent data suggest that high dose chemotherapy (hdct) and autologous stem cell transplantation (asct) may be of benefit in patients with aids-related lymphoma (arl). we report on a patient with refractory arl successfully salvaged by hdct and asct. in august a -year-old man presented with a rightlower quadrant abdominal mass. he was known to be hivpositive since (cdc category c ). highly active antiretroviral therapy (haart) was initiated in . his latest cd cell counts was /µl and his hiv-load < copies/ml. abdominal ct-scanning confirmed a lesion of x cm in greatest dimension localized in the right iliac fossa. the spleen was enlarged at . x . cm. a biopsy revealed a stage iia diffuse large b-cell lymphoma. from august to december the patient received courses chop resulting in a partial remission. because a pet-scan indicated residual active lymphoma more courses r-choep (rituximab, cyclophosphamide, doxorubicin, vincristine, etoposide, prednisone) were applied and a complete remission (cr) was achieved. months later he relapsed with an isolated tumor localized in the right iliac fossa. the lymphoma increased in size under courses of bendamustine/rituximab. in july the patient received courses of cisplatin/cytarabine/dexamethasone (dhap) supported by g-csf. a sufficient number of peripheral blood stem cells were harvested after the st cycle ( . x /kg cd + cells). the patient experienced stable disease and received another course ifosfamide/etoposide/epirubicin (iev). however, progressive disease caused an incomplete paresis of his right leg. in september hdct (beam) was administered followed by asct ( . x /kg cd + cells). haart was continued throughout the transplant period with the hiv viral load remaining negative. the patient developed neutropenic fever, a central venous catheter-related infection (koagulase-negative staphylococci), bacteremia (klebsiella pneumonia) and toxic enteritis who grade . broad spectrum antimicrobial therapy was given for weeks. the granulocyte count reached . x /l on day + and platelets x /l on day . a ct-scan performed weeks after asct showed a partial remission. moreover, the paresis of his right leg has disappeared. the patient is doing well and currently undergoes consolidating radiotherapy to the initial tumor bulk. hdct and asct may be of benefit in selected cases of refractory arl. angioimmunoblastic t-cell lymphoma: treatment in two cases a. lópez de la guía, m. martin-salces, a. kerguelén, r. de paz, t. cobo, d. hernandez, j. g-bustos, m. canales, f. hernandez-navarro h.u. la paz (madrid, e) introduction: angioimmunoblastic t-cell-lymphoma (ail) is one of the mature t-cell neoplasm defined in the real-and who-classifications. although patients with angioimmunoblastic t-cell lymphoma (ail) have a poor prognosis with conventional treatment , there are no generally accepted treatment guidelines of proven effectiveness because of low frequency. in that way, it would be necessary to determinate new lineage-treatment to improve unfortunately evolution. methods: between and we reported the good development with high-dose chemotherapy (hdct) and autologous haematopoietic stem cell transplantation in two patients with refractory in our centre. results: the age at transplantation was and years-old respectively. treatment prior to bone marrow transplantation in one case was initially prednisone alone and cladribine, cyclofosfamide and prednisone for cycles, and the other one was chemotherapy based of increased dose of schedule epoch; in this case was necessary secondary treatment with ifosfamide and etoposide ife for cycles. cd + selected autologous peripheral blood stem cell transplantation was given like third line of treatment in the two cases. the regimen for the mobilization of peripheral blood stem cells (pbsc) included ifosfamide, etoposide and g-csf. in one case the median yield of pbsc was , x cd +cells/kg and . x cd +cells/kg and the other one was , x cd +cells/kg and . x cd +cells/kg. the conditioning treatment consisted of beam regimen. there was none treatment-related death. post-taspe complications were herpes zoster infection and biclonal gammapathy igg kappa and igg lambda in one case, and no evidence of acute complications in the second patient. the patients remain in complete remission after a following time of and months respectively and there is no evidence of relapsed. conclusions: our cases confirm previous experience that ail is susceptible to high-dose chemotherapy and cd + selected autologous peripheral blood stem cell transplantation, but more cases and longer observation time as well as better selection of patients with refractory ail would be necessary to determinate the standard treatment. ( )imperial college (london, uk) introduction: we describe the novel use of beam campath conditioning in an autologous setting. we used alemtuzumab ( campath h ) in combination with beam conditioning in two patients who underwent autologous peripheral blood stem cell transplantation for aggressive t cell lymphoma. case a: a year old lady of carribean origin presented with htlv- driven atll. she was treated with courses of chop + dacluzimab with good pr. she received dhap salvage therapy with stem cell collection followed by a beam campath h autograft. she developed cmv reactivation treated with valganciclovir. her day + re-staging showed cr with significant reduction in htlv- proviral load from copies to . copies/ pbmcs. case b: a year old south american presented with massive splenomegaly & pancytopenia. he was diagnosed with gd hepatosplenic lymphoma and was started on chop chemotherapy regimen with initial response followed by progression after cycles. he received dhap chemotherapy followed by autologous peripheral blood stem cell transplant with beam campath h conditioning. post transplant, he developed cmv reactivation managed with valganciclovir. he received splenic irradiation weeks post transplant and at months post transplant, he remains in complete remission. role of alemtuzumab (campath h)-campath h targets all lymphocytes expressing cd . this includes gd t cells. htlv- predominantly infects cd + t cells ( % to % of the proviral load). the expression of cd on htlv- infected cells has not been studied in humans but in nod/scid mice developing htlv- positive tumours, the 'leukemic' cells express cd to a high level. conclusion: the use of beam campath h conditioning regimen in an autologous setting is a novel approach in selected patients with aggressive t cell lymphomas. it merits further investigation. unrelated bone marrow transplantation in a child with high-risk anaplastic large cell lymphoma: case report g. tezcan, v. hazar, v. uygun, a. kupesiz, a. yesilipek akdeniz university school of medicine (antalya, tr) the use of allogeneic stem cell transplantation in non-hodgkin lymphoma patients is not yet clearly defined, especially in children and adolescents. for patients who are in need of bmt, to have a chance for human leucocyte antigen (hla) identical sibling to serve as an allogeneic donor is only %.. for others, transplant from a matched unrelated donor (mud) is an alternative option but with higher risk for gvhd and graft rejection as well as infectious complications and organ toxicity. here, we describe a case of alcl treated with unrelated cord blood transplantation. year old boy admitted with the complaint of mass in right medial thigh region was diagnosed as alcl and chemotherapy was started. at the beginning of second course, because of progression, chemotherapy was changed and daclizumab was added. by the beginning of third chemotherapy course, he was in remission and chemotherapy was continued while an unrelated donor was being searched since hla matched sibling donor was not available. at the end of forth course, one antigen mismatched cord blood was found and following marrow recovery, he was referred to our center for stem cell transplantation while he had been still in remission. the patient was conditioned with total body irradiation ( total cgy) given in six fractioned doses in association with etopside ( mg/kg) and cyclophosphamide ( mg/kg). he was infused with . x /kg mnc and . x /kg cd + cells. cyclosporine, metotrexate and atg were used for gvhd prophylaxis. engraftment was achieved for neutrophil and thrombocyte on posttransplant day and , respectively. on posttransplant day , venooclusive disease developed and it was treated with defibrotide. he has not been experienced any sign of gvhd and he has been still in remission by the end of posttransplant three months. he was the first pediatric alcl case to our knowledge who was treated with unrelated cord blood transplantation. the increasing number of volunteer stem cell donors and although in very limited cases successful results published in the literature regarding the unrelated stem cell transplantation in this group of patients, make this therapeutic option acceptable in the treatment of high risk or treatment failure patients although more data and long term follow up in larger series are needed. high-dose chemotherapy and autologous stem cell transplantation for malignant lymphoma: influence of disease status at time of transplant s. genadieva-stavrik, l. cevreska, a. pivkova, z. stojanoski, o. karanfilski, b. georgievski clinical center (skopje, mk) high-dose chemotherapy with autologous stem cell transplantation (asct) could improve the survival of patients (pts) with malignant lymphoma, but the optimal time of asct is still not clear. this therapeutic method is considered as the optimal strategy in high risk patients in remission and relapsed malignant lymphoma patients. however the potential role in chemoresistent disease and primary refractory relapse is still controversial and optimal timing and indication is still to be assessed. methods: we analyzed consecutive adult patients with malignant lymphoma who underwent asct in a period - . m.hodgkin (hd) was diagnosed in pts, non-hodgkin lymphoma (nhl) in pts. the graft was non-purging, peripheral stem cell. the median follow-up was months (from to months). in hd group the conditioning regimen was beam in all cases. status at asct was complete remission (cr) in cases, chemo sensitive relapse in cases and primer refractory disease in cases. patients in complete remission are patients with adverse prognostic factors for hodgkin disease. in nhl group the conditioning regimen was beam in cases, and other chemotherapeutic regiment in cases. high grade histology was diagnosed in all patients, diffuse large bcell pts, anaplastic large t-cell , lymphoblastic pts. status at asct was complete remission (cr) in cases, chemosensitive relapse in cases, and refractory disease in cases. according to the international prognostic index (ipi) which is validated scoring system predictive of survival in various types of de novo aggressive non-hodgkin lymphoma patients with cr has been evaluated as high risk patients. results: permanent complete remission was achieved in out of pts ( %) with cr at asct in out of pts ( %) with chemosensitive relapse. complete remission could not be achieved in refractory disease. conclusion: the retrospective analysis of our data showed that the asct in patients with malignant lymphoma is an effective salvage therapy. status of the disease at the time of asct is prognostic factors which strongly influenced the outcome of asct. the study suggests that the best results are achieved if asct is performed in complete remission compared with refractory/relapsed disease. patients with malignant lymphoma and initial high risk prognostic factors should be transplanted in first complete remission. autologous stem cell transplantation after radioimmuntherapy with ibritunomab-tiuxetan (zevalin) followed by high-dose cyclophosphamide is feasible in lymphoma patients with end-stage renal disease w. stein, f. gottschalk, o. knigge, u. aurich, m. wernicke, m. kiehl clinic frankfurt/oder (frankfurt/oder, d) we report on a year old male patient suffering from follicular lymphoma grade ii, stage iiia, first diagnosed in . initial therapy consists of cycles r-chop (rituximab, cyclophosphamide, doxorubicine, vincristin, and prednisolone) with partial response. in april patient became symptomatic with abdominal pain and a ct scan demonstrates enlarged lymph nodes paraaortocaval and a left hydronephrosis necessitating ureter splint. after verification of lymphoma relapse two cycles of dexabeam (dexamethasone, bcnu, etoposide, ara-c, melphalan) chemotherapy were initiated followed by stem cell harvest and as lymphoma shows progression we added one cycle of r-dhap (rituximab, dexamethasone, high dose ara-c, cisplatin) again without any s response. due to further lymphoma progression patient became haemodialysis dependent as creatinine level increase to µmol/l and urea level to . mmol/l. furthermore, he suffers from oliguria, ascites and severe edema of the legs, scrotum, and abdomen. as no standard chemotherapy with a clear chance of response was feasible we decided to start a high dose chemotherapy followed by autologuous stem cell transplantation. conditioning regiment consist of radioimmunotherapy with ibritunomab-tiutexan ( , gbq) days prior to high dose cyclophosphamide ( mg/kgbw/day) on days - and - . on day , x cd + cells per kg bw were given. platelet engraftment was observed on day + ( . /µl) and day + ( . /µl), respectively, as neutrophil exceed /µl on day + . patient did not show any problems associated with either radio-chemotherapy or neutropenia. as edema and ascites disappear during neutropenic phase kidney function resolve weeks post transplant. mrt scan shows a very good partial response with residual lymph nodes paraaortocaval. from this case we conclude that in refractory follicular lymphoma radioimmunochemotherapy followed by high dose cyclophosphamide as conditioning regiment even in the case of renal failure necessitating haemodialysis is feasible and highly effective. twenty patients (male: female= : , ages ranging - with a median of ) received high dose chemotherapy followed by peripheral blood stem cell transplant (pbcst) at inha university hospital. their diseases consisted of diffuse large b cell lymphoma (dlbcl, n= ), peripheral t cell lymphoma (ptcl, n= ), lymphoblastic lymphoma (lbl, n= ), anaplastic large cell lymphoma (n= ), burkitt lymphoma (n= ), nasal type extranodal nk/t cell lymphoma (n= ), mycosis fungoides (mf, n= ), and nodal marginal zone lymphoma (mzl, n= ). most of them were in stage via (n= ) or vib (n= ), and others were in stage ie (n= ), bulky iia/bulky iiae (n= ), or iiia (n= ). three patients had disease extended to cns at the very beginning of the treatment. chemotherapy including chop, r-chop, promace/cytabom, copblam-v, dhap, high dose methotrexate, or regimens for acute lymphoblastic leukemia (patients were exposed to a median of courses of chemotherapy ranging - ) yielded cr (n= ), pr (n= ), cr (n= ), cr (n= ), and pd (n= ) right before pbsct. a total of ( allogeneic and autologous) pbsct were performed for patients. allogeneic pbsct was carried out in case of disseminated mf, recurrent ptcl after autologous pbsct, and recurrent ptcl in cr after tandem autologous pbsct. a patient with lbl received double autologous pbsct. the conditioning regimen was cbv (cyclophosphamide,bcnu, etoposide) for most autologous pbsct, cytbi (cyclophosphamide and total body irradiation) for mf, fludarabine based chemotherapy in other allogeneic settings. radiotherapy was given before or after pbsct in patients (brain in , abdomen in , mediastinum in , and nasal cavity in ). a fatal veno-occlusive disease developed in mf patient who died even after orthotopic liver transplant. fatal septicemia in patients at immediate post-pbsct period hampered proper evaluation of the treatment efficacy. a median disease free survival duration was months (range ~ +), and overall survival duration . months (range ~ +). all the patients died of disease who had metastatic disease in their brains. as of this writing, of ( %) are alive disease free at a medial of months (range ~ ). it is of note that, among them, a ptcl patient who received triple pbsct is alive disease free at months post-transplant, a patient with dlbcl in cr at months, a patient with disseminated mzl in cr at months, and a patient with ptcl in cr at months. a. mousavi, s. samiee, m. iravani, b. bahar, m. jahani, k. ali moghaddam, a. khodanbandeh, i. bibordi, a. ghavamzadeh horc (tehran, ir) the non-hodgkin's lymphomas (nhls) are cancers of the cells that populate lymph nodes. it is classified according to its histology, its immunophenotype, cytogenetic and molecular biology. most nhls are cancers of b-lymphocytes. although some of patients with nhl are cured with chemotherapy with or without radiotherapy, the ones who relapse and those with primary refractory disease have poor outcomes with salvage regimen. over the past years, several clinical trials using high dose chemotherapy or chemoradiotherapy with autologous stem cell transplantation or allogeneic stem cell transplantation in this setting have been reported. approximately % of patients appear to be cured using this approach. high dose therapy/autologous stem cell transplantation is standard therapy in two scenarios, in relapsed or refractory non-hodgkin's lymphoma and in patients with refractory aggressive lymphoma whose disease is shrinking with second-line chemotherapy. here we report patients with nhl, who have undergone high-dose chemotherapy (hdct) followed by hematopoietic cell transplantation (hct). of all the patients ( . %) were male, and ( . %) were female. the median age was years old; with minimum and maximum of and y/o respectively. the majority of patients who had immunophenotype study were diagnosed with b-cell lymphoma ( . %). patients ( . %) received autologous stem cell transplantation and ones ( . %) had peripheral blood as graft type. before (hct) . % of patients were in first complete remission. (including cr with salvage therapy) the conditioning regimen for the majority of patients ( . %) was ccnu, etoposide, ara-c (cytarabin) and melphalan. the median duration of hospitalization for autologous transplanted patients was days which was days for allogeneic transplanted ones. transplant mortality rate in the first days was %. the median follow-up duration was days, with minimum and maximum of and days respectively and during this period the overall survival (os) is . % and the disease free survival rate (dfs) was . %. zevalin therapy of a non-hodgkin relapsed lymphoma patient following an autologous peripheral stem cell transplantation l. rejto ( ) the yttrium- ( y) -labelled ibritumomab tiuxetan (zevalin, idec-biogen, san diego ca) is an accepted therapy for relapsed, or therapy-refracter b-cell non-hodgin lymphomas, but it is not officialy recommended in patients who have failed an autologous stem-cell transplant. patients with recurrent lymphoma following an autologous transplant have limited treatment options. cases so far only have been described in the literature whose relapse following autologous peripheral stem-cell transplantation (apsct) has been treated with zevalin. in the present paper the authors discuss the case of a year-old male patient, who underwent lymph-node biopsy due to generalised lymphadenomegaly. the histology test proved positive for cd follicular lymphoma. following an cycle chop (cyclophosphamide , doxorubicine, oncovin, prednison), a cycle fnd (fludara, mitoxantron, dexamethasone), later a cycle dhap (dexamethasone, high-dose ara-c, cisplatin) therapy, autologous peripheral stem-cell transplantation was performed. six months later due to a relapse a cycle r-cepp (rituximab, cyclophosphamide, etoposide, prednison, procarbasine), then a cycle hyper-cvad therapy was applied. the patient (had no compatible sibling) proved to be therapy-refractory, therefore zevalin treatment was started (after a preparatory rituximab therapy mbq zevalin was applied). the zevalin treatment did not caused any unusually serious side effect. during the months since the start of the zevalin therapy the patient has been in a complete haematological remission. in nhl, in case of a relapse following apsct zevalin therapy has proved to be a good alternative. the use cd- monoclonal antibody in the treatment of bcell non-hodkin lymphoma with autologous stem cell transplantation o. tarabar, l. tukic, d. stamatovic, z. tatomirovic, b. balint, b. cikota, z. magic medical military academy (belgrade, cs) introduction: recent trials have shown that cd- monoclonal antibody, rituximab (r) may be effectively associated with autologous stem cell transplantation (asct) in the treatment b cell non-hodgkin's lymphoma (nhl). aim: to analyze the efficacy of the incorporation of r at different steps of autologous transplantation (r-asct) programs in patients (pts) with high-risk nhl. methods: between march to december r+asct was applied for the treatment of pts with b cell nhl ( low grade; diffuse large cell lymphoma -dlcl). asct were performed after chop induction chemotherapy (ct) with r (r-chopx ) in pts or without r in pts. in the time of asct complete remission (cr) had pts and others pts had partial remission (pr). regimen mobilization were g-csf with cy ± vp- in pts, eshap ct in pt, r+cy in and megar-choep in pt. a single infusion of r ( mg/m² ) used as in vivo purging days prior cy or day of therapy megar-choep. the average number of colected mobilized cd + cells was , x /kg bm (range , - , ). all pts received cvb conditioning regimen. the posttransplant immunotherapy consisted of a single dose r every months (m) started m following asct in pts. sequential monitoring of minimal residual disease (mrd) during and after treatment was performed by pcr. five of seven pts are available for mrd. results: at a median follow-up of months ( - ), pts are alive. after r+asct treatment, out of available pts became pcr negativ (low grade lymphoma- ; dlcl- ). four pts with dlcl are still in complete remission (cr) and two of them in molecular remission (mr) and months (m). the both pts with low grade lymphoma relapsed and m after transplantation, the one of them never attained pcr negativity and second pt reverted to pcr positivity after m. this therapy is well tolerated, with no adverse effects on hematological recovery of incidence of infections. conclusion: this therapy was effective in the subset of pts with high risk dlcl. r+asct treatment is able to eliminate mrd, whereas pcr positivity is associated with a high risk of relapse. double syngeneic transplantation in plasma cell leukaemia c. barrenetxea, m. callis, s. iraheta, e. sanchez, v. pons vall d´hebron (barcelona, e) introduction: plasma cell leukemia (pcl) is a rare disorder, characterized by circulating clonal plasma cell. it accounts for less than % of all plasma cell dyscrasias and has a fatal prognosis. it can be primary or secondary, when there was a previously diagnosed plasma cell dyscrasia. the median survival is - moths in the first case and moths in the second. case: we present a years old man, diagnosed in november , with multiple myeloma iga kappa, bence jones +, that presented weight loss, retinal hemorrhages, respiratory distress, hepatomegaly, splenomegaly, osteolytic lesions, the cariotype showed hyperploid, chromosome monosomy, translocations t ( , ) and t ( , ). first, he was treated with tree cycles of a drug's combination with melphalan, carmustine, vincristine and dexametasone with no response, therefore, was changed to cyclophosphamide, adriamycin, vincristine methotrexate y citarabine. after two cycles, the patient got complete remission. the patient had a twin brother, and we decided, to do a double transplantion to consolidate the response. the first transplantation was condicionated with carmustine, etoposide, citarabine and melphalan (beam), and the second with cyclophosphamide and total body irradiation; the patient remained in complete remission. eight months after second transplant was admitted to the hospital with disorientation, bradipsiquia, headache, and sensoriomotor loss of lower extremity. laboratory examination showed differential count of leukocytes, haemoglobin and platelets were normal, ldh increase, absence of monoclonal gammophaty in blood and urine by inmunofixation, and the brain's computerized tomography showed multiple intraparenchymatous lesions, with peripheral edema, in both cerebral hemispheres, confirmed by magnetic nuclear resonance, all of that suggested a neoplastic disease. these lesions were biopsied with the result of multiple myeloma lambda. the patient died one day after biopsy because intracranial hypertension. conclusion: plasma cell disease has poor prognosis, and transplantation could be a good option for some patients, but in our case we only achieve to extend life a few months. delayed engraftment following autologous stem cell transplantation: risk factors apart from stem cell dose r. fineman, n. haddad, t. zuckerman, i. avivi, t. faibish, m. markovitz, l. dan, d. abu zemach, l. akria, j.m. rowe rambam medical center (haifa, il) background: the use of mobilized peripheral blood stem cells results in prompt engraftment of all three cell lines in both autologous and allogeneic transplantation compared to bone marrow. the stem cell (cd +) dose is known to be a major factor for bone marrow recovery. although reinfusion of ≥ x cd + cells/kg is considered more than adequate in terms of rapid engraftment, there are still patients who engraft after day , despite sufficient cd + cell dose. we looked for other factors contributing to delayed engraftment apart from the stem cell dose. methods: retrospective data on lymphoma and multiple myeloma patients with delayed engraftment (absolute neutrophil count > /ul after day ) after autologous transplantation were analyzed. factors including stem cell dose, infectious complications and outcome were evaluated. median age at hsct: ( - ), pts > . diagnosis: mds= , saml= . twenty-seven pts received a median of ( - ) chemo cycles before hsct; pts received only red cells transfusions allo, haplo, mud). hsct source: pbsc %) pts; , , and after auto, allo, haplo and mud, respectively. causes of death: infection , disease progression , ards , other . all pts evaluable for disease status after hsct (n°= ) were in cr at first marrow examination median os from hsct is ( - ), median. at last update pts ( . %) are alive in cr after a median follow-up of days haplo and mud, respectively. r results of high-dose chemotherapy followed by autologous stem cell transplantation in children with advanced neuroblastoma in paediatric bone marrow transplant centres in poland from wojcik ( ), k. kalwak ( ), e. gorczynska ( ), d. turkiewicz ( ), a. dyla ( ) pl) purpose: postransplant morbidity and clinical outcome in children with advanced neuroblastoma (nbl) who underwent high dose chemotheraphy followed by autologous stem cell transplantation were investigated. patients: the total children with stage iii/iv neuroblastoma treated in five bone marrow transplantation units in poland from to were analysed. median age of childen was , years (range - , years). children were transplanted in complete/partial remission (cr/pr), in patients megachemotherapy was a part of the treatment of relapse (> cr/pr). aphaeresis was done in patients. bone marrow was collected in pts; bone marrow and stem cells were transplanted in patients melfalan + etoposide + carbo in pts.; melphalan in pts.;thiotepa + ctx + carbo in patient and thiotepa + topotecan + carbo in patient. results: ( , %) children are still alive at median observation time months. ( , %) children died dfs) is , ; expected -year os and dfs is , and , respectively. os at months in the group of children transplanted in cr/pr was , ; dfs , . os and dfs expected at years were , and , respectively. in children transplanted > cr/pr estimated os was , , dfs , at months and expected -year os and dfs were , and , respectively. conclusions: megachemotherapy followed by autohsct in patients with advanced neuroblastoma has not many adverse effects. estimated -year os and dfs are higher in the group of children results: a total of patients underwent an autologous stem cell transplantation for lymphoma and multiple myeloma at the rambam medical center between the years and ( and respectively). indications for transplantation included a chemosensitive relapse in lymphoma high risk patients and rarely also for refractory patients. in myeloma auto transplants were performed for patients achieving good response upon initial chemotherapy. patients with lymphoma received the beac/beam conditioning and patients with myeloma were given melphalan mg/m². patients showed a delayed engraftment, of them were patients with multiple myeloma and only with lymphoma. median time to engraftment was days (range - ). in patients the cd + cell dose was > x /kg and in one patient it was x /kg. patients died of severe infections, one with lymphoma on day without engraftment, the other with myeloma although engrafted on day . the outcome of other patients was uncomplicated. there were no detectable differences in the clinical course or toxicity among those who engrafted eraly or the late engrafters. conclusion: the use of mobilized peripheral stem cells shortens time to engraftment. using high doses of stem cells is safe, but there remains a significant risk of delayed engraftment in % in myeloma compared with < % in lymphoma patients. considering the homogenicity of the groups and the high stem cell dose reinfused, it is likely that bone marrow microenvironment, known to be impaired in multiple myeloma, has a role in fascilitating engraftment. a single-centre experience in autologous stem cell transplantation for patients with multiple myeloma h. kasparu, j. könig, o. krieger, m. girschikofsky, d. lutz elisabethinen hospital (linz, a) from oct. to aug. we performed autologous stem cell transplantations (abct) in multiple myeloma (mm)-patients (age (median: - ) years; female: , male: ). following conventional chemotherapy patients (transplanted between - or patients not eligible for a tandem transplantation concept due to late infections (n= ) or toxic side effects (carditoxicity ( ), neurotoxicity ( ), dermatitis ( ), smds ( )) underwent a single course of abct and patients multiple courses of transplantations ( double and tripple abct). no significant differences between both groups were seen according to age, sex distribution or stage of disease at time of abct. more patients who relapsed after conventional treatment ( vs. pts) were included in the single abct group. the conditioning chemotherapy consisted of mel mg/m² for single and double abct and mg/m² for tripple abct. all patients were transplanted with peripheral stem cells. the time to granulocyte recovery > , g/l lasted - days (median: ) and to platelet recovery > g/l - days ( median: ) without a difference between the consecutive numbers of transplantation. all patients but one in each group responded to transplantation: cr pts., pr pts., trm pt. (single abct group) and cr pts., pr pts., failure pt.( multiple abct group), resp. in the single abct group pts. ( %) relapsed after - months (med: mo) , in the multiple abct group pts. ( %) relapsed so far after - months (med: months). the median observation time is shorter in the multiple abct group ( vs. months). the median pfs and os in the single abct group is and months, resp. in the multiple abct group median pfs lasted months, the median os is not reached yet. both transplantations were well tolerated even in older patients. there is a trend to longer pfs in patients after multiple abct, but due to different observation periods no final conclusions can be drawn concerning os. autologous haematopoietic stem cell transplantation for the treatment of multiple myeloma o. tarabar, l. tukic, d. stamatovic, m. elez, v. glavicic, l. simic, b. balint, s. marjanovic medical military academy (belgrade, cs) high dose therapy with autologous stem cell transplantation (asct) is the treatment of choice for patients (pts) with multiple myeloma (mm). we report here our centre experience in pts with mm who have undergone asct. between december and september pts ( m/ f) with a median age of years (range - ) were transplanted. most pts ( %) were in stage iii (durie-salmon). before transplantation, pts have received line of chemotherapy ( - cycles vad). at the time of autograft % pts were responders (partial remission or very-good partial remission) and the others had minor response/refractory disease. all pts received peripheral blood stem cell support after conditioning with melphalan ( pts) or melphalan associated to cyclophosphamide and busulfan (by/cy +m). three months (m) after asct pts received interferon ( mu/s.c. t.i.w) and pts thalidomide ( - mg daily) as maintenance therapy. after transplantation a complete (cr) or very-good partial response was achieved in / pts ( %); all other treated pts experienced a reduction of m-component > %. with the median follow up of , m (range - ), % pts were alive. to date, in cr are still pts with the median duration of remission from asct of m. seven pts relapsed or progressed during to m after asct. two pts died from transplant related complication. asct is safe and effective procedure not only in chemotherapy sensitive pts with mm but also in resistant cases. our date confirm that asct is the current gold standard therapy for many pts with mm. dose intensity and efficacy of treatment in patients with multiple myeloma e. darskaya, s. bondarenko, a. smirnova, b. afanassyev pavlov state medical university (st. petersburg, rus) mm patients were included in our study. induction therapy was vad-d-d or idad-d-d. patients with mm were undegone intensification dexabeam. patients received cycle dexabeam. was sensitive to the -st line therapy, dexabeam resulted in ( %) cr, ( %) ncr and ( %) pd. patients was unsensitive and the resulte was ( %) cr, ( %) pr, ( %) sd, ( %) na. cycles dexabeam received patients, of them were in pr after the first line therapy, -in sd, -in progression of disease. ( %) sensitive patients achieved ncr, patients, resistant to the first line therapy achieved ( %) cr, ( %) pr, ( %) -sd. patients of mm underwent asct, patients ( , %) after conditioning regimen of melphalan mg/m², patients ( %) -melphalan mg/m², patients ( , %) - mg/m². patients with mm received tandem asct. -years dfs in sensitive to induction and intensification therapy patients was %, dfs in group of the patients with progression or relapse before asct was no longer that . year. p = . . the -years duration of a plateau phase of the patients, sensitive to the first line therapy and intensification, after -st asct was %, the duration of a plateau phase of the patients with progression before asct, was no longer that . year. p = . . the -years duration of a plateau phase of the patients, who received melphalan mg/m² as conditioning regimen, was %. in patients, received mg/m² melphalan and less it was no longer that . years.s years dfs in group of the patients with tandem asct was % in comparison with % in group of the patients with single asct. p = . . the conclusion: our preliminary results are that the increasing of dose intensity improves the efficacy of treatment of patients with multiple myeloma. successful myeloablative allogeneic haemopoietic stem cell transplantation in a patient with end-stage renal failure on haemodialysis a. alfred ( ) introduction: end stage renal failure (esrf) has conventionally been considered a relative contraindication to hsct. although increasing numbers of patients undergoing autologous procedures with hd are being reported, documented experience in the allogeneic hsct setting remains extremely limited. to the best of our knowledge only three previous case reports have been published. case report: a year old male, treated with regular hd for esrf from , was diagnosed with myelodysplastic syndrome associated with monosomy (ipss int- ) in . after extensive counselling with the patient, careful consideration of donor issues, and collaboration between bmt and renal teams, a decision was made to proceed with allogeneic hsct with curative intent. in april , the patient underwent myeloablative conditioning with total body irradiation gy and cyclophosphamide mg/kg followed by transfusion of g-csf mobilised allogeneic pbsc from his hla and abo-matched sister (cd cell dose = . x /kg). recipient and donor cmv serology was negative. during conditioning, hd was performed on a daily basis to optimise biochemistry. the patient was closely monitored for cyclophosphamide cardiotoxicity with ecg and echocardiography. oral mesna was used to prevent haemorrhagic cystitis. hd was subsequently maintained at the regular thrice weekly schedule. graft-versus-host disease (gvhd) prophylaxis was ciclosporin and methylprednisolone. ciclosporin levels were little affected by hd and there were no significant problems maintaining the therapeutic range. otherwise careful consideration was given to drug dosing in relation to hd. regimen related toxicity was no more than expected in a patient with normal renal function. engraftment was prompt and discharge was on day + . mild acute and chronic ghvd has been managed with additional corticosteroids. there have been two inpatient re-admissions for post-transplant complications. bone marrow examination at three months post hsct confirmed trilineage engraftment and % female karyotype with no evidence of monosomy . the patient is presently stable, in remission and on reducing immunosuppression at over months post-hsct. conclusion: this case highlights the feasibility of myeloablative conditioning and allogeneic hsct in carefully selected patients with esrf on hd. ethical considerations should incorporate the additional impact on the donor of a potential future histocompatible living donor renal transplantation. liver complications in hematopoietic transplantation (hct) setting may be life threatening, and hepatitis b virus (hbv) infection increases the risk of hepatic complications in patients undergoing hct. we describe a year old white women who was treated years before with chemotherapy for non-hodgkin disease achieving complete remission. she acquired hbv infection because of a blood red cell transfusion. five years after complete remission she developed anemia, piastrinopenia and leucopenia. bone marrow biopsy histology excluded lymphoma relapse but revealed multilineage myelodisplasia with excess of blasts. cytogenetic examination detected qdeletion. according to ipss score myelodisplasia was classified at high risk. since an high viral b load ( copies/ml), despite normal alt and ast values, the patient underwent lamivudine prophylaxis. hct was considered and his brother resulted full hla match. he was succesfully mobilized with g-csf. before transplantation viral b load did not decrease so antihepatitis b prophylaxis was changed from lamivudine to adefovir dipivoxil that was also employed as the only antiviral prophylactic treatment along all the procedure. patient underwent reduced conditioning regimen: e.v. busulfan ( , mg/kg weight) plus fludarabine ( mg/m²) and rabbit atg fresenius ( mg/kg).graft versus host disease (gvhd) prophylaxis consisted in cyclosporin and metotrexate. , x /kg cd and , x /kg monocleated were infused. neutrophil engraftment was at + , while platelet engraftment at + . the patient developed steroid sensitive cutaneous grade ii acute gvhd. complete chimerism was achieved at + . at day she developed abrupt alt and ast increase (> iu/l), without bilirubin increase and a concomitant viral b load more than copies/ml. lamivudine was reintroduced and adefovir was continued. one week later transaminases began to decrease and at day both alt and ast were under iu/ml, while viral b load was x and the patient is well at day . a reduced conditioning regimen with fludarabin and e.v. busulfan together with rabbit atg revealed to be safe and efficacious in this year old patient. moreover lamivudine and adefovir association was able to control hbv replication. adefovir alone successfully avoided every other viral infection (hsv, cmv etc.)along transplant procedure and it did not impair engraftment. haematopoietic stem cell transplantation in poor prognosis mds and saml: prolonged patients survival is achievable, but refining of infections management and reduction of treatment-related toxicities is required to improve patients outcome m. tassara, a. crotta, l. camba, f. lunghi, m. marcatti, j. peccatori, m. bregni, f. ciceri, m. bernardi san raffaele scientific institute (milan, i) introduction: allogeneic hematopoietic stem cells transplantation (hsct) is the therapy of choice for poor prognosis mds and saml patients (pts); a hla identical, related (allo) or unrelated (mud), or haploidentical familial (haplo) donor is available for most pts. autologous (auto) hsct is an alternative for pts without a donor. prolonged overall survival (os) and disease free survival (dfs) are reported in a minority of mds/saml pts after hsct. major drawback of allo, haplo and mud hsct is the high incidence conclusions: prolonged os and dfs are achievable with hsct in poor prognosis mds and saml pts, also in the elderly. prevention of pts contamination before hsct, mainly from aspergillus, and reduction of early trm, mainly in the haplo subset of pts, could improve pts survival. trials for primary/secondary anti-fungal prophylaxis are ongoing and reduced-toxicity conditioning regimens are under investigation at our institute. auto in cr is an alternative if hsct from a donor is not feasibile; to reduce the relapse rate after auto an experimental maintenance treatment should be proposed. high-dose chemotherapy and autologous peripheral blood stem cell transplantation followed by a successful extremity sparing surgery in a case of osteosarcoma arising in osteogenesis imperfecta s. ataergin, f. arpaci, k. erler, b. demiralp, a. kaya, a. ozet, m. basbozkurt gata (gulhane) faculty of medicine (ankara,tr) objective: osteosarcoma (os) arising in osteogenesis imperfecta (oi) is reported as only nine proven cases in the english literature; among these one case had a limb sparing surgery, but none underwent a high-dose chemotherapy (hdc) and autologous peripheral blood stem cell transplantation (apbsct). methods: we recently demonstrated the effectiveness of hdc and apbsct in localized osteosarcomas. this case is the first one of os occurring in oi who underwent a hdc and apbsct. results: a- -year male patient had been followed-up since his birth with type i oi and had recurrent traumatic bone fractures. he was admitted to orthopedics clinic with complaints of bone pain, edema and swallowing in right proximal tibia. an incisional bone biopsy revealed conventional osteosarcoma. the stage was iib and two cycles of neoadjuvant chemotherapy consisting with cisplatin mg/m², adriamycin mg/m², ifosfamide . g/m² and mesna . g/m² were administered in three consecutive days, every three weeks, according to our institutional treatment protocol for osteosarcoma. fifteen days later, stem cells were mobilized using g-csf ( µg/kg/day in two doses) and collected by cobe spectra cell separator (cobe bct inc., lakewood, co). the amount of cd + cells was . x /kg. hdc (ifosfamide g/m², carboplatin . g/m², etoposide, . g/m² and mesna on a dose of - % of the total ifosfamide dose were given in dividing doses on six consecutive days) and apbsct were performed thereafter. extremity sparing surgery (wide en-bloc resection and reconstruction prosthesis) was applied after the neutrophil and platelet engraftment. postoperative tumor necrosis rate was % (by histopathology) and the same induction chemotherapy regimen was given for three additional cycles as consolidation chemotherapy. conclusion: the patient is still disease-free with a good functional score and no fracture has occurred in months after the last follow-up. improved survival in neuroblastoma by autologous peripheral blood stem cell transplantation: a single-centre experience h. kook, j.y. kim, h.j. baek, d.k. han, h.r. yi, h.j. kim, t.j. hwang chonnam university hwasun hospital (hwasun, kor) objectives: neuroblastoma is the most common extracranial solid tumor of childhood, and its outcome in advanced diseases has been very poor. in this study, the author evaluated the treatment outcome and prognostic factors in advanced neuroblastoma. methods: the study group comprised of patients who were diagnosed and treated with neuroblastoma at chonnam national university hospital from january, to may, . data were obtained from the retrospective review of the medical records. patients were classified according to the evans group. the conventional treatment including surgery, radiotherapy, pre-and post-operative chemotherapy was given to stage i, ii, and iii patients. for stage iv, relapsed patients, high-dose chemotherapy followed by autologous peripheral blood stem cell transplantation (pbsct) was administered. the chemotherapy consisted of cisplatin, doxorubicin, etoposide, and cyclophosphamide (ccg , ). conditioning for pbsct was modified vamp-tbi(cisplatin, doxorubicin, etoposide, melphalan, and total body irradiation). all patients who completed cytotoxic therapy were then either received no further therapy or treated with -cis-retinoic acid for six months. results: among patients, were males and females. the median age at diagnosis was . months (range, - months). the primary sites were the adrenal glands in patients, followed by retroperitoneum in , and thoracic cavity in . most of the patients were in advanced stages: stage iii in ; stage iv in . autologous pbsct was done in cases. the -year event-free-survival (efs) rate was % in all study patients with % for stage i, % for stage ii/iii, % for stage iv, % of iv-s. in cases with stage iv neuroblastoma, the efs rate at years after diagnosis was better among the patients who underwent autologous pbsct than among the patients who received chemotherapy ( % vs. %; p = . ). also, efs was better in patients who received -cis-retinoic acids after pbsct than those who did not ( % vs. %; p < . ). conclusion: treatment with high dose chemotherapy and autologous pbsct improved efs among children with advanced neuroblastoma. in addition, treatment with -cisretinoic acid was beneficial for patients who underwent transplantation. prospective randomized study is warranted to further improve survival for subset of advanced patients who might fail to current management strategies. key: cord- - iu pr authors: nan title: the th annual meeting of the european society for blood and marrow transplantation: physicians – oral session date: - - journal: bone marrow transplant doi: . /s - - - sha: doc_id: cord_uid: iu pr nan methods: study aim was to evaluate the schedule of ist given in combination with pt-cy as gvhd-prophylaxis post-haplo for acute leukemia (al) and reported to the alwp/ebmt registry. patients were divided into groups: received cyclosporine a-mycofenolate-mofetil(csa-+mmf) initiated at day+ (group- , n= ) or csa +mmf both started at day+ (group- , n= ) and tacrolimus + mmf from day+ (group- , n= ). transplants were performed from - and median follow up is months (range - ). pt-cy was given on day+ and day+ in group- and on day+ and day+ in group- and . results: acute myeloid leukemia (aml) was the most common indication for haplo ( %) and approximately % of patients were transplanted in cr . there were some differences among groups: patients in group- were younger (median age years, p< . ) were transplanted in more recent year ( , p< . ), received more frequently a regimen based on tbf (thiotepa, fludarabine and busulfan) ( %, p< . ) and bone marrow (bm) as source of stem cells ( %, p< . ), with no atg ( %, p< . ). probability of os at years was %, % and %, for the groups, respectively, p= . . probability of lfs and grfs at years were % and %, % and %, % and %, for the groups, respectively, (lfs p= . , grfs p= . . overall the cumulative incidence (ci) of grade ii-iv acute gvhd was %, % and %, for the groups, respectively, p< . , and the ci of chronic gvhd was %, % and %; p= . . the ci of relapse at years was %, % and % (p= . ) and background: patients with acute myeloid leukaemia (aml) often achieve remission but subsequently die of relapse driven by chemotherapy resistant leukemic stem cells (lscs). here we hypothesized that lscs must also escape immunosurveillance to initiate and maintain cancer and investigate the interplay with nkg d, a danger detector expressed by cytotoxic lymphocytes such as natural killer (nk) cells that recognizes stress-induced ligands (nkg dl) of the mic and ulbp protein families on aml cells. methods: de novo aml were stained with antibodies against mica, micb and ulb / / or an nkg d-fc chimeric protein recognizing pan-nkg dl. nkg dl pos and nkg dl neg aml cells sorted from the same patient were analysed in colony forming assays, leukemogenesis assays in nsg mice, by rnaseq, gene expression arrays, qrt-pcr and targeted next generation sequencing. aml cells co-cultured or not with nk cells (control or anti-nkg d pre-treated) were co-stained for additional stem/immunological markers. parp expression was analysed by qrt-pcr and immunoblot, and binding to nkg dl promoters by chromatin immunoprecipitation. parp inhibition (parpi) in aml cells was performed in vitro or in vivo using sirnas or inhibitors (ag- , veliparib) . results: heterogeneous nkg dl expression was detected among leukemic cells of the same patient (fig. a) . interestingly, when compared to nkg dl pos subpopulations, nkg dl neg aml cells isolated from the same patient showed immature morphology, enhanced in vitro clonogenicity ( ± colonies vs. ± , p< . , n= aml patients) and selective abilities to initiate leukemia (nkg dl neg , / , %; nkg dl pos , / , %; p< . , fig. b , n= aml patients) and survive chemotherapy in nsg mice devoid of functional nk cells. in mice, nkg dl neg aml cells generated both nkg dl pos and nkg dl neg progeny of which again only latter was able to induce leukemia in re-transplant assays. similar leukemia-specific mutations were detected in nkg dl neg compared to nkg dl pos aml cells from the same aml but published lsc (fig. c ), hsc and genes stemness score signatures were specifically enriched in nkg dl neg subfractions. nkg dl neg cells enriched for lscs defined by alternative markers (cd + , cd -, gpr + ) but could identify cells with functional and molecular lsc activity also in cd non-expressing aml (n= analyzed patients). nkg dl expression was repressed by parp recruitment at nkg dl promoters. parp inhibition (parpi) induced nkg dl surface expression in lscs and co-treatment with parpi and nk cells (but not with either alone) suppressed leukemogenesis in patient derived xenograft (pdx) models (fig. c ) cotransplanted with nk cells. low nkg dl surface or high parp mrna expression associated with poor outcome in aml patients. furthermore, nkg dl neg and cd + lscs showed reduced expression of other immune stimulatory molecules (e.g. cd , cd , cd , cd ) and different expression of immune or inflammatory response gene signatures (gsea). conclusions: these data indicate that lscs escape nk cell recognition by selectively suppressing the surface expression of nkg dl and other immunostimulatory molecules. absence of nkg dl can identify lscs across genetic aml subtypes (including cd negative amls). this lsc specific mechanism of immune evasion could be overcome by treatment with parp inhibitors, which in conjunction with functional nk cells holds promise to eradicate lscs and promote immune-mediated cure of aml. disclosure: c.l.: sanofi, novartis, otsuka (consultancy); roche (research funding) background: in contrast to imatinib, data on the use of nd and rd generation tyrosin kinase inhibitors (tki) in the treatment of minimal residual disease (mrd), molecular and hematological relapse (mr/hr) after allogeneic stem cell transplantation (sct) in philadelphia chromosome positive (ph+) acute lymphoblastic leukemia (all) are scarce. methods: we performed a retrospective, ebmt registry based analysis, including patients with documented use of nd or rd generation tki given for persisting mrd, mr or hr after allosct in - choice of tki, efficacy, and toxicity of tki and patient outcome were analysed. results: patients (female , male ) were identified, out of which had also received a tki ( % imatinib) before allosct. median age at transplant was years ( - ), % were transplanted in first complete remission (cr ), % of the patients were in molecular cr. conditioning was myeloablative in % and reduced in %, donors were matched siblings ( %), unrelated ( %), haploidentical ( %) and cord blood ( %). after allosct, patients developed hr, mr and had persisting mrd. for treatment, patients received dasatinib (n= ), nilotinib (n= ) and ponatinib (n= ). median interval between diagnosis of persisting mrd or mr/hr and first application of a tki was days, median duration of tki treatment was days (range - ). fifty-eight patients were treated with tki only (dasatinib, n= , nilotinib, n= , ponatinib, n= ) , while received additional treatment such as dli, chemotherapy, or second allosct. main toxicities of dasatinib were effusion, edema, or other pulmonary complaints ( - % of patients) and infections ( %). no particular side effects were reported for nilotinib and ponatinib (no vascular events). dose reduction of tki was required in %. response rates were % (entire cohort) and % (patients receiving tki only). for the entire cohort, -and -year overall survival (os) from first application of tki was % and %. two-year os was comparable in patients treated for persisting mrd/mr and for hr ( % and %). among patients treated with tki only, / -year os was %/ %. rate of cgvhd was % for the whole population and % for the tki alone cohort. conclusions: the use of nd and rd generation tki, given alone or in combination with other therapies for treating persisting mrd, mr or hr after allosct in ph+ all was not associated with increased toxicities. dasatinib was the most frequently used drug. outcome compared favorably with published results on relapse after allosct in ph negative all, suggesting that treatment with tki could improve survival after post-transplant relapse, even when given as single therapy. type of relapse did not influence response rates and outcome. disclosure: nothing to declare multi-state modelling of the interplay between remission-induction chemotherapy and consolidation with allosct in newly diagnosed aml patients reduced rate of relapse in the clofarabine arm, discovering a significant difference between the treatment arms in the hazard of relapse only after allosct (hr . , % ci . - . , p-value = . ), and not before allosct (hr . , . in addition, we found increased nrm in the clofarabine arm before allosct (hr . , % ci . - . , p-value = . ), and not after allosct (hr . , % ci . - . , p-value = . ). these effects are statistically significantly different (interaction test hr . , % ci . - . , p-value = . ). at two years after registration, . % ( % ci . - . ) of the patients in the control arm, and . % ( . - . ) in the clofarabine arm were alive relapse-free in cr without allosct, while . % ( . - . ) and . % ( . - . ) were alive relapse-free after allosct. conclusions: presented results suggest that the rate of relapse after allosct is lower among patients whose induction therapy includes clofarabine. these results could possibly be explained by higher rate of mrd negativity achieved in the clofarabine arm before proceeding to allosct. we also observed a higher nrm rate in cr before allosct in the clofarabine arm, indicating that the favorable effect of clofarabine on relapse may be compromised by toxicity. clinical trial registry: hovon study is registered at netherlands trial registry #ntr disclosure: nothing to declare haploidentical transplant with post-transplant cyclophosphamide for t-cell acute lymphoblastic leukemia: outcome strongly correlates with disease status; a report from the ebmt acute leukemia working party background: partial tandem duplication of mll (mll-ptd) is an infrequent mutation in aml which produces a number gain of ' acetyltransferase domains of kmt a protein as a result of a repeated exon - / gene sequence. mll-ptd leads to a disturbed histone acetylation and upregulation of determined hox genes. mll-ptd aml defines a specific aml entity, distinguishable from cases of aml with mll rearrangement, with a characteristic pattern of co-mutations, including a high association with flt -itd (q-y sun et al., leukemia ) . prognosis of mll-ptd-aml is remarkably poor, with initial chemoresistance and high relapse rate; as a consequence, allogeneic hematopoietic cell transplantation (allohct) in early phase is recommended to overcome its high-risk prognostic impact (jp patel et al., nejm ; v grossmann et al, blood ) . nonetheless, studies focusing on transplant outcome have not been previously addressed. methods: for this purpose, we analysed the outcome of mll-ptd aml adult patients reported to ebmt who had received an allohct from matched related or unrelated donors in cr during the period - . molecular screening of mll-ptd was performed locally, but the presence of this mutation was verified specifically by a focused questionnaire among participating centres. results: overall, we identified patients fulfilling inclusion criteria (median age, . years; / female patients). most patients harboured an intermediate risk cytogenetics ( / ; % of available) and ( / , % of available) patients presented with concomitant flt -itd. donor was a matched sibling in transplants ( %) and an unrelated donor in the remaining cases. conditioning was myeloablative (mac) in procedures ( %) and reduced intensity in ( %). in vivo t-cell depletion was used in ( %) of transplants. at two years, cumulative incidence of relapse (cir) was % ( % ci: - ), and non-relapse mortality (nrm) % ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ; -day acute gvhd of grade iii-iv was % ( - ) and -yr chronic gvhd was % (extensive, %, - ). two-year overall survival (os) was % ( - ), with leukemia-free survival (lfs) and graft-and relapse-free survival (grfs) of % ( - ) and % ( - ) , respectively (figure) . multivariate analysis did not identify any prognostic factor for lfs; notably, presence of flt -itd, conditioning regimen intensity or donor type did not influence outcome. conclusions: these results support the potential clinical benefit of allohct performed in cr in patients with mll-ptd aml, with a decreased relapse risk compared to previously reported series, suggesting the existence of a tangible graft-versus-leukemia effect (gvl) in this setting. disclosure: nothing to declare o safety and efficacy of reduced intensity conditioning regimen combined with anti-thymocyte globulin and post-transplantation cyclophosphamide as graft versus host disease prophylaxis for acute myeloid leukemia background: we aimed to evaluate the safety and efficacy of the use of reduced intensity conditioning regimen (ric) combined with anti-thymocyte globulin (atg) and posttransplant cyclophosphamide (ptcy) for graft versus host disease (gvhd) prophylaxis in patients diagnosed with acute myeloid leukemia (aml). methods: one hundred four adult patients were included. all patients received the same ric regimen including fludarabine ( mg/m /day day - to - ), busulfan ( . kg/ m /day day - and - ), and total body irradiation ( cgy) (day - ) combined with rabbit-atg ( . mg/kg: day - to - ), ptcy ( mg/kg/day: day + ,+ ), and cyclosporine. unmanipulated peripheral blood stem cells were infused. last follow-up was november . median follow-up was months (range - ). results: findings are summarized in figure . one year overall survival (os) progression-free survival (pfs) and non-relapse mortality (nrm) was . % ( % ci . - . ), . % ( % ci . - . ) and . % ( % ci . - . ) respectively. main causes of death were relapse ( %) and infection ( %). three patients had residual disease prior-transplant and they had a significant worst os (p= . ) and rfs (p= . ). patients who had karnofsky performance status (kps) ≤ % had a significantly worse os (p= . ) and pfs (p= . ). the achievement of ≥ % chimerism of background: indications for hematopoietic stem cell transplantation (hsct) for adults with all evolve over time and vary among countries. methods: the goal of this study was to assess general trends in the number of various types of hscts performed between years and in europe. data reported to the ebmt registry were used for this analysis. in addition, we evaluated hsct rates with respect to the incidence of all in selected european countries. results: altogether, first allogeneic (n= ) or autologous (n= ) hscts were performed in the period - . comparing years - and - , the number of allohscts performed in first cr increased by %, most prominently for transplantations from unrelated ( %) and mismatched related donors ( %). the number of hscts from matched sibling donors increased by %, while the number of autohscts decreased by %. the increase of the use of allohsct, irrespective of the disease stage, was stronger for ph-pos ( %) than ph-neg all ( %). among patients aged > years, the number of allohsct increased by % while among younger adults ( - years), by %. between and , peripheral blood was used as source of stem cells in % cases of allohsct, compared to % between - . the use of bone marrow decreased from % to %, respectively. the proportion of allohsct with reduced-intensity conditioning (ric) increased from % to %. among myeloablative transplantations, regimens based on total body irradiation were the preferable option (app. % over the whole study period). in contrast, among ric regimens, the use of chemotherapy predominated ( % between - ) . in most of analyzed individual countries, the estimated rates of allohsct (no. hsct per newly diagnosed all) for patients in cr increased over time. however, the values for a period - varied strongly, being highest in finland ( . ), followed by the netherlands ( . ) and sweden ( . ) while lowest in russia ( . ) . conclusions: results of our analysis indicate continued trend to increased use of allohsct for adults with all, which may be attributed to increasing availability of unrelated donors. however, it may also be speculated that the introduction of tyrosine kinase inhibitors allowed higher proportion of patients with ph-pos all proceeding to transplantation. finally, the implementation of ric regimens contributed to wider use of allohsct among older adults. limitations of the analysis include any assumptions made regarding all incidence for the specified time period and possible variation in reporting to the ebmt registry from different countries over time. year type of hsct hsct - hsct hsct - hsct hsct - hsct hsct - hsct hsct - disclosure: sg has received honoraria for amgen. as and sw are employees of amgen and own shares in amgen. mm has received honoraria for speaking for amgen and pfizer. outcome of allogeneic-hsct in adult patients with phpositive-all in the era of tki: a retrospective analysis of università degli studi e ospedale maggiore policlinico di milano, milano, italy, genova, italy, azienda socio sanitaria territoriale papa giovanni xxiii, bergamo, italy background: we conducted a retrospective, nationwide analysis to describe the clinical outcome of adult patients with philadelphia chromosome-positive acute lymphoblastic leukemia (ph+all) undergoing allogeneic hematopoietic stem cell transplantation (hsct) after being treated with a tki based therapy. methods: a total of patients were included in the study. the median age at hsct was (range: - ). all patients ( %) received tki before hsct (performed between - ) . of these patients, ( %) were in cytologic complete remission (cr) while ( %) had an active disease at the time of hsct. molecularly measurable residual disease (mrd) was negative in patients ( %) at the time of hsct. the donor was unrelated in % of cases. the prevalent source of stem cells was peripheral blood ( %). the conditioning regimen was myeloablative in % of cases (tbi-based in %) and included atg in % of cases. results: with a median follow-up after hsct of . months (range: , the probability of overall survival (os) at , and years was . %, . %, and . %, respectively, with a median os of months. progression free survival (pfs) at , and years was . %, . % and . %, respectively. os and pfs were significantly better in patients with cr and mrd-negativity at the time of transplant compared to patients with cr but mrd-positive ( % os not reached vs. months, p= , ; % pfs not reached vs months, p= . ). the cumulative incidence of relapse (cir) at years was significantly lower in patients with cr and . %, p= . ). the non relapse mortality (nrm) after , , and years was . % ( %ci: . - . ), . % ( %ci: - . ), and . % ( %ci: - . ), respectively. the subgroup of patients with mrdnegative both at hsct and at rd month after hsct had a better outcome ( year os %). conversely, the patients who underwent hsct with active ph+all had a median os and pfs of and months, respectively. background: haploidentical (haplo) donors have expanded patient transplant access. however, outcome of patients with acute lymphoblastic leukemia (all) undergoing allogeneic stem cell transplant (asct) with haplo donors in argentina has not been reported. we aimed to analyze the outcome of asct in patients with all, particularly results with haplo donors. methods: we collected data from patients with an all diagnosis who underwent asct in first complete remission (cr ) and subsequent remissions (cr +) in centers in argentina, affiliated to gatmo, between and patients that underwent asct with matched donors (sibling and unrelated) and haplo donors (with post-transplant cyclophosphamide) were included. both donor categories were compared in terms of overall survival (os), nonrelapsed mortality (nrm) and cumulative incidence of relapse (cir). graft versus host disease (gvhd) was also evaluated. multivariate analysis was performed by cox regression for os and fine-gray for ci of competing events. a further propensity score (ps) adjustment was performed by donor group. results: in a -year period, patients were included (mean age y; range - ; male . %); ( %) during last years. all phenotype was b ( %) and t ( %). at diagnosis, / ( %) had cns involvement and / ( %) were philadelphia chromosome positive. asct was performed in cr (n= ; %) and in cr + (n= ; %) after a median time from all diagnosis to asct of and months, respectively. comorbidity index (hct-ci) was - in / ( %). donors were matched (n= ; %; related and unrelated) and haplo (n= ; %). conditioning regimen was myeloablative in / ( %; patients with total body irradiation), and this conditioning was more frequent in matched ( %) than haplo ( %) (p= . ) donors. two-years os was % ( %ci - ) for the entire population; % ( ci - ) for matched donors and % ( % ci - ) for haplo donors (p= . ). in the multivariate analysis, pretransplant status (cr vs cr +; hr . , p< . ), cns status at diagnosis (yes vs no; hr . ; p= . ) and unrelated donors (yes vs no; hr . ; p= . ) were independently associated with os; donor category had not impact in the os. by adjusting the ps term (roc area . ), no difference was found by donor category. twoyears nrm was % ( %ci - ) for matched and % ( %ci - ) for haplo (p= . ) donors; older donors (p= . ) and unrelated donors (p= . ) were associated with higher nrm. two-years cir was % ( %ci - ) for matched and % ( %ci - ) for haplo (p= . ) donors; only male donors were associated with higher cir (p= . ). ci of grade - acute gvhd was % vs % (p= . ) and chronic gvhd was % vs % (p= . ) for matched and haplo donors respectively. in both groups, matched and haplo donors, the half of deaths were due to relapse. background: relapse is the most important cause for treatment failure in pediatric b-precursor acute lymphoblastic leukemia (bcp-all) occurring in - % of patients. mechanisms of ineffective graft-versus-leukemia (gvl) effects or t-cell responses against all remain to be investigated. methods: we analyzed parameters of immunosurveillance in bone marrow (bm) samples of pediatric patients to identify potential mechanisms of t-cell suppression. expression of co-stimulatory/ co-inhibitory molecules was analyzed to identify implications for gvl. expression was correlated with clinical outcome ( years mean followup) . t-cell immunoglobulin and mucin-domain containing- (tim- ) overexpression and crispr/cas -mediated knockout (ko) in primary t cells were performed to analyze its role for anti-leukemic t-cell functionality. to induce an interaction of t cells with leukemic blasts, anti-cd /-cd bispecific t-cell engager (bite) was added and t-cell activation/ proliferation were analyzed. fold change (fc) was created by comparing levels of t-cell activation/ proliferation before vs. after co-culture. transcriptome analysis of primary bm samples identified expression levels of known tim- inducers. results: flow cytometric analyses of bcp-all samples showed increased tim- expression on cd + bm t cells at initial diagnosis in patients with relapse in the course of disease. multivariate analysis confirmed -fold increased relapse risk in tim- high (n= ) vs. tim- low expressing (n= ) patients. pd- expression on bm t cells alone had no impact on relapse-free survival (rfs), whereas patients with high percentage of tim- /pd- double positive cd + bm t cells showed significantly decreased rfs ( . % vs . %). co-culture experiments revealed that tim- is induced in primary t cells by contact with leukemic cells (mean tim- expression . % vs. . % on t cells with vs. without addition of leukemic cells, n= ). transcriptome analysis was performed to identify expression levels of known tim- ligands/ inducers in bm samples with high vs. low tim- expression. no significant differences in expression levels of high-mobility-group-protein b (hmgb ), carcinoembryonic antigen-related cell adhesion molecule (cea-cam ) or galectin were observed. known tim- inducers il- , il- , il- , il- or transforming growth factor beta (tgf-β ) were not differentially expressed indicating that another mechanism must be responsible for tim- overexpression. tim- overexpression and crispr/ cas -mediated tim- ko were performed to analyze functional relevance of tim- expression in an in vitro leukemia model. t cells of healthy donors were co-cultured with leukemic cells and anti-cd /-cd bite to induce anti-leukemic t-cell response. tim- ko t cells showed significantly increased activation compared to wildtype t cells (fc of cd expression . vs. . , n= ) . in contrast, proliferation of tim- overexpressing t cells was significantly impaired (fc . vs. . , n= ) , whereas tim- ko t cells showed higher proliferation levels compared to controls (fc . vs. . , n= ) . conclusions: tim- expression on cd + bm t cells is a strong predictor for pediatric bcp-all relapse and is induced by t-cell interaction with leukemic cells. tim- expression decreases anti-leukemic t-cell activation and proliferation and thus constitutes a new mechanism of immune escape and potentially insufficient gvl effects in pediatric bcp-all. targeting the tim- axis can be of interest to improve future immunotherapy of advanced bcp-all. disclosure: nothing to declare. abstract already published. multiparameter flow cytometric minimal residual disease before myeloablative allogeneic hematopoietic cell transplantation in acute myeloid leukemia influences patients survival in first and second complete remission background: the growing evidence from the literature strongly suggest that multiparameter flow cytometric (mfc) minimal residual disease (mrd) assessment in aml can be used to risk-stratify patients at the time of allogeneic hematopoietic stem cell transplantation (allohct). we sought to determine the significance of mfc-mrd status in patients with aml in first or second complete remission (cr) treated with myeloablative conditioning (mac) allohct at six centers of the polish adult leukemia group (palg). methods: mrd was assessed by -color ( -color since ) mfc performed on bone marrow aspirates obtained as routine assessment before allohct. all consecutive patients undergoing mac allohct were included in the analysis if they underwent pre-hct mfc-mrd analysis from may until january . the abnormal population was quantified as a percentage of the total cd + white cell events. residual disease at a ≥ . % level was considered mrd-positive (mrd+) . results: we identified adult patients (median age years, range - ) with aml undergoing allohct from either hla-identical sibling (n= ) or unrelated donor (n= ), in cr (n= ) or cr (n= ), who were conditioned with i.v. busulfan given in myeloablative dose ( , - , mg/kg) in combination with flu (n= ) or cy (n= ). gvhd prophylaxis consisted of calcineurin inhibitor combined with mtx plus atg in allohct from unrelated donors. positive mrd status before allohct was detected in / ( %) pts in cr and / ( %) pts in cr . the mrd(-) and mrd(+) groups did not differ in terms of gender, age, eln cytogenetic and molecular genetic risk, first and second cr, conditioning regimen, hsc source, and type of donor. the -year overall survival (os) for mrd(-) and mrd(+) patients were % and % (log-rank p= . ). the respective -year leukemia-free survival (lfs) were % and % (log-rank p= . ). in univariate and multivariate cox proportional hazard model the only significant adverse prognostic factors for lfs were mrd(+) (hr . , p= . ) and high eln genetic risk (hr . , p= . ) . the same factors significantly influenced os [hr . , p= . and hr . , p= . for mrd(+) and high eln risk, respectively] . conclusions: our findings confirm that pre-transplant residual disease at a ≥ . % level assessed by mfc is independent risk factor for both lfs and os in patients undergoing allohct. in addition, the results of our study show that mac allohsct outcomes in patients with aml in first and second cr are significantly influenced by both mfc-mrd status and eln cytogenetic and molecular genetic risk. [[o image] . leukemia-free survival according to pre-transplant minimal residual disease level] disclosure: nothing to declare o second allogeneic stem cell transplantation in acute lymphoblastic leukemia patients in second complete remission or relapse: a study on behalf of the alwp / ebmt background: second allogeneic transplantation (hsct ) is a therapeutic option for patients (pts) relapsing (rel) after first hsct (hsct ) however most of the available data is in acute myelogenous leukemia (aml) and there is very limited data on hsct in patients (pts) with all. therefore, the alwp of the ebmt performed a large registry analysis to study the outcome of hsct in pts with all. methods: we studied pts receiving hsct as a salvage treatment between the years (y) - for rel following hsct in cr . median follow-up of surviving pts was months (iqr: - ). results: pts ( %) were males and median age at hsct was . years (range: - ). median time from hsct to hsct was ( - ) days (d) and from rel to hsct it was ( - ) d. at the time of hsct ( %) pts were in cr while ( %) had advanced disease. ( %) pts received sibling donor (msd) and ( %) unrelated donor (ud) hsct ( / - ; / - , missing hla- ). in % of pts with available data the hsct was performed from the same donor. the majority of pts with available mrd data transplanted in cr ( / pts) were mrd negative pre-hsct . karnofsky performance status was > in % of pts. % were transplanted with pb graft. ( %) pts received chemotherapy based conditioning (reduced intensity %, myeloablative %) while it was tbi based in ( %) pts. ( %) pts received in vivo t-cell depleted (tcd) grafts. % of the pts engrafted. acute graft-versus-host disease (agvhd) ≥ ii and ≥ iii-iv occurred in % and % of the pts. incidence of y total and extensive chronic gvhd was % and %, respectively. main causes of death were leukemia recurrence in %, gvhd in % and infections in %. at and y, the cumulative incidence of nrm, ri, lfs, os and grfs were % & %: % & %, % & %, % & % and % & %, respectively. in mva no factor predict nrm. in multivariate analysis, ri was independently associated with time from hsct to rel, agvhd ≥ii after hsct and in vivo tcd and lfs the prognostic factors were time from hsct to rel and kps≥ at hsct . factors associated with os were age (per years), time from hsct to rel, ric at hsct , kps> at hsct and ud vs msd. longer time internal from rel to hsct and in vivo tcd was associated with inferior cgvhd. lastly for grfs the prognostic factors were time from hsct to rel, agvhd ≥ii after hsct , ric at hsct and kps≥ at hsct . conclusions: results of hsct in all pts with rel or cr are poor with only % os and % grfs at y with very high ri of %.the prognostic factors are similar to those previously reported for hsct in aml. the future goals are to prevent and treat relapsed all by mrd driven novel monoclonal antibodies and car-t cell therapy. disclosure: nothing to declare. aplastic anaemia outcomes of allogeneic stem cell transplantation (hsct) for older patients (> years) with severe aplastic anaemia using alemtuzumab-based ('fcc') regimen: king's college hospital experience background: treatment of older patients with severe aplastic anaemia (saa) is problematic with poor long-term survival after treatment with antithymocyte globulin (atg) and/or ciclosporin (csa). use of fludarabine, low dose cyclophosphamide (cy) and atg ('fcatg') conditioning suggests better outcomes among older patients transplanted from matched sibling donors compared to high dose cy/ atg conditioning, but gvhd remains a serious concern. we have transplanted saa patients aged > years, predominantly from unrelated donors, using alemtuzumabbased ('fcc') regimen. methods: from our fcc saa database of patients, aged > years were transplanted between - . median age was years (range - ); aged - and aged ≥ years. donor was matched sibling (msd) in ( %), / matched unrelated (mud) in ( %), / unrelated (mmud) in ( %). conditioning was fludarabine mg/m x , cy mg/m x , alemtuzumab . mg/kg daily from day - to - . post graft immune suppression was csa alone. gy tbi was added to fcc for mmud hsct. all patients received peripheral blood (pbsc) as stem cell source. / ( %) patients were hla alloimmunised. pb telomere length (tl) by multiplex qpcr measured in patients, was < st centile in ( %), < th centile in ( %) and normal in ( %) patients. first line hsct was performed in / ( %) msd and / ( %) among unrelated donors. hct comorbidity index (htc-ci) score was - in ( %); in ( %) and > in ( %). results: three patients had invasive fungal infection at time of hct and died day + to , and one patient died at months from multiorgan failure with recurrent parainfluenza virus and cmv. median cd chimerism was % ( - ), % ( - ) and % ( - ) at day + , iyr and yr post hsct. one late graft failure at months was associated with low csa blood levels, and was followed by successful nd transplanted with no gvhd. -year os was %, compared to % among patients aged < years (p= . ). os was % and % for patients aged - and ≥ year, respectively, p= . . os for msd, mud and / mmud was %, % and %, respectively. htc-ic score of > was associated with worse os of % compared to % with score < , p= . . outcomes were comparable irrespective of telomere length ( % vs % for normal vs short telomere, p= . ). cumulative rates of acute and chronic gvhd were % and %, respectively. all cases of acute gvhd were confined to skin and grade i/ii only, and no cases of severe chronic gvhd. ( %) patients needed dose reduction of csa with addition of mycophenolate due to renal dysfunction. rates of cmv and ebv reactivation were % and % respectively, with no cmv or ebv disease. conclusions: fcc conditioning regimen enabled high survival and low risk of gvhd among older patients with hct-ci score < and who did not have established invasive fungal disease at time of hsct. clinical trial registry: not applicable disclosure: no conflicts of interest to declare haploidentical transplantation with post-transplant cyclophosphamide (haplo-ptcy) for patients with acquired or inherited bone marrow failure syndromes (bmf): the experience from curitiba, brazil background: availability of unrelated donors as well as time to find a donor and the costs related to graft acquisition are important limitations in countries with ethnical minorities and fewer resources. methods: we describe the experience of transplants in patients(pts) with acquired or inherited bmf submitted to an haplo-ptcy transplantation between . and . . the median age was ys, % were male and % were cmv positive. haplo-ptcy was the st transplant for pts, second or third for pts. diagnosis: fanconi anemia (fa,n= ), acquired severe aplastic anemia (asaa,n= ), telomere diseases (n= ); other inherited bmf (n= ). all pts had failed prior therapies and % had previous blood transfusions. the majority received a ric regimen with low dose tbi (n= , %). donors were father(n= ), mother(n= ), other relatives(n= ). bone marrow was the stem cell source in pts. all pts received gvhd prophylaxis that included ptcy followed by cyclosporine and mycophenolate mofetil. fa pts received a modified preparatory regimen and ptcy at a total dose of mg/kg (n= ) or mg/kg (n= ) while other bmf received mg/kg results: fa pts: pts did not receive atg in the preparatory regimen and all engrafted, despite the presence of donor specific antibodies(dsa) in pts. three pts had aml and are in remission and ys after transplant. pts died due to gvhd (n= ); toxoplasmosis/cmv pneumonia (n= ) or relapse (n= ). / pts are alive with a median follow-up(fu) of . ys. in the subgroup of fa pts receiving r-atg(n= ), pts presented primary or secondary graft failure(gf), none had dsa and all died despite a nd haplo-ptcy with different donors. pts had advanced disease and are in remission at the last fu. / pts are alive at a median of ys after transplant. eight pts died due to gvhd (n= ); rsv pneumonitis (n= ) and gf (n= ). all pts transplanted with asaa are alive and fully engrafted at a median of . ys after transplant and none developed gvhd, nine out of pts transplanted for other inherited bmf are alive at a median of ys after transplant. gf occurred in pts, all received a nd haplo-ptcy from different donors and are alive and engrafted. pts died due to gvhd (n= ), gf (n= ) and tma (n= ). altogether cmv reactivation occurred in pts ( %), at a median of days (range: - ) and hemorrhagic cystitis in pts ( %) at a median of days (range: - ). after transplant. conclusions: haplo-ptcy for pts with acquired or inherited bmf should be offered for those who need an immediate transplant but lack a matched donor. % of pts are alive at a median fu of ys but gvhd is a major complication for pts with inherited bmf, especially fa. new approaches to gvhd prophylaxis and treatment are needed in order to improve quality of survival for these pts. disclosure: nothing to declare relationship between plasma rabbit anti-thymocyte globulin level and response to immunosuppressive therapy in patients with severe aplastic anemia: results of a multicenter, prospective, randomized study background: patients with acquired aplastic anemia (aa) who do not have hla-matched donors receive immunosuppressive therapy (ist) with anti-thymocyte globulin (atg). previous studies have suggested several variables that predict response to ist. however, no studies have investigated the plasma atg level as a variable. in this study, we assessed the relationship between plasma rabbit atg (r-atg) level and response to ist in patients with severe aa. methods: patients with severe aa who required initial ist were enrolled from may to october . the ist regimen included r-atg (thymoglobulin®, sanofi, cambridge, . or . mg/kg/day for days) and cyclosporine a ( mg/kg/day for minimum months). plasma r-atg level was measured using a rabbit igg elisa kit on days and . response rate was defined as complete and partial responses at months. receiver operator characteristic curves were generated to discriminate between response and no response to ist. results: a total of patients (aged . - . years) were randomized; and patients received . and . mg/kg of r-atg, respectively. in the . mg group, the response rate was %, which was comparable with that in the . mg group ( %) (p = . ). plasma r-atg level greatly varied in both groups. median r-atg level on days and after ist was . ( . - . ) and . ( . - . μg/ml), respectively, which was not significantly different between two dosages of atg groups (day , p = . ; day , p = . ). according to the r-atg level, response rates were significantly higher in the group with higher r-atg level than in that with lower atg level (day , % vs. %, respectively; p = . and day , % vs. %; p = . ) (figure) . cut-off levels at days and were . and . μg/ml, respectively. the vast majority ( %) of patients with levels higher than cut-off levels on day responded to ist. in multivariate analysis, higher atg levels at day were independent favorable predictors of response to ist at months (or = . ; % ci: . - . ; p = . ). there were no significant differences in the kinetics of lymphocyte subsets among patients treated with different dosages of atg. however, higher atg level was associated with lower cd + t and regulatory t cell numbers for the entire -month period. conclusions: the present data indicate interindividual variability in plasma r-atg level. higher atg level resulted in improved response to ist and correlated with prolonged immune reconstitution. individualized dosing of atg via a pharmacokinetic model may improve the response rate to ist and reduce the number of patients who require allogeneic stem cell transplantation following ist. clinical background: radiation and dna alkylating agents used in hematopoietic cell transplantation (hct) can cause organ damage, malignancy and death. these risks are heightened in patients with genetic bone marrow failure (bmf) syndromes driven by defects in cellular proliferation or dna repair, including dyskeratosis congenita (dc), which arises from impaired telomere maintenance. we hypothesized that proliferative defects in hematopoietic cells of patients with bmf and very short telomeres might permit myeloid engraftment following hct without the need for radiation or dna alkylating agents. we conducted a multi-center prospective trial (nct ) evaluating engraftment after hct without these agents. methods: we enrolled bmf patients with genetic validation of dc or lymphocyte telomere length < st percentile by flow-fish. we performed hct using bone marrow allografts from related or unrelated donors matched at or of hla alleles after a preparative regimen consisting of only fludarabine and alemtuzumab. graft versus host disease (gvhd) prophylaxis consisted of cyclosporine a and mycophenolate mofetil. the primary endpoint of the trial was donor myeloid engraftment, defined as an absolute neutrophil count ≥ cells/μl by day + and donor myeloid chimerism > % by day + . [[o image] . engraftment and survival after radiation-and alkylator-free hct for bmf with very short telomeres] results: twenty patients (age . - . years old at hct) received treatment between august -october at institutions. eighteen of the patients received unrelated donor grafts ( matched, single-allele mismatched). primary myeloid engraftment was achieved in of patients ( %) at a median days post-hct (range - days). the single patient with primary graft failure had dc-related liver disease and hypersplenism; in this case, splenectomy at day + promptly revealed donor myeloid engraftment. of the other patients, had sustained myeloid engraftment, with a median post-hct follow-up of months (range - months). three patients had secondary graft failure. two of these had early graft rejection and underwent successful repeat hct using higher intensity regimens. the third patient maintained high donor chimerism after primary engraftment but developed severe neutropenia in the setting of multiple viral reactivations, and died of a fungal infection days post-hct. there was one other death, due to dc-related gastrointestinal complications months post-hct. none of the patients who engrafted durably under the protocol regimen had acute gvhd. four had chronic gvhd ( limited, extensive), treated successfully with limited courses of topical or oral steroids. conclusions: we conclude that this radiation-and alkylator-free hct conditioning regimen is an effective strategy for bmf in patients with dc or very short lymphocyte telomeres. eliminating dna damaging agents may reduce hct complications including gvhd and enable transplant in patients with high-risk comorbidities. clinical background: the standard treatment of acquired aplastic anemia (aa) is either intensive immunosuppressive therapy (ist) or allogeneic hematopoietic cell transplantation (hct). as supportive measures, red blood cells and platelet transfusions are the mainstay of therapy and patients are often multitransfused, which in turn can lead to anti-human leukocyte (hla) alloimmunization. in acquired aa the rate of hla-alloimmunization has previously shown a higher frequency in patients with aa compared to hematological malignancies. however, these results date back before the general introduction of leukoreduction of blood products and photochemical pathogen reduction of platelet components, and are based on cell-based assays. in recent years, leukoreduction and pathogen reduction of blood products became standard in switzerland and the solid-phase assay (luminex® technology) is now widely available to test for hla-antibodies, allowing a more extensive and detailed characterization of hla-antibodies. with these techniques, less is known on the exact incidence of hla-antibodies and their magnitude, associated cofactors and its impact on treatment outcomes in acquired aa. methods: we retrospectively investigated aa patients treated with ist (n= ) and/or hct (n= ) at the university hospital of basel and the university children's hospital of basel (switzerland) regarding hla antibodies since the introduction of testing with the luminex® at our center in . at least one hla antibody measurement before and/or after therapy was available per patient. all patients received leukoreduced blood products and as of platelets treated with intercept® (uv+ amotosalen). results: overall, hla-antibodies were detected in ( %) patients with a higher rate of hla alloimmunziation by severity of aa (p< . ). the median number of hlaantibodies in each patient before therapy (i.e. ist or hct) was (iqr - ). in patients undergoing hct hlaantibodies were more frequent before treatment start as compared to patients with ist treatment (median (iqr - ) versus . (iqr - . ), p< . ). differences between treatments remained after adjusting for all covariates (p< . ). there was no statistically significant difference regarding the hla antibody mean fluorescence intensity (mfi) between the two treatment forms (overall mean mfi of +/- ). the highest mean hlaantibody mfi before therapy was (+/- ) with a maximum of . females showed a significantly higher number of hla-antibodies (p< . ) and also higher mean mfi (p< . ). furthermore, the number of pregnancies was associated with higher numbers of hla-antibodies (p< . ), however the number of transfusions did not have significant impact on hla-antibody number and mfi. regarding outcome, there was no significant association between the number of hla-antibodies and engraftment as well as bleeding events. conclusions: hla-alloimmunization is still frequent in patients with acquired aa but today number of pregnancies and gender seem to be more important for development of hla-alloimmunization than number of transfusions. interestingly, patients treated by hct show a higher rate of hla-alloimmunization before treatment start in comparison to ist, thereby emphazing the importance of blood management and donor selection in hct in acquired aa as hla-antibodies can cause platelet refractoriness and can represent donor-specific antibodies in the setting of mismatched hct (e.g. haploidentical). disclosure: nothing to declare. health-related quality of life in systemic sclerosis before and after autologous hematopoietic stem cell transplant -a systematic review background: autologous hematopoietic stem cell transplantation (ahsct) for severe rapidly progressive systemic sclerosis (ssc) allows significant regression in skin and lung fibrosis and improvements in overall and event free survival up to years after transplant. we undertook this study to synthesize the evidence on changes in healthrelated quality of life (hrqol) associated with ahsct for ssc. methods: autologous hematopoietic stem cell transplantation (ahsct) for severe rapidly progressive systemic sclerosis (ssc) allows significant regression in skin and lung fibrosis and improvements in overall and event free survival up to years after transplant. we undertook this study to synthesize the evidence on changes in healthrelated quality of life (hrqol) associated with ahsct for ssc. results: the search returned articles. eight were selected: uncontrolled phase i or ii trials, cohort studies and rct (assist, astis, scot). hrqol data from ssc patients treated with ahsct and with intravenous cyclophosphamide (cyc) as a comparator were extracted. hrqol was assessed using the health assessment questionnaire-disability index (haq-di) (n= patients), the short-form health survey (sf- ) (n= patients) and the euroqol dimensions (eq- d) (n= patients). hrqol was analyzed as a secondary outcome in all studies. quality of the data was assessed as high. haq-di improved significantly more with ahsct compared to cyc (- . vs - . , p= . at years in astis; % vs % improved at . years in scot). scores also improved pre-post ahsct in the uncontrolled studies (ranging from - . to - . points at one year (all p< . ), and up to - . points at . years (p< . )). sf- physical component summary scores were significantly better in subjects treated with ahsct compared to cyc (between group differences ranging from points at one year in assist and . points at years in astis (p= . ); % vs % improved at . years in scot (p= . )). similar differences pre-post treatment scores were also reported in an uncontrolled study (increase of points, p< . ). in assist, there was a trend for the sf- mental component summary score to improve in the ahsct arm ( vs , p= . ) and worsen in the cyc arm ( vs , p= . ) at one year. there were no significant differences between the ahsct and cyc arms in astis and scot with . and . years of follow-up, respectively. post-treatment scores improved significantly compared to pre-treatment in an uncontrolled study (from to points, p= . ). astis showed a significant difference in the index-based utility score of the eq- d ( . , p< . ) and a non-significant difference in the general health visual analogue scale ( . , p= . ) at two years, in favour of ahsct compared to cyc. conclusions: although there is heterogeneity in the reported data, ahsct in ssc was consistently associated with marked and sustained improvement in hrqol. this analysis provides additional compelling data for the role of ahsct in ssc when assessing patient's point of view. clinical trial registry: we hypothesised that ahsct induces a rebooting of thymic function, resulting in the re-development of a functional, tolerant immune system. we aimed to examine cellsurface and dna markers of recent thymic emigrants (rte's) longitudinally in a cohort of ms patients post-ahsct in order to identify markers that correlate with a durable treatment response. methods: peripheral blood mononuclear cells (pbmncs) were collected from patients enrolled in the phase ii trial at st vincent's hospital sydney for ahsct in ms (moore et al, jnnp in press). a multicolour flow cytometry panel to optimally identify rte's was performed on patient samples at , , , and month timepoints, allowing us to track changes in thymic output longitudinally following ahsct. dna markers of thymic function -sj:b trec ratio was performed in the same cohort of patients, on the same bio-banked sample to enhance the validity of observed changes. statistical analysis was performed with graphpad prism. results: a sustained, significant increase in rte's and sj:b trec was detected between pre-transplant and month post-transplant specimens (p = . ). in patients where similar analysis was able to be performed at months a trend to increased markers of thymic output was observed. a correlation between rte's and sj:b trec was observed (r= . , p = . ). contrary to other publications in the field, trec as a marker of thymic output did not appear to be lower when patients were analysed by age (< yrs vs. > yrs). greater thymic output as determined by sj:b trec was observed at all timepoints in patients who had evidence of sustained disease remission as opposed to patients who experienced disease relapse. conclusions: we have seen evidence that sustained thymic reactivation is a component of immune reconstitution following ahsct for ms. previous studies have only demonstrated thymic activity at months but this study confirms that thymic activity remains prominent at and even months. this thymic regeneration may contribute to a durable clinical response in patients with ms post hsct. clinical background: allogeneic hsct offers the potential replacement of an aberrant immune system. this retrospective study assessed long-term outcomes of this strategy in patients treated for severe autoimmune diseases (ads), reported to the ebmt registry. methods: among the total patients who received allogeneic hsct between - , we received detailed questionnaires on long-term outcomes from patients. the diagnosis of ad was hematological (n= ) and nonhematological (n= ), among pediatric (= ) and adult (= ) populations. the median age of patients at hsct was . years (pediatric: median . years, range . - . ); adult : median . years, range . - . ). all patients were refractory to previous immunosuppressive therapies (median of lines of treatments, range - ), and eight of them received a previous autologous transplant. the graft source was pbscs in , bm in , and cord blood in patients. donors were as follows; % mrd, % mud, % mmrd, % syngeneic and % cord blood. conditioning was mac in and ric in patients. serotherapy with atg was given in patients, while patients received alemtuzumab. post-transplant gvhd prophylaxis was cyclosporine-based for the majority of patients (n= ). results: median follow-up was months (range . - months). toxicity profile was similar to allogeneic hsct in other contexts. the incidence of grades ii-iv acute gvhd was . % ( % ci: . - . ) at -days; severe acute gvhd was reported in . % of them. cumulative incidence of chronic gvhd was . % ( % ci: . - . ) at -year; extensive manifestations were reported for % of chronic gvhd. overall graft rejection rate was . %. seven secondary ad and one case of new malignancy (lymphoma) occurred. viral reactivations were reported in a total of patients, including cmv (n= ), ebv (n= ), adenovirus (n= ), bk virus (n= ), hsv (n= ), hhv (n= ) and vzv (n= ). seven cases of invasive fungal infection were reported (one aspergillosis and three candidiasis). ten bacterial infections (only patients developed infection from gram-negative bacteria) and four pneumonia were observed. at the last follow-up complete clinical response was obtained in . % of patients, while partial remission was reported in . %. relapse incidence (ri) was . % ( % ci: . - . ) at the last follow-up. post-hsct autoimmune disease specific treatment was required for patients. in subgroup analysis among different diseases, the os rates were similar between immune cytopenia and other ads. at years, os was % ( % ci: . - . ) , nrm was . % ( % ci: . - . ) and pfs was . % ( % ci: . - . ), with no differences between immune cytopenia ( . %) and other ads ( . %). by multivariate analysis, only one prognostic factor remained significantly associated with long-term outcomes: a more recent year of transplant (better os, p= . ; lower chronic gvhd, p= . ). conclusions: this large retrospective survey of the ebmt registry confirms the potential of allogeneic hsct to produce long-term disease remission in a large proportion of refractory ads, with acceptable toxicities and nrm. results: twenty-seven patients were evaluated before and at months after transplant, of which were additionally evaluated at months. at and months after ahsct, patients presented significant improvement of mrss (p< . ), mip (p< . ), mep (p< . ), mwt distance (p= . ), and physical (p< . ) and mental (p= . ) components of the sf- , when compared to pretransplant evaluations. no changes were observed in fvc after treatment. despite a transient decline in dlco at months (p< . ) after transplant, dlco levels at months were not different from baseline (pre-transplant). significant correlations were observed between the mwt distance and physical component score of quality of life (r= . , p< . ). no significant correlation was observed between pulmonary function and the other evaluated variables (mrss, respiratory muscle strength, physical capacity and quality of life). conclusions: ahsct significantly improves the functional status of ssc patients in the first year of follow-up. although the pulmonary function remained stable after ahsct, there was significant increase in the physical capacity and quality of life of patients. these results can be interpreted as positive outcomes of ahsct for ssc. disclosure: nothing to declare. model of multidisciplinary and multicenter approach for hsct for children with multiple sclerosis: long- background: hsct for children with multiple sclerosis (ms) proved effectiveness and safety. it is required to improve the results by analysis of long-term follow-up and late effects. several challenges identified in multidisciplinary collaboration for successful treatment as well as a problem of switching these patients to the adult healthcare. we aimed to create a model of organization of help for children with severe refractory multiple sclerosis based on multidisciplinary and multicenter approach. methods: fifteen children with ms received autologous hsct (ahsct) from january to may . gender: females - , males - . mean length of ms prior ahsct was . ± . months and mean age of ms debut was . ± . years old. all patients had severe refractory ms treated with corticosteroids, interferons, plasmapheresis and mitoxantron with negative results. these patients presented signs of neuroinflammation. mean baseline edss before the start of mobilization was . ± . . procedures included mobilization with the help of cyclophosphamide and filgrastim and conditioning: cyclophosphamide mg/kg and hatg mg/kg. all patients received at least x ˄ /kg cd + hematopoietic stem cells (mean . x ˄ ± . x ˄ ). we analysed the incidence and nature of late effects in patients with at least one year of follow-up (based on the standard protocol for late effects). fertility preservation proposed for patients. ahsct as well as pre-and posttransplant care was done by multidisciplinary team involved both transplant and neurological team. technology of transfer patients to adult center for post-transplant observation was identified. results: all patients survived. mean time to engraftment was ± . days. eleven patients experienced culture negative fever, one patient -cystitis, and one patient had cmv reactivation within days of the transplant. no patient experienced an edss increase post-hsct above baseline, and all patients improved. mean improvement of edss was . ± . during the first days after ahsct (fast recovery). in-time transplanted patients improved better. improvement confirmed by immunological data (increasing of immune regulation index and t-regs in comparison with the baseline). median follow-up period was months ( - months) . four patients ( . %) experienced exacerbations (both neurological and mri) in median of years ( - years) after ahsct. no onsets of secondary autoimmune disease and malignancies was seen. cardiocascular late effects were seen in patients and endocrine -in patients (all females). all these late effects were successfully treated. patients after the age of years old were transferred to partnering adult center. this center uses the same protocol for hsct (in adults) and posttransplant observation. detailed scheme of transfer developed. conclusions: ahsct for pediatric patients with severe refractory ms appears to be safe and effective method and in-time hsct can significantly improve the outcomes. most of the patients did not experienced exacerbations. late effects found in patients were successfully treated. we provide a best care for these patients in both childhood and adulthood by transferring them to adult center. thus, a unique multicenter and multidisciplinary model of care for children with severe refractory ms was found. disclosure background: neuromyelitis optica spectrum disorder (nmosd) is an inflammatory central nervous system disorder characterized, despite immunotherapy treatments, by life-long, severe, and disabling attacks of optic neuritis and myelitis. the aim is to determine if autologous nonmyeloablative hematopoietic stem cell transplantation could be an alternative treatment option. methods: following stem cell mobilization with cyclophosphamide ( g/m ) and filgrastim, patients were treated with cyclophosphamide ( mg/kg) divided as mg/kg intravenously (iv) on day - to day - , ratg (thymoglobulin) given iv at . mg/kg on day - , mg/kg on day - , and . mg/kg on days - , - , and - (total dose mg/kg), and rituximab mg iv on days - and + . unselected peripheral blood stem cells were infused on day . aqp -igg antibody status was determined by clia validated elisa or flow cytometry assays. cell killing activity was measured using a flow cytometry based complement assay. results: twelve (eleven aqp -igg positive) patients were treated with a median follow-up of months. ten patients are more than five years post-transplant. at five years, % of patients were relapse-free off all immunosuppression (p< . ). at one and five years after hsct, edss improved from a baseline mean of . to . (p< . ) and . (p< . ), respectively. nrs improved after hsct from a baseline mean of . to . at one year (p< . ) and . at five years (p< . ). the sf- quality of life total score improved from mean . to . (p= . ) and . (p= . ). aqp -igg serostatus converted to negative in nine patients and complement activating and cell killing ability of patient serum was switched off. two patients remained aqp -igg seropositive (with persistent cell killing ability) and relapsed within two years of hsct (p< . ). conclusions: prolonged drug-free remission with aqp -igg seroconversion to negative following nonmyeloablative autologous hsct warrants further investigation in larger randomized controlled trial. clinical trial registry: identifier: nct clinicaltrials.gov disclosure: nothing to declare o abstract already published. autoimmune haemolytic anaemia after haematopoietic stem cell transplantation in children: a french multicenter study background: autoimmune cytopenias (aic) are a rare but serious complication of haematopoietic stem cells transplantation (hsct). the auto immune haemolytic anaemia (aiha) is the most frequent of these complications in paediatrics and is difficult to treat. so far, there has been no nationwide post transplantation aiha study. methods: this observational, retrospective and multicentric study focused on french paediatric cases of aiha after hsct between january and january . data was collected from national reference databases and direct interview of physicians. the inclusion criteria were patients between to years old who developed an aiha or an evans syndrome after hsct. data concerning patient, primary diagnosis, hsct procedure, pre-transplant viral status, blood group compatibility, characteristics of cytopenia, delay transplant-cytopenia, graft vs host reaction. laboratory characteristics and therapies were analyzed as well as the response to first, second or third line treatments. results: paediatric hsct were performed in french paediatric centers between and . among them, children developed an aic: aiha, pancytopenia and evans syndromes. the median age at hsct was , years old ( , [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] ) . average delay between transplantation and aic was days ( - ). the median follow-up after transplant was , months ( - ) . patients were transplanted for non-malignant disease ( benign hemopathies, immunodeficiencies) and for a malignant one. stem cell source was bone marrow for % of patients, peripheral blood stem cells for % and cord blood for %. the donor was unrelated for % of hsct, matched related for % and % of procedures were haploidentical. % of patients received als as part of their conditioning regimen, it was myeloablative in % (tbi based for %) and reduced intensity in %. direct anti-globulin test (dat) was positive to igg +/-c d for % of patients, patients had a blood group incompatibility ( minor and major). % of patients had an acute gvh and chronic gvh on time of aiha was find in % of patients. for % of patients the first line treatment was a combination of steroids ( mg/kg/day), intravenous immunoglobulin and rituximab ( mg/m per week). for % of patients a second or third-line treatment was required (imurel, cellcept, cyclophosphamide, sirolimus, campath, bortezomib). patients died ( relapse, infections and severe anaemia, multiple organ failure). conclusions: this is the first study describing precisely the aiha post hsct in pediatrics in france along with the various treatments used. it's a rare but severe complication with multiple risk factors associated with a high mortality rate and no standardized therapy at this time. further studies would allow us to understand the disease better in order to prevent its occurrence and treat it more efficiently. disclosure: nothing to declare background: the prognosis of relapsed/refractory acute lymphoid leukemia (all), non hodgkin lymphoma (nhl) or chronic lymphocytic leukemia (cll) is very poor, particularly in patients relapsing after autologous (autohct) or allogeneic hematopoietic cell transplantation (allohct). in the last decade, several chimeric antigen receptor anti-cd (car ) constructs have been developed. two of them (tisa-cel and axi-cel) are already approved by the fda and ema for all and nhl. however, the availability and affordability of these commercial carts remains a challenge in europe. methods: the first academic pilot clinical trial (clinicaltrials.gov nct ) using our fully academic (a b :cd : - bb:cd z) car was approved by the spanish agency of medicines on may/ . eligibility criteria included r/r all (adult and pediatric), nhl and cll who had failed standard available therapy. the primary objective of the study was safety, and secondary objectives were response rate and its duration (progressionfree survival). here we report an updated analysis of this trial. results: as of december/ we have included patients in the study. of these, we performed lymphoapheresis to and we processed the cells of , although for the moment we have only infused to of them. the diagnoses of these patients were all in ( adults), nhl in and cll in . median age was years and % were women. of the patients with all, had relapsed after allohct, while / patients with nhl had relapsed after autohct. after conditioning with fludarabine ( mg/ m ) and cyclophosphamide ( mg/m ), we infused . - x ari- cells /kg, first as a single infusion (first patients, cohort ), and then in - fractions (last patients, cohort ). we have observed ( %) cases of severe cytokine release syndrome (crs) and the non-relapse mortality (nrm) was % ( / ) . these deaths were due to crs ( ) and pseudomembranous colitis ( ), and all happened in cohort . we had no cases of severe neurotoxicity, and grade ii neurotoxicity was only seen in patients. of the patients with active all at inclusion, had enough follow-up for efficacy analysis, and all of them achieved a cr ( of them with negative mrd). with a median follow-up of . months (range, . - . ) , relapses were observed on days + (cd +), + and + (cd -), leading to a progression-free survival of % at months. in patients with nhl/cll we have documented cr in , one of them (nhl) relapsing at day + . one nhl patient has not been evaluated yet and the other patients did not respond and have died due to their lymphoma. the cll patient, refractory to lines of treatment, achieved a cr, which is maintained at day + . conclusions: treatment with ari- cells is effective with a response rate of . % in all (pfs of % at y) and % in nhl/cll. we also observed cases of severe crs in % and a nrm of % of patients, though dose fractioning seems to improve safety profile. still, longer median follow-up and further patient inclusions are needed. clinical background: bcma car-t cells have demonstrated substantial preclinical and clinical activity for relapsed/ refractory multiple myeloma (rrmm) patients. in different clinical trials, the overall response rate (orr) varied from % to %. complete remission (cr) rate varied from % to %. previous studies indicated higher car-t cell expansion in vivo achieved better remission. here we developed a bcma car-t cell product manufactured via lentiviral vector-mediated transduction of activated t cells to express a second-generation car with - bb costimulatory domain. methods: our trial (chictr ) was initiated to evaluate the safety and efficacy of autologous bcma car-t treatment for rrmm. the enrolled rrmm patients either had received at least prior treatment regimens, including a proteasome inhibitor, an immunomodulatory agent, anti-cd monoclonal antibody or were double or triple-refractory, and have over % bcma expression. patients were subjected to a lymphodepleting regimen with flu daily for days ( mg/m , d to d ) and cy ( mg/ m , d- to d- ) prior to the car-t cell infusion (d ) at a dose range of - x car+ cells/kg. results: as of the data cut-off date (nov th, ), patients, median age . ( to ) years old, with a median of . ( . to . ) years since mm diagnosis, were infused with bcma car-t cells. patients had a median of prior regimens (range to ) ( figure a ). all the patients were eligible for initial evaluation of early clinical response with a median observation period of ( . to ) weeks. orr was %. all the patients achieved mrd negative in bone marrow by flow cytometry within - weeks after car-t infusion. partial response ( pr, . %), very good pr ( vgpr, . %), and complete response ( cr, . %) within weeks post car-t infusion were achieved. durable responses from weeks towards the data cut-off date were found in / patients ( . %). the disease progressed in patient from pr by week ( figure b ). one patient died of severe infection by day . all patients had detectable car-t expansion by flow cytometry from day post car-t cell infusion. expansion peaks were found between day to day . the peak car-t cell expansion in cd + lymphocytes of peripheral blood (pb) varied from % to % with a median percentage of . % ( figure c ). the peak absolute number of car-t cells in pb varied from to per μl with a median number of per μl. cytokine release syndrome (crs) was reported in all the patients, including with grade , with grade and with grade . car-t -related aes were pancytopenia ( . %), fever ( . %), nausea ( . %), heart failure ( . %). no patient died of crs complication. conclusions: our data showed bcma car-t treatment is safe with prominent efficacy. we also observed high expansion level and long term persistence of bcma car-t cells contribute to potent anti-myeloma activity. these initial data provide strong evidence to support the further development of this anti-myeloma cellular immunotherapy. clinical trial registry: chictr disclosure: nothing to declare chimeric antigen receptor (car)-mediated bcl b suppression in lymphoid progenitor cells propagates natural killer-like cell development background: the transcription factor b cell cll/lymphoma b (bcl b) is indispensable for t lineage development of lymphoid progenitors. pre-fabricated t cell products, allowing for a wider choice of effectively targetable antigens, being applicable to a wider range of patients, and minimizing the risk of long-term sequalae from on target/off tumor effects would be highly desirable. here we hypothesized that antigen receptor engineering of hematopoietic stem cells would fundamentally impact lymphoid progenitor cell fate and as a consequence biological properties thereby allowing to generate an ubiquitously available lymphoid cell product for targeted immunotherapy across mhc barriers. methods: murine hematopoietic stem cells were transduced with a broad panel of st and nd generation murine cars containing different costimulatory domains and numbers of active immune receptor-based activation motifs (itams) against cd in a lentiviral backbone and consecutively differentiated into lymphoid progenitors using the op -dl feeder layer system. resulting products were comparatively assessed in vitro and in vivo (facs, functional assays, microarray gene transcript analysis, western blotting, timely controlled transgene expression, leukemia challenge experiments, in vivo lymphoid depletion experiments) upon co-transplantation into a mhc-mismatched myeloablative transplantation model for relapsed leukemia. results: car expression on early ex vivo generated lymphoid progenitors suppresses bcl b ( figure b ) and leads to decreased t cell-associated gene expression. concomitantly, suppressed bcl b permits lymphoid progenitors to acquire nk cell-like properties ( figure a ) and upon adoptive transfer into hematopoietic stem cell transplant recipients they differentiate into carinduced killer cells (carik) that mediate potent antigendirected antileukemic activity across mhc barriers ( figure c ,d). a cd , but not - bb, costimulatory domain and active itams are critical for a functional carik phenotype. conclusions: these results give important insights into differentiation of lymphoid progenitors driven by synthetic car transgene-expression and inform the potential use of ex vivo generated carik as an "off-the-shelf" product for targeted immunotherapy. disclosure: m.v.d.b has ip licensing with seres therapeutics and juno therapeutics. m.v.d.b. has also received honorariums from flagship ventures, novartis, evelo, seres, jazz pharmaceuticals, therakos, amgen, merck & co, inc., acute leukemia forum (alf), and dkms medical council (board) and research support and has stock options with seres therapeutics. a.g. has received research support from aprea therapeutics and infinity therapeutics. all remaining authors have no conflict of interest. [[o image] . figure : car-induced killer (carik) cells provide strong anti-leukemia effects in vivo.] (a) c bl/ (b ) recipients received x b tcd-bm only or additionally with x syngeneic im z lymphoid progenitors. numbers of nk . + carik cells and frequencies of cd + tcrβ + progeny within the tom+ gate in spleens on day (im z , n= ; itom, n= ). statistics was performed by using students t-test (twotailed). data represent means ± s.e.m. (b) western blot analysis for bcl b in lysates from itom lymphoid progenitors, im z -lymphoid progenitors or b wt thymocytes. (c) recipients co-transplanted with either syngeneic (syn) or mhc class i and ii mismatched (allo) im z -expressing lymphoid progenitors (n= mice, respectively) received . x c -mcd leukemic cells on day after transplantation and monitored for survival. (d) car-lymphoid progenitor-co-transplanted mice were treated with weekly i.p. injections of either an anti-nk . antibody (clone: pk ; μg/dose) or with pbs for control (n= per group). all mice received . x c -mcd cells on day after transplantation and were monitored for survival. survival curves were compared using mantel-cox (log-rank) test. significant differences are indicated by *p < . , ***p < . , ****p < . . characterization of an hla-dpb specific t-cell receptor for adoptive immunotherapy sebastian klobuch , elisabeth neidlinger , carina mirbeth , wolfgang herr , , simone thomas , background: hla-dpb mismatches occur in up to % of allogeneic hematopoietic stem cell transplantations from hla / matched donors and were shown to be associated with a decreased risk of leukemia relapse. therefore, targeting hla-dpb mismatched antigens by donor t cells seems to be an attractive strategy to enhance graft-versusleukemia effect. we recently established a reliable method to generate and isolate hla-dpb mismatch reactive t cells receptors (tcr). tcr-modified t cells showed leukemia eradication of primary human aml blasts in a xenogeneic nod/scid/il rgc -/-(nsg) mouse model. however, human fibroblasts used as surrogate cells for graft-versus-host (gvh) reactivity were also recognized by hla-dpb -specific t cells upon ifn-γ pretreatment, which up-regulates hla-class ii expression on these cells. in this project, we aim at the isolation of tcrs recognizing mismatched hla-dpb only on hematopoietic cells, which might lower their risk for gvhd. methods: naive-enriched cd t cells were stimulated with autologous dendritic cells expressing allo-hla-dpb alleles upon rna transfection. to drive the outgrowth of cd t-cell populations expressing 'cd -independent' tcr which allow the redirection of both cd and cd t cells, we blocked the cd /hla interactions by the addition of cd binding antibody. allo-hla-dpb reactive cultures were analyzed for their recognition of primary aml blasts in ifn-γ elispot as well as chromium-release assays. highly reactive cd t cell populations were further analyzed for their ifn-γ secretion against nonhematopoietic cells. tcrs from most promising cd t cell clones with an hla-dpb specific recognition of hematopoietic but not non-hematopoietic cells were isolated and further analyzed. results: two cd -independent t cell clones with reactivity to the hla-dpb * : mismatch allele specifically lysed hla-dpb * : + primary aml blasts. most importantly, one of these t cell clones did not show ifn-γ secretion upon co-culture with ifn-γ pretreated primary fibroblasts. therefore, we isolated this tcr and transferred it into cd and cd t cells from healthy donors. tcr dp re-directed cd and cd t cells specifically recognized and lysed primary aml blasts from several hla-dpb * : + patients in vitro. again, cells of non-hematopoietic origin (fibroblasts) were not recognized even after ifn-γ pretreatment and hla-dp upregulation. to optimize tcr expression and activity, we exchanged the constant domains of the tcr by their murine counterparts. this modification not only led to a higher ifn-γ production and lysis of aml blasts, but also induced recognition of ifn-γ pretreated fibroblasts. however, tumor cell lines overexpressing hla-dpb * : were also recognized by cd t cells engineered with the wild-type or murinized tcr dp , suggesting that recognition of hematopoietic and non-hematopoietic cells is rather triggered by the avidity between the t cell and the target cell than by the tcr target epitope. conclusions: in conclusion, our data suggest that allo-hla-dpb specific tcrs are powerful therapeutic off-theshelf reagents in allogeneic t-cell therapy of leukemia. the isolation of allo-hla-dpb specific tcr without crossreactivity to non-hematopoietic cells might be one strategy to avoid hla-dpb specific gvh reactivity upon inflammatory situations (e.g. viral infections). however, our data also indicate that finding of the most suitable tcr candidate is challenging. disclosure: nothing to declare. abstract already published. infusion of memory t cell (cd ra-depleted) dli improves cmv-specific immune response early after abt cell-depleted hsct: first results of a prospective randomized trial background: abt cell depletion effectively prevents severe gvhd in mismatched hsct, but in a proportion of cases delayed immune recovery leads to increased infection risk and nrm. we've shown in a pilot study that infusion of low-dose memory t cells (cd -ra depleted) is safe after engraftment among recipients of ab t cell-depleted grafts (pmid: ). we initiated a prospective trial to directly test the efficiency of this approach. we report here an interim result of a prospective, randomized, single-centre trial (nct ). methods: a total of paediatric patients with malignant disorders (all, n= ; aml, n= ; nhl, n= ; acute mixed lineage leukemia, n= and mpd, n= ) were enrolled between october and september . patients were randomly assigned to receive cd ra-depleted dlis (experimental arm), n= , or not (control arm), n= . median age at hsct was . years, m:f ratio - : . the conditioning consisted of either treosulfan (n= ) or tbi (n= ) in combination with fludarabine and thiotepa. gvhd prophylaxis included tocilizumab at mg/kg at day , abatacept at mg/kg at day , + , + and + , and bortezomib at , mg/m at days - , - , + , + . neither antithymocyte globulin nor calcineurin inhibitors were used. donors were hla-haploidentical (n= ) or matched (n= ). all donors and % of the recipients were cmv seropositive. pmbc grafts were split and tcrαβ/cd depletion and cd ra depletion were performed with clinimacs prodigy. the median dose of cd + cells was x /kg, αβt cells - x /kg. in the experimental arm memory dlis were infused on day at x /kg and on days + , + , + , + at x /kg. in the control arm patients received dli after engraftment to prevent relapse (n= ) or treat infections (n= ). primary endpoints were the cumulative incidence (ci) of cmv viremia (> copies/ml) by day + and the ci of acute gvhd grade ≥ ii. results: median follow-up for survivors was year ( , - ). engraftment of wbc and platelets was achieved in pts, one patient died at day + . wbc and platelets engrafted at a median of days and days, respectively. the incidence of cmv viremia was % ( - ) overall, % ( - ) in the experimental arm vs. % ( - ) in the control arm, p=ns. the ci of agvhd ≥ grade ii was % ( - ) overall, % ( - ) in the experimental arm vs. % ( - ) in the control arm, p=ns. two patients died, one per treatment arm, resulting in % ( - ) ci of trm at year among the whole cohort. causes of death were preengraftment bloodstream infection and disseminated adenovirus infection. patients randomized to experimental arm acquired anti-cmv reactivity significantly earlier, according to ifn-g elispot assay on day + after hsct (p= , ). conclusions: co-infusion of donor-derived memory dli with the αβ t cell-depleted graft is safe and improves recovery of virus-specific immune responses. replacement of atg with targeted blockade of cd /cd costimulation and il- receptor does not compromise engraftment and gvhd control, and is associated with low rate of non-relapse mortality. [[o image] . disclosure: nothing to declare o conditioning prior to cd -specific car ( - z) t cells predicts response and survival in pediatric relapse/refractory (r/r) b-all background: cd -specific car t cells have demonstrated clinical benefit in patients with r/r b-cell acute lymphoblastic leukemia (b-all). several factors have been associated with response including conditioning chemotherapy, cd / ratio, and post infusion car t cell expansion. methods: we studied the feasibility of a multi-center trial of a msk-developed cd -specific car ( - z) for the treatment of r/r b-all, the toxicity following infusion, and performed predictor analysis for optimal response. pediatric and young adult patients with r/r b-all were eligible for infusion. patients received a cyclophosphamidebased (cy) conditioning of high dose (hd; g/m ) or low dose (ld; . g/m ) chemotherapy. outcomes of interest were complete response (cr), overall survival (os). variables considered were conditioning regimen (hd-cy vs ld-cy), pre-treatment disease burden (mrd vs morphologic), complete remission (cr) status, absolute lymphocyte count (alc) change, and in vivo car t cell expansion. results: patients were included; patients received hd-cy and received ld-cy prior to car t cell infusion. among evaluable patients (n= ), cr or cr with incomplete count recovery was demonstrated in % and % for hd-cy vs ld-cy cohorts respectively (p= · ). os was superior in the hd-cy cohort as compared to the ld-cy cohort (median os = not reached; nr) vs. · months (p= · ; figure ). lymphodepletion (delta alc: prior to and following cy) was significantly higher in the hd-cy cohort as compared to the ld-cy cohort (p< · ; figure ). the in vivo car t cell expansion (peak car t cell vector copy number/ml) in peripheral blood was higher in the hd-cy cohort as compared to the ld-cy cohort (p= · ; figure ). to less extent, disease burden prior to treatment with conditioning chemotherapy and car t cells impacted response. disease response was % ( / ) in low disease burden group (mrd-cohort) compared to % ( / ) in the high disease burden group (morphologic cohort) (p= · ). os was also superior in the low disease burden group (median os = nr) compared to high disease burden group (median os = . months; p= · ). combined response for hd-cy/mrd was % ( / ), hd-cy/morphologic % ( / ), ld-cy/mrd % ( / ), and ld-cy/morphologic % ( / ). grade iii/ iv toxicity occurred in % ( / ) of patients including severe cytokine release syndrome (scrs) in % of patients and severe car-associated neurotoxicity in % of patients. conclusions: in this preliminary analysis we demonstrate that dose intensity of conditioning chemotherapy positively correlated with response and overall survival for patients treated with car t cells and confirms, to a lesser extent, pre-treatment disease burden impacts both response and overall survival. clinical trial registry: nct disclosure: the authors acknowledge william lawrence and blanche hughes foundation provided funding for the conduct of this study with juno therapeutics providing funding for an extension cohort of patients. k.j.c. has received research support from juno therapeutics; has consulted, participated in advisory boards, or participated in educational seminars for juno therapeutics, and novartis. r. j.b. m.s. i.r. are co-founder, stock holders, and consultants for juno therapeutics. c.s.s. has received research support from juno therapeutics; has consulted or participated in advisory boards for juno therapeutics, kite and novartis. early and late hematologic toxicities in children and adults treated with cd -car t cells elad jacoby , , shalev fried , , abraham avigdor , , bella bielorai , , amilia meir , michal besser , , jacob schachter , , avichai shimoni , , arnon nagler , , , amos toren , background: autologous t cells transduced with cd directed chimeric antigen receptors show notable remission rate in advanced patients, leading to approval by the fda and ema for treatment of relapsed and refractory acute lymphoblastic leukemia (all) and non-hodgkin lymphoma (nhl). the most common adverse events reported are cytokine-release syndrome (crs) and neurotoxicity. here we study and profile of hematologic toxicity of patients following locally produced car t cells. methods: we studied the first patients treated on a phase b/ clinical trial using cd car-t cells for b-cell malignancies, focusing on hematologic toxicities (neutropenia, thrombocytopenia and anemia), from the initial lymphodepleting regimen till progression or an additional treatment was administered. cytokine levels were studied using milliplex map, human cytokine/chemokine panel ii. results: between july and march , patients were enrolled on the trial, and patients who received car-t cells and survived more than days were included in this analysis. neutropenia, thrombocytopenia and anemia occurred frequently ( %, % and %, respectively) after car t cell infusion, and were associated with a prolonged or biphasic nature: in % of patients hematologic toxicity occurred or were ongoing after days from cell infusion, and in % (neutropenia) and % (thrombocytopenia), two trough levels were noted, the second trough occurring after day + . later events of cytopenia, following more than days from car infusion and in absence of further therapy, occurred in % (neutropenia), % (thrombocytopenia) and % (anemia requiring prbc transfusion) of patients. we identified a strong correlation between the late hematologic toxicities (thrombocytopenia and neutropenia, p= . , thrombocytopenia and anemia, p< . , anemia and neutropenia p= . ). on univariate analysis, factors affecting late cytopenia were prior hsct (p= . , . and . for anemia, thrombocytopenia and neutropenia respectively) and higher crs grade p= . , . and . for late anemia, thrombocytopenia and anemia respectively). diagnosis (all vs nhl) or age were not correlated to the incidence of early or late post-car cytopenia. to further study potential causes of late events in patients who were in remission, and in absence of signs of hemophagocytosis, patients' serum was analyzed for chemokine panel at different time points. as expected, serum thrombopoietin levels were correlated with the platelet count. we observed that only in late events (more than days from infusion) sdf- serum levels correlated to neutrophil count (r = . , p= . ). conclusions: cytopenia are common events following cd car t cell therapy, and may have a prolonged and even biphasic course. patients at risk include those following a recent hsct or with high grade crs. late neutropenia events which occurred later than expected recovery from conditioning chemotherapy and following resolution of crs, were correlated with serum sdf levels, similar to prior observations with late onset neutropenia related to rituximab (dunleavy, blood background: acute myeloblastic leukemia (aml) represents % of all leukemias of western countries, being the second malignant hemopathy in pediatric population. in the last decades, the survival rate has maintained around %, being relapse the main problem. it has been highlighted the role of immune system for the control of aml and new therapeutic strategies have been developed. in this setting, the use of alloreactive natural killer (nk) cells could play a key role not only in the context of hematopoietic stem cell transplantation (hsct) but also as adoptive immunotherapy in patients with minimal residual disease. in this project we proposed including the cellular therapy with haploidentical activated and expanded nk cells as adjuvant therapy in pediatric patients affected by aml in complete remission and without indication of hsct. methods: it is a multicentre, open, prospective and no randomized phase ii clinical trial, to evaluate the efficacy and safety of allogenic nk cells infusion from haploidentical donor after a lympho-ablative chemotherapy in pediatric patients affected by low and intermediate risk aml in first complete cytological remission. patients were treated according to spanish protocol (sehop ) including inductions plus consolidation cycles. after chemotherapy patients received the infusion of activated and expanded nk cells (nkae) after a lympho-ablative treatment based on cyclophosphamide mg/kg and fludarabine mg/kg. nk cells were obtained from ml of peripheral blood from haploidentical donors selected based on alloreactivity of kir inhibitors and kir activators receptors. nk cells were activated and expanded for weeks trough co-culture with the cellular line k -mbil - bbl suitable for clinical use in humans (good manufacturing practices, gmp). results: at this time products in patients have been infused, with the following characteristics: nk cells (cd + cd + cd -) . ± . %; t lymphocytes (cd + cd -cd + ) . ± . %. one product was rejected for quality criteria. a mean of . ± . x nk cells/kg and . ± . x t cells/kg were infused. median age of the patients were years ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) . treatment and infusions were well tolerated. the main adverse event was neutropenic fever in one patient. after a median follow up of days ( - ) all the patients are alive. only one patient showed relapse at day + after nk cells infusion and was submitted to an hsct from a matched unrelated donor. conclusions: the preliminary results of this trial suggest that nk cells infusion as consolidative strategy in pediatric patients with aml, is feasible and secure. the therapeutic effect should be confirmed in a larger cohort of patients. clinical trial registry: clinicaltrials.gov (nct ) and eudract ( - - ) . disclosure: authors have no conflict of interest. the trial was funded by "fundación mutua madrileña" grant o tcrαβ/cd depleted and il- stimulated donor cell infusions can exert anti-tumor effects after allosct without inducing gvhd in vitro and in vivo background: natural killer (nk) cells and γδ t-cells have been shown to play a significant role in gvl effects after allogeneic stem cell transplantation (sct). both cell types are not restricted by mhc molecules which makes them unlikely to elicit graft versus host disease (gvhd) even in mismatched sct and therefore are suited also for cell-based posttransplant immunotherapy. incubation with cytokines strongly enhances their spontaneous and antibody dependent cellular cytotoxicity (adcc). methods: thus, we investigated the efficacy of a combination of il- stimulated nk-and γδ t-cells from mismatched donors in vitro and based on urgent medical need in ultra high risk patients. results: in vitro data: spontaneous cytotoxic activity and adcc with tcrαβ/cd depleted pbmcs from healthy donors against ls (neuroblastoma) and nalm- (leukemia) target cells after overnight stimulation with il- was significantly increased compared to non-stimulated cells (e: t ratio : ; ls cells: % and % vs %; nalm- cells: % and % vs %). the results for cd a assays are in line with the cytotoxicity approaches. nk and γδ t-cells showed a significantly stronger stimulation after il- incubation +/-adcc, as measured by secreting cytokines (cd a, infγ, tnfα). moreover, patients at extremely high risk for relapse received nk-/ γδ t-cell infusions after previous transplantation from haploidentical (n= ) or matched donors (n= ); diagnoses: t-all, relapse after st sct n= ; b-lineage-all, cr after nd sct, n= ; aml, relapse after st sct, n= ; relapsed neuroblastoma, pr after nd sct, n= . for that purpose magnetic microbeads and the clin-imacs ® device were used for tcrαβ/cd depletion of leukapheresis products from the original donors. afterwards, remaining cells (nk, γδ t and myeloic cells) were stimulated with il- ( ng/ml) overnight. efficacy of the stimulation procedure was measured in leukapheresis products: after incubation with target cells (ls) both, nkcells as well as γδ t-cells showed a significant increase in cd a secretion compared to non-stimulated effectors (n= ) from healthy donors (nk-cells: % vs %; γδ tcells: % vs %). a median number of . x nk-cells and . x γδ t-cells and . x residual tcrαβ cells/ kg were infused. four patients had received therapeutic mabs (anti-gd , anti-cd or anti-cd ) within hours after cell infusion to induce adcc. side effects were treatable cytokine release syndrome in patients; no gvhd occurred. infused cells could be tracked by cd expression for up to days in peripheral blood. outcome: patients with t-all and neuroblastoma responded and achieved another cr, patients maintained their previous cr for up to now / days; patients progressed/relapsed after / / days. additional therapies were applied in / patients. patients are alive (median follow up: days after infusion), / are disease free. conclusions: infusion of a combination of nk/ γδ tcells can increase anti-tumor effects and adcc with appropriate therapeutic mabs without inducing gvhd even after mismatched sct. further investigations are needed to evaluate the role of this cellular therapy. disclosure: nothing to declare o abstract already published. identifying chimeric antigen receptor (car) centers and car activity in europe, a survey on behalf of the cellular therapy & immunobiology working-party (ctiwp) of ebmt background: the use of chimeric antigen receptor (car) t cells has shown outstanding efficacy in patients with relapsed/refractory b-cell lymphoid malignancies, as reported by many groups in united states (us) and china. car-t cell based activity in europe is still at early stages. few european academic and pharma-sponsored clinical trials are currently opened to inclusions, and access to the two ema-approved autologous car-t products remains limited for the majority of european centers, since they must undergo a tedious and non-harmonized qualification process imposed by the manufacturer as part of the drug label. improved awareness of car-t centers and ongoing car-t activities in europe are essential to promote european networking activities, improve competitiveness of eu medical centers, and develop clinical trials, education, accreditation, and registration/long-term follow-up, thus fulfilling the needs of stakeholders. methods: a survey questionnaire was prepared by the committee members of the ebmt cellular-therapy-&-immunobiology-working-party (ctiwp). questions were formulated to identify car-t centers and car-t based activity in europe, and to know the characteristics of centers and the organization for implementing a clinical car-t program. using monkey survey, the survey was sent to ebmt-centers. results: centers replied to the survey. patients (all %, nhl %, mm %, cll %, aml . %, solid tumors %) have been treated so far in ebmt centers from countries. most of these centers are jacie-accredited ( % for clinical activities; % for cell collection activities). additional centers reported plans to start car-t cell administration within the following -months. most patients were treated in the setting of academic clinical trials ( . %l) or pharma sponsored trials ( %l). few patients ( %) received marketed car-t cells. pcr ( %) and flow-cytometry ( %) were used to monitoring the persistence of car-t cells after infusion. cytokine levels were measured in % of the cases, and ngs was performed in % of the centers. half of the centers reported to use "point of care" manufacturing. patient management was mainly carried out by bone marrow transplant (bmt) physicians ( %). however, % of the centers reported to have also implemented a "car-t team", as a multidisciplinary team managed by a coordinator, with weekly clinical meetings. in more than % of the centers, there were specific rules for transferring patients to the intensive-care-unit. in contrast, only % of the centers had a nurse coordinator for management assistance of these patients. % of the centers plan to expand the capacity of their own department for car-t program. conclusions: this survey is the first attempt to gather information about car-t cell activity in europe. this first results confirms active and growing involvement and high quality standards of european centers having car-t cell program. the expanding use of these complex atmp approaches could be facilitated through harmonization of the clinical practice, shared analysis of good quality data, and by a centralized european clinical trial office. the cellular therapy car-t form is readily available to capture in more detail safety and efficacy of this intervention and will allow sharing in a transparent process registry data with stakeholders. methods: this is an open-label, phase study (chictr ). eligible patients had rrmm, confirmed as bcma-positive by flow cytometry or pathological examination. autologous t cells transduced with a bcmatargeted chimeric antigen receptor (car-bcma t cells) are used( × ^ /kg car+t cells), following pre-infusion treatment--intravenous administration and lymphodepletion conditioning (fludarabine and cyclophosphamide). the primary outcome measure is incidence of adverse events (aes) especially clinical events, including crs, neurotoxicity and coagulopathy, et al. additional outcome measures are duration and expansion of bcma car t cells. results: patients (median age of , range to ) with rrmm have received treatment. the median %pc in bm at enrollment was . % (range % to %). / had high tumor burden, defined as ≥ % bone marrow plasma cells. / had prior autologous stem cell transplant. as of data cut-off (dec , ), crs grade - occurred in patients, grade - occurred in . surprisingly, not a few patients developed coagulation disfunction, manifesting elevated d-dimer, prolonged aptt, pt, tt, and decreased fibrinogen, with case of organ bleeding. laboratory values correlating with crs reaching grade - (n= ) compared to those with milder crs (n= ) included peak ferritin (mean: . vs . ug/l, p< . ), peak il- (mean: . vs . pg/ml, p< . ), peak il- (mean: . vs . pg/ml, p< . ), and peak ifn-γ(mean: . vs . pg/ml, p< . ). no significant difference was seen in peak crp, il- and tnf-α. patients with crs - had a higher △aptt (mean: . vs . s, p< . ), △d-dimer (mean: . vs . ug/l, p< . ) and △fibrinogen (mean: - . vs - . g/l, p< . ) compared to those with milder crs. there was no significant difference in △pt and △tt.the changes of d-dimer and aptt were in line with the indictors of crs, such as crp, il- and il- in some individual cases. the significant correlation between aptt and crp (p< . ) appeared on patients ( . %), while the significant correlation between il- and aptt as well as between il- and d-dimer only appeared on patients ( . %). conclusions: early safety and efficacy results of bcma car t therapy in rrmm are encouraging. aes of coagulation were observed. changes of aptt, d-dimer and fibrinogen of patients with high crs level were more significant than those with milder crs. on the individual level, aptt correlated with crp in the course of therapy. given extensive cross-talk between inflammation and coagulation, coagulation-related indictors are more convenient for monitoring crs. also, our study suggests the importance in diagnosis and early management for coagulopathy to avoid crs-related mortality. clinical trial registry: chictr http:// www.chictr.org.cn/abouten.aspx disclosure: nothing to declare o abstract already published. point-of-care production of cd car-t cells in an automated closed-system bioreactor: report of first clinical experience background: cd car-t cell products are approved as therapy for advanced b-cell malignancies. the predominant manufacturing model is a centralized industrial-type production process. an alternative approach to car-t cell production and delivery to the patient is via a point-of care manufacturing process. methods: between february to november a total of pts with relapsed/refractory b-cell malignancies ( female, male[gr ], median age y) were screened, pts were enrolled for single center, phase i-ii trial of safety and clinical efficacy of automatically produced cell therapy product, were eligible for compassionate use of therapy. seven patients had relapsed b-all after hsct, pts had refractory relapse after chemotherapy, pt had refractory induction failure, pt had refractory primary mediastinal bcell lymphoma (pmbcl). eight patients received previous blinatumomab infusion. eight patients had high disease burden > % blast cells in bm, median % ( - ), pts had minimal residual disease (mrd) in bm. the clinimacs prodigy t cell transduction (tct) process was used to produce cd car-t cells from fresh patientderived leukapheresis product. automatic production included cd /cd selection, stimulation with macs gmp t cell transact, transduction with lentiviral (second generation cd . - bb zeta) vector (lentigen, miltenyi biotec company) and expansion over days in the presence of texmacs gmp medium and macs gmp il- /il- combination. final product was administered without cryopreservation after fludarabine/cyclophosphamide preconditioning. all patients received prophylactic tocilizumab at mg/kg. results: all production cycles were successful. median transduction efficiency was % ( - ). median expansion of t cells was fold ( - ). cd /cd ratio in the final product was , . the cell products were administered at * /kg of car-t cells in pts, * /kg in pts, * /kg in pts. cytokine release syndrome occurred in ( %) pts: grade i in pts, grade ii in pts, grade iv in pts. car-t cell related encephalopathy occurred in ( %) pts, including one fatal brain edema. grade i neurotoxicity had pts, grade ii - pt, grade v - pt. four patients were admitted to the intensive care unit (icu). three patients died until day ( due to sepsis, due to fatal brain edema and sepsis, on autopsy in the brain vessels of this patient were found klebsiella pneumoniae emboli) twelve patients were evaluable for response at day . three pts had persistent leukemia, without evidence of car-t expansion. mrd-negative responses were achieved in cases among evaluable cases with bone marrow involvement. patient with pmbcl had a decrease in metabolic activity on pet/ct scan. cd (-) relapse after initial response was registered in case at day. conclusions: production of car-t cells with the clinimacs prodigy tct process is a robust option that provides a point-of-care manufacturing approach to enable rapid and flexible delivery of car-t cells to patients in need. robustness and consistency of this approach provides a solid basis for multi-center academic trials in the field of adoptive cell therapy. disclosure: nothing to declare background: in the last decade, several prognostic scoring models such as the international prognostic scoring system (ipss), the dynamic ipss (dipss) or dipss-plus have been developed for diagnosed primary myelofibrosis (pmf) and are currently used for decision finding regarding allogeneic stem cell transplantation. furthermore, the prognostic relevance of mutation profile resulted in a mutation-enhanced system (mipss ) in transplant-age pmf patients ( years or younger). for secondary myelofibrosis, the mysec-pm was developed. while all scoring systems were developed in nontransplant populations and may be useful in decision finding regarding transplantation, uncertainty remains regarding usefulness of these systems to predict posttransplant outcome and thus counseling patients whether to proceed to transplant. methods: bone marrow or peripheral blood samples were obtained before transplantation and mutations were detected using next-generation sequencing. the following myelofibrosis-associated genes were sequenced: jak , calr, mpl, asxl , idh / , cbl, dnmt a, tet , sf b , srsf , u af , ezh , tp , nras, kras, runx , and flt . cytogenetic analysis and reporting were performed according to the international system for human cytogenetic nomenclature criteria using standardized techniques. we examined myelofibrosis patients, of whom had pmf and smf at time of transplantation. a training cohort of patients was used to create a clinical-molecular transplant scoring system (mtss) predicting survival from cox models, internally validated by use of bootstrap and cross-validation. model discrimination was measured by the concordance index (c). the final mtss was externally validated in a cohort of patients and was furthermore applied to posttransplant non-relapse mortality as a secondary objective. results: multivariable analysis on -year survival identified age ≥ years, karnofsky performance status < %, platelet count < x /l and leukocyte count > x /l at time of transplantation, hla-mismatched unrelated donor, asxl mutated and calr-/mpl-unmutated genotype being independent prognostic factors for outcome. the uncorrected concordance index for the final survival model was . ( . - . ), and bias-corrected indices were similar. a weighted score of was assigned to transplantation from an hla-mismatched unrelated donor and an calr-/mplunmutated genotype, whereas a score of was assigned to older age, leukocytosis, thrombocytopenia, asxl mutation, and poor performance status. the mtss consisted of four distinct risk groups showing -year survival in the validation cohort of % ( - %) for low (score - ), % ( - %) for intermediate (score [ ] [ ] , % ( - %) for high (score ), and % ( - %) for very high risk (score > ), respectively (p < . ). increasing score was predictive of non-relapse mortality (p < . ) and remained applicable to pmf ( conclusions: we show here that this internally and externally validated mtss accurately discriminated different risk for death and may improve counseling patients with respect to transplant compared with currently existing systems, as well as facilitate design of clinical trials in the transplant setting. disclosure: nothing to declare. response to up-front azacitidine in juvenile myelomonocytic leukemia (jmml): interim analysis of the prospective european multicenter study aza-jmml- background: jmml is a chemotherapy-resistant neoplasia of early childhood. allogeneic hematopoietic stem cell transplantation (hsct) is the only curative therapy, saving approximately % of these children. relapse is the major cause of treatment failure, with chemotherapy prior to hsct being notably unsuccessful. novel therapies controlling the disorder prior to hsct are urgently needed. methods: we conducted a phase , multicenter, open-label study to evaluate pharmacodynamics, safety, and antileukemic activity of azacitidine monotherapy prior to hsct in patients with newly diagnosed jmml. azacitidine was administered at mg/m /day intravenously on days - of a -day cycle for to cycles. the primary endpoint was the number of patients with clinical complete remission or clinical partial remission (cpr) at cycle day (c d ); secondary endpoints included overall survival following hsct. results: eighteen jmml patients ( ptpn -, nras-, kras-, nf -mutated) aged . - . years were enrolled. median (range) wbc, platelet count and spleen size were: . ( . - . ) × /l, ( - ) × /l, and ( - ) cm below the costal margin, respectively. dna methylation class (lipka et al. nat comm ; n= ) was high, intermediate, or low in , , and patients, respectively. sixteen patients completed cycles of therapy and of them completed cycles. two patients discontinued treatment before completing cycles due to disease progression. six patients ( %) experienced ≥ grade or manageable adverse event, consistent with the known azacitidine safety profile. eleven patients ( %) achieved cpr at c d and had progressive disease either at c d or prior. importantly, of the patients who needed platelet transfusions before or shortly after treatment initiation did not require transfusions at the time of hsct. seven of these platelet responders had normalized their platelet count (≥ × /l). palpable spleen size decreased in responders by a median of . cm after cycles and ranged from - cm below the costal margin after cycles. sixteen patients received allo-hsct from a family or compatible unrelated donor following a busulfan-(n= ) or treosulfan-based (n= ) preparative regimen after a median of days ( - ) from last azacitidine dose. thirteen transplanted patients were leukemia-free at median follow-up of . months ( . - . ) after hsct. two children (both high methylation class) given hsct relapsed after the allograft. sixteen of the patients were alive at a median follow-up of . months ( . - . ). one patient who discontinued treatment before cycle died from disease progression, and non-responder died from graft failure. conclusions: this study shows azacitidine monotherapy was well tolerated in children with newly diagnosed jmml. although the long-term advantage of azacitidine therapy remains to be fully assessed, both the decrease in spleen size and significant platelet responses observed demonstrate that the drug was effective in jmml and provided clinical benefit to jmml patients in this study. this clinical trial has shown that azacitidine therapy prior to hsct may be considered for patients with jmml. clinical trial registry: nct disclosure: charlotte niemeyer is a member of a board of directors or advisory committee and provides consultancy for celgene. claudia rössig is a member of a board of directors or advisory committee at amgen, eusapharm, roche, celgene, novartis, pfizer, bms; honoraria from amgen, roche, pfizer. andré baruchel provides consultancy (includes expert testimony) with celgene, novartis, servier, jazz pharma; research funding and honoraria from shire; honoraria from novartis, jazz pharma. susana rives has received honoraria (for talks in industria sponsored satellite symposia) from novartis, jazz pharma, baxalta, shire, servier; speaker's bureau at novartis, jazz pharma, baxalta, shire, servier, amgen, erytech pharma; membership on an entity´s board of directors or advisory committees at novartis; received travel and accommodation expenses for medical congresses from novartis, jazz pharma, baxalta, shire, servier, amgen, erytech pharma. marco zecca has received honoraria from chimerix and jazz pharma. marry m. van den heuvel-eibrink received honoraria from celgene (consultation fee). bouchra benettaib, noha biserna, jennifer poon, mathew simcock, meera patturajan are employees of and hold stock or other equity ownership in celgene. christian flotho, daniel lipka, jan starý, karsten nysom, gérard michel, thomas kilngebiel, franco locatelli, giuseppe basso, concetta micalizzi, irith baumann, markus schmugge liner have nothing to disclose. ruxolitinib before allogeneic hematopoietic stem cell transplantation (hsct) in patients with myelofibrosis: a preliminary descriptive report of the jak allo phase ii study marie robin , , raphael porcher , corentin orvain , background: allogeneic hematopoietic stem cell transplantation (hsct) is the only curative treatment for patients with myelofibrosis; os is from to % depending on age, comorbidities, disease status and type of donors. jak / inhibitors have been reported to decrease constitutional symptoms and spleen size in one half of patients with a possible advantage of os as compared to best current treatment. retrospective studies show that ruxolitinib has been used in patients before hsct with apparent good tolerance. methods: in , we initiated a phase ii prospective french collaborative (filo and sfgm-tc) trial testing the role of ruxolitinib given before allogeneic hematopoietic transplantation in patients with myelofibrosis. patients could be included if they were intermediate or high risk according to lille score or ipss. patients had to start ruxolitinib after inclusion and were transplanted in case a donor has been identified within months. primary aim was progression-free survival at one year after hsct. results: patients could be included. ( %) had a primary myelofibrosis and ( %) had a myelofibrosis secondary to essential thrombocythemia (n= ) or polycythemia vera (n= ). at time of inclusion, ( %) patients had constitutional symptoms, ( %) had palpable splenomegaly, ( %) patients had hemoglobin < gr/dl, ( %) patients had peripheral blast cell count > or = at % and ( %) had thrombocytopenia < g/l. median follow-up was months. at months, one patient has died, patients had no donor, had an hla matched sibling donor, had an hla matched unrelated donor and had a / hla mismatched unrelated donor. ( %) patients with a donor at months could be transplanted of whom underwent a splenectomy before transplantation. conditioning regimen was fludarabine mg/m in combination with melphaln mg/m . gvhd prophylaxis was cyclosporine and mycophenolate mofetil with atg in unrelated donor. partial response under ruxolitinib was % while the whole complete response incidence was % at one year. os and pfs at years were % ( % ci: - ) and % ( % ci: - ). os was significantly better in patients with hla matched sibling donor as compared to patients without a donor or with an unrelated donor (p= . , figure ). cumulative incidence of acute gvhd on day was % of whom % had grade - acute gvhd. cardiogenic shock generally associated with severe sepsis or "sepsis like" syndrome occurred in patients: patients were not transplanted because of this complication while in the other patients cs occurred within the days after transplantation. three renal failures secondary to tumor lysis syndrome were declared just after conditioning regimen initiation. figure . overall survival in jak allo patients. conclusions: os was very good in patients with hla matched related donor and significantly better than in patients with an unrelated donor. more analyses are currently ongoing to determinate the potential role of cytokine release in peri-transplantation time, as well as specific myelofibrosis and quality of life questionnaires assessing the impact of ruxolitinib in patients. background: while cytogenetics may influence outcome in primary myelofibrosis (pmf) from diagnosis which lead to several prognostic systems incorporating different cytogenetic risk classifications, its definitive role specific after allogeneic stem cell transplantation is still unclear. here, we aim to evaluate the role of currently existing cytogenetic risk classifications included in the dynamic international prognostic scoring system (dipss)-plus and in the mutation-enhanced system (mipss -plus version . ). methods: in this multicenter retrospective study, we examined pmf patients undergoing allogeneic stem cell transplantation. current cytogenetic risk stratifications used in dipss-plus and mipss -plus version . were evaluated. according to dipss-plus, an unfavorable karyotype includes + , - / q, i( )q, - / q, p-, inv ( ), and q rearrangements while mipss -plus version . consisted of very high risk (vhr: single/multiple abnormalities of - , i( q), inv( )/ q , p-/ p . , q-/ q , + , or other autosomal trisomy, not including + /+ ), favorable (normal karyotype or sole abnormalities of q-, + , q-, chromosome translocation/duplication or sex chromosome abnormality including -y), and unfavorable (all other abnormalities). results: the median follow-up period of all pmf patients was months and the median time from diagnosis to transplant was months. after five years, ( %) patients had died. the median age at transplant was years and % of patients were male. most allografts were applied using peripheral blood ( %) and were received from an hla-matched unrelated donor ( %), followed by mismatched unrelated ( %), matched related ( %), and mismatched related or haploidentical ( %). reduced intensity conditioning was applied to % of patients. splenectomy before transplant was undergone in % and ruxolitinib was received by %. frequencies according to driver mutation genotype were: calr ( %), jak ( %), mpl ( %), and triple negative ( %). asxl mutation was present in % while % had more than two mutations overall. ( %) patients had normal karyotype. most frequent abnormalities were trisomy in ( %), trisomy in ( %), deletion in q in ( %), deletion in q in ( %), chromosome in ( %), chromosome in ( %), and chromosome translocation/duplication in ( %). more than two abnormalities were present in ( %) patients. high cytogenetic risk category according to dipss-plus was present in ( %) while ( %) had vhr and ( %) had unfavorable risk according to mipss -plus version . . regarding outcome, an unfavorable karyotype according to dipss-plus showed year os of % ( - %) vs % ( - %), with causespecific hazard being . (p= . ). -year relapse and nonrelapse mortality rates were not significantly different showing % and % for unfavorable karyotype vs % and % (p= . and . ). with respect to the three-tiered classification of mipss -plus version . , -year os of vhr was % ( - %) and unfavorable was % ( - %) vs favorable risk of % ( - %), with cause-specific hazard of . (p= . ). relapse and non-relapse mortality were % and % for vhr, % and % for unfavorable, and % and % for favorable risk (p= . and . ). conclusions: current cytogenetic risk stratifications do not predict outcome in pmf after allogeneic stem cell transplantation. [[o image] . background: the aim of this study was to assess the outcome of patients with myelofibrosis allografted before and after , in two transplant centers (genova san martino and rome gemelli). methods: we have studied patients, divided in two groups: - (n= ) and between and (n= ). the age was significantly older in the most recent group ( vs years, p< ), and there was a greater use of alternative donors ( % vs %, p< . ), and more patients with dipss-r high score ( % vs %, p= . ). the transplant score (based on transfusions > and splee size > cm) was intermediate-high risk in % and % (p< . ) of patients respectively. the conditioning regimen was a combination of thiotepa, busulfan fludarabine (tbf) in % and % of patients grafted before and after (p< . ). the transplant risk score (ts) was based on spleen size (> cm) and pre-graft transfusion (> ) as previously described (bone marrow transplant. mar; ( ) : - ) results: outcome. the cumulative incidence (ci) of acute grade ii-iv ( %) and of chronic gvhd ( %) was comparable in the two time periods. the year ci of non relapse mortality (nrm) was reduced overall from % to % (p= . ) and the year ci of relapse from % to % (p= . ). as a consequence the overall year actuarial survival has improved from % to % (p= . ). predictive factors. the following factors predicted survival in multivariate cox analysis: high risk transplant score (hr . , p= . ), high risk dipss-r (hr . , p= . ), the use of tbf (hr , , p= . ), alt donor (ht . m p= . ), donor age > years (hr . , p= . ) and abo match (hr . , p= . ). dipss and transplant score. when combining dipss-r (high) and transplant score (int -high) we could identify groups : score (dipss not high and ts not high) (n= ) score (either high dipss or high ts) (n= ), score (both dipss a ts high risk) (n= ) . nrm was %, % % in these group (p= . ), relapse was %, %, % (p= . ) and year survival was %, %, % (p< . ). in the current transplant era (> ) the year disease free survival of these groups is %, %, % (p< . ). abo matching further increases dfs for score patients. nrm mortality for these groups is currently %, %, % (p= . ). conclusions: the year survival of allografts in patients with myelofibrosis has improved overall from % before , to % beyond , despite the current use of % alt donors. predictive factors are the transplant score (transfusions and spleen size), dipss-r and the use of alkylating agents (tbf), the latter being the major change in the transplant eras. patients received ruxolitinib before the transplantation, were transfusion-dependent. all the patients underwent pbsc infusion, except one who received bmsc. in cases the source was a sibling donor, in a mud, in a hla-mismatched ud ( mismatch). conditioning regimen was mostly based on the combination of singlealkylating agent and fludarabine ( bu/flu; mel/flu/ bcnu; mel/flu; treo/flu) with atg infusion. six patients received a fully myeloablative schema, reduced-intensity conditioning. only one patient received conditioning with tbi. results: before the transplant, a panel of molecular analysis based on next-generation-sequencing revealed, in addition to the mpl mutation, alterations in asxl ( %), srsf ( %), sf b ( . %), ezh ( %), idh ( . %), idh ( . %), tet ( . %), tp ( . %). after the transplant, the incidence of acute gvhd was %; only patients ( . %) experienced acute gvhd grade - . chronic gvhd was registered in % of cases ( patients: mild, moderate, severe). relapse occurred in only case. nrm incidence was . % of cases, occurring in the first year after transplant. with a median follow up of months, -year overall survival was . %, and -years pfs reported the same value, beeing the only relapse at > years after transplantation. the relapse occurred in the only patient who harbored mutations in both asxl and ezh genes. conclusions: these retrospective data suggest that the particular group of mpl-mutated myelofibrosis may have a good outcome after stem cells transplantation. in particular, our data revealed very low rate of disease relapse ( . %) in comparison with the available historical controls regarding myelofibrosis in toto. [ background: a significant proportion of cml patients undergoing allogeneic stem cell transplantation (allo-hsct) restart tki following transplant to prevent relapse. there is however no data to support if tki can be discontinued following allo-hsct and whether such patients can safely discontinue tki remains controversial. practices varies among transplant centres depending on countrywide practices, centre experience, duration of molecular remission, patients 'wish and analysis of retrospective data on the outcome of patients who discontinue tki after transplant may provide further insight to help elaborating future guidelines. the present study objective is to investigate the outcome of patients with cml who discontinue tki therapy after restarting tki following allo-hsct. this retrospective study may be helpful to support future guidelines. methods: through the ebmt database of patients who received an allo-hsct between march and september , we identified cml patients who restarted tki treatment post allo-hsct and stopped it after at least months of tki therapy. results: out of patients who discontinued tki, were in first chronic phase (cp ) at the time of transplant, in second or third chronic phase (cp and cp ) and in accelerated or blastic phase (ap/bp) or primary refractory disease. one patient had missing data at the time to transplant. allo-hsct conditioning was of reduced intensity (ric) in patients and myeloablative (mac) in patients, including tbi in patients. tki therapy was (re)started in all patients after a median time from transplant of . months (range, . to . months) for a duration of . (range, . - . months). after a median time from tki discontinuation of . months (range . - . ), -years progression free survival (pfs) and overall survival (os) were % ( %ci to %) and % ( %ci to %) respectively. patients in cp at the time of transplant had a significantly higher -years os compared to those in cp / cp or ap/bp, % vs % ( %ci to %) and % ( %ci to %) respectively (p< . , figure ). causes of death (cod) in the cp / and ap/bp groups were relapse ( / and / respectively) and nrm ( / and / respectively, missing cod in each group). conclusions: post-transplant tki discontinuation appears safe in patients who receive an allo-hsct while still in first chronic phase. however, such approach in patients who transform to advanced phases before allo-hsct remains a matter of concerns given the high incidence of post allo-hsct relapse. further analysis to identify reason for restating tki post-transplant and for subsequent discontinuation will be performed after further data is collected through the ongoing data quality initiative (dqi) in cml. [ background: allogeneic stem-cell transplantation (allo-sct) is a well-established treatment modality for high-risk hematopoietic malignancies. however, the optimal intensity of myeloablation with a reduced-toxicity conditioning regimen to decrease relapse rate after allo-sct without increasing trm has not been well established. thiotepa is an alkylating compound with antineoplastic activity and immunosuppressive properties, as well as the ability to penetrate the blood brain barrier. thiotepa has become an integral part of the thiotepa, busulfan iv (busilvex), and fludarabine (tbf) conditioning regimen, which is being used with increasing frequency, particularly for haploidentical and cord-blood transplants. however, few studies have focused on analyzing the effect of thiotepa dose in the tbf conditioning. methods: the aim of this study was to evaluate the optimal dose of thiotepa, as part of the tbf conditioning for allo-sct in adults with aml transplanted in complete remission (cr), by comparing the transplantation outcomes of patients who received mg/kg thiotepa and days of busilvex ( . mg/kg) (t b f) versus those who received mg/kg thiotepa with days of busilvex (t b f) or days of busilvex ( . mg/kg) (t b f) using a large dataset from the ebmt registry. results: we identified aml patients allotransplanted between january and june from matched related or unrelated donors or t replete haplo-identical donors. patients ( %) received (t b f); patients ( %) received (t b f); the remaining patients ( %) received (t b f). all the patients were in cr at transplant. median follow-up was months (iqr: - ). outcomes are summarized in the table . acute gvhd grade ii-iv was %, % and % (p= . ) respectively. at years the non-relapse mortality (nrm) was %, % and % respectively (p= . ); the relapse incidence (ri) was %, % and % (p= . ) respectively; the leukemia free survival (lfs) was % vs % vs % (p= . ) respectively and the overall survival (os) was % vs % vs % (p= . ) in the groups, respectively. the year grfs was %, %, and % respectively (p= . ). in multivariate analysis, acute gvhd was higher for patients receiving t b f (p= . ; hr . ) or t b f (p= . ; hr . ) as well as for patients receiving transplant from haploidentical donor or peripheral blood stem cells, whereas nrm was higher for older patients (p= . ; hr . ), patients receiving t b f (p= . ; hr . ) or haploidentical transplant (p= . ; hr . ). importantly, os was lower for older patients (p= . ; hr . ) or for patients receiving t b f (p= . ; hr . ). the multivariate analysis was adjusted to all the different factors between the groups. conclusions: t b f is associated with higher incidence of acute gvhd compared to t b f whereas t b f associated with higher nrm, a higher incidence of acute gvhd and a lower os compared to t b f. with the limitation of the retrospective nature of registry data, these results suggest that a lower dose-intensity of thiotepa and busilvex in the tbf regimen in general may yield better results in aml patients transplanted in complete remission. background: thethiotepa-busulfan-fludarabine (tbf) based conditioning regimen is widely used in t-cell repleted haploidentical transplantation (haplo) with post-transplant cyclophosphamide. however, the use of anti-thymocyte globulin (atg) has not been well established. it decreases the incidence of graft versus host disease however some claim that it's at the cost of increased relapse. we conducted this multi centric study to compare the outcomes of patients who underwent haplo with tbf conditioning regimen with atg to those without. methods: this is a multicentric retrospective study. data was collected from centers, the american university of beirut medical center, hospital saint antoine paris, institute paoli calmette marseille, and humanitas research hospital milan. we included all consecutive adult patients who underwent haplo with tbf conditioning. the conditioning consisted of thiotepa mg/kg per day infused on days - and/or - , fludarabine mg/m infused on day - to day - ; and busulfan mg/m infused on day - to day - . graft versus host disease (gvhd) prophylaxis consisted of post transplantation cyclophosphamide mg/ kg per day on day + and day + , cyclosporine on day + and readjusted according to level, and mycophenolate mofetil mg every hours beginning on days + to + or + depending on the center. patients who received atg received a dose of . mg/kg per day. results: we included a total of patients, of whom ( %) received atg (group ) as part of the conditioning chemotherapy. patients who received atg had a younger median age compared to the second group without atg (group ) ( and years respectively; p value . ). ( % vs %) of each group had acute leukemia, and ( % vs %) were in complete remission at the time of transplant, while patients ( %) in the group had progressive disease at transplant. patients ( . %) had an intermediate disease risk index (dri). in the atg group, patients ( %) compared to ( %) in the other group received mg/kg thiotepa, while ( ) and ( %) received mg/kg respectively. peripheral blood stem cells were the most common graft source in both groups ( % and % respectively). at a median follow-up of . months, patients receiving atg had a statistically significant decreased risk of acute graft versus host disease (agvhd) (rr . ; p value . ), and non-relapse mortality (nrm) at months (rr . ; p value . ). atg also resulted in higher progression and overall survival at months, which was not statistically significant ( . % and . %; p value . , with . % and . %; p value . respectively). conclusions: atg as part of the pre-transplantation conditioning leads to significant reduction in agvhd and nrm at months without significant effects on pfs or os. disclosure background: high-risk leukemia is associated with poor prognosis and inferior outcome. in elderly or comorbid patients allogeneic sct with myeloablativ conditioning regimen as the most effective treatment option is not available. sequential regimen combining cytoreductive therapy with ric has shown high antileukemic activity for high-risk patients with acceptable toxicity profile. this study is based on the observation that antileukemic effects have been described previously for the nucleoside analogue clofarabine. methods: the trial was designed as an investigatorinitiated prospective, multicenter, open-label, two-arm, parallel-group phase ii study comparing clarac to flamsa regimen. flamsa regimen consists of fludarabine ( mg/ m , days - to - ), amsacrine ( mg/m , days - to - ) and cytarabine ( mg/m , days - to - ). clarac regimen consists of clofarabine ( mg/m , days - to - ) and cytarabine ( mg/m , days - to - ). both cytoreductive therapies were combined with bu/cy (busulfan, x . mg/kg, days - to - and cyclophosphamide mg/kg, days - to - ). as gvhd-prophylaxis atg, csa and mmf were used. patients with high risk aml or advanced mds (ipss ≥ int ) with contraindication for conventional conditioning or refractory to induction therapy were eligible. primary objective was to demonstrate that event-free survival is improved by clarac instead of the flamsa. secondary objectives were overall and relapse-free survival, mortality rate, safety profiles and cardiac toxicity. results: between and , patients were recruited, patients did not meet the in-/exclusion criteria. a total of were randomized with patients each in the clarac and flamsa group. mean time to event was . ± . days for flamsa and . ± . days for clarac, respectively (p= . , figure ). in total of the adverse events were serious with fatal outcome of patients in the clarac and patients in the flamsa group. cardiac toxicity was observed in patients in the clarac treatment arm, whereas patients were affected in the flamsa treatment arm (p= . ). overall survival for clarac was numerically, but not statistically inferior to flamsa (p= . ). a part of / ( . %) patients died until the end of the study in the clarac treatment arm, whereas only / ( . %) died in the flamsa treatment arm (p= . ). conclusions: this study did compare two different conditioning regimens for allogeneic stem cell recipients with high risk aml/mds. patients have been included and were randomized. the treatment arms were wellbalanced at study baseline for relevant covariates. superiority of the clarac treatment regimen over the flamsa regimen could not be demonstrated. consistently hazard ratios for event free survival, overall survival and relapsefree survival were in favor of the control group with flamsa treatment. no differences were found regarding cardiac toxicity, rate of engraftment, or chimerism. regarding general safety parameters, no relevant differences between the two treatment strategies were found. clinical trial registry: background: currently, there is no direct evidence to recommend specific conditioning intensities in myelofibrosis undergoing allogeneic stem cell transplantation. moreover, recent risk stratifications for diagnosed myelofibrosis (mf) included specific mutation profiles as prognostic factors. using clinical-molecular data from four different centers from germany and france, we sought to determine whether molecular genetics have an impact on outcome after reduced intensity (ric) and myeloablative conditioning (mac) stem cell transplantation in mf. methods: previously published methods were used to sequence myelofibrosis-associated genes (i.a. calr, jak , mpl, asxl , srsf , ezh , idh / , dnmt a, tet , tp ). risk stratifications according to dynamic international prognostic scoring system (dipss), mutation-enhanced system (mipss ), genetic inspired prognostic system (gipss), prognostic model for secondary myelofibrosis (mysec-pm), cytogenetics as well as other clinical and transplant-specific variables were included in analyses. cox model with hazard ratios (hr) was used for survival (os) and cumulative incidence for relapse and non-relapse mortality (nrm). risk ratios (rr) were obtained for subgroup analysis. results: the study included mf patients ( primary and secondary mf) of whom received ric and mac. the median follow-up was months in ric and months in mac and the median age was and years. patients receiving ric were at higher risk according to dipss, mipss , and mysec-pm, whereas frequencies of driver mutation genotype (calr, jak , mpl, triple negative) as well as asxl mutation were similar. hla-mismatched unrelated donors were more frequent in ric. most common conditioning regimens for ric were bu/flu ( %) and flamsa ( %), and flu/mel ( %), treo/flu and tbi/flu ( %, respectively) for mac. no significant difference was found for ric versus mac with respect to os ( % vs %; p= . ), relapse ( % vs %; p= . ) or nrm ( % vs %; p= . ). early relapse within five months was increased after ric ( % vs %). including molecular variables, ric showed higher but not significantly different os rates in patients with < mutations, in triple negative driver mutation genotype, asxl mutation ( figure ). when evaluating patients with an asxl mutation, those patients who had only one asxl or one additional mutation seemed to benefit from ric showing -year os of % vs % for mac (p= . ), whereas no difference was identified when more than two additional mutations were present ( % vs %; p= . ) . furthermore, in patients with an asxl mutation and one additional driver mutation (calr, mpl, jak ), year os was significantly different showing % in ric vs % in mac (p= . ). regarding current risk stratifications, ric showed significantly improved os only in high risk dipss and mysec-pm, whereas no difference was found regarding the remaining systems such as dipss-plus, mipss and gipss ( figure ). conclusions: the evaluation of different conditioning intensities in mf did not favor ric or mac regarding currently existing risk stratifications. with respect to molecular genetics, a proportion of patients specifically harboring up to two mutations including asxl may benefit from ric with respect to os. [ background: sequential conditioning regimens (sr) have shown substantial activity in relapsed/refractory acute myeloid leukemia (r/raml). the original prototype sr is the flamsa-cytbi regimen. various modifications of this protocol have been developed in recent years (see table ). in the current study, we compared the outcomes of patients suffering from r/raml that had received their first allogeneic stem cell transplantation (allosct) after conditioning with a sr. methods: adult patients with r/raml who had received their first allosct following sr conditioning between and and were reported to the ebmt registry were analyzed. patients were grouped according to the type of sr used as shown in table . the flamsa-cytbi group served as comparator for all others. the primary endpoint was leukemia-free survival (lfs). secondary endpoints were overall survival (os), relapse incidence (ri), nonrelapse mortality (nrm), refined graft-versus-host-diseasefree, relapse-free survival (grfs), acute (a)gvhd and chronic (c)gvhd. multivariate analysis was done using cox regression. results: patients' characteristics are detailed in table . there were more patients with transformed nhl in the beam ( %vs %). the median time to neutrophil count > . g/l and platelet counts > g/l were ( - ) and ( - ) days for beeam and ( - ) and ( - ) days for beam, respectively. twenty-nine ( %) patients in the beeam and ( %) patients in the beam groups relapsed after a median time to relapse of and months, respectively. after a median follow-up from transplant of months, ( %) patients died, ( %) in the beeam and ( %) in the beam groups, respectively. the main causes of death was lymphoma in patients, (beeam: , beam: ), infections in , (beeam: , beam: ), secondary cancers in patients (beeam). there were no significant differences between beeam and beam regimens for os: hr= . [( . - . conclusions: in this matched pair analysis, beeam and beam resulted in equivalent nrm, lfs and os suggesting that both conditioning regimens may reasonably be employed in patients with dlbcl. the higher relapse rate following beeam requires further evaluation. a prospective randomized study will be required to confirm the equivalence of the two regimens. [[o table] . background: inflammatory bowel diseases (ibd) are thought to increase the risk and severity of graft-versus-host disease (gvhd) and non-relapse mortality (nrm) after allogeneic hematopoietic stem cell transplantation (allo-hsct). thus, ibd have been included in pre-transplant comorbidity risk scores, although formal analysis of allo-hsct outcomes in patients with ibd are lacking. methods: with this background, we designed an ebmt registry retrospective case-controlled analysis to assess outcomes of allo-hsct in patients with ibd. the aim was to compare the incidence of gvhd, nrm and overall survival (os) after allo-hsct in the groups of patients. each patient with ibd was matched with controls according to following factors: patient sex, patient age, disease, intensity of conditioning, donor type and hla disparity, cell source and year of transplant. group comparisons were done using logrank test or gray test for competing risks outcomes. results: between and , patients with ibd who underwent allo-hsct for a hematologic malignancy were reported to ebmt. the cohort comprised males and females, with a median age of years (range, to years) at the time of transplantation. the most frequent malignancies in the ibd group were acute leukemia ( patients; %) and myelodysplastic/myeloproliferative neoplasm ( patients; %). the donor was an identical sibling for patients ( %), and a matched unrelated donor for patients ( %). patients ( %) received a myeloablative conditioning regimen while patients ( %) received a reduced-intensity conditioning regimen. with a median follow-up of months (range, - ) for the patients with ibd and months (range, - ) for controls, the cumulative incidence of grade ii-iv acute gvhd was % for the patients with ibd and % for controls (hazard ratio (hr) for ibd versus controls, . ; % ci, . to . ; p= . ). the cumulative incidence of extensive chronic gvhd at months was % in patients with ibd and % in controls (hr, . ; % ci, . to . ; p= . ). nrm at months was % for the patients with ibd and % for controls (hr, . ; % ci, . - . ; p= . ). the relapse incidence at months was % in patients with ibd and % in patients without ibd (hr, . ; % ci, . - . ; p= . ). os at months was % for the patients with ibd and % for matched controls (hr, . ; % ci, . - . ; p= . ). finally, gvhd-free relapse-free survival (grfs) at months was % for the patients with ibd and % for matched controls (hr, . ; % ci, . - . ; p= . ). conclusions: we report the largest matched-controlled study of allo-hsct in patients with ibd conducted so far. contrary to our expectations, we found no significant differences in acute gvhd, nrm or os between the ibd group and controls. however, ibd patients had significantly more extensive chronic gvhd than the control group. our results suggest that allo-hsct should not be contraindicated if ibd alone is considered a comorbidity. however, ibd patients have a higher risk for developing severe forms of chronic gvhd, which could considerably impair their long-term quality of life. clinical background: the easix (endothelial activation and stress index) score is associated with non-relapse mortality (nrm) and overall survival (os) after reduced intensity (ric) allohct (luft et al, lancet haematol ). we aimed to validate the prognostic ability of easix in a cohort of both unmodified and cd -selected allohct. methods: between april and december , adult patients (pts) underwent unmodified ric or non-myeloablative (nma) allohct (n= ) with uniform gvhd prophylaxis of sirolimus/tacrolimus and low/ dose mtx or myeloablative conditioning (mac) allohct using ex vivo cd -selection (n= ) with the clinimacs cd reagent system (miltenyi biotech) as gvhd prophylaxis. the easix formula (ldh*creatinine/thrombocytes) was calculated at multiple timepoints (pre-allohct, day , day and onset of acute gvhd). a log transformation using base (log ) was applied to all easix variables to reduce skew. a one unit increase in log easix is associated with a doubling (one-fold increase) of easix on the original scale. relapse and death or relapse, were considered competing risks for nrm and acute gvhd, respectively. results: median age was . years ( . - . ) and most pts were male ( %). most pts had myeloid malignancies ( %) and received mac ( %). with a median follow up of . years ( . - ) among survivors, and -year os were . % ( % ci, . - . ) and . % ( % ci, . - . ), respectively. the and -year nrm were . % ( % ci, . - . ) and . % ( % ci, . - . ), respectively. the -year cumulative incidence of grade - , - and - acute gvhd was . % ( % ci, . - . ), . % ( % ci, . - . ) and . % ( % ci, . - . ), respectively. causes of death in pts at last follow up included relapse ( %), gvhd ( %) and infection ( % higher easix score at d , d and at onset of acute gvhd was significantly associated with increased risk of death and nrm (table and figure ). conclusions: higher easix scores at day , day , and at onset of acute gvhd are associated with higher nrm and inferior os with a more prominent association in calcineurin inhibitor-based unmodified allohct compared to cd -selected allohct. endothelial damage is an important contributor to poorer outcomes after allohct and easix formula provides an easy complimentary tool to predict outcomes in these patients. background: subsequent malignant neoplasms (smns) are one of the most important complications of hematopoietic stem cell transplantation (hsct) and result in considerable morbidity and mortality. the reported rate of smns after hsct in adults ranges between - % at -years. there is limited data on smns after pediatric, adolescent and young adult (aya) hsct, where the potential years of life gained is greater than among older adults. the objective of this study was to assess the incidence and types of smns in a cohort of survivors of childhood and aya hscts that were performed for malignant indications. methods: all hsct patients (age - years at time of transplant) who survived at least -years after hsct in the province of ontario, canada between and were identified from transplant centers. clinical data were linked to provincial administrative databases and the ontario cancer registry that identifies cancer cases based on pathology reports and electronic patient records. results: four-hundred and forty-six -year allogeneic hcst survivors were included in this study. of them, ( %) developed smns at a median follow up of . years (range: . - . years). the -year cumulative incidence of smn was % ( . - . %) and the -year cumulative incidence of smn was % ( . - . %). several patients developed more than smn. the most common smns were: papillary carcinoma of the thyroid (n= ); secondary leukemia/lymphoma (n= ); squamous cell carcinoma of the skin/oral mucosa (n= ); and adenocarcinoma of colon/lung (n= ). ten other types of smns were found including sarcoma, melanoma, nerve sheath tumor and breast cancer. nine survivors died at a median of . months after smn diagnosis. the -year cumulative incidence of smn for acute lymphoblastic leukemia survivors who received total-body irradiation was % ( . %- . %). conclusions: our findings corroborate the observation that children and aya who undergo allogeneic hsct are at a significant risk for developing smn. careful observation in the survivorship period is required for potential early detection. clinical and and who survived at least years post-hct while continuing follow-up at our centre. we documented performance status, comorbidities, number of medications and occurrence of secondary malignancies at years, as well as survival following the -year time-point. results: the median age of the cohort at years post-hct was years (range - ), ( %) of patients underwent transplant using a related donor. eighty patients ( %) underwent hct for cml, ( %) for aml, ( %) for all, ( %) for aplastic anemia, ( %) for other indications. bone marrow grafts were used in ( %) patients. myeloablative conditioning was used in ( %) patients. individual comorbidities were categorized into five major groups: endocrine ( %), cardiac ( %), secondary cancer ( %), psychosocial ( %), and other organ dysfunctions ( %). at the year mark, median karnofsky performance status was (range - ). no comorbidities were seen in ( %) patients. the most frequent individual comorbidities were dyslipidemia (n= , %), hypertension (n= , %), osteoporosis (n= , %) and hypothyroidism (n= , %). at the time of the -year post-hct follow-up, the median number of medications patients were taking was (range - ). follow-up data after the -year mark was available for ( %) patients. median follow-up of survivors after the -year mark was months (range - months). fiveyear overall survival of the patients was % ( %ci - %) and at years was % ( % ci - %). when grouped by age at the -year mark, there was no significant difference in -year os survival between ages - (n= , -year os %), - (n= , -year os %) and - (n= , -year os %) (p= . ). when grouped by the number of concurrent comorbidities, there was a significant difference in os between the groups with - (n= ), - (n= ) and ≥ comorbidities (n= ) ( -year os %, % and % respectively, p= . , figure ). when grouped by the number of medications patients were on at the -year mark, there was a borderline significant difference between the groups on - (n= ), - (n= ) and ≥ (n= ) different medications ( -year os %, % and % respectively, p= . ). conclusions: long-term allogeneic hct recipients may develop a number of long-term comorbidities that negatively influence survival even past the -year mark. these findings warrant the continuous long-term medical followup of allogeneic transplant patients, regardless of age or time that has lapsed post-hct. background: no standard procedure is in use to predict sinusoidal obstruction syndrome/venoocclusive disease (sos/ vod). recently the sos/vod cibmtr clinical risk score (age, karnofsky, sirolimus, hepatitis b/c, conditioning regimen, disease type) has been established using the cibmtr database (biol blood marrow transplant. ; : ) . the endothelial activation and stress index (easix), based on the simple formula 'ldh(u/l) x creatinine(mg/dl) / thrombocytes ( /l)', has been proven to predict mortality after gvhd (lancet haematol ; :e ) . the aim of the current study was to test prediction of sos/vod by easix compared with the cibmtr score in two independent european cohorts. methods: sos/vod was defined according to the ebmt criteria. the capacity of easix and of the cibmtr score for predicting sos/vod was tested retrospectively in consecutive adult patients undergoing allosct at a single institution between and (training cohort). the primary endpoint was prediction of sos/vod by the cibmtr score or by easix-d (easix measured at the day of allosct). results were validated in an independent cohort of adult allosct recipients from another single institution transplanted between and . incidence of sos/vod was assessed by uni-and multivariable cox regression analyses using age, easix and the cibmtr score as confounders. results: sos/vod was diagnosed in patients ( . %, median onset day + ) in the training cohort and in patients ( . %, median onset d + ) in the validation cohort, respectively. in the training cohort, increasing easix-d was significantly associated with sos/vod incidence on multivariate analysis (hr per log increase . , %ci . - . , p= . ) . similarly, easix-d predicted the incidence of sos/vod in the validation cohort (hr per log increase . , % ci . - . , p< . ). also, the cibmtr score showed an association with sos/vod incidence which was however significant only in the training cohort (hr per log increase . , % ci . - . , p= . ) but not in the validation cohort, (hr per log increase . , % ci . - . , p= . ). these results are visualized by comparing easix-d and cibmtr-vod scores in patients grouped according to later vod development within the observation period ( figure , kruskal-wallis tests; a-b easix-d and c-d cibmtr risk score). the association of easixd with vod incidence was independent from cibmtr score. conclusions: easix-d is very easy to test and predicted sos/vod in two separate cohorts of allosct recipients independent of the cibmtr-vod score. patients with high easix-d scores might be candidates for clinical evaluation of intensified strategies to prevent sos/vod after allosct. [[o image] . figure ] disclosure: the authors thank jazz pharmaceuticals for providing financial support that was used for data collection. the company had no active part in this project, did not have access to data and was not involved in analyses or writing/editing of this abstract. early hyperbilirubinaemia without sos/vod -a link between endothelial dysfunction and early mortality after allogeneic transplantation thomas luft , david schult , joshua majer-lauterbach , sihe jiang , aleksandar radujkovic , peter dreger , olaf penack university hospital heidelberg, heidelberg, germany, charité universitätsmedizin berlin, berlin, germany background: endothelial dysfunction is a risk factor for early mortality after allogeneic stem cell transplantation (allosct) and is linked to transplant-associated thrombotic microangiopathy (tam). similar to tam, diagnosis of sinusoidal obstruction syndrome / venoocclusive disease (sos/vod) is based on consensus criteria, and several scores have been proposed that include earlier or later stages of the diseases. early hyperbilirubinaemia occurs frequently after allosct. the pathophysiology is often elusive, and only in a subset of allosct recipients with hyperbilirubinaemia sos/vod is identified as the underlying mechanism. the aim of the current study was to explore clinical impact and pathophysiological context of early hyperbilirubinaemia without sos/vod. methods: in two independent cohorts of and patients, serum bilirubin levels before allosct and on days , , , , , and were retrospectively retrieved. sos/vod was defined according to the ebmt criteria. patients with at least one bilirubin value of > mg/dl between days - were grouped as "early hyperbilirubinaemia". we assessed overall survival (os), non-relapse mortality (nrm) and time to relapse (ttr) with and without early hyperbilirubinaemia depending on coincident sos/vod, tam, and refractory acute gvhd, and we investigated the impact of statin-based endothelial cell prophylaxis (pravastatin plus ursodeoxycholic acid). serum bilirubin levels were correlated with the endothelial activation and stress index (easix, 'ldh(u/l) x creatinine(mg/dl) / thrombocytes( /l)'), measured before allosct (easix-pre) or on the day of transplantation (easix-d ). similarly, bilirubin levels were correlated with the sos/vod cibmtr clinical risk score, and with serum markers of liver damage (alanine transaminase, alt, gamma-glutamyltransferase, ggt) and endothelial cell distress markers (angiopoietin- ). results: early hyperbilirubinaemia was diagnosed in ( %) and ( %) patients of the training and validation cohort, and vod diagnostic criteria were met by % and % of patients with early hyperbilirubinaemia, respectively. in all patients, early hyperbilirubinaemia was associated with increased nrm and os (nrm, training: hr . , %ci . - . , p< . ; nrm validation: hr . , %ci . - . , p< . ). increased nrm in recipients with hyperbilirubinaemia without sos/vod was independent from tma or refractory acute gvhd ( figure ). easix-pre and easix-d correlated with day - bilirubin in both cohorts. easix-pre, easix-d and the cibmtr-vod score predicted risk of early hyperbilirubinaemia. however, only easix predicted risk of nrm in patients without sos/vod. endothelial protection with statins and ursodeoxycholic acid was associated with reduced incidence of and reduced nrm after early hyperbilirubinaemia. furthermore, pre-transplant angiopoietin- correlated with early hyperbilirubinaemia, whereas alt and ggt did not. conclusions: early hyperbilirubinaemia represents a novel risk factor for nrm independent of tma and refractory acute gvhd, even in patients not meeting the diagnostic criteria for vod. the endothelial relationship of this condition is underlined by the observation that angiopoietin- , easix-pre and easix-d but not markers of liver damage associate with the incidence of early hyperbilirubinaemia. therapeutic strategies aiming at normalization of endothelial dysfunction after allosct are attractive. as a first example, our data demonstrate reduced incidence of early hyperbilirubinaemia and reduced nrm thereafter in allosct recipients prophylactically treated with statins and ursodeoxycholic acid in the peri-transplant period. [[o image] . figure background: accumulating evidence has suggested complement activation in transplant-associated thrombotic microangiopathy (ta-tma), mainly in the pediatric setting. to further understand its pathogenesis in adults, we hypothesized that both complement and neutrophils are activated in ta-tma as previously observed in distinct thrombotic disorders. methods: we enrolled adult ta-tma (international working group/iwg criteria), acute and/or chronic graftversus-host-disease (gvhd) and control hematopoietic cell transplantation (hct) recipients in a : : ratio (january -june ). complement activation was detected in patient sera and plasma with the modified ham test (mham), soluble c b- and activated c fragments; while neutrophil activation with neutrophil extracellular traps (nets) using extracellular dna and myeloperoxidase/ mpo-dna. results: we studied ta-tma, gvhd and control patients. ta-tma patients suffered from severe acute and/or extensive chronic gvhd. ta-tma presented at median + ( - ) day post-transplant. full donor chimerism was evident in all patients. no significant difference in transplant characteristics was observed among groups, except for the significantly lower gvhd rate in the control group. c b- and mham levels were significantly increased in ta-tma compared to gvhd (p= . and p= . , figure a ) and control patients (p= . and p= . , figure b ), while no significant difference was observed in activated plasma c levels in the plasma. in the multivariate analysis, c b- levels were an independent predictor of ta-tma diagnosis (p= . ). we next sought to determine the cutoff value of c b- that distinguishes ta-tma from other hct recipients. in the receiver-operating characteristic curve, we found a significantly high area under the curve ( . , p= ). values higher than . ng/ml conferred a specificity of % with a very high sensitivity of . % for ta-tma diagnosis. regarding nets, both extracellular and mpo-dna were significantly increased in patients with ta-tma compared to gvhd (p< . and p= . , figure c ) and control patients (p< . and p= . , figure d) . interestingly, increased complement activation markers (c b- and mham) were strongly associated with mpo-dna (r = . , p= . and r = . , p= . , respectively) and extracellular dna (r = . , p= . and r = . , p= . , respectively). lastly, we studied changes of complement and neutrophil activation in patients that received complement inhibition by eculizumab. despite delayed initiation in the first two patients ( and days post ta-tma diagnosis respectively), we observed laboratory response including evidence of reduced hemolysis, schistocytosis and transfusion needs. both extracellular dna and c b- levels were significantly reduced post doses of eculizumab (p< . ). however, all patients succumbed to complications of endstage renal disease and infections after a median of ( ) ( ) ( ) ( ) ( ) ( ) ( ) doses of eculizumab. conclusions: our findings demonstrate for the first time a crosstalk between complement and neutrophils in adult ta-tma. in addition, we were able to set a cut-off c b- value for distinguishing complement activation in unselected patients diagnosed with the iwg criteria. although complement inhibition by eculizumab seems to hinder this pathophysiological process, further studies are needed to clarify changes and identify optimal therapeutic targets in this complex setting. [[o image] . background: hepatic vod/sos with multi-organ dysfunction (mod; typically, renal or pulmonary) may be associated with > % mortality. defibrotide is approved for treating severe hepatic vod/sos post-hematopoietic stem cell transplantation (hsct) in patients aged > month in the european union, and for hepatic vod/sos with renal or pulmonary dysfunction post-hsct in the united states and canada. per prescribing guidelines, defibrotide mg/kg/day ( divided doses) is recommended for ≥ days. this pooled analysis examined time to complete response (cr) of vod/ sos and mod symptoms relative to day of defibrotide initiation in patients who received defibrotide mg/kg/day. methods: time to cr and safety data were pooled from studies that included patients with vod/sos and mod post-hsct who were treated with defibrotide: a phase trial (n= ) and a phase , randomized dose-finding trial (n= receiving mg/kg/day). duration of therapy in patients who discontinued due to cr in an expanded-access program (t-ind) in vod/sos patients with and without mod post-hsct (n= ) was analyzed separately due to differences in the patient population and data monitoring protocol (eg, cr data by day were not collected). vod/sos diagnosis was defined by baltimore criteria/biopsy for the phase and studies; in the t-ind, modified seattle criteria also was permitted. minimum recommended treatment duration was ≥ days (phase ) or ≥ days (phase , t-ind). results: the pooled phase and trials had patients with cr (n= and n= , respectively) and patients without cr (n= and n= , respectively). of patients with cr ( figure) in the phase and trials, the median time to cr was . days (range: - ); of these, . % achieved cr after or more weeks of treatment. in the t-ind, patients discontinued treatment due to cr (median time to discontinuation, days after initiation of defibrotide; range, - ), with patients ( . %) discontinuing on or after week of treatment. in the phase and studies (n= ), patients ( %) had treatment-related adverse events (traes); most common were gastrointestinal (gi) hemorrhage ( . %), epistaxis ( . %), hypotension ( . %), and pulmonary alveolar hemorrhage ( . %). in the t-ind (n= ), patients ( . %) had ≥ trae; most common were pulmonary hemorrhage ( . %), gi hemorrhage ( . %), epistaxis ( . %), and hypotension ( . %). conclusions: among patients with cr in the phase and studies, a significant number of patients achieve cr ( %) after weeks of persistent treatment, highlighting the importance of continued therapy as per label indication. support: jazz pharmaceuticals figure. time to cr among patients achieving cr in defibrotide phase and studies] trial registry: clinicaltrials.gov: nct , nct , and nct disclosure: paul g. richardson has served on advisory committees and as a consultant, and has received research funding from jazz pharmaceuticals. angela r. smith and leslie lehmann have nothing to disclose. nancy a. kernan received grants from gentium during the conduct of the study, and her research was supported by the national cancer institute of the national institutes of health under award number p ca ; the content is solely the responsibility of the author and does not necessarily represent the official views of the national institutes of health. she has a research grant from jazz pharmaceuticals. robert ryan and william tappe are employees of jazz pharmaceuticals and hold stock and/or stock options in jazz pharmaceuticals plc. stephan a. grupp has served on a steering committee and as a consultant to jazz pharmaceuticals. sperm counts and prevalence of testosterone substitution at long-term follow-up after myeloablative allogeneic hsct in childhood background: little is known about the long-term effects of pediatric hsct on the male reproductive axis. we investigated sperm counts and prevalence of testosterone substitution twenty years after pediatric hsct and aimed at identifying risk factors for azoospermia and testosterone substitution. methods: this cross-sectional follow-up study included two national cohorts of adult males (≥ years) treated with myeloablative allogeneic pediatric hsct before age between - in denmark and finland. the study included medical history; physical examination including testicular volume and screening for chronic graft-versushost-disease (cgvhd); sex hormones and a semen sample. cumulative (pre-hsct and hsct) gonadal irradiation including tbi and cumulative cyclophosphamide equivalent doses (ced) were estimated from patient files. results: / ( %) of eligible patients (age - years) participated with a median (range) follow up time of years. % had malignant diagnoses and % were treated with tbi-conditioning. delivered a semen sample. results of semen and sex hormone analyses are listed in table . / ( %) had detectable sperm counts ( . - . million), of these were treated with chemotherapy only and with tbi ( with gy tbi and with - gy tbi including with gonadal shielding). in patients with detectable sperm in the ejaculate, increase in sperm counts was associated with time from hsct (β= . million per year %ci ((- . )- . ), p= . , time range - years) indicating late spermatogenic recovery. testicular irradiation was a strong risk factor for azoospermia (or= . %ci ( . - . ), p< . ) and testosterone substitution (or= . %ci ( . - . ), p= . ) and no patients with cumulative testicular irradiation doses > gy had detectable sperms, figure . pre-pubertal stage at hsct was a risk factor for later testosterone substitution (or= . %ci ( . - ), p= . ). risk of testosterone substitution was associated with time from transplantation (or for + year . %ci ( . - . ), p= . ). cumulative ced adjusted for testicular irradiation was not a risk factor for azoospermia or testosterone substitution, nor was cgvhd. [[o image] . figure ] conclusions: late spermatogenic recovery is possible - years following myeloablative hsct but depends on cumulative testicular irradiation dose; azoospermia was present in all patients treated with > gy. pre-pubertal stage at hsct increases the risk for later testosterone substitution supporting the hypothesis that pre-pubertal leydig cells are more sensitive to irradiation than more mature ones. additionally, the risk of testosterone substitution increased with time from transplantation indicating a potential early androgen insufficiency in male hsct recipients. thus, close follow-up and focus on cumulative irradiation doses are needed. disclosure background: hematopoietic stem cell transplantation (hsct) has become a standard component of therapy for several malignant indications. as hsct may improve survival for some cancers, the risk for late complications is of increasing concern. the frequency of hospitalizations can serve as a proxy measure of severe morbidity. however, knowledge regarding late hospitalizations is limited. the objectives of the study were to describe health care utilization as measured by hospitalizations beyond years following transplant in survivors of pediatric/adolescent and young adult (aya) hsct for a malignant indication. methods: we linked data from ontario hsct centers and provincial health care utilization data housed at institute for clinical evaluative sciences (ices) to describe all hospitalizations and their indications. we also described intensive-care unit admissions. the study population included ontario residents with cancer age - years who underwent hsct between and , who survived more than years from transplant (index date). hospitalizations were described from the index date until dec or death. results: the cohort consisted of survivors who were followed for a median of . years from index date (interquartile range (iqr) . - . ). indications for transplant included: acute lymphoblastic leukemia (n= , . %); myeloid malignancy (n= , . %); and lymphoma (n= , . %). of these, ( . %) received a related-donor bone-marrow hsct. at the time of hsct, ages were: - (n= , . %); - (n= , . %); and - (n= , . %) years. there were patients ( %) with at least hospitalization beyond years from hsct. there were a total of hospitalizations, resulting in a hospitalization rate of . per follow-up year. average length of hospital stay was . days. a total of intensivecare unit admissions were documented among ( . %) patients. the most common indications for hospitalization were: graft-versus-host disease (gvhd) (n= , . %), relapse (n= , . %), infection (n= , . %), orthopedic procedures/fractures (n= , . %), benign neoplasm (n= , . %) and subsequent malignant neoplasm (n= , . %). among those who did not relapse, / ( %) were hospitalized. at years following hsct, the proportion of patients hospitalized was . %. an underlying diagnosis of acute myeloid leukemia (aml) (p= . ) and chronic gvhd (p= . ) were associated with increased rate of hospitalization. in the follow-up period, ( . %) patients died. conclusions: we identified a high rate of late hospitalization in pediatric/aya survivors who underwent hsct for a malignant indication, even among those without relapse. a diagnosis of aml and chronic gvhd were associated with increased risk for hospitalization. careful observation in the survivorship period is required for potential prevention of hospitalization. clinical background: engraftment syndrome (es) is a clinical complication characterized by inflammatory signs and symptoms occurring during neutrophil recovery after stem cell transplantation (sct). its incidence varies depending on the clinical criteria used. the objective of this study was to analyze the incidence, clinical characteristics, risk factors and clinical outcomes of es after haploidentical-sct with post-transplant cyclophosphamide (haplosct) in a single center. methods: consecutive haplo-sct performed between - in our center were retrospectively reviewed. gvhd prophylaxis was performed with cyclophosphamide mg/kg/day days + and + , mmf and csa from day + . g-csf was started in all cases from day + until engraftment. cases were excluded from the analysis ( due to death before engraftment and due to primary graft failure). maiolino and spitzer´s diagnostic criteria were used to define es. results: characteristics of the transplant included are shown in table . the es incidence was . %, with median time to diagnosis of days (iqr, ( ) ( ) ( ) ( ) ( ) . median time to neutrophil engraftment in the es cohort was days (iqr, ( ) ( ) ( ) ( ) ( ) ( ) ( ) . fever ( %) and skin rash ( %) were the most frequent clinical findings. there were other cases of fever and skin rash during the peri-engraftment period with a final diagnosis of gvhd considering clinical outcome. patients ( %) received high doses of corticosteroids, with favorable response in % of cases. of note, % cases also needed intensive supportive care. there were no deaths secondary to es. univariable analysis showed a higher risk of es with use of brother/sister as cell donor ( % cases of es; p= , ). no other risk factors were identified. no association was noted with acute or chronic gvhd. there was no significant difference in nrm, overall survival and progression-free survival between es and non-es patients. conclusions: in our experience, es is a frequent complication of haplosct with post-transplant cyclophosphamide. to our knowledge, this is the largest study including only haploidentical sct. most cases of es had a self-limited course or good response to corticosteroids, and there were no associated mortality. however es can progress to multi-organ dysfunction with need of intensive supportive care. in our analysis, incidence of es was higher with the use of sibling haplo-donors, and further studies are needed to confirm these results. we have not found relationship between es and gvhd. specific biomarkers may contribute to an early identification of this entity in order to install therapeutic measures. disclosure: nothing to declare features and outcome of early cardiac toxicity associated with post-transplant cyclophosphamide in allogeneic stem cell transplantation background: data on risk factors and incidence of early cardiac events (ece) after post-transplant cyclophosphamide (pt-cy) are scarce. thus, we compared clinical outcomes between patients who received pt-cy and patients who did not in a cohort study including all consecutive patients allografted in our center. methods: we analyzed all ece occurring within months after hsct in patients. transthoracic echocardiography and ekg were performed before hsct, at day + and in case of ece. prior to transplant, patients ( . %) had at least one cardiovascular risk factor, ( . %) cardiovascular disease history and ( . %) left ventricular systolic dysfunction (lvsd) (defined by left ventricular ejection fraction < %). median age was years (range, - ) and . % of patients were males. patients were transplanted for aml (n= , %), all (n= , %), lymphoma (n= , %), multiple myeloma (n= , . %), mds (n= , %) and mpn (n= , %). disease risk index was high or very-high in patients ( %). conditioning regimen were mac (n= , %), ric (n= , %) or sequential (flamsa-like) (n= , %). donors were matched related (n= , %), unrelated (n= , %) or haploidentical (n= , %). stem cell source was peripheral blood (n= , %) or bone marrow (n= , %). gvhd prophylaxis included cyclosporine in all patients associated with mycophenolate mofetil (n= , %), short courses of methotrexate (n= , %) and/or antithymocyte globulin (n= , %). in the pt-cy group, patients received pt-cy at mg/kg/day for at least day and patients for days, including patients with unrelated donor and patients with matched related donor, either because of hla-mismatch or renal insufficiency, or inclusion in a clinical trial. results: in univariate analysis, cumulative incidence of ece was . % in the pt-cy group and . % in the no pt-cy group (p< . ). the main complication was lvsd ( % of patients in the pt-cy group and % in the no pt-cy group, p< . ). other ece included acute pulmonary edema (n= , %), arrhythmia (n= , %), pericarditis (n= , %) and coronary artery syndrome (n= , %) in the whole patient group. ece resolved in patients ( %). cardiovascular risk factors and the cumulative doses of anthracycline were not significantly associated with the incidence of ece. in multivariate analysis, the main factors associated with ece were the use of pt-cy [hr= . , % ci . after a median follow-up of . months (iqr, - ), the -year cumulative incidences of nrm, relapse, os and dfs were % vs. %; % vs. %; % vs. %; % vs. % in the pt-cy and no pt-cy groups, respectively (p values non significant). at last-follow-up, patients have died. the main causes of death were disease relapse (n= ), gvhd/infection (n= ) and ece (n= ). conclusions: incidence of ece is significantly higher in the pt-cy group. in elderly patients or with a history of pretransplant cardiac event, an alternative to pt-cy should be considered to prevent ece. disclosure: nothing to declare severe iron overload, measured by liver mri at the preallo-hsct, significantly impaired the long-term outcome of the procedure methods: once approved by the clinical trials and ethics committee, a liver mri was systematically offered to patients who were admitted to undergo an allo-hsct in our center. among the pts consecutively transplanted between june and july , pts signed the informed consent and underwent a pre-hsct mri to assess the hepatic iron load. a signal intensity ratio (sir) method was employed for the measurement. results: pts were male, and were female. median age was years (range: - ). the baseline diseases were: aml, nhl, all, mds, cmpd, mm, and bmf. underwent alternative donor transplants ( unrelated, and haplo-identical), and hla-id family donor transplants. stem cell source was pb in , and bm in cases. conditioning regimen was intensive in pts, and ric in ; no non-myeloablative allo-hsct were performed. based on hepatic iron overload at pre-hsct mri, the patients were classified into the following groups: pts ( %) showed severe io (lic > micromol/g or . mg/g), pts ( %) moderate io (lic - micromol/g or . - . mg/g) and pts ( %) no significant io (lic < micromol/g or . mg/g). as shown in table , majority of patients with severe iron overload had been heavily transfused, and had a high pre-hcst ferritin level. surprisingly, pre-hsct chelation had been employed only in pts ( %) of this group. overall mortalities at days + , + and + in the global series, and in severe versus non-severe io group are reflexed in table . conclusions: ) our data shows that pre-allo-hsct iron-overload correlates with previous prbc transfusion load; ) it also makes evident that io is an important risk factors for post-transplant mortality. ) our real-life study reflects that only a minority of heavily transfused pts had received chelation therapy previously to the allo-hsct. ) considering the relevance of pre-allo-hsct iron overload, we strongly suggest to referring physicians to employ chelation therapy for patient candidates to transplant during the treatment of the underline disease. background: neurologic complications (ncs) are associated with relevant morbidity and mortality after allo-hsct. the aim of this study was to analyze the incidence, characteristics, risk factors and outcomes of patients developing ncs after allo-hsct. methods: we evaluated consecutive adult patients (> years) who underwent allo-hsct at our center between january and december . we collected data on neurological symptoms, diagnostic methods, time of onset and cause. nc was defined as any neurological event that occurred after starting the conditioning regimen and before relapse. nc due to central nervous system (cns) infections o neoplastic infiltration were excluded. we considered both cns and peripheral nervous system (pns) complications. results: the current series comprised allo-hsct from matched sibling donor (msd), from umbilical cord blood (ucb), matched unrelated donor (mud) and haploidentical donor (haplo). median age was years and most patients had acute leukemia ( %). median follow-up of surviving patients was months (range, - ). there were differences in median follow-up according to the donor source, being longer in ucb and msd, and months respectively, and shorter in mud and haplo, and months respectively (p< . ). overall, ncs were documented in cases, after msd transplants, after ucb, after mud, and after haplo. cns complications ( %) were more common than pns events. the most frequent nc was encephalopathy ( %) followed by myopathy ( %) -year and -year cumulative incidence of ncs was % and %, respectively. -year cumulative incidence was % after msd, % after ucb, % after mud, and % after haplo transplants (p= . ). conclusions: ncs are common and diverse after allo-hsct. ncs were more frequent in recipients allografted from alternative donors, recipients older than years, and in those developing gvhd. cns complications, but not pns, are associated with poor os. disclosure: nothing to declare. abstract already published. using ciclosporin's area under the curve (auc) to predict risk of acute kidney injury in non-myeloablative haematopoietic stem cell transplantation vaidie julien , jean-baptiste woillard , stéphane girault , pierre marquet , franck saint-marcoux , arnaud jaccard , pascal turlure background: non myeloablative allogeneic stem cell transplantation (hsct) by limiting toxicity, can be proposed to elderly patients. however, acute renal injuries related to anti-calcineurin, which are frequent in this population, can negatively impact the outcome. currently, the exposure indexes to follow and the target to use are not consensually established. however, using the area under the curve (auc) for therapeutic drug monitoring (tdm) is theoretically the best method to describe the patient's exposure. the primary objective of this study was to determine an auc target for ciclosporin associated to the occurrence of acute kidney injury in hsct patients. methods: we retrospectively studied all consecutive patients who received a non-myeloablative allogeneic stem cell transplantation at limoges university hospital from june to december . patients received fludarabine mg/m /day between d- and d- before allograft and busulfan . mg/kg/day at d- and d- . gvh prophylaxis consisted in rabbit anti-lymphocyte serum at the dose of . mg/kg at d- and d- , and ciclosporin at the beginning dose of mg/kg per os twice a day. mycophenolate mofetil was added for patients with hla-matched or mismatched unrelated donors. tdm of ciclosporin was done based on trough concentration (c ) twice a week with concomitant renal evaluation using creatininemia. dose adjustments were done in according to the sfgm-tc recommendations and renal tolerance. ciclosporin's auc was evaluated at day , day and day after allograft using bayesian estimation from a limited sampling strategy and a population pharmacokinetics model previously published. the association between ciclosporin auc and acute kidney injury was investigated using a joint model. a roc curve was then constructed to investigate an auc threshold associated with the best sensibility/specificity ratio for acute kidney injury (aki . interestingly, a very low correlation was observed between ciclosporin c and auc (r² = . for the overall period). additionally, an higher intra-individual variability was observed with c than auc (coefficient of variation= % and % respectively). conclusions: we report in this study that a ciclosporin auc= . mg*h/l could be used as a high threshold for aki. new evaluations of auc in prospective studies are needed to better define the relevance of this marker in clinical practice. disclosure background: human cord blood (cb) provides an attractive source of hematopoietic stem cells for allogeneic transplantation of patients with a variety of diseases. a sufficient hematopoietic stem cell (hsc) dose, currently measured as cd + cells/kg recipient, is essential for successful engraftment. nevertheless, the frequency of true hsc within the cd + population and the dynamics of their clonal offspring remain poorly understood and may differ between donors. methods: here, we use cellular barcoding and multiplexed high-throughput sequencing to determine the frequency of repopulating cells among cd + cells from individual human cb donors, and to quantify their contribution to each of the blood lineages over time in murine nod/scid/il ry -/-(nsg) xenografts. results: in total, we detected a median of . (range - ) clones in blood and . (range . - ) clones in bone marrow, corresponding to hspc frequencies of : to : . the number of retrieved clones correlated to barcoded cd + cell dose (spearman r= . , p= . ), yet could vary up to four-fold between mice transplanted with the same cell dose. clonal patterns in blood early after transplant differed markedly from those at later time points, and became increasingly deterministic over time. the majority of clones displayed multilineage output, yet clones with bias towards lymphoid or myeloid lineages were also present. similar to recent data on murine hsc clones, human cb clones were distributed asymmetrically across different bone marrow sites. conclusions: in conclusion, the frequency of nsgrepopulating cells among cord blood cd + cells is low, and highly variable between individual cb donors. heterogeneity in hsc frequency, proliferation and lineage fate decisions may contribute to (non-)engraftment upon hsct. future research will be aimed at identifying the underlying mechanisms guiding hsc behavior upon transplantation and expanding our findings to human hsct recipients. [[o image] . background: chronic granulomatous disease (cgd) is a rare genetic immune disorder that leaves patients susceptible to life-threatening infections, chronic inflammation and often long hospital stays. x-linked cgd (x-cgd) is caused by mutations in cybb encoding the gp phox subunit of the phagocyte nadph-oxidase, which regulates cell ph and ionic content for efficient microbial killing. allogeneic hematopoietic stem cell transplant (hsct) has been a potentially curative approach for x-cgd patients, but is often complicated by lack of hla-matched donors and risks of graft versus host disease, graft rejection, and procedure-related fatality. previous attempts at autologous ex vivo gene therapy for x-cgd using gammaretroviral vectors have met with limited efficacy due to transient engraftment of gene corrected cells, gene silencing, and mutagenic activation leading to myelodysplasia. here we report on patients with a history of severe x-cgd-related complications, who received autologous ex vivo gene therapy (gt) using a novel self-inactivating lentiviral vector (g xcgd lv) designed to limit the risk of mutagenesis through preferential expression of the missing g phox subunit in mature myeloid cells. methods: similar trials of gt with g xcgd lv were initiated in the uk (n= , plus compassionate use patient) and in the usa (n= , sites). all patients had histories of severe, persistent infections, and inflammatory disease. g-csf and plerixafor-mobilized cd + selected hematopoietic stem and progenitor cells were transduced ex vivo with g xcgd lv. subjects received near-myeloablative conditioning with single agent busulfan, targeted to net area-under-the-curve of , ng/ml*hr. freshly prepared or cryopreserved quality-tested gene-modified cells, manufactured on-site, were administered intravenously. primary endpoints were efficacy, as determined by percent of oxidase positive granulocytes by dihydrorhodamine [dhr] flow cytometry, and safety at months. results: we report results for patients ( - years) with - . years of follow-up; additional patients were treated but died within months of gt from complications deemed related to pre-existing disease-related co-morbidities (severe pulmonary disease, anti-platelet antibodies). gt was welltolerated, only one serious adverse reaction (a systemic inflammatory process at engraftment of functional neutrophils) was reported as possibly related to gt. patients experienced typical conditioning-related transient neutropenia, thrombocytopenia and mucositis. there has been no molecular evidence for clonal dysregulation or gene silencing through cpg dinucleotide methylation. followup demonstrated sustained stable persistence of - % oxidase (+) neutrophils for > months in / surviving patients (one, who remains clinically well, had a decline to < % after months) [ figure] . these patients have maintained restoration of biochemical function and immunity (defined as ≥ % of oxidase (+) by dhr) as of december . patients have been well, without new x-cgd-related infections, and are successfully weaned off prophylactic antibiotics. conclusions: these results are the first demonstration of effective autologous lentiviral gt at months in severely affected x-cgd patients without evidence of genotoxicity. corrected neutrophil function has been observed in patients for > months and has been associated with significant clinical improvement, freedom from infections, and resolution of chronic inflammation. results are supportive of extended clinical trials evaluating the safety and efficacy of g xcgd lv-based gene therapy. updated results from the ongoing northstar- (hgb- ) trial of lentiglobin gene therapy in patients with transfusion-dependent β-thalassemia and non-β /β genotypes franco locatelli , alexis thompson , , janet kwiatkowski , , suradej hongeng , john porter , martin sauer , adrian thrasher , isabelle thuret , heidi elliot , ge tao background: allogeneic hematopoietic stem cell (hsc) transplantation is potentially curative for patients with transfusion dependent β-thalassemia (tdt); however, it is associated with risks of morbidity and mortality and is limited by donor availability. gene therapy has the potential to be an effective treatment option for patients with tdt, but without some of these limitations. lentiglobin gene therapy contains autologous cd + hscs transduced ex vivo with the bb lentiviral vector encoding β-globin with the t q amino-acid substitution. lentiglobin is being studied in patients with tdt and non-β /β genotypes in the ongoing, phase northstar- study (hgb- ; nct ). methods: northstar- is enrolling patients with tdt who had a history of ≥ ml/kg/year of red blood cells (rbcs) or ≥ rbc transfusions/year. to generate drug product (dp), autologous cd + cells were collected by apheresis after g-csf and plerixafor mobilization and transduced with the bb lentiviral vector. patients received myeloablative conditioning with single-agent busulfan before dp infusion. primary endpoint was the proportion of patients achieving transfusion independence (ti, weighted average hemoglobin [hb] ≥ g/dl without rbc transfusions for ≥ months). patients are followed for years and offered participation in a long-term followup study. statistics are presented as median (min-max). results: as of september , patients (age years; patients ≥ years) have been treated (follow-up . [ . - . ] months). patients received a cell dose of . x ( . - . ) cd + cells/kg with a dp vector copy number of . ( . - . ) vector copies/diploid genome and % ( - %) of cells were transduced. baseline liver iron content (lic) was . ( . - . ) mg fe/g dw. outcomes by age and baseline lic will be reported. times to neutrophil and platelet engraftment were ( - ) and . ( - ) days, respectively; patient ( . month follow-up) and patients ( . - . months follow-up) had not achieved neutrophil and platelet engraftment, respectively, at time of analysis. of patients with ≥ months of follow-up, ( ≥ years old) stopped chronic rbc transfusions and two have achieved ti. at last study visit, total hb in these patients was . - . g/dl consisting of . - . g/dl gene therapy-derived hb, hba t q . one treated patient had no rbc transfusions for months, then re-initiated transfusions due to low hb. non-hematologic grade ≥ adverse events post-infusion in ≥ patients included stomatitis, febrile neutropenia, epistaxis, pyrexia, and veno-occlusive liver disease (vod). one grade event of serious prolonged thrombocytopenia after platelet engraftment was considered possibly related to lentiglobin. the three grade serious vod events were attributed to myeloablative conditioning ( table ). the events resulted in extended hospitalization and resolved following defibrotide treatment. there were no deaths or graft failure and no evidence of vector-mediated replication of competent lentivirus or clonal dominance. conclusions: in northstar- , / patients with tdt and non-β /β genotypes treated to date produced sufficient gene therapy-derived hb, hba t q , to stop chronic transfusions following lentiglobin gene therapy. total hb in patients off rbc transfusions remains stable at > g/dl. the safety profile of lentiglobin remains generally consistent with myeloablative busulfan conditioning. background: metachromatic leukodystrophy (mld) is an ultra-rare and devastating demyelinating lysosomal storage disease caused by mutations in the arylsulfatase a (arsa) gene, currently with no approved treatment. we report an interim analysis of the safety and efficacy results of early-onset mld subjects treated with experimental autologous, ex-vivo, lentiviral-mediated hematopoietic stem cell gene therapy (hsc-gt) followed for up to years posttreatment, as part of an ongoing, open-label, study. the study has completed the enrollment period; follow up visits are currently on-going. methods: gt consists of a formulation of autologous cd + cells transduced ex vivo with a self-inactivating lentiviral vector encoding for the human arsa gene and administered intravenously after busulfan conditioning. twenty early-onset mld subjects (pre-symptomatic or early symptomatic) were enrolled and treated ( late infantile [li] and early juvenile [ej]). co-primary efficacy endpoints were improvement in gross motor function measure (gmfm) score ( %) and a significant increase in arsa activity in peripheral blood mononuclear cells (pbmcs), evaluated months after treatment. safety endpoints include engraftment failure and long-term safety and tolerability of lentiviral-transduction. results: / subjects are alive after a clinical follow-up of - years. two ej subjects, treated after onset of symptoms, died due to rapid disease progression -and months post-treatment. there was no treatment-related mortality, no evidence of abnormal clonal expansion, and no adverse events related to the medicinal product. durable and stable engraftment of gene-corrected cells were observed beginning -month post-treatment, with persistent vector copy number in cd + bone marrow cells and pbmcs throughout the follow-up for all subjects. reconstitution of arsa activity in the hematopoietic system was observed in both populations (li and ej), stabilizing at normal to supranormal levels within three months. arsa activity in csf showed a similar pattern; normal levels were observed - months post-treatment, demonstrating effective enzymatic production in the central nervous system (cns). the majority of li and ej subjects treated before the onset of overt symptoms showed normal motor development, stabilization of motor dysfunction or a significant delay in disease progression, as measured by gmfm total score and gross motor function classification (gmfc)-mld. cognitive function (measurements included performance and verbal iq scores) was maintained within normal range for most subjects, independent of their symptomatic status at the time of treatment. improvement or stabilization of central demyelination and peripheral nervous system (pns) abnormalities were observed in most subjects treated. conclusions: this interim analysis demonstrates that hsc-gt continues to be a safe and well-tolerated treatment for all mld subjects treated with a clinical follow-up ≤ years. all subjects achieved high levels of multi-linage engraftment, polyclonal hematological reconstitution and central and peripheral arsa activity reconstitution within or above normal levels. patients treated prior to symptom onset achieved a sustained clinical benefit in motor and cognitive function as well as on instrumental biomarkers of pns and cns demyelination, suggesting that autologous, ex-vivo hsc-gt is a highly promising therapeutic approach for li and ej mld pre-symptomatic subjects. further research is needed to support the benefit:risk profile in ej patients. clinical trial registry: nct https://www.clinicaltrials.gov/ct /show/nct ? term=nct &rank= disclosure: the san raffaele telethon institute for gene therapy (sr-tiget) is a joint venture between telethon and ospedale san raffaele (osr). ada-scid gene therapy (strimvelis) was licensed to glaxosmithkline (gsk) in and received european marketing authorization in . alessandro aiuti is the pi of the ada-scid long-term follow up clinical trial sponsored by gsk. strimvelis was licensed to orchard therapeutics (otl) in april . lentiglobin gene therapy in patients with sickle cell disease: updated interim results from hgb- background: β-globin gene transfer may reduce or eliminate complications in patients with sickle cell disease (scd). lentiglobin gene therapy (gt) comprises drug product (dp) made from autologous hematopoietic stem cells (hscs) transduced with the bb lentiviral vector (lvv) encoding β-globin with an anti-sickling t q substitution (hba t q ). the safety and efficacy of lentiglobin gt in adults with scd is being evaluated in a phase study, hgb- (nct ). patients were initially treated with dp made from bone marrow harvested (bmh) hscs (group a, fully enrolled), then from dp made from bmh hscs but using a refined manufacturing process (group b, fully enrolled), and subsequently from plerixafor mobilized hscs (group c, currently enrolling). methods: adults with severe scd (history of recurrent vaso-occlusive crisis, acute chest syndrome, stroke, or tricuspid regurgitant jet velocity of > . m/s) were enrolled. autologous cd + cells, collected by bmh or apheresis following mobilization with μg/kg plerixafor, were transduced with bb lvv. after myeloablative busulfan conditioning (area under the curve goal of [range - ] μm*min daily), patients were infused with the transduced cells and monitored for safety and efficacy. summary statistics are median (min-max). results: as of may , , patients had hscs collected, patients had dp manufactured and patients were treated. eleven patients ( in group a, in group b) underwent bmh and patients ( in group b [who also had bmh], in group c) underwent mobilization/ apheresis. median of . ( . - . ) x and . ( . - . ) x cd + cells/kg were collected per bmh (n= ) and per mobilization cycle (n= ), respectively. eighteen grade adverse events (aes) in patients were attributed to bmh and grade aes in patients to mobilization/apheresis. dp and treatment characteristics are shown in table . dp characteristics were improved in group b and group c vs group a. the safety profile post-dp infusion was consistent with myeloablative conditioning and underlying scd; most common non-hematologic grade ≥ aes were stomatitis, febrile neutropenia, and vasoocclusive pain. no grade ≥ dp-related aes, graft failure, veno-occlusive liver disease, replication competent lentivirus detection or clonal dominance were reported. at last visit (table ) , hba t q levels were higher in group b ( . - . g/dl) vs group a ( . - . g/dl). in group c patients at the -month visit, hba t q ( . [ . - . ] g/dl) levels were equal to or exceeded hbs levels ( . [ . - . ] g/dl). in group c patient at the -month visit, hba t q was . g/dl and total hb was . g/dl. conclusions: these data support the safety and feasibility of plerixafor-mediated hsc collection in patients with scd. hgb- protocol changes have improved lentiglobin dp characteristics yielding higher hba t q levels. additional data will determine the clinical effect of increased hba t q /hbs ratios. background: the prognosis of most patients with chemotherapy-refractory or multiply-relapsed cd + (a cell membrane protein) non-hodgkin's lymphoma (nhl) or hodgkin lymphoma (hl) still remain poor. targeting cd with monoclonal antibodies in hl and anaplastic large cell lymphoma (alcl) was shown to induce remarkable clinical activity; however, occurrence of adverse events (mainly neuropathy) may result into treatment discontinuation in many patients. immunotherapeutic approaches targeting cd by chimeric antigen receptor (car) has been demonstrated to be of value in two independent clinical trials (pmid: ) (pmid: ), although clinical benefit was sub-optimal. methods: we designed two rd generation, clinical-grade retroviral vectors carrying the cassette anti-cd singlechain variable fragment linked via cd hingetransmembrane domain, to the signaling domains of two costimulatory domains, namely either cd / - bb or cd /ox and cd -ζ. the inducible caspase- (icasp ) safety switch was also included in the constructs with the goal of promptly controlling undue toxicity. as a selectable marker, we added, in frame with the car molecule, a peptide derived from cd antigen. the in vitro anti-tumor efficacy was evaluated by using karpas , l or hdml- , in both short-term cytotoxic assay (represented by cr release assays) and long-term co-cultures ( days). cytokine profile upon antigen stimulation was characterized, as well as tcell exhaustion and memory marker profile. to assess the expansion, persistence, and antitumor effect of car.cd t cells in vivo, we used a nsg mouse model engrafted i.v. with human lymphoma cell lines (karpas and l ) genetically modified with ff-luciferase, this allowing the monitoring of tumor growth by ivis imaging system. persistence of car.cd t cells was evaluated every days, together with a deep characterization of memory profile and policlonality of persisting t cells. results: independently from the costimulatory domains cd /ox or cd / - bb, the generated retroviral vectors showed similar transduction efficiency of t cells ( . ± . % and . ± . %, respectively). nevertheless, car.cd incorporating cd .ox costimulatory domains was associated with more stable expression of the car over time, during extensive in vitro culture ( . ± . % vs . ± . % car+ t cells at days after transduction; p= . ). this finding was also associated with the evidence that car.cd -cd .ox t cells showed a superior anti-lymphoma in vitro activity as compared to car.cd -cd . bb t cells, when challenged at very high tumor/effector ratio ( : ). moreover, antigen-specific stimulation was associated to high levels of th cytokine production, with car.cd -cd .ox t cells secreting a significantly higher amount of ifnγ ( . ± . pg/ml), il ( . ± . pg/ml) and tnfα ( . ± . pg/ml) as compared to car.cd -cd . bb t cells ( . ± . pg/ ml, p= . ; . ± . pg/ml, p= . ; . ± . pg/ml, p= . ; respectively). in nsg mouse lymphoma models, we proved that car.cd -cd . ox t cells had an extensive superior anti-tumor control than car.cd -cd . bb t cells, leading to a significant reduction of bioluminescence at day ( . x vs . x , p= . ) and an increased overall survival of the treated mice ( % vs %, at days, p= . ). conclusions: overall, these data indicate that, in the context of car.cd t cells, the costimulatory machinery of cd .ox is crucial for improving both persistence and ultimately the antitumor efficacy of the approach. disclosure: nothing to declare background: acute graft-vs-host disease (agvhd) is a serious complication of allogeneic hematopoietic stem cell transplantation (allo-hsct). less than % of patients (pts) achieve sustained responses with first-line corticosteroid (cs) treatment. retrospective studies demonstrated clinical benefit with the janus kinase (jak) /jak inhibitor ruxolitinib (rux) in pts with steroid-refractory (sr) agvhd. here we present -month follow-up data from patients enrolled in reach (nct ), a phase trial evaluating rux plus cs in sr agvhd. methods: reach was a single-arm, open-label, multicenter study. eligible pts were aged ≥ years and developed grade ii-iv sr agvhd following allo-hsct from any donor source for hematologic malignancies. sr agvhd was defined as gvhd that progressed after days or had not improved after days of primary treatment with methylprednisolone ≥ mg/kg/d (or equivalent), development of gvhd in another organ after receiving ≥ mg/kg/d methylprednisolone for skin or skin plus upper gastrointestinal gvhd, or inability to tolerate cs taper. pts received rux mg twice daily (bid), with optional increase to mg bid in the absence of cytopenias. the primary endpoint was day overall response rate (orr), and the key secondary endpoint was -month duration of response (dor). orr was defined as the proportion of patients demonstrating a complete response (cr), very good partial response, or partial response. results: the study enrolled pts. median age was years, and . % were male. treatment was ongoing in pts ( . %) at data cutoff ( jul ) . at day , orr was . % (cr, . %). responses were observed irrespective of agvhd grade and sr criteria. best orr at any time during treatment was . % (cr, . %). median (range) time to response was ( - ) days. median dor with minimum months follow-up was days for both day responders ( figure ) and for pts who had a best overall response at any time during treatment. four pts ( . %) had malignancy relapse. overall, nonrelapse mortality at months was . %; results varied by day response (day responders, . %; other responders, . %; nonresponders, . %). median overall survival had not been reached for day responders. the most frequently reported hematologic treatment-emergent adverse events (teaes) were anemia ( . %), thrombocytopenia ( . %), and neutropenia ( . %). cytomegalovirus infection ( . %), sepsis ( . %), and bacteremia ( . %) were the most frequently reported infections. fatal rux-related teaes were sepsis and pulmonary hemorrhage ( pt each) and were attributed to both rux and cs. conclusions: in this first prospective trial of rux in sr agvhd, orr was . % by day and . % at any time during treatment. responses were rapid and durable. the ae profile was consistent with expectations for rux and pts with sr agvhd. rux represents a promising therapeutic strategy. background: chronic graft-versus-host disease (cgvhd) remains a major complication after allogeneic hematopoietic cell transplantation (allo-hsct). over the last decade, clinical success in patients with cgvhd has been hampered by the lack of insight into the complex pathobiological mechanisms of the disease and the paucity of specific therapeutic targets. although, it is now evident that the clinical manifestations of cgvhd are the result of a highly complex immune pathology involving both donor b cells and t cells as well as other cells. current work on immune cells involved in cgvhd pathobiology is limited by the number of parameters that conventional flow cytometry (fcm) can analyze because of cell autofluorescence and fluorescent dye spectral overlap. mass cytometry time-offlight (cytof) substitutes rare earth elements for fluorophores to label antibodies, which allows simultaneous measurements of more than parameters in single cells without correction for spectral overlap, and empowers us to understand cgvhd biology at the single-cell level. methods: we used mass cytometry with extensive antibody panels to perform in-depth immune profiling of peripheral blood samples from patients following allo-hsct, in which patients were without cgvhd, patients experienced moderate cgvhd and patients experienced severe cgvhd. the involved organs in patients with cgvhd are skin, liver and lung. results: we simultaneously stained cells with antibody panels created for this study. the t cell panel was designed to identify different populations of naive, memory, effector, regulatory, and exhausted t cells. the panels also included markers for the identification of b cells, natural killer cells, nkt cells, dc cells, plasma cells, granulocytes, and myeloid cells. in million measured cells, we identified immune cell phenotypes, in which there were t cell phenotypes, b cell phenotypes, monocyte phenotypes and granulocyte phenotypes. to generate a comprehensive view of the immune ecosystem of cgvhd, we generated two-dimensional maps of the data using the dimensionality reduction algorithm t-sne. this analysis showed a strong overlap between cgvhd of moderate and severe grades, but seperation from patients without cgvhd. seven immune compositions were identified to be cgvhd-associated. five distinct immune cells were correlated with specific cgvhd-involved organs (skin or lung), thereby presenting an in-depth human atlas of the immune cells in this disease. conclusions: this study revealed potential biomarkers and targets for immunotherapy of cgvhd and validated cytof as a valuable tool that can be used for immune profiling of cgvhd. disclosure: nothing to declare o t cell costimulation blockade with abatacept for acute graft-versus-host disease prevention in matched and mismatched unrelated donor transplantation: results of the first phase trial background: we performed a phase trial in adults and children to test abatacept for agvhd prevention ('aba '), based on our promising preclinical and pilot patient data. methods: aba had cohorts: a) hla mismatched (' / ', n= ), a single-arm study with pre-specified cibmtr matched analysis (vs cni+mtx or cni+mtx+atg). b) hla-matched (' / ', n= ) , randomized double-blind, comparing cni+mtx+placebo vs cni+mtx+aba ('aba'). abatacept dosing was mg/kg on d - , + , + , + . aba was designed as a screening phase trial, with relaxed type error ( . ) and standard type error ( . ). power analysis assumed reduction of gr - agvhd from %--> % in / s and %--> % in / s. median follow up = days ( / s) and days ( / s) . results: reduced grade - agvhd: aba was associated with decreased d gr - agvhd. in / s, gr - agvhd = . % (aba) vs % (cni+mtx) and % (+atg), ( -sided p = . , . ). in / s, gr - agvhd = . % in aba vs . % in placebo, ( -sided p = . ). reduced grade - agvhd in / s: aba was associated with decreased d gr - agvhd. in / s, gr - agvhd = % (aba) vs % (cni+mtx) and % (+atg, -sided p = . , . ). in / s, gr - agvhd = . % in aba vs . % in placebo ( -sided p = . ). chronic gvhd: for / s, yr cgvhd = . % (aba) vs . % (cni+mtx) and . % (+atg, p = . , . ). in / s, data collection is ongoing. safety indicators: there was no difference in neutrophil or platelet engraftment, cmv and ebv reactivation between aba and controls. cumulative incidence of relapse in / s at yr . % (aba) vs . % (cni+mtx) and . % (+atg) (p= . , . ). in / s, at yr, it was . % (aba) vs . % (placebo, p = . ). statistically significant survival advantage in / s: for / s, yr non-relapse mortality (nrm) = . % (aba) vs . % (cni+mtx) and % (+atg, p = . , . ). for / s, nrm = . % vs . % at yr (p = . ). severe agvhd free survival at months for / s = % (aba) vs % (cni+mtx) and % (+atg, p = . , . ). for / s = . % (aba) vs . % (placebo, p = . ). for / s, relapse-free survival (rfs) = . % (aba) vs . % in cni+mtx and . % in +atg (p = . , . ). for / s, rfs = . % for aba vs . % (placebo, p = . ). for / s, overall survival (os) = % (aba) vs . % (cni+mtx) and % (+atg, p = . , . ). for / s, os = . % (aba) vs . (placebo, p = . ). conclusions: our results suggest that short-course aba can safely prevent agvhd without compromising relapse. despite the modestly sized study, the comparative event size for / s was high enough that the protective effect of aba against gr - agvhd was highly significant. for / s, there was a statistically significant improvement for gr - gvhd and a trend toward an advantage in all parameters. for both cohorts, severe agvhd free survival was statistically-significantly improved. these results are the first to demonstrate efficacy of in vivo t cell costimulation blockade in preventing agvhd. background: in the nih cgvhd diagnostic classification, patients with gvhd after months are classified as either late agvhd or cgvhd. to date, this is only a clinical classification, with no biological differences identified. recently, the pbmtc / applied biomarker in late effects of children and adolescent (able) study completed accrual of pediatric allogeneic hematopoietic cell transplantation (hct) patients. we used day biomarkers to identify biological differences between cgvhd and late agvhd. methods: the pbmtc /able study with centers in canada, us, and europe prospectively collected peripheral blood samples at , , month post hct and at the onset of cgvhd in children. a comprehensive analysis for previously identified cgvhd immune cell markers by flow and cytokines by elisa on plasma and streck tubes shipped overnight and centrally evaluated at bc children's hospital. clinical data was collected centrally with a thorough central clinical adjudication by the pbmtc study committee. of those enrolled, were evaluable at day and classified as a) late agvhd (n = ), b) cgvhd (n = ), and c) controls that did not develop cgvhd (n = ). univariate analysis was performed comparing late agvhd, cgvhd, and no gvhd controls. significant differences were defined as a biomarker with both a roc auc ≥ . and p value ≤ . compared to controls. results: the profile of cgvhd included a cluster of abnormalities in memory and transitional b cells, conventional naïve and follicular helper t cells, and a loss of both recent thymic emigrant regulatory t cells and cd bright nk regulatory cells. four inflammatory cytokines, st , aminopeptidase n, cxcl and mmp (see table ) were increased. patients clinically identified as late agvhd had a more restricted biomarker pattern of limited b cell abnormalities and st . conclusions: late acute gvhd is limited to restricted b cell and elevation of st . cgvhd is characterized by the identical b cell abnormalities but with the additional loss of regulatory function in cd bright nkregs and rte treg cells. with the loss of regulatory function in cgvhd, there is an increase in cd lo b cells, follicular t helper cells, and additional cytokines. these prognostic markers findings may suggest therapeutic targets that differ for late agvhd compared to cgvhd. background: we are reporting the outcome of patients with steroid refractory acute graft versus host disease (sr-gvhd), treated with an anti-cd monoclonal antibody (begelomab r ). methods: twenty-eight patients were enrolled in two pilot studies eudract no. - - and no. - - , whereas patient were treated on a multicenter follow up compassionate use of the antibody. the median age of the patients was respectively and years. at the time of anti-cd treatment, gvhd was overall recorded as grade ii in patients, grade iii in and grade iv in patients. in the pilot sudies patients had failed line of treatment, wheas in the follow up compassionate use, patients had failed one line (n= ), two lines (n= ), three lines (n= ) or lines of treatment (n= ). results: there were no adverse events attributable to the antibody. day response was recorded in % and % in the pilot studies and follow up patients. response in grade ii gvhd was evaluable only in the pilot studies ( %); response in grade iii gvhd was recorded in % and % patients in the two groups; response in grade iv gvhd was recorded in % and % of patients in the two groups. overall there were % responses for skin and liver stage - , and % responses for gut stage - gvhd. the cumulative incidence of non relapse mortality (nrm) at months was % and %. for day responders, this figure was % and %, for non responders it was % and % in the two groups. the overall survival at year was % for the pilot studies and % for the follow up patients. conclusions: in conclusion, begelomab induces a high remission rate on day+ in patients with sr-gvhd, including a significant proportion of patients wih severe gut and liver gvhd. clinical background: cgvhd is characterized by an imbalance between effector and regulatory arms of the immune system that results in overproduction of inflammatory cytokines including il- and il- . moreover, a persistent reduction in the number of regulatory t (treg) cells limits the ability of the immune system to recalibrate this pro-inflammatory environment. kd is an orally available rho-associated coiled-coil kinase (rock ) selective inhibitor. in vitro data suggest that kd modulates immune homeostasis by shifting the th /treg balance towards a treg phenotype. methods: kd - is an open-label phase a study in patients with steroid-dependent cgvhd after no more than prior lines of treatment. three cohorts (c : mg qd (n= ), c : mg bid (n= ), and c : mg qd (n= )) were enrolled. the primary endpoint is overall response rate (orr), defined per the nih consensus criteria. results: as of -september- , the median duration of treatment was , and weeks for c , c and c , respectively. the median age was years (range - ) and median time from cgvhd diagnosis to kd treatment was months. % of patients had received ≥ prior lines of therapy and % had ≥ organs involved at baseline. patients remain on treatment with kd , with median duration of treatment of weeks (n= ), weeks (n= ) and weeks (n= ) for each cohort, respectively. the orr was % in c , % in c and % in c . responses were rapid, with % of responders achieving a response at the first assessment ( weeks). among responders, %, % and % have sustained responses for ≥ weeks in each cohort, respectively. responses were observed across all affected organ systems, including crs in upper gi, lower gi, esophagus, mouth, skin, joints/fascia, eyes, and liver. two patients with lung cgvhd achieved pr. % of patients achieved reductions in corticosteroid dose and patients discontinued corticosteroid treatment while receiving kd . % of responders achieved a clinically meaningful improvement (≥ -point reduction) in the lee symptom scale (lss) score. kd has been well tolerated. commonly reported aes (≥ % patients) were urti, ast/alt elevations, fatigue, nausea and diarrhea. grade ≥ aes occurring in > patients were ggt elevations (n= ) and hyperglycemia (n= ). no saes were considered related to study drug. two patients discontinued treatment due to aes considered possibly related to kd (headache, diarrhea). three fatal events occurred (relapse of leukemia; lung infection; cardiac arrest); none were considered related to kd . no increase in incidence of infection was observed. consistent with the postulated kd mechanism of action, th cells decreased and treg cells increased in patients receiving kd background: antithymocyte globulin (atg) treatment significantly decreases later development of chronic graftversus-host disease (cgvhd). one phase trial evaluating atg was the canadian bmt group (cbmtg) study that found that atg treatment resulted in significantly less cgvhd and dependence on immunosuppressive treatment at year. the exact mechanism by which atg decreases cgvhd is not known. we hypothesized that using known prognostic day cgvhd biomarkers in the wellcontrolled cbmtg trial represented an optimal approach to understand atg's biological impact on cgvhd in humans. methods: a separately developed cbmtg cgvhd biomarker study opened while cbmtg was ongoing and accrued patients (n = ; atg treated and n = controls) of the cbmtg patients. samples were collected at , , and months and at the onset of cgvhd and evaluated at bc children's' hospital research institute, vancouver, bc. patients were evaluated for day immune cellular markers previously associated with later cgvhd including: a) naive helper t (th) cells, b) recent thymic emigrant (rte) th cells; c) cd low b cells; d) cd bright nkreg cells; and e) treg cells. frequencies in the atg treated and control group were evaluated to detect significant difference using non-parametric t-test mann-whitney test. results: patients of this subpopulation (aged - years) were shown to be representative of the larger cbmtg study population. the atg treated group had a significant decrease in total t cells, rte th cells, and naïve th cells at day compared to the control population (p < . each population -see table ). atg treatment had no impact on tregcells, cd + b cells, and cd lowb cells but there was a significant increase in cd bright nkreg cells (p < . ). we evaluated the ratio of naïve th effector cells to the regulatory nkregcell population and saw a > fold difference the atg treated group (naïve th cell:nkregcell ratio = . ) compared to untreated patients (ratio of . ; p < . ). in this small population we found that the naïve th cell:nkregcell ratio was also high prognostic for later development of cgvhd ( . vs. . ; p < . ). conclusions: these results suggest that atg's major mechanism of action is related to its ability to simultaneously inhibit naïve th cells and enhance cd bright nkreg cells after transplantation. while these results require confirmation, they support strategies that target the ratio of nk reg cell and naive cd + t cells to modulate cgvhd. clinical trial registry: nct disclosure: none of the authors have any conflicts of interest to declare o immune reconstitution -based score at diagnosis of cgvhd predicts gvhd severity and overall-survival: a novel prognostication tool for gvhd treatment tailoring background: allogeneic stem cell transplantation (hsct) survivors are at a relevant risk of developing chronic gvhd (cgvhd), which importantly affects quality of life and increases morbidity and mortality. early identification of patients at risk of development of severe cgvhd related morbidity would be a relevant tool to tailor preventive strategies. we have previously demonstrated the role of immune reconstitution (ir) as predictive biomarker of occurrence of cgvhd. the aim of this study was to evaluate the prognostication power of ir at cgvhd onset through a new ir-based score. methods: we analyzed clinical data from adult patients consecutively undergoing first allogeneic hsct transplant between january and december at our institution. a written consent was given for the use of medical records for research. patients were divided into a test cohort ( pts) and a validation cohort ( pts). median follow-up for surviving patients was years.we built a cox multivariate models for os in patients with cgvhd of any severity. variables included in the models were: patient age (according to median value), r-dri score, type of donor (matched related donor vs matched unrelated vs haploidentical), main gvhd prophylaxis platform (atg-based vs ptcy-based vs neither of the two), ir values (cd , cd , nk, iga, igm according to median values) at cgvhd diagnosis, history of prior agvhd, karnofsky ps, plt < . /μl, alc< /μl, eos < /μl.once we identified the variables independently predicting os by multivariate analysis, we derived a formula for a prognostic risk index by using the β coefficients found in the model. each patient was then assigned a score and we defined three groups of os risk (low, intermediate and high) by dividing the score into three classes using the first and third quartiles. finally, to evaluate predictive performance of the ir-score we calculated the receiver operating characteristics (roc) curve via the area under the curve (auc), to summarize the ir-score ability to correctly classify events and non-events. results: patients ( test-cohort, validationcohort) were evaluated for cgvhd and outcome. our multivariate model defined the variables independently predicting os at cgvhd onset: cd + count > / mcl, nk count < /mcl, igm < . g/l, karnofsky ps < %. final score was calculated as follows: , (if cd > / mcl) + , (if nk < /mcl) + , (if igm < , ) + , (if karnofsky < ). low risk patients were defined as having a score ≤ . , intermediate > , and ≤ . , high risk > . . the y-os for low risk patients was %, for intermediate % and for high risk % in the test-cohort and %, % and % in the validation-cohort ( figure a-b) . the roc curve analysis supports the validity of the ir-score in our cohort of patients -auc . %, with % confidence intervals higher than %. furthermore ir-score was able to stratify across nih-severity classification (figure c). conclusions: immune-reconstitution score at diagnosis of cgvhd predicts gvhd severity and overall-survival. irscore could be adopted to identify patients at high risk and modulate cgvhd treatments accordingly. disclosure: chiara bonini has research contract with intellia therapeutics. the other authors declare that they have no conflicts of interest. haploidentical transplantation with sirolimus-based gvhd prophylaxis and unmanipulated pbsc graft: background: haploidentical transplantation has emerged as a viable option for patients lacking a fully matched donor. we firstly explored the association of sirolimus and atg, later followed by sirolimus with pt-cy as gvhd strategy. herein, we describe long-term outcomes of haploidentical hsct using sirolimus-based gvhd prophylaxis. methods all patients received sirolimus and mmf as gvhd backbone prophylaxis plus atg in patients, and pt-cy in . conditioning regimen was based on treosulfanfludarabine; recipients of pt-cy transplants were more likely to receive a regimen intensified by a nd alkylating agent (melphalan or thiotepa). median follow up was longer in atg group ( vs monhs, p< . ). there were no differences in dri. results: the majority of patients reached the neutrophil ( % in atg group vs % in pt-cy group) and platelet ( % vs %) engraftment within days after hsct. immune-reconstitution was broad and fast, reaching more than /ml cd + t cells within a median of vs days. the two groups were similar in terms of survival and main transplant outcomes. in the atg group, the cumulative incidence of grades ii-iv and iii-iv acute gvhd at days was % and %. corresponding rates after pt-cy were % and %. the cumulative incidence of overall and severe chronic gvhd was % and % at years in atg group and % and % after pt-cy .the cumulative incidences of relapse and nrm in atg group were respectively % and % at years. corresponding rates after pt-cy were % and %. in atg group, -year os was %, while grfs was %. the corresponding probabilities after pt-cy were % and %. the only difference reported was a better pfs in favour of pt-cy ( % vs %, p= . conclusions: extended follow-up in patients confirms sirolimus-based gvhd prophylaxis as feasible and safe in haploidentical hsct based on unmanipulated pbsc graft. both atg and ptcy association to sirolimus provide an effective prevention of gvhd and translate into a similar long-term overall survival. a significant advantage of sir-pt-cy on relapse rate warrants further investigation. background: steroid-refractory graft-versus-host disease (sr-gvhd) is still responsible for high mortality in patients undergoing allogeneic stem cell transplantation; a number of agents is currently available in case of steroid-refractoriness, yet there is so far no consensus about a standard second-line treatment and overall survival (os) remains poor.α -antitripsyn (αat) is a circulating -kda serine protease inhibitor found to enhance the production of anti-inflammatory cytokines and to favor the expansion of regulatory t-cells; it has therefore been tested in situations of altered tolerance and disproportionate inflammation, including gvhd. two studies showed that treatment of sr acute gvhd with αat is feasible and effective. methods: we retrospectively analyzed a series of patients who received exogenous αat for sr acute gastrointestinal gvhd or overlap gvhd with acute gut features. sr-gvhd was defined and graded according to standard criteria. αat was administered intravenously at a loading dose of mg/kg at day followed by mg/kg /day every other day for a total of doses. response to treatment was defined according to published criteria; os was estimated with the kaplan-meier method.a panel of cytokines and immune cell subsets were measured before treatment and once weekly during treatment by a luminex assay and by flow cytometry, respectively. results: sixteen patients were treated for gut gvhd between september and march . median age was years (range - ). αat was administered at a median time of days from transplantation (range - ) and of days from gvhd onset (range - ) . acute gvhd was scored as grade ii in % of patients, grade iii in %, grade iv in %. sixty-seven percent of patients had already received one or more lines of treatment other than steroids, including ruxolitinib, etanercept, atg; orr was % with a cr rate of %; median time to best response was days (range - ), with a continued orr at day of %. the overall rate of gastrointestinal responses was %. median follow-up of living patients was days (range - ); median os was days and -year os was % ( % ci % - %).the most common infectious event was cmv reactivation ( %); grade - infectious complications were recorded. there was no quantitative deficiency of blood aat levels before treatment ( . g/l ± . ); blood αat rose significantly during and after treatment. baseline αat level didn´t differ between responding and nonresponding patients. a cytokine profile was evaluable in patients; no statistically significant increase or decrease in cytokine plasma concentration after αat infusion was observed. surprisingly, a decrease in circulating t regs after exposure was found (p= . ), regardless of patients´responding status. conclusions: treatment with αat was safe and effective in a cohort of sr-agvhd high risk, pre-treated patients and should be considered as a possible alternative. changes in the cytokine milieu and t-cell subsets shown in murine models were not observed in a real-life setting. table . peripheral blood was used as graft source in % of the patients in the atg group and in % in the pt-cy group. gvhd prophylaxis consisted in atg mg/m days - to - , mtx days + , + , + and + , and csa from day - in the atg group. the pt-cy group received cyclophosphamide mg/kg/d on days + and + , followed by either csa or tacrolimus and mycophenolate mofetil (mmf) from day + in patients ( %), cyclophosphamide on days + and + combined with csa or tacrolimus from day in patients ( %), or cyclophosphamide on days + and + combined with tacrolimus and sirolimus from day + in patients ( %). cumulative incidence at days of grade ii-iv ( % vs %, p= . ) and iii-iv ( % vs %, p= . ) acute gvhd, were significantly higher in the mtx-csa group ( figure a ). there were no differences in the -year cumulative incidence of chronic moderate to severe gvhd between the atg and the pt-cy group ( % vs %, p= . ). after a median follow-up of months for the atg group and months for the pt-cy group, -year overall survival (os) was higher in the pt-cy group ( % vs %) although not statistically significant (p= . ) ( figure b) . we found no differences between both cohorts in -year event-free survival (efs) ( % and %, p= . ) and the composite endpoint of gvhd-free and relapse-free survival (gfrs) ( % vs %, p= . ). the -year cumulative incidence of relapse was significantly higher in the pt-cy group ( % vs . %, p= . ) and non-relapse mortality (nrm) at -years was higher in the atg group ( % vs %) but not statistically significant (p= . ). conclusions: in our experience, in spite of the limited number of patients, gvhd prophylaxis using pt-cy combined with additional immunosuppression after mud hsct, using mostly peripheral blood as graft source, reduced the cumulative incidence of agvhd compared to standard prophylaxis with mtx-csa. prospective studies with longer follow-up are needed to confirm these observations. disclosure: nothing to declare o abstract already published. methods: consecutive patients who underwent allo-sct for hematological malignancies between january and august were included in this prospective singlecentre protocol. all patients had at least one baseline risk factor predicting development of severe gvhd (e.g. hla mismatch, fem-to-male sex mismatch). pt-cy in combination with a second immunosuppressive drug was used as gvhd prophylaxis. results: patients characteristics are summarized in table . median age was (range, - ) years, with male patients ( %) receiving a graft from a female donor. more frequent allo-sct indications were acute leukemia and mds ( . %) followed by nhl ( %). eleven patients ( %) were transplanted in advanced status. donor was a sibling, matched unrelated or mismatched unrelated in %, % and % respectively. seven patients ( %) received a myeloablative conditioning regimen including tbi ( or . gy) while the remaining ( %) received a ric/rtc regimen based on fludarabine, busulfan or melphalan +/-thiotepa ( mg/kg). median follow-up for survivors was days (range: - ). median time to neutrophil and platelet engrafment were + ( - ) and + ( - ) days, respectively (g-csf not routinely used). early toxicity was low, without cases of thrombotic microangiopathy, only cases of drug-related renal failure ( . %) and case of possible vod. before the introduction of mini-thiotepa in the ric protocols (flubu/ flumel) there was case of primary graft failure (gf) and cases of late graft failure ( / ; %); cases were successfully regrafted with the mini-thiotepa ric. there have been no further cases of gf after its introduction (n= evaluable cases + second salvage allorics). additional potential risk factors for gf were cd + ≤ x e /kg (p= . ) and a high lymphocyte count at stem cell infusion (p= . ). the ci of grade - acute gvhd at day + was % ( %ci: - ) with only cases of refractoriness to steroids. of the evaluable patients, only developed moderate chronic gvhd leading to a yr-ci of %. nrm was . % at yr and the ci of relapse was % ( % ci: - ). all relapses occurred in patients with intermediate/ high rdri. yr-os was % and the estimated -year dfs was . %. conclusions: these outcomes confirmed the feasibility of both ric and mac allo-sct using pt-cy as soc with a single immunosuppressive drug in patients at high risk of gvhd. an important observation was the high rate of gf with "classical" alloric platforms (flubu/flumel), which appears to be lower after introducing the mini-thiotepa. although these data need confirmation in larger cohorts, the current results suggests that pt-cy may pave the way to improving the quality of life of transplant survivors by markedly reducing severe gvhd. protection of the endothelium during steroid-refractory gvhd background: clinical data demonstrated that endothelium related factors predict mortality after the diagnosis of agvhd, suggesting that the endothelium may be involved in the pathobiology of steroid-refractory agvhd (sr-agvhd) (j clin oncol. mar ; ( ) methods: intestinal biopsies from patients after allosct. murine agvhd models balb/c→b , b →bdf and lp/ j→b with and without steroid treatment. immunostaining, electron microscopy, light sheet fluorescence microscopy (lsfm), facs. in vivo and in vitro assays for endothelial dysfunction. treatment with phosphodiesterase type inhibitor (pde ) in sr-agvhd models. results: we found a significant higher percentage of apoptotic vessels in duodenal and colonic mucosa biopsies of patients with grade iii-iv agvhd compared to no gvhd ( figure a ). in murine experimental agvhd, we detected severe microstructural endothelial damage and reduced endothelial pericyte coverage accompanied by reduced expression of endothelial tight junction proteins leading to increased endothelial leakage in agvhd target organs. during intestinal agvhd, colonic vasculature structurally changed, reflected by increased vessel branching and vessel diameter ( figure b) . we analysed human biopsies and murine tissues from sr-gvhd vs. naive (untreated) agvhd and found significantly lower lymphocyte infiltration in sr-gvhd, demonstrating low inflammatory activity ( figure c ). our findings suggest that endothelium-related and t cell independent mechanisms play a previously unrecognized role during sr-agvhd, providing the rationale for t-cell independent treatment strategies. as a first example for such an approach, we tested the endothelium-effective pde inhibitor sildenafil and found reduced apoptosis as well as improved metabolic activity of endothelial cells in vitro. in accordance, sildenafil treatment resulted in improved survival and reduced target organ damage during experimental sr-agvhd ( figure d ). conclusions: we show profound endothelial involvement after allo-hsct and demonstrate that endothelialprotection with sildenafil ameliorates sr-agvhd, providing a novel non-immunosuppressive treatment approach. these results can serve as rationale for translational development of endothelium-based therapies for sr-agvhd. disclosure: the authors declare no confilct of interest relevant to this study o abstract already published. the therapeutic effect of immune-modifying microparticles in an acute graft-versus-host disease model john galvin , sara beddow , stephen miller university of illinois chicago, chicago, il, united states, northwestern university, chicago, il, united states background: inflammatory monocytes are recruited to target organs during acute graft versus host disease (agvhd). as seen in other autoimmune disorders, inflammatory monocytes play an important role in antigen presentation and cytokine production. these actions allow for a sustained activation and proliferation signal to t-cells. previous studies have shown that imp treatment in mouse models of colitis, encephalitis, myocardial infarction and peritonitis markedly reduced monocyte accumulation in the affected end-organs --promoting tissue repair; reducing disease symptoms and increasing survival. therefore, our objective was to test clinical outcomes after imp treatment in a mouse model of agvhd. methods: murine agvhd model: balb/c mice were given cgy total body irradiation, irradiated balb/c mice were transplanted with × c bl/ bone marrow cells and × c bl/ spleen cells via tail vein.imp treatment: imps were made with plga (phosphorex inc, hopkinton ma) was administered to the recipient mice ( . mg/kg body weight) by iv daily starting from day to day after bone marrow transplantation (bmt). pbs at the same volume was used as vehicle control. in vivo bioluminescence imaging: mice were given an intraperitoneal injection of luciferin ( mg/kg body weight) and then anesthetized and imaged using the ivis imaging system (xenogen). imaging data were analyzed and quantified with living image software (xenogen). results: imp treated mice had significantly less severe acute gvhd symptoms (average score of . ) than the untreated bm+sp group (average score . ) starting at the time of imp treatment (days - ) and remained with significantly reduced symptoms for the day course (figure ). imp treatment also rescued bm+sp mice from agvhd associated mortality with a -day overall survival of % compared to % in the untreated bm+sp group (figure ). intestinal tissue from the imp treated mice compared to the bm+sp mice demonstrated less evidence of agvhd (an average score of . and . , respectively). hepatic tissue from the imp treated mice compared to the bm+sp mice demonstrated less evidence of agvhd (an average score of . and . , respectively). imp treatment also significantly reduced inf-γ levels in the intestinal tissues of treated mice compared to untreated bm +sp mice. in the mice infused with lymphoma cells (a -luc), imp treatment reduced agvhd symptoms and death while preserving the gvl effect. conclusions: our results demonstrate that imps significantly reduce symptoms and mortality in a murine model of agvhd while preserving gvl. the reduction in inflammatory monocytes with imps leads to a reduction in inflammatory cytokines, hepatic lymphocyte infiltration and intestinal mucosal denudation. these findings highlight the potential of imp therapy as a specific and potentially safe treatment in acute gvhd. [[o image] . figure background: despite significant improvements in the supportive sickle cell disease (scd) causes substantial morbidity and mortality. allogeneic hematopoietic stem cell transplantation (hsct) is currently the only curative option but is only offered if a matched sibling donor (msd) is available. with a msd availability of < % t cell depleted hsct from a haploidentical donor (t-haplo-hsct) is a potential alternative. methods: patients (pts) with advanced stage scd (asscd) were transplanted with a cd + /cd + or αβ/cd + depleted t-haplo-hsct ( pts, median age years, range - years) or with bone marrow (bm) from a msd, pts, median age , range - years). indication for hsct was asscd with multiple scd related complications. all pts underwent exchange transfusion before hsct. in all pts the conditioning regimen consisted of treosulfan, thiotepa, fludarabine and atg. immunosuppression was carried out with cyclosporine a or tacrolimus and mycophenolate mofetil. the control group received a msd bm allograft. results: in the t-haplo-sct group the pts received a cd + or αβ + t-and cd + b-cell depleted peripheral stem cell allograft with , x cd + cells/kg body weight (range , to , ) . all pts with a median follow up of months (range - months) in the msd group and from pts with a median follow up of months (range - months) in the t-haplo-sct group are alive. engraftment was achieved in all pts with stable chimerism over %, except for pts with a stable mc in the t-haplo sct group and patient in the msd group, but complete engraftment of red cell precursor in the bm. all pts are off immunosuppression with a stable almost complete chimerism. the conditioning regimen was well tolerated with no case of high-grade transplant related morbidity. the post-transplantation complications were comparable in both groups. one patient developed after severe rotavirus gastroenteritis a severe cmv pneumonitis and succumbed to an uncontrolled cmv pneumonitis. one patient in the t-haplo sct group suffered from a late graft failure and developed a macrophage activation syndrome. he died in a septic event.none developed a glucksberg grade iii-iv agvhd and in the t-haplo sct group pts ( %) and in the msd group pts ( %) developed a steroid sensitive mild to moderate cgvhd with symptoms of fasciitis, oral as well as mild cutaneous gvhd. in both groups no severe or steroid refractory cgvhd was observed. conclusions: our results demonstrate increasing evidence for the safety and efficacy of cd + /cd + or αβ + /cd + depleted haploidentical hsct in asscd. the treosulfan based conditioning regimen was an excellent alternative to busulfan with a low incidence of transplant related morbidities and therefore most suitable for pts with scd. these results open the option of a curative therapy for almost all scd pts without a msd. disclosure: nothing to declare. abstract already published. hematopoietic stem cells o abstract already published. comparison of outcomes post allogeneic hematopoietic cell transplantation using fresh versus cryopreserved peripheral blood stem cell grafts background: cryopreservation is routine practice with autografts, however in the allogeneic hct setting the effects of cryopreservation have not been thoroughly investigated. we sought to compare allogeneic hct outcomes using fresh versus cryopreserved grafts in a large single centre cohort. methods: between and , we retrospectively reviewed consecutive adult patients who underwent allogenic peripheral blood hct at our centre. outcomes assessed included platelet (≥ x e /l) and neutrophil (≥ . x e /l) engraftment, occurrence of acute graft-versushost disease (gvhd) in the first days, overall survival (os), cumulative incidence of relapse (cir) and non-relapse mortality. results: median follow up of survivors was months (range - months). fresh grafts were received by patients, received cryopreserved grafts, median age at hct was and years respectively. transplant indication was myeloid malignancy in ( %), lymphoid in ( %) patients. myeloablative regimens were used in ( %) patients. the majority of fresh grafts were from unrelated donors ( %) while most cryopreserved grafts were from matched related donors ( %). in vivo t-cell depletion was performed in % of fresh and % of cryopreserved transplants. median time to neutrophil engraftment for fresh versus cryopreserved grafts was and ( - ) days respectively, while median time to platelet recovery was ( - ) and days, respectively. for fresh versus cryopreserved grafts, grade ii-iv acute gvhd was seen in % and %, respectively (p= . ) while grade iii-iv acute gvhd was seen in % of patients in both groups (p= . ). on univariate analysis, os for the entire cohort at years was % ( %ci - %) and at years was % ( %ci - %). two and year os was % ( %ci - %) and % ( %ci - %) respectively for fresh grafts and % ( %ci - %) and % ( %ci - %) respectively for cryopreserved grafts (p= . ). cumulative incidence of relapse (cir) of the entire cohort at years was % ( %ci - %) and at years was % ( %ci - %). two and year cir was % ( %ci - %) and % ( %ci - %) respectively for fresh grafts and % ( %ci - %) and % ( %ci - %) respectively for cryopreserved grafts (p= . , figure ). [[o image] . figure ] multivariable analysis for os verified no significant difference between fresh versus cryopreserved grafts (p= . ). for cir however, cryopreservation was the only independent predictor of relapse (hr . for cryopreserved, %ci . - . , p= . ), while for nrm cryopreservation was not an independent predictor of increased risk (p= . ). when the multivariable analysis was repeated for related donor transplants only (n= , cryopreserved grafts, fresh), this confirmed the independent increased relapse risk for cryopreserved grafts (hr . , p= . ) . conclusions: we confirmed on univariate and multivariable analysis that there is no significant difference in os between allogeneic transplants performed with fresh versus cryopreserved peripheral blood stem cell grafts, however there is a significant increase in relapse risk associated with cryopreservation. disclosure: nothing to declare. background: high dose post-transplantation cyclophosphamide(ptcy) used in haploidentical transplantation (haplo-sct) has demonstrated to be highly effective in acute and chronic gvhd prophylaxis; however it is associated with high relapse rates. anti-t-lymphocyte globulin (atg-fresenius®) is also effective as gvhd prophylaxis but its benefit in overall survival (os) and relapse free survival (rfs) is unclear. the aim of this study was to compare the effectiveness of two gvhd prophylaxis regimen employed in high risk transplantation: ptcy-haplo-sct and low doses of atg-f used in peripheral blood(pb) and mismatched transplantation. the primary endpoint was to evaluate the incidence and severity of agvhd and cgvhd. as secondary endpoints we analysed the os, rfs and grfs (considering as events severe agvhd, systemic therapy-requiring cgvhd, relapse or death). we also evaluated mortality related to transplantation(trm) and post-transplant complications. methods: we retrospectively analysed allo-sct performed in our institution between and . we analysed two cohorts: haplo-sct with ptcy ( mg/ kg, days + ,+ ) followed by tacrolimus and mycophenolate (mmf); and pb and/or mismatched transplants with low dose atg-f ( mg/kg days - ,- ,- ) associated to calcineurin inhibitors starting on day - , with short course mtx (days + ,+ ,+ ) or mmf. in both cohorts, mmf was stopped on day + and calcineurin inhibitors were tappered on day + . comparing both groups, we found differences in diagnosis (lymphoproliferative disorders vs ,p= . ), high dri-score ( vs , p= . ), previous transplantation ( vs ,p= . ), reduced-intensity conditioning regimen ( vs ,p< . ) and bone marrow as stem cells source ( vs , p< . ). results: median time to neutrophil engraftment was similar in both groups: vs days. conversely, median time to platelet recovery was longer in ptcy cohort ( vs days, p= . ). there were no differences in agvhd incidence (ptcy . % vs atg . %) or severe agvhd (ptcy . % vs atg %, p= . ). the global cgvhd incidence was pcty . % vs atg %. mild, moderate and severe cgvhd incidence was . %, . % and . %, for ptcy vs %, % y % for atg(p= . ).with a median follow-up of months ( months for ptcy and months for atg) the os at and was: pcty . % and %, and atg . % and . %,p= . ). rfs ( and months) was: % and . % for ptcy, and, % and % for atg(p= . ). grfs at and months was . % and . %, for ptcy patients and . % and % for atg patients (p= . ). ptcy cohort seemed to develop more relevant non-infectious complications, but there were no differences among infectious complications. image . trm was similar in both cohorts: pcty . % vs atg . % (p= . ). we neither found differences in early toxic mortality (< days): pcty . % vs atg . % for atg (p= , ). conclusions: regardless the different transplant scenarios in which they were used, ptcy and low dose atg seem to be equally effective in the prophylaxis of severe forms of acute and chronic gvhd, offering similar grfs. moreover,they show similar early toxicity and rate of infectious events. disclosure: we have nothing to disclose. conclusions: emapalumab treatment promoted disease control, blunting the exacerbated immune response typical of the disease, with a favorable safety and tolerability profile. the results of the study also suggest that emapalumab may contribute to optimize post-transplant outcome of patients given hsct. based on these data, emapalumab received marketing authorization in the us from the food and drug administration for the treatment of patients with primary hlh with refractory, recurrent or progressive disease or intolerance with conventional therapy. background: allogeneic hematopoietic stem cell transplantation (hsct) is a potentially curative treatment for some inherited disorders, including selected primary immunodeficiencies (pids). in the absence of a wellmatched donor, hsct from a haploidentical family donor (hifd) may be considered. various approaches are being developed to mitigate the risks of graft failure and graftversus-host disease (gvhd) and to speed-up immune reconstitution. among those, high-dose, post-transplant cyclophosphamide (ptcy) is increasingly used in adult recipients. however, data on ptcy in children and those with inherited disorders in particular are scarce. methods: we reviewed the outcome of children transplanted with hifd and ptcy for pid (n= ) or osteopetrosis (n= ) in a single center. median (range) age was . years ( . to )). patients in our series had major risk factors for poor post-hsct outcome such as active viral infections at the time of transplantation (n= ), ebvrelated lymphoproliferation in partial remission (n= ), previous kidney transplant (n= ). hsct with ptcy was a primary (n= ) or a rescue procedure after graft failure (n= ). conditioning regimen was myeloablative in most primary hscts and non-myeloablative in all rescue procedures. results: after a median follow-up of . months, of the patients engrafted. twenty-one patients are alive and have been cured of the underlying disease. the two-year overall survival rate was . %. the cumulative incidences of acute gvhd grade ≥ ii, chronic gvhd and autoimmune disease were %, . %, and . %, respectively. there were only two cases of grade iii acute gvhd, all cases of cgvhd were limited and allowed to stop systemic immune-supression, autoimmunity consisted in autoimmune hemolytic anemia in remission at last follow up, vitiligo and thyroiditis. the cumulative incidence of blood viral replication and life-threatening viral events were % and . %, respectively. there was evidence of early t cell immune reconstitution including early anti-viral responses. conclusions: in the absence of an hla-identical donor, hifd hsct with ptcy is a viable option for patients with life-threatening inherited disorders. disclosure: nothing to declare background: griscelli syndrome (gs) is a very rare autosomal recessive disease, characterized by skin hypopigmentation and silvery-gray hair. gs type (gs ) patients suffer immunodeficiency and potentially fatal episodes of macrophage activation known as accelerated phases during early childhood. the only curative treatment modality for these patients is allogeneic hematopoietic stem cell transplantation (hsct). we report the outcome of hsct in children with gs . to date, this is the largest cohort of gs patients who have undergone transplantation at a single center. methods: we retrospectively reviewed consecutive patients with gs who underwent hsct at our institution between january -december . median age at diagnosis and at transplant was . ( - ) and . ( . - . ) years respectively. prior to hsct, ( %) and ( %) patients had life-threatening accelerated phases (hlh) and cns involvement respectively. all such patients were treated with chemotherapy and achieved remission at the time of hsct. the source of grafts was matched related marrows in ( . %) pts and partially mismatched unrelated cord in ( %) pts. two patients received haploidentical and matched unrelated marrows respectively. conditioning regimens were myloablative doses of busulfan/cyclophosphamide and busulfan/fludaribine in ( %) and ( %) patients respectively. all patients received gvhd prophylaxis. growth factors were administered in ( . %) and median cd dose used was . ( . - . ) ^ and . ( . - . ) ^ per kg of body weight for marrow and cord respectively. results: post infusion rate of engraftment was . % ( cases) with median time to neutrophil and platelets recovery were ( - ) and ( - ) days respectively. cumulative incidence of acute gvhd was . % ( ), with an overall grade of i, ii and iv as %, % and % respectively. the post-transplant course was complicated by cmv infection, ebv viremia and veno-occlussive disease in ( %), ( %) and ( %) patients respectively. chimerism studies at the last contact were available for patients. full donor cell chimerism ( %) was seen in ( %) of the transplanted patients. post-transplant two patients experienced disease reactivation at a median time . ( - ) days. with a mortality rate of . % ( ) and a median follow-up time of . months, five-year cumulative probability of overall survival (os) for our cohort of patients was . ± . . transplant related mortality counted as death within day was . % ( ). prominent causes of death were septic shock followed by ards. cumulative probability of five years overall survival was significantly better in those who did not have hlh prior to sct ( . ± . vs. . ± . , p-value: . ). of the patients with neurological involvement before hsct, survived with residual sequelae in patients. os at five years was . ± . and . ± . (p-value: . ) in pts with and without cns involvement at presentation respectively conclusions: hsct in patients with gs is potentially curative with long-term, disease-free survival. early hsct before the development of the accelerated phase showed better result. [ background: primary immunodeficiencies (pids) are heterogeneous inborn disorders resulting in impaired cellular and/or humoral responses. pid present a wide range of clinical manifestations, ranging from infections to autoimmune, inflammatory and/or malignant complications. allogeneic stem cell transplantation (allosct) is curative for pediatric pid with an excellent safety. in adults on the other hand, this therapeutic approach remains controversial. methods: this is a retrospective, monocentric study of consecutive adult patients with pid who underwent an allosct between and . the objective was to assess the feasibility, effectiveness and safety of this procedure. results: twenty-two ( %) of patients presented an inherited t or b cell deficiency and ( %) a phagocyte impairment. twenty ( %) patients had a genetic diagnosis. besides infectious complications, ( %) patients had an history of lymphoma and ( %) an history of autoimmune/ inflammatory complication. the median age at transplant was years (range - ). twelve ( %) patients received a myeloablative conditioning (mac), ( %) a reduced intensity conditioning (ric) and ( %) had no conditioning (second transplant for severe combined deficiency). mac included fludarabine (flu)/buslfan (bu, dosage: . mg/kg) based regimen (n= ) and bu ( . mg/kg)/cyclophosphamide (cy) based regimen (n= ). ric included based flu/bu (range dosage: . - . mg/kg) regimen (n= ), flu/cy based regimen (n= ), thiotepa/flu/bu regimen (n= ) and low dose tbi based regimen (n= ). among the patients who received a conditioning, ( %) received alemtuzumab and ( %) received antithymocyte globulin as part of the conditioning. the stem cell source was bone marrow for ( %) and peripheral blood stem cells for ( %). the donor was matched related for ( %), matched unrelated for ( %), mismatch unrelated for ( %), and haplo-identical for ( %). all assessable patients had a successful engraftment. eight ( %) presented a grade ii-iv acute gvhd (one grade iii, grade iv), and ( %) a chronic gvhd ( limited and extensive). with a median follow-up of months ( - months) post-transplant, the -years overall survival (os) was . %. the transplant related mortality was . % with deaths. among them, both patients transplanted with an haplo-identical donor died. all deaths occurred in the firstyear post-transplant. thus, the use of an haploidentical donor was associated with an adverse outcome. conversely, neither the conditioning nor the stem cell source was associated with a worse outcome. except one patient with an history of aggressive b cell lymphoma who relapsed few months after allosct, no patient with an history of lymphoma relapsed. after a salvage treatment, the relapsing patient remained in complete remission years later. at last follow-up, all surviving patients had a stable, mixed donorrecipient or full donor chimerism without any sign of active infection. conclusions: allosct in the pid setting is an effective therapeutic with an acceptable toxicity and should be considered in case of severe infectious, inflammatory and/or malignant complications in young adult patients with pid when an appropriate donor is available. disclosure methods: we analyzed the results of allogenic hsct with tcrαβ/cd graft depletion in patients with various pid (excluding classic scid) who received hsct from may to september in our center. the median age at hsct was , years (range , - , ). patients received hsct from matched unrelated, haploidentical donors, -siblings. the conditioning regimens included: fludarabin (flu) mg/m with alkylator (treosulfan (treo) - g/m ) in patients, with alkylators (treo - g/m with melphalan mg/m or thyotepa mg/kg) in . twenty-five patients received alkylators with addition of g-csf mg/kg and plerixafor mg/kg. seventeen patients with nijmegen breakage syndrome (nbs) received reduced intensity conditioning with busulfan mg/kg or treo mg/m , cyclophosphamide mg/kg and flu mg/m . in all but patients serotherapy was used: patients -horse atg - mg/kg, -rabbit atg (thymoglobulin) mg/kg, - mg/kg anti cd monoclonal antibodies. in patients two or more immunosuppressive drugs were used after transplantation (combination of calcineurin inhibitor (cni) with short course of methotrexate, or mycophenolate mofetil, or abatacept), patients received one immunosupressive agent (cni). from november no posttransplant immunosuppression was used ( patients). primary end points were: incidence of acute gvhd, graft failure (gf) and transplant related mortality (trm). because of the supposed influence of conditioning regimen on gf and trm incidence, to the analysis of gf and trm were included only patients with similar conditioning regimen with alkylators (excluding patients with nbs, with alkylator and addition of g-csf and plerixafor in conditioning regimen). median follow up after hsct was , years (range , - , years). results: overall survival (os) in patients was , ( % ci , - , ). cumulative incidence (ci) of acute gvhd in patients with two and more immunosupressants was , ( % ci , - , ), with one - , ( % ci , - , ), with no immunosupression - , ( % ci , - , ), p= , . ci of graft failure was: in patients with two and more immunosuppressive agents (n= ) - , ( % ci , - , ), with one (n= ) - , ( % ci , - , ), without immunosuppression (n= ) - , ( % ci , - , ), p= , . ci of trm in patients with two and more immunosuppressants was , ( % ci , - , ), with one - , ( % ci , - , ), without immunosuppression- , ( % ci , - , ), p= , . conclusions: we conclude that in our group of pid patients presence or absence of immunosuppression after hsct with tcrαβ/cd graft depletion and its extent made no significant difference on the incidence of acute gvhd, graft failure and trm. considering these results hsct with tcrαβ/cd graft depletion without posttransplant immunosupression could be recommended for patients with pid. disclosure background: lps-responsive beige-like anchor protein (lrba) deficiency is a severe primary immunodeficiency with a variable clinical phenotype, including features overlapping with common variable immunodeficiency, autoimmune lymphoproliferative syndrome, and immune dysregulation polyendocrinopathy enteropathy x-linked syndrome and an association with lymphoma. recent findings strongly support hsct in patients with severe presentation of lrba deficiency. however, there are no up-to-date follow-up and survival data of patients not undergoing transplantation beyond previous publications of large cohort studies. this study was conducted to increase the knowledge on transplant experience and to elucidate the clinical course of both transplanted and untransplanted patients with lrba deficiency under various targeted treatment modalities. methods: we performed an international inborn errors working party-and european society for immunodeficiencies-wide survey in to collect information about the hsct experience and the clinical course of both transplanted and untransplanted patients with lrba deficiency. we included an assessment of the disease activity and treatment responses with a specially developed immune deficiency and dysregulation activity (idda) score, which weighs the sum of organ involvement (scored by - ) by days of hospitalization and performance scores, taking into account the type of therapy and response per treatment phase, which we herewith introduce. data were obtained in accordance with the declaration of helsinki and an institutional review board review by means of retrospective chart review by local physicians. results: we analyzed the data of lrba deficient patients from centers, including unpublished patients. patients of our cohort underwent hsct between and . overall survival of patients undergoing transplantation (median fu months) was % ( / patients); all deaths were due to transplant-related mortality and occurred within months of hsct. . % ( / ) of untransplanted patients ( - years) are currently alive. out of the surviving transplanted patients (n= ) are in complete remission, in good partial remission (no treatment) and in partial remission (receiving therapy), implying that out of transplanted patients ( . %) are currently without treatment. only out of living untransplanted patients ( . %) do currently not receive therapy. most commonly used drugs in lrba deficiency were steroids ( . %), sirolimus ( . %) and abatacept ( . %). disease activity, measured by idda score, was significantly lower in patients treated with abatacept (p< . ). no immunosuppression-associated malignancy was detected in our cohort. analyzation of the idda score of all living patients revealed significantly lower disease activity in patients transplanted and alive, than in those who did not undergo hsct (p= . ). conclusions: in conclusion our findings do, concordant with previous results, support hsct, especially in patients with a severe phenotype of lrba deficiency. however, the question of hsct indication and optimal time point cannot yet be generally resolved. in terms of conventional treatment, abatacept is clearly favorable in untransplanted patients. nevertheless, it is not available in many countries and the long-term dependency on this treatment with its associated potential risks remains. finally, our newly introduced idda score for assessment of disease activity may also prove useful for other syndromes with immune dysregulation. disclosure: nothing to declare background: cytomegalovirus (cmv), is a major complication of allogeneic hematopoietic stem cell transplant (hct) recipients. to overcome morbidity associated with cmv infection, antivirals are given, and while they successfully suppress viremia, they do so in the absence of immune reconstitution. a different approach is to use a vaccine to bolster immunity to cmv early post-hsct and sustain it at least until the patient is immune reconstituted. we selected a highly attenuated, non-proliferating viral vector referred to as modified vaccinia ankara (mva) to insert immunodominant cmv antigens pp , ie -exon , and ie -exon . this novel vaccine (triplex) demonstrated excellent tolerability and immunogenicity in a clinical study of healthy adults (la rosa, et al. blood ) . these findings prompted us to conduct a test of vaccine protective efficacy in at-risk hct recipients in a multi-center, randomized and placebo-controlled phase clinical trial (nct ). methods: eligible patients were cmv-seropositive and undergoing hct for hematologic malignancies from matched related/unrelated donors without the use of exvivo or in-vivo t cell depletion. the randomization ( : ) was stratified by donor serostatus and hct center. patients were enrolled pre-hct, and on d were requalified for eligibility based on lack of prior cmv reactivation, ≥grade gvhd, high-dose steroids, or other significant transplantrelated morbidity. after randomization patients received triplex or placebo injections on d and d post-hct and followed for one year. the primary endpoint is defined as any cmv reactivation [≥ cmv genome copies (gc)/ ml], low-level reactivation prompting antiviral therapy, or cmv disease prior to d post-hct. based on the objective of reducing reactivation from % to % or from % to % on the vaccine arm, a sample size of was determined to provide at least % power at a one-sided . level of significance by log-rank test. results: the patient clinical/hct characteristics were balanced between the vaccine arm (n= ) and placebo arm (n= ). the vaccine was well tolerated with no significant difference in grade - aes or saes in both arms. triplex displayed potent immunogenicity, as many vaccine recipients with either cmv-positive or -negative donors showed strong reconstitution of both cd and cd cmv-specific immunity that initiated soon after the first injection and was elevated for at least d post-hct. most notably there was a reduction of primary endpoint-defining reactivation through d in the vaccine arm ( events, . %) compared to the placebo arm ( events, . %, p= . ). greater detail regarding protocol-specified secondary endpoints including time to viremia, its duration, frequency and duration of antiviral drug treatment, late cmv viremia, time to engraftment and incidence of acute and chronic gvhd, relapse and non-relapse mortality will be presented. conclusions: in summary, triplex was well tolerated in seropositive hct recipients, and was significantly better than placebo in preventing viremia cases accompanied by improved cmv-specific t cell reconstitution. background: due to the risk of invasive pneumococcal disease (ipd) after allogeneic transplant, the ecil guidelines recommend doses of pneumococcal conjugate vaccine (pcv) starting months after transplant, followed by either one polysaccharide -valent vaccine (psv ) dose or a th pcv in case of chronic gvhd. however, no data strongly support recommendations from the nd year. the aim of this study was to assess the anti-pneumococcal antibody (ab) concentrations, years after different schedules of vaccination. methods: patients who received allogeneic hct more than months ago and were seen in consultation between jan-nov were screened for igg ab concentrations for the pneumococcal serotypes shared by pcv , pcv , and psv ( , b, v, , c, f, f) , using modified enzyme linked immunosorbent assay (wernette et al. ) . patients were considered protected if their ab concentrations were > . μg/ml for the serotypes tested. from , all patients received pcvs during the st year of transplant, then either one dose of psv or (from ) a th pcv. for patients transplanted for more than months when pcv became available, we locally recommended > pcv and one psv . results: sixty-one patients were assessed at a median of . years (range: . - . ) after transplant. the mean age was years (range: - ), m/f sex ratio , . most patients had acute leukemia ( , . %) or lymphoproliferative diseases ( , . %). the donor was hla-identical in ( %) cases. conditioning regimen was myeloablative in ( %), reduced-intensity or non-ma in ( . %) each. none patient had experienced ipd since transplant nor had received immunoglobulins for at least months or rituximab for at least months at time of assessment. fifty-four patients ( . %) received consecutive pcv: ( . %) during the first months and ( %) thereafter. five ( %) patients received pcv and only one patient ( . %) received a unique dose of pcv. the st pcv was administered at a median of . (range: - ) months after hct. fifty-seven ( . %) patients received one psv at a median of months (range: . - ) and ( %) of them received a nd psv thereafter. finally, / ( %) patients received the recommended program. the overall protection rate was / ( . %) at a median time of . years after hct. no differences were observed between patients who received the recommended program ( / , %) and the others ( / , %). however, this latter group was often vaccinated > years after hct and therefore not protected for years, but may have developed a better response to only one or pcv. in all groups, the ab concentrations were heterogeneous from one serotype to an other. recipient age, donor age, underlying disease, conditioning, donor type, previous gvhd, lymphocyte count or gammaglobulin serum levels were not associated with seroprotection. conclusions: although the early antipneumococcal vaccination program is now well established, half of the patients vaccinated according to the guidelines were not protected anymore years later. prospective studies are needed to establish an optimal long-term programme. disclosure: nothing to declare , letermovir vs. placebo) and was well tolerated overall. we evaluated risk factors associated with development of cs-cmvi to further inform the clinical use of letermovir prophylaxis and future trial designs. methods: the participants without detectable cmv dna at randomization (primary efficacy population) were analyzed. cs-cmvi was defined as cmv viremia requiring antiviral preemptive therapy (pet) or cmv diseasepatients without cs-cmvi who died or withdrew from the trial on or before week were censored at the time of those events for this analysis. potential risk factors for cs-cmvi by week post hct were examined using univariate and multivariate cox proportional hazards models. candidate covariates were included in the multivariate cox model if they were associated with cs-cmvi in the univariate analysis or had been previously identified in the literature as significant risk factors. to avoid collinearity, the trial high-risk for reactivation of cmv stratification covariate was replaced with an updated covariate that only included patients who underwent hct with a mismatched donor, cord blood, ex-vivo t-cell depletion, or received alemtuzumab. the haploidentical hct and matched unrelated donor type were considered separately from the trial high-risk cmv categories. graft-versus-host disease (gvhd) and systemic glucocorticoid exposure were modeled as time-dependent covariates. race was dichotomized into asian and non-asian. the effect of study treatment was not included in the model due to nonproportional hazards. instead, letermovir treatment was used as a stratification variable. hazard ratios and % confidence intervals (ci) were calculated. results: there were cs-cmvi events ( . %) among patients by week post-hct for an incidence rate of . / patient-days ( % ci, . - . conclusions: donor cmv seronegativity, haploidentical hct, gvhd, glucocorticoid use, atg use, and asian race conferred significant risk of cs-cmvi through week post-hct in a phase study of letermovir prophylaxis. these results identify hct patient groups that would benefit the most from letermovir prophylaxis. methods: adult patients (age - ) with de novo aml in complete remission (cr) perusing t-cell replete haplo-hsct with atg+g-csf protocol were consecutively enrolled at peking university people's hospital between and . all patients were evaluated for donor-patient cmv serostatus before hsct. cmv dna emia were positive for > copies/ml cmv by pcr tests on peripheral blood (pb). refractory cmv infection was defined as cmv dnaemia lasting for > weeks in spite of treatment. pb samples of patients with refractory cmv infection ( relapse and without aml relapse) were sequencing for cmv derived small rna. cmv-mir-us - /ul- d were then chosen as target mirnas and tested by stem-loop taqman qpcr in consecutive patients (n= ). cumulative incidence of relapse (cir) and treatment related mortality (trm) were calculated using competing risks, cox model was tested with patient age, sex, wbc count at diagnosis, cr status, courses to achieving cr, cytogenetic risk group, minimal residual diseases before hsct, cmv infection, cmv-mir-us - / ul- d expression. us - /ul- d were further explored for the association with t cells and natural killer (nk) cells reconstitution and target mrna validation. results: in the total cohort of patients, the -year overall survival (os) and leukemia-free survival (lfs) were . % and . %, cir and trm were . % and . %. cmv infection occurred in . % recipients at a median time of weeks after haplo-hsct, and the duration was weeks. . % recipients experienced refractory cmv infection. cmv infection (yes vs. no, refractory vs no) did not affect cir in univariate analysis, while in multivariate analysis, cytogenetic risk group was identified as the only independent prognostic factor affecting cir (hr . , % ci, p= . ). in the cohort testing for cmv microrna, . % patients were identified expression of us - , . % patients for ul- d, while . % patients had co-expression. the relapse incidence was significantly lower in patients with vs. without co-expression of mir-us - /ul- d. co-expression of mir-us - /ul- d was the only independent risk factor for reducing cir (hr . , % ci, . - . ; p= . ). us - /ul- d as also found to promote reconstitution of nkg c+ nks. luciferase assay identify plzf as target of us - while klrc targeted by mir-ul- d, which suggest potential role of shifting nkg c/nkg a balance to nkg c domination. conclusions: the present study is the first prospective trial to evaluate cmv infection on relapse for aml patients following t-cell replete haplo-hsct with atg+ g-csf protocol. our results suggested cmv-mir-us - /ul- d co-expression rather than cmv infection reduces the relapse incidence, the stronger gvl effect might be associated with strengthening nkg c+ nk cells reconstitution and alloreactivity via plzf/klrc pathway. [[o image] . mir-us - /ul- d reduces relapse in aml after haplo-hsct] clinical trial registry: chictr-och- disclosure: nothing to declare o a randomised, placebo-controlled phase study to evaluate the efficacy and safety of asp , a first-inclass, dna-based vaccine in cmv-seropositive allogeneic haematopoietic cell transplant recipients adjudicated cmv-specific avt or adjudicated cmv eod and mortality through year post transplant. results: overall, patients were randomized, of whom received ≥ dose of randomized treatment (asp n= ; placebo n= ). there was no statistically significant difference in the proportion of patients who achieved the primary endpoint between the asp (n= [ . %]) and placebo (n= [ . %]) groups, respectively (odds ratio . ; % confidence interval p= . ). there were no statistically significant differences between groups for any secondary endpoints (table) . the incidence of teaes was similar between groups, except for a greater incidence of drugrelated teaes in the asp group (n= [ . %] ) compared with placebo (n= [ . %]) attributed to injection site-related teaes. mean t-cell response to pp increased over time in both groups and was significantly greater with placebo compared with asp (p= . ). there was no statistically significant difference between groups for mean gb-specific antibody response. conclusions: asp did not demonstrate efficacy in the reduction of overall mortality and cmv eod through year post transplant. asp demonstrated a similar safety profile to placebo, with the exception of injection siterelated teaes that were more frequent in the asp group. participants in this study will be followed up to . years post transplant for long-term safety assessments. disclosure: jm: personal fees and non-financial support from astellas, basilea, cidara, f g, gilead, merck and hsct) and autologous hsct (auto-hsct). during the observation period probable and proven rare ifd (eortc/msg criteria) cases were diagnosed in children and adults with hematological malignances and non-malignant hematological diseases after allo-hsct (n= ), auto-hsct (n= ), and chemotherapy (n= ). the median age was ( - ) y.o., males - %(n= ). the median follow up time for rare ifd cases was months; for survivors - months. results: incidence of rare ifd in hsct recipients was , %, it was higher after allo-hsct ( , %) than auto-hsct ( , %) (p< , ). in eight patients, this complication developed after ct and four of them proceed to allo-hsct. the most frequent underlying diseases were acute lymphoblastic leukemia ( %) and acute myeloid leukemia ( %). the median time of onset of rare ifd after allo-hsct was ( - ) days, auto-hsct - ( - ), after start of . etiology of rare ifd was identified by culture in % cases: rhizopus spp. - %, paecilomyces spp. - %, fuzarium spp. - %, malassezia furfur - %, trichosporon asahii - %, scedosporium apiosperium - %, scopulariopsis gracilis - %, rhizomucor pusillus - %, lichtheimia corymbifera - %, mix rare ifd with rhizopus spp. + paecilomyces spp. - %, paecilomyces spp. + fuzarium spp. - %. % cases (mucormycosis) were diagnosed by microscopy. in % cases rare ifd developed after or in combination with invasive aspergillosis, and patients had both preexisting invasive aspergillosis and co-infection with mucormycosis. the main site of infection were lungs ( %), the main clinical symptom -febrile fever ( %). antifungal therapy was used in all patients: lipid amphotericin b - %, lipid amphotericin b + caspofungin - , %, voriconazole - , %, posaconazole - , %, lipid amphotericin b + posaconazole - %, and echinocandins - , %. surgery was used in % patients. overall survival at weeks from the diagnosis of rare ifd was %. the -weeks overall survival was better in patients after ct and auto-hsct ( %) than allo-hsct ( %), p= , . conclusions: the incidence of rare ifd in hsct recipients was , % and depends on type of transplantation. rare ifd is a late complication after chemotherapy and hsct and usually develops after or in combination with invasive aspergillosis. higher incidence and worst prognosis rare ifd is observed in allo-hsct recipients. disclosure: nothing to declare o incidence and outcome of kaposi sarcoma after hsct: a retrospective analysis on behalf of idwp background: kaposi's sarcoma (ks) is an angioprolipherative disease which occurs in immunosuppressed patients, often associated with infection by human herpes . in solid organ transplantation (sot) ks is relatively common, the risk being - folds higher than that of general population and representing around % of secondary cancers. also hematopoietic stem cell transplantation (hsct) is a risk factor for the development of ks but until now only few case reports were published. we assessed retrospectively the incidence, the clinical characteristics, and the outcome of ks in the ebmt database. methods: the cases of ks were identified by ebmt registry (promise) and by inviting all ebmt centers to notify ks cases. the clinical features, type of therapy, survival rate and causes of death were retrieved from of promise or, if lacking, by specific case report form sent to participating centers. the center response rate was / ( %). results: fourteen centers reported patients with ks, all ks were diagnosed from to , but one case that was diagnosed in . the analysis was limited to / patients because ks was diagnosed before hsct in patients: they were patients, females and males who developed ks after allo hsct ( ) and auto hsct ( ); moreover, ks occurred after a second hsct in patients and after a third hsct in patient. the search of hhv in tumour tissue was done in cases and resulted positive in / . the underlying disease were: % leukemia, % lymphoma, % myeloma, % myelodysplastic syndrome (mds), % bone marrow failure. the source of stem cells was bone marrow in patients ( %) and peripheral blood in patients ( %). the median age at ks diagnosis was . years (range . - . ) . considering the number of hsct performed in the participating center from to , the incidence rate was . % in allogeneic transplantations ( / ), . % in autologous transplantations ( / ) and . % ( / ) in the whole group. the interval of time between hsct and the development of ks was months (range - . - . ) . the organ involvement was: % skin ( patients), % lymph nodes ( ), % gingival ( ) . apart from withdrawal of immunosuppression, patients received chemotherapy, patients received radiotherapy, and patient received radio and chemotherapy; moreover patients received antiviral treatment with ganciclovir or foscarnet. eight patients ( %) are alive whereas patients ( %) died at a median time of . months, range . - . . the causes of death were infection in cases, secondary malignancy/ptld in and relapse/progression in whereas no case of death directly associated to ks. conclusions: ks is a rare complication of the immunosuppressive status related to hsct that generally occurs within the first year after hsct. the low prevalence and the rarity of this complication do not justify the adoption of screening program for hhv- . on the other hand, the role of the virus in febrile status in immunosuppressed patients and the risk factors for the development of ks are not well known. disclosure background: autologous stem cell transplantation (auto-sct) is considered the standard treatment for patients with relapsed or refractory (r/r) hodgkin lymphoma (hl). for those with high-risk disease, an alternative consolidation strategy with allogeneic sct (allo-sct) could be a potential option to improve the outcome. however, allo-sct with a reduced-intensity conditioning (ric) needs around months for the graft-versus-lymphoma effect (gvl) to develop, thus in patients with an aggressive hl the disease might progress before this happens. in this setting, a tandem auto-ric-sct approach has the potential of combining cytoreduction to keep the lymphoma under control and the potential benefit of a gvl effect. to better understand the safety and efficacy of a tandem auto-ric-sct approach we conducted a retrospective analysis of patients treated with this strategy between january and december and reported to the ebmt registry. methods: patients were included if they had received an auto-sct followed by a planned ric-sct in < months with no disease relapse between the procedures. the primary endpoint was progression-free survival (pfs) after the tandem procedure. secondary endpoints were overall survival (os), cumulative incidence of non-relapse mortality (nrm), incidence of relapse (ir) and graft versus host disease (gvhd). results: one-hundred and thirty patients [ % male, median age at auto-sct, years (range: - )] fulfilled the inclusion criteria. the median time between diagnosis and auto-sct was months (range: - ) and the median number of lines prior to auto-sct ( ) ( ) ( ) . disease status at auto-sct was complete response in %, partial response in % and the remaining % were transplanted with active disease. the median time from auto to allo-sct was months ( - ). forty percent underwent an identical sibling allo-sct, % unrelated and % haplo. tbi was used in % of the patients as a part of ric. gvhd prophylaxis was cyclosporine-methotrexate in % of the patients, cyclosporine-micofenolate mofetil in % and post-transplant cyclophosphamide in the remaining %. % of the patients engrafted after ric-sct. after a median follow-up of months ( - ), % of the patients background: allogeneic stem cell transplantation (allo-sct) is a valid option in patients with refractory/relapsed lymphoma but gvhd remains the major cause of mortality and morbidity. calcineurin-inhibitors (cni) combined with methotrexate or atg is the conventional strategy for preventing gvhd, resulting in an incidence of cgvhd of - %. post-transplant cyclophosphamide (pt-cy) reduces the risk of severe cgvhd and improves survival in acute leukemia patients receiving allo-sct from matched sibling (msd) or unrelated donors (ud). the aim of this retrospective registry-based study was to compare pt-cy-based gvhd prophylaxis to standard prophylaxis in the setting of msd or ud for patients with lymphoma. methods: three thousand eight hundred sixty-four lymphoma patients undergoing an hla -id sct(hla identical stem cell transplantation) registered in promise were included in the study (table ) . outcomes between pt-cy vs no-pt-cy were compared a) with a multivariate cause-specific cox model adjusted on ric/mac, donor type, source of stem cell, age of the patient, donor gender, patient gender, diagnosis and, disease status at sct; and b) by matching one pt-cy patient ( ) with two no-pt-cy patients ( ) using the same covariates. results: in univariate analysis, comparing pt-cy and no-pt-cy, the -day ci of grade - agvhd was % and % (p= . ), the -year ci of non-relapse mortality (nrm) was % and % (p= . ) and the -year relapse incidence (ri) was % and % (p= . ), respectively. the -year ci of cgvhd was % vs % (p= . ), -year pfs for pt-cy and no-pt-cy was % and % (p= . ) and % and % (p= . ) , respectively. in multivariate analysis, prophylaxis with pt-cy was not associated with a reduced risk of agvhd, overall or extensive cgvhd, nrm, ri, nor with an improved pfs or os. likewise, in the matched-pair analysis pt-cy did not impact on any of these outcomes. conclusions: this study demonstrates that in lymphoma patients who underwent an hla-id sct, gvhd prohphylaxis strategies employing pt-cy-based achieve equivalent transplant outcomes to those seen with cni-based strategies. myeloablative conditioning may contribute to disease control after stem cell transplantation in blastic plasmacytoid dentric cell neoplasia background: blastic plasmacytoid dentric cell neoplasia (bpdcn) is a rare and clinically aggressive hematopoietic malignancy, which preferentially involves the skin, the bone marrow, and, occasionally, the lymph nodes. it mainly affects elderly patients and has a poor prognosis with conventional chemotherapy. the treatment for this condition is heterogeneous, some patients treated with lymphoma-type regimens and others receiving intensive acute leukemiatype chemotherapy. although preliminary case series suggest that hematopoietic stem cell transplantation (sct) could provide sustained disease control in patients with bpdcn, the role of sct and potential graft-versus-tumor effects (gvt) in this condition is yet undefined. methods: between and patients were included in an ebmt prospective non-interventional study (nis) on the value of sct in bpdcn. these were compared with patients with bpdcn registered with the ebmt during the same time period outside the nis. no differences with regard to overall survival (os), line of treatment or year of transplant was observed between patients included in the nis and the remainder from the ebmt database, and they were therefore analyzed together. results: one hundred and forty-two patients were treated with an allogeneic sct (allosct) and patients with autologous or syngeneic sct (autosyn). the median follow-up was months with no differences between the allosct and autosyn groups. disease status at sct was complete remission (cr) in %, and % patients received the sct as part of first-line treatment. two-year os after autosyn was % ( % ci - %), whereas it was % ( % ci - %) following allosct. of those patients who received an allosct, % were transplanted from an unrelated donor (ud), and % from an identical sibling donor (sib). reduced intensity conditioning (ric) was used in % and myeloablative conditioning (mac) in % of the patients. in allotransplanted patients, multivariate competitive risk analysis of nrm and relapse incidence considering age, conditioning intensity, line of treatment, remission status prior to allosct and donor type revealed a statistically significant reduction of relapse incidence (p= . , hr . ci . - . ) without increased nrm for patients who received mac compared to ric. multivariate cox regression analysis for os considering the same co-variates confirmed that mac was associated with a significantly reduced mortality risk (p= . , ci . , hr . - . ) along with being in cr at allosct (p< . , hr . , . conclusions: this study confirms on a large data set that sct is an effective and potentially curative treatment for patients with bpdcn. the superiority of mac and the efficacy of autosyn suggest that apart from gvt, highdose therapy might be an important contributor to long-term disease control in this condition. clinical background: allogeneic hematopoetic cell transplantation (allo-hct) is a curative therapy for patients with relapsed/ refractory and high-risk non-hodgkin lymphoma (nhl). however, no large studies have evaluated the trends in the utilization of allo-hct in elderly nhl patients (≥ years). using the cibmtr registry we report here a time trends analysis of allo-hct use in elderly nhl subjects methods: we identified nhl patients (≥ years of age) undergoing a first allohct during - in the united states (u.s.). study cohort was divided into the following time-periods for analysis; - vs. - vs. - . primary outcome was overall survival (os). secondary outcomes included progression-free survival (pfs), relapse/progression (r/p) and non-relapse mortality (nrm). results: baseline patient characteristics are shown in table- . during the three study time-periods median patient age ( - -year), use of reduced-intensity conditioning regimens and proportion of patients with chemosensitive disease remained stable. in the most recent era ( - ) a higher proportion of patients had t-cell nhl, history of prior autografts, good performance status (kps - ) and high hct-ci, while fewer subjects received a hlamatched sibling hct. the cumulative incidence of day grade - acute graft-versus-host disease (gvhd) for - vs. - vs. - cohorts was % vs. % vs. % respectively (p= . ). the cumulative incidence of chronic gvhd at year was % vs. % vs. %, in similar order (p= . ). the -year probabilities of nrm and r/p of - vs. - vs. - time-periods were % vs. % vs. % (p= . ) and % vs. % vs. % (p= . ), respectively (figure) . the -year probabilities of pfs and os ( - vs. - vs. - showed significantly improved outcomes in the most recent time-periods as following: % vs. % vs. % (p= . ) and % vs. % vs. % (p< . ), respectively (figure) . on multivariate analysis, compared to the - cohort, the - cohort showed a % reduction in the risk of mortality (rr= . , %ci= . - . , p= . ). the most common cause of death was relapse of primary disease in all time-periods. conclusions: utilization of allo-hct has steadily increased in elderly nhl patients in the u.s. since . in the recent years despite decline in the use of hlamatched sibling donors and transplanting elderly patients with higher hct-ci and more heavily pretreated disease, survival outcomes have improved. age alone should not be a determinant for allo-hct eligibility in nhl. methods: four medical experts who had managed patients with dlbcl using different car t-cell therapy protocols and products independently reviewed the extracted adverse event data from the case report forms and re-graded crs using the more commonly used lee scale (lee, blood, ) and the nt grading for encephalopathy (modified car t related encephalopathy syndrome (mcres) (neelapu, nature reviews in clinical oncology, ) while blinded to the original trial grading and other experts' grading. re-grading assessments and disagreements concerning the assigned lee and nci ctc grades were later discussed and reconciled among reviewers during a live meeting. as per the investigational charter, in cases that could not be reconciled, the most conservative final assessment of any reviewer determined the final grading for any individual case. results: crs: of ( %) patients were originally recorded as crs by penn scale, and were regraded as crs by lee criteria. of subjects received anticytokine therapy overall. with this blinded reassessment, more patients were categorized as grade (lee vs penn: vs ), fewer patients as grades and ( vs and vs , respectively) and the same number of patients as grade ( vs ). nt: results were compared with data on nt determined by nci ctc criteria of tisagenlecleucel, in which nt was broadly defined as the occurrence of any nervous system or psychiatric ae (eg, anxiety, dizziness, headache, peripheral neuropathy, and sleep disorder). of subjects were identified as having nt by ctc criteria: ( . %) patients were identified as having grade / nt, ( . %) patients as having grade nt,& ( . %) patients as having grade nt. evalution by mcres grading system revealed low rates of encephalopathy/delirium: ( . %) patients had grade / nt, ( . %) patients had grade nt, and ( . %) patients had grade nt. no grade events were seen and the presence of crs was associated with higher likelihood of concomitant nt. conclusions: these results demonstrate difficulties in identifying cross protocol toxicity assessments. harmonized grading scales being developed for future studies will facilitate comparison of the safety profiles of different car t-cell and other immune effector cell products. further analyses are ongoing of these data with the new asbmt consensus crs guidelines. recent studies show that mouse bone marrow tregs localize in the hematopoietic stem cell (hsc) niche, where they contribute to hscs maintenance and promote donor engraftment and b cell lymphopoiesis. we are investigating if human tregs promote b cell reconstitution and immunity in preclinical models and in haplo-hct patients. methods: b cell reconstitution was analysed monthly by facs in bone marrow (bm) and peripheral blood (pb) samples from patients who underwent either treg/tcon haplo-hct ( patients), or t-cell depleted haplo-hct ( patients) or haplo-hct with high dose post-transplant cy (ptcy, patients). diagnosis was acute leukemia in patients, lymphoma in and multiple myeloma in . pb total immunoglobulin (ig) and anti-cytomegalovirus (cmv) igm were also monitored together with cmv viremia. for the mouse model, donor derived human treg and cd + hematopoietic stem cells (hscs) were coinfused in sublethally irradiated ( gy) immune-deficient nsg mice and donor engraftment and b cell reconstitution were analysed in mouse pb twice a month by facs. results: b cell reconstitution was faster after treg/tcon haplo-hct when compared to other haplo-hct protocols. b cell counts were higher in pb of patients that received treg/ tcon haplo-hct (p = . ) and were comparable to those of healthy subjects by months after transplant ( ± cells/ mm , fig. a ). we could detect early frequencies of cd + cd + cd + cd + common lymphoid progenitors, cd + cd + cd + cd -pre/pro-b, cd + cd + cd + cd + pre-b, and pro-b cells in the bm of these patients, that resulted in an increased production of cd + cd + cd -igm + immature b cells, cd + cd + cd + igm + transitional b cells and cd + cd + mature b cells. we used a mouse model of xenotransplantation to understand whether donor b cell reconstitution in treg/ tcon haplo-hct is boosted by a treg-mediated effect on donor human hscs. we found that infusion of human tregs facilitated donor hsc engraftment. hsc-derived mature b cells were rapidly abundant and easily detectable days after hsc infusion in pb of treg-treated animals. to evaluate donor b cell function we analysed ig production in response to cmv reactivation in transplanted patients. post-transplant hypogammaglobulinemia was rapidly corrected in treg/tcon haplo-hct patients. total igm were higher compared to other haplo-hct protocols and reached normal levels by months after transplant ( ± mg/dl, fig. b ). cmv reactivation rate was similar among haplo-hct protocols (~ %), but it occurred later after treg/tcon immunotherapy ( +/- days vs +/- days). new production of anti-cmv specific igm was documented in % of cmv seropositive patients +/- days after treg/tcon haplo-hct, while anti-cmv specific igm were undetectable after other haplo-hct protocols within the first months after transplant. [ background: allogeneic cell transplantation (hsct) success prediction is partly based on minimal residual disease (mrd) and hematopoietic chimerism testing. we developed a droplet digital pcr platform (dpcr) for the simultaneous detection of mrd and hematopoietic chimerism. methods: a panel of deletion/insertion polymorphic markers and frequent molecular targets used for mrd testing: npm , runx -runx t (t ; ), dnmt a, mll-ptd, cbfß-myh (inv ), kras, mll-af , idh / , ckit, bcr-abl (t ; ), evi and wt expression were included in a single dpcr platform for mrd and hematopoietic chimerism analysis. a total of patients were evaluated with a mean follow-up of days (range: - days). results: hematopoietic chimerism analysis revealed mixed chimerism (mc) in patients ( % of all patients). mc was detected more frequently in patients with reduced intensity conditioning ( %) when compared with full conditioning ( %). the mean percentage of host derived dna in peripheral blood was % (range: . - %). three different patterns of mc were observed: increasing, decreasing or stable mc. in those patients with stable or decreasing mc (n= ), as well as patients with complete donor chimerism (n= ) the molecular targets used for mrd monitoring were not detectable. in out of patients, increasing mc was detected. this group of patient showed in addition either a positive mrd marker, increased wt expression or both. in patients with increasing mc, a positive mrd marker and increased wt expression hematologic relapse of the underlying disease was observed. in patients with increasing mc and a positive mrd marker, we analysed whether mc or the molecular target for mrd was first detected. in % of the cases mc was detected before the molecular marker used for mrd assessment, while in % of the patients mrd positivity was detected before mc. in the remaining patients ( % of the patients) mc and mrd positivity were detected simultaneously. the mean time between either mc or mrd detection in peripheral blood and relapse was days (range: - days). patients with increasing mc and mrd positivity, whether or not they responded to treatment, showed a similar kinetic pattern for the chimerism and mrd markers. in those patients that responded to molecular relapse treatment (n= ) the mean time to achieve complete donor chimerism or mrd pcr negativity was days and days respectively. conclusions: the combination of mrd and chimerism markers in a dcpr platform represents a sensitive and accurate diagnostic tool for the comprehensive assessment of the molecular remission status after hsct. in addition, by using the developed dpcr platform costs and turnaround times can be reduced. disclosure background: minimal residual disease (mrd) monitoring can help to indicate impending relapse of acute leukemia after allogeneic hematopoietic stem cell transplantation (allo-hsct). because impending relapse can be altered with early detection of low-volume disease and timely therapies, preemptive intervention is a reasonable option for patients with mrd which can spare those in remission from further therapy. chemotherapy plus donor leukocyte infusion (chemo-dli) is the most important preemptive intervention, but it may lead to several complications, such as severe graft-versus-host disease (gvhd) and pancytopenia. hypomethylating agents (hmas) represent another potential preemptive intervention, but it only delayed the time to hematologic relapse and the long-term efficacy may be unsatisfactory. thus far, few studies had identified the longterm efficacy of preemptive intervention with drugs in patients with mrd after allo-hsct.two prospective studies (nct and nct ) reported that preemptive interferon-α (ifn-α) treatment can help clear minimal residual disease (mrd) and prevent relapse after allogeneic hematopoietic stem cell transplantation (allo-hsct). in this extension study, we aimed to identify the long-term clinical outcomes of preemptive ifn-α treatment in acute leukemia patients who were mrd positive after allo-hsct (n= ). methods: mrd was monitored by multiparameter flow cytometry (mfc) and taqman-based reverse transcriptionreal time polymerase chain reaction (pcr). a patient was considered to be mrd-positive when a single bone marrow sample tested positive by pcr or mfc. recombinant human ifn-α- b injections were administered subcutaneously for cycles (twice or thrice weekly in every -week cycle). results: the -year cumulative incidence of total chronic graft-versus-host disease (cgvhd) and severe cgvhd after ifn-α treatment was . % ( % ci, . - . %) and . % ( % ci, . - . %), respectively. the -year cumulative incidence of relapse and non-relapse mortality (nrm) after ifn-α treatment was . % ( % ci, . - . %) and . % ( % ci, . - . %), respectively. the year probabilities of disease-free survival and overall survival (os) after ifn-α treatment were . % ( % ci, . - . %) and . % ( % ci, . - . %). in multivariate analysis, severe acute gvhd was associated with a higher risk of nrm and poorer os, and mild to moderate cgvhd was associated with a lower risk of relapse and better survival. conclusions: these data confirmed that preemptive ifnα treatment showed long-term efficacy in patients who were mrd-positive after allo-hsct. clinical trial registry: the study was registered at http://clinicaltrials.gov as #nct and #nct . disclosure: the authors declare no competing financial interests. a phase ii clinical trial of leuprolide for enhancement of immune reconstitution after ex vivo cd + cell-selected allogeneic hct with tbi-based conditioning gnrh agonist use, which has been associated with thymic cellular degeneration in mouse (velardi, jem ). as direct gnrh antagonism might circumvent this effect, a follow-up phase ii trial evaluating peri-transplant degarelix for enhancement of immune reconstitution is in progress. background: in allogeneic hematopoietic stem cell transplantation (allo-hsct) recipients, cytomegalovirus (cmv) reactivation and disease are frequent causes of morbidity and mortality, that may be evaded by cmv-specific t cell reconstitution. methods: we designed a prospective, single-center observational study to assess if the kinetic and quality of cmv specific t-cell reconstitution impact the incidence and severity of cmv reactivations. we report data on the first consecutive patients affected by hematological malignancies receiving allo-hsct followed by cyclophosphamide and rapamycin between december and august . patients received allo-hsct from family (sib-lings= ; hla haploidentical= ), unrelated hla matched (n= ) donors or cord blood (n= ). the cmv serostatus of host (h) and donor (d) pairs was: h + /d + (n= , %), h + /d -(n= , %) and h -/d + (n= , %); h -/dwere excluded. cmv dnaemia was assessed weekly in whole blood (wb). absolute numbers of polyclonal and cmv-specific t cells were quantified by flow cytometry using trou-count™ tubes (bd) and dextramer® cmv-kit (immu-dex), respectively, in the graft and fresh wb at days - , + , + , + , + , + , + and + . dextramers permit the identification of cmv-specific lymphocytes restricted for several hla class i molecules: a* : / * : /* : /* : and b* : /* : /* : . these alleles allowed the longitudinal evaluation of ( %) patients. results: at a median follow-up of days post-hsct, ( %) patients experienced a cmv-related clinically relevant event (cre, median + days), including patients ( %) with cmv disease. for each time-point, we compared the absolute number of cmv-specific lymphocytes in patients experiencing or not a subsequent cre. at + days, we observed lower cmv-specific cd + t cells in patients prone to reactivate cmv than in not reactivating patients (median cmv-specific cd + cells/ ml= . vs . , p= . ). furthermore, patients with any dextramer positivity at + days displayed a lower incidence of cre compared with subjects who were negative (cre probability: . vs . , p= . ). patient stratification based on different thresholds of dextramerpositive cells confirms the inverse association between cre and cmv-specific immunity (cre incidence in patients with: cells/ml= . , < cells/ml= . , ≥ cells/ml= . ; p= . ). we observed a higher cre incidence in cmv h + /dpairs than in h + /d + ( . vs . , p= . ). taking advantage of the hla mismatched-hsct setting, we then dissected cmv-specific t-cell response according to hla restriction elements (h/d=shared n= , drestricted n= , h-restricted n= ). in h + /d + pairs, we observed a fast and similar kinetic of reconstitution of cmv-specific lymphocytes restricted by h/d and d hlas. conversely, in h + /dpairs, we detected only cmv-specific cd + lymphocytes restricted for h/d haplotypes. hostrestricted cells remained undetectable for the first days after hsct. conclusions: when the donor is cmv seropositive, a rapid and effective reconstitution of cmv-specific d-and h/d-restricted memory t cells occurs. if the donor is cmv seronegative, only h/d-restricted lymphocytes are observed early after allo-hsct in h + /dpairs. these findings indicate that cmv reactivation can prime h/d-restricted t cells presumably educated in the donor thymus; conversely, d-and h-restricted donor-derived lymphocytes have not yet undergone neither cross-priming nor thymic education, which might be required for full protection from cmv. disclosure: c.b. received research support from molmed s.p.a. and intellia therapeutics. l.b. is employed by immudex aps. non of the other authors has any relevant conflict of interest to disclose. o -azactidine is safe and effective therapy for prevention of disease relapse in high-risk patients with acute myeloid leukemia and myelodysplastic syndrome following allogeneic stem-cell transplantation ivetta danylesko , noga shem-tov , adriana del-giglio , ronit yerushalmi , arnon nagler , avichai shimoni background: allogeneic stem-cell transplantation (sct) is curative approach in patients with aml or mds. however, disease recurrence is the major cause of treatment failure. azacitidine has been used in standard treatment of mds and also in aml patients not eligible for standard chemotherapy. there is limited data on the safety and efficacy of azacitidine after sct. we explored the use of low-dose azacitidine for prevention and treatment of relapse of aml/ mds after sct. methods: patients in cr after sct who were considered to be at high-risk for relapse were given azacitidine at mg/m for days every days, planned for years. patients with overt relapse after sct were given - mg/ m for days until progression. results: ninety-four patients, median age years ( - ) were given azacitidine after sct from hla-matched sibling (n= ), matched-unrelated (n= ) or haploidentical (n= ) donors. diagnosis was aml (n= ) or mds (n= ). the conditioning regimen was myeloablative (n= ) or reduced-intensity (n= ). patients were given prophylactic azacitidine; were in cr after sct but at high-risk for relapse due to active disease (n= ) or positive minimal-residual disease (mrd) (n= ) prior to sct, or poor-risk cytogenetics (n= ). seven patients were given azacitidine as secondary prevention after achieving cr with chemotherapy for post-transplant relapse. patients were given azacitidine pre-emptively for mixed-chimerism, positive mrd after sct or early relapse (< % marrow blasts). patients in the combined prophylactic/ preemptive group (n= ) started azacitidine in a median of . months ( . - . ) after sct and received a median of courses . patients were also given donor-lymphocyte infusions (dli) concomitantly with azacitidine. are still on therapy, died or progressed, stopped after long remission, stopped due to patient request and only discontinued due to toxicity. of patients given prophylactic azacitidine remained in cr. of patients given azacitidine preemptively achieved cr and remained in cr. with median follow-up of months , of the patients in the prophylactic/ pre-emptive group are alive and died. the estimated -year os and pfs are % ( %ci, - ) and % ( %ci, - ), respectively. the expected pfs in this group of high-risk patients is less than %. patients were given azacitidine for overt relapse after sct. patients were given a median of courses ( - ) and were also given dli. patients achieved cr, stable disease and progressive disease. patients are still on therapy, died or progressed and had to discontinue due to hematological toxicity. with median follow-up of months , are alive and are progression-free. and % ( %ci, - ), respectively. conclusions: azacitidine is safe and effective therapy when used prophylactically in high-risk aml/mds patients to prevent relapse or preemptively to treat mrd or early relapse after sct. azacitidine maintenance may improve outcome in this high-risk patient group and should be further explored. results of azacitidine treatment in overt post-transplant relapse are limited. disclosure: nothing to declare novel mass cytometry analysis identifies reciprocal changes in nkreg and cd em as the dominant early immune reconstitution signature associated with subsequent acute gvhd after ric-ahst background: treatment failure after allogeneic haematopoietic stem-cell transplantation (ahst) using reducedintensity conditioning (ric) results from either too much alloreactivity and harmful acute graft-versus-host disease (agvhd) or not enough (reducing graft-versus-tumour effects). studies have identified individual reconstituting immune cell subsets associated with development of clinical alloreactivity but the functionally dominant parameters remain unknown. we therefore used mass cytometry (ms) technology to determine multiple parameters simultaneously to identify dominant cellular immune reconstitution signatures associated with development of clinical alloreactivity after ahst. methods: phenotypic markers identifying > t, b and nk cell subsets known to influence alloreactivity were combined in a single ms panel. peripheral blood from patients with haematological cancer was analysed at d+ after t-replete hla-matched ric-ahst using uniform conditioning. test samples were spiked with cd barcoded healthy control cells. three complementary high-dimensional analytic tools were used to identify immune signatures in cd + lineage + cells across the whole cohort and identify differences between patients grouped by subsequent occurrence of agvhd. results: significant batch effects were effectively reduced with a novel r-based algorithm normalising data to control cells. unsupervised clustering analysis using phenograph and flowsom algorithms identified and phenotypically distinct clusters respectively. diversity analysis demonstrated lower cluster diversity (p= . ) in the patients who subsequently developed agvhd consistent with perturbation of phenotypic clusters in these patients. comparison of individual cluster abundance identified cluster groups significantly different between patients who subsequently developed agvhd and those who did not. a cluster containing cells with a cd bright cd neg cd +/regulatory/tolerant nk cell (nkreg) phenotype was significantly reduced in patients who subsequently developed agvhd using both phenograph and flowsom algorithms (p< . ). a differentiating cell population with this phenotype was also identified in forward analysis using the citrus algorithm. notably, this reduction in nkreg was accompanied by a significant increase in abundance of a cluster of ccr + cd ra -ccr -cd effector memory t-cells (tem) in patients who subsequently developed agvhd. there was a significant negative correlation (p= . ) between nkreg and tem. in contrast there was no inverse correlation between cd regulatory t-cells and cd em. these changes were independent of clinically significant cmv reactivation. finally, we determined the impact of time to agvhd on this novel immune signature. reciprocal changes in nkreg and cd tem abundance were more significant in patients who developed agvhd before d , consistent with a dynamically evolving immune signature. conclusions: we show proof-of-concept that a novel pipeline can be applied to ms data to measure multiple immune reconstitution parameters after ahst. importantly, this pipeline identified concomitant loss of nkreg and increase of cd tem in patients who subsequently developed agvhd. this is consistent with loss of nk cell-mediated control of alloreactive cd tem cells as the dominant immune process preceding the development of agvhd after ric-ahst. our data provide mechanistic insight into evolution of alloreactivity and support the development of strategies to maintain or expand nkreg numbers early post-transplant to reduce harmful agvhd. [[o image] . dominant immune reconstitution signature early after ric-ahst associated with subsequent acute gvhd] multiple myeloma o abstract already published. impact of high-risk cytogenetics in newly diagnosed multiple myeloma undergoing upfront stem cell transplantation: a study from the ebmt chronic malignancies working party background: current consensus identifies t( ; ), t( ; ), t( ; ), gain and/or deletion in chromosome , and del( / p) as high-risk cytogenetics in newly diagnosed multiple myeloma (ndmm). however, evidence on outcome of specific abnormalities after transplantation as first-line treatment is limited. we analyzed high-risk ndmm patients reported to the european society for blood and marrow transplantation (ebmt) registry undergoing upfront stem cell transplantation. methods: upfront transplantation was defined as first autologous transplant within months from mm diagnosis. survival and cumulative incidence were calculated from date of first transplant ( % confidence interval). end points were progression-free survival (pfs), overall survival (os), relapse and non-relapse mortality (nrm). cox model with hazard ratios (hr) was used for multivariable os analyses and cumulative incidence method for relapse incidence and nrm. results: within the ebmt registry, high-risk ndmm patients according to cytogenetics underwent single autologous (n= ), tandem autologous (n= ), autologous-allogeneic (n= ) stem cell transplantation between and . the median follow-up of all patients was months ( % ci, - months), the median age was years (range, - years) and the median time between diagnosis and transplantation was . months (range, . - . months). frequencies according to cytogenetic were: del( ) (n= , %), t ( ; ) (n= , %), gain or deletion in chromosome (n= , %), t( ; ) (n= , %). two or more cytogenetic abnormalities were documented in patients ( %). % of patients were male and most patients had igg ( %) or iga ( %) paraproteins. a karnofsky performance status < % had % of patients while frequencies according to international staging system (iss) i/ii/iii were %/ %/ %. complete remission (cr) at time of transplantation was achieved by %. in univariable analysis, presence vs absence of del ( ) conclusions: in ndmm patients with at least one highrisk cytogenetic abnormality undergoing upfront transplantation, outcome was similar between del( ) and t( ; ) while the presence of two or more high-risk cytogenetic abnormalities showed significantly worse os compared with only one high-risk abnormality. disclosure: nothing to declare. the role of renal impairment at diagnosis in multiple myeloma undergoing autologous transplantation. a retrospective analysis of the cmwp background: renal impairment (ri) is frequent in newly diagnosed myeloma patients and is considered to be a risk factor for worse overall survival. with active myeloma therapy renal function often improves or even normalises. however, it is unclear whether renal impairment at diagnosis is a persisting biological risk factor or rather a potentially reversible organ complication. methods: from the ebmt calm study database all myeloma patients having received a first autologous transplant between and with information on renal function both at diagnosis and at transplant were extracted. renal function was classified according to the calculated glomerular filtration rate (gfr) rate as normal ("normal", gfr > ml/min), moderately impaired ("moderate", gfr - ml/min) or severely impaired ("severe", gfr < ml/min). categorial variables were tested by chi-square test. os was determined from transplantation and calculated by kaplan meier with logrank testing. results: patients fulfilled the selection criteria and were included. at diagnosis, patients had normal, moderate and severe ri. median age at diagnosis was , and years in the ri subgroups. genetic information was available in only a subgroup of patients. t( ; ) was present in %, % and % respectively and del was found in %, % and %. bortezomib-based induction therapy was given in %, % and % of cases (p= . ). os differed significantly between the ri groups with a median of , and months, respectively (p< . , fig ) . in contrast renal function at transplant had no impact on os with a median of months (no ri at transplant), months (moderate ri at transplant) and not reached (severe ri at transplant). most of the patients with severe ri at diagnosis had improved their renal function by the time of transplantation. however, this did not positively impact on os: patients with no ri at transplant had a median os of months (n= ), while it was months for moderate ri (n= ) and not reached in patients transplanted with persisting severe ri (n= ,p< . ). conclusions: from this large analysis including almost myeloma patients renal impairment at diagnosis has been found to be a risk factor for os, while renal function at transplant did not impact on post transplant survival. these findings support the safety and efficacy of autologous transplantation in patients with severe ri at transplant. on the other hand improving renal function between diagnosis and transplant does not seem to improve prognosis.. our analysis supports the notion that ri at diagnosis appears to be a surrogate parameter for a more aggressive disease course. disclosure: nothing to declare fig. oerall survival according to ri at diagnosis] analysis of outcomes in patients with myeloma who had a second allohct either for disease relapse or graft failure: an ebmt cmwp study background: the options for patients with myeloma (mm) who relapse or develop graft failure after an allosct are limited. a second allohct is occasionally feasible though is high-risk. we performed a retrospective analysis to assess outcomes in this cohort. methods: data on patients with mm who underwent a second allohct at ebmt centres between and were obtained from the ebmt registry. results: a total of patients ( m, f) with mm ( % igg, % iga, % lc) underwent a second allosct. the median (range) age at the first allohct was . ( . - . ) years and . % were >/=pr. when comparing the indications for the nd allohct -relapsed mm ( %) or graft failure ( %) -patients with graft failure were significantly more likely to have received a mismatched (related/unrelated) or unrelated donor for the st allosct ( % vs. %) (p= . ), were more commonly female ( % vs. %) (p= . ) and were more likely to have had a ric as opposed to a mac allosct ( % vs. %) (p= . ). the median (range) interval between the st and nd allohcts was . ( . - . ) months in cases of graft failure and . ( . - . ) months for those who had relapsed. at a median ( % ci) follow-up of . ( . - . ) months following the second allohct, overall survival (os) was % ( - %) at two years and % ( - %) at five years. there was no difference in os at five years based on the indication for allohct: % ( - %) for relapse and % ( - %) for graft failure (p= . ) (figure ). neutrophil engraftment following the nd allohct was achieved by day + in % ( - %) and % ( - %) of the relapse and graft failure patients, respectively. the cumulative incidence of agvhd (ii-iv) and cgvhd (at five years) following the nd allohct was % ( - %) and % ( - %), respectively.relapse-free survival was % ( - %) at years, % ( - %) in those transplanted for disease relapse and % ( - %) in those transplanted for graft failure (p= . ). the cumulative incidence of relapse and nrm at five years was % ( - %) and % ( - %), respectively. the five-year os following the second allosct was % ( - %) for those relapsing between and months after the first allohct and % ( - %) for those relapsing later than months (p= . ).on univariate analysis, os at five years was superior in patients who had had hla identical sibling donors as opposed to other donor sources: % ( - %) vs. % ( - %) (p< . ). on multi-variate analysis, donor source (hla identical sibling vs. other) remained a predictive factor for os (p= . ). conclusions: in this high-risk mm cohort, one quarter of patients remained alive five years after the nd allohct with similar outcomes seen following disease relapse and graft failure. however, the relapse-free survival rate was low in those transplanted for relapsed mm. later relapses after the first allohct appear to fare better and the best outcomes are seen using matched sibling donors. a nd allohct therefore remains an option to be considered for selected mm patients. disclosure: nothing to declare minimal residual disease (mrd) ratio before and after autologous stem cells transplantation (asct) in multiple myeloma (mm) riccardo boncompagni , michela staderini , chiara nozzoli , elisabetta antonioli , barbara accogli , riccardo saccardi careggi university hospital, florence, italy, background: in the last ten years, multiparametric flow cytometry (mfc) has been standardized and routinely applied for the detection of mrd as a prognostic factor in mm patients across different lines of therapy. we assessed the mrd carried out before and after asct in a series of consecutive mm patients in order to investigate whether the ratio of the two determinations might increase the prognostic potential. methods: we collected bone marrow samples for mrd assessment at the end of induction therapy and months after asct from mm patients treated between and achieving at least a very good partial remission (vgpr) with a bortezomib-based induction therapy, according to the most recent international myeloma working group (imwg) criteria (kumar s et al, lancet oncol ) . mfc-determined mrd was evaluated according to euroflow recommendations (kalina t et al, leukemia ) . all patients were examined with fluorodeoxyglucose positron emission tomography/computed tomography (fdg-pet/ct) scan before and after the asct. results: post-induction therapy mrd was found predictive of post-asct mrd status. indeed, patients transplanted in a mrd positive status had a significantly increased risk to maintain a mrd positivity status after transplantation (odds ratio -or - , , p = , ). detection of post-asct mrd had a negative impact on median pfs ( months vs not reached respectively, p = , ). in cox-regression analysis, a complete remission status (cr) with an undetectable mrd after the asct resulted to be the major protective factor from relapse (hazard ratio -hr - , , p = , ), while patients with a detectable mrd before and after the asct had the worse pfs ( months, hr , ; p = , ). risk analysis showed different pfs risk groups: "high" for the patients with mrd detectable before and after the asct, "intermediate" for patients with mrd positivity before the asct who achieve a negativity after, and "low" in the case of mrd undetectable before and after. in our study, response evaluated by fdg-pet/ct showed no correlation with pfs. conclusions: multiparametric flow cytometry is a relatively recent method to assay mm mrd, and its role in mm therapeutic path is still under investigation. according to our data, a detectable mrd after the asct is a major relapse risk. interestingly we found that it can be early predicted by the post-induction mrd status and its negativization after asct has a modest impact on this. therefore, we support the concept of treatment escalation when a cr is not reached after the induction treatment, in order to undergo to the asct in the best possible response. however, double mrd determination before and after vtd, with ≥ % of patients achieving a best response of ≥vgpr. the orr in patients receiving 'other' induction therapies was % and the ≥vgpr rate was %. finally, following auto-sct, the orrs for patients receiving vtd induction were around %, %, and % in lines - , , and +, respectively. conclusions: this analysis provides prospective, realworld data on therapy of patients with mm receiving auto-sct. vtd is the most widely used induction regimen prior to auto-sct. moreover, the response rates are in line with reported rates in phase iii clinical trials. while other induction regimens are being developed, vtd is likely to remain a standard of care, because access to novel agents will continue to vary greatly from country to country due to factors such as affordability, local guidelines/restrictions, and regulatory decisions. background: car-t cell therapy against the cd antigen is a breakthrough treatment for patients with relapsed/ refractory (r/r) b-cell non-hodgkin lymphoma (nhl). despite impressive outcomes, non-response and relapse with cd negative disease remain challenges. through dual b-cell antigen targeting of cd and cd , with a first-in-human bispecific lentiviral car-t cell (lv . car), we attempt to improve response rates while limiting relapses due to cd antigen loss. production was optimized with point of care automated manufacturing using the clinimacs prodigy, a compact gmp compliant tabletop device in an iso clean room. methods: patients were treated on our phase dose escalation + expansion trial (nct ) to demonstrate feasibility of point of care manufacturing and safety of a bispecific bb/cd z lv . car t cell for adults with r/r b-cell nhl. safety was assessed by incidence of dose limiting toxicities (dlts) within days postinfusion. dose was escalated in incremental + fashion with a starting dose of . x cells/kg and a target cell dose of . x cells/kg. lymphodepletion was with fludarabine mg/m x days and cyclophosphamide mg/m x day. patients received either fresh uncryopreserved car-t cell infusions (n= ) or cells thawed (n= ) after cryopreservation. results: patients have completed treatment: patients in dose escalation and patient in dose expansion. median age was years ( - ) and histology included dlbcl in patients, mcl in patients, and cll in patients. in dose escalation, patients were treated at . x cells/kg, patients at . x cells/kg, and patients at . x cells/kg with no dlts to report. no patient experienced grade - cytokine release syndrome (crs) or grade - neurotoxicity (ntx) allowing start of a dose expansion cohort at the . x cells/kg level. in total, patients had grade - crs and patients had grade - ntx. mean time to crs was day + post-infusion and no patient required icu level care. patients required - doses of tocilizumab. the day overall response rate for all patients was % with / achieving a complete response (cr) and / achieving a partial response (pr). all patients in cr remain in remission, the longest months from treatment. car-t persistence is demonstrated in figure . two patients had progressive disease (pd) at day and patients with pr, eventually progressed. all progressing patients underwent repeat biopsy, and all retained either cd or cd positivity. target dose lv . car t cells were produced in all patients indicating % feasibility of our manufacturing process. conclusions: phase results from the first-in-human bispecific lv . car t clinical trial demonstrate that near patient manufacturing and infusion of . x cells/ kg is safe for further investigation with no dlts among treated patients. point of care production logistics aided the administration of fresh car-t cells in the majority. with limited toxicity and % sustained response in this relapsed refractory population, this approach to car-t production and dual b-cell targeting merits further investigation. o car-t cell therapy bridging to allogeneic hematopoietic cell transplantation for patients with refractory and relapsed acute lymphoblastic leukemia jia chen , , yi fan , , yang xu , , suning chen , , huiying qiu , , xiaowen tang , , yue han , , chengcheng fu , , depei wu , background: refractory and relapsed (r/r) acute lymphoblastic leukemia (all) always leads to a dismal outcome. allogeneic hematopoietic cell transplantation (hct) is the only potentially curative modality for r/r all, but the long-term survival post-hct remains unsatisfying. car-t cell therapy targeting to cd produces promising response for r/r all patients, but the recurrence is the major concern. we investigated the effectiveness of a tandem protocol using car-t cell therapy followed by allogeneic hct. methods: we conducted a prospective study to enroll the patients with r/r all. major inclusion and exclusion criteria are: ) definitely diagnosed as all; ) primary refractory (failed to achieve cr after induction) patients or relapsed patients with no response for salvage therapy; ) with an available donor for allogeneic hct; ) without severe organ dysfunction or uncontrolled infection. patients enrolled received car-t cell therapy targeting to cd with a total dose of ~ × /kg of recipient weight, and the preparative regimen before car-t cell infusion consisted of fludarabin and cyclophosphamide. after the evaluation at days post car-t cell infusion, patients started the allogeneic hct procedure using a myeloablative conditioning (modified bu/cy). the control group consisted of patients with r/r all and underwent allogeneic hct without car-t therapy in the same time frame. results: totally patients were enrolled in this study from december, through april, , including primary refractory patients and relapsed patients (details in table ). twenty patients ( . %) achieved cr after car-t cell therapy, and patients ( %) developed grade or higher crs. no irreversible toxicities emerged and all the patients moved to the hct procedure. when comparing with the control group (figure ) , the -year cumulative incidence of relapse was . ± . % for the trial group versus . ± . % for the control group (p = . ), and -year overall survival was . ± . % versus . ± . % (p = . ). conclusions: we concluded that car-t cell therapy bridging to allogeneic hct is a promising approach for r/r all patients, which leads to both high response and low risk of relapse. besides, car-t cell therapy is a safe modality as salvage treatment for r/r all patients, which had little negative impact for following hct. methods: to test the hypothesis that haploidentical hct would be a valid option for high-risk pediatric aml patients lacking a matched donor, we designed a diseasespecific, multi-centre study. we retrospectively analyzed consecutive patients under years with high-risk aml underwent matched sibling donor (msd) (n= ) or haploidentical donor (hid) hct (n= ) between july, and dec, . a : ratio matched pair analysis was implemented with the following matching factors: cytogenetic risk, disease status (cr /cr />cr ), age and sex of patients, sex of donor, and graft type. results: all patients achieved myeloid recovery with a median time of d and d for msd cohort and hid group (p= . ). the cumulative incidence of grade ii-iv acute graft-versus-host-disease (gvhd) in msd cohort ( %) was significantly lower than in hid group ( %, p= . ); the incidence of chronic gvhd was comparable between the two groups. the cumulative incidence of relapse in msd cohort ( %) was significantly higher than in hid group ( %, p= . ); the incidence of nrm was and % (p= . ), respectively. the -year overall survival ( % versus %, p= . ) and leukemia free survival ( % versus %, p= . ) were comparable in msd-hct compared with hid-hct. in a multivariate analysis, hid-hct remained a significant factor for reduced relapse rate (hr . ( . - . ), p= . ) in comparison with msd-hct. in subgroup analysis for patients with known cytogenetics and transplanted in the first complete remission (n= ), the cumulative incidence of relapse in msd cohort ( %, n= ) was significantly higher than in hid group ( %, n= , p= . ); and leukemia free survival ( % versus %, p= . ) were significantly lower in msd-hct compared with hid-hct. in a multivariate analysis among these subgroup of patients, hid-hct remained a significant factor for increased lfs (hr . ( . - . ), p= . ) in comparison with msd-hct. conclusions: in conclusion, unmanipulated haploidentical-hct achieves outcomes comparable to those of isd-hct for high-risk pediatric aml patients and even exerts greater gvl effect in some circumstances. such transplantation was proved to be a valid alternative treatment for high-risk pediatric aml patients lacking a matched donor. larger prospective studies are needed to confirm these findings. disclosure: nothing to declare. background: allogeneic hematopoietic stem cell transplantation (hsct) is the only curative option for patients with beta thalassemia major. although limited study in the literature has evaluated the impact of age on success of transplantation, more data are needed. in this study, we aimed to evaluate the effect of age of the patients on transplant outcome in cases who underwent hsct with the diagnosis of thalassemia major. methods: all cases who underwent stem cell transplantation with thalassemia major were included. all thalassemia major patients with a median age of years (range month, . years) underwent allogeneic hsct using myeloablative conditioning regimen. cyclosporine and methotrexate were used as gvhd prophylaxis. in total, patients underwent hsct at age younger than and patients underwent at age older than years and all patients were assigned to two different groups according to transplantation age. the distribution of donor type and stem cell sources by age groups is shown in table . no statistical difference was found between the two age groups in terms of donor type and stem cell source. patients in two different age groups were compared with cox regression analysis in terms of overall survival, thalassemia free survival and thalassemia-gvhd free survival. results: a total of patients; patients under years of age, patients aged and over, were engrafted and remained transfusion independent with full or mixed chimerism. four patients, two from each age group, did not engrafted and had primary rejection. four patients under seven years of age developed secondary rejection, whereas in the group of patients older than years of age, patients experienced secondary rejection (p< . ) . a total of patients developed acute gvhd ( . %)and their rates were similar in both age groups ( . % vs %). chronic gvhd development rates in two group was also similar ( % vs %). the median follow-up time was months (range . - months). the -year overall survival rates (os), thalasemia-free survival rates (tfs), thalassemia-gvhd free survival rates (dfs) were shown in the table . both thalassemia-free survival and thalassemia-gvhd free survival were higher in patients who underwent transplantation under seven years of age . there was no difference in overall survival. conclusions: the results of our study show that the rates of rejection are high, thalassemia free and thalassemia / gvhd free survival are low in patients who underwent stem cell transplantation over seven years of age. in the light of successful transplantation results from unrelated donors, the delay in age of transplantation in thalassemia patients should also be avoided. disclosure: nothing to declare o hla-haploidentical transplantation with regulatory and conventional t-cell adoptive immunotherapy in pediatric patients with high-risk acute leukemia background: post-transplant relapse is still a major cause of treatment failure in high-risk acute leukemia (al) patients. in order to separate the gvl effect from gvhd, we investigated the role of a thymic-derived cd + cd + foxp + regulatory t cells (tregs). the perugia center reported results from adult high-risk al patients who received an hla haploidentical t-cell-depleted hematopoietic transplant and adoptive immunotherapy with donor tregs and conventional t cells (tcons) (and no posttransplant pharmacologic immunosuppressive gvhd prophylaxis) (di ianni et al., blood , martelli et al. blood . adoptive immunotherapy with tregs and tcons prevented post-transplant leukemia relapse and largely protected patients from gvhd. in this report we present a pediatric cohort of high risk leukemia patients who received a haploidentical treg/tcon-based hematopoietic transplant. methods: twelve pediatric patients, median age of nine years (range, - ) with high-risk acute leukemia underwent hla-haploidentical stem cell transplantation with regulatory and conventional t-cell adoptive immunotherapy between september and december . eleven had all (three ph+), one secondary aml. seven patients were transplanted in cr ( ph+ all, all in cr after second-line induction, all with extramedullary leukemia, all with t( ; ), secondary aml after medulloblastoma), two patients in cr , three in cr . median time from diagnosis to transplantation was . months (range, - ), median time from relapse to especially for aml pts. our analysis suggests that early αβ t cell recovery is associated with a relatively low nonrelapse mortality and relapse rate. disclosure: nothing to declare background: enhancing stem cell performance can improve the results of hematopoietic stem cell transplant (hsct) for diseases in which engraftment is unpredictable, and in patients receiving pre-hsct conditioning regimens of progressively decreasing intensity. in a series of preclinical studies, we explored the use of tat-myc, a chimeric recombinant protein, to improve the performance of hematopoietic stem cell grafts. methods: tat-myc recombinant protein encompasses amino acids from the n-terminal nuclear localization domain of hiv-tat, coupled to the entire coding sequence of c-myc, with an appended histidine tag to aid in protein purification. the construct was expressed in bacteria and purified to pharmacological grade purity under glp conditions. results: brief ( hour) culture of fibroblasts or hematopoietic cells in medium containing microgram/ ml tat-myc results in rapid nuclear localization of the recombinant protein, whence it disappears within hours as measured by western blot. flow cytometric assays have been validated to measure the uptake of tat-myc recombinant protein into nucleated marrow cells. marrow homing of tat-myc recombinant protein-treated murine marrow increased -fold as compared to that of control cells. incubation of activated murine t cells with tat-myc recombinant protein conferred resistance to granzyme b cytotoxicity, but did not protect cells from effects of cyclophosphamide. tat-myc recombinant protein-treated marrow could be serially transplanted in mice for three generations. murine bone marrow harvested from fluorouracil treated mice and briefly incubated with tat-myc recombinant protein outcompeted control marrow when transplanted in sub-lethally irradiated immunedeficient mice even at ratios of : treated:control cells. t-and b-cell reconstitution following transplantation in immune deficient mice was superior following tat-myc vs control incubation of murine marrow. engraftment of human umbilical cord blood (ucb) cells in sub-lethally irradiated immune deficient mice was markedly improved following tat-myc incubation as compared to that of control ucb cells. the transforming potential tat-myc protein was extensively explored. tat-myc culture hematopoietic cells did not display aneuploidy in cytogenetic or spectral karyotypic analyses. intramuscular injection of micrograms of tat-myc protein in p +/mice for consecutive weeks did not result in tumor formation. to exaggerate potentially transformative effects of tat-myc protein, murine marrow was co-incubated with both tat-myc and tat-bcl recombinant proteins prior to transplantation into irradiated immune deficient mice. mice were followed for weeks and none developed malignancies. serially transplanted marrow incubated with both tat-myc and bcl- proteins did not result in malignancies in recipient mice. of > mice that have been exposed to tat-myc recombinant protein in our experiments, none has developed a tumor. conclusions: in preclinical studies, brief incubation with tat-myc recombinant protein enhances homing, engraftment and immune reconstitution of murine and human cells in recipient mice following transplantation. brief exposure to the recombinant protein (as opposed to transduction of the myc gene) does not cause malignant transformation of cells. we are currently developing clinical trials using tat-myc protein to enhance engraftment following hsct. disclosure: greg bird, brian turner, thomas payne, yosef refaeli are employees and or shareholders in taiga biotechnologies. jerry stein has received laboratory support from taiga biotechnologies graft γδ t-cell receptor sequencing identifies public clonotypes associated to hsct efficacy in aml patients and unravels cmv impact on repertoire distribution lucas cm arruda , ahmed gaballa , michael uhlin , , relapse patient group had an increased proportion of long cdr sequences ( - nucleotides) compared to relapse patients [ . %vs . % (p= . ) and . %vs . % (p= . )]. grafts from cmv-positive donors presented significantly reduced diversity (inverse simpson's di: . vs . , p= . ), decreased proportion of cdr sequences having , , and nucleotides [ . %vs . % (p= . ), . %vs . % (p= . ), . %vs . % (p= . ) and . %vs . % (p= . )], and an increase of sequences with - nucleotides ( . %vs . %, p= . ). hyperexpanded clones took up . times more space in the cmv positive grafts ( . % vs . %, p= . ), who presented a skewed non-gaussian distribution. for all samples, the segments trgv and trgjp were the most frequent. nonrelapsing group received grafts with lower usage of trgv , trgv and trgjp segments and higher usage of trgjp compared to relapse patients [ . %vs . % (p= . ), . %vs . % (p= . ), . %vs . % (p= . ) and . %vs . % (p= . ) ]. cmv-positive donor grafts presented a lower trgv and trgjp expression ( . %vs . %, p= . , and . % vs . %, p= . ) as well as a higher trgjp gene usage ( . %vs . %, p= . ). the v -jp combination was the most frequent pairing in all samples. non-relapse patients received grafts with lower usage of the pairing v -j , v -j , v -jp [ . %vs . % (p= . ), . % vs . % (p= . ) and . %vs . % (p= . )] and higher usage of v -jp pairing ( . %vs . %, p= . ) than relapse patients. the tcr usage of the sequence pairs v -j , v -jp , v -jp , v -jp, and v -jp was lower in cmv-positive grafts [ . %vs . % (p= . ), . %vs . % (p= . ), . %vs . % (p= . ), . %vs . % (p= . ) and . %vs . % (p= . )]. we identified public clones shared exclusively between the grafts received by non-relapsing patients in addition to four private over-represented sequences exclusively present in grafts given to nonrelapse patients, taking from . % to . % of the trg repertoire and longer than nucleotides. we also identified five private over-represented and one public cdr sequence associated to cmv infection. additionally, cmv-positive grafts presented the highest percentage or repertoire taken by private over-represented clones, ranging from . % to . %. conclusions: our findings show that the trg composition is not associated to agvhd incidence, cmv infection reshapes the trg repertoire and several public sequences are associated to clinical remission. disclosure: the authors have nothing to disclose. background: pediatric patients with high-risk alveolar rhabdomyosarcoma (arms) above the age of years cannot be cured by conventional therapies. immune cells targeting erbb with a chimeric antigen receptor (car) were recently considered for these patients. cytokineinduced killer (cik) cells already capable of natural killer (nk)-like anti-tumor capacity additionally redirected with an erbb car may provide overall disease control in these high-risk tumors. methods: erbb -car modified cik cells were generated from conventional cik cells (wt-cik) by lentiviral gene transduction on day of culture. the codon-optimized car sequence consists of an igg heavy-chain signal peptide, an erbb -specific antibody fragment scfv (frp ) and a modified cd α hinge region, as well as cd transmembrane and intracellular domains and a cd ζ intracellular domain. x luciferase gene-transduced rh (arms) cells were engrafted in immunodeficient nod/scid/γc -(nsg) mice. mice were randomly selected into different treatment groups (dbps on day + , . x wt-cik or erbb car-cik cells on days + and + , . x wt-cik or erbb car-cik cells on days + and + ). mice were monitored by bioluminescence imaging (bli) until day + . tumor engraftment and immune cell homing at tumor sites were analyzed by facs, chimerism and immunohistochemistry analyses. results: human rms xenografts were established in all mice treated with dbps only. control-mice showed a median survival of days. human rms was identified in all analyzed organs, with the highest tumor burden seen in livers of dbps-treated mice. mice injected with wt or erbb -car cik cells on days + and + showed a significant improved (p < . and p < . ) disease-free survival, respectively. furthermore, no signs of tumor engraftment were shown by bli in erbb car-cik cell treated mice while some of the mice treated with wt-cik cells developed positive tumor signals between weeks and . in out of ( %) wt-and in all ( of , %) car-cik cells treated mice no residual tumor cells were identified by pcr-based analysis. in contrast, tumor cells were detectable in all mice with delayed anti-tumor treatment applied on day + and + . however, tumor growth was lower in these groups. correspondingly, bli showed delayed tumor engraftment in mice with wt-and even more with car-cik cell treatment given on day . treatment on day resulted in a significantly improved survival of erbb -car cik cell treated mice (p < . ), while survival was not improved after wt-cik cell infusion (p > . ). within all treatment groups, immune cells were detected by chimerism and facs analyses. facs analyses showed a significant increase of nk-like t cells (p < . and < . , wtand erbb -car cik cells). additionally, a higher, but not significant, amount of effector memory and stem cell memory t cells were detected. conclusions: these pre-clinical in vivo results indicate that erbb -car redirection of cik cells improves both homing and nk-like cytotoxicity of cik cells in the presence of erbb -positive tumors, implying that this therapy may represent a step forward in the treatment of patients with resistant, relapsed and advanced rms. disclosure: michael merker, juliane wagner, vida meyer, thomas klingebiel, winfried s. wels and eva rettinger have nothing to declare. peter bader declares the following potential conflicts of interest: novartis (consultancy: included expert testimony, speaker bureau, honoraria), medac (research funding, patents and royalties), riemser (research funding), neovii (research funding), amgen (honoraria) . genesis -a phase iii randomized double-blind, placebocontrolled trial, evaluating safety and efficacy of bl- and g-csf in mobilization of hcs's for autologous transplantation-multiple myeloma hemda chen , zachary d. crees , keith stockerl-goldstein k , abi vainstein , ella sorani , osnat bohana-kashtan , john f dipersio biolinerx, tel aviv, israel, washington university in st. louis, st. louis, wa, united states background: cxcr mediates retention of hematopoietic stem cells (hscs) in the bone marrow (bm) niche. bl- , a novel, high affinity cxcr antagonist is a potent mobilizer of hscs to the peripheral blood with numerous potential clinical applications, including mobilization of cd + cells for autologous hsc transplantation (auto-hsct) in multiple myeloma (mm). this study aims to evaluate the efficacy of single dose bl- plus g-csf in mobilization of ≥ . x cd + cells/kg in up to apheresis sessions for auto-hsct in mm. methods: a phase iii study composed of an open-label, single-arm lead-in part followed by a randomized, doubleblinded, placebo-controlled part . eligible mm patients age - will receive g-csf ( μg/kg; sc) daily for up to days and one dose of bl- ( . mg/kg; sc) or placebo on day followed by up to apheresis sessions; and if needed a second dose of bl- or placebo on day followed by up to apheresis sessions. progressive disease at time of ldc did not respond although car-t cells could be seen morphologically under the microscope. this might be explained by multidrug related phenomenon protecting refractory leukemia from car-t cell attack. conclusions: commercial available car-t cell product tisagenlecleucel (kymriah®) showed high efficacy in r/r-all patients to re-induce cr. clinical trial registry: commercial available car-t cell product tisagenlecleucel (kymriah®) showed high efficacy in r/r-all patients to re-induce cr. disclosure: pb: novartis (consultancy: included expert testimony, speaker bureau, honoraria); medac (research funding, patents and royalties); riemser (research funding); neovii (research funding); amgen (honoraria) . aj: novartis and bluebird: (consultancy) . all other author declare no coi. the main causes of death were sct-related in % and disease in %. -months pfs, os, ir and nrm were % ( - ), % ( - ), % ( - ) and % ( - ), respectively. cumulative incidence of grade - acute gvhd at days after ric-sct was % ( - ) and chronic gvhd at months % pfs and ir were influenced by patient sex (p= . and . ) and disease status at allo-sct ) and stem cell source (p= . ); acute grade - gvhd by donor type (p= . ) and chronic gvhd by allo-sct conditioning (p= . ) and donor sex (p= . ). conclusions: this is the largest series analysing the efficacy and safety of a tandem auto-ric-sct approach in r/r hl. the low nrm and ir with promising pfs and os suggest that this might be an effective post-transplant cyclophosphamide-based gvhd prophylaxis compared to standard prophylaxis in patients with lymphoma receiving hla identical transplantation: a retrospective study from the lwp of ebmt luca castagna ebmt lymphoma worky party clinical trial registry: nct disclosure: richard t. maziarz: honoraria, membership of advisory committee and research funding employment: oregon health & science university (ohsu); the potential conflict of interest re: consultant services to and payment from novartis has been reviewed and managed by dava oncology; honoraria and research funding: genentech; membership on an entity's board of directors or advisory committees membership on an entity's board of directors or advisory committees and research funding: novartis honoraria and membership on an entity's board of directors or advisory committees: nordic nanovector. vadim v. romanoff: employment: novartis employment: novartis. james signorovitch: employment: analysis group, which received research funding from novartis. solveig g. erickson: employment: novartis. david g. maloney: research funding other: scientific advisor: kite pharma, novartis disclosure: nothing to declare o multiple myeloma treatment in real-world clinical practice: a focus on induction regimens prior to autologous stem cell transplantation from the prospective, multinational, non-interventional emmos study cic, ibmcc (usal-csic) state budget healthcare institution of moscow. city outpatient clinic of healthcare use of unmanipulated hla-haploidentical donor transplants (haplo-hsct) is constantly increasing in the last years. few cases of haplo-hsct using posttransplant-cyclophosphamide (pt-cy) for pediatric patients were reported by single center and registry studies, with an incidence of grade ii-iv agvhd ranging from % to % and cgvhd approaching up to %, although with low incidence of extensive disease. methods: we investigated the outcomes of children (< = y) undergoing haplo-hsct using pt-cy as gvhd prophylaxis disease status at haplo-hsct was cr for %, cr % and advanced for %. poor-risk cytogenetics was reported in % of aml, and % had ph+ all all patients received pt-cy, in association with tacrolimus/mmf in % or csa/mmf in % or engraftment rate was % with patients experiencing graft failure. cumulative incidence (ci) of day- acute gvhd grade ii-iv and grade iii-iv were % and % respectively, and ci of -y chronic gvhd was % ( % extensive disease). -y ci of nrm was % and relapse %. disease recurrence and infections were the most common causes of death. y-os and lfs were % and %. y-os was % and % (p= . ) for aml and all; it was %, % and % (p< . ) for patients transplanted in cr , cr and advanced disease. for y-lfs, no significant difference was found according to the type of conditioning regimen ( % macchemotherapy-based, % mac-tbi based and % for ric, p= . ). the use of pbsc was associated with higher ci of grade os: cr vs cr conclusions: pt-cy is effective in preventing severe gvhd in children with leukemia receiving an unmanipulated haploidentical-donor transplant. disease status remain the most important factor for outcomes. the use of pbsc as stem cell source increases the risk of grade ii-iv agvhd. the effect of long-term complications, and morbidity related to gvhd results: all patients achieved primary, sustained fulldonor engraftment (median neutrophils engraftment days, range - ; median platelets days, - ). five patients ( %) developed ≥ grade agvhd ( / had concomitant hcv hepatitis and developed liver agvhd), none developed cgvhd. the immune recovery was good in all patients despite immune suppressive therapy in patients with agvhd causes of nrm were: agvhd, invasive aspergillosis, thrombotic microangiopathy. nine of the patients are alive at a median follow-up of months ( - months), cgvhd/ leukemia-free survival is %. conclusions: these preliminary data in very high risk pediatric patients showed that hla-haploidentical transplantation with regulatory and conventional t-cell adoptive immunotherapy russian federation background: relapse, gvhd and associated non-relapse mortality are the main obstacles to successful hsct in children with leukemia. αβ t cell depletion was developed to prevent gvhd and improve immune reconstitution in recipients of mismatched grafts either melphalan (n= ) or thiophosphamide (n= ) or etoposide (n= ) were added, fludarabine was used in all pts. two types of gvhd prophylaxis were used: type (n= ): hatg mg/kg and post-hsct tacro/mtx, type (n= ): thymoglobulin(ratg) mg/kg the median dose of cd + cells was x /kg, aβ t cells - x /kg. results: five patients ( , %) died before engraftment due to septic event. primary engraftment was achieved in all evaluable pts ( %) with full donor chimerism. among the whole cohort the ci of gvhd grades ii -iv and iii -iv was - ), p= , . ratg was also effective in prevention of cgvhd: ci at year after hsct was % vs %, p= , . -year ptrm was in the group with available immune reconstitution data (n= ) αβ t cell recovery at day + was associated with a trend to decreased incidence of relapse, ci of relapse was % ( % ci: - ) in those with αβ-t cell count < median vs % ( % ci: - ) in those with αβ-cell count >median italian bone marrow donor registry hematopoietic stem cell transplantation (hsct) from hla-a, -b, -c and -drb -matched unrelated donors ( / ud) is performed across hla-dpb barrier in more than % of cases. clinically tolerable (permissive) mismatches (mm) at hla-dpb locus have been classified by different immunogenetic models. here we compare the prognostic value of these models ii) a similar model subdividing alleles in tce groups (tce , crocchiolo iii) differences in "delta functional distance" scores of polymorphic aminoacids in hla-dpb peptide-binding groove (crivello while the first three models were applicable to all hla-dpb mm patients, the latter was restricted to of them. the tce model appeared the most restrictive one, with only % of mm considered to be permissive. median follow-up was . y. results: hla-dpb permissive (p) mm pairs defined by tce model (n= ) had superior -y overall survival (os) and gvhd-free & relapse-free survival (grfs) compared to non-permissive (np) mm (n= ) ( ± % vs ± % cgvhd (hr . , p . ) and extensive cgvhd (hr . , p< . ). the predicted hla-dpb mismatched allele expression in the recipient was associated with -d ci of grade≥ agvhd: ± % in high expression (n= ) versus ± % in low expression (n= ), p< . . this was confirmed in multivariate analysis for grade≥ agvhd (hr . , p< . ), however, without higher hazards for trm and overall mortality. the overlap among the four models and their adjusted hr for os is shown in figure . conclusions: functional hla-dpb matching is of prognostic value in / ud-hsct outcomes. in our cohort, tce appears superior to other models in predicting survival and stratifying risks of trm and cgvhd however, it remains unclear how outcomes of patients with all treated with a haploidentical donor (haplo) compare with hla matched unrelated donor (mud) transplants. methods: we, therefore retrospectively compared outcomes of patients with all who underwent haplohct with ptcy, reported from the participating centers (hit-rc and ebmt) from / to / , with a matched cohort of patients ( : ) who underwent mud-hct and were reported to ebmt. patients were matched for sex, age at transplant (≤ or > ), disease type (b-all vs. t-all), disease stage (cr vs. cr vs. other), disease risk (high vs. others), philadelphia chromosome status (positive vs negative), and conditioning regimen (mac vs. ric/nma) in multivariable analysis, os, pfs, nrm, and relapse rate were not statistically different between patients receiving hct from haplo or mud, regardless of the intensity of the conditioning regimen; (table ). conclusions: in conclusion, in this large retrospective analysis, outcomes of patients with all undergoing transplant from a haploidentical donor with post median follow-up was , months and was similar in both cohorts (p= . ). -year overall survival (os) was . % for the all patients and did not differ between transplants from a ird or a crd ( % vs %, p= . ; table i). -year progression-free survival (pfs) and gvhd/relapse-free survival (grfs) was % and %, respectively. -year non-relapse mortality (nrm) was % and was similar between ird and crd (p= . , table i). -months cumulative incidence of grade - acute gvhd, grade - acute gvhd and -year moderate-severe chronic gvhd was %, % and %, respectively. again, there was no difference between crd and ird transplants in terms of grade - acute gvhd and moderate-severe chronic gvhd (table i) conclusions: our results confirm previous findings that a crd haploidentical transplant is a viable option for haplo-sct when a first-degree donor is available. main long-term outcome are not different in this matched pair analysis. extending our analysis to a larger cohort of patients receiving a crd is warranted to confirm our preliminary results. references: disclosure: the authors have no conflict of interest to disclose stem cell mobilization, collection and engineering o a recombinant chimeric protein, safely enhances graft performance following hematopoietic stem cell transplantation median follow-up time for survivors was months. results: overall survival (os) and relapse-free survival (rfs) rates at years after transplantation were % %, respectively. cumulative incidences of non-relapse mortality (nrm) and relapse at years were % and %, respectively. in multivariable analysis, performance status (ps) over (ps> vs ps < = , hr . , p= . ) and lymphoma progression at transplantation (cr/cr-u/pr vs others, hr . , p= . ) showed significant negative impacts on os. cbt was strongly associated with better os compared to unrelated bmt/pbsct (hr . , p= . ) and comparable to related bmt/pbsct. lymphoma control status at transplantation was significantly associated with relapse (relapse/induction failure vs cr/cr-u/pr, hr . , p= . ). poor ps over at transplantation (ps> vs ps < = , hr . , p= . ) and reduced-intensity conditioning (ric) regimen (ric vs myeloablative, hr . , p= . ) were associated with higher risk of nrm. conclusions: allo-hsct could improve overall survival of patient with mature t-or nk-cell lymphomas, if performed at appropriate timing with good disease control of partial remission (pr) or better. cord blood unit could be a favorable alternative donor source when related donors are not available. on the other hands, ric regimen did not decrease the risk of nrm in allo-hsct for mature t-and nk-cell lymphoma patients in our setting. out study also suggested that major problem of allo-hsct is still a high frequency of relapse after transplantation. better disease control is mandatory to improve the outcomes of allo-hsct for mature t-or nk fabio ciceri , luca vago , katharina fleischhauer unite´de recherche mixte en sante´(umr_s) , inserm netherlands, institute of hematology and blood transfusion ptcy is largely adopted as gvhdprophylaxis backbone in haploidentical transplantation. the encouraging results prompted investigations to assess ptcy also in unrelated donor (ud) setting. high-resolution hla-matching contributes to ud-hsct success; however, due to the selective in-vivo deletion of alloreactive t-cells, ptcy could modulate hla-matching impact on ud-hsct. methods: we compared the outcomes of acute leukemia patients receiving / (n= ) and / (n= ) hlaallele matched ud-hsct with ptcy gvhd-prophylaxis table illustrates patients' characteristics. the power to detect a -years % difference grfs between the groups was % (alpha -sided= %). results: outcome endpoints at years were not different between ± %, p= . ; lfs: ± % and ± %, p= . ; os: ± % and ± %, p= . , respectively). the -day ci of grade≥ and grade≥ agvhd were comparable for / and / ud ( ± % and ± %, p= . and ± % and likewise, the -y ci of cgvhd and extensive cgvhd were similar between the -y ci of trm was ± % after / and ± % after / ud-hsct (p= . ). relapse incidence at -y was ± % for / and no interaction was found between donor type and additional atg use. variables associated with grfs were active disease (hr . compared to st cr, p< - ) and karnofsky ps≥ % (hr . , p< - ). conclusions: in the present series of acute leukemia patients transplanted with ptcy, we report comparable survival with / and / hla-matched ud-hsct, across all disease stages, suggesting that this platform could alleviate the detrimental effect of single hla-allele mismatching. these results warrant prospective comparative trials of ptcy versus standard use of atg-based gvhd disease status: cr disclosure: nothing to declare o myeloablative conditioning for first allogeneic hsct in pediatric all: ftbi or chemotherapy? -an update of the retrospective multicenter ebmt-pdwp study rose-marie hamladji , cristina diaz de heredia , elena skorobogatova czech republic, hôpital robert debré and paris-diderot university methods: this update was done to extend the time of follow-up (fu, date of analysis: /oct/ ). to compare outcomes of ftbi vs cht-conditioning, we performed a retrospective ebmt-registry study. children aged - years (y) after mac for first allo-hsct of bm/ pbsc from msd/ud in cr /cr between - were included. propensity-score-weighting was used to control pretreatment imbalances of observed variables. this statistical method ensured that analyzed groups differed only in the parameter under investigation (here: conditioning) busulfan/cyclophosphamide/ etoposide (bu/cy/eto) was the most frequently applied cht-regimen in cr ( ( %)) and bu/cy in cr ( ( %)). the remaining conditionings bu/ cy ( ) or other-chemo ( ) with median-fu of . , . and . y. in weighted univariate analysis, -y-os was . % after other-chemo, . % after bu/cy and . % after ftbi. in weighted cox-model, pts having received other-chemo had a higher risk to experience an event compared to ftbi (hr= . , p=< . ). -y-lfs was coxmodel, pts having received bu/cy and other-chemo had a higher risk to experience an event compared to ftbi (hr= . , p=. ; hr= . , p< . ). -y-nrm (range: . % (bucy) to . % (other-chemo)) did not show significant differences in weighted cox-model. conclusions: this recent study-update ensured a substantial fu. we confirmed the clear superiority of ftbiconditioning compared to cht with regard of lfs and ri for all-pts undergoing allo-hsct in cr . for pts in cr we could not find significant differences between ftbi and cht-conditioning. these retrospective findings are currently re with data monitoring committee (dmc) review after each cohort. part will include patients randomized : . results: part enrolled patients, median age following these promising results, dmc recommended early continuation to part of the phase iii trial. conclusions: the genesis lead-in results demonstrate bl- is a potent mobilizer of hscs, with potential to improve mobilization rates while minimizing mobilizationrelated healthcare costs. clinical trial registry: nct disclosure: hemda chen -medical director, biolinerx abi vainstin -vp of medical affairs, biolinerx ella sorani-vp of r&d, biolinerx osnat bohana-kashtan-project manager markus y. mapara , john f. tisdale , julie kanter , janet l. kwiatkowski , , lakshmanan krishnamurti , manfred schmidt , alexandra l. miller , francis j. pierciey jr. background: phlh is a rare, genetic, hyper-inflammatory syndrome driven by high production of interferon (ifn)-γ. emapalumab (ni- ) is a monoclonal antibody that neutralizes ifn-γ and is developed as a treatment for hlh.methods: this open-label phase / study (nct , data cut-off is july ) includes patients ≤ years old with phlh based on genetic confirmation, family history, or presence of ≥ / hlh- diagnostic criteria. patients were treatment-naïve ( ) or had failed previous conventional hlh therapies ( ) . emapalumab ( mg/kg intravenously every - days, increased up to mg/kg based on clinical and laboratory response parameters) was administered with dexamethasone ( - mg/m /day with tapering permitted). treatment duration was weeks with possible shortening to a minimum of weeks or extension up to allogeneic hematopoietic stem cell transplantation (hsct). the primary endpoint, overall response rate (orr), was objectively assessed based on normalization or ≥ % improvement in pre-defined criteria: fever, splenomegaly, cytopenias, hyperferritinemia, fibrinogen and/or d-dimer levels, central nervous system (cns) abnormalities, with no sustained worsening of scd levels. an exact binomial test at one-sided . significance level was used to evaluate the null hypothesis that orr be at most %. following completion of the study, patients entered an extension phase (nct ).results: patients (median age . yr, range . - yr) entered the study with broad spectrum of phlh clinical abnormalities, > % with cns involvement. mutations in phlh-associated genes were present in % of patients. orr was significantly higher than the pre-specified null hypothesis, thus meeting the primary efficacy endpoint (table) . response rates based on investigator's clinical judgement were . % and . % in the two groups.emapalumab was safe and well tolerated. mild to moderate infusion-related reactions occurred in % of patients. infection caused by pathogens potentially favored by ifn-γ neutralization occurred in patient (disseminated histoplasmosis, resolved with treatment). no off-target effects were observed. most patients proceeded to hsct ( % of patients received myeloablative conditioning and % reduced intensity conditioning) with favorable outcome (engraftment in % and % of patients, respectively) and % of patients receiving hsct were alive at year post transplant.background: asp is a first-in-class, dna-based vaccine designed for the prevention of cytomegalovirus (cmv) infection; it contains two plasmids encoding glycoprotein b (gb) and phosphoprotein (pp ). this study aimed to investigate the efficacy, safety and immunogenicity of asp compared with placebo in allogeneic haematopoietic cell transplant (allo-hct) recipients.methods: this was a randomised, double-blind, placebocontrolled phase study. cmv-seropositive allo-hct recipients received five intramuscular ml doses of mg/ ml asp or placebo (phosphate buffered saline) in a : ratio on days − to − , to , ± , ± and ± , relative to the day of transplant (day ). plasma cmv viral load was determined through year and analysed at the central laboratory. treatment-emergent adverse events (teaes) were recorded through days after the last randomized treatment injection. immunogenicity was measured by t-cell responses to pp and gb-specific antibody levels through year after the first randomized treatment injection. the primary endpoint was the proportion of patients with a composite of all-cause mortality and adjudicated cmv end-organ disease (eod) through year post transplant. secondary endpoints were cmv viraemia rate, adjudicated cmv-specific antiviral therapy (avt) rate, a composite of protocol-defined cmv viraemia and adjudicated cmv-specific avt use, first occurrence of background: haploidentical stem cell transplantation (haplo-sct) with post-transplant cyclophosphamide (pt-cy) represents a potential curative strategy for patients with hodgkin lymphoma (hl) when a matched related or unrelated donor is not available . while bone marrow (bm) was originally the preferred stem cell source, more recently peripheral blood stem cell (pbsc) is more often used. some retrospective studies suggest that the risk to develop acute and chronic graft-versus-host-disease (gvhd) is higher with pbsc than bm, while pbsc may reduce the risk of relapse . here we analyzed the effect of stem cell source in hl patients receiving haplo-sct with pt-cy, with the aim to evaluate if the final outcome is modified by the use of pbsc or bm.methods: from april to january , consecutive patients with poor prognosis hl received a haplo-sct with pt-cy either from a pbsc (n= ) or bm (n= ). the two cohorts were similar for most characteristics, but the pbsc group had more patients with an unfavorable hematopoietic stem cell transplant comorbidity index (hct-ci) score ≥ (p= . ) and had received a non myeloablative conditioning (nmac; p= . ).results: cumulative incidence of neutrophil> /ul at day + and of platelet > /ul at day + were % ( % ci: - ) and % ( % ci: - ), respectively, with no significant differences between the pbsc and bm cohorts. with a median follow-up of . months, there was no difference between pbsc and bm graft in terms of cumulative incidence of grade - acute gvhd ( % vs %, p= . ), grade - acute gvhd ( % vs %, p= . ) and moderate-severe chronic gvhd ( % vs %, p= . ). this was also confirmed by multivariate analysis. in the whole population, the -year overall survival (os), -year progression-free survival (pfs) and -year gvhd/relapse free survival (grfs) rates were %, % and %, respectively. we observed a trend for improved os ( % vs %, p= . ) and pfs ( % vs %, p= . ) for recipients of pbsc relative to bm cells, but pre-transplant disease status was the only significant variable by univariate analysis (table i) . by multivariate analysis, pre-transplant active disease status, transplant from a bm and hct-ci ≥ remained the only independent predictors of adverse outcome in terms of os; pfs and grfs (table i) . nonrelapse mortality was not affected by graft source both by univariate and multivariate analysis, while pre-transplant disease status was the only variable affecting the chance of disease relapse.conclusions: overall these data suggest that pbsc is associated with better outcome, in terms of os, pfs and grfs, relative to bm cells as graft source for patients undergoing haplo-sct with pt-cy. in addition, the risk of acute and chronic gvhd is not increased after pbsc relative to bm graft.references disclosure: the authors have no conflict of interests to disclose o consensus crs and neurotoxicity regrading of "juliet": phase ii prospective study of tisagenlecleucel therapy in patients with relapsed/ refractory large b cell lymphoma background: t-cell depletion using ex vivo cd + cell selection reduces gvhd risk after allogeneic hct, but delayed immune reconstitution, particularly t-cell reconstitution, has limited improvement in survival. sex steroids negatively impact lymphopoiesis, likely by thymic atrophy, and our preclinical models have shown that androgen blockade with the gnrh agonist leuprolide enhances thymopoiesis (goldberg, ji ; velardi, jem ) . we hypothesized that peri-hct leuprolide could improve immune reconstitution among recipients of cd selected hct.methods: this was a phase ii clinical trial of leuprolide in cd -selected myeloablative allogeneic pbsct for hematologic malignancies in patients aged - (nct ). all participants received conditioning with tbi cgy, thiotepa mg/kg, and cyclophosphamide mg/kg; antirejection prophylaxis with rabbit atg - . mg/kg; and palifermin mg/kg/d on days - to - and to + . patients received a -month depot of leuprolide . mg - weeks before conditioning and a second depot months later. primary endpoint was an absolute cd + count >= by months post-hct. patients who died, relapsed, or otherwise did not have flow data available at day + were excluded from primary endpoint analysis but included in outcome analyses. we excluded flow data after secondary cell infusions (dli, ctls, second hct, cd + cell boost) but followed recipients of these interventions for survival analysis. descriptive statistics summarized absolute levels of lymphocyte subset counts at select time intervals. a kruskal-wallis rank sum test compared counts among patients who received leuprolide/palifermin, historical controls who received palifermin alone, and historical controls who received neither. kaplan-meier functions estimated os/ rfs. cumulative incidence functions estimated gvhd/ nrm.results: thirty-two patients received at least one dose of leuprolide. median age was years (range - ). twenty-six( %) had acute leukemia, ( %) mds/mpn, and ( %) cml. all but one had an / -matched donor. at median follow-up of months among survivors (range - ), estimated y os was %( %ci - %) and rfs %( %ci - %). ci of trm at y was %( %ci - %). ci of grade iii-iv acute gvhd was %( %ci . - %). of patients with evaluable flow data, % achieved a cd + count >= by +/- days, not significantly different from historical controls. median lymphocyte counts at +/- days did not differ significantly among groups (table; figure) .conclusions: this phase ii study did not demonstrate significant quantitative improvement in immune reconstitution after leuprolide with palifermin in recipients of tbibased cd -selected hct. tcr sequencing to identify possible improvement in t-cell diversity in this cohort is forthcoming. one potential explanation for these results lies in the initial surge in sex steroid levels immediately after background: the prospective, multinational, noninterventional emmos study aimed to document, and describe real-world treatment regimens and disease progression in patients with mm at different stages of the disease.methods: adult patients initiating any new mm therapy between and were eligible. a multi-staged patient/site recruitment model was applied to minimize selection bias, and enrolment was stratified by country, region, and practice type. patients' medical/disease features, treatment history and remission status were recorded at baseline, and prospective data on treatment, efficacy and safety were collected electronically every months until years after last enrolment. responses were investigatorassessed. overall findings from emmos were previously reported. here, we are presenting additional analyses focusing on the induction regimens used in the subgroup of patients who proceeded to auto-sct frontline.results: a total of patients ( with stem-cell transplant [sct] and without) were enrolled. patient demographics/baseline characteristics were as expected. of recipients of sct after enrolment, ( %) underwent auto-sct frontline. % of the auto-sct patients were aged ≤ years. among these frontline auto-sct patients, the majority had a single transplant ( %). the most frequent induction regimen was bortezomibthalidomide-dexamethasone (vtd; n= ; %), bortezomib-dexamethasone (vd; n= ; %), bortezomib-cyclophosphamide-dexamethasone (vcd; n= ; %), doxorubicin-bortezomib-dexamethasone (pad; n= ; %), and cyclophosphamidedexamethasone-thalidomide (ctd; n= ; %). only % of patients received a bortezomib-lenalidomidedexamethasone (vrd) induction regimen, while lenalidomide was shown to be the most frequently used agent in lines and at time of relapse. in the vtd subgroup, most patients received mg thalidomide dose during induction. the majority of administration schedule was based on days cycles, while a few other schedules were seen corresponding to days cycles or to delay due to adverse events or other specific reasons. the most prevalent number of vtd cycles was and ( % and %, respectively). lower or higher number of cycles was only marginal ( % and %, respectively). out of the patients with vtd induction, % achieved cr as best response, % ncr or vgpr and % pr. after auto-sct, the best overall response rates (orr) at any time during frontline therapy were > % for those patients whose treatment included university medicine greifswald, greifswald, germany background: we explored the effect of dinutuximab beta (ch . /cho) on outcome within the hr-nbl /siopen trial population by comparing an era prior immunotherapy availability.methods: the analysis cohort consists of the immunotherapy population (ip) ( - ) and a matched control population (cp) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) . all patients had rapid cojec induction, up to two tvd courses and high-dose chemotherapy (bumel or cem) followed by autologous stem cell reinfusion (hdc/ascr) within months since diagnosis; local control included surgery and local radiotherapy ( gy) followed by maintenance with six cycles of isotretinoin. ip patients had additional five cycles of dinutuximab beta short-term infusions with or without subcutaneous interleukin- . cp patients had to be part of the hdc randomization. the median time between ascr and initiation of immunotherapy was days in the ip. only patients without progressive disease at this landmark time point were included in the cp. median follow-up was . years (iqr: . to . years) for eligible patients.results: the y-efs of the ip ( patients) was % ± % compared to %± % for the cp ( patients; p< . ). both populations were balanced for sex, age, stage , mycn amplification and response prior hdc. multivariate analysis showed an independent higher risk for the cp (p= . , hr . ), for cem (p= . ; hr . ), a response < cr prior to maintenance therapy (p= . , hr . ) and for > metastatic compartment at diagnosis (p< . , hr . ). after adjustment for risk factors, the benefit of immunotherapy was confirmed in bumel-(p= . ; hr . ) and in cem-treated patients (p= . ; hr . ).conclusions: results suggest a patient benefit from dinutuximab beta based immunotherapy with or without il within the hr-nbl trial.clinical trial registry: the trial was registered with clinicaltrials.gov (number nct ) and eudract (number - - ) . https://www.siopen-r-net.org disclosure: the academic data supported apeiron to obtain the dinutuximab beta product licensure in may in the european union (ema). siopen and ccri had an agreement in place with apeiron regarding the provision of academic data. ruth ladenstein and holger lode acted as consultants for apeiron on behalf of siopen for the ch . /cho development.the other authors declared no conflicts of interest. out of ( %) received serotherapy with alemtuzumab (n= ) or atg (n= ) before viral reactivation. where possible immunosuppression was withdrawn in combination to bcv therapy. viral load was detected in blood by pcr as copies/ml. data on response to bcv was divided into complete response (cr) with undetectable virus in blood and resolution of symptoms, partial response (pr) with at least log drop in viral load after bcv, no response (nr) with no change in the viral load and stationary disease and progressive disease (pd) with evidence of at least a log rise in viral load by pcr or organ disease progression.results: the median viral load at the start of bcv was . million copies/ml (range: - million copies/ml). bcv was used as a first line treatment in cases and as second line in cases after failure of first line therapy (n= ), toxicity (n= ) or both(n= ). / ( %) patients had evidence of viral induced organ disease at time of bcv administration; adv disease (encephalitis, pneumonitis, hepatitis and colitis), ganciclovir resistant cmv retinitis and bk haemorrhagic cystitis. / ( %) patients achieved either a cr (n= ) or pr (n= ) and / ( %) patients with organ disease achieved a cr.two patients with adv disease and pr received donor derived cytotoxic t lymphocytes to achieve cr. at a median follow-up of months (range: - . ), patients who were in cr or pr did not show any evidence of viral reactivation after bcv discontinuation despite no evidence of immune reconstitution (ir). four patients had evidence of disease progression with significant rise in viral load while on bcv therapy and all died. the patient with ganciclovir resistant cmv attained cr of cmv retinitis.among patients with adv viraemia ± disease; ( %) achieved either cr (n= ) or pr (n= ). nine cases had concomitant bk± cystitis at the time of bcv therapy and all had nr. toxicity was observed in / cases; renal impairment (n= ), transaminitis (n= ) and diarrhoea (n= ). median cd , cd and cd were persistently low both pre-bcv and at the end of treatment; cells/ul vs cells/ul, cells/ul vs cells/ul and cells/ul vs cells/ul; p= . , p= . , p= . respectively (figure ). at last follow-up, / ( %) were alive. of these, viral infection related mortality was / ( %).conclusions: bcv is an effective and well tolerated treatment in immune compromised patients with adv infection with a response rate of %.[[o image] . background: although the impact of donor graft composition on clinical outcome after hematopoietic stem cell transplantation (hsct) has been studied, little is known about the role of intra-graft γδ t-cell receptor (tcr) repertoire on clinical outcome following hsct. using high-throughput sequencing we sought to analyze the tcr γ-chain (trg) repertoire of γδ t-cells within donor stem cell grafts and address its potential impact on clinical response in the corresponding patients.methods: we analyzed twenty peripheral blood stem cell grafts from matched unrelated donors and classified as cmv-positive/negative. the respective acute myeloid leukemia recipients were followed for disease relapse and acute graft-vs-host disease (agvhd) development post-hsct. γδ t-cells were isolated using magnetic beads and the gdna extracted for next-generation sequencing (immunoseq, adaptive biotechnologies). trg characteristics were assessed using vdjtools, vdjviz, tcr and immunoseq analyzer platforms.results: deep sequencing showed similar median total/ unique reads in all grafts as well as similarly low unique cdr trg ratios. grafts presented multiple clonal overrepresentations, with no differences on tcr richness between patient groups. grafts received by the non-background: prognosis of mature t-and nk-cell lymphomas remains poor despite development of novel therapeutic agents. accordingly, these lymphomas are still good candidates of allogenic hematopoietic stem cell transplantation (allo-hsct) to achieve long-lasting remission of the diseases. however, the analysis of transplantations for these lymphomas is scarce, mainly due to the rarity of these lymphomas. hence, we analysed the data of these transplantations operated in different japanese institutions as a multi-institutional joint research and examined factors that affected the outcomes in aims of figuring out better transplant strategies against these lymphomas.methods: a total of patients who received allo-hscts for mature t-and nk-cell lymphomas (ptcl-nos, n= ; nk/t cell lymphoma nasal type, n= ; aitl, n= ; alcl, n= ; eatl, n= ; other lymphomas, n= ) from to in institutions were examined. median age at transplantation was (range, - ). forty patients received transplantation from related bone marrow transplantation (bmt) / peripheral blood stem cell background: children, adolescents and young adult patients with all with second relapse, relapse after allogeneic sct or patients with primary refractory disease have a poor prognosis with conventional treatment concepts. in this patient group several studies using second generation cd chimeric antigen receptor t-cells (car-t cells) demonstrated high efficacy with two year survival rates of up to %. recently, two different car-t cell products were approved by the fda and in also by the ema in europe: tisagenlecleucel (kymriah®) for the treatment of patients with b-precursor all who are i) refractory, ii) in second relapse or iii) who relapsed after allogeneic sct (relapsed/refractory all; r/r all) as well as for diffuse large cell lymphoma (dlbcl) and axicabtagen ciloleucel (yescarta®) for the treatment of b-cell lymphoma. here we report our first results using commercially available car-t-cell product tisagenlecleucel (kymriah®) in patients with all which were treated by the university hospital for children and adolescents frankfurt am main (n= ), the department of medicine iii, university hospital lmu munich (n= ), and the von hauner kinderspital, lmu munich, germany (n= ).methods: between october and december eleven patients received apheresis for car-t cell generation. nine patients suffered from r/r c-all, and two from r/r bprecursor all. eight patients had relapsed after allogeneic hsct, one patient each suffered from first r/relapse, second r/relapse or from primary r/all. in / ( %) patients car-t cell production was successful after one and in patients ( %) after a second apheresis.median patients' age was . years ( . - . ). between apheresis and start of lymphodepleting chemotherapy (ldc), / patients received low dose chemotherapy according to the frapostall protocol (willasch et al. ) and one patient was treated with inotuzumab. production slots were immediately available, resulting in turn-around-time from apheresis to product delivery of - weeks. disease status at start of ldc was cr w/o mrd (n= ), cr mrd pos. (n= ), cri (n= ), persistence of blasts (n= ), and disease progression ( - % blasts, n= ). ldc consisting of flu-cyc was given to / patients, one patient did not receive ldc.results: car-t cells could be transfused to / patients at a median dose of . mio/kgbw ( . mio . ). in one patient, in whom a second viral transduction procedure was necessary; neither ldc nor car-t cell transfusion could be given because of diseases progression and deterioration of the patient's general condition. cytokine release syndrome (crs) grade i was observed in one patient; / patients did not develop crs. cytokine related encephalopathy syndrome (cres) grade ii was observed in / patients. at day + / patients ( %) achieved mrd negative cr. the two patients with key: cord- - gux gsi authors: nan title: physicians abstracts date: - - journal: bone marrow transplant doi: . /bmt. . sha: doc_id: cord_uid: gux gsi nan introduction: the incidence of cgvhd after allogeneic sct is higher when peripheral blood stem cells are used as stem cell source. there is a strong need for preventing cgvhd after asct without increasing the risk of relapse. materials (or patients) and methods: we performed a multicenter, multinational, open-label, randomized study comparing anti-lymphocyte globulin (atg-fresenius s ) mg/kg on day - , - and - with no atg in patients with aml (n ¼ ) or all (n ¼ ) in st complete remission (cr; n ¼ ) or nd cr (n ¼ ) who received peripheral blood stem cells from their hla-identical sibling after standard tbi ( gy)/ccclophosphamide ( mg/kg) or busulfan ( mg/ kg)/cy ( mg/kg) based myeloablative conditioning regimen. standard gvhd prophylaxis consisted of cyclosporine a and a short course of mtx ( mg/m on day þ , þ , þ and þ ). the primary study aim was to compare the cumulative incidence of cgvhd at years after asct. results: out of randomized patients from centers and nations were withdrawn before conditioning and asct due to leukemia progression, or cancellation of the donor. patients were analyzed for safety and efficacy; were randomized to atg and to non-atg. the treatment groups were comparable regarding recipient and donor age and sex, cmv serostatus, disease (aml vs all), st or nd cr. the median time to leukocyte ( . x e /l) and platelet ( x e /l) engraftment was significantly delayed in the atg group ( vs. days, po . and vs days, po . ). the incidence of acute gvhd grade i-iv was % for the atg arm and % for the non-atg arm (p ¼ . ) and for severe grade iii/iv acute gvhd % and %, respectively (p ¼ . ). the cumulative incidence of cgvhd at years was % ( % ci - %) in the atg and % ( % ci - %) in the non-atg arm (po . ). regarding limited and extensive cgvhd the ci at years was % vs % (p ¼ . ) and % vs % (po . ), respectively. there was no difference in infectious complications, cmv and ebv reactivation between both arms. the cumulative incidence of therapy related mortality at years was % ( % ci - %) for the atg arm and % ( % ci - %) for the non-atg arm (p ¼ . ), resulting in year relapse-free and overall survival of % ( %ci - %) and % ( % ci - %) for the atg group and of % ( % ci - %) and % ( % ci - %) for the non-atg group (p ¼ . and p ¼ . , respectively). chronic gvhd free survival at years was % for the atg and % for the non atg arm (po . ). a composite endpoint cgvhd and relapse-free survival at years was in favor for the atg treated patients ( % vs. %, p ¼ . ). conclusion: this is the first randomized cgvhd prevention study providing evidence that atg-fresenius s ( x mg/kg) is highly effective in preventing limited and extensive cgvhd in hla-identical sibling pbsc transplantation when used within a myeloablative preparative conditioning regimen. the use of atg-fresenius did not result in an obvious increase of infectious complications and relapse, resulting in similar overall survival rates, but improved cgvhd/relapse free survival. introduction: the establishment of large transplant registries and introduction of novel statistical techniques have paved the way for large scale data analysis. nevertheless, contemporary tools for risk prediction of transplant related mortality (trm) following allogeneic (allo) hematopoietic stem cell transplantation (hct) are of limited clinical use, owing to a sub-optimal predictive accuracy. apart from inherent procedural uncertainty, methodological factors impeding prediction might be attributed to the statistical methodology used, number and quality of features collected, or simply the population size. using an in-silico approach (i.e. iterative computerized simulations), based on machine learning (ml) algorithms, we aimed to define prediction limiting factors of day trm and rank variable contribution. ml is a field of artificial intelligence dealing with the construction and study of systems that can learn from data, rather than follow explicitly programmed instructions. commonly applied in complex data scenarios, such as financial settings, it may be suitable for outcome prediction of hct. materials (or patients) and methods: study population was a cohort of , adult acute leukemia allo-hct recipients from the ebmt-alwp. twenty four variables were analyzed, including recipient, donor and transplant characteristics. study design involved two phases. the first, focused on development and comparison of several ml based prediction models of day trm. in the second phase, a repetitive computerized simulation was applied. factors necessary for optimal prediction were explored: algorithm type, size of data set, number of included variables, and performance in specific subpopulations. models were assessed and compared on the basis of the area under the receiver operating characteristic curve (auc). results: six ml based prediction models for day trm were developed on the entire dataset. optimal aucs ranged from . - . . depending on the algorithm used for prediction model development, the in-silico experimental system yielded the following results: predictive performance plateaued on a population size ranging from n ¼ - ; a range of - ranked variables, selected by a separate feature selection algorithm, were necessary for optimal prediction; disease status and donor type were consistently top ranking variables. predictive performance of models developed for specific subpopulations, ranged from . to . for patients in second complete remission and patients receiving reduced intensity conditioning respectively. conclusion: we present a novel experimental system for assessment of prediction boundaries in hct. the present approach has clinical implications. we show that predictive performance of day trm is unlikely to improve with the data routinely gathered on hct recipients, as different algorithms reach approximately the same performance. in addition, an exhaustive search for variable importance, reveal that few variables "carry the weight" with regard to predictive influence. predictive performance converged when sampling more than patients, reflecting the importance of large registry studies. overall, it seems we have reached a point of predictive saturation. improving predictive performance will likely require additional types of input like genetic, biologic and procedural factors. disclosure of interest: none declared. introduction: allogeneic stem cell transplantation (sct) can provide long-term disease control in selected patients with relapsed refractory nhl. restricted availability of a matched sibling donor limits its use especially for patients with rapidly progressing disease in whom unrelated donor search cannot be awaited. because data of alternative donor transplants in nhl is sparse, we aimed to compare outcome of haploidentical and cord blood transplants with conventional relatedand matched unrelated donor transplats for nhl. materials (or patients) and methods: information of patients with mantle cell lymphoma (mcl), dlbcl, t-cell lymphoma (tcl) and follicular lymphoma (fl) who received an sct from a sibling donor (sib), / matched unrelated donor (mud), haplo-identical donor (haplo) or cord blood (cord) between and was downloaded from the ebmt database. results: patients with nhl were identified in the ebmt database meeting the inclusion criteria. received a transplant from a sib, from a mud, from cord ( mcl, dlbcl, fl, tcl) and from a haplo donor ( mcl, dlbcl, fl, tcl) . active disease (po . ) and karnofsky index (ki) below % was also more common in the haplo group (p ¼ . ). other variables were balanced. median follow-up after sct for all patients was months (ci to ). relapse incidence after conventional transplants (sib, mud) and alternative donor transplants (haplo, cord) was not significantly different within the whole group (haplo: hr . % ci . - . p ¼ . ; cord: hr . % ci . - . , p ¼ . ) and across all studied disease entities. in contrast, nrm incidence was not significantly different between sib and mud, but significantly higher with alternative donor transplants (haplo: hr . % ci po . ; cord: hr . % ci po . ) . with the exception of fl where mud in addition to haplo and cord transplants had a significantly higher nrm incidence than sib transplants. because patients who received a haplo transplant had more commonly active disease at transplant and worse ki, we performed multivariate modeling of relapse-and nrm incidences adjusting for the above mentioned covariates. no different relapse incidences between donor groups was observed. nrm incidence in contrast, was significantly higher in mud (reference sib, hr , p ¼ . ) and cord (reference sib, hr , p ¼ . ) but not in haplo transplants. most interestingly, acute gvhd incidence was significantly increased in mud compared to sib (p ¼ . ) transplants but not in haplo (p ¼ . ) or cord (p ¼ . ) transplants. multivariate adjustment for diagnosis (mcl, dlbcl, fl, tcl), remission prior to sct, ki (kio % vs. %) and conditioning intensity (ric vs. mac) did not reveal worse os for haplo but a worse os for cord (hr po . introduction: retrospective studies in mds/saml suggest that reducing the intensity of the conditioning regimen prior to allogeneic stem cell transplantation reduces the risk of nonrelapse mortality but is associated with a higher risk of relapse, but prospective randomized studies for mds are lacking so far. materials (or patients) and methods: within the chronic malignancies working party (cmwp) of ebmt, we performed a prospective randomized trial comparing a busulfan based (busulfan mg/kg orally or equivalent dosis intravenously (iv) plus fludarabin mg/m ) reduced intensity conditioning regimen (ric) and a standard myeloablative busulfan (busulfan mg/kg orally or equivalent dosis iv plus cyclophosphamide mg/kg) based regimen (mac) in patients with mds or saml (o % blasts). between may and december , patients were included from centers and nations and could be analysed. major inclusion criteria were: mds (according to fab: ra, rars, raeb, raeb-t), cmml, and saml, blasts less than %, matched related or unrelated donor (hla / , mismatch was allowed), age - years (for unrelated) and - years (for hla-identical sibling). included patients were stratified according related vs unrelated donor and blast countoor than %. the primary endpoint of the study was year non-relapse mortality.the median age of the patients was . years (r. - y) . donors were hla-identical sibling (n ¼ ), matched unrelated (n ¼ ) and mismatched related or unrelated (n ¼ ). the patients were well distributed in both arms regarding age, gender, ipss risk profile, number of blasts at transplantation, donor source and mismatch donor. results: graft failure occurred in after mac and after ric. median time to leukocyte ( . x e /l) occurred after days after mac and days after ric and and platelet ( x e /l) engraftment occurred after days after mac and days after ric, respectively. acute gvhd ii-iv was noted in % after ric and % after mac. chronic gvhd was seen in % after ric and % after mac. the cumulative incidence (ci) of non-relapse mortality (nrm) after year was % ( % including pediatric/adolescent cases o years), with myeloablative conditioning ( %; tbi and non-tbibased), from hla-identical siblings ( %), and using ex vivo t-cell depletion. in addition to the above matching criteria, cases and controls were also balanced for other factors such as donor gender and gender mismatch, cmv serostatus, in vivo t-cell depletion and karnofsky's performance status. compared to hiv-neg, hiv-pts had lower rates of neutrophil engraftment ( . % vs . %, p ¼ . ), higher incidence of grade iii-iv acute gvhd ( % vs %, p ¼ . ), higher nrm at day ( % vs %, p ¼ . ) and years ( % vs %, p ¼ . ), and similar incidence of relapse ( % vs %, p ¼ . ). overall, hiv-pts had poorer pfs ( % vs %; p ¼ . ; hr . [ . - . ] ) and os ( % vs %; p ¼ . ; hr . [ . - . ]) at years than hiv-neg cases.outcomes within hiv-pts were comparable for myeloablative vs reduced intensity conditioning and among peripheral blood, bone marrow and cord blood stem cell sources (data not shown). finally, while hiv-pts' outcomes were comparable for allo-hct from hla-identical siblings and alternative donors (os: % vs %; p ¼ . ) , the use of such alternative donors in hiv-pts was less common than in hiv-neg hct recipients ( % vs %, p ¼ . ). conclusion: this study showed that the outcome of allo-hct is poorer in hiv-pts than in the general population, primarily driven by higher nrm, and in keeping with their inferior overall life expectancy despite haart. even so, allo-hct is feasible in hiv-pts with hematologic indications, with a % os at years. in view of the current reduced use of alternative donors despite comparable results to matched sibling donors, hiv-pts requiring an allo-hct should be granted access to donor search and consideration for transplantation at the same level as hiv-neg counterparts. these data are key to inform allo-hct strategies in hiv-pts, including its investigational use to eradicate hiv infection. disclosure of interest: none declared. introduction: haplo-hsct is a therapeutic option for patients with high risk hematologic neoplasms with the advantages of quick availability, easy programation and logistics, and a committed donor. it has shown promissing results in patients diagnosed with relapsed or refractory hodgkin lymphoma (hl) at least comparable to allogeneic transplant from siblings or unrelated donors (burroughs lm et al. biol blood marrow transplant ; : - . materials (or patients) and methods: we retrospectively evaluate the results of haplo-hsct with iv busulfan (bux) based ric regimens (fludarabine mg/m x days (- to - ), cyclophosphamide , mg/kg x days (- to - ) , bux , mg/ kg x (bux )or days (bux ) on days - to - ) and gvhd prophylaxis based on ptcy ( mg/kg on days þ and þ ) and a calcineurin inhibitor plus mycophenolate from day þ performed in geth centers to patients diagnosed with relapsed or refractory hl. results: from march- , haplo-hsct have been performed in patients diagnosed with relapsed or refractory hl in geth centers. median age was years ( - ), % were males and all were in advanced phases of their disease, after a median of prior treatment lines ( ) ( ) ( ) ( ) ( ) ( ) ( ) . autologous hsct was previously employed in %, and allogeneic hsct in %. five patients ( . %) have received more than prior transplants. disease status at haplo-hsct evaluated by pet was complete remission in ( %) and persistent disease in ( %). bone marrow was employed in ( %) and peripheral blood in ( %), without t-cell depletion in all cases. the haploidentical donor was patients mother ( ), father ( ), siblings ( ) or daughter ( ) . the ric regimens employed were bux in ( . %) and bux in patients ( . %). median neutrophils engraftment was day þ ( - ) and platelets k was day þ ( - ). graft failure with autologous reconstitution happened only in patient ( . %). the day þ cumulative incidence (ci) of non-relapse mortality (nrm) was % ( / ) and % ( / ) at year posttransplant. the day þ ci of grade ii-iv acute gvhd was %, and grade iii-iv was . %. chronic gvhd ci was . % at year, being extensive in %. after a median follow-up for survivors of months , the event-free survival (efs) was . % and overall survival (os) was %. the -year ci of relapse or progression was %. factors related with better year efs were cr prior to haplo-hsct ( % vs %; p ¼ . ) and receiving less than treatment lines prior to haplo-hsct ( % vs . %; p ¼ . ). no significant differences were seen when comparing bux against bux in terms of nrm, efs or os. conclusion: haplo-hsct with ptcy and bux based ric conditioning in relapsed or refractory hl patients, renders long-lasting remissions with acceptable toxicity and gvhd, obtaining better results in those transplanted in cr and with less than treatment lines prior to haplo-hsct. disclosure of interest: none declared. introduction: b-lineage acute lymphoblastic leukemia (all) is the most common childhood cancer. although this disease can be successfully treated in % of patients, prognosis for primary refractory or relapsed disease is very poor. even after allogeneic stem cell transplantation (sct), relapse rates are considerable and correlate significantly with persistent minimal residual disease (mrd) prior to and after sct. mrd constellations represent favorable effector-target ratios and thus might be optimally suited for immunotherapeutic intervention with therapeutic antibodies. materials (or patients) and methods: we developed an fcoptimized cd antibody ( g sdie) and produced it in pharmaceutical quality. g sdie mediates markedly enhanced antibody-dependent cellular cytotoxicity (adcc) through its improved capability to recruit fcgriiia bearing effector cells including nk cells and gd t cells. g sdie was applied on compassionate use to mrd-positive pediatric patients with relapsed or refractory all (cr n ¼ , zcr , n ¼ ). results: side effects such as headache and fever were negligible. in all patients complete cd þ b-cell depletion was observed during therapy. after discontinuation of g sdie therapy b cell counts recovered rapidly to normal levels. in / patients mrd was reduced byz log or fell below mrd-detection threshold of - over the course of treatment. / responders were receiving additional treatment. patients relapsed, patient died of cns chemotherapy associated toxicity and patient died of late posttransplant sepsis. patients have been in sustained remission for - days (median follow-up days). in initial cytotoxicity screenings, performed in an allogeneic setting, significantly increased lysis of all blasts by pbmc after adding g sdie was observed. nk cells and gd t cells were identified as main effector cell populations. cd expression on patient blasts was confirmed by quantitative flow cytometry (mean . x molecules/ cell, ± . x ). cytotoxicity assays using patient pbmc on autologous blasts confirmed sustained functionality of patient effector cells over the course of g sdie treatment. cytotoxicity assays were performed using pbmc from transplanted patients obtained at different time points of g sdie treatment. lysis of autologous all blasts was increased when g sdie or autologous patient serum taken after antibody infusion was added. after infusion of mg/m - mg/m g sdie serum half-life was h - h. serum levels of g sdie remained above saturating concentrations ofz ng/ml (ec ¼ ng/ml) until the following application in the bi-weekly treatment cycle. notably, in / analyzed patients under g sdie therapy, a down-modulation of cd surface expression on the leukemic blasts was observed. in vitro antigenic shift assays on patient blasts showed considerable but very heterogeneous shift of cd surface expression. furthermore, a positive correlation between cd surface expression levels and g sdie mediated lysis was observed. these observations hint at in vivo tumor escape mechanisms and moreover indicate selective pressure exerted by immunotherapy with g sdie, underlining its therapeutic potential, but also delineating possible limitations. conclusion: promising anti-leukemic effects of the g sdie antibody have been observed in vitro and in vivo. we are currently preparing a clinical phase i/ii trial. disclosure of interest: none declared. introduction: allogeneic hct with myeloablative conditioning is considered a standard of care for adults with high risk acute lymphoblastic leukemia (all). however, with improving results of conventional-dose chemotherapy and the introduction of novel agents on one hand, and the improvement in transplantation techniques on the other, the indications for allohct require re-evaluation, taking into account patient-and procedure-related factors associated with the risk of nonrelapse mortality (nrm). the aim of this study was to analyze the results of myeloablative allohct treatment for patients with all according to recipient age and donor type. materials (or patients) and methods: patients treated with allohct in first complete remission during the period - were included. the outcomes were analyzed for the periods - , - and - , in various age groups, separately for hla matched sibling (msd, n ¼ ) and unrelated donors (urd, n ¼ ). results: for msd-allohct recipients treated during the period - , the following two-year probabilities of os were obtained: % for the - years old (y.o.) group, % for both the - y.o. and - y.o. groups and % for the - y.o. group. the incidence rates of nrm were %, %, % and %, respectively for those same age groups. in comparison with the - period, significant improvements were observed for all age groups except for the - y.o. patients. the improved survival rates were a consequence of reduced nrm and a tendency towards a reduced risk of relapse. among urd-allohct recipients, the os was % ( - y.o.) , % ( - y.o.) , % ( - y.o.) , and % ( - y.o.) , while the respective incidence rates of nrm were %, %, % and %, the improvement of os over time was documented for - y.o. (p ¼ . ) and - y.o. ( . ) patients due to the reduced incidence of nrm with no significant effect on relapse. in a multivariate analysis adjusted for disease-, patient-, donorand procedure-related factors, transplantations performed for the period - , when compared to - , were associated with significantly reduced risks of the overall mortality (hr ¼ . , p ¼ . ), treatment failure (either relapse or nrm, hr ¼ . , p ¼ . ), and nrm (hr ¼ . , p ¼ . ) and showed a trend towards reduced risk of relapse (hr ¼ . , p ¼ . ). the overall mortality was reduced for transplants with tbi-based compared to chemotherapy-based conditioning (hr ¼ . , p ¼ . ) as a result of reduced risk of relapse (hr ¼ . , p ¼ . ). type of donor (msd vs. mud) had no significant effect on survival (hr ¼ . , p ¼ . ). conclusion: results of allohct for adults with all improved significantly over time in all age groups, mainly due to the reduction of nrm. importantly, results obtained with matched unrelated transplants were comparable to sibling transplants. total body irradiation should still be considered as the preferable type of myeloablative conditioning for all. disclosure of interest: none declared. cmv seronegativity is associated with a % decrease in -year survival in patients undergoing reduced intensity sibling-donor transplantation for treatment of myeloid malignancy d. j. lewis ,* , c. holmes , k. peggs , a. peniket , m. nikolousis , s. nagra , g. pratt , , c. craddock , , r. malladi , , p. moss , school of cancer sciences, university of birmingham, birmingham, university college hospital, london, oxford university hospital, oxford, birmingham heartlands hospital, centre for clinical haematology, university hospital birmingham, birmingham, united kingdom introduction: reduced intensity (ric) allogeneic stem cell transplantation is a highly effective treatment for acute myeloid leukaemia and the immunological 'graft versus leukaemia' effect is believed to be a major mechanism of disease control. cytomegalovirus reactivation has been suggested to reduce the rate of disease relapse in acute myeloid leukaemia (aml). we investigated the influence of cmv serostatus on the clinical outcome of patients who underwent t cell depleted ric allografts for myeloid malignancy, and went on to examine reconstitution of lymphoid subsets. materials (or patients) and methods: we studied patients who underwent ric allografts for aml (n ¼ ) and mds (n ¼ ) from four uk centres, with fludarabine and melphalan conditioning þ /-alemtuzumab. overall survival was calculated. relapse rate and non-relapse mortality were calculated with competing risk analyses. results: the median overall survival was . years. the relapse rate of the entire cohort was % at year with a nonrelapse mortality of %. the overall survival for the 'cmv at risk' group was . % at year compared to . % for cmv (-/-). this difference was most marked in patients transplanted from sibling donors (n ¼ ); the overall survival at year was % in cmv-at-risk patients (n ¼ ) compared to only % in the cmv seronegative group (n ¼ ) (p ¼ . ). this % increment in survival was due to a % reduction in the rate of disease relapse in patients that were cmv-at-risk ( year relapse rate of % versus %). there was a non-significant trend towards improved overall survival in those that experienced cmv reactivation amongst the cmv-seropositive patients ( year os % versus %, p ¼ . ) , mainly due to a reduction in the rate of relapse ( year relapse rate % versus %). because of this large difference in relapse risk, we went on to examine the effects of cmv serostatus and alemtuzumab use on reconstitution of lymphocyte subsets at months post transplant. alemtuzumab led to -fold decrease in the t cell count at months compared to transplants in which t cell depletion was not used. however, within this alemtuzumab group, positive cmv serostatus in the donor or recipient was associated with a relative fold and fold increase in the cd þ and cd þ t cell count compared to cmv seronegative pairs. indeed, cd þ t cells were virtually undetectable at this time point in cmv seronegative transplant/donor pairs. conclusion: cmv seropositivity is markedly beneficial for patients who undergo a sibling donor reduced intensity allograft for myeloid malignancy and in whom alemtuzumab is used for conditioning. this effect is most likely to be due to the profound influence of chronic viral replication on boosting t cell immune reconstitution in the early post transplant period. cmv seronegative patients with aml should be considered at risk of impaired survival for certain subgroups of stem cell transplant. disclosure of interest: none declared. introduction: allogeneic stem cell transplantation is the only curative option for patients with high risk acute myeloid leukemia (aml) and for those experiencing relapse. either matched sibling donor (msd) or unrelated donor (ud) is indicated. materials (or patients) and methods: with the aim to compare the outcomes of both strategies we have retrospectively analysed adults with aml receiving either msd (n ¼ ) or ud (n ¼ ) in ebmt centers from - . for ud, were / hla matched and were / . median follow up was (range - ) months. there were statistical differences between the groups. compared to msd recipients, ud transplants were older ( vs years, p ¼ . ), were performed more recently ( vs , p ¼ . ) , had longer interval between diagnosis to transplant ( vs months, p ¼ . ), had more often secondary aml ( % vs %, p ¼ . ) and were transplanted with higher proportion of cmv negative donors ( % vs %, p ¼ . ). peripheral blood stem cells (pbsc) was used as graft source in % of patients in both groups, p ¼ . . conditioning regimen was more frequently myeloablative for patients transplanted with a msd ( % vs %, p ¼ . ). msd received more often busulfan and cyclophosphamide as mac ( %) or a tbi based regimen ( %). for ud, bu-fludarabine was the most frequent conditioning used ( %). results: cumulative incidence (ci) of neutrophil engraftment was similar ( % vs % for msd vs ud, respectively, p ¼ . ). grade ii-iv acute gvhd was % vs % (p ¼ . ) and ci of chronic gvhd was % vs % (p ¼ . ) for msd and ud, respectively. for msd and ud respectively, ci of relapse (ri) was % vs %, p ¼ . ; ci of non-relapse mortality (nrm) was % vs %, p ¼ . . probability of leukemia-free survival (lfs) at years was % vs %, p ¼ . , and overall survival (os) was % vs % p ¼ . , respectively. chronic gvhd as time-dependent variable was associated with lower ri (hr . , p ¼ . ), higher nrm (hr . , p ¼ . ), and higher os (hr . , p ¼ . ). according to hla-match for msd, / ud and / ud, ci of relapse (ri) was % vs % vs %%, p ¼ . ; ci nrm was % vs % vs %%, p ¼ . and probability of lfs at years was % vs % vs %, p ¼ . , respectively. in a multivariate analysis adjusted for differences between the groups, ud was associated with lower ri (hr . , p ¼ . ) and higher lfs (hr . , p ¼ . ) compared to msd. when analyzing according to hla-match, there was no differences for patients transplanted with an ud / or a / for ri (hr . , p ¼ . ) , and lfs (hr . , p ¼ . ) . the other factors independently associated with better outcomes were the interval between diagnosis and transplant (ri hr . , po . ) , and lfs (hr . , po . ) . conclusion: unrelated donor transplant was associated with better lfs due to lower ri compared to msd for high-risk patients with aml transplanted in first relapse. there were not differences for the ud / match probably due the graftversus-leukemia effect in the setting of patients transplanted with active disease. disclosure of interest: none declared. introduction: allogeneic reduced intensity transplantations (rict) in elderly patients (pts) with aml in cr has become a commonly used treatment modality. however, no prospective studies to date have been reported to support use of this strategy. the aim of this prospective, multicenter study is to compare outcomes of patients receiving rict with patients being treated with conventional chemotherapy. in an amendment allowed also the use of matched unrelated donors. the study is currently ongoing in countries with a total of pts included. only pts included as per the original protocol are accounted for in this interim analysis which focuses on safety. materials (or patients) and methods: the study was designed by the transatlantic leukemia group (tralg) consisting of centers associated with the swedish and canadian bmt groups, and centers from germany, norway, finland and new zealand. patients should have intermediate or high risk aml in cr and not being eligible for a myeloablative transplant due to age or comorbidities, and have at least one potential sibling donor. date of inclusion was defined as the date of hla-typing of the first sibling, and pts were allocated to the donor (d) or no-donor group (nod) based on hla match. overall survival at yrs is the primary endpoint; secondary endpoints include rfs, gvhd, non-relapse (nrm) and transplant related (trm) mortality. conditioning consisted in the vast majority of patients of fludarabine mg/m iv and busulphane mg/ kg po or . mg/kg iv. gvhd prophylaxis was cya/methotrexate or cya/mmf. the study started to accrue patients in . results were analyzed on an intent to treat basis. results: from pts were included in sweden (n ¼ ), canada (n ¼ ), germany ( ) , norway ( ), new zealand ( ) and finland ( ) . patients were excluded from analysis; favorable risk, protocol violations and pts; females, males, median age ( - ) yrs, saml, were allocated to the d group (n ¼ ), or the nod group (n ¼ ). in the d group, pts ( %) did not reach transplant, of these died ( after relapse, other causes). cytogenetic risk groups were categorized as per eln criteria. the distribution of total disease risk categories was % adverse, % intermediate, % unknown. median follow-up of surviving pts at dec th was . ( . - . ) yrs after inclusion. ninety-one pts relapsed ( %). kaplan-meier estimates of os and rfs in the whole group at yrs were % and % respectively. for adverse and intermediate risk pts os and rfs at yrs were % and %, and % and %, respectively. in the transplanted pts acute gvhd grade - occurred in % of pts, grade - in %. extensive gvhd occurred in %, limited in %. pts ( %) have died, without and after aml relapse. in the nod group ( %) pts died from non-relapse causes. in the d group, pts died before transplant, while the trm was / ( %) with causes of death: gvhd, other. conclusion: selected patients with aml in cr tolerate rict or standard management with low mortality. disease control remains a major issue. this multicenter prospective protocol will continue to accrue patients until relevant conclusions can be drawn comparing a rict to standard treatment. the current rapid inclusion of pts and participation of new sites in australia, greece and estonia will help to complete the study. disclosure of interest: none declared. introduction: the wilms' tumor gene (wt ) is overexpressed in % of acute myeloid leukemias (aml) and myelodysplastic syndromes (mds) and proved to be a good marker for minimal residual disease (mrd) monitoring. although allogeneic haematopoietic stem cell transplantation (allo-hsct) is the most effective treatment for aml/mds, disease recurrence remains a major problem. after allo-hsct, quantitative wt monitoring can represent a useful tool to detect mrd and tailor immunotherapeutic strategies. materials (or patients) and methods: we included in this retrospective analysis pts with aml/mds ( and respectively) overexpressing wt and allotransplanted in our institution from / to / . twenty two pts were transplanted from a matched related donor, from a matched unrelated donor, from a mismatched related donor and from cord blood units. / pts received a reduced toxicity conditioning treosulfan-based. at the time of transplant, / pts ( %) were in cr, while / ( %) had active disease. median follow up (fu) of our cohort was s days (range, - days). wt mrd monitoring was performed by rq-pcr and considered positive for values copy numbers of wt per e abl [ ] . after allo-hsct, detection of positive wt was followed by immunomodulatory therapeutic interventions according to the time from transplant, the presence of active graft-versus-host disease (gvhd) and the general clinical conditions: tapering and/or discontinuation of immunosuppressive drugs (is), donor lymphocytes infusions (dli), administration of hypomethylating agents. median time to disease relapse was calculated from the time of detection of wt value above the threshold. results: at day post allo-hsct, out of pts ( %) were in cr. forty-five out of ( %) cr pts had wt levels persistently negative during follow up (fu) and / remained in cr at the last fu. sixty-four out of pts ( %) had at least one increase of bm wt levels above the threshold during observation and were evaluated for preemptive treatment. in / ( %) is was tapered until suspension and/or dli þ /-azacytidine was administered. median time from first wt positive value to treatment was days (range, - days). in out of these pts ( %) wt level normalized, whereas pts ( %) progressed to overt disease. all the that normalized wt remained in cr. no grade iii or iv acute gvhd was observed, while severe chronic gvhd occurred in / pts ( %). median time to disease relapse for the treated pts was days (range, - days). in / pts ( %) with a post allo-hsct wt positivity, the presence of an active form of acute or chronic gvhd prevented from applying further immunotherapy strategies. among these pts, remained in cr, relapsed. median time to disease recurrence for these pts with gvhd was days (range, - days). finally in / pts ( %) concurrent clinical issues (e.g. active infection) did not allow any attempt to prevent relapse and / relapsed. median time to disease relapse for untreated pts was days (range, - days). conclusion: in our series, pre-emptive immune-modulatory maneuvers targeting wt levels proved to be feasible and safe and of potential clinical benefit to postpone overt relapse in high risk aml/mds pts post allo-hsct. introduction: allogeneic stem cell transplantation (sct) with both myeloablative (mac) and reduced intensity conditioning (ric) is effective therapy in aml. several studies have shown that leukemia-free survival (lfs) is similar after sct with mac and ric. however, there is paucity of data on the long term outcome (beyond years) following ric due to the relative recent introduction of this approach. the alwp of ebmt published in leukemia the largest study until that time, comparing the outcome of aml patients (pts) given ric (n ¼ ) and mac (n ¼ ). the median follow-up was months. in multivariate analysis, non-relapse mortality (nrm) was significantly lower and relapse rate was significantly higher after ric resulting in similar lfs. materials (or patients) and methods: in order to better predict long-term outcome and late events we have now updated sct outcomes in a larger cohort of pts, age z years (n ¼ ) after sct from matched sibling donors, in the years - with a median follow up of . years ( . - ) . results: pts were given ric and mac regimens. the median age at sct was ( - ) and ( - ) years, respectively (po . ). % of ric recipients had advanced disease at sct compared to % of mac recipients. the percentage of pts in cr and cr / later cr was % and % compared to % and %, respectively (p ¼ . ). ric recipients were more likely to receive pbsc ( % vs %, po . ) and in vivo t-cell depletion ( % vs %, po . ). % and % had poor-risk cytogenetics, respectively (p ¼ . . , po . ) and poor cytogenetics (hr . , po . ) . the conditioning regimen did not predict -year lfs. nrm rates were higher after mac than ric throughout the late post sct course, while relapse rates were only mildly decreased at the late phases. in pts surviving leukemia-free years after sct the subsequent nrm was % and %, respectively (p ¼ . ). subsequent relapse rates were % and % (p ¼ . ) and lfs was % and %, respectively (p ¼ . ). in pts surviving leukemia-free years after sct, subsequent nrm was % and %, respectively (p ¼ . ). subsequent relapse rates were % and % (p ¼ . ), and lfs was % and %, respectively (p ¼ . ). conclusion: lfs remains similar after ric and mac even years after sct. the trend for excess nrm with mac remains throughout the late course, however excess relapse after ric is more obvious in the early phases. pts remaining leukemia-free years after sct can expect excellent subsequent outcome with both regimens. long-term follow-up studies (beyond years) are of significant importance when assessing sct outcomes. disclosure of interest: none declared. myeloablative busulfan based strategies in transplantation particularly of children with non-malignant diseases. a reduction in short term mucosal and hepatic (sos) toxicity and the absence of long term pulmonary toxicity have been demonstrated. it is unclear, however, whether this reduction of toxicity is accompanied by an equal or inferior myeloablative capacity compared to busulfan based myeloablative regimens. materials (or patients) and methods: we performed a retrospective analysis of consecutive patients with nonmalignant diseases transplanted in german pediatric transplant centers (hamburg, hannover, munich and ulm) with a treo based regimen (treo and fludarabin (flu) ± thiotepa (tt)±serotherapy) in the period between january st and june th . results: we identified patients with inborn errors including primary immunodeficiencies, hemoglobinopathies, hemophagocytic lymphohistiocytosis, mucopolysaccharidosis and osteopetrosis. the median age at transplantation was . years ( . - ) . all pts received treo/flu. tt was added in of these pts, serotherapy in pts. the os after years was %, the efs %. the incidence of agvhd ii- iv was % and . % for cgvhd - . primary engraftment of donor cells was present in % of patients. however, mixed chimerism at any time point was found in % and disease recurrence in %. an additional cellular therapy (act: stem cell boost, dli, or nd transplant) was applied in % of patients. act was more often needed after transplantations from mmfds and muds than from msd/mfds (p ¼ . ). there was a significant difference in patients who received alemtuzumab as serotherapy early (d- to d- ) versus late (d- or later) during conditioning, with % act in the early and % act in the latter group (p ¼ . ). cell source (bm or pbsc) and addition of tt did not affect the cumulative incidence (ci) of act. conclusion: there is an excellent overall survival and efs with treo based conditioning in the transplantation of patients with non-malignant diseases. despite a good primary engraftment, a high rate of mixed chimerism, disease recurrence and need for additional cellular therapy was observed. interestingly, early administration of serotherapy was correlated with a higher probability of stable donor engraftment and has to be considered as a relevant factor for transplant outcome. a randomized controlled prospective study comparing conditioning regimens with treo against busulfan in nonmalignant diseases is needed. disclosure of interest: none declared. long-term outcomes of reduced-intensity conditioning allogeneic stem cell transplantation for adult high-risk acute lymphoblastic leukemia in first complete remission s. lee ,* , k. introduction: reduced-intensity conditioning (ric) allogeneic stem cell transplantation (sct) has emerged as an option designed to lower nonrelapse mortality (nrm) for older patients and those with comorbid conditions. however, the role of ric-sct in adult acute lymphoblastic leukemia (all) remains unclear because the interpretation of transplantation outcome is mainly limited by the small sample size, short follow-up duration, various regimens for conditioning and graft-versus-host disease (gvhd) prophylaxis, and the heterogeneity of the criteria used to select patients for ric-sct. previously, we conducted a phase trial of ric-sct in adults with high-risk all and showed the potential role of this strategy, especially in patients in first complete remission (cr ). here, we report on the updated results of ric-sct by analyzing consecutive adult high-risk all transplants in cr . materials (or patients) and methods: during the period between and , consecutive patients in cr (median age, years [range, - years]) were given an identical ric regimen consisting of fludarabine ( mg/m in total) and melphalan ( mg/m in total). the indications for ric-sct were advanced age (z years; n ¼ ; . %) and comorbid conditions (n ¼ ; . %). graft sources were peripheral blood stem cells (n ¼ ; / -matched sibling donor, / -matched unrelated donor, / -matched unrelated donor) and bone marrow (n ¼ ; / -matched unrelated donor, / -matched unrelated donor). the median time-to-transplantation was days (range, - days). gvhd prophylaxis was attempted by administering calcineurin inhibitors (cyclosporine for sibling donor transplants, tacrolimus for unrelated donor transplants) plus methotrexate. antithymocyte globulin ( . mg/kg in total) was administered to the patients who received allele-mismatched unrelated donor grafts. if residual leukemia was detected in the absence of gvhd at months after transplantation, calcineurin inhibitors were rapidly discontinued. results: fifty-two patients developed grade ii to iv acute gvhd ( grade ii, grade iii, grade iv). the cumulative incidence of acute gvhd at days was . %. of the patients who survived for at least days with sustained engraftment after transplantation, developed chronic gvhd ( limited, extensive), resulting in a -year cumulative incidence of . % . after a median follow-up of months (range, - months), the cumulative incidence of relapse (cir) and nrm at years were . % and . %, respectively, and the -year disease-free survival (dfs) and overall survival (os) rates were . % and . %, respectively. within the cohort of ph-negative all transplants (n ¼ ), the -year cir, nrm, dfs, and os rates were . %, . %, . %, and . %, respectively. in a subgroup of ph-positive all transplants (n ¼ ), the -year cir, nrm, dfs, and os rates were . %, . %, . %, and . % respectively, and for these patients, minimal residual disease kinetics (early-stable molecular response vs late molecular response vs poor molecular response) and the presence of chronic gvhd were closely related to cir and dfs. conclusion: our data suggest that ric can be considered as a reasonable choice for providing a long-term disease control for adult high-risk all patients in cr . disclosure of interest: none declared. substitution of tbi by intravenous busulfan for elderly aml/mds patients within the flamsa-ric protocol is feasible and yields comparable results m. schleuning ,* , d. judith , i. burlakova , h. baurmann , r. schwerdtfeger , g. stuhler centre for hematopoietic cell transplantation, dkd helios klinik, wiesbaden, germany introduction: the flamsa-ric protocol is a highly effective conditioning protocol for high risk myeloid leukaemia patients (pts). however, many of these pts are beyond the age of . in this population the use of total body irradiation (tbi) might be toxic, even with the reduced dose of cgy, as described in the original flamsa-ric protocol. in an attempt to further reduce toxicity for elderly ( y) or comorbid pts we substituted tbi by intravenous busulfan (ivbu; x . mg/kg bw) within the flamsa-ric protocol. materials (or patients) and methods: retrospective study to analyze the results of ivbu in comparison to those achieved in pts receiving the classical flamsa-ric protocol with tbi during the same time period. results: from november to october pts with high-risk aml or mds received an allogeneic stem cell transplant after flamsa-ric conditioning. eighty-three pts (median age y) received tbi and ninety pts (median age s y) received ivbu. in the tbi group pts suffered from aml and pts from mds and in the ivbu group diagnoses were aml in and mds in pts. unfavourable cytogenetics or molecular genetics were found in % of tbi and % of ivbu pts, respectively. in the tbi group % were transplanted with active disease as compared to % in the ivbu group. in both groups stem cell grafts from unrelated donors were used in approximately % of pts. all pts engrafted. after a median follow-up of . y for surviving pts the probability of leukaemia-free survival at y after transplant is % in the tbi group and % in the ivbu group (p ¼ . ). twentyseven relapses occurred in the tbi and in the ivbu group. non-relapse mortality (nrm) for the tbi cohort was % and for the ivbu group % at day þ (p ¼ . ) and % and % during the entire observation period (p ¼ . ). the difference in nrm is probably owing to the older age of the pts in the ivbu group and was due to more infectious complications. no significant difference in survival was observed in mud or sibling transplants in either group. conclusion: in conclusion, substitution of tbi by ivbu is feasible with no enhanced relapse rates observed and should be further evaluated in prospective clinical trials also for younger pts. introduction: to ascertain the therapeutic potential of non-tbi-based conditioning for cd þ hpc-selected, t celldepleted allografts, we conducted a trial comparing our standard regimen, arm (a) cgy hftbi þ thiotepa, mg/ kg/day x days þ cyclophosphamide mg/kg/day x days vs. arm (b) busulfex . mg/kg/ h x (dose adjusted) þ melphalan mg/kg/day x þ fludarabine mg/m /day x and arm (c) clofarabine mg/m /day x þ melphalan mg/m /day x þ thiotepa mg/kg/day x , as preparation for t-cell depleted cd þ pbsc transplants from gcsfmobilized leukocytes fractionated with the clinimacs cd þ reagent system. materials (or patients) and methods: primary endpoints were engraftment, gvhd, transplant-related mortality (trm) and yr os and dfs (confer table) . stratification of pts to arms a (standard), b or c was based on the patient's disease, disease stage and clinical factors such as age, prior therapy or comorbidities enhancing risks of tbi. arm b was the non-tbi arm predominantly used for myeloid and arm c for lymphoid malignancies. prior to transplant, recipients of hla-matched or non-identical transplants received rabbit thymoglobulin at . mg/kg/day x or days respectively, to prevent graft failure. no gvhd drug prophylaxis was given post transplant. results: a total of consecutive patients, accrued between / / and / / , were analyzed ( in arm a, in arm b, in arm c). these pts have been followed for a median of . months. donors were related or unrelated and hla-matched for % of the patients and - hla alleles disparate for %. median age for the entire group was . years, with older pts predominating in the non-tbi groups (medians arm a, . yrs; arm b, . yrs; arm c, yrs). the cd þ pbsc transplant provided a mean dose of . x cd þ progenitors/kg (range . - . ) and . x cd þ t-cells/kg (range . - . ). all pts engrafted; but pts ( . %) in arm b experienced late graft failure, one of whom was reconstituted after a secondary graft. overall the incidence of grade ii-iv acute gvhd was %, % and % for arm a, b and c respectively. trm at year was % in arm a, and % in arm b and % in arm c. two year os and dfs for each arm are: arm a - . % and %; arm b - % and % and % and % for arm c. for the pts who received standard risk transplants (i.e., pts with high risk forms of aml, all or nhl in o cr, aml in o cr, mds ra/rcmd, cml in o cp or mm in cr ), year os and dfs are: arm a - % and %; arm b - % and %; arm c - % and %, with relapse rates at yrs of arm a - %, arm b - . %, and arm c - . %. cumulative incidence of relapse (cir) stratified by risk group reveals that the probability for relapse is significantly higher for the pts with high risk disease (p o . ), whereas the cumulative incidence of non-relapse mortality (nrm) is comparable (p ¼ . )( figure ). the median time to relapse has not been reached in either group. the estimates of relapse at year were . % ( % ci, . - . % ) for high risk pts and . % ( % ci, . - . %) for the other group. the estimated -year cir was . % ( % ci, . - . %) for the group at high risk and . % ( % ci, as measured from allosct prior to dli bm progression and focal progression patterns were highly similar with cumulative incidences of bm progression of %± % and %± %, and of focal progression of % ± % and % ± %, after and months post-transplant, respectively. in contrast, as measured from dli bm progression and focal progression patterns showed strong dissimilarity: at and months after dli cumulative incidences of bm progression were %± % and %± % respectively, whereas focal progression was % ± % and % ± %, respectively, illustrating a potent immunological response in bm with only limited effect of dli on focal lesions. conclusion: disease progression patterns of multiple myeloma after tcd-ric allosct diverged from initially similar bm and focal progression patterns in the absence of alloimmune responses towards disease control in bm with focal progression after dli. this finding illustrates failure of donor lymphocytes to target extra-medullary/focal disease in multiple myeloma. disclosure of interest: none declared. engineered t cells modified to express a cs- -specific chimeric antigen receptor (car) confer anti-myeloma activity in vitro and in pre-clinical in vivo models introduction: adoptive immunotherapy with t cells that were modified by gene-transfer to express tumor-targeting chimeric antigen receptors (cars) has therapeutic potential in advanced b-cell malignancies. we are pursuing the glycoprotein cs (slamf /cd ) as candidate target for car t cells in multiple myeloma (mm), due to its restricted high level expression on malignant plasma cells in a significant proportion of mm patients. here, we evaluated the anti-mm function of t cells that we modified with cs -specific cars in vitro and pre-clinical in vivo models. materials (or patients) and methods: we constructed two cs -specific cars with antigen-binding domains derived from the huluc and luc mabs that target distinct cs epitopes, each comprising a signaling module of cd zeta and a cd co-stimulatory domain, and encoded both constructs in lentiviral vectors for gene-transfer. results: cd þ and cd þ t-cell lines expressing the huluc and luc cs -cars could be readily generated from healthy donors and mm patients (n ¼ / ), and propagated and expanded in vitro with similar kinetics as t cells expressing a cd -specific car that we included as a reference. in functional experiments, cd þ t cells expressing either of the cs -cars conferred specific high level lysis of mm lines (mm .s, nci-h and opm- ), primary mm, k that had been stably transfected with cs , but not native cs -negative k . we also detected high level production of ifng and il- (cd cd ), and productive proliferation after stimulation with cs þ target cells, with significantly superior anti-mm reactivity mediated by the huluc compared to the luc cs -car construct. we confirmed the anti-mm efficacy of cs -car modified t cells using a xenograft model in immunodeficient mice (nsg/mm .s). mice were inoculated with fireflyluciferase labeled mm .s myeloma by tail vein injection and days later, when mice presented with disseminated disease, administered a single dose (i.v.) of a cell product consisting of equal proportions of cd þ and cd þ t cells modified with either the optimal cs -car, the cd -specific car or mock transduced. we observed rapid and durable complete rejection of established mm from bone marrow and resolution of extramedullar mm manifestations in all of the mice treated with cs -car t cells (n ¼ ), whereas mice treated with cd -car t cells, or control t cells had to sacrificed due to progressive disease (n ¼ / ). of interest, in this in vivo model, we observed similarly effective anti-mm responses mediated by cs -car t cells that had been derived from healthy donors and mm patients. conclusion: our data suggest the potential of t cells expressing cs -specific cars to confer anti-mm activity in clinical settings. the experience with anti-cs mab huluc , that as single agent has only minute anti-mm activity, indicates targeting this molecule will be safe and not be associated with toxicity to normal tissues. we observed stronger anti-mm reactivity with cs -cars targeting a proximal (huluc ) rather than a distal (luc ) epitope on cs protein, in line with our previous observation that the targeted epitope on a given antigen affects tumor recognition of car t cells. experiments to analyze the function of our cs -cars against panels of primary mm in our nsg model are ongoing to inform our efforts of clinically translating car t-cell therapy in this entity. ; ) , t( ; ), del p by fish and/or del q by karyotyping]. all pts had to achieve at least a partial response from preceding salvage chemotherapy (n ¼ ) or second salvage auto hsct (n ¼ ). pts underwent allo tcd hsct with busulfan ( . mg/kg x doses), melphalan ( mg/m x days), fludarabine ( mg/m x days) and rabbit atg ( . mg/ kg x days). tcd was performed by positive cd selection (isolex) followed by rosetting with sheep erythrocytes for the initial pts ( - ) and by cd þ enrichment by the miltenyi device in pts thereafter, achievingo cd þ /kg for all grafts. none of these pts received immuno suppressive therapy post tcd hsct. pts with / hla matched donors were also eligible to receive low doses of dli ( x e - x e cd þ /kg) no earlier than mos post allo hsct. results: pts with a median follow up of . mos (range: . - . mos) of survivors are reported, median age years (range - ). all pts engrafted promptly (median d þ , range þ -þ ).trm (grade ii-iv) at mos is % ( % ci: % - %). acute gvhd was % ( % ci: % - %) and chronic gvhd was not observed in any pt. the overall survival (os) and progression-free survival (pfs) with their % confidence intervals (ci) are shown in table . factors associated with worse outcome were disease status and number of previous treatments prior to tcd hsct. ( ) . in this study, we have analyzed the molecular consequences of del( )(p ), an abnormality we and others have previously shown to have an adverse impact on survival of mm patients ( ) ( ) ( ) . materials (or patients) and methods: in a cohort of patients that were diagnosed with mm between and , we have investigated the clinical impact of del( )(p ) on time to progression (ttp) and overall survival (os). moreover, response rate of patients to st line bortezomib treatment was investigated. we have also analyzed the expression profiles of genes located near the p region in patients with and without del( )(p ). additionally, we have analyzed the in vitro response of primary mm cells with and without the deletion to bortezomib-mediated killing and sensitization to trail/apo l-triggered apoptosis in an attempt to understand why mm patients carrying p deletion respond poorly to bortezomib treatment. results: we found that mm patients carrying del( )(p ) deletion had significantly shorter ttp compared to patients without the deletion (p ¼ . ) and most importantly these patients had significantly shorter os compared to patients without the deletion (p ¼ . ). in a cohort of patients, we observed that patients with del( )(p ) (n ¼ ) responded poorly to bortezomib, % showing no response while % of patients without the deletion (n ¼ ) responded to bortezomib treatment. in vitro analysis revealed that mm cells from patients with del( )(p ) show higher resistance to bortezomib treatment possibly due to upregulated expression of genes such as ptk b, ccdc , rhobtb , nfkb, myc and bcl while showing downregulated levels of tp and scara when compared to mm cells without the deletion. furthermore, we have observed that mm cells with del( )(p ) express higher levels of the decoy death receptor, trail-r and fail to upregulate the pro-apoptotic death receptors trail-r and trail-r that are located in the p region. as a result, mm cells with del( )(p ) were largely resistant to bortezomib and trail/apo l-mediated apoptosis. conclusion: substantiating the clinical outcome of the patients, our data provides a potential explanation regarding the poor response of mm patients with del( )(p ) to bortezomib treatment. furthermore, our clinical evaluation suggests that including immunomodulatory agents such as lenalidomide in the treatment regimen may help to overcome this negative effect, providing an alternative thought in planning treatments of patients with del( )(p ). introduction: autologous stem cell transplantation is the standard treatment in patients with multiple myeloma (mm). however, there is discrepancy over the optimal mobilization regimen. therefore a randomized study was conducted to compare cellular composition of the collected grafts as well as early hematopoietic and immune recovery in mm patients receiving g-csf with or without low-dose cyclophosphamide for mobilization of blood grafts after induction with lenalidomide, bortezomib and dexamethasone. materials (or patients) and methods: thirty patients with mm were included into this prospective multicenter study. there were males and females with a median age of years (range - ). fourteen patients were mobilized with cyclophosphamide plus g-csf (arm a) whereas sixteen patients were mobilized with g-csf alone (arm b). melphalan mg/m was used as high-dose therapy and patients having graft cd þ cell countso x /kg (measured before freezing) were scheduled to receive g-csf after the graft infusion. cryopreserved graft samples were analyzed with a flow cytometry for t and b cells (cd /cd /cd /cd ) as well as for nk cells (cd /cd þ cd /cd ). also cd þ cell subclasses were analyzed (cd /cd /cd /cd ). complete blood counts were evaluated on day þ and one month post-transplant and a flow cytometry for blood lymphocyte subsets (t, b, nk) was performed one month after the graft infusion. results: the blood grafts in arm a contained significantly higher amounts of cd þ cells and the grafts of the arm b contained significantly higher proportion of primitive cd þ cd þ cd cells and t, nk and b lymphocytes ( table ) . the median amount of infused cd þ cells was comparable between the arms ( .  /kg in group a vs. .  /kg in group b, p ¼ . ). the number of platelets was slightly lower in the group b at d þ (p ¼ . ) otherwise the course of early hematological and immune recovery was comparable between the groups. the use of g-csf alone instead of a combination with cyclophosphamide seems to enrich the blood grafts with significantly higher number of t and b lymphocytes and a higher proportion of more primitive stem cells. the hematological and immune recovery was comparable between the arms. the possible effects of graft composition in long-term patient outcomes will be further evaluated in the ongoing goa study (graft and outcome in autologous stem cell transplantation). disclosure of interest: none declared. introduction: pet is a useful tool that allows deeper assessment of response beyond that measured by m protein levels. it has been reported to predict outcome following both asct. to be able to integrate pet-ct negativity to internationally accepted response criteria the cut-off level needs to be validated by independent investigators. this prospective study was initiated to elucidate the prognostic role of pet-ct in the asct setting utilizing the cut-off found in our patients in ankara university ( . ) comparing with those initially reported by barlogie et al ( . ) and zamagni et al ( . ) materials (or patients) and methods: consecutive patients diagnosed and transplanted in ankara university with pre-and post-asct fdg-pet-ct imaging were included. patients were: median age . þ /- . (m/f: / ), iss i/ii/iii: / / , renal impairment ( , %), bone involvement ( , %), del q ( . %), t ( ; ) and/or p ( . %), ldh high( , %), induction with bortezomib ( , %). pre-asct clinical response vgpr: . %, post-asct clinical response vgpr: . %. overall survival (os): median: months ( . - months). pasw statistics for windows program was used for statistical analysis. results: as reported previously roc analysis revealed . as a significant cut-off level (p ¼ . ; os). pet-cr was defined fdg uptake less than . or . depending on the analysis. post-asct pet ( . ) was predictive for pfs (p ¼ . ) but not os (p ¼ . ) . however pet ( . ) was predictive for os (p ¼ . ) but not pfs. depending on the cut-off more (suv . : / ) or less (suv . : / ) patients met the criteria for pet-negativity (or remission) following asct. expert pet assessment resulted with pet-cr / similar to the suv . frequency. as shown in figure patients to converted to cr after asct (positive/negative group) displayed a better pfs than those who had reached cr prior to asct. this analysis was significant if cut-off was . but not . . expert assessment was also able to differentiate patients with better prognostic features. . - . ). leukaphereses were analized for mrd: the median plasma cells value was , % ( , - , %); in pts pcs wereo , % (cut off for mrd negativity). conclusion: mobilization with cy-bor-dx þ g-csf and borbased asct is safe and effective in elderly mm patients. this schedule allows the collection of an adequate dose of cd þ cells, with a very low rate of mobilization failure ( %), also in elderly mm pts. a low rate of clonal pcs contamination in the harvest was also observed. this approach allows to perform asct in most elderly pts, achieving high response rate and promising outcome with a short term treatment: weeks compared to the non-asct programs ( weeks in the vmp program). disclosure of interest: none declared. mica expression levels were investigated in n ¼ gut biopsies with sybr green s qrt-pcr. histological grades of the gastrointestinal gvhd (gi gvhd) were determined by the pathology department at the university clinic, regensburg and severity was grouped by assigning an apoptotic score ( ¼ absence of apoptosis, ¼ maximum apoptosis). a protein biochip array (evidence investigator s , randox) was utilised for measuring mica serum levels and evaluated in n ¼ samples from allo-hsct patients collected at pretransplantation, day- , day þ , day þ and months post transplantation. results: our analysis showed that the methionine allele in rs was associated with an increased risk of relapse (p ¼ . ). the same allele was also found to be associated with a reduced overall survival (p ¼ . ) which was more severe for non-t cell-depleted allo-hsct (p ¼ . ). vice, versa, the presence of the valine allele was associated with the development of agvhd (p ¼ . ). in the gut, mica expression was investigated in patients treated with low doses of steroids (r mg/kg), as high dose steroid treatment strongly suppressed mica expression. higher levels of mica were associated with an apoptotic score ¼ (no apoptosis) (p ¼ . ) and the absence of active gi gvhd (p ¼ . ). increased soluble mica levels at months post-transplantation were significantly associated with agvhd (p ¼ . ). conclusion: mica molecules have been shown to play prominent roles in immune processes and therefore are also potential agvhd biomarkers. in this study, we showed that the methionine (mica- met) allele was associated with the incidence of relapse while the valine (mica- val) allele was associated with an increased risk agvhd. a low overall survival for patients who did not have had the t cell depletion treatment was also associated with the presence of the methionine (mica- met) allele. in the gut of patients treated with low doses of steroid, mica gene expression levels were higher with the absence of gvhd. this may indicate that the isoforms are able to meditate nk-cell and t cell inactivation, and down-regulate nkg d with high levels of soluble mica contributing to the development of agvhd. eleven patients needed gvhd treatment: pts received ganciclovir iv (gcv mg/kg/ h/ days), pts valganciclovir per os (vgcv mg/ h/ days). both gcv and vgcv were effective in control clinical manifestations of gvhd in a median of days (range - ) and resulted in a significant reduction in numbers of circulating tk-cells, without reduction of cd þ tk-negative lymphocytes resulting in no effect on long-term immune reconstitution. in five patients additional concomitant treatment with low-dose steroid (prednisone o . mg/kg per day for a median of weeks) was given. a pt who presented severe gut and liver gvhd and one pt who received at transplantation an high dose of unmanipulated lymphocytes ( . x /kg) -were succesfully treated with a combined therapy of prednisone and cyclosporine or rapamicine in association with gcv. one patient developed a severe classic de novo c-gvhd, with sclerodermatous lichenoid skin and mouth features plus moderate dry-eye symptoms that was successfully treated with vgcv and a transient course of mycophenolate mofetil ( g per day) over a months period. no cases of quiescent or progressive c-gvhd was observed after a median follow-up of days (range / ). conclusion: in our long-lasting clinical application of haploidentical tk-cells, an effective induction of immune reconstitution and a complete control of gvhd, provided a long-term immunosoppressive therapy free survival in absence of gvhd related deaths or longterm complications. introduction: despite major improvements in allogeneic hematopoietic cell transplantation (allo-hct) over the last decades, severe corticosteroid-refractory acute and chronic graft-versus-host-disease (gvhd) still remains a life-threatening complication characterized by high mortality rates ( - %). since preclinical and early clinical evidence indicated anti-inflammatory effects of ruxolitinib, we collected the outcome data from multiple stem cell transplant centers using the jak / inhibitor ruxolitinib as salvage treatment in patients suffering from corticosteroid-refractory gvhd. materials (or patients) and methods: a total of stem cell transplant centers in germany, france, switzerland and united states reported outcome data from patients who received ruxolitinib for corticosteroid-refractory gvhd (skin, mucosa, intestine, liver, lung, musculoskeletal) between / and / . patients were classified as having acute (n ¼ ) or chronic (n ¼ ) gvhd. the median number of previous gvhd-therapies was for acute gvhd (range: - ) and for chronic gvhd (range: - ). results: the overall response rate was . % ( / ) in acute gvhd comprising crs ( . %) and prs ( . %). in chronic gvhd the overall response rate was % ( / ). clinical improvement was rapid with a median time to response of . ( - ) weeks and ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) week after initiation of ruxolitinib treatment in acute and chronic gvhd, respectively. all responders were in persistent remission at last follow-up. the median follow-up was ( - ) and ( - . ) weeks for acute and chronic gvhd patients, respectively. non-responders (acute gvhd: / , chronic gvhd: / ) received other salvage therapies. cytopenias (anemia, leukopenia or thrombocytopenia) and cmv reactivation were observed during the time of ruxolitinib treatment in both acute ( / , . % and respectively / , . % ) and chronic ( / , % and respectively / , %) gvhd patients, sometimes however cytopenias already preceded ruxolitinib treatment. ruxolitinib treatment was stopped or reduced in patients because of cytopenia. cmv was controlled by antiviral therapy in all patients. ruxolitinib responders died, one because of leukemia relapse weeks after ruxolitinib was stopped, and two from gvhd progression. in one of the patients who died, ruxolitinib was stopped due to impossibility of oral drug application. conclusion: overall, these data collected in multiple centers using different strategies for gvhd prophylaxis and treatment suggest that ruxolitinib is a very promising agent in the treatment of corticosteroid-refractory acute or chronic gvhd and may be successfully used to treat a major subset of patients beyond nd line of gvhd treatment. a prospective randomized multicentre clinical trial testing therapeutic jak / inhibition as salvage treatment in gvhd is planned to verify the efficacy of ruxolitinib and to identify potential biomarkers that may be predictive for response. disclosure of interest: none declared. transplant-associated renal microangiopathy is associated with a high risk of refractory acute gvhd and characterized by a specific biomarker signature introduction: there is increasing evidence that endothelial damage is involved in the pathogenesis of steroid-refractory graft-versus-host disease (refgvhd). recently, serum soluble st (suppressor of tumorigenicity, il- receptor), an independent risk factor of cardiovascular death, has been identified as a risk factor of refractory gvhd. however, the pathomechanism of endothelial cell dysfunction which is associated with mortality from gvhd is yet poorly characterized. renal transplant-associated microangiopathy (tma) is another endothelial complication of allosct, and its association with severe gvhd and with biomarkers of endothelial damage (st , scd (soluble thrombomodulin)), and endothelial function (vegf) is investigated in this study. materials (or patients) and methods: evidence for renal tma was studied in a cohort of patients who underwent allosct between and at our institution and who have provided informed consent for this observational study. criteria to diagnose renal tma included an otherwise unexplained % rise in creatinine and lactate dehydrogenase (ldh) levels (or a pre-existing ldh above u/l), a % drop in platelet counts (or a pre-existing platelet count below /nl) and at least % schistocytes. cytokines were measured in sera taken prior to allosct and on the indicated days thereafter and stored at minus c. statistical analyses were performed using spss and included a cumulative incidence analysis of causespecific hazards and the non-parametrical median test of independent probes. results: both renal tma and refgvhd were rare complications after allosct but were significantly associated with each other (tma only / ( . %), refgvhd only / ( . %), both / ( . %), chi . ). median time intervals from allosct to renal tma and refgvhd were . ( . - . ) months and . ( . - . ) months respectively. in the overlap group, gvhd onset usually occurred before renal tma (- . months, - . to . ) nrm rates were significantly increased in all three cohorts but approached % in patients with both complications. serum stm levels as well as soluble st levels increased between transplantation and day /day after allosct in all cohorts, refgvhd, tma and both. in contrast, vegf levels (day ) were significantly lower specifically in patients with tma with or without refgvhd, but not in patients with refgvhd without tma. conclusion: this study identifies renal tma as an endothelial cell dysfunction associated with extremely high mortality rates in the context of gvhd. the absence or presence of renal tma defines two separate subsets of refgvhd with a different prognosis. biomarkers of endothelial damage or vulnerability, such as stm, st , and vegf can help to dissect tma and refgvhd, and might be useful to identify and guide management of patients with endothelial dysfunction who are at high risk of fatal complications after allosct. disclosure of interest: none declared. introduction: snps of the key cytokines and chemokines involved in the pathogenesis of agvhd have become an object of major interest recently. here we present snps rs cc/ct/tt and rs aa/ag/gg of the nf-kb gene in association with agvhd. materials (or patients) and methods: in our single-center study we analyzed patients allografted for the following hematological malignancies: aml ( %), all ( %), cll ( %), mds ( %), cml ( %), nhl ( %), imf ( %), cmml ( %), and other hematological disorders ( %) between - . the median age of the study group was ( - ) years. patients were allografted after myeloablative ( %), non-myeloablative ( %) and reduced (intensity/toxicity) conditionings ( %). gvhd prophylaxis was done by solo cyclosporine-a ( %), cyclosporine-a with mycophenolate mofetil ( %) and cyclosporine-a with short-methotrexate ( %). ''in vivo'' t-depletion with thymoglobuline was used in % of recipients. patients were allografted from hla identical donors (related %) with median age ( - ) years. the female donor/male recipient combination represented % of all pairs. the grafts contained median . ( . - . ) x cd þ cells/kg and median . ( . - . ) x mnc /kg. snps analysis was done from genomic dna isolated from edta-treated peripheral blood. genotyping was performed with sequenom massarray platform using allele-specific maldi-tof mass spectrometry assay (sequenom, san diego, ca, usa). primers were designed using the sequenom snp assay design software version . for iplex reactions. univariate analysis was performed to find significant difference in agvhd among the patient groups with different nf-kb profiles. the asymptotic pearson's chi-square test was used in cross tabulation with significance level set to . . results : out of patients patients ( %) are still alive, patients ( %) died ( died of transplant-related complications). the median post-transplant follow-up was . ( . - . ) years. agvhd developed in patients ( %), grade iii-iv in of them ( %). both nfkb snps were completely correlated in the sense that knowing the genotype of one fully determined the genotype of the second one ( patients did not follow this correlation and were excluded from the analysis). consequently, we defined a common predictor nf-kb which codes the information carried by both nf-kb snps in the following way: all patients carrying the rs cc genotype were also positive for rs aa allele and were marked as nf-kb ¼ i, all patients carrying the rs ct genotype were also positive for rs ag allele and were marked as nf-kb ¼ ii and finally all patients carrying the rs tt genotype were also positive for rs gg allele and were marked as nf-kb ¼ iii. the nf-kb profile was found to be significantly associated (p ¼ . ) with agvhd in the following way: patients in the nf-kb ¼ i group are more probable to suffer from agvhd than in the nf-kb ¼ iii group. conclusion: this is the first report showing the association of nf-kb gene snps with agvhd. the transcription factor nf-kb has been implicated in the regulation of cellular stress and inflammatory signals. according to our pilot data patients with inherited genetic abnormalities of the nf-kb gene may be prone to agvhd. patients.the two groups -control/treatment -were well balanced in terms of diagnosis (p ¼ . ), and disease phase (p ¼ . ) : the most frequent diagnosis was aml (n ¼ ), followed by all (n ¼ ) and mds (n ¼ ). the median age for control/treatment was years ( - ) vs yers ( , , (p ¼ . ); the proportion of patients over years was % in the control and % in treatment group (p ¼ . ). the donor type in the control/treatment arms was as follows: hla identical siblings n ¼ /n ¼ , unrelated cord blood (cb) n ¼ /n ¼ , unrelated donor (ud) n ¼ /n ¼ , and haploidentical family donors (haplo) / (p ¼ . ). skin biopsies.a skin biopsy before randomization, was not mandatory: it was performed in patients. gvhd was diagnosed as proven, probable and possible respectively in s %, % and %of the patients.these different reports were equally distributed in treatment and controls (p ¼ . ). results: the cumulative incidence of acute gvhd grade ii (primary end point), was % in controls and % in treatment patients (p ¼ . ). this difference was maintained in different subgroups. the ci of transplant related mortality (trm) was % (control) vs % (treatment) (p ¼ . ), despite a non significant younger median age in treatment patients. excess mortality in the treatment arm was due to an excess incidence of infections. actuarial year survival was % (control) vs % (treatment) (p ¼ . ). cases of death in the control/treatment groups were as follows: gvhd % - %; infection, % %; interstitial pneumonia, % - %; toxicity, % - %; leukemia relapse % - % (p ¼ . ). there was no significant difference in trm among different centers (p ¼ . ) progression of gvhd and skin biopsies.the proportion of patients progressing to gvhd grade ii þ was simlar in proven, probable, possible gvhd ( %, %, % ) (p ¼ . ). however the proportion of gvhd related deaths was % for proven gvhd, and % for probable/possible gvhd. introduction: graft versus host disease (gvhd) is a common and severe complication after allogeneic stem cell transplantation. during pregnancy, the placenta and the fetal membranes function as an immunological barrier, protecting the fetus from the mother's immune system. we have isolated stromal cells from the decidual layer of term placentas. decidual stromal cells (dscs) are of maternal origin and strongly inhibit the alloreactivity of t-cells in vitro. the effect is mainly contact dependent, and decreases the production of several key cytokines involved in the cytokine storm promoting the continuation of gvhd. materials (or patients) and methods: to investigate the effect of dscs on acute gvhd we enrolled patients diagnosed with acute gvhd and clinically non-responsive to standard therapy. the protocol was modified after patients, the dscs were then thawed and infused in infusion solution with albumin instead of ab-plasma and given repeatedly and earlier upon diagnosis. this led to the formation of two treatment groups (group n ¼ and group n ¼ ), which were compared to matched historical controls (n ¼ ). we also performed a retrospectively corrected analysis of steroid refractivity. results: group received a median of infusion on day after standard treatment compared to group , who received a median of infusions (p ¼ o , ) on day (ns). no adverse events related to the treatment were observed. at weeks after treatment, % of the patients in group hade responded to the treatment. in contrast, all patients in group responded (p ¼ o , ). all patients in the treatment groups received fungal prophylaxis. the overall cumulative survival (os) at one year was % for the controls as compared to % for group and % for group (p ¼ o . ). when the groups were corrected for steroid-refractivity, the os was % for the controls, % for group and % for group (p ¼ o . ). in the steroid-refractory control group (n ¼ ), the risk of dying from gvhd at one year was %, whereas no patients in the steroid-refractory group died from gvhd(n ¼ , p ¼ o . ). in the last months, no patients have died from acute gvhd at our center. in conclusion, dscs might be an effective treatment of acute gvhd. the infusion should be prepared in albumin, given as early as possible and in repeated doses. to confirm these striking findings, we will extend the followup time and enroll more patients in our study. disclosure of interest: none declared. oral session: stem cell source and donor type o unmanipulated haploidentical stem cell transplantation after reduced intensity or ablative conditioning regimen for the treatment of acute leukemia-a report from the acute leukemia working party of the ebmt m. introduction: haploidentical hematopoietic stem cell transplantation(haplo-hsct)is feasible option for patients with acute leukemia(al)at high risk of relapse who do not have hla-matched related or unrelated donors. haplo-hsct was associated with severe acute graft-versus-host disease (agvhd) in unmanipulated transplants and a high incidence of graft rejection in t-cell depleted transplants because of the high frequency of t cells that recognized major class i or ii hla disparities between donor and recipient. two approaches were developed to overcome these problems:megadose of t-cell depleted hematopoietic progenitor cells without any posttransplant immunosuppression and t-cell replete grafts with innovative pharmacological prophylaxis of agvhd.posttransplant cyclophosphamide(ptcy)is regarded as a gvhdspecific immunosuppressant in adults but its feasibility is unknown in children. purpose: to evaluate the feasibility and outcome of haplo-hsct in children and adolescents with acute leukemia in active disease depending on conditioning regimens and methods of harvesting haplo-graft. primary end points: overall survival (os), transplant-related mortality (trm). secondary end points: engraftment rate, agvhd, cgvhd, relapse rate. materials (or patients) and methods: patients(range from - y.o. median y.o.)with al(all- pts, aml- pts)in progressive disease (pd)(cytoreduction chemotherapy (ctx) prior conditioning regimen- pts, without- pts) were analysed.mac þ atg regimen based on giac protocol received pts, mac þ ptcy mg/kg on d þ , þ - pts, ric þ atg regimen based on flu- pts, ric þ ptcy mg/kg on d þ , þ - pts. all pts received prophylaxis of agvhd based on csa- pts, tac- pts, tac þ sir- pts. g-csf-primed t-cell replete bm was used as a graft source in pts (median cd þ cells , x /kg), g-csf mobilized peripheral blood (cd þ selected by clinimacs, miltenyi biotec) and g-csfprimed bm- pts (median cd þ cells , x /kg). [o ] results: -year os was , %. ptso y.o. had significantly higher os vs pts y.o. , % and , % respectively (p ¼ , ). -year os in pts receiving ctx prior conditioning regimen was % vs , % in pts with leukemic burden(p ¼ , ). significantly difference were observed in year os in pts transplanted g-csf-primed t-cell replete bm , % vs % in pts after g-csf mobilized peripheral blood and g-csf-primed bm (p ¼ , ). pts receiving ric þ ptcy had -year os , %, ric- , %, mac- %, mac þ ptcy- % (p ¼ , ). trm after haplo-hsct in pts with al in pd was %. engraftment was sustained in , % pts. full donor chimerism was achieved in , % pts on d þ . median anc engraftment ( , x /l) d þ ,plt recovery( x /l) d þ . cumulative incidence of grade - agvhd was , %,cgvhd- , %. cumulative incidence of relapse was , %. conclusion: haplo-hsct g-csf-primed unmanipulated bm is an effective method of achieving remission with good sustained engraftment rate in children and adolescents with resistant disease. ric regimen followed by t-cell replete haplo-hsct with ptcy on d þ , þ was associated with good os, low incidences of gvhd and trm. before any definitive conclusions can drawn, a randomized study is required. disclosure of interest: none declared. the detection of donor specific anti-hla antibodies in recipients of unmanipulated haploidentical blood and marrow transplantation is predictive of poor graft function introduction: our previous study suggest that choosing young, male, non-inherited maternal antigen-mismatched donors is reasonable following unmanipulated haploidentical blood and marrow transplantation (hbmt). recently, a correlation between the presence of donor-specific anti-hla antibodies (dsa) and graft failure has been demonstrated in haploidentical transplant settings. in our protocol, approximately % patients can achieve sustained, full donor chimerism. however, poor graft function (pgf) remains one of complications after unmanipulated hbmt. therefore, we determined the effect of dsa on primary pgf in order to provide further evidence for donor selection. materials (or patients) and methods: three hundreds and fourty-five patients with hematological diseases receiving hbmt were enrolled in this prospective study. the median age of the patients was years (range, - years). these patients were randomly selected as training group (n ¼ ) and validation group (n ¼ ). peripheral blood serum were collected pre-conditioning regimens. dsa were determined using the luminex-based assay. results: in all patients, the percentages of dsa positive cases were . % ( / ). the incidence of dsa in female patients was higher than that of male cases ( % vs. %, p ¼ . ). in the training set, a cutoff value of dsa (mfi ¼ ) were developed. multivariate analysis showed that the presence of dsa (patients with mfiz vs. cases with mfiz ) was associated with primary pgf. in the validation set, the association of dsa with primary pgf following transplantation was also confirmed. the association of pgf with inferior overall survival (os) was demonstrated both in the training group and in the validation group. in all patients, the median time to platelet recovery in dsa positive (mfiz ) patients was slower than that of dsa negative ones ( days vs. days, p ¼ . ). the incidence of primary pgf and primary graft failure was . % and . %, respectively. dsa positive patients experenced higher incidence of primary pgf ( % vs. introduction: transplacental trafficking of maternal and fetal cells during pregnancy establishes long-term, reciprocal microchimerism in both mother and child because of exposure of the two immune systems to the non-self alloantigens (maloney et al., j clin invest. ). studies show the immune system in the mother is capable of being sensitized by paternal histocompatibility antigens. for example, antibodies directed against paternal hla-antigens (van rood et al., nature. ) and t lymphocytes directed against paternal major (van kampen et al., hum immunol. ) and minor histocompatibility antigens (verdijk et al., blood. ) are frequently detected in multiparous women. we previously demonstrated mother/child immune interactions positively influenced the outcome of mother to child hla haploidentical t cell-depleted hematopoietic transplantation. in a series of adult and pediatric patients we demonstrated mother donors conferred protection against leukemia relapse and improved transplant related mortality (trm), which was largely due to infection, and improved survival (stern et al., blood ). materials (or patients) and methods: the kaplan-meier method evaluated leukemia-free survival. cumulative incidence estimates were used for relapse and trm, as they are competing risks. multivariate analysis assessed the impact of diverse variables on transplantation outcomes. results: we analyzed the outcomes of adult acute leukemia patients after t cell-depleted haploidentical transplantation. when compared with transplantation from all other family members, transplantation from mother donors was associated with significantly lower trm (largely infectious) ( % vs % from all other donors, p ¼ . ). multivariate analyses demonstrated transplantation from mother donors was an independent factor predicting improved survival (hazard ratio . , % confidence interval . to . , p ¼ . ). in an attempt to elucidate the mechanism, we analyzed donor t cell repertoires that were specific for cmv antigens presented by recipient antigen-presenting cells (by elispot and by limiting dilution cloning). unlike all other donor/recipient pairs, mothers possessed cmv-specific cd cell clones that killed child's and father's cmv-pulsed dendritic cells (dcs). such clones were nonalloreactive as they did not kill the child's or father's non-cmv-s pulsed dcs. mothers also possessed cd t cell clones that produced ifn-gamma in response to child's and father's cmvloaded dcs. such clones were non-alloreactive as they did not respond to child's or father's non-cmv-pulsed apcs. thus, mothers possessed a t cell repertoire that recognized cmv antigens also when presented by the unshared, father's, hla haplotype. in fact, they showed twice as many t cells that recognized cmv antigens presented by the child's apcs than all other donor/recipient pairs (po . ). conclusion: therefore, pregnancy resulted in the generation of an additional t cell repertoire that specifically recognized pathogen antigens presented by the unshared paternal hla haplotype antigens on the child's apcs. apparently, upon mother to child t cell-depleted hematopoietic transplantation, such repertoire expands over time and helps reduce infectious mortality. further studies are needed to elucidate the mechanisms underlying mother t cell selection/education by paternal hla haplotype antigens on the child's apcs. disclosure of interest: none declared. uni-directional and bi-directional non-permissive hla-dpb t cell epitope group mismatches have similar risk associations in / matched unrelated donor hct , was analyzed after separating uni-directional from bi-directional non-permissive mismatches. non-permissive mismatches were defined as unidirectional hvg when the donor but not the patient carried an hla-dpb allele from a tce group not present in the patient, and vice versa as uni-directional gvh. bi-directional nonpermissive mismatches were present when none of the hla-dpb alleles in patient and donor were from the same tce group. the associations with clinical endpoints of overall survival (os), transplant related mortality (trm), relapse, acute gvhd (agvhd) and chronic gvhd (cgvhd) were studied using multivariate proportional hazards methods. results: the number of transplants with permissive or nonpermissive uni-directional hvg, uni-directional gvh and bidirectional mismatches was , , and , respectively. in the trm analysis, non-permissive uni-directional hvg (hr . , p ¼ . ) and gvh mismatches (hr . , p ¼ . ) had significantly higher relative risks (rr) of trm compared to the permissive group. the bi-directional group had similar rr (hr . , p ¼ . ). in pairwise comparisons, there were no statistical differences between the uni-and the bi-directional non-permissive groups for any of the outcomes tested. introduction: there are several alternative sources of donor stem cells available for patients (pts) who need an allo-sct and especially for those who lack a hla-matched donor. outcomes of mismatched-unrelated-donor (mm-urd) transplant have recently improved, and a comparison between matched and mm-urd sources in a uniform cohort of pts has not been performed after ric regimens. materials (or patients) and methods: pts, aged z year, who underwent fully matched or mm-urd ric pbsct or bmt from - were included in the study. all donors were hla-matched ( / ) or mismatched at one or two-loci ( / or / ). the kaplan-meier-estimator, the cumulative incidence function and cox proportional hazards regression models were used where appropriate. results: in total pts receiving matched or mismatched ric-urd allo-sct were included in the study (aml , all ). / hla-matched pts were compared with recipients receiving / (n ¼ ) or / (n ¼ ) after ric mm-urd allo-sct. median age of / , / and / hlamatched recipients were years. higher female donor to male recipients were in / cohorts ( %) compared to / ( %) and / ( %) group (p ¼ . ). more pts with cr and advanced-disease were among / and / cohort compared to / matched donor recipients (cr , and %; advanced disease , , % respectively; p ¼ . ). also higher percentage of pts with secondary leukemia were in / cohorts compared / and / matched donor ( , , %; p ¼ . ). percentages of engraftment ( %, %, %, p ¼ . ) were no different between the groups. acute gvhd grade ii-iv was %, %, %, and grade iii-iv , , and %, respectively for / , / and / matched donors, respectively (p ¼ . and . ). in univariate analysis, -year survival rate was significantly higher for pts receiving / donor ric-urd allo-sct in cr compared to / or / mm-urd (os: %, %, %, p ¼ . ; lfs: %, %, %, p ¼ . , respectively). however, among the cr and advanced disease groups there were no differences in outcome between fully matched or mm-urd ( / or / ) donor (cr þ : os %, %, %, p ¼ . ; lfs %, %, %, p ¼ . ; advanced-disease: os %, %, %, p ¼ . ; lfs %, %, %, p ¼ . , respectively). there was no difference in ri between the groups and nrm was higher after fully matched donor compared to mm-urd ( / or / ) only in pts with cr diseases (p ¼ . ). multivariate analysis showed higher nrm after / (hr . , p ¼ . ) compared to fully matched donor and no difference in nrm between / vs. / mm-urd. there was no difference in ri between / vs. / or / vs. / mm-urd donor. os and lfs were superior after fully matched donor vs. / mm-urd (os: hr . , p ¼ . ; lfs: hr . , p ¼ . ). however, there was no difference in adjusted os and lfs between / vs. / mm-urd ric allo-sct. chronic gvhd rate was not different between matched or mm-urd allo-sct groups. conclusion: despite the limitations of a retrospective registrybased study, our analysis shows no significant outcome difference between / and / mm-urd allo-sct after ric regimen in patient aged z year. in the absence of prospective data, we conclude that mm-urd ric allo-sct is a therapeutic option for acute leukemia pts not having fully matched donor. disclosure of interest: none declared. introduction: the use of minors as hsc donors is medically and legally accepted and is increasing. however, there is a lack of understanding of the physical and psychosocial effects of pediatric hsc donation. the goal of this investigation was to longitudinally investigate hrqol in this group. materials (or patients) and methods: participants were related pediatric donors (n ¼ ) who donated at domestic u.s. centers between / and / . data were collected from donors and their parents via structured telephone interviews at pre-donation, and weeks and year post-donation. a healthy age/gender matched pediatric sample was generated from existing data for normative comparisons. interviews gathered socio-demographics, psychosocial characteristics, and multidimensional hrqol using the well-validated pediatric quality of life inventory (pedsql) which produces a total score and physical, emotional, social, school and psychosocial subscores. t-tests were used to compare hrqol from donor self-report, parental proxy-report, and the normative sample across the three assessment points. mixed logistic models were used to examine the effects of pre-donation variables on post-donation hrqol. results: donors were - yrs (median ¼ yrs) and all but one were sibling donors. most parental respondents were mothers ( %; median age yrs), % were married, and % had at least a bachelor's degree. donor vs proxy. across all hrqol domains except emotional functioning and at all three assessment time points, donor self-reported hrqol was significantly lower than that reported by parental proxies. donor vs norm. at pre-donation, as compared to the normative sample, donors reported significant hrqol deficits across multiple subdomains and in total hrqol (t ¼ - . ,po. ). at weeks post-donation, donors reported deficits in physical functioning (t ¼ - . , po. ) and total hrqol (t ¼ - . ,po. ). at year post-donation, donors reported deficits in physical (t ¼ - . ,po. ) and school functioning (t ¼ - . ,po. ). donors at hrqol risk. across the three assessment time points, %, %, and % of donors respectively had self-reported pedsql total scores below the standard cutoff indicating significant clinical risk of poor hrqol -scores below the cutoff are similar to those of chronically ill children. sixteen percent, %, and % were below the cutoff at only one, two, or all three assessments respectively. the youngest donors ( - yrs) were at significantly greater risk of being below the cutoff than were their older counterparts with %, % and % of this group below cutoff at each of the three assessments respectively. in multivariable analyses, the pre-donation factor most strongly and consistently associated with below cutoff pedsql scores at weeks and year post-donation was pre-donation donor self-reported pedsql score (likelihood ratio: . ,po. ; . ,po. ). conclusion: these findings suggest that there may be significant hrqol deficits among pediatric hsc donors and that in this particular context, parents are not able to accurately report those deficits. these findings also indicate that research to identify predictors of poor hrqol and the development of interventions to screen and address hrqol deficits are urgently needed. disclosure of interest: none declared. introduction: in allo hct patients (pts) with disseminated adv disease, mortality is reported to be up to %. antiviral treatment (tx) usually consists of iv cidofovir (cdv), which has a significant risk of nephrotoxicity. bcv is an orally-available, lipid-conjugate of cdv with no evidence of nephrotoxicity in clinical trials. the pilot portion of the phase advise (cmx - ) study was initiated in march to enroll b allo hct and other immunocompromised adv pts with, or at risk of progression to, disseminated adv disease, to guide the final study design. as of nov , subjects have been enrolled and entered into the database, including allo hct pts ( with disseminated adv disease), solid organ transplant pts and ''other'' pts. preliminary safety and virologic results for the allo hct pts with disseminated disease are described. materials (or patients) and methods: all subjects receive open-label bcv mg (z kg) or mg/kg (o kg) twiceweekly for wks, extendable up to wks for pts at high-risk of relapse, and are followed for wks post-tx. adv dna viral load (vl) in plasma is measured using a quantitative pcr test (limit of detection [lod] log c/ml). results: baseline (bl) characteristics for the subjects are: median (range) age ( . , ) y, % o y; % male; median (range) plasma adv vl . (o lod to . ) log c/ml (n ¼ ); % adv positive by qualitative pcr in respiratory secretions, % in urine, % in stool; % with cmv in plasma, % ebv in plasma and % bkv in urine; % received prior iv cdv. as of nov , subjects had completed tx and had discontinued tx prematurely. the most common reasons for tx discontinuation were death (n ¼ ) and adverse event ([ae] n ¼ ). median (range) tx duration was ( , ) days (n ¼ ). virologic response in cdv-naïve and exposed subjects with detectable plasma adv vl at bl are summarized in the table. in subjects with positive adv pcr at bl, % ( / ) cleared adv in respiratory secretions, % ( / ) in urine and % ( / ) in stool. through dec , % ( / ) of allo hct subjects with disseminated adv disease had died, with a median -day observation period for living subjects. no death was attributed to bcv. aes leading to permanent tx discontinuation attributed to bcv were vomiting and abdominal pain in subject, and acute gvhd in subject. median (range) change in adv vl from bl (log c/ml) median (range) time to minimum on-tx (days) proportion ? log reduction in adv vl or to undetectable at nadir minimum on-tx last on-tx cdv-naive (n= ) - . (- . , . ) - . (- . , + . ) ( , ) % ( / ) cdv-exposed (n= ) - . (- . , + . ) - . (- . , + . ) ( , ) % ( / ) s conclusion: the observed mortality rate was % for allo hct pts with disseminated adv disease in advise, which is lower than literature rates reported for this pt population ( - %; ison , sandkovsky ). bcv showed potent virologic activity in cdv-naïve and exposed pts with no new safety concerns. these preliminary data support expansion of the pilot portion to a definitive phase study. introduction: the guidelines for immunization of hematopoietic stem cell transplant (hsct) recipients recommend doses of anti-pneumococcal conjugate vaccine (pcv) from - months after transplant, followed by a dose of polysaccharide -valent (ppv ) vaccine at months in case of no chronic graft-versus-host disease (gvhd), or an additional pcv dose in case of gvhd. however, due to lack of long-term data, there is no recommendation for boosts after months. our goal was to assess the retainment of anti-pneumococcal antibodies in allogeneic hsct recipients vaccinated years ago. materials (or patients) and methods: in , the idwp published the results of the idwp trial that compared the immune response assessed one month after doses of pcv , started either at , or at months after myeloablative hsct . additionally, all patients received dose of ppv at or months after transplant. all surviving patients had been assessed for anti-pneumococcal antibodies against the vaccine-serotypes months after hsct. this study was the basis of the current guidelines for anti-pneumococcal immunization after allogeneic hsct. the present study included surviving patients from the idpw , who were assessed for antibody levels against the pcv -antigens and against of the ppv antigens (pn and pn ), between . and years after transplant, i.e. to years after the last assessment in the initial study. the mean age was y ( - ), and / had acute leukemia. only had chronic gvhd (limited: , extensive: ) and had suffered a leukemia relapse. eleven ( %) had received an additional dose of ppv at a mean time of . years ( - years) after transplant, according to local procedure. the rates of persistent responses to all antigens of pcv were . % for an ab cut-off of . mg/ml, and % for a cut-off of . mg/ml. compared to the response rate at months after transplant, these rates were not significantly decreased but showed important serotype-specific variability. similar findings were observed for pn and pn antibody levels. neither the recipient or donor age, donor type, source of stem cells, gvhd, nor the administration of an additional dose of ppv (given to / patients) influenced the maintenance of the response. the timing of the initial vaccination was the only parameter influencing the long-term response; patients who were vaccinated lately after hsct (from months) had a significantly better maintenance of the response than patients vaccinated early (from months) after transplant. the patients who were not responders at months and who received an additional dose of ppv at , and months after transplant, respectively, did not respond. conclusion: in long-term hsct survivors without severe chronic gvhd vaccinated against s pneumoniae according to the current guidelines, the specific immunity is not fully maintained a decade later. patients, who received an additional ppv dose after months post-transplant, do not seem to benefit from this boost. boosts with pcv should be explored. so far, the optimal schedule of anti-pneumococcal vaccination in hsct recipients after months remains to be established. references : early cytomegalovirus reactivation -a potential factor for early robust t cell reconstitution and possibly a prognostic factor for agvhd after hsct p. r many previous studies have shown that agvhd puts patients at risk of cmv reactivation, most likely due to more intensive immunosuppression. however, recent studies and case reports also show that that cmv-r could be a risk factor for agvhd. here, we studied the effect of cmv-r on t cell reconstitution in patients with and without agvhd. materials (or patients) and methods: cmv r þ /d þ patients transplanted - in our institution were included in this study. all the patient samples were monitored for the cmv viral load (cmvpp expressing cells/ , leukocytes) and t cell reconstitution (cd , cd , cd and all available hla specific cmv tetramers for each patient) within the first days after hsct. patients were subdivided into five groups: group : no agvhd but cmv-r (no-agvhd-cmv-r), group : agvhd after cmv-r (agvhd-after-cmv-r), group : agvhd before cmv-r (agvhd-before-cmv-r), group : agvhd but no cmv-r (agvhd-no-cmv-r) and group : no agvhd-and no cmv-r (no-agvhd-no-cmv-r). results: the characteristics for onset of cmv reactivation and agvhd in the different subgroups are provided in table . cd , cd , cd and cmv specific t cells were analyzed on day ± days after hsct. in order to investigate the potential influence of cmv-r in the absence of agvhd on t cell reconstitution, we compared the t cell numbers in the groups cmv-r þ /-subsequent agvhd (i.e. group þ ) with group (no-agvhd-no-cmv-r). we found significantly more cd (p ¼ . ) and cd t cells (p ¼ . ) in groups þ compared to group . there were no differences in t cells between the groups with cmv-r þ /subsequent agvhd (i.e. groups þ ) compared to the groups with agvhd þ /-subsequent cmv-r (i.e. groups þ ). moreover, there were no differences in t cell numbers between the groups with agvhd þ /-subsequent cmv-r (i.e. groups þ ) compared to group . to study the impact of cmv-r on t cell reconstitution in patients with subsequent agvhd we compared the t cell numbers in group (agvhd-after-cmv-r) with group (no-agvhd-after-cmv-r). there were significantly more cd t cells (p ¼ . ) and a trend for more cd , cd and cmv-ctls in group (agvhd-after-cmv-r) compared to group (no-agvhd-after-cmv-r). subsequently, we compared the potential influences of cmv-r and of agvhd on t cell reconstitution. we found significantly more cd (p ¼ . ) and cd t cells (p ¼ . ) in group (agvhd-after-cmv-r) compared to group (agvhd-no-cmv-r). moreover, we studied the overall potential influence of cmv-r in the presence of agvhd on t cell reconstitution. we found a trend for more t cells (cd , cd , cd and cmv-ctls) in group (agvhd-after-cmv-r) compared to group (agvhdbefore-cmv-r). in conclusion, patients with agvhd after cmv-r had considerably more cd t-cells on day compared to patients with cmv-r but no agvhd and significantly more cd and cd t cells compared to patients with agvhd but no-cmv-r. these results suggest that early cmv-r enhances overall t cell reconstitution which could be a potential risk factor for developing agvhd after hsct. single and double cbt were used in and cases, respectively. tbi was part of the conditioning regimen in cases ( %). in vivo t-cell depletion by atg was used in % of patients. at least one pcr with a viral load log/ml of blood was sufficient to define hhv- reactivation after the graft. the impact of hhv- reactivation on cbt outcomes has been studied as a time-dependent variable. in multivariate analysis, hhv- was independently associated with graft failure (hr: . conclusion: our study confirms that hhv- reactivation is a risk factor for graft failure in cbt recipients after myeloablative conditioning regimen. this result has to be confirmed prospectively and in the setting of reduced-intensity conditioning cbt. this paves the way also to test prospectively the indication of ganciclovir or foscarnet use as anti-hhv- prophylaxis in cbt recipients. disclosure of interest: none declared. introduction: candida is the second more frequent cause of invasive fungal infection in haematological immunocompromised hosts, especially in the patients who undergo an haematopoietic stem cell transplantation (hsct). the aim of this study was to analyse retrospectively the outcome of patients with candida infections acquired in the first days after allogeneic hsct. materials (or patients) materials (or patients) and methods: in prospective study allohsct recipients were included from dec to jul . the median age was y.o., males - %. most of pts had high-risk acute leukemia ( %). allohsct from mud were performed in %, mrd - %, haplo - %, mmud - %, predominantly with ric ( %). eortc/msg criteria for diagnosis and response to therapy were used. since active diagnostic strategy, including bronchoscopy with bal, in pts with ct-scan lung lesions before allohsct has been introduced to the routine practice. ''active ia'' is the ia diagnosed just before hsct. results: incidence of ia before allohsct was , % (n ¼ / ). according to eortc/msg criteria % of pts had probable ia and % proven ia. the main sites of infection were lungs - %, central nervous system - %, and colon - %, other localizations were observed mostly in a combination with lung involvement: sinuses - %, spleen - %, and liver - %. antifungal therapy before allohsct was administrated in % pts (voriconazole - %, other - %) with the median duration of therapy - months. complete response to antifungal therapy was registered in ( %) pts, partial response or stabilization in ( %), and ''active ia'' in ( %) pts. after allohsct all pts received antifungal therapy with voriconazole (first line - %, continuation of treatment - %, and secondary prophylaxis - %). median length of treatment was days ( - ). cumulative incidence of relapse or progression of ia after allohsct was , % (n ¼ ). relapse of underlying disease was the main risk factor for the relapse or progression of ia after allohsct ( % vs %, p ¼ , ). progression of ia after allohsct was treated with voriconazole mg per day (n ¼ ) and combination vori þ caspo (n ¼ ). relapse of the ia after allohsct was treated with voriconazole mg per day (n ¼ ). no toxicity of the antifungal treatment was registered. complete response was achieved in pts, and stabilization - . -weeks overall survival (os) after the start of antifungal therapy was %. two pts died with the progression of the underlying disease. days os after allohsct was %, -year os after allohsct was %. there was no significant difference in os in pts with or without ia before allohsct. conclusion: incidence of the ia before allohsct was , %. cumulative incidence of relapse or progression of ia after allohsct in pts with proven and probable ia before allohsct was , %. relapse of the underlying disease was the main risk factor for relapse or progression of ia after allohsct. secondary prophylaxis with voriconazole should be used in pts with ia before allohsct. relapse or progression of ia after allohsct didn't impair os. ia is not a contraindication for allohsct. disclosure of interest: none declared. pt. patients had acute kidney injury which was managed conservatively without the need for renal replacement therapy. / survivors have normal lft's, patients have a residual mild increase in transaminases due to cgvhd, whereas patient has a moderate increase in lft's due to cgvhd. conclusion: busulphan based conditioning were the most important risk factor for vod. myelofibrosis had a strong trend towards causing vod (p- . ). early intervention with defibrotide along with supportive management was able to completely resolve vod in most of the cases and the day mortality was only / ( %). only death was directly attributed to vod and death each due to sepsis and biopsy proven drug induced liver failure. disclosure of interest: none declared. validation of two new prognostic scores to predict nonrelapse mortality in patients undergoing reduced-intensity conditioning allogeneic hematopoietic cell transplantation p. barba introduction: in , new pretransplant predictive models of non-relapse mortality (nrm) for patients undergoing allogeneic hematopoietic cell transplantation (all-hct) have been created based on modifications of the hct comorbidity index (hct-ci) and the ebmt score. the first model (hct-ci/ age) consisted of the addition of an extrapoint for patients years to the hct-ci (sorror et al. jco. ). the other model developed by the alwp of the ebmt combined categories of the hct-ci and the ebmt score into an integrated score (versluis et al. leukemia. ). none of these models have been validated in independent cohorts. materials (or patients) and methods: we analyzed the predictive capacity of these new models and compared it with the hct-ci and the ebmt score in a population of reducedintensity conditioning allo-hct (allo-ric) consecutively transplanted patients in spanish centers during an year period ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) . the scores of all models were calculated by a single investigator as originally defined. risk-groups stratification was also performed as originally defined, except for the ebmt score in which patients were divided into low-(score - ), intermediate-(score - ) and high-risk (scores ) according to percentiles , and . for the hct-ci/age patients scoring points (n ¼ ) were grouped with those scoring - points. the predictive capacity of all models was calculated by means of the harrell's c-statistic and were compared by calculating a z-score and p values from the estimated standard error. results: a total of patients were included. median age at hct was years (range - ). most patients received allo-hct from hla identical sibling donors (n ¼ , %) mainly for acute myeloid leukemia and myelodisplastic syndromes (n ¼ , %). median follow-up for survivors was . years (range . - ) . the median hct-ci/age and the alwp model scores were (range - ) and (range - ), respectively. the median hct-ci, ebmt scores were (range - ) and (range - ), respectively. risk group distribution of patients according to each model is summarized in introduction: systemic inflammatory response syndrome (sirs) is defined as an inflammatory state induced by infections or toxic damages. sirs is diagnosed when two or more of the following criteria are met: body temperatureo c or c, heart rate beats/minute, tachypnea breaths/minute or paco o mmhg, leukocyteso cells/ mm or cells/mm or presence of % immature neutrophils. the goal of this study was to assess the incidence of sirs early after an allogeneic stem cell transplantation (allosct) (from day to hematopoietic recovery) and evaluate whether sirs may influence the occurrence of acute s graft-versus-host disease (agvhd) and non-relapse mortality (nrm introduction: pure red cell aplasia (prca) after allogeneic hematopoietic stem cell transplantation (hsct) is a relatively rare complication after major abo incompatible allogeneic transplant. although reduced intensity transplant was a potential risk factor for prca, the impact of stem cell source has not been fully evaluated. we conducted a retrospective risk factor analysis for developing prca in major abo incompatible transplant including cord blood transplantation (cbt) and reduced-intensity conditioning. materials (or patients) and methods: we reviewed the medical records of adult patients who underwent allogeneic hsct for the first time at the toranomon hospital from to . prca after hsct was defined as anemia with low reticulocyte counts (o %) in peripheral blood for more than days after transplantation in association with neutrophil engraftment and a lack of erythroid precursors in bone marrow. results: one-hundred and sixty-three patients with major or bi-directional abo incompatibility who achieved neutrophil engraftment and survived more than days after hsct were included in this study. seventy four patients received reducedintensity conditioning, patients underwent cbt, did bone marrow transplantation (bmt) and did peripheral blood stem cell transplantation (pbsct). reticulocyte engrafted (reticulocyte ¼ %) in patients with a median time of days after hsct during this study, which was significantly longer after cbt compared to bmt/pbsct ( days vs. days, p ¼ . ). in patients, reticulocyte count remained o % beyond days post-transplant, of whom were diagnosed as prca with a cumulative incidence of . %. prca was not observed in cbt patients, and the cumulative incidence of prca was significantly lower after cbt compared to bmt/pbsct ( % vs. is the only curative treatment in fa. immune reconstitution after hsct is increasingly recognized as a critical determinant of morbidity and mortality in hsct. the aim of the study was to better understand the kinetics of immune reconstitution in children with fa who underwent allogeneic hsct after a fludarabine based reduced intensity conditioning regimen. materials (or patients) and methods: in this study, lymphocyte subgroups of children who underwent hsct were evaluated before hsct and , , , , and months after hsct. children with fa (n: ) comprised the study group and children with non-malignant diseases (n: ) comprised the control group. in addition to classical lymphocyte subgroups; activated t lymphocyte subgroups including cd / ( þ ), cd / ( þ ), cd /hla-dr( þ ), cd / ( þ ), cd / ( þ ) t lymphocytes were evaluated in study and control groups. results: when absolute levels of lymphocyte subgroups were evaluated in children with fa, cd ( þ ) lymphocyte count returned to pre-hsct levels at months. cd ( þ ) t lymphocyte count reached to pre-hsct levels at months. cd ( þ ) t lymphocyte count returned to pre-hsct levels at months. cd ( þ ) b lymphocyte count turned to pre-hsct levels at months. cd / ratio returned to pre-hsct levels at months. cd / ( þ )cd ( þ ) nk-t and cd / ( þ )cd (-) nk lymphocytes returned to pre-hsct levels within month after hsct. among hsct related complications; acute gvhd developed in / ( . %) children in study group and in / ( . %) children in control group. on the other hand, chronic gvhd developed in / ( . %) children in study group and in / ( . %) children in control group. when specific subgroups reflecting lymphocyte activation were evaluated in study and control groups; activated cd / ( þ ) nk lymphocyte and cd / ( þ ) t lymphocyte count returned to pre-hsct levels at months in both groups. while activated cd /hla-dr( þ ) t lymphocyte count returned to pre-hsct levels at months in both groups; activated cd / hla-dr( þ ) t lymphocyte count was higher at , , and months in control group. activated cd / ( þ ) t lymphocytes returned to pre-hsct levels at months in study group and those returned to pre-hsct levels after months in control groups. cd / ( þ ) activated t lymphocyte count was higher at months in study group (pr . ). cd / ( þ ) activated t lymphocytes reached pre-hsct levels at months in control group and at months in study group. besides, cd / ( þ ) activated t lymphocyte count was higher at and months in control group (pr . ). in this study, we show that the kinetics of recovery of the lymphocytes subgroups in children with fa after hsct follows those patterns also described for children with other diseases: early recovery of nk cells ( month), followed by effector cytotoxic t cells ( months) and b cells ( months) , and finally, cd ( þ ) t-helper cells ( months). high levels of cd /dr( þ ) activated t lymphocyte count at , , and months and high levels of cd / ( þ ) t lymphocyte count at and months in control group are attributable to the high frequencies of acute and chronic gvhd in control group than those of study group. disclosure of interest: none declared. introduction: although hla haploidentical hsct has been largely employed in children with life-threatening nonmalignant disorders, the survival of patients given this type of allograft has been reported to be inferior to that of patients transplanted from a compatible unrelated volunteer (uv). we implemented a novel method of ex vivo t-and b-cell depletion based on the selective elimination of ab þ t cells and b cells. we herein report an update of children with non-malignant disorders who were given this type of allograft. materials (or patients) and methods: twenty-two patients were males and females, median age at hsct being . years (range . - . ) . nine patients had severe combined immunedeficiency (scid), fanconi anemia (fa), severe aplastic anemia (saa), thalassemia major, hemophagocytic lymphohistiocytosis (hlh) and each immunedeficiency with polyendocrinopathy enteropaty x-linked (ipex), kostmann syndrome, hyper ige syndrome, osteopetrosis, swachmann-diamond syndrome and congenital amegakaryocytic thrombocytopenia (camt). all patients were transplanted from of the parents ( from the mother and from the father), the median number of cd þ and ab þ t cells infused being . x /kg and x /kg. the original conditioning regimen consisted of treosulphan and fludarabine (flu) þ thiotepa in ( scid, ipex, camt, kostmann syndrome and swachmann-diamond syndrome), flu and cyclophosphamide þ single dose tbi in ( fa and saa) and busulphan, flu and thiotepa in ( thalassemia, hlh, osteopetrosis and hyper ige syndrome). no patient received immunosuppression after hsct. all patients received fresenius rabbit atg ( mg/kg/day) on days - through - before allografting and rituximab ( mg/m ) to prevent ebv-related ptld on day - . results: all patients but engrafted, the median time to reach neutrophil and platelet recovery being days (range - ) and days (range - ), respectively. the patients ( with saa and each with thalassemia, fa, hlh and osteopetrosis) who had primary graft failure were successfully re-transplanted ( from the same parent, from the other relative and from an -hla locus disparate uv figure a ). in the latter group, agvhd started median days after hsct and was zgrade iii and steroid refractory in / cases. a higher agvhd rate was also observed in male patients who received a graft from a female donor (f- m mismatch) compared to all other sex-matches ( / vs. / , p ¼ . ). in multivariate cox regression analysis, both omission of mtx and f- m mismatch were associated with agvhd occurrence. in line with this, the protective effect of mtx was most pronounced in the subcohort of patients with a f- m mismatch: grade ii-iv agvhd occurred in / f- m mismatched non-mtx recipients whereas / f- m mismatched mtx recipients developed agvhd (p ¼ . ). all relapse occurred in / patients ( %). we did not observe differences in relapse rate between non-mtx recipients and mtx recipients ( / vs. / , p ¼ . , figure b) . although non-mtx recipients were more often transplanted for all in nd remission, pre-transplantation minimal residual disease (mrd) status did not differ between the groups. in line with earlier reports, mrd positivity was the most important risk factor for all recurrence. in mrd positive patients, the omission of mtx and agvhd occurrence did not prevent relapse after hsct. conclusion: mtx prophylaxis reduced the occurrence of severe agvhd, without compromising relapse free survival in pediatric all patients after t cell replete hla-identical bone marrow transplantation. prevention of agvhd reduces morbidity and the need for high-dose immunosuppressive agents, allowing for alternative immunotherapy-based therapeutic interventions in individuals at high risk for disease recurrence. introduction: hematopoietic stem cell transplantation (hsct) has contributed to improved outcome in childhood acute leukemia (al). however, post-hsct relapse is associated with a dismal prognosis and its optimal treatment remains unclear. we aimed to compare patients' related factors and treatment strategy, in case of relapse or progression post-allogeneic hsct in children with al in a recent ten-year period. materials (or patients) and methods: a total of children who received a first allogeneic hsct for all or aml from january to december experienced a relapse or progression thereafter. they were treated in the centers of the sfgm-tc, among them cases were analysable. primary endpoint was overall survival (os) after diagnosis of relapse or progression post first hsct whatever the treatment post relapse was. . lymphocyte (sub)populations were analysed frequently post transplantation by flow cytometry. cd þ t-cell recovery was defined as appearance ofz cells/ml, b-cell and nk cells recovery asz cells/ml. the median active atg serum concentration at time of t-cell or b-cell reappearance was . au/ml, the maximum level was au/ml. in % of the patients, nk cells re-appeared when the median active atg level was higher ( . au/ml) up to a maximum level of au/ml (see figure) . for alemtuzumab the median concentration at t-cell recovery was . mg/ml, max. . mg/ml, but in % of the patients nk cells reappeared at a higher concentration up to . mg/ml. for both drugs, t-cell recovery was significantly correlated with the serum level of the drug (po . for atg, and po . for c h), whereas a significant correlation was absent for nk cells. conclusion: atg and alemtuzumab are both able to deplete t-, b-, and nk cells. for both drugs, the exposure is highly variable even in patients with an equal weight receiving the same dose. here, we report that t-cell recovery is closely related with serotherapy exposure. nk-cells are the first cells that re-appear post hsct . all of the patients received the same ric regimen based on the use of fludarabine in combination with melphalan and antithymocyte globulin (atg). prophylaxis against gvhd was achieved via cyclosporine and methylprednisolone. results: all patients were engrafted. the median times to neutrophil and platelet engraftments were days (range: - ), and days (range: - ), respectively. patients underwent hsct from hla matched sibling donors (n ¼ ), full matched other related donors (n ¼ ), unrelated matched donor (n ¼ ) and unrelated mismatched donor (n ¼ ). the source of graft were peripheral blood (n ¼ ), bone marrow (n ¼ ) and cord blood (n ¼ materials (or patients) and methods: twenty-nine patients with miop were treated by hsct at the university childreń s hospitals in paris (n ¼ , since ) and ulm (n ¼ , since ). miop was caused by mutations in tcirg (n ¼ ), clcn (n ¼ ), snx (n ¼ ), rank (n ¼ ), and fermt (n ¼ ); age at transplant was between and months (median months); donors were haploidentical family donors (n ¼ ), mud (n ¼ ), phenoidentical relatives (n ¼ ), and msd (n ¼ ). thiotepa and serotherapy was added to the busulfan and fludarabine based regimen in all patients with donors other than msd. results: all but patients showed a primary and sustained engraftment; of patient, who rejected their haploidentical graft, could be rescued by a second graft from the second parent. severe vod, which had to be treated by ascites puncture, was seen in patients only. only one case of gvhd and no case of chronic gvhd was observed. cause of death in patients were liver toxicity in conjunction with cmv and fungal infection after prolonged aplasia (mud transplant at age of month) and complications in conjunction with engraftment failure ( introduction: bmt is the only proven curative treatment available for haemoglobinopathies. however, the number of patients who can benefit is seriously restricted by the lack of hla-matched related donors not suffering from the condition and the limited number of unrelated donors available for the ethnic groups in which these conditions are prevalent. in order to expand the donor pool, haploidentical transplantation with a post-infusion of stem cells cyclophosphamide approach has been developed for young adults, but whilst well tolerated it has resulted in relatively high rates of rejection and the need for a prolonged period of immunosuppression . materials (or patients) and methods: consecutive parental haploidentical transplants ( for sickle cell disease and one for b halassaemia major) were performed at st. mary's hospital, london, from june to november . the median age was . years of age (range to ). all patients lacked a suitable hla-matched related donor and an unrelated search had not identified a / or / donor. endogenous haemopoieis was suppressed with hypertransfusions, hydroxycarbamide mg/kg and azathioprine mg/kg for at least two months pre-transplantation. the conditioning included fludarabine mg/m , thiotepa mg/kg was added, cyclophosphamide mg/kg, tbi gy and atg (thymoglobulin) . mg/kg. gvhd prophylaxis was provided with cyclophosphamide mg/kg on days þ and þ , mmf and sirolimus. the minimum follow-up was days post-transplantation and half of the patients are days post-transplantation and have completed all treatment. the source of stem cells was g-csf primed bone marrow in all cases, aiming x tnc/kg. results: all patients engrafted, though one patient subsequently suffered secondary graft failure following macrophage activation syndrome and died. the median neutrophil engraftment was days (range to ). all surviving patients are cured from the manifestations of the original disease. none of the patients suffered vod, though infectious complications occurred at a higher rate than seen for related transplants for the same conditions at our institution. five patients had no evidence of acute or chronic gvhd. five patients developed stage acute gvhd (median presentation day þ , range to ) responding to topical steroids; one patient suffered skin gvhd stage on day þ and one patient gut gvhd stage on day þ , both treated with msc. all patients responded to first line treatment with no recurrence of disease. all patients but one achieved % donor fraction both in whole blood and t cells. one patient requires immunosuppression beyond day þ with stable % donor fraction in whole blood and % in t cells. introduction: bone marrow transplantation (bmt) offers a definitive cure for thalassemia in over % of low-risk children with a matched related donor. many centers currently incorporate thiotepa in busulfan-or treosulfan-based bmt regimens for thalassemia. this combination, however, may permanently impair fertilty in most patients. in the era of increasingly effective supportive care in which many thalassemia patients may have children, this is concerning. very longterm follow up studies have shown how the standard bu-cy regimen may be associated with birth rates comparable to the control population (la nasa et al. blood ). this study retrospectively compares bmt outcomes in two groups of low risk patients (defined as livero cm and ageo y) with severe thalassemia (st) (defined as a thalassemia syndrome with spontaneous hemoglobino g/dl), receiving oral busulfan ( mg/kg), cyclophosphamide ( mg/kg) and either thiotepa ( mg/kg) (tt-bu-cy) or rabbit atg (fresenius mg/kg or thymoglobulin mg/kg total doses from day - to - ) (atg-bu-cy) as preparative regimen. standard cyclosporine and short-term methotrexate plus low dose methylprednisolone were used for gvhd prophylaxis. materials (or patients) and methods: this is a retrospective multicentre comparative study of the safety and efficacy of substituting tt with atg in low-risk st patients undergoing matched-related bmt. between january and july , a group of patients were transplanted after conditioning with tt-bu-cy, while between august and july , patients were conditioned with atg-bu-cy. results: the actuarial overall survival in the tt-bu-cy and atg-bu-cy groups is . % and . %; thalassemia-free survival is . % and . % at a median follow up of . and . months respectively, with no statistically significant difference by logrank test. conclusion: substituting thiotepa with atg in the standard bu-cy context seems safe and effective. higher fertility rates are expected for patients on the atg-bu-cy regime. disclosure of interest: none declared. materials (or patients) and methods: data has been collected retrospectively from infants with scid who received transplants at different centers over a -year period . the differences between groups were compared by using chi-square or fisher's exact test, where appropriate. a p value of less than . was considered statistically significant. results: boys ( %) and (% ) girls with scid whose ages ranged between . - months (median months) at the time of diagnosis were transplanted. parental consanguinity was identified in ( %) of infants. % of the patients had received bcg vaccination before diagnosis. b þ and b-phenotypes were detected in . % (n ¼ ) and . % (n ¼ ) respectively, while ada deficiency was recognized in . %, rd (reticular dysgenesis) in . % of the cases. rag , jak , rag and artemis defects were the leading genotypes among the patients with molecular diagnosis ( . %; n ¼ ). out of , patients ( . %) had either a matched sibling or a family donor, while ( . %) and ( . %) children received haploidentical (mmfd) and mud transplants respectively. the hsct source was bone marrow in ( . %), peripheral blood in ( %) and cord blood in ( . %) of the patients. among a total of ( . %) retransplants, received a second transplant while children received a boost only. children survived, died and was lost to followup. the overall survival rate was . % over a years period. it increased from % ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) to % (p ¼ . ) during the latter years ( - ) and even to , % during the last years ( - ). the survival rates with relation to donor types were as follows: msd ¼ . % (n ¼ ), mrd ¼ . % (n ¼ ), haploidentical . % (n ¼ ) and mud % (n ¼ ). age at diagnosis significantly (r months or months) influenced the survival rate of the patients (p ¼ . ). immunophenotype did not seem to have an effect on survival rate and immunoglobulin (ig) requirement following hsct did not differ between b þ vs b-phenotypes (p . ). conclusion: this is the first multicenter study with the largest data obtained from scid patients transplanted in turkey. the median age at diagnosis was months, b-phenotype and rag were the most common among other defects. age at diagnosis ( months), and donor type (haploidentical) (po . ) were two major factors significantly related to poor outcome. expanded donor availability, advances in intensive care facilities, diversity of transplantation centers and specialized teams are among major factors contributing to the longterm outcomes of hsct. however, newborn screening is of paramount importance in ensuring early diagnosis and timely transplantation thus improving the survival of scid patients in turkey. disclosure of interest: none declared. oral session: stem cell mobilization & regenerative medicine haematology, bmt unit, hospital clínico universitario virgen de la arrixaca, imib, university of murcia, surgery, hospital clínico universitario virgen de la arrixaca, haematology, bmt unit, hospital clínico universitario virgen de la arrixaca, imib, university of murcia, murcia, spain introduction: amniotic membrane (am) is a non-tumorigenic tissue attributed with various biological properties (low immunogenicity, anti-inflammatory, anti-fibrotic and antimicrobial effects) related to its ability to synthesize and release cytokines and growth factors. am, that is usually discarded after birth, is in our experience an easily obtained tissue which processing, storage and management can be included in the daily routine of the cryobiology laboratory. ma can be used as a ''biologic bandage'' for healing management of chronic wounds in diabetic and non-diabetic patients. in our hospital there is an ongoing clinical trial to study the use of am to improve epithelialization (nct ). here, we describe the results obtained prospectively after the compassionate use of cryopreserved human amniotic membrane allografts in patients with chronic diabetic foot ulcers. materials (or patients) and methods: am was obtained from healthy mothers who had programmed an elective caesarean operation for obstetric reasons after they signed the informed consent. donors were screened by reviewing their medical records and by performing laboratory test to discard transmissible disease agents. am processing was done under sterile conditions in the gmp facility; the process involves: washing the am to eliminate blood traces, cut am into several fragments, sew each fragment on an impregnated dressing sheet and introduce them on cryopreservation bags adding the cryoprotectant solution based on human albumin, tc- medium and % dmso. the am fragments were storaged at - c and delivered once we the negative viral serology of the donor was confirmed months later. cryopreserved am was applied to six consecutive patients with diabetic foot ulcers under a compassionate use program of the diabetic foot unit from may to november . wound size reduction and rates of complete healing were evaluated. results: patients were aged between and years. they were affected by grade ii diabetic foot ulcers in the wagner ulcer classification scale that had failed previous treatments for periods between months and years. am was applied weekly or every ten days until complete healing or partial reepithelialization of the ulcers. a median of ( - ) cryopreserved am fragments were applied for an average treatment period of days. the mean size of the ulcer was reduced by %.l wreduced in size by %. at last follow-up, out patients have total epithelialization of the ulcer. no adverse events related to its application were observed. conclusion: our results show that the application of cryopreserved amniotic membrane is a feasible and safe treatment in complex diabetic foot ulcers. more rapid healing may decrease clinical operational costs and prevent long-term medical complications. furthermore, the treatment achieves re-epithelialization of long evolution wounds that were not reached with conventional therapies. disclosure of interest: none declared. bone as a regulator of human hematopoietic stem cell (hsc) trafficking: study of biochemical markers of bone remodeling and angiogenic cytokines during hsc mobilization, in patients with lymphoma and myeloma p. tsirkinidis ,* , e. terpos , g. boutsikas , a. papatheo-dorou introduction: bone is not considered just a structural, supportive tissue of bone marrow, but an hsc-niche regulator. data regarding the role of bone turnover in hsc mobilization in humans are scarce. the aim of the present study was to study bone remodeling and vessel equilibrium during hsc mobilization in lymphoma and multiple myeloma patients. materials (or patients) and methods: forty-six patients ( lymphoma and multiple myeloma) were studied. serum samples were collected at two time points: before mobilization (pre-mobilization sample) and on the day of hsc collection, which coincided with the peak circulating cd counts (collection sample). in / patients, additional serum samples were collected, between mobilization and collection. the following molecules were measured by elisa in patients' sera: ) bone resorption markers: carboxyterminal telopeptide of collagen type (ctx), aminoterminal telopeptide of collagen type (ntx), tartrate resistant acid phosphatase isoform b (tracp- b), ) bone formation markers: bone alkaline phosphatase (balp), osteocalcin (osc), osteopontin (opn), ) the osteoblastogenesis inhibitor dickkopf- (dkk- ) ) the osteoclastic regulators: receptor activator nf-kb ligand (rankl), osteoprotegerin (opg), ) angiogenic cytokines: angiopoietin- (angp ), angiopoietin- (angp ), angiogenin (ang). values were compared with non-parametric methods. patients who had either a collection of cd þ cells o . x /kg, or a circulating cd count peak o /ml were considered poor mobilizers. results: the comparison of the aforementioned molecules between the pre-mobilization and collection samples revealed: balp (p ¼ . ) and opn (p ¼ . ) increased significantly, while osc, a marker of bone turnover, and dkk- decreased significantly (p ¼ . and p ¼ . , respectively). these findings reveal a significant increase of bone formation during mobilization. rankl (p ¼ . ) and opg (p ¼ . ) increased significantly, leading to an increase of rankl/opg ratio (p ¼ . ), consistent with osteoclastic activation. however, there was no evidence of increased osteoclastic activity, as ctx decreased significantly (p ¼ . ), while both ntx and tracp- b did not change. angp- showed a dramatic reduction (p ¼ . ), while angp- increased (p ¼ . ), resulting in a significant decrease of the angp- /angp- ratio, a finding indicating vessel destabilization during mobilization. these results were further supported by the intermediate measurements, which showed a straightforward alteration of bone metabolism early in hsc mobilization. poor mobilizers had significantly higher ctx levels both at premobilization (p ¼ . ) and collection samples (p ¼ . ), higher ntx levels at collection (p ¼ . ), lower angp- premobilization (p ¼ . ) and higher osc at collection (p ¼ . ) compared to good mobilizers. thus, ctx, ntx s and angp- pre-mobilization levels may serve as reliable predictors of poor mobilization. conclusion: our study showed for the first time in humans, that bone plays a dynamic role during hsc mobilization: bone formation and vessel destabilization are the two major events and osteoblasts seem to be the orchestrating cells during this process. osteoclasts are stimulated, but not fully active. moreover, some of these markers may identify poor mobilizers. disclosure of interest: none declared. introduction: allogeneic bone marrow transplantation is a curative treatment for leukemia and genetic disorders. although some patients are cured of their underlying illness, they are at risk of developing potentially fatal graft-versus-host disease (gvhd). a recent phase randomized trial conducted by the canadian bmt group (n ¼ ) comparing the impact of g-csf mobilized peripheral blood (pb) to bone marrow (bm). a representative group of donor samples were evaluated and identified that high concentrations of cd bright nk cells in the donor product is strongly associated with a lack of development of both acute and chronic gvhd (odds ratio . ; p ¼ . ) in g-csf stimulated bm. we found that the cd bright nk population was cd (nkp ) positive with comparable expression of cd in both sub-populations consistent with regulatory nk cells (nk reg ). we hypothesized that alternate strategies utilizing the mobilizing agent, plerixafor, may enrich for cd bright nkp cells in pb thus reducing the risk of gvhd and circumventing the need of harvesting donor cells from the bm. materials (or patients) and methods: we performed a pilot study to examine the impact of plerixafor, that blocks the sdf- cxcr interaction, in mobilizing cd bright nk reg cells in the pb and bm, to determine the whether we can optimize a donor source with the highest potential for post transplant tolerance, resulting in a lack of both agvhd and cgvhd. we recruited healthy adult human volunteers. five subjects received one dose of plerixafor at micrograms/kg/day and pb and bm samples were harvested prior to, at , and hours after plerixafor administration. the subsequent participants each received daily doses of granulocyte colony stimulating factor (g-csf) at micrograms/kg/day for days, followed by a dose of plerixafor on day . pb and bm samples were collected prior to g-csf, after g-csf, and at and hours after plerixafor administration. we used multi-parametric flow cytometry to identify cd bright nk cells. the phenotype of cd bright nk cells was cd bright cd þ (nk þ ) cd -cd perforingranzyme b -. results: we observed a significant rise in pb nucleated cells at both and hours after plerixafor administration with a peak increase at hours. plerixafor alone induced a significantly higher proportion of cd bright nk reg cells in pb when compared to g-csf for days ( . vs. . fold, p ¼ . ) and g-csf for days followed by plerixafor treatment ( . vs. . fold, p ¼ . ). moreover, cells collected hours after plerixafor alone had a significant increase in the proportion of cd bright nk reg cells ( . vs. . fold, p ¼ . ) in pb compared to bm. this study suggests that plerixafor alone is able to mobilize a high number of cd bright nk reg cells in pb harvested after hours. we were not able to increase that population in a similar manner by using g-csf alone or g-csf plus plerixafor. these findings suggest that allogeneic donor mobilization of peripheral blood may give a product with a low rate of gvhd potentially superior to g-csf stimulated marrow and warrants further testing in a randomized clinical trial. disclosure of interest: none declared. introduction: only small studies have compared performance of the novel spectra optia apheresis system (terumo bct) with the widely used com.tec (fresenius healthcare) and the late cobe spectra (terumo bct) device in allogeneic stem cell collections. our collaborative working group compared performance data of all devices from two collection centers to analyze device-as well as center-specific performance parameters. materials (or patients) and methods: we analyzed firstday apheresis collections in g-csf-stimulated healthy donors that were performed in cologne (cgn) using spectra optia mnc (n ¼ ), cobe spectra mnc (n ¼ ), or com.tec (n ¼ ) and in dü sseldorf (dus) using spectra optia mnc (n ¼ ) or cobe spectra mnc ( ). peripheral blood and product samples were analyzed in center-specific laboratories. results: in both centers, irrespective of the apheresis device, similar yields of cd þ cells per kg donor bodyweight were reached. in cgn, collection rates (collected cd þ cells per kg donor bodyweight per cd þ cell count in peripheral blood before apheresis) were similar between the three apheresis systems. the continuously collecting cobe spectra yielded more cd þ cells over time (cr per h: , ± , ) than the discontinuously collecting apheresis systems ± , , po , ) and spectra optia mnc ( , ± , , po , ). the com.tec collected cd þ cells with highest efficiency (ce : ± % vs ± %, po , and ± %, po , for spectra optia mnc and cobe spectra, respectively). in comparison to cobe spectra ( ± min), procedure times were longer when using com.tec ( ± min, po , ) or spectra optia mnc ( ± min, po , ) systems. the product purity, measured as percentage of mnc was highest in products collected with the spectra optia mnc ( ± % vs ± %, po , vs ± %, po , for cobe spectra and com.tec, respectively). the relative differences between cobe spectra and spectra optia collection performance parameters were similar in the dus apheresis center: shorter procedures with higher cr per h for cobe spectra, higher mnc purity in spectra optia mnc products. absolute results differed between the two centers. conclusion: the highest collection efficiency (ce ) was seen when the com.tec device was used. however, this was accompanied by low product purity. compared to any other apheresis device that has been analyzed in this study the novel spectra optia allows collection of higher mnc purity apheresis products from allogeneic donors. however, this was associated with a significant prolongation of procedure timethe major disadvantage of this device. the cobe spectra collects more cells over time by allowing higher mean inlet flow rates despite inferior collection efficiency. therefore, a continuous collection procedure on the spectra optia device that allows high inlet flow rates would be an ideal collection setting in terms of collection efficiency and collection rate per unit of time. disclosure of interest: none declared. introduction: our group has previously established the safety and effectivity in terms of cd þ cell recovery and viability of the automated washing of cryopreserved hematopoietic progenitors (hp) with the sepax â device (biosafe) using normal saline supplemented with . % albumin (nsa), as well as the absence of infusion-related events and of a negative impact on engraftment dynamics (sánchez-salinas et al. ). in our present study we compare this solution with a ready-to-use free of human derived components solution: % hydroxyethyl starch / . in . % sodium chloride injection (voluven â , fresenius kabi). materials (or patients) and methods:: peripheral blood hp units apheresis cryopreserved using autologous plasma plus % dmso corresponding to autologous peripheral blood hp transplants were studied. after rapid thawing in a water bath at c, an automatic wash with the sepax ( washes cycle) was performed using either nsa ( units) or voluven ( units). nucleated cell levels determined by an hematology analyzer, flow cytometry cd þ cell counts, trypan blue cell viability and granulocyte macrophage (gm-cfu) and erythroid (e-bfu) colony forming cell cultures were performed on aliquots collected prior to and after the washing technique. statistical analysis was performed using descriptive statistics and a simple-measures anova. results: the mean total nucleated cell (tnc) and cd þ cell recovery was , % ± , , and , % ± , respectively for nsa and , % ± , and , % ± , for voluven. the mean gm-cfu and e-bfu cell recovery was , % ± , and , % ± , respectively for nsa and , % ± , and , % ± , for voluven. the mean viability recovery was . ± , for nsa and , ± , for voluven without differences between both solutions (p , ). there were no significant differences between both solutions in none of this parameters in spite of the tnc significant loss (p o , ), there were no significant differences between the pre and post-washing cd þ cell numbers (p ¼ , ), gm-cfu (p , ), e-bfu (p , ) or viability (p , ). in contrast with the % of untoward reactions recorded in our historical data of dmso containing cell infusions, we observed just three adverse effect with the washed cells with voluven ( , %) and none with nsa. one patient experienced a epileptic fit related to the infusion speed, another two suffered grade nausea and transient hipotension respectively. median time to neutrophil engraftment ( cells/ml) and platelet engraftment ( . cells/ml) for nsa were , ± , and , ± , days respectively and , ± , and , ± , for voluven. when comparing both solutions, there were no significant differences in neutrophil engraftment (p , ) or in platelet engraftment (p , ). conclusion: both nsa and voluven are equally effective for washing cryopreserved hp, ensuring a good cd þ cell recovery and preserving their viability and engraftment potential. both solutions avoid the dmso infusion related adverse events. as so, voluven constitutes an excellent alternative free of human-derived products and ready-to-use solution to our previous nsa standard washing solution. references: sánchez-salinas a. et al. transfusion. nov; ( ) : - disclosure of interest: none declared. introduction: ruxolitinib is a jak inhibitor that was recently approved for treatment of primary and secondary mf and shows impressive symptom control by suppression of inflammation. ruxolitinib is also a promising drug for treatment of acute and chronic gvhd. the immune-modulatory effects of ruxolitinib are at least in part due to an inhibitory effect on dendritic cell biology (heine et al., blood ) . dendritic cells (dcs) are important antigen-presenting cells. upon antigen contact they migrate into the draining lymph nodes to prime t cells. the aim of this study was to define the impact of the jak inhibitor ruxolitinib on dc migration. materials (or patients) and methods: cd þ cells isolated from human buffy coats were differentiated for seven days in the presence of gm-csf and il- to modcs and finally matured with lps. murine bone marrow-derived dcs (bmdcs) were generated by flushing bone marrow from femur and tibia of mice, plating the cell in medium containing gm-csf and maturing the cells with lps. migration of dcs was analyzed in transwell assays or dynamically by time-lapse microscopy within three dimensional collagen gels towards ccl- gradients. signaling events were analyzed by western blot to evaluate changes in phosphorylation levels. results: d migration of ruxolitinib-exposed dc is dosedependently reduced in vitro. by analyzing the migratory phenotype of human modcs within three dimensional collagen gels, ruxolitinib-exposed dcs are still able to sense chemokine gradients and form lamellipodia at the leading edge of the cell, whereas the retraction of the uropod is inhibited. additional in vivo studies could show that the jak inhibitor potently reduces homing of bmdcs into draining lymph nodes in mice. sirna knockdown experiments revealed that this inhibitory effect is jak -and jak -independent. on a molecular level we could show a reduced phosphorylation of rho-associated protein kinase (rock) in ruxolitinib-treated modc upon ccl- stimulation. finally, the migration phenotype of modc exposed to ruxolitinib could be mimicked by the rock inhibitor y- . conclusion: rhoa family members are key proteins controlling important steps of cell migration, such as protrusion formation at the front of the cell and consequently cell polarization. rock is a downstream effector of rhoa and leads to stabilization of the actin cytoskeleton via cofilin and actomyosin ii contraction by the myosin light chain. the observed reduction of rock phosphorylation may reveal an important mechanism of ruxolitinib-induced inhibition of dc migration. our current efforts focus on the validation of rock as offtarget jak / -independent target kinase of ruxolitinib as potential mediator of the observed effects, which may at least in part also explain the potential therapeutic impact of ruxolitinib for therapy of gvhd. introduction: alemtuzumab, a monoclonal cd -antibody, is used for t-cell depletion (tcd) in the context of allogeneic hematopoietic stem cell transplantation (hsct) to prevent graft versus host disease (gvhd). when we applied this approach in a clinical trial, nearly half of our patients developed acute gvhd (agvhd) overall grade i-ii . since regulatory t cells (treg) play a major role in controlling gvhd, we hypothesized that they might be functionally impaired in our patients after alemtuzumab based treatment and further investigated on these cells. materials (or patients) and methods: we analyzed peripheral blood samples of patients with agvhd, patients with chronic gvhd, and patients who never developed gvhd after alemtuzumab-mediated tcd. treg were identified as cd þ cd þ cd þ cd -or cd þ cd þ cd þ foxp þ and subsets described by expression of cd . since cd is linked to the membrane by a glycosylphosphatidylinositol (gpi)-anchor, we used flaer to stain for gpi-anchors themselves. to further investigate treg activation, we stained additional markers: cd , cd , gitr, cxcr , ccr , ctla- , garp and granzyme. to observe how treg reconstitute after hsct, we analysed samples from patients with agvhd at different time points after transplantation and correlated our findings with the clinical course of gvhd. treg function was evaluated in cfse-suppression-assays: patient derived ex vivo treg were facs-sorted according to their cd expression and incubated with proliferating cfsestained cd -effector t cells from healthy donors. reduced cd -proliferation was used as indirect marker for the suppressive function of the cd positive and negative treg. results: patients with agvhd showed significantly elevated percentages of cd negative treg: mean . % (range . - . % ) in comparison to only . % (range - . %) in patients with chronic our without gvhd. by flaer-staining we confirmed that loss of cd correlates with the absence of gpi-anchors on the cell surface, these cells do also lack other gpi-anchored proteins (e.g. cd , cd ). patients who overcame agvhd over time reconstituted gpianchor positive treg, whereas gpi-anchor negative treg remained the dominant population in patients with ongoing agvhd. the fraction of activated garp positive treg was mainly detected among the gpi-anchor positive treg population. all other markers showed a heterogeneous expression profile with a tendency towards lower expression on gpi-anchor negative treg. suppression-assays showed a higher suppressive capacity of gpi-anchor positive ex vivo treg in contrast to gpi-anchor negative treg of the same patient. conclusion: cd negative / gpi-anchor negative treg reconstituted in patients after alemtuzumab mediated tcd and mainly persisted in patients with acute gvhd. these treg were less likely to express the activation marker garp. functional assays demonstrated that gpi-negative treg were functionally impaired -in patients developing acute gvhd, these impaired treg represented the major treg-population. our preliminary data suggest that gpi-anchor negative treg, like other gpi-anchor negative t-cell subpopulations (previously shown), are functionally altered. we hypothesize that this might promote acute gvhd. these gpi-anchor negative treg could be useful to diagnose and guide immunosuppressive therapy in patients with acute gvhd. disclosure of interest: none declared. introduction: we previously showed that enhanced human invariant nkt (hinkt) lymphocytes recovery after allogeneic stem cell transplantation was associated with a reduced risk of acute graft versus host disease (agvhd). using a humanized mouse model of gvhd, we aimed to determine whether hinkt cell subsets could be involved in the regulation of allogeneic immune response and to elucidate their mechanisms of action. materials (or patients) and methods: human inkt were obtained by in vitro expansion of total peripheral blood mononuclear cells (pbmc) from healthy donors followed by electronic sorting of cd þ and cd -inkt subsets among the cd þ cd d-tetramer positive total inkt population. xenogeneic gvhd was assessed in gy irradiated nod/scid/ gamma-c -/-(nsg) mice previously injected with hpbmc alone or hpbmc enriched with cd -(pbmc þ inkt -) or cd þ (pbmc þ inkt þ ) hinkt cells. the effects of hinkt lymphocytes on the survival and phenotype of human monocyte derived dendritic cells (hmo-dc) were assessed by flow cytometry (on the expression of annexine v, propidium iodide, and that of cd , cd , cd , respectively). the effects of cd or cd þ hinkt cell subsets on the expression of the t cell activation marker cd and on the cytokine expression profile of cd þ t lymphocytes was assessed in vivo in the nsg model and in vitro, during the allogeneic mixed lymphocyte reaction (mlr). results: in vivo, nsg mice transplanted with pbmc þ inkt cells showed a significantly prolonged overall survival in comparison to nsg mice transplanted with pbmc alone (p ¼ . ) or with pbmc þ inkt þ inkt cells (p ¼ . ). improved survival observed with hpbmc þ inkt cells was associated with lower clinical and histological gvhd scores. in vitro, at inkt/ dc ( n) ratio, hcd -inkt cells significantly increased the apoptosis of mature hmo-dc compared to the cd þ inkt subset (p ¼ . ). hmo-dc apoptosis in the presence of cd -inkt cells was contact dependent ( % in contact vs. % in transwell, po . ), occurred as early as hours after cell-contact and was associated to the degranulation of cd -inkt cells as shown by cd expression. while h inkt cd lymphocytes could inhibit the maturation of hmo-dc in vitro, their cd þ counterpart strongly induced high levels of expression of cd , cd and cd on mo-dc in a contact dependent way. when allogeneic mlr was performed, in the presence of cd -inkt lymphocytes, proliferating alloreactive t cd þ lymphocytes showed a lower median fluorescence intensity of cd (p ¼ . ), and intracellular inf-g (p ¼ . ) il- (p ¼ . ) and il- (p ¼ . ) in comparison to mlr without addition of inkt cells. compared to nsg mice injected with pbmc alone, mice injected with pbmc þ inkt cells showed lower cd expression on circulating t cd lymphocytes (p ¼ . ), and reduced intracellular expression of il- on circulating t cd lymphocytes (p ¼ . ). conclusion: these results are in line with our clinical observations and suggest that h cd -inkt cell subset could directly modulate the allogeneic immune response by downregulating antigenic presentation and subsequently t cell activation and th /th cytokine production. the nsg preclinical mouse model suggest that developing a cellular therapy based on inkt cd lymphocytes might be interesting for the prevention of human agvhd. disclosure of interest: none declared. in vivo expansion of host type regulatory t cells via a selective tnfr agonist protects from acute gvhd medicine ii, wuerzburg university hospital, pathology, wuerzburg university, wuerzburg, germany introduction: cd þ foxp þ regulatory t cells (tregs) suppress graft-versus-host disease (gvhd) after allogeneic hematopoietic stem cell transplantation (allo-hct). by controlling the magnitude of the alloresponse tregs still allow for efficient anti-leukemia (gvl) effects of transplanted conventional t cells in preclinical mouse models. current clinical study protocols for donor tregs in the treatment or prophylaxis of gvhd rely on their ex vivo expansion and infusion in high numbers. here we present a fundamentally novel strategy for inhibiting gvhd based on the in vivo expansion of recipient tregs prior to allo-hct, exploiting the crucial role of tumor necrosis factor receptor (tnfr ) in treg biology. materials (or patients) and methods: to expand tregs in vivo we developed a highly selective tnfr agonist (that would not bind to tnfr ) and treated allo-hct recipients two weeks before allo-hct. expansion of radioresistant host type tregs, alloresponses of conventional t cells, and tumor progression were monitored with bioluminescence imaging, fluorescence microscopy, and flow cytometry in different mhc major mismatch mouse models (c bl/ , h- b balb/c, h- d and fvb/n, h- q c bl/ , h- b ) of gvhd and gvl. results: in vitro, this new tnfr -agonist expanded natural tregs from wild type but not from tnfr -deficient mice. accordingly, a human variant of this tnfr -specific agonist expanded human tregs in vitro. in vivo treatment of healthy mice with the murine tnfr -agonist significantly increased treg numbers in secondary lymphoid organs and peripheral tissues, particularly in the gastrointestinal tract, a prime target of acute gvhd. consequently, pre-treatment of recipient mice with this novel tnfr -agonist expanded host-type radiation resistant tregs prior to allo-hct in two models across mhc barriers. tnfr agonist pre-treatment resulted in significantly prolonged survival and reduced gvhd severity when compared to tnfr -deficient recipients or untreated allo-hct recipients. this was accompanied by reduced donor t cell proliferation and infiltration into gvhd target organs. in vivo depletion of host derived tregs completely abrogated the protective effect of tnfr -agonist pre-treatment. while in vivo tnfr -agonist pre-treatment protected allo-hct recipients from gvhd, antitumor effects of transplanted t cells remained unaffected in two different murine b cell lymphoma models. conclusion: our novel strategy demonstrates that the expansion of host tregs by selective in vivo tnfr -activation significantly improves the outcome after allo-hct and results in prolonged tumor-free survival. disclosure of interest: none declared. introduction: we reported that mir- expression is upregulated in donor t cells during agvhd and mice receiving mir- knock-out (ko) donor splenocytes do not exhibit lethal gvhd and have improved survival as compared to mice receiving wild type (wt) splenocytes. while we showed that mir- does not affect the alloreactive or homeostatic proliferative potential of t cells, a significant decrease in the expression of the homing receptors ccr , cxcr , and s p were found on mir- -ko t cells, suggesting that the loss of mir- could impair the migration of these cells to the peripheral target organs resulting in less agvhd. here, we further investigate the impact of mir- expression in t cell migration and elucidate the t cell population responsible for agvhd modulation. materials (or patients) and methods: lethally irradiated balb/c or b d f recipients were infused with t cell depleted bone marrow (bm) cells ( x ) and gfp expressing mir- ko or gfp-b wt t cells ( x ). transplants with cd þ ( x ) only t cell subsets were performed to identify the lymphocyte population that contributes to the mir- mediated effects on agvhd. results: on days , and post transplant, recipient mice were sacrificed, and tissues harvested in order to study the kinetics of mir- ko t cell migration following allogeneic hsct. there was a dramatic decrease in t cell infiltration of peripheral organs (peyer's patches, liver, lung and skin) in recipients of mir- -ko t cells as compared to wt t cells as evidenced by confocal microscopy of gfp labeled donor cells. we reasoned that these effects could be due to the modulation of ccr and other chemokine receptors by mir- . we found that a recently described long noncoding rna (lincr-ccr - as) located in the vicinity of several chemokine receptor encoding genes including ccr , ccr and ccr and that is involved in chemokine regulation and t cell migration has conserved potential mir- binding sites. we then set out to determine if mir- regulates the expression of this lncrna in relationship with ccr expression. there was a significant decrease in ccr mrna expression in mir- -ko versus wt donor t cells obtained from recipient mice at the time of agvhd, but no significant difference in the levels of lincr-ccr - as. a luciferase reporter assay, however indicate that there was an interaction between mir- and lincr-ccr - as. our result does not exclude the possibility that mir- might influence the activity rather than lincr-ccr - as levels, a hypothesis that is currently being tested. to identify the lymphocyte population involved in mir- mediated modulation of agvhd we performed a b into f transplant using cd þ t cells alone as the source of donor t cells. median survival of recipients of t cell depleted bm þ wt cd þ t cells (n ¼ ) was days as compared to % survival of all recipient mice of mir- ko cd þ t cells (n ¼ ) on day , (p ¼ . ). recipients of mir- ko cd þ t cells exhibited also significant lower agvhd clinical (po . ) and pathological scores (po . ) than wt recipients. conclusion: our data suggest that mir- may exert its modulating effects in agvhd by affecting t cell migration. our results also point to the cd þ t cell subset seems to play an important role in the mir- regulation of agvhd. experiments are currently underway to determine the role of mir- in regulatory t cells and cd þ t cell subsets in the modulation of agvhd. disclosure of interest: none declared. oral session: solid tumors introduction: hdct has a recognized indication in the salvage setting of advanced gct and is steadily utilized worldwide. while the prognostic impact of response to prior lines of ct (i.e. definition of chemoresistance) is ascertained, that of response to induction/mobilization ct preceding single or multiple hdct cycles is unknown. we present the results of a retrospective study of the ebmt- stwp. materials (or patients) in the multivariable model for pfs, tumor primary (overall p value ¼ . ), igcccg category (overall p value ¼ . ), and progression to induction (hr: . , %ci: . - . , p ¼ . ) were significantly prognostic. the latter was also significantly prognostic for os (hr: . , %ci, po . ) together with the transplant setting (overall p value ¼ . ). results for response to induction ct were confirmed in the models that included missing data. the c-index of the model was . for pfs and . for os. conclusion: progression to induction ct prior to hdct was independently and significantly associated with shorter pfs and os, while response or progression to prior ct lines was not. this information could have important implications to refine patient eligibility to transplantation and enhance the prognostic risk grouping. these data need external validation. disclosure of interest: none declared. kir-ligand incompatibility in the graft-versus-host direction improves progression-free survival in patients with primary high risk neuroblastoma after umbilical cord blood transplantation with nonmyeloablative conditioning y. takahashi introduction: donor nk cells expressing inhibitory killer cell immunoglobulin-like receptors (kirs), which do not recognize their cognate ligands ('kir ligands') on recipient targets, are free from hla inhibition, resulting in a decreased incidence of relapse and improved the outcome after hla haploidentical stem cell transplantation (hsct) (ruggeri, blood ) or umbilical cord blood transplantation (ucbt) (willemze, leukemia ) in leukemia patients. venstrom reported that significant association between kir/hla genotypes predictive of missing kir ligands have a better outcome without allogeneic hsct following anti gd monoclonal antibody therapy in patients with high risk neuroblastoma (venstrom, clin cancer res ). these observations led us to start the clinical trial of allogeneic ucbt from kir ligand incompatible donor for patients with high risk neuroblastoma in . materials (or patients) and methods: eligibility criteria of this study were newly diagnosed stage iv neuroblastoma patients with one of the following ) chemo-resistant disease defined as mibg positive score after courses of induction chemotherapy, ) older age defined as years old and older at diagnosis or ) mycn amplification defined as greater than copies per tumor cell. kir ligand mismatched ucbt donor was prospectively selected from the japan cord blood bank network according to the genotyping of hla-c or b as previously reported (willemze, r. leukemia ). we scheduled ucbt with reduced intensity conditioning regimen about three months after conventional high dose chemotherapy followed by autologous peripheral blood stem cell transplantation. reduced intensity conditioning regimen for cbt was flu, l-pam mg/m and tbi gy. tacrolimus and methotrexate were used for gvhd prophylaxis. single inhibitory kir expressed nk cells with no corresponding recipient's hla were monitored by flowcytometry before and after ucbt to access the expansion of alloreactive nk cells in vivo. we retrospectively analyzed the outcome of patients with high risk neuroblastoma treated in nagoya university hospital between april and june . results: fifty four patients were treated before dec. . after this study started in jan. , patients underwent kir ligand incompatible cbt who match eligibility criteria ( chemo-resistant, mycn amplification and older age) and patients received standard treatment. all patients achieved engraftment in ucbt group and no patients developed grade iii or more acute gvhd and chronic gvhd. only two patients died in this group because of bu related lung toxicity. surprisingly, no patient in this group relapsed with the median follow-up period of ( - ) months. on the other hand, cumulative incidence of relapse in others was . %. year progression free survival in cbt group was significantly better than in others ( . % vs . %, p ¼ . ). single inhibitory kir expressed nk cells significantly expanded after cbt (p ¼ . ). finally, multivariate analysis revealed only kir ligand incompatible cbt and stage iii were significant better covariate factors for relapse. conclusion: this is the first report that kir ligand incompatible allogeneic ucbt significantly reduced the incidence of relapse in high risk neuroblastoma. disclosure of interest: none declared. better prognosis for brca-mutated breast cancers treated with high-dose chemotherapy and autologous hematopoietic progenitor cell transplantation. a singleinstitution retrospective study l. boudin , , , * , a. goncalves , , , j. m. extra , , r. sabatier , , h. sobol , , f. eisinger , , j. moretta , , c. tarpin , , j. camerlo , , b. calmels , , c. lemarie , , a. granata , , , , f. bertucci , , , a. madroszyk , , p. viens , , , c. chabannon , , introduction: hereditary brca genetic mutation is responsible for approximately - % of breast cancers (bc).the objective of this study was to compare the outcome of brca-mutated (brca mut ) versus brca wild type or unknown (brca wt/uk ) bc following treatment with high dose chemotherapy (hdc) and autologous hematopoietic progenitor cell transplantation (act). materials (or patients) and methods: all female patients treated for breast cancer (bc) with hdc and act at institut paoli-calmettes between and were included. patients were divided into groups depending on the indication of hdc with act: metastatic breast cancer (mbc) or high risk breast cancer (hrbc) including inflammatory breast cancer (ibc) and/or massive lymph node involvement (lni). all patients were examined for the presence of known brca or mutations. information regarding patient, tumor characteristics and treatment was also collected. the main objectives were the analysis of overall survival (os) and progression-free survival (pfs) according to the brca mutation status in the two groups. survival curves were generated using kaplan-meier method and compared using log-rank test. results: a total of patients were included, mbc and hrbc ( lni and ibc). among mbc and hrbc, ( brca , brca ) and ( brca , brca ) patients were brca mut , respectively. in mbc, median age was respectively (range - ) and years (range - ) for brca mut and brca wt/uk ; % of brca mut patients had triple negative (tn) bc subtype (her -negative and hormonal receptor-negative) versus only % in brca wt/uk individuals. in brca mut and brca wt/uk median number of metastatic sites was and , with % and % visceral metastases, respectively. in hrbc, median age for brca mut and brca wt/uk was (range - ) and (range - ), respectively; % of brca mut had tn bc subtypes versus , % of brca wt/uk . ibc represented % of hrbc in brca wt/uk and % in brca mut . in mbc, % (all except one) of brca mut remained alive at years; the median overall survival (os) was not reached, compared with a median os of . years for brca wt (ci % ¼ . - . ), (p ¼ . log-rang test); median progression free survivals (pfs) were , months (cl % ¼ in brca mut patients versus , months (cl % ¼ . - . ) , and in brca wt/uk patients (p ¼ . log-rank test). in hrbc, years probabilities of os were respectively (all patients alive) vs for brca mut and brca wt/uk individuals, while years probabilities of disease free survival (dfs) were ) and , respectively (p ¼ . log-rank test). conclusion: the prognostic impact of brca mutation in bc remains controversial. in this series of mbc and hrbc, we have reported an outstanding survival outcome in a small subset of patients with documented brca mutation. in spite of a higher proportion of tn and more aggressive features, brca mut bc had a better outcome than their brca wt/uk counterpart, the difference reaching statistical significance in mbc population. hypersensitivity to dna-damaging agent, possibly due to defect in homologous recombination associated with brca mutation, could explain these results. despite the low numbers of cases in our series, hdc with act, may be considered as an option for brca mut , women with advanced bc. disclosure of interest: none declared. introduction: paclitaxel-based regimens are now commonly employed for second or third-line salvage therapy for gct. this might have an impact on the results with subsequent salvage hdct in these patients (pts). the ebmt-stwp is sponsoring a retrospective study on the outcomes of hdct administered in the last years. hence, we aimed to study outcomes with hdct after relapse to paclitaxel-ct to identify the level of chemoresistance in these pts. materials (or patients) and methods: data have been collected from ebmt registry, including european centers. eligibility included adult male patients (pts) with gct, and treatment with second or further-line hdct between the years and . both paclitaxel used in prior ct lines of therapy and in induction-mobilization regimens pre-hdct were considered. multivariable cox regression analyses (mva) were undertaken to evaluate the association of prespecified factors (site of tumor primary, igcccg category, response to induction ct, response to prior lines [chemosensitive vs chemorefractory], line of hdct, year of hdct) including prior-paclitaxel therapy with progression-free (pfs) and overall survival (os). results: since / , pts have been registered, of them were suitable for present analysis. pts ( %) received hdct in second-line, ( %) in third and ( %) beyond the third-line. hdct regimens were as follows: ( %) hd-cbdca-vp (ce), ( %) adding ifosfamide (cei), ( %) other mixed regimens. ( %) were taxane-containing. pts received a single hdct course, pts double and multiple courses ( missing). median follow up was months (iqr: - ). prior paclitaxel was significantly associated with shorter os in the univariable model (p ¼ . ). however, on mva prior paclitaxel-therapy was not significantly prognostic for both pfs and os, as shown in the table. a separate model evaluated the interaction between prior paclitaxel-therapy and taxane-containing hdct: no significant interaction was found (p ¼ . . - . . conclusion: the administration of paclitaxel-based regimens before hdct did not affect pfs/os. results were confirmed when excluding pts who were administered taxane-containing hdct. line of hdct was not significantly prognostic too. there is no evidence to disallow patients who have been treated with taxanes in second or third line to receive hdct as futher salvage therapy. these data need external validation. disclosure of interest: none declared. favorable outcome of a cohort of metastatic breast cancer patients treated with high dose chemotherapy and autologous transplantation at a single institution does not change the prognostic significance of histopathological subtypes l. boudin introduction: studies of high dose chemotherapy (hdc) in breast cancer (bc) often lack of biomarker information, notably the human epidermal growth factor receptor (her ) status. the objective of this study was to evaluate the outcome of patients affected with different subtypes of bc following treatment with hdc and autologous hematopoietic progenitor cell transplantation (act). materials (or patients) and methods: all female patients treated for metastatic breast cancer (mbc) with hdc and act at institut paoli-calmettes between and were included. patient, tumor and treatment characteristics were collected. patients were categorized in three subtypes based on hormonal receptor (hr) and her status of the primary tumor: luminal (l), (hr þ /her -), her (her þ , any hr), and triple negative (tn) (her -and hr-). main study endpoints were overall survival (os) and progression-free survival (pfs) categorized by bc subtypes. treatment related mortality (trm) was also evaluated. survival curves were generated using kaplan-meier method and compared using log-rank test. results: a total of mbc patients are included; median age was (range - ); metachronous and synchronous mbc were % and %, respectively. % patients received hdc with act as part of their first-line treatment following diagnosis of metastases. median number of metastatic sites was (range - ), including % of visceral metastases. l, her and tn subtypes were , and % of patients, respectively. all but one her patients ( %) received trastuzumab during the metastatic phase of the disease before ( %) and/or after ( %) oral session: acute leukemia introduction: the outcome of patients years old with all is poor with no clear recommendations regarding the indications for hct. these patients are usually considered ineligible for myeloablative allohct and offered transplantation with reduced intensity conditioning (ric). autologous hct is an alternative. however, the role of both treatment options has not been established so far. the aim of this study was to retrospectively compare results of ric-allohct and autohct in all years old and to identify factors affecting outcome. materials (or patients) and methods: patients treated with ric-allohct from either hla-identical sibling (n ¼ ) or matched unrelated donor (n ¼ ) and treated with autohct in first complete remission between and have been included in this analysis. median age in both groups was ( - )y and ( - )y, while median interval from diagnosis to hct was . months and . months, respectively. the proportion of ph( þ ) all among those with reported cytogenetics was % and %, respectively. results: with a median follow-up of months, the probability of os at two years was % for ric-allohct and % for autohct (p ¼ . ), while lfs rates were % and %, respectively (p ¼ . ). relapse incidence at two years was comparable for ric-allohct and autohct ( % vs. %, p ¼ . ) while non-relapse mortality was significantly reduced for autohct ( % vs. %, respectively, p ¼ . ). the advantage in favor of autohct was significant for ph(-) all (os: % vs. %, p ¼ . ; lfs: % vs. %, p ¼ . ) while no significant differences could be observed for ph( þ ) all (os: % vs. %, p ¼ . ; lfs: % vs. %, p ¼ . ). in a multivariate analysis adjusted for recipient age and gender as well as interval from diagnosis to transplantation the use of autohsct was independently associated with reduced risk of mortality (hr ¼ . , p ¼ . ), treatment failure (hr ¼ . , p ¼ . ) and non-relapse mortality (hr ¼ . ; p ¼ . ) with no effect on relapse incidence (hr ¼ . , p ¼ . ). in the ric-allohsct subgroup lfs was negatively affected by female donor/male recipient combination (hr ¼ . , p ¼ . ). lfs rates for both sibling and mud transplants were comparable ( % vs. %, p ¼ . ). the use of peripheral blood cells compared to bone marrow was associated with reduced risk of relapse (hr ¼ . , p ¼ . ). in the autohsct setting there was a tendency to higher risk of treatment failure with increasing recipient age (hr ¼ . , p ¼ . ). other variables including type of conditioning (tbi-based vs. chemotherapy-based) did not affect survival in any of the study cohorts. conclusion: considering poor overall prognosis of all patients years old, results of both ric-allohct and autohct appear enhancing and both types of transplantation may be considered valuable treatment options. potential advantage of autohct as suggested by results of our analysis should be further explored including data on disease-related prognostic factors and the status of minimal residual disease. disclosure of interest: none declared. introduction: the karyotype of the leukemic cells at diagnosis is one of the strongest prognostic factors in acute myeloid leukemia (aml). but the major cytogenetic risk categorizations were based on the large clinical studies of conventional chemotherapy for aml. in this retrospective study, we analyzed the influence of the cytogenetics of aml at diagnosis on the outcomes of allogeneic hematopoietic cell transplantation (hct). materials (or patients) and methods: from the database of jshct, we extracted the data of adult patients with aml who receive first hct between and . a total of , recipients were included. additional survey for the recipients who had been reported to have cytogenetic abnormalities at diagnosis (n ¼ , ) confirmed the data of karyotype of , cases. results: cytogenetics at diagnosis were categorized into (a) normal karyotype ( . %), (b) inv( ) or t( ; ) ( . %), (c) t( ; ) ( . %), (d) q abnormality ( . %), (e) complex karyotype ( ¼ abnormalities) ( . %), (f) monosomal karyotype (mk) ( . %), and others. overall survival (os) and cumulative incidence of relapse at years of all cases were . % and . %, respectively. these recipients were classified into groups: favorable (fav) included (b) (n ¼ ); intermediate (int), (a), (c), t( ; ) and others (n ¼ , ); unfavorable- (unf- ), (d) except t( ; ), and (e) except (f) (n ¼ ); unfavorable- (unf- ), (f) (n ¼ ). adjusted os at years were . % in fav, . % in int, . % in unf- and . % in unf- ( figure) . risk of overall mortality (rom) and risk of relapse (rr) were compared among these four groups, adjusted by age, sex, disease status at transplantation and stem cell source. in fav group, rom (hr . , , p ¼ . ) and rr ( . , . - . , p ¼ . ) were significantly lower than in int group. on the other hand, compared with int group, recipients in unf- and unf- group had significantly (po . ) higher rom (unf- . , . - . ; unf- . . - . ) and rr (unf- . , . - . ; unf- . , . - . ). these results showed that the karyotype of leukemic cells at diagnosis was also one of the powerful prognostic factors in hct for aml. but the impact of each cytogenetic presentation on the outcomes of transplantation may have some differences from that of chemotherapy. to improve this risk stratification system, molecular abnormalities of aml at diagnosis are also considered for the influence on outcomes of hct. disclosure of interest: none declared. late intensification with autologous hsct after nonmyeloablative beam conditioning has shown to be effective approach in adults with t-cell acute lymphoblastic leukemia treated by non-intensive protocol: results of the rall study group e. parovichnikova ,* , l. group is conducting the all- trial where the main principle is non-intensive but non-interruption treatment and autologous hsct after beam followed by maintenance in t-cell all pts without hla-identical donors (ebmt , op- ). the autologous hsct was planned for all patients as late high dose consolidation (on the - weeks of the protocol). in patients with early and late t-cell immunophenotype having hla-identical siblings allogeneic bmt was an option. from nov, , till nov, , centers enrolled all patients. in phenotype was unknown ( , %). b-cell precursor phenotype was diagnosed in , % (n ¼ ), t-cell precursor phenotypein , % (n ¼ ); biphenotypic al -in , % (n ¼ ). t-cell all patients were young - y ( - ); male gender prevailed - f/ m. t-all subtypes distribution in our study was as follows: ( , %) patients had early t-all (t-i/ii), ( , %)thymic (t-iii), ( , %) -mature (t-iv). for the whole group medians were: hb- g/l ( - ), l - , * / l ( , - ), plt - * / ( - ), b/m blasts - , % ( - ), ldh - iu ( - ). no born marrow involvement was detected in pts, b/m blasts - % -in (so t-lbl - , %). cytogenetics was available in , % of pts (n ¼ / ), , % of them (n ¼ / ) had normal (nk), pts -abnormal karyotype, pts ( %) had no mitosis. mediastinum involvement was registered in / ( , %) and cns disease -in / ( , %). results: induction and follow-up data were available in pts. cr was achieved in / ( , %): after prephase ¼ , after st ind.phase ¼ , after nd ind.phase ¼ . induction death occurred in n ¼ ( , %). primary resistance and even progression during induction was registered in n ¼ ( , %). pts proceeded to autologous hsct, almost all (n ¼ ) were successfully harvested at a median time of weeks. pts were poor mobilizers and bone marrow exfusion was carried out. no data on mrd level at time of harvesting is available so far. auto-hsct was applied at a median months from cr. allogeneic hsct from sibling donors was performed in cr pts ( with ti/ii and with t-iv). no deaths in allo-hsct group have occurred. the land mark analysis at months for chemotherapy group (n ¼ ) and at day of hsct for transplanted groups was done. disease free survival constituted for chemotherapy - % at years, and % -in both transplanted groups. none of the transplanted patients relapsed so far. the overall and diseasefree survival in the whole cohort of t-cell all patients was % and %. in a multivariate analysis only autologous hsct influenced dfs. conclusion: our study demonstrates that t-all may be treated by non-intensive, but non-interruption approach. even without high dose consolidation -years os and dfs (land mark analysis) constituted %. autologous hsct with beam conditioning after mild induction/consolidation followed by prolonged maintenance seems to add benefit to the overall optimistic results decreasing the relapse rate from % to % within years. disclosure of interest: none declared. with a median follow up from time of hla typing of months (range, . - ) for all patients and months (range, - ) for survivors, the -and -year probability of survival for all eligible aml patients was ± % and ± %, respectively. at the same time points, for the transplanted patients the probability of survival from time of graft was ± % and ± %, respectively. the probability of survival was particularly dismal for the patients transplanted in advanced disease phase ( ± % at yrs), whereas for the patients transplanted in early (cr þ cr ) phase the -yrs probability of survival was ± % and, by excluding the low number of cb recipients, it was ± % for hla id sib, ± % for hrd and ± % for mud recipients (p ¼ ns). conclusion: rtn policy allows a donor identification in % of all evaluable aml patients and provides an allogeneic transplant to % of them with no substantial differences in s terms of long-term survival between initially eligible and definitively transplanted patients or by comparing the different donor stem cell sources. transplant results should be given following information on the specific transplant policy and only the itt analysis allows to know the real impact of each transplant program. disclosure of interest: none declared. materials (or patients) and methods: oral pan was administered in two sequential schedules, either thrice weekly (tiw) every week (a) or every other week (b). arm a, in which pan was started at a dose of mg tiw and escalated to mg tiw using a þ design was followed by arm b, in which pan doses increasing from mg tiw to mg tiw were investigated. pan was initiated between day þ and þ after hsct and given for up to year. eligibility criteria included: ancz , /ml, plateletsz , /ml, adequate organ function and no severe gvhd. dlt was defined as prolonged g hematologic toxicity or any non-hematologic toxicityzg unrelated to disease progression within days of the first pan dose. results: pts ( aml, mds), median age yrs (range, - ), are evaluable for mtd. cytogenetics were classified as low (n ¼ ), intermediate- / (n ¼ ) or adverse risk (n ¼ ) according to eln criteria. pan was started a median of days (range, - ) after hsct from a mrd (n ¼ ), mud (n ¼ ) or mismatched donor (n ¼ ) which was performed in active disease (n ¼ , median bone marrow blasts %, range, - ), cr (n ¼ ) or cr (n ¼ ). the rp d was mg tiw in arm a and mg tiw every other week in arm b based on dlts: fatigue g at mg, colitis and nausea/emesis g each at mg (arm a), diarrhea and headache g at mg each (arm b). pan-related or unrelated g / aes were reported in of pts ( %) and included hematologic toxicity ( %), laboratory alterations ( %), infections ( %), constitutional ( %) and gastrointestinal symptoms ( %) and neurologic, pulmonary, pancreatic/hepatobiliary or vasculary toxicity ( - % each). aes were rapidly reversible after interruption and/or dose reduction (n ¼ ) and no study-related deaths occurred. fifteen pts ( %) have completed one year of pan, pts discontinued treatment prematurely after - days due to aes (n ¼ ) or relapse (n ¼ ). prophylactic or preemptive dlis ( - ) were administered to pts ( %). eleven pts ( %) developed mild (n ¼ ), moderate (n ¼ ) or severe (n ¼ ) chronic gvhd. patients died of aml (n ¼ ) or severe chronic gvhd (n ¼ introduction: our previous research has provided the evidence that an autoreactive immune system can be ?reset? into a healthy, tolerant state by immunoablative treatment to eradicate pathogenic effector cells, followed by transplantation of hematopoietic progenitor cells (hsct). here, we present the clinical and serologic responses and long-term immune reconstitution in patients with severe ads for up to years after receiving immunoablation and asct. materials (or patients) and methods: since , patients with refractory autoimmune diseases (sle, n= ; ssc, n= , ms, n= ; polychondritis, n= ; panniculitis, n= , granulomatosis with polyangiitis, n= , and chronic inflammatory demyelinating polyneuropathy, n= ) received a cd + -selected autologous stem cell transplantation after immunoablation with atg (neovii, formerly fresenius) and cyclophosphamide as part of a prospective monocentric phase i/ii clinical trial. autoantibody titers were evaluated with elisa, peripheral tand b lymphocyte subsets immunophenotyped using multicolor flow cytometry. results: with a median follow-up of years after immune reset (range, months to years), of patients ( %) achieved a progression-free survival (pfs), defined as survival without major organ failure. % of these patients showed durable clinical remissions for up to years despite discontinuation of immunosuppressive treatment, while % of patients had a stabilization of their underlying disease under maintenance therapy. disease relapse occurred in three sle patients (at , , and months, respectively), two of whom died later from uncontrolled disease and infection, respectively. of patients died from infection (n= ) or cardiac failure (n= ). anti-dsdna antibodies completely normalized in all sle patients and ana significantly declined from a median titer of : at baseline to : six months after transplantation in patients with connective tissue diseases. recovery of recent thymic emigrants (rtes) was completed between and months after immune reset, reaching on average . to . times the baseline levels. recurring foxp + tregs had significantly higher expression levels for cd and cd ra compared to age-matched healthy controls, indicating their recent thymic origin. numeric recovery of the naÿve b cell compartment was completed within year after immune reset in responding patients. conclusion: these data confirm our assumption that the reprogramming of an autoreactive immune system into a juvenile and self-tolerant immune system is feasible and associated with long-term remissions. our findings propose that chronic autoimmunity is not an end point depending on continuous treatment with specific anti-inflammatory agents, but may be cured by combining specific targeting of autoreactive memory and effector cells with a reactivation of thymic activity. a future challenge is to make this therapeutic approach attractive for a larger number of patients by reducing the rate of severe infections. this may be achieved by either using a more selective graft purging, e.g., depletion of t cell receptor alpha/beta and cd + cells from apheresis products with the clinimacs device, or an adoptive transfer of microbe-or virus-specific memory t and/or b cells. disclosure of interest: none declared. in the present report we summarize the long term effects of transplantation. materials (or patients) and methods: the mobilization protocol included cyclophosphamide and g-csf. the conditioning consisted of cyclophosphamide ( mg/kg/day on days - ,- ,- , - prior to transplantation) and antithymocyte globulin (thymoglobulin -of . mg/kg/day given on day - , and . mg/kg/day given on days - ,- , - and - ) except for three patients, who received atg fresenius at adjusted doses. results: one of the transplanted patients died during neutropenia due to the sepsis and its complications. the mean time of observation of remaining patients as of december was months (range - months). the independence of exogenous insulin after the transplantation was achieved in of them. three patients were lost to follow up. the median time without exogenous insulin for patients in follow up was months (range - months). four out of ( %) remain in remission of diabetes (exogenous insulin free) with median follow up of months (range - months). notably, three patients in the series were treated with atg fresenius (due to transient problems with thymoglobulin availability) -the median follow-up of these patients is months with months of remission of diabetes. the treatment led to significant reduction of hba c and good glycemic control in patients. no attempt was made to repeat procedure in relapsing patients. the hsct leads to remission of new onset t dm in majority of patients but the response in most of them is limited to average of months. patients receiving atg fresenius tended to have longer remission but small number prevented drawing conclusions. there is a need to develop procedure to either prolong response or to effectively treat the relapse of diabetes. disclosure of interest: none declared. hematopoietic stem cell transplantation for refractory crohn's disease: feasibility and toxicity j. introduction: autologous hematopoietic stem cell transplantation (hsct) is a salvage treatment for severe refractory crohń s disease (cd) patients. we evaluated feasibility and toxicity of autologous hsct for refractory crohn's disease (cd). materials (or patients) and methods: in this prospective study, refractory cd patients with an aggressive course despite medical treatment, impaired quality of life and no surgical options were included. hematopoietic stem cells were mobilized with cyclophosphamide and g-csf and collected by leukapheresis from pheripheral blood. in a second step, conditioning regimen with cyclophosphamide and rabbit antithymocite globulin (ratg) was used and previously collected stem cells were infused. toxicity and complications during the procedure and within first-year after transplantation were evaluated, along with the impact on safety after the introduction of supportive measures over the study. results: twenty-six patients were enrolled. during mobilization, patients presented febrile neutropenia, including bacteremia and septic shocks. neutropenia median time after mobilization was days. five patients withdrew from the study after mobilization and patients entered into the conditioning phase. hematopoietic recovery median time for neutrophils ( . x /l) was days and for platelets ( x /l) was days. ninety-five percent of patients suffered febrile neutropenia and patients presented worsening of perianal cd activity during conditioning. among non-infectious complications, patients presented antithymocyte globulin reaction, patients developed mucositis and patients had hemorrhagic complications. changes in supportive measures over the study, particularly antibiotic prophylaxis regimes during mobilization and conditioning, markedly diminished the incidence of severe complications. during the first -month follow-up, viral infections were the most common observed complications, and one patient died due to systemic cytomegalovirus infection. conclusion: autologous hsct for refractory cd patients is feasible but extraordinary supportive measures need to be implemented. we consider that this procedure should only be performed in highly-experienced centers. disclosure of interest: none declared. autologous haematopoietic stem cell transplantation in aggressive multiple sclerosis: a uk cohort from two centres j. clay ,* , p. . (range . - . ) , whilst the remaining had a deterioration of . - . materials (or patients) and methods: cb units (cbu) were frozen and thawed across the participating banks, according to local sops; pre-freezing and post-thaw samples were assessed for clonogenic potential (cfu) and cd þ recovery and viability through standard ishage protocol (si) (brocklebank am ). a modified ishage gating strategy (mi) was developed by reduction of the debris gate and by extension of lympho-monocytes gate in order to detect cd þ events with abnormal physical parameters (cd þ gate). facs sorting was performed to collect cd þ /cd þ events included inside (p ) and outside(p ) the si cd gate. sorted cells were then characterized by fc, confocal microscopy analysis and cfu assay in order to confirm that cd þ cells detected out of the si cd gate are not an artifact. confocal microscopy analysis was performed by labeling with a cd ab recognizing a different epitope and marked with a nuclear counterstain. results: si and mi strategies showed no significant discrepancies when determining cd þ absolute number and viability in pre-freezing cbus. the recovery of viable cd þ cells assessed by si or mi in the thawed products did not show any differences statistically significant, whilst the recovery of total cd þ cells was significantly lower in si ( , ± %) than in mi ( , ± . %)-analyzed samples (mean±sd, po , with pair t test). this difference resulted in a lower cd þ viability with mi than si ( , % ± . vs , % ± %, respectively, po . ). fc post-sorting analysis showed that the sorted p (si cd gate) population was cd dim cd þ cd þ cd - -aad-, whereas p (mi extended cd gate) showed the same markers expression but a -aad þ staining, therefore determining such events as cd þ dead cells. this data was confirmed by confocal microscopy analysis. finally, p events showed a % clonogenic efficiency, whereas p events were not able to generate any colony. conclusion: a cd þ count after cbu thawing lower than the fresh sample is often reported, usually with a high rate of viability. we showed that the standard methodology for cd þ counting leads to the exclusion of a number of dead cd þ cells, thus resulting in a viability overestimation. we propose here a modification of the ishage counting standard for thawed samples, which provides a more reliable assessment of cd þ viability. the content of pdc in bm grafts had a significant beneficial effect on post-transplant survival, an association not seen in gpb grafts . the % improved survival seen in recipients of more donor bm pdc was due to decreased deaths from graft rejection and acute and chronic gvhd. donor bm pdcs in murine bmt favorably modulated t-cell activation, decreasing gvhd through gamma-interferon-dependent upregulation of ido, while preserving gvl effects . the present study tested whether the biological activity of pdc from different graft sources are explained by the differential expression of homing receptor molecules or immuneregulatory pathway genes. materials (or patients) and methods: facs isolated pdc from allogeneic stem cell grafts obtained from bm, gpb, umbilical cord blood (ucb) and following mobilization with plerixafor (plxpb) samples. expression of chemokine receptor, integrin and selectin were measured by flow cytometry. gene expression on flow cytometry-sorted pdc was analyzed with illumina chips. differentially expressed genes using a -sided t-test comparing groups of replicate samples with po . were selected for exploratory analyses of regulatory pathways. mhc mismatched mouse h b-h k transplants compared outcomes with pdc from ccr ko vs wt donor. indirect presentation of allogeneic peptides by pdc and classical dc was studied using h b-h b/d transplants with h kd peptide recognition by tea transgenic donor t-cells. results: pdc from bm and plxpb grafts had higher cd l expression but lower ccr and cxcr expression than pdc from gpb and ucb grafts. high cxcr and low ccr expression were seen in pdc from all graft sources. while differences in ccr expression between pdc from bm vs gpb grafts suggested homing differences might explain observed differences in trm associated with the pdc content of the allograft, donor pdc from ccr ko mice had equal ability to modulate gvhd as pdc from wt mice. pdc in grafts from both human and mice were phenotypically immature and were relatively ineffective in activating t-cells through indirect presentation of alloantigen peptide, indicating that immune regulatory effects might be more important than antigen presentation. supporting this hypothesis, gene expression patterns of human pdc from bm showed lower expression levels of genes related to adaptive immunity, and higher [o ] expression of genes associated with innate immunity, than pdc from gpb, ucb, or plxpb grafts. conclusion: functional differences in immuneregulatory genes explain some of the observed differences in transplant outcome when stratifying recipients based upon between the content and source of donor pdc. bm pdc appear to be polarized towards activation of innate immunity while pdc from other graft sources are more polarized towards antigen presentation. novel approaches of stem cell mobilization, such as plerixafor, may increase the content of immature pdc enriched for expression of that might be effective in recapitulating the beneficial effects of bm pdc with respect to preventing early posttransplant transplant related mortality. references : results: with a median follow-up of , and months in the / urd, / urd and ucb, hematopoietic recovery was slower in ucb compared to urd recipient: the cumulative incidence of neutrophils recovery at day and platelet recovery  /l at day were respectively % and % in ucb against % and % in / urd and % and % in / urd (po . ). while there was no significant difference in the day cumulative incidence of grade - agvhd: % in / urd, % in / urd and % in ucb, the years cumulative incidence of cgvhd was significantly lower in ucb recipients: % against % in / urd (po . ) and % in / urd (p ¼ . ). in multivariate analysis, there was no statistically significant difference in nrm between ucb recipients and / recipients (hr . , p ¼ . ) or / recipients (hr . , p ¼ . ) . in multivariate analysis, the cumulative incidence of relapse/progression was significantly lower in / urd recipients compared to ucb recipients (hr . , p ¼ . ) and there was also a trend towards a decreased incidence of relapse in the / urd recipients (hr . , p ¼ . ). compared to ucb transplants there was no significant difference in pfs after / urd (hr . , p ¼ . ) and / urd (hr . , p ¼ . (table ) . results: there were no significant differences in pre-transplant characteristics between both groups ( table ) . the day cumulative incidence of neutrophil engraftment was similar in both groups ( % vs %), with a median time of and days, respectively (p ¼ . ). similarly, the -day cumulative incidence of platelet engraftment was % vs %, in a median time of and days, respectively (p ¼ . ). four cases among the haplo-cord group showed primary cb graft failure ( of them showing third party donor cells engraftment) who were rescued by a second cb transplant in two cases and haplo-sct in two cases. there were no graft failures in the haplo-sct group. grade ii-iv acute gvhd was more frequent in the haplo-sct group ( % vs %, p ¼ . ) and chronic gvhd showed a higher tendency in the haplo group ( % vs %, p ¼ . ). with a median follow-up of months ( - ) in the haplo-cord group and months in the haplo-sct group, os was % and % (p ¼ . ), respectively. efs was % vs % (p ¼ . ), relapse rate was % vs % (p ¼ . ) , and trm was % vs % (p ¼ . ), respectively. conclusion: in our experience, albeit differences in follow-up and limited number of patients, myeloablative single cord supported by hla-mismatched cells and haploidentical transplantation with post-transplant cyclophosphamide both offer valid alternatives for patients with acute leukemia. engraftment rates are similar in both groups as well as relapse rate. early toxic mortality is higher in the haplo-cord group while gvhd rates seem to be higher in the haplo-sct patients. further analysis including larger series and longer follow-up are needed to confirm these observations. disclosure of interest: none declared. introduction: studies on conditioning regimens prior to autologous stem cell transplant (asct) in lymphomas generally present a major pitfall, the miscellaneous mix of histologies composing the series, resulting in a reduced statistical power when focusing on a specific subset. we previously demonstrated (visani et al, blood ) the safety of a new conditioning regimen with bendamustine, etoposide, cytarabine, and melphalan (beeam) prior to asct in resistant/relapsed lymphoma patients (eudractnumber - - ). the regimen showed long-lasting significant antilymphoma activity, with a -year pfs of %. materials (or patients) and methods: we thus designed a single histology, phase ii study to evaluate the efficacy of the beeam conditioning in resistant/relapsed diffuse large b-cell non-hodgkin lymphoma (dlbcl) patients. the study was registered at at european union drug regulating authorities clinical trials (eudract) n. - - . the primary endpoint of the study is to evaluate the -year complete remission rate. fixing the lowest acceptable rate as % and the successful rate as %, with a significance level a ¼ . and a power -b ¼ . , the sample size was estimated in patients. results: until now, patients (median age years, range - ) were enrolled. at the time of writing, we have data available for patients with resistant/relapsed diffuse large b cell non-hodgkin lymphoma were consecutively enrolled in the study. briefly, / patients had advanced stage disease (iii-iv), were primary refractory and had relapsed after a median number of lines of therapy (range: - ). eleven patients had or more relevant comorbidities (range: [ ] [ ] [ ] [ ] [ ] . patients were in ii or subsequent cr after salvage therapy, whereas were in pr and had progressive disease. a median number of . x cd þ /kg cells (range , - , ) collected from peripheral blood was reinfused to patients. all patients engrafted, with a median time to anc . x /l of days. median times to achieve a platelet count x /l and x /l were and days respectively. ten out of patients presented a fever of unknown origin ( %), whereas patients ( %) presented a clinically documented infection. all patients received g-csf after transplant for a median time of days (range: - ). one patient died due to an incomplete hematological recovery after transplant, producing an overall transplant related mortality of . %. thirtyeight out of patients are evaluable up to now for response to treatment. / ( . %) obtained a cr, / a pr, whereas / did not respond to therapy. after a median follow-up of months after transplant (range - ), / patients were refractory, / relapsed, and / ( %) are still alive, in continuous cr. conclusion: in conclusion, this data provide evidence that the beeam regimen is safe and has promising efficacy in resistantrelapsed aggressive diffuse large b cell lymphoma. pts with hl at risk of relapse post-asct (clinicaltrials. gov #nct ). the primary results showed that bv significantly improved progression-free survival (pfs) per independent review vs. placebo. efficacy analyses by investigator review were performed in subgroups as these data may provide useful information for making treatment decisions. materials (or patients) and methods: the aethera trial is a phase , randomized, double-blind, placebo-controlled study. after asct, pts received bv . mg/kg q wk or placebo for up to cycles. pts were enrolled in of high-risk categories: refractory to frontline (fl) therapy, relapse o mos after fl therapy, and relapse z mos after fl therapy with extranodal disease. the primary endpoint was pfs per independent review. pfs by investigator was also evaluated. results: pts were randomized at sites in the us and europe. median age was yrs (range, - ) and % were male. asct conditioning regimens were beam: %, cbv: %, or other regimens: %. % of pts received radiation as part of their transplant regimen. all pts had completed the treatment phase as of aug . subgroup analyses by demographics, stratification factors, pt characteristics, and number of risk factors showed that pfs by investigator was improved across all groups; the hazard ratio comparing bv to placebo was o for all analyses (table ) . in an ad hoc analysis, pts with increasing numbers of risk factors for progression had progressively greater pfs benefit with bv vs. placebo. the most common adverse events (aes) in the bv arm (vs. placebo) were peripheral sensory neuropathy ( % vs. %) and neutropenia ( % vs. %). a higher incidence of herpes simplex and zoster infections were observed with bv ( %) vs. placebo ( %); otherwise, opportunistic infections were rare and balanced between the arms. the incidence of pulmonary toxicity (by meddra smq) was low, but slightly higher with bv vs. placebo ( % vs. %). two pts died r days of last dose of bv; cause of death was acute respiratory distress syndrome. response to salvage therapy pre-asct cr . ( . , . ) pr . ( . , . ) sd . ( . , . ) hl status after frontline therapy refractory . ( . , . ) relapse o months . ( . , . ) relapsez months with extranodal disease . ( . , . ) treatments pre-asct . ( . , . ) b symptoms after failure of frontline therapy . ( . , . ) extranodal disease at time of pre-asct relapse . ( . , . ) risk factors a z . ( . , . ) z . ( . , . introduction: although high-dose chemotherapy (hdc) is the gold standard for the treatment of many relapsed or refractory lymphomas, the outcome is still unsatisfactory in some subsets of patients with adverse prognostic features. we treated high-risk patients with a tandem strategy associating debulking with hdc followed by autologous stem cell transplantation (asct) and subsequent allogeneic sct (tandem auto-allo) materials (or patients) and methods: adult patients consecutively treated at two centers were included. criteria for receiving tandem auto-allo were: hl and nhl refractory to first-line therapy, less than cr after first salvage treatment, relapse after prior asct, multiple relapses, histology of transformed follicular, mantle-, t-and nk-cell lymphoma and documented infections during hospital stay in ( %), without significant differences between beam and hd-mel groups. among the patients with active disease before asct, the overall response rate was % (n ¼ responders) and those in cr were ( %). no difference was observed in terms of response in the beam and hd-mel groups (p ¼ . ). allogeneic sct donor was either hla-identical (n ¼ ), mud / (n ¼ ), haploidentical (n ¼ ) or cord blood (n ¼ ). -y os of entire cohort was % ( % ci: - ), -y pfs was % ( - ), rates of acute gvhd grade - and chronic gvhd were % and % respectively. trm rate was % (n ¼ ). no difference between beam and hd-mel group was observed for os ( % and % respectively, p ¼ . ) or trm ( % and %, p ¼ . ). of note, os of patients in cr before and after asct and os of those obtaining cr after asct was superimposable ( figure ) figure) . a multivariate analysis revealed that late allo-hsct was a unfavorable prognostic factor for os with a statistical significance (hazard ratio, . ; % ci, . - . ; p ¼ . ( ) or complete response after autosct after a third line of chemotherapy ( pts). donors of allosct were hla identical siblings in % of patients, matched unrelated in % and haploidentical donors in % of patients. median age at transplant was (range, - ). at allosct, % of patients were in cr, % were in pr, % in sd or pd. % patients performed allosct having relapsed o months from autosct or with primary refractory disease, % having relapsed months after autosct. results: median follow-up of surviving patients was . years (range, . - . ) . overall survival (os) was % at year, % at years and % at years of follow-up. in multivariate analysis, only disease status before allosct significantly impacted the os (p ¼ . , hr . , ci % . - . ) , whereas donor and timing or relapse/refractoriness did not change os (p ¼ . , hr ¼ . , and p ¼ . , hr ¼ . , respectively better control of the disease: this has occurred together with changes in donor type (from unrelated to family haploidentical), in gvhd prophylaxis (from mtx cya atg to post transplant cyclophosphamide), and in the conditioning regimen, combining thiotepa and intravenous busulfan; in addition dipss was somewhat lower in more recent patients, and also spleens were smaller. we believe these data suggest that alternative donor grafts are currently a therapeutic option for patients with mf. disclosure of interest: none declared. long follow-up data show that survival rate after allogeneic stem cell transplantation in patients with cll is higher for younger patients because of significantly lower nrm during the whole follow-up period: a retrospective study of the cmwp m. van gelder ,* on behalf of cmwp . among pediatric pts, % were aged - months, % were aged - y, and % were aged - y. day þ survival data by age group are available for pts post-hsct and pts post-ct (table) , with rates of svod post hsct of % in pediatric pts and % in adults. hpse has been shown to be involved in inflammation and may therefore play an important role in the pathogenesis of vod of the liver. the purpose of this study was to identify an association between hpse gene single nucleotide polymorphisms (snps) and vod of the liver after allogeneic hsct in childhood. materials (or patients) and methods: children (median age, years) who underwent allogeneic bone marrow (n ¼ ) or peripheral blood stem cell transplantation (n ¼ ) in a single center and their respective donors were genotyped of hpse for rs and rs using taqman real-time polymerase chain reaction. the donor was hla-matched unrelated in % of transplants and hlaidentical related in % of transplants. conditioning regimen was myeloablative in all cases and based on busulfan in % of transplants or total body irradiation in % of transplants. two forms of post-transplant immunosuppression predominated, cyclosporine a and methotrexate in % of transplants and cyclosporine a alone in % of transplants. results: donor/patient pairs were analyzed. cell samples from the patient were available in cases and from the donor in cases. genotype ag of hpse rs snp was found in patients ( %), aa in patients ( %), and patients were homozygous for gg ( %). analysis of hpse rs snp revealed a similar distribution for tc (n ¼ , %) and tt (n ¼ , %) and a frequency of patients ( %) for cc. vod of the liver was observed in / patients ( %). if vod of the liver was diagnosed all of our patients were treated with defibrotide as early as possible. altogether, / patients with vod of the liver ( %) died of vod, whereas / patients with vod of the liver ( %) survived vod. early medical intervention is most probably the reason for this high cure rate. patients with hpse genotypes gg or ag of rs (g a) had a significantly reduced incidence of vod of the liver on day after hsct compared to patients with genotype aa ( % vs. %, p ¼ . ). in addition, incidence of vod in patients with genotype cc or ct of rs (c t) was significantly decreased in comparison to patients with genotype tt ( % vs. %, p ¼ . ). interestingly, no patient with genotype cc developed vod. because both snps co-occur in vivo, we generated subsets: aa-tt, gg-cc and a group with remaining snp combinations. we found significant differences between all three patient groups (p ¼ . ). patients with aa-tt showed the highest incidence of vod of the liver ( %), while vod was not observed in patients with gg-cc ( %) and residual combinations were numerically in-between ( % allogeneic, n ¼ ) were considered at risk of hbv reactivation for the following criteria of positivity: ) donor and/or recipient anti-hbc and/or ) anti-hbe and/or ) anti hbs þ /-. lamivudine prophylaxis was given to out of patients ( %). overall, patients, autologous and allogeneic recipients, developed hbv reactivation at a median time of months (range, - ) after hsct. of these patients, were at risk of reactivation and of them were not receiving prophylaxis. one was hbv-negative at the time of pretransplant screening. the cumulative incidence of hbv reactivation was , % for patients at risk and %.for the entire hsct population studied. one patient reactivated hbv infection during lamivudine prophylaxis. hbv isolate genotype was studied in all reactivated patients. two isolates showed hbsag escape mutations ( n, y, i, k, r, e, r k, t s) and in lamivudine drug resistance mutations ( s and l l), of them resulted hbv genotype d and one resulted genotype f. conclusion: the low rate of reactivation in our cohort is probably due to the accuracy of pre-transplant donor/ recipients screening and to the extensive prophylaxis program. however, a case of hbv reactivation occurred in a patient negative at serological screening (sero-negative hbv occult infection). the detection of immune-escape hbsag mutations, associated with lack of recognition by neutralizing immune activity and by diagnostic assay in most hbvreactivated patients, supports the role of these mutations in the process of immune-suppression driven hbv reactivation. disclosure of interest: none declared. introduction: polyomavirus (pv) hemorrhagic cystitis (hc) is a severe complication after haploidentical hematopoietic stem cell transplantation (haplo hsct). in the setting of solid organ transplantation, the use of tacrolimus (fk) has been associated with higher risk of pv-hc compared with cyclosporine a (csa). the aim of our study was to investigate risk factors of pv-hc in haplo-hsct recipients, with particular focus on immunosuppressive agent used as graft-versus-host disease (gvhd) prophylaxis. materials (or patients) and methods: we retrospectively analyzed the incidence, risk factors and outcome of pv-hc in consecutive adult patients (pts) undergoing haplo hsct due to hematological malignancies between and at our two institutions. all hscts were t-cell replete and included post-transplant high-dose cyclophosphamide as part of gvhd prophylaxis. pv-hc was defined as pv detection in urine by pcr testing in association with clinical symptoms of hc without other concurrent genitourinary conditions. variables tested as potential risk factors to develop a pv-hc were: conditioning regimen, age, diagnosis, stem cell source, levofloxacin prophylaxis, acute gvhd, interval between diagnosis and hsct, pre-hsct therapy lines, previous subdiaphragmatic radiotherapy, n. of cd þ cells infused, neutrophil and platelet engraftment, fk vs csa-based immunosuppressive regimen. results: main pts' characteristics are shown in table . after a median follow-up time of months from transplantation, ten ( %) pts developed a pv-hc. pv-hc occurred in / ( %) pts receiving fk and / ( %) pts receiving csa (p ¼ . ). etiology was bk virus in ( %) cases and jc virus in one ( %) case. median time of pv-hc diagnosis was ( - ) days post-hsct. in the multivariate analysis, myeloablative (mya) or reduced-intensity conditioning (ric) regimens (hr ¼ . , % ci: . - . ; p ¼ . ) and fk-based immunosuppression (hr ¼ . , % ci: . - . ; p ¼ . ) were the only two independent factors associated with pv-hc. regarding treatment, two pts received cidofovir therapy, one pt benefited from immunosuppression tapering, whereas only supportive therapy was required for the remaining pts. notably, all pts achieved complete remission of symptoms. . ( . - . ) relapse who would benefit from further treatment intensification early after transplant. introduction: the prognosis for patients with hematologic malignancies (hm) who relapse after allohct is dismal, and immune checkpoint modulation with ctla blockade is a novel strategy to enhance the graft versus tumor (gvt) effect. materials (or patients) and methods: this is a phase i/ib study of the ctla blocking antibody ipilimumab to treat patients with any hm histology who relapse after allohct. the primary objectives are to determine the mtd and evaluate safety. secondary objectives include a preliminary evaluation of efficacy and changes in immune cell phenotype. ipilimumab was given at mg/kg or mg/kg iv q weeks for cycles of induction, followed by maintenance dosing q weeks up to year. disease-specific response criteria were assessed at the mid-point ( weeks), end of induction ( weeks), and throughout maintenance. immunophenotyping was performed by -color flow cytometry and analyzed by facsdiva. results: twenty-three patients are enrolled to date. in the phase i portion, patients were treated at mg/kg and patients were treated at mg/kg. an mtd was not reached, and patients subsequently enrolled in the phase ib expansion cohort at mg/kg. the median number of prior therapies excluding transplant was (range - ), and patients had received prior therapy for their post transplant relapse. histologies included chl (n ¼ ), aml (n ¼ ), nhl (n ¼ ), and mds (n ¼ ), and patient each had mm, mpn, and all. the median age at enrollment was (range - ). immune-related adverse events (iraes) were observed in patients, and included itp (n ¼ ), diarrhea (n ¼ ), pneumonitis (n ¼ ), and colitis (n ¼ ), all of which were generally reversible with steroids with most patients resuming ipilimumab dosing. three dlts have been observed, including cases of chronic gvhd (both liver, mild) and one patient death due to presumed sepsis in the context of severe iraes, including grade colitis and grade pneumonitis. nine patients remain on treatment, and patients discontinued due to progressive disease, patients due to cgvhd, and patient with trm. in an interim efficacy analysis, / ( . %) patients evaluable for response had anti-tumor activity with clinical benefit. four patients achieved formal responses by disease-specific criteria, including a chl patient with pr with dramatic reduction in nodal and extranodal disease with a complete marrow response at weeks (baseline % involvement), a mm patient with a pr with near resolution of a lung plasmacytoma, and two patients with aml who achieved cr by weeks. the median follow-up time among survivors is . months, and month overall survival is currently %. immunophenotyping studies revealed that the ratio of regulatory t cells to conventional t cells decreased between % and % after ipilimumab treatment. conclusion: multiple doses of ipilimumab given to patients with relapsed hm after allohct were generally well-tolerated, with cgvhd and iraes observed in a small number of patients. anti-tumor activity was observed, both in lymphoid malignanices and for the first time in myeloid malignancies, including patients with aml who achieved cr. the ratio of regulatory to conventional t cells decreased with ctla blockade. the phase ib expansion cohort at mg/kg continues to accrue and will provide additional efficacy, safety, and correlative data. disclosure of interest: none declared. adverse risk cytogenetic (p ¼ . ). in patients who relapsed between and months post-transplant more than one course of chemotherapy to achieve cr (p ¼ . ), adverse risk cytogenetics (p ¼ . ) and flt itd positivity (p ¼ . ) all predicted for relapse. finally only cmv positivity predicted for relapse risk for relapse more than months post-transplant (p ¼ . ). conclusion: this study demonstrates for the first time that a complex interaction of disease specific and transplant factors determine the kinetics of relapse post-transplant. in addition to identifying that heterogenous factors related to transplant characteristics and disease biology determine the timing of disease relapse these data confirm the importance of early intervention post-transplant in patients allografted for aml and and will assist in the design of novel therapeutic strategies. disclosure of interest: none declared. introduction: bone marrow (bm) is the recommended stem cell source in hematopoietic stem cell transplantation (hsct) for bone marrow failure (bmf). recent large studies in aplastic anemia showed that the use of peripheral blood stem cells (pbsc) increased the risk for chronic graft-versus-host disease without reducing graft failures and lead to an inferior survival after hsct. however, a substantial proportion of hsct is still performed with pbsc. ease of collection and a more rapid engraftment might favor pbsc in the short term and more background information to clarify the situation is warranted. materials (or patients) and methods: the current global practice of hsct for bmf was studied for potential macroeconomic factors associated with the selection of stem cell source. introduction: the outcome of alternative donor hematopoietic stem cell transplantation (hsct) for the patients with severe aplastic anemia (saa) who lack an hla-matched familial donor has improved recently. this study was aimed to compare the treatment outcome of immunosuppressive therapy (ist) with alternative donor hsct in children with saa. materials (or patients) and methods: medical records of saa patients who received ist and/or alternative donor hsct from umbilical cord blood (ucb), haploidentical peripheral blood stem cells (pbsc), unrelated bone marrow (ubm), or unrelated pbsc (upbsc) between june and december were reviewed, and data were analyzed retrospectively. the kaplan-meier method was used to calculate the estimated survival rates which were compared with log-rank test. results: of patients with saa, patients received ist and/ or alternative donor hsct. nineteen patients received ist as an initial treatment modality, of whom patients failed to respond at months from the initiation of treatment. thirtyfour patients received alternative donor hsct either following ist (n ¼ ) or as frontline therapy (n ¼ ; ubmt, upbsct, ucbt, haploidentical pbsct). the failure-free survival rate (ffs) of ist group was . %, while that of frontline hsct group was . % (po . ). patients who received hsct following ist showed an inferior event-free survival rate as compared with those who received frontline hsct ( . % vs . %; p ¼ . ). conclusion: the outcome of alternative donor hsct in our cohort was higher than usually expected, especially in those who received hsct without prior ist. these results suggest that frontline alternative donor sct might be a better treatment option than ist for children with saa who lack an hla-matched familial donor. disclosure of interest: none declared. increased risk of development of malignancies in adult patients treated with antithymocyte globulin as first line treatment for aplastic anaemia during a follow-up period of thirty years. j. v. d introduction: adult aplastic anemia (aa) is considered to be an immune-mediated disease with bone marrow aplasia and pancytopenia. patients can be treated with either hematopoietic stem cell transplantation (hsct) or immunosuppressive therapy (ist) with antithymocyte globulin (atg) and achieve long-term survival. treatment related long term toxicity and outcomes should guide the decisions about first line therapy. a long-term toxicity of both treatments is the development of malignancies. follow-up studies until years after ist in aa patients showed cancer development in up to % of patients. analysis of cohorts with a longer duration of follow up are lacking and the incidence of malignancies in aa patients was never compared with a control population and the contribution of second line hsct to this risk is not clear. we assessed the long-term cumulative incidence of malignancies in a cohort of adult patients with aa who were treated with atg as first line treatment at leiden university hospital between and with hsct and death as competing risks and compared this incidence to cancer development in the general dutch population. materials (or patients) and methods: patients treated with first line atg between and were entered in this single-centre retrospective cohort study. primary endpoint was the cumulative incidence of malignancies with death and second line treatment with hsct as competing risks. secondly, the cumulative incidence in our cohort was compared to that of the sex-and age matched general dutch population derived from the dutch cancer registration, using standardized incidence ratio (sir). furthermore, several patient-dependent (age), disease-dependent (severity of disease) and treatmentdependent (addition of ciclosporin to atg) variables were assessed as possible prognostic factors for cancer development, using cumulative incidence curves, gray's test and the cox proportional hazard model for the cause-specific hazard. spss statistics . and r were used for all data analyses. ethical approval for data collection and data analysis was obtained. results: median length of follow-up of surviving patients without hsct was . years (range . - . years) , with a cumulative malignancy rate of % after years. the incidence of malignancies was increased compared with the general dutch population (sir: . ( % ci: . - . )). the cumulative incidence of developing mds/aml was % after years (occurring at - years after start of treatment). in the multivariate analysis, age at time of ist was significantly associated with a higher malignancy rate (hr: . per years; p ¼ . ). hr for addition of ciclosporin was . (p ¼ . ) and hr for nsaa vs (v)saa hr was . (p ¼ . )). the risk of the development of malignancies in aa patients treated with atg is increased compared to the risk in the age and sex matched dutch population for both solid and hematologic malignancies. we are not able to discern whether the increased risk is due to the treatment with ist or due to the biology of the disease. long follow up studies in age matched aa patients receiving hsct are needed to compare this long term risk between both treatments. these results underscore the recommendation in international guidelines to follow all patients with aa after initial ist lifelong. disclosure of interest: none declared. hematopoietic stem cell transplantation, molecular biology, dmitriy rogachev center for pediatric hematology, oncology and immunology, moscow, russian federation introduction: results of matched unrelated and, to a lesser extent, haploidentical transplantation in severe aplastic anemia have improved over the last decade. the major factors behind this improvement are high-resolution hla-typing, intensified immune suppression with fludarabine and alemtuzumab and modern supportive care. we implemented a new graft manipulation method, tcr-alpha/beta depletion, to further improve gvhd control and overall results of transplantation in a group of saa patients, refractory to two courses of atg/csa immune suppression. materials (or patients) and methods: twenty six patients with saa, median age , ( , - ) years, male/ female, received mud ( ) or haploidentical ( ) transplantation with tcr alpha/beta and cd depletion between . . and . . . patients received a median of atg/csa courses. core preparative regimen included cyclophosphamide mg/ kg x , fludarabine mg/kg x , atg (horse) mg/kg x and total lymphoid irradiation gy total dose. one patient received alemtuzumab instead of atg due to anaphylaxis. in all cases g-csf-mobilized pbsc were used as graft source. tcr-alpha/beta and cd depletion were performed with clinimacs plus according to manufacturer's recommendations. final graft contained x of cd þ /kg and x of tcra/b þ /kg. post-transplant immune suppression included short methotrexate and tacrolimus till day þ - . results: primary engraftment was registered in all patients, median days for neutrophils and for platelets. secondary graft failure/rejection developed in patients, both transplanted from mud. both patients were salvaged with a second mud transplantation. acute gvhd was registered in patients, grade skin in and grade skin þ gut in patients. cumulative incidence of agvhd grade - was %( %ci: - ). chronic gvhd was registered in patient, cumulative incidence %( % ci: , - ). two patients ( from each cohort) died of cmv pneumonia at and months respectively. in both cases cmv sero status was d-/r þ . at year follow-up cumulative incidence or trm is %( %ci: , - ), km estimte of overall survival is %( %ci: - ), %( %ci: - ) in the mud group, %( %ci: - ) in the haplo group. conclusion: tcr alpha/beta depletion is a highly effective method of gvhd prophylaxis in saa patients. use of this platform makes alternative donor transplantation in saa a safe therapeutic option. further improvement will depend on new strategies to control viral infections. disclosure of interest: none declared. tp: ( ) vs ( ) cm/s, respectively; (po . ), and were decreased more significantly following hsct than on tp: mean(sd) d: - ( ) vs þ ( ), respectively. the percentage of patients with normal velocities was significantly higher post-hsct ( / ) than in the tp arm ( / ) (p ¼ . ) conclusion: this prospective national trial comparing tp vs. hsct in sca-patients with a history of abnormal velocities shows for the first time that hsct significantly results in a greater decrease in velocities than tp, and has very little toxicity. these results suggest hsct is the treatment of choice for sca-children with a history of abnormal-tcd and genoidentical donor. disclosure of interest: none declared. introduction: busulfan (bu) is the backbone of the conditioning regimen for patients with sickle cell anemia (sca) undergoing bmt. patients with sca might predispose to transplant-related neurological and pulmonary toxicities due to pre-existing disease-related cerebrovascular and lung injury. bu therapy appears to be an important contributing factor in this context. to date, no studies have evaluated the pharmacokinetic (pk) parameters of intravenous bu (ivbu) for subsequent doses in patients with sca. materials (or patients) and methods: we studied ivbu pk parameters and clinical outcomes of children with sca undergoing bmt from hla matched siblings. the median age of patients was . (range, . - . ) years. conditioning regimen consisted of bu/cy /atg (n ¼ ) or flu/bu/cy (n ¼ ). six patients ( %) had stroke, ( %) exhibited gliosis due to previous brain injury, ( %) had repeated acute chest syndrome, and ( %) patients were on chronic transfusions. ivbu was administered every h for days with pk-guided dose adjustment to target a conservative area under the concentration versus time curve (auc) range of - mmol*min. the role of glutathione s-transferase (gst) polymorphisms has also been investigated. results: all patients had sustained engraftment. a repeated measures anova showed that the first-dose bu clearance ( . ml/min/kg) was significantly higher than after dose ( . ml/min/kg; po . ), dose ( . ml/min/kg; po . ), and dose ( . ml/min/kg; po . ). such differences in clearance have never been described in patients with sca. after the first-dose, % of patients achieved the target range.none of the patients developed hepatic vod. no patient developed grade z toxicity. there was no association between gst polymorphism and the pk of bu. we adapted a new dose-adjustment strategy targeting exposures to the lower end ( mmol*min) of the auc range after the first dose of bu to avoid unnecessary dose increases on subsequent days due to differences in clearance. this strategy enabled most patients ( %) to maintain the auc within therapeutic range following dose adjustments. a -year probability of os and sickle-cell free survival were % ( % ci, - %). there was no correlation between any bu pk parameters and survival, toxicity, acute gvhd, chronic gvhd, and transplant-related mortality. conclusion: this study found that the pk behavior of ivbu in children with sca is characterized by significantly higher bu clearance after the first-dose compared with subsequent daily doses. this finding allowed us to adapt a new dose adjustment strategy after the first dose of bu, which effectively prevented subsequent dose readjustments. conservative auc range and targeting exposures to the lower end of the range after the first dose was associated with negligible toxicity, and high engraftment and sickle cell-free survival rates. disclosure of interest: none declared. however, the probability to find a suitable door is only - %. since most of these patients (pts) could not find the potential donor, we would like to investigate the haploidentical donor hsct (hapo-sct) in thalassemia. materials (or patients) and methods: between jan and december , a total of severe thalassemia patients (pts) underwent hsct in our center. sixty five pts underwent mrd-hsct, pts underwent mud-hsct and pts underwent haplo-hsct. for haplo-hsct, subjects were male and were female. the median age was yrs (range; - ). thirteen of received stem cells from mother and from father. ten of were high risk class . these high risk class pts received hydroxyurea mg/kg/d for at least months prior to hsct. all pts received courses of pre-transplant immunosuppression (ptis) consisting of fludarabine (flu) mg/m /d together with dexamethasone (dex) mg/m /d for days. after courses of ptis, all pts received a reducedtoxicity conditioning (rtc) regimen consisting of thymoglobulin . mg/kg/d (d- to d- ) , flu mg/m /d i.v. (- to - ) each dose immediately followed by busulfan (bu) mg/ m once daily i.v. (d- to d - ) gvhd prophylaxis consisted of cyclophosphamide (cy) mg/kg/d (d þ to d þ ). tacrolimus or sirolimus was given for months to yr started together with mycophenolate mofetil on d þ , the latter was quickly tapered after months. t-cell repleted peripheral blood stem cells (pbsc) were given to all pts, targeting a cd þ dose of - x cells/kg. results: sixteen of were engrafted with full donor chimerism ( %) while pts suffered graft failure. however, these pts received second transplant on day þ with minimal conditioning regimen and additional pbsc after which they achieved full donor chimerism. the median time to neutrophil engraftment was days (range; - ) . six pts had acute gvhd gr i, grade ii and gr iv. only one had extensive chronic gvhd. at this time, of pts survive thalassemia-free and have sustained full donor chimerism ( %). event free survival (efs) and overall survival (os) rates are %. the median follow up time is months (range - ). the efs rates among mrd, mud and haplo-hsct in our center are %, % and % respectively (p ¼ . ). conclusion: haploidentical hsct for high risk thalassemia pts with our novel approach is safe and should be considered as modality to secure thalassemia-free survival with a low risk for graft rejection and treatment-related mortality. in view of our results, we suggest that all thalassemia pts even those with high risk class features should be offered the chance for cure with hsct. disclosure of interest: none declared. introduction: allogeneic hematopoietic stem cell transplantation (hsct) is a curative option for many patients with hematological malignancies. graft versus host disease (gvhd) mediated by donor alloreactive t cells remains a major obstacle and limits its wider application. it is now well established that t regulatory cells (tregs) are critical for the maintenance of self-tolerance, and a deficiency or dysfunction in these cells is thought to be a key factor in the development and progression of gvhd. numerous murine studies have shown the benefit of adoptive transfer of tregs in the setting of hsct. notably, the success of this approach is improved if the transferred tregs are specific for antigens expressed by the host. a robust method to generate homogeneous populations of alloantigen specific tregs in humans would be a major step towards realizing the goal of using these cells to suppress gvhd. traditional approaches to generate antigen specific t cells include repetitive cycles of re-stimulation with antigen in vitro, sorting of tetramer positive cells, or over-expression of a t cell receptor for a specific antigen. these methods produce limited cell numbers and require modification for each individual patient depending on their own tissue type and that of the transplanted patient. we hypothesized that a more efficient approach to generating antigen specific tregs would be to genetically engineer them to express alloantigen-specific chimeric antigen receptors (cars). materials (or patients) and methods: we generated an hla-a specific car by cloning the immunoglobulin heavy and light chain variable regions from an anti-hla-a antibodysecreting hybridoma. these regions were fused into a single chain antibody, which was then linked to intracellular signalling domains from human cd and cd zeta. surface expression and antigen-specificity of the a -car was confirmed by flow cytometry to detect expression of a myc epitope tag on the extracellular portion of the car and binding to an hla-a tetramer, respectively. cd þ cd hi cd ra þ naïve tregs (ntregs) and cd þ cd lo cd ra þ naïve tconv (ntconv) cells were sorted from the blood of hla-a donors, stimulated with artificia apcs and transduced with lentivirus encoding the a -car or a control car specific for her . results: transduced tregs maintained their expected phenotype, including high levels of foxp , ctla- , cd and helios, and low levels of cd and il- compared to tconv cells. to test antigen specificity, the transduced t cell lines were stimulated with k cells expressing hla-a or her : only k cells expressing hla-a stimulated proliferation of a -car expressing tregs and tconv cells. car-stimulated tregs also suppressed the in vitro proliferation of car-stimulated tconv cells. to demonstrate the in-vivo suppressive capacity of a -car tregs, nsg mice were reconstituted with hla-a þ pbmcs in the absence or presence of different ratios of control-transduced or a -car transduced tregs. preliminary results suggest that a -car tregs have a superior ability to delay the onset of gvhd compared to polyclonal tregs. conclusion: human tregs can be efficiently transduced to express functional, alloantigen-specific cars that confer antigen-specific specific suppression of effector t cells both in-vitro and in-vivo. these data supports the feasibility of this strategy to re-direct tregs for transplant-related therapies. disclosure of interest: none declared. car spacers including ngfr domains allow efficient t-cell tracking and mediate superior antitumor effects m. casucci ,* , l. falcone , b. camisa , f. ciceri , c. bonini , a. bondanza innovative immunotherapies unit, experimental hematology unit, clinical hematology and bone marrow transplantation unit, san raffaele hospital scientific institute, milano, italy introduction: chimeric antigen receptors (cars) frequently include an igg -ch ch spacer conferring optimal flexibility for antigen engagement and allowing the selection and tracking of car-expressing t cells. a serious drawback of ch ch -spaced cars is however their interaction with fcg receptors (fcgrs). indeed, this antigen-independent binding may lead to the unintended elimination of cells expressing these receptors (mainly phagocytes), foster the development of non-specific immune reactions and drastically decrease the efficacy of the strategy due to the premature clearance of transduced t cells in vivo. materials (or patients) and methods: we designed and constructed novel car backbones by substituting the igg -ch ch spacer with regions from the extracellular portion of the low-affinity nerve-growth-factor receptor (lngfr), differing for the length and potential binding to ngf. in particular, we used our recently developed cd v -specific car as a model for comparing the antitumor activity of the different lngfr-based designs both in vitro and in vivo. results: after transduction, all constructs could be identified on the t-cell surface using anti-lngfr antibodies, indicating that they were correctly processed, mounted on the cell membrane and still recognized by anti-ngfr antibodies. as a consequence, all the lngfr-based spacers allowed selecting car-t cells with immune-magnetic beads coupled to anti-ngfr antibodies, without interfering with their expansion and functional differentiation after activation with cd /cd beads plus il- and il- . most importantly, lngfr-spaced car-t cells maintained potent cytotoxic, proliferative and cytokine-release activity in response to cd v -expressing leukemia and myeloma cells, while lacking antigen-independent recognition through the fcgrs. noticeably, even at supraphysiological ngf concentrations, the lngfr-spaced cd v -car. z car t cells were not induced to proliferate, indicating the absence of signaling via soluble ngf. strikingly, lngfrspaced car-t cells better expanded and persisted in vivo compared to ch ch -spaced car-t cells and mediated superior antitumor effects in a well-established tumor disease model. interestingly, we demonstrated that the premature disappearance of ch ch -spaced car-t cells was due to engulfment by mouse phagocytes, a phenomenon not occurring with lngfr-spaced t cells. in conclusion, we have demonstrated that the incorporation of the lngfr marker gene directly in the car sequence allows for a single molecule to work as a therapeutic and as a selection/tracking gene and shows an increased efficacy/safety profile compared to the igg -ch ch spacer. disclosure of interest: none declared. university children's hospital, würzburg, clinic of pediatric oncology, heinrich-heine-university düsseldorf, university children's hospital, essen, university children's hospital, münster, department of pediatrics, jena university hospital, germany introduction: viral infections represent an important cause of morbidity and mortality in immunosuppressed patients post hematopoietic stem cell transplantation (hsct). as viral infections often remain refractory to pharmacologic treatment, alternative treatment strategies such as immunotherapy are required. adenovirus (adv) is the predominant diseasecausing pathogen in pediatric hsct. materials (or patients) and methods: in a clinical trial we analyzed safety and efficacy of ex vivo adoptive t-cell transfer (act) with hexon-specific t cells, predominantly of t helper - phenotype. thirty patients suffering from chemo-refractory adv disease or viremia after hsct were treated with advspecific t cells generated by ifn-g capture technique. results: adv-specific t cells were successfully isolated in % of cases using the adenoviral hexon antigen and were directly infused into the patients without further ex vivo expansion steps. adv-specific t-cell grafts were composed of a mixture of naïve, central memory, effector memory and effector t-cell populations with a predominance of late effector stages, indicating proliferation and effector potential of the transferred t cells. in all thirty patients, act was feasible without acute toxicities or significant onset of graft-versus-host disease. act led to antiviral immunity in vivo up to months with viral control, resulting in complete clearance of viremia in % of patients with antigen-specific t-cell responses. efficacy of adoptive t-cell transfer was independent of the initial t-cell dose. after a follow up of months post act, overall survival was markedly increased in responders (mean: days, survivors) as compared to non-responders who all died shortly after act (mean: days, no survivors). adv-related mortality was % in non-responders compared to . % in responders (z log reduction of dna copies/ml post act), indicating a strong correlation between virus-specific immunity and virus control. conclusion: in conclusion, ex vivo act of adv-specific t helper - cells was well tolerated and led to successful and sustained restoration of t-cell immunity, correlated with virological response and protection from virus-related mortality. this cellular immunotherapy is a short-term available and broadly applicable treatment. disclosure of interest: none declared. [o ] putkonen , t. kuittinen , j. pelkonen , , p. mäntymaa , k. remes , v. varmavuo , e. jantunen o o comparison of ric-allohct and autohct for years old patients with acute lymphoblastic leukemia: an analysis from acute leukemia working party of the ebmt s united kingdom, erasmus mc-daniel den hoed cancer centre double-blind, placebo-controlled phase study of brentuximab vedotin in patients at risk of progression following autologous stem cell transplant for united states, szent istvan & szent laszlo corporate hospital hematology & stem cell dept united states, department of bone marrow transplantation & oncohematology, maria sklodowska-curie institute of oncology united states, istituto nazionale dei tumori, milano, irccs azienda ospedaliera universitaria united states introduction: the aethera trial was initiated to evaluate whether brentuximab vedotin (bv) can prevent progression in conclusion: bv improved post-asct pfs across all subgroups of pts. pts with more risk factors had the most benefit from bv. aes were consistent with the known safety profile of bv references: . lion t. adenovirus infections in immunocompetent and immunocompromised patients european guidelines for diagnosis and treatment of adenovirus infection in leukemia and stem cell transplantation: summary of ecil- adoptive transfer and selective reconstitution of streptamer-selected cytomegalovirus-specific cd þ t cells leads to virus clearance in patients after allogeneic peripheral blood stem cell transplantation lowest numbers of primary cd þ t cells can reconstitute protective immunity upon adoptive immunotherapy donor ebv status has introduction: epstein-barr virus (ebv) has been a major cause of post-transplant lymphoproliferative disorder after allogeneic stem cell transplantation (hsct). the impact of the donor (d) and recipient (r) serologic status on survival, relapse-free survival, relapse incidence, non-relapse mortality and incidence of graft-versus-host disease was unknown so far. objective: we analyzed the influence of the donor's and recipient's ebv status on allo-hsct transplant outcomes. materials (or patients) and methods: , allo-hscts performed due to acute leukemia ebv-seropositive donors also had no influence on relapse-free survival ), and non-relapse mortality hsct recipients receiving grafts from ebv-seropositive donors had higher risk of acute gvhd after adjusting for confounders (donor type, conditioning, stem cell cource, patient age, gender match, t-cell depletion, year of transplant), d þ serostatus had an impact on development of acute gvhd p ¼ . , and for cgvhd all patients engrafted, with a median time to neutrophil and platelet recovery of days ( - ) and days ( - ), respectively. post-hsct recovery of lymphocyte subsets was broad and fast: median time to cd /ml was days, to cd /ml days and to cd /ml days. circulating t cells comprised naïve and memory subsets, with a recovery of cd recent thymic emigrants (rtes) from day . all patients had a significantly higher proportion of rtes at day and compared to their pre-hsct levels, suggesting an improvement in their thymic function after hsct. with a median follow-up for living patients of months ( - ), the -year cumulative ci of nrm and relapse were % and %. three of the acute leukemia relapses were hla-loss variants. notably, one was observed for the first time in all. ci of agvhd grade ii-iv and iii-iv at months were % and %, while -year ci of cgvhd was % conclusion: myeloablative haplohsct with pbsc, pt-cy and sirolimus is a valid option for patients with aggressive/ advanced disease. the acceptable rates of gvhd and nrm as well as the favorable immune reconstitution profile open the way for combining it with novel immunomodulatory/ cellular therapies to improve dfs in patients at high risk for o factors determining the kinetics of disease relapse after allogeneic stem cell transplantation (allo-sct) for acute myeloid leukaemia (aml): a survey from the acute leukaemia working party of the ebmt c o a novel quantitative pcr approach targeting insertion/ deletion polymorphisms (indel-pcr) for chimerism quantification: finally high sensitivity and quantification capacity together a post-hematopoietic stem cell transplantation (sct) chimerism monitoring is important to assess engraftment, anticipate relapse and provide information on the development of graft versus host disease, facilitating therapeutic intervention. the aim of this study was to test the technical efficacy and clinical utility of a novel quantitative pcr approach targeting insertion/deletion polymorphisms of note, analysis of artificial mixtures provides evidence of significantly (z logs) higher sensitivity by indel-pcr ( . %) than by str-pcr ( %, table ). moreover, indel-pcr shows unprecedented quantification capacity (table ). out of the samples analyzed, were positive and negative by both methods, while were positive only by indel-pcr ( % with o % recipient). hematological relapse occurred in patients, molecular relapse/persistence in patients. all of them presented a positive indel-pcr (with increasing %r in / ) and a negative str-pcr result in the sample before relapse (table ). the patients in complete remission, although presented positive indel-pcr, showed stable or decreasing %r chimerism dynamics in / (data not shown). conclusion: this novel indel-pcr is a simple and accurate technique that, in comparison with the current gold standard str-pcr, shows very good concordance and provides higher rates of informative loci per patient, as well as unprecedented combined sensitivity and quantification capacity introduction: the immune recovery after cd þ cell selection is slow and patients tend to remain susceptible to opportunistic infections for several months after hsct. to hasten and improve post-transplant immune reconstitution broad repertoire various strategies under this approach, a rapid immunological reconstitution and very promising outcome have been reported in pediatric patients. with the aims of confirming these results even in adults, we have recently launched this programme and here we report our preliminar clinical data in leukemia patients we have so far treated over the past months. materials (or patients) and methods: twenty-two patients, median age years (range - ), with aml (n ¼ ), all (n ¼ ), mds (n ¼ ) entered the study. eight patients were in cr , in cr , and in advanced-stage disease at transplant. conditioning consisted of atg , mg/kg from day - to day - , treosulfan gr/sqm from - to - , fludarabine mg/sqm from - to - and thiotepa mg/kg on days - and - . no patient received any post transplantation pharmacologic prophylaxis for gvhd. ten mg/kg g-csf was used to mobilize pbpcs from one-haplotype mismatched donors ( mothers median cd þ cell/ml counts at , and days since the transplant were , and , respectively. cmv antigenemia reactivation occurred in cases (in , cmv serology was unfavourable). no patients has so far developed cmv disease. invasive fungal disease was prevented in all cases using l-amb-based prophylaxis over the neutropenic phase. overall, patients have so far died ( relapse, non-hematologic causes). survive ( diseasefree, in early relapse) at a median follow-up of months (range - ) (fig. ). conclusion: the infusion of ab/cd -depleted grafts was safe and effective also in adult setting, resulting into rapid engraftment and fast immunological reconstitution haploidentical stem cell transplantation in very poor risk cytogenetics acute myeloid leukemia: results in consecutive patients cytogenetic prognostic risk was defined according to the revised medical research counsil (mrc) classification, from diagnostic bone marrow samples with standard methods and in accordance with international system of human cytogenetics guidelines. os and disease free survival (dfs) were calculated using the kaplan-meier methods. results: median age of the patients at time of transplant was years (range to ).cytogenetics:chromosome abnormalities pts, monosomal karyotype pts and complex karyotyping pts non relapse mortality (nrm) was % at year after transplant.estimated lfs from day þ after transplant was . % and % at years. conclusion: haploidentical stem cell transplant (haplo-sct) is a valid treatment option for the patients with very poor risk aml. references: haploidentical, unmaipulated, primed bone marrow for pts with high risk hematologic malignancies haploidentical transplantation using t cell replete pbsc and myeloablative conditioning in pts with high risk hematologic malignancies who lack conventional donors kodera on behalf of the worldwide network of blood and marrow transplantation o graft depletion of tcralpha/beta and cd in matched unrelated and haploidentical transplantation for severe aplastic anemia: high survival with low incidence of graftversus-host disease and transplant-related mortality m at enrollment all patients were on tp and paired analysis showed that mean (sd) maximum velocities had significantly decreased (po . ) under tp: ( ) cm/s vs ( ) cm/s at tp initiation. following hla-typing, without genoidentical donor were included in the transfusion arm and with genoidentical donor were transplanted in hsct-centers. during the months follow-up, no stroke was observed but one patient in the tp arm experienced a hyperammonemic reversible coma, without mri/mra alteration. in the hsct arm, all patients successfully engrafted, one grade ii and two grade iii acute gvhd, and no chronic gvhd were observed. complications were seizures (n ¼ ), cmv (n ¼ ) or ebv replications (n ¼ ), hemorrhagic cystitis (n ¼ ), aspergillosis (n ¼ ), prolonged but reversible thrombopenia (n ¼ ), transitory hemolytic anemia (n ¼ ) granda ospedale maggiore policlinico, hematology and bone marrow transplantation unit metabolic syndrome (ms) is defined as a clustering of five factors including ( ) hyperglycaemia ( ) hypertriglyceridaemia; ( ) low hdl cholesterol; ( ) hypertension; ( ) obesity (high waist circumference or body mass index (bmi)). it is associated with raised risk of cardiovascular disease (cvd) and is increasingly recognised in patients after hct and revised guidelines for long-term hct survivors recommend screening for ms. previous studies have been small and the definition of ms variable, although harmonised criteria are now agreed materials (or patients) and methods: this was an ebmt approved cross-sectional, non-interventional study of consecutive hct patients aged þ years and a minimum of year post-transplant attending routine follow-up hct and/or late effects clinics in centres. centres completed med c forms incorporating routine recording of the ms parameters (given above) as well as performance status (ecog); evidence of cardiovascular events using the harmonised definition of ms (at least / factors), the prevalence of ms was . %. there was no difference in time since hct but there was a significant difference in prevalence by age at diagnosis, hct, follow-up (all po . with increasing age). as expected, statistical differences (po . ) between patients with and without ms were observed for bmi routine screening and early intervention may reduce the risk of cardiovascular events in hct survivors, and should ideally be tested in a randomised controlled trial setting. meanwhile, screening and management of reversible features of the metabolic syndrome should be robustly integrated within routine hct long-term follow-up care. references: disclosure of interest: none declared. introduction: the introduction of less toxic conditioning regimens for hematopoietic cell transplantation (hct) has led to an increase in eligible patients, although their benefit on patient's perceived wellbeing remains unclear. we aimed to prospectively study patients' quality of life (qol) and emotional wellbeing (ew) in consecutive hct recipients depression and anxiety (hads), and sleep quality (psqi) at pre-hct, at hospital discharge (hd) and at months post-hct. results: out of transplanted patients, ( %) consented to participate. those who refused (n ¼ , %) more frequently had active disease at hct ( % vs. % for pr/cr, p ¼ . ) and/or had a prior hct among included patients ( % men; median age received an auto-hct (n ¼ at hd; n ¼ at þ m) received an allo ric-hct (n ¼ at hd; n ¼ at þ m) at baseline, clinically significant depressive symptoms were reported by % of the patients, with a slight increase ( %) at hd and at þ m (p ¼ . ). again, there was a strong interaction between depressive symptoms and hct groups in the early post-hct phase (p ¼ . ); depression decreased in auto-hct after hd and, on the contrary, it increased in the same time-point in mac-hct (p ¼ . ). borderline differences were seen between auto-and ric-hct (p ¼ . ) but not between ric-and mac-hct (p ¼ . ). clinically significant anxiety was observed at baseline in % of the patients and significantly decreased at the time of hd ( %) and afterwards ( %) conclusion: mac-hct recipients reported the greatest impairment in the parameters studied; other variables such as gender, age and baseline ew/qol should be considered for specific psychological/clinical follow-up and eventual intervention unit of molecular and functional immunogenetics, unit of hematology and bone marrow transplantation conflict with: scientific consultant of molmed s.p.a. o erbb -chimeric antigen receptor (car) modified cytokineinduced killer (cik) cell intervention for refractory solid tumors after allogeneic stem cell transplantation m already at day of culture, up to % of transduced cells showed surface expression of the erbb -car (n ¼ ). there were no significant phenotypic differences (cd þ /cd þ /cd þ /cd þ /tcr-a/b and tcr-g/d) between unmodified, empty control vector and erbb -car transduced cik cells. erbb -car cik cells efficiently lysed erbb -overexpressing breast carcinoma cells (mda-mb ) in a hour short-term cytotoxicity (europium release) assay in vitro. compared with unmodified and empty-vector transduced cik cells e:t; . % vs. . % specific lysis, n ¼ ; po . ). long-term cytotoxicity analysis ( h, brightfield imaging cytometry) demonstrated comparable results even at low effector to target ratios of : ( . % vs. . % specific lysis, n ¼ ). comparable results for shortand long time cytotoxicity could be obtained for all other tested rhabdomyosaroma cell lines in vitro. conclusion: erbb -car engineered cik cells are highly specific and efficient against erbb -antigen expressing tumor cell lines in vitro. our experiments may help to develop an approach for improved treatment of patients with high-risk adoptive t-cell immunotherapy with hexon-specific thelper- cells as a treatment for refractory adenovirus infection after allo-sct -safety and efficacy results from a anna children's hospital university children's hospital, frankfurt, dr. von haunersches kinderspital introduction: allogeneic hematopoietic cell transplantation (hct) offers the chance of cure for patients with nontransformed follicular lymphoma (fl) but is associated with the risk of non-relapse mortality (nrm). the aim of this study was to identify subgroups of fl patients who benefit from hct. materials (or patients) and methods: the minimum essential a data of consecutive patients who received hct for fl between - were extracted from the database of the german registry ''drst''. diagnosis of fl was verified by contact with reference pathologists. results: the median patient age at time of transplantation was years (range - ). prior to allogeneic hct / patients ( %) had undergone autologous hct. at time of hct, patients ( %) had sensitive disease while patients ( %) had chemorefractory disease (rd). engraftment was achieved in % of evaluable patients. day nrm was %. the median follow-up of surviving patients was . years (range . - . ) . estimated , , , -year overall survival (os) was %, %, %, and %, respectively. the corresponding estimates for efs were %, %, %, and %, respectively. % of the patients with rd at time of transplantation survived long-term. of the n ¼ patients with documented cr after hct only n ¼ ( %) relapsed. only two late relapses (beyond year ) were diagnosed among the patients with a follow-up years. patients with chronic gvhd (irrespective of stage) had a lower risk of relapse, if transplanted in cr, and a higher chance to achieve cr, if transplanted in pr or with pd (no chronic gvhd: / , chronic gvhd: / , p ¼ . ). therefore a reduced intensity conditioning approach might be considered in future prospective trials. hsct is most successful prior to leukemic transformation. given implications for treatment decisions and donor selection gata mutation screening should be performed on all patients with molecularly undefined mds and bmf disorders and potential related donors. disclosure of interest: none declared. pre-transplant weight loss predicts non-relapse mortality and relapse rates in patients with myelodysplastic syndrome after allogeneic stem cell transplantation a. radujkovic , * introduction: we have recently provided evidence that weight loss and minor metabolic changes prior to allosct were able to predict relapse and death of acute myeloid leukemia patients using data from two independent patient cohorts. this retrospective study investigated the influence of pre-transplant weight loss on the outcome of mds patients after allosct in three independent cohorts. materials (or patients) and methods: a total of patients ( % male) with a median age of years were included into the analysis. patients have been diagnosed with mds according to who criteria and received an allosct between and in three different german referral centers (heidelberg, dresden and berlin). weight data were raised from medical records by three independent researchers in three independent institutions. weight loss (expressed in percent) was calculated on the basis of recorded weight data at the time of allosct and the maximum weight in the time period of - months prior to allosct. the mds who subtype was ra(rs)/rcmd in patients ( %), raeb in patients ( %) and raeb in patients ( %). according to ipss %, % and % of the patients were in the risk groups intermediate- , intermediate- and high, respectively. the majority of the patients (n ¼ , %) was previously untreated. nineteen patients ( %) and patients ( %) received hypomethylating agents and induction type chemotherapy prior to allosct, respectively. thirty-one patients ( %) received transplants from related donors, patients ( %) from matched unrelated donors and ( %) from mismatched unrelated donors. ninety-three patients ( %) received reduced intensity conditioning and patients ( %) received standard myeloablative conditioning. survival times were measured from date of allosct. overall survival (os), relapse-free survival (rfs), relapse incidence and non-relapse mortality (nrm) were calculated from date of allosct to the appropriate endpoint. cox regression analysis was applied for os, rfs, relapse and nrm. relapse and nrm were considered as competing risks. results: estimated median follow-up at the time of analysis of surviving patients was months. a total of ( %) patients experienced weight loss % with ( %) patients losing more than % weight in the period of - months prior to allosct. patient, disease and transplant characteristics did not differ between patients with weight loss ( %, n ¼ ) and those without (n ¼ ). in multivariate analysis, weight loss and donor type were independently associated with shorter os and rfs (po . and po . , respectively). nrm was predicted by donor type (p ¼ . ), ipss (p ¼ . ) and pre-transplant weight loss (p ¼ . ) in multivariate analysis. furthermore, weight loss was also an independent predictor of relapse (cause-specific hr . %ci po . ) . in a mixed effect model with weight loss as outcome only ipss prior to allosct had significant impact on weight loss (p ¼ . introduction: hla-c-encoded kir ligands (c /c ) have been identified as important factors for the outcome of unrelated allogeneic hsct: in a previous retrospective study cml recipients bearing at least one c ligand showed worse survival when compared to c c recipients (hr . , po . ), especially when peripheral blood progenitor cells (pbpc) were used or in advanced disease stages. these initial findings were confirmed in a second cohort in advanced aml/cml, (but not in mds or all/nhl) receiving pbpc. notably, hla-c allele matching contributed differentially to the transplantation outcome: it was beneficial in c patients, but was detrimental in c recipients (increased trm, hr . , po . ; increased relapse, hr . , p ¼ . ). we hypothesized that c patients have a high frequency of immuno-competent nk cells (icnk) enabling eradication of residual disease due to the genetically hard-wired sequential acquisition of kir receptors during early reconstitution phase post hsct, with c -specific nk cells emerging first. alloreactive t cells -resulting from hla-c mismatch-might thus not have an additional beneficial effect in c patients and might even be detrimental (e.g. increased gvhd), but may serve an important function in relapse control in c patients. the lack of disease control in hla-c-matched c patients would thus be explained by a combination of insufficient numbers of icnk cells in the early phase and the lack of alloreactive t cells later post hsct. consequently, this group exhibited poorest clinical outcome of all four groups defined by recipients kir ligands and hla-c allele matching in the investigated cohorts. materials (or patients) and methods: the aim of the present retrospective study was to determine the influence of hla-c allele matching on the background of hla-c encoded kir ligand status in a large patient cohort (n ¼ , provided by cibmtr). statistical analysis was performed by cibmtr. patients received unrelated allografts between and , with the majority of patients after ( %). % of the recipients were younger than y, % younger than y. % of patients had early, % intermediate, and % advanced disease (aml %, all %, cml %, mds %). % received bone marrow, % pbpc. % received a myeloablative conditioning. endpoints were os, agvhd ii-iv and iii-iv, extensive chronic gvhd, relapse, dfs, and nrm. due to multiple comparisons and multiple endpoints, po . was considered as significant. all models were adjusted for significant clinical covariates. stratification was used in cases of non-proportional hazards. patient-donor pairs were classified according the recipient hla-c kir ligand expression (c c , or c ( ¼ c c or c c )) and the degree of the hla-c allele match: results: introduction: hepatic veno-occlusive disease (vod), also called sinusoidal obstruction syndrome, is a potentially fatal complication of hematopoietic stem cell transplantation (hsct). severe vod (svod), clinically characterized by multiorgan failure (mof), has been associated with a % mortality rate; it may develop in a substantial number of highrisk patients (pts). defibrotide (df), a sodium salt of complex single-stranded oligodeoxyribonucleotides, is thought to protect injured endothelium and to restore thrombo-fibrinolytic balance. in a phase trial in svod, df improved complete response rate and survival at day þ post hsct vs historical controls, with a favorable safety profile. in the european union, df is approved for treatment of severe hepatic vod in hsct therapy in adults and children. in the us, df is available through an expanded access, protocol-directed treatment ind (t-ind) collecting data on safety/efficacy in children and adults with svod and non-severe vod post hsct or post chemotherapy (ct). the original t-ind protocol required vod diagnosed by baltimore criteria (total bilirubin z . mg/dl with z of hepatomegaly, ascites, or % weight gain) plus mof (renal and/or pulmonary) following hsct; the study was amended to include non-severe vod (ie, materials (or patients) and methods: in a single center retrospective study patients who underwent allogeneic hematopoietic stem cell transplantation (hsct) for various diseases ( % acute leukemia) were genotyped for cyp b (c g) expression and their influence on outcome was analyzed. genotyping of cyp b (c g) was performed by real-time pcr.results: patients ( %) were genotyped as homozygous wild-type (wt) gene c/c, ( %) as heterozygous genotype c/g and ( %) as homozygous gene mutated g/g. calculated genotype frequencies did not differ from that reported earlier by other studies for caucasians. patients' demographic and treatment characteristics showed no difference between the three groups except that cyp b cc was more common in females % than in males % (po . ).five-year estimate for overall survival (os) was þ % for the cc group and þ % for the c/g-and g/g groups (po . ). surprisingly, this difference was primarily evident in males (po . ), where the group with cyp gene mutations did significantly worse ( -year estimate for os: þ % vs. þ %), whereas it was virtually absent in females (p ¼ . ). trm and rr were higher for the group with mutated genes in regard to the group with wt gene (although not significant). . in phylogenetic analysis of the e gene, a two-step pcr amplification of almost the entire adenovirus e gene was performed using primers designed from the known sequence of the hadv a reference strain (am . ). results: all patients had been admitted to the ward, but the two last patients (patient and ) had no timely connection to other known hadv a cases ( figure) . in addition, four of the patients ( , , and ) made visits to the out-patient clinic on the same day as one or several other hadv positive patients.sequencing the hexon gene resulted in % homology between the patient samples but also to the reference strains of hadv a (accession number ab . (cmv) is an important cause of morbidity after allogeneic hematopoietic stem cell transplantation (hsct). the latent virus reactivates in immune-compromised patients, due to both post-hsct immunosuppressive therapy and impaired t-cell reconstitution/function. here we report the impact of mismatches in hla-molecules between donor and recipient on cmv-reactivation and cmv-specific immune reconstitution. materials (or patients) and methods: this retrospective study included patients, who received a transplant from a matched related donor (mrd n ¼ ), matched unrelated donor (mud n ¼ ), mismatched unrelated donor (mmud n ¼ ) or mismatched related donor (mmrd n ¼ ). hlatyping ( / ) of patients and donors was conducted via highresolution multiplexed pcr. blood samples were routinely monitored for cmv pp antigen expressing cells per , leukocytes. cmv-cytotoxic lymphocytes (cmv-ctl) reconstitution was analyzed in the blood from patients at days þ , þ , þ , þ after hsct, using hla-cmv tetramers. results: the fisher's exact test was used to analyze the data. the outcome was that hla mismatch (class i or ii) showed significant influence on recurrent (multiple) cmv reactivations (mcmv-r) (po . ). we analyzed the relative risk (rr) in the subgroups with different levels of hla-matching for cmv-r and mcmv-r. the group transplanted from mrd served as reference (ref. ) . shortly, we found significantly higher risk for mcmv-r in the mmud group (rr . , % ci . - . , p ¼ . ). in the mrd ( %) patients had mcmv-r, while in the mud ( %), in the mmrd ( %) and in the mmud group ( %) had mcmv-r. furthermore, we investigated the mean numbers of cmv-ctl/ ml of blood in the groups with different levels of hla-match. we divided cmv-ctl levels into ranges: o , - and cmv-ctl/ml. in the mmud group we observed a trend for an increased risk for the lack of cmv-ctl ( ( %) patients; rr . , % ci . to . , p ¼ . ) compared to ( %) patients from the mrd group. significantly less patients ( ; %) had more than cmv-ctl from the mmud group (rr . , % ci . to . thus the focus of the present study was the selection of hadvstreptamer þ t-cells and ebv-streptamer þ t-cells.materials (or patients) and methods: cells from leukapheresis healthy donors were prepared in large ( À  ) and small ( x ) cell batches. whereas the larger batch was directly labelled with streptamers to select hadv and/or ebvspecific t-cells (large-scale), the smaller batch was used to generate in vitro virus-specific t-cell lines before streptamerlabelling for streptamer selection (small-scale). isolation of hadv-and/or ebv-specific t-cells was performed using the clinimacs device.results: the purity of hadv-streptamer þ t-cells among cd þ cells, obtained from large-scale selection was only . %, but reached up to % when hadv-and ebvstreptamers were applied simultaneously. a further increase in purity of hadv-specific t-cells reaching up to % was achieved by small-scale selection. all final products fulfilled the microbiological and chemical release criteria. ifn-g-response indicating functional activity was seen in / hadv and / ebv large-scale selections and in / hadv small-scale selections.conclusion: the use of hadv-streptamers for clinical applications is feasible particularly when combined with other streptamers or when performed after a previous in vitro expansion period. in this cohort of t-cell replete haplo hscts using post-transplant high-dose cyclophosphamide, we found that a higher intensity of conditioning (mya or ric vs non-mya) as well as the use of more immunosuppressive calcineurin inhibitor (fk vs csa) were both significantly associated with a higher incidence of pv-hc. results: in cd þ lin -cd cells, probes were deregulated between patients without agvhd and patients with agvhd ( of this probe were up-regulated and were down-regulated, po . , fold change . ) . in cd þ lin -cd þ cd progenitors, probes were deregulated between patients without agvhd and patients with agvhd ( of this probe were up-regulated and were downregulated, po . , fold change . ). probes were deregulated in both cd þ lin -cd þ cd and cd þ lin -cd populations. genes from ribosome protein biogenesis, translation machinery (eef d, eef g, eif k) and cell cycle (ccnd , cdk ) were over-expressed in cd þ lin -cd þ cd and in cd þ lin -cd populations from patients without agvhd compared with those from patients affected by agvhd and from healthy donors. expressions of genes from the oxidative phosphorylation metabolic pathway (ndufs , sdha, atp a ) and genes involved in stress resistance (btg , mgst , hpx) were specifically increased in cd þ lin -cd þ cd lymphoid progenitors and not in cd þ lin -cd non-lymphoid progenitors from patients without agvhd compared with patients suffering from agvhd and from healthy donors. we show for the first time that circulating lymphoid t-cell progenitors undergo profound changes in metabolism favoring energy production and response to stress after allo-hsct in humans. these mechanisms are abolished in case of agvhd, indicating a persistent cell-intrinsic defect in addition to the impact of agvhd on the bone marrow environment. disclosure of interest: none declared. introduction: post transplant interventions such as donor lymphocyte infusion (dli) or administration of pharmacological agents, represent important novel strategies with the potential to reduce the risk of disease relapse after allo-sct in acute myeloid leukaemia (aml). such approaches are critically dependent on timely intervention post-transplant but despite this the factors determining the kinetics of disease relapse in patients allografted for aml have not been defined. materials (or patients) and methods: adults who received an allo-sct for aml in first complete remission (cr ) between and were studied. patients were transplanted using a sibling donor and from an adult matched-unrelated donor. patients received a myeloablative conditioning (mac) regimen and a reduced intensity (ric) regimen. a series of landmark analyses were performed at , and months in order to identify prognostic factors of relapse for patients alive and well at the beginning of each time interval. the probabilities of relapse were calculated by using the cumulative incidence estimator to accommodate for death as a competing risk. factors predicting relapse were studied using cox regression model including time dependent variables. a backward stepwise procedure was used for variable selection. results: with a median follow-up of months, patients relapsed. the year cumulative incidence of relapse was % [ %ci: - ]. overall % of patients destined to relapse did so within the first year post-transplant. the overall factors predicting disease relapse for the whole population were more than one course of chemotherapy to achieve cr , flt itd positivity, adverse risk cytogenetics, shorter interval from cr to transplant. the occurrence of acute gvhd grade ii or greater (p ¼ . ) and chronic gvhd (p ¼ . ) were both associated with a lower risk of relapse. using landmark analyses the factors determining relapse at different stages post transplant were observed to differ. in the first months post-transplant the significant factors determining relapse risk were: patient age (p ¼ . ), prolonged interval from diagnosis to cr (p ¼ . ), flt itd positivity (p ¼ . ), adverse risk cytogenetics (p ¼ . ) and use of in vivo t cell depletion (p ¼ . ). the only factors observed to determine relapse risk between and months post-transplant were introduction: the number of haematopoietic stem cell transplants being performed worldwide is increasing as is interest in side effects. despite the increase in published data on late effects in the last decade, data on very long term survivors ( years) is lacking. in this study we describe the outcome of all the patients transplanted at our centre more than years ago. materials (or patients) and methods: between june -january , patients had received allogeneic sct for haematological malignancies at the hammersmith hospital. most patients ( / ) were transplanted for cml with cy/ tbi conditioning and the majority were in chronic phase at sct (n ¼ ). at the time of analysis in december , / ( %) patients had died. of presumed survivors, patients had moved abroad and an additional patients were considered lost to follow up as they had had no contact with our centre within years of the study. of the remaining patients, detailed follow up information was available for . results: the majority of deaths ( / ) occurred within years of sct. a further patients died between - years after sct the most frequent cause of death being relapse ( / ) and infection ( / ). between - years after sct there were deaths; the most frequent causes were relapse (n ¼ ), second malignancy (n ¼ ) and gvhd (n ¼ ). between - years there have been deaths and the most frequent causes were second malignancy (n ¼ ) and respiratory (n ¼ ). the latest recorded relapse was at . years. for survivors for whom we had detailed follow up information the median follow up time was y months (range y months - years months). the median age at follow up was y (range - ). / patients had had a diagnosis of cancer at the following sites: skin (bcc or melanoma) n ¼ , oral or tongue n ¼ , oesophagus n ¼ , breast n ¼ and a further patient had had testicular and bladder cancer. / patients had dyslipidaemia and / were being treated for hypertension. / were diabetic and / were hypothyroid. of male patients, had low testosterone levels requiring treatment. / had vascular complications including three with ischaemic heart disease one of whom also had pvd, two with venous thrombosis, one with a tia and two patients with renovascular disease. dxa data was available for / of these patients and bmd was recorded as low (osteopenia or osteoporosis) in / . conclusion: we conclude that late deaths more than y after sct are more likely to be due to second malignancy than relapse. appropriate screening identifies a large number of abnormalities in the surviving patients most of which are amenable to treatment. disclosure of interest: none declared. the aim was to evaluate the survival and late toxicities, defined as any disease condition other than lymphoma occurring after at least months after asct. the median patient age at asct was years (range, - ). all patients relapsed after at least one chemotherapy line (previous treatments range - ), % of them received radiotherapy. at asct, % of patients were in cr, % in pr, % stable, % in progression. full dose beam was given to % of patients while % received dose reductions for comorbidities. results: the median follow-up was . years (range . - . ) . the -year os and pfs were and % (median not reached for both). the non-lymphoma-associated mortality was %, % and % at , and years of follow-up. the os was impacted in multivariate analysis by disease status before asct (p ¼ . , hr . , ci % . - . ), and radiotherapy (p ¼ . , hr . , ci % . - . ) . pfs was impacted by female gender (p ¼ . , hr . , ), pre-transplant disease status (p ¼ . , hr . , ), and radiotherapy (trend, p ¼ . , hr . , . none of the factors analyzed impacted late non-lymphoma-associated mortality, except for a trend given by age (p ¼ . ). late toxicities after beam asct occurred in % of patients, and included infection ( % -most frequently pulmonary), hypogammaglobulinemia ( %), pulmonary complications ( % -mostly reduction of pulmonary function tests scores), metabolic syndrome ( %), cardiovascular complications ( %), second tumors ( %), hypothyroidism ( %), diabetes ( %), chronic kidney failure ( %), hepatitis b reverse seroconversion ( %) and ocular complications ( %). the cumulative incidence of second tumors was , , and % at , , and years of follow-up, and reached a plateau of % at years of follow-up. patients had a second cancer, of whom had a solid tumor (skin [ ] , colorectal, prostate, lung [ each], breast and oropharingeal [ each]), a hematologic tumor (secondary mds or aml [ ] or nhl [ ] ). age (p ¼ . , hr . per year, ci % . - . ) , and male gender (p ¼ . , hr . favorable for female sex, ci % . - . ) , increased risk of a second tumor. of patients who died without lymphoma, died of second tumors, died of cardiovascular complications, of late infections, and for other causes. in multivariate logistic regression, the incidence of second tumors was associated with age (p ¼ . ), and there was a trend for patients receiving radiotherapy for late cardiovascular complications (p ¼ . ). conclusion: beam conditioning is associated with a % crude incidence of late effects, mostly infections, hypogammaglobulinemia, and pulmonary complications. the most important preventive measures for late mortality could be the screening for cancer, especially for older patients, screening for heart disease particularly for patients receiving radiotherapy, and prompt and aggressive treatment of late infections. disclosure of interest: none declared. introduction: the advent of highly active antiretroviral therapy (haart) in had led to a suppression of hiv viral load, to an improved immune function resulting in a significant reduction of opportunistic infections and hiv related morbidity and mortality. consequently, more intensive treatments, including autologous stem cell transplantation (asct), have been extended also to the hiv-positive population. however, in the literature data are scarce concerning the long-term events (incidence of lymphoma relapse, of second cancers and aidsdefining conditions) in hiv-positive patients (pts) affected by relapsed lymphoma who underwent asct. materials (or patients) and methods: we treated hivpositive pts affected by relapsed/refractory lymphomas with asct consecutively in our cancer center. ten pts died during or early after asct due to progressive disease ( pts), chemotherapy toxicity ( pt) and infection ( pts). we analyzed the post-transplantation long-term data of hiv-positive lymphoma pts, reaching a complete response after asct. eighteen pts were male ( %) and pts were female. our cohort of pts included non-hodgkin's lymphomas (nhl) and hodgkin's lymphomas (hl), respectively. twenty-two pts ( %) received one, and pts received two second-line chemotherapy regimen before asct, respectively. the majority of the pts were submitted to a single (beam conditioning regimen) and only two pts to a tandem asct procedure (high dose melphalan followed by beam conditioning regimen). all pts received haart concomitantly to cancer treatment. results: two pts experienced a lymphoma relapse, after . and . years from asct, respectively. three pts presented with a secondary malignancy ( pt an anal squamous cell cancer, pt a squamous cell carcinoma of the larynx and pt a cin , respectively), with a median time of . years from asct. eight pts had opportunistic infections (oi): pts developed a pneumocystis carinii pneumonia, pt a cytomegalovirus pneumonia, pt a mycobacterium avium complex pneumonia, pt a herpes simplex chronic ulcer, pts cutaneous relapsing herpes zoster, respectively. the median time of oi appearance was . years (iqr: . - . ). two pts died: one of lymphoma relapse, the other of car accident. with a median of -years follow up (iqr: . - . ) the os and pfs of the entire sample of pts were % and % at years, respectively. our results may be summarized as follows: ) out of pts are still alive and in long-term complete remission after asct. these data confirm the long-term efficacy of asct in hiv-positive pts affected by relapse/ refractory lymphoma. ) the appearance of oi is earlier than that of second malignancies after asct. ) the secondary malignancies developed by our pts are non-aids-defining cancers, in agreement with the increased incidence reported by the literature in the haart-era and at least two cases are linked to a viral pathogenesis (hpv for both anal cancer and cervical cancer precursor lesion). ) both oi and second malignancies in our pts series were successfully managed and cured and the only long-term death occurred due to lymphoma relapse. disclosure of interest: none declared. introduction: allogeneic hematopoietic cell transplantation (hct) is an effective therapeutic option for high risk hematological malignancies; % of those who survive the first years are expected to become long-term survivors. the prevalence of chronic health conditions approaches % among hct survivors and that for severe or life-threatening conditions exceeds %. materials (or patients) and methods: a standardized followup of hct survivors is applied at our center, according to jacie standards. here we report the analysis of data collected between nov and nov in adult patients (ptsmedian age at follow-up y -r - ) who underwent an hct between and at our institution. data on items were prospectively collected in an institutional database. a written consent was given by pts allowing the use of medical records for research in accordance with the declaration of helsinki. results: overall % of pts received an haplo, % a mud and % a match related hct; pts deceased in the last year ( because of disease relapse, of late major infectious complication, of second cancer). at a median follow-up of y (r - ; cumulative follow-up y) we observed: -chronic graft-versus-host-disease (c-gvhd): at a median follow-up of y (r - ) ( %) pts are presenting c-gvhd features. according to nih consensus criteria cases were classified as mild, moderate, severe. median number of involved organs (r - ), pts were experiencing skin lesions, eyes impairment, mouth alterations.-late infectious manifestation: ( %) pts present late infections, pts deceased. pneumonia was reported in pts, varicella zoster virus reactivation in , encephalitis in ( virus related, toxoplasma related), hepatitis in , ebv reactivation in .-second cancers: second malignancies were diagnosed in ( %) pts, pts are actually under work-up for diagnosis. nonmelanoma skin cancer was the most frequent diagnosis ( cases); pts were diagnosed with cervix cancer, with prostate cancer, with lung cancer and with bladder cancer. single cases of thyroid, parathyroid, colon, gastric, kidney, larynx, endometrial and breast malignancies were also reported. all pts were treated according to standard policy for general population, / are alive.-thyroid dysfunction: ( %) pts presented overt hypothyroidism.-cardiovascular diseases: arterial diseases were reported in pts, atrial fibrillation in and cardiomyopathy in pts -overall %.-metabolic syndrome (ms): ( %) pts were presenting features of ms ( / features among hypertension, dyslipidemia -raised triglycerides and lowered high-density lipoprotein cholesterol-, raised fasting glucose and central obesity). -secondary hemosiderosis: iron overload was documented (with mri and blood parameters) and treated in pts ( %). according to donor source no difference were observed (chisquare test -p ns) except for higher incidence of moderate/ severe gvhd incidence in match related hct (p . ) as compared to alternative hct. conclusion: hct survivors are at a defined relevant risk of developing long-term complications that have a direct impact on quality of life, morbidity and mortality. introduction: long-term survivors of allogeneic hsct now form an expanding and unique patient population with often complex physical and psychological late effects (le) and associated unmet needs. despite international guidelines , optimal delivery models of le services are unclear from clinical, organisational and economic viewpoints. materials (or patients) and methods: in order to scope current models of care for le service delivery within the uk, we undertook a survey of the nhs adult allogeneic hsct centres during . centres were invited to participate in an online survey composed of questions examining service organisation, multi-disciplinary team (mdt) provision, access to other specialist services and patient engagement. results: a % response rate was achieved from programme directors or delegated specialist staff. around half of centres also treated patients r years and all centres had achieved or were working towards jacie accreditation. in % of centres, the le service was led, coordinated and delivered by consultant medical staff, with the remainder being nurse-led. most centres ( %) provided follow-up in a dedicated allograft or le clinic for the first year, but thereafter attrition resulted in b % of patients being followed after years, and b % after years. most centres had easy access to medical specialities necessary for le management, but specialist interest in long-term hsct complications was uncommon. only % of centres had access to a le mdt, often limited to patients o years. despite specific jacie competency s standards, a third of centres had held no le educational event in the previous three years. most centres ( %) had an sop for long-term monitoring and le management, with the focus predominantly on physical le with only % including a formal psychological screening assessment. only % of centres had audited the performance of the sop. screening for endocrinopathies, iron overload and cardiovascular complications was near universal, but access to mammography and cervical smear testing was more limited. revaccination rates were high, but only % of centres routinely tested antibody responses. despite recommendations, most ( %) centres never used standard templates to communicate le risk to gps or referring consultants. only % of centres had a patient support group accessible to hsct survivors with equivalent numbers having undertaken patient satisfaction surveys related to le service provision. the most commonly perceived barriers to implementation of le services were funding of psychological and other clinical staff and extra investigation costs.conclusion: this survey has demonstrated variation and limitations in the provision of long-term follow up of allogeneic hsct survivors within the uk nhs. although patients are seen in specialist clinics and have access to other specialities, there are limitations in sustaining long-term screening, mdt working, education, audit and patient engagement, as well as perceived barriers to resourcing staff and investigations. further work is warranted to optimise effective, sustainable and affordable models of care for delivery of le services in this expanding specialised patient population. references: . majhail et al. biol hematology department, hospital sant pau, hematology department, hospital vall d'hebrón, epidemiology department, hospital de sant pau, hematology department, hospital del mar, barcelona, spain in vivo and to unravel the requirements for their long-term persistence directly in humans. materials (or patients) and methods: we studied the immune system of patients who underwent haploidentical hsct and infusion of donor lymphocytes transduced to express tk suicide gene (median dose: . x cells/kg) for high-risk hematologic malignancies. in case gvhd, proliferating tk-cells were promptly eliminated upon ganciclor (gcv) administration with complete resolution of the adverse reaction without immunosuppressive treatments. results: at a median follow-up of years after hsct (range - . ), a complete recovery of nk cells, b lymphocytes and ab or gd t cells was observed. the cd þ and cd þ t cell compartment of tk patients were characterized by level of naïve and memory cell comparable to age and sex matched healthy controls. tk-cells were detected in all patients, at low levels (median ¼ cells/ul), even in patients treated with gcv. ex vivo selection of pure tk-cells confirmed the presence of functional transduced cells, thus directly demonstrating the ability of memory t cells to persist for years. importantly, gcv sensitivity was preserved in long-term persisting tk-cells, independently from their differentiation phenotype. longitudinal follow up revealed that tk-cells circulated in patients at stable levels and displayed a conserved phenotype comprising effector memory (t em ), central memory (t cm ) and stem memory (t scm ) t cells. the low level of ki- positivity suggested the maintenance of a pool of gene-modified memory cells through homeostatic proliferation. polyclonality was demonstrated by sequencing among tk-cells of thousands of diverse tcrs with a broad usage of v and j alpha and beta genes. the number of tk-cells persisting at the longest follow-up did not correlate with the amount of infused cells, but instead with the peak of tk-cells measured within the first months after infusion, suggesting that antigen recognition is dominant in driving in vivo expansion and persistence of memory t cells. accordingly, we documented the persistence of cmv and fluspecific tk-cells only after post-transplant cmv reactivation or after flu infection. we observed that the number of infused t scm cells positively correlated with early tk-cell expansion and with their long-term persistence, suggesting that t scm might play a privileged role in the generation of a long-lasting immunological memory. conclusion: after infusion, gene-modified memory t cells persist for up to years within a physiological immune system. antigen exposure and a t scm phenotype were associated with long-term persistence of infused tk-cells. further studies on tk-cell tcr repertoire and vector integrations are currently being performed to elucidate the in vivo dynamics of infused memory t cells. use of zoledronic acid after tcrab/cd -depleted haploidentical transplantation to enhance gd t cells anti-leukemia effect p. merli introduction: hsct is a potentially curative option for a number of malignant disorders; however, up to % of patients lack a suitable hla-matched either related or unrelated donor. in order to optimize haploidentical transplantation, we recently developed a new method of graft manipulation (i.e. tcrab/cd negative selection), which retains in the final product large numbers of effector cells, namely nk and tcrgd lymphocytes. relapse remains the main cause of treatment failure. based on preclinical data showing bisphosphonates-mediated improvement of tcrgd cells-blast killing through accumulation of phosphoantigens, we started a prospective trial based on post-transplant infusion of zoledronic acid, with the aim of enhancing tcrgd cells anti-tumor effect. materials (or patients) and methods: enrolled in the study were pediatric patients (median age at transplantation . years, range - ) affected by either all and aml ( and patients, respectively) at very-high risk for relapse/trm due to disease status (cytogenetic/molecular characteristics, lack of remission or previously failed hsct). all of them underwent a tcrab/cd -depleted hsct from an hla-haploidentical donor (one of the two parents). according to the model of kir/kir ligand mismatch, patients were transplanted from an nk-alloreactive donor. the median number of infused gd þ t cells was . x /kg (range . - . ) . zoledronic acid was administered monthly at the dose of . mg/kg per dose (maximum dose mg), starting from day þ after transplantation. patients underwent zoledronic acid infusions, together with oral calcitriol and calcium supplementation, in the outpatient unit. results: a total of infusions were administered with a mean of . infusions per patient (range [ ] [ ] [ ] [ ] [ ] ; only one episode of symptomatic hypocalcemia (at first administration) occurred and was rapidly corrected with parenteral calcium supplementation. none of the patients experienced de novo onset or worsening of previously developed acute gvhd, this finding supporting the observation that gd t-lymphocytes do not cause gvhd. in the study period, six patients relapsed and died due to infectious complications. with a median follow up of months (range - ) the -year kaplan-meyer estimate of os and lfs were . % (se . ) and . (se . ), respectively ( figure a) . the cumulative incidence of relapse and trm were . % and . %, respectively (figure b) . repeated infusions of zoledronic acid (i.e. more than ) seem to offer an advantage in terms of dfs ( . % vs . %, p ¼ . ), although the difference was not statistically significant (figure c and d) .conclusion: these data suggest that the infusion of zoledronic acid after tcrab/cd -depleted haplo hsct is safe. repeated infusions of zoledronic acid after haploidentical hsct seems to be more effective in preventing leukemia recurrence. more patients and a longer follow-up are needed to establish the efficacy of this approach. disclosure of interest: none declared. inducible t-cell receptor expression in precursor t-cells for leukemia control introduction: the co-transplantation of hematopoietic stem cells (hs) with those that have been engineered to express tumor-reactive t cell receptors (tcrs) and differentiated into precursor t cells (prets) may optimize tumor reduction. since expression of potentially self-(tumor-) reactive tcrs will lead to negative selection upon thymic maturation, we investigated whether prets forced to express a leukemia-reactive tcr under the control of a tetracycline-inducible promoter would allow timely controlled tcr expression thereby avoiding thymic negative selection. materials (or patients) and methods: using lentiviral vectors, murine lsk cells were engineered to express a tetracyclineinducible tcr directed against a surrogate leukemia antigen. tcr-transduced lsk cells were co-cultured on t cell development-supporting op -dl cells to produce prets. lethallyirradiated b /ncrl recipients received syngeneic t celldepleted bone marrow and  syngeneic or allogeneic (b .a) tcr-engineered prets. an otherwise lethal leukemia cell (c ) challenge was given days later. results: after in vivo maturation and gene induction up to % leukemia free survival was achieved in recipients of syngeneic tcr-transduced prets (po . ) as shown in figure a. importantly, transfer of allogeneic gene-manipulated prets increased the survival of recipients (po . ) without inducing graft versus host disease (gvhd). nontransduced prets provided significantly lower leukemia protection being not significantly superior to the pbs controls. the progenies of engineered prets gave rise to effector and central memory cells providing protection even after repeated leukemia challenge. in vitro transduction and consecutive expansion of mature t cells required at least  cells/ recipient to mediate similar anti-leukemia efficacy, risking the development of severe gvhd if of mismatched origin, and providing no long-term protection. importantly, while transgene induction starting immediately after transplant forced cd þ t cell development and was required to obtain a mature t cell subset of targeted specificity, late induction favored cd differentiation and failed to produce a leukemiareactive population due to missing thymic positive selection. conclusion: co-transplanting tcr gene-engineered prets is of high clinical relevance since small numbers of even mismatched hs can be transduced at a reasonable cost, expanded in vitro, stored if needed, and provide potent and long lasting leukemia protection. disclosure of interest: none declared. key: cord- -yg salqt authors: nan title: oral sessions and working party date: - - journal: bone marrow transplant doi: . /bmt. . sha: doc_id: cord_uid: yg salqt nan basic science award: o mt -mmp and reck inversely regulate haematopoietic progenitor cell egress a. avigdor* ( ) , y. vagima ( ) , p. goichberg ( ) , o. kollet ( ) , s. shivtiel ( ) , m. tesio ( ) , a. kalinkovich ( ) , i. petit ( ) , o. perl ( ) , e. rosenthal ( ) , i. resnick ( ), i. hardan ( ) , a. nagler ( ) , t. lapidot ( ) ( )the chaim sheba medical center (tel-hashomer, il) ; ( ) the weizmann institute of science (rehovot, il); ( )hadassah medical center (jerusalem, il) hematopoietic progenitor cell release to the circulation is the outcome of signals provided by cytokines, chemokines, adhesion molecules, and proteases. yet, the mechanisms of progenitor cell egress during g-csf mobilization are not fully understood. membrane type- metalloproteinase (mt -mmp) and its endogenous inhibitor, reck, are established key regulators of tumor cell motility. we detected higher mt -mmp and lower reck expression on circulating human cd + progenitors and maturing leukocytes as compared to immature bone-marrow (bm) cells. mt -mmp expression was more prominent on cd + cells obtained from pb of g-csftreated healthy donors whereas reck was barely detected. g-csf mobilization in nod/scid mice, previously engrafted with human cells, increased mt -mmp and decreased reck expression on human progenitors and maturing leukocytes, in a pi k/akt -dependent manner, resulting in elevated mt -mmp activity. blocking mt -mmp function impaired g-csf mobilization, while reck neutralization promoted egress of human cd + progenitors. targeting mt -mmp expression by sirna or blocking its function reduced the in-vitro sdf- induced migration of human progenitors via matrigel and impaired the bm homing capacity of transplanted human progenitors in nod/scid mice. in accordance, neutralization of reck function facilitated the migration of human bm cd + cells in vitro. furthermore, following g-csf mobilization, we also observed a reduction of cd on human cd + progenitors in the bm of chimeric mice. this was accompanied by accumulation of cd cleaved products of molecular weights, expected for mt -mmp activity, in the bm supernatants. blocking mt -mmp function in chimeric mice resulted in less cleavage of cd upon g-csf mobilization, whereas in the absence of a mobilizing signal, increasing mt -mmp activity by reck ab injection facilitated cd proteolysis on the bm cells. finally, mt -mmp expression correlated with the number of cd + cells, collected on the first apheresis day in consecutive healthy donors and patients mobilized with g-csf. in conclusion, our results indicate that g-csf inversely regulates mt -mmp and reck expression on cd + progenitors, resulting in net increase in mt -mmp activity. mt -mmp proteolysis of cd diminishes progenitor adhesion to bm components, leading to cell egress. these previously undefined cell autonomous changes in the course of g-csf treatment might serve as target for new approaches to improve mobilization. morbidity and mortality associated with treatment-related organ toxicity is a major factor limiting success of allogeneic hematopoietic stem cell transplantation (hsct). conservative therapeutic strategies have been ineffective in part, resulting in high rates of progression or complete organ failure and death. over the last decades, solid organ transplantation (sot) has been increasingly used for the treatment of terminal organ failure in hsct recipients. to date, information regarding the use of sot as treatment attempts in patients after hsct is limited. as well the risk factors accounting for the necessity of sot after hsct as well as the incidence and outcome of this therapy are not well defined. a questionnaire survey was carried out within ebmt centres. centres participated in this survey, covering allogeneic hsct between and . cases of sot were identified. in more detail, liver-, kidney-, lung-, and one heart transplantations were performed in different centres. indications for liver transplantation were infections leading to cirrhosis (n= ), sinusoidal obstruction syndrome (n= ), and gvhd of the liver (n= ). rejection of the transplanted liver or terminal organ failure occurred in one patient respectively. other complications after liver transplantation were infections (n= ), bleedings (n= ) and kidney failure (n= ). most kidney transplantations were performed because of chronic kidney failure due to drug toxicity (n= ). transplant rejection and/ or kidney failure did not occur. interestingly, two of the kidney donors were also stem cell donors for the transplant recipient. lung transplantation was performed in all cases because of bronchiolitis obliterans and/ or gvhd which led to respiratory failure. rejection occurred in patients and in one patient terminal transplant failure occurred. other major complications in the lung transplant recipients were kidney failure (n= ) and infections (n= ). heart transplantation was performed in one patient because of pre-terminal heart insufficiency due to drug treatment. in summary, very few sot for terminal organ failures were performed. the overall survival of patients receiving an organ graft after hsct was . % at years with a median follow up time of months. complications after sot, like infections and organ rejection were frequent, but manageable. we conclude that sot offers a viable therapeutic option for patients who develop terminal organ failure after hsct. influence of immunisation timing on the response to conjugate-pneumococcal vaccine after allogeneic stem cell transplant: final results of the ebmt idwp trial c. cordonnier* ( ) , m. labopin ( ) , v. chesnel ( ) , p. ribaud background: pneumococcal infections are causes of death after sct. the efficacy of the polysaccharide -valent vaccine (ppv ) is limited before mo after sct, or if graftversus-host disease (gvhd) . previous studies with the wyeth heptavalent conjugate prevnar® vaccine (pcv ), using different schedules after allogeneic sct, showed a response around - % of the patients. however, the optimal timing of vaccination is yet not defined, and pneumococcal infection may occur early in the first months after sct. our objective was to show that the response to early (e) ( mo) immunization is not inferior to a late (l) ( mo) immunization. methods: patients ≥ year old and at months after allogeneic myeloablative sct were randomized to receive doses of pcv at month interval, followed by a ppv months later, from (e) or from (l) months after transplant. the primary endpoint was the % of responders (≥ . µg/ml of each of the pcv serotypes) month after the rd dose of pcv (s ). ab levels were blindly measured by elisa. all patients were followed until months after transplant, or until death, whichever occurred first. results: patients were randomized: in the early (e), and in the late (l) group. most patients were adults with acute leukemia, transplanted from an hla-identical donor. patients were evaluable for the primary endpoint (e: , l: ). the response rate was respectively % vs % at s , and not inferior in the early, when compared to the late group ( % ci; . ) . however, at months, significantly less e patients were still protected when compared to l patients ( / ; % s / ; %, p=. ). in the lack of difference in the response between groups, the groups were pooled to analyse the impact of transplant characteristics on the percentage of responders at s . donor age (> y) and chronic gvhd were the only factors impairing the response in the multivariate analysis. conclusion: the ab response month after doses of pcv after allogeneic sct is about % and non inferior when started at than at months. we therefore recommend starting immunization at months to offer an earlier protection. however, the e vaccination offers a significantly lower protection at years, suggesting the need for a boost during the second year after e immunization. the authors are grateful to the safety committee: d. engelhard, p. reusser, and p. reinert depletion of the autoreactive immunological memory followed by autologous haemopoietic stem cell transplantation in patients with refractory sle induces long-term remissions through de novo generation of a juvenile and tolerant immune system t. alexander ( ) , a. thiel ( ) , g. massenkeil ( ) , a. sattler ( ) , s. kohler ( ) , h. mei ( ) , h. radtke ( ) , g.r. burmester ( ) , a. radbruch ( ) , r. arnold ( ) , f. hiepe* ( ) ( )charité -universitätsmedizin berlin (berlin, de) ; ( ) german arthritis research center (berlin, de) clinical trials have indicated that immunoablation followed by autologous haemopoietic stem cell transplantation (asct) has the potential to induce clinical remission in patients with refractory systemic lupus erythematosus (sle). to elucidate the mechanisms mediating the beneficial long-term clinical responses, we investigated the immune reconstitution in sle patients receiving asct as part of a monocentric phase i/ii clinical trial. seven patients with sle were evaluated during a long-term follow-up (median follow-up period months) who were immunoablated with cyclophosphamide and rabbit antithymocyte globulin, followed by transplantation of purified autologous cd + haemopoietic stem cells. previous failure of conventional immunosuppression, including cyclophosphamide, had been an inclusion criterion. humoral immunity was evaluated, peripheral t and b lymphocytes were immunophenotyped and frequencies of t lymphocytes specific for distinct antigens of interest were assessed after short-term stimulation ex vivo. in all patients clinical and serological remission was observed, accompanied by disappearance of anti-dsdna autoantibodies and protective antibodies from serum. one patient developed a relapse months after asct. in the responding patients, cd + cd ra+ cd + t cells, i.e. recent thymic emigrants, recurred with a doubling in absolute counts compared to agematched healthy controls until -yr post-transplant (p= . ). absolute numbers of cd + foxp + regulatory t cells (tregs) normalised after asct and tcr repertoires cd + t cells displayed a broad clonal diversity as compared to the pretransplant status. early after asct, often high frequencies of virus-specific effector t cells were detected. autoreactive t cells specific for nucleosomes or smd were not detectable. a normal b cell compartment developed within months after therapy, as compared to the pre-existing b cell deficiencies, which had included naive (igd+) b cell lymphopenia (p= . ), relative predominance of memory (igd-) b cells (p= . ) and expansion of cd high cd plasma blasts. our data demonstrate that the long-term therapy-free clinical remissions observed in sle patients after complete immunoablation and asct are accompanied by a loss of immunological memory and a fundamental reset of the immune system. depletion of the autoreactive memory and reactivation of thymic education probably are the basis for regeneration of self-tolerance and clinical remission. m. themeli* ( ), l. petrikkos ( ) , m. waterhouse ( ) , h. bertz ( ) , n. zoumbos ( ) , j. finke ( ) , a. spyridonidis ( ) ( )university of patras medical school (rion-patras, gr); ( )freiburg university medical center (freiburg, de) we previously demonstrated frequent genomic alterations measured by microsatellite instability (msi) in non-neoplastic epithelial tissues of pts who underwent allogeneic hematopoietic cell transplantation (hct) but not in pts after autologous hct (blood ; : ) . confirmation in larger independent patient cohort and in an in vitro system was needed. buccal samples from unselected pts obtained - days (median ) after allogeneic hct were analysed for msi. control subjects ( healthy and pts after auto-hct, samples) were negative for msi. msi was observed in ( %) allo-transplanted pts. msi+ pts were significantly older than msi-patients (median age y vs y, p< . ). by using logistic regression analysis we found that the relative risk for msi was -fold higher in pts who experienced extensive chronic gvhd as compared to pts with no gvhd. although the median follow up in msi+ pts was significantly lower than in msi-( vs days, p< . ), secondary malignancy ( skin-and adeno-ca but none in the oral cavity) was diagnosed in ( %) of the msi+ pts) and only in ( %) msi-pt (p< . ). other clinical features were not significantly different between msi+ and msi-pts. in an vitro mutation analysis model we tested the hypothesis that an alloantigenic stimulus is substantially involved in the mutation process. briefly, keratinocyte (hacat) cells were transfected with a plasmid vector carrying a neomycin selectable marker, a hygromycin resistance (hygr) sequence and a (ca) repeat. in this system, dna slippage mutations become detectable after hygromycin treatment as hygr+ colonies. the mutant fraction was expressed as the number of hygr+ colonies corrected for relative cell survival. untreated cells served as controls. treatment of stably transfected hacat cells with tnfa ( - ng/ml, h), tgfb ( - ng/ml, h) and supernatant from a mixed lymphocyte culture (mlc, h) didn't cause any detectable induction of genomic instability (gi). treatment with h ( - µÌ, - hours) resulted in a time and dose dependent gi induction (max . fold). cocultivation of hacat cells with stimulated lymphocytes from mlc resulted in a . fold induction of the mutant fraction. in conclusion, our in vivo and in vitro data indicate that "alloantigenic reactions may induce genomic instability in the allotransplanted pts which might predispose them to secondary neoplasia. the ebmt risk score predicts outcome after allogeneic hsct in all haematological disease categories and is independent of stem cell source or conditioning intensity a. gratwohl*, m. stern, j. apperley, t. de witte, j. passweg, v. rocha, a. sureda, r. brand, d. niederwieser information on factors associated with outcome after allogeneic hsct is a prerequisite for risk adapted strategies. five key factors form the basis of the ebmt risk score: stage of the disease (early , intermediate , advanced ), age of the patient (< y , - y , > y ), time interval from diagnosis to transplant (< y , > y ), histocompatibility (hla-id sibling , others ) and donor recipient gender combination (other , female donor for male recipient ). they were identified and validated in several independent series of cml patients, but not yet in other diseases. we examined patients, y of age (median, - y range), . % male with an allogeneic hsct for aml ( ; . %), all ( ; . %), cml ; . %), mds ( ; . %), mps ( ; . %), lymphoma ( ; . %), myeloma ( ; . %) or saa ( ; . %) between and . donor was a hla id sibling in . %, other donor in . %. stem cell source was bone marrow in . %, peripheral blood in . % and cord blood in . %.conditioning was standard in . %, reduced in . %. each risk factor was tested individually by multivariate analysis and confirmed as cumulative dose response risk in all subcategories with two exceptions: stage was not applicable in saa, time interval was not applicable in patients in st cr. cumulative incidence of transplant related mortality (trm) at years increased with the risk score from . % (score , pts) to . % (score , pts), . % (score , pts), . % (score , pts), . % (score , pts), . % (score , pts) and . % (score / , pts). after stratification by risk score, underlying disease had only a minor impact on the rate of trm. inside the risk score categories, trm improved significantly during the period of observation (rr (rr - rr rr - rr since . ). absolute trm rates declined less markedly ( - %; - %; since %) due to a shift towards higher risk patients in more recent years. the ebmt risk score separated risk categories in all diseases, for all donor types, for all stem cell sources and for patients with reduced or standard conditioning. these data show that risk categories for outcome after allogeneic hsct can be defined. they can be integrated into risk assessment algorithms and form the basis for individualised risk adapted strategies when transplant and non transplant strategies are available as treatment options. working party solid tumours reduced-intensity allogeneic transplantation for breast cancer d. blaise* ( ) , a. gonçalves ( ) , s. fürst ( ) , j.o. bay ( ) , c. faucher ( ) , m. michallet ( ) , j.m. boiron ( ), j.y. cahn ( ) , n. gratecos ( ) , m. mohty ( ) , c. chabannon ( ) , g. gravis ( ) , b. esterni ( ) , j.m. extra ( ) , p. viens ( ) ( )institut paoli-calmettes (marseille, fr) ; ( ) centre jean perrin (clermont ferrand, fr) ; ( )chu edouard herriot (lyon, fr) ; ( )chu haut lévèque (bordeaux, fr); ( )chu jean minjoz (besançon, fr); ( )chu cimiez (nice, fr) we initially treated pts with allo sct for advanced metastatic breast cancer. all pts (age: ( - )) underwent asct after the same reduced intensity conditioning (ric) (fludarabine ( mg/m ), busulfan ( mg/kg) and thymoglobulin ( , mg/kg) or tli ( cgy)) from a hlaidentical sibling (bm: %; pbsc: %) followed by csa. prior to asct a median of lines of treatment ( - ) were administered over a period of days . nine pts underwent autologous sct at a median time of days ( - ) prior to asct. all pts were measurable and had a median of metastatic sites ( - ) (liver: %, bone: %, lung: % and brain: %): according to recist criteria, ( %) and ( %) pts had progressive (pd) and stable disease (sd) respectively. none of the pts died from trm. two of them achieved partial remission (pr) at and days respectively (objective response (or): % ( - ). all pts but eventually progressed and died from disease ( year overall survival (os): %( - )). results are dramatically different in regards to disease status at time of transplant. while outcome is uniformly poor for pts with pd, patients with sd achieved a % ( - ) or rate for a % ( - ) os at years with ( %) patients surviving more than years ( +, -and -), which is significantly different from pts with pd. we established that ric-asct can be safely performed in brc pts, whereas highly pd pts do not benefit from this approach. we run a second trial in less advanced disease to confirm encouraging results (present accrual: pts). all patients will be presented. however it seems that curability in brc will be achieved only in pts in the initial disease phase: target population would need a careful selection on individual prognosis factors indicating their poor short term poor outcome: this represents the ultimate goal for future investigations. supported in part by a grant from the french ministry of health (phrc ; phrc ) and a special grant (pole areca) from the association pour la recherché contre le cancer (arc) we have utilized autografting to achieve maximum tumor reduction before proceeding to non-myeloablative allografting. this strategy could provide the benefit of a conventional allograft, but with reductions in the typical acute toxicities and associated mortality of myeloablative conditionings. between september and april , we enrolled patients with metastatic breast carcinoma. median age was years. at the time of autografting, the patients had received a median of (range, - ) previous chemotherapy lines; patients had received hormone therapy, and seven patients had undergone radiotherapy on bone lesions. the primary endpoint of this study was the decrease of non-relapse mortality (nrm) from the current - % noted after myeloablative allografting. patients received autografting at a median of months (range, - ) from the diagnosis of breast cancer. no patient died after transplant. one patient who had been in complete remission and two who had been in partial remission before autografting remained in complete or partial remission. no non-relapse mortality was noted in the first days after non-myeloablative allografting. thirteen patients achieved full chimerism. five patients ( %) developed grade ii-iii acute gvhd, while six patients developed chronic gvhd (five patients with extensive disease) and needed intensive immunosuppressive therapy. we have recently reported a subsequent patient transplanted from her hla-identical sister. disappearance of liver, adrenal, mediastinal, pleural, and diffuse nodes and bone metastases, observed simultaneously with clinical chronic gvhc months after non-myeloablative allografting, suggested a profound graft-versus-tumor effect. renal cell carcinoma (rcc) has recently been identified as being a target for gvt effect. since there has been a number of publications describing gvt effects in patients with rcc undergoing mostly reduced intensity transplantation. at the nhlbi, patients have been conditioned with cyclophosphamide ( mg/kg x ) and fludarabine ( mg/m² x ) then transplanted with a g-csf mobilized blood stem cell allograft from their hla identical or single antigen mismatched related donor. twenty-nine of patients have had disease regression consistent with a gvt effect ( . % cumulative incidence of a complete response + partial response). a better understanding of the immune cells and their target antigens that mediate tumor regression could potentially lead to the development develop more effective hct approaches for solid tumors. recently, t-cells with in vitro tumor cytotoxicity patterns consistent with recognition of minor histocompatibility antigens and tumor restricted antigens have been identified in some responding patients. the identification of tumor restricted antigens targeted by donor immune cells could lead to the development of transplant approaches that enhance gvt effects while avoiding gvhd through tumor vaccination or the adoptive infusion of in vitro expanded donor t cells with tumor antigen specificity. we detected rcc-reactive cd + t-cells by elispot analysis in the blood of several responding patients with metastatic rcc following hct that were absent before transplantation. we successfully generated donor cd + t-cell clones from lymphocytes obtained from these patients that have direct cytotoxicity against the patient's rcc cells. in one responding patient, cytotoxic t-lymphocytes and t-cell clones with rcc-specific tumor cytotoxicity were isolated from the blood after transplantation. utilizing cdna expression cloning, we identified an hla-a -restricted -mer peptide (named ct-rcc- ) to be the target antigen of these rcc-specific tcells (takahashi y, harashima n. et al-j clin invest in press) ct-rcc- -specific t-cells were detected by tetramer analysis in the patient's blood after tumor regression but not before hct. tetramer analysis of hla-a + rcc transplant recipients showed ct-rcc reactive t-cells expanded significantly in all responders in contrast to the non-responders, where only / showed an increase in ct-rcc- reactive t-cells. the genes encoding the ct-rcc antigen were found to be derived from a human endogenous retrovirus (herv)-e previously unknown to be expressed in any human tissues; this herv-e was found to be expressed in the majority of rcc tumor lines and fresh rcc tissue but not in normal kidney cells or other normal tissues. this is the first solid tumor antigen identified using allogeneic t-cells from a patient undergoing hct. these data suggest this herv-e is transcriptionally active in rcc, encoding an immunogenic antigen that is over-expressed in rcc which could be a potential target for cellular immunity. update of the results of high-dose chemotherapy as primary or salvage therapy in germ cell tumours g. rosti* ( ) , u. de giorgi ( ) , p. pedrazzoli ( ) , m. bregni ( ) ( cisplatin-containing regimens cure nearly % of patients with advanced germ cell tumors. hdct has been extensively used in the last years with somehow controversial results. the ebmt it- study on relapsing good-risk patients has not shown any difference in overall survival comparing four courses of standard second line therapy versus one late intensification single shot approach, even if patients achieving cr did significantly better if randomized in the high-dose arm. a phase iii us trial in patients with poor prognosis, treated upfront, even if not showing an advantage for hdct, has shown a significant difference for those with unsatisfactory marker decline. an input of the possible role of hdct in relapsing/refractory patients came from the retrospective data of the indiana university of tandem hdct with carboplatin and etoposide in a large series of consecutive men with metastatic testicular cancer that had progressed after receiving cisplatin-containing combination chemotherapy. this study shows % and % four-year disease-free survival in patients who received hdct as second-line or third-line or later therapy, respectively. as it is a retrospective review, one may argue that the results are biased by patient selection. this does not seem to be the case, however, as even patients with very poor prognosis achieved long-term disease-free survival - % of survivors were classified high-risk by the igcccg classification and % had platinum-refractory disease. it is important to note that all patients in this series received peripheral-blood progenitors as sources of hematopoietic stem cells. this strategy allowed a rapid engraftment, thereby permitting the administration of two courses of high-dose in addition, peripheral-blood progenitors were enriched for cd + hematopoietic cells, a procedure which may have a role in eliminating possible cancer cells from the graft. we believe that, on the basis of the robust data provided by einhorn and colleagues, a well-designed randomized trial of hdct versus conventional-dose chemotherapy should be performed in patients with poorprognostic clinical features who relapse after initial chemotherapy. at present, there should be no debate on the use of tandem-hdct in patients with cisplatin-refractory germ-cell tumors and those who have failed second-line therapy. gitmo and igg (italian group for germ cell cancer) are planning a network of centres in italy to refer such patients for the tandem hdct. autologous stem cell transplantation international multiple sclerosis trial (astims, eudract number - - , supported by ebmt; www.astims.org) is now a multicenter, prospective randomized phase ii study. the primary endpoint is the number of new t lesions on mri. the investigational treatment comprises mobilization with cy and g-csf and conditioning with beam followed by asct and atg compared to monthly i.v. pulses of mitoxantrone at mg followed by gr of methylprednisolone. at the moment patients have been enrolled (january ): in barcelona, in genova, in modena, in florence, in chieti, in reggio calabria and in bergamo. the astims trial is therefore still going on, with the aim to arrive within years from now at the new target of enrolled cases. in the meantime, more than are the ms treated in the world with ahsct and a few phase i/ii are running with the aim to identify the clinical characteristics of the patients who can really take advantage from the procedure or to evaluate the efficacy of low intensity conditioning regimens. the study of all the cases treated in italy in the last years with the same regimen (beam and atg), same inclusion criteria and followed by the same neurohematological teams involved in the prospective study supported by gitmo, showed that patients with a relapsing remitting clinical course respond significantly better than secondary progressive cases to ahsct, indicating the population of patients who have to be selected in the future for the design of prospective studies with a clinical endpoint. the astis trial j.m. van laar* ( ) , d. farge ( ) , a. tyndall ( ) , o. astis investigators ( ) ( ) newcastle university (newcastle, uk); ( ) hopital st louis (paris, fr); ( )basel university hospital (basel, ch); ( )jcuh (middlesbrough, uk) background: high dose immunosuppressive therapy (hdit) and hematopoietic stem cell transplantation (hsct) is a novel treatment for patients with severe systemic sclerosis (ssc) . previous studies showed durable responses in two thirds of patients up to yrs after hsct ( ) . this treatment modality is now further investigated through the astis-trial (autologous stem cell transplantation international scleroderma trial), a prospective, controlled, randomized trial to compare safety and efficacy of hdit + hsct versus monthly i.v. cyclophosphamide in ssc patients at risk of major organ failure or early mortality. objectives: to evaluate whether hdit + hsct is superior over conventional treatment in terms of safety and efficacy in ssc patients, and to assess potential predictive factors of response. methods: ssc patients with early active diffuse disease with or without major organ involvement are eligible. ssc patients randomized to the transplant arm undergo mobilization with cyclophosphamide x g/m², conditioning with cyclophosphamide mg/kg, rbatg . mg/kg, followed by reinfusion of cd + selected autologous hsct. those randomized to the control arm are treated with x monthly i.v. bolus cyclophosphamide mg/m². the primary endpoint is event-free survival, defined as survival until death or development of major organ failure during years follow-up. progression-free survival is the main secondary endpoint. results: one hundred eleven ssc patients have been enrolled in centers per january : male, female, mean age yrs, mean modified rodnan skin score , mean disease duration , yr, mean vc %, mean dlco %. sixty-one patients were randomized to the transplant arm, to the control arm. no unexpected toxicities have yet been observed in either arm with a median follow-up of months (range . grade , toxicities occurred in / transplant patients and in / controls (p= . ). atg-related toxicity led to its discontinuation in / transplant patients. two fatalities in the transplant arm were categorised as probably treatment-related. conclusion: the ongoing astis trial has enrolled patients sofar. treatment-related mortality and number of patients with serious adverse events of stem cell transplantation are lower than previously reported in registry analyses. references: . van laar jm, farge d, tyndall a, on behalf of the ebmt/eular scleroderma study group. the astis-trial, hope on the horizon. ann rheum dis ; : . standard nih or eurolupus cyclophosphamide (cy) protocols and mycophenolate mofetil (mmf) as induction therapy in severe bilag a sle is still associated with % failure, % relapse and % to % death at years in the absence of a single standard treatment worldwide for refractory sle, phase i-ii studies analysed the use of: a) rituximab (anti cd mab) in more than patients showing complete to partial early response around % with relapse in to % of the cases; b) autologous hematopoietic stem cell transplantation (hsct) since under the auspices of the joined ebmt-eular working party, reporting durable remission with reduced or no immunosuppressive drug requirement in %, one-third of whom later relapsed to some degree with a ± % (n= / ) overall survival at years for sle among the hsct procedures registered: in in the ebmt data base. the north american, mostly single centre experience showed higher rates of remission with also some relapses. maintenance immunosuppression after induction of remission may decrease the return of disease activity. this was the basis of the ebmt approved astil trial: a prospective randomized open, multicenter, phase ii b study to compare the efficacy of autologous hsct with rituximab as remission induction, followed by mmf ( g /day) as maintenance in both arms for severe sle patients with disease duration ≤ years since the diagnosis and sustained or relapsed active bilag a sle. this analysed describes the outcome of pediatric patients receiving hematopoietic stem cell transplantation (hsct) to treat severe refractory autoimmune cytopenias. the registry of the ebmt contains data on patients receiving transplants. patients had autoimmune haemolytic anemia ( ), evans's syndrome ( ) , immune thrombocytopenia ( ), pure red cell aplasia ( ) and autoimmune lymphroliferative synrome ( ) . patients were males with a median age at diagnosis of years (range . - years) and a median age at transplant of . years ( - years) . the median disease duration prior the transplant was months (range - months) and all patients failed multiple prior treatments. transplant were autologous for and allogeneic for patients, of these transplanted from an hla identical donor, from a family mismatched donor and from a matched unrelated donor. one patient received autologous transplant while another patient received an allogeneic transplant because a relapse after the first autologous transplant . the stem cell source was mobilized pbsc in transplant, bone marrow in and cord blood in patients. the graft was t depleted in of recipients of autotransplant and of allotransplant recipient. the conditioning regimen used were heterogeneous. patients died of treatment related mortality, in the allo and in the autologous group for a trm of % eights patients had a complete and continous response after the transplantation although of these died for secondary malignancy. patients relapsed after the procedure ( in the allo and in the autologous group) and one of these died for disease progression. patients were not evaluable for response. the present analysis has some limitations because treatment protocols, mobilization and conditioning regimen were heterogenous and doesn't allows a detailed analysis of these factor moreover these preliminary data suggest that autologous and allogeneic hsct may induce response in half of patients with severe autoimmune cytopenia of long duration unresponsive to several therapeutic options. b long-term follow-up of autologous stem cell transplantation for juvenile idiopathic arthritis n.m. wulffraat wilhelmina children's hospital (utrecht, nl) the majority of children with juvenile idiopathic arthritis can nowadays be treated adequately. however despite the use of combinations of antirheumatic drugs, corticosteroids and the newer so called biologicals (blocking the tnf, interleukin or interleukin pathways) a proportion of children with arthritis remain resistant also to these therapies and suffer from a very severe, debilitating and potentially fatal disease. for such children autologous stem cell transplantation (asct) is successfully performed since . here we describe the long term outcome of the initial cohort of children with resistant juvenile idiopathic arthritis, treated with asct. the initial cohort of children was treated with a conditioning regimen containing cyclophosphamide, anti thymocyte globulins and low dose total body irradiation. overall favourable responses were seen, with a drug free remission rate of - %. in the more recent years late relapses were noted with lower percentages for drug free long term outcome. special emphasis is given on cases showing very late relapses, occurring after and years. the observed relapses are often less severe compared to the situation before sct and can be treated successfully with conventional drugs in the majority of cases. more recently, asct was performed in jia children with a fludarabin containing regimen in stead of low dose tbi. with a to year follow up, these patients are all in drug free full remission. allogeneic transplant with an hla matched family donor was reported in jia cases. follow up of and year is sofar show clinical disease remission and tapering of medition. in conclusion, given the favourable long term outcome, sct remains a valuable treatment option for children with drug resistant jia. s multipotent mesenchymal stromal cells in the treatment of autoimmune diseases a. tyndall* ( ) , f. dazzi ( ) multipotent mesenchymal stromal cells ( msc) isolated from the bone marrow and other sites are currently being studied to determine their potential role in the pathogenesis and/ or management of autoimmune diseases. in vitro studies have shown that they exhibit a dose dependent antiproliferative effect on t and b lymphocytes, dendritic cells, natural killer cells and various b cell tumour lines, an effect which is both cell contact and soluble factor dependent. these soluble factors include tgf beta, il- , indoleamine , dioxygenase, il- ra, and hla-g among others. anti proliferative and immunomodulatory mechanisms are probably multiple and most likely due to the induction of arrest of the cell cycle in g /g . a plethora of phenotypic definitions and experimental conditions accounts for some of the variation in in vitro phenomena being reported previous assumptions that msc are immunoprivileged have been challenged by recent animal data in non immunosuppressed hosts. animal models of autoimmune disease and tissue injury (ischemic kidney, chemically induced lung fibrosis and liver toxicity) have mostly shown a positive clinical response, with some early warning signs in a melanoma model concerning tumour surveillance. a limited number of patients suffering from acute graft versus host disease have been treated with msc as well as sporadic case reports and small uncontrolled series in multiple sclerosis and crohns disease. prospective phase i trials are starting in multiple sclerosis and crohns disease and being considered in inflammatory rheumatic diseases. an international interdisciplinary data base has been developed to exploit the collective experience. sirolimus is associated with veno-occlusive disease of the liver after myeloablative transplantation c. cutler*, k. stevenson, h. kim, p. richardson, v. ho, c. revta, r. ebert, d. warren, j. koreth, p. armand, e. alyea, r. soiffer, j. antin dana-farber cancer institute (boston, us) veno-occlusive disease of the liver (vod) is an uncommon but important cause of mortality after allogeneic transplantation. to determine if use of the immunosuppressive mtor inhibitor, sirolimus, is a risk factor for vod, we performed a retrospective review of vod incidence and risk factors at our institution since , when we began using sirolimus. methods: review of electronic medical records of all transplant patients undergoing tbi-based myeloablative transplantation with adult stem cell donors was performed. results: patients transplanted between / and / were identified and stratified by sirolimus use ( exposed, unexposed). sirolimus patients received sirolimus/tacrolimus ± methotrexate; all others received tacrolimus/methotrexate as gvhd prophylaxis. there were no differences in the age, gender, donor-recipient gender match or diagnoses between cohorts. sirolimus patients were more likely to have unrelated or mismatched donors, were more likely to have received pbsc (p< . for both) and were less likely to have gr. ii-iv acute gvhd ( vs. %, p= . ) in comparison with non-sirolimus patients. the incidence of vod in the sirolimus group was % and was . % in the unexposed (rr . , p= . ), but vod occurred later among sirolimus patients ( vs. days, p= . ) . among mrd recipients, the rr was . ( . vs. . %, p= . ). when adjusted for age, gender match, stem cell source, hla match (mrd vs. urd/mismatch), and transplant risk (standard vs. high), sirolimus use remained a significant risk for vod (adjusted or . , p= . ). cause-specific mortality related to vod was similar in sirolimus and non-sirolimus patients. despite the increase in vod, treatment-related mortality was similar in all sirolimus and non-sirolimus patients and among mrd sirolimus and non-sirolimus patients at year ( vs. %, and vs. %, both p=ns). in addition, there was a trend towards increased overall survival (os) for all sirolimus patients ( yr os vs. %, p=ns) and for mrd sirolimus patients ( yr os vs. %, p= . ). in a cox regression model, age > (p= . ), donor match (p= . ) and vod (p< . ) but not sirolimus use (p= . ) were significantly associated with overall survival. conclusions: sirolimus is associated with vod after tbibased myeloablative transplantation. despite this association, transplant outcomes appear equivalent or better than standard tacrolimus/methotrexate based immunosuppression. physical health can be compromised in very long-term survivors after hsct compared to their respective donors but not mental health: a paired analysis t. daikeler, a. rovo*, m. stern, j. halter, j.d. studt, a. buser, d. heim, j. rischewski, m. medinger, a. tyndall, a. gratwohl, a. tichelli university hospital (basel, ch) with the improvement of prognosis, health status and functional well being of long term survivors after hsct become an important issue. we performed a cross-sectional prospective study on long-term survivors and their respective sibling donors at a median follow up of . years (range - ) after hsct. the median age of the recipients and donors at time of the study was . ( - ) and . years ( - ) respectively. both recipients and donors were seen on the same day for evaluation. the short form- ® (sf- ) health survey, which provides a generic health status measurement through items assessing concepts of health was used. three of the items measure physical health (pf, rp, bp), two measure both physical and mental health (gh, vt), and three measure mental health (sf, re, mh). in addition there are two summary scores for physical (pcs) and mental (msc) health. for statistical analysis norm-based scoring (nbs) was applied, where is the mean score, and the standard deviation of a defined general population. paired analysis between donors and recipients were performed for detecting differences. all scored items of recipients as well as of the donors were within the range of one standard deviation of the norm-based population. all scores concerning physical well being except one, (rp), were statistically lower in the recipients than in their donors. in contrast, there was no difference in scores concerning mental well being. this is confirmed by the summary measurements of physical health (pcs) with , in the recipients and , in the donors , (p= . ) and mental health (mcs) with , versus , (p= . ) . physical health (pcs) was lowest in patients with severe chronic gvhd compared to their donors ( , versus , ) (p= . ), age older than years at hsct , versus , (p= . ), older than years at the last control , versus , (p= . ) and for female patients , versus , (p= . ) . none of the factors had a statistical impact on mental heath status (mcs, p> . ). in summary, quality of life of long term survivors after hsct measured with the sf- questionnaire is still within the normal variation of the general population. however, when compared to their respective donors, the physical health status is significantly compromised in the recipients. severe gvhd, older age and female gender are associated with an inferior physical health status. a retrospective analysis of sexual function, fertility and endocrine status in male long-term survivors of allografts for haematological malignancy i.h. gabriel*, r. szydlo, m. klammer, r. patterson, n. swan, n. salooja, e. olavarria, e. kanfer, d. marin, a. rahemtulla, j.f. apperely imperial college healthcare nhs trust, hammersmith hospital (london, uk) steady improvements in the outcome of allogeneic stem cell transplantation (allo-sct) have resulted in significant numbers of long-term survivors and an increasing focus on factors impacting quality of life (qol). post transplant infertility and sexual dysfunction are two such factors. using a questionnaire we audited male survivors of allografting for haematological malignancies at our centre. men ( %) responded. the median age at sct was . yrs. the median time from transplant was . yrs and % were > years from sct ( % > years). % had returned to full pre-sct sexual activity, however, a number of problems in sexual function were reported. % complained of new persistent erectile dysfunction (ed) (normal prevalence - %). ed affected men of all ages ( %, % and % at < years, - years and > years respectively). ed was seen in % of recipients of tbi vs % of those who had not received tbi. % experienced penile glans dryness, previously unreported which appeared to be closely associated with chronic gvhd(p . ). in addition urethral constriction, phimosis, genitourinary infection and peyronies's disease all complicated sct. % of men reported reduced libido with % of ed patients reporting normal libido. % and % of men suffered premature or painful ejaculation respectively. % of male survivors described a negative impact of infertility on themselves or their partners and % had utilised assisted fertility/donor insemination. other problems included dyspareunia, inability to use condoms, sicca syndrome, and poor body image. factors forming barriers to new, or worsening existing, relationships were most prominent in younger patients. compared to values pre-transplant fsh was raised in % by months post-transplant and remained high at yrs (p= . ), indicating long-term damage to sertoli cells. lh was significantly elevated at months in % compared to baseline (p= . ) and normalised by yrs suggesting leydig cell recovery. testosterone levels were normal in % of men at months. lack of physician continuity, language, presence of visiting fellows/students and the impression that physicians are only concerned with managing malignancy were barriers discussing these issues at routine follow-up appointments. a high prevalence of sexual dysfunction exists post-sct. increased awareness of these complications and their effects on patients and their families would permit prompt and appropriate management. secondary malignancies in recipients of allografts for chronic myeloid leukaemia in chronic phase using hlamatched sibling or volunteer donors i.h. gabriel*, s. avery, r. szydlo, n. salooja, e. olavarria, e. kanfer, m. klammer, a. rahemtulla, j. goldman, j.f. apperley imperial college healthcare nhs trust, hammersmith hospital (london, uk) during the last decade imatinib has replaced allogeneic sct as first line treatment for chronic myeloid leukaemia (cml) and second generation tyrosine kinase inhibitors now compete with allografting as second line therapy. however over the same period there have been steady improvements in the outcome of allografting and therapeutic choices are increasingly complex. improved information regarding the long-term side effects of both chemotherapy and allografting might help inform decision-making. we now update the incidence of secondary malignancy in consecutive patients of median age . years, who underwent allografts for cml in first chronic phase ( from sibling and from unrelated volunteer donors). the median follow-up is . yrs. all patients received cyclophosphamide with either total body irradiation (tbi) (n= ) or busulphan (n= ). gvhd prophylaxis was cyclosporin, methotrexate, ± t-cell depletion. the probability of developing a secondary malignancy post allo-sct was determined by the method of cumulative incidence (ci). the overall incidence in this group was . % with patients developing new tumours. % were diagnosed within years, % within yrs, % within yrs. men and women were equally susceptible to secondary malignancies. the overall incidence of developing a new malignancy in our cohort is . % which is higher than in an age-matched population. involved sites included: skin ( including melanoma, squamous cell and basal cell carcinoma), breast ( ), high grade b-lymphoma, ( ), tongue ( ), colo-rectal ( ), cervix ( ), osteosarcoma ( ), testis ( ), bladder ( ), mucoepidermoid ( ), oesophagus ( ), lung ( ), penile shaft ( ), and mds ( ). patients had malignancies. pre-bmt treatment consisted of hydroxyurea alone in , busulphan in and interferon in and some combination of these in others. the ci of secondary neoplasia increased with time reaching . %, . %, . % and . % at , , ,and yrs respectively. patients have died, directly attributable to the secondary tumour. as all patients had the same genetic abnormality and received only minimal prior cytotoxic therapy, this increase in neoplasia is likely to be due to sct. these data highlight the need for close follow up and screening. prospective evaluation of oxygenation index and noninvasive-ventilation in patients suffering from acute lung injury after allogeneic transplantation m. wermke*, s. schiemanck, g. höffken, g. ehninger, m. bornhäuser, t. illmer university hospital (dresden, de) objectives: respiratory dysfunction is a major cause of allogeneic transplant-related-mortality (trm). little is known about how to recognize and treat acute lung injury (ali) in this setting. we present the first prospective randomized clinical trial evaluating the use of non-invasive-ventilation (niv) for treatment of ali in allogeneic transplantation. methods: all patients (n= ) undergoing allogeneic transplantation at a single center were investigated from to . oxygenation-index (pao /fio ) was monitored twice daily from the beginning of the conditioning regimen. patients meeting criteria for ali were randomized to receive either oxygen or intermittent niv with positive end-expiratory pressure. patients not responding to assigned therapy were allowed either to switch from oxygen to niv or proceeded with treatment on intensive care unit (icu). results: of eligible individuals, were randomized to receive oxygen and to niv, patients withdrew consent. oxygen-, niv-and control-( patients without ali) group were well balanced regarding known risk factors for trm. only tbi was significantly more frequent in patients with ali. patients with ali showed significantly shortened short-and long-term overall survival (os) when compared to controls ( -day-os vs. %; median-os vs. months). oxygenation index was not only a major adverse prognostic factor; it also suggested ali long before clinical parameters raised suspicion. of patients randomized to niv only ( %) did not respond and had to be transferred to icu. in contrast of patients ( %) assigned to oxygen did not improve. of these, switched to niv but only patients had to be transferred to icu. thereafter, there was no significant difference between both groups in need for intubation (oxygen: patients, niv: patients). niv did not lead to improved survival in our study ( -day-os niv: %, oxygen: %; median-os niv: months, oxygen: months). survival of intubated patients was poor, as only of intubated patients survived for more than days. conclusion: oxygenation-index is an easily measurable early indicator of ali and poor survival in allogeneic transplantation. niv seems to reduce the need for icu and consecutive intubation in patients not responding to oxygen. we were not able to show significant survival benefits for niv-patients, presumably because patients not responding to oxygen were allowed to switch to niv. what to do when the first allogeneic stem cell transplantation fails? m. kedmi, i.b. resnick, b. gesundeheidt , l. drey, s. samuel, r. or, m.y. shapira* hadassah -hebrew university medical center (jerusalem, il) the failure of allogeneic stem cell transplant (allo-sct) is usually cumbersome, we have retrospectively evaluated our experience in a nd allo-sct. patients: out of allo-transplantees, patients ( males, females, median age . y( m- . y)) underwent or more allo-sct. the indications for the st transplant were acute leukemia ( ), chronic leukemia ( ), lymphoma ( ), other malignancies ( ) and non malignant ( ). the st to nd interval was d to . y (median d). the most frequent indications for the nd sct were basic disease ( ), rejection ( ) and engraftment failure ( ). the nd sct conditioning was radiation based ( )or chemotherapy based in the others and was myeloablative or reduced intensity (ric) in and respectively. in of the scts the original donor was used. nd donor matching was full in transplants (family- , unrelated- ) or mismatched in transplants ( and , family and unrelated donor respectively). graft source for the nd sct was bm ( ) and pbsc ( ). of the grafts were t cell depleted. gvhd prophylaxis was given in nd procedures. the median survival from nd sct was d. despite the low rate of gvhd prophylaxis used only and of the patients developed agvhd and cgvhd respectively. / patients ( %) transplanted survived a year after the nd sct (figure ). trm was % including sepsis, liver and renal failure, neurologic toxicity, rejection and gvhd. factors indicating higher chance for survival were non malignant disease, longer time between procedures (more then y), hla matching and the use of ric (figure ). age at transplantation, the indication for transplantation (relapse vs. others), the development of acute gvhd, radiation based vs. chemotherapy based conditioning, gvhd prophylaxis (either pharmacological or t cell depletion) or graft source were not shown to indicate better or worse prognosis. with a median follow up of . y, patients ( . %) are still alive, out of which are disease free. conclusion: although highly toxic, a nd allo-sct may be beneficial to some patients and lead to long term survival. it seems that the patients whom will gain the most out of nd allo-sct are those who had a longer period between scts, have a matched donor or have non malignant diseases although patients with malignant diseases (some with refractory disease) survived at least a year from the nd procedure while in cr. it is also noteworthy that the use of ric improved outcome. s o treatment of donor graft failure with autologous or allogeneic stem cell boost or a second allogeneic transplantation based on chimerism testing a. shimoni*, n. shem-tov, a. rand, e. ribakovsky, r. yerushalmi, i. hardan, a. nagler chaim sheba medical center (tel-hashomer, il) donor graft failure (gf) is a life-threatening complication of allogeneic stem cell transplantation (sct). we performed this analysis to determine the rate and outcome of gf in the era of modern sct. we retrospectively reviewed data of scts from hla-matched siblings (n= ), matched-unrelated (mud, n= ) or alternative donors (mismatched-related, haplo-identical and cord-blood, n= ) performed in a single institution since / . gf was diagnosed in patients (pts), cumulative incidence (ci) . % ( %ci . - . ). gf was determined when anc had not reached . x * /l by day (primary gf, n= ) or when anc decreased irreversibly after engraftment (secondary gf, n= ). ci of gf was . %, . % and . % after sct from siblings, mud or alternative donors, respectively (p< . ) but was similar following myeloablative or reduced-intensity conditioning ( . % and . %, respectively). pts with a predominant donor population in chimerism testing were given donor cell boost with no additional conditioning (n= ). pts with a predominant host population were given autologous back-up cells (n= ) or a second sct from a different donor (sibling- , haplo- , mud- ) with nonmyeloablative conditioning. pts survived > week after second graft infusion and are evaluable for engraftment. pts engrafted within a median of days (range, - ). the probability and pace of engraftment was similar in the different approaches. pts ( %) were able to be discharged home and died; early after diagnosis of gf with no intervention, within one week of second graft infusion and prior to engraftment, with no engraftment and early after engraftment from infection (n= ), organ failure (n= ) and gvhd (n= ). with a median follow-up of months (range, - ), are alive and additional pts died (relapse- , gvhd- , infection- ). the projected -year survival for all pts was % ( %ci - ). interestingly, pts given autologous cells had donor cell recovery, had spontaneous autologous reconstitution within weeks, died within months (gvhd- , infection- ) with persistent donor cells and remained complete donor until she relapsed years later. in conclusion, treatment of gf with a chimerism directed method can salvage a subset of pts with gf. reserving autologous and/or donor backup cells or an alternative donor is advisable in pts at high-risk of gf. the observation of allogeneic recovery after autologous boost is intriguing and of unknown mechanism. at home autologous stem cell transplantation for haematological malignancies: the role of preparative regimens f. fernández-avilés*, m. rovira, c. martínez, a. gaya, c. gallego, a. hernando, s. segura, l. garcía, j. güell, m. valverde, e. carreras, e. montserrat hospital clínic (barcelona, es) background: the aim of this study was to investigate the impact of the most commonly used regimens on the engraftment, toxicity and readmission rate after autologous stem cell transplantation (asct) in patients managed at home. patients and methods: at home asct (since day + ) was offered to all patients with a good performance status, a travelling time to the hospital of less than minutes, and a caregiver available h per day. in all patients the preparative regimen was administered at the hospital. patients with lymphoma were treated with intensified beac or beam (mg/m ) (bcnu , etoposide , cytarabine , and cyclophosphamide (cy) or melphalan ), patients with myeloma received melphalan mg/m and patients with leukaemia total body irradiation (tbi) gy and cy mg/kg. all patients received the same supportive care, including prophylactic i.v. ceftriaxone once daily. indications for re-admission to the hospital were: patient's or caregiver's desire; uncontrolled nausea, vomiting or diarrhea; mucositis requiring total parenteral nutrition or i.v. morphics; persistent fever; hemodynamic instability, pneumonia, or cardiac and/or respiratory distress. results: seventy-five patients were included in this study. forty-five received beac (n= ) or beam (n= ) (n= , group a), melphalan (group b) and tbi-cy (group c). recovery (days) of granulocyte count above . x /l (group a: ( - ), b: ( - ) and c: ( - )) was significantly faster for patients from group a (a vs. b, p= . ; a vs. c, p= . ). fever occurred in %, % and % of patients in the groups a, b and c, respectively (a vs. b, p= . ). the median (range) days with fever were ( - ), ( - ) and ( ) ( ) ( ) ( ) ( ) for a, b and c groups, respectively (a vs. b, p= . ; a vs. c, p= . and who mucositis grade upper to was observed in %, % and % (a vs. b, p= . ). in group a, ( %) patients needed re-admission by pneumonia (n= ) and persistent fever (n= ), as compared to only ( . %) because of persistent fever in the rest of patients (p= . ). conclusions: despite a faster granulocyte recovery, patients managed at home after beac-beam presented a higher incidence and duration of febrile neutropenia, severe mucositis, and high rate of hospital re-admission, especially when compared with patients receiving melphalan. based on this information we have established a more frequent and strict follow-up of patients having received beac-beam and managed at home. outcome of allogeneic stem cell transplantation in first remission for philadelphia chromosome-positive acute lymphoblastic leukaemia following three schedules of imatinib-based chemotherapy b. wassmann, n. goekbuget, h. pfeifer, d.w. beelen, j. dengler, n. kröger, m. stelljes, k. kolbe, w. bethge, m. bornhäuser, h.-j. kolb, m. lübbert, m. stadler, h. serve, d. hoelzer, o.g background: allogeneic sct in cr is considered the best curative treatment option in philadelphia chromosome-positive acute lymphoblastic leukaemia (ph+all) pts., but relapse and transplant related mortality (trm) remain significant limitations. although imatinib (im) is considered standard element of front-line therapy, the impact of different treatment schedules is not known. objective: to evaluate overall survival (os), trm and relapse risk (rr) in ph+all pts. following allogeneic sct in cr (n= ) after three different im-based chemotherapy regimens. patients: in a prospective, multicenter gmall study, three successive pt. cohorts received im according to one of three schedules: in cohort a (n= ) im was administered alternating with chemotherapy beginning immediately after induction phase ii (ipii)(med. age ( - )yrs.). in cohort b (n= )im was started after completion of induction phase i (ipi) and given concomitantly with chemotherapy throughout ipii, st consolidation (cons.i) and up to yrs.). pts. in cohort c (n= ) received front-line im starting after a day prephase and continued parallel with ip i+ii, cons.i up to yrs.). results: the proportion of pts. transplanted in cr was ( %), ( %) and ( %) in cohorts a, b and c, respectively. median follow-up since allogeneic sct was ( . - ) , ( . - )and ( - ) mo.. median os was mo., mo. and not reached. estimated os at and mo. was %, % and % and % and %, respectively. trm ( / ; %) was mainly due to infections (n= ; %) or gvhd (n= ; %). / ( %) trm deaths occurred in pts. aged > yrs.. trm at and mo. in pts. aged ≤ yrs.(n= ) was % and % compared with % and % in pts. aged > yrs.(n= )(p= . ). rr was significantly lower in pts. with low or undetectable pretransplant bcr-abl levels ( % at mo. vs. % at mo., p= . ). conclusion: all tested schedules enable allogeneic sct in a high proportion of pts.. there was a trend towards superior os in cohort c with no significant difference in trm in the cohorts. age and pretransplant mrd levels had the greatest impact on treatment outcome: trm was significantly higher in pts. > yrs. both low and negative pretransplant bcr-abl levels were predictive of a low probability of relapse, whereas high levels were associated with a % relapse incidence within the st year. the impact of posttransplant im on relapse risk remains to be determined. impact of flt -itd on the outcome of hla-identical stem cell transplant for adult aml with normal cytogenetics: substantial probability of leukaemia-free survival despite increased relapse risk. a retrospective analysis of the ebmt-alwp s. brunet *, m. labopin, a. gratwohl, a. buzyn, j. harousseau, j. jouet, g. socié, a. rambaldi, m. mothy, g. cook, j. sierra, v the prognosis of patients with acute myeloid leukemia (aml) and flt internal tandem duplication (flt -itd) is poor. it is unclear whether this mutation has impact on the outcome of allogeneic stem cell transplantation (allo-sct) for aml in patients with normal cytogenetics (nc). different cooperative groups have obtained controversial results. for this reason, registry data including large numbers of patients are of interest. we analysed the predictive value of flt -itd on relapse incidence (ri), non-relapse mortality (nrm) and leukemia-free survival (lfs) in patients with aml in st cr with nc who underwent a myeloablative allo-sct from hla-identical siblings and were reported to the ebmt. the series included patients, negative itd/flt and ( %) flt /itd positive reported between and with a median follow up time of and months, respectively. no significant differences were observed between the two groups regarding gender, median age ( vs yrs), fab classification, number of induction courses to achieve cr, interval diagnosis to transplant, conditioning regimen, in vitro t-cell depletion, female donor to male recipient and cmv serostatus. in contrast, patients with flt -itd+ had higher leukocyte counts (wbc) at diagnosis ( vs . x /l, p= . ) and more frequently peripheral blood was the source of stem cells ( % vs %, respectively). univariate (table ) and multivariate analyses demonstrated the adverse impact of flt -itd for lfs (hr . , p= . ) and relapse incidence (hr= . , p= . ). it is remarkable, however, % of flt /itd positive patients transplanted for aml in first cr remain alive and disease-free at years. other independent prognostic factors for lfs and relapse were: more than one induction course to achieve first cr. for relapse, higher wbc (hr= . , p= . ) was also a significant prognostic factor. in summary, this analysis shows an adverse impact of flt -itd on transplantation outcome. despite this finding % of flt /itd positive aml patients in first cr and normal karyotipe remain disease-free at years, a proportion that seems higher than with other treatment options. it is becoming clear that a higher lymphocyte count one month after allogeneic stem cell transplantation (sct) is associated with better transplant outcome in patients transplanted from an hla-identical sibling. however, a predictive role of the day post-transplant absolute lymphocyte count (lc ) in unrelated transplants is not defined. we studied the relationship between lymphocyte counts and other engraftment parameters on outcome in patients with myeloid leukemia ( acute myeloid leukemia, chronic myeloid leukemia, and myelodysplastic syndrome) receiving myeloablative sct from an hla-a, -b and -dr matched unrelated donor at karolinska university hospital, stockholm from - . median recipient age was years (range . - ), patients ( %) were under years. conditioning consisted of cyclophosphamide with busulphan (n= ) or total body irradiation (n= ). bone marrow (bm) was given to patients and mobilized unmanipulated peripheral blood stem cells (pbsc) to . median stem cell dose was . x /kg ( . - . ). sixty-three patients ( %) received g-csf post-graft. immunosuppression used post graft was cyclosporine with four doses of methotrexate in patients and other treatments in . overall survival at years was % and relapse-free survival was %. the incidence of agvhd grades ii-iv was % in patients with an lc of < . x /l, % if the lc was . - . x /l and % in patients with an lc > . x /l (p= . ). transplant related mortality (trm) was % in patients with an lc < . x versus % (lc of . - . x ) and % (lc > . x )(p< . ). survival was significantly higher in patients with an lc > . x /l, compared to patients with an lc . - . x /l, and patients with < . x /l ( % vs. %, vs. % p= . and . respectively). when analyzed as a continuous variable in multivariate analysis, a higher lc was associated with a lower incidence of acute gvhd grades ii-iv, improved survival, less relapse and higher relapse-free survival (see figure) . these results indicate that the lc is a robust prognostic factor for transplant outcome in matched unrelated as well as matched related sct for myeloid malignancies receiving either bm or pbsc with or without irradiation conditioning. further research to identify the transplant conditions leading to prompt lymphocyte recovery might lead to global improvements in sct outcome in unrelated sct. s o gitmo survey on the outcome of acute myeloid leukemia patients receiving autologous stem cell transplantation in the old and recent era: final results of the multivariate analysis a. olivieri*, b. bruno, g. meloni, m. falda, a. rambaldi, w. arcese, e. alessandrino, r. scime', r. lemoli, m. cimminiello, a. bacigalupo, a. bosi on behalf of gruppo italiano trapianto midollo osseo (gitmo) gitmo registry data files from acute myeloid leukemia (aml) adults, autotransplanted from to , have been evaluated to assess the outcome according to age, conditioning and stem cell source; patients received one autologous stem cell transplantation (asct); two or more asct; allogeneic transplant after failure of asct. patients were categorized in cohorts basing on the transplant era (before or after ); the two cohorts were significantly different as regard: age distribution ( % pts older > yrs transplanted after vs % pts > yrs before ); less pts < yrs received asct after ( % versus %); preparative regimens: tbi or bu-cy were less common in the recent era; status of disease: % received asct in rst or nd cr before , compared to % after ; stem cell source: more pts ( %) received pbsc after versus % before . with a minimum yrs followup, os did not differ in the two cohorts: % in the patients autotransplanted after vs % in the autotransplanted before . as a preliminary analysis suggested a possible role of pbsc in reducing the overall nrm after (from % to % after ), we made a multivariate analysis to evaluate the main variables (age, stem cell source, transplant era, disease status at transplant and type of conditioning) influencing the outcome, in terms of os, nrm, dfs and relapse incidence. age> yrs significantly worsened nrm ( , hr), os ( , ) and dfs ( , hr) regardless the transplant era; asct performed in advanced disease (> nd cr) was associated with a , times increased risk of relapse and nrm. stem cell source did not significantly influence os and nrm, but bm source, instead of pbsc, was associated with significantly reduced risk ( , hr; p= . ) of relapse. finally tbi regimens were associated with increased nrm compared to bu-cy ( , hr). some conclusions can be drawn from this survey: -the advanced age (> yrs) remains an adverse factor both for nrm, os and for lfs; -these data definitely show that pbsc are associated with increased risk of relapse after asct, without major impact on os; -regimens including tbi are not recommended being associated with an increased nrm without a significant reduction of relapse. the main efforts in the future should be aimed to reduce the relapse incidence probably by designing new conditioning regimens (and by targeting the minimal residual disease post-transplant) and by a cautious use of pbsc in patients receiving a short consolidation. up-front allogeneic stem cell transplantation as part of induction therapy in newly-diagnosed high-risk acute myeloid leukaemia -an update of a prospective phase ii trial u. platzbecker*, m. füssel, m. schaich, t. illmer, b. mohr, j. schetelig, a. kiani, c. theuser, c. thiede, g. ehninger, m. bornhäuser university hospital (dresden, de) poor-risk cytogenetic aberrations, bad response to the first cycle of induction chemotherapy (ic) or the presence of an flt- receptor mutation define high-risk aml (hr-aml) and result in an increased risk of failure of long-term disease control. as a matter of fact, only a minority of this patient group proceed to hsct due to treatment failure or death from infectious complications during ic. therefore, there is substantial need to improve the outcome of these patients with hr-aml. we report results of an ongoing prospective trial evaluating an "early" hsct applied during induction-chemotherapy induced aplasia in forty (n= ) newly-diagnosed hr-aml patients with a median age of years ( - ). a median of days (range - ) after the first (n= ) or second (n= ) cycle of ic patients received a reduced-intensity regimen that was based on fludarabine combined with either busulfan (n= ) or melphalan (n= ) followed by allogeneic g-csf mobilized peripheral blood stem cells (pbsc) from related (n= ) or unrelated (n= ) donors. twenty-six patients were not in complete remission before conditioning therapy was started with a median marrow blast count of % (range - ). patients with unrelated grafts received antithymocyte globulin and gvhd prophylaxis was performed with cyclosporine a only in all patients. all patients engrafted and went into remission. acute gvhd grade ii-iv occurred in % and extensive chronic gvhd in % of patients. with a median follow-up of months (range - ) the probability of overall and disease-free survival at months is %. early allogeneic hsct as part of primary induction therapy seems to be an effective strategy in high-risk aml patients. comparison of up-front versus minimal residual disease triggered imatinib after stem cell transplantation for philadelphia chromosome-positive acute lymphoblastic leukaemia: interim results of a randomised phase iii study b. wassmann*, h. pfeifer, w. bethge, m. bornhäuser, j. dengler, m. stadler, d. beelen, n. basara, r. schwerdtfeger, k. schäfer-eckart, l. uharek, h. serve, d. hoelzer, o.g. ottmann on behalf of the gmall study group background: detection of minimal residual disease (mrd) following allogeneic sct for philadelphia chromosomepositive acute lymphoblastic leukemia (ph+all) is highly predictive of evolving relapse. we have previously shown that only about % of pts. with ph+all who convert to mrd positivity after allogeneic sct achieve renewed pcr negativity in response to interventional imatinib (im), started a median of mo. after sct. failure to rapidly achieve a complete molecular response was almost invariably associated with hematologic relapse. we hypothesized that earlier initiation of im in the setting of a lower leukemic cell burden would increase response rates and improve dfs. objective: to determine whether the earliest possible initiation of im after sct is superior to delayed im triggered by reappearance of bcr-abl transcripts with respect to feasibility, tolerability and duration of molecular and hematologic remission. patients: six wks. after allogeneic sct pts. were randomly assigned to receive im either up-front (cohort )(n= ) or subsequent to detection of bcr-abl positivity as determined by real time quantitative and/or nested rtpcr (cohort )(n= ). target dose of im was mg, but a lower dose of mg was permitted if deemed necessary because of tolerability. im was scheduled for a total duration of one year of pcr negativity. results: to date, patients have been enrolled. of the pts. randomized to each cohort, each underwent sct in first complete remission (cr ), each in cr . im was started in / pts. in the up-front im and / in the mrd-triggered cohort, with most pts. receiving mg im ( / pts. and / pts., respectively). med. time from sct to start of im was (cohort a) and days (cohort b). after a med. follow-up of ( - ) and ( - ) days in cohorts a and b, respectively, none of the pts. transplanted in cr and of with sct in cr relapsed. three pts. (cohort a) died in cr, only one of whom had actually received im. im was discontinued prematurely in / pts. in the im up-front arm and / in the mrd-triggered im cohort, due mostly to gastrointestinal toxicity and gvhd. conclusions: with rigorous monitoring of mrd, both schedules of post-sct im (up-front versus mrd-triggered) are associated with a remarkably low relapse rate, although follow-up is still short. tolerability of im is poorer than generally experienced in non-transplanted patients. the routine use of im after sct is a promising strategy to improve outcome of pts. with ph+all. we compared reduced intensity conditioning (ric, n= ) with myeloablative conditioning (mac, n= ) in patients with aml undergoing hematopoietic stem cell transplantation (hsct), using hla-a, -b and -drb identical unrelated donors, transplanted between to and reported to the ebmt. in the ric group, median age was higher, transplant was performed more recently, time from diagnosis to transplant was longer, t-cell depletion was less commonly used, and more patients received pbsc vs. bone marrow (p< . ). in patients below years of age in cr , transplant-related mortality (trm), relapse and leukemia-free survival (lfs) were similar in the two groups. in patients in cr - below years of age, -year probability of relapse was % in the mac group, compared to % in the ric patients (p= . ). in these patients, trm was % vs. %, and lfs was % vs. % at two years in the two groups, respectively (ns). in patients with aml advanced disease below years of age, trm, relapse and lfs did not differ significantly between ric and mac patients. at two years, lfs was % with mac vs. % using ric (p= . ). in patients above years of age in cr and advanced disease, relapse was not statistically different in the two groups. in cr - , mac patients above years had a probability of relapse of % vs. % for ric patients (p= . ). there were no statistically significant differences in trm or lfs, using mac or ric in patients above years. to conclude, using ric as an alternative to mac in patients with aml undergoing transplants with matched unrelated donors resulted in: no significant difference in trm in patients above and below years of age, significantly increased risk of relapse in patients treated with ric in cr - . lfs was similar using the two conditioning regimens. higher incidence of relapse with peripheral blood as source of stem cells in adult patients with acute myelocytic leukaemia autografted in first remission n.c. gorin ( ) in the past years the modalities of autologous stem cell transplantation (asct) in first remission (cr ) for patients with acute myelocytic leukaemia (aml) have evolved: the age limit for asct has been gradually extended up to years . total body irradiation (tbi) pretransplant has declined in favour of chemotherapy (ct). leukapheresis products (pb) have replaced bone marrow (bm) as source of stem cells. we were interested in evaluating the potential impact of these modifications on the outcome post asct. a total of asct ( bm, pb) were reported from january to december . % of all pb transplants were done after . therefore, we compared bm versus pb in patients patients transplanted after : patients received bm and pb. the median follow up in the two groups were and ( - ) months respectively. patients receiving pb were older ( y vs ,p< . ), had more aml of the m , , categories (p< . ) and received less tbi ( % vs %, p< . ). by multivariate analyses, age was a significant factor with lower trm, higher relapse incidence (ri) and lower lfs above years. failure to reach cr within days was associated with a higher (ri) and a lower lfs. the use of pb as compared to bm significantly resulted in a higher ri ( ± % vs ± %, p= . ), and a lower lfs ( ± % vs ± %,p= . ) with only a trend for a slight reduction in trm ( ± % vs ± %, p= . ). tbi was not a significant factor for outcome. we finally focused on good risk patients (cr within the first days): the population consisted of patients grafted with pb and with bm. patients receiving pb were older (p< . ), and received less tbi ( p< . ). again in this good risk population the ri was significantly higher with pb ( ± % vs ± %, p< . ), with a trend for a worse lfs ( ± % vs ± %,p= . ). we conclude that the shift to pb as a source of stem cells in the past years has resulted in increasing the relapse incidence possibly through mobilisation of leukemic cells and /or insufficient purging of the autograft. s experimental stem cell transplantation/ stem cell research o co-transplantation of placental derived mesenchymal stromal cells produces superior engraftment of umbilical cord blood compared to double unit umbilical cord blood transplantation s. hiwase, p. dyson, s. young, b. to, i. lewis* imvs (adelaide, au) double-unit umbilical cord blood transplantation (ucbt) has been shown to overcome some of the limitations of ucbt, particularly in adult recipients. whether this simply reflects a cell dose effect has not been established. co-transplantation of mesenchymal stromal cells (mscs) has also been suggested as a means of enhancing engraftment and may be appropriate in patients where two suitably matched cords cannot be identified. in this study we have directly compared engraftment rates of double-unit ucbt with msc cotransplantation. mscs were obtained from placental tissue by enzymatic digestion and isolated by plastic adherence. placental mscs demonstrated fibroblastic morphology, immunophenotype and differentiation potential similar to bone marrow derived mscs. in a nod/scid mouse model groups of mice were compared: group received x cd + cells from a single cord unit (u ); group received x cd + cells from u + x mscs; group received . x cd + cells from u + . x cd + cells from u ; group received . x cd + cells from u + . x cd + cells from u + x mscs. in independent experiments mean engraftment rates were: group %, group %, group %, and group %. hence msc co-transplantation produced superior engraftment when compared with either single unit (p= . ) or double unit transplantation (p= . ). combining results demonstrated the superiority of msc co-transplantation with mice receiving mscs showing mean engraftment of % compared to mice who did not receive mscs having engraftment of % (p= . ). transplantation of double ucbt did not improve engraftment when compared with a single ucbt of equivalent dose. additionally, the quality of engraftment was enhanced with msc co-transplantation producing superior engraftment of cd + cells. in conclusion, at equivalent cell dose single and double ucbt lead to similar engraftment, suggesting the enhanced engraftment seen with double ucbt reflects a cell dose effect. msc co-transplantation enhances engraftment of both single and double unit cords and may be a potential strategy to be explored in the clinic. infusion of allogeneic mesenchymal stromal cells can delay but not prevent gvhd after murine transplantation m. kambouris *, b. turner, l. sinfield, h. cullup, d. hart, k. atkinson, a. rice mater medical research institute (south brisbane, au) multipotent, mesenchymal stromal cells (msc) are emerging as a means of immunosuppression for patients with steroid refractory graft-versus-host disease (gvhd). despite clinical use, pre-clinical data is still lacking. we established an in vivo model using msc to control gvhd to determine their mode of immunosuppression. we showed in a mixed lymphocyte reaction that msc are highly immunosuppressive and significantly reduce t cell proliferation. we then examined the effects of donor-derived intraperitoneally (ip) or intravenously (iv) injected msc on gvhd in a myeloablative conditioned, full mismatched model of haematopoietic stem cell transplantation (hsct) [ubi-gfp/bl (h- b)->balb/c (h- d)]. x donor-derived msc/mouse were injected hrs pre or hrs post hsct then mice were monitored daily for gvhd. only mice given msc ip hrs post hsct showed a signficant delay in death from gvhd, where median survival was increased by days (day vs day , p< . , (fig ) . we then investigated if msc delivered ip pre or post hsct altered cytokine-driven trafficking of cellular effectors known to play a role in gvhd via timed sacrifice. control mice (hsct and no msc or msc and no hsct) were also sacrificed daily for days following hsct. we found that msc given post-hsct enter an environment of significantly increased activated dendritic cells (dc) in the spleen ( fig ) and at day and day post hsct, mice given msc pre hsct had increased levels of activated splenic dc compared to mice treated with msc hrs later. at day , we also saw more ifn-gamma in spleen washings in mice treated pre hsct compared to controls. we then determined the role of msc in gvhd control in a minor mismatch model [ubi-gfp/bl (h- b)->balb.b (h- b)]. msc were given on day , or ; or to mice with established gvhd (> % weight loss with other surrogate gvhd markers more than days post hsct). mice given msc therapy after gvhd onset showed no increase in survival compared to controls. however, msc administered prophylactically at day ( x /mouse) or day ( x /mouse) but not at day , showed significantly increased survival compared to controls suggesting that the timing of msc administration is important for their impact on gvhd. in summary, ip injection of msc influences gvhd and survival and may have in vivo influence on activated dc. elucidation of the mechanism by which msc control gvhd may ultimately lead to wider application of their use to assist hsct. s o immune reconstitution after cord blood transplantation by direct intrabone injection of cells a.m. raiola*, a. ibatici, v. pinto, a. kunkl, f. guialn, f. gualandi, d. occhini, a. dominietto, c. di grazia, s. bregante, t. lamparelli, m. mikulska , g. piaggio, m. podestà, f. frassoni, m. van lint, a. bacigalupo s. martino's hospital (genoa, it) introduction: cord blood transplantation (cbt) has been increasingly used to treat hematological malignancies. since march we have been investigating a pilot study of cbt in adults by injecting cb cells directly into the bone to overcome the cell dose barrier. pre-clinical transplant models have shown that intrabone (ibm) injection reduces the incidence of gvhd in mhc disparate donor/recipient pairs, suggesting that the way of transplant may affect the immune reconstitution. we have analysed immune reconstitution (t/b/nk cells) at different time-point after ibm-cbt in adults recipients. materials & methods: thirty-three consecutive patients with advanced haematological malignancies a single-unit graft cbt. median age was years ( - ) and follow-up was months ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) for patients survivors longer than days after cbt. median cell dose was . tnc/kg (range, . - ). donor/recipient hla matching was highly disparate: / in %, / in %, / in %. all but pts received a myeloablative conditioning regimen (tbi cgy in / pts). gvhd prophylaxis consisted of csa, mmf and pretransplant atg. immunophenotype on peripheral-blood lymphocyte subsets was assessed on day + , + + , months and yr after cbt. results: early death occurred in pts. all were evaluable and engrafted % donor. overall survival was % and relapse-related death was % ( pts). median time to neutrophil and platelet recovery was and days, respectively. grade ii acute gvhd was seen in cases ( %) and grade iii in case ( %). moderate/extensive chronic gvhd developed in / ( %). on day + , median cd + and cd + count were as low as and cells/mmc, respectively and remained below normal values up to months. at year, cd + and cd + count normalized ( and cells/mmc). nk values reached values as high as and cells/mmc on day + and + , then remained stable up to yr. b cells were extremely low up to day+ , peaked to on day + and slighty expanded over time. conclusion: data from our series showed that immune reconstitution is as delayed as in patient who received cb grafts intravenously, with prolonged t-cell lymphopenia and bcells spike since day+ (komanduri, blood ). we also confirmed early nk expansion which could be due to the high donor/recipient pair hla disparity associated with the low cd + count. if this may have an impact on the relatively low relapse incidence observed in advanced patients warrants further studies. the effect of mesenchymal stem cells in a rat model of experimental arthritis is dose-and time-dependent e. yannaki ( ) , a. papadopoulou* ( ), m. yiangou ( ) , e. athanasiou ( ) , i. batsis ( ) , a. athanasiadou ( ) , a. xagorari ( ) , a. anagnostopoulos ( ) , a. fassas ( ) ( )george papanicolaou hospital (thessaloniki, gr); ( )aristotle university (thessaloniki, gr) mesenchymal stem cells (mscs) have recently generated a great deal of excitement, due to their potential of differentiation into a broad spectrum of tissues and immune regulation by non-mhc-restriction. the latter capacity has resulted in an effective control of gvhd in several studies; however, the role of mscs in autoimmune diseases has not been extensively investigated in animal models. we aimed to explore the effect of rat mscs on cultured fibrobiast-like synoviocytes (fls) and t-cells from the spleen after adjuvant arthritis induction (aa) as well as their in vivo immunomodulatory potential in a rat model of aa resembling human rheumatoid arthritis. culture of aa-fls in the presence of supernatant from syngeneic msc culture when compared to fls in the absence of supernatant, reduced the aa-fls proliferation (p= . ) as well as the proliferation of cona-stimulated aa t-cells (p= . ). coculture of activated t-cells with syngeneic mscs produced stronger inhibition (p= . ) in a dose-dependent manner. the inhibitory effect of allogeneic mscs in activated aa t-cells was even stronger and similarly dose-dependent, either by secreted agents (p= . ) or by cell to cell contact (p= . ). low doses of mscs ( . - x cell/recipient) administered iv, intrasplenic or intrabone marrow, at single or multiple infusions, didn't produce significant differences in disease scores of msctreated as compared to control rats. on the other hand, repeated, higher dose ( x cell/rat), iv infusions of syngeneic or allogeneic y+mscs to female recipients, before the onset of aa (d and d post aa), resulted in significantly lower arthritic scores with maintenance of a rather normal joint architecture with mild focal synovial hyperplasia/pannus formation and reduced abnormal chondropiasia or bone erosion as compared to control rats. in contrast, symptom worsening occurred when the same cell dose was injected after arthritis onset (d and d post aa). at the time of sacrifice (d post aa), no y+mscs were detected in the spleen or in cultured fls from synovial membrane, by pcr or fish, suggesting that the observed benefit was not due to mscs homing to the target tissues or that migration of mscs may have happened earlier. our data suggest mscs as a potential new therapeutic approach for autoimmune arthritis. however, the benefit of infused mscs seems to be dose-and time-dependent and further studies are required before the results can be clinically translated. objectives: mesenchymal stem cells (msc) suppress alloantigen-induced proliferation, interferon-gamma (ifngamma) production and cytolytic killing in vitro and infusion of third-party msc appears a promising therapy for acute gvhd. however, little is known about the specificity of immunosuppression by msc and in particular the effect on cell-mediated immunity to infectious pathogens. we have studied the effect of msc on virus-specific t-cell responses. results: peripheral blood mononuclear cells (pbmc) from normal donors were stimulated for days with autologous lymphoblastoid cell lines (lcl) (ebv), pp peptides (cmv), an adenoviral vector ad f (ad) or allogeneic pbmc (allo), in the presence or absence of third-party msc (msc/pbmc ratio : ). msc significantly suppressed proliferation in response to allo (mean % suppression, p= . ), but had less effect on the response to ebv (mean % suppression, p= . ) and no suppression of the response to cmv or ad. msc had no effect on expansion of ebv and cmv pentamerspecific t cells after stimulation with their cognate antigen. elispot assays demonstrated that msc significantly inhibited ifn-gamma production in response to allo (mean sfc/ cells ± without msc, ± with msc) and to a smaller extent to ebv ( ± vs ± ), but not to cmv ( ± vs ± ). established ebvspecific cytotoxic t-cells (ebv-ctl) from donors were stimulated with autologous lcl with or without msc for days. the percentage of ebv-pentamer specific ctl was not affected by the presence of msc (mean . % vs . %). msc did not inhibit ifn-gamma production by ebv-ctl ( ± vs ± ). furthermore, cytolytic killing of lcl by ebv-ctl was not suppressed by msc (mean specific lysis ± . % vs ± . % at e:t ratio : ). finally, we studied anti-cmv immunity in patiens who received msc for acute gvhd of the gut evolving into chronic gvhd, with a good transient response. pbmc from these patients showed persistence of pp -pentamer positive t-cells and retained ifn-gamma response to cmv post msc infusion (sfc / pbmc pre-msc, at month and at months). conclusion: msc have little effect on t-cell responses to cmv, ebv and ad, which contrasts to their strong immunosuppressive effects on alloreactive t cells. these data have major implications for immunotherapy of gvhd with msc and suggest that the effector functions of virus-specific t-cells may be retained after msc infusion. intracerebral transplantation of human mesenchymal stem cells after hypoxic-ischaemic brain damage in rat h. huang* ( ), j. fan ( ), x. lai ( ) , q. yu ( ) , w. ding ( ) , y. wang ( ) ( )zhejiang university school of medicine (hangzhou, cn); ( )zhejiang chinese medical university (hangzhou, cn) mesenchymals stem cells (mscs) have dramatic potential of proliferation and multiple differentiation, and recently, neural transdifferentiation of mscs provoke extensive interest. our study was designed to explore the migration, differentiation and the therapeutic benefit of hmscs after hypoxic-ischemic brain damage in rats. hmscs were prelabeled with bromodeoxyuridine (brdu) for h before transplantation. animal models of hypoxic-ischemic brain damage (hibd) were built in one month old wistar rats, and three days after hypoxia-ischemia, hibd rats in hmscs-treated group (n= ) received intracerebral transplantation of ± hmscs, while hibd rats received pbs of the same volume in control group (n= ). in sham-operated group (n= ) and hibd group (n= ) which just had hypoxic-ischemic brain damage but did not receive any treatment. all of the groups did not receive any immunosuppression agents. behavior tests (alternative electro-stimulus y-maze, by single blind method) indicated that pace memory capacity of rats in hmscs-treated group is significantly better at weeks post-transplantation. he straining also showed that the damage caused by hypoxicischemic had lager pathological changes with bulks of neural cells necrosis and neuropil cavitations formation, even disappearance of normal architecture in cortex and hippocampus, however, it would be significantly improved in pathology in hmscs-treated group. hmscs stayed around the local area of transplantation in three days after transplantation, then migrated mainly along the ventricular system. four weeks later, hmscs were observed to migrate to the parenchyma and distribute throughout the cerebra. three days after transplantation, some hmscs expressed gfap in the local area of transplantation, and a few of cells expressed neurofilament (nf) near blood vessel. four weeks after transplantation, brdu and gfap co-express cells increased which mainly distributed in the local area of transplantation, the cortex, hippocampus and ependymal layer. nestin positive cells did not be detected at any time point. conclusion: after being transplanted intracerebrally, hmscs could migrate to the lesion area, express nf and gfap which indicated their differentiation into mature neurons and neurogliocytes, and promote tissue repair and functional recovery of hibd. furthermore, these results suggested that hmscs could be a promising treatment for hypoxic-ischemic brain damage. identification and enrichment of human bone marrow derived mesenchymal stem cells by monoclonal antibody, zuc x. lai, h. huang*, l. huang, j. cao, f. zeng, j. zheng zhejiang uninversity school of medicine (hangzhou, cn) human mesenchymal stem cells (mscs) could be well isolated and expanded from bone marrow and have been widely studied, however, there is no specifically definitive marker of mscs till now. in our previous study, a novel murine monoclonal antibody (mcab) zuc was produced by hybridoma technology, which was specifically reactive with human mscs, while showed negative cross-reactivity when screened against a variety of human tissues. flow cytometric analysis showed that zuc antigen expression by cultured mscs and mononuclear cells derived from bone marrow were . ± . % and . ± . % respectively, and western blotting demonstrated the molecular mass of antigen was about kd. zuc antigen positive and negative cells were separated from bone marrow mononuclear cells by immunomagnetic activated cell sorting, and plated respectively in human mscs medium consisting of % fbs, lg-dmem. the purity of the recovered fractions for zuc by macs was . ± . % by flow cytometry. the positive cells have adhered to culture flask in vitro, and the quantity of adhered cells that had fibroblast-like morphology increased and proliferated during primary expansion period, while the negative cells were observed as round shape cells without any proliferation. it was demonstrated that zuc antigen positive cells continued growth with spindle-shape, expansion in long-term culture. phenotype of zuc antigen positive cells was analyzed by flow cytometry, the culture-expanded positive cells were uniformly positive for cd , cd , cd , cd , and lack typical hematopoietic antigens such as cd , cd , cd , hla-dr, which demonstrated that zuc positive cells sorted from bone marrow mononuclear cells were mscs. with proper medium, the zucs antigen positive cells could be successfully induced to differentiate into adipocytes, osteoblasts, and neuro-like cells which were positive of neuron markers such as nestin, nse and nf-m. conclusion: zuc mcab was a specific surface marker against human mscs for cell sorting. the zuc antigen positive cells separated from bone marrow mononuclear cells had potential capacity of high proliferation and multiple differentiation. everolimus enhances the immunomodulatory properties of cd positive selected human mesenchymal stem cells r. racila*, w. melchinger, j. finke, r. marks medical hospital university of freiburg (freiburg, de) immunomodulatory properties of human mesenchymal stem cells (msc) suggest their use as a potent cellular therapeutic agent for acute graft versus host disease in allogeneic hematopoietic stem cell transplantation (hct). everolimus is a mtor inhibitor which is currently being tested for gvhd prophylaxis, but its effect on mscs is not yet understood. since mtor inhibition by everolimus might interfere with protein biosynthesis, we tested whether mscs maintain their immunomodulatory properties in the presence of everolimus. in order to obtain a homogenous msc population, positive selection of cd + cells from bone marrow aspirates from volunteers using anti-cd antibody coated magnetic beads were used. the one-step procedure resulted in an enrichment of cd highcd +cd lowcd -msc with copurification of cd lowcd -cd high hematopoietic cells. expansion culture resulted in a pure cd lowcd lowcd highcd highcd low msc population. we assessed the immunomodulatory properties of cd lowcd lowcd highcd highcd low mscs when cocultured with autologues and allogeneic t cells stimulated with anti-cd /cd antibodies or third party peripheral mononuclear cells. the presence of msc in allogeneic t cell cultures did not result in a proliferative t cell response. moreover, stimulation of allogeneic msc/t cell cultures with anti-cd /cd antibodies resulted in suppression of t cell proliferation as determined by thymidine incorporation. a - % reduction of maximal t cell proliferation was seen when a ratio of : (msc to t cell) was used. in order to examine the effect of mtor inhibition onto the immunomodulatory properties of msc, we incubated the msc for hours with nm everolimus. after several washes the everolimus treated and irradiated msc were used for coculture (ratio : ) with stimulated allogeneic t cells. interestingly, everolimus treated mscs showed enhanced suppressive properties and a - % reduction of maximal t cell proliferation. we conclude that everolimus enhances the immunomodulatory properties of cd positive selected human mscs and the clinical use of this mtor inhibitor might result in significant immunsupressive activity in allogeneic hct. fibrosis regression after autologous haematopoietic stem cell transplantation in systemic sclerosis l. vija, f. verrecchia, o. verola, a. de raignac, d. sibon, l. michel , d. farge st louis hospital (paris, fr) background: significant regression of clinical skin sclerosis, as assessed by repeated measure of the rodnan modified skin sclore (mrss), has been shown after autologous stem cell transplantation (abmt) in diffuse scleroderma (ssc) patients. objectives: to analyse wether ) the clinical and histological extent of skin fibrosis can regress after hsct and ) extra cellular matrix organization and pro-fibrotic signals elicited by tgf-b in ssc fibroblasts derived from skin biopsies vary according to the modified rodnan skin score (mrss) before and after hsct . methods: ssc patients underwent skin biopsies with simultaneous measure of mrss before or after treatment by immunosuppressive drugs with or without autologous hsct. the histological presence and distribution of sclerosis was assessed as : = no fibrosis, + = light fibrosis, ++ = moderate fibrosis, +++ = extensive fibrosis within the papillary (pd), the superficial reticular dermis (srd), the median reticular dermis (mrd) and the deep reticular dermis (drd). human fibroblasts were established by explanting skin punch biopsy (ssc patients or healthy controls) in dulbecco's modified eagle's medium and cells were used for experiments between passages and . immunoblotting analyses with anti-phospho smad (mayo clinic, rochester, mn), anti-smad (clinisciences, ca), and anti-bactin antibodies (santa cruz biotech, ca) and rt pcr for col a , col a and pai- have been performed. results: double blind optic microscopy analysis of the biopsies seriate sections on standard matrix stains allowed to define histological subgroups : with grade weak fibrosis, with grade moderate fibrosis and with grade severe fibrosis with significant correlation (p< . ) between the grades of fibrosis and the mrss. in skin fibroblast cultures, smad phosphorylation levels, representative of tgf-b receptor activity, increased in parallel with the mrss. when compared to pretransplant values, a significant regression of the degree of fibrosis was observed both in the papillary and in the reticular dermis after hsct (n= ), correlated with a decreased mrss. conclusion: in ssc patients, we confirm that the histological extent of skin fibrosis correlates closely with the mrss and further demonstrate that both parameters regress after treatment by hsct. the extent of tgf-b signaling activation in ssc skin fibroblasts appears to parallel the severity of disease. we report here an update of the phase multicenter, prospective study of high-dose immunosuppressive therapy and autologous haematopoietic cell transplantation in nonprimary progressive ms patients showing high disease activity on the basis of both brain magnetic resonance imaging (mri) and sustained clinical deterioration despite conventional therapies. the treatment consisted of stem cell mobilization with cyclophosphamide ( g/m²) and filgrastim, and conditioning with bcnu ( , -bis( -chloroethyl)- -nitrosourea), cytosine arabinoside, etoposide, and melphalan (beam) followed by antithymocyte globulin (atg, mg/kg) and infusion of unmanipulated peripheral blood stem cells (pbscs). the median follow-up is now months (range - ). confirmed progression-free survival was evaluated according to edss disability score as the probability of being alive without clinical progression as compared to baseline. progression-free survival is , % at , years after hsct (figure ). seven patients achieved a sustained improvement in their edss score, while patients remained stable and worsened of at least , points in edss score. disease progression was not associated to clinical relapse. only one patient showed a clinical relapse at + months, spontaneously remitted without deterioration of the edss score.since transplantation patients were treated only with symptomatic therapy. hsct was capable to induce sustained progression and treatment-free survival in a large cohort of rapidly deteriorating ms patient with a modest transplantrelated toxicity. experimental animal data and single case reports have documented that allogeneic haematopoetic stem cell transplantation (hsct) potentially alters the course of severe autoimmune disease (aid). experience with allogeneic hsct for these indications is still limited. we therefore undertook a retrospective analysis of the ebmt database to identify patients, which received allogeneic hsct for aid. we identified patients (pts), who underwent allogeneic hsct for aid between and . follow up was requested by a questionnaire, sent to the referring physicians, asking for diagnosis, transplant procedure, complications, disease status and last follow up. the completed questionnaire was available at the time of abstract submission for pts. median age at transplantation for all pts was . ( . - . ) years; ( male/ female/ not known). sixteen hsct were performed for non-haematological aid and for haematological aid. three pts had two hsct for aid. eight pts had a previous autologous hsct. mean follow up time was . months (iqr . - . ) . responses were complete for pts ( %), partial for ( %), no change for ( %) and unknown for pts ( %). complete or partial response rate was independent of underlying disease; (haematological vs. non-haematological aid) (p= . ), gender (p= . ) or age (p= . ). cyclophosphamide based conditioning was associated with a better response rate (p= , ), while atg had no influence on the response. overall mortality was %. three patients ( %) died of progression of aid, eight patients ( %) died of treatment related reasons. the median followup time of non-survivors ( pts) was . months (range . - . ) and of survivors ( pts) . months (range . - . ) . kaplan maier curves displayed similar survival rates for haematological and non-haematological aid. best survival was observed for twin donors ( pts all survived) and matched donors (related or unrelated). tbi conditioning showed a trend to a higher mortality (p= . ). in conclusion allogeneic hsct has the potential to induce complete remission in refractory aid. the high mortality rate limits its general use. for selected patients allogeneic hsct based on cyclophosphamide conditioning from a matched donor could be an option and should therefore be prospectively evaluated. patients with intractable form of ms were included in the phase ii clinical trial involving the high dose chemotherapy with autologous peripheral blood progenitor cell (pbpc) support between and . patients underwent high dose conditioning beam. t cell depletion in vitro was performed on grafts depending on number of harvested progenitors and available resources. patients with not purged graft received atg mg/kg i.v. d+ , d+ after pbpct. in patients pbpc mobilization failed (cyclophosphamide g/m + g-csf). one patient refused transplantation after improvement in disability following mobilization. median follow-up is months ( - ). median edss (expanded disability status scale) of grafted patients at the time of inclusion was . ( . - . ), median edss of grafted patients at last follow up was , ( . - . ). two patients died and months resp. after transplantation because of progression of ms and the cause not related to transplantation, respectively. two patients were lost for follow up. there was no treatment related mortality. at last follow up, the significant improvement (by . point and more on edss) remains in patient, stabilisation of the disease occured in patients ( %), patients gained disability significantly (by . point and more on edss). in patients occured transient significant neurological improvement lasting - months. the development of disability between the group grafted with in vitro purged graft and the group with atg i.v. was not significant. cumulative survival without significant deterioration was % in the group with purged grafts and , % in the group without purging in vitro (p= , ). among mobilized but not transplanted patients improved by . point on edss, patients worsened by . point. three serious adverse events related to transplantation were observed: respiratory failure after the onset of mucositis in the patient with severe pontomesncephalic impairment, sepsis with respiratory failure following bilateral pulmonary hemorrhage, both patients needed temporary artificial ventilation and recovered. bleeding related to the inhibitor of fviii occured in patient, remmission has been achieved after the immunosuppressive therapy. as majority of patients with otherwise intractable ms at least stabilized in their disability, we consider the results to be promising and requiring confirmation in a randomized trial involving also less handicapped patients. long-term results of a gitmo retrospective study on haematopoietic stem cell transplantation for paroxysmal nocturnal haemoglobinuria s. santarone*, e. di bartolomeo, a. bacigalupo, e. tagliaferri, a. iori, a. risitano, s. tamiazzo, f. papineschi, a. rambaldi, a. spagnoli, e. angelucci, p. di bartolomeo on behalf of the gitmo allogeneic haematopoietic stem cell transplantation (hsct) may cure paroxysmal nocturnal hemoglobinuria (pnh). in this study we report the results of allogeneic hsct in patients ( males and females) affected by pnh who were transplanted between july and may . the median age at time of hsct was years ( - ). the median time from diagnosis to hsct was months ( - ). all patients had received various treatments before hsct including steroids, immunosuppressive drugs and growth factors. twenty-one patients were transfusion-dependent. the median number of packed red blood cells and platelet concentrates received before hsct was ( - ) and ( - ) respectively. the median peripheral hematological counts at transplant were: polymorphonucleates (pmn) ( - ) x /l, hemoglobin , g/dl ( . - ), platelets (plt) ( - ) x /l. thirteen patients developed thromboembolic episodes before hsct. twenty-four patients were transplanted from hla identical sibling and from matched unrelated donors. the donor's median age was years ( - ). the conditioning regimen was myeloablative for patients (busulfan and cyclophosphamide), whereas patients received a reduced intensity conditioning including fludarabine, cyclophosphamide, melphalan and total body irradiation. as graft-versus-host disease (gvhd) prophylaxis, patients received cyclosporine (csa) alone and were given csa and short course methotrexate. two patients received t-cell depleted marrow cells. twenty patients were given bone marrow cells (median nucleated cells . ( , - , ) x /kg) and received peripheral blood stem cells (median cd + cells . ( . - . ) x /kg). twenty-five patients engrafted with a median time of ( - ) days to reach > . x /l pmn and days to reach > x /l plt. the probability of developing grade ii-iv acute gvhd and extensive chronic gvhd was % and % respectively. the transplant related mortality at months was %. causes of death were infection in patients, acute gvhd in , chronic gvhd in , multi-organ failure in and ebv-related disease in . as of december , patients are alive with complete hematological recovery and no evidence of pnh at a median follow-up of months ( - ). the -year kaplan-meier probability of disease-free survival for the patients transplanted from related donor is %. no patient developed thromboembolic disease following hsct. this study confirms that hsct is a curative treatment for the majority of patients with pnh. subcutaneous alemtuzumab is safe and effective for treatment of global or single-lineage immune-mediated marrow failure: a pilot study from the working party aplastic anaemia (wpsaa) a. risitano*, l. marando, c. selleri, b. serio, e. seneca, a. camera, l. catalano, g. scalia, l. del vecchio, a. iori, s. maury, a. bacigalupo, g. sociè, a. tichelli, j. marsh, h. schrezenmeier, j. passweg, b . rotoli on behalf of the wpsaa acquired marrow failure syndromes may globally affect all hematopoietic lineages, as in aplastic anemia (aa), or may selectively involve single lineages, as in pure red cell aplasia (prca) or in agranulocytosis (agr). standard treatment for these conditions includes immunosuppression (is), because of their common cellular immune-mediated pathophysiology. here we report a phase ii/iii pilot study with alemtuzumab (mabcampath®) (ale) followed by low-dose cyclosporine a (csa) as alternative is regimen in patients suffering from marrow failure ( aa, prca and agr). all patients received subcutaneous ale as escalating dose in consecutive days ( - - - -[ ] mg, total dose for aa and for prca and agr), premedicated by betamethasone, clorpheniramine and paracetamol. six patients received one or more additional courses as a result of relapse, so a total of courses were administered. all patients started oral low dose csa ( mg/kg) on day . an intensive anti-infectious prophylaxis has been exploited, which included oral valgancyclovir and cotrimoxazol as anti-cmv and anti-p. carinii agents, respectively. all patients completed the treatment with no relevant injection-related side effect (with exception of fever in some cases), nor significant clinical or laboratory abnormality. a complete lympho-ablation was observed in all patients within - days, which persisted for several weeks; transient worsening of neutropenia and/or thrombocytopenia were observed in some patients. at a median follow-up of months, infectious events were irrelevant: in cumulative patient-months, hsv and flu have been recorded, all resolving quickly. no cmv reactivation was demonstrated. immune reconstitution was delayed up to several months, especially affecting the cd + compartment. patients with adequate follow up (more than months) were assessed for treatment efficacy: aa patients showed cr, pr, nr. in the prcas, there were cr and nr; out of agrs obtained cr. among responding patients, / saas, / prcas and / agr experienced relapses, which were successfully treated by additional course of ale. in conclusion, subcutaneous ale is a feasible and safe is regimen for patients suffering from immune-mediated marrow failure syndromes. preliminary results suggest excellent response rate, and support efficacy even in case of relapse; such favorable risk-to-benefit ratio predicts for this regimen a leading position in the future is strategies. stable mixed chimerism has been described as being beneficial in patients with aplastic anemia after stem cell transplantation, as these patients were protected from severe graft versus host disease and from rejection, the major drivers of mortality in these patients. patients with marrow failure syndrome do not need full donor chimerism as they do not benefit from a graft-versus-malignancy effect. here we describe a protocol resulting in high rate of stable mixed s chimerism in patients conditioned with cyclophosphamide + atg and transplanted with peripheral stem cells depleted by campath with add-back of t-lymphocytes post-transplant at different dose levels, according to donor-recipient relationship. we included patients in this protocol, median age was , ( - ) years, were male, donors were identical siblings in , mismatched related in matched unrelated in and mismatched unrelated in , respectively. stem cell dose was . ( . - . ) cd x /kg. time to neutrophil engraftment was ( - ) days and to platelet engraftment ( - ) days. median follow-up of surviving patients is days ( - ) acute grade ii gvhd was observed in patient (with the mismatched unrelated donor), this patient died subsequently, while all other patients remain alive. there is mild chronic gvhd in , none of the patients lost the graft. chimerism analysis showed full donor chimerism in , transient mixed chimerism (defined as mixed chimerism developing into full donor chimerism) in , and stable mixed chimerism in , limited to mononuclear cells in and observed in the granulocyte and mononuclear compartment in . mixed chimerism remained stable for up to years. a protocol of conditioning with cyclophosphamide and atg and gvhd prophylaxis with t-cell depletion using campath and t-cell addback may induce stable mixed chimerism in a large proportion of patients with aplastic anemia with low gvhd and graft failure risks. i. resnick* ( ), a. maschan ( ), m. shapira ( ), p. trakhman ( ), e. skorobogatova ( ), d. balashov ( ), m. aker ( ), r. or ( ) ( introduction: ten years ago we introduced a fludarabine (flu) based conditioning for hematopoietic stem cell (hsc) and umbilical cord blood transplantation [ , ] for fanconi anemia (fa). this approach dramatically improved results of fa treatment. following initial case reports and small series [ , ] , two large studies were published: by an italian group and of european experience [ , ] . aim: here we present an update of our two centers' experience of hsc transplantation for fa. methods: thirty patients (age . to , median . years) underwent allogeneic hsc for treatment of fa. used conditioning protocols were flu based in : flu or mg/m + cyclophosphamide (cy) mg/kg or busulfan (bu) mg/kg or both + antithymocyte globulin (atg) atgam mg/kg or fresenius mg/kg. an alternative conditioning ( patients) was cy-tai/tli or bu cy . follow up till december , was to months. results: in the group of patients who got flu based protocols disease free survival (dfs) is % vs % in pre-fludarabine protocols; p= . ; or= . ( . - . ). difference of dfs in patients transplanted from matched family donor ( %, n= ) vs matched unrelated donor ( %, n= ) was insignificant. causes of death were acute graft-vs-host disease (gvhd) grade ≥ (n= ) in combination with sepsis (n= ), multiorgan failure (n= ), liver veno-occlusive disease (n= ), bleeding (n= ), chronic extensive gvhd (n= ), secondary malignancy (n= ). there was no clear advantage in using a combination of cy and bu compared with single agent protocols (dfs out of patients). three patients suffered a rejection and underwent a nd transplant ( are alive and well). all surviving patients are disease free and in an excellent clinical condition. conclusion: combination of fludarabine with atg and low dose cy and/or bu is safe, demonstrates low rejection rates and is well tolerated by fa patients. lower doses of flu ( mg/kg) can be safely used without a pronounced risk of rejection. references: .kapelushnik j et al. bone marrow transplant. ; : . .aker m et al. j pediatr hematol oncol. : . .bitan m et al. biol blood marrow transplant. ; : . .maschan aa et al. bone marrow transplant. ; : . .gluckman e et al. biol blood marrow transplant. ; : . .locatelli f et al. haematologica ( ) results: cb recipients were more likely to have advanced leukemia at conditioning (relapse or induction failure, cb vs. bm = % vs. % in aml patients, and cb vs. bm = % vs. % in all patients, p< . , p= . , respectively). the proportion of all with philadelphia chromosome abnormality was higher ( % to %, p= . ) among cb recipients. human leucocyte antigen (hla) was serologically mismatched in % of patients with aml and % of patients with all among cb recipients, while hla a, b, and dr were all matched genotipically for bm recipients. in controlled comparisons using multivariate analyses, among patients with aml, higher rate of treatment-related mortality (trm) (hazard ratio [hr]= . , % confidence interval [ci], . - . , p= . ) was observed in recipients of cb, which contributed to decreased overall survival (hr= . , %ci, . - . , p= . ). relapse rate did not differ between the two groups of aml patients (hr= . , %ci, . - . , p= . ). in patients with all, the relapse rate was higher with marginal significance among cb recipients (hr= . , %ci, . - . , p= . ). there was no significant difference between these groups for trm (hr= . , %ci, . - . , p= . ) or overall mortality (hr= . , %ci, . - . , p= . ) for patients with all. there was no increase in the incidence of acute graft versus host disease in cb recipients among patients with either aml or all despite hla disparity. conclusions: matched or mismatched cb is a favorable alternative stem cell source for patients without a suitable donor. we found different outcomes between patients with aml and all, which indicates the importance of disease-specific analyses in alternative donor studies. decreasing trm is required to improve the outcome for cb recipients, particularly for patients with aml. . the original minneapolis conditioning regimen was used in ( %) pts and modified in ( or gy tbi: pts; atg : ). characteristics of the grafts: a single unit was infused in pts ( %), two in ( %). hla compatibility was / in pts, / in , / in , ≤ / in ; pts were abo matched. infused nucleated cells (nc) was . x /kg ( - ): . x /kg in single units and . x /kg in double units. csa and mmf were used for gvhd prophylaxis in all pts. results: neutrophils recovery was ± % at a median of days ( - ) ; % pts experienced autologous recovery; % had mixed and % full donor chimerism at d+ . univariate analysis indicated the low weight, previous transplantation, double units and hla compatibility as significant factors for neutrophil recovery; however multivariate analysis did not find any significant factor. acute gvhd was observed in ± % of pts: , and pts had grade ii, iii or iv agvhd respectively and chronic gvhd in %. non relapse mortality was ± % at months; relapse: ± %; overall survival: ± %. causes of death were relapse in pts, gvhd in pts, venocclusive disease and multiorgan failure in , infections in and other toxicity in . dfs at and months were ± % and ± %, respectively. multivariate analysis identified independent risk factors: hla disparity( + vs + ), cell dose (< . x /kg) and age (> y). this assessment of ucbt after nma confirms the good results of the minneapolis group (brunstein et al. blood ) . few events were observed between and mo and dfs remains high and ucbt is a good option in absence of other source of stem cells. background: cord blood transplants (cbt) are associated with delayed or failed engraftment in a significant proportion of patients. we hypothesized that direct intrabone (i.b.) transplant of cord blood cells could improve hematologic recovery. methods: unrelated cb cells were selected for consecutive patients ( patients. / , patients / hla and one pt. / antigen matched). median transplant cell dose was . x /kg (range . - . ). cb cells were concentrated in syringes of ml each and infused in the supero-posterior iliac crest (spic) ( patients bilaterally; patients monolaterllay) under rapid general anesthesia ( min. with propofol). patients' median age was years ( - ); had acute leukaemia ( with refractory or relapsed disease and high risk first remission leukemia); chronic myeloid leukemia in advanced phase; refractory hodgkin's disease; lnh in advanced phase and aplastic anemia/mds. most patients (n= ) were prepared with conventional tbicyclophosphamide. results: the infusion of cells i.b. in spic was uneventful. six patients are not evaluable because they died of multiorgan failure within days from transplant. the patient with saa/mds did not engrafted and was re-transplanted. all the other patients surviving more than days engrafted ( %). median for pmn engraftment (> . x /l) was day ( - ), whereas for platelets (> x /l) it was day (range - ). four patients died of infection; one patients died of ptld on day + . four patients relapsed and died. twenty-two patients are alive in remission at a median follow up of months (range - ). from day + full donor chimerism was documented in cd , bone marrow cells and progenitor cells from both the injected and non-injected spic; from day + , cfc progenitors reached the values of normal individuals in bilateral sites documenting the colonization of the hematopoietc system and possibly an improvement of seeding efficiency. only patients experienced acute gvhd grade ii and grade i; patients have moderate chronic gvhd and one patient extensive cgvhd. conclusion: direct intra-bone transplant of cb cells overcomes the problem of graft failure even when low numbers of hla mismatched cb cells are transplanted. low incidence/severity of acute gvhd was observed. nearly all patients for whom a cb unit was searched were able to undergo cbt. this approach may change the policy of hemopoietic cell transplants. background: the impact of donor-recipient abo matching on outcomes after allogeneic stem cell transplantation (sct) has been a matter of controversy. objective: to evaluate whether abo matching has a significant influence on overall survival(os) in patients(pts) receiving sct for hematologic malignancies. methods: we conducted an individual patient data-based meta-analysis using a pooled dataset extracted from centers(ctrs) not included in previous pooled analyses. we analyzed pts who had received bone marrow or peripheral blood transplantation for hematological malignancies. the primary endpoint was os, comparing pts receiving an abo matched (ma) graft with those receiving a major (mj), minor (mi), or bidirectional abo mismatched (bi) graft using a multivariate (mv) cox model. in addition, os and mortality within days were analyzed with adjustment for confounders. results: a total of sct, including ma, mj, mi, and bi cases, were analyzed. they included related sct (rsct) and unrelated sct (ursct) from western ctrs; rsct and ursct from asian ctrs. the median age of recipients was years (range, - ). the probabilities of os [ % confidence interval(ci)] at years among ma, mj, mi, and bi groups were % ( - ), % ( - ), % ( - ), and % ( - ) with a median followup of months (range, - ). overall, adverse impact on os was observed for bi group [hazard ratios (hr) adjusted by age and sex:mj . ( . - . ), mi . ( . - . ), and bi . ( . - . )]. among recipients of rsct, we observed no significant difference in os between the ma group and any other group. in contrast, mi and bi groups among ursct recipients experienced worse os. sources of heterogeneity were pts who received bone marrow; who had acute leukemia; who underwent transplant at asian ctrs. in mv analysis of os and early mortality adjusted for confounders, mi and bi groups showed inferior os among the subset of ursct, and an increased risk of early mortality was observed only among mj group regardless of donor type [hr ( %ci): mj . ( . - . ), mi . ( . - . ), and bi . ( . - . )]. conclusion: our meta-analysis demonstrates an overall adverse association between bi transplants and survival, largely accounted for by adverse impact of minor or bidirectional abo mismatching on os in ursct. abo mismatching appeared to have little or no impact on os in rsct. larger studies including ursct of various ethnic backgrounds should be performed. intra-bone marrow injection of umbilical cord blood: no impact on rate of haematopoietic recovery c. brunstein, j. barker, d. weisdorf, t. defor, j. wagner university of minnesota (minneapolis, us) neutrophil engraftment after umbilical cord blood (ucb) transplantation is slower than that observed after adult peripheral blood or marrow transplantation.in order to augment engraftment, we evaluated the safety and potential efficacy of ucb intra-bone marrow injection(ibmi). it was hypothesized that direct ibmi to the marrow microenvironment would reduce hematopoietic stem cells (hsc) loss and improve homing. based on our prior experience with two partially hla matched ucb units, the median time to neutrophil engraftment was days (r: - ). trial success required: ) absence of severe adverse events within hours of ibmi, and ) a -day reduction in days to neutrophil recovery (i.e., median anc> /mcl by day ). based on these measures of success, patients were planned. all received cyclophosphamide mg/kg, fludarabine mg/m²/day and total body irradiation cgy with mycophenolate mofetil and cyclosporine a immunoprophylaxis. all pts received ucb units randomly assigned to either iv infusion or ibmi. the ibmi unit was volume reduced to ~ ml with ~ ml infused into each posterior superior iliac crest under local anesthesia at the pts bedside. ten pts were evaluable ( were disqualified due to inability to volume reduce the ibmi unit and ibmi unit was not randomized). median recipient age was years ( - ) and median weight . kg ( - ). seven had acute leukemia and had non-hodgkin's lymphoma. median infused cell doses for the iv and ibmi units were . x tnc/kg ( . - . ) and . x tnc/kg ( . - . ), respectively. pts received two / (n= ), two / (n= ) or two / (n= ) hla matched units. no adverse events were reported with the ibmi procedure. nine of engrafted. median time to neutrophil and platelet recovery (> , /mcl) was ( - ) and ( - )days, respectively. complete chimerism was observed with one unit engrafting long term (ibmi unit in and iv unit in ). seven of evaluable pts had acute graft-versus-host disease (grade ii in and grade iii in ). with a median follow up of months, the probability of survival is %( %ci, - %) at year. based on this interim analysis, the study was prematurely discontinued based on the fact that the predetermined required rate of neutrophil recovery (median days) was unlikely with high degree of certainty. while technically easy and safe, neither neutrophil nor platelet recovery after ibmi were not improved to warrant additional patient accrual. j. fernandes*, v. rocha, d. setubal, m. bierings, m.a. champagne, r. pasquini, g. socié, primary graft failure (pgf) is a fatal complication after allogeneic hematopoietic stem cell transplantation (hsct). unrelated donor cord blood transplantation (ucbt) is frequently reported as having a delayed neutrophil recovery and the incidence of pgf varies from - %. second transplants for pgf have been associated with a high treatment related mortality (trm) and a poor outcome, with an overall survival (os) varying from to %. we retrospectively analyzed patients who received a second ucbt following pgf after a first ucbt for hematological diseases, between and . were defined as pgf patients that did not achieved a neutrophil count > x /l until days after ucbt or that received a second transplant for pgf in this period. patients having relapsed in the first days after ucbt were excluded. median age was . years (range, - ). diagnoses were: acute leukemia (n= ), myelodysplasia (n= ) and bone marrow failure syndromes (n= - =aplastic anemia and =fanconi anemia). patients received either one (n= ) or two (n= ) unrelated cord blood units matched - / (hla a and b low resolution and drb allelic typing) as donors for the first transplant. median time between first and second transplants was days ( - ) and median follow-up after the second transplant was months. concerning the second transplants, patients received mismatched unrelated bone marrow grafts, haploidentical related grafts, single ucb and double ucb grafts. t-cell depletion was used in cases. conditioning regimen was of reduced intensity in the majority of cases, the most frequent being fludarabine-based regimens. low-dose tbi was administered to patients and serotherapy (alg/atg/other moab) was used in patients. median time to neutrophil recovery was days ( - ) and of patients engrafted ( of receiving a single cb unit, of receiving double cb units and of other alternative donors). chimerism analysis for these patients at engraftment showed: =complete donor and =mixed chimerism. twelve patients ( %) developed acute graft-versus-host disease (gvdh) grades ii-iv and chronic gvhd was observed in of the evaluable patients ( %). at year, trm was %± % and os was %± %. in conclusion, these results suggest that second transplants in this set of extremely high risk patients are effective and should be considered early following primary graft failure after ucbt. umbilical cord blood (ucb) is increasingly used as a source of hematopoietic stem cells (hsc) for transplantation in patients (pts) with leukemia. initial reports suggested that ucb transplantation was associated with a - % leukemia freesurvival (lfs). therefore, we evaluated the outcomes in consecutive patients transplanted with ucb after a myeloablative therapy between and for the treatment of acute myeloid leukemia (aml,n= ), acute lymphoblastic leukemia (all,n= ), chronic myeloid leukemia (cml,n= ), and myelodysplastic syndrome (mds,n= ). conditioning consisted of either cyclophosphamide (cy) mg/kg, fludarabine (flu) mg/m²/day and total body irradiation (tbi) cgy with mycophenolate mofetil (mmf) and cyclosporine a (csa) immunoprophylaxis or the same cy/tbi with equine anti-thymocyte globulin (atg) mg/kg and methylprednisolone (mp) immunoprophylaxis. the median age was yrs (r: . - ), median weight . kg (r: - ). most were male ( %), cytomegalovirus seropositive ( %). grafts were composed of two units in % and hla mismatched at one ( %) or two ( %) antigens. the median infused cell doses were . x nucleated cells/kg (r: . - ), . x cd cells/kg (r: . - ), . x cd cells/kg (r: . - . ). pts were classified as standard risk (acute leukemia in cr - or cml in first chronic phase, n= ) or high risk (n= ). median follow-up for survivors is . years (r: . - . ). engraftment occurred in patients ( % ( %ci, - %). incidence of grades ii-iv and iii-iv acute and chronic gvhd was %( %ci, - ), % ( %ci, ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) and % ( %ci, - %), respectively. incidence of treatment-related mortality at years and relapse at years was % ( %ci, - ) and % ( %ci, - ), respectively. lfs was %( %ci, - ) and overall survival was %( %ci, - ) at years. in multivariate analysis, disease risk group was the only risk factor for poor lfs (rr . ; %ci, . - . ;p<. ) and not year of transplant, pt age and weight, nc and cd cell dose, treatment regimen, gvhd, cmv serostatus and hla disparity. clearly, lfs after ucb transplantation, despite hla mismatch, represents a valuable alternative to bone marrow and peripheral blood especially for pts requiring urgent transplant or lacking an hla matched unrelated adult volunteer donor. double umbilical cord blood transplantation using reduced-intensity conditioning: a single-centre experience double umbilical cord blood transplantation (ducbt) reduces the time to engraftment and as a result may reduce associated transplant-related mortality. in this report, we describe our experience with ducbt after reduced intensity conditioning. methods: subjects were treated on sequential protocols between - . all patients received fludarabine ( mg/m²/d x ), rabbit atg ( . mg/kg/d x ) and melphalan ( mg/m² x ) as conditioning. gvhd prophylaxis began on day- and was administered through day and then tapered. cyclosporine/mycophenolate mofetil (cy/mmf) was given to the first patients, and sirolimus/tacrolimus (siro/tac) was given to the remaining patients. all patients received / or better allele level hla-matched cord units and a supportive care regimen of antiviral, antibacterial and antifungal prophylaxis. g-csf was administered from day+ through engraftment. results: the median age was yrs ( - ) and patients ( %) were male. malignant diagnoses were similar although there was a higher proportion of advanced lymphoid malignancy in the siro/tac group. subjects received a combined total of . x tnc/kg and . x cd cells/kg, without cohort differences. engraftment kinetics did not differ based on gvhd prophylaxis, with neutrophil engraftment (anc of ) at a median of days ( - ), and platelet engraftment ( /microl) at a median of days ( - ). patients experienced graft loss between days and ( cy/mmf, siro/tac). grade ii-iv acute gvhd occurred in % ( % cy/mmf, % siro/tac, p = . ). chronic gvhd occurred in %, without cohort differences. -day trm was %, and the risk of relapse at year was % (cy/mmf %, siro/tac %, p = . ). atypical and viral infections were common, and ebv-ptld occurred in patients. with a median follow-up of months, relapse-free and overall survival at year are and %, without cohort differences. causes of death included infection( ), relapse( ), ebv-ptld( ), and other( ). conclusions: dubct following reduced intensity conditioning results in early engraftment and low -day treatment related mortality. the use of siro/tac gvhd prophylaxis is associated with less gvhd than cy/mmf prophylaxis. this approach may lead to a higher relapse incidence, although this may be explained by differences in subject baseline characteristics. dubct should be considered for all adults in whom a suitable adult donor does not exist. ( ) rna encoding the wilms tumor protein (wt ) is overexpressed in the vast majority of patients with acute myeloid leukemia (aml) and peptide vaccination studies have demonstrated that wt is a target for active specific immunotherapy. preclinical data from our laboratory and that of hans stauss have shown that wt mrna-electroporated dendritic cells (dc) stimulate wt -specific t cells in vitro (van driessche a et al. leukemia ; : - . therefore, we started a phase i/ii dose escalation trial in which patients with aml received intradermal injections with wt rnaloaded dc. feasibility, safety, immunogenicity and antileukemic activity of the vaccine were investigated. high risk aml patients (all but one of them in complete remission) underwent a leukapheresis and monocytes were isolated using cd immuno-magnetic beads by clinimacs. dc were generated in -day cultures in clinical-grade medium supplemented with serum, gm-csf and il- and matured with pge and tnf-alpha. keyhole limpet hemocyanin (klh) was added during maturation as a cd + t-helper antigen. mature dc were harvested, electroporated with wt mrna and used as vaccines. eight patients received four biweekly dc vaccines. a delayed-type hypersensitivity (dth) test was performed weeks following the last vaccination. patients were monitored for minimal residual disease (mrd) by analyzing wt rna expression in peripheral blood by qrt-pcr. before and after the vaccination cycle, peripheral blood was collected for immunomonitoring purposes. there was successful dc generation and vaccine production in all patients selected. no serious adverse events or toxicity was seen. a decrease in wt rna expression was observed during the course of the vaccination in / patients who had an increased wt mrna level in peripheral blood at the start of dc vaccination. a vaccine-specific immune response was demonstrated in / patients by dth both to klh as well as to wt . preliminary data from immunomonitoring in pre-and post-vaccination t cell samples from patients show a mixed t helper (th) /th response towards the klh and the wt protein following vaccination. we conclude that vaccination of aml patients with wt rnaloaded dc is feasible and safe. furthermore, the vaccine elicits anti-vaccine t-cell responses in vivo and a decrease in wt rna expression levels was observed during mrd monitoring in some vaccinated patients, strongly suggestive of antileukemic activity of the dc administered. reduced-intensity allogeneic stem cell transplantation using related haploidentical donors for relapsed or refractory lymphomas: evidence of anti-tumour activity in poor prognosis disease p. corradini*, c. carniti, a. raganato, a. vendramin, l. farina, v. montefusco, r. milani, m. milanesi, p. longoni, l. gandola, c. lombardo, a. dodero istituto nazionale dei tumori (milan, it) haploidentical stem cell transplantation (haplo-sct) has disclosed new possibilities for the treatment of hematologic malignancies in patients (pts) who do not have a hla compatible sibling or a fully matched unrelated donor. the role of haplo-sct in relapsed lymphomas is, at present, unknown. we devised a new strategy potentially useful also for pts without nk alloreactive donors (nk alloreactivity in lymphomas is untested). we report the results of a phase i-ii trial evaluating early add-backs of cd -depleted donor lymphocytes (dlis) (from x up to x cells/kg from day+ to day + at monthly intervals) after a reduced intensity conditioning (ric) regimen in pts with relapsed lymphomas. ex-vivo and in-vivo t-cell depletion were carried out by cd + cell selection and alemtuzumab, respectively. histologies were non-hodgkin's lymphomas (nhl, n= ) [chronic lymphocytic leukemia (cll), n= , aggressive nhl (hg-nhl), n= and follicular cell lymphoma (fcl), n= ] and hodgkin's disease (hd, n= ). the median age was years (range, - ). pts received a median number of lines and % failed autograft. at median follow-up of months (range, - months), pts ( %) were alive (n= in durable cr) and died of disease (n= ) or non-relapse mortality (n= ). two-year os and pfs of the whole group were % ( % ci, %- % ) and % ( % ci, %- %), respectively. the main factors influencing -year os were age (≤ years: os of %) and chemosensitivity of disease at time of sct ( % versus % for chemosensitive versus chemorefractory, p< . ). before dlis, only of pts ( %) developed acute gvhd. cd -depletion, perfomed using an immunomagnetic method, reduced the content of cd + tcells by a median value of . log ( . - ). a total of cd depleted dlis were administered to pts. three pts ( %) developed acute gvhd grade ii-iv following dlis. eight pts receiving cd -depleted dlis never experienced a relapse. we observed a statistically significant expansion of cd + and cd +, but not of cd + and nk cells, at day + following cd -depleted dlis. eight pts had measurable trec/µg at year after sct. spectratyping of t cells demonstrated a polyclonal vß repertoire: pre-transplant %, post-transplant %. escalated doses of cd -depleted dlis increase the level of cd + in the first months after haplo-sct and are feasible. this is the first report of long-term remissions in lymphomas after ric haplo-sct. k. perruccio*, f. topini, a. tosti, a. carotti, t. aloisi, f. aversa, m. f. after haploidentical stem cell transplantation, immune recovery is slow due to decaying thymic function and extensive t-cell depletion of the graft which is needed to prevent graft-versus-host disease (gvhd). consequently, infectious related mortality is about - %. to address this problem, we investigated the efficacy of adoptive immunotherapy after photodynamic purging of alloreactive t cells (theralux tm , kiadis pharma, amsterdam, the netherlands) in preventing gvhd and improving immune s reconstitution. optimized protocol conditions provided , ± (mean ± sd)-fold allodepletion, full retention of tregulatory cells, and preservation of pathogen-specific t-cell responses (against aspergillus, candida, cytomegalovirus (cmv), adenovirus (adv), herpes simplex virus (hsv), varicella zoster virus (vzv), toxoplasma antigens). here we present the preliminary results of a clinical trial. escalating doses of photodynamically allodeleted donor t cells, i.e., . x /kg, . x /kg, x /kg and x /kg, were infused into groups of haploidentical transplant recipients. only patient developed grade iii agvhd at the x /kg cell dose and responded to immune suppressive treatment. immune assessment analyses revealed that infusion of cell doses equal or greater than x /kg are associated with significant reconstitution of t-cell counts and appearance of pathogen-specific t-cell responses. three weeks after infusion, cd + and cd + t cells were ± /cmm and ± /cmm (versus ± /cmm and ± /cmm respectively, in patients receiving t-cell doses below x /kg, p = . ). aspergillus, candida, cmv, adv, hsv, vzv, toxoplasmaspecific cd + and cd + t-cell responses had recovered to frequencies within the normal ranges while they were absent in patients who received t cell doses under x /kg (p = . ). in conclusion, this study demonstrates the feasibility, safety and preliminary indications of efficacy of adoptive immunotherapy after photodynamic purging of alloreactive t cells in recipients of haploidentical stem cell transplantation. a larger study will evaluate the impact of these t-cell infusions on transplant related mortality and disease free survival. leukemic stem cells (lsc) are a self-renewing subpopulation of malignant cells which is thought to play a central role in the pathogenesis of acute leukemia. lsc are likely contributing to both disease initiation and relapse and therefore represent an important therapeutic target. resistance of lcs to standard chemotherapy agents is an important consideration for the development of new therapies. in this study, we asked whether lsc of acute myeloid leukemia (aml) can be recognized and eliminated by the anti-tumor function of natural killer (nk) cells. using cd dimcd +cd -as a phenotypic characteristic of the lsc population in aml, we purified these cells from peripheral blood of patients with de novo aml and examined their sensitivity to alloreactive nk cells. for this purpose, the recently described nk cell lines with single kir specificities and mismatched with respect to hla-class i allotype of target leukemic cells (diermayr et al, blood online) were employed. using the hematopoietic colony forming unit (cfu) assay in methylcellulose, we demonstrated that purified lsc gave rise to hematopoietic colonies which were maintained upon a serial passage. when lsc were preincubated with alloreactive nk cells the growth of aml cfu was significantly reduced. next, we demonstrated that cd dimcd +cd -aml cells do not express the cell surface ligands, mic and ulbp, for the major nk cell activating receptor nkg d. incubation of purified cd dimcd +cd -cells with histone deacetylase (hdac) inhibitor valproic acid (va) for days, resulted in an up to fold upregulation of mic or ulbp ligands in / tested aml samples. the va treatment had no impact on aml cfu formation in the absence of nk cells, but abrogated the clonogenic growth when applied together with alloreactive nk cells. importantly, the effect of va was limited to lsc since normal hematopoietic stem cells were unaffected, both with respect to expression of nkg d ligands as well as the colonyforming efficiency. taken together, our results indicate that aml lsc are susceptible to nk cell-mediated cytotoxicity and that expression of nkg d ligands makes lsc more accessible to nk cells. we propose that adoptive transfer of alloreactive nk cells in combination with pharmacological use of hdac inhibitors merit evaluation as novel approaches to prevent relapses of aml with nk cell immunotherapy. these approaches are now evaluated in an in vivo model of human aml transplanted into nod/scid mice. survival in patients receiving a transplant from an hlamismatched donor (at / alleles) can be significantly improved by selecting a donor mismatched for hla-dpb b. shaw* ( , ) , n. mayor ( ) recipient/donor hla matching is an important determinant of outcome in transplantation using volunteer unrelated donor (vud). there is evidence that matching for hla-a, -b, -c, -drb , -dqb results in a beneficial outcome. the impact of matching for hla-dpb is more controversial and few studies have considered the additive effect of --dpb . we investigated the outcome, dependent on the degree of hla matching at alleles, in recipients of vud transplants for leukaemia. the patients had aml ( ), all ( ) and cml ( ). the majority of patients had t cell depletion as gvhd prophylaxis. patient/donor pairs were matched for / alleles. of these, were matched in addition for dpb . pairs had a single allele mismatch ( / ). of these, were matched and were mismatched for dpb . pairs had multiple mismatches (≥ ), in dpb was matched. we found a significant difference in overall survival when comparing these groups (fig , p= . ). survival was not significantly different in the / matched transplants dependant on dpb matching (p= . ). however, there was a significant difference in survival dependant on dpb matching within the hla mismatched group (n= ) ( years: % dpb mismatched compared % dpb matched, p= . ). in particular, in pairs with a single hla mismatch at -a, -b, -c, -drb , -dqb , the presence of a dpb mismatch improved overall survival ( years: %) compared to dpb match ( %, p= . ). in multivariate analysis including disease, stage, patient/donor age, patient cmv status and gender, the significant survival benefit of dpb matching persisted (or . ; % ci . , . ; p= . ). this effect appeared to be mediated both by a decrease in relapse in the dpb mismatched pairs ( yrs: % compared to % in dpb matched pairs, p= . ) and a decrease in trm ( yrs: % compared to % in dpb matched pairs, p= . ). no differences in acute or chronic gvhd were seen. we conclude that in patients who receive hla-mismatched grafts (at hla-a, -b, -c, -drb , -dqb ), a donor who is dpb mismatched in addition results in a superior outcome compared to one who is dpb matched. as physicians frequently have a choice between mismatched donors, we believe that selection by dpb within this group will be possible and beneficial. additionally, dpb is the only hla allele for which matching status impacts on disease relapse. we speculate that this may suggest a difference in the functional mechanisms of dpb . the cxcl /cxcr chemokine pathway specifically targets alloreactive t-cells to the bone marrow of mice with leukaemia r. van der voort, v. verweij, f. maas, j. vos, m. philippens, t. de witte, h. dolstra* umc st radboud (nijmegen, nl) allogeneic stem cell transplantation is an effective treatment for patients with leukemia. this therapeutic effectiveness is largely attributed to the graft-versus-tumor (gvt) response during which alloreactive donor t cells eradicate minor histocompatibility antigen (miha)-expressing tumor cells in the bone marrow (bm). unfortunately, subsets of alloreactive t cells recognize miha in healthy tissues, such as the gut, skin and liver, resulting in graft-versus-host disease (gvhd). thus, there is a strong need to separate gvt responses from gvhd. an appealing strategy to achieve this goal would be to increase the migration of alloreactive t cells to the tumorcontaining bm and/or block the infiltration of gvhd-prone organs. although the homing receptors for migration into the gut and skin are extensively studied, those involved in t cell trafficking to the bm are largely unknown. here, we characterized the pathway that alloreactive t cells exploit to infiltrate leukemia-containing bm. t cells were isolated from bm and spleen (control organ) from mice with or without acute myeloid leukemia (aml) that had previously received an allogeneic bm-transplant (bmt). in addition, we used naïve mice as controls. the expression of activation markers and a panel of putative homing receptors was determined by flow cytometry, and the level of several chemokines by real-time pcr and elisa. migratory behavior was analyzed by transwell migration assays and flow chamber assays. where bm of naïve and bmt mice contained few t cells, amlcontaining bm was heavily infiltrated with cd + and cd + t cells with an effector/effector memory phenotype. interestingly, bm of mice with leukemia predominantly recruited cd + and cd + t cells expressing the chemokine receptor cxcr . in contrast, the number of cxcr + t cells in bm from naïve or bmt mice without aml, and in all spleens was generally low. furthermore, the ligand for cxcr , cxcl , was expressed by activated macrophages and dendritic cells and its expression level increased in bm of mice with aml as compared to controls. finally, we show that cxcr + t cells are strongly attracted by soluble cxcl and show increased binding to the endothelial adhesion molecule vcam- upon stimulation with cxcl . in conclusion, our data suggest that the cxcl /cxcr axis is a novel pathway that specifically regulates the recruitment of alloreactive t cells into leukemia-containing bm. the pd- /pd-l co-inhibitory pathway is involved in functional impairment of alloreactive cd + t-cell responses against leukaemia w. norde ( ) , f. maas ( ) , h. fredrix ( ) , a. schattenberg ( ), m. kester ( ) , f. falkenburg ( ) , r. van der voort ( ) donor lymphocyte infusion (dli) following allogeneic stem cell transplantation (sct) is a potent treatment for hematological malignancies. the effectiveness is attributed to the graftversus-tumor (gvt) response, during which donor cd + t cells become activated by minor histocompatibility antigens (miha). consequently, these alloreactive cd + t cells clonally expand, acquire effector functions and kill mihapositive malignant cells. after the response, most cd + t cells die through apoptosis, while a small population remains to form a pool of long-lived memory cells. unfortunately, tumor relapses still occur despite the presence of miha-specific memory t cells in the patient for many years, suggesting that these memory t cells become impaired over time contributing to tumor evasion. recently, a crucial role for the co-inhibitory pd- /pd-l pathway in inhibiting the function of virus-specific cd + t cells was demonstrated in chronic viral infections. here, we investigated the role of the pd- /pd-l pathway on the functionality of miha-specific cd + t cells. initially, we analyzed expression of activation and co-signaling molecules on miha-specific ctl following restimulation with irradiated miha-positive ebv-lcl plus il- . we observed that pd- expression was upregulated on miha-specific ctl within hours after stimulation. in addition, we analyzed miha-specific cd + t cells from leukemia patients after dli. we observed increased levels of pd- on miha tetramer-positive cd + t cells, compared to tetramer-negative cd + t cell subsets in the same patient. knowing that sustained expression of pd- is associated with diminished proliferative capacity of virusspecific cd + t cells, we determined whether blocking the ligation of pd- could increase the capacity of miha-specific cd + t cells to proliferate in vitro. interestingly, we revealed that the addition of anti-pd-l antibody augments the proliferation of miha-specific cd + memory t cells. notably, addition of peptide alone or in combination with an irrelevant antibody did not induce proliferation of these cells. finally, we observed that primary leukemia cells and various hematological cancer cell lines express the pd- -ligand pd-l following stimulation with ifng. these data suggest that pd- /pd-l interactions in the tumor environment may impair miha-specific cd + t cell function, and intervening with this co-inhibitory pathway may result in improved gvt immunity after allogeneic sct. the graft versus-leukemia-effect in b-cell chronic lymphocytic leukemia (b-cll) appears to be less efficient compared to myeloid leukemias. since the aberrant coexpression of cd in association with cd is a hallmark of b-cll, we asked if both molecules might serve as a combinatorial t-cell target. b-cll cells as third party in mixed-lymphocyte reactions (mlr) induced t-cell anergy. we used recombinant proteins comprising a single chain (sc) fvcd antibody (ab) fused to human interleukin (il)- via the hinge region of human immunoglobulin (ig)g in conjunction with a bispecific s cd x ab (bsab; clone okt xhd ) to saturate cd and cd binding sites on b-cll cells. consistent with the absent immunogenic potential, b-cll cells pretreated with scfvcd ab, native il- and cd x bsab were not recognized by allogeneic resting t cells. in contrast, b-cll coated with the dimeric scfvcd -il- and cd x bsab induced profound stimulator cell dependent allogeneic and autologous t-cell proliferation. in addition, t cells from fresh peripheral blood mononuclear cell samples from b-cll patients (n= ) pretreated with dimeric scfvcd -il- and cultured in the presence of cd x bsab expanded by - log within days. notably, addition of soluble exogenous il- to both types of culture remained inferior compared to the cd -targeted il- delivery. t-cell proliferation and expansion also resulted in cell-mediated cytotoxicity. in conclusion, a dual targeting approach using aberrantly expressed tumor cell surface molecules for membrane delivery of dimeric il- molecules using a fusion cytokine in conjunction with a bispecific antibody as shown for b-cll represents a strategy to reverse tumor-induced t-cell anergy. hurler's syndrome (hs), the most severe form of mucopolysaccharidosis type-i causes progressive deterioration of the central nervous system and death in childhood. allogeneic-stem cell transplantation (sct) before the age of two years halts disease progression and prolongs life. graft-failure and mixed-chimerism ( - %) limit the success of sct for hs. unrelated-cord blood transplants (ucbt) are suggested to be a good alternative option for bone marrow, however, little is known about risk factors for outcomes after ucbt for this disease. we have analyzed hs children that received an ucbt from to and were reported to eurocord or transplanted at duke university. median age at ucbt was , ( , - ) yrs, and median follow up was ( - ) mths. the donor was hlaidentical (hla-a and b by low resolution and hla-drb by high-resolution) in cases ( %) and incompatible in cases ( %: most with ( %) and ( %) hla disparities). the median nucleated cell dose/kg and cd +/kg at infusion were respectively . ( - )x and , ( , - )x . with the exception of patients, all received a busulfan/cyclophosphamide (+fludarabine: ) regimen. all patients received atg or campath ( ). results: median days to neutrophil and platelet recovery were ( - ) and ( - ) days, respectively. mixedchimerism was found in only %. all patients had normal enzyme levels after engraftment. in multivariate analyses for neutrophil recovery, a cd ±dose of > . x /kg (hr= . ; p= . ) was associated with increased probability of recovery. acute-gvhd (grade ii-iv) was observed in %, while chronic-gvhd was seen in % at years. two years overall survival (os) and disease/event free survival were % and %, respectively. for years os, time from diagnosis to ucbt more than months was associated with increased risk of death ( % for those children transplanted earlier and % for those transplanted later: p= , ). in conclusion, outcomes following ucbt for hurler syndrome are encouraging. ucb appears to be a good alternative allogeneic stem cell source to transplant children with hs. earlier transplantation and higher cell dose are associated with better outcomes after ucbt for hs patients. introduction: transplant for mpsih has been associated with high rates of morbidity, mortality and graft failure. some of the difficulty of the transplant reflects pre-transplant co-morbidities including significant cardiac dysfunction, respiratory including upper airway compromise and visceral organ enlargement. in recent pharmacological enzyme replacement therapy (ert) has been available that will ameliorate many of these comorbidities. manchester protocol. we give weekly doses of ert prior to conditioning and continue until donor cell engraftment. patients and methods. we have performed transplants in patients in manchester. of these patients have received ert with transplant. in of these there has been full intensity conditioning with pk guided oral (hd) busulfan ( mgs/m²/dose, n= ) or iv busulfan with pk monitoring but not adjustment. results: of these patients received oral busulfan and the most recent have received iv busulfan. of these patients are alive with full donor cell engraftment. one patient experienced primary graft failure following a cord transplant and was successfully transplanted with a second cord and despite achieving full donor chimerism died of adenovirus months after the graft. donor source for these patients was mud cord in , other mud in (pbsc donations) and matched family donors in (pbsc in ). efs survival for this group is therefore % with a median follow up of months. our engrafted survival rates for previous patients treated on previous protocols are %, including second grafts to maintain donor chimerism. previous patients receiving ert and reduced intesity grafts experienced rejections. administered busulfan was significantly higher in those receiving hd busulfan -they received a mean of mgs/kg compared with the previous dose of mgs/kg -(p< . , students t-test). discussion: manchester is the largest centre for mps transplants in europe and this is the largest collection of patients treated with both ert and sct. there were concerns that pre-transplant ert would increase rates of graft rejection. these data refute those concerns. we believe that the improving results reflect reduced patient co-morbidity after ert, improved delivery of full intensity chemotherapy with busulfan and better donor matching including use of unrelated cord blood as a donor source. s o haematopoeitic stem cell transplantation for chronic granulomatous disease -a single-centre experience e. soncini* ( ), m. slatter ( ) , l. jones ( ), s. hughes ( ), t. flood ( ) , d. barge ( ) , g. spickett ( ) chronic granulomatous disease (cgd) is a primary immunodeficiency affecting phagocytes; mutations in genes coding for subunits of nicotinamide adenine dinucleotide phosphate cause failure of oxygen metabolite generation, resulting in impaired microbial killing, susceptibility to bacterial & fungal infections and lung & gut inflammation. antibacterial & antifungal prophylaxis with cotrimoxazole & itraconazole improve short and medium term survival but do not cure cgd; patient quality of life is burdened by frequent hospitalizations, recurrent diarrhoea, inflammatory lung damage, failure to thrive and a % risk of death by the third decade of life. haematopoietic stem cell transplantation (hsct) can cure the disease. we report our single centre experience. between and , patients underwent hsct for cgd in newcastle. a retrospective analysis of medical records to november reviewed age at hsct, donor type, hla matching, conditioning, immuno-reconstitution, significant post-hsct complications, growth, lung function and outcome. received bu/cy, flu/melph, flu/bu, bu/mel conditioning. had sibling, unrelated donors. had / hla-match, / hla-match (a, c mismatch), / hla-match (dr & dq, hvg direction only). received marrow, pbsc and cord blood transplants. of ( %) patients were alive with functioning neutrophils with follow up . - . years. / with significant inflammatory lung disease had an improvement in lung function. all patients with colitis had resolution of symptoms, with impressive catch-up growth. patient had cytopenias secondary to post hsct-cmv infection, there were no other significant infections, only had significant gvhd, liver that resolved with treatment and gut gvhd that resolved with prolonged steroid treatment, with resulting osteoporosis and avascular hip necrosis. patients died, both of complications relating to active fungal infection. / survivors are off prophylactic medication with excellent life quality. hsct is curative for patients with cgdcomplications are less frequent and outcome better in those without pre-existing infection or inflammation. hsct should be considered early in the treatment of cgd, particularly if a suitable donor is available. long-term outcome and quality of life after haematopoetic stem cell transplantation in osteopetrosis: a single-centre experience d. moshous ( ) background: malignant infantile osteopetrosis (miop) is caused by defective osteoclast function. impaired bone resorption is leading to increased bone density resulting in progressive marrow failure and extramedullary haematopoiesis. the abnormal bone remodelling generates also compression of cranial nerves, especially the optical nerve, causing various degrees of visual impairment up to complete blindness, starting early in life. some patients present also neurological deterioration which is not clearly explained by the pathophysiology of miop. the genetic basis of op is heterogeneous, until now four genes with autosomal recessive inheritance have been identified: tcirg , clcn , ostm and rank-l, but there seem to be also autosomal dominant forms. in the absence of treatment, children with miop die early in life often because of bleeding or infection. the only curative treatment available consists in haematopoetic stem cell transplantation (hsct). patients and methods: we report on hsct which were performed in patients ( females and males) in our unit between and . stem cell sources were bone marrow, peripheral blood stem cells and one unrelated cord blood unit. the donors were haploidentical related donors ( / ), hla-identical siblings ( / ), related matched donors ( / ), mud ( / ) and unrelated cord blood ( / ). the majority of patients received hsct before the age of three months ( days to months), with a mean age at first transplant of , months. two patients have been transplanted at an older age, at years months and years months respectively. conditioning consisted mostly in busulfan and cyclophosphamid based regimens, in cases fludarabine was added, atg was used in cases. the injected cell number varied from , - , x nucleated cells/kg, corresponding to , - , x cd + cells. outcome: for two patients the follow up is still too short to conclude. out of evaluable patients survived (overall survival: %). the causes of death were interstitial pneumonia in patients, veno-occlusive disease in patients, multiorgane failure in patients, rejection in patients, transplanted related toxicity in patients, bleeding in and infection in patient. survival was better in the setting of hlamatched hsct / ( , %) versus hla-mismatched hsct / ( %. three patients received , one patient hsct. detailed data especially on long-term neurological development will be provided. ( ) ). there was no graft rejection; mixed donor chimerism occurred in %. the incidence of an acute gvhd ≥ ii was %, that of a chronic gvhd ii % with no difference between mrd and mud. cgvhd was associated with more than expected mri changes years after hsct (p < . ). moreover, patients with unexpected disease progression often displayed poor immunologic reconstitution and late viral problems. of pts. (= %) in group ii/iii remained neurologically stable with good quality of life; of these continued to visit a regular school. all pts. in group i deteriorated after hsct; however, out of pts. stabilized and full dementia could be prevented. conlusion: the above results confirm the role of a favorable disease stage for hsct outcome, as already shown (c. peters, blood, ) . in addition, the data demonstrate the importance of transplant-associated factors (e.g. cgvhd). in our view, the superior neurologic outcome after mrd-bmt allows a less restrictive indication for hsct in more advanced ccald patients. j.-s. kühl* ( ), g. strauß ( ), b. weschke ( ), w. köhler first successful bone marrow transplantation for x-linked chronic granulomatous disease using a sibling donor selected after preimplantation female sexing and hlamatching j. reichenbach* ( ), h. van de velde ( ), m. de rycke ( ), c. staessen ( ) , p. platteau ( ), p. baetens ( ), t. güngör ( ), ( ), i. libaers ( ) ( objective: allogeneic hematopoietic stem cell transplantation (hsct) from an hla-identical donor is currently the only proven curative treatment for chronic granulomatous disease (cgd). hsct with alternative donors is associated with higher morbidity and mortality. the objective was therefore to perform in vitro fertilization (ivf) and preimplantation hlamatching combined with female sexing for hsct in x-linked cgd. methods: ivf followed by preimplantation genetic diagnosis (pgd) was used to identify a female hla-genoidentical embryo in a family who needed a suitable donor for their boy affected with cgd. two pgd cycles were performed. results: in the second pgd cycle two hla-genoidentical female embryos were transferred and a pregnancy was obtained. the mother had a normal delivery of a healthy girl at term. conventional hsct had to be performed, due to insufficient cell numbers in the cord blood source, at age months of the donor and age / years of the recipient and resulted in complete stable donor chimerism and immunological reconstitution up to months post hsct. conclusion: hsct after ivf and combined female sexing and hla matching offers a new therapeutic option for patients with x-linked primary immunodeficiency such as cgd needing hsct but lacking an hla-genoidentical donor. a multicentric comparative analysis of outcomes of hla identical related cord blood and bone marrow transplantation in patients with beta-thalassemia or sickle cell disease n. kabbara* ( ), f. locatelli ( ) , v. rocha ( ) most patients with beta thalassemia major (tm) or sickle cell disease (scd) can be cured by hematopoietic stem cell transplantation (hsct) from either cord blood (cb) or bone marrow (bm). one advantage of cb is the absence of risk associated with donation. in order to compare outcomes after hsct with cb or bm, we studied patients with tm or scd who received hla identical sibling cb (n= ) or bm (n= ) allografts between and . in order to avoid center and period effect, only centers that performed both types of hsct during the same period were included. we compared the incidence of hematopoietic recovery, acute and chronic graft-versus-host disease (gvhd), disease-free survival (dfs) and overall survival (os) after cb and bm transplantation. compared to bm, cb recipients were significantly younger (median of . y versus . y), smaller ( kg vs kg), and were transplanted more recently (in vs ) . the bm group consisted of ( %) scd and ( %) tm patients, and the cb group, ( %) scd and ( %) tm patients. the indications for transplantation in scd were not statistically different between cb and bm groups. more tm patients belonging to pesaro ii-iii risk classes received bm ( %) compared to cb ( %) (p= . ). there were also differences in the conditioning regimen (more frequent use of atg/alg in the bm group and of fludarabine and thiotepa in the cb group) and gvhd prophylaxis (more methotrexate-containing therapy in the bm group compared to the cb group). in addition, the nucleated cell content was times higher in bm compared to cb. the table below shows the non-adjusted univariate analysis for outcomes for all patients according to the stem cell source used. in tm patients, the year-dfs rates were % and % for bm and cb recipients, respectively, and in scd patients, % and %, respectively. in a multivariate analysis adjusted for age and type of hemoglobinopathy, dfs was not statistically different between cb and bm recipients (rr= . , p= . ). in conclusion, patients with tm or scd had excellent outcomes after hsct whether they received stem cells of cb or of bm from an hla identical sibling. these results strongly suggest that cb transplantation from hla identical siblings should be pursued when possible to avoid the discomfort and risks of a bone marrow harvest. s o communication biases during informed consent for unrelated bone marrow transplantation in adult thalassaemia patients g. caocci* ( ), s. pisu ( ) ( ), g. la nasa ( ) ( )r. binaghi hospital -asl (cagliari, it); ( )cagliari university (cagliari, it); ( )gimema data center (rome, it); ( )oxford university (oxford, uk) although there are numerous guidelines to evidence-based medicine, few explain how to build the information into patient oriented decision-making. several factors may hamper physician-patient communication and compromise the informed consent process. in an attempt to eliminate some of the barriers to understanding, we investigated the main factors of communication between physicians and adult thalassemia patients transplanted from an unrelated donor twenty-five transplanted thalassemia patients and physicians were given a questionnaire to investigate the communicated, perceived and recalled risk for mortality and graft-versus-host disease (gvhd), the acceptable risk percentage, besides the motivation and external influences underlying the choice of undergoing hsct. the risks of dying or gvhd perceived by the patients were significantly lower than those communicated by the physicians (p=. , p<. , respectively). also the perception of severe gvhd as a life-threatening condition was much lower (p=. ). younger patients perceived a significantly higher risk of dying than older patients (p=. ). females perceived a significantly higher risk of developing gvhd or dying than males (p<. in both cases). the median percentage communicated for the risk of dying was significantly higher than the percentage remembered by the patients ( % vs %; p=. ). the median percentage considered to be acceptable for the risk of dying was significantly higher in the patients compared to the physicians ( % vs %; p=. ). hence, the balance between the risks and the benefits of the transplantation procedure had not been fully understood by the patients. therefore, it is important that physicians make every effort to overcome communication bias, heuristics and distorted processes of remembering or understanding that could possibly influence the informed consent and decisionmaking process. in adults with aml (n= ), % received an hla identical family hsct, % an autograft, % an hla matched unrelated hsct, % an hla mismatched hsct( % haplo, % hla unrelated and % cord blood). in adults with all (n= ), % received an hla identical family hsct, % an autograft, % an hla matched unrelated hsct, % an hla mismatched hsct ( % haplo, % hla unrelated and % cord blood). for adults in first cr with all or aml given a hla identical sibling donor, -years overall survival (os) was %; it was % for those given an autograft and % for those patients given a hla matched unrelated transplant. for adults with aml given an unrelated cord blood or a haplo-transplant in first cr, overall survival at years were % and % respectively. many studies have been performed and published on behalf of the alwp with the collaboration of many ebmt centres. thanks to the ebmt centres, physicians and data managers, we have been able to collect specific information for those studies. therefore, in , studies and one editorial have been published in the most important journals in the field of medicine, hematology and transplantation, retrospectives and prospective studies are on going and of abstracts were oral presentation at the last ash meeting. moreover, the establishment of subcommittees within the alwp, in the different fields of transplantation has brought enthusiasm among participants and increased the number of participants to up to in the alwp meetings. we would like to thank w arcese, b labar and o ottman for their input as heads of alternative donor, developing centres and phi+all subcommittees, respectively. they have accomplished two years as subcommittee's heads. we would like to welcome a nagler, s giebel and j esteve as new heads of alternative donor, developing centres and molecular markers subcommittees, respectively. i take also this opportunity to thank m mohty and c schmid for their activity in the alwp. we hope that the enthusiasm inside the alwp can motivate young physicians and more ebmt centres to join us with their ideas and input to perform important studies for the ebmt community and importantly to continue improving the field of transplantation for patients in need. the main objective of this new subcommittee is the analysis of the impact of main molecular markers on the outcome of stem-cell transplantation. several molecular lesions described in recent years are contributing to a biological characterization of acute leukemia and provide relevant prognostic information. in this regard, mutations of flt- , nucleophosmin (npm ) or cebpa genes, frequently found in the large subgroup of aml with normal karyotype, arise as the most relevant prognostic factors in this subset of patients. nonetheless, the precise role of different transplant modalities for each of these aml categories remains to be defined. thus, a recent analysis performed within the alwp confirmed a higher risk of relapse associated with flt- itd after myeloablative allotransplant transplant from an hla-identical sibling, although the procedure seemed to benefit a significant proportion of flt- itd aml patients transplanted in first cr, according to the survival plateau observed. nonetheless, the optimal transplant policy in these molecularly-defined high-risk patients is not firmly established. the sc is also aimed to investigate the results of transplant in patients with rare subtypes, such as aml with translocation t( ; )/dek-can or aml with q /evi rearrangement. given the low frequency of these entities, only those studies performed in large cooperative groups have the potential to elucidate the role of different transplant strategies for these specific subtypes. another objective of the sc is the design of biologically oriented studies analysis on the outcome of transplant in adult all, which might help to clarify the controversial role of transplant in early phases of the disease. thus, specific studies for wellrecognized prognostic categories of adult all such as mllrearranged, hypodiploid varieties or results of transplant among different t-all subtypes can be sponsored by the alwp. finally, the emergence of molecularly targeted therapy such as tyrosine kinase inhibitors in philadelphia positive all or conjugated anticd monoclonal antibody in aml are changing the natural history of the disease and, therefore, might modify indications and results of transplant in patients treated with these agents. reduced-intensity conditioning m. mohty* institut paoli-calmettes (nantes, fr) in the last decade, the so-called non-myeloablative or reduced-intensity conditioning (ric) regimens for allogeneic stem cell transplantation (allo-sct) have emerged as an attractive modality to decrease allo-sct-related toxicities. indeed, ric allo-sct represents an attempt to harness the well-documented immune graft-versus-tumor (gvt) effect while attempting to overcome toxicity. the work of different pioneering groups rapidly proved that this approach is feasible in several disease settings or patients' categories, and had the added benefit of expanding the transplant option to patients who are ineligible for myeloablative allo-sct. unfortunately, and despite several thousands of patients receiving ric allo-sct reported to international registries, the true value of ric allo-sct in the management of haematological malignancies is, as yet, difficult to delineate. several reasons can help understanding these difficulties. a consistent definition of "non-myeloablative" or 'ric' regimens is still lacking. the different ric regimens comprise a continuum that overlaps with standard myeloablative regimens. where in theory, reduction of the "inflammatory" component of the conditioning as in ric regimens may lead to the reduction of the incidence of gvhd, gvhd remains a matter of concern after ric allo-sct, raising questions about the need for continuous immunosuppression and its corollary of long-term infectious complications. also, the notion or definition of "ineligibility" for conventional standard allo-sct is not clearly defined. the goal of the ric subcommittee within the acute leukemia working party of ebmt is to address through retrospective and prospective studies, the specific role of ric allo-sct in net health outcomes that should include, in a specific disease setting, an analysis of disease-free survival and overall survival balanced against treatment-related toxicity, quality of life, complications and death. transfusion of donor lymphocytes (dlt) for treatment of leukaemia relapse after allogeneic stem cell transplantation (sct) has been a milestone in the field of immunotherapy against malignant disease. it can be regarded as the proof of principle for the graft-versus-leukemia effect. however, in contrast to the impressive results obtained in cml, results in acute leukemias have been inferior, although there is a small subgroup of patients that obviously responded. the immunotherapy subcommittee of alwp has set the goal to study the role of cellular immunotherapy more in detail with respect to different diseases as well as distinct variants of allogeneic sct. hence, a large retrospective analysis of dlt in relapsed aml has been performed, and similar investigations have been started in all. further, the role of dlt after haploidentical and ric transplants is in the focus of ongoing studies. finally, the conditions and the results of prophylactic or preemptive dlt, given after sct to chimeric patients in cr or in a minimal residual disease status, are currently investigated. use of hsct for patients with acute leukemia is increasing in europe. therefore many ebmt centers have been established in all countries, mainly in the new eu countries. although the number of transplants in eastern countries is growing and the results are improving, the scientific activity appears still insufficient. enhancement of the scientific collaboration in eastern europe is a principle goal of the developing centres subcommittee. to reach this goal it is planned to create a panel of young investigators representing major centres from the region. this panel is expected to discuss current clinical practice as well as to share laboratory experience. on this background it will be able to propose and coordinate multicentre studies including: ) prospective clinical trials, ) retrospective analyses, ) biological studies on hsct for acute leukaemia. other goals of this subcommittee is to evaluate outcomes of hsct performed in eastern countries as well as to develop studies and methods for analyzing the role of center effect on the hsct outcomes. with these objectives two studies have been performed. in the first one, we have analyzed patients (aml, ; all, ) treated with hla matched related donor in first complete remission in countries, from to . with this study we have observed that results of mrd-hsct for acute leukemia in eastern europe improved over time, as a consequence of decreased nrm. another study performed was the update of the lancet paper which has demonstrated a center effect in major ebmt centers. we have updated this study and we were able to show a persistent centre effect but improvement of lfs of hla identical hsct for adults with aml in first cr over the period - . other study proposals have been discussed such as the use of economic and social markers in order to evaluate transplant outcomes in the different eu countries in collaboration with yearly survey of a gratwohl. currently, we can find an alternative donor for almost all patients without an hla identical sibling donor, such as hla matched unrelated, haploidentical and cord blood unrelated donor. the objective of this subcommittee is to study the feasibility of those different strategies and their outcomes and compare their results. with the active collaboration with eurocord, we are able to perform studies of unrelated cord blood transplantation. a recent study in collaboration with eurocord, that will be presented during this meeting, has shown the impact of kir ligand mismatching on decreasing relapse and improving lfs in ucbt recipients with acute leukemia. also comparative studies of outcomes after ucbt and haplo have been performed in children and adults. one of the main objectives of this subcommittee is to improve the hla data of the database and therefore be able to study the impact of specific hla alleles on outcomes of hsct recipients, such as the impact of permissive hla-dpb alleles. with the collaboration with the immunobiology wp and donor registries, we hope to improve the data of hla high resolution typing of unrelated donors. other projects are i) to study the influence of gene polymorphisms with outcomes of alternative hsct ii) to compare various conditioning regimens in this setting; iii) to target therapy with the novel drugs preand post-alternative transplants and iv) to compare immune reconstitution after mud vs. haplo vs. cbt. update of the registration study v. rocha*, e. polge, m. arat, v. koza, h. wandt, t. ruutu, a. ferrant, g. milone, p. defabritis, w. arcese, l. verdonck, a. bosi, b. allione, a.l. herr, f. pinto, c. arrais, s. nabhan, g. socie, g. dini, e. gluckman, m the registration study is a joint prospective, multi-center, non randomized trial of ebmt al and pds wps and eurocord. primary aim is to evaluate the contribution of different strategies of treatment (matched sibling donor;msd and an alternative hsct) for adults with al. secondary aim are to compare, in a first step, policy of each centre, feasibility of each transplant modality, time to transplant. in a second step, to compare in a intent to treat analysis lfs, rr and trm according to different transplants strategies. the date of the registration of the patients is considered as the date of hla typing test. for each patient, a specific questionnaire has to be completed specifying the initial strategies and each change in those strategies : at registration, after - months after the resgitration and , each months during the first year after hsct, and twice a year for the following years. the intention to treat will be defined, for each patient by the choice of the treatment planned and reported by the each center and the real modality performed according to the last option, and donor availability. between december and december , adults, enrolled by ebmt centrers, were registered for the study, had aml, all and biphenotipic al. hla typing was performed for patients at diagnosis, after first cr, in primary refractory la, and in more advanced phase ( in cr or more and in relapse). a msd was found for patients. the choice registered for the remaining patients was: alternative donor (search for ud, cb, or haplo); autologous hsct ( pts). chemotherapy alone ( pts).the registration of patients has been closed on december .the follow up of patients will proceed for years. the final analysis will be performed on december . a randomised phase iii study comparing conventional chemotherapy to low dose total body irradiation-based conditioning and haematopoietic cell transplantation from unrelated donors as consolidation therapy for elderly (> y) patients with aml in first complete remission d. niederwieser* on behalf of osho /fhcrc/hovon sakk/austrian/french and alwp-ebmt study objectives: efficacy of allogeneic related and unrelated hematopoietic cell transplantation (hct) after reduced intensity conditioning as a consolidation treatment for patients with aml in complete remission or refractory anemia with excess of blasts (raeb) in comparison with a non-transplant approach. patients with a matched sibling or with an unrelated donor, who have entered cr , will be eligible for randomization in a (transplantation): (non-transplantation) fashion. patients without a donor will receive post-remission therapy without transplantation. blaise, f. frassoni, m. kuenz, b. rio, n. russel, p. rebulla, g. sanz, j. garcia, j. cornelissen, c. navarette, d. niederwieser, a. nagler, g. socié, a. sureda, v umbilical cord blood (ucb) cells from unrelated donors have several advantages as compared to other sources of stem cells for allogeneic use, such as prompt availability, decreased risk of graft-versus-host disease and easy collection with little risk to the mother or to the newborn. ucb has been shown to contain sufficient progenitor cells to provide durable engraftment. however, because of low infused cell doses, single ucb transplantation (ucbt) in larger children and in adults may be associated with delayed engraftment and increased risk of graft failure. transplantation of double cord blood units (ducbt) represents a strategy to overcome this limitation. the results of both pediatric and adult ducbt studies suggest that this approach is safe, and is associated with higher engraftment rates and improved transplant outcome, in both the nonmyeloablative and myeloablative settings, as compared to single ucbt. some preliminary data also indicates a reduction in relapse with double ucb transplantation. in this view, we will discuss a proposal of a randomized trial comparing single versus double ucbt for patients with hematological malignancies. a writing committee has been established to discuss all the aspects of this protocol that will be presented during the alwp session. overall the leukaemia free survivals (lfs) at five years for patients autografted in first (cr ) and second (cr ) remission have been respectively and % in adults and and % in children, with some improvement in outcome after . recent prospective trials from single institutions or national groups have confirmed a lfs of % at years in adult patients autografted in cr . several randomized studies in the pre "high dose ara-c" era had shown superior outcomes of asct over conventional chemotherapy. the only randomized study using hdara-c in the chemotherapy arm (us intergroup study,p cassileth nejm , m slovak blood ) indeed has shown better results for asct in good risk patients by cytogenetics, better results for hdara-c in intermediate risk patients and better results for genoidentical allogeneic transplants in the high risk group. numerous retrospective studies using the ebmt registry have shown equivalence for lfs and overall survival (os) when comparing asct to allogeneic transplantation with unrelated donors and/or allogeneic transplants with reduced intensity conditioning (ric), with a higher relapse incidence (ri) for asct but a reduced transplant related mortality (trm). despite this situation, the annual reports of ebmt activities indicate a clear drop in asct for aml associated to a steady increase of allogeneic transplant activity, using ric and all cell sources including cord blood. the practice of asct itself has changed over the years: less than % of the patients still receive marrow as a source of stem cells and mobilization of stem cells in peripheral blood has become the s standard. total body irradiation (tbi) which has been shown to have the highest anti leukemic effect is used in only % of the patients for conditioning. in vivo purging consisting of chemotherapy consolidation courses given before mobilisation is not standardized, and higher cd + cell yields have been shown to contain leukemic cells and to be associated with a higher relapse incidence (n feller, leukemia ) . a recent retrospective survey of the awlp-ebmt (presented at this meeting) indeed indicates that the number of consolidation courses given before autografting with peripheral blood influences the outcome. in patients autografted with marrow, peripheral blood early (interval cr -transplant< days) and peripheral blood later (> days) , the lfs at years have been respectively , and %. the source of stem cells and the interval from cr to transplant have been significant in multivariate analyses. another recent retrospective survey from alwp-ebmt (nc gorin, jco, in press) has compared asct to allogeneic stem cell transplantation in patients with core binding factor mutations (inv and t( ; )) and shown equivalence in outcome for patients transplanted in cr with lfs around % at years. overall the data accumulate to confirm previous and already old observations that asct remains an interesting alternative therapeutical approach for the treatment of : -older patients -patients considered for ric and/or unrelated donors transplants, with bm,pb or cord blood -chemosensitive aml (so called good risk patients) including thoses reaching cr rapidly (rapid remitters) and those carrying a cbf mutation. since pb has replaced almost totally pb as source of stem cells, for practicality reasons, all effort should be made at increasing in vivo purging aggressively, with, as much as feasible, minimal residual disease monitoring by molecular biology. there is still a need for randomized studies in good and intermediate risk patients, while high risk patients should be allografted whenever possible. in the largest prospective mutlicentre study initiated by the ebmt and comparing filgrastim-mobilized peripheral blood progenitor cell (pbpc) to bone marrow, patients transplanted with pbpc developed more often chronic gvhd. however outcomes, which where overall and disease free survival, relapse and transplant related mortality were similar in both cohorts. at the time of first publication we were aware that long term observation was needed to ultimately determine the role of both source of stem cells. indeed, outcome differences due to difference in the composition of the graft might become apparent only years after transplantation. we therefore planed a follow-up study in order to asses the long term outcomes, general health status, social integration as well as the occurrence of malignant and non-malignant late effects in both treatment groups. the patient accrual of the initial study took place between february and september , including patients transplanted for leukaemia from their hla-identical sibling. in september we sent out a questionnaire to all centres which had participated to the study. the questions included information on survival, cause of death, relapse, secondary malignancies, chronic gvhd and its treatment, the occurrence of non-malignant late effects such as hypothyroidism, bronchiolitis obliterans, sicca syndrome, blood counts at last follow-up as well as question on general health status and social integration. so far, ( %) from the centres responded, including of patients ( %) alive at -year follow-up. ninety-eight of the patients reported patients are alive, and have died after the rd year of follow-up (gvhd n= , relapse n= , other n= ). chronic gvhd is still present in patients ( %), secondary malignancies occurred in cases ( %), non-malignant complications in ( %). the median karnofski score as a measure of general health was > % (range - ). eightyeight ( %) patients have returned to work or to school. during the late effect working party meeting in florence, italy, we will provide preliminary data on the longterm outcome, and compare both patients groups, patients transplanted with pbpc to those who had received bm. this is retrospective multicenter analysis on the incidence and risk factors of cardiovascular events after allogeneic hematopoietic stem cell transplantation (hct), in long term survivors treated in ebmt transplant centers. these patients received hct between and and surviving ≥ year after transplantation. the median age of the patients at last follow-up or at time of a vascular event was years (range - years), and the median follow-up time years ( - years). twenty ( . %) out of patients developed an arterial event in at least one arterial territory. the cumulative incidence of first arterial event years after hct was % ( % ci, %- %). the cumulative incidence for patients with a high global cardiovascular risk score, including arterial hypertension, diabetes, dyslipidemia, physical inactivity and smoking was %, as compared to % in those with a low risk score. in univariate analysis, older age at last follow-up, all cardiovascular risk factors taken individually, and acute gvhd were associated with a higher risk of a cardiovascular accident. in multivariate analysis age older than years at last follow-up, and patients with a high global cardiovascular risk score had a . -fold and . -fold increase, respectively of the relative risk for an arterial event after hct. thus, long term survivors after allogeneic hct are likely to develop cardiovascular risk factors, and present an increased risk for premature cardiovascular accidents after allogeneic hct. update on the phase ii/iii study of the incidence and outcome of vod with the prophylactic use of defibrotide in paediatric stem cell transplantation (vod-df study) s. corbacioglu* university children's hospital (ulm, de) along with graft versus host disease (gvhd) and cytomegalovirus (cmv) infection, veno-occlusive disease (vod) is one of the most frequently encountered serious complications after stem cell transplantation. the currently reported overall incidence of vod ranges from % to more than % in children who have undergone stem cell transplantation. defibrotide (df) is a polydisperse oligonucleotide derived from porcine intestinal mucosa with antithrombotic and protective properties on the microvasculature but minimal hemorrhagic risk. in large, multicenter, international phase i/ii trials targeting patients with severe vod df has emerged as a promising therapy for vod. the vod-df study is a prospective international multicentre phase ii/iii study with the aim to assess the beneficial effect of defibrotide on the incidence, morbidity and mortality of vod in children at high risk to develop vod after myeloablative stem cell transplantation (nih trial number: nct , eudract number: - - ) . the study, co-sponsored by gentium and the ebmt, is open for recruitment since january with currently participating centres in countries. the prospective recruitment period is years with a sample size of patients. the current recruitment status as of january is patients with an average recruitment rate of patients per month. in november the first data safety monitoring board (dsmb) meeting evaluated safety data and mortality rates of the first study patients who completed days of follow-up. as conclusion of this meeting the types of adverse events described were considered typical for this patient population with no unexpected toxicities. no significant difference was observed in the number of adverse events between the prophylaxis arm compared to the control arm. therefore it was unanimously concluded that there were no safety considerations of concern arising from the defibrotide-treated patients on the prophylactic arm at the present time. for the determination of the finale sample size an interim analysis will be performed at patients. the recruitment of this sample size should be completed by early this year. in the meantime the following centres either completed or will complete the initiation process in order to join the study: ankara, antalya, istanbul, and prague. the registration study is a joint prospective, multi-center, non-randomized trial of ebmt al and pds wps in cooperation with the eurocord. primary aim is to evaluate the contribution of different strategies of treatment (msd and alternative hsct) for children with al. secondary aim are to compare, in a first step, policy of each centre, feasibility of each transplant modality, time to transplant. in a second step lfs, according to different transplants strategies, rr, trm will be compared. the date of the registration of the patients is considered as the date of hla typing test. for each step, a specific questionnaire has to be completed: at registration, after - months, at hsct; each months during the first year after hsct, and twice a year for the following years, followup forms will be completed. the intention to treat will be defined, for each patient by the choice of the treatment reported to be planned by the centre in each questionnaire, the real modality performed according to the last option, and donor availability. between december and november , children, enrolled by ebmt centres, were registered for the study. children had all (at diagnosis or in cr , patients; at first relapse or in cr , patients; in more advanced disease), or aml (at diagnosis or in cr , patients; with refractory disease, at first relapse or in more advanced phase, patients). a msd was found for children, the choice registered for the remaining children was: chemotherapy alone ( pts it is estimated that every year in europe about - children become donors of hematopoietic progenitor cells for siblings undergoing allogeneic stem cell transplantation. the aim of the study is the analysis of donor safety and occurrence of early side effects related to bone marrow (bm) or peripheral blood stem cells (pbsc) collection in pediatric siblings qualified to be a donor. the study is based on questionnaires send to transplant centers. following endpoints are analyzed: complications during anesthesia, complications related to bm collection, complications of catheter placement, complications related to apheresis, number of nights spent in hospital and psychological issues in donor immediately after stem cell collection and during one year follow-up. since january , donors (median age years, range months - years) were registered. the source of stem cells was pbsc and bone marrow, in and patients, respectively. preliminary data indicate that the rate of complications is very low, thus the procedure is safe for donors. due to low patients accrual, retrospective donors registration over a period of year before the start of the study is being proposed. the th meeting of the ebmt pediatric diseases working party will be held june - , in poznan (poland) and for the first time will be accompanied by the meeting of ebmt pediatric nurses. the second announcement with detailed information concerning preliminary scientific programme, call for abstracts, registration, hotel accommodation, travel, important dates etc. is available on the meeting web page www.bokiz.pl/ebmt-pds-wp- , which is also accessible using the link from the ebmt pds wp web page. physicians sessions (monday and tuesday, june - ), nurses sessions (tuesday, june ), and joint session (wednesday, june ) will create an excellent opportunity to summarize recent progress in the field of hsct from pediatric point of view, to initiate new ideas, and to improve an understanding and collaboration between nurses and physicians involved in pediatric hsct. apart from that the meeting will be a unique platform for initiation new trials, interaction, and cooperation. more than outstanding speakers will present various aspects of topics of the meetings. however, organizers expect also a lot of interesting original oral as well as poster presentations, and an exciting and stimulating discussion. therefore, all participants are invited and encouraged to submit their own experimental and clinical results for presentation during the meetings. the deadline for abstract submission is monday, march , . early registration is possible until monday, march , . twenty grants from the ebmt covering registration fees and local accommodations will be awarded by the scientific committee on a competitive basis to physicians or nurses (information concerning application for grants is available on the meeting web page). pharmaceutical companies, manufactures of technical equipment and software as well as publishers are invited to display their products at the industrial exhibition which will part of the meeting. detailed information about poznan, including information how to get to poznan, is available on the web page www.city.poznan.pl. it is a real honor and a great pleasure to invite everybody interested and involved in pediatric hsct to poznan to attend the th meeting of the ebmt pediatric diseases working party and st meeting of the ebmt pediatric nurses, poznan (poland), june - , . why is important to built a paediatric nurse group inside pds wp? s. calza* ( ), v. van de crommert ( ) ( )children's hospital (genoa, it); ( ) transplantation group initiated a prospective, randomized, parallel-group, open-label phase iii, multicenter study, comparing vtd (arm a) with td (arm b) for mm patients progressing or relapsing after autologous transplantation. inclusion criteria were: patients in first relapse after at least one autologous transplantation, including those who may have received velcade or thalidomide before transplant. exclusion criteria: subjects with severe neuropathy or non secretory mm. patients will participate ( in each arm). primary study end point was time to progression. secondary end points included toxicity, response rate, eventfree survival and overall survival. treatment was scheduled as follows: velcade . mg/m² will be given as an i.v bolus on days , , and followed by a -day rest period (days to ) for cycles ( months) and then on days , , , followed by a -day rest period (days to ) for cycles ( months). in both arms, thalidomide will be given at mg/day per os for one year and dexamethasone mg/day per os four days every three weeks for one year. thrombosis prophylaxis was strongly recommended as well as valacyclovir prophylaxis (in arm a) against reactivation of varicelle zoster virus. patients reaching remission could proceed to a new stem cell harvest. however, transplantation, either autologous or allogeneic, could only be performed after achieving the one year treatment. response was assessed by ebmt criteria, with additional category of ncr. adverse events are graded by the nci-ctcae, version . . as of january , , patients entered the study. in france (ifm - study), in italy, in germany, in switzerland (a sakk study), in belgium, in austria and the czech republic, in the uk. are assessable: males, females; median age: yrs (range - ), number of autologous transplant: one: , two: . of these patients, were randomly assigned to receive vtd and to receive td. treatment was discontinued in patients. an interim toxicity analysis was planned to be performed when the first hundred patients had been included. this interim analysis is ongoing (january ). other preliminary studies demonstrate that vtd is a highly active and relatively well tolerated regimen. the combination is used in the relapse setting, as well as first line, consolidation and maintenance. in this protocol, the starting doses of velcade and thalidomide are relatively high and the duration of treatment is long. we will assess the superiority of vtd over td in the relapse setting. protocol eu-dract number: - - . tyrosine kinase inhibitors (tkis) are the accepted first line treatment of choice for the majority of patients with chronic myeloid leukaemia (cml) although allogeneic stem cell transplantation (sct) remains the only potentially curative procedure. although second generation tkis have shown promising results, in young patients resistant or intolerant to imatinib and with a low ebmt score allogeneic sct is still considered the best second line therapy. however, despite its curative potential, a significant proportion of patients relapse following allogeneic sct. the gold standard for the treatment of relapse following allogeneic sct is the infusion of lymphocytes from the transplant donor (dli). however, dli may result in significant complications including graft versus host disease and bone marrow aplasia and using escalating doses protocols, responses may be protracted. there is consequently a need for an alternative treatment for patients that relapse following sct, that is both efficacious, faster acting, easier to administer and safer. dasatinib has been shown to be effective in treating patients that are resistant or intolerant to imatinib and as a result constitutes a good candidate treatment option for imatinib intolerant or resistant patients that relapse following sct. the cml sub-committee has launched a prospective phase ii to investigate the efficacy of dasatinib in this setting. this study will concentrate on patients years of age with ph+ cml whose disease has relapsed after transplantation from an hla-identical sibling or an hla-matched unrelated donor and have not responded to withdrawal of immunosuppressive treatment where this is possible. enrolled subjects will be commenced on dasatinib mg qid and receive treatment for months. continuation of treatment beyond months will be at the investigator's discretion. the primary end-point will be complete molecular remission at months secondary end-points will include complete cytogenetic remission rates, overall survival and proportion of patients requiring dli. donor lymphocyte infusions will be administered to all patients in whom dasatinib has been discontinued indefinitely or there is evidence of disease progression during dasatinib therapy or there is evidence of disease relapse after initial response tom dasatinib. the study will open in the summer of and will aim to recruit patients in years. interested ebmt centres are invited to participate. ( ) allogeneic hematopoietic stem cell transplantation (hsct) as first line therapy for patients with chronic myeloid leukaemia (cml) has been replaced by imatinib. the role as second line therapy in patients who failed imatinib treatment is a matter of debate. second generation tyrosine kinase inhibitors (tki) have already proven their efficacy in this setting. early transplant related mor¬tality of allogeneic hsct is considered to be too high. however, transplant outcome of young cml patients with a low risk for transplant related mortality has not been analyzed recently. method: in order to better counsel patients with a hlaidentical sibling confronted with this situation we performed a retrospective analysis of transplants reported to the ebmt be¬tween and . we selected for patients who had a low risk (ebmt-) score for transplant related mortality and who were transplanted from an hla identical sibling. we analysed the outcome of those only who were transplanted in st chronic phase and who received best current treatment, defined as standard conditioning, no t-cell depletion and bone marrow as stem cell source. results: patients ( % of all patients transplanted for cml in this time period) with a median follow up of months ( - months) who fulfilled these criteria were identified. they were % males and % females with a medium age of years (range to years). % ( patients) were less than years old and % were above the age of . the time interval from diagnosis to transplantation was less than year in % of patients. about one third each had an ebmt risk score , or . data were obtained from teams in countries. the probability of survival at years in a competing risk model was % ( % c.i. - ) with a cumulative incidence of death without relapse of % at months and no additional death from transplant related mortality thereafter until months of follow-up ( figure ). -years post hsct, % of patients were estimated to be alive after relapse (and hence the relapse free survival was %). conclusions: these results show the current transplant outcome which is achievable by selecting only patients with a low risk for transplant related mortality. in this context, the data shown is valid even without information on pre-and/or posttransplant therapy. allogeneic hsct is a valuable option as second line therapy after imatinib failure for cml patients with a low transplantation risk. t prolymphocytic leukemia (t-pll) is a rare, aggressive neoplasia of t lymphoid lineage which is characterized by poor survival of less than one year. incidental reports suggest that both autologous and allogeneic hematopoietic stem cell transplantation (hsct) might be effective in this disease. a comprehensive retrospective analysis of patients registered in the ebmt database will be a subject of independent presentation at this meeting. however, given the limitations of conventionally collected registry data (dubious follow-up information and extreme heterogeneity), and realizing the impossibility of performing an international formal prospective trial under the current regulatory framework, we developed a new concept of two complementary projects: the first is called "ebmt prospective observational audit on allogeneic and autologous transplantation in t-pll" and means that transplant centers will be encouraged to register their patients with t-pll very timely with the ebmt, followed by mandatory submission of ebmt medb and follow-up forms. the second is the "ebmt/eln frame of orientation for allogeneic and autologous transplantation in t-pll". its purpose is mainly to avoid transplants in situations where they are very unlikely to be successful and to avoid excess heterogeneity of eventual transplants performed, thereby facilitating scientific analysis. this expert opinion-based framework covers criteria for the diagnosis of t-pll, transplant eligibility, pre-transplant remission induction strategies, remission requirements, timing of hsct, donor compatibility criteria, conditioning, gvhd prophylaxis, and mrd monitoring. with these two complementary components it should be possible to largely improve the usual quality of registry-based data and to generate scientifically sound knowledge on hsct in an orphan disease such as t-pll. primary plasma cell leukaemia (pcl) is a rare variant of plasma cell dyscrasia, associated with poor prognosis, median survival - months, significantly shorter than for multiple myeloma. treatment with alkylating agent therapy is ineffective though polychemotherapy may offer modest improvement in survival. autologous transplantation is now widely used in the treatment of pcl and this report summarises the european blood and marrow transplant (ebmt) experience. a retrospective study was carried out of patients with common type myeloma ( % igg, % iga and % light chain) and patients with pcl undergoing first autologous transplant between and . all patients were reported using med-a (limited data set) or med-b (extensive data set) forms and included in the study regardless of availability of complete data. comparisons used the chi-squared test for categorical data and mann-whitney test for continuous data. overall survival and progression-free survival were calculated using the kaplan-meier method and comparisons made using the log-rank test. relapse/progression and death without relapse or progression were computed by the proper non-parametric estimator for outcomes with competing risks and compared by the gray test. there was no significant difference in age, gender, calcium or albumin at presentation. haemoglobin was significantly lower ( g/dl v g/dl, p= . ), creatinine significantly higher ( micro mol/l v micro mol/l, p= . ) and b microglobulin significantly higher in the pcl group. there was no difference in graft type or use of total body irradiation but the pcl group was transplanted closer to diagnosis ( . v . months, p= . ). while no significant difference in engraftment, pcl patients were more likely than myeloma patients to enter cr post-transplant. overall survival for pcl patients was greatly inferior to myeloma patients - . (ci . - . , p= . ) v . months (ci . - . ), attributable to response of short duration and increased relapse-related mortality. this is the largest reported study of pcl patients and suggests improved outcome with use of autologous transplantation. it is however dispiriting to note that outcome is greatly inferior to that in myeloma despite likely pre-selection for fitness of the pcl group. there is urgent need for collaborative study of alternative approaches including highly effective induction with novel agents and optimal stem cell transplant strategy. myeloma patients from centres that had undergone hla typing were included in the trial. study inclusion was at the time of conditioning for first autologous transplant at the achievement of a response status of at least stable disease after vad-like induction treatment of previously untreated patients. patients with an hla-identical sibling were allocated to the auto+allo (aual)-arm (n= ) and patients without a matched sibling donor to the auto (au)-arm (n= ); single or tandem autografting was optional. conditioning for asct was melphalan mg/m , and for allo ric was fludarabine mg/m x plus tbi gy. the accrual period was from february to february . the two treatment groups were well matched for the standard prognostic parameters such as beta- -microglobulin, karyotype, gender, mm subtype, stage, albumin, creatinin, calcium and response status at transplantation. median age at transplantation was significantly higher in the au-arm ( vs years). the cr rate was % in the aual-arm and % in the au-arm (p= . ). cumulative months non-relapse-mortality was % in the aual-and % in the au-arm (p= . ). at years after transplantation, there was no significant difference between the treatment arms with respect to os (aual %, au %), rfs (aual %, au %) or relapse rate (aual %, au %).. however, looking at the os curve for all patients in the aual-arm, a survival plateau on the %-level seems to be emerging from years and onwards. we conclude that no significant differences in outcome was observed in this early analysis, but longer follow-up is warranted before any definite conclusions can be drawn. updated results will be presented. allogeneic stem cell transplantation (sct) can cure patients with mds or aml. the major disadvantage of allogeneic stem cell transplantation is the high treatment related mortality. recently the introduction of dose-reduced conditiong followed by allogeneic stem cell transplantation has lowered the treatment related mortality in comparison to standard conditioning, but a prospective comparison between both approaches is lacking. the subcommittee mds of the clwp launched a multicenter, prospective phase iii-study comparing dose-reduced versus standard conditioning followed by allogeneic stem cell transplantation from related or unrelated donors in patients with mds or secondary aml. the primary endpoint is treatment related mortality at one year. the hypothesis is that a dose-reduced conditioning will reduce the non-relapse mortality from % to % at one year after allogeneic stem cell transplantation. a total of patients is needed to achieve this goal. patients should have mds or saml (less than % blasts) and should be eligible for standard and dose-reduced conditioning and aged - years if donor is a hla-matched unrelated donor (hla-a, hla-b, hla-drb and hla-dqb ) (one mismatch allowed)and aged - years if donor is a hla-matched related donor ((hla-a, hla-b, hla-drb and hla-dqb ) (one anti¬gen-mismatch allowed). the patient will be randomised between a dose reduced conditioning (arm b) and a standard conditioning (arm a). the standard conditioning (arm a) consisted of busulfan ( mg/kg bw orally or . mg/kg intravenously) and cyclophosphamide ( mg/kg) and reduced conditioning consisted busulfan ( mg/kg bw orally or . mg/kg intravenously) in combination with fludarabine ( mg/m²). so far the protocol is activated in germany, the netherlands, russia and italy. detailed protocol as well as recruitment will be presented. a randomised trial of rabbit anti-thymocyte globulin, given on day+ after alternative donor transplants a. bacigalupo*, f. ciceri, p. di bartolomeo, t. lamparelli, g. milone on behalf of the gitmo background: transplant mortality (trm) can be predicted by using laboratory values, on day+ after an allogeneic hemopoietic stem cell transplant (hsct) (bone marrow transpl. ; : ) : increased mortality is mainly, but not exclusively, due to increased acute graft versus host disease (gvhd). in a pilot study, a low dose of rabbit anti-thymocyte globulin (atg) given on day+ reduced gvhd and trm. aim of the study: the aim of this study was to test in a propsective randomized trial, whether intervention on day+ in patients with a high risk score would reduced the risk of trm and gvhd. patients: eligible were patients undergoing hsct from family hla mismatched (n= ), or unrelated donors (n= ). all patients received atg . mg/kg x pre-transplant, were stratified for intermediate and high risk day+ score, and were randomized to receive an additional dose of ratg ( , mg/kg day + and day+ ) (n= ) or no additional treatment (n= ) (control arm). the two groups were balanced for age, disease phase, day+ score, and donor/recipient sex mismatch. results: the predictive value of day+ score on trm was confirmed in the control arm ( % vs % for intermediate and high risk patients, p= . ), whereas in the day+ atg arm , there was no difference ( % vs %, p= . ). trm was overall reduced from % in the control arm to % in the atg day+ arm (p= . ) : the difference was more pronounced in patients with early disease and high risk on day ( % vs % p= . ), and in hsct with female donors in male recipients ( % vs %, p= . ). acute gvhd grade iii-iv was reduced overall from % to % (p= . ), and chronic gvhd was reduced % to % (p= . ). conclusions: we confirm that patients with different risk of trm can be identified on day+ after hsct: in patients at greater risk of trm, the administration of atg on day+ reduces gvhd and trm. additional intensified supportive care (including anti-infectious treatment ) may further reduce trm in patients with a high risk score on day+ . our group reported a strong association of polymorphisms (snps) within the antibacterial defense receptor nod /card with severe gvhd following allogeneic sct. however, functional studies explaining these effects were so far missing. therefore, we analysed gastrointestinal biopsies from controls, patients (pts) prior to sct and pts following sct. biopsies post sct were obtained at the time of first symptoms indicating gastrointestinal gvhd. slides were evaluated microscopically for occurrence of apoptotic cells, loss of crypts and infiltration with lymphocytes, eosinophils and neutrophils. in addition, immunohistochemical analyses for cd , cd , cd , mib and cd expression were performed. all pts and donors were typed for presence or absence of nod /card snps. semiquantitative histological results were compared with clinical parameters such as stage of gvhd, use of corticosteroids and nod /card status. comparison of controls with pts post-transplant showed a significant increase of apoptotic cells / crypt loss associated with enhanced lymphocyte and neutrophil infiltration. whereas the number of cd cells in the lamina propria significantly increased after sct, cd cell numbers were strongly diminished. within post-transplant biopsies, loss of crypts (score . in gvhd - , . in gvhd and . in gvhd / ), changes in neutrophil infiltrates (score . in gvhd - , . in gvhd and . in gvhd / ) and reduction of cd infiltrates (score . in gvhd - , . in gvhd and . in gvhd / ) were clearly correlated with stages of gastrointestinal gvhd, whereas cd cells showed an increase and cd positive cells were unchanged in pts with severe gvhd. presence of nod /card snps themselves resulted in a significant reduction of neutrophils (p . ) and cd cells (p . ) but had no impact on further parameters. this effect could not be explained indirectly by more severe gvhd in pts with nod /card snps and was confirmed in multivariate analysis. our data indicate for the first time functional changes in gastrointestinal biopsies from pts after allogeneic sct in relation to the nod /card genotype. the observed reduction of neutrophils and cd cells may result from a reduced expression of chemokines attracting these cells to inflammatory sites, as il- production is strongly regulated by nod /card dependent activation of nf-kappab. presence of cd cells and neutrophils may be required to prevent dysregulated inflammation. introduction: the nih staging and response criteria offer for the first time criteria for standardized diagnosis and staging of severity as well as evaluation of physical functioning and quality of life (qol) of chronic graft-versus-host disease (cgvhd). we present the interim analysis of a prospective germany multicenter validation study on the nih staging criteria in cgvhd. methods: patients (median age years, range - ) after allogeneic hematopoetic stem cell transplantation (hsct) for hematologic malignancies were evaluated according to the nih criteria based cgvhd activity assessment, the lee cgvhd-symptom-scale, fact-bmt, human activity profile (hap), sf , berliner social support scale (bsss), item adjective measure ( -am), hospital anxiety and depression scale (hads) and the nccn-distress-thermometer. enrolment occurred between day and year after hsct or in the presence of active cgvhd without time limit. follow-up surveys were conducted at , , , , , and months after baseline survey. at all time points disease status, co-morbidities and medication were documented. results: sixty five patients had cgvhd (mild n= , moderate n= , severe n= ) while patients did not have cgvhd. the comparison of the severity grading (mild-moderatesevere) of the physician and severity grading of the patient revealed a high correlation (p< . , r=. ), while the comparison of point scale of patient and physician revealed differences between patient and physician in the range of - points (physician), where patients graded more severity compared to physicians. the cgvhd nih consensus grading correlated inversely with fact physical well being (r=. , p < . ). the hap maximum activity score correlated inversely with severity of cgvhd (p< . , r=. ). the cgvhd symptom scale summary score correlated with physician severity grading (r=. , p<. ), the fact-g score (r=. , p< . ), mental health (r=. ), energy and vitality (r=. ) and hap maximum score (r=. , p < . ). beside fact physical and functional well being the hap maximum score was independent of other aspects of qol. discussion: the results demonstrate, that severity of cgvhd as assessed by the nih consensus grading correlates with impairment of physical well being as well as daily activities and qol. since the cgvhd symptom scale covers severity of cgvhd and aspects of qol it should be applied together with the hap in clinical routine. naturally occurring regulatory t cells (tregs) have been reported to play an important role in modulating graft-versushost disease (gvhd), a major complication after allogeneic haematopoietic stem cell transplantation. despite striking findings in animal models supporting the therapeutic use of tregs in gvhd, the data from human studies is limited and their mechanism of action remains elusive. in this study, treg modulation of cd + lymphocyte induced in situ graft-versushost reactions (gvhr) was evaluated using a unique in vitro human gvhd model. tregs were defined as cd +cd hifoxp + and isolated from buffy coat of healthy blood donors using robosep following rosettesep enrichment of cd + cells (stemcell technology). isolated tregs were expanded in vitro with anti-cd cd mab coated dynabeads (invitrogen) prior to use. the alloreactive immune reactions were set up by co-culturing "donor" cd + lymphocytes with hla unmatched allogeneic "recipient" monocyte derived dcs (mo-dc) in the absence or presence of "donor"-derived tregs ( : ratio for treg: cd + lymphocytes) for - days. following magnetic depletion of mo-dc and tregs, allo-antigen stimulated "donor" cd + lymphocytes were co-cultured for days with "recipient" skin tissue. the severity of histopathological changes in skin tissue was scored as grades i-iv according to the lerner gvhd grading system. in out of experiments the presence of tregs significantly reduced the severity of skin gvhr from grade iii to grade i. the levels of ifng, tnfa and il- cytokines in the supernatants from the primary co-culture of allogeneic mo-dc and cd + lymphocytes were significantly reduced in the presence of tregs (ifng: pg/ml vs . pg/ml, p< . ; tnfa: . vs . , p= . and il- : . vs . , p= . ). following allogeneic mo-dc stimulation there was a . and . fold reduction in the percentage of cd + lymphocytes expressing the activation marker cd + ( . vs . , p= . ) and intracellular ifng ( . vs . , p= . ) in the presence of tregs. cd + lymphocyte proliferation measured by h-thymidine incorporation and cfse dilution was found to be markedly suppressed ( %- % inhibition) in the presence of tregs in alloreactions. further investigations are underway to explore the mechanisms and characterise the modulation of gvhr by tregs in an allo-antigen specific setting. donor-recipient hla class i ligands and kir-haplotype a are associated with severe acute graft-versus-host disease in unrelated haematopoietic stem cell transplantation for beta-thalassaemia r. littera ( ) killer immunoglobulin-like receptors (kirs) regulate the activity of human natural killer cells, mainly through recognition of hla class i molecules. two broad haplotypes of kir genes have been defined. the a haplotype is characterised by a single activating kir gene ( ds ), whereas the b haplotype is characterised by two or more activating kir genes ( ds , ds , ds , ds and ds ). many studies have investigated the impact of kirs and their ligands on hematopoietic stem cell transplantation (hsct) in patients affected by acute myeloid or acute lymphoblastic leukemia. however, the results of these studies remain controversial. allogeneic hsct in talassemia patients offers an ideal study model since this cohort of patients is not biased by the variability of conditioning regimens and the different clinical and immunologic characteristics of patients transplanted for oncohematologic disorders. we studied thalassemia patients transplanted from an unrelated donor. the conditioning regimen was the same in all patients. donor and recipient pairs were typed for the hla-a, b, cw, drb , drb , drb , drb , dqa , dqb and dpb loci using high resolution molecular typing techniques. kir genes were typed using kir-gene-specific primers. out of transplanted patients, are alive and well (disease-free survival . %), rejected and died. twentytwo patients ( / - . %) developed acute graft-versushost disease (agvhd). in of these patients agvhd was grade iii-iv. patients who were heterozygous for hla-cw groups (hla-cwasn ) and (hla-cwlys ) had a higher risk of developing acute gvhd than c /c or c /c homozygotes ( / vs / ; rr= . ; % ci: . - . ; p=. ). conversely, / patients who rejected were c /c or c /c homozygotes (rr= . ; % ci = . - . ; p=. when compared with heterozygotes). these findings confirm the results of a previous study performed on a cohort of thalassemia patients. in the present study, of the patients ( . %) with severe grade iii-iv agvhd had been transplanted from donors who were homozygous for kir haplotype a (rr= . ; % ci: . - . ; p=. ). in conclusion, it would seem that c /c heterozygosity associated with donor homozygosity for the a haplotype is likely to favour donor alloreactivity and thereby increase the risk of severe gvhd. analyses of these genetic markers may help modulate conditioning regimens and the intensity of gvhd prophylaxis in patients undergoing unrelated hsct. interleukin- (il- ) is an immunoregulatory cytokine secreted predominantly by activated t-helper (th ) cells. it suppresses the cytotoxic action of macrophages, inhibits the production of pro-inflammatory cytokines and is a central mediator of allergic inflammation. a single nucleotide polymorphism (snp) exists within exon of the il gene at position + . the a allele of this snp and high il- levels have been linked with several inflammatory conditions and il- mixed lymphocyte culture (mlc) levels have been associated with graft-versus-host disease (gvhd) following haematopoietic stem cell transplantation (hsct). consequently the roles of the il + snp and il- mlc levels in hsct were examined in this investigation. polymorphism studies were carried out on a cohort of hsct recipients and donors from transplant centres across europe. il genotyping was performed using pcr and rflp analysis. il- levels were measured in a cohort of mlc supernatants using a cytometric bead array and correlated with gvh reaction (gvhr) grades from an in vitro model of gvhd. in all statistical analyses p values < . were regarded as being significant. multivariant analysis of the whole hsct cohort demonstrated that the il + a allele was significantly associated with the development of both acute (grades iii-iv) and chronic gvhd (p= . and p= . respectively). these associations remained significant when the cohort was stratified for transplant type and conditioning regimen. significant associations were also observed in a subset of patients diagnosed with cml; in hla-matched siblings possession of the il + a allele was linked with a decreased susceptibility to chronic gvhd, whereas in mud transplants possession of the a allele was a risk for chronic gvhd. depending on the subset analysis, several clinical factors were also significantly associated with gvhd. analysis of the mlc data demonstrated that il- levels increased with gvhr grade, with significantly higher levels being observed in mlc supernatants with gvhr grades iii-iv (p= . ). to our knowledge this is the first investigation examining the roles of both il- mlc levels and the + snp in hsct. the findings are encouraging, indicating that il- may be involved in the immunopathology of gvhd. consequently, il- levels, as well as snp analysis could provide key pretransplant information on gvhd prognosis and be potential novel targets for post-hsct gvhd therapy. allogeneic hematopoietic stem cell transplantation (allo-hsct) is a curative treatment for hematologic malignancies, but the application is slimited due to major complications, such as severe graft versus host disease (gvhd). diagnosis of acute gvhd is based on clinical features and biopsies, a proteomic pattern specific for agvhd has been published and evaluated blindly on samples collected from patients undergoing allo-hsct at mhh between and . the majority of the patients included were transplanted for hematological malignancies (n= ), for hematopoietic failure syndromes (saa, n= ; pnh n= ; omf, n= ). forty-five patients were treated with dose-reduced conditioning regimens; gvhd-prophylaxis consisted of cyclosporin (csa) plus methotrexate (mtx) or mycophenolic acid (mmf), and antibodies respectively. most patients were transplanted from matched unrelated donors (mud, n= ), received stem cells from matched related (mrd, sib, syngeneic), from haplo-identical related, and from mismatched related donors. based on the positivity of the agvhd pattern we initiated a pilot trial of pre-emptive therapy, treating patients upon pattern positivity with mg prednisone/kg bw and compared the outcome of the treated group to those patients who did not receive pre-emptive therapy. in patients were transplanted and screened for agvhd at mhh, but not pre-emptively treated. forty one patients developed agvhd ( %), ( %) had agvhd >ii and were treated with standard protocols. eight patients developed agvhd grade iii or iv ( / = %) and of these died. between april and june patients were transplanted at mhh. in patients the agvhd proteomic pattern showed a clear correlation for gvhd >ii. twenty six ( %) of transplanted patients had agvhd >ii, received pre-emptive therapy, while were treated upon clinical signs of agvhd according to standard treatment protocols.two patients in the preemptive treatment group ( / = . %; of pre-emptively treated: %) had agvhd iii or iv and died ( / = . %; / : %). of the standard therapy patients developed agvhd grade iii or iv and to date of these have died ( / = % or / of the standard therapy group= %). thus, taken together our results indicate that pre-emptive treatment may decrease the severity of agvhd and probably leads to a better overall survival. donor-derived t cells emigrating from the graft after solid organ transplantation have been shown to promote immunological tolerance thereby improving long-term graft survival. however, donor t cells are also able to induce lifethreatening allo-reactive graft-versus-host disease (gvhd) in the transplant recipient. the exact mechanism by which donor t cells influences this delicate balance between tolerogenicity and allo-reactivity has not been elucidated. we observed two liver transplant patients with severe gvhd who developed a donor t-cell chimerism in peripheral blood and bone marrow up to %, which is far above those of previously described cases. this enabled us to isolate donorderived t cells in sufficient numbers to allow for a detailed analysis of phenotypic and functional features ex vivo. we found that the donor t cells died by apoptosis over time without any evidence of rejection by host t cells. interestingly, the host-versus-donor reactivity appeared to be selectively impaired, as anti-viral t cells were still detectable in the host repertoire. these results indicate that graft t cells are able to specifically eliminate donor-reactive t cells from the host repertoire thereby preventing donor cells from subsequent rejection. since substantial donor t-cell chimerism persisted in both patients beyond resolution of gvhd, we investigated potential mechanisms of immunotolerance. we observed that the recovery from gvhd was not accompanied by an expansion of immunosuppressive cd /cd /foxp -positive t cells in peripheral blood. however, we obtained formal evidence that host-reactive donor t cells were controlled by an alternative negative regulatory pathway, executed by the immunoinhibitory receptor programmed death- (pd- ) and its ligand pd-l . we did not only find an exceptionally high level of pd- expression on host-reactive donor t cells ex vivo, but also discovered that blocking pd-l on host cells significantly enhanced anti-host reactivity by donor cd t cells in vitro. we thus suggest the interference with the pd /pd-l pathway as a novel therapeutic strategy to control host-reactive donor t cells in solid organ transplant-associated gvhd. our observations might be also of relevance for other clinical scenarios of misdirected allo-reactivity, such as graft rejection as well as severe gvhd after allogeneic hematopoietic stemcell transplantation. ( ) purpose: to determine risk factors of outcomes after umbilical cord blood transplantation (ucbt) for patients with advanced lymphoid malignancies. patients and methods: we evaluated adult patients (median age, years) who underwent unrelated donor ucbt for lymphoid malignancies. ucb grafts were antigen hla mismatched in %, and were composed of one (n= ) or two (n= ) units, with a median cell dose of . x nucleated cells/kg and . x cd cells/kg. diagnoses were non-hodgkin lymphoma (nhl, n= ), hodgkin lymphoma (hl, n= ), and chronic lymphocytic leukemia (cll, n= ), with % having advanced disease and % having failed a prior autologous transplant. sixty-four percent of patients received a reduced-intensity conditioning regimen and % low-dose total body irradiation (tbi). median follow-up was months. results: cumulative incidence (ci) of neutrophil engraftment was % by day , with greater engraftment in recipients of higher cd +/kg cell dose ( % vs. %, p= . ). ci of non-relapse-related mortality (nrm) was % at year, with a lower risk in patients treated with low-dose tbi ( % vs. %, p= . ). ci of relapse or progression was % at year, with a lower risk in recipients of double unit ucbt ( % vs. %, p= . ), and those with chemosensitive disease ( % vs. %, p= . ) and indolent nhl ( % vs. %, p= . ) . the probability of progression-free survival (pfs) was % at year, with higher survival in those with indolent nhl ( % vs. %, p= . ), chemosensitive disease ( % vs. %, p= . ) and who received low-dose tbi ( % vs. %, p= . ). conclusion: ucbt is a viable treatment for adults with advanced lymphoid malignancies. diagnosis of indolent lymphoma, chemosensitive diseases, and use of low-dose tbi and were factors associated with significantly better outcome. positron emission tomography scan performed before reduced-intensity conditioning allogeneic stem cell transplantation has a prognostic value in patients with relapsed and chemosensitive hodgkin's lymphoma or aggressive non-hodgkin lymphoma a. dodero* ( ), r. crocchiolo ( ) ( ) ( positron emission tomography (pet) scan using fluorodeoxyglucose [ f-fdg] has a recognised prognostic value in patients (pts) with hodgkin lymphoma (hl) or aggressive non-hodgkin lymphoma (hg-nhl) receiving chemotherapy or autologous stem cell transplantation (sct). thus, we retrospectively assessed the prognostic role of pet scan before reduced-intensity conditioning allogeneic sct. between and , consecutive pts with hg-nhl or hl, responding to salvage therapy, were evaluated with a pet scan before allografting. presence (pet-pos) or absence (pet-neg) of abnormal f-fdg uptake was correlated to progression-free survival (pfs) and overall survival (os).interpretation of pet scan was obtained with visual assessment alone by a nuclear medicine physician (evaluation of maximal suv in pet-pos cases is ongoing). median age of pts was years (range, - years). histologic subtypes included: hg-nhl [b phenotype (n= ), t phenotype (n= ), other (n= )] and hl. fortyseven pts ( %) were allografted from a hla-identical sibling donor, from a haploidentical donor and from an unrelated donor. sixty-eight pts ( %) failed autograft, the median number of prior regimens was (range, - ). pet scans were performed at a median of days prior to allograft: out of pts were pet-pos [hg-nhl (n= ), hl (n= )] whereas were pet-neg [hg-nhl (n= ), hl (n= )]. pts with pet-pos or pet-neg scans were well balanced in terms of diagnosis, previous treatments, and type of donor. at a median follow-up of months (range, - months), pts are alive and died [toxicity n= , disease n= ]. overall, the estimated -year pfs in pts with pet-neg or pet-pos scans were % ( % ci, % - %) versus % ( % ci, % - %), respectively (p< . ). for hg-nhl pts, the estimated -year pfs was % for pet-neg as compared to % for pet-pos (p< . ) whereas for hl pts, the estimated -year pfs was % as compared to %, respectively (p= . ). a statistically significant higher cumulative risk of relapse was observed in pts with pet-pos scan before allograft as compared to the pet neg scan ( % versus %, p< . ). the estimated -year os in pts with neg or pos pet scans were % ( % ci; % - %) versus % ( % ci; %- %), respectively (p< . ). our study shows a better pfs and os for pts being pet neg before allografting. pet scan should be incorporated in pretransplant work-up to validate our findings prospectively. l. rigacci*, a. bosi, b. puccini, p. corradini, l. castagna, n. cascavilla, g. milone, a. bacigalupo, r. scimè, g. specchia, a. rambaldi, p. leoni, f. ciceri, a. levis, s. guidi, b. bruno, r diffuse large b cell lymphoma (dlbcl) is the most common lymphoid malignancy in adults. autologous hematopoietic stem cell transplantation (ahsct) has been shown to be an effective therapy for patients with dlbcl who relapsed after complete remission (cr). patients who relapse after an ahsct have a very poor prognosis and usually can not be cured with standard or high dose chemotherapy. allogeneic hematopoietic stem cell transplantation (allohsct) has shown to be effective in the rescue of lymphoma patients relapsed after conventional or high dose therapies. according to this data we have analysed all patients with diagnosis of dlbcl who have performed an allohsct from to after an ahsct relapse. seventy-five patients were selected from the data-base, were male ( %), presented a diagnosis of dlbcl and were anaplastic large cell lymphoma. the stem cell donor was related in patients ( %) and unrelated in patients ( %). the stem cell source was peripheral blood in cases and bone marrow in cases. the conditioning regimen was conventional in patients and reduced intensity in patients. the median time between ahsct and allohsct was months (range - months). twenty-five patients ( %) performed allohsct after the obtainment of at least a partial remission or a controlled disease, ( %) were treated with active disease and in cases the data was not available. after allohsct % of patients obtained a response (cr or pr) and did not have evidence of disease, % of patients did not respond and progressed. the treatment related mortality (trm) was %, out ( %) patients died in conventional regimen and out ( %) in reduced intensity arm. acute graft versus host disease (agvhd) was observed in patients (grade iii-iv in patients), and chronic (cgvhd) in %. after a median follow-up of months from the diagnosis (range - months) and a median follow-up of months after allohsct (range - months) the overall survival was %. the overall survival was significantly higher in patients treated with reduced intensity conditioning in comparison with patients treated with conventional conditioning (p: . ). this retrospective study confirms that allo-transplant it is feasible and it could be really effective in a poor prognosis group of patients. moreover the use of reduced intensity conditioning improves these results. ( ) median time from diagnosis to mud-sct was months (range, - ). % of the patients had failed previous autologous transplant (asct), and % were transplanted with chemorefractory disease. peripheral blood was the source of hematopoietic stem cells in % and reduced intensity conditioning regimens (ric) were used in % of the cases. after a median follow up for living patients of months, the estimated -year non-relapse mortality (nrm), relapse rate (rr), progression free survival (pfs) and overall survival (os) for the whole series were %, %, % and . %, respectively. grade ii-iv acute graft-versus-hostdisease developed in % of patients. patients selected for ric protocols were older (median age of years vs years, p = . ) and more heavily pre-treated; % had failed autograft compared with % in the conventional conditioning(cc) group (p = . ). despite these unfavorable factors, nrm for patients receiving ric was significantly lower than observed in patients treated with a conventional regimen: % vs % at years (p = . ). however, this advantage was offset by an increased rr in patients undergoing ric-mud ( -yr rr: % vs %, p = . ), resulting in a very similar pfs and os for both types of conditioning regimens. the prognostic factor with highest impact on pfs was refractory disease at transplantation (rr = . ; %ci . - . , p = . ). patients transplanted with chemosensitive disease had a yr pfs of % irrespective to the conditioning regimen applied, whereas patients transplanted with chemorefractory disease had a year pfs of % only. in conclusion, mud sct provides a true chance for cure for select patients with dlbc lymphoma who failed conventional therapies, particularity if transplanted with cemosensitive disease. ric, knowm to be associated with a reduced nrm rate, should be especially considered in patients with chemosensitive patients, whereas cc might be the preferred option for patients with more aggressive disease. prospective evaluation of f-fluorodeoxyglucose (fdg) positron emission tomography as a predictor of residual disease and subsequent relapse in patients with diffuse large cell ymphoma and hodgkin's lymphoma undergoing hdc and asct s. akhtar*, a. al-sugair, y. al kadhi, a. al-zahrani, m. abdelsalam, s. bazarbashi, d. ajarim, i. maghfoor king faisal specialist hosp. & res. centre (riyadh, sa) background: there is emerging data indicating poor outcome in diffuse large cell ymphoma (dlcl) and hodgkins lymphoma (hl) patients with positive f-fluorodeoxyglucose (fdg) positron emission tomography (pet) before high dose chemotherapy (hdc) and autologous stem cell transplant (asct). we initiated this prospective trial to evaluate the impact of pet as a predictor of post hdc residual disease and relapse in patients with dlcl and hl undergoing hdc asct. patients and methods: from july to june , patients with relapsed or refractory dlcl and hl were enrolled as a potential candidate for hdc asct. patients did not have hdc asct due to progression ( ), refusal ( ), noncompliance ( ) or other reasons ( ). all eligible patients received eshap as salvage chemotherapy, responding dlcl or responding / stable hl patients received same chemotherapy as mobilization regimen for stem cell collection followed by beam as hdc. prior to the initiation of eshap, each patient had ct scan + other radiological studies if needed and pet ( ct- and pet- ), after - cycles of salvage chemotherapy / prior to hdc asct (ct- and pet- ) and approximately days post hdc asct or earlier if clinically indicated (ct- and pet- ). patients had hdc asct and of them had both pet- and pet- available at the time of this analysis. results: patients characteristics are male: , female: , dlcl: and hl: . relapsed and refractory . median age at asct is years ( to ). median follow-up from asct is months. patients were pet- negative prior to hdc asct. at the time of this evaluation, of these patients, ( %) had an event, event free survival (efs) %. patients were pet- positive prior to hdc asct, ( %) had an event, efs %. efs for pet- negative vs pet- positive has p value of . . using kaplan-meier method, positive pet- has % probability of an event vs % for a negative pet scan (p= . ). efs for ct- negative vs ct- positive patients is % vs %, p = . . kaplan-meier for ct- positive showed % probability of event vs % for ct- negative, p= . . conclusions: prior to hdc asct, positive pet scan indicates high risk of residual disease or progression. many of these patients are likely to suffer from treatment failure. these patients are potential candidate for more aggressive and experimental therapies. for many patient with pre-transplant negative pet, post transplant pet scan can be omitted. radioimmunotherapy with haemopoietic stem cell transplantation for treatment of malignant non-hodgkin lymphoma: multicentre study on thirty patients a. mele ( ) ( ), g. console ( ), n. cascavilla ( ) , p. scalzulli ( ) /kilograms (range . - . ). all patients engrafted. the median number of red blood cell and platelet transfusion were ( - ) and ( - ), respectively. the median time to platelet and neutrophil counts higher than x /l and . x /l were (range, - days) and days (range, - ) , respectively. mucosites occurred in all patients (grade iii and iv in and cases). febrile neutropenia occurred in % of cases. six pneumonitis and blood stream infections were documented. one patient developed an atrial fibrillation. twenty-one of patients were valuable for -day response. the -day overall response was % with % of cr. four early deaths before day- occurred: case for septic shock (day + ), for viral encephalites (day + ) and for progression disease (day + , ). the kaplan-meyer estimated treatment related mortality (trm) is %. seven cases ( cr, pr and not response) progressed at a median follow-up of days post hst (range, - ). twenty-four of patients are alive at a median follow-up of days post hst (range, - ). six patients died ( %): for progression, in cr for ards (day + ) and for trm. sixteen ( %), ( %) and of patients are alive in cr, pr and progression, respectively. one case is not valuable for response (day+ ). we analyzed the characteristics of alive patients in cr: had aggressive lymphoma and were at least in pr before hst (p=ns). the kaplan-meyer estimated y-efs is %. conclusion. the use of rit plus transplant induces % of or ( % cr) with sustained engraftment, an acceptable extra-haematological toxicity and a rapid immunological recovery in patients who failed to achieve cr after first line chemotherapy. the power of this program needs to be assessed in a larger series of patients. hdt/asct has been planned either in front-line pts with high prognostic score/bulky mass/stage iii-iv or at relapse. first line therapy were mainly abvd/mine for hd and acvbp/chop ± rituximab for nhl. fdg-pet was performed after - chemotherapy cure. mine and dhap were the most frequent salvage chemotherapy used for refractory front-line therapy pts. the conditioning regimen was mainly bicnu-etoposide-aracytine-melphalan. pts ( %) were pre-hdt/asct pet negative and positive ( %). / pre-hdt/asct positive pts ( . %) converted to negative by additional cross chemotherapy. after hdt/asct, / others pre-hdt/asct positive pts ( %) converted to post-hdt/asct pet negative. one negative pre-hdt/asct pet converted to positive. residual disease of positive pts was mainly treated by local radiation. after a median follow-up of . years (range . - . ) after pre-hdt/asct pet, pts relapsed, dead (with of relapses), remained resistant disease. survival was measured from pre-hdt/asct pet to death (overall survival os) or relapse/death (event-free survival efs) with censoring time at the time of last follow-up. median os and efs for the two groups were not reached. estimated years os was % and %, respectively for pre-hdt/asct pet negative and positive pts. estimated years efs was % and % respectively for pre-hdt/asct pet negative and positive groups. a positive fdg-pet after induction chemotherapy is highly predictive of poor survival in hd and nhl pts but an additional risk-adapted treatment strategies before hdt/asct by salvage cross chemotherapy and after hdt/asct by targeted radiation may improve pre-hdt/asct pet positive pts outcome. donor lymphocyte transfusions (dlt) after allogeneic stem cell transplantation have been shown to be very effective in treatment of recurrent mylogenous leukemia but displayed limited use in chronic lymphocytic leukemia (cll) and highly malignant non-hodgkin lymphoma (nhl). here we studied whether bi (fbta ), a novel trifunctional bispecific antibody targeting cd on lymphoma cells and cd on t cells could induce graft-versus-leukemia / lymphoma responses in combination with dlt or mobilized peripheral blood stem cells (pbsct) after allogeneic transplantation in these diseases. six patients ( cases each with p mutated cll, and highgrade nhl) refractory to standard therapy were treated with escalating doses of bi (range - µg) followed by dlt or sct. in out of patients, a prompt, but transient clinical and hematological response was observed. side effects (fever, chills and bone pain) were tolerable and appeared at lower dose levels in cll (> µg) than in high grade nhl (> µg). the cytokine profile was characterized by transient increases of il- , il- and il- . neither human anti-mouse antibodies (hamas) nor graft-versus-host disease (gvhd) developed allowing repeated treatment courses. in summary, the trifunctional antibody bi induced prompt antitumor responses in extensively pretreated, p mutated, alemtuzumab and rituximab refractory patients indicating its therapeutic potential. d. stachel*, k. kirby, l. corey, m. boeckh fred hutchinson cancer research center (seattle, us) background: although cmv viral load is a predictor of cmv disease in hct recipients, regulatory agencies presently do not accept it as primary endpoint for studies that evaluate new therapeutics. the aim of this study was to examine whether cmv viral load predicts transplant-related mortality (trm) or overall survival (os) in the era of preemptive therapy. methods: consecutive patients following a first hct at fhcrc between and were analyzed; of them were cmv seropositive. patients underwent weekly testing for cmv viremia by culture and quantitative pp antigenemia (ag) during the first days after hct. preemptive antiviral therapy was given for any pp ag. using univariate and multivariable cox models, we analyzed the association of initial, mean, peak cmv load, and cmv area under the curve (auc) for their association with os and trm at year after hct. viral load parameters were analyzed in quartiles. pp ag results are expressed as cells/ , cells. results: trm occurred in . % of all patients and . % of seropositive recipients. in a model that included cmv seropositive recipients, the adjusted hazard ratios (hr) for the upper quartile of initial viral load (ag > ), peak viral load (ag > ) and auc (ag> ) were . ( % ci . - . ), . ( % ci . - . ), and . ( % ci . - . ), respectively. the statistical models were adjusted for recipient and donor age, hla match, donor relationship, year of hct, acute/chronic gvhd and postengraftment neutropenia (to account for the toxicity associated with preemptive therapy); additional factors evaluated (but not significant in univariate analysis) were race, donor cmv serostatus, cell source, type and intensity of conditioning, t cell depletion, risk of the underlying disease, and gvhd prophylaxis. similar models that included all patients and those that used os as endpoint also showed significant associations for first and peak viral load and the auc. results are presently being validated in a separate cohort that underwent pcr surveillance. conclusion: initial and peak viral load as well as the viral auc are independently associated with trm and os in allogeneic hct recipients receiving preemptive antiviral therapy. the peak viral load showed the strongest association with trm and os. these data further support the use of parameters of viral dynamics as primary endpoints for studies that evaluate immune augmentation or drug prevention strategies. objectives: to evaluate the effect of immunoglobulin (ivig) and cytomegalovirus-hyperimmune immunoglobulin (cmv-ivig) prophylaxis in patients undergoing hematopoietic stem cell transplantation (hsct). methods: systematic review and meta-analysis of randomized controlled trials comparing systemic ivig or cmv-ivig with placebo or no intervention for prophylaxis in patients undergoing hsct. the cochrane library, medline, conference proceedings and references were searched until . the primary outcome was all-cause-mortality at end of follow-up. two reviewers independently appraised the quality of the trials and extracted data. relative risks (rr) with % confidence intervals were estimated and pooled. results: twenty trials fulfilled inclusion criteria, assessed ivig and assessed cmv-ivig. for ivig vs. control there was no difference in all-cause mortality, rr . ; . - . , trials, fig . results were similar when only patients following allogeneic hsct were assessed ( trials), when mortality was assessed at day - ( trials) or after years ( trials), and when high-dose ivig was used ( trials). trials' methodological quality did not impact results. overall, there was a reduction in the number episodes of interstitial pneumonitis (ip), rr . ; . - . , trials, although the reduction in cmv infections was not statistically significant, rr . ; . - . , trials. there was no difference in clinically or microbiologically documented infections, rr . ; . - . and rr . ; . - . , respectively, trials both). there was no difference with regard to acute graft vs. host disease (gvhd), rr . ; . - . , trials. veno-occlusive disease (vod) was significantly more frequent with ivig, rr . ; . - . , trials, fig , an open-label randomised study of oral valganciclovir versus intravenous ganciclovir for pre-emptive therapy of cytomegalovirus infection after allogeneic stem cell transplantation l. volin* ( ), l. barkholt ( ) cytomegalovirus (cmv) remains a leading cause of infectious complications after allogeneic stem cell transplantation (sct) and without preventive measures patients with cmv infection have a significant risk to develope cmv disease. a preemptive treatment approach with iv-ganciclovir or ivfoscarnet is used to restrict the use of potentially toxic drugs for patients at risk for cmv disease. recently, valgan-ciclovir, an oral prodrug of ganciclovir, has become available. the nordic bmt group started a study with the aim of demonstrating that treatment outcome of the first cmv dnaemia after sct with oral valganciclovir is not inferior to that obtained with iv-ganciclovir. the primary study endpoint was achievement of quantitative cmv pcr (qpcr) negativity on day of treatment or earlier. qpcr was carried out weekly for the first three months after sct, and if positive, patients were randomized to receive either oral valganciclovir mg twice a day or ganciclovir iv mg/kg every h for days (adjusted according to renal function). qpcr was studied twice a week. if qpcr was negative on day - of treatment, the treatment was discontinued, and if negativity was not achieved but the copy number was decreasing, maintenance treatment once daily was administered for another days. thereafter the treatment was carried out according to local policy. of the patients receiving an allogeneic sct, patients were treated for a hematological malignancy and for a solid tumor. the patients in the poand iv-groups did not differ by recipient/donor cmv serostatus, age (median vs years), diagnosis, donor (sibling/ unrelated), or the time of pcr positivity (day + and + after sct, respectively). the median cmv dna copy number/ml at diagnosis was ( - ) in the pogroup, and ( - ) in the iv-group. the maximum copy number/ml was ( - ), and ( - ), respectively. the incidence of agvhd ≥ grade was % in the po-group and % in the iv-group. the primary endpoint of the study was achieved in / ( %) of the popatients and / ( %) of the iv-patients. during the study three patients with a solid tumor de-veloped cmv disease, two in the po-group and one in the iv-group. oral valganciclovir therapy seems not to be inferior to iv-ganciclovir therapy in the preemptive treatment of the first cmv dnaemia after allogeneic sct, and the practical advantages of oral treatment are obvious. genetic variations in the nod /card gene are associated with bacteraemia and sepsis after allogeneic stem cell transplantation m. grube, d. veith, g. rogler, j. brenmoehl, h. bremm, j. hahn, r. andreesen, e. holler university hospital regensburg (regensburg, de) purpose: single nucleotide polymorphisms (snps) of the nod/card gene resulting in a diminished nuclear factor-kappab (nf-kappab) response to bacterial cell wall products have been recently associated with an increased incidence of crohn`s disease as well as transplant related mortality and gvhd following allogeneic transplantation. in addition, a recent study from our group revealed nod /card variants as independent predictors of death from septic shock in nonhematological patients. therefore, we now analysed the direct interaction of nod /card variants with febrile bacteremia and sepsis in patients receiving allogeneic sct. experimental design: we retrospectively analyzed donor/recipient pairs for single nucleotide polymorphisms (snps) of the nod/card gene (snp , and ). bacteremia was determined by bacterial growth in the blood culture. septic syndrome was determined by classical criteria. results: / ( %) patients had unmutated snp`s (wild type group), in / ( %) patients either the recipient or the donor had any mutated snp (r or d any mutation). / ( %) patients showed febrile bacteremia, whereas / ( %) developed lethal sepsis syndrome. the cumulative incidence of bacteremia was % in the wilde type group and % in patients with any mutation (r or d any mutation). the cumulative incidence of lethal sepsis syndrome was % in the wilde type group and % in the patients with any mutation (r or d any mutation) (p< , ). in a more detailed analysis of individual donor and recipient snps, presence of snp in the recipient and any snp in the donor (either snp , or ) were signifikant risk factors whereas the highest cumulative incidence of sepsis was found if the donor had a snp mutation (p < , ). since bacteremia/sepsis syndrome even occurred in patients without gvhd or prior to onset of gvhd our data argue against an induction of bacteremia/sepsis syndrome secondary to gvhd. in line with this view, a multivariate analysis including gvhd showed snp`s of the nod/card gene as independent risk factors for lethal sepsis syndrome (p< . ). conclusion: our results suggest that defective signalling of nod /card either in epithelia (recipient) or monocytes/macrophages (donor) may be directly involved in a diminished antibacterial defense. translocation of bacteria may be an important step in subsequent sct related complications. infection control interventions for cancer patients: efficacy evaluation through systematic review and metaanalysis m. paul*, a. schlesinger, a. gafter-gvili, l. leibovici rabin medical center; beilinson campus (petah-tikva, il) background: currently used infection control measures for hematological cancer patients are frequently applied with an unclear evidence basis resulting in a heterogeneous application of these interventions. methods: systematic review and meta-analysis. included were all prospective comparative studies assessing the effects of non-pharmacological interventions applied for infection prevention and control among cancer patients following chemotherapy. interventions were classified according to the transmission modality targeted by the intervention; airborne, contact or endogenous flora. the primary outcome assessed for all interventions was -day all-cause mortality. relative risks (rr) with % confidence intervals are reported. results: we identified studies assessing protective environment (pe) including high particulate air filtration (hepa) ± laminar airflow ( randomized trials, hematological cancer, for hsct), studies comparing outpatient vs. inpatient management for hsct (all nonrandomized) and miscellaneous studies assessing diet, footwear, gowns or specific room design ( randomized trials, for hsct). pe resulted in a remarkable reduction in allcause mortality at day : rr . , . - . overall and rr . , . - . in adequately randomized trials alone. the rr was . , . - . for -day mortality and . , . - . for the longest follow-up, up to years. similar survival benefit was observed for allogeneic hsct patients and other patients. significant reductions were observed for all infection-related outcomes, except mold infections (table) . when isolating the effects of the different transmission modalities it became apparent that endogenous suppression using antibacterial (usually combined with antifungal) prophylaxis was the major contributor to the beneficial effect of pe ( figure) . outpatient hsct ( allogeneic and autologous) was non-inferior to inpatient treatment, rr . , . - . for mortality; rr . , . - . for infections; and rr . , . - . for bacteremia. conclusions: protective isolation offers an overall significant benefit to the patient, since the assessment of all-cause mortality encompasses both infection, cancer and treatmentrelated outcomes. the additional value of hepa and strict contact isolation over antibiotic prophylaxis is unclear and probably depends on the local prevalence of mold infections. outpatient hsct appears safe and should be explored in randomized trials. m. dettenkofer*, r. babikir, h. bertz, a.f. widmer, w.v. kern, e. meyer, p for surveillance of nosocomial bloodstream infections (bsi) and pneumonia during neutropenia in adult patients undergoing bone marrow transplantation (bmt) or peripheral blood-stem cell transplantation (pbsct), an ongoing multicenter surveillance project was initiated by the german national reference centre for surveillance of nosocomial infections in (onko-kiss). methods: nosocomial infections are identified using cdc definitions for laboratory-confirmed bsi and modified criteria for pneumonia in neutropenic patients [for detailed information see : cid ; : - , or in german language: http://www.nrz-hygiene.de/surveillance/onko.htm]. results: over the -month period from july up to june centres participated. altogether , patients with , neutropenic days were investigated. of these, , ( %) had undergone allogeneic and , ( %) autologous bmt or pbsct. the mean length of neutropenia was . days ( . d after autologous and . d after allogeneic transplantation). in total, bloodstream infections and cases of pneumonia were identified. site-specific incidence densities are shown in the table. there was a trend to lower incidence densities over the five years reported. following allogeneic transplantation, . bsi/ patients and . cases of pneumonia/ patients occurred whereas following autologous transplantation . cases of bsi/ patients and . cases of pneumonia/ patients were observed. the main pathogens associated with bsi were coagulase-negative staphylococci ( %). conclusions: the ongoing onko-kiss project adds to the improvement of quality of care in hct-patients by providing sound reference data on the occurrence of bsi and pneumonia during neutropenia. since , surveillance is extended to neutropenic patients with acute leukemia to allow participation for centres not performing hct. t-cell-mediated immunity is an essential host factor in the control of hcmv latency in patients undergoing an allohematopoietic-stem-cell transplantation. our aims were to identify patterns of hcmv-specific immune responses associated with multiple or prolonged reactivations. we analyzed recipients during the course of infection/reactivation and latency. the cd +t-cell responses were weekly determined using hla-class i tetramers together with extended phenotypic analyses. our results showed that recipients from unrelated donors were more susceptible to multiple reactivations and that the donor hcmv serologic status influenced the occurrence of prolonged reactivations. we found that the lack of hcmv-specific t-cells during the first episode of reactivation was associated with multiple further reactivations. the sequential phenotypic follow up showed that patients with uncontrolled reactivations were unable to develop hcmv-specific t-cells of the late differentiation phenotype cd ra+cd -cd -. our data indicate that the longitudinal evaluation of cd and cd ra expression within the tetramer positive subset could help to identify patients developing a protective immune response. the evaluation of hcmv-specific immune responses during the first episode of reactivation, together with extended phenotypes could improve immune monitoring, especially in recipients from unrelated donors and other situations at risk of uncontrolled viral reactivation. invasive aspergillosis in allogeneic stem cell transplant recipients from alternative donor: incidence, risk factors and outcome a.m. raiola*, m. mikulska , b. bruno, m.t. van lint, f. gualandi, d. occhini, a. dominietto, c. di grazia, s. bregante, a. ibatici, t. lamparelli, e. furfaro, f. frassoni, c. viscoli, a. bacigalupo s. martino's hospital (genoa, it) introduction: invasive aspergillosis (ia) remains an important complication with high morbidity and mortality in patients undergoing haematopoietic stem cells transplant (hsct) according the donor type. materials and methods: we determined, with retrospective analysis, the incidence, risk factors for ia and outcome in patients who received hsct from unrelated donor (mud), family mismatched donor, or cord blood between january and december in our unit. the diagnosis of ia was documented as proven or probable according to the european organisation for research and treatment of cancer eortc/niaid international consensus. we have also considered proven ia the presence of fungal invasion on autopsy. results: a total of patients were included in the study, with a median follow-up of days after hsct, (range - days) (types of hsct: matched unrelated %, mismatched related %, mismatched unrelated %, cord blood %). there were cases of proven and probable ia, with prevalence of , %. the median time to onset of was day + (range: - ), with ( %) cases diagnosed in neutopenia (before take o secondary neutropenia). there were cases diagnosed on autopsy as a proven disease while antemortem they were classified as probable ( ) or possible ( ); no clinical suspicion was present in patient. the diagnosis of probable ia was made by galactomannan platelia test in patients and by sputum culture in . in patients both criteria were present. in cases ia was present within lungs only, whereas patients developed disseminated ia. multivariate analysis identified the following risk factors for ia: late take of neutrophil cells (p= . ) and steroid therapy (p= . ). among patients with ia, died and deaths were related to the mould infection. mortality was % (p< . ). multivariate analysis, among patients with ia, for overall survival identified the following risk factors: atg use in conditioning regimen (p= . ) steroid therapy, relapse, iga and cholinesterase at diagnosis of ia (p= . , . , . and < . , respectively). conclusions: the prevalence if ia remains high among alternative hsct recipients, with few risk factors for ia. clinical and radiological presentation is highly aspecific and invasive procedures are rarely feasible. about concern survival of ia immuno deficiency at diagnosis seems to be important. henze ( ) minimal residual disease (mrd) quantified prior to allogeneic sct has been shown to predict outcome in children with relapsed all in retrospective meta analysis. based on these results intense discussions were started as to whether transplant procedures should be adapted according to the mrd levels pre transplant. within the all-rez bfm group we have started a prospective trial evaluating the impact of pre-transplant mrd load in a well defined group of children who received their transplant in second or subsequent remission. between march and july , children with relapsed all treated according to the protocols all-rez bfm or and receiving allogeneic sct in nd (n= ) or rd cr (n= ) have been enrolled. mrd quantification was performed within days prior to sct by real time pcr using t-cell receptor and immunoglobulin gene rearrangements as clone-specific targets with at least marker with a sensitivity of - . probability of event free survival (pefs) and cumulative incidence of relapse (cir) in patients with mrd ≥ - was . (± . ) and . (± . ) compared with . (± . ) and . (± . ) in patients with mrd < - (p [efs, log-rank]= . ; p [cir, gray] < . ). clinical and therapeutical parameters were equally distributed between both subgroups. the difference in pefs and cir was more prominent in intermediate risk patients s (n= ) compared to high risk patients s /s /cr (n= ). thus, s patients with mrd ≥ - (n= ) showed pefs of . (± . ) and cir of . (± . ), where as patients who entered transplantation with mrd < - (n = ) had a pefs of . (± . ) and a cir of . (± . ); (p[efs] = . ; p[cir] < . ). high risk patients s / /cr who entered transplantation with a mrd load of < - (n= ) showed a pefs and cri of . (± . ) and . (± . ), respectively. in contrast, pefs and cri were . (± . ) and . se (± . ) in patients who entered transplant with higher mrd load of > mrd- . multivariate cox regression analysis revealed mrd as the only independent parameter predictive for efs (p= . ). mrd prior to allogeneic sct proves to be the most important risk factor for outcome post transplantation. new strategies with modified sct procedures including conditioning regimen, graft manipulation, and gvhd prophylaxis and or post transplant intervention strategies are warranted and will be performed to improve prognosis in patients with a high risk of relapse. acute lymphoblastic leukemia (all) remains the most common indication for unrelated donor bone marrow transplantation in children. there have been major advances in conditioning regimes; supportive care and availability of donors, all of which have made transplant a viable treatment option for patients with relapsed or high-risk disease. we retrospectively analyzed data from consecutive allogeneic transplants for all performed at the royal bristol hospital for children from october until september . transplants were performed in patients with all. disease status at the time of transplant was complete remission(cr)- , n= ; cr- , n= , cr- , n= , children not in complete remission n= .(see table ) all patients were pre-treated on the current mrc-ukall protocols for the time period. eighty-six patients received stem cells from identical siblings or relatives. matched unrelated donors were used in children, mismatched unrelated donors in , haplo-identical family donors were used in children. included in these totals are umbilical cord blood donations. cyclophosphamide and total body irradiation was used as conditioning therapy with the exception of children under the age of two and children receiving second transplants. ciclosporin a was used for graft verses host disease (gvhd) prophylaxis. short course methotrexate was given to mismatched and sibling donors. in the unrelated group t-cell depletion was effected using campath monoclonal antibodies. the majority of patients died due to disease relapse. the incidence of chronic graft verses host disease was low with correspondingly high karnofsky scores. this study offers a unique opportunity to analyze data from a large number of patients consistently pretreated according to united kingdom national leukaemia protocols and with a uniform approach to transplant conditioning, gvhd prophylaxis and supportive care over a year period. background: intravenous bu has been used in the two dutch pediatric stem cell transplantation centers in various myeloablative regimens with different targets for the area under the curve (auc) and different dosing regimens of bu (once or four times daily). we retrospectively analyzed the association between bu exposure expressed as auc and clinical outcome. methods: all children, transplanted between - , receiving intravenous bu as part of a myeloablative regimen, were included. patients were separated into quintiles based on the total auc of four days of treatment. the association with the primary endpoints death, graft-failure or relapse, efs and the secondary endpoints (acute graft-versus-host disease grade - (agvhd), veno-occlusive disease (vod) and mucositis grade - ), were tested using uni-and multivariate cox regression analysis. the lowest auc group was used as index group. results: patients were included ( malignant indications; non-malignant indications). median age at transplantation was . years (range . to years). the overall efs and survival were % and % respectively. in multivariate analyses a total auc between . - mg*h/l was associated with highest efs ( %, p= . ) and survival ( %, p= . ). a lower auc bu was significantly associated with higher incidence of relapse or graft failure, whereas in the highest auc-percentile a high incidence of treatment related mortality (trm) was seen. covariates were hla-disparity and age. hla-mismatched donors and older children showed a significantly lower efs, the latter mainly due to a higher trm. agvhd occurred in %, vod in % and mucositis in % of patients. these side effects significantly correlated with a high auc bu in combination with the addition of melphalan to the conditioning regimen. the once daily dosing versus fourtimes daily did not influence any of the results. in conclusion a total auc of bu between . - mg*h/l is associated with highest efs in malignant and non-malignant disease. a lower auc correlated with more relapse/graft failure especially in hla-mismatched donors. a higher auc was associated with more trm negatively influencing efs especially in older children. the occurrence of severe agvhd, vod and severe mucositis was mainly related to combination of bu and melphalan. dosing of busulfan in children by tdm to a target of . - mg*h/l might improve efs. mechanical ventilation in a recent cohort of children after allogeneic haematopoietic stem cell transplantation: risk factors and outcome s. van gestel*, c. bollen, m. bierings, j. boelens, n. wulffraat, h. van vught university medical center utrecht (utrecht, nl) introduction: in previous studies, median intensive care unit (icu) mortality in children after hematopoietic stem cell transplantation (hsct) was % (range % - %). it has been suggested that icu mortality decreased over time, but we could not confirm this in a recent meta-regression analysis (van gestel et al, submitted) . conclusions on mortality trends were limited, since recent outcome data from children requiring mechanical ventilation after hsct were scarce. objective: to assess risk factors for and outcome of mechanical ventilation in a recent cohort of children after allogeneic hsct. design: retrospective chart review. setting: a pediatric icu and hsct centre in a university hospital in the netherlands. patients and methods: all children who received an allogeneic hsct between january and april were included. patients who required endotracheal mechanical ventilation for more than hours were identified. potential risk factors for the requirement of mechanical ventilation and for icu mortality were recorded. uni-and multivariable analyses were done to identify risk factors. results: hscts were performed in patients. thirtyfive patients ( % of hsct recipients) received mechanical ventilation on occasions. none of the potential risk factors was significantly associated with icu admission. there was a trend towards an increased probability of icu admission over the years. this suggests that transplantations were carried out in sicker patients over time. sixteen admissions resulted in death on the icu, giving a case fatality rate of %. icu mortality was mainly associated with (persistence of) multiple organ failure on the first days of admission. none of the pre-admission transplantation characteristics significantly influenced icu mortality. based on their pediatric risk of mortality (prism) scores, our patients had a higher acuity of illness on icu admission than patients in previous studies. six-month survival in patients discharged from the icu was %. conclusion: we found a significantly lower icu mortality in a recent cohort of children after allogeneic hsct compared to previous studies, even though our patients were sicker. this can most probably be explained by improvements in transplantation medicine and icu treatment strategies. results from our study are promising, but need to be confirmed in other, preferably multi-centre, studies. early and comprehensive vaccination coverage in paediatric recipients of related and unrelated stem cell transplantation following coadministration of the pneumoccoccal conjugate vaccine prevenar™ and the hexavalent combination vaccine infanrix hexa™ r. meisel ( ) children undergoing allogeneic hematopoietic stem cell transplantion (ahsct) lose protective immunity to vaccinepreventable disease and thus are at significant risk for lifethreatening infections. however, important issues on reimmunization after pediatric ahsct remain controversial due to the lack of prospective clinical trials. in the prospective multicenter vaccination trial ikast (nct ) a total of pediatric allohsct recipients (median age . ( . - . ) years) were therefore immunized with a primary series of three monthly doses of the hexavalent tetanus, diphtheria, pertussis, poliomyelitis, haemophilus influenzae type b and hepatitis b combination vaccine infanrix hexa ( vcv; glaxosmithkline pharma) along with the heptavalent pneumococcal conjugate vaccine prevenar (pcv ; wyeth pharma). vaccination was started at months after transplantation, irrespective of immunosuppressive therapy or gvhd, with a subsequent booster dose at months. immunogenicity was analysed by assessment of antibody concentrations and adverse events were prospectively collected. prior to immunization only . % and . % of patients (pts) exhibited protective antibodies towards pcv and vcv vaccine antigens, respectively. as a result of highly significant increases in mean antibody concentrations (p< . ) protection to all pcv and vcv antigens was achieved in . % and . % of pts after primary immunization within the st year after allohsct, and this was independent of patient age, conditioning regimen, stem cell source, and most importantly, donor type (related vs. unrelated) and in vivo t cell depletion (all p-values> . ). nine months later, prior to booster vaccination, . % and . % of pts retained protective antibody concentrations. however, mean antibody concentrations had dropped by a factor of . - . (p< . ) except for pneumococcal serotype , thus underlining the need for subsequent vaccination. following booster immunization, antibody concentrations increased . - . fold (p< . ) indicating robust memory responses, and . % and % of pts achieved protective antibody levels against all pcv and vcv antigens. vaccination was well tolerated with no vaccine-related saes. our data show that early immunization of pediatric ahsct recipients according to our simple revaccination schedule is safe and provides early and comprehensive vaccination coverage during the first years following ahsct from both related and unrelated donors. treosulfan-containing regimens achieve high rates of engraftment associated with low transplant morbidity and mortality in children with non-malignant disease and significant co-morbidities b.f. greystoke* ( ), s. bonanomi ( ) ( ), p. naik ( ), k. rao ( ) , n. goulden ( ) , p. amrolia ( ) , r.f. wynn ( ) , p.a. veys ( ) ( )pendlebury children's hospital (manchester, uk); ( )great ormond street hospital (london, uk) treosulfan is an alkylating agent with both immunosuppressive and myeloablative properties, which has recently been introduced as a conditioning agent in allogeneic and autologous haemopoietic stem cell transplantation (hsct). early studies have been performed principally in adult patients with malignant disease in whom the drug appears to be well tolerated with a low incidence of regimen-related toxicity. we report the use of treosulfan in consecutive children undergoing sct for non-malignant disease: immunodeficiency (n= ), metabolic disease (n= ), osteopetrosis (n= ) and other (n= ). patients received a total treosulfan dose of or gms/m²/patient given in divided doses on successive days. a range of other conditioning agents and serotherapy was administered to patients who underwent family donor sct (n= ), or unrelated donor sct (n= ). bone marrow (n= ), peripheral blood (n= ) and cord blood (n= ) were used as a stem cell source and in one patient a combination of marrow and cord blood from the same sibling donor was used. one patient ( %) died of early transplant-related complications. transplant morbidity was limited and there was no vod other than in this patient. mucositis was mild and many patients did not require parenteral nutrition. nappy rash with ulceration was noted in many patients. twenty-eight patients ( . %) established donor cell engraftment which was full in ( . %). in patients ( %) there was stable donor engraftment, including sufficient lineage specific chimerism, to correct the underlying condition. four patients required additional transplant procedures to maintain adequate donor-derived haemopoiesis, and two patients are under consideration for further hsct. twentyseven patients survive with a median follow up of days. there were late deaths due to progression of the underlying disease, gvhd or infection. treosulfan based conditioning regimens achieve excellent engraftment with reduced regimen-related toxicity and in children with non-malignant disease at high risk for both regimen-related toxicity and graft failure. preliminary results indicate that in a haploidentical setting cd /cd -depleted grafts may be advantageous regarding engraftment and immunreconstitution. since effector cell with potential antileukemic activity are cotransfused, such grafts may be suited in particular for patients with insufficient remission. nk cells have been shown both in vitro and in vivo to mediate positive effects regarding engraftment, gvhd, immunreconstitution and relapse and are an important part of the cd / depleted grafts. we analyzed nk activity in cd / depleted grafts, which was low compared to fresh isolated pmncs. several cytokines and cytokine combinations for activation of the grafts were tested and the strongest enhancement in nk activity could be obtained with il- . we therefore developed a protocol for the overnight incubation of the grafts according to gmp. stimulated grafts were transplanted seven times either at day or as an immunotherapeutical approach after transplantation and were well tolerated. median cell counts for infused cells/kg bodyweight were x cd +/ -, x cd +, x cd +, x cd + and , x cd + cells. no occurrence of agvhd was seen in these patients after infusion of the cells. enhanced nk activity after stimulation could be demonstrated in vitro against k or primary leukemic blasts. nk cells showed increased proliferation capacity in brdu assay and [ h]-thymidine incorporation assays, especially after additional stimulation with il- while t cells showed only a moderately enhanced proliferation against third party stimulators or with okt . therefore activation of haploidentical t cell depleted grafts with il- is a promising tool to enhance the activity of infused nk cells and should be further investigated in clinical trials. background: development of an effective strategy for patients with epmd. methods: patients (pts) were registered. median age is . years (yrs) ( . - ). primary site was extremity in pts and axial/other in pts ( % in the pelvis). metastatic spread was bone marrow (bm) only in pts, bone only in pts and bone & bm in pts, other (mainly lymph nodes) metastatic sites in pts. tumour volume was above ml in pts. six vide induction cycles were completed by %. local treatment included surgery when possible and/or radiotherapy (rx) as indicated. recommended hdt was busulphan (bu) mg/m² and melphalan (mel) mg/m² with pscr. median follow up is years (range, - . ). results: partial remission or better was achieved after cycle in %. the overall survival at years is %± . in patients with bm/bone metastasis, significantly favourable univariate factors in the unselected cohort at diagnosis (dx) were age < yrs (event free survival at yrs (efs) %, p< . ), metastatic site (efs: bm only %, bone only %, bm+bone %, p= . ), single bone lesions only (efs %, p= . ), primary tumour site (efs: extremities %, chest/spine/hn %, abdomen pelvic %, p= . ) and tumour volume of < ml (efs %, p< . ). ). multivariate analysis identified four major risk factors at dx: primary tumour volume > ml p< . (rr ) and > bone metastases p= . (rr . ), age above p= . (rr . ), bm metastasis p= . (rr= . ). for pts receiving bumel it is noteworthy that pts of < a and epmd achieved an efs of % in comparison to older counterparts > a (efs %, p= . ). conclusion: further strategy refinement and validation of hdt appears necessary within investigational , ideally randomised studies. allogeneic haematopoietic cell transplantation for chronic lymphocytic leukaemia with p deletion: a retrospective ebmt analysis j. schetelig*, a. van biezen, r. brand, d. caballero, r. martino, m. itala, j. garcía-marco, l. volin, n. schmitz, r. schwerdtfeger, a. ganser, f. onida, b. mohr, s. stilgenbauer, m. bornhäuser, t. de witte, p. dreger on behalf of the chronic leukemia working party, ebmt purpose: patients with advanced chronic lymphocytic leukaemia (cll) and p deletion have a very poor prognosis even after intensive chemotherapy. while allogeneic hematopoietic cell transplantation (hct) has the potential to cure patients with advanced cll it is not known whether this holds true for patients with p deletion. patients and methods: patients with p-cll who had received hct were identified by an ebmt-based survey. baseline data were downloaded from the ebmt database. additional information on the course of the disease, the cytogenetic diagnosis and last follow up was collected by a questionnaire. data were analysed as of february . results: patients were identified. twelve patients with autologous hct, haplo-identical donors or the detection of p-after hct were excluded from further analysis. patients had received an allogeneic hct between march and july from a matched sibling donor (n= ) or an alternativ donor (n= ). the median age at hct was years. the diagnosis of deletion p-was made by fish in % and by conventional banding in % of patients. the median interval between first diagnosis and detection of pwas . years and the median interval between detection of p-and hct was . years. patients had received a median of chemotherapy regimens, including fludarabine in % of patients. at hct, % of patients were in remission. reduced intensity conditioning was applied in % of patients. % of the patients received peripheral blood stem cells. gvhd prophylaxis was performed heterogeneously. one patient experienced primary graft failure. acute gvhd grades ii to iv occurred in % of patients and extensive chronic gvhd in % of patients. after a median follow-up of months (range, to months) of patients who are alive, were in complete remission, in partial remission and patients had progressive disease at last follow up. -year overall survival and progression-free survival was % ( % ci, % to %) and % ( % ci, % to %). the cumulative incidence of relapse at years was % ( % ci, % to %). no additional relapse occurred in six patients with a follow-up between and . years. conclusion: allogeneic hct has the potential to induce longterm disease-free survival in selected patients with advanced p-cll. given the otherwise very dismal outcome of this disease, prospective studies on allogeneic hct earlier in the course of p-cll seem warranted. separating patients with myelofibrosis with myeloid metaplasia into risk groups for allogeneic haematopoietic cell transplantation -results of a german multicentre analysis f. collenbusch, r. schwerdtfeger, m. schleuning, c. schmid, j. finke, m. stadler, m. bornhaeuser, d. messerer-schmid, h objective: we aimed to derive risk factors for allogeneic hematopoietic cell transplantation (allohct) in patients with s myelofibrosis with myeloid metaplasia (mmm) from retrospective analysis. patients: between and patients (pts) from german centers, median (md) age ( - ), were grafted from related and unrelated donors. pts had primary, secondary mmm. at allohct pts were dupriez score , score and score , needed red cell transfusions. chromosomal and molecular analysis was available from and pts resp.: pts had favourable, unfavourable cytogenetics, pts were jak mutated. conditioning was of standard, intermediate and reduced intensity in , and cases. pts received bm, pbsc. boosts and donor lymphocyte transfusions were given for relapses, incomplete chimerisms and one graft failure. results: at a md follow up of ( - ) days probability of overall and relapse free survival (os and rfs) was and %. neither time from diagnosis to transplant, age, bsymptoms, marrow fibrosis, splenomegaly, jak , hemoglobin < g/dl, platelets < /nl, ldh, comorbidity (cci or hct-ci), conditioning, type of donor nor graft source were predictive for os. log rank test showed a trend towards lower survival in cases with dupriez score ≥ (p= . ) and transfusion dependence (p= . ). solely circulating blasts > % (p= . ), monocytes > /nl (p= . ) and cytogenetics (p= . ) were predictive for survival after allohct. cox regression analysis within pretransplant variables revealed cytogenetics (p= . ) and monocytes (p= . ) as independent factor for os. for rfs only cytogenetics retained independence (p= . ). combining cytogenetics, monocytes, blasts and transfusion dependence to a risk score from to allowed to discriminate between good ( - ), intermediate ( ) ( ) and high risk ( ) pts for os ( , , %, p= . ) and rfs ( , , %, p= . ). following allohct pts with limited cgvhd had a projected os and rfs of % and % as opposed to % rfs for no and % rfs for extensive cgvhd, resp. (p= . ). pts treated with cellular immunotherapy achieved a partial and a complete response. conclusions: pts with mmm can be scored as low, intermediate and high risk for allohct according to pretransplant disease characteristics. evidence for a potent graft-versus-mmm-effect suggests exploitation of adjuvant cellular immunotherapy to improve current results. syngeneic bmt for cml in chronic phase: update of the seattle results a. fefer*, j. radich, t. gooley, l. holmberg, m.e. flowers, s. pavletic, r. storb, f. appelbaum fred hutchinson cancer research center (seattle, us) to determine the efficacy of syngeneic bmt as treatment for chronic phase cml (cml-cp), we reviewed the results of all patients (pts) we transplanted between and , before the gleevec era. of pts, died within months (mos)due largely to pulmonary problems. of the pts who went into a complete cytogenetic remission (cr), relapsed at a median time of years after bmt, but remained in cr for up to (median, ) years. all pts tested were negative by pcr when last tested at - (median, ) years. thus, enduring cytogenetic and molecular cr's were achieved with syngeneic bmt, in the absence of an allogeneic graft vs leukemia (gvl) effect. at years, the estimated probability of relapse, overall survival and relapse-free survival was %, % and %,respectively. to assess the influence, if any, of the conditioning regimen on the relapse rate, we subdivided the pts into groups transplanted in time periods with different conditioning regimens, as follows: group i (n= ), - , dimethylbusulfan (dmb) mg/kg iv, cyclophosphamide (cy) mg/kg/day x and total body irradiation (tbi) gy at a single exposure; group ii (n= ), - , cy mg/kg/d x & tbi gy/day x ; group iii (n= ), - , busulfan (bu) mg/kg/day x po, cy mg/kg/day x and tbi gy/day x . the median time from diagnosis to bmt for the groups was , & months, respectively. relapses occurred in pts in group i at a median of years, pts in group ii at a median of eayr, and pts in group iii at and years. thus, the relapse rate with the cy/tbi regimen was significantly higher than that for the regimens containing dmb or bu (hazard rate (hr) of relapse for group ii vs i was . , p= . , and for group ii vs iii, hr . , p= . ). however, cy/tbi was less toxic, with no early bmt-related deaths in contrast to / and / deaths in groups i and iii. finally, we compared the risk of relapse in groups i and iii combined i.e.twins conditioned with dmb/bu-containing regimens, with that of cml-cp pts who underwent bmt from allogeneic matched siblings after cy/tbi at our center. the risk of relapse is quite similar (hr= . , p= . ). these results suggest that either the dmb or bu can eradicate a number of cml cells similar to that destroyed by an allogeneic gvl effect and/or that the dmb or bu can somehow induce a gvl effect exerted by syngeneic cells. we have previously shown that levels of the polycomb group (pcg) gene bmi- rna were significantly higher in advanced phase than in chronic phase (cp) chronic myeloid leukaemia (cml). in addition, in patients treated with hu and ifn-a, low bmi- expression was associated with an improved overall survival (os) (blood ). here, we investigated whether bmi- and other previously established prognostic genes (cd , pr- and ela- ) are implicated in the prognosis of cml in the context of allogeneic stem cell transplantation (allo-sct). we studied cp-cml patients who received allo-sct from hla-identical related donors. cd , pr- , ela- and bmi- expression was assessed by q-rt/pcr in the recipients pbmcs collected before allo-sct. the median expression level for each gene was used to segregate the patients into groups (low: gene expression <median, and high: >median). the median fu post-allo-sct was . (range, . - . ) years. the median ebmt-gratwohl score was . none of the tested genes showed any significant association with engraftment or with graft rejection. cd , pr- and ela- expression was not associated with os. however, in contrast to our previous findings in the non-allo-sct setting, patients displaying a high bmi- expression level prior to allo-sct had significantly better os than those with low expression (p= . ). when bmi- was included in a multivariate survival model and adjusted for the other prognostic variables, a high expression was found to be an independent marker associated with better survival (rr= . , %ci; . - . ;p= . ). given the impact of bmi- expression level on os, without a significant association with relapse, and since neither bmi- , nor the other genes showed any significant association with leukemia-free survival, we assessed their impact on trm. there was a striking and significant association between acute gvhd and bmi- expression, not only in overall incidence (low bmi- : grade - (n= ), grade (n= ), grade - (n= ); high bmi- : grade - (n= ), grade (n= ), grade - (n= ); p= . ), but also in cumulative incidence at day ( % vs. %, p= . ). in multivariate analysis, a low bmi- expression level was associated with an increased risk of grade - acute gvhd (rr= . , %ci; . - . ; p= . ). these results suggest that bmi- can serve as a biomarker for predicting outcome in cp-cml patients receiving allo-sct. such measurement allows for tailored therapeutic intervention, including informed recommendation for allo-sct in patients failing tyrosinekinase inhibitors. haematopoietic stem cell transplantation in tprolymphocytic leukaemia: a retrospective ebmt analysis w. wiktor-jedrzejczak*, r. brand, a. van biesen, e. carreras, v. leblond, g. cook, m. ethell, a. nagler, t. ruutu, m.l. brune, j. schetelig, t.m. de witte, p. dreger on behalf of the clwp ebmt t prolymphocytic leukemia (t-pll) is a rare, aggressive neoplasia of t lymphoid lineage which is characterized by poor survival of less than one year. incidental reports suggest that both autologous and allogeneic hematopoietic stem cell transplantation (hsct) might be effective in this disease. however, no larger series on the efficacy of hsct in t-pll has been reported to date. therefore the purpose of the present study was to analyze the outcome of transplants for t-pll registered at the ebmt database. results: eleven patients who had undergone autologous transplantation (auto-sct) and patients who had undergone allogeneic transplantation (allo-sct) were identified from the database and verified as t-pll. in the auto-sct group, there were males and females, with a median age of ( - ) years. patients were transplanted within the st year of diagnosis, and the remainder in later phases. patients received tbi in their conditioning, while received chemotherapy only. engraftment and recovery was prompt. there was one case of non-relapse death (nrm). patients relapsed within the first post-transplant years, translating into a median event-free survival (efs) of months and a median overall survival (os) of months. a single patient had prolonged disease control until relapse after months.in the allo-sct group ( males/ females), median age was ( - ) years; patients ( %) had been transplanted within the st year of diagnosis. status at allo-sct was complete or partial remission in patients ( %), and more advanced disease or unknown in the remainder. patients ( %) received reduced intensity conditioning. donors were hla-identical siblings in patients ( %), matched unrelated donors in patients, and a mismatched unrelated donor in one patient. engraftment was documented in of patients ( %) with information available. -year nrm was %. of relapses observed occurred during the first post-transplant year, mainly in patients transplanted in advanced disease, translating into biphasic survival curves with initial steep decline (median efs months, median os months) but still % efs at months suggesting sustained disease control in a subset of patients. conclusions: these data indicate that allo-sct may provide effective disease control in selected patients with t-pll. prospective investigation of allo-st in eligible patients with this otherwise fatal disorder seems to be warranted. high-dose chemotherapy with autologous stem-cell support versus standard-dose chemotherapy: metaanalysis of individual patient data from randomised adjuvant breast cancer trials t. demirer*, n.n. ueno, m. bregni, d with autologous haematopoietic stem cell transplantation (hsct) for the treatment of primary poor risk breast cancer (bc) patients (pts) has lost favour over recent years in the oncology community. this is mainly due to the worrisome high mortality and morbidity of the procedure, and the lack of a clear survival benefit in early randomized studies. recently reported trials have demonstrated that hdc with hsct could still have a role in selected subgroup of pts, namely pts with ≥ positive axillary lymph nodes (ln) and in pts with her- negative tumours. aim of this study is to re-evaluate toxicity and efficacy of hdc with hsct in a large cohort of pts receiving hdc in italy between january , and december , . bc pts receiving hdc for poor risk bc were identified in the gitmo registry. in patients with > ln, a thorough data set including biological characteristics, toxicity and follow up was available. median age was years ( - ), % of pts were pre menopausal at treatment, % had an endocrine responsive tumours and % had a her- + tumour. median number of positive axillary ln was ( - ), with % of pts having ≥ ln+. % of pts received alkylating agents-based hdc as a single procedure while % received epirubicin or mitoxantrone-containing hdc, usually within a multi-transplant program. transplant related mortality (trm) at days was . %, while late cardiac and secondary tumour related mortality were around % overall. with a median follow up of months, median disease free survival (dfs) and overall survival (os) in the entire population were . and . year, respectively. exploratory subgroup analysis demonstrated that os was significantly better in endocrine responsive tumours (p= . ), while menopausal or her- status did not affect survival. in poor prognosis pts with er, pgr and her- negative tumours (median ln+ = ), median os was months. median os was significantly better (p= . ) in patients receiving multiple transplant procedures, this effect being particularly evident in the ≥ ln+ population. in conclusion, our retrospective analysis suggests that hdc with hsct has lower trm than expected and high efficacy in well defined subgroups of patients. multiple transplants seem more active than single hdc procedures. this analysis could be useful in selecting well defined patient populations in which to re-address the role of hdc as adjuvant treatment. long-term follow-up of metastatic renal cancer patients undergoing reduced-intensity allografting m. bregni*, m. bernardi, p. servida, a. pescarollo, r. crocchiolo, e. treppiedi, f. ciceri, j. peccatori scientific institute san raffaele (milan, it) objective: stem cell transplantation from a hla-compatible sibling donor has been advocated as adoptive immunotherapy for cytokine-resistant, metastatic renal cancer (rcc). however, the recent introduction of several targeted therapy compounds has reduced the interest in this therapeutic strategy. we have reanalyzed our transplant series with the aim to detect long-term benefit form allografting. methods: from february to may , patients with cytokine-refractory rcc received a reduced-intensity allograft from an hla-id sibling donor. median age was years; most ( ) had clear-cell histology. median number of previous treatments was ( - ). median days from diagnosis to allograft were . all patients received a thiotepa, fludarabine, and cyclophosphamide conditioning regimen, and a cyclosporine-based gvhd prophylaxis. six patients received dli at escalating doses for progressing or nonresponding disease. results: one-year-os was % ( % ci: - ), and y-os was % ( % ci: - ). at a median observation time of months, patients are alive, one in cr, one in vgpr, and three with disease. we have analyzed the correlation of the following variables with survival: pre-transplant disease status, age at transplant, time from diagnosis to transplant, total infused cells, infused cd + cells, infused cd + cells, bm chimerism at + , post-transplant disease status at + and + , best response, day of best response, csa withdrawal day, infusion of dli, occurrence of gvhd. at univariate analysis, numbers of cd +/kg infused cells, best response, and disease status at + significantly correlated with survival. at multivariate analysis, only disease status at + retained statistical significance (p= . ), while cd + cells/kg retained marginal significance (p= . ). conclusion: twenty percent of cytokine-refractory rcc patients are alive at a median ( - ) months after allografting. all these patients have received > x cd + cells/kg, and had stable or responding disease at + after transplant. three patients are receiving sorafenib, and two are in cr/pr without further therapies. reduced-intensity allografting is able to induce long-term disease remission in a fraction of relapsed rcc patients. it is unknown if relapse or pd after targeted therapy will be susceptible to allograft-mediated gvt effect. the place of allografting in the treatment of metastatic rcc, alone or in combination with targeted therapies, needs reappraisal. haploidentical family donors represent the ideal solution to offer to every patient with high risk leukemia the potential cure of marrow stem cell transplantation. extensive application of haploidentical transplantation (haplo-sct) is limited by high rate of late transplant related mortality (trm) and relapse associated with the delayed immune reconstitution secondary to the procedures for severe graft-vs-host-disease (gvhd) prevention. in a haplo-sct phase i-ii multicenter, open, no-randomized, trial sponsored by molmed spa, we infused donor lymphocytes genetically engineered to express the suicide gene herpes simplex thymidine kinase (tk-dli) to induce early immune reconstitution, while selectively controlling gvhd. between september and september , patients (pts) -median age -with high-risk hematologic malignancies were enrolled, out of pts were in complete remission (cr). after myeloablative conditioning regimen, pts received a median x /kg cd + and . x /kg cd + (median time to engraftment: weeks). no immune reconstitution were observed in absence of tk-dli. twenty-seven pts received tk-dli: pts obtained prompt immune reconstitution with cd +> /mcl at day+ (median) from haplo-sct and day+ from tk-dli. eleven pts developed gvhd ( acute gvhd grade i-iv and chronic gvhd) that was always abrogated by the suicide gene induction. the -year trm was %, with last infectious event at day post transplant, for pts treated with tk-cells, and % for pts who didn't receive tk-cells (p< . ). immune reconstitution obtained with tk-cells infusion correlated with: . rapid development of a wide t-cell repertoire, . detection of high frequencies of t-cells specific for opportunistic pathogens, . abatement of the incidence of infectious adverse events (ae) and serious ae. the years lfs was % for pts who achieved immune reconstitution and % for pts who failed immune reconstitution (p:< . ). this strategy is feasible and effective in providing immune reconstitution in haplo t-cell-depleted setting. in uni-and multi-variate analysis both status at transplant and immune reconstitution are significant risk factor. infusion of tk-cells could significantly extend the application of haplo-sct. a randomized phase iii study comparing tk-dli versus any t cell repletion strategy after haplo-hsct in high risk acute leukemia is now starting. ie- and pp specific peptide pools for the generation and expansion of cmv-specific donor t-cells after haploidentical stem cell transplantation: feasibility and first clinical experiences s. ganepola* ( ), p. reinke ( ) , c. gentilini ( ) , m. hammer ( ) , m. schmidt-hieber ( ), c. tietze-bürger ( ), k. freyberg ( ), d. volk ( ) , e. thiel ( ) , l. uharek ( ) ( )hematology/oncology/transfusionmedicine (berlin, de); ( )charite berlin campus mitte (berlin, de) for immunocompromised patients, rapid reconstitution of cytotoxic t cell function is mandatory for the control of human cytomegalovirus (cmv) infection and disease. the cmvassociated proteins pp and ie- are important components of a relevant cmv-directed immune response. here we report the first clinical experiences concerning feasibility and safety with a novel method for the generation of cmv-specific t-cells by stimulation with a peptide-pool containing pp and ie- . material and methods: after apherisis of - . x pbmc from healthy donors ( igg positive, igg negative) and patients with proven cmv disease, cells were further processed under gmp-conditions. cells were stimulated with peptide pools representing the pp (ul ) and ie- (ul ) proteins and ifng producing cells were selected with the cytokine-capture-assay (miltenyibiotec) and further expanded in cell-culture conditions on a -well-plate. after performance of intern and extern quality controls, cells were freshly retransfused and/or kryoconserved in defined portions for further redonations. results: in / rounds we could expand cmv-specific t-cells. expansion-rates ranged from . fold to . fold (median . ) of the initial cell count. in a median time of days (range - days), cell counts expanded from . fold up to . fold (median . ). facs-analysis at the end of the culture revealed that in median % of the cells were cd + (range . - . %) and, respectively, % were cd + (range . - . %). in cytotoxicity assays, a lysis of - % (median %) of lcl-line cells could be achieved. so far, two patients with therapy refractory cmv disease received in vivo expanded polyvalent t-cells against multiple (ie- /pp ) cmv epitopes. no serious adverse events occurred during the first days after administration and cmv antigenemia was decreasing in one patient. however, since both patients finally died from a septical multi-organ failure, we were not able to assess the long lasting impact of the manoeuvre on the control of cmv-disease. conclusion: selection and subsequent expansion of cmvspecific t-cells is possible with this method which allows an up to -fold expansion of cd + cells. the use of pp and ie- specific peptide pools for the prevention or treatment of cmv infection is feasible and could be of superior effectiveness as compared to approaches directed against a single cmv epitope. lmp -specific tcr gene therapy for hodgkin's lymphoma d.p. hart, s. thomas, s. xue, h.j. stauss, e.c. morris* university college london (london, uk) ebv-positive hodgkin lymphoma typically demonstrates latency ii antigen expression, characterised by loss of most ebv antigens except for the latent membrane protein (lmp) and and the ebna- protein. reed sternberg cells expressing lmp can be a target for antigen-specific immunotherapy, but lmp -specific autologous ctl for adoptive immunotherapy are difficult to generate due to poor immunogenicity. t cell receptor (tcr) gene transfer using retroviral vectors containing the tcr alpha and beta chain genes can reproducibly redirect the antigen specificity of a t cell population. the aim of this study was to generate a retroviral tcr construct suitable for the rapid and efficient production of lmp -specific ctl. retrovirally introduced tcrs compete with endogenous tcrs for cd molecules required for assembly of the tcr complex. this competition may limit surface expression of the introduced tcr resulting in a transduced t cell with poor functional avidity. in an attempt to generate a 'highly competitive' lmp -tcr the following modifications were made to the retroviral vector construct: i) nucleotide sequences were codon-optimised for efficient translation in human cells; ii) the constant region of each tcr chain was altered to contain murine sequences to enhance cd binding; and iii) the tcr alpha and beta chain genes were linked by a self-cleaving a sequence. the unmodified hla-a -restricted lmp -specific tcr was poorly expressed in primary human t cells (up to . % of viable cd + t cells, as detected by facs analysis using monoclonal anti-vbeta antibodies), suggesting that it competed inefficiently with endogenous tcr chains for cell surface expression. however, retroviral transfer of the modified lmp -tcr into human t cells improved lmp tcr expression to - % cd + t cells. the transduced cells bound hla-a /lmp pentamer, showed peptide-specific ifngamma and il production and killed target cells displaying the lmp peptide. importantly, expression of the introduced lmp -tcr suppressed expression of almost the entire repertoire of endogenous tcr combinations, including 'mis-paired' tcrs. 'mis-paired' tcrs contain an introduced alpha chain paired with an endogenous beta chain and vice versa. the antigen specificity of such mispaired tcrs is unknown and could be auto or allo-reactive. modified tcr sequences, producing 'dominant' tcrs may improve the efficacy and reduce the potential risks of tcr gene therapy a novel form of adoptive immunotherapy. allogeneic stem cell transplantation in patient with major histocompatibility complex class ii immunodeficiency: a single-centre experience h. al-mousa*, z. al-shammari, a. al-ghonaium, h. al-dhekri, s. al-muhsen, r. arnaout, a. al-seraihy, a. al-jefri, a. al-ahmari, m. ayas, h. el-solh king faisal specialist hospital (riyadh, sa) background: major histocompatibility complex class ii (mhc ii) deficiency is a rare combined immunodeficiency disease. allogeneic bone marrow transplantation (bmt) is considered the only available curative treatment. survival rate post bmt is lower than other forms of primary immunodeficiencies. these differences were observed for both bmt performed with matched and non-matched donors. patients and methods: between june and august , thirty two children with mhc ii deficiency underwent thirty seven bmt procedures at king faisal specialist hospital and research centre, riyadh, saudia arabia. five patients required second bmt trial. median age at bmt was months (range, - months). the source of stem cells was unmanipulated marrows from hla-genoidentical siblings in pts, hla-phenotypically identical related donors in pts and umbilical cord for patient. conditioning was with one of regimens, regimen a ( pts): busulfan (bu), cyclophosphamide (cy) and etopside (vp- ), regimen b ( pts): bu/cy and anti-thymocyte globulins (atg), regimen c ( pts): bu/cy/vp- and atg and regimen d ( pts): fludarabine, melphalan and atg. median cd dose was . x /kg (range, . - . x /kg). graft-versus-host disease (gvhd) prophylaxis consisted of cyclosporine (csa) and methorexate (mtx) in pts, csa alone in pts and csa and steroid in patient. results: pts had adequate immune reconstitution and sustained engraftment (assessed by short tandom repeats) ranged from - % for lymphoid line and - % for myeloid line. seven pts ( pts were post second transplant) died secondary to sepsis, multiorgan failure and primary disease. acute gvhd was seen in pts and pts developed chronic gvhd. the overall disease free survival rate was % with a median follow up of . years (range . - . years). low survival rate ( %) was seen in patients who underwent second bmt. conclusion: bone marrow transplantation (bmt) can cure the disease, provided it is performed before complications leading to severe organ failure develop. second bmt is associated with high rate of mortality. further studies and long term follow up are required to determine the appropriate conditioning regimen. to analyze the effects of cd -trail+ cells on tumor vasculature, tumorbearing mice were perfused with sulfo-biotin and tumor endotelial cells (tec) were then revealed by horseradish peroxidase (hrp)-conjugated streptavidin. as compared with cd -mock-or soluble (s)trail-treated mice, a -hour treatment with cd -trail+ cells significantly (p ≤. ) reduced microvessel density ( ± vs ± vs ± vessel per x tumor cells, respectively) and increased the thickness of the vessell wall ( . ± µm vs . ± µm vs ± µm, respectively), suggesting that cd -trail+ cells induce an early vascular disruption leading to a progressive disintegration of the vascular bed. confocal microscopic imaging of tumor sections double-stained with anti-cd and anti-trail-r showed that this receptor was expressed by - % of large tumor vessels. interestingly, upon treatment with cd -trail+ cells, but not strail, tunel staining revealed an extensive apoptosis of tec. forty-eight hours following injection of cd -trail+ cells, a -fold increase of apoptotic index was detected, which was associated with extensive necrotic areas ( % to % of tissue section). these data show that: (i) tumor homing of cd -trail+ cells induces extensive vascular damage, hemorrhagic necrosis and tumor destruction; (ii) the antitumor effect of cd -trail+ cells is mediated by both indirect vascular-disrupting mechanisms and direct tumor cell killing. targeting of a therapeutic suicide gene to human alloreactive memory t-cells with stem-cell features requires il- and il- a. bondanza* ( ), l. hambach ( ) in a phase ii clinical trial investigating the prophylactic infusion of suicide gene-modified donor t cells after haploidentical hemopoietic cell transplantation (haplo-hct), we observed a rapid and effective immune reconstitution. after activation with anti-cd antibodies, t cells were modified with a retroviral vector (rv) encoding for the herpes simplex thymidine kinase (tk). tk+ cells displayed an effector memory (em) phenotype (cd ra-cd l-, cd ±cd +, il- ±ifn-g+). when needed, graft-versus-host disease (gvhd) was controlled upon administration of the prodrug ganciclovir (gcv). the graft-versus-leukemia (gvl) effect was substantial in patients transplanted in remission, but failed to cure patients in relapse. genetic modification with rv is limited to memory t cells. em tk+ cells have a reduced alloreactivity. central memory (cm) t cells (cd ra-cd l+, cd +cd +, il- +ifn-g±) share many characteristics with stem cells, namely the ability to selfrenew and to differentiate into effector cells. recently, it has been proposed that alloreactivity may be confined to memory t cells with stem-cell features. since alloreactivity is the common ground of both graft-versus-host disease (gvhd) and the gvl effect, crucial to the success of the strategy is the suicide gene-modification of cells with such properties. we found that addition of cd costimulation on cell-sized beads and the use of homeostatic cytokines, such as il- and il- , generates central memory (cm) tk+ cells. cm tk+ cells were highly alloreactive, both in vitro and in vivo in a humanized animal model of gvhd based on the grafting of human skin onto nod/scid mice. gcv administration abrogated gvhd. stimulation of cm, but not of em tk+ cells with autologous dendritic cells pulsed with hla -restricted peptides from the minor histocompatibility alloantigen (mhag) ha- or h-y efficiently induced mhag-specific t cells that lysed natural ligand expressing hla-a + targets. a fraction of mhag-specific tk+ cells expressed il- ra. only il- ra+ mhag-specific tk+ cells could self-renew and differentiate into effector cells. when infused in nod/scid mice harboring human mhag+hla-a + leukemia, tk+ mhag-specific t cells significantly delayed disease progression. altogether, these data suggest that targeting of a suicide gene to human alloreactive memory t cells with stem-cell features requires il- and il- and warrant their use in the clinic for a safe and powerful gvl effect. loss of foxp expression after in vitro expansion of human cd +cd +cd -regulatory t-cells p. hoffmann* ( ), t.j. boeld ( ) , r. eder ( ) , j. huehn ( ) , s. floess ( ) in animal models the adoptive transfer of donor-type cd +cd + regulatory t cells (treg) protects from graftversus-host disease (gvhd) after allogeneic stem cell transplantation (sct), as shown by us and several other groups. exploring this strategy in human sct, we currently perform a first phase i clinical trial using freshly isolated treg. for future trials requiring large cell numbers for repetitive treatments, we described in vitro culture conditions that permit a more than -log polyclonal expansion of treg (blood : ; ) . a highly enriched starting population proved to be crucial for the generation of pure treg cell products, a criterion fulfilled by the naïve, cd ra+ subpopulation of cd +cd high t cells (ra+ treg) (blood : ; ). an alternative isolation strategy for treg relies on the exclusion of cd + cells, as activated cd +cd + conventional t cells express high levels of cd while cd +cd + treg show no or only weak expression. for a direct comparison of the two isolation protocols, we sorted cd +cd +cd low/neg t cells (cd -treg) and ra+ treg from the same leukapheresis products and analyzed the cells after and weeks of expansion. whereas both populations were > % foxp + upon isolation, only ra+ treg maintained foxp expression throughout the expansion period ( % [range: to %; n= ] after and % [range: to %; n= ] after weeks). in contrast, cd -treg cultures contained only % (range: to %; n= ) foxp + cells after weeks and highly variable and significantly lower numbers of foxp + cells than ra+ treg cultures after weeks ( % [range: to %; n= ; p= . ]). further analysis identified cd ra-foxp + memory-type cells within the cd -treg starting population as a major source of foxp -as well as il- -and ifngamma-producing cells emerging during in vitro culture. in addition, we analyzed the dna methylation status of a defined region within the foxp locus termed tsdr (t reg-specific demethylation region), which has been shown to be demethylated exclusively in natural t reg cells (baron et al., eji : ; ) . upon isolation, ra+ and cd -treg showed complete demethylation of the tsdr, but only ra+ treg maintained this demethylated status throughout the entire culture period. based on these findings, we suggest that expansion of naive cd ra+cd +cd high t cells represents the most promising strategy for adoptive treg cell therapies. several eortc aml trials were based on a x factorial design. in elderly patients, aml- trial was set-up to assess the value of g-csf during/after induction, and of infusional vs non-infusional mini-ice as consolidation. in the eortc-gimema aml study (patients ≤ years old), comparing anthracyclines in induction and consolidation, the value of allosct vs autosct has been assessed based on intent-totreat analysis. the "donor vs no donor" comparison was performed, overall, and according to cytogenetic subgroups and age. in the eortc-gimema aml (age ≤ yrs) study the accrual for the st question (hd-ara-c vs sd-ara-c in induction) had to be expanded, due to insufficient number of patients randomized for the nd one (il- vs no il- after autosct). new statistical considerations were set-up for the st question as the trial expanded to patients. statistical planning of clinical trials: an emphasis on sample size determination a. latouche* ebmt (paris, fr) an essential step when planning a trial is the calculation of the sample size or the number of patients to recruit to detect a relevant effect with sufficient power. patients enrolled in a clinical trial may experience exclusive failure causes, which defines a competing risk setting. the main decision regards the relevant quantity to assess the treatment effect: either cause-specific hazard or cumulative incidence function. the analysis should then be performed according to the method used for sample size computation, if one does not want an under-powered trial. from practical point of view, the practitioner has different options to plan a study accounting for competing risks. in this work, we thus compare these approaches. in medical applications multi-state models arise in attempting to understand complex disease or treatment processes. in these models individuals can move among a finite number of states defined by specific conditions of health, often including death. they are natural extensions of the traditional survival models in which transitions between multiple states are considered. multi-state models give more insight into the disease-recovery process because they take into account the occurrence intermediate events. in this talk i discuss some of the advantages (and disadvantages) of multi-state models and their application to bone-marrow transplantation. long-term follow-up of hcv infected patients; updated results of the ebmt prospective study p. ljungman*, a. locasciulli, a. békássy, l. brinch, i. espigado, a. ferrant, i. franklin, v. gomez-garcia de soria, j. o'riordan, m. rovira, p. shaw, h many long-term survivors after sct are infected with hcv. it has been reported that a high proportion of these patients develop late occurring liver cirrhosis. treatment options include interferon with or without the addition of ribavirin but the knowledge about these options' effectiveness is limited. therefore in , the idwp of the ebmt initiated a prospective study of hcv infected sct patients having survived at least months after sct. the intention is to every five years ask for follow-up on living patients. patients were included in the study cohort between and . this is the rd report of this cohort. due to limited follow-up data, patients have been excluded. thus, the report is based on patients with a median follow-up of years ( . - . ). the kaplan-meier estimate of survival is . %. patients have died of whom have been reported to have died from liver associated complications including one after liver transplantation. two additional patients are alive after liver transplantation. patients have been treated with interferon with or without the addition of ribavirin or with ribavirin alone. / patients became hcv pcr negative of whom have relapsed with positive hcv pcr. the side effect profile was similar to other patient populations treated for hcv. conclusion: in this prospectively followed cohort, the prognosis of chronic hcv infection was quite favourable. treatment is feasible and associated with acceptable efficacy and toxicity. current status of the apbmt registry y. kodera*, a. yoshimi, r. suzuki, y. atsuta, l.l. chan, a. li, p.-l. tan, w.y.k. hwang, t.v. binh, t.j. chiou, a. ghavamzadeh, l. dao-pei, p.- the data was submitted through the national registries in my, jp, and tw. in other countries/regions, the data was collected either by apbmt data center or by regional coordinators. the total number of hsct has been steadily increasing in most of the countries/regions and most dramatically in cn and in ir. the annual number of hsct of reported was , (data of countries/region), which was doubled in these years; of those, % were allogeneic and % were autologous. the number of allogeneic sct has been steadily increasing year after year; meanwhile the number of autologous sct has been recently stable. the distribution of related and unrelated donors (ud) differed widely among the countries (ud= % [vn] to % [jp] in ) . ud-sct is increasing consistently in cn, jp and hk, but not in other countries/regions. interestingly, cord blood appeared to be a common stem cell source in asia, accounting for % of ud-hsct ( % in cn to % in ir/vn, data of ). unrelated donor peripheral blood stem cell transplantation is not common in asian countries/regions except for china. in summary, this simple survey provided us basic information on current situations and trends of hsct in asia, which may be helpful to advance the patient registry. recently apbmt developed an electric data collecting system called trump and the group also agreed to share the common basic survey items with ebmt and cibmtr (med-a/ted) and join the global transplant activity survey under the umbrella of world-wide group for blood and marrow transplantation (wbmt). these international collaborations may facilitate future advances of the hsct. background: ebv associated ptld is a serious complication after sct.several risk factors may increase the risk of ptld. analysis of ebv dna viral load has been suggested to be useful for monitoring and used as the basis for preemptive therapy with rituximab with the aim to reduce the risk for ptld. methods: patients who underwent sct from until were included in this analysis. ebv serological mismatch between donor and recipient, primary ebv infection, cord blood grafts, and diagnosis of lymphoma were regarded as risk factors and used to split the patients into a high risk and a standard risk group. before (early group), patients were tested on clinical suspicion of ebv associated symptomatic infection while after (recent group) high risk patients were monitored by quantitative pcr while standard risk patients were to be sampled on clinical suspicion of symptomatic ebv infection only. patients were analyzed; patients in the early and in the recent group. in the early group were classified as high and as standard risk. in the recent group, were defined as high and as standard risk. the ebv viral load was measured in serum by a taqman pcr technique. after , rituximab was to be given in high risk patients at an ebv dna level of > . copies/ml. in the early group and in standard risk patients of the recent group, rituximab was given on suspicion of ebv symptomatic infection. results: a total of blood samples were analyzed for ebv dna. more samples (median in the early and in the recent group) were taken in the high risk compared to the standard risk group (median and samples, respectively). in the early group, / ( %) high risk patients compared to / ( %) standard risk patients had ebv detected at least once. the corresponding numbers in the recent group were / ( %) in the high risk compared to / ( %) in the standard risk group. in the early group, / ( . %) received rituximab. while / ( . %) in the recent cohort was given rituximab. high risk patients received more often rituximab than standard risk patients in both the early and recent cohorts (data not shown). in the early group ( %) developed ptld of whom died ( . %) while in the recent group, / ( . %) developed ptld of whom died ( . %). conclusions: a strategy of targeted monitoring to high risk patients can be safely utilized with a low risk for development and fatal outcome of ptld. a retrospective ebmt survey on the use of cidofovir for bk-related haemorrhagic cystitis after allogeneic haematopoietic stem cell transplant s. cesaro* ( ), y. koc ( ) bk virus has been recently associated to post-engraftment hc. cidofovir (cdv) is often used for bk-related hemorrhagic cystitis (bk-hc) treatment although data on safety and efficacy are scarce for this indication. we collected retrospectively the experience of cdv for bk-hc among the ebmt centres. episodes of bk-hc in patients from centres were recorded. all patients but had been transplanted for a malignant disease. median age at haematopoietic stem cell transplant (hsct) was yrs (range - ) and / patients were children or adolescent (age < yrs). the source of stem cell was pb in %, bm in %, and cb in % whilst the type of donor was sibling in %, unrelated donor in % and other related donor in %. myeloablative regimen + tbi was used in % of patients. bk-hc occurred after a median of days from hsct (range - ) but only episodes were diagnosed after day + . hc was scored as follows: grade i, grade ii, grade iii, grade iv, and lasted in median days (range - ). concurrent morbidity was represented by fever in %, hypertension in %, vod in % and ttp in % of episodes. most of patients received hyperhydration plus rc and plt transfusion during hc whilst only of episodes were treated with bladder irrigation through vesical catheter. cdv was started a median of . days after hc (range - before to days after hc) and was administered for a median of doses (range - ). the most frequent schedule was - mg/kg/weekly or fortnightly with probenecid (used in % of all treatments). patients received intravesical cdv via catheter. cdv toxicity was reported in % of episodes and was limited to kidney: grade i, grade ii, grade iii, grade iv. complete resolution or clinical improvement of hc was reported in ( %) and ( %) of episodes, respectively. although the datum is not available for all episodes cdv treatment was associated to negativization of bk viremia in % ( / ) and of bk viruria in . % ( / ) of episodes. we conclude the cdv therapy is a potential useful option for the treatment of bk-hc with limited moderate or severe kidney toxicity. prospective data are needed to define the best timing and schedule of treatment and the predictive factors for clinical and virological response. single-centre prognosis analysis and validation of the seattle, french and strasbourg prognosis indexes of invasive aspergillosis in adult patients with haematological malignancies or after haematopoietic stem cell transplantation r. parody*, r. martino, f. sanchez, j.l. piñana, j. sierra sant pau hospital (barcelona, es) in this retrospective study we analyzed the outcomes of adults hematological patients who had a history of proven (n: ), probable (n: ) and possible (n: ) invasive aspergillosis (ia) between january and june , at santa creu i sant pau's hospital of barcelona. forty-one patients ( %) were recipients of an allogeneic hematopoietic stem cell transplantation (allohsct). the main goal of the study was to analyze the prognosis factors predicting the outcome of -month aspergillosis free survival (afs) and overall survival (os), in order to compare the results with previously published prognosis indexes. analysis was performed in all patients and separately in allohsct and non-allohsct patient cohort. a total of sixty seven patients ( %, recipients of allohsct)) died in the first months after ia diagnosis [median days: (range - )]. invasive aspergillosis was identified as the main cause of death in patients ( %, recipients of allohsct) [median days: (range ]. according with autopsy findings, cases ( possible and probable) could be recategorized as proven cases. diagnosis of ia at or before had a negative impact in both -month afs and -month os. in all patients and allo-hsct patients, variables (excluded the year of diagnosis) decreased -month afs in multivariate analysis: (i) disseminated ia and monocytopenia < . x /l, respectively (ii) impairment of one organ, (iii) impairment of or more organs and (iv) high-doses steroids and an alternative donor, respectively. a risk model for progression was generated for each group, according with the presence of - factors,( and % afs), - factors, ( and % afs) or factors, ( % afs)[p< . ]. in base of similar results previously published our prognosis index model could be validated with both the french and seattle models for allo-hsct recipients and with the strasbourg model for hematological patients. one-year microbiological survey with molecular typing method of pseudomonas aeruginosa in a bmt unit b. bartolozzi* ( ), r. fanci ( ) pseudomonas aeruginosa is one of the most common nosocomial pathogens in intensive care and in oncohematological units and it still represents an important cause of morbidity and mortality. molecular epidemiological survey is a very important tool to understand the nosocomial pattern of each hospital and to identify outbreaks. from may to june we performed a "real time" aflp (amplified fragment-length polymorphism) monitoring of all p.aeruginosa strains isolated in patients within the bmt unit of florence. the patients admitted in the unit during this period were ( autologous transplants, unrelated allogeneic transplants and identical sibling allogeneic transplants. p.aeruginosa was responsible for ten bloodstream infections (bsis) and of two microbiologically documented infections without bacteremia. the aflp analysis showed the persistence within the unit of two clones of p.aeruginosa. one was responsible of two outbreak episodes in patients allocated in the same single room. the first episode occurred in may-october involving patients; a team consisting in the bmt head physician, the bmt head nurse, the infection control officer and the infection control physician was actively involved in a strict bacteriological surveillance that allowed to isolate the same strain from a irgasan soap sample. for this reason, hyperclorination of the water network, sink tap changing, water filters installation and weekly soap monitoring were performed. nevertheless the same clone was isolated after several months in a water sample, showing the inefficacy of the control measures taken. in may-june the same p.aeruginosa clone was isolated in three infected patients and in a shower tap sample. hydraulic works inside the room were performed and until now no other case of p.aeruginosa was observed. the other persistent clone was responsible of contamination of sample of irgasan soap taken from the ward cloakroom (collected from november until april ) and of a bsi in a patient (march ) . our results showed that p.aeruginosa is largely and consistently found in environment which may represent an important source of infection, difficult to eradicate. moreover, punctual microbiological surveillance and molecular typing methods are essential to early detect nosocomial outbreaks, to identify p.aeruginosa reservoir and to guide in decision making in order to understand and possibly stop the epidemic chain. donor lymphocyte infusion (dli) following t cell depleted allogeneic stem cell transplantation (sct) is an effective treatment for relapsed hematological malignancies. dli often results in a profound graft versus leukemia (gvl) effect with relatively limited graft-versus-host disease (gvhd). both gvl reactivity and gvhd are likely to be mediated by donor derived t cells recognizing polymorphic minor histocompatibity antigens (mhag) on patient cells. it has been hypothesized that t cells recognizing mhags expressed on non-hematopoietic cells are responsible for gvhd, whereas t cells recognizing hematopoiesis restricted mhags may be selectively involved in gvl reactivity. to analyze the diversity of alloreactive t cells involved in gvl and gvhd we analyzed the immune response in a patient responding to dli following hla-matched sct for aml. six weeks after dli, gvhd limited to skin and mouth developed coinciding with a rapid and sustained conversion to % donor chimerism. we clonally isolated and characterized activated hla-dr expressing t cells at the onset of gvhd ( % of the circulating t cells). in total cd + t cells clones and cd + t cell clones were tested for alloreactivity as defined by cytotoxicity or ifng production upon recognition of patient derived ebv-lcl and not donor derived ebv-lcl. % of the cd + t cell clones and % of the cd + t cell clones were identified as alloreactive t cell clones. next, recognition of patient skin-fibroblasts as a target for gvhd was determined both in the absence or presence of ifng to mimic an inflammatory environment. none of the t cell clones recognized unmanipulated fibroblasts. only after co-culture with ifng, % of the alloreactive cd + t cell clones but none of the cd + t cell clones reacted with patient fibroblast. by blocking studies, panel studies and t cell receptor-vb analysis the diversity of this immune response was determined. and different reactivities were found for the cd + t cells and cd + t cells, respectively. in conclusion, isolation of activated t cells identified a very polyclonal immune response directed against multiple mhags in all hlaclass i and hla-class ii alleles. despite the polyclonality of this immune response, reactivity of the t cell clones was relatively hematopoiesis specific, resulting in % donor chimerism, persistent complete remission and only moderate gvhd limited to the skin. since t cell recognition of fibroblasts was observed only upon co-culture with ifng, this may reflect a secondary reactivity induced by the ongoing gvl effect. background and aims: chronic graft-versus-host disease (cgvhd) is a recognized cause of genital complications in the vulva and vagina in female patients (pts) after allogeneic stem cell transplantation. however, clinical features and consequences of genital cgvhd are poorly described in the literature. in a retrospective study, we assess prevalence, symptomatology and effect on sexual life of genital cgvhd. in a prospective study, we try to prevent progression and sequelae of genital cgvhd. here we report early results from these ongoing studies. patients and methods: all women allografted - in the western region of sweden are asked to participate in a study comprising (i) structured anamnesis and validated questionnaires for the identification of depression and sexual dysfunction; (ii) gynecological examination; (iii) biopsies for pathological examination. prospective pts are seen by the gynecologists up to years post-transplant. clinical findings of genital cgvhd is a dry, thin, painful mucosa, inflammation, lichenoid bands, vulvar synechiae and vaginal stenosis. systemic and local estrogens do not resolve those signs. histology findings were those of chronic inflammation, lichenoid reaction and fibrosis. results: fifty-eight consecutive pts with - yrs of follow-up were approached and so far women have been examined. median age was ( - ) yrs) and follow-up post-transplant was . ( - ) yrs. fifteen pts ( %) had a clinically diagnostic cgvhd, and using pathology examination (n= ) the diagnosis was confirmed (n= ) or suspected (n= ). another of the pts had signs suggestive of cgvhd and histology was confirmatory in one and suspect in cases. two of pts had vulvar synechiae and ( %) had vaginal adhesions. in the prospective study, pts have been included and the follow-up is now ( - ) months. after months of follow-up (n= ) clinically cgvhd had been diagnosed or suspected in cases. local corticosteroid or takrolimus cream was commenced in / cases. female sexual distress scale intervention performed as sceduled revealed sexual dysfunction at at least once in ( % of all) and beck depression inventory indicated depression ( %). conclusions: genital signs and symptoms, including vaginal stenosis, are common features of cgvhd and are associated with sexual dysfunction and depression. gynecological surveillance and early intervention may reduce the risk of severe sequelae after genital cgvhd. langerhans cell chimerism early after t-cell depleted allogeneic haematopoietic stem cell transplantation k. schneiker*, t. schmitt, a. konur, j. hemmerling, k. bender, e. von stebut, a. hadian, k. kolbe, c. huber, w. herr, r.g. meyer university clinic mainz (mainz, de) skin is the most frequently affected organ in acute graft versus host disease (gvhd). data from murine studies suggest that the interaction of residing host epidermal langerhans cells (lc) and donor t cells is crucial for the initiation of acute gvhd. in an ongoing clinical protocol applying alemtuzumab-based t cell depleted (tcd) allogeneic stem cell transplantation (sct) we observed acute skin gvhd occurring very early after transplantation despite of low t cell counts in the peripheral blood (meyer, blood ; : ) . we therefore intended to analyze the lc chimerism in these patients. up to now, lc-chimerism analysis in humans has been performed by the detection of the sex-chromosomes restricting it to sex-mismatched donor / recipient pairs. in our patient-population, this would limit the analysis to less than / of the patients. consequently, we aimed at a method to isolate lc from small skin samples with high purity for a sensitive str-based chimerism analysis of general applicability. epidermal skin layers were prepared from mm punch biopsies by digestion with dispase i. they were further split and used for both immunofluorescent staining and digestion with trypsin to generate a single cell suspension and cd a/mhc-class ii-positive lc were subsequently sorted by flow cytometry. the density of lc on day + after hsct was much lower compared to before transplantation or to that of healthy individuals. but still, lc could be purified in all analyzed patients. the isolated lc numbers ranged from to > . we confirmed that the purity was exceeding . % in patients in a facs-reanalysis, thereby reproducing the findings with skin of healthy individuals. applying two alternative str-based protocols, we obtained reliable results for lc chimerism in of patients and could detect signals with as few as isolated cells. in patients, the majority of isolated lc was of donor origin whereas the other patients had predominantly host lc. after day + post hsct, further patients showed only a few remaining lc of host origin. in summary, we have established a sensitive method that enables the chimerism analysis on highly purified lc independent of sex-mismatched donor / recipient pairs. our results on a few patients' samples cannot yet be related to clinical events. but the method allows the investigation of lc´s chimerism and potentially of other tissue-resident antigen presenting cells to study their impact on gvhd in humans. we and others have data on the activation of coagulation and fibrinolysis during and after allogeneic stem cell transplantation (sct), which may have pathogenic implications in the subsequent recovery. we characterised the outcome including graft versus host disease (gvhd) in association with adaptive mechanisms of anti/coagulation after allogeneic sct in patients with a hematological malignancy. they were given myeloablative conditioning with cyclophosphamide and total body irradiation. patients received the transplant from a sibling and from an unrelated donor. gvhd prophylaxis consisted of cyclosporine were "unresponsive, non-progressive". the prognostic features, including cytogenetics, were similar in both groups. % of the patients responded to the first hdt (cr/ncr %, pr %, mr %). patients were given a second transplant ( "auto", "allo"). % who received a second "auto" up-graded their response (cr %, pr %, mr %) while % who underwent "allo-ric" increased their response (cr %, pr %). median survival of the whole series was years. patients progressing while on therapy had a shorter survival than the "no-change" group (median yrs vs not reached, p= . ). finally, the "non-responsive, nonprogressors" patients had similar survival than the with chemosensitive disease intensified with hdt. conclusions: ) hdt in patients with primary refractory mm results in a low cr rate, ) patients progressing while on initial therapy have a short survival despite the intensive approach and ) patients with "non-responding, non-progressive" disease have similar survival than chemosensitive patients. whether this good outcome is due to the impact of hdt or to the natural history of a more indolent disease remains to be further investigated. background: primary amyloidosis (al) responds poorly to conventional therapy and has poor prognosis. autologous stem cell transplantation (asct) results in a significant response rate although the procedure is hampered by a high transplant-related morbidity and mortality. aim. to analyze the outcome of a series of patients with al who underwent asct at a single institution during a years period. patients and methods: thirty four patients ( m, f; median age years, range: - ) who received an asct between november and september were included. fourteen patients had received previous therapy and were newly diagnosed. in % of the patients the light chain was of lambda type. the median number of involved organs was (range, - ) including kidney ( patients), heart ( ), liver ( ) , peripheral nerve ( ), autonomic system ( ) and gastrointestinal tract ( ). thirty-eight percent of the patients had more than organs involved. all patients were mobilized with g-csf alone and the intensive regimen consisted of mel- in patients and mel- in patients. the median interval between diagnosis and asct was months. results: the overall transplant-related mortality (trm) was %. there was a trend towards a higher trm in patients transplanted from november -april (n= , trm: %) as compared to those transplanted from june -sept (n= , trm %) (p= . ). in the patients who underwent asct before nov. (minimum follow-up of year) the response and survival on an intent-to-treat basis were as follow: cr ( %), pr ( %), no response ( %), early death ( %).organ response was observed in patients ( %). the median response duration and the overall survival were and months, respectively. favorable prognostic features for survival were: cardiac septum < mm (p= . ), normal beta m (p= . ) and normal nt-probnp (p= . ). updated results on the overall series of patients will be presented at the meeting. conclusion: ) asct in al results in significant cr rate with prolonged response duration an overall survival and ) although trm is high, there is a trend towards a lower trm over the years this most likely reflecting both a better patient´s selection and general management. outpatient-based peripheral blood stem cell transplantation for patients with multiple myeloma a. ghavamzadeh, m. khani*, a. karimi, k. alimoghadam, a. manokian, r. maheri, m. asadi, f. afshar, a. shamshiry hematology, oncology and bmt resarch center (tehran, ir) intrduction: the aim of this study was to explore the feasibility and safety and cost-benefit of performing asct on an outpatient basis. material and methods:total of patients affected by mm and in complete remission (cr) or partial remission (pr) were selected to receive asct on an out-patient or in-patient basis . in the in-patient group , patients received mg/m² and mg/m² melphalan as conditioning regiment respectively . in out-patient group patients received mg/m and patients recived mg/m melphalan. in out-patient group all the patients were programmed to go home the day after asct and to be rehospitalized in the case of febrile neutropenia or other sever toxicities. we used caregiver, general physician, staff nurse as an out-patient and visit team and also unequipped routine house of the patients during neutropenia. results :median ages were ± . years, median hospital stay were , . days in in-patient and out-patient respectively. there were not significant difference between these groups in aphresis days ,granulocyte colony stimulating factor(gcsf) requirement for mobilization and mononoclear cell (mnc) or cluster of differentiation(cd) + cell parameters (p< . ), but statistically significant reduction in total cost and hospitalization (p< . ).there were also significant reduction (p< . ) in parentral antibiotic,blood product requirement and need for total parentral nutrition. conclusion :many different authores have explored the feasibility of autografting patient on an outpatient basis. the ease of administration of hdm as well as the lack of excessive extramedullary toxicity, including nausea and vomiting renders patients with mm more suitable for outpatient management, in the present study, we describe an outpatient program based on management of the patient in his/her house during aplastic phase . our results clearly indicate that such a proceder is feasible and safe in a patient population with a median age of years. with an accessible caregiver, the most frequent cause of readmission in other study were febrile neutropenia and sever mucositis need tpn. in particular, it is worth nothing that transplant cost, requirement for blood product and hospital stay decreased significantly in this method of transplantation. treatment of multiple myeloma with sequential autologous and low dose total body irradiation based allogeneic unrelated sct c. pfrepper*, t. lange, r. krahl, m. cross, h.-k. al ali, w. pönisch, n. basara, d. niederwieser university hospital of leipzig (leipzig, de) purpose: the limited availability of related donors poses an important limitation in the treatment of multiple myeloma (mm). the use of unrelated donors would alleviate this problem. furthermore, outcome after unrelated transplants following reduced intensity conditioning (ric) has been shown to be superior in other diseases including cll and aml, due to increased graft-versus malignancy reaction. in this study, the feasibility of autologous followed by allogeneic unrelated ric sct was tested and the results compared to autologous followed by related allogeneic ric sct and allogeneic ric sct after relapsing following autologous sct. patients and methods: we report the outcome of mm patients with a median age of (range - ) years treated with high dose chemotherapy (melphalan mg/m² on day - ) followed by autologous sct. subsequent ric-sct was performed using fludarabine ( mg/m²/d from days - to - ) and tbi ( gy on day ) followed by cyclosporine ( . mg/kg twice daily from day - ) and mycophenolate mofetil ( mg/kg daily from day ). eleven patients received an unrelated graft (group ) and a related graft (group ) within median (range - ) months from the autologous hct. ten further patients (group ) underwent either related or unrelated sct (range - ) months after insufficient response following autologous transplantation and additional chemotherapy. results: durable engraftment was obtained in % of all patients. overall survival (os) at years was ± %, ± % and ± % (p= . ) in groups , and , respectively at a median follow up of (range - ) months. nine patients ( %) were in cr, patients ( %) in pr, patients ( %) had progressive disease after allogeneic sct. however; progression free survival (pfs) was ± %, ± % and ± % in the three groups, respectively (p= . ). non relapse mortality (trm) was not different between the three groups, but relapse incidence was lowest in patients after unrelated transplantation. conclusion: we conclude that autologous sct followed by low dose tbi based sct from matched unrelated donors provides rapid and sustained engraftment for mm patients comparable to that of related donors. our limited data suggest a tendency to higher pfs following unrelated compared to related sct. these results provide the basis for a phase iii study designed to compare auto-allo unrelated to auto-auto sct. background: a significant proportion of patients with multiple myeloma have a long-lasting response after autologous stemcell transplantation (asct). however, others relapse relatively quickly. the aim of this study was to determine if the presence of monoclonal plasma cells (mpc), detected by flow cytometry, in the apheresis product, and in pre-and posttransplant bone marrow samples predicts a shorter time to progression (ttp). patients and methods: we included patients diagnosed with multiple myeloma and treated with high dose therapy followed by asct between november , and february , . in all casas, flow cytometric analysis was performed in the apheresis products and in bone marrow samples, both prior and after asct. the variables evaluated as possible prognostic factors were: age, sex, type of monoclonal component, durie-salmon stage, iss stage, presence of mpc (cd +/cd +/ cd -/cd -or cd dim) in apheresis products, and in bone marrow samples ( days before, and days after asct), and accomplishment of a complete response prior or after asct. results: patients were included: male, female. median age at asct was years (range, - years). twenty-nine patients had progressed. on univariate analysis, age . ); p< . ], iss stage [or . ( . - . ); p= . ], mpc in apheresis products (p= . ), mpc in pretransplantation bone marrow (p= . ), mpc in postransplantation bone marrow (p= . ), and the achievement of complete response prior and after asct (p= . ) had a negative impact on time to progression. on multivariate analysis, the presence of mpc in apheresis [or . ( . - . ), p= . ] and the presence of mpc in pretransplantation bone marrow (or . , p= . ) were identified as independent predictors of a shorter ttp. conclusions: the presence of mpc in apheresis products and in pretransplantation bone marrow were identified as independent predictors of shorter ttp. both parameters can identify a group of patients with multiple myeloma which could benefit from most aggresive conditioning regimens or additional maintenance therapies. long-term follow-up of patients with systemic al amyloidosis treated with high-dose melphalan and autologous stem cell transplantation after induction and mobilisation chemotherapy s. o. schonland*, t. bochtler, m. hansberg, a. mangatter, j.b. perz, a.d. ho, h. goldschmidt, u. hegenbart university of heidelberg (heidelberg, de) introduction: longterm survival in al amyloidosis (al) patients (pts) has been shown recently after high-dose melphalan (hdm) (sanchorawala, blood ) in % of the patients. however, no advantage of hdm compared to conventional chemotherapy has been observed in a randomized multicenter trial (jaccard, nejm ) . this was probably due to high transplantation related mortality (trm) and a substantially longer time to the start of chemotherapy in the hdm group. whether treatment intensification using induction and mobilization chemotherapy prior hdm can improve results has not yet been definitively shown. methods: we have updated pts with al (perz, bjh ) who received vincristine/adriamycine/dexamethasone as induction and additional mobilization chemotherapy (mostly cyclophosphamide based) prior to hdm in our center from until . inclusion criteria were age < years, nyha stage < iii and a who performance status < . median age was years; median number of involved organs was . fifteen and pts had symptomatic renal or cardiac involvement, respectively (on dialysis at hdm, n= ). results: the median overall survival (os) has not been reached at a median follow up of months post hdm (figure). trm was %. a hematological response (hr) to induction and mobilization chemotherapy occurred in % ( % of pts achieved a complete remission (cr)). hr and cr rate increased to % and % after hdm, respectively. median time to cr from hdm was months. two patients had already an organ response (or) after induction and mobilization chemotherapy. or was observed in % and the median time to or was months post hdm (range - months). pts with cr have an estimated os of % after years; for pts not reaching cr the median os is months (p= . ). hematological relapse or progression occurred in about half of the patients; however pts ( %) are still in first cr with a sustained or and a median follow-up of almost years. conclusion: we observed a very high cr and or rate using this intensive treatment approach. patients with cr after hdm have an excellent survival. therefore, the role of induction therapy in al, e.g. with bortezomib or lenalidomide, should be further investigated in randomized trials in experienced centers. the main goal of treatment in al remains achievement and maintenance of a cr of the underlying plasma cell disorder. the results of reduced-intensity conditioning allogeneic stem cell transplantation (ric allo-sct) for multiple myeloma (mm) are still under considerable debate. while ebmt data did not support the universal use of ric for mm allografts, the italian randomized multicenter study suggested that in newly diagnosed myeloma, survival in recipients of a hematopoietic stem-cell autograft followed by ric allo-sct from an hlaidentical sibling is superior to that in recipients of tandem stem-cell autografts. the aim of this multicenter retrospective national study was to identify prognostic factors for outcome of high-risk patients with mm after allo-sct prepared by ric. data from patients (median age years, range - ), who received grafts from a sibling (n= ) or unrelated donor (n= ) were analyzed. at time of transplant, only patients ( %) received ric allo-sct in cr or vgpr, while patients ( %) were transplanted in pr. patients were transplanted either in stable disease (n= ) or were in refractory/progressive disease (n= ). all patients have received at least one autologous transplant prior to ric allo-sct. the graft source was pbscs in the majority of patients (n= ). % of the patients received the seattle fludarabine and low dose tbi ric regimen, while % of patients received fludarabine, busulfan and atg. patients ( %) died of transplant-related complications. the incidences of grade - acute gvhd and extensive chronic gvhd were % and % respectively. at years, overall and progression free survivals (os, pfs) were % ( %ci, - ) and % ( %ci, - ) respectively. disease status (cr, pr, sd vs. progressive) was significantly associated with overall survival (p= . ; fig. below) . in multivariate analysis, disease status at time of ric allo-sct, was the strongest parameter associated with an improved os and pfs (p= . and p= . respectively). despite its obvious caveats, the relatively low trm observed in this series, suggest that there is still space to investigate ric allo-sct for mm. however, ric allo-sct appears to result in a durable response only if it is applied early in the disease history, especially when patients are still chemosensitive. since the latter results are also expected to be further improved with the systematic and early use of maintenance therapies (bortezomib and/or lenalidomide) after ric allo-sct, randomized or quasirandomized prospective studies are still warranted. the role of reduced-intensity conditioning (ric) allogeneic stem cell transplantation (allo-sct) for adult patients with acute lymphoblastic leukaemia (all) is still under debate. all encompasses a group of chemosensitive diseases, raising concerns that significant reduction of the intensity of the preparative regimen may have a negative impact on leukemic control. in this multicenter retrospective study, the outcome of adult (age at transplant > y.) patients with all who underwent transplantation in complete remission (cr) with an hla-identical sibling donor, were analyzed according to types of conditioning: ric in patients, and standard myeloablative conditioning (mac) (or high-dose) in patients. both groups were comparable in terms of gender, cr status (cr and cr ), interval from diagnosis to allo-sct, and r/d cmv serostatus. patients in the ric group were older (median y. vs. y in the mac group;p< . ). most of the patients in the mac group received high dose tbi ( %), while the majority of the ric regimens included either low-dose tbi or were atg+chemotherapy-based regimens. the majority of patients ( %) from the ric group received pbscs. in the mac group, the stem cell source consisted of bone marrow in % of patients. with a median follow-up of months (range, , the incidences of grade ii-iv and grade iii-iv acute gvhd were: %, %, and %, % in the mac and ric groups respectively (p=ns). the cumulative incidence of nonrelapse mortality at years (nrm) was % (mac) vs. % (ric) (p= . ). the cumulative incidence of relapse at years was % (mac) vs. % (ric) (p= . ). however, the latter differences did not translate into any significant difference in term of leukemia-free survival (lfs) at years: % (mac) vs. % (ric) (p= . ). in multivariate analysis for lfs, the status at transplant was the only factor associated with an improved lfs (p< . , rr= . , %ci, . - . ). the results of this study suggest that ric regimens may reduce nrm rate after allo-sct for adult all when compared to standard mac regimens, but with a higher risk of disease relapse and no impact on lfs. the latter represent promising findings, since patients who received ric are likely to have serious comorbidities, which led the transplantation center to choose ric, and surely most of these patients would not have received a standard allo-sct in most institutions. therefore, ric allo-sct for adult all (> y.) may represent a valid therapeutic option when a conventional standard conditioning is not possible, warranting further prospective investigations. reduced-intensity conditioning allogeneic stem cell transplantation for patients with acute myeloid leukaemia: long-term results of a "donor" versus "no donor" comparison m. mohty *, h. de lavallade, j. el-cheikh, p. ladaique, c. faucher, s. fürst, n. vey, d. coso, a.m. stoppa, j.a. gastaut, c. chabannon, d. blaise institut paoli-calmettes (marseille, fr) the issue of possible higher relapse rates after reducedintensity conditioning (ric) allogeneic stem cell transplantation (allo-sct) is still under considerable debate. this report describes the updated long term results (initial publication, leukemia ) of consecutive acute myeloid leukaemia (aml) patients, diagnosed between and in a single centre. using a genetic randomization through a "donor" versus "no donor" comparison, our aim was to assess the true benefit of ric-allo-sct for adult aml. in this series, patients ( %; "donor" group) had an "identified" hlaidentical sibling donor, while the remaining patients had no hla-matched related donor ("no donor" group). as per institutional policy, hla-mud were not considered during the study period. no significant differences in patients or aml features were found between the two groups. in the "donor" group, patients ( %; median age, (range, - )) could actually proceed to the ric-allo-sct. the current median follow-up is months. in an "intention-to-treat" analysis, the km estimate of leukemia-free survival (lfs) was significantly higher in the "donor" group as compared to the "no donor" group (p= . ; % versus % at years). when restricting the analysis to patients who could actually receive the ric-allo-sct (median follow-up, months from time of allo-sct), the difference in lfs was also significant between this group of patients ("transplant" group) and the remaining patients ("no transplant" group; p= . ; % versus % at years). no major toxicities were encountered during ric administration (fludarabine, busulfan and atg), and only patients died from toxicity, for a cumulative incidence of trm of % ( %ci, - %) at last follow-up. this relatively low trm translated towards a significantly higher overall survival (os) in the "transplant" group as compared to the "no transplant" group (p= . ). in the "intention-to-treat" analysis, os was still significantly higher in the "donor" group as compared to the "no donor" group (p= . ; fig. below) . after controlling for all relevant factors, in the multivariate analysis, only actual performance of ric-allo-sct (p= . ; rr= . ; %ci, . - . ), was significantly predictive of an improved lfs. based on these long term results, we conclude that if a matched related donor is identified, ric-allo-sct should be proposed since it represents a valid and potentially curative option for aml patients not eligible for standard myeloablative allo-sct. graft failure after reduced-intensity conditioning b. hertenstein*, e. dammann, r. brand, a. van biezen, d. niederwieser, t. de witte, t reduced intensity conditioning (ric) regimens use doses of chemotherapy and/or radiotherapy which per se do not eradicate the malignant cells but which should be nevertheless sufficient to allow sustained engraftment. the incidence of graft failure might therefore be higher in ric and may be dependent on the regimen used. to evaluate the incidence of graft failure and the potential risk factors in patients transplanted after ric, data files of such transplantations were selected from the clwp data base. in all records chemotherapeutic drugs and dosages as well as tbi dosages were individually checked and only records meeting the ebmt definition of ric were included. the analysis was further restricted to patients surviving > days. ric transplants were identified. a chemotherapeutic ric regimen was used in patients (flu/bu , flu/mel , flu/cy ), tbi only in and a combination of chemotherapy and tbi in patients. stable engraftment occurred in patients ( . %), no engraftment in patients ( . %) and a graft loss in patients ( . %). survival for patients with graft failure was % at days and % at one year. patients received a second transplant and survival at one year was % for these patients. graft failure was lowest in patients with myeloma ( . %) and highest in patients with mds/mps ( . %). with the use of bone marrow as graft source graft failure was much higher than with peripheral blood stem cells ( . % vs. . %) . graft failure occurred in . % of transplants from hla-identical siblings, in . % from matched unrelated donors and in . % from mismatched unrelated donors. there was no difference between the different conditioning regimens, neither between chemotherapy alone, tbi or combined regimens ( . % vs. . % vs. . %) nor between the major chemotherapy regimens themselves (flu/bu . %, flu/mel . %, flu/cy . % for hla-id siblings). abo match and donor-recipient sex mismatch had no impact on graft failure. in multivariate analysis underlying disease and graft source were significantly correlated with graft failure. in summary the incidence of graft failure after ric is low. mds/mps as underlying disease and the use of bone marrow as graft but not the type of the conditioning regimen were risk factors for graft failure. ( ), h. esperou ( ) , m. attal ( ) , n. milpied ( ) , b. lioure ( ) , p. bordigoni ( ) , i. yackoub-agha ( ) , j. bourhis ( ) , b. rio ( ) , e. deconninck ( ) , m. renaud ( ) , n. raus ( ) , d. blaise ( ) ( )hôpital edouard herriot (lyon, fr); ( )société française de greffe de moelle et de thérapie cellulaire (saint-denis, fr) this report updates a retrospective study from sfgm-tc registry concerning patients who underwent allogeneic hematopoeitic stem cell transplantation (hsct) after reduced intensity conditioning (ric) from hla identical siblings ( %) and unrelated donors ( %) for hematological malignancies. at time of conditioning, patients were in cr, in pr, in stable disease (sd) and in progressive disease (pd). as conditioning, patients received fludarabine and tbi ( grays), patients fludarabine, busulfan and atg and patients other regimens. after transplant, patients ( %) developed an acute gvhd grade ii (grade ii: , iii: and iv: ). a chronic gvhd was present in patients ( %) ( limited and extensive). with a median followup of months, the and -year probability of overall survival (os) were . % ( - ) and %( - ) respectively and the and -year probability of event-free survival (efs) were %( - ) and % ( . - ) respectively. the trm at year, years and years was % ( - ), % ( . - ) and % ( - ). a mixture model, gfcure with splus statistical package determined the percentages of long-term survivors and its adequacy was verified graphically. the probability to be a long-survivor was % ( . - . ) (fig. ) and to be a long event-free survivor was % ( - ) (fig. ) . the multivariate analysis has tested recipient and donor age, disease status pre-transplant, number of transplants before rict, hsc source, sex matching, hla matching, cmv status and abo compatibility. the only factor which had a significant impact on long-term survival after rict was the disease status just prior conditioning: pr versus cr: hr: . [ . - . ] p< . and pd versus cr: hr: . [ . - . ] p< . . in conclusion, these updated data demonstrate that allogeneic hsct after ric was able to possibly cure % of patients with haematological malignancies and the most important factor to take into account remains to be in cr pre-transplant. the haematopoietic cell transplantation-specific comorbidity index predicts survival and non-relapse mortality in lymphoma and myeloma patients receiving a ric allograft l. farina* ( ), b. bruno ( ) the allogeneic hematopoietic cell transplantation-specific comorbidity index (hct-ci) has been recently developed to identify patients at high risk of morbidity and mortality after an allogeneic stem cell transplant (allosct). reduced-intensity conditioning (ric) regimens have decreased non-relapse mortality (nrm) in heavily pre-treated patients. we performed a retrospective study to assess whether comorbidities, according to hct-ci, may influence the outcome of lymphoma and multiple myeloma patients undergoing a ric allosct. between and , patients received a ric allosct from a hla identical sibling (n= ) or unrelated (n= ) donor in three italian transplant units. median age at transplant was years (range, - ) and % of the patients were or older. diseases included non hodgkin's lymphoma (n= ), multiple myeloma (n= ) and hodgkin's lymphoma (n= ). median number of previous treatments was (range, - ). disease risk according to kahl et al. (blood ) was low, intermediate and high in %, % and % of patients respectively. patients with hct-ci of , - and ≥ were ( %), ( %) and ( %), respectively. variables included in multivariate analysis were age (< or ≥ ), disease risk (low, intermediate and high), number of previous lines of therapy (≤ and > ), hct-ci ( , - and ≥ ), karnofsky perfomance status (ps, > % and ≤ %). oneyear os and pfs were %, %, % and %, %, % in patients with hct-ci of , - and ≥ respectively. cumulative incidence of nrm was %, %, % at year and %, % and % at years, whereas relapse mortality was %, %, % at year and %, % and % at years. by multivariate analysis only karnofsky ps (p= . ) and hct-ci (p= . ) were correlated with os, whereas pfs was influenced by disease risk category (p= . ), number of previous therapies (p= . ), and both karnofsky ps (p= . ) and hct-ci (p= . ). interestingly hct-ci was the only significant factor that could predict nrm (p= . ) while karnofsky ps failed to show a significant correlation (p= . ). of note, age (≥ ) was not statistically significant either in os, or pfs or nrm. although the data need to be confirmed in prospective trials, these results showed that hct-ci may be a useful tool to predict os, nrm and also pfs after ric allosct in lymphoma and myeloma patients. in the clinical setting hct-ci and not age should be used for patient risk assessment. venous thromboembolism (te) occurs as a consequence of genetic and environmental factors. important genetic risk factors are deficiencies of natural anticoagulants, antithrombin-iii (at-iii), protein c (pc) and ps, and genetic mutations of factor v leiden (fvl) and prothrombin (pth a ). thrombotic complications after hct are usually catheter-related thrombosis (crt), pulmonary te (pte) and deep vein thrombosis (dvt). our aim in this study was to evaluate the effects of the deficiencies of atiii, pc and ps, and the gene mutations of fvl and pth on the incidence of the development of te complications and liver sinusoidal obstruction syndrome (sos) at the early or late period post-hct. in our center, pre-transplant work-up includes routine thrombophilia tests. between apr -jan patients (m/f: / , median age: years) admitted to our transplantation center were retrospectively analyzed for the relation of the presence of thrombophilia and the frequencies of the occurrence of a te complications and liver sos. all but patients (n= ) had an hla identical sibling donor. the ratios of the detection of the plasma activation level below % for pc, ps and at-iii were . % ( / ), . % ( / ) and . % ( / ), respectively. gene mutations were studied in patients prior to transplant and the frequencies of gene mutations were detected in . % patients (n= ) either fvl (n= ) or pth (n= ) gene. none of the patients had both mutations, fvl and pth. at the peri-or posttransplantation period we observed venous te in patients ( crt, pe and dvt), liver sos in patients and myocard infarction at the early period in only one patient. in out of patients with low pc activity crt occurred, and liver sos was observed in only one patient. in out of patients with genetic mutation developed a te complication, crt and pte. crt occurred in of patients with fvl mutation, while crt and pte among patients with pth mutation were observed. liver sos was seen in only one patient with pth heterozygote positive. we found the presence of low pc level or genetic mutation increased the frequency of the development of te (or: . and . , respectively), but no effect on the liver sos. in conclusion, the use of thromboprophylaxis at peri-transplant period in patients with genetic mutations is still a controversial topic; which should be elucidated with controlled studies. iron overload (io) is an adverse prognostic factor in patients who undergo allogeneic haematopoietic stem cell transplantation (hsct) for thalassemia and appears to play a similar role in patients with other hematological disorders. estimation of io is primarily based on serum ferritin, however many confounding factors particularly in hsct recipients may result in frequent ferritin overestimation. aim of the study was to quantify io by uperconducting quantum interference device (squid) after hsct and evaluate the impact on infections and gvhd; additionally the feasibility of iron-depletion has been investigated in this pilot study we evaluated io in consecutive adult patients who received hsct from a matched sibling (n= ) or a matched unrelated donor (n= ). primary diagnosis included aml/mds in % of cases. assessment of io after hsct included serum ferritin and in those with hyperferritinemia (> ng/ml), liver iron concentration (lic) was evaluated by squid magnetic susceptometry. io assessment was performed in patients in remission at a median time of days after hsct (range - days). median serum ferritin was ng/ml (range - ).thirtynine of ( %) patients had serum ferritin level > ng/ml; lic (microgfe/g liver wet-weight) evaluated by squid was available for / patients with elevated ferritin values. overall, patients ( %) had moderate (lic levels - microg/gww) to severe (lic levels> microg/gww) io; median lic values were microg/gww (range - ). nine patients ( %) had normal lic values (lic< ) despite high ferritin. patients with lic> received a median of packed red blood cells before study evaluation (range - ). the rates of bacteremias, invasive fungal disease and chronic gvhd were higher among patients with lic> as compared to patients with lic< ( %, % and % vs %, % and % respectively; p=ns). eighteen of the patients with lic levels > were treated by regular phlebotomy. for of these the phlebotomy program is still ongoing, while patients completed the program with ferritin normalization; one patient was switched to deferasirox due to poor tolerance. in cases phlebotomy was considered contraindicated; patients who relapsed did not receive any treatment. our preliminary data show that one third of hsct recipients may present moderate/severe io as assessed by squid, and iron-depletion resulted feasible in % of these subjects. a trend for higher rates of infectious complications and chronic gvhd was observed in patients with io. despite recent advances, mortality rates after allogeneic hematopoietic stem cell transplantation (hsct) remains high and cannot be accurately predicted. several reports from the seattle group suggested the use of comorbidity indexes (ci) to provide valid and reliable scoring of pre-transplant comorbidities that predicted non-relapse mortality (nrm) and overall survival [blood (cci) , (hct-ci) & ann int med (pam)). however, whether such indexes could be used by other groups, and if one index better predicts survival than another, is yet unknown. hct-ci and pam required grading according to pulmonary function tests (pft), which were lacking for a majority of our patients. we thus designed a modified hct-ci and a corrected pam, without pft. survival curves were estimated using kaplan-meier method. cumulative incidences (ci) of non-relapse mortality were analyzed, with relapse treated as competing event. the likelihood ratio statistics in proportional hazards models was computed for each index, with its associated p-value, as a measure of association between the comorbidity indexes collapsed into risk groups and the outcomes. discriminative performance of comorbidity indexes was then evaluated by the c-index. standard errors of c-indexes were computed from that of somers' dxy rank correlation coefficient. p-values for comparison of c-indexes were obtained using a nonparametric bootstrap procedure. we thus retrospectively studied patients ( male, female) who received their first allogeneic hsct at saint louis hospital between april and december . the median age was years ( - ), patients ( %) were transplanted from a matched related donor and patients received an hsct for intermediate or high-risk diseases ( %). using cci, % of our patients had indexes of or more; median reduced hct-ci was ; and median corrected pam score was . the discriminative properties of the ci were rather low in our patient population with c-index of . , . and . for the cci, reduced hct-ci and corrected pam indexes, respectively. comparison of patients-and transplant-characteristics between our and seattle group's cohorts however revealed significant differences with more children, more cord blood hsct and hsct for fanconi anemia in saint louis. finally, direct comparison of scoring items and multivariate analysis revealed that age, unmatched related donor and hepatic disease were associated with nrm in our cohort. primary graft failure occurs in - % of unrelated cord blood transplantation (ucbt) patients. its occurrence is associated with increased risk of death due to relapse (when associated with autologous recovery) or complications of pancytopenia. salvage transplant is possible in patients with absence of neutrophil recovery (anr): clinical tools are required to promptly identify these patients at risk. we conducted this retrospective study to identify risk factors for anr. from to , consecutive children underwent a first ucbt in our center. anr was defined as absolute neutrophil count ≤ /microl in patients alive without disease on day + . patients for whom a salvage transplant conditioning regimen was initiated before day + were excluded from this analysis (n= , of them are late survivors). ninety one ( ) children were eligible for analysis. median time for neutrophil recovery (≥ /microl) was days (range, to ). anr occurred in / ( . %) and was confirmed by bone marrow biopsy. of these patients, underwent a nd transplant, one of which succeeded. five ( %) died between day + and + of transplant complications that occurred at a median of days (range - ) of their first transplant. in of them, complications were related to persistent neutropenia and/or lymphopenia. our current strategy of waiting until day + for a nd transplant decision carries a significant risk of mortality in patients with anr. when analyzed for demographics and graft characteristics, the group with anr did not differ significantly from the group with neutrophil recovery. the specificity and sensitivity for anr prediction of white blood cell (wbc) < /microl at different time points is as shown in table . wbc < /microl on day + and day + carries a risk of day + anr of % and %, respectively. the negative predictive value of wbc < /microl on day + is %. therefore, in order to reduce the risk of mortality associated with anr, we now proceed to nd transplant evaluation for children with wbc < /microl by day + , and initiate the therapy on day + if wbc counts remains < /microl. purpose: the aim of this retrospective study was to determine whether pre-transplant iron overload assessed by serum ferritin level predict the non-relapse mortality (nrm) and overall survival (os) rate after haematopoietic cell transplantation (hsct). patients and methods: patients with hematological disorders underwent myeloablative or reduced-intensity conditioning followed by the first hsct from related or unrelated donors at keio bmt program from to . patients were excluded from this analysis because ferritin and crp levels were not available or measured at the time of transplant. demographic data and information on comorbidities was obtained from the keio bmt database. comorbidity was scored according to the hct-specific comorbidity index (hct-ci) proposed by the seattle group without any modifications. results: median value of ferritin was ng/ml(range - ng/ml). the percentage of patients with ferritin ng/ml did not vary significantly among diseases. in all but patients crp were within normal range at the time of measuring ferritin. there was a strong relationship between pretransplant ferritin and os. the -year os for patients was . % in the first quartile (ferritin - ng/ml) (q ), . % in q ( - ng/ml), . % in q ( - ng/ml), and . % in q (more than ng/ml) (p= . ). -year os was also significantly higher for those with ferritin level more than ng/ml compared with those with ferritin equal or less than ng/ml ( . % vs . % p= . ). the -year nrm was also significantly associated with pretransplant serum ferritin level. the -year nrm was . % for patients with ferritin more than ng/ml, and . % for those with ferritin equal or less than ng/ml (p= . ). the mean score of hct-ci was . for those in q , . in q , . in q , and . in q . the mean score of hct-ci was significantly higher in patients with hyperferritinmia (ferritin more than ng/ml) than those without ( . vs . p= . ). conclusion: these results suggested that pretransplant level of ferritin could be a marker of predicting nrm, os after hct and thus useful for patient counseling before hct. whether the addition of ferritin level into hct-ci scoring system give a better prediction of posttransplant nrm and os needs further investigation. increased incidence of renal impairment after radioimmunotherapy as part of the conditioning regimen before allogeneic stem cell transplantation t. zenz* ( ), r. schlenk ( ) intensifying the conditioning regimen with radioimmunotherapy (rit) is feasable for high risk leukemias and mds patients. while additional radiation exposure by rit is greatest in the bone marrow, significant exposure of the kidney occurs and an increased frequency of bmt nephropathy has been observed in patients receiving rit using the rhenium labelled anti-cd antibody. in order to obtain a precise picture of the incidence of renal impairment we compared two large cohorts receiving allogeneic stem cell transplantation (sct) with or without the use of rit from our center. between and , patients with a median follow-up of months received hla-identical or mismatched allogeneic sct. of these received an intensified conditioning regimen with a rhenium (n= ) or yttrium labelled (n= ) anti-cd antibody. the cohorts were similar with respect to sex, stem cell source and use of nephrotoxic medication. there was an increased proportion of patients with aml, all, and haploidentical donors in the group receiving rit (p< . ), while the conventional group had more patients with cml, tbi in the conditioning regimen, cyclosporine use and incidence of gvhd (p< . ). the clinical characteristics were documented for years post sct. we defined relevant renal insufficiency as a creatinine level above µmol/l and the failure to reach creatinine values below to exclude intermittent drug toxicities. the incidence of renal failure was % in patients with conventional conditioning regimens and % in patients receiving rit (p= . ). the competing risk analysis showed a significant increase in the cumulative incidence of nephropathy (p= . ). even though the dosimetry showed decreased renal doses in patients with y this did not lead to a decreased incidence of renal impairment. other factors as tbi, cyclosporine use, donor source or gvhd did not influence the development of renal failure. in a multivariate model including rit, csa, donor source, gvhd and the combination of csa and rit, the combination of rit and csa was the only significant risk factor (p= . ). in conclusion, rit lead to an increased frequency of continuous renal impairment. the combination of cyclosporine with rit appeared detrimental while tbi (with renal shielding) did not lead to an increased incidence of renal failure. strategies to decrease the incidence of renal impairment may include nephroprotection with ace inhibitors or alternative immunosuppression. prospective evaluation of oral mucositis in allogeneic stem cell transplant recipients receiving conventional myeloablative conditioning or reduced-toxicity conditioning with treosulfan and fludarabine h. uotinen*, l. volin, e. juvonen, a. nihtinen, t. ruutu helsinki university central hospital (helsinki, fi) oral mucositis (om) is a frequent and often severe complication of allogeneic stem cell transplantation (sct). the aim of this study was to evaluate the incidence and duration of severe (who grade iii-iv) and ulcerative (grade ii-iv) om in patients receiving different conditioning regimens. between sept and nov , patients were prospectively observed daily from the start of conditioning until hospital discharge. conventional (convent.) myeloablative (ma) conditioning (cy/tbi n= , bucy n= ) was given to patients ( aml, all, mds, cml, mm, mf, others) and treosulfan (treo)-based conditioning to ( mds, aml, mm, others) patients. treo-based conditioning consisted of treo x g/m² (n= ) or x g/m² (n= ) and fludararabine (fld) mg/m². treo-fld conditioning is regarded as ma at least at the treo dose level of x g/m² but with reduced non-hematological toxicity. treobased conditioning was given to patients not eligible for convent. full intensity conditioning because of age, comorbidity or heavy preceding cytotoxic treatment. there were no significant differences in the om results with the two treo dose levels used. the median age of the patients was ( - ) years in the convent. ma group and ( - ) years in the treo group. in the convent. ma group the donor was sibling in cases and unrelated in cases. in the treo group the donor was sibling in cases and unrelated in cases. the incidence of severe om was % in the convent. ma group and % in the treo group (p< . ). severe om episodes had a mean duration of ( - ) days. the incidence of ulcerative om was % in the convent. ma group and % in the treo group (p< . ). the ulcerative om episodes had a median duration of ( - ) and (range - ) days (p= . ), respectively. om reached maximum who grade on day in the convent. ma group and on day in the treo group after the start of conditioning. the median duration of hospitalisation from the sct was ( - ) days in patients with ulcerative om vs. (range - ) days in patients without (p= . ). in conclusion, conditioning with treosulfan and fludarabine caused significantly less severe and ulcerative mucositis than conventional ma regimens, and the duration of ulcerative mucositis was shorter. this supports the role of treo-fld conditioning for allogeneic transplantation especially in the treatment of patients with increased risk of toxic complications. patients receiving cbt had significantly slow neutrophil and platelet recovery in multivariate analysis. the incidences of acute and chronic gvhd were not significantly different. unrelated bmt showed better trm ( % versus % at year, p< . ), relapse ( % versus % at years, p< . ) and dfs ( % versus % at years, p< . ) results compared with cbt. next, we analyzed bmt (n= ) and cbt (n= ) recipients who have taken total body irradiation (tbi; > gy) containing myeloablative regimen and calcineurin inhibitors (cyclosporine or tacrolimus) plus methotraxate (mtx) for gvhd prophylaxis without history of prior transplants. in this subpopulation, multivariate analysis revealed no significant difference between bmt and cbt in dfs ( % and % at years; hazard ratio (hr): . ; % confidence interval (ci): . - . ; p= . ). no statistically difference was also seen in trm ( % at year after bmt and % at year after cbt; hr: . ; %ci: . - . ; p= . ). however, the cumulative incidence of relapse was significantly lower in bmt than in cbt ( % and % at years; hr: . ; %ci: . - . ; p= . ). the current japanese registration data in mds showed overall results of unrelated bmt were better than those of unrelated cbt by competing risk regression models. these data also suggest that unrelated cbt could be safely and effectively used as same as unrelated bmt when adequate transplant procedures are selected. comparative studies have shown that cb transplant is characterized by a lower risk of gvhd. cb units have been reported to contain tregs, but minimal data are available on these cells. aim of this study was to compare the suppressive functions of tregs expanded from cb units with those expanded from the peripheral blood (pb) of patients who have undergone an allogeneic sct. tregs were purified from mononuclear cells obtained from cb units or pb using the cd +cd + regulatory t-cell isolation kit (miltenyi biotec) and expanded for days in -well u-bottom plates coated with anti-cd and anti-cd moabs plus il- . to assess the suppressive functions, expanded tregs from cb units or pb were seeded with naïve autologous effector t cells stimulated with allogeneic dendritic cells (dc) pulsed with apoptotic leukemic blasts, then incubated with [ h]-thymidine and counted in a beta-counter. suppressor activity was measured as [ h]-thymidine incorporation in the presence or absence of tregs. the proportion and the immunophenotypic analysis of tregs present in the cb units (n = ) -in terms of expression of surface cd , cd , cd l, cytoplasmic ctla- and foxp was comparable to those obtained from the pb of allografted patients (n = ). in addition, tregs from cb units and from the pb of these patients showed an equivalent expansion capacity [mean fold increase (range), cb units . ( . - ); pb patients . ( . - . ) ]. on the contrary, preliminary data show that tregs expanded from cb units (n = ) exert a higher suppressive function on the proliferative reaction of t cells stimulated by allogeneic dc compared to tregs expanded from the pb of allografted patients (n = ) [mean fold reduction (range), cb units . ( . - . ); pb patients . ( . - . ) ]. moreover, immunofluorescence analysis demonstrated that tregs expanded from cb units (n= ) are highly positive for cytoplasmic il- (mean %, range - ) and negative for ifn-gamma. these results indicate that tregs contained in cb units exert a potent suppressive function in mixed lymphocyte reaction culture assays and offer further insights into the understanding of the biology of cb transplant. double negative tregs are reduced in allo-transplanted patients developing graft-versus-host disease b. serio* ( ), z. mciver ( ), a. risitano ( ) , c. selleri ( ) , j. maciejeski ( ) ( )cleveland clinic (cleveland, us); ( ) federico ii university of naples (naples, it) during development of a healthy immune system, central tolerance is induced in the thymus by the negative selection of t lymphocytes that have a high affinity for self antigens. after bmt, central tolerance may be impaired by thymic involution and conditioning regimens resulting in dysregulated alloreactivity. various subpopulations of regulatory t cells including the tregs or suppressor t cells with cd +cd +foxp + phenotype (foxp tregs), nk t-cells and the cd +cd -cd -cd -a/atcr+ t regs (double negative [dn] tregs) had led to the mechanisms of peripheral tolerance. we investigated behavior of dn following human allogeneic hsct with regard to the occurrence of graft versus host disease (gvhd) and restoration of t cell receptor (tcr) repertoire. a cohort of patients was investigated; underwent matched unrelated hsct and received matched sibling grafts. frequency of dn and tcr repertoire of cd and cd cells was measured serially and at the time of diagnosis of gvhd by flow cytometry. our patient population demonstrated skewing of tcr repertoire and we identified a very strong and linear relationship between total number of vâ family expansions and the grade of gvhd (grade - , n= , p=. ); this relationship held true when considering separately both the cd and cd compartments (p=. and p=. , respectively). the median frequency of dn tregs was calculated to be . % of the total cd + t-cell population (range . - . ). by using the total number of vâ family expansions as a gauge of alloreactivity, we observed a reduced number of dn tregs in those individuals that developed or more vâ family expansions (n= , mean . vs . cell/ìl, p=. ). in addition we also noted a significant difference in both the mean percentage and mean absolute values of dn tregs for those individuals that developed gvhd (grade > ) when compared to those that did not (n= , . vs . %, p=. , and . vs . cells/ml, p=. respectively). the size of dn treg compartment inversely correlated to the grade of gvhd (grade - ) as both percentage and absolute value (n= , p<. , and p=. respectively). in conclusion we found that vb family expansion are associated with degree of alloreactvity and gvhd. in addition we show that dn tregs are reduced after bmt suggesting their important role in peripheral tolerance and alloreactivity. this work contributes to better define the regulatory role of dn and to develop future therapeutic applications. patients with myelofibrosis. a study of the mds subcommittee of the chronic leukaemia working party of the de witte on behalf of the mds subcommittee of the chronic leukemia working party of the european group for blood and marrow transplantation hla disparities were / match (n= ), / (n= ), and / (n= ). , respectively. twenty-one patients received cyclosporine, and patients had tacrolimus alone as gvhd prophylaxis from day c . neutrophil engraftment was achieved in % (median day; ). cumulative incidence of pir, grade - and grade - acute gvhd were , and %, respectively. estimated -years survival was % ( % ci: - %) sierra hospital de la santa creu i sant pau (barcelona, es) background: neurological complications (nc) of allogeneic hematopoietic stem cell transplantation (allo-hsct), involving central (cns) and/or peripheral nervous system (pns) are common and remain life-threatening in most cases. their incidence and characteristics after allo-hsct have not been well defined we excluded patients with nc due to cns relapse of their malignancies as well as encephalopaty in the context of pre-mortem multi-organ failure cns complications included seizures in cases, non-focal encephalopaties, meningoencephalitis and strokes or hemorrhages. pns complications consisted of cases of neuralgia/neuritis and cases of demyelinating axonal neuropathies ( guillain-barré syndrome) piñana is supported by grants isciii (cm / ) o haematopoietic stem cell transplantation for advanced primary mds in children: results of the ewog mds study group b. strahm* ( ) ); ( )wilhelmina children´s hospital and methotrexate, and additional methylprednisolone in case of a sibling donor. the median clinical follow-up time for the patients was months ( - months). several anti/coagulation activities associated with endothelial cell activation were serially assessed up to months: prothrombin fragments + (f + ), thrombin time (tt), fviii:c, activated protein c-protein c inhibitor (apc-pci) complex and protein c act (pc). during conditioning (on day- ) as an early sign of thrombin generation, f + and apc-pci complex increased - fold. after engraftment, fviii:c increased steadily to reach its high maxi-mum on day + ( % ± %, median±sd, p< . ). interestingly, pc rose ( % ± %) in parallel with fviii:c with a -fold individual variability. after the engraftment fviii:c and pc were highly interrelated. gvhd developed in patients and was predicted by early low pc activity (< %) during conditioning (p= . ) (or= . ). gvhd was also associated with elevated level of f + (> . nmol/l) (p= . ) and was predicted by short tt (< s) (p= . ) (or= . ) after the transplantation (on day + ). no patient whose f + was < . nmol/l (n= ) developed gvhd. elevated level of f + (> . nmol/l) after the transplantation associated with non-relapse mortality. patients with the highest thrombin generation after sct all died ( - months). in con-clusion, early up-regulation of thrombin generation and down-regulation of pc associated with the appearance of gvhd. after allogeneic sct procedure there is an intimate relation between endothelium regulated coagulation and development of gvhd, suggestive of a new therapeutic target. final results from a phetema study l. rosiñol* ( ) , j.j. lahuerta ( ) , a. sureda ( ), j. de la rubia ( ), j. , m. hernández-garcía ( ), b. hernández-ruíz ( ), j.a. , j.l. bello ( ), d. carrera ( ), m.j. peñarrubia ( ) , e. abella ( ) , a. león ( ), c. poderós ( ) , j.c. j. besalduch ( ) , r. , i. p. ribas ( ), j. san miguel ( ) , j. bladé ( ) ( )hospital clinic (barcelona, es) ; ( ) background: two randomized trials showed that tandem transplant result in a significantly longer efs and os in patients failing to achieve complete remission (cr) or near-cr with a single transplant. however, other studies failed to show survival benefit from a second transplant but there was no survival plateau. promising results have been reported using dose-reduced intensity conditioning (allo-ric), especially after debulky with an autologous transplant.aim: to investigate the efficacy in terms of response upgrading and survival from a second transplant intensification in patients with chemosensitive disease who failed to achieve cr or near-cr with a first transplant. patients and methods: patients diagnosed with mm from oct to dec younger than years received courses of vbmcp/vbad and responding patients were intensified with busulphan/melphalan or mel- followed by stem cell support. patients not achieving cr or near-cr were planned to undergo a second transplant (second auto with cvbcyclophosphamide, etoposide and bcnu -or mel- intensification or an allo-ric with fludarabine/mel- conditioning, if sibling donor available. results: eighty-five patients received a second autologous transplant while underwent an allo-ric. the cr rate was significantly higher with allo-ric ( % vs. %, p= . ). there was a trend towards a higher trm with the allogeneic procedure ( % vs. %, p= . ). after a median follow-up of months for alive patients, there were no significant differences in efs ( mos. vs. nt reached) and os ( mos vs. not reached for autologous tandem vs. auto/allo-ric). however, there is a plateau in the "allo-ric" group beyond years of the second procedure not observed in the autologous arm. a final update will be presented. conclusions: ) an allo-ric transplant after an autologous procedure results in a significantly higher cr rate than a second autologous transplant, and ) although we found no significant differences in survival between the two transplant modalities, there is a plateau in the allogeneic group. ( ) background: it has been assumed that patients with primary refractory myeloma benefit from early high-dose therapy/stem cell support (hdt. however, in the reported series patients with "unresponsive-progressive disease" vs those "nonresponding-non progressing" were not analyzed separately aim: response and survival after early hdt in the two populations of truly primary refractory multiple myeloma (i.e., patients with progressive disease versus those with "no change" or "stable disease" while receiving the initial therapy). patients and methods: from oct to dec , patients with mm received cycles of vbmcp/vbad and at least one transplant. of the patients were refractory to vbmcp/vbad. these resistant patients were scheduled to receive a tandem transplant, the first with bu- /mel- or mel- and the second "autologous" with cvb (ciclophosphamide(etoposide/bcnu) or mel- or "allo-ric" (if donor available) with fluda/mel- . response and progression were defined by the ebmt criteria. results: of the primary refractory patients had progressive disease under the initial chemotherapy while allogeneic stem cell transplantation (sct) is the only curative treatment approach in patients with myelofibrosis. the major limitation is the high treatment related mortality, which exclude mainly elderly patient from this treatment procedure. to determine the toxicity and efficacy of a dose-reduced conditioning regimen, consisting of busulfan ( mg/kg), fludarabine ( mg/m²) and anti-thymocyte globulin (atg fresenius: x mg/kg for related and x mg/kg for unrelated sct) followed by allogeneic sct, we performed a prospective multicenter trial in elderly patients with myelofibrosis in centers in three countries. from to , patients with a median age of years (r., - ) were included and were evaluable for outcome. risk profile was low risk with constitutional symptoms ( %), intermediate risk (n= %) and high risk (n= %). all but patients received peripheral blood stem cells as graft source either from related (n= ) or unrelated donor (n= ). all but one ( %) patient showed leukocyte and platelet engraftment after a median of and days, respectively. the median duration of leukocyte aplasia was days (r., - ). acute graft-versus host disease (gvhd) grade ii o iv occurred in % and severe agvhd iii/iv in %, while chronic gvhd was seen in % of the patients. non-relapse mortality at year was % ( % ci: - %) and significantly lower for patients younger than years of age ( % vs %, p< . ) and for patients with low risk vs intermediate/high risk disease ( % vs %, p= . ), while a higher nrm was seen for patients transplanted from hla mismatched donors ( vs %, p= . ). the cumulative incidence of relapse at year was % ( %ci: - %) and influenced by time from diagnosis to sct of less or more than months ( vs %, p= . ). patients with splenectomy had higher incidence of relapse ( vs %, p= . ). the estimated year overall and event-free survival was and %, respectively. the overall survival was influenced by age less than years ( % vs %, p= . and low vs intermediate/high risk ( % vs %, p= . and hla mismatch ( vs %, p= . ), while no impact on survival was seen for cytogenetic abnormalities, jak mutation status and donor (related vs unrelated) these results of a prospective multicenter study show excellent outcome of a busulafan/fludarabine based reduced conditioning regimen followed by allogeneic stem cell transplantation in patients with myelofibrosis. graft rejection after stem cell transplantation following reduced-intensity conditioning is influenced by the underlying disease, the donor type, disease stage and the cd content of the graft g.-n. franke*, a. mikolajewska, s. leiblein, h.-k. al-ali, e. hennig, w. pönisch, d. niederwieser, t. lange university of leipzig (leipzig, de) objectives: stem cell transplantation (sct) after reduced intensity conditioning (ric) is routinely used as a curative approach for older and medical impaired patients with haematological malignancies. in contrast to sct with conventional conditioning, graft rejection (gr) remains an important issue. we analyzed patients with sct after gy total body irradiation (tbi) with or without fludarabine (flu) conditioning to identify risk factors for gr. patients and methods: patients with a median age of (range - ) years, underwent allogeneic sct (bm= , pbsc= ) from a related (n= ) or unrelated (n= ) donor for aml (n= ), cml (n= ), nhl/mm (n= ), mds/mps (n= ) or other diseases (n= ). conditioning regimen consisted of gy tbi at day and flu mg/m from - to day - (n= ) followed by cyclosporin a and mycophenolate mofetil. results: ( . %) patients developed primary (n= ) or secondary graft failure (n= ) defined as a t-cell chimerism of < % donor cells. in univariate analysis and also in multivariate analysis, diagnosis of cml (p= . ), unrelated donor (p= . ), early disease stage prior to sct (p= . ) and low cd cells in the graft (p= . ) were identified as independent predictors for gr. the relative risk (rr) to experience gr was . , . , . and . , respectively. even in a subgroup analysis with pbsc recipients only, cml, unrelated donor and disease stage were associated with higher risk of gr. furthermore, a lower cd count increases the risk of gr by a rr . (p= . ) in the multivariate model (median . x /kg bw). an increase in cd cells was not associated with increased incidence of acute gvhd grad ii-iv until day (p= . ). in contrast, the cd -content of the graft had no impact on gr either as categorical (>median vs. <median) or as continuous variable (p= . ). conclusion: in contrast to given risk factors for graft rejection, the t-cell content of the graft can be individually requested in every patient. since higher cd cells in the graft reduces the risk of gr without subsequent acute gvhd in our cohort of patients, we recommend the administration of cd cells > . x /kg bw especially in patients with cml, unrelated donor and early disease stage in ric-sct. unrelated cord blood transplantation for adult patients with acute myeloid leukaemia/myelodysplastic syndrome using a reduced-intensity conditioning regimen consisting of fludarabine, melphalan and total body irradiation k. masuoka*, k. ishiwata, m. tsuji, s. takagi, h. yamamoto, d. katoh, y. matsuhashi, s. seo, n. matsuno, n. uchida, a. wake, s. miyakoshi, s. taniguchi toranomon hospital (tokyo, jp) objectives: to evaluate the efficacy of unrelated cord blood transplantation (ucbt) in a reduced intensity (ri) conditioning regimen consisting of fludarabine ( mg/m²), melphalan ( mg/m²) and total body irradiation ( cgy), we analyzed retrospectively the results of adult patients with acute myeloid leukemia (aml)/ myelodysplastic syndrome (mds) in our hospital. patients and methods: we reviewed medical records of patients with aml/mds who had received single cord blood unit between november and july at toranomon acute leukaemia / myelodysplasia o myeloablative allogeneic stem cell transplantation with unrelated donors for high-risk acute myeloid leukaemia: no increase in relapse with alemtuzumab-depleted grafts p. kottaridis* ( ), k. thomson ( ) the role of t cell depletion in myeloablative allogeneic stem cell transplantation for acute myeloid leukemia (aml) remains uncertain. dose intensification may compensate for any potential loss of graft-versus-leukemia effect, and the use of t cell replete grafts is associated with significant morbidity and mortality, particularly when using unrelated donors. this study describes the results in patients with high-risk aml transplanted with an alemtuzumab-containing myeloablative regimen, using unrelated donors. median age at transplant was years and patients ( %) had donors mismatched at - hla loci. all patients were high-risk for relapse, as defined by induction failure, adverse cytogenetics, >cr , or secondary disease ( preceding myelodysplastic syndrome [mds] , therapy-related). two patients had untreated relapse at the time of transplant, and of the remaining ( %) had refractory disease. the conditioning regimen was cyclophosphamide mg/kg for days, fludarabine mg/m² for days and total body irradiation (tbi) . gy in fractions over days. stem cell source was peripheral blood in and bone marrow in , and graft-versus-host disease (gvhd) prophylaxis was with mg alemtuzumab added to the stem cells prior to infusion and cyclosporin mg/kg. median follow-up for surviving patients was months. estimated event-free survival (efs) was % at yr and % at years. acute gvhd grade ii occurred in % (no grade iii-iv) and extensive chronic gvhd in %. non-relapse mortality (nrm) was % at years with no events beyond months post-transplant, and estimated relapse risk was % at years with no events beyond months. for efs and relapse risk, the only significant variable was chemosensitivity pre-transplant, with inferior efs ( % at years for chemosensitive patients vs % for chemorefractory, p= . ) and worse relapse risk ( % at years for chemosensitive patients vs % for chemorefractory, p= . ) in those not in complete remission at transplant. there was no impact on nrm or relapse risk depending on the presence of hla mismatch, or of significant gvhd (>grade i acute, or any chronic gvhd). use of this regimen therefore permits the successful transplantation of patients with high-risk disease and hla-matched/mismatched unrelated donors, with minimal acute gvhd, relatively low nrm and no evidence of an excessive relapse rate, particularly in those in complete remission at transplant. haematopoietic stem cell transplants (hsct) using unrelated donors (ud) has become an important and viable option in the treatment of acute leukaemia (al). we have previously shown an increased risk of relapse with hla-dpb matching and independently, with nod /card genotype. in light of these data, we have analysed a larger ud-hsct cohort in order to establish the impact on outcome when both variables are considered. hla and nod /card typing was performed on al ud-hsct pairs. transplants were between and . diagnoses were all ( %) and aml ( %). % of the cohort were / hla matched and % were also hla-dpb matched. myeloablative conditioning regimens were used in % of transplants. % of conditioning protocols included t-cell depletion. bone marrow was used in % of transplants, the remaining % using peripheral blood stem cells. two forms of post-transplant immunosuppression predominated, cyclosporine a and methotrexate ( %) and cyclosporine a alone ( %). based on our previous data, the cohort was grouped according to their relapse risk, group (dpb matched; nod /card snp, n= ), group (dpb matched; nod /card wild-type (wt) or dpb mismatched; nod /card snp, n= ) and group (dpb mismatched; nod /card wt, n= ). disease relapse differed significantly between the three groups ( year: group %, group %, group %, p= . ). this finding persisted in multivariate analysis where being in either group or was protective towards relapse as compared to group (rr . ; p= . and rr . ; p= . respectively). in group (high risk), this resulted in decreased overall survival (os) ( % vs % in group , rr . ; p= . ). the best os was seen in group (low risk) where in addition to low relapse, there was increased acute and chronic graft-versus-host disease (gvhd) (p= . and p= . respectively). in this cohort, limited cgvhd was s associated with reduced relapse (p= . ) and better os (p< . ). in accordance with our theory, being hla-dpb matched and nod /card snp predicts for the worst outcome with significantly increased relapse and reduced os. the ideal pairing is hla-dpb mismatched and nod /card wt. these data suggest that prospectively typing al patients for both variables will allow the prediction of transplant outcome and will allow the effects of being independently hla-dpb matched or nod /card snp to be offset by intelligently selecting a suitable, less precarious donor. (t ) is the most common chromosomal abnormality in aml. the prognostic impact of t as a sole aberration in aml remains unclear, conferring either an intermediate or a poor prognosis. indeed, patients with t occurring with other cytogenetics abnormalities seem to have the prognosis conferred by the accompanying aberration. the aim of this study was to describe the results of allogeneic transplantation in a large series of aml patients exhibiting isolated or associated t and to compare outcome with intermediate risk aml patients exhibiting normal caryotype and receiving allo-hsct. aml patients were identified (males n= ; females n= ) with isolated (n= ) or associated (n= , favourable group n= , intermediate n= ; high-risk n= ; unknown n= ) t , allografted with an hla identical sibling (n= ) or an unrelated donor (n= ) between and and reported to the ebmt. median age was years (range: - ). median interval between allo-hsct and diagnosis was days (range: - ). the proportion of patients in cr , cr /cr or active disease was %, % and %. myeloablative and reduced intensity conditionings were performed in and patients respectively. gvhd prophylaxis consisted of csa/methotrexate or csa/mmf in %. engraftment was observed in patients ( %). grade ii and iii/iv acute gvhd occurred in % and %. chronic gvhd developed in %. with a median follow-up of months (range: - ), -year lfs, relapse rate and nrm were %, % and %. status at transplant (cr vs others) (p< . , hr . , % ci: . - . ), female sexe (p= . , hr . , % ci: . - . ) and hla sibling donor (p= . , hr . , % ci: . - . ) were significant predictors for better lfs. -year lfs was similar between aml patients with isolated or associated t ( % vs %, p= . ). whenconsidering only patients allografted in cr , -year lfs was similar between isolated t aml patients ( %, n= , median follow-up months), associated t aml patients ( %, n= , median follow-up months) and aml patients with normal caryotype ( %, n= , median follow-up months). we conclude that allogeneic transplantation in first linetherapy is a valid therapeutic option in patients exhibiting isolated or associated t . isolated or associated t do not confer bad prognosis and aml patients exhibiting such aberrations have to be considered as intermediate risk patients as they likely have the same outcome of aml patients with normal karyotype allografted in cr . regimen intensity in acute myelogenous leukaemia: addition of cgy total body irradiation to a myeloablative fludarabine/busulphan/thymoglobulin allogeneic transplant regimen reduces relapse without increasing transplant-related mortality j. russell* ( ), w. irish ( ) , l. savoie ( ) some attempts to intensify myeloablative stem cell transplant (sct) conditioning protocols for acute myelogenous leukaemia (aml) have reduced relapse only at the expense of increased transplant-related mortality (trm). a regimen of intravenous busulphan, fludarabine and thymoglobulin has been well tolerated but followed by a substantial relapse rate. we report the results of a study to enhance the antileukaemic effect of this protocol by adding a low dose of total body irradiation (tbi). patients were treated between and with fludarabine mg/m daily x and intravenous busulphan . mg/kg daily x . had additional total body irradiation (tbi) cgy x on day - or . graft-versus-host disease (gvhd) prophylaxis was cyclosporine a, methotrexate and thymoglobulin (genzyme) . mg/kg total dose. median age was (range - ) years in tbi recipients, ( - ) for the non-tbi group. there was no difference in the proportions of sct with good risk (gr, cr or cr ) recipients ( % vs %), alternative (unrelated or mismatched related) donors ( % vs %) cmv +ve donor +/or recipient ( % vs %), female donors to male recipients ( % vs %) and high risk (hr) recipient cytogenetics ( % vs %) between the tbi and no tbi groups respectively. more tbi recipients received blood cells ( % vs %, p = . ) and consequently higher cd + cell doses (median . x /kg, range . - . vs median . /kg, range . - . , p < . ). follow-up of survivors was - months (median ) for tbi recipients and - months (median ) for those not given tbi. there was no difference in incidence of acute gvhd grade ii-iv at % vs ± %, acute gvhd grade iii-iv at % vs % and chronic gvhd at % vs % with and without tbi respectively. outcomes at years are shown in the table: after adjusting for all the above risk factors the cox proportional hazard ratio for relapse was . ( % ci . - . ) in favour of tbi (p = . ). there was no effect of gvhd on relapse. the impact of tbi on relapse without affecting trm resulted in a significantly decreased risk of mortality with a hazard ratio of . ( . - . , p = . ). the hazard ratio for disease-free survival was . ( . - . , p = . ). this regimen allows some intensification by adding a low dose of tbi without an effect on trm but a reduction in relapse, confirming the importance of regimen intensity in transplantation for aml. s o long-term survival in patients suffering from aml with a complex aberrant karyotype after early allogeneic stem cell transplantation using the flamsa-ric regimen: results from a prospective phase ii trial c. schmid* ( ), m. schleuning ( ) introduction: patients suffering from acute myeloid leukemia (aml) with a complex aberrant karyotype (i.e. ≥ cytogenetic aberrations) usually show poor response to chemotherapy and have a grim prognosis. allogeneic stem cell transplanttaion (allosct) is the treatment of choice, although the unfavorable prognostic value of a complex karyotype was preserved in most studies. in contrast, in the flamsa-ric pilot trial, a post-hoc subgroup analysis revealed similar results for patients with a complex karyotype as compared to more favorable cytogenetic subgroups (schmid, schleuning et al, jco ) . therefore, a prospective phase ii trial for patients with complex karyotype aml was initiated within the german aml cooperative group, to evaluate the role of allosct, performed as early as possible after diagnosis. the flamsa-ric preparative regimen contained the sequence of intensive cytoreductive chemotherapy (flamsa), followed three days later by reduced intensity conditioning (ric; gy tbi; cyclophosphamide, atg). patients and methods: patients from centers (median age: , , range - years), with a median of (range: - ) cytogenetic aberrations were included. median time from diagnosis to transplantation was days. eight patiens had received one, had received two courses of conventional chemotherapy. stage at start of flamsa-ric was cr in , first cytogenetic relapse in , and primary induction failure in patients, including with persistent disease after high-dose arac. donors were hla-identical siblings in , matched unrelated donors in , and -ag-mismatched unrelated donors in cases. results: nineteen patients engraftet (median day: + ), one died in aplasia. patients achieved molecular cr, regenerated with blasts, and one had cytogenetically persistent disease. agvhd developed in patients, but reached > grade ii only in . five patients developed cgvhd. relapse occurred in patients. after a median follow up among survivors of (range: - ) months, patients have died from leukemia (n= ) or treatment-related causes (n= ). overall survival at one and two years from transplantation is % and %, the corresponding leukemia free survival is % and %. conclusion: to our knowledge, this is the first trial specifically addressing patients with a complex aberrant karyotype. although the numbers are small, the results suggest a promising activity of early allosct in this otherwise very unfavorable subgroup of patients with aml. the iron (fe) chelator, desferrioxamine (dfo), has been shown to have both antiproliferative and apoptotic effects in tumor cells. fe depletion results in g /s cycle arrest and cell apoptosis and dfo acts as hypoxia-mimetic agent by accumulating the hypoxia-inducible factor- alpha protein which regulates the cellular response to hypoxia by cessation of growth. in this retrospective study we evaluated the effect of dfo administration and iron overload (iro) in relapse incidence in consecutive patients (pts), allografted for malignant diseases (myeloid: , lymphoid: ). the disease phase was early in , intermediate in and advanced in pts. thirty received non ablative and ablative conditioning. peripheral blood ( ) or bone marrow ( ) grafts were donated from siblings, matched unrelated and haploidentical donors. graft vs host disease (gvhd) prophylaxis was consisted of cyclosporine or prograf plus methotrexate. non-responders or relapsed pts within months post allotransplant, were excluded. according to our center policy, pts with established engraftment and iro, as evidenced by ferritin levels> ng/ml, elevated liver enzymes or/and liver mri indicating hemosiderosis, receive dfo. among / pts with iro, were treated with dfo ( mg/kg x days/week, iv or sc for months at least). the year relapse rate (rr) was significantly lower in pts treated with dfo than in non-treated pts ( % vs. %, p= , ) and this benefit was restricted to myeloid malignancies only. in a multivariate analysis we examined dfo, chronic and acute gvhd, disease phase, graft source, type of donor, origin of malignancy and preparative regimen as risk factors for relapse. dfo administration retained its significance (p= , ) while the absence of cgvhd and the advanced disease phase were significant factors for relapse (p< , ). in order to explore whether iro affects the relapse incidence we separately examined the fe-overloaded pts who received no chelation ( ) vs the non-fe-overloaded pts ( ). the -year rr for untreated pts was % vs % for those with ferritin values < ng/ml (p= , ). iro remained as significant risk factor for relapse in the multivariate analysis (p= , ). this is the first clinical study to investigate the role of dfo in relapse incidence post allotransplant suggesting a possible role for dfo therapy post-transplant. prospective studies are needed to clarify if dfo may have a role in relapse prevention. equivalent disease-free survival results after cbt and bmt from unrelated donor using tbi containing myeloablative regimen and calsinulin inhibitors plus mtx methods in patients with mds in japan: multivariate analysis by competing risk regression models s. takahashi*, t. yamaguchi, m. monna-ooiwa, s. taniguchi, h. akiyama, t. morii, y. nagari, y. takaue, s. okamoto, k. miyamura, h. sao, t. nagamura, s. kato, t. kawase, y. the result of single institutional analysis in japan has shown cord blood transplantation (cbt) is promising in adults with hematologic malignancies including myelodysplastic syndrome (mds) and is comparable with bone marrow transplantation (bmt) from unrelated or related donors. we evaluated safety and efficacy of both bmt and cbt from unrelated donors using the data of mds patients within the japan marrow donor program and the japan cord blood bank network registry database and related those to biological and procedural factors. clinical data of patients with mds including transformed acute myelogenous leukemia who received unrelated bmt (n= ) or unrelated cbt (n= ) between and were collected. the median periods of follow-up for survivors were months for bmt and months for cbt. we analyzed the hematopoietic recovery, incidences of graft-versus-host disease (gvhd), risks of transplant-related mortality (trm), relapse and disease-free survival (dfs) using competing risk regression models.advanced primary myelodysplastic syndrome (mds) represents a subgroup of childhood mds characterized by bone marrow (bm) dysplasia and an increased blast count in peripheral blood (pb) and/or bm. allogeneic hematopoietic stem cell transplantation (hsct) is the only curative treatment. here we report the results of patients (pts) ( males/ females) enrolled in the prospective ewog mds study. according to the highest who type prior to hsct pts were classified as raeb ( ), raeb-t ( ) and mdr-aml ( ). median age at diagnosis was . yrs ( . - . ) and the median time from diagnosis of advanced mds to hsct was . mo ( . - . ) . cytogenetics revealed monosomy in pts, a complex karyotype in pts, other abnormalities in pts, no abnormalities in pts and was unknown in pts. intensive chemotherapy was given to pts prior to hsct. children were transplanted from an hla matched family donor (mfd), while the remaining were given an allograft from a matched or ag mismatched unrelated donor (ud). the preparative regimen included busulfan, cyclophosphamid and melphalan in all cases. stem cell source was unmanipulated bm, pb and cord blood in , and pts, respectively. cyclosporine-a alone was used as graft versus host disease (gvhd) prophylaxis in most children transplanted from a mfd, while the majority of pts transplanted from an ud were given cs-a, mtx and anti-thymocyte globulin. with a median follow up of . yrs ( . - . ) out of pts are alive and disease free, experienced relapse, and died of transplant related complications. the -year probability of event-free survival (efs) is . [ . - . ] while the -year cumulative incidence of transplant-related mortality (trm) and disease recurrence are . [ . - . ] and . [ . - . ], respectively. the cumulative incidence of grade ii-iv gvhd is . [ . - . ]. there is no significant difference in efs, trm and disease recurrence according to donor, mds subtype, karyotype or the treatment applied prior to hsct. age at hsct > yrs, an interval from diagnosis of advanced mds to hsct ≥ mo and the presence of gvhd contribute to a significant increase in trm. these results indicate that a large proportion of children with advanced mds can be rescued by allogeneic hsct. trm remains a main cause of treatment failure especially for children older than yrs. for these patients new strategies to reduce trm will have to be defined. introduction: patients with a history of an allogeneic haematopoietic stem cell transplant(hsct)have an increasing risk to develop a secondary cancer.the association of the hla system to cancer is well known.sibling donors share the same hla alleles, and the question is,if they have an increased tumour incidence compared to the general population.this is of concern because the possibility of a tumour induction as a result of mobilisation with g-csf has been discussed. methods: all hla-identical sibling pairs with an hsct from to for a malignant disease and living in switzerland were evaluated. general data of the donors were taken from the patient's charts of the transplant unit.donors were contacted per call or mail and asked about their current health status.in case of a malignant cancer,information on date of diagnosis,localisation,treatment was obtained.data were compared with the age-,and sex adapted cancer incidence rate of the swiss association of cancer registries(sacr).the same information was retrieved for the patients. results: pairs were identified,in ( %)the donors ( men(m), women(w))could be contacted. median observation time was . years(y)(range - y). donors were < y, between - y, between - y, between - y and > y old, hence % were < y old.seventeen ( %)donors, bone marrow and peripheral blood donors,had developed a total of cancer of different localisations (mamma,prostate,skin,bone marrow, colon,bronchus,stomach,bladder,orl).according to the incidence rate of the sacr, . tumours in m and . in w would have been expected, (m) (rr . ) and tumours (w) (rr . ) were found in donors between - y.in the age category - y, . tumours in m and . in w were expected and (rr . )respective (rr . )observed.in the subgroup of men between - y . tumour were expected and s observed (p< . ).no donor > y developed a tumour. of the donors had died, from the tumour, of cardiac disease.in patients a secondary tumour was diagnosed post hsct. conclusion: we observed a definite number of donors with a malignancy after hsc donation. absolute numbers were similar to the numbers of tumours in their transplanted siblings, observed versus expected rates were similar to an age and sex matched population except in male donors between - y by now. data from this single centre cohort remain yet inconclusive but underline the need for international collaborative donor follow up. the number of transplants is consistently increasing for aml, all, and lymphoid malignancy in all countries/regions except for vietnam. meanwhile, the number of hsct for nonmalignant diseases such as aplastic anemia and hemoglobinopathy has been stable in all countries except for iran, in which country it is recently increasing. the proportion of hemoglobinopathy in all hsct differed widely among countries; % in japan and china, % in singapore, % in taiwan, % in hong kong, % in malaysia, % in vietnam, % in iran, (all hsct until , only in taiwan) . the trends changed quite differently among countries and regions over time for other diseases, most strikingly in cml. in japan, the number of hsct for cml had consistently increased until (n= in ) and then rapidly decreased due to introduction of imatinib (n= in ). in contrast, it is dramatically increasing in china (n= in china (n= in to n= in and stable in other countries. the number of hsct for mds and multiple myeloma is consistently increasing only in japan (and iran for myeloma), but not in other countries. hsct for solid tumors had been commonly performed only in japan, but the number has decreased since (n= in , n= in ) . in summary, hsct is developing in most of the asian countries/regions. however, disease indications and trends differed widely among them probably due to different economics situations, health service systems, and availability of donors and agents such as imatinib for cml. short-and long-term side effects in the healthy donors of allogeneic haemopoietic peripheral cells mobilised with lenograstim: a single-centre experience m. martino*, g. console, e. massara, e. spiniello, i. callea, f. gatto, g. messina, t. moscato, r. fedele, g. irrera, p. iacopino u.o. ematologia con trapianto (reggio calabria, it) healthy allogeneic donors, who were mobilized with lenograstim and underwent hemopoietic peripheral cells (hpc-a) collection at our institution, were enrolled in a shortand long-term surveillance protocol for a year period. to date, donors have been assessed with a median followup of months ( - ) ; for subjects, the follow-up is > months and for subjects is > months. healthy donors received lenograstim at a median dose of . µg/kg (range - ). bone pain was reported as the most common adverse event ( . % of donors). common associated symptoms included fatigue ( . %), fevers ( % ), headache ( . %), nausea ( . %), insomnia ( . %). spleen size increased in . % of donors (> cm exceeding the marginal cost at physical examination ). all donors experienced side effect resolution within - days of lenograstim discontinuation. leukocyte mean peak values were x /l and the nadir of platelet counts reached mean values of x /l and; however, such a decrease was not complicated by bleeding manifestations. the hemoglobin concentration decreased slightly but not significantly. leukocyte and platelet counts returned to normal values in about one week. no vascular disorders and cardiac disease occurred. long-term observation included adverse events in donors after days from hpc-a mobilization and any neoplastic or not disease developed any time post donation. donors showed persistent, slight leucocytopenia until the second month, with recovery in the fourth month of follow-up. donors showed an ast and alt result . times the upper limit of normal until the second months of follow-up. transit ischemic attack occurred in donor, ( months post after donation). autoimmune event has been reported months post-g-csf (anckylosing spondylitis); donor with a history of chronic obstructive pulmonary disease developed a secondary polyglobulia ( months post-g-csf); donor developed a gastric tumor, months post-donation. no hematological malignancy was observed. in conclusion, our main findings are that the primary toxicity of g-csf administration is bone pain and that no cardiovascular events was related to the donation. with a median follow-up near to years, no hematological disease was observed in our cohort of donors. effect of the search of an unrelated stem cell donor in patients with high-risk leukaemia: prospective, singlecentre study on intention to treat analysis a.p. iori* ( ), v. valle ( ) allogeneic stem cell transplant (hsct) plays a major role in the treatment of acute leukemia (al) patients with high risk (hr) features at diagnosis or in ≥ nd complete remission (cr). between and , patients -median age years ( - ) -with hr al followed at our center and lacking a family hla compatible donor were addressed to a search of hsc donor through the bmww registry and cord blood banks. the aim of this prospective study was to assess the effect of the search on the outcome of patients with hr al on an intention to treat analysis. forty-three patients started the s search in i cr, in ii cr, in >ii cr, in relapse. fifty-five % of the patients who entered into the study in cr showed a disease relapse at a median of ( - ) and ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) months from the start of the search for patients in i-ii cr and > ii cr, respectively. sixty-nine patients ( %) underwent an hsct, % in a more advanced phase compared to the start of the search. nineteen of the patients ( %) who started the search in relapse obtained a cr and underwent an hsct. globally, % of patients failed to undergo a transplant because of lost eligibility due to disease progression. for the entire population, the year survival probability and dfs were %. disease progression was the major cause of death. when the variables affecting outcome parameters were analyzed in univariate analysis, the occurrence of relapse during the search period and the transplant procedure affected os (p= . ) and dfs (p< . ). a more advanced phase of the disease at the start of the search and no transplantation were the factors negatively affected the relapse (p= . , p< . ). for patients who underwent an hsct, the factors which negatively affected os and dfs were a relapse during the search and a more advanced disease phase at transplant. both these factors plus the disease phase at the start of the search affected the relapse for transplanted patients. in conclusion, by decreasing the length of the search ( months for patients in i-ii cr and months for patients with a more advanced phase of the disease) the risk of relapse can be reduced, thus increasing the possibility of carrying out a transplant and the transplant success. moreover, starting the search in relapse we may obtain the cr while waiting for the hsct. therefore, during the search of an hsc donor the "timing" of the transplant must be considered the major strategic factor for patients with hr al. the goal of this prospective study was to evaluate the impact of donor's and recipient's genetic polymorphisms regarding components of innate immunity on outcome of allohsct. consecutive patients with hematological malignancies, aged ( - )y, treated with allohsct from either sibling (n= ) or matched unrelated (n= ) donors were included. the conditioning was myeloablative. gvhd prophylaxis consisted of csa, mtx ± atg. donors and recipients were tested for single nucleotide polymorphisms(snp) , , of the nod /card gene, tlr ( ), tlr ( ), tlr (stop codon c t) and il r( ), as well as kir genotype. in addition, immune reconstitution was studied. os rate at y was significantly lower in allohsct with at least one activating kir mismatch compared to transplants with full compatibility ( %vs. %, p=. ). in particular, the presence activating killer immunoglobulin-like receptors (kir) in the donor with its absence in the recipient (d+r-) was associated with decreased rates of os ( %vs. %, p=. ) and dfs ( %vs. %, p=. ), as well as increased nrm ( %vs. %, p=. ). kir ds and kir ds d+r-mismatches resulted in increased risk of grade ii-iv acute gvhd, while kir ds and kir ds d+r-mismatches were associated with increased risk of chronic gvhd. d+r-activating kir mismatches correlated with increased cd +/cd + t cell ratio up to day + . in all cases of incompatibility regarding kir ds , kir ds and kir ds , t cells with expression of respective receptors could be detected up to days after allohsct. the presence of snp of the nod /card gene in the recipient was associated with decreased probability of os ( %vs. %, p=. ) and dfs ( %vs. %, p=. ) as well as increased nrm ( %vs. %, p=. ) and grade iii-iv acute gvhd ( %vs. %, p= . ). in a multivariate analysis adjusted for other potential risk factors, increasing number of d+r-activating kir mismatches as a continues variable appeared to independently influence os, dfs, nrm, grade ii-iv acute gvhd, and chronic gvhd. recipient snp of nod /card was predictive for os, dfs, nrm, grade iii-iv acute gvhd, and chronic gvhd. snps of tlr and il r genes had no impact on outcome. conclusions: both activating kir d+r-mismatches and recipient snp of nod /card appear to enhance alloreactivity and independently influence survival after allohsct. evaluation of these polymorphisms may contribute to better donor selection and optimization of the allohct procedure. the interpretation of the role of hla-dpb in unrelated haematopoietic stem cell transplantation is subject to discussion. we have investigated the role of hla-dpb allele matching on haematopoietic stem cell transplantation (hsct) outcomes in recipients who were hla -a, b, c, drb , dqb matched with their unrelated donors at the allelic level ( / ). additionally, we analysed the association of polymorphic amino acids mismatches of dpb molecule with hsct endpoints, and the permissiveness concept of zino and colleagues (zino et al., ) . dpb allele mismatches were significantly associated with an increased incidence of acute graft-versus-host disease (agvhd), transplant-related mortality (trm) and worse overall survival (os). we observed that the mismatch at amino acid position increased the risk for both agvhd and trm. risk factors for agvhd also included mismatches at positions , , , and . this is, to our knowledge, the first report of an in vivo effect of single amino acid mismatches on hsct outcomes. in our study, grouping of allelic mismatches into permissive and nonpermissive categories and their association with transplantation endpoints proved relevant for trm but not for other clinical endpoints. g.f. torelli*, r. maggio, n. peragine, m.s. de propris, b. lucarelli, m.g. mascolo, m. screnci, s. salvatori, l. malandruccolo, a.p. iori, a. guarini, r. foà sapienza university (rome, it) studies in mouse models of stem cell transplant (sct) have shown that the infusion of culture-expanded regulatory t cells (tregs) can be effective in preventing and suppressing gvhd, while still allowing a gvl effect. cord blood (cb) stem cells are now broadly used in the unrelated sct setting and key: cord- -e phpqes authors: nan title: cis annual meeting: immune deficiency & dysregulation north american conference date: - - journal: j clin immunol doi: . /s - - -z sha: doc_id: cord_uid: e phpqes nan mutations in the genes encoding proteasome subunits (psmb , psmb , psma and psmb ) have been identified as the cause of candle syndrome. these mutations lead to malfunction of the proteasome, which results in buildup of cellular waste products. it is hypothesized that dysregulation in the interferon (ifn) signaling pathway in response to this waste is the driving mechanism of the inflammatory response, and may serve as a therapeutic target in these patients. objectives: to describe a case of suspected candle syndrome successfully treated with tofacitinib. methods: retrospective chart review was conducted with respect to diagnosis, treatment and response. results: a -month old caucasian male was admitted to the hospital for evaluation of profound anemia. his medical history was significant for extreme prematurity (born at weeks from premature labor), intraventricular hemorrhage grade iv that resulted in hydrocephalus needing ventriculoperitoneal shunting, and developmental delay. he was noted to have a hemoglobin of . g/dl during a neurosurgical evaluation for routine shunt revision. he developed hemodynamic decompensation and required hospital admission for packed red-blood cell transfusion. review of systems was remarkable for intermittent pruritic macular rash, daily temperature fluctuations (fever to hypothermia), joint pain/swelling/ stiffness of multiple sites, poor weight gain, irritability, irregular breathing, abdominal distention, and regression of gross motor milestones (no longer rolling over, sitting without support, or pulling up to stand). workup excluded infections and lymphoproliferative malignancies, and he met clinical criteria for systemic juvenile idiopathic arthritis. his initial laboratory studies showed systemic inflammation (wbc x ^ /ul, hgb . g/dl, platelets x ^ /uul, crp . mg/dl, sedimentation rate mm/h, ferritin ng/ml). imaging studies revealed serositis with right-sided pleural and pericardial effusions. he also had myositis supported by imaging and elevation of muscle enzymes (ast u/l, aldolase . u/l). the patient was started on pulse iv methylprednisolone mg/kg daily x days, followed by oral prednisolone mg/kg/day and anakinra mg/kg/day with partial improvement and he was discharged home. he was readmitted . weeks later due to concerns for macrophage activation syndrome (ferritin , ng/ml) in the setting of a gastrointestinal infection and anakinra was increased to . mg/kg/day. however, he continued to have persistently elevated inflammatory markers and so the dose was increased again to mg/kg/day. three months after initial presentation, he had an upper respiratory and ear infection and became ill with generalized rash, increased work of breathing, and poor perfusion. anakinra was considered a treatment failure at that time. he required several doses of pulse steroids and initiation of tocilizumab mg/kg iv every weeks with improvement on systemic symptoms. methotrexate mg/m² weekly was added soon after for persistent arthritis and inability to wean systemic steroids. he continued to have abnormal inflammatory indices, including ferritin ( , ng/ml) and il- levels ( , pg/ml, normal - ). proband only whole exome sequencing revealed a single heterozygous mutation in the psmb gene (c.- g>a), a published pathologic variant. based on this finding, the patient was started on tofacitinib . mg orally twice a day with a dramatic response. laboratory markers of inflammation normalized, and he was able to walk within the first month of treatment. further genetic testing to detect an additional proteasome subunit variant, as well as functional testing on a research basis to demonstrate an interferon signature are being pursued. conclusions: this case highlights the value of early genetic studies in patients with autoinflammation so that initiation of targeted therapy is not delayed in efforts to achieve control of symptoms and evade future complications. this case also illustrates the challenges in diagnosing monogenic autoinflammatory disorders in young patients that present with recurrent fevers, generalized rash, arthritis, and systemic inflammation that mimic systemic juvenile idiopathic arthritis. our experience contributes to the understanding of janus kinase inhibition in type i interferonopathies. of his recurrent infections and etiology of myasthenia gravis. results of the ct chest are notable for a thymoma. thymectomy with biopsy reveals benign pathology with a mixture of type a and b cells. he continues to have persistent fatigue, generalized weakness, diplopia, diarrhea and recurrent respiratory infections after thymectomy. immunoglobulin and lymphocyte subset panels reveal hypogammaglobulinemia with absent b cells. endoscopy reveals villous atrophy and blunting without evidence of celiac disease, inflammatory bowel disease or infection suggesting autoimmune enteropathy. the constellation of clinical and laboratory features are consistent with good syndrome with evans syndrome, seronegative myasthenia gravis and autoimmune enteropathy. the patient is started on immunoglobulin replacement therapy and pyridostigmine with resolution of recurrent infections and improvement of fatigue, generalized weakness and diplopia. three years later his fatigue and evans syndrome recur with new onset loss of appetite and a thirty pound weight loss. repeat immunologic labs were notable for elevated cd , borderline low cd and highly elevated cd cells with low absolute number and fraction naïve cd and cd cells suggesting worsening combined immunodeficiency with peripheral t cell expansion. a bone marrow biopsy reveals large granular lymphocytic (lgl) leukemia and he is started on methotrexate. serum antibodies targeting ifn, and il- are negative four years after removal of thymoma. conclusions: this case is consistent with a classic presentation of good syndrome represented by thymoma, t and b cell-mediated immunodeficiency, increased susceptibility to infections and autoimmune manifestations of evans syndrome, myasthenia gravis and autoimmune enteropathy. in this case the combination of evans syndrome, autoimmune enteropathy and lgl leukemia as malignancy further worsen prognosis and is typically not seen together in good syndrome. this case depicts well the crossroad of infection, autoimmunity and malignancy in late onset immunodeficiencies. introduction/background: dedicator of cytokinesis (dock ) deficiency is a known cause of autosomal recessive hyper-ige syndrome with a combined immunodeficiency. most of the mutations in dock are lossof-function homozygous or compound heterozygous point mutations or deletions. dock deficiency has been associated with low lymphocyte counts with impaired antibody responses, as well as eosinophilia, recurrent bacterial and cutaneous viral infections, malignancies, and severe atopy. we report the case of a year old man with history of hyper-ige syndrome, severe atopy, eosinophilia, and antibody deficiency, phenotypically atypical for dock , who was noted to have two variants of unknown significance in the dock gene. objectives: we report the case of a year old man with history of hyper-ige syndrome, severe atopy, eosinophilia, and antibody deficiency, phenotypically atypical for dock , who was noted to have two variants of unknown significance in the dock gene. methods: a year-old man presented to us for evaluation of known hyper-ige syndrome. he had a long history of elevated ige, peripheral eosinophilia, severe atopic dermatitis, food allergies, asthma, severe eczema since early childhood which failed to respond to methotrexate, mycophenolate, cyclosporine, and omalizmuab, but ultimately responded to intravenous immunoglobulin (ivig). his infectious history included mrsa skin infections and one episode of pneumonia, and he reported a history of fungal skin infections but the history was unclear. initial immune workup revealed eosinophilia of , ige level (as high as , ), igg level , igm level , and iga level . he had no random antibodies to streptococcus pneumonia serotypes, but he had protective antibodies to diphtheria and tetanus. lymphocyte subsets showed cd , cd , cd , cd . he had normal mitogen stimulation to pha but decreased mitogen stimulation to candida. dna testing for a stat mutation was negative. results: we found two missense variants of uncertain significance in the dock gene ( .p.v i, nm_ . :c. g>a and .p.l v,nm_ . :c. c>g ). the first variant had previously been reported in the clinvar database as a variant of uncertain significance, and the second variant had not been previously reported in the literature to our knowledge. our assay could not determine if the two dock variants were on the same allele or on different alleles. dock protein expression testing is currently pending. conclusions: our patient presented with history of elevated ige, eosinophilia, atopy, severe eczema, and cutaneous mrsa and fungal infections. he was noted to have variants of uncertain significance in the dock gene. homozygous or compound heterozygous pathogenic variants in dock are associated with an autosomal recessive hyper-ige syndrome and combined immunodeficiency with clinical features of recurrent bacterial infections, cutaneous viral infections, severe atopic disease, as well as susceptibility to malignancy. our patient does not have all the typical features of dock deficiency and he seems to have a less severe phenotype. notably, he does not have the cutaneous viral infections or malignancy often seen in dock mutation hyper-ige cases. our case demonstrates new missense mutations, which have not previously been described in the literature, possibly causing a milder phenotype of dock deficiency. a case of igm deficiency and adult-onset still's disease negative. previous biopsy of her cervical and thoracic lymphadenopathy was unremarkable for malignancy. during her hospitalization, serum immunoglobulins were performed, which demonstrated normal levels of igg and iga, with igm level of < mg/dl (reference range - mg/dl), consistent with selective igm deficiency. liver function tests revealed an elevated aspartate aminotransferase (ast) of u/l and an alanine aminotransferase (alt) of u/l with a total bilirubin of . mg/dl and an alkaline phosphatase of u/l. her ferritin was elevated at g/l. the patient fulfilled yamaguchi criteria for aosd with three major criteria of evanescent rash, intermittent fevers in a quotidian pattern, bilateral arthralgias in the hips, knees, and ankles. she also met two minor criteria of liver abnormalities and lymphadenopathy. conclusions: selective igm deficiency is an uncommon immunodeficiency disorder associated with increased risk for autoimmune disorders. the recognition of co-morbid autoimmune illnesses in an immunodeficient patient is often complicated by a paucity of examples in the literature and potential confounding of laboratory serology analysis. we report the first case of a patient with selective igm deficiency and aosd. introduction/background: anaphylaxis to protamine is an uncommon but life-threatening complication of cardiac surgery and insulin therapy. here we present a case of recurrent protamine hypersensitivity during vascular surgery. objectives . recognize clinical signs of protamine hypersensitivity . recognize recurrent hypersensitivity to protamine as a serious complication of anesthesia methods: a year old man with a history of diabetes, previously on nph insulin, hypertension, hyperlipidemia, chronic smoking, and peripheral artery disease with multiple vascular interventions was admitted to undergo a right lower extremity saphenous vein graft bypass. three years earlier during a similar intervention, the patient had developed intraoperative hypotension after protamine sulfate administration. protamine was subsequently held for additional surgeries, however the patient was able to tolerate protamine with slower infusion one year later. for the current vascular surgery, the patient was pretreated the day of surgery with diphenhydramine and dexamethasone, and a test dose of protamine was infused prior to full dosing. the patient initially appeared to tolerate the full protamine dose, but quickly developed facial erythema and angioedema. due to concern for laryngeal edema he remained intubated and was transferred to the surgical intensive care unit, where he received additional diphenhydramine and dexamethasone. his symptoms resolved and he was successfully extubated the next morning. results: anaphylaxis to protamine is an uncommon but lifethreatening complication of cardiac surgery and insulin therapy. protamine sulfate is a polypeptide used widely to neutralize heparin anticoagulation during cardiac and vascular surgeries, and in nph insulin. severe anaphylactic or anaphylactoid reactions caused by injection of protamine sulfate are well documented in literature, and the product contains a black box warning for such. the pathophysiologic mechanisms underlying these reactions are not clear, but ige-mediated hypersensitivity appears to play a role in many reactions, and prior sensitization or cross-sensitization (eg, to fish) have been suggested. type b adverse drug reactions are idiosyncratic drug reactions and are often unpredictable, as in our patient who previously tolerated protamine but subsequently developed an adverse reaction. hypersensitivity reactions during anesthesia should be thoroughly studied to identify the responsible drug and minimize exposure in recurrent surgeries. conclusions: this case illustrates the potential for severe reactions even with newer protamine formulations, and highlights the unpredictable nature of type b adverse drug reactions. it is important for clinicians to exhibit awareness of the potential adverse effects of protamine sulfate in such situations. introduction/background: x-linked lymphoproliferative syndrome type (xlp- ) is a rare primary immune deficiency caused by loss of function in the x-linked inhibitor of apoptosis protein (xiap). common reported manifestations include recurrent hemophagocytic lymphohistiocytosis, splenomegaly, crohns-like inflammatory bowel disease, and transient hypogammaglobulinemia without reductions in major t cell or b cell repertoires, with the exception of inkt cells and mait cells. however, with only~ known cases worldwide, we are likely only beginning to understand the phenotypic spectrum of this disease. objectives: to describe additional manifestations of xlp- that expand our current understanding of its phenotype. methods: a year-old male with adult-onset, treatment refractory ulcerative colitis was evaluated in the immunology clinic for a history of recurrent sinopulmonary infections, skin abscesses, and recurrent ebv and vzv infections. extensive laboratory testing was performed in the course of his evaluation, including lymphocyte immunophenotyping, lymphocyte proliferation and cytotoxicity studies, quantification of total immunoglobulin levels and specific antibody function, hiv testing, and genetic testing. results: laboratory testing was significant for persistent cd lymphocytopenia ranging from - cells/mcl (rr: cells/mcl). total b cell count was normal but b cell subsets showed an elevation in the percentage of naïve b cells (range: . . %), low non-switched memory b cells (range: . - . %, rr: . - . %), and low to low-normal switched memory b cells (range: . %- . %, rr: . - . %), a pattern that has been seen in some autoimmune diseases. genetic testing with a commercial immune deficiency panel (invitae corp) showed a pathogenic mutation in xiap [exon , c. c>t (p.arg *)]. this mutation has previously been reported to cause a premature stop codon and reduced xiap function. the patient was referred for hematopoietic stem cell transplant and is currently awaiting transplant with a matched unrelated donor. conclusions: xlp- is typically reported as having normal t cell, b cell, and nk cell counts, but the presence of persistent cd lymphocytopenia in this patient illustrates that this is not always the case. our patient also had abnormalities in his b cell repertoire that have not been previously reported in xlp- . additionally, xlp- has been associated with crohns disease and celiac-like bowel diseases, while our case indicates that the phenotype may also include ulcerative colitis. ( ) submission id# a case report: enteroviral encephalitis as a consequence of partial humoral immunodeficiency in a chronic lymphocytic leukaemia patient treated with rituximab hadeil morsi, st immunology st , oxford university hospitals introduction/background: enteroviral (ev) infections are prevalent and usually self limited or cause mild gastrointestinal manifestations. however , in the context of primary antibody deficiency , rare cases has been reported to develop meningoencephalitis and been linked to poor outcome with fatality or chronic course. ev meningoencephalitis is even far rare reported in the era of rising secondary humoral immunodeficiency as a consequence of b cell depleting therapy e.g. rituximab and lymphoproliferative malignancies. limited treatments for ev encephalitis are available to date, apart from intravenous immunoglobulin replacement which has variable efficiency. objectives: studying such rare cases of ev meningoencephalitis as a consequence of antibody deficiencies would help to develop guidelines for intravenous immunogobulin replacement for treating these infections to improve outcome as well as predicting patients at higher risk who should be considered for prophylactic immunoglobulin therapy. methods: herein, we report a rare case of proven enteroviral meningoencephalitis following rituximab based therapy for b-cell chronic lymphocytic leukaemia and an uneventful six months period of follow up. he was found to have persistent absent b cells six months after completing six cycles of fludarabine, cyclophosphamide and rituximab therapy. interestingly, he had partial pneumococcal igg serotypes deficiency, whilst his total igg, igm and iga were all within normal limits throughout the course of the disease. the patient was treated empirically with intravenous immunoglobulin when his subtle confusion progressed to overt behavioural changes. initially his level of consciousness continued to deteriorate and he was not communicating. results: fortunately enough, the patient did have a remarkable improvement of gcs within couple of days and a slower recovery of higher mental functions e.g. memory and calculations in the next couple of months. conclusions: early suspicion and detection of entervorial meningoencephalitis in patients at risk of secondary antibody deficiency is crucial for timely ivig replacement and better outcome. patients with haematological malignancies and those on b cell depleting immunotherapy should be screened for pneumococal igg serotypes as part of secondary immunodeficiency workup. further studies on enteroviral neurological meningo/encephalitis are required to optimise ivig therapy and prognostication. furthermore, such studies provide an important asset to reveal the underlying mechanisms for humoral/b-cell mediated protective response against ev compared to other t-cell mediated viral immunity, whilst highlighting the mechanisms of immunodeficiency in cll and immunotherapy. ( ) submission id# a comparison of immune reconstitution following human placenta-derived stem cells (hpdsc) with umbilical cord blood transplantation (ucbt) vs. ucbt alone in pediatric recipients with malignant and non-malignant diseases introduction/background: ucbt is a safe and effective treatment in children (geyer/cairo et. al bjh, ) . however, due to a limited concentration of hematopoietic progenitor cells (cd +) in ucb, ucbt has been associated with delayed hematopoietic reconstitution and a higher incidence of engraftment failure. hpdscs contain a rich population of hpcs, are low in hla class i/ii expression and t-cells, and have regenerative, anti-inflammatory, and immunosuppressive properties (cairo et al bmt, ) . objectives: to determine whether ucbt + hpdsc (vs. ucbt alone) is associated with enhanced hematopoietic and immune cell reconstitution in children with malignant and non-malignant diseases. methods: immune cell reconstitution at days + , , and was assessed in children who received ucbt with hpdscs at nymc (nct , ind# ). minimum tnc was x ^ /kg ( / hla match) or . x ^ /kg ( - / hla match). immune cell subset counts at these time points were compared to those from a historical population of pediatric recipients of ucbt alone (geyer/cairo et. al bjh, ) . results: twenty four patients years were enrolled. mean age was (range, . [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] years. malignant diseases = , non-malignant diseases = . fourteen patients received myeloablative conditioning (mac) and ten patients received reduced toxicity conditioning (rtc). there were no severe adverse events associated with hpdsc infusion. two patients with non-malignant disease receiving rtc using alemtuzumab experienced primary graft failure. probability of neutrophil engraftment was . %, median day . of evaluable patients at day , the probability of platelet engraftment in neutrophil engrafted patients was %, median day . ( - ) . at days , , and , mean percent donor chimerism in whole blood was , , , and %, respectively. average percent of whole blood hpdsc chimerism was % at day and < % at beyond day . one patient with malignant disease relapsed. month overall survival was . %. there was no significant difference in cd , cd , cd , cd and cd immune cell reconstitution following ucbt + hpdsc vs. ucbt alone (image ). conclusions: these results suggest that ucbt ± hpdsc results in similar immune cell reconstitution. a larger cohort with extended follow-up would be required to confirm these preliminary findings. supported by a grant from celgene cellular therapeutics. a decade of disseminated abscesses due to mycoplasma faucium in a patient with activated pi k syndrome (apds ) introduction/background: pik r monoallelic mutations are known to be responsible for apds-like syndrome, a rare form of primary immunodeficiency presenting as combined immunodeficiency or hyper-igm like phenotype. this study reports a patient carrying heterozygous pik r mutation with early onset and long-term disseminated abscesses due to mycoplasma faucium in both peritoneal abscess and skin, with generalized involvement in neck and both upper extremities. objectives: we describe clinical management of retroperitoneal and skin abscesses before molecular diagnosis was available in a patient with a primary immunodeficiency. identification by rdna s in so-called sterile abscesses may confirm the clinical suspect of an oportunistic infection. furthermore, this study offers insight on the pik r suspicion even in the absence of higm-like phenotype. methods: a -year-old girl with -year history of recurrent peritoneal effusion, which had been drained repeatedly was admitted in our institution for a -year history of multiple supurative cutaneous and lymph node-abscesses (fig & a-c). she had prior diagnosis of agammaglobulinemia under standard subcutaneous immunoglobulin replacement therapy and subcutaneous interferon-gamma treatment. on physical examination at the age of years, she was stunted (weight and height below the rd percentile), with facial, arm skin abscesses and right fistulized axillary lymphadenopaties, - cm hepatomegaly and giant splenomegaly results: she had her first immunological work up at the age of years during one isolated episode of knee arthritis and first episode of skin abscesses. serum immunoglobulins revealed panhypogammaglobulinemia (igg< . mg/kg, iga < . mg/kg, igm < . mg/kg) with low b cell count. on her back, there was a x cm, elastic, neither painful nor tender mass. after proper assessment by ct scan and mri she had her retroperitoneal abscess drained percutaneously, and healed with sclerotherapy (percutaneous alcohol and polidocanol instilation) by the interventional radiologist ( fig b) . analysis of drained pus as well as pus of skin abscesses was made by s rdna pcr, having coincidence of . % with mycoplasma faucium . combination antibiotic therapy (doxycycline and ciprofloxacin) was started with favourable response. unfortunately skin abscesses then relapsed. t-cell phenotype only showed t-cell lymphopenia with senescent (tem & temra expansion) phenotype. whole exome sequencing revealed a heterozygous mutation, previously reported (c. + g>t) conclusions: in summary, this report emphasizes the suspicion of a combined immunodeficiency in the presence of multiple abscesses by mycoplasma, the usefulness of rdna s in order to achieve proper objectives: we describe a -year-old male patient with novel heterozygous mutation of ep gene; his first manifestations were initially characterized by infections, cytopenia and hypogammaglobulinemia suggesting a common variable immunodeficiency (cvid), but later on, persisting lymphopenia was suggestive of a combined immunodeficiency. methods: the patient was born to unrelated healthy italian parents at weeks gestation with adequate weight for gestational age. shortly after birth, he underwent several surgical procedures due to interventricular defect, aortic coarctation, double outlet right ventricle, open botallis duct, and gastroesophageal reflux. at the age of four, he came to our attention due to stomatitis. clinical examination revealed dysmorphisms (microcephaly, wide forehead, sparse eyebrows, high nasal root, low-hanging columella, thick lips, micrognathia), splenomegaly (spleen diameter . cm at abdominal ultrasound), and severe developmental delay. in the course of the infectious episode, blood tests showed leukopenia associated with neutropenia (white blood cells . /mm ; neutrophils / mm ) and thrombocytopenia (platelets . /mm ). analysis of bone marrow aspirate revealed normal differentiation of both myeloid and erythroid lineages. treatment with high doses immunoglobulin resulted in increase of platelet counts (up to /mm after month), while neutrophil counts spontaneously returned to normal when the infection resolved. however, thrombocytopenia relapsed ( /mm ) after months and intravenous high-doses of corticosteroids did not achieve normal platelets count. despite oral corticosteroid treatment started at the age of six, two episodes of autoimmune hemolytic anemia occurred. during the following six years of follow-up, the patient experienced recurrent infections (stomatitis, upper respiratory tract infections, and skin abscesses), but none of the episodes has required hospitalization. but, at the age of ten, he was admitted to the hospital because of severe cultures negative diarrhea. despite immunoglobulin replacement therapy was started at the age of fourteen, he was admitted twice due to bilateral pneumonia requiring continuous positive airway pressure and, a few months later, acute respiratory failure with evidence of mycoplasma pneumoniae and rhinovirus infections. immunological evaluation under chronic corticosteroid treatment at different time points showed persisting lymphopenia, with lymphocyte counts ranging from /mmc to /mm , thrombocytopenia (platelets ranging from /mm to /mm ), undetectable anti-diphteria and anti-tetanus toxoid antibodies, and splenomegaly. interestingly, analysis of isohemoagglutinins, revealed low titers of anti-a ( : ) at years of age, but normal immunoglobulins (igg mg/dl, iga mg/dl, and igm mg/dl). at the age of seven, reduced mitogen proliferation, hypogammaglobulinemia (igg mg/dl; iga mg/dl, igm mg/dl), increased cd +tcr+cd cd t-cell counts ( . / . %) and impaired fas mediated apoptosis as measured in two separate assays (table ) . at the age of fourteen, evaluation of b-cell subsets showed increase of cd locd lo cells and reduction of switched memory b-cells. analysis of t-cell compartment unveiled a decreased proportion of cd +ccr +cd ra+ recent thymic emigrants (rte) cells and ccr +cd ra+ naive cells, with prevalence of effector memory t-cells (ccr -cd ra-) ( table ) . interferon signature gene expression showed borderline levels of ifi (data not shown). because of the decrease of igg and the infectious episodes ivig treatment was started at age of fourteen. a molecular investigation performed by whole exome sequencing (wes) revealed a novel heterozygous missense mutation (nm_ . :c. t>c , p.met thr) in the exon of the gene ep encoding the histone acetyltransferase (hat) protein p . results: few immunological reports are available in rsts patients - . in keeping with previous data , , , our patient presented with progressive b-and t-cell lymphopenia, hypogammaglobulinemia with poor antibody response but also reduced naïve t cells, evans syndrome, splenomegaly, and defective lymphocyte apoptosis with increased dnt. at the age of seven, the patient presented the features of cvid. flow cytometry revealed expansion of cd hicd locd lo b cells, that is frequently associated with splenomegaly in cvid patients , and reduced switched memory b-cell, previously reported in a rsts patient with crebbp mutation (fig. s ). lougaris et al. reported expansion of cd locd lo b-cells in nf-kb haploinsufficiency , and this suggests in our opinion that alterations in the nf-kb pathway due to ep mutations may affect b-cell differentiation. compared to healthy controls and cvid patients with predominant infectious complications, upregulation of interferon responsive genes in cvid subgroup with noninfectious complications (i.e hematologic autoimmunity, lymphoproliferation) and lymphopenia with reduced total b cells and switched-memory b cells has been demonstrated . borderline levels of ifi expression under corticosteroid treatment represent a novel finding in rsts. interferon signature may identify and better characterize subgroup of rsts patients with autoimmune cytopenias and lymphopenia. at the age of fourteen, analysis of lymphocyte subsets revealed decreased total cd +, cd +, cd +, and of both naïve cd + and cd + cells. elevated and persisting igm levels were also observed (fig. s ). according to our data, increased igm levels may be related to high proportion of terminal differentiated igm+ cells. these data (infections requiring hospitalization, immune dysregulation, lymphopenia, reduced naïve t cells, and reduced proliferation to mitogen) together with clinical history (fisher evans syndrome and lymphoproliferation) and exclusion of known syndromic immunodeficiencies, suggested a diagnosis of combined immunodeficiency . conclusions: our case underlines the value of wes in patients with difficult phenotype-genotype correlation. no rsts typical traits were present and, prior to wes, several syndromes and immunodeficiencies were excluded. our report expands the phenotypic spectrum of ep mutations, thus in syndromic patients with clinical and immunological overlap between cvid and cid ruling out ep mutation should be advisable. furthermore, immunological work-up should be taken into consideration in rsts patients, in order to early identify immunological abnormalities that may lead to severe immune-hematological complications. introduction/background: interferon gamma receptor (ifngr )-related disorders are rare variants of mendelian susceptibility to mycobacterial diseases. although hematopoietic stem cell transplantation (hsct) is curative, it is complicated by high rates of delayed or failed engraftment thought to be due to high concentrations of interferon (ifn)-gamma. umbilical cord blood transplantation additionally increases risk of graft failure. objectives: describe a pediatric patient with non-functional ifngr who successfully underwent umbilical cord blood transplantation. methods: direct clinical care of described patient with additional electronic medical record chart review. results: the patient is a -month-old boy of yemeni descent who initially presented with significant hepatosplenomegaly and extensive lymphadenopathy, including a large mediastinal mass. he then developed salmonella enteritidis sepsis requiring numerous antimicrobials, vasopressor support, intubation and continuous renal replacement therapy. his evaluation showed a hyperinflammatory state with elevations in ferritin, ifn-gamma, scd , il- , il- , il- , il- and il- levels. maximal ferritin and ifn-gamma levels reached . ng/ml and pg/ml (normal < pg/ml), respectively. flow cytometry revealed normal expression of ifngr and il r but absent ifn-gammastimulated stat phosphorylation, suggesting defective ifngr signaling. genetic testing showed a previously unreported homozygous mutation in ifngr (c. + t>c) which affects a donor splice site in intron and is predicted to cause absent protein function. dexamethasone and a single dose of alemtuzumab ( . mg/kg) were given to decrease inflammation. he then underwent allogeneic hsct using a / human leukocyte antigen matched umbilical cord unit following a reduced-toxicity conditioning regimen of alemtuzumab ( . mg/kg), fludarabine ( mg/m ) and busulfan (auc mg/l*h). plasma ifn-gamma was undetectable prior to starting conditioning and on the day of transplant. neutrophil engraftment occurred on day + with day + posttransplant chimerism analysis of peripheral blood myeloid cells showing the presence of donor cells only. conclusions: these early results suggest that umbilical cord blood transplantation may be feasible in patients with ifngr -related disorders provided adequate control of inflammation is gained prior to transplant. introduction/background: introduction: btk is a cytoplasmic tyrosine kinase that activates phospholipase c (plc ) via phosphorylation, which ultimately leads to the activation of nfk, which is essential for b cell development and survival. mutations in btk lead to x-linked agammaglobulinemia (xla). in addition to the pleckstrin homology and tyrosine kinase domains, btk contains two src homology domains, sh and sh , which are essential for btk function. we describe a novel btk mutation (c. a>t) resulting in v a substitution in the sh domain that results in aberrant xla function with nearly normal btk protein expression. objectives: case report/results: the male proband presented with recurrent otitis media, persistent fevers and neutropenia beginning in the first year of life with an igg level of mg/dl and a lack of b cells ( cell/mm ), as demonstrated by flow cytometry. btk protein expression in monocytes, also determined by flow cytometry, was equivalent to controls. family history is significant for a maternal uncle with history of recurrent sinus infections and pneumonias with low iga and igm, low to normal ige, and an absent vaccine response. flow cytometry also showed an absence of b cells and essentially normal btk protein expression in his monocytes compared to controls. targeted high throughput sequencing of both proband and the uncle revealed a previously unreported missense mutation in exon , leading to the substitution of an aspartic acid residue for a valine (v d.) the mutation is in the highly conserved sh domain of btk (conservation phylop conservation score . .) the probands mother and his sister were shown to be carriers of the same mutation and had normal serum immunoglobulin levels and normal numbers of b cells. methods: we hypothesized that if the btk v d mutant protein was non-functional, the female carriers of the mutation would only express wild type (wt) btk in their b cells while their monocytes would express both wt and mutant btk. to test this hypothesis cd + b cells and cd + monocytes were purified from pbmc by fluorescence activated cell sorting (facs) from the sister, cdnas were generated from the respective populations of cells, and the btk cdna was sequenced using high-throughput sequencing. results: at a read-depth greater than , , cd + b cells demonstrated btk expression only from the wt allele (~ % wt btk) whereas both the wild type and mutant allele of btk were expressed at approximately equal levels in monocytes. conclusions: discussion: these results define a novel mutation in btk that nominally affects protein expression, but alters function. the v a substitution is found in the d structural element of the sh domain that is part of a hydrophobic phosphotyrosine binding pocket. a mutation in the adjacent residue, y s, has been shown to alter protein conformation and decrease binding affinity to plc by roughly -fold resulting in xla. our study, using a carrier harboring the c. a>t mutant btk, demonstrated that in contrast to mononuclear cells, b cells only expressed the wt allele. this is consistent with the loss of function of v a btk protein, thereby causing xla in both the proband and affected uncle. introduction/background: pyoderma gangrenosum (pg) is often associated with systemic autoimmune diseases but it has rarely been reported with common variable immune deficiency (cvid). while genetic analysis has been increasing in both disease domains, there has been little investigation into the genetic components associated with the cooccurrence of these entities. heterogeneous nfkb mutations have recently been identified in familial cases of cvid, though rarely have they been associated with pg. objectives: this case describes a novel nfkb mutation that may link both cvid and pg, and bolsters the recent identification of heterogeneous nfkb mutations in cvid. methods: a -year-old woman with a history of frequent skin infections in childhood presented with persistent, infected wounds following cholecystectomy. upon admission, she was started on broad spectrum antibiotics but continued to have fevers and leukocytosis. labs were also notable for elevated crp ( ; normal (n): . - ) and esr ( ; , with low c ( ; n: - ), c (< ; n: - ) , and ch (< ; n:> ). wound cultures grew multi-drug resistant coagulase negative staphylococcus. despite broad spectrum antibiotics, the wounds failed to heal. dermatology was consulted and punch biopsy revealed a dense neutrophilic infiltrate and no identifiable pathogens, which supports the diagnosis of pg. the patient was started on high dose steroids ( mg/kg/ day) and had a rapid response with decreased skin inflammation and lesion expansion. unfortunately, the patient developed posterior reversible encephalopathy syndrome (pres) on steroids and therefore pg treatment was changed to infliximab as recommended by dermatology. further laboratory testing found that the patient also had low igg ( ; n: - ) and iga ( ; , and a diagnosis of cvid was made given her clinical history of recurrent skin infections. genetic testing was pursued to evaluate additional pg therapy options and this revealed a heterozygous mutation in nfkb (c.a g; p.q q), located base pairs upstream of the splice donor site for exon twenty-one. while this mutation has not been previously identified as a pathogenic variant, similar mutations in this gene have been linked to autosomal dominant cvid. the patients father also carried a similar mutation but without any evident clinical phenotype. results: nfkb plays a crucial role in both immune and inflammatory responses. this case highlights a novel mutation in nfkb that has not been previously described as a disease-causing change. other mutations resulting in nfkb haploinsufficiency have been associated with cvid and rarely with concurrent pg, as in this case. based on the location of the mutation, it is expected that the variant causes obliteration of the normal splice site and therefore results in defective mrna that encodes p / p . interestingly, studies on p knockout mice show decreased levels of igg, iga, and ige but not igm and our patient similarly had low levels of igg and iga but normal igm. conclusions: further studies are needed to determine if there are other links to this novel nfkb mutation in patients with cvid and pg. ( ) submission id# a rapid flow cytometric analysis of dna repair proteins reveals a radiosensitive phenotype in bcl b deficiency associated with severe combined immunodeficiency (scid). introduction/background: we present the second report in the literature of a patient with immunodeficiency, dysmorphic features, growth retardation, and a homozygous variant in the dna ligase i (lig ) gene with associated absence of full-length lig protein. results: this is a now year old girl who was the fourth child of parents who are first cousins. she has one healthy older brother, a second older brother who died within hours of birth of meconium aspiration, and an older sister who died at six months of age of an upper respiratory illness / pneumonia after a history of congenital anemia, poor weight gain, cardiomegaly and hepatomegaly. she was born at weeks and spent the first five weeks of life in a neighboring hospital neonatal intensive care unit (nicu) for hepatomegaly, mild cardiomegaly (previously identified on fetal ultrasound) and congenital anemia (requiring transfusions). her exam was and remains notable for weight, height, and head circumference below rd percentile, prominent forehead, hypotelorism with epicanthal folds, downslanting palpebral fissures, and low set, posteriorly rotated and prominent ears. after discharge to home from the nicu, her course was subsequently complicated by poor weight gain, chronic diarrhea beginning after her first rotavirus vaccine, and multiple deep vein thromboses. she then became critically ill at months of age with respiratory failure, and was transferred to our institution for respiratory oscillator support. absolute lymphocyte count on admission to our institution was . k/μl (total wbc . k/μl, anc . k/μl) with agammaglobulinemia. she was diagnosed with and treated for pneumocystis jirovecii pneumonia, gradually weaned from oscillator to room air, and was discharged home five weeks later on . mg/kg every other week igg replacement. she has had no serious infections requiring hospitalization in the months since. alc has remained persistently below . k/μl with a corresponding uniform deficiency of t, b, and nk cells and no detectable trec positive t-cells. whole exome sequencing identified homozygosity for a c. g>a coding region variant in the lig gene not previously reported in the literature. a fibroblast cell line was successfully established and western blot shows an absence of full-length lig protein. further molecular characterization is in progress. conclusions: this second reported case provides further evidence for dna ligase i deficiency as a distinct clinical entity comprising immunodeficiency, dysmorphic features, and growth retardation. introduction/background: chronic mucocutaneous candidiasis (cmc) is associated with a heterogeneous group of primary immunodeficiencies. autosomal dominant stat gain-of-function (gof) mutations have been identified in up to % of patients with cmc. these mutations lead to impaired il a/f t cell immunity although the underlying mechanism is unclear. there seems to be no genotype-phenotype correlation. recently, jak inhibitor therapy has been reported to improve cmc and autoimmunity in patients with stat gof mutation. objectives: we describe an infant with cmc associated with a novel stat gof mutation. results: a -month-old girl was referred to our immunodeficiency clinic with chronic diaper rash since weeks of life, failure-to-thrive, and history of labial abscess complicated by rectolabial fistula. she was subsequently diagnosed with food protein-induced enterocolitis syndrome triggered by cows milk-based formula. laboratory evaluation revealed normal cbc with differential, lymphocyte subsets, mitogen response, immunoglobulin levels, antigen response to candida, and neutrophil oxidative burst assay. whole exome sequencing identified a de novo heterozygous variant in stat (c. t >c, p.cys arg) . further evaluation of this mutation revealed increased gas (gamma activation sequence) reporter activity in response to ifng stimulation suggesting that this is a gain-of-function mutation. the patient later developed significantly elevated liver enzymes while on fluconazole treatment, candida parapsilosis sepsis, granulomatous lesions in the liver, splenic lesions, intermittent thrombocytopenia and n o r m o c y t i c a n e m i a . s e p s i s a n d l i v e r l e s i o n s r e s o l v e d on amphotericin treatment but other findings, including tpn dependency persisted. we are planning to initiate a jak inhibitor therapy, ruxolitinib. conclusions: this case is a possible genotypic and phenotypic expansion of cmc due to stat gof. professor, the university of british columbia introduction/background: b cell cll/lymphoma b (bcl b) is a zinc finger protein transcription factor with a multitude of regulatory functions in the integumentary, central nervous, cardiac, and immune systems. it is critical for t cell lineage commitment, development, differentiation, survival, and function. in addition, it also specifies the identity and function of innate-like lymphocytes, including t cells, innate lymphoid cells (ilcs), and invariant natural killer t cells (inkt). however, little is known about its function in the human immune system, especially in the context of immune disorders. objectives: to understand the immunopathogenesis of a novel p.c y bcl b variant. methods: research study protocols were approved by our institutional research ethics board. two members of the family were enrolled (the index patient and her father). written informed consent for genetic testing and participation was provided by the parents for the child. genetic, bioinformatic, proteomic, and biochemical analyses were performed. results: we have identified the second described case of immune disease caused by a de novo heterozygous damaging variant of bcl b (p.c y). this young girl presented with intellectual disability, microcephaly, severe atopy, eczema, alopecia totalis, and brittle nails. extensive clinical immunophenotyping of patient blood showed initially unremarkable b and t cell populations. however, the patient possessed abnormal rare innate-like lymphocyte populations (inkt, dn t cells). using mass cytometry (cytof), a technique capable of concurrently analyzing parameters in a single cell, we were able to examine various innatelike lymphocyte populations, including t cells, ilc - , and nk cells. we found that the patient possessed severely compromised numbers of t cells, thus potentially implicating the p.c y variant in t cell development and function. conclusions: the identification of decreased t cells in a patient with a p.c y variant of bcl b suggests that bcl b is important for human t cell development and provides novel insights into the roles of both bcl b and t cells in regulating atopy and autoimmunity. introduction/background: adenosine deaminase deficiency caused by mutations in ada gene is a newly recognized disorder. it is associated with a spectrum of vascular and inflammatory phenotypes, ranging from early onset recurrent stroke to systemic vasculopathy or vasculitis. objectives: we describe a year old female patient with features of early onset immune thrombocytopenia (itp), autoimmune hemolytic anemia (aiha), chronic splenomegaly and variable abdominal lymphadenopathy. she was diagnosed with evans-syndrome and treated with rituximab at and month of age. from years of age she developed recurrent infections, hypogammaglobulinaemia with specific antibody deficiency, progressively decreasing class-switched memory b cells, and increased cd +cd -cd -//t cells ( %). differential diagnosis included common variable immunodeficiency (cvid) or autoimmune lymphoproliferative syndrome and therefore a broad search for causative genetic defect was initiated. the parents are first cousins of middle-eastern origin suggesting an autosomal recessive inheritance. patient was stable on long-term mycophenolate mofetil (mmf) and immunomodulatory dose ( g/kg/ month) ivig treatment. methods: genomic dna of the patient was sequenced with next generation sequencing technology. a panel of genes linked to primary immunodeficiency was analyzed. the identified variant was confirmed by sanger sequencing. results: genetic testing revealed a homozygous pathogenic mutation in the ada gene with one base pair duplication in exon (c. dup. p.arg alafs* ) that creates a frame shift starting at codon arg . the new reading frame ends at a stop codon positions downstream, likely resulting in a truncated protein. plasma ada activity of the patient was markedly reduced ( . mu/ml, normal . - . ) and confirmed the diagnosis of ada deficiency. the parents of the patient are heterozygous carriers of the same mutation. unlike most previously reported cases, this patient had an extended phenotype with no neurological evidence of vascular pathology, however brain mri revealed two silent lacunar infarct or vasculitis related changes. we speculate whether the long-term mmf or ivig therapy might be protective against vasculitis. conclusions: ada deficiency may present with a wide spectrum of clinical phenotypes beyond classical vasculopathy. the diagnosis should be considered in patients with hematological autoimmune disease, splenomegaly and/or cvid like presentation. better understanding of pathophysiology of ada deficiency may help diagnosis and targeted treatment. professor, university of california, los angeles ca introduction/background: adenosine deaminase (ada) deficiency as a cause of severe combined immunodeficiency (scid) is distinct from other forms of scid in several ways. historically, survival and clinical outcome of infants with ada scid have been inferior compared to infants with other scid genotypes. there are multiple treatment modalities available for ada-scid, including enzyme replacement therapy (ert), allogeneic hematopoietic cell transplant (hct) and experimental autologous transplant of gene corrected cells (in recent years preceded by low dose busulfan), designated as gene therapy (gt). in addition, there is a growing body of evidence for effects of ada deficiency on non-immunologic organ systems that may contribute to the historically poorer outcomes of these infants. therefore, it is important to evaluate the cohort of patients with ada scid separately from other scid cases. objectives: to capture incidence and treatment trends and to compare outcomes following available treatments for this rare inborn error of metabolism and other forms of scid, the primary immune deficiency treatment consortium (pidtc), a network of north american immunology and transplant centers, has collected standardized data for analysis. methods: ada scid patients, first treated between through , were enrolled from centers (range - subjects/site). ada accounted for % of the total pidtc scid patients treated during that time. patients were entered into either a retrospective protocol (pidtc , n= ) or a prospective protocol starting in (pidtc , n= ) . ada-scid patients who received an hct as first therapy entered either of two strata, as with other scid patients in pidtc studies, based on whether their initial presentation met definitions for typical (n= ) or leaky scid (n= ); in contrast, patients initially treated with either ert or gt were entered into a separate stratum (n= ). results: sixty-four patients ( % of all ada-scid enrollees) had ert as first therapy, but only in this cohort received ert as sole therapy; went on to have subsequent hct (n= ) or gt (n= ). there were various combinations of treatment cycles among these ada-scid patients. most received hct {+/-subsequent treatments} ( %), ert followed by hct {+/-subsequent treatments} ( %), or ert followed by gt {+/-subsequent treatment} ( %); several patients received multiple successive treatment modalities, representing either failure of initial treatment or planned progression from ert to cellular therapy. two-year survival has improved over time from % in ( - to % - (p= . ) (figure ). the survival for all other non-ada scid patients registered by pidtc over these two eras were: % ( - ) and % ( - ) . hct (either as sole therapy or after ert) accounted for > % of cellular therapies between and ; in contrast, since , gt was used as commonly as hct (n= vs. n= , respectively). there was a trend toward better two-year survival for patients receiving gt as first cellular therapy since ( %, n= , all after initial ert) compared to those receiving hct over the same time period ( %, n= , either as first therapy or after ert), although this did not achieve statistical significance (p= . ). conclusions: this study reveals the improved prognosis for patients with ada scid in recent years and the emergence of gt as a new treatment modality. further analyses are investigating the impacts of prior infection and treatment modality, including effects of conditioning, on outcomes (survival, event free survival, clinical outcomes and completeness of immune reconstitution) for ada-scid in successive eras. this study may identify optimal treatment approaches for future ada scid patients. sponsored by the pidtc, a member of the rare diseases clinical research network (rdcrn) and funded by u ai (niaid and ordr, ncats, nih). dbk has potential financial conflict of interest as a member of the scientific advisory board for orchard therapeutics and an inventor on intellectual property which ucla has licensed to orchard therapeutics related to gene therapy for ada scid. jp discloses that her spouse is employed at invitae, a dna sequencing company. intern, imam abdulrahman bin faisal university introduction/background: patients with diabetes mellitus are immunologically vulnerable population to develop different types of microbial infections. immunization has an important role in infection prophylaxis. in fact, vaccines containing thymus-dependent antigens protect patients with diabetes as they produce massive and complex immune response and feature immunologic memory. the recommended vaccinations for patients with diabetes mellitus are influenza vaccination yearly and pneumococcal vaccination. in observational studies, influenza vaccine has been shown to be similarly effective in adults < years of age with diabetes as in older patients with or without diabetes [ ] . among immunocompetent elderly, vaccine efficacy of the -valent pneumococcal conjugate-vaccine (pcv ) was modified by dm with higher vaccine efficacy among subjects with dm [ ] . the hepatitis b vaccination should be given to unvaccinated adults with diabetes mellitus who are ages to years. for older patients administration only after assessment of benefits and risks of acquiring hepatitis b virus (hbv). in fact, one review suggests that dm is associated with the progression of severe liver outcomes in adults with hbv [ ] . on the other hand, tetanus and diphtheria vaccinations should be updated. in addition to, vaccinations such tick-borne encephalitis, meningococcal infections and other infections that put in risk diabetic patients travelling abroad. accordingly, theres a variability of vaccines that can offer a preventive method to reduce morbidity, mortality, and medical expense. in our multicenter study among eastern province saudi arabia evaluated the degree of adherence of the physicians to the immunization recommendations for adult patients with diabetes mellitus type and type to increased awareness of the immunization importance in diabetic patients. objectives: to increased awareness of the immunization importance in diabetic patients. in fact, patients with diabetes mellitus are immunologically vulnerable population to develop different types of microbial infections. immunization has an important role in infection prophylaxis. methods: this is a cross sectional study involving adult patients with type and type diabetes mellitus using a questionnaire. patients will be recruited from outpatient clinics including the primary care clinics and inpatient words of king fahd hospital-al khobar and other centers in the eastern province saudi arabia. after an informed consent, baseline data will be collected. patients will be then asked if they received the recommended vaccines and who was the provider. their knowledge regarding the needed immunization will be also tested. they will then be asked about frequency of upper respiratory tract infections and pneumonia they had over the last years. results: we are expecting to find low adherence to the recommended immunizations by the physician. we may also find that patients who received the vaccines has low incidence of related infections. conclusions: increased awareness of the immunization importance in diabetic patients and adherence to immunization as part of standard care of adult patients with diabetes mellitus that offer a preventive method to reduce hospitalizations, mortality, and medical expense. - , - , and - bases. repeatability and reproducibility of the assays were . and . , respectively. . % of the target regions were covered with over x sequencing depth. we showed that the assays had . sensitivity to detect single-exon deldups and . sensitivity to detect copy number aberrations covering two or more exons. using acmg guidelines for variant classification a diagnosis was established in % patients that were sent for comprehensive panel analysis. conclusions: conclusions: our results demonstrate the analytic validity of the developed tests and show that the technology is well-suited for clinical diagnostics of inherited eye disorders. it also demonstrated a cost-effective diagnostic tool to simultaneously diagnose various types of mutations from snvs to copy number variations. introduction/background: loss of function (lof) and null mutations in orai and stim cause a rare autosomal recessive immunodeficiency by abolishing calcium release-activated calcium (crac) channel function and store-operated ca + entry. the clinical presentation is characterized by scid-like disease, dental enamel defects, muscular hypotonia and anhidrotic ectodermal dysplasia. objectives: here we present the outcome of calcium assessments performed on lymphocytes from an adult patient with unusual infections and a purported novel single pathogenic variant in orai . methods: the ca + response in lymphocytes following activation by varying concentrations of non-cross-linked anti-cd was assessed by flow cytometry. results: the patient was a year old female with a history of seasonal and medication allergies and recurrent sinus infections, who had recently developed an acute infection of her right first metatarsal joint. cultures from the joint space grew neisseria gonorrhea and atypical mycobacteria at two different time points. hiv screening was negative. follow-up testing with the mantoux test and quantiferon gold suggested that she also had latent tuberculosis infection, for which she was started on rifampicin therapy. immunologic evaluation revealed normal complete blood count and differential, normal t, b and nk cell counts, normal immunoglobulin g, a, and m levels, as well as normal responses to polysaccharide vaccines and normal t cell proliferative responses to mitogen and antigen stimulation. however, given the identification of atypical organisms from joint fluid cultures as well as latent tuberculosis (despite the lack of significant risk factors), genetic testing was recommended by her local physicians to rule out an underlying primary immunodeficiency. an invitae primary immunodeficiency panel identified a single novel pathogenic variant in orai . she was then referred to our institution for further evaluation. so far, individuals identified as heterozygous for lof or null mutations in either orai or stim have lacked any phenotype associated with crac channelopathy. however, there have been reports of abnormalities in calcium response in parents of a number of these patients, who are heterozygous for the disease-causing mutation. to examine the functional effect of the observed mutation in our patient, her freshly isolated pbmcs were loaded with indo- and the ca + response of her lymphocytes were assessed by flow cytometry. this showed a dose-dependent decrease in the patients t cell response to non-cross-linked anti-cd in comparison to the normal control, i.e. there was a clear decrease in the patients ca + response at microgram/ml in comparison to the normal control, but the decrease was rectified upon stimulation with microgram/ml or more of anti-cd . conclusions: these findings provide functional support for the identification of a new pathogenic mutation in orai . nevertheless, it is not yet clear if the mutation has any mechanistic role in the patients recent clinical presentation. introduction/background: patient : years old boy, born to nonconsanguineous parents, with hypothesis of autoimmune encephalitis (vasculitis), which was not characterized. csf has already been routed to barcelona times and to vienna time. refractory epilepsy remains (uses drugs yet). he had continuous fever during the entire hospitalization. he had adhd (took ritalin for years) and central auditory processing deficit. one year ago he began to have fever for days, improving later but evolving with bilateral otitis, predominantly on the left ear, accompanied by sinusitis. soon after that he began presenting epilepsy that worsened severely. he was interned and took acyclovir ev, associated with hydantoinate (had stevens-johnson by the drug, suspending and improving quickly). in march had uti + amo, several infections requiring meropenem + vanco, among others. received flebogamma mg/ day/ days, twice; he also received weekly rituximab for a few months. in use of carbamazepine, clobazan, phenobarbital, levotiracetam and vigabatrin times a day. personal antecedents: allergic rhinitis, bronchial asthma, recurrent otitis media, iga deficiency. patient : male, years old, born to non-consanguineous parents, with history of repeated infections in the upper respiratory tract from one year of age with prolonged dry cough and sore throat in all episodes, treated with dexamethasone without improvement, followed by antibiotics with resolution of the condition. at age five, in february , he had a new episode of sore throat and cough that lasted three months, improving spontaneously thereafter. in june , new episode of sore throat with elevated fever and whitish plaques in the tonsils, being prescribed benzetacil, without improvement in three days. he returned to the same emergency room, the antibiotic was replaced by zinnat (axetil-cefuroxime), but within the hospital he began to convulsionate, entering into an epileptic crisis, being hospitalized for days in this service and being transferred to another hospital specialized in pediatrics, intensely investigated and treated with partial improvement of the condition. he remained in coma, not walking and talking for some months, recovering slowly with physical therapy and speech therapy. of relevant exams have: reduced iga, but before it was normal (probably induced by anticonvulsants); full-body magnetic resonance imaging demonstrates generalized lymphadenomegaly and hepatosplenomegaly; pet-ct showing signs of hypoperfusion in temporal (right), occipital and cerebellum regions (suggestive of hypoperfusion -vasculitis?) my first impression was of possible mevalonate kinase (mvk) or autoimmune lymphoproliferative syndrome (alps) deficiency. with these tests described above, i believe that the first diagnostic suspicion is that the epilepsy was triggered by hemophagocytosis in the central nervous system and consequent extremely severe epilepsy, triggered by ebv infection. objectives: to compare the clinical and genetic similarities and differences of both patients. methods: both patients wer submitted to whole exome sequencing looking for the genetic alterations associated to the disease of these patients. results: wes of patient showed an allelic variant in the gene of rai (c. g.a; p.arg gln) possibly pathogenic and that could be related to the clinical features of the patient. wes of patient showed an allelic variant in the gene of cd (c. g>a; p.arg his) heterozygous and of uncertain significance. another allelic variant was found in the gene of btk (c. g>a; p.arg gln) classified as of uncertain significance and hemizygous (as btk is in chromosome x). the expression of both proteins evaluated by flow cytometry is normal, decreasing but not abolishing the possibility of pathogenicity. conclusions: the similarity of the clinical presentations is striking, but the genetic alterations are totally different, leading to the presentation of this abstract. (inf-) have been associated with adult onset immunodeficiency in patients of asian origin. pathogens that cause infections in these patients include mycobacterium avium-intracellularae (mai), non-typhoidal salmonella, cytomegalovirus, penicillium marneffei, and varicella zoster virus. methods: chart review of one patient results: we present a thirty-two-year-old filipino female, with sjogrens syndrome, penicillin and vancomycin allergy, and shellfish allergy suffering from recurrent mai spinal osteomyelitis. after three months of conservative management of back pain, mri showed an abscess at l /l and l / s vertebrae, which was diagnosed as acid fast bacilli on biopsy. she was treated for mycobacterium tuberculosis with rifampin, isoniazid, pyrizinamide, and ethambutal with subsequent change in antibiotic therapy after six weeks once cultures grew mai. mri showed spread of abscess to l -s vertebrae. three months into treatment, she was found to have a new abscess at a different spinal site, and antibiotics were again changed. two months later, she had recurrence of disease with multiple large iliopsoas abscesses, cutaneous fistulas, insufficiency fractures of the sacrum bilaterally, and osteonecrosis of the l vertebra, requiring extensive surgical debridement. medications were adjusted and she was referred to immunology eight months after initial presentation to infectious disease. laboratories were notable for elevated igg ( mg/dl) and iga ( mg/dl) and decreased cd ( cell/ul) and cd / ( cell/ul) cells. serum electrophoresis showed low albumin, elevated gamma fraction, and polyclonal gammopathy. specific antibody titers, lymphocyte proliferation assay, ch , and immunofixation were within normal limits. cytokine panel was significant for elevated il- receptor cd ( pg/ml), il- ( pg/ml), and il- ( pg/ml), and normal tnf, inf, il- , il- , il- , il- , il- , il- , il- , and il- . further serologic testing was positive for autoantibodies to inf-. patient continues treatment with iv antibiotics and is awaiting enrollment in a rituximab trial. conclusions: autoantibodies to inf-should be considered in patients of asian origin presenting with adult onset immunodeficiency, particularly those with severe or recurrent infection with mai. a high level of suspicion is required to make this diagnosis: failure to consider this disease entity leads to delay in diagnosis with potentially significant consequences for the patient. allergy/immunology, university of south florida at johns hopkins all childrens hospital introduction/background: autoimmune and inflammatory conditions are common in cvid. these have been associated with increased morbidity and mortality. objectives: we sought to further understand and evaluate the prevalence of autoimmune and rheumatologic manifestations in patients with common variable immunodeficiency (cvid). methods: we performed a retrospective analysis of cvid patients with rheumatologic/autoimmune complications in the partners healthcare cvid cohort. we evaluated baseline patient characteristics as well as autoimmune and rheumatologic complications in this cohort of patients. results: in the partners cvid cohort, / ( %) had autoimmune or rheumatologic disease. autoimmune cytopenias were reported in / ( %) patients, including coombs positive autoimmune hemolytic anemia (n= ), idiopathic thrombocytopenic purpura (n= ), and autoimmune neutropenia (n= ). autoimmune thyroid disease was reported in / ( %) patients, including hypothyroidism (n= ) and hashimotos thyroiditis (n= ). inflammatory arthritis was present in / ( %), most commonly seronegative rheumatoid arthritis (ra) (n= ), followed by inflammatory arthritis (n= ), seropositive ra (+rf or +ccp antibody) (n = ), psoriatic arthritis (n= ), and juvenile idiopathic arthritis (n= ). systemic autoantibody disease was diagnosed in patients ( %), with diagnoses including vasculitis (n= ), systemic lupus erythematosus (n= ), polymyalgia rheumatica (n= ), antiphospholipid syndrome (n= ), mixed connective tissue disease (n= ), crest/ scleroderma (n= ), myositis (n= ), sjogrens syndrome (n= ), and discoid lupus erythematosus (n= ). inflammatory neuropathy was diagnosed in patients, with small fiber polyneuropathy (n= ), uveitis (n= ), myasthenia gravis (n= ), bells palsy (n= ), and multiple sclerosis (n= ). autoimmune skin conditions were diagnosed in patients with diagnoses including psoriasis (n= ), alopecia (n= ), and vitiligo (n= ). while the mean igm was higher in the patients with autoimmune/rheumatologic manifestations than in other cvid patients ( vs mg/dl), this difference did not reach statistical significance (p= . ). conclusions: autoimmune and rheumatologic complications are present in over half of patients with cvid. increased vigilance for autoimmune and rheumatologic complications is important as survival outcome are worse in cvid patients with non-infectious complications as previously described. further evaluation of these patients to understand the mechanism of immune dysregulation is essential, as this may promote targeted therapies and improve clinical outcomes. introduction/background: immunoglobulin concentrates have been successfully used for decades to treat patients with primary or secondary immunodeficiency disorders. this treatment has substantially decreased the frequency of life-threatening infections in these patients. octanorm is a newly developed maltose-formulated subcutaneous immune globulin (human) . % liquid for the treatment of patients with primary immune deficiency (pid) and secondary immune deficiency (sid). objectives: biochemical and physico-chemical properties were investigated. methods: molecular size distribution of monomers, dimers, polymers and fragments were determined (according to european pharmacopeia (ep) monograph . ) by size exclusion chromatography (sec). igg and igg subclass concentrations were quantified by respective nephelometric methods. functionality of the igg was demonstrated by measurement of fc function, opsonophagocytosis and fc gamma receptor binding assays. dynamic light scattering measurement and size exclusion chromatography were used to characterize the integrity of the igg molecule. measurement of potential procoagulant activity was done by natem and tga (fxia-like activity). the capacity of the octanorm manufacturing process to robustly inactivate/remove pathogens was investigated in spiking experiments with prions and viruses. results: octanorm contains more than % of human igg and is characterized by an especially low content of polymers and aggregates, low viscosity, low isoagglutinin titres, low iga and igm contents with a broad spectrum of antibodies against infectious agents. it has a distribution of immunoglobulin g subclasses closely proportional to that in native human plasma. in the final product, potential procoagulant activity is not detectable. functionality and physico-chemical properties of the igg molecules were demonstrated by state-of-the-art methods. virus safety of octanorm is obtained via a combination of three validated orthogonal methods as part of the manufacturing process: cold-ethanol fractionation, solvent/ detergent (s/d) and ph treatment. a substantial depletion of prions during the manufacturing process was demonstrated. conclusions: octanorm is a state-of-the-art subcutaneous immunoglobulin. based on the excellent stability the intended shelf life of octanorm is months stored at + °c to + °c protected from light. within its total shelf life the product can be stored at room temperature up to + °c for up to six months. efficacy and very good tolerability of this new subcutaneous normal immune globulin . % were shown in a clinical phase iii study performed in centers in north america and europe. professor, sapienza university of rome introduction/background: primary antibody deficiencies (pad) are characterized by defective ig production resulting in high susceptibility to bacterial infections, especially caused by s. pneumoniae and h. influenzae. there is a limited evidence on the rate of microbial airway epithelial colonization and on the role of bacterial carriage on the development of recurrent respiratory tracts infections in such populations. objectives: the aim of this study was to investigate the prevalence of s. pneumoniae and haemophilus influenzae colonization in pad adults in italy and its clinical and immunological correlates. methods: nasopharyngeal and oropharyngeal swabs were obtained from cvid and patients with idiopathic primary hypogammaglobulinemia (iph) over years of age and under ig replacement treatment during the period october -april . presence of s. pneumoniae and h. influenzae was investigated using conventional cultural methods and rt pcr. s. pneumoniae isolates were serotyped by the quellung reaction; capsular type of h. influenzae isolates was determined by pcr. the pattern of associations between the two species and potential risk factors were investigated. respiratory infections rate was recorded over months of follow up. results: among cvid prevalence of carriage assessed by traditional culture was % and % for s. pneumoniae and h. influenzae, respectively. rt pcr allowed to identify a higher rate of carriage of s. pneumoniae and h. influenzae compared to standard culture. cvid and iph had not different rate of pneumococcal colonization, whereas cvid had higher rate of h. influenzae carriage identified by a culture methods and rt pcr. no synergistic association between s. pneumoniae and h. influenzae colonization was observed. among cvid, s pneumonia and h. influenzae carriage were associated to low iga and igm levels. cvid under antibiotic prophylaxis did not have an increased prevalence of carriage. no association was found between carrier status detected by culture and having chronic lung disease, bronchiectasis or the rate of infections during the follow up. rt pcr identified merely the association between igm levels and h. influenzae carriage. antibiotic resistance from isolated stains was also assessed. conclusions: this is the first study assessing the prevalence of s. pneumonia and h. influenzae carriage in cvid. objectives: to discuss clinical challenges in a subject with congenital hair hypoplasia detected on newborn screening with preserved t-cell mitogen responses but declining naive t-cell counts. parents are jehovah's witnesses, which adds complexity to clinical decisionmaking. methods: immune evaluation included complete blood count, lymphocyte subsets, flow cytometry to define t-cell subsets, immune globulins, repeat trec assay from peripheral blood, human immunodeficiency virus (hiv) and cytomegalovirus (cmv) dna pcr, screen for maternal engraftment, chromosome microarray analysis, lymphocyte proliferation to mitogens, t-cell proliferation to interleukins, t-cell receptor v-beta diversity analysis by spectratyping, and thymic ultrasound. results: repeat trec assay from peripheral blood on day confirmed undetectable trecs. cd + thymic emigrants were low at (reference - cells/ul). naïve cd + and cd + t cell compartments declined as did naïve cd + t cells from to cells/ul and naïve cd + t cells from to cells/ul. lymphocyte proliferation to mitogens was preserved except for the cd + response to pokeweed mitogen. interleukin proliferation of cd + lymphocytes was slightly decreased after stimulation with anti-cd ( %, normal > %). however, t-cell proliferation with other interleukins (il- , anti-cd +il- , anticd as %cd ) was preserved. the t cell receptor repertoire had intermediate diversity. antibody replacement therapy was prescribed for declining igg with no history of severe infections except for rhinovirus prior to discharge from the nicu at weeks, during which she required continuous positive airway pressure (cpap) therapy. the viral load for cmv and hiv dna were undetectable. breastfeeding was discontinued early because the mothers cmv serology revealed positive cmv igg. the child has had intermittent anemia, which is common in patients with chh. [ ] tests for autoimmune hemolytic anemia were not performed because the patient required ivig from an early age. the parents of the child are jehovahs witnesses, complicating requests for blood transfusions. conclusions: this case highlights challenges in clinical decision making for a newborn with chh identified on nbs. t-cell function is preserved, but declining naïve t cell counts and absent trecs lead us to consider hematopoietic stem cell transplant (hsct). indication and timing of elective hsct is unclear and may depend on the natural progression of disease, including infections and anemia division of immunology and allergy, the hospital for sick children introduction/background: cartilage-hair hypoplasia (chh), caused by mutations in the ribonuclease mitochondrial rna-processing (rmrp) gene, is associated with diverse immune abnormalities including combined immune deficiency (cid). most patients with chh are managed with supportive measurements, while few have received allogeneic hematopoietic stem cell transplantations (hsct). the progression of the immune abnormalities and the impact of hsct in patients with chh and cid have not been well characterized. objectives: to characterize the progression of the immune abnormalities and the impact of hsct in patients with chh and cid methods: the clinical and laboratory findings of siblings diagnosed in infancy with chh and cid due to the common a>g mutation in rmrp, including the effects of hsct performed in of them, were compared. results: both patients suffered from recurrent respiratory infections at early age with reduced t cells numbers and responses. patient # immune function continued to deteriorate leading to hsct from an hla-matched sibling at . years of age. the patient suffered acute and chronic graft versus host disease of the skin with residual mild joint contractures and scleroderma-like skin changes. seven years after hsct patient # has normal immune function. immune evaluations of patient # in the first years of life indicated mild improvement. the patient did not have a suitable related hsct donor and the family elected to continue with supportive care. at years of age, patient # is clinically well and thriving with persistent t cell abnormalities. conclusions: close monitoring of immune function in early life for patients with chh and cid as well as the availability of suitable donors assists in determining management, including hsct introduction/background: leukocyte adhesion deficiency (lad) represents a group of distinct inherited disorders, which inhibit the normal extravasation of neutrophils and their recruitment to sites of infection or inflammation. objectives presentation of case: the patient is a girl of years old with no history of primary inmunological disease (pid) in family members, including a healthy brother (currently years old). she received all the immunization schedule until year old (according to peruvian schedule). she had several admission to the hospital since newborn; the st hospital admission was at days of life with diagnosis of: sepsis, pneumony, onphalitis (leukocytes count: ), she had more hospital admission and the leukocytes count were always above . in summary, she presented other infections besides the ones admitted at hospital like: episodes of sepsis, episode of pneumony, episodes of cellulitis (left eye, left elbow, right thigh and vaginal ( ), episodes of otitis, episodes of tonsillitis, episodes of diarrhea, episodes of rhinoadenoiditis, episodes of sinusitis, episodes of gingivitis and episode of whooping cough. she received different antibiotics for treatment, even broad-spectrum as vancomycin, meropenem and cefepime. the diagnosis of leukocyte adhesion deficiency (lad) was made at year months by clinical features like delaying in separation of umbilical cord, recurrent infections and persistent leukocytosis> . flow citometry was taken at years old, resulting cd b/cd : , %. results: discussionf: or the severe phenotype, in which leukocytes express < % of normal levels of cd , death occurs at an early age because of severe infection unless patients receive bone marrow transplant. however, it does not happened with her so we suspected in a possible reversion in lad so we took a second flow citometry at years old and the results were: total leukocyte: ; linfocyte: cd b+/cd +: , %; granulocyte: cd b+/cd +: , % and the conclusion was c receptor (cd b/cd ) absent in linfocyte, monocyte and granulocyte. molecular study was taken at years old, resulting a homozygous substitution c. c>t identified in exon , causing a nonsense mutation: p.arg , confirming lad diagnosis. she is, currently, receiving profilactic antibiotic and antimicotic which has reduced considerably recurrent infections. it seems that our patient may have a mixed phenotype due to a clinical expression, which may not require hematopoietic cell transplantation (hct) despite features of the severe type. conclusions conclusion: we conclude that the clinical evolution of this patient is unsual because she has severe lad , she has not transplanted yet, profilaxis treatment has improved to decrease frecuency of infections, she exceeded life expectancy and last flow citometry confirmed that lad was not reverted. , ph.d. , luigi d. notarangelo, md , troy r. torgerson, m.d. ph.d. , ph.d. , hans d. ochs, m.d. , m.d., ph.d. introduction/background: patients with x-linked hyper-igm syndrome (x-higm) due to cd ligand (cd l) deficiency often present with low blood neutrophil counts. however, even when not neutropenic and despite immunoglobulin (ig) replacement therapy, cd l-deficient patients are susceptible to life-threatening infections by opportunistic pathogens, suggesting impaired function of phagocytes, and requiring novel therapeutic approaches. objectives: to analyze whether peripheral neutrophils from cd l-deficient patients display functional defects and to explore the in vitro effects of recombinant human interferon (rhifn)-on such cells. methods: we investigated the microbicidal activity, respiratory burst and transcriptome profile of neutrophils from cd l-deficient patients. in addition, we evaluated whether the lack of cd l in mice also affects neutrophil responses. results: neutrophils from cd l-deficient patients exhibited defective respiratory burst and microbicidal activity which were significantly improved in vitro by rhifn-. similar to humans with cd l deficiency, cd l-deficient mice were found to have defective neutrophil responses. moreover, neutrophils from cd l-deficient patients showed reduced cd protein expression and a dysregulated transcriptome profile suggestive of impaired differentiation. conclusions: our data suggest a non-redundant role of cd l-cd interaction in neutrophil development and function that could be improved in vitro by rhifn-, indicating a potential novel therapeutic application for this cytokine. methods: in this study, through the use of healthy livers, paired peritumoural tissues (pt) and intratumoural tissues (it) from hcc patients. results: increased expression of cd on nk cells was observed in intratumoral but not peritumoral regions, along with increased expression of its ligand cd and a poor prognosis. human cd + nk cells exhibited functional exhaustion, showing decreased ifn-and tnf-productions, impaired cytolysis in response to in vitro stimulation, and high gene expression of il- and tgf- with low expression of t-bet, il- , perforin and granzyme b by global transcriptomic analysis of sorted cd + and cd -nk cells. blocking tgf- specifically inhibited cd expression and reversed the dysfunction of nk cells. in addition, we compared other two receptors, cd and tigit, which share common ligand cd with cd , and found cd plays a more important role in nk exhaustion. conclusions: these findings indicate that human cd + nk cells have features of functional exhaustion, suggesting that cd -cd blockade has the potential to restore immunity against liver tumors by reversing nk cell exhaustion. introduction/background: mutations in ncf (encoding protein p phox of the nadph oxidase) result in an autosomal recessive form of chronic granulomatous disease (cgd), a rare genetic disease with impaired phagocyte production of reactive oxygen species and recurrent infections. diagnosis of p phox cgd is based on abnormal dihydrorhodamine assay and absence of p phox protein by immunoblotting; however, these assays fail to diagnose carriers of ncf mutations. instead, carrier status is inferred after the birth of a child with p phox cgd. furthermore, identification of the specific genetic defect in patients with p phox cgd is complicated by two highly conserved (> %) pseudogenes. the ncf gene has a gtgt at the start of exon , while the pseudogenes (ncf b and ncf c) delete one gt (gt). in p phox cgd, the most common mutation in ncf is gt causing c. _ delgt; p.tyr fsx . sequence homology between the wild type gene and pseudogenes precludes using standard sanger sequencing to identify specific mutations in ncf . objectives: to identify phenotypic and genotypic differences that facilitate the diagnosis of patients and carriers with p phox cgd. methods: expression of p phox in neutrophils is determined by fixing and permeabilizing whole blood with intraprep, and then incubating with either anti-p phox antibody or its corresponding isotype. alexafluor -conjugated secondary antibody is used to detect the target antigen. expression of p phox is based on the mean fluorescence intensity of cells within the neutrophil population gated using forward and side light scatter. differential expression of p phox by flow cytometry is validated using quantitative immunoblotting. to screen for the gt mutation, a droplet digital polymerase chain reaction (ddpcr) with two distinct probes recognizing either the wild-type gtgt sequence or the gt sequence was used to quantitate the ratio of gtgt vs. gt copies. a second ddpcr reaction established copy number by comparing one probe for an invariant region of ncf /ncf b/ncf c and a second probe for the single-copy telomerase reverse transcriptase gene, tert. the results of these two assays were combined to determine the total number of gtgtcontaining and gt-containing ncf copies. results: analysis of p phox expression in permeabilized neutrophils determined that neutrophils from p phox cgd patients had negligible p phox expression; neutrophils from p phox cgd carriers exhibited~ % of p phox expression compared to healthy volunteers independent of the mutation in ncf . of all p phox cgd patients tested by ddpcr, . % ( / ) exhibited copies of gtgt, . % ( / ) exhibited copy of gtgt (compound heterozygotes with non-gt mutation), and . % ( / ) exhibited copies of gtgt (two non-gt mutations). moreover, ddpcr can identify the carriers among kindreds within the gt p phox cgd families. unexpectedly, among normal subjects tested, only . % exhibited the expected copies of gtgt per total ncf /ncf b/ncf c copies, designated / ; a significant number exhibited more than two copies of gtgt ( . % with / , . % with / ); others exhibited ncf /ncf b/ncf c copy number variation ( . % with / and . % with / ). conclusions: flow cytometric analysis of neutrophil intracellular p phox staining provides a quick method to identify patients and carriers of p phox cgd. droplet digital pcr can be used to identify patients and carriers of p phox gt, the most common mutation in p phox cgd copy number variation is observed at the ncf locus among normal subjects tested. introduction/background: defects in the cd subunits of the tcr/cd complex account for a small percentage of the scid presentations. cd deficiencies are characterized by profound and t-cells lymphocytopenia, and normal numbers of b-and natural killer (nk) cells in the peripheral blood (t-b+nk+ scid). thymocyte development from double negative stage to double positive stage results arrested. affected individuals typically present with severe and opportunistic infections in the early infancy. objectives: to evaluate possible underlying immunodeficiency disorder in the setting of chronic ebv infection. methods: here we report our experience of a patient with an atypical presentation of cd deficiency. results: a -year-old girl previously healthy, first generation americanborn of gambia immigrants was referred to our clinic for further evaluation of chronic ebv infection. one year before presentation, she developed bilateral parotid enlargement, cervical, axillar, and hilar lymphadenopathy, bronchiectasis, and pulmonary nodules. relevant previous studies included: ebv viremia (pcr quant , copies), ebv igg and ea positive whereas igm and ebna were negative. peripheral blood phenotyping was not suggestive for malignant process and cervical lymph node biopsy was consistent with a reactive process without evidence of a clonal lymphoproliferative disorder and bal studies positive for ebv only. hiv and tb both negative. she was otherwise thriving with no history of recurrent infections and family history was negative for consanguinity or immunodeficiency. our evaluation revealed: normal blood counts, ebv pcr quant , copies, normal immunoglobulin levels and vaccine titers. she had a normal lymphocyte subsets, but skewed cd ra/ro consistent with low thymic output (very low cd naïve t cells ( . %), low cd + naïve t cells ( . %), and elevated temra ( . %)), poor mitogen, and antigen responses. all these findings were suggestive of a scid like phenotype; therefore, scid next generation sequencing panel was pursued, and showed a novel homozygous splice site mutation (c. + g>t) in the cd gene. conclusions: a homozygous mutation in the cd gene might not necessarily imply profound t-cell lymphopenia. though this patient did not present with classic clinical course, and immune findings; her inability to clear ebv with persistent significant viremia does support a t-cell immune deficiency. she remains at risk for ebv-associated lymphoproliferative disorder, infections, and autoimmunity. bmt will be a curative treatment option. however, it is difficult to predict evolution of clinical phenotype given the atypical presentation. this case illustrates the importance of contextual interpretation of clinical findings, laboratory data, and genetic analysis for treatment approach. introduction/background: pol is multi-subunit polymerase that includes both pole with catalytic activity and additional pole , and . this holoenzyme plays a key role in proofreading damaged dna and is required for proper dna replication in proliferating cells, such as lymphocytes. germline mutations are linked to rare cause of primary immunodeficiencies whereas somatic mutations are described in colon cancer. primary immunodeficiencies are reported pole- deficiency in members of a large consanguineous french kindred and a palestinian female with fils (facial dysmorphism, immunodeficiency, livedo, and short stature). all reported cases with pole deficiency have homozygous intronic splice site variant (c. + a>g) that result in a deletion of exon which lead to subsequent frame shift (from p.s v onwards) and a premature stop codon at position ; this transcript results in a degraded product. the proportion of the pole transcript in t lymphoblasts is significantly lower ( %) in patients then carriers or healthy individuals. objectives: hereby we describe the clinical progression and treatment challeneges of the palestinian female with pole- deficiency secondary to homozygous g.g + a>g substitution. results: initially patient presented with viral and recurrent ear infections and cmv viremia. with age, patient had less episodes of infections even with intermittent pause in immunoglobulin replacement therapy (igrt), however had multiple admissions for fever of unknown origin with negative cultures but increased ferritin level ( ng/ml) and low platelet count ( , count/ml). autoimmune and inflammatory complications were not reported among the french kindred. her skin also worsened with poor wound healing and scarring throughout that hinders igrt via subcutaneous route. furthermore, igrt with intravenous administration has resulted in symptoms of aseptic meningitis likely related to the underlying inflammatory state. in addition, patient shows decline in immune dysfunction. initially, she had normal immunoglobulin g (igg) however by years of age, she developed hypogammaglobulinemia with low igm unlike in the french family with patients. also, pneumococcal, diptheria, tetanus titers were non-protective off of immunoglobulin replacement therapy (igrt). beyond low switched memory b cells, patient also developed low b cell by yo age. t cell dysfunction continued to decline from decreased lymphocyte proliferation to antigens at yo age to fully absent lymphocyte proliferation to antigens and mitogens. naïve cd and cd compartments continue to be preserved. currently management challenges include treatment strategies for thrombocytopenia, inflammation and progressive skin disease that complicates the proper selection of route for optimal igrt. conclusions: beyond progression of immunological decline, our patient developed inflammatory phenotype with age. the progression of her immunological decline may be related to further decrease in the proportion of the wild type pole transcript and yet to be examined. overall, clinical follow up is essential in patients with pole- deficiency as phenotype can change with age and may pose new challenges. longitudinal follow up studies are needed to uncover the potential role of germline pole and pole pathogenic variants in cancer susceptibility. introduction/background: x-linked hyper-igm syndrome (xhigm) is one type of primary immunodeficiency diseases, resulting from defects in the cd ligand/cd signaling pathways. objectives: here, we retrospectively reviewed clinical, laboratory and genetic characteristic of xhigm in chinese population, thus further improving diagnosis and treatment for xhigm. methods: we collected and analyzed chinese patients, who were diagnosed and followed up in hospitals affiliated to shanghai jiao tong university school of medicine from to . targeted gene capture combined with next-generation sequencing technology and sanger sequencing were used to find out related gene mutation. results: the median onset age of these patients was months (range: days months). thirty-six percent of them had positive family histories, with a shorter diagnosis lag. the most common symptoms were recurrent sinopulmonary infections ( patients, %), neutropenia ( patients, %), protracted diarrhea ( patients, %), and oral ulcer ( patients, %). ten patients had bcgitis. six patients received hematopoietic stem cell transplantations and four of them had immune reconstructions and clinical remissions. twenty-seven unique mutations in cd l gene were identified in these patients, with novel mutations. conclusions: to our knowledge, this report provides the largest cohort of patients with xhigm in china. mutation analysis is an important tool for xhigm diagnosis. infants with scid are asymptomatic at birth and unless prompt diagnosis of the disease is made they may horrifically be vaccinated. simultaneous appearance of two live vaccine associated infections in one person is rarely reported. objectives: in this study we present two infants with scid, who received bcg and oral polio vaccines early in life before the diagnosis of immune deficiency was made. both patients developed localized and disseminated infections originating from the bcg vaccine (bcgitis and bcgiosis, respectively) and in addition were diagnosed with chronic fecal secretion of vaccine-derived polio virus (vdpv); alarmingly, in both cases, the vdpv underwent reverse mutation of attenuated sites to the neurovirulent genotype. the rarity of concomitant infection from two live vaccines in one recipient, together with the multiple complexities originating from these infections in immunodeficient infants, led us to report these cases and to inquire the pathogenesis that underlies this unique condition. methods: immunological evaluation:: cell surface markers of peripheral blood mononuclear cells (pbmcs) were measured by immunofluorescent staining and flow cytometry serum concentrations of immunoglobulins were measured using nephelometry. quantitative analysis of the tcr v repertoire was performed by means of flow cytometry. quantification of t cell receptor excision circles (trecs) was determined by real-time quantitative (rq)-pcr. genetic analysis: genetic diagnosis of scid was made for patient by direct sanger sequencing of candidate genes and retrieval of mutation in rag , and for patient by wes (whole exome sequencing), followed by validation of the dna cross-link repair c (dclre c) mutation using sanger sequencing. poliovirus detection and characterization: stool samples were collected monthly and transported to the national poliovirus laboratory located at israel central virology laboratory (icvl) for polio detection and characterization. results: in both patients, immunological workup revealed undetectable serum iga and igm levels with normal igg levels (table ) , lymphocyte immune-phenotyping using flow cytometry revealed complete absence of t and b cells with presence of nk cells (table ) . trecs, a dna marker of naive t cells and thymic output, were absent in both patients. the diagnosis of scid was made. we initiated prophylactic antibiotic (trimethoprim-sulfamethoxazole) and anti-fungal (fluconazole) treatment, as well as monthly intravenous immunoglobulin (ivig) infusions. genetic workup: the t-b-nk+ scid phenotype in patient led us to search for a mutation in the rag complex genes. indeed, a sanger sequencing of the rag gene, revealed a g t homozygous mutation which predicts an amino acid substitution from glycine to valine in position (fig. ). for patient , who had a similar t-b-nk+ scid phenotype, we identified a homozygous mutation in the dclre c gene (del. bp, c. -cttt) (fig. ) , using wes. the genetic evaluation confirmed the diagnosis of scid due to rag deficiency in patient and artemis deficiency in patient . clinical course: during hospitalization patient developed disseminated bcg related disease with skeletal lesions involving the phalanx, tibia and maxillary bones, as well as involvement of the spleen, liver and pancreas. anti-tubercular therapy with isoniazid, rifampicin, ethambutol and ciprofloxacin was initiated. due to lack of response, empirical trial of g-csf (granulocyte colony-stimulating factor), in order to enhance macrophage activity . the patient showed good response to this combination therapy. patient developed a palpable rigid mass on her left shoulder with surrounded redness at the site of the bcg vaccine at the age of months. the clinical diagnosis of bcgitis was established and triple antitubercular therapy with isoniazid, rifampicin and ethambutol was initiated with good response. throughout their hospitalization, both infants suffered from intermittent diarrhea. pcr for enterovirus was performed and detected the presence of type and type vaccine-derived poliovirus (vdpv) in patient and , respectively. during the follow-up, stool samples collection revealed accumulation of several polio virus mutations and some of the neuro-virulence attenuation sites were reverted to the neuro-virulent genotype .fortunately, both patients did not show any signs of flaccid paralysis. precautionary measures of isolation were taken to prevent spread of the vdpv. eventually, both patients underwent allogeneic bone marrow transplantation (bmt): patient had bmt without pre-conditioning, from a matched sibling donor. due to engraftment failure, a second bmt was repeated, this time successfully. on follow up examination her t cell repertoire showed a normal tcr v polyclonality and trec was detected, indicating the emergence of new t cells. due to ongoing low immunoglobulin levels this patient is still on regular ivig-infusions and prophylactic antibiotic treatment, as well as isoniazid. currently, she is well, her bcgitis is not active and her stool specimens are negative for polio. patient underwent an urgent haplo-identical bmt with alpha-beta t cell depletion without pre-conditioning, due to her unstable medical condition. flow cytometry analysis six months post bmt revealed lymphopenia of . % ( /mm³) with low mature t cells (cd %) and absent b cells. trecs were barely detected. microsatellite analysis as a marker for engraftment revealed a stable donor chimerism of %. the patient is still on immunosuppressive therapy doing well, her bcgitis is not active and her stool specimens are negative for polio. conclusions: these cases highlight the importance of early recognition of scid by neonatal screening or thorough family anamnesis, and the need to further defer the timing of administration of live vaccines. introduction/background: measurement of b cells, b cell subsets and specific antibodies produced in response to vaccination are key tests used to investigate immune system function. specific antibody production may indicate b cell functionality. objectives: in this study the correlation between the different b cell subsets and antibody responses to pneumovax® in an immunocompromised population was investigated. methods: b cell subsets were assessed by flow cytometry and pneumococcal responses measured using the vacczyme pneumococcal capsular polysaccharide (pcp) igg, iga and igm elisas (the binding site group, birmingham, uk) in primary immunodeficiency patients (pid) vaccinated with pneumovax®. lower limits of normal were defined as follows: b cells: . %; naive b cells: . %; non-switched memory b cells: . %; switched memory b cells: . %; pcp igg: mg/l; pcp iga: u/ml; and pcp igm: u/ml. results: the correlation coefficients between percentage of b cells/b cell subsets and igg, iga or igm pneumovax® responses ranged from - . to . . the percentage of the cohort achieving a normal b cell and normal igg, iga or igm response to pneumovax® was %, % and % respectively. b-cell responses were measureable in the remaining patients but they did not produce normal concentrations of pcp igg, iga or igm. further stratification of patients who achieved normal percentage of switched or un-switched b cells but who failed to achieve normal igg, iga or igm responses to pneumovax® were %, % and %, respectively, for un-switched and %, % and %, respectively, for switched b cells. conclusions: the combined measurement of b cells and response to vaccination are required to provide a detailed insight into these disorders. introduction/background: this is a -year-old boy of african american heritage with multiple congenital malformations who was diagnosed with very early-onset inflammatory bowel disease, which proved to be resistant to treatment. subsequent testing revealed a heterozygous mutation in exon of ctla . this variant has not been previously reported in the literature in individuals with ctla -related disease. heterozygous mutations in ctla cause a disease of immune dysregulation. clinical presentation is variable and may be characterized by enteropathy, hypogammaglobulinemia, granulomatous lymphocytic interstitial lung disease, lymphocytic organ infiltration in non-lymphoid organs, autoimmune cytopenias, and recurrent infections. early onset colitis has been reported with ctla -related disease and is unique to our patient's initial clinical presentation. objectives: this is a -year-old boy with cloacal exstrophy of the urinary bladder, omphalocele, imperforate anus, polydactyly, and sacral agenesis who was diagnosed with very early-onset inflammatory bowel disease at six months of age. he underwent cloacal exstrophy closure, omphalocele repair, and colostomy placement in the first week of life. at six months of age, he presented with dark tarry stools. upper endoscopy and colonoscopy revealed polyps, ileitis, and colitis. he was p-anca positive and started on sulfasalazine. unfortunately, he continued to have symptoms suggestive of active colitis, prompting a change to prednisone and azathioprine. despite therapy, his colitis persisted leading to chronic bloody diarrhea and growth failure. initial immune evaluation consisted of a normal complete blood count, serum immunoglobulin, lymphocyte subsets, and neutrophil oxidase burst assay. foxp analysis by flow cytometry showed a moderately elevated percentage of foxp +cd + cells in the cd + t cell population, but the regulatory t cell immunophenotype was normal. because suspicion was high for a monogenic immunologic disease to explain his symptoms, genetic sequencing was performed. a candidate gene panel was sequenced by next generation sequencing, and a heterozygous mutation in exon of ctla (c. g>a; p.arg gln) was found. this variant has not been previously reported but is predicted to be pathogenic in exac and polyphen databases. results: the patients diagnosis of ctla- haploinsufficiency associated with very early-onset inflammatory bowel disease has provided opportunity for targeted treatment of his specific molecular defect. given his poor response to treatment thus far, the patient will be started on abatacept. abatacept is fda approved for the treatment of rheumatoid arthritis but has been used successfully for the treatment of disease-related manifestations of ctla- haploinsufficiency. abatacept is a ctla- fusion protein formed by the igg fc region linked with the extracellular domain of ctla- ; it replaces the defective protein in ctla- haploinsufficiency. in addition, given other manifestations of ctla- haploinsufficiency including lymphoproliferative disease in non-lymphoid organs, particularly the brain and lung, we have initiated further evaluation of these organs to evaluate for disease-specific manifestations. conclusions: the protein cytotoxic t lymphocyte antigen- (ctla- ) is an essential negative regulator of t cells. heterozygous mutations in ctla cause a disease of immune dysregulation. clinical presentation is variable and may be characterized by enteropathy, hypogammaglobulinemia, granulomatous lymphocytic interstitial lung disease, lymphocytic organ infiltration in non-lymphoid organs, autoimmune cytopenias, and recurrent infections. inflammatory bowel disease may be associated with certain variants in ctla , but the literature remains limited, both by number of papers published as well as by ethnic subsets studied. clinicians who are presented with children who have early-onset colitis, and particularly inflammatory bowel disease that is difficult to treat, should consider possible genetic abnormalities, such as ctla- haploinsufficiency, as these can impact therapeutic decision-making and outcomes. introduction/background: dock deficiency is an autosomal recessive combined immunodeficiency syndrome associated with recurrent infections, eczema and other atopic diseases. the infections are usually viral, bacterial and fungal resulting in predominantly cutaneous and sinopulmonary manifestations. homozygous or compound heterozygous deletions or mutations in the dock gene ( p ) lead to abnormal cytoskeletal organization and impaired function of dendritic cells and lymphocytes. an aniline derivative belonging to the group of synthetic sulfones, dapsone has been employed in the treatment of chronic skin diseases characterized by an accumulation of neutrophils and eosinophils. methods: chart review of one patient results: we present a four-year-old male with severe eczema, persistent asthma, allergic rhinitis, as well as peanut and egg allergy suffering from recurrent skin abscesses and prurigo nodularis. abscesses began at age months and required prolonged courses of antibiotics, eight in total prior to presentation. other infectious history included otitis media and lymphadenitis. there was no history of pneumonia or other severe infections. skin abscesses responded to oral antibiotics, but recurred shortly after completing extended courses of treatment. laboratory results including quantitative immunoglobulins, specific antibody titers, myeloperoxidase staining, neutrophil oxidative burst and complement were within normal limits. laboratories were notable for elevated ige ( iu/ml) and eosinophilia ( eosinophils/microl). lymphocyte immunophenotype was significant for mild elevations in cd and cd . dock genetic sequencing by genedx revealed a heterozygous missense mutation in exon (c. c>a, amino acid change p.ala asp). abscess cultures grew methicillin sensitive staphylococcus aureus (mssa) and enterococcus faecalis. mssa was sensitive to ampicillin/sulbactam, cefazolin, gentamycin, moxifloxacin, oxacillin, rifampin, tetracycline, and vancomycin. isolate was resistant to ciprofloxacin, clindamycin, erythromycin, levofloxacin, penicillin, and trimethoprim/ sulfamethoxazole. the patient was initially treated with emollients, mupirocin washes, topical steroids, anti-histamines, bleach baths, and cephalexin three times a day. he improved clinically but was unable to tolerate cephalexin for more than ten days secondary to abdominal pain. cephalexin, with addition of probiotic, was attempted several months later and again had to be discontinued because of abdominal pain and vomiting. due to limited antibiotic options, dapsone was started after ruling out glucose- -phosphate dehydrogenase deficiency. dapsone was initiated at a dose of . mg/kg, and the patient was monitored weekly for hemolytic anemia. after two weeks of treatment, anemia was noted and the dose was decreased to mg/kg. he has continued on dapsone mg/kg once a day, with significant improvement in abscess number and severity. he has not required other systemic antimicrobials since starting dapsone. conclusions: dapsone may be considered as a treatment option for children with heterozygous dock mutation and recurrent abscesses, particularly those requiring prophylaxis, long term treatment and lacking antibiotic options. introduction/background: autosomal dominant hyper-ige syndrome (ad-hies) is a rare complicated primary immunodeficiency disease (pid). signal transducer and activator of transcription (stat ) gene mutation is found to cause ad-hies. tlr / signaling plays multiple roles in b cell proliferation, activation, class-switch recombination, and cytokine and antibody production. however, little is known about b cell response to tlr / agonist in patients with ad-hies. objectives: here, we aim to study the response of b cells from ad-hies patients to the tlr / agonist. methods: pbmcs were isolated from peripheral blood of ad-hies patients and age matched healthy controls. pbmcs were stimulated with tlr and tlr agonist (r and cpg odn , respectively), then b cells were analyzed for proliferation, the expression of certain surface markers (cd , cd and cd ), intracellular immunoglobulin levels (igm and igg) and intracellular cytokine levels (il- and il- ) by flow cytometry. results: in response to tlr / agonist, proliferative capabilities of b cells were reduced in ad-hies patients compared with those in age-matched healthy controls. besides, defective costimulatory molecule cd expression was observed in b cells from ad-hies patients. furthermore, significantly lower igm and igg levels, and il- production was detected in b cells from ad-hies patients. however, there was no significant difference in b cell apoptosis between ad-hies patients and healthy controls. conclusions: these data demonstrated that stat gene mutations in ad-hies patients contributed to impaired b cell tlr / signaling, and further affected b cell proliferation, activation, cytokine secretion and antibody production. introduction/background: antibody function is most commonly measured by a rise in antibody titers in response to antigen introduced by vaccination or natural infection. specific antibody deficiency is defined as normal serum levels of immunoglobulins with reduced or absent antibody response to antigens, often after administration of the pneumococcal vaccine polyvalent (pneumovax® ). a paucity of information exists about measurement of antigen-antibody binding, or avidity, as a measure of antibody function, including persons with recurrent sinopulmonary infections who have normal response to immunization with pneumococcal vaccine polyvalent. objectives: the aims of this study are to identify and evaluate children with recurrent sinopulmonary infections who had appropriate rise in pneumococcal antibody titers following immunization with pneumococcal vaccine polyvalent but low response by avidity, and to assess response with igg replacement therapy in these patients. methods: a retrospective chart review involved eight children with recurrent sinopulmonary infections with discordant pneumococcal antibody and avidity results following vaccination with pneumococcal vaccine polyvalent. these eight children subsequently received igg replacement therapy. results: the mean age of subjects was . (range - ) years. the mean number of serotypes with a normal antibody response (> . ug/ml) among children following immunization with pneumococcal vaccine polyvalent was . (range - ) of serotypes while the mean number of serotypes with a normal avidity response ( . ) was . (range - ) of serotypes. igg replacement was administered subcutaneously in children. the mean igg level was mg/dl. local reactions were all mild and observed in / ( %) children. no serious adverse events were reported. all children experienced a marked reduction in respiratory illnesses while on igg replacement therapy. conclusions: discordance between pneumococcal antibody titers and pneumococcal avidity titers was identified in eight children with recurrent respiratory illnesses. in children with recurrent sinopulmonary infections despite normal antibody response to pneumococcal vaccine polyvalent, measurement of pneumococcal avidity may identify patients with poor pneumococcal antibody function. igg replacement in these children was well tolerated and associated with a decrease number of respiratory infections. introduction/background: exome sequencing (es) is a powerful genomic tool that can be used to identify novel molecular causes of disorders with multiple etiologies. immunologic disorders are clinically and genetically heterogeneous, and therefore present unique diagnostic challenges both in the clinic and in the laboratory. objectives: the objective of this study was to assess the utility of es for determining the genetic etiology of immunological disorders and describe diagnostic yield and outcomes of exome sequencing (es) for patients with immunologic disorders and immunologic phenotypes. methods: a retrospective review was performed of individuals referred for clinical es for primary abnormalities of the immune system and individuals with additional phenotypes where multiple immunologic features were reported as part of the clinical picture, as determined during internal curation. analysis of clinical es data was performed by boardcertified clinical geneticists and all variants reported were confirmed by a secondary methodology. positive es outcomes required a pathogenic or likely pathogenic variant in a gene with autosomal dominant or x-linked inheritance, or compound heterozygous or homozygous pathogenic or likely pathogenic variants in a gene with recessive inheritance. results: the most common clinical indications for es in this cohort were hypogammaglobulinemia ( %), neutropenia ( %), immune dysregulation ( %), lymphopenia ( %), and combined immunodeficiency ( %). the gender distribution was % male (n= ) and % female (n= ); % of cases were pediatric (< years, n= ) and % were adult (>= years, n= ). positive results were reported in cases ( %), comparable to the overall diagnostic yield of es at our laboratory (retterer et al., ) . this included / ( %) of cases submitted as proband-only, / ( %) submitted as a trio, and / ( %) submitted as duo, quad or alternative family structure. diagnostic results spanned different genes and recurrently reported genes with identified pathogenic variants included flg (n= ), rag (n= ), sbds (n= ), lrba (n= ), stat (n= ), and pik cd (n= ). variants possibly associated with the phenotype, but not considered diagnostic, were reported in % of cases (n= ), while % of cases (n= ) had reportable findings in a candidate gene. conclusions: these results support that exome sequencing for individuals with immunologic-based phenotypes has similar diagnostic utility as the overall rate for clinical es. immunologic es cases with trio family structures have a higher diagnostic yield than proband-only cases, as inheritance information improves confidence in classification of variants as pathogenic or likely pathogenic. genetic heterogeneity, as demonstrated by the large number of distinct genes represented in this cohort of diagnostic es cases and rapid candidate gene discovery make es a valuable tool for genetic diagnosis in patients with immunological disorders. introduction/background: vaccination response to the -valent polysaccharide vaccine (ppsv ) is often used in the diagnosis of common variable immunodeficiency (cvid). unfortunately, ppsv titers are often difficult to interpret and many cvid patients are started on igg replacement therapy (igrt) before adequate evaluation. unlike ppsv titers, the enzyme-linked immunosorbent spot assay (elispot) is independent of igrt and can provide an ex vivo functional measurement of specific antibody production on the b cell level. objectives: develop and test an elispot assay to better determine vaccination response to ppsv compared to ppsv titers in cvid patients on igrt, healthy controls, and igrt patients without immunodeficiency. methods: an elispot assay was successfully optimized and used to evaluate the ppsv -specific b cell response in healthy adult controls. elispots were performed on day , and day (when plasmablasts are best evaluated). ppsv titers were measured on day , day , and day . for igrt patients, flow cytometry for b cell subpopulations will be performed to further validate the assay. results: normal controls demonstrated a significant increase in ppsv antibody spot forming units (sfu) between day and after ppsv vaccination. ppsv titers showed generally robust initial titers at day , and no significant change at day , with day results pending. conclusions: here we optimized an elispot assay that functionally measures the specific antibody response to ppsv in normal controls with ppsv titer results pending. we are actively recruiting patients on igrt (both with cvid and without immunodeficiency) for comparison. we hope to validate our assay as a useful alternative to ppsv titers that may be particularly useful when patients are on igrt. previous studies have demonstrated bal may be a sensitive diagnostic method for treatment failures of clinically diagnosed pneumonias, even if performed under treatment with empiric antibiotics, and can lead to a culture-directed change in antimicrobial therapy in the majority of cases. however, it has also been reported that at least in one piddchronic granulomatous disease (cgd)the diagnostic yield of bal was inferior to that of other diagnostic methods (marciano et al., ) . further information on the diagnostic yield of bal and other invasive procedures to obtain a specific organism diagnosis in pidd patients with suspected pulmonary infection is needed. objectives: to characterize the yield of diagnostic procedures used in pidd patients with pneumonia or other suspected pulmonary infections at the ucsf benioff childrens hospital. methods: we screened our database of pidd patients (encompassing patients seen from september , to september , cared for by the pediatric immunology service at the university of california, san francisco to identify patients with history of at least bal or other invasive diagnostic procedure (fna or open lung biopsy) for etiologic diagnosis of suspected pulmonary infection. if multiple bals were performed during a single episode of illness, only the first was used for this analysis. results: we identified pidd patients with history of at least one bal or other invasive diagnostic procedure, for a total of events. most procedures (n= ) were performed at our institution, with documented in outside hospital records. patient diagnoses included cgd ( ), nemo deficiency ( ), cvid ( ), stat -deficient ad-hies ( ), dock deficiency ( ) , and mhc class ii deficiency ( ) . of bals, / ( %) grew a predominant organism, but only / ( %) were positive for an organism believed to be causative by providers and/or for which the overall result of the bronchoscopy affected antimicrobial treatment. bal yield was highest in patients with a clinical and/or radiologic diagnosis of pneumonia ( / , %). yield was poor ( / , %) in minimally or chronically symptomatic patients referred for bal for interval changes on chest ct (i.e. suspected fungal infection). our nemo deficiency cohort had the highest rate of positive organism isolation by bal ( / , %) for diagnosis of pulmonary infection. in our cgd cohort, / ( . %) bals grew a causative organism. lung biopsy yielded positive organism isolation in / cases ( / in cgd, burkholderia cepacia and nocardia cyriacigeorgica; / in cvid) and fna in / cases of cgd (aspergillus fumigatus). fna or biopsy was done concurrently or after bal in cases; in / ( %), fna or biopsy, but not bal, was positive for a causative organism. conclusions: at our institution, bal overall had a % rate of causative organism isolation in pidd patients, but had up to a % rate of organism isolation in those with clinical and/or features of pneumonia. our rate of causative organism isolation was slightly higher than in previous reports. however, in specific instances, biopsy was still required to make a definitive diagnosis. bal may have limitations in certain populations of pidd patients, such as in cgd, but it may be a reasonable starting point in the diagnosis of pneumonia or worsening pulmonary disease in pidd. prospective research is needed to evaluate whether fna or lung biopsy, though more invasive, could result in overall shorter time to institution of appropriate directed therapy and shorter hospitalizations for specific pidd patients. introduction/background: prior to the introduction of newborn screening, cases of severe combined immunodeficiency often presented with severe or disseminated infections. herein, we report an infant from india who presented for evaluation and treatment of a periorbital mass, presumed to be malignant. however, he was found to have disseminated bacille calmette guerin (bcg), as well as multiple other infections, and was eventually diagnosed with x-linked scid. results: a -month old boy was born to unrelated parents in india. he initially presented with growing periorbital mass and fever for two weeks. pet scan showed hypermetabolic areas in the bone marrow, spleen, mesenteric lymph nodes, and left shoulder, along with the periorbital mass. biopsy of the mass revealed numerous b lymphocytes without malignant transformation. there was no evidence for malignancy on bone marrow biopsy, though numerous granulomas and a significant decrease in t lymphocytes were seen. peripheral blood flow cytometry showed a complete lack of t cells. the bone marrow biopsy was reexamined, and innumerous acid-fast bacilli were found. cultures grew mycobacterium bovis, and he was diagnosed with disseminated bcg disease. genotyping revealed a novel splice site variant in il rg, consistent with x-linked scid. further evaluation revealed multiple other infections. these included extended-spectrum beta lactamase-produce e. coli bacteremia, human metapneumovirus, cytomegalovirus, and pneumocystic jiroveci. he was also tested for vaccine-associated poliovirus because he had received the oral polio vaccine, but this was negative. four-drug therapy was started for the bcg, and the periorbital lesion completely resolved within several weeks. additionally, he was treated with intravenous ribavirin for the human metapneumovirus, and sulfamethoxazole-trimethoprim for the p. jiroveci. although the cmv was initially treated with ganciclovir, the virus eventually developed resistance and required treatment with foscarnet. due to his many infections, he underwent haploidentical stem cell transplant plus donor lymphocyte infusion from mom. this transplant failed to engraft, but a matched unrelated donor was eventually found, and our patient received a second bone marrow transplant. he currently is showing signs of engraftment, and is continuing to be treated for his multiple infections. conclusions: disseminated mycobacterial disease due to the bacille calmette-guérin (bcg) vaccination has been noted in cases of xlinked scid previously, often consisting of lymphatic, skin, and pulmonary manifestations. however, there is a paucity of published cases presenting with multiple other co-infections. nor are there reports of disseminated bcg presenting as a localized mass. this case highlights the unique considerations when evaluating a patient with immunodeficiency from another country, where vaccination practices and epidemiology differ. specifically, unusual presentations of infections or masses may warrant investigation for severe immunodeficiency. prototypic t-b+nk+ immune phenotype is caused by mutations in the iil ra gene. the il signaling has an important role during t-cell development in the thymus, contributing to cell proliferation and survival. in addition, in mouse models, the rearrangement of the t cell receptor genes, specifically the gamma locus (trg), has been shown to be regulated by il signaling. similar to other scid phenotypes, patients with il ra deficiency are predisposed to acquire opportunistic infections early in life and to display poor outcome and death, unless their immune system is restored by mean of hematopoietic stem cell transplantation (hsct). while most patients with il ra mutations have full il ra deficiency resulted in a severe t cell depletion, some have a partial deficiency with residual cells or leaky phenotype, or even present symptoms later in life. objectives: here we report two non-related infants detected by the israeli national newborn screening program for scid. despite having similar il ra missense mutation (f l) they displayed distinct clinical and immunological course, resulted in a completely different treatment approaches; observation in one patient with an unusual recovery, and hsct in the other patient methods: patient lymphocytes were examined for subset counts, thymic output (via excision circles), t cell receptor repertoire diversity (tcrb) and il ra expression and function. the pathogenic il ra mutation was found by whole exome sequencing (wes). high throughput immunesequencing was performed to characterize the trg repertoire. results: we established patients' diagnosis by validating the pathogenic il ra mutation, and showing profoundly impaired t cell immune work up and abnormal il ra expression and function, determined by stat phosphorylation assay. all these measurements improved over time for the patient with less severe clinical presentation, while remained low in the patient with the severe phenotype. characterization of their trg immune repertoire using high throughput immune-sequencing revealed restriction of t cell receptor repertoires of both patients upon their initial diagnosis, compared to healthy controls. however, skewed usage of variable (v) gene segments and abnormalities of the cdr length distribution were more prominent in the patient with the severe phenotype. conclusions: these studies illustrate the gap that exists in our understanding of other non-genetic parameters that may influence disease course and severity in patients harboring a similar genetic defect. furthermore, the results reinforce the role of il- signaling not only in cell proliferation but also in trg rearrangements introduction/background: dock immunodeficiency syndrome is primary immunodeficiency disease caused by loos of function mutations in the dock gene, which was known to play a critical role in the survival, proliferation and function of several types of immune system, especially lymphocyte. dock immunodeficiency syndrome is the most common cause of autosomal recessive hyper-immunoglobulin e syndromes (hies) and mainly expressed as recurrent infections and severe allergic disease affecting the skin. in addition, autoimmune features including systemic lupus erythematosus, hemolytic anemia or idiopathic thrombocytopenic purpura may be presented in dock immunodeficiency syndrome. objectives: we report a case of -month-old boy diagnosed with dock immunodeficiency syndrome, which was initially expressed as sle without recurrent skin infections. methods: a child with atopic dermatitis was admitted to another hospital because of fever lasting more than days accompanied by swelling of the hands and foot. he developed whole body edema, perioral purpura and oliguria. complete blood count was normal and blood urea nitrogen , creatinine and albumin levels were normal on his laboratory findings. however, c-reactive protein level was high at . mg/dl, coagulation parameters were abnormal (prothrombin time . sec; activated partial thromboplastin time . sec, d-dimer . ug/ml). so he was transferred to our hospital for further examination and treatment on the th day of fever. additionally, ulceration of the tonsil and maculopapular rashes on the abdomen and both legs were observed in physical examination. we suspected a meningococcal infection and administered antibiotics. however, no bacterial isolates were identified in the blood and csf culture test. fever persisted despite the administration of antibiotics. we checked immunoglobulin level, complement level and autoantibodies based on fever of unknown origin. immunoglobulin g, m and a were normal, complement fractions c , c , and ch were low at . mg/ dl, . mg/dl and . u/ml, respectively. antinuclear antibodies were positive at : with homogenous fluorescence. anti-ds dna antibody was positive at . . the tests for anti-ssa, anti-ssb, anti-ribonucleoprotein, anti-scleroderma , and anti jo antibodies were all negative. he developed leukopenia and thrombocytopenia over time. results: he was treated with steroid satisfied diagnostic criteria for systemic lupus erythematosus (sle) and fever subsided. he was confirmed lupus nephritis by renal biopsy later. because of onset of sle at the young age, we performed diagnostic whole exome sequencing and multiplex ligation-dependent probe amplification assays. conclusions: he was confirmed dock gene deletion (a deletion on one allele and point mutation on the other allele). he is preparing for hematopoietic stem cell transplantation due to autoimmunity and nonreversible parenchymal organ damage form infections although he has not yet experienced a life-threatening infection. introduction/background: during immunological investigation, it is important to distinguish those individuals who may hav e hypogammaglobulinemia (hypo) without fulfilling the criteria for the severe antibody deficiency common variable immunodeficiency (cvid) e.g. unspecified hypogammaglobulinemia from those with cvid. objectives: since low igg concentrations may support a diagnosis of cvid, we sought to investigate whether the measurement of additional igg subclass antibodies (iggsc) may provide further discrimination between patients with cvid and those with hypo. methods: iggsc concentrations were measured in serum samples from cvid patients (n= , : . m:f, median age . years, range - ) and hypo patients (n= - , : . m:f, median age . years, range - ). results: cvid patients had lower median iggsc concentrations for all iggsc: igg mg/dl (range - ) vs mg/dl (range - ); igg - mg/dl (range - ) vs mg/dl (range - ); igg - mg/dl (range - ) vs mg/dl (range - ); igg mg/dl (range . - ) vs mg/dl (range . - ). this was significantly lower for igg and igg (p= . and . respectively). a higher percentage of cvid patients had iggsc below the lower limit of the normal range compared to hypo patients: igg ( vs . %); igg ( vs . %); igg ( vs . %); igg ( . vs . %). % of cvid patients had low concentrations of or more iggsc vs only . % of hypo patients (p= . ). . % and . % of hypo patients had low levels of or iggsc, respectively. conclusions: igg subclass measurements may have some utility in distinguishing cvid patients from hypogammaglobulinemia patients. introduction/background: the thymus is often removed during cardiac surgery for repair of congenital heart disease, but the extent of tissue removed varies between procedures and surgeons. previous studies have shown decreased t cell counts after thymectomy but there is limited data on the effect of thymectomy on t cell receptor excision circle (trec) levels and infection risk. objectives: to determine the effect of partial and complete thymectomy during cardiac repair surgery on trec levels and infection risk. methods: a retrospective study of electronic medical records was performed on children who received cardiac surgery before age one at new york presbyterian/morgan stanley childrens hospital between / / and / / . patients with heart transplant or primary immunodeficiency were excluded. data was recorded on trec levels (abnormal trec < copies/μl on new york state newborn screen), number of positive cultures, viral pcr panels, and infiltrates on chest x-ray. patients were followed for a minimum of six months after cardiac surgery. study was irb approved. results: cardiac surgery was performed on patients and data was available for patients. of patients included, had a partial and had a complete thymectomy. trec levels after surgery were recorded for patients. only % of patients had an abnormal trec level on newborn screen. there was no difference between partial and complete thymectomy on risk of abnormal trec (p = . ) and mean total number of infections at months (p = . ). conclusions: thymectomy rarely causes low trec levels in children undergoing cardiac surgery. complete thymectomy does not significantly increase infection rates in these children compared to partial thymectomy. these findings are possibly due to presence of ectopic thymic tissue or thymic regeneration and are reassuring for children undergoing complex cardiac surgeries. however, long-term follow-up of these children will be necessary to determine residual function of the thymus and clinical response. introduction/background: new sequencing techniques have revolutionized the identification of the molecular basis of primary immunodeficiency disorders (pid), not only by establishing a gene-based diagnosis, but also by facilitating defect-specific treatment strategies, improving quality of life and survival, and allowing factual genetic counseling. because these techniques are generally not available for physicians and their patients residing in developing countries, collaboration with overseas laboratories has been explored as a possible, albeit cumbersome, strategy. objectives: we sought to determine whether blood collected by guthrie cards could be shipped across continents by regular airmail to a cliaapproved laboratory for confirmatory testing. methods: blood was collected and blotted onto the filter paper of guthrie cards by completely filling three circles. we enrolled male patients with presumptive x-linked agammaglobulinemia (xla) cared for at the vietnam national children's hospital, their mothers and several sisters for carrier analysis. dbs were stored at room temperature until ready to be shipped together, using an appropriately sized envelope, to a cliacertified laboratory in the us for sanger sequencing. the protocol for sanger sequencing was modified to account for the reduced quantity of gdna extracted from dbs. results: high-quality gdna could be extracted from every specimen. btk mutations were identified in of patients studied, confirming the diagnosis of xla in % of the study cohort. type and location of the mutations were similar to those reported in previous reviews. the mean age when xla was suspected clinically was . years, similar to that reported by western countries. two of mothers, each with an affected boy, had a normal btk sequence, suggesting gonadal mosaicism. conclusions: dbs collected on guthrie cards can be shipped inexpensively by airmail across continents, providing sufficient high-quality gdna for sanger sequencing overseas. using this method of collecting gdna we were able to confirm the diagnosis of xla in of vietnamese patients with the clinical diagnosis of agammaglobulinemia. introduction/background: znf is a positive regulator of stat expression. it has recently been described that nonsense mutations in znf account for the stat -like phenotype in four autosomalrecessive kindred. patients presented with reduced stat expression and diminished th cell numbers, in absence of stat mutations. objectives: here, we decribed a turkish case having nonsense mutation in znf developed dual malignancy in years. results: a -year-old female patient presented with severe eczema, recurrent cold skin abscesses, herpetic skin lesions, sinopulmonary infection, otitis media and hearing loss. the parents are cousins. two younger sisters of the index case had eczema and recurrent skin infections since their infancy period. physical examination of the patient revealed severe eczema, high palate, micrognathia, maxillary hypoplasia and hearing loss. laboratory findings showed reversed cd /cd ratio, high serum ige level ( . u / l) and low igg level ( mg / dl). the patient was diagnosed as hyper ige syndrome and ivig therapy ( mg / kg, every weeks) was initiated because of igg subgroup deficiency and recurrent sinopulmonary infections. at the age of , a polypoid mass filling the left nasal cavity was detected in her examination. paranasal sinus ct revealed a mass obliterating the left nasal cavity, left ethmoid sinus and frontal sinus. immunohistochemical stains showed a small round cell malignant tumor in the nasal cavity. she was treated with chemoradiotherapy successfully. a homozygous nonsense mutation has been detected at exon in the znf gene (c. c> t) (kindly provided by grimbacher's lab) very recently. she developed papillary thyroid carcinoma two years after completing the cancer therapy. conclusions: the relationship between znf defect and cancer development is unknown. the development of a second malignancy in this patient for a short time of completing the therapy might imply us a tendency for malignancy in znf patients. additionally, the likelihood of increased radiosensitivity in these patients should be taken into consideration. introduction/background: prometic % igiv contains purified igg, % as monomer; with a distribution of igg subclasses proportional to that in native human plasma. we report the interim results from a phase trial in the usa of prometic % igiv in adults and children with pidd. objectives: this was a phase , single-arm, open-label, multicenter trial to evaluate the safety, tolerability, and efficacy of prometic % igiv in adults and children with pidd. methods: adults and pediatric subjects with pidd on a stable dose of igg replacement therapy ( - mg/kg) for at least months with serum igg trough levels > mg/dl were included. subjects received prometic % igiv every to weeks for approximately year at the same dose and schedule as their previous igg replacement therapy. results: an interim analysis was conducted when data were available on adult subjects who received at least one dose of prometic % igiv (total of infusions), with subjects receiving at least months of treatment (exposure = . subject years). at this time, pediatric exposure was only . subject years. there were no serious bacterial infections (sbis) reported, and rate/yr of infections other than sbis was . which was comparable to rate while on commercial product ( . ) all subjects achieved an igg trough level > mg/dl. there were no deaths, and no subject had a study drug-related serious adverse event or an adverse event that resulted in permanent discontinuation of study drug. a total of adverse reactions (ar) ( . /infusion) occurred in subjects ( . %), with infusions ( . %) associated with an ar. most infusions ( . %) were completed without a rate reduction. most ars were mild or moderate in severity, with severe ars ( . /infusion) occurring in subjects ( . %). the most frequent ars were headache ( . % subjects or . /infusion) and fatigue ( . % subjects or . /infusion). conclusions: in adults treated with prometic % igiv, there were no sbis and infusions were well tolerated. director, sean n. parker center for allergy and asthma research at stanford naddisy foundation, professor of medicine and pediatrics, stanford university introduction/background: desensitization to food allergies is being studied in clinical trials using oral immunotherapy (oit). there are limited data regarding the immune changes associated with successful oit. epigenetics involves heritable changes in gene function without modification of the underlying dna sequence. this is mediated by methylation, histone modification, or changes in microrna. objectives: to study methylation changes in the loci of four key genes of immune cells involved in allergy, interleukin (il- ), interferon gamma (ifn-g), forkhead box protein (foxp ), and interleukin (il- ), comparing baseline to post-oit. methods: we completed a phase , randomized, placebo-controlled, multi-food oit trial using omalizumab, an anti-ige biologic, to facilitate desensitization for multi-food allergic individuals. double-blind, placebo-controlled food challenges (dbpcfcs) to multiple foods were conducted at entry and after weeks of treatment, the primary endpoint. omalizumab (n= ) or placebo (n= ) was administered for weeks, with oit for - foods starting weeks after the beginning of omalizumab or placebo. after weeks ( weeks of oit), participants underwent dbpcfcs to their offending foods. treatment failures (n= ) were offered open-label omalizumab. pyrosequencing of bisulfite treated genomic dna purified from pbmcs from each participant at baseline and post-oit was undertaken to investigate changes in methylation. results: forty-four participants achieved successful desensitization, defined as passing dbpcfcs to or more foods following oit. we found that the - cpg site in the il- promoter region is hypermethylated over time during successful multi-food oit (fdr-adjusted p < . by wilcoxon signed-rank test). the median % of methylation at baseline was . (interquartile range . %) and was . post oit (interquartile range . %). there were no statistically significant (with a significance level of fdr adjusted p value of . ) changes in the il- , foxp , or ifn-g loci in the cpg sites we studied. conclusions: these preliminary results suggest that one immune mechanism involved in successful desensitization may involve suppression of th function by hypermethylation of il- in immune cells in the peripheral blood. introduction/background: atopic dermatitis (ad) is a common chronic inflammatory skin disorder afflicting from infancy to adults with itching, scratching, and lichenification. objectives: we investigated the effects of esculetin from fraxinus rhynchophylla on atopic skin inflammation. methods: for induction of atopic skin inflammation, we exposed the ears of female balb/c mice with house dust mite (dermatophagoides farinae extract, dfe) and , -dinitrochlorobenzene (dncb) during weeks. results: oral administration of esculetin reduced dfe/dncbinduced atopic skin inflammation symptoms based on ears swelling and scratch numbers. the immunoglobulin (ig) e, igg a, and histamine levels in serum were decreased and inflammatory cell infiltration in skin tissue was reduced by the esculetin. it suppressed th , th and th responses by inhibiting the production of inflammatory cytokines such as tumor necrosis factor (tnf)-, interferon (ifn)-, interleukin (il)- , il- , il- and il- in the ear tissue. further, we investigated the effects of escueltin on activated keratinocytes, one of the most representative cells for studying pathogenesis of acute and chronic atopic skin inflammation. as results, esculetin suppressed gene expression of th , th and th cytokines and activation of nuclear factor-b and signal transducer and activator of transcription in tnf-/ifn-stimulated keratinocytes. conclusions: taken together, the results imply that esculetin attenuated atopic skin inflammation, suggesting that esculetin might be a potential therapeutic candidate for the treatment of ad. introduction/background: autoimmunity is often seen in common variable immune deficiency (cvid) with immune thrombocytopenic purpura (itp) being the most frequent manifestation at a prevalence of - % in cvid patients. in such patients, itp is often recognized and treated long before cvid, the implications of which are unknown. primary itp is a clinicopathologic diagnosis which includes an evaluation of other conditions that may mimic it including cvid. currently, it is unknown how frequently cvid is evaluated during the diagnostic workup of itp and what percentage of those patient actually have cvid. objectives: the two main objectives of this study were to determine the number of itp patients that had an igg level checked during their clinical course and if the globulin fraction can be used as a marker for hypogammaglobulinemia in itp patients at the time of diagnosis. methods: a retrospective chart review was undertaken at a large academic medical center of patients with a new diagnosis of itp between january and january . igg levels were collected and globulin fractions were calculated as the difference between serum total protein and albumin within days of the initial itp diagnosis. results: six hundred and twenty-three patients were found to have a new diagnosis of itp in the given timeframe. of these, only ( . %) had igg levels checked at any point during their clinical course. twelve of the ( . %) had hypogammaglobulinemia with only of the ( %) having a formal immunologic follow-up evaluation. two were diagnosed with a primary immunodeficiency ( cvid and ctla deficiency). globulin fractions were calculated on patients at the time of itp diagnosis. mean calculated globulin fraction in hypogammaglobulinemic patients was . (range . . ), versus . (range . . ) in patients without hypogammaglobulinemia (p= . ). conclusions: the diagnosis of cvid is often delayed from the onset of symptoms which can include autoimmune conditions such as itp. our data indicate that clinicians do not routinely check igg levels at the time of itp diagnosis which should be considered standard of care based on the current guidelines. our data suggest that although calculated globulin fractions were significantly lower in hypogammaglobulinemic patients, the variability was substantial and hypogammaglobulinemic patients would be missed using this as an indicator of low immunoglobulins. future directives include a prospective study using igg levels checked at the time of itp diagnosis, with formal evaluation for cvid in any hypogammaglobulinemic patients to evaluate the true prevalence of cvid among itp patients. introduction/background: autoimmune lymphoproliferative syndrome (alps) is a disorder characterized by immune dysregulation due to the rupture of lymphocyte homeostasis, which occurs as a result of mutations in the apoptotic pathway mediated by fas. this disease is sometimes misdiagnosed due to its variable phenotypic expression and the overlapping of symptoms with many other hematological and immunological disorders. ( ) a patient diagnosed with evans-fisher syndrome (sef) was referred to the laboratory for immunity studies. this is a patient who from infancy had multiple admissions for severe sepsis, with severe anemia and severe thrombocytopenia. in the evaluation of the history of the disease by the work team, a series of clinical data was observed that suggested the possibility of the patient presenting an alps, so it was decided to incorporate as part of the study, the quantification of the cells t cd + cd -cd -(double negative t cells or dnt) that express tcr +. the differentiation pathways of dnt tcr + and the role of fas in this process are not clear, some authors hypothesize that they may be represented by direct descendants of chronically activated positive simple t cells, with deregulated co-receptors, in a state of differentiation in which they were destined to perish by fas-mediated apoptosis. ( ) ( ) ( ) ( ) the population of tcr + dnt cells required for diagnosis must be derived from the particular study of each laboratory in their populations, but several working groups agree that the pathological limit values are , % in total lymphocytes or , % in cd + lymphocytes. ( ) evidence shows that dnt alps cells are not simply accumulated in senescent withdrawal; they and their precursors remain active and proliferate under the influence of activation signals. ( ) although numerous genetic deficiencies lead to lymphoproliferation of t cells, only that caused by a defective fas pathway is dominated by negative double t cells. ( ) in clinical immunology, the distinction between cd ra + and cd ro + cells is particularly useful for determining the state of the "naive" cell compartment in relation to its thymic origin. primary immunodeficiency disorders are characterized by a decrease / absence of thymus performance, and often involve a decrease in cd ra + t lymphocytes. scientific evidence suggests the important role of the "naive" subpopulation in the origin and maintenance of self-reactive effectors in the periphery. ( ) regulatory t cells are able to effectively control autoreactive t cells, especially when negative thymic selection is defective. its differentiation and function is controlled by foxp . the decrease or absence of regulatory t cells leads to autoimmune diseases, specifically those mediated by cd + t cells, and to lymphoproliferation characterized by multiorgan inflammation and other autoimmune disorders. ( , ) objectives general: establish the diagnosis of a pediatric patient with suspected primary immunodeficiency due to dysregulation, an autoimmune lymphoproliferative syndrome (alps). specific: evaluate the tcr + dnt population, relevant in the diagnosis of alps. quantify cell populations that exhibit markers of activation, differentiation and regulation of the relevant t cells in the study of this disease. methods: monoclonal antibodies (mab) cd fitc / cd pe / cd pc triple labeled (beckman coulter), tcrapc, cd pe, cd rapercp, cd rofitc, cd fitc, cd rope, cd apc, cd f, cd pe, cd percp, cd pe, cd pe, cd percp, cd fitc, cd fitc were used , hla dr and the cd fitc / cd pe / foxp apc regulatory t cell kit, all from miltenyi biotec. the trial included the use of a healthy control. both samples were processed in unison to ensure reproducibility. the samples were acquired in a beckman coulter gallios cytometer, with the use of the kaluza program, version . . the analysis strategy included the formation of overlapping "gate" windows for the quantification of dnt tcr + and regulatory t cells. results: the patient studied showed . % of tcr + negative double t cells, in relation to . % of the healthy control. the presence of this population of t cells in patients diagnosed with evans syndrome, even in the absence of lymphoproliferation, is consistent with alps. ( ) (graph ) the presence of increased tcr + dnt and lymphoproliferationexpressed as lymphadenopathies and splenomegaly of noninfectious or malignant cause, of at least months of evolution; plus the typical immunohistological findings found in the patient's lymph node biopsy (paracortical hyperplasia), the presence of autoimmune cytopenias (hemolytic anemia and thrombocytopenia) and hypergammaglubulinemia, led to the "probable diagnosis" of alps in the patient studied. the accumulation of dnt in the lymph nodes and other peripheral organs is accompanied by qualitative changes in the composition of the t cell repertoire, so that the immunophenotype performed included markers of activation, differentiation and regulation. the evaluation of the activation on t lymphocytes and nk cells showed that there was a slight decrease in the expression of the receptor for il- , cd . the cd + cd + population was expressed in greater percentalthough discretely-in the patient than in the healthy control, which was directly related to the presence of hypergammaglobulinemia, elevated iga and the presence of autoantibodies. it was also highlighted by the high expression of the cd + cd + autoreactive population in the patient studied ( . % vs . %). in order to know the impact on cell differentiation, the subpopulations of effector and memory cells for cd + and cd + t lymphocytes were evaluated by combining the cd ra and cd ro isoforms. cd ra is expressed in naive t lymphocytes. particularly, the cd + cd ra + population has an essential function as a suppression inducer, and it is diminished in the patient studied, in relation to the control. ( . % vs. . %, respectively) the same behavior was shown by the cd + cd ro + memory and effector cells, . % vs . %. in the cd + population itself, it was possible to confirm the presence of a clone that expressed both receptors (cd ra and cd ro), probably a temra population (terminally differentiated effector memory cells). when comparing the results, it was unexpectedly found that there was a slight decrease between patient and control ( . % vs . %). as the characteristic phenotype of this cell population could not be corroborated, conclusive assessments could not be made. the combination of cd isoforms was also used to study cd + t lymphocytes, but similar behavior between patient and healthy control was observed. (the lab results are shown in table ) the analysis of the regulation of the immune response showed a decrease in the cd + cd + population and the expression of the foxp transcription factor in the patient with respect to the control. ( . % vs. . %, . % vs. . %, respectively) (graph ) conclusions . the markedly high quantification of cd -cd -tcr + t lymphocytes allowed to define the "probable diagnosis" of autoimmune lymphoproliferative syndrome in the patient under study. . the increased activation of cd + cells and the presence of the cd + cd + self-reactive population, largely responsible for autoimmunity and lymphoproliferation in the alps, could be confirmed. . the decrease in the cd + cd ra + t-cell suppressor-inducing population evidenced corroborated its involvement in this disorder by primary immunodeficiency, in relation to the thymus dysfunction that originates and maintains self-reactive effectors in the periphery. . the decrease in the expression of the foxp transcription factor observed, points towards a low regulation of the response that leads to autoimmunity and to lymphoproliferation, specifically mediated by cd + t cells. allergy and immunology arnp, nicklaus children's hospital allergy and immunology division director, nicklaus children's hospital introduction/background: specific antibody deficiency syndrome is characterized by a weak antibody response to bacterial polysaccharide antigens when no other immune system abnormalities can be found. low titers to pneumococcal vaccine have become one of the most frequently recognized immune abnormalities in pediatric patients with recurrent sinopulmonary infections. nonetheless, insufficient data and lack of consistent testing of the response to pneumococcal polysaccharides continues to affect the optimal diagnosis and management of this specific antibody deficiency. objectives: to characterize the pre-and post-immunization igg antibody trend for each specific serotype included in the pneumococcal -valent conjugate vaccine (pcv ), as well as others that are routinely tested, in a cohort of pediatric patients with recurrent sinopulmonary infections. secondarily, to understand differences in the immune response to the vaccine booster between age groups. methods: this retrospective review identified patients with recurrent sinopulmonary infections. in this cohort, required an immune workup, and were found to have low pneumococcal titers needing a pcv vaccine booster. baseline pneumococcal serotype-specific antibody titers at initial visit and weeks after the vaccine booster were obtained. patients were categorized by age: years, - , - , and - . an adequate response to the pneumococcal conjugate vaccine was deemed to be a -fold increase over baseline and/or a post-immunization titer of . μg/ml or greater. results: overall, pcv booster provided a significant improvement in the number of protective titers, increasing from . ( % ci: . - . ) serotypes at baseline to . ( % ci: . - . ) serotypes at weeks (p < . ). this increase correlated with improved clinical outcomes % showed no signs of recurrent infection after the first booster and % after a second dose. all those who did not improve clinically suffered from co-morbidities (genetic abnormalities and rheumatologic diseases). post-immunization antibody concentrations were significantly higher than at baseline for all serotypes (p < . ) and only , n, and f did not exhibit a greater than -fold increase (p > . ) weeks following the booster. across age groups, only , f, and v showed pre-immunization differences in titers. there were no differences between ages in post-immunization titer levels for all serotypes. similarly, all age groups had a comparable number of baseline titers (p = . ) and at follow-up (p = . ). conclusions: in pediatric patients with recurrent sinopulmonary infections, an additional pneumococcal booster proved to be effective in the protection of these children from further infections. the pcv booster substantially increased titer levels and concentrations, and significantly improved clinical outcomes, independent of age. this investigation has provided us with a better understanding of the response after booster vaccination, and its role in the protection of patients from recurrent sinopulmonary infections. further studies are needed to elucidate whether a fifth dose of pcv should be optional as part of the vaccination schedule of patients with recurrent sinopulmonary infections. introduction/background: many primary immunodeficiencies (pids) share overlapping presentations, complicating clinical diagnosis. due to the ability to include many genes in one assay and the rapid turnaround time that these panels allow, expanded next-generation sequencing (ngs) panels are valuable in facilitating the diagnosis of patients with pids. objectives: we aimed to determine the clinical utility of an expanded ngs panel for the genetic diagnosis of patients with suspected pids. methods: we performed a retrospective analysis of the clinical utility of a -gene pid ngs panel used in a clinical diagnostic laboratory. from april to october , panels were ordered for patients with suspected or known pids. results: seventy-four pathogenic or likely pathogenic (p/lp) variants were identified in ( . %) patients. eight ( . %) of the p/lp variants were copy number variations ( deletions, duplications). fifty-two patients ( %) had p/lp variant, and patients ( . %) had p/lp variants. of positive patients, . % (n = ) were heterozygous carriers for autosomal recessive conditions where a second variant was not identified. twenty-two patients had p/lp heterozygous variants in genes with autosomal recessive and autosomal dominant inheritance patterns, in which the positive findings may or may not explain the patient's phenotype. for example, variants in this category were heterozygous variants in tnfrsf b (taci). genetic diagnoses were established or likely in % of patients with p/lp variants ( . % of all patients). five patients were heterozygous for a single p/lp variant and a variant of unknown significance in the same autosomal recessive gene. four patients in whom a genetic diagnosis was determined were also heterozygous carriers for a second, unrelated condition. one patient was found to have two distinct genetic diagnoses. variants of uncertain significance (vus) were identified in most ( . %) patients. the average turnaround time from test requisition to return of results was days. in total, % percent of genetic diagnoses were for conditions that are treatable with hematopoietic cell transplantation. conclusions: these results illustrate the utility of broad ngs panels for the diagnosis of patients with pids. introduction/background: severe combined immunodeficiency (scid) is a life-threatening immune deficiency manifest by extreme susceptibility to infection. early diagnosis and definitive treatment with either hematopoietic cell transplant (hct) or, in select cases, gene therapy (gt) has been shown to significantly improve survival. newborn screening for scid has allowed for the opportunity to promptly identify these patients before significant infections occur. at ucsf, newly diagnosed infants with scid are admitted to the hospital for management and remain in isolation for definitive treatment and until adequate immune reconstitution occurs. previous work by our group demonstrated up to % of these parents experience psychosocial trauma manifested by depression and post-traumatic stress disorder (ptsd). the psychosocial challenges that contribute to depression and ptsd in these parents have not been qualitatively described. objectives: to understand the range of experiences and feelings of parents/caregivers of infants with scid diagnosed by newborn screening throughout their prolonged hospitalization and isolation in order to better support patients, parents, and their families. methods: voluntary participation was elicited from parents of children with scid who were status post hct/gt for one year or longer. semi-structured, in-person interviews lasting approximately - minutes were conducted with parents; interviews were recorded and transcribed. parents were asked to discuss their experiences from first notification of an abnormal screening result through discharge after hct or gt. emerging themes were identified from the transcribed interviews. results: we interviewed mothers and fathers of infants with scid. six infants received hct, while one underwent gt for ada-scid. all children were alive and well at the times when interviews were conducted. overall, once admitted, parents reported feeling well supported by the medical team and support staff. however parents identified a number of stressful events. uniformly reported key stressors included: receiving the first phone call regarding their childs abnormal newborn screening results, preparing for hct, coping with prolonged isolation, and transitioning from hospital isolation to care with ongoing isolation at home. other challenges described reflected the additional stressors of caring for a newborn, including coping with postpartum depression. overall, we identified three major themes encompassing the challenges faced by parents of hospitalized scid patients: (i) loss of normalcy and control over multiple aspects of life; (ii) prolonged waiting periods (especially the wait between diagnosis and hct and between hct and evidence of t cell engraftment); and (iii) perceived lack of guidance on realistic expectations during the hospital stay . parents sought and relied on peer support from other scid parents to learn about coping with the nuances of daily life as a parent of an infant with scid. conclusions: we identified multiple psychosocial stressors and challenges uniquely faced by parents and caregivers of infants diagnosed with scid by nbs. parents described barriers to caring for their own physical and mental health, which can be especially harmful to parents experiencing postpartum depression. recognizing these challenges allowed for the identification of opportunities to improve both healthcare delivery to their children and institutional support for future families affected by scid (table i) . emphasis should be placed on providing parents with scid-specific resources as early as the time of diagnosis, connecting parents with scid support networks, and facilitating access to psychosocial and mental health services for caregivers introduction/background: chronic granulomatous disease (cgd) is caused by a genetic defect that impairs phagocyte function. this disease results in recurrent infections and granuloma formation. rarely do patients develop cutaneous symptoms, unless associated with autoimmune disorders such as systemic erythematous lupus. previously described cutaneous findings include granulomas, abscesses, photosensitivity, malar rash, discoid lupus, vasculitis, and rarely vesicular rashes. here we describe two term infants diagnosed with x-linked cgd who present, in addition to frequent infection, with a unique papulopustular skin rash initially diagnosed as non-classic appearing eczema refractory to usual eczema treatment and antibiotics. objectives: to characterize cutaneous findings in x-linked cgd and emphasize the importance of considering further work in patients who present with similar rashes in conjunction with concerning features for primary immunodeficiency. methods: each infant was diagnosed with cgd based on abnormal dhr testing and/or genetic evaluation. after obtaining consent from both families, we have documented photographs of the development of rash in two newly diagnosed infants with cgd. one infant underwent cutaneous biopsy with resulting pathologic evaluation. results: our first patient presented at months of age with episodic fever with chronic leukocytosis, iron deficiency anemia, thrombocytosis, elevated inflammatory markers, proctocolitis with elevated calprotectin, and rash. egd with flexible sigmoidoscopy showed no signs of inflammatory bowel disease, the left colon had macroscopically raised erythematous lesions but was microscopically normal with no signs of active colitis. he was initially diagnosed with fpies and eczema in the setting of diagnosis of various infections (otitis, upper respiratory illness). the skin rash was described as non-pruritic, generalized pink-purple papulopustular lesions with a pink base, most prominent on upper and lower extremities, mostly sparing the trunk. skin biopsy histopathology revealed an essentially unremarkable appearing epidermis and within the dermis, there was a superficial perivascular lymphohistiocytic infiltrate. eosinophils and plasma cells were not abundant. the histologic changes were thought to be non-specific, but could be seen in a drug reaction or urticaria. a viral exanthem could also demonstrate these changes. giemsa stain and a stain for mast cell tryptase revealed normal numbers of mast cells in the biopsy specimen. he was not on any oral medications at the time and did not respond to treatment with topical moisturizers, oral antihistamines, topical steroids, or any standard for eczema care. he had a maternal uncle who passed away in infancy from infection. heightened clinical suspicion for primary immunodeficiency led to obtaining a neutrophil respiratory burst assay which was consistent with cgd and genetic testing was positive for x-linked cgd. pathogenic mutation nucleic acid change was c. c>t; hemizygous; amino acid alteration was p. arg>stp within the cybb gene locus. he had no other features of auto-immune disease including negative ana obtained later in his clinical course and his colitis diagnosed as cgd-associated colitis. his rash did not respond to systemic treatments for his colitis, oral antibiotics, and during hsct. our second patient presented at months of age with persistent fevers, inguinal lymphadenopathy, leukocytosis, elevated inflammatory markers, non-bloody diarrhea and rash. there was a family history of autoimmune gi disease. he presented to the hospital with fever of unknown origin with concern for infection, atypical kawasakis, and drug rash. his rash was described as a blotchy, pink, papular rash most prominent on the upper arms but also present on lower arms, legs, and chest, faint on face. there is some induration and thickness to the rash in some areas (confluent on the arms with more erythema and induration) and more faint pink papules (scattered on legs) elsewhere. a biopsy of his inguinal lymph node showed granulomatous lymphadenitis with neutrophilic abscess formation, and culture was positive for serratia marcescens. neutrophil respiratory burst assay was consistent with cgd and genetic testing was positive for x-linked cgd, mutation with the cybb gene. pathogenic nucleic acid change was c. + g>a; hemizygous; amino acid alteration was p. deletion of exon . the rash remained unchanged with treatment for infection, initiation of antifungal and bacterial prophylaxis, along with topical steroid therapy. conclusions: in patients who present with frequent infections and who have unusual cutaneous findings that do not fit with common infant rashes, consideration of work up for cgd should be pursued. specifically, generalized papulopustular rash with a negative skin biopsy can be misdiagnosed as atopic dermatitis, and in the right clinical context cgd should be considered. introduction/background: chronic lung disease in common variable immunodeficiency (cvid) is heterogeneous, and it is a leading cause of morbidity and mortality in this population. currently, the diagnosis and monitoring of chronic lung disease in cvid rely on radiographic findings, biopsy and/or pulmonary function tests. fractional exhaled nitric oxide (feno) is a noninvasive biomarker of airway inflammation, and it has been widely utilized to aid the in-office diagnosis, characterization, and management of inflammatory airway disease, such as asthma. however, exhaled nitric oxide in cvid patients with chronic pulmonary complications has not been examined. objectives: we aimed to determine fractional exhaled nitric oxide levels in cvid patients. methods: we measured exhaled nitric oxide in cvid patients with or without chronic lung disease. results: feno measurements were obtained in cvid patients (mean age: , range - ). five patients had no known lung disease; patients had chronic lung disease (bronchiectasis, n= ; lung granulomas, n= ; hepatopulmonary syndrome, n= ). four patients were on inhaled steroid (bronchiectasis, n= ; lung granulomas, n= ), patients were on systemic steroid (lung granulomas, n= ; hepatopulmonary syndrome, n= ), and patient was on oral budesonide (no known lung disease). feno was elevated (> parts per billion [ppb]) in of patients, of whom had known chronic lung disease. patients with granulomatous lung disease had higher feno (mean . ppb, range - ppb) compared to patients without known lung disease (mean ppb, range - ppb, p= . ), patients with bronchiectasis (mean . ppb, range - , p= . ), and the patient with hepatopulmonary syndrome ( ppb). feno levels remained elevated in granulomatous lung disease in patients with inhaled or systemic steroid use. conclusions: this is the first report of feno measurements in cvid patients. feno is elevated in a subgroup of cvid patients, and it may differ according to the underlying lung pathology. further investigation is warranted to determine the utility of feno in the diagnosis and management of chronic lung disease in cvid. professor of pediatrics, university of british columbia introduction/background: whole exome sequencing (wes) has revolutionized the discovery, diagnosis, and treatment of primary immune deficiency diseases (pids), a group of disorders with high genetic and phenotypic heterogeneity. current bioinformatic analysis approaches for wes rely heavily on population databases to exclude variants present in the populations represented by these databases. this approach may confound the ability to detect true disease causing variants, and conversely, flag variants that are absent from population databases only by virtue of originating from minority populations. ultimately, the pathogenicity of novel variants can only be mechanistically established through rigorous biochemical and functional validation. objectives: to evaluate the pathogenicity of a novel homozygous variant in caspase recruitment domain family member (card ) (c. g>t, p.c y), in a patient with features of combined immunodeficiency (cid). methods: research study protocols were approved by our institutional review board. six members of one family (the affected child, healthy siblings and their parents) were enrolled. written informed consent for genetic testing and participation was provided by the parents for their children. genetic, bioinformatic, biochemical and immunological investigations were performed. results: targeted sanger sequencing confirmed the presence of the homozygous card variant c. g>t, p.c y in the index patient (and subsequently in one apparently healthy sister). each parent was found to be a heterozygous carrier of the variant. nfkb activation in vitro was found to be normal for both the index patient and sister, and was indistinguishable from unaffected family members. conclusions: although multiple in silico tools predicted the c. g>t, p.c y card variant to be pathogenic, the patients b and t cells did not have aberrant nfkb activation in vitro, as would be predicted given the central role of card in nfkb activation. this practical experience highlights how imperative it is to functionally characterize novel variants found by wes, even when variants are predicted to be damaging. chief, clinical immunology -faculdade de medicina do abc introduction/background: gata is a zinc finger transcription factor essential for embryonic and definitive hematopoiesis. heterozygous mutations leading to gata deficiency were first described in . the age of clinical presentation ranges from early childhood to late adulthood, with most of them in adolescence to early adulthood. patients present clinical findings as monocytopenia, nontuberculous mycobacterial infections, myelodisplasia, viral (mainly hpv and ebv), fungal and bacterial infections. patiens may arise with many phenopytes, showing how complex is the effect of this transcription factor. objectives: we report a patient with gata mutation. methods: report: a year old female patient was referred due to a mycobacterial non-tuberculosis pneumonia. she was a healthy child until puberty. at the age of she presented genital herpes and recalcitrant vulvo-vaginal warts (hpv). at , she complained of lower back pain with no diagnosis for months, medication was unnefective, and she developed ischemic stroke. hemiparesia was mild and temporary. endocarditis with no agent identified was also diagnosed. therapy with acetilsalicilic acid was mantained until the age of . five years later, vaginal hpv spread through vulve and anal region evolving to neoplasia. she was submitted to surgery, radio and chemotherapy until october . one year later, profound cytopenias led to hospitalization. during that time, she had cough and fatigue and pulmonary tuberculosis was diagnosed. despite therapy with rifampin, isoniazid and pyrazinamid, there was no improvement. bronchoalveolar lavage was positive for mycobacterium avium. main immunological evaluation showed: monocytes ( %), t cells /mm ; cd +: ( , %), cd +: ( %); b cells /mm ( , %); nk: /mm ( , %); normal immunoglobulin levels; incomplete response to pneumococcus serotypes; igm and igg positive for cmv; negative serologies for hiv, ebv and htlv / . molecular analysis identified an heterozygous mutation c. c>tp.(thr met) in gene gata . results: we report a patient with gata mutation. the same gata mutation was previously described in national institute of health nih-usa (n= ) and in australia (n= ). conclusions: it took almost years for diagnosis suspicion. gynecologists should be warned about this diagnosis in order to improve patients prognosis. early diagnosis is crucial with adequate prophylaxis, prompt treatment of infection, surveillance of malignancy, and, moreover, family screening and genetic counseling. this is the first report of this mutation in latin america. introduction/background: severe combined immunodeficiency (scid) due to adenosine deaminase (ada) deficiency was the first human monogenic disease to be approached with gene therapy, and ongoing research advances over years led to the approval by the european medicines agency of a stem cell gene therapy product for its treatment using a gammaretroviral vector (gv), strimvelis®. despite the high success rate using gvs for ada gene transfer without vector-related complications, the development of leukoproliferative complications from the use of gvs in gene therapy of other disorders led us to develop lentiviral vectors (lvs) to deliver the corrective ada sequence/cdna (corrigan-curay et al, mol ther ; modlich et al, mol ther .). lvs, typically derived from hiv- and devoid of all viral genes, can be produced in a self-inactivating (sin) configuration, in which the viral long-terminal repeat enhancers are absent, eliminating the major identified cause of insertional oncogenesis due to gvs. we developed a lv (efs-ada) that carries a normal human ada cdna (codon-optimized) and demonstrated in pre-clinical studies its efficacy to transfer and express the ada protein, with evidence of significantly decreased potential for insertional mutagenesis compared to gammaretroviral vectors using the immortalization (ivim) assay and in murine bone marrow transplant models (carbonaro et al, mol ther, ) . this new lv was evaluated for safety and efficacy in parallel clinical trials of gene therapy for ada scid performed at sites in the u.s (university of california, los angeles and the national institutes of health {nih} clinical center) and u.k. (university college london/great ormond street hospital) enrolling subjects between and . we report here results from the patients treated in the u.s. between - . objectives: this is a prospective, non-randomized phase i/ii clinical study to assess the safety and efficacy of efs-ada lentiviral vector stem cell gene therapy in ada-scid subjects older than month. methods: subjects with ada-scid were enrolled, screened to document eligibility and underwent bone marrow harvest ( - cc/kg). bone marrow was processed to isolate cd + cells, which were pre-stimulated by overnight culture in serum-free medium containing c-kit ligand, flt- ligand, and tpo followed by culture with the efs-ada lentiviral vector overnight. subjects received a single dose of busulfan ( mg/kg) iv for reduced intensity conditioning. cells were removed from culture, washed, formulated and administered by iv infusion at least hours after busulfan administration. enzyme replacement therapy (ert) with pegylated bovine ada (peg-ada) was continued for one month post-transplant and then stopped. subjects were followed over months to assess safety and efficacy end-points. results: with - months follow-up, overall survival is %. eventfree survival has also been %, as all subjects are alive, remain off peg-ada ert, and none have required a second transplant. successful engraftment of gene-corrected cells was observed in all subjects at months, and persisted over the months of observation, based on vector gene marking in granulocytes and peripheral blood mononuclear cells (pbmcs), changes from baseline in rbc ada enzyme activity, and levels of metabolic detoxification of deoxyadenosine nucleotides. immune reconstitution was observed in all subjects and was sustained over the two years of observation, based on improvement of peripheral blood absolute lymphocyte counts and lymphocyte subsets (t, b and nk cells, and naïve cd + t cells). eighteen of patients have been able to stop receiving immunoglobulin replacement therapy. subjects who had routine infections all recovered with standard of care treatment, and there were no severe or opportunistic infections. conclusions: conclusions: gene therapy using the efs-ada lv has a favorable safety profile and was efficacious in this trial. a current followon trial at ucla is using a cryopreserved formulation of the cell product and pharmacokinetic-adjusted busulfan dosing, sponsored by the california institute for regenerative medicine (clin - ) and orchard therapeutics. acknowledgements: this study was supported by research grants from the national institutes of health (u ai ; p hl - ; nhlbi gtrp rsas # and ) and the california institute for regenerative medicine (cl - ; fa - ); support from the ucla david geffen school of medicine human gene and cell therapy program and the ucla eli & edythe broad center of regenerative medicine and stem cell research; and funding from the nhgri intramural program. dbk has potential financial conflict of interest as a member of the scientific advisory board for orchard therapeutics and as an inventor on intellectual property which ucla has licensed to orchard therapeutics related to gene therapy for ada scid. introduction/background: x-linked severe combined immune deficiency (scid-xl) is a rare monogenic primary immunodeficiency disorder (pid) where male infants are born without an adaptive and innate immune system. it is a life-threatening disease due to patients inability to fight viral and bacterial infections. scid-xl is driven by any of the two hundred known pathogenic mutations in the interleukin gamma receptor (il rg) gene, which function is required for proper development of t, b and nk cells. the most effective treatment for scid-xl, if performed in the first few months of life, is allogeneic hematopoietic stem cell transplantation (allo-hsct). this treatment is limited by absence of match donors, incomplete immune reconstitution, graft versus host disease and the need for long-term immunosuppression. an alternative curative treatment for scid-xl would be genome editing-based gene therapy by ex vivo genome correction of the patients long-term hematopoietic stem cells (lt-hscs) prior to autologous stem cell transplantation (auto-sct). objectives: here we report a proof-of-concept genome editing-based approach for correcting scid-xl disease. methods: using a crispr/cas -raav platform, we deliver a fulllength codon optimized il rg complementary dna (cdna) at the endogenous start site in cd + hematopoietic stem and progenitor cells (hspcs). results: using an optimized genome editing protocol we achieved > % genome editing as early as h and a median of % genome targeting while retaining > % viability and promoting greater than % ex vivo expansion of cd + hspcs from healthy donors. we demonstrate that our approach retains proper il rg signaling function in t-cells derived from healthy male donors and rescues the lymphopoietic defect from a patients derived mobilized cd + hspcs both in vitro and in vivo. we further show robust in vivo primary human engraftment potential and multi-lineage hematopoietic reconstitution of il rg gene targeted hspcs: a median of % (bone marrow), % (spleen) and % (liver) of il rg genome targeted engrafted cells was achieved at four months following primary transplantation into nsg mice and a median of . % - % was detected at months from secondary transplants of il rg targeted hspcs, thus achieving clinical levels of editing of lt-hscs. lastly, our observation of ( ) an intact hematopoiesis derived from il rg targeted cd + hspcs combined with ( ) a normal karyotype analysis and ( ) our deep analysis of potential off-target activity that showed only off-target sites of no known functional significant with < . % frequency of off-target activity presents strong evidence for the safety of our genome editing approach. conclusions: in sum, this pre-clinical study provides specificity, toxicity and efficacy data supportive of continued development of genome editing to treat scid-xl. objectives: to determine the risk of gvhd in msd hsct for scid patients compared to matched related donor (mrd). methods: retrospective cohort study comparing msd with mrd and the outcome of gvhd in all scid patients who underwent hsct between and . all statistical analysis was done using ibm spss statistic software. results: scid patients underwent hsct, ( %) received gvhd prophylaxis. gvhd occurred in ( . %); / ( %) had gvhd prophylaxis compared to / ( %) that did not, p value = . . acute gvhd occurred at a higher rate in msd / ( . %) compared to mrd / ( . %) p value = . . we analyzed outcome also according to period of hsct. first periods was to ; hsct, msd: , mrd: ; all had gvhd prophylaxis and there was no difference in gvhd. the second period was to : hsct, had msd and had mrd, gvhd prophylaxis was used in . % in msd and % in mrd, p value = . . gvhd was significantly higher in the msd ( . %) compared to mrd ( . %) odds ratio of . ( ci . to . ) p value = . . conclusions: gvhd prophylaxis in msd transplant may have a role to be considered in scid patients. introduction/background: xiap deficiency (also known as x-linked lymphoproliferative disease type xlp ; mim: ) is an x-linked primary immunodeficiency associated with mutations in the gene encoding the x-linked inhibitor of apoptosis (xiap; mim: ). the pathophysiology is characterized by immune dysregulation, usually triggered by epstein-barr virus (ebv) infection. primary ebv infection is followed by hemophagocytic lymphohistiocytosis (hlh) with high grade persistent fever, splenomegaly, hematologic cytopenias and hepatitis. most patients die during this acute phase, and those who survive usually evolve with hypogammaglobulinemia, recurrent infections, cytopenias, inflammatory bowel disease (ibd) and low counts of inkt cells. dysbiosis of intestinal microbiota is believed to fuel ibd and possibly contribute to the initiation and/or perpetuation of the disease. experimental studies have provided solid evidence to support a role for the indigenous gut microbiota in the pathogenesis of autoimmune diseases, thereby raising the possibility that an altered gut microbiota is an environmental risk factor for xiap disease. objectives: in this study, we sought to investigate the identity and abundance of the bacteria in gut microbial communities in a years old male patient with xiap deficiency. case presentation: at years of age, the patient presented with positive serology of active ebv infection, hlh, severe hepatitis, encephalitis and myocarditis. after recovery, the patient evolved well with few manifestations for several years. approximately years ago, the patient showed slow progression of hypogammaglobulinemia predisposing him to infections of the upper and lower respiratory system that required intravenous immunoglobulin replacement. immunological evaluation revealed reduction (but not absence) of inkt cells. at that time, the patient presented with intermittent diarrhea and abdominal pain that became more frequent and severe. after evaluation as ibd, the patients had been treated but without much improvement of diarrhea or resolution of his pain. after approximately months, the patient presented noted pain and fistulas lesions at the scrotal and gluteal regions. the exact causes of ibd in xiap deficiency are not known, but the abnormal activation of the mucosal immune system due to exaggerated response to the commensal bacteria associated to the dysregulation of nod and nod signaling might play an important role in the development and maintenance of the inflammatory status. methods: the gut bacterial composition was assessed by targeted metagenomic from the patients stool sample, collected before initiation of ibd antibiotics therapy. the s rrna amplified and its sequences were analyzed using a bioinformatic pipeline based on mothur software. we determined the bacterial community composition using , filtered reads using illumina miseq platform. results: according to the ezbiocloud database, the obtained dataset included operational taxonomic units at % dissimilarity, distributed among the following groups: bacteroidetes ( . %) with . % of b. dorei, , % b. vulgates, and . % b. fragilis, firmicutes ( . %), proteobacteria ( . %), bacteria_uc ( . %), actinobacteria ( . %). conclusions: increased abundance of bacteroides species including b. dorei and b. vulgatus have been implicated in inflammation in several gut diseases such as ulcerative colitis, irritable bowel disease, and celiac disease. although this experience is limited to a single patient, the results of the present study suggest an association between altered gut microbiota and the pathogenesis of ibd in xiap disease and may be of relevance to the future development of novel therapeutic strategies for xiap deficiency. ( ) submission id# hematopoietic stem cell transplantation in patients with primary immune regulatory disorders: a primary immune deficiency treatment consortium (pidtc) and inborn errors working party (iewp) study introduction/background: primary immune regulatory disorder (pird) is a newly recognized group of immune-mediated diseases with prominent features of autoimmunity, autoinflammation, and non-malignant lymphoproliferation in addition to immunodeficiency. the clinical manifestations of pirds are frequently difficult to manage and hematopoietic cell transplantation (hct) can be considered as a treatment option, often in those with the most severe disease. we sought to aggregate data from patients who have undergone hct for genetically defined pird or features of immune dysregulation. objectives: we sought to aggregate data from patients with pird who have undergone hct in order to determine the quantity of patients, clinical manifestations, indication and hct, and overall outcome. methods: a questionnaire based survey was sent to all primary immunodeficiency treatment consortium sites and hct referral centers in europe to determine the quantity and characteristics of patients with pirds who have undergone hct. the survey captured clinical manifestations, timing and indication of hct, strategy of hct, and outcomes from - . results: patients from centers ( in north america and in europe) were included with either known genetic defects or considered to have immune dysregulation regardless of gene defect. known genetic defects were identified in subjects, while had symptoms of immune dysregulation but lacked a genetic diagnosis. the mean age of onset of disease was years (range - years). clinical manifestations included gastrointestinal disorders ( %), failure to thrive ( %), dermatitis ( %), hematologic cytopenias ( %), and lymphoproliferative disease ( %). recurrent infections ( %), immunodeficiency ( %), autoimmunity ( %), and autoinflammation ( %) were also common. organ specific autoinflammation occurred most commonly in the lung ( %) and brain ( %). the median age of hct was years ( - years). graft sources included matched unrelated donors ( %), matched related donors ( %), mismatched unrelated donors ( %) and haploidentical donors ( %). reduced or minimal intensity conditioning was used in % of transplants. five-year overall survival was % and the majority of survivors had resolution of symptoms that led to transplantation. among those patients that died, infection was the most common cause. conclusions: based on our survey data, pird patients commonly develop clinical features of autoimmunity, autoinflammation, and susceptibility to infection at a young age. hct can be successful and lead to disease resolution. however, further studies to define the appropriate patient, timing of hct, donor selection and pre-hct conditioning regimen are necessary to improve outcomes of patients with pird. introduction/background: pathogenic variants in tnfrsf b (taci) are relatively common (found in about % of the population) but have been seen in about - % of cvid patients, interestingly in both homozygous and heterozygous states. recent articles suggest that the heterozygous state increases the risk of developing autoimmunity due to an effect on autoreactive b cell selection and activation. objectives ) illustrate the importance of genetic testing in patients with difficulty to treat inflammatory disease ) present a case report of inflammatory bowel disease associated with a pathogenic mutation in the tnfrsf b gene results: yo wf was diagnosed with crohns disease due to the chronic abdominal pain, vomiting, and bloody stools since years of age. intestinal biopsy revealed inflammatory changes in the entire intestine with active, submucosal lymphoid hyperplasia, neutrophilic cryptitis with focal areas of crypathic damage, and submucosal epithelioid granulomas more predominantly seen in the colon and rectum. she was started on immunosuppressant medications but developed anaphylactic reaction to both infliximab and adalimumab. she was then treated with azathioprine, mesalamine, methotrexate, and oral budesonide. despite these medications, she continued to have frequent relapses, - episodes a year and required periodic systemic corticosteroid bursts. other biologics, vedolizumab and ustekinumab, were also tried without success, and she subsequently underwent a colectomy. her postoperative course was complicated by ards, poor abdominal wound healing, and sepsis. due to her complicated clinical course, immune work up was performed which revealed a normal cbc and lymphocyte subpopulations, but hypogammaglobulinemia with low isohemagglutinin titers and specific antibody levels. comprehensive genetic testing ruled out chronic granulomatous disease and other known primary immunodeficiencies but revealed a rare missense mutation in tnfrsf b (taci). this variant, c t (p.cys arg) (rs , exac . %) is likely pathogenic. this heterozygous variant has been seen in both cvid cases and unaffected relatives but significantly more common among cvid patients. moreover, the studies on b cells of these relatives showed impaired function. increased number of autoreactive b cells were also found in the bone marrow of heterozygous individuals and these cells could give a risk of developing autoimmunity. conclusions: in difficult-to-treat autoimmune diseases, identifying the underlying immune defect may aid in the treatment decision. in this case, b cell targeted treatment such as anti-cd monoclonal antibody could be beneficial. introduction/background: defects in immunoproteasome caused by biallelic or digenic loss-of-function mutations in proteasome catalytic subunits cause an autoinflammatory disease identified as chronic atypical neutrophilic dermatosis with lipodystrophy and elevated temperature (candle) associated to an increased interferon type i gene signature. the proteasome maturation protein (pomp) is a chaperone for both standard and immuno-proteasome assembly and is critical for the incorporation of catalytic subunits. here, we characterize and describe pomp-related autoinflammatory immunodeficiency disease (praid) in two unrelated patients and identify the underlying genetic mechanism of disease. objectives: determine the genetic cause and mechanism of disease in two patients with pomp variants . methods: whole-exome sequencing (wes) was performed to identify a genetic cause of our patients dysregulatory syndrome. proteasome assembly and catalytic function was assessed by sds-page and native gel respectively, using patient derived cell lines. expression of interferon type i-induced genes was measured by rt-qpcr. pomp protein was identified by western blot. results: we identified two unrelated individuals with a unique syndrome characterized by neonatal onset autoinflammation, neutrophilic dermatosis, autoimmunity, and combined immune deficiency with severe systemic viral and bacterial infections. immunologic evaluation for both individuals revealed elevated immunoglobulins, low cd + t cell numbers and extremely low b cell counts with persistently high titers of autoantibodies and increased expression of interferon type i-induced genes. in both individuals, truncating heterozygous de novo frameshift variants in pomp were identified by wes and confirmed by sanger sequencing. most mrna transcripts with premature termination codons should undergo nonsense-mediated decay (nmd), however in both of our patients, cdna sequencing revealed these transcripts escaped nmd. the expression wildtype and truncated versions of pomp protein was further confirmed by western blot. transfection of mutant constructs into an otherwise healthy cell line recapitulated an increased interferon signature suggesting a dominant negative mechanism. conclusions: we define praid in two unrelated individuals characterized by neonatal onset immune dysregulation and combined immunodeficiency caused by truncating variants in pomp in which transcripts that escape nmd result in a truncated protein that leads to a dominant negative (i.e antimorphic) allele. to our knowledge, praid is the first inherent defect of immunity mechanistically characterized by nmd escape. introduction/background: parvovirus viremia may occur in pediatric heart transplant patients who underwent thymectomy and developed secondary t cell lymphopenia. high dose intravenous immunoglobulin (hdivig) has been used to treat parvovirus infection in these cases. objectives: we aim to review different routes of immunoglobulin treatment in pediatric cardiac transplant patients with parvovirus viremia and compare the patients immunological phenotype. methods: data from three pediatric heart transplant patients with parvovirus viremia in a tertiary care center was reviewed including t cell counts, parvovirus viral load, route, dosage, and frequency of immunoglobulin treatment. results: all three patients received hdivig. patient and tolerated the treatment and viremia improved. patient developed recurrent aseptic meningitis from hdivig treatment and his viral load remained > million copies/ml. compared to the other two cases, patient had a much lower t cell count that likely contributed to the persistence of viremia. to improve his quality of life and reduce healthcare costs, a facilitated subcutaneous immunoglobulin (scig) treatment option was explored. scig treatment was well tolerated and led to a dramatic decrease in parvovirus viral loads in patient . conclusions: most pediatric cardiac transplant patients with persistent chronic parvovirus viremia respond well to hdivig, scig may serve as an alternative treatment option in refractory cases, especially in those with severe t cell lymphopenia. severe combined immunodeficiency (scid), by detecting tcell receptor excision circles (trecs) from dried blood spots (dbs) on routine newborn screening (nbs). this has lead to improved estimates of the incidence and prevalence of scid, decreased diagnostic delay, and improved patient outcomes. preliminary studies outside the us have demonstrated that nbs can be adapted to include screening for b-cell deficient infants before symptom onset by quantifying kappa-deleting recombination excision circles (krecs) on dbs. objectives: we report results of the initial characterization of a high throughput triplex trec/krec/rnasep assay run in , samples in new york state (nys). methods: dbs from anonymous, de-identified infants were included in the current study. dna from patients with confirmed primary immunodeficiencies (n= ), including, but not limited to, x-linked agammaglobulinemia (n= ) and scid(n= ) were obtained from the centers for disease control and prevention, and clinical immunologists working with the nbs programs in massachusetts, minnesota, wisconsin, and nys, were used as positive disease controls. all dbs were extracted and processed according to current nys nbs protocols. a trec/krec/rnasep triplex assay was designed and optimized to minimize reagent use, and maximize target amplification. cycle threshold (ct) was determined, and error detection cutoffs were identified to optimize sensitivity. results: pcr efficiency, assay quantification, intra-assay reproducibility, and error detection rates all met nys nbs standards. error detection rate for the triplex trec/krec/rnase p assay is %, comparable to the current error detection rate of % for the current duplex trec/rnase p assay. samples falling into the error detection range are repeated (analysis in process) to determine a receiver operating characteristic curve. conclusions: we show that the high-throughput trec/krec/rnase p triplex assay is feasible in a large, racially and ethnically diverse population in nys. compared to the current duplex assay, this assay has favorable performance characteristics and provides additional immunologic characterization. due to assay optimization, we were able to add the krec test at no additional cost. work is underway to further characterize other assay parameters such as sensitivity and specificity, in preparation for adoption of the triplex assay as part of routine nys nbs. highly accurate wiskott-aldrich syndrome diagnosis via rapid flowbased was protein staining samuel chiang , sue vergamini , ammar husami , marianne ifversen , kejian zhang , jack bleesing, md, phd , yenan bryceson , rebecca marsh, md introduction/background: wiskott aldrich syndrome (was) is a rare xlinked hemizygous disease commonly associated with symptoms of immune deficiency. diagnosis is based on clinical parameters including thrombocytopenia and reoccurring infections, but currently does not include any disease specific marker. as the only permanent treatment for was is hematopoietic stem cell transplantation, it is imperative that a swift and accurate diagnosis be made. absent or lowered was protein (wasp) levels have been reported in sporadic was cases. however, no systemic evaluation exists to date on the accuracy of wasp quantification for was diagnosis. objectives: to determine the accuracy of wasp staining in predicting was genetic abnormalities. methods: we retrospectively evaluated results from a rapid whole blood flow cytometry based assay on a cohort of suspected was patients and compared relative wasp staining levels to was genotype. roc curves as well as accuracy calculations were generated. results: a total of patients with normal and patients with a genetic abnormality in was were collected. missense mutations were most common but insertions, deletions, and gross mutations were also found ( fig a) . comparing was sequencing results to whole blood wasp expression levels provided an . % sensitivity and % specificity for a combined accuracy of . % when juxtaposed against genetic sequencing. when variants of unknown clinical significance (vucs) were removed, the sensitivity improved to . % (fig b) . conclusions: staining for wasp is a quick, simple, and accurate assay for the prediction of genetic was defects. introduction/background: cytotoxic t lymphocyte antigen (ctla ) is an inhibitory co-receptor essential for regulatory t cell (treg) function and a central regulator of t cell proliferation and expansion. ctla haploinsufficiency is a recently described autosomal dominant disease, in which heterozygous ctla mutations result in severe immune dysregulation with variable age of onset and a wide array of clinical manifestations. herein we describe atypical findings in a patient with a novel pathogenic variant in ctla . objectives ) to understand whether the novel ctla variant identified is pathogenic and ) to describe eosinophilic gastrointestinal inflammation and exocrine pancreatic insufficiency as possible manifestations of ctla haploinsufficiency. methods: next generation sequencing (blueprint genetics ©) was used to identify the ctla variant. polyphen and sift (blueprint genetics ©) were used for in silico analysis for the prediction of the effect of this genetic variant on protein structure/function. flow cytometry was used to evaluate ctla expression of regulatory t cells. results: a -year-old boy with type diabetes mellitus and autoimmune thyroiditis presented with abdominal pain, diarrhea, and weight loss. initial studies revealed markedly elevated peripheral blood eosinophils (> cells/μl) and exocrine pancreatic insufficiency (< μg elastase per gram of stool). prominent eosinophilic inflammation was appreciated in biopsies of the stomach, duodenum, jejunum, and terminal ileum. no parasitic infection or inciting drug/food trigger was identified. additional blood studies revealed normal total quantification of t cells but with increased memory t cells ( %, cd +cd +cd ro+) and decreased treg ( %, cd + cd +cd hifoxp hi). b cell quantification and serum immunoglobulin (ig) levels were unremarkable, save a modestly elevated ige level ( iu/ml). comprehensive next-generation sequencing of genes associated with primary immune deficiency revealed a novel heterozygous missense mutation (c. g>a, p.asp asn) affecting the last nucleotide in the ligand binding domain (exon ) of ctla . subsequent analyses revealed decreased ctla expression in the patients t cells compared to healthy controls as well as evolving hypogammaglobulinemia. treatment consisted of methylprednisolone and parenteral nutrition followed by sirolimus and abatacept to which the patient responded favorably. conclusions: we report a -year-old boy with a history of type diabetes mellitus and autoimmune thyroiditis presenting with hypereosinophilia, eosinophilic gastroenteritis, and exocrine pancreatic ins uff iciency as unique m anifestat ions o f ctla haploinsufficiency. although not previously reported in individuals with ctla haploinsufficiency, peripheral blood eosinophilia and eosinophilic inflammation of the gastrointestinal tract have been observed in patients receiving ipilimumab (ctla blocking antibody) suggesting a potential mechanism for the aforementioned findings. severe exocrine pancreatic insufficiency is a rare but observed manifestation in individuals with type diabetes mellitus. whether severe exocrine pancreatic insufficiency would be expected t o o c c u r m o r e f r e q u e n t l y i n i n d i v i d u a l s w i t h c t l a haploinsufficiency and type diabetes mellitus is unclear; however, our reported case and surveillance of others with ctla haploinsufficiency could elucidate incidence and prevalence of this manifestation. introduction/background: mild asymptomatic hypogammaglobulinemia during pregnancy is a well-described phenomenon due to hemodilution. in patients with known humoral primary immunodeficiency such as common variable immunodeficiency, women require an upwards titration of their immunoglobulin replacement dose. isolated symptomatic hypogammaglobulinemia during pregnancy in a patient is not well described in the literature. objectives . understand the physiology of igg during pregnancy. . define a rare entity with a likely genetic predisposition that manifests as hypogammaglobulinemia isolated in pregnancy. . management of this entity with defining goals of treatment. results: year old gravida para female presented with progressive hypogammaglobulinemia restricted to pregnancy starting with rd pregnancy, recurrent otitis media requiring sets of myringotomy tubes, recurrent sinusitis, and streptococcal pharyngitis. her son has common variable immunodeficiency (cvid) requiring immunoglobulin replacement (igrt). prior to conception, her igg was mg/dl. during th week of pregnancy, her igg level was mg/dl. during weeks of pregnancy, she complained of fatigue, and developed an episode of sinusitis that required different antibiotic treatments. at that time, her igg was mg/dl. she was started on subcutaneous igrt maintain igg troughs of > mg/dl. she remained infection-free during her pregnancy and igrt was stopped a few months after delivery with her serum igg level returning to pre-pregnancy levels. patient was initially evaluated during her rd pregnancy with recurrent streptococcal pharyngitis. at that time, her igg was mg/dl. diphtheria, haemophilus influenza, mumps, measles and rubella titers were protective, tetanus titer was non protective with / antipneumococcal titers protective. she was vaccinated with pneumovax and tdap with development of protective titers and remained infectionfree. igrt was not given and post-delivery, her igg levels improved to mg/dl. she was seen one and half years later, for prenatal counseling for her th pregnancy. her immune evaluation included an igg . she demonstrated protection to tetanus, diphtheria and streptococcal pneumoniae. at weeks of gestation, she developed recurrent upper respiratory infections requiring antibiotics. her igg was mg/dl. at weeks of gestation, her igg was mg/dl. igrt was recommended, but patient refused at that time. after delivery, igg improved to mg/dl. conclusions: decreased immunoglobulin levels during pregnancy are welldescribed phenomenon which can be attributed to hemodilution of pregnancy. igg transport to fetus generally begins in the second trimester and reaches its pinnacle in the third trimester. patients with known cvid require dose adjustments (higher) during pregnancy as they can be more symptomatic during this time. here we describe a patient who has an almost normal immune evaluation except for mildly low iga during absence of pregnancy, but during pregnancy, develops recurrent infections with significantly low igg level. her family history of a son with cvid, new daughter with low igg levels like to be transient hypogammaglobulinemia of infancy (thi), another daughter with thi suggests a genetic b-cell defect that manifests as cvid with mildly low iga and hypogammaglobinemia during metabolic stress such as pregnancy. there is only one similar report of a single pregnancy of transient symptomatic hypogammaglobulinemia during pregnancy. such patients should be adequately worked up and treated during pregnancy with igrt to decrease maternal and fetal mortality. introduction/background: card encodes a scaffold protein in lymphocytes that links antigen receptor engagement with downstream signaling to nf-b, jnk, and mtorc . germline mutations in card are known to give rise to distinct primary immune disorders in humans, including scid (null mutations), b cell expansion with nf-b and t cell anergy (benta; gain-of-function mutations), and severe atopic disease (loss-of-function, dominant interfering mutations). objectives: here we report our experience with an expanded cohort of patients harboring novel heterozygous card mutations that extend beyond atopy to include other immunologic phenotypes not previously associated with card mutations. methods: cell transfections and primary t cell assays were utilized to evaluate signaling and function of card variants. results: we demonstrate that in addition to severe atopy, heterozygous missense mutations in card associated with dominant negative activity can present with immunologic phenotypes similar to those observed in stat lof, dock deficiency, common variable immune deficiency (cvid), congenital neutropenia, and immune dysregulation, polyendocrinopathy, enteropathy, x-linked (ipex) syndrome. evaluation of rare or novel card variants found in affected patients showed that dominant negative activity was largely confined to the card or coiled-coil domains, but did not always manifest in atopic disease. conclusions: these results illuminate a broader phenotypic spectrum associated with card mutations in humans, and underscore the need for functional studies to demonstrate that rare gene variants encountered in expected and unexpected phenotypes must nonetheless be validated for pathogenic activity. introduction/background: increasing number of states have been screening for severe combined immune deficiency (scid) as part of the expanded newborn screening program for nearly a decade. in the era of newborn screening, patients with scid present often asymptomatically and are prepared for early hematopoietic stem cell transplantation (hsct). with advances in genetic testing, mutations in over genes have been associated with development of the scid or leaky-scid phenotype. objectives: to present unique cases of hypomorphic x-linked scid where no known pathogenic mutations were identified on initial genetic testing, the il r gamma-chain as protein was expressed, but subsequent testing months later revealed pathogenic il rg mutation affecting either translation or protein function. to expedite earlier identification of pathogenic il rg mutation, we propose screening with x-inactivation studies in maternal t lymphocytes and assessing gamma-chain function by evaluating il r signaling in select male infants with abnormal newborn screen for scid, specifically those with scid phenotype but no identified pathogenic gene mutation on initial genetic testing, and the presence of gamma-chain expression. male patient a presented during the first week of life after his newborn screen was found to be abnormal with undetectable t cell receptor excision circle (trec) count. a phenotype of t-b+nk-scid was established and the patient was sent for bone marrow transplant evaluation. genetic testing revealed a novel hemizygous missense mutation in il rg (p.glu gln), which was a variant of unknown significance. the patient had common gamma-chain expression by flow cytometry on b and nk cells. he underwent haploidentical hsct with his father as donor. unfortunately, his transplant was complicated by prolonged neutropenia, slow t cell reconstitution, and eventual graft failure. to review his next treatment options, it was necessary to prove that his il rg mutation was pathogenic. male patient b presented with concern for an underlying immune deficiency after being hospitalized for peumocystis jirovecii pneumonia. his newborn screening for scid was inadequate at birth and follow up was delayed. retrospective analysis of the newborn screening card at birth confirmed absence of trecs. his phenotype was also t-b+nk-, consistent with x-linked scid. he also proceeded to hsct with a haploidentical parent donor. despite hhv- viremia pre-hsct which persisted posttransplant, he has had appropriate t cell engraftment. comprehensive genetic testing on whole exome level did not reveal any known mutations contributing to his phenotype. patient did have expression of gamma-chain by flow cytometry on t, b and nk cells. however, further testing revealed an il rg utr deletion of aa that, based on similar findings from a prior study can possibly lead to mrna abnormalities. to expedite the association of scid phenotype with x-linked disease, implying gamma-chain pathology, we obtained x-inactivation studies in maternal t-cells that showed severe skewing in both cases. furthermore, il r signaling was impaired on b-cells in each case. the combination of these two assays proved that both patients carry a pathogenic il rg mutation. conclusions: in the era of newborn screening for scid, we are discovering that phenotypic variability of scid patients can be very broad and caused by hypomorphic mutations in the common chain gene. exonbased genetic testing cannot exclude all variants and novel variants of unknown significance have to be evaluated by additional assays, including functional studies for causal effect. it is important to expedite early proof of association with gamma-chain pathology, especially in the era of gene therapy. we propose that in male infants with abnormal scid newborn screening and no known or previously described pathogenic mutation on genetic screen, evaluation continues for hypomorphic il rg mutations. the probability of this process can be increased by a simple screening test for x-inactivation of maternal t lymphocytes. allergy / immunology, allergy / immunology associates inc. / case western reserve university introduction/background: igg -related disease (igg -rd) is an immunologic disorder with multiple clinical presentations previously thought unrelated. it is characterized by the frequent presence of tumor-like swelling of the affected organs and several histopathological findings including tissue lymphoplasmacytic infiltrates with predominantly igg -positive plasma cells and lymphocytes, storiform fibrosis and obliterative phlebitis. humoral immunodeficiency is a term that encompasses several disease entities associated with impaired antibody production. it is suspected in patients who present with recurrent, frequently severe, sinopulmonary infections with encapsulated bacteria, which leads to evaluation of quantitative immunoglobulin levels and vaccine responsiveness. despite a few reports of primary immunodeficiency in patients with serum igg elevation, no adult case has been reported of igg -rd in a patient with concomitant humoral immunodeficiency. objectives: to present a unique case with the presence of concomitant igg -related disease and humoral immunodeficiency. methods: comprehensive chart review of our patient and all performed exams. literature review for igg -related disease, igg elevation in humoral immunodeficiency and concomitance of igg -related disease with humoral immunodeficiency. results: our patient is an -year-old caucasian male with relevant past medical history of chronic bronchitis who was referred to our practice after several episodes of pneumonia in the previous years, with six courses of antibiotics just in the year prior for recurrent sinopulmonary infections. blood tests revealed hypergammaglobulinemia and low level of vaccine responsiveness. chest ct showed multiple bilateral pulmonary nodules and hilar and mediastinal lymphadenopathy. sinus ct showed left maxillary sinus opacification. pulmonary function testing was normal. he later presented with left eye edema, proptosis, diplopia, and painless submandibular salivary gland enlargement. laboratory investigation showed an igg of mg/dl, igg of mg/dl. the patient denied any history of pancreatitis or abdominal pain and abdominal ultrasound was normal. biopsy of a salivary gland was normal. mri of the left orbit was obtained, showing lacrimal gland enlargement. based on the patients recurrent infections, lack of response to tetanus immunization, and limited, non-sustained response to pneumococcal immunization, the patient was started on ivig therapy. the patient was also diagnosed with possible igg -rd based on his salivary gland enlargement and orbital disease in association with hypereosinophilia and increased plasmablast levels. oral prednisone mg daily was started for four weeks, later followed by slow steroid taper (reducing mg every two weeks) with considerable improvement in left eye swelling and proptosis. a few months after discontinuation of the steroids, the orbital disease returned to its previous severity. left lacrimal gland biopsy confirmed igg -related disease, with many areas showing greater than one hundred igg -positive plasma cells per high-power field. after another course of steroids, oral prednisone was weaned to a maintenance dose of mg daily and the patient became asymptomatic from his ophthalmologic complaints with normalization of his ophthalmologic exam. his last checked igg was mg/dl. igg was still elevated at . mg/dl , but given controlled symptoms the patient was spaced to monthly ivig infusions and continued on that daily steroid dosage. conclusions: our patient, initially diagnosed with a humoral immunodeficiency, was later also diagnosed with biopsy-proven igg -related disease, which is a novel association of this two diseases in an adult patient. previous rare reports of association between elevated serum levels of igg and patients with concomitant humoral immunodeficiency were in the presence of isolated igg elevation and not in the presence of igg related disease. this novel association creates a therapeutic dilemma since the patient in question is hypergammaglobulinemic, yet needs ivig, which can lead to side effects such as thrombosis due to a hyperviscous state. the description of additional concomitant cases of both diseases and further understanding of their pathophysiology will be crucial to create awareness and obtain earlier diagnosis, to refine therapeutic options and design adequate treatment protocols. igm and iga anti-pneumococcal capsular polysaccharides as prognostic tool for common variable immunodeficiency: a longitudinal study. professor, sapienza university of rome introduction/background: the clinical spectrum of cvid ranges from a poorly symptomatic form to severe phenotypes characterized by high susceptibility to infections, autoimmunity, granulomatous inflammation, lymphoproliferative disorders, and malignancies. due to high prognosis heterogeneity, prognostic factors are required. objectives: with the aim to identify additional prognostic factors, we evaluated the anti-polysaccharide iga and igm responses by elisa assay in cvid in a longitudinal study over a -year period. methods: patients were immunized at baseline with the -valent pneumococcal polysaccharide vaccine (pneumovax®). twenty healthy donors (hd) were also included. results: as expected, cvid patient had lower igm/iga response than hd. for cvid, four immunological phenotypes were identified by postvaccination igm and iga levels: igm and iga responders ( %), igmhigh responders ( %), igm-low responders ( %) and non-responders ( %). to simplify, we analysed igm-high group with igm and iga responders and igm-low with non-responders. during the follow up, concomitant cvid-related conditions, immunoglobulin serum levels, respiratory infections and outcome were recorded by medical files. cvid igm-low/non-responders developed more frequently respiratory, gastro enteric and autoimmune manifestation and malignancies in comparison to igm-high/igm and iga responders (respectively, pneumonia: % vs % ; chronic diarrhea: % vs %; autoimmunity % vs %). autoimmune cytopenias were not found in the igm-high/igm and iga responders group. eleven ( %) patients died during the study time. survival analysis according to the igm/iga responder status showed that the -years estimated survival for igm-high/igm and iga responders vs igm-low/non-responders group was respectively: % vs %, % vs %, % vs %, % vs %, % vs %, % vs %, % vs %. interesting, in our series only two deaths were due to infective complications: five were consequent to malignancies, one to autoimmune cytopenias and three to not-cvid related conditions. conclusions: in conclusion, even if patients could not raise the protective humoral level, in cvid the anti-polysaccharide iga and igm responses could represent a prognostic factor, individuating groups of patients with less immunological impairment, lower risk of comorbidities and better survival. introduction/background: gaucher disease (gd) is a rare autosomal recessive disorder characterized by a defective function of the catabolic enzyme -glucocerebrosidase (gba) leading to a progressive accumulation of its substrate-glucocerebroside (gc) -in various organs in particular in mononuclear phagocite system. hepatosplenomegaly and cytopenia represent the most common features of the disease. moreover, gd patients also show hyperinflammatory features -secondary to machrophages engorgement and actviation-hypergammaglobulinemia, and a immune-dysregulation involving b , t and nk cells. since clinical phenotpye can be subdolous, symtoms can overlap with alps, however, few data are available on specific immunity pattern in these patients. objectives: to evaluate immune-phenotype and other alps parameters in a cohort of patients with gd methods: we evaluated lymphocytes subsets, immunophenotypic and serological features of alps (dnts, tcr alfa/beta b , b-memory cells, tregs/hla-dr ratio, il- , il- ), and test of apoptosis in a cohort of patients with gd followed-up at igg. results: patients ( in treatment, not) were studied. dnts and tcr alfa/beta b + resulted to be > . % of t-lymphocytes and > % in / ( %) and in / ( %), respectively. b-memory cells and t-regs/hla-dr ratio were < % and < in / patients ( %). / evaluable ( %) had all these parameters concomitantly alterated. / ( %) evaluable patients were resistant to apoposis. il- was pathological in / ( %) patients. all patients had normal levels of il- and sfas. conclusions: this study shows that some patients with gd may present an immune-dysregulation pattern that can overlap with alps features. therefore, the differential diagnosis of gd should be taken into consideration by clinicians during diagnostic work-up of patients with an alpslike phenotype. introduction/background: allogeneic hematopoietic stem cell transplantation (hsct) using unrelated and haploidentical donors is complicated by increased rates of graft-versus-host disease (gvhd) and slow immune reconstitution. selective depletion of alpha/beta t lymphocytes and b cells is a recently developed method of graft manipulation that retains mature natural killer (nk) and gamma/delta t lymphocytes, both of which may exert a graft-versus-leukemia effect and protection against life-threatening infections. objectives: to describe the rate and quality of immune reconstitution, incidence of transplant-related complications, including viral reactivation and gvhd, and overall outcomes following tcr-alpha/beta-and cd depleted hsct for hematologic malignancy in pediatric patients. methods: forty patients of median age . years ( . - . ) underwent hsct for acute myeloid leukemia (n= ), acute lymphoblastic leukemia (n= ), and myelodysplastic syndrome (n= ). grafts were from unrelated (n= ) and haploidentical (n= ) donors. tcr-alpha/beta and cd depletion was performed with the miltenyi clinicmacs plus system. median cd + cell dose was . x /kg ( . - ), and median cd + cell dose was . x /kg ( . - . ). conditioning was with myeloablative busulfan or total body irradiation, cyclophosphamide, and thiotepa. twenty of unrelated donor hscts and / haploidentical hscts also included antithymocyte globulin x . no patient received post-transplantation gvhd prophylaxis. all but patients received rituximab on day + per protocol for recipient positive epstein barr virus (ebv) serology. results: all patients engrafted. median time to neutrophil engraftment was ( - ) days, and median time to platelet engraftment was ( - ) days. one patient experienced graft rejection on day + , and twelve patients relapsed at a median of ( - ) days. overall survival was / ( %) at a median of . ( . - ) months follow-up. two ( %) patients developed grade iii or higher acute gvhd, and ( %) patients developed extensive chronic gvhd. cumulative incidence of cytomegalovirus (cmv) and adenovirus reactivation were / ( . %) and / ( . %), respectively. nine ( . %) patients developed bk hemorrhagic cystitis +/-viremia. ebv reactivation was not observed. median total, myeloid, t cell, and b cell donor chimerism were all % (ranges - %, - %, - %, and - %) at year post-hsct. immune reconstitution of all cells lines was rapid ( table ) . eighteen of ( %) patients had detectable t cell receptor excision circles (trecs) by months with a median trec count of ( - ) per ^ cd t cells, and recovery of the naïve t-cell compartment was observed by months in / ( %) of patients. t cell function as measured by response to pha was normal by months in / ( %) patients and continued to increase steadily with time. despite rituximab on day + for / ( . %) patients, there was rapid b cell reconstitution. nineteen of ( . %) patients had present switched memory b cells at months, and / ( . %) surviving patients are off immunoglobulin replacement at a median of ( - ) months. conclusions: selective tcr-alpha/beta and cd depletion of haploidentical and unrelated grafts results in high engraftment and rapid immune reconstitution with low incidence of gvhd in children with hematologic malignancy. cd depletion and routine post-hsct rituximab on day + are effective at preventing ebv reactivation. introduction/background: hyper-igd syndrome (hids; ) is an autosomal recessive disorder characterized by recurrent episodes of fever associated with lymphadenopathy, arthralgia, gastrointestinal disturbance, and skin rash. the diagnostic hallmark of hids is a constitutively elevated level of serum immunoglobulin d (igd), although patients have been reported with normal igd levels. the disease is associated with mutations in the gene of mevalonate kinase. objectives: male patient, years old, born to non-consanguineous parents, with a history of severe diarrhea since childhood, followed by respiratory infections and pneumonias. moreover he presented several episodes of severe abdominal pain, with intestinal obstruction since seven months old, needing surgical intervention due to acute abdomen. this picture repeated several times until years of age, being submitted to new surgeries due to intestinal suboclusion. at and years he had gastroenteritis and then pancreatitis. years ago new severe acute gastroenterocolitis with suboclusion, submitted to laparotomy and resection of a little part of the gut. he has frequent diarrheas, triggered by coffee and tea. usually evacuates times a day. he was evaluated by several pediatric immunologists in childhood, who distrusted alpha heavy chain disease. he had an intense reaction to the bcg vaccine, and then did not make any more vaccines (only opv (sabin) in the campaigns). he was tonsillectomized at years of age. he had measles, chickenpox, and mumps (at years old). cellulitis at years, repeating several times since then. he presented improvement of pneumonias but still has sinusitis and otitis (approximately times a year introduction/background: pitthopkins syndrome is a rare neurological disorder caused by mutations the tcf gene on chromosome q . clinical features include severe intellectual disability, constipation, microcephaly, and seizures. features distinguishing it from other neurodevelopmental syndromes such as rett syndrome and angelman syndrome include breathing abnormalities (either apneic episodes or hyperventilation) and atypical facial features. typical facial dysmorphism includes bitemporal narrowing, deep-set eyes, an m shaped upper lip, and widely spaced teeth. although a very rare diagnosis (slightly over reported cases), it is not known to be associated with underlying humoral or cellular immunodeficiency. there is only one report of igm abnormalities described in a patient with pitt-hopkins syndrome. objectives: to present a case of pitthopkins syndrome with humoral immunodeficiency. methods: this is a case presentation of a patient with pitt-hopkins syndrome requiring immunoglobulin replacement therapy. results: year old female with genetically diagnosed pitt-hopkins syndrome who presented to our office for immunological evaluation in the setting of recurrent sinopulmonary infections. she was placed on chronic antibiotics by the department of otholaryngology for approximately two years prior to presentation. she was found to be hypogammaglobulinemic (igg: mg/dl, iga: mg/dl, igm: mg/dl), had non-protective titers to streptococcus igg antibody ( / titers protective greater than . mcg/ml) despite booster vaccination with pneumovax, and had borderline tetanus ( . iu/ml) and diphtheria titers ( . iu/ml). given this laboratory evaluation and her recurrent illness, immunoglobulin replacement therapy (igrt) was started. whole exome sequencing was completed to assess for any other genetic cause of her immunodeficiency. the only abnormality was her previous known pathologic variant p.q hfsx c. _ delga (gln his) in exon in the tcf gene. she continued to have infections despite therapeutic igrt. chronic antibiotic treatment was initially tapered, however needed to be reintroduced as ivig alone was not stopping her infections, despite a igg level over mg/dl. conclusions: humoral immunologic deficits are not known to be associated with pitt hopkins syndrome. there has been one case report of a patient with poliomyelitis-like syndrome following an asthma attack in a patient with pitt hopkins syndrome, which was treated with igrt and resulted in a nearly complete recovery. however, igrt was not used for reasons of underlying immunodeficiency. to our knowledge this is the first patient with pitt hopkins syndrome with persistent hypogammaglobulinemia and frequent infections requiring immunoglobulin replacement therapy. it remains unclear why the patient continued to have infections despite igg levels > mg/dl, yet with the combination of igrt and prophylactic antibiotics the patient remains healthy. introduction/background: gata deficiency is a rare disease that typically presents in late childhood or early adulthood with heterogeneous phenotypes including emberger syndrome. emberger syndrome is characterized by lymphedema and predisposition for myelodysplastic syndrome (mds) and acute myeloid leukemia (aml). over % of patients with gata mutations have immune deficiencies. definitive diagnosis of gata deficiency is made by gene sequencing, and treatment includes infection control and potentially hematopoietic stem cell transplant (hsct). objectives: the goal of this report is to contribute data to the small documented cohort of patients with gata deficiency to aid in diagnosis and management of this rare, heterogeneous disorder. methods: we present a case series of three siblings with identical gata mutations with variable phenotypes. a usidnet query resulted patients with gata mutations. results: three siblings ( year-old female (a), year-old male twin (b), and year-old female twin (c)) and their mother had congenital deafness. clinical symptoms include (a) h n influenza requiring mechanical ventilation, warts, and hypogammaglobulinemia, (b) streptococcus pyogenes neck abscess, warts, acne, and mds, and (c) lymphedema and acne. all three patients had absolute monocyte count (amc) of and lymphopenia without documented lymphoid cell dysfunction. a mutation on exon , c. delg, confirmed the diagnosis. all three patients were started on mycobacterial prophylaxis with azithromycin and recommended hpv vaccination. the usidnet query average age of symptom onset was years, and average age at diagnosis was . years. . % ( / ) of patients had a family history of gata deficiency, % ( / ) of patients had warts, . % ( / ) had lymphedema and only patient had sensorineural deafness. % ( / ) of patients had amc of . functional data was limited. conclusions: life threatening infections as well as hematologic malignancies have been reported in patients with gata deficiency, which can be successfully treated with hsct. to our knowledge, the gata mutation detected in this family has not been previously reported. the clinical presentation in these three patients was heterogeneous despite identical genotypes, and diagnosis occurred years after initial symptoms. variable phenotypes were found in the usidnet gata deficiency cohort as well. a high index of suspicion for the disorder and early recognition of clinical manifestations and laboratory abnormalities may aid in timely diagnosis of gata deficiency, with potential for improved outcomes. consulting medical advisor, immune deficiency foundation introduction/background: as individuals with antibody deficiencies age they are susceptible to developing shingles. patients with antibody deficiencies are advised not to receive live viral vaccines such as zostavax, the shingles vaccine. objectives: our survey aimed to determine the frequency of shingles and use of zostavax in common variable immunodeficiency (cvid) and hypogammaglobulinemia patients. methods: , email invitations delivered to members of the immune deficiency foundation database requesting participation in an online survey about zoster, influenza and varicella experiences. data from individuals age years old or older with cvid (n= ; mean age years old; % female; % white non-hispanic) or hypogammaglobulinemia (n= ; mean age years old; % female; % white non-hispanic) were analyzed. results: close to one fifth ( %, n= ) of adults age or older with cvid or hypogammaglobulinemia (n= ) had received shingles vaccination. the majority of those who were vaccinated reported receiving zostavax once ( %, n= ), while % (n= ) received a booster vaccination as well. mild side effects (e.g., skin rash and muscle pain) were only reported by % (n= ) of vaccine recipients after receiving their first vaccination. no side effects were reported after receiving the booster vaccination and no hospitalizations were reported as a result of receiving zostavax. when comparing shingles diagnosis and shingles vaccination, of the adults age years old and older, % (n= ) had been diagnosed with shingles, and of those diagnosed % (n= ) did not receive zostavax. of adults age years old or older, % (n= ) reported a shingles diagnosis. similarly, the cdc reports almost % people in the united states will develop shingles during their lifetime. more than half ( %, n= ) of those ever diagnosed with shingles reported experiencing shingles once, % (n= ) had shingles twice and % (n= ) had shingles three or more times. respondents with more than three shingles episodes were more likely to report their rash lasted more than two months and their blisters became infected. conclusions: almost % of adults with cvid or hypogammaglobulinemia reported receiving zostavax despite recommendations against vaccinations for immunodeficient individuals. however, side effects in those pi patients who received the shingles vaccine appears minimal. though it is possible these individuals were vaccinated prior to diagnosis of their pi; additional patient and physician education on live vaccines and immunodeficiency may be needed as well. the approval of the new non-live virus component varicella zoster vaccine may be of benefit to patients with pi. introduction/background: inclusion body myositis (ibm) is a rare disorder characterized as an inflammatory myopathy with endomysial inflammation and numerous red-rimmed vacuoles seen on biopsy. five cases of ibm have been described in the literature in patients with common variable immunodeficiency (cvid). objectives: to make immunologists aware of ibm as a complication of cvid, that may be incorrectly diagnosed as myositis or autoimmune neuropathy. methods: case description results: we report a -year-old man with common variable immunodeficiency on gammaglobulin replacement who presented complaining of progressively worsening lower extremity pain, weakness, and fatigue. he states that over the last couple of years, it has become difficult climbing and descending steps, arising from a seated position, and has begun experiencing frequent falls. his creatinine kinase level was found to be elevated at u/l and he continued to be lymphopenic with total lymphocyte counts ranging from to k/ul (cd + t-cells %, cd + t-cells %, cd + bcells %). his esr ranged from to mm/h. anti-glutamic acid decarboxy antibody (gad) was initially elevated at . u/ml and rose to . u/ml within months suggestive of a neuropathy. based on an electromyography (emg) and a muscle biopsy, he was diagnosed with polymyositis. he was treated with high dose steroids with no improvement. his intravenous gammaglobulin dose was then increased from mg every weeks to mg per day for straight days every weeks. months later, his creatinine kinase level dropped into the normal range (< u/l), however, he continued to complain of worsening weakness. physical exam showed decreased muscle bulk in forearms and quadriceps bilaterally, lack of a quadriceps tendon reflex, strength / in flexor digitorum profundus and / in hip flexors, and a broad-based gait. histology and electron microscopy of a repeat muscle biopsy identified rimmed muscle vacuoles as typically noted in inclusion body myositis. conclusions: inclusion body myositis is a potential rare complication of common variable immunodeficiency. it can mimic polymyositis and inflammatory demyelinating disorders. high dose steroids and ivig are of no clinical benefit in ibm, despite decreasing serum creatinine kinase levels, which may raise a false impression of a clinical benefit. objectives: in this abstract we present the case of the development of generalized cmv infection in a child with scid. girl n. at the age of months entered the children's infectious clinical hospital with complaints of cough, high febrile temperature for days, refusal to eat. from the anamnesis of life the girl from the st pregnancy, birth, was born full term in weeks gestation, birth weight g. for months of life, a bad increase in body weight was noted and at the time of admission, the weight in months was g. according to the parents, the child had atopic dermatitis. from the anamnesis of the disease on . . , the temperature rose to . °c, there was a cough and a mucous discharge from the nose. then the child refused to eat, the body temperature rose to . °c. at this time, the girl's mom was borne by the ari. january , patient was hospitalized in the hospital with a diagnosis: acute respiratory viral infection, acute rhinitis, pharyngitis, acute bronchitis, toxicosis of - degrees. acute pneumonia? atopic dermatitis, infant form. on january , , due to the worsening of the condition associated with the increase in oxygen (o ) -dependence, the child was transferred to the department of anesthesiology and resuscitation. methods: in the general analysis of blood upon admission, leukocytes are . x / l, hemoglobin is g / l, platelets are x / l, esr is mm / hour, stabs are % (abs. - . x / l), segmented - % (abs- . x / l), lymphocytes - % (abs . x / l), monocytes - % (abs - . x / l). in a biochemical study, the total protein is g / l, total bilirubin is . mol / l, urea is . mmol / l, creatinine is mol / l, lactate dehydrogenase is u / l, alt is u / l, asat - e / l, crp - . mg / l. radiography of the lung from / / -data in favor of interstitial pneumonia. the study of the acid-base ph state is . , pc is . mmhg, po is . mmhg, lactate is . mmol / l. a blood test was performed using the elisa and pcr method for markers of hsv, cmv, enterovirus and toxoplasmosis. ultrasound of the abdominal cavity revealed moderate hepatomegaly, signs of thickening of bile, splenomegaly. moderate diffuse changes in the renal parenchyma (toxic-inflammatory?). the minimum amount of free fluid in the abdominal cavity. ultrasound of the brain revealed signs of subependimal microcyst on the right. according to the immunogram, a sharp decrease in cd + % ( - %) was detected, activated t-lymphocytes (cd + hla-dr +) were . % ( - %), t helper / inducers (cd + cd - . % ( - %) and t suppressors / cytotoxic (cd + cd -) . % ( - %), a high ratio of tx / tc (cd + cd +) was detected . % ( . - . ), cytotoxic non-t cells (cd -cd +) - , , an increase in the number of b-lymphocytes (cd +) - . % ( - %), natural killers (cd + cd +) - . % ( - %), natural t-killers (cd + cd + cd +) - . ( - %), leukocyte gates (cd + cd -) - % ( - %). the absolute content of t-lymphocytes was . x / l, b -lymphocytes - . x / l. the number of thymic migrants (cd + cd ra + cd +) was not detected ( %). according to the results of the immunogram the diagnosis is made: severe combined immunodeficiency (t-b + nk +). / / ct scan of the chest was diagnosed ct signs of a polysergic two-sided inflammatory process in the lungs (figure ). when blood was sown for sterility on january , , staphylococcus epidermidis was isolated in an amount of , sensitive to linezolid, gentamicin resistant to amoxicillin, amoxicillin / clavulonic acid, and ciprofloxacin. on january , , cmv dna was detected in an amount of . × copies / ml. results: since the arrival clarithromycin was administered at a dose of mg / kg per day in divided doses from / / to / / . from / / to / / . change of antibacterial therapy for azithromycin intravenous at a dose of mg / kg per day once a day. . . - / / the state of the child is very severe with negative clinical and laboratory dynamics despite the ongoing therapy. antibacterial therapy was changed to meropenem in a dose of mg / kg intravenously every hours, linezolid at a dose of mg / kg per day and oseltamivir at a dose of mg / kg per day from / / to / / . . . - . . substitution therapy with an octagam in a dose of . g / kg was intravenously dripped. the patient's condition without significant dynamics. based on the results of pcr on cmv, ganciclovir was administered at a dose of mg / kg intravenously drip times a day. / / due to a decrease in platelet count, the platelet mass is transfused and there was a rash all over the body at night, which is associated with the development of the "graft versus host" reaction (gvhr). despite the ongoing therapy, a fatal outcome occurred. the main diagnosis: primary immunodeficiency (severe combined immunodeficiency, t b + nk +). complications: sepsis. septic shock. spon: ards, renal failure, dis, thrombocytopenia, anemia . two-sided lower-lobe pneumonia. generalized cmv infection. gvhd, acute dermal form. concomitant: atopic dermatitis, infant form. conclusions: the peculiarity of the described clinical case was that the patient's first symptoms of scid developed in the first months of life and were manifested by a bad weight gain, atopic dermatitis and the development of a life-threatening generalized cytomegalovirus infection with the development of bilateral low-grade pneumonia, respiratory insufficiency and acute cutaneous gvhd form, after transfusion of unirradiated platelet mass. an expanded immunological study confirmed the diagnosis of scid. methods: this study included patients ( boys and girls) aged months to years. the reasons for entering the hospital were manifestations of severe hepatitis (in children), acute respiratory infection ( children) and patient with symptoms of infectious mononucleosis. all patients were examined according to clinical protocols and given the severity and atypicality of the course of any infectious diseases, patients underwent immunological examination of the blood and they were consulted by an immunologist. all children were diagnosed with congenital immunodeficiency. results: in cases, the trigger for the realization of the immunodeficiency state was infection (e. meningoseptica + kl. pneumonia + b. pertussis; cmv; veb); in patients, giant cell hepatitis occurred. in patients, despite the ongoing therapy, the disease had an unfavorable (lethal) outcome ( patient with hepatitis and patient with generalized cmv infection). conclusions: thus, it should be noted that timely diagnosis of a congenital defect of the immune system and thus timely therapy will avoid adverse outcomes. interferon gamma (actimmune®) effects on severe burkholderia cepacia pneumonia in variant x-linked chronic granulomatous disease professor of pediatrics, university of utah associate professor of clinical pediatrics, keck school of medicine at the university of southern california professor of medicine, university of utah introduction/background: interferon gamma (ifnγ; actimmune®) has been proven to significantly decrease the overall number of infections in patients with chronic granulomatous disease (cgd) when given prophylactically (nejm : , ) . therapy with ifnγ has also been employed to treat severe overwhelming infections in some instances, such as severe aspergillosis with success (jid : , ) . we report here, two patients with very severe burkholderia cepacia (b. cepacia) infection, one of whom was placed on a respirator for approximately two weeks and another who was on extracorporeal membrane oxygenation for an extended period of time. both were treated with ifnγ (actimmune®) in addition to appropriate antimicrobial therapy in an attempt to affect these lifethreatening infections. objectives: the objective of this presentation is to describe two very severe variant x-linked cgd patients with b. cepacia pneumonia who were treated with ifnγ. in addition, we measured both super oxide production as well as nitric oxide production in the stimulated or unstimulated phagocytes from these patients in the presence or absence of interferon gamma. methods: case histories of both patients were reviewed in respect to the severity of their infection, the time spent on a respiratory or extracorporeal membrane oxygenation, the antimicrobial therapy administered, and the clinical results following the administration of interferon gamma as adjunctive immunomodulatory treatment. a standardized neutrophil oxidative burst assay was employed using cytochrome c reduction to measure super oxide production. in addition, nitric oxide was measured in the phagoctyes of the patients after stimulation with phorbol myristate acetate (pma) in the presence or absence of ifnγ using daf- fluorescence dye to detect the production of intracellular nitric oxide. results: a -year-old male developed a left lobar pneumonia and was admitted to primary children's medical center's intensive care unit and treated with iv cefotaxime, clindamycin, later, vancomycin and azithromycin were added. ct scan revealed a left-sided pneumonia and moderate parapneumonic effusion. subsequently, the patient decompensated, was intubated, and placed on respirator therapy. broncheoalveolar lavage and blood grew b. cepacia. neutrophil dihydrorhodamine fluorescence (dhr) demonstrated an intermediate broad peak of fluorescence with a small peak of unactivated cells, while the mother's dhr showed a broad intermediate peak suggesting the carrier state of variant x-linked cgd. both the patient, a . month old younger brother, and the carrier mother were found to have a g to a splice site mutation in exon of the gp -phox gene at position c. confirming the diagnosis of x-linked variant cgd. after approximately weeks on the respirator, ifnγ (actimmune®) therapy was instituted with significant improvement of the patient's lung function. he was taken off the respirator approximately days after the ifnγ therapy was instituted. following addition of ifnγ to his pma-stimulated neutrophil, there was a % increase in superoxide production and a fold increase in nitric oxide in his monocytes. the second patient was a -year-old male who presented with fever and cough and was diagnosed with right-sided middle and lower lobe pneumonia with cavitations. bronchoalveolar lavage grew b. cepacia and nocardia. following increasing ventilatory and circulatory collapse he was placed on extracorporeal membrane oxygenation and treated with - antimicrobial agents. after days of such therapy, ifnγ therapy was initiated and he was weaned from ecmo after days and has remained essentially healthy since then when he is on ifnγ prophylaxis. dhr revealed a broad intermediate peak in the patient and normal and intermediate peaks in the mother suggesting variant x-linked cgd in the patient and the carrier state of x-linked variant cgd in the mother. targeted sequencing revealed a g to a splice site mutation in exom of the cybb gene at position c. confirming the diagnosis of x-linked variant cgd. following addition of ifnγ to this patient's pma stimulated phagocytes, there was a % increase in superoxide production and a % increase in monocyte nitric oxide production. conclusions: two male variant x-linked cgd patients with splice site mutations in the cybb gene and severe life-threatening b. cepacia pneumonia, one on a respirator and one on ecmo were administered ifnγ (actimmune®) and each responded dramatically within - days recovering their respiratory capacity and coming off of assisted ventilation and ecmo, and have continued to do well when on ifnγ (actimmune®) therapy. introduction/background: hyqvia is a recombinant human hyaluronidase (rhuph )-facilitated subcutaneous immunoglobulin (ighy) % replacement therapy for patients with primary immunodeficiency diseases (pidd). objectives: to acquire long-term safety data on ighy, and assess prescribed treatment regimens and administration in routine clinical practice, a global postauthorization safety study (pass) is being conducted. methods: this is an ongoing prospective, non-interventional, open-label, uncontrolled, multicenter study initiated in the united states in november to assess local and systemic effects of ighy within a routine clinical setting. patients aged years with pidd who have been prescribed and/ or have started ighy are eligible for enrollment. patients are followed according to standard clinical practice and their treatment regimen is at the discretion of the treating physician. the presence of anti-rhuph antibody titers is evaluated on a voluntary basis. results: as of august , patients had been enrolled at us study sites. there were no serious aes which were deemed treatment related. sixteen patients experienced a causally related non-serious local ae ( . %; . events/patient-year, . events per infusion) and patients experienced a causally related non-serious systemic ae ( . %, . events/patient year, . events per infusion). of the patients with immunogenicity data, had positive binding antibody test to rhuph (titers : ); no neutralizing rhuph antibodies were detected. conclusions: this interim analysis of prospectively-collected data of ighy use in routine clinical practice indicates that ighy is well tolerated with no treatment-related saes and has not been associated with neutralizing anti-rhuph antibodies in patients with pidd. introduction/background: idiopathic thrombocytopenic purpura (itp) and/or hemolytic anemia accompanied by splenomegaly occurs in up to % of patients with common variable immunodeficiency (cvid). treatments include steroids, other immune suppressants and rituximab. however, in some that do not respond, splenectomy may be performed. while splenectomy is known to be associated with an increased risk of infections or thromboembolic events, studies in other conditions (hemolytic disorders, hereditary spherocytosis, etc), also suggest an increased risk of pulmonary arterial hypertension (pah) after this procedure. objectives: while splenectomy is known to be associated with an increased risk of infections or thromboembolic events, studies in other conditions (hemolytic disorders, hereditary spherocytosis, etc), also suggest an increased risk of pulmonary arterial hypertension (pah) after this procedure. methods: we report three cases of pah following splenectomy for cytopenias in patients with cvid. results: the first is a yo female, with long standing cvid complicated by interstitial lung disease, nodular regenerative liver disease and itp post splenectomy. the second is a yo man with severe bronchiectasis, cirrhosis/nodular regenerative liver disease and itp post splenectomy. the third is a yo woman with cvid (taci compound) complicated by cirrhosis/nodular regenerative liver disease, lung nodules and evans syndrome. all developed severe pah requiring chronic medications. pah in these patients is best classified as multifactorial, group v. conclusions: whether due to thrombus formation, continued cytopenias, and/or vascular changes, we suggest that pah may be a long-term complication of splenectomy in complex cvid. connective tissue, skeletal and vascular abnormalities. two isoforms exist, stat , a amino acid protein, and stat , a amino acid protein produced by alternative splicing of exon resulting in a frame shift and truncated protein at the c-terminus. objectives: we follow patients from families with stat mutations leading to altered c-terminal proteins. the patients have high ige but milder features of ad-hies. methods: clinical data were collected. stat sequencing, stat functional assays as well as lymphocyte phenotyping were performed. results: patient is a year old man diagnosed with hies as a child due to eczema, recurrent boils and high ige ( s iu/ml). he has tortuous and dilated coronary arteries, however denies lung infections, cmc, retained teeth, scoliosis, minimal trauma fractures, or hyperextensible joints. as an adult he developed avascular necrosis of both hips. whole exome sequencing revealed a novel splice mutation, c. + g>t at the end of exon causing skipping of the nucleotide exon as well as utilization of the stat alternative splice acceptor in exon , resulting in a nucleotide deletion. the mutant stat protein product has a amino acid, in-frame deletion encompassing both y and s phosphorylation sites. no stat is made from the mutant allele. lymphocyte phenotyping was unremarkable, however total stat protein levels were decreased in ebv transformed b cell lines and there was decreased y phosphorylation after stimulation. patient (family ) was healthy until diagnosed with severe, refractory coccidiodies pneumonia complicated by pneumothorax with prolonged bronchopleural fistulae at age years. this led to an immune evaluation in which he was found to have elevated ige ( iu/ml, ref . - . iu/ ml). he had one perianal abscess, primary teeth requiring extraction and mild scoliosis, but denies cmc, bacterial pneumonias, eczema, or minimal trauma fractures. stat sequencing revealed a single base insertion in the transactivation domain, c. _ insc causing a frameshift in the stat isoform, p.r pfsx , occurring immediately after the s phoshporylation site. the deletion occurs within the alternatively spliced region of exon , providing an intact, wild-type stat . stimulation with il- or il- showed reduced pstat at y , and elevated pstat which is often seen in other ad-hies patients. lymphocyte phenotyping was unremarkable and th cell analysis showed low-normal levels of th cells while ebv transformed b cells showed reduced total stat levels. his mother and infant sister also share this mutation -the month old infant had normal ige, and intermittent rashes; his mother has high ige ( iu/ml), recurrent sinopulmonary infections (complicated by tobacco use) but without bronchiectasis or pneumatocele, and denies cmc with the exception of pregnancy related vaginal candidiasis. conclusions: loss of function and gain of function mutations in stat lead to distinct syndromes, but it appears that stat mutations affecting the isoform expression, such as those reported here, can also lead to immune dysregulation with incomplete features of ad-hies. these stat mutations will allow us to better understand the relative roles of the isoforms stat and stat in somatic and immune cell signaling. physicians and is associated with immunological defects aswell. mutationsin the kmt d and kdm a genes are the most common genetic changes that lead to kabuki syndrome but for many cases the genetic basis remains unknown. objectives: recognize the varied presentation for a unique immunodeficiency syndrome methods: this is a case series. results: this is a case series describing three patients with ks and their clinical presentations which predominantly involve immunodeficiency and autoimmunity. our first patient is a -year-old male with autoimmune hemolytic anemia (aiha) at a young age, recurrent respiratory infections and hypogammaglobinemia which led to a diagnosis of cvid at the age of six. in addition, he has dysmorphic facial features, intellectual disabilities, and short stature but it was not until his late twenties where he was found to have a missense mutation in kmt d (p.arg cys) which has been described in patients with ks. the second patient is a -year-old female with hypogammaglobinemia, evan's syndrome, short stature, and severe complications which include granulomatous-lymphocytic interstitial lung disease, pulmonary hypertension, and chronic kidney disease. she was also found to have a missense mutation in kmt d (p.arg cys) in her early thirties and passed away from complications of her disease. the third patient is a -year-old female with a history of low iga/igg and poor vaccine titers, aiha, neutropenia, pulmonary nodules, and developmental delay who was diagnosed with cvid in her mid-twenties. for her immunodeficiency and autoimmunity, she was treated with immunoglobulin replacement, rituximab and cyclosporine and was found to have a missense mutation in kmt d (p.cys trp). this mutation was a de novo mutation in this patient and has also been reported in another patient with ks. conclusions: these cases highlight that the presentation of ks is varied and frequently includes immunodeficiency and autoimmunity in addition to the characteristic short stature and developmental delay. a diagnosis of ks remains challenging due the diversity of symptoms and disease severity, the need for genetic testing, and due to overlapping clinical presentations with other developmental conditions. thus, often times, like in these cases, the diagnosis of ks is delayed. chief medical officer, rocket pharma introduction/background: lad-i is a rare disorder of leukocyte adhesion, resulting from itgb gene mutations encoding for the beta- integrin component cd . cd deficiencies prevent integrin dimerization and endothelial leukocyte adhesion, essential for extravasation and antimicrobial activity. severe lad-i (< % of normal neutrophil [pmn] cd levels) is characterized by recurrent serious infections and early mortality unless treated by allogeneic hematopoietic stem cell transplant (hsct). mortality for severe lad-i was reported as % by age in an initial multicenter retrospective study. moderate lad-i ( - % of pmn cd levels) is more indolent; although most patients (pts) survive childhood with recurrent skin and mucosal surface infections; mortality by age can exceed %. lad-i is characterized by umbilical cord complications (delayed separation and omphalitis), poor wound healing and leukocytosis. objectives: reports regarding lad-i have been published in recent decades but no recent comprehensive prognostic assessments are available. we sought an updated understanding of severe lad-i with emphasis on prognosis in the absence of hsct, hsct outcomes and association of cd expression with clinical features. methods: we created a database of all published lad-i cases via pubmed searches and review of available references. results: three hundred twenty-three lad-i cases were reported between - in publications ( case-reports; largest series n= ). the nations reporting the most cases were iran (n= ), usa (n= ), and india (n= ); the highest number of publications were from us centers ( ). pts were considered to have severe lad-i, moderate and were not classified. pmn cd expression levels was reported for cases and was < % in patients ( %) and >= % in pts. four pts with cd > % were considered to have severe lad-i (cd % range . . ). gender was noted for pts; ( %) were male. age at presentation was reported for cases. for pts with cd < %, median presentation was age m (range . - m); for pts with cd >= %, median presentation was age m (range . - m). infection details and cd % were available for ( %) cases. the most frequent infections in pts with cd < % were respiratory tract ( %), sepsis ( %) and otitis media ( %) and for pts with cd >= % they were periodontal ( %), otitis media ( %) and sepsis ( %). perianal skin infections and necrotic skin ulcers were noted in > %. umbilical complications were more frequent in severe lad-i ( of pts with cd < % [ %] and of with cd >= % [ %; p = . ]). for severe lad-i pts with years of follow-up (or death prior to y), there was correlation between absence of umbilical complications and survival to m (p < . ). wbcs were reported in cases (median x /l; range x /l). there were limited correlations between cd expression and wbc (r < . ) and between cd and cd expression (r < . ). mutation analyses were reported in cases with > gene locations noted and mutations on exons , and accounting for % of specified cases. in cases, cd expression was > %; in of cases where cd expression was noted, at least one cd moiety was reported as < %. we sought to understand whether prognosis for severe lad-i in the absence of hsct is similar to the initially-reported % survival to age . there were severe lad-i cases (per investigator assessment or cd < %) for whom survival to years was reported, of whom died prior to age ( % mortality). mortality was similar for the subset of cases reported since ( %, deaths). early mortality was substantially lower in patients with cd >= % and the majority of pts with cd > % survived to adulthood. outcomes for pts who received hsct were consistent with recent series; phenotypic correction was reported in % of pts with hla-matched sibling donors. mortality was % overall ( % for hlamatched sibling recipients). for pts receiving haploidentical hsct there was % mortality and % received subsequent hsct. conclusions: severe lad-i remains a life-threatening condition with limited -year survival in the absence of allogeneic hsct. umbilical complications and granulocytosis are frequent early manifestations; respiratory tract, ear, sepsis, oral and skin infections are common. hsct is potentially curative; transplant-mortality and other complications are frequent, especially in haploidentical recipients. diverse itgb mutations result in lad-i, and genetic evaluation may be valuable for diagnosis and prognosis. rapid identification of pts with potential lad-i (unusual or severe infections in infancy, granulocytosis and umbilical complications) is essential to enable referral to centers with disease expertise. objectives: to determine the phosophorylation of stat in ad-hies patients with known stat mutations. methods: peripheral blood mononuclear cells (pbmcs) were collected from ad-hies patients under irb-approved institutional research protocols. pbmcs were then stimulated with il- or il- for , and minutes. cells were surface stained for cd , cd , cd , cd and cd . they were then fixed, permeabilized and stained with anti-y -stat to evaluate for phosphorylation of stat at position y . cells were then washed and data acquired using flow cytometry. results: pbmcs from one ad-hies patient with a stat sh domain mutation (p.y c) demonstrated normal y phosphorylation after il- stimulation. pbmcs from another ad-hies patient with a dbd mutation (p.h y) exhibited highly reduced stat phosphorylation after il- stimulation and absent phosphorylation after il- stimulation. conclusions: these findings highlight that, in the context of ad-hies, the domain location of the stat mutation does not predict stat phosphorylation potential following stimulation and challenges the current paradigm. ( ) submission id# cheng sun associate professor, institute of immunology introduction/background: as the predominant lymphocyte subset in the liver, natural killer (nk) cells have been shown to be highly correlated with the outcomes of patients with hepatocellular carcinoma (hcc). previously, we reported that nk cells were decreased and functional deficiency in hcc. however, the mechanism underline remains unknown. objectives: in this study, through the use of healthy livers, paired peritumoural tissues (pt) and intratumoural tissues (it) from hcc patients, methods: we have evaluated the expression of cd and its co-ligand receptor btla on hepatic cd + t cells and nk cells. results: decreased expression of cd on nk cells was observed in intratumoural but not pt regions, along with nk cell dysfunction, poor prognosis and tumour metastasis. human cd + nk cells exhibited functional activated, high capacity of ifn-secretion and nk mediated immunity by global transcriptomic analysis of sorted cd + and cd -hepatic nk cells. blocking tgf- specifically reversed the ifn-production of cd + nk cells. in addition, this decreased cd expression is predominantly on cd bright nk cells. conclusions: these findings indicate that cd expression reduction contributes to nk cell exhaustion and tumour immune escape, suggesting that cd has the therapeutic potential for fighting liver cancer. introduction/background: the low number of circulating lymphocytes in the blood is a marker for cellular immunodeficiency in young children. ethnicity also affects the lymphocyte count and ethnicity-specific lymphocyte norms have been used in many countries. this study analyzed the lymphocyte counts in a large cohort of infants and young children from the arabian peninsula. objectives -to define the normal lymphocyte counts in arab children -to define the possible cutoff lymphocyte count that define lymphopenia. methods: this is a cross-sectional analysis of the lymphocyte counts in , arab children. the age groups were: day, - months, - months, - years, - years, - years, - years, and - years, % females. we analyzed the first blood count performed during their visit to the abu dhabi seha ambulatory healthcare services between april and october . the median, th percentile and th percentile counts were calculated. the th percentile lymphocyte count was used to define lymphopenia. the kolmogorov-smirnov test, a non-parametric test, was used to compare lymphocyte counts between groups. statistical significance was defined by a two-tailed p< . . results: the median counts were higher during infancy. the variability (disparity) of the counts (reference intervals) progressively decreased from birth to years of life. the th percentile lymphocyte counts were relatively constant from birth to years ( . - . x /l) and from to years ( . - . x /l), table . the lymphocyte counts were similar in boys and girls. the lymphocyte counts were compared to those from five other studies. conclusions: arab children have lower lymphocyte counts ( th percentile) than children in the united states, brazil and south africa, but their counts are similar to children in china and uganda. our study results support the development and use of ethnicity-specific lymphocyte count standards. the implication of our results is that using these lower cutoff values for lymphopenia will prevent a large number of arab children from having unnecessarily investigated for immunodeficiency. introduction/background: chronic granulomatous disease (cgd) is a rare phagocytic defect caused by mutations in the nadph oxidase system leading to reduced or absent reactive oxygen species production. in addition to specific infectious susceptibility, patients with cgd are predisposed to hyperinflammation in response to infectious agents, autoimmunity, colitis, and other forms of autoinflammation. cgd patients with mutations in p phox are at increased risk of diabetes and cardiovascular disease. hyperinflammatory and auto-inflammation responses are often difficult to predict and manage. intracellular adhesion molecule- (icam- ) and e-selectin are endothelial adhesion markers that facilitate the adhesion and transendothelial migration of leukocytes and elevations in these markers have been associated with cardiovascular disease, glomerular injury, and thrombotic events. expression of adhesion molecules is induced by pro-inflammatory cytokines and are associated with a hyperinflammatory state. objectives: to determine if e-selectin and icam- are elevated in patients with cgd and to determine of endothelial adhesion markers could serve as a biomarker of inflammatory disease in cgd patients. methods: thirty-eight pediatric and adult subjects with cgd ( xlinked (xl-cgd), p phox deficient and p phox deficient, and x-linked cgd carriers were enrolled. e-selectin (pg/ml) and icam- (ng/ml) were measured from the plasma of patients via sandwich elisa. results: all cgd patients had histories of severe infection, active infection, chronic colitis, or other autoimmune disease at time of evaluation. nine p phox deficient cgd patients had history of diabetes and/or early onset cardiovascular disease. one subject with x-linked cgd had undergone hematopoietic stem cell transplantation (hsct). plasma levels of e-selectin were significantly elevated above healthy controls (median , pg/ml) in subjects with xl-cgd (median , pg/ ml, p< . ), p phox deficient cgd (median , pg/ml, p= . ) and p phox deficient cgd ( , pg/ml). plasma levels of icam- were also elevated above healthy controls in subjects with xl-cgd (median . ng/ml), p phox deficient cgd (median . ng/ml), and p phox deficient cgd ( . ng/ml), although none were statistically significant. plasma quantities of e-selectin and icam- increased further in those p phox deficient patients with diabetes and/or cardiovascular disease (median e-selectin , pg/ml, icam- . ng/ml) but neither reached statistical significance. e-selectin and icam- quantities in female carriers of xl-cgd and in xl-cgd cured by hsct were similar to values found in healthy controls. conclusions: immune dysregulatory features and hyperinflammation in cgd can be difficult to predict and manage. the endothelial adhesion markers e-selectin and icam- are elevated in patients with xl-cgd and p phox deficient cgd that worsens with presence of early onset cardiovascular diseases and resolves post-hsct. elevations in e-selectin and icam- in the serum of cgd patients may serve as surrogate markers of inflammation and suggest a chronic endotheliopathy in cgd patients. introduction/background: the phenotypic presentation of ctla haploinsufficiency was only recently described. the management of these patients in the medical literature is limited to anecdotal case reports. we aimed to detail our experience with short and long term immunomodulatory therapy in treating autoimmune cytopenias in the background of ctla impairment. objectives: we aimed to assess the efficacy of mtor inhibitors in treating autoimmune cytopenias in patients with ctla haploinsufficiency. methods: we retrospectively identified patients with proven ctla mutations and documented refractory autoimmune cytopenias while receiving care at nih clinical center (from july to august ). the complete (cr) and partial (pr) clinical response was assessed after six weeks of treatment and defined as hgb > g/dl, platelets > k/ul and hgb > g/ dl, platelets > k/ul, respectively without transfusion requirement. results: all analyzed patients failed or exhibited disease recurrence on at least one prior medical therapy, including: corticosteroids, rituximab, romiplostim and eltrombopag. the initial response rate to evrolimus and/or sirolimus was % ( cr and pr). one of the partial responders had recurrence of idiopathic thrombocytopenic purpura at four months while on rapalogs. overall, we used mtor inhibitors in patients for a total of patient years to treatm multiple modalities. the top three most common recorded adverse events were: clostridium difficile colitis (n= , in patients), lipid abnormalities (n = , patients required treatment) and bacterial pneumonia (n= , in patients). conclusions: our limited retrospective data suggests that mtor inhibitors might be efficacious in the treatment of autoimmune cytopenias in ctla haploinsufficent patients. further prospective studies are required to assess safety and efficacy of mtor inhibitors in this patient population. association with frequent upper respiratory tract infections and pharyngitis. from years of age she continued to have recurrent febrile episodes in the absence of infection, with fever of - degrees celsius on a monthly basis, normally lasting - days. she also developed episodes of urticaria, temporally unrelated to her febrile episodes. the rash was noted to be induced by exposure to the cold, and would generally last - days with some clinical response to antihistamines and naproxen. her clinical picture then progressed and she developed joint pain and swelling, particularly affecting her hands and feet. over the following years the patient continued to have recurrent episodes of urticaria as well as progressive joint involvement and limitation. there was some initial improvement with naproxen and prednisone. her symptoms were refractory to subsequent treatment with methotrexate, leflunomide, infliximab and tocilizumab and she remained corticodependent. immunological work up including lymphocyte phenotype, immunoglobulins, vaccine responses to both protein and live vaccines and ch were normal. there was no evidence of raised inflammatory markers with esr - mm/h, crp < . mg/l and ferritin ug/l. autoimmune workup including ana, ena, anti-dna, anca and c and c was normal. the initial differential diagnoses included systemic juvenile arthritis, periodic fever syndrome or cryopyrin associated periodic syndromes (caps). the patient therefore had a periodic fever gene panel that identified a heterozygous mutation in exon of nlrp , c. c>t (p.arg trp). whilst mutations in nlrp are known to be associated with familial cold autoinflammatory syndrome, this was reported as a variant of unknown significance. we therefore proceeded with functional in vitro testing to demonstrate pathogenicity. the patients monocytes showed increased secretion of il b upon stimulation with lps as compared to healthy donors. when tested in a luciferase reporter assay, the mutated nlrp partially lost the capacity to inhibit the nfkb pathway. overall these in vitro studies show that this nlrp mutation results in defective regulation of the inflammatory response. the patient was commenced on anti-il therapy with canakinumab, with no clinical improvement so was therefore discontinued. we report a case of familial cold autoinflammatory syndrome due to a mutation in exon of nlrp , that to this point has remained refractory to multiple treatment modalities. functional testing was able to demonstrate mutation causality and defective regulation of the inflammatory response. objectives: authors aim to evaluate the spectrum of clinical phenotypes associated with nemo hypomorphic mutations within the usidnet registry. methods: investigators obtained demographic, laboratory, and clinical data on patients with a defect in nemo within the usidnet registry. results: there were male patients within the usidnet registry with a diagnosis of eda-id attributed to nemo hypomorphic mutation. of these, were associated with a known variant in nemo (e x, f l, m v), and were associated with previously unreported variants (d v, e del, c. - g>c). for patients, a mutation was not specified. most reported having an affected family member (n= , %). median age of symptom onset and diagnosis were year (iqr . - y) and years (iqr - y), respectively. median age at most recent visit was years (iqr - y). infections, additional clinical features, treatments, and outcomes are summarized in table . skin manifestations (n= , %) and pulmonary complaints (n= , %) were common, with eczema (n= , %) and asthma being most prevalent (n= , %). gastrointestinal conditions (n= , %) were also frequently reported, and included non-specific diarrheal illness, enteropathy, colitis, enteritis, and inflammatory bowel disease. neurologic features (n= , %), including seizures, hearing defect, peripheral neuropathy, and encephalopathy, were unexpectedly common, suggesting a previously unrecognized disease association. conclusions: we observed that allergic diseases, including asthma and eczema, were common in patients with nemo mutation. notably, varied neurologic features were more prevalent than previously reported. this study highlights the potential of cross-institutional registry analysis to deepen our understanding of extremely rare genetic diseases. coordinated effort across institutions is required to better characterize the spectrum of clinical phenotypes associated with hypomorphic mutations in nemo. mycobacterial lysate (ml) and purified protein (ppd) in the diagnosis of patients with mendelian susceptibility to mycobacterial disease (msmd) elimination of this infection depends mainly on the success of the interaction between macrophages and infected t lymphocytes. patients with mendelian susceptibility to mycobacterial disease (msmd) present severe and recurrent infections due to impaired signaling of the ifn/il- axis. objectives: our aim was to evaluated the ifn/il- axis of patients with clinical history suggestive of msmd using mycobacterial lysate (ml) and purified protein (ppd) by elisa assay. methods: samples of patients (n= ) with a clinical history suggestive of msmd arrived in our laboratory. for the diagnosis, ml of blood diluted ( : ) in rpmi culture medium supplemented were used. the samples were distributed in two different plates, one used in the dosage of il ( h) and the other in the ifn dosage ( h). thus, they were stimulated with ml ( ug/ml for the h plate and ug/ml for the h plate) and with ppd ( ug/ml for the h plate and ug/ml for plate of h). at the same time, in half of the wells stimulated with lm and ppd, the cytokine ifn ( iu/ml) or il p ( ng/ml) were added in the plate. next, the plates were incubated at °c and % co² and the supernatant were collected and quantified by the elisa assay. the results were evaluated by the statistical mann whitney u test. results: six of the patients presented alterations in the evaluation of the ifn/il- axis. the age of the diagnosis of male patients ranged from to years. the only two female patients diagnosed were and years old. the clinical history was heterogeneous: had lymph node hyperplasia, pneumonia, colitis, herpes zozter, another had a urinary tract infection and bcgitis. the pathogens isolated were the m. tuberculosis, m. abscessus, m. gordonae, m. genavense and m. konsossi species found in different patients. statistical analysis of the ifn-/il- axis evaluation by elisa was performed.the results of the dosage of ifn were significant in samples with lm and lm plus il- (***). similar results were observed in samples treated with ppd and ppd plus il- (**) when compared with healthy controls. in the il- dosage, statistical difference was observed in the samples with lm (***) and with lm plus ifn (*). in samples stimulated with ppd, the results did not show statistical differences (ns.) but ppd + ifn (*) were significantly different. conclusions: six patients were diagnosed by the evaluation the il- / ifn axis. the use of micobacterial lysate (ml) showed reliable results to the diagnosis of patients with il /ifn pathway defects. the genetics diagnosis will be performed. introduction/background: invasive infections due to mycobacterial species are a feared complication in patients with t-cell deficiency or phagocyte disorders, and treatment is frequently complicated by antimicrobial resistance. cd + th t-cell immunity is known to be critical to antimycobacterial defense; accordingly, adoptive t-cell immunotherapy with mycobacteria-specific t-cells (mst) may be a beneficial therapy for combating these infections. objectives: to determine if ex vivo expansion of t-cells targeting common mycobacterial antigens is feasible from healthy donors, and whether the same antigens are recognized by patients with primary immunodeficiency (pid) and invasive mycobacterial infections. methods: peripheral blood mononuclear cells (pbmc) from healthy donors were pulsed with overlapping -mer peptide libraries encompassing five mycobacterial antigens (ag b, ppe , esat- , cpf , adk) and expanded for days with cytokines il- and il- . expanded msts were tested for specificity against the targeted antigens via ifn-g elispot, multiplex cytokine analysis, and flow cytometry. pbmcs from pid patients with invasive mycobacterial infections were similarly tested for presence of t-cells recognizing the tested mycobacterial antigens. a minimum of spots per x cells above negative control was considered specific on elispot. results: ten healthy donors and eight patients with pid were tested. specificity against - mycobacterial antigens (median ) was confirmed in all ten healthy donors, with a mean . -fold cellular expansion during the -day culture. msts were predominantly cd + t-cells (mean/sd: +/- %), with both central memory (mean/sd . +/ - . %) and effector memory (mean/sd . +/- . %) populations. there was no clear difference in antigen specificity between bcg immunized (n= ) and bcg naïve (n= ) healthy donors. six of pid patients had no detectable immunity to tested mycobacterial antigens. one patient with combined immunodeficiency has a low detectable specificity to ag b (mean spot forming colonies[sfc]), and a patient with nfkb haploinsufficiency mounted a response against ag b (mean sfc) and ppe (mean sfc). conclusions: mycobacteria-specific t-cells can be rapidly expanded from healthy donors utilizing a protocol that could easily be translated to a good manufacturing practices facility. the majority of tested pid patients lacked immunity to the targeted antigens. adoptive immunotherapy with msts derived from third-party healthy donors may be a beneficial adjunctive therapy for pid patients with invasive mycobacterial infections. is an innate immune deficiency, primarily affecting the phagocytic compartment, and presenting with a diverse phenotypic spectrum ranging from severe childhood infections to monogenic inflammatory bowel disease. dihydrorhodamine (dhr) flow cytometry is the standard diagnostic test for cgd, and correlates with nadph oxidase activity. while there may be partial genotype correlation with the dhr flow pattern, in several patients, there is no correlation. objectives: in such patients, assessment by flow cytometric evaluation of nadph oxidase-specific (nox) proteins provides a convenient and rapid means of genetic triage (table) . methods: we performed dhr flow cytometry and nox flow cytometry on granulocytes and monocytes of cgd patients. results: phenotypic and laboratory patient data shown in table. *p had decreased p phox (% and mfi) monocytes, but not in granulocytes. all other siblings (p , p , and p ), and mother (p ) had relatively higher p phox (%) monocytes, but still lower than the healthy control. p had normal %p phox monocytes, comparable to control. however, p -p , and p had normal p phox (mfi) in monocytes and granulocytes. p - , and p have normal %gp phox+ granulocytes, while p and p have modestly decreased %gp phox in monocytes. p - , and p have all moderately decreased gp phox protein (mfi) in granulocytes and monocytes compared to healthy control, with a single population for protein expression. p , p and p have bimodal populations for gp phox in monocytes but not in granulocytes, with a larger positive population, and a much smaller negative population. the data from p - suggest that the amount of gp phox does not necessarily correlate with neutrophil oxidative burst, as measured by dhr. also, not all cybb variants affect p phox protein expression, though both proteins are membrane-bound. the cyba vus in p does not appear to have affected p phox protein expression in either monocytes or granulocytes. conclusions: the atypical clinical presentation of some cgd patients can make genotype-phenotype correlation with dhr flow data challenging. genetic testing, while necessary, can take several weeks. however, nadph-oxidase specific-protein flow assessment offers a rapid alternative to identification of the underlying genetic defect, and can be utilized as a reflex test to an abnormal dhr flow. further, it can provide insight into correlation between oxidative burst relative to protein expression in granulocytes and monocytes. head, division of immunology and allergy, the hospital for sick children introduction/background: adenosine deaminase (ada) is a ubiquitous enzyme important for purine metabolism. few studies have indicated that ada deficiency, in addition to causing profound lymphopenia and susceptibility to infections, is associated with neutrophils abnormalities. objectives: determine whether ada deficiency directly affects neutrophils and what are the mechanisms involved. methods: peripheral blood (pb) and bone marrow (bm) from . weeks old ada-deficient (ada-/-) mice that closely recapitulate the phenotype observed in ada-deficient patients, as well as ada+/-littermates were used to study neutrophils development and function. some experiments were supplemented with -week old ada-/-mice, maintained until weeks of age with ada enzyme replacement, and littermates. results: the number of neutrophils in pb of ada-/-mice at . and weeks of age was similar to ada+/-mice. the function of pb neutrophils from ada-/-mice, determined by oxidative burst, was also normal. the percentage of lin-/c-kit+/sca + hematopoietic progenitor cells in bm demonstrated significant reduction in ada-/-mice compared to littermates ( . ± . % and . ± . %, respectively, p= . ). moreover, expansion of bm isolated from ada-/-mice in methylcellulose resulted in significantly less cd b+/ly- g+ neutrophils compared to healthy controls ( . ± . % compared to . ± . %, respectively, p= . ). proliferation of bm cells, determined by brdu incorporation into cells dna, was higher in ada-/-mice than in littermates, possibly contributing to the normal neutrophil numbers in pb of ada-/-mice. conclusions: ada deficiency directly affects neutrophil development. further studies will help understand the significance of these effects and potential therapies for ada-deficient patients. objectives: we present here the clinico-pathologic features of a -yearold female with a heterozygous fancd gene deletion/mutation with evidence of cellular and humoral immune dysregulation. methods: we evaluated the patient using standard immunology anatomic, cellular, and biochemical functional assays. results: the patient has multiple dysmorphias including total anomalous venous return (repaired), mesomelia, absent ear canal, radial ray dysplasia, and short stature. her medical history is significant for an episode of pneumococcal sepsis despite adequate vaccination. whole exome sequencing demonstrated deletion of exons - and a pathologic mutation (c. g>a, p.arg gln). repeated blood samples and immunophenotyping demonstrated severe lymphopenia. there were markedly low cd + t-cell counts with a low cd :cd ratio ( . ). changes in the composition of the b-cell population included: significantly diminished absolute total b-cells, elevated immature cells, low levels of transitional cells, and undetectable advanced b-cell populations. there was no immunogenic response to pcv- or varicella/tetanus/ diphtheria vaccination. the nk-cell count was unaffected and demonstrated normal spontaneous and stimulated cytotoxic response. bone marrow analysis demonstrated hypocellularity without dysplasia. conclusions: we report here a pediatric patient with a novel fancd deletion/mutation presenting with severe lymphopenia in two cell compartments (b and t cells) and susceptibility to invasive bacterial infection. the findings are suggestive of combined immune deficiency. the cellular immune profile suggests that fancd may be involved in the transition of immature b and t cells to mature cells, a process that requires substantial dna recombination. additional genetic and biochemical evaluation is needed to further characterize this rare clinical finding. introduction/background: familial hemophagocytic lymphohistiocytosis type (hlh) is a rare fatal condition due to a mutation in the pfr gene on chromosome q - inherited in an autosomal recessive pattern which results in overactivation of the immune system. symptoms usually manifest before year of age. a -year-old previously healthy male presented with acute onset of bilateral lower extremity pain and weakness, with subsequent inability to ambulate over a -day period. neurological exam demonstrated decreased lower extremity power, sensation, absent patellar and achilles reflexes, and a wide-based gait. objectives: not applicable methods: not applicable results: laboratory data was significant for neutropenia and thrombocytopenia, elevated levels of ferritin and serum cd , and a positive ebv pcr. patient was initially treated with ivig for possible ebv-driven guillain-barre syndrome with some improvement in neurologic symptoms as well as thrombocytopenia. a mass of retroperitoneal lymph nodes were noted on spinal mri. testing was negative for alps and bone marrow biopsy was negative for leukemia/lymphoma. further work-up revealed absent perforin expression in cytotoxic cells and normal sap protein expression on staining, with poor nk function. genetic testing revealed a pathogenic mutation in the pfr gene with additional variant of unknown significance. the patient was treated with rituximab for persistent ebvand returned with worsening lower extremity weakness. on mri, focal enhancements were found in the brain as well as worsening mass compression of the lumbosacral nerve roots. nerve root biopsy showed histiocytes with hemophagocytosis and a dense lymphohistiocytic infiltrate which stained positive for cd , cd , and granzyme b, with some loss of cd and no perforin. bone marrow biopsy was negative for hemophagocytes. conclusions: the patient was diagnosed with worsening familial hlh with cns involvement. hlh-directed chemotherapy (dexamethasone, cyclosporine, etoposide, intra-ommaya methotrexate/hydrocortisone) was started and hsct was performed. familial hlh is a rare and often lethal disorder that generally presents at a very young age. the acute onset and severity of presentation as occurred in this previously healthy adolescent is uncommon. familial hlh should be considered even in older patients with unexplained overactivation of the immune system. is the most profound form of primary immune deficiency, and is usually fatal in the first year of life without treatment. newborn screening for scid using quantitative analysis of t-cell receptor excision circles (trecs), has become the accepted method to facilitate early diagnosis and treatment in most of the united states, as scid babies typically do not make trecs. ikbkb deficiency is a rare form of autosomal recessive scid found in the northern cree first nations people of canada, where t cells develop normally but are non-functional. trec analysis is expected to be normal in ikbkb scid, and does not identify these cases. objectives: the objective of our study was to determine the feasibility of targeted genetic newborn testing for ikbkb deficiency. methods: we implemented a pilot project of prospective targeted genetic testing for the previously described homozygous ikbkb mutation (c. dupg) in newborns from small northern manitoba communities. between and , dna was extracted from dried blood spots of newborns, and targeted sanger sequencing of the mutationharbouring ikbkb exon was performed. results: all infants born in the selected communities underwent testing. fifty-five infants ( . %, or / . ) were found to be heterozygous carriers. one affected infant was identified, and underwent hematopoietic stem cell transplant before onset of infections. our findings are consistent with the predicted homozygosity for this mutation ( / . x / . x / = / births). conclusions: we demonstrated that targeted newborn testing for ikbkb deficiency was feasible, and provided the first prospective estimate of the ikbkb mutation carrier frequency in select manitoba northern cree first nations populations. we suggest that if we are to capture all babies with scid in manitoba, future newborn screening should be universal and include both trecs and direct mutation testing for population-specific mutations, including the first nations ikbkb mutation. high throughput analysis for trecs and targeted mutations will be introduced for universal newborn screening in manitoba. introduction/background: immunoglobulin class-switch recombination (csr) and somatic hypermutations (shm) are prerequisites of antibody and immunoglobulin receptor maturation and diversity within the adaptive immune system. the mismatch repair (mmr) machinery, consisting of homologues of mutsa, mutla, and mutsb (msh /msh , mlh /pms , and msh /msh , respectively) and other enzymes, is involved in csr, e.g. as backup of nonhomologous end-joining repair of activation-induced cytidine deaminaseinduced dna mismatches, and furthermore, in addition to errorprone polymerases, in the repair of shm-induced dna breaks. in line, a varying degree of antibody deficiency, from iga or selective igg subclass deficiency, to common variable immunodeficiency and hyper-igm syndrome have been shown in small numbers of patients with constitutional mmr deficiency (cmmrd) in addition to the known severe cancer predisposition due to genomic instability of patients with biallelic loss-of function mutations in one the mmr components. objectives: to elucidate the clinical relevance of primary immunodeficiency (pid) in cmmrd, we collected history and laboratory data of a novel cohort of consecutive patients from families with homozygous mutations in pms (n= ), msh (n= ), and mlh (n= ) reported to the consortium care for cmmrd (c cmmrd) between and , most of whom manifested with typical malignancies during childhood. methods: retrospective chart review according to a specific questionnaire and extended routine immunological analyses were performed with irb approval from the medical university of graz, austria. results: none of the presented patients fulfilled any classical or extended clinical warning signs of pid (infections, immune dysregulation, inflammation). furthermore, analyzing multiple specific laboratory parameters of the humoral and cellular immune system, we could not detect a uniform pattern of abnormalities. importantly, our data do not confirm previous suggestive evidence of iga or igg subclass deficiency, a specific antibody formation, or a b memory cell maturation defect. results of next generation sequencingbased detection of impaired class switch recombination and somatic hypermutations are pending. the t cell subsets and receptor repertoires were unaffected. together, neither clinical nor laboratory parameters were suggestive of pid in the present series of novel cmmrd patients. conclusions: we conclude that patients with cmmrd do not generally show a clinically relevant pid that could facilitate early diagnosis. on the contrary, these data support the prospect of potentially successful immune therapy of malignancies in the context of cmmrd. introduction/background: ada- deficiency is immune dysregulation diseases caused by an autosomal recessive mutation on cecr gene characterized by polyarteritis nodosa, childhood-onset, early-onset recurrent ischemic stroke and fever. objectives: report a new mutation on the cecr gene resulting in ada- deficient children. methods: female, -year-old, presented history of multiple ischemic strokes at one-year-old associated with recurrent fever and livedo racemosa. she has no siblings and parents are not consanguineous. results: the laboratory evaluation shows red cell= . x /ml, hemoglobin= . g/dl, leucocytes= . x /ml, neutrophils= . x / ml, lymphocytes= . x /ml and plateletes= x /ml. ige< ui/ml, igg= mg/dl, igm= . mg/dl and iga= . mg/dl. subsets lymphocytes shows cd += . %, cd /cd = . , cd + = % and cd / = . %. due the clinical history, we performed cecr gene sequence homozygous substitution located at position - of acceptor splice site intron , c. - g>a. this is a high conservative region with no alteration along phylogenetic studies and predicted to be pathogenic. to confirm the functional alteration, the ada- activity was tested in dried plasma spot showing . mu/g protein confirm the gene loss of function and the mutation pathogenicity. conclusions: the authors presented a novel mutation of cecr gene, the first one described in splice site causing gene loss of function and confirmed by extremely reduced ada activity. introduction/background: severe combined immunodeficiency (scid) is the most severe form of primary immunodeficiency characterized by severe, life threatening infections during early infancy. scid is a medical emergency associated with significant mortality if hematopoietic stem cell transplantations is not instituted early in the course of the disease. scid is a genetically heterogeneous disease caused by mutations in more than different genes. different genes are implicated in different ethnic populations and geographical locales depending on the rates of consanguinity and endogamy in these populations. however, following the institution of newborn screening of scid in almost all state of the us and the widespread use of next generation sequencing in primary immunodeficiency diseases ar-scid due to mutations in rag and rag genes are found to be more prevalent than reported earlier. objectives: we performed a retrospective analysis of scid cases diagnosed at our centre and referred to us from other centres to determine the clinical, immunological and genetic basis of the disease in these cases. genetic variants both recurrent and novel were analysed in detail. methods: fifty six ( ) of the suspected patients met the esid diagnostic criteria. the clinical features, immunological defects and the gene sequencing results of these patients were analysed. gene sequencing was performed at the our centre and other collaborative centres at dept of pediatrics and adolescent medicine, queen mary hospital, hong kong, national defense medical college, saitama japan, kazusa dna research centre, chiba, japan and duke medical university centre, usa. mutations were detected in of the patients. mutations were classified as recurrent or novel after checking different databases such as exome aggregation consortium (exac), human gene mutation database (hgmd) and other relevant scid databases. the effect of novel, previously unreported mutations was determined using in-silico prediction tools such as sift and polyphen . functional studies were also performed in few cases to determine the effect of novel mutations. results: mutations were detected in patients. mutations were more common in genes causing autosomal recessive form of scid than the x-linked variant. mutations were detected in il rg gene in patients followed by mutation in the rag gene in patients, dclre c in patients and rag in patients. mutations were also detected in ada gene ( patients, mutations), il r ( patients), stim , pnp and nhej ( patient each). nine novel mutations were detected. three in il rg gene, in rag , in ada and one each in the nhej and il r gene. conclusions: autosomal recessive form of scid was more common in our cohort compared to x-linked form of the disease. mutations in rag and rag genes were the commonest ( patients) followed by mutation in il rg gene ( patients). nine novel mutations in different pid genes were detected in our cohort of scid patients introduction/background: rasgrp is a guanine-nucleotide exchange factor which phosphorylates ras-gdp to the activated form ras-gtp in response to t-cell receptor stimulation, resulting in ras activation. mutations in the gene coding for rasgrp have been recently described in four patients with profound t-cell deficiency, resulting in recurrent bacterial and viral infections, autoimmunity and malignancy. here we describe a two-year-old male presenting with recurrent sino-pulmonary infections, found to have two variants in rasgrp , one not previously described. objectives . to describe a case of combined immunodeficiency with two pathogenic compound heterozygous rasgrp mutations. . to compare the clinical phenotype of rasgrp deficiency of our patient with that of previously described cases. . to argue for early hematopoietic stem cell transplantation in view of the increased susceptibility to epstein barr virus (ebv) induced lymphoma in patients with rasgrp deficiency. methods: a two-year-old male was referred to seattle children's immunology clinic for recurrent otitis media and two episodes of pneumonia. the diagnosis of combined immunodeficiency was considered based on a profound t-cell deficiency during immune evaluation and he was started on azithromycin and tmp/smx prophylaxis. at age ½ years he was hospitalized with a bladder outlet obstruction and found to have two abdominal masses. biopsies were obtained and he was diagnosed with an ebv driven b cell lymphoproliferative disorder. in addition, his csf and bone marrow were considered positive based on pcr and staining, respectively. treatment with cyclophosphamide, prednisone, rituximab, and intrathecal methotrexate was initiated. due to poor csf ebv clearance, intrathecal therapy was escalated to rituximab. results: initial laboratory evaluation showed elevated igg ( mg/dl), normal number of cd b-lymphocytes, and adequate response to tetanus, prevnar- and varicella vaccine. b-cell phenotyping showed elevated immature/transitional b-cells. a profound t-cell defect was identified with cd t-cell lymphopenia ( /mm ), elevated cd t-cells ( /mm ), inverted cd /cd ratio ( . ) and absent proliferation in response to mitogens (pha, anti-cd ) and antigen (tetanus). forty-three percent of peripheral blood t-cells were / positive. t-cell phenotyping revealed decreased cd and cd naïve t-cells with elevated proportion of cd + t-effector memory t-cells. cervical lymph node and retroperitoneal mass biopsies showed atypical lymphoproliferation without malignant transformation. exome sequencing revealed two variants in rasgrp . the first variant (c. + g>a) is located at a splice site predicting an unstable transcript targeted for degradation. the second variant (c. c>t) is a novel mutation resulting in a stop codon. conclusions: recurrent sino-pulmonary infections are often a presentation of antibody deficiency. in this case, further investigation showed a profound t-cell defect, resembling that reported in patients with homozygous nonsense mutations in the catalytic domain of rasgrp and patients with homozygous insertion mutations leading to a premature stop codon at the bzip domain. our patient had biallelic mutations in rasgrp downstream of the catalytic domain, both leading to unstable transcripts. he developed an ebv induced atypical lymphoproliferative disorder, a complication reported in one rasgrp deficient patient whose disease progressed to b cell lymphoma and unsuccessful hsct. another patient developed ebv induced lymphoproliferative disorder after hsct for ebv-positive hodgkin lymphoma. two additional patients presented with recurrent infections, developed b-cell lymphoma and one was successfully transplanted. we are preparing the patient for hsct after chemotherapy for his lymphoproliferative disease to correct the underlying immune defect given that these patients are at high risk of developing lymphoma following ebv infection. introduction/background: immunodeficiency-centromeric instabilityfacial anomaly is a group of rare genetic disorders typically involving agammaglobulinemia. type four is caused by variants in the hells gene. the five patients previously reported with icf have fit the phenotype of agammaglobulinemia. here we report a patient with novel phenotype including neutropenia and neuroblastoma. objectives: describe a unique presentation of immunodeficiencycentromeric-instability-facial anomaly syndrome to further expand our understanding of this disease. methods: retrospective chart review results: six-month old male was transferred to our tertiary care facility for ongoing chronic respiratory infection, chronic diarrhea, and failure to thrive. his past medical history was significant for -week prematurity due to rupture of membranes requiring a two month nicu stay for bronchopulmonary disease. upon discharge, he was bottle feeding and on room air. he had recurrent congestion for three months with two courses of antibiotics and one and a half weeks of diarrhea leading up to admission for difficulty breathing. he was found to have multiple infections including rhinovirus and parainfluenza virus on nasal wash, pjp pneumonia, norovirus, and pseudomonal cellulitis of his nose causing significant destruction. although previous laboratory studies revealed a normal absolute neutrophil count (anc), his anc quickly dropped to cells/ul. his igg, iga, and igm were undetectable. while his total b cell count ( cells/ul) was normal, he lacked any switched memory b cells. he had near normal total t cell count (cd cells/ul) and cd count ( cells/ul) and a markedly decreased cd count ( cells/ul) and poor proliferative response to low concentrations of phytohaemagglutinin and pokeweed mitogen. a . cm x . cm x . cm paraspinal mass was found on chest ct, which was subsequently characterized as mibg-avid with a curie score neuroblastoma. metastatic evaluation including bone marrow aspirate and biopsy was negative for malignancy. however, marked granulocytic hypoplasia and maturation arrest were present suggesting severe congenital neutropenia or, less likely, immune-mediated. whole exome sequencing detected homozygous variant of unknown significance in the hells gene (p.m t). he was treated with intravenous immunoglobulin and g-csf with clinical and laboratory improvement. his neuroblastoma was initially observed, then subsequently removed due to a > % increase in size. pathology confirmed mycn non-amplified favorable histology. he remains in remission months after resection. he is currently awaiting bone marrow transplant for his immunodeficiency. conclusions: the significance of this case report is the novel presentation of icf . neutropenia and malignancies have been reported in immunodeficiency-centromeric-instability-facial anomaly syndrome (icf ) but not icf . this case report thus expands upon the clinical picture of icf patients to include neutropenia and malignancies, and further describes the immunodeficiency. associate professor, university of south florida -johns hopkins all childrens hospital introduction/background: identification of newborns with severe combined immunodeficiency using state wide newborn screening (nbs) began in florida in . abnormal results require extensive confirmatory diagnostic testing and prophylactic antimicrobial medications are needed to effectively evaluate and treat the infant. several barriers have been identified within government-sponsored health insurance programs that impede delivery of these evaluations and medications, often resulting in delays and/or inpatient hospitalization in order to provide timely and appropriate care. objectives: determine the cost differential between the initial evaluation and treatment of a scid patient detected by newborn screening in the inpatient versus outpatient setting. methods: the cost utilization of inpatient versus outpatient management of newly identified scid patients from nbs were analysed to include the cost of confirmatory testing and initiation of prophylaxis within the inpatient versus outpatient setting. laboratory tests included assessment of t cell immunity with quantitative and functional assessment, immunoglobulin measurement, genetic testing for scid variants, evaluation for maternal engraftment, and hla typing. medications included ig supplementation, pentamidine, fluconazole, and acyclovir. we compared the actual cost of inpatient stay and inpatient evaluation versus the approximate cost that would have been accrued if the patient were not admitted to the hospital. results: from - , infants with government-sponsored health insurance had abnormal nbs and were confirmed to have scid after evaluation at our institution. all infants were admitted into the hospital for initial evaluation and initiation of appropriate medications for an average of days. total average cost of medication administration for days was $ , , total cost of laboratory testing was $ , , and average inpatient stay averaged $ , per patient. conversely, the cost that would have been accrued in the outpatient setting for medication would have been $ , for days. laboratory testing costs would be no different as an outpatient. in total, the cost for inpatient evaluation was $ , versus $ , as an outpatient. conclusions: standard laboratory assessments and medications are necessary for infants identified with scid by population based nbs. despite government sponsoring of the florida nbs program, unnecessary barriers exist by government sponsored insurers that lead to a delay appropriate care. inpatient admission alleviates these barriers, but significantly increases cost. we advocate that standard ambulatory scid outpatient evaluation and initial treatment be authorized in children identified with scid through nbs without delay. patients with cd g mutations reveal a role for human cd g in treg diversity and suppressive function. introduction/background: integrity of the tcr/cd complex is crucial for positive and negative selection of t cells in the thymus, and for effector and regulatory functions of peripheral t lymphocytes. genetic defects that reduce, but do not abrogate tcr signaling, are associated with a variable degree of immune deficiency and immune dysregulation. in particular, while cd d, cd e, and cd z gene defects in humans present mainly with severe immune deficiency, cd g mutations lead to milder phenotypes, mainly characterized by autoimmunity. however, the role of cd , encoded by cd g, in establishing and maintaining immune tolerance has not been elucidated. objectives: we aimed to investigate abnormalities of treg cell repertoire and function in patients with genetic defects in cd g with evidence of clinical autoimmunity. methods: high throughput sequencing (hts) was used to study composition and diversity of the t cell receptor (trb) repertoire in treg, conventional cd + (tconv), and cd + cells from patients with cd g mutations and in healthy controls. treg function was assessed by studying their ability to suppress proliferation of tconv cells. results: treg cells of patients with cd g defects had reduced diversity, increased clonality, and reduced suppressive function. the trb repertoire of tconv cells from patients with cd g deficiency was enriched for hydrophobic amino acids at position and of the cdr , a biomarker of self-reactivity. overlap between treg and tconv cell repertoires was observed in cd g mutated patients. conclusions: the treg and tconv cell repertoire of patients with cd g mutations is characterized by a molecular signature that may contribute to the increased rate of autoimmunity associated with this condition. introduction/background: a common concern with b cell-depleting therapies is their potential effect on humoral immunity. although there have been reports of prolonged hypogammaglobulinemia in adult patients receiving rituximab, little is know about this phenomenon in children. objectives: we sought to assess humoral immunity in children receiving rituximab and determine risk factors leading to low immunoglobulin levels and infections. methods: we conducted a retrospective study on all pediatric patients ( years) who received rituximab for the first time between january to december in a single tertiary pediatric hospital. charts were reviewed and data was collected prior to rituximab treatment and at , , and > months after treatment. patients who received rituximab after hematopoietic cell transplantation (hct) or for a malignancy and those with an underlying primary immune deficiency (pid) at the time of treatment were excluded. results: in total, patients received rituximab during the study period. of those, were excluded (hct: n= , lymphoma: n= , pid: n= ). sixtyeight patients were eligible. indications for rituximab treatment were renal disease (n= ), neurologic disease (n= ), hematologic disease (n= ), rheumatologic disease (n= ), ebv control (n= ), other (n= ). one patient who died from autoimmune encephalitis days after rituximab was excluded from the follow-up study. at any time after rituximab treatment, low igg was present in / ( . %), low iga in / ( . %), and low igm in / ( . %) of patients. over a year after their last rituximab dose, / ( . %) of patients still had low b cell counts for age, and / ( %) had low memory b cell counts (cd + among cd + cells: mean value = . % +/- . % sd). hospitalisation for infection was required in / ( . %) patients in the year following rituximab treatment, which was associated with having either low igg ( . % vs . %, p= . ) or low iga ( . % vs . %, p= . ), but not with low igm levels ( . % vs . %, p= . ). also, receiving a treatment with more than one rituximab cycle was a risk factor for low igg ( . % vs . %, p= . ). conclusions: hypogammaglobulinemia following rituximab treatment was frequent, and the presence of low igg and iga were associated with a higher risk of serious infection in this context. introduction/background: subcutaneous immunoglobulin (scig) replacement therapy for patients with primary immunodeficiency (pid) is usually administered once a week. however, a variety of dosing regimens can be used to provide flexibility for patients. objectives: we used pharmacokinetic (pk) analysis to evaluate the pk characteristics of weekly and biweekly (once every weeks) scig administration in patients with pid. methods: this pk substudy was part of a prospective, open-label, phase study (nct ) in patients with pid treated with igpro (hizentra®, csl behring, bern, switzerland). a noncompartmental analysis of serum igg concentrations was used to calculate pk parameters and compare pk outcomes on weekly and biweekly dosing. results: of the patients included in the pk substudy, provided samples for both weekly and biweekly regimens. the dose-adjusted area under the concentration-time curve was comparable for both treatment regimens: . and . (h*g/l)/mg for the weekly and biweekly regimens, respectively. the igg clearance was also similar, being . for the weekly and . ml/h for the biweekly regimen. median peak igg concentrations occurred later with the biweekly regimen ( . days) compared to . days for the weekly regimen. igg trough levels were close for both treatment regimens, with arithmetic means slightly lower for biweekly than for weekly regimens, at . vs. . g/l respectively. the minimum igg concentrations within a dosing interval were also comparable, with arithmetic means of . and . g/l for the weekly and biweekly treatment regimens, respectively. conclusions: biweekly and weekly hizentra® administration at the same total monthly igg doses resulted in similar igg exposures. pharmacokinetics, efficacy, tolerability and safety of a new subcutaneous human immunoglobulin . % in primary immune deficiency introduction/background: patients with primary immune deficiencies (pid) require life-long replacement therapy with immunoglobulins (ig) to prevent severe infections and irreversible complications. in addition to safety and efficacy, tolerability and convenience of administration of ig products are essential factors in patient acceptance. a new . % ig preparation (octapharma, lachen) was developed for subcutaneous administration (scig) derived from the established manufacturing process of octapharmas intravenous ig (ivig) brand octagam®. objectives: primary outcome was to assess efficacy of a new . % subcutaneous human immunoglobulin preparation in preventing serious bacterial infections. secondary endpoints included evaluating tolerability and safety, determining the pk profile, the number and rate of other infections and changes in quality of life measurements. methods: a prospective, open-label, single-arm phase study involving patients was conducted at centers in north america and europe. pid patients who were stable on ivig treatment for at least months and with igg trough levels . g/l underwent a -week wash-in/wash-out period consisting of weekly scig doses . times the previous ivig dose (based on published conversion rates for marketed scig products), followed by a -week efficacy period ( scig infusions in total). of patients enrolled had complete pharmacokinetic assessments at different time points: before the switch from ivig to scig (pkiv), after the wash-in/wash-out phase (pksc ) and at week of the efficacy period (pksc ). results: patients (age: - years; mean age . years; . % female) receiving a total of , scig infusions ( . g/kg/week in young children ( years and < years of age) and . g/kg/week in adults; average overall: . g/kg) were included in the full analysis sets. no serious bacterial infections were recorded. among the other infections observed during the efficacy period only one infection was graded as severe (bronchiolitis due to rsv virus), which led to hospitalization ( days). all other infections were mild ( . %) or moderate ( . %) in intensity. infection rate per person-year was . . of the reported adverse events, only were assessed as being related to the study drug; all of these events were non-serious. five non-study drug related serious adverse events were reported in patients ( . %). serum igg trough levels were nearly constant during the study with a minimum trough level of . g/l and mean trough plasma concentrations of . ± . g/l and . ± . g/l for pksc and pksc . median igg trough levels after scig treatment were . to . g/l higher compared to ivig treatment prior to enrollment. a dosing conversion factor (dcf) of . was determined by auc (area under the curve) measurements, allowing dose adjustment to achieve bioequivalence between ivig and scig dosing. improved quality of life measurements, utilizing sf- v , were observed in both physical and mental health parameters when compared from the first to last scig infusion. conclusions: this study demonstrated that the new subcutaneous human normal immunoglobulin . % is well tolerated, safe and effective in patients with pid. introduction/background: atopic dermatitis (ad) is a chronic, relapsing, inflammatory skin disorder with associated pruritus that affects percent of children in the united states. severe atopic dermatitis refractory to conventional therapy can be concerning for an underlying immunodeficiency, especially in infants. increased incidence of hypogammaglobulinemia has been associated severe ad. a handful of cases describe a correlation with transient hypogammaglobulinemia of infancy (thi), but a thorough immunological evaluation is often missing to further understand this relationship between ad and characterization of thi. objectives: define various phenotypes of thi with their clinical presentation and laboratory findings. compare thi vs. thi associated with severe a.d. methods: a case series of six patients was conducted at a single academic center from / / to / / for patients with severe atopic dermatitis with low igg levels. all available immunological laboratory data were retrospectively collected during this time period. descriptive statistical analysis was utilized for data comparison. results: of the six patients, four had no infectious history. of the remaining two, one had recurrent skin abscesses associated with his poorly controlled atopic dermatitis requiring oral antibiotics and one patient had two episodes of staphylococcus aureus superinfection of the eczema. at the time of presentation, the mean age was months with mean igg of mg/dl and mean ige of , ku/l. iga and igm were within normal age cut offs. all patients had normal protein and polysaccharide specific antibody titers after completion of vaccination series. mean cd + count was , /ul with normal cd +, cd +, cd + and cd + cell counts. three patients were tested for lymphocyte mitogen proliferation and complement function which were normal. two patients, who were tested, had normal phagocyte work up. mean age of igg improvement to igg> mg/dl, was months. patients had total protein and albumin levels with mean . ku/l and . ku/l, respectively which eventually normalized. four patients improved with skin care and dietary modification to hypoallergenic formula. one patient improved with extensively hydrolyzed formula and three patients improved with amino acid formula. one patient improved with aggressive skin care. one patient, who was noncompliant with dietary recommendations and aggressive skin care, did not improve. conclusions: one prospective study described an increased incidence of hypogammaglobulinemia in patients with atopic dermatitis compared to controls regardless of the severity. no further prospective studies have characterized hypogammaglobulinemia in this population. according to the primary immunodeficiency practice parameters, children with thi often present with frequent viral and bacterial respiratory illnesses, low igg levels and normal vaccine responses. in one study, the period of hypogammaglobulinemia spontaneously corrects to normal by mean age of months with all patients reaching normal levels by months. these patients may have low igm or iga and decreased t or b cells, which eventually normalize. management is often with antibiotic prophylaxis and if refractory to prophylaxis or unable to tolerate, igg administration (igrt) based on severity of symptoms is recommended. we describe a less severe variant of thi associated with severe atopic dermatitis and characterized by very transiently decreased igg levels with earlier resolution than typical thi, normal igm and iga levels, normal specific antibody levels and normal t, b and nk cells. these patients typically do not manifest recurrent viral or bacterial respiratory illnesses. they also do not require antibiotic prophylaxis or igrt. severe atopic dermatitis maybe a positive prognostic indicator for patients with thi and is associated with an earlier self-resolution of hypogammaglobulinemia, lack of typical infections, normal iga, igm and normal t and b cell numbers. a possible mechanism for thi associated with severe ad may be transdermal loss of protein which was supported with mildly low total protein and albumin levels rather than immaturity of the immune system in patients with true thi. introduction/background: in recent years it was found that heterozygous mutation in pik cd gene produces an autosomal dominant primary immunodeficiency characterized by onset of recurrent sinopulmonary and other infections in early childhood with defects in both b-and t-cell populations and a special susceptibility to uncontrolled viral infections. many patients develop chronic lymphoproliferation and there is also an increased susceptibility to b-cell lymphomas. here we present a patient assumed as a common variable immunodeficiency with heterozygous mutation in pik cd gene. objectives: to describe a case of a patient assumed as a common variable immunodeficiency with heterozygous mutation in pik cd gene. results: year old woman with personal history of severe and recurrent upper and lower respiratory infections, chronic pulmonary disease with bilateral bronchiectasis, chronic diarrhea without diagnosis, mild osteopenia, focal lesion in right hepatic lobe, atopic dermatitis and anemia. she was following up in other center and in she was diagnosis with common variable immunodeficiency (cvid) and started treatment with intravenous immunoglobulin (ivig), but she referred low adherence to it. she did not referred history of lymphoproliferation nor significant viral infections. she has a daughter with spherocytosis who required esplenectomy and also had bronchiectasis and cvid, she deceased at years old because pulmonary infection. other daughter and sons referred healthy. in our first immunologic studies we found severe hypogammaglobulinemia (igg mg%, no dosable iga and igm) with absent of b cells in peripheral blood. we started with high doses of ivig ( mg/k/month) and antibiotic prophilaxis with improvement of the functional respiratory test and without new infections. we are planning colonoscopy to study her chronic diarrhea. thinking that her clinical picture could be other than cvid we order a genetic study. a nextera exome capture and next generation sequence with illumina hiseq was made and an heterozygous mutation in pik cd gene (chr : . . , p.pro ala) was found. family and functional studies are still pending. conclusions: due that clinical presentations of primary immunodeficiencies are becoming more complex, its diagnosis is a challenge for immunologist now a days. studies with next generation sequence is a very useful tool in indefinite cases, especially when more than one member in the family are involved. chief, laboratory of clinical immunology and microbiology, national institute of allergy and infectious diseases, national institutes of health introduction/background: heterozygous gain-of-function mutations in pik cd as well as heterozygous pik r mutations that affect interaction of p with p lead to constitutive hyperactivation of the pi k pathway and cause activated pi k delta syndrome type or type (apds , apds ), respectively. we describe a female with apds with short stature, diffuse lymphadenopathy, recurrent upper respiratory tract infections, elevated igm, persistent ebvand cmv viremia and disseminated toxoplasmosis who gave birth to a genetically affected daughter with severe congenital toxoplasmosis. objectives: to characterize the molecular and cellular defects underlying severe toxoplasmosis in this family methods: investigation of the molecular basis of the disease was performed through whole exome sequencing, and results were validated by sanger sequencing. functionality of the pi k pathway was assessed by analyzing akt and s phosphorylation in freshly isolate b cells with and without stimulation with anti-igm. an excisional lymph node biopsy from the affected mother was stained with anti-pd antibody to detect t follicular helper cells, and with igm and igg specific antibodies to analyze the proportion of isotype-specific b and plasma cells results: whole exome sequencing of maternal dna with targeted analysis of pid genes identified a heterozygous mutation at an essential donor splice site of pik r (nm_ . :c. + g> a). sanger sequencing confirmed the presence of this mutation in both the mother and her child. functional studies on b cells freshly isolated from both patients confirmed an increase in baseline akt and s phosphorylation, suggesting constitutive activation of the pi k-mtor signaling pathway. a lymph node biopsied from the mother contained numerous pd- + tfh cells and igm+ plasma cells conclusions: toxoplasma gondii is an obligate intracellular parasite that is usually only symptomatic in immunocompromised hosts. severe toxoplasmosis has been reported in the following primary immunodeficiencies: cd ligand deficiency, tap deficiency, cvid, nfkb deficiency, and immunodeficiency due to anti-ifn-autoantibodies. importantly, toxoplasma infection was previously reported in a month-old infant with apds , and ocular involvement has been described in a -year-old patient with apds . this, however, is the first report of systemic and severe congenital toxoplamosis in a mother and child with apds. to evade innate host defenses, t. gondii induces the activation of the pi k/akt signaling pathway, reducing intracellular reactive oxygen species and creating an intracellular environment that is hospitable to parasite survival and proliferation. therefore, we postulate that the pik r mutation may lead to a hospitable cellular environment for toxoplasmosis replication. this work was partially supported by the division of intramural research, niaid, nih (protocol # -i- ). additional authors of this work are: ottavia delmonte and kerry dobbs, from the laboratory of clinical immunology and microbiology, niaid, nih. introduction/background: aplaid (autoinflammation and plc-gamma- -associated antibody deficiency and immune dysregulation) is a term that was proposed for the newly discovered autoinflammatory condition resulting from a pathogenic missense variant, ser tyr, in the cterminal sh (csh ) domain of plc-gamma- in order to distinguish it from plaid, a distinct clinical entity that results from intragenic deletions of portions of the csh domain of the same protein. plc-gamma- is a phosphodiesterase that is predominantly expressed in hematopoietic cell lines and acts on pip to produce ip and dag in the pkc and ras/raf/ erk pathways. the only formally published case of aplaid described a father-daughter pair with an autoinflammatory clinical syndrome affecting the skin, mucosa, eyes, pulmonary and gastrointestinal systems. objectives: to describe the aplaid phenotype resulting from a novel genetic variant of the phosphodiesterase plc-gamma- in two unrelated families methods: clinical review and case presentation results: we report a -year-old female with a long-standing history of recurrent pneumonia, cellulitis and cystitis with obstructive lung disease characterized as bronchiolitis and dynamic airway collapse, with negative alpha- -antitrypsin testing. she had a history of childhood onset granuloma annulare and pressure-induced urticaria, as well as episcleritis. immune testing revealed low igm ( mg/dl), elevated baff levels and a low percentage of cd + memory b-cells, prompting sequencing of plcg , which identified a c. t>a, p.met lys variant in the calcium binding c domain. her -month-old daughter, who has a history of bullous skin lesions, failure to thrive, febrile episodes and recurrent respiratory infections was likewise found to have this plcg variant. like her mother, the daughter also has low igm and elevated baff levels. similar symptoms of recurrent sinopulmonary infections and hypogammaglobulinemia, have been described in an unrelated family that shares this same plcg variant in a doctoral thesis by rozmus. this work also describes functional analysis, in which this particular variant of plc-gamma- was demonstrated to have dysregulated plcgamma- activity leading to aberrant intracellular calcium signaling and increased apoptosis of immature b-cell subsets. conclusions: we describe our experience in evaluation and treatment of this family with a previously undiagnosed disorder. together, these new cases add to the expanding body of knowledge regarding plc-gamma- and its importance for the development of autoinflammatory and primary immunodeficiency conditions. ( ) submission id# present a case.non-celiac gluten sensitivity is an emerging entity with symptoms similar to celiac disease, but without positivity in specific diagnostic tests. it is considered more common than celiac disease patients with sigad have a greater risk of concomitant autoimmune disorders than health individuals. sigad was previously to be associated with celiac disease, but not usually in non -celiac gluten sensitive. objectives: describir a case ataxia non celiac gluten sensitivity, methods: clinic case description. results: he patient has negative serology test for gluten, but clinically respond as celiac disease. conclusions: all patient has negative serology test for gluten, but clinically respond as celiac disease, could has non-celiac disease, this case described here ,it suggest than this entity should to thought before than the celiac disease, since not-celiac disease is it more common. associate professor, federal university of rio de janeiro introduction/background: primary cutaneous actinomycosis is a rare condition caused by gram-positive filamentous bacteria and generally occur after traumatic inoculation. objectives: to report an unusual etiology of skin lesions in a patient under anti-tnf-alpha therapy. methods: we report the case of a -years-old female receiving conventional doses of adalimumab for ankylosing spondylitis who presented, two weeks before referral for dermatological assessment, with an erythematous nodule with purulent discharge on right pretibial region and pruritic erythematous plaques with scaling and peripheral pustules in the trunk and nose tip. results: a fungal etiology was suspected and scraping specimens from several cutaneous lesions were submitted to direct microscopic examination and culture, which were negative for fungi. cutaneous biopsy of the pretibial lesion was sent for histopathology and microbial cultures. adalimumab was interrupted and empirical therapy with oral terbinafine ( mg/day) was started with close clinical follow-up. a folliculitis reaction pattern without granulomas was observed during histopathological examination and gram-positive cocci were isolated from biopsy sample and further identified as saccharopolyspora sp. by molecular typing. sulfamethoxazole+trimethoprim was added and discontinued after one week due to a cutaneous rash. terbinafine ( mg/day, p.o) was used for months with complete clearing of all skin lesions and very good tolerability. spondyloarthritis signs and symptoms were unremarkable and no anti-inflammatory or immunomodulating treatment was necessary. conclusions: increased risk of skin actinomycotic infections in patients under anti-tnf therapy is not consistently reported in the literature. nevertheless, they should be included as a differential diagnosis of atypical skin lesions in individuals under anti-tnf therapy. where we aim to determine the prevalence, incidence, characteristics, treatment and outcomes of primary immunodeficiency disease (pid) patients in qatar. pids are rare heterogeneous disorders of the immune system that result in an increased susceptibility to infection, immune dysregulation and occasionally, to cancer. objectives: determine the range of pids with important epidemiological data in qatar after analyzing the database and creating a registry. methods: this is a retrospective study of pid patients followed at hamad medical corporation from to using medical records. all patients who were diagnosed with a pid irrespective of age were included. patients were classified according to the international union of immunological societies expert committee on pid. the data is captured under sectionsa) patient demographics including age, gender, ethnicity b) clinical presentation, c) immunodeficiency profile including age at diagnosis, type of immunodeficiency, family history d) treatment modality and e) lab/genetic data. results: we registered patients ( females and males) over a span of four years. mean age at onset, diagnosis and diagnostic delay were . , . and years respectively. majority of the patients were arabs % followed by people from the asian subcontinent %. antibody deficiency was seen in %, immune dysregulation %, well defined immunodeficiency (at, hige, digeorge,wiskott aldrich) % and t/b cell cid %. rare diagnoses (ipex, msmd) were recorded whereas no cases of toll like receptor and complement deficiency were seen. consanguinity rate was (n = , % = ) and first degree cousin marriage (n = , % = ). family history was positive in % (n= ) of the patients. maximum diagnostic delay was seen in scid ( % > months) and agammaglobulinemia ( % > months).during the patients life, infection was the most common presenting complaint ( %) followed by sinopulmonary disease ( %) and gi-tract manifestations ( %). the most common infections were pneumonia ( %), otitis media and conjunctivitis ( % each) followed by failure to thrive ( %) and sepsis ( %).microbial isolates particularly seen as causative agents of infections were p.aeruginosa and salmonella ( %) each, mrsa and e.coli ( %) each. a genetic defect was confirmed in % of ataxic telangiectasia and % of scid patients. active infections were treated and prophylactic antibiotics were prescribed in cases ( %). prophylactic antibiotics were prescribed to % of patients with immune dysregulation and to % of well-defined syndromes. out of patients who had hsct, % had successful transplants. ivig was given to % of the total pid patients with humoral immunodeficiency patients receiving the most ( %). one patient had gene therapy and two required interferon gamma treatment for msmd. mortality rate at st year of life was % whereas total mortality rate was %, excluding cases that passed away before pid was diagnosed. conclusions: the estimated prevalence of pid in qatar is found to be per . over the years, physicians have become increasingly aware of pid and survival rate has improved. initiation of newborn screening for scid and agammaglobulinemia will lead to earlier diagnosis and initiation of therapy with better outcomes. introduction/background: agammaglobulinemia is typically associated with a near absence of b cells secondary to a developmental block in bone marrow, and, usually but not always, manifests in early life. most of such patients are males with x-linked agammaglobulinemia (btk deficiency). females with agammaglobulinemia, of either autosomal recessive or dominant inheritance, are rare. objectives: we present and discuss the differential diagnosis for the conflicting clinical and immunological phenotypes of two adult females who present with infections, cytopenias, and agammaglobulinemia with low to normal b cell count. methods: retrospective chart review of clinical and laboratory data results case : a -year-old female who, at age years, had evans syndrome (autoimmune thrombocytopenia and hemolytic anemia) that resolved after steroid and high-dose gammaglobulin treatment. at age years, immunologic workups revealed a complete absence of serum immunoglobulins (igg, iga, igm) and low b cells ( %) (normal range - ). however, three years prior, patient had detectable igm ( mg/ dl). b cell subset analysis showed an expansion of cd hi lo b cells ( %, normal . - . %), with a marked decrease in switched memory b cells ( . %, normal - %). additional immunophenotyping revealed a reduced frequency of naïve cd + ( . %, normal > %), cd +( %, normal > %) t cells, and normal lymphocyte proliferative responses to mitogens and anti-cd , anti-cd /anti-cd , and anti-cd /il- . case : a -year-old female with recurrent upper respiratory tract infection, undetectable igg, iga, igm and ige, and no response to vaccinations (tetanus and pneumococcal). thrombocytopenia ( x count/microliter) was noted once during her thirdtrimester pregnancy without evidence of pre-eclampsia. postpartum cd + t cell count was low ( cell/microliter, normal - cell//microliter). immunophenotyping revealed normal b cell count with reduced frequency of naïve cd + ( . %, normal > %) and cd + ( %, normal > %) t cells. conclusions: we report two adult females who present in early adulthood with recurrent infections and cytopenias. both had agammaglobulinemia and decreased naïve t cells suggestive of late-onset combined immunodeficiency (locid). the presence of peripheral b cells makes autosomal recessive defects in b cell receptor signaling (lamda , iga, igb, igm, blnk) less likely. differential diagnosis of locid with cytopenias includes ctla- haploinsufficiency, gain-of-function pik cd mutations, ikaros defects and autosomal recessive rag deficiency. both patients remain at risk for developing autoimmune complications. a molecular diagnosis will facilitate targeted therapy for the underlying defect. professor, director of the laboratory of childhood immunology, ku leuven introduction/background: complement factor properdin (cfp) is a soluble glycoprotein which has a unique known role as a positive regulator of the alternative complement pathway by binding and stabilizing the inherently labile c /c convertase enzymes. mutations in the cfp gene lead to aberrant protein expression or to expression of a dysfunctional protein which results in high susceptibility to pyogenic infections especially neisseria meningitidis. properdin-deficient individuals are at greater risk of fulminant meningococcal disease, with mortality rates as high as %. objectives: case report: a year old belgium boy from nonconsanguineous parents presented with achronic purulent cough and recurrent infections of upper and lower airways. he suffered from a severe pneumonia at the age of . ct thorax showed bronchiectasis of the right middle lobe and left lower lobe. in addition, he had recurrent acute otitis media since the age of year old. immunological work-up showed an absent ap activity, with normal ch , c and c . results: genetic and functional analysis: sanger sequencing of cfp gene identified a hemizygous c. t>g, p.y g (cadd score . , msc_cadd . ) mutation in the patient and his mother. properdin elisa (hycult biotech) showed absent and % of the normal healthy control value of serum properdin concentrations in patient and the healthy mother, respectively. conclusions: conclusion: we diagnosed properdin deficiency in a y old boy presenting recurrent lower and upper respiratory tract infections. ap testing, added to ch , should therefore be part of initial workup for patients with recurrent severe respiratory tract infections. indeed early diagnosis allows for appropriate prolonged antibiotic prophylaxis and immunization to reduce the risk of fatal meningococcal disease, to reduce or prevent organ damage and to allow for genetic counselling in the family. - , - , - and - ) and child depression inventory (cdi) were implemented to both children and parents in addition to sociodemographic data form; beck depression inventory (bdi), beck anxiety inventory and zarit caregiver burden scale were implemented only to parents. results: the depression inventory parent and child form values of the patient group with primary immunodeficiency were significantly higher than the control group (p= , and p= , ). according to beck depression and anxiety inventories, it was seen that the depression and anxiety inventory scores of the parents of the patient group were higher than the control group (p= , and p= , respectively). it was determined that the cdi parent form scores of the patients with hospitalization history were statistically higher than the patients without hospitalization history ( , ± , and , ± , ; p= , ). bdi scores were significantly higher in the group which received ivig (p= , ). while the quality of life of the patients compared to their parents was perceived as worse than the healthy children in all dimensions (p= , p< , and p< , ), the quality of life of the children was worse only in psychosocial and total quality of life fields (p< , p= , and p= , ). it was seen that quality of life of children physical health scores of only the patients with hospitalization history were statistically significantly lower ( , ± , and , ± , ; p= , ) . while no statistically significant difference was found in terms of quality of life of the child scores between the groups which received and did not receive ivig replacement treatment, psychosocial and total qualify of life scores of parents were statistically significantly lower in the group which received ivig replacement treatment (p= , p= , ). zarit care giver burden scale scores were similar in patient and control groups. although both groups were on the limits of mild to moderate caregiving burden, it was seen that the scores of the group which received ivig were significantly higher than the group which did not receive ivig (p= , ). conclusions: as a conclusion, we think that it will be appropriate to inform and monitor the entire family in relation to psychosocial difficulties and care giving burden they may experience in time as well as the medical aspects of the disease in order to develop a holistic approach to children with primary immunodeficiency. introduction/background: chronic lung disease is the most common complications of cvid, affecting - % of patients. it includes bronchiectasis affecting % of patients and granulomatous lymphocytic interstitial lung disease (glild) in to %. both are associated with an increased morbidity and mortality. pulmonary functional studies and ct scans have been proposed as screening procedures for lung involvement in cvid. the definitive diagnosis of glild is established histologically, but it is too invasive in patients with radiological abnormalities and no symptoms. objectives: we aim to describe the pulmonary complications of our cohort of patients with cvid comparing them with subjects affected with other types of hypogammaglobulinemia. methods: we reviewed all clinical records of the patients with a diagnosis of hypogammaglobulinemia of any cause until december . we looked at all pulmonary function tests (pft), -minute walk tests and ct scans performed. we classified patients according to the ct scan pulmonary disease pattern. we compared the demographic data and pulmonary characteristics of each group and intended to describe similarities and differences between them. results: we collected patients and ct scans from patients. patients were grouped for further analysis as follows: no ct performed: ; normal ct: ; bronchiectasis: ; glild: and bronchiectasis + glild: . eight patients ( %) had no ct including six with cvid, one with x linked lymphoprolipherative disease (xlp) and one had a syndromic deletion in chromosome . the median age of symptom onset was . yo, with a median delay to diagnosis of year. nine subjects ( %) had ct scans with no chronic disease, six had cvid, rituximab induced hypogammaglobulinemia (rih) and mgus with hypogammaglobulinemia (mguswh). the median age of symptom onset was years old with a median delay to diagnosis of years. nine patients ( %) presented persistent bronchiectasis, with cvid, with an igg deficiency (sigg d) and with xlp. median age at symptoms onset was yo and median delay to diagnosis years. seven patients ( %) had glild; all afected with cvid. median symptoms onset was years old, and median delay to diagnosis was years. one patient ( %) was included in the bronchiectasis and glild group. she was referred with a diagnosis of rih, in the context of a pulmonary malt lymphoma. however, her ct did not show a typical ct lung lymphoma pattern, lymphocyte monoclonality was never shown and she had a reduced pre-treatment low gamma globulin concentration and a history of respiratory infections since adolescence, we believed her diagnosis is cvid. pft with an obstructive pattern identified patients with bronchiectasis (p< . ) and patients with glild had a significantly lower basal and post minute walk o saturation (p< . , n= ) conclusions: using a systematic approach, we identified that roughly % of patients with hypogammaglobulinemia have chronic pulmonary ct abnormalities. half of them had bronchiectasis, that were associated to hypogammaglobulinemia of any cause, a longer disease course and a reduction in fvc and fev with a shorter distance reached in the minute walk test. the other half had glild, spirometries in this group were useless, but o saturation was significantly lower, basal and after the minute walk test. we found a high frequency of pulmonary disease in our cohort and a disease progression study is now in place. hies) is a primary immunodeficiency characterized by eczema, sinopulmonary infections, and musculoskeletal and vascular abnormalities. care of patients with this disease is largely supportive with the use of prophylactic antibiotics and topical eczema therapies. the use of replacement immunoglobulin is increasing. hematopoietic stem cell transplant (hsct) is being considered more frequently, but many questions remain regarding which patients should undergo transplant. as pulmonary complications are a leading cause of morbidity and mortality in this disease, and potentially improved with replacement immunoglobulin and hsct, we sought to examine more closely the patients with more frequent pulmonary hospitalizations and structural lung disease. objectives: to determine the rates of pulmonary complications in our large cohort of ad-hies patients, and examine the relationship between immunologic markers and pulmonary disease. methods: we retrospectively reviewed the records of ad-hies patients seen more than one time at nih between and . there were pediatric patients under the age of years. we reviewed the number of and cause for hospitalizations, and reviewed radiology reports for structural lung disease including bronchiectasis and pneumatoceles. we correlated these findings with specific antibody responses and lymphocyte phenotyping, such as the number of memory b lymphocytes. results: the patients range in age from years to years, with a median age of years. patients had chest cts, with percent having bronchiectasis, and percent having cavitary lesions. percent of patients had both. these abnormalities were more prevalent in patients with low memory b cells. percent of patients receive replacement immunoglobulin. in patients not receiving replacement ig, percent had appropriate response to pneumococcal vaccine. during this time period, there were hospitalizations, the majority of which were associated with pulmonary infections. the rate of overall hospitalization was higher in the group with low memory b cells (p= . ), but there was no difference associated with age. conclusions: immunologic abnormalities may assist in determining the long-term prognosis of patients with ad-hies, and can be considered in management plans such as the use of immune globulin and consideration for hsct. introduction/background: purine nucleoside phosphorylase (pnp) deficiency is an autosomal recessive disorder affecting the purine salvage pathway causing a (severe) combined immunodeficiency disorder, autoimmunity, and neurological symptoms. patients typically present in the first few years of life with frequent infections and a failure to thrive. decreased plasma levels of uric acid are suggestive, while neutropenia is a rare complication. prognosis is generally poor with few patients making it to adulthood without treatment. treatment options are limited to general supportive care and hematopoietic cell transplantation. methods: we present the cases of three patients, one sister and two brothers, from a consanguineous french canadian family. they presented in early adulthood with nearly identical histories of repeated pneumonias and chronic rhinosinusitis. one patient had necessitated filgrastim to treat a parainfectious neutropenia. they did not report any neurological abnormalities or symptoms of autoimmunity. results: a primary immune deficiency was suspected. initial evaluation showed normal immunoglobulin levels, a lack of response to vaccination, positive ebv ebna iggs, and auto-antibodies. the patients were severely lymphopenic with reduced numbers of t, b and nk cells ( l: , cd + %, cd + %, cd + %, cd + %, cd -cd + %). in particular, they had a severe depletion of naive t-helper recent thymic emigrant cells (cd +cd ra+cd + %). additional studies revealed increased urinary inosine, guanosine, deoxyinosine, and deoxyguanosine, severely reduced erythrocyte pnp activity ( % residual activity), and a normal uric acid level. a homozygous, missense mutation, c. c>g (p.his asp) was found and described as pathogenic in silico. the diagnosis of pnp deficiency was made; tmp-smx prophylaxis and ivigs were started. conclusions: theses cases are atypical for a number of reasons. the first is the relatively benign immunological course, the lack of neurological symptoms, and the advanced age at the time of the diagnosis, possibly due to residual enzymatic activity. in addition, pnp deficiency has been classically described as causing a marked reduction of t cells with a relative sparing of b cells. however, b cell lymphopenia have been reported, particularly with latter presentations. while this specific mutation had been identified once previously in the literature, as a compound heterozygote, these are the first confirmed cases of homozygotes. the pathogenicity of the mutation was not only suggested in silico, it was confirmed biochemically. in addition, in vitro functional studies have demonstrated the importance of his for the binding of pnp to its substrate, particularly in the formation of the early transition state. indeed, pnp (p.his asp) has been shown to have a greatly reduced affinity for its substrates and catalytic activity. further study is needed to identify the optimal management for these patients. introduction/background: whole exome sequencing has become an integral part of diagnosis and treatment of rare immunodeficiency diseases. one of these diseases is tetratricopeptide repeat domain a(ttc a) mutations; a rare disorder associated with multiple intestinal atresias and severe combined immunodeficiency. histologic assessment of organ biopsies of patients with ttc a deficiency suggest it may play a role in multiple organs as it is expressed in the cytoplasma of intestinal, thymus and pancreatic cells. many patients with ttc a deficiency have moderate to severe combined immunodeficiency. nine patients described in the literature with ttc a mutations have been treated with hematopoietic stem cell transplant, mostly at a young age. objectives: this case report describes a rare presentation of an adolescent with ttc a deficiency, her clinical presentation, immune evaluation and treatment with eventual referral to bone marrow transplant at the age of . methods: retrospective chart review under irb approval was performed. whole exome sequencing performed at baylor texas childrens hospital and mutations were confirmed by sanger sequencing. extensive immune and nk cell phenotyping were performed by flow cytometry and nk cell function was tested using standard cr release cytotoxicity assays. results: a year old female was referred to immunology for severe refractory warts, history of severe diarrhea with epithelial dysplasia but no atresia, failure to thrive requiring parenteral nutrition until years old and multiple infections with staphylococcus aureus and candida. at years of age, she developed rapidly worsening pulmonary function that improved with monthly pulse methylprednisone. her immune phenotype demonstrated a combined immunodeficiency including severely low cd , cd , b and nk cells. she had no igd-cd + memory b cells and undetectable igg, iga and igm, isohemagglutinins and vaccine titers to diphtheria and tetanus. her mitogen proliferation response was low and she had abnormal tcr v-beta repertoire. she was referred to the texas childrens hospital center for human immunobiology for the nk cell evaluation and reasearch clinic (near) the patient was found to have impaired nk cell lytic function and terminal maturation. this was demonstrated by the decreased frequency of cells that matured to the cd dim subset and was accompanied by decreased expression of the lytic effector molecule perforin and the fc receptor cd . this phenotype was conserved when we isolated cd + hematopoietic precursors and performed nk cell differentiation in vitro. whole exome sequencing demonstrated a compound heterozygous mutation in the ttc a gene, including a c. + _ + aagt mutation, which has been previously described as a pathologic founder mutation in the french canadian population. the second mutation is a previously unreported c. g>a (p.e k), a variant of unknown significance. mutations were confirmed by sanger sequencing, and parents are carriers of the mutations. she was referred for hematopoietic stem cell transplantation with a / unrelated donor option. conclusions: ttc a deficiency is a genetic mutation associated with high morbidity and mortality leading to gut atresia and dysfunction in combination with severe immunodysfunction. we have described a unique case of ttc a deficiency with a novel mutation which is associated with intestinal epithelial dysplasia with no atresias and a late presentation and evolution of combined immune deficiency. nk cell assessments show significantly impaired terminal maturation suggesting a critical role for ttc a in human nk cell development. introduction/background: ras-associated autoimmune leukoproliferative disorder (rald) is a rare condition with significant overlap of clinical and laboratory findings with malignant disorders, notably juvenile myelomonocytic leukemia (jmml) and chronic myelomonocytic leukemia (cmml), but with much better prognosis without specific therapy. we report a case of rald in a month-old male infant originally suspected to have jmml. results a -month-old male infant, born to an unrelated ethiopian father and turkish mother, presented with splenomegaly and leukocytosis ( , /l), monocytosis ( , /l), and thrombocytopenia ( , /l); hemoglobin was gm/dl and fetal hemoglobin concentration . %. family history indicated early death of three paternal uncles, two at age - months and one at age years. jmml was suspected and bone m a r r o w a sp i r a t i o n w a s p e r fo r m e d ; h ow e v e r, r e s u l t s o f immunophenotypic testing with flow cytometry analysis and cytogenetic testing with fish did not support this diagnosis. patient had a normal b level. serum igg ( . mg/dl [nl . - . mg/dl]) and igm ( . mg/dl [nl . - . mg/dl] levels were elevated and iga was normal. lymphocyte subset analysis showed % cd [nl - %], % cd [nl - %], % cd [nl - %] positive cells with normal percentages of cd and cd /cd cells; there were no cd -/ cd -t cells detected. given the findings of leukocytosis, monocytosis, splenomegaly, hypergammaglobulinemia, and a dearth of cd -/cd -t cells, the patient appeared to meet criteria for rald. mutation testing with next generation sequencing showed an nras missense mutation [c. g>t; pgly cys] in % of t cells and % of myeloid cells. the presence of the nras mutation, in the absence of other typical jmml findings or rasopathy syndrome features, supported the diagnosis of rald. there was no therapeutic intervention and, at one year of age, the patient was reported to remain clinically well; he relocated to ethiopia with his parents. conclusions rald is a nonmalignant clinical syndrome originally classified as a subtype of autoimmune lymphoproliferative syndrome (alps) but subsequently distinguished to be a separate entity due to lack of double negative t-cells, a mutation in fas/fasl/caspase- , or consistently elevated serum b levels. though also present in % of jmml patients, a somatic mutation in ras signaling protein (kras or nras), which controls b-cell tolerance and production of autoantibodies, is present in all reported cases of rald. while all previously reported cases of rald had somatic kras or nras mutations, our patient's nras mutation was present in more than % of t cells and myeloid cells, suggesting a heterozygous germline mutation; this has not previously been reported in rald and needs to be further confirmed ( ) the primary objective of this study was to evaluate hct outcomes in patients with was who underwent hct since in north america. we hypothesized that survival after hct from alternative donors such as cord blood has improved compared to published results, but that overall survival would be superior in patients who undergoing hct at a young age. methods: patients were enrolled on pidtc protocol , a multicenter retrospective natural history study of patients treated for was in north america since . clinical features, disease status, hct type and post-hct outcomes were analyzed in patients who underwent hct at pidtc centers between - . descriptive statistics such as median and range for continuous variables and counts and percentages for categorical variables were used to summarize characteristics of the study population. in addition, kaplan-meier curves were used for estimating survival probabilities results: diagnosis of was was confirmed by an expert review panel for eligibility. mutation in the was gene was available for patients, including nonsense (n= , %), frameshift (n= , %), missense (n= , %), splicing (n= , %), gross deletion (n= , %), in-frame deletion (n= , %), pr complex (indel) (n= , %). donor types included matched sibling (n= , %), unrelated adult volunteer bone marrow or peripheral blood stem cells (n= , %), umbilical cord blood (n= , %) and other related donors ( each, phenotypically matched related, haploidentical) . median age at time of hct was . months (range, . . months). the vast majority of patients received busulfan containing conditioning regimens ( %), with some receiving other myeloablative ( %) or reduced intensity regimens ( %). with a median follow-up of . years, overall survival was excellent with -year and -year survival probabilities of % ( % ci, - %) and % ( % ci, - %), respectively. survival was similar at year for recipients of hct from matched sibling ( %, % ci, - %), matched unrelated donor ( %, % ci, - %) or umbilical cord blood ( %, % ci, - % see figure) . importantly we confirmed that survival at year was better in patients who were < years old (n= ) compared to those who were years old (n= ) at the time of hct ( % versus %, respectively p= . ). this difference persisted when only unrelated donor recipients were analyzed ( % vs %, p= . ). overall the percentage of patients in our study who underwent hct at a young age was high ( %) compared to the literature ( % in moratto et al, , blood) . the rate of second hct was only % (n= ). cumulative incidence of acute grade - at days, acute grade - at days and chronic graft-versushost disease at year were % ( % ci - %), % ( % ci - %) and % ( % ci - ), respectively. conclusions: outcome of hct for was since shows excellent overall survival for all donor types, including umbilical cord blood with very low rates of second hct. importantly, hct at a younger age (< years old) continued to be associated with superior survival supporting the provision of hct earlier in the course of the disease. further analysis of the complete cohort is planned to determine whether age at hct has decreased in the modern era compared to pre- and to analyze factors associated with platelet and immune reconstitution, donor chimerism and autoimmunity. introduction/background: comel-netherton syndrome is a rare disease hallmarked by congenital ichthyosis, atopy, trichorrhexis invaginata (bamboo hair) and, within the past years, has been defined as a primary immunodeficiency. specific, rare genetic polymorphisms (c. t>a, pl h) in the fc receptor iiia (fcgr a) on natural killer (nk) cells have been shown to decrease nk cell function. patients with these defects present with susceptibility to severe and recurrent viral infections, however, not all fcgr a mutations result in this clinical phenotype. here we report on a child with a mixed genotype, presenting with congenital ichthyosis, atopy and recurrent bacterial and viral infections. methods: immunophenotyping of lymphocyte subpopulations were evaluated by flow cytometry. genomic dna was sequenced using next generation sequencing. all detected variants were then sequenced using sanger sequencing. cd dual epitope assay and evaluation of nk cell maturation was also performed. intravenous immunoglobulin replacement therapy was initiated. results: our patient presented at months of age for evaluation of food allergy. she has congenital ichthyotic erythroderma with pruritic atopic dermatitis-like skin eruptions and a history of hypernatremic dehydration immediately following birth, thin and easily broken hair and a history of failure to gain weight with appropriate catch-up growth following formula fortification. she has had multiple episodes of acute otitis media and recurrent upper respiratory tract infections associated with wheezing. t-, b-and nk cell quantitation by flow cytometry was normal, including naïve and memory b cells. immunoglobulins and vaccine antibody levels to haemophilus influenza type b, tetanus and streptococcus pneumoniae were normal. lymphocyte proliferation to mitogens was normal. natural nk cell cytotoxicity was initially decreased, however, subsequent cytotoxicity testing performed at months of age was normal. whole exome sequencing revealed a homozygous mutation in spink (c. - a>g) and a homozygous missense mutation in the first immunoglobulin domain of fcgr a (c. t>a), both variants of unknown significance and under additional investigation. this homozygous mutation in spink was not detected in the patients healthy, unaffected sibling. conclusions: this is a case of an infant with a clinical phenotype consistent with comel-netherton syndrome, however genotyping has revealed homozygous mutations in spink and fcgr a, suggestive of a possible mixed genotype. a recent large study investigating the use of whole exome sequencing to identify variants implicated in primary immunodeficiency found that in % of families, more than gene contributed to the immunodeficiency phenotype. it should therefore be kept in mind that variability in an individuals clinical phenotype may be attributable to the presence of a mixed genotype. introduction/background: cartilage-hair hypoplasia (chh) is a rare autosomal recessive disease caused by mutations in the rmrp gene and can manifest with scid. allogeneic hct is a curative therapeutic option for scid associated with chh. bordon et al. reported an overall survival of % ( / patients) following hct with a predominantly myeloablative conditioning regimen with busulfan and cyclophosphamide. data on outcomes of hct using a ric regimen are limited. objectives: we herein report our experience with allogeneic ric hct for scid associated with chh. methods: we reviewed records of all patients who underwent allogeneic hct for scid associated with chh at our institution, with a ric regimen containing alemtuzumab, fludarabine and melphalan. results: five patients ( male, female) underwent allogeneic ric hct for chh at median age of months (range, months years). all patients had biallelic mutations in the rmrp gene, and met pidtc criteria for scid, prior to hct. two patients were diagnosed by newborn screening for scid. one patient received serotherapy only (rituximab mg/m daily x days, anti-thymocyte globulin . mg/kg daily x days) conditioning for initial hct but developed graft failure and subsequently received a ric regimen for second hct. all patients received a ric regimen consisting of alemtuzumab mg/kg over days(n= ) or mg/kg over days (n= ) and fludarabine mg/kg (weight < kg, n= ) or mg/m over days (n= ), and single dose of melphalan . mg/ kg(weight < kg, n= ) or mg/ (n= , dose reduced by % for preexisting sclerosing cholangitis with grade liver fibrosis). patients received matched (n= ) or - allele mismatched (n= ) bone marrow grafts and all but grafts were from unrelated donors. all patients received cyclosporine and steroids for gvh prophylaxis. all patients engrafted with full donor chimerism. three patients developed mixed chimerism, but continue to maintain donor t cell chimerism > %. two patients developed vod of the liver (one patient developed mild vod whereas the patient with pre-existing sclerosing cholangitis developed severe vod). one patient developed grade skin gvhd and one patient developed limited chronic skin gvhd. none of the patients developed liver or gi-gvhd. all patients remain alive at a median follow up of years (range months- years) with good t-cell and b-cell immune reconstitution. conclusions: our experience suggests that allogeneic ric hct with alemtuzumab, fludarabine and melphalan for scid associated with chh is curative, offers durable t-cell engraftment, low gvhd along with excellent survival and might be preferable over a myeloablative conditioning regimen, to further limit toxicity in young infants, especially in the era of newborn screening. refractory thrombocytopenia in a patient with wiskott-aldrich syndrome despite hematopoietic stem cell transplantation: eltrombopag as a therapeutic option. mauricio chaparro-alzogaray , marcela estupiñan-peñaloza , gisela barros-garcía , oscar correa-jimenez pediatric hematologist/oncologist, hsct unit, fundación homi hospital de la misericordia pediatric hematologist/oncologist, fundación homi hospital de la misericordia pediatrics resident, universidad nacional de colombia introduction/background: wiskott-aldrich syndrome (was) is an xlinked disorder characterized by: immunodeficiency, eczema and hemorrhage due to thrombocytopenia [ ] . hematopoietic stem cell transplantation is the treatment of choice, with an overall survival of approximately % regardless of the source of the stem cells [ ] . eltrombopag has been used as a pre-transplant stabilization therapy [ ] . in our knowledge, there are no reports of its use for the management of post-transplant autoimmune cytopenias in was. objectives: to present a clinical case in which the complexity of diagnosis and management of wiskott-aldrich syndrome is highlighted, as well as the usefulness of eltrombopag in post-transplant persistent thrombocytopenia. methods: clinical case presentation and review of literature. results: clinical case: -month-old infant with a history of thrombocytopenia identified at the third day of life (positive serology and viral load for cmv), bacteremia due to serratia marcescens at months, intermittent diarrhea from months, multiple platelet transfusions, ivig cycles, and ambulatory management with corticosteroids; who consulted the er for a -day of bloody diarrheic stools, generalized petechiae and fever. he was irritable with generalized petechiae in the lower extremities, diaper area dermatitis with signs of superinfection. admission cbc: wbc: /mm , neutrophils /mm , lymphocytes /mm , monocytes /mm , hb . g/dl, ht . % mcv . fl, mch . g/dl, mcmh . %, platelets /mm mpv fl. bone marrow aspiration was performed that ruled out proliferative syndrome. immunological profile with normal immunoglobulins and flow cytometry showed t lymphocytosis with cd / cd inversed ratio, direct coombs was positive. he progressed to refractory thrombocytopenia, with intracranial bleeding and transfusion platelet requirement every h. molecular diagnosis of wiskott-aldrich syndrome was made and it was decided to carry out an allogeneic transplant of unrelated umbilical cord. he showed adequate response, % chimerism; however, he persisted with important cytopenias, without response to management with ivig and corticosteroids, so that on day + he restarted eltrombopag that he received prior to transplantation. from day + he received rituximab for weeks. he continued with eltrombopag months post-transplant. currently without cytopenia and without complications, he completed the post-transplant year without additional complications. conclusions: was represents a great diagnostic challenge in pediatric clinical practice. despite the therapeutic option of transplantation of hematopoietic stem cells, patients may persist with different complications, within these; autoimmune cytopenias will require additional therapies. eltrombopag, in addition to being used as a pretransplant transient measure, is useful for the management of post-transplant persistent thrombocytopenia in these cases. references: . immunol allergy clin north am. ; ( ) methods: retrospective study of medical records in two centers of immunology results: in our center we have p with pid, with made retrospective study of medical records, up today we have registered p ( %). according to this record, these diseases are distributed in the following way: predominantly antibody disorders: p ( %), predominantly t cell deficiencies: p ( , %), phagocytic disorders: p ( . %), complement deficiencies: p( . ), other well pid: p( . ), autoimmune and immune dysregulation syndromes: p( . %), unclassified immunodeficien cies: p( . %). predominantly antibody deficiencies are the most common pid, which comprise more than half of our all p. these group is represented by: specific iga deficiency (sad): p, specific igg deficiency: p, transient hipogammaglobulinemia: p, common variable immunodeficiency (cvid): p, agammaglobulinemia linked x: p, agammaglobulinemia unknown causes: p, secondary hipogamma globulinemia: p, selective igm deficiency: p, subclasses deficiency: p, cd l deficiency: p, hyper igm unknown causes: p, other hyogammaglobulinemia: p. among them, selective iga is the most common pid.in our cohort of unclassified immunodeficiency, we have p with auto inflammatory syndrome, such as family mediterranean fever, hyper igd syndrome and candle like-syndrome and p with nk deficiency. in all our pid p, are under replacement gammaglobulin (gg)treatment, p use intravenous gg and p use subcutaneous gg conclusions: the lasid registry model represents a powerful tool to improve health policies, showing that are under diagnosed and should receive more attention. more data are needed to define the exact prevalence of pid to avoid underestimation of these diseases due to under reporting. as different reports in different countries, in our centers predominantly antibody deficiencies are the most prevalent. although the number of patient diagnosed with pid, is growing. many physicians still know little about these disorders. introduction/background: a number of anticonvulsant medications have been shown to cause hypogammaglobulinemia. lamotrigine is a phenyltriazine anticonvulsant medication that is approved to treat seizure disorders and bipolar disorder. objectives: we describe a patient who developed hypogammaglobulinemia secondary to lamotrigine use. methods: we performed a chart review and case-based literature review. results: a- -year old female presented to immunology clinic for evaluation of panhypogammglobulinemia. she was initially evaluated by general internal medicine for lightheadedness and fatigue and was found to have serum igg of mg/dl ( - mg/dl), igm mg/dl ( - mg/dl) and iga mg/dl ( - mg/dl). the patient reported a history of recurrent sinus infections occurring around twice per year. she reported resolution with oral antibiotics. she denied any history of pneumonia and was never hospitalized for treatment of infection. she did report increased number of recurrent upper respiratory infections; around - per year for the last several months. she denied any family history of primary immune deficiency. she was never prescribed corticosteroids or immuno-modulators. her past medical history was significant for bipolar disorder type for which she was prescribed lamotrigine mg oral twice daily seven months prior to her initial visit. the medication was prescribed prior to the onset of recurrent sinus infections. she had protective post-vaccination titers against tetanus, diphtheria, and acellular pertussis and non-protective pre-vaccination pneumococcal titers. post vaccination pneumococcal titers were not obtained due to the cost of the pneumonia vaccine. the patient was started on prophylactic azithromycin three times weekly. lamotrigine was discontinued and the patient switched to lurasidone due to concern for anticonvulsant-induced panhypogammaglobulinemia. her serum immunoglobulin levels increased after . months off of lamotrigine (igg mg/dl, igm mg/dl, iga mg/dl), and she is currently asymptomatic without further infections. conclusions: lamotrigine is likely responsible for the reversible hypogammaglobulinemia in this patient. serial immunoglobulin levels should be checked in all patients who experience recurrent sinopulmonary infections while on lamotrigine. in two separate reports, lamotrigine induced hypogammaglobulinemia began within months and months of starting therapy respectively. in another study, hypogammoglubinemia was reported in % of patients taking lamotrigine. further studies are needed to accurately describe onset and frequency of hypogammgeobulinemia in these patients. currently, it is unclear whether post vaccine titers are protective in patients with lamotrigine induced hypogammaglobulinemia. introduction/background: the association of vaccine strain rubella virus with cutaneous and sometimes visceral granulomatous disease has been reported previously in patients with various primary immunodeficiency disorders (pids). the majority ( / ) of these pid patients with rubella positive granulomas had dna repair disorders, namely ataxia telangiectasia (at) (n= ) or nijmegen breakage syndrome (nbs) (n= ) or rag (n= ) and rag (n= ) deficiency. objectives: to support this line of inquiry, we provide additional descriptive data on the previously reported nbs patients as well as additional previously unreported patients with rubella virus induced cutaneous granulomas and dna repair disorders as well as additional previously unreported pid patients with rubella virus induced cutaneous granulomas. methods: we provide in-depth descriptive data on the previously reported nbs patients as well as additional previously unreported patients with rubella virus induced cutaneous granulomas and dna repair disorders including at (n= ) and dna ligase deficiency (n= ). we also provide in-depth descriptive data on additional previously unreported pid patients with rubella virus induced cutaneous granulomas, including cartilage-hair hypoplasia (n= ), mhc class ii deficiency (n= ), whim syndrome (n= ), and coronin- a deficiency (n= ). results: the median age of the patients is . years (range - ). the majority are females ( %). cutaneous granulomas have been documented in all cases while visceral granulomas (spleen and liver) were observed in cases. t cell and b cell lymphopenia as well as hypogammaglobulinemia or impaired antibody formation were present in most patients. all patients had received rubella virus vaccine. the median age at presentation of cutaneous granulomas was months (range - ). the median duration of time elapsed from vaccination to the development of cutaneous granulomas was months (range - ). the diagnosis of rubella was made by pcr in % of patients and by immunohistochemistry in the remainder. one patient was confirmed to have vaccine strain rubella virus. hematopoietic cell transplantation was reported in three patients. rubella associated complications did not contribute to death among those patients who died ( %). conclusions: of the now cases, ( %) share the diagnosis of a dna repair disorder and confirm that chronic rubella virus infection is associated with cutaneous granuloma formation. analysis of patients with dna repair disorders and other pids with this complication will help clarify determinants of rubella pathogenesis, identify specific immune defects resulting in chronic infection and may lead to defect-specific therapies. introduction/background: introduction/background: hizentra® is a % liquid igg product approved for subcutaneous administration in adults and children greater than two years of age who have primary immunodeficiency disease (pidd). limited information on use of hizentra® is available for children who have received hematopoietic stem cell transplantation (hsct). objectives: objectives: the aims of this study are to determine the safety and efficacy of hizentra® in pediatric patients post hsct, and to characterize reasons for switch from intravenous igg (ivig) to subcutaneous (scig) delivery following hsct. methods: methods: a retrospective chart review involved pidd infants and children (mean age . [range . to . ] months) status post hsct who received hizentra®. ten patients received hizentra® by pump administration, and patients by manual push. results: results: hizentra® administered weekly to children included an average of infusions (range - ). the mean dose was mg/kg/ weeks. the mean igg level was mg/dl while on hizentra® in patients compared to a mean trough igg level of mg/dl in patients during immunoglobulin administration prior to hizentra® of most children. four patients naïve to igg therapy were started on hizentra®. average infusion time was . (range - ) minutes for manual entry and (range - ) minutes for pump entry, and the average number of infusion sites was . (range to ). local reactions were mild and observed in / ( . %) children. ten patients had no local reactions. no serious adverse events were reported. the rate of serious bacterial infections (sbi) was . per patient-year while receiving hizentra®, similar to reported efficacy studies. the reasons for switch from ivig to scig in patients (some patients had multiple reasons) were improved igg serum levels and physician desire for steady state serum igg levels (n= ), loss/lack of venous access (n= ), patient/caregiver preference (n= ), home site of care preference (n= ), and physician preference for scig (n= ). conclusions: conclusions: hizentra® is a safe and effective option in children who have received hsct. reasons for switch from ivig to scig included improved serum igg levels, desire for steady state serum igg levels, and patient/caregiver preference. consulting medical advisor, immune deficiency foundation introduction/background: it is currently unknown how many persons with primary immunodeficiency (pi) receive the seasonal influenza vaccination, and what proportion becomes ill. additionally, the effect of immunoglobulin (ig) replacement therapy on the frequency of influenza diagnosis and severity of symptoms is not known. objectives: the current study sought to measure the prevalence of seasonal influenza vaccination and diagnosis among persons with pi, specifically those with antibody deficiencies: x-linked agammaglobulinemia (xla), common variable immunodeficiency (cvid) and hypogammaglobulinemia. methods: , email invitations were delivered to members of the immune deficiency foundation database requesting participation in an online survey regarding their zoster, influenza and varicella vaccination experiences. data from persons with xla (total n= ; children age< ; n= ; adults age> , n= ; % male; % white non-hispanic), cvid (total n= ; children age< , n= ; adults age> , n= ; % female; % white non-hispanic), and hypogammaglobulinemia (total n= ; children age< , n= ; adults age> , n= ; % female; % white non-hispanic) were analyzed for the - influenza season. results: overall, % (n= ) of the sample received a seasonal influenza vaccination during the - influenza season. persons with xla were less likely to receive an influenza vaccination ( %, n= ), than persons with cvid ( %, n= ) and hypogammaglobulinemia ( %, n= ). a fifth of respondents ( % overall, % of children and % of adults) who attend school and/or work stayed home at some time to avoid seasonal influenza. though the majority ( %, n= ) were not diagnosed with influenza, when stratified by age, children were twice as likely to be diagnosed than adults. of the children sampled, % (n= ) were diagnosed with seasonal flu compared to % (n= ) of the adults that were sampled. the role of replacement ig therapy in protection against flu can best be examined in the patients with xla since these patients cannot make specific antibodies to vaccine and only % (n= ) received the vaccine. although % (n= ) of the individuals with xla were exposed to flu, only % (n= ) were diagnosed with influenza; two of whom were not receiving ig therapy at time of diagnosis. concerning influenza severity, individuals receiving ig therapy at the time of flu diagnosis tended to have modestly milder symptoms than those not receiving ig treatment (e.g., less likely to report sore throat ( % versus %, p<. )), but the sample size is too low to draw firm conclusions. conclusions: a high proportion of antibody deficient persons received a seasonal influenza vaccination for the - influenza season. in addition to vaccination, many individuals attempted to avoid influenza infection by remaining home from school and/or work. there was a suggestion that ig replacement therapy may partially protect xla patients from symptomatic flu, although the role of cell mediated immunity in protection against flu is not clear. a prospective study could determine if ig replacement therapy partially protects against clinical flu symptoms in xla and cvid patients using two groups those that receive the current flu vaccine vs. those that do not receive the flu vaccine. introduction/background: fanconi anemia (fa) is an autosomal recesive or x-linked genetic disorder, characterized by cytopenia or bone marrow failure, this will lead to a severe anemia, neutropenia and thrombocytopenia, requiring frequent interventions with diversified therapies, including hematopoietic stem cell transplant (hsct). a complete immune reconstitution is requiere for a success transplant. one of the main upcoming events asociated after the hsct, is the secundary immunodeficiency, which is asociated with a significant morbility and mortality in the patients. to rich a successful alogenenic-hsct, is impending a complete reconstitution of the t cells immunity, for this it is crucial the presence of factors like; thymic activity of the hsct recipient, biological features of the allograft (eg, degree of histocompatibility, number and type of infused donor t cells) and preparative regimens. objectives: describe the kinetics of the immune reconstitution in fa patients after alogenic-hsct, as well as the infections associated during this process and other comorbilities. methods: we decribed the lympocyte population (t, b and nk) in pb measured by flow cytometry in fa patients on days + , + , + , + , + and + post alogenic hsct from a match related donor. the conditioning was base on fludarabine mg/m , cyclophosphamide mg/kg and antithymocyte globulin rabbit mg/kg. infectious-disease survillance for virus was determined by the quantification by dna pcr-rt for cmv, ebv, adenovirus, as well as the presence of galactomannan and candida sp antibodies, or isolation of fungus or bacterial culturing in the cases of infectious disease of known or uknown aetiology. results: the lymphocyte population mesurement was performed in a total of five fa patients who undergo to an allogenic hsct, all of them received stem cells from a match related donor and the source was bone marrow. successful engraftment was observed in all patients, there were no deaths reported. after the hsct was performed, the kinetics of recovering for the distinct lymphocytes subsets was the the following: nk cells (cd +cd +) were the first to recover, followed by cd+ t cells, b lymphocyte and finally cd+ t cells (figure ) all of them rich normal values and remain stable. three out of five patients presented infectious disease: two of them were cmv positive, one patient has a concurrent detection of adenovirus, and in the other was detected aspergillus, in both of them it was presented at day + . the third one developed acute gvhd which progressed to a chronic gvhd, at day + was diagnosed on him listeria monocytogenes meningitis, at that momento he has just cd+ +cd and cd + reconstituded. there is no new infectous disease detected in any of the patients after they reconstituted cd + t cells. conclusions: complete immune reconstitution is the decisive for the presence of several morbilities and mortalities, mainly because of oportunistic infecctions and gvhd. we show that the kinetics of recovery of the different populations of lymphocytes follows those patterns also described for patients with other hematological malignancies: early recovery of nk cells, followed by effector cytotoxic t cells and b cells, and finally, cd + t-helper cells. the utility of post-transplant monitoring of pb-lymphocyte subsets for improved follow-up of patients undergoing bmt and prevent opportunistic infections. introduction/background: early detection of primary immunodeficiency diseases (pids) before serious infection is of utmost importance and a question of patient survival. one of the main organs involved in pids is skin with mucocutaneous manifestations being one of the common complaints in pids. the presenting skin symptoms may serve as an essential element in early diagnosis of pids. objectives: this study aims to determine characteristics, frequency, nature and incidence of skin manifestation in pid patients seen in qatar. methods: this retrospective study was conducted at hamad medical corporation the only tertiary hospital in qatar from january to july .the subjects included were pid patients < years old, who had dermatological complaints. information collected included dermatological diagnosis, gender, age at onset of signs and symptoms, age at definite diagnosis, family history of related disease, any lab results such as pathology or viral studies obtained from the clinical medical record. patients were diagnosed and classified according to clinical and laboratory criteria by the international union of immunological societies primary immunodeficiency committee. results: a total of patients were studied and skin/mucocutaneous manifestations were found in patients ( %) with male to female ratio of . : . age at onset of skin manifestations ranged from to years. skin manifestations were divided in two categories according to presentation before pid diagnosis (primary manifestations n= , %) and during the disease period (secondary manifestations n= , %). the type of pids predominantly having primary manifestation were scid (n= , %) cvid and cgd (n= , %, each), xla, at and griscelli syndrome (n= , %, each).various manifestations which were atypical in their presentation and persistently found in patients who did not respond to any effective treatment (n= ) led to the basis of clinical immunological study and diagnosis including rare diagnosis such as ipex. whereas secondary manifestations were primarily reported in scid (n= , %) cgd, at and digeorge syndrome (n= , %, each). the nature of skin manifestations varied in both groups, primary manifestations notably had % cutaneous infections comprising of thrush ( %) stomatitis ( %) viral rashes ( %) bacterial rashes ( %) impetigo ( %) and fungal rash ( %). eczema/atopic dermatitis were % and other minor miscellaneous skin alteration cases found were apthous mouth ulcer ( ) erythroderma ( ) gvhd like rash ( ) alopecia ( ) psoriasis ( ) and scleroderma ( ) . secondary manifestations were identified as % cutaneous infections % eczema and infantile seborrheic dermatitis ( ) pruritus ( ) gvhd and alopecia ( ) each. the causative microorganisms were confirmed in % of the cases via lab cultures whereas common infections such as chicken pox and herpes were validated through clinical symptoms. none of the cases were biopsied. overall, highest skin infection was seen in ataxia telangiectasia ( ). other pids with prominent cutaneous manifestations included cgd ( ), digeorge syndrome ( ), hyper ige syndrome ( ) and scid (rag or , cases). more than two types of skin manifestations were found in % patients over the due course of their illness. conclusions: an awareness of various cutaneous and skin disorders associated with pid which are persistent and unresponsive to treatments among dermatologist and family physicians is crucial to raise suspicion for early detection, timely management and prevention of complications. introduction/background: ataxic telangiectasia (at) is a rare neurodegenerative autosomal recessive disease associated with immunodeficiency, poor coordination and disability. ataxia telangiectasia has a diverse clinical heterogeneity which may often lead to an incorrect diagnosis or late detection of the disorder resulting in exposing the patient to unnecessary radiation from various sources. objectives: we present a female child from the asian subcontinent that was seen for the first time with ulcerative skin lesions, failure to thrive and marked lymphopenia. methods: a search of the pubmed database was carried out, using different combination of the terms "ataxia", "telangiectasia", typical , atypical and "presentation" results: four-year-old pakistani girl, product of first degree nuptial presented with generalized vesicular rashes mainly on abdomen and scalp which turned into ulcers with residual hypo pigmented lesions and diarrhea. she had short stature, failure to thrive (weight and height < th percentile) and no previous infection or family history of primary immunodeficiency diseases (pid). mild delay in milestones was observed especially in speech. immunizations were up-to-date including bcg. on examination she had mild ocular telangiectasia but no cutaneous involvement. investigations revealed lymphopenia ( . ) and eosinophilia. workup for diarrhea including duodenoscopy revealing subtotal villous atrophy however the biomarkers for celiac disease were negative. immune system workup showed high igg levels, normal iga, ige and igm level. igg subclasses: low igg , and , antibody titers to h.influenza and pneumococcus vaccine were relatively low. lymphocyte subsets showed low cd ,cd and high nk cells ( %), low naïve t cells ( %) and inverted cd /cd ratio.t cell function with post pha was . % but had normal response to cd . alfa feto protein was requested due to continued low cd count and the level was found to be high: . iu/ml. next-generation sequencing (ngs) showed homozygous mutation for trp fs in chromosome of atm gene confirmed by whole exome sequencing. immediate treatment with ivig was started leading to mark improvement of skin lesions, diarrhea and weight gain. conclusions: the index of suspicion of at should be highlighted when deciphering lymphocyte subset. currently there is no neonatal screening for pid diseases and next generation sequences in qatar. once implemented it may prove beneficial in discovering cases from early infancy and reaching upon a definite diagnosis. introduction/background: chronic granulomatous disease (cgd) is a genetic disorder of the nadph oxidase complex in phagocytes which results in impaired production of microbicidal reactive oxygen species that can lead to recurrent life-threatening bacterial and fungal infections. the majority of affected patients in the united states have a x-linked defect in the cybb that encodes gp phox protein followed by an autosomal recessive defect in the ncf gene that encodes p phox . x-linked female carriers show populations of neutrophils and following lyonization, and severe skewing of x-chromosome inactivation can get cgd type infections. a recent study has shown that the lower dihydrorhodamine (dhr) percent in female carriers predicts a higher infection risk but carrier state on its own predicts autoimmunity results: a -year-old female presented for an evaluation of recurrent fevers. she had a history of several lobar pneumonias in childhood. at age , she developed erythematous bumps on her abdomen that were initially non-bothersome but later became painful. she then developed recurrent fevers, chills, and rapid weight gain. biopsy of the skin lesions showed subcutaneous panniculitis-like t cell lymphoma. she was treated with chemotherapy and found to be in complete remission. she had no previous family history of recurrent infections. at the completion of her chemotherapy, her -year-old son became septic in the hospital and was then diagnosed with cgd from mutated cybb gene. post chemotherapy, the patient had a prolonged course with an atypical lung infection that required bronchoscopy and video thorascopic surgery, however no bacteria was ever identified. she did improve after a long course of ivantifungals. evaluation for immunodeficiency was done after she had a recurrence of low grade fevers, cough, and fatigue. dhr flow cytometry with phorbol myristate acetate (pma) was done which showed . % neutrophil oxidative burst activity after stimulation, consistent with a highly skewed lyonization pattern in x-linked cgd. conclusions: this case demonstrates an interesting lesson in a woman with recurrent infections since childhood whose clinical picture was confounded by her history of malignancy. the abnormal lung infections prior to malignancy were alarming and gave cause to do additional immunology testing. however, it begs the questions, if she did not have a son, would she have been diagnosed with severe x-lined carrier disease. introduction/background: measurement of the specific antibody response following vaccine challenge provides clinicians with a better understanding of the adaptive immune response in individuals undergoing immunological evaluation. objectives: we hypothesised that after classification of primary immunodeficiency (pid) patients based on vaccine response (vr), further division using igg subclass (iggsc) - measurements may identify additional patients with abnormal b cell function and patients with different frequencies of infection at presentation. methods: vrs and serum iggsc concentrations were quantified using the vacczyme anti-pneumococcal polysaccharide (pcp) igg elisa and human iggsc liquid reagent kits (the binding site group limited, uk) in pid patients ( : . m:f, median age . years, range - ). all patients were immunised with pneumovax® (sanofi pasteur msd). the lower limits of published normal iggsc ranges were used as cut-off values (igg . g/l, igg . g/l and igg . g/l). pcp concentrations equal to or less than mg/l post-vaccination was considered an abnormal response. results: agreement between vr and iggsc measurements was % (p= . ). the frequency of respiratory tract infections at presentation among pid patients with a normal pcp igg vr was % and % among patients with an abnormal vr. subsequently, four separate groups could be identified by including iggsc measurements. the frequencies of infections at presentation were for the vr+/iggsc+ group (n= ) % vs. % for the vr+/iggsc-group (n= ); and % for the vr-/iggsc+ group (n= ) vs. % for vr-/iggsc-individuals (n= ). conclusions: these results confirm that vr and iggsc measurements are independent serum biomarkers of humoral immunity. taken together the vr and iggsc results provide more detailed information about the immune status that may influence diagnosis, treatment and monitoring decisions. introduction/background: hizentra® is a % liquid igg licensed for subcutaneous administration in adults and children greater than two years of age who have primary immunodeficiency disease (pidd). subcutaneous immunoglobulin (scig) use in autoimmune conditions is reported. stiff person syndrome (sps), a rare neurologic disorder characterized by fluctuating muscle spasms and rigidity, is mediated by autoantibodies to glutamic acid decarboxylase (gad). symptoms of sps have been shown to improve after administration of intravenous immunoglobulin (ivig) however, there is a paucity of information regarding use of scig in sps. objectives: the aim of this study is to describe the use of hizentra® in patients with sps, including indications for scig, clinical characteristics of patients, and clinical and laboratory response to scig. methods: a multicenter retrospective chart review examined patients with stiff person syndrome treated with hizentra®. results: sps was diagnosed in patients at and years. both patients were started on ivig, steroids, and rituximab prior to initiation of weekly hizentra® (ages and years). the average dose of hizentra was mg/ kg weekly. the average igg level while receiving ivig was , . mg/dl, similar to the average igg level while on hizentra was , mg/dl. the number of administration sites ranged from - , and duration of infusions ranged from - minutes. serum anti-gad antibody levels prior to hizentra® were u/ml and u/ml respectively. anti-gad antibody level during treatment with hizentra (available for one patient) was > , u/ml, and , u/ml following discontinuation of hizentra®. one of the reasons for switching to hizentra® was lack of response while on ivig. the average number of hizentra infusions were . patients reported improvement in spasticity related to sps while on hizentra®, and one patient had improvement in seizures. one patient discontinued hizentra® in favor of intravenous immunoglobulin (ivig) due to physician preference. the most common side effects were local reactions including pain, pruritus, and redness. no serious adverse events were reported. conclusions: hizentra® was associated with improved symptoms in sps in both patients including decreased spasticity, and improved seizure frequency in one patient. serum anti-gad levels did not decrease following administration of hizentra. hizentra® was well tolerated in patients with sps, with most side effects reported as mild. hizentra® may be considered as an alternative to ivig treatment in patients with sps. introduction/background: combined immunodeficiency (cid) has been associated with a spectrum of secondary gastrointestinal manifestations, including infectious and non-infectious causes. abatacept, a soluble ctla -igg fusion protein targeting t cell activation, has demonstrated benefit in the treatment of autoinflammatory manifestations in patients with cid due to underlying heterozygous germline mutations in ctla and lrba. objectives: herein, we report two patients with cid characterized by low immunoglobulin levels, low class-switched memory b cell counts, and low peripheral naïve cd + t cell counts. both patients suffered from severe and persistent diarrhea complicated by protein-wasting and malnutrition, ultimately diagnosed as biopsy-consistent autoimmune enteropathy. the clinical history of patient was also notable for granulomatous lung disease and autoimmune cytopenias. methods: a t-regulatory cell disorder was considered in both cases, however, ctla and lrba were found to be unaffected by whole exome sequencing (patient ) and/or flow cytometry (patient ). due to progressive worsening of the autoimmune enteropathy despite management with chronic steroids (both patients), combination rituximab/mycophenolate mofetil (patient ), and total parenteral nutrition (tpn) complicated by recurrent infections (patient ), abatacept was trialed at mg sc weekly. results: in both patients, the start of abatacept therapy produced a dramatic improvement as measured by decreased stool frequency, improved weight gain, and decreased protein-wasting. patient has been maintained on this monotherapy for over one year, with improvement in the gastrointestinal as well as pulmonary autoinflammatory complications. the only documented adverse event has been hepatitis b reactivation, managed with tenofovir and abatacept continuation. patient has required azathioprine/abatacept combination therapy for clinical stabilization and is without a significant adverse event to date. conclusions: we conclude that abatacept may be a relatively safe and effective therapeutic in the management of severe autoimmune enteropathy in the background of cid, even when used outside of the classical clinical context of ctla or lrba haploinsufficiency. objectives: as increasing the outbreak of allergic diseases, study for treatments comes to the force. in this research, we aimed to assess the effects of sg-sp , a derivative of gallic acid, on mast cell-mediated allergic inflammation using various animal and in vitro models. methods: ovalbumin-induced systemic anaphylaxis and immunoglobulin e (ige)-induced passive cutaneous anaphylaxis are standard animal models for immediate-type hypersensitivity. oral administration of sg-sp hindered the allergic symptoms in both animal models. these inhibitions were deeply related to the reductions of histamine and interleukin- . results: sg-sp reduced degranulation of mast cells and expression of inflammatory cytokines in a dose dependent manner. sg-sp showed better anti-allergic effects compared to gallic acid and dexamethasone. down regulations of intracellular calcium level and nuclear factor-b activation by sg-sp were causative of the reduction of allergic mediators. to anticipate the exact target of sg-sp , phosphorylation of proteins involved in mast cell signalling was assessed. sg-sp suppressed the activations from lyn and was aggregated with high affinity ige receptor (fcri). conclusions: from these results, we assured that sg-sp directly interact with fcri. all together, we propose that sg-sp might be a therapeutic candidate for allergic disorders. ( ) submission id# systematic assessment of pain in patients with primary immune deficiency using validated pain questionnaires: a prospective study. introduction/background: the number of primary immunodeficiency (pid) patients is rising dramatically because of good medical care as well as increased awareness. around in live births are affected. more than disorders have been discovered so far and this number is expected to rise in the coming years. as with any other chronic illness, pid patients are also prone to acute and chronic pains. chronic pain is a big challenge and lack of understanding of the etiology and underlying mechanisms limit our ability to diagnose or treat it effectively. objectives: to systematically assess chronic pain in patients with pid using validated questionnaires and to try to understand the underlying mechanisms of neuropathic versus non neuropathic pain. methods: short-form mcgill pain questionnaire (sf-mpq) is a recognized way to ascertain different pain characteristics as well as severity. a validated arabic version of the sf-mpq was used to prospectively assess chronic pain in patients with pid. furthermore, a validated arabic version of the neuropathic pain questionnaire-short form (npq-sf) was also used to assess neuropathic pain and to differentiate it from nonneuropathic pain. a total of patients with pid were included. results: males: females were . %: . % respectively. mean age was . years. commonest diagnosis was combined immune deficiency in % of the patients followed by common variable immune deficiency which was . %. chronic pain was found in % of the patients that participated in the study. % of the patients who complained of pain found it to be tiring and exhausting. % each had aching or heavy or sharp pain. % had cramping pain, % had tender pain and % characterized their pain as throbbing or burning. % felt shooting pain, % had splitting pain and % had gnawing pain. % found it to be frightening and % described it as being sickening in itself. the commonest pain complaint was abdominal pain in % of the patients followed by headache % and chest pain %. pain attributed to neuropathy was present in about . % of the study population. most patients described that they had been experiencing pain for at least - years. conclusions: this is the first international study to understand the prevalence, duration and severity of chronic pain among pid patients. a significant number of patients reported ongoing pain. this is the first time any kind of pain has been studied systematically in the patients with primary immune deficiency. treating pain should have a major impact on improving the patients quality of life. introduction/background: primary immunodeficiency (pidd) patients, particularly those with severe t cell defects, are at increased risk of infections. bone marrow transplant also contributes to significant t cell lymphocytopenia, which can be associated with similar risks. several prophylactic measures, which vary per institution, are placed on these patients in order to prevent infections. we report on a likely outbreak of rapid growing nontuberculous mycobacteria (ntm) at our institution, with a suspected association to tap water objectives: to recognize nontuberculous mycobacteria as a threat to patients with immunodeficiency disorders methods: case series of consecutive patients admitted to one institution with similar infections results: in a period of months, / to / , patients admitted to our institution were found to have rapid growing ntm species on central line culture or bronchoalveolar fluid. these cultures were obtained due to fever and symptoms of systemic infection. per institutional practices no peripheral blood samples were obtained at the time of initial culture. all had significant t cell dysfunction: wiskott-aldrich syndrome ( ), an undefined combined immunodeficiency ( ) , and three patients post bone marrow transplantation (bmt), one months ( days) post an unrelated cord blood transplant for farber syndrome, one days out of a matched related transplant for epstein barr virus (ebv)-associated lymphoproliferative disorder, and one patient with high risk neuroblastoma days out of his second tandem autologous transplant. both allogeneic transplant recipients had received serotherapy. all species were rapid growing, identified as mycobacterium mucogenicum (n= ), immunogenum (n= ), or abscessus-chelonae complex (n= ). the patients had not shared rooms, caregivers, invasive procedures, or medications from the same batch. three ( %) cases met cdc criteria for hospital acquired infections (hai). all patients were in general ward rooms, and were on standard precautions given diagnosis had not been established (n= ) or they were considered outside the typical window of strict bmt precautions (n= ). ntm species were subsequently isolated from hospital tap water. this has resulted in a significant increase in infection precautions, including the use of sterile water only for cares, as well as bottled water for drinking (with no use of ice machines), for all patients being evaluated for pidd or with significant lymphocytopenia. conclusions: ntm are a threat to patients with pidd. tap water is a potential source of mycobacterial infections in pidd patients. minimizing exposure risk to water sources containing ntm is very important in this population. patients with concern for pidd or significant t cell lymphocytopenia should take steps to avoid ntm exposures, including the use of sterile water for cares, and bottled water for drinking. introduction/background: purine nucleoside phosphorylase (pnp) deficiency is a rare autosomal recessive condition leading to severe combined immunodeficiency and neurologic impairment, typically presenting in early childhood. the condition is progressive and typically fatal in the first or second decade of life without hematopoietic stem cell transplantation (hsct). objectives: to illustrate the clinical and laboratory presentation of pnp deficiency with a novel mutation in the pnp gene via a case study. results: case: a four year old female of hispanic descent, with a past medical history of spastic diplegia, initially presented with chronic nasal congestion, recurrent sinusitis, and cough. laboratory studies were significant for an alc cells/mcl ( - cells/mcl). she was lost to follow-up for two years, before returning to medical attention for pneumonia. evaluation of lymphocyte subsets at age years revealed cd + cells/mcl ( - cells/mcl), cd + cells/mcl ( - cells/ mcl), cd + cells/mcl ( - cells/mcl), and cd + cells/mcl ( - cells/mcl). t cell mitogen stimulation to pha measured by flow cytometry was % of control. pnp activity was nearly absent and urine guanosine and inosine were significantly elevated. gene sequencing revealed a homozygous c. a>g mutation in the pnp gene. prophylactic antimicrobials were started. despite strong recommendation for hsct, the patient was again lost to follow up until age years, at which time she was found to have progressive lymphopenia, bronchiectasis, and developmental delay. t cell mitogen stimulation to pha measured by thymidine uptake assay was % of control and pnp functional activity was undetectable. the patient underwent a / matched unrelated hsct six years after her initial presentation. one year post-transplant, the patient continues on immunoglobulin replacement therapy. her course was complicated by cutaneous gvhd, treated successfully with topical corticosteroids. conclusions: this case study illustrates the progressive nature of pnp deficiency in the first decade of life. our patient is notable in that she survived without significant medical intervention to the age of years. her presentation at age years was not unlike those previously reported in the literature, with muscle spasticity, ataxia, and recurring bacterial infections. to the authors knowledge, this case reports a novel mutation in the pnp gene. introduction/background: severe congenital neutropenia (scn) is a primary immunodeficiency disease characterized by early onset recurrent infections, persistent severe neutropenia and congenital genetic defect. severe idiopathic neutropenia (sin) is a rare disease defined by persistent severe neutropenia, in the absence of an identifiable etiology. objectives: here, we aim to find out clinical, laboratory, genetic characteristic and remission status in children with scn and sin in chinese population. methods: in this study, we enrolled chinese children who experienced severe neutropenia longer than months without any virus infection or auto-immune antibodies from june to july in hospitals affiliated to shanghai jiao tong university school of medicine. their clinical, laboratory and molecular characteristics were analyzed and the patients were followed up to observe their remission status. targeted gene capture combined with next-generation sequencing technology was used to find out related gene mutation. results: patients in this study had a mean age of . ± . months. molecular analysis revealed that patients had associated mutations of scn, including elane and g pc . among patients with continuous follow-up, one died for unknown reason. ten patients have recovered from sin (r-sin) with mean neutropenia duration of . ± . months. scn patients had more frequent infection ( . ± . times per year) than sin ( . ± . times per year, p= . ) and r-sin patients (p= . , . ± . times per year). scn patients had significantly higher count of anc and monocytes than sin (p= . ) and r-sin patients (p= . ). however, there was no difference in anc and monocytes counts between sin and r-sin patients. bone marrow examinations demonstrated a myeloid maturation arrest at the myelocyte-metamyelocyte stage in scn patients, while most of sin and r-sin patients were normal. conclusions: our study indicated that, patients with mild infection, lower anc, monocytes count and normal bone marrow are likely to be sin. whereas others with relatively more severe infection, higher anc, monocytes count and maturation arrest in bone marrow are inclined to be scn. introduction/background: patients with qualitative and/or quantitative defects of humoral immunity often require immunoglobulin (igg) replacement therapy (igrt). usual starting doses range between - mg/kg and the dose is adjusted as needed depending on the patient. there is a paucity of information about whether and how extreme bmi (obese or underweight) and route of administration may affect a patients rate of infections. objectives: the objective of the study is to determine whether rate of infection is associated with bmi, dose or route of administration in patients receiving igrt. methods: this is a retrospective chart review from december -october . we included patients between the age of month and years old who were evaluated initially and had at least one follow up evaluation. data reviewed included the route of administration, dose, infection history, igg serum levels, height and weight to calculate bmi, gender, age and diagnosis requiring immunoglobulin replacement therapy. participants were excluded if the type of immunodeficiency is unknown or if the participant had incomplete data for the requested data fields. the number of infections between visits was modeled using poisson regression as a function of dose, route of administration and the bmi with the log of the follow up interval as an exposure offset. results: eighty-five patients were eligible, and preliminary results for patients are presented here. the mean infection rate per weeks of follow up was . in obese patients, . in overweight patients and . in underweight/normal weight patients adjusted for route of administration; however these rates do not differ significantly from one another. the mean infection rate per weeks of follow up differed by administration route; . infections for ivig versus . infections for scig when adjusting for bmi (p< . ). the mean infection rate per weeks of follow up was not associated with dosage. conclusions: overweight patients may experience more infections than obese or underweight patients, regardless of administration route. ivig patients may have a lower rate of infections compared to scig patients regardless of bmi. work is ongoing to complete analyses for the remainder of the eligible patient population. ( ) submission id# introduction/background: primary immunodeficiencies (pids) are a rare group of genetic disorders associated with a tendency to infectious diseases and an increased incidence of cancer. there is no data regarding the prevalence of malignancies in patients with pid in turkey. objectives: in this study, we aimed to evaluate patients who diagnosed as pids and developed malignancies, and calculate the estimated frequency of malignancies associated with pids in turkey. methods: forty five patients who were diagnosed with malignancy in the follow-up period of pid at the four tertiary immunology clinics between and years were included in the study. the data were obtained retrospectively from the hospital records and the database of esid online patient registry. results: the prevalence of malignancies in patients with pid was found as . % ( / ). the male to female ratio of the patients was / , the median age was . years (minimum: . , maximum: ) and the median age at which patients get diagnosed with malignancy was years (minimum: . , maximum: ) . there was no cancer history in their family members. the most common type of pid which associated with malignancy was ataxia telangiectasia (n= , . %). non-hodgkin lymphoma was the most common malignancy (n = , . %) in our group (table ) . ebv quantitative pcr was positive in lymphoma cases ( . %). the median number of x-rays and ct scans in patients with at and bloom syndrome before malignancy developed were (minimum: , maximum: ) and (minimum: , maximum: ), respectively. two cases had dual malignancies (papillary thyroid cancer and anal adenocancer). twenty cases were treated with chemotherapy, cases with hematopoietic stem cell transplantation, cases with radiotherapy, and cases with surgical treatment, treatment information of patients was not reached. remission was detected in cases, while resistance to therapy in cases and recurrences in two patients were observed. four patients are still on chemotherapy. twenty cases died. conclusions: the tendency of malignancy in patients with pids is due to the deficiency in the immune response that lead to failed surveillance against oncogenic viruses, premalignant/malignant cells, or both. lymphoid malignancies are the most common malignancies associated with pids. pids-associated malignancy incidence has increased in recent years because of that improved survival of the patients. this study is the largest cohort investigating the association of malignancy in patients with pid in turkey. additionally, we first reported tendency to malignancy in a patient with znf deficiency. introduction/background: next-generation sequencing (ngs) has become an integral tool in the evaluation of primary immunodeficiency disorders (pid). we describe a patient with a previously described pathogenic foxp variant who met clinical and laboratory criteria for cvid. objectives: describe a patient with a previously described pathogenic foxp variant who met clinical and laboratory criteria for cvid. results: case description: an -year-old male born premature at weeks presented with a history of recurrent infections. family history was negative for immunodeficiency. the patient developed recurrent acute otitis media beginning at year of age, three episodes of pneumonia beginning at years of age, and recurrent sinus infections requiring treatment on average four times a year beginning at years of age. initial immunologic evaluation at age was notable for: igg mg/dl (reference - mg/dl), igm mg/dl (reference - mg/dl), iga mg/dl (reference - mg/dl), ige iu/ml (reference - iu/ml). lymphocyte subpopulations were normal. specific responses to vaccines showed: protective antibody titers to diphtheria, but not to tetanus or pneumococcal antigens. he did not respond to booster vaccination and was started on ivig with significantly reduced frequency of infections. at age , while on ivig, he developed oral ulcers (biopsy consistent with ulcerative eosinophilic granuloma), abdominal pain, and recurrent arthralgias involving ankles, elbows, hips and the sacroiliac joint. magnetic resonance imaging (mri) was consistent with sacroilitis. subsequent imaging was consistent with chronic relapsing osteomyelitis (crmo). gastrointestinal biopsies showed severe active chronic pangastritis with antralized oxyntic gastric mucosa with enterochromaffin cell hyperplasia; suggestive of autoimmune gastritis. plasma cells were present throughout the gastrointestinal tract. ngs (bcm-ngs, baylor miraca genetics laboratory, houston tx) identified a hemizygous pathogenic missense variant, c. g>a (p.r q) in the x-linked foxp gene, that has been reported previously in patients with immunodysregulation, polyendocrinopathy, enteropathy, x-linked (ipex) syndrome. flow cytometry studies showed a decreased percentage of treg cells of total cd expressing cells, . % (reference . - . %), but normal foxp expression within these cells, % of treg cells with foxp expression (reference - %). interestingly, treg cell subset phenotyping obtained at the same time showed a normal percentage of natural tregs, . % cd t cells (reference . - . %), as well as normal percentage of naive tregs, . % cd t cells (reference . - . %). up to this point, he had not had any signs of diabetes, thyroid disease or frank enteropathy involving the small or large intestine. conclusions: we report a pathogenic foxp variant, occurring in a patient with a cvid-like phenotype, autoimmune gastritis, and an association with crmo. this case demonstrates the increasing utility of ngs, which can profoundly impact prognostic and therapeutic considerations. ( ) submission id# the natural history of patients with profound combined immunodeficiency (pcid): interims analysis of an international prospective multicenter study. immunodeficiency in this patient group. so far, patients were transplanted after enrolment, overall patients died ( in the hsct group, in the non-hsct group). analysis of the hsct decisions revealed the divergent decisions in patients with similar disease burden, favoring an ongoing prospective matched pair analysis of patients with similar disease severity with or without transplantation. so far, neither the genetic diagnosis nor simple measurements of t-cell immunity emerged as good predictors of disease evolution. conclusions: the p-cid study for the first time defines and characterizes a group of patients with non-scid t-cell deficiencies from a therapeutic perspective. since genetic and simple t-cell parameters provide limited guidance, prospective data from this study will be an important resource for guiding the difficult hsct decisions in p-cid patients. ( ) submission id# the plasma contact system and its role in common variable immunodeficiency: an explorative study. introduction/background: a growing body of evidence suggests that the contact system is involved in the activation of various vascular and immunological pathways and acts as an interface to help regulate allergic reactions, coagulation, complement, innate immunity and inflammation. as demonstrated in mice experiments, contact activation and high molecular weight kininogen (hk)-derived peptides increased homing of t and b lymphocytes into lymph nodes, which suggests an important area of research for understanding the contact systems role, specifically fxii, in immune-mediated inflammation and immune dysregulation. this novel mechanism prompted further inquiry into its role of various human disease states characterized by inflammation. plasma hk cleavage has been proposed as a useful and minimally invasive biomarker in various inflammatory disease states. this pathway has not been explored in cvid, in which inflammatory complications are found in one-third of patients with an unidentified genetic cause. characterizing the contact system biomarkers in cvid patients could elucidate a role in pathogenesis. objectives: assess the presence of contact activation at baseline in sera from cvid patients with and without inflammatory complications compared to healthy controls. methods: cvid patients were recruited in the outpatient setting and the measurement of cleaved plasma hk (chk) levels was determined by western blot analysis, under reducing conditions, with quantitation of total and chk bands using an odyssey imaging system (licor). a one-way anova test for differences among the studied groups will be applied. c inhibitor levels, c and c levels and high-sensitivity crp were also measured as comparable biomarkers for inflammation. results: to date, cvid patients were studied, with and without inflammatory complications. repeated determinations of cleaved hk% (chk%) revealed an average of . % (range: . - . %) in cvid patients with inflammatory complications and those without complications averaged . % (range: . - . %). healthy controls had an average chk of . % (n= , range: . - . %). conclusions: cleaved kininogen detected in the sera of cvid patients was found at similar levels compared to healthy controls (chk< %). findings suggest that systemic activation of the contact system might be absent in cvid, however, future considerations include developing detection methods for local tissue activation. ( ) submission id# the underlying primary immunodeficiencies and lung diseases, and low cd and cd counts are associated with recurrent pneumonia in hiv negative lymphopenia patients. clinical fellow, yale university school of medicine introduction/background: lymphopenia can be considered as primary or acquired immunodeficiency. lymphopenia is associated with a considerable increase in susceptibility to infections and the treatment focus for lymphopenia is mainly consists of prophylaxis and treatment of opportunistic infections. aids is the most well known cause of acquired lymphopenia and the cd count serves as an effective surrogate marker for disease progression and guideline for prophylaxis for hiv positive lymphopenia patients (hplp). hiv negative lymphopenia patients (hnlp) have been following the same prophylaxis guideline for hplp patients. however, it is unclear whether same prophylaxis guideline will be appropriate for both groups since the underlying immune mechanisms are different between these two groups. objectives: we aimed to define the optimal treatment and prophylaxis guideline for hnlp. in this study, we compared the clinical phenotypes and absolute counts of lymphocyte subsets between hnlp with recurrent pneumonia (pna), recurrent upper respiratory infection (uri) and no pulmonary infection. methods: electronic medical records of hnlp (n= ) seen at an academic immunology clinic between the year of and were reviewed retrospectively. lymphopenia was defined as absolute cd count less than /μl. the age, absolute counts of cd , cd , cd , cd , nkt and nk cell counts, history of antibiotic or antiviral prophylaxis use, autoimmune disease, lung disease, immunosuppressive therapy use, hypogammaglobinemia and ivig therapy use were compared between patients with recurrent pna (n= ), recurrent uri (n= ) and no pulmonary infection (n= ). results: this study showed that patients with recurrent pna had significantly lower absolute cd , cd , nk cell counts and age compared to the patients with recurrent uri ( table ) . none of the clinical phenotype was significantly different between these two groups. when we compared the patients with recurrent pna vs no infection, all lymphocyte subset counts and age were similar between these two groups except the frequency of underlying lung disease which was significantly higher in the recurrent pna group (table ) . lastly, we grouped the recurrent pna and uri groups together as the pulmonary infection group. when we compared the pulmonary infection group with the no infection group, lymphocyte subset counts were not significantly different, however, infection group showed significantly higher incidence of pid and the trend of lower rate of is therapy use than the no infection group (table ) . conclusions: the initial study has several limitations. this was a retrospective study from a single clinic and patient population was limited to patients. despite these limitations, we believe that this study provides valuable messages regarding prophylaxis guideline for pulmonary infection in nhlp; the underlying pids including icl and cvid, lung diseases particularly bronchiectasis and the absolute cd and cd counts less than /μl and / μl, respectively, are associated with recurrent pna and the patients with these risk factors likely will benefit from antibiotic prophylaxis. in addition, patients with acquired lymphopenia due to chronic use of is therapies without underlying pid or lung disease less likely develop pulmonary infection despite of low cd and cd counts. further investigations are crucial to elucidate the clinical significance of our initial observation by increasing the patient population and analysis of detailed immunologic and genetic profile of these patients which will likely reveal immunological markers and genes that are involved in the pathogenesis of both primary and hiv negative acquired lymphopenia. director, neuro-oncology program, division of hematology/oncology, ucsf benioff children's hospital oakland, ca introduction/background: gata is a hematopoietic transcription factor, required for the development and maintenance of a healthy stem cell pool. heterozygous mutation of gata has been associated with different but overlapping syndromes affecting both myeloid and lymphoid cell lines including aplastic anemia, myelodysplastic syndrome, acute myeloid leukemia, pulmonary alveolar proteinosis and lymphedema. it is also associated with immunodeficiency and susceptibility to mycobacterial, fungal, and viral infections. hematopoietic stem cell transplantation (hsct) is the only available cure which should ideally occur before patients develop neoplasia, severe infections or lung disease. objectives -to describe the clinical presentation of a genetically confirmed case of gata mutation -to describe the characteristic hematological and immunological profile of patients with gata mutation -to emphasize the importance of high index of suspicion and early diagnosis in improving outcome with hsct methods: case: a -year-old female presented with prolonged fever, fatigue, nonspecific rash, and unremarkable clinical exam. laboratory evaluation was significant for mild pancytopenia with profound monocytopenia. bone marrow analysis was remarkable for hypocellularity without evidence of dysplasia or malignancy. peripheral blood flow cytometry showed decreased t-and b-cells and absent nk cells. results: the constellation of pancytopenia, marked monocytopenia and absent nk cells, were suggestive of gata deficiency. sanger gene sequencing of gata revealed a heterozygous nonsense mutation (p.arg *). conclusions: mutation of gata is the underlying defect in overlapping clinical syndromes and is associated with immunodeficiency and malignant predisposition. incidence of organ dysfunction, infections and neoplasia increases with age. confirming the diagnosis during the phase of marrow hypocellularity/monocytopenia and pursuing hsct prior to malignant transformation may improve patient outcome. introduction/background: reduction in child mortality has kept pace with improved immunization and nutritional status. however, there are children with severe infections who have no identifiable reason. the search for children with pid was initiated in a tertiary care referral hospital from a region with no documentation of the prevalence of this disorder, but with a high rate of consanguinity. financial constraints, poor availability of laboratory facilities or therapeutic options locally, limited awareness among pediatricians and vast distances between premier centres were major obstacles. objectives to identify children with primary immune deficiency and study the spectrum of pid in north kerala to prevent infectious and other complications in these children to provide curative therapy whenever feasible methods . acquisition of knowledge and skills to diagnose and manage pids . establishment of an immune deficiency clinic . liaison with centres across the country offering diagnostic tests and stem cell transplant . formation of project team and submission of a project to the central government . improvement of diagnostic facilities at home institution results: children with recurrent, severe or persistent infections or with two or more esid warning signs were screened for pid. severe combined immune deficiency was diagnosed in children, wiskott -aldrich syndrome in children, chronic granulomatous disease in children and x linked agammaglobulinemia in children and leucocyte adhesion deficiency in children. prophylaxis with ivig was initiated in children and stem cell transplant was done for children. conclusions: in a country with resource constraints, limited awareness among health care providers and vast distances, it is possible to make a difference to the lives of families of children with pid by networking across centers with expertise in immunological and molecular genetic diagnostic methods and life -saving therapeutic modalities like stem cell transplantation. introduction/background: complete thymic aplasia is a rare cause of scid, and requires thymic transplantation for curative treatment. because thymic transplantation is not widely available, there can be significant delay between diagnosis and curative therapy placing the infant at risk of invasive life threatening infections. objectives: describe modalities of therapy for adenoviremia in a patient with scid due to thymic aplasia. methods: chart review was performed. treatment including hlapartially matched third party cytotoxic t cell therapy and matched related hematopoietic cell transplant were assessed for treatment of life threatening adenoviremia in a scid patient with thymic aplasia. we identified an infant with a mutation in tbox transcription factor (tbx ) (c. _ dup ) by way of state newborn screening for scid. she had severe t cell lymphopenia and abnormal t cell function. at months age, she developed adenoviremia with associated fulminant hepatitis, and an initial viral load of million copies/ml. despite prolonged therapy with cidofovir, viral load increased to as high as million copies/ml. treatment with two infusions of partially hlamatched third-party cytotoxic t cells specific to adenovirus ( / and / hla matched, with hla class ii-mediated antiviral restrictions) led to partial clinical improvement without viral clearance. due to continued severe adenovirus-related hepatic dysfunction, unmanipulated bone marrow from an hla-identical sibling was infused without conditioning ( million cd + cells/kg and . x cd + cells/kg), at months of age. after an initial surge in adenoviral loads attributed to massive viral lysis, the degree of viremia progressively declined and was < , copies/ml within weeks of marrow infusion. antiviral tcell activity against adenovirus was detected at low level in peripheral blood via ifn elispot at weeks post-marrow infusion. she subsequently developed acute cutaneous and hepatic gvhd responsive to tacrolimus and steroids without recrudescence of viral illness. conclusions: delay in curative therapy in scid substantially increases risk of invasive life threatening infections. one strategy of allogeneic hct can be to eradicate severe infection in scid by providing the necessary t cell directed therapy against infectious agents. antigen-specific partial immune reconstitution can be achieved with hct in patients with thymic aplasia but concern regarding the development of full immunologic t cell diversity in athymic patients remains. senior technician, sanquin bloodsupply amsterdam introduction/background: chronic granulomatous disease (cgd), an inherited disorder of granulocyte function caused by a failure of intracellular superoxide production, normally presents as severe recurrent bacterial and fungal infections in the first years of life. the majority of affected individuals are diagnosed before the age of years, although patients may remain undiagnosed until adulthood despite the early onset of the symptoms objectives: investigation of fungal infection in adult cgd patients methods: nbt and dhr for detection of cgd and molecular analysis for detection of type of mutation in cgd and fungal characterization. results: we report here the detection of causative fungal infections in adult patients with cgd. in the first patients we found paecilomyces formosus infection in an adult patient ( years old female) with undiagnosed cgd who was referred to the shahid beheshti university hospital (tehran, iran) complaining of cough, dyspnea and fever for weeks prior to admission. microscopic analysis revealed branching solitary phialide with ellipsoidal conidia with long chain arrangement and many chlamydospores which mostly resemble botryotrichum species but during subcultures on potato dextrose agar (pda) and sabouraud dextrose agar (sda), phialides typical of paecilomyces species appeared. typically, paecilomyces spp. rarely cause infections in humans and if these fungi are detected in blood urine or cerebrospinal fluid cultures they are considered as contaminants. the second patient was a -year-old man was referred to the hospital with weight loss, fever, hepatosplenomegaly and coughing. he had previously been diagnosed with lymphoadenopathy in the neck at age and prescribed antituberculosis treatment. bal and serum galactomannan tests were negative. low, subnormal levels of ros were produced following stimulation of purified neutrophils with the phorbol ester pma. genomic dna was extracted and gene scan was used to determine the ratio between the number of exon sequences of neutrophil cytosolic factor (ncf ) gene, which encodes p phox, and the number of -ncf exon sequences. in addition, the fungal culture was disrupted with glass beads and dna was extracted. the dna sequence results were blasted using the ncbi genebank database, which showed % similarity to an aspergillus terreus isolate in the gene bank fungal library with accession no . conclusions: thus, despite its current relative rarity in older patients, the presence of fungal infection is changing our understanding of the diagnosis, management and outcome of cgd. greater appreciation of the potential of fungal infection in older cgd patients is important in regions, such as iran, where tuberculosis is endemic and that sarcoidosis and cgd are considered as differential diagnosis. the demonstration of the successful patient-orientated treatment after using sequencing to confirm cgd and to identify the presence of the specific infectious agent emphasises the importance of adopting this approach across the region. introduction/background: primary immunodeficiencies (pi) represent a heterogeneous group of over genetically distinct disorders which interrupt normal host-defense mechanisms and predispose to significant morbidity and mortality. presently, we are only able to screen for severe t-cell deficiencies at birth; however, the most common forms of pi often go undetected for years leading to adverse patient outcomes and excessive healthcare spending. given the relatively high incidence of pi in the general population, informatic measures could be useful for determining individual risk for pi and facilitating earlier, correct diagnosis and appropriate treatment across the spectrum of pi. objectives: the purpose of this study was to test the jeffrey modell foundations spirit (software for primary immunodeficiency recognition intervention and tracking) analyzer on the texas childrens health plan. major aims were to identify individuals with medium-high risk for pi, assess the clinical characteristics of at risk patients and determine if risk identification led to diagnosis of pi over a month period. methods: after removing all known pi diagnosed patients from the database, , individual texas childrens health plan enrollees were screened for risk of pi with the spirit analyzer using relevant, weighted icd and icd codes. patient characteristics are shown in table . following identification of medium-high risk (mhr) individuals, letters were sent to their primary care physicians to alert them of patient risk. a second analysis of the mhr individuals was performed months later. detailed chart reviews were conducted on mhr individuals to further assess clinical features of this group. the study was approved by the baylor college of medicine institutional review board (study h- ). results: of the original cohort, ( . %) were identified as mhr for pi. from that group, ( . %) were accessible for analysis and ( . %) had electronic health records for review. in the months following the first analysis, ( . %) were diagnosed with a pi. (figure ) another patients had concerning diagnoses coded warranting further investigation. (figure ) concerning diagnoses included: cellulitis( ), abscess ( ), recurrent otitis media( ), recurrent sinusitis ( ), osteomyelitis ( ), and mastoiditis ( ) among others. in total patients had a pi diagnosis or a history concerning for pi ( . % of main cohort; . % of mhr cohort). conclusions: the spirit analyzer is effective at identifying persons at risk for pi and facilitates diagnosis. potential mhr yield by the analyzer is over %. these patients are treatable and will benefit from targeted intervention once identified. the analyzer can also highlight concerning conditions worthy of additional assessment. future work should focus on longitudinal healthcare outcomes for patients diagnosed with pi via spirt and physician perspectives on the utility of the tool. introduction/background: genetic variants in card contribute to several diseases caused by dysregulation of the adaptive immune system. dominant-negative, loss-of-function and gain-of-function variants in card variants lead to primary immunodeficiency disease. however, primary immunodeficiency disease caused by card gain-of-function variants often progresses to b-cell malignancy. the clinical course and treatment options depend on the type of card mutation. unfortunately lymphocyte immunophenotyping and traditional proliferation assays can't distinguish the variant effect or predict the likelihood of malignancy. objectives: to use multiplexed functional assays for determining variant effect to distinguish between dominant-negative, loss-and gain-offunction effects in card . our ultimate goal is to generate a variant function map that can be used to guide diagnoses and treatment of immune dysregulation caused by mutations in card . methods: we co-delivered crispr/cas ribonucleoprotein complexes with libraries of single-stranded oligonucleotide repair templates thus generating lymphoma cell populations containing all possible singlenucleotide variants (~ different protein coding changes) in the nterminal amino acids of card . following culture with and without bcr pathway inhibitors, we used next generation sequencing to quantify variant abundance before and after culture, both with and without bcr pathway inhibitors. results: due a requirement for card in these lymphoma cells, those with dominant-negative and loss-of-function card variants grow more slowly, whereas those with gain-of-function variants grow faster in the presence b cell receptor pathway inhibitors. by tracking the relative abundance of each variant in the population by next generation sequencing over multiple conditions, we determined the functional effect of each. we assessed the functional effects of thousands of card variants in parallel. this enabled us to confirm several previously reported gain-offunction and dominant-negative variants in card , as well as identify several additional novel variants. finally, we evaluated previously undescribed dominant-negative, loss-of-function and gain-of-function variants during differentiation of primary human b cells and during nf-b signaling. conclusions: the results of our experiments demonstrate the utility of multiplexed functional assays for determining variant effect in proteins where distinguishing between dominant-negative, loss-and gain-offunction effects are required to guide diagnoses and treatment. introduction/background: primary immune-deficiencies (pids) are a heterogeneous group of nearly monogenic inborn errors of immunity. in recent years, whole exome sequencing (wes) became a valuable diagnostic approach for the identification of molecular defects in patients with clinical manifestations suggestive of pid. this approach provides a definitive diagnosis and may help in genetic counseling, prenatal diagnosis and pre-symptomatic identification of patients with a potentially lethal disease. the diagnostic yield using wes was found to be~ % in rare mendelian disorders and~ % in pids. we present here a markedly higher yield, of %, in pids with a high percentage of consanguinity ( % in this study). objectives: using the whole exome sequencing (wes) approach in israeli pid patients with high percentage of consanguinity to identify the genetic causes underlying their diseases for better diagnosis and clinical management. methods: wes was performed on genomic dna obtained from wbc of immune deficient patients with potential genetic causes. the sequencing data was analyzed bioinformatically. each of the discovered mutations was validated and the familial segregation was confirmed, using sanger sequencing. results: the probands ( males and females) ranged in age from weeks to years with a mean of . years. of them, patients are jewish ( %), palestinians ( %) and ( %) is from a greek ethnicity (cyprus). based on their clinical and immunologic phenotype at the time of initial evaluation, patients were assigned to one of seven pid groups: (i) humoral immunodeficiency patients ( %); (ii) combined immunodeficiency (cid) ( %); (iii) cid with syndromic features ( %); (iv) scid ( %); (v) congenital defects of phagocytes ( %); (vi) immune dysregulation ( %) and (vii) phenocopy of pid ( %). we identified mutated alleles, all in coding regions, that are highly likely to be causative in of the patients, achieving a % molecular diagnostic yield. among the patients, the mode of inheritance in patients ( %) is autosomal recessive and in is compound heterozygote ( %). four patients ( %) harbor a non-inherited mutation on one allele, either de novo or somatic. the inheritance of the mutation in one patient ( %) is x-linked. the high rate of bi-allelic inheritance ( % of the alleles) is mainly due to the high frequency ( %) of consanguinity among the studied cohort. twenty eight mutated genes were identified in this study. of them, were found to be novel in causing an inherited disease in man. interestingly, some genetic defects in known genes were found in patients with atypical phenotypes. in patients ( % of the total number of patients and % of the wes diagnosed) the discovery of the genetic cause led to a change of therapy, towards a more targeted and personalized one. the revised treatments included bone marrow transplantation, conditioning protocols, reduced intensity of immune suppression, and prevention of unnecessary treatments due to their possible deleterious outcome. conclusions: except of being a useful tool for diagnosing and deciphering novel or atypical forms of pids, our wes study demonstrates an immediate and powerful impact on patient therapy in pids. early intervention with bone marrow transplant or gene therapy is critical and best when the infant in uninfected. newborn screening for scid and t-cell lymphopenia has been implemented in states. the screening test is performed from dried blood spots collected at birth and involves pcr quantification of circular dna byproducts of t-cell receptor gene rearrangement, t-cell receptor excision circles (trec). trecs are generated during t-cell maturation in the thymus and are indicative of naïve t-cell output. assays and protocols to measure trecs vary by state and there is no standardized guideline at this time. washington state is unique in that it is one of few states where all newborns undergo two or more independent newborn screens, the first by which igrt treatment helps in these patients is unreported. here we present the first case of an mbl deficient patient with other complicating conditions (low igg and multiple cf polymorphisms, ciliary dyskinesia) who did not show improvement on igrt, thus refuting current literature. results: a -year-old caucasian male patient presented due to recalcitrant warts of the hands and feet. he had childhood otitis media status post tympanoplasty, mild molluscum contagiosum, and eczema. also, history of migraines with a negative brain mri. a trial of weekly pegylated interferon alpha mcg for his warts was discontinued due to significant neutropenia and depression. physical exam was notable for bulky skin colored warts on the lateral and dorsal fingers and dorsal hands including periungual skin. clusters of verrucae were noted on the feet. initial laboratory testing was notable for mild hypogammaglobulinemia with protective specific antibodies. lymphocyte subset analysis revealed a predominantly t cell lymphopenia with decreased naïve and memory cd and cd subsets, normal absolute numbers of b cells but with low memory subsets, and normal numbers and function of nk cells. whole exome sequencing ultimately revealed homozygous r q mutations in the cecr , a mutation previously described as causing ada deficiency. ada activity was about %. conclusions: ada deficiency is a relatively newly defined genetic defect, with a clinical phenotype that continues to evolve with newly diagnosed cases. our patient had not had evidence of vasculitis or stroke, but had recalcitrant warts with lymphopenia as his primary presentation. approach to therapy for those without vasculitis or significant cytopenias remains unknown. introduction/background: whole exome sequencing has added greatly to our library of primary immune deficiencies. however, interpretation of findings is not always straightforward. clinicians need to understand the limitations of this diagnostic tool and be familiar with the next steps in order to achieve a diagnosis. objectives: a year old male presents for evaluation of bronchiectasis of bilateral lower lobes and poor growth. past medical history was significant for gerd with poor weight gain during infancy and childhood, oral thrush, recurrent respiratory infections and bilateral partial hearing loss. the bal showed abundant neutrophils and was positive for strep pneumoniae and haemophilus influenza. methods: serum immunoglobulins were normal with an elevated iga ( ) and mildly elevated ige ( ). tetanus igg was protective ( . iu/ml) but post-vaccine responses to the pneumococcal polysaccharide vaccine were poor with protective titers achieved to / serotypes. lymphocyte phenotyping was remarkable for a complete absence of b and nk cells. cd :cd ratio was inverted ( . ) and the majority of the cd + t cells were memory phenotype. mitogen proliferation was essentially normal. due to the absence of b cells and despite normal total serum immunoglobulins, the patient was started on sc ig weekly and antibiotic prophylaxis with trimethoprim-sulfamethoxazole and fluconazole. clinically he demonstrated good weight gain and a reduction in cough and respiratory symptoms. results: whole exome sequencing was performed. a single base mutation in ada (c. t>c) was identified in one allele. this mutation (p.l p) is known to be deleterious and when homozygous is associated with typical scid. however, this mutation was not present on the other allele. secondary analysis looking for large deletions and duplications failed to identify any abnormality in sequence in the normal allele. but the expression of this allele was markedly diminished causing us to suspect that its function might be impaired. functional testing was performed and demonstrated that there was no ada activity in the patients red blood cells, thus confirming a diagnosis of combined immune deficiency due to ada deficiency. conclusions: null mutations in ada result in an absence of ada function with profound cd cell lymphopenia and dysfunction. clinically affected infants have typical scid. hypomorphic mutations may lead to partial function of ada with more variable immune defects. interestingly, most patients with ada deficiency have neurologic complications, frequently hearing loss. we believe that most likely this patient has had some degree of spontaneous reversion of the mutation in the normal allele leading to a less severe phenotype. as reported previously in a case involving a mutation in il rg chain, normal function was not restored and the patient remains with a combined immune deficiency. this case highlights an important limitation of whole exome sequencing and the need for confirming the impact of a genetic defect with a functional assay. introduction/background: the autosomal dominant hyper-ige-syndrome (ad-hies) is a rare primary immunodeficiency and multisystem disorder resulting from heterozygous loss-of-function mutations in the stat gene. ad-hies is characterized by skeletal dysplasia, recurrent pulmonary and skin infections (e.g. staphylococcal abscesses, eczematoid dermatitis) due to an increased susceptibility to bacteria and fungi. since many patients do rather well on anti-infective prophylaxis and supportive care, and early case reports suggested no benefit of allogeneic hematopoetic stem cell transplantation (hsct), ad-hies patients are rarely referred for hsct. the literature still contains only a handful of patients who underwent hsct. all these patients had experienced severe disease related complications before hsct and consequently the benefit of hsct was reported to be variable. objectives: currently, the general consensus is to only consider patients with severe pulmonary disease for hsct. it could however be postulated that transplanting patients earlier in their disease course and correcting their immunodeficiency before permanent organ damage due to infectious complications has occurred may extend life-expectancy and improve the quality of life of ad-hies patients. methods: we report on a year old female ad-hies-patient who presented with bronchiectases following recurrent pneumonias, one pneumatocele, and normal pulmonary function tests. her past medical history also included recurrent skin infections, serious haemoptysis, pathological fractures and atopic eczema. considering that lung infections are the major life-limiting complication in ad-hies and are potentially positively influenced by hsct, our patient had enquired about a transplant. after extensive discussion of the pros and cons and obtaining full informed consent from her and her family, she underwent an elective hsct. she received bone marrow from an hla-matched sibling donor after a reduced-intensity conditioning consisting of alemtuzumab ( x , mg/kg), treosulfan ( x g/ m ), fludarabine ( x mg/m ) and thiotepa ( x mg/kg), and graftversus-host disease (gvhd) prophylaxis with cyclosporine a (csa) and mycophenolate mofetil (mmf). results: the peri-transplant course was complicated by acute lower gastrointestinal tract bleeding and renal failure of unknown origin. continued kidney function impairment led to early tapering and discontinuation of csa on day + after hsct in the absence of any acute gvhd. neutrophils and platelets engrafted on days + and + respectively. she is currently on day + , free of gvhd or infection, exhibiting full donor chimerism and recovered kidney function. in view of the preexisting pulmonary damage she currently remains on antibiotic prophylaxis and inhalation therapy. conclusions: this ad-hies patient who underwent hsct with few pre-hsct disease complications and relatively little permanent organ damage may add to our understanding of whether early hsct will lead to improvement of quality of life and possibly increased life-expectancy in ad-hies patients. it remains to be elucidated whether her rather uncommon peri-hsct complications are connected to her underlying disease. future research should be directed at identifying ad-hies patients at high risk of severe pulmonary complications early, so these could be referred for timely hsct. objectives: this is a case study of a patient who had refractory ibd symptoms and recurrent infections who was found to have xiap deficiency and mefv variant mutations. methods: a year old male presented at age with recalcitrant ibd unresponsive to multiple medications including steroids, mesalamine, azathioprine, infliximab, adalimumab, methotrexate, and vedolibumab. in addition, he developed severe infections from a combination of immune dysregulation and immunosuppressant medications. currently he is on ustekinumab but still has severe abdominal symptoms. patient does not have a history of hemophagocytic lymphohistiocytosis (hlh). family history was significant for ibd in both his mother and sister. he has had persistent lymphopenia which ranged between to cells/ mm . t/b/nk panel showed decreased cd t cells ( cells/mm ) with normal cd , cd , and cd /cd ratios. b and nk cells were normal in quantity. t cell antigen/mitogen assays showed normal response to all mitogens (pha, pwm, con a) and most antigens (tetanus, candida, hsv, vzv, adv) but low response to cmv antigen. igg was elevated at , , but iga, igm, and ige were normal. his ebv dna pcr is negative. results: exome sequencing revealed a novel xiap hemizygous variant at position c. g>a (p.=?) of unknown significance and a mefv heterozygous variant. functional testing performed at medical college of wisconsin showed no expression of xiap on lymphocytes and a defect in nod pathway. given the xiap deficiency, bone marrow transplant was discussed as an option for refractory ibd and prevention of hlh. rituximab was also offered to decrease the possibility of hlh. currently the patient is in the decision making process of both treatment options. conclusions: genetic evaluation with clinical exome in early onset refractory ibd, family history of ibd, and recurrent infections demonstrated a novel mutation in the xiap gene with possible contribution of mefv variant as well. the absence of xiap protein expression and abnormal functional assay of the nod pathway confirm the pathogenicity of the mutation. identification of this genetic variant will help guide future therapeutic options and prognosis for this patient. introduction/background: x-linked agammaglobulinemia (xla) is a primary immunodeficiency disease caused by mutations of bruton tyrosine kinase (btk), which is essential in b cell maturation. xla is typically associated with bacterial infections of upper and lower respiratory systems, enteritis and increased risk for malignancies. objectives: to present the evolution and treatment of a complicated flexispira (helicobacter bilis) infection with delayed diagnosis in an xla patient. methods: clinical and laboratory features of a patient with xla evaluated at the national institutes of health. results: a -year-old male patient with xla presented for initial evaluation of indurated lower extremity and torso lesions. he was diagnosed with xla at age years secondary to bacterial sepsis pneumonia and empyema. after starting ivig at age , he was well without significant infections except for recurrent otitis media. at years of age, he developed right leg edema below the knee, which progressed to patchy skin thickening and discoloration. a tissue biopsy at years of age revealed marked fibrous thickening of the subcutaneous septum with diffuse infiltrate of eosinophils with negative cultures for bacterial, fungal, and mycobacterial infections and thought to be consistent with eosinophilic fasciitis. mri demonstrated infiltration in the superficial and deep muscle compartments with fibrosis. symptoms persisted despite empirical treatment with iv antibiotics and steroids. at years of age his left leg became involved with similar findings, while under treatment including multiple immunosuppressants (dapsone, methotrexate, tacrolimus, hydroxychloroquine, iv cyclophosphamide, remicade, cytoxan, and enbrel) targeting eosinophilic fasciitis. at age , he developed ulcerations over the left shin and ankle and was diagnosed with chronic multifocal osteomyelitis based on mri findings. physical exam was notable for bilateral leg swelling below the knees with woody appearance and induration, hyperpigmentation, and tenderness. indurated and nodular lesions without discoloration were noted above the waist line, on right forearm and above right nipple. skin biopsies of right lower extremity and right forearm were positive for numerous spirochetal-like organisms with warthin-starry stain. treatment was initiated with meropenem and gentamicin. gentamicin was discontinued due to vestibular ototoxicity six weeks later and doxycycline was added. initial response was followed by worsening of symptoms and intolerance to treatment, which led to the addition of tigecycline and azithromycin. with continued progression, seven months into initial evaluation, chloramphenicol and nitazoxanide were added to the regimen. due to the persistence of flexispira organisms on skin biopsy warthin-starry stains, fresh frozen plasma (ffp) was introduced four months later ( months after initial evaluation) as an adjunctive treatment. cultures performed at the cdc were negative; however, pcr and sequencing resulted in identification of helicobacter bilis. units of ffp was infused weekly for over three years, then reduced to every two weeks, with goal igm levels > mg/dl. subsequently, labs including cytopenias, inflammatory markers, and immunoglobulins improved as well as a negative warthin-starry stain. symptoms have improved with almost complete resolution of findings with only residual small areas of discoloration over both lower extremities. conclusions: xla immune deficiency is associated with flexispira (helicobacter bilis) infections, with typical appearance of discoloration and induration, which may evolve to osteomyelitis due to delayed treatment. although typically observed over the lower extremities, immunosuppressive treatment may lead to further expansion above the waist line. approach to therapy with weekly to bimonthly ffp infusions in addition to antibacterial treatment has proven to be beneficial in controlling the infection. higher igm levels resulting from the ffp may also provide antibacterial effects. introduction/background: x-linked severe combined immunodeficiency (scid) is a well described primary immunodeficiency associated with mutations in the common gamma chain. patients with x-linked scid classically present with profoundly low or absent t cells and nk cells with a variable number of b cells. the lymphocytes that are present typically have a proliferation index < % control when stimulated with mitogens and antigens. patients must undergo corrective therapy with bone marrow transplant (bmt) or gene therapy to avoid the life-threatening infections that are associated with the nearly absent adaptive immune system. objectives: a -week-old boy presented to childrens national immunology clinic for initial evaluation of critical result on newborn screen. methods: targeted partial exome sequencing was performed on a -dayold patient who was picked up via trec assay on the maryland newborn screen. flow cytometry was completed at childrens national and proliferation studies completed at cincinnati childrens hospital diagnostic immunology lab. results: flow cytometry revealed markedly decreased lymphocytes with nearly absent cd + t cells and low cd + t cells with a r e l a t i v e i n c r e a s e i n c d + c d r o t c e l l s ( r a t i o cd ra:cd ro: %: %). the b and nk cells were within the reference range for age. mitogen proliferation studies showed a mild decrease to pha and normal responses to pokeweed and cona ( cpm, cpm, and cpm, respectively). partial exome sequencing revealed a hemizygous nonsense substitution in il rg (c. c>t, p.r ) . maternal engraftment accounted for % of the t cells. the patient was started on prophylaxis with ivig, bactrim, and fluconazole with the plan to proceed with bone marrow transplant. as patient approached bmt maternal engraftment became absent in whole blood and repeat proliferation studies revealed normalization of the response to pha (stim index) with continued normal responses to cona and pokeweed. the patients flow cytometry values and ratios remained unchanged. patient completed a reduced intensity preparative regimen of busulfan, fludarabine and alemtuzumab prior to receiving his / matched unrelated donor bone marrow transplant. conclusions: it remains to be determined why initial proliferation studies showed > % function with improvement over time in a patient with a well-described genetic mutation causing scid ( ) director, division of intramural research, nih/niaid introduction/background: the advent of next-generation sequencing (ngs) has led to a proliferation of newly discovered genetic diseases and expanded phenotypes of known immunodeficiencies. availability of specific gene panels or whole exome sequencing (wes) with targeted analysis based on broad phenotypes, coupled with clinicians increasing awareness has led to higher utilization of ngs. published reports from high throughput sequencing labs indicate exome analysis identifies causative mutations in only - % of probands. results: we have seen several patients who underwent high quality ngs in whom causative mutations were not identified. two separate families with multigenerational histories of leukemia, aplastic anemia, myelodysplastic syndrome, and cytopenias suggestive of gata deficiency had myeloid gene panel screens at commercial labs without causative mutations identified. targeted gata sequencing in the first family identified a novel change in gata , c. g>t, p.l l. cdna analysis demonstrated this synonymous variant resulted in aberrant splicing leading to a frameshift and premature termination. the wes bioinformatics pipeline failed to recognize the splice mutation. in the second family, pcr amplification spanning the terminal exons revealed a shortened pcr product with a base deletion fully encompassing the penultimate exon and leading to a amino acid in-frame deletion. the deletion spanned all capture probes for the exon resulting in only the wild-type allele being captured and sequenced. additionally, capture kits targeting only coding regions of genes fail to capture deep intronic mutations such as those seen in gata (hsu, ) or in the untranslated region of ikbkg, encoding nemo, (mooster, ; hsu, submitted) . lastly, even with good capture and sequencing, the presence of pseudogenes may confound downstream sequence alignment as seen in ncf , encoding p phox preventing recognition of disease causing mutations. conclusions: with ngs becoming more widely available as a clinical diagnostic tool, it is important to remember that wes results, unlike many laboratory tests, are not binary. inadequate bioinformatics pipelines, deletions, intronic or untranslated mutations, and pseudogenes can all mask the presence of causative mutations. targeted panel captures or analysis will miss novel genes. astute clinicians need to recognize the limitations of the current technology and pursue alternate assays when suspicions warrant. a cell based assay for the detection of autoantibodies to il- in human serum. matt phillips, phd , vijaya knight, md, phd senior scientist, national jewish health director of immunology and complement adx labs, national jewish health introduction/background: patients with chronic candida infections are typically deficient in some aspect of il- signaling. one mechanism, which has recently come to light, is through the production of il- autoantibodies, particularly in patients who already suffer from specific autoimmune diseases. objectives: our objectives are to develop a diagnostic assay to accurately and easily detect il- autoantibodies in patient serum. methods: we developed a cell-based reporter assay using hek blue il- cells (invivogen) to detect the ability of patient serum to block il- receptor signaling. once stimulated with il- , the cells secrete alkaline phosphatase (ap) into the surrounding media which is detected by invivogen hek blue media and an absorbance reader. addition of serum containing blocking antibodies inhibits secretion of ap. results: we were able to demonstrate, using a single patients serum with known il- autoantibodies, the inhibition of il- signaling in hek blue il- reporter cells. we further characterized the sensitivity of our assay with a commercially available anti-il- monoclonal and found it to be sensitive to between . x - m and . x - m. conclusions: loss of il- signaling can lead to problematic immune deficiencies including difficulty in clearing extracellular pathogens such as candida. some people who have an immune deficiency in the il- pathway may have developed autoantibodies to il- and thus have difficulty generating an appropriate immune response. we have developed a relatively low maintenance, cost effective, and simple test for detecting il- autoantibodies in human serum. alternations in repertoire of t and b cell subsets in patients with partial recombination activating gene (rag) deficiency with autoimmunity and history of viral infections introduction/background: patients with partial deficiency of recombination-activating genes or (rag / ) can present with a wide spectrum of primary immunodeficiencies including combined immunodeficiency with granuloma and/or autoimmunity (cid-g/ai). prior case reports have highlighted alterations in b and t cell compartments; however comprehensive characterization of t and b cell receptor repertoires of lymphocyte subsets regarding diversity and autoreactivity has not been reported. objectives: defects in v(d)j recombination due to rag deficiency results in a skewed t and b cell repertoire that may be further modified by viral infections and promote inflammatory or autoimmune phenotype. methods: peripheral t and b cell compartments were sorted from two patients with combined immunodeficiency secondary to hypomorphic rag and rag mutations. b cells were stimulated with cd l, cpg and il- to transition to antibody secreting cells (ascs), mimicking viral infection. repertoire analysis and single cell cloning of bcr heavy and light chain variable regions from sorted b cell populations has been performed. repertoire of t cell subsets (treg and follicular helper) were also examined results: we noted skewing towards proximal j usage in all b cell compartments (mature naïve, marginal zone, cd -/low and memory) of two rag deficient patients compared to healthy controls. b cell clones with v - v genes with low rate of somatic hypermutation expanded during b cell development. after in vitro stimulation mature naïve b cells from rag deficient patient were capable of transitioning to antibody secreting cells and enriched for polyreactivity to dsdna, insulin, lps and ifn cytokine ( to %) compared to healthy control ( to %). in connection to altered b cell compartments, restricted repertoire of regulatory t cells and an expanded and skewed follicular helper t subset were detected. conclusions: our data indicate that patients with partial rag deficiency have skewed t and b cell subsets that can further be altered towards antibody secretion and polyreactivity after stimulation such as viral infections. chief of the division of allergy and immunology, division of allergy immunology, the childrens hospital of philadelphia, philadelphia, pa usa introduction/background: the clinical features of q . deletion syndrome include virtually every organ of the body. t cell lymphopenia, as a consequence of thymic hypoplasia, is the most commonly described immunologic feature and is most prominent in childhood. later in life, t cell exhaustion may be seen and secondary deficiencies of antibody function have been described in patients with q . deletion syndrome. objectives: the role of deletion breakpoints in determining q . deletion syndrome immunophenotype is unknown. in this study, we examined the effect of q . deletions with and without tbx on lymphocyte counts. methods: lymphocyte counts were compared between total q . patients with tbx -containing deletion (a-b, a-c, a-d deletions), and a total of patients with tbx -noncontaining deletion (b-d, c-d, d-e, d-f deletions). lymphocyte counts of patients with q . deletions were compared to a set of patients with a q . duplication including the tbx locus. lymphocyte subset counts for each group were analyzed by t-test. results: cd counts were significantly lower in the tbx -deleted cohort compared to the other two cohorts (mean cells/mm in tbx deleted cohort, cd cells/mm in the non-tbx deleted cohort, and in the duplication cohort, p< . for all). similarly, cd counts were lower in the tbx -deleted patients compared to the other two cohorts. there were no significant differences in cd , cd , and nk cell counts between the three cohorts. conclusions: these represent the first data to examine t cell counts in q . deletion syndrome patients with different breakpoints. our data highlights an important role for tbx or other genes in the a-b region in regulating t cell production. paracoccidioidomycosis associated with a heterozygous stat mutation and impaired ifn-immunity introduction/background: mutations in genes affecting ifn-immunity have contributed to understand the essential role of this cytokine in the protection against intracellular bacteria and fungi. however, inborn errors in stat , which controls il- responses, have not yet been reported. objectives: to determine the underlying genetic defect in a family with a history of paracoccidioidomycosis (pcm) disease. methods: genetic analysis was performed by whole-exome sequencing and sanger sequencing. stat phosphorylation and translocation from the cytosol to the nucleus, as well as ifnrelease by patient lymphocytes were assessed. the effect on stat function was evaluated by site-directed mutagenesis using a lymphoblastoid b cell line (b-lcl) and u a cells. microbicidal activity of patient monocytes/macrophages was also analyzed. results: a heterozygous missense mutation, c. a>t (p.e v) in stat was identified in the index patient and her father. patients and fathers lymphocytes showed reduced stat phosphorylation and nuclear translocation as well as impaired ifn-production. in accordance, b-lcl and u a cells carrying the stat mutant displayed reduced stat phosphorylation. patient's and father's pbmcs and macrophages (alone or in the presence of t cells) displayed impaired fungicidal activity compared with those from healthy controls that improved in the presence of recombinant human (rh) ifn-, but not rhil- . conclusions: our data suggest autosomal dominant stat deficiency as a novel inborn error of il- -dependent ifn-immunity associated with susceptibility to pcm disease. profound b cell lymphopenia in gof-stat that improves post ruxolitinib associate professor, university of south florida -johns hopkins all childrens hospital introduction/background: subjects with gain of function signal transducer and activator of transcription (gof-stat ) mutations have a variable clinical phenotype including combined immunodeficiency (cid). ruxolitinib, a janus kinase / inhibitor has been successful at treating immune dysregulation in subjects with gof-stat . two subjects with profound b cell and/or t cell lymphopenia as a major manifestation are described, one of which was successfully treated with ruxolitinib. objectives: to discuss gof-stat mutations, their effect on the immune system, and the potential benefit of ruxolitinib in these subjects. methods: retrospective chart review was performed. results: subject (c. a>g) is a year old male with a history of recurrent shingles, chronic mucocutaneous candidiasis (cmc), pneumocysitis jiroveccii pneumonia, varicella zoster meningitis, severe enteropathy, cerebral aneurysm and lymphoproliferation, and autoimmune hypothyroidism. he has profound lymphopenia predominantly affecting t and b cells (cd + cells/ul, cd + cells/ul, cd + cells/ul, cd + cells/ul, cd + cells/ul). subject (c. g>t) is a year old male with a history of severe cmc, recurrent pneumonia, enteropathy, and autoimmune thyroiditis. he had severe b cell lymphopenia (cd + cells/ul, cd + cells/ul, cd + cells/ul, cd + cells/ul, cd + cells/ul). treatment with ruxolitinib . mg bid led to clinical improvement of enteropathy and increased b cell counts in subject (cd + cells/ul). ruxolitinib has not yet been initiated in subject . both subjects were treated with anti-microbial prophylaxis and immunoglobulin supplementation. conclusions: combined immunodeficiency with variable degrees of b and t cell lymphopenia and hypogammaglobulinemia can be profound in subjects with gof-stat mutation. despite proper anti-microbial prophylaxis, this immunodeficiency can lead to severe infections. in addition to treating the autoimmune and immune dysregulatory features, treatment with ruxolitinib can improve the cid present and potentially reduce infectious susceptibility. igg -related disease (igg -rd), its common mimickers and response to anti-il -(reslizumab) treatment rachel eisenberg, md , arye rubinstein, md-phd fellow in allergy and immunology, montefiore medical center chief division of allergy and immunology, albert einstein college of medicine and montefiore hospital, bronx, ny, usa introduction/background: we describe a complicated case of igg related disease (igg -rd) both in its presentation and novel treatment objectives: to review the common mimickers of igg -related diseases which often lead to delayed diagnosis and treatment. to discuss novel therapeutic treatments for igg -related disease results: a -year-old woman with a history of thyroid disease, sicca symptoms, lipodystrophy, relapsing parotid enlargement, asthma and erdheim chester syndrome initially presented with recurrent bacterial and fungal sinusitis despite multiple sinus surgeries. immunologic workup was notable for lymphopenia of /ml, cd count of ( - cells/ul normal range) and elevated igg of ( . - . normal range). imaging was notable for nasal septal perforations and hypoplastic maxillary sinuses. there was high suspicion for igg disease however the patient was lost to follow up during which time she developed cachexia, eosinophilic pleural effusions ( % eosinophils), lung mass and a parotid mass with predominant t cell infiltrate misdiagnosed as follicular lymphoma. features consistent with igg -rd included > % ratio of igg /igg and a predominant t cell infiltrate a biopsied lung mass showed igg plasma cell > /hpf also consistent with igg -rd. bone marrow biopsy was within normal limits. the patient was treated with rituximab, an effective treatment for igg -rd. on treatment her igg levels normalized, however she developed recurrent large eosinophilic lung effusions requiring repeat drainage. fractional exhaled no (feno) was elevated to ppb. she was started on reslizumab at a dose of mg/ kg resulting in marked improvement in her respiratory status along with normalization of peripheral eosinophilia and reduction of feno to the normal level of ppb. conclusions: igg -rd is a fibro-inflammatory condition which can affect any organ system and is diagnosed via tissue histology showing igg positive plasma cells and a typical morphologic pattern. this case outlines the common mimickers of igg -rd often leading to a delayed diagnosis. before the final diagnosis of ig -rd was made by us, the patient carried multiple diagnoses including: thyroid disease, recurrent parotid enlargement and seronegative sjogrens. these diagnoses in hindsight may have been mikuliczs syndrome, kuttners tumor and/or riedels thyroiditis which are common manifestations of igg -rd. chronic sinusitis, atopic diseases, peripheral eosinophilia and destructive osseous lesions as noted in our patient are seen in up to % of patients with igg -rd. destructive bony lesions and eosinophilia can mimic granulomatous polyangiitis, which was ruled out in our patient. her cachectic appearance and diagnosis of lipodystrophy can be explained by destruction of osseous tissue in the craniofacial skeleton which was later confirmed on imaging. lymphoid inflammatory infiltrates are commonly seen in igg -rd and are can be misdiagnosed as a follicular lymphoma as in our case. a novelty in this case is the successful treatment with reslizumab targeted at the eosinophilic component of the disease. on reslizumab our patients asthma was for the first time controlled, pleuritis improved, fractional exhaled no (feno) normalized and her cachexia is improving. treatment with reslizumab should be considered in patients with igg -rd who manifest with eosinophilic respiratory disease. introduction/background: ikbkb deficiency (c. dupg in exon ) is a rare autosomal recessive form of severe combined immune deficiency (scid) originally described in canadian infants of northern cree descent. ikbkb scid is characterized by normal lymphocyte development, but impaired t-cell activation, along with innate immune defects. objectives: to report the clinical presentation, immunologic phenotype, and outcomes for patients with confirmed or suspected ikbkb scid due to this founder mutation. methods: we retrospectively reviewed hospital records dating back to of patients with confirmed homozygous ikbkb mutations, as well as patients suspected to be affected due to their clinical presentation and family relations to molecularly confirmed cases. results: fifteen patients were included. they presented early in life (average age months) with invasive and disseminated viral, bacterial, mycobacterial, and fungal infections. patients had concurrent and multiple infectious organisms, with a notable predilection for candida, gram negative organisms, and mycobacteria. four patients in our cohort received bcg vaccination at birth, resulting in fatal disseminated m. bovis infection in all, and additional patients succumbed to atypical mycobacterial infection following hematopoietic stem cell transplant (hsct). one newborn was identified in through new initiatives for targeted newborn screening for the mutation. immunologic features at presentation included normal to elevated lymphocyte counts with normal to elevated cd , cd , and cd t cells. when tested, response to pha varied from absent ( / ), to low ( / ), to normal ( / ). most patients had hypogammaglobulinemia, most often of the igg isotype ( / ). six had assessment of trec levels, and all had values above thresholds for screening programs. eight patients died before they could undergo hsct, and received transplants in the setting of ongoing severe, lifethreatening infections. only patient underwent hsct prior to the onset of infection. in our cohort, there are only long-term survivors. conclusions: ikbkb deficiency is a severe form of scid with early onset of invasive and disseminated multi-organism infection. the immunologic phenotype is characterized by normal to elevated lymphocyte numbers which do not meet pidtc criteria for scid, variable (and sometimes normal) mitogen response, normal trec levels, and low igg levels. the disease is universally fatal without hsct, however, conclusions regarding efficacy and long-term outcomes of hsct are uncertain given the small sample size. immune -dysregulation mimicking systemic lupus erythematosus in a patient with lysinuric protein intolerance introduction/background: lysinuric protein intolerance (lpi) is an inherited aminoaciduria caused by defective amino acid transport in epithelial cells of the intestine and kidney due to bi-allelic, pathogenic variants in slc a . the clinical phenotype of lpi includes failure to thrive and multi-system disease including hematologic, neurologic, pulmonary and renal manifestations. individual presentations are extremely variable, often leading to misdiagnosis or delayed diagnosis. here we describe a patient that presented with suspected immunodeficiency in the setting of early-onset systemic lupus erythematosus (sle), including renal involvement, who was subsequently diagnosed with lpi post-mortem. objectives: describe a clinical a patient with lysinuric protein intolerance that presented as early-onset systemic lupus erythematosus (sle), including renal involvement and primary immunodeficiency. methods: after informed consent was obtained, dna samples were obtained from the proband and his parents. trio whole exome sequencing was performed to identify a cause of early onset autoinflammation resembling systemic lupus erythematosus. results: the male proband had a history of failure to thrive starting at months of age, recurrent bacterial otitis media, and one episode of severe bacterial pneumonia requiring hospitalization. he presented at months of age with multifocal pneumonia, anemia (hgb . mg/dl) and mild thrombocytopenia. initial laboratory studies revealed low albumin ( . mg/dl), elevated ldh ( ), and mild hepatomegaly. renal and liver function testing was initially normal. immunologic evaluation for suspected primary immune deficiency showed normal immunoglobulin titers, low c ( ) and low c ( . ). lymphocyte phenotyping revealed low b cell counts ( . % of total lymphocytes) with t cells and nk cells within the normal range. despite antibiotic therapy, the patient worsened, developing fevers, a generalized erythematous rash, edema and nephrotic syndrome with oliguria. renal biopsy uncovered glomeruli with accentuated, global thickening and diffuse, peripheral capillary loops, as well as focal spiculated defects, there was endothelial swelling and other signs of acute damage including epithelial flattening, adluminal irregularity and extensive intraluminal proteinaceous detritus. endocapillary proliferative lesions, extracapillary crecents or tubular atrophy was not observed. immunofluorescence studies were positive for c , igg and c q granular deposits, mainly at the mesangium, interpreted as lupus nephropathy. endothelial swelling and massive, subepithelial electron-dense deposits with spike formation from the basement membrane were noted on electron microscopy, mimicking a stage ii membranous pattern of injury. autoantibodies included ana ( : ), anti-dsdna, smith, ssa and rnp were positive in agreement with the diagnosis of sle. immunosuppressive therapy with high dose iv corticosteroids and cyclophosphamide was initiated. despite this, the patient developed pancytopenia, elevated ferritin levels, increased triglycerides, and low fibrinogen. bone marrow biopsy displayed erythrocyte phagocytosis by macrophages, confirming a diagnosis of hemophagocytic lymphohistiocytosis (hlh). the patient subsequently died despite aggressive immunosuppression with high dose methylprednisolone and high dose iv immunoglobulin and dialysis. samples were collected from the deceased patient and his parents for research whole exome sequencing. trio analysis identified compound heterozygous missense variants in slc a . ammonia levels were not evaluated during the patients hospitalization. conclusions: lysinuric protein intolerance is a severe metabolic disorder that can present with protean systemic features including primary i m m u n o d e f i c i e n c y. i m p a i r e d l y m p h o c y t e f u n c t i o n , hypocomplementemia, immune-mediated glomerulonephritis, autoantibodies, and hlh are known complications of lpi. exactly how ineffective amino acid transport triggers these systemic inflammatory features is not yet understood. lpi should be considered in the differential diagnosis of early-onset sle, particularly in the absence of response to immunosuppressive therapy. director, national institute of immunohaematology introduction/background: chronic granulomatous disease (cgd) is a primary immunodeciency disorder with recurrent pyogenic infections and granulomatous inflammation resulting from loss of phagocyte superoxide production. mutations in any one of the five structural genes of the nicotinamide adenine dinucleotide phosphate (nadph) oxidase complex viz. cybb and cyba encoding for membrane bound gp phox and p phox; and ncf , ncf , and ncf encoding for cytosolic components p phox, p phox, and p phox respectively, have been found to cause cgd. the relative incidence of these gene defects varies significantly depending on the ethnic background of the population. identification of molecular defect is important for patient management as well as for prenatal diagnosis in the affected families. the present study was aimed at studying the pattern of underlying genetic defects in a cohort of indian patients affected with cgd. objectives .to identify the underlying genetic defect in patients with chronic granulomatous disease in india . clinical,immunological and molecular characterisation . to utilise this information for genetic counselling and prenatal diagnosis of the affected families methods: eighty-seven (n= ) patients with abnormal nbt and dhr were included in this study. in case of male patients, mothers were first screened for carrier status to rule out x-linked cgd (xl-cgd). those patients where mother is not showing mosaic pattern were suspected for autosomal recessive cgd (ar-cgd) and were screened for the ratio of ncf gene to pseudo ncf gene by genescan analysis. additionally, evaluation of nadph oxidase components expression by flow cytometry also helped us to determine the underlined genetic defect and it is validated by dna sequencing of respective genes. results: eighty-seven patients were molecularly characterized to identify disease causing mutation which includes: novel mutations in cybb, in cyba, in ncf gene in our cohort. . % (n= ) of the patients belonged to xl-cgd. . % (n= ) of the patients are suspected to have ncf gene defect among which; homozygous delgt mutation was identified in patients. . % and . % patients showed abnormal p phox and p phox expression suggesting defect in cyba gene and ncf gene respectively. spectrum of mutations involve: % of delgt mutations, % nonsense, % missense, % deletion, % insertion, % other than homozygous delgt mutations. male to female ratio is . : . consanguinity is noted in % of the patients. conclusions: despite the male predominance ar-cgd is more common ( %) as compare to xl-cgd ( %) in this cohort of indian patients, which is distinct from the western data. % are the novel mutations suggesting, a wide heterogeneity in the nature of mutations in indian cgd patients. flow cytometric evaluation of nadph oxidase component is used as a secondary screening test to identify cgd sub-group. molecular characterisation of cgd genes was not only used in the confirmation of diagnosis but also in genetic counselling and pre-natal diagnosis in affected families. novel nlrc gain-of-function mutation presenting with neonatal enterocolitis and autoinflammation, with positive clinical response to rapamycin and anakinra. senior investigator, viral immunology section, national institute of neurological disorders and stroke senior investigator, translational neuroradiology section, national institute of neurological disorders and stroke staff clinician, neuroimmunology clinic, national institute of neurological disorders and stroke staff clinician, laboratory of clinical immunology and microbiolology, niaid, nih introduction/background: cytotoxic t-lymphocyte antigen- (ctla- ) is an essential negative regulator of the immune response and its function is critical for immune homeostasis. uni-allelic mutation in the ctla gene leading to reduced function or expression of ctla- , termed ctla- haploinsufficiency, can lead to systemic immune dysregulation with wide spread clinical disease, but with variable clinical penetrance. the neurological manifestations of ctla- haploinsufficiency are not known. objectives: to perform detailed phenotyping of the neurological manifestations of ctla- haploinsufficiency. methods: a retrospective review and prospective collection of clinical, imaging, cerebral spinal fluid, and pathological specimens was performed in a cohort of genetically confirmed patients (n= ) with ctla mutations who are followed at the national institutes of health. neurological symptoms and exams were collected on patient visits and from historical records. the data collected included brain mris and spinal cord mris that were visually inspected for evidence of inflammation. cerebral spinal fluid values were obtained from patients including flow cytometry in patients. pathological tissue from brain biopsies of inflammatory lesions was examined from patients. results: central nervous system (cns) inflammation was found in / ( %) of the cohort. common clinical manifestations from the patients with cns inflammation were headaches / and seizure / . focal deficits were rare. mri findings included contrast enhancing neuroinflammatory lesions in the brain / , brainstem/cerebellum, and spinal cord / . figure a -c show representative inflammatory lesions. lesions were multifocal in / patients and / had recurrent inflammatory lesions on longitudinal follow up. lesions were, at times, extremely large, / with a lesion > cm^ . leptomeningeal enhancement (lme) was seen in / patients and clearly preceded intraparenchymal lesion development in patients. figure d shows a site of lme (green chevron) that develops into an intraparenchymal lesion (yellow chevron), mris are separated by days. spinal fluid analysis showed a lymphocytic pleocytosis (mean cells/mm^ ) with the presence of oligoclonal bands in patients. pathological features included a mixed cellular infiltrate, predominantly lymphocytes or plasma cells, with little evidence of demyelination or necrosis (figure e). conclusions: the neurological manifestations of ctla- haploinsufficiency include recurrent and, at times, severe neuroinflammation. however, even large lesions and lesions in eloquent anatomical locations had little to no focal clinical defects resulting in a striking clinical-radiological dissociation. future studies into the mechanisms of cns-related disease may reveal important information related to peripheral and central immune system functioning. conditioning with anti-cd immunotoxin in a mouse model of hypomorphic rag deficiency allows complete reconstitution of the immune system with lack of toxicity enrica calzoni, md , cristina corsino, technician , marita bosticardo, phd , yasuhiro yamazaki, md phd , hsin-hui yu, md phd , lisa ott de bruin, md , john manis, md , rahul palchaudhuri, phd , david scadden, md , luigi d. notarangelo, md treated mice and % in cd -sap/ treated mice. at sacrifice, in the bm we observed strong selective advantage for donor b cells at all developmental stages, but in particular in the most mature subsets, in both cd -sap and cd -sap/ treated mice, rescuing the block in development at pre-b cell stage found in untreated f l mice. donor engraftment in bm hsc reached levels around % and % in cd -sap and cd -sap/ treated mice, respectively. donor chimerism in t and b cells in the spleen was also higher than % in both cd -sap and cd -sap/ groups. in the thymus, full donor chimerism was achieved in both cd -sap and cd -sap/ treated mice, starting at the dn stage and persisting at dp, sp and sp . importantly, in both treatment groups, t cell development was corrected both in terms of subset distribution and absolute numbers to levels comparable to those of wt mice. finally, the thymic epithelial cell compartment was also fully reconstituted, with a normal number, distribution and maturation of both cortical and medullary thymic epithelial cells. conclusions: in conclusion, we show here that conditioning with cd -sap immunotoxin, alone or in combination with rads tbi, is safe and leads to full reconstitution of the immune system in rag hypomorphic mice, suggesting that this conditioning regimen should be considered for testing in clinical setting. introduction/background: foxp is a key transcription factor for the maintenance of immune tolerance. foxp mutations result in dysfunction of foxp + regulatory t cells (tregs) causing immune dysregulation, polyendocrinopathy, enteropathy, x-linked (ipex) syndrome, a severe early onset autoimmune disease, which can be fatal if not promptly diagnosed and treated. our recent international study analyzing the longterm outcome in i.e. patients of the two currently available treatments, pharmacological immune suppression and allogeneic hematopoietic stem cell (hsc) transplantation, showed poor long-term disease-free survival or overall survival limitations, respectively (barzaghi f. et al, jaci, ) . ipex syndrome is a good candidate for gene therapy as it has been demonstrated that reconstitution of wild-type treg cells can control the disease. however, foxp expression is highly regulated, and its safe and physiological expression in treg and teffector (teff) cells is challenging. lentiviral-mediated (lv) foxp gene transfer successfully converts ipex patients-derived cd + t cells into treg-like cells (cd lv-foxp t cells) with stable suppressive capacity (passerini l. et al, sci transl med, ) . these ex vivo converted tregs are ideal as a short term cell-based therapy for ipex patients, but this approach does not reestablish regulated foxp expression in teff cells, that also likely contribute to the ipex pathology. thus, we are further characterizing cd lv-foxp t cells and, at the same time, developing gene editing strategies for ipex, whereby autologous t cells or hscs are genetically modified or corrected, respectively, and reinfused into the patients. objectives: to provide more effective treatments for ipex patients, we are i) optimizing lv-foxp gene transfer in t cells to be suitable for clinical use, and ii) establishing a novel foxp gene editing in hscs and testing both approaches in preclinical models. methods: lv-foxp gene transfer can be obtained in cd + t cells activated polyclonally or in an antigen-specific manner. the vector construct is bidirectional, foxp expression is under the ef a-promoter and the truncated form of ngfr, used as marker gene, is under cmv promoter. foxp gene editing is performed using a combination of crispr/cas , a chemically modified sgrna targeting foxp , and an aav packaged homologous donor dna template and the efficacy and safety of the resulting construct is tested in different cell types in vitro and in humanized mice. results: we demonstrate that cd lv-foxp t cells can successfully be generated specific to different antigens. this result opens to new potential clinical benefit of cd lv-foxp t cells with more safe and specific regulatory effect than polyclonal cd lv-foxp t cells. we are currently adapting the protocol to optimal in vitro production for clinical use and assessing dose, survival and efficacy of the cd lv-foxp t cells using different in vivo models. due to the wide distribution of identified mutations throughout the foxp gene, we have designed a gene editing strategy that uses homology directed repair to insert the coding sequence of the foxp gene at the start codon of the endogenous mutated foxp gene. this strategy permits regulated expression of the inserted wild-type, functional foxp protein in patient cells independent of the location of the downstream mutation. using this site-specific gene knock-in, we find that the system effectively targets expression of foxp in different cell types, namely tregs, teff cells and primary human cord blood-or bone marrow-derived hscs. gene editing of normal donor and ipex tregs and teff cells allowed us to test for regulated gene expression and for establishment of normal treg suppressor function and t cell proliferation upon activation. additionally, preliminary results demonstrate that gene edited hscs can be transplanted into nsg mice for long-term reconstitution. conclusions: our results show the feasibility of different gene therapy approaches for ipex syndrome. in addition, they suggest that cd lv-foxp t cells, either polyclonal or antigen-specific, could be applied not only in ipex but also in immune mediated diseases of different origins. the results from the foxp gene editing support the use of crispr/ cas to treat ipex syndrome patients with autologous edited hscs. this gene editing approach may also be applied to treat other pediatric monogenic blood and immune disorders. human pi kgamma deficiency with humoral defects and lymphocytic infiltration of barrier tissues andrew takeda, bs , william comrie, phd , yu zhang, phd , paul tyler, bs , koneti rao, md , carrie l. lucas, phd graduate student, yale university clinician, niaid, nih assistant professor of immunobiology, yale university introduction/background: the phosphatidylinositol -kinase (pi k) signaling pathways play a key role in transducing signals from a diverse array of stimuli by producing the pip second messenger. class ib pi k is primarily activated by g protein-coupled receptors (gpcrs), and this class is comprised of the p gamma catalytic subunit in complex with the p or p regulatory subunit. in contrast to the class ia pi k subunits, inherited mutations in the genes encoding the class ib subunits have not been described. objectives: given the leukocyte-restricted expression pattern of p gamma, we hypothesized that mutations affecting this kinase may be found in cohorts of patients with rare immunodeficiency disorders. our objective was to identify such mutations and determine molecular, biochemical, and cellular derangements in patients with mutated p gamma. methods: we used whole-exome sequencing of families to identify inherited gene mutations and determined the mechanistic basis of disease using biochemical assays to assess effects on protein function and cellbased assays to define functional defects with disease relevance. results: we identified a patient (here called a. ) harboring compound heterozygous mutations in pik cg, the gene encoding p gamma, who presented in early life with autoimmune cytopenias and eczema and, at the age of years, developed cryptogenic organizing pneumonia and prominent t cell infiltration of the lungs. she also has a history of skin infections, lymphadenopathy/splenomegaly, eosinophilia, defective antibody production, and more recently, lymphocytic colitis. she inherited a frameshift pik cg mutation from her mother and a missense mutation resulting in an r p amino acid substitution from her father. expression of p gamma protein was lost, and stability of its p binding partner was reduced. despite defective t cell signaling responses to chemokines (i.e., gpcr stimulation), chemotaxis of patient t cell blasts in vitro was normal. intriguingly, the frequency of peripheral blood treg cells was low in patient a. , and her cd t cells more frequently expressed the tissue-homing cxcr chemokine receptor. consistently, serum levels of cxcr ligands were elevated in patient a. . moreover, we found augmented inflammatory cytokine production from m -polarized macrophages differentiated from patient a. monocytes or from thp cells treated with p gamma inhibitor or stably expressing pik cg shrna. conclusions: we report the first human with loss of pi kgamma activity and present her clinical presentation with notable t cell infiltration of barrier tissues. based on our analyses, we propose that loss of p gamma activity in humans causes t cell-intrinsic effects of reduced tregs and increased tissue-homing propensity and the t cell-extrinsic effect of augmented inflammatory responses in macrophages. together, these consequences of p gamma deficiency drive aberrant accumulation of t cells in lung and gut. introduction/background: pulmonary disease is a frequent complication across many primary immunodeficiencies (pidds), however its impact on the quality of life (qol) in pidds is not well characterized. objectives: to ascertain the types of infectious and non-infectious pulmonary complications occurring in pidds and to determine how these complications affect qol. methods: we analyzed the pulmonary complications, disability descriptions, and clinical status of subjects with pidds in the usidnet registry using descriptive statistics. karnofsky or lansky performance indices (n= ) and promis qol data (n= ) were also analyzed. the t-test/mann-whitney test and chi square test were utilized to compare continuous and categorical variables, respectively. results: infectious pulmonary disease was reported in a majority of subjects ( . %), most commonly pneumonia ( . %) and bronchitis ( . %). non-infectious pulmonary disease was reported in . % of all subjects, most commonly asthma/reactive airway disease ( . %), bronchiectasis ( . %) and interstitial lung disease ( . %). pulmonary insufficiency was listed as a cause of disability in . % of all subjects with pidds, with highest rates of this disability in subjects with immune dysregulation ( . %). lower karnofsky/lansky performance scores were observed in subjects with pneumonia, lung abscess, bronchiectasis, interstitial lung disease, and emphysema/copd as compared to without these disorders (p< . ). promis qol metrics were largely similar among subjects with and without pulmonary disease, although physical function scores were significantly worse in those with copd/emphysema (mean= . +/- . ) as compared to without (mean= . +/- . , p = . ). promis physical function scores were also worse in subjects with non-infectious pulmonary disease (mean = . +/- . ) compared to those with infectious pulmonary disease only (mean = . +/- . , p= . ). a significantly greater percentage of patients with a history of copd/emphysema ( . % vs. . %) or interstitial lung disease ( . % vs. . %) were deceased as compared to those without a history of these disorders (p< . ). conclusions: both infectious and non-infectious pulmonary disorders cause significant morbidity in pidds and are associated with higher mortality in this population. infectious and non-infectious pulmonary complications were often associated with worse karnofsky/lansky scores while there was limited impact on promis qol measures. latin-american consensus on the management of patients with severe combined immunodeficiency, part : supportive measures during the time from diagnosis to definitive treatment. juan carlos bustamante ogando , armando partida-gaytán , francisco espinosa rosales , lasid "consensus on scid" study group pediatric allergy and clinical immunology specialist, clinical immunologist and researcher at primary immunodeficiency research unit, national institute of pediatrics pediatric allergy and clinical immunology specialist, researcher at primary immunodeficiency research unit, national institute of pediatrics pediatric allergy and clinical immunology specialist, president, fundación mexicana para niñas y niños con inmunodeficiencias primarias (fumeni) result in a majority of patients with late diagnosis, more comorbidities and reduced access to curative treatments. the interventions during such period are vital to keeping optimum health status to improve the probability of success of curative therapies. many interventions are not supported by clinical trials, are based mainly on clinical experience, and there are no clinical guidelines to standardize such treatments. objectives: to generate a consensus on the supportive care of patients with scid, from the diagnosis until a curative treatment is given, under a latin-american perspective taking into account particular challenges for our region. methods: in a first step, we gathered available information about scid diagnostic and therapeutic guidelines from two sources: a) literature search and b) personal communications with pid experts from europe and usa. next, we developed an expert consensus through a modified delphi technique (electronic and anonymous). we used google® forms® to gather the information and microsoft office excel® for the analysis of agreement through kappa coefficient and rounds concordance through repeated measures analysis of variance (anova). results: we gathered an expert panel of subjects from latin-american countries (argentina, brazil, chile, costa rica, mexico, and peru) including the primary centers caring for scid patients. we generated a document with agreed diagnostic and therapeutic interventions grouped in topic-domains (i.e. protective and isolation methods to decrease the risk of infections, antimicrobial prophylaxis, immunoglobulin treatment, immunizations, nutritional aspects, antimicrobial treatment, blood derivatives use, routine laboratory workup, imaging and other studies, conventional multidisciplinary approach). we also included nonagreed interventions, but where relevant arguments are shared, to allow for particular clinical scenario decisions. conclusions: this is the first document of its type, and it intends to standardize clinical care of latin-american patients with scid, reduce disease burden and ultimately improve health outcomes. we see this effort as a starting point for the continuous improvement of our professional care to such patients and is intended to help as a tool not only for immunologists but for primary care physicians and other specialists involved in scid patient's care. this work will hopefully be published during as a lasid collaborative work, and it will help as a guide for clinicians caring for scid patients not only in latin america but in other world regions. also in the future, this consensus may be improved by collaboration from immunologists worldwide. no significant difference in hospitalizations one year before vs. year after treatment for prophylactic antibiotics (p= . ) or igrt (p= . ). baseline igg was higher in prophylactic antibiotics vs. igrt ( . vs. . mg/dl, p= . ) . sex, severity of sad, igg subclasses deficiency, and lymphocyte counts were not significantly different between treatment groups. conclusions: prophylactic antibiotics are not inferior to igrt in preventing infections in some sad patients. while, clearly some patients with sad will need igrt, our date indicate that larger prospective studies are needed to identify patients who will benefit most from igrt vs prophylactic antibiotics alone. richard and barbara schiffrin presidents distinguished professor of microbiology and director, institute for immunology, university of pennsylvania introduction/background: t cell thymic development is dependent on signals received via the pre-tcr complex and we here report the first case of pre-tcr alpha (ptcra) autosomal recessive t cell immunodeficiency in an infant with a positive scid newborn screen (nbs). objectives: we sought to uncover the mechanistic links between ptcra mutations and immune dysfunction. methods: the patient was tracked clinically, with serial clinical immunophenotyping and t cell function testing. in addition, we performed deep immunophenotyping with mass cytometry and single cell rna sequencing to delineate the molecular circuitry underlying her immune phenotype. results: the patient presented with t cell lymphopenia and impaired response to mitogen stimulation. hsct was considered, but she did not meet clinical criteria and remained healthy, so she was watched closely on prophylaxis while awaiting genetic testing. response to serial mitogen stimulation remained between~ - %, response to serial cd /cd activation was normal and tcrv spectratyping was normal. whole exome sequencing revealed two mutations in ptcra. no prior human cases of ptcra deficiency have been published, but a mouse model bears a striking resemblance to this case (fehling et al, nature, ) , with elevated t cells and decreased t cells. her mitogen stimulation responses became persistently normal around years of age with stable t cell lymphopenia, elevated t cells and normal switched memory b cells. anti-fungal prophylaxis was halted, and she remained on atovaquone alone with persistent t lymphopenia. she was able to mount an antibody response to rabies vaccine at . years and was weaned off scig replacement and is planned to initiate vaccination. deep immunoprofiling with mass cytometry (cytof) demonstrated a unique immunophenotype. single cell rna sequencing confirmed normal cd and cd tcr clonotypic diversity but increased clonotype diversity in t cells and increased and transcript levels. in addition, cd naïve, cd memory and cd naive t cells demonstrated both increased numbers of expressed genes and transcriptomic diversity, with altered cytoskeletal and tcr proximal signaling pathways across t cell subsets versus control. this may reflect a peripheral role for ptcra, a durable imprint of thymic signaling events mediated by ptcra, evidence of homeostatic proliferation or a combination of the above. conclusions: scid nbs led to identification of homozygous variants in ptcra causing a novel t cell immunodeficiency characterized by t cell lymphopenia, altered proximal tcr and cytoskeletal signaling and increased number of altered t cells. we will continue to pursue the mechanism of these mutations by developing ipsc and studying their t cell differentiation capacity in vitro, as well as further defining her immunometabolic phenotype. rag hypomorphic mouse mutants show partial preservation of thymocyte development but peculiar abnormalities of thymic epithelial cell phenotype introduction/background: the recombination-activating gene (rag) and rag proteins are essential for v(d)j recombination. in the absence of these proteins, the development of b and t cells is blocked at early progenitor stages, resulting in severe combined immunodeficiency (scid). hypomorphic mutations in rag , allowing residual activity, result in delayed-onset combined immunodeficiency with residual development of t and b lymphocytes, associated with autoimmunity and/or granulomas (cid-g/ai). objectives: to study in details the effect of rag hypomorphic mutations at the early stages of t cell development, we have generated mouse models carrying mutations described in patients with cid-g/ai (r q, r w, f l) (niaid animal protocol: lcim e). methods: we performed an extensive evaluation of the thymic phenotype in the mouse models. results: the number of total thymocytes was found to be drastically reduced in all three models. however, two of these mouse models (r q and f l) retained a significant level of rag activity, and resulted in the development of mature t cells in the thymus, while the mouse model carrying the r w mutation had minimal rag activity and presented a phenotype more similar to that of complete rag knockout mice. in r w mice, almost all thymocytes were blocked at the double negative (dn) stage and there were virtually no mature t cells, as found in rag ko mice. on the other hand, r q and f l mice presented double positive (dp) and single positive (sp) and sp cells. the cross talk between t cells and thymic epithelial cells (tec) in the thymus is fundamental for the development and maturation of both types of cells. in rag -/-mice, and consequently in the absence of mature t cells, tecs cannot complete their maturation, and the mtec subset is virtually absent. these results were also observed in the r w mouse model. instead, in r q and f l mice, the residual rag gene activity allows development of a reduced number of mature t cells. although the number of tecs was markedly reduced in r q and f l mice, ctecs and mtecs were both present, but with an excess of ctecs. furthermore, mtecs were predominantly mhc-iihigh (mtechi), and only a minority of mtecs were mhc-iilow (mteclo) cells, the opposite of what found in adult wt mice. finally, mtechi cells from rag mutant mice were found to express aire to levels and frequencies comparable to those of wild-type (wt) mice. conclusions: our results show that tec in mouse models carrying rag hypomorphic mutations are affected both in terms of absolute numbers and in terms of subset distribution and maturation state. to further investigate the functional consequences of impaired cross-talk between thymocytes and tecs in rag mutant mice, we have performed rnaseq in sp and tecs sorted from r q, f l and wt mice. analysis of the gene expression profile in tec may thus provide novel insights in the mechanisms that govern normal and pathologic thymic t cell development. vedolizumab for autoimmune enteropathy in primary immunodeficiency: a case series of outcomes introduction/background: gastrointestinal complications are common in patients with primary immunodeficiency. infections are the leading cause, but autoimmune enteropathies including inflammatory bowel disease (ibd)-like colitis, sprue-like enteropathy, and nodular lymphoid hyperplasia (nlh) have been recognized in a subset of these patients. to date, there is no established treatment for these noninfectious disorders. vedolizumab is a humanized monoclonal antibody that binds to the alpha- beta- integrin, inhibiting the migration of memory tlymphocytes across the endothelium into inflamed gastrointestinal parenchymal tissue. it is fda approved as first-line therapy for inflammatory bowel disease. the safety and efficacy of treating autoimmune enteropathy with vedolizumab in patients with concurrent primary immunodeficiency (pid) has not previously been reviewed. objectives: to review the outcomes of a series of patients with hypogammaglobulinemia and autoimmune enteropathy following vedolizumab therapy methods: patients ( male, female) at mount sinai with enteric biopsies demonstrating inflammatory enteropathy with t cell infiltrates have been treated with vedolizumab. results: five of the seven patients completed induction therapy. one patient was recently started on therapy. therapy was aborted in one patient who developed acute hepatitis during induction. another developed severe cytomegalovirus enteropathy, prompting discontinuation. two patients discontinued therapy due to response failure. at present, two patients remain on therapy at months with symptomatic improvement. conclusions: vedolizumab was effective in cases, but had no benefit or deleterious side effects in subjects. its effectiveness in another patient is presently under investigation. introduction/background: ataxia telangiectasia (at) is an immunodeficiency most often associated with t cell abnormalities and abnormalities in serum immunoglobulin levels, primarily iga. there is a subset of patients with a hyper-igm phenotype, some with cutaneous granulomas, which may reflect a distinct clinical phenotype. a yearold female presented for evaluation of concern for immunodeficiency because of frequent illnesses, presumed to be viral. she was found to have an ataxic gait, some speech delay, mild ocular and ear pinna telangiectasia, and an ulcerative rash on the left upper and right lower extremity. initial blood work showed elevated -fetoprotein levels ( ng/ml), elevated serum igm ( mg/dl), low igg (< mg/dl), and iga ( . mg/dl). objectives: to determine if the atm mutations in this patient are associated with perturbations in the frequencies, distributions and functions of b and t cell subsets which account for the observed phenotype. methods: next generation sequencing was used to identify the mutations in the atm gene. b and t cells were purified from the patients peripheral blood by positive selection. intracellular staining for foxp and t-bet was performed. b cells were activated in the presence of polyclonal f(ab) rabbit anti-human igm, multimeric, soluble, recombinant-human cd l, gardiquimod (tlr agonist), or cpg (tlr agonist). the treg suppression assay was carried out by co-culturing cd +cd hicd lo tregs and cd +cd cd + responder t cells at a : ratio in the presence of beads loaded with anti-cd , anti-cd , and anti-cd for . days. results: next generation sequencing revealed two pathogenic mutations in the atm gene, a novel mutation creating a premature stop codon [c. dela,(p.lys asnfs )], and a nonsense mutation [c. c>g, (p.tyr ter)]. proliferative responses of pbmc to mitogens (pha, cona, pwm) were reduced to roughly half of the control responses; the response to tetanus was normal whereas the response to c. albicans was absent. serum cytokine analyses demonstrated elevations in levels of tnf ( . pg/ml) and il- ( pg/ml); levels of ifn, il- , il- , il- , and il- were below the limits of detection. b cell abnormalities included markedly increased percentages of cd locd lo cells ( %) expressing t-bet and fas. activation of these cd /low b cells through the b cell receptor, tlr and tlr , and cd was decreased in response to all of the stimuli as evidenced by a lower percentage of b cells expressing the activation markers cd and cd relative to healthy control samples. the frequency of unswitched cd +igd+ memory b cells was also increased ( %). among the naive b cells, the proportion of cd + cd cd cd +igmhi transitional b cells that newly emigrated from the bone marrow (bm) was found to be diminished to . % of the naive b cell compartment. in the t cell compartment, there was a decreased frequency of total cd + cells but normal absolute numbers of cd +cd + t cells. there was also a decreased proportion of naive cd +cd +cd rocd l+ t cells and a striking increase in the cd +cd +cd ro+ memory t cells ( %). this appeared to be largely attributed to the increased proportion of cd +cd +cd ro+cd l effector memory t cells ( %). the circulating t follicular receptor (ctfh) cell frequency in the patient was -fold higher ( %) than the average for healthy donors but icos expression levels were normal. treg frequency was decreased but suppressive capacity was not impaired. conclusions: the mutations in atm described here add to the growing understanding of the heterogeneity in degree and complex nature of the immunodeficiency seen in patients with at. these mutations resulted in perturbations in frequencies and distributions of normal and atypical b and t cell subsets, which can explain some immunologic aspects of the clinical phenotype in this patient. the immunophenotype seen here may also differentiate at patients with granulomas from those without cutaneous lesions. supported in part by grifols, the joanne siegel memorial fund, the dreizessen fund (to ewg), grants from niams t ar - (km) and niaid r ai (em). introduction/background: a -month-old male presented with pancytopenia, b cell deficiency and developmental delay. he was born at weeks with weight of . kg. he was severely anemic with a hb of . , and transfused on day of life. he received hep b and bcg vaccines without complications. a month later he had a hb of . with a febrile illness. a bone marrow aspiration performed at days, showed dyserythropoiesis without hemophagocytosis, and normal numbers of precursors. t and b cells were decreased. further evaluation with repeat bone marrow showed decrease in all cell lineages. exome sequencing of the family showed homozygous variant in mysm (c. _ delp. (lys arg/s* ) omim: * ) in the patient. both parents and hla-matched sister, were heterozygous for the same variant in mysm . treatment consistent of replacement immunoglobulin, packed rbcs, and g-csf. there was no history of recurrent viral or severe bacterial infections except for - episodes of urinary tract infections, which were treated with antibiotics. physical exam revealed low set ears, sunken and wide set eyes, depressed nasal bridge, mild micrognathia, frontal bossing, and cm x cm cafeau-lait spot noted behind left knee. he was pancytopenic with a wbc count ranging /mcl to /mcl, and anc ranging from /mcl to /mcl (on intermittent g-csf). hb = . gm/dl requiring transfusions every - weeks, and platelet count was , /mcl. b cell deficiency was confirmed with total b cell count of cells/mcl. b-cell maturation was essentially normal. t cell counts were normal with ageappropriate distribution of naïve and memory t cells, and t cell function. there was normal t cell receptor repertoire diversity. objectives: to assess defect in dna repair using a flow cytometry-based assay in a patient with mysm deficiency. methods: patients with mysm deficiency are reported to have increased genomic instability. deb testing, and telomere length analysis revealed normal results. defects in the dna repair pathway were assessed using a flow cytometry-based assay measuring phosphorylation of atm (patm), smc (psmc ) and h ax (gh ax) without irradiation, or h or h after low-dose ( gy) radiation using a cs source. the analysis was performed in t, b and nk cells. results: the patient had higher patm and psmc in t cells compared to the experimental controls (hc) at h post-irradiation. also, the amount of gh ax in nk cells was significantly higher than hc at h post-irradiation. interestingly, the patients b cells showed approximately % of b cells with constitutive gh ax even without irradiation, and this subset increased slightly to % at h after irradiation. the mfi (amount) of gh ax also increased at this time-point. at h post-irradiation, there was normal dephosphorylation in healthy control lymphocyte subsets. however, the patients t cells did not de-phosphorylate completely and showed higher residual patm, and psmc in both t and b cells. also, both t and b cells, at h, demonstrated a small subset of t cells ( %) with constitutive gh ax without irradiation, which increased to % after irradiation. there was also an increase in gh ax mfi in the irradiated sample. in b cells, % showed constitutive gh ax without irradiation at h, and this increased to % after irradiation, with a corresponding increase in mfi. conclusions: in summary, this rapid flow analysis revealed defects in the dna repair pathway, including higher patm, psmc and h ax phosphorylation in t, b and nk cells at h post-irradiation. at h, only t cells showed a residual subset with patm expression. but, psmc , a downstream target of atm, revealed higher levels in t, b and nk cells at h post-irradiation. this assay, which allows lineage-specific analysis, permitted dissection of dna repair defects, in individual lymphocyte subsets revealing heterogeneity within the cell subset to radiation susceptibility. the practical benefit of this rapid multi-parameter flow assay is selection of appropriate conditioning regimen for hematopoietic transplantation, as was the case with this patient. this has significant practical implications for treatment of patients with radiosensitive immunodeficiencies. ctla- haploinsufficiency-associated inflammation can occur independently of t-cell hyperproliferation introduction/background: cd and ctla provide opposing proliferative signals to t cells. we identified an -year-old female subject (s ) with heterozygous deletions of cd and ctla and multi-organ inflammatory disease characterized by a lack of t cell infiltrates in affected organs. inflammatory disease was remarkably responsive to s ctla -ig therapy. objectives: our goal was to characterize the immunologic consequences of combined deletion of cd and ctla , specifically assessing t cell proliferation and treg function in comparison to patients with alps associated ctla haploinsufficiency. we further sought to explain how this s s inflammatory diseases could occur without a pathologic t cell infiltrate and why they were amenable to ctla -ig therapy. methods: we performed phenotypic analyses of subject t cells and innate lymphoid cells (ilcs). we functionally characterized subject t cells. we created serum cytokine profiles. we stained and analyzed tissue biopsies. results: cd and ctla expression on s t cells were half that of control t cells. s t cells were hypoproliferative. s tregs were scarce and lacked suppressive function similarly to alps tregs. s tregs could suppress autologous t responder cells, likely due to their poor proliferative capacity. s colonic biopsies featured significantly fewer infiltrating intraepithelial lymphoid cells than biopsies from an alps patients. unlike alps patients whose colonic gland infiltrates were overwhelmingly t cells, s intraepithelial lymphoid cells were neither t cells nor b cells, suggesting the presence of ilcs. indeed, a greatly expanded population of type innate lymphoid cells (ilc ) and prototypical ilc cytokines were identified in s peripheral blood. ilc frequency and cytokine levels decreased in response to treatment with ctla -ig, corresponding with marked improvement in enterocolitis, hepatitis and pericarditis. conclusions: we report a novel genetic syndrome of combined cd /ctla deletion and describe the immunolopathologic correlates of this disease. dual ctla -and cd -haploinsufficiency results in a phenotype of multi-organ inflammatory disease characterized by ilc expansion in the setting of t-cell hypoproliferation and quantitative and qualitative treg defects. our patients clinical response to ctla -ig parallels published mouse studies and suggests the existence of additional stimulatory b receptor(s) preferentially expressed on ilc s over conventional t-cell populations. diagnosis of radiosensitivity and dna repair defect in dna ligase iv deficiency with a rapid flow cytometry assay. introduction/background: dna ligase deficiency (lig -scid) is one of several monogenic defects affecting dna repair, and causing lymphopenia (t-b-nk+) and a radiosensitive scid (rs-scid) phenotype. the assignment of a timely diagnosis is vital in the management of patients with rs-scid. laboratory assessment of radiosensitivity is laborious, and utilizes fibroblasts (non-hematopoietic) or lymphoblastoid cell lines, and can take several weeks to months for results. objectives: we demonstrate for the first time, the application of a flow cytometric-based kinetic analysis of phosphorylated h ax (h ax) in lymphocyte subsets, especially nk cells, for the diagnostic assessment of lig -scid. methods: simultaneous measurement of multiple dna repair markers phosphorylated (p) atm, smc and h ax (h ax) was performed by flow cytometry to assess dna repair defects in a -year-old korean female. the patient was evaluated for recurrent fevers, chronic respiratory tract infections, chronic diarrhea, and rash. genetic testing revealed compound heterozygous variants (nm_ , c. g>t, p.trp cys and nm_ , c. a>t, p.asp val) in lig . functional assessment (phosphorylation) was measured in t and nk cells (b cells were absent), before irradiation (background control), or after low-dose ( gy) irradiation ( and hours). results: we observed maximal h ax generation at hour post-irradiation, with progressive dephosphorylation at hours post-irradiation in healthy controls. the patient showed normal frequencies (%) of t cells and nk cells positive for h ax ( . % and . % respectively); (controls (n= ) t cells = . % and %; nk cells = . % and . %), but increased intracellular levels (mean fluorescence intensity, mfi) of h ax (t cells = . and nk cells = . ) compared to controls (t cells = . and . ; nk cells = . and . ) at hour post-irradiation. however, more importantly, at hours post irradiation there was a lack of dephosphorylation in a substantial proportion of lymphocytes ( % of t cells and % of nk cells) compared to healthy controls (t cells= . % and . %; nk cells = . % and . %). further, while there was dephosphorylation of h ax at h in patient lymphocytes as compared to h, the amount, as measured by mfi, remained elevated at h (t cells = . , nk cells = . ) compared to controls (t cells = . and . ; nk cells = . and . ). the data from patm and psmc were uninformative for the evaluation of lig -scid. conclusions: flow-based kinetic analysis of h ax is a useful marker for the diagnosis of lig -scid, and can be performed with a small amount ( cc) of blood, and provides a result in - days, facilitating rapid assessment of radiosensitivity in this condition. human plcg haploinsufficiency results in nk cell immunodeficiency and herpesvirus susceptibility objectives: we aimed to investigate the cause of disease in three patients from two kindreds with recurrent or severe herpesvirus infections and nk cell dysfunction. methods: we used exome sequencing and mass cytometry (cytof), as well as traditional immunologic techniques, to investigate the genetic causes, immune cell subpopulations/signaling, and nk cell function of these patients. we additionally used mouse models, crispr cell lines and in vitro assays to assess the role of plcg haploinsufficiency in disease. results: kindred a consisted of two patients presenting with hsv susceptibility and autoimmunity. kindred b consisted of one patient with severe cmv myocarditis and adenoviral hepatitis. both kindreds were evaluated for nk cell function and showed reductions in target killing in spite of normal cytotoxic granule degranulation against the same target. microscopy analysis suggested that granule mobility was reduced in at least one kindred. cytof revealed reductions in plcg phosphorylation after receptor crosslinking in the nk cells of both kindreds. kindred a also presented with a reduction in naïve b cells without perturbations in immunoglobulin output, b cell memory formation or class switching. trio whole exome sequencing was performed and revealed rare heterozygous plcg mutations in both kindreds. functional analysis, as well as mouse and crispr models, support a functional haploinsufficiency as a cause for nkd in these patients. conclusions: heterozygous loss-of-function point mutations in plcg have not been previously investigated as a cause of nk cell deficiency or recurrent herpesvirus infection. thus, these patients represent a novel immunodeficiency involving plcg haploinsufficiency, nk cell dysfunction, and herpesvirus susceptibility. hypomorphic rag mutations alter the pre-immune repertoire at early stages of lymphoid development introduction/background: human rag deficiency is associated with a spectrum of clinical phenotypes. while the most severe forms of rag deficiency manifest with severe combined immune deficiency or omenn syndrome since the first weeks of life, more recently patients have been identified who present to medical attention at a much older age predominantly with symptoms of autoimmunity and/or inflammation. many of the mutations associated with this atypical syndrome are found in the c-terminal domain (ctd) of the rag gene and allow residual development of t and b cells. these patients have an abnormal peripheral t and b cell repertoire, but how this is affected by abnormalities in the composition of the pre-immune repertoire vs. antigen-mediated selection and homeostatic proliferation in the periphery is unknown. objectives: in order to investigate whether mouse models with hypomorphic mutations in the rag ctd recapitulate the phenotype observed in patients with cid-g/ai, and to study how these mutations affect repertoire composition, cell selection and survival during t and b cell development, we generated three mouse models carrying homozygous rag mutations (f l, r q, and r w), corresponding to human mutations (f l, r q, r w) previously reported in patients with late-onset combined immune deficiency with granuloma and/ or autoimmunity (cid-g/ai). methods: mice were generated using crispr/cas mediated gene editing. t and b cell development, including apoptosis was studied by flow cytometry. immunoglobulins in naïve mice, baff levels and specific antibody responses were measured by elisa. serum igm autoantibodies were measured using a microarray (utsw). analysis of t cell receptor (trb) repertoire in several sorted t cell populations and immunoglobulin heavy chain (igh) repertoire in pre-b cells was performed by adaptive biotechnologies. in order to be able to detect both dj and vdj rearrangements, pro-b cells and spleen b cells were sequenced using high-throughput genome-wide translocation sequencing-adapted repertoire sequencing (htgts-rep-seq). analysis of vk-jk rearrangements in pre-b cells was performed by pcr amplification. results: immunological characterization showed partial development of t and b lymphocytes, with persistence of naïve cells, preserved serum immunoglobulin, but impaired antibody responses and presence of autoantibodies, thereby recapitulating the phenotype seen in patients with cid-g/ai. by using high throughput sequencing, we identified marked skewing of igh v and trb v gene usage in early progenitors, with a bias for productive rearrangements after selection occurred, and increased apoptosis of b cell progenitors. this suggested that more alleles remained in germline configuration. moreover, in the rearranged igh loci, the distal v gene segments were preferentially rearranged already at the earliest stages of b cell development, a finding that has not been previously reported. in addition, rearrangement at the igh locus was impaired, and polyreactive igm antibodies were detected. conclusions: in conclusion, this study demonstrates that hypomorphic rag mutations reported in cid-g/ai cause abnormalities of the primary b and t cell repertoire. these changes may affect survival and selection of t and b cells, and thereby contribute to the immune dysregulation often seen in patients with cid-g/ai. senior clinician, nih/niaid/lcim introduction/background: autosomal dominant hyper ige syndrome (ad-hies) is a primary immunodeficiency due to loss of function stat mutations. disease manifestations include recurrent skin and pulmonary infections, eczema, mucocutaneous candidiasis, as well as tion and in vitro studies demonstrated that the enhanced signaling could be controlled by ruxolitinib, an approved jak / inhibitor. informed by these experimental data, the patients were treated with ruxolitinib with remarkable improvement in a variety of clinical end-points, including hematological profiles and growth parameters. conclusions: this characterization of a human jak gain-of-function mutation expands our current understanding of the role of jak in eosinophil biology, hematopoiesis and immune function. chronic granulomatous disease, ornithine transcarbamylase deficiency and x-inactivation is a primary immune deficiency characterized by defects in the nadph oxidase enzyme complex resulting in a susceptibility to a narrow spectrum of bacteria and fungi. mutations in cybb encoding gp phox and located at xp . are associated with the most common form of cgd. deletions and rearrangements in this region are associated with other genetic diseases such as mcleod syndrome (xk), retinitis pigmentosa (rpgr), duchenes muscular dystrophy (dmd), ornithine transcarbamylase deficiency (otc), and x-linked mental retardation (tspan ). patient phenotype depends on the extent and position of the deletion, creating a "contiguous x-chromosome gene deletion syndrome. objectives: we report a four-year-old female, who presented with symptoms of x-linked cgd and otc deficiency with a large, contiguous multi-gene deletion on the x-chromosome. methods: we report a four-year-old female, who presented with symptoms of x-linked cgd and otc deficiency with a large, contiguous multi-gene deletion on the x-chromosome. the patient is the second born of a set of non-identical triplets from a clomiphene assisted pregnancy at weeks gestation. (weight at birth: lbs oz). after a day stay in the nicu, and cpap for one day she was discharged home. by the second month, she began having problems gaining weight and had persistent vomiting. at months, she was admitted with a pneumonia diagnosed as methicillin resistant staphylococcus aureus by lung biopsy. during that hospitalization, the mother noted an enlarging lesion on the infants left hand which was biopsy proven serratia marscecens osteomyelitis and was treated with intravenous cefepime for weeks. at this point a dihydrorhodamine assay (dhr) showed only . % positive cells (nl - %) and she was diagnosed as a carrier of x-linked cgd. bactrim was initiated for antibacterial prophylaxis but it was discontinued due to recurrent diarrhea; she was unable to tolerate antifungal prophylaxis as well due to liver function abnormalities. she continued having frequent vomiting episodes, irritability, failure to thrive and development delay. ulcerations in esophagus were confirmed by egd and colonoscopy, probably related to persistent emesis. diarrhea was unresolved. at months of life she was admitted with acute encephalitis, a serum ammonia level of and elevated urine orotic acid. results: given the demonstrated carrier status for cybb and apparent otc deficiency, comparative genomic hybridization was performed, revealing a deletion from xp . to xp . this . mb loss includes genes that are known to cause disease. since the diagnosis, the patient has been on reduced protein diet, resolving her diarrhea, she has subsequently grown and is on th percentile for weight and height. her dhr is now % positive. at years she was diagnosed with cone rod dystrophy. currently she is on bactrim for cgd prophylaxis and l-citrulline for the otc deficiency. she continues to gain weight, and has shown great improvement in her development delay and no new cgd-related infections have recurred. conclusions: this case reminds us that x-linked carriers with large deletions may be symptomatic and genetic analysis to determine other affected genes can be important for medical management. introduction/background: wiskott-aldrich syndrome (was) is a rare and severe x-linked disorder with variable clinical phenotypes correlating with the type of mutations in the was gene. the long-term prognosis of this syndrome is generally poor, with hematopoietic stem cell transplantation (hsct) remaining the only curative choice. the syndrome is poorly characterized in china. objectives: we retrospectively reviewed patients with was referred to our hospital from to , and summarize their clinical manifestations and genetic features. methods: sixty-four children suspected to be was from unrelated families were enrolled in this study. the clinical data of children were reviewed in the present study. distribution of lymphocyte subsets from peripheral blood and was protein (wasp) expression in peripheral blood mononuclear cells was examined by ow cytometry (fcm). wasp mutations were identified by direct sequencing of pcramplified genomic dna results: among patients with primary immunodeficiency diseases (pid), ( . %) were finally diagnosed as was with gene identified. the mean time of diagnosis was . months (range, . - . ). the common onset clinical manifestation was diarrhea, and most patients had recurrent upper respiratory tract infection, otitis media, pneumonia, and skin abscess. one patient had nephrotic syndrome and no patient with malignancy. all patients had classical was phenotype with was clinical scores - . total mutations in wasp were identified, including novel mutations. six patients received hsct, five survived, with one died because of gvhd. compared with the other patients without wasp mutations, was patients had lower numbers of cd + t cells and b cells, and higher eos and ige level. there was a negative association between the number of b cells and the was clinical scores. conclusions: in china, diagnosis of was has improved over the last decade, although a much higher number of cases had been expected. establishing more diagnostic centers dedicated to the care of pid will facilitate early, correct diagnosis and better care of was in china. regulatory cells (tregs) are precisely quantified by measuring demethylation of the treg-specific-region of foxp . more recently, we have identified highly cell type-specific dna regions of demethylation for further cell populations. objectives: this novel technology allows the implementation of differential immune phenotyping into the newborn screening procedure. here, we aimed at epigenetically quantify multiple immune cell types in different biological samples including dried blood spots and samples from patients with pid with immundysregulation, where currently no approach is available. methods: using cell type-specific demethylation sites, we developed epigenetic qpcrs for quantification of t-, treg, b-, nk-, monocyte and granulocyte cell population. epigenetic qpcr is applicable for relative and absolute quantification in whole blood and dried blood spots using isolated, bisulfite converted dna. results: we demonstrated > % concordance with flow cytometric analyses of the same fresh blood samples from healthy subjects. we have validated the method in children with symptoms of immundysregulation resulting from monogenic defects, including for example foxp , cd , stat , ctla , and leading to treg/teffector cell imbalance where treg deficiency has been difficult to assess by flow cytometry. furthermore, we tested dried blood spot (guthrie card) samples from healthy newborns and patients with diverse pids and correctly identified / pid patients, indicating that this method holds promise for newborn screening. conclusions: the method we established for immune cell quantification based on epigenetic cell type-specific markers, is feasible and reliable in biological samples either fresh, frozen or archived, including dried blood spot. the analysis of further immune cell types introduces an innovative opportunity to diagnose a variety of pids and immunodysregulatory disorders as early as newborn screening. pancytopenia and immunodeficiency with mds in an infant due to samd l mutation we present our data on behalf of the pcid study consortium of the inborn errors working party worth profound combined immunodeficiencies (p-cid) are inherited diseases with impaired t-cell function leading to infections, immune dysregulation or malignancies. genetic, immunologic and clinical heterogeneity make patient specific decisions on indication and timing of hematopoietic stem cell transplantation (hsct) difficult. objectives: since the pcid study recruits non-transplanted p-cid patients aged - years to prospectively compare natural histories of age and severity-matched patients with or without subsequent transplantation and to determine whether immunological and/or clinical parameters may be predictive for outcome methods: our prospective/retrospective international observational multicenter study recruits pediatric p-cid patients to identify biomarkers and clinical parameters that are predictive of outcome. results: so far > growth hormone deficiency comprised the majority of neuroendocrine cases ( %, / ). other notable neurologic diagnoses included headache ( %, / ), seizure ( %, / ), cerebrovascular accident ( %, / ), and neurologic tumors ( . %, / ). in addition to somatic neurologic conditions, many cvid patients ( . %, / ) had reported diagnoses of depression, anxiety, and post-traumatic stress disorder. conclusions: our findings suggest that neurologic diagnoses are more common in cvid patients than previously recognized. patients with neurologic autoimmune disease appear to have a more severe phenotype with earlier age at symptom onset. many cvid patients had depression, anxiety clinical outcomes of human herpesvirus reactivation after hematopoietic stem cell transplantation prognostic factors and outcome of epsteinbarr virus dnaemia in high-risk recipients of allogeneic stem cell transplantation treated with preemptive rituximab cytomegalovirus in hematopoietic stem cell transplant recipients daratumumab controls life-threatening post-hsct autoimmune haemolytic anaemia objectives: we present a month old pancytopenic male who was diagnosed with myelodysplastic syndrome (mds) with monosomy due to samd l mutation. methods: whole exome sequencing was performed on a male, who presented at months of age with findings concerning for a bone marrow failure (bmf) syndrome despite a normal bmf genetic panel. results: the patient presented at months of age, with severe pancytopenia, fevers, e. coli bacteremia, pancolitis, and echtyma gangrenosum. bone marrow showed severe aplasia with occasional macrophages.he was treated with antibiotics, as well as steroids, etoposide and cyclosporine for presumed hemophagocytic lymphohistiocytosis (hlh). a bone marrow failure and hlh genetic panels were normal conclusions: this is one of the first reported cases of samd l mutations causing mds since its initial discovery earlier this year. samd l mutation should be considered in patients who present with pancytopenia and monosomy mds. as new defects continue to be identified, further evaluation outside of typical bmf panels may be relevant primary immune deficiency disease in patients over age : an analysis from a proprietary immunology patient registry roger ucla school of medicine director objectives: to characterize the prevalence of pidd among older individuals using a patient database maintained by the consortium of independent immunology clinics (ciic), comprised of specialty immunology outpatient practices in the us. methods: patients with pidd were identified in the ciic database using icd- codes d conclusions: our data suggest that pidd in patients over age may be more prevalent than previously reported introduction/background: the patient is a -month-old boy, born at + weeks gestation to non-consanguineous parents of italian origin. he was admitted to the intensive care unit at days of age with profuse bloody diarrhoea, weight loss, severe metabolic acidosis and acute renal failure. he had a rapid respiratory deterioration necessitating intubation, ventilation and inotropic support. the patient developed features of macrophage activation syndrome with: (i) prolonged fever > . °c, (ii) hepatosplenomegaly, (iii) bicytopenia (anaemia and thrombocytopaenia), (iv) hypertriglyceridemia, (v) high ferritin ( , ) and (vi) haemophagocytosis on bone marrow smear. he also presented with three interesting features (i) a macular erythematous rash that slowly resolved and was replaced by reticulo-livedoid rash, (ii) a marked hypereosinophilia and (iii) no significant elevation of hladr/cd + t cells on lymphocyte immunophenotyping ( - %). the patient underwent rectal biopsy which confirmed the presence of eosinophils, but without significant inflammation or architectural changes. stool microscopy showed presence of partially necrotic intestinal epithelial cells. immune work up demonstrated global t lymphopaenia without balanced subpopulation and eliminated a familial hemophagocytic lymphohistiocytosis (normal perforin and cd a expression on cd + t-cell and nk cells). circulating foxp + cd +cd lowcd + t cells were within normal range. a dihydrorhodamine reduction assay was normal. chief, laboratory of clinical immunology and microbiology, national institute of allergy and infectious diseases, national institutes of health introduction/background: hematopoietic stem cell transplantation (hsct) has been used for the treatment of hematologic malignancies and primary immunodeficiencies (pid), for several decades with increasing efficacy. however, toxicity related to conditioning regimens based on the use of chemotherapy and/or irradiation to ensure engraftment of donor cells, remains a significant problem. recently, an alternative, potentially low-toxicity, approach has been proposed, which makes use of an immunotoxin targeting cd -expressing cells, which include hsc and more mature leukocytes. this approach is particularly attractive for leaky forms of severe combined immune deficiency (scid), with residual production of dysfunctional t and/or b cells, such as atypical forms of rag deficiency. objectives: we have developed a mouse model carrying a hypomorphic mutation in the rag gene (p.f l) resulting in a combined immunodeficiency with signs of autoimmunity, recapitulating the phenotype seen in patients. methods: using this model, we have tested the efficacy of conditioning with an anti-cd immunotoxin (cd -sap) alone or in combination with low irradiation ( rads; cd -sap/ ), and compared these regimens to a myeloablative dose of irradiation ( rads) [niaid protocol lcim e]. following conditioning, the mice were transplanted with wild-type (wt) bone marrow (bm) lineage-negative cells and followed over time to evaluate immune reconstitution. results: conditioning with cd -sap alone or with cd -sap/ led to a consistent engraftment of donor t and b cells in the peripheral blood (pb) of f l that increased overtime, reaching % donor chimerism at weeks. myeloid (cd b+) and nk cells in pb of f l mice also showed high level of donor engraftment that remained stable at around % in cd -sap around hours of life and the second after one week. there is no data on the utility of one versus two screens for scid screening. here we present our data evaluating whether patients with scid or t-cell lymphopenia were identified on the first or second trec screen. objectives . determine the benefit of a second trec screen in identifying scid and t-cell lymphopenia at birth. . examine outcomes of trec screening in washington state since implementation. methods: results of scid newborn screening performed in washington state between january and june were reviewed retrospectively with the staff of the washington department of health laboratory where the trec assay is performed. trec thresholds (copies/μl) were defined as follows: absent ( ), low ( - ), borderline ( - ) and normal (> ). all trec assays were run with a beta-actin control to assure sample adequacy. a screen is considered abnormal if there is one low/ absent trec or two borderline trec. newborns with abnormal trec screening have follow-up diagnostic testing consisting of lymphocyte flow cytometry to evaluate numbers of naïve and mature t cells, b cells, and nk cells, performed at seattle childrens hospital. results: a total of positive trec screens were found in washington state between january and june . five patients who did not have diagnostic flow cytometry testing were excluded from the analysis (one protocol deviation, one lost to follow-up and three who died before testing could be performed). the first screen was abnormal in forty three patients, while the second screen was abnormal in patients. five patients had an abnormal third or fourth trec drawn for other newborn screen follow-up. three patients with scid were identified, all with abnormal values on first screen. fortyfive patients with t-cell lymphopenia were identified; from the first screen and from the second screen. there was one patient with mhc ii deficiency was missed by both first and second screens because she did not have t-cell lymphopenia. the false positive rate with the first screen was % versus % with subsequent screens. the false positive rate dropped to % with two abnormal trec. the positive predictive value of scid or t-cell lymphopenia with the first abnormal trec was % versus % with two abnormal trec. average age of collection among infants with a positive screen was . hours for the st nbs and . days for the nd nbs. live viral vaccines were postponed in three patients who had an abnormal secondary screen (one with idiopathic t-cell lymphopenia, one with ectrodactyly-ectodermal dysplasia-clefting (eec) syndrome and one with q . deletion). one of these was started on pjp prophylaxis. conclusions: the practice of obtaining a second nbs from all newborns in washington state has led to increased identification of patients with tcell lymphopenia but did not result in identification of additional patients with scid. the false positive rate of the first and subsequent newborn screens was similar and decreased in patients with two abnormal trec. interventions including delaying live viral vaccines and pjp prophylaxis were instituted in patients who had a normal initial trec but abnormal secondary screen and documented t cell lymphopenia. two trec screens in all newborns can result in identification of additional patients with t-cell lymphopenia who may require intervention and additional follow-up. it is not yet clear whether the cost of a second mandatory scid newborn screen is balanced by the additional sensitivity gained by this approach. introduction/background: mannose-binding lectin (mbl) is a multimeric lectin that recognizes a wide array of pathogens independently of specific antibody, initiates the lectin pathway of the complement system, and acts as a proinflammatory mediator. mbl deficiency is reported to increase the frequency of infections in patients with impaired immune systems or cystic fibrosis (cf) patients. mbl replacement is experimental and unavailable. immunoglobulin replacement therapy (igrt) is controversial in the treatment of mbl deficiency; however, there are no reports of its efficacy or role in this condition. we describe a cf carrier patient with mbl deficiency, ciliary dyskinesia and mildly low igg who did not respond to igrt. objectives . understand when mbl deficiency can be symptomatic. . define the relationship between mbl deficiency and cf. . describe the treatment options of mbl deficiency. . define the efficacy of igrt and mbl deficiency. methods: a single case report. results: year old girl with mbl deficiency, cf carrier state with polymorphisms ( - a>g cf variant with t/ t and m v polymorphism) and ciliary dyskinesia (diagnosed by ciliary biopsy) who initially presented at eight years of age with recurrent sinusitis, recurrent otitis media status post tympanostomy tube placement, reactive airway disease and tracheomalacia. she had nine episodes of recurrent sinusitis with six negative sinus cultures and one positive for pseudomonas aeruginosa. she required a total of nine courses of antibiotics. labs showed several mbl levels < ng/ml on three occasions, normal ch , ah , igg - mg/dl (low normal for age), normal iga, igm, t cells, b cells, nk cells, and robust specific antibody titers. despite adequate pulmonary hygiene including nebulized levalbuterol, budesonide, dornase alfa, ipratropium, hypertonic saline and compression vest twice daily, she continued to have recurrent bronchitis and cough. in addition, even with sinus rinses and intranasal corticosteroid, she continued to have - episodes of sinusitis yearly. for this reason, she underwent bilateral total ethmoidectomies and maxillary antrostomies with modest reduction in frequency of sinus infections and symptoms. however after two years, her bacterial sinusitis recurred. four episodes were positive for methicillin staphylococcus aureus or pseudomonas aeruginosa. this did not improve significantly with a trial of intranasal mupirocin. due to increased frequency of bacterial sinusitis refractory to traditional therapy, she was started on subcutaneous igrt dosed approximately mg/kg/month dosing maintaining igg troughs around mg/dl. after a six month trial, she did not have improvement in the frequency of sinusitis, bronchitis and otitis media. she required - courses of antibiotics for bacterial upper respiratory tract infections and igrt was stopped. prophylactic antibiotics and repeat sinus surgery were instituted. conclusions: majority of the patients with low/deficient mbl levels do not manifest significant symptomology due to the redundancy of the innate immunity. the increased susceptibility to infections is thought to be due to additional factors that compromise other components of the immune system. specifically in cf patients, mbl deficiency is associated with earlier colonization with pseudomonas, more rapid decline in lung function and earlier death secondary to end-stage lung disease. there are no reports of combined mbl and cf carrier symptomatic patients similar to this patient. there are no validated age-corrected values for mbl levels in pediatric patients. the clinical relevance of these levels to infection frequency, severity, or treatment of mbl deficiency remains to be proven. it has been proposed that < ng/ml is considered deficient in children. mbl therapy is still experimental and not commercially available. management when provided for severe or frequent infections includes prompt treatment with antibiotics, prophylactic antibiotics, appropriate vaccinations, and a trial of igrt. there are no reported cases describing the efficacy of igrt in mbl deficiency, and the mechanism subsequent genetic analysis identified the presence of a de-novo heterozygous mutation in the nucleotide binding domain of nlrc (c. g>c, p.val leu). a mutation involving this amino-acid position has already been described but with a different substitution pattern in a boy and his father who presented with mas (p.val ala) ( ) . in order to prove the causality of this mutation, our team generated thp- cell lines expressing the two different mutations through gene-editing with the crispr system. in this system the mutation p.val leu, as well as the p.val ala mutation, were responsible for spontaneous activation of caspase , as evidenced by flica assay. the patient was treated with iv methylprednisone mg/kg/day. he continued to have progression of his inflammatory state, and was therefore commenced on anakinra. following confirmation of mutation, he was started on rapamycin, reasoning that (i) through autophagy induction, rapamycin could potentiate the action of anakinra ( , ) and (ii) through mtor inhibition counteract the effect of il- on t-cells ( ). with a combinatory therapy of anakinra up to mg/kg/day and ramapycin (with trough levels of - ng/l), the patient showed a marked clinical improvement, allowing weaning of steroids and establishment of enteral feeds. ferritin levels reduced to - ng/ml. we observed a significant decrease in il- plasmatic level following treatment initiation (pre-vs post-treatment levels of pg/ml and pg/ml, respectively). we report a novel nlrc gain-of-function mutation, presenting with neonatal enterocolitis and autoinflammation with improvement under combinatory therapy of anakinra and rapamycin. to our knowledge this is the first case to report the use of rapamycin in this disease, with what appears to be encouraging results. further studies are required to elucidate the potential role of rapamycin in the management other inflammasome disorders. introduction/background: allogeneic hematopoietic stem cell transplantation (hct) is currently standard treatment for patients with severe combined immunodeficiency (scid), with -year overall survival > % for typical scid (heimall blood ). previous studies revealed that poor clinical outcomes correlated with poor long-term t cell reconstitution, including low cd t cell counts and low naïve cd + t cell counts (pai nejm ) . we hypothesized that t cells developing in a poorly reconstituted immunologic environment would show features of chronically activated t cells with increased expression of inhibitory co-receptors. we further hypothesized that the intensity of the conditioning regimen would correlate with the expression of inhibitory co-receptors. objectives: to characterize the t cell phenotype of scid patients at > years post-hct and to investigate the impact of conditioning regimen on the quality of t cell reconstitution and the expression of inhibitory coreceptors methods: we analyzed scid patients - yrs (median yrs) after hct. we excluded from the analysis patients with chronic graft-versushost disease (gvhd), chronic dna viral infections or patients who had received donor lymphocyte infusion or boost in the months prior to study. scid genotypes (n) included il rg/jak ( ), rag /rag / dclre c ( ), ada ( ), il r ( ) and other/unidentified ( ). nine patients had received a reduced intensity (ric) or myeloablative (mac) conditioning regimen, while had received either no conditioning or immunosuppression only (none/is). poor t cell reconstitution was defined as cd t cell counts below cells/mm ( patients). t cell phenotype, including expression of inhibitory co-receptors, was assessed by flow cytometry. results: compared to patients with cd counts above cells/mm , patients with low cd counts had low naïve cd ra+ccr + t cells (p= . ) and high cd ra-ccr -t effector memory (tem) cells (p= . ), low numbers of naïve thymic cd ra+ cd + t cells, low numbers of trecs and a less diverse t cell repertoire (p< . ). additionally, they had an increased frequency of cd t cells expressing pd ( % vs %), ctla ( % vs . %), cd ( % vs %, p= . ), and b ( % vs %, p= . ) inhibitory co-receptors. increased inhibitory receptor expression was associated with a differentiation profile skewed toward a tem phenotype, reduced t cell diversity, increased markers of t cell activation, and the development of a highly exhausted cd high pd- high t cell population. more importantly, a fraction of ccr + cd ra+ cd t cells expressed pd ( % vs %, p= . ) and b ( % vs %, p= . ) in patients with low cd counts, suggesting that some naïve t cells of poorly reconstituted patients were chronically activated. inhibitory receptor expression did not increase with hla disparities between the donor and recipient, a history of gvhd after transplant or the infection status of the patient prior to transplant. however, inhibitory co-receptor expression was correlated with conditioning regimen, with increased frequency of b + cd t cells in unconditioned patients ( % vs % in none/is and ric/mac patients respectively, p= . ). conversely, ric/mac conditioning was associated with higher naïve cd t cell numbers (p= . ), higher naïve thymic cd ra+ cd + t cell numbers (p= . ), a more diverse t cell repertoire (p= . ) and low expression of inhibitory co-receptors. ric/mac conditioning was also associated with improved naïve t cell generation and limited expression of inhibitory receptors in il rg/jak patients ( % vs % for b , % vs % for cd in ric/mac versus none/is il rg/ jak patients respectively), a genotype permissive to t cell engraftment. conclusions: collectively, our results suggest that the expression of inhibitory co-receptors may be a biomarker of poor t cell reconstitution in transplanted scid patients. further, we propose that lack of conditioning limits t cell reconstitution, which correlates with increased expression of inhibitory receptors on circulating cd t cells. antibodies targeting inhibitory receptors are now available in clinical trials to treat cancer and viral infections. it will be necessary to evaluate the relationship between inhibitory receptor expression, t cell function and clinical outcome, to see if a selected group of scid patients could benefit from these immunotherapies. wallace chair, chief of allergy immunology, children's hospital of philadelphia introduction/background: prophylactic antibiotics (abx) and immunoglobulin replacement (igrt) are commonly used to treat specific antibody deficiency (sad), but the optimal therapy is not established. objectives: to compared outcomes (number of infections and hospitalizations) in sad treated with igrt vs. prophylactic antibiotics. methods: two-center, retrospective chart review of sad patients from jan -may . we excluded patients with hypogammaglobinemia and/or other immunodeficiency diagnosis. characteristics and treatment were reported, rates of infections/hospitalizations among treatment groups were compared using linear regression model. results: sad patients included. mean age was years, % were females. ( . %) received prophylactic antibiotics, ( . %) received igrt, ( . %) did not receive any specific treatment. number of infections decreased from . (year before treatment) to . (year after treatment) in prophylactic antibiotics group (p= . ), and from . to . in igrt group (p< . ). various musculoskeletal and vascular abnormalities. little is known of gynecologic-obstetric complications, but with improved therapies women are living longer making reproductive health more pertinent. objectives: to learn more about obstetric and gynecological health in women with stat loss of function. methods: we prospectively interviewed and retrospectively reviewed medical records of adult women with ad-hies evaluated at the nih between - . results: of patients aged - years (mean years), women were interviewed, and chart reviews were performed on . age of menarche in our cohort was consistent with the national average ( . vs. . years). five of patients ( . %) reported having worsening lung symptoms with menstruation, and of participants ( %) reported worsening eczema during menstruation. with regard to routine health maintenance, of women reported having regular cervical cytology testing; ( %) reported an abnormal result. of these , had hpv that responded to treatment or was hpv only not requiring treatment; had reactive changes due to yeast and ascus that was subsequently normal. mastitis and breast abscesses occurred in women. eleven women reported vulvar cysts/abscesses requiring drainage. nine women had progestin-releasing iuds placed, in some to suppress menses-associated vulvar eczema/abscess flares; no infectious complications were reported from progestin iud use. over % of women chose not to conceive given underlying disease. of women with pregnancies, women had live births, of ( %) had miscarriages, and of ( %) experienced recurrent pregnancy loss. three women whose pulmonary symptoms worsened during pregnancy were diagnosed with progression of parenchymal lung disease post-partum. one woman experienced worsening of skin manifestations. other reported postpartum complications included one wound infection (after caesarean) and one hemorrhage leading to hysterectomy. conclusions: as women with ad-hies are living longer, significant infectious and disease-related exacerbations related to both menstruation and pregnancy were observed in this patient population. it is important to focus on maintenance of their gynecologic and obstetric health and carefully monitor for these morbidities. finally, while these women may choose to attempt pregnancy, the risk of recurrent pregnancy loss and worsening disease warrants discussion. rna sequencing identifies aichi virus as the cause of chronic infection with lymphoproliferation in a patient with x-linked agammaglobulinemia objectives: we here present an xla patient with a complicated course in whom we detected aichi virus (aiv ). methods: case report: the patient was diagnosed with xla at age years, based on agammaglobulinemia with absent b cells and a known pathogenic mutation in btk (c c>t, p.r c). he had been suffering from recurrent respiratory and gastrointestinal infections since the age of months. he was started on immunoglobulin (ig) substitution, which resulted in complete control of infections with igg trough levels at g/l. however, at years of age he developed unexplained fever, refractory temporal epilepsy, hepatitis, progressive nephromegaly with chronic renal failure, splenomegaly, episodic diarrhea and growth failure. ultrasound identified multiple focal lesions in the liver, spleen and kidney. a liver biopsy showed severe chronic hepatitis with initial perisinusoidal fibrosis. serial kidney biopsies showed variable oligoclonal cytotoxic t cell infiltrates, suggestive of chronic viral infection. standard diagnostics failed to reveal a pathogen in blood or stool samples and on biopsies. results: we finally resorted to rna sequencing on a kidney biopsy sample. this technique identified aiv , a kobuvirus of the family picornaviridae, with a high read number. subsequently pcr confirmed the presence of aiv in both liver and spleen. results for cerebrospinal fluid, blood and feces are pending. conclusions: aiv is a picornavirus responsible for self-limiting gastroenteritis in humans. confirmatory pcrs on blood, csf and stool samples are ongoing. however, given the unequivocal result of the rna sequencing and confirmatory pcrs in other affected organs, we believe that the complications in this xla patient can be explained by aiv chronic infection. these findings confirm the potential of next generation sequencing techniques to identify infectious agents in patients with primary immunodeficiency. characterization and successful treatment of a novel autosomal dominant immune dysregulatory syndrome caused by a jak gain-offunction mutation. introduction/background: janus kinase (jak ) plays an essential, nonredundant role in the jak/stat signaling cascade, a key pathway in the control of hematopoiesis and immune function. significant progress has been made in elucidating the role of jak , but gaps in our knowledge still persist. to date, somatic gain-of-function mutations in jak have been linked to t-cell acute lymphoblastic leukemia. objectives: to understand and treat human jak gain-of-function mutations. methods: research study protocols were approved by our institutional review board. four members of the family (the affected children and their parents) were enrolled. written informed consent for genetic testing and participation was provided by the parents for their children. genetic, bioinformatic, biochemical and immunological investigations were performed. results: we describe the first known patients carrying a germ-line gainof-function mutation in jak . the clinical phenotype includes severe atopic dermatitis, markedly elevated peripheral blood eosinophil counts with eosinophilic infiltration of the liver and gastrointestinal tract, hepatosplenomegaly, autoimmunity, and failure to thrive. functional analysis established the gain of function phenotype caused by the muta- introduction/background: herpesviridae infection after hsct for hematologic malignancies, specifically cytomegalovirus (cmv), epstein-barr virus (ebv) and human herpes virus- (hhv- ), have been associated with various outcomes including post transplant lymphoproliferative disorder (ptld), graft-versus-host disease (gvhd) and mortality ( ) ( ) ( ) . patients with primary immunodeficiency may have different incidence and outcomes with respect to herpesviridae given their differences in age at transplant, conditioning choices and underlying disease susceptibility to this viral family. objectives: the objective of this study was to describe the incidence and outcomes of cmv, ebv and hhv- post hsct for the primary immunodeficiency population. methods: a single center retrospective chart review of primary immunodeficiency registry patients (research ethics board protocol no. ) who received hsct from january december was undertaken. patients who received gene therapy were excluded. antiviral prophylaxis was given according to institutional protocol. demographic and clinical data were collated and analyzed with microsoft excel . the primary outcome was incidence and time to dnaemia for cmv, ebv and hhv- . results: sixty one patients who underwent hsct for primary immunodeficiency from january -december were reviewed. diagnoses are noted in table . the average age of transplant was . months (range . - . ). transplant recipients received busulfan and cyclophosphamide conditioning ( with anti-thymocyte globulin (atg), with alemtuzumab added), transplants received busulfan, fludarabine and either atg or alemtuzumab, and received atg alone. six transplants were unconditioned. . % ( / ) of patients developed gvhd requiring systemic immune suppression. the overall incidence of cmv, ebv and hhv- post hsct for primary immunodeficiency was . % ( / ), . % ( / ) and . % ( / ) respectively. in those with severe or profound combined immune deficiency the incidence of cmv was . % ( / ), ebv . % ( / ), and hhv- . % ( / ). in the patients with chronic granulomatous disease, cmv incidence was % ( / ), and ebv and hhv- were both % ( / ). in recipients with pre-transplant negative pcr for ebv, cmv and hhv- (r-), time to dnaemia post transplant is seen in figure . ptld was seen in patients with rag and il r, both of whom were d+r-for ebv status with ebv dnaemia, and one of whom died attributed to ptld. two year mortality for all patients was % ( / ), with mortality . % ( / ) for those with either cmv, ebv or hhv- dnaemia vs . % ( / ) in those without (p= . ). disseminated cmv prior to transplant was attributed to cause of death for one case. conclusions: cmv incidence was rare, likely from screening for cmv negative hsct donors. cmv, ebv and hhv- dnaemia was not associated with differences in mortality in this cohort. ebv incidence was common, and ptld incidence was similar to previously published outcomes for hsct for other disease ( ) . the advent of cytotoxic t cell therapy for ebv may help abrogate this risk. professor, senior physician, ulm university medical center, pediatrics, germany introduction/background: new-onset aiha occurs in - % of pediatric patients post-hsct. incomplete immune recovery may predispose to immune dysregulation following hsct including autoimmune cytopenias. although prednisolone or other immunosuppressive drugs control most episodes, some patients respond incompletely to first or second line therapies including rituximab. objectives: we describe an innovative therapy for post-bmt aiha refractory to proteasome inhibition. three patients responded to anti-cd antibody (daratumumab) therapy after failing treatment with bortezomib. methods: we retrospectively evaluated data from three patients treated with daratumumab for post-transplant aiha. patients and were treated according to the positive response reported for patient . results: aiha occurred between - months following hsct. daratumumab was curative in patients, the third one had only transient response and relapsed months after this treatment had been initiated. following daratumumab patients no longer required any prbc transfusions. conclusions: in potentially life-threatening aiha in the context of hsct daratumumab may be an effective rescue therapy in combination with rituximab. early post-natal thymus development is strictly dependent on the level of foxn expression in tec these infants present severe t cell lymphopenia early in life, but in most cases their immune system gradually normalizes. however, the role of foxn haploinsufficiency in causing this phenotype is unclear. objectives: to analyze t cell development and tec phenotype in nu/+ mice of various age, in order to investigate whether foxn haploinsufficiency in mice results in impaired thymic development early in life, followed by progressive normalization, thereby recapitulating the human phenotype. methods: we analyzed groups of nu/+ and +/+ littermates, divided by age: day, - days, weeks. the number of etp and the distribution of cortical and medullary tecs (ctecs, mtecs) were analyzed by flow cytometry. maturation of mtecs was further assessed by staining for mhc-ii and aire. real-time pcr was used to analyze the expression of ccl , cxcl , dll , and scf, four key foxn target genes. the study was performed in accordance to niaid animal protocol lcim e. results: at day and - days of life, nu/+ mice showed a dramatic reduction of etps, both in terms of frequency and absolute numbers, as compared to +/+ mice. however, by weeks of age, the frequency and count of etps were comparable in nu/+ and +/+ mice. a slight but significant reduction in the frequency and absolute count of mtecs was observed in all three groups of nu/+ mice. additionally, the ratio between mtec expressing high levels of mhcii (mtechi) and those expressing low levels of mhcii (mteclo) was always higher in +/+ mice as compared to nu/+ mice. moreover, mtechi cells in nu/+ mice expressed lower levels of aire, the gene crucial for thymic negative selection of autoreactive t cells. finally, as compared to wild-type littermates, nu/+ mice showed reduced thymic expression of ccl , cxcl , dll , and scf at day . reduced expression of ccl persisted at day , but normalized at weeks. conclusions: these data indicate that foxn haploinsufficiency in mice leads to impaired thymic colonization by etps and abnormalities of tec differentiation and maturation early in life, followed by progressive normalization, thereby recapitulating what observed in newborns with heterozygous foxn mutations. these observations have important implications for the management of these infants, who should be monitored closely without rushing to definitive treatment for scid. introduction/background: foxn is the master regulator gene for the development and maturation of the thymic epithelial cells (tecs). by inducing the expression of chemokine receptors such as ccl and cxcl , foxn also allows the migration of early thymic progenitor (etp) cells from the bone marrow. lack of foxn leads to the nude (nu)/severe combined immunodeficiency (scid) phenotype in humans and mice. recently, a number of newborns have been identified with low t cell receptor excision circles (trecs) at birth, associated with heterozygous foxn mutations. these infants present severe t cell lymphopenia early in life, but in most cases their immune system gradually normalizes. however, the role of foxn haploinsufficiency in causing this phenotype is unclear. objectives: to analyze t cell development and tec phenotype in nu/+ mice of various age, in order to investigate whether foxn haploinsufficiency in mice results in impaired thymic development early in life, followed by progressive normalization, thereby recapitulating the human phenotype. methods: we analyzed groups of nu/+ and +/+ littermates, divided by age: day, - days, weeks. the number of etp and the distribution of cortical and medullary tecs (ctecs, mtecs) were analyzed by flow cytometry. maturation of mtecs was further assessed by staining for mhc-ii and aire. real-time pcr was used to analyze the expression of ccl , cxcl , dll , and scf, four key foxn target genes. the study was performed in accordance to niaid animal protocol lcim e. results: at day and - days of life, nu/+ mice showed a dramatic reduction of etps, both in terms of frequency and absolute numbers, as compared to +/+ mice. however, by weeks of age, the frequency and count of etps were comparable in nu/+ and +/+ mice. a slight but significant reduction in the frequency and absolute count of mtecs was observed in all three groups of nu/+ mice. additionally, the ratio between mtec expressing high levels of mhcii (mtechi) and those expressing low levels of mhcii (mteclo) was always higher in +/+ mice as compared to nu/+ mice. moreover, mtechi cells in nu/+ mice expressed lower levels of aire, the gene crucial for thymic negative selection of autoreactive t cells. finally, as compared to wild-type littermates, nu/+ mice showed reduced thymic expression of ccl , cxcl , dll , and scf at day . reduced expression of ccl persisted at day , but normalized at weeks. conclusions: these data indicate that foxn haploinsufficiency in mice leads to impaired thymic colonization by etps and abnormalities of tec differentiation and maturation early in life, followed by progressive normalization, thereby recapitulating what observed in newborns with heterozygous foxn mutations. these observations have important implications for the management of these infants, who should be monitored closely without rushing to definitive treatment for scid. introduction/background: immunoglobulin g rd is an immunemediated disease most commonly seen in middle-aged and older men. clinical features include autoimmune pancreatitis, salivary gland disease, orbital disease and retroperitoneal fibrosis. pathologic features include a lymphoplasmacytic infiltrate enriched in igg -positive plasma cells and fibrosis in a storiform pattern. laboratory evaluation usually reveals an elevated serum igg concentration, and glucocorticoids are often used as treatment in early stages of disease. however, disease may recur off of steroids, and prolonged illness or steroid non-responsive disease is usually treated with rituximab. objectives: the objective of this case presentation is to discuss a presentation of a igg rd is a young adult male. results: year old male adopted from thailand, initially presented to hospital with five days of intermittent abdominal pain, night sweats and worsening fatigue. ct abdomen and pelvis revealed bronchiectasis in lung bases, hepatic masses and soft tissue infiltration in the porta hepatis extending into the liver, pulmonary nodules (read as possible metastases), bilateral renal masses, retroperitoneal nodes and ileocolic intussusception secondary to possible lymphoma. one year prior to presentation he developed new onset bilateral cervical lymphadenopathy. biopsy revealed a heterogenous lymphoid population and lymphoma was ruled out. he had a repeat lymph node biopsies during the admission, which was all negative for malignant cells, and was compatible with reactive lymphoid tissue with plasmacytosis. immunophenotype showed % lymphocytes with normal cd /cd ratio, and % cd -,cd -, tcr ++ t cells. immunohistochemical stain of a lymph node fine needle biopsy showed a mixture of cd + b and cd + t cells, abundant cd +, igg+ plasma cells(pcs) , with some igg+ pcs(ct of the neck and chest was completed for possible staging and revealed cervical lymphadenopathy, nasal polyposis, a soft tissue mass vs. enlarged lateral rectus muscle (left orbit), and mediastinal lymphadenopathy. a diagnosis of autoimmune lymphoproliferative syndrome (alps) was presumed by the hematologic/oncologic team given his elevated igg ( , mg/dl) and elevated vitamin b (> ) with multiorgan involvement. alps panel revealed a mutation in faslg (allele , c.- c>t) which is a variant of uncertain clinical significance. alps criteria/scorewas + for / criteria (cd +cd +/hla dr ratio < . ). he had been referred to the nih for further management of alps before initial presentation to our office. immunologic work up revealed immunoglobulins of igg: , mg/dl, iga: mg/dl, igm: mg/dl; / protective streptococcal titers (> . mcg/ml); hib: . mg/l; negative quantiferon gold, and hiv-serology. at that point igg rd vs. castleman disease was suspected instead of alps. further work up showed a normal il- level, and human herpes virus was also negative. igg subsets showed an elevated igg ( mg/dl), igg ( mg/dl), igg ( mg/dl) and normal igg ( . mg/dl). excisional lymph node biopsy was recommended to rule out castleman syndrome or igg rd. a left salivary gland was removed and showed mainly igg positive pcs with no multicentric lymphocytic infiltrates. dilution of the patients serum to determine if there was a prozone affect that minimized the igg level showed an elevated igg level of . mg/dl. this is in the st percentile of all cases of igg rd in terms of serum igg concentration. he was diagnosed with igg rd, a form previously called mikulicz disease, which is comprised of lacrimal and parotid gland enlargement. rituximab was given initially because of the severity of his disease. after two cycles he showed decreased lymphadenopathy, notable weight gain and marked decrease in fatigue. conclusions: igg rd is a rare and complex immunologically based disease process rarely seen in children or adolescents. patients with igg rd often undiagnosed at initial evaluation. normal serum igg levels are seen in % of patients with igg rd and thus a normal igg level should not be used as a biomarker to make the diagnosis of igg rd or in treating this disease. furthermore, the possibility of having a prozone affect in measuring igg needs to be considered and evaluated. meticulous correlation of clinical, pathologic, and imaging findings is required to make the diagnosis. modelling human immune deficiency from novel missense mutations with orthologous heterozygous mutations engineered in mice by crispr/cas introduction/background: next generation sequencing has resulted in substantial progress in identification of mendelian immune deficiency syndromes. in some cases, however, putative causal mutations occur in single kindreds, or even individual patients. under these circumstances, functional analysis of patient derived cells combined with in vitro analysis of genetically manipulated cell lines can provide additional evidence in support of genetic causation, but this might not be conclusive. objectives: understanding how genetic defects result in complex syndromes of immune deficiency and immune dysregulation can be impossible to achieve in vitro. one method for overcoming these obstacles is to generate accurate mouse models of human immune deficiency methods: mouse models of human immune deficiency are a valuable tool in which the murine genome is engineered to introduce a mutation orthologous to that discovered in the patient. we have applied this strategy to elucidate causation and mechanism of immunological defect in several mutations affecting the nf-kb pathway. results: so far, defects in both canonical and non-canonical pathways of nf-kb activation have been shown to cause immune deficiency, often associated with immune dysregulation. we describe a known defects and novel putative defect identified in the canonical nf-kb pathway conclusions: crispr-cas mouse models can be used to elucidate mechanism of disease and provide compelling evidence that mutations are causative. introduction/background: primary immune deficiencies (pid) with or without immundysregulation are rare diseases resulting from monogenetic aberrations leading to either infections or autoimmune manifestations or both. early diagnosis and treatment are crucial for reducing morbidity and mortality. beside genetic diagnosis not commonly yet performed, current standard methods for early diagnosis are dependent on either fresh samples or limited to certain cell types. to overcome those limitations, especially in newborn screening, a novel technology of methylation-based qpcr can be applied. among the known epigenetic modifications, dna demethylation is the most stable and genomic loci with highly cell type specific demethylation sites can be identified. differential methylation can be measured from blood samples of limited availability, or with suboptimal storage. we previously showed that thymic-derived t population, and determine whether there are unique diagnostic and treatment considerations within this demographic. whitney goulstone , elizabeth tough executive director, canadian immunodeficiencies patient organization lpn, alberta health services introduction/background: primary immunodeficiency diseases (pids) represent a significant collection of immune system disorders that increase susceptibility to infection, which in some cases are serious or life-threatening. patients with pid often require immunoglobulin g (igg, commonly referred to as ig) replacement therapy to prevent infections and associated comorbidities. pid treatment, in addition to symptoms and associated social and emotional impacts, has a significant impact on patients quality of life (qol). information available on the real-world diagnosis, management, and outcomes of the canadian pid patient population is limited. objectives: to better understand diagnosis and treatment of canadian patients with pid, we surveyed canadian patients with pid associated with the canadian immunodeficiencies patient organization (cipo) the primary goal of the survey was to gain insight into the canadian pid patient population with regards to demographics, diagnosis, treatment, including regimes, qol, and communication and support . methods: the authors conducted a cross-sectional survey to measure health-related qol in a cohort of patients with pid. eligible participants were identified through the canadian immunodeficiencies patient organization (cipo). the questionnaire consisted of questions that covered patient-reported outcomes including diagnosis, qol, treatment regimes, and communication. results: surveys were returned by patients with pid. participants conveyed significant impact on qol including personal, occupational, financial, emotional, and social impacts as a result of pid symptoms, risks, and treatment logistics, limitations, and side effects. the most common diagnoses were related to b-lymphocyte disorders ( . %). common treatments include intravenous immunoglobulin (ivig; in hospital) and subcutaneous immunoglobulin (scig; at home), in addition to antibiotics and antifungals as required. respondents reported feeling average before treatment on a scale of - (mean . ± . ) with increased health after treatment (mean . ± . ). respondents felt current treatment was convenient (mean . ± . ) and were comfortable with self-infusions (mean . ± . ). conclusions: patients with pid are not uncommon in the canadian community, and in these patients pid is associated with a significant impairment in qol. experiences range with regards to a particular treatments advantages and disadvantages, cost, travel, and convenience. respondents hope to achieve improved qol through the following solutions: better treatment, improved infusions, gene modification, more research and clinical trials, a cure, and education and outreach. improved financial, medical, and social supports were also requested. introduction/background: severe combined immunodeficiency (scid), the most severe form of t cell immunodeficiency, is detectable through quantification of t cell receptor excision circles (trecs) in dried blood spots (dbs) obtained at birth. for many professional, humanitarian and financial reasons, newborn screening (nbs) for scid is warranted. implementation of this screening test is highly important where high frequency of consanguinity is known to exist. objectives: since october , israel has conducted national scid nbs. this important, life-saving screening test is available at no cost for every newborn in israel. methods: herein, we describe two years results of the israeli scid newborn screening (nbs) program. validation includes cbc, lymphocyte subsets, trec (in a different method), t cell receptor (tcr) repertoire and response to mitogenic stimulation. whole exome sequence (wes) for genetic detection, and next generation sequencing (ngs) to demonstrate tcr clonality are used as well, in some unsolved cases. results: of , births screened, ( . %) had abnormal trec in their first screen ( terms, pre-terms), ( . %) had a repeated abnormal trec also in their second screen and were referred for a validation process. fourteen scid patients were diagnosed, so far, through the nbs program in its first years, revealing an incidence of : , births in the israeli population. consanguine marriages and muslim ethnic origin were found to be a risk factor in affected newborns, and a founder effect was detected for both il r and dclre c deficiency scid. other diagnoses were made as follows: cases were found to have t cell lymphopenia; cases were diagnosed with syndromes; cases were found to have secondary t cell lymphopenia; cases were pre-term infants and cases were considered as false positive with normal evaluation. lymphocyte subset analysis and trec quantification in the peripheral blood appear to be sufficient for confirmation of typical and leaky scid and ruling out false positive results. detection of secondary targets (infants with non-scid lymphopenia) did not significantly affect the management or outcomes of these infants in our cohort. we could also report several perspectives regarding t cell development in non immunodeficient newborns that emerged from the accumulated data. conclusions: already in a short term, trec nbs in israel has achieved early diagnosis of scid and other conditions with t-cell lymphopenia, facilitating management and optimizing outcomes. this program has also enabled gaining insights on t cell development in health babies. (hct) . surprisingly, % of these infections were acquired during the interval between confirmation of the scid diagnosis and hct (heimall, blood ). to investigate further, in late we surveyed pre-hct management practices for scid patients at pidtc centers. physicians representing north american centers responded, including immunologists, transplant specialists and who identified as both. % of centers lacked a standard procedure for the management of infants with a positive nbs result. to confirm the scid diagnosis, basic t, b and nk cell flow cytometry was performed by most. testing for naïve t cells was generally included, but testing of lymphocyte mitogen proliferation was inconsistent. specialists were notified of a patients positive nbs test at median age . days ( - days) , and management of patients as scid was started at median age days ( - days). when unconditioned hct was anticipated, % of respondents began planning as soon as possible after diagnosis, while % awaited genetic testing results before hct. respondants consistently implemented pre-hct prophylaxis with trimethoprim/sulfamethoxazole ( %), fluconazole ( %) and immunoglobulin infusions ( %), although timing of initiation varied. palivizumab was used by %. there was little consensus regarding viral monitoring. although most physicians screened for cmv by blood pcr ( %), only about half routinely screened for ebvor adenovirus. while % of physicians started prophylaxis against double-stranded dna viruses in all patients, % did so only in selected situations, such as active genital hsv at the time of delivery, or when the mother was cmv-seropositive. although all centers used only acyclovir as antiviral prophylaxis, doses and timing varied widely: from - mg/kg/day, divided into or doses, continued in most centers until immune reconstitution. finally, % of physicians recommended that cmvseropositive mothers stop breast-feeding. there was no consensus on where patients should reside prior to hct. hospital or home were favored equally, although need for a reliable family was indicated by % as a criterion for home-based management. for hospitalized patients, % of centers required patients to be in a reverseisolation/positive-pressure room and over half required staff to wear gown, gloves, and mask. with regard to visitors, % required parents/ relatives to perform hand hygiene. approximately % did not require a gown, gloves, or mask for visitors. finally, there was no consensus on allowing siblings or friends to visit, but the majority permitted grandparents or other adult relatives. this survey revealed wide variability in the diagnostic pathway, viral surveillance, and isolation practices for scid patients, although pretransplant prophylaxis with immunoglobulin, fluconazole, and trimethoprim/sulfamethoxazole were utilized consistently. we conclude there is considerable opportunity to develop diagnostic and pre-hct management pathways for scid. prospective tracking of management practices could reveal which are important for avoiding pre-hct infections. evidence-based practice guidance is needed to maximize the potential to bring each scid patient identified via nbs to hct infection-free. ikaros/ikzf is an essential transcription factor expressed throughout hematopoiesis. in humans, somatic mutations in ikzf are linked to b-cell acute lymphoblastic leukemia (all), and germline heterozygous haploinsufficient mutations cause common variable immunodeficiency-like disorder with incomplete penetrance. herein, we report seven unrelated patients with an earlyonset novel combined immunodeficiency associated with de-novo, fully-penetrant, germline heterozygous dominant negative mutations affecting amino acid n in ikzf dna binding domain. patients presented with different infections, but pneumocystis jirovecii pneumonia was common to all. one patient developed a t-cell all. additional findings included decreased b-cells, neutrophils, eosinophils and myeloid dendritic cells as well as t-cell and monocyte dysfunction. t-cells exhibited a profound naïve/recent thymic emigrant/ t-helper phenotype and were unable to evolve into effector memory cells; monocytes failed to respond to different stimuli or facilitate t-cell activation. this new defect expands the spectrum of human ikzf -associated immunodeficiency diseases from haploinsufficient to dominant negative. the beta subunit of the il- receptor (il rb; cd ) is essential for il- and il- mediated signal transduction in a variety of hematopoietic cell types including t and nk cells. here we report the clinical and immunologic phenotypes of two siblings born to consanguineous parents harboring a variant in il rb, resulting in autoimmunity, with lymphoproliferation of cd + t and cd hi nk cells, and decreased regulatory t cell frequency. the proband presented with inflammatory enteropathy, failure to thrive, and disseminated cmv infection at months of age, and later developed atopy, lymphocytic interstitial pneumonitis, and red blood cell autoantibodies. his younger sister presented with severe autoimmune hemolytic anemia and cmv viremia at months of age, and later developed lymphocytic interstitial pneumonitis. whole exome sequencing and chromosomal microarray studies revealed a homozygous deletion within the highly conserved wsxws motif of the extracellular domain of il rb (c. _ delcctggagcc, p.pro _ser del). the deletion results in reduced il rb expression (cell surface and intracellular) in t and nk cells. the functional consequences of the defect include complete impairment of stat phosphorylation through the il- receptor but only partial impairment through the il- receptor, as well as a compensatory increase in serum il- and il- levels (> pg/ml). cd + t cell proliferation responses to in vitro t cell receptor (tcr) stimulation were reduced compared to an age-matched, healthy control, but partially rescued by supraphysiologic levels of il- and il- . despite reduced cd + t cell proliferation responses, the proband displayed a t cell population skewed toward cd + t cells, with an oligoclonal expansion of effector memory cells. arguing against effects of pervasive cmv infection alone, the sister displayed similar phenotypic and functional abnormalities at birth, prior to her cmv infection. our data suggest that the identified hypomorphic mutation in il rb results in a survival advantage for those cd + t and cd hi nk cells most sensitive to the exuberant serum il- and il- levels produced in response to the defect. these surviving cd + t and cd hi nk cells have great lymphoproliferative potential, leading to multisystem autoimmunity and inflammatory complications. therefore, we describe il rb deficiency as a novel primary immunodeficiency disease with prominent immune dysregulation and selective cd + t and cd hi nk cell lymphoproliferation. key: cord- - c ut authors: nan title: cis annual meeting: immune deficiency & dysregulation north american conference date: - - journal: j clin immunol doi: . /s - - -z sha: doc_id: cord_uid: c ut nan abstract/case report text oral lichen planus (olp) is a t-cell mediated chronic inflammatory tissue reaction in which presentation can range from asymptomatic plaques to painful, erosive, bullous, or ulcerative lesions. here, we present a year-old female with a novel ctla- variant, multiple autoimmune conditions, and unusual tongue lesions. our patient was healthy until years of age when she developed hashimoto's thyroiditis. at , she developed psoriasis. at , she was diagnosed with alopecia totalis and epstein-barr virus (ebv) with resultant and persistent anemia, thrombocytopenia, lymphopenia and neutropenia. she had chronic abdominal pain and diarrhea since age . esophagogastroduodenoscopy revealed lymphocytic esophagitis and active duodenal inflammation with increased intraepithelial lymphocytes. colonoscopy revealed mildly active chronic colitis with eosinophils. whole exome sequencing revealed a heterozygous c. dela (p.q rfs* ) pathogenic mutation in exon of ctla- . family history is remarkable: father (splenomegaly and psoriasis) and brother (autoimmune hemolytic anemia) have ctla haploinsufficiency with the same mutation. abatacept was initiated with re-growth of hair, improvement in cytopenias, improvement in psoriasis, and some reduction of gastrointestinal symptoms. since her abdominal pain persisted repeat endoscopies after six months of abatacept revealed persistent active lymphocytic esophagitis with some improvement in inflammatory injury in her duodenum and colon. physical exam revealed glossitis with a gel-like coating and ulceration on her tongue, xerosis along her face and scalp without other abnormalities ( figure) . she denied recent dental procedures, appliances, or tongue biting. her wbc ranged from - x ^ cells/l and hemoglobin . - . g/dl. absolute lymphocyte count ranged from . - . x ^ cells/l. immunologic evaluation revealed low iga and pan-low lymphocyte subsets (table) . ebv pcr ranged from - , copies/ml. tongue scraping revealed candida dubliniensis and she responded to days of fluconazole. two months later, she developed painful white patches along her tongue and subsequent kilogram weight loss recalcitrant to viscous lidocaine, antacids, and days of fluconazole. incisional tongue biopsy revealed ulceration with underlying granulation tissue with lymphocyte and plasma cell infiltration consistent with olp ( figure) . periodic acid-schiff diastase stain and grocott stain were negative. aerobic culture was normal. no fungus was isolated within days. epstein-barr encoding region in situ hybridization was negative. two weeks of topical dexamethasone lead to temporary improvement. her tongue lesions waxed and waned over the following months. due to persistent psoriasis, methotrexate was initiated without worsening in her tongue lesion. to our knowledge, this is the first case of olp reported in a patient with ctla- haploinsufficiency. ctla- haploinsufficiency may present with variable clinical phenotypes including increased risk of ebv viremia and malignancies. therefore, after ebv and malignancy are ruled out, olp may be a prudent diagnosis to consider in a ctla insufficient patient with unusual oral lesions. mutations of bcl b appear to be associated with lymphoproliferation and autoimmunity as well as susceptibility to herpes virus infections. additional research focusing on characterization of dna binding sites of bcl b as well as the downstream expression of associated target genes is needed. these data combined with longitudinal analysis of additional patients with confirmed bcl b mutations, will help clarify determinants of bcl b pathogenesis and highlight potential therapeutic strategies. consanguineous marriages in tribal cultures, such as that in the united arab emirates significantly increase the prevalence of autosomal recessive disorders. premarital genetic screening and counseling, thus, are expected to reduce the frequency of these diseases. in this pilot study, diagnostic exome sequencing was used in the premarital screening program to identify recessive pathologic variants preventable by premarital counseling. a total of pathologic or likely pathologic variants were identified in studied emiratis ( couples), averaging . variants per person. four percent of the persons had negative diagnostic exome sequencing; the remaining had one to eight variants per person. of the distinct variants, ( %) were novel. twenty ( %) couples had pathologic or likely pathologic variants of inborn errors of immunity (iei). two couples ( %) had iei pathologic or likely pathologic heterozygous variants in both partners imposing risk for autosomal recessive disease in the offspring. other eighteen couples ( %) had pathologic/ likely pathologic heterozygous variants present in only one person of the couple. total of sixteen ( . %) iei variant identified and eight ( %) were novel. fourteen known phenotypic iei diseases were recognized (table. ). these preliminary results support a need for nationwide premarital genetic screening, and primary immunodeficiency registry to identify common and novel pathogenic variants with high heritability rate. these results will aid adopting a preand post-connectional reproductive carrier counseling to reduce autosomal recessive diseases. also, it will assist the diagnosis of these complex diseases in our community. table pathologic or likely pathologic variants of primary immunodeficiency (pid). abstract/case report text background: chronic granulomatous disease (cgd) is a primary immunodeficiency disorder caused by defects in the phagocytic nadph oxidase complex, leading to increased susceptibility to infection and inflammatory or autoimmune disease. up to % of patients have gastrointestinal (gi) involvement and meet diagnostic criteria for inflammatory bowel disease (cgd-ibd). objectives: we analyzed cgd patients from the united states immunodeficiency network (usidnet) registry to determine whether ibd may change the presentation, treatment, and outcomes of cgd patients, as compared to those without ibd. methods: a retrospective evaluation of cgd cases from the usidnet registry was completed. cgd-ibd was defined as the presence of any major physician-reported inflammatory, non-infectious gi tract disease manifestation, including crohn disease, ulcerative colitis, ibd endoscopy findings, gi fistulas, gi strictures, gi obstruction, and proctitis. demographic information, genotypes, symptoms and conditions, infections, antimicrobial therapies, immunomodulator use, and allogeneic hematopoietic stem cell transplantation (hsct) data were analyzed. results: patients with a diagnosis of cgd were identified. met criteria for ibd; were categorized in the non-ibd group. crohn disease and colitis were the most common gi disease manifestations in the cgd-ibd group (n= ), followed by gi fistulas (n= ). cgd-ibd patients had an increased average frequency of infections ( . events/patient) compared to the cgdnon-ibd group ( . events/patient). in both groups, lower respiratory tract infections were the most common infection type and aspergillus was the most common organism. enteric organism infections were more common in ibd patients. temporal data regarding the timing of infections were not available. immunomodulators, including biologics and interferon-gamma, were used at a significantly higher rate in ibd patients compared to non-abstract/case report text down syndrome (ds) is characterized by the occurrence of three copies of human chromosome (hsa ). these patients often develop chronic mucocutaneous candidiasis (cmc) and autoimmune thyroiditis, mimicking patients with heterozygous gain-of-function (gof) stat mutations, which enhance cellular responses to the three types of interferon (ifn). hsa contains a cluster of four interferon receptor (ifn-r) genes: ifnar , ifnar , ifngr and il rb. a gene dosage effect at these four loci may contribute to the infectious and autoimmune manifestations observed in individuals with ds. we report high levels of ifn-αr , ifn-αr and ifn-γr expression on the surface of monocytes and ebv-transformed-b (ebv-b) cells from ds patients. levels of ifn-ɣr , encoded by a gene on chromosome , were similar in the immune cells of ds patients and healthy controls. total and phosphorylated stat (stat and pstat ) levels were constitutively high in unstimulated and ifn-α-and ifn-γ-stimulated monocytes from ds patients, although less so than those in patients with gof stat mutations. following stimulation with ifn-α or -ɣ, but not with il- or il- , pstat and ifn-ɣ activation factor (gaf) dna binding activities were significantly higher in the ebv-b cells of ds patients than in controls, this response resembling the dysregulated responses observed in patients with stat gof mutations. plasma type i ifns concentrations were high in about % of the ds patients tested. a genome-wide transcriptomic analysis involving principle component analysis and a comparison of interferon modules was performed on circulating monocytes. it showed that ifn-stimulated genes (isgs) were expressed more strongly in ds than in controls. ds monocytes have intermediate levels of ifn-α-and ifn-γ-induced isgs relative to monocytes from healthy controls and from patients with gof stat mutations. by contrast to patients with gof stat mutations, circulating th counts were normal and the proportion of terminally differentiated cd + t cells was high in ds patients. the constitutive upregulation of type i and type ii ifn-r, at least in monocytes of ds patients, may therefore contribute to the autoimmune diseases observed in these individuals. scid screens (positive screen defined as trec values less than units/ul per statewide criteria). results forty nine neonates were identified with low trec values. ( %) of these infants had repeat trec screening, ( %) of which were found to have a positive second trec screen. lymphocyte subsets were evaluated in of these infants and of which were noted to have lymphopenia (defined as absolute lymphocyte count less than ). infants were noted to have low cd levels (defined as < cells/ul) and were noted to have low cd levels (defined as less than cells/ul). of note, % of the infants with cd and cd lymphopenia had normal repeat trec levels. of the infants were noted to have low b cell levels (defined as < cells/ul). infants had quantitative immunoglobulin levels and of these two were noted to have igg levels less than . of note one infant was diagnosed with partial digeorge syndrome via microarray. in our study population, no infants were diagnosed with scid. discussion our study shows that testing for trec levels on newborn screen may be beneficial in identifying not only scid, but also other immunologic conditions. infants in our study had evidence for both cell mediated and humoral immunodeficiency which necessitated further workup and follow up from allergy and immunology specialists. it may be beneficial to develop further programs to track infants identified with abnormal trec levels on newborn screens to determine if they develop signs of immunodeficiency syndromes later in life. abstract/case report text the patient was transferred to our adult clinical immunology transition clinic for low igg and iga, elevated igm and b cell lymphopenia, treated with subcutaneous gamma globulin. his medical history was relevant for recurrent respiratory infections since two years-old, failure to thrive and developmental delay. he also developed chronic auto-immune hemolytic anemia (aiha) at ten years old, accompanied by prominent lymphoid hyperplasia. our initial evaluation at the age of twenty years old showed massive polyadenopathy and splenomegaly. work-up confirmed flair-up of aiha. at that point, the diagnosis of apds was raised. he also had mild intellectual impairment and dysmorphic features such as a mild degree of ocular depression, deep-set eyes, vaguely triangular face, small chin, but had normal stature. a customized panel for usual genes involved in classic hyperigm syndromes, apds, noonan and kabuki syndromes came back negative. at years old, an urgent coloscopy was performed because of acute abdominal pain and showed diffuse ileal lymphoid hyperplasia. biopsies confirmed reactional lymphoid hyperplasia without infection nor malignancy. a second genes ngs panel associated with pid identified a heterozygote mutation in tap ; expression of hla class was normal on flow cytometry. the patient was then started on sirolimus for an "apds-like syndrome" despite the lack of genetic confirmation. six months after introduction of mtor inhibitor, his abdominal pain had completely disappeared. tep scan showed complete resolution of axillar, retroperitoneal and inguinal lymph nodes and significant regression of splenomegaly. a third large non-biased + genes ngs panel revealed a pb de novo deletion that included the splice site of pik r exon typically involved in apds : c. _ + del p.(asp glufs* ) , which was missed by the first two panels. indeed, oligonucleotide-selective sequencing technology used for the previous panels was associated to mapping errors of short reads and difficult detection of large deletions. interestingly, the patient also presented some but not all dysmorphic features of short syndrome which is related to pik r haploinsufficiency. in this new era of genetic testing, this case is a reminder that we need to be aware of the pitfalls of genetic tests and that clinical judgment is still our best diagnostic tool. abstract/case report text intro: patients with chronic granulomatous disease (cgd) are theorized to have a lower risk of malignancy related to their lack of free radical formation. there are relatively few reports of malignancy described in patients with cgd. we report three cases of malignancies in the large cohort cgd population at the national institutes of health followed between - to add to the seven cases described in the literature. case : a -year-old man with x-linked chronic granulomatous disease with a history of severe inflammatory bowel disease, who presented with progressive left-sided chest pain in , decreased appetite and weight loss. transthoracic lung biopsy showed atypical cells and a pet/ct showed abnormally dense mesentery and widespread hypermetabolic abnormalities. a mesenteric biopsy showed metastatic pancreatic adenocarcinoma. palliative care was initiated. patient expired four months after diagnosis. case : a -year-old man with x-linked chronic granulomatous disease and severe inflammatory bowel disease requiring total proctocolectomy and who had been remotely treated with infliximab, presented in with right upper quadrant pain. abdominal ultrasound and mri of the liver showed multiple liver lesions. biopsy of these lesions revealed hepatocellular carcinoma. patient underwent two courses of radiolabeled itrium spherules. however, his disease progressed and he expired approximately five months after diagnosis. c a s e : a n -y e a r-o l d m a n w i t h x -l i n k e d c h r o n i c granulomatous disease and inflammatory bowel disease who presented in with fevers, abdominal pain and pancytopenia. during the course of his hospitalization, he developed sepsis which led to his demise. on autopsy, an incidental finding of papillary thyroid carcinoma was made. discussion: these three patients all had poorly controlled inflammatory bowel disease. additionally, patients with cgd are typically exposed to higher doses of radiation, leading one to expect higher rates of radiation induced malignancies. however, there are still relatively few case reports of cancer in the cgd population. tissue biopsy is necessary for diagnosis. due to end organ damage secondary to the underlying disease in the first two cases, treatment options were limited. managing infections during chemotherapy can be complex due to drug interactions with chemotherapeutic agents. abstract/case report text myeloperoxidase (mpo) deficiency is the most common inherited defect of phagocytes that impairs microbial killing since the toxicity of the respiratory burst is dampened without myeloperoxidase release from the azurophilic granules. a significant portion of these patients remain asymptomatic, however there is a clinically variable phenotype that can present if they do become symptomatic. fungal infections with candida strains appear to be the most frequently reported. we present an adulthood case of recurrent invasive candidal disease due autosomal recessive myeloperoxidase deficiency from a pathogenic missense variant in the mpo gene (c. c>t (p.arg trp)). a -year-old caucasian male was in his normal state of health without any major illnesses until years of age when he was diagnosed with candida osteomyelitis of the heel, followed by cryptococcal meningitis the following year which ultimately required a ventriculoperitoneal shunt. in , he had a prolonged hospitalization after presenting with lethargy, headache and vomiting that culminated in seizure activity and prompted an emergency room visit. imaging at the time showed ventriculomegaly, and fluid from the shunt revealed yeast, but no bacteria. he was started on broad spectrum antifungal therapy and admitted for further management. cerebral spinal fluid and blood cultures confirmed invasive candida albicans meningitis. during this hospitalization, he also developed sepsis secondary to serratia marcescens. because of the pathogens that were being isolated, our service was consulted. of note, our patient does not have diabetes mellitus. a neutrophil oxidative burst assay showed an absent respiratory burst compared to control. a primary immunodeficiency panel to identify genetic variants was also sent to invitae. variants in cyba, cybb, ncf , and ncf were not identified, making chronic granulomatous disease less likely. peroxidase staining was negative on neutrophils and normal on eosinophils, suggesting a diagnosis of mpo deficiency. this led to mpo gene sequencing for deletion and duplication analysis. a homozygous pathogenic variant consistent with a molecular diagnosis of a mpo related condition was identified. immunoblotting of patient-derived immune cells demonstrated an absence of mature enzyme. although not typically indicated, given the severity of his presentation, our patient remains on fluconazole for long term prophylaxis. his younger brother also had a history of invasive disease with candidaosteomyelitis and meningitis. a neutrophils oxidative burst assay showed similar results in his brother and similar results with peroxidase staining, also suggesting a diagnosis of mpo deficiency. confirmatory genetic testing has not been performed yet. their father, who reported severe skin infections with candida, had peroxidase stains performed on neutrophils and eosinophils which were both normal. we have presented a patient without a significant history of diabetes mellitus who developed invasive disease from candida and serratia and was ultimately diagnosed with myeloperoxidase deficiency. abstract/case report text introduction: juvenile xanthogranuloma (jxg) is an often benign, histiocytic proliferative disorder of the mononuclear phagocytic system. patients typically present with localized cutaneous lesions. systemic disease, especially central nervous system involvement, rarely occurs but has significant morbidity and mortality risk. no standard evaluation nor therapy regimen exists for systemic jxg and little is known about the genomic alterations underlying its pathology. case report: a full-term male infant presented at months of age with post-prandial abdominal pain, fevers, altered mental status and weight loss. abdominal ultrasound and ct identified renal masses. a chest ct was obtained showing a paraspinal mass with possible neural foramina extension. mri brain and total spine was consistent with diffuse leptomeningeal disease involving the left frontal convexity, brainstem, cerebellum, and multiple cranial nerves. abnormal enhancement was also present along the entire surface of the spinal cord extending into the cauda equina with additional enlargement of the cervical/upper thoracic cord with intramedullary enhancing masses and a right paraspinal mass. renal biopsy yielded a pathologic diagnosis of disseminated jxg. integrative clinical sequencing of the mass identified a somatic driving alk rearrangement (kif b-alk in-frame fusion). tumor and matched germline dna sequencing did not detect any alterations in the ras/mapk pathway. bone marrow biopsy was negative for disease with cerebrospinal fluid analysis showing numerous monocytes and macrophages consistent with jxg. the patient was started on therapy consisting of systemic dexamethasone, intrathecal methotrexate/hydrocortisone and systemic intravenous cytarabine. his first cycle was complicated by pseudomonas aeruginosa bacteremia and gangrenous cellulitis of the perianal region, treated with systemic/topical antibiotics and topical gm-csf. following completion of the initial cycle of therapy, the patient was noted to have declining neurologic status, including seizure-like activity. repeat mr imaging revealed worsening cns disease with new subdural fluid collection and progression of leptomeningeal enhancement and intramedullary cervical lesion. in light of disease progression, the decision was made to continue dexamethasone treatment, but add adjunct intrathecal cytarabine, and transition to targeted alk inhibition via daily oral ceritinib, given its predicted cns penetrance followed by ceritinib in combination with systemic intravenous clofarabine. significant clinical and radiographic improvement was noted with the new targeted treatment regimen. ceritinib therapy was tolerated well overall after a % dosing reduction made for initial grade gastrointestinal toxicity and grade hypertriglyceridemia (non-life threatening but level > mg/dl). following continued treatment with daily ceritinib and completion of cycles of clofarabine therapy, our patient experienced complete disease remission. he continues to do well on daily ceritinib monotherapy with plan to complete an additional year of therapy. conclusion: our report highlights the potential benefit of real-time integrative clinical sequencing in the management of systemic histiocytic lesions, specifically non-langerhans cell conditions. it has the potential to identify novel somatic genetic alterations, other than the typical lchassociated braf mutations of the mapk pathway, that may be therapeutically targetable. treatment with nd generation alk-inhibition in our pediatric disseminated jxg patient was a novel, biologicallyrationale management approach with minimal toxicity and potentially contributed to his complete remission. abstract/case report text background: c glomerulonephropathy (c gn) is a progressive kidney disease with the predominant pathological feature of c deposits around the glomerular capillaries. c gn patients suffer from dysregulated activation of the alternative pathway as the result of autoantibodies or congenital genetic defects that stabilize cleavage of c . despite therapy involving immunosuppression and complement-pathway inhibition, the prognosis for c gn is poor. we report a patient with autoantibody-mediated, refractory c gn who demonstrated no improvement on rituximab but achieved sustained remission on bortezomib. follow up studies after one year demonstrated clearance of the culprit autoantibody, normalization of c levels, and improved pathologic appearance of the kidneys. this case supports the idea that c gn is frequently driven by pathogenic autoantibodies that may not clear with rituximab alone. plasma cell directed therapy has the potential to clear these autoantibodies and halt the progression of disease. case presentation: we report the case of a hispanic male with chronic renal dysfunction initially diagnosed with membranoproliferative glomerulonephritis on renal biopsy at years of age. despite cellcept and prednisone, over the next years, he had worsening proteinuria and an increase in protein-to-creatinine ratio. renal biopsy suggested c gn, and lab studies revealed a factor-h autoantibody and c level below the assay limit of detection. after initiating eculizumab, the proteinuria temporarily improved; however, the proteinuria eventually worsened, and he was referred to immunology. we hypothesized that the factor h-binding autoantibody was the cause of dysregulated c cleavage and disease progression, and blocking the terminal complement pathway with eculizumab would not halt upstream c -mediated kidney injury. at the age of , rituximab and plasmapheresis were administered to clear the factor-h autoantibody. three months after rituximab administration, the factor-h autoantibody level decreased to the normal range, but he continued to have significant proteinuria with low serum albumin and undetectable c level. we concluded that the relevant autoantibody was not solely produced by differentiating memory b cells, so we decided to target the plasma cell compartment. bortezomib was started at the age of , and eculizumab was continued given his initial response to treatment. after adding bortezomib, factor h autoantibody levels dropped below prior levels and serum c level normalized. renal biopsy at the age of showed evidence of imp r o v i n g c d e p o s i t i o n a n d l e s s p r o m i n e n t g l o m e r u l a r hypercellularity, with stable mesangial hypercellularity, interstitial fibrosis, tubular atrophy, and sclerotic glomeruli. although his proteinuria did not worsen, it remained persistent, suggesting that earlier introduction of bortezomib could have prevented disease advancement. there has been no further progression of kidney failure. conclusions: the majority of c gn patients harbor autoantibodies to components of the alternative pathway of complement. this case provides evidence that at least some of these autoantibodies are indeed the cause of complement dysregulation, and thus are prime targets for therapy. b cell targeting therapies may be inadequate to decrease autoantibody levels for some patients. early initiation of bortezomib, or other plasmacell directed therapy, may effectively induce complement normalization and disease remission in these cases. inhaled corticosteroid and a long-acting bronchodilator. he has no family history of immunodeficiencies or congenital disorders. computed tomography(ct) chest showed bronchiectasis. his complement studies and isohemagglutinin titers were also normal. his serum immunoglobulin(ig) and lymphocytes on presentation are shown in table . he had a poor response to polysaccharide pneumococcal vaccination. he was diagnosed with combined igg /igg subclass/iga deficiency and was started on immunoglobulin replacement therapy and prophylactic rotating antibiotic therapy. thereafter, his clinical course markedly improved with a reduction in the frequency of rti's as well as the number of bronchiectasis exacerbations. there was high suspicion for an underlying genetic disorder based on his constellation of neurodevelopment disorders and immunodeficiency. cytogenetic evaluation with array comparative genomic hybridization(cgh) analysis showed duplication of xq and xq consistent with mds. discussion: mds is caused by duplications involving the mecp gene locus of the x chromosome at xq . it has a % penetration rate in males whereas females act as carriers and are usually unaffected. rarely, cases of de novo mutations causing mds have been reported. chromosome microarray analysis is currently the best initial clinical test when mecp duplication syndrome is suspected. management needs a multidisciplinary approach involving geneticists, neurologists, ophthalmologists, physical medicine and rehabilitation specialists, psychologists, gastroenterologists, and allergy and immunology specialists. prophylactic treatment with ivig and antibiotics has been the standard of care for immunodeficiency in these patients. prognosis is guarded and most male patients die in the mid to late 's because of severe rti's secondary to immunodeficiency. conclusions: this case confirms the association of mds with combined iga and igg subclass deficiencies. clinicians should consider pursuing genetic evaluation for mds in patients with neurodevelopmental disorders and immunodeficiency because the diagnosis of the syndrome can change the overall approach to management and expectations in prognosis. abstract/case report text introduction: c nephritic factor is an autoantibody that binds to the alternative pathway c convertase (c bbb). this results in unchecked overactivation of the alternative complement pathway, which can lead to renal disease, partial lipodystrophy, retina disease, and frequent infections. in this case, we present a patient with partial lipodystrophy and low c , subsequently found to have c nephritic factor. case description: a year old female presented with a month history of low c levels. she was diagnosed months ago with poststreptococcal glomerulonephritis (psgn) after presenting with hematuria and elevated aso titers. she had c levels drawn - months after time of diagnosis and c level was low at (normal range - ), which was consistent with psgn. it was rechecked months after time of diagnosis and was still low. she was referred to rheumatology at this time and was found to have a positive ana titer : . tests for lupus and anti-phospholipid syndrome were negative. c normalized to the low-normal range at months after time of diagnosis to . her pediatrician checked to make sure it remained normal around months after initial diagnosis and c was low again at . c was normal at . she was referred to immunology for further evaluation. during this time she was asymptomatic with no fevers, infections, hematuria, rashes, joint pain, or joint swelling. she has no history of hospitalizations other than the first for psgn. mother denied family history of autoimmune disorders. physical exam: physical exam was notable for abnormal subcutaneous facial fat with normal fat distribution in the rest of her body. the rest of the exam was unremarkable with normal cardiac, pulmonary, abdominal, and skin exam. testing: c level was rechecked and low at . c nephritic factor was elevated at . (normal range . - . ). alternate pathway complement (ah ) was confirmed twice and was undetectable, < (normal level greater or equal to ). total hemolytic complement (ch ) was low at (normal level - ). other complement levels were checked and c q, c , c , c , c , c , c , and c complement were within normal range. discussion: the overactivation of the alternative complement pathway by c nephritic factor can result in various clinical manifestations, such as c glomerulopathy and acquired partial lipodystrophy in predominantly the face and the upper torso. the exact mechanism of how c nephritic factor is related to facial and upper body lipodystrophy is not known. one proposed mechanism is that adipocytes in the face and upper body produce more factor d, which is a complement protein utilized by c nephritic factor. overactivation of the alternative complement pathway on the adipocyte then leads to formation of the membrane attack complex, resulting in adipocyte lysis. eye disease, such as retinitis pigmentosa and macular degeneration can develop. c nephritic factor can also lead to more frequent infections and renal disease. patients need to be closely monitored. if patients develop c glomerulopathy, they may need to be considered for immunomodulatory therapy, such as steroids and other immunosuppressants. project manager/ucsf benioff children's hospital senior clinical research associate/ucsf benioff children's hospital associate professor/department of clinical pharmacy, ucsf staff research assistant iv/ucsf benioff children's hospital senior supervisor/ucsf benioff children's hospital laboratory specialist/ucsf benioff children's hospital research specialist/ucsf benioff children's hospital assistant professor/ucsf benioff children's hospital clinical professor/ucsf benioff children's hospital assistant professor/ucsd rady children's hospital staff pediatrician/tuba city indian health service staff pediatrician/phoenix children's hospital associate professor/seattle children's hospital chief, genetic immunotherapy section/niaid, nih professor/university of minnesota professor/ucsf benioff children's hospital abstract/case report text background: artemis-deficient scid (art-scid) represents % of all scid, but occurs in / births in navajo and apache native americans. artemis protein, encoded by dclre c, is essential for repairing dna double-stranded breaks, including those generated during v(d)j recombination of antigen receptor genes as t and b cells develop. artemisdeficiency causes not only t-b-nk+ scid, but also increased sensitivity to alkylating drugs and radiation. art-scid is the most difficult scid to treat with allogeneic hematopoietic cell transplantation (hct) due to high rates of rejection and gvhd, incomplete immune reconstitution, and toxicity following intensive conditioning regimens. as an alternative, we developed a self-inactivating lentiviral vector containing the human artemis promoter and dclre c cdna (aproart). we are evaluating its toxicity and efficacy in a phase i/ii gene transfer trial in art-scid patients. methods: newly diagnosed infants with art-scid and older patients with insufficient immunity despite prior allogeneic hct were eligible if organ function was acceptable. infants needed to have no matched sibling donor and be at least months old at conditioning. cd + cells were isolated from bone marrow or cytokine-mobilized peripheral blood, cultured with cytokines, transduced x with aproart, and cryopreserved. patients received daily doses of busulfan, targeted for a cumulative exposure (cauc) of mg*hr/l, with infusion of thawed cells on the following day. results: we treated newly diagnosed infants (art - & - ) with median age . m (range . - . ) and previously-treated patients (art - ) ( . y, . y and . y), with a median follow-up of . m (range . - . ). the mean (sd) bu cauc was . ± . mg*hr/l. patients received a median of . x aproart-transduced cd + cells/kg (range . - . ). the average vector copy number (vcn) and transduction efficiency in the marrow grafts exceeded those in the pbsc grafts: . ± . copies/cell vs . ± . (p= . ) and ± % vs ± . % (p= . ), respectively. there were no serious busulfan side effects. all patients had transduced peripheral blood leukocytes by w and of developed gene marking in t, b, nk and myeloid cells by w (fig. ) . gene-corrected cd , cd , cd / ra/ccr , cd and cd cells appeared in of patients (fig. ) , with art having t, nk and myeloid marking without b cells at m post infusion. normalization of lymphocyte proliferation to pha occurred in the evaluable (> w) infants (fig. ) , all now outpatients off isolation. two infants and previously treated child developed autoimmune hemolytic anemia (aiha), with requiring immunosuppressive therapy. infections included rhinovirus at presentation in art that resolved with t cell reconstitution. after discharge art acquired and recovered from norovirus and art acquired and recovered from cmv and rotavirus. analyses of insertion sites and t cell receptor diversity are pending. conclusion: infusion of aproart-transduced autologous cd cells into art-scid patients pretreated with very low exposure busulfan resulted in multilineage engraftment of transduced cells with evidence for t and b cell immune development. aiha, the only complication to date, occurred early and appears to resolve following restoration of t cell immunity. these encouraging results suggest potential effectiveness of ex vivo gene therapy for art-scid. ( ) submission id# mailan nguyen, md , susan canny, md, phd , andrea ramirez, md , ivan chinn, md abstract/case report text background: systemic lupus erythematosus is a heterogeneous disorder of the immune system. systematic genetic evaluation of patients with childhood-onset sle (csle) has begun to identify phenotypic clusters of csle patients with classic sle-causing genetic variants, as well as revealed unexpected genetic mimics of lupus. we report patients diagnosed with csle with similar typical and atypical lupus features, who were subsequently found to carry pathogenic nras variants that are the cause of ras-associated autoimmune leukoproliferative disorder (rald). cases: all patients ( females, male) presented at < years of age (average age . months, range - months) with antinuclear antibodies, anti-double-stranded dna antibodies, autoimm u n e h e m o l y t i c a n e m i a , s e v e r e t h r o m b o c y t o p e n i a , antiphospholipid antibodies, hypocomplementemia and nephritis. additionally, the patients all displayed fevers, organomegaly, lymphadenopathy and hypergammaglobulinemia. two out of patients had a malar rash, leukopenia, lymphopenia, anti-smith antibodies, serositis or arthritis. no patient had oral or nasal ulcers or photosensitivity. despite the fevers, lymphoproliferation and systemic autoimmunity, the patients did not display overwhelming immune dysregulation (peak ferritin - ng/ml). interestingly, the patients were found to have monocytosis ( - %), as has previously been reported in rald. double negative t cells were within normal range in the patients in which this was tested. all patients required aggressive immune modulation for control of their disease manifestations. two of the developed severe infections, specifically pneumococcal sepsis, during therapy. current follow-up covers an average of . years (range . to years). the patients responded to corticosteroids and were given sequential trials of various steroid-sparing therapies. in general, they appeared to benefit from both b cell depletion and t cell-directed modalities (cyclosporine, rapamycin), which are not first line therapy in csle. unfortunately, patient developed a fatal pulmonary infection while on treatment; her underlying disease was felt to be quiescent. due to the early-onset of disease, each patient was selected for genetic evaluation ( by exome sequencing, by gene panel). this lead to the discovery of pathogenic nras variants (c. g>a, p.g d) in all patients, assumed to be somatic, although this was confirmed in only case. conclusion: ras-associated autoimmune leukoproliferative disorder can present indistinguishable from csle with positive autoantibodies, immune cytopenias, arthritis, nephritis and hypocomplementemia. clinicians should consider evaluating for rald in csle patients who present at an early age ( < years) with predominant features of lymphoproliferation and hematologic abnormalities, particularly monocytosis. t cell-directed therapy with cyclosporine or rapamycin should be considered for rald. ( ) human ctla loss-offunction causes dysregulation of foxp + regulatory t (treg) cells, hyperactivation of effector t cells, and lymphocytic infiltration of target organs. patients also exhibit progressive loss of circulating b cells, associated with an increase of predominantly autoreactive cd (lo) b cells and accumulation of b cells in non-lymphoid organs. inherited human ctla loss-of-function demonstrates a critical quantitative role for ctla in governing t and b lymphocyte homeostasis. ( ) this case highlights the importance of next generation sequencing (ngs) in diagnosing and managing complex presentations with multi-system involvement. case presentation: patient was diagnosed with diffuse large b cell lymphoma at age and treated with rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisone (r-chop) in / . he underwent autologous stem cell transplant with preparative carmustine, etoposide, cytarabine, and melphalan (beam) in / . he subsequently developed recurrent giant condyloma acuminata following transplantation requiring surgical resections, refractory immune thrombocytopenic purpura (itp) requiring aggressive systemic steroids and high dose ivig at least yearly, experiencing hypogammaglobulinemia, recurrent sinopulmonary infections, disseminated herpes zoster, and kaposi sarcoma. in / , he underwent ct/pet, revealing extensive hypermetabolic lymphadenopathy and splenomegaly. bone marrow biopsies were negative for lymphoma, although showed a slightly hypocellular marrow ( - % cellularity), % blasts, and eosinophilia without peripheral eosinophilia. repeated evaluations for hiv, syphilis, histoplasma, cmv, hhv , hhv , bartonella, coxciella, brucella, htlv, toxoplasma were negative. htlv and antibodies had been negative prior to transplantation. tonsillectomy / due to progressive enlargement showed reactive follicular hyperplasia with focal acute tonsillitis without granulomas or viral inclusions. repeated lymphocyte enumeration and proliferation studies were normal. a repeat pet scan / revealed persistent diffuse lymphadenopathy involving the neck, chest, abdomen, and pelvis. left lung biopsy revealed non-caseating granulomas without lymphoma. stains for ebv were negative. repeat pet scan / indicated disease progression prompting a left axillary excisional lymph node biopsy, revealing ebv lymphadenitis with large, reactive follicles with interspersed inflammation and loosely formed granulomas and cd positive b cells within the follicles. ebv blood pcr was negative. afb and fungal stains were negative on all biopsies. in / , his igg was ( - ), iga ( - ), and igm ( - ) with only out of protective serotypes to pneumococcus post-vaccination at . or greater. in / , igg was ( - ). custom ngs panel showed a heterozygous missense variant in ctla c. c>g (p.s r) located in the transmembrane domain. this variant of uncertain significance is suspicious and strongly suggests the diagnosis of ctla -related autoimmune lymphoproliferative syndrome . patient was referred to the national institute of health, where he received a bone marrow transplant. conclusion: primary immunodeficiency diseases comprise a group of highly heterogeneous immune system diseases and around forms of pid have been described. ngs has recently become an increasingly used approach for gene identification and molecular diagnosis of human diseases guiding treatment to patients who may otherwise have poor outcomes. ( ) ( ) submission id# erik newman, md , cullen dutmer, md allergy and immunology fellow/university of colorado and children's hospital colorado assistant professor of pediatrics/section of allergy & immunology, children's hospital colorado, university of colorado school of medicine, aurora, co, usa abstract/case report text introduction: inherited defects of the complement system are rare disorders that can result in unique susceptibility to infections with select bacteria. patients with a deficiency of a complement protein early in the complement pathway (affecting c qrs, c , c , factor h, or factor i) have increased susceptibility to infection with encapsulated bacteria, most notably streptococcus pneumoniae and neisseria species. in contrast, patients with a deficiency of a complement protein at the terminal end of the complement pathway (affecting c , c , c , c alpha/beta/gamma, or c ) almost universally present with severe, recurrent, or disseminated neisseria species infections. most genes encoding complement proteins are found on autosomes, in which specific complement deficiencies result from biallelic mutations. although particular complement deficiencies occur at higher frequencies in certain populations, the prevalence of specific complement deficiencies is unknown in many parts of the world, especially in underdeveloped regions, including sub-saharan africa. herein, we describe a young congolese boy with an atypical presentation of c alpha deficiency. case description: a -month-old congolese boy with consanguineous parents (first cousins) presented with recurrent infections. prior to an evaluation of his immune system, he was hospitalized five times. his infections included episodes of acute otitis media, bacterial pneumonia, and viral pneumonitis. a bronchoscopy revealed diffusely edematous airways and growth of candida albicans, moraxella catarrhalis, and streptococcus pneumoniae in bronchoalveolar lavage cultures. concurrently, his respiratory pcr panel was positive for adenovirus. his initial immune evaluation included assessments of his serum immunoglobulin levels, vaccine titers (tetanus, diphtheria, haemophilus influenzae, and streptococcus pneumoniae), neutrophil oxidative burst, lymphocyte subsets, and ch , in which only his ch was abnormal ( u/ml). his ch remained low on repeat assessment ( u/ml), at which time an ah was pursued and also returned with a low result ( % of normal). a complement system genetic panel identified a homozygous intronic variant in c a (c. - g>a). functional confirmation of the variant revealed that the patient had a significantly decreased c level ( mcg/ml) and absent c function. discussion: we present a case of a young congolese boy with c alpha deficiency and a clinical presentation atypical for defects in terminal complement proteins. our patient presented primarily with recurrent respiratory infections, including streptococcus pneumoniae pneumonia, but without a preceding history of meningococcal disease. while it is well established that patients presenting with terminal complement pathway defects have an increased susceptibility to meningococcal disease, it is less clear if they have increased susceptibility to pneumococcal infections. occurring between exons and , the homozygous intronic variant in c a identified in our patient is predicted to result in abnormal splicing. while the allele frequency of this mutation is relatively high in the african population ( . ), functional confirmation of the variant demonstrated a decreased c level and absent c function that support the pathogenesis of the mutation. the discrepancy between the allele frequency and reported disease cases could be explained in part by varying clinical manifestations seen in c deficiency or underrecognized disease in sub-saharan africa. abstract/case report text rationale: scid is a syndrome characterized by profound t, b, and (in some cases) nk cell defects that is universally fatal unless immune reconstitution is achieved. a total of scid infants have been given allogeneic bone marrow transplantation at duke university medical center without pre-transplantation chemotherapy or post-transplantation graft-versus-host disease (gvhd); % received t cell-depleted haploidentical parental marrow and ( %) are known to be deceased. post-transplantation follow-up ranged from months to years. the aim of this cross-sectional study is to characterize the clinical status of a large cohort of survivors treated at a single medical center. methods: clinical status was assessed by detailed questionnaires delivered by mail or electronically. adult (≥ years old) and pediatric questionnaires were based on patients' age. patients were also contacted by telephone and evaluated at clinic visits. molecular type of scid, demographics, type, date and age at transplant were obtained from a clinical database. results: fifty questionnaires were completed to date from survivors ranging in age from to years. twenty-nine/ were adults ≥ years at the time of the questionnaire. genetic defects were known for all patients-xlinked scid was the cause in about half ( figure ). twenty-three of patients were on immunoglobulin replacement. thirty of reported having received immunizations, and about half of those received live vaccines. fifteen of reported they were taking no regular medications; reported taking prophylactic antibiotics. >we found substantial scholastic achievement, with / adult patients reporting college attendance. two had post graduate education including doctorate level degrees. occupations included physician, nurse, factory worker, musician, teacher, and engineer. one patient had children. twenty-seven / adult patients shared their height and weight and % ( / ) had a healthy bmi (bmi . - . ), while % ( / ) were overweight, and % ( / ) were underweight (< . ) . in pediatric patients, the average age and sex-adjusted bmi was at the th percentile and only had a bmi that was < th percentile. thirty-four/ patients reported seeing an immunologist regularly. in the adult group, % reported no longer seeing an immunologist. the health conditions reported were similar to those common in the general population, and included rashes, warts and mouth ulcers. most reported these were transient, self-resolving issues. thirteen of ( %) reported having adhd, higher than nih reported rates which estimate adhd in . % of adults and % of children). ten of ( %) reported having anxiety, similar to the nih reported prevalence of . % in the general population. / (~ %) reported having no active concerns about their health. conclusions: overall, our findings are consistent with those in the last update done by railey et al, j. peds. : [ ] [ ] [ ] [ ] [ ] [ ] [ ] in this population. patients are doing well with most problems similar to those common in the general population. most have a healthy bmi. adhd had a higher prevalence than in the general population. more than / of scid patients are not seeing an immunologist regularly, and a majority do not have any active concerns. . genetic causes of scid in the patients whose questionnaire data are presented. x-linked scid was the most common, followed by ada and il- r deficient scid clinical fellow/national institute of allergy and infectious diseases (niaid/nih) research nurse/nih-nhgri professor of pediatrics and allergy and immunology/ann & robert h. lurie children's hospital of chicago chief, laboratory of clinical immunology and microbiology/national institute of allergy and infectious diseases, niaid/national institutes of health, nih abstract/case report text rationale: myopathy has been occasionally documented in patients with primary immunodeficiency (pid). however, data on frequency and patient characteristics associated with myopathy are lacking. we performed a descriptive analysis of patients with primary immunodeficiency (pid) in the usidnet having myopathy as a feature of their primary disease. methods: the usidnet registry was queried for the spectrum of myopathic disorders in pid patients that had been entered into the registry as of november , . results: a total of pid patients with myopathy were identified, of which ( . %) were female. median age at onset of symptoms related to pid was years (range . - years, iqr . - years). median age of diagnosis of pid was . years (range . - years, iqr - . years). age at onset of myopathic disorders was not known. twenty-eight ( . %) patients had a diagnosis of common variable immunodeficiency (cvid), ( . %) had agammaglobulinemia, patients each ( . %) were diagnosed with severe combined immunodeficiency, hypogammaglobulinemia, or 'hlh and pigmentary disorders', while patients ( . %)were reported in each category of combined immunodeficiency (cid), autoimmune lymphoproliferative syndrome (alps), and autoinflammatory disease. thirty-five patients ( . %) had a causative gene variant identified attributable to pid. the most common variant identified was btk ( patients) followed by aire, lyst, cybb ( patients each) and pi kcd ( patients). eighteen individual patients had other variants identified ( figure ). patients had cellulitis or skin/ subcutaneous tissue infection, patients had a 'skin or subcutaneous tissue abscess', had pyoderma gangrenosum, and patient with eczema herpeticum. within this cohort of patients, the most common myopathy listed was myositis ( ) followed by 'muscle weakness' ( ), dermatomyositis ( ) , myalgia/s ( ) , myalgia/myositis ( ), myopathy ( ), polymyositis ( ) , steroid-induced myopathy ( ) . no patient had an infectious myositis or muscle abscess listed. eighteen patients ( . %) had a myopathic disorder at the time of diagnosis of their pid. thirtyone patients ( . %) received prednisone, ( . %) received hydrocortisone and ( . %) received dexamethasone. two patients had a diagnosis of adrenal insufficiency. nine patients ( . %) underwent hematopoietic stem cell transplantation. nine patients ( . %) died; median age of death years (range . - . years, iqr . - . years). ). one patient with chronic granulomatous disease had myopathy due to duchenne muscular dystrophy, which was listed as a cause of death. no other myopathic disorders were listed as a cause of death for the other patients. conclusion: myopathy and inflammatory myopathic disorders occur at relatively high frequency in pid, and may be present even at the onset of clinical symptoms. the underlying etiology can be speculated to be multifactorial. further subgroup analysis is warranted to elucidate possible variant-specific or treatment-associated characteristics of myopathy in pid. laboratory studies at years revealed normal igg/iga/igm but markedly elevated serum ige ( , ku/l), anemia (hb . g/dl), thrombocytopenia ( x /l) and lymphopenia ( cells/l), with low t and b cell counts, very low proportion of naïve t cells, skewed repertoire of cd + t cells, undetectable trec levels, and impaired t cell proliferation to mitogens and antigens. there was an elevated percentage of circulating plasmablasts ( . %) and of dysreactive cd low cd low b cells ( . %). at the age of , hsct with reduced intensity conditioning was performed from her phenotypically hla-matched father, with improvement of t and b cell count and function. whole exome sequencing (wes) identified a homozygous missense variant in the mannosidase alpha class b member (man b ) gene (p.asp asn), that segregates with disease in the pedigree. the man b asp residue is evolutionary conserved. the p.asp asn allele has a minor allele frequency of . in gnomad, with no homozygotes. the cadd score for this variant is . , significantly higher than the mutation significance cutoff score ( . ). man b is involved in the lysosomal degradation of glycoproteins and demannosylation of free n-glycans. in particular, man b cleaves man glcnac to generate man glcnac . serum n-glycan profiling revealed elevated man /man and man / man in the patient. n-linked and free glycan profiling by mass spectrometry (ms) showed accumulation of man glcnac , man glcnac and man glcnac glycans in patient fibroblasts as compared to control cells, consistent with defective lysosomal glycoprotein degradation. lentiviral transduction of wild-type man b into patient fibroblasts led to normalization of the n-linked glycan profile, with reduction of man glcnac from . to . times control levels, and of man glcnac from . to . control levels, indicating rescue of the impaired deglycosylation ( figure a ). western-blotting demonstrated defective n-glycosylation of lamp and icam proteins in patient fibroblasts ( figure b) , which were corrected upon lentiviral transduction of wild-type man b ( figure c ). overall, our results indicate that loss of man b enzymatic activity leads to dysregulation of deglycosylation and abnormal mannosylation of glycans. in conclusion, we have demonstrated that man b deficiency accounts for a novel autosomal recessive cdg with prominent features of immune deficiency and immune dysregulation. abstract/case report text heme oxygenase- (hmox ) is a rate-limiting enzyme that catalyzes the degradation of heme to carbon monoxide, ferrous iron, and biliverdin, which becomes bilirubin. these byproducts are implicated in inflammation, cell homeostasis, and antioxidant defense( ). hmox -deficiency is an extremely rare autosomal recessive disorder with a complex presentation of a wide spectrum of symptoms, including hemolytic anemia and hyperinflammation, requiring genetic testing for confirmed diagnosis ( ) . we report the fifth known case of hmox -deficiency ( ) ( ) ( ) , a boy who presented at years of age with aspects of the characteristic phenotype, but also had early onset asplenia, interstitial lung disease, and previously undocumented immune deficiency. patient's presentation was notable for hyperinflammatory exacerbations triggered by viral and bacterial infections as well as vaccinations. episodic flares occurred every few months lasting weeks to months with fevers of - f, hypoxia, leukocytosis above , /mm , hemolytic anemia with negative coombs, thrombocytosis exceeding million/ mm , transaminitis, hemoglobinuria, hyperferritinemia to , ng/ml, and elevated ldh to , iu/l. immune evaluation revealed normal immunoglobulin levels and adequate vaccine titers to both protein and carbohydrate antigens. although class switched populations were normal, b-cell phenotyping showed absent immature and transitional b-cells, low mature memory, and reduced cd + memory b-cells at % (normal > %). mitogen stimulation with phytohemagglutinin and anti-cd were decreased ( . % of control and . % of control, respectively). t-cell phenotyping demonstrated cd population heavily skewed to immaturity with % of cells with naïve phenotype cd ra+cd +ccr +. there were few effector-memory t cells and the cd population was skewed towards immaturity with > % of the cells naïve. liver biopsy was performed secondary to hepatomegaly yielding mild to moderate sinusoidal fibrosis. bone marrow biopsy revealed a normocellular marrow with % blasts, increased megakaryocytes, and extensive hemophagocytosis. natural killer cell function was very low, while soluble il- ra level was normal. further workup for hemoglobinopathies, metabolic defects, congenital disorders of glycosylation, lysosomal storage disorders, wilson's disease, autoimmune hepatitis, inherited and autoimmune hypercoagulability disorders, connective tissue disorders, myositis, and myopathies were all unremarkable. imaging demonstrated asplenia and howell-jolly bodies were present. hemophagocytic lymphohistiocytosis (hlh) genetic testing showed no variants. he was suspected to have systemic juvenile idiopathic arthritis (sojia) with episodes of macrophage activation syndrome. the frequency of his autoinflammatory flares increased such that he was corticosteroid dependent by age , having failed methotrexate, azathioprine, and anakinra. he was started on tocilizumab with laboratory improvements, but his lung disease progressed and became oxygen dependent. lung biopsy confirmed nonspecific interstitial pneumonitis (nsip) with cholesterol granulomas also seen in sojia. ultimately, chronic lung disease led to his death at age . whole exome sequencing yielded a paternal frame shift hmox and maternal splice donor hmox resulting in absence of protein. bone marrow transplantation (bmt) in hmox deficient mice have rectified phagocytotic defects and thereby their autoinflammatory phenotype, but no human reports for bmt treatment of hmox -deficiency has been described. here we describe a phenotype expansion for hmox deficiency to include not only asplenia and hepatomegaly, but also interstitial lung disease with cholesterol granulomas and inflammatory flares. abstract/case report text introduction: mutations in the gene encoding signal transducer and activator of transcription (stat ) cause autosomal dominant hyperimmunoglobulin e syndrome (ad-hies) characterized by recurrent skin and sinopulmonary infections, atopic dermatitis, and elevated serum immunoglobulin e (ige) levels. treatment is largely aimed at controlling symptoms and preventing infections with no standard of care. there is a paucity of literature describing the utilization of biologic therapies in the ad-hies patient population. we present patients from one family with ad-hies successfully treated with monoclonal antibody therapies targeted at il- , il- and il- . case descriptions: patient : -year-old female with stat lof c. c>t (p.arg trp) with a history of atopic dermatitis and asthma requiring - steroid courses per year with frequent school absences. she developed a severe pruritic rash covering her upper body months ago that failed to respond to antihistamines and topical antibiotics prescribed by her primary care provider. given her poorly controlled asthma and our concern for a follicular type morphologic variant of atopic dermatitis, dupilumab was initiated. her scoring atopic dermatitis (scorad) prior to initiation of biologic therapy was . and improved to . following doses ( weeks) of dupilumab with clear dramatic improvement in her skin and quality of life ( figure ). she also reports decreased asthma severity with no steroid courses, reduced albuterol usage, and significant decline in school absences since initiation of dupilumab. patient : -year-old female with stat lof c. c>t (p.arg trp) who is the sister of patient . she has a history of severe asthma requiring frequent emergency department visits, hospitalizations, and - steroid courses per year despite therapy with high-dose fluticasone-salmeterol. spirometry prior to april demonstrated an obstructive pattern with an fev ranging from - %. she was initiated on mepolizumab in april . subsequent spirometry demonstrates an fev average of % with a range of - %. she had one hospitalization in early but otherwise no hospitalizations for asthma since initiation of biologic therapy. patient : -year-old female with stat lof c. c>t (p.arg trp) who is the paternal first cousin of patients & . she has a history of severe atopic dermatitis with associated pruritus and picking behaviors, poorly controlled despite daily triamcinolone application. she previously failed ultraviolet therapy and crisaborole. she also has a history of severe asthma requiring - steroid courses per year despite high-dose fluticasone-salmeterol. dupilumab was started in may . scorad prior to initiation monoclonal antibody therapy was . and declined to . following weeks ( doses) of dupilumab therapy with marked improvement in skin appearance and pruritus ( figure ). discussion: we present three cases of ad-hies caused by stat loss-of-function mutations treated successfully with monoclonal antibody therapies targeted at il- or il- and il- . to the best of our knowledge, there is no published data describing the use of these biologic agents in the treatment of ad-hies. future studies are needed to clarify the role of these cytokines in the pathogenesis of ad-hies and to elucidate clinical indications for biologic therapy in this patient population. informed consent was obtained from all individual participants included in the study. abstract/case report text ctla- is a potent inhibitor of t cell proliferation that competes with costimulatory receptor cd for its ligands cd and cd expressed on antigen presenting cells. heterozygous loss-offunction mutations in ctla- have been identified in patients with lymphocytic infiltration of multiple nonlymphoid organs (lo et al). the patient is a -year-old jordanian male born at term to nonconsanguineous parents, hospitalized at mo for lll pneumonia, and at mo and at mo he was evaluated in the ed and diagnosed with non rsv-bronchiolitis with lll infiltrate thought to be secondary to atelectasis. at yo he developed lll pneumonia and respiratory failure requiring picu admission. he was treated with ceftriaxone. after discharge, he had weeks of intermittent fever, progressive fatigue, productive cough, and ftt. he received courses of cefdinir, clindamycin, and tmp-smx without improvement. chest ct revealed left lung consolidation, lll bronchiectasis, lul tree in bud opacities, and hilar lymphadenopathy. bronchoscopy with bal revealed no bacterial growth and no acid-fast bacilli. srrna ngs was positive for h. influenzae. lung biopsy demonstrated acute and chronic bronchiolitis with bronchiolitis obliterans and intraluminal polyps, with lymphocytic infiltration involving the bronchi and bronchioles. the lung parenchyma showed airspace filling with foamy macrophages and chronic interstitial inflammation. acid fast and fungal stains were negative. he was treated with systemic steroids for bronchiolitis obliterans with noted improvement. the severity of lung disease at such an early age prompted an immune evaluation. sweat test, anca, anti-pr and hiv were negative. total immunoglobulins were normal for age, and titers to s. pneumoniae, diphtheria and tetanus were protective. lymphocyte enumeration revealed elevated t and nk cell numbers for age. lymphocyte proliferation to pha, pwm, candida and tetanus were normal. dihydrorhodamine assay was normal. b cell phenotyping was normal. there was normal expression of cd , hla-dr and cd on activated t cells; of note the patient was on systemic steroids when tested. invitae gene pidd panel revealed a variant in ctla : c. g>a (p.gly glu) that has been shown to be pathogenic in one patient (schawb et al). flow cytometry showed normal frequency of t follicular helper cells and t regulatory cells compared with controls, however ctla- expression by t regulatory cells was lower than control. due to the severe and progressive nature of the patient's lung disease, therapy with x weekly azithromycin and abatacept mg sq weekly was initiated. we report a case of ctla- haploinsufficiency presenting with recurrent pneumonia and bronchiolitis obliterans in a -year-old child. based on patient registry data, our patient appears to be the youngest child diagnosed with ctla- haploinsufficiency reported in the literature to date (schawb et al). notably, our patient l a c k s o t h e r f e a t u r e s c o m m o n l y d e s c r i b e d i n c t l a - haploinsufficiency, including autoimmune cytopenias, gastrointestinal disease, lymphoproliferation, and hypogammaglobulinemia. this case illustrates the importance of consideration of this diagnosis in young children with severe lung disease without other evidence of immune dysregulation. our hope is that prompt recognition and early treatment administration will prevent disease progression and further decrease in pulmonary function. splenectomy, he had an episode of pneumococcal meningitis at age , and sepsis of unknown origin at age . over the past several years he has developed chronic tinea corporis, onychomycosis, and otitis externa infections despite numerous antimicrobial regimens. at age , the patient developed urinary retention, walking, and balance difficulties. he was found to have diffuse white matter changes on mri, elevated wbc, and positive oligoclonal bands. initially, he was diagnosed as progressive ms treated with steroids with partial improvement. csf microbiology studies including afb stains, bacterial, fungal, mycobacterial cultures, cryptococcal antigen, vdrl, t. pallidum particle agglutination (tppa), as well as, pcr for cmv, ebv, vzv, enterovirus, hsv - , jc virus and t. pallidum were all negative. peripheral blood studies included mycobacterial blood culture, pcr for cmv, ebv, hhv- , in addition to serology for cryptococcal antigen, and coccidioides species, all of which were negative. additional neurological complications include granulomatous uveitis and oscillopsia, which he developed around age . immune evaluation performed at age revealed low igg and igm, and the patient was started on grams of monthly ivig. cbc with differential was notable for normal monocyte count and thrombocytopenia, mild neutropenia (table ). immunophenotyping revealed absent b cells and nk cells, while the cd t cells were elevated. cd t cells were normal (table ) . at age , whole-exome sequencing identified a heterozygous missense mutation in gata c. c>t, p.(arg trp). after the diagnosis of gata haploinsufficiency, he was found to have myelodysplastic syndrome with multilineage dysplasia (mds-mld) on bone marrow biopsy. he is currently awaiting bone marrow transplant discussion: we present a -year-old male with cytopenias, splenomegaly, leukoencephalomyelopathy, granulomatous uveitis, and recurrent fungal infections found to have a pathogenic heterozygous missense mutation in gata . leukoencephalomyelopathy in gata haploinsufficiency has been associated with jc virus and ebv infection. our patient did not have any evidence of a chronic csf infection. to our knowledge, myelopathies have not been reported with gata c. c>t, p.(arg trp). this case highlights the variable nature of presentation in gata haploinsufficiency, and the need for clinical awareness of this entity in order to facilitate early diagnosis and appropriate therapy. immunoglobulins* white blood cells . x * /l (n: - ) magnetic resonance imaging (mri) of the brain and spine showed numerous enhancing parenchymal nodules (figure one). brain or spinal biopsy was requested, but not recommended by our neurosurgery service. lumbar puncture evaluation was performed. cerebral spinal fluid showed no bacterial or fungal elements. both quantiferon gold for tuberculosis and three consecutive sputum cultures for acid fast bacilli were negative. he continued his sirolimus and the intravenous immunoglobulin replacement was increased to two grams/kg. the patient was cleared from respiratory isolation and discharged after two weeks in our facility with mild improvement in his neurologic status. within five days he was sent to a nationally renowned hospital. at this facility, extensive evaluation for his neurologic deficits were performed including culture and pcr for bacteria, virus, mycobacteria from csf bone marrow, lymph node, blood, and induced sputum. these were noncontributory. he was given high dose corticosteroids for two days. one of our facility's sputum cultures was reported with acid fast bacilli. but sputum mycobacterium tuberculosis pcr was negative. repeat mri scans of the brain and spine showed improvement (figure ), so no brain biopsy was performed. he was sent back to our facility with the recommendation to start a targeted pi kinase inhibitor on compassionate grounds as he was not eligible for the clinical trial because his weight was less than kg. but pretreatment abdominal ct revealed multiple low-density lesions scattered throughout the liver (figure ) not previously seen on prior noncontrast ct four months prior. discussion: although this gain in function mutation of the pi kδ signaling pathway disorder has been well characterized, this is a rare report of a patient with pasli immunodeficiency with central nervous system and later liver lesions pet imaging showed subcarinal, mediastinal, retroperitoneal lymphadenopathy, splenic enlargement to cm and bilateral lung nodules ( figure ). excisional biopsies of left axillary and left lower lobe of lung were performed and showed low-grade b-cell lymphoma (mucosal) and underlying lymphoproliferative disease. invitae alps and cvid panels ( genes) revealed a heterozygous variant of unknown significance in exon of fas (c. a>g(p.asp gly) unlike most fas mutations causing alps, this mutation is in the extracellular region rather than the death domain . the c. a>g variant has been reported in a single patient with alps phenotype, affecting fas protein function by inhibiting binding to fas ligand (fas-l), reducing fas-l induced apoptosis . this fas c. a>g mutation was found in alps affected brother and was absent in the unaffected father. the patient's mother passed away prior to testing. since diagnosis, the patient's malt lymphoma has been treated with rituximab weekly for the first month and then monthly. he continues to receive monthly ivig for hypogammaglobulinemia. after two years, ct demonstrates a significant decrease in pulmonary nodules and splenomegaly ( figure ). the patient's forced vital capacity (fvc) improved from . l ( % predicted) to . l ( % predicted). conclusion: we report the first case of malt lymphoma seen in a patient with alps. it is unknown whether the unique fas c. a>g mutation in the non-death domain contributes to malt lymphoma progression. we propose malt lymphoma is a malignant transformation of chronic inflammation that has the potential to occur in patients with alps. in the future, improved knowledge of mechanistic pathways of inflammation in lymphoma development and progression is important in the optimal management of alps. abstract/case report text background wiskott-aldrich syndrome (was) is a rare x-linked disorder characterized by combined immunodeficiency, eczema, microthrombocytopenia, infections, autoimmunity and increased risk of hematological malignancies. gene therapy (gt) using autologous cd + cells is an emerging alternative treatment with possible advantages over standard allogeneic hematopoietic stem cell transplant. we report the outcomes of a phase i/ii clinical trial in which was patients underwent gt using a self-inactivating lentiviral (sin-lv) vector expressing the human was cdna under the control of a . kb fragment of the human was promoter. subjects and methods: five patients with severe was (clinical score - ) were enrolled (table ) . cd + cells were transduced ex-vivo and reinfused after conditioning with busulfan and fludarabine. two subjects (p , p ) had autoimmunity pre-gt, manifested as skin vasculitis and autoimmune cytopenias. results: all subjects were alive at median follow-up of . (range . - . ) years. multi-lineage vector gene marking was sustained over time. all had clinical improvement of eczema, infections and bleeding diathesis. was protein (wasp) expression was increased over baseline but remained below normal levels. proliferation of t cells in response to anti-cd improved post-gt. humoral immune deficiency improved, with normalization of igm, and independence from ig replacement and vaccine responses in those tested. platelet levels increased to > x cells/ul in only the two subjects with a vcn ≥ in transduced stem cells. podosome formation in monocyte-derived dendritic cells was near absent pre-gt and improved in all subjects post-gt, but only reached healthy control levels in the subjects with highest vcn. in contrast to other trials using this sin-lv, two patients (p and p ) had flares of autoimmunity post-gt, offering the opportunity to study the poorly understood mechanistic features of immune dysregulation in this disease. selfreactive vh - -expressing b cells and cd lo b cells remained elevated in most patients. however, despite wasp expression in foxp + tregs, those with autoimmunity had poor numerical recovery of t cells and tregs at the time of clinical symptoms ( fig a) . in addition, il- producing regulatory b cells (bregs) were highly deficient pre-gt, recovered in subjects who did not experience autoimmunity, but failed to recover in p and p ( fig b) . moreover, transitional b cells, which are enriched in bregs and are potent inducers of treg populations, also recovered poorly in those two subjects ( fig c) . there have been neither severe gt-related adverse events nor abnormal clonal expansion in transgene-marked cells to date. conclusion in summary, our data confirm and extend the safety and efficacy of gt in correcting disease manifestations associated with was, with the longest overall follow-up reported so far in studies using sin-lv. in addition, our findings suggest that higher vcn is needed in order to correct myeloid compartments such as platelets and monocytes. finally, we report the novel finding of the restoration of bregs and suggest that recovery of this compartment, along with tregs, is protective against development of autoimmunity post-gt. overall, these data suggest a mechanism for breakdown of immune tolerance in was with important therapeutic implications and prognostic value. this is an -year-old hispanic female who initially presented with failure to thrive, recurrent fevers and intermittent cough with episodes of perioral cyanosis. symptoms started at age months and were attributed to recurrent viral and bacterial infections. at months old, she was hospitalized with fever and hypoxemia (o saturations %). cxr showed prominent interstitial lung markings and she was diagnosed with pneumonia. ct scan confirmed cxr findings and ruled out anatomical anomaly. she was lost to follow up for years, and re-presented with worsening respiratory status. a repeat ct scan demonstrated worsening interstitial thickening. immune workup, including quantitative immunoglobulins, ch , lymphocyte subsets and vaccine response titers (pneumococcal and tetanus), was unremarkable, except for elevated igg levels. genetic testing for surfactant dysfunction mutations was negative. thoracoscopic lung biopsy revealed interstitial fibrosis, pas-positive granular alveolar proteinosis, type ii cell hyperplasia, and lymphoid follicles. at age , she was admitted for a pericardial effusion. rheumatology was consulted for evaluation frequent fevers and persistently elevated inflammatory markers, with concern that the pericarditis was autoinflammatory. she had an elevated ana (> : homogeneous pattern), il- ( . pg/ml), and igg ( mg/dl) at that time. she had an atypical anca pattern with positive myeloperoxidase antibodies. anti dsdna, smith and scl were negative. she was treated with steroids and hydroxychloroquine with some improvement in her oxygen requirement. one year later she had an additional episode of pericarditis, treated with colchicine. a few months later, she was admitted with newonset gross hematuria and elevated serum creatinine (to mg/ dl). kidney biopsy showed anca vasculitis with glomerulonephritis ( % crescents, no scarring or fibrosis). she provisionally received a diagnosis of microscopic polyangiitis, with lung and kidney involvement. she did not have peripheral vasculopathy. she was started on cyclophosphamide, rituximab, and iv steroid pulses. cyclophosphamide was discontinued due to recurrent episodes of posterior reversible encephalopathy syndrome (pres) after infusion. her igg level decreased as she developed nephrotic range proteinuria. a primary immunodeficiency genetic panel was sent to evaluate for monogenic immune dysregulation syndromes and revealed a tmem gene mutation (c. g>a) which has previously been reported in other subjects with savi (stingassociated vasculopathy of infancy syndrome). sting is a cytosolic dna sensor that leads to type i interferon production upon stimulation. this gain-of-function mutation was confirmed by measuring interferon signature gene expression at the nih (fig ) , and her diagnosis was revised accordingly. the patient was started on a jak-inhibitor (tofacitinib) to block interferon signaling. unfortunately, the patient is now deceased, due to overwhelming infection and multi-organ system failure. conclusion: genetic testing can be crucial in aiding the diagnosis of complex patients with immune dysregulation and can provide an opportunity for targeted therapy, which should be employed as soon as able to stop disease progression. abstract/case report text background: activated phosphoinositide -kinase δ syndrome (apds- ) was first described in as a monogenetic immune dysregulation syndrome with a variable phenotype. increased sinopulmonary and herpesvirus infections are well described, but fungal infections such as candidiasis have been rare. to date, disseminated histoplasmosis has not been described. history: a yo caucasian male who was previously diagnosed with common variable immunodeficiency (cvid) in late childhood due to recurrent sinopulmonary infections presented with recurrent fever, pancytopenia, severe splenomegaly, and lymphadenopathy. urine histoplasmosis antigen and beta-d-glucan were elevated. a bone marrow biopsy demonstrated granulomatous inflammation. transbronchial biopsy of a subcarinal lymph node was consistent with granulomatous disease. this led to a diagnosis of disseminated histoplasmosis. he was treated with amphotericin b and then months of itraconazole, with improvement of his symptoms. he was admitted to the hospital about years later when he presented with fatigue, fever, chills, dark urine, and scleral icterus. he was found to have an acute worsening of chronic anemia with a hemoglobin of . g/dl. due to elevated ldh, presence of schistocytes on peripheral smear, and undetectable haptoglobin, he was diagnosed with autoimmune hemolytic anemia, despite a negative direct coombs. a bone marrow biopsy specimen was hypercellular with marked erythroid predominance, with normal flow cytometry and no blasts identified. infectious workup was negative. ct chest during the workup revealed new right hilar and mediastinal lymphadenopathy, in addition to calcified right hilar and subcarinal lymph nodes, bronchiectasis, and stable hepatosplenomegaly. transbronchial biopsy of lymph nodes showed benign lymph nodes with calcified necrotizing granulomata and presence of non-viable fungal species, presumably "old" histoplasmosis. family history: family history was significant for mom dying at years-old from undefined cns infection. immune labs: · panlymphocytopenia: absolute lymphocyte count of /ul, cd + t cells /ul, cd + /ul, cd + /ul, cd + /ul, cd +cd + /ul. cd +/cd + ratio . · decreased class-switched memory b cells and plasmablasts · elevated t central memory cells and activated (hla-dr+) cd + and cd + t cells · hemoglobin . g/dl, platelets , /ul, anc ranging from /ul to /ul · iga mg/dl, igm mg/dl; reportedly had low igg prior to initiating ivig in childhood · ebv pcr and cmv pcr negative genetics: · pik cd (c. g>a), consistent with diagnosis of autosomal dominant apds- . discussion: gain-of-function variants leading to increased pi kδ activity have been shown to cause both b and t cell dysfunction, leading to impaired immunologic responses to bacterial and viral infections. recurrent sinopulmonary infections and herpesvirus infections are commonly seen and while mucocutaneous candidiasis has been reported in cohorts of patients with pik cd, other fungal infections are not common. severe disseminated histoplasmosis infections have been described in primary immunodeficiencies characterized by signaling defects in the il- /ifn-γ pathway, stat deficiency, cd l deficiency, gata deficiency and in stat gain-of-function mutations. to our knowledge, disseminated histoplasmosis has not been previously reported in patients with pik cd immunodeficiency. abstract/case report text the reported case represents the first case of nbas disease detected by newborn screening program for primary immunodeficiency, based on krec assay. the patient came to our attention due to the complete absence of krecs and normal trecs on dbs (dried blood spot) while hospitalized for low weight at birth ( , g), intolerance for enteral feeding, hepatosplenomegaly, slightly elevated liver transaminase, head and face eczematous dermatitis. during the st month, he also presented klebsiella pneumoniae urinary tract infection and methicillin-resistant staphylococcus aureus sepsis. peculiar phenotypic features including triangular face, proptosis, flat philtrum, mild retrognatia, hirsutism, loose and slightly wrinkled skin, and apparent reduction of subcutaneous fat were noticed at birth. complete blood count showed lymphocytopenia, marked hypereosinophilia. serum immunoglobulin g (igg) were markedly decreased, iga and igm were undetectable. extended immune-phenotyping showed complete absence of cd + cells, low count of cd + lymphocytes, and reduced natural killer (nk) levels. at month of age a colonoscopy was carried out for persistent diarrhea and reduced tolerance to enteral feeding. the histological examination of mucosal intestinal biopsies showed signs compatible with autoimmune enteropathy. for this reason immunosuppressive therapy with rapamycin was started without consistent clinical amelioration. many cvc-sepsis occurred in the last months, associated with persistent gastrointestinal symptoms and severe growth restriction. despite the absence of experience data in literature for nbas syndrome, we retain that hsct represents the only resolutive therapy for him. abstract/case report text case: a -year-old female with asthma and allergies presented to immunology clinic with a history of chronic fatigue and sinusitis. fatigue occurred daily every - weeks and was described as not feeling rested even after hours of sleep. chronic sinusitis required - prolonged antibiotic courses per year. nasal cultures grew methicillin-sensitive and -resistant staphylococcus aureus and haemophilus influenzae type b (hib). three separate sinus surgeries over the prior few years reduced her sinus symptoms. other infectious history was significant for recurrent urinary tract infections with e. coli and klebsiella, recurrent otitis media as a child, and a diagnosis of transient hypogammaglobulinemia of infancy that resolved at years of age. review of systems revealed axillary lymphadenopathy for - days twice per year not related to infection. she had longstanding eczema that responded to topical tacrolimus, multiple environmental allergies, and recently diagnosed asthma that improved with inhaled budesonide/formoterol. as a teenager she received allergy immunotherapy for a few years but stopped due to frequent adverse reactions. family history revealed that father died from cancer. physical exam was unremarkable. laboratory evaluation demonstrated normal igg mg/dl, igm mg/dl, iga mg/dl, elevated ige mg/dl, normal t and nk cell enumeration, mildly low total b cells ( cells/mcl, . %), and normal b cell subsets (cd +igm+cd - %, cd +igm+cd + %, cd +igm-cd + %). tetanus antibody titer was protective, but hib antibody titer was undetectable at < . mcg/ml with marginal response after vaccination ( . mcg/ml). pneumococcal serotype specific igg levels (mayo) were mostly undetectable with of serotypes protective at baseline and only protective post vaccination with pneumovax . cd / cd blastogenesis was poor. due to poor antibody response and continued sinus infections she was started on igg replacement. her fatigue and sinus symptoms improved moderately but she continued to require antibiotics and sinus ct scans continued to demonstrate significant disease. a focused exome sequencing panel was pursued and a novel heterozygous card variant was found (c. g>t, p.r l). discussion: the card /bcl /malt (cbm) complex is a critical signaling adapter that facilitates several downstream immune responses predominately through nf-kb. mutations in several different domains of card result in a clinical entity collectively referred to as card -associated atopy with dominant interference of nf-kb signaling (cadins). cadins is associated with a broad range of clinical manifestations but most have marked atopy with infections, poor t cell proliferation, and varying levels of poor antibody response. both our variant (p.r l) and a previously reported pathogenic variant in the same amino acid (p.r g) involve a change from a charged arginine to a non-polar amino acid in the critical bcl / card binding interface. iκbα degradation was not present in b cells from our patient ( figure ) confirming the functional defect in nf-kb signaling. thus, we present a novel variant that fits cadins both clinically and genetically. clinicians should be aware of cadins when patients present with recurrent infections in the setting of significant allergic disease. i b degradation assay. stimulation: μl of whole blood was stimulated in a ml facs tube with ng/ml phorbol -myristate -acetate (pma; sigma, cat# p ) at °c for , , or min, at which point ml of pre-warmed x lyse/fix buffer (bd, cat# ) was added. cells were fixed for min at °c, centrifuged and washed twice with facs buffer (pbs supplemented with % fbs and mm edta). staining and permeabilization: fc receptors were blocked for min at rt (human trustain fcx; biolegend), followed by a min stain on ice with anti-cd af (clone rpa-t ; biolegend), and anti-cd bv (clone hib ; biolegend). cells were washed with facs buffer and permeabilized for min on ice with ml phosflow perm buffer ii (bd biosciences, cat# ) that had been precooled to - °c. after permeabilization, two ml facs buffer was added and the samples were centrifuged. after three additional washes, the cells were stained with anti-iкbα pe (clone /ikba/mad- ; bd biosciences) for min at rt. samples were washed three times and data were collected on a cytek dxp flow cytometer. data were analyzed with flowjo software. abstract/case report text introduction: wiskott-aldrich syndrome (was) is a rare, but well-defined x-linked disorder. loss-of-function mutations in the was gene result in classic was and x-linked thrombocytopenia (xlt), while gainof-function mutations lead to x-linked neutropenia (xln). classic was phenotypic features include recurrent infections, microthrombocytopenia and eczema along with increased susceptibility to autoimmune disorders and malignancy. most males with classic was are diagnosed in early childhood and early death can result from its various clinical manifestations. case: we present a -year-old male who was referred to immunology for hypogammaglobulinemia. as an infant he had moderate eczema, and at the age of two was diagnosed with immune thrombocytopenia (itp) with baseline platelets of - x ^ /l. infectious history was notable for one episode of pneumosepsis and recurrent otitis media, influenza, and herpes labialis infections. around the age of , he was diagnosed with common variable immunodeficiency (cvid) based on the finding of low immunoglobulins. he developed diffuse large b cell lymphoma at age , and was treated with cyclophosphamide, doxorubicin, vincristine, prednisone and rituximab (chop-r). at age he developed abdominal pain with bloody stools. investigations confirmed an endoscopic and pathologic diagnosis of ulcerative colitis. due to the severity of his disease, he has required maintenance therapy with vedolizumab. he was again noted to have hypogammaglobulinemia at which point he was referred to immunology at our centre. there was no significant family history of immunodeficiency, malignancy or autoimmunity. blood work was notable for normal white blood cell and lymphocyte counts, platelets of x ^ g/l, low igg at . g/l ( . - . g/l) with normal iga, igm and ige. lymphocyte subsets including t, b and nk cells were within the normal range. genetic testing was performed and he was found to have a known pathogenic mutation in the was gene (c. g>a, p.asp asn) which has been previously reported in association with was and xlt. he has since been placed on immunoglobulin replacement and has been referred for consideration of hematopoietic stem cell transplantation. discussion: was is a rare syndrome that can have a similar phenotype to other immunodeficiency disorders including cvid, omenn syndrome and ipex (immune dysregulation, polyendocrinopathy, x-linked). individuals with cvid present with hypogammaglobulinemia and recurrent infections, and these individuals also have an increased susceptibility to autoimmune disorders, gastrointestinal disease and malignancies, especially lymphoma. although eczema is a common disorder, its presence in addition to features of early onset thrombocytopenia, immunodeficiency, autoimmunity and/or malignancy in male patients should heighten the suspicion for was. it is important to make the diagnosis of was as hematopoietic cell transplantation and gene therapy are potentially curative treatment options. abstract/case report text secondary immune deficiencies (sid) are caused by varied mechanisms and are common in patients with hematological malignancies such as chronic lymphocytic leukemia (cll) and multiple myeloma (mm). in this setting, both the disease and its treatment (such as b cell ablation therapy) contribute to the development of secondary antibody deficiency. infections remain a major cause of morbidity and mortality in cll and mm patients. this underscores the need for early recognition and stratification of risks in order to guide appropriate treatment, including immunoglobulin replacement therapy (igrt). new guidelines for the use of human normal immune globulins in sid patients were implemented by the european medicines agency (ema) in . despite these new guidelines, significant variations remain across european countries in the assessment and approaches aiming to achieve reduction in infection burden, including different strategies for initiation, dosing and discontinuation of igrt. the same is true for north america where igrt is widely used off-label to prevent infections in patients with sid due to hematological disease or other reasons. in order to address this variability, a task force comprising both immunologists and hemato-oncologists drafted statements aiming to test for consensus. statements were related to six major areas: definition of infections, measuring igg levels, initiating igrt, igrt dosing, scig usage and discontinuing igrt. this was followed by an international delphi consensus exercise in three rounds which aimed to develop recommendations on how to diagnose, treat and follow-up patients with antibody deficiency associated with hematological malignancies. the first delphi round consisted in testing the statements with a panel of sid specialists and subsequently their comments were used by the task force to refine the statements. in the second delphi round, the refined statements were presented via phone interviews to the same panel to assess their level of agreement with each statement (ranging from "i totally disagree" to "i totally agree"). consensus was considered to be reached per statement if % of the experts agreed with each statement overall. the cut-off for overall agreement was "i somewhat agree". if the expert chose level or less the reasons underpinning his/her choice were discussed. consensus was achieved for all statements on level ("i somewhat agree"). only statements did not achieve consensus on level "i mostly agree". in delphi round , panelists who had not "mostly agreed" with these five statements were given the opportunity to reconsider their assessment based on the feedback from other panelists, which was shared with them. the panelists then chose to maintain or refine their assessment. analysis of the full results on the six key areas identified by the task force will be presented at the conference to offer recommendations and help guide the management of sid in patients with hematological malignancies. abstract/case report text introduction: mast cells (mcs) are hematopoietic-derived immune cells, whose precursors migrate within tissues reaching maturation and differentiation. masitinib, a selective tyrosine kinase inhibitor, is efficient in controlling the survival, differentiation, and degranulation of mcs. aim: to optimize mast cell-differentiation from human bone marrow (bm) hematopoietic stem cells, and to find best cell culture conditions for proliferation, differentiation, and maintenance of mcs, which is important when studying particularly mcs' response to cytotoxic compounds. material-methods: to produce mcs in vitro, the first method (m ) we used was a modified semi-solid culture method ( ). briefly; human bm mononuclear cells (mncs) were obtained with ficoll gradient from bm sample of a patient with idiopathic thrombocytopenic purpura. colonyforming unit (cfu)-mast was developed from mncs in methylcellulose medium supplemented with scf ( ng/ml) + il- ( ng/ml), and il- ( ng/ml; only first week). - weeks later mast cell colonies were transferred into suspension cultures, in which mcs matured and multiplied up to - weeks and were used in experiments till th week of culture. on the other hand, in our second method (m ); mncs were separated by ficoll, seeded in well-plates with imdm containing fbs %, pen/ strep, and a little amount of methylcellulose, and incubated at o c, %co . cultures were then supplemented with imdm (fbs %) + scf ( ng/ml) + il- ( ng/ml) on day ; and imdm (fbs %) + scf ( ng/ml) + il- ( ng/ml) + il- ( ng/ml) on day . beginning on day till the end, imdm (fbs %) + scf ( ng/ml) + il- ( ng/ml) were added to cultures. for both methods, morphological assessment of colonies/cells were evaluated under an inverted microscope (figure and ). verification of mcs was performed by immunoflorescence staining for anti-tryptase andchymase antibodies, and by toluidine blue staining. macrophages were verified by anti-cd- immunoflorescence staining. mcs were exposed to masitinib or dmso for the evaluation of dose-related effects of masitinib, and cytotoxicity was evaluated by mtt assay. results: in m , culture conditions were easier to handle compared to m . in m , high amounts of mcs in immature and pre-mature forms were appeared as early as - days, and peak levels of proliferation rate was around - weeks of culture, which was about weeks earlier than m . culture could be maintained till weeks in both methods. although mcs are non-adherent cells, in liquid method adherent bm cells such as fibroblasts, endothelial cells and mesenchymal stem cells have adhered to the plate and grown up, providing an attachment site for mcs and serving as a natural bm nest, mimicking in-vivo environment, for mcs to grow and proliferate ( figure ). attachment of mcs has provided medium exchange available without changing culture dishes. when mcs were exposed to masitinib ( . , , and μm/μl), approximate survival rates were %, %, %, respectively. discussion: in our liquid medium method, the adherent bm cells not only provided a natural nest supporting mc development and differentiation, they also served as an attachment site for mcs. as the cells slightly adhered, when trypsinized shortly, they easily detached and used for experiments. and we also report for the first time that adding a little amount of methylcellulose to the liquid medium provides ease of aggregation of cfus, and easy development of mcs. we suggest that our liquid culture may be superior to semi-solid method, that it is faster and easier to handle. in studies subjects crossed over to subcutaneous (sc) igiv-c %, and in the third study crossover was to immune globulin sc (human), % caprylate/chromatography purified (igsc %). a total of pi patients from these studies were included in the poppk analysis and serum igg concentrations were included in the final pk analysis. the pk of igg following iv and sc administration was adequately described by a two-compartment model with first-order elimination from the central compartment. administration of igiv was modeled as an infusion directly into the central compartment. absorption of exogenous igg from the depot site of sc infusions into the central compartment was modeled as a first-order process with an absorption rate constant (ka). the full model was constructed by incorporation (forward selection process) of covariates of interest into the model. after completion of the covariate model development, the final model showed that igg pk was not influenced by (a) the igsc formulation used in the different studies ( % vs. %), (b) gender, and (c) age (pediatric vs. adult). body weight was identified as a significant covariate having an effect on clearance and volume of distribution. based on the final pk results, serum clearance of igg for the reference population was estimated to be . l/day. the volume of distribution of the central and peripheral compartments accounted for . l and . l, respectively. the intercompartmental clearance was . l/day, and the absorption constant from the depot (ka) was . day- . the absolute bioavailability of igg after sc administration was calculated as . %. the developed method was used to evaluate alternative dosing intervals following sc administration. the equivalent of a weekly igsc maintenance dose administered , , , , or times per w e e k , o r b i w e e k l y p r o d u c e d o v e r l a p p i n g s t e a d y -s t a t e concentration-time profiles and similar area under the concentration versus time curve (auc), maximum concentration (cmax), and minimum concentration (cmin) values. the results of the evaluation and simulations for igg exposure following a switch from igiv-c % dosing (every -or -weeks) to sc dosing further suggest that a range of dose-adjustment factors (daf), from : to : . would be sufficient to provide clinically effective trough igg concentrations throughout the course of treatment at various treatment frequencies. current us product labeling for igsc % specifies a daf of : . for transitioning immune globulin dosing from iv to sc, and specifies igsc % dosing frequencies of weekly or more frequently ( - times per week). in this poppk analysis all sc dosing regimens evaluated theoretically would provide viable alternative administration options for maintaining adequate immunoprotection in pi patients with dosing flexibility over a range of regimens. however, in % or more, no causative gene can be found going down to undefined inflammatory syndromes (uis). anti-il drugs (ail d) revolutionized some il- mediated diseases, such as traps, caps, hyper-igd/mkd and fmf. nevertheless, treatment response among disorders are not the same as well as no specific study was designed for uis. papa et al, , recently suggested the use of anakinra for the treatment of uis, especially those refractory/intolerant to colchicine with severe or very symptomatic phenotype. this paper aims to retrospectively report for the first time the experience with canakinumab in monogenic and multifactorial disorders in a single, private center in brazil. patient and methods: patients's records that received canakinumab from january to december at clinica croce, ima-brazil, were revised. demographic and clinical data were extracted and descriptively described. all statistical analysis are presented as: average (minimal; maximum; standard deviation). results: a total of patients with autoinflammatory diseases were enrolled and % (n= ) are female. of them, % (n= ) patients had a monogenic disease: % (n= ) caps, % (n= fmf), % (n= ) mkd, % (n= ) homozygous nlcr and % (n= ) pami syndrome. multifactorial disorders were % (n= ) patients : % (n= ) recurrent idiopatic pericarditis, % (n= ) schnitizler syndrome and % (n= ) uis. the average age of the first symptoms was , years ( ; ; , ) and the average age of diagnosis was , years ( ; ; , ) while the aveage of diagnosis delay was , years ( ; ; , ). all patients had used, prior to anti-il , corticosteroids with % prevalence of cushing syndrome and % (n= ) tried at least one steroid sparing agent without clinical success due to: intolerance or non-effective disease control or side effects. in the fmf group (n= ) % tried colchicine prior to canakinumab and this drug was not effective to % because of amyloidosis status and in % colchicine-induced hepatitis was observed. canakinumab was effective for disease control in % (n= ) considering: control of clinical manifestations, amyloidosis reversion and normalization of acute reactants markers. the only side effect observed during the follow up were acute flu-like symptoms and psicomotor agitation ( , % , n = ). the average time of follow up is of , months ( ; ; , ). canakinumab could be discontinued in just one patient with uis. conclusions: this is the first report of canakinumab use for autoinflammatory disorders in brazil. canakinumab is an effective and safe drug for monogenic and multifactorial disorders control. no serious adverse effect could be observed in the years maximum follow up of this drug. neither, no specific infectious disease more prevalent in south america, such as yellow fever, dengue, zika or chikungunya was observed. abstract/case report text a -year-old caucasian male with autosomal recessive hyper igm syndrome type (higm ) due to aicda mutation, diagnosed at age , presented with a newly developed mediastinal mass. he receives routine ivig, pulmonary function tests (pft's) and chest x-rays. at age , patient was noted to have cervical and inguinal lymphadenopathy. ct scan indicated left mediastinal, hilar and pleural lymphadenopathy with soft tissue infiltration around the descending thoracic aorta and esophagus. biopsy indicated no evidence of a lymphoma or infection. years after initial workup, routine pft's showed a declining diffusion capacity by %. patient complained of intermittent chest pain but displayed no clinical symptoms of cough, dyspnea, dysphagia or reflux. ct scan which revealed an extensive illdefined soft tissue mass extending from the thoracic outlet to the level of the esophageal hiatus that encased vascular structures resulting in narrowing and occlusion of left upper lobe pulmonary artery and left lower lobe pulmonary arteries respectively. imaging demonstrated homogenous ventilation to bilateral lungs and decreased perfusion in the left lung compared to the right lung. infectious workup was negative for atypical infections. biopsy revealed miced cellular infiltrate with no predominenant cell type or evidence of malignancy, consistent with previous lymph node biopsy years prior. cd and cd stains revealed aggregates and scattered b-cells and t-cells, respectively. removal of mass was proposed but due to the ambiguous borders and location, surgical excision was not possible. patient was given doses of rituximab ( g), doses mg/kg pulse steroids hours apart, and daily sirolimus (level was adjusted based on sirolimus level). follow-up ct scan indicated significant interval improvement with - % reduction of the soft tissue mass. blood flow in the left lower lobe pulmonary artery has still not returned. this may be due to collaterals and may be a separate problem from compression due to the mass. cytotoxic t-lymphocyte antigen (ctla- ) is an inhibitory immune regulator critical for governing t and b cell homeostasis. heterozygous ctla mutations can cause a syndrome of immune dysregulation with a variable clinical phenotype including hypogammaglobulinemia , autoimmune cytopenia and endocrinopathies, lymphoproliferation, predisposition to malignancy, tissue specific lymphocytic infiltration of brain, lung and gi tract as well as colitis. methods: we retrospectively reviewed medical records of all patients with ctla haploinsufficiency evaluated at the nih between - . a pathologic variant in ctla was confirmed in all patients. we analyzed frequency of campylobacter species detected in the stool samples by pcr based biofilm rapid array as well as reflex bacterial stool and blood cultures when available. results: forty-six patients aged - years were evaluated at the nih between and . six of patients ( %) had at least one episode of campylobacter species associated acute or worsening diarrhea, with one patient also having campylobacter bacteremia. all patients with positive campylobacter species in stool samples had clinical histories and/or endoscopic biopsy findings consistent with enteropathy or colitis predating the incidence of campylobacter infection. two of the six patients ( %) had recurrent or chronic campylobacter infection, while four of the six patients ( %) had multiple gastrointestinal pathogens detected by stool pathogen screening at various times. conclusions: campylobacter species infection of the gastrointestinal tract seem to occur at an increased incidence in our ctla haploinsufficient cohort. to the best of our knowledge, this is the initial report for the association between ctla haploinsufficiency and campylobacter species infection of the gastrointestinal tract. although ctla- is a critical immune checkpoint involved in mucosal immune homeostasis and gut microbiota-immune system cross talk, the underlying mechanism predisposing to campylobacter infection in ctla- deficient patients remains to be explored. our study suggests screening of stool for campylobacter species in patients with ctla haploinsufficiency associated enteropathy. ( ) submission id# abstract/case report text ikaros transcription factor and ikaros family members are critical for development of lymphocytes and other blood cell lineages. full length ikaros (isoform ) contains six c h zinc fingers (zf), four nterminal dna binding zf and two c-terminal dimerization zf. somatic ikaros mutations and deletions have been associated with increased predisposition to b-acute lymphoblastic leukemia (all) as well with poor disease prognosis. recently, germline ikaros mutations affecting the n-terminal dna binding domain and acting in a haploinsufficiency or dominant negative manner were reported to be associated with common variable immunodeficiency (cvid) and combined immunodeficiency (cid), respectively. herein we describe a novel set of germline heterozygous ikaros allelic variants affecting the c-terminal dimerization domains in four unrelated families. clinical manifestations include hematopoietic cytopenias presenting as evans syndrome, and hematologic malignancies including t-cell all and burkitt lymphoma; other manifestations observed were b-cell lymphopenia and hypogammaglobinemia, but recurrent or severe infections were not prevalent or characteristic. we demonstrate that mutants affecting dimerization abolish ikaros homodimerization as well as heterodimerization with ikaros family members aiolos and helios. these variants also affect dna binding at dimerization sites and pericentromeric targeting. opposed to previous allelic variants reported, dimerization changes alter post-translational sumoylation and gene transcription regulation. our data show that mutations affecting ikaros dimerization are mainly associated with cytopenias and/or malignancies, have a different mechanism of action than previously reported variants, present with incomplete clinical penetrance, and contribute to the growing spectrum of genotype-phenotype ikaros associated diseases. introduction: there has been much discussion regarding the return of secondary findings in genetic sequencing research. opinions differ on whether researchers should return secondary findings to participants at all and if so, what the best method is to do so. we have opted to systematically identify and return pertinent secondary findings to participants in our cohort of patients with immune-mediated diseases that undergo exome sequencing. additionally, exome sequencing may determine multiple or other genetic diagnoses in addition to the primary diagnosis, which we call "incidental findings." here, we discuss the secondary and incidental findings discovered in our cohort thus far. methods: individuals in our protocol underwent consent for exome sequencing, including a discussion of the possibility of secondary findings. exome sequencing data was analyzed, and variant pathogenicity was scored using the acmg criteria (richards et al); variants determined to be likely pathogenic, pathogenic, or otherwise clinically important were confirmed via clia-certified sanger sequencing. confirmed variants were returned to participants. we then queried internal databases for cases involving secondary and incidental findings. results: as of november , exome sequencing, interpretation and reporting had been completed for participants. we detected a total of secondary findings in ( . %) participants, including variants in apob, brca ( ), brca ( ), dsp, fbn , kcnh , ldlr, mybpc ( ), ryr , pkp ( ), and vhl. additionally, we detected possible dual/multiple genetic diagnoses in ( . %) participants, some of which explained an unusual clinical presentation or symptom. these included individuals with variants in multiple immune-related genes, including one individual with variants in gata and tnfrsf a, and those with variants in genes related to multiple organ systems, including an individual with variants in ifngr and sco . discussion: exome sequencing in this cohort detects not only important secondary findings, but also discovers a significant portion of individuals with multiple genetic diagnoses. notably, exome sequencing may provide further context or explanation for unusual phenotypic presentation and help determine specific symptom etiology even when a primary genetic etiology is already known. additionally, these secondary and incidental finds may be important to consider when delineating risks and symptoms of novel or recently-discovered conditions. abstract/case report text background: immune dysregulation and lymphoproliferative disorders including alps like disease, hlh ebv driven lymphoproliferative disease leading to rare lymphomas require a multidisciplinary approach utilizing expertise in immunology and hematology/oncology to care for these patients as we learn the molecular etiology of their underlying disorders. at texas children's hospital, the immunology lymphoproliferative evaluation and diagnostic (ilead) clinic was created to provide a comprehensive clinical and research approach to caring for patients with these rare disorders. in an effort to streamline care and access, we recently on-boarded an advanced practice provider (app) . methods: a chart review was conducted months before and after onboarding the app for ilead patient visits. we reviewed the following patient care and access parameters to determine increase in efficient and effective patient care as well as improved access to the clinic. these parameters included: referral process, time of referral placement to appointment, number of patient visits, wait time in clinic, lab interpretation and reporting time for disseminating results to families, and collaboration process with other specialties. results: within months of the app starting our average wait from placing the referral to first appointment fell by an average of %. in addition, we created an algorithm to prioritize patients with immediate need to be seen. by streamlining the referral process and patient priority, we developed a "pre-clinic" conference process by which all patients are reviewed and preliminary plans are made prior to the patient's arrival. this has translated into our ability to increase the number of patients seen in clinic from to and decreased the wait time in clinic by approximately minutes. since the app started, no patient has been in clinic for more than minutes. this has also led to an increase in rvu generation. in terms of efficiency in patient care, all labs are now ordered while in the room with the patient by the app and physician providers. in turn, all labs are resulted directly to the app who reviews labs, collaborates with physicians for care and reports to families in a timely fashion within - weeks of labs being resulted compared to greater than month previously. to improve collaborator communication and post visit plans, a post-visit clinic summary was created. this has been effective in reducing the time to other specialty referrals, follow up visits and effective care for ongoing clinical needs. conclusions: the addition of an app in our ilead multidisciplinary clinic which provides specialized care for patients with immune dysregulation and lymphoproliferative disorders effectively increases work productivity of providers and enhances patient care by increasing access to care, decreasing wait time in clinic and time of reporting of results and future plans. the app with knowledge and expertise in immunology and immune dysregulation is a cost effective way to enhance provider and patient support. with the overwhelmingly positive results, future plans include expanding our multidisciplinary clinic to other services that care for patients with suspected immune deficiency. abstract/case report text introduction: pediatric lymphoproliferative disorders represent a clinically and genetically heterogeneous group of conditions. misdiagnosis and delayed diagnosis can contribute to substantial morbidity and mortality. identification of molecular etiologies and underlying disease mechanisms may facilitate timely interventions and guide targeted or curative therapies. methods: the study was performed through retrospective chart reviews in accordance with all local ethics and irb committees. the study was designed to investigate a cohort of pediatric patients who met criteria for non-malignant lymphoproliferative disorders from texas children's hospital and collaborating centers for underlying genetic etiologies. results: a total of affected individuals from families met criteria. distribution between male and females was nearly equivalent: males (n = ) and females (n = ). approximately half of the cohort was hispanic (n = ). overall kaplan meier survival was % (n = ). whole exome sequencing was performed in all subjects and available family members. likely disease-causing genetic defects were identified in of families ( %). within these families, ( %) carried variants in genes in international union of immunological societies established primary immunodeficiency diseases. potential novel genetic causes of immune deficiency or immune dysregulation were also discovered. mechanistically, all of the implicated genes had roles in modulating lymphocyte activity; initial activation, cytoskeletal organization, or apoptosis of lymphocytes; or regulation of inflammation. all subjects less than one year of age had an identified gene in one of the three mechanistic categories with the dominant mechanistic genetic category being defective control of lymphocyte signaling ( %). in addition, % of patients between and years of age were found to have a potential genetic diagnosis underlying the lpd, with a more equal distribution of mechanistic categories compared to patients greater than years of age where only % have a genetic cause. other important disease manifestations identified were ebv-associated disease in subjects ( %) and subjects ( %) met hlh- criteria. conclusion: primary immunodeficiency diseases and other genetic abnormalities of the immune system underlie a significant percentage of pediatric lymphoproliferative disorder cases. greater than % of patients less than years of age have a genetic etiology underlying the lymphoproliferative disorder. many of these gene defects can be treated with targeted therapies or hematopoietic stem cell transplantation. genetic testing therefore plays an essential role in the diagnosis and management of children with these conditions. abstract/case report text a -year-old gentleman with a history of immune dysregulation polyendocrinopathy enteropathy x-linked (ipex) syndrome with known pathogenic variant in foxp presented to our emergency department with two witnessed episodes of tonic-clonic seizures earlier that day. he has had a longstanding history of recurrent infections and autoimmune conditions since birth, and was being treated with monthly ivig infusions and sirolimus while awaiting bone marrow transplantation. his symptoms on admission included foaming at the mouth, generalized shaking, bladder incontinence, and tongue biting that lasted about five minutes. family reported recent sores inside his mouth and lips, but denied any recent fevers, neck pain, headaches, chest pain, abdominal pain, nausea, vomiting, and sick contacts. he lives on a farm with livestock and reportedly had recent tick exposure. his last ivig infusion was two weeks prior to admission, at which time he also received inactivated flu vaccine. in the ed, a third seizure was witnessed by multiple medical providers. he subsequently received lorazepam and lacosamide with interval improvement. he underwent diagnostic lumbar puncture, as well as extensive evaluation for infections. he was started on empiric antibacterial and antiviral meningitis coverage. analysis of the csf showed a lymphocytic pleocytosis; bacterial cultures and hsv / pcr were negative, as was a -pathogen meningitis/ encephalitis panel performed by pcr. eeg was negative for seizure-like activity, and brain mri showed mild atrophy without sclerosis in the left hippocampus. subsequently, anti-infectious therapy was stopped, and patient was discharged with outpatient followup scheduled for suspected non-infectious aseptic meningitis that was potentially triggered by flu vaccination versus ivig. on day four post-discharge, however, pcr for ehrlichia chaffeensis in the serum returned positive, and he was started on oral doxycycline. ehrlichiosis is a rare tick-borne illness that may cause various nonspecific symptoms including fever, headaches, myalgias, and generalized malaise. most prevalent in the mid-atlantic regions of the united states, tick-borne ehrlichia spreads through the mononuclear phagocytic system and can infiltrate many organs including the kidney, liver, lungs, and heart. csf penetration can cause sometimes fatal meningoencephalitis. aseptic meningitis due to ehrlichiosis has been described in recent literature. cases in hiv patients and transplant patients on chronic immunosuppressive therapy have been severe, resulting in organ dysfunction in many instances and death in a few. however, this case marks the first documented ehrlichia infection in a patient with primary immunodeficiency. this patient's presentation of aseptic meningitis and clear exposure history fits the clinical picture. his relatively benign course could be due to preserved t effector function not seen in persons with hiv or transplant patients with significant immunosuppression. patients with ipex usually present with autoimmunity and allergies, but are also prone to significant infections. it is important to perform a comprehensive workup, including testing for atypical infections, in patients with immune dysregulation syndromes who present with symptoms of unclear etiology. special attention should be paid to patients who live in areas with known endemic exposure risks. empiric antibiotic therapy may need to be considered early to prevent delays in treatment. abstract/case report text introduction autosomal recessive hypomorphic mutations in pgm have been described to result most commonly in either hyper-ige or severe combined immunodeficiency (scid) clinical phenotypes in humans, with one report of an individual with combined immunodeficiency without atopy. herein, we describe a series of individuals newly diagnosed with pgm deficiency functionally confirmed using lectin-based flow cytometric analysis of peripheral blood mononuclear cells, that broadens the associated clinical phenotypes to confirm cid without atopy and childhood evans syndrome. in addition, we present new disease-causing pgm variants, and functionally confirm the pathogenicity of a fourth (p.i t). classical hies phenotype cases . and . identify sisters of spanish descent with a classical hyper-ige phenotype. the younger sibling demonstrated severe atopic dermatitis, mild-moderate asthma, multiple food allergies, one episode of itp, and adhd. the older sibling demonstrated atopic dermatitis, skin infections, and c. albicans otomastoiditis. the siblings were found to have the damaging compound heterozygous variants p.t i and p.q x in pgm . case is a year-old guatemalan boy with prominent atopy including asthma, allergic rhinitis, food allergy, elevated ige, atopic dermatitis, as well as oral hsv who was found to be homozygous for the damaging pgm variant p.i t. cid phenotype with a paucity of atopy case is a -year-old turkish girl who is the daughter of a consanguineous union. she presented with infantile nephrotic syndrome at months of age, and subsequently developed leukopenia, neutropenia, and low igg. complications include bronchiectasis, sinusitis, pseudomonas urinary tract infection, and inflammatory skin lesions without atopy. she was found to be homozygous for the damaging pgm variant p.r h evans syndrome case is a -year-old girl from guatemala. she developed multilineage autoimmune cytopenias including immune thrombocytopenic purpura (itp), autoimmune hemolytic anemia (aiha) and autoimmune neutropenia (ain) at the age of years, refractory to multiple treatments and finally responding to mycophenylate mofetil. she has a history of mild eczema but is without other atopy and suffered from multiple invasive bacterial infections. an additional patient, case , was diagnosed with coombs positive aiha and itp at age years refractory to multiple treatments and finally responsive to cyclosporine. cytopenias recurred year later, resulting in hypoxic brain injury. he died of infectious complications at the age of years. both patients were found to be homozygous for the damaging pgm variant p.i t. discussion this is the first report of pgm deficient individuals presenting with evans syndrome as a primary presentation without additional pathology. while disease-associated mutations appear to cluster around the key conserved domains of the protein, no clear genotype-phenotype correlation is readily observed. in addition to autoimmune cytopenias, pgm deficient individuals have also been reported with splenomegaly, lymphoma, and ebv viremia. thus, in particular for children with lymphoproliferative disease, pgm deficiency should also be considered in the differential diagnosis. abstract/case report text introduction: meningitis is a life-threatening manifestation of cryptococcus neoformans (c. neoformans). it occurs in increased frequency in those with impaired cell-mediated immunity, especially those with hiv/aids. infection with c. neoformans has been seen in previously healthy individuals diagnosed with idiopathic cd lymphopenia (icl). icl is defined by an absolute cd + count of less than cells/m on multiple occasions, usually to months apart, without other apparent cause such as hiv infection, immunodeficiency, or immunosuppressive medications. case description: our patient is a previously healthy -yearold female with cryptococcus meningitis and fungemia. her course was complicated by elevated intracranial pressure requiring extraventricular drain. she was treated with amphotericin and flucytosine for month. notably, the patient was also found to have moraxella catarrhalis (m. catarrhalis) bacteremia without identifiable source. she denied history of environmental risk factors, was not up to date on cancer screening, and recently returned from a trip to italy. initial evaluation revealed lymphopenia ( cells/ul), low cd + ( cells/ul), cd + cells ( cells/ul), and cd / + ( cells/ul), but normal cd + ( cells/ul) and cd + cells ( cells/ul). hiv, ana, leukemia/ lymphoma flow cytometry panel was negative. she also had a normal lymphocyte proliferative responses to pha ( . %), normal cd ra:ro, and protective tetanus titers ( . iu/ml), but only / protective pneumococcal serotypes. initial immunoglobulins demonstrated slightly low igg ( mg/dl). laboratory studies months after presentation demonstrated improved lymphopenia ( cells/ul) continued low cd + cells ( cells/ul), but normalized igg levels ( mg/dl). followup labs also demonstrated decreased cd + b cells ( cells/ ul) and insufficient response to polysaccharide vaccine ( / pneumococcal serotypes). three months after discharge, she is continued on daily fluconazole without recurrence of infections although she still has diplopia and headache. discussion: in a review of cryptococcosis in patients with icl, of them had cryptococcal infection in both the cns and blood. of these patients, was cured, improved, relapsed and then improved, and died. three of these patients were treated with amphotericin and flucytosine. five of these patients had underlying disease and had notable infections with vzv, tb, or hpv, however other infections such as m. catarrhalis were not mentioned. m. catarrhalis bacteremia has been described in children with underlying immune dysfunction and respiratory infection as well as secondary to pneumonia with m. catarrhalis. in cases of m. catarrhalis bacteremia in adults, most had underlying malignancy and/or neutropenia, predisposing respiratory factors, or source for infection. conclusion: this report of c. neoformans meningitis and m. catarrhalis bacteremia in the setting of icl is unusual in that to our knowledge, m. catarrhalis bacteremia has not been reported in icl. cases like this also raise the question as to whether some laboratory abnormalities are secondary to infection, treatment, or underlying disease. it is important to report these cases with icl in order to group disease phenotypes, as continued monitoring and data collection of these cases may lead to discovery of new disease processes. abstract/case report text cytokines play critical roles in regulating the development, survival, differentiation and effector function of immune cells. cytokines exert their function by binding specific receptors on the surface of immune cells and typically activating intracellular jak/stat signaling pathways, resulting in induction of specific transcription factors and regulated expression of target genes. in order to differentiate into an appropriate effector fate, lymphocytes need to integrate multiple signals often provided concomitantly by numerous cytokines that activate shared transcription factors. how these signals are balanced and regulated to yield the optimal class of immune response remains to be completely determined. inborn errors of immunity, or primary immunodeficiencies (pids), result from germline mutations in defined genes, leading to loss-of expression, loss-of function, or gain-of function of the encoded protein. pids are characterised by defects in immune cell development, or their differentiation into effector cells during immune responses, thereby rendering patients not only highly susceptible to infectious diseases, but also autoimmunity, autoinflammation, allergy and cancer. pids are thus an unprecedented model to link defined monogenic defects to immune dysregulation in clinical settings. indeed, pids have unequivocally revealed non-redundant roles of single genes, molecules, signaling pathways and lymphocyte subsets in host defense and immune regulation, and formed the basis of better therapies for immunopathologies. our indepth analysis of inborn errors of immunity of cytokine signalling pathways have identified fundamental requirements for generating long-lived humoral immune responses in humans. here, i will present data relating to our recent studies of how inactivating mutations in il r, il r, znf , stat , stat , and stat , disrupt or dysregulate the generation and function of human memory b cells and tfh cells, thereby precipitating humoral immunity, as well as allergic disease and autoimmunity. abstract/case report text introduction: flow cytometry is a powerful diagnostic tool for detecting hematologic malignancies in a variety of patient specimens including body fluids and lymph node aspirates. cytopathologists are frequently confronted with lymphocyterich effusions, and the definite decision of whether the lymphocytosis is of a purely reactive nature or a presentation of an indolent lymphoma may be an extremely difficult based on microscopy alone. moreover, small proportions of malignant cells that may be missed out by routine morphology can be detected by flow cytometry. objective: the purpose of this study was to evaluate the usefulness of multiparametric flow cytometry immunophenotyping (fci) to confirm the presence of leukemia or lymphoma cells in body fluids and fna specimens. methods: body fluids and fna specimens simultaneously obtained for fci, cytologic analysis and real time pcr from patients were submitted to our flow cytometry laboratory from january to september . the samples studied were body fluids ( pleural fluids and ascitic fluids) and fna samples ( enlarged lymph nodes and lung mass).four color fci method was performed and the following fluorescent monoclonal antibodies were used: cd , cd , cd , cd , cd , cd , cd b, fmc , kappa and lambda light chains, cd , cd , cd , cd c, cd , cd a, cd , cd , cd , cd , cd , tdt, cd , cd , cd , cd , cd , cd , bcl , cd . fci analysis was performed on a beckman coulter cytomics fc flow cytometer using software cxp to analyze data. the cases were diagnosed as leukemia or lymphoma as per tuberculosis ( case). ascitic fluid (n= ) samples showed positivity for angioimmunoblastic t-cell lymphoma ( cases) and dlbcl ( case). fna of lymph nodes (n= ) were positive fort-lymphoblastic lymphoma ( cases), angioimmunoblastic tcell lymphoma ( cases), dlbcl ( cases), hodgkin lymphoma ( case), nodular lymphocyte predominant hodgkin lymphoma ( case), peripheral t-cell lymphoma (nos) ( case), splenic b-cell marginal zone lymphoma( case), tuberculosis ( cases). one fna of lung mass were tumor of neural cell origin. both immunophenotype and cytomorphology positive for malignancy were in / ( . %) cases.cytomorphology was negative/ suspicious in / ( . %) cases, of which both cytomorphology a n d i m m u n o p h e n o t y p e n e g a t i v e w e r e ( % ) a n d cytomorphology negative but immunophenotype positive cases were ( . %). mtb dna was detected in pleural fluid in case and fna sample in cases. conclusion: multiparametric flow cytometry by using comprehensive panel of monoclonal antibodies is a useful diagnostic test to evaluate body fluids or fna as it can demonstrate small malignant populations that may be missed out by routine cytomorphology. clinical laboratory geneticist/department of genetics, university of groningen, groningen, the netherlands abstract/case report text the phenotypes of primary antibody deficient (pad) patients range from milder (e.g. specific antibody deficiency) to severe (e.g. x-linked agammaglobulinemia) deficiency of the immune system. pad patients form a clinically, immunologically as well as genetically heterogeneous group. often, the genetic background has not been elucidated; it probably is not monogenetic in a large subgroup of patients. pad patients suffer most frequently from recurrent bacterial infections of the respiratory or gastrointestinal tract due to immune deficiency, but may also have varying degrees of autoimmune and lymphoproliferative comorbidities due to immune dysregulation. unfortunately, a standardized description of pad phenotypes is lacking rendering robust definitions of pad-subtype diagnoses, including cvid, difficult. this impairs the formation of homogeneous cohorts that can form the starting point for future clinical and genetic research. the pad subgroup of the human phenotype ontology (hpo) immune mediated disorders consortium supported by ern rita and esid is addressing the gaps in standardized phenotypic description of pads. using the hpo dataset, literature mining, and esid, iuis and omim classifications, we aimed to reevaluate and complete the pad-related hpo terms to allow efficient data exchange and matching of phenotypically similar pad patients. as a principle, it was decided to avoid the ongoing variance in pad-subtype definitions and to build the pad-related hpo tree based as much as possible on unambiguously interpretable items. 'hypogammaglobulinemia' was deleted as hpo term, and replaced by separate hpo terms such as 'decreased total igg in blood', subdivided in 'transient' vs. 'chronic', and '(near) absent' vs. 'partially decreased' (the same for igg , igg , igg , igg , iga and igm). 'decreased specific antibody level in blood' was specified further into 'decreased natural antibody level to blood group antigens in blood', subdivided in '(near) complete' vs. 'partial' absence (the same for protein, polysaccharide and protein-conjugated polysaccharide vaccination). relevant hpo terms related to infection and to specific organ manifestations like bronchiectasis, autoimmunity and lymphoproliferation were re-evaluated and completed, and will be linked to pad diseases in the hpo online system by the pad subgroup experts. once finalized, existing pad cohorts will be classified according to the new hpo pad-related terms, and studied by clustering technologies (example of two patients shown in figure ; white = absent, color = present). acceptance and widespread use of this pad-related hpo tree for standardized phenotyping will be essential to empower future multicenter clinical research and related genetic discoveries as well as support clinicians in diagnosing pad through the linkage of hpo terms to pad disease entities. senior investigator oral immunity & infection section/nih/nidcr abstract/case report text leukocyte adhesion deficiency type (lad ) is an autosomal recessive disorder characterized by the inability of granulocytes to emigrate from the bloodstream to sites of inflammation. lad is caused by mutations in the itgb gene ( q . ), encoding the beta- -integrin, cd , which is essential for firm adhesion of leukocytes to the endothelium. in lad survival is compromised, morbidity from inflammatory lesions is high, and treatment is poor. the moderate form of lad is often managed with antibiotics for prophylaxis and during acute infections. after infancy severe gingivitis and chronic periodontitis are universal. periodontal findings affect primary and permanent teeth, causing intense oral mucosal (gingival) inflammation and destruction of tooth supporting bone, which are hallmarks of the disease periodontitis. blocking the il- /il cytokines, which are up regulated in lad gingiva, can reduce bacterial load and resolve inflammatory gingivitis. ustekinumab binds to the shared p subunit of human il- and il- , cytokines that modulate lymphocyte function, including t helper (th) cells and th subsets, thereby blocking them. objective: explore the effect of ustekinumab on lad inflammatory disease. method: prospective study using ustekinumab for oral inflammation. patients receive five doses over year, -or mg depending of weight. results: (two patients have enrolled, p is > year post treatment, p is still on study) patient characteristics · patient : age at diagnosis, yrs.(itgb mutation c. delt (null)); cd (%pmn control): . %; cd a(%pmn); . %. at the initiation of the protocol ( yrs old) wcc: . k/ul; crp: . · patient : age at diagnosis, yrs.(itgb mutation c. a>g,p.g s c. c>t,p.a v) cd (%pmn control): %; cd a(%pmn): . %. at the initiation of the protocol ( yrs old) wcc: . k/ul; crp: . response: patient : oral ulcers before treatment: episodes every two months. during ustekinumab therapy: · oral ulcers: episode in a year · reduction in bleeding on probing: . % · gingival index reduction: % patient : oral ulcers before treatment : monthly. during ustekinumab therapy: · oral ulcers: none in first months · reduction in bleeding on probing : . % · gingival index reduction: . % safety: no significant adverse events were documented during the therapy p had a previous skin lesion that flared leading to iv antibiotics. p had a previous sebaceous cyst drain spontaneously. discussion: two patients showed improvement in chronic periodontitis and a substantial decrease in oral ulcers while on ustekinumab. no clear safety signals were seen. durability of these findings is still unknown. ustekinumab in lad deficiency appears to be safe and potentially effective. post doctoral fellow/servicio de inmunología, inst. multidisciplinario de investigación en patologías pediátricas (imipp), hospital de niños ricardo gutiérrez. chief resident/servicio de inmunología-hospital de niños "dr. r.gutierrez" immunologist/centro de inmunología clínica "dra. liliana bezrodnik y equipo"-servicio de inmunología htal. de niños "dr. r.gutierrez" immunologist/centro de inmunología clínica "dra. liliana bezrodnik y equipo" immunologist/servicio de inmunología-hospital de niños "dr. r.gutierrez" abstract/case report text introduction: primary immunodeficiencies with dysregulation associate defects in the immune homeostasis leading to inappropriate immune response (lack or excess) that causes autoimmunity, allergy and/or inflammation. impairment of different subsets of t and b compartments may be associated with these pids. aim: ) describe t and b memory compartment of pid patients (pts) with dysregulation: cd deficiency, stat gof, stat b deficiency, ctla variant, pi kcd variant and cvid-like (with no molecular defect) and compare them with a group of healthy donors (hd). ) associate ctfh profile with b cell compartment impairment. results: ) pts showed a significant decrease of naïve cd + t cells (cd ra+cd +) ( , % vs , %) (p < . ) with expanded central memory t cells (cd ra-cd +) ( . % vs . %) (p < . ); cd + t cells had higher levels of activation markers (cd +hla-dr+) ( , %vs , %) (p < , ). pts showed a significant increase of circulating follicular t cells (ctfh) (cd ra-cxcr +) compared with hd (mean , % vs , %) (p < , ) with pd- overexpression (p < . ). stat gof, ctla , pi kcd and cvid-like pts showed a skew towards ctfh (cxcr +). regulatory t cells (cd +cd ++ foxp +) were absent in cd and stat b deficiency and decreased in the other pts. within cd + cells, although effector memory (cd ra-cd -) (p < . ), temra (cd ra+ cd -) (p < . ) and hla-dr+ cd + (p < . ) subsets showed a significant increase compared with hd, the behaviour was variable between different mutations. regarding b cell compartment, pts with stat gof, pi kcd and cvid-like showed a severe impairment of switched-memory b cells (sw-mbl) (cd +igd-igm-); the stat b deficient patient had increased frequencies of this subset, while ctla pts had a variable b defect. ) lower sw-mbl values were significantly associated with lower values of ctfh cells (p < . ) (r= . ). cd low b cells were exclusively high in cvid-like pts, and transitional b cells were increase in pi kcd and almost all cvid-like pts. discussion: in summary, patients with dysregulatory syndromes associate a defect of t and b homeostasis (survival, activation and differentiation). specific mutations can differentially affect the quantity and/or the quality of ctfh. there is a strict association between the differentiations of tfh with th profile with the generation of sw-mbl. these alterations may play a role in the pathophysiology of primary immunodeficiencies with b lymphocyte functional impairment. immune monitoring of lymphocyte subsets of patient with dysregulation may approach to the diagnosis of specific monogenic mutations. objective: the purpose of this study is to increase awareness and improve diagnosis of primary immune deficiency (pid) in the heterogenous group of patients with autoimmune cytopenia (aic) by identifying clinical characteristics and laboratory biomarkers that distinguish those with underlying pid, disease activity and guide mechanism-based targeted therapy. methods: patients with aic (autoimmune hemolytic anemia (aiha), immune thrombocytopenia (itp), or autoimmune neutropenia (ain)) were referred to our immune dysregulation team and prospectively enrolled during - . detailed immune phenotyping (igg, iga, igm, lymphocyte subsets, vaccine titers, lymphocyte proliferation to mitogens/ antigens), serum lipopolysaccharide (selps) and autoantibodies were measured and/or collected by chart review and genetic testing for pid was pursued. results: from to , patients were enrolled; two subjects were removed due to parental request or lack of aic diagnosis. of the remaining patients, ( %) were classified as "aic-pid" based on genetic testing and/or immune phenotyping; ( %) were classified as aic-only, and ( %) were asymptomatic family controls. the patients were predominantly children (ages - years, average age . years); % ( / ) were male. among patients who have had genetic testing to date (n= )( %), pathogenic genetic mutations were confirmed in / ( %) of patients. mutations include fas/fasl (n= , including family members without aic), ctla (n= ), q (n= ), and one patient each with nfkb , was, pole- , pi k, casp , card , and cgd; the remainder of aic-pid patients were classified as combined immune deficiency or common variable immune deficiency based on immune phenotyping. lymphocyte subsets (cd +t, cd +t, cd +b, cd + nk) and immune globulins (igg, iga, igm) tended to be lower in aic-pid patients vs aic-only (p < . ). evans syndrome was more common in aic-pid patients ( / , %) compared to aic-only ( / , %). lps was elevated in the serum of aic patients compared to healthy controls (mean vs pg/ml, p < . ). excluding partial digeorge syndrome patients (average lps pg/ml), selps levels were significantly higher in genetically-defined untreated pid patients (average pg/ml) vs. other pid (average pg/ml)(p= . ) or patients with aic alone (average pg/ml)(p= . ). studies are ongoing on specific subsets that are linked to immune dysregulation (switched memory b cells, t-regulatory cells, double negative t cells, t follicular helper cells) and the use of soluble il- as a biomarker of disease activity. conclusions: a high fraction of aic patient were identified with underlying pid in our study. basic immune evaluation with immunoglobulin levels and lymphocyte subsets expedited diagnosis of pid. genetic evaluation distinguished a group of patients with aic-pid and highly elevated lps level, reflecting high bacterial load, which may distinguish them from the rest the aic cohort. the source of bacterial lps can be multifactorial and is yet to be determined. our studies continue focusing on biomarkers that can be applied to the heterogenous group of patients with aic. this will allow early detection and timely initiation of targeted therapies. investigator/dermatology branch, niams, nih head, dermatology consult service/dermatology branch, niams, nih chief, fungal pathogenesis section/laboratory of clinical immunology and microbiology, nih abstract/case report text introduction/background: autoimmune-polyendocrinopathy-candidiasis-ectodermal dystrophy (apeced) is a monogenic autoimmune disease resulting from biallelic mutations in the aire gene. although typically characterized by the classic triad of chronic mucocutaneous candidiasis, hypoparathyroidism and adrenal insufficiency, we recently reported that the clinical spectrum of the syndrome is far broader and that incorporation of an adjunct triad of apeced rash, intestinal dysfunction, and enamel hypoplasia in the classic triad could lead to earlier diagnosis (ferre et al., jci insight, ). among the adjunct triad manifestations, apeced rash occurs in % of american apeced patients by age , most often developing in the first year of life. objectives: to report and describe the clinical features of apeced rash as the first manifestation in a -month old patient with apeced. methods: following enrollment in a niaid irb-approved protocol ( -i- ) the patient was evaluated with history and physical examination, aire sequencing, measurement of interferon-autoantibodies, and skin biopsy with immunohistochemical analyses. results: a -month-old girl with a family history of genetically confirmed apeced in her -year old sister developed discrete circular, maculopapular erythematous lesions on her torso that spread to the face, arms, and legs while sparing the palms and soles. the rash was partially blanching, non-painful and non-pruritic and was preceded by low-grade fever ( □c) without other accompanying symptoms. she had not received medications or vaccinations prior to the rash onset. the lesions increased in size with associated central clearing and resolved . months after onset. the rash recurred with similar appearance times over months with each recurrence lasting between days and . months. as with the first rash episode, recurrences were often preceded by fever ( - □c) without accompanying symptoms or inciting factors. neither topical nor oral antihistamines improved the rash. aire sequencing identified the same compound heterozygous mutations (c. _ del and c. c>t) that the sister has. high titers of interferon-□ autoantibodies were measured in serum. skin biopsy revealed superficial perivascular chronic inflammation and intraepidermal lymphocytes composed predominantly of mixed cd and cd t lymphocytes with few perivascular b lymphocytes. no eosinophils or vasculitis was observed. myeloperoxidase immunostaining revealed extensive karyorrhexis. laboratory studies revealed normal white count and esr, negative anti-ige receptor antibody, and positive anti-ige antibody. at months, she developed oral candidiasis as second manifestation of apeced, thus reaching a diagnostic dyad when applying our proposed expanded diagnostic criteria. she has not developed hypoparathyroidism or adrenal insufficiency; thus, she has not yet reached a classic diagnostic dyad. systematic screening for these endocrinopathies will be needed to avoid lifethreatening complications of acute endocrine failure. conclusions: we report the clinical and histologic features of apeced rash manifesting as the first disease component of apeced in a month old girl. apeced should be considered in the differential diagnosis of recurrent erythematous maculopapular urticaria-like eruptions characterized by mixed lymphocytic and neutrophilic infiltration unresponsive to antihistamines. our case illustrates the clinical utility of incorporating the expanded diagnostic criteria of apeced rash, enamel hypoplasia and intestinal dysfunction into the classic diagnostic triad, which can lead to earlier apeced diagnosis. who presented with prolonged severe neutropenia despite g-csf and seven hospitalizations for febrile neutropenia in the span of ten months. prior to his neutropenia, patient was on monthly ivig, with igg trough - in the past year. he was evaluated for bmt in but declined. patient was first found to be neutropenic in aug when he was admitted with pseudomonas thigh abscess, hsv stomatitis and rhinovirus infection. he was treated with broad-spectrum antibiotics with improvement in neutropenia. the following month, he was hospitalized again with neutropenic fever, left axilla pseudomonas abscess and rhinovirus infection. he underwent bone marrow biopsy revealing left shifted myeloids with decreased maturing forms and t cell predominant lymphoid aggregates, suggestive of autoimmune neutropenia vs. hyper igm syndrome associated with neutropenia. anti-neutrophils antibodies were negative. he was then admitted the following month ( / ) with febrile neutropenia with cxr concerning for viral pneumonitis vs. atypical pneumonia. he was started on g-csf therapy with significant initial response in anc. however, this response was short-lived as he was again admitted in / with febrile neutropenia and upper respiratory rhinovirus infection. he was continued on daily g-csf. due to persistently normal anc for approximately three weeks, he was weaned off g-csf in / . in / and / , he had two more hospitalizations for febrile neutropenia. g-csf was restarted with dose uptitrated to mcg/kg during his hospitalization in april. he was found to be thrombocytopenic with splenomegaly on abdominal ultrasound. anti-platelet antibodies and repeat anti neutrophil antibodies were not detected. he was discharged with close follow up with immunology and hematology. due to his age, he was transitioned to penn allergy/immunology in / . there was close communication between chop allergy/immunology, chop hematology and penn allergy/immunology during this transition period. patient was admitted to hup in / with febrile neutropenia (despite higher dose of g-csf), rhinovirus infection, pseudomonas sinusitis and ct chest findings suggestive of possible fungal pneumonia. due to persistent neutropenia refractory to g-csf treatment, hematology was consulted and repeat bone marrow biopsy showed hypercellular bone marrow with markedly left shifted granulocytic hyperplasia, compatible with g-csf therapy. flow cytometry showed no evidence of plasma cell neoplasm. dose of ivig was adjusted and increased based on his weight. per hematology, he also received an additional high dose ivig g/kg x days for presumed immune mediated neutropenia with immediate increase in anc. despite anc of for days,anadditional g/kgofivigimprovedhisancto> within hours of his first dose. thrombocytopenia also improved to normal range. since then, patient has been on monthly - mg/kg ivig with no recurrence in neutropenia. this patient's prolonged persistent neutropenia with immediate response to high dose ivig is suggestive of autoimmune neutropenia, which should be taken into consideration in hyper igm patients with persistent neutropenia. abstract/case report text background: children with digeorge anomaly (dga) represent a heterogenous group, often classified as either partial dga (pdga) or complete dga (cdga) based upon the degree of thymic hypoplasia. this paucity of t-cell parameters and function has serious implications for infection risk, autoimmunity, and malignancy. however, there are limited studies stratifying children with dga by these subgroups, especially in regard to immune function and subsequent infection risk. study design: single-center, retrospective cohort analysis evaluating the relationship between pdga and cdga to infection risk with particular focus on infection-related hospitalization, pathogenic organism identification, and antimicrobial resistance profiles. the source population includes all pediatric patients < years of age diagnosed with either pdga or cdga while receiving care at duke university from january , to june , . the final analysis sample included patients. methods: to evaluate the differences in immune function between dga subgroups, we will report the proportion of low ( < th percentile for age) t cell immune biomarkers for both subgroups and compare populations using a chi-squared test. to compare per year incidence of infectionrelated hospitalization for dga subgroups, a poisson model with number of hospitalizations per patient as the outcome, an offset equal to the time at risk for hospitalization, and either pdga or cdga diagnosis as the exposure will be used. models will be bivariate. we will report an incidence rate ratio (irr) and % confidence interval ( % ci). to evaluate the impact of cellular and humoral immune function on infection-related hospitalization, we will use poisson models where the outcome is the number of hospitalizations per patient, an offset equal to the time at risk for hospitalization, and low immune biomarker as the exposure. all models will be bivariate. we will report an irr and % ci. infection type and resistance profiles will be completely descriptive. results: as expected, children with cdga had a significantly higher probability of a low ( < th percentile for age) values for total t cells (cd +), helper t cells (cd +cd +), cytotoxic t cells (cd +cd +), and naïve helper t-cells (cd +cd ra+cd l+) as well as a significantly lower probability of low pan memory t-cells (cd +cd ro+) compared to children with pdga. no differences were detected in the percentage of low natural killer (nk) cells (cd +cd +) or b cells (cd +) between subgroups. cdga patients had a significantly higher incidence of hospitalization per year ( . ( . , . )) compared to pdga patients ( . ( . , . )). the irr is . ( . , . ). across both subgroups, the incidence of hospitalization was higher in dga patients who had low helper and naïve t-cells. there is ongoing analysis into hospital-related infection and resistance profiles. notable frequencies include bacteremia ( > %), invasive viral disease ( > %), and opportunistic infections ( > %). conclusions: children who had cdga were % more likely to have an infection requiring hospitalization than children who had pdga, emphasizing the need for thymus transplant for cdga. further analysis of infection type and patient outcomes is critical to enhancing management of this unique patient population. abstract/case report text antibody cross-reactivity among flavivirus has been documented. in recent times zika virus has been emerging in pockets of the mosquito-infested regions, while southwestern saudi arabia is known for arthropod-borne viral diseases and we do not know the incidence or even presence of zika virus in this region. it is restricted to predict the igm and igg antibody detection ranges owing to limited data and colossal cross-reactivity among the zika and other flaviviruses. we tested sera from pregnant women irrespective of their clinical presentation for zika and dengue igm, igg respectively. the zika positive samples were further confirmed by plaque reduction neutralization tests (prnt). from our results, . % ( ) cases were positive for zika igm against . % ( ) positivity to igg. when these samples were assessed for dengue igm and igg, we observed . % ( ) seropositivity for igm and igg respectively. there was no single sample positive for both igm and igg of zika or dengue. however, we observed one sample positive for both zika and dengue igm. upon mapping the overlapping serotiters, there was no significant correlation observed between the dengue igm and igg. whereas zika igg positive sample showed high serotiter for dengue igg indicating the contribution of cross-reactivity for observed zika positivity. screening for the incidence of zika, therefore, becomes particularly hard in a population that has the presence of pre-exposure of dengue and this cross-reactivity makes it hard to determine the zika incubation and antibody prevalence confounded with other flaviviruses. abstract/case report text introduction humoral pid diagnostic protocol includes the analysis of the immune response to different protein and polysaccharide antigens (ags) ( ) . although the analysis of the immune response against the polysaccharides vaccine from pneumococca has been the standard method, the use of s. typhim vi vaccine has appeared as a good alternative ( ) . in this report we show the results obtained with the use of s.typhim vi in adults patients attending the pid outpatient clinic. material and methods patients with humoral-suspected pids were challenged with typhoid polysaccharide vaccine (typhim vi®; sanofi-pasteur). serum was obtained on basal and after weeks of vaccination. specific igg levels against s.typhim were measured using "vacczyme tm human anti-salmonella typhi vi igg enzyme immunoassay kit" (binding-site). results a total of adult patients attended the pid clinics during nov -nov . from those, patients were fully evaluated using a humoral-suspected pid algorithm that includes the s.typhi vaccination. in total male and female patients completed the protocol and were analyzed. twenty patients were considered as responders (ratio pre/ post > x) whereas patients were non-responders. discussion the main advantage of assessing polysaccharide immune response using s.tyhpim is the usual lack of specific igg at the moment of the initial evaluation. in this serie, just one patient has a high basal level (# -vaccinated in the past). thirteen patients had basal levels below the detection limit of the test ( , u/ml) and patients between , and , u/ml, that has been described as a cut otf level for nonimmunised individuals (personal experience, , ). regarding the polysaccharide immune response as a tool to distinguish pid vs non-pid patients, the results showed a good correlation between those non-responders with more clinical relevant pid diagnostics. seven non-responders patients were subsequently diagnosed with a primary (* on table i ) and/or secondary id (** on table i ). despite this, there were patients that we could have classified as strong responders (ratio > x, absolute specific igg postvaccination level > u/ml) and patients considered as weak responders (ratio post/pre > x, absolute specific igg postvaccination level < u/ml). strong responders were considered non-pid after including other clinical investigations and laboratory tests (cell subpopulation study, pcp response) whereas weak responders group consisted in some "minor" forms of pid, like isolated igm or ig subclasses deficits. more patients are needed to confirm this functional classification of pid patients regarding their s.typhi immune response. abstract/case report text activated pi kδ syndrome (apds) is a primary immunodeficiency characterized by recurrent respiratory infections, as well as increased risk of chronic viremia with herpes family viruses, benign lymphadenopathy and b cell lymphoma. it is caused by heterogeneous germline gain-of-function mutations which ultimately lead to the hyperactivation of the phosphoinositide- -kinase δ (pik δ). pik δ exists as a heterodimer composed of a catalytic and a regulatory subunit. it interacts with b cell receptors, t cell receptors, costimulatory and cytokine receptors, and is a key player in a signaling pathway involved in cell growth, proliferation and survival. apds is caused by mutations in the pik cd gene, affecting its protein product p δ (catalytic subunit). apds is caused by mutations in the pik r affecting p a (regulatory subunit). short syndrome is a rare multisystem disorder characterized by short stature, hypertextensible joints, ocular depression, reiger anomaly and tooth eruption delay. the primary causes of short syndrome are heterozygous loss-of-function mutations in the pik r gene. the combination of apds and short syndrome is very rare, with only few cases described in the literature. in this report we present a teenager with a pathogenic variant in the pik r gene, and phenotypic characteristics of both apds and short syndrome. our patient is a -year-old female with a history of growth delay and short stature, delay tooth eruption, recurrent sinopulmonary infections and hypogammaglobulinemia. evaluation performed at a prior institution for recurrent infections revealed low igg levels. she did not initiate therapy at that time and was lost to follow up for several years. at the time of our initial evaluation she reported continued recurrent episodes of upper respiratory infections and sinus infections requiring antibiotic treatment that often did not clear the infections. her physical exam was relevant for short stature ( %ile, z=- . ), low weight for age ( < %ile, z=- . ) and hyperextensibility. her facial features were significant for prominent forehead and triangular face. given concern for immune deficiency, a complete immune evaluation was obtained. her workup revealed low igg levels, with igm and iga within normal limits. she did not have protective titers to s. pneumoniae, h. influenza or diphtheria and tetanus. after administration of vaccine boosters, she was able to generate a response to all vaccines except for tetanus. she had remarkably low absolute b cells ( cells/ul) and percentage ( %), and low cd :cd ratio ( . ). she was started on amoxicillin prophylaxis and monthly ivig replacement therapy. invitae immunodeficiency panel genetic testing was sent and revealed a pathogenic loss of function variant in an intronic splice site in the gene pik r (c. + g>c). after initiating treatment with ivig, her sinus infections significantly improved and she has not had any further episodes. igg levels have remained within normal limits with monthly ivig therapy. this pathogenic variant had been previously associated with apds ; however, it had not been associated with short syndrome. the mechanisms that link both conditions is yet to be identified. this case report emphasizes the importance of screening for comorbidities associated with short syndrome in apds patients, and vice versa. finding the genetic diagnosis for patients with suspicion of primary immunodeficiency (pid) is becoming increasingly important in the management of primary immunodeficiency and estimating the risk for family members. we constantly increase the diagnostic yield for pids by improving the sequencing technology, updating the panels with new genes discovered related to pid, and finding diagnoses from difficult to sequence regions and regions with high homology. here we report our experiences with nearly patients suspected with pid. moreover, we provide a case example, how we increase the diagnostic yield by developing unique techniques for specific genes which cannot be reliably analyzed by ngs alone. diagnostic yield including all immunology related panels was . % ( / ). the majority of the tested individuals were males ( / , . %) and the most common age of testing was between to years ( / , . %). the highest diagnostic yield . % ( / ) is in children from ages to years, whereas in patients over years of age the diagnosis was found for only . % ( / ) of the patients. in two patient cases, our cnv detection algorithm indicated a homozygous deletion in the index patient samples potentially covering the whole ncf gene. additional bioinformatic analysis targeting specifically two coding positions that differ between the ncf gene and the two pseudogenes showed that all reads in those positions originated from the pseudogenes. homozygous deletion in the ncf gene was further confirmed by sanger sequencing two regions in ncf with primers that specifically bind to either ncf or the pseudogenes. while clean ncf sequences from both regions were obtained for a control sample, no ncf -specific amplification product was obtained for the index patient samples. pseudogenes were amplified and sequenced successfully in both index patient samples and positive control samples. loss-of-function of ncf is a well-established mechanism leading to cgd and by overcoming the difficulties regarding ncf deletion detection by ngs, we can improve diagnostic rate in individuals affected with cgd. gata deficiency can lead to a broad spectrum of clinical and hematological phenotypes; in some cases, nk cell deficiency is the primary manifestation, resulting in a greatly increased susceptibility to viral infections and malignancy. gata -deficient patients, particularly those who suffer from severe viral infections, have reduced frequencies of peripheral blood nk cells and loss of function in the existing nk cells. specific loss of the less mature (cd ^bright) nk cell subset is a hallmark of the immune phenotype in gata deficiency, suggesting that generation or survival of nk cell precursors is impaired. given the remarkable spectrum of clinical phenotypes in gata -deficient patients and the poorly understood biology underlying their nk cell defect, we sought to characterize circulating nk cells on a single-cell level. we performed single-cell (sc)rnaseq of lineage-depleted innate lymphocytes from a patient with gata deficiency. as expected from flow cytometric phenotyping of peripheral blood cells from this and other patients, scrnaseq revealed decreased representation of canonical cd ^bright cells. within the cd ^dim population, we identified two nk cell populations that were seemingly unique to the gata deficient patient relative to a healthy donor. pathway analysis defined the first of these populations (population ) by the expression of genes associated with cellular response to stress, extracellular stimulus and inflammation, as well as programmed cell death and regulation of proliferation and apoptosis. the second population (population ) was defined by genes associated with nk cell chemotaxis, cytokine responses and interferon signaling. to extend our findings, we performed scrnaseq of additional healthy donors and analyzed an additional gata -deficient individual who was clinically asymptomatic (yang et al. ). of note, we detected population in seemingly healthy cmvnegative individuals, suggesting it was not uniquely a result of gata deficiency but associated with an inflammatory response and not related to adaptive nk cells generated in response to cmv infection. population , on the other hand, only appeared in our symptomatic gata -deficient patient. we additionally performed bulk gene expression analyses from an unrelated gata -deficient patient that confirmed the altered expression of genes associated with both novel cell populations. current efforts are focused on better defining the functional response of nk cells in these patients and confirming the identification of our novel populations by mass cytometry (cytof). together, our data define the heterogeneity and complexity of nk cells in gata deficient and healthy individuals. perforations have been reported with tocilizumab, a monoclonal antibody of the interleukin (il- ) receptor, suggesting that il- signaling plays a role in intestinal wall integrity. as il- signals through stat , we sought to investigate the potential association between lof stat and intestinal perforations, as well as the incidence and outcome in our patient cohort. methods: we performed a retrospective chart review of patients with lof stat (n= ) followed at our institution, looking for those with non-malignancy associated spontaneous gastrointestinal perforations. the demographic information, stat mutation, comorbidities at the time of perforation, clinical presentation, management, and clinical outcomes were compiled results: ten lof stat patients were identified as having documented intestinal perforations, an approximate rate of %. one perforation was the initial presentation of diffuse large b cell lymphoma (dlbcl) of the duodenum and liver, and was excluded from the rest of the analysis. the other nine perforations occurred between to years old (mean: ), and % were female. stat mutations were localized to the dna binding domain (n= ) and the sh domain (n= ). two of the perforations occurred while inpatient for lung infection. another occurred while recovering from pneumonia at home. two perforations were associated with the initial diagnosis of diverticulitis (at age and ). one perforation occurred in the terminal ileum, one in the cecum, one in the transverse colon, and six in the sigmoid. five patients underwent primary closure of the bowel. four patients required a temporary ostomy, with subsequent successful ostomy reversal. only one patient has since died of pulmonary hemorrhage, the other patients are alive with a mean of years post perforation follow-up, with no recurrence of perforation. one patient with prior sigmoid resection required ileal resection post perforation due to as massive intestinal bleed. conclusion: spontaneous gastrointestinal perforations occurred in our lof stat cohort at a rate of approximately %. one case was associated with malignant infiltration of the gastrointestinal tract and two cases were associated with diverticulitis both known risk factors for perforation. although the pathogenesis of the perforations in lof stat remains unclear, the connective tissue phenotype likely contributes as well as the association with diminished il- signaling, as has been demonstrated with the perforations and tocilizumab. abstract/case report text background granulomatous and lymphocytic interstitial lung disease (glild) is a life-threatening complication that occurs in patients with common variable immunodeficiency (cvid) and monogenic cvid-like disorders, but the optimal treatment is unknown. objective to determine if the use rituximab and azathioprine (rtx-aza) or rituximab and mycophenolate mofetil (rtx-mmf) would improve the radiographic abnormalities as determined by high-resolution computed tomography (hrct) of the chest and/or pulmonary function tests (pfts) in patients with cvid and glild. methods this is a retrospective study of patients seen from july to december with cvid and glild who completed immunosuppressive therapy (rtx ( mg/m ) for weeks, repeated at -month intervals for or total courses, and aza ( . - . mg/kg/day) or mmf ( mg- mg-bid) for months). complete pfts and hrct scans were performed prior to therapy, at the conclusion of therapy, and periodically thereafter. hrct scans were blinded, randomized, and scored independently (in pairs) by two radiologists. all patients underwent whole exome sequencing (wes). number (percentage) and median (interquartile range) were reported for categorical and continuous variables, respectively. differences between pre-and post-treatment and between relapse and post-relapse hrct scores and pft parameters were analyzed with wilcoxon signed ranks test. kaplan-meier survival curves were also done. unadjusted one-sided p-values < . were considered statistically significant. results the glild cohort (n= ) had a : female predominance, and age at glild diagnosis was ( - ) years (table ) . autoimmunity was present in the majority of patients, with thrombocytopenia ( ( %)) the most common manifestation. enteropathy ( ( %)), inflammatory bowel disease ( ( %)), and nodular regenerative hyperplasia of the liver ( ( %)) were also present. splenomegaly ( ( %)) was present in the majority, but polyarthritis ( ( %)) was notably absent. twenty ( %) patients had been previously treated with systemic steroids. hrct scores substantially improved between pre-and posttreatment for rtx-mmf (p= . ) and rtx-aza (p < . , figure ). fev (p= . ), fvc (p= . ), and tlc (p= . ) also improved, but dlco (p= . ) was unchanged (figures and ). excluding two ( %) patients who died . and years after therapy of respiratory failure ( ( %)) and septicemia ( ( %)) respectively, / ( %) patients relapsed . ( . - . ) years following therapy with an estimated % relapse rate after years ( figure ). as of december , of patients that relapsed showed improvement in hrct scores (p= . ), and the remaining patients are still undergoing retreatment ( figure ). four ( %) pneumonias occurred during immunosuppressive therapy, all with severe restrictive lung disease . eight ( %) patients had a damaging mutation in a gene known to predispose (tnfrsf b, n= ( %)) or cause a cvid-like primary immunodeficiency (ctla : ( %); kmtd : ( %); birc : ( %)). immunosuppressive treatment improved the hrct scores regardless of the absence (p < . ) or presence of a damaging mutation (p= . ) ( figure ). conclusion combination chemotherapy appeared to be effective in improving the radiographic abnormalities and pulmonary function of patients with cvid and glild. a majority of patients had sustained remissions, regardless of the presence or absence of a monogenic disorder. iqr=interquartile range, cvid=common variable immunodeficiency, glild=granulomatous and lymphocytic interstitial lung disease, vats=video-assisted thoracoscopic surgery, tbx=transbronchial biopsy, ms=mediastinoscopy excluding b cell malignancy, pft=pulmonary function test, h/o=history of, dz=disease, nrh=nodular regenerative hyperplasia, ibd=inflammatory bowel disease table . baseline patient characteristics since its first description the number of cases has increased progressively [ ] . although described as a predominant antibody deficiency [ ] , various complex phenotypes have been associated with mutations in this gene [ ] . case description. female patient with no remarkable history until years old, when she suffers from a persistent fever associated with purulent abscesses in venipuncture areas and hyperleukocytosis with neutrophilia, so she was treated for about months in hospitals in the city of barranquilla before she was referred to our institution. the patient's clinical picture consisted of persistent fever unresponsive to broad-spectrum antibiotic treatments, skin abscesses, left subphrenic abscess and toes osteomyelitis. the microbiological studies documented a bacteremia by acinetobacter baumannii and isolation in bone marrow of candida parapsilosis. during her care stay in barranquilla, she was approached as a chronic granulomatous disease versus job's syndrome, she received two doses of immunoglobulin with partial control of symptoms. due to the recurrence o f fever, abs ces ses and hyperleukocytosis, they decided to refer to our institution for further studies. upon admission to our institution, the patient presented nutritional compromise, with spontaneous resolution of fever but persistence of high acute phase reactants, with significant improvement of leukocytosis. all the cutaneous lesions she presented were debrided at the site of remission. immunoglobulin levels, lymphocyte populations and dyhidrorhodamine test were normal. she remained with no weight gain, constipation, abdominal distension and hepatic involvement with elevated liver enzymes and prolonged coagulation times. new bone compromise was documented. inflammatory bowel disease, neoplastic or chronic infectious disease involvement was ruled out. during the stay in our institution no microbiological isolation was documented. skin, colon and bone tissue biopsies were performed and extra-institutionally performed liver biopsy were examined, showing as a single common finding leukocytoclastic vasculitis in all tissues. given the heterogeneous nature of the condition, the diagnostic possibility of an immune dysregulation disorder was considered and a therapeutic trial with nsaids and prednisolone at mg/kg/day was started, as well as genetic studies by exome sequencing. the exome results documented a novel mutation in the pik cd gene [c. t> a (p.phe tyr)] as probably pathogenic. the patient has presented a clinical improvement and a significant decrease in inflammation markers. at the moment, we are waiting for the performance of functional tests to define the definitive therapy for this patient. conclusions. this case description highlights the diagnostic difficulties that face in developing countries, where the nonavailability of functional testing has implications on the diagnosis opportunity and establishment of optimal therapeutic for patients with complex diseases such as primary immune regulatory disorders abstract/case report text background: autoinflammatory syndromes, a wide family of diseases, defined as attacks of inflammation that are unprovoked (or triggered by a minor event) and are primarily related to dysregulation of the innate immune system. periodic/recurrent fever syndromes were the former name of these diseases. however, only in two conditions: cyclic neutropenia (cn) and periodic fever, aphthous stomatitis, pharyngitis, and adenitis (pfapa) are febrile episodes truly periodic. for pfapa, although diagnostic criteria differ and there is no consensus research definition, patients are usually not difficult to recognize based on clinical course and presentation . high index of suspicion and understanding the parental experiences and descriptions of febrile episodes is imperative in facilitating early recognition and timely diagnosis. aims: to standardize and summarized the clinical presentation of pfapa, based on parental descriptions and providers observation of febrile episodes. methods: utilizing a query for the icd- diagnosis code m . ( + ) we identified a cohort of children diagnosed and managed for periodic fever, excluding those with monogenetic mutation (e.g. blau, majeed) and those with chronic illness. we reviewed the charts for documented parental report and provider observation of febrile episodes. standardized signs and symptoms were recorded for each patient [ table ]. results: a cohort of children, boys ( %) and girls ( %) with documented, cyclic episodes of fever > , was identified. the average age at diagnosis was . +/- . years. classic symptoms were reported or observed in % of patients ( ). more than half ( %, patients) had documentations of other symptoms, usually reported by parents to occur sporadically during some fever episodes. decreased oral intake and general "ill appearance" was reported by parents in % of patients. when reporting the time intervals, parents usually reported similar length for each episode, typically between - days, and regular interludes, typically between - weeks. the findings are summarized in table . discussion: the findings presented here are in concordance with previously published data describing pfapa as a syndrome affecting young, generally healthy children with identical episodes of fever lasting for a few days, recur with regularity. our data support the approach that parental observation is fundamental in identifying the unique pattern of illnesses. engaging the parents with directed interview is crucial to establish this clinical diagnosis in a timely fashion, prevent misdiagnoses of future febrile episodes as presumptive infections, and avert unnecessary antibiotics courses. this cohort adds the observation that up to % of patients who display this identifiable pattern of illnesses, do not present with aphthous stomatitis, pharyngitis, or adenitis. these classic symptoms although common, are not the rule. the cyclic chronicity of febrile episodes associated with general ill appearance (but not lethargy), and decreased po, in an otherwise healthy child is the clinical gold-standard of this condition. abstract/case report text heterozygous mutations in nfkb are frequently identified among immunodeficient patients with highly variable clinical symptoms. in a world-wide collaborative effort, we characterized the clinical and cellular phenotype and the management of of these patients harboring distinct nfkb variants. nfkb encodes the transcription factor precursor p which is processed to p (canonical pathway). known pathogenic variants cause p haploinsufficiency (due to protein decay) or p -skipping (with expression of p -like forms). most variants however are single amino acid changes with yet unknown effects. all sequence changes were assessed in silico for their probability of pathogenicity including variants which were additionally tested in vitro. these analyses include the sub-cellular protein localization (microscopy), protein expression, stability and processing (western blotting), transcription factor activity (reporter assay) and dna-binding ( figure ); the associated unadjusted odds ratio (or) was not significant, suggesting igrt had restored cases to a similar baseline infection risk as the controls. in a multivariate conditional logistic regression model adjusting for the significantly higher occurrence of risk factors in cases compared with controls in the preindex period, cases were associated with a % lower adjusted odds of major/severe infections in the post-index period versus controls (or= . ; p= . ). conclusions: patients who required treatment with igrt (privigen®/ hizentra®) had previously experienced more major/severe bacterial infections than those not needing treatment with igrt. however, igrt was associated with a reduction in the risk-adjusted odds of major/severe bacterial infections compared with non-igrt treated sid patients with haematological malignancies. abstract/case report text introduction: real-world data are lacking as far as identifying patients with secondary immunodeficiency (sid)/hypogammaglobulinemia who may benefit most from interventions to protect them from potentially fatal infections. this study aimed to identify risk factors for major/severe infections in patients with sid with underlying haematological malignancies. methods: a retrospective database analysis was conducted using the iqvia real-world data adjudicated claims -us database (study period: january -september ). inclusion criteria were adults newly diagnosed with sid (first diagnosis termed the index date), with ≥ months continuous health plan enrolment pre-index (baseline period) and a minimum of months' data post-index (mean: days), with chronic lymphocytic leukaemia, multiple myeloma and/or non-hodgkin's lymphoma and without claims for any ig therapy in the -month baseline period. patient characteristics in the -month baseline period were assessed. over the post-index period, antibiotic/antiviral use and frequency of infections were assessed. the frequency of major/severe infections was determined using diagnosis codes for bacterial, viral, fungal, parasitic, other or unspecified causal pathogen infections. major/severe infections were defined as those requiring inpatient hospitalisation with an infection diagnosis code and/or use of intravenous (iv) antibiotics or iv antivirals in an outpatient setting. a multivariate cox proportional hazards (ph) model evaluated baseline patient characteristics associated with risk of major/severe infections post-index. results: a total of , patients met the inclusion criteria. the mean age of patients was years and . % were male. in the -month baseline period: . % of patients received cancer treatments and . % of patients received antibiotics ( . % iv antibiotics). a total of . % of patients experienced any infection, . % experienced ≥ infections and . % experienced major/ severe infections. the mean number of infections over the baseline months was . for any infection (at the unique diagnosis code level) and . for major/severe infections (unique hospitalisations with any infection diagnosis code and/or unique days with an outpatient iv antibiotic or iv antiviral). in the post-sid diagnosis period, . % of patients had major/severe infections; of the major/ severe infections, . % were identified as bacterial, . % were viral, . % were fungal, while . % did not have a causal pathogen specified. a total of . % of patients experienced one severe/ major infection and . % experienced ≥ severe/major infections ( figure ). the mean annualised number of major/severe infections post-index was . . receiver operating characteristic (roc) curve analysis to optimise sensitivity versus false positives in identifying those at risk of major/severe infections post-index identified a cutoff point of three bacterial infections in the baseline pre-index period as a potential optimal trigger to consider treatment to avoid major/severe infections post-index ( figure ). the multivariate cox ph analysis suggested that hospitalisations, infections (≥ ), or antibiotic use in the -months pre-index (prior to sid diagnosis) were predictive of major/severe infections post-index (post-sid diagnosis) (all p < . ). conclusion: infections are common in patients with haematological malignancies and sid. key baseline predictors for major/severe infections in patients with an sid diagnosis were a history of infections, hospitalisations or antibiotic use. unfortunately, ms/ms detection is limited by the extremely low (e.g., pmol/l) protein concentrations in blood cells. peptide immunoaffinity enrichment coupled to selected reaction monitoring (immuno-srm) is a robust method for quantification of low abundance proteins in complex matrices, including dried blood spots (dbs). in a study of patients, immuno-srm reliably identified wiskott-aldrich syndrome (was) and x-linked agammaglobulinemia (xla) patients using direct quantification of proteins responsible for disease (front. immunol., ). we further expanded our approach for x-linked chronic granulomatous disease (x-cgd), ada and dock deficiency. marker proteins representing platelets, nk cells, and t-cells have also been analyzed to provide additional information about disease processes. these results demonstrate the utilization of immuno-srm as a sensitive platform for multiplexed signature peptide quantification and its potential for pidd newborn screening and clinical diagnosis from dbs. methods: candidate peptides were selected based on ms/ms sensitivity and uniqueness in the proteome. anti-peptide monoclonal antibodies (mabs) were then generated for peptide enrichment from dbs. blood from normal controls, xla, was, xl-cgd, dock and ada deficiency patients was collected after consent on filter paper, dried, and stored at - °c. proteins were extracted from dbs, digested with trypsin, and enriched using mabs bound to magnetic beads. the enriched peptides were then eluted and analyzed with a waters xevo tq-xs. results: a multiplexed immuno-srm panel has been generated for screening eight signature peptides representing five pidd-specific and three cell-type specific proteins from dbs. limits of detection and quantification were femtomoles of peptide, the assay showed a broad linear range, and intra-assay and inter-assay coefficients of variation were < %. in samples from xla, was, xl-cgd, dock and ada deficiency patients, signature peptides are significantly reduced relative to normal controls and patient identification had excellent agreement with clinical and molecular diagnosis. also included in the multiplex panel are cell specific markers for platelets (cd ), t-cells (cd ɛ), and nk cells (cd ). diagnostic cutoffs for each peptide concentration have been established. in was patients, cd levels were significantly reduced consistent with characteristic thrombocytopenia. immuno-srm also has the ability demonstrate the effects of pidd treatment. a was patient analyzed before and after bone marrow transplant showed normalized was protein and cd after treatment. two ada deficiency patients showed normal levels of ada enzyme after rbc transfusion. finally, a high-throughput (ht) immuno-srm method screens pidd-specific peptides in a . -minute runtime meeting high volume nbs workflow requirements. this ht method returned identical results to the standard immuno-srm pidd panel. conclusions: the data herein demonstrate the feasibility of using immuno-srm as a broad clinical diagnostic for identifying and studying pidd patients from easily collected and shipped dbs. significantly, ht immuno-srm workflows represent a promising potential option for nbs of pidds and other congenital disorders. chief, laboratory of clinical immunology and microbiology/national institute of allergy and infectious diseases, niaid/national institutes of health, nih abstract/case report text we have previously used the artificial thymic organoid (ato) system, based on the d aggregation and culture of a delta-like canonical notch ligand -expressing stromal cell line (ms -dll ) with cd + cells, to study t cell differentiation from cd + cells obtained from patients carrying defects that are intrinsic to hematopoietic cells (rag - , ak , il rg) or that affect thymus development (digeorge syndrome). we now report results of in vitro t cell differentiation of cd + cells obtained from patients with either haploinsufficiency or dominant negative (dn) mutations of the ikzf gene. ikzf is an essential transcription factor expressed throughout hematopoiesis and involved in both lymphocyte and myeloid differentiation. heterozygous germline mutations in ikzf give rise to distinct clinical phenotypes, depending on the nature of the mutation. in particular patients with ikzf haploinsufficiency present with common variable immunodeficiency (cvid) associated with b cell immune deficiency, b-all susceptibility, and autoimmune manifestations. no clinical t cell defects are evident among these patients, except for elevated naive and central memory cd +cd + t cells. in contrast, patients carrying dn ikzf mutations present with combined immunodeficiency (cid) characterized by the presence of an increased proportion of naïve t cells, associated with defective generation of memory t cells, impaired t cell activation, signaling and proliferation, reduced t-helper (th) polarization, and susceptibility to pneumocystis pneumonia. different mouse models of ikzf mutations have been developed, however their phenotype does not fully match what reported in patients, and in some models indicates a more severe defect in t cell development. to address these controversies and to gain novel insights into the effects of distinct ikzf mutations on human t cell development, we used the ato system to analyze progression of t cell development from cd + cells obtained from one patient with ikzf haploinsufficiency and one patient with dn ikzf mutation. both patients showed a similar early block in t-cell differentiation a t p re -t c el l s ta g e. ho w ev e r, th e p a ti e nt wi t h i kz f haploinsufficiency showed a more pronounced leakiness, with a residual production of cd +tcrab+ cells, which could account for the milder t-cell phenotype presented in this type of patients. interestingly, the dn patient presented an increased accumulation of cd -cd b-cd aa+ cells. these results show an unexpected role for ikzf in humans in early stages of t-cell differentiation and indicate ikzf as a necessary factor for the induction of cd b expression in t cells. abstract/case report text background: pediatric acute liver failure without an identifiable cause (indeterminate palf/ipalf) is associated with increased rates of liver transplant and mortality. aplastic anemia (aa) may develop weeks after the diagnosis. the immunologic mechanisms that contribute to disease pathogenesis have not been clearly elucidated. we report detailed immunophenotyping of a patient with ipalf/aa. case: a previously-healthy -year-old male was admitted for acute hepatitis presenting with jaundice and hepatosplenomegaly. evaluation for infectious, toxic, metabolic, autoimmune, and rheumatologic disorders was negative. he was pan-lymphopenic (cd +alc cells/μl) with an inverted cd :cd ratio of . on admission. liver biopsy showed severe portal, interface, and lobular inflammation characterized by activated sinusoidal macrophages and perforinexpressing cd +t-cells. compared to a healthy control, the percentage and number of peripheral blood cd +t-cells expressing perforin ( %v. %, v. cells/μl) and granzyme-a/b ( %v. %, v. cells/μl) was also increased, while percentages of perforin+ ( %) and granzyme-a/b+( %) nk cells were normal. bone marrow (bm) showed % cellularity with rare hemophagocytosis. serum cytokine analysis demonstrated il- pg/ml, il- -binding-protein pg/ml, cxcl pg/ml, and sil- rα u/ml, consistent with smoldering hemophagocytic lymphohistiocytosis (hlh), but he did not meet hlh diagnostic criteria. genetic sequencing did not identify pathogenic variants in genes associated with primary immunodeficiencies. he was diagnosed with ipalf and treated with three doses of anakinra and two weeks of ruxolitinib, followed by prednisone - mg/kg/day and intravenous immunoglobulin g/kg/month. immunophenotyping performed after two months of therapy showed persistent inversion of the cd :cd ratio with small expansions of cd +cd -cd -cells and tcr··+t-cells. in the cd +t-cell subset, there was a substantial paucity of naïve cells, with effector memory t cells (tem) being more abundant than central memory t cells (tcm). in the cd +t-cell subset, the majority of cd ro+cells were tem with no detectable tcm, and % of all cd +t-cells were cd +temra. in both cd + and cd +t-cell subsets, activated (hla-dr+) and senescent (cd +) subpopulations were increased, and the majority of cells expressed the exhaustion marker pd- . the hepatic inflammatory infiltrate similarly reflected repetitive antigenic stimulation, with expansion of cd +cd +t-cells. quantitative immunoglobulins and total memory b-cells and plasmablasts (cd +cd +) were normal for age. however, there were no circulating iga-memory b-cells and a reduced number of iggswitched memory b-cells (table ) . given the severity of his phenotype and bm hypocellularity ( %), allogeneic hct was performed using a matched-related-donor ( / ) with conditioning of flu+cy+alemtuzumab. at d+ , he shows improved liver function but persistent pancytopenia, with transfusion-dependence for platelets. discussion: to our knowledge, this is the first description of detailed immunophenotyping in blood from a patient with ipalf/ aa. other studies have identified distinguishing hepatic infiltrates and cytokine/chemokine profiles that suggest excessive activation of cytotoxic t-lymphocytes and macrophages contribute to disease pathogenesis (alonso et al, ). our preliminary data supports this hypothesis and expands the spectrum of immune dysregulation in the t and b cell compartments, proposing a primary immune etiology. immune dysregulation may be concordant with hyperinflammation and cytokine storm, the latter offering potential therapeutic targets. early diagnosis and treatment of immune dysregulation may prevent development of aa. background: x-linked agammaglobulinemia (xla) is one of the first inborn errors of immunity identified, with thousands of patients described to date. infections originally dominated the clinical phenotype, but early diagnosis and immunoglobulin replacement allowed for long term survival as well as recognition of late-onset complications. nodular regenerative hyperplasia (nrh) of the liver is a silent cause of non-cirrhotic portal hypertension. nrh underlying pathophysiology remains blurry and the disease has no specific treatment. nrh has been increasingly reported in primary immunodeficiency but data in xla are very limited. objectives: to assess and characterize nrh in patients with xla. methods: we retrospectively reviewed the medical records of all xla patients referred to the nih between and . hepatology evaluation and liver biopsies were performed when clinically indicated. patients were stratified into nrh+ or nrhgroups, according to their nrh biopsy status (patients with no liver biopsies were classified as unknown). laboratory values are presented as medians. fisher's exact test and mann-whitney test were used to compare categorical and continuous variables, respectively. results: twenty-one xla patient records were reviewed, with a median age at start of follow-up (f/u) of y and a median duration of f/u of years. eight patients underwent at least one liver biopsy of whom ( % of nih xla cohort) were nrh+. the median age at nrh diagnosis was y ( - ). among patients who had liver biopsies, alanine aminotransferase (alt) levels were mildly elevated in all, while alkaline phosphatase (alp) levels were only increased in nrh+ patients (p= . ). both nrh+ and nrhgroups had similar aspartate aminotransferase (ast) levels at baseline but higher values were observed at the end of f/u in the nrh+ group ( vs. u/l, p= . ). persistently low platelet count ( < k/μl for more than months), mildly to highly elevated hepatic venous pressure gradient (hvpg) and either hepatomegaly and/ or splenomegaly were present in all nrh+ patients. in opposition, neither persistently low platelet counts, nor hepato-or splenomegaly were present in the two nrh-patients evaluated. hvpg was normal in the only nrh-patient tested. all-cause mortality was higher among nrh+ patients ( / , %) than in the rest of the cohort ( / , % among nrh-and unknown patients, p= . ). conclusions: based on our retrospective analysis, nrh appears as an underreported, frequent and severe late-onset complication in xla, which is highly associated with increased mortality. persistent thrombocytopenia, elevated alp, elevated hvpg, hepato-and/or splenomegaly were common in liver biopsyproven xla/nrh+ patients and distinguish them from xla/ nrh-patients. based on nrh prevalence, severity, lack of specific treatment and poor outcome in xla, immune-reconstitution (rather than igg replacement and infectious prophylaxis) should be considered early in this population in order to prevent fatal long term complications. abstract/case report text introduction: barth syndrome (bths) is an x-linked recessive disorder caused by a mutation in the tafazzin (taz) gene resulting in an inborn error of cardiolipin phospholipid metabolism (an important mitochondrial inner membrane lipid). it is commonly characterized by intermittent neutropenia and cardiac and skeletal myopathies. we present a case of bths with associated lymphopenia and hypogammaglobulinemia, which has not been previously described in the literature. case report: a two-month old male, born full term with normal newborn screening, was first admitted for rsv bronchiolitis. at this time, patient underwent an echocardiogram given his older brother with hydrops had died hours after birth and on autopsy was found to have dilated cardiomyopathy (dcm). patient was similarly noted to have dcm and thus had whole exome sequencing done that showed a hemizygous mutation in the taz gene (c. g>a). this novel variant resulted in early termination of the protein (p.trp ter) with concern for loss of function. in regard to patient's first year of life, he had frequent uri symptoms, episodes of acute otitis media requiring tympanostomy tubes, but no documented pneumonias or other serious bacterial infections. patient also had gross developmental delay, particularly motor, and feeding difficulties with persistent failure to thrive requiring g tube placement. his absolute neutrophil count ranged from - cells/mm in the first year. at age months, patient was found to be in acute decompensated heart failure with concern for myocarditis (ck , u/l, troponin i . ng/ml) as well as acute hypoxic respiratory failure with respiratory cultures growing pseudomonas. he was incidentally found to have an igg level of mg/dl (normal for age - ) and treated empirically with ivig. when seen by immunology, further workup showed persistent b cell lymphopenia (absolute cd of - /mm ). he also had a low initial nk cell count ( - /mm , later normal) with normal cd and cd t cell counts. tetanus and hib titers could not be assessed as he had recently received ivig. his igg trended up to mg/dl a few days after initial ivig and then subsequently dropped to mg/ dl, with a level of mg/dl two weeks following initial dose. workup for gastrointestinal or renal losses of immunoglobulin were negative. he also shortly after developed enterobacter bacteremia. his igg levels at this time continue to remain around mg/dl. he subsequently required a heart transplant at age months for his dcm. after transplant, he continued to improve from a cardiac standpoint, but his lymphopenia persisted and each time he was weaned off ivig, his hypogammaglobulinemia persisted at - mg/dl thus requiring additional ivig replacement over the course of the next months. the remainder of immunoglobulins were normal initially, but the igm slowly dropped over time to - . mg/dl. patient was started on weekly subcutaneous immunoglobulin replacement at months, doing well clinically at age -month follow up. conclusion: here we present a patient with bths, with a novel variant, who had b-cell lymphopenia as part of his presentation with persistent hypogammaglobulinemia requiring ivig replacement. year fellow/ucla associate professor/division of allergy, immunology, and rheumatology, university of california los angeles chief of pediatric allergy and immunology/harbor-ucla chief, laboratory of clinical immunology and microbiology/national institute of allergy and infectious diseases, niaid/national institutes of health, nih biologist/laboratory of clinical immunology and microbiology, division of intramural research, national institute of allergy and infectious diseases, national institutes of health project scientist/ucla abstract/case report text a -year-old female presented for combined immunodeficiency. at years of age she was diagnosed with rag hypomorphism and started on ivig. as a child, she was hospitalized for pneumonias and cryptococcal meningitis. she suffered sinusitis, hepatitis, tooth abscess, cmv and herpes stomatitis. later, she experienced recurrent cutaneous abscesses, utis, vaginal yeast infections, and hidradenitis. she twice hospitalized recently for pneumonias and diagnosed with mycobacterium abscessus on bronchoscopy. she suffers onychomycosis, osteomyelitis and oral and esophageal candidiasis with odynophagia. on exam, she had white plaques on tongue and buccal mucosa. she had hyperpigmented plaques on forehead and cheeks and thickened nails. immune evaluation was significant for lymphopenia with alc and thrombocytopenia with platelets k. b cells were nearly absent ( absolute count) and nk cells were low at absolute count. ige was absent, igm mg/dl, iga mg/dl and igg mg/dl (on replacement). her total cd + count was , cd + t cells were low at %, but cd + cells normal at %. the cd + t cells were mostly memory phenotype, which probably reflects lymphopenia-induced proliferation of a small number of clones. her cd + t cells also had an elevated amount of memory cells for age, but still had presence of naive cd + t cells. as expected with perpetual lymphopeniainduced proliferation, there was evidence of terminal memory (temra) in the cd + lineage. proliferative responses of t cells were modest. cd + t cells did respond to pokeweed, but less to pha and cona. there were no antigen specific responses. trecs were normal. esr was mildly elevated at . of note, her liver enzymes were elevated with alkaline phosphatase and ast , presumably secondary to prolonged fluconazole use. w es r e v e a l e d a k n o w n p a t h o g e n i c v ar i a n t i n s tat (nm_ . : c. c>g (p.n k)) as well as a heterozygous variant in rag p.m t. the stat mutation is de novo and was previously published as a gain of function mutation. however, when we performed validation studies to evaluate cd + cells with stimulation to ifna, the patient had decreased pstat as compared with control. va . analysis was performed to evaluate rag defect and showed % of t cells with va . expression confirming that the rag defect is not clinically significant. she developed severe thrombocytopenia refractory to platelet transfusions and ivig. she was started on ruxolitinib which improved platelet counts. however, she presented with shortness of breath, persistent tachycardia and was found to have cmv carditis and hepatitis significant for echocardiogram with ef %. cmv pcr is improving with last check iu/ml after month of therapy with ganciclovir. we now are looking for evidence of socs to explain the decreased stat phosphorylation. genetic testing is critical when evaluating a patient with immunodeficiency. our patient demonstrates that genetic mutations cannot be taken at face value and should be evaluated and validated fully to optimize patient care. fellow/university at buffalo / oishei children's hospital abstract/case report text opportunistic infections (oi) are commonly seen in patients undergoing hematopoietic cell transplantation (hct). different strategies for antimicrobial prophylaxis are often employed in the transplant setting to reduce the likelihood of encountering infection. the predisposing risks for infections include the expected neutropenia and lymphopenia following conditioning, prolonged defects in cell-mediated and humoral immunity during the engraftment period, and iatrogenic immunosuppression by medications for graft versus host disease (gvhd). we report the case of a -year-old male with acute lymphoblastic leukemia, which relapsed to chronic myelogenous leukemic blast crisis, and failed a subsequent allogeneic hct with central nervous system relapse. he was subjected to a second allogeneic hct. his immediate post-second transplant course was complicated with skin and gut gvhd, and infection and/or reactivation of coronavirus, respiratory adenovirus, epstein-barr virus, and human herpesvirus . while the herpesviral infections were controlled with antivirals and rituximab, adenovirus c infection proceeded to involve the gastrointestinal tract, and proved persistent over several months despite use of cidofovir. the patient's gvhd and transplant-associated thrombotic microangiopathy necessitated use of further immunosuppressants, including the complement protein c -binding eculizumab (an inhibitor of formation of the terminal c b- complex), ruxolitinib (a janus kinase [jak] / inhibitor) and low-dose interleukin- . h i s c l i n i c a l c o u r s e w a s f u r t h e r c o m p l i c a t e d b y stenotrophomonas maltophilia gut colonization and subsequent bacteremia, as well as multiple gram-positive bacteremia courses. at around day + , there was a life-threatening pericarditis with pericardial effusion and respiratory distress, associated with pneumocystis and stenotrophomonas being isolated from bronchoalveolar lavage. this occurred despite the patient being on pentamidine prophylaxis. the patient eventually recovered on trimethoprim/sulfamethoxazole therapy. we discuss the various risk factors potentially contributing to each oi in this illustrative case. in particular, complement and jak inhibitor therapy are fairly new drugs approved for other indications, whose off-label use in transplant patients is increasing. both have recently been associated with certain oi in the literature, as they are in this patient. abstract/case report text background: autoimmune lymphoproliferative syndrome (alps) is characterized by chronic nonmalignant lymphadenopathy, splenomegaly, hepatomegaly, cytopenias, and other autoimmune manifestations. typically, the biomarker profile of patients with alps includes elevated tcr αβ+ dnt cells, serum igg, serum b , serum il and soluble fas ligand (sfasl). hdl cholesterol can also be significantly low. alps is caused by lymphocyte accumulation due to defects in the fas-mediated apoptosis signaling pathway. these defects cause resistance to physiological apoptosis in lymphocyte populations that results in chronic lymphoproliferation. the molecular defect underlying most alps etiologies is attributed to heterozygous germline or somatic (limited to dnt cell subpopulation) pathogenic single nucleotide variants (snv) in fas. we describe copy number variants (cnvs) at the fas locus underlying alps in unrelated families. methods: through the centralized sequencing initiative at at the national institute of allergy and infectious diseases (niaid), patients undergo genomic workup to identify molecular defects contributing to clinical phenotypes of immune system disorders. all patients receive exome sequencing and a subset of patients also receive array-cgh analysis. patients and results: we performed exome sequencing on patients with a clinical diagnosis of alps. for patients with no molecular defect through exome, we performed cnv analysis. in this cohort, we identified three patients with a copy number variant involving the fas locus. all patients presented with splenomegaly and lymphadenopathy in childhood with ages of onset ranging from months to years old. all patients experienced anemia, autoimmune neutropenia, and thrombocytopenia. they had biomarker evidence showing elevated serum b levels, sfasl levels, and elevated αβ+dnt cell populations. they were found to have very low hdl cholesterol in early childhood ranging from - mg/dl ( - mg/dl). all patients had negative family histories for lymphoproliferative disorders and immunodeficiency. these patients had clinical presentations and biomarker profiles similar to alps patients with germline and somatic fas variants. patient : we detected a~ . mb copy number loss encompassing all of fas. parental studies were not performed. patient : we detected a~ . mb copy number loss encompassing all of fas. parental studies showed this to be maternally inherited. in addition, prior karyotype testing of the bone marrow showed the same deletion. patient : we detected a~ . mb copy number loss encompassing exons - of fas. parental studies were not performed. these results are consistent with the pathogenic nature of copy number variant losses involving fas. the mechanism of disease in these patients is consistent with haploinsufficiency. in family , the mother harboring the fas deletion is unaffected. this is consistent with prior observation of reduced penetrance within a family in alps. conclusion: these three cases harbored causative deletions in fas in the presence of biomarkers indicative of alps and negative results for germline and somatic genetic variant testing. these patients demonstrate that copy number variant analysis should be pursued if there is robust clinical and biomarker evidence of alps as it can lead to a molecular diagnosis and appropriate treatment when exome or next generation panel based fas sequencing is inconclusive. abstract/case report text rationale: the thymus is essential for the development of tcells. patients with thymoma have decreased aire expression and have an abnormal thymic microenvironment where the negative selection of t-cells is compromised, resulting in a broad spectrum of autoimmune-mediated diseases. besides myasthenia gravis, which is found in to % of patients with thymoma, other autoimmune diseases have been reported including erythroblastopenia, systemic lupus erythematosus, inflammatory myopathies, thyroid disorders and good's syndrome. recent studies have described additional autoimmune conditions such as pneumonitis in thymoma patients. we identified a patient who developed chronic cough post-thymectomy and was found to have lymphocytic pneumonitis with associated autoantibodies against lung antigen kcnrg and lung immunopathology consistent with apeced pneumonitis, which implies a common pathogenic mechanism between these conditions. methods: we describe a patient with thymoma who developed autoimmune pneumonitis associated with kcnrg autoantibodies and a characteristic pattern of immunopathology recently described in patients with monogenic disorder caused by primary aire deficiency (apeced) and secondary aire deficiencies (thymoma, rag deficiency). results: patient is a -year-old male with no significant past medical history who was in good state of health until age when he was diagnosed with and received treatment for guttate psoriasis (resolved with uv therapy) and alopecia areata. at age , he developed severe abdominal pain and weight loss. he had an abdominal ct performed that showed chronic pancreatitis and thymoma. one month later, the patient underwent thymectomy and subsequently, underwent ercp and pancreas biopsy, revealing atrophic pancreatitis with negative staining for lgg and lgg . at that time, he was started on pancreatic enzymes with improvement of abdominal symptoms. following thymectomy, he developed persistent dry cough and recurrent symptoms of sinusitis which did not respond to several courses of oral antibiotics to treat his positive culture for pseudomonas. he had a negative work up for vocal cord dysfunction and cystic fibrosis, and negative autoantibodies against ifn-gamma, il- a, and gm-csf. for work up of chronic cough, the patient underwent ct imaging of the chest which revealed diffuse peri-bronchial thickening, mucus plugging, and tree-in-bud nodularity through most of his lungs. he underwent bronchoscopy with bal which revealed n o r m a l b r o n c h i a l m u c o s a a n d a i r w a y n e u t r o p h i l i a . endobronchial biopsies showed basement membrane thickening and dramatic lymphocyte infiltration in intraepithelial and submucosal areas. his bal cultures revealed mycobacterium intracellulare/chimaera. patient was also tested for autoantibodies against lung-specific bactericidal/permeabilityincreasing fold-containing b (bpifb ) and the potassium channel regulator kcnrg that have been associated with the development of pneumonitis in patient with apeced, thymoma and rag deficiency, and was found to have kcnrg-targeted autoantibodies. conclusions: thymoma is a disease associated with secondary aire deficiency. this case illustrates common clinical, radiographic, histological, and autoantibody features in thymomaassociated and apeced-associated pneumonitis, indicating that disorders with primary and secondary aire deficiencies may have common pathogenetic mechanisms. bpifb and kcnrg should be included in the autoantibody profile testing of patients with thymoma and lung disease. immune suppression and antimycobacterial antibiotic treatment are planned. abstract/case report text introduction/background: activated phosphoinositide -kinase δ (pi kδ) syndrome (apds) is a primary immunodeficiency caused by a gain-of-function mutation in the pik cd gene that encodes the p δ catalytic subunit of pi kδ. it is characterized by recurrent respiratory tract infections, lymphoproliferation, nodular mucosal lymphoid hyperplasia, enteropathy, ebv and/or cmv infection, reduced t cell function and high levels of igm. there is not evidence of this disease in peruvian patients. methods: a case series of two pediatric patients with apds. results: the first patient is a girl of non-consanguineous parents. family history shows four maternal uncles died at pediatric ages with unknown diagnosis. at the age of , she presented lymphadenopathy and fever being treated as cat scratch disease without improvement of symptoms. months later, she was hospitalized due to anemia, mild hepatosplenomegaly, ascites and chronic diarrhea and diagnosed with gastrointestinal tuberculosis (tb). a hepatic biopsy only showed reactive hepatitis. however, the patient did not improve her symptoms despite anti tb treatment. years later, she was hospitalized for lymphadenopathy, pancytopenia, chronic diarrhea, ascites and severe hepatosplenomegaly. cmv igg was positive and lymph node biopsy revealed paracortical and follicular lymphoid hyperplasia due to ebv infection without neoplastic proliferation. low cd + t and cd + b cells and high igg levels were found (table ) . at this time, it was suggested the diagnosis of apds which was confirmed by next generation sequencing (ngs) identifying a heterozygous mutation in the pik cd gene (c. g>a, p.glu lys). she was treated with sirolimus and ivig for years. the symptoms persisted despite treatment and died at the age of . the second patient is a -year-old girl also of non-consanguineous parents. family history includes eczema (father) and colorectal cancer (mother). she has had recurrent respiratory infections, chronic diarrhea and poor weight gain since months old receiving symptomatic treatment only. at the age of , she was hospitalized for persistent pneumonia ( p s e u d o m o n a s p o s i t i v e ) , l y m p h a d e n o p a t h y a n d m i l d hepatosplenomegaly. a ct scan showed bilateral bronchiectasis and the sweat chloride test was negative. based on this, a diagnosis of cystic fibrosis was made and treatment was started. however, a genetic study only showed heterozygous mutations in the cftr gene (g d and g x). year later, she presented a neck-located skin abscess. at the age of , she was hospitalized for complicated pneumonia, diarrhea, lymphadenopathy, ascites and severe hepatosplenomegaly. multiple polyps in the duodenum and colon with lymphoid hyperplasia were detected, ebv igm and igg were positive and a lymph node biopsy showed paracortical hyperplasia without neoplastic proliferation. cd + t cells and igm levels were increased (table ) . a diagnosis of apds was suspected and ivig was started. ngs showed the same mutation as the first patient (c. g>a, p.glu lys). conclusion: apds should be considered in patients with recurrent respiratory tract infections, lymphoproliferation, enteropathy and abnormal immunologic function without another explanation. ngs is a useful tool to identify these cases in low-income countries. acknowledgments: we thank drs. raif geha and janet chou, division of immunology, boston children's hospital, harvard medical school for the genetic diagnosis. background: granulomatous-lymphocytic interstitial lung disease (glild) is an increasingly recognized pulmonary complication associated with common variable immunodeficiency (cvid) but the natural history and long term prognosis remains poorly defined. imaging findings with computed tomography (ct) are heterogeneous and visual features do not consistently predict a patient's progression to fibrotic lung disease. computer-aided lung informatics for pathology evaluation and rating (caliper) provides an objective analysis of lung parenchymal texture and quantifies the extent of normal lung, along with abnormal features such as honeycombing, reticular/consolidative and groundglass opacity. this may be useful in cvid patients to monitor changes in character or extent of disease and may facilitate early intervention before the disease becomes more aggressive or advanced. case description: our patient is a -year-old non-smoking female with cvid who has been followed for her cvid and associated interstitial lung disease. for more than twenty years, she has had varying abnormalities found on chest ct and these appear consistent with glild. specifically, she has had variable regions of mixed consolidation, ill-defined nodularity and septal thickening. the changing morphology and distribution made assessment of overall severity and extent of fibrosis versus parenchymal infiltration inconsistent. for clinical decision support we used caliper to analyze the current ct ( ) and compared caliper results for previous ct data. caliper provided a comprehensive analysis of the extent and characteristics of parenchymal features, and objectively determined normal and abnormal regions, some of which were not visually apparent. the caliper color overlay was able to highlight subtle regions of ground-glass opacity in areas that visually were regarded as uninvolved lung and quantify the extent of the reticular densities/ consolidation over time. caliper does not differentiate reticulation from consolidation, does not detect nodularity or septal thickening, and ct imaging cannot distinguish inflammation from fibrosis. however, caliper has the power to quantitatively assess overall disease extent and demonstrate subtle abnormalities that would otherwise have been dismissed as normal, given relative sparing compared to other regions. caliper may also provide evidence for disease progression or therapeutic response that is not otherwise radiographically apparent. conclusion: caliper assessment may be a useful tool as an adjunct for a patient with glild to help quantify the extent and character of lung parenchymal involvement. this information may serve as an important guide for clinicians in the assessment of successful management and early intervention to prevent irreversible fibrosis. patients had a trial of fingolimod without any beneficial changes in immune status. both patients receive pneumocystis jirovecii pneumonia prophylaxis with sulfamethoxazole-trimethoprim. conclusions: these results indicate that s pl deficiency due to sgpl mutations is a syndromic primary immunodeficiency leading to profound lymphopenia and hypogammaglobulinemia. our data emphasize the importance of sphingolipid metabolism for an efficient immune response and the need for more studies to delineate the exact mechanisms on how this happens in humans. after d at °c there was a median - % (range - % to + %; p= . ) change in activity. after d - % (range - % to + ; p= . ), d - % (range - % to + %; p < . ), d - % (- % to + %; p < . ) and after d - % (- % to + % p < . ). a °c stability of d was determined from the median percentage reduction; total allowable error adjusted stability data indicated a °c stability of d. samples stored at - °c following repeat freeze thawing saw a freeze/thaw cycle dependent decrease in ch activity. after freeze/thaw cycle there was a median - % (range - % to + %; p= . ) change, cycles - % (range - % to + %; p= . ), cycles - % (range - % to + %; p < . ), cycles - % (range - % to - %; p < . ) and after cycles - % (range - % to - %; p < . ). allowable error adjusted stability data indicated a maximum of freeze/thaw cycles. conclusion: sample storage and handling can have a significant impact on functional complement assessments. room temperature storage should be avoided unless samples will be analysed on the day of collection, °c storage is tolerable providing that assessment is within d; freezing samples at - °c with limited freeze/thaw analysis would be optimal. however, further investigations into longer-term storage at - °c and - °c would be beneficial. conclusions: gi disease is common in cvid affecting % of patients in our cohort. gi+ cvid patients have a higher frequency of autoimmune manifestations than those without gi complaints. the odds of itp, hypothyroidism, and evans syndrome all showed significantly increased odds in the gi+ group. the results of our study may have implications for both gastroenterologist and immunologist. recurrent infections especially those of the sinopulmonary tract are often the trigger for cvid evaluation. autoimmune and gi symptoms however may be the initial presentations of cvid and overlooked until other more recognizable manifestations evolve. the combination of gi issues and autoimmunity especially thrombocytopenia, evans syndrome, and hypothyroidism should include cvid in the gi differential. for the immunologist, a cbc is standard in the work-up of cvid and may reveal autoimmune cytopenia. evaluation for autoimmune disease and in particular hypothyroidism is not. given our findings an initial immune work up specifically for thyroid disease may be indicated. is the transcriptional factor for many cytokines such as il- , responsible for t cell and neutrophil defense again fungal infection. stat mutation leads to defect of neutrophil proliferation and chemotaxis to inflammatory site as well as production of antimicrobial peptides by respiratory epithelial cells. the poor tissue repair in the cavitary lesions and bacterial superinfection in patient's lung created a culture dish for fungal growth and dissemination. traveling to the endemic area and patient's noncompliance to antifungal prophylactic treatment further increased the risk of histoplasmosis infection. pediatric immunologist/john hunter children's hospital abstract/case report text we present the case of a month old boy, the first child to his nonconsanguineous parents of european descent. he first presented at months of age with a cellulitis of his right fourth finger culture positive for staphylococcus aureus which responded to a prolonged course of flucloxacillin. at months he presented with norovirus positive gastroenteritis leading to a brief admission and slow resolution. the first of two severe episodes of oral stomatitis and respiratory distress occurred at months of age. hsv was isolated from the oral lesions and blood culture during that admission was positive for kingella kingae. no cardiac or bone involvement was identified. a more severe episode of oral stomatitis occurred two months later (age months) swab positive for an enterovirus (not typed). due to airway compromise and rapid deterioration he was admitted to the pediatric intensive care unit. again, kingella kingae was cultured from blood cultures with no obvious focal systemic source. the only notable clinical finding was rapid deterioration and, in retrospect, the absence of any significant recorded fever ( < oc). crp elevation was observed (max. mg/l) and neutropenia was found with each of the more severe infectious presentations but recovered in the interval. baseline immunological investigations were normal (lymphocyte subsets, naïve t cell populations, lymphocyte proliferation, serum immunoglobulins and vaccine responses). serial measurement of circulating neutrophils did not identify a cyclical pattern and they were morphologically normal. a panel of genes relevant to primary immunodeficiency (invitae©) revealed a homozygous mutation in irak ((c. c>t (p.gln *)) which leads to a premature stop codon. this is a known pathogenic mutation leading to disease and is most prevalent in the european population (allele frequency (gnomad) = . ). prophylaxis with sulfamethoxazole / trimethoprim and amoxicillin was commenced along with monthly ivig. he has been well since diagnosis with no further severe infectious presentations. functional testing is underway to assess in vitro host viral defence in our patient and potential novel mechanisms relevant to this rare innate immunodeficiency. case studies will be presented on the five cases of fmp that were diagnosed and treated in . potential exposures were identified in four out of five cases: gardening exposure in one case and vaping exposure in three cases. all five were male, age range - . four were gp deficient, and one was p -phox deficient. historically, the vast majority of cases of fmp could be traced to a significant gardening exposure such as lawn mowing or spreading mulch. this was the first year that we saw patients with no identifiable gardening exposure in the setting of significant vaping exposure. with vaping at epidemic levels, especially among teenagers and young adults, it is important to consider that a vaping history is potentially a risk factor for fmp and counseling regarding the potential risks of vaping should be included in infection risk modification for all patients with cgd. abstract/case report text background: granulocyte-macrophage colony-stimulating factor (gm-csf) plays a critical role in macrophage and dendritic cell maturation and host defense against fungus. autoantibodies to gm-csf are associated with susceptibility to cryptococcus and nocardia infections as well as pulmonary alveolar proteinosis (pap) in otherwise healthy individuals. we report a case of a -year-old previously healthy female who presented with cryptococcal meningitis and was found to have autoantibodies against gm-csf. case presentation: weeks prior to admission, our previously healthy -yearold taiwanese female developed a headache associated with tinnitus and visual changes. the headache worsened over the next few weeks and she developed photophobia, phonophobia, and severe nausea/vomiting. at presentation, her exam was notable for papilledema, bilateral cn vi palsy and right foot & left hand paresthesia. mri brain showed ring-enhancing lesions in the anterior frontal lobe, caudate head, and the inferior globus pallidus. she underwent a diagnostic and therapeutic lp. opening pressure was elevated at and csf studies were notable for low glucose, elevated protein, pleiocytosis ( % lymphocytes) and positive cryptococcal antigen. csf culture grew cryptococcus gattii. ct chest revealed a right upper lobe and a left lower lobe nodule. workup: cbc with diff was unremarkable. hiv was negative. lymphocyte subsets were unremarkable with only mildly decreased nk cells, normal immunoglobulin panel including ige, protective titers to tetanus, diphtheria, and ppsv . targeted genetic sequencing did not identify any known mutations in primary immunodeficiency. notably, anti-gmcsf autoantibodies were detected by elisa and were able to neutralize gm-csf phosphorylation of stat detected by flow cytometry. autoantibodies to ifn-γ were not detected. management: patient was initiated on a -week course of liposomal amphotericin b and flucytosine. her csf cultures were cleared of cryptococcus after days of treatment, but her hospital course was complicated by persistently symptomatic intracranial hypertension, worsening pleiocytosis, and elevated cytokine levels in the csf, all of which were consistent with post-infectious inflammatory syndrome (piirs). she received therapeutic lps - x/week until subsequent ventriculoperitoneal shunt placement. concurrently, methylprednisolone was administered for days with a gradual prednisone taper. these interventions led to improvements in her symptoms, including diplopia, and reduction in opening pressures and inflammatory markers in the csf. lifelong fluconazole prophylaxis was recommended. from a pulmonary standpoint, she remained asymptomatic without signs of pap and has had normal pulmonary function tests (normal dlco) and stable chest imaging. conclusion: in otherwise healthy hiv-negative patients presenting with extrapulmonary cryptococcus or nocardia infections, autoantibodies to gm-csf should be suspected and testing for functional autoantibodies to gm-csf (and ifn-γ) should be sent, as genetic testing will not pick up this disease entity. genetic testing should be considered to rule out gata deficiency and x-linked cd l deficiency. idiopathic cd lymphopenia can be ruled out with lymphocyte enumeration. immediate treatment of cryptococcosis is not necessarily different from patients without gm-csf autoantibodies. long-term prophylaxis (fluconazole if presenting with cryptococcus; trimethoprim-sulfamethoxazole if with nocardia) is likely warranted in addition to monitoring for the development of pap. recognizing piirs in patients with cryptococcal meningitis and management with corticosteroids are critical steps. abstract/case report text background: non-infectious complications cause most morbidity and mortality in common variable immunodeficiency (cvid). cvid with complications (cvidc) is defined by elevated t helper (th ) responses attributed to increased circulating microbial products resulting from mucosal iga deficiency. however, complications do not uniformly occur in those with iga deficiency. objective: we tested whether cvidc occurs preferentially in those with hyper-responsiveness to microbial stimuli, manifested by elevated nf-κb-driven cytokines and resultant th responses in cvid patients with increased circulating microbial products. methods: we applied unbiased high-throughput seromics and mass cytometry, cellular and molecular biology approaches, and clinical record review in a subject cvid cohort. results: cvidc was defined by increased nf-κb-driven cytokines that promote th immunity in blood in association with elevated soluble cd , a marker of circulating microbial products, and elevated tnf production by peripheral blood mononuclear cells stimulated with lipopolysaccharide. this cytokine upsurge was associated with mutation of full-length nfkb p gene product ( delt) but not mutations that also involved the nfkb p product involved in transactivation. cytokine elevation corresponded with increased cd +cd -monocytes expressing higher cd and hla-dr and more central and effector memory cd + t cells, t cell chemoattractants, and t cellpredominant tissue pathology. those with granulomatous or neutrophilpredominant, rather than t cell, pathology had the highest tnf. tnf antagonism improved neutrophilic gastritis in cvid with nfkb delt after t cell targeted therapy failed. conclusion: nf-κb dysfunction underlies th immunopathology and tnf-associated innate inflammation in cvidc. both forms of nf-κb immune dysregulation may divergently shape cvid immunopathology. staff clinician/laboratory of clinical immunology and microbiology, immunopathogenesis section, national institute of allergy and immunology, national institutes of health, abstract/case report text introduction: patients with autoantibodies to ifn-γ develop severe and progressive infections with intracellular pathogens, despite aggressive antimicrobial treatment. we describe the use of daratumumab (anti-cd , targeting plasma cells) in a patient with autoantibodies to ifn-γ and progressive disseminated mycobacterium avium infection. she had progressive disease despite treatment with multi-drug antimycobacterials rituximab, and bortezomib. methods: clinical symptoms, total cd /cd , anti-ifn-γ autoantibody titers, and specific imaging were obtained before and after treatment with daratumumab. anti-ifn-γ autoantibody titers were determined by serial -fold dilutions of plasma and measuring anti-ifn-γ autoantibody levels by a particle-based technique as previously described. results: a -year-old filipino woman had progressive disseminated m. avium with extensive bone and soft tissue involvement (calvarium, ribs, bilateral arm soft tissue, paraspinal muscles, bilateral glutei, left inferior pubic ramus, bilateral iliac bones, sacrum, and bilateral humeri) and a tracheo-esophageal fistula. she received bedaquiline, azithromycin, ethambutol, tedizolid, moxifloxacin, clofazimine and meropenem as well as rituximab g once monthly for months. despite these she had progression of clinical and radiographic disease. bortezomib . mg/m twice weekly for weeks was added, but discontinued for ast and alt elevations. rituximab was continued to maintain cd numbers undetectable but clinical and radiographic disease progressed. while on rituximab, total igg level and anti-ifn-γ autoantibody levels decreased from mg/dl to mg/dl and to , respectively. while on bortezomib, total igg levels remained stable ( mg/dl to mg/dl) and anti-ifn-γ autoantibody levels fell slightly ( to ). after starting daratumumab, there was clinical and radiographic improvement, with reduced pain and disappearance of multiple soft tissue lesions. igg levels decreased from mg/dl to mg/dl and anti-ifn-γ autoantibody levels decreased from to . adverse effects of daratumumab were urticaria, pruritus and shortness of breath after the first infusion and aseptic meningitis after the th infusion. conclusions: daratumumab resulted in clinical and radiographic improvement of disseminated m. avium in a patient with rituximab and bortezimib-refractory autoantibodies to ifn-γ. daratumumab is another potentially effective therapeutic agent for anti-ifn-γ autoantibodies. abstract/case report text next-generation sequencing (ngs) is now routinely used as a clinical diagnostic tool. however, regions of high sequence homology continue to be a major challenge for short-read technologies. regions within ikbkg, ncf , sbds, c a, c b, coro a, fcgr a, fcgr b, pms , slfn , slfn , stat b, unc b , and ups are not available by standard ngs. we discuss strategies for analysis of these special regions. we have developed a strategy for supplementing our disease targeted panels which are performed using capture chemistry and a standard reference file. the supplemental method uses gene specific long range amplicon and a special gene specific reference file for alignment. the genes of interest are separated from their homologous counterparts using specific long range amplification primers. multiple amplicons may be pooled together and prepared for sequencing on an illumina miseq instrument using truseq nano dna library prep. bioinformatic analysis proceeds with a custom reference file in which non-specific regions of homology have been removed. this allows reads to be uniquely mapped despite significant homology; a requirement for variant calling. we prepared specific amplicon for several homologous gene targets including the ikbkg gene and the ikbkg pseudogene (ikbkgp ). both amplicons were sequenced in separate reactions and were compared with the standard capture method. variants which are not called in the standard-capture method due to poor mapping scores (non-uniquely mapped reads) are called in the amplicon method. in the capture method, the variants are visualized in the bam as a mixture of gene and pseudogene, while gene and pseudogene variants are clearly separated and identified in the amplicon method. due to high variability in alignment, many homologous regions do not provide reliable copy number variant (cnv) results and must be removed from cnv analysis. however in some situations, we are able to creatively leverage cnv analysis to identify alleles that mis-align to the pseudogene. the pathogenic ncf gt deletion in exon appears to resemble a copy number deletion event when present as reads from one allele mis-align to the ncf b and ncf c pseudogenes. complement genes, c a and c b, share alignment due to their high homology with each other. cnv analysis in normal samples represents four alleles rather than two alleles. cnv events may have a weak signal with no indication of which gene is affected. variant frequencies from the capture and supplemental pcr analysis can be used in tandem with cnv analysis to detect events and may indicate which gene is affected. we plan to include these strategies in our new inborn errors of immunodeficiency gene panel (ieigp) which will enable us to provide a more comprehensive analysis than is currently available. the im diagnosed were inflammatory bowel disease-like (n = , with perianal fistula in / ), mouth ulcers (n = ), discoid lupus (n = ), autoimmune dermatitis (n = ) and eczema (n = ), chronic lung disease (n = ) and granulomas (pulmonary n = ; ocular n = ; bladder n = ; oropharynx n = ). three patients presented more than one site of inflammatory disease. all patients were treated with systemic or topical immunosuppressive or immunomodulatory therapy, most of them corticosteroids. five patients underwent hematopoietic stem cell transplantation (hsct), median age at hsct was years ( - ), and two died month after hsct. conclusions: although infections are more frequent and have a major impact on patient morbidity and mortality, im are increasingly prevalent in patients with cgd. awareness regarding this possible comorbidity is of major importance, since earlier diagnosis and adequate treatment may be crucial for patients survival and quality of life. there is a gap in clinical knowledge regarding associations between specific pid and different rheumatological diseases. in this study, we are reporting the incidence of various rheumatological conditions reported in a large pid population using the usidnet (united states immunodeficiency network) registry. methods: we used the retrospective usidnet registry to conduct the analysis. we included all primary immunodeficiency patients with physician diagnosed rheumatological diseases. results: the total number of pid patients in our query was . ( . %) patients had a diagnosis of rheumatological disease. this cohort included ( . %) female and ( . %) male patients. rheumatologic complications were highest in the interferonopathies ( . %), complement deficiencies ( . %) and autoimmune lymphoproliferative syndrome (alps) ( . %). additionally, disease patterns were noted to be different in each pid. dermatomyositis was found to be the most common rheumatologic condition in patients with x-linked agammaglobulinemia (xla) with a rate of . %, which was remarkably higher than the reported prevalence in the united states ( . %). alps patients had a higher ( . %) numbers of sjogren syndrome diagnoses as compared to the general population ( . - . %). systemic lupus erythematosus was increased in patients with mucocutaneous candidiasis ( . %) as compared to the general population ( . %) and other pids. rheumatoid arthritis (ra) was reported in patients with specific antibody deficiency ( . %), common variable immunodeficiency (cvid) ( . %) and alps ( . %). wiskott-aldrich syndrome patients had the highest numbers of cases diagnosed with vasculitis ( . %). . % of patients with severe combined immunodeficiency (scid) had reported rheumatologic disease. juvenile rheumatoid arthritis (jia) and systemic sclerosis were reported in . % of patients with digeorge syndrome. conclusions: this study reports that higher numbers of rheumatologic diseases are diagnosed in pids compared to the general population. the incidence of different rheumatological disease was variable based on the pid diagnosis. early diagnosis of these diseases is crucial, given the high risk of irreversible complications. limitations of our study include possible selection bias as majority of cases were enrolled from tertiary care centers. abstract/case report text background disorders of immune dysregulation are associated with autoimmune features. this feature could potentially have an impact on the outcome post hematopoietic stem cell transplantation (hsct). hsct, although curative, can be challenging with the underlying immune dysregulation resulting in significant morbidity and mortality. we present the journey through hsct for these children and the factors affecting the outcome. we analysed the data on children up to the age of years diagnosed to have a disorder of immune dysregulation through gene mutation analysis and who underwent hsct at our centre from to . results . xiap mutation a -year-old boy underwent a haploidentical hsct from his father using fludarabine, treosulfan, and gray radiotherapy with post-transplant cyclophosphamide. after initial complete chimerism and cytomegalovirus reactivation responsive to valganciclovir, he developed progressive diarrhoea almost months post-hsct. a rectal biopsy confirmed cmv reactivation and features of inflammation. he has since been treated for the same and is on follow up for inflammatory bowel disease. his chimerism had dropped to % and has remained stable. the second child is a -year-old girl who underwent tcr alpha/beta depleted haplo sct and is months post-hsct, with no features of gvhd or infections, and is doing well with complete chimerism. . il r deficiency three boys aged eight months, one year, and two years of age, diagnosed to have il r deficiency underwent hsct. all three children needed nasogastric tube feeding, parenteral nutrition, and vigilant monitoring for electrolyte disturbances. in the first two children, we had performed tcr alpha/beta depleted pbsc transplants from their haplo matched fathers. the -year-old engrafted by d+ and is doing well two years post hsct with complete chimerism, no gvhd, and infections. his autoimmunity, including recurrent skin scarring, has resolved entirely. the -month-old, however, had primary graft failure and succumbed to his illness. the -year-old boy underwent matched unrelated donor hsct and engrafted by d+ with completed chimerism documented on three occasions. he, however, had secondary graft failure around d+ , and he succumbed to the illness. . lrba deficiency an -month-old girl with lrba deficiency had presented at four months of age with excessive sweating, hepatosplenomegaly, and recurrent chest infections. she was started on monthly intravenous immunoglobulin replacement and abatacept. she received myeloablative conditioning with thiotepa, treosulfan and fludarabine and underwent a matched sibling donor hsct. she engrafted by d+ and has been well ten months post hsct with complete chimerism, no gvhd, and infections. conclusion disorders of immune dysregulation are a heterogeneous group with a varied spectrum of immune dysfunction. myeloablative conditioning is essential, and there is a high risk of cytokine release syndrome and the need for supportive care. the autoimmune features need to be followed for progression in organs other than the hematopoietic system and may require interventions. as long-term data evolves, more precise definitions for patient and donor selection will enable improving outcomes. we performed a retrospective observational analysis of case records of children up to years of age, diagnosed to have variants of scid, and underwent hsct at our centre from to . results . zap deficiency a -month-old girl presented with oral thrush and submandibular cellulitis from one week of life with failure to thrive. she underwent a tcr alpha/beta depleted haploidentical hsct. conditioning included treosulfan/thiotepa/fludarabine/anti-thymocyte globulin. she engrafted by d+ ; now three years post-hsct with complete donor chimerism without gvhd or infections. . orai- mutation a -month-old girl presented with failure to thrive, generalized hypotonia, oral thrush, and recurrent respiratory infections. she underwent haplo-sct with post-transplant cyclophosphamide with pbsc from her haplo-matched father. conditioning included fludarabine/treosulfan. she had cytokine release syndrome grade , which responded to tocilizumab. she had hypertension throughout the peri-engraftment period and had an episode of pres with seizures. her symptoms abated with neutrophil engraftment by d+ . the post-transplant period was complicated by grade skin gvhd and cytomegalovirus reactivation. she has remained disease-free with complete chimerism three years post-hsct. her hypotonia is steadily improving with physiotherapy. . cernunnos-xlf deficiency a -year-old male presented with recurrent infections from years of age, aplastic anemia diagnosed at years of age, subsequent transformation to acute myeloid leukemia at years of age. he had developed multiple fusarium abscesses during the neutropenic period post-chemotherapy for aml. he was referred for a matched sibling sister hsct when in remission. conditioning included fludarabine/treosulfan. he engrafted by d+ with complete chimerism. he developed progressively worsening skin, gut, and liver toxicity secondary to chemotherapy and succumbed to the illness two months post-hsct. . ikzf mutation an -month-old girl presented with failure to thrive, massive splenomegaly, persistent pneumonia, anemia, and thrombocytopenia. she underwent a matched sibling donor pbsc transplant after myeloablative conditioning with thiotepa/treosulfan/fludarabine. she engrafted by d+ , following which all her symptoms abated. she had secondary graft failure two months post-hsct and succumbed to her illness. . mhc class ii deficiency (bare lymphocyte syndrome) three children, aged months, two years, and four years underwent matched sibling donor hsct. myeloablative conditioning with thiotepa/treosulfan/fludarabine resulted in engraftment. the first child died of invasive intestinal aspergillosis days post-hsct. the other two children are well months post-hsct with complete chimerism without gvhd or infections. the two-year-old girl received one cycle of pre-transplant immunosuppression with fludarabine/dexamethasone to prevent graft rejection pre-hsct as she was referred for a second transplant. conclusion children with scid have traditionally been transplanted using reduced intensity (ric) conditioning with immunomodulation. scid variants require myeloablative conditioning with a vigilant follow up for the detection of graft rejection. radiation sensitive scid associated with dna breakage repair defects require ric and close monitoring for gvhd. advances in hsct, including supportive care and haplo-sct, have provided a ray of hope for these hitherto rare conditions. j clin immunol abstract/case report text immune dysregulation, polyendocrinopathy, enteropathy, x-linked (ipex) (omim # ) is a monogenic autoimmune disorder that occurs due to loss of function variation in foxp causing dysfunctional t regulatory cells. although immunosuppression is a mainstay of treatment for autoimmunity, ipex treatment is frequently limited by insufficient response to therapy or side effects of immune suppression. we present a year old male with ipex whose prior immunosuppressive treatment was complicated by inefficacy and medication side effects, requiring a new approach to treat his colitis and erosive dermatitis. he initially presented with infantile diabetes and subsequently developed dermatitis, squamous cell carcinomas, alopecia totalis, and colitis. his clinical diagnosis of ipex was confirmed by foxp sequencing, demonstrating known pathogenic variant c. g>a (p.ala thr). this variant has been described in ipex affected individuals in multiple publications (ref ). his variant affects at the frkhead domain of foxp and has been associated with others with severe psoriasiform dermatisis and alopecia universalis (ref ). his prior immunosuppressive therapies included at different times combinations of corticosteroids, tacrolimus, sirolimus, azathioprine, infliximab, adalimumab, rituximab, dupilumab, and oral mesalamine. the relative efficacy of these agents based on experiences in a cohort of ipex patients was reviewed in (ref ), with the exception of duplimab, which was not listed in that review. for our patient, management of his widespread autoimmunity has been limited by toxicity or lack of efficacy of medications. notably, his dermatitis had no improvement with duplimab, consistent his low total ige and lack of allergic manifestations. at age , after initiation of treatment with sirolimus, he had spontaneous colonic perforation requiring descending colectomy. after stabilization of his colonic perforation, his multi-disciplinary team of allergy-immunology, gastroenterology, and dermatology initiated tofacitinib. tofacitinib is small molecule inhibitor of janus kinase (jak) signaling pathways that mediate cytokine driven autoimmune activation. it is fda approved to treat rheumatoid arthritis, psoriatic arthritis and ulcerative colitis. the decision to use this jak inhibitor was due to its fda approved use for ulcerative colitis, to target our patient's colitis and his other autoimmune manifestations, specifically his dermatitis. its off label for primary immune dyregulatory disorders including candle, stat -gain of function and stat -gain of function disorders has been published (ref ), but thus far its use to treat autoimmunity due to ipex has not been published. he experienced leukopenia while on mg of tofacitinib, which resolved after lowering his dose. currently, he has had improvement in his colitis and dermatitis, and partial improvement in alopecia. he has been on tofacitinib mg daily for months, with only prednisone mg daily as additional immune suppression. as the number and types of selective immune modulators increases, there is continued need to share the experiences of treating physicians of which therapies have been successfully able to decrease disease manifestations with tolerable side effect profiles. we present a year old male with ipex syndrome with severe dermatitis and colitis complicated by colonic perforation despite standard immunosuppressive therapy, who is safely and effectively being treated with tofacitinib. is not frequently associated with autoimmunity, likely due to impaired il- and il- pathways. however, in our relatively large cohort of lof stat patients, we have noted an increased incidence of systemic lupus erythematosus (sle) diagnoses and sle-like symptoms. herein, we characterized the clinical and laboratory features of the patients in our cohort with sle and sle-like disease, with the aim to better understand the pathogenesis by evaluating ifn stimulated genes and neutrophil net formation. methods a retrospective chart review was performed of patients with lof stat to identify those with sle and sle-like presentations, and included clinical features, laboratories including inflammatory markers, auto-antibodies, and complement levels. rt-pcr was performed for interferon stimulated genes (isgs) from neutrophils and pbmcs of lof stat patients with and without sle, and healthy controls. neutrophil net formation was assessed for lof stat patients with and without sle, and healthy controls. results out of a cohort of patients, five patients (ages - ) were identified who carried the diagnosis of sle, and with slelike disease (ages - ). for those with sle, age of presentation was - years, of were female. clinical features included nephritis ( ), alopecia ( ), autoimmune cytopenias ( ), arthritis ( ), discoid rash ( ), and raynaud ( ). all had positive auto-antibodies, and of had low c and/or c . for those with sle-like disease, age of presentation was - , and of were female. clinical features included alopecia ( ), autoimmune cytopenias ( ), raynaud( ), and nephritis ( ). all had positive autoantibodies, and of had low complements. lof stat patients with and without clinical features of sle had increased expression of isgs from both pbmcs and neutrophils. increased spontaneous net formation was observed for lof stat patients both with and without sle symptoms. discussion although autoimmunity is not a common finding in lof stat , we have identified sle or sle-like disease in about % of our cohort, with a high incidence of kidney disease, including one patient who required kidney transplant. the interferon signature and net formation were unexpectedly high in both the patients with and without the sle features. ongoing studies include whole exome sequencing for possible second mutations or modifiers, the role of ige in the kidney disease, and further autoantibody detection. the increased ifn signature raises the question about jak-stat modulation for therapy. chief, genetic immunotherapy section/niaid, nih abstract/case report text chronic granulomatous disease (cgd), a rare immunodeficiency with decreased reactive oxygen species (ros) production, increased susceptibility to infection, and increased mortality is caused by mutations in any one of distinct phagocyte oxidase (phox) components of the nadph oxidase, nox . in the past, identification of the specific protein defect was primarily determined by immunoblotting using specific antibodies to the phox proteins. recently, however, we have shown using fluorescenceactivated cell sorting (facs) analysis of neutrophils in whole blood permeabilized and stained with specific anti-p phox antibody that p phox protein expression was absent in p phox cgd patients and significantly reduced in p phox cgd carriers [kuhns et al. . blood adv. ( ): - ]. these findings demonstrated that determination of phox protein expression by facs analysis provide an alternative to immunoblotting and can aid in the identification of p phox cgd patients and carriers. we now have extended these studies to patients and carriers with p phox cgd. facs analysis of p phox expression in permeabilized neutrophils demonstrated that p phox expression was absent in four patients with different mutations in ncf [two patients homozygous for c. e (+ ) g>a, one patient homozygous for c. _ del aag, p.glu del; and one patient compound heterozygous for the mutations, e (+ ) g>a and c. _ del aagaaggac]. moreover, the expression of p phox in nine p phox cgd carriers was significantly reduced > % compared to expression in neutrophils from healthy volunteers. another cytosolic phox protein, p phox, has been shown to associate with p phox in a : molar ratio [tsunawaki et al. . biochem biophys res comm. ( ): - ]. the expression of p phox was reduced in both carriers and patients with mutations in ncf . despite reduced expression of p phox and p phox, neutrophils isolated from carriers of p phox cgd exhibited normal dihydrorhodamine (dhr) oxidation after stimulation with phorbol ester and fell within the normal range for ros production (measured by luminol-enhanced chemiluminescence) after stimulation with either fmlf, opsonized zymosan, or phorbol ester with one notable exception. included in this cohort of p phox carriers was a p phox cgd patient (homozygous for a gt deletion at the start of exon in ncf ) who also carried a heterozygous damaging mutation in ncf [c. a>t; p. asn ile]. normal ros production in the presence of reduced p phox and p phox expression suggest that these proteins are not rate-limiting components for maximum nox activity in neutrophils. finally, determination of the expression of specific phox components by facs analysis of permeabilized neutrophils from whole blood provides a rapid and alternative approach to immunoblotting to determine the specific protein defect in cgd, and, importantly, one that could be easily established in most clinical labs. funded by nci contract no. n d . the original clinical observation that defined patients with hyper-ige syndrome (hies) was the presentation of cold abscesses ("job's syndrome"), which indicated a deficient inflammatory response. mutations in the stat gene have now been identified in most classic autosomal dominant hies patients, but we do not fully understand how these mutations cause the clinical presentation. since the discovery of stat mutations, research on hies focused largely on the adaptive arm of the immune system and suggested that the innate immune defects could be secondary. for example, the discovery that there is a th cell and il- cytokine deficiency in hies provided a possible explanation to the neutrophil chemotaxis defects in hies, as il- is one of the chemokines critical for neutrophil recruitment in vivo. the goal of this study was to investigate myeloid cells from hies patients. first we used c a, fmlp, il- , cxcl , and cxcl to study neutrophil chemotaxis in vitro. responses to c a, fmlp, and il- were equally robust in hies compared to healthy controls, demonstrating that neutrophils from patients are capable of efficient directed migration in vitro. neutrophils from all hies patients responded to cxcl and cxcl significantly below that of the healthy controls. cxcl and cxcl are cxcr -specific chemokines. these results indicated a neutrophil intrinsic cxcr -specific defect. we also found that patient-derived cells express comparable levels of cxcr on the cell surface, suggesting a cxcr chemokine receptor signaling defect. after identifying a neutrophil defect in hies, we wanted to get a broader view of myeloid cells in hies in addition to identifying the cxcr specific defect. stat is a transcriptional regulator, therefore we performed transcriptional profiling of hies and healthy control-derived neutrophils and monocytes. as it was shown before, the expression of stat was not different between patients and controls, since hies is usually caused by the decrease in stat activity not by decrease in expression. we found, however, an increase in stat and stat expression as well as significant changes in the expression of genes regulated by interferons. increased expressions of stat / in both neutrophils and monocytes likely provide and explanation for the increase in interferon regulated genes. multiple genes were identified as potential regulators of cxcr signaling. the balance between the stat and stat signaling has long known to be a regulator of immune cell activation, especially in t cells, but less studied in myeloid cells. stat and stat / signaling pathways crossregulate each other in healthy cells. we propose that in hies the decreased stat signaling leads to not only changes in expression of effector (e.g. inflammatory) genes, but also decreases expression of genes in the regulatory (negative) feed-back loop, which are required for decreasing stat / activity. therefore, the immune cell defects caused by decreased stat activity are compounded by the increase in stat / activity. increase in stat / signaling can cause pathologies in the absence of stat defects, as well as further decrease stat signaling, thus contributing to hies. interfering with stat / signaling in hies may represent a therapeutic opportunity. abstract/case report text rationale: t-cell receptor excision circles (trecs) testing on newborn screening (nbs) has been vital for identifying patients with severe combined immunodeficiency (scid). we aimed to determine whether one or more abnormal trecs result on a nbs might predict higher mortality rates despite the absence of an identifiable underlying etiology. methods: newborns with a positive trecs nbs result without the diagnosis of scid or q . deletion syndrome born from october to december were included (n= ). newborns were divided into three groups: group infants had a subsequent normal repeat screen (n= ); group infants did not undergo repeat screening as the majority expired before a repeat screen could be conducted (n= ); group infants had a normal initial screen but subsequent abnormal screen (n= ). cases were matched : to controls on gestational age, birth weight, nicu status, race, birth quarter, and birth year. nbs records were linked to birth and death certificate records. demographic characteristics were compared and mortality rates were calculated between the groups. results: the mortality rate of group was . %, group was . % and group was . %. when compared with matched controls, there was no difference in the mortality rate of group when compared to the control group. there was a significant difference in the mortality rate between cases and controls in both group (p < . , % ci . , . ) and group (p < . , % ci . , . ). the apgar scores in group infants were comparable to their matched controls. infants in group (p = . ) and group (p = . ) had significantly lower apgar scores than the controls. the majority of the infants in all three groups were less than weeks gestation, however, group had a higher percentage of infants born very premature (less than weeks). there was no significant difference in maternal age, maternal education, prenatal care status, cigarette use, or maternal steroid use between the cases and controls in all three groups. conclusions: infants with an initial abnormal screen who had a subsequent normal repeat screen did not have an increased rate of mortality compared to their matched controls (group ). however, group infants (with unresolved repeat screen) and group infants (with a first abnormal value on a repeat screen) did have increased mortality rates when compared to their controls. overall, an abnormal trecs level on nbs without a confirmed negative repeat screen, was associated with higher mortality in our study population. further studies will be needed to determine if the trecs assay can serve as a predictor for mortality in newborns with an abnormal screen. abstract/case report text introduction: primary immunodeficiency refers to a heterogeneous group of diseases characterized by altered function or composition of the immune system, and are grouped into adaptive or innate system defect. immunoglobulin g subclass immunodeficiencies (iggscs) are classified as a b-cell-related adaptive system disorder and are therefore associated with recurrent sinopulmonary infections with encapsulated bacteria, presenting with pneumonia, recurrent bronchitis, rhinosinusitis, and herpes zoster. its primary mechanisms are still unclear, although the cause for this deficiency might be related to gene deletions, transcription errors, or be an effect of allotype. igg immunodeficiency reaffirms its association with the patient's clinical condition and is often associated with igg deficiency. objective: to evaluate the prevalence of igg immunodeficiency in ferraroni's clinic, classify it by gender, age, igg dosage and other subclasses, correlate it with igg immunodeficiency and the clinical presentations presented by the patients under analysis. method records of patients with igg immunodeficiency whose clinical pictures were followed throughout years were evaluated, patients aged from to years. all tests were done at the same laboratory and all patients have consented to be part of this study, which has been approved by the ethics committee. results twenty-four patients with igg deficiency, , % (n= ) were women and , % (n= ) were male, with average of and years, respectively. the average of igg was . mg/dl, and that of igg was mg/dl. of the patients evaluated, . % had upper airway infections (sinusitis, rhinitis, otitis and tonsillitis), % herpes simplex, . % asthma. less prevalent cases were reported as . % of patients had bronchiectasis, . % candidiasis and . % herpes zoster. . % presented the association of igg and igg deficiency. discussion: the role of specific igg deficiency in the infectious setting is still unknown, but it usually occurs in association with other isotypic deficiencies and sinopulmonary infections. furthermore, the igg subclass is relevant on the study of environmental antigens -suggesting its involvement with allergic disordersand has been described in association with other diseases, such as chronic mucocutaneous candidiasis, ataxia-telangiectasia and allergic colitis. igg deficiency is related to increased susceptibility to bacterial infections. studies show a correlation between igg and igg immunodeficiency that generally imply clinical features characterized by recurrent infections by encapsulated bacteria. the data obtained through the analysis of patients' charts corroborated this information, since it was evident that most of the patients had really similar clinical conditions. conclusion: igg deficiency has a direct correlation with higher prevalence of upper airway infections, such as rhinitis, sinusitis and pneumonia, and with an increased incidence of allergic disorders, here presented by our cohort. additionally, research suggests that hies may cause impaired cd + t cell function. we hypothesized that a low percentage of both th and th cells would be predictive of hies and would differentiate hies from atopic disorders. to evaluate this hypothesis, we examined the percentage of th , th , and ifng+cd + t cells, laboratory parameters, and genetic diagnoses from a large cohort of patients to determine which parameters distinguish patients with stat loss-of-function variants. methods: we conducted a retrospective, multi-institutional chart review of over patients who received a th assay at the medical college of wisconsin clinical immunology research laboratory. the th assay is performed by activating pbmcs with pma/ionomycin/brefeldin a and staining for cd , cd , ifng and il- a. the following parameters were included in the chart review: the percentage of th , th , and cd +ifng+ cells, immunoglobulin levels, atopy scores, infectious history, and genetic diagnoses. results: using logistic regression, we demonstrated that the percentage of th , cd +ifng+, and th cells were positively correlated with age, and percentage of cd +ifng+ cells was higher in females than males. we found that the percentage of th and th cells were decreased in both atopic disease and hies, with hies having the lowest values. interestingly, one subject with a stat gain-of-function (gof) variant had an elevated percentage of th and th . in addition, we determined that ige levels were inversely correlated with the percentage of th , cd +ifng+, and th cells, while iga and igm were positively correlated with the percentage of th cells. several different monogenic defects characterized by increased fungal infections exhibited a low percentage of th including tatton-brown-rahman syndrome and cornelia de lang syndrome. conclusions: we confirmed that the percentage of th cells is low in both hies and atopy in a large cohort of subjects, and that the percentage of th cells may be helpful in distinguishing hies from atopic disease. however, since the percentage of th , cd +ifng+, and th cells correlate with age, caution should be used when testing young children. the inverse correlation between ige levels with th and th responses suggests that similar pathway(s) may drive both hies and atopy. additionally, the decreased th responses in stat lof and increased th responses in stat gof hies raise questions about the role of stat in regulating ifng levels. we also identified patients with different genetic disorders with fungal infections in which the th percentages were low, suggesting that the th test may be useful in evaluating individuals with unusual fungal infections. abstract/case report text background:primary and secondary autoimmuune neutropenia (pan/san) are well described entities. several autoimmune neutropenias do not fit the criteria of either pan or san showing peculiar characteristics mainly for older age at onset and/or for duration of the disease; moreover they are not associated , at least at the beginning, with autoimmune markers/diseases aim of the study: to describe a cohort of subjects affected with autoimmune neutropenia, defined as "atypical" (aan), registered in the italian neutropenia registry (inr) and to compare these data with those from subjects diagnosed with pan still in the inr. patient and methods: subjects with neutropenia and positivity of indirect antibodies against neutrophils (registered in the inr from to ) lasting for more than years, or diagnosed after years of age ( up to y), without any associated autoimmune, signs/markers were considered eligible for the present study. results: data from patients were collected: / subjects ( %) were defined as aan and / ( %) as pan. among aan affected patients %, were "long lasting" aan, while % were defined as "late onset" aan .the degree of neutropenia in aan group was mild in %, moderate % and severe in % of the subjects . leukopenia at onset was a common hall mark seen in % of aan patients (median values /mm³ ; range - /mm³ ) especially in the "late onset" aan if compared with pan and " long lasting" one ( p= . ). as for clinical features, almost half of the aan cohort suffered from recurrent or "significative infections", while severe episodes (namely sepsis, meningitis , osteomyelitis , pneumonia, deep abscess or flemmon) were shown in % being more frequent, but non significantly higher than those reported in the pan group ( % )( p=ns) interestingly, recurrent apthae were significantly more seen in the "late onset" aan group if compared with the "long lasting" aan (p= . ). during follow up, markers and/or symptoms of autoimmunity appeared in % of the aan cohort, being another element of peculiarity in respect to pan (p < . ). as for immunological pattern in aan, immunoglobulin values were lower than the references for age in %,while were above them in % of the cohort . lymphocytes subsets evaluation showed decreased value of cd +cd + cells in % of cases, followed by depletion of cd -cd +cd + subtype in %, cd +cd + in % of cases and cd + cd + in % . preliminary study on b memory and t-reg cells values, showed a quantitative deficiency respectively of in % and % of the studied subjects. mutation analysis performed by ngs in % of the subjects identified pathogenic variants of : taci ( ), tinf ( ) and lrba ( ) .comparison between pan and aan is detailed in table . conclusions atypical neutropenia in childhood is a disorder which show many difference with pan; indeed appears an epiphenomenon of a complex immunological disturbances rather than a disease itself. occasionally mutations of genes of immunodeficiency/disimmunity can be demonstrated abstract/case report text background: medications treating ra typically include systemic corticosteroids used to treat inflammation flares, and disease modifying therapies (dmards). traditional dmards include methotrexate, leflunomide, hydroxychloroquine, and sulfasalazine. recently, biologic/immuneresponse modifiers have come to the forefront for overall therapeutic benefit, however, an unfortunate side-effect may be the risk of increased immunosuppression. this study seeks to determine the occurrence rates of immune deficiencies among patients initiating ra therapies. methods: using the pharmetrics plus commercial claims database from - , ra patients (icd- and - codes: m , m ) over the age of were indexed on their first use of a new biologic therapy. all patients were required to have enrollment six-month pre and one-year post index. cohorts of patients were grouped by medication: methotrexate, adalimumab, etanercept, and rituximab. ra patients receiving adalimumab, etanercept, and rituximab were allowed concomitant use of methotrexate, but could not use any other biologic medications in the post period. a minimal adherence of % was required of all biologic treated ra patients. an additional cohort of ra patients untreated with biologic therapies was indexed on their first ra diagnosis within the time window and used as a control. ra patients with comorbid conditions who would also require biologic treatment were excluded including crohn's disease and ulcerative colitis. between group comparisons were made with the no treatment group as the referent. to account for differences in age, gender, and elixhauser comorbidity conditions patients in each cohort were matched : to the rituximab group. results: , ra patients met inclusion criteria: , in the methotrexate group, , receiving etanercept, , receiving adalimumab, and receiving rituximab. a total of , in the treated groups and , in the no biologic treatment group. demographic information including age and gender were significantly different but numerically similar between the groups, with rituximab group having the highest proportion of female patients but limited dispersion with the lowest proportion being in the etanercept group. healthcare utilization metrics highlighted a significantly higher average number of office visits ( . , sd: . vs no treatment . , sd: . , p < . ) and a higher proportion of rituximab patients being hospitalized ( . % vs no treatment . %, p < . ). the diagnosis of immune deficiency was highest among the rituximab group with . % followed by methotrexate . %, adalimumab . %, etanercept . %, and no treatment . %. after matching, similar rates were seen for healthcare utilization to the pre-match results. the post-match odds of being diagnosed with immune deficiency were significantly greater for the rituximab group (or . , ci: . - . ) than the no treatment group. conclusions: the purpose of dmards is to modulate the immune system and decrease autoimmunity in ra. however, this treatment may lead to significant immunosuppression. this study suggests that treatment with certain biologic/immune-response modifier therapies may be associated with higher rates of healthcare utilization. in particular, the increased post-treatment diagnostic coding of immune deficiency demonstrates the heightened awareness among healthcare providers of the chronic immunosuppressive potential of rituximab. evaluation of potential secondary immunodeficiency pre-and post-dmard use should be incorporated into routine practice. abstract/case report text introduction: autoimmune lymphoproliferative syndrome (alps) is a rare inherited disorder of lymphocyte homeostasis due to a fasmediated apoptosis and characterized by non-infectious and nonmalignant lymphoproliferation, autoimmunity, and secondary malignancies (national institute of health criteria). in spite of recent progress, one third of alps patients still remain gene orphan and they have been previously categorized as alps-u. in some cases, patients fitting alps diagnostic criteria have been shown to carry mutation on genes involved in other immune-dysregulation syndromes. aims: the aim of this study is to compare the clinical and immunological features, and the outcome of a cohort of alps patients with mutations on the typical causing genes (fas, fasl, fadd and casp )-here defined as alps-g -vs the ones without a molecular diagnosis or carrying mutations on other genes (both defined as alps-u). patients and methods the demographic, clinical, biochemical, genetic informations and details about treatment are derived from the alps italian network. search of mutations was performed with sanger pcr and/or next generation sequencing techniques (extended to immunodeficiency genes panel). results: alps patients were registered in our data base; the genetic analysis was performed in subjects ( %): / pts ( %) were alps-g and the remaining ( %) alps-u. six-teen out of ( %) alps-u patients resulted to carry mutations on other genes (lrba, stat +cecr, ctla , baffr, taci, nmlrc , ikbkg, gaucher), and the remaining ( %) were negative. the alps-u subjects showed a more complex phenotype compared to the alps-g group, which was characterized by multi-organ involvement (p= . ) and positivity of autoimmune markers (p= . ). (table ). cytopenia affecting one or more haematopoietic lineages was present in both groups ( % and %) with no significant difference, apart from lymphocytopenia that was more frequent in alps-u group (p= . ) ( table ) . as for lymphocyte subets and immunoglobulin dosage no differences were shown within the two groups. vitamin b and il- were more frequently raised in alps-g group (p= . , p= . ) (table ). four out of ( %) patients did not require any treatment. first-line treatment (steroid or intravenous immunoglobulins) controlled the disease only in / ( %) cases. the response rate to second line therapy -micofenolate mofetile (mmf) or rapamycin-was % and % in alps-g and alsp-u group, respectively. moreover, target therapies or drug combinations were more commonly applied in alps-u subjects (p= . ) ( table ) . conclusions: our study showed that alps-u subjects, despite the alps phenotype, represent distinct clinical entities and that genes associated with other immune-dysregulation syndromes are frequently represented in this group ( / , %). the identification of such disorders is crucial for the management of second-line treatment and/or the administration of target therapies abstract/case report text background: we have shown previously that allergic reactivity to ovalbumin (ova) could be regulated in mice following perturbation of immune networks using combinations of an immune ig along with antiidiotypic ig. we have explored features of this regulation including: its persistence after cessation of administration of combined igs; the ability of heterologous igs to produce immunoregulation; a role for treg induction in regulation; and the ability to attenuate responses in mice presensitized to an allergic stimulus. methods: balb/c mice were sensitized to ova. mice also received weekly injections of immune ig or anti-idiotype ig (at separate sites) from either homologous (mouse) or heterologous (human) sources. in the latter case pooled ivig (given im, hence hereafter imig) was used as a source of anti-idiotype ig, and human anti-tet as immune ig. injections of the ig were given from the time of ova sensitization (to attenuate development of immunity), or after pre-sensitization of mice (to attenuate existing allergic responses). all mice were assayed for development of ova-specific serum ige and igg, as well as the production of ova-induced il- , il- , il- , il- and il- in splenocytes cultured for hrs. in studies examining possible mechanism(s) responsible for inhibition of immunity mice received, in addition to the ig treatments described, infusion of depleting anti-cd , and/or anti-cd antibodies, or a mab to tnfsfr , known to expand tregs implicated in regulation of allo immunity. results: combinations of both heterologous and homologous immune igs and anti-idiotype igs attenuated ova allergic responses in both naïve and pre-sensitized mice. this attenuation persisted in mice greater than weeks after cessation of treatment with the igs used. finally, depletion of either cd or cd cells ameliorated the suppressive effect seen, while the combination of anti-cd and anti-cd essentially abolished suppression. suppression was further enhanced by anti-tnfsfr mab. conclusions: we conclude that the combine ig treatment protocols used produced a long-lasting suppression of allergic immunity, even in pre-sensitized animals. the effects seem to depend upon induction and expansion of tregs and represents a novel approach to treatment of allergic disease in humans and other animals. abstract/case report text background wiskott-aldrich syndrome protein (wasp) is found in the cytoplasm of hematopoietic cells but can transit to t lymphocyte nuclei at distinct developmental timepoints. wasp deficiency is a rare, x-linked combined immunodeficiency disease. affected patients display qualitative but not quantitative t cell defects. we report two immune deficient subjects with nearly identical exon frameshift mutations in was, the gene encoding wasp. one subject lacked circulating t cells, the other possessed several distinct cd t cell populations each expressing quantitatively different amounts of wasp. objective to determine how similar was mutations can cause scid in one person and generate b and t cells with heterogenous wasp expression in another. methods to identify somatic was mutations, we deeply sequenced was exons, introns, promoters and ' untranslated regions at , read depth in genomic dna from various b and t cell populations of each subject and their unaffected relatives. we confirmed genomic variants were transcribed and translated by sequencing was transcripts and analyzing wasp in primary cell lysates, both fractionated and not. to model our subjects' diseases we transfected primary cells and cell lines with mutant was transcripts and then measured viability and nuclear localization via confocal microscopy. results deep sequencing of genomic dna revealed all of subject one's cells carried the same germline exon frameshift was mutation. the mutation was incorporated into subject one's was transcripts and translated into a truncated form of wasp, which was relegated primarily to the cell nucleus. subject two possessed three distinct cd t cell subsets that each carried either the germline exon frameshift was mutation or a variety of somatic mutations that circumvented frameshift wasp expression. evasion strategies included exon skipping, adoption of a cryptic exon splice site and reversion to wild type amino acid sequence. subject two incorporated somatic mutations into was transcripts which encoded either stable near full-length proteins or unstable non full-length ones. subject one's sister and subject two's mother, who both carried the germline exon frameshift mutation, produced only wild type transcripts and proteins. conclusion we report two patients with was mutations encoding truncated wasp. if expressed, truncated wasp localized to the cell nucleus, and this was associated with t cell developmental arrest and severe combined immune deficiency. if, through a variety of epigenetic and somatic strategies, t cells could avoid expression of truncated wasp, they would survive but display phenotypical abnormalities and functional defects. patients with pidds show a higher susceptibility to hematopoietic malignancies, in particular to non-hodgkin lymphomas (nhl) that, generally, account for approximately - % of paediatric cancers and their incidence increases with age. recently new gene defects responsible for pidds with lymphoproliferation as a key clinical sign have been identified. our goal is to investigate possible immune-mediated mechanisms underlying malignant lymphoproliferation in children who did not show other typical symptoms of pidds. we retrospectively selected and reviewed the clinical history of nine patients with nhl ( burkitt lymphoma, large b cell lymphoma and lymphoblastic tcell lymphoma). immunophenotyping and exome analysis of known pidds genes were performed after lymphoma remission. six out of nine patients showed a mild hypogammaglobulinemia at time of presentation, not noticed before. moreover, one patient had history of recurrent respiratory infections, one of hematologic autoimmunity and two of nine were ebv-positive at diagnosis. preliminary results show an aberrant b cell phenotype in four patients; exome analysis reveals a novel heterozygous genetic variation in ikzf gene in one patient with burkitt lymphoma and autoimmune cytopenia was identified. concerning the remaining patients, further studies are ongoing. a detailed review of clinical history of paediatric patients affected from nhl as well as an impaired immunophenotyping can be important indicators of immune-mediated disorder underlying lymphoproliferation and helpful signs of possible pidds that should promptly be investigated by genetic analysis. this will allow an appropriate diagnosis and disease management. abstract/case report text introduction: caspase activation and recruitment domain (card ) encodes a scaffold protein that links antigen receptor activation to intracellular signaling. dominant heterozygous loss of function (lof) mutations in card cause a syndrome of severe atopic dermatitis, elevated ige, and allergic disease. atopic dermatitis can be difficult to control leading to substantial morbidity. dupilumab is a humanized monoclonal antibody that blocks il- and il- signaling approved for treatment of refractory atopic dermatitis. we present a case of a -year-old female with card deficiency successfully treated with dupilumab. case: a -year-old puerto rican female with history of recurrent sinopulmonary infections with episodes of pneumonia, moderate persistent asthma, food allergies, recurrent skin boils, and severe atopic dermatitis was referred for further management and evaluation for autosomal dominant hyper-ige syndrome (ad-hies). her atopic dermatitis was refractory to conventional therapy with topical corticosteroids, twicedaily emollient use, and bleach baths; it was also refractory to immunosuppression with mycophenolate mofetil and cyclosporine. on exam the patient exhibited coarse facial features and a high palate. she had eczematous lesions on the face, trunk, and extremities (scorad ). laboratory evaluation showed: eosinophilia ( cells/ul), elevated ige (> ku/l), low igm ( mg/dl), and elevated iga ( mg/dl). lymphocyte subsets and mitogen response were normal but antigeninduced proliferation was abnormal. autosomal dominant hyper ige score was indicating a high likelihood of ad-hies. no mutations in stat were identified and th cell expression was elevated. dedicator of cytokinesis (dock ) deficiency was also considered but dock protein expression was normal. further genetic testing revealed an base pair deletion in card (c. _ del) predicted to be pathogenic. the combination of the patient's phenotype and large deletion was consistent with card deficiency. despite continued immunosuppression with cyclosporine and aggressive skin care, the patient's atopic dermatitis was still severe and poorly controlled. off label (patient < years) treatment with subcutaneous dupilumab mg every weeks was initiated. at last follow-up, months after dupilumab start, the patient had substantial improvement in dermatitis with clear skin on the face, trunk, and extremities (scorad ). cyclosporine was discontinued and topical medications were applied less frequently. discussion: hypomorphic heterozygous dominant negative loss of function mutations in card have recently been associated with severe atopic dermatitis and allergic disease. treatment of atopic dermatitis in card deficiency remains challenging, but dupilumab appears to be an effective alternative to refractory disease. longer follow-up and a larger cohort of card -lof patients treated with dupilumab are necessary to understand the long-term efficacy and safety for use of dupilumab in these patients. abstract/case report text purpose: the micromilieu within premalignant respiratory papillomas supports persistent hpv / infection and disease recurrence in recurrent respiratory papillomatosis (rrp). these patients show polarized (th -/treg) adaptive immunity in papillomas and blood, enriched immature langerhans cell (ilc) numbers, and overexpressed cox /pge in the upper airway. to better understand the adaptive and innate dysregulation in rrp, we studied blood-derived monocytes, ilcs, and tissue-derived ilcs from rrp patients and controls. experimental design: monocyte subpopulations were isolated, differentiated into ilcs, activated, and then assessed by flow cytometry. monocytes were induced to differentiate into ilcs with/ without added pge , and then activated by il- γ, pge , pge +il γ, or lps. ilc cd expression was identified by flow cytometry. monocyte-derived ilcs, papilloma, foreskin, and abdomen skin ilcs, were also analyzed by qpcr for select chemokine/cytokine mrna expression after isolation, hrs later in culture, and again after poly(i:c) or tnfα stimulation. results: the three monocyte sub-populations differed between patients and controls, and patients' monocytes generated fewer ilcs. classical monocytes generated most, but not all ilcs. pge levels were higher in rrp plasma, and added pge reduced control, but not patients' monocyte-ilc differentiation. pge had no effect on ilc maturation identified by cd expression. papilloma-derived ilcs expressed low ccl- , and high ccl- mrna and were unresponsive to poly(i:c) or tnfα. tissuespecific cytokine/chemokine responses between ilcs from papillomas, foreskin and abdominal skin differed. only papilloma ilcs expressed il- γ after isolation, and they up-regulated ccl mrna hrs later without further stimulation. conclusions: monocyte/ilc innate immunity is impaired in rrp, in part due to increased pge exposure. the immunosuppressive papilloma micromilieu likely alters ilc responses that skew, hpv / -specific th /treg adaptive immunity in rrp. abstract/case report text introduction: familial mediterranean fever is a hereditary auto inflammatory disorder that typically manifests with recurrent fevers, abdominal pain and in some patients there is an associated with amyloidosis leading to eventual renal failure. while there are several common mutations in the mefv gene that when homozygous give these classic symptoms, patients with atypical mutations or heterozygous mutations often have a different clinical course. we present identical twin siblings with compound heterozygous mefv mutations but differing clinical phenotypes. case description: the index patient is a year old girl, conceived via ivf, who began having fevers at age . . her fevers occurred every weeks for months before she was referred to immunology for evaluation. her parents describe her as happy and otherwise not ill appearing during these episodes. genetic testing for familial mediterranean fever revealed compound heterozygous e q and p s mutations in the mefv genes. initiation of colchicine therapy in the affected sibling has resulted in a complete resolution of her symptoms. a trial off colchicine resulted in return of cyclic fevers. her identical twin sister was also tested, and carries the same mutation, but is still asymptomatic. this created great concern amongst their parents who had genetic testing prior to undergoing ivf that revealed no parental mutations in mefv. in consultation with genetics the mother was tested again through the same laboratory that had performed testing on the children. this revealed an identical mutation in mom who is also asymptomatic. conclusions: although classic homozygous mefv mutations have resulted in well described fever syndromes, there is considerably less data on heterozygous and compound heterozygous mefv mutations. in these two identical siblings only one patient has a classic manifestation of familial mediterranean fever. while it is possible that the other twin will develop similar symptoms later on in life, it is also possible that another factor is necessary to trigger symptoms in this unusual genetic presentation of fmf. in addition this case highlights the importance of understanding the testing method used by the laboratory performing the genetic testing. while the mother was initially reported as negative the laboratory that performed her testing only tested for the most common mefv mutations. more complete testing, that included the entire gene sequence, revealed that she did contain an mefv mutation in e q, which although more rare is thought to be pathologic when combined when combined with a second mutation. abstract/case report text there are several lines of evidence that link the pi k/akt/mtor signaling pathway to primary immunodeficiencies. hyperactivation of the pi k/akt/mtor/s k signaling pathway in immune cells can be the consequence of dominant gain-of-function mutations in the genes encoding for pi kδ that cause the activated pi kδ syndrome (apds). patients with these mutations may develop immunodeficiency and immune dysregulation as well as neurodevelopmental delay and growth retardation. in addition, mutations of genes within the pi k-akt-mtor pathway were also known to cause megalencephaly and segmental cortical dysplasia. mutations in akt , a member of the akt family of proteins and a downstream effector of pi k-mediated signaling, was shown to be associated with autosomal dominant megalencephalyassociated syndromes. here we describe a year old girl, born to consanguineous healthy parents, who presented with megalencephaly, developmental delay, hypotonia, cervical lymphadenopathy and hepatosplenomegaly. the patient had recurrent hospital and icu admissions for idiopathic thrombocytopenia (treated with ivig), recurrent laryngitis, recurrent peritonsillar abscess, preorbital cellulitis, conjunctivitis with purulent discharge, otitis media, pneumonia with pleural effusion (required drainage), metapneumovirus pneumonia with respiratory failure, recurrent skin cellulitis, and abscesses that grew mrsa (required drainage). in addition, the patient is known to have asthma and allergic rhinitis. mri of the brain showed megalencephaly, ventriculomegally, thin and dysplastic corpus callosum, a normal cerebellum, and myelination appropriate for age. immunoglobulin levels, lymphocyte subsets and the oxidative burst test were all within normal limits. cmvand ebv were not detected. bacterial cultures grew mrsa (skin), strept. pneumoniae, h. influenzae, and e. coli (urine). extensive metabolic workup was done, which was inconclusive (metabolic/mitochondrial diseases). whole exome sequencing identified an akt variant c. g>a; p.(asp asn) in exon . the variant was identified in the patient but not in the parents and it was confirmed by sanger sequencing. further molecular testing concluded that the variant is caused by a de novo mutation during early development. although pathogenic variants in akt gene were shown to be associated with megaloencephaly-associated syndromes, no associations with immune deficiency have be reported. functional studies will be pursued to confirm the link between the clinical phenotype and the identified variant in the akt gene. complex is a critical signalling adaptor that regulates lymphocyte activation, proliferation, survival, and metabolism. primary immunodeficiencies affecting each component (termed 'cbm-opathies') result in broad clinical manifestations ranging from combined immunodeficiency (cid) to atopic disease or lymphoproliferation. we present the laboratory and clinical findings of two canadian first nations patients found to be homozygous for the same novel card mutation (c. c>t; p.r *) causing complete card deficiency. results: we recently identified an -month-old boy who presented with a severe case of entero/rhinovirus bronchiolitis with interstitial lung disease and a -year-old boy with a history of severe pulmonary infections with bronchiectasis (including pjp), chronic sinusitis, candidiasis, invasive bacteremia, and severe ileo-colitis and oral ulceration requiring total colectomy. testing of both patients demonstrated absent tregs, elevated naïve b cells with absent memory b cells, and panhypogammaglobulinemia. next generation sequencing revealed that both patients were homozygous for the same novel variant of card (c. c>t; p.r *), which rendered card protein undetectable by immunoblot. card deficiency was confirmed by stimulating patient b cells with phorbol -myristate acetate (pma) and ionomycin and immunoblotting for signalling proteins in both the nf-κb (ikkα/β, iκbα, p ) and mapk (mek / , mkk , jnk / , erk / ) pathways as well as cleavage substrates of the malt paracaspase (relb, cyld, bcl , hoil ). nf-κb and jnk activation were completely absent and m a lt p a r a c a p a s e a c t i v i t y w a s l o s t . f u r t h e r m o r e , c oimmunoprecipitation experiments revealed that card was required for optimal malt association with bcl in response to stimulation. to define the impact of card deficiency on the b cell transcriptome, rna-seq experiments were performed. this revealed an inability to upregulate critical genes involved in immunity and tolerance (e.g. cd lg, ctla , il , il ), decreased enrichment in cytokine pathways (e.g. ifn-α, il- , tgf-β), and decreased enrichment in malt -dependent genes. furthermore, rna-seq confirmed the developmental block observed in patient b cells and suggested that b cells were halted at the centroblast to centrocyte transition. both patients ultimately underwent hematopoietic stem cell transplantation (hsct), which restored lymphocyte signalling and activation as measured by nf-κb, jnk, and malt paracaspase substrate cleavage. conclusions: we have presented the most comprehensive clinical and molecular characterization of human card deficiency to date. these two cases highlight the crucial role of card in regulating b cell development, function, and humoral responses, as confirmed by signalling and transcriptomic analyses. furthermore, hsct is potentially helpful for these patients as assays performed on post-transplant cells demonstrated restored signalling and activation. abstract/case report text introduction: primary immune deficiencies (pid) can have a significant impact on the quality of life of patients and their families. as more patients with pid are surviving to adulthood, the need to monitor them closely and ensure they are transitioned appropriately is even more crucial. we compared the perspectives of pediatric and adult immunologists toward the transition of patients with pid at our institution. methods: pediatric allergy/immunology providers at lurie children's hospital and adult allergy/immunology physicians at northwestern university both in chicago, il completed respective surveys anonymously (www.surveymonkey.com). questions were derived from the validated 'attitude' and 'quartt' instruments for transition. respondents were asked to rate their level of agreement on a -point likert scale, ranging from strongly disagree to strongly agree. results: overall, pediatric and adult providers participated (response rate . %). of total respondents, % thought the transition process should be initiated at age - . about % of the adult immunologists selected and older, whereas pediatric providers would begin earlier; . % of pediatric providers note they would initiate transition at age - . both pediatric and adult immunologists agreed that patients should be transferred when the provider felt they were ready ( %) and when they were in stable condition ( %). both adult and pediatric immunologists selected transfer of complete medical file as a preferred communication method for transition. other strategies preferred by adult providers were a referral letter with brief summary of medical history ( . %) and staff meeting with pediatric and adult immunologists ( . %), whereas pediatric providers would prefer a joint outpatient dedicated transition clinic ( . %). pediatric and adult immunologists, patient, parent, and transition liaison were considered the most important active participants in the transition process. the most prominent barriers to a formal transition were unavailability of a transition coordinator or nurse specialists ( %) or of all disciplines of the interdisciplinary team ( %), and limited time ( %). on the other hand, limited demand (too few patients) was strongly rejected as a barrier. all participants agreed during transition patients should be educated about medications and their side effects, their condition and related potential future complications, and symptoms that require seeking health care. over % of participants also agreed that education about how to set up ivig and further insurance needs were important. all participants agreed that the transition process should include assistance on how to promote the patients' independence and self-management skills, medication management/adherence, and understanding of immunoglobulin replacement and side effects. other important transition components included knowing how frequently lab draws are required for monitoring ( %) and having a written individualized transition plan ( %). pediatric providers also thought having an email or telephone help line would be beneficial. conclusion: this study adds to the growing body of literature examining attitudes of immunologists toward transition, and it highlights important transition components and barriers. further work is ongoing to determine the transition needs identified by patients and parents and define markers for successful transfer in order to build a transition policy at our institution specific to immunodeficiency patients. abstract/case report text introduction: bronchiectasis (bq) is an abnormal and irreversible dilatation of bronchi secondary to repeated cycles of airway infection and inflammation. predominantly antibody deficiency is the main group of primary immunodeficiencies (pid) in adults and had been reported up % of subjects with non-cystic fibrosis bronchiectasis (ncfb). hypergammaglobulinemia (igg level higher than , mg/dl) had been observed in . % of ncfb cases (retrospective data). diagnostic delay and inappropriate management of patients with predominantly antibody deficiency can lead to irreversible lung damage or even death from serious infections. the effect of hypergammaglobulinemia on ncfb is unknown. here we present the frequency of immunoglobulin abnormalities (pad and hyperigg) in adults with ncfb in cali, colombia. methods: we present preliminary data of a descriptive prospective study that will include patients with ncfb. women and men > and < years old will be included. all volunteers will be evaluated by a clinical immunologist, complete blood count and serum igg, iga, igm and ige levels will be determined. according with clinical suspicious, igg subclasses, anti-pneumococcal igg response and b cell subpopulations will be performed. the project will be executed in months. written informed consent has been obtained for all subjects included. this project count with irb approvals at universidad del valle and hospital universitario del valle. results: a total of ncfb cases have been included in the study. the mean age was . years ( - years) with a female:male ratio : . moderate-severe dyspnea was observed in / cases (medical research council -mrcdyspnea scale to ). recurrent pneumonia was found in / cases ( %). the main etiologies of bronchiectasis were: post-infection / ( %); idiopathic / ( %); autoimmunity / ( %); primary immunodeficiency / ( %) asthma / ( %); copd / ( . %); primary ciliary diskinesia / ( %); reflux / ( %) and others. primary immunodeficiencies, . % of ncfb cases, were classified as: predominantly antibody deficiency ( cases) including cvid cases, igm deficiency cases, igg subclasses deficiency case, selective iga deficiency case and hypogamaglobulinemia case. combined immunodeficiency (dock deficiency) case. interestingly igg hypergammaglobulinemia was observed in / cases ( . %) suggesting humoral immune response deregulation. conclusion to the best of our knowledge this is the first prospective study evaluating the etiology of non-cystic fibrosis bronchiectasis (ncfb) in colombia. hypergammaglobulinemia and predominantly antibody deficiencies affect % of adults with ncfb in colombia. our study reinforced the necessity to evaluate humoral immune response in patients with bronchiectasis. conflict of interest: authors disclosure any potential financial conflict of interest related to this abstract. acknowledgements: this investigator-initiated research was supported with a grant from baxalta us inc, a member of the takeda group of companies bt - /iir-col-bxlt- . abstract/case report text background: early-onset inflammatory bowel disease (eoibd) is defined as ibd diagnosis in children less than years of age. the occurrence of autoimmune disease in children (where it is relatively rare, compared to adults) may be caused by a highrisk predisposition gene (monogenic disorders). mayo clinic children's center has a unique care model, where a patient who is referred for eoibd meets with a team of physicians, including gastroenterology, immunology, genetics, and nutrition. we describe our experience of our eoibd clinic from an immunologic perspective. methods: we conducted a retrospective cohort study through emr chart review of pediatric patients who were referred to our eoibd program ( - ). first diagnosis of ibd under the age of was the inclusion criteria. we assessed the presentation, clinical correlates, and immunologic evaluation. approval was obtained from mayo's institutional review board. data abstraction and analysis was done using the software jmp. results: pediatric patients met the inclusion criteria, with ( %) males and ( %) females. the median age of ibd diagnosis was years ( - years range). median values and the distribution of variables used in the nutritional and immune evaluation were assessed (fig ) . nutritional assessment was remarkable for low to low normal hemoglobin and ferritin levels. vitamin d and albumin levels were overall within the normal range. growth parameters indicated that the median bmi percentile was ( - ). with immune and genetic screening, one patient was found to have x linked chronic granulomatous disease (cgd). immune evaluation of other patients was overall within normal limits. fecal calprotectin served a reliable non-invasive biomarker for inflammation with the median being . ( . - . ). of these patients underwent gi pathogen panel testing of which ( %) tested negative, four ( %) tested positive for c. diff, and two ( %) others to shiga toxin-producing e. coli. it was also noted during the chart review that most patients had poor disease control despite undergoing treatment with various anti-inflammatory and immunosuppressive drugs. the patient diagnosed with cgd underwent bone marrow transplantation. a higher proportion of patients referred to our program in recent years underwent a more comprehensive multispecialty evaluation. conclusion: awareness of monogenic causes of inflammatory disorders in children has increased in recent years. it is also important to rule out intestinal infections that can act as ibd mimic. identifying monogenic disorders and other ibd mimics helps with targeted therapy and symptom improvement in these patients who have a difficult-to-treat disease. the group of children with eoibd, regardless of whether there is an inborn error of immunity, suffers from very high morbidity and a high burden of disease. comprehensive immune-nutrition assessment of eoibd patients paves way for further in-depth immunogenic assessments and allows for global management. abstract/case report text background: chronic granulomatous disease (cgd) is a primary immunodeficiency (pid) affecting the nadph oxidase system in phagocytes resulting in increased susceptibility to catalase-positive organisms. holland presented the first report of dual impact of cgd and hiv in a patient with disseminated nocardiosis. because their cgd patient admitted to history of iv drug use, he was frequently screened for hiv. case: a -year-old african american male with known cgd tested positive for hiv by western blot in the ed in when he presented with complaints of intermittent fever and cervical lymphadenopathy. his cgd was diagnosed by nbt blood testing at years of age. he had frequent skin infections and fever prior to diagnosis. clinically, he did so well that his cgd diagnosis was questioned by his immunologists. however, cgd was confirmed by additional abnormal nbt tests and, ultimately, dhr flow cytometry testing. during his second infectious disease consultation for hiv, at age , he disclosed that he was bisexual. previously, the patient was screened for hiv and hiv antibodies in due to anal fissure. he was screened again in for marked cervical and supraclavicular lymphadenopathy. his cd + t cell absolute count was noted to be low ( /mm ) in at age . until , his prior hiv screenings were negative. during his cgd treatment course as an adult, he was known to be variably adherent with administration of interferon gamma due to adverse effects, particularly pain at the site of injection and malaise. at the time of his positive hiv western blot in , his cd + t cell count was / mm . after starting hiv antiretroviral treatment, his viral load became undetectable. at age , he had burkholderia cepacia pyelonephritis resulting in left nephrectomy. sepsis from b. cepacia was fatal (positive blood cultures without known primary source) in at age . his recent viral load was still undetectable and cd + count was /mm . summary: our case reveals the complexities of treating a patient with both primary and acquired immune deficiencies. it illustrates the importance of taking a thorough social and sexual history starting in adolescence, including those patients with pid. patients with pid should be followed closely by a primary care physician, in addition to an allergist-immunologist and infectious diseases specialist, to ensure age appropriate medical and developmental screening. the recognition of pids is improving due to better screening, awareness, and treatment. currently, the rate of hiv infection is highest among young homosexual african american males. it remains important to understand the epidemiology of primary and acquired immunodeficiencies to best identify those at highest risk. ( ) submission id# phosphatase and tensin homolog (pten) hamartoma tumor syndrome identified by newborn tcell receptor excision circle screening for severe combined immunodeficiency δ) subunits that are critical for cellular signaling. heterozygous gain-offunction (gof) mutations in pik cd (encoding p δ) result in activated pi k δ syndrome (apds ), while heterozygous loss-of-function (lof) mutations in pik r (encoding p α) result in activated pi k δ syndrome (apds ). given its role as a negative regulator of the pi k signaling pathway, heterozygous lof mutations in pten (encoding phosphatase and tensin homolog, pten) result in a clinical phenotype that approximates that of apds /apds and is therefore referred to as activated pi k δ syndrome-like (apds-l). however, sequelae of heterozygous pten lof mutations extend beyond the immune system and include a group of disorders collectively known as pten hamartoma tumor syndrome (phts). although severe t cell lymphopenia at birth would be unexpected in apds , apds , or apds-l, below normal t cell receptor excision circle (trec) counts have been reported in apds , but only in individuals outside of the neonatal period. herein, we describe an infant girl with a low trec count at birth who was found to have phts. case description: a -day-old girl, born at a gestational age of weeks, was found to have a low trec count of /microliter (normal => ). a second trec count obtained at weeks of age resulted as /microliter. arguing against a diagnosis of severe combined immunodeficiency (scid), flow cytometric analyses performed at weeks of age revealed only a modestly diminished cd + t cell count ( /microliter; cd + and cd +) with a normal percentage of naïve and memory cd + t cells ( % and %, respectively). by months of age, her cd + t cell count dropped to /microliter, which was accompanied by a significantly decreased percentage of naïve cd + t cells ( %). sequencing and deletion/duplication analysis was pursued via a commercially available -gene panel aimed at genetically defined primary immunodeficiency (pid), in which no clearly pathogenic mutations were identified. over the following months, the patient was noted to have macrocephaly, tall stature ( th percentile), axial hypotonia, and gross motor delays. sequencing and deletion/duplication analysis was then pursued via a commercially available -gene panel aimed at genetically defined macrocephaly and overgrowth syndromes, in which a hemizygous pathogenic mutation in pten (c. a>g, p.gln arg) was identified. subsequent flow cytometric analyses demonstrated findings characteristic of apds-l, including expanded transitional and cd lo b cells, decreased isotype switched memory b cells, increased effector memory t cells, a lowered threshold for intracellular calcium mobilization upon b cell receptor engagement, and increased basal akt (protein kinase b) and s (ribosomal protein s ) signaling. discussion: we report the first case of phts identified by newborn trec screening for scid. as pten is not included in most commercially available, scid-or pid-tailored gene panels, phts would be missed by conventional genetic testing. therefore, analysis for variants in pten should be considered in neonates with low trec counts, macrocephaly, developmental delay, and other suggestive sequelae. abstract/case report text primary (or familial) hemophagocytic lymphohistiocytosis (hlh) is a rare, life-threatening hyper-inflammatory syndrome affecting mainly young children. it is caused by mutations in genes involved in the granule-dependent cytotoxic pathway, inducing extreme inflammation and massive tissue infiltration by activated t cells and macrophages. standard chemotherapy-based treatment regimens are toxic and induce remission in only % of patients. to this day, hsct is the only available curative treatment, but the inability to efficiently control the inflammation in many patients prior to transplantation often leads to graft failure, with transplant-related mortality around %. thus, the development of new, more potent and less toxic anti-inflammatory regimens would be a major advancement in the treatment of hlh. here, we hypothesize that combination therapies targeting several jak-dependent cytokines will be more effective than monotherapy to reduce the life-threatening symptoms induced by this pathology. using a perforin-deficient (pko) mouse model, we first tested the effects of blocking antibodies against ifnγ, the dominant cytokine secreted during hlh, in combination with antibodies targeting other highly elevated cytokines, such as il- and il- , on the manifestations of the disease. we found that anti-il- r and anti-il- antibodies, when used in combination with anti-ifnγ antibodies, did not significantly improve the symptoms of hlh compare to anti-ifnγ antibodies alone. further, we found that targeting the jak-stat signaling pathway with ruxolitinib, a specific inhibitor of jak and jak , molecules downstream of ifnγ and il- , but not il- signaling, was as beneficial as anti-ifnγ monotherapy. next, we tested the efficacy of ruxolitinib in combination with anti-il- antibody, as this later cytokine is not jak-dependent and was shown to drive macrophage activation syndrome in other contexts. unfortunately, this combination did not result in better symptom resolution than the use of ruxolitinib only. in contrast, combination therapy using ruxolitinib and anti-ifnγ antibodies showed a striking synergistic effect on the resolution of most disease manifestations, to such an extent that our pko mice presented a clinical phenotype indistinguishable than that of a c bl control mice. our findings demonstrate that jak-dependent cytokines are the main cytokines driving the progression of hlh in pko mice. collectively, our results suggest that anti-ifnγ antibodies and ruxolitinib, although effective independently, should be used in combination to more efficiently suppress hlh progression. these results are particularly relevant since the emapalumab, an anti-ifnγ monoclonal antibody was recently approved by the fda for the treatment of hlh while ruxolitinib will soon be in clinical trials for this indication. this project was supported by funds from the fondation de cancérologie charles bruneau and the canadian institutes of health research (mop- ). abstract/case report text introduction: transcription factor (tcf ), also known as transcription factor e -alpha (e a), is a helix-loop-helix transcription factor which plays a critical role in lymphopoiesis. tcf is required for b and t lymphocyte development. defects in tcf have been associated with agammaglobulinemia , autosomal dominant, characterized by low levels of immunoglobulin and early onset recurrent bacterial infections. deletion or diminished activity of tcf may also play a role in lymphoid malignancies. runs of homozygosity (roh) are contiguous stretches of homozygous genotypes at consecutive polymorphic dna marker positions. roh are important reservoirs of homozygous deleterious variation. the homozygosity heterogeneous hmm (h m ) algorithm was specifically developed for analyzing whole exome sequencing (wes) data. the branch point sequence (bps) is an essential splicing signal located - bases upstream of splice acceptor sites. while bps variants are rare, they may result in aberrant pre-mrna splicing and genetic disorders. these variants may be overlooked by standard wes analysis methods because they are intronic and the mammalian bps is a degenerate motif. objective: describe the method used to identify a bps variant in consanguineous brothers with immunodeficiency, including early onset recurrent infections and b-all, hypogammaglobulinemia, t and nk lymphocytosis, low b cells, and low naïve t cells. methods: wes was performed for all family members. data were analyzed using standard read mapping, variant calling and annotation methods. roh were analyzed using the h m algorithm (magi et al. ). roh from the siblings was intersected (bedtools) and the output was submitted to the genomic oligoarray and snp array evaluation tool (v . ). variants were confirmed by sanger sequencing. results: no candidate disease variants were detected in the coding regions, ' or ' splice sites, or utrs for both brothers; however, analysis of intersected roh revealed a homozygous tcf intronic variant within a putative bps (tcf c. - a>t). sanger sequencing of the mutant cdna revealed activation of a cryptic splice site. heritability for routine childhood vaccines has been shown to range from - %. the genetic component of vaccine response suggests we should be able to predict vaccine response in infants with biomarkers. methods: multi-center study of infants born vaginally at full term and followed through months of life. cord blood was collected at birth & peripheral blood was collected at and months of life. six-month collection was weeks post administration of routine vaccinations, while -month collection was immediately prior to receiving the -month booster vaccines. b cell subsets were analyzed with flow cytometry. vaccine titers and cytokines were measured via multi-plex elisa. study was irb approved. results: our data confirmed the immaturity of the newborn humoral immune system with a lower overall b-cell abundance, a predominance of naïve b cells, an inability to class-switch and produce igg or iga, and a th bias. maturation was observed over the first year with increasing overall b-cell abundance, frequency of memory b-cells producing iga, igg, and igm, and frequency of plasmablasts. scd levels also increased throughout the first year due to microbial translocation reflecting establishment of the microbiome. conversely, cord blood contained high levels of baff, april, scd l, il- , and il- , with levels decreasing thereafter. all infants displayed evidence of humoral immune system activation after getting -month vaccines. total plasmablast levels peaked weeks after receipt of month immunizations. a decrease in total plasmablasts was evident between and months, although levels remained above those at birth, corresponding with the need for -month booster vaccinations to maintain long-lasting immunity. il- and ifn-gamma had a significant positive correlation with memory b cells and plasmablasts at subsequent time points suggesting that these cytokines play a role in b cell differentiation and vaccine response. baff and april cytokines were elevated at birth, consistent with germinal center formation and underwent a compensatory decrease thereafter. april & scd levels in cord blood significantly correlated with higher tetanus titers at months suggesting that vaccine response may be predicted by cytokine biomarkers at birth. response to vaccines was also dynamic with il- levels being significantly correlated with tetanus titers at weeks after receipt of month immunizations. conversely, scd l levels did not correspond to b cell development consistent with a known b cell hyporesponsiveness to cd l in infants. conclusion: humoral immune development is both predictable and dynamic. biomarkers in the cord blood, produced by the infant, are predictors of b cell development and vaccine response in infancy. background: adult-onset immunodeficiency with anti-ifnɣ autoantibodies is a newly described immunodeficiency syndrome characterized by disseminated nontuberculous mycobacterial and other opportunistic infections in previously healthy middle-aged individuals typically from southeast asia. it is caused by the presence of autoantibodies directed against the cytokine ifnɣ, which is required for intracellular pathogen killing by macrophages as well as phosphorylation of the transcription factor stat , which is involved in cell survival gene expression. successful treatment of the immunodeficiency has been described in prior case reports with immunomodulatory therapies, including rituximab. however, there are no standard recommendations for dosing or timing of these agents, or recommendations for longterm monitoring of disease activity. case presentation: a -year-old laotian woman with a history of type diabetes and possible prior hepatitis c infection presented to the immunology clinic for evaluation of immunodeficiency. in the two years prior to presentation, the patient was diagnosed with mycobacterium avium infection involving the parotid gland and lymph nodes of the neck, mycobacterium avium complex bacteremia, histoplasma capsulatum involving the lymph nodes of the neck, and leukocytoclastic vasculitis of the lower extremities. as a child and young adult, she had no severe or recurrent illnesses and did not suffer from any chronic disease. preliminary immunologic testing demonstrated normal t, b, and nk cell subsets, elevated immunoglobulin g, a, and m levels, and protective titers to tetanus, diphtheria, and / pneumococcal serotypes. measurement of anti-ifnɣ autoantibodies was positive, which led to the diagnosis of adult-onset immunodeficiency with anti-ifnɣ autoantibodies. the patient was treated with four doses of monthly rituximab with resolution of the anti-ifnɣ autoantibodies, restoration of normal stat phosphorylation, and depletion of cd -positive b cells. after a -year period of being lost to follow-up, during which she continued to receive rituximab every months at the direction of a local provider, the patient re-presented to the immunology clinic to re-establish care. at that time, the patient had no evidence of anti-ifnɣ autoantibodies based on titers and normal stat phosphorylation. cd -positive b cells remained depleted. the patient also confirmed subjective clinical improvement and denied any interim infectious complications. conclusion: this case provides an example of successful treatment of a patient with adult-onset immunodeficiency with anti-ifnɣ autoantibodies with rituximab. it also highlights the utility of ifnɣ functional testing with stat phosphorylation, which may be used to monitor disease activity and to make decisions about ongoing immunomodulatory treatment. it is necessary to monitor additional immunological markers to refine the diagnosis of a secondary post-lt immunodeficiency to take preventive measures before infections occur. we sequentially measured t lymphocytes and antibodymediated immunity in a -year-old male receiving a lung transplant for idiopathic pulmonary fibrosis to determine which immune indicators could improve the identification of a secondary immunodeficiency. methods: t and b cell numbers, igm, igg, iga, ige and igg subclasses and specific antibodies to s. pneumonia capsular polysaccharides were assessed over a months-long post lt period. the clinical progress, infections and pulmonary function were monitored prior to transplantation and at regular intervals thereafter. results: the patient had progressive idiopathic pulmonary fibrosis starting with an episode of pulmonary hypersensitivity years earlier. he developed increasing respiratory failure progressing to complete dependency requiring a lt in june . he was treated with prednisone, mg/ day continuously for months, then decreased to mg/day. other immunosupressants included mycophenolic acid and tacrolimus and on/ off antibiotics that eventually led to severe tendinitis at - months post lt. at ½-month post lt he developed an early onset bronchiolitis obliterans syndrome (bos) that was controlled by increasing the prednisone dose. sequential immunologic evaluation showed his igg dropping from , mg/dl to after two weeks and then remaining stable at that level for the rest of the observation period. igm and iga had minor variations and ige remained very low. igg fell from - mg/ml pre-lt to - in weeks and remained stable at that level thereafter. antibodies against s. pneumoniae polysaccharides started high, between - ·g/ml for all serotypes and fell rapidly in the first weeks post lt, then continued a steady decline with > % serotypes falling < . ·g/ml at months. twelve of pneumococcal serotype antibodies increased above . ·g/ ml after igg replacement at months. cd t lymphocytes decreased from ± , cells/ul to - at month, remaining at that number after that. cd /th cells increased from - cells/ul to at months when prednisone was tapered down and then decreased to after increasing the prednisone dose again. this decrease coincided with a reduction in bos manifestations. conclusions: immune monitoring revealed an independent decrease of immunoglobulins with a stronger decrease in igg and specific pneumococcal antibodies. the role of th cell increase in developing bos needs further investigation. stat is a frequent target of cancer therapies due to its role in certain malignancies for cell proliferation and metastasis. with lof stat , decreased incidence of some cancers may be expected, however increased rates of lymphoma are described. we sought to describe the incidence and spectrum of malignancy in our relatively large lof stat cohort. methods: we performed a retrospective analysis of lof stat patients evaluated at the nih clinical center to determine the type of malignancies diagnosed, treatments received, and outcomes following therapy. results: a total of patients with malignancies were identified (cancer incidence %). six patients ( %) were diagnosed with non-hodgkin lymphoma (nhl); with diffuse large b-cell lymphoma (dlbcl) and with burkitt lymphoma (bl) with age at diagnosis ranging from years to years with median age of years. pathology staining for ebv was available in four patients; all of whom were negative by eber. all dlbcl patients received da-epoch-r for - cycles, and all achieved complete remission. five of patients with lymphoma are alive and disease-free. one patient died of heart failure years post chemotherapy without disease relapse. two patients were diagnosed with papillary thyroid carcinoma at ages and , one of whom was subsequently diagnosed with nhl. two other patients were diagnosed with basal cell carcinoma of the skin at ages and ; both of whom had prior voriconazole exposure. conclusion: malignancy, most commonly nhl, occurs in patients with lof stat mutations. nhl should be considered in patients with progressive lymphadenopathy, and thyroid carcinoma should be considered in patients with thyroid nodules. patients treated with voriconazole are at an increased risk of skin cancer and require careful skin monitoring. as survival increases, it will be important to monitor the incidence of malignancies diagnosed, as it is possible that decreased stat signaling may prove to be protective of some cancers, such as colon and breast carcinoma, in which increased stat signaling is implicated in pathogenesis. abstract/case report text introduction: recurrent pneumonia is defined as or more episodes of pneumonia in one year or more than pneumonias throughout life (with radiological resolution between episodes). in retrospective studies, up to % of adult subjects with recurrent pneumonia coursed with primary immunodeficiencies (pid). prospective studies evaluating the etiology of recurrent pneumonia are scarce. diagnostic delay and inappropriate management of patients with pid (predominantly antibody deficiency for example) could lead to irreversible lung damage or even death from serious infections. here we present the frequency of primary immunodeficiencies in adults with recurrent pneumonia in cali, colombia. methods: we present preliminary data of a descriptive prospective study that will include patients with recurrent pneumonia. women and men > and < years old will be included. all volunteers will be evaluated by a clinical immunologist, complete blood count and serum igg, iga, igm and ige levels will be determined. according with clinical suspicious, igg subclasses, anti-pneumococcal igg response and b cell subpopulations will be performed. the project will be executed in months. written informed consent has been obtained for all subjects included. this project count with irb approvals at universidad del valle and hospital universitario del valle. results: a total of recurrent pneumonia cases have been included in the study. the mean age was . years ( - years) with a female:male ratio : . moderate-severe dyspnea was observed in / cases (medical research council -mrcdyspnea scale to ). non cystic fibrosis bronchiectasis was found in / cases ( %). the main etiologies of recurrent pneumonia were: primary immunodeficiency / ( %); asthma / ( . %); autoimmunity / ( . %); primary ciliary diskinesia / ( . %) and others. hypergammaglobulinemia represented / ( %) of cases. primary immunodeficiencies, % of recurrent pneumonia cases, were classified as: predominantly antibody deficiency ( cases) including cvid cases, igm deficiency cases, selective iga deficiency cases, igg subclasses deficiency case, hypogamaglobulinemia case and agammaglobulinemia case. combined immunodeficiency: dock deficiency case and ataxia telangiectasia case. conclusion: to the best of our knowledge this is the first prospective study evaluating the etiology of recurrent pneumonia in colombia. predominantly antibody deficiencies and igg hypergammaglobulinemia affect % of adults with recurrent pneumonia in colombia. this study allows us diagnosed more than new cases of adult onset pid. immunological evaluation is critical in the assessment of patients with recurrent pneumonia. conflict of interest: authors disclosure any potential financial conflict of interest related to this abstract. acknowledgements: this investigator-initiated research was supported with a grant from baxalta us inc, a member of the takeda group of companies bt - /iir-col-bxlt- . abstract/case report text introduction: lysinuric protein intolerance (lpi) is an autosomal recessive metabolic disorder due to pathogenic mutations in slc a . it is distinguished by decreased plasma concentrations and increased urinary excretion of lysine, arginine and ornithine and can present with multiorgan involvement and a spectrum of immune deficiency. we present a five-year-old female with lpi, early-onset juvenile systemic lupus erythematous (sle), hemophagocytic lymphohistiocytosis (hlh), and granulomatous skin lesions that were positive for vaccine-strain rubella. methods: retrospective chart review was conducted. laboratory investigations included lymphocyte immunophenotyping by flow cytometry, lymphocyte proliferation to mitogen, quantitative serum immunoglobulins, vaccine titers, autoantibodies, metabolic studies, and genetic evaluation by next generation and whole exome sequencing. results: a five-years-old female of mixed native american and african american race presented at years of age with severe failure to thrive, history of recurrent fevers, joint swelling, recurrent skin lesions, severe anemia and neutropenia, and hypergammaglobulinemia. upon further evaluation, she demonstrated hyperferritinemia and ana, rnp, smith, and ss-a autoantibodies and was diagnosed with early-onset juvenile sle. laboratory immune evaluation revealed age-appropriate lymphocyte subpopulations and lymphocyte proliferative responses to mitogens and antigens, markedly elevated igg, iga, igm with no associated monoclonality, and protective tetanus and pneumococcal titers. given her severe clinical manifestations at an early age, concern for immunodeficiency prompted further genetic evaluation with next generation dclre c sequencing, which was negative, and whole exome sequencing, which revealed two heterozygous mutations in slc a , consistent with lpi. laboratory metabolic evaluation was also consistent with a diagnosis of lpi. she continued to experience recurrent cutaneous lesions on her upper and lower extremities. biopsy findings were consistent with a granulomatous lesion and subsequently identified by the cdc to have vaccine-strain rubella infection. due to recurrent pneumonia and concern for pulmonary alveolar proteinosis (pap), pulmonology was consulted and eventually confirmed pap, and she has required home oxygen supplementation. given her history of recurrent infections and vaccine-strain rubella infection, supplemental ivig was initiated. her sle has been fairly refractory to medical management, including systemic corticosteroids, mycophenolate, rituximab, and cyclosporine. she recently developed hlh at years of age and is currently maintained on canakinumab, mycophenolate, and systemic corticosteroids, yet continues to have sle and pap that has been difficult to control. conclusion: the range of clinical and immunologic findings in lysinuric protein intolerance has varied widely in the literature. our patient presented with early-onset juvenile sle and did not develop hlh and pap until years after initial presentation. despite relatively normal cellular immunity by laboratory evaluation, our patient was identified to have chronic infection with vaccine-strain rubella virus, indicating severe t cell dysfunction, and poses challenges for future immunomodulatory treatment. introduction: x-linked immunodeficiency with magnesium defect, ebv infection, and neoplasia (xmen) disease is caused by lossof-function (lof) mutations in the magnesium transporter (magt ) gene. it is a rare x-linked combined immunodeficiency and selective congenital disorder of glycosylation. clinical manifestations include chronic ebv viremia, recurrent bacterial and viral infections, lymphadenopathy, splenomegaly, autoimmunity, liver and central nervous system (cns) abnormalities. magt deficiency was first noted to result in chronic ebv infection and an increased susceptibility to ebv+ lymphomas. we recently recognized merkel cell carcinoma at a very young age in two xmen patients, leading to our review of the malignancies in this cohort. methods: we reviewed the records of male patients ( seen at the nih) with confirmed hemizygous lof mutations in magt for diagnosis of malignancy, therapy, and outcome. results: we identified malignancy in patients of with magt deficiency ( %). four patients had hodgkin's lymphoma (hl) (ages - years), three had non-hodgkins lymphoma (nhl) (ages - years), one had kaposi sarcoma ( years) , one patient developed eber-negative liposarcoma (age years) after receiving chemo and radiotherapy for severe lymphoproliferative disease (lpd) at age years and two had merkel cell carcinoma at exceedingly young ages ( and years). all patients had chronic ebv viremia. they all received treatment according to established protocols. currently, all except for three patients are alive and in remission, including one post-hsct. overall malignancy survival of %. it is important to note that three patients who did not have malignancy had ebv lpd so severe that it warranted treatment with a malignancy protocol, with one mistaken as having lymphoma. conclusion: xmen immune deficiency, an x-linked glycosylation disorder, is a multisystem disease associated with increased susceptibility to malignancies. initially, ebv driven lymphoproliferation and lymphoma was described with xmen; however, with increasing diagnoses, more malignancies are being recognized. all the recognized malignancies are associated, at least in part, with dnaviruses, including ebv, hhv- , and merkel cell virus. understanding the clinical phenotype and pathogenesis of this disease will improve monitoring and early diagnosis of malignancies for patients with magt deficiency. abstract/case report text background: primary immune deficiencies (pid) constitute a heterogeneous group of over individually rare congenital diseases that involve genes coding for proteins of the immune system, and which result in increased susceptibility to infection, inflammation, autoimmunity, allergy and cancer. the complexity of the diagnostic task, and the intrinsic biases and limitations of the human mind, can be aided by computational tools. among the available machine learning approaches, decision tree algorithms select the best node to split based on entropy and information gain; random forests build dozens or thousands of decision trees randomly to improve accuracy and reduce overfitting. aim: to implement a machine learning-assisted clinical decision support system for the diagnosis of pid. methods: with a local database of patients with suspected iei, we built a decision tree using c . dtc, and a random forest on python . (jupyter notebook, scikit, mathplotlib, pandas, numpy). the database was obtained by conducting an electronic search on medsys of patients with the term "immunodeficiency" in their electronic medical records, and then hand-picking cases in which a pid had been confirmed or ruled out. it consisted of patients, of which had been diagnosed with iei. we first split the dataset randomly into training ( %) and testing ( %) sets. the decision tree was tasked with classifying correctly pid or not. after running the algorithm in the training set, we evaluated in the testing set through cross-validation. results: accuracy was greater than % for the dataset (pid/not). . for the dtc with levels. the attribute with the lowest gini coefficient was low iga ( . ). accuracy for the random forest classifier was . with trees. feature importance was highest for lung infection ( . ), high igg ( . ), low iga ( . ), skin infection ( . ), no isolate ( . ), and allergy ( . ); it was lowest for consanguinity, high igm, central nervous system infection, parasites and no infections. during the random generation of trees, accuracy reached up to %. discussion: we built two classification models. decision trees lend themselves more easily to learning and deriving rules of thumb from their sequences. random forests are more robust and better suited for categoric (as opposed to binary) classification. we next want to develop a chatbot, currently under construction, that will ask relevant questions in optimal sequence, and extract undiagnosed patients with suspected iei, based on statistical "red flags". we also have preliminary results of this process applied to a usidnet database with over , patients, and are also working on multinomial logistic regression and naïve bayesian classifiers for this and other databases. abstract/case report text objectives: acute viral respiratory infections (avri) are associated with significant healthcare resource use and cost. the use of intravenous immunoglobulin (ivig) may be an effective treatment for immunosuppressed patients and reduce overall healthcare resource utilization. the goal of this study was to assess hospital resource utilization associated with ivig use among patients hospitalized for avri. methods: using data from the - premier hospital database, we identified patients hospitalized with a diagnosis of avri [respiratory syncytial virus (rsv), parainfluenza virus, rhinovirus, or metapneumovirus], and who had an immune deficiency (chemotherapy treatment, transplant, primary immunodeficiency disorder (pidd), specific antibody deficiency, other immunodeficiency, or disorders of the immune or lymphatic systems). patients receiving ivig within the first hours were compared to patients who did not receive ivig at all. due to the nature of the need to better understand the treatment effect associated with ivig, we used an inverse probability weight-based regression model. since there were substantially more controls than cases, we randomly drew , controls. a logistic regression model was developed to adjust for factors associated with the probability of ivig use within hours of admission. this propensity score was then used to weigh subsequent models to assess length of stay (total and icu) using negative binomial models and logistic regression for inpatient death. results: a sample of , immunocompromised inpatients were identified, receiving ivig within the first hours of admission and , who did not receive ivig. the ivig group was older (mean age vs , p < . ), had more antiviral use ( % vs %, p < . ), and had less cancer ( % vs %, p < . ). after adjustment for immunity type (transplant, cancer), rsv, pidd, age, prednisone, antiviral use, ribavirin use, urban hospital setting, teaching status, intubation and lung disease, patients with ivig use had . less days of hospitalization (p= . ) and . less days in the icu (p= . ) than non ivig users. conclusions: this data analysis suggests that hospital length of stay and icu length of stay were significantly shorter for immunocompromised patients hospitalized for acute viral respiratory infections who were administered ivig within the first hours of admission, as compared to patients who did not receive ivig. it is possible that ivig use may have an impact on hospital resource utilization and costs. future prospective studies would help further assess the role of ivig in patients hospitalized with acute viral respiratory infections. associate professor/yale university abstract/case report text background: cd ligand deficiency is an x-linked combined immunodeficiency associated with opportunistic infections and increased risk of malignancies. expansion of memory cd + t-cells with senescent features is known to be associated with chronic immune stimulation including aging, chronic infection and malignancy. cd + t-cell characteristics of cd l deficient (cd ld) patients in relation to their clinical history have not been described. objective: we studied correlation between cd + t-cell senescence with clinical histories of cd ld patients. methods: we analyzed the frequency and phenotypic characteristics of peripheral cd + t-cell subsets in four cd ld patients ( , , and years old (yo)) and healthy controls (hcs). t cell excision circle (trec) counts and telomere lengths of the patients and hcs were measured using quantitative pcr. in-depth analysis of cd + t-cells of the yo patient and hcs was done using high-dimensional cytometer time of flight analysis (cytof). results: three patients ( , and yo) with histories of recurrent infections and poor compliance with immunoglobulin therapy (ivig) showed an increased frequency of effector memory cd + t-cells with the senescent phenotype compared to age matched hcs. whereas yo patient with excellent ivig compliance starting at infancy did not show any senescence phenotypes of the cd + t-cells. the telomere length and trec count of each patient correlated with the degree of cd + t-cell senescence and their current ages, respectively. in-depth analysis showed similar expression patterns of molecules related to senescence and cytotoxicity in cd + t-cells including cd , t-bet, eomes, granzyme b and perforin in the yo patient and mid-elderly hcs. conclusion: our findings suggest that prompt diagnosis and compliance with ivig starting at the infancy may prevent early onset cd + t-cell senescence in cd l deficiency. abstract/case report text introduction: immunoglobulin g -related disease (igg -rd) is an immune-mediated fibroinflammatory condition that affects multiple organs. when igg -rd is found in the ocular adnexa, the term "igg related ophthalmic disease (igg -rod)" is used. objective: our case describes a patient with igg -rod without systemic involvement. case: mr. x is a -year-old male with a pmh of cml (on imatinib) and allergic rhinitis who presented to clinic with orbital swelling for twenty years. his swelling had always been responsive to steroids, but would return once steroids were tapered. patient was diagnosed with biopsy proven cml in and is currently taking imatinib. because his peri-orbital edema persisted, a right lacrimal gland biopsy was done which showed "marked lymphocytic infiltrate of soft tissue with lymphoid follicles, many plasma cells, and eosinophils. no atypical histiocytes." flow cytometry was negative for malignancy. results: crp . mg/l. esr mm/hr. igg elevated at mg/dl. ct chest from and ct chest, abdomen, pelvis from were without fibrotic changes. assessment: when diagnosing igg -rd, we categorize diagnosis into three levels (possible, probable, or definite) by three criteria (clinical manifestation, elevated serum igg , and histopathology). this is detailed as follows: clinical exam showing organ specific swelling or masses, elevated serum igg (> mg/dl), and histopathology with either lymphocyte and plasmacyte infiltration and fibrosis or infiltration of igg + plasma cells (ratio of igg +/igg+ cells ≧ % and ≧ igg + plasma cells per high power field). not all these components are required for diagnosis, but meeting histopathologic criteria makes diagnosis more probable. our patient's disease was localized to his eye, and patients with igg -rod have unique diagnostic criteria. these criteria are similar to the criteria for igg -rd, but emphasize enlargement of the ocular adnexa, less frequent fibrosis, and ≧ igg + plasma cells per high power field. our patient's histopathology revealed a lymphoplasmacytic infiltrate, but lacked storiform fibrosis or obliterative phlebitis. his serum igg level was mg/dl, and his biopsy was positive for an igg +/igg+ ratio of % and more than igg + plasma cells per high power field. based on this, he meets criteria for igg -rod. conclusion: igg -rod is a rare condition that is usually associated with systemic organ involvement. our case is unique, as no systemic disease has been detected. we also suspect our patient has been living with igg -rod for several years, as his orbital swelling began in high school. it is important to note that he has been on imatinib, a tyrosine kinase inhibitor, for treatment of his cml. imatinib inhibits c-abl and platelet-derived growth factor receptor, tyrosine kinases involved in profibrotic pathways. patient's lack of fibrosis could also be due to his longstanding use of this drug. it is also possible that he has a rare form of igg -rod without systemic involvement. a limited number of such cases have been reported, but no consensus has been made on why disease course was localized. our patient was started on rituximab, and his serum igg decreased to mg/dl after the first cycle. we hope his disease achieves remission. informed consent: informed consent was obtained from all individual participants included in the study. abstract/case report text platelet abnormalities with eosinophilia and immune-mediated inflammatory disease (plteid) is a recently discovered combined immunodeficiency with inflammatory and allergic manifestations with few cases reported. we describe a female patient with compound heterozygous mutation in arpc b gene with suggestive clinical findings of plteid. a -year-old girl presented with chronic diarrhea since neonatal period, with bloody stools and failure to thrive. she also presented atopic dermatitis, recurrent cutaneous and mucosal ulcers, recurrent respiratory infections ( episodes of otitis media, pneumonias) and many episodes of mucocutaneous candidiasis. family history revealed a sibling deceased in the second month of life, who presented a similar clinical picture and a paternal uncle and second degree cousin that died in the first year of life. there is no history of consanguinity. laboratory evaluation revealed peripheral eosinophilia ( /mm ), normal platelet numbers with low platelet volume ( , fl -reference value , - , fl), normal igm levels with elevated igg ( mg/dl-rv - mg/dl), iga ( mg/dl -rv - mg/dl) and ige ( iu/ml -rv a) associated with plteid. t h e i n f a n t r e c e i v e s a n t i m i c r o b i a l p r o p h y l a x i s w i t h sulfamethoxazole-trimethoprim and fluconazole, intravenous immunoglobulin replacement and was referred to hematopoietic stem cell transplantation (hsct). this case was the first one described in brazil and highlights the importance of seeking for a genetic diagnosis in patients with complex clinical phenotypes. precise diagnosis can impact on treatment approach. live vaccines are generally contraindicated in patients with combined immunodeficiency (cid). however, in less severe cid, such as partial dgs, those vaccines can be considered depending on the immunologic status of the patient. there are recommendations regarding to measles, mumps, rubella (mmr) and varicella vaccines, but yellow fever vaccine (yfv) is generally contraindicated in this population. considering the severity of the yellow fever disease and the absence of specific treatment, the use of this vaccine is an important topic for debate in cases of patients from endemic areas. objective: this study aimed to describe the use of yfv and other live attenuated vaccines in patients with dgs, associating it with their immunological profiles and the presence of adverse effects. methods: retrospective study of medical records of patients with dgs confirmed by mlpa or fish, followed in a pediatric reference center for primary immunodeficiencies between and . collected data included: demographic characteristics, medical history, history of immunization with live vaccines, postvaccination adverse reactions and immunological profile, including immunoglobulins levels, serologic vaccination responses, lymphocyte immunophenotyping, lymphocyte proliferation responses to mitogens and prophylactic treatments (antibiotic or immunoglobulins). results: thirty-five patients with confirmed dgs and median age of years ( - y) were included ( m: f). thirty-three children ( %) received mmr vaccine: nine presented t lymphopenia. two of the patients had cd < , one of them with normal mitogenic proliferation response and the other was not tested. three of the patients had low immunoglobulins levels ( / low igg, / low igm and / low iga), and one of them received intravenous immunoglobulin (ivig). twentynine of had normal serologic vaccination responses. adverse effect was only reported by one patient, who had one episode of fever after the administration of all vaccines. yellow fever vaccine was administrated to children ( %): had t cell l y m p h o p e n i a ( b u t c d > ) , a n d a n o t h e r p a t i e n t h a d hypogammaglobulinemia and received ivig and prophylactic antibiotics. twelve of showed adequate serologic responses to mmr and hepatitis b. only patient reported mild reaction (tremors) two days after the yfv administration. the same patient had normal t cells, immunoglobulins and vaccine responses. twenty patients ( %) received bacillus calmette-guerin vaccine (bcg), ( %) received oral polio, ( %) rotavirus and ( %) received varicella vaccine. no severe adverse events were documented in any patient that received live vaccines, and no patient developed measles, mumps, rubella or yellow fever diseases as a consequence of administration of the vaccine. conclusions: in this cohort of pediatric patients with dgs, yfvand other live vaccines were well tolerated, and no severe adverse events were reported, suggesting that widespread contraindication of yfv may endanger unvaccinated patients with less severe phenotype living in endemic areas. immunological evaluation and individualized decisions are always recommended, and further studies are needed to assess the safety of the yfv in this pediatric population. abstract/case report text introduction: lad-i is a rare inherited disorder of leukocyte (primarily neutrophil) adhesion to endothelial cell surfaces, migration, and chemotaxis resulting from itgb gene mutations encoding for the β -integrin component, cd . severe lad-i (i.e., cd expression on < % of neutrophils) is characterized by recurrent serious infections, impaired wound healing, and childhood mortality. although allogeneic hematopoietic stem cell transplant (allohsct) is potentially curative, its utilization and efficacy are limited by hla-matched donor availability and risk of graftversus-host disease (gvhd). rp-l - (clinical trials.gov # nct ) is a phase / open-label clinical trial evaluating the safety and efficacy of autologous cd + cells transduced with a lentiviral vector (lv) carrying the itgb gene encoding for cd (chim-cd -wpre) in severe lad-i. methods: pediatric patients ≥ months old with severe lad-i (demonstrated by cd expression on < % neutrophils and at least one prior significant bacterial or fungal infection) are eligible. peripheral blood (pb) hematopoietic stem cells are collected via apheresis after mobilization with granulocyte-colony stimulating factor (g-csf) and plerixafor. cd + hspcs are selected, transduced with chim-cd -wpre lv, and cryopreserved. myeloablative conditioning with busulfan (therapeutic drug monitoring (tdm) dosing with adjustments to enable target area under the curve (auc)) is administered over days, followed by infusion of the thawed investigational drug product (rp-l ). patients are followed for safety assessments including replication competent lentivirus (rcl) and insertion site analysis (isa), and for efficacysurvival to age ( months) and at least -year post-infusion without allohsct, increase in neutrophil cd expression, pb vector copy number (vcn), decrease in infections and/or hospitalizations, and resolution of skin or periodontal abnormalities. results: an initial lad-i patient (age years) with recurrent severe infections and documented itgb mutations has been treated as of november . baseline cd , cd a, and cd b expression were < %. mobilization and apheresis procedures were performed successfully and busulfan conditioning was administered at the target auc. investigational product was comprised of . x e cd + cells/kg with vcn of . copies/cell (liquid culture), and was infused without complications. no serious treatment-emergent adverse events were reported. neutrophil engraftment ( consecutive days of anc ≥ ) was observed days post-infusion. pb pmn cd expression months posttreatment was . % with comparable cd a and cd b expression levels; pb cd (myeloid) vcn at . months was . . safety and efficacy data months post-treatment will be available at the time of presentation, in addition to preliminary data regarding a potential additional patient. conclusion: preliminary evidence demonstrates that rp-l enables itgb genetic correction with robust cd /cd neutrophil expression in this frequently fatal primary immunodeficiency. abstract/case report text introduction: the complement system plays an integral role in the innate immune system and links innate and adaptive immunity. complement deficiencies, hereditary or acquired, are rare. acquired deficiencies are more prevalent, occurring in nephrotic syndrome, reduced hepatic synthesis or transiently in sepsis/viremia. they are also seen in the presence of autoantibodies known to cause depletion of complement factors, such as c nephritic factor (c nef). c deficiency is associated with infection susceptibility, particularly to encapsulated bacteria, and immune complex disease. case description: a year old male was evaluated for recurrent infection. in childhood, he had recurrent sinusitis, otitis media requiring tympanostomy tube placement and persistent pharyngitis despite tonsillectomy. as a teenager, he developed glomerulonephritis, progressing to end stage renal disease and requiring transplant at age . the kidney allograft failed years later, with biopsy demonstrating recurrent glomerulonephritis. the patient was transitioned to peritoneal dialysis and later hemodialysis, due to recurrent pd-related infections. his adult course was complicated by recurrent methicillin sensitive staphylococcal aureus (mssa) catheter and soft tissue infections (cellulitis and abscess), sinusitis, sepsis (streptococcal, mssa and tularemia), multifocal pneumonia and a left below knee amputation for osteomyelitis that required revision surgery. patient reported other autoimmune phenomena including a presumptive diagnosis of vasculitis and possible lupus-like syndrome. the constellation of recurrent infections and autoimmune features was most concerning for an early complement deficiency. prior work up was notable for low c , ch and ah with normal c , factor h and factor i. extensive laboratory work up revealed normal c q, c level and function, serum immunoglobulins, vaccine titers, factor b and factor d levels. atypical hus (ahus) panel revealed a heterozygous silent variant in exon of cfh and a heterozygous polymorphism within an intron in mcp/cd , seen with increased prevalence in the patient population with ahus. wes was notable for a variant of uncertain significance in the vcl gene only. c level and function were markedly decreased, alongside low ch and ah . both sc b- level and c nephritic factor were elevated. a diagnosis of acquired c deficiency due to c nef was made and patient was started on bactrim prophylaxis. he has remained free of serious infection since starting antibiotic prophylaxis. discussion: c nef stabilizes the alternative pathway c convertase, c bbb, increasing its half-life and blocking dissociation. this leads to unregulated consumption of c with subsequent deficiency. c nef has been associated with c glomerulopathy, infection and partial lipodystrophy. however, there is marked heterogeneity in clinical phenotypes with reported asymptomatic individuals. our patient's glomerulonephritis likely represents c glomerulopathy. case reports and series of successful treatment of c glomerulopathy with rituximab and eculizumab have not commented on immune outcomes beyond the kidney. other potential therapeutic strategies include plasma cell depletion with either bortezomib or daratumumab. further study is needed to evaluate these therapies influence on both reversal of c depletion and overall impact on immune function in the setting of c nef. abstract/case report text introduction: patients with heterozygous signal transducer and activator of transcription (stat ) gain of function (gof) pathogenic variants exhibit an array of phenotypes including susceptibility to viral, bacterial, fungal and mycobacterial infections, autoimmunity, and cancer predisposition. progressive disseminated histoplasmosis (pdh) is well-described to affect infants. however, no reports have evaluated underlying monogenic immune dysregulation in previously healthy infants presenting with pdh. we report an infant who presented with pdh and associated hemophagocytic lymphohistiocytosis (hlh) leading to the diagnosis of a heterozygous stat gof mutation. case report: a previously healthy -month old male presented with persistent fever, pancytopenia, transaminitis, elevated ferritin, hepatosplenomegaly and coagulopathy. his clinical and laboratory evaluations were concerning for hlh syndrome. he had no prior history of immune hyperactivation or atypical infections. secondary causes of hlh were investigated, and patient was diagnosed with pdh based on marked histoplasma antigenemia. targeted genetic testing did not reveal a genetic etiology of familial hlh. he was successfully treated with a pulse and taper of dexamethasone as well as liposomal amphotericin b with transition to itraconazole. immunologic evaluation at the time of initial presentation demonstrated increased mean channel fluorescence for both perforin and granzyme noted in his nk cells. his nk function was decreased; however, he had a normal cd a degranulation assay. his b-cell panel demonstrated low non-switched memory b-cells, low switched memory b-cells and low total memory b-cells. given his extreme immune activation with histoplasmosis, abnormal immunologic testing, and persistent lymphopenia despite resolution of his infection, a primary immunodeficiency next generation sequencing panel was sent. the results demonstrated a pathogenic variant in stat (c. c>t; p.ala val). this single nucleotide variant has been previously shown to be pathogenic (clinvar). abstract/case report text introduction: cytotoxic t lymphocyte antigen- (ctla- ) is known to have an important role as a negative regulator of immune responses, participating in the control of regulatory t cells and effector t cells. in mice its absence is associated with fatal autoimmunity and several ctla- mutations, leading to low or absent ctla- expression, have been shown in humans to be associated with a phenotype that includes hypogammaglobulinemia (with recurrent respiratory infections) and several manifestations of autoimmunity (enteropathy, granulomatous lymphocytic interstitial lung disease, organ infiltration, splenomegaly, autoimmune cytopenias, lymphadenopathy, amongst others), in an autosomal dominant mode of transmission. one of the published mutations, c.c t, that results in an alanine to valine substitution (p.a v), with a highly conserved alanine at that position, had a cadd score of and was associated with the phenotype above, and was shown to be associated with a low expression of ctla- on regulatory t cells and with low ctla- function (reduction of ctla- -mediated transendocytosis). methods: after irb approval, we searched for ctla- mutations present in the biome biobank· biorepository, containing whole exome sequencing data on patients, with data obtained using illumina· v hiseq sequencing platform. sifting through all the ctla mutations in the data, we identified four patients with the c.c t mutation described above. extensive chart review of the four patients was performed. results: four patients were found with the ctla- c.c t mutation. none of them had any of the described phenotypical characteristics of ctla- deficiency. patient is a -year-old male with history of coronary artery disease, atrial fibrillation, stroke, hypertension, brain aneurysm, chronic kidney disease, gout and depression. patient is a -year-old female with history of morbid obesity. patient is a -year-old female with history of hypertension, obesity, pre-diabetes, dyslipidemia and iron deficiency anemia. patient is a -year-old female with history of peripheral artery disease, hypertension, dyslipidemia, chronic kidney disease and lung cancer. conclusion: prior literature has attempted to characterize the clinical penetrance of ctla- mutations, suggesting it to be around %, with that number applying to different mutations in ctla- mutation carriers. we screened a large biorepository of more than thousand patients for ctla- patients and identified four patients that carry one of the best described ctla- mutations, previously validated from a functional standpoint and associated with a severe phenotype. none of the four patients demonstrated any of the previously described phenotypical characteristics, and all four have ages above the median age of onset of years. with the increasing use and broad population application of genetic studies, it is crucial to define the value of identifying presumed pathogenic variants in the absence of the adequate phenotype, with all the prognostic, therapeutic and ethical considerations it may imply. prior case reports of pil patients with b-cell malignancies have discussed treatment regimens with chemotherapy, radiation, and/or surgery, but neither the use nor the outcomes of allogeneic hematopoietic stem cell transplantation (hsct) in the management of recurring b-cell malignancies have been readily reported. case description: a -year-old man with pil and an accompanying history of lymphopenia, hypoproteinemia, hypoalbuminemia, and hypogammaglobulinemia was diagnosed with diffuse large b-cell lymphoma (dlbcl) of the liver following a preceding history of burkitt lymphoma of the ileum at years of age and dlbcl of the liver at years of age, in which each malignancy was genetically distinct. in addition, the patient had a history of benign nodular adenomatoid hyperplasia of the thyroid at years of age that required a hemi-thyroidectomy. treatment considerations for the patient included chimeric antigen receptor t-cell therapy, autologous hsct, and allogeneic hsct, in which allogeneic hsct was ultimately pursued. prior to hsct, the patient was lymphopenic ( cells/microliter) with significant t-cell lymphopenia ( cells/microliter) and an increased proportion of memory t-cells ( % of his cd + t cells were cd ro+), as well as hypogammaglobulinemic (igg mg/dl; iga mg/dl; igm mg/dl). immediately following treatment of his dlbcl with rituximab, ifosfamide, carboplatin, and etoposide, the patient underwent a matched-related sibling donor hsct with a preparative regimen of busulfan, thiotepa, and fludarabine. now months status-post hsct, the patient has maintained full-donor chimerism and has no evidence of graft-versus-host disease or malignancy. as expected, hsct has not corrected abnormalities in certain parameters associated with his pil, as he continues to display significant hypoproteinemia, hypoalbuminemia, and hypogammaglobulinemia, but he has an improved lymphocyte count ( , cells/microliter). discussion: there is no definitive or curative treatment for pil; furthermore, the genetic etiology of pil remains unknown. supportive regimens to help mitigate or offset manifestations of pil exist, such as adherence to a low-fat diet with medium-chain triglyceride supplementation, but there are no therapies available to prevent or reduce the risk of developing bcell malignancies in this patient population. although previous case reports have detailed successful treatment of b-cell malignancies in pil patients with chemotherapy, radiation, and/or surgery, there are no published consensus guidelines regarding management of b-cell malignancies in the setting of pil, especially if recurrent in nature. for non-pil patients with chemotherapy-refractory disease, or recurrent disease following autologous hsct, allogeneic hsct is a potentially curative option. herein, we describe a pil patient with a history of multiple b-cell malignancies who underwent a successful allogeneic hsct, indicating that allogeneic hsct may be an effective treatment option for similarly affected patients. abstract/case report text introduction diarrhea in young infants is common and generally self-limited. in persistent cases, the differential diagnosis is broad and includes infections, food protein-induced allergic proctocolitis, congenital diarrheas and enteropathies. in addition to monogenic inflammatory bowel diseases, many cellular, humoral, and combined immunodeficiencies should be considered, including but not limited to cvid, ipex and ipex-like phenotypes, lad, dyskeratosis congenita, intestinal lymphangiectasia, omenn syndrome, cartilage hair hypoplasia, cgd, il- axis defects, aid deficiency and wiskott-aldrich syndrome. case presentation a full term infant born after an uncomplicated pregnancy to nonconsanguineous honduran parents presented with non-bloody, non-bilious vomiting and dehydration at days of life. the infant later developed frequent loose stools, some of which were bloody, and failure to thrive. his family and prior medical history, including newborn screen, were normal. an extensive workup was initiated which showed: -persistent and severe anemia with a hemoglobin nadir of . mg/dl -hypoalbuminemia requiring multiple infusions -elevated alpha- -antitrypsin and calprotectin level in stool -profound hypogammaglobulinemia with normal iga, igm and ige for age -normal gross and histologic findings on esophagogastroduodenoscopies and colonoscopies besides a gastric ulcer thought not be the cause of his anemia -normal abdominal imaging including ultrasound, ct angiography and mri -no source of bleeding on meckel scan or exploratory laparotomy -normal dhr assay, g pd level and positive myeloperoxidase stain -immunophenotyping showing t cell lymphocytosis affecting cd + more than cd + compartment, with normal lymphocyte proliferation to mitogens -normal sweat chloride level genetic testing was initiated with a targeted immunodeficiency panel which showed variants of unknown significance in adar, dock , lyst, ptprc and tbx genes, none of which adequately explained his presentation. whole exome sequencing showed that he was a compound heterozygote in the dgat gene. a pathogenic variant c. + t>c (ivs + t>c) was inherited from the father and a likely pathogenic variant c. g>c (p.r p) was inherited from the mother. patient was diagnosed with dgat deficiency, an inborn error of lipid metabolism resulting in protein-losing enteropathy (ple). under gastroenterology's guidance, a low fat diet was initiated, resulting in rapid improvement in stool consistency, weight gain, albumin level and stool alpha- -antitrypsin level. he remains on subcutaneous immunoglobulin replacement therapy for ongoing hypogammaglobulinemia. conclusion: protein-losing enteropathies commonly present with intractable diarrhea and significant laboratory derangements due to malabsorption including hypogammaglobulinemia. as a result of these findings and since many of the etiologies are immunologic in origin, immunologists are an integral part of the evaluation of such cases. in cases where immune system interrogation reveal normal results, genetic testing is crucial in guiding the diagnosis. in our case, whole exome sequencing not only provided the diagnosis but also characterized a variant that was previously of unknown significance as likely pathogenic. dietary management provided rapid improvement in growth and nutritional status. ongoing monitoring will reveal if this management also assists in igg level maintenance and hematologic abnormalities or if even more stringent control of dietary fat will be required abstract/case report text background: foxp gene mutations are associated with immune dysregulation polyendocrinopathy x-linked (ipex) syndrome, a rare xlinked monogenic disease of immune dysregulation and autoimmunity. the classic presentation consists of severe enteropathy, dermatitis, and endocrinopathies (commonly early onset insulin dependent diabetes mellitus). clinical presentation and severity can be variable even in family members with the identical variant. we present a patient with ipex symptomatology and a hemizygous variant in the polyadenylation (polya) signal of foxp that is classified as a variant of uncertain significance (vus). this specific variant was reported in a single case study in which the patient improved after hematopoietic stem cell transplantation (hsct). case presentation: a month-old ex -week gestation boy was admitted with lethargy, hypovolemia, electrolyte disturbances, and acute kidney injury. he developed persistent diarrhea and vomiting after receiving rotavirus vaccine. his family history is significant for early deaths of three maternal unclesone stillborn, one death at months and another at years from unknown gastrointestinal problems. he demonstrated peripheral eosinophilia (to . k/ul), elevated ige, and anemia requiring multiple transfusions. he developed severe enteropathy with hypoproteinemia requiring total parenteral nutrition, continual albumin infusions and maintenance of npo. he had generalized edema, respiratory distress requiring high flow nasal cannula, and repeatedly spiked fevers with negative infectious evaluation. acute kidney injury improved but renal ultrasound showed persistent nephrocalcinosis. endoscopy yielded biopsies demonstrating duodenitis with severe villous atrophy, scanty isolated intraepithelial eosinophils and neutrophils, a few crypts with mucin, reactive epithelial changes and increased lamina propria eosinophils. colon biopsies showed mucosa with focally increased lamina propria eosinophils with scanty neutrophils and surface epithelium without cryptitis. esophagitis with reactive epithelial changes, spongiosis, and many intraepithelial eosinophils was also present. the patient's lymphocytes showed unremarkable proliferation to pha and pwm. cd + cd + t cells demonstrated intracellular foxp expression by flow cytometry. a commercially-available immunodeficiency targeted panel revealed that he was hemizygous for a vus in foxp (exon , c.* a>g non coding). this variant is also referred to as an aauaaa>aauaag or aataaa>aataag change in the polya site. he was also heterozygous for vus at these additional loci: cd a c. c>t abstract/case report text introduction: implantation of allogeneic cultured thymus, partially depleted ex vivo of t cells, can result in naïve t cell development in patients with complete digeorge syndrome (dgs). in a few patients, early and transient skin rash, often characterized as "atypical dgs" or late autoimmune manifestations have been reported following implantation. here we describe a patient with complete dgs who developed immune reconstitution inflammatory syndrome (iris) or atypical dgs following thymus implantation. case description: a female patient was diagnosed at birth with complete dgs due to absent t cell receptor excision circles (trec), hypoplastic thymus, profound hypocalcemia with hypoparathyroidism and cardiac defects. the patient also had microretrognathia, oral motor dysfunction, sialorrhea, recurrent aspirations and reflux requiring a gastro-jejunum feeding tube, low-set ears with right ear microotia, semicircular canals atresia, alopecia and mal-rotated kidneys. prior to thymus implantation, the patient was thriving, had no skin rash, no eosinophilia and no t cells. detailed genetic analyses, did not reveal a cause for her syndrome. at months of age pulmonary aspergillosis was diagnosed presumptively. at months of age the patient received an allogeneic t-cell depleted thymus implant from a male donor, without prior conditioning or post-implantation immune suppressive medications. the procedure was uneventful and the patient returned home after days. results: four months after implantation, a pruritic maculopapular rash appeared on the head and trunk that spread to the extremities including the palms and soles. there was no lymphadenopathy or splenomegaly. an infectious etiology could not be found. eosinophilia and an increase in liver enzymes were noted. there was an increase of cd + and cd + t cells with predominantly memory phenotype, which had been undetectable month earlier. analysis of t cell diversity showed a restricted repertoire with expansion of two v-beta families. there was no evidence of donor cells to suggest graft versus host disease. skin biopsy showed minimal superficial perivascular inflammatory infiltrate composed mainly of cd + histiocytes and rare cd + t cells. the patient was treated with prednisone and cyclosporine. a liver biopsy was performed weeks after initiation of treatment that showed moderate and diffuse peri-portal ductular reaction but no duct associated lymphocytic infiltrate or significant duct epithelial injury or ductopenia. the skin rash rapidly resolved with desquamation, while the liver enzyme abnormalities persisted for two more months. cyclosporine and prednisone were weaned over months. t cell numbers, their response to stimulation and diversity have since normalized, as well as trec and naïve t cell production. the patient is producing appropriate antibodies to protein and polysaccharide vaccines. sixteen months after implantation the patient developed grave's disease with markedly elevated free-t , undetectable tsh and elevated antibodies to the thyroid receptor, which rapidly normalized with ongoing methimazole treatment. the patient is currently months after the implantation and is free of infections, thriving and developing appropriately. conclusions: this patient developed atypical dgs or iris, often associated with autologous and allogeneic hematopoietic stem cell transplants, organ transplants or effective treatment of hiv, after successful thymus implantation for complete dgs. abstract/case report text congenital disorders of glycosylation are a rare group of genetic disorders due to defects in protein glycosylation. phosphoglucomutase (pgm ) is an enzyme necessary for the synthesis of uridine diphosphate n-acetylglucosamine, an important precursor for protein glycosylation. patients with autosomal recessive pgm deficiency have a multisystemic disorder characterized by a neurologic impairment and clinical features classically observed in autosomal dominant hyper-ige syndrome due to stat mutations; including recurrent pneumonias, skin abscesses, elevated levels of ige, and abnormalities in connective tissues and bones. we hypothesized that gp , a highly glycosylated protein and coreceptor of the cytokine il- , would be weakly expressed on pgm deficient cells, due to impaired glycosylation. we studied pgm -deficient patients from kindreds and showed that il- -driven stat phosphorylation was impaired in their pbmcs and ebv-transformed b cells. accordingly, the induction of socs target gene was significantly decreased. in contrast, the patients had normal stat phosphorylation and socs induction downstream of il- , a cytokine whose signaling is independent of gp . flow cytometry and immunoblotting showed significantly lower gp expression in peripheral t-cells and ebv-transformed b cells from pgm -deficient patients compared to healthy donors. we did also show that in vitro inhibition of n-glycosylation, using tunicamycin in ebv-transformed b cell line from healthy donor, alters gp -mediated signaling. collectively, our findings demonstrate that defective glycosylation in pgm -deficient patients results in reduced expression of gp and consequently, impaired gp dependent stat phosphorylation and defective il- signaling. this may account for the overlapping clinical features shared by pgm and stat deficient patients. abstract/case report text introduction: there are no known effective therapeutic modalities for patients hospitalized with moderate to severe acute viral respiratory infections, and treatment is primarily supportive. intravenous immunoglobulin (ivig) has been reported in limited cases to be used in this setting, especially in immunocompromised patients. the primary objective of this retrospective study is to compare clinical and economic outcomes among immunocompromised patients hospitalized with viral respiratory infections who received ivig to those who did not receive ivig at a large academic center hospital. methods: we performed a double-center, retrospective cohort study of all immunocompromised patients who were hospitalized for acute documented respiratory viral infections between and . we divided patients into two groups: those who received ivig therapy for respiratory infections, and those who did not receive ivig therapy. data on age, gender, immune status, viral type, immunosuppression type, respiratory support, microbiological data, length of hospital stay (los), icu los, as well as death and readmission rates were extracted from medical records. in order to adjust for severity bias typically present in observational data such as these, we employed inverse probability weighting (ipw) using all collected baseline covariates. outcomes (death, length of stay in hospital and icu, readmission) were examined using a series of logistic and poisson regression models adjusting for baseline covariates and employing ipw. results: a total of individual hospital admissions were analyzed; patients received ivig and did not receive ivig. there were no significant differences between the two groups in terms of mean age, gender . average age was . , % were female, . % were transplant patients of which . % had lung transplant, . % had liver transplant, . % had bone marrow transplants (bmt), . % had kidney transplant, . % had heart transplant and . % had both solid organ and bmt. . % of patients had a hematologic malignancy, and . % had a primary immunodeficiency. the most common isolated respiratory virus w a s r h i n o v i r u s ( . % ) , f o l l o w e d b y r s v ( . % ) , parainfluenza ( . %) and metapneumovirus ( . %). overall, the use of ivig as associated with a significantly shorter icu length-of-stay, with an (or=- . , p= . ), and a higher hospital readmission rate. in the sub-analysis of patients who received ivig within the first hours of hospitalization (n= ), ivig use was associated with a significantly shorter icu los (or=- . , p= . ), significantly shorter overall hospital los (or=- . , p= . ), and no significant change in readmission rate. conclusions: to our knowledge, this is the first retrospective cohort analysis evaluating the effect of ivig in immunocompromised patients hospitalized with respiratory viral infections. the results suggest that immunocompromised patients receiving ivig may have a shorter hospital and icu los, especially if ivig is provided within the first hours of admission. this may result in reduced healthcare costs. this study is limited by its retrospective nature, and the potential bias that patients treated with ivig are sicker to start with. future prospective studies are suggested to further evaluate these findings. ( , ) . the most common precipitant in children is medication, followed by infection ( , ) . although a clear association between mycoplasma pneumoniae and sjs has been established, there is a scarcity of literature exploring the role of this infection in recurrent sjs in children ( ) ( ) ( ) . case presentation: a -year-old female with prior history of sjs was admitted for mucosal and skin lesions in the setting of community acquired pneumonia. her past medical history included sjs with eye involvement, secondary to mycoplasma pneumoniae (ig m positive), occurring five years prior to this admission. she also had frequent episodes of acute otitis media and sinusitis in early childhood. family history was negative for immunodeficiency. her clinical presentation included respiratory symptoms and fever for days treated with ceftriaxone, followed by cefdinir and levofloxacin. her fever improved the day prior to admission, but she developed conjunctival injection, ocular pain, and ulcerative lesions in her mouth and nares. on physical examination, she had low grade fever with mucosal lesions including conjunctival erythema with serous discharge, painful blisters and denudated skin in lips, perioral area, nares, tongue and oropharynx. initial testing included negative blood hsv pcr, blood culture, rapid antigen testing for group a streptococcus and influenza a/b, and elevated crp in . mg/dl and esr mm/hr. right lower lobe p n e u m o n i a w a s c o n f i r m e d w i t h a c h e s t r a d i o g r a p h . nasopharyngeal pcr and serum igm were positive for mycoplasma pneumoniae. she had a mildly elevated anticardiolipin igm ( mpl), a mildly decreased c ( mg/dl) and a negative ana. she was diagnosed with recurrent sjs secondary to mycoplasma pneumonia infection. she completed treatment with levofloxacin for mycoplasma pneumonia, and received cyclosporine and high-dose methylprednisolone. she had bilateral amniotic membrane transplantation to prevent corneal ulceration . she was discharged after clinical improvement, and recurrent oral lesions were noted at followup. immunological work up as an outpatient revealed normal serum immunoglobulins, normal lymphocyte subsets and low pneumococcal titers with adequate response post-vaccination. sjs secondary to mycoplasma pneumonia infection has predominance of mucosal involvement over rash, which was observed in our patient ( , ) . some case series reported a recurrence of sjs up to % within a -year follow up. almost half of patients with recurrent sjs developed multiple sequelae ( , ) . early diagnosis of sjs, especially in those with prior history of sjs, helps to provide appropriate supportive care, monitoring of complications and treatment of possible superinfections ( , ) . conclusions: there is limited information in the literature regarding the role of mycoplasma pneumoniae associated recurrent sjs in children. it is possible that these episodes are triggered by and/or immune predisposition. there is ongoing discussion regarding whether these clinical presentation should be labeled sjs secondary to mycoplasma pneumonia infection or, depending of the skin involvement, m. pneumonia-associated mucositis (mpam) and m. pneumonia-induced rash and mucositis (mirm) ( , , ). mycoplasma should be treated appropriately in patients with recurrent sjs. abstract/case report text background: growing access to genetic testing has facilitated the genetic evaluation of primary immunodeficiencies but has also greatly increased the number of variants of uncertain significance (vus) encountered in clinical practice. interpreting the significance of vus requires multiple lines of evidence. w e d e s c r i b e a n e u t r o p e n i c i n d e x p a t i e n t w i t h hypogammaglobulinemia, unusual hpv susceptibility, and dual heterozygous pathogenic loss-of-function nfkb and heterozygous missense cxcr vus. family analysis showed the nfkb variant was inherited from his mother, while the novel cxcr variant was present in his father and sister. all four patients presented with recurrent infections, warts, and hypogammaglobulinemia. (figure ) the nf-κb gene encodes p /p transcription factor of the canonical nf-κb pathway, the most common autosomal dominant monogenic cause of common variable immunodeficiency (cvid). cxcr is a g-protein-coupled chemokine receptor with cxcl as cognate ligand. autosomal dominant pathogenic gain-offunction cxcr variants lead to impaired receptor downregulation and retention of neutrophils and other leukocytes in the bone marrow defining whim (warts, hypogammaglobinemia, infections, and myelokathexis) syndrome. all cxcr pathogenic variants truncate the carboxyl-tail of the cxcr receptor, a region responsible for receptor internalization, with the exception of one missense non-truncating variant p.e k. case series: the proband (p ) is a -year-old male with history of recurrent bacterial respiratory tract infections, warts, moderate neutropenia, thrombocytopenia and hypogammaglobulinemia requiring immunoglobulin replacement therapy (igrt). bone marrow biopsy didn't show myelokathexis. next-generation panel sequencing identified a novel heterozygous missense cxcr (c. c>a, p.s y) vus. the serine residue is highly conserved up to zebrafish. this variant was present in heterozygous form in two cases in gnomad database ( , alleles). additional whole-exome sequencing revealed a heterozygous pathogenic nfkb variant (c. dup, pa sfs* ) located in the nterminal rel homology domain, consistent with nfkb loss-offunction. both, the patient's sister (p ) and their father (p ), carry the heterozygous cxcr vus but not the pathogenic nfkb variant, and have history of warts, hypogammaglobinemia, and recurrent infections. the hpv susceptibility is particularly striking in p manifesting by genital warts and hpv-positive oropharyngeal cancer. bone marrow evaluation didn't identify myelokathexis in p (p is pending). the mother of the index case (p ) has cvid requiring igrt and immunomodulation. she shares the nfkb variant with p but is negative for the cxcr vus. extensive t and b cell phenotyping revealed low class-switched memory b cell count ( - counts/ul) in all subjects, and loss of transitional and mature naïve b cells in p and p with nfkb variant. proband b cells showed the highest tendency for apoptosis ( - %) within the family. we describe members of a family with similar presentation (infections, hypogammaglobinemia, warts), however variable combination of nfkb and cxcr variants, where either genetic defect or their combination could explain the clinical phenotype. biochemical consequence of our novel cxcr variant is pending. as the proband showed the most severe immune phenotype and neutropenia, we hypothesize that cxcr has a synergistic effect on nkfb loss-of-function. the contribution of cxcr vus of the clinical phenotype of the two other family members is yet to be determined. background: the yield of diagnosis by exome sequencing for some primary immunodeficiencies (pid) has been less than the typical diagnostic rate for clinical exome analysis (~ - %). the relatively low diagnostic rates for certain subtypes of the pids may be attributed to variable expressivity and/or an incomplete understanding of the genetic basis, among others. additionally the extent of multiple diagnoses and phenotypyic expansion in pid is not well explored. cohorts with highresolution clinical and genetic data are instrumental for exploring these questions. we evaluated the use of human phenotype ontology (hpo)annotated datasets to systematically address the prevalence of these issues using a cohort of individuals with pid who participated in research exome sequencing at the niaid. results: we generated a phenotype dataset of individuals with pids by annotating the clinical features of these subjects obtained from electronic health records (ehr) with hpo terms. exome sequencing of these individuals identified probands with a pathogenic or likely pathogenic (p/lp) variant in a gene associated with the respective clinical presentation. we identified probands where the same gene harbored a p/lp variant in at least three unrelated individuals. we used the clinical and genetic data of individuals in the following areas: ) we identified p/lp variants in aire, pik cd, nlrp , fas, ctla , gata , cybb, stat and tnfrsf b in at least ten patients that explained their clinical presentations. this dataset allowed us to characterize variable expressivity of diseases associated with these genes by capturing the variability in the observed hpo terms among probands with p/lp variants in the same gene. dimensional reduction of clinical features of probands allowed us to cluster patients sharing similar phenotypic profiles. we found clinical presentation of individuals with monoallelic p/ lp variants in aire were relatively less variable and clustered more compactly compared to that of individuals with gata variants. ) the extent of multiple diagnoses in pid is not well explored. the benchmark cohort we developed allowed us to identify candidates for multiple diagnosis or phenotype expansion by comparing the phenotype profile of each patient expressed in hpo terms to the hpo terms typically observed for a given pid. for example, we identified gain-of-function pathogenic variant in pik cd in a patient that explained the clinical features of the pid observed in this patient. however, the patient also displayed developmental delay, congenital hemiplegia, cerebral palsy and absent speech. these features are not known to be associated with pik cd variants, making this individual a candidate for > genetic diagnoses. conclusions: we developed a benchmark dataset where clinical features of patients were described using hpo terms. this dataset allowed us to quantify variable expressivity for certain pid subtypes and to systematically identify potential candidates for multiple diagnosis or phenotypic expansion. abstract/case report text warts, hypogammaglobulinemia, recurrent infections and myelokathexis syndrome is a rare combined immunodeficiency due to autosomal dominant gain-of-function mutations of cxcr chemokine receptor. the late diagnosis of whim syndrome in two ukrainian adolescents highlights the diagnostic challenges in this disease. patient , year-old girl, had recurrent pneumonia since the first year of age; overall she had episodes of pneumonia. she has suffered from chronic bronchitis for last several years. she had recurrent otitis media and chronic pyelonephritis. neutropenia was revealed when she was year old. during episodes of bacterial infections she occasionally had normal value of neutrophils. the girl does not receive any treatment. patient , year-old boy, had three episodes of pneumonia when he was , and year old. others symptoms include recurrent herpetic infection, warts on the hands. since years of age he has haven persistent low neutrophil counts. the child was followed by hematologist and since years of age he has received g-csf ( mg/kg) twice a month. both children have leukopenia - cells/mm , neutropenia - - cells/mm , lymphopenia - cells/mm , low number of bcells - - cells/mm . hypogammaglobulinemia was not prominent in both children, they have slightly decreased level of igg ( , g/l), normal level of igm ( , - , g/l), patient has low level of iga , g/l. patient does not have protective level of antibodies to diphtheria and tetanus anatoxin, and anti-hbs antibodies were absent despite complete immunization. bone marrow aspirate revealed hypercellular marrow with granulocytic hyperplasia which was characterized by hypersegmented nuclei and cytoplasmic vacuolization of neutrophils. on molecular analysis of cxcr , heterozygous mutation c. c>t (p.arg *), known as r x mutation, was detected in both patients, confirming the diagnosis of whim syndrome. replacement therapy with intravenous immunoglobulin was started in both children together with antibacterial prophylaxis and g-scf. vaccination with -valent vaccine against hpv infection was recommended for both patients. whim syndrome is very rare immunodeficiency but may be underdiagnosed. the awareness about rare forms primary immunodeficiency is very important in clinical practice for early diagnosis and treatment. methods: clinical providers recruited from nicer institutions electively completed web-based survey questions related to provider characteristics as well as initial diagnostic evaluation of itp, aiha, ain and es via securequestionpro® software. likert scales ranging from ("rarely" < %), ("sometimes" to %), ("half the time" % to %), ("frequently" to %), and ("almost always" to %) were used to ascertain frequency of evaluation for each diagnostic study. statistical analysis and plotting was done using rv . . . plots were created using packages ggplot , v . . and ggiraphextra v . . . mean likert scale scores were calculated for each study for each suspected disease and plotted on radar charts. results: the survey was completed by providers, including hematology/oncology ( . %), rheumatology ( . %), allergy/ immunology ( . %) and other sub-specialties ( . %). a slight majority of physicians ( %) were fellows or within years of graduation; physician extenders and clinical pharmacists were also respondents. the majority ( . %) of respondents indicated that ≤ new immune-mediated cytopenia patients were seen at their institution annually. the vast majority of respondents ( . %) reported evaluating ≤ new es patients per year at their institution with % evaluating ≤ cases annually. collated data from all respondents showed that in all disease states, the primary evaluation was focused on peripheral destruction mechanisms; the majority of patients are only "sometimes" or "rarely" evaluated for bone marrow failure syndromes, connective tissue disease, immunodeficiency and non-malignant lymphoproliferative disorders, but when done were more likely in es ( figure ). evaluations were biased by sub-specialty with higher degrees of connective tissue focus by rheumatology and immunodeficiencies by allergy/immunology (table ) . genetic sequencing was "frequently" or "almost always" sent in . % of itp, . % of aiha, . % of ain and . % of es p a t i e n t s . p e r s o n a l o r f a m i l y h i s t o r y o f a u t o i m m u n e / hyperinflammatory disease, malignancy or cytopenias most strongly influenced the decision to send genetic testing. lack of insurance coverage/negative financial impact on the patient and concerns about the inability to resolve variants of uncertain significance were the biggest barriers for obtaining genetic testing. conclusions: current practices in the evaluation of immunecytopenias are heterogeneous by sub-specialty and globally limited in scope with few patients being evaluated for underlying etiologies. in particular, despite a known high frequency of pathogenic variants in es, less than a third of patients are undergoing sequencing, highlighting a need to reduce barriers to genetic testing. development of a consensus guideline with multi-disciplinary engagement to harmonize an optimal evaluation for patients with immune-mediated cytopenias is needed. interferon regulatory factor- (irf ) binding protein- (irf bp ) was originally identified as a transcriptional co-repressor of irf ( ). mutated irf bp was identified in a -member family with recurrent sinopulmonary infections, progressive hypogammaglobulinemia, and poor response to protein vaccines ( ) . we have now identified additional families ( subjects) with irf bp mutations. clinical histories show an expanded phenotype with / having chronic gastrointestinal disease; with gastrointestinal manifestations as the initial clinical complaint. five had granulomata in liver(x ), spleen, lung(x ) and gastrointestinal tract. five out of six tested had poor pneumococcal vaccine responses and four patients reported viral infections including varicella zoster(x ), influenza a and sapovirus. irf bp is a amino acid protein containing a highly conserved cterminal protein-protein interaction ring domain (rd). constraint metrics from gnomad indicate mild tolerance to missense changes and intolerance to loss-of-function alleles. we identified categories of mutations: rd mutation or deletion (n= patients), null alleles (n= ) and non-rd missense changes (n= ). functional studies assessing the ability to affect nfatdriven luciferase expression were performed. rd mutations ( / ) had more profound loss-of-repression than wild-type, while missense changes had lesser, but still measurable effects. further, mutation categories and functional studies correlated with clinical phenotypes. of patients with rd mutations, / had infections as presenting symptoms, / tested had hypogammaglobulinemia and / were diagnosed with cvid. one patient with a missense rd mutation had only an infectious phenotype (pulmonary mycobacterium avium) with slight decrease in immunoglobulins; in functional studies this mutation had the least effect of the rd mutations. haploinsufficient patients reported respiratory infections ( / ), recurrent urinary tract infections ( / ), gastrointestinal disease ( / ) and hypogammaglobulinemia ( / ). in contrast, / patients with non-rd missense changes presented with gastrointestinal complaints while only patients had infections (recurrent bronchitis, shingles). gi disease prevalence is consistent with high levels of irf bp expression in the colonic crypt cells (human protein atlas). to confirm this, immunohistochemical staining of colon biopsies from two patients was performed, identifying epithelial and glandular cells of the colon. irf bp is involved in multiple processes, including the negative regulation of nfat signaling( ), tcr signaling( ), inflammatory macrophages ( ) , and pd-l transcription ( ) . interaction with the glucocorticoid receptor affecting anti-inflammatory and metabolic transcription ( ) has also been reported. these observations highlight the irf bp response to type-i interferons (irf ) and tcr stimulation (nfat), regulation of inflammatory macrophages and co-regulation of glucocorticoid receptor mediated signaling. the expanding role of irf bp in multiple biologic systems correlates with the broad clinical presentation we observed in our patients. further studies utilizing irf bp mutation knock-in mice will help characterize the gastrointestinal, lung and immune pathology seen in our cohort. abstract/case report text common variable immunodeficiency (cvid) is a disorder of antibody deficiency arising from over genetic lesions. the clinical presentation of patients with cvid varies from recurrent, severe infections to autoimmunity. the immune dysregulation in cvid is especially difficult to treat and the lifespans of patients suffering from autoimmunity are much shorter than those without such complications. unfortunately, we have no way to identify which patients fall into which categories, or even know how many sub-categories of cvid there are. therefore, the field requires a method to classify patients into categories to precisely recognize and aggressively treat the more severe phenotypes. we address this goal by integrating analyses of patient exomes with analyses of cellular signaling. by analyzing stimulation assays with phospho-protein mass cytometry and high-dimensional data analytics, we aimed to elucidate signaling and phenotyping deficiencies in patients with cvid. importantly, our panel identifies all circulating immune cell subsets in whole blood. in eosinophils, we found amplified responses of pp , pstat , and cleaved caspase- in response to tlr / stimulation. we found additional amplified responses of pstat and pstat in cd lo monocytes. this finding suggests a previously unidentified role for eosinophils and cd lo monocytes to contribute to the pathophysiology of cvid. we found abormal numbers of memory b cell counts, total switched b cell counts, and igm+, cd + b cell (plasmablasts) counts between cvid patients and healthy controls. cd expression on b cells was significantly reduced in cvid patients as well. these b cell results mirror findings from prior, seminal studies on cvid. notably, we have found higher pd- expression in the effector cd t cells of patients. integrating phenotype data, genetic analysis, and mass cytometry data will provide a deeper understanding of each patient's phenotype and how the are clustered. we also expect that a better understanding of alterations in the exomes and functions of the circulating immune cells of cvid patients will lead to new therapeutic approaches. abstract/case report text objectives: primary atopic disorders are monogenic disorders leading to profoundly dysregulated allergic responses. studying patients with these disorders has been instrumental in expanding our understanding of the pathogenesis of allergic inflammation with therapeutic implications for common polygenic versions of allergic disease. clinical findings: we have identified a now -year old boy who presented with severe eczema, extremely high blood eosinophil counts ( . x cells/l, normal range: - . x cells/l) after birth and very high serum ige levels ( υg/l, normal range: - ug/l) since birth. known allergic disorders and parasitic infections are ruled out. given the extreme phenotype, whole exome sequencing was performed on the trio of patient and parents, and the patient was found to have a homozygous mutation in the evolutionarily conserved fibronectin iii domain of the osmr gene (c. t>a, p.v d) (figure ). osmr encodes oncostatin m receptor-beta, a component of both the osm type ii receptor and the il receptor, and is important for keratinocyte cell proliferation, differentiation, apoptosis and inflammation. mutations in osmr have been reported in association with familial primary localized cutaneous amyloidosis, however this condition was ruled out in this patient through skin biopsy which showed no amyloid deposits. methods and results: we modelled the c. t>a osmr mutation in hek cells and observed a loss of expression of the osmr receptor on the cell surface (with normal intracellular protein levels). this observation was mirrored in primary fibroblasts obtained from the patient. signal transduction through phosphorylation of stat and stat and gene expression (il and ccl measured via qpcr) was absent after stimulation with osm in patient fibroblasts. these signaling defects were rescued using a lenti-viral transduction approach to introduce the wild-type (wt) osmr gene. whole transcriptome analysis using rna sequencing confirmed that osm mediated jak-stat signalling pathways were deficient in the patient fibroblasts and were rescued after lenti-viral transduction of wt osmr. rna sequencing analysis also suggested significantly enhanced expression of genes in the nf-κb signalling pathway (e.g.: il and cxcl ) and decreased expression of genes in the tgf-β signalling pathway (e.g.: smad and smad ) in patient fibroblasts at baseline. this was also rescued upon lentiviral transduction. conclusion and future directions: our findings shed light into the disease mechanism of a novel primary atopic disorder, caused by a homozygous missense mutation in osmr. abstract/case report text -year-old caucasian female presented to immunology clinic with hypereosinophilia, eosinophilic esophagitis, peptic ulcer disease, severe gi bleeds, and chronic hepatitis. healthy throughout childhood, with minimal infectious history. in adolescence developed chronic severe myalgias and nsaid overuse, to which the peptic ulcer disease and bleeding were attributed. parents healthy and non-consanguineous. son with severe bleeding episodes and small stature. on exam she weighed lb, bmi . sclerae anicteric. tongue deeply furrowed. cervical nodes palpable. heart and lung exam normal. no hepatosplenomegaly. no clubbing of the digits or edema. skin was clear. wbc , /ul, eosinophils /ul, hemoglobin g/dl, normal platelet count. however, platelet aggregation testing abnormal. bone marrow normocellular, and flow cytometric and molecular analysis did not show hematolymphoid malignancy, primary hypereosinophilic syndrome, or systemic mastocytosis. lymph node biopsies did not show lymphoma or aberrant t cell populations. noted to have chronically elevated creatine phosphokinase, ranging from - u/l over two years at our institution. deltoid muscle biopsy showed non-specific myelopathic changes. an adult dystrophy immunostaining panel was normal. ultrastructure examination showed no abnormal storage material. a genetic panel for metabolic myopathies failed to reveal a cause. total igg, iga and igm normal. ige elevated at ku/l, and igg subclasses showed igg elevated at mg/dl. flow cytometry showed normal t, b and natural killer cell numbers. normal proportions of naïve, mature and activated t cells. vaccine response assessment was normal. evaluation for autoimmune/rheumatologic diseases was negative. liver biopsy demonstrated findings consistent with primary or secondary sclerosing cholangitis (without increased igg staining). given her inflammatory phenotype, additional genetic analysis was sent, assessing for primary immunologic disorders. this identified heterozygous variants of uncertain significance in ctla (c. t>a; ps t), zap (c. c>g; p.d e), and stim (c. t>c; p.l s). analysis of the ctla variant in vitro revealed that it was expressed normally. foxp expressing regulatory t cells were present in normal proportions in vivo and appeared phenotypically normal. this variant was found in her unaffected father. the zap variant is present in population databases (rs , exac . %), and was felt unlikely to be clinically relevant. the stim l s variant, although not shown previously in human patients, has been previously shown in vitro to be a gain of function mutation [ ] [ ] [ ] . furthermore, familial analysis revealed that this was a de novo mutation arising in the patient, and present in her son. humans with other gain of function mutations in stim and the orai channel it activates have overlapping syndromes including storkmorken syndrome, tubular aggregate myopathy and york platelet syndrome, characterized by chronic myopathy and platelet aggregation defects [ ] . the stim l s mutation is predicted to cause constitutive stim activation and calcium influx and likely provides an explanation for the patient's chronic myopathy and abnormal platelet aggregation. neither eosinophilic disease, nor cholangitis, have been described previously in stim gain of function-related diseases. it is unclear whether these issues are related to this novel stim mutation, or to other genetic or environmental influences. treatment of diseases caused by overactive crac channels is challenging as no pharmacologic inhibitors are yet clinically available. nomid/cinca syndrome is one of the periodic syndromes associated with cryopyridines. it is a defect in the innate immune system causing excessive activation of the inflammasome, with consequent il- secretion and neutrophil recruitment. clinically, damage occurs to organs such as the skin (neutrophilic urticaria), central nervous system (meningitis and deafness) and joint (arthritis). levy et al. ( ) evaluated a large series of patients and median onset age was . years, while the median age at diagnosis was years, although the symptoms initiate in the first days of life. treatment includes corticosteroids, which act by nonspecifically blocking all inflammatory cytokines, or by blocking il- specifically. if early diagnosis and treatment of the disease is not made, natural evolution leads to motor and adaptive disability and death in % of cases already in adolescence due to infection, neurological complications or secondary amyloidosis. we report a -month-old male child from nonconsanguineous parents who presented shortly after birth, multiple scaling and erythematous lesions throughout the body, evolving with following symptoms: abdominal abscess, hepatitis, meningitis and pioarthritis. laboratory tests showed elevation of inflammatory tests (esr, crp, amyloid protein a) and leukocytosis. the diagnosis was suspected at the nursery where the patient remained hospitalized for days. a personalized multigene panel was requested. it was identified the variant p.gly val, heterozygous for nlrp gene, not described in the literature, confirming the diagnosis of cinca/nomid syndrome. after discharge, it was introduced prednisolone ( , mg/kg/day) and antiinterleukin- (il- ). after the second dose, skin lesions and joint edema regressed, weight gain, and neuropsychomotor development improved. this case reports a very early diagnosis of nomid/cinca syndrome. it warns neonatologists and pediatricians about the need of precocious recognition of the syndrome, probably improving the prognosis of the patient. professor/university center health abc abstract/case report text background: leprosy affects more than , people worldwide. brazil represents the rd. country in the world in leprosy frequency and maranhão state is an hyperendemic region. the city of imperatriz (ma) stands out as a reference center in the care of these patients. according to few reports, lectin pathway of complement system may play a role in susceptibility to leprosy. mannose binding lectin (mbl) and ficolins (fcns) recognize patterns of sugars and acetylated residues (pamp), respectively, in a wide variety of pathogens, including m. leprae. high levels of ficolins and mbl may act unfavorably promoting the spread of m. leprae. the present study evaluated the role of ficolin and mbl in m.leprae patients and contacts. methods: a cross-sectional case-control analytical study was carried out, evaluating clinical and epidemiological data and serum levels of mbl and fcn (elisa) from july to april . the study was approved by ethics committee and informed consent forms were signed before sample collection. data analysis was performed using the spss . for windows statistics program. results: we evaluated serum samples ( patients and healthy family contacts), . % were female, % under years old, % african-brazilian, % of the families had more than contacts at home. clinical data showed multibacillary forms in . %; dimorphic ( %) and virchowian clinical forms ( . %), up to affected nerves in ( . %) and more than lesions in ( . %). it was observed that ( . %) had a reaction, being type ( %) more predominant. disability grade was found in patients ( . %). in children under years, . % were multibacillary, . % dimorphic and % undetermined; ( . %) also had reactions, % type reaction and degree of disability in . % of children with the disease. the evaluation of serum fcn and mbl levels for the patients (n = ) and contacts (n = ) were . ng/ml and . ng/ml, (p = . ), and . ng/ml (p) and . ng/ml (c) (p = . ), respectively. there were lower values of fcn in patients with type reaction (sudden and intense inflammatory processes) versus no reaction ( . ng/ml vs . ng/ml) (p = . ) and in patients with disability grade (severe sequelae) versus disability grade ( . ng/ml vs . ng/ml) (p = . ). higher fcn values was observed in patients with no disability ( . ng/ml) (p = ). mbl concentrations were higher for patients above years in comparison with patients below that age ( . ng/ml vs . ng/ml)(p = . )) and correlated with the occurrence of a multibacillary clinical form. conclusions: mbl and fcn levels were not different in the patients and contacts of m. leprae, nevertheless the presence of severe forms with sequelae (reaction type and disability grade ) were associated with lower levels of fcn . in addition, it is possible that lower mbl levels could influence the higher frequency of multibacillary disease below years old. abstract/case report text introduction: hyper ige syndrome (hies) is a primary immunodeficiency characterized by elevated ige levels. symptoms can range from severe eczema, recurrent skin infections or pneumonias, and typical dysmorphic facies. there have been wide non-immunologic presentations in patients with hies, including retained primary teeth, scoliosis, craniosynostosis, arterial aneurysms and joint hyperextensibility. an association between hies and autoimmune hemolytic anemia (aiha) has further been described in the literature. however, there have been no reported cases of hies in association with iron deficiency anemia and concurrent pica. we present a unique case of a patient with a history of eczema, recurrent skin infections and pica found to have hies and iron deficiency anemia. case presentation: a -year-old boy with a history of allergic rhinitis presented to the allergy & immunology clinic for evaluation of chronic eczema and recurrent skin infections. the patient had a history of multiple hospitalizations requiring intravenous antibiotics for cellulitis and superinfected eczema since he was an infant. symptoms were refractory to the use of multiple skin barrier ointments and oral antihistamines. his mother further noted that for the past two months prior to initial evaluation, he developed a fixation with eating crayons, baby powder and chewing on drywall. physical exam was notable for a dysmorphic face, broad based nose, pale nasal mucosa with ample clear discharge, high-arched palate and lower incisor supernumerary teeth. his skin was characterized by generalized dryness, lichenification and scaly desquamation with boils on extensor surfaces of knees and elbows. initial screening for hies via t-helper functional assay was consistent with decreased expression of il- . genetic testing revealed stat s g missense pathogenic variant consistent with hies. cbc was also notable for decreased hemoglobin at . g/l and mcv of fl. patient was diagnosed with concurrent hies and pica in the setting of iron deficiency anemia. iron supplementation was started and patient's pica improved. discussion and conclusion: our patient with hies had a peculiar initial presentation with the classic signs and symptoms of hies and pica. the diagnosis of hies can often be delayed due to the wide range of clinical presentations. to our knowledge, the association of hies with iron deficiency anemia and pica has been underreported in literature. screening for anemia should be considered when evaluating patients with hies in order to rule out comorbid iron deficiency anemia which can be easily treated with iron supplementation. abstract/case report text introduction: common variable immunodeficiency is a primary immunodeficiency with variable and diverse phenotypic presentations. the two main phenotypes include a group which primarily exhibits recurrent infections and a group with or without infections and primarily inflammatory and autoimmune complications. the latter, may lead to a delay in diagnosis and is associated with poorer outcomes and higher morbidity and mortality. ( ) another group of patients present with t-cell defects, lung disease, autoimmunity, and infections and may be diagnosed as having cvid but instead can have mutations in lrba or pi kinase. this subset of patients has been referred to as "cvid-like" in the literature. ( ) case presentation: patient is an year old female who initially presented to an outside facility due to days of fatigue, fever, and abdominal pain. upon presentation, she was found to have massive splenomegaly, hepatomegaly, and an abnormal chest x-ray showing mediastinal lymphadenopathy and pleural effusion. laboratory results demonstrated pancytopenia, hypogammaglobulinemia, and low b cells, t cells, and nk cells via flow cytometry. she was transferred to our institution for further work up. she did not have any prior history of recurrent infections, asthma/lung disease, or autoimmune conditions. initial ct of the chest was consistent with granulomatous lymphocytic interstitial lung disease. patient was diagnosed with common variable immunodeficiency with granulomatous lymphocytic interstitial lung disease and was treated initially with high dose ivig, corticosteroid taper, rituximab, and imuran. she had interval worsening of pft and lung disease as shown by ct scan. genetic panel for cvid and related conditions revealed variants of unknown significance. one heterozygous mutation in blnk gene (c. g>a) and one heterozygous mutation in lrba gene (c. g>a). she was started on infliximab with plans to repeat ct scan in months. discussion: mutations in both blnk and lrba have been associated with primary immunodeficiency. mutations in blnk, which is located on chromosome , have been associated with autosomal recessive agammaglobulinemia. homozygous or compound heterozygous mutations in lrba on chromosome , can lead to lrba deficiency which encompasses a wide range of clinical presentations including hypogammaglobulinemia, autoimmune disease, inflammatory bowel disease, antibody deficiency, organomegaly, and recurrent infections. ( ) without genetic testing, the clinical presentation can be difficult to distinguish from common variable immunodeficiency. the patient presented has clinical features that can be seen with mutations in both blnk and lrba, however she is heterozygous for both mutations. further analysis, including measurement of lrba protein expression, is needed to further define her underlying immunodeficiency so appropriate treatment can be administered. abstract/case report text a month-old, previously healthy, unvaccinated male presented with one week of diarrhea and cough and was admitted for dehydration and hypoxemia. his mother and sister both had a history of incontinenti pigmenti (ip). on physical exam, he was alert, afebrile, with tachypnea and subcostal retractions. enterovirus/rhinovirus and parainfluenza were detected, but he became progressively hypoxemic and eventually required intubation and high-frequency oscillatory ventilation. chest x-ray showed multifocal bilateral airspace opacities. empiric treatment for pjp with trimethoprim/ sulfamethoxazole and glucocorticoids was started. tracheal aspirate pcr confirmed p. jiroveci. hiv rna pcr was negative. ivig was started due to suspicion for primary immunodeficiency. although his respiratory status gradually improved, he subsequently developed multiple skin lesions. skin biopsy grew mycobacterium szulgai. m. szulgai osteomyelitis of the right fibula and the left nasal bone was also detected, indicating hematogenous spread of the infection. he was started on four-drug anti-mycobacterial therapy and interferon-gamma (actimmune) at doses ranging from μg/m^ three times weekly to μg/m^ qod. immune work-up revealed t-cell lymphopenia [cd +/cd + /μl ( - , /μl) and cd +/cd + /μl ( - , /μl)] with an abnormally increased proportion of memory cd t-cells compared to naïve cells for age. b-cell numbers were normal, and nk cells were decreased [cd +cd +/cd - /μl ( - /μl)]. nk cell lytic function by k lysis was normal, whereas cd a degranulation was decreased. the serum igm level was normal [ mg/dl ( - mg/dl) whereas iga [ mg/dl ( - mg/dl)] and igg [ mg/dl ( - mg/dl)] were elevated. mononuclear cell cytokine response to ligands for tlr -tlr , tlr -tlr , tlr , tlr , and tlr -tlr was normal. dna sequencing revealed a novel nonsense mutation in exon of the ikbkg (p.gln ter (q x) (cag>tag): c. c>t, confirming the diagnosis of nemo deficiency, which was suspected based on the infectious disease presentation and the maternal history of ip. the diagnosis was further supported by signs of ectodermal dysplasia of teeth that appeared starting at months of age. he underwent hsct using bone marrow from a / matched unrelated donor after conditioning with atg, busulfan, fludarabine and rituximab. actimmune therapy was continued until days prior to transplant. for gvhd prophylaxis, he received tacrolimus and low-dose methotrexate. he achieved full donor chimerism post-transplant and has had no significant gvhd. interesting features of this case include the prominence of ip in mother and sister, which is usually due to female heterozygosity for an ikbkg null allele. such null alleles when inherited by the male fetus are embryonic lethal. our patient's nonsense mutation would be expected to result in severely impaired ikbkg protein expression and function. however, the fact that he had was born at term and initially was healthy coupled with his preservation of normal tlr function suggests that his ikbkg allele is likely to be a hypomorphic mutation. studies are in progress using ebv-transformed b-cell lines from the patient to evaluate ikbkg expression and function. also of interest, our patient was able to tolerate relatively high doses of interferon-gamma therapy without inflammatory side effects or an adverse impact on engraftment or gvhd. abstract/case report text background: primary atopic disorders are caused by genetic mutations that skew the immune system towards severe allergic disease. germline gain-of-function (gof) mutations in jak are a newly described monogenic cause of severe atopy, with affected patients demonstrating profound eosinophilia and allergic inflammation. our initial report of this novel condition identified a dramatic clinical response to the combined jak / inhibitor ruxolitinib. we aimed to determine the long-term clinical response to ruxolitinib in patients carrying a germline jak gof mutation, and to characterize the effect of enhanced jak signaling on t lymphocyte effector functions and hematopoiesis. methods: clinical outcomes were evaluated in two pediatric patients carrying the c. c>a (p.a d) gof mutation in jak after . years of ruxolitinib treatment. t cell phenotyping was performed using extracellular surface marker and intracellular cytokine staining by flow cytometry, and by gene expression signature profiling of rna sequencing data. to evaluate the effect of enhanced jak activity on myelopoeisis, we reprogrammed jak gof patient-derived peripheral blood mononuclear cells into induced pluripotent stem cells (ipsc) and performed directed myeloid differentiation. rna sequencing was performed on rna collected during ipsc myeloid differentiation and from whole blood of affected patients before and after ruxolitinib treatment. results: long-term use of ruxolitinib was associated with improved growth, reduced eosinophilia, and control of allergic inflammation without significant infectious complications, however, anemia represented a dose-limiting adverse effect. t cell immunophenotypic analysis revealed severe t helper (th) cell skewing towards a th phenotype preruxolitinib treatment, in keeping with the allergic clinical manifestations. analysis of myeloid differentiation revealed an increased myeloid to erythroid ratio in colonies derived from jak gof ipscs compared to controls. rna sequencing analysis of jak gof human whole blood and ipscs compared to controls revealed upregulation of cytokine and cytokine receptor genes implicated in allergic inflammation and early eosinophil precursor commitment, including csf- and the interleukin- receptor. reactome pathway analysis of genes upregulated in both jak gof ipsc and whole blood compared to controls showed enrichment of several pathways including interferon alpha/beta, interleukin- /- and interleukin- signaling. conclusions: this work demonstrates a critical role for jak in atopic immune dysregulation, specifically driving a th phenotype and eosinophilia. combined jak / inhibition can reverse much of the allergic inflammation, with dramatic clinical effects. this has important implications for our understanding of the pathogenesis and potential therapeutic targets for early life allergic immune dysregulation. had severe combined immunodeficiency (scid) and/or severe disease in association with their combined immunodeficiency (cid) necessitating haematopoietic stem cell transplantation (hsct). we present clinical and laboratory features of new zealand patients from the same family with a novel heterozygous missense variant in rac [c. t>g, p.ile ser (i s)]. the index patient (p -age y, m) has a history of infectious gastroenteritis, staphylococcal aureus conjunctivitis, recurrent otitis media and recurrent herpes simplex virus (hsv)- cutaneous infections. his siblings (p age y, m; p -age y, f) and his mother (p age y, f) all have a history of recurrent viral (hsv- ) and bacterial (staphylococcal aureus, streptococcal pyogenes) cutaneous infections and/or recurrent sinopulmonary infections that respond to empiric antimicrobial therapy. their neutrophils all had enhanced superoxide production in response to stimulation by fmlp and pma as compared to healthy controls'. these findings suggest that rac i s is an activating mutation causing notable abnormalities in neutrophil morphology and nadph oxidase activation similar to other recently reported mutations. this novel mutation expands the phenotypic spectrum of rac activating mutations. clinical management of affected patients needs to be tailored to their phenotype and disease severity. background and aims: heterozygous mutations in cytotoxic tlymphocyte antigen- (ctla ) are associated with recurrent infections, lymphoproliferation, autoimmunity and lymphocytic infiltration of target organs. disease penetrance can be highly variable even among related family members carrying the same ctla mutation. our evaluation of a subset of the ctla patient cohort followed at the national institutes of health (nih) revealed that % of ctla mutation carriers have gastrointestinal (gi) manifestations which include diarrhea and diffuse lymphocytic enteropathy. our aim was to determine whether the intestinal microbiome, metagenome and metabolome could distinguish patients with ctla haploinsufficiency (ctla -h) based on disease severity, and the presence or absence of gi manifestations. methods: clinical metadata and fecal samples were collected from healthy individuals (n= ) and patients with ctla -h (n= ). patients with ctla -h were classified as having minimal (n= , only endocrine and/or dermatological manifestations) or systemic disease (n= , hematological and multi-organ involvement). they were further classified based on whether they had a history of enteropathy (n= ) or active gi disease ( < bowel movements per day and/or blood or mucus in stool) at time of sampling (n= ). metabolomic profiling (using a panel of metabolites) and s rrna gene sequencing (v region) was performed on fecal samples (total samples: ; number of reads/sample: , to , ; median: , ). a subset of samples were subjected to shotgun metagenomic sequencing based on findings from the s rrna gene sequencing analysis. results: all patients with ctla -h and a history of enteropathy or active gi disease also had systemic disease. fecal samples from patients with a history of enteropathy had a distinct microbial community structure (fig. ) which was significantly less diverse (fig. ) compared to healthy individuals and patients with minimal vs. systemic ctla -h. patients with a history of enteropathy had significantly higher relative abundance of bacterial taxa including shigella-escherichia (fig. ) . shotgun metagenomic sequencing confirmed that samples from patients with a history of enteropathy were dominated by subsets of identified escherichia coli strains, all of which share genes coding for specific types of virulence factors such as curli fibers (facilitate uptake into host cells), flagellar proteins (increase motility) and enterobactins (increase bacterial iron transport). meanwhile, samples from patients without active gi disease at the time of collection were enriched for several taxa including bacteroides nordii and akkermansia muciniphila compared to patients with ctla -h and active gi disease (fig. ) . metabolomic analyses showed that asparagine, -hydroxybutyrate, cytosine and cystine were enriched in samples with abundant e. coli, whereas samples without e. coli were enriched in metabolites involved in pyrimidine (holm p= . ), purine (holm p= . ), and alanine/aspartate/glutamate metabolism (holm p= . ) (fig. ) . conclusions: fecal samples from ctla -h patients with a history of enteropathy were heavily colonized with e. coli strains that are associated with a specific metabolomic profile and that share virulence factor genes that may facilitate host invasion. these data suggest that the microbiome and metabolome can distinguish patients with ctla -h and gi disease, and support the potential use of antibiotics or even antimetabolites to treat ctla -hrelated enteropathy. the dna polymerase delta (pol δ) complex is essential for leading and lagging dna strand synthesis. its catalytic subunit (pold ), carries both polymerase and exonuclease activities and plays a crucial role in dna replication and repair. heterozygous pold mutations have been associated with inherited colorectal cancer and mandibular hypoplasia, deafness, progeroid features and lipodystrophy (mdpl) syndrome. more recently a biallelic loss of function mutation in pold (p.r c) that impairs the stability of the pol δ complex, has been reported in related subjects with recurrent infections, deafness and combined immunodeficiency (cid) with t-cell lymphopenia, cd + t cell oligoclonality but preserved b cell proliferation. we report here a second family in which a novel biallelic missense mutation in pold gene was associated with cid. the proband is a -year-old boy born to consanguineous pakistani parents. since infancy he suffered from failure to thrive and recurrent infections, including episodes of pneumonias, multiple otitis media, sinusitis, recurrent cellulitis at the g tube site, bk viruria and shingles. live and dead vaccines were well tolerated. at years of age sensorineural hearing loss together with profound leukopenia (anc cell/μl, alc cells/μl) and hypogammaglobulinemia ( mg/ dl) were identified. intermittent ivig replacement and antimicrobial prophylaxis were initiated. immunophenotyping at years of age showed severe t cell lymphopenia ( cd + cells/μl, cd + cells/μl, cd + cells/μl, figure . volcano plot of metabolites present in fecal samples enriched with e. coli vs. samples without e. coli. c e l l s / μ l , c d + c d h i f o x p + c e l l s / μ l ) , a n d hypogammaglobulinemia (igm mg/dl, igg mg/dl, iga < ). physical exam was remarkable for multiple acquired nevi in the groin area, teeth abnormalities and global developmental delay. whole exome sequencing analysis revealed a homozygous pold missense variant (nm_ c. c>g, p.q e) absent in public databases (cadd score of ). parents were heterozygous. tcr-vβ family expression was normal in both cd + and cd + t cells, but the proportion of t cells expressing vα . (encoded by the distal trav - gene) was markedly reduced (less than %), consistent with impaired vdj recombination at the tra locus and/or with defective thymocyte survival. constitutive expression of γh ax was observed in t and nk cells after h and h of culture in unirradiated conditions. at h post-irradiation ( gy), reduced levels of p-atm were detected in t and nk cells, and lack of atm, smc and h ax phosphorylation was observed in a subset of b cells, suggesting inability of these cells to mount an effective dna repair response. bone marrow examination showed normal trilineage hematopoiesis but decreased proportion of cd -cd + mature b cells and increased proportion of pre-b cells. conclusion: we report the second mutation associated with autosomal recessive pold deficiency. our findings broaden the understanding of the mechanisms underlying the immune defect in this disease to include b cell maturation arrest in the bone marrow and a dna repair defect that may support the generation of a restricted tcr repertoire in the thymus and increased malignancy risk. abstract/case report text following allogeneic hematopoietic cell transplantation (hct) for scid, the development of a diverse t cell repertoire is essential for optimal immune recovery. high-throughput sequencing (hts) of the trb repertoire is the best tool for the evaluation of clonotype dynamics during immune reconstitution as compared to cdr spectratyping and staining of vβ families. we investigated whether longitudinal hts analysis of trb would accurately assess development of tcr repertoire diversity over time and reflect the quality of t cell reconstitution following hct for scid. we wanted to study the effect of conditioning regimen, scid genotype, donor type on tcr diversity post hct. we hypothesized that repertoire diversity may represent an early biomarker to predict long-term immune reconstitution vs. need for a second intervention. we assessed if the trb repertoire post-hct carried a molecular signature of selfreactivity. methods: the composition and diversity of trb repertoire of scid infants, pre-hct and at d, and mo and yearly posttreatment(s) was studied by hts. median time of follow-up was mo. subjects were part of a prospective study of scid by the primary immunodeficiency treatment consortium. equal amounts of total rna extracted from peripheral blood was used as template to semi-quantitatively amplify trb rearrangements. the vdj statistics file (past program) was used to calculate a shannon entropy (h) index of repertoire diversity and simpson ( -d) index of repertoire clonality. results: trb sequence analysis of scid patients showed poor diversity at baseline, followed by improvement to normal complexity (h index > . ) after hct. similar kinetics of development of trb diversity were seen in patients with il rg, jak , and il r defects (n= ) as in those with rag and artemis defects (n= ). in the latter group, however, hct with no conditioning or immune suppression only was associated with persistently lower diversity than hct with conditioning (p < . ), a difference not found in the il rg/jak /il r group (fig. ) . hct from a matched donor ( / conditioned) correlated with higher diversity than hct from a mismatched donor ( / conditioned) (p= . ). having > cd + t cells/ul at mo post-hct correlated with higher trb diversity at and mo post-hct (p < . ). the trb repertoire d post-hct was enriched for the presence of central cysteines at the apex of the cdr (p < . ), a biomarker of self-reactivity ( fig. ). an h-index of . or lower at d after hct predicted need for second intervention (hct or gt) (fig. ) . conclusions: analysis of trb diversity allows for detailed assessment of development of a diverse t cell repertoire following cellular therapies for scid and confirms the need for patienttailored treatment strategies based on scid genotype. t-cell repertoire d post-hct is characterized by a molecular signature that may contribute to the increased rate of autoimmunity early post-transplant. furthermore analysis of trb diversity at d post-hct may identify patients at risk for failure of sustained immune reconstitution, thus prompting a second intervention without delay. abstract/case report text background atopic dermatitis is a chronic, multifactorial, relapsing inflammatory skin condition which is one of the main known health problem worldwide. atopic dermatitis lesions are frequently colonized by staphylococcus aureus and staphylococcus epidermidis. their susceptibility to form biofilms, ability to form adhesive skin colonies which lead to extremely resistant to antibiotics and immune responses. formation of skin biofilm resulted in complex bacterial communities that have unique effects on human keratinocytes, mouse fibroblasts and host immunity. aims: the aims of this study to confirm the specificity of s. aureus or its secreted factors in induction of pro-inflammatory cytokines il- , tslp and toxicity on human keratinocytes and mouse fibroblast. the second aim to study the inhibitory effect of co-culture of s. epidermidis with s. aureus in term of production of pro-inflammatory cytokines and toxicity. method and materials: human epidermal keratinocytes and mouse embryonic fibroblasts cell lines from t were used as a control strain to examine production of inflammatory response (il- and tslp) and cell death induced by s. aureus in the presence and absence of s. epidermidis. tslp and il- were detected by elisa and the apoptosis of s. aureus and s. epidermidis on these cells was evaluated by flow cytometry. result: recent findings propose the important role of skin biofilms in the pathogenesis of atopic dermatitis. s. aureus have been found to induce secretion of pro-inflammatory cytokines and cause apoptosis of human keratinocytes and mouse fibroblasts. presence of s. epidermidis as skin biofilm found to protects the human keratinocytes and mouse fibroblasts from induction of proinflammatory cytokines and cytotoxicity. conclusions and future work: s. aureus are essential in production of inflammatory response and cell death of mouse fibroblasts and human keratinocytes. future work will be carried out to identify the soluble factors that responsible in induction of pro-inflammatory cytokines. in addition, more studies are needed to be able to understand the mechanism by how s. epidermidis reduce the induction and cytotoxicity caused by s. aureus. j clin immunol in adult patients, in whom arbitrarily defined diagnostic criteria for antibody deficiency syndromes are not fulfilled, is subject to interpretation and decision differences reported by immunologists world-wide. in this study, we explored whether training in one particular program would decrease the variability in diagnostic and treatment approaches seen in the responses to two nationwide questionnaires in the uk and the usa. methods: a -minute online survey originally administered to a cross-sectional sample of us allergists/immunologists (usa/i) in january, , was also answered by a/i subspecialists who had trained in the last years at the louisiana state university health science center allergy immunology training program in new orleans (laa/i). respondents were asked questions on patient assessment, antibiotic use, initial igrt, and immune response assessment in decisionmaking to prescribe igrt. usa/i participants were recruited from the dynata physician professional panel. laa/i participants were recruited by the louisiana primary immunodeficiency network (lapin). results: overall, laa/i had consensus responses to the various practice questions close to % of the time, but outliers were always present, as was also observed in the usa/i. there was a higher frequency in the reported care of patients as described in the questionnaire by laa/i. over % of laa/i assessed vaccine responses prior to commencing igg replacement vs only % of usa/i p < . . all la a/i used the pneumococcal vaccine for assessment purposes while few used tetanus and hemophilus influenza, and none used meningitis or salmonella vaccines. these vaccines were still used by some of usa/i. a high level of concordance was observed among all respondents in that only few regarded pneumococcal antibody testing as the definitive test to commence igrt. high resolution chest ct scan was used more often by laa/i before starting igrt. assessment of effectiveness of igrt was decided after only months by more usa/i, vs laa/i, who tended to wait months to decide to continue or discontinue igrt. conclusions: all a/i responders saw a significant number of patients who do not conform to strict diagnostic criteria for antibody deficiency syndromes. there is diversity in the approach of usa allergists/ immunologists in determining the indication for igrt for non-classical antibody deficient patients. laa/i responses made it obvious that post graduate influences always play a role in shaping the way a/i practice evolves after graduation. drawing on clinician experiences through questionnaires offers a valid contribution to developing consent approaches to improve patients' clinical conditions. diagnostic criteria and treatment guidelines would benefit from practice-based realistic recommendations based on a/i experience. abstract/case report text background: autoimmune lymphoproliferative syndrome (alps) is a rare genetic disorder secondary to a defective fas-mediated apoptotic pathway of mature lymphocytes. it is characterized by chronic nonmalignant lymphoproliferation in the form of lymphadenopathy and/or splenomegaly, autoimmune manifestations such as cytopenias, increased risk of lymphoma, and expansion of tcrαβ+ cd -/cd -(dnt)t-cells. germline or somatic pathogenic variants in fas, fasl, and casp are well described genetic defects associated with alps. the definitive diagnosis for alps, based on the revised nih diagnostic criteria, include both required criteria (chronic non-malignant, non-infectious lymphadenopathy, splenomegaly, or both and elevated tcrαβ+ dnt t-cells) and one of the primary accessory criteria (defective lymphocyte apoptosis or mutation in the genes mentioned above). patients who do not meet the current diagnostic criteria are considered for alps-related disorders. case presentation: we report a -year-old male who presented with recurrent infections, splenomegaly and chronic lymphadenopathy since month of age. due to its chronicity he was evaluated by multiple specialists for malignant and infectious causes. hematological workup including bone marrow biopsy was unremarkable except for an elevated ldh level. infectious workup identified a past cmv infection. clinical course is pertinent for chronic splenomegaly which was identified incidentally at . years of age during an evaluation for intussusception. family history is pertinent for a father with recurrent infections, paternal grandmother with thrombocytopenia of unknown cause requiring platelet transfusions, and paternal cousin with neutropenia. there is no family history of lymphomas. history of chronic lymphoproliferation and recurrent infections prompted an evaluation for lymphoproliferative disorder. full immune workup was notable for elevated plasma il- and il- , normal immunoglobulin levels, lymphocytes subsets, vitamin b level, soluble fasl, and relative frequency (%) but borderline increased absolute count of tcrαβ+ (dn) t-cells. in addition, he was noted to have presence of anti-platelet antibodies, poor lymphocyte proliferation to antigens, and low pneumococcal antibody titers. genetic testing with a pid gene panel identified a likely pathogenic heterozygous variant in prf c. del (p.his thrfs* ), a heterozygous variants of uncertain significance in casp c. c>t (p.pro leu) and stim c. a>g (p.thr ala). the casp variant is present in alleles in gnomad ( k total allele count) and reported deleterious by sift. discussion: unlike the typical alps presentation, characterized by dominantly lymphoproliferation and autoimmunity, our patient's clinical phenotype is striking for recurrent infections, abnormal t-cell function, and poor antibody response. our patient does not the meet diagnostic criteria for alps due to normal relative frequency of dn t-cells. however, presence of elevated of il- , il- , platelet autoantibodies raise concern for alps-related disorder. in addition, family history of recurrent infections and cytopenias raises concern for familial autoimmunity and alpslike phenotype. although casp is associated with autosomal dominant and autosomal recessive alps, the role of this vus is yet to be determined. conclusion: we continue to investigate the pathogenicity of our novel casp vus. further studies include pedigree analysis, fas apoptosis assay and apoptosis pathway testing to assess for the etiology of this alps-related disorder. (word count , max ) abstract/case report text rationale: ocrelizumab is a recombinant anti-cd monoclonal antibody, which binds to a different, but overlapping cd epitope than rituximab. there have been increasing reports evaluating hypogammaglobulinemia and morbidity and mortality in patients receiving rituximab, but there is a paucity of data on hypogammaglobulinemia in patients treated with ocrelizumab. methods: we performed a retrospective review of patients who received ocrelizumab in our healthcare system. we evaluated the demographics, indication for ocrelizumab, frequency of immunologic evaluation, and h y p o g a m m a g l o b u l i n e m i a p r e -a n d p o s t -o c r e l i z u m a b . hypogammaglobulinemia was stratified as mild (igg < mg/ dl or less than lab reference range), moderate (igg < mg/ dl) or severe (igg < mg/dl). results: we identified patients who received ocrelizumab for multiple sclerosis (average number of ocrelizumab cycles = ; range - cycles). there were ( %) female patients, with a mean age of years old (range - ; standard deviation ± t-cells. tnfα, ifnγ, and il- were not statistically different between cgd patients and healthy controls. tnfα, ifny, il- , and il- a expression in patients with cgd who had active colitis or history of colitis were increased as compared to cgd patients without a history of colitis but did not reach statistical significance. in two patients, il- a expression that was elevated pre-hct normalized post-hct. discussion: the mechanism for increased susceptibility to inflammatory disorders in patients with cgd has not been well elucidated. our results agree with previous studies demonstrating increased il- and il- a production from cd + t-cells in patients with cgd indicating a proinflammatory state in these patients at baseline. also, there appears to be an increase in tnfα, ifny, il- , and il- a expression from cd + t-cells that correlates with presence of inflammatory disease vs. those without inflammatory disease indicating that these cytokine perturbations may be able to serve as biomarkers of disease activity. a larger sample size with prospective collection will be analyzed in the future. abstract/case report text background: glucose- -phosphatase catalytic subunit (g pc ) deficiency, is characterized by severe congenital neutropenia, recurrent bacterial infections, mild intermittent thrombocytopenia and a high incidence of congenital cardiac and uro-genital defects. we report the case of a -yo male with chronic neutropenia and thrombocytopenia, who was found to have homozygous pathogenic variants in g pc (c. del, p.phe serfs* ). unique to this case is the patient's long history of misdiagnosis of evans syndrome (chronic autoimmune neutropenia with thrombocytopenia). case presentation: a -yo male with reported diagnosis of chronic autoimmune neutropenia and thrombocytopenia since age was referred to our a b clinic with concern for an underlying immune dysregulation syndrome. he had a history of oral ulcers, gingivitis, recurrent bacterial infections (otitis media, pneumonia, skin abscess) concomitant with severe neutropenia ( < cells/ul), for which he had received treatment with systemic steroids and g-csf since he was years old. he also had history of asthma and short stature, thought to be secondary to his chronic systemic steroid use. we present the first chilean patient with stat gof immunedysregulation . moreover, to our knowledge this is the first stat gof patient presenting with lymphomatoid granulomatosis. this is a severe pulmonary disease in which primary immunodeficiencies including stat gof should be considered in the differential. in this case rituximab successfully resolved pulmonary nodules and respiratory symptoms. there was persistence of mild hepatosplenomegaly but otherwise clinical stability and monitored expectantly until the age of . at that time he presented with fever, left knee and ankle arthritis. he underwent arthrocentesis of the left knee and left ankle, both aspirates were sterile, with notable leukocytosis with heavy neutrophilic predominance. an extensive rheumatologic and infectious workup was non-diagnostic. both sil- r and il- were elevated, , units/ml ( < ) and pg/ml ( < ), respectively. he was treated with systemic corticosteroids, ultimately arthritis resolved after months. at age he presented for the first time with periorbital pain and conjunctival injection of the left eye that persisted after minor trauma. he was found to have nongranulomatous uveitis, which responded ultimately to systemic corticosteroid. he then presented at age with fever and right knee and great toe arthritis. again he underwent arthrocentesis which revealed aseptic arthritis, and at that time was started on anakinra (anti il- β) and prednisone. there was clinical improvement over several weeks followed by return of right knee arthritis, coupled with onset of symptomatic uveitis of the left eye. despite systemic corticosteroids and anakinra and il- blockade, the patient was again admitted shortly thereafter to the hospital with arthritis, fevers, rash and abdominal pain. there was concern for evolving hlh and the patient was ultimately transferred to cincinnati children's for further evaluation and treatment. pertinent inflammatory biomarkers at that time included sil- r of , units/ml and il- level of , pg/ml. in addition to anakinra and systemic corticosteroids, the patient was started on tadekinig alfa (recombinant human il- binding protein) as part of a prospective study. hlh flare ultimately resolved without use of antineoplastic agents, and the patient was discharged home. soluble il- r levels since normalized, and il- levels decreased to less than pg/ml. the patient has been doing well on anakinra and tadekinig alfa, though continues to experience mild to moderate right knee effusion. this case suggests il- inhibition may be an effective therapeutic approach for patients with xiap deficiency. in the absence of neurological involvement and infectious trigger, ruxolitinib was initiated at a dose of mg/m /day, in combination with dexamethasone ( mg/m /day). this treatment led to rapid normalization of the neutropenia ( hours), complete resolution of the splenomegaly ( days) and disappearance of hlh biological markers (triglycerides levels in week, activated hla-dr+ cd t cells in weeks, fibrinogen levels in month), without the need for etoposide or serotherapy. dexamethasone was weaned every two-weeks and stopped after weeks. ruxolitinib was well-tolerated with no side effects. while in complete remission of her hlh, the patient then received alemtuzumab ( . mg/kg total dose) and a fludarabine-based myeloablative conditioning regimen. ruxolitinib was weaned over one week, and a / unrelated transplant was performed with success. the immediate posttransplant period was complicated by a veno-occlusive disease that responded rapidly to defibrotide and a corticosteroidresistant skin and ocular graft-vs-host disease (gvhd) despite a prophylaxis with ciclosporine and mycophenolate mofetil. gvhd was controlled by the reintroduction of ruxolitinib. at months post-hsct, her chimerism is % donor. to our knowledge, this case is the first description of a patient with primary hlh successfully treated in first intent by a combination of dexamethasone and ruxolitinib prior to hsct. our observation suggests that this targeted and less-toxic treatment regimen, that does not include etoposide nor high-dose alemtuzumab, is effective, well-tolerated and could be used in first intent to treat primary hlh. abstract/case report text presentation a -year-old girl, presented in ambulatory consultation, with a -year history of recurrent fever, influenza-like symptoms (sore throat, malaise), associated with self-limited painful genital ulcers (just within the period of fever). the first episode was characterized for an fournier's infection, requiring in-hospital treatment, multiple surgical procedures, antibiotics and hyperbaric oxygen therapy. after that catastrophic debut, she was diagnosed approximately episodes per year pharyngitis (with fever, malaise and sore throat) treated with corticoids, antibiotics and topic medication. the last year, noticed that every episode of fever (total of ) were associated with one or several genital lesions. the patient had no relevant medical history, she didn't receive long-term medication, she received all immunizations, she was sexually inactive, and hadn't apply any topic medication or product on the vulva, there was no trauma history, psychological medical history or sexual abuse. episodic gynecologic examination showed her labia minor several lesions, fibrinous, soft ulcerations on their inner aspect, these lesions had a symmetrical appearance, known as kissing lesions; no vulvar swelling, vaginal discharge or lymphangitis were noticed. there were no other skin or mucous membrane lesions (figure ). investigations viral (hiv, hbv, hcv, ebv, cmv) and treponemal (tpha-vdrl) serologies were negatives. erythrocyte sedimentation rate (esr) and pcr analysis within ulcers episodes were positive. specific antibodies (cardiolipin, anti ro/ss-a, anti la/ss-b, anti ccp, anti ena, anti gliadin, anti tpo anti tpo) serologies were negative. otherwise, important elevation immunoglobulin d was observed ( , mg/dl, twice the normal value): mild elevations of immunoglobulin m and immunoglobulin a were observed. serum subtypes of immunoglobulin g and immunoglobulin e were normal. leukocytosis with monocytes elevation and an increase of lymphocytes b were present. (table ). discussion lipschütz ulcers are uncommon and an often unknown entity for physicians, but it is important to recognize and include it in the differential diagnosis of vulvar ulcerations. this condition is characterised by self-limited painful ulcerations of the vulva or lower vagina in adolescent or young women, non-sexually transmitted, and usually preceded by influenza or mononucleosis-like symptoms. hyperimmunoglobulin d syndrome (hids) is characterized for unremitting fever lasting four to seven days and the presence of palpable tender lymphadenopathy, splenomegaly, arthralgia/arthritis, abdominal pain, and mucocutaneous manifestations. laboratory findings suggestive of hids include elevated age-specific serum immunoglobulin d (igd) and/or immunoglobulin a (iga) levels, elevation of acute phase reactants, and urinary excretion of mevalonic acid during, but not between, attacks. the diagnosis is established if an elevated age-specific level of igd is detected. iga levels are typically measured at the same time but are not required for diagnosis. elevated serum igd is not specific for hids and can occur in patients with certain neoplastic, infectious, heritable, and idiopathic disorders. in the present case report, the patient was treated with colchicine, with favorable evolution and free from new events. levels of ig d, platelets and monocytes remain high. we describe a young female patient presenting recurrent lipschütz ulcers, fever and elevation of serum immunoglobulin d, suggesting that hids could be associated with genitalia ulcers. ( ), and transmission in vivo in extremely low birth weight infants was considered ( ) . based on these considerations, the risk/benefit was considered favorable for restarting pasteurized donor breastmilk feeds. given his small size and young age, we had significant concerns about using ganciclovir prophylaxis and opted to hold this and monitor weekly cmv pcr. the child has been titrated up on these feeds, is gaining weight appropriately, and has had weekly cmv pcrs which are negative x since restarting donor breastmilk. t cells, last checked at weeks of age ( weeks gestational age) remain essentially absent. given the lower risk of nec in premature infants with breastmilk-based enteral feeds, a broader, multi-institutional study is warranted to best examine the safety of pasteurized donor breastmilk in infants with scid and complete digeorge syndrome. transfected with a reporter plasmid (for luciferase), wt or mutant-stat plasmids. nk cell cytotoxicity was measured by cr release assay. we used multiparametric immune profiling to dissect the effect of stat -gof mutations on nk cell developmental phenotype. results: similar to our previous studies, we observed higher levels of stat phosphorylation after two hours of stimulation from the dbd mutation compared to the ccd mutations. the stat activity assay confirmed gain of function observed by flow cytometry, but this activity was higher in k q mutant and d e mutant (ccd-closer to dbd) than v i mutant. all patients demonstrated low nk cell lytic unit compared to healthy donors. interestingly, we observed a correlation between low lytic unit and lower numbers of cd dim perforin + cd + nk cells; much lower in patient with k q mutation. stat -gof patients showed a significant decrease in total nk cell numbers and impaired nk cell maturation was characterized by low expression of cd , and higher levels of immature nk cell markers (cd , nkg a, cd b). conclusions: these data suggest that impairment of nk cell function is affected by the location of the stat mutation and continues to be the case in novel mutations identified. the identification the genotype/ phenotype correlation in the spectrum of the nk cell defect in stat gain-of-function mutants may help to better understand the molecular basis for stat activation and/or function to predict clinical manifestations of disease and ultimately treatment regimens. mutation specific analysis after an amniocentesis showed that one twin was a carrier (l m) and the other was a compound heterozygote (r c and l m). after birth, twin a had a mildly low erythrocyte ada level ( . nmol/h/mg; normal range + ) with normal metabolites and a normal immunophenotype, similar to both parents. twin b showed a normal absolute lymphocyte count and mitogen proliferation, normal t lymphocyte subsets, mildly low b and nk cells with % naïve t cells and a normal trec assay. erythrocyte ada levels were absent in peripheral blood, with mildly elevated metabolites [daxp= . μmol/ml rbc (normal < . ) and %axp= . (normal < . )]. weekly recombinant ada enzyme replacement therapy (ert) was started at week of life with subsequent normalization of the metabolites by week . absolute lymphocyte, t cell subsets were normal at birth but continued to rise slightly above normal range after starting ert. b and nk cell counts were mildly low at birth but normalized by week . genetic testing confirmed the prenatal genotypes in the twin girls. the patient is now months old and doing well with no history of infections. her twin was not an hla match and family is currently awaiting gene therapy approval. discussion: ada deficient patients show substantial clinical and metabolic heterogeneity that tends to correlate with the genotype but phenotypic discordance occurs even within the same genotype. we describe an infant with prenatally diagnosed compound heterozygous mutations in the ada gene (grade-i: r c and grade-ii: l m). ada alleles are graded from -iv with increasing ada expression and decreasing severity respectively. there are reports of children with grade i/iii allele combinations with delayed, late and partial phenotypes. two siblings have been reported with l m allele (grade iii) in combination with a different grade i allele (r q), presenting with combined immunodeficiency at and months. the specific allele combination from our patient has not been previously reported, however, we expected that the grade-i allele likely would be more deleterious than the grade-iii allele. in our case, predicting a future phenotype remains a challenge, creating a dilemma regarding management strategies. however, with only mild metabolite elevations in our patient after birth, we may speculate whether the prenatal diagnosis with early ert precluded the development of a full immunophenotype and it remains to be seen whether nonimmune sequeli may be prevented. conclusion: children with compound heterozygous mutations in the ada gene can pose diagnostic and therapeutic challenges, especially due to the associated metabolic and clinical phenotypic variability. early recognition and treatment may potentially alter long-term morbidity and mortality. (ipex)-like phenotype. immunodeficiency is often combined with impairment of the humoral and cellular compartments. hematopoietic cell transplant (hct) can resolve disease-related manifestations in stat -gof, but overall survival is poor and there is a high rate of secondary graft loss in transplanted patients. jakinibs are a class of medications that block cytokine-induced jak/stat activation. ruxolitinib preferentially inhibits jak and jak and has been used as precision-directed therapy for treatment of stat -gof related manifestations with success in stabilizing and in some cases reversing organ-specific manifestations. the utility and safety of jakinibs for long term treatment of stat -gof and in the prevention of disease-related manifestations is not known. as such, hct is often pursued for patients once disease-related manifestations are controlled with jakinibs. we present a patient with stat -gof mutation with gradual secondary graft loss following hct years ago, that has had continued disease progression despite chronic ruxolitinib treatment. case presentation this is a years-old male diagnosed with a de novo heterozygous stat mutation (c. a>g/a) at age , years following hct for ipex-like disease. he has been treated with ruxolitinib for the last three years. this patient initially presented at months of age with wasting enteropathy, failure to thrive, early-onset type diabetes and hypothyroidism. he had frequent upper respiratory infections during childhood including mycobacterium fortuitum mediastinal lymphadenitis. at years he underwent / matched, unrelated bone marrow transplant following reduced-intensity conditioning. mixed donor chimerism was present in the first days following hct, and he continued to have a slow progressive decline of donor chimerism with full graft loss ( % whole blood donor chimerism) by age . at age , enteropathy returned leading to cachexia and tpn dependence. concurrently, he had recurrent upper respiratory tract infections, lymphopenia, and hypogammaglobulinemia. imaging showed bronchiectasis and lung function was consistent with obstructive lung disease (fev : . l fvc: . l dlco: . ml/min/mmhg). initiation of ruxolitinib at age resolved his enteropathy with discontinuation of tpn and > -pound weight gain. enteropathy has not returned. pulmonary clearance measures have also been employed. dlco initially improved (dlco: . ml/min/mmhg) but obstructive lung pattern continued (fev : . l fvc: . l). after initial improvement, dlco began to decline. over the last years and despite treatment with ruxolitinib, lung function has deteriorated with worsened fev ( . l), fvc ( . l), and dlco ( . ml/min/mmhg). with this progressive decline, the family is now pursuing second hct. discussion jakinibs apply precision-directed therapy for immune dysregulatory features of stat -gof. their use leads to substantial disease control and clinical improvement but does not prevent disease progression. jakinibs should be used as a bridge to definitive therapy with hct in patients with stat -gof mutation. a recent large registry study showed a higher risk of infections (hr . , % c.i . - . ) in children with thymectomy as compared to surgery controls, in addition to demonstrating differences in the risk of cancers, autoimmunity and atopy. limited small studies have described some risk factors for altered immune consequences; however, specific predictors of infections among children with congenital heart disease (chd) undergoing thymectomies have not been systematically assessed. among children with chd and thymectomy, we sought to characterize children with and without reported infections within years postthymectomy and identify predictors of bacterial and viral infections. methods: using a retrospective chart review (institutional irb approved) from / / and / / , we identified children with chd that underwent thymectomy and excluded any known conditions associated with immunodeficiency and those with less than -month follow-up post-thymectomy. first absolute lymphocyte count (alc) after thymectomy was stratified using a cutoff at % of the lower limit of age-adjusted normal values (alc value < % vs alc value > % of the lower limit of age-adjusted normal levels). we sought to assess predictors of reported bacterial (positive blood, cerebrospinal fluid, respiratory cultures and chest-x-ray confirmed pneumonia) and viral infections (positive viral pcr tests) within years postthymectomy. results: we identified children with chd who had thymectomies, of which, % ( / ) were male. the median age at thymectomy was months (interquartile range months- . years); % ( / ) underwent a complete thymectomy; and % ( / ) developed a chylothorax within week post-thymectomy. a substantial proportion of children had an alc below % of the lower limit of age-adjusted normal levels after thymectomy ( % [ / ] pre-thymectomy vs % [ / ] postthymectomy). among children with chd post-thymectomy, % ( / ) and % ( / ) reported bacterial and viral infections within years, respectively. children with post-thymectomy alc values below % of the lower limit of age-adjusted normal levels had higher odds of reported bacterial (or . , % c.i . - . , p= . ) and viral (or . , % c.i . - . , p= . ) infections post-thymectomy as compared to those with an alc greater than % of the lower limit of ageadjusted normal levels (multivariate logistic regression). there was no association with the type of thymectomy (partial vs complete), age at thymectomy, weight at thymectomy, sex or prematurity. conclusions: among children with congenital heart disease with no known immunodeficiency undergoing thymectomy, alc below % of age-adjusted normal levels post-thymectomy may be associated with higher odds of bacterial and viral infections. a retrospective study design with a small sample size poses several limitations; however, this study suggests that post-thymectomy absolute lymphocyte values may be a potentially useful marker to identify higher risk patients in this population. radiological assessments esp. in the ct chest is commonly performed, but has associated radiation exposure and pulmonary function testing, at times, maybe insensitive to small changes in lung pathophysiology. many pids may have overlapping features with short telomere syndromes (sts) a, which are accelerated aging syndromes affecting hematopoietic, pulmonary, hepatobiliary and/or immunological systems, unified by a high cell turnover in these organs. clinical assessment of ageappropriate telomere length (tl) is performed using flow cytometry & fluorescence in-situ hybridization (flowfish). methods: we retrospectively analyzed telomere lengths in lymphocytes and granulocytes using the flow cytometry and fish method .flowfish testing was done at reference laboratories in johns hopkins university (jhu, usa).approval was obtained from mayo's institutional review board. data abstraction and analysis was done using the software jmp. results: patients were included in our analysis with females ( %) and males ( %).the median lymphocyte count of our cohort was . ( . - . ).the telomere length was strongly associated with the presence of lung disease (p= . *) and the presence of interstitial lung disease closely paralleled the changes in telomere length (delta-as compared to age adjusted normal percentiles lengths). shorter lymphocytic telomere length was associated with more severe reduction on total lung capacity (tlc; p= . *). conclusion: shorter lymphocytic telomere length served as a reliable biomarker for interstitial lung disease in pid patients. this may open up newer avenues for assessment of aging pathways in pid and may offer the option of using senolytic therapies in pids. mutations in the il- receptor common gamma chain gene (il rg) result in x-linked severe combined immunodeficiency (scid). the common gamma chain is shared by il- , il- , il- , il- , il- and il- receptors. x-linked scid typically presents with low or absent t and nk cells and normal or elevated numbers of b cells. we report a case of x-linked scid with elevated b and nk cell numbers (t-b+nk+). the male patient had an abnormal newborn screen for scid in north carolina. lymphocyte enumeration performed at days of life showed cd + cells/mm , b cells/mm , and nk cells/mm . he had no naïve t cells. repeat lymphocyte enumeration two weeks later showed that the cd + count had increased to /mm . only . % ( cells/mm ) were cd ra+ naïve t cells. he continued to have elevated b cell and nk cell numbers. chimerism studies revealed the presence of % female cells in mitogen-stimulated pbmc by fluorescence in situ hybridization, indicating the presence of transplacentally transferred maternal cells. lymphocyte proliferation responses to pha and cona mitogen stimulation were very low (less than % of normal). immunoglobulin levels were igg mg/dl, igm mg/dl, and undetectable iga and ige. genetic studies revealed a missense mutation in il rg, c. c>t, resulting in an amino acid substitution (p.ala val) in the extracellular domain. family testing showed that the patient's mother was a carrier for this variant. the father and the two healthy older brothers did not have this variant. of note, the family history was significant for lateral maternal male early deaths. at weeks of age, the patient received an unfractionated bone marrow transplant from his hla-identical brother without conditioning or gvhd prophylaxis. at the time of this report's submission, he is weeks post-transplantation and has had successful engraftment (whole blood-cd + fraction was composed of > % donor cells). he also now has normal t cell proliferation in response to mitogens and normal levels of all immunoglobulins. genetic defects that cause primary immunodeficiency can have variable phenotypic presentations. the patient's phenotype was atypical in that he had elevated nk cell numbers. to further evaluate these cells, we checked for stat phosphorylation following il- stimulation of abstract/case report text background: stat gain-of-function (gof) mutations cause a multisystem disease of early onset autoimmunity and lymphoproliferation, severe post-natal growth restriction, and recurrent and/or invasive infections. treatment of the autoimmune and auto-inflammatory features of stat gof patients relies heavily on immunosuppression and is often challenging. the full scope of phenotypes, treatments and outcomes may be broader when analyzing a substantially larger cohort than those already reported. methods: we gathered and analyzed data on patients from centers world-wide with confirmed gof mutations in stat . retrospective chart reviews were performed in accordance with all local ethics and irb committees to determine clinical manifestations, immunophenotype, treatment regimens, success of treatment methods, and overall survival. funcitonal transcriptional activity was assessed by luciferase reporter assay on each individual mutation. results: fifty-nine individual mutations were identified and all conferred gof by a validated luciferase assay. there were mutations in the nterminal domain, in the coiled-coil domain, in the dna binding domain, in the sh domain, and in the transactivation domain with the overwhelming majority being missense mutations. median age at presentation was approximately years; % of subjects are male and % are female. immunodysregulatory features presented in all patients. autoimmune cytopenias were the most common occurring in % of subjects (n= ), followed by lymphoproliferation in % (n= ) with increased frequencies of double negative (cd -cd -)t cells being found in % of of patients tested, enteropathy in % (n= ), endocrinopathy in % (n= ), interstitial lung disease in % (n= ), dermatitis in % (n= ), and inflammatory brain disease in . % (n= ). growth failure was present in % (n= ) with half of those patients having concurrent enteropathy. infections were reported in % of the cohort to include recurrent and/or invasive viral, bacterial, opportunistic, fungal, and mycobacterial infections. prominent abnormalities of immunophenotyping included t cell ( %) and b cell ( %) lymphopenia with reduced t cell proliferation in response to mitogens or antigens in % of those evaluated patients. fifty-nine percent of the patients hypogammaglobulinemia while % exhibited poor specific antibody responses to recall antigens. overall survival was % at data collection.treatment of stat gof patients often included multiple agents: ivig , chronic and pulse steroids, mtor inhibitors, calcineurin inhibitors, rituximab, mycophenolate mofetil, alemtuzumab, tocilizumab, and jakinibs. those started on jak inhibition showed improvement in clinical symptoms and, to date, there are stat gof patients on targeted jak inhibition. thus far, patients have undergone bone marrow transplant with a % survival rate. discussion: stat gof mutations were first reported in to cause a heterogeneous syndrome of autoimmunity and lymphoproliferation with immunodeficiency and infection susceptibility. earlier treatment with targeted therapy such as jak inhibitors has led to reduced disease morbidity. we report the largest cohort of stat gof patients collected through a multi-national collaboration of the longitudinal data and natural history of stat gof disease. understanding the heterogeneity of presentation and key features that will lead to proper diagnosis and early treatment in an effort to prevent long term disease associated sequelae. we present the case of month old male with a novel heterozygous mutation in tcf and two previously unreported phenotypes: ) absent circulating cd + b cells yet preserved immunoglobulin synthesis and vaccine responses and ) significant thrombocytopenia that improved with immunosuppression. there is also a striking family history of two half-sisters who died during early infancy with similar clinical and lab findings and the same genetic change. the infant boy was born at term with respiratory failure and generalized rash. at birth he had thrombocytopenia ( k/ul) and lymphopenia ( / ul). initial absolute cd + t cell count was low ( /ul), yet he had normal thymic output and proliferative responses to mitogens ruling out scid. cd + b cells were < /ul and bone marrow biopsy revealed decreased hematagones, yet he had a normal igm level ( . mg/dl) elevated iga ( mg/dl), and elevated ige ( iu/ml). igg levels were initially obscured by maternal igg and ivig; in turn he made positive titers to diphtheria and tetanus vaccination. the infant has maintained his own igg production. rapid genome sequencing revealed a heterozygous predicted deleterious vous in tcf , in the second transactivation domain (c. c>t, p.pro ser). the same change in tcf was identified in the deceased half-sisters as well as the father: all infants had different mothers, suggesting autosomal dominant inheritance. the sisters had similarly severe thrombocytopenia and absent circulating b cells; their causes of death were not completely understood. the year-old father has normal platelet levels, very low cd + b cells ( /ul), elevated igg ( , mg/dl) and ige ( , iu/ml), and normal levels of iga and igm. the father also has an elevated number of cd + t cells ( , /ul) with an increased percentage of t cells expressing hla-dr ( %). in the months after birth, the infant boy continued to require frequent platelet transfusions. despite the persistent t lymphopenia, there was evidence of increased t cell activation with elevated levels of soluble il- r ( pg/ml) and increased percentage of cells expressing hla-dr ( %) cd ( %), cd ( %), cd ( %), and cd ( %). a -day trial of prednisone was associated with an increase in his platelet count to > k/ul. he was switched to rapamycin as a steroid-sparing agent, and his platelet count has remained > k/ul for several weeks without transfusions. interestingly, his b cell counts also improved after the steroid trial ( /ul) and his absolute lymphocyte count is normalizing on rapamycin. a potential mechanism could be rapamycin decreasing t cellmediated destruction of platelets or b cells. reassessments of t cell activation markers and b cell phenotyping while on rapamycin will be done in the future. in contrast to multiple published cases of tcf mutations associated with complete agammaglobulinemia and absent b cells, we present a case of an infant with absent b cells yet preserved humoral function as well as severe thrombocytopenia responsive to rapamycin. in collaboration with colleagues at nih, studies are underway to understand whether/how the unique change in tcf is related to either phenotype described above. abstract/case report text background: childhood-onset, chronic, multi-system inflammatory diseases are increasingly being characterized as monogenic inborn errors of immunity. arpc b deficiency is a recently described, rare combined immunodeficiency characterized by recurrent/severe infections, a variety of autoimmune manifestations and platelet defects. we describe a case of arpc b deficiency identified in an adult patient with recurrent ulcers/ bechet-like disease, non-malignant lymphoproliferation and intermittent microthrombocytopenia. patient case: at year of age, our female patient was diagnosed with behcet disease based on a history of bloody stools at months, oral ulcers at months and vulvar lesions at year. she underwent rheumatology evaluations for inflammatory arthritis, episcleritis, eczema, vasculitic ulcerating nodules of the trunk, perineum and extremities, and verrucae forming flat plaques similar to epidermodysplasia verruciformis without a unifying diagnosis. other infections include otitis media, sinusitis, pseudomonas ecthyma gangrenosum, cervical lymphadenitis, and pneumonia. at years old, the patient was referred to our immuno-hematology comprehensive program clinic with a concern for malignancy versus a primary immune regulatory disorder (pird). she had a -month history of drenching night sweats, urticarial plaques, edema in her extremities and diffuse cervical, axillary and inguinal lymphadenopathy. past complete blood counts showed intermittent mild microthrombocytopenia. lymph node biopsies were negative for a neoplastic process but identified plasmacytosis, including focally increased iga-kappa+ plasma cells. expert review of the lymph node biopsy, and further evaluation excluded multicentric castleman disease. consideration was also given to autoimmunune lymphoproliferative syndrome (alps)-like disorders; however, her alps flow cytometry panel was nondiagnostic. her basic immune evaluation showed severe t cell lymphopenia (cd + cells/ cm, cd + cells/cm, cd + cells/cm) with adequate b and nk cells, normal lymphocyte proliferation to pha and pwm, and dysgammaglobulinemia with igg g/dl, iga g/dl, igm g/ dl and ige g/dl. due to concern of an underlying pird, a primary immunodeficiency panel was sent for gene analysis with negative results. however, trio clinical exome sequencing identified biallelic variants in the gene arpc b. one allele has a truncating, nonsense pathogenic variant in exon denoted as c. g>t, p.glu ter. the other allele has a likely pathogenic variant in intron denoted as c. - a>g, resulting in disruption of the canonical splice acceptor for exon . this is predicted to cause exon skipping, with an in-frame deletion of amino acids coded by exon . conclusion: this case highlights the value for evaluation for pirds in patients presenting with behcet-like disease, particularly in the context of other autoimmune manifestations and/or microthrombocytopenia. it also underscores that patients with arpc b deficiency may present with chronic non-malignant lymphoproliferation. moreover, this patient emphasizes the value of exhaustive genetic testing for complex immunologic phenotypes. abstract/case report text lipoyltransferase gene defect is associated with severe mitochondrial dysfunction disrupting lipoic acid biogenesis. clinical manifestations associated with early seizures, hypotonia, cardiomyopathy and pulmonary hypertension and encephalopathy. early neonatal death due to sepsis and cardiovascular collapse is commonly seen. the patient is a week preemie male with congenital heart disease who developed severe intractable lactic acidosis on day of life with increased excretion on organic acids of -methyl- , dihydroxybutyric acid. a mitochondrial disorder , echs or hibch deficiency was suspected. at mo of age the patient was admitted for apneic spells and respiratory compromise. he was found to have elevated crp associated with rhinovirus infection and gram-negative bacteremia. due to the history of failure to thrive and sepsis, immunology was consulted. immunologic work up indicated normal b, t and nk cells with normal dhr, but showed agammaglobulinemia. the patient was started on ivig and whole exome sequencing was done. molecular analysis showed compound heterozygote mutations in the lipt gene: c. g>a (p.arg gln) and c. t>g (pval gly). subsequent biochemical analysis also showed biochemical abnormalities consistent with lipt defect. lipoyltransferase is an enzyme involved in activation of a number of enzymes requiring lipoic acid. it is involved in lipoic acid synthesis. lipoic acid is required for the activity of pyruvate dehydrogenase, alpha-ketoglutarate dehydrogenase, and branched-chain alpha-ketoacid dehydrogenase. the literature indicates that most patients with lipt defect have a severe, often fatal course. the patient is now almost years old and has stable clinical course without any major infections. he certainly has significant hypotonia and developmental delay. in conclusion, we are presenting the first case of lipt gene mutations associated with agammaglobulinemia who responded well to ig supplementation therapy. our immunologic findings in this case highlights the importance of immunodeficiency work up in challenging cases. as we see more cases lipt gene mutations, we will better understand the clinical spectrum. abstract/case report text a now -year-old male was initially evaluated for concerns regarding food allergy, eczema, food protein-induced enterocolitis syndrome, and failure to thrive. he had reactions of varying severity to multiple foods. these usually involved immediate urticaria or prolonged vomiting, diarrhea, and abdominal pain. ige and skin prick testing was performed to suspected foods and was positive to milk, egg, pork, wheat, peanut, pecan, coconut and corn. these foods had historically caused reproducible immediate symptoms. testing was negative to other suspected foods. he developed an oral aversion and extremely restricted diet. symptoms of abdominal pain, hematochezia, rashes, arthralgias, headaches, fatigue, dyspnea, and palpitations increased. urticaria and severe abdominal pain with vomiting and diarrhea continued intermittently without identifiable triggers on a restricted diet. laboratory markers demonstrated elevated inflammatory markers, anemia, iron deficiency, vitamin b deficiency, and vitamin c deficiency (scurvy). gastroenterology work up did not identify any pathology. gastrointestinal symptoms did not respond to treatment with multiple gerd medications or oral steroids. baseline tryptase was elevated. low histamine diet was initiated and repeat tryptase remained elevated. fractionated tryptase revealed normal mature (beta) tryptase with elevated total tryptase, negative genetics for c-kit mutation, normal urine prostaglandins. family members had tryptase levels drawn. one parent and sibling had elevated tryptase levels, while the other parent's tryptase was normal. hereditary alpha tryptasemia syndrome is defined by elevated blood tryptase levels and symptoms involving multiple organ symptoms. patients with elevated tryptase levels without symptoms are defined as having hereditary alpha tryptase trait. there is significant variability regarding which patients are symptomatic. organ symptoms that may be involved include skin, gastrointestinal, neurologic, connective tissue, cardiac, neuropsychiatric. severe allergic reactions such as anaphylaxis can occur. increased blood levels of the protein tryptase are caused by extra copies of the alpha tryptase gene (tpsab ). treatment is usually directed at specific symptoms, antihistamines, and mast cell stabilizers. research continues into additional treatment options. this patient was started on cromolyn and long-acting antihistamine. his gastrointestinal symptoms and rash/urticaria improved, and he began tolerating a small, but increased, variety of foods. the majority of his constitutional symptoms of fatigue, arthralgias, weakness resolved as he began gaining weight, and hemoglobin, vitamin c and b normalized. his sibling was evaluated and noted to have food allergy, asthma, abdominal pain, gerd, and eczema. she was also started on cromolyn and antihistamines which improved her gastrointestinal symptoms. parent with elevated tryptase was recommended to be evaluated further with allergist. this is an example of a patient with elevated tryptase and multiple organ system involvement. some of his signs and symptoms responded to mast cell stabilizing and antihistamine medications. patients with history of recurrent episodes of allergic reactions to foods and multiple constitutional symptoms would benefit from baseline tryptase levels. family members should also be tested if the patient has elevated tryptase. multiple studies have been published looking at the rates of scid in the united states. the estimated rate of scid prior to screening was per live births. post screening implementation, on average rates of scid were found to be closer to in live births. results : development of a t cell receptor excision circles (trec) scid screen in alberta involved the screening of anonymous term neonates using quantitative pcr for trecs. the cycle threshold for the control gene, rnasep, was set at . as % of our population had a cycle threshold < . ( % ci [ . , . ]). from those bloods spots with adequate dna, a final trec cut off of was chosen, as it would give an accuracy of . %, and fairly low false positive rate of . % ( % ci [ . , . ]). since starting a population based screen for scid in june of , we have identified cases of scid and cases of low trec not caused by scid. to date we have detected one case of reticular dysgenesis, cases of ada scid and one case of x-linked scid. other causes of lymphopenia in the neonatal period detected with abnormal trecs include one syndrome associated with variably affected cellular immunity (charge) and cases of secondary lymphopenia including four cases of prematurity, three cases of diaphragmatic hernia or gastroschisis, four patients with underlying cardiac disease, and one patient with severe hydrops. discussion : canada has multiple unique populations with increased risk of scid. the estimated rate of scid in canada prior to implementation of a population based screen was . per live births. the rate within canada's first nations, métis and inuit populations is . per live births. prior to scid screening, alberta had cases of scid identified between - with an estimated rate of per live births. to date, our screen in alberta has identified cases of scid with a rate of per live births which is significantly higher than previously estimated. given that early diagnosis and definitive management through bone marrow transplant or gene therapy has been shown to reduce mortality this screen will help reduce morbidity and mortality in this vulnerable population. abstract/case report text introduction: human herpesvirus (hhv- ) has the ability to integrate its genome into host telomeres. if this integration occurs in gametes, then the virus can be genetically transmitted and offspring will carry a copy of chromosomally integrated hhv- (cihhv- ) in each somatic cell. this can lead to false attribution of infectious and non-infectious presentations of hhv- , and make the diagnosis of active hhv- infection difficult. we present the case of a patient with meningoencephalitis attributed to hhv- and persistently elevated blood levels of hhv- by pcr concerning for primary immunodeficiency who was discovered to have cihhv- . case description: a -year-old female who carried a past diagnosis of hhv- meningoencephalitis was seen in immunology clinic for follow up of persistently elevated levels of hhv- dna in her blood by pcr. she was born at weeks and as an infant had failure to thrive (ftt), anemia and a varicella like rash after varicella immunization. at the age of , she received flumist vaccine and developed a fever the following day. over the next few days, she developed lethargy, altered mental status, headache, photophobia, seizure, papular rash and oral ulcers. she was admitted to the hospital and csf studies were consistent with viral meningoencephalitis ( wbc, l, m, rbc) although hsv, cmv, ebv and enterovirus were negative. she was treated for presumed hsv encephalitis with days of iv acyclovir and days of high dose steroids. one week after discharge, she again developed papular rash on feet, headache, oral ulcers and lethargy. she was admitted and csf studies this time showed only wbc but positive hhv- . blood and skin swab were also positive for hhv- . immunology was consulted while admitted and work up for primary immunodeficiency was initiated. her work up was normal including responses to vaccine titers, complement studies, moderate in t cells, monocytes, neutrophils, lung and muscle, and low in skin, liver, heart and kidney tissues. analysis of each individual gene expression level by nanostring in comparison with healthy controls demonstrated significantly higher levels of the following irgs: ddx , epsti , gbp , ifi , isg , ly e, oas , oas , oas , rsad , rtp and socs . whole blood rna-seq was performed in patients and pathway analysis with the differentially upregulated genes demonstrated an enrichment of intraluminal vesicle formation and negative regulation of apoptotic signaling pathways. stimulation of pbmcs with the tlr ligands poly i:c, odn, and lps induced a fold increase in ifi and a -fold increase in ifna and ifnb transcription compared to baseline. one patient had constitutive upregulation of stat and stat in monocytes and of stat and stat in t cells. conclusion: we describe a novel immunedysregulatory disease caused by de novo truncating variants in samd l that presents similar to candle with neutrophilic pannicultis and points to an important role of samd l on regulation of adaptive and innate immune responses. acknowledgements: this work was supported by the nih irp of niaid abstract/case report text introduction: ataxia telangiectasia (at) is caused by a defect in the atm gene which is responsible for repair of damaged dna. it is a rare, devastating neurodegenerative disease that results in ataxia and telangiectasias, particularly of the sclera and skin. those with at are at increased risk for immunodeficiency and cancer. the immunodeficiency is variable and may result in deficiency in humoral and cellular immunity in some patients. here we present a patient with ataxia telangiectasia and hypogammaglobulinemia on immunoglobulin therapy who developed recurrent urinary tract infections (uti) and sepsis. case presentation: the patient is a -year-old female with ataxia telangiectasia and hypogammaglobulinemia who presented with three episodes of uti, one of which resulted in prolonged hospitalization due to sepsis and acute kidney injury (aki). she presented with days of flank and back pain and was hospitalized for days for e coli uti. she improved on iv antibiotics and was discharged home to complete treatment with oral ciprofloxacin. due to persistent emesis, she was readmitted weeks later with urosepsis and aki with a creatinine of . mg/dl, over times her baseline creatinine. after additional antibiotics and iv fluids, she improved clinically, renal function normalized and she was discharged home. renal ultrasound was unremarkable with no anatomical abnormalities. she was relatively healthy prior to this with only one episode of bacterial pneumonia in . she receives weekly subcutaneous immunoglobulin therapy dosed at mg/kg with normal igg levels ( , , mg/ dl). at baseline, she had high igm ( mg/dl) and low iga ( < mg/ dl) levels, as well as decreased t cells but normal nk and b cells ( cells/ul cd +, cells/ul cd +, cells/ul cd +, cells/ul cd +cd +, and cells/ul cd +). she has hyperglycemia (on metformin), hypertriglyceridemia (on atorvastatin) and hypertension (on losartan). she is thin and wheelchair bound with bilateral telangiectasias to the sclera, neck, and chest. she has occasional eye bleeding and epistaxis, presumably from her telangiectasias. she has good hygiene and good adherence to medications. she voids voluntarily, has no indwelling urinary catheter and is not sexually active. discussion: patients with ataxia telangiectasia may have frequent viral and bacterial infections, most frequently upper and lower respiratory tract infections, as well as wart and skin infections. based on our review, this is the first reported case of ataxia telangiectasia with hypogammaglobulinemia on immunoglobulin therapy with recurrent utis complicated by urosepsis and aki. despite adequate igg levels on immunoglobulin therapy, our patient continued with recurrent utis. it is uncertain whether her non-ambulatory status, hyperglycemia or related immunodeficiency are the causes for her increased susceptibility to utis. the literature reports patients with at and bladder wall telangiectasias can result in significant hematuria, and perhaps this may be a source of entry for bacteria and consequent development of uti. this suggests that patients with ataxia telangiectasia and recurrent utis may benefit from renal ultrasound and possible cystoscopy to better visualize telangiectasias. we recommend consideration of workup for recurrent utis in patients with ataxia telangiectasia. abstract/case report text objectives: patients with partial rag deficiency frequently present with humoral autoimmunity suggesting breach in tolerance mechanisms and subsequent expansion of autoreactive b cell clones. here we aim to trace polyreactive b cells and their descendants at b cell developmental stages through our in-house bioinformatic pipeline, immchaintracer (ict). methods: the b cell receptor (bcr) was expressed as monoclonal antibodies from single sorted mature naive b cells (n= - per donor) from patients with hypomorphic rag deficiency and healthy donors. xthe recombinant monoclonal antibodies were screened for polyreactivity (dsdna, insulin, lps and ifnα) by elisa. in parallel, igh repertoires were deep sequenced from sorted mature naïve, activated naïve and memory b-cell compartments. our in-house assembled bioinformatic pipeline called immchaintracer (ict) was applied to track down the descendants of cloned autoreactive igh sequences in repertoires of subsets above. results: igh sequences (n= including polyreactive, nonpolyreactive clones) from mature naive b cell from six patients with partial rag deficiency and healthy donors (n= including polyreactive, non-polyreactive) were analyzed with our novel inhouse bioinformatic approach to track lineage fate in repertoire at specific developmental stages. interestingly, . % of the patients' sequences and their descendants were identified in their mature naive, activated naive or memory b cell repertoires, while none of the analyzed healthy donor clones were found at later subsets. furthermore, genealogical analyses of related clones revealed lineage expansion and progressive positive antigen selection of the autoreactive clones in the patients. conclusions our findings demonstrate that peripheral tolerance checkpoint is broken in hypomorphic rag patients. our novel method enables tracing the fate of autoreactive naive b cells in the effector repertoires. we have shown that impaired b cell tolerance allows the expansion and combination of sirolimus and rituximab therapy controlled his autoantibody production which was an important goal for his autoimmune condition. we present a new treatment approach for anti-nmda receptor encephalitis. rituximab was tried on these cases before but the combination of sirolimus and rituximab therapy was never given before. we now recommended that on refractory cases of anti-nmda receptor encephalitis, combination of rituximab and sirolimus therapy can be tried. ( ) submission id# mosaic variants in immune function genes identified through exome sequencing stable with no interim infections and is doing well on thyroid hormone replacement. the current plan is for the patient to undergo a stem cell transplant for the arpc b deficiency as he is at high risk for recurrent infections and severe disease. although this gene mutation is rare, review of the current literature describes patients with this condition that have undergone stem cell transplant and have done well. at this time, this seems to be the best option for management, and it may potentially be curative. abstract/case report text introduction: common variable immunodeficiency (cvid) is a disorder characterized by impaired immunoglobulin production and frequent or recurrent infections, but also associated with an increased risk for developing malignancies such as lymphomas. although intravenous and subcutaneous immunoglobin g replacement has been successful in reducing the number of bacterial infections and prolonging survival, it fails to address other complications that arise from this disorder. we report a case of a patient with cvid who developed mycosis fungoides (mf). mf is a rare form of cutaneous t-cell lymphoma, occurring in about in , to , individuals, lack of treatment could potentially be fatal. case presentation: a -year-old caucasian woman with a history of ulcerative colitis, allergic rhino-conjunctivitis and tonsillectomy was referred to the immunology clinic for evaluation of low serum immunoglobulins. there was no family history of infections or immune deficiencies, but paternal grandfather had colon cancer and maternal grandmother had lung cancer. the patient reported frequent episodes of bronchitis, and sinus infections. immunizations were up to date for her age. medications included azathioprine, cetirizine, fluticasone nasal spray, hyoscyamine, montelukast, lactobacillus, omeprazole, and olopatadine ophthalmic solution. no history of frequent use of systemic steroids. initial serum immunoglobulins revealed normal ige and igm but low iga ( mg/dl, normal range - mg/dl) and low igg ( mg/dl, normal range - mg/dl). cbc with differential, lymphocyte subsets, c and c levels were normal. while she had adequate protective titers against haemophilus influenza type b, diphtheria and tetanus, titers against pneumococcus were < % protective and she failed to mount an adequate response to pneumococcal polysaccharide vaccine. given her diagnosis of cvid, she started scig ( mg/kg every weeks). a year later, she developed a bilateral nonpruritic rash in the abdomen and upper trunk. initial skin biopsy suggested a drug reaction. a subsequent biopsy revealed a superficial perivascular lymphoid infiltrate with focal epidermotropism and positive t cell receptor gamma gene rearrangement, consistent with mf. testing for cell t receptor beta gene rearrangement was negative. while mf treatment consisted of triamcinolone . % ointment, treatment for ulcerative colitis transitioned from azathioprine to vedolizumab. conclusions: cutaneous t cell lymphomas, although uncommon, can be seen in cvid. there are several reasons for the increased risk of lymphoma in cvid. the role of chronic infections and the development of lymphoma as of yet, is not clear. skin reactions to scig products in the areas of infusion are relatively common and resolve promptly. high index of suspicion is crucial in obtaining tissue sample to confirm or rule out malignancy therefore avoiding delaying proper treatment. abstract/case report text patients with lipopolysaccharide responsive beige-like anchor protein (lrba) deficiency present with a plethora of immune related defects including a defective humoral response characterized by low numbers of switched memory b cells and plasma cells, as well as an impaired production of antibodies, leading to recurrent infections. however, the molecular mechanisms behind the defective b cell response remain unknown. to gain better insights into the possible roles of lrba in b cell physiology, we screened for lrba-interacting proteins using computational predictions. twenty-seven proteins involved in vesicle trafficking and autophagy were identified as potential lrba-interacting partners. to validate those potential lrba interactions, we performed coimmunoprecipitations and proximity ligation assays (pla), finding that endogenous lrba interacts with the phosphoinositide kinase regulatory subunit (pik r ) in b cells. pik r (aka vps ) is the regulatory subunit of vps , the catalytic subunit of the pi k-iii complex, which acts as a positive regulator of autophagy by producing phosphatidyl inositol- phosphate (pi( )p). autophagy is a catabolic mechanism essential for cell survival and plasma cell differentiation. in fact, we observed that reduced lrba impaired the production of pi( )p upon autophagy induction. in addition, we observed in both lrba-deficient hela and b cells reduced mobility, abnormal accumulation and increased size of autophaghosomes, accompanied by an atypical lysosomal positioning. these abnormalities are due to a blockade of the autophagosome-lysosome fusion, as detected by reduced lc -ii lipidation upon autophagy induction in the presence of lysosome inhibitors. interestingly, lrba-deficient hela and b cells exhibited enhanced activity of mammalian target of rapamycin complex (mtorc ) signaling, a key suppressor of autophagy whose activation possibly contributes to defective autophagy. taken together, b lymphocytes lacking lrba can form autophagosomes but they fail to fuse with lysosomes. thus, we propose a role of lrba at late stages of autophagy through the binding to pik r . abstract/case report text apds caused by gain-of-function mutations (gof) in the genes (pik cd and pik r ), encoding for the p δ and p subunits of phosphoinositide -kinase δ (pi kδ), results in hyperactivation of the pi k/akt/mtor/s k pathway and lead to immune dysregulation, lymphoproliferation and immunodeficiency. apds manifests with respiratory tract infections, bronchiectasis, susceptibility to herpes group viruses, autoimmunity, cytopenia, lymphoproliferation and lymphoma. gastrointestinal system manifestations include enteropathy, colitis, and liver disease. eosinophilic esophagitis (eoe) or eosinophilic gastrointestinal disease (egd) have been under diagnosed in reported apds cohorts. objectives: to review the incidence, demographics and relevant clinical data for eosinophilic gastrointestinal disease in a single center apds cohort. methods: review of clinical and laboratory findings from apds patients followed at the nih clinical center, from to . results: patients were either historically diagnosed or actively studied at our center for egd. incidence of all egd is % in our cohort and all patients had mutations in pik cd, none in pik r . most patients also had multiple gi manifestations. conclusion: immunopathology and genetic predisposition leading to eoe is complex. eosinophilic gi disease including eoe appears to represent significant gi pathology in apds. this implies that activation of pi k pathway may be directly involved in the etiology of eoe. abstract/case report text introduction: dominant negative mutations in stat (lof stat ; job's syndrome) cause a primary immune deficiency characterized by eczema, recurrent skin and lung infections, and connective tissues and skeletal abnormalities. over the last several years, vascular abnormalities causing tortuous and aneurysmal middle-sized arteries have increasingly been recognized. our institution has been imaging prospectively the coronary and cerebral arteries since - for brain imaging and from - for heart imaging. the purpose of this review is to provide an update on the extent of clinical manifestations noted in hies (continued) pain improved but rapidly worsened with tapering of steroids. the pet/ ct at that time, showed extensive hypermetabolic areas in lymph nodes, femurs, left acetabulum, left pubic ramus, right ischium, sacrum, both iliac bones, eight rib, t and t , both clavicles, both humeral, manubrium, and extension into musculature. initial biopsies were culture negative, s was positive for mac. she was referred to the national institutes of health where she was diagnosed with autoantibodies to interferon-γ . prior to referral, she initially was treated with azithromycin, ethambutol (need to discontinue due adverse effects), amikacin, rifampicin, linezolid, with not no evidence of clinical response. she underwent debridement of epidural anterior abscess to t . surgery involved t - laminectomy, and curettage on several bones. she presented unable to walk secondary to pain and neuropathies. initial laboratory: crp: mg/l; wc . ; hgb: . g/dl; ana : . (strongly positive) cd : ul her first course of rituximab consisted of doses of gm at d , , and monthly thereafter for a total of doses with clinical and radiographic improvement. she was maintained on optimal antibiotics therapymeropenem, rifampin, azithromycin, moxifloxacin, and clofazimine. two years after she completed rituximab, she presented to her home hospital with increased left hip pain and biopsy grew mac. retreatment with rituximab failed to show clinical improvement. her medical regimen was augmented with tedizolid and bedaquiline, however no iv antibiotics were added. rituximab was reinitiated, after the progression of symptoms despite treatment with rituximab; bortezomib, was trialed using the schedule based on the multiple myeloma literature. she completed full cycles (two at the nih, three at home). she subsequently has had clinical improvement and is working again and has not had progressive neurologic decline. the titers of antibodies to interferon-· did not follow the clinical improvement. however, we plan to keep her cd + cells zero and continue the bortezomib given her clinical and radiologic improvement. abstract/case report text for patients with primary immunodeficiencies (pid), finding a genetically defined diagnosis can be critical for prognosis, treatment, and counseling. however, for many patients, determining a genetic etiology remains elusive despite routine gene panel and exome sequencing because of an inability to resolve variants of uncertain significance (vus). crispr-based genome editing could be used to address this need by introducing patient-derived vus into primary human immune cells for further study; however, existing techniques are limited by poor efficiency. we recently developed novel non-viral techniques for large gene editing in primary human t cells and hematopoietic stem cells (hscs). we achieve up to fold greater efficiency than existing tools using crispr cas ribonucleoprotein nanoparticles that are non-covalently linked to homology directed repair (hdr) template dna. whereas mutation analysis has previously been limited to expensive and time-consuming animal models and transformed cell lines, we now have the ability to rapidly recreate any mutation in the native gene locus in otherwise healthy primary human cells. we demonstrate this ability by using our technique to knock-in well characterized loss-of-function mutations in jak and il rg and gain-of-function mutations in jak that are known to cause severe combined immunodeficiency and tumor growth, respectively. we show that these recreated mutations have the expected effects on t cell proliferation and intracellular signal transduction in the setting of il- stimulation. we then use our technique to investigate a prototype case of an adult patient with the unusual combination of common variable immunodeficiency, inflammatory arthritis and uveitis, and neutrophilic urticaria. genetic testing in this patient had previously revealed heterozygous coding vus's in four genes previously associated with a pid disease, including jak , but none of the specific variants have been previously reported. it is thus not clear which mutation (if not more than one) causes this patient's dysregulated cell activation, which limits targeted treatment options with kinase inhibitors or future gene or cell therapy. by knocking our patient's jak variant into primary human t cells and comparing these cells to those carrying wild-type, known loss-of-function, and known gain-of-function mutations, we are able to rapidly characterize the functional impact of our patient's variant and isolate its effect on his complex phenotype. this in vitro genetic engineering approach thus allows patient-specific vus to be modeled directly in primary human immune cells with a rapid turnaround time that is relevant for clinical applications, including molecular diagnosis and screening of pharmacologic or gene therapies. further, similar strategies could be leveraged as a potential basis for future gene correction therapy. abstract/case report text definition : "at variants" comprise a heterogeneous group characterized by the later onset of clinical symptoms, a slower progression, a prolonged lifespan compared to most patients with at and decreased levels of chromosomal instability and cellular radiosensitivity. in these patients, telangiectasia and / or immunodeficiency may be absent, while neurological features are present.( ). material and methods :a years old girl, born out of a non-consanguineous parents with clinical picture consisting in progressive alteration of the march ( months), associated to exotropia, no weight gain or height, alteration of balance while she is sitting, she walks by herself. she has acute bronchiolitis. results and discussion: brain magnetic resonance without alterations. abdominal ultrasonography and cpk normal. ophthalmologist assessment found exotropia, he did not find telangiectasia. motor and sensitive neuro-conduction were reported normal. alpha fetoprotein ng / ml increased. karyotype xx, non-structural alterations. normal auditory-visual evoked potentials. whole exome sequencing (wes): identified the small homozygous pathogenically deletion ¬c. + _ + del taag; p.? have a spelling effect in the splicing. it is not present in the population database or not as a known variant in function mutation of smad has been shown to increase smad phosphorylation in the nucleus in fibroblasts. further research is needed to examine the role of this mutation in t and b lymphocytes, given the interesting immunological phenotype of this patient. publisher's note springer nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations. outbreak of mycoplasma pneumoniae-associated stevens-johnson syndrome characterization of children. with recurrent episodes of stevens johnson syndrome recurrent stevens-johnson syndrome secondary to mycoplasma pneumoniae infection recurrence and outcomes of stevens-johnson syndrome and toxic epidermal necrolysis in children syndrome after mycoplasma pneumoniae infection a review of causes of steven-johnson syndrome and toxic epidermal necrolysis in children submission id# expanded phenotype and genotype-phenotype correlations bsc(hons) msc bmbs mrcp dominant activating rac mutation with lymphopenia, immunodeficiency, and cytoskeletal defects an autosomal dominant scid form due to a gain of function mutation in the rac gene national institute of allergy and infectious diseases (niaid) professor biochemistry/molecular pharmacolog/nyu langone health canada head of the ircm bioinformatics core facility/montreal clinical research institute (ircm) associate investigator/national human genome research institute (nhgri) division of intramural research (dir), national institute of allergy and infectious diseases (niaid) translational and functional genomics branch/chief and senior investigator of the translational and functionanational human genome research institute (nhgri) division of intramural research (dir), national institute of allergy and infectious diseases (niaid) formula versus donor breast milk for feeding preterm or low birth weight infants cytomegalovirus (cmv) inacƟvaƟon in breast milk: reassessment of pasteurizaƟon and freeze-thawing prevenƟon of cytomegalovirus transmission via breast milk in extremely low birth weight infants submission id# the costa rican registry for primary immunodeficiencies department chief/department of pediatric immunology and rheumatology, national children´s hospital "dr. carlos sáenz herrera national children´s hospital "dr. carlos sáenz herrera" cid % (n= ), cid with syndromic features % (n= , at ), antibody deficiencies % (n= , xla ) ada deficiency, cd l-, flh- (syntaxin deficiency), ipex and osteopetrosis. nineteen patients developed malignancies, mainly non-hodgkin lymphomas among at patients conclusions: during the study period, pid were registered, mainly at and osteopetrosis cases. scid represented % and xla % of patients allergy and immunology/ department of pediatric immuunology and texas children's hospital director immunogenetics program/department of immunology, allergy and rheumatology at baylor college of medicine associate professor, director food allergy program/department of pediatric immuunology shearer center for human immunobiology associate professor/department of pediatric immuunology, allergy, and retrovirology granuloma faciale: successful treatment of nine cases with a combination of cryotherapy and intralesional corticosteroid injection intralesional steroid injection: a novel method to treat the symptoms of idiopathic granulomatous mastitis long-term effectiveness of intralesional triamcinolone acetonide therapy in orofacial granulomatosis: an observational cohort study sarcoidosis in the periocular scar as the first finding of systemic sarcoidosis: clinical-radiological characteristics a case of scar sarcoidosis of the eyelid submission id# null mutations: a gene dosage effect shubham goel, phd , hye sun kuehn clinical centre, nih, staff scientist/immunology service certified molecular geneticist/national institute of allergy and infectious diseases, national institutes of health genetic counselor/national institute of allergy and infectious diseases, national institutes of health division of intramural research; chief, immunopathogenesis section/laboratory of consultant/university hospital freiburg center for pediatrics and adolescent medicine cci -center for chronic immunodeficiency medical director professor/division of blood and marrow transplantation and cellular therapies new south wales, australia. professor of medicine and pediatrics/icahn school of medicine at mount sinai senior investigator/laboratory of clinical immunology and microbiology, national institute of allergy and infectious diseases, national institutes of health functional nk assay, t/b/nk panel, tlr assay phd staff scientist/translational autoinflammatory disease section, lcim, niaid, nih, bethesda, md research fellow/translational autoinflammatory disease section post-baccalaureate fellow/translational autoinflammatory disease section pediatric rheumatologist/department of pediatric infectious diseases and immunology shupyk national medical academy for postgraduate education alder hey children's nhs foundation trust hospital, liverpool, uk pediatric rheumatologist/department of pediatrics dra. liliana bezrodnik y equipo dra. liliana bezrodnik y equipo"-servicio de inmunología htal. de niños hospital de niños ricardo gutiérrez pathology department/children's hospital of philadelphia, philadelphia, pa post-doctoral fellow/translational autoinflammatory disease section results: we identified patients with de novo frameshift variants in samd l (c. dela, p.i lfs* reduced toxicity allogeneic hct with a busulfan, fludarabine regimen: a promising approach for non-cgd primary immune deficiencies requiring myeloablation? phd , blachy davila saldana college of medicine assistant professor/division of bone marrow transplant, immunedysregulation and immuno-hematology program, aflac cancer and blood disorders center instructor of pediatrics/division of bone marrow transplantation and immune deficiency, cincinnati children's division of bone marrow transplantation and immune deficiency, cincinnati children's national institutes of health certified molecular geneticist/national institute of allergy and infectious diseases genetic counselor/national institute of allergy and infectious diseases, national institutes of health operations manager and genetic counselor/national institute of allergy and infectious diseases collaborative bioinformatics resource/national institute of allergy and infectious diseases genome in a bottle analysis team/genome in a bottle consortium division of intramural research (dir), national institute of allergy and infectious diseases (niaid) medical genomics and metabolic genetics branch/national human genome research institute, national institutes of health table clinical and demographic characteristics, severity measures, and previous treatments for the pid cohort geographic region, n (%) north central , ( . ) ( . ) , ( . ) northeast , ( . ) ( . ) , ( . ) south ) ( . ) , ( . ) high-potency oral antibiotics , ( . ) ( . ) , ( . ) systemic high-dose corticosteroids copd, chronic obstructive pulmonary disease cd / cd : . , lymphocytes b , natural killer cell/mm . conclusions: although the atm activity corresponding to this new splicing mutation is unknown, it is presumed that it has some residual function, since splicing mutations is associated with better neurological prognosis have been reported (pidd) . at the immunodeficiencies research unit we discuss and pursue the molecular and genetic diagnoses for patients with suspected pid from all around the country. starting this year, we are processing and analyzing our own patients' wes results at the unit. evaluating the diagnostic yield of wes, as a measure of effectiveness or quality control, may result in process optimization and perhaps allow for better patient selection and resource allocation. objective: to describe and characterize our patients with suspected pidd whose dna samples were sent out for whole-exome sequencing; to analyze and compare our wes diagnostic yield after the first batches of patients; to identify patient attributes that may predict a positive diagnostic wes result. methods: genomic dna was obtained from whole-blood samples of patients with suspected pidd from hospitals in mexico city, monterrey and puebla. wes was performed using a ng sequencer (illumina hiseq) in new jersey (admera health, llc), with % coverage and a x depth of the idt xgen library, human genome version (december ). two fastq files for each patient sample were transferred back to our unit, where the bioinformatic workflow was completed. we used galaxy in the cloud for quality control, mapping & alignment, and detection of variants; variant effect predictor to process, map, annotate and filter variants; and igv (broad institute) and genome browser (ucsc) for visualization. we defined diagnostic yield as the proportion of patients with a genetic diagnosis after analysis of their wes results. we performed multivariate logistic regression, tree partitioning algorithm and linear abstract/case report text a -year-old male with x-linked chronic granulomatous disease (cgd) complicated by severe perianal disease and proctocolitis presented with two weeks of open draining lesions on the thighs, bilateral inguinal regions, and gluteal cleft. the wounds became excruciating and prevented normal ambulation. the patient was admitted for iv antimicrobial therapy, local wound care and systemic steroids. wound cultures, throughout his course, yielded growth of klebsiella pneumonia, candida parapsilosis, malassezia globose, escherichia coli, enterococcus faecalis and staphylococcus epidermidis allowing for directed antibiotic and antifungal therapies. despite improvement, the wounds persisted after several weeks of treatment. faced with recalcitrant cutaneous lesions despite aggressive systemic and topical therapies, we looked to alternative options. noting that other granulomatous diseases show response with intralesional corticosteroid therapy, we considered this for our patient ( , ) . for example, patients with idiopathic granulomatous cheilitis had a complete response after three monthly injections of intralesional corticosteroids ( ) . sarcoidosis patients also improve with intra-granuloma corticosteroid injection ( , ) . our patient received mg triamcinolone acetonide injections two separate occasions, administered in multiple open lesions at eight-week intervals. the cutaneous lesion improvement was gradual and complete resolution of the first open wound was noted fifty-two days from initial steroid injection. to our knowledge, intralesional glucocorticoid therapy has not previously been used to treat cutaneous disease in cgd patients. we are reporting the first cgd patient with successful lesion resolution following steroid injection as part of therapy. as such, we believe this case is significant and suggests that direct lesion injection with glucocorticoids can add to treatment options for cgd patients with recalcitrant cutaneous disease. serum igg, alone or in combination with iga and/or igm, were reduced in all patients. t-follicular helper cells were reduced only in patients carrying biallelic mutations (a.ii. , a.ii. and a.ii. ) but not in any patients with monoallelic tcf null mutation. t cell enumeration and function by means of proliferation was normal in all mutation+ individuals. no mutated (truncated) protein expression was detected from patients with either biallelic or monoallelic tcf null mutations. however, wildtype tcf protein was detectable in about half amount in heterozygous patients. cdna data showed either / or / wt/mutated transcripts ratios in homozygous or heterozygous individuals, respectively, suggesting mutated proteins instability; and all together, protein haploinsufficiency for the heterozygous cases. ex-vivo, cd l and il -induced plasmablast differentiation was found to be reduced in / patients tested ( biallelic patient a.ii. and monoallelic patients a.i. , b.i. and c.i. ). moreover, decreased igg, iga and igm production in vitro correlated with reduced plasmablast cell differentiation.in conclusion, all individuals carrying either mono-or biallelic null mutations have immunological penetrance of the b cell defect. however, while clinical penetrance was complete in patients with biallelic mutation, it was partial for those with monoallelic tcf null mutation suggesting a gene dosage effect for clinical penetrance. in addition, our study emphasizes that tcf is relevant to the plasmablast differentiation process as well as for ig production. further studies are being conducted to evaluate the individual roles of e and e on the immune and clinical features. background: heterozygous gain-of-function (gof) mutations in the stat gene result in a hyperphosphorylated state where patients develop recurrent or persistent chronic mucocutaneous candidiasis (cmc), other cutaneous mycosis, bacterial infections, disseminated dimorphic fungal infections and autoimmune disease. furthermore, the nk defect we characterized illustrated an immature cd dim nk cell subset with decreased expression of cd , perforin, cd , and impaired cytotoxic capacity associated with increased susceptibility to viral infections observed in these patients. methods: in this study, we evaluated patients with novel stat mutations (d e mutation, located in coiled-coil domain and k q mutation, located in dna-binding domain). a third patient with the previously reported v i mutation (ccd) was also recruited for this study. in vitro, pbmcs from these patients were stimulated with ifn-α for , , and minutes and levels of phospho-stat on cd dim nk cell subset were measured by flow cytometry. the stat activity (firefly and renilla luciferase activities) was evaluated in u a-stat deficient cells abstract/case report text background: t cell lymphopenia associated with genetic syndromes can be identified with low t cell receptor excision circle (trecs) up on the newborn screen for severe combined immune deficiency (scid). jacobsen syndrome (js), q terminal deletion, is a rare genetic disorder seen in / , births characterized by facial dysmorphisms, platelet abnormalities, neurologic complications, immune system abnormalities including t and b cell defects. we report an infant with js found to have low trecs on nbs and review the immune phenotypes of our cohort of patients with js. methods: a retrospective chart review of all patients with js seen by the allergy immunology service at a large tertiary referral center from / / - / / was performed in accordance with irb standards. result: the index patient had two newborn screens hours after birth and two weeks later in accordance with texas state law. the first nbs resulted normal trecs while the second nbs had low trecs. a third nbs was done per protocol and again showed low trecs. subsequently, lymphocyte subsets at months of age showed severe t-cell lymphopenia: cd cells/ dl ( %), cd cells/dl ( %), cd cells/dl, and low recent thymic emigrants (cd +cd ra+ccr +cd +) cells/dl ( %) with normal lymphocyte mitogen proliferation. a chromosomal microarray (cma) revealed a q deletion known to cause js.over the five year study period we evaluated seven patients with js referred to our center. the majority of patients ( %) presented to clinic with history of recurrent infections including recurrent pneumonia, sinusitis, otitis media, skin abscesses and warts. t-cell lymphopenia was found in of ( %), / ( %) had abnormal lymphocyte proliferation (mitogens and antigens) and met criteria for pjp prophylaxis. in addition, / ( %) had antibody deficiency requiring igg replacement therapy. of the cases reviewed, only patients were born during the period of time that texas was performing the nbs. conclusion: jacobsen syndrome can present with a spectrum of immune defects most notably t cell lymphopenia and antibody deficiency. these patients can present at birth with low trecs. this cohort analysis highlights the importance of considering chromosomal genetic syndromes with features of primary immunodeficiency in evaluating patients with low trecs. further evaluation of larger cohorts gathered from neurology or genetics clinics at multiple centers would be helpful for future study in identifying those who need close immunology care. abstract/case report text introduction: recognized as sentinels of the immune system, mast cells (mcs) and dendritic cells (dcs) are both derived from hematopoietic/ progenitor stem cells in bone marrow (bm). the crosstalk and direct contact between these cells have been well documented and play an important role in modulating immune response. we showed that presence/absence of il- directs cell fate; whether progenitor cells will be differentiated into mcs or dcs. we also report an easy method in which in vitro generation of dcs is possible without external induction of gm-csf or il- . material-methods: to produce mcs and dcs in vitro, briefly bm samples were collected from patients with idiopathic thrombocytopenic purpura., bm mononuclear cells (mncs) were seperated by ficoll gradient, and seeded in plates with imdm medium containing fbs %, pen/ strep, and a little amount of methocult, and incubated in oc, % co (day ). the treatments on the following days were as follows: day , imdm (fbs %) + scf ( ng/ml) + il- ( ng/ml); day , imdm (fbs %) + scf ( ng/ml) + il- ( ng/ml) + il- ( ng/ml). on day , groups were formed: group i: imdm (fbs %) + scf ( ng/ml) + il- ( ng/ml) + il- ( ng/ml), group ii: imdm (fbs %) + scf ( ng/ml) + il- ( ng/ml), group iii: dmem + fbs %. the cultures were evaluated every days under an inverted microscope. verification of mcs was performed by toluidin blue and tryptaseimmunoflorescence staining. macrophages were verified by cd- immunoflorescence staining. dendritic cells of different stages of abstract/case report text introduction a reduced toxicity busulfan, fludarabine regimen with alemtuzumab or anti-thymocyte globulin (gungor et al.) was efficacious in patients undergoing allogeneic hct for cgd. we report our experience with a similar approach for patients with non-cgd primary immune deficiencies needing a reduced toxicity myeloablative approach. methods we retrospectively reviewed records of consecutive patients who underwent allogeneic hct for primary immune deficiencies with a preparative regimen containing busulfan, fludarabine and alemtuzumab or anti-thymocyte globulin(atg), at three transplant centers between - . busulfan was given either every hours over days with target auc of to μmol/min (based on q hr. dosing) or twice daily over days with target auc of to μmol/min (based on q hr. dosing) or once daily over days with a target auc of - μmol/min (based on q hr. dosing). fludarabine mg/m to mg/ m was given divided over - days. serotherapy included alemtuzumab . - . mg/kg or atg . mg/kg given divided over days. gvhd prophylaxis consisted of cyclosporine and mycophenolate mofetil. results forty patients (was= , hlh= , cd l deficiency= , ipex/veoibd= , scn= , ifngr def./cid/x-scid/msn / lad= ) received busulfan, fludarabine and alemtuzumab or atg for allogeneic hct (first hct in patients and second hct in the patients with hlh). median age was . years (range, . years - . years). patients received a graft from an hla-matched related (n= ), unrelated (n= ), or single allele mismatched related or unrelated donor (n= ). all except one patient engrafted at a median of days (range, - days). one patient developed veno-occlusive disease and two patients developed diffuse alveolar hemorrhage. notably, it was the second transplant for all patients. eight patients ( %) developed grade - acute gvhd and patients ( %) developed chronic gvhd. one patient developed primary graft failure and two patients secondary graft failure. nineteen patients ( %) maintained full donor (> %) chimerism following allogeneic hct. twenty patients ( %) developed mixed chimerism, predominantly in the t-cell lineage, but t-cell donor chimerism progressively increased post-hct. at -year post-transplant, of patients ( %) with mixed chimerism had donor myeloid chimerism > % and t-cell chimerism > %. two patients underwent a second transplant for graft failure. there were deaths in the cohort. overall survival was %( of ) and event free survival was %( of ) at year. conclusion our experience suggests that a reduced toxicity busulfan, fludarabine regimen with alemtuzumab or atg as serotherapy offers a promising approach with low toxicity, durable myeloid engraftment, low incidence of grade - gvhd and excellent survival and can be considered for a variety of primary immune deficiencies where myeloablative hct is desired. abstract/case report text this is an -year-old patient who has the diagnoses of anti-nmda receptor encephalitis with associated catatonia. the patient has history of a pineal gland germinoma diagnosed in august after months of double vision. however, after several months, the patient developed difficulty in sleeping, anxiety and nightmares. the patient presented to emergency department in february with personality changes. he was diagnosed with nmda encephalitis based on the clinical findings as well as presence of elevated serum and csf anti-nmda antibodies. the patient was initially treated with high dose systemic steroids with poor response. due to the worsening of his clinical condition, he was started on plasmapheresis, but had poor response to this therapy as well. these treatments are known standard treatment options for this condition. during his hospital stay, different therapies were discussed. cyclophosphamide was one of the treatment options, but because of the side effect profile and severe toxicity, we recommended different treatment modality. we started the patient on rituximab as well as sirolimus therapy to suppress both t and b-cell responses. after receiving two doses of rituximab in addition to daily sirolimus, the patient showed improvement of his symptoms. and declining nmda receptor antibody titers. this treatment plan was chosen for autoantibody mediated encephalomyelitis due to the fact that rituximab has inhibitory effect on naive b cells, but not on the proliferation of memory b cells and sirolimus has profoundly inhibiting role on memory b cells as well as a t-cell responses. a mosaic gene variant is one which is present in some, but not all, cells within an individual. they are increasingly associated with a number of diseases, including a number of primary immunodeficiency disease (pid). here we systematically analyzed mosaic variants in known or putative immunodeficiency genes from exome sequencing data from individuals. lofreq was used to identify mosaic variants with a variant allele fraction (vaf) of . - . ( . is the vaf for a non-mosaic, heterozygous variant). we removed variants from extreme read-depth (> standard deviations), unmappable, repeat-rich, and duplicated genomic regions. the average number of detected variants per mb was . and the total number of genomic locations with variants was , . mosaic variants were underrepresented in exonic regions, suggesting that coding variants may be deleterious. more mosaic variants were detected in saliva exomes compared to peripheral blood exomes (p-value < x - ), suggesting tissue-specific mosaic variants in buccal epithelial cells and white blood cells of saliva. to understand the clinical relevance of these findings, the variants were further filtered to include only nonsynonymous variants found in fewer than % of samples, leaving a total of , variant locations. of the remaining variant locations, had a pathogenic assertion in the human gene mutation database, and were in international union of immunological societies pid genes. further variant interpretations and clinical correlations are underway. these data suggest that mosaic variants in pid genes are common, vary by location of collection, and may have clinical diagnostic relevance. abstract/case report text introduction/background: the arpc b (actin related protein / complex subunit b) gene is a protein coding gene prominently expressed in blood cells and is necessary for the assembly and maintenance of the human actin-related protein / complex (arp / ). actin polymerization plays a central role in many immune functions including proliferation and differentiation of immune cells, migration, intercellular and intracellular signaling and activation of both innate and adaptive immune responses. defects in the actin cytoskeleton affect hematopoietic cells in the bone marrow and the immune response giving rise to a distinct primary immune deficiency, which is phenotypically similar to wiskott-aldrich syndrome (was). arpc b deficiency clinically presents as a severe multisystem disease which includes platelet abnormalities, recurrent infections, failure to thrive, inflammatory changes in the intestine, eczema, cutaneous vasculitis, eosinophilia, and elevated inflammatory markers. arpc b deficiency is rare and has only recently been described in the literature. here we present a clinical case of a patient found to have a pathogenic arpc b mutation via whole exome sequencing. case report: here we describe a month old somalian boy who presented to the immunology team at months of age with hematemesis, hematochezia, melena, failure to thrive, atopic dermatitis, hypothyroidism, autoimmune thrombocytopenia and recurrent infections concerning for a primary immune deficiency. family history was notable for parental consanguinity and an older sibling with a similar presentation of hemorrhagic gastroenteritis who died in kenya around months of age due to complications of his symptoms. the initial primary immunodeficiency evaluation revealed normal inflammatory makers, normal igg and protective vaccine (prevnar and tetanus) response. iga and ige were elevated at mg/dl and . ku/l respectively. flow cytometry was remarkable for t cell lymphopenia (cd , cd , and cd ) with reduced naïve cd and cd t cells. b and nk cell count were normal. alps panel and was protein expression was unremarkable. whole exome sequencing was performed and revealed homozygotic mutation of arpc b c. t>c, ivs + t>c which was predicted to be a pathological variant. subsequently, dhr flow cytometry with fmlp showed significant increase of dhp fluorescent (mfi . when compared to control mfi . ) consistent with findings from other arpc b deficiency patients. at the time of the most recent clinic visit, the patient has remained abstract/case report text introduction: activation of the nod-like receptor family cardcontaining protein (nlrc ) leads to the formation of an inflammasome. the inflammasome, a large cytosolic multiprotein complex of innate immunity, promotes proteolytic cleavage, maturation, and secretion of pro-inflammatory cytokines, including il- and il- . a gain-of-function mutation (gof) in the gene encoding nlrc or nlrc -inflammasomopathy is characterized by hyperinflammation with persistent elevated il- , infantile enterocolitis, and early-onset macrophage activation syndrome (mas). objectives: to describe phenotypic variation among three siblings with a novel nlrc gof variant and to expand our current understanding of the clinical manifestations of the disease methods: clinical and laboratory features were studied in three siblings of a hispanic family with a novel nlrc gof variant. results: a novel variant, c. g>a (p.arg gln) on the nlrc gene, was identified in a -year-old male with recurrent febrile episodes since one year of age. his laboratory findings showed highly elevated esr, crp, il- , and fecal calprotectin. his endoscopic finding was unremarkable. the recurrent fever partially responded to canakinumab. a -year-old sister with ileocolonic crohn's disease for two years was found with the same nlrc variant and highly elevated il- . crohn's disease was well controlled after adding infliximab infusion to methotrexate therapy. a -year-old sister, who has been asymptomatic and healthy, was tested with the same positive nlrc variant and highly elevated il- . the nlrc variant is inherited from their father, who currently has a diagnosis of psoriasis vulgaris. the il- levels of the three siblings show in the figure patients with flh- develop the classic hlh phenotype early in life, with periods of remission. the pathogenesis is associated with a defect of perforin-dependent cytotoxicity. t-ctl and nk cells fail to remove abnormal cells, consequently, an uncontrolled proliferation and activation of cd + t cells and macrophages develops and generates an inflammatory cytokine storm and a solid organs infiltration. mortality of fhl- is very high without treatment; allogeneic hsct is the only curative therapy. we report the outcome of a child with fhl- , four years after his treatment with an allogeneic hsct using an hla haploidentical donor. case report: a boy born in december , previously healthy, the only child of parents without consanguinity. he has a normal family and perinatal history. when he is years old, he begins suffering from with fever, arthralgia, hepatosplenomegaly and pancytopenia. he meets the diagnostic criteria for hemophagocytic lymphohistiocytosis (hlh) and received treatment with iv methylprednisolone and entered in remission of its hlh. six months later, he restarts his clinical picture of hlh and j clin immunol receives treatment with the hlh- protocol, which leads to remission. ayear later he presents a new episode of hlh that responds to ivig, cyclosporin a and dexamethasone. the possibility of primary hlh is then suggested. when he is years old, the molecular diagnosis is made with the identification of a mutation in the stx gene, the case was classified as fhl- due to syntaxin deficiency. in october , a hsct was performed. at the age of , his clinical status is evaluated and laboratory studies show that his immunodeficiency due to syntaxin deficiency was cured. his -year-old mother, hla haploidentical was used as a donor and a protocol developed for the treatment of osteopetrosis was applied. the hsct protocol did not use a tcell depleted graft. hstc conditioning was done with melphalan days - and - , fludarabine days - to - , anti-thymocyte globulin days - and - , and cyclophosphamide day - . (figure) . clinical and demographic characteristics, including the use of a novel claims-based weighted algorithm (risk vital sign; rvs) and those initiating ivig and scig, were described. stratified analysis based on pid diagnosis codes was performed. probability of receiving available ig treatments based on baseline characteristics was evaluated by logistic regression and propensity score methods. results: selected clinical and demographic characteristics, severity measures, and previous treatments between the overall pid population (n= , ), pid patients initiating scig (n= , ), and pid patients initiating ivig (n= , ) are presented in the table. patient characteristics and previous treatments tended to be stable, although hypertension, obesity, and corticosteroid use increased during the study period. new ig users tended to be older and female, with increased depression, dyslipidemia, and hypertension than all pid patients. new scig users had more diagnoses of respiratory (e.g., asthma, copd) and inflammatory (e.g., arthritis, fibromyalgia, inflammatory bowel disease) comorbidities and less cancer than all pid patients. new scig users compared with new ivig users had increased asthma and copd, fibromyalgia, and inflammatory bowel disease and decreased cancer and peripheral vascular disease. previous corticosteroid use was higher in ig users than all pid patients. among scig users, prior pid treatments of iv antibiotics, and oral high potency antibiotics were similar to all pid patients. ivig users had higher iv antibiotic and antifungal use. rvs was initially developed to identify patients likely to have undiagnosed pid. this analysis applied rvs to patients diagnosed with pid to assess severity. rvs based on -year history in the overall pid cohort was predominantly low, with only . % of patients scoring in the medium and high ranges. rvs was increased in incident ig users, with ( . % medium/high for scig; . % medium/high for ivig). in other markers of severity, scig users had more sinusitis and ivig had more pneumonia than all pid. ig users had fewer abscesses, cellulitis, and otitis media than the full pid cohort. conclusions: this exploratory analysis showed a trend toward increased hypertension, inflammatory and respiratory comorbidity, higher rvs, and previous corticosteroid treatment in patients initiating on ig compared with all pid patients. results could be confounded based on pid diagnosis codes used and warrants further research. author disclosures: mp, cas, and zh are employees and stockholders of the takeda group of companies. jo is a consultant to takeda. jbl is an employee of rti health solutions, an organization funded by takeda to conduct this research. mer was an employee of rti health solutions at the time this research was conducted.presenting author: colin anderson-smits submission topic: immunoglobulin replacement therapy patients in a larger cohort group and to determine if there are patterns of disease not previously reported. methods: we performed a retrospective chart review of patients with lof stat (n= ) followed at the national institutes of health. we specifically looked at tortuosity, aneurysms, and dilation of both coronary and cerebral arteries. epidemiologic information, stat mutation, co-morbidities, and laboratory information were reviewed along with imaging studies, specifically brain mri, brain mra, heart mri, and coronary ct. results: most recently, patients with hies are found to have vascular abnormalities including tortuosity, dilatation, narrowing, and aneurysms of middle sized, cerebral, and coronary arteries. in an effort to determine the extent of vascular involvement in addition to miscellaneous organ involvement, we are reviewing a cohort of patients with hies who were evaluated at the nih. of these patients, are women and are men. of these patients, two have passed away due to vascular events leading to their deaths. there are four patients under the age of , patients between the ages of and , patients between the ages of and , and three patients above the age of (age range - , mean age . ).of the patients, five of these patients were found to have abnormal brain mri/mra at an approximate rate of . %. two of these patients were found to have at least one cranial aneurysm, two of these patients were found to have a level of narrowing or stenosis, and one patient was found to have dilatation.in terms of coronary abnormalities, of the ( . %) patients were noted to have at least one coronary abnormality including dilatation, aneurysm, or tortuosity on heart mri or coronary ct. eight patients ( . %) were found to have dilatation of which four patients were female and patients were male. of the patients, ten patients ( . %) were found to have at least one aneurysm. there were patients ( %) that were found to have at least mild tortuosity. conclusion: vascular abnormalities in our lof stat patients occurred at an exceedingly high rate-cerebral and coronary artery, . % and . % respectively. due to this, patient's with lof stat should be considered for screening with brain and heart imaging. currently, there are no guidelines which outline the appropriate timeline for screening in these patients however following these patients over time will allow us to determine the most appropriate interval for imaging follow up. abstract/case report text year old woman with personal history of severe and recurrent upper and lower respiratory infections, chronic pulmonary disease with bilateral bronchiectasis and several micro nodules, chronic diarrhea without diagnosis (colonoscopy with mild colitis, without cmv. no bacterias no parasits were found in stools), mild osteopenia, focal lesion in right hepatic lobe, atopic dermatitis and anemia. she was followed up in other center and in she was diagnosed with common variable immunodeficiency (cvid) and started treatment with intravenous immunoglobulin (ivig), but with low adherence to it. she did not have referred history of lymphoproliferation nor significant viral infections. she had a daughter with spherocytosis who required esplenectomy and also had bronchiectasis and cvid diagnosis, she deceased at years old due to pulmonary infection. one years old son has anemia. her other daughter and son are healthy. objective: to describe unique management of recurrent viral respiratory infections in a qds patient. case: -year-old male with a history of qds complicated by truncus arteriosus, vsd, hypoparathyroidism, asthma, and autism was followed for a history of "frequent pneumonias." he had daily rhinitis and a history of frequent otitis media which resolved after tympanostomy tube placement at age y. however, over the next two years, he had admissions with various respiratory viral infections resulting in respiratory distress and prolonged oxygen need. viruses detected during these separate admissions varied and included parainfluenza , metapneumovirus, rsv, b and coronavirus nl , coronavirus hku , and rhino/enterovirus ( times total). he was previously on a prophylactic course of antibiotics which made no difference in his symptoms. laboratory evaluation showed protective adaptive immunity with normal immunoglobulin numbers for age (igg mg/dl), along with normal b and t cell numbers. he had protective pneumococcal titers and mounted a normal mitogen response. while his nk cell numbers were normal, his nk t cells were low. his tlr functioning also appeared normal. in order to decrease his overall illness burden and to keep him out of the hospital, ivig infusions (~ mg/kg) were initiated monthly. shortly after initiation of treatment, his nasal purulence and drainage resolved and his family noted that he became more active and playful. treatment was continued for months, during which time he had only one episode of influenza infection needing inpatient management. he has been off of ivig for months without recurrence of his viral infections. conclusion: in this patient, severe recurrent viral respiratory infections despite apparently normal adaptive and cellular immunity presents a unique management dilemma. this was not an issue of recurrent bacterial infections as prophylaxis did not make a difference in the frequency of his infections. his nk t cells were low, which could have contributed to his frequent viral infections as nkt cells are known to play a role in viral immunity. the successful use of ivig treatment in his case points to a different use for ivig, namely for the anti-respiratory virus antibodies which are presumably contained within the formulation.given this finding, it may be prudent to consider ivig in management of qds patients, even with normal immune evaluation, in order to decrease risks of complications associated with severe and recurrent viral respiratory infections. abstract/case report text introduction: smad is a critical downstream signaling molecule for transforming growth factor-β (tgf-β) and bone morphogenic protein (nmp ). initially, smad , also known as dpc deleted in pancreatic cancer locus , was described as a tumor suppressor gene, and somatic deletion of the smad is seen in % of pancreatic carcinomas. subsequently, a germline point mutation in the smad gene (p. i v) was reported to cause myhre syndrome (mim# ). myhre syndrome is an autosomal dominant disease characterized by cognitive impairment, hearing loss, and musculoskeletal anomalies. the immunological phenotype of these patients has not been previously described, despite the critical role of tgf-β in regulating t cell response and the prevention of excessive inflammation. case report: a -year-old boy with myhre syndrome was referred to immunology clinic for evaluation of recurrent ear infections. he developed acute otitis media infections as an infant and had tympanostomy tubes placed at one year of life. he also had a recurrent sinus infections. at two years of age, he was diagnosed with autism and sensory neuronal hearing loss. brain mri showed a mildly hypoplastic pituitary gland, and a thickened corpus callosum with decreased myelination. given these findings, whole-genome sequencing was performed, which revealed a heterozygous de novo mutation in smad (p.i v / c. a>g) consistent with the diagnosis of myhre syndrome. through age , he was in the th percentile for height until he was started on growth hormone, which he responded to robustly. he is now he is in the th percentile for height. he had adenoidectomy due to sleep-disordered breathing at the age of years. he is maintained on montelukast and inhaled corticosteroids for treatment of rhinitis and mild persistent asthma. he is on atenolol for the treatment of primary hypertension. on physical exam, he has facial dysmorphisms, thickened skin, and contraction of the fingers consistent with myhre syndrome. immunologic elevation showed significant hypogammaglobulinemia (igg mg/dl), low iga ( mg/dl) and normal igm mg/dl. igg subclasses showed low igg and igg at mg/dl and mg/dl respectively. although he was fully vaccinated, his tetanus antibody was low at . iu/ml. however, this improved after repeat vaccination to . iu/ml. total t and b lymphocyte counts were normal; however, his memory cd and cd t cells were low for age at . % and . %, respectively. additionally, his switched memory b cell count was low at . %. conclusion: smad gain of function (myhre syndrome) can lead to impaired memory t and b cell formation with significant hypogammaglobulinemia and low iga. although the patient was able to respond to protein vaccination (tetanus), it is not clear if he will be able to maintain a long-term response. in a previous study, a similar gain of key: cord- -oecpqf j authors: nan title: aspho abstracts date: - - journal: pediatr blood cancer doi: . /pbc. sha: doc_id: cord_uid: oecpqf j nan myelodysplastic syndrome (mds) and frequently arise in the context of inherited bone marrow failure (bmf) syndromes, such as shwachman diamond syndrome (sds). monosomy /del( q) is associated with high grade mds and propensity to progress to acute myelogenous leukemia, a major cause of morbidity and mortality for patients with inherited bmf. development of non-transplant strategies to treat bone marrow failure without simultaneously stimulating outgrowth of malignant clones remains a major challenge. objectives: the aim of this study is to investigate the molecular consequences of del( q) in the context of bmf with the goal of developing more effective treatments. design/method: to study the biological and molecular consequences of monosomy/del( q) in bmf, induced pluripotent stem cells were generated from sds patients (sds-ipsc) . a deletion of the mds-associated region of the long arm of chromosome was then introduced using a previously published modified cre-lox approach. results: the sds ipsc phenocopied bone marrow failure with slow proliferation and impaired hematopoietic differentiation. we next explored whether deletion of q conferred a relative fitness advantage within the context of bone marrow failure. proliferation of the sds-del( q) ipscs was reduced below that of both the isogenic sds ipscs and normal controls without an increase in cell death. sds-del( q) demonstrated reduced hematopoietic differentiation compared with isogenic sds cells. these data demonstrate that deletion of q fails to confer a relative growth advantage relative to isogenic sds ipscs and results in further impairment of hematopoiesis. to gain insight into the mechanisms of del q-associated clonal evolution in sds, we performed rna sequencing (rnaseq) of sds+/-del( q) ipsc. expression of tgf pathways and their downstream targets were reduced in sds-del( q) ipscs compared to isogenic sds ipsc. single cell rnaseq analysis of primary sds bone marrow cells confirmed that the tgf pathway is hyperactivated in sds. western blot analysis showed increased phospho-smad levels in sds ipscs compared to sds-del( q) and normal controls, while total levels of smad were unchanged. pharmacological targeting of tfg with small molecule inhibitors resulted in selective improvement of sds hematopoietic colony formation and myeloid differentiation without stimulating outgrowth of the isogenic sds-del( q) cells or normal controls. these results demonstrate that del( q) reverses the tgf pathway hyperactivation of sds. furthermore, inhibition of tgf selectively rescues hematopoiesis in sds but not in isogenic del q cells, suggesting a potential strategy to treat bone marrow failure without stimulating del q clonal outgrowth. background: standard therapy of medulloblastoma consists of treatment with alkylating agents and radiation after surgical resection. although a statistically significant increase in survival is reported with this regimen, / rd recur and become resistant this class of agents ultimately leading to mortality. large numbers of somatic mutations were observed in recurrent medulloblastoma (rm) after alkylating agent and radiation treatment. high mutation rates in tumors can have twofold effect; ) a large number of non-synonymous mutations that have no role as drivers can still cause functional tumor antigens increasing the neoantigen burden and immunogenicity. moreover, ) such tumors can gain mutations in canonical or non-canonical dna repair pathways leading to a gain in the number of mutations as seen in case of glioblastoma, this can lead to even higher accelerated mutational rate. evidences suggest that high mutational load can cause higher neoantigen burden thereby making the tumor more susceptible to immune checkpoint inhibition. we propose that post therapy recurrent medulloblastoma gain mutational signature and immunophenotype of malignancies demonstrating clinical response to immune checkpoint therapy. objectives: ) rm has molecular signatures identical to tumors with high immunogenicity and clinical response to immune check point inhibition. ) rm has the immune inflammatory phenotype; harboring high percentage of tumor infiltrating lymphocytes (tils), macrophages and monocytes. design/method: to test our hypothesis, we downloaded the raw bam files of previously published data from international cancer genome consortium (icgc) . this set of about matched primaries and recurrent medulloblastoma cases forms our discovery cohort. we have called somatic variants using the gatk pipeline by the broad institute. to validate our key findings, we have procured human medulloblastoma specimens and are conducting whole exome sequencing. the primary assays utilized to assess immunogenicity are immunohistochemical (ihc) staining of formalin fixed and embedded recurrent medulloblastoma tissue to identify tils, tumor associated macrophages and other markers. mg/m had dlts of dyspnea (grade )/hypoxia (grade ) but no dlts were observed in any other cohort. adverse events were generally mild to moderate, consistent with the safety profile observed in adults. across the desc cohorts, plasma concentrations were dose-proportional and steady state concentrations were lower on day vs. day . mean systemic exposure in the mg/m cohort was ∼ -fold greater compared with the adult rp d of mg bid. a pk:pd relationship between tazemetostat exposure and h k me levels in peripheral blood monocytes and granulocytes was observed in the desc phase. consistent and significant post-dose reductions in h k me occurred at doses ≥ mg/m . further analysis of twelve patients treated at the rp d confirmed that h k me inhibition was maximally inhibited. doses - mg/m showed confirmed objective responses (cr/pr) per recist/rano in patients with es (n = ), chordoma (n = ), and atrt (n = ). background: previous studies established that the platelet/ fibrin(ogen) axis promotes metastatic potential by impeding the clearance of newly formed micrometastases by natural killer (nk) cells. however, multiple important questions remain, including the potential of fibrin(ogen) to promote metastasis through interactions with cells other than platelets (e.g., inflammatory cells), and the fundamental question of whether fibrin polymerization is required for metastasis. objectives: determine the role of fibrin polymerization and fibrin(ogen) engagement of integrins iib and m in metastasis. design/method: we performed experimental and spontaneous metastasis assays in immunocompetent mice carrying specific fibrinogen structure/function alterations. results: expression of a mutant fibrinogen lacking the binding motif for the leukocyte integrin m (fib - a) significantly decreased metastatic potential relative to wildtype fibrinogen, suggesting a role for fibrin(ogen)inflammatory cell interactions mediated by m in metastasis. to directly determine the importance of thrombinmediated fibrin polymerization in metastasis, we analyzed metastatic potential in fibaek mice, which carry a form of fibrinogen essentially "locked" in the soluble state due to a mutation in the a chain thrombin cleavage site. metastatic potential in fibaek mice was diminished relative to control mice, speaking to the importance of thrombin-mediated fibrin polymerization in the metastatic process. however, the fibaek mice retained significant metastatic potential relative to complete fibrinogen deficiency, indicating that fibrinogen monomer retains significant prometastatic properties. in order to better define the role of fibrin(ogen)-platelet interactions in metastasis, we compared metastatic potential in control and fib Δ mice, carrying a form of fibrinogen lacking the chain binding motif for the platelet integrin iib . surprisingly, this mutation had no impact on metastatic potential. together, these studies suggest fibrinogen plays a multifaceted role in metastasis. fibrin(ogen)-leukocyte interactions mediated by m appear to have a role in metastasis. previous studies showed that macrophages promote the metastatic potential of circulating tumor cells, which may represent at least one important m expressing cell type whose prometastatic behavior is influenced by fibrin(ogen) interactions. these studies show that thrombin-mediated fibrin polymerization promotes metastasis, but soluble fibrinogen retains some significant prometastatic capacity. surprisingly, loss of the fibrinogen chain iib binding motif had no impact on metastasis. given the established importance of platelets in metastasis, these findings suggest that fibrin (ogen) is capable of platelet stabilization through mechanism(s) independent of this iib binding motif. platelets may bind polymerized fibrin at other sites, and/or fibrin interactions with other matrix proteins capable of binding iib are sufficient to support platelet functions required for metastasis. the role of platelets in hemostasis and thrombosis is well defined, but it is becoming increasingly evident that platelets also assist in host defense and inflammation. platelets participate in the innate immune system through direct antimicrobial activity and interactions with effector cells (chapman , garlanda , kapur ). in the adaptive immune system, platelets recruit and costimulate t-cells, and promote b-cell differentiation and antibody class switching (kapur , morrell ). the question remains: which mechanisms influence platelet immune function and are they developmentally regulated? preliminary studies in the palis lab have revealed significant dif-ferences in embryonic versus adult platelet gene expression, including regulators of immune and inflammatory responses such as beta -microglobulin (b m) and major histocompatibility complex class i (mhc ). mhc is expressed on all cell surfaces except red blood cells and its molecular chaperone b m is a marker of inflammation highly expressed in platelet alpha granules (zufferey ). preliminary data from the morrell lab reveals a mass release of b m during platelet activation, which drives monocyte differentiation to an inflammatory phenotype through tgfb receptor signaling. we therefore sought to determine whether developmental changes in platelet b m expression mediate differences in platelet-mediated monocyte activation. with trilineage hematopoiesis with a predominance of early myeloid precursors, with full maturation. microarray, elane and sbds sequencing and deletion/duplication analyses were negative. immunologic evaluation was significant for agammaglobulinemia and an absence of memory (cd +cd +) b cells. a gene primary immunodeficiency panel revealed two variants of unknown significance-c. g>a and c. g>t in dnmt b; one previously reported in association with icf . parental testing demonstrated parental heterozygosity. centromeric instability was confirmed in mitogen stimulated lymphocytes showing characteristic, multibranched chromosomes containing at least arms of chromosome and joined near the centromere. decondensation of the qh and qh regions and triradial configuration of chromosome was noted, and a diagnosis of icf syndrome was made. the patient was started on monthly intravenous immunoglobulin (ivig). prophylaxis for pneumocystis jiroveci pneumonia and respiratory syncytial virus was initiated. a / matched sibling hsct is being planned. demonstrated the diagnosis of high grade osteosarcoma. the patient was started on multi-agent chemotherapy with planned a whole femur prosthesis at time of local control. cases of osteosarcoma have been described in the literature in patients with nf (median age; years, range - years) with slightly male predominance ( cases). the femur was the most common site of involvement ( cases). four patients died of metastatic disease despite surgery and multi-agent chemotherapy. conclusion: nf represents a major risk factor for development of malignancy and uncommonly osteosarcoma in adolescents and adults. we report a rare case of an extensive involvement of osteosarcoma of the left femur in a child with known diagnosis nf . this presentation should alert the pediatric oncologists to monitor for bone tumors in patients with nf by physical exam and detailed medical history. hasbro children's hospital, providence, rhode island, united states background: dysautonomia is a paraneoplastic syndrome most commonly described in adult malignancies. despite current therapies aimed at symptoms management, it is often debilitating. we present a case of a -year-old girl who initially presented with autonomic dysfunction and was subsequently found to have hodgkin lymphoma. objectives: describe hodgkin lymphoma presenting with dysautonomia and discuss symptom management with rituximab design/method: case report a year-old-girl presented with severe symptoms of orthostatic hypotension necessitating prone positioning to prevent syncopal episodes. additionally, she reported anhidrosis, xerostomia, urinary retention, and constipation. she had unmanageable peripheral neuropathic pain despite multiple analgesia medications. initially, it was suspected that her symptoms were caused by an atypical presentation of guillain-barre syndrome. she was treated with intravenous immunoglobulin g, without response. due to a suspicion of a paraneoplastic syndrome a positron emission test/cat scan (pet/ct) was performed and revealed widespread fdg-avid nodal and splenic disease. pathology from a thoracoscopic biopsy of a mediastinal lymph node demonstrated classical hodgkin lymphoma. she was classified as stage ivb. a paraneoplastic panel obtained during the first cycle of chemotherapy revealed elevated anti-amphiphysin antibodies and glutamic acid decarboxylase (gad) antibodies. therapy was initiated with abe-pc (doxorubicin, bleomycin, etoposide, prednisone, cyclophosphamide) ; vincristine was held given her significant neuropathy. due to persistence of autonomic symptoms following her first cycle and presence of antiamphiphysin and gad antibodies, rituximab was incorporated into her treatment. following two cycles abe-pc, she had a rapid early response by fdg-pet/ct. she completed an additional three cycles of abd-pc. end of therapy imaging demonstrated complete response with a single persistent mildly fdg-pet avid lymph node (deauville ) and her antibodies were negative. she continues treatment of maintenance rituximab with significant improvement, but not resolution, of her orthostatic hypotension. at this time, the patient can ambulate with assistance. constipation and urinary retention have fully resolved and, her peripheral neuropathy, xerostomia, anhidrosis have improved. conclusion: this is rare case of a pediatric hodgkin lymphoma patient developing dysautonomia associated with antiamphiphysin and glutamic acid decarboxylase antibodies and subsequently managed with chemotherapy and rituximab. clinicians should be suspicious of a paraneoplastic syndrome when a neurologic disorder fails to improve with standard treatment. results: labs obtained at an outside hospital one month prior to presentation showed absolute neutrophil count (anc) and hemoglobin . g/dl. she presented to our institution with days of fever, hepatomegaly cm below costal margin, a white plaque on her tongue, and circumferential perianal ulceration. labs were significant for anc and hemoglobin . g/dl. anti-granulocyte antibody testing was positive. bone marrow biopsy showed arrest of neutrophil maturation. after initiation of filgrastim ( . mcg/kg/day), her anc increased to > and repeat bone marrow biopsy demonstrated left shifted myelopoiesis. biopsy of her oral lesion demonstrated invasive actinomyces prompting a prolonged course of antibiotics. biopsies of her oral and anal lesions were reported as myeloid sarcoma without mll rearrangement. chemotherapy was not initiated due to complete resolution of both lesions within weeks of initiating filgrastim and appropriate antibiotic coverage. she has not developed any further lesions concerning for malignancy. testing for common genes associated with severe congenital neutropenia and autoimmune lymphoproliferative syndrome was negative. her immunoglobulin levels and the measurement of age-appropriate vaccine responses were normal. after her lymphocyte subpopulation analysis indicated a selective deficiency in cd positive t-lymphocytes (absolute cd cell count ), the severe combined immunodeficiency panel from genedx showed compound heterozygous mutations in results: a male infant was born with a large thigh mass. the child was clinically well aside from restricted movement of affected leg. mri showed mass expanding into pelvis without other lesions. an interventional-radiology guided core biopsy of the mass was reported as high-grade spindle cell sarcoma without etv rearrangement. surgery was deferred because of concern that it would result in excessive morbidity. the mass was treated with vincristine and dactinomycin per infantile fibrosarcoma protocols. after months of therapy, no significant change in size of the mass was noted on physical exam or imaging. repeat biopsy was obtained to confirm diagnosis and allow for expanded tumor testing. this biopsy showed triphasic distribution of adipose, fibrous and mesenchymal tissue consistent with fhi with rare sarcomatous foci. additional chemotherapy was deferred and the child was followed clinically. his tumor has remained approximately the same size and still unresectable. next generation sequencing of tumor utilizing panel based technology revealed braf-erc fusion consistent with braf activating mutation. this mutation was confirmed by fluorescent in situ hybridization (fish) probe for braf. braf and mek inhibitors have been pursued as treatments to decrease size of tumor and allow for resection. conclusion: braf mutations have been characterized in a variety of malignancies. inhibition of braf and downstream signaling components has produced promising results in a variety of patients. this is the first case report of a braf mutation in a fhi. although management of fhi is typically surgical, this does suggest a potential therapeutic target and may allow for improved surgical outcomes especially in cases where up-front surgery would result in unacceptable morbidity. genetic sequencing of fhi and other rare tumors is an important tool and has the potential to identify mutations amenable to targeted therapies. background: icf is a rare autosomal recessive disorder characterized by hypo-or agammaglobulinemia and often opportunistic infections suggesting t-cell dysfunction. it is further categorized into subtypes - based on mutations in dna methylation. mutations in the helicase-lymphoid specific (hells) gene, which is required for t-cell proliferation and participates in de novo dna methylation, are characteristic of icf type (icf ). of approximately reported cases of icf, less than percent are characterized as icf . while malignancy has been reported in icf (angiosarcoma, acute lymphoblastic leukemia), and icf (hodgkin lymphoma), here we describe the diagnosis and management of an icf patient with neuroblastoma and neutropenia, which has not been previously described. objectives: describe a novel phenotype and mutation of icf and its management to further expand our understanding of this disease. results: a month ex- week premature male with bronchopulmonary disease and failure to thrive presented with acute respiratory failure in the setting of recent viral bronchiolitis with associated chronic diarrhea. he was subsequently diagnosed with multiple infections including pjp pneumonia, norovirus, parainfluenza, rhinovirus, and pseudemonal cellulitis. he presented with profound neutropenia and agammaglobulinemia with presence of b and t cells on lymphocyte phenotyping. ct revealed a paraspinal mass that was mibg-avid on further study, strongly suggesting neuroblastoma. bone marrow was normocellular and negative for malignancy, however revealed marked granulocytic hypoplasia and maturation arrest concerning for severe congenital or, less likely, immune-mediated neutropenia. metastatic workup was negative. whole exome sequencing revealed a homozygous variant of unknown significance (c. t>c) in the hells gene, portending a working diagnosis of icf syndrome. immunoglobulin supplementation, pentamidine prophylaxis, and g-csf were initiated. he was able discontinue g-csf after months of treatment. his neuroblastoma, initially categorized as l , met criteria for observation. however, followup mri revealed interval growth nearing the spinal canal. he underwent tumor resection, confirming mycn non-amplified, favorable histology neuroblastoma. after infectious prophylaxis and immunologic support were initiated, he incurred two other hospitalizations, the first for g-tube cellulitis and the second for parainfluenza respiratory illness. he now has stable neutrophil counts off g-csf and remains in remission from neuroblastoma. current plan is to proceed with bone marrow transplantation for immunodeficiency. conclusion: icf has not previously been described with neutropenia or neuroblastoma. this report not only describes a novel mutation and phenotype of icf and the management thereof, but also reveals the potential curative role of bone marrow transplantation in such disease. staten island university hospital -northwell health, staten island, new york, united states background: desmoid tumors are rare tumors that arise from highly differentiated fibroblasts. they occur in isolation or as part of the disease spectrum of familial adenomatous polyposis (fap) . fap mutations between codons - typically correlate with increased extraintestinal disease such as desmoid tumors and upper gastrointestinal polyps. we describe a patient with a large intra-abdominal desmoid tumor who is heterozygous for a c. c>t (p.arg cys) apc gene mutation. we are not aware of any other patients reported with this germline apc mutation presenting with a desmoid tumor. objectives: to discuss a novel apc mutation and the presentation of a rare case. design/method: review of clinical presentation, genetic analysis and management of a rare tumor. a -year-old female with no significant medical history presented with abdominal asymmetry and intermittent pain. she reported urinary urgency, shortness of breath, early satiety, decreased appetite and a -pound weight loss over the course of months. ct scan of the abdomen demonstrated a × cm abdominal tumor abutting the local organs but no presence of bowel obstruction. a biopsy revealed a spindle cell neoplasm favoring fibromatosis. there was no known family history of fap, colon cancer, or desmoid tumors. apc gene mutation analysis demonstrated a c. c>t (p.arg cys) heterozygous gene variant. due to size and location of the tumor, it was initially deemed unresectable. the patient was started on a course of monthly liposomal doxorubicin. she tolerated the initial cycles well and interval ct after cycles of chemotherapy revealed a % decrease in tumor volume. variability exists in phenotypic presentation with regards to the location of the afp mutation locus. while fap mutations associated with desmoid tumors typically have changes in the - codon region, our patient presented with a heterozygous mutation resulting in a missense mutation at codon . due to the change in polarity and size, the mutation is not considered to be of conservative nature. we are only aware of one other report of this mutation, which occurred in an individual with a personal and family history of colon cancer. we are not aware of any patients with desmoid tumors who also have this germline apc gene mutation. our case report highlights an apc gene mutation that is not well-described; we are not aware of any other cases of this mutation reported in patients with desmoid tumors. future evaluation and tracking of this mutation may lead to the determination of further clinical significance. background: over time, advanced care planning for location of death has been associated with increased deaths at home rather than in the hospital. in some cases, however, complex management and symptom control can prevent families from achieving their goal of keeping their child out of the hospital and at home at the end of life. ascites is a sequelae of many conditions including malignancy that might lead to significant morbidity. increasingly, interventional procedures are being utilized. peritoneovenous "denver" shunts are placed internally with one end in the peritoneal space and the other buried within a major vessel such as the svc. a one-way valve and pump buried under the skin allows the patient to pump fluid from the peritoneal to the vascular space. the shunt is used frequently in adults, but has not seen much use in pediatric oncology patients. objectives: to describe a case of a terminally ill patient with refractory wilms tumor with ivc involvement who received symptomatic relief with denver shunt placement. results: an -year-old female was diagnosed with relapsed, refractory, metastatic wilms tumor with pulmonary and hepatic involvement, with tumor extension to the hepatic veins and ivc. multiple chemotherapeutic regimens and palliative radiation to the ivc were administered, but her disease continued to progress, leading to pressure on the portal vein and portal hypertension. the resulting ascites was causing the patient significant pain and was difficult to manage. the patient's code status was changed to dnr/dni after discussion with her mother, who identified a desire to have the child die at home as comfortably as possible. a peritoneovenous shunt was placed in order to control the patient's pain and avoid frequent medical procedures and therapies. despite initial anxiety, the patient was able to utilize the pump and achieve significant improvement in her ascites and pain. she was able to spend the remaining six weeks of her life at home. ascites is a common phenomenon of end stage disease. peritoneovenous shunts are a treatment modality that may be considered to allow for pain control at the end of life for pediatric oncology patients with ascites. the procedure is relatively low risk, allows for self-control of the pump to maintain comfort, and is easy enough to use by the patient or family. background: extraneural metastases (enm) from pediatric glioblastoma multiforme (gbm) are rare, with an estimated frequency of . %. etiologic factors include multiple neurosurgical procedures and sarcomatous dedifferentiation. their occurrence can seriously affect the patient's quality of life and survival. while enms have been well documented in adults, pediatric cases have not been previously summarized. a year old male with a cerebral gbm developed extension of disease outside of the neuraxis approximately months post initial presentation and at the time of disease progression. metastases included exracranial temporal lesions, cervical and mediastinal lymph nodes and s of s bilateral lung nodules. a large pleural-based soft tissue metastatic focus was identified on imaging when the patient presented with respiratory distress secondary to a right tension pneumothorax, which was recognized and managed promptly. we summarize the main reported cases in literature to better define risk factors for and evaluate the proposed mechanisms underlying these systemic metastases. design/method: we performed a literature review on the pubmed database using the terms gbm and enm. patients under years of age who met the weiss criteria for the diagnosis of enm from primary cns tumors were included. results: our patient fulfilled two of the three weiss criteria with confirmed gbm at the primary site with all enm in the temporal soft tissue and cervical lymph nodes displaying histopathologic features similar to the primary cns tumor. the intrathoracic adenopathy and lung nodules detected upon chest imaging during workup for respiratory distress were assumed to represent additional metastatic foci. our literature review identified pediatric patients with enm from gbm with a median age of years (range . - years) and a slight female predominance ( % females vs. % males). the most common sites of metastases reported were pleura/lungs, bones, lymph nodes and liver. in of patients, metastases were associated with csf shunting. conclusion: pediatric oncologists should have an increased index of suspicion when caring for patients with gbm, particularly those who have undergone shunting procedures and present with systemic symptoms including bony pain, respiratory changes, transaminitis or cytopenias which should prompt timely investigation for enm. although enm of cns tumors carry very poor prognosis, their diagnosis has potential therapeutic importance because treatment of metastatic lesions may alleviate symptoms and improve the quality of life. additional studies may be warranted to evaluate the incidence of enm that can provide valuable insight into the pathogenesis and biology of high-grade gliomas. nicklaus children's hospital, miami, florida, united states background: sinusoidal obstruction syndrome (sos) has been reported in patients undergoing intensive chemotherapy and as a complication post-hematopoietic stem cell transplan-tation. sos may be complicated by portal hypertension, hepatorenal disease or multi-organ failure. however, despite treatment, there may be further potential complications that can be anticipated in patients with history sos. we report two patients with history of sos presented with post-procedural bleeding after gastric tube placement. we believe that their presentations may be associated to their previous diagnosis of sos. design/method: pubmed search was done with search for terminology including "sinusoidal obstruction syndrome" "defibrotide", and "bleeding". papers relevant to our cases were selected for literature review. results: case : a year-old female with history of desmosplastic medulloblastoma status-post resection and intensive chemotherapy was diagnosed with sos one month after her second part of planned tandem transplant. she was managed with paracentesis and defibrotide. due to malnourishment, patient had a gastric tube placement months after she completed therapy and had an episode of upper gastrointestinal bleeding postoperatively from the g tube site. case : similarly, a year-old male diagnosed with anaplastic medulloblastoma status post resection and adjuvant multiagent chemotherapy. his treatment course was complicated with sos after the second cycle of induction chemotherapy which responded to -day course of defibrotide. likewise, the patient had a major bleeding event from the g-tube site approximately two months after sos diagnosis. defibrotide was discontinued in both cases before g-tube placement. both patients had no previous history of bleeding disorders or relevant family history. in addition, comprehensive laboratory evaluations were within normal limits before both procedures. in sos, there is blockage of fluid out of the liver that leads to congestion, ascites, ischemia of the liver, and post-sinusoidal portal hypertension. two related causes of sos should be considered as an explanation for g-tube bleeding. similar patients should have close monitoring postoperatively or if possible surgical intervention should be delayed until the sos process has been evolved. nicklaus children's hospital, miami, florida, united states background: the development of treatment related acute myeloid leukemia (t-aml) and myelodysplastic syndromes (t-mds) is a potential complication after cytotoxic chemotherapy or radiation therapy. the incidence of development of t-aml/t-mds varies from - % depending on the treatment regimen used. cutaneous myeloid sarcoma (ms) is a common presentation of extramedullary leukemia and usually occurs in the setting of aml. we report a rare case of cutaneous ms in an adolescent female after successful treatment for ovarian yolk sac tumor (yst) stage i with bep (bleomycin, etoposide and cisplatin) therapy. the ms was managed only with biopsy and close observation. design/method: a pubmed search was conducted for queries including t-aml/t-mds, cytotoxic agents, cutaneous myeloid sarcoma, regression. relevant papers were selected for literature review. a year-old female was diagnosed with a left ovarian yolk sac tumor, for which she underwent left salpingooophorectomy and successfully completed cycles of bep over months. during routine follow-up months after initiation of treatment for ovarian yst, she was noted to have a small, non-tender, indurated nodule on the left side of her upper back approximately cm in diameter. punch biopsy of the skin nodule was performed and pathology was positive for cutaneous myeloid sarcoma. at the time of next follow-up less than one month later, the skin lesion had resolved. two subsequent bone marrow aspirates were performed one month apart and were negative for leukemic involvement or mds. examinations and work-up including whole body pet with ct scan were negative for evidence of disease. although cutaneous ms can be regarded as the herald of systemic myeloid disease rather than a localized process, our patient was monitored periodically with physical exam and laboratory evaluations. she remains free of disease more than four years after the presentation of cutaneous ms without any further treatment. spontaneous regression ms has been previously reported. the authors would like to stress that a conservative approach with close observation could be an option in cutaneous ms even with history of chemotherapy exposure. nesreen ali, iman sidhom, sonia soliman, sherine salem national cancer institute, cairouniversity, egypt children cancer hospital egypt, egypt background: acute leukemia is the commonest malignancy in childhood. the coincidental occurrence of leukemia with hemophilia is extremely rare. hemophilia is a congenital rare x linked bleeding disorder. the main complication of the two diseases is bleeding diathesis which may be lifethreatening due to many factors, deficiency of coagulation factors in hemophilic patients, thrombocytopenia from disease and chemotherapy in leukemic patients, certain cytotoxic drugs such as asparaginase which may result in coagulation disorders and infection which may lead to disseminated intravascular coagulation. objectives: reporting such a case is imperative to set up treatment guidelines for prevention of bleeding and to optimize the therapeutic approach for these patients. design/method: seventeen years old boy, presented to children cancer hospital egypt in june with pallor and multiple ecchymoses.he was diagnosed with precursor b acute lymphoblastic leukemia, cerebrospinal fluid (csf) was free, the chromosomal analysis revealed hypodiploidy , xy. he had moderate type of hemophilia a since birth, factor viii level was . % at time of diagnosis, coagulation profile revealed prolonged partial thromboplastin time (normal - ), factor viii was low %, prothrombin concentration and prothrombin time were normal % and seconds, virology screening for hepatitis b core igg/igm, hbs ag, hiv and hc igg /igm were negative.the patient started induction total xv sjcrh protocol, factor viii unit/kg was given at presentation before doing bone marrow aspiration(bma), csf and as a prophylactic before intramuscular asparaginase injection, intrathecal and bma. it was given immediately within hours before the procedures and platelets transfusion was given regularly to maintain platelets count about , . the minimal residual disease by flow cytometry was . % and . % at d and d induction. results: our patient received his induction and reintensification chemotherapy without any major bleeding event which reveals the success of our guidelines for the prevention of bleeding. he developed very early relapse at w maintenance by the same clone. he received salvage chemotherapy but didn't achieve remission and died out of disease and resistant clone. the development of leukemia on top of hemophilia is a major problem. bleeding complication during chemotherapy can be prevented by regular prophylactic factor viii and platelets concentrate transfusion with good supportive care. life threating bleeding complication may be correlated with the severity of hemophilia. we need to collect data about the biology of leukemic cells, complications, and cause of death to optimize care for these patients. background: mucoepidermoid carcinoma (mec) is a rare malignancy that arises from exocrine glands in the upper aerodigestive tract and tracheobronchial tree. conventionally, mec diagnosis is based on histology, with prognosis based on the extent of resection and detection of metastases. mec is characterized by a translocation of chromosomes q and p resulting in a fusion between the mect and maml genes, that occurs in - % of cases. this fusion transcript has been recognized to have a favorable impact on disease features and prognosis of mec. however, recent studies indicate that high grade mec can have mect -maml fusion positivity and multiple other genomic imbalances that have not been studied in much detail. owing to the rarity of mec tumors, more definitive data related to the clinical and prognostic significance of these molecular markers are limited. objectives: . identify the presence or absence of mect -maml fusion in the tissue of our patient. . analyze the incidence of the fusion in mec cases in children and young adults retrieved from the iowa cancer registry. . determine if fusion status correlates with clinical, pathological and outcome data in our cohort. design/method: we describe the case of a year-old caucasian male who presented with recurrent pneumonia, persistent cough and radiographic evidence of right lobar collapse. bronchoscopy revealed an endobronchial lesion and the patient underwent right upper lobe sleeve resection. pathology report was consistent with low grade muco-epidermoid carcinoma. we retrieved archived formalin-fixed paraffinembedded (ffpe) specimens of pediatric and young adult mec cases (ages - ) reported in iowa from - using the iowa cancer registry. testing for the mect -maml fusion in the index case and ffpe specimens will be done using a custom-designed laboratory validated next generation sequencing (ngs) assay with the ability to detect novel fusion partners. clinical, pathological and outcome data (age, sex, tumor site, tumor size, nodal metastases, clinical stage, histologic grade, treatment and follow up) will be analyzed to correlate with fusion status. the mect -maml fusion tested positive in our index patient. we will obtain irb approval to test for the fusion in the archived ffpe specimens and correlate clinical, pathological and outcome data. conclusion: mect -maml fusion is a frequent event in mec that has prognostic and potential therapeutic applications in adults. the results of this study may enlighten the clinical management of mec in children and young adults. children 's mercy hospital, kansas city, missouri, united states background: mutations in the samd gene are associated with a rare syndrome comprising of myelodysplasia, infection, restriction of growth, adrenal hypoplasia, genital phenotypes and enteropathy (mirage syndrome). diagnosis is made through exome sequencing. in the largest reported case series, of eleven patients diagnosed with mirage syndrome, two developed loss of chromosome . given the potent growth restricting activity of samd mutants, the loss of chromosome is considered the first documentation of adaptation by aneuploidy mechanisms in humans and led to myelodysplastic syndrome (mds), with deaths occurring from related complications at and years of age. objectives: to report a case of mirage syndrome with congenital thrombocytopenia progressing to bone marrow failure, managed uniquely with bone marrow transplantation. results: male born at weeks gestation with prenatal diagnosis of iugr, two vessel cord, oligohydramnios was found to have ambiguous genitalia, adrenal insufficiency, partial panhypopituitarism and congenital thrombocytopenia with bone marrow showing absence of megakaryocytic precursors. severe thrombocytopenia was present from birth. bone marrow evaluation demonstrated a hypocellular marrow with markedly reduced megakaryocytic and myeloid precursors and no evidence of myelodysplasia. he required gastric tube placement for failure to thrive, had a laryngeal cleft repaired and developed focal segmental glomerulosclerosis. mpl gene testing for congenital amegakaryocytic thrombocytopenia was negative. testing for fanconi anemia, shwachman-diamond syndrome and dyskeratosis congenita was also negative. approximately % of cells had loss of heterozygosity on chromosome q. exome sequencing showed that he is heterozygous for a de novo gain of function variant, c. g>a (p.arg gln), identified in the samd gene, confirmed by sanger sequencing and consistent with a diagnosis of mirage syndrome. at years of age, he developed pancytopenia requiring frequent transfusions with platelets and packed red blood cells. he underwent a successful bone marrow transplant at years of age without significant complications, and remains transfusion independent without cytopenias greater than months from bone marrow transplantation. conclusion: it is imperative to pursue work up for persistent congenital thrombocytopenia in a stepwise multidisciplinary manner. to the best of our knowledge, this is the first case of mirage syndrome associated bone marrow failure treated with bone marrow transplant. due to the individual rarity of mirage syndrome and pediatric myelodysplastic syndrome, it is important to maintain an index of suspicion given their association and explore bone marrow transplant as a therapeutic option. results: the patient demonstrated disease regression, initially, and continued without disease progression for months. the regimen has been well tolerated with only minimal side effects of dry skin (ctcae grade ) and a transient episode of brief erythrodysesthesia (ctacae grade ) that resolved spontaneously. the combination of sorafenib and capecitabine was effective and well tolerated in this adolescent patient with fl-hcc. our observations, although in a single patient, lend support for further testing of this novel oral chemotherapy regimen in patients with fl-hcc, a disease for which there is no effective standard chemotherapy approach. background: epstein-barr virus (ebv) is a ubiquitous virus associated with a broad range of malignancies due to its oncogenic potential. history of organ or bone marrow transplantation, immunosuppressive therapy, and primary or acquired immunodeficiency syndromes increases the risk of ebvassociated tumors. epstein-barr virus associated smooth muscle tumors (ebv-smt) are unique and rare neoplasms typically discovered in immunocompromised patients. most information related to pathogenesis and therapeutic options is limited to case reports and case series of adult patients. there are several gene expression pathways that ebv utilizes, the most notable of which is the mammalian target of rapamycin (mtor) pathway. the mtor pathway performs a key role through integrating various cell growth signals and factors to regulate protein synthesis and metabolism related to smooth muscle proliferation. sirolimus is an immune modulating therapy that targets the mtor pathway to block activation of lymphocytes. objectives: several case reports have demonstrated shortterm clinical remission of ebv-smt in adult patients with the use of sirolimus. we report the first case of long-term background: bilateral neuroblastoma is characterized as neuroblastoma arising in both adrenal glands, a rare presentation with little data on its genetic make-up. a two-monthold patient was diagnosed with bilateral neuroblastoma in our clinic. her risk assignment was based on biopsy of the left adrenal lesion, which showed mycn amplification, an unfavorable genetic marker. treatment regimen was intensified accordingly and after courses of chemotherapy tumors were excised. patient went on to receive a stem cell transplant and immunotherapy. with no knowledge of genetic similarity between the two tumors it is unclear whether biopsy of the right lesion would have yielded similar results or whether bilateral biopsies are needed for risk assessment of bilateral neuroblastoma. objectives: utilize whole exome sequencing (wes) to characterize the genomic signature of bilateral adrenal neuroblastomas excised following chemotherapy treatment. design/method: paraffin-embedded samples from left (l) and right (r) tumors underwent wes at the broad institute. we analyzed resulting data including somatic variant calls, indel mutations, and copy number variants (cnvs) using ingenuity software to evaluate and compare differences between the two tumor samples. preliminary analysis of the data shows important descriptive information on the two tumor samples. out of somatic mutations in the r tumor cells and mutations in the l tumor cells, only two common somatic mutations were present. out of cnv calls in the r tumor and in the l tumor, cnvs were common between the two tumors, or % of each tumor's cnv calls. there was a fold higher frequency in gains versus losses. the median size of the common cnvs was , (range to , , bp). cancerrelated genes with increased copy numbers included transcription factors, receptors for signal transduction pathways, and histone methylation proteins. conclusion: preliminary analysis of the wes results of the two adrenal tumors show some genomic divergence. because the tumor tissue was exposed to chemotherapy prior to excision it is difficult to determine whether genomic divergence is a result of independently originated tumors or subsequent adaptation to chemotherapy of a clonal cell population. the high number of common cnvs in the two tumors points to a common cell of origin, however the low number of common somatic mutations does not fit that picture. a future study to help elucidate the question will be wes of the original biopsy tissue to provide information on tumor mutations prior to the effects of chemotherapy. baylor college of medicine, houston, texas, united states background: although there has been significant improvement in the overall survival rates of children with cancer many children will still die from their illness or complications secondary to treatment. research surrounding the deaths of children who succumb to their disease is warranted to ensure we are providing the best care possible for these patients. objectives: this case series aims to explore pediatric cancer deaths by focusing on perhaps the most extreme cases of high intensity end of life care. we explore those patients whom we know are dying or our very likely to die as evidence by their do not resuscitate (dnr) orders. in all of these cases despite the patients very grim prognosis, their great likelihood of death and limitations placed of resuscitation methods all patients continued end of life care in the pediatric intensive care unit (picu). the primary medical records of all children with a cancer diagnosis who died between february , and january , in the picu with a dnr order seven days or earlier prior to death. each medical history included disease-directed treatment history and response with particular attention to the events surrounding the terminal admission. results: eight patients met criteria for this study representing . % of all cancer patients who died during this time period and . % of those who died in the icu. the average time between dnr and death is . days ( days - days). the average length of terminal admission was . days ( day - days). the average time between diagnosis and dnr is . months ( months - months). the average time between diagnosis and death is . months ( months - months). conclusion: these cases highlight the journey that patients, families and providers endure leading up to death. medical care is complex, there are very few absolutes that are encountered when caring for patients and decisions around limiting or withdrawing medical care are made in a context of the prior journey. . these cases help to understand the complexity of death and how two seemingly opposite ideals can be congruent in the event of an anticipated death. most of these cases show the need for improved anticipatory guidance surrounding death and greater consideration for de-escalation of care when death is expected. the hospital for sick children, toronto, ontario, canada background: rhabdomyosarcoma (rms) is the most common soft tissue sarcoma in children, with embryonal (erms) and alveolar (arms) representing the most common subtypes. arms tumors are associated with inferior outcome when compared to erms, and they are characterized in about % of the cases by a t ( ; ) or t( ; ) chromosomal translocation with creation of a pax -foxo or pax -foxo fusion gene, respectively. it is increasingly clear that the pax-foxo fusion status is an important poor prognostic factor, thus the histological classification tends to be replaced by the fusion status, particularly in terms of risk stratifica-tion in contrast to arms, there are no recurrent chromosome alterations in erms; however, there are multiple numerical chromosome changes that are frequent in these tumours: gain of chromosome , , and have been found in to % of emrs karyotypes. moreover, erms tumors show frequently allelic loss, the .p . chromosomal region being the most frequently involved. recently, novel gene fusions have been described also in erms tumours. these fusions involved mainly the ncoa and or the vggl genes. the rearrangement partners are variable, and include, i.e. pax ( q ), srf ( q ) and tead ( p ). objectives: to present a patient who died as a consequence of brain metastases while on therapy in the setting of an foxo negative rms and the identification of a new translocation t( ; )(q ;q ). design/method: case report and retrospective review of the literature. we report a case of pelvic embryonal rhabdomyosarcoma in a -month old boy. he was treated as per cog arst intermediate risk group, but unfortunately was found to have a large cerebellar tumour during the course of his chemotherapy treatment and he subsequently passed away. a novel translocation between chromosomes and was observed in of metaphase cells by g-band analysis in the autopsy sample of the brain lesion. breakpoints of the translocation were estimated to be at q and q . there were no additional clonal chromosome abnormalities in the tumour cells. conclusion: erms tumors with fusion genes involved have been exclusively described in patients less than months of age; they seem to be associated with spindle cell histology and, a favorable outcome. in our patient, a novel ( ; ) translocation was found and clinically, the patient had a dismal outcome. further studies are indicated to inquire whether this finding is of significance in term of prognosis for these patients. children 's national medical center, washington, district of columbia, united states background: iatrogenic immunodeficiency-associated lymphoproliferative disorders (lpds) are a group of lymphoid s of s proliferations or lymphomas that are well known to be associated with an immunosuppressed state. these disorders most commonly occur following hematopoietic or solid organ transplantation (called post-transplant lymphoproliferative disorders or ptld), but cases have also been described during the treatment of autoimmune and rheumatologic disorders by immunosuppressive and immunomodulatory medications. these disorders are strongly associated with infection by the epstein-barr virus (ebv) as a result of impaired immune function in the immunosuppressed state. while this phenomenon has been well documented in autoimmune conditions, cases affecting pediatric patients while on antileukemia chemotherapy are lacking. background: atypical teratoid/rhabdoid tumor (at/rt) of the central nervous system (cns) in children younger than years old has a prevalence of % to % and accounts for . % of all pediatric cns tumors. only - % of patients have leptomeningeal dissemination. rhabdomyosarcoma is the most common soft tissue tumor in childhood, but represent only - % of all pediatric cancers. rarely, it can metastasize or even directly extend into the cns, but typically, cases of cns involvement arise either from parameningeal areas or other primary sites. primary spinal or meningeal rhabdomyosarcoma is extremely rare. objectives: our objective is to describe two unique cns malignancies presenting as rare, primary leptomeningeal disease. design/method: case a -month-old female presented with vomiting, fatigue and listlessness, despite a normal head ct and brain mri. csf showed hypoglycorrhachia and mild pleocytosis. ceftriaxone was started, but she developed nuchal rigidity and cranial nerve vii palsy. repeat brain mri showed evolving leptomeningeal enhancement concerning for meningitis. she gradually developed worsening opisthotonus and ultimately a brain biopsy of the temporal lobe was consistent with at/rt. case a -year-old male presented with new generalized tonic-clonic seizure activity and intermittent headaches with photophobia, phonophobia, and vomiting. brain mri was significant for enhancement of interpenducular and suprasellar cisterns extending to the optic nerves and chiasm most consistent with meningitis. neurosurgery ultimately placed a lumbar drain for hydrocephalus, and a tissue biopsy demonstrated primary meningeal rhabdomyosacroma. results: in case , our patient's temporal lobe biopsy demonstrated grade iv malignant tumor cells consistent with atypical teratoid/rhabdoid tumor. fish demonstrated a homozygous deletion of smarcb ( q . ). she was started on chemotherapy per the dana farber at/rt protocol but ultimately was discharged home on hospice. in case , our patient's lumbar arachnoid biopsy demonstrated cellular tumor consistent with group iiia embryonal rhabdomyosarcoma. immunostaining was positive for cd , desmin, myogenin, and myo-d with neural markers ema and gfap highlighting the meninges but without a neural component to the tumor. he completed craniospinal radiation to gy total with lumbar boost to . gy total. he is currently receiving chemotherapy per arst protocol. conclusion: these two cases are particularly instructive because of their similar initial presentations and neuroimaging, but with very different and unique diagnoses. university of iowa, iowa city, iowa, united states background: ebf -pdgfrb fusion causes ph-like b-cell acute lymphoblastic leukemia (b-all), which has a philadelphia positive phenotype without the bcr-abl translocation. this is one of several mutations associated with ph-like b-all and leads to downstream overexpression of tyrosine kinase. ebf -pdgfrb fusion accounts for about % of children with ph-like b-all. patients with ph-like b-all previously had poorer outcomes with conventional chemotherapy. the addition of tyrosine kinase inhibitors (tki), like imatinib, has improved the outcome for many patients predicted to have tki sensitive mutations. objectives: to review clinical characteristics and outcomes of two cases of ph-like b-all at the university of iowa stead family children's hospital and to compare these outcomes to similar cases reported in the literature. design/method: a retrospective chart review was performed for two cases of ph-like b-all diagnosed and treated at the university of iowa stead family children's hospital. results: both patients were males diagnosed at years of age with high wbc count ( , and , ) and positive for ebf -pdgfrb gene fusion. patient (pt ) was cns b at presentation while patient (pt ) was cns negative; neither had testicular involvement. both started treatment according to cog protocol aall . peripheral blasts cleared by induction day for pt and induction day for pt . at end of induction, pt had m bone marrow and pt had m bone marrow but mrd %. dasatinib was started induction day for pt and induction day for pt . pt was still not in remission at end of consolidation; bone marrow cell culture for tki resistance showed best response to dasatinib. pt proceeded to anti-cd car t-cell therapy followed by tbi-based matched unrelated donor bone marrow transplant. pt had negative mrd at the end of consolidation and continues chemotherapy according to aall , dasatinib arm. both patients are currently clinically well. our patients had the same tyrosine kinase gene fusion and similar initial clinical courses. while both patients had persistent disease at end of induction, pt had almost % blasts while pt had significant reduction of disease burden before starting tki. pt showed good response with the addition of dasatinib while pt did not. these findings suggest that response to conventional chemotherapy may potentiate the effect of tki and may predict overall outcome. there are likely additional factors which must be taken into account when determining response to tki for patients with ph-like b-all which have not yet been identified. background: medulloblastoma is the most common malignant brain tumor of childhood. classically, medulloblastoma presents as a well-defined mass lesion in the cerebellum, with a high rate of metastatic dissemination. primary leptomeningeal medulloblastoma (plmb) is an exceedingly rare type of medulloblastoma presentation with a dismal prognosis in which patients present with isolated leptomeningeal disease without an associated mass. to our knowledge, only three pediatric and three adult cases of plmb (ages - years) have been reported, all of which died within months of diagnosis. this is the first case of plmb to report a molecular classification. objectives: to report the case of a pediatric patient with plmb in which histopathologic and molecular characterization was performed and to describe the patient's treatment and clinical course. design/method: retrospective review of the patient's electronic medical record and review of the literature. a -year-old boy presented with headache, vomiting, diplopia, and fatigue. physical examination revealed upward gaze palsy, left-sided extremity and facial weakness, and ataxia. magnetic resonance imaging (mri) of the brain revealed diffuse cerebellar leptomeningeal enhancement and edema without an identifiable mass and moderate hydrocephalus. mri of the spine and cerebral spinal fluid analysis were normal. a diagnosis of cerebellitis was rendered, and the patient underwent placement of a ventriculoperitoneal shunt. an extensive infectious, neurologic, rheumatologic, and oncologic workup did not identify an etiology. empiric antibiotics, high-dose steroids, and intravenous immunoglobulin therapy yielded minimal improvement. two months later, repeat mri of the brain performed for declining mental status demonstrated progressive thickening of cerebellar leptomeningeal disease. a suboccipital craniectomy with decompression and cerebellar biopsy were performed. pathologic examination revealed a diagnosis of plmb, classic histology, non-wnt/non-shh, without gain/amplification of myc/mycn, and p wild type pattern. craniospinal radiation to cgy with a cgy boost to the posterior fossa was delivered with concurrent carboplatin/vincristine over six weeks. two months following chemoradiation, mri of s of s the brain demonstrates significantly reduced pathological leptomeningeal enhancement of the cerebellum, and the patient is awaiting initiation of systemic chemotherapy while recovering from a surgical wound infection. conclusion: plmb is extremely rare but should be considered in patients with cerebellitis and diffuse leptomeningeal involvement who are refractory to medical management or in whom an etiology has not been identified. cerebellar biopsy is recommended early to enable timely treatment and improved outcomes. molecular classification should be performed in cases of plmb to further characterize this disease, inform treatment decisions, and improve clinical outcomes. background: primary intracerebral osteosarcoma is extremely rare and limited to case reports. ptpn gain of function is associated with noonan syndrome, which has increased risk of multiple cancer types including brain tumors, but osteosarcoma has never been described. ptpn mutations have been reported in many cancers as both oncogenes and tumor suppressors, however no ptpn mutations have been described in osteosarcoma. pdgfr-a is a growth factor receptor whose activation is implicated in several malignancies. pdgfr-a and ptpn concurrent mutations are described in glioblastoma. there is no known link between holoprosencephaly, noonan syndrome, and osteosarcoma. we report a case of multifocal intracerebral osteosarcoma in a child with lobar holoprosencephaly and chronic subdural hemorrhage and discuss the genetic changes found in the tumor. design/method: a seven-year-old caucasian female, with a known diagnosis of lobar holoprosencephaly, chronic subdural hemorrhage and well controlled seizure disorder presented with status epilepticus shortly after completing antibiotic therapy for infection of subdural hematoma. mri showed diffuse dural thickening with mass lesions in the frontal lobe, temporal lobe, and the parasagittal region, the largest of which was contiguous with the subdural space but none of the lesions were associated with bone on mri or by direct neurosurgical visualization. tissue obtained for concern for recurrent infec-tion resulted in a diagnosis of high grade osteosarcoma. dna analysis was performed to help guide treatment choice. results: standard metastatic work-up was negative for skeletal primary tumor or metastatic lesions outside of the brain. she was treated with high dose methotrexate for two cycles per modified aost . despite maximal supportive care, she quickly developed rapid tumor growth as well as intratumoral hemorrhage with resultant herniation and death from respiratory failure just three months after diagnosis. tumor gene sequencing discovered three mutations with described roles in cancer: pdgfra d >vr, kdm a loss of exons - , and ptpn a v. conclusion: to our knowledge, primary multifocal extraosseus intracerebral osteosarcoma has not been previously described. despite known cns penetration of high dose methotrexate, this tumor proved resistant and aggressive. holoprosencephaly is associated with a multitude of known genetic drivers, but none are found in this case. furthermore, the genetic changes in this tumor are not typical for osteosarcoma. pdgfr-a over-expression is described in osteosarcoma, but is not clearly correlated with worse overall survival. further research is required to determine the role of ptpn in osteosarcoma. background: anaplastic lymphoma kinase (alk) encodes a receptor tyrosine kinase whose activation induces pathways associated with cell proliferation, angiogenesis, and cell survival. alk rearrangements are rare in neuroblastoma, while alk mutations and gene amplification occur more frequently. alk mutations have been found to be associated with increased alk protein expression that is associated with a worse prognosis. alk is commonly mutated in neuroblastoma at three hotspots (f , r , and f ). the eml -alk rearrangement has mostly been associated with lung adenocarcinomas, with only a few cases of non-lung cancers found. it has never been reported in neuroblastoma. multimodal therapy and to report the successful management of treatment related iron overload. results: a -year old male presented with abdominal swelling and ct showed a right kidney mass and bilateral lung nodules. he underwent right radical nephrectomy with lymph node sampling. pathology was reviewed centrally and revealed wilms tumor with diffuse anaplasia with rhabdomyosarcoma arising within the stromal component and of nodes positive. he received adjuvant intensive chemotherapy and radiation to the hemiabdomen and whole lungs. the -week chemotherapy regimen was vincristine, doxorubicin, cyclophosphamide (per cog arst ) alternating with carboplatin and etoposide (per cog aren revised uh- ). treatment was complicated by multiple episodes of fever and neutropenia and anorexia requiring g-tube placement. post-therapy, he had persistent neutropenia and thrombocytopenia without related complications. every months for evaluations he underwent a bone marrow which revealed normocellular marrow with maturing trilineage hematopoiesis. evaluation for a bone marrow failure syndrome was unrevealing. starting at months into therapy and all posttherapy imaging showed splenomegaly. he received units of packed red blood cells through the duration of therapy. he was diagnosed with iron overload based on serum ferritin and imaging, including t *mri. he received therapeutic phlebotomy for years with normalization of serum iron studies, t * of the heart, and liver iron concentration. he is more than years from completing therapy with no evidence of recurrent disease. asymptomatic cytopenias persist and he has no evidence of iron overload. conclusion: though a rare development, clonal sarcomatous transformation can occur in wilms tumor. our patient's tumor was successfully treated with intensive multimodal therapy targeting the diffusely anaplastic wilms and the rhabdomyosarcomatous component. treatment-related iron overload in a pediatric patient with a solid tumor was successfully treated with phlebotomy. consideration should be given to screen patients with solid tumors who receive multiple packed red cell transfusions for iron overload at the completion of cancer therapy. primary children's hospital, university of utah, salt lake city, utah, united states background: malignant solid tumors are less frequently encountered in infants. primitive myxoid mesenchymal tumors of infancy (pmmti) are a myofibroblastic malignancy and cases are rarely reported in the literature. cure is achieved in the majority of cases with surgical resection, however treatment for unresectable cases remains an enigma. recently published literature postulates that the newly discovered bcor duplication found in pmmti is tumorigenic via an epigenetic pathway. this molecular signature resembles that of clear cell sarcoma of the kidney (ccsk) and the growing number of bcor mutated sarcomas. a similar chemotherapeutic backbone and local control used for ccsk, has been proposed for the unresectable subset of pmmti. utilizing this approach a month-old with relapsed disease has remained disease free for months. however, given the rarity of this disease and the lack of published literature, there is no known standard of care treatment for unresectable and/or recurrent ppmti. we report a case of unresectable recurrent pmmti, a rare infant tumor, with less than cases reported. design/method: medical record, radiological studies, pathology and literature was reviewed. results: our patient is a now month-old female who presented with constipation and lower extremity weakness in the first weeks of life. an mri demonstrated a large lumbar epidural mass with spinal cord impingement. given prolonged (> days) neurological symptoms and location, emergent chemotherapy was initiated. biopsy showed a bcor positive, primitive myxoid mesenchymal tumor of infancy (pmmti). she was treated with ifosfamide, carboplatin and etoposide, and demonstrated clinical and radiographic response. we gave two additional cycles of cyclophosphamide, carboplatin and etoposide until surgical resection was feasible followed by two post-surgical cycles of chemotherapy. unfortunately, four month post-therapy mri demonstrated two new lesions; an unresectable paraspinal soft tissue mass and a left iliopsoas groove mass. given bcor association and reported successful therapy with vincristine, doxorubicin, cyclophosphamide alternating with ifosfamide and etoposide, we elected to incorporate vinca-alkaloid and anthracycline into her regimen. she is being treated with vdc/ie with plan for radiation consolidation. conclusion: pmmti is a locally aggressive tumor, for which surgical resection is curative. for those not amendable to resection, best care practices are still being determined. we report a case of pmmti initially responsive to chemotherapy, but not curative. this is the second case to conclusively demonstrate chemo-responsiveness. bcor mutation seems to be a common feature of this cancer; its role in the pathogenesis and as a target is an area of investigation. medical college of wisconsin, milwaukee, wisconsin, united states background: atypical teratoid/rhabdoid tumors (atrt) are central nervous system (cns) tumors that most commonly occur in very young children. there is no widely accepted standard of care for atrt patients, and while survival rates are improving they are historically poor. patients with metastatic disease to the spine at diagnosis have a worse prognosis, and for patients > years old, the presence of metastatic disease often results in the use of craniospinal radiation. the importance of correctly identifying metastatic disease at diagnosis aids in decision making and can have both prognostic and therapeutic implications. mr imaging at diagnosis is used to identify metastatic disease; however, here we present a case of diffuse leptomeningeal enhancement that spontaneously resolved after resection of a primary supratentorial atrt. objectives: to describe the resolution of diffuse leptomeningeal enhancement after resection of a primary atrt tumor in a -month-old prior to any adjuvant therapy. results: a -month-old male presented with a month history of vomiting and weight loss, regression of gross motor developmental milestones, and left hemiparesis. a brain mri demonstrated a × . × . cm solid and cystic right atrial mass with diffusion restriction and post-contrast enhancement. smooth diffuse enhancement was noted along the surface of the brainstem and within the interpeduncular fossa. a spine mri demonstrated diffuse circumferential post-contrast enhancement along the surface of the entire spinal cord. the patient underwent a successful near total surgical resection of the primary mass. pathology confirmed the loss of ini- staining in tumor cells, consistent with a diagnosis of atrt. no immediate adjuvant radiation or chemotherapy was given. repeat imaging was completed days after resection. brain mr demonstrated expected post-operative changes within the surgical cavity without definitive residual mass or leptomeningeal enhancement. spine mr demonstrated complete resolution of the previously seen circumferential enhance-ment along the entire spinal cord. csf evaluation at that time was negative for tumor cells. after recovery from surgery, chemotherapy treatment was initiated. conclusion: leptomeningeal enhancement at the time of diagnosis of atrt has historically been considered clear evidence of metastatic disease. this case raises questions about the previously accepted etiology of these imaging changes and suggests that widespread leptomeningeal enhancement should be carefully interpreted in future patients with similar imaging findings. in this setting, clinicians should consider repeat imaging following primary surgical resection in order to provide appropriate prognostic information and inform therapeutic decisions. poster # primary ewings sarcoma of cervical cord mimicking cauda equina syndrome sucharita bhaumik, joshua chan nyu winthrop hospital, mineola, new york, united states background: ewing's sarcoma (es) is a malignant primary bone tumor usually involving long bones. primary es of spine is quite uncommon ( . %) and its location in the cervical spine is even more rare. cauda equina syndrome (ces) is symptoms due to damage to the bundle of nerves below the end of the spinal cord known as the cauda equina (low back pain, radiating shooting pain down the legs, paraplegia, and loss of bowel or bladder control). it often occurs with lesions of lumbosacral spine. treatment with high-dose steroids may provide pain relief and improved neurologic function (by reducing edema) while awaiting diagnostic studies objectives: to demonstrate an unusual clinical presentation and emergent management of cervical es presenting with ces like symptoms. : year old male presented with a left sided posterior neck mass. soon after, he developed weakness of left arm, urinary and stool retention and inability to walk or bear weight in both legs. on physical exam a left tempero-occipital × cm fixed, non-tender, non-fluctuant mass was noted as well as motor and sensory impairment of left upper extremity, bilateral spastic paraplegia and loss of sphincter control. mri cervical spine showed a left cervical tumor with moth eaten appearance involving the vertebral bodies of c -c , adjacent muscles, displacing vital structures of the neck and compressing the cervical spinal cord. the thoracic and lumbosacral spine had no disease involvement. due to rapidly worsening spinal cord compression he was emergently treated with high dose steroids. he gained back all function in his extremities and regained bowel and bladder control. this eliminated need for urgent neurosurgical intervention. results: biopsy of the neck mass showed small blue round cells consistent with es with ewsr gene rearrangement. staging work up revealed no additional metastatic involvement. he then initiated treatment for localized es with systemic chemotherapy and radiotherapy and has had excellent response to treatment so far. conclusion: this is the first known case of non metastatic primary cervical es mimicking ces where an acutely enlarging mass presented with rapidly progressive neurologic deficits due to compression of anterior spinothalamic tract. in these unusual presentations of ces without lumbodorsal involvement it is important to consider cervical lesions. early rapid steroid initiation should be considered while awaiting biopsy results to prevent worsening cord compression followed by es focused treatments. this increases the chance of a successful outcome. the initial improvement with steroids may confuse the tumor with being a lymphoma children 's mercy hospital, kansas city, missouri, united states background: von willebrand disease (vwd) is a relatively common bleeding disorder with a high degree of genotypic and phenotypic variation. bleeding is usually mucocutaneous but can be severe and include muscle and joint bleeds especially in type vwd patients. most common bleeding management consists of desmopressin, anti-fibrinolytics, and/or plasma-derived antihemophilic factor/von willebrand factor (ahf/vwf) complex. a recombinant vwf has become available in the last few years. anaphylaxis and inhibitor development in vwd are rare. objectives: to describe the rare clinical manifestation of anaphylaxis to factor concentrate in a patient with severe type vwd. results: a -year-old female with severe type vwd [baseline vwag %, activity < %, factor viii (fviii) %] originally presented with heavy menstrual bleeding (hmb) leading to anemia requiring blood transfusion. she underwent placement of a levonorgestrel-releasing intrauterine device (lngiud) and began norethindrone. her hmb continued despite the lngiud and an increase in norethindrone dosing. plasma derived ahf/vwf complex was administered, which she had previously received. following the infusion, the patient developed anaphylaxis with hives, wheezing, tachycardia, and itching requiring doses of diphenhydramine and dose of hydrocortisone with resolution of symptoms. subsequently, she received recombinant vwf without incident. however, due to her low fviii level, she also required treatment with a full length recombinant fviii product. she again developed hives and itching after this infusion. she has since received recombinant vwf with recombinant fviii/fc fusion protein without further allergic reaction. there was no evidence of an inhibitor with her most recent post-infusion vwf level was %, factor viii %. conclusion: anaphylaxis to plasma derived factor products has been documented far less frequently within the vwd population compared to those with hemophilia and is typically seen in those with large gene deletions, usually with type disease. therefore, similar type vwd patients with severe disease may benefit from gene sequencing. it is unclear in this patient's case to which aspect of her treatment she is allergic, as she reacted to plasma-derived ahf/vwf and full length recombinant fviii, but not recombinant vwf or recombinant fviii/fc fusion protein. we hypothesize that she may be allergic to an epitope in the fviii b domain, or that the presence of fc fusion may have had a protective effect. further investigation including genetic analysis is planned. nodules. biopsies were consistent with neuroendocrine carcinoma, large cell type (g ). next generation sequencing revealed a khdrbs -braf fusion. he received conventional cytotoxic chemotherapy regimens both with cisplatin/doxorubicin, capecitabine/temozolomide, and doxorubicin/etoposide, but achieved a minimal response followed by rapid disease progression, massive ascites, and renal failure secondary to bilateral ureteral obstruction. results: based on his prior genomic testing, therapy with single agent mek inhibitor (trametinib) was initiated. this produced a rapid, dramatic response with greatly reduced disease burden at all sites, resolution of ascites and return to completely normal activity within months. this response lasted for approximately months before the tumor again progressed. further therapy with an erk inhibitor was ineffective, and the patient expired from progressive disease. located on the chromosome q , the braf oncogene, as part of the ras/mapk pathway, is involved in cellular proliferation, differentiation, migration, and apoptosis. braf mutations are recognized in a wide range of adult malignancies: thyroid cancers, non-small cell lung cancer, cholangiocarcinoma, ovarian cancers, and multiple myeloma. braf mutations have also been described in adult neuroendocrine carcinoma of the colon. trametinib is a highly specific inhibitor of mek /mek , a downstream mediator in the braf pathway. it has demonstrated activity in a number of tumors including advanced melanoma and gliomas. trametinib was chosen for this patient based on his atypical braf fusion. we believe this is the first documented case of its successful use in neuroendocrine carcinoma in the pediatric population. conclusion: this case demonstrates the presence of braf fusion in a case of pediatric neuroendocrine carcinoma and significant response to single agent mek inhibition in this context. this cases raises the question as to whether the combination of a targeted inhibitor, in addition to either conventional chemotherapy or other braf inhibitors, might offer a better approach to therapy than current treatment options. albany medical center, albany, new york, united states background: warm autoimmune hemolytic anemia (waiha) is characterized by autoantibody, and occasional complement binding of protein antigens, on the surface of red blood cells at temperatures ≥ oc resulting in targeted destruction. we describe the case of a year old male with a history of evan's syndrome, poor immune response to vaccines and lymphoid hyperplasia, presenting with altered mental status and severe anemia, found to have a warm igg pan agglutinin with evidence of both intra and extravascular hemolysis. his course was complicated by respiratory failure requiring intubation, pulmonary emboli, enterococcus bacteremia and hypertension. he received multiple transfusions with only transient increases in hemoglobin. the aiha was refractory to multiple rounds of treatment with high dose steroids, ivig, rituximab, cyclophosphamide, bortezomib, plasma exchange and mycophenolate mofetil (mmf). objectives: given the refractory nature of our patient's aiha the decision was made to trial eculizumab, a monoclonal antibody targeting c complement, preventing its cleavage and activation, and shown to be effective in treatment of atypical hemolytic uremic syndrome and hemolysis due to an igm cold agglutinin. prior to eculizumab infusion, ch and sc b- assays were significantly elevated. design/method: the patient was given two doses of eculizimab days apart. results: his hemoglobin steadily rose independent of red cell transfusions with a corresponding decrease in reticulocyte count, ldh and ch levels. the patient has remained stable with a normal hemoglobin ( - g/dl) on maintenance steroids and mmf. although we cannot definitively conclude that eculizumab directly caused his recovery, the clinical course post-eculizumab suggests this may be an efficacious treatment for aiha. genetic testing showed monoallelic frameshift mutation of the nfkb gene and monoallelic missense mutation of the dock gene. given the role of nfkb in both immunodeficiency and autoimmunity, it is thought that the patient's phenotype is due to nfkb haploinsufficiency and he is currently considering hematopoietic stem cell transplant. st. joseph's regional medical center, paterson, new jersey, united states background: heterozygous -thalassemia typically manifests as thalassemia minor, characterized by mild microcytic hypochromic anemia with minimal clinical ramifications. coinheritance of -globin gene triplication has been reported to exacerbate the clinical and hematological phenotype ofthalassemia trait, due to increase in the alpha/non-alpha-chain imbalance. reported phenotypes range from asymptomatic thalassemia minor to moderate thalassemia intermedia, usually diagnosed in adulthood without transfusion dependence. this combination has been described in mediterranean, european and asian populations, but rarely reported in hispanics. objectives: to report two cases of unusually severethalassemia intermedia in hispanic patients with heterozygosity for triplicated -globin gene and a ( )-thalassemia allele. results: case : sixteen-month-old male of mexican descent presented with persistent microcytic anemia and jaundice. peripheral smear showed nucleated rbcs with basophilic stippling and target cells. hemoglobin electrophoresis revealed: hba- %, hbf- %, hba - . %. -globin gene testing revealed heterozygosity for ( ) mutation (ivsi-i, g→a). given the unusually severe anemia, -gene testing was performed which showed -globin gene(anti . ) triplication ( / ). at four years, he had splenomegaly and bilateral maxillary prominence. head ct showed irregular contour of the parieto-occipital region due to medullary expansion. due to significant persistent anemia ( - g/dl) and progressive bony deformities of the skull, patient began chronic transfusions at age eight after family declined splenectomy.case : fifteen-year-old female, of peruvian and honduran descent, presented for evaluation prior to cholecystectomy for gallstones and recurrent ruq pain. father had known thalassemia trait. her hb was . g/dl with hypochromia, microcytosis, and target cells. electrophoresis indicated -thalassemia trait (hba- %, hba - . %, hbf- . %), confirmed by gene testing (heterozygous for a ( ) mutation in codon c>t). given jaundice and gallstones, -globin gene analysis was ordered showing triplication ( / ). ruq pain resolved post-cholecystectomy, but she developed persistent painful splenomegaly. she began hydroxyurea to increase gamma-globin production and decrease excess alpha chains, but it was discontinued due to hematological toxicity. due to recurrent luq pain and progressive splenomegaly, she underwent laparoscopic splenectomy at age with resolution of symptoms and improved hemoglobin. conclusion: -globin gene testing should be considered in -thalassemia carriers with an atypical clinical presentation including hispanic patients. the wide variability in the phenotypic expression of (anti . ) mutation andthalassemia trait suggest interplay of other genetic factors which remain undefined. the clinically significant presentation amongst certain subjects, as in our two cases, makes it imperative to identify these factors to aid in phenotype prediction and genetic counseling. ashley bonheur, shivakumar subramaniyam, jogarao vedula, sucharita bhaumik nyu winthrop hospital, mineola, new york, united states background: wilms tumor (wt) is one of the most common solid malignant neoplasms in children. a diverse range of genes and mechanisms are implicated in wt pathogenesis. predisposing syndromes result from a disruption of wt gene, crucial for renal and gonadal embryogenesis. another gene is wt gene locus at p , an area of imprinting. the p tumor suppressor gene on chromosome p . is seen in patients with anaplastic histology. in addition to these genes, whole and partial chromosome gains of q, , q, , , & and losses of p, p, q, q, as well as loss of heterozygosity (loh) are commonly seen. some genetic markers appear to be predictive of outcome and are now incorporated into the assigning of risk-directed therapy. patients with loh at chromosome p and q are treated with more intensive chemotherapy, as they have been associated with increased risk of relapse and mortality. objectives: to describe a new complex translocation involving chromosome , , and in a case of pediatric wt. design/method: a four-year old female presented with abdominal pain and emesis. on exam, patient had a firm and large abdominal mass. radiologic studies revealed a complex lobulated right renal mass. right radical nephrectomy was performed. histopathologic studies showed wt with triphasic histologic features with blastema predominance, invasion of the lymphovascular and perinephric adipose tissues, perinephric lymph node involvement and no anaplasia. chest ct scan showed bilateral lung metastases. tumor cytogenetics showed an abnormal karyotype, a complex translocation of , , and . the rearrangement occurred due to translocation between chromosomal bands q and q , with an insertion of q - on the q region. pcr based genotyping using microsatellite markers additionally identified loh for chromosome p and q . the patient was treated for high risk stage iv wilms tumor with favorable histology and received intensive chemotherapy and radiation therapy to the flank and the lungs. she is now in remission months after, with no evidence of recurrence on surveillance scans. complex translocations associated with wt have not been rigorously studied. a question for further study is whether there is any relationship between recurrence potential with a complex translocation compared to common chromosomal abnormalities. further knowledge of the molecular pathology and genetic changes in wt will help the development of new targeted therapies, as well as new biomarkers to aid diagnosis, risk stratification, and monitoring of treatment and relapse. results: a week-old girl was referred for evaluation of an abnormal newborn screen. mother was a known carrier of hb khartoum trait while father was a known carrier of thalassemia trait. patient's hemoglobin quantification performed by capillary zone electrophoresis showed hbf %, hb variant %, and no detectable hba. the hb variant ran in the d zone, a pattern consistent with mother's hb. alkaline agarose gel electrophoresis banding pattern showed f/s. acid agarose gel electrophoresis pattern showed v/f. later testing revealed abnormal isopropanol stability with + precipitation at minutes. this electrophoresis pattern is consistent with the pattern previously reported of hb khartoum. clinically, the patient is a healthy, active child whom we have followed for two years. she has not had any significant anemia outside of her physiologic nadir. she has not had any hemolytic episodes, and her bilirubin levels have always been within the normal range conclusion: to the best of our knowledge, this is the only reported case of hb khartoum/ thalassemia. the proline to arginine substitution of hb khartoum introduces a charged group on the chain at the site of contact. the resulting unstable chains can dissociate into monomers and favor the formation of methemoglobin, leading to hemoglobin instability. we had wondered if this unstable hemoglobin might result in clinical hemolysis when challenged with oxidative stress, such as in periods of infection. however, in the two years we have followed this patient, she has never had a hemolytic episode. at two years of age, she has hbf . %, hb khartoum . %, and hba . %. whether hbf elevation is protective from oxidative stress remains to be determined as we continue to follow this child. university of puerto rico -medical science campus, san juan, puerto rico, united states background: gm gangliosidosis is a lysosomal disorder caused by -galactosidase deficiency due to mutations in the glb gene. it is a rare autosomal recessive neurodegenerative disorder with an incidence of about : , - : , live births worldwide. this neurological disorder has three clinical forms. gm type , or infantile form is characterized by psychomotor regression by the age of months, visceromegaly (hepatosplenomegaly), macular cherry red spot, facial and skeletal abnormalities, seizures, and profound intellectual disability. we present a -year-old female with gm type and acute lymphocytic leukemia (all). design/method: she was diagnosed with gm type at the st months of age and family history was remarkable for an older sister with gm type . diagnostic studies reveal homozygous exon of the glb gene for a sequence variant defined as c. c>t, predicted to an amino acid substitution p.aarg cs. results: patient presented to our hospital with petechiae in lower extremities, pallor and intermittent tracheal bleeding. physical examination shows a hemodynamically stable girl that is chronically ill dependent of mechanical ventilation, severe mental retardation and scatter petechiae at upper and lower extremities. laboratory workup revealed severe normocytic anemia (hgb: . g/dl) with immature peripheral cells and thrombocytopenia ( × /l). serum chemistry revealed increase ldh ( u/l), increase hepatic enzymes (ast: u/l), normal uric acid level. there was no evidence coagulopathy. chest x ray was unremarkable except for evidence of chronic pulmonary illness. abdominal sonogram hepatosplenomegaly. during hospitalization, bone marrow aspirate and biopsy was performed which was diagnostic of b cell acute lymphoblastic leukemia (all) with . % lymphoblast and orderly myeloid/erythroid maturation. flow cytometry: % b lymphoblast with aberrant phenotype c/w b-acute lymphoblastic leukemia. karyotype revealed hyperdiploid female of favorable prognosis. cytogenetic by fish: hyperdiploid all with extra copies of runx and igh (no bcr-abl translocation). family was oriented about the new diagnosis and the dismal prognosis in conjunction to her primary condition. parents agree on no chemotherapy treatment for all with only supportive treatment. to this date, there is no evidence in literature that has previously described association of gm and leukemia. life expectancy of patient's primary condition is null therefore, correlation with leukemia might not be a coincidental finding. this patient opens the possibility of malignancy as part of gm type thus, malignancy diagnosis should be considered as part of their medical lifetime course. university of south florida, tampa, florida, united states background: hematological manifestations related to hiv infection are not uncommon, with thrombocytopenia having an estimated prevalence of - %. the pathophysiology is likely multifactorial. studies suggest that the primary mechanism may be immunologic resulting in accelerated platelet destruction. additional theories suggest that infection of megakaryocytes may also play a role causing inadequate platelet production. treatment of hiv-related thrombocytopenia is challenging. first-line treatments include initiation and optimization of antiretroviral therapies, immunoglobulin (ivig), and glucocorticoids. however, this approach is not effective in all patients and second line treatment options are less well studied, particularly in the pediatric population. objectives: we aim to present and discuss the case of a year old patient with perinatally acquired hiv- infection and persistent thrombocytopenia who, after failing first line therapies, showed normalization of platelet count on the novel thrombopoietin receptor agonist, eltrombopag. design/method: a retrospective chart review of the case patient's medical record was conducted. additionally, a thorough literature review was performed on this topic including the pathophysiology of hiv related thrombocytopenia and its treatment modalities. the patient required monthly ivig infusions for about year, but did not show a sustained response, often with platelet count dropping to less than , in between infusions. after initiation of mg eltrombopag daily the patient showed a sustained increase in platelet count (range , - , ). during a brief week lapse in eltrombopag treatment his platelet count dropped to , . upon re-initiation of therapy his count increased to , . the patient has remained asymptomatic, off of ivig for over one year, with undetectable hiv viral load and greater than cd t cell counts. no side effects or grade laboratory abnormalities were reported. conclusion: treatment of hiv-related thrombocytopenia can be challenging. first line therapies, including ivig and glucocorticoids, are not effective in all patients. several other treatment modalities have been utilized, including anti-d immunoglobulin, dapsone, danazol, interferon alfa, vincristine, thrombopoetic growth factors including romiplostim and eltrombopag, or splenectomy, but these are less well studied. this represents the first reported case of a pediatric patient with hiv who showed a positive response to eltrombopag with a sustained improvement in platelet count and no adverse effects from treatment. eltrombopag may be a safe alternative to first line therapies in those patients with hiv and refractory thrombocytopenia, however additional studies are needed. university of illinois college of medicine at peoria, peoria, illinois, united states background: achromobacter xylosoxidans is a gram negative rod with peritrichous flagella which causes rare opportunistic infections most commonly encountered by immunocompromised patients. it is primarily associated with uncomplicated bacteremia, cather-associated infections, and pneumonia. most reports of bacteremia associated with a. xylosoxidans are nosocomial, associated with neoplasm, and occurring mainly in adults. most reported infections with a. xylosoxidans in children are associated with cystic fibrosis. there are very few reported cases of septic shock from a. xylosoxidans bacteremia and pneumonia in the pediatric oncology population. objectives: to describe a rare case of a. xylosoxidans septic shock in a pediatric patient with relapsed neuroblastoma results: a -year old boy with history of stage iv highrisk neuroblastoma underwent standard frontline therapy with chemotherapy, hematopoietic stem cell transplant, radiation therapy, and immunotherapy, followed by a dfmo trial for maintenance. his -month follow-up scans demonstrated relapse and he was subsequently treated with additional chemotherapy, surgical resection, and mibg therapy, crizotinib for an eml -alk fusion and finally ifosfamide, carboplatin and etoposide (ice). he developed neutropenic fevers and was started on cefepime, vancomycin and fluconazole. blood cultures were initially negative. on the th day of fever, his previously scheduled pet scan was performed during hospitalization and showed new pulmonary opacities. he did not have respiratory symptoms, but therapy was escalated to meropenem, vancomycin and amphotericin. emergent bronchoscopy was performed the same day, with all bacterial and fungal cultures remaining negative. overnight, he developed tachypnea and saturations in the upper s, requiring nasal cannula. ir-guided lung biopsy was performed the next day, a flexible bronchoscopy was done to remove blood clots in the airway, the patient was placed on a ventilator, femoral lines were placed, granulocytes ordered and pressors were started for deterioration to presumed septic shock. arterial and femoral lines were placed but patient continued to have hemodynamic instability on multiple pressors. the following day, blood and respiratory cultures returned positive for results: at days after the start of iti, the inhibitor was < . bu and continued undetectable months after initiation of iti therapy. in this patient, iti with high-dose plasma-derived factor viii and von willebrand factor (vwf) complex was well tolerated and effective. genetic analysis confirmed a large factor viii gene duplication of exons to . we believe our patient developed inhibitor so quickly ( exposure days) due to the possibility of this mutation causing a frameshift that introduces a premature termination codon. this might be functionally similar to a deletion in the factor viii gene which poses the highest risk for inhibitor development in patients with severe hemophilia a. this variant has only been identified previously in two unrelated patients diagnosed with severe hemophilia a. this duplication is not listed in dbsnp variant database, nor observed in the general population database. our case proves the effectiveness of this method for patients with severe hemophilia a and an inhibitor. it also shows that more research is needed to identify patients at risk for inhibitor development. background: mercaptopurine ( -mp) is a prodrug that is a core component of maintenance chemotherapy for patients with a diagnosis of acute lymphoblastic leukemia (all). suppression of the neutrophil count is used to demonstrate adequate dosing of -mp during this phase of therapy. bone marrow suppression is mediated by the active metabolite -thioguanine ( -tgn), whereas the metabolite -methylmercaptopurine nucleotides ( -mmpn) has been shown to cause hepatotoxicity. allopurinol has been used infrequently in all maintenance therapy in the setting of skewed metabolism when adequate myelosuppression is difficult to achieve due to excessive hepatic toxicity. when given in combination with allopurinol a reduced dose of -mp may result in both increased -tgn levels and decreased -mmpn levels. objectives: describe the characteristics and clinical course of patients treated with allopurinol and reduced dose -mp during maintenance chemotherapy for all. we performed a retrospective chart review of patients at aflac cancer and blood disorders center of children's healthcare of atlanta with new diagnoses of b or t-cell all who received allopurinol during maintenance chemotherapy. we identified eleven patients with b-cell or tcell all who received allopurinol adjunctive therapy during maintenance chemotherapy at a single institution between - . these patients received adjunctive allopurinol for - weeks (median weeks) with reduced -mp ( - % of full dose). all ten patients with genetic testing for thiopurine s-methyltransferase (tpmt) had wildtype genotype associated with normal enzyme levels. indications for allopurinol use were most commonly unfavorable -mp metabolite levels, transaminitis (n = ), pancreatitis (n = ) and hyperbilirubinemia (n = ). favorable metabolite shift was achieved in all patients. liver enzymes improved in of patients with transaminitis after initiation of allopurinol/reduced -mp. three patients who experienced pancreatitis during maintenance did not have recurrence after initiation of allopurinol ( of these patients previously reported). six patients developed pancytopenia while on allopurinol, and two of those patients developed pancytopenia severe enough to require allopurinol cessation. four patients developed isolated anemia (hgb < . g/dl) without thrombocytopenia or severe neutropenia. no patient has experienced a recurrence of leukemia. overall, treatment with allopurinol and reduced dose -mp was successful in producing a favorable -mp metabolite distribution and reducing toxicity. therapy was generally tolerated; however a major and notable side effect was pancytopenia, in two cases severe enough to stop allopurinol treatment. anemia may be more prominent with allopurinol usage. allopurinol effect is variable among individual patients despite normal tpmt genotypes. baylor college of medicine, houston, texas, united states background: congenital sideroblastic anemia, b-cell immunodeficiency, periodic fevers and developmental delay syndrome (sifd) is a rare inherited sideroblastic anemia syndrome, first described in with clinically similar cases. genetic variations of trnt were identified as causative. objectives: to present an unusual presentation of a patient with sifd complicated by diagnosis of concomitant alpha thalassemia trait. design/method: retrospective chart review. a five month old male infant was referred to our hematology center for evaluation of elevated hemoglobin barts identified on newborn screen. despite numerous attempts, blood work was unable to be collected. at seven months of age he had microcytic anemia (hemoglobin . g/dl, mean corpuscular volume fl) more severe than what would be expected with alpha thalassemia trait. no variant hemoglobin was identified with isoelectric focusing or high performance liquid chromatography. by nine months of age he developed growth failure, intermittent emesis with fevers, developmental delays (predominantly gross motor), hearing loss, a disproportionally large head and coarse, thinning hair. over the next ten months, he was seen by numerous specialists for seemingly unconnected problems including sensorineural hearing loss, elevated liver enzymes and growth hormone deficiency. alpha globin analysis revealed deletion of two alpha globin genes. at months of age, he was admitted with one week of fevers, jaundice, and emesis. peripheral blood smear showed microcytic hypochromic anemia with marked anisopoikilocytosis including target cells, elliptocytes, tear drops, spherocytes, poikilocytes, marked polychromasia, and coarse basophilic stippling. given the inconsistency of his laboratory findings with the diagnosis of alpha thalassemia trait and clinical syndromic findings, bone marrow biopsy was performed which revealed rare ringed sideroblasts. one month later whole exome sequencing revealed trnt splicing variant c. - c>g and novel missense variant c. a>t consistent with sifd. hemoglobin barts on newborn screen with moderate to severe microcytic anemia directed initial diagnostic work-up towards variant alpha thalassemia. as additional medical conditions developed the focus shifted to a unifying syndrome. compared to previously described cases, our patient was diagnosed at an older age, presented with anemia rather than episodes of febrile illnesses, and had rare sideroblasts on bone marrow examination. diagnosis in this case led to identification of the novel c. a>t variant in his sister who had similar, but milder, features. sifd is a rare disease with variable phenotypic severity making diagnosis challenging without high index of suspicion which is crucial for appropriate management. wiseman, blood, . chakraborty, blood, background: cholelithiasis is uncommon in childhood. cholelithiasis is known to occur more frequently in children with predispositions, including female sex, obesity, parenteral nutrition, previous abdominal surgery, use of oral contraceptives, family history of gallstones, chronic hemolytic anemias, hepatobiliary disease, or exposure to specific drugs. although there have been occasional case reports linking cholelithiasis to childhood leukemia or leukemia therapy, the prevalence and risk factors of cholelithiasis in patients with childhood leukemia remain unclear. objectives: to estimate the prevalence of cholelithiasis in patients diagnosed with childhood acute lymphoblastic leukemia (all), and to evaluate possible risk factors for the development of cholelithiasis in patients with childhood all. we performed a computer-assisted review of the electronic medical records of patients diagnosed for b or t-cell all at children's healthcare of atlanta in the period from to . patients with diagnoses of cholelithiasis, cholecystitis or who had a cholecystectomy were identified. possible risk factors of age, sex, bmi, history of abdominal surgery and parenteral nutrition use were abstracted. patients with underlying chronic hemolytic anemia or pre-existing gallbladder disease were excluded. results: seventeen cases of cholelithiasis and cases of cholecystitis without documented cholelithiasis were identified. among patients with cholelithiasis, were female. median age at diagnosis of cholelithiasis was . (range . - . ) years. seven patients had no symptoms referable to cholelithiasis at the time of diagnosis. the median age of leukemia diagnosis among these patients was . (range . - . ) years. the median interval from diagnosis of leukemia to gallbladder disease was . years. four patients had bmi over the th percentile for age. two patients had a prior history of intraabdominal surgery. no patient received oral contraceptive pills. six patients received parenteral nutrition for more than days. there was no documented family history of cholelithiasis. seven patients did not receive any cholelithiasis directed therapy. two patients were managed with medical management only, with endoscopic retrograde cholangiopancreatogram with stone extraction, and with cholecystectomy. our study estimates the prevalence of cholelithiasis in childhood lymphoblastic leukemia to be . %, higher than the reported prevalence in the general pediatric population of . - . %. although our cohort size is small, it appears that all therapy and supportive care modalities associated with all are likely to play a larger role in the development of cholelithiasis than known predisposing factors in the general population. further studies are warranted. background: an uncommon side effect of intravenous immunoglobulin (ivig) administration is clinically apparent, sometimes severe hemolysis. we describe a severe case of coombs-positive hemolytic anemia secondary to ivig administration. ivig is a blood derivative manufactured from pools of , to , individual plasma donations. ivig is not abo-type restricted, so anti-a, anti-b and anti-a,b isoagglutinins are detectable. objectives: to describe a rare but serious type of transfusion reaction leading to gross hemolysis after ivig administration. results: a -year-old male with a past medical history of obstructive sleep apnea and obesity was admitted to the pediatric intensive care unit for adenoviral pneumonia and subsequent respiratory failure requiring mechanical ventilation. he had a complex hospital course with many complications including acute respiratory distress syndrome (ards), septic-shock, and coombs-positive hemolytic anemia. the patient was treated with commercial ivig (baxter/baxalta) -mg/kg daily for five days. he had two isolated episodes of severe hemolysis in relation to ivig administration requiring multiple transfusions of packed red blood cells (prbc). examination of pre-transfusion peripheral blood smear showed spherocytosis with rouleaux formation and large clumped rbc aggregates. the patient's blood type was classified as blood group a, rh-negative and his initial prbc transfusions were of this type. subsequently, the patient's coombs test was found to be positive using polyspecific and anti-igg typing sera. the patient's antibody screen against reagent group o screening cells was negative ruling out autoimmune hemolytic anemia. however, type specific anti-a antibodies were detected in his plasma as well as the acid eludate prepared from the coombs-positive red blood cells. it was concluded that the patient's hemolysis was due to anti-a antibodies presumed to arise from ivig. the patient's rbc transfusions were changed to o-negative blood and the hemolytic process resolved. the patient ultimately died due to complications of ards. although hemolysis is a known side effect of ivig, it is rarely considered when deciding to administer ivig. in addition, it has rarely been described in the pediatric population. ivig is used in the treatment of a growing number of medical conditions. due to the critical nature of many of these patients, hemolysis secondary to ivig may not be considered and continued blood transfusions with the patient's specific blood type may be used. it is crucial to remember that severe hemolysis can occur from ivig, and the importance of transfusing with blood group o, rh-negative blood when applicable. university of maryland medical center, children's hospital, baltimore, maryland, united states background: coagulopathy is a well-described complication of acute promyelocytic leukemia (apml), and remains a leading cause in induction failure. with treatment, coagulopathy associated with apml has been shown to rapidly improve. multiple organ dysfunction syndrome (mods) in apml, including acute respiratory distress syndrome (ards), has been associated with infection, traumatic injury, malignant infiltration, and cytokine release syndrome. when mechanical ventilation is no longer sufficient, extracorporeal membrane oxygenation (ecmo) can be considered; however, coagulopathy, severe end-organ damage, and malignancy are all relative contraindications to initiation of treatment. we report the case of a -year-old female presenting in respiratory failure, disseminated intravascular coagulopathy (dic), with intracranial hemorrhage, and mods, diagnosed with apml, successfully treated with ecmo therapy. design/method: retrospective case analysis and literature review. our patient, a -year-old female was admitted in respiratory failure and altered mental status, following a fall shortly prior to presentation. initial laboratory values were notable for pancytopenia, dic, and acute renal failure. a non-contrast head ct showed left temporal lobe intraparenchymal hemorrhage. she was diagnosed with apml by peripheral smear, later confirmed by fish for t( : ), and was started immediately on high-risk induction chemotherapy as per cog protocol aaml , including all-trans retinoic acid, arsenic trioxide, idarubicin, and dexamethasone. cvvhd was required for acute renal failure. despite maximal respiratory support, she remained hypoxemic, with oxygenation index of , pao /fio ratio of . ecmo was initiated hours after start of induction, hours after admission. coagulopathy resolved on day of induction, ecmo was discontinued after days, mechanical ventilation and cvvhd were stopped after days and she continued to improve, eventually achieving remission with few neurologic side effects. despite relative contraindications to ecmo, this patient was successfully treated with ecmo without significant neurologic side effects. the correction of her coagulopathy was multifactorial: ) restoration of adequate oxygen delivery via ecmo improving endothelial function; ) successful organ support to allow sufficient response to induction chemotherapy with atra leading to the terminal differentiation of leukemic blasts; ) complement and contact system activation through contact with ecmo circuitry. this case illustrates that ecmo can still be considered in patients despite coagulopathy and end organ damage. sinai hospital of baltimore, baltimore, maryland, united states background: primary polycythemia vera is an extremely rare diagnosis in the pediatric patient and is defined by a marked elevation of red blood cells due to erythropoietin-independent mechanisms. presentations of this disorder range from the asymptomatic person to severe thrombotic events, such as budd-chiari syndrome or cerebrovascular stroke. mutations in the jak gene are found in adult and pediatric patients with polycythemia vera; however, the jak v f mutation is less commonly identified in pediatric patients. we describe an otherwise healthy -year-old female who presented with a significantly elevated total erythrocyte count, hemoglobin, and platelets, incidentally discovered upon routine annual blood work obtained by her pediatrician. design/method: this is a report and discussion of a rare case. demonstrated cellular marrow with trilineage hematopoiesis and no dysplasia. cytogenetics were not assessed. his hemoglobin and platelet count recovered but leukopenia and neutropenia persisted. follow-up evaluation at three months revealed fevers, ongoing cytopenias, a one-month of a nodular skin rash on the trunk and extremities resembling erythema nodosum, and hepatitis (peak alt and ast of , and , , respectively). following clinical evaluation, a skin biopsy was performed and was remarkable for atypical lymphocytes within the subcutis with t-cell markers, a high ki- , and positive tia- , perforin, and -f immunoperoxidase stains. negative stains for cd , cd , and ebv were noted. these results are consistent with sptcl. additional evaluation did not support a diagnosis of hlh. a staging evaluation was performed. pet-ct showed widespread hypermetabolic subcutaneous activity in the legs, trunk and skull and diffuse marrow hyperplasia. bone marrow demonstrated involvement with precursor b-cell acute lymphoblastic leukemia, with a mll gene rearranagement. his skin biopsy was retrospectively stained with tdt, cd , pax- , cd a, and cd with negative results, and a blood smear taken at the time of the skin biopsy did not demonstrate leukemic cells. conclusion: this is the first report of a patient with sptcl having a synchronous malignancy. the patient is doing well, currently in the maintenance phase of treatment for his all, and his skin disease has resolved on pet-ct. while it is possible that his presentation was a function of chance, the possibility of an underlying immune dysfunction or cancer predisposition warrants further investigation. cincinnati children's hospital medical center, cincinnati, ohio, united states background: hereditary xerocytosis (hx) is a rare red blood cell (rbc) dehydration disorder, characterized by variable hemolysis and propensity to iron overload. hx is often misdiagnosed as hereditary spherocytosis (hs). while splenectomy is curative for hs, it is relatively contraindicated in hx due to a substantial thromboembolism risk, signifying the importance of delineating these diseases. blood smear abnormalities are variable and often insufficient to make an accurate diagnosis. osmotic-gradient ektacytometry and genetic confirmation are critical in distinguishing these overlapping disorders. objectives: describe a family with hx, initially misdiagnosed as hs. discuss the importance of distinguishing these disorders and the utility of ektacytometry in making this distinction. design/method: a -year-old caucasian male was diagnosed with hs after presenting with prolonged neonatal jaundice starting on the first day of life. he described mild scleral icterus and history of intermittent jaundice and dark urine, without need for transfusions. his father, paternal uncle and paternal grandmother were all diagnosed with hs during childhood and underwent cholecystectomy. additionally, his father underwent splenectomy for abdominal pain. the child's blood counts revealed compensated anemia (hb . gm/dl) and reticulocytosis (arc × /mcl) with increased mcv ( . fl) and mchc ( . gm/dl). blood smear showed increased polychromasia and poikilocytosis with rare spherocytes and few stomatocytes. while the child had normal ferritin, his father had iron overload (ferritin ng/ml) despite no prior transfusions. osmotic-gradient ektacytometry profile of the child and father's rbcs showed a characteristic left-shifted, bell-shaped curve with decreased omin and ohyp, diagnostic of hx. the family is currently undergoing genetic studies. despite clinical similarities between hs and hx, distinguishing these diseases has significant management implications. hx is a disorder of rbc permeability, causing shortened rbc survival. stomatocytes on blood smear can raise suspicion for hx, but are insufficient to make an accurate diagnosis. identifying characteristic biomechanical membrane properties using osmotic-gradient ektacytometry is the gold standard for clinical diagnosis, which can then be confirmed by molecular studies. hs and hx can be easily and reliably distinguished using ektacytometry, as both disorders have very distinctive curves representing different rbc deformability patterns. after hx diagnosis was made, we counseled the family against splenectomy, as the risk of thromboembolism is significantly increased in hx compared to hs, and the father was diagnosed with iron overload. conclusion: hx is commonly misdiagnosed as hs. this case highlights the importance of making this distinction, and the utility of osmotic-gradient ektacytometry in reliably distinguishing these conditions. penn state health children's hospital, hershey, pennsylvania, united states background: relapsed acute myeloid leukemia (aml) presenting as an isolated central nervous system myeloid sarcoma (cns ms) is very rare and its treatment is not well-defined. thiotepa, vinorelbine, topotecan and clofarabine (tvtc) has been successful for re-induction therapy to induce remission prior to hematopoietic stem cell transplant (hsct). objectives: to describe our experience in utilizing tvtc therapy in two children with no extramedullary disease at initial diagnosis who presented with relapsed aml as intracranial myeloid sarcomas. results: case : month-old female was diagnosed with flt negative aml and completed treatment per the children's oncology group (cog) aaml study on the low risk arm without bortezomib. cerebral spinal fluid (csf) negative at diagnosis. fish testing positive for tcf gene deletion of unknown significance. mrd was undetectable after induction i and remained undetectable after each cycle. nine months off therapy, recurrent headaches prompted mri imaging which revealed two posterior fossa masses. csf and bone marrow testing were negative. stereotactic biopsy of the larger mass confirmed recurrence of aml. patient underwent two cycles of tvtc with a total of seven doses of intrathecal cytarabine with almost near resolution of the cns ms. completed cranial radiation and proceeded to allogeneic stem cell transplant with unrelated cord marrow donor and is disease free at approximately day + .case : year-old female diagnosed with flt and mll negative aml and completed treatment per cog aaml study on the low risk arm without bortezomib. csf negative at diagnosis. mrd was undetectable after induction i and completed therapy without complications. two months off therapy, a retrospective analysis of her diagnostic bone marrow by the cytogenetic laboratory to test a new panel identifying novel q partners revealed a cryptic insertional : (mllt /mll(kmt a) translocation. at four months off therapy, acute mental status changes prompted mri imaging which revealed two intracranial ms and lumbar spine involvement. resection of the larger lesion for symptomatic relief confirmed the mllt /mll(kmt a) fusion. csf positive for blasts and marrow negative for relapsed disease. patient completed two cycles of tvtc with a total of seven doses of it cytarabine with near resolution of cns disease (only mm contrast enhancement in the medulla). she received craniospinal radiation and is awaiting improvement in her cardiac function before proceeding to hsct. conclusion: tvtc is a successful reinduction regimen for relapsed aml with cns ms prior to hsct. background: acute severe anemia can be a life-threatening medical condition. the differential is quite broad for possible etiologies of acute severe anemia, including autoimmune hemolytic anemia (aiha) and atypical hemolytic uremic syndrome (ahus). autoimmune hemolytic anemia is an antibody-mediated process that targets the protein antigens located on the surface of red blood cells. treatment options for aiha include corticosteroids, with up to % of patients being responsive, with some requiring splenectomy. atypical hemolytic uremic syndrome is a medical urgency, defined as the triad of microangiopathic hemolytic anemia, thrombocytopenia, and acute kidney injury. the etiology is usually due to genetic causes, or less commonly, due to autoantibodies or idiopathic reasons. prognosis is very poor. objectives: differentiating between autoimmune hemolytic anemia and atypical hemolytic uremic syndrome can be a time-sensitive diagnostic dilemma while the patient is in critical condition, but this important delineation can vastly alter therapeutic options. design/method: here we discuss two cases highlighting the diagnostic workup involved in differentiating between atypical hemolytic uremic syndrome and autoimmune hemolytic anemia. patient a is a -year-old male who presented in extremis with severe anemia, uremic encephalopathy, and severe acute renal injury requiring hemodialysis and multiple blood transfusions. patient b is a -month-old male, who also presented in extremis with respiratory failure secondary to adenovirus/rhinovirus/enterovirus, with acute progressive renal failure and microangiopathic hemolytic anemia, requiring hemodialysis and cardiorespiratory support. : patient a underwent a full hematologic and infectious disease workup. subsequent laboratory studies confirmed enteropathogenic e.coli (epec) in the patient's stool; blood cultures remained negative. renal biopsy results were consistent pigment nephropathy. bloodwork indicated positive direct coombs. patient a was ultimately treated with steroids mg/kg/day, with significant improvement. patient b also included a full hematologic work-up, including adamts activity and ahus genetic panel, as well as full infectious disease work-up. subsequent laboratory test-ing revealed blood cultures growing streptococcus pneumoniae, with adamts activity at % (adult ref range: >/ = %), and normal complement levels. imaging findings also supported diagnosis of ahus. the management of a critically ill patient with acute severe anemia requires a thorough hematologic and infectious disease work-up. while molecular and genetic are helpful in definitive diagnosis of ahus, the utility of such results is limited by time. overlapping clinical presentation of a patient in extremis due to acute severe hemolytic anemia with progressive renal failure presents a rather broad differential, with time-sensitive treatment and prognostic implications. the favorable response to steroids delineates aiha from hus. background: d- -hydroxyglutaric aciduria (d- -hga) is a rare metabolic disorder characterized by developmental delay, hypotonia, and bi-allelic mutations in d- hydroxyglutarate dehydrogenase (d hgdh) or isocitrate dehydrogenase (idh ). metaphyseal chondromatosis with d- -hydroxyglutaric aciduria (mc-hga) is a type of d- -hga that has been previously reported in seven patients (omim ; pmid ), three of whom had somatic mosaicism for r variants in isocitrate dehydrogenase (idh ). we describe a -year-old boy with mc-hga who subsequently developed acute myeloid leukemia (aml) and was found to have a r variant in idh in a leukemic bone marrow sample. we report the first case of aml with this metabolic disorder. design/method: a -year-old hispanic boy presented with short stature, developmental delay, abnormal skin pigmentation, and unilateral congenital cataract. workup revealed multiple skeletal enchondromatosis and elevated urine d- -hydroxyglutaric acid levels. he was diagnosed with mc-hga. no pathogenic variants in d hgdh, idh and idh were identified in peripheral blood. germline testing with biopsies of skin lesions was declined by the family. two years later, he presented with streptococcal sepsis and pancytopenia. blasts were noted on peripheral smear. bone marrow morphology was consistent with acute myelomonocytic leukemia (∼ % blasts). chromosome analysis showed normal xy, and molecular testing by pyrosequencing idh and idh revealed a r c variant in idh ( % mosaicism). the patient is being treated as per the cog study aaml . end of induction i bone marrow aspirate was hemodiluted, but there was no obvious residual disease by flow cytometry ( . - . % sensitivity) or morphology. the previously identified idh variant was no longer detectable (limit of detection < %). although targeted therapy for aml with idh mutation is currently in phase i clinical trials in adults, there is no safety or efficacy data for using idh inhibitors in children. treatment with ivosidenib is therefore not currently an option for our patient. conclusion: this is the first case of aml reported with this rare metabolic disorder. somatic r variants in idh have been identified in three other mc-hga cases. this same mutation leads to the accumulation of d- -hydroxyglutarate in gliomas and aml. without any confirmed germline mutation or somatic mosaicism testing of multiple specimen sources, we can only speculate that the patient has an underlying somatic idh mutation associated with mc-hga which subsequently led to leukemogenesis. we present the first case of this association, to increase index of suspicion for development of aml in children with metabolic disorders associated with variants in idh . background: congenital combined deficiency of the vitamin k-dependent coagulating factors (vkcfd) is a rare heterogeneous autosomal recessive bleeding disorder. vkcfd is caused by mutations in the genes of either gamma-glutamyl carboxylase (ggcx) or vitamin k epoxide reductase complex (vkorc), which are responsible for the gammacarboxylation of vitamin k dependent proteins (vkdps) allowing for their activation. the clinical presentation ranges from no bleeding to intracranial hemorrhage. to date, vkcfd has been reported in few patients worldwide. objectives: we report a case of a girl with novel homozygous mutation of the ggcx gene, highlighting her clinical and biochemical characteristics with a review of the literature. a -month-old girl of consanguineous emirati parents, presented to our hospital with a history of bleeding from puncture site after receiving her second-month vaccine. that was associated with episodes of mild mucosal bleeding. review of systems was negative for jaundice, steatorrhea and failure to thrive and physical exam was unremarkable. investigations revealed markedly prolonged pt and aptt with high inr. fibrinogen, hemoglobin and platelets were always normal. activities of vitamin k-dependent factors including fii, fvii, fix, fx, protein c and s were all low. a measurement of proteins induced by vitamin k absence (pivka-ii) was done and came very high. this was associated with a mild elevation in liver enzymes but normal liver function test. the picture was supporting vitamin k deficiency, and as a result, she was started on oral vitamin k supplements of mg/day. she responded partially to vitamin k and required higher doses to stabilize her inr. after excluding acquired causes and due to her requirement of high doses of vitamin k, a mutation in either ggcx or vkorc genes was suspected. genetic analysis was conducted for her which revealed a novel missense homozygous mutation in the ggcx gene (c. a>t) confirming the diagnosis of combined deficiency of vitamin k-dependent clotting factors type . the asymptomatic parents were both heterozygous for the same mutation. results: she is currently stable on mg/day of vitamin k supplements. conclusion: vkcfd is a rare bleeding disorder with an overall good prognosis due to the availability of several effective therapeutic options. the function of the mutated gene is unknown. our patient demonstrated a partial response to vitamin k supplements suggesting presence of a residual carboxylation capacity and a possible role of this gene in the enzymesubstrate interactions. university of alabama at birmingham, birmingham, alabama, united states s of s background: gata is a zinc finger transcription factor that plays a critical role in the regulation of hematopoiesis and lymphatic angiogenesis. mutations leading to gata deficiency (gd) have been linked to a variety of clinical conditions. patients with gd have a striking predisposition to develop myelodysplastic syndrome (mds), acute myeloid leukemia (aml), or chronic myelomonocytic leukemia (cmml). acute lymphoblastic leukemia (all) has not been associated with gd, although the association of bcell all and gd has been previously reported. objectives: to describe a unique association of gata deficiency and t-cell all in a young child. results: an -year-old female presented with a one-week history of fever and malaise. she had a significant past medical history of verruca plantaris and self-resolving leukopenia associated with febrile illnesses. significant family history included sister with neutropenia and human papilloma virus (hpv) infection, and mother with neutropenia, monocytopenia, atypical mycobacterial infections, and hpv infection. peripheral blood revealed hemoglobin . g/dl, hematocrit . %, platelets , /ul, and white blood cell , /ul (neutrophils /ul, lymphocytes /ul, monocytes /ul). patient underwent a bone marrow biopsy demonstrating lymphoblast infiltration. flow cytometry analysis demonstrated monoclonal lymphoid blast population that co-expressed cd , cd , cd , nuclear tdt, cd , however, lacked expression of cd , cd , cd , cd , hla-dr, or myeloperoxidase. findings were consistent with tcell all with aberrant myeloid markers. cytogenetics analysis revealed ,xx,dic( ; )(p . ;p . ). patient began treatment as per children's oncology group aall and achieved remission at the end of induction. course of therapy was complicated by episodes of fever, reciprocating junctional tachycardia, asparaginase-associated thrombosis, viral meningitis, recurrent episodes of verruca plantaris, and resistant streptococcus pneumoniae or haemophilus parainfluenza infections causing chronic cough. later, she was also found to have low igm levels; after completion of therapy, she developed monocytopenia. lymphocyte subset panel revealed absent b cells, decreased number of natural killer (nk) cells, and cd /cd inversion. further work-up included gata sequence analysis that showed heterozygous nonsense mutation (c. c > t/c; reference nm_ ) likely resulting in gata haploinsufficiency. patient continues to be in remission, is receiving monthly immunoglobulin replacement and is on azithromycin for atypical mycobacterial prophylaxis. surveillance bone marrow biopsies have shown no evidence of mds or leukemia, however, have demonstrated persistent hypocellularity. the possibility of undergoing an allogeneic bone marrow transplant is actively being discussed given its curative potential. clinicians should be aware that t-cell all may be associated with gata deficiency. cincinnati children's hospital medical center, cincinnati, ohio, united states background: treatment for severe hemophilia a is centered on factor viii (fviii) replacement therapy. development of an alloantibody (inhibitor) against fviii is a significant treatment complication occurring in as many as - % of patients. high titer inhibitors render treatment with factor viii ineffective, necessitating the use of bypass agents that may not achieve hemostasis with the same efficacy. considering the substantial ramifications of inhibitor development on treatment, eradication of inhibitors is of great importance to achieve adequate hemostasis in this patient population. desensitization by immune tolerance induction (iti) is the primary method of inhibitor elimination. however, not all patients respond to iti. immunomodulation may be considered as the next line of therapy, although controversy remains in regards to agent selection and use. objectives: there is incomplete data on the use of immunomodulation therapy for inhibitor eradication in severe hemophilia a. we present a case of a pediatric patient with severe hemophilia a and high titer inhibitor who failed initial iti therapy to better illustrate potential treatment options for the future. design/method: a retrospective chart review was performed on a patient with severe hemophilia a at cincinnati children's hospital medical center. results: an -year-old caucasian male with severe hemophilia a secondary to intron inversion, was initially diagnosed following extensive bleeding after circumcision at birth. he was identified as having an inhibitor ( bethesda units (bu)) at months of age after exposure days of treatment. he failed multiple attempts of iti, with recombinant and plasma-derived (pd) fviii. he was advanced to immunomodulation therapy in combination with pdfviii, however demonstrated anaphylaxis to rituximab and ofatumumab. he underwent tolerization to rituximab, and received a six month course with a partial response (nadir of . bu). months following last dose of rituximab, a rising inhibitor titer ( . bu) was found. mycophenolate mofetil (mmf) was initiated with subsequent inhibitor stabilization and a decreasing titer ( . bu) over the course of the following year. mmf has been well tolerated without major side effects or infection throughout therapy. conclusion: development of an inhibitor against fviii is a considerable complication in patients with severe hemophilia a. use of immunomodulatory therapies following iti failure remains controversial. mmf has not been well studied in this patient population. we report a case of a patient who is being successfully treated with mmf with minimal side effects. further prospective studies should be considered to further define the role of mmf immunomodulation therapy. background: down syndrome (ds) children with aml (ds aml) have higher cure rates than their non-ds counterparts. outcomes for refractory/relapsed cases, however, remain dismal. somatic mutations of the gene encoding the transcription factor gata in ds aml patients are responsible for the observed hypersensitivity of ds aml blasts to cytosine arabinoside (ara-c). in view of excellent survival rates (approaching %) of ds aml patients, the ongoing children's oncology group (cog) aaml study seeks to determine the feasibility of treating standard risk (minimal residual disease/mrd negative) ds aml patients using a reduced dose ( -fold decrease) ara-c backbone. although results from japanese trials with this approach are promising, north american and european data are conflicting. although chromosome rearrangements in ds aml do not appear to carry the same adverse prognostic significance as in non-ds aml, monosomy in ds aml patients has been associated with a moderately worse outcome. isochromosome q, however, is rare and has only been reported in previous cases of ds aml. objectives: to report our institutional experience of very early relapse involving cases of ds aml patients treated per the reduced dose ara-c arm ( . g/m ) of the aaml study. design/method: we hereby report the disease course and cytogenetics of the above ds aml patients. : patient is a month old caucasian female who had gata mutation negative aml. patient is a -year old caucasian male whose chromosomal analysis revealed isochromosome q ( copies of the long arm of chromosome ). both patients achieved negative mrd (< . %) after induction i chemotherapy with thioguanine, low-dose ara-c and daunorubicin and proceeded per the reduced dose ara-c arm of aaml . patient relapsed immediately after completion of chemotherapy. salvage chemotherapy with mitoxantrone/high dose ara-c (hidac) failed to induce a second remission and the patient subsequently died of disease. patient relapsed within months from end of therapy. the patient underwent salvage chemotherapy utilizing a hidac backbone and remains in disease remission. the noted very early relapse following a reduced dose ara-c regimen in our above ds aml children suggests that testing for gata mutation and chromosome rearrangements may play a useful role in the development of future risk-stratified treatment strategies for ds aml. university of rochester, rochester, new york, united states background: in developed countries in the st century, severe nutritional deficiency is not an often considered differential diagnosis of unexplained childhood anemia. aside from iron deficiency anemia, vitamin deficiency severe enough to impact hematopoiesis is uncommon in the general pediatric population. here we present the unique case of a -monthold infant who presented with intermittent emesis, failure to thrive (ftt), developmental delay, macrocytic anemia, and neutropenia which was initially concerning for a congenital bone marrow failure syndrome. instead, she was discovered to have an underlying, potentially familial deficiency of b . objectives: . to describe the unique case of an infant with b deficiency. . to outline the importance of including b deficiency in the differential diagnosis of unexplained megaloblastic anemia in children. a -month-old exclusively breastfed infant presented for gastroenterology evaluation due to persistent emesis and poor weight gain over the course of months. her history was notable for delayed developmental s of s milestones and hypoactivity. marked pallor prompted hematologic evaluation, which revealed concern for macrocytic anemia (hemoglobin . g/dl, mcv ), reticulocytopenia ( . × ^ / l), and neutropenia (anc . × ^ /l). an otherwise reassuring physical examination and laboratory evaluation was notable only for the discovery of an undetectable b level and marked hyperhomocysteinemia ( mol/l). her hemoglobin (hgb) continued to decline (to . g/dl) over the first few days after presentation, and she required red blood cell (rbc) transfusion. within only a few days of initiation, daily cyanocobalamin injections resulted in a robust reticulocytosis response, improved hgb, immediate normalization in the neutrophil count, and resolution of hyperhomocysteinemia. additional history and laboratory evaluation from the patient's mother revealed a concurrent, asymptomatic maternal b deficiency as well as a history of a need for b supplementation in the maternal grandfather, raising concern for an inherited etiology. despite the rarity of vitamin-deficient hematologic abnormalities in the general pediatric population, b deficiency should be considered as a potential cause of an otherwise unexplained megaloblastic anemia, especially in the setting of concurrent ftt and neurodevelopmental delay. a detailed family history should be obtained in such cases and may have helped to prevent this patient's clinical sequelae had the deficiency been discovered sooner. our patient has experienced a favorable clinical response to b supplementation, attesting to the importance of vitamin b in early childhood growth and development. background: peg-asparaginase is universally utilized in the treatment of pediatric acute lymphoblastic leukemia (all). despite its high efficacy in this disease, it is associated with hypersensitivity and allergy in - % of patients. protracted anaphylaxis has been described in circumstances such as severe food allergy with ongoing allergen exposure; however, it has not yet been described in relation to peg-asparaginase. we describe the first reported case of protracted anaphylaxis after peg-asparaginase administration, provide guidance as to time course and management of protracted anaphylaxis, as well as evidence that erwinia asparaginase may be safely administered even in this high risk population. objectives: to provide guidance regarding the duration, course and management of protracted, severe anaphylaxis after peg-asparaginase therapy. a year old male with very high risk all presented for consolidation therapy with peg-asparaginase (intramuscular) and vincristine. one hour after administration, he developed generalized hives and angioedema, for which he was given diphenhydramine. he then quickly developed progressive hives, angioedema, subjective throat and chest tightness, and wheezing. he was treated with diphenhydramine, epinephrine, albuterol, and methylprednisolone with resolution of symptoms. one hour later, symptoms recurred and the patient became hypotensive; he was retreated with methylprednisolone and epinephrine, and was transferred to the pediatric intensive care unit (picu). in the picu, he was placed on an epinephrine drip, and continued on methylprednisolone, diphenhydramine, cetirizine, albuterol, and ranitidine. the epinephrine drip was successfully discontinued after hours, and his other medications were gradually weaned over the course of two weeks. of note, the patient did have st segment changes in his electrocardiogram during the first hours of anaphylaxis. these were associated with normal ventricular function as per echocardiogram, and resolved within one week. this patient has subsequently tolerated multiple doses of erwinia asparaginase (intramuscular) without premedication. this patient was acutely managed in the pediatric intensive care unit with steroids, anti-histamines, and continuous infusion epinephrine. symptoms consistent with severe anaphylaxis including hives, angioedema, throat and chest tightness, wheezing, and hypotension persisted for a total of four days before finally resolving. he has thus far tolerated multiple doses of erwinia asparaginase without any symptoms of allergy, hypersensitivity, or anaphylaxis. protracted severe anaphylaxis after peg-asparaginase therapy can be successfully managed with multi-agent therapy, including antihistamines, steroids, and continuous infusion epinephrine. re-challenge with an alternate form of asparaginase may be tolerated, even in a patient with protracted anaphylaxis to peg-asparaginase. ucsf benioff children's hospital oakland, oakland, california, united states background: vincristine (vcr) is widely used in pediatric cancers. unlike most cytotoxic agents, hematopoietic toxicity is uncommon. vcr-induced anemia has been observed but its mechanism has not been well studied. vinca alkaloid-induced membrane changes were seen in early studies of hereditary spherocytosis (hs) and anecdotal cases suggest vcr may increase hemolysis in such patients. here we describe a case involving severe vcr-induced anemia in a patient with hs and an explanation as to the mechanism. objectives: to describe the mechanism of vcr-induced anemia in hs. design/method: case report. a year-old female with hs was diagnosed with t-lymphoblastic lymphoma. she had required packed red blood cell (prbc) transfusions as a neonate and thereafter had done well without episodes of acute hemolysis or aplasia. complete blood counts (cbc's) demonstrated a compensated hemolysis, and she did not require further transfusions until she commenced chemotherapy. by the start of maintenance she had received many more prbc transfusions than the average patient. intermittent drops in hemoglobin (hb) did not correlate with any particular agent, and she had stable, mild splenomegaly. a clear pattern emerged during maintenance. her hb was - g/dl at monthly clinic visits, when she received vcr, intermittent intrathecal methotrexate, and corticosteroids. within - days, her hb dropped to . ± . g/dl, and reticulocyte count decreased from . to . ± . %. transfusion at day corrected hb, and the reticulocytes and hb returned to baseline. white blood cell and platelet counts did not change after vcr. blood samples from pre, immediately post, and days post vcr were analyzed and rbc characteristics and markers of hemolysis were not significantly different. ektacytometry showed identical curves, indicating no change in rbc deformability. in vitro incubation of patient blood samples with vcr also did not affect the osmotic deformability, confirming that a change in rbc rigidity was unlikely the reason for the drop in hb. these data indicate that a dysregulation of erythropoiesis was responsible for the anemia after vcr, rather than damage of peripheral rbc's. in most patients, maintenance therapy for lymphoblastic lymphoma does not cause severe anemia, likely because a temporary reduction in erythropoiesis in patients with a normal rbc survival and low reticulocyte count is not noticed. however, in a patient with decreased rbc survival and a brisk reticulocytosis, a disruption in rbc generation is more apparent. in conclusion, vcr administration to patients with an rbc disorder warrants close observation for potentially severe vcr-induced anemia. background: the addition of tyrosine kinase inhibitors (tki) to conventional chemotherapy has improved outcomes for pediatric patients with philadelphia chromosome-positive (ph+) acute lymphoblastic leukemia (all), however there remains an increased risk of relapse compared to other types of childhood all. typically, in relapsed disease the philadelphia chromosome persists and several mechanisms of resistance involving acquired mutations of the bcr-abl chimeric oncoprotein have been reported. objectives: describe a unique case of a pediatric patient with ph+ b-precursor all relapsing with b-precursor all without the philadelphia chromosome. results: an -year-old boy was diagnosed with ph+ bprecursor all with the presence of the t( ; )/bcr-abl translocation by cytogenetics and fluorescence in situ hybridization (fish), respectively. additional abnormalities included gains of runx and loss of one copy of etv . a remission bone marrow with negative minimal residual disease (mrd) was achieved at the end of induction with dasatinib and the esphall chemotherapy backbone. duration of tki therapy was two years post diagnosis. nearly one year after the completion of therapy, cytopenias prompted a bone marrow investigation. relapsed b-precursor all was established by immunophenotyping, however fish analysis did not identify the bcr-abl rearrangement. moreover, quantitative reverse transcriptase pcr was negative for the bcr-abl fusion transcript. again fish analysis of the bone marrow revealed multiple additional copies of runx and mono-allelic loss of etv , similar to the initial diagnostic sample. the patient was re-induced per aall anticipating a ph+ all relapse. however, with confirmation of the loss of the ph+ clone, tki therapy was not re-initiated. due to positive mrd of . % at the end of re-induction therapy, the patient was salvaged with blinatumomab therapy and subsequently underwent an allogenic stem cell transplant with a sibling donor. conclusion: this is the first known report of a pediatric patient with ph+ b-precursor all who developed recurrent b-precursor all without the philadelphia chromosome. the persistent findings of gain of runx and loss of etv makes it unlikely that a second unrelated b-precursor all developed following successful treatment of the original disease. this case highlights the possibility of a genetically distinct subclone present at the onset of disease that shared abnormalities of runx and etv but did not contain the philadelphia chromosome. nevertheless, the subclone harbored leukemogenic potential in the absence bcr-abl expression. it is plausible that the predominant clone present at diagnosis was effectively treated with dasatinib and extinguished, but the bcr-abl -negative clone persisted in the face of tki therapy. background: ligneous conjunctivitis is a rare form of pseudomembranous conjunctivitis that develops specifically in patients with type plasminogen deficiency. lack of plasmin activity in those patients result in defective fibrinolysis and formation of fibrin-rich membranous material/ masses that develops on the palpebral conjunctiva as well as other sites in the body.current management involve surgical excision of the masses that is usually complicated by multiple recurrences. recently, use of topical plasminogen concentrates helped delaying recurrence, but currently, those concentrates are not commercially available. we report on a -year-old omani girl, with hypoplasminogenemia who required optimization of plasminogen level at the time of surgery to delay/ prevent recurrence. objectives: case report on the peri-operative use of ffp versus cryopricipitate transfusion as an alternative replacement of plasminogen during surgical excision of ligneous conjunctivitis. design/method: pharmacokinetic study was performed to assess plasminogen recovery after ffp ( ml/kg) and precipitate ( bag/ kg) transfusion results: plasminogen levels remained subnormal after either ffp or cryoprecipitate administration. with ffp, the maximum concentration reached was almost % of normal. although half-life of plasminogen is known to be - . days, the patient seemed to have a high catabolic rate after receiv-ing cryoprecipitate, with plasminogen levels reaching basal levels within hours. because of the better recovery profile with ffp, we opted to give ffp before and after surgery. peri-operative management included ffp transfusion at ml/kg/ hours one day before and for days post operatively, followed by ml/kg once daily from day - , then ml/kg on th post-operative day. topical treatment was initiated using antibiotic and steroids ed on the day of surgery, followed by heparin ed on the second day. on follow up, she used topical heparin, cyclosporine, prednisolone, and topical lubricant eye drops for variable duration. clinical picture remained stable for almost year post operatively, when she started to develop recurrence of ligneous lesions again. background: ponatinib (inclusig®, ariad pharmaceutical) is a rd generation multi-targeted tyrosine kinase inhibitor (tki) approved for treatment of adults with chronic myeloid leukemia (cml) and philadelphia chromosomepositive acute lymphoblastic leukemia (ph+ all) resistant to or intolerant of other tkis. ponatinib has numerous drug-drug interactions and a black box warning for associated serious adverse vascular events and hepatotoxicity. for this reason, ponatinib use has been confined to specific high-risk populations. however, in patients who prove refractory to other therapies, the potential benefits of ponatinib may outweigh risks. to date, ponatinib has not been studied in the pediatric/adolescent and young adult (aya) population. furthermore, literature describing the use of ponatinib alone or in combination with other agents in pediatric oncology patients is scarce. objectives: to describe a single institutional experience using ponatinib in the pediatric patients with ph+ all. design/method: two cases of ponatinib use in pediatric ph+ patients resistant to other tkis were identified at our institution and are described. peripheral blood samples obtained from both patients identified bcr-abl p fusion transcripts and sanger sequencing was used to identify resistant mutations. results: our first case is a -year-old female who received upfront multi-agent chemotherapy plus dasatinib for ph+ all. relapse was confirmed on end-of-therapy bone marrow evaluation, thus bcr-abl mutation testing was performed and revealed a t i mutation. ponatinib was initiated then discontinued after one week due to clinically significant fluid retention with peripheral edema and bilateral pleural/pericardial effusions. the second case is a lateadolescent female with ph+ all who relapsed -years after stem cell transplant (sct). following relapse, tki therapy included both imatinib and dasatinib. due to persistence of bcr-abl fusion transcript despite tki therapy she was switched to ponatinib. shortly following initiation of ponatinib she developed a diffuse, maculopapular rash, which persisted despite dose reduction, resulting in ultimate discontinuation of the drug. bcr-abl mutation testing identified f l and f v resistance-conferring mutations. to date, there is scant existing literature detailing the use of ponatinib in pediatric patients. appropriate dosing is undefined and side effect profile not well described, particularly when used concurrently with other chemotherapeutic agents. thus, this case series reporting the response to and toxicity of ponatinib in pediatric ph+ all patients has important clinical implications. additionally, this is the first report of a pediatric ph+ all patient with documented t i mutation underscoring the importance of bcr-abl mutational testing, particularly at the time of relapse. cooper university hospital, camden, new jersey, united states background: myh -related disorder is a rare autosomal dominant disease, encompassing several subtypes: may hegglin anomaly, epstein syndrome, fechtner's syndrome, and sebastian syndrome. heterozygous mutations are seen in the gene encoding non-muscle myosin heavy chain iia (nmmhc-iia) which is involved in cell motility as well as functions to maintain cellular shape and integrity. the presentation of myh -rd is mainly characterized by macrothrombocytopenia, but various related expressions exist: nephritis often leading to renal failure, cataracts and sensorineural deafness ( ). a -year-old girl with history of extensive dental caries, hyperactivity, and speech delay due to suspected hearing loss was incidentally found to have thrombocytopenia at the time of genetic evaluation. she did not have any bruising or excessive bleeding. she did not respond to observation, immunoglobulins, or steroid therapy. her platelet count remained persistently low ( - k/ul). she underwent extensive evaluation to rule out platelet disorder vs. coagulation defect. her peripheral smear showed enlarged platelets by giemsa stain but no inclusion bodies were noted in granulocytes. her platelet aggregation and platelet surface glycoprotein by flow cytometry were negative. her coagulation profile was also normal. objectives: this case report summarizes the complexity in diagnosing myh -rd in a pediatric patient. design/method: since a unifying diagnosis for her clinical presentation was not apparent, whole exome sequencing (wes) was undertaken. results: wes revealed the r c heterozygous pathogenic variant, located in exon in the myh gene. myh gene alteration explained the patient's clinical features of macrothrombocytopenia and hearing loss. this mutation was paternally inherited, and her father demonstrates mosaicism. he was asymptomatic with normal platelet count but his morphology showed enlarged platelets with no inclusion bodies in granulocytes. when dealing with patients who have mild or no symptoms of bleeding diathesis but evidence of persistent macrothrombocytopenia, considering a platelet disorder belonging to myh -rd can help delineate certain predisposing syndromes and guide clinical management. patients are likely to benefit from early genetic testing while receiving supportive therapy. wes can highlight syndromes and provide information on recurrence risk for families. the renal and hearing abnormalities are indistinguishable between epstein and fechtner's syndromes, but the pathogenic variants differ ( ). the genotype-phenotype correlation implies that our patient may have either syndrome, although clinical features compatible with nephritis have yet to manifest. patients should be monitored closely for long-term progression of myh disease, and treatments should be initiated accordingly. we present an -year old female evaluated by genetics at birth due to prenatal microcephaly. chromosomes and microarray were normal. at age she developed standard risk pre-b-cell acute lymphoblastic leukemia (all). she completed treatment in and has been doing well in the interim, remaining in complete clinical remission. during and after treatment she exhibited developmental delay and neurocognitive deficits. at age her height and weight were at or below the th centile and head circumference was below the nd centile (approximately standard deviations below the mean and corresponding to the th centile for a -month-old girl). bone age was appropriate. she had a distinctive triangular face with micrognathia and a pointed nose resembling a seckel-like syndrome. the patient also had clinodactyly of the th toes, zygodactylous triradius involving the nd and rd left toes, tendency to sydney line in the right palm and a radial loop in the left middle finger. the patient's unique clinical presentation prompted a more thorough genetic evaluation, which led to a novel finding we feel is clinically significant with regard to the development of malignancy. design/method: whole exome sequencing (wes) was performed on the patient as well as her biological parents (trio). a de novo heterozygous mutation in the gene pcdh with potential relation to the phenotype was discovered. this c. dupa variant causes a frameshift starting with codon asparagine , changing this amino acid to a lysine residue and creating a premature stop codon at position of the new reading frame denoted p.asn lysfsx . this variant is predicted to cause loss of normal protein function via protein truncation or nonsense-mediated mrna decay. conclusion: pcdh is a member of the protocadherins family which is important in cell-to-cell adhesion and synaptic function in the central nervous system and is highly expressed in areas of the brain involved in higher cortical function and speech. aberrant expression of protocadherins has been associated with the development of malignancies in many organ systems. with regards to leukemia, the methylation status of this gene at diagnosis has been implicated in the prognosis of all and could be used as a biomarker to predict relapse. this patient's de novo mutation and clinical presentation are unique to what has been previously presented in the literature. we feel that this mutation is a clinically significant finding that may shed light on the role of this gene in the development of hematopoeitic malignancies. background: acquired hemophilia a (aha) is an uncommon and potentially life-threatening hemorrhagic disease characterized by sudden onset of bleeding in patients with neither personal nor family history of bleeding dyscrasia. it is usually seen in adults with autoimmune diseases, solid tumors, lymphoproliferative diseases, pregnancy or during the postpartum period; occurrence in the pediatric population has rarely been reported. we report a case of an otherwise healthy teenager who was found to have aha when he presented with acute onset of atraumatic soft tissue hematoma. results: a -year old male of middle eastern descent with history of congenital absence of the right external ear, but otherwise in good general health, presented to our emergency department with a three day history of progressive worsening of right lower leg pain, swelling, and paresthesia, without preceding history of trauma. evaluation by the pediatric orthopedics service documented significantly elevated compartment pressures, necessitating immediate four-compartment fasciotomy. pre-operative labs were significant for prolonged activated partial thromboplastin time (aptt) of . ( . - . ) seconds with normal prothrombin time (pt) and international normalized ratio (inr). ptt did not correct on mixing studies, suggesting the presence of a circulating anticoagulant. factors xii and xi were in the normal range; factor ix was elevated, ( - ). factor viii level was % and fviii inhibitor level was . bethesda units (< . ), confirming the diagnosis of aha. work up for autoimmune disease was negative. his bleeding and surgical hemostasis were managed with recombinant factor vii (novoseven) mcg/kg every hours for hours post operatively, with gradual interval prolongation. factor viii antibody eradication was managed with prednisone mg/kg/day. factor viii and inhibitor levels normalized by day of hospitalization. recombinant factor vii was discontinued; steroids were gradually tapered and discontinued at discharge (hospital day ). conclusion: acquired hemophilia is likely an underdiagnosed condition in pediatrics. while it is typically seen in adults with underlying autoimmune disease, solid tumors, lymphoproliferative disease, or during pregnancy or the postpartum period, pediatric cases may have no identifiable etiology. this case highlights the importance of considering this diagnosis in any patient with unexplained bleeding regardless of their age, so as to intervene early and prevent adverse consequences. university of oklahoma, oklahoma city, oklahoma, united states background: myeloid neoplasms associated with eosinophilia is a rare subtype of chronic leukemia characterized by clonal eosinophilia. the true incidence is unknown due to its rarity and possible classification as idiopathic hypereosinophilia syndrome. the most common chromosomal aberrations involve platelet-derived growth factor receptors (pdgfrs). we report one such rare case in a pediatric patient. most of the pediatric management of this entity is derived from adult case reports and case series. objectives: to describe a case of chronic leukemia presenting as eosinophilia results: a previously healthy year old caucasian male presented with a several week history of migrating joint pain, splenomegaly, and abnormal blood counts with leukocytosis, thrombocytopenia and absolute eosinophilia. white blood cell differential showed myeloid precursors suggestive of chronic myeloid leukemia. bone marrow evaluation showed % blasts and % eosinophils. bcr-abl testing was negative, ruling out cml. fish analysis for eosinophilic clonality revealed deletion of chic gene, resulting in fip l /pdgfra fusion gene, diagnostic for myeloid neoplasm with eosinophilia associated with pdgfr abnormalities. treatment was started with tyrosine kinase inhibitor (tki), imatinib mg daily. within months, fish analysis for fusion gene was negative. after approximately months of daily imatinib, he was switched to maintenance dose of mg weekly. he is approximately months since diagnosis and doing well on maintenance imatinib. in , the who revised its classification of some chronic eosinophilic leukemias to myeloid and lymphoid neoplasms associated with eosinophilia and rearrangement of pdgfra, pdgfrb, fgfr . the most common abnormality is the fip l /pdgfra fusion gene. other less common abnormalities include fusion genes kif b-pdgfra and etv -pdgfrb and point mutations in pdgfra . some features of chronic eosinophilic leukemia include absolute eosinophilia, splenomegaly, elevated vitamin b and tryptase levels, and organ damage from eosinophil infiltrates and cytokine release. patients with rearrangements or mutations involving pdgfra are usually very responsive to imatinib. starting doses have not been well studied or established. experts recommend co-administration of corticosteroids during the first few days of imatinib therapy in patients with a history of cardiac involvement and/or elevated serum troponin levels to prevent myocardial necrosis, a rare complication of imatinib therapy in eosinophilic patients. fortunately our patient did not have cardiac involvement and to date has not exhibited signs of chronic tki toxicity. conclusion: myeloid neoplasms with eosinophilia constitute a rare form of chronic leukemias. they are often associated with pdgfr abnormalities and are usually very responsive to tyrosine kinase inhibitor therapy. walter reed national military medical center, bethesda, maryland, united states background: germline samd l mutation is a rare cause of constitutional bone marrow failure with a unique propensity for clonal evolution to monosomy and mds. objectives: previous case series have demonstrated diverse clinical outcomes in patients with a germline samd l mutation. our case presents a novel samd l mutation (p.val leu). additionally, the case highlights the challenges in clinical decision making for a patient with a gene mutation that is known for clonal evolution towards monosomy with risk of progression to myeloid malignancy, but also known for self-correction through uniparental disomy or inactivating mutations which results in disease remission. design/method: a retrospective chart review and review of the literature was performed. dna was isolated from peripheral blood and used for whole exome sequencing. a peripheral blood sample from the patient's mother and father showed no samd l mutation. skin biopsies of the patient and parents were evaluated for uniparental disomy or new mutations. to determine the pathogenicity of this novel mutation, the specific samd l mutant dna was transfected into the human embryonic kidney cell line to assess its role in inhibiting cell proliferation. our patient presented at months of age with pancytopenia and hypocellular bone marrow in the setting of s of s sepsis. he had evidence of dysfunctional immune activation with hemophagocytosis and elevated soluble il with simultaneous severe hypogammaglobulinemia. analysis of the peripheral blood showed no increase in chromosomal breakage, normal telomere length, and normal flow cytometry. gene testing for primary hemophagocytic lymphohistiocytosis and inherited bone marrow failure were negative. after the patient recovered from his presenting illness, a repeat bone marrow biopsy demonstrated improved cellularity with myelodysplasia and cytogenetics significant for monsomy .whole exome testing demonstrated a novel samd l mutation. the patient continued to require intermittent ivig and failed to demonstrate appropriate leukocytosis with intermittent infections. on repeat bone marrow evaluation over the course of months, the patient demonstrated no evidence of evolution towards self-correction and had a persistent monosomy clone. the patient is scheduled to undergo a matched unrelated donor bone marrow transplant. our case highlights the unique clinical picture associated with constitutional marrow failure and clonal evolution secondary to a novel samd l mutation which is thought to cause pancytopenia by inhibiting cellular proliferation and often results in the development of monosomy which rescues hematopoiesis but with a risk for malignancy. background: notable labs developed a flow cytometricbased assay with a custom robotic platform to test fdaapproved drugs for anti-cancer activity against individual patient's tumor cells. this personalized assay is a potential method for identifying novel agents and drug combinations to treat aml patients who have failed standard therapies. objectives: to present the case of a teen who underwent successful treatment of relapsed aml post-sct with bortezomib, panobinostat, and dexamethasone-a regimen selected based upon results of notable lab testing. results: a -year-old male with m -aml had an isolated bone marrow relapse months after completion of scheduled therapy. at relapse, his aml was flt -itd positive. he achieved a second remission with negative mrd and underwent matched sibling donor bmt after busulfan/cyclophosphamide conditioning. bma performed on day + was mrd positive ( . %). repeat bma done on day + showed . % mrd. he started sorafenib on day + . he received donor lymphocyte infusion (dli) on day + , then received cycles of azacitadine (aza) followed by dli. marrow mrd by flow after sorafenib alone, sorafenib with dli, and sorafenib with aza/dli were %, . %, and . %, respectively. treatment was complicated by varicella meningitis, grade i skin agvhd, febrile neutropenia and c. difficile colitis, and metapneumovirus pneumonia. despite extremely low levels of leukemia (marrow mrd . %), notable lab testing performed on the patient's leukemia cells from marrow collected after aza/dli/sorafenib revealed sensitivity of his leukemic blasts to a combination of bortezomib, panobinostat, and dexamethasone. because of prolonged cytopenias, multiple infectious complications, and persistently positive mrd, he discontinued aza/dli/sorafenib and on day + started bortezomib . mg/m iv on days , , , and ; panobinostat mg po on days , , , , , ; and dexamethasone mg po on days , , , , , , , and . chemotherapy cycle started days later. he tolerated treatment without side effects and with resolution of rash and cytopenias. he achieved full donor chimerism, negative flt -itd, and complete remission by morphology and flow after two cycles. notable lab testing is a powerful tool for evaluating the sensitivity of small populations of leukemic blasts to novel drug therapy. results from notable lab testing may serve as a useful guide for treatment selection after failure of standard aml therapy. this patient achieved morphologic and mrd remission post-sct with bortezomib, panobinostat, and dexamethasone-a regimen predicted to be efficacious based upon notable lab results. maria ahmad-nabi, christine knoll, sanjay shah, esteban gomez, lori wagner phoenix children's hospital, phoenix, arizona, united states background: development of inhibitors in patients with factor ix deficiency (fixd) is a well-recognized complication occurring in - % of patients. within this subset a small percentage can develop anaphylaxis to factor. desensitization with cyclophosphamide, an alkylating agent used in the management of various oncologic malignancies, and reported for use in factor viii desensitization has been previously unreported for use in desensitization in patients with fixd. rituximab, an anti-cd antibody, however has been used. objectives: to induce immune tolerance (it) in patients with inhibitors to factor ix with either novel or under reported methods using cyclophosphamide and/or rituximab. we report a case series of patients at phoenix children's hospital with fixd who achieved it with cyclophosphamide and/or rituximab. results: patient one was a year old male with severe fixd, who at the time of desensitization had inhibitor levels of bu. he was desensitized with cyclophosphamide, then admitted for infusion of recombinant factor ix. he experienced a few minor symptoms of intolerance including an urticarial rash which was self-limited, and hemarthrosis of the right elbow on day which responded to novo . he tolerated the remainder of his infusion without issues. he continued recombinant factor ix daily, and returned to clinic for monthly cyclophosphamide for months. he did develop urticaria with hemarthrosis and spontaneous muscle bleeds which were tempered with zantac, zyrtec, solumedrol, and benadryl. he remained without a recurrence of inhibitors, however did have intermittent hemarthrosis of his ankles thereafter requiring prophylactic twice daily dosing recombinant factor ix. patient two was a year old male with severe fixd and a family history of anaphylaxis to factor causing early death in all male relatives with the disease. he had never received factor ix and did not have a detectable inhibitor prior to desensitization. he successfully underwent desensitization to recombinant factor ix with rituximab in the icu, and returned to clinic for weekly infusions x . he experienced no adverse reactions concerning for anaphylaxis. he continued to tolerate factor ix products without evidence of intolerance, development of inhibitors, and continues on as prophylactic dosing of recombinant factor ix every other day. our experience at a single institution proves cyclophosphamide as a novel agent for inducing it in those with fixd and anaphylaxis. it also provides further evidence that rituximab can desensitize patients with severe fixd. differences include longer duration for cyclophosphamide therapy ( months vs month). background: cartilage-hair hypoplasia (chh) is an autosomal recessive chondrodysplasia associated with defective cell-mediated immunity caused by mutations in the ribonuclease mitochondrial rna processing (rmrp) gene. cancer incidence is -fold higher in patients with chh than in the general population, especially non-hodgkin lymphoma. the use of rituximab, an anti cd antibody, results in decreased host b-cell number and impaired humoral function for - months. the safety of rituximab in pediatric patients with cancer and immunodeficiency is not well documented. a diagnosis of underlying immunodeficiency may discourage physicians from using rituximab due to the risk of severe bacterial infection or viral re-activation. objectives: to report a case of burkitt lymphoma in a young adult female with chh and defective cellular immunity successfully treated with rituximab. results: an -year old amish female with disproportionate short stature presented to our center for management of stage iv biopsy proven burkitt lymphoma with myc rearrangement. she had presented a week earlier with cervical, occipital, and submandibular lymphadenopathy, splenomegaly; fevers, night sweats, and weight loss for - weeks. on exam, her height was three feet associated with brachydactyly, mild bowing of the legs, normal size head without frontal bossing, fine and sparse hair. she had normal intelligence. her pattern of dysmorphisms was suggestive of chh (genetic testing not performed at time of diagnosis). pet-ct scan showed stage iv disease with involvement of cervical lymph nodes, spleen, iliac bone and bone marrow. treatment with standardintensity fab/lmb therapy (group c) with the addition of rituximab was initiated. she had an incomplete response to cop (∼ % reduction of tumoral masses) but achieved complete remission after copadam . her course was complicated with severe varicella zoster but she completed therapy and remains in complete disease remission for months after treatment completion. genetic testing subsequently performed proved homozygosity for chh with a n. a>g variant. she had no other opportunistic infections during or after therapy. conclusion: the use of rituximab was both safe and beneficial in our patient despite defective cell mediated immunity secondary to chh suggesting that rituximab may be safe to use in patients with cellular immune deficiencies. background: hemophilia a and b are bleeding disorders characterized by deficiency in factor viii or ix, respectively. spontaneous or provoked hemarthrosis is a known complication of hemophilia. repetitive episodes of hemarthrosis can lead to debilitating hemophilic arthropathy. lyme disease is a tick-born infection which is endemic to increasing parts of the united states. chronic lyme disease, the phase in which lyme arthritis typically develops, occurs months to years after initial infection and is characterized by swelling of one or more large joints generally in the absence of systemic symptoms. objectives: review cases of hemophilia a and b patients with episodes of provoked hemarthrosis refractory to intensive recombinant factor replacement therapy found to have concurrent lyme arthritis. design/method: we report two clinical cases and review relevant literature. results: first, we report a year-old male with moderate hemophilia a with a provoked knee hemarthrosis which failed to improve despite months of intense factor replacement therapy requiring multiple hospitalizations. factor replacement regimens included twice daily standard half-life recombinant factor viii products or daily to every other day extended half-life recombinant factor viii products with trough levels aimed as high as - %. factor viii pk studies were obtained for dosing, to confirm adherence, and to evaluate for subclinical inhibitors (inhibitor testing was negative). given protracted symptoms additional workup for hemarthrosis was pursed. lyme titers were positive for ( )igg, though negative for igm. he was treated with days of doxycycline during which time hemarthrosis greatly improved on examination and imaging, and he was able to recover function through physical therapy. second, we report a year-old male with moderate hemophilia b who required multiple hospital admissions for a provoked knee hemarthro-sis with no improvement in symptoms despite weeks of daily or twice daily factor replacement with standard halflife recombinant factor ix products aiming for % correction. we performed inhibitor testing (which was negative) and pk studies to assess for non-detectable inhibitors, dosing and adherence. lyme testing was positive for ( )igg, though negative for igm. he was treated with amoxicillin for days during which time hemarthrosis significantly improved on examination and imaging. diagnosis and follow-up imaging studies for both patients included mri and serial bedside ultrasounds performed as per uc san diego school of medicine mskus guidelines. background: relapse/refractory aml following allogeneic hematopoietic stem cell transplant (hsct) holds a high mortality rate. current relapse/refractory therapy modalities for younger patients may include re-induction with a clofarabinebased regimen followed by second allogeneic hsct. even for patients who undergo second hsct, the five-year survival rate is dismal. new therapies, including small molecule inhibitors, are being studied in the post-hsct relapse setting or those unfit for hsct with promising results. venetoclax is a small molecule inhibitor that has received breakthrough designation for aml treatment in elderly patients objectives: to report a young adult aml patient with relapse post hsct who was successfully re-induced with topotecan, vinorelbine, thiotepa, clofarabine (tvtc) and has sustained remission with venetoclax maintenance therapy. this approach appears to be unique in terms of reported literature. results: our patient is now a -year-old female noted to have mll rearranged aml at initial diagnosis when she was years old. she underwent chemotherapy consisting of cytarabine/daunorubicin according to standard + . due to persistent disease, she was re-induced with g-csf, clofarabine, and high-dose cytarabine (gclac) which put her in cr. her course was complicated by sepsis, colitis, gastrointestinal bleed, deep venous thrombosis, and transfusionassociated circulatory overload. given her co-morbidities, she received another cycle of clofarabine/cytarabine, and then proceeded to reduced intensity allogeneic hsct, according to bmt ctn . the patient tolerated hsct well and experienced no transplant-related complications, including no acute or chronic gvhd. unfortunately, she relapsed about month's post-hsct. initial salvage therapy consisted of another course of g-clac, but due to persistent disease the decision was made to re-induce her with topotecan, vinorelbine, thiotepa, and clofarabine (tvtc). during this time however, she was found to have extensive infection with a fusarium species requiring a course of anti-fungal therapy. bone marrow evaluation showed no residual disease with an mrd of < . %. once the absolute neutrophil count recovered, the patient was started on single-agent venetoclax for maintenance therapy, which has been well-tolerated. she remains in morphologic remission for over months. we describe herein a young adult with multiply relapsed aml wherein tvtc re-induction, followed by maintenance with venetoclax were safely used in the post-hsct setting. venetoclax therapy in the relapsed aml setting warrants further study. background: vitamin b deficiency is uncommon in children in developed countries, especially in the absence of risk factors like malabsorption or inadequate dietary intake. it often presents with non-specific symptoms and signs and can elude diagnosis. the recognition and treatment of vitamin b deficiency is critical as it can lead to bone marrow failure as well as severe neurological and developmental problems in children. to increase index of suspicion of vitamin b deficiency anemia in children. we report a rare case of vita-min b deficiency anemia in a child who presented with a severe macrocytic anemia, with signs of hemolysis and concern of malignancy. design/method: an almost three-year-old previously healthy girl presented with a few day history of fever, emesis, fatigue and pallor. she had no dysmorphic features, hepatosplenomegaly or lymphadenopathy on exam, growth and development were normal. laboratory findings showed severe macrocytic anemia (hemoglobin . grams/dl; mcv . fl) with reticulocytopenia. signs of intravascular hemolysis were present with elevated lactate dehydrogenase ( , units/l) and haptoglobin below assay limit. immune-mediated hemolysis was ruled out. initial picture of a hemolytic anemia was compounded by other findings of moderate neutropenia, mild thrombocytopenia and peripheral smear showing occasional blasts. further workup was done with a broad differential diagnosis that included leukemias, hemolytic anemias, bone marrow failure syndromes, and specific deficiencies. results: workup revealed abnormally low vitamin b levels along with significantly elevated homocysteine and methylmalonic acid levels indicating functional vitamin b deficiency. bone marrow evaluation showed megaloblastic anemia and dyserythropoiesis consistent with vitamin b deficiency, and ruled out leukemia. vitamin b deficiency can cause a hemolytic anemia like picture secondary to intramedullary hemolysis due to ineffective erythropoiesis. myeloid precursors are also affected which can lead to neutropenia, thrombocytopenia, and abnormal peripheral blood cells. in our patient, initial symptomatic anemia was treated with blood transfusion, followed by intramuscular vitamin b injections with normalizing lab values. so far, workup for an etiology for vitamin b deficiency is negative except for an equivocal range of anti-parietal cell antibodies raising concerns for pernicious anemia; however it is rare in this age group. another rare condition is an inborn error of the cobalamin transporter. she is currently on oral vitamin b supplementation and further workup will be planned based on response. conclusion: this case highlights the importance of early consideration and thorough evaluation of vitamin b deficiency in children with unclear etiology of anemia, so that prompt treatment can be initiated. memorial hospital/ university of miami, miami, florida, united states background: despite great success in the treatment of acute lymphoblastic leukemia (all), the outcomes for patients with relapsed all remain poor. prognostic indicators include timing and site of relapse. blinatumomab, is the first agent in its class that simultaneously binds cd -positive cytotoxic t cells to cd -positive b cells resulting in lysis of malignant cells. however, mechanisms of leukemia resistance to blinatumomab are unclear. objectives: to describe a case with multiple sites of extramedullary (em) relapse during blinatumomab therapy. results: a -year-old hispanic male with philadelphia positive, cd -positive b-precursor cell all refractory to chemotherapy, had failed a bone marrow (bm) and was placed on blinatumomab and imatinib. he achieved minimal residual disease (mrd)-negative systemic remission, but during his fifth cycle developed bilateral periorbital masses. biopsies confirmed cd -negative isolated em relapsed disease, which was treated with radiation therapy (rt). there was notable resolution of em disease and he continued systemic therapy. subsequently, he presented with a painful left scapular swelling. imaging showed muscle and lung parenchymal em relapse with cd -positivity confirmed on histology. he continued on blinatumomab with localized rt while awaiting car-t cell therapy. his bm mrd remained negative until he developed systemic mrd-positivity with cd -positive blasts following the sixth cycle. primary resistance to blinatumomab is poorly understood. it is proposed that expansion of cd -negative clones or downregulation of cd following blinatumomab may play a role. this was observed in our patient's periorbital relapse; but subsequent em and systemic relapses were cd -positive, consistent with the co-existence of multiple clones in relapsed all. it has also been postulated that em relapse could be linked to the failure of blinatumomab or t cells to migrate to em sites of disease or drug inactivation by the microenvironment. the second em relapse in our patient, with cd -positive disease suggests this as a possible mechanism of relapse. this was reported in patients with cd positive non-hodgkin lymphoma (nhl), and higher doses of blinatumomab however, have shown promising results in this population. despite blinatumomab's effectiveness in inducing remissions in patients with refractory/relapsed all, it appears to have limitations in patients with em disease. these may arise either from the multiclonality associated with relapsed all or due to the emergence of resistance to blinatumomab, including failure to migrate to em sites. background: cyclic neutropenia is a rare hereditary disorder, characterized by recurrent neutropenia, cycling at about week intervals, with variable associated symptoms including oral ulcers and fever. there are reported cases of cyclic neutropenia associated with chronic inflammation leading to development of reactive aa amyloidosis. one patient also presented with amyloid goiter. we report a new case of cyclic neutropenia with associated renal and thyroid amyloid. design/method: a -year-old female presented with a month history of thyromegaly, and recurrent aphthous ulcers associated with fevers. laboratory workup showed severe neutropenia, anemia, azotemia, and abnormal thyroid function, with an absolute neutrophil count - / l, hemoglobin - . g/dl, serum creatinine - . mg/dl, and uric acid - . mg/dl. thyroid stimulating hormone was elevated - . iu/ml, and normal free t . urinalysis showed + protein, + blood, and - urine red blood cells/hpf. chest radiograph showed mild narrowing of the trachea from thyroid compression. bone marrow biopsy showed a hypocellular marrow, with tri-lineage hematopoiesis, left shifted myeloid maturation with very rare mature neutrophils. both renal biopsy and thyroid fine needle aspiration revealed abundant amyloid. of note, her father had aa amyloidosis, resulting in end-stage renal disease (esrd) requiring hemodialysis, and recurrent aphthous ulcers. the family history suggested a familial predisposition. genetic testing revealed a pathogenic elane c. a>t gene mutation with autosomal dominant inheritance confirming the diagnosis of cyclic neutropenia. we treated our patient with daily granulocyte colony stimulating factor to reduce the burden of chronic inflammation induced by cyclic neutropenia, and to preserve renal and other end organ function affected by further amyloid deposition. results: proband with elane gene mutation positive cyclic neutropenia, amyloidosis of thyroid and kidney, with a positive paternal history of aa amyloidosis resulting in esrd. cyclic neutropenia may result in chronic inflammatory states leading to secondary amyloidosis. university of kentucky, lexington, kentucky, united states background: overall survival of burkitt lymphoma (bl), regardless of stage, is greater than % in the pediatric population when treated with multi-agent chemotherapy. adenovirus is a common, usually self-limited infection within the pediatric population; however, findings can vary within an immunocompromised host. hepatitis is a rare complication, with very few reports of radiologic findings in this patient population. we discuss a three year old male with history of bl who presented with clinical and radiographic evidence of relapse but was found to have adenovirus hepatitis. design/method: a case report of a patient with bl in complete remission after completion of standard of care chemotherapy, who presented with return of high fever, elevated ldh, transaminitis and hepatic lesions. we describe the hepatic imaging and pathology consistent with adenovirus hepatitis in this immunocompromised host. our patient presented at three years old with a six week history of worsening abdominal pain and fevers. he was found to have a right sided pleural effusion, multiple lesions of the liver, and diffuse abdominal lymphadenopathy; biopsy of lymph tissue was consistent with bl. he completed therapy per anhl arm b and was in a complete remission at the end of planned therapy. one month after completion of therapy, he returned with high fever, abdominal pain and transaminitis, similar to his initial presentation. ct scan showed multiple hypodense discrete lesions throughout the liver and re-accumulation of right sided pleural effusion. ldh peaked at u/l (uln u/l). uric acid remained within normal limits. bilirubin peaked at . mg/dl, conjugated . mg/dl. liver biopsy was performed, showing smudgy nuclei with immunohistochemical staining positive for adenovirus. there was no evidence of lymphomatous involvement. resolution of hepatic lesions and transaminitis, with normalization of ldh and fever, occurred with symptomatic treatment alone. adenovirus is known to cause systemic disease in immunocompromised patients and rarely hepatitis. no pediatric patients with discrete hepatic lesions secondary to adenovirus have been reported in the literature. three cases of discrete hepatic lesions have been reported in adult immunocompromised patients, two with fatal fulminant liver failure and one who required cidofovir. this case demonstrates that a common pediatric viral infection can present with lesions concerning for metastatic disease in a pediatric lymphoma patient. prompt diagnosis is vital in the management of these patients when recurrent lymphoma is in the differential. background: heparin induced thrombocytopenia (hit) is an immunologic process in which antibodies bind a heparin complex and cause a paradoxical hypercoagulable state. ramifications of this process may include a multitude of thrombotic events and bleeding complications secondary to platelet consumption. in our patient, hit manifested as increased bruising, an acute decrease in platelet count, and continual clotting of her crrt circuit. hit, although rare in pediatrics, should be included in the differential for children with thrombocytopenia who have received heparin products. to present a unique case report of a critically ill pediatric patient who developed hit in the presence of multiorgan system failure and to discuss the challenges encountered with identification of an alternative anti-coagulant. results: a yo obese, caucasian female child presented to our facility with bilateral pulmonary emboli (of unclear etiology). initially, she was started on a continuous heparin infusion, but was transitioned to enoxaparin within days without issue. five days after enoxaparin was initiated, the patient developed acute kidney injury (evidenced by increasing creatinine) attributable to her biventricular heart failure. due to her need for continuous renal replacement therapy (crrt), she was transitioned back to a continuous heparin infusion. whereas her initial platelet count on transition was normal, she developed severe thrombocytopenia ( , ul) within hours. due to intermediate risk but low suspicion for hit, pf antibodies were sent which were positive. after much discussion, she was transitioned to an argatroban infusion which was titrated according to ptt levels. within hours, her platelet count normalized. at discharge, she was prescribed apixaban for anti-coagulant management. conclusion: hit is an uncommon presentation in the pediatric population. given its rarity, there is often a delay in diagnosis which increases risk of complications such as bleeding, stroke, and limb ischemia. even if the diagnosis is suspected or proven, there may be challenges in initiating alternative agents as limited data exists on pediatric options. as argatroban remains the treatment of choice for patients with hit, experience in pediatric patients is limited, and dosing recommendations have been extrapolated from adult studies. anecdotal data exists for use of bivalirudin in children, although studies, primarily, focus on use in specific cardiac cases. in our patient's case, choice was further complicated by renal failure. this case study highlights the need for further research regarding the identification of a secondary anti-coagulant agent for use in pediatric patients with hit. background: subcutaneous panniculitis-like t-cell lymphoma (sptl) is a rare form of non-hodgkin's lymphoma characterized by infiltration of cytotoxic t-cells into subcutaneous tissue. sptl occurs in both adults and children and can present in both patient populations as either alpha/beta or gamma/delta subtypes. patients with the gamma-delta phenotype have an overall poorer survival, although the exact etiology is unclear. interestingly, both subtypes of sptl can present with secondary hemophagocytic lymphohistiocytosis (hlh), and this is associated with a worse prognosis. currently, there are no standardized treatment protocols for sptl, and clinical management includes watchful waiting, corticosteroids/immunosuppression, chemotherapy, and stem cell transplant. the primary objective was to compare how two patients with the same diagnosis responded acutely to therapy. we performed a retrospective chart review of two pediatric patients at our institution who were diagnosed with alpha/beta sptl and secondary hlh. we examined each presentation, treatment course, and outcome. we then completed a brief review of the current literature describing treatment of and outcomes for sptl with secondary hlh. results: these two patients presented in a similar manner with signs and symptoms of hlh. each was then subse-quently diagnosed with alpha/beta sptl after biopsy of cutaneous nodules and each had diffuse disease, as measured by pet. however, they demonstrated vastly different acute responses to therapy. one patient was pre-treated with systemic glucocorticoids before receiving definitive chemotherapy and tolerated therapy well as an outpatient. the other patient started systemic chemotherapy without steroid pretreatment and developed severe cytokine storm characterized by hypotension, cardiac dysfunction, multi-organ failure and cytokine elevation. both patients achieved complete remission (cr) after treatment with chop chemotherapy and remain disease-free - months off therapy. in patients presenting with sptl and secondary hlh, we propose that initial treatment with antiinflammatory or anti-cytokine therapy can decrease, or even prevent, the possibility of life threatening cytokine release as a result of cytotoxic chemotherapy. background: congenital dyserythropoietic anemia type ii (cda ii) is a rare autosomal recessive disorder, rarely presenting in the neonatal period. iron overload often occurs as a late sequela of ineffective erythropoiesis and intramedullary hemolysis. objectives: to report the novel use of iron chelation in an infant with cda ii associated with severe iron overload. the patient is a -month-old, former -week infant with prenatal non-immune hydrops and transfusion-dependent fetal anemia who presented with persistent anemia, reticulocytopenia, hyperbilirubinemia, liver dysfunction, and hyperferritinemia. his initial ferritin was . ng/ml, tibc ug/dl, and transferrin mg/dl. his bone marrow biopsy showed trilineage hematopoiesis and erythroid dyspoiesis characterized by binucleation of late-stage precursors. genetic testing revealed a compound heterozygous missense mutation and splice site mutation in the sec b gene, confirming the diagnosis of cda ii. initial liver biopsy revealed mild portal fibrous expansion, and abundant hepatic iron deposition. his ferritin continued to increase, peaking at , ng/ml, along with liver enzymes peaking at an alanine aminotransferase (alt) of u/l and aspartate aminotransferase (ast) of u/l. ferriscan showed an elevated estimated liver concentration of . mg/g dry tissue. repeat liver biopsy months later showed giant cell hepatitis with worsening mild portal fibrosis and hemosiderosis. additionally, tissue liver iron concentration was mcg/g dry weight. cardiac t * mri revealed mild cardiac iron deposition. given his significant degree of iron overload, deferoxamine was used to reduce hemosiderosis and liver morbidity in preparation for bone marrow transplantation. the patient received deferoxamine mg/kg/day iv x days/week for three months, without any clinically significant adverse events. blood counts and hepatic and renal function were monitored weekly without any abnormalities. growth parameters and liver enzymes significantly improved while receiving chelation therapy. as a noninvasive, cost-effective method, serum ferritin levels were monitored monthly to gauge response to treatment. despite receiving blood transfusions every - weeks, serum ferritin decreased to ng/ml and liver enzymes decreased to alt u/l and ast u/l prior to bone marrow transplantation. we report the use of deferoxamine in a patient with cda ii less than years of age, for treatment of iron overload. our patient tolerated deferoxamine well without significant adverse events or organ toxicity. deferoxamine may be a well-tolerated method of reducing iron burden in young patients with iron-loading pathologies. background: low grade gliomas with kiaa- -braf fusions typically have a favorable prognosis with infrequent rates of high grade transformation, low rates of metastasis and even lower rates of extra cns metastasis. while highgrade transformation has been reported for tumors with braf v e mutations and cdkn a deletions, it has not been pre-viously reported in gliomas with kiaa- -braf fusions. while there are case reports of high-grade cns malignancies metastasizing through a ventriculo-peritoneal (vp) shunt, low-grade gliomas metastasizing in this manner are extremely rare. objectives: to describe a unique case of peritoneal tumor dissemination of a braf fusion positive high grade neuroepithelial tumor in a child with a vp shunt placed for multifocal braf fusion positive low grade astrocytomas results: an eight-year-old male was initially diagnosed with multifocal low-grade astrocytomas of the hypothalamus and c -c spinal cord. initial testing revealed the kiaa- -braf fusion, but no cdkn a or braf v e mutation. initial surgical management included a vp shunt and resection of the cervical spinal lesion. he received vincristine and carboplatin, followed by transition to vinblastine given new thoracic metastatic lesions after months of therapy. at months after diagnosis, scans were concerning for diffuse leptomeningeal progressive disease and new intracranial lesions, necessitating craniospinal radiation. following a near cr, he presented months later with acute onset of abdominal pain. a ct scan revealed peri-renal and perirectal soft tissue masses, confirmed by exploratory laparotomy to be peritoneal tumor dissemination of high grade neuroepithelial tumor. a kiaa -braf fusion was noted and confirmed by rt-pcr, identical to that seen in the original cns tumors. additional findings included deletion of chromosome p (without q loss) and heterozygous and homozygous deletion of cdkn a found by fish. brisk mitotic activity justified a high-grade designation. salvage chemotherapy consisted of cycles of ice with subsequent resolution of pet-avid disease and only minimal peri-nephric tissue remaining. given the favorable response, surgical resection and multiple tissue biopsies were performed which documented no residual active disease. the shunt was revised and he started trametinib for maintenance. we present a unique case of peritoneal dissemination of high grade neuroepitheial tumors with the same kiaa- -braf fusion as multifocal low grade astrocytomas in a child with a vp shunt. this raises suspicion for tumor metastasis and transformation to a higher grade malignancy versus two distinct diseases, which may be indicative of an underlying cancer predisposition. texas children's hospital, houston, texas, united states background: polycythemia is a common referral to hematology. it is important to evaluate for a high oxygen affinity hemoglobinopathy, ensuring appropriate testing is performed for early diagnosis and avoidance of additional tests and procedures. a year old mexican female presented with an elevated hemoglobin and hematocrit, symptoms of plethora of her hands and feet, chest pain, palpitations, and fatigue. further confounding the picture, she also had significant menorrhagia and iron deficiency. she was diagnosed with the rare high oxygen affinity hemoglobin new mexico variant, only previously described once in the literature in a year old black boy. objectives: the patient initially presented at age with a hemoglobin of . g/dl and a hematocrit of . %. initial work up consisted of a hemoglobin electrophoresis which diagnosed sickle cell trait, a co-oximetry panel which was normal, and erythropoietin level of mu/ml, also normal. she was then lost to follow up and re-referred at age . she is a competitive basketball athlete, and at that time, she presented with a hemoglobin of . g/dl, and hematocrit of %. erythropoietin level continued to be normal at mu/ml. design/method: cardiology was consulted regarding chest pain and palpitations with a normal evaluation. chest x-ray was also normal. a bone marrow aspirate and biopsy was performed with results significant for mild erythroid hyperplasia and mild reticulin fibrosis. jak mutation, von hippel lindau, bpgm, and hereditary erythrocytosis mutations including phd , hif a, and epor mutation analysis were sent, all of which were normal. testing to mayo clinic for p rbc oxygen dissociation returned low at mmhg ( - mmhg normal range) and subsequently a hemoglobin electrophoresis identified a hemoglobin variant leading to beta globin gene sequencing. results: patient found to be heterozygous for hemoglobin new mexico, with . % hb new mexico and . % hba, and . % hba . there was no evidence of hbs. when evaluating patients with polycythemia, maintaining a high index of suspicion for high affinity hemoglobinopathies may eliminate further unnecessary and invasive testing for patients. caution should be used when using hemoglobin electrophoresis testing since hb new mexico is known to migrate similarly to hbs on hplc with minimal change that may not be detected in regular laboratories. most high affinity hemoglobinopathies are reported to not have significant symptoms. in this case, our patient complains of fatigue, occasional palpitations and plethora of hands and feet. we will need to further follow this patient for possible attributable symptomatology. divya keerthy, simone chang, warren alperstein, patricia delgado, claudia rojas, ofelia alvarez, matteo trucco university of miami jackson memorial hospital, miami, florida, united states background: improved technology is enabling detection of previously unidentified translocations and mutations in otherwise unclassified sarcomas. one such mutation is the bcl- co-repressor -internal tandem duplication (bcor-itd) allowing for the new classification of bcor positive undifferentiated round cell sarcomas (urcs). this sarcoma has a similar appearance to clear cell sarcoma of the kidney (ccsk), potentially representing an extra-renal manifestation of this tumor, but their clinical pathologic features are not identical. objectives: this case highlights how recombinant polymerase chain reaction (rt-pcr) and bcor immunohistochemical staining can ease the diagnosis of this rare sarcoma. results: a month-old female presented for right sided pre-septal cellulitis and a temporal subcutaneous mass. the detection of multiple other subcutaneous nodules on exam raised the concern for malignancy and she was admitted for evaluation. she had two subcutaneous masses on her abdomen, with more cutaneous masses on her legs, back, shoulder, cheek and submandibular areas. she lacked spontaneous lower limb movement and had bilateral clonus. imaging confirmed multiple masses throughout the body including paravertebral area from t to l , bilateral adrenal glands, left kidney and muscles of upper and lower extremities. initial differential included neuroblastoma, infantile myofibromatosis, rhabdomyosarcoma or atypical presentation of a renal tumor. however, synaptophysin and chromogranin stains were negative. with standard immunohistochemistry, the tumor could be only broadly classified as "undifferentiated sarcoma" maintaining the diagnostic challenge. using rt-pcr in the setting of a morphologically primitive round cell neoplasm with strong bcor expression, two external institutes simultaneously diagnosed the tumor as bcor-urcs. the primary lesion is unknown but potentially may have arose from the kidney. bcor-urcs has a heterogeneous histology with tumor cells appearing monomorphic in nests of - cells separated by septa with uniform nuclei. there is frequently an "orphan annie eye" appearance and sparse cytoplasm to the cells. diagnosis cannot be made solely on evaluation of this nonspecific histology. rt-pcr uses the genetic abnormality in undifferentiated sarcomas to narrow the differential and bcor immunohistochemical staining provides further context. bcor has significant diagnostic value given its sensitivity and specificity in urcs. another potential marker includes ywhae-nutm b fusions, which occur in smaller subset of cases, but requires further study. rt-pcr has helped further classify tumors leading to the diagnosis of a rare undifferentiated sarcoma with bcor overexpression. while this technology is beneficial, its availability is limited. if accessibility improves, earlier identification and treatment may be possible maximizing the chance for a positive outcome. background: hematohidrosis is a rare condition that mimics bleeding disorders. cases present with oozing blood tinged fluid from various sites like eyes, ears, nose, skin, etc. reported causes of this condition were stress or fear, physical activity, psychological disorders. the condition is self-limited and don't affect the general condition of the patients, but it may contributes to psychosocial problems and may increases their stress and anxiety. so this condition needs to be promptly treated. to test the response of this disease and the associated headache to propranolol treatment. design/method: our case female patient years old st offspring of non consanguineous marriage, was admitted with recurrent episodes of oozing blood tinged fluid from eyes, ears and nose months before admission, about . - ml from each orifice, lasted - minutes and subsided spontaneously. it could involve the sites simultaneously or - sites. the number of attacks was - times per day then gradually increased to - times per day. later on the patient developed a bleeding attack from umbilicus. these attacks were aggravated by stress and physical activity and decreased with rest and sleep. the condition was associated with severe headache involving the whole head, throbbing in nature of gradual onset, increased by physical activity and relieved by analgesics. the condition was not associated with vomiting, blurring or diminution of vision, ocular pain, eye discoloration. no earache, tinnitus or diminution of hearing. there was no other form of discharge from eyes, ears or nose. no history of ecchymotic patches, bleeding from other orifices or blood product transfusion. no history of trauma, drug intake, fever or rash. no symptoms of other system affection. past history of recurrent attacks of epistaxis and two operations were done that passed without remarkable bleeding. no similar condition in the family physical examination was free, no evidence of psychological problems. complete blood count, coagulation profile, platelets function, factor and c.t brain were normal. oozing fluid from the patient was analyzed showed the same components as blood. results: our case started oral propranolol . mg/kg/day based on its use in similar cases in literature. the frequency of attacks and headache reduced then stopped after months of treatment and didn't recur after stoppage of propranolol. propranolol can treat this condition successfully. further investigations are needed to determine the link between this condition and severe headache our case was suffering from. background: wilms tumor is the most common renal solid tumors of childhood and is derived from primitive metanephric cells located in the kidney. primary extra-renal wilms tumors (erwt) are extremely rare, estimated to comprise . - % of all wilms tumors. despite similar histologic appearance intrarenal and erwts differ in embryologic tissues of origin. erwts arise from the more primitive mesonephric or pronephric origin and, therefore, can develop anywhere along the craniocaudal migration pathway of these primitive tissues, most often retroperitoneal, inguinal/genital, lumbosacral/pelvic and mediastinal. these tumors are typically staged and treated per national wilms tumor study (nwts) guidelines, and, by definition, are stage ii or greater due to location beyond the kidney borders. based on the cases reported in the literature, outcomes for erwt are comparable to renal wilms tumors with an % local recurrence rate and an % two-year event-free survival. we report the first case of a stage iii testicular extrarenal wilms tumor in an -month-old male with an intrabdominal undescended testis who underwent complete surgical excision followed by chemotherapy and inguinal radiation. results: a full term -month old male underwent orchipexy for an undescended left testicle. the testicle was noted to be grossly abnormal with a pea-sized thickened tissue adherent to the upper pole and a separate mass outside of the scrotum on the superior epididymis. both masses were removed, and s of s pathology demonstrated wilms tumor with favorable histology and negative margins. ct imaging of the chest, abdomen and pelvis were negative for a primary renal tumor, local residual disease, pathologic lymph node enlargement or distant metastases. the tumor was classified per nwts as stage iii due to tumor removal in multiple pieces. the patient completed dd- a treatment with vincristine, doxorubicin and dactinomycin per aren with cgy left inguinal radiation. he is currently months off therapy without clinical or radiographic evidence of recurrent disease. primary erwt is an extremely rare malignant neoplasm associated with challenges in diagnosis, staging and treatment. based on the cases reported in the literature, outcomes are similar to that of intrarenal wilms tumor. there are four pediatric paratesticular wilms tumors reported in the literature and, to the best of our knowledge, this is the first case of stage iii testicular wilms tumor successfully treated with dd- a chemotherapy and radiation. in erwt, nwts guidelines for staging and treatment should be applied with evaluation of both kidneys to exclude an intrarenal primary tumor. background: patient is a yo f, with esrd secondary to atypical hus versus ttp, who presented with thrombotic microangiopathy, aki, thrombocytopenia and anemia after a living unrelated donor kidney transplant. patient initially had downtrending creatinine. on post-op day , hematology was consulted for an increasing ldh and drop in platelets. peripheral smear was notable for an absence of schistocytes. yet, biopsy of the kidney revealed microthrombi. the patient was diagnosed with a thrombotic microangiopathy. plasmapharesis was initiated on day # , at which time ms r was noted to have significantly elevated creatinine. plasmapharesis did not yield any correction in labs and significant bruising developed. patient was started on eculizimab; plasmapharesis was stopped. shortly after, creatinine, anemia and thrombocytopenia corrected to levels at which she was discharged. overall, patient was found to have progressive anemia, thrombocytopenia, an increasing creatinine and ldh ( s) concerning for atypical hus, despite absence of schistocytes on peripheral smear. she responded well to eculizimab, with correction of hematologic changes during induction. she was discharged on eculizimab and continued to respond with normalizing platelet counts and hemoglobin. the differential in light of patient's thrombotic microangiopathy and thrombocytope-nia also included ttp. yet, adamts remained normal. dic was unlikely given normal fibrinogen level and d-dimer. objectives: presentations of atypical hus vs ttp. discuss eculizumab as a treatment of atypical hus. highlight atypical presentations of illness in transplant patients. results: despite absence of schistocytes by smear, pt was diagnosed with atypical hus based on presentation and after failing plasmapharesis, she responded well to eculizumab. though her presentation was abnormal, her response to this antibody that blocks the complement cascade suggests that she was experiencing a complement-mediated process. there are rare documented cases in the literature of atypical hus without schistocytes. hemolytic uremic syndrome (hus) is characterized by hemolytic anemia, thrombocytopenia and acute kidney injury. atypical hus is a diagnosis of exclusion, not due common etiologies such as shiga toxin. among atypical causes are complement-mediated forms, caused by an antibody to complement factor. in addition to plasmapharesis, renal transplant and supportive care, the mainstay of treatment for atypical hus is eculizumab (an antibody that blocks the complement terminal cascade). this case describes a patient unique in that, she was diagnosed with atypical hus without any schistocytes by smear. secondly, she responded to eculizumab, with unremarkable gene studies. finally, this case highlights that transplant patients often have unique presentations. nicklaus children's hospital, miami, florida, united states background: synovial sarcoma is a spindle cell tumor categorized as a soft tissue sarcoma. the chromosomal translocation t(x; ) leading to the ss -ssx fusion protein is unique to this sarcoma. it is a slow growing tumor with common recurrences and often, at presentation, with evidence of metastatic disease. if resection is not feasible, then neoadjuvant with adjuvant chemotherapy is recommended. metastasis carries an unfavorable prognosis given synovial sarcoma historically does not respond well to chemotherapy. trabectedin is a well-tolerated alkylating agent currently indicated for the treatment of liposarcoma and leiomyosarcoma. we present a -year-old male with metastatic synovial sarcoma to the lungs that progressed and was refractory to chemotherapy. he was administered trabectedin as a form of palliative chemotherapy, with significant clinical and radiographic response. design/method: pubmed search was done with search for terminology including "synovial sarcoma" and "trabectedin". papers relevant to our case were selected for literature review. a -year-old male patient presented with a large right axillary mass. initial imaging showed a heterogeneous multiseptated mass invading the subscapularis and teres major muscles along with innumerable lung nodules. biopsy confirmed diagnosis of monophasic synovial sarcoma. the patient was started on protocol arst with ifosfomide, mesna, doxorubicin. he completed cycles followed by radical resection and sessions of radiation. due to progression of disease multiple chemotherapy regimens were tried including topotecan and cyclophosphamide, protocol advl with lorvotuzumab, and pazopanib. imaging of the chest continued to show significant progression of metastasis. the patient's clinical status deteriorated with worsening respiratory status, requiring l of oxygen therapy, and inability to ambulate. he was started on trabectedin . mg/m for palliative care. after cycles of treatment patient was no longer requiring oxygen and was ambulating without assistance. radiological imaging showed significant reduction in number and size of lung nodules. trabectedin is a recently approved alkylating agent for the management of sarcomas resistant to first line treatment. response in synovial sarcoma is scarcely documented in the pediatric population. epidemiology places the most common age group in the young adults and children. our case opens the doors to further consideration of the use of trabectedin in the pediatric patient with metastatic synovial sarcoma. background: gata is an x-linked gene that plays critical role in hematopoiesis. mutations of gata gene can be associated to various blood disorders including diamond blackfan anemia, cytopenia, congenital dyserythropoietc anemia and acute megakaryoblastic leukemia. we report a patient with macrocytic anemia and platelet dysfunction who carries a novel gata mutation that has not been reported. results: a now -month-old male with complex medical history including prematurity at weeks, dysmorphic features, global developmental delay, hyperinsulinism, hypogonadotropic hypogonadism, growth hormone deficiency, micropenis, failure to thrive, patent ductus arteriosus status post ligation, and severe hypotonia, was referred to hematology at months old for resolved, transient thrombocytopenia and macrocytic anemia since month of age. chromosomal microarray showed chromosome deletion of q . , which is the rps gene. he doesn't have a family history of diamond blackfan anemia (dba), despite mom having the same rps mutation. he was then diagnosed with dba. his lab workup showed mild macrocytic anemia (hgb . g/dl, mcv fl), normal to inappropriately low reticulocyte count, normal white blood cell and platelet counts, hgf %, erythroid ada . eu/gm hgb (elevated). he has abnormal pfa- , with prolonged closure time of both adp and epinephrine. he had low von willebrand antigen and ristocetin cofactor activity. he has severe pancreatic insufficiency. bone marrow biopsy showed normocellular marrow with trilineage hematopoietic maturation, without ringed sideroblasts. since mother has the same rps gene mutation, maternal labs were done and showed no evidence of macroytosis or anemia. the diagnosis of dba was questioned. whole exome sequencing did not identify any pathogenic sequence changes in the coding regions of rps gene, but detected a gata mutation r w, which was reported variant of uncertain significance. his mother shares the same mutation and is asymptomatic, but she may not be affected since gata iis xlinked. his father doesn't harbor the gata mutation. conclusion: gata gene encodes zinc finger dna binding hematopoietic transcription factor, which is important during erythroid differentiation. gata mutation r w has not been reported in literature and is a novel variant of gata mutation, which might be contributing to this patient's clinical picture. further studies are warranted to confirm gata mutation r w to be a pathogenic sequence change. alexander boucher, tomoyuki mizuno, alexander vinks, greg tiao, stuart goldstein, james geller cincinnati children's hospital medical center, cincinnati, ohio, united states background: hepatoblastoma (hb), the most common pediatric primary hepatic malignancy, can be associated with specific congenital syndromes. recently, chronic kidney disease and genitourinary anomalies have been linked to hb. cisplatin is a key chemotherapeutic agent in treating hb but its renal clearance and toxicity profile can limit its use for those with end-stage renal disease (esrd). objectives: using an institutional case series, we present data using cisplatin for hb in dialysis-dependent esrd and define recommended dosing for future use. design/method: a chart review of patients with concurrent hb and esrd on dialysis treated with cisplatin at our institution was undertaken. demographic data, diagnostic history, tumor pathology, alpha fetoprotein (afp), hearing assessments, dosing schema, treatment outcomes, and therapyrelated toxicities were reviewed. total cisplatin levels were collected at time points within days after each infusion. free cisplatin levels were also collected for infusions, as were dialysate cisplatin levels. pk parameters were generated using bayesian estimation with a published population pk model as a priori information. results: three patients meeting these criteria were identified. each had "low risk" (non-metastatic resectable) disease at presentation and underwent upfront resections. all had congenital renal anomalies with esrd prior to their hb diagnosis. all cisplatin infusions were given over hours, followed hours later by hemodialysis. patients and received cisplatin at % of children's oncology group's ahep weight-based dosing ( . mg/kg). patient received % of ahep body surface area-based dosing ( mg/m ) during cycle but required a second dose reduction ( mg/m ) for cycle due to prolonged cisplatin exposure (total area under the curve mg⋅h/l; average for all seven evaluable cycles mg⋅h/l) and early sensorineural hearing loss at - hz. no other hearing loss in any patient was identified; mild toxicities also included grade - emesis and grade neutropenia and thrombocytopenia. the median (range) of clearance, volume of distribution at steady-state, and elimination half-life at terminal phase for total platinum were . ( . - . ) l/hour/ kg, . ( . - . ) l/ kg and ( - ) hours, respectively. patients and received cycles with rapid afp normalization. patient required an additional cycles, for a likely second primary hb year after initial therapy. cisplatin can be used successfully in pediatric patients with esrd on hemodialysis to treat hb with minimal morbidity using % standard mg/kg-based dosing ( . mg/kg), achieving pharmacologically appropriate cisplatin exposures. background: treatment for immune thrombocytopenia (itp) has been grouped into rescue and maintenance therapy and often is reserved for patients with bleeding, severe thrombocytopenia, or for improvement in quality of life. splenectomy is considered one of the more invasive but definitive treatments with success rates of - %. treatment of itp can be more difficult in the setting of previous treatment with immune modulation or when the patient is immunocompromised and not a candidate for splenectomy. objectives: present an interesting case of a patient with an autoimmune disease that presented with severe thrombocytopenia, un-responsive to rescue therapy, and requiring emergent splenectomy in the setting of acute intracranial hemorrhage (ich). a year old female with a history of juvenile dermatomyositis presented with a fine purpuric rash on her extremities, wet purpura, and a platelet count of k/ l. bone marrow evaluation at that time was consistent with itp. she was on cyclosporine and plaquenil for dermatomyositis. platelets failed to increase after three doses of intravenous immunoglobulin and high dose steroids. following a two week course of oral prednisone and eltrombopag, she presented with persistent severe thrombocytopenia of k/ l, anemia of . g/dl, and a lower gi bleed. she was started on amicar, novo-seven, rituximab, and given platelet transfusions with no improvement in bleeding. subsequently, she developed a subdural hematoma with midline shift. surgery performed an emergent open splenectomy with concurrent continuous platelet transfusion. results: she was monitored closely post operatively and, due to ich, transfused to maintain platelets greater than k/ l. by week post-op she had normal platelet counts off transfusions. all medications were stopped within three days of discharge. she represented eight days later with abdominal pain and thrombocytosis and was found to have a portal vein, splenic vein and mesenteric vein thrombosis. she was started on lovenox therapy and admitted for monitoring due to her history of ich. it is unknown whether our patient's underlying immune dysregulation and history of treatment with immunosuppressive medications may have contributed to her unresponsiveness to multiple therapeutic agents. in addition, her significant bleeding did not allow us to fully evaluate her response to second tier therapy. this adds to the scarcity of literature of itp response in pediatric patients with autoimmune disease, and may support more aggressive therapy upfront in these patients. background: multivisceral organ transplantation involves concurrent transplantation of the stomach, pancreas, liver, and intestine with splenectomy, and has been classically used in the pediatric population for infants with intestinal failure from disorders affecting foregut integrity. while there is some data demonstrating its efficacy in adults with low-grade abdominal malignancies, it has not been traditionally used for hepatocellular carcinoma treatment. to describe a unique pediatric case of multivisceral organ transplantation as definitive therapy for refractory fibrolamellar hepatocellular carcinoma in an adolescent male. a year old male presents with a history of fibrolamellar hepatocellular carcinoma, tumor invasion of the portal vein, severe portal hypertension complicated by bleeding esophageal varices and hypersplenism. he had two treatments with yttrium- radioembolization, without significant response. he completed six cycles of traditional chemotherapy in combination with sorafenib with resolution of petavidity, but minimal decrease in tumor size and continued portal hypertension. since his disease remained relatively stable for over years, he was evaluated and listed for multivisceral organ transplantation. at approximately years and months after diagnosis, he underwent en bloc liver, pancreas, stomach, small bowel, and colon transplant with splenectomy. a single lymph node was positive for malignancy at the time of resection. in addition to expected post-transplant complications, he also developed skin only acute graft versus host disease at weeks after transplant, treated successfully with a thymoglobulin course. he clinically improved and was back to his baseline activity level, on full oral feedings within months post-transplantation. at three and six month post-transplantation, there is no concern for relapsed hepatocellular carcinoma on comprehensive imaging and evaluation. he is maintained on protocol immunosuppression and posttransplant support. we present the first known case of successful multivisceral organ transplantation in the treatment of refractory pediatric fibrolamellar hepatocellular carcinoma. background: hematohidrosis is a rare disorder that presents with spontaneous excretion of whole blood from intact skin or mucosa. diagnosis is based on clinical observation of the occurrence with the proven presence of erythrocytes and other blood components, without other abnormalities to account for the phenomenon. the existing literature is scarce and consists of primarily case studies. most reports describe bleeding from facial sites around the eyes, ears, and nose. the available literature suggests anxiety and physical or emotional stress reactions as the most common inciting events. little evidence exists regarding the ideal therapeutic approach, however propranolol has been used successfully to reduce bleeding frequency and severity in multiple case reports. a specific genetic etiology has not been elucidated, and no familial cases have previously been reported. we present a pair of half-siblings, both of whom presented with spontaneous cutaneous and mucosal bleeding before two years of age, and report on preliminary results of propranolol therapy. tanzania. at months of age, he became ill and developed spontaneous bleeding from his ears, nose, and scalp. he continued to have frequent bleeding episodes, usually related to illness or physical distress. a bleeding diathesis work-up was unremarkable, however some episodes were severe enough to require transfusions. the patient was subsequently diagnosed with hiv and hepatitis b, presumably acquired via unscreened blood product transfusions. patient b is an infant female born to the same mother as patient a, with a different father. she was healthy until two months of age when she developed spontaneous bleeding from the hairline, eyelids, ears and genital/rectal area. bleeding episodes were nearly always associated with irritability and crying. extensive coagulation workup was unremarkable. results: propranolol therapy was started in both patients, titrated to a goal of mg/kg/day. in both patients, the frequency and duration of bleeding episodes significantly improved. patient b continues to have milder occasional bleeding episodes from her eyes, ears and scalp but has significantly less discomfort and irritability during the episodes. conclusion: to our knowledge, there are no prior reports involving two related patients with hematohidrosis. this case series suggests that there may be a genetic predisposition which has yet to be identified. propranolol has shown effectiveness in reducing symptom frequency and severity. background: gliomas are the most common central nervous system tumors in children. they are classified into different grades based on genotype (idh, braf, tsc, etc.). lowgrade gliomas such as oligodendrogliomas, astrocytomas, and mixed oligoastrocytomas are classified as grades i and ii. of the molecular level alterations this case report focuses on the braf v e mutation. braf is a member of the raf family of serine/threonine protein kinases and it plays an important role in cell survival, proliferation and terminal differentiation. objectives: here we discuss two cases where dabrafenib, a braf kinase inhibitor, was utilized in the management of gliomas. the cases focus on the use of dabrafenib late versus early in disease course. design/method: patient jl is a year old female who was diagnosed with a low-grade glioneuronal tumor (c -t with a metastatic lesion to the brain) in . jl was treated with chemotherapy, radiation, and surgical resection. despite treatment, the patient's disease progressed. she developed lower extremity dysfunction, urinary incontinence, poor truncal control, and hydrocephalus. dabrafenib was started after the braf v e mutation was confirmed. patient lg is a year old female who presented in november with left facial and upper extremity weakness. ct and mri scans demonstrated a mixed solid and cystic lesion extending from the optic chiasm and hypothalamus to the right thalamus and posterior basal ganglia with additional involvement of the right cerebral peduncle. neurosurgical intervention was undertaken and dabrafenib was started after the braf v e mutation was confirmed. results: patient jl's mri scans have demonstrated improvement of the spine with diminished areas of enhancement along thecal margins, decreased volume and enhancement within the trigeminal plate cistern and resolution of ependymal enhancement within the right ventricle. the patient's most recent mri exhibits no disease progression in head or spine. jl has shown improvement clinically since starting dabrafenib. patient lg has shown improvement in strength and recent mri of the brain has shown resolution of enhancement along surgical resection margins, decreased hyperintensity along the inferomedial aspect of the right basal ganglia and no new enhancements. conclusion: low grade gliomas can alter a person's quality of life and even lead to life threatening complications. often the standard chemotherapy, radiation and surgery don't prevent these complications. genetic analysis can help clinicians target therapy towards certain mutations such as braf v e. dabrafenib has shown to decrease tumor burden, early utilization as therapy can help prevent morbidity and mortality. children's hospital of pittsburgh of upmc, pittsburgh, pennsylvania, united states background: copper is an essential cofactor in enzymatic reactions essential to proper hematologic, skeletal, neurologic and vascular function. copper requirements in children over the age of are mg/day, which is readily acquired in a typical diet. copper deficiency is known to occur in patients with the rare x-linked mutation and in older individuals with gastrointestinal bypass surgery; however, it is rarely reported in other conditions. objectives: to highlight individuals with autism spectrum disorders or developmental delay with a limited dietary repertoire are at risk for copper deficiency, thus a high index of suspicion must exist in order to diagnose the disorder. design/method: a y/o boy with a prior diagnosis of global developmental delay and oral aversion presented with slowly progressive fatigue, weakness, gait instability, and weight loss. his longstanding feeding difficulties were refractory to intensive feeding programs. his daily diet consisted of - oz of milk and - individual servings of butterscotch pudding ( - calories/day, . mg iron/day). initial complete blood count demonstrated white blood cell count of . , absolute neutrophil count of , hemoglobin of . , mean corpuscular volume of < , reticulocyte count of . , platelet count of . review of his peripheral blood smear revealed microcytic, hypochromic red cells without marked fragmentation, anisopoikilocytosis and ringed sideroblasts; there were no morphologic abnormalities of his leukocytes or platelets. iron studies demonstrated ferritin of , total iron binding capacity of , and % iron saturation. he had no evidence of b , folate deficiency or blood loss. additional evaluation revealed a serum copper level of (range - ), and cerulosplasmin of . (range - ). results: once a diagnosis of copper deficiency was made, the patient promptly began a course of parenteral copper repletion. he received iv copper mcg/kg/day x days then weekly intravenous infusions. given his malnutrition, a gtube was placed to begin oral copper repletion and enteral nutrition. within weeks his copper level improved as well as his blood counts. unfortunately, although his blood counts and copper levels normalized, his neurologic status remains below his old baseline although, he has made gains in his gross and fine motor abilities. conclusion: acquired copper deficiency in the pediatric population is a rare event but given the hematologic and neurologic consequences, prompt recognition and treatment is important. this patient's clinical course demonstrates the need to have a high index of suspicion of concomitant nutritional deficiencies other than those routinely evaluated such as iron, b and folate. background: lymphoepithelioma-like thymic carcinoma (lelc) is a rare, aggressive neoplasm with a high rate of invasion, metastasis and recurrence. there are no known curative therapies for metastatic lelc. we report the case of a -year-old male who presented with metastatic ebv positive lelc. sites of disease included a large primary anterior mediastinal mass and metastases to hilar lymph nodes, lungs and liver. he was initially treated with cisplatin and fluorouracil followed by mediastinal radiation. he had a partial response to therapy but his end of therapy scans showed disease progression in lungs, liver, and hilar, supraclavicular and axillary lymph nodes. objectives: molecularly targeted therapies tailored to the patient's genetic profile offer a novel approach to obtain improved survival outcomes. design/method: the patient enrolled on a precision medicine trial, nmtrc : molecular-guided therapy for the treatment of patients with relapsed and refractory childhood cancer (nct ). in this study, tumor/normal whole exome sequencing and tumor rna sequencing were performed and a molecular report detailing the results of genomic and gene expression analysis was generated. a treatment plan was designed within a molecular tumor board comprising oncologists, pharmacists, genomicists, and molecular biologists with domain expertise. results: exome sequencing revealed somatic coding point mutations and no structural mutations (focal copy number changes or translocations). candidate somatic driver mutations included tp s x and r w as well as kit n k. both genes have been previously implicated in thymic carcinoma. rna expression analysis demonstrated aberrant activation of biological pathways, including overexpression of kit, hdac , and , tyms, and dhfr. the molecular tumor board selected the combination of pemetrexed ( mg/m ) on day of a day cycle, imatinib ( mg daily), and vorinostat ( mg days - , - , and - ) . on day of cycle , he was admitted with a herpes zoster infection and imatinib was discontinued in order to reduce risk of herpes zoster recurrence. imaging after cycles showed a complete metabolic response on f- fdg pet and a partial response by ct size criteria. as of december , the patient had received cycles of pemetrexed and vorinostat. scans in december showed an increase in the size and metabolic activity of two right lower lobe pulmonary nodules. there were no new sites of disease and imatinib was re-started. background: systemic lupus erythematosus (sle) is a chronic autoimmune disease that affects multiple organ systems and is associated with many different autoantibodies. patients can present with vague constitutional symptoms including fever, rash, fatigue, and weight loss. some of the various hematologic manifestations of sle include anemia of chronic disease, leukopenia, autoimmune hemolytic anemia (aiha), and idiopathic thrombocytopenic purpura (itp). these can be the presenting signs of sle. evans syndrome (es), a disease characterized by itp and aiha, is a rare hematologic manifestation of sle. neurofibromatosis (nf ) is a relatively common neurocutaneous disorder. these patients are at risk of developing benign and malignant tumors. its association with autoimmune disorders, including sle, remains rare. objectives: there are few cases in the literature that have patients with the combination of sle and nf . this is the only case that has a patient with sle, nf , and es. results: a -year-old caucasian female presents with two months of vaginal bleeding, weight loss and petechiae. her exam is remarkable for petechiae and café au lait macules. laboratory findings show severe anemia and thrombocytopenia. she receives blood and platelet transfusions during stabilization, and a bone marrow aspirate is performed to rule out a malignancy which is negative. based on the presence of thrombocytopenia and a positive coombs test, an autoimmune process such as es is considered. screening tests for sle reveal positive antinuclear and anti-double stranded dna antibodies as well as low complement. she receives intravenous immunoglobulin and methylprednisolone and eventually her vaginal bleeding slows and her counts recover. she begins sle therapy with hydroxychloroquine and azathioprine. due to the presence of café au lait macules on her exam, a genetics evaluation is performed and the patient is also diagnosed with nf . to date, there are seven cases of sle with nf reported in the literature, only two of which are pediatric cases. there are no reports of the combination of sle, nf , and es. conclusion: es is a rare hematologic manifestation of sle but can be the initial presentation of this disease. one large study estimates % of childhood-onset sle cases are observed to have es. screening for sle should be considered in all es patients even in the absence of typical clinical findings. association of nf and sle has been rarely described. whether this association reflects a causal relationship or is coincidental needs more investigation. (lube, ped blood & cancer, ) . university of california san diego, la jolla, california, united states background: high grade glioma (hgg) has poor outcomes in adults and children. extraneural metastases are very rare in hgg, and poorly characterized with only a few small case series in adults and only isolated case reports in pediatrics. no genomic data has previously been published for any children with hgg who develop extraneural metastases. objectives: our objective is to describe the natural history of two children with hgg and bony extraneural metastases, comparing their clinical characteristics as well as whole exome sequencing data for both tumors. this information would suggest similar patients should be monitored closely for extraneural metastasis and may benefit from more systemic therapy. design/method: we present a case series of two patients who presented with hgg and had development of bony metastases less than six months after initial diagnosis. both patients had molecular profiling with whole exome sequencing (wes). the first patient was an -year-old male with a tumor found in the left lateral ventricle invading into the fornices, hypothalamus, and left midbrain, who had subtotal resection. bony metastasis were found at . months after diagnosis, and he died months after diagnosis. he initially received radiation, followed by nivolimuab. the second was a -year-old female with a tectal/pineal tumor and multiple spinal cord metastases, who had subtotal resection. she developed bony metastasis at . months after diagnosis and died months after diagnosis. her histologic diagnosis was pineoblastoma, revised to hgg after whole exome sequencing. she received craniospinal radiation followed by chemotherapy per acns (cisplatin, vincristine, and cyclophosphamide) for cycles. when she failed to respond satisfactorily to this therapy, wes of tumor was performed and the findings were consistent with hgg. treatment was transitioned to temodar and lomustine after hgg diagnosis was given. she had ongoing progressive disease despite this therapy as well as trials of nivolumab, everolimus, and vorinostat. neither patient had extraneural metastasis at presentation. in both tumors, whole exome sequencing identified the h f a k m mutation. both tumors also had additional known mutations associated with hgg but no other overlapping mutations. this case series represents the first description of the genetic alterations of pediatric hgg patients who developed extraneural metastases. while h f a k m is a common mutation in pediatric midline hgg, especially dipg, and is associated with more aggressive disease, there has not been an association with extraneural metastasis prior to this series. background: deferiprone-induced agranulocytosis is a well -known albeit rare side effect of the drug. incidence of agranulocytosis varies from . - . %, while milder neutropenia is reported in . % of patients treated with deferiprone. deferasirox is unknown to cause such a complication. clinical trials and post marketing side effect monitoring studied possible correlations between different risk factors and development of agranulocytosis. unfortunately, no studies directly addressed a special risk in a community with background of ethnic neutropenia, like oman. objectives: to report on the incidence of neutropenia among omani children with b thalassemia using different iron chelators design/method: a retrospective study conducted on patients < year-old with b thalassemia treated with different iron chelators. electronic patients records were reviewed to detect episodes of neutropenia either mild (anc . -< . /cmm), moderate (anc . -< ), severe < . , or agranulocytosis anc = ). data were collected including sex, age, personal or family history of ethnic neutropenia, iron chelating agent, infective complications, management and outcome. detailed clinical, laboratory ± radiological information were reported for patients who developed life-threatening agranulocytosis. among young patients with b thalassemia, treated between - in squh, neutropenia, was reported in patients ( . %).severe neutropenia was encountered on occasions in patients ( / : . %) ( on deferiprone including episodes of agranulocytosis, on defersirox, on combined chelation, and off chelation). moderate neutropenia was encountered in patients ( / : . %), on occasions: deferiprone ( ), deferasirox ( ), combined chelation ( ), and episodes off chelation. mild neutropenia was more prevalent, encountered in patients ( . %) on occasions ( on deferiprone, on defersirox, on combined chelation, and off chelation) of patients exposed to deferiprone, patients had neutropenia ( %), higher than previously reported. deferiproneinduced agranulocytosis was encountered in patients ( / = . %). three of them had life threatening complications. one patient developed pneumonia complicated by rupture of pulmonary artery aneurysm-massive hemoptysis, who recovered fully after catheter embolization. the second had facial cellulitis and treatment with gcsf was complicated by frequent ventricular extrasystoles. the third had sepsis, disseminated herpes simplex and required admission to icu for inotropic support. in a community with background ethnic neutropenia, neutropenia is more common to be encountered among thalassemic patients, both on and off chelation therapy. careful monitoring of anc and rational choice/modification of chelating agents is required for optimal management of iron overload and to avoid life threatening complications. objectives: this case control study aimed to evaluate the systolic and diastolic cardiac function in groups of children with ti: non transfused group and a group that received early regular blood transfusion comparing them to healthy controls. design/method: thirteen regularly transfused patients with ti with a mean age of . + . years were compared with eight patients who are non-transfused or minimally transfused (< rbcs transfusion/year); mean age . + . years and healthy controls with a mean age of . ± . years. clinical parameters and standard echocardiographic and tissue doppler imaging (tdi) were compared. results: young non-transfused ti patients had a statistically significant higher peak late diastolic velocity of the left ventricular inflow doppler, a mitral valve a wave duration over the pulmonary vein a wave duration ratio and the pulmonary s of s vein s/d velocities ratio compared to the transfused group with p values of . , . , . respectively. in addition, they have a lower e/a ratio of the mitral valve inflow and a larger left atrial to aortic diameter ratio compared to the control group with p values of . and . respectively. the diameters of the right and left outflow tract were significantly larger in the non transfused group with a trend to have a higher cardiac index compare to the transfused group. systolic function was similar in the studied groups and none of the patients had evidence of pulmonary hypertension. young patients with ti who are receiving early regular blood transfusion have normal systolic function. diastolic function assessment revealed indicators of an abnormal relaxation of the left ventricle in the non transfused group which indicate diastolic dysfunction. the abnormalities affected multiple diastolic function parameters which give an indication that the changes are clinically significant. a statistically significant increase in the diameters of the outflow tracts are likely attributed to high cardiac output status in nontransfused ti patients as they had a trend to have a higher cardiac index. these findings support the early commencing of regular blood transfusion therapy for ti patients to prevent serious cardiac complications in adult life. background: in the -week sustain study, crizanlizumab . mg/kg significantly reduced the frequency of scpcs versus placebo ( . vs . , p = . ) and increased the time to first on-treatment scpc ( . vs . months, p = . ) in patients with sickle cell disease (scd). to evaluate time to first scpc in sustain study subgroups and the likelihood of not experiencing scpc for the duration of the trial using post hoc analyses. design/method: sustain was a randomized, double-blind, placebo-controlled, phase study (nct ). inclusion criteria were: scd patients aged - years; - scpcs in previous months; concomitant hydroxyurea use permitted if ≥ months and stable dose for ≥ months. patients were randomized : : to receive intravenous crizanlizumab . mg/kg, . mg/kg, or placebo. study treatments were administered on days and , then every weeks to week , with the final assessment at week . median time to first scpc after first dose was summarized for crizanlizumab . mg/kg or placebo in these subgroups: - or - scpcs in previous months; scd genotype; and hydroxyurea use at baseline. hazard ratios (hrs) for crizanlizumab . mg/kg versus placebo were calculated based on cox regression analysis, with treatment as a covariate. descriptive statistics were used to summarize the frequency of patients who were scpc event-free for the duration of the study by prior scpc events, scd genotype, and hydroxyurea use at baseline. : patients received crizanlizumab . mg/kg and received placebo. there was a meaningful delay in time to first scpc with crizanlizumab . mg/kg versus placebo observed in the entire study population. the effect was present in both scpc subgroups, and the largest treatment difference was observed in hbss scd versus other genotypes ( . vs . months; hr: . ). in patients taking hydroxyurea who experienced - scpcs in the previous year, time to first onstudy scpc was longer with crizanlizumab . mg/kg versus placebo ( . vs . months; hr: . ). a greater proportion of patients treated with crizanlizumab . mg/kg were scpc event-free versus placebo in each of the analyzed subgroups. one third of patients who were taking hydroxyurea and treated with crizanlizumab . mg/kg were scpc event-free during the study versus . % with placebo, possibly suggesting an additive effect. with crizanlizumab . mg/kg, there was a clinically meaningful delay in time to first scpc and an increased likelihood of being scpc-free versus placebo in all subgroups investigated. cincinnati children's hospital medical center, cincinnati, ohio, united states background: shwachman-diamond syndrome (sds) is an inherited marrow failure syndrome associated with increased risk of myelodysplasia (mds) and acute myeloid leukemia (aml). objectives: this multi-institutional retrospective study investigated clinical features, treatment, and outcomes of sds patients who developed mds or aml by central pathology review. design/method: nine individuals presented with aml ( male, female), mds-eb / ( males, females, with mds ( male and female), and one male with isolated persistent somatic tp mutation. one mds-eb and mds patient progressed to aml. median age (years) at diagnosis of mds was (range . - ), mds-eb / was (range . - ) and aml was . (range . - ). complex cytogenetics were noted in / aml cases, with one having normal cytogenetics. complex clonal cytogenetic abnormalities were noted in of mds-eb /eb patients and clonal abnormalities in of mds patients. follow up was available for aml patients; are deceased. received chemotherapy with intent to proceed to hematopoietic stem cell transplant (hsct). four failed to achieve remission and died with disease without proceeding to transplant. one patient proceeded to hsct without prior chemotherapy. four of six transplanted subjects died with relapsed disease. treatment related mortality was largely infectious or gvhd. the sole surviving aml patient had normal cytogenetics, achieved remission with chemotherapy and underwent hscts with separate stem cell infusions due to two primary graft failures. he remains alive in remission more than years after diagnosis. of the mds-eb / patients, underwent ric hsct, three of whom are alive, one died of infection. the fifth patient has stable disease on continued decitabine monotherapy for . years. of mds patients with treatment data, had upfront hsct therapy, upfront chemotherapy and had no therapy. three patients required ≥ hscts all due to graft failure. follow up is available for , of whom are deceased, with relapsed disease. treatment related mortality was largely infectious or graft failure. one individual died of hepatic failure unrelated to mds. seven mds patients are alive in remission. in summary, prognosis is poor for patients with sds who develop aml due to resistant disease and treatment-related complications. better markers for risk stratification are needed to identify patients who would benefit from early transplant. novel therapeutic strategies are urgently needed to improve outcomes of sds patients with mds or aml. background: unlike primary myelofibrosis (pmf) in adults, which is associated with somatic mutations in jak , mpl, or calr, myelofibrosis in children is rare and the underlying genetic mechanisms remain elusive. here we describe families with autosomal recessive congenital macrothrombocytopenia with focal myelofibrosis (cmtfm) due to germline mutations in the megakaryocyte-specific immune receptor tyrosine-based inhibitory motif (itim) receptor g b-b. objectives: to characterize the clinical phenotype, histological features and identify the causative gene for cmtfm. we performed affymetrix snp . genotyping on the index family to identify shared regions of homozygosity by descent. whole exome sequencing (ws) was performed on all three pedigrees to identify potentially causative mutations. we studied affected children from families, with macrothrombocytopenia, anemia, mild leukocytosis and a distinctive pattern of bone marrow (bm) fibrosis centered around clusters of atypical megakaryocytes. affected children had mild to moderate bleeding symptoms and required platelet and red cell transfusions. none showed evidence of extramedullary hematopoiesis, and all were negative for mutations in jak , mpl, and calr. snp genotyping identified multiple statistically non-significant genomic loci, including the region of the major histocompatibility locus (mhc) on chromosome p (lod = . ). we focused on this region because affected individuals in two families shared a common homozygous human leukocyte antigen (hla) type and had congenital adrenal hyperplasia (cah) due to -hydroxylase (cyp a ) mutation; the cyp a and hla loci are located at p . and p . - p . . wes revealed homozygous frameshift mutations in the megakaryocyte and platelet inhibitory receptor g b-b, encoded within the candidate linkage region. we identified two distinct g b-b frameshift mutations (c. _ + dup; p. fs and c. inst; p. fs) in individuals within these three families. no other mutations that segregated with the phenotype were identified. to validate g b-b as a potential disease-causing gene, we evaluated g b-b expression in bm biopsy specimens from affected patient and control samples by immunohistochemical staining using a monoclonal antibody. g b-b was strongly s of s and selectively expressed in megakaryocytes of control samples, but completely absent in clinically affected individuals. a murine knockout that lacks g b-b has a strikingly similar phenotype with macrothrombocytopenia, myelofibrosis and aberrant platelet production and function, further affirming the causality of g b-b mutations. we showed that autosomal recessive loss-offunction mutations in g b-b cause cmtfm, uncovering the molecular basis of this rare disease. loss of g b-b-dependent inhibition of megakaryocyte activation likely underlies the distinctive focal myelofibrotic phenotype and might be important in other forms of marrow fibrosis. cardinal glennon, saint louis, missouri, united states background: intrauterine transfusion is the method of choice for management of fetal anemia due to red blood cell alloimmunization. despite the decrease in prevalence of anemia due to rhesus d alloimmunization with prophylactic administration of anti-rhd immunoglobulin in rh d negative patients, maternal red red blood cell alloimmunization with other type of red blood cell antigens remains an important cause of fetal anemia. newborn who received intrauterine transfusion for hemolytic disease may have prolonged postnatal transfusion requirement. objectives: -to evaluate clinical outcome of fetuses and newborns who received intrauterine transfusions. -to determine the need of packed red blood cell transfusions until months of age. we conducted a retrospective case series study of all intrauterine transfusions due to anemia secondary to red blood cell alloimmunization performed in our regional center ssm in st louis missouri, between april and january . we evaluated the indications, diagnosis, gestational age, and frequency of intrauterine transfusions, along with the infant's gestational age at birth, duration of admission, timing of blood transfusion and monitoring of hemoglobin. results: intrauterine transfusions were performed in patients. the most common causes of alloimmunization were due to d antibodies (n = , %) and kell antibodies (n = , . %). the median gestational age of the first intrauterine transfusion was . weeks, and the median pre-transfusion hemoglobin was . g/dl. the gestational age at the first intrauterine transfusions was found to be significantly correlated with the number of postnatal transfusions (r = . . p = . ). the median gestational age at birth was found to be weeks ( . - . weeks), with a hemoglobin of . ( . - . ). in our population, patients ( %) received postnatal transfusions, of which were during the first weeks of life, and close monitoring follow up with a hematologist was established in patients at their discharge from the nursery/nicu. one neonatal death occurred and severe morbidity due to severe anemia occurred in one infant. despite the continuing risk factor for persistent anemia, only patients had follow up hemoglobin monitored by their primary care provider. conclusion: infants with anemia due to red blood cell alloimmunization treated with intrauterine transfusion should be monitored closely via regular complete blood count for persistent anemia due to suppression of fetal erythropoiesis. sebastian hesse, piotr grabowski, juri rappsilber, christoph klein dr. von hauner childen's hospital, lmu university hospital, munich, munich, germany background: neutrophil granulocytes are the most abundant leukocytes in the peripheral blood. validated diagnostic options for these cells are limited, leaving many patients with functional neutrophil defects without a defined diagnosis. objectives: here we evaluate proteomics as a new diagnostic tool to investigate defects of neutrophil granulocytes. we analyzed neutrophil granulocytes from children with severe congenital neutropenia (scn) associated with elane mutations, children with chronic granulomatous disease (cgd) with cyba ( ) or cybb ( ) mutations and children with leukocyte adhesion deficiency (lad) due to itgb mutations. in addition we collected samples of children with genetically undetermined neutrophil defects. neutrophils from healthy individuals served as controls. cells were isolated from fresh venous blood using negative selection (purity > %). whole cell proteome analysis was done by data-independent acquisition. showed a correlation coefficient of ∼ . . principal component analysis demonstrated unequivocal separation of the proteome of healthy and diseased cells. differential expression analysis showed minimal proteome aberrations in lad with deficiency in cell surface receptors and upregulation of alpl (total downregulated proteins: / total upregulated proteins: ). analysis of neutrophils from cgd patients also showed limited proteome aberration. cyba and cybb were both diminished independent of genotype, whereas protein clusters around a stat / centered network were increased (total down: / up: ). neutrophils with elane mutations showed the gravest proteome disturbance (total down: / up: ) with an upregulated translational apparatus (srpdependent ribosomes and protein folding complexes) and increased mitochondrial proteins. proteins of each granule subset were dysregulated and metabolic pathways upregulated. a detailed analysis of the proteome from patients with genetically undefined diseases is currently ongoing. one patient with clinical phenotype of cgd was found to have no mutations of nadph oxidase members in whole exome sequencing but critically low levels of ncf on protein level. heterozygosity mapping showed autozytocity in the ncf region warranting current efforts to sequence promoters and intronic regions of the gene. mass spectrometry based proteomics promises exciting new insights into monogenic disease of neutrophil granulocytes and may offer new diagnostic options, in particular in synergy with genome sequencing. by virtue of our international care-for-rare alliance, open to new partners, we hope that our proteome focus may lead to better delineation of as yet unknown disease of neutrophil granulocytes. background: warm autoimmune hemolytic anemia (aiha) is an igg mediated disease. although it can be post-viral, it is often idiopathic and can also be a forme fruste for malignancy or an autoimmune disease. initial management includes steroids. it often relapses on steroid wean and can be refractory to the use of second line treatment such as rituximab. objectives: abatacept (ctla- -ig fusion protein, ctla- mimetic) has been used to ameliorate autoimmune manifestation associated with ctla- haploinsufficiency. we used abatacept as a novel therapeutic agent to manage patients with refractory aiha. design/method: a retrospective case series of two patients at phoenix children's hospital with severe refractory aiha. results: patient , a previously healthy year old female, presented with weeks of icterus, fatigue, and hemoglobinuria. spleen was enlarged cm below the costal margin. laboratory evaluation demonstrated: hemoglobin . g/dl, mild leukopenia /microliter, platelets , /microliter, reticulocytosis . %, positive direct coombs' test, mycoplasma igm and igg positive. bone marrow evaluation showed a hypercellular marrow. she continued to need packed red blood cell (prbc) transfusions despite receiving high dose steroids, ivig and rituximab from may-july . in august, she started sirolimus decreasing her transfusion requirement. after starting abatacept ( mg/kg/dose bi-monthly for three doses and then monthly) in october, she maintained hemoglobin of - g/dl without transfusion. patient , a previously healthy month old male, presented with one week of progressive fatigue, jaundice, and poor feeding. splenomegaly was absent. laboratory evaluation revealed hemoglobin . g/dl, leukocytosis , /microliter, platelets , /microliter, reticulocytosis . %, negative direct coombs' test, and non-specific reactivity on antibody screen. evaluation for inherited hemolytic anemia including a next generation sequencing panel was negative. further evaluation by blood bank showed + positive coombs' for c d due to a warm antibody. cold agglutinin disease was ruled out. bone marrow evaluation was normal. he received high dose ivig as a steroid sparing agent but continued to require prbc transfusions weekly. when prednisone did not seem to slow down hemolysis, treatment with abatacept was initiated and he has not required transfusions for two months. steroids are being weaned. we present successful treatment of two refractory aiha cases with abatacept. patient is steroid and transfusion free and continues on monthly abatacept and sirolimus. patient is also transfusion free and continues on a steroid taper. ctla- is crucial for suppressive function of treg cells. abatacept by binding to cd / seems to enhance treg activity ameliorating autoimmune hemolysis. children's minnesota, minneapolis, minnesota, united states s of s background: transfusional iron overload is common in patients receiving chronic red cell transfusions. as a result, iron chelation is required to minimize toxicity from iron overload. chelation with a single agent can be inadequate at controlling or reducing iron burden. when combination therapy is required deferoxamine may be added to oral chelation. deferoxamine is generally given subcutaneous over - hours for - days a week at - mg/kg/day. many patients struggle to remain compliant with this schedule which has prompted trials of intravenous high-dose (hd) deferoxamine. prior reports of short-term hd deferoxamine have shown minimal side effects however, prolonged use of hd deferoxamine has known toxicity. when compliance is a concern, our center has used hd deferoxamine infusions at mg/kg/hr x hours every to weeks. objectives: evaluate the safety and efficacy of hd deferoxamine at our institution to help guide future therapy. design/method: a retrospective review was completed of patients previously treated with hd deferoxamine between april and september at children's minnesota. final sample included patients ages to years with underlying diagnosis of thalassemia ( ) and diamond-blackfan anemia ( ). deferoxamine infusions were given for hours every - days with a mean length of treatment of days. results: all patients were on combination therapy with deferasirox, however deferasirox was held during deferoxamine infusion. mean pre-deferoxamine liver iron concentration (lic) was . mg/g and mean post lic was . mg/g (p = . ). ferritin mean pre-deferoxamine was ng/ml compared with mean post ng/ml (p = . ). two patients had possible allergy, leading to deferoxamine discontinuation. one patient developed hives, eye swelling and cough while the other had emesis and cough. another patient experienced facial nerve palsy of unclear etiology, which did not recur with resumption of deferoxamine. no respiratory complications were seen. results showed significant decrease in iron burden following combination therapy with high dose deferoxamine and deferasirox. no significant pulmonary, liver, renal, vision, or hearing toxicities were observed. three patients reported reactions to deferoxamine infusions. however, one of these was able to successfully continue deferoxamine without further incident. short-term, hd deferoxamine was effective at reducing lic in combination with oral chelation but requires further evaluation to assess for potential increased risk of toxicity. short-term hd deferoxamine may be considered in the setting of poor compliance of subcutaneous administration or inadequate chelation with single agent therapy. further studies are needed to clarify ideal dosing, timing and risk of toxicity. background: immune thrombocytopenia (itp) is the most common cause of symptomatic thrombocytopenia in childhood but remains a diagnosis of exclusion warranting further evaluation if atypical findings are present. two male children ( months and years old) with newly diagnosed immune thrombocytopenia (itp) were found on initial evaluation to have persistent elevations of lactate dehydrogenase (ldh), alanine aminotransferase (alt), and aspartate aminotransferase (ast). these serum enzyme abnormalities cannot be attributed to itp. in the setting of thrombocytopenia, elevated transaminases and ldh create diagnostic complexity for the hematology/oncology provider as their elevation raises concern for malignancy, hemolytic disease, and other systemic diseases. to raise awareness about an unexpected pattern of duchenne muscular dystrophy in patients undergoing evaluation for itp. to expand the differential of a hematologist/oncologist when abnormal labs support a nonhematologic diagnosis design/method: this case-series of two patients with their clinical and laboratory findings were discovered with retrospective chart review. results: after a thorough evaluation for hemolytic anemias, liver disease and infectious etiologies was negative, bone marrow and liver biopsies were considered. eventually, both children were found to have severely elevated serum creatine kinase (ck). skeletal muscle has the highest concentration of ck of any tissue. thus, significant ck elevation is almost exclusively attributable to muscle injury and is the most sensitive and specific enzyme for diagnosis of muscle disease. referral to a neuromuscular specialist and further genetic testing confirmed the diagnosis of duchenne muscular dystrophy in both children allowing initiation of appropriate interventions. to date, there is no clear genetic predisposition to itp in patients with muscular dystrophy although further investigation may be needed. hematology/oncology providers should consider obtaining a serum ck to rule out muscle disease in any male child with unexplained elevations of serum ldh and/or aminotransferases, as it provides an easy and inexpensive, non-invasive approach to screening. additionally, clinical history and physical examination can aid in the diagnosis of muscular dystrophy, with gross motor delay, abnormal muscle bulk, gower's sign, and proximal muscle weakness all possible findings. objectives: to identify the range of cbcs in patients with ds without infections, hematologic or immune disorders and to create more accurate reference ranges for total white blood count; hemoglobin; hematocrit; mcv; platelet count and absolute neutrophils (anc), lymphocytes, monocytes, eosinophils, and basophils. design/method: a retrospective investigation of healthy pediatric patients with ds who received a cbc between and as part of their medical care at a single, large, pediatric teaching hospital. the study group consisted of children with ds (male = , . %; mean age = . years, sd = . ) at time of blood draw. initially children were reviewed for possible participation in the study; however, patients were excluded due to not meeting the study's inclusion criteria. descriptive statistics were performed on demographic and clinical characteristics. kruskal-wallis h tests, anova, and t-tests were run to determine the significant associations between independent means. results: a significant difference in absolute neutrophils between racial groups, f( , . ) = . , p = . , was observed. there was an increase in anc from . +/- . with african americans to . +/- . in the other racial groups and to . +/- . with caucasians. differences were also found in anc in hispanics/latinos versus non-hispanic/latinos. the results were higher in non-hispanics and latinos, a significant difference of -. ( % ci, -. to -. ), t( ) = . , p = . . preliminary kruskal-wallis h tests run determined that there were significant differences between age groups for total white blood cell, hemoglobin, hematocrit, platelets, lymphocytes and anc. further studies are being run to evaluate in which age groups these differences lie and create reference ranges by age, race and sex. conclusion: among patients with ds, there are differences between racial groups and age groups. this data has been compared to previously established reference ranges for cbcs, but we are currently establishing healthy cbc controls which we will use to validate the reference ranges. these ranges will be published to help guide providers in workup and management of patients with ds. background: transfusion is a critical part of the care provided in the neonatal intensive care unit, but it is not without risks. low birth weight and premature infants can become anaemic from an immature haematopoietic system and frequent phlebotomy. these infants often receive multiple red blood cell transfusions. identifying infants more likely to require such intervention is important in ensuring the appropriate usage of this scarce resource. to determine whether birth weight, gestational age, gender, length of stay and mode of delivery can predict red cell concentrate (rcc) transfusion, units required, donor exposure and time to exposure. design/method: a retrospective chart review of all infants born below weeks gestation and/or birth weight less than , g who received a red blood cell transfusion between july and july in the cork university maternity hospital neonatal unit. results: infants met the inclusion criteria, ( . %) received a rcc transfusion. our study showed lower gestational age (p< . ) and lower birth weight (p< . ) infants are more likely to be transfused. donor exposure increases with a lower birth weight (p = . ). multivariate analysis showed infants with a lower gestational age (or - . per day; p< . ); lower birth weight (or - . per g; p< . ) and a longer length of stay (or . per day; p< . ) are more likely to receive a higher number of rcc transfusions. the time to first rcc transfusion is shorter in those with lower birth weight (or . per g; p< . ) and lower gestational age (or . per day; p< . ). gender and mode of delivery were not found to be predictors of red blood cell transfusion in this study. conclusion: low birth weight and premature infants are more likely to receive a rcc transfusion during admission to the neonatal unit. our study highlights predictors of rcc transfusion, donor exposure and time to transfusion. these can be used in identifying at risk infants, counselling parents and in anticipating transfusion requirements. emily southard, r. grant rowe, david williams, akiko shimamura, taizo nakano children's hospital colorado, aurora, colorado, united states background: the mecom locus encodes transcription factors that regulate hematopoietic stem cell self-renewal and maintenance. overexpression of mecom has been noted in - % of acute myeloid leukemia, several solid tumors, and denotes a poor prognosis. mutations that reduce mecom expression or that disrupt protein function, however, have been implicated in the development of bone marrow failure (bmf) through undefined pathways. an association between mecom mutations and radioulnar synostosis with amegakaryocytic thrombocytopenia (rusat) syndrome has been reported, however further characterization of this phenotype has yet to be explored. to characterize the phenotypic spectrum of a cohort of pediatric patients with novel mecom mutations. we performed a retrospective review of five patients with mecom mutations who were referred to hematology at children's hospital colorado or boston children's hospital. clinical, laboratory, and genetic data was collected on subjects and available family members. results: four of subjects were identified in infancy presenting with congenital cytopenias or physical dysmorphisms that prompted broad genetic screening. platforms for genetic detection included microarray, targeted genetic panels, and whole exome sequencing. three of subjects with cytopenias presented with congenital thrombocytopenia, of whom rapidly progressed to severe aplastic anemia. four of subjects presented with congenital anomalies, of whom demonstrated radioulnar synostosis. additional dysmorphic features identified include craniofacial (low set ears x ), cardiac (pda x , vsd x , aortic root dilation x ), pulmonary (pulmonary hypertension x , arteriovenous malformations x ), and developmental delay. one subject presented at age years with acute pancytopenia, hypocellular marrow, no dysmorphisms, and a mecom variant of unknown signif-icance. the identified mecom mutations include one . mb deletion involving several genes including mecom, one variant affecting a splice acceptor consensus sequence predicted to disrupt splicing, and three novel missense mutations, tyr cys, arg thr, and tyr cys, all of which were absent from public databases and were predicted in silico to be deleterious. we describe the phenotypic spectrum of patients with novel mecom variants. a subset of patients lacked radio-ulnar synostosis and had presence of additional systemic anomalies, demonstrating a varied clinical phenotype that is not isolated to rusat syndrome. a centralized publically accessible database to share clinically annotated mecom variants, together with analysis by experts in mecom function would advance our understanding of the clinical interpretation of mecom variants. mecom should be considered in the differential diagnosis of bone marrow failure and we advocate for the inclusion of mecom in targeted sequencing panels. cairo university, cairo, egypt background: beta thalassemia is regarded as a serious public health problem in the mediterranean region, southeast asia, and the middle east. however, very few studies have been conducted to assess the quality of life (qol) among thalassemia major patients. objectives: to assess the quality of life among b-thalassemia major patients using short form (sf)- questionnaire and to determine the factors associated with their quality of life. design/method: a cross-sectional study was conducted among thalassemia major patients who were attending the hematology outpatient clinic at cairo university hospital, during the study period. data were collected between october and march . the quality of life was assessed for patients aged ≥ years. the mean age of the studied group was . ± . years. the majority ( . %) had one monthly blood transfusion. the mean total score of sf- was . ± . . general health perception domain was the most affected domain with mean score, while vitality was the least affected one. there was no statistically significant difference between males and females regarding different quality domains except for vitality where the mean score was significantly higher in males than females (p = . ). age at onset of disease, and at first blood transfusion were the most documented factors positively correlated with the quality of life among the enrolled thalassemia patients. conclusion: the quality of life in thalassemia major patient was found to be compromised. all thalassemia patients should undergo assessment of the quality of life so that interventions focusing on the affected domains can be implemented. background: international adoption of children with special needs has become more prevalent in recent years leading to tremendous growth in the number of u.s. thalassemia patients adopted from foreign countries. currently % of the , thalassemia patients registered in the cooley's anemia foundation (caf) patient database have been adopted from foreign countries, primarily china. as this population continues to grow, further information is needed in order to provide these families with best supportive care. the primary goal of this study is to characterize the socio-demographics and health statuses of adopted children with thalassemia and their families. a secondary goal is to describe adoptive families' motivations, experiences, challenges, and support resources. design/method: a redcap survey was accessed by families of adopted children with thalassemia through the caf website and caf social media from january to august . following a four-question screen, eligible subjects were directed to complete an adoption questionnaire. families who had at least one adopted child with thalassemia receiving care at a participating thalassemia treatment center or hematology office in the u.s. were considered eligible. descriptive statistics were analyzed using sas . . respondents who were ineligible or who provided incomplete data were removed from the dataset prior to analysis. of survey respondents, qualified and completed the survey. these households had adopted a total of children with thalassemia ( . % male), most from china ( . %), where they had been living in orphanages ( . %). legal guardians identified primarily as christian ( . %). the majority had completed post-secondary education ( . %) with reported household incomes greater than $ , ( . %). most adoptive families were connected to an adoption group or community including online groups, local support groups, and adoption networks ( . %). commonly cited challenges were: ) volume of frequent medical appointments, ) insufficient support from their local care centers, and ) financial burdens. the reality of care for the population of adopted patients with thalassemia in the u.s does not seem to match the expectations set by their providers. we are hopeful this data will be used to assist adoptive families navigating the complexities of thalassemia care. the findings suggest that this population would benefit from additional outreach, education, guidance, and advocacy resources -especially in the early stages of adoption and during initiation of post-adoption medical care. background: in many higher-income countries, thalassemia major has become a chronic disorder; many outcomes are different in emerging countries with more limited resources. most analyzes of health-related quality of life (qofl) in thalassemia have been conducted in high-income settings. objectives: to assess the impact of health status on qofl in thalassemia patients in an emerging country. we assessed qofl in randomly-selected patients ( thalassemia major; with hemoglobin e thalassemia; five thalassemia intermedia) at the national thalassemia center in kurunegala, sri lanka where approximately patients are managed. treatment is free, but compared to north america/europe, access to tertiary staff and other resources are limited. overall, control of body iron as estimated by serum ferritin concentration (mean± sem, ± g/l) was not optimal in many patients. to understand the impact of health status on qofl, we used the sf v health survey, analyzing scores of physical function, pain, general health, social functioning, emotional and mental health, to generate overall physical and mental component scores. results: compared to reports from higher-income countries (american journal of hematology ; : - ), physical function scores (mean±sd, . ± . ) were similar in sri lankan patients; indeed, in three categories (physical role, social function, emotional role), sri lankan scores were slightly higher. by contrast, compared to scores from higherincome settings, those estimating bodily pain, general health, and mental health were significantly lower, resulting overall in a significantly lower physical component score in sri lankan patients. male sri lankan patients reported higher scores than females, and somewhat surprisingly, in four categories (physical function, physical role, social function and emotional role) reported higher scores than those obtained in higher-income settings. lower scores in physical functioning, leading to an overall lower physical component score, were recorded by females. patients with hemoglobin e thalassemia reported generally poorer qofl than those with thalassemia major. the lack of differences in qofl in patients with "high" and "low" hemoglobins was likely related to low pre-transfusion hbs (mean±sem, . ± . g/dl) in nearly all patients. these early data in a small cohort of thalassemia patients in an emerging setting suggest that in many patients bodily pain, reduced mental health, and poorer views of general health affect overall qofl. prospective studies in larger cohorts including evaluation of adequacy of transfusions and chelation therapy, complications, and overall accessibility of care may guide approaches to improve qofl in lower-income settings of thalassemia care. geetanjali bora, anand prakash dubey, tarun sekhri, mammen puliyel, aparna roy maulana azad medical college, new delhi, delhi, india background: in the last two decades, the presence of osteopenia has been described in optimally treated patients with transfusion dependent thalassemia, the pathogenesis of which seems to differ from osteopenia in non-transfused patients. the prevalence rate of low bone mineral density (bmd) in pediatric population is highly variable amongst studies done worldwide. furthermore, the role of metabolic and endocrine factors in determining bone mass in this population is not well understood. objectives: to assess bmd in subjects with transfusion dependent beta thalassemia by dual-energy-x rayabsorptiometry and find its co-relation with clinical, biochemical and hematological parameters. design/method: this is a comparative cross-sectional study and includes patients with transfusion dependent beta thalassemia between ages to years enrolled from a thalassemia day care center in the year - . at the time of enrollment age, sex, bmi z scores, pubertal staging, duration and type of chelation therapy were noted. enrolled subjects were scanned for bmd at lumbar spine l - and left femoral neck using dexa scan. the bmd was expressed in mean values and z scores. age, bmi, ethnicity and gender matched historic controls were used to generate z scores. ml of pre transfusion fasting venous blood samples were obtained to test for serum calcium, phosphate, alkaline phosphatase, pth, thyroid function panel, serum ferritin and serum igf- levels. mean values for pretransfusion hemoglobin and serum ferritin over last months were calculated. results: total no of subjects , median age . years, male ( %), female ( %), ethnicity % asian, bmi < rd centile ( %), pre pubertal %, all receiving transfusion and chelation therapy. prevalence of low (z score < - sd) and very low (< - . sd) bmd was %, % at l -l respectively and %, % at left femoral neck respectively. there was trend of lower bmd z scores with advancing age. statistically significant co-relation (p value < . ) was found between low bmd and low mean pretransfusion hemoglobin, serum phosphate, igf - and vitamin d levels conclusion: a sizable proportion of children and adolescents with transfusion dependent thalassemia have suboptimal bone mineral density and this decline may start as early as - years of age despite being on transfusion regimen highlighting the importance of yearly dexa screening and optimization of pre-transfusion hemoglobin, vitamin d and igf levels. vanderbilt university medical center, nashville, tennessee, united states background: it is well described that iron deficiency anemia (ida) can co-present with thrombocytosis or thrombocytopenia, though cases of thrombocytopenia are less frequent than thrombocytosis. prior reports of thrombocytopenia have included adult and pediatric patients with menorrhagia ( - ), menorrhagia due to uterine fibroids ( ), or other gynecologic abnormalities ( ). our cases highlight the pattern of ida, thrombocytopenia, and menorrhagia in the setting of significant menstrual clotting without observed gynecologic abnormalities in african-american adolescents. objectives: to describe the clinical course of three adolescent females with severe ida, menorrhagia, and thrombocytopenia. results: our cases included three female african-american patients ages - who presented with severe anemia and concurrent thrombocytopenia in the setting of menorrhagia. all three patients reported heavy and prolonged menstrual cycle bleeding with significant clots. two of the three were admitted for transfusions at presentation and noted to have significant menstrual bleeding with continued blood loss requiring additional transfusions until bleeding was controlled with estrogen therapy. these two patients were evaluated with pelvic ultrasounds revealing a prominent endometrium in both patients and hyperechoic material consistent with a clot in one patient. average hemoglobin on presentation was . gm/dl ( . - . ), average platelet count was , /mcl ( , - , ), and average mcv was ( - ). all had severe iron deficiency with an average ferritin of ng/ml ( - ) subsequently treated with oral iron. one patient had a prior history of ida that required transfusion and had subsequent normalization of her complete blood count. two patients had subsequent thrombocytosis before normalization of their platelet counts. two patients received platelet transfusions: one due to recent neurosurgical intervention with a higher goal platelet count and the other to help control menstrual bleeding after a nadir platelet count of , . a review of the clinical history and red cell indices pointed to ida and ongoing blood loss from menorrhagia as the reason for the bicytopenias. the thrombocytopenia in these cases may have been exacerbated by consumption of platelets in the significant clots all three patients reported. it is reasonable to treat with iron supplementation and supportive care which may include transfusions or management of menorrhagia with oral contraceptives or other hormonal methods. background: sickle cell disease is one of the most common inherited red blood cell disorders, yet many are not aware of their carrier status. the american college of obstetricians and gynecologists' guidelines recommend that pregnant women of african, mediterranean and southeast asian descent be screened for hemoglobinopathies with a cbc and hemoglobin electrophoresis . however, adherence to this practice and frequency of improper screening with sickledex is unknown. proper screening and counseling can impact families' knowledge, allowing for establishing relationships with pediatric hematology providers earlier. objectives: we sought to assess prenatal hemoglobinopathy screening practice patterns and methods of obstetrics & gynecology (obgyn) and family medicine providers in the nyc regional area. design/method: a cross-sectional electronic survey was administered to obgyn and family medicine practitioners from four nyc institutions. questions focused on prenatal hemoglobinopathy screening practices using case scenarios with variations on parental trait status and ethnicities. chisquare analyses were used to compare the two provider groups on categorical variables. there were total responses; surveys were complete, of which were obgyn and family medicine providers. respondents were mainly from academic medical centers, with the majority being faculty ( % of the obgyns and % of family medicine). no significant difference was found in frequencies of screening patients with a positive family history of a hemoglobinopathy. when asked about screening practices for patients without a personal/family history of a hemoglobinopathy, % of obgyns versus % of family medicine providers "always" screened for hemoglobinpathies (p = . ). when analyzed by ethnic background, there were significant differences by group in screening patients of white ( % vs %), black ( % vs %), mediterranean ( % vs %), and asian descent ( % vs %) (p≤ . for all). however, in cases where the hemoglobinopathy carrier status of both parents was known, there was no difference in screening with a hemoglobin electrophoresis. furthermore, > % of all respondents use sickledex for screening in the case scenarios. conclusion: this pilot survey highlights a difference in the methods and likelihood of prenatal hemoglobinopathy screening based on the type of prenatal care provider. screening differences can lead to variations in prenatal guidance, diagnostic procedures, informed decision-making and knowledge of families referred to pediatric hematology clinics. this is the first study analyzing prenatal screening for hemoglobinopathies in obgyn and family medicine. improving prenatal screening practices by collaborating with hematologists may decrease health care disparities and allow for earlier relationship building with pediatric hematology. . acog, opinion# , poster # hermansky-pudlak syndrome: spectrum in oman background: hermansky-pudlak syndrome (hps) is a rare autosomal recessive disorder, characterized by the triad of oculocutaneous albinism, a hemorrhagic diathesis resulting from storage pool-deficient platelets, and accumulation of ceroid/lipofuscin-like material in various tissues. before , nine different types of hermansky-pudlak syndrome were identified, which can be distinguished by their signs and symptoms and underlying genetic cause. in , a tenth type was defined based on mutations in the ap d gene. hps type is characterized in addition by severe neutropenia and recurrent sinopulmonary infection. the disease is more common in puerto rico, and this is the first report from oman. to describe the clinical, laboratory and genetic characteristics of hps sub-types in oman, including the first cases of hps type . design/method: this is a retrospective study, including cases with hps that had been suspected clinically and confirmed through genetic mutation analysis. clinical data included sex, age at presentation, initial clinical presentation (skin, eyes, development, neurological involvement, bleeding tendency, recurrent infections) and course of disease. laboratory data (complete blood counts, platelet and absolute neutrophil counts, coagulation screening, platelet function tests by platelet function analyzer, and platelet aggregation studies using different agonist had been recorded. pcr and next generation sequencing for genetic confirmation by testing mutations in hps , ap b , hps , hps , hps , hps , dtnbp ,, bloc s , bloc s genes had been done. results: seven omani cases with hps have been identified ( males and females). their age ranged between (at birth) to years. two patients had hps type , patient had type , while the other cases had hps type . no other sub-types were encountered in oman. all patients were products of consanguineous marriage. one patient had adrenal hge, while the others had mild hemorrhagic phenotype, characterized by recurrent bruising and mild epistaxis. laboratory testing confirmed variable platelet aggregation defects with different platelet agonists. all patients had characteristic hypopigmentation, iris transillumination, nystagmus, and foveal hypoplasia. both patients with hps type had the same homozygous mutation in the ap b gene (c. _ delta), and presented with severe neutropenia. early diagnosis and initiation of gcsf on one of them improved outcome and prevented the development of complications. late diagnosis in the other patient resulted in the development of bronchiectasis as a result of recurrent sinopulmonary infections. background: sickle cell disease (scd), a genetic disorder characterized by defective sickle hemoglobin (hbs), triggers red blood cell sickling, hemolysis, vaso-occlusion, and inflammation. ischemic injury from scd starts in infancy and accumulates over a lifetime, causing pain, fatigue, and progressive end-organ damage that culminates in early mortality. voxelotor (gbt ) is an oral, once-daily therapy that modulates hemoglobin's oxygen affinity, thereby inhibiting hemoglobin polymerization. objectives: to assess the safety, pharmacokinetics, and efficacy of voxelotor in pediatric patients with scd. design/method: this ongoing study is being conducted in parts: part a: a single dose of voxelotor mg in pediatric and adolescent patients; part b: multiple doses of voxelotor mg/d or mg/d for weeks in adolescents. part b's primary objective is to assess the effect of voxelotor on modifying anemia. secondary objectives include measuring other markers of disease modification, such as hemolysis; daily scd symptoms, using a patient-reported outcome (pro) measure; and safety. results: as of november , , patients ( females) had received voxelotor mg and patients ( females) had received voxelotor for ≥ weeks. the median age for the patients was years, % were receiving hydroxyurea (hu), and % had ≥ painful crises in the past year. data for hemolysis measures are available for patients who received voxelotor for weeks. six of the patients achieved a hemoglobin (hb) response of > g/dl increase. laboratory markers of hemolysis improved concordantly; the median reductions in reticulocytes and indirect bilirubin were % and %, respectively. ten of patients showed reduction in total symptom scores (tss) at week , with a % median reduction in tss from baseline. there were no treatmentrelated serious adverse events (aes) or drug discontinuations due to aes. voxelotor mg for weeks in adolescents with scd, the majority receiving hu, demonstrated consistent, sustained efficacy on hb levels and measures of hemolysis; > % of patients showed a > g/dl improvement in hb. improvement in tss in mildly symptomatic patients suggests that the pro is sensitive to treatment effect and supports use in the ongoing hope phase study. voxelotor's reassuring safety profile is consistent with results in adults. these interim results support ongoing clinical evaluation of voxelotor as a potential disease-modifying therapy for adults and children with scd. supported by global blood therapeutics. background: acute kidney injury (aki) is a common complication in sickle cell disease (scd), and a potential risk factor for sickle nephropathy. aki is associated with acute decline in hemoglobin (hb) during vaso-occlusive pain crisis and acute chest syndrome (acs). it is unclear which pathologic factor plays a stronger role in aki development during hb drop: increase in free heme during vaso-occlusive events secondary to hemolysis or hb decline itself. objectives: to investigate if hb decline alone is associated with aki, we tested if the renal function of patients with scd worsened during parvovirus b -induced transient aplastic crisis (tac), in the absence of accentuated hemolysis. design/method: with irb approval, a retrospective study of patients who had laboratory confirmed parvovirus-b was conducted. serum creatinine (scr), both during and within months from the tac event, was collected. comparisons of the clinical and laboratory characteristics were analyzed using the wilcoxon test for continuous variables. aki was defined as an increase in scr by ≥ . mg/dl or a % increase in scr from baseline. to evaluate differences in change in hb on aki risk, changes in scr during tac were compared to those during pain crisis or acs admissions by fitting a generalized linear mixed model for binary outcome. a comparative sample of acs events and vaso-occlusive pain crisis were used to estimate rates of aki according to hb levels. results: three ( %) of the patients with scd developed aki during tac. no association was identified between change in hb from baseline to tac event (p = . ). no cases of aki were identified until hb decreased < . g/dl or the change in hb was ≥ . g/dl from baseline. next, we developed a model to evaluate the impact of change in hb from baseline for patients admitted with tac, pain crisis or acs on aki. with a g/dl decrease in admission hb from baseline, patients with tac had a % probability of developing aki, while acute chest syndrome and pain crisis would have a % and % probability, respectively. our data suggest that aki is still prevalent during parvovirus b -induced tac. however, the risk of aki during a tac event is and times lower than that from severe anemia induced by acute chest syndrome and vasoocclusive pain events, respectively. hemolysis-induced anemia during scd crisis appears to have a more significant role in the development of aki as compared to agenerative anemia. background: the natural history of hemoglobin e beta thalassemia (hbethal), the commonest form of severe beta thalassemia worldwide, has been examined in very few longterm studies. previously, we reported findings in hbethal patients in sri lanka. objectives: to evaluate longterm requirements for transfusion and splenectomy, complications and death in hbethal patients. design/method: all available patients were reviewed - times annually over years. results: patients ( %) died, aged (mean ± sem) . ± . years; the (known) causes commonly included iron overload ( ) and infection ( ); patients surviving patients are aged . ± . years. of patients originally classified by severity (group the mildest, and group the most severe, phenotypes), ( %) were assessed as mild (groups and ), of whom transfusions had been discontinued in . ultimately, / ( %) resumed transfusions, often following shifts to increasingly severe phenotypes including increasing intolerance to anemia. age at resumption of transfusions (following a transfusion-free interval of . ± . years) was . ± . years; in the more severe groups and , regular transfusions were stopped in / patients and resumed in / ( %), at younger ages ( . ± . years) and after shorter transfusion-free periods ( . ± . years) than in "milder" patients. mid-parental height (mph) was ultimately achieved in %. patients ( %) were splenectomized; updated analysis of responses to splenectomy (originally "group " patients), showed that splenectomy (at . ± . years) was followed by an extended, but impermanent, transfusion-free interval ( . ± . years); % patients resumed transfusions, usually related to exercise intolerance or poor growth. in groups and , complications of anemia and ineffective erythropoiesis, including leg ulcers (in % and %) and gallstones ( % and %), were more frequent than in groups and ; fractures were observed ( - %) across all groups, except for regularly-transfused group patients ( %). pulmonary artery pressures > mm were recorded in % patients. evaluation of patients with hbethal requires observations over years, without which definition of patients as "mild" or "severe" may be misleading. while in many patients transfusions may be withheld or reduced in frequency, troublesome complications may surface with advancing age even in "milder" patients. although individual consideration of transfusion requirements is critical, the availability of effective chelation, where this can be provided without prohibitive cost, may alter the balance of risks and benefits of regular transfusions in hbethal. (premawardhena a. lancet ). background: social determinants of health (sdh) are environmental and socioeconomic factors, such as access to food and housing that affect health outcomes. pediatricians are increasingly screening for sdh as part of primary care visits, however less is known about screening for sdh in pediatric hematology. evidence suggests that sdh play a role in disease severity for children with scd, who face significant socio-economic and racial disparities. the goal of our quality improvement (qi) project was to increase the percentage of patients with scd who were connected to community resources for unmet social needs. design/method: we based our intervention on the successful implementation of wecare in our institution's pediatric primary care clinic. eligible patients were identified at the start of each clinic session. on arrival the parent was given a self-reported screening tool for six sdh (childcare, education, employment, food, utilities and housing). results were entered in the electronic health record by the physician or social worker who then printed a pre-existing resource list for patients with a positive screen. we used a series of plan-do-study-act (pdsa) cycles to study tests of change. we tracked process measures (percentage of patients screened, percentage of patients with an unmet social need who received a resource sheet), outcome measures (percentage of patients with an unmet social need who connected with a community resource) and balancing measures (staff, patient and provider satisfaction). run charts were reviewed weekly and then monthly to inform further tests of change. examples of pdsa cycles include who gave the paper survey to patients (social worker or physician versus medical assistant) and length of time between surveys ( to months). results: between august and december screening rates improved from % to %. of the patients screened, % report at least one unmet social need; of those % received a targeted list of community resources in the first month of the project, and % in the fifth month. finally, % of patients reached by phone had connected with a community resource within weeks of the clinic visit. we have successfully implemented universal screening for sdh for patients with scd in our urban pediatric hematology clinic without requiring extra staff. next steps include further pdsa cycles to connect more patients to appropriate resources, and tracking improvement in health care utilization outcomes from addressing sdh in this vulnerable patient population. background: the clinical manifestations of sickle cell disease (scd), chronic hemolytic anemia, and vaso-occlusion occur as a direct result of sickle hemoglobin (hbs) polymerization. voxelotor (gbt ) is a first-in-class, oral, oncedaily investigational agent designed to modulate hemoglobin's oxygen affinity in a targeted approach to inhibit hbs polymerization. objectives: to examine the pharmacokinetics (pk), safety, and dosing of voxelotor in children (aged - years) and adolescents (aged - years) with scd from part a of the gbt - study. design/method: gbt - is an ongoing, open-label, phase a study in patients aged - years with scd (sickle cell anemia or sickle beta zero thalassemia). part a of this study (the focus of this abstract) is examining pk of singledose ( mg) voxelotor. pk samples to measure whole blood and plasma voxelotor concentrations were collected up to days following single-dose administration. separate population pk (ppk) models were developed to describe the concentration versus time profiles of voxelotor in whole blood and plasma using nonlinear mixed effects modeling (non-mem, version . ). ppk modeling and physiologically based pk (pbpk) modeling were used to simulate voxelotor pk parameters and support dose selection for future evaluation in younger children. : part a included adolescents ( females; median age years [range - ]) and children ( females; median age . years [range - ]). mean weight was . kg (range - kg) and . kg (range - kg) in adolescents and children, respectively. voxelotor was well tolerated with no drugrelated grade ≥ adverse events (ae) or serious aes. a compartment model with first-order absorption best described the pk of voxelotor (and was the same model structure used for adults with scd). voxelotor pk exposures in adolescents were comparable to those observed in adults, but higher exposures were observed in children. ppk and pbpk modeling support the use of a weight-based dosing strategy in younger children (aged < years) in future trials. adult voxelotor doses can be used in adolescents. however, based on higher pk exposures, a lower weight-based dosing strategy is recommended in children. ppk and pbpk modeling provides an innovative approach to minimize experimental dosing in children and accelerate dose selection of voxelotor in ongoing and future clinical studies. this abstract is supported by global blood therapeutics. background: hydroxyurea (hu) reduces rates of acute complications, and improves long term outcomes in patients with sickle cell disease (scd) and is now fda approved for children. through previous work we have increased the number of eligible patients on hu in our clinic, however accessing a compounding pharmacy remained a significant barrier to hu adherence for infants and children who cannot swallow capsules. objectives: the objective of our quality improvement project was to improve adherence to hu among pediatric patients with scd at our urban safety net hospital by addressing barriers to obtaining liquid hu. design/method: to begin we met with the leadership of our outpatient pharmacy which offers mail order delivery. however, like most retail pharmacies, they do not have the necessary protective equipment to compound liquid hu. through a series of discussions, we began a unique partnership with our institution's inpatient chemotherapy pharmacy who compounds the liquid hu and delivers it to the outpatient s of s pharmacy, who then dispenses liquid hu to families. using a series of plan-do-study-act (pdsa) cycles we tracked adherence by calculating the medication possession ratio (mpr), defined as the percentage of days in a given period of time that each patient had their medication on hand. the mpr for liquid hu mpr among enrolled patients was tracked by pharmacy staff and reviewed monthly. additional pdsa cycles included adding automatic refills and reminder calls by pharmacy staff and improving communication about delivery. we also tracked patient satisfaction. results: between march and december , a total of thirty pediatric patients were enrolled in our program for on-site compounding and free mail order delivery of liquid hu. mpr for liquid hu is currently . % among enrolled patients, significantly higher than the mpr of % reported in the literature, and has risen steadily since the beginning of the project. families are highly satisfied with the program, specifically appreciating the convenience of mail order delivery, saving on delivery fees, and reminder calls when refills were due. by compounding and dispensing liquid hu directly from our institution's outpatient pharmacy we have significantly improved adherence to this hu therapy in our high-risk population. next steps include analysis of change in clinical outcomes for patients enrolled in this program. as adherence to hydroxyurea is associated with decreased acute care utilization and cost, programs such as ours could play a crucial role in reducing the excessive costs and ed utilization among this patient population. background: experience with the iron-chelator deferasirox is reported widely in higher-income settings. by contrast, real-life experiences in emerging countries are infrequently reported. objectives: to evaluate, in a non-trial setting, the real-life response to deferasirox in an emerging country. design/method: in sri lanka's national thalassemia center which manages patients without tertiary staff, quantitative evaluations of body iron or estimates of extra-hepatic iron, the records of patients who began deferasirox in / were retrospectively reviewed. results: baseline assessments (mean±sem) indicated substantial iron loading [serum ferritin (sf) , ± ug/l; serum alt ± . u/l (normal ≤ u/l)]. deferasirox was introduced at low doses ( . ± . mg/kg/day); many patients started at < mg/kg and, after months, doses remained ≤ mg/kg/day in % patients. after months, sf in % patients remained > , ug/l; only by months had (mean) sf declined to < , ug/l ( ± ; p< . ). similarly, mean alt normalized (to ± u/l) only by months. death and complications were not systematically recorded by staff who had been charged, without provision of additional resources, with the introduction of this new drug in hundreds of patients. these results contrast to those in sri lanka's tertiary thalassemia center where, in patients following the introduction of deferasirox ± . mg/kg/day, sf declined rapidly, even in relatively less ironloaded patients (from , ± to , ± g/l after months; p = . ). these findings underscore the importance, during the implementation of new drug regimens in lowerincome centers with marginal resources, for investments in methods to quantitate body iron burden, hands-on educational initiatives to guide day-to-day management by competent but non-expert staff, and data systems to record efficacy, effectiveness, toxicity and compliance. such investment is critical to optimising therapy and improving complications in thalassemia patients worldwide: even in sri lanka, where resources directed to thalassemia management are greater than in most of asia, results in the oldest living cohort (born - ) indicate under-treatment [elevated iron burdens (sf , ± ug/l) and high prevalences of diabetes ( %) and hypothyroidism ( %)]. even in a younger cohort (born - ) which has benefitted from improved treatments, the prevalence of many complications exceeds those reported from high-income settings. over the next decade, and two decades after the who declaration that the impact of thalassemia on global mortality and morbidity is underrecognized, increased investments by governmental and nongovernmental sources will be necessary to improve outcomes for asian patients with thalassemia. background: a major barrier to success in hydroxyurea (hu) treatment of patients with sickle cell disease (scd) is non-adherence. objectives: to optimize hu adherence in patients with scd. design/method: a care model was designed by the sickle cell (qi) team at children's hospital to improve hu adherence among scd patients. the original model included bimonthly family phone contact, monthly dispensing pharmacy phone contact and lab monitoring. adherence measures included obtaining hu from pharmacy monthly, completion of monthly labs, hb f percentage and mcv, and mtd achievement. from / - / , several pdsa cycles refined our care model. a one-year follow-up survey gathered feedback on the care model. the first-year data involved ∼ patients. the biggest improvements resulted from making pharmacy calls before patient/family calls, shipping liquid hu to outlying patients, and tracking call time/content. the qi goal was % hu adherence by / . the % baseline adherence rate increased to % by / , and has remained in that range. the completion rate of patient/parent phone calls increased from % the first month to % at six months. pharmacy prescription pick-up has increased from % to % per month. lack of liquid hu availability was overcome by shipping the medication to the patient's home. parental hesitance to share information by phone, especially with qi team members with whom they had no established relationship, was overcome by having the longtime sickle cell nurse do many of the early calls. however, survey feedback showed families became comfortable with several clinic personnel calling. the calls gave families the opportunity to ask questions about their child and/or get additional information about scd. the calls also provided an opportunity for seasonal flu shot or tcd testing reminders. the surveys gave information on the optimal time of day to reach each family, providing individualization and further increasing the percentage of completed calls. two families surveyed said they no longer needed two calls a month because they were now able to remember to pick up hu, administer it, and get labs on their own. this qi project has not only improved hu adherence, but also fostered health education/counseling, increased patient/parent satisfaction, and enhanced service utilization. medical team member and patient/family comments demonstrate that it has helped build relationships and trust between families and the medical care system. based on survey feedback, we will further individualize care to increase adherence rate and sustain improvements. cincinnati children's hospital medical center, cincinnati, ohio, united states background: the thalassemias are a heterogeneous group of genetic blood disorders caused by mutations that decrease or eliminate the synthesis of the -and/or -globin subunits of hemoglobin. the phenotype of thalassemia depends on the interaction of the -and -globin gene clusters, because both loci determine the -/ -chain balance. for example, a -thalassemia phenotype can be more severe than expected when coinherited with -globin gene triplication (copy number gain), which exacerbates the -/ -globin imbalance. objectives: describe four individuals with an incorrect diagnosis of -thalassemia trait who were later properly diagnosed by comprehensive genetic testing to have -thalassemia intermedia caused by heterozygous -thalassemia mutations coinherited with triplicated -globin loci. design/method: sequence analysis of the -globin (hba /hba ) and -globin (hbb) genes, and copy number variation analysis of the -and -globin gene clusters by multiplex ligand-dependent probe amplification. results: four unrelated individuals of northern european ancestry were evaluated for signs and symptoms not explained by a diagnosis of -thalassemia trait (previously made by a pediatric hematologist), including growth delay, splenomegaly, moderate anemia, marked elevation of hemoglobin f, thalassemic facies, reticulocytosis, and/or indirect hyperbilirubinemia. genetic testing revealed that all were heterozygous ( / ) for the same, single -globin mutation [hbb.c. c>t (p.q *)] and also heterozygous for an -globin triplication ( / anti- . ). their previous diagnoses of thalassemia trait had been made by complete blood counts, hemoglobin electrophoresis, and/or sequence analysis of the -globin genes only. these individuals' phenotypes ranged from moderate anemia only to multiple stigmata of thalassemia, demonstrating the phenotypic variation of a thalassemia genotype. correct diagnosis was made at an average age of . years. a trial of chronic transfusions was initiated for one patient for growth failure. all were educated about the potential for exacerbations of anemia, gallstones, osteoporosis, and iron overload (even without transfusions). parental genetic testing was recommended to assess reproductive risk, because inheritance of this complex genotype can be apparently autosomal dominant. conclusion: heterozygosity for a -thalassemia mutation does not necessarily indicate -thalassemia minor or "trait". when coinherited with -globin gene triplication, a symptomatic form of -thalassemia can occur. correct and timely diagnosis of thalassemia requires careful consideration of the degree of anemia and examination for organomegaly, bony changes, and jaundice. sequence analysis and copy number variation analysis of both the -and -globin gene clusters is key. hematologists need to be aware of this diagnostic possibility and how to test for it to prevent inaccurate or delayed diagnosis. background: the burden of healthcare costs for sickle cell disease (scd) is nationally estimated at over $ billion. the major components of these costs are inpatient and emergency center (ec) visits, many of which are potentially avoidable. in several chronic conditions, a subset of patients account for most of the avoidable encounters. identifying these patients is the first step in targeted care delivery. objectives: to measure and analyze scd patient utilization patterns in the ec and inpatient at texas children's hospital (tch). we identified all individuals under years old with any encounter at tch associated with an international classification of disease (icd)- or code for scd, including hgb ss, hgb sc, and hgb s/beta thalassemia. for each patient, we identified all inpatient and ec encounters in the days prior to their most recent encounter. finally, each encounter was classified as associated with pain, acute chest syndrome (acs), or "other" using an algorithm of discharge diagnosis codes and pharmaceutical delivery. the total number of scd-associated ec and inpatient encounters over the prior year was calculated for each patient. we stratified each patient according to their utilization patterns: low ( - encounters), intermediate ( - encounters), and high (≥ encounters). we identified unique patients with scd that had at least one encounter from july until june . there were , scd-related encounters in the days prior to their most recent encounter. most ( %, n = ) patients exhibited low-utilization patterns and % (n = ) were intermediate. finally, a small subset ( %, n = ) demonstrated high-utilization patterns and accounted for % of all encounters. high-utilization was associated with older age and public payment mechanisms. pain encounters were predominantly in pre-adolescents and teenagers with high-and intermediate-utilization patterns. acs was most frequent in pre-teens and younger teens in the intermediate-utilization group. finally, the youngest-aged high and intermediate users presented for other reasons such as febrile episodes and splenic sequestration. our findings reflect national trends in that a significant portion of encounters are attributed to a small subset of patients exhibiting a high-or "super-" utilization pattern. at our institution, scd super-utilization is associated with older age and pain. we also identified a group of infants and toddlers with frequent encounters for fever. to comprehensively address this burden, it will be important to design interventions targeted toward age and specific medical needs. background: background: the rarity of diamond blackfan anemia (dba) has hindered describing the spectrum of disease, identifying predictive correlations, and guiding datadriven recommendations. long-term toxicities from steroid or transfusion therapy that start in childhood remain the major clinical problems in patients with dba who do not receive stem cell transplant. objectives: objective: to define the dba patient population at st. jude children's research hospital including treatment responses and toxicities to help inform recommendations on treatment and monitoring. design/method: method: medical records were reviewed for all patients with dba treated at st. jude between and for diagnostic testing, treatment types and regimens, and outcomes. two-sample t-test or wilcoxon rank sum test was used to compare continuous variables in two groups depending on the normality of the data tested by shapiro-wilk test. results: a total of patients with dba were identified with a median age of . years (range months - years) at last follow up. a ribosomal protein gene mutation was identified in / patients ( %) with an rps mutation / ( %). thirteen different congenital malformations were described in / patients ( %). fourteen of twenty ( %) patients treated with corticosteroids had an initial response and of those achieved full remission. three patients became steroid-refractory and were unable to wean to an acceptable dose. five of twenty patients continue on lower-dose steroids. five patients currently require no therapy. univariate analysis revealed no statistically significant genetic predictors of response or remission, however, / rpl patients responded to steroids with / ( %) in long-term remission. ten patients are maintained on chronic transfusions and have undergone successful hematopoietic stem cell transplant. nineteen of treated patients ( %) had a treatment-related toxicity. patients on steroids were more likely to have short stature than patients on transfusions or in remission (p = . ). severe bone mineral density deficit occurred in / ( %) patients, in before age years. eight patients had hepatic iron overload, in one documented by age years. other severe toxicities included restrictive cardiomyopathy from iron overload, pathologic fracture, diabetes mellitus, and premature ovarian failure in one patient each. this genotypically and phenotypically heterogeneous dba cohort had a high rate of treatment-related toxicities, notably growth retardation, bone density loss, and hepatic iron overload even in very young children. these findings underscore the need for early standardized monitoring. background: patients with sickle cell disease (scd) face worsening morbidity and mortality between ages and , when they must transition from pediatric to adult healthcare.( ) an effective curriculum addressing disease knowledge, educational and vocational skills, self-efficacy, and social supports is critical to a successful transition. traditional didactic approaches have not led to durable knowledge retention. ( ) technology-based methods have been attempted, but the best educational approach remains unknown. objectives: . to understand how adolescent and young adult (aya) patients with scd view existing transition education. . to include patient preferences in improving our transition curriculum. we developed a qualitative survey to assess patient views of existing approaches for learning about scd and their opinions about preferred transition topics. thirty patients with scd aged to years old were recruited between january and december . responses were managed using redcap electronic data tools hosted at the university of rochester.( , ) qualitative and quantitative data analyses were performed, including independent t-testing to compare responses between age groups. results: approximately % of subjects were under years of age, while % were or older. seventy-one percent had a computer, and . % had a cell phone, with most reporting daily use. subjects reported greatest satisfaction with learning from their doctor during clinic visits ( . % agree or strongly agree) and websites on a cell phone ( . % agree or strongly agree); the least popular methods were online chat rooms and microsoft® powerpoint presentations. satisfaction was similar across age groups. recommended transition topics were viewed positively, with subjects ranking highest understanding their bloodwork ( . % agree or strongly agree) and understanding laws protecting students with chronic disease ( . % agree or strongly agree). older subjects ( - years old) agreed more strongly with learning about opioid addiction and understanding differences between adult and pediatric doctors than did younger subjects ( - years old) (p < . ). this pilot study was successful in helping us to understand the educational needs of aya patients with scd. preliminary data underscore the importance of education provided by the pediatric hematologist. our results also suggest that the optimal use of technology-based methods requires further investigation and that tailoring transition education by age group may be useful. background: similar to patients with transfusion-dependent beta-thalassemias (tdt-beta), survivors of hemoglobin barts hydrops fetalis (homozygous alpha- -thalassemia, tdtalpha) will require lifelong transfusions of erythrocytes. we have previously shown that a transfusion strategy that is based on the guidelines developed for tdt-beta (conventional transfusion) is suboptimal for these patients owing to the differences in the pathophysiology of anemia in the two conditions: in tdt-alpha, conventional transfusion strategy will lead to a gradual increase in non-functional hbh with subsequent tissue hypoxia and hemolysis. an aggressive transfusion strategy that was based on reduction of hbh and increase in "functional" hemoglobin level resulted in improvement of tissue oxygenation and reduction of hemolysis but was associated with significant increase in transfusional iron burden [amid et al, blood ] . objectives: to define the optimal chronic blood transfusion targets for hbh% and functional hemoglobin in patients with tdt-alpha. design/method: following research ethics board approval, longitudinal data of patients with tdt-alpha ( males, median age . ( . - . ) were retrospectively collected. variables of interest included total pre-transfusion hemoglobin, hbh%, and "functional" hemoglobin [measured as total hemoglobin x ( -hbh/ )]. outcome variables were lactate dehydrogenase (ldh, marker of hemolysis), and soluble transferrin receptor (str, marker of erythropoiesis). hemoglobin analysis was done using high-performance liquid chromatography and capillary zone electrophoresis. we examined the association of "functional" hemoglobin with str, and hbh% with ldh, using repeated-measures anova to adjust for the effect of multiple testing. we constructed receiver operating characteristic curve and calculated the area under the curve to define the best cut-off values for variables of interests. there was a strong association between functional hb and str, as well as hbh and ldh. the optimal cut-off for "functional" hemoglobin that was associated with str < . mg/l was g/l (auc = . , sensitivity and specificity of . % and % respectively). the optimal cut-off for hbh to supress ldh to < u/l was % (auc = . , sensitivity and specificity of . % and % respectively). the optimal pre-transfusion hbh% for reduction of hemolysis was % and the optimal "functional" hemoglobin to adequately supress erythropoiesis was g/l. to meet these hbh% and functional hb targets by simple blood transfusions, patients with tdt-alpha would require a hypertransfusion regimen with a minimum pre-transfusion total hb of g/l and consequently high transfusional iron burden. an alternative approach using exchange transfusion to reduce hbh% and improve functional hemoglobin would be associated with less volume of transfusion and potentially better long-term outcome. hospital sacre coeur, milot, haiti background: initial results of work developing a pediatric sickle cell disease (scd) clinic at the hôpital sacré coeur (hsc) in milot, northern haiti were presented at aspho . the purpose of this clinic is for a pediatrician with a special interest in scd to provide scd care, advising on trait and managing disease with penicillin prophylaxis (pcn) and hydroxyurea therapy (hu) for select patients. this clinic was started in collaboration with a us based hematologist and support from yale-new haven hospital. objectives: to describe the success and challenges of providing pcn and hu in the scd clinic at hsc through a review of patient records. design/method: since this clinic's inception, a database of patients, with basic clinical information has been kept and made accessible, through 'drop-box', to the us hematologist. the records of those that presented to the clinic were reviewed. the hemoglobin diagnosis was made either by clinical history and sickle cell prep or by hemoglobin electrophoresis through alpha laboratory, port-au-prince, haiti. results: ninety-nine individuals were seen in the first years of the program. fifty-six underwent a hemoglobin electrophoresis. of these , are ≤ years old. thirty-two were started on pcn vk, of which / ( %) were ≤ years old. eleven patients were started hu therapy. all patients on hu have shown progressive increases in hemoglobin. there have been no clinical complications of hu therapy. none of the patients taking hu have required hospitalization or transfusion in . three patients (not on hu) were hospitalized in for complications of scd (osteomyelitis, pain). in , with less than half the numbers in the program, there were admissions for severe anemia, pain, stroke and splenic sequestration. with ongoing external support and a local reputation for excellence in sickle cell care, the clinic at hsc has been able to expand services and improve the health of a growing number of patients with scd. early data suggests that pcn and hu therapies are helping to reduce complications and improve quality of life. challenges to date have included lack of funding for transportation to clinics, for hospitalizations and to cover the cost of electrophoreses. at the same time as continuing providing excellent care and gathering data, it is crucial to explore opportunities for collaboration and cooperation in ways that will assure that the clinic can become independently sustainable while continuing to improve the quality of life for the individuals it serves. background: ykl- is an inflammatory glycoprotein expressed by infiltrating macrophages in various inflammatory conditions. it has been found to be elevated in patients with different pathological conditions like acute and chronic inflammations, increased remodeling of the extracellular matrix (ecm), development of fibrosis and cancer. several studies have found elevated ykl- concentrations in sera of patients with liver diseases such as hepatic fibrosis by hepatitis c virus. it has been suggested that ykl- concentrations reflect the degree of liver fibrosis. to evaluate serum ykl- levels in patients with -thalassemia and its relation to viral hepatitis, liver stiffness as assessed by transient elastography (fibroscan, fs) and hepatic iron concentration. design/method: a prospective study included patients with -tm ( males and females) with mean age . ± . years (range: - years). serum ferritin level, liver enzymes (alt and ast), hbs ag, anti hcv ab and serum ykl- using elisa kit were evaluated. all patients were subjected to liver mri t * to detect liver iron content by the sequence and transient elastography (fibroscan, fs) to assess degree of liver stiffness. results: mean fibroscan value was ( . ± . ) kpa with a median . (range . to ) kpa. ( %) patients were categorized as f - and ( %) were stage f - , ( %) patients had severe fibrosis. their median serum ferritin was ng∖ml, with ( %) patients had values exceeding g/l. median cardiac t * was . with patients had values below ms, and the median lic was . mg/g dw with patients showed readings above mg/g dw. nyl- was evaluated as a marker of inflammation and liver fibrosis and showed mean value . (± . ) pg/ml, and range from to pg/ml. mean ykl- was significantly higher among males (p = . ), patients on chelation therapy (p = . ), patients on dfs (p≤ . ), in those with abnormal liver enzymes, splenectomised patients, patients with hbv sero-positivity, those with moderate elevation of t * and patients with high grades of liver fibrosis (p< . ). ykl- showed positive correlation with the rate of transfusion, lic, ferritin, alt and ast but negative correlation with weight, height and t *. roc curve analysis revealed that the cutoff value of ykl- at pg/ml could differentiate -tm patients with and without viral hepatitis with . % sensitivity and specificity of . %, area under the curve (auc) . , positive predictive value . and negative predictive value . (p< . ). roc curve analysis revealed that the cutoff value of ykl- at pg/ml could detect -tm patients with liver cirrhosis with . % sensitivity and specificity of . %, area under the curve (auc) . , positive predictive value . and negative predictive value . (p< . ). conclusion: serum ykl- levels are elevated in patients with -thalassemia and can detect patients with active viral hepatitis and liver stiffness. background: the most common splenic complication in pediatric patients with sickle cell disease (scd) is acute splenic sequestration (ass), which has often been managed with splenectomy. although splenectomy has been a treatment of choice for years, long-term vascular complications have not been thoroughly evaluated. pulmonary hypertension (phtn) is a severe complication of scd. in adults with scd, phtn has been associated with a -month mortality rate of approximately %. it has been reported that splenectomized patients with hemolytic disorders are at even greater risk of phtn. several medications exist to treat phtn, but with few studies of their efficacy or toxicities in patients with scd. additionally, these patients are often treated with either chronic prbc transfusions or hydroxyurea (hu) to raise hemoglobin, reduce hemolysis, and prevent vaso-occlusive events. objectives: to evaluate effect of chronic prbc or hu vs. no intervention, on tricuspid regurgitant jet velocities (trv) in pediatric patients with scd and history of splenectomy. design/method: retrospective chart review of splenectomized patients with hbss followed at marian anderson center at st. christopher's hospital for children, philadelphia, between and . we analyzed trvs ( hu, prbc, and from control group receiving neither treatment) from patients ( hu, prbc, neither). mean age at echo was . +/- . . data was analyzed with linear correlations and analysis of variance (anova), including the post hoc test of least significant difference (lsd) for all pairs of treatment groups. results: trv was not significantly correlated with age at time of assessment or with time between splenectomy and trv. univariate anova among groups yielded trv means of: . +/- . cm/s (hu), . +/- . (prbc), . +/- . (neither). we found a notable difference as the mean of the hu group was almost cm/s lower than the others, but no overall statistically significant association for any of the groups exists. however, when we performed post hoc tests to adjust for multiple comparisons and looked at all pairings within the anova, we found that the lsd between the hu and the prbc groups was statistically significant (p = . ), and that a trend exists between the hu group and the neither treatment group (p = . ). our data suggests that treatment with hu is correlated with a reduction in trv in pediatric patients with scd who underwent splenectomy. given these promising results, we believe our data warrants further study with larger treatment groups. nancy olivieri, gaurav sharma, susmita nath, rajib de, tuphan kanti dolai, prakas kumar mandal, abhijit phukan, amir sabouhanian, robert yamashita, angela allen, david weatherall, prantar chakrabarti background: hemoglobin e thalassemia (hbethal), which accounts for % of all severe beta thalassemia worldwide, has an estimated prevalence of . / , in west bengal, from which little information about clinical findings has been reported. objectives: to document clinical and laboratory findings in patients with hbethal, ultimately to improve resources for clinical management. design/method: we reviewed records from: a database recording patient names; clinic charts; "special" charts containing additional details; and, in transfused patients, transfusion day-care records. additionally, because in india's public hospitals original lab/imaging reports are commonly retained at home, % of families were interviewed to provide additional information. we excluded records of patients aged < years and patients aged < years who had not been reviewed since . results: while at least one visit had been recorded in , hbethal patients at nrs hospital, most patients are not regularly reviewed there. we examined charts [ ( %) aged ≥ years; ( %) aged - years; % male], representing approximately % of regularly-reviewed patients. most families ( . %) reported monthly incomes (< , indian rupees), below the monthly cost of living ( , rupees) in kolkata. mean (±sem) hemoglobin was . ± . g/dl. % patients were receiving eight or more transfusions per year; from , % had been treated with deferasirox, . ± . mg/kg/day. iron control estimated by serum ferritin concentration ( . ± g/l) was highly variable. a total of % patients were splenectomized. a substantial obstacle to documenting complications was the lack of recording, in any of the five sources, of many relevant parameters: for example, the status of sexual maturation (normal, delayed, or absent) was documented in less than %, and measurements of fasting blood glucose in less than %, of records. where recorded, complication rates were high: delayed/abnormal sexual maturation was recorded in % patients aged > years; in the patients aged > years and those aged - years, respectively, hypothyroidism was recorded in % and %, and elevated serum alt in % and %. in most evaluable patients > years, height was measured between the rd- th percentiles. cardiac findings, rarely documented, included pulmonary hypertension and reduced left ventricular ejection fractions in a few patients. despite dedicated attention to many aspects of thalassemia care, insufficient documentation limited a clear understanding of the current morbidity in hbethal patients. investment in personnel and technology will be critical to record relevant information, ultimately to improve clinical management, over the next decade. children's hospital of richmond at vcu health, richmond, virginia, united states background: sepsis is a common cause of death in children with sickle cell disease (scd). recommendations for care of fever in children with scd include immediate medical evaluation including blood culture and initiation of broad-spectrum antibiotic therapy. the increasing availability of pcr-based respiratory pathogen panels (rpp) provide the opportunity to rapidly identify viral causes of fever. the role for rpps in identifying the source of fever in children with scd and how it affects provider practice is not well studied. ( ) to determine the epidemiology of respiratory virus-associated fever in children with scd and ( ) to determine whether a positive rpp is associated with reduced risk of bacteremia in this population. this was a single-center, retrospective cohort study. we identified and reviewed the medical records of all children with scd seen in our emergency department (ed) with temperature ≥ . oc at home or in the ed from january , , through september , , as well as, all febrile children for whom rpps were sent since the introduction of rpps april . we reviewed the results of blood cultures, rpps, chest radiographs, and ed notes and discharge summaries to identify sources of infections. independent t test and chi-square analysis were used as appropriate to compare results using spss©. overall, the rate of bacteremia was %. there were no cases of bacteremia among children with positive rpps. % of children with negative rpps had true bacteremia. a positive rpp did not reduce the likelihood of bacteremia (p . ). patients with bacteremia had higher presenting temperatures than those without bacteremia ( . oc vs . oc, p . ). the most common rpp findings were rhinovirus/enterovirus ( %), human metapneumovirus ( %), and influenza a ( %). sending an rpp did not affect admission rate ( % and % respectively, p . ); however, likelihood of admission was lower in patients with positive rpps ( % vs %, or . [ . - . ], p . ). length of stay (los) was shorter in patients for whom an rpp was not sent ( . vs . days, p . ). as previously reported, bacteremia in febrile children with scd is very low, but remains a serious concern, particularly in the setting of high fever (> oc). a positive rpp did not reduce the odds of bacteremia, but did have a sta-tistically significant impact on both admission rate and los. more work is needed to understand how rpp results impact provider decision-making and care for children with scd. cincinnati children's hospital medical center, cincinnati, ohio, united states background: diffuse myocardial fibrosis is a common, if not defining, feature of the heart in sickle cell anemia (sca) that is strongly associated with diastolic dysfunction. we found diffuse myocardial fibrosis in every patient in a sca cohort (n = ) ranging in age from to years (niss ). the treatment and prevention of this complication of sca has not been studied before. objectives: because diffuse myocardial fibrosis must begin in early childhood, we hypothesized that early initiation and uninterrupted use of disease-modifying therapy for sca can prevent it. design/method: we use cardiac magnetic resonance imaging (cmr) to measure the myocardial extracellular volume fraction (ecv) to quantify diffuse myocardial fibrosis in individuals with sca who have been treated, uninterrupted, with hydroxyurea or chronic transfusion therapy since ≤ years of age. two comparison groups were used: individuals with sca who have not been treated with disease-modifying therapy since ≤ years of age (n = ) and controls without sca (n = ). results: we studied individuals ( m/ f) with a mean age of . years (range - ). mean age at the start of diseasemodifying therapy was . ± . years (range - ). only had evidence of mild diffuse myocardial fibrosis (ecv . ); the other had no detectable diffuse fibrosis (all had ecv < . , the upper limit of normal). mean ecv was . ± . , which was significantly lower than the ecv of individuals with sca who have not received early uninterrupted therapy ( . ± . ; p = . ) and not statistically different from normal controls ( . ± . ; p = . ). none had macroscopic fibrosis by late gadolinium enhancement or evidence of myocardial hemosiderosis by t * imaging. no patient had diastolic dysfunction by echocardiographic classification, right heart catheterization, or both. disease-modifying therapy for sca can prevent diffuse myocardial fibrosis, and possibly diastolic dysfunction, if started in early childhood. prospective trials of disease-modifying and anti-fibrotic therapy are planned to prevent diffuse myocardial fibrosis, which can be monitored noninvasively by cmr, and improve outcomes in sca. (niss, blood, ) . background: a statewide sickle cell surveillance system (sscss) was developed with the goal of determining the prevalence of sickle cell disease (scd) in indiana and the level of care that patients receive throughout the state. persons with scd are at high risk of infection, especially with encapsulated organisms, as well as at increased complications from influenza. utilizing sscss data, the relationship between vaccination status and mortality was explored. to determine if vaccination status is associated with mortality in persons with scd. the project was granted a waiver of consent by the st. vincent irb. death certificates were obtained to identify cause of death. deceased patients (cases) were matched by age, gender, and sickle genotype to living patients (controls). vaccination data were collected from the medical record and the children and hoosier immunization registry program (chirp) through the date of death for each case. cases and controls were assigned a point for completion of the pneumococcus, meningococcus and haemophilus influenza type b (hib) vaccine series and one point if the influenza vaccine was given within a year prior to death of the cases [max vaccine status score (vss): ]. total points were compared between the cases and controls. two tailed t-tests to compare means of continuous data and wilcoxon signed-rank test to compare ordinal data. one thousand forty-eight individuals were included in the sscss. six hundred and seven ( . %) were seen at one institution and included in this analysis (mean age = years). thirty-three of the ( . %) were deceased at the time of analysis. six point one ( . )% of controls and . % of cases received a vss of . the mean vss for cases was . ± . and . ± . for controls. thirty point three ( . ) % of controls had a vss of one or more, compared to % of cases (p = . ). patients who died of infection [streptococ-cus (n = ), pseudomonas (n = ) and unidentified organisms (n = )] were not up to date on vaccination against encapsulated organisms, but two had received the influenza vaccine in the year prior to death. in this sample, mortality occurred exclusively among adult patients, which is consistent with current patterns in developed countries. among these adults, vss and mortality rates were not related. limitations to the study include small sample size and potential incompleteness of vaccine records. vaccination rates and other standard of care indicators should be explored in a larger cohort of patients to determine associations with mortality. background: sickle cell disease (scd) is a genetic disorder resulting in acute and chronic complications, including delayed puberty. delayed puberty can have adverse physical and psychosocial effects on affected children and families. there are no published reports from ghana on pubertal timing in children with scd. the aim of this cross-sectional study was to describe pubertal changes in children with scd at korle bu teaching hospital (kbth), accra, and compare these findings to those in a control group without scd. design/method: children with scd and children with hb aa, ages - years, were consecutively recruited and matched for age, sex and socioeconomic status. investigator-administered questionnaires were used to obtain demographic data for all participants and information on menarche (girls only). pubertal status was assessed by physical examination using tanner staging. testicular volumes were determined in boys using a prader orchidometer. body mass index (bmi) and socioeconomic status (ses) of participants were analyzed to determine if there were any associations with tanner stage. of the with scd, ( . %) were hb ss and ( . %) hb sc. females comprised . % (cases and controls). mean age at onset of breast development was significantly delayed in girls with scd ( . ± . years) compared to controls ( . ± . years) but there was no significant age difference at onset of pubic hair development. mean age at menarche was significantly delayed in girls with hb ss ( . ± . years) and hb sc ( . ± . years), compared to those with hb aa ( . ± . years). in boys, the mean ages at onset of puberty were significantly delayed in those with scd ( . ± . years, for genital development and . ± . years, for pubic hair development), compared to those without scd ( . ± . years and . ± . years, respectively). mean testicular volumes were significantly lower in cases compared to controls, across all age ranges (p< . ). mean bmi in both cases and controls were similar at onset of breast development in girls. however, in boys with and without scd, mean bmi values were significantly different at pubertal onset. in univariate analysis, ses was not associated with tanner stage for both genital and breast development. mean ages at pubertal onset were significantly delayed in children with scd. longitudinal studies are needed to further characterize any associations with bmi and determine potentially modifiable risk factors affecting pubertal onset in scd. background: sickle-cell disease (scd) is a life-threatening genetic disorder associated with multiple chronic and acute complications. specific monitoring and treatment for children is a major part of the medical focus, but there remains a lack of real-world evidence of the disease burden and practice patterns among the pediatric scd population. objectives: to examine the clinical burden and management of scd among pediatric patients. design/method: a retrospective claims study was conducted using the medicaid analytic extracts database from jan - dec . pediatric patients (aged < years) with scd were identified using icd- -cm diagnosis codes ( . - . , . - . ). the first observed scd diagnosis during the identification period was designated as the index date. patients were required to have continuous medical and pharmacy benefits for at least months pre-and months post-index period. patient data were assessed until the earliest occurrence of the following events: disenrollment, death, or the end of the study period. patient demographic and baseline clinical characteristics, clinical outcomes (mortality, incidence of pain crisis, complications), scd management, and healthcare utilization were examined. all variables were analyzed descriptively. results: a total of , patients met the study inclusion criteria, with a mean age of . years. most patients were black ( . %) and had a charlson comorbidity index score of ( . %). mortality during follow-up was . in personyears, and the event rate of pain crisis in the inpatient setting was . in person-years. the three most common complications after pain crisis (highest rates in person-years) were fever ( . ), infectious and parasitic diseases ( . ), and asthma ( . ). rates of life-threatening complications were also examined in person-years, including acute chest syndrome ( . ), stroke ( . ), splenic sequestration ( . ), pulmonary hypertension ( . ), and pulmonary embolism ( . ). . % of patients were prescribed antibiotics during the one-year post-index period. other frequent medications utilized among children were folic acid ( . %), nonsteroidal anti-inflammatory drugs ( . %), opioids ( . %), and hydroxyurea ( . %). . % of patients had a blood transfusion within one year post-index date. patients had frequent health care utilizations in the inpatient ( visit), emergency room ( visits), office ( visits), and pharmacy ( visits) settings during the one-year follow-up period. pediatric scd patients are burdened with a high rate of complications including pain crisis. in addition, patients utilized a substantial amount of health care resources including outpatient office care and acute care visits. background: novel use of hydroxyurea in an african region with malaria (noharm, nct ) is a randomized controlled trial of hydroxyurea for very young children with sickle cell anemia living in uganda. during year , study participants received blinded study treatment of hydroxyurea or placebo; those receiving hydroxyurea had no increased risk of malaria, but had both laboratory and clinical benefits. during year , all study participants received openlabel hydroxyurea treatment. to assess the effects of open-label hydroxyurea treatment in a very young population of children with sickle s of s cell anemia living in uganda. study endpoints included the rates and severity of malaria infections, clinical sickle-related events, and laboratory effects. design/method: all children in the noharm trial were enrolled at mulago hospital sickle cell clinic in kampala uganda. during year , all children received open-label fixeddose hydroxyurea ( mg/kg/day) for months, after previously receiving either hydroxyurea or placebo for months. results: a total of children entered year of the noharm trial and received fixed-dose hydroxyurea, including males and females, at an average age of . ± . years. among children previously on placebo, there were malaria events in children, including with severity grade ≥ , and three deaths (two acute chest syndrome, one sepsis). clinical adverse event rates dropped from . to . per patient year, and hospitalizations were reduced from to . expected hematological benefits of increased hemoglobin, mcv, and fetal hemoglobin, along with decreased neutrophils and reticulocytes, were rapidly achieved. laboratory adverse events were infrequent at . events per patient-year, and only half of those were dose-limiting hematological toxicities. among children previously on hydroxyurea, there were malaria events in children, including with severity grade ≥ , and two deaths (one acute chest syndrome, one sepsis). clinical adverse event rates and hospitalizations were maintained at low rates, the hematological benefits of hydroxyurea continued throughout the extended treatment period, and dose-limiting toxicities remained infrequent. fixed-dose hydroxyurea treatment of young children with sickle cell anemia living in uganda is associated with no increased risk for malaria. clinical and laboratory benefits occur, including children previously on placebo who crossed-over to hydroxyurea treatment. future studies should focus on the optimal dosing and monitoring strategies, in an effort to determine the overall feasibility and safety of introducing hydroxyurea therapy across sub-saharan africa. background: acute chest syndrome (acs) is the second most common cause of hospitalization in patients with sickle cell disease and is a leading cause of morbidity and mortality. in mid- , an algorithm was implemented at cohen children's medical center to initiate transfusions within four hours of diagnosis of acs in order to improve patient outcomes. objectives: the aim of this project was to analyze the effect of early blood transfusion on the outcomes of patients with acs. we focused on the number of total transfusions, need for exchange transfusion, need for intensive care unit (icu) stay, and length of hospitalization. design/method: a retrospective chart review was completed on patients admitted to ccmc with a primary diagnosis of sickle cell disease and a secondary diagnosis of either acs or pneumonia during the years of - . data from the three years directly prior to implementation of the algorithm was compared to data from the three years directly after implementation of the algorithm. a total of patients were analyzed, of which belonged to the pre-algorithm group and to the postalgorithm group. patients from the post-algorithm group had a higher incidence of transfusions ( % with a mean transfusion number of . pre versus % with a mean of . post) as well as exchange transfusion ( % pre versus % post). the post-algorithm group had a shorter overall length of stay (mean of . days pre versus . days post). while the overall percentage of patients requiring an icu admission was similar in each group ( % pre versus % post), the post-protocol group had a lower likelihood of requiring an icu admission for reasons outside of line placement for exchange transfusion, most commonly for icu-level respiratory support ( % pre versus % post). despite a higher total number of transfusions, early recognition and transfusion for acs can lead to decreased lengths of hospitalization as well as decreased need for icu-level respiratory support. further studies comparing different center's clinical practice guidelines are necessary to improve the standard of care. background: novel use of hydroxyurea in an african region with malaria (noharm) was the first placebocontrolled randomized clinical trial of hydroxyurea in sub-saharan africa. in noharm, young children with sca received either hydroxyurea or placebo during year , followed by open-label hydroxyurea for all study participants during year . an ancillary noharm project was designed to determine if hydroxyurea treatment lowers transcranial doppler (tcd) velocities and possibly reduces stroke risk in this very young cohort. objectives: to perform tcd screening on the noharm cohort, measuring the time-averaged mean velocity (tamv) at the end of both year and year . we hypothesized that the maximum tamv would be lower for noharm study participants receiving hydroxyurea compared to those receiving placebo, and that key clinical and laboratory parameters would also influence tcd velocities. design/method: all children enrolled in noharm were eligible to undergo tcd examination at two study time points: month - when they were completing the blinded treatment phase, and again at month - at the end of the open-label treatment phase. tcd measurements included tamv readings from the main intracranial arteries: middle cerebral artery, distal internal carotid artery, and bifurcation on tcd. all tcd examinations were scored and classified as normal (less than cm/sec), conditional ( - cm/sec) or abnormal (greater than or equal to cm/sec), with higher scores correlating to greater risk of stroke. results: at the end of year , tcd exams were conducted of which were suitable for analysis ( hydroxyurea, placebo). based on the maximum tamv, the median velocity was cm/sec (iqr - ) for children on hydroxyurea and cm/sec (iqr - ) on placebo, p = . . maximum tamv values had negative correlations with hemoglobin concentration (- . ), fetal hemoglobin (- . ), and oxygen saturation (- . ); positive correlations were noted with age ( . ) and absolute neutrophil count ( . ). at the end of year , tcd exams were conducted and all were suitable for analysis; the median velocity was cm/sec on open-label hydroxyurea treatment, regardless of previous blinded treatment. all correlations with tamv were maintained except for age. conclusion: compared to placebo, hydroxyurea treatment for young children with sca living in uganda was associated with lower tcd velocities, which have been correlated in other studies with lower risk of primary stroke. tcd velocities were correlated with hematological and clinical parameters that can be improved by hydroxyurea therapy. children's hospital of richmond at virginia commonwealth university, richmond, virginia, united states background: acute chest syndrome (acs), defined by respiratory symptoms and a new pulmonary infiltrate, is a serious complication of sickle cell disease (scd). acs can occur during hospitalization for non-pulmonary conditions, such as a vaso-occlusive crisis or after surgery. nih clinical practice guidelines encourage incentive spirometry (is) which decreases the incidence of acs. it is additionally widely accepted that early, frequent ambulation in post-operative and pneumonia patients decreases the length of stay (los). to decrease acs events in children with scd at our children's hospital, we aimed for is use in % of ageappropriate pediatric sickle cell admissions. design/method: a multidisciplinary team examined inpatient acs prevention practices, including is, at children's hospital of richmond. key drivers were identified, including educational awareness of patients and healthcare staff, order placement, and documentation. we aimed for all scd patients ≥ months of age hospitalized with any admission diagnosis to participate in is with the use of a traditional incentive spirometer or similar age-and ability-appropriate devices (e.g. positive expiratory pressure devices, bubbles, and pinwheels). we secondarily aimed to increase activity events, specifically ambulation and out of bed time. educational and outreach tools included patient informational brochure and incentive program, and staff informational sessions and reference materials at workstations. a disease-specific order set was implemented including desired is and activity orders. data were collected prospectively may through november , during which pdsa cycles were conducted. admissions during the corresponding months of the previous year were reviewed for comparison. independent t-test analysis was performed using graftpad prism statistical analysis software. results: improvements reaching statistical significance included increase in is order placement from % to % of admissions (p < . ), and admissions with documented is use increased from % to % (p < . ). los decreased from a mean of . days to . days (p . ). post-admission development of acs also decreased from % to % of admissions, but did not reach statistical significance (p . ). there was an additional increase in appropriate activity order placement and documentation of activity events. conclusion: improving education and outreach to patients and staff, including implementation of a disease-specific order set, can improve is use and activity events. the decline seen in incidence of acs development during hospitalization, though not statistically significant, and the decreased los are encouraging, and efforts continue to improve on these trends. background: painful vaso-occlusive crises (voc) are a frequent and debilitating complication of sickle cell disease (scd) and are thought to occur due to progressive blockage of the microvasculature with rigid sickle shaped red blood cells. any trigger that decreases the microvascular blood flow (mbf) can promote entrapment of sickled cells in the microvasculature and progression to voc. exposure to cold wind and changes in weather are common triggers of voc and are associated with increased frequency of hospitalizations for pain in patients with scd. there is limited experimental data on the physiologic effects of these factors on peripheral perfusion in scd. to study the effect of graded thermal stimuli on the peripheral mbf in scd. design/method: scd and control (healthy or sickle trait) subjects aging to years were exposed to their individual threshold temperatures for heat and cold detection, heat and cold pain via tsa-ii thermode that was placed on the thenar eminence. mbf was measured on the contralateral thumb using photo-plethysmography (ppg). the vasoconstriction response within the complex ppg signal was detected using cross-correlation technique. mean mbf was derived from the ppg amplitude during each of these stimuli and compared to baseline mbf. cross correlation analysis showed that cold pain caused significant vasoconstriction response in % of the subjects, followed by heat pain ( %), cold detection ( %) and heat detection ( %).there was a significant drop in the mbf during cold pain (p < . ), heat pain (p < . ), heat detection (p = . ) and cold detection (p = . ) when compared to baseline mbf, with cold pain causing the greatest drop in mbf. thermal sensitivity and mbf responses were comparable between scd and controls. conclusion: exposure to graded thermal stimuli causes a progressive drop in mbf with exposure to cold pain eliciting the strongest vasoconstriction response. vasoconstriction occurred in the contralateral hand at an average of seconds after the stimuli, suggesting a neurally mediated mechanism. although there was no significant difference in vasoconstriction responses between scd and controls, the drop in mbf in patients with sickle cell disease can increase the likelihood of entrapment of the sickled red blood cells, leading to vaso-occlusion. these findings are consistent with extensive reports in literature that exposure to cold weather is associated with a higher frequency of voc. this suggests that neurally mediated vasoconstriction is likely an important factor in the pathophysiology behind cold exposure leading to voc in scd. background: vaso-occlusive crisis (voc) is a major cause of hospital admissions in children with sickle cell disease (scd). although the use of clinical biomarkers in voc has been studied, especially with regards to acute chest syndrome (acs), there is less data regarding overall voc severity prediction. in addition new biomarkers such as platelet to lymphocyte ratio (plr), neutrophil to lymphocyte ratio (nlr), and lymphocyte to monocyte ratio (lmr) have been little studied with regards to scd. objectives: to identify whether admission laboratory values, changes from well baseline laboratory values, and new biomarkers such as plr, nlr, and lmr could predict severity of vaso-occlusive crisis in children with sickle cell disease admitted with voc. design/method: this was a retrospective single center observational study of admissions of voc in children aged - years with hbss or hbs-b thal from september to november excluding those on hyper-transfusion protocol or having an admission diagnosis of acs. univariate analysis was done using student's t-test, mann-whitney non parametric test, or fischer's exact test as appropriate depending on the distribution between admission laboratory data of complete blood count (cbc), reticulocyte count, comprehensive metabolic panel, lactate dehydrogenase (ldh), change from well baseline cbc values within months previously, plr, nlr, lmr, and the development of complicated voc. complicated voc was defined as the development of secondary acute chest syndrome, prolonged admission duration > days ( hours), requirement of blood transfusion, and readmission within days. results: a total of admissions were studied. fifty-nine ( . %) were female. of the , ( . %) were complicated with no significant differences in sex (p . ) or age (p . ). univariate analysis revealed significant elevations in total bilirubin (p . ), ldh (p . ), and platelet count (p . ) in those with complicated voc. there is also significant difference in the percentage change of platelet count from baseline with greater decline in uncomplicated voc (p . ). there were no significant differences in plr (p . ), nlr (p . ), or lmr (p . ). conclusion: elevations in total bilirubin, ldh, and platelet count in admission laboratory values are associated with developing complicated voc. in addition, those with complicated voc present with significantly less decline in platelet count from baseline well cbc. plr, nlr, and lmr do not seem to be useful predictive biomarkers for severity of voc. background: sickle cell disease (scd) causes health problems of varying frequency and severity. the only validated biomarker for children with scd is transcranial doppler. if reliable predictors existed for scd severity, children with scd could be treated according to risk category. many patients with scd face psychosocial or economic hardships, but these factors have not been evaluated as risk markers for medical or functional severity of scd. objectives: the goal of this project was to develop and stratify a preliminary list of psychosocial risk factors for health outcomes that could be used as scd severity predictors. st. vincent institutional review board. a list of potential psychosocial risk factors for adverse health outcomes was compiled based on assessment materials utilized by the sickle safe program (indiana's hemoglobinopathy newborn screening follow-up program). this list of items was distributed to child abuse prevention ( ) and scd ( ) experts, who ranked each item on a likert scale of (least important) to (most important). mean scores were calculated using spss version ; assessments were retrospectively analyzed to determine psychosocial risk factor frequency. risk factors occurring in ≥ % of homes were considered high frequency events. overall, there was high agreement among experts on the risk factors that were considered the most important predictors of severe scd outcomes. the risk factor with the highest frequency ( %) was eligibility for public assistance programs. fifteen risk factors were rated ≥ by the experts. four ( . %) were high frequency events occurring in ≥ % of homes: a child with hbss or hbs thalassemia not taking hydroxyurea ( %); parent report that they had treated a fever (> ®f) at home in the past months ( %); tobacco use by someone in the household ( %); and the family reporting significant psychosocial stressors in the past year ( %). tobacco use in the home was significantly correlated with several other risk factors (smoking during pregnancy [r = . ], other health concerns in the child [r = . ], and child having health insurance [r = - . ]), suggesting that it is part of a constellation of health risk. in general, the risk factors that were rated as most important for health outcomes occurred less frequently in the sample. this study represents important progress toward identifying a group of psychosocial risk factors for scd severity, which is a necessary first step for future investigation of empirical relationships between candidate risk factors and scd outcomes. unitversity of cartagena, cartagena, bolivar, colombia s of s background: sickle cell disease is an autosomal recessive disorder characterized by a mutation in the -globin chain, which produces hbs. acute and chronic complications as aplastic crisis, acute chest syndrome, priapism, stroke, leg ulcers and primary/secondary prevention of stroke can be treated with simple transfusion or exchange transfusion. the latter offers advantages as lower iron overload, post-treatment hbs goal control, lower viscosity and improved microvascular circulation. but it is not a widely-used option because is associated with technical difficulties. objectives: standardization of a new partial exchange transfusion protocol in a group of patients with sickle cell disease, within the framework of a chronic transfusion program. design/method: this is a prospective descriptive study, which included patients under years with sickle cell disease ( hbss, hbs-tal), with indication of partial exchange transfusion in a chronic transfusion program, according to the institutional protocol; patients who fulfilled the inclusion criteria were enrolled in the study between february and december . a registry of the medical and technical complications was made in each of the procedures. a database was constructed in excel, and the graph-pad prism® version oc software was used for statistical analysis. the sequence is as follows: isovolemic phlebotomy and transfusion of packed red cells. depending of the recent hemoglobin level ( hrs), we do the phlebotomy there: hb: - . : cc/kg, hb: - . : cc/kg, hb> : cc/kg; isovolemic solution (ns , %) there: hb: - . : cc/kg, hb: - . : cc/kg, hb> : cc/kg and packed red cell transfusion there: hb: - . : cc/kg, hb: - . : cc/kg, hb> : cc/kg. the safety of this exchange transfusion protocol was analyzed in patients with sickle cell disease ( procedures). there were no differences in the sex distribution, and the median age was years. % of the population was homozygous. the indication of transfusion was . %( / ) primary stroke prevention, . %( / ) secondary stroke prevention and . %( / ) was other reason. a low percentage of complications was found ( . %); of which, those of medical origin (hypotension and nausea/vomiting) were only presented in . % of the total procedures. the standardization of this protocol was safe and its use could be extended to other low-income centers that treat patients with sickle cell disease that need chronic transfusion program including patient with hemoglobin level until gr/dl. we suggest do studies for measure the security and efficacy of this protocol in patients with acute complications. background: clinical trials that aim to achieve pain reduction have challenges achieving clinical endpoints as pain has no quantifiable biomarkers and may be unrelated to scd. furthermore, the threshold of seeking medical care differs between patients and vocs that occur at home are missed. we present a non-interventional, longitudinal study to identify vocs in patients with scd. objectives: to examine the longitudinal relationship between pros and biomarkers in subjects with scd before, during, and after a self-reported voc event, in order to build a model of in-home and clinical voc and to collect longitudinal pros and biomarker data from subjects that span voc events in the home, clinic and the hospital. design/method: longitudinal measures of pain, fatigue, function, activity, and biomarkers from scd patients in steady state and voc were studied over a six month period. patients self-reported pain, fatigue, function, and medication use using a novel epro tool. voc was reported in real-time, triggering a mobile phlebotomy team. blood was collected sequentially after self-reported voc (at home or hospital). blood samples were drawn two days after resolution of voc, as reported by the patient. during non-voc periods, blood was drawn every weeks to establish a baseline. biomarkers included leukocyte-platelet aggregates and circulating microparticles, cell and soluble adhesion molecules, cytokines, inflammatory mediators and coagulation factors. patients wore an actigraphy device to track sleep and activity and rest. results: twenty-seven of thirty-five patients experienced a total of days with voc > hr, of which only days resulted in healthcare utilization. voc days had significantly higher pain and fatigue scores. voc days were associated with significantly decreased functional scores, with significantly greater decreases during vocs requiring medical contact compared to at-home vocs. different activity profiles were identified for non-voc, at-home voc and medical contact voc days by actigraphy monitoring. at-home voc days exhibited increased daytime resting compared to non-voc days. medical contact vocs had decreased average and peak activity, and increased daytime resting compared to non-voc days. a sleep fragmentation index trended up for both at-home ( %) and medical contact voc days ( %). significant changes during voc days were observed in: c-reactive protein ( % increase), nucleated rbc ( % increase), monocyte-platelet aggregates ( % increase) and neutrophil-platelet aggregates ( % increase), interleukin- ( % increase), interleukin- ( % increase) and tnfalpha ( % increase). the identification and assessment of at-home vocs through use of epros, actigraphy and biomarkers is feasible as demonstrated by this innovative at-home study design. background: risk-stratifying sickle cell disease (scd) patients and demonstrating response to disease-modifying therapies is challenging due to the phenotypical heterogeneity of scd. a pathogenic role for procoagulant von willebrand factor (vwf) via excess vwf high molecular weight multimers (hmwm) has been proposed, with variable reports of increased vwf and hmwm in crisis vs. steady-state in adults, but less so for vwf in children with scd. moreover, vwf and multimers have not been studied in sickle trait. objectives: our pilot study evaluated the potential for vwf antigen (vwf:ag) and hmwm on densitometric tracings to serve as biomarkers for disease severity or treatment response in children and young adults with scd compared to sickle trait (hbas) siblings. design/method: we evaluated vwf:ag, vwf multimers and retrospective clinical data from hbss, hbsc and hbas subjects at steady state. one hbsc subject also had a crisis sample. median scd age was years ( . - . years). % were female. scd severity was judged by annual vasoocclusive and acute chest events, or stroke/elevated tcd. eight of ( hbss and hbsc) took hydroxyurea. four hbss subjects had severe scd, all of whom were chronically transfused. results: mean vwf:ag (normal - iu/dl) was higher for hbss ( +/- . ) and severe hbss ( +/- . ) compared to hbsc ( +/- . , p = . and . , respectively); however, lacked statistical significance when compared to hbas ( +/- . , p = . and . , respectively). vwf:ag was elevated in / ( %) steady-state, including / ( %) with "severe" disease on chronic transfusion and / ( %) taking hydroxyurea, in hbsc crisis but no hbsc / ( %) at baseline. vwf:ag was high in / ( %) hbas siblings. four ( %) had increased hmwm at baseline: hbss/severe disease/chronic transfusion, hbss/hydroxyurea and hbsc untreated. hmwm were increased only during vaso-occlusive crisis in hydroxyureatreated hbsc subject. no ultra-large hmwm were observed. in this preliminary study, in young scd subjects, vwf:ag trended higher in hbss vs. hbsc and in severe hbss participants at a single time-point, but serial evaluations at baseline, in crisis and with optimized diseasemodifying therapy are needed to determine the potential of vwf:ag and hmwm as biomarkers for severity or treatment response. surprisingly, vwf:ag was high in some sickle trait subjects. since hbas is associated with some health challenges such as increased thrombosis risk, further examination of vwf and endothelial dysfunction in sickle trait may provide novel insights into its role as a biomarker. background: the national heart lung & blood institute(nhlbi) guidelines for acute management of voe recommends rapid evaluation and treatment of pain, including administration of a parenteral opioid within -minutes of triage or -minutes from registration, pain reassessment & repeat opioid delivery within - -minutes. inf use has been increasing in peds due to its rapid onset and ease of administration. objectives: to evaluate ped utilization of inf & its effect on intravenous (iv) opioid administration and pain control for the treatment of voe. design/method: a retrospective review of emr was performed on children with scd± years presenting to a ped with voe (pain scores on a - scale) from jan-june . variables studied were median time (iqr, %ci) from ped arrival to first-parenteral-opioid-administration, time-to-first-iv-opioid, first & final pain score, disposition and readmission rate. time-to-first-iv-opioid was also compared to historical data (jan-dec ,n = ) prior to inf protocol initiation. . additionally, % patients received iv opioids within minutes of ed arrival in the inf+iv opioid vs. % in the iv opioids alone group (p< . ). no differences in -hour-returnrates were found in any of the groups, including inf alone group. conclusion: use of inf in the ped for voe is an excellent strategy to shorten time-to-first-parenteral-opioidadministration, improve pain scores & improve adherence to the nhlbi guidelines. however we had distinct unexpected findings: ( ) delays in iv opioid delivery after inf use & ( ) inf alone appeared to provide sufficient pain control without iv opioids for disposition home in % of voe patients. whether the latter reflects insufficient pain management or that there is a milder subgroup for whom inf alone is sufficient, requires further investigation. this study illustrates our experience with a ped-based inf protocol in terms of unanticipated delays in iv opioids and also discharges after inf alone. efforts are underway to further improve use of inf in voe management. st. christopher's hospital for children, philadelphia, pennsylvania, united states background: folate supplementation is commonly included as standard management in patients with sickle cell disease. however, clear evidence supporting the clinical benefits of this practice is lacking. a single study demonstrated improvement on the occurrence of repeat dactylitis at a higher dose of folic acid. to compare clinical outcomes in pediatric patients with sickle cell disease treated with folate supplementation versus those who were not. design/method: this study was a retrospective chart review that included patients to years old with sickle cell disease type ss and s followed at st. christopher's hospital for children. data collected included information about folate supplementation, red cell indices and the presence or absence of clinical outcomes including vaso-occlusive crisis requiring hospitalization in the last six months, acute chest syndrome, infections, asthma, sleep apnea, nephropathy, cerebral vascular disease, stroke and avascular necrosis. analysis of variance (anova) was used to evaluate mean differences between age, number of infections, number of voc events, hemoglobin, reticulocyte count, and mean corpuscular volumes. additionally, chi square analysis was implemented to evaluate differences in folate and non-folate groups for left ventricular remodeling (lvr), sickle cell nephropathy, asthma, obstructive sleep apnea (osa), nocturnal hypoxia, and avascular necrosis (avn). mean differences between the folate and non-folate groups were compared for patients on and off hydroxyurea therapy. one hundred and seven patients met inclusion criteria following review of clinical data. of the patients included in the study, patients were found to be taking folate ( %), while patients were not ( %). statistical analysis showed that there were no significant differences in the incidence of clinical outcomes between patients on folate versus those who were not on folate. of the patients who were not on hydroxyurea, hemoglobin levels were significantly higher in patients on folate versus those who were not (p = . ), but not significantly different for the patients on hydroxyurea. this study suggests that folate supplementation makes no significant impact on the red blood cell indices of anemia nor on the incidence of adverse clinical outcomes in children with sickle cell disease. however, a larger prospective study is needed to guide future considerations for folate supplementation in sickle cell patients in the clinical setting. background: tanzania ranks rd globally for the number of infants born annually with sickle cell disease (scd) but lacks a national newborn screening program. the prevalence of sickle cell trait (sct) and scd is highest in the northwestern regions around lake victoria served by bugando medical centre (bmc) a teaching and consultancy hospital in mwanza. bmc also houses the hiv early infant diagnosis (eid) laboratory that tests dried blood spots (dbs) from hivexposed infants. dbs can be tested for hiv and then retested for sickle cell trait and disease. to determine the prevalence of sickle trait and disease by region and district in northwestern tanzania using existing public health infrastructure. secondary objectives explored associations between sct, scd, malaria and hiv. design/method: the tanzania sickle surveillance study (ts ) is a prospective year-long cross-sectional study of hivexposed infants born in northwestern tanzania, whose dbs collected by the eid program are tested at bmc and available for further testing of sct and scd. samples from children ≤ months of age were tested by isoelectric focusing (ief) and scored independently by two tanzanian staff as normal, sct, scd, variant, or uninterpretable. dbs samples scored as disease or variant were repeated. over the course of months, ief gels have been run. a total of , dbs samples have been scored, including , from children less than -months old. the overall prevalence of sct is . % and the prevalence of scd is . %, along with . % hemoglobin variants. quality of the laboratory results is extremely high, with only . % dbs samples yielding an uninterpretable result. geospatial mapping of the first , samples revealed a regional scd prevalence ranging from . % up to . % among the regions served by bmc. the prevalence of sct and scd is very high in northwestern tanzania. geospatial mapping will identify high prevalence areas where targeted newborn screening can be started using existing public health infrastructure with minimal start-up cost and training. further data will enhance the accuracy of the map to the district level. background: pediatric patients with sickle cell disease (scd) could develop obstructive, restrictive or mixed abnormalities of pulmonary function (pf). several publications report progressive worsening of pf over time, which could lead to severe morbidity in adult patients with sickle cell disease. in adults with sickle cell anemia up to - % of mortality is related to lung disease. early intervention aimed at improvement of lung function could significantly decrease morbidity and possibly improve life expectancy. among disease modifying approaches commonly used in scd are hydroxyurea (hu) and chronic prbc transfusions. both interventions lead to increase of hemoglobin, decrease of hbs fraction, leading to decreased hemolysis. reports of effect of hu on pulmonary function are conflicting with some suggesting no effect and others proposing a slower decline of pulmonary function. the goal of our study is to evaluate effect of disease modifying therapies, like hu and chronic prbc on change of pulmonary function in pediatric patients with sickle cell disease. design/method: this study utilized a retrospective chart review of children with scd who had multiple pfts. we analyzed pfts from patients done during clinic visits. scd patients were divided into three treatment groups: hydroxyurea, chronic transfusions or neither. data was analyzed with linear correlations and analysis of variance (anova). comparison were made between the three groups specifically observing the changes in absolute numbers on pfts over time using the first and last pft the patient had. results: there were a total of patients with multiple pfts (ranging from - ); control ( ), hydroxyurea ( ) and chronic transfusion ( ). the mean changes of the control, and hydroxyurea for the pft parameters fev (- . the chronic transfusion group demonstrated a small improvement in pfts over time for fev ( . ), fvc ( . ), fef - ( . ), however there was a decline in fev /fvc (- . ). however, there was no statistically significant (p-value < . ) in the difference in any pfts parameters between any of the groups. in children with scd there is a decline of pf parameters over time. although no significant differences were seen between the three groups it appears chronic transfusion may improve or limit the decline in pfts. larger studies need to be done to evaluate difference in pf decline in patients with scd patients. background: the use of mobile technology in health care has been a growing trend. patients with chronic diseases such as sickle cell disease (scd) require close monitoring to provide appropriate treatment recommendations and avoid complications. we conducted a feasibility study for patients with scd hospitalized for pain using our self-developed mobile application (tru-pain: technology resources to better understand pain) and a wearable activity tracker. subjective symptoms such as pain and objective data such as heart rate (hr) were measured. we aimed to ) correlate nursing recordings with mobile technology recordings; ) get feedback from patients about usability. design/method: we enrolled patients with scd > years old and < hours from admission for uncomplicated vasoocclusive crisis, excluding patients admitted to icu. patients were given an ipad and a wearable device. they were instructed to record in the application at least once per day and to keep the wearable on, removing only to charge. prior to discharge, patients completed a feasibility questionnaire. we enrolled patients, % females, median age . (range to ) who were admitted for a median days (range to ) for uncomplicated pain crisis. patients used the application throughout hospitalization and made one entry/day (range to ). pain scores recorded via tru-pain correlated well (r = . , p< . ) with pain scores recorded in emr. there was an average of , data points recorded per day, by the wearable, with a maximum of , data points/day. the median amount of hours of wearable data per day was . (maximum of . ). the hr recorded via the wearable correlated significantly with the hr recorded in emr (r = . , p-value < . ). as for usability, % of patients indicated never having a problem with the technology, % found tru-pain 'very easy' or 'somewhat easy' to use, and % were 'very satisfied' with their participation in the study, indicating that it helped them track their pain. our pilot study during hospitalization shows strong potential for using tru-pain for patients with scd. pain data from application and hr from wearable correlated well to the emr data. according to the feedback received, our application was easy to use and helped patients track their pain. despite limitations of battery life, the use of wearable technology is feasible, providing additional data such as activity. we are optimistic that we can continue to improve our tru-pain system to help improve care in patients with scd. background: hydroxyurea, chronic blood transfusion, and bone marrow transplantation can reduce complications, and improve survival in sickle cell disease (scd), but are associated with a significant decisional dilemma because of the inherent risk-benefit tradeoffs, and the lack of comparative studies. these treatments are underutilized leading to avoidable morbidity and premature mortality. there is a need for tools to provide patients high-quality information about their treatment options, the associated risks, and benefits, help them clarify their values, and allow them to share in the process of informed medical decision making. objectives: to develop a health literacy sensitive, web-based, decision aid (ptda) to help patients with scd make informed choices about treatments, and to estimate in a randomized clinical trial the acceptability and effectiveness of the ptda in improving patient knowledge, involvement in decisionmaking and decision-making quality. design/method: we conducted qualitative interviews of scd patients, caregivers, stakeholders, and healthcare providers for a decisional needs assessment to identify decisional conflict, knowledge, expectations, values, support, resources, decision types, timing, stages, and learning, and personal clinical characteristics, and to guide the development of a ptda. transcripts were coded using qsr nvivo . stakeholders completed alpha and beta testing of ptda. we conducted a randomized clinical trial of adults, and of caregivers of pediatric patients to evaluate the comparative efficacy of the ptda, vs. standard of care. results: ptda (www.sickleoptions.org) was developed per decisional needs described by stakeholders and finalized following alpha testing, and beta testing by and stakeholders respectively. in a randomized trial of subjects considering various treatment options, qualitative interviews revealed a high level of usability, acceptability, and utility in education, values clarification, and preparedness for decision making of the ptda. a median % rated the acceptability of ptda as good or excellent and provided narrative comments endorsing the acceptability, ease of use, and utility in preparation for decision making. the ptda met international standards for content, development process, and efficacy with the exception of having a full range of positive and negative experiences in patient stories. compared to baseline ptda group had statistically significant improvement in preparedness for decision making (p = . ) and informed subscale of decisional conflict (p = . ) but not for decisional self-efficacy, knowledge, choice predisposition, or stages of decision-making. a ptda for patients with scd developed following extensive engagement of key stakeholders was found to be acceptable, useful, easy to use, to improve preparedness for decision making, and decrease decisional conflict. background: painful vaso-occlusive crisis (voc) accounts for the majority of emergency department (ed) visits and hos-pitalizations in sickle cell disease (scd). we are interested in studying mental stress and associated autonomic nervous system (ans) imbalance that cause vaso-constriction as possible triggers of scd pain. to this end, we developed a mobile phone application (app) to record daily pain frequency and intensity as clinical endpoints that might be predicted by ans parameters measured in the laboratory. in particular, we think that the aura may represent ans instability that precedes or even triggers change in blood flow and voc. objectives: to assess the feasibility of using an app to evaluate frequency and severity of voc and its potential association with mental stress and presence of aura. design/method: an app was developed for both ios and android systems to allow patients to track pain, stress, and aura. the idea was to create an app that was easy to use with the intent to only capture pain episodes, rather than detailed description of the pain. all scd patients were eligible and a parent version was available for younger children. de-identified data was automatically transferred to a hipaa compliant database via a cloud-based server interfaced to the main research project database. a feedback questionnaire was implemented after at least a month of utilization to assess usability. of the scd patients enrolled, participants utilized the app and of the participants that provided feedback indicated the app was easy to navigate. the mean pain scale was out of (standard deviation . ) for those that entered they had pain that day. although the mean stress level was out of , there was a statistically significant correlation between increasing stress levels and increasing pain scores (p < . ). aura was reported by patients, with patients reporting more than episodes. moreover, on days aura was present there was greater incidence that pain was present as well (p < . ). however, there was no statistically significant association between pain intensity and presence of an aura (p = . ). conclusion: consistent with prior research, reported pain intensity is significantly associated with reported stress intensity. although there was an association between presence of aura and pain, it did not seem to correlate with pain intensity. this uniquely designed app can monitor scd pain clinically and help understand the role of sickle dysautonomia in the genesis of scd pain. university of florida college of medicine, gainesville, florida, united states background: evidenced-based guidelines recommend the emergent evaluation of fever in children with sickle cell disease (scd). as the prevalence of bacteremia has decreased, outpatient management has become more common. however, fever can sometimes herald other complications of scd, such as acute chest syndrome, vaso-occlusive pain crisis, splenic sequestration, or aplastic crisis. institutional practices regarding fever management in scd remain variable, and little is known about the clinical outcomes of children hospitalized for uncomplicated fever. objectives: the primary objective was to determine the rate of bacteremia or scd-related complications per febrile episode in children with scd admitted to a single institution between january and june for uncomplicated fever. this was a retrospective cohort study of febrile patients up to years of age with scd, any genotype, admitted to the university of florida during the defined study period. eligible patients were identified by a database search using admitting diagnosis codes for scd and fever based on the international classification of diseases th and th revisions. encounters were manually reviewed to confirm eligibility. patients were excluded if they had other indications for hospitalization apparent at the time of admission, such as an acute vaso-occlusive episode requiring parental narcotics, asthma exacerbation, or additional complications of scd. the database search identified encounters, of which were excluded based on confounding indications for hospitalization. sixty-three eligible patients accounted for hospitalizations. the median age was years (range weeks- years); . % were male. mean duration of hospitalization was . days (range - days). eight positive blood cultures were identified; six of these were classified as contaminants. bacteremia or the development of a scd-related complication was identified in ( . %) admissions. these included acute chest syndrome (n = ), bacteremia (n = ), splenic sequestration (n = ), and red cell transfusion (n = ). exploratory analyses of potential predictors of bacteremia or scd-related complications showed no association with the presenting white blood cell count or degree of fever (p = . ). of the patients classified as having a scd-related complication, % had hemoglobin ss disease and % had at least one prior documented complication. % of the patients transfused had at least one prior transfusion. conclusion: while improvements in preventative care have substantially lowered rates of bacteremia in children with scd, fever warrants careful evaluation for other acute scdrelated complications. providers should consider inpatient observation in select cases. additional studies are warranted to define subsets of patients suitable for outpatient fever management. background: children with sickle cell disease (scd) exhibit lower neurocognitive functioning than healthy peers, even in the absence of stroke. among the domains commonly affected, working memory (wm) seems particularly affected by disease processes and wm deficits have significant implications for academic achievement and disease selfmanagement. few interventions to improve working memory in pediatric scd have been evaluated. to determine the effects of cogmed, a homebased computerized wm training intervention, in children with scd using a randomized controlled trial design. design/method: participants (ages - ) with scd completed a baseline neuropsychological assessment and those with wm deficits were randomized to either begin cogmed immediately or enter an -week waitlist. cogmed is a homebased intervention completed on an ipad that consists of increasingly challenging exercises targeting visual-spatial and verbal wm, practiced over sessions. at the end of training, participants completed a post-intervention neuropsychological assessment, including tests of visual-spatial and verbal wm from the wechsler intelligence scale for children-fifth edition (wisc-v). results: ninety-one participants (m age = . , sd = . ; % female; % hbss) enrolled in the study; % (n = ) exhibited wm deficits and were randomized to either begin cogmed immediately or wait - weeks before starting cogmed. among those that have received the intervention and reached the end of their training period (n = ), participants ( %) completed at least cogmed sessions, ( %) finished at least sessions, and finished at least sessions ( %). the mean number of completed cogmed sessions was . (sd = . ). paired samples t-tests revealed significant improvements on the working memory index (t[ ] = - . , p = . ) and on the digit span (t[ ] = - . , p = . ), and spatial span-backward (t[ ] = - . , p = . ) subtests. improvements were especially pronounced for participants completing at least sessions. partial correlations controlling for respective baseline scores indicated that the number of cogmed sessions completed was positively correlated with post-test scores on digit span (r = . , p = . ) and spatial span-backward (r = . , p = . ) subtests. among participants who completed at least cogmed sessions, % scored in the average range or higher on the working memory index at the post-intervention assessment, compared to % at baseline. results support the efficacy of cogmed in producing significant improvements in wm. a dose-effect was observed such that participants who completed more cogmed sessions had greater improvements in wm. home-based cognitive training programs may ameliorate scd-related wm deficits but methods for motivating and supporting patients as they complete home-based interventions are needed to enhance adherence and effectiveness. background: sickle cell disease is associated with myriad complications that lead to significant morbidity and early mortality. hydroxyurea has been used successfully to reduce the incidence of these complications and has led to significant improvements in quality and duration of life. at children's minnesota we recommend hydroxyurea in all patients with hb ss/s thalassemia as early as months of age with a goal of starting all patients before months of age. objectives: the purpose of this study was to evaluate the use of hydroxyurea therapy in young patients with sickle cell disease, with particular attention to those children less than one year of age. design/method: a retrospective chart review was conducted on patients less than years of age with sickle cell disease who began hydroxyurea therapy between january , and december , . the study population was divided into three cohorts based upon age at hydroxyurea initiation: cohort ( - year), cohort ( - years), and cohort ( - years). outcomes included laboratory data, clinical events (hospitalization, dactylitis, pain crisis, transfusion, splenic sequestration, acute chest syndrome), and toxicity occurring in the first years of life. results: a total of patients were included in cohorts (n = , mean age . months), (n = , mean age . months), and (n = , mean age . months). patients in cohort had higher hemoglobin (p = . ) and mcv (p = . ) and lower absolute reticulocyte count (p = . ) when compared to cohort . the wbc (p = . , < . ) and anc (p = . , . ) were significantly lower compared to both older cohorts. however, no patient had therapy held because of neutropenia. the mean baseline hemoglobin f in cohort was . % compared to . % and . % in cohorts and respectively (p = . , p< . ). the mean duration of therapy in cohort was . months, compared to . months in cohort (p = . ) and . months in cohort (p = . ). during this time, hb f levels remained higher in cohort (mean . %) compared to cohorts and (mean . %, p = . and mean . %, p = . ). patients in cohort experienced fewer hospitalizations (p = . ), pain crises (p = . ), and transfusions (p = . ). there was no difference in toxicity between groups. hydroxyurea was used safely in infants to months of age and resulted in more robust hematologic responses and a decrease in sickle-related complications when compared with patients starting hydroxyurea later in life. children's national health system, washington, district of columbia, united states background: children with sickle cell disease (scd) have a significantly greater risk of silent or overt cerebral infarction than the general population. infarcts are associated with declines in cognitive functioning and academic achievement. while infarcts are reliably identified using mri, scans are expensive and occasionally necessitate sedation. moreover, mri's are not recommended for routine monitoring of cerebral infarcts. additional tools are needed for discriminating the presence of a cerebral infarct that are brief, noninvasive, inexpensive, and repeatable. objectives: to evaluate differences in performance on cogstate, a computerized neurocognitive assessment, in patients with scd with and without history of cerebral infarct. design/method: participants included children with scd ages - (m = . , sd = . ; % female; % s of s hbss) enrolled in a cognitive intervention trial. participants completed the cogstate pediatric battery, which measures processing speed, sustained attention, verbal learning, working memory, and executive functioning. history of silent or overt infarct was determined via health record review. participants also completed measures of intelligence (iq) and math fluency. results: participants' standard scores across most neurocognitive measures were lower than expected compared to the standardization sample (mean iq = . , sd = . ). thirty percent of participants (n = ) had a documented history of cerebral infarct. participants with a history of cerebral infarct scored lower on cogstate tasks measuring sustained attention (t[ ] = . , p = . ) and executive functioning (t[ ] = . , p = . ), as well as on a measure of math fluency (t[ ] = . , p = . ). receiver operating characteristic (roc) analyses demonstrated that the cogstate task measuring sustained attention was a fair discriminant of patients with and without a history of infarct (auc = . , ci = . - . , p = . ), whereas iq score was not (auc = . , ci = . - . , p = . ). cogstate processing speed and sustained attention tasks fairly discriminated between patients with at least average or below average intelligence (auc = . , ci = . - . , p = . and auc = . , ci = . - . , p = . , respectively). finally, the cogstate processing speed task was good at discriminating between at least average or below average math fluency (auc = . , ci = . - . , p< . ). multiple tasks in the cogstate pediatric battery appear to adequately identify patients with a history of cerebral infarcts. in addition, cogstate tasks appear to be fair predictors of impairments in iq and academic achievement outcomes. cogstate is inexpensive and can be easily administered in a medical setting with minimal training in approximately minutes. results support the potential for cogstate to be used as a screening tool for medical and neuropsychological abnormalities in children with scd. st. christopher's hospital for children, philadelphia, pennsylvania, united states background: cardiovascular disease contributes to the morbidity and mortality of patients with sickle cell disease (scd). hydroxyurea therapy in scd has known clinical efficacy including improving anemia, decreasing episodes of vasoocclusive crisis and acute chest syndrome, and decreasing mortality. effect of hydroxyurea on cardiac function in children with scd is not well studied. an earlier study suggested the protective effect of hydroxyurea on left ventricular (lv) hypertrophy in scd. we hypothesized that hydroxyurea use would be associated with decreased lv remodeling and improved cardiac function. we aimed to evaluate the association between hydroxyurea use and lv remodeling and cardiac dysfunction in children with scd. design/method: we completed a retrospective study of patients with scd who were to years old, followed at st. christopher's hospital for children and had an echocardiogram completed in the past months. data collected included gender, bmi, scd genotype, hydroxyurea use, chronic transfusion use, and d and doppler echocardiographic parameters. cardiac structure, geometry, systolic function, and diastolic function echocardiogram parameters were included. analysis of variance (anova) tests were performed to assess for statistical significance of differences in cardiac parameters between patients with and without hydroxyurea use. analysis of covariance (ancova) tests were performed to control for age. results: demographic and echocardiogram data was collected on all patients who met inclusion criteria. of the patients included, ( %) were on hydroxyurea therapy. patients on hydroxyurea had significantly lower mean relative wall thickness (p = . ) and significantly higher mean peak early lv filling velocities (p = . ) and peak early lv filling/septal annuli early peak (e/ea) velocities (p = . ); however, only the e/ea velocities remained significant when controlling for age (p = . ). mean peak early lv filling velocities approached significance when controlling for age (p = . ). hydroxyurea therapy resulted in a significantly higher e/ea velocity, suggesting that these patients had worse diastolic function. it is possible that the patients initiated on hydroxyurea already had worse disease manifestations than those not on hydroxyurea, possibly accounting for the decreased diastolic function. when controlling for age, hydroxyurea use did not result in significant differences in cardiac structure parameters, systolic function parameters or cardiac geometry. prospective studies and larger sample size are needed to validate our findings, examine for additional statistically significant differences, and develop preventive strategies for cardiovascular disease in children with scd. background: acute chest syndrome (acs) is now the leading cause of death in children with sickle cell disease; mortality in the u.s. is reported to be - % and is mostly due to respiratory failure. early transfusion improves clinical outcomes. although patients with concurrent asthma are considered at increased risk for poor outcomes, risk factors for respiratory failure in pediatric acs have not been well-defined. to determine whether specific epidemiological and clinical features of children hospitalized with acs are predictive of the need for mechanical ventilation. design/method: data from the kids' inpatient database were reviewed to identify patients age < years with a discharge diagnosis of acs for the years , , , and . outcomes were defined by the international classification of diseases, ninth revision, clinical modification code. data were weighted to estimate total annual hospitalizations according to hospital characteristics in the united states. trends in healthcare costs, length of hospital stay, transfusion, and mechanical ventilation use were analyzed using multivariable linear regression. in addition, multivariable logistic regression was used to ascertain specific clinical or epidemiologic factors associated with mechanical ventilation use after adjusting for patient and hospital characteristics. the total hospitalizations for acs were , in ; , in ; , in ; and , in . reported use of mechanical ventilation ranged from . % to . % and was associated with non-black compared to black children (or, . ; %ci, . to . ) and the fall season (or, . ; %ci, . to . ), but not with age, preexisting asthma or hb-genotype. comorbidities of obesity (or, . ; %ci, . to . ), obstructive sleep apnea (or, . ; %ci, . to . ) and heart disease (or, . ; %ci, . to . ) were associated with mechanical ventilation use. the use of simple and exchange transfusion during all acs admissions ranged from . % to . % and . % to . %, respectively. among pediatric acs patients, those with obesity, obstructive sleep apnea or heart disease were at increased risk for respiratory failure and might benefit from early intervention (e.g., transfusion). surprisingly, asthma in children with acs does not appear to be a distinct risk factor for respiratory failure, and further studies are needed to clarify whether differences in treatment approach (e.g., addition of corticosteroids, bronchodilators) might impact on acs progression and/or severity even in high risk patients without asthma. objectives: to compare pulmonary functions between aa and k children with scd and to assess if a high hb f level contributes to better function. design/method: a cross sectional study was done on children with scd (hb ss disease) followed in comprehensive sickle cell programs. aa patients were followed at brookdale hospital, ny and k patients were followed in mubarak hospital, kuwait. children between the ages of and years who had pulmonary function tests (pft) done as a routine screening were enrolled. pft was done using spirometer and plethysmography. patients with congenital or anatomical lung abnormality, heart disease, pulmonary disease such as acute chest syndrome, acute asthma or pneumonia within weeks were excluded. results: there were children ( in each group) with scd,. restrictive pattern on pft was seen in / ( %) of aa vs. / ( %) of k (p> . ). obstructive pattern was seen in / ( %) of aa vs. / ( %) of the k group (p> . ). in both groups, children ( %) had normal pft. three/ ( %) in the aa group had a hb f> % as compared to / ( %) in the k group (p< . ). abnormal pft was noted in / children ( %) in each group. hbf was > % in / ( %) in the aa group vs. / ( %) in the s of s k group (p< . ). in patients with abnormal pft, mean hbf was . ± . in aa group, compared to . ± in k group (p< . ). conclusion: abnormal pft is highly prevalent among children with scd in both groups. aa children are more likely to have restrictive disease and k to have an obstructive pattern. level of hbf did not seem to protect k patients from abnormalities on pft. this finding should emphasize the importance of performing pft as part of the initial evaluation of all children with scd. background: sickle cell disease (scd) is a life-threatening disease with varied clinical spectrum and severity leading to premature death. there is a lack of validated prognostic marker in scd. recent evidence suggests that inflammation and platelet adhesion plays a critical role in the pathophysiology of vaso-occlusion in scd. elevated mean platelet volume(mpv) values are associated with a higher degree of inflammation in many disease states but it's effect on sickle cell disease or it's severity is unknown. objectives: to analyze the role of mpv in predicting disease severity/mortality in pediatric patients with scd. design/method: this is a single center retrospective study and included patients with sickle cell disease between months and years of age during a -year period ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) . demographic information, lab data and clinical information including acute chest syndrome (acs), priapism, transfusions, sepsis, pain crisis, avascular necrosis were collected. all laboratory data were collected in steady state with no crisis in the recent past months. the disease severity score/probability of death was calculated using a validated model to predict risk of death in sickle cell disease (sebastiani et al. blood ) . pearson test was used to analyze correlation between mpv and probability of death. results: total no. of patients = ; male ( . %); female ( . %). median age is . years. all patients were of african-american origin. disease severity, hb ss - ( %); hb sc - ( . %) and sickle-beta thalassemia ( . %). patients on hydroxyurea has significantly lower mpv, p = . and this is independent of hb f levels. mpv has a significant positive correlation with the probability of death, p = . and correlation coefficient, r = . . on subgroup analysis, the correlation is even more significant in the age group between and years, p = . , r = . . using linear regression model, with probability of death as a dependent variable and hydroxyurea, mpv as independent variables, mpv maintains a significant association with probability of death (p = . ). conclusion: mpv is an independent biomarker predicting disease severity and probability of death in pediatric patients with sickle cell disease. hydroxyurea a known disease ameliorating agent is associated with lower mpv values. this effect is independent of the levels of fetal hemoglobin and may be due to anti-inflammatory effect of hydroxyurea or effect on the platelets. background: major success with initial qi projects by the sickle cell care team at children's hospital has precipitated ongoing inclusion of the qi approach to many other aspects of patient care. objectives: to optimize scd patient care utilizing qi processes. design/method: success of the scd qi team's initial project on transcranial doppler studies (tcds) and a second more complex project on hydroxyurea (hu) adherence, led to additional projects on completion of key immunizations, rbc phenotyping, and vitamin d level testing. using similar processes and principles from the hu adherence project, plan-do-study-act (pdsa) cycles were used to conduct smallscale tests of change. patient chart prep sheets, created for bi-monthly pre-appointment chart prep meetings, were significantly modified to include these focused care qi objectives. because of difficulty with emr database capability, data collected from the emr was tracked in excel spreadsheets or other unique tracking vehicles for the various parameters. for example, due to the clinic's diffuse, geographically scattered population, many separate non-shared primary care emrs, and lack of a mandatory state immunization registry; immunization records needed to be retrieved from pcps, outlying hospitals, public health departments, and fqhcs, and added to the emr and excel database. starting in / , all such data was collected and updated monthly. in one year's time ( - ) , the average immunization completion rate for seven key immunizations (pcv , pcv , hepatitis a, hepatitis b, meningococcal a, meningococcal b, and hpv) has increased by %. the biggest improvements were a % and % increase in completion for meningococcal a and meningococcal b, respectively. completion rate for rbc phenotyping rose from . % to . %. patients with at least one vitamin d lab test increased from . % to . %. since starting the tcd project in , the percent of patients who have completed their annual tcd has gone from a baseline of % to a sustained value of > %. conclusion: these qi projects have not only increased adherence to national recommendations for care of scd patients, they have helped establish a scd clinic methodology to create and implement sustainable processes. having the focused care initiatives prominently displayed on the patients' chart prep sheet serve as a reminder to medical team members to check the status of that item. this methodology is currently being used to formulate additional qi projects on annual renal function parameters and specialty visits, such as annual eye and dental exams. background: dominican republic has a high burden of sickle cell disease, and - % of children with homozygous hbss (sickle cell anemia, sca) will develop primary stroke. transcranial doppler (tcd) ultrasonography is an effective screening tool for primary stroke risk, but is not routinely available in dominican republic. hydroxyurea and blood transfusions are available, but no prospective screening and treatment program for stroke prevention has been implemented to date. ( ) to screen a large cohort of children with sca living in dominican republic, using tcd to identify elevated stroke risk; ( ) to determine the effects of treatments for stroke prevention (hydroxyurea for conditional velocities and transfusions for abnormal velocities). we hypothesized that both hydroxyurea and blood transfusions will decrease elevated tcd velocities and help prevent primary stroke. design/method: stroke avoidance for children with república dominicana (sacred, nct ) features a research partnership between cincinnati children's hospital and robert reid cabral children's hospital in dominican republic. the protocol, consent forms, and redcap database were prepared collaboratively and translated into spanish, and then irb approval was obtained at both institutions. in the initial prospective phase, children receive tcd screening over a -month period; those with conditional tcd velocities (maximum time-averaged velocity - cm/sec) receive fixed-dose hydroxyurea at mg/kg/day, followed by dose escalation to maximum tolerated dose, while those with abnormal tcd velocities (≥ cm/sec) receive monthly transfusions for stroke prevention. results: a total of children were enrolled in sacred, with an average age of . ± . years. initial tcd screening revealed ( . %) normal, ( . %) conditional, ( . %) abnormal, and ( . %) inadequate velocities. among children ( males, females, average age . ± . years) who initiated hydroxyurea at mg/kg/day for conditional tcd velocities, completed six months of treatment with expected hematological benefits including significant increases in hemoglobin concentration ( . to . g/dl) and fetal hemoglobin ( . to . %). no clinical strokes have occurred in the treatment group. repeat tcd examination after -months of hydroxyurea treatment revealed % ( / ) with previous conditional velocities had normal tcd velocities. the prevalence of conditional tcd velocities in the dominican republic is high, indicating an elevated stroke risk among children with sca. hydroxyurea treatment is associated with improved hematological parameters, lower tcd velocities, and probable decreased stroke risk. sacred is an important prospective and collaborative research trial providing epidemiological data regarding tcd screening, stroke risk, and hydroxyurea effects among children with sca. background: red blood cell aggregation is a rheologic property that explains the shear-thinning behavior of blood. at lower shear rate blood flow, red cells tend to aggregate, s of s whereas in higher shear rate blood flow, these aggregates are dispersed. this property is especially important in the venous system, where low shear rate blood flow predominates. there is inconsistent data in the literature concerning aggregation and aggregability in sickle cell disease (scd). objectives: because the lorrca and myrenne instruments have been shown to be similarly effective methodologies in red cell aggregation measurements, we aimed to determine whether the measurement of aggregation indices in scd, by myrenne and by lorrca, is consistent in our lab. design/method: we measured aggregation in blood samples corrected to % hematocrit. aggregability was measured using kda dextran in the myrenne but not the lor-rca. aggregation index using lorrca was measured in patients with scd and healthy subjects enrolled in a study of blood flow between and . aggregation and aggregability using the myrenne was measured in patients with scd and healthy subjects enrolled in a separate study of blood flow between and . results: using lorrca, we found that aggregation index in patients with scd was less than that of healthy subjects (p< . ). in the myrenne, aggregation at stasis was slightly higher in patients with scd compared to healthy subjects (p = . ) but aggregation at low shear rotation was not different. aggregability was higher in the patients with scd compared to healthy subjects at both stasis and low shear rotation (p< . ). red cell aggregation is an important determinant of low shear blood flow. deoxygenated venous blood is particularly important to low shear blood flow in patients with sickle cell disease. we found that two different aggregometers predict different aggregation results for scd. it is unclear why there is a systematic difference between the two methods, but there are some possibilities. first, the syllectogram in the lorrca is generated by the backscatter of light from the laser, while the myrenne measures transmitted light. second, the distance between the bob and cup in the lorrca is microns, while the gap between plates in the myrenne is microns, which might affect the disaggregation of red cells. further work is needed to understand the differences in red cell aggregation and aggregability when using these instruments, particularly when using aggregation as a predictor of blood flow and tissue perfusion. background: children with sickle cell disease (scd) are at risk of acute splenic sequestration crisis (assc). assc is a life-threatening complication characterized by splenomegaly, pain and severe anemia. assc most often occurs in young children with the most severe forms of scd and one-third of patients will have more than one episode. treatment is based primarily on expert opinion and includes blood transfusion and surgical splenectomy. objectives: we plan to assess the clinical practice patterns of physicians treating children with assc. design/method: a survey study was performed. the survey included six scenarios of severe scd with variation in age, hydroxyurea-use, and episode number of assc; questions focused on the acute and chronic management of assc. the survey was disseminated on three occasions over a six-month period, using an online survey tool, surveymonkey, to pediatric hematologist-oncologists participating in the american society of pediatric hematology-oncology hemoglobinopathy special interest group. the survey had a response rate of % ( / ). most respondents were recent graduates ( %; / ) practicing in academic urban centers with greater than sickle-cell patients. seventy-nine percent ( / ) recommended hydroxyurea initiation in - m/o with severe scd. prophylactic penicillin after surgical splenectomy was continued by % ( / ) after years. for the acute management of assc results did not vary despite patient age, hydroxyurea use, and the number of previous assc episodes. simple transfusion was preferred by % ( / ), with % ( / ) recommending slow transfusion and % ( / ) recommending routine simple transfusion. for the chronic management of assc, results varied based on patient age and the number of previous assc episodes. for a m/o after the first episode, % ( / ) recommended observation and % ( / ) hydroxyurea initiation. for a m/o with any prior episode of assc, % ( / ) recommended chronic transfusion therapy and % ( / ) surgical referral for splenectomy. for a y/o after the first episode, % ( / ) recommended surgical splenectomy and % ( / ) increasing hydroxyurea dose. for a y/o with any prior assc episode, % ( / ) recommended referral for surgical splenectomy. in this survey, we found most providers continue to recommend simple transfusions for assc and surgical splenectomy after two episodes. the majority of providers continue to delay referral for surgical splenectomy until age two, but earlier referral in children under two and use of chronic transfusion therapy were also reported. variability in chronic management highlights the need for further research of splenic sequestration. background: developing therapies for sickle cell disease (scd) is challenging in part because the accepted endpoint, vaso-occlusive crisis (voc), occurs infrequently, does not measure full disease burden, and is a measure of healthcare utilization. in phase / studies of patients with scd, voxelotor (gbt ) has demonstrated increased hemoglobin (hb) levels and reduced hemolysis and has been safe and welltolerated. voxelotor is being evaluated in the ongoing hope phase trial. objectives: to report the innovative phase / hope trial design with novel primary and secondary outcomes to accelerate drug development. design/method: hope (nct ) is a phase , randomized, placebo-controlled, multicenter study of oral voxelotor in patients with scd (aged - years) with baseline hb . - . g/dl and - episodes of voc in the prior year. to accelerate clinical trials to support drug development, the study combines a phase exploratory, dose-selection phase (group ) with a pivotal phase (groups / ). patients in group will be randomized : : to voxelotor or mg/day or placebo. analysis for dose selection will occur when the final patient has received weeks of treatment. group will continue enrollment with randomization : : until dose selection based on analysis of the group cohort. group will allow for a seamless transition into group , which will randomize patients : to the selected dose or placebo. the final data analysis set will include group patients who received placebo or the selected dose and all group patients. the primary endpoint is an objective laboratory measure and surrogate of clinical benefit, increase in hb > g/dl, from baseline to weeks based on voxelotor mechanism of action (inhibition of hb polymerization). this trial is the first to use a patient-reported outcome (pro), the -item sickle cell disease severity measure, as a secondary endpoint. this novel electronic pro, developed specifically for the hope study following fda guidance, will evaluate changes in scd symptom exacerbation and total symptom score from baseline to weeks. additional secondary endpoints include measures of hemolysis, rates of voc, transfusions, and opioid use. the study was designed to enable selection of pro-defined symptom exacerbations or traditionally defined voc as the key secondary endpoint after the group analysis. results: this study is ongoing. the hope trial, expected to complete enrollment by late , will evaluate the efficacy and safety of voxelotor compared with placebo in patients with scd. supported by global blood therapeutics. background: inflammation, coagulation activation, oxidative stress and blood cell adhesion are elements of sickle cell disease (scd) pathophysiology. patients with scd have low levels of the omega- fatty docosahexaenoic acid (dha) and eicosatetraenoic acid (epa) in plasma and blood cell membranes. dha is a bioactive fatty acid with anti-inflammatory, anti-blood cell adhesion and anti-oxidant properties. altemi-atm is a novel dha ethyl ester formulation with a proprietary delivery platform (advanced lipid technology® (alt®)) that enhances oral dha bioavailability. the scot trial investigated the effects of altemiatm in children with scd. objectives: to demonstrate the effects of altemiatm on blood cell membrane omega- index and selected biomarkers of inflammation, coagulation, adhesion and haemolysis associated with scd. s of s design/method: children with scd, aged - years (n = ), were enrolled. subjects were randomized to receive either placebo or one of three daily oral doses of altemiatm ( - , - or - mg/kg/day dha) for two months. the effects of altemiatm on red blood cell (rbc), white blood cell and platelet membrane omega- fatty acids index (total dha + epa levels) were assessed after four weeks of treatment. the effects of altemiatm on markers of inflammation, adhesion, coagulation, and hemolysis were assessed after eight weeks of treatment. cell membrane dha and epa concentration was determined by using lc-ms/ms method. the percent changes from baseline on blood cell membrane omega- index and select scd biomarkers were compared between the three dose groups and placebo using a mixed-model repeatedmeasures (mmrm) analysis with baseline blood cell membrane omega- index, hydroxyurea use, and treatment as fixed effects and patient as a random effect. after four weeks of treatment, blood cell membrane dha and epa levels were significantly increased in all altemiatm doses (p< . ). after eight weeks of treatment, significant reductions were observed in se-selectin (p = . ), and d-dimer (p = . ) in patients exposed to altemiatm dose level vs. placebo. hemoglobin was significantly increased at altemiatm dose level versus placebo. plasma high-sensitivity c-reactive protein, lactate dehydrogenase, soluble vascular cell adhesion molecule- and white blood cell count showed improvement after weeks of treatment in all three altemiatm doses levels but did not reach significance. conclusion: treatment with altemiatm enriches dha and epa in blood cell membranes of patients with scd and improves select sickle cell disease biomarkers of blood cell adhesion and thrombin generation. these findings provide insight into the mechanisms of action of altemiatm in sickle cell disease. brown university -hasbro children's hospital, providence, rhode island, united states background: despite clinical advances in the treatment of sickle cell disease (scd) in pediatric and young adult patients, pain remains a significant source of disease-related morbidity. physical therapy has been shown to be useful for the treatment of pain in children and young adults with various chronic illnesses of which pain is a significant component, however no data exists regarding potential benefits of physical therapy in pediatric and young adult patients with scd. objectives: to query healthcare providers and others involved in the care of pediatric and young adult scd patients regarding possible benefits of and barriers to physical therapy as a potential treatment modality. design/method: we conducted a web-based survey of healthcare providers within the new england pediatric sickle cell consortium (nepscc) in an attempt to identify potential benefits of and barriers to outpatient physical therapy in this patient population. results: nearly % of survey participants felt that physical therapy had the potential to be "somewhat beneficial" or "very beneficial" in pediatric and young adult patients with scd. a majority of physicians reported having referred patients with scd for physical therapy in the past. the most frequently identified perceived potential benefits included improved functional mobility, improvement of chronic pain symptoms, decreased use of opiates, improved mood symptoms, improved acute pain symptoms, and improved adherence with medications and clinic visits. significant perceived barriers identified included lack of transportation, time constraints, patient lack of understanding, and difficulty with insurance coverage. our study indicates that healthcare providers have an overwhelmingly positive view of the use of physical therapy in the management of pediatric and young adult patients with scd. significant barriers exist which need to be addressed. future research should focus on patient and parent perspectives regarding physical therapy, as well as a randomized controlled trial of a physical therapy intervention in this patient population. background: vitamin-d deficiency is fast becoming increasingly recognized in patients with sickle cell disease (scd). while it is estimated that these patients are five times more likely to develop vitamin-d deficiency, the exact clinical significance of this is largely unknown. given that this deficiency can be inexpensively and easily treated, our study sought to establish the prevalence of vitamin-d deficiency in our patient population and its relationship with disease severity. objectives: to estimate the prevalence of vitamin-d deficiency in patients with scd in our institution and to analyze their disease severity in relation to their vitamin-d level. design/method: through retrospective chart review we analyzed subjects that represent a cohort of patients followed at the adult and pediatric hematology services at university of miami with known diagnosis of scd that had a vitamin-d level drawn between january st, and august st, . we conducted a cross-sectional study and recorded the first vitamin-d level during this period. patient demographics, medical and social history information were collected along with laboratory data. the number of admissions for vaso-occlusive crisis (voc) and acute chest syndrome within one year preceding the collection the vitamin-d level was also recorded. results: a total of charts were reviewed, adult charts and pediatric charts. after exclusion, patients were enrolled. subclinical vitamin-d deficiency is only evident on laboratory blood testing of vitamin-d ( -hydroxy) and according to this laboratory result patients were classified as sufficient (≥ ng/ml), insufficient (< to ng/ml) and deficient (< ng/ml). out of the cases, . % ( / ) were deficient, . % ( / ) were insufficient and . % ( / ) were optimal. after statistical analysis two negative correlations were identified, increasing vitamin-d levels with decreasing white blood cell count (ci %- . (- . , - . )and decreasing incidence voc (ci %- . (- . , - . ). conclusion: this study confirms that there is a significant prevalence of vitamin-d deficiency in patients with scd. furthermore, the results of this investigation proved that vitamin-d deficiency is associated with acute pain and leukocytosis in patients with scd. given the multitude of confounding factors that affect vitamin-d absorption and intake, multivariate analyses are required to truly further investigate this relationship. texas children's hospital, houston, texas, united states background: hemophagocytic lymphohistiocytosis (hlh) is a rare but life-threatening condition of hyper-inflammation that is characterized by splenomegaly, cytopenias, hyperferritinemia, hypertriglyceridemia, hemophagocytosis and coagulopathy. although timely diagnosis is imperative, it is often challenging as these individual signs and symptoms may occur in a variety of clinical conditions. to report a case of undiagnosed sickle cell anemia presenting with severe ebv viremia and associated hemophagocytic lymphohistiocytosis results: a -month-old previously healthy male presented with respiratory distress, increased fatigue, and a focal seizure following a two-week history of cough and lowgrade fevers. physical exam was consistent with hypovolemic shock and revealed significant splenomegaly. laboratory testing revealed severe hypoglycemia, acidosis and electrolyte disturbances including hyperkalemia, hyperphosphatemia, and hyperuricemia. labs showed a leukocytosis (wbc , ), severely low hemoglobin ( . ), and platelets of , . coagulation testing revealed prolonged pt/inr and ptt, hypofibrinogenemia and a highly elevated d-dimer. additional workup was completed to determine etiology of acute presentation, given broad differential diagnosis. infectious studies were consistent with an acute ebv infection (plasma ebv pcr > , ). elevated levels of soluble il- and ferritin completed / criteria for the diagnosis of hlh. bone marrow evaluation showed trilineage hematopoiesis with no abnormal blast population or hemophagocytosis. results from hemoglobin electrophoresis sent from the initial cbc sample were notable for hbs . %, hbf . %, and hba of %, confirming the diagnosis of sickle cell disease. the patient was started on hydroxyurea and penicillin and splenomegaly resolved. with supportive care, he demonstrated gradual improvement in symptoms and laboratory abnormalities, including normalization of soluble il- , ferritin, cd , il- levels, immunoglobulins, and declining ebv titers. nk cell function has remained abnormally low, not eliminating the possibility of acquired hlh despite spontaneous improvement. conclusion: splenic sequestration associated with sickle cell disease in combination with acute infectious mononucleosis could have explained many of the presenting symptoms including anemia, thrombocytopenia, and splenomegaly. however, it does not explain the unusually high ebv titer and degree of inflammation meeting diagnostic criteria for hlh, which raises concern for an underlying immunologic abnormality such as x-linked lymphoproliferative disorder (xlp). although testing for xlp was negative, he will require s of s continued monitoring in the future for signs of relapse. this case illustrates the complexity of diagnosing lymphohistiocytic disorders and the significant overlap in presentation between these disorders and other medical conditions. background: vaso-occlusive crisis (voc) is one of the most distressing occurrences in patients with sickle cell disease (scd). patient controlled analgesia (pca) is recommended by nih and expert opinions favor its early use. we aim to review the use of pca in patients with voc and to evaluate if its early use is associated with faster pain control and reduced length of stay (los). design/method: this retrospective single center study included all pediatric patients admitted and treated with pca for a severe voc from to . "early" use was defined as start of pca within hours of arrival in the emergency department (ed) and "late" use after hours. time to reach adequate analgesia was defined as oucher, verbal scale or faces pain scale < / obtained twice consecutively in a -hours interval. time to reach adequate analgesia and los were compared between early-pca and late-pca groups. results: a total of patients presented episodes of voc treated with pca during the study. sixty-one episodes ( %) were treated with early-pca and ( %) with late-pca. both groups were comparable in terms of age ( . vs . years old), gender ( . % female vs . %), hemoglobin phenotype ( . % hbss vs . %), but median pain score at admission was higher in early-pca than in late-pca ( / vs / , median difference ( % ci , ). early-pca was associated with a median reduction in los of . days ( % ci . , . ) (median early-pca los . vs late-pca . days). time to reach analgesia could be evaluated only in a subset of patients ( in early-pca and in late-pca group). although time to reach adequate analgesia tended to be shorter in the early-pca group, it was not statistically different: median . hours vs . hours, difference of . ( % ci - . , . ). side effects were observed during ( . %) pca treatments ( / ( . %) episodes in early-pca, / ( . %) in late-pca group) among which ( . %) were significant adverse events. these were observed in patients who required interventions: desaturations requiring oxygen without intubation, neurologic abnormalities (hallucinations, visual abnormalities, no stroke), urinary retentions. conclusion: early use of pca for severe voc was associated with a reduced length of hospital stay despite that these patients had higher pain score on admission. prospective studies are needed to support these positive outcomes. background: acute chest syndrome is one of the leading causes of death in children with sickle cell disease - . while the cause of acute chest syndrome most commonly is not identified, fat embolism and infectious causes are believed to be most common. with an extremely high mortality rate, rapid identification and initiation of therapy is essential for survival. case presentation: we describe the case of an -year-old female with sickle cell sc disease who was admitted for vasoocclusive pain crisis and quickly progressed to multi-system organ failure due to fat embolism syndrome and parvovirus b infection objectives: the case highlights the presentation and diagnosis so other providers can optimize outcomes for those with this under-recognized syndrome design/method: her parvovirus studies returned after days which showed: parvovirus b dna pcr detected; parvo igg . (positive > . ); and igm . (positive > . ). the patient experienced an approximately . g/dl drop in hemoglobin( . to . g/dl/ hrs) with progressive thrombocytopenia (from , to , /ul) and a peripheral smear showed microcytic,normochromic red cells with nucleated rbcs and occasional nuclear budding, slight polychromasia, schistocytes, and polymorphic cells with toxic granules that suggested leukoerythroblastosis. she was emergently transferred to the regional quaternary care hospital for ongoing ecmo therapy where she experienced a change in her pupillary exam prompting a stat ct scan that showed severe, diffuse cerebral edema with transtentorial herniation. the decision was made to withdraw life-sustaining therapies and her family refused a post-mortem autopsy examination. fat embolism syndrome is a severe and uncommonly recognized complication of sickle cell disease, seen most commonly in those with a non-ss phenotype and previous mild disease course who present with severe, unrelenting vaso-occlusive pain episode and/or acute chest syndrome that progresses to respiratory distress with altered mental status and cutaneous changes. rapid identification and initiation of exchange transfusion therapy should be initiated with clinical suspicion because of the extremely high mortality rate. although previously considered rare, it needs to be considered in the differential diagnosis of more commonly encountered complications of sickle cell disease. background: patients with sickle cell disease (scd) experience vaso-occlusive crisis (voc), which results in extreme pain, often requiring opioids and admission. genetic and environmental factors affect the frequency and severity of these episodes. previous research has born conflicting evidence on whether environmental temperature is contributory. edmonton, alberta is the northern most city with a population over a million in north america. there is an increasing sickle cell population which is exposed to extreme winter conditions. this provides a suitable population and atmosphere to study the influence on cold external temperatures in scd. this study sought to identify if pediatric patients with scd, experience greater morbidity in cold external temperatures. board approved retrospective case control series. patients were identified through a clinical database, and emergency visit, phone call and admission data was collected over a fiveyear period. the average, minimum and change in temperature on day of presentation, and hours prior, was collected from the government of alberta, and was statistically analyzed using descriptive statistics, to determine the relation to vaso-occlusive events. results: one-hundred and eighteen patients were identified, and voc events reviewed. the mean patient age was . years of age with a range from . - years old. the female to male ratio was equivalent with female ( . %) and male ( . %) voc events. eight records ( %) had docu-mented cold exposures. the analysis between the temperature and the frequency of events did not yield significant correlation. average and minimum temperature on day of admission had the largest percentage of voc events occur at mild temperatures, from - . to • c and - . to respectively. change in temperature on day of admission, and hours had the largest percentage of voc events at a mild to moderate change in temperature of - degrees. data at & hours prior to admission showed similar results. secondary data analysis accounting for the lower proportion of extreme weather days in comparison to moderate temperate days showed no significant impact. there was no correlation of average, minimum or change in temperature on day of admission, or hours prior. multiple cofounding factors likely contribute to these results. as it was a retrospective study many confounding and precipitant factors may not be recorded or identified. a prospective study to better record specific cold exposure is warranted. children's national health system, washington, district of columbia, united states background: achieving optimal anticoagulation with unfractionated heparin (ufh) in pediatric patients receiving extracorporeal membrane oxygenation (ecmo) is often challenging due to antithrombin (at)-mediated heparin resistance (hr). intermittent at dosing during pediatric ecmo support does not maintain adequate at levels. continuous at infusion (cati) presents an alternative strategy to achieving consistent goal at levels and optimizing heparinization. however, cati during pediatric ecmo has not been adequately studied. objectives: to describe our center's experience with an ecmo cati protocol. design/method: in , we modified our ecmo anticoagulation protocols to include ufh titration according to anti-factor xa (anti-fxa) levels and cati in patients with at-mediated hr. the cati rate was calculated using baseline and goal at levels while accounting for the circuit volume. cati was administered with ufh into the circuit via a s of s y-infusion set. at and anti-fxa levels were monitored every hours. recombinant at (r-at) concentrate was used at our center until with subsequent transition to a plasmaderived at (pd-at) concentrate. due to the longer half-life of pd-at concentrate, the protocol was modified so cati is stopped once target at and anti-fxa levels are achieved. we conducted a retrospective study of all patients who received cati during ecmo support at our center. data are reported as median and interquartile range and compared using the mann-whitney u test. two-tailed p-value < . was considered statistically significant. since , patients [ males, age month ( . - )] on ecmo support received catis ( rat, pd-at) per our protocol ( patients received pd-at infusions during one ecmo run). the duration of cati was hours ( - ). cati administration led to significant increases in at and anti-fxa levels from baseline of % ( - ) and . units/ml ( . - . ) to the first level within goal of % ( - ) and . units/ml ( . - . ), respectively (p< . ). the respective times to achieve goal at and anti-fxa levels were hours ( - ) and hours ( - ). the respective peak at and anti-fxa levels were % ( - ) and . units/ml ( . - . ). during cati, no patient required circuit change, patient developed cannula thrombosis and patients experienced non-fatal major bleeding. conclusion: cati in pediatric patients receiving ecmo support with close monitoring of at and anti-fxa levels was associated with significant rapid increase in at, optimization of heparin effect, and reduction in thrombotic complications without increase in major bleeding compared to prior reports. a prospective study of this at dosing strategy is warranted. children's hospital of orange county, orange, california, united states background: inherited factor xiii (f ) deficiency is a rare bleeding disorder with wide heterogeneity in clinical manifestations ranging from mild bruising, and mucosal and umbilical stump bleeding to spontaneous, severe intracranial bleeding. the bleeding phenotype is influenced not just by zygosity of the fxiii mutation alone, but also by co-inheritance of variants in other clotting protein genes that also play a major role in clot formation and stability. we present a series of three siblings found with f a gene variant and platelet dysfunction linked to bleeding phenotype. design/method: retrospective chart review of the index case, coagulation studies and whole gene sequencing. the index patient presented at two years of age with a subdural hematoma after a fall, requiring emergent craniotomy. a week after initial evacuation, she re-bled, prompting an extensive work-up for potential bleeding disorders, including f activity, von willebrand profile, comprehensive fibrinolysis panel, pai- antigen level, platelet mapping thromboelastogram (plt-teg), and f genetic analysis. the patient's identical twin and older sibling, who had symptoms of bruising, underwent a similar evaluation. the index patient demonstrated consistently low f activity ( - %), and platelet function testing revealed decreased response to adp agonists. the twin and older sibling had normal f levels, and only slightly decreased response to adp in platelet studies. whole gene analysis of f and other genes on our next generation panel, revealed several intronic deletions in the index patient that were not shared by her siblings, which likely account for her decrease in circulating f levels. her symptoms have responded well to monthly treatment with factor concentrate. all three children shared the f variant, pro leu, previously described as a risk factor for intracranial hemorrhage. the f mutation, pro leu, has been associated with intracranial hemorrhage in young women, but the presence of the variant alone may not be enough to cause a severe bleeding phenotype. family studies identified novel deletions in the index patient which may account for her decreased f levels, which would have been overlooked with standard sequencing. future studies, including evaluation of 'platelet' f levels, should be performed when platelet dysfunction is detected. further laboratory and clinical evaluation is required to delineate the long term implications of the interaction of even mild f deficiency if present with additional clotting disorders such as the platelet function defect in these siblings. background: acquired hemolytic anemia can occur due to mechanical shearing of red blood cells and is classically seen in patients with prosthetic heart valves. there are reports of this same traumatic effect with other repairs, including annuloplasty. following valvular procedures flow disturbances can exist across the valve that lead to shear stress and hemolysis. although von willebrand disease (vwd) is typically seen due to an inherited disorder in the pediatric population, flow disturbances in the setting of valve abnormalities can lead to acquired von willebrand syndrome (avws). von willebrand factor multimers become unfolded and elongated in the setting of shear stress resulting in increased susceptibility to cleavage by adamsts- . specifically, loss of high molecular weight multimers (hmwms) can lead to a syndrome akin to type a vwd. objectives: to describe a case of mechanical hemolysis with acquired type a vwd design/method: a -month-old girl with history of hypoplastic left heart syndrome and severe tricuspid valve insufficiency underwent norwood procedure, blalok-taussig shunt placement and subsequently a bidirectional glenn and tricuspid valve annuloplasty. during the following month she requires weekly red blood cell (rbc) transfusions due to intermittent anemia. she also experienced bloody stools and dark urine. laboratory evaluation was notable for normocytic anemia, reticulocytosis, elevated lactate dehydrogenase, and low haptoglobin consistent with hemolytic process. immune-mediated hemolysis from transfusion reaction or presence of autoimmune or alloimmune antibodies testing was negative. to investigate gi bleeding, work up for vwd revealed normal vw activity and antigen but with loss of high molecular weight multimers consistent with acquired type a vwd. in consultation with cardiology, it was felt her tricuspid valve insufficiency jet could be leading to mechanical hemolysis and avws. a repeat echo showed persistent moderate tricuspid insufficiency but no other significant changes. due to the patient's continued need for weekly rbc transfusions she was subsequently trialed on pentoxifylline which is used in adult patients to decrease blood viscosity and increase erythrocyte flexibility in patients with mechanical hemolysis. her transfusion needs remained the same and the medication was discontinued after two weeks. she required one transfusion a week later but no transfusions since that time. although not commonly seen in pediatric patients, the diagnosis of mechanical hemolysis accompanied by avws should be pursued in a patient with congenital heart disease with significant anemia and/or bleeding. the work up in these patients is difficult as echocardiograms can be inconclusive thus an extensive hematologic evaluation is usually necessary. objectives: our aim was to assess incidence of and potential risk factors for central line-related dvt at our institution between - . additionally, our goal was to analyze if that incidence differed between the three central line types and identification of line-specific risks. design/method: a retrospective chart review of central line placements in pediatric patients at cleveland clinic between - was conducted. data included demographics, potential risk factors, line characteristics and any related thrombotic events. the study cohort consisted of lines in pediatric patients aged - years of age. there were . thrombi ( % ci . - . ) per , line days. statistically significant risk factors for thrombus include diagnosis group (liquid tumor highest rate of %, solid tumor lowest at %), type of line (picc %, broviac %, and mediport %), location of line, greater number of lines per patient, peg asparaginase ( % vs %), sepsis, and history of procoagulant state. line characteristics such as lumen size and number of lumens were not identified as a significant risk. there was a significantly higher rate of thrombus in than in the previous years when pooled ( % in vs . % from - , p = . ). the incidence of dvt in pediatric patients at our institution was highest with broviac lines, and significant risk factors in our patient population included liquid tumor, femoral vein location, peg asparaginase, sepsis, and history of a procoagulant state. the incidence of thrombi was highest in , and therefore highlights the urgent need for improvement in nationwide hospital practices to minimize risk of thrombi formation and early detection in the higher-risk s of s populations. there is still much to be learned regarding the characteristics specific to different central lines, which would influence thrombi formation. nyu winthrop hospital, mineola, new york, united states background: pediatric immune thrombocytopenic purpura (itp) is an autoimmune disorder with platelet counts < causing increased risk for significant hemorrhage. there is increased immunologic platelet destruction due to production of specific autoantibodies along with inhibition of platelet production. few randomized trials exist to guide management and ultimately each patient requires an individualized treatment plan. itp may be acute (diagnosis to m) or chronic (> months). one of the treatments of chronic itp is laparoscopic splenectomy (ls), which is very well tolerated. a rare complication of ls is splenosis, an autotransplantation or implantation of ectopic splenic tissue within the abdominal cavity or in any other unusual body compartment. splenosis is sometimes associated with relapsed itp due to preserved immune activity. the usual management of symptomatic splenosis is surgical resection. objectives: to describe medical management in a young patient with itp relapsed due to extensive unresectable splenosis following ls design/method: our patient was originally diagnosed at years with itp and was treated with ls at years of age for chronic severe thrombocytopenia and persistent bleeding not responding to first line therapies. she tolerated it well and had a complete response (cr) defined as a platelet count of > measured on occasions > days apart and absence of bleeding. she maintained a normal platelet count for twelve years after which she relapsed (loss of response after cr) with severe thrombocytopenia and hematuria necessitating high dose steroids. ct scans showed multiple wellcircumscribed soft tissue masses in the left lower quadrant adjacent to uterus and left ovary, involving left omentum and the anterior abdominal wall partly. findings were confirmed by damaged rbc nuclear scan to be splenosis. during laparoscopy the splenosis lesions were deemed too extensive and were not resected completely to avoid postoperative morbidity. she was started on sirolimus around the same time for treatment of her relapsed itp and steroids were weaned off. results: eight months since beginning sirolimus with therapeutic levels she remains in cr with no bleeding and has not required any steroids, immunoglobulins or anti d immunoglobulin. conclusion: sirolimus is a safe and effective steroid-sparing agent in treatment of chronic itp. this is the first instance of a patient with poorly resectable splenosis responding well to medications for itp. more data is needed regarding the longterm efficacy of such an intervention and whether it will eliminate the need for a second surgery in relapsed itp patients with extensive splenosis. background: storage pool disorders affecting platelets result in bleeding symptoms related to a deficiency or defect in alpha granules or delta granules. in delta-storage pool disorders (dspd,) there is a deficiency of the delta granules and their constituents, which results in the inability of platelets to properly activate as well as lack of proper constriction of blood vessels during bleeding episodes. amongst patients with dspd, females most commonly present with menorrhagia, while males tend to present with epistaxis and easy bruising. the international society on thrombosis and hemostasis (isth) developed a screening bleeding assessment tool (bat) for mild bleeding disorders, shown to be a validated tool in children. diagnosis of dspd is classically made with a platelet electron microscopy (pem) value < . delta granules per platelet (dg/pl), but recently lower diagnostic thresholds of dg/pl or even . dg/pl have been suggested. objectives: evaluate the correlation between pem and bleeding scores, and also examine various cut-off values used to diagnose and risk stratify patients with dspd. design/method: retrospective chart review of pediatric patients followed by hematology with a diagnosis of dspd was performed. clinicians obtained bleeding scores for each patient as standard of care in the hemostasis clinic. quartile ranges were established to appropriate three stages of severity based upon bleeding scores. statistical analysis was performed using software r and exploratory data analysis to evaluate for a correlation. results: amongst all patients, the average bat score was . and pem was . dg/pl. the average bleeding score for pem between . dg/pl and dg/pl was . , while the average bleeding score for pem below dg/pl was . . the correlation coefficient between pem and bleeding scores is . . using a threshold of dg/pl, % of patients would have met diagnostic criteria. quartile ranges for the bleeding scores are as follows: st quartile was - , nd quartile was - , and rd quartile was > . conclusion: patients with a more marked granule deficiency do not exhibit a more severe bleeding phenotype, suggesting proper platelet function is not solely determined by granule quantity in these patients. bleeding severity may be more appropriately assessed with bleeding scores rather than pem values, and using quartile ranges may aide in risk stratification and therapeutic interventions for dspd patients. further work remains to determine the optimal diagnostic threshold of pem dspd in pediatric populations. texas children's hospital, houston, texas, united states background: warfarin management has many challenging aspects including pharmacogenomics, food and drug interactions, lack of standardized dosing, patient compliance, tracking lab results from multiple lab locations, and the potential for significant bleeding or thrombotic complications. a literature review revealed limited data highlighting anticoagulation monitoring workflow and emr documentation and specifically, no data in the pediatric population. historically, the texas children's hospital cardiology and hematology centers were each documenting anticoagulation data within the epic tm system differently. epic's tm original design for anticoagulation documenting resulted in the necessity to duplicate documentation in order to see at-a-glance critical anticoagulation monitoring information. objectives: the objective of this project was to standardize inr documentation across departments to reduce the risk of patient safety events and improve workflow. design/method: a workgroup assembled consisting of nurses from the cardiology and hematology departments, along with staff members from the epic tm is support group. the workgroup identified current documentation practices, available epic tm tools, and brainstormed ideas to streamline and improve both documentation with the current epic tm tools. physician partners were identified in cardiology, hematology and coagulation laboratory to gain their input. a new anti-coag (ac) encounter was developed and first made available in an epic tm practice environment, then once approved, epic tm written education and training session were completed by both departments' staff. results: surveys were sent to health care providers in the cardiology and hematology centers prior to the new ac encounter, and also to health care providers six months after implementing the ac encounter. six responses were received for each survey. the pre-implementation survey showed the most problematic part of the documentation system for anticoagulation was no single place in the emr to find a complete anticoagulation picture. post ac encounter implementation survey results revealed more health care providers using the epic tm inr reminder pool, less time needed to compile a report of three months of anticoagulation information, less time needed to document individual encounters, less locations needed to document ac information and decreased amount of types of documentation used. standardized ac encounters improves workflow with less time needed to document and compile information, less types of documentation utilized and easier access to patients ac information. next steps include retrospective review of patients' inr time in therapeutic range to determine if there was an impact on patient compliance and continue to evaluate and modify the ac encounter to enhance user friendliness. caitlin tydings, jennifer meldau, christine guelcher, carole hennessey, eena kapoor, michael guerrera, yaser diab s of s children's national health system, washington, district of columbia, united states background: venous anatomic abnormalities (vaas) are considered a risk factor for developing deep vein thromboses (dvts) that occur as a result of significant alterations in venous blood flow. identification of predisposing vaas can be challenging. hence, diagnosis can be delayed or overlooked especially in pediatric patients. dvts in children or adolescents with predisposing vaas have been only described in sporadic case reports and small case series. objectives: to describe characteristics and outcomes of dvts in pediatric patients with underlying vaa treated at our center. design/method: we conducted a retrospective chart review of all pediatric patients with objectively confirmed extremity dvt treated at our institution over a -year period from to and identified all patients with underlying vaas. patients were managed according to standardized institutional protocols based on published guidelines. post-thrombotic syndrome (pts) was assessed at our center using the manco-johnson instrument. relevant data were collected and summarized using descriptive statistics. during the study period, of pediatric patients ( %) [ females, median age years (range - )] diagnosed with extremity dvt at our center were found to have an underlying vaa. vaas included may-thurner anomaly ( patients), venous thoracic outlet obstruction ( patients) and inferior vena cava (ivc) atresia ( patients). additional provoking factors were identified in patients at time of presentation. dvt locations included upper extremity veins ( patients), lower extremity veins ( patients) and lower extremity veins and ivc ( patients). the majority of dvts [ patients, ( %)] were completely occlusive. high risk thrombophilia (defined as inherited deficiency of antithrombin, protein c, or protein s, or antiphospholipid antibody syndrome) was present in patients ( %). all patients were treated with therapeutic anticoagulation with patients continuing indefinite anticoagulation. endovascular interventions were performed in patients and included percutaneous pharmacomechanical thrombectomy and/or catheter-directed thrombolysis ( patients), balloon angioplasty ( patients) and stent angioplasty ( patients). surgical interventions included thoracic decompressive surgery ( patients) and surgical thrombectomy ( patient vvas represent an important risk factor for developing extensive extremity dvt in adolescents. this special population is at risk for short-term and long-term com-plications. early identification and correction of vaas may improve outcomes. however, multicenter, prospective studies are needed for developing optimal evidence-based treatment approaches. alexander glaros, roland chu, sureyya savasan, meera chitlur, madhvi rajpurkar, yaddanapudi ravindranath children's hospital of michigan, detroit, michigan, united states background: acute budd-chiari syndrome (bcs) is a rare thrombotic emergency in children, and etiologies/treatment are less well-defined than in adults. in adults, a systematic approach including anticoagulation, relief of venous obstruction, and treatment of the underlying cause has proven successful. more recently treatment has tilted towards aggressive surgical interventions, which carry significant risk and are often not feasible. objectives: review our experience with three different patients with bcs and suggest a mechanistic based approach to treatment. the records of three patients with bcs were reviewed and their presentations, etiologies, treatment, and outcomes were reported. results: patient a was a -year-old female with paroxysmal nocturnal hemoglobinuria who presented with recurrent worsening abdominal pain over several months. narrowing of inferior vena cava (ivc) and hepatic veins was noted on imaging. liver transplant was not considered surgically feasible. she was treated with eculizumab, steroids, and anticoagulation with restoration of hepatic venous flow in weeks. patient b was a -year-old male with several weeks of right upper quadrant pain, fatigue, and pre-syncopal episodes, with a history of blunt abdominal trauma from football scrimmage weeks earlier. he was found to have near complete occlusion of the ivc and hepatic veins. liver transplant was not considered feasible. he was successfully treated with anticoagulation alone. patient c was a -yearold male with acute myeloid leukemia in induction cycle who developed severe pancytopenia; typhlitis was diagnosed and managed medically. days later he acutely decompensated, arrested, and was placed on extra corporeal membrane oxygenation, and imaging showed complete occlusion of the portal vein, hepatic veins, and ivc to the level of the atrium, with bilateral pulmonary emboli. emergency liver transplant or catheter based interventions was deemed not feasible. treatment with eculizumab was considered for presumed inflammation induced complement activation (c mg/dl [normal - ]; ch was u/ml [normal - ]) as a trigger for thrombosis, but the patient progressed quickly and died before it could be initiated. our experience with bcs shows that invasive interventional options and liver transplant may not be feasible in most patients for multiple reasons. rapid diagnosis and aggressive etiology-based medical management are paramount to successful treatment of this rare complication. eculizumab may be considered in treating bcs with complement activation not only due to innate disorders, but also secondary to acute inflammation when proper laboratory evidence is present. background: platelet aggregation studies are the gold standard for the diagnosis of platelet function defects during the evaluation of a patient with bleeding problems. the platelet aggregation test measures how well platelets clot in response to different concentrations of epinephrine, adenosine diphosphate (adp), collagen, arachidonic acid and ristocetin. because platelet function defects are often under-recognized and under-diagnosed in the pediatric patient, the true incidence is unknown. we report our experience in the diagnosis of platelet defects at our institution over a -year period in order to add some clarity to the limited pediatric data available. objectives: our primary objective is to document correlations/trends between less well-known platelet function abnormalities and clinically significant bleeding at our institution over a -year period. design/method: after appropriate irb approval obtained, we performed a retrospective chart review of all children who had platelet aggregation testing done from to . data collected included demographics (age, sex, race), personal and family history of bleeding, screening for coagulation defects and platelet aggregation test results. symptoms examined in our data were limited to epistaxis and heavy menstrual periods. for each of these symptoms, results were further analyzed to those with abnormal responses to adp and epinephrine. patients with existing bleeding diagnoses and those with incomplete medical records were excluded. we identified patients. of the patients with epistaxis, % had abnormal platelet aggregation testing while only % of those with heavy menstrual periods had abnormal results. within our population, abnormal platelet function assay (pfa- ) results or race did not appear to correlate with abnormal platelet aggregation testing. in the cases of epistaxis, sex was also noncontributory. our preliminary results suggest that platelet aggregation testing was more useful in predicting platelet defects in those with a clinical bleeding history of epistaxis as opposed to heavy menstrual periods. for other presenting symptoms, platelet aggregation testing did not offer diagnostic benefit. abnormal response to adp in the platelet aggregation test was the most common finding in our population; the clinical significance of which is not well understood. going forward, we plan to document whether abnormal results correlated significantly with the subsequent final diagnoses of our patients. background: decision making for severe hemophilia a in previously untreated patients (pups) has recently become a significant ethical debate. recombinant factor viii (rfviii) products previously were recommended to avoid transmission of blood borne pathogens associated with plasma-derived fviii (pdfviii) products. however, the increased incidence of fviii alloantibody inhibitors with rfviii products compared to pdfviii products has challenged this former standard of care. despite the support of the medical and scientific advisory council, recommendations considering pdfviii products for a pup remains controversial. design/method: we used a modified utilitarian approach involving clinical, public health, and research ethics. shared decision making permeates the framework to maximize understanding, minimize bias, respect informed consent or dissent, and provide care that aligns with patient and family values when medically and practically feasible. the framework has three tiers. first, it evaluates whether resources are scarce or abundant for equitable resource allocation. if fviii products are scarce, we s of s recommend developing a central supply for emergency use and then evaluating the needs of the severe hemophilia a patients. prioritization of who receives the factor products would be decided by a designated team based on the availability of the factor products and clinical scenarios, with no preference given to those on research trials. however, if resources are abundant, treatment for acute bleeding and standard of care prophylaxis measures, including primary prophylaxis, could continue. the second tier accounts for whether there is a new infectious epidemic or concern where a pathogen cannot be eliminated. if there is, healthcare and public health workers may limit the use of pdfviii products. if not, pdfviii and rfviii products are to be equally considered. the third tier evaluates whether the clinical scenario is emergent or not. if there is acute, emergent bleeding, the immediately available resource should be used, along with bypassing and/or adjuvant resources as needed until the bleeding has resolved or improved. to align with patient and family preferences, attempts to have both pdfviii and rfviii products available at similar costs in institutions would be ideal. this ethical framework endeavors to balance autonomy, beneficence, nonmaleficence and justice in helping guide discussions among providers, pups with severe hemophilia a, and their families. disclaimer: findings and conclusions are those of the author(s) and do not necessarily represent the official position of the centers for disease control and prevention, emory university, or children's healthcare of atlanta. background: von willebrand disease (vwd) is a common bleeding disorder which affects up to % of the population without gender predilection. bleeding associated with this condition results from a deficiency or abnormality in von willebrand factor interfering with formation of primary hemostasis. ehlers-danlos syndrome (eds) is a group of rare inherited connective tissue disorders which may have an associated bleeding manifestation without abnormalities in coagulation testing. bleeding symptoms reported in eds result from capillary and tissue fragility. joint hypermobility syndrome (jhs) is an inherited condition which is nearly indistinguishable from eds iii. reports of coinheritance of vwd and eds or jhs are infrequent. the objective of this retrospective study was to review patients with coexisting vwd and eds or jhs at the indiana hemophilia and thrombosis center in order to describe the type and severity of bleeding symptoms, physical examination findings, and pertinent laboratory data. design/method: the electronic medical record database of the indiana hemophilia and thrombosis center was queried for patients with a diagnosis of vwd and one of the following descriptors: hypermobility syndrome, hypermobility, hypermobile joints, or ehlers-danlos syndrome. the records of identified patients were reviewed for demographics, type and severity of bleeding symptoms, beighton scores (bs), vwd antigen, ristocetin cofactor, factor viii levels, vwd multimer pattern, vwd subtype, genetic testing for eds, and family history of eds. results: a total of patients with dual diagnoses of vwd and eds and patients with vwd and hypermobility were identified with this query. two patients had completed genetic testing for eds, and one had a col a gene mutation identified. significant bleeding symptoms in the vwd and eds group included hematuria and postoperative hemorrhage. two of these patients had delayed wound healing postoperatively. seven of the patients identified to have type i vwd and jhs had moderately severe and somewhat unusual bleeding episodes reported including hematuria, hematemesis, and hemoptysis; of these patients had significant perioperative bleeding. females composed % of the vwd and eds group and % of the vwd and jhs group. conclusion: coinheritance of vwd and eds is an uncommon phenomenon. patients with vwd and eds or jhs may have atypical and moderately severe bleeding, especially with procedural intervention. incorporation of bs into the assessment of patients with bleeding disorders is useful to identify potential inherited collagen disorders, as diagnosis of these conditions may impact clinical management. in the year-long phase ii study (ro fd ), / khe patients responded. patients were followed for years after study completion, collecting data on growth and development, complications of therapy, unexpected toxicities, and need for continuing sirolimus. objectives: after study therapy treatment of one year, objectives include: . assess long term toxicity over the - year period after study therapy completion . assess unexpected toxicity . assess overall condition of the patient . assess need for restart or continuation of sirolimus therapy design/method: prospective follow-up of patients with a diagnosis of khe from institutions. inclusion criteria: follow-up for - years post-study. results: follow-up included data at year (n = ) and - . year (n = ) time points. average age at the start of treatment was months. of patients were available for follow up. four patients are no longer on sirolimus: one patient completed study therapy and remains off treatment (ot) ( years), required years of treatment and is now . years ot and required an additional treatment course prior to successful discontinuation now and months ot. of the patients still on sirolimus, all restarted medication for symptoms of pain, swelling and/or edema interfering with quality of life and have made an average of . attempts to discontinue sirolimus. no patient had reoccurrence of kmp. all patients had improvement in clinical and radiologic appearance of khe but all have residual lesions noted on imaging and/or clinical exam. no unexpected toxicity, growth delay, developmental issues or other long term toxicity of sirolimus was noted. conclusion: this is the first prospective data on long-term follow up of khe patients treated with sirolimus. although numbers are small, sirolimus is well tolerated; however, over half the patients were still on medication at - year follow up. this stresses the need for continued long term follow up in these young patients and investigation of the mechanism of sirolimus effect. nationwide children's hospital, columbus, ohio, united states background: recent studies have identified that adult persons with hemophilia (pwh) have a higher prevalence of hypertension and renal disease than the general population. while hematuria is a known complication of hemophilia a and b (ha, hb), its long-term impact on pwh is not currently known. by annually screening our patients with urinalysis, our pediatric center identified that just under half of our patients demonstrated hematuria over a four-year period. motivated by a desire to identify early markers of hypertension and renal disease, we sought to determine if this finding is reflected in the pediatric hemophilia population as a whole. objectives: establish the population-wide prevalence of hematuria in pediatric pwh. design/method: we used the pediatric health information system (phis) database, which contains clinical and resource utilization data for inpatients from hospitals nationwide, to analyze the prevalence of hematuria, hypertension, renal disease and related diagnosis codes in pediatric pwh who were admitted from january to september . results: during the five-year period, , unique pediatric pwh accounted for , admissions. while the majority of admissions were for bleeding or infectious concerns, ( . %) patients had an affiliated admission code for hematuria. for admissions as a whole, the median age was years with % of those admitted being infants, % toddlers, % children, % adolescents, % older than . we identified % of admissions were for ha with the remaining % were for hb. there were ( %) admits in which a bypassing agent was administered. the median length of stay for persons with hematuria was days compared to days for nonhematuria/other bleeding. there were ( . %) admissions with hypertension reported; though, only patients received an antihypertensive medication during that admission. additionally, only ( . %) admissions reported a diagnosis code of renal disease. our study demonstrated that pediatric pwh are experiencing hematuria. in general, only patients with persistent hematuria require hospital admission so we suspect this data underrepresents the numbers of pwh experiencing hematuria that is managed in the outpatient setting. we also suspect that hypertension is grossly underreported and undertreated in pediatric pwh. additionally, there are a low number of patients experiencing renal disease requiring hospital admission among this cohort. given that there is little research into the long-term impact of hematuria in hemophilia, we feel these findings support the need for further vigilance of our pediatric pwh. background: gla and gsd can aggressively destroy bone, with significant impact on morbidity and mortality. the mtor inhibitor, sirolimus has been shown to be effective in the treatment of these diseases. based on the addition of mtor inhibition to bisphosphonate therapy in metastatic cancer therapy, regimens have been used for refractory or high risk gla and gsd but there is heterogeneity of diagnosis, and variability of drug regimens and assessment of effectiveness. objectives: . assess the variability of clinical features of gla and gsd . assess the heterogeneity of diagnosis . assess drug regimens and response assessment across multiple institutions design/method: we conducted a retrospective review from institutions of cases of gla and gsd treated with sirolimus and a bisphosphonate for at least months with assessment of clinical features, treatment protocols, response regimens and side effects. results: patients included gla (n = ) and gsd (n = ). the average age at diagnosis was years. clinical features included effusions: gla (n = ), soft tissue lymphatic malformations: gla (n = ), gsd (n = ), multiple splenic lesions: gla (n = ), and soft tissue swelling at the site of bony lesion: gsd (n = ). the presenting symptom in patients was pain with patients (gla) presenting with shortness of breath. fracture was noted in patients: gla ( ), gsd ( ). diagnostic and/or response imaging included mri, ct, bone scan, skeletal survey and dexa scan. treatment consisted of: initial sirolimus use with the addition of bisphosphonate secondary to worsening disease (n = ), initial therapy with other agents (interferon, chemotherapeutic agents, radiation) and change to sirolimus and bisphosphonate secondary to toxicity (n = ), sirolimus and bisphosphonates (n = ) and sirolimus, bisphosphonates and interferon (n = ). seventeen patients had stable disease and patients had improvement of disease. sirolimus protocol was standard; however, bisphosphonate protocol varied in dosing and frequency. side effects were tolerable and expected with no grade iii or iv toxicity. sirolimus and bisphosphonates are a safe and effective therapy for gsd and gla. a consistent medication regimen, redefined response and an improved radiologic classification will be important for the development of a prospective clinical trial. background: hemophilia a is a bleeding disorder from the deficiency of clotting factor viii. the most significant sequelae of hemophilia a is the tendency to develop hemarthrosis that incites joint destruction. the prevalence of overweight and obesity has been increasing in the general and hemophilia population and leads to several morbidities including arthropathy. this is a particular concern for hemophilia a as arthropathy is a consequence of joint bleeding. objectives: the purpose of this study was to detect the relation between body mass index (bmi) and joint health endpoints in a pediatric hemophilia population. design/method: participants in this study included patients from the hemostasis and thrombosis center at children's hospital los angeles. participants were pre-screened and approached for this study during routine follow-up appointments. patients aged - years old who have been diagnosed with hemophilia a, including mild, moderate, and severe, qualified for the study. informed consent was obtained from the patients or parents before enrollment. joint health was objectively measured by physical therapists from children's hospital los angeles using the hemophilia joint health score (hjhs). an hjhs total score is calculated by assessing: swelling, duration of swelling, muscle atrophy, crepitus on motion, flexion loss, extension loss, joint pain, and muscle strength in major joints. subjective data was also obtained by patients recording their annual bleed rate within the past year. of the patients, ( %) were normal weight, ( %) overweight, and ( %) obese. we used chi-square analysis to compare joint scores across bmi classifications (chi square = . , df = , p-value = . ). although, this did not approach statistical significance, the average hjhs score in patients who had a hjhs > shows an increasing trend among bmi classifications: . in normal bmi patients, . in overweight bmi patients, and . in obese bmi patients. the average number of annual bleeds in those with positive values show: in normal bmi patients, in overweight bmi patients, and in obese bmi patients. although a positive effect of adiposity was found in the joints of hemophilia a pediatric patients, the effect shows there was not enough evidence to conclude a difference. future studies are needed to address whether obesity has an effect on hemophilia and to determine whether overweight/obesity can lead to further complications in hemophilic joints. background: stagnant blood flow in slow-flow vascular malformations (vm), particularly in their venous components, can lead to localized intravascular coagulation (lic) that is characterized by elevated d-dimer levels, low fibrinogen and decreased platelet count this coagulation derangement can lead to localized thrombosis or bleeding which can result in pain, functional limitations, and possible progression to disseminated intravascular coagulopathy (dic). the treatment of vm and their associated coagulopathy has proven difficult. patients with complex vm are frequently managed with sirolimus, an mtor inhibitor, and have clinical benefits, including reduction of pain and improvement in functional impairment. it is possible that some of these improvements from sirolimus could be secondary to improvement in the coexisting lic. objectives: this study assessed the use of sirolimus to manage the coagulopathy seen in slow-flow vm. design/method: we reviewed charts of patients with vm who are followed in the vascular anomalies center at arkansas children's hospital and were started on sirolimus. efficacy was objectively assessed through improvement of ddimer, fibrinogen and platelet count. three sets of lab values (pre-sirolimus, - months post-sirolimus, and most recent) were obtained for each patient when available. we identified a total of patients who had been prescribed sirolimus. eighteen were excluded based on underlying condition other than slow-flow vascular malformation and for inadequate medical records. a total of patients ( combined vascular, venous) were included in the study. all had elevated d-dimer levels (mean . mcg/ml feu, median . mcg/ml feu, range ( . - . )) prior to treatment. two patients had an associated low fibrinogen (below mg/dl), indicating severe lic. with treatment, ( . %) patients showed an overall decrease in d-dimer levels with an average decrease of . mcg/ml feu between pre-and post-sirolimus labs, and an average decrease of . mcg/ml feu between pre-sirolimus and most recent values. the two patients with low fibrinogen prior to treatment showed a decrease in d-dimer levels (mean decrease of . mcg/ml feu) and an increase and normalization in fibrinogen (mean increase . mg/dl) after beginning sirolimus. no patient had thrombocytopenia. we report that treatment with sirolimus was effective in improving coagulopathy associated with slowflow vm as evidenced by decreased d-dimer levels and increased fibrinogen and/or platelets. long-term use of this medication in this population may decrease the bleeding and thrombotic complications that these patients experience, especially following invasive vascular procedures. background: safety and efficacy of bay - , a sitespecifically pegylated b-domain-deleted recombinant factor viii, in previously treated adolescents and adults aged - years with severe hemophilia a was demonstrated in the phase / protect viii study and ongoing extension. objectives: this subanalysis examines the efficacy and safety of bay - in adolescents in protect viii and the ongoing extension study (data cutoff, january ). design/method: in protect viii, patients (including adolescents) received bay - on demand or as prophylaxis for weeks. prophylaxis regimens for weeks - were twice-weekly ( - iu/kg), every- -days ( - iu/kg), or once-weekly ( iu/kg) infusions based on bleeding during a -week run-in period of iu/kg twice-weekly prophylaxis. patients continued their prophylaxis regimens in the extension or changed regimens at any time. results: twelve patients aged - years were included in the protect viii intent-to-treat population; s of s additional patient discontinued after dose (included in safety population). for patients receiving prophylaxis before study enrollment, median (range) number of total and joint bleeds in the months before study entry was . ( - ) and . ( - ), respectively. ten patients ( . %) had target joints at baseline (median [range], [ - ] per patient). during weeks - of protect viii for the entire time patients remained on their designated prophylaxis dosing frequency, the median (quartile [q] ; q ) annualized bleeding rate (abr) for patients receiving twice-weekly (n = ), every- -days (n = ), and once-weekly prophylaxis (n = ) was ( ; . ), . ( ; . ), and . ( ; . ), respectively (overall prophylaxis [n = ], . [ . ; . ]). two patients switched from once-weekly to twice-weekly (n = ) or every- -days prophylaxis (n = ), and number of bleeds decreased from to in one patient and to in the other. all patients from the main study continued in the extension; mean abr in the extension was . and varied by dosing regimen (twice weekly [n = ], . ; every days [n = ], . ; once weekly [n = ], . ). two patients changed from every- -days to once-weekly prophylaxis during extension (mean abr, . ). one patient had a nonneutralizing antibody to bay - at baseline; end-of-study titers were negative. no patient developed anti-peg antibodies or factor viii inhibitors or experienced a serious adverse event related to bay - during the main study or extension. in previously treated adolescents with severe hemophilia a, bay - prophylaxis was effective in prevention of bleeds, with less bleeding overall versus prestudy, and was generally well tolerated. funded by bayer. cincinnati children's hospital medical center, cincinnati, ohio, united states background: vascular malformations (vms) consist of a heterogeneous group of congenital disorders characterized by the abnormal development of blood and/or lymphatic vessels, which cause a broad spectrum of clinical manifestations. although considered benign, vms are frequently associated with cutaneous complications that can cause significant morbidity such as nodular overgrowth, skin thickening, pruritus, oozing or bleeding of lymphatic blebs and secondary infection. oral sirolimus has shown to be effective in the treatment of complicated vascular malformations but has known side effects and need for frequent laboratory monitoring. currently, there are limited studies on the use of topical sirolimus for the treatment of cutaneous manifestations of vascular malformations. objectives: to evaluate the efficacy and safety of topical sirolimus in vms with cutaneous complications and propose indications for use. design/method: this is a retrospective review of medical records of patients with vascular malformations treated with topical sirolimus from january to december . response was determined by subjective and objective improvement. results: twenty-four patients, ( %) females and ( %) males, with vascular malformations and cutaneous manifestations were treated with topical sirolimus. age ranged from - years. indications for treatment were: blebs ( %, n = ) causing either leaking, bleeding, pain, pruritus, swelling or recurrent infection; nodular overgrowth % (n = ); pyogenic granuloma % (n = ); bleeding % (n = ) and cosmetic % (n = ). treatment course ranged from - months. no major side effects were reported. one patient reported burning and itching sensation. regarding clinical response: % (n = ) patients had improvement in cutaneous lesions; % (n = ) had a stable lesions; and % (n = ) stopped treatment due to side effects. for prior/concomitant treatment: % (n = ) had prior surgery, laser or sclerotherapy; % (n = ) had concomitant oral sirolimus. of the patients not receiving concomitant systemic sirolimus, only % (n = / ) had been on oral sirolimus. of these patients, % (n = / ) had a very good response to topical treatment. : topical sirolimus appears to be beneficial and well-tolerated with a minimal side effect profile for the treatment of cutaneous manifestations of vascular malformations as a single agent or as adjuvant therapy with systemic sirolimus when symptoms are not adequately controlled. further studies are needed to prospectively analyze efficacy and safety of topical sirolimus in this patient population. objectives: to evaluate the safety and efficacy of long-term romiplostim in children with itp. design/method: all patients received weekly sc romiplostim from - g/kg to target platelet counts of - × ( )/l. median (min-max) treatment for the patients was ( - ) weeks for a total of patient-years, or . years per patient. at baseline, median (min-max) age was ( - ) years; % were female; . % had prior splenectomy. median (min-max) average weekly dose was . ( . - . ) g/kg, including escalation to a stable dose; patients started on g/kg. reasons for discontinuing romiplostim (n = , %) included consent withdrawn (n = ), required other therapy (n = ), and ae (n = ) (asthenia, headache, dehydration, and vomiting in one patient and anxiety in the other; none treatment related). fifty four serious aes occurred in patients but were treatment related in one (concurrent grade thrombocytopenia, grade epistaxis, and grade anemia). anti-romiplostim neutralizing antibodies were detected in one patient who discontinued to receive other therapy; antibodies were absent on retesting. from week on, median platelet counts remained > × ( )/l; median platelet counts were > × ( )/l from weeks - . nearly all ( %, / ) patients had ≥ platelet response (platelet counts ≥ × ( )/l, excluding ≤ weeks after rescue medication). most ( %, / ) patients had a platelet response ≥ % of the time and % ( / ) did ≥ % of the time. sixty ( %) patients (or caregivers) self-administered romiplostim. fifteen ( %) patients had treatment-free periods of platelet counts ≥ × ( )/l for ≥ weeks (ie, remission); these patients ( girls, boys) had had itp for a median (min-max) of . ( . - ) years, none had prior splenectomy, and had received romiplostim for . ( . - ) years. all had platelet counts > × ( )/l for ≥ months and / for ≥ months; the median (min-max) duration of being ≥ × ( )/l was ( - ) weeks. of baseline characteristics such as sex, platelet counts, itp duration, and number of past itp treatments ( , , , > ), only age < years was predictive of developing treatment-free periods ≥ weeks (p = . ). in this seven-year open-label extension, > % of children with itp achieved a platelet response and romiplostim was well tolerated. importantly, % of patients were able to discontinue all itp medications for ≥ months. funded by amgen inc. background: sirolimus is an immunosuppressive drug that is widely used in solid organ and bone marrow transplantation, and more recently for the treatment of vascular and lymphatic anomalies. sirolimus has been associated with decreased immunity in the transplant setting in patients that have received other immunosuppressive drugs or were immunosuppressed from previous chemotherapy. the effects of sirolimus on the immune system in chemotherapy naïve children who have not received other immunosuppressive agents are not well understood, and there is variability in the approach to fever and pcp prophylaxis. to understand the effects of sirolimus on the immune system of patients with non-complicated vascular or lymphatic anomalies by evaluating anc, alc prior to and after sirolimus therapy. design/method: multi-institutional retrospective review was done to include patients with non-complicated vascular or lymphatic anomalies. those with effusions/ascites, multiorgan involvement, or history of vascular-anomaly-related infections prior to treatment were excluded. results: twenty patients with kaposiform hemangioendothelioma (n = ), generalized lymphatic anomaly (n = ), cloves syndrome ( ), and simple vascular malformation (n = ) were included. age at initiation of sirolimus treatment ranged from . - years. male to female ratio was : . sirolimus was initiated due to extensive disease, lack of response to steroids or bisphosphonates, pain, dment, lymphatic drainage, and prevention of ongoing overgrowth. prior to the start of sirolimus (sir- ) the mean anc was and alc was . the target level of sirolimus varied by indication and patient, and ranged from to . after the st steady state level, month after sirolimus (sir- ) the mean anc decreased to and alc was . at months after sirolimus (sir- ) the mean anc was and alc was . the first sirolimus levels (sir- ) mean was . ; and sir- level was . . nine patients were placed on pcp prophylaxis at the start of sirolimus. none of these patients had an infectious complication while on sirolimus at a median f/u of months. one patient had mild neutropenia (anc > ) which normalized after discontinuation of pjp prophylaxis. conclusion: in this small cohort of patients we found that the anc and alc level in patients with non-complicated vascular or lymphatic anomalies at sir- was not different from the sir- or sir- . prospective studies that specifically track anc, alc, igg, and lymphocyte function should be conducted to better understand the effects of sirolimus in the immune system. this data will allow for uniform recommendations regarding prophylaxis and management of febrile episodes. background: acute infections and the associated systemic inflammation can increase the risk of venous thromboembolism (vte) and in certain well-defined clinical scenarios may be the primary trigger of vte in pediatric patients. pediatric data on vte in the setting of acute infection are sparse. objectives: to describe characteristics and outcomes of vte in pediatric patients with acute infections. we conducted a retrospective chart review of all pediatric patients with objectively confirmed vte treated at our institution since and identified all patients in whom an acute infection was identified as a vte trigger. patients were managed according to standardized institutional protocols based on published guidelines. relevant demographic, clinical and laboratory data were collected and summarized using descriptive statistics. since , acute infection was identified as a trigger in of vtes ( %) diagnosed at our center. the median age at time of vte diagnosis in this group was . years (interquartile range . - ). males were more commonly affected than females, representing % of cases. neonatal vte events accounted for % of cases. sepsis was the most common acute infection to be identified as a vte trigger [ / cases ( %)]. most vte events ( %) associated with acute infections were considered hospital-associated vtes. at time of vte diagnosis, % of patients were critically ill. extensive vte (defined as completely occlusive thrombosis involving > venous segment) occurred in % of patients. acute infection was deemed to be the primary trigger for vte in / patients ( %). infection-associated vtes in this cohort included cerebral sinus venous thrombosis due to sinus or cns infection ( patients, %), septic throm-bophlebitis ( patients, %), lemierre's or lemierre's-like syndrome ( patients, %) and osteomyelitis-associated deep vein thrombosis ( patients, %). systemic anticoagulation was prescribed in / patients ( %). anticoagulationrelated major bleeding occurred in / patients ( %). vte complications included vte recurrence ( patients, %), vte progression ( patient), acute pulmonary embolism ( patients) and arterial ischemic stroke ( patients). our study indicates that acute infection is a common risk factor for pediatric vte, especially in critically ill children, and can be the primary trigger in a significant proportion of vte cases associated with acute infections. anticoagulation appeared to be overall safe in this population and was associated with low rates of serious vte-related acute complications. however, our study also suggests that this population may be at increased risk for vte recurrence and anticoagulation-related major bleeding. background: epithelioid hemangiomas (eh) are rare benign vascular tumors that occur in soft tissues and bone and present between the third and sixth decades of life. a subset ( %) of eh harbor fos rearrangement. eh has been described in children, but little is known about the long-term outcomes of pediatric eh. the main objective is to obtain data to be used for improved understanding of this rare disease in order to provide standardization of care and development of future research studies. board-approved retrospective review of clinical, pathologic, and radiographic characteristics, and treatment outcomes in patients diagnosed with eh between and . results: eight patients were male; mean age at diagnosis was . years (range: - ). lesions involved the lower extremities (n = ), cranium (n = ), pelvis (n = ), and spine (n = ). multifocal disease was identified in five patients. the most common presentations involved significant localized pain and neurologic symptoms: headache, cranial nerve injury, loss of consciousness. radiographic studies identified variable features, such as multifocal lytic bony lesions with sclerotic margins, enhancing soft tissue component, and surrounding inflammatory edema. histologically, all specimens were composed of vascular channels lined by epithelioid endothelial cells without significant cytologic atypia; solid cellular areas (n = ). endothelial cells were positive for cd and egr, and negative for camta . fos rearrangement was assessed in only one specimen and detected. mean follow-up time was days (range: - ). patients were treated with surgical resection, intravascular embolization, bisphosphonates, propranolol, interferon, and sirolimus. one patient treated with interferon and one with sirolimus exhibited partial response for mean follow-up of . days. although eh is a benign neoplasm, it is difficult to manage without standard protocols and portends considerable morbidity. our findings suggest medical management, particularly sirolimus, may benefit these patients; however, long-term follow-up is needed in treated children. novel fos inhibitors are in development and may benefit patients with fos rearrangement. penn state health children's hospital, hershey, pennsylvania, united states background: central venous catheters (cvc) are often required in critical care settings in order to provide a secure point of access for life sustaining care. clinical studies identify cvc presence as the single most important risk factor for deep vein thrombosis (dvt) in children. venous thromboembolic event (vte) incidence rates in critically ill children with a cvc range from . - % and . - . per catheter days depending on the population studied. per institutional protocol, the penn state health children's hospital picu (hershey, pa) utilizes a low dose continuous infusion of unfractionated heparin (ldufh) at units/kg/hr as prophylaxis against cvc-related vte and to maintain line patency. the efficacy of this approach has never been evaluated. to determine if ldufh for prophylaxis results in lower incidence of cvc-related vte, catheter dysfunction and central line associated blood stream infection (clabsi) without increasing morbidities. to determine if the incidence of catheter related vte is lower than historical published data, a retrospective chart review was conducted utilizing the institutional electronic medical record for all patients in , aged - . years, who had a cvc during a picu admission. secondary objectives such as the incidence of catheter dysfunction, clabsi, and any associated bleeding complications are also being analyzed. results: interim data analysis revealed cvcs ( nontunneled cvc, totally implantable devices, tunneled lines, peripherally inserted central catheters [picc] ) in total patients with a median age of . years. overall vte incidence was . % ( / ) with vtes associated with non-tunneled cvc and with piccs. sixty one percent of non-tunneled cvcs received ldufh and % ( / ) of the patients with vtes associated with non-tunneled cvcs did receive ldufh prophylaxis. vte incidence rate of nontunneled cvcs with ldufh was . % ( / ) and . per picu catheter days. the only other vte events identified within our study cohort were in the picc group where two patients experienced vte, one of which was receiving ldufh. clabsi incidence was . % ( non-tunneled cvc, tunnel cvc, picc). no major bleeding complications were associated with ldufh. preliminary data demonstrates ldufh is efficacious in preventing cvc-related vte in comparison to published reports. further analysis will compare another similar sized and acuity level picu which does not practice the same method. background: fibroadipose vascular anomaly (fava) is a rare, challenging disorder associated with pik ca mutations. fava often causes painful replacement of muscle and soft tissues with fibrotic and adipose tissue and is associated with ectatic draining veins. treatments for focal lesions are surgical excision, cryoablation or sclerotherapy and the role of medical therapy is unclear. some fava lesions are too extensive or directly involve neurovascular structure, resulting in refractory pain. objectives: to retrospectively evaluate the efficacy of sirolimus in patient with residual symptoms after procedural therapies for fava design/method: retrospective review of individual cases from institutions of fava refractory to other therapies treated with sirolimus for at least months. cases were s of s identified by polling member of the aspho vascular anomalies special interest group. results: all seven patients report improvement on sirolimus therapy. all patients had received prior procedures, including sclerotherapy ( patients), cryoablation ( patients) and/or resection ( patients). mean age at sirolimus initiation was y (range - y). mean length of therapy is . months (range - months). six patients were treated with bid dosing and one adult received daily dosing. goals of sirolimus were improvement in pain or musculoskeletal dysfunction. pain and function improved in all patients, including discontinuation of narcotic use and resumption of participation in sports. time to symptom improvement ranged from - weeks. in four patients for whom dose was lowered, pain recurred in all four and responded to restarting or increasing sirolimus dose. while all patients do not have pre-and postsirolimus imaging, decrease in fava lesion size is seen in cases with available imaging. sirolimus side effects are similar to prior reports, most commonly mouth sores, elevated lipids and acne. we report the first known data supporting a role of sirolimus in refractory fava cases. sirolimus is welltolerated and initial improvement is rapid, within weeks of initiation. whether sirolimus has a role in upfront therapy to reduce lesion size prior to procedures deserves further study. objectives: to assess platelet responses in children with itp receiving romiplostim. design/method: eligible children had itp for ≥ months, ≥ prior therapy, and screening platelet counts ≤ × ( )/l or uncontrolled bleeding. weekly dosing was from - g/kg to target platelet counts of - × ( )/l. bone marrow biopsies were evaluated in europe at baseline and after or years (cohorts and ). as of mar , patients received ≥ dose. at baseline, median (min-max) age was ( - ) years, itp duration was . ( . - . ) years, and platelet count was ( - ) × ( )/l; patients ( %) had had prior splenectomy. the median (q , q ) % time with a platelet response (platelet count ≥ × ( )/l, no rescue medications past weeks) in months - was % ( %, %) (primary endpoint). over the course of the study, % ( / ) of patients had a platelet response. four patients maintained platelet counts ≥ × ( )/l with no itp medications for ≥ weeks. median (min-max) treatment duration was ( - ) weeks for patient-years in total. median (min-max) average weekly romiplostim dose over the course of the study was . ( . - . ) g/kg; the median dose was g/kg at year (n = ) and g/kg at years (n = ). most ( %) patients initiated self-administration. sixty-four patients ( %) discontinued treatment, most frequently for lack of efficacy (n = ), patient request (n = ), and adverse event (ae) (n = ). fortyone ( %) patients had serious aes (saes) including epistaxis ( %) and decreased platelet count ( %). five patients had treatment-related saes: headaches, abdominal pain, and each of presyncope and neutralizing antibodies (ab). there were cases of neutralizing ab to romiplostim (of patients tested), but none to tpo; / had continued elevated platelet counts and in / cases ab were not found on retesting. for cohort , of patients with baseline bone marrow biopsies, had evaluable on-study biopsies scheduled for year; patient had an increase from grade to . there were no findings of collagen or abnormalities. in this interim datacut of a romiplostim openlabel study in children with itp, % of children had a platelet response. overall, the median dose was . g/kg; the median romiplostim dose over time reached g/kg. no new safety signals were observed over patient-years. funded by amgen inc. background: hepatic hemangiomas are benign vascular tumors without a medical home, managed by multiple specialties. the diagnosis has been assigned historically to various vascular lesions affecting the liver with completely different clinical presentations, resulting in difficult standardized management. objectives: the consensus steering committee identified an acute need of clear definitions and evaluation guidelines using the updated international society for the study of vascular anomalies (issva) classification. the goal was to formulate recommendations that will be adopted by all specialties involved in the care of children with hepatic hemangiomas. design/method: we used a rigorous, transparent consensus protocol, with input from multiple pediatric experts in vascular anomalies from hematology-oncology, surgery, pathology, radiology and gastroenterology. in the first section, we precisely define the subtypes of hepatic hemangiomas seen in children (congenital and infantile) using clinical course, histology and radiologic characteristics. inclusion and exclusion limits to the diagnosis are noted. the following two sections describe these subtypes in further detail, including complications to be considered during monitoring and respectively recommended screening evaluations. conclusion: while institutional variations may exist for specific clinical details, a clear understanding of the diagnosis of hepatic hemangiomas affecting the pediatric population and the possible complications that require screening during the monitoring period should be standard. as patients with hepatic hemangiomas are managed by different medical and surgical specialties, a multidisciplinary consensus based on current literature, on the data extracted from the liver hemangioma registry and on expert opinion was required and was accomplished by this manuscript. objectives: to investigate the association between routine prophylaxis with bay - and bleeding outcomes after adjusting for key patient and pharmacokinetic (pk) characteristics. design/method: the leopold kids study evaluated safety and efficacy of bay - prophylaxis in previously treated boys aged ≤ years with severe hemophilia a. patients received bay - - iu/kg x/wk (n = ) or > x/wk (n = ) and were followed up for - months. prophylaxis dose and frequency were assigned by investigators. pk parameters, including area under the curve (auc), half-life, and clearance, were derived from a population pk model and reflect predicted pk values with a -iu/kg dose. patient characteristics were compared between the x/wk and > x/wk groups using wilcoxon rank sum or chi-square tests. negative binomial regression was used to model the association between prophylaxis frequency and annualized bleeding rate (abr) for total bleeds, first without adjustment and then adjusting for age, pk parameters, and bleed history. results: mean ± sd age for patients in this analysis was . ± . years. patients receiving prophylaxis x/wk had more bleeding episodes in the months before study entry (mean ± sd, . ± . [median, . ] for x/wk vs . ± . [ . ] for > x/wk; p = . ) and were more likely to have been treated on demand ( % vs %; p = . ). pk parameters were similar between the x/wk and > x/wk groups. without adjustments, abr during the study was % higher in the x/wk group compared with the > x/wk group (rate ratio [rr], . ; % ci, . - . ; p = . ). abr was % lower in the x/wk group (rr, . ; % ci, . - . ; p = . ) after adjusting for age, auc, and number of bleeds in the prior months. conclusion: abr was numerically lower but not significantly different between the x/wk and > x/wk groups after adjusting for age and pk parameters. these findings suggest that even among patient groups that are homogeneous with respect to age, pk, and bleed history, further individualization of bay - prophylaxis based on other characteristics may help reduce bleeding episodes even at a lower treatment frequency. larger real-world studies are needed to verify these findings. funded by bayer. stanford, palo alto, california, united states s of s background: vascular malformations may be of lymphatic, arterial, venous or capillary endothelial origin. they may be simple or complex, with complex malformations being a combination soft tissue and skeletal overgrowth. although likely present at birth, these malformations often become symptomatic with puberty or infection, and range from little or no clinical impact to life threatening symptoms. in malformations primarily of venous origin, pain may be significant and hypothesized to be caused by phlebolith development (intra-malformation thrombi), inflammation, consumptive coagulopathy, vascular engorgement, and endothelial proliferation. anti-angiogenic and anti-platelet therapies have been reported to relieve pain. however, the use of anticoagulation for pain is not well described. objectives: to report clinical features and outcomes of patients with vascular malformations of venous origin treated with anticoagulation for pain. we performed a retrospective review of patients with vascular malformations followed by the hematology service between january and december who were treated for pain with anticoagulation. pain relief was determined both by wong-baker pain scales and patient report. clinical data were extracted from electronic medical records. we identified five patients with venous malformations (vm) who had received anticoagulation for pain. four patients were female and median age was years old (range to years old) at time of initiation of anticoagulation. all five patients had vm of the extremity, two with vm of the lower extremity, and three patients had vm of the upper extremity. two patients had concomitant coagulopathy and demonstrated decreased d-dimer after initiation of anticoagulation. four patients received enoxaparin, and one adult patient received rivaroxaban. all patients reported improvement in pain after administration of anticoagulation. one patient exhibited mild epistaxis and bruising at the injection site. there was no significant bleeding or other complications. pain is a significant complication in patients with venous malformations. our case series suggests that anticoagulation is a safe and effective therapy for pain relief in this population. further investigation is indicated to compare the effect of anticoagulation to other therapeutic interventions such sclerotherapy, surgery, and sirolimus in the treatment of pain associated with venous malformation. maria ahmad-nabi, christine knoll, sanjay shah, lucia mirea phoenix children's hospital, phoenix, arizona, united states background: estimates of the incidence of dvt in patients with osteomyelitis range widely from %- %, however risk factors and outcomes of dvt in this cohort have not been thoroughly established. objectives: this study aims to estimate the incidence of dvt in patients with osteomyelitis, and to assess risk factors and outcomes of dvt in this cohort. design/method: after irb approval, a retrospective chart review was conducted for patients aged - years seen at phoenix children's hospital between - with icd / codes for osteomyelitis. exclusion criteria included chronic recurrent multifocal osteomyelitis, and chronic dvt. demographics, clinical factors and outcomes were compared between osteomyelitis patients with and without dvt using the fisher-exact and wilcoxon-rank sum tests, as appropriate for the data distribution. results: a total of study subjects with osteomyelitis had a mean (standard deviation) age of . ( . ) years. dvt was present in ( % of ) patients, and ( %), ( %) and ( %) patients received anticoagulation for < , - and ≥ weeks, respectively. patients with vs without dvt were more likely to be male ( % vs %; p-value = . ), and had significantly higher rates of bacteremia ( % vs %; p-value = . ). rates of central lines were comparable between dvt and non-dvt patients ( % vs %; p-value = . ); however patients with dvt vs without dvt had significantly longer mean length of stay ( vs days; p-value < . ) and higher rates of icu admission ( % vs %; p-value < . ). the incidence of dvt among osteomyelitis pediatric patients was estimated at %, with risk increased by male sex and bacteremia. patients with dvt had significantly higher rates of icu admission and longer length of hospital stay. many of these patients had standard practice management of their dvt with - weeks of anticoagulation. our data highlights the need for recognition of high risk patients, and the need for future efforts targeting dvt prophylaxis. baylor college of medicine, houston, texas, united states background: lymphatic malformations (lm) frequently occur in the head and neck and can often be disfiguring and even life-threatening. management options include observation, surgery, sclerotherapy, and sirolimus. the optimal sequence of therapeutic interventions has not been determined due to the lack of comparative clinical trials or established guidelines. thus, prenatal planning with a multidisciplinary team is beneficial. we present a case series of ten children with head and neck lms evaluated in at our multidisciplinary vascular anomalies center. a chart review was performed to assess treatment modalities and recent trends. results: seven of patients ( %) with head and neck lms were diagnosed prenatally. six patients required an ex utero intrapartum treatment procedure. all patients were started on sirolimus at a median age of . months (range days - years). four patients most recently started on sirolimus were less than months of age at the time of initiation. six patients underwent partial excision of lm during the first year of life; none of whom received sirolimus prior to surgery. sirolimus was discontinued in one patient given chronic clostridium difficile infections, and non-compliance in another patient. five patients received sclerotherapy. tracheostomy was necessary in six patients; one patient was de-cannulated after months on sirolimus. all patients have had radiographic and clinical improvement of lm with varying treatment modalities. current clinical observations show improved response with sirolimus and demonstrate tolerability of sirolimus at a young age. conclusion: treatment of pediatric head and neck lms is challenging and a multidisciplinary approach is necessary. as the majority of patients are diagnosed prenatally, prenatal planning and discussion of potential use of sirolimus is beneficial. availability of vascular anomalies experts in the prenatal/neonatal period offers the best management results, and early initiation of sirolimus should be considered for complex lesions. long-term follow up is warranted to investigate the efficacy and timing of treatment options. yale school of medicine, new haven, connecticut, united states background: to mitigate transfusion of pathogencontaminated platelets, amotosalen, a synthetic psoralen compound, is added to sdp components. exposure to uv-a light activates amotosalen and crosslinks dna/rna base pairs, preventing replication of a broad spectrum of viral, bacterial, and other pathogens that may contaminate platelets. pr-sdps were fda approved for clinical use with no age restrictions in . we initiated use of pr-sdps in november of for all patients. we retrospectively analyzed usage of pr-sdp vs conventional (non-pr) platelets (cp) in neonatal and pediatric patients with thrombocytopenia to compare hemostatic efficacy and the incidence of transfusion reactions (tr) for these products, after one year of a dual platelet inventory. design/method: since pr-sdp were fda-licensed, no irb approval was required; pr-sdp and cp were both considered standard of care. we evaluated transfusions for all pediatric patients age - years who received any platelet transfusion between november and november . we determined the volume (mean ml ± sd) of each type of platelet component transfused, the number of platelet transfusion episodes, and reported trs based on cdc hemovigilance guidelines. a subgroup analysis was performed for thrombocytopenic neonates ( - months). results: patients - years who received only cps (n = ) received a total of , ml of platelets ( ± ml/patient) over transfusions ( . ± . episodes/patient). for comparison, in patients who received only pr-sdp, a total of , ml of platelets ( ± ml/patient, p = . ) were infused over transfusions ( . ± . episodes/patient, p = . ). for neonates ( - months, n = ) who received only cps, , ml of cps ( ± ml/ patient) were transfused over episodes ( . ± . episodes/patient). for comparison, those who received only pr-sdp (n = ), received , ml of pr-sdp ( ± ml/patient, p = . ), transfused over episodes ( . ± . episodes/patient, p = . ). for all recipients - years (n = ), including additional patients who received both cp and pr-sdp, there were three reported allergic trs over transfusion episodes, while no allergic reactions were reported with pr-sdp transfusions. one febrile tr was reported to cp transfusion, while three were reported for pr-sdp. in conclusion, pr-sdps, in our pediatric population age - years, were comparable to cp products in regards to volume and episodes of platelet transfusions, and incidence/type of transfusion reactions. pr-sdp were safe and effective for use in this pediatric patient population. background: vascular anomalies are classified as either vascular tumors or vascular malformations. fibro-adipose vascular anomaly (fava) is a newly described entity which presents with distinct clinical, radiographic and histopathologic findings. we present a case in which the diagnosis of fava was complicated by a persistent low platelet count secondary to immune thrombocytopenia (itp). to describe a challenging diagnosis of a novel vascular anomaly (fava) complicated by severe thrombocytopenia. a year old male presented to hospital with bruising and left thigh pain related to a remote sports injury. blood work revealed a platelet count of × /l, but with an otherwise normal complete blood count. the following were also normal: aptt and fibrinogen; d dimer levels were slightly increased. he was treated with one dose of ivig ( . mg/kg) for presumed itp and responded well with his platelet count increasing to × /l. he returned to hospital weeks later with recurrent thrombocytopenia and worsening leg pain. an ultrasound of the left thigh revealed a . cm x . cm x . cm lesion within the vastus medialis. the diagnosis of an intramuscular hematoma secondary to persistent thrombocytopenia was made. the patient presented with multiple episodes of thrombocytopenia over the next several months. his itp did not respond to oral prednisone ( mg/day for days). he continued to have short-lived responses to ivig requiring infusions every other week as his platelet count would fall below × /l. his leg pain progressed, restricting him to a wheelchair. further imaging by mri brought into question the diagnosis of a hematoma and a biopsy of the thigh lesion was performed. the results were consistent with a diagnosis of fava; this was subsequently excised. conclusion: this is a unique case where a vascular anomaly was misdiagnosed as a hematoma due to a patient's persistent thrombocytopenia and history of an injury. fava is a newer entity which, unlike other vascular anomalies, has not been linked to thrombocytopenia or a localized consumptive coagulopathy. after excision of the fava, the patient's chronic pain, and mobility resolved, though his itp persisted. objectives: this preliminary, exploratory analysis of realworld administrative data was conducted to determine units dispensed and factor replacement product-related direct expenditures associated with a currently marketed shl or ehl rfix product. design/method: de-identified claims data from the commercially available truven health marketscan® research u.s. claims database were used to identify direct expenditures and number of international units (ius) dispensed for all patients aged - years with a diagnosis code of icd- . /icd- d who used nonacog alfa or eftrenonacog alfa during the study period (june , to july , ). reference weight measurements from the centers for disease control and prevention national center for health statistics' (cdc nchs) anthropometric data were used to estimate product dispensation on an iu per kg basis. the nonacog alfa and eftrenonacog groups comprised and patients, respectively. the median [iqr] age in the two groups was . [ . ] and . [ . ] years, respectively. while of the patients in the eftrenonacog alfa group had > calendar quarter of available data, only of the patients in the nonacog alfa group had > available quarter. the median rfix product dispensation per quarter was , ius (iqr, , ius) in the nonacog alfa group and , ius (iqr, , ius) in the eftrenonacog alfa group. incorporating attributed weight values, the median rfix product iu dispensation per kg per week was . iu/kg/wk (iqr, . iu/kg/wk- . iu/kg/wk) in the nonacog alfa group, and . iu/kg/wk (iqr, . - . iu/kg/wk) in the eftrenonacog alfa group. applying wac prices (eftrenonacog alfa = $ . /iu; nonacog alfa = $ . /iu), the calculated estimates of $/kg/week were $ and $ in the nonacog alfa and eftranonacog alfa groups, respectively. conclusion: preliminary real-world data derived from a large u.s. claims database revealed differences in product dispensation and factor product-related expenditures among pediatric patients with any severity of hemophilia b to whom an shl or ehl rfix product was prescribed. refinements of these data, potentially to exclude instances of sporadic usage, may shed light on real-world dispensation of rfix products among pediatric hemophilia b patients. background: vascular malformations can be classified as simple (including capillary, venous, lymphatic, arteriovenous), combined, malformations of major named vessels or associated with other anomalies. multiple modalities including laser treatments, sclerotherapy, embolization, surgery and pharmacological intervention (with mtor inhibitors like sirolimus) have been used for treatment of vascular malformations. these interventions have been used alone or in combination with varied outcomes. we present our institution's experience with a multimodal approach to simple and combined vascular malformations. design/method: we performed a retrospective chart review of patients with vascular malformations who were referred to our center for an interventional radiology evaluation from june -july . we included patients (age at presentation: months - years), referred initially for interventional radiology procedures (irp) for vascular malformations. all patients had symptoms of pain and/or swelling/deformity. diagnosis of was based on vascular imaging (doppler ultrasound, mri/a/v). nine patients had venous malformations (vm), five had macrocystic lymphatic malformations (lm), six had lymphatic-venous malformations (lvm), and two arteriovenous malformations (avm). patients initially underwent interventional radiology procedures. all the vm patients responded to sclerotherapy alone. three patients with lm responded to sclerotherapy alone, remainder required surgical intervention. one avm patient responded well to embolization, the other needed surgical resection after embolization. four lvm patients underwent irp with minimal improvement in symptoms ( - procedures attempted), surgical resection was attempted in patients with poor response and patients were started on sirolimus ( . mg/m /dose twice a day). all lvm patients started on sirolimus have responded well (decreased pain and swelling); time to initial symptom response ranged from weeks - month from starting medication. in this case series, patients with simple vm responded well to sclerotherapy alone, avm and lm patients needed irp and/or surgery for complete response. complex lvm did not respond well to surgery or irp; . % had improvement in clinical symptoms with addition of sirolimus to the treatment regimen. response to various modalities of treatment varied based on the type of vascular malformation. a multidisciplinary approach to management of vascular malformations is essential to provide multimodal therapeutic options for rapid symptom relief and improve the quality of life of these fragile patients, especially those with complex malformations. background: von willebrand disease (vwd) is the most common bleeding disorder in humans, affecting ∼ % of the united states' population. desmopressin (ddavp) is a longacting vasopressin analog that induces vasoconstriction and release of vwf. ddavp is used in patients with vwd and as a surgical prophylaxis, but carries anti-diuretic properties. to avoid electrolyte imbalance and hyponatremia, fluid restrictions are recommended in the hours post-ddavp administration. objectives: this study sought to examine perioperative practices and outcomes following ddavp administration and a fluid restriction protocol in a population of pediatric patients with von willebrand disease. design/method: a retrospective chart review was conducted for patients with von willebrand disease who underwent surgical procedures at children's hospital of pittsburgh of upmc between january , and december , . patient age, sex, weight, diagnosis, surgical procedure, total fluids administered, and post-operative sodium level were recorded. the primary outcomes noted were the proportion of patients exceeding % of the recommended fluid consumption for the -and -hour periods post-ddavp s of s administration, as defined by local guidelines. secondary outcomes were the presence of any bleeding requiring an er visit or readmission or hyponatremic seizures within hours of ddavp administration. results: data was compiled for patients ( females, males). the mean age was . years (sd . years), median age was years (range to years). procedures included dental ( ), otolaryngology ( ), orthopedics ( ), gastrointestinal ( ), plastics ( ), neurosurgery ( ), ophthalmology ( ), dermatology ( ), general surgery ( ) and gynecology ( ). % of patients exceeded % of the fluid volume recommended for the first -hour period post-ddavp administration while still in the surgical setting. no patients exceeded % of the fluid volume recommended for the total -hour period post-ddavp administration. post-operative sodium levels were obtained in only of patients. no patients returned to the er or were admitted for bleeding in the hours post-ddavp administration. no patients returned to the er or were admitted for hyponatremia or seizures in the hours post-ddavp administration. maintenance of a fluid restriction protocol effectively deterred negative outcomes in this cohort. however, a significant fluid volume was administered in nearly a third of patients despite the restrictions. given the risk of hyponatremia, and limited compliance with fluid restrictions, postoperative sodium levels should be recorded in following ddavp administration to assess the possibility of a hyponatremia and to reinforce the importance of fluid restrictions and their communication. results: a male fetus required in utero insertion of a pleuroamniotic shunt for bilateral pleural effusions diagnosed antenatally by ultrasound. shortly after delivery at term, he developed respiratory distress and was found to have reaccumulation of the pleural effusions. blood work on day of life showed a platelet count of , / l, which then decreased precipitously. he demonstrated schistocytes on blood-smear, signs of consumptive coagulopathy with hypofibrinogenemia and high d-dimers, and compensatory reticulocytosis. he required multiple transfusions and admissions to the intensive care unit for respiratory support. investigations ruled out congenital ttp, neonatal alloimmune thrombocytopenia, and noonan syndrome. given high clinical suspicion for an underlying vascular lesion causing kmp, a full body mri without contrast was undertaken. this showed a focal area of suspicious signal intensity in the upper paraspinal musculature. an ultrasound and mri with contrast demonstrated an extensive infiltrative vascular lesion involving the paraspinal musculature, prevertebral space, posterior extrapleural space, mediastinum, and neck. the child was commenced on prednisone ( mg/kg/day) and rapamycin ( . mg/m twice/day). there was no clinical or laboratory improvement after one month. a biopsy was performed which confirmed khe. in the second month of rapamycin therapy, the platelet count gradually normalized and the patient was discharged from hospital at . -months of life. prednisone was weaned off at . months of life. a repeat mri at months showed significant reduction in the khe. he is now almost years into therapy and doing well. conclusion: this is a unique case of khe with kmp that initially presented with extensive and recurrent pleural and pericardial effusions. this case demonstrates the importance of suspecting an underlying vascular malformation in the presence of kmp. our patient had a delayed but overall good response to rapamycin. further studies investigating duration of rapamycin therapy is key for the optimal management of these patients. rosa diaz, donald mahoney, lakshmi srivaths, donald yee texas children's hospital, houston, texas, united states background: since von willebrand disease (vwd) is the most common inherited bleeding disorder, it must co-exist with other less common bleeding disorders in some dually affected patients. however, reports of combined deficiencies in factor viii (fviii) and von willebrand factor (vwf) are rare. objectives: to study the prevalence and bleeding phenotype of combined deficiencies of fviii and vwf in males with hemophilia a in a hemophilia treatment center. design/method: we retrospectively reviewed the electronic medical records of males with hemophilia a followed at our institution during the past years. the primary and secondary outcomes for the study were ( ) the prevalence of combined fviii and vwf deficiencies and ( ) the bleeding phenotype of these patients. we identified vwf deficiencies in % (n = ) of the patients with hemophilia a. most (n = , %) patients were tested for vwf deficiency as part of the initial hemostatic evaluation, but one-third were tested due to clinical concern for inadequate response to fviii concentrate. the median duration of follow up was . years (range . to . ). patients were referred to our clinic at a median age of months (range to years) for evaluation of easy bruising (n = , %), mucosal (n = , %) and surgical bleeding (n = , %). primary diagnoses included with severe, moderate and mild discrepant hemophilia a. secondary diagnoses included with low vwf activity, type vwd and with type unclassified. patients experienced episodes of musculoskeletal (n = , %), mucocutaneous (n = , %) and cns bleeding (n = , %). a total of patients received factor prophylaxis. half of the patients were initially treated with fviii concentrates but subsequently changed to combined fviii/vwf products due to the frequency of breakthrough bleeding despite good compliance. all patients are on combined fviii/vwf products at the time of this review. a total of ( %) of this cohort developed chronic joint disease manifest as decreased range of motion and/or abnormal mri findings. combined deficiencies of fviii and vwf were present in % of our center's hemophilia patients. these patients exhibited a severe bleeding phenotype as evidenced by the high frequency of hemarthrosis, need for prophylaxis and high prevalence of chronic joint disease. while the optimal treatment strategy remains to be elucidated, early recognition of a combined deficiency may have important clinical implications, particularly in patients who demonstrate a suboptimal response to fviii concentrate alone. background: childhood neutropenia is heterogeneous and may be congenital or acquired. cerebral cavernous malformation (ccm ) is a neurovascular malformation disorder where lesions consist of low flow, dilated capillary endothelial channels with increased permeability, predisposing to hemorrhage and thrombosis. programmed cell death protein (pdcd ) activity has been implicated in glia and neuron migration, and recently linked to the dysregulation of the actin and microtubule cytoskeleton, thereby affecting cellular morphology and migration. variants of pdcd encoding pdcd have been associated with ccm . ccm causes a greater and earlier disease burden than other ccms, with % presenting younger than years. some patients have associated extra-neuronal manifestations, suggesting that pdcd plays a role in other tissues. we describe a patient with significant blood cytopenias associated with ccm . design/method: retrospective chart review to obtain patient data. results: an -month old female presented with seizure and was found to have multiple intracranial cystic lesions and abscesses due to s. pneumonia serotype f. during her treatment, she developed anemia (hemoglobin . - . g/dl), thrombocytopenia (platelets , - , cells/l), and profound neutropenia (absolute neutrophil counts of zero). initial bone marrow evaluation revealed a normocellular marrow but with marked granulocytic hypoplasia and % hematogones on flow cytometry. florescent in situ hybridization excluded cytogenetic changes characteristic of myelodysplastic syndrome. further evaluation included testing for neutrophil antibodies, chromosome breakage, and telomere length and results were normal. whole exome sequencing excluded mutations affecting congenital neutropenia genes, but detected a de novo pdcd variant (c. + g>a), thereby diagnosing ccm . the neutropenia has responded well to granulocyte colony stimulation factor (gcsf), which is still needed at months of age. moreover, the thrombocytopenia has progressed, requiring periodic platelet transfusions. over time, the bone marrow hematogone population has decreased to % at months of age, though the granulocytic hypoplasia persists. conclusion: our case describes the first patient with neutropenia and thrombocytopenia associated with ccm . we hypothesize the pdcd variant is the etiology of bone marrow dysfunction due to its role in actin and microtubule cytoskeleton formation, akin to the pathophysiology of xlinked neutropenia. supportive features of an underlying genetic cause of marrow dysfunction include the persistence of cytopenias beyond infection resolution as well as presence of hematogones. hematogones were previously reported to occur in patients with other congenital neutropenia disorders, indicating they could be a feature of congenital neutropenia and may be reactive to surrounding cell apoptosis. further testing of pdcd role in hematopoiesis should be explored. background: - % of adult women will suffer from heavy menstrual bleeding (hmb) during their lifetime. % of women with inherited bleeding disorders suffer from hmb. there is a paucity of data about hmb among adolescents and young adults (aya), a population in which hmb may have large social and educational effects. objectives: to study the social and academic implications of hmb in an aya population. design/method: this is a questionnaire based survey conducted in a medium-sized city in california. we recruited females - years of age from one high school and from local university. the questionnaire was set up in research electronic data capture (redcap) at our institute which allowed us to obtain objective data about the respondents' menstrual cycles. a link was sent to the high school students via their online portal schoolloop and to the university students via social media and word of mouth. data was collected over weeks from may to august . we received replies, some were not complete. using regression analysis, data was analyzed from respondents in the age group of - (with a mean age of ) years. we developed a composite score for hmb based on factors including saturation levels, number of pads, duration of bleeding, soaking of a pad within two hours, passage of clots, size and number of clots, and gushing sensation. we conducted statistical analysis of the drivers and implications of hmb based on the composite score. results indicate that having a relative with hmb, having other bleeding problems, and having anemia are drivers of higher hmb score. the results also indicate that hmb adversely affects quality of life as measured by participation in sports, social activities, after-school activities, tiredness, absenteeism, and gpa. hmb is also associated with increased rates of anemia and use of anti-depressants. hmb-driven anemia further adversely affects gpa. under-represented minorities are more likely to have a higher hmb score, as well as an increased adverse effect of hmb on gpa. the results suggest that the social costs of hmb are pervasive in the aya population, and especially pronounced among minorities. a relative with hmb is a significant driver of heavy menstrual bleeding. a hemostatic screen should be included when assessing the aya population with hmb. johns hopkins all children 's hospital, st. petersburg, florida, united states background: propranolol is a non-cardioselective beta blocker medication frequently prescribed for hemangiomas and hyperthyroidism. propranolol inhibits types i and ii iodothyronine deiodinases, enzymes that convert bioinactive thyroxine (t ) into bioactive triiodothyronine (t ). hypothyroidism is a well-recognized complication of diffuse hepatic hemangiomas that produce type iii deiodinase, an enzyme that converts t into bioinactive reverse t and t into diiodothyronine. thyroxine is typically selected for replacement in this population, even though doses up to % above physiologic may be necessary. we hypothesized that low dose, nearly physiologic t would be safer and equally effective because it bypasses propranolol's impact on the pituitarythyroid axis. we report an infant with diffuse hepatic hemangiomatosis and acquired hypothyroidism successfully treated with propranolol, prednisone, and triiodothyronine. design/method: a mo healthy female presented with abdominal distension, poor oral intake, and hepatomegaly. mri confirmed diffuse hepatic hemangiomatosis, the largest lesion measuring . cm by . cm. thyrotropin (tsh) was elevated at . (reference range* . - mcgiu/ml), total t # (rr - ng/dl), and total t ^ . (rr - mcg/dl). treatment was started with prednisone ( mg/kg/day) for three weeks, propranolol ( mg/kg/day) and t ( . mcg/kg/day). the t dose was slowly titrated to a maximum of . mcg/kg/day. thyroid hormone levels rapidly improved on t replacement. after two weeks, the tsh was . , tt , and tt . . after eight months, the tsh was . , tt , and tt . . at twelve months, the tsh dropped to . , tt , and tt . , suggesting decreased tumor production of type iii iodothyronine deiodinase. liver mri confirmed fewer hemangiomas, largest being . cm by . cm. the patient's t dose was reduced. both propranolol and t were discontinued after twenty-four months of treatment. one year off all therapy, this child has normal growth and development, only two < . cm hepatic hemangiomas and no evidence of hypothyroidism (tsh . ; tt ; tt . ). conclusion: t at near physiologic doses corrects the consumptive hypothyroidism associated with diffuse hepatic hemangiomas. t replacement is preferable to thyroxine due to its lower risk of rebound hyperthyroidism as the hemangiomas involute and type iii deiodinase production declines. there are two prior case reports describing t use without t , one employing propranolol and the other utilizing steroids for hemangioma management. this is the first case report with long term follow-up of a child treated with multimodal therapy including propranolol, prednisone, and triiodothyronine. *rr = reference range; #tt = total t ;^tt = total t background: multifocal lymphangioendotheliomatosis with thrombocytopenia (mlt) is a rare congenital disorder first described in that is characterized by multiple vascular abnormalities commonly involving the skin and gastrointestinal tract as well as consumptive coagulopathy often resulting in gi bleeding in infancy( ). to describe an unusual presentation and successful management of mlt in a neonate. design/method: baby h was born at full term after a pregnancy complicated by maternal sinus venous thrombosis requiring anticoagulation beginning at weeks. at birth, she was diagnosed with multiple hemangiomas based on clinical exam. at two weeks of age, she developed melena and hematemesis. cbc revealed platelet count of and she was referred to the ed. abdominal ultrasound was concerning for abnormal hepatic waveform; cxr showed multiple pulmonary nodules. workup revealed no other lesions and no further hematologic abnormalities. biopsy of presumed hemangioma ultimately revealed a smooth muscle-lined vascular proliferation without glut- immunoreactivity, consistent with mlt. her early course was complicated by an acute hemodynamically significant gi bleed; esophagogastroduodenoscopy identified six bleeding vascular malformations within the stomach that were injected with epinephrine and sclerosed with successful hemostasis. she received multiple prbc and platelet transfusions. central access was obtained and she was started on oral sirolimus based on previous reports of successful use in management of vascular malformations given its antiangiogenic and immunosuppressive effects ( ). she has tolerated it well with no evidence of toxicity and has achieved a partial response with stable of hemoglobin > and platelet count > . cutaneous lesions have diminished in intensity and she has had no further signs of gi bleeding. she receives pentamidine for pcp prophylaxis. she continues to have appropriate growth and development. we describe here an unusual presentation of an already rare disease. while cutaneous and gi lesions are typical of mlt, pulmonary involvement is not well-described in the literature. early identification of tissue-based diagnosis enabled timely stabilization and treatment of the patient. five months later, she continues to tolerate sirolimus and has shown significant response with diminished coloration of cutaneous lesions, stable blood counts, and no further bleeding. mlt is a relatively newly-recognized disorder with significant phenotypic variability. given that bleeding secondary to a kasabach-merritt-type consumptive thrombocytopenia is the major cause of morbidity and mortality in the first year of life in children with mlt, it is essential to recognize the diagnosis and initiate appropriate treatment as early as possible. north, arch background: patients with generalized joint hypermobility (jhm) may experience easy bruising or bleeding given the association between these symptoms and abnormalities in collagen, a required component of primary hemostasis. heavy menstrual bleeding (hmb) is a common initial presentation for females with underlying hemostatic defects and may be the sole manifestation of a bleeding disorder. however, limited reports describe jhm as a cause of hmb, leading to under recognition. objectives: to describe the clinical characteristics and management of young women presenting with hmb in the setting of jhm. design/method: this study utilized our hmb research registry. we included subjects - years, seen in the nationwide children's young women's hematology clinic between february and november with both hmb and jhm. medical records were retrospectively reviewed for history of presentation, menorrhagia impact questionnaire (miq): a validated quality-of-life tool for females with hmb, medication profiles and relevant laboratory studies. results: twenty-five patients met inclusion criteria (median age years, range - ) with an average beighton score of . (range to ). participants presented an average of . years (range months to years) after menarche despite % of patients reporting heavy to very heavy menses since menarche. according to the miq responses, most participants expressed hmb-associated limitations in physical activities ( %), social activities ( %), and work or school activities ( %). of the participants, % reported bleeding symptoms in addition to hmb, most commonly easy bruising ( %), epistaxis ( %) and cutaneous bleeding ( %). forty percent of young women presented with anemia due to chronic blood loss. results of hemostatic testing were unremarkable, with the exception of one patient who was also found to have type von willebrand disease. additionally, % of females reported arthralgia, with knees and ankles the most commonly affected joints. at time of presentation, % of participants reported failure of initial therapies and most patients ( %) were managed long-term with oral hormone therapy. in a small population of young women found to have jhm who initially presented with hmb, patients were likely to have prior bleeding symptoms as well as substantial delays from menarche to timing of presentation at our young women's hematology clinic despite limitations in activities of daily life. greater awareness of the associations between bleeding symptoms and jhm, despite typically normal hemostatic laboratory results, is necessary so that patients can more easily be identified and receive appropriate therapy. the objective is to determine the impact of cl care practices involving the home environment on ambulatory clabsi rates. design/method: information for the pi was collected through a comprehensive survey that was completed annually by the ccbdn member hospitals. responses to the questions about cl care practices involving the home environment were selected from the pi for . ambulatory clabsi rates and ambulatory total bloodstream infection (bsi) rates were obtained from another ccbdn database. the proportion of hospitals that did or did not employ a particular cl care practice was tallied. the mean ambulatory clabsi rate and mean ambulatory total bsi rate of the hospitals that did or did not employ a particular cl care practice were compared using generalized linear model techniques assuming an underlying negative binomial distribution. results: twenty-five hospitals submitted responses to the questions about cl care practices involving the home environment. one hospital was excluded for lack of bsi data. sixty-three percent of the hospitals programmatically educated parents about all aspects of the cl care bundle. the mean ambulatory clabsi rate for the hospitals that educated parents was significantly lower than that of the hospitals that did not ( . infections/ cl days vs. . infections/ cl days; p = . ). the mean ambulatory total bsi rate was also significantly lower ( . infections/ cl days vs. . infections/ cl days; p = . ). the mean ambulatory clabsi rates and mean ambulatory total bsi rates were not significantly different for the other cl care practices. conclusion: an analysis of cl care practices involving the home environment reveals that parental education of all aspects of the cl care bundle is associated with a lower ambulatory clabsi rate and lower ambulatory total bsi rate. this finding highlights the importance of systematically teaching family members the proper method of handling cl. background: children undergoing chemotherapy are at a high risk for developing nausea. dr. amy baxter in collaboration with pediatric oncology patients and nurses, developed and validated a pictorial nausea rating scale for children aged - years, called the baxter retching faces (barf) nausea scale. staff nurses at a large, academic, pediatric hospital located within washington, d.c., have identified variability in nursing assessment and documentation of chemotherapy induced nausea and vomiting (cinv) in pediatric oncology patients. the purpose of this quality improvement project was to utilize the barf scale to standardize assessment and documentation of nausea in pediatric oncology patients receiving chemotherapy. the primary aims of this project were to: assess feasibility of the barf scale in clinical practice; increase nursing knowledge about cinv through education sessions; increase documentation of nausea assessments through the use of the scale. the secondary aim of this project was to: increase the recognition of nausea through the use of a standardized assessment tool. design/method: the pdsa model was used to guide the design and implementation plan. in the first phase of the project data was collected to identify the prevalence of nausea in patients admitted for chemotherapy in the prior three months. education sessions discussing cinv and the utilization of the barf scale were conducted. pre and post assessment of nurses' knowledge of cinv and documentation were assessed. in the second phase the barf scale was implemented into practice. nurses were asked to utilize the barf scale to assess and document nausea scores in patients, aged to years, receiving chemotherapy. at the end of the implementation period nurses were surveyed about the feasibility of the scale. post data was collected to identify the prevalence of nausea documented in the electronic health record. this project was undertaken as a quality improvement initiative at children's national and it does not constitute as human subjects research. as such it was not under the oversight of the institutional review board. results: all data has been collected; however complete data analysis will be conducted in the upcoming weeks. background: sickle cell disease (scd) is the most common inherited blood disorder in the united states (us); however, there are few quality measurements to evaluate scd practice. in , the nhlbi published guidelines that include two key interventions for children with sickle cell anemia (sca): the use of transcranial doppler (tcd) screening for stroke prevention and hydroxyurea (hu) to prevent scd pain crisis. we conducted a national survey of scd management sent to providers in over institutions in the us to better assess knowledge of the guidelines and barriers to hu counseling and tcd screening guideline implementation. it was hypothesized that the barriers to tcd screening are different than barriers to hu counseling and prescribing. a -question anonymous survey was sent to providers by mail (follow-up by email). survey themes included nhlbi guidelines knowledge and comfort with understanding and implementing both tcd screening and hu use. the response rate was % ( / ) however one survey was incomplete. thus, were analyzed in the final data set. all of the respondents are in active practice, % s of s in academics and all care for children with scd. the majority of providers ( %) felt "very" or "extremely" confident in their knowledge of tcd screening and interpretation. similarly, % of providers felt "very" or "extremely" familiar with hu dosing and management. for tcd screening, % of providers estimated their screening rates were > % and % providers felt their annual screening rates were - %. the two biggest barriers to tcd screening noted by providers (of moderate to extreme significance) included: lack of support staff ( %) and lack of time during a patient visit ( %). regarding hu prescribing practices, % of providers offered hu to at least % of children with sca over nine months of age. the biggest barrier to hu prescribing noted by % of providers was concerns about patient adherence or access to the medication. only % providers felt that lack of support staff was a moderately significant barrier to hu prescribing. the pediatric scd providers surveyed all have access to the nhlbi guidelines. despite widespread guideline knowledge, there are different barriers for tcd screening versus hu prescribing, which prevent optimal implementation. as a result, although both recommendations are from the same nhlbi guideline, they likely will require different implementation strategies (systems-based interventions for tcd screening; interventions to improve patient adherence for hu counseling) to improve outcomes. background: invasive fungal disease (ifd) is a major cause of mortality and morbidity among pediatric immunocompromised patients such as those who receive chemotherapy or hematopoietic stem cell transplantation. the current diagnostic 'gold standard' of ifd remains culture of infected tissue obtained by biopsy. noninvasive biomarker testing for galactomannan or , -beta-d-glucan (bg) can have low sensitivity and does not provide species-level identification. nextgeneration sequencing (ngs) of cell-free plasma is a promis-ing noninvasive approach to providing species-level identification of ifd via a blood test and can further guide specific treatment. objectives: describe the incidence of positivity for fungal specific pathogens on ngs analysis in a high-risk immunocompromised pediatric population and correlate results with other 'standard' infectious studies if performed. design/method: immunocompromised pediatric patients with suspected ifd were enrolled and plasma was collected at time of enrollment. ngs was performed on extracted dna in cell-free plasma (karius, redwood city, ca). after removing human reads, remaining sequences were aligned to a curated database including pathogens. organisms present at a significance-level above a predefined threshold were reported. results: twenty-seven samples from enrolled patients have been processed thus far. of these subjects, were enrolled for prolonged febrile neutropenia (≥ hours) despite broad-spectrum antibiotics, for recrudescent febrile neutropenia, for abnormal imaging, and with other findings. after evaluation of routine studies performed, patients met criteria for proven ifd, for probable ifd, and for possible ifd using eortc/msg guidelines. the ngs plasma test identified the same pathogen as cultured from infected tissue or blood in % ( / ) of the proven cases. in the probable cases, pneumocystis jirovecii was identified in a patient with a positive bg ( pg/ml) and pneumonia. among the possible cases, toxoplasma gondii was detected in a patient with prolonged febrile neutropenia and lung imaging suggestive of ifd. additionally, candida glabrata was isolated in a patient with prolonged febrile neutropenia but no other criteria for ifd. numerous pathogens were also identified that could explain the above clinical parameters, including hsv , cmv, vzv, hhv , ebv, bk polyoma virus, and ureaplasma parvum. the cell-free plasma ngs test can detect invasive fungal infections from blood. the test identified fungi from proven ifd, detected pathogens in both probable and possible ifd cases, and is a useful diagnostic tool in the evaluation of ifd. supplies and sample shipment and processing supported by karius, inc. baylor college of medicine, texas children's hospital, houston, texas, united states background: practicing medicine is a lifelong learning process. as noted in the institute of medicine's seminal report, 'to err is human,' adverse outcomes do not typically result from individual recklessness; rather, they result from faulty systems, processes, or conditions that provide an environment conducive to making a mistake, or failing to prevent one. learning to systematically review errors and translate lessons learned into quality improvement (qi) initiatives is a critical component of practice-based learning and improvement for practitioners at all career levels. objectives: to develop a methodical, self-reflective and nonthreatening approach to incident analysis and translation of lessons learned into qi initiatives. design/method: we used a validated, structured case audit approach, modified from szostek et al: ) review all documentation relating to the case and identify all health care providers involved; ) interview stakeholders, including those who directly provided and supported care; ) use a qi tool to conduct a root-cause analysis; ) identify a systems issue that contributed to the outcome; and ) propose systems-level interventions and prioritize initiatives based on effort-yield projections. results: pdsa cycle : plan: establish a committee to ) identify potential cases, ) triage cases for conference presentation, ) determine timing and frequency of conferences, ) develop a training manual, ) record identified qi initiatives. do: we established a quarterly section-wide meeting to which all members of the pediatric hematology/oncology service are invited, including administrative and nursing leadership. we developed a training manual and structured presentation template. prioritized cases were discussed in advance during multidisciplinary case review sessions, and presented by senior fellows who were instructed to focus discussion on potential opportunities for qi. study: we identified cases, meeting criteria for mmi presentation. qi initiatives identified from this conference resulted in a number of systemic practice changes; however, we encountered challenges to sustaining these changes over time. act: objectives for the next pdsa cycle are to ) establish a method for tracking the adherence to recommended changes in practice, ) maximize sustainability by integrating qi initiatives into institutional qi leadership and practice standardization committees. we have successfully implemented an mmi conference that meets out of institute of medicine quality domains: safety, effectiveness, patient-centeredness, timeliness, and efficiency. a standardized, consistent approach to mmi presentations that includes identification of contributing factors and specific qi implications has the potential for improving both provider education and patient care/safety. johns hopkins university, baltimore, maryland, united states background: receiving a cancer diagnosis is a life-changing event for patients and caregivers, although little is known about the experience. while some oncologists receive dedicated training in delivering this bad news, the initial conversation is often with a primary pediatrician, and these providers often feel they do not receive adequate training in the communication of a cancer diagnosis. objectives: our objectives were two-fold: first, to better define the experiences of caregivers/patients when told of a cancer diagnosis, and to query how caregivers/patients believe providers can improve the disclosure of this bad news. secondly, to assess what, if any, training primary pediatricians received in this skill, and to assess how comfortable providers in various settings and stages of training are with communicating cancer diagnoses. design/method: from november - , semistructured, in-depth interviews were conducted with pediatric oncology patients and caregivers of patients (n = ) diagnosed in the past year regarding their experiences receiving the diagnosis at our institution. in addition, pediatric residents (n = ), outpatient pediatric primary care physicians and pediatric emergency medicine physicians (n = ) were interviewed regarding their experiences delivering cancer diagnoses. interviews were analyzed following principles of thematic analysis. interviewers with patients and caregivers had two common themes: ) all emphasized their wish for direct and thorough information; ) both patients and caregivers emphasized the gratitude they felt for physicians who gave them hope by emphasizing the good prognosis of their child's cancer. lack of training in this area, as well as lack of comfort delivering this news was common will all providers. additionally, providers report variable approaches to giving bad news, including ) whether to tell caregivers separately or tell the child and parents together, and ) whether to give favorable prognostic information. additionally, attending physicians also differed significantly in their approaches to teaching residents. while some believed residents should give the news to gain experience, others felt that this is not appropriate if residents are inexperienced. only one resident reported ever receiving feedback on his communication skills in this type of discussion. conclusion: we plan to build on these interviews to develop a national survey of patients, caregivers, and providers to better understand the issues surrounding this discussion. we will use the findings to develop a communication curriculum for pediatric residents, focusing on the discussions that occur in the outpatient setting by primary pediatricians. background: human papilloma virus (hpv), common in both females and males, is responsible for pathologies ranging from benign genital warts to cervical and penile cancer. hpv strains and are responsible for , malignancies each year in the united states, and one third of them arise in men. pharmaceutical companies have now developed a vaccine that will help prevent the virus-associated malignancies. the cdc initially recommended that females ages - years receive the vaccine series, then starting in they expanded the eligibility to males ages - years. despite being widely available and highly publicized, only % of eligible females receive the full vaccine series. objectives: this study aims to assess the knowledge of hpv, the attitudes towards the hpv vaccine, and identify barriers preventing its full utilization. once identified, we aim to overcome the barrier(s) in order to improve vaccination rates in eligible adolescents. we distributed a standardized questionnaire to the parents of eligible female and male patients in our pediatric hematology-oncology clinic. it assessed the parents' knowledge of hpv and the vaccine, their views of the vaccine, and reasons why they may oppose it. results: approximately % of parents claim they have been educated about hpv, mostly by their primary care physician. however, % did not know what disorders hpv caused; % felt the vaccine should not be added to the typical vaccine schedule; % of parents do not intend to vaccinate their child. of those that opposed the vaccine, one-third were concerned about potential side effects and nearly % feel they do not have enough information. additionally, % of parents are not aware that the vaccine is available at their child's doctor and only % of parents have discussed the hpv vaccine with their child's doctor. the largest barrier to the utilization of the hpv vaccine that we have identified appears to be lack of educa-tion. as a result, we have begun distributing the cdc's hpv and vaccine patient guide to our patients' families as an intervention. we are currently in the process of re-administering our survey to these families after implementing the intervention to assess its success in increasing both knowledge and utilization of the hpv vaccine. cancer institute, chennai, chennai, tamilnadu, india background: rasburicase is a recombinant urate oxidase enzyme approved for use in tumor lysis syndrome (tls) and it acts by reducing serum uric acid levels. using rasburicase at the recommended dose of . mg/kg/day for days is expensive and it is not known whether this extended schedule is clinically beneficial compared to a single fixed dose of . mg. the aim of the present study was to evaluate the efficacy of single dose rasburicase . mg in prevention and management of tls. design/method: rasburicase is available as single use . mg vial. at our institution a single dose of rasburicase . mg irrespective of bodyweight has been used in adults and in children a dose of . mg/kg (maximum . mg) has been used since for prevention and management of tls and subsequent doses are given based on biochemical response and clinical condition. we retrospectively analysed the case records of patients who had received rasburicase from january to january . the study included patients with hematological malignancies who received rasburicase. children accounted for . % (n = ) patients and males comprised % (n = ). rasburicase was used prophylactically in ( . %) patients, for laboratory tls in patients ( . %) and for clinical tls in ( . %) patients. single fixed dose rasburicase prevented laboratory/clinical tls in % of the prophylactic group and prevented clinical tls in % of the laboratory tls group. none of the patients in prophylactic and laboratory tls group developed clinical tls. however, majority of the patients with clinical tls required more than one dose rasburicase. single dose of . mg ( vial) rasburicase is efficient in preventing and managing laboratory tls and is economically viable in resource constrained settings. nicole wood, lauren amos, nicholas clark, chris klockau, karen lewing, alan gamis children's mercy kansas city, kansas city, missouri, united states background: medication reconciliation for newly diagnosed oncology patients is complicated and cumbersome. these patients are often admitted on no medications, and leave on multiple. chemotherapy and supportive medications are crucial. despite numerous individuals overseeing this process, prescribing errors or omissions still occur. when reviewing the literature, improvement occurs when there is an interprofessional and standardized process to medication reconciliation. objectives: this project's aim was to improve the accuracy of the discharge medication reconciliation process from % to % from february -august . the process measure was the percentage of patients discharged with an accurate checklist. additional time for staff spent in completing the checklist and avoiding an increased error rate by changing the prescribing process were followed as balancing measures. we created a discharge medication checklist which included a list of required home medications prescribed by the resident, ideally hours prior to discharge. it required fellow or attending review and pharmacy to review the list and educate the family. checklists were collected monthly and reviewed against the electronic medical record (emr) for accuracy. results: six pdsa cycles were completed. there were errors during the data collection time frame. in pdsa cycle , a patient received acetaminophen for pain control which is avoided at home. in addition, this patient received diphenhydramine instead of ondansetron, which is preferred as an antiemetic. in pdsa cycle , a patient with a pending diagnosis was sent home with acetaminophen. of note, this patient did not have a checklist completed upon discharge. this project provides a novel and important method to standardize the discharge medication reconciliation process in a complex patient population. it clarifies which types of medications these patients need, provides pharmacy teaching to families which was not done previously, and prescribes discharge medications to families sooner. after the first medication reconciliation error, the checklist was revised. no further errors were made following revision, with the exception of one patient without a completed checklist at dis-charge. our accuracy rate increased from % at baseline to % following implementation. we are in the process of making the checklist electronic and accessible in the emr. in the interim between the end of data collection and implementation into the emr, a leukemia patient was sent home without an epinephrine pen, further demonstrating the importance of this standardized discharge process. for this reason, we have re-instituted the checklist until the electronic version is available. background: survivors of pediatric cancer are at risk of losing pre-existing protective antibodies to vaccine preventable diseases. in a prior study, % of children < years lost humoral immunity to measles as a result of chemotherapy induced alterations in immune system. measles in recipients of immunosuppressive chemotherapy has mortality rates up to %. because of volitional vaccine refusal, there has been a dramatic increase in measles infection from cases in to in , including several statewide outbreaks. small pediatric oncology practices frequently share floor/clinic space with the general pediatric patients putting them at risk for measles since virulence starts hours prior to symptoms. there is no standard protocol for revaccinating post-chemotherapy patients. to assess measles risk based on serial humoral immune status in a cohort of pediatric oncology patients receiving intensive chemotherapy design/method: patients < years age with known vaccination status receiving intensive chemotherapy between july -june at our institution's pediatric oncology practice were included in this prospective study. serial measles igg antibodies were measured at diagnosis, months and months after initiation of chemotherapy using elisa. measles immunity was defined per lab standards. a comparison of pre-chemotherapy and serial post-chemotherapy immunization titers was made for all patients by diagnosis. the study population consisted of children ( male); patients had all, non-hodgkin lymphoma, sarcoma and other solid tumors. two patients ( . %), both unvaccinated had non-protective measles antibody levels at s of s baseline. of the remaining patients, . % patients ( leukemia, lymphoma and sarcoma) lost protective antibody titers at months after initiation of chemotherapy and . % ( leukemia, lymphoma and sarcoma) at months after initiation of therapy. % of the remaining patients who retained measles antibody titers within protective range at months also demonstrated a steady decline in antibody titers at and months from therapy initiation. the loss of protective measles humoral immunity occurred significantly more often in patients with leukemia compared to other malignancies. oncology patients in our practice undergoing intensive chemotherapy demonstrated progressive waning of protective measles igg titers. our data suggests that it should be standard practice to check all patients for measles humoral immunity prior to starting chemotherapy and at completion. larger studies need to be performed to establish guidelines for revaccinating post-chemotherapy pediatric patients, an intervention that is easily applicable and of low cost. background: the accurate determination of glomerular filtration rate (gfr) is important to screen for acute kidney injury, to dose chemo-therapy, and to identify risk for chronic kidney disease.being correlated with inulin clearance, measured gfr by iohexol plasma disappearance (igfr) is a new gold standard for measurement of gfr in pediatric cohort studies. igfr is based on the clearance of an exogenous marker and is unaffected by endogenous compounds or a patient's muscle mass. we compared igfr with -hour urine creatinine clearance ( crcl) and gfr estimating equations using serum creatinine (scr) and serum cystatin c (cystc) in pediatric patients with cancer. we recruited participants who were ages to yrs, continent of urine, and diagnosed with a malignancy in the past years. eligible subjects had stable kidney function for at least two weeks prior to the assessment of igfr. consented subjects had baseline assessments including height, weight and vital signs. blood samples were obtained for serum chemistry, and time zero iohexol. igfr determined by ml iohexol solution infused over - minutes followed by ml of sterile saline. blood was drawn at , , and minutes.at the same time of igfr, the crcl was collected. igfr was calculated using a two-compartment model and area under the curve. we compared igfr to published gfr equations (schwartz et al, kidney int ). results: ten subjects ( female/ male) agreed to participate. the distribution of diagnoses for the subjects: all = , lymphoma = , brain tumors = and hepatocellular carcinoma = . six patients were off therapy. the lower gfrs are noted in patients who had malignancies other than leukemia, likely due to the use of cisplatin based therapy. the average igfr was ml/min/ . m^ whereas crcl was . ml/min/ . m^ ; demonstrating the crcl overestimates gfr compared to igfr. comparing igfr to univariate equations using scr, cystc, and the multivariate equation with both, the univariate cystc equation correlated well with igfr; the others overestimated igfr. we found that crcl overestimated igfr. the univariate cystc equation better correlated to igfr than equations with scr. the poor performance of scr based methods to assess gfr might be due to decreased muscle mass and inadequate nutritional status. creatinine-based determinations of gfr alone, may not be accurate in this population. further study is needed to determine if igfr should be a standard of care to assess gfr in children with cancer particularly who are receiving nephrotoxic medications and incontinent of urine. background: pediatric oncology patients undergoing chemotherapy through indwelling venous catheters are at increased risk for severe sepsis especially when neutropenic due to chemotherapy. rapid triage and early recognition are essential because delayed initiation of antibiotics and fluids in these patients or delayed transfer to higher level of care after initial stabilization is associated with poor clinical outcome. our pediatric oncology out-patient clinic is designated as an article unit whereby the providers can initiate and give treatment such as intravenous fluid, antibiotics, chemotherapy and blood products. objectives: global aim-optimize management of early sepsis and decreased morbidity, mortality and hospital length of stay in the high risk pediatric oncology patients. smart aim-improve timely management with initiation of fluids and antibiotics and transfer of septic patients to higher levels of care by % in months in above patients design/method: multidisciplinary team with physicians and nurses was created. retropective chart review of sepsis patients treated at the clinic from april to october was done using an audit sheet to identify the barriers in the delivery of care. three patients were identified and data analyzed prior to intervention; two were analyzed post interventions. a key driver diagram was created by the group to drive intervention. a process map was designed to identify the different steps in the care of these patients to pinpoint areas needing improvement. different timed data points were used starting from time of arrival to clinic, time to antibiotics and fluids and time to transfer to higher level of care. rapid pdsa cycles were done to improve the processes and delivery of care. run charts were created. there was an improvement close to the goal of % for all data points used. pdsa cycles for improvement included conducting frequent mock codes with appropriate feedback real time coaching and process planning with nursing staff. we partnered with pharmacy for close loop communication with clinic staff and we improved communication between physicans at different levels. conclusion: sepsis in neutropenic pediatric oncology patients is deadly and can be reversed with timely management at different levels. given the promising results of the above project, we want re-inforcement of the processes to be a part of the daily practice of first line clinical staff. eventually we will extend the principles learnt in management and triage of sepsis to other outpatient emergencies chemotherapy related anaphylaxis background: chemotherapy-induced nausea and vomiting (cinv) is a common side effect in children receiving antineoplastic chemotherapy. recommended prophylactic antiemetic medications are based on the classification of chemotherapy emetogenicity. however, despite appropriate use of these antiemetic agents, some patients will still experience nausea and/or vomiting. children's oncology group clinical practice guidelines recommend the addition of olanzapine to prophylactic regimens for management of breakthrough cinv. objectives: our pediatric hematology oncology center implemented a quality improvement (qi) project aimed to increase the use of olanzapine in pediatric cancer patients years of age and older receiving moderately or highly emetogenic chemotherapy and experiencing breakthrough cinv over a month period. design/method: this qi project was conducted utilizing plan-do-study-act (pdsa) cycles. for the first pdsa cycle, baseline data was collected through chart review to determine the rate of olanzapine use for breakthrough cinv over a month period from july to december . breakthrough cinv was defined as use of or more doses of antiemetic agents other than those given for cinv prophylaxis. guidelines for treatment of breakthrough cinv were reviewed with pediatric hematology/oncology attending physicians and fellows. flyers were created that listed chemotherapy regimens considered moderately and highly emetogenic. if a patient experienced breakthrough cinv, a flyer was to be placed in the patient's roadmap binder to signal olanzapine should be added to the next chemotherapy block. data was collected over a month period in september following this first intervention. the second pdsa cycle consisted of didactic education and training of pediatric oncology nurses as well as pediatric residents regarding the addition of olanzapine for breakthrough cinv. rates of olanzapine use were then collected from october through november . results: olanzapine use increased from . % at baseline to . % after the first pdsa cycle ( = . , p = . ). after the second pdsa cycle, olanzapine use increased another . % to . % ( = . , p = . ). the administration of olanzapine was successfully increased by modifying patients' roadmaps after patients experienced breakthrough cinv as well as with education and training of pediatric oncology staff, fellows, residents, and nurses. background: venous thromboembolism (vte) is increasingly affecting children. according to an administrative database study, there was a % increase in the incidence of vte among children admitted to free-standing children's hospitals in the united states from to . risk factors for hospital-acquired vte are well-known and well-studied in adults, with evidence-based preventative measures available. similar guidelines are lacking for children. objectives: there is an ongoing national-initiative to develop and institute methods for screening and preventing hospitalacquired vte in children. in / , nationwide children's hospital instituted an electronic screening form required for all patients admitted ≥ hours. patients were scored and riskstratified based on eight risk-categories. a summated score was used to determine the vte risk level, and used to make prophylaxis recommendations for patients ≥ years; as well as patients ≥ years who were admitted to an intensive care (icu), surgical, or trauma unit. the purpose of this irb exempt, quality improvement initiative was to retrospectively review our experience with this risk-stratification tool. results: hospital-acquired vte events occurred in unique subjects. median age at vte diagnosis was years. only ( %) vte occurred in children ≥ years of age. ( %) vte were deep vein thrombosis (dvt), and ( . %) involved pulmonary embolism. vte was most common in subspecialty units including the pediatric and cardiac icus ( . %); neonatal icu, ( . %); and hematologyoncology, ( . %). ( %) vte were associated with central venous catheters (cvc) and events ( %) were associated with altered mobility. congenital heart disease/heart failure was the most common chronic medical condition associated with vte ( ( . %) events); whereas infection and trauma/surgery were the most common acute medical conditions associated with vte ( ( . %) and ( %) events, respectively). during ( %) events, subjects scored a summated score ≥ . in summary, in this single institution, prospectively maintained database, cvc remains the most common risk factor for vte, followed by cardiac disease, infection and trauma/surgery. most subjects who developed vte scored high (score ≥ ) on our screening tool. only a small proportion of vte occurred in patients older than years and thus eligible for thromboprophylaxis. our results indicate that future vte prevention endeavors should include these age groups in addition to exploring more aggressive prophylactic modalities including pharmacological prophylaxis. background: pediatric fellows are required to have active engagement in quality improvement (qi) activities, and yet a national acgme review found most trainees had "limited knowledge of qi methods" and "limited participation in interprofessional qi teams". the twenty fellows in our pediatric hematology/oncology training program identified blood culture utilization as their qi priority. our institution recently introduced a hospital-wide decision algorithm to guide providers regarding when to obtain blood cultures. there is often a low threshold to obtain blood cultures in immunocompromised pediatric oncology patients, but these are often low-yield or result in falsepositives. our fellows spearheaded a project to implement the algorithm in the inpatient pediatric oncology population and improve the proportion of appropriately drawn blood cultures. we investigated how appropriately the algorithm was being utilized on the inpatient pediatric oncology floor prior to and after several educational steps aimed at disseminating the algorithm to members of the care team. our primary endpoint was to quantify the proportion of culture episodes drawn "inappropriately", with a goal of reducing inappropriate episodes to ≤ %. the algorithm was initially introduced to the nursing staff and residents covering the twenty-bed inpatient unit in september . qi project planning took place with upper level fellows in january . fellows and faculty received intensive training on the algorithm in july-august . we then conducted a retrospective chart review of blood culture episodes drawn between august and november . upper level fellows scored ∼ culture episodes as to whether the decision to culture and number of cultures drawn were "appropriate" or "inappropriate", and catalogued the indications for culture episodes and if applicable, why the episode was found to be inappropriate. additionally, fellows discussed inappropriate culture episodes with the team onservice, to provide direct feedback on where the algorithm failed. results: between august -december on average cultures/ patient-days were drawn. forty-nine percent of culture episodes were inappropriate. from january -october , following targeted education on the algorithm, the rate of blood cultures drawn decreased to cultures/ patient-days. the average proportion of inappropriate culture episodes fell to . %, representing a % decrease in inappropriate culture utilization. correct application of a decision algorithm for blood culture utilization can reduce total cultures drawn on an inpatient pediatric oncology unit. fellow-led education of the multi-disciplinary team decreases the rate of inappropriate culture episodes as well as provides active engagement in qi. background: inadequate understanding of sickle cell disease (scd) is common and can affect patients' compliance and therefore their morbidity and mortality, especially after transition to adult care. optimal clinical care for scd includes disease education, which can be difficult given the breadth of possible topics and limited time in clinic. it is unclear how best to provide personalized, efficient education for adolescents with scd. this quality improvement (qi) study aimed to implement a questionnaire-based system to improve patients' knowledge of their scd and documentation of education by the nurse or physician. the study objective was to improve provider documentation and patient knowledge about their scd by identifying patients' gaps in comprehension. by january , the study aimed to increase education documentation from % to %. by april , the study aimed to increase use of a smart phrase for education documentation from % to %. by june , the study aimed to increase patients' knowledge about their disease by %. design/method: twenty-one scd patients enrolled on an irb approved qi study, with twenty active patients. our comprehensive team generated a questionnaire with knowledgebased questions for two age groups: - and - years old. at each comprehensive visit, a questionnaire was distributed, with at least -month intervals. the provider scored questionnaires and reviewed two educational topics, with wrong answers taking priority. plan-do-study-act (pdsa) cycles included pdsa# : patients completed questionnaire. pdsa# : a smart phrase addressing questionnaire topics was created and shared with providers. pdsa# : patients received education handouts during clinic education. documentation in clinic notes was the process measure and questionnaire scores was the outcome measure. results: pdsa# is complete, pdsa# has four patients remaining, and pdsa# is ongoing. due to variable visit frequency, there are multiple concurrent cycles. after pdsa# , free text documentation was completed an average of % over the course of months. after pdsa # documentation increased to % within months and questionnaire scores increased from an average of % to %. of the questions that patients got wrong on their first visit, they were significantly more likely to improve on retesting if the topic was taught to them than if it was not addressed ( % vs. %, p = . ). we are currently completing pdsa# and collection of post pdsa# data. questionnaire-based scd education coupled with standardized smart phrases improves patients' scd knowledge and documentation by providers. further improvement in knowledge is expected with the addition of handouts. background: exposure to suffering can have a profound impact on the wellness of caregivers, often referred to as the "cost of caring". this cost is especially high in pediatric hematology/oncology. repeated exposure to suffering has the potential to negatively impact resilience and increases the risk of burnout, thus impacting quality of care and patient satisfaction. we have developed a peer support team utilizing the critical incident stress management (cism) model. this model has been successfully used in other professions that frequently face traumatic events such as fire fighters, police and emergency medical technicians. the h.o.p.e.s. team (helping our peers endure stress) consists of volunteer multidisciplinary staff members who have received training to provide both group and peer support following any 'critical incident' that may impact one or more staff members. we hypothesize that implementation of the h.o.p.e.s. team will improve staff resilience, decrease overall rates of burnout and improve compassion satisfaction. s of s design/method: we are using both empiric metrics and anecdotal reports to assess the impact of the h.o.p.e.s. team. prior to the activation of the team, all pediatric hematology/oncology clinical staff members were surveyed using validated tools to assess their levels of resilience, burnout, secondary trauma and compassion satisfaction (proqolv and brief resilience scale). they were also asked to rate the number of times they had experienced critical incidents, as well as their perceived level of distress after dealing with traumatic events. after the h.o.p.e.s. team has been functional for months, we will send the same survey to staff members to measure changes, paying special attention to resilience and rates of burnout and compassion satisfaction. results: enthusiasm for development of the team has been high. of people approached to volunteer their time to participate in the multidisciplinary team agreed, including attending physicians, fellows, nurses, nurse practitioners, child life specialists, social workers, clergy and psychologists. all volunteers participated in a -day training conducted by an instructor from the international critical incident stress foundation. engagement in the first staff survey has been high, with of responding to date. data collection is ongoing. clinical staff in pediatric hematology/oncology may be particularly vulnerable to burnout and decreased resilience by repeatedly witnessing suffering and trauma. peer support interventions following critical incidents may lead to increased resilience and compassion satisfaction while decreasing rates of burnout. enthusiasm for the development of a peer support team has been high. background: monthly blood transfusions are an indicated therapy for pediatric patients with sickle cell disease with certain complications. maximizing transfusion efficiency in a busy infusion clinic requires: ensuring that appropriate blood units are available in the hospital blood bank; laboratory specimens are obtained from patients in advance; and coordination of clinic appointment and nursing availability. we sought to improve clinic efficiency through identifying ways to better communicate with patients/families regarding upcoming laboratory and transfusion appointments, and to assess the efficacy of implementing a web-based personalized text reminder (pinger.com). we measured the baseline frequency with which transfusion appointments were missed by families, moved to later within the week, or delayed due to late labs. a convenience sample of patients receiving monthly transfusions received a questionnaire about patient/parent preferences for appointment reminders and barriers to keeping appointments. those patients/parents who did not opt-out of an additional text reminder received personalized texts from their care team reminding them of lab and transfusion appointments. rates of missed/moved/delayed appointments were compared between the group receiving the additional text messages and the group only receiving standard, hospitalgenerated appointment reminders (telephone call). results: forty-one families ( patients) responded to the survey, capturing information on % of patients receiving chronic transfusion therapy. thirteen families ( %) declined the additional text reminders. families reported a preference for text reminders ( %), more often than email ( %) or telephone ( %), and % of families wanted to receive reminders for both transfusion and laboratory appointments. the majority ( %) of families reported competing work/life priorities as the reason for missed/late appointments. other families noted transportation/travel ( %), fear/illness/pain ( %), and lack of reminders ( %) as the reason for missed appointments. at baseline (twelve weeks), . % of appointments were missed on a weekly basis (range - of available per week), . % were moved, and % of appointments were delayed. during our intervention period (twelve weeks), % were missed, . % were moved, and . % were delayed (combined, both groups). there was no difference in missed ( . % texted vs . % standard), moved ( . % texted vs . % standard) or delayed ( . % text vs . % standard) appointments. though families at our center reported a preference for a text-based reminder, personalized text reminders for appointments did not improve clinic efficiency as measured by missed, moved or delayed transfusion appointments. there was no improvement in appointment adherence in the group receiving personalized texts in addition to standard hospital reminders. university of utah, salt lake city, utah, united states background: childhood cancer outcomes have improved significantly, in large part due to multi-institution collaborative clinical trials run by the children's oncology group (cog). approximately half of eligible children with cancer will enroll on a therapeutic trial, but little is known about the factors affecting caregiver decision-making regarding enrollment or how well the required elements of informed consent are conveyed during the consent process. objectives: . assess coverage of ten of the required elements of informed consent for cog therapeutic trials. . describe factors affecting caregiver decision-making regarding therapeutic trial enrollment. we surveyed families of children who were offered enrollment onto a phase cog therapeutic study for an initial cancer diagnosis in the previous months. fisher's exact or wilcoxon rank-sum tests were utilized to compare demographic and other motivating factors related to enrollment decision-making. results: seventy participants were surveyed. regarding of the basic required elements of informed consent, % knew the trial involved research, % knew consent was required, % knew the enrollment length for the trial, % knew they could continue care independent of enrollment, % knew who to contact with questions, % knew there were options besides enrollment, % knew they could withdraw at any time, % knew the information was confidential, % knew there were risks associated with the trial, and % knew there were benefits. of all participants, % (n = / ) enrolled onto a therapeutic study. among enrollees, % (n = / ) of the primary caregivers had completed college compared to % (n = / ) of those not enrolled (p = . ). when asked about factors impacting their decision, % (n = / ) of those enrolled said they felt there were no risks or did not know if there were risks associated with the study compared to % (n = / ) of those choosing not to enroll (p = . ). of those enrolled, % (n = / ) reported the physician recommendation "somewhat" or "strongly" affected their decision to enroll compared to % (n = / ) of those not enrolling (p = . ). of those who enrolled, % (n = / ) reported feeling pressured to enroll while % (n = / ) of those not enrolled reported pressure (p = . ). of enrollees, % (n = / ) reported they did not have enough time to decide compared to % (n = / ) of those not enrolled (p = . ). failure to convey all required elements of informed consent highlights possible deficiencies in the consent process for cog therapeutic trials. caregivers' perception of being pressured and lack of time to make an informed decision may impact clinical trial enrollment. background: abnormal uterine bleeding (aub) is a frequent adolescent gynecologic complaint. however, limited research exists to guide management, and acute care varies. we sought to improve emergency care for adolescents with aub by developing a clinical effectiveness guideline (ceg) and assessing its impact on quality of care. design/method: a stakeholder engagement group consisting of members from the departments of hematology/oncology, adolescent medicine, general pediatrics, and emergency medicine designed a ceg algorithm for emergency aub management. pediatric residents received ceg training and their knowledge and attitudes were assessed using pre and post intervention surveys. icd- and codes identified electronic health record data for patients presenting to the pediatric emergency department (ed) for aub months before and after ceg implementation. pre-pubertal patients and those with vaginal bleeding from trauma were excluded. a weighted, -point scoring system consisting of prioritized aspects of history, laboratory studies and management was developed to quantify the quality of care provided. t-test, chi square test, wilcoxon rank sum test, and a run chart were used for analysis. of the patients identified, met inclusion criteria. there were % of patients currently using some form of contraception, while . % had bleeding related to a current or recent pregnancy. median aub quality care scores were pre-and post-intervention (p = . ). run chart data showed no shifts or trends (overall median score, -points). both pre and post-implementation, points were deducted most frequently for not assessing personal/family clotting disorder history and inappropriate use/dosing of oral contraceptives. we successfully designed and implemented a ceg and educational intervention for aub management in a pediatric ed. these data suggest our ceg may be an effective tool to improve emergency aub care for adolescents, though additional cycles are needed. background: high-dose methotrexate (hd-mtx) is a common chemotherapy administered inpatient at most centers. its administration is particularly susceptible to error due to the need for frequent drug levels with resulting changes in supportive care. errors can prolong patient stay and cause patient harm. objectives: global aim-to reduce the length of stay (los) of hd-mtx admissions. smart aims-to increase the percentage of patients whose pre-hydration fluids are started by am from % to % by / / , and to increase the percentage of patients who receive hd-mtx by pm from % to % by / / . we used rapid process improvement methods to target earlier methotrexate administration. a key driver of prolonged los was hypothesized to be drug levels returning overnight rather than in the day time due to delayed hd-mtx start. changes implemented have included scheduling hd-mtx patients as the first patients of the day for their exam in clinic and scheduling labs to pass for hd-mtx on the day prior to admission. there are ongoing pdsa cycles to change the location of pre-hydration start from the inpatient room to the clinic exam room in order to meet hd-mtx administration time goals. we are piloting two different education materials to improve patient experience. one explains hd-mtx levels in a red/yellow/green stoplight format and the other reminds patients how to prepare for the admission. other interventions regarding how we test urine ph and safety checks in the ordering process for history of delayed clearance are in the planning stage. the project is ongoing, but as of / / , we start methotrexate by pm % of the time which is improved from a baseline of %. when the project was started, pre-hydration was never started before am. now, fluids are started by am % of the time. pdsa cycles are ongoing and we have yet to sustain reductions in los, but some months have shown decreased los by as much as hours from baseline measurements. rapid cycle improvement can be utilized to decrease los hd-mtx admissions. this has important financial implications as well as the potential to reduce secondary harm from unnecessary time in the hospital. pediatric cancer centers should schedule hd-mtx admissions first thing in the morning so that data regarding kidney injury and drug clearance can be interpreted by the day team and children are not cleared for discharge in the middle of the night. background: education and training for interdisciplinary pediatric oncology providers requires training in principles of palliative and end-of-life (eol) care. the experiences of bereaved parents can inform and enhance palliative care educational curricula in uniquely powerful and valuable ways. the objective of this study is to present an innovative palliative care educational program for oncology providers facilitated by trained bereaved parents who serve as volunteer educators in local and national palliative care educational forums and to describe how incorporation of bereaved parents in these educational forums affects participant comfort with communication and management of children at the eol. design/method: survey tools were adapted to determine how bereaved parent educators affected participant experiences in different educational forums: institutional seminars on pediatric palliative and eol care, role-play based communication training sessions, and an international symposium on pediatric palliative oncology. pre-and post-session surveys with incorporation of retrospective pre-program assessment item to control for response shift were used in the evaluation of institutional seminars and communication training sessions. results from feedback surveys sent to all attendees were used to appraise the participants experience in the international oncology symposium. results: involvement of trained parent educators across diverse, interdisciplinary educational forums improved attendee comfort in communicating with, and caring for, patients and families with serious illness. importantly, parent educators also derive benefit from educational with interdisciplinary clinicians. integration of bereaved parents into palliative and eol care education is an innovative and effective model that benefits both interdisciplinary clinicians and bereaved parents. background: poorly controlled chemotherapy-induced nausea and vomiting (cinv) significantly impairs patients' quality of life and contributes to ongoing medical costs through increased length of stay in the hospital or readmissions and outpatient visits for control of nausea, vomiting or dehydration. lack of adherence to national evidenced-based guidelines that dictate antiemetic prescribing for variably emetogenic chemotherapy leaves patients vulnerable to increased cinv and its ensuing complications. objectives: to review our institution's antiemetic prescribing practices and their consistency with the antiemesis guidelines from the national comprehensive cancer network (nccn) and children's oncology group (cog)-endorsed supportive care guidelines and to further develop tools to increase adherence to these national-based guidelines to improve control of cinv. we performed a retrospective chart review of inpatient chemotherapy encounters. we evaluated emetogenicty of chemotherapy (high, medium, low), initial antiemetic regimen ordered, number of as needed medications required and adherence to national evidenced based guidelines tailored to each level of emetogenicity in the prescription of antiemetics. results: fifty-five total inpatient chemotherapy encounters were reviewed over months. eighteen of these encounters were considered to have been highly emetogenic chemotherapy (hec) with the remaining of these considered to be moderately emetogenic. only out of hec encounters completely included all guideline-recommended agents. there was a demonstrable lack of consistency across providers with dosing of aprepitant and most as needed medications. there was significant variation in order of first, second and third line anti-emetics ordered -with lorazepam and promethazine being used most frequently. with an aim of improving antiemetic prescribing practices for our patients, we are currently rebuilding chemotherapy treatment plans in our electronic medical record to incorporate antiemetic drug order sets that follow evidenced-based guidelines for variably emetogenic chemotherapy. this will be used in conjunction with an education initiative about best practices in supportive care for all prescribers of antiemetics. review of our department's recent inpatient chemotherapy encounters show we are falling short in following nationally recommended standards for appropriate antiemetic coverage during chemotherapy. identification of these deficiencies allows for implementation of quality initiatives to improve prescriber adherence to evidenced-based guidelines for better control of cinv. background: there are currently no consensus guidelines for the management of pediatric oncology patients presenting with fever without neutropenia. historically, these patients had been treated similarly to neutropenic patients with empiric antibiotics. while there has been a shift towards reducing unnecessary empiric treatment, there has been limited research into the outcomes associated with withholding empiric iv antibiotics in this patient population. we assessed the safety and efficacy of our institution's current protocol of observing well-appearing patients who present with fever without neutropenia and compared the outcomes of the patients who did and did not receive empiric iv antibiotics. design/method: this was a prospective, single-institution cohort study. patients were included if they were currently undergoing chemotherapy for an oncologic diagnosis and presented initially as an outpatient with fever and nonneutropenia (defined as anc ≥ cells/mm ). for each episode we recorded lab and blood culture results, signs and symptoms of initial presentation, and clinical outcomes, including antibiotic administration and hospital admission. results: a total of episodes of well-appearing patients with fever without neutropenia were identified. compliance with the institutional protocol was high; . % of patients were observed without receiving empiric iv antibiotics. the majority of patients were discharged home and there were no serious complications or infectious deaths. the incidence of positive blood cultures was low ( . % including several likely contaminants), despite the presence of central venous catheters in the majority ( . %) of patients. there were no significant differences in age, oncologic diagnosis, central s of s line access, anc value, or incidence of bacteremia between patients who did and did not receive empiric iv antibiotics. patients who were admitted to the hospital were significantly more likely to have received iv antibiotics (p < . ) despite documentation of a reassuring exam. however, admitted patients who initially received iv antibiotics were just as likely to discharge within hours compared to patients who were observed. we propose that empiric iv antibiotic administration in febrile, non-neutropenic, otherwise well-appearing patients is unnecessary. our study demonstrated no adverse consequences of observation and no significant differences in clinical outcomes between patients who did and did not receive iv antibiotics aside from rate of hospitalization. this supports the practice of observation without empiric antibiotics for such patients. background: children with hepatoblastoma (hb) undergo repetitive computed tomography (ct) scans to determine response to treatment and assess for relapse. this imaging exposes children to radiation, anesthesia, and imposes financial and emotional burden. objectives: review our institutional experience to determine if afp measurements are sufficient to assess response to treatment and detect relapse. we conducted a retrospective chart review of all patients diagnosed with hb at our institution between - . data collected included serum afp, total number and type of imaging studies during and post treatment, and how relapse or progressive disease was detected. results: thirty-one patients were diagnosed with afp positive hb. during therapy, ct scans were performed: to assess for response to therapy or surgical planning (average scans/patient) and due to concern for progression with rising afp. off therapy, surveillance ct scans were performed (average of . scans/patient) and ( %) included the chest in patients with no lung metastasis at diagnosis. relapsed patients averaged . surveillance scans, . of which were done before relapse was noted on imaging. there were no cases of radiographic evidence of relapse without a prior increase in afp. during treatment, response to therapy based on imaging correlated with a decline in afp in all patients, arguing that repetitive scans are not needed in this setting unless required for surgical planning. only of scans performed during off therapy surveillance displayed evidence of relapse, all of which were preceded by rise in afp. our study represents the largest cohort of hb patients. prior studies suggest similar results, but included fewer patients, lower stage of disease and less than years of surveillance monitoring. at our institution, the cost of a ct c/a/p is $ , with reimbursement varying from - %. in comparison, the cost of an afp measurement is $ . . many scans also require anesthesia and result in emotional toil for families concerned about this procedure as well as the results. thus, afp demonstrates greater sensitivity, with significant cost savings and decreased emotional burden, and should be used for monitoring both during and off therapy, replacing routine serial imaging. background: we observed that our practice of drawing daily blood cultures in hospitalized patients with fever and neutropenia was wasteful; it resulted in excessive negative cultures that did not add to patient care. the smart aim of this quality improvement project was to reduce the number of negative blood cultures drawn on hospitalized patients with fever and neutropenia by % in months. design/method: after reviewing published evidence suggesting drawing daily blood cultures in febrile neutropenic patients was unnecessary, a new blood culture guideline was implemented: cultures were drawn at presentation for fever with neutropenia and, if negative at hours, repeat cultures were not drawn except for clinical change, new fever after being afebrile > hours, or antimicrobials were being changed/broadened. to impact key drivers, we educated staff and changed blood culture order sets to require providers to select a reason for ordering the culture and to eliminate a nursing order to draw daily cultures with fever. we compared the number of blood cultures drawn per central linedays (/ -cld) and the proportion of positive versus negative cultures pre-guideline (july -may ) and postguideline (june -december ). we calculated the cost savings from reducing cultures. to assess patient safety, potential septic events without a corresponding positive blood culture were reviewed. data were analyzed by service (oncology and stem cell transplant). a chi-square test was used to compare rates. in stem cell transplant patients, pre vs. postguideline, there were vs. total cultures drawn/ -cld; vs. positive ( % decrease, p = . ) and vs. negative cultures/ -cld ( % decrease, p< . ). in oncology patients, pre vs. post-guideline, there were vs. total cultures drawn/ -cld; vs. positive ( % decrease, p = . ) and vs. negative cultures/ -cld ( % decrease, p< . ). the decreased positive culture rate among oncology patients may be due to decreased culture contaminants and/or the effect of a concurrent initiative to decrease clabsi in that group. there were safety concerns; however, chart review concluded that the guideline did not lead to missed infections in these patients. for the first months of the guideline, the total cost savings in blood cultures was $ , . . the implementation of our new blood culture guideline successfully led to a substantial reduction in the collection of negative cultures and a cost savings without compromising the detection of bacteremia in hospitalized pediatric patients with fever and neutropenia. background: there are various evidence-based guidelines for treatment of adult cancers, such as the nccn guidelines. previously, care was standardized for most new diagnosis pediatric cancer patients through enrollment on a clinical trial. with decreasing clinical trial availability and enrollment and few, if any, evidence-based guidelines for pediatric cancer, care standardization is challenging for pediatric cancers. objectives: to assess consistency of care, as determined by plan of treatment by diagnosis, for pediatric patients receiving chemotherapy for newly diagnosed cancer at a single center. design/method: patients with a new cancer diagnosis at a large, tertiary care pediatric oncology center in calendar year were identified through reports from the chemotherapy order entry (coe) system. reports included diagnosis (recorded through standardized options) and the plan of treatment. chart review was used to exclude patients who started treatment elsewhere and patients being treated for relapse, to clarify diagnosis if the standardized options in coe were unclear, and to clarify treatment plan if needed. data was entered and analyzed in a redcap database. specific diagnoses were clustered into higher level disease groups and the distribution of treatment plans for patients within each was determined. this project was deemed exempt from irb approval for human subject research as a qualifying quality improvement project. of the patients with a first chemotherapy order in , were excluded due to one or more reasons: stem cell transplant ( ), transfer of care ( ), relapse ( ), and other ( ). an additional patients were excluded because < patients/year/diagnosis. there was no cns tumor disease group with > patients. thus, patients with hematologic malignancies or non-cns solid tumors are the focus of this analysis. for patients with intermediate risk rhabdomyosarcoma, the plan of treatment was the standard arm of a cog protocol, arst for patients and arst for subsequent patient after protocol activation. for all other diseases including lymphoblastic leukemia/lymphoma (excluding infants), classical hodgkin lymphoma, aml (excluding trisomy and apml), stage iii/iv burkitt lymphoma/diffuse large b-cell lymphoma, posttransplant lymphoproliferative disease, wilms tumor, rhabdomyosarcoma, ewings sarcoma, osteosarcoma, neuroblastoma, and retinoblastoma, only one treatment plan per risk category was used. conclusion: this analysis demonstrates highly consistent chemotherapy treatment at a single center for patients with hematologic malignancies and non-cns solid tumors. next steps include exploring strategies to group diagnoses for cns tumors and assessing the quality of evidence supporting the treatments given. background: rapid initiation of empiric antibiotics in patients with fever and neutropenia has been shown to reduce morbidity and mortality. current practice guidelines call for the initiation of antibiotics in these patients within sixty minutes and time-to-antibiotic (tta) has been suggested as a quality-of-care measure. many institutions, including our own, face barriers to meeting this time limit. objectives: utilizing a quality improvement model, determine barriers and implement an intervention to reduce the time-to-antibiotics for pediatric febrile patients with suspected neutropenia who present to the emergency department (ed) at our institution. we have identified and implemented an intervention utilizing the plan-do-study-act model for quality improvement. a twelve-month retrospective review was conducted to evaluate the efficacy of the current practice algorithm at our large, academic tertiary-care hospital. subjects identified were pediatric oncology patients undergoing active chemotherapy who presented to the ed with febrile neutropenia. we identified two specific barriers, triage level assignments and delay in ordering antibiotics. to address these barriers, we have created a wallet sized "fever card" that patients were instructed to show upon arrive to the ed. in collaboration with the ed staff, efforts were also made to educate all pediatric staff on the use of the fever card. post-intervention data collection is currently underway and pre-and post-intervention antibiotic delivery times will be compared. the pre-intervention cohort consisted of thirty-three encounters with a mean time-to-antibiotic delivery of minutes, or seventy-five minutes greater than the accepted standard of care. only one patient received antibiotics within sixty minutes of arrival. post-intervention data collection is currently underway. since identifying two barriers to meeting the standard of care at our institution, we have implemented a quality improvement measure that empowers patient families to direct appropriate triage in the ed as well as simplifying the treatment protocol for ed providers. we expect to identify an improvement in time-to-antibiotics from the pre-intervention to the post-intervention period. background: sickle cell disease (scd) is a genetic disorder in which sickle hemoglobin (hbs) triggers multiple downstream effects, including red cell sickling, hemolysis, vaso-occlusion, and inflammation. scd, a lifelong disease initiated at birth with injury that accumulates over time, causes significant end-organ damage and clinical complications that are undertreated and associated with early death. homozygous mutation (hbss) causes the severe form of scd. individuals with scd are at increased risk of infection, stroke, and retinopathy. clinical guidelines for pediatric patients with scd recommend prophylactic penicillin use (ages - ), annual screening for stroke with transcranial doppler (tcd) imaging (ages - ), and annual ophthalmology exams to assess for retinopathy (ages ≥ ). there are limited real-world data on implementation of these nhlbi-based recommendations. objectives: to describe utilization of penicillin, tcd screening, and ophthalmology care in children with hbss disease. medicaid administrative claims databases were used to identify us patients aged - years at first indication of hbss recorded in each calendar year from to . patients were required to have medical and pharmacy benefits for the calendar year in which they were identified and for months prior to their first recorded hbss indication. prior year utilization of penicillin, tcds, and ophthalmologist visits was measured for each annual cohort. annual cohorts included - commercial (mean age . years, % female) and - medicaid (mean age . years, % female) patients with hbss disease. fewer than half of all patients had received a tcd scan in the previous year, with similar rates seen across all age groups for both payers. ophthalmologist visits increased as patients aged, and while patients aged - years had the highest proportion with an ophthalmologist visit in both payer populations, the overall implementation remained low. in contrast to the low use of tcd and ophthalmology visits, penicillin use was highest in the - year age group: > % use in any given year for both payers. conclusion: although our data demonstrated high penicillin use in the - year age group, consistent with guidelines there is an opportunity to improve implementation of other guidelines-based recommended screening. for example, tcd screening can identify children at risk of scd-related stroke in order to initiate preventive therapies. further research to understand potential barriers to proper screening and to evaluate strategies to improve awareness, adherence, and implementation of recommended screenings in children with scd is warranted. supported by global blood therapeutics. background: childhood cancer therapy has improved where there are many long-term survivors. while psychosocial difficulties in pediatric cancer survivors are recognized, the prevalence of these problems at initial survivorship presentation is unclear. objectives: to examine the prevalence of overall internalizing symptoms (e.g., depression/anxiety) in pediatric cancer survivors presenting to a survivorship clinic and to examine how this is mitigated by receiving psychological services and by evidence of parental depression/anxiety. design/method: pediatric cancer survivors attending their first visit at the reach for survivorship clinic at vanderbilt (ages - ) were included. survivors' parents ( % female) completed the child behavior checklist (cbcl), beck depression inventory-ii, and beck anxiety inventory. survivors > years completed a self-report. the wilcoxon rank-sum and pearson's test were used for univariate analyses. the effect size and % confidence intervals (ci) estimated from the multivariable linear regressions were reported. results: childhood cancer survivors a median of years old and . years off therapy were included. thirty one survivors ( %) showed at least borderline clinical internalizing problems (t score > ) on the cbcl, but only of these patients ( %) reported receiving psychological services. nine other survivors with normal t score ≤ also reported receiving psychological services. parental depressive and anxiety symptoms were correlated to the parental report of survivor overall internalizing symptoms (spearman = . , p = < . and = . , p = < . respectively), however they were not correlated to survivor selfreports. furthermore, parents with mild to severe depressive symptoms or mild to severe anxiety symptoms were more likely to rate their child as having higher overall internalizing symptoms (p = . ; p = . , respectively). multivariable linear regression showed that when adjusted for age, gender, cancer diagnosis and time off treatment, reported utilization of psychological services ( = . , % ci [ . , . ],p = . ), and parent depressive symptoms ( = . , [. , . ],p< . ) were significantly associated with child overall internalizing symptoms. in an otherwise identical alternate model substituting parental anxiety for parental depression, parental anxiety was also a significant risk factor ( = . , [. , . ], p< . ). alternatively, parent anxiety/depressive symptoms were not significantly associated with child self-report of internalizing symptoms. childhood cancer survivors have an elevated prevalence of experiencing internalizing symptoms but seldom report receiving psychological services. childhood cancer survivors' parents with anxious/depressed symptoms are more likely to rate their children as having more internalizing problems, compared to patient self-reports. ongoing longitudinal analyses will help clarify the best timing for potential interventions. background: life expectancy for adults with sickle cell disease (scd) has remained unchanged over the past years despite improvements in pediatric scd survival. at greatest risk are the adolescents and young adults (ayas) transitioning from pediatric to adult care. allen county ranks rd in scd incidence among the counties in indiana, and has board certified pediatric hematologist-oncologists. when children "age out" of the pediatric system, there are few providers knowledgeable about managing adults with scd in the region. a novel partnership between hematologists and the family medicine residency program in allen county was initiated to educate family medicine residents (fps) about scd, hydroxyurea (hu), and management of scd-related complications with the goal to increase the number of knowledgeable providers to care for adults with scd. to determine the effectiveness of online learning modules in educating fps about hu, best practices for aya scd care and transition. three online learning modules about scd (comprehensive care of ayas with scd, hu, best practices in aya transition) were developed and cme-accredited. electronic pre-and post-tests were distributed to fps with five questions for each module covering: contraception; screening tests; hu indications, dosing and monitoring; developmental milestones and scd knowledge assessments. the st vincent irb reviewed the protocol and granted a waiver of consent. results: twenty-six fps ( %) completed the pre-and posttests. over two-thirds correctly identified the clinical benefits of hu on both assessments. knowledge about the rationale for hu therapy increased after the completion of the hu module ( % correct on pre-test vs. % on post-test, p = . ). the proportion of correct responses increased for all comprehensive aya scd care post-test questions, but only the leading cause of death and the priapism-related questions reached statistical significance ( % vs. %, p = . ; % vs. %, p = . , respectively). the proportion of correct responses for of the transition-focused questions was unchanged ( % for both), while the proportion of correct post-test responses on the self-care assessment question significantly increased ( % vs. %, p = . ). after module completion, fps were able to correctly identify common scd complications and why hu is an effective treatment for individuals with scd. the best practices of transition clinic module may need modification to improve physician understanding of the intricacies in establishing and maintaining a scd transition clinic. overall, online training is effective at educating fps and could be used to increase the number of providers knowledgeable about scd care. background: survival rates for pediatric hodgkin lymphoma (hl) exceed % with contemporary therapy. studies of pediatric hl survivors treated in the s- s have shown increased risk for treatment-related chronic health conditions. risk-adapted therapy, including tailored radiotherapy, has been developed to reduce long-term morbidity while maintaining excellent survival. little is known about chronic conditions associated with contemporary therapy presenting during the first years from therapy completion (early outcomes). objectives: to analyze survival and early outcomes of pediatric hl patients treated with contemporary therapy. we conducted a retrospective review of hl patients diagnosed < years of age at our institution from - . three-year overall (os) and event-free (efs) survival were calculated with kaplan meier statistics using sas . . results of standardized screening for targeted toxicities that developed between - years from therapy completion were identified and graded per ctcae criteria. censoring occurred at date of death, years from therapy completion, or december , . data from the last collection point were used for prevalence calculations in cases with multiple evaluations. we identified patients ( % male; % non-hispanic white; mean age at diagnosis . ± . years) with a median time since therapy completion of . years (range . - . ). initial treatment included: ( %) chemotherapy only and ( %) multimodality treatment. all patients received anthracyclines (median dose mg/m ) and % received alkylating agents (median cyclophosphamide equivalent dose [ced] mg/m ). the -year os was % with an efs of % ( % chemotherapy only, % multimodality treatment; p = . ). patients with relapsed/refractory disease received salvage treatment including chemotherapy only (n = ), multimodality therapy (n = ), or multimodality treatment including stem cell transplant (autologous n = ; autologous+allogeneic n = ). no patients developed thyroid dysfunction, cardiac dysfunction, subsequent neoplasm, or male gonadal dysfunction during the study period. pulmonary dysfunction was limited to ctcae grade . anti-mullerian hormone (amh) below the normal range was found in / pubertal females who received ced ≥ mg/m compared to / females who received ced < mg/m . two of the females with low amh also had follicle stimulating hormone > iu/ml. this study is the first to evaluate early outcomes in pediatric hl survivors. the results indicate contemporary chemotherapy and a lower rate of radiotherapy utilization lead to excellent -year survival rates with minimal early toxicities. females exposed to ced ≥ mg/m are at increased risk for gonadal dysfunction and should be prioritized for fertility preservation approaches prior to initiation of cancer therapy. background: cancer is one of the leading disease-related causes of death among individuals aged < years in the united states. recent evaluations of national trends of pediatric cancer used data from before , or covered ≤ % of the us population. objectives: this study describes pediatric cancer incidence rates and trends by using the most recent and comprehensive cancer registry data available in the us. design/method: data from us cancer statistics were used to evaluate cancer incidence rates and trends among individuals aged < years during - . data were from states and covered % of the us population. we assessed trends by calculating average annual percent change (aapc) in rates using joinpoint regression. rates and trends were stratified by sex, age, race/ethnicity, us census region, county-based economic status, and county-based rural/urban classification, and cancer type, as grouped by the international classification of childhood cancer (iccc). we identified , cases of pediatric cancer during - . the overall cancer incidence rate was . per million; incidence rates were highest for leukemia ( . ), brain tumors ( . ), and lymphoma ( . ). rates were highest among males, aged - years, non-hispanic whites, the northeast us census region, the top % of counties by economic status, and metropolitan counties. the overall pediatric cancer incidence rate increased (aapc = . , % ci, . - . ) during - and contained no joinpoints. rates increased in each stratum of sex, age, race/ethnicity (except non-hispanic american indian/alaska native), region, economic status, and rural/urban classification. rates were stable for most individual cancer types, but increased for non-hodgkin lymphomas except burkitt lymphoma (iccc group ii(b), aapc = . , % ci, . - . ), central nervous system neoplasms (group iii, aapc = . , % ci, . - . ), renal tumors (group vi, aapc = . , % ci, . - . ), hepatic tumors (group vii, aapc = . , % ci, . - . ), and thyroid carcinomas (group xi(b), aapc = . , % ci, . - . ). rates of malignant melanoma decreased (group xi(d), aapc = - . , % ci, - . -- . ). this study documents increased rates of pediatric cancer during - , in each of the demographic variables examined. increased overall rates of hepatic cancer and decreased rates of melanoma are novel findings using data since . next steps in addressing changing rates could include investigation of diagnostic and reporting standards, host biologic factors, environmental exposures, or potential interventions for reducing cancer risk. increasing pediatric cancer incidence rates may necessitate changes related to treatment and survivorship care capacity. background: while childhood cancer treatment modalities have improved, the delayed effects of cancer treatment continue to compromise the quality of life in survivors. metabolic syndrome (ms) is diagnosed based on the presence of three of the following findings -obesity, dyslipidemia, hypertension and insulin resistance per the world health organization (who) criteria. the increased risk of ms among childhood cancer survivors was first reported in the 's and is known to increase the incidence of cardiovascular disease in these individuals. objectives: assess the frequency of ms in childhood cancer survivors at our institution. . we conducted a retrospective chart review on pediatric cancer survivors, - years of age, who had been treated at sri ramachandra medical institute and research foundation between august and august . patients who received at least one year of treatment with s of s chemotherapy and/or radiation and surgery were included. medical history, family history of diabetes, cardiovascular diseases, and hypercholesterolemia, tanner staging, weight for height (< y per who criteria), bmi (> y per indian academy of pediatrics iap), blood pressure (nhlbi criteria), fasting blood sugar levels and lipid profile were obtained from the charts. statistical analysis of the data was done using ibm spss statistical software (version ). results: patients were studied, . % were male. . % were under years of age, . % between - years and . % above years. leukemia survivors comprised . % of the sample and non-leukemic's were . %. . % were treated with chemotherapy alone, . % with radiotherapy and chemotherapy, and . % underwent surgery with radiotherapy and chemotherapy. hypertension was found in . % of the study group, dyslipidemia in %, impaired fasting blood glucose in . % and . % were found to be obese. % of the study group was diagnosed with ms based on who criteria. conclusion: % of our study population was found to have ms per who criteria. individual metabolic complications were detected in % of the population. acute lymphoblastic leukemia (all) survivors appeared to be at high risk in our population. ms has been known to increase cardiovascular complications in cancer survivors. a multidisciplinary team approach to management of these patients is important to closely monitor and manage the long-term complications related to ms such as type diabetes and atherosclerosis. such an approach is essential to decrease long term morbidity and mortality from ms in this vulnerable population. background: the -year survival rate for childhood cancer exceeds %. however, up to % of these children require admission to the pediatric intensive care unit (picu) within three years of diagnosis. these children account for approximately % of all picu deaths, with mortality being higher for those post-hematopoietic stem cell transplant (hsct). national guidelines recommend that providers share informa-tion regarding prognosis and treatment options within the first hours of icu admission. these prognostic goals of care conversations (pgocc) are critical to the care of children with malignancies, a subpopulation at risk for increased mortality. to determine the frequency of pgocc as well as describe differences in patient characteristics and critical care therapies by pgocc status. design/method: a retrospective cohort study was conducted using the university of michigan virtual picu system database. picu admissions lasting longer than hours for patients ages to years between july , and june , with an oncologic diagnosis and/or hsct were identified. data on pgocc, patient demographics, diagnoses, picu interventions, and outcomes were recorded and compared between children with pgocc and those without using chi square test for categorical variables and kruskal-wallis test for continuous data. of picu admissions, % were male; the mean age was . years. the leading diagnoses were acute lymphoblastic leukemia ( %), acute myeloid leukemia ( %), lymphoma ( %), neuroblastoma ( %), and brain tumors ( %), and % of patients were post-hsct. pgocc was documented in ( %) patients. in comparison with patients who did not have a pgocc, children with a pgocc were more likely to be readmitted to the picu ( % vs. %, p < . ) and more likely to have had relapse of disease ( % vs. %, p< . ). patients with a pgocc had higher severity of illness scores (p = . ), higher use of non-invasive ( . % vs. . %, p = . ) and invasive conventional ventilation ( . % vs. . %, p< . ), and high frequency ventilation ( . % vs. . %, p < . ). also, patients with pgocc were more likely to receive continuous renal replacement therapy ( . % vs. . %, p< . ), arterial catheterization ( . % vs. . %, p< . ), and cardiopulmonary resuscitation ( . % vs. . %, p< . ). in only in critically ill children with hematologic-oncologic disease is pgocc held. children with pgocc were sicker and received more critical care interventions. future research is needed to evaluate the content of pgocc. background: central nervous system (cns) tumors and autism spectrum disorder (asd) represent significant disease cohorts in the pediatric population. asd diagnoses in children have a prevalence of %, in every children in the united states. additionally, more than , cns tumors are reported in children age to years in the united states with brain tumors being the most common solid tumor and the leading cause of death among all childhood cancers. the genetic etiology of autism and cns tumors is complex. specific gene alterations present in certain cancers have similarly been described and suspected to play a role in asd subtypes. targeted therapy panels, like foundation one (fo), have been beneficial in guiding treatment for some cancers based on distinct gene alterations. given the genetic overlap, the potential for therapeutic benefit and crossover from such actionable gene target panels merit further exploration in asd and cns tumors. we aim to identify and describe genetic alterations with known actionable targets in cancer therapy from fo as potential diagnostic, therapeutic and research targets for neurodevelopmental diseases. we plan to discuss the common genetic alterations between our cancers and neurodevelopmental diseases described in the literature. fo data was extracted and compared to the literature. each reported gene alteration from fo plus the keywords "autism", "psych" were used on pubmed to search for a suspected association if any with a neurodevelopmental disorder. results: twenty-one patients representing a cohort of six unique (astrocytoma-five, ependymoma-six, gbm-four, glioma-three, nerve sheath tumor-one, etmr-two) cns tumors were investigated. fo produced eighty total with sixty unique gene alterations. thirty-one ( %) of these yielded at least one published, suspected association to a neurodevelopmental disorder. the most common gene alterations were tp -four, cdkn a/b-five and braf-four. the main functional categories were cellular: proliferation, structure, differentiation and degradation; chromatin modeling; histone transcriptional modification; dna methylation and repair; strna; and neural signaling. sixty unique gene alterations were found in our cns tumor set using foundation one. thirty-one ( %) of these discrete alterations paired with at least one description in the literature as having been similarly altered in an asd subtype. many of these alterations have actionable targeted therapies presented through foundation one for our cns tumors and may be a relevant guide in the future of targeted therapy and research in asd subtypes. monoclonal antibody therapy usage is associated with significantly improved survival in b-cell nhl aya patients. although the usage has increased in the aya population from to , the magnitude of the increase is low. factors that affect the use of mab include race and insurance s of s type. further research is warranted to identify why privately insured patients are less likely to receive these drugs. background: prevention of chemotherapy-induced nausea and vomiting (cinv) remains a challenge despite advances in pharmacotherapy and the development of cinv clinical practice guidelines by the pediatric oncology group of ontario (pogo) that have been endorsed by the children's oncology group. achieving control of cinv in pediatrics further is complicated by the difficulty young children have vocalizing their symptoms. use of a validated nausea-assessment tool in conjunction with improved adherence to evidence-based guidelines may result in better quantification of symptoms and reduction of both nausea severity and vomiting frequency for pediatric patients undergoing chemotherapy. the pediatric nausea assessment tool (penat) has been validated for children ages - , and its integration into clinical practice may help optimize cinv control. objectives: this single-institution study sought to improve control of cinv in patients admitted for chemotherapy by standardizing the antiemetic regimens prescribed by all providers according to an institutional cinv algorithm developed from the pogo guidelines. we hypothesized that treatment using a standardized guideline would improve cinv control in patients admitted for chemotherapy. a baseline cohort of admissions for chemotherapy completed penat assessments and cinv diaries prior to receiving chemotherapy, four times daily during each admission, and daily for days following completion of chemotherapy from may , to january , . providers then were provided an institutional cinv treatment algorithm based on the pogo guidelines and received education at departmental meetings on appropriate implementation of this algorithm. a second cohort of admissions completed penat assessments and cinv diaries in a similar fashion from july , to december , . results: complete control of vomiting markedly improved following cinv guideline implementation ( % vs %, p <. ) with treatment failure also significantly reduced ( % vs %, p <. ). after controlling for the degree of emetogenicity of chemotherapy received, a patient was . times more likely to vomit prior to guideline implementation (or . , ci . - . ). there was no difference in nausea control, even after adjusting for the emetogenicity of chemotherapy. conclusion: control of chemotherapy-induced vomiting (civ) improved following widespread implementation of an institutional cinv treatment algorithm at a single institution. the severity of nausea reported remained unchanged which may reflect the difficulty of assessing nausea or an inadequate sample size. future research may focus on cinv treatment management through the use of guidelines specifically for breakthrough cinv and delayed cinv. background: aspho's professional development committee (pdc) recognized pediatric hematologists-oncologists (phos) serving in the united states (us) military have unique professional development needs that may not be addressed by aspho or a similar professional society. these individuals may also encounter challenges when transitioning to a civilian career. however, barriers to professional development have not been systematically characterized. the objectives were to characterize the number of phos with current or prior military service (mphos) and to identify any unmet professional development needs. design/method: a working group consisting of pdc members and both senior and early career mphos was formed. initial comments were solicited by email from known mphos regarding potential gaps in professional development and interest in working with aspho to improve support of mphos. a survey was developed and piloted with four members of the advisory group, questions were revised based on their feedback, and a final version was distributed via the aspho website and online community forum. targeted emails were sent to mphos identified through aspho and military databases. eligibility to complete the survey included ) completion of a fellowship in pediatric hematologyoncology, and ) current or prior service as an active duty military provider. quantitative and qualitative information were collected, including demographic data and perceived barriers to professional development. responses were summarized using descriptive statistics. results: sixty-five mphos were identified and surveys were completed for a % response rate. respondents were engaged in a variety of professional activities; % were male, % were serving active duty commitments, and % felt there were professional development gaps. areas of concern were categorized into nine themes with the most concerning being ) limited civilian knowledge of mpho practices ( % of participants), ) inability to attend professional society meetings ( %), and possibility of deployment ( %). participants expressed a desire for educational products to meet their specific needs and for networking opportunities with civilian colleagues. qualitative analyses identified concerns about low patient numbers and practice size. a subset of mphos perceive significant gaps in professional development. additional research is needed to better define areas for intervention, but many of the concerns align with those of similarly sized civilian programs and may be addressed through professional society networking opportunities, such as an aspho special interest group. background: infertility is an established cause of distress and has a negative impact on quality of life among childhood cancer survivors. the american society of clinical oncology has established guidelines on fertility counseling for individuals of reproductive age diagnosed with cancer, with the goal of improving reproductive and psychosocial outcomes. studies have shown that instituting a fertility team that can provide counseling and discuss fertility preservation (fp) options results in improved patient satisfaction in patients with cancer. objectives: the goal of this study was to examine predictors of referrals to the multidisciplinary fertility team, and documented fp interventions among these patients. design/method: an irb-approved retrospective medical record review was performed at a large pediatric academic center. all patients with new cancer diagnoses receiving chemotherapy were included from january (when the fertility team was established) to present. a standardized abstraction form was used to collect information about: age at diagnosis, gender, cancer type, whether a fertility consult was placed, and documented fp interventions. data were summarized descriptively and comparisons were made using nonparametric statistical methods. results: patients met inclusion criteria, of which ( %) were male. cancer types were as follows: leukemia/lymphoma, cns tumors, sarcomas, embryonal tumors, and langerhan's cell histiocytosis (lch). the mean age was . years, (range < - years). overall, % of all patients had a consultation with the fertility team. patients were significantly less likely to have a fertility consult if they were younger (p< . ). further, there were differences in the consultation rate between diagnoses, with % of sarcoma patients completing a consult, compared to % of those with cns tumors, % of those with embryonal tumor, % of those with leukemia/lymphoma and none of the patients with lch. our findings show that many children, adolescents, and young adults newly diagnosed with cancer are still not receiving fertility counseling despite: ) an expanding body of literature supporting the need to provide this counseling, ) guidelines published by several organizations recommending discussions about infertility risk and fp options, and ) presence of a multidisciplinary fertility team. specific strategies need to be developed to improve access for younger children, and for disease groups in whom fertility consults are underutilized, such as youth with cns tumors, embryonal tumors, and leukemia/lymphoma. background: socioeconomic status (ses) has on impact on overall survival in the pediatric oncology population. unfortunately, data are insufficiently detailed to explain the mechanism behind this phenomenon. how parents handle the health management demands placed on them at the time of a child's cancer diagnosis may represent a point of differentiation in health outcomes. objectives: determine the association between socioeconomic factors, cancer literacy, and parents' understanding of home emergency management and their responses to instances of pain, nausea, and fever. in a prospective observational study of parents whose children were newly diagnosed with cancer, we obtained demographic information and, using a validated instrument, (dumenci, ) we evaluated cancer literacy. we tested understanding of the education parents received about home emergency management with a -item multiple-choice vignette-based questionnaire focused on actions needed in home scenarios. we then followed parents' actual behavior through periodic phone calls assessing instances of nausea, pain, and fever and their responses to these episodes. results: preliminary analysis of participants showed an average score of on the -item parental understanding questionnaire (range - ). variables associated with increased score were college-level education by . points ( % ci [. to . ]), private insurance by . points [. to . ] and adequate cancer literacy by . points [. to . ]. actual behavior reported by families indicated that married parents and those with income above $ , were less likely to treat instances of pain by % ( % ci [ to ]) and % [ . to ], respectively. white parents, those with college-level education, and those with adequate cancer literacy were less likely to treat instances of nausea by % [ to ], % [ to ] and % [ to ], respectively. no associations were found between socioeconomic markers and parental responses to instances of fever. our findings suggest an association between demographic and socioeconomic markers and improved parental understanding of home emergency management. paradoxically, the same markers show a decrease in treatment response to pain and nausea. larger prospective studies are needed to link this behavior pattern to health outcomes, and help inform the extent of ses impact on home emergency management. emory university/children's heathcare of atlanta, atlanta, georgia, united states background: cardiovascular disease is a leading cause of morbidity and mortality in childhood cancer survivors (ccs). previous research showed wide practice variation in referral patterns to cardiology from the survivor clinic and in recommendations from cardiologists about the need for further testing or exercise restrictions. to develop a cardio-oncology algorithm in order to standardize referrals to cardiology and provide guidelines for cardiologists evaluating pediatric ccs. design/method: survivorship and cardiology experts developed a weighted scoring system for pediatric ccs who received cardiotoxic therapy based on time since treatment and risk factors identified by the children's oncology group (cog) and american heart association (aha). the cardiooncology algorithm assigned a score of - . the score range was categorized to guide cardiology referral: screening echo only ( - ), consider cardiology referral ( - ), recommend cardiology referral ( - ), and regular cardiology follow-up (≥ ). the algorithm also provides recommendations to cardiologists for screening and exercise modifications based on the score. after establishment of the algorithm, a convenience sample of institutional survivor clinic patient charts were retrospectively reviewed from the first month of each quarter from april -march to validate the algorithm, evaluate referral patterns to cardiology, and assess cardiology recommendations. the retrospective chart review evaluated patients ( % male; % non-hispanic white; % leukemia survivors; median age at diagnosis . years [range - . ]; median time off-therapy . years [range . - . ]). patients ( %) received anthracyclines (median dose mg/m , range - ) and ( %) received cardiac radiation. assigned cardio-oncology scores resulted in: % echo only, % consider cardiology referral, % recommend cardiology referral, and % regular cardiology followup. when evaluating detection rates of late effects by cardiooncology score, survivors ( %) had an abnormal echo: / echo only, / consider referral, / recommend referral, and / regular cardiology follow-up. assessing referral patterns prior to initiation of the algorithm revealed forty-two survivors ( %) referred to cardiology: / echo only, / consider referral, / recommend referral, and / regular cardiology follow-up. of the patients seen by a cardiologist at our institution, had further diagnostic testing ordered (i.e., stress test) and received exercise restrictions. a cardio-oncology algorithm and guidelines will standardize cardiac care for survivors by assigning a score to guide referral and cardiology practice after referral. prospective clinical use has begun and review will occur in one year to determine changes in detection rates of cardiac late effects, referrals, and recommendations from cardiologists. oregon health and science university, portland, oregon, united states background: delirium affects - % of patients (pts) in pediatric intensive care units (picu) and is associated with increased length of stay, decreased attention in school, and post-traumatic stress disorder. the diagnostic and statistical manual of mental disorders (dsm v) defines delirium as a "disturbance of consciousness […] with reduced ability to focus, sustain or shift attention" due to an underlying medical condition. despite the medical complexity of the hospitalized pho population, there are no published prospective studies looking at delirium in these pts. hypothesizing that delirium is under recognized in the pho population, we designed a year-long prospective study using a validated screening tool to determine the frequency of delirium in hospitalized pho pts and to identify associated clinical factors. design/method: baseline frequency of pts with symptoms suggestive of delirium was determined through retrospective chart review using a data mining program of electronic medical records (emr). for the prospective study, pho and picu nurses were trained to use the cornell assessment for pediatric delirium and to record scores within the emr on all pho pts once every -hour shift. predetermined demographic and clinical variables were entered daily into a red-cap database on all hospitalized pho pts. results: baseline frequency of delirium, without active screening, was determined to be . % of hospitalized pho pts. in the first months of the prospective study, consecutive admissions occurred among unique pho pts: oncology, hematology, and stem cell transplant pts. pts had at least positive delirium screen, for a prevalence per admission of . %. statistically significant variables associated with delirium, at p < . by univariate logistical regression, included prolonged length of stay, pt location (picu vs pho unit), and fever. adjusting for length of stay, administration of benzodiazepines and opiates were also significantly associated with delirium, p = . and . , respectively. on average, nurses completed delirium screening in % of each pts' -hour shifts. study accrual ends in jan and final data analyses will be reported in the abstract presentation. conclusion: delirium does occur in the pho hospitalized population and screening by trained nursing staff is feasible. pts at highest risk appear to be pts with prolonged hospital stays, picu admissions, or frequent use of benzodiazepines/opioids. routine screening should improve our recognition of delirium and allow us to promptly intervene, or prevent delirium in an effort to avoid potential acute and long term consequences. background: with high survival rates for children and adolescents with hodgkin lymphoma (hl), treatment regimens are now designed to maximize cure while decreasing risk of long-term health outcomes associated with chemotherapy and radiation therapy. within contemporary treatment regimens, the comparison of toxicities experienced by patients receiving chemotherapy plus radiotherapy (crt) versus only chemotherapy (co) has not been studied extensively. objectives: this study examines select self-reported adverse health outcomes in survivors of contemporarily-treated pediatric hl to better understand the balance between efficacy and toxicity associated with chemotherapy and radiation therapy. (cog) ahod that evaluated a response-based treatment paradigm in pediatric hl. patient who received initial chemotherapy were randomized based on early response to continued chemotherapy, chemotherapy plus radiotherapy or augmented chemotherapy plus radiotherapy. patients completed self-report questionnaires on health problems at , , , and years following therapy. we examined selected patient-reported pulmonary, gastrointestinal (gi), cardiac and endocrine outcomes. kaplan-meier survival curves were used to determine probability of survival without the selected adverse health outcome. log-rank tests were used to compare the co versus the crt group. results: a total of , enrolled patients, patients in the co group and patients in the crt group, completed , questionnaires at a median of . years after s of s completion of therapy (q , q : . , . ) which were analyzed. the cumulative -year incidence of endocrine dysfunction was significantly greater in the crt group versus those in the co group ( % versus %; p< . ), driven by the incidence of hypothyroidism ( % versus %; p< . ). there were no significant differences in cardiac ( % versus %; p = . ), pulmonary ( % versus % p = . ), and gastrointestinal dysfunction ( % versus %; p = . ) between the co and crt patients. conclusion: this study demonstrates low cumulative incidence overall of organ dysfunction early post completion of contemporary therapy for hl. the addition of radiation therapy significantly increased risk for hypothyroidism, but with no higher risk noted for cardiac, pulmonary or gi dysfunction. limitations include self-report status, potential selection bias, and relatively short latency period following end of therapy. longer follow-up is needed to determine more delayed risks for organ dysfunction in order to best define the balance between therapeutic efficacy and long-term adverse health outcomes related to chemotherapy and/or radiation therapy. background: identification of an organism via bronchoalveolar lavage (bal) or respiratory tract biopsy (rtb) has historically been considered the gold standard for diagnosis of invasive fungal infection (ifi); however, data previously published by our group showed that these procedures infrequently lead to a change in management in children with an oncological diagnosis or undergoing hematopoietic stem cell transplant (hsct). there is also a paucity of data on the cost of ifi in this population. to compare the costs of work-up and management of pulmonary ifi diagnosed based on ct scan alone versus ct scan or chest x-ray prompting a bal or rtb. design/method: we collected cost data on patients at ann & robert h. lurie children's hospital of chicago undergoing chemotherapy or within months of hsct who were suspected of having an ifi between and . in order to include sufficient time to account for post-procedure compli-cations but avoid including costs unrelated to ifi, data were included for days from the day of their diagnostic scan or procedure. cost data was available for of the patients previously studied. thirty-six of these patients were diagnosed with suspected ifi based on ct only and patients underwent bal or rtb. when evaluating specific costs, inpatient beds costs were higher in the bal and rtb group (median $ , versus $ , , p = . ), yet there was only a trend towards higher costs for antifungal agents (median $ , versus $ , , p = . ) and respiratory support (median $ versus $ , p = . ). many of the initial ct scans were not captured in the -day evaluation period for the bal or rtb group based on the study design; however, even when accounting for ct scans up to a week prior these procedures, the total cost of ct scans was higher in the ct only group (median $ versus $ , p = . ), as they had more scans. despite this, total costs were significantly higher for patients who underwent bal or rtb versus ct scan only (median $ , versus $ , , p < . ). combined with our previous data that bal and rtb infrequently leads to a change in management in children with an oncological diagnosis or undergoing hsct suspected to have an ifi, the significantly higher costs associated with these procedures makes these invasive diagnostic techniques even less desirable. batra, pediatr blood cancer, . background: while infants > months of age with acute lymphoblastic leukemia (all) have a poor prognosis, infants with acute myeloid leukemia (aml) fare better despite more intensive therapy. there are limited data on this difference, particularly differences in supportive care requirements during induction therapy for infants. objectives: to compare induction mortality and resource utilization in infants relative to non-infants aged < years, separately for all and aml. design/method: we used previously established cohorts of children treated for new onset all or aml at children's hospitals in the us contributing to the pediatric health information system. patients with down syndrome were excluded. follow-up started on the first day of induction chemotherapy and continued until the earliest of: days after commencement of chemotherapy, start of the subsequent course, or death. high acuity of presentation, defined as icu requirements involving or more organ systems within the first hours following initial admission were compared using log binomial regression. -day inpatient mortality was compared using cox regression. resource utilization rates (days of use per inpatient days) were compared using poisson regression. results: a total of all ( infants, non-infants) and aml ( infants, non-infants) were included in the analyses. infants were more likely to present with high acuity compared to non-infants for both all ( % and %, rr = . , % ci: . , . ; p< . ) and aml ( % vs %; rr = . , % ci: . , . ; p = . ). infants with all had higher inpatient mortality compared to non-infants even after accounting for differences in acuity of presentation ( . % vs . %, adjusted hr = . % ci: . , . ; p = . ). in contrast, inpatient mortality was more similar for infants and noninfants with aml ( . % vs . %, adjusted hr = . % ci: . , . ; p = . ) and comparable to rates among infants with all. infants with all and aml had higher rates of utilization of fresh frozen plasma, cryoprecipitate, diuretics, supplemental oxygen, and ventilation relative to non-infants. infants with all also had higher rates of total parenteral nutrition, ecmo, and patient controlled analgesics compared to noninfants. infants with all experienced significantly higher induction mortality compared to noninfants, a difference not entirely explained by acuity at presentation. differences in ru among infants may reflect higher presentation acuity and greater treatment related toxicity. further work is needed to elucidate the contribution of treatment related toxicity to early mortality in infants with all. background: fever in a child with cancer is a medical emergency due to the significant risk of a serious bacterial infection. many attempts have been made to risk stratify these patients. the respiratory pathogen panel (rpp) is a panel of polymerase chain reaction tests that identify seventeen common respiratory viruses and three bacterial infections. samples are taken via nasopharyngeal swab. rpps are frequently sent, but we do not have data to determine whether a positive result can lead to stratification to a lower risk of bacterial infection. ( ) to determine the epidemiology of respiratory virus-associated fever in pediatric oncology patients ( ) to determine whether a positive rpp is associated with reduced risk of bacteremia in this population. this was a single-center, retrospective cohort study. we identified and reviewed the medical records of all pediatric oncology patients seen in our emergency department (ed) with fever from the introduction of the rpp in april to september , . we reviewed the results of blood cultures, rpp, chest radiographs, and discharge summaries to identify sources of infection. we also identified the patients' cancer diagnosis, age, absolute neutrophil count (anc), and absolute lymphocyte count (alc). results: positive rpps were found among pediatric oncology patients who presented to the ed with fever. the most common positive rpp findings were rhinovirus/enterovirus (rev) ( %), parainfluenza ( %), influenza ( %), coronavirus ( %), and polyviral ( %). among patients with a positive rpp, % had bacteremia compared to % bacteremia among all pediatric oncology patients with fever (or . [ . - . ], p . ). all cases of bacteremia were associated with rev. there was no bacteremia identified in patients with rpps positive for other viruses (or . [ . - . ], p . ). rev positivity did not confer a lower risk of bacteremia than rpp negative patients ], p . ). anc (p = . ) and alc (p = . ) less than , and number of patients with severe neutropenia (p = . ) were not statistically different between the rev and non-rev positive rpp groups. rpps positive for viruses other than rev reduced the likelihood of bacteremia in febrile pediatric oncology patients in the ed setting. patients with bacteremia may have concurrent infection with rev. a larger study is warranted to determine if positive rpp results can inform clinical management of a child with febrile neutropenia. emily mueller, anneli cochrane, seethal jacob, aaron carroll s of s background: the usage of mobile health (mhealth), which refers to the application of mobile or wireless communication technologies to health and healthcare, has grown exponentially in recent years. mhealth tools have been used by caregivers of other vulnerable populations, but little has been focused on caregivers of children with cancer. objectives: to conduct a survey to understand the mobile technology usage, barriers, and desired mhealth tools by caregivers of children with cancer. we conducted a mailed cross-sectional paper survey of caregivers of all children who were diagnosed with cancer at riley hospital for children between june, and june, . the survey contained questions, both fixed and open-ended, in both english and spanish. up to three rounds of surveys were sent to those who did not respond. of the respondents, they were primarily parents ( . %), median age was . years (range - ), and most were white ( . %) and non-hispanic/latino ( . %). the top three annual household income brackets included $ , to $ , ( . %), $ , to $ , ( . %) and under $ , ( . %). the majority had an education: . % college graduates, % graduate degree, and . % high school education or ged. nearly all respondents owned a smart phone ( . %) and . % owned a tablet. the majority used an ios operating system ( . %), while . % reported use of a device with an android operating system. all caregivers reported use of at least one mobile website/app regularly for their personal use. while . % of respondents reported no barriers to mobile technology use, the top barrier selected was "data limitations" ( . %). overall, . % wanted at least one medical managementrelated website/app: medical knowledge ( . %), healthcare symptom tracking/management ( . %), and medication reminders ( %). healthcare system-related desires were high, as . % wanted access to their child's medical record and . % wanted a website/app to facilitate better communication with medical providers. there were no significant associations between socioeconomic status (income or education) with barriers or types of websites/apps desired by caregivers. since the vast majority of caregivers use mobile technology with minimal barriers, future research should focus on designing an mhealth tool to address the medical management needs by caregivers of children with cancer. by supporting caregivers through this type of mhealth tool, it could positively impact patient clinical outcomes through greater adherence to medications and treatment protocols. background: in children with fever and neutropenia, early initiation of targeted antibiotic therapy improves outcomes, yet there are no standards for choice of empiric antibiotics. in our institution implemented an early empiric ceftriaxone (eec) protocol to reduce time to antibiotic administration in febrile hematology-oncology patients who are potentially neutropenic when the absolute neutrophil count is not yet know. ceftriaxone is given immediately after obtaining blood for culture and lab studies. in patients found to be neutropenic, ceftriaxone is discontinued and cefepime is initiated. the purpose of this retrospective study was to evaluate our eec protocol in neutropenic patients by assessing ceftriaxone sensitivity of positive blood cultures and comparing rates of adverse outcomes with a cohort of patients treated prior to implementation of the protocol. we are now conducting a prospective study to more thoroughly investigate antibiotic sensitivities of organisms isolated from blood cultures of neutropenic patients. design/method: hematology-oncology patients with at least one positive blood culture between january and december were identified. patient demographics, neutrophil count, antibiotic treatment, isolated organisms and sensitivities, and adverse outcomes (increased respiratory support, hypotension requiring intervention, and icu admission) were obtained by retrospective chart review. fisher exact test was used to compare dichotomous variables between patient groups. we are now prospectively identifying febrile neutropenic patients with positive blood cultures and performing antibiotic sensitivity testing to several antibiotics commonly used as empiric therapy for febrile neutropenia. results: retrospectively, we identified neutropenic patients with a total of bacterial isolates from blood cultures. of organisms isolated, were tested for sensitivity to ceftriaxone and ( %) were not sensitive, / ( %) of gram-positive cultures and / ( %) of gram-negative cultures. ten of ( %) eec patients had an adverse outcome versus / ( %) of non-eec patients (p = . ). notably, % of eec patients required icu admission versus % of non-eec patients (p = . ). thus far our data obtained prospectively is revealing similar rates of ceftriaxone resistance with / cultures not sensitive to ceftriaxone ( %, ci . %- . %). in our retrospective study, no statistically significant difference was seen in overall adverse outcome rate between the two cohorts, though icu admission rates were significantly higher in eec patients. ceftriaxone resistance rates were high in tested isolates, which is further supported by preliminary data from our ongoing prospective study. given these data, eec may not be effective at improving outcomes in febrile neutropenic pediatric hematology-oncology patients. background: approximately in children diagnosed with cancer will die of their disease, despite advances in treatment. results: two focus groups of six parents each met in june . the parents were predominantly female ( female, male) and had lost their children an average of . years prior (range - . years). two parents were in the same family. nearly all patients were offered palliative care ( / ), all were offered hospice and most died at home ( at home, in the icu). parent discussion uncovered six broad themes: beneficial provider qualities, optimal communication, helpful systematic supports, struggles to feel like a good parent, struggles with a loss of control and unmet needs. parents appreciated providers who were consistent, reliable and honest. parents desired communication that was sensitive to the needs of the patient and family with a balance of hope and realism. parents appreciated the tangible supports pro-vided by social work and the emotional support of child life both for the patient and their siblings. some parents struggled to define and advocate for their child's quality of life, especially when it led to disagreeing with the medical team. several parents expressed frustration with unfamiliar caregivers in the hospital, especially trainees. they expressed a strong desire for more anticipatory guidance about the end of life including how to discuss it with their children. they also wished for a cancer-specific support group for bereaved parents. conclusion: bereaved parents of pediatric oncology patients in our focus groups appreciated consistent, reliable providers who communicated with a balance of realism and hope. they appreciated the tangible and emotional support they received and wanted more anticipatory guidance at the end of their child's life. these results can help guide clinical care, especially in communities without strong palliative care support. further research is needed to develop interventions to improve end of life care. background: clinical trials involving human subjects depend on informed consent (ic) to ensure ethical protections for participants. parents of children with cancer often lack full understanding of the basic elements of ic for clinical trials. additionally, the stress of their child's cancer diagnosis may affect their decision-making capabilities. this is especially problematic as these children rely on parents to fully comprehend clinical trials and weigh their benefits and risks. physician communication is critical for effective family-centered care. the acgme mandates that training programs teach and assess trainees' communication skills. however, there are currently no published curricula aimed at training pediatric hematology/oncology fellows to deliver ic effectively for cancer clinical trials. to develop and pilot-test a simulation-based curriculum to enhance communication skills of pediatric s of s hematology/oncology fellows in the delivery of ic for cancer clinical trials. we developed, tested, and implemented the curriculum from to in two phases. in phase- , we reviewed literature on simulation-based curricula and completed a needs assessment to create a clinical scenario and full curriculum using standardized patients. using miller's pyramid model, fellows' assessments included: immediate de-brief, surveys to assess pre/post confidence and knowledge of the basic ic elements ("knows" and "knows how"), and -degree summative assessments compiled from fellow self-assessments, faculty, and standardized patients ("shows how"). after initial testing and refinements done with fellow, in phase- , we implemented the curriculum with our fellows. likert scale ( strongly disagree- strongly agree) and basic p values are reported. results: fellows gave high mean ratings for training relevance ( . ) and standardized patients' preparedness ( ). almost all ( . ) reported they have used the knowledge gained in their clinical practice. increase in self-reported confidence (pre/post) was noted in all domains: general -describing possible benefits of the clinical trial . / vs. . / (p = . ), risks and potential side effects . / vs. . / (p = . ), and explaining alternatives . / vs. . / (p = . ); research -discussing purpose of the clinical trial . / vs. . / (p = . ), and randomization . / vs. . / (p = . ); and family-centered -addressing emotions during ic . / vs. . / (p = . ), and delivering bad news . / vs. . / (p = . ). summative evaluation mean ratings for all fellows were . (range . - . ). our novel simulated-based ic curriculum, significantly increased fellows' self-reported confidence and skills during ic delivery. importantly, our ic curriculum addressed not just research-related content but also management of parental emotional needs during the ic discussion. next phase includes kirkpatrick model program evaluation and dissemination across other training programs in our institution. national kaohsiung normal university, kaohsiung, taiwan, province of china background: taiwan's childhood cancer foundation reported in that the -year survival rate of childhood cancer was %. as a result, many childhood cancer survivors were back in school after treatment. however, childhood cancer survivors' educational outcomes suffered because of their long-term absence from school and late effects of cancer and cancer treatment. a few school reentry protocols have been developed by the nursing professionals in taiwan to facilitate students' return to school but remained experimental in nature and hardly accessible. parents, students, and teachers were left to their own devices to make individual school reentry plans. objectives: this study aimed to examine and uncover the commonalities among three middle school students' successful school reentry experiences from their teachers' perspectives and to analyze the factors contributing to their success. design/method: this is a qualitative interview study. indepth semi-structured interviews were conducted with three middle school teachers in december about their perceptions, observations, and experiences working with adolescent childhood cancer survivors. the students were two boys with leukemia and one girl with bone cancer. they were diagnosed in the first year of middle school when they were - years old and returned to school for the third and the final year. these students met the following criteria for successful school reentry: regular school attendance, average/above average academic performance, friendship maintenance, and high school diploma. the theme -bring the class to the hospital was found to be the key to the adolescents' successful return to school. without a prescribed school reentry protocol and in the face of limited bedside education services, the homeroom teachers, as links between school, home, and hospital, brought the class to their hospitalized students. they doubled as bedside teachers conducting lessons at the hospital or students' homes, became friends with the parents, witnessed firsthand the students' pain and triumph during treatment, brought the students back to school for visits and celebrations, delivered the classmates' wishes and news to the students, encouraged and welcomed classmates' visits to the hospital, and, together with parents and other teachers, developed flexible school reentry schedules for the students. this on-going study demonstrated the critical roles and functions of homeroom teachers in successfully bringing the students back to school during and/or after cancer treatment. further analysis will be focused on how and why these three homeroom teachers were able to carry out this unexpected task on top of their already full workload. jennifer kesselheim, shicheng weng, victoria allen, collaborative group fellowship program directors dana-farber/boston children's cancer and blood disorders center, boston, massachusetts, united states background: a novel, -module, case-based curriculum entitled "humanism and professionalism for pediatric hematology-oncology" (hp-pho) aims to foster pho fellows' reflection on grief and loss, competing demands of fellowship, difficult relationships with patients and families, and physician well-being and burnout. in small group facilitated sessions, fellows work to identify coping strategies and explore how the challenges of fellowship influence both their own doctoring and the patient experience. objectives: to administer the hp-pho curriculum in a prospective, cluster-randomized trial, measuring whether exposure to this educational intervention, compared to standard conditions, fosters humanism and professionalism and improves satisfaction with training. design/method: pho fellowship programs (n = ) were cluster-randomized to deliver usual training in humanism and professionalism (control) or the novel curriculum (intervention) during the - academic year. the primary outcome measure was the pediatric hematology-oncology self-assessment in humanism (phosah). secondary measures included a -point satisfaction scale, the maslach burnout inventory (mbi), the patient-provider orientation scale, and the empowerment at work scale. participating fellows were pre-tested in summer and post-tested in spring . a change score was calculated for each study instrument. we compared each outcome between arms using mixed effect models adjusted for pre-test score as a fixed effect and site as a random effect. results: randomization yielded intervention and control fellows. the two arms did not significantly differ in distribution of fellow age, gender, or post-graduate year. the intervention sites successfully administered of ( %) modules. change scores on the phosah were not significantly different between the control and intervention arms (adjusted mean difference = . ; % confidence interval [ci] - . , . ; p = . ). compared to the control arm, fellows' exposed to the curriculum gave significantly higher ratings on several items within the satisfaction scale including satisfaction with their training on "physician burnout" (adjusted mean difference = . ; % ci . , . ; p< . ), "physician depression" (adjusted mean difference = . ; % ci . , . ; p< . ), "balancing professional duties and personal life" (adjusted mean difference = . ; % ci . , . ; p = . ), and "humanism overall" (adjusted mean difference = . ; % ci . , . ; p = . ). change scores on other secondary measures were not significantly different between study arms. conclusion: exposure to the hp-pho curriculum did not alter fellows' self-assessed humanism and professionalism. however, the curriculum proved feasible to administer and intervention fellows expressed higher levels of satisfaction in their humanism training, indicating the curriculum's positive impact both for fellows and their learning environment. background: recent work has documented significant levels of unmet needs among adolescents and young adults with cancer, particularly psychosocial challenges during the transition to adulthood, (e.g., abrupt disruption to school and social life, and social isolation). given that adolescents and young adults drive mobile app use, a mobile-phone may be an ideal way to deliver a psychosocial intervention to adolescents and young adults with cancer. to use a patient-centered approach to inform a mobile-based mindfulness and social support intervention for adolescent and young adult patients with cancer. design/method: participants were ten aya with sarcoma ( % female; % adolescents); parents of the five adolescents, and six healthcare providers (n = ). formative research involved three steps: ( ) in-depth interviews were conducted with ten aya with sarcoma; parents of the five adolescents, and six healthcare providers (n = ). ( ) adaptations were made to an existing mindfulness app which offers a program for youth. modifications included creating a -week "mindfulness for resilience in illness" program, with relaxation exercises, and the addition of videos featuring two sarcoma survivors as program hosts. content was informed by the mindfulness curriculum for adolescents, learning to breathe. ( ) a private facebook usability group was organized to (i) elicit beliefs about the mindfulness app and potential future enhancements, and (ii) promote social support. results of the in-depth interviews revealed themes around adolescents' functioning and coping, including body image concerns; recurrence-related anxiety; anger over loss; and being overwhelmed by medical information. themes from the interviews were incorporated into a demonstration version of the mobile app. a patient-centered approach is widely recommended in the development of mobile-based health behavior change interventions and may be a useful way to inform development of a mobile-based mindfulness and social support intervention for adolescents and young adults with cancer. background: medical trainees consistently report suboptimal instruction and poor self-confidence in communication skills. despite these deficits, few training programs provide comprehensive pediatric-specific communication education, particularly in the provision of "bad news." an in-depth survey to examine the historical experience and communication needs of pediatric fellows was conducted at a large academic pediatric center as the first step towards the development of a comprehensive communication curriculum. to determine the previous educational and clinical experiences of pediatric subspecialty fellows, assess their levels of comfort in the context of various communication topics, and query potential modalities and topics for future communication training. design/method: the needs assessment survey was developed using previously developed and validated questions and review of the literature. the survey was reviewed by internal and external pediatric oncology and palliative experts and pre-tested with a subset of trainees to enhance content validity. results: thirty-two out of a total of fellows completed the survey ( % completion rate), of which % were pediatric hematology-oncology or subspecialty fellows. most fellows had participated in previous teaching sessions ( %), including those involving role play or simulation ( %). however, few fellows had received feedback from senior clinicians on their communication skills ( % of fellows had received feedback ≤ times). on a scale of -x, with indicating "not well prepared," the mean score for of communication items was < . fellows felt least prepared to lead discussions around informed consent for experimental therapies, end of life care, and autopsy. fellows indicated that didactic educational sessions and additional coursework were less useful strategies for improving their communication skills, whereas small group role play sessions with faculty and/or bereaved parent educators were most useful. fellows' overall communication preparedness score was not correlated with post-graduate year but was positively associated with the number of times they previously had delivered bad news to patients and families. fellows requested additional training on many topics, with greatest interest in learning skills to optimize communication with an angry patient or family. additional topic requests included placing limitations on resuscitation, withdrawing/withholding further therapy, and ageappropriate inclusion of patients in difficult discussions. despite self-report of prior communication skills training, pediatric subspecialty fellows felt underprepared to participate in difficult discussions with patients and families. learners identified role-playing and coaching with real-time feedback from other physicians and bereaved parents as more useful training strategies as compared to didactic sessions. background: when children die of cancer, parents must adjust to their child's absence amidst the lingering turmoil of what preceded their death: witnessing their child undergo painful treatments, making difficult decisions, and anticipating a devastating loss, all the while hoping for a recovery. adjustment to a child's death, as depicted by current bereavement literature, necessitates making meaning of one's loss. professional care staff can help parents make sense of their child's illness, and in turn, of their own parental experience during treatment. however, the extent to which relationships with professional care team members influence parents' ability to make sense of, and successfully cope with, their loss has not been examined. objectives: to examine how bereaved parents' interactions with their deceased child's pediatric oncology professional care team have impacted their grief symptoms design/method: to better understand how interactions with professional care staff relate to parents' grief outcomes, we conducted a mixed-methods study examining staff impact on parental grief. thirty participants whose children died of cancer one to three years ago completed an in-depth interview and psychometrically validated surveys measuring meaningmaking, depression, and grief symptoms. results: correlational analyses of the measures found that an increase in meaning making was associated with lower depressive and grief symptoms. a content analysis of the interviews found that many participants regarded staff "like family," had on-going relationships with staff after their child died, and described various ways staff interactions during treatment and after the child's death helped them make sense of their loss. in particular, participants described how interactions with staff have helped them find benefits in their loss and learn to create a new relationship with their child despite their physical absence. quantifying the interview data and statistically analyzing it along with the measures found that participants' increased frequency of describing staff's positive impact on their grief correlated with higher meaning-making scores and lower grief symptom scores. our study found that bereaved parents who lost their children to cancer were articulate in sharing their experiences of staff engagement and communication during treatment, offering numerous examples of how staff aided them in making meaning of their loss that were reliably associated with their subsequent grief. we hope the results of this mixed methods research encourage further study of the importance of staff interaction with families during the critical period of their children's care, and the lasting impact this can have regardless of the treatment outcome. memorial sloan kettering cancer center, new york, new york, united states background: although resiliency has been recognized as necessary for healthcare professionals, trainees feel unprepared for the emotional challenges inherent in caring for sick and dying patients. compounded by long hours, challenging work environments, and lack of formal training on handling emotionally difficult situations, many institutions are recognizing the need for interventions to reduce trainee distress. the goals of this fellow-led quality improvement initiative were: ) to determine whether there is a need for emotional support amongst pediatric hematology and oncology fellows, ) to provide formal resiliency and debriefing sessions, and ) to measure feasibility, acceptability and effectiveness of implemented curriculum. design/method: an anonymous survey to determine need for resiliency and debriefing sessions following a traumatic event was distributed to active pediatric hematology & oncology fellows at memorial sloan kettering cancer center in january . once need was established, an intervention consisting of a formal curriculum was developed and initiated in june , involving: ) scheduled and ad hoc debriefing sessions in response to traumatic events (including patient death, codes, interpersonal conflicts, end-of-life care); led by a psychiatrist and social worker with fellows and a pediatric oncologist mentor in attendance, and ) a resiliency didactic curriculum, led by a palliative medicine specialist, focused on skills such as contesting cognitive distortions and mindfulness. the effectiveness of these sessions will be measured using follow-up anonymous surveys at months (currently underway) and months post-initiation of intervention. the initial survey demonstrated most trainees ( / ) were present at or more deaths during their training, while less than half of respondents had attended a post-event debriefing session. % of respondents felt there was not sufficient emotional support from the institution for physicians caring for dying patients. a separate pre-intervention survey found all respondents ( / ) expressed a need for regular debriefings, and nearly all anticipated that they would benefit from such debriefings. concerns identified by trainees that would preclude participation in the curriculum included preference to deal with emotional situations privately and time constraints. trainees identified a need for formal debriefings and resiliency skill development. the program was easily implemented, and is both feasible and acceptable with good attendance. feedback received at the -month mark will determine deficits and possible improvements to the curriculum. the -month survey will measure effectiveness of the program and whether it should be continued. background: acute kidney injury (aki) is a common but under-recognized complication among patients with leukemia. it is associated with prolonged hospital stays, increased mortality, progression to chronic kidney disease, and delays or changes in cancer therapy which may affect a patient's prognosis. however, data on aki in pediatric patients with cancer is still lacking overall. we investigated the incidence of aki in patients who were newly diagnosed with all at our center from january to september . we performed a retrospective chart review of all patients who were newly diagnosed with all from neonate to years in our facility. we determined the incidence of aki in our population using the kidney disease: improving global outcomes (kdigo) diagnostic criteria. we also assessed for nephrotoxic exposures, nci all risk stratification and risk of aki, and tumor lysis syndrome (tls). we identified patients diagnosed during the study period who met inclusion criteria. median follow-up time was . months (range . - . ). the cohort was predominantly male ( . %) and hispanic ( . %). our analysis showed . % had aki by kdigo criteria ( % grade , . % grade , and % grade ), . % had aki on presentation, and % had multiple aki episodes during the study period. older age and longer length of hospitalization were associated with aki (p = . and p = . , respectively). there was no association between aki and nci all risk classification, contrast exposure, hyponatremia, elevated white blood cell count, uric acid levels, antimicrobial therapy, or diuretic use in this study. conclusion: aki was a common finding in our study population. the majority had grade aki by kdigo criteria. however, aki was associated with older age and a longer length of stay. further study is needed to determine the short-and long-term impact of aki on pediatric patients with all. st. jude children's research hospital, memphis, tennessee, united states background: in some regions, the availability of trained pediatric oncologists is a limiting barrier for the care of children with cancer. in , the unidad nacional de oncología pediátrica (unop) and the universidad francisco marroquín school of medicine in guatemala established a pediatric hematology/oncology fellowship program sponsored by st jude children's research hospital to provide central america and the caribbean with well-trained specialists. a systematic analysis of the impact of fellowship programs in pediatric oncology has never been done, especially in the context of a regional education program. objectives: this study sought to analyze the impact of the unop fellowship program based on the regional number of providers, pediatric cancer centers and patient volume. in addition, it sought to characterize the jobs and scientific output of the graduates. the impact will be evaluated in the context of a cost analysis. to define the volume of providers, pediatric cancer centers and patients, the directors of pediatric cancer centers in central america were sent an online survey to obtain these data. all the centers contacted maintain an updated hospital-based patient registry. in addition, the graduates of the fellowship program were also sent an online survey, asking about their job at graduation, current role and scientific productivity. the cost analysis will include assessment of direct costs including salaries and stipends for away rotations, as well as the indirect costs of faculty time spent teaching. since the establishment of the unop fellowship program, the region has more providers for pediatric cancer (p< . ) and centers treat a larger volume of patients (p< . ). two new centers have opened with graduates of the program. all but one graduate practice pediatric oncology ( / ) and the majority do it in their country of origin ( / ). no graduate practices outside of this region. almost half of the graduates ( %) hold a leadership role at their institution. the majority of their time is spent in the public sector (> %). the majority of graduates participate in clinical research ( %) and have participated in the creation or implementation of therapeutic protocols ( %). on average, the graduates have published peer-reviewed articles since completion of training. the unop fellowship program has had a favorable impact on pediatric cancer care in the region, contributing to the capacity to treat a larger volume of patients. graduates practice pediatric oncology in the region in the public sector, frequently hold leadership roles and are scientifically productive. background: abandonment of treatment is a major cause of treatment failure and poor survival in children with cancer in low-and middle-income countries. the incidence of abandonment in peru has not been reported. objectives: the aim of this study was to examine the prevalence and associated factors of treatment abandonment in pediatric patients with cancer of peru. we retrospectively reviewed the sociodemographic and clinical data of children referred between january and december to the two main tertiary centers for childhood cancer, located in lima, peru. definition of treatment abandonment was used from the siop (international society of paediatric oncology) podc (paediatric oncology in developing countries) abandonment of treatment working group recommendation. results: data of children diagnosed with malignant solid tumors and lymphomas were analyzed, of which ( . %) abandoned treatment. univariate logistic regression analysis showed significant higher abandonment rates in children living outside the capital city, lima (p< . ); prolonged travel time to a tertiary center (> hours; or . , p = . ); living in a rural setting (or . ; p< . ) and lack of parental formal job (or . ; p = . ). according to cancer diagnosis, children with retinoblastoma were more likely to abandon compared with other solid tumors. in multivariate regression analyses, rural origin and lack of formal parental employment were independently predictive of abandonment. conclusion: treatment abandonment prevalence in our country is high and closely related to socio-demographical factors. treatment outcomes could be substantially improved by strategies that help prevent abandonment of therapy based on these results. st. jude children's research hospital, memphis, tennessee, united states background: to improve the quality of a pediatric hematology/oncology fellowship program, a systematic assessment must be performed that can evaluate its current state and identify areas of opportunity, as well as modifications over time. unfortunately, widely agreed-upon metrics of quality for pediatric hematology/oncology fellowship programs currently do not exist. this is particularly important in this field due to the global shortage of specialists. for this reason, an assessment instrument that is applicable throughout the world must be created. objectives: the st. jude global education program assessment tool (epat) is a novel instrument that seeks to evaluate pediatric hematology/oncology fellowship programs around the world in systematic and objective way. epat will help determine key performance indexes that are relevant for quality education in pediatric hematology/oncology fellowship programs and establish the framework for improvement. design/method: firstly, key domains to be evaluated for program assessment were identified a priori based on the continuum of pediatric hematology/oncology fellowship programs in the context of geography and educational structure. subsequently, questions were formulated to evaluate these key domains, seeking to assess elements involved in ensuring competence in clinical practice, academic productivity and regional impact. due to the novelty of this tool and the lack of defined metrics of quality, epat relies on expert opinion in a two-step process: internally in the department of global pediatric medicine at st. jude children's research hospital and, subsequently, from a panel of experts in global pediatric oncology and medical education from around the world. ten key domains were identified to evaluate all aspects relevant to training programs around the world, regardless of educational and geographic context. questions have been created to assess these domains and, to make epat quantitative, these have assigned weights with a value reflective of their relative importance. this grading system allows for a score in each key domain, permitting monitoring of changes over time. epat is currently at the stage of external expert review, and subsequently will be piloted in five fellowship programs around the world to provide different geographical and patient care contexts for its validation. once epat is finalized, it will be distributed to pediatric hematology/oncology fellowship programs around the world to be applied. epat proposes a novel strategy to assess training programs in a systematic way that includes all aspects relevant for a training program in a global context. this tool will help guide improvements in pediatric hematology/oncology fellowship programs and assure a well-trained workforce. background: with the improvement in pediatric oncology patient survival and outcomes in the past several decades, monitoring for recurrence and long-term effects of therapy has become even more important. the utilization of personalized treatment summaries and survivorship care plans (scps) is one way to communicate this information with patients and families. the american college of surgeons commission on cancer (coc) created a standard regarding provision of scps to % of eligible patients by december , as a metric for accreditation of all cancer centers. the standard applies to all patients with stage i, ii, and iii cancer diagnoses and requires creation of the scp within one year of diagnosis or six months of completing treatment. during implementation at our pediatric cancer center, we identified barriers to use of the guidelines in the childhood cancer setting. objectives: define eligibility for an scp for pediatric oncology patients to include all patients with curative intent and to deliver scps within six months of finishing therapy. design/method: using chart review and a cancer center registry query, we identified childhood cancer patients potentially eligible for an scp by collecting stage, goal of therapy, and dates of treatment. all patients with curative intent were deemed eligible for an scp regardless of stage i-iv. patients being followed in the oncology clinic for posttreatment surveillance and care were included even if they had received an scp in the survivorship program or were greater than six months off therapy at time of implementation. as expected in the pediatric oncology population, acute lymphoblastic leukemia (all) was the most common diagnosis comprising . % of patients. all is stratified into risk groups instead of surgical staging categories, and treatment duration is greater than one year, unlike many adult-onset malignancies. these differences required interpretation of the guidelines to apply to our pediatric population for all and other pediatric diagnoses with non-surgically based staging. our pediatric oncology clinic has to date provided scps to of eligible patients by adapting the guidelines to focus on patients with curative intent to receive an scp by six months off therapy. cancer staging guidelines and goals for curative intent as well as lengths of treatment vary between the pediatric and adult populations. the coc guidelines require adaptation for optimal applicability to the pediatric oncology population. background: education in communication for fellows in fields that require difficult discussions with families are few in nature. adult learning pedagogies such as role play are under-utilized in medical education, and have been shown to be as effective as traditional teaching methods such as lecture. an -module course for fellows in hematology/oncology, hospice and palliative medicine, radiation oncology, and pediatric hematology/oncology was implemented in january/february . fellows participated in the program. topics covered including fundamentals of communication, coping and spirituality, delivery of bad news, communicating with families, sexual dysfunction during treatment, palliative care/death and dying, and burnout. objectives: overall goal of this course is to foster holistic physicians who views their patients as people with cancer, not cancer patients, and physicians that can communicate effectively with their patients throughout the disease continuum. by the end of the course, learners should be able to practice the fundamental principles of good communication. design/method: fellows initially participated in a pre-course osce to establish baseline skills. osce was facilitated by the center for learning and innovation at northwell, and included actors portraying a pediatric patient and family member to whom the fellow had to break bad news. two months later, the course was carried out over the span of eight weeks and included didactic sessions followed by minutes of role play scenarios. five of the eight modules included role play, with faculty members serving as simulated patients. after the course, a second breaking bad news osce was held. both osces were filmed, and feedback was given by the on-site actors. additionally, faculty members were given access to the videos in an on-line format and were given an evaluation tool to assess the fellows' performance pre-and post-intervention. fellows were given subjective surveys pre-and post-course as well. results: subjective data from participants showed a noticeable increase in comfort level in all areas on the pre-and post-course survey. data obtained from osce videos showed improvement in communication skills as assessed by sps and faculty members using a new evaluation tool developed by faculty. initial first-run data shows that this course is successful in improving communication skills as well as increasing fellows' comfort level across several domains of communication. future directions for our course include improving and validating our assessment tool, expanding our topic base to include more aya and pediatric scenarios, faculty development for improved role play, and investigating impact on practice after course completion. background: acute lymphoblastic leukemia (all) is the most common form of childhood cancer with approximately children diagnosed each year. survival rates have improved significantly over the past several years. children with all are at risk for developing musculoskeletal complications during and after completion of treatment, which can contribute to impaired activity, elevated body mass index (bmi), and risk for complications. interventions involving physical activity could improve musculoskeletal strength as well as overall health in these children. the aims of this study are to examine the feasibility of a directed physical activity program for children with newly diagnosed all during the initial intensive phase of therapy and to evaluate the overall health and quality of life of children participating in the directed physical activity program. design/method: all subjects will receive education materials about the importance and safety of physical activity and a nutrition handout. all subjects will also participate in the directed physical activity program under the supervision of a trained physical therapist for at least minutes every week for weeks. the program will entail four stations including a cardiovascular, balance/proprioception, strength and flexibility, and coordination and cardio. feasibility will be assessed by tracking the participation rate throughout the study period. other assessments will be made at study entry, at the end of weeks of physical activity initiative and months after completion of the intervention. assessments include overall strength and flexibility, weight, height, bmi, blood pressure and performance scores. descriptive statistics will be used for this study. results: a total of patients, male and female, enrolled in the study over a . month period. patient ages ranged from - years. half of the patients enrolled have completed the week program and all patients had stability or improvement of their physical functioning scores. further data collection and analysis is ongoing. patients in the early intensive phase of all therapy are at risk for complications that can affect their physical functioning. a directed physical activity protocol may improve their overall physical functioning. patients may not need specific physical therapy; however a directed physical activity program appears to be beneficial for these patients. the main roadblocks to successful completion of the program were difficulty with scheduling, strain on the parents and patient from treatment, unplanned admissions for fever, as well as nausea and fatigue at time of visit. albany medical center, albany, new york, united states background: communication skills are a core competency highlighted by the acgme. increasing resident confidence in delivering difficult news has been shown to lead to more s of s effective communication. currently, the majority of residency programs lack formal training in communication skills. our objective was to demonstrate feasibility and efficacy of integrating a standardized-patient based training program for communication skills into the curriculum of pediatric residents design/method: to date, pediatric and medicine/pediatric residents have participated in the program during the intern year. the program consists of three, two-hour long sessions, in which each resident is given several opportunities to act out case scenarios with a standardized patient. scenarios included informing a parent of their child's new cancer diagnosis and disclosure of a positive hiv test to a teenager. residents received post hoc peer to peer, and preceptor to learner feedback. pre and post-program surveys were completed by residents. results: following course completion residents reported an increase in confidence in multiple areas of communication including giving a difficult diagnosis (p< . ), discussing a poor prognosis (p< . ), responding to different patient/family member emotional responses i.e. crying or anger (p< . ), and organizing vital information to be relayed (p< . ). in conclusion, communication skills training of pediatric residents is feasible and provides a platform for developing valuable skills not taught elsewhere within the curriculum. background: for children with cancer, transitioning back to school during or after treatment can be challenging. literature supports the need for school re-entry programs to ease this transition. however, these programs vary widely among pediatric cancer institutions with little data addressing their program components. data from this study provides information on current school re-entry programs across these institutions. objectives: one objective of this study was to assess for correlation between the presence of a school re-entry program and other factors, such as geographic location and institution size. a second objective was to establish a list of differences between institutions' school re-entry program components. finally, we aimed to describe current school reentry practices, as well as program benefits and perceived areas for improvement. states with membership in the children's oncology group were offered enrollment in this study. a member of each institution was invited to participate in a survey established by the research team. this person was closely associated with the institution's school re-entry practices. each interview queried institution demographics, as well as program components (e.g., participants, target audience, resources). comment was also collected on program benefits and potential for improvements. analysis of transcripts was performed using pearson's correlation to assess for relationships between institution size, geographic location, and program presence. grounded theory was used for analysis of benefits and improvements. results: thirty-nine of forty-one pediatric institutions who were offered enrollment participated in this study. twentynine institutions ( %) indicated the presence of a school reentry program, and ten ( %) stated they had none. no correlation was found between institution size and the presence of a school re-entry program (p = . , ns). there was also no correlation found between institution location and the presence of a school re-entry program (p = . , ns). a major theme surrounding the benefits of having a program included education for the returning student's peers. for those with programs, perceived improvements included increasing staffing and the ability to offer more services. the results do not support the hypothesis that the presence of a school re-entry program is influenced by the size and geographic location of the treating institution. however, data seem to suggest that available staffing may influence the presence of a program. future studies are needed to address other potential influences, as well as to take an evidence-based approach to determine the effectiveness of the interventions present in these programs. cohen children's medical center/ zucker school of medicine at hofstra-northwell, new hyde park, new york, united states background: genetics/genomics is evolving at an extremely rapid pace. current advances lead to individual algorithms toward disease treatment for each disease with multiple branch points. fellows learn only a fraction of the knowledge and there is no formal approach to teaching critical analysis of information and application algorithms toward disease. additionally, as knowledge evolves extremely rapidly, any approach must teach self-acquisition and application of evolving discoveries. objectives: to create, implement and evaluate a novel curriculum for genetics/genomics targeted toward pediatric hematology/oncology fellows design/method: the curriculum includes four components: ) genetic and genomic medical knowledge, with one initial team-based learning session and weekly online multiple choice questions; ) essential pathways, which will teach molecular pathways common in oncogenesis and relevant to targeted therapy in microteaching sessions with using auditory, visual and tactile learning; ) knowledge acquisition and clinical judgment, to allow learners to gain experience into researching data available, then developing and prioritizing potential treatment plans using problem-based learning sessions in which they will stage a patient, research treatment options, prioritize and present findings; and ) synthesis to demonstrate independent ability to research and recommend therapy through an independent project in which the learner, given a case, will present the case and research findings, genetics/genomics, molecular pathways and make recommendations for therapy in molecular tumor board for faculty and fellows. to evaluate, we plan to recruit to institutions, match for size of programs and implement in half and evaluate nd and rd year fellows in both groups by mcq exam and satisfaction surveys. the creation of a multi-module, adult-learning based curriculum for genetics and genomics in pediatric oncology is feasible. implementation and evaluation are necessary to demonstrate efficacy. background: neuroblastoma is the most common extracranial solid tumor in children. chimeric anti-gd antibody ch . (dinutuximab) therapy has improved the survival of children with newly diagnosed high-risk, neuroblastoma patients as well at the time of first relapse/progression. acute neuropathic pain is a well-documented side effect of dinutuximab administration. however, additional adverse effects including sensorimotor neuropathy, ocular symptoms, and behavioral changes have been described. the incidence and severity of these effects are currently not well-documented in pediatric patients. with improved long term survival of patients receiving this modality, it is important to look for the potential late effects of dinutuximab. objectives: to determine the incidence and severity of neurologic, ophthalmologic, or behavioral changes after dinutuximab administration at our institution. we performed a retrospective chart review using our electronic medical record. we included all patients with high-risk neuroblastoma between the ages of and years at our institution diagnosed between and who received dinutuximab. patients with history of opsoclonus-myoclonus syndrome or gross sensorimotor neuropathy prior to receiving dinutuximab were excluded. we examined clinical documentation for subjective reports and objective exam findings of neurologic, ophthalmologic, or behavioral changes. we also looked for referrals made to neurology, ophthalmology, physical medicine & rehabilitation (pm&r), and psychology. : twenty-two patients met inclusion criteria. at the time of chart review, patients were alive and were deceased. eighteen patients received dinutuximab per anbl ; patients received dinutuximab per anbl . of these patients, patients reported symptoms of interest and reported multiple symptoms. six patients reported symptoms that began at least months after completing dinutuximab. nine patients had objective findings on exam, including decreased deep tendon reflexes, abnormal pupils, and nearsightedness. for patients, referrals were made to ophthalmology, pm&r for neuropsychologic testing, or neurology. two patients who reported symptoms of interest were not referred to a specialist. conclusion: neurologic, ophthalmologic, and behavioral symptoms were commonly reported and demonstrated on exam among pediatric patients with high-risk neuroblastoma who received dinutuximab. it is important to identify these effects so that appropriate specialist referrals can be placed for adequate management of these changes. we recognize that these symptoms may not be solely due to dinutuximab as these patients receive other agents including opioids, so a prospective trial is needed to further evaluate the long-term effects of dinutuximab and to determine how best to screen for these effects. akron children's hospital, akron, ohio, united states background: pediatric cancer is the leading cause of diseaserelated death in children in the united states (u.s.). in , over fifteen thousand children were diagnosed with cancer in the u.s. this population is at high risk for malnutrition due to the multimodal therapies they receive: surgery, chemotherapy, radiation therapy, antibody therapy, and/or bone marrow transplant. adverse effects of these therapies include taste changes, loss of appetite, diarrhea, vomiting, and/or mucositis, making it difficult for the children to be able to consume adequate amounts of nutrition during therapy. there is no "gold standard" measurement tool for identifying patients at risk for malnutrition. nutritional status is not frequently evaluated as a component of clinical trials. assessment of anthropometric measurements (weight, height, z-scores) at diagnosis, as well as over the duration of treatment, can assist in the early identification of malnutrition. the incidence and prevalence of malnutrition in this population is unknown at akron children's hospital. the purpose of this study is to describe the nutritional status and provision of nutritional support therapies in pediatric patients during their first year post new oncologic diagnosis. objectives: identify the incidence and prevalence of malnutrition across oncologic diagnostic categories over the first twelve months post diagnosis. we performed a retrospective records review of all patients newly diagnosed with cancer in at akron children's hospital. demographic and anthropometric data was collected at time of diagnosis and nutritional status categorized by z score. anthropometric and nutrition support data was then collected every two months for the first year after diagnosis along with incidence of unplanned inpatient admissions. results: a total of patients were included in the analysis, with . % malnourished at time of diagnosis; . % developed malnutrition the first year. patients with solid tumors represented % of patients with pre-existing or acquired malnutrition. overall, % of patients received at least one nutritional support modality. patients with pre-existing or acquired malnutrition had a non-significant increase in unplanned admissions (p = . ). our study demonstrated that patients with solid tumors were found to be at increased risk of pre-existing and acquired malnutrition, followed by leukemias, and experienced higher incidence of unplanned admissions in the time period observed. prospective, multi-center replication of this study, including detailed collection of nutrition therapies is recommended to guide development of diagnosis specific nutrition support guidelines. background: pediatric and young adult oncology patients treated with intense chemotherapy have a high incidence of transfusional iron overload. iron deposition can lead to heart failure/arrhythmias, liver abnormalities, endocrine dysfunction, ineffective erythropoiesis, and increased cancer and mortality risk. however, there is a paucity of data regarding recommendations for management of transfusional iron overload in these cancer survivors. consequently, long-term complications of transfusional iron overload specific to these patients have not been assessed. objectives: to assess screening and phlebotomy-based treatment algorithms for this population. design/method: a retrospective chart review of pediatric and young adults who completed oncology management, had iron overload, and initiated phlebotomy treatment was conducted. tiered screening occurred in patients that received at least packed red blood cell (prbc) transfusions. patients were recommended for evaluation and possible phlebotomy if: ( ) liver iron concentration (lic) > mg of iron/gram dry weight liver tissue by ferriscan and/or ( ) cardiac mri t * < ms. during phlebotomy, iron status was assessed quarterly and phlebotomy discontinued with lic < or normalization of ferritin/imaging lic verification. descriptive statistics were employed to report the characteristics of the study population. spearman correlations were utilized to describe associations between transfusions, lic, ferritin, iron saturation and number of phlebotomy sessions. results: twenty five survivors underwent phlebotomy. the mean age was . years (sd . ) and ( %) were female. oncologic diagnoses: all ( %), aml ( %), nhl ( %), ewing sarcoma ( %), osteosarcoma ( %), neuroblastoma ( %) and cns ( %). patients received a median of . (iqr - ) transfusions. median number of phlebotomy sessions was (iqr - ) over . years (iqr . - . ). prior to phlebotomy, median lic was . mg/g (iqr . - . ) and ferritin was . ng/ml (iqr - ) . no patients demonstrated abnormal cardiac t * mri (n = ). ( %) patients completed phlebotomy. one discontinued due to poor vascular access. no patients developed iron deficiency. lic was reduced by a median of . mg/g (iqr . - . ) and ferritin by ng/ml . correlation between number of transfusions and phlebotomy sessions was poor (r = . ). conclusion: management guidelines are lacking for transfusional iron overload in pediatric and young adult survivors of cancer. we demonstrate a phlebotomy algorithm that is effective and tolerated. correlation between number of transfusions received and phlebotomy treatments was poor, necessitating serial assessments. using this management algorithm, prospective studies can evaluate the effect of iron removal on iron overload complications in this patient population. penn state children's hospital, hershey, pennsylvania, united states background: cancer therapy leads to an impaired immune system that takes time to recover. it is important to ensure that these survivors have adequate immunity to prevent common yet potentially severe childhood illnesses. no validated guidelines currently exist for surveillance testing or re-immunization in this population. retrospective analysis involving a small cohort of pediatric cancer patients treated at penn state children's hospital showed % of patients screened for varicella immunity after therapy completion did not have adequate disease titers. to determine the proportion of pediatric cancer survivors who have lost humoral immunity to previously received vaccines; to determine the rate of response to single dose boosters or full vaccine series in seronegative subjects after one booster. design/method: pediatric cancer survivors treated at the children's hospital who are at least months from completion of cancer therapy are prospectively tested for antibody levels to hepatitis b, tetanus, varicella, measles, and strains of pneumococcus ( , b, v, c, f, and f). samples are analyzed by the cdc for measles and varicella avidity. seronegative subjects by commercial studies, are eligible to receive booster vaccines. titers are rechecked at least weeks after boosters to re-evaluate immunity; if still seronegative, subjects will receive the entire vaccine series. titers are finally tested at least weeks after the final dose of the vaccine series. immunity analyzed after therapy, after boosters, and after vaccine series. results: of pediatric cancers survivors who completed therapy, % were non-immune to hepatitis b, % nonimmune to > % of pneumococcal strains tested, % nonimmune to measles, % non-immune to varicella, and % non-immune to tetanus. of subjects who received mmr vaccine after therapy and prior to study enrollment did not have protective antibodies to measles. of the subjects who received varicella vaccine after end of therapy and prior to study enrollment, did not maintain protective antibody levels. cdc results for measles and varicella are pending, as well as repeat studies after vaccine boosters and series. conclusion: a significant percentage of pediatric cancer survivors do not retain immunity to hepatitis b, pneumococcus, measles, and varicella. after one booster, a high percentage of subjects did not develop protective immunity to varicella. only subject did not have immunity to tetanus, which is consistent with the high immunogenicity of tetanus toxoid. formal guidelines are needed to protect this population from vaccine-preventable illness post-therapy. children's hospital of richmond at virginia commonwealth university health system, richmond, virginia, united states background: childhood cancer survivors are at risk for being overweight. diet and physical exercise are important in maintaining a healthy lifestyle and weight; however, it has been reported that cancer survivors are less active than their peers. one reason for this may be that there are no clearly established risk-based exercise recommendations for cancer survivors. another reason may be that providers tend to focus s of s recommendations for exercise more towards patients who are overweight. objectives: to describe changes in physical fitness of childhood cancer survivors who exercise. design/method: 'moving forward' is a wellness and physical fitness program that the center for care beyond the cure at chor offers in partnership with the ask childhood cancer foundation and the ymca. the program is available for any childhood cancer survivor between y and y age, being seen at our center. survivors define their fitness or wellness goals and then work with a trainer once a week (at least) for min sessions throughout the year to achieve these goals. baseline and ongoing measurements for core strength, endurance, overall strength and balance were collected. the average of each of the parameters of all participants were compared from the beginning to the end of the program. over the year, there was a % increase in endurance as measured by the average of the miles walked in minutes, % increase in core strength as measured by the average number of sit-ups in secs, an % and % increase in overall strength as measured by the average weight lifted by leg press and the average weight lifted by chest press, and a % increase in balance as measured by the average number of seconds balancing on a single leg. in addition, each child had actually gained weight in the process with an approximately % increase in the average of the weights of all children. there are benefits to regular exercise beyond weight control, and improvements in physical fitness can be seen even without weight loss. regular physical exercise results in improved physical fitness and should be universally advocated to all patients. determining insulin resistance, measuring changes in fatigue and wellness perception following exercise are future directions that we intend to explore. dana-farber cancer institute, boston, massachusetts, united states background: improvements in adolescent and young adult cancer patient (aya) survival rates and quality of life outcomes have lagged behind those of children and older adults, highlighting a need for research targeting this unique population. current literature supports the value of strong ayaclinician communication, notably in facilitating therapeutic alliance, however little is known about aya communication priorities during cancer care and barriers to optimal ayaclinician communication. objectives: to explore aya and oncology clinician communication priorities and to identify barriers and facilitators to aya-oncology clinician communication. design/method: semi-structured interviews were held with aya cancer patients and survivors (ages - years) from a single large academic institution and oncology clinicians (physicians and nurse practitioners) from academic institutions in the northeastern united states. interviews were conducted in english by phone or in person. all interviews were audio-recorded and transcribed verbatim. analyses were aided by nvivo software. ayas identified a wide range of topics as important to discuss with clinicians. the most frequently identified topics were ) side effects of treatment (with an emphasis on physical appearance and function, n = ), ) social issues (including friendship, family, and school, n = ), ) looking ahead to the future (n = ), and ) sexual & reproductive health (including future fertility, contraception, and romantic relationships, n = ). clinicians prioritized ) cancer treatment and side effects (n = ), ) emotional and psychological health (n = ), and ) sexual and reproductive health with a focus on fertility risk and fertility preservation (n = ). aya reported facilitators to good communication including an open and long-established relationship with the clinician (n = ) and clinician engagement in age-appropriate and patient-directed conversations (n = ). barriers included parental presence during visits (n = ). clinicians reported barriers including ) clinician discomfort (not feeling wellequipped to discuss psychosocial topics such as sexual health, spirituality, and relationships with peers, n = ), ) presence of parents/family (n = ), and ) perceived patient discomfort discussing specific topics (such as sexual health, n = ). clinicians acknowledged the need for collaborative efforts with additional team members (i.e. nurses, psychosocial providers) to assist in meeting aya communication needs. conclusion: aya and clinician-reported communication priorities are largely aligned. however, ayas emphasize some topics, such as social function, appearance, and sexual health that are not highly prioritized by clinicians, which may result in gaps in care for ayas in treatment and in survivorship. these data identify opportunities for intervention, including clinician education, patient and family education, clinic-based intervention, and systems-based changes that can be developed and tested. background: primary care physicians (pcps) cite lack of knowledge and inadequate communication with the oncology team as major barriers to providing recommended surveillance for late effects of treatment to childhood cancer survivors. a standardized telephone handoff to pcps posttherapy is a potential strategy to increase survivorship care by pcps through interactive communication. to determine the feasibility of a structured telephone communication using the situation, background, assessment, and recommendation (sbar) communication tool delivered by a trained oncology nurse to increase pcp knowledge and willingness to provide survivorship care. design/method: from / / to / / , a registered nurse expert in childhood cancer survivorship attempted to contact by telephone the pcps of the most recent patients attending yale's childhood cancer survivorship clinic that were < years old, english-speaking, and ≥ years posttreatment. all pcps had been previously sent an individualized survivorship care plan (scp) that listed the patient's previous treatment history and recommended surveillance tests. upon successful contact and after confirming receipt of the scp, the nurse explained the definition of late effects, description of patient's diagnosis and treatment history, and associated potential late complications and schedule of recommended surveillance tests. the pcp was also asked about his/her ability and willingness to provide needed surveillance for late effects in the future. overall, of pcps were successfully contacted with a median of phone call (range: - ) that lasted a median of minutes (range: - ) after a median of business day (range: - ). no pcps ended the call mid-conversation. all pcps were receptive and expressed appreciation for the call. twenty-five of ( %) pcps expressed an understand-ing of the material discussed and endorsed belief in their ability and willingness to provide late effects surveillance for their patients. no pcps questioned discussing their patient's care with a nurse versus a physician. interactive, structured communications between nurses and pcps by telephone are feasible and are associated with high-levels of pcp confidence in providing survivorship care. background: childhood cancer (cc) admissions account for % of non-newborn pediatric hospitalizations. these hospitalizations are longer and more expensive than other hospitalizations. admission payer (medicaid or commercial) reflects both health policy and sociodemographic status. the objective of this study was to determine if length of stay (los) or cost of cc admissions differed by payer. we used the kids inpatient database, a sampling of all pediatric hospital discharges in the united states. analysis for this study was limited to admissions containing a cancer diagnosis in any discharge icd- codes. admissions were further subcategorized by discharge codes according to diagnosis (leukemia, lymphoma, solid tumor and brain tumor) and reason for admission (chemotherapy, procedure, infection, non-infectious toxicity or "other"). charges were converted to costs using cost-to-charge ratios. multivariable linear regression models were performed to control for age, gender, race, reason for admission, and diagnosis. results: there were , weighted admissions for children with a cancer diagnosis in . of these admissions, . % had medicaid, . % had commercial insurance, and less than % had other payers. the mean los for medicaid admissions was . days ( % ci . - . ), compared with . days ( % ci . - . ) for commercial insurance. surgical admissions accounted for the largest difference in length of stay with medicaid admissions being . days longer than those covered by commercial insurance ( . days vs . days), however, the difference was significantly different for all reasons for admission. in multivariable analysis admissions associated with commercial insurance were % shorter s of s (p< . ), accounting for approximately one hospital day, than admissions associated with medicaid after controlling for other variables including race. the mean overall cost for medicaid admissions was $ , ( % ci - ), compared with $ , ( % ci - ) for commercial insurance. in the multivariable model, cost was collinear with race. conclusion: los and cost of admissions associated with medicaid differed from those associate with commercial payers. medicaid admissions were % longer on average than commercial insurance, accounting for a difference in length of stay of approximately one day although the difference varied with the reason for hospitalization (chemotherapy, surgical procedure, infection, other toxicity, other). costs of admissions were not independent of race. further investigation into potential explanations for this difference including differential access to home care needs, outpatient reimbursement differences, social indications for prolonged hospitalization, and provider biases, is warranted. background: pediatric cancer is a major cause of morbidity and mortality among children surpassed only by accidents. despite improved outcomes in high income countries (hic) survival rates remain poor in the developing word. there are various diagnostic and therapeutic limitations contributing significantly for the survival gap. the main objective of the study is to to evaluate the outcomes of pediatric cancer in armenia and identify diagnostic and therapeutic limitations in the country. we conducted a retrospective study among (≤ years old) children with cancer (solid tumors and hematological malignancies), who were diagnosed and treated at the clinic of chemotherapy of muratsan hospital complex of yerevan state medical university between and . those patients, who didn't receive chemotherapy for any reason were not included in the study cohort. epidemiological, social, medical information was collected through the patient charts review. this included patient age at diagnosis, sex, place of residence (city vs village), the educational level and employment status of parents, type of cancer, stage, presentation of symptoms, first medical specialty consulted and the time consulted, initial work-up, the type of treatment received, information on the diagnosis/treatment received abroad. results: at our clinic during the mentioned period of time the majority of patients presented with hematologic malignancies- %. ( . %) patients had information on diagnosis delay. average delay in diagnosis was about days. in % of cases the first contact with "healthcare system" was through pediatrician, and in % with surgeon. out of relapsed patients received salvage treatment in armenia and abroad. from those who stayed for treatment in armenia patients survived. majority of relapsed patients had acute lymphoblastic leukemia. from leukemia patients immunophenotyping and cytogenetics were available for ( . %) patients; the majority of missing cases were between and , when these diagnostic modalities were not available or affordable in the country. ( %) patients received part of diagnosis and/or treatment abroad. the most frequent reason for going abroad was bone marrow transplantation, otherwise none available in armenia. out of patients were lost to follow-up, patients had a fatal outcome. patients were in remission at a median follow up of . years. conclusion: unavailability of cancer registry and several essential diagnostic/treatment modalities, luck of multidisciplinary care and palliative support, high rate of out-of-pocket expenses were among the main challenges of pediatric cancer care in armenia. background: adverse drug reactions (adrs) are increasingly recognized as important and sometimes irreversible complications of cancer treatment. anthracyclines and cisplatin are effective chemotherapeutic agents, but their use can be limited by cardiotoxicity (anthracyclines) and ototoxicity (cisplatin) in up to % of patients. genetic variants that can be used to predict who is most at risk of developing these adrs have been discovered and replicated. objectives: to create pharmacogenetic risk prediction models for anthracycline and cisplatin toxicities and discuss results with oncologists to facilitate incorporation into treatment decision-making when appropriate. design/method: risk prediction models were developed from the linear regression of strongly-predictive genomic variants (odds ratios ≥ ) discovered and replicated in at least three patient populations. these models were used to assess an individual patient's genomic risk of developing cardiotoxicity from anthracyclines or hearing loss from cisplatin. risk results were returned to oncologists showing where the specific patient's genetic risk of toxicity lies on a continuum between the lowest and highest risk groups across all studied patients using a multi-gene model. interviews were conducted with patients, families, and oncologists to determine how results were valued and utilized. results: patients have been genotyped and had their genetic risk results returned to their oncologists. the first patients have been characterized to determine the impact these test results have had on their clinical care. results were described as being useful in decision-making by patients and/or oncologists in % of cases. additionally, for patients in the most extreme risk groups (highest and lowest risk), a change in treatment plan was ordered % of the time for cisplatin patients and % of the time for anthracycline patients. this included increased cardiac and audiological monitoring, the addition of a protective agent, or choosing an alternative treatment protocol if the risk outweighed the benefits of remaining on the current treatment plan. in interviews, patients indicated that they felt more involved in decision making, and felt reassured by understanding their genetic risk of toxicities. genetic risk prediction models for anthracycline cardiotoxicity and cisplatin ototoxicity were highly utilized by patients and oncologists in decision-making. results were found to be an important tool for informing patients of the risk of adrs during cancer treatment, and resulted in patients and their families feeling more involved in decision-making. background: childhood cancer survivors are at increased risk of developing executive dysfunction, and low socioe-conomic status (ses) has been identified as one of the mediators of executive functioning. previous studies have used traditional measures of ses, such as parents' education level, family annual income and occupation. but more recently, area based socioeconomic measures like block group poverty status are deemed to be more useful in monitoring of social inequalities in health in the united states. block groups are statistical divisions of census tracts and generally contain between and , people. the current study aims to understand the association of block group poverty status (percentage of households in family's block group of residence living below the federal poverty level) with executive functioning among cancer survivor children. design/method: we used a retrospective cohort of childhood cancer survivors. relevant information was collected from the medical record, administrative data sets and parent-filled surveys. address information was geocoded using arcgis . to obtain data on the block group poverty status. a priori cut-points were set to represent block groups with families living below poverty level at %, . % to . %, and ≥ . %. executive functioning were assessed through a parent-rated instrument, the behavior rating inventory of executive functions (brief). multiple linear regressions were used to determine the relationship between block group poverty status and the brief scores. results: data was examined from families of childhood cancer survivors, ranging in age from to years. in this sample, . % families reported an annual income <$ , , . % reported income between $ , and $ , while . % reported annual income ≥$ , . primary care giver of . % of cancer survivors had more than more high school education, and . %, . % and . %, of families were living in a block groups with %, . - . % and ≥ % poor households respectively. block group poverty level was not significantly associated with annual income levels (spearman's rho = . , p = . ), or parental education level (spearman's rho = - . , p = . ). in a step-wise multiple linear regression, there was no statistically significant association seen between block group poverty status and executive functioning after adjusting for co-variables in the final model. future prospective study with a bigger sample size, longer follow up period and more robust measures of the executive functioning like a clinician administered test are needed to understand the effect of block group poverty status on executive functioning. to d completion was . days (range - ). all parents strongly agreed/agreed that d was helpful and would recommend d participation to another family. ten parents ( %) reported time spent on d was "just right." no parent felt more worried due to the intervention, though parent found d participation stressful. this interim analysis suggests that parents have a favorable d experience and recommend the intervention. to date, < % of enrolled parents fail to participate. d shows promise as an acceptable interdisciplinary communication intervention targeted to the early treatment period for childhood cancer. children 's hospital and research center oakland, oakland, california, united states background: screening echocardiograms are recommended by children's oncology group (cog) guidelines to assess for anthracycline-induced left ventricular (lv) systolic dysfunction. the yield of screening echocardiograms during chemotherapy and in the immediate post-therapy period is uncertain. objectives: to assess the incidence of lv dysfunction detected by screening echocardiograms during chemotherapy and in the immediate post-therapy period, defined as - months off-therapy. design/method: children diagnosed with cancer between january -march who received anthracycline chemotherapy were identified. echocardiograms were performed as per protocol, institutional and cog guidelines, and were reviewed retrospectively. lv dysfunction was defined as fractional shortening (fs) < % or ejection fraction (ef) < % ( ) results: in this cohort (n = , median age years), the most common diagnosis was all ( . %), followed by aml ( . %). of echocardiograms, ( . %) were performed during treatment and in the immediate posttreatment period. thirty-eight ( . %) patients had a > % decrease in fs compared to their pre-treatment echocardiograms. none of these patients required any treatment modification or cardiac medications. only patient ( . %) had echocardiogram-proven lv dysfunction discovered on a screening echocardiogram during her treatment course. she eventually died due to multi-organ failure following septic shock. this patient was receiving treatment for aml and had received mg/m of doxorubicin-equivalent anthracyclines at the time of the abnormal echocardiogram. one patient with metastatic ewing sarcoma had borderline lv dysfunction with a fs of % detected a month before completion of therapy. she had received mg/m of doxorubicin equivalent anthracyclines at the time of the abnormal echocardiogram. she did not require any therapy modification or additional cardiac medications. serial echocardiograms done on this patient have shown stable ventricular function. no off-therapy screening echocardiograms identified lv dysfunction. in our experience, the yield of echocardiograms to detect anthracycline-related cardiac dysfunction during treatment and in the immediate post-therapy period is very low. one patient developed lv dysfunction during treatment and one had borderline fs, while no lv dysfunction was identified within months of completing chemotherapy. though fs decreased in % of patients, none required intervention. further study is needed to optimize the use of echocardiography screening in children treated with anthracyclines. references: . landier w et al. jco . background: platinum-based chemotherapy increases the risk of sensorineural hearing loss in children with cancer. little is known about the impact of hearing loss on cognitive and emotional functioning in survivors. to determine the association of severe/profound hearing loss after platinum-based chemotherapy with ) cognitive impairment and ) emotional distress (i.e. anxiety and/or depression). cross-sectional study of all patients attending yale's childhood cancer survivorship clinic ≥ years off therapy for cancer diagnosed at < years and treated with cisplatin and/or carboplatin, but with no history of cns tumor, cranial radiation, congenital hearing loss, or developmental delay. hearing loss severity and hearing aid data were abstracted from audiograms and detailed clinical history. cognitive impairment was defined as behavior rating inventory of executive function t score ≥ , assessment by neuropsychologist, and/or history of special education. emotional distress was determined by brief symptom inventory t score ≥ (global or two subscales) or behavioral and emotional screening system t score ≥ , psychologist interview, and/or history of psychotropic medication/psychotherapy. the most recent available patient data were used. logistic regression with sas software, version . was performed. results: overall, patients ( % female, % white) met eligibility criteria with a median age of . years (iqr = . ) at diagnosis and . years at evaluation (iqr = . ) after a diagnosis of sarcoma ( %), neuroblastoma ( %), or other ( %) for which % received cisplatin and % received carboplatin. fifteen patients ( %) had severe/profound hearing loss in at least one ear. patients with severe/profound hearing loss had a significantly increased risk of cognitive impairment (or = . ; % ci = . - . ), but not emotional distress, compared to patients without severe/profound hearing loss. there was no significant association between age at diagnosis, current age, time since diagnosis, sex, race, ethnicity, or diagnosis with either cognitive impairment or emotional distress. similarly, there was no significant interaction between ) age at diagnosis and hearing loss or ) sex and hearing loss with either cognitive impairment or emotional distress. ten of the ( %) patients with severe/profound hearing loss in at least one ear were recommended hearing aids, of which ( %) reported compliance most of the time. we conclude that severe/profound hearing loss is significantly associated with cognitive impairment, but not emotional distress, in childhood cancer survivors. our data supports the need for interventions to improve hearing in these patients, including compliance with hearing aids. background: who grade anaplastic astrocytoma is a high grade glioma dependent on vascular endothelial s of s growth factor (vegf) mediated angiogenesis for its growth and infiltration. bevacizumab is a recombinant humanized monoclonal antibody which binds vegf-a and inhibits angiogenesis. common adverse effects of bevacizumab are hypertension, proteinuria, thrombosis and bleeding. while animal model based studies have shown that bevacizumab may impair ovarian function the effects of bevacizumab therapy on human fertility are not clear. since the physiology of pregnancy involves neovascularization/angiogenesis it is recommended that conception be avoided for at least months following exposure to bevacizumab. to describe the course of a young adult who became pregnant after receiving bevacizumab and radiation therapy for treatment of an anaplastic astrocytoma. a year old woman diagnosed with a localized hemispheric who anaplastic astrocytoma was treated with chemotherapy and radiation (temozolomide/ . gy) followed by cycles of bi-weekly bevacizumab/temozolomide. patient opted not to pursue fertility preservation prior to initiation treatment. she experienced bevacizumab-associated proteinuria and hypertension during treatment but received all protocol mandated doses (cumulative doses: bevacizumab = mg/kg; temozolomide = . gm/m ). she had a spontaneous unassisted pregnancy months after completing treatment. her pregnancy was uneventful and she was normotensive throughout. fetal ultrasonography at , , , weeks revealed no abnormality of the brain, heart, great vessels, kidney, extremities, placenta and umbilical cord. at weeks she delivered a female infant via cesarean section (birth weight: grams, apgars: and ) excessive post-partum hemorrhage was not reported. placenta was bi-lobed and weighed g. histological analysis revealed normal placental villous development and maturation and two small infarcts. conclusion: exposure to bevacizumab in our patient had no detrimental effect on fertility and on placental/fetal vascular development. we hope this report will add to the existing data on the effects of bevacizumab therapy on fertility. children's healthcare of atlanta, emory university school of medicine, atlanta, georgia, united states background: reports of malnutrition incidence and prevalence in young cancer patients are variable and not well established. previous research suggests children, especially less than years old, treated with intensive cancer-directed therapy are at higher risk for malnutrition. however, no standardized assessment has been used to evaluate risk in this population. objectives: we aim to assess the trends of weight-for-age for patients following cancer diagnosis. this study will be the first to use a standardized measure of treatment intensity (intensity treatment rating scale, itr- ) and will assist in targeting interventions for identification and treatment of malnutrition. design/method: this observational, retrospective study obtained data through the center's pediatric cancer registry and electronic medical record. patients were classified by tumor type (brain or non-brain tumor) and treatment intensity (itr- ). itr- incorporates diagnosis, chemotherapy, radiation, and surgery, beginning with lowest intensity ( ) to highest intensity ( ). inclusion criteria included new cancer diagnosis - at less than years old, with weight obtained and available within days of therapy start date. incomplete data, alternate growth charts, or treatment intensity of , were excluded. weight was obtained at start of therapy and through years after treatment initiation (approximately days) and converted to z-scores adjusted for age and sex. weight trajectories were modeled using generalized linear mixed models with subject-specific random intercepts and spline functions. separate functions were constructed for subgroups of interest (tumor type and itr). results: there were patients included: patients with brain tumors ( . %) and with non-brain tumors ( . %). of included patients, had treatment intensity of ( . %), of ( . %) and of ( . %). over the observation period, , valid weights were recorded. at initiation of treatment, no difference existed between z-score by tumor type (p = . ) or by intensity ( vs. , p = . ; vs. , p = . ; vs. , p = . ). tumor type did not affect z-score through the follow up period. z-scores were higher for intensity rating vs. and vs. (p = < . and p = . respectively) at days after the start of treatment and persisted through days (p = . and p< . respectively). higher treatment intensity is associated with decline in z-score and failure to return to baseline. future directions include further analysis on specific risk factors and timing of weight loss, longer-term follow-up of weight trends, and targeted interventions for identification, prevention, and treatment of malnutrition. objectives: asses the pt requirements for bleeding episodes in a prospective cohort of pcp using a < × e threshold compared to a < × e /l threshold in a historical cohort. we collected pt data in all pcps treated at our center between january/ through december/ . diagnosis, prescription for pt (prophylaxis vs bleeding disorder), plt count and transfused units were assessed for each pt. pcps treated from january/ through june received prophylactic pt with a < × e threshold (cohort a), and pts treated from july/ through december/ received prophylactic pt with a < × e threshold. pts done for procedures and pts with concomitant hemorrhagic pathology were excluded. we compared the number of pts prescribed as prophylaxis vs bleeding episode between cohorts. data analyzed: graphpad prims . ®. statistical analysis: percentages with confidence interval (ci); t-student test (parametric variables) and mann-whitney test (nonparametric variables). statistical significance: p< . . we reviewed pts ( in cohort a, cohort b) in patients. % had acute leukemia, % received and auto or allo hsct. diagnoses and the proportion of patients undergoing hsct was comparable in both cohorts. the average number of pts per patient was , in cohort a and , in cohort b (p = ns), but a significant difference was found when hsct patients were excluded from this comparison ( , pt per patient in cohort a vs , in cohort b, p = , ), which resulted in an estimated , % reduction in pts prescription. furthermore ( , %) pts were prescribed for bleeding episodes in cohort a versus ( , %) in cohort b (p = ns). patients receiving hsct in the entire group ver-sus those not receiving hsct had similar pt requirements for bleeding episodes ( % vs , % p = ns) conclusion: a < × e plt count threshold for prophylactic pts is safe in pcp in chemotherapy and hsct. it can result in a significant reduction in pt usage. key words: platelets, transfusions, prophylaxis, cancer, childhood. ucsf benioff children's hospital oakland, oakland, california, united states background: transition of care for adolescent and young adult (aya) survivors of childhood cancer from pediatric to adult-oriented long-term follow-up (ltfu) is complex. loss to follow-up is common, and little is known about the success rates among different models. the survivors of childhood cancer program (sccp) at ucsf benioff children's hospital oakland employs a community-based model for transitional care. our multidisciplinary team provides aya survivors a comprehensive treatment summary and recommendations, then facilitates transition to primary care or adult oncology ltfu programs. evaluate the success rate for transition of care among aya survivors of childhood cancer in our ltfu program, and identify barriers to successful transition. design/method: aya patients seen from november to august in the sccp with intent to transition were asked by email or telephone if they had followed up with their designated provider. the primary outcome was successful transition, defined as establishing care within months of their visit. patients were also asked about barriers to transition and to rate the new provider's familiarity with their cancer history and ltfu needs. results: transition was intended for patients. eightyseven were contacted and responded. of these, ( %) successfully transitioned, while ( %) were lost to followup. ages ranged from to years, at to years since completion of therapy. ten ( %) transitioned to a primary care provider, ( %) to an adult oncology ltfu program, and ( %) to a pediatrician. patients rated their new provider's knowledge above average ( . ) on a -point scale from poor ( ) to excellent ( ). survivors lost to follow up indicated the following barriers to transition: loss/change of insurance ( ), inability to find a provider ( ), too busy/forgot ( ), problems with transportation ( ), concerns about cost/copay ( ), and s of s other ( ). twelve patients requested further assistance with transition. conclusion: two-thirds of responding patients successfully transitioned. more work is needed to overcome various barriers to transition for one third of aya survivors. albany medical center, albany, new york, united states background: the transition from active treatment, to offtherapy follow-up, is a stressful event for parents of children with cancer. the psychosocial needs of parents after therapy have received limited attention in the united states with only published quantitative studies, the largest with parents. we have secured funding for and recruited a transition care coordinator (tcc) to investigate this further. objectives: our objective is to assess and screen parents at the end of their child's treatment, and to develop interventions to support parents during this time and thereafter. design/method: after informed consent, a standardized questionnaire, the psychosocial assessment tool (pat . ), was administered to parents at end of therapy (t ), months later (t ) and year later (t ). the tcc provided "universal" intervention to all families with an end of therapy binder containing a treatment summary, follow-up roadmaps, information on late effects, and survivor scholarships. based on their pat . scores, some parents were provided intervention specific to symptoms (targeted intervention for scores - . ) or referred to a behavioral health specialist through the clinic social worker for counseling (for scores > ). results: analysis of pat data showed that % of parents (n = ) scored in the targeted or clinical ranges; % of parents scored in those ranges at pat . significant gender differences were revealed with the mean score for men of . and for women of . . this was confirmed by showing statistical significance (p = . ) when analysis was conducted for only a subgroup of data composed of couples (n = ). analysis of pat data by couples (n = ) showed the mean score for men was . and for women was . (p = . ). gender differences were most apparent in caregiver stress reaction questions that focused on ptsd symptoms. when the subgroup of couples' scores (n = ) for caregiver stress reaction at pat was analyzed, there was a significant difference (p = . ) in caregiver stress reaction with a mean of . for men versus . for women. [note: subcategory scores range from to ]. this study was initiated in october using a tcc and the pat . screening tool. the results suggest greater stress on mothers after therapy, with a substantial proportion of parents having symptoms of ptsd after therapy. background: hodgkin lymphoma (hl) is a common childhood cancer characterized by an inflammatory microenvironment. chemotherapy and radiation may exacerbate this inflammation and contribute to the development of late effects (pneumonitis or pulmonary fibrosis). in a heterogeneous cohort of childhood cancer survivors exposed to pulmonarytoxic therapy, no association between pro-inflammatory cytokines and late pulmonary dysfunction was observed. our objective was to test this association in a relatively uniform cohort of survivors of hl, given the well-recognized proinflammatory background of this disease. objectives: to characterize off-therapy pulmonary function in survivors of hl treated with contemporary therapy, and to investigate its association with persistent systemic inflammation. design/method: blood samples, clinical data, and pulmonary function tests were obtained from survivors of hl ≥ months off therapy. lung function score (lfs), a validated method for assessing degree of pulmonary dysfunction on a scale of i to iv, was determined from diffusion capacity and forced expiratory volume in one second (fev ). for a control group, blood samples from patients with benign, noninflammatory hematologic conditions were used. plasma concentrations of inflammatory cytokines were measured on a luminex platform (emd millipore). associations between clinical features or cytokine levels and lfs i (normal) vs. ii-iv were evaluated using logistic regression or wilcoxon rank sum tests, respectively. results: of survivors (mean age at diagnosis: years, range: - ; mean time off therapy: . years, range: . - ), % were categorized as lfs ii (mild dysfunction), % as lfs iii (moderate dysfunction), and no survivors as lfs iv (severe dysfunction). higher lfs was associated with female sex (p = . ) but not other demographic, disease, or treatment factors. forty-eight survivors had blood samples collected at a mean age of . years (range: - ) with a mean time since treatment completion of . years (range: . - . ). of controls, the mean age at time of blood collection was years (range: - ). survivors did not have significantly elevated cytokine levels compared to controls. female survivors of hl ≥ months off therapy are at increased risk of pulmonary dysfunction. neither evidence for pulmonary dysfunction, as measured by lfs, nor duration of time off therapy were related to systemic inflammation in this study. pulmonary function deterioration and clinical pulmonary symptoms are rarely observed immediately following therapy but increase over time. future studies may consider exploring the contribution of systemic inflammation to pulmonary late effects in survivors farther off therapy, when risk for this late effect is greater. background: thyroid carcinoma is a very rare tumor in pediatrics, accounting for . - % of childhood carcinomas in the united states and europe. we aim to detect the risk of second malignancies among pediatric thyroid cancer survivors. the cohort analysis consisted of pediatric cancer patients aged less than years diagnosed with a primary thyroid cancer and identified by site code icd- - : c , reported to a seer database between and . they were followed up by death or the end of the study period (december , ) . out of patients diagnosed primarily with thyroid carcinoma, there were patients who had incidences of subsequent malignancies. the mean age of patients at initial diagnosis of thyroid cancer was years. females ( . %) had significantly higher incidence of second malignancies (sm) than males ( . %). the overall standardized incidence ratio (sir) of sm in thyroid pediatric patients was higher than expected (sir = . ). some specific sites showed significantly higher incidences: salivary gland (sir = . ), gum and other mouth (sir = . ) and kidney (sir = . ). the overall risk of sm in patients received radioactive iodine was higher than expected (sir = . ). the cumulative inci-dence of sms from the initial diagnosis of thyroid cancer was calculated with the survival methodology of competing risk, death treated as a competing event. cumulative incidence of sm was . % [ % ci ( . , . %)] at years and substantially expanded after years, reaching . % [ % ci ( . , . %)] at years. the cumulative incidence of each tumor type at years was . % [ % ci ( . , . %)] for breast cancer, . % [ % ci ( . , . %)] for salivary gland, . % [ % ci ( . , . %)] for each one of kidney and cervix uteri and . % [ % ci ( , . %)] for each one of ovary and melanoma of the skin. cumulative incidence of sm was stratified based on race, gender and radiotherapy exposure, but there was no statistical difference in each of them. conclusion: race, gender, histological subtypes, and radioactive iodine may play an important role as prognostic factors for developing sm among pediatric thyroid cancer survivors. identification of underlying mechanisms that raise the risk of sm is important for both treatment and follow-up strategy. background: the ethical practice of informed consent requires it be both voluntary and understood by the research participant. in pediatric oncology, parents must undergo informed consent to enroll their child with cancer into clinical trials, but often it can be difficult to understand especially for parents with low english proficiency. previous research has shown that parents of children with cancer have difficulty understanding voluntariness, and that parental satisfaction with informed consent does not always correlate with adequate comprehension. objectives: to examine socio-demographic and contextual correlates of comprehension of informed consent, voluntariness, and satisfaction in parents who consented to participation of their child in a cancer clinical trial. we focused on characterizing differences between non-hispanics and hispanics, the fastest growing ethnic group in the u.s. design/method: parents/guardians (n = ) of children aged - years with newly diagnosed cancer, who had consented to participation of their child in a clinical trial for cancer treatment at rady children's hospital-san diego were s of s prospectively recruited. parents completed questionnaires assessing comprehension, voluntariness, satisfaction, health literacy, socio-demographics, and acculturation level, if hispanic. comprehension was surveyed at baseline and longitudinally at months. comprehension, voluntariness and satisfaction outcomes were analyzed by socio-demographics, health literacy, and acculturation level using logistic regression. results: of the participants surveyed, ( . %) were hispanic and ( . %) were non-hispanic. we found that higher health literacy was associated with greater objective comprehension (p< . ), voluntariness (p< . ), socioeconomic status (p< . ), and acculturation (p< . ). hispanics reported lower objective comprehension (p = . ), voluntariness (p = . ), health literacy (p< . ) and ses (p = . ) compared to non-hispanics. spanish-speakers reported lower voluntariness (p = . ), health literacy (p< . ), and acculturation (p< . ) compared to englishspeakers. at the -month follow-up, comprehension in hispanics significantly improved (p = . ) compared to their baseline comprehension. satisfaction was moderately high across all subgroups and was not significantly impacted by socio-demographics, health literacy, or acculturation. in this study, with equivalent numbers of hispanic and non-hispanic participants, we found that hispanic and spanish-speaking parents of children with newly diagnosed cancer had inadequate informed consent comprehension, voluntariness and health literacy despite high satisfaction. our study suggests that hispanics and individuals with limited english proficiency are not making truly informed decisions for their child with cancer. to ensure the ethical practice of research in pediatric oncology, the informed consent and decision-making process must be improved with culturally and linguistically interventions for these underserved populations. memorial sloan kettering cancer center, new york, new york, united states background: pediatric oncology patients undergo repeated bone marrow aspirations and biopsies (bma/bx). these potentially painful procedures can exacerbate anxiety and distress. standard practice at memorial sloan kettering (msk) department of pediatrics is to use propofol, which has amnestic but no analgesic properties. we sought to evaluate whether the addition of local anesthetic would improve patient experience with bma/bx. the purpose of reppair: reducing procedural pain and improving recovery of quality of life (qol) (nct ) is to evaluate the efficacy of local anesthesia with ropivacaine in reducing procedural pain and improving post-procedure qol in pediatric neuroblastoma patients undergoing bma/bx with general anesthesia. reppair is a prospective, randomized, crossover clinical trial that opened for enrollment october . eligible patients were - years old with neuroblastoma. participants were observed on trial for two sequential bm procedures; one procedure with intervention a: propofol alone (pa), and the other with intervention b: propofol plus ropivacaine (p+r). participants were randomized to intervention sequence ab or ba and were blinded to the order of interventions. participants and recovery room (rr) nurses, who were also blinded, followed a standardized postprocedure pain management algorithm. the primary endpoint was percentage of participants requiring opioid analgesia in the hours post-procedure. secondary endpoints included total opioid in hours, non-opioid analgesia use, pain scores, time to first opioid, and short-term qol. qol was assessed by a parent-proxy metric that evaluated pain interference with sleep, physical, emotional, and social recovery. as of january , patients were assessed for eligibility and patients were randomized ( have completed both procedures). for the primary endpoint, a slightly higher proportion of participants required opioid for pa than p+r ( % versus %, p = . ). pain scores in the rr were significantly higher for pa than p+r (median [ th, th percentile]: [ , ] versus [ , ], p = . ). there were no statistically significant differences in total opioid or non-opioid analgesia, -and -hour pain scores, median time to first opioid, or pain interference scores. there were no adverse events. conclusion: preliminary findings of the reppair trial suggest that local anesthesia does not reduce the need for opioid analgesia or improve short-term qol in pediatric patients undergoing bma/bx with general anesthesia. local anesthesia did improve pain scores in the immediate recovery period. final results of this study will help establish evidence-based guidelines and optimize the experience of pediatric patients with bone marrow procedures at our center. background: children with advanced cancer experience a range of symptoms throughout treatment or at end of life, some of which are poorly controlled. minimizing suffering, including effective symptom management, in children with advanced cancer is a central value for pediatric oncology clinicians. patient-reported outcomes have been used in symptomrelated research in pediatric oncology patients; however the majority of literature specific to symptoms during palliative care and end of life for children and adolescents with advanced cancer is based primarily upon medical record reviews and to a lesser extent, patient self-report. the purpose of this study was to prospectively describe symptom frequency, severity, and level of distress in children/adolescents with advanced cancer using patient selfreport and parent proxy. design/method: a prospective cohort design was used for this study. five pediatric oncology institutions from across the united states participated. children and adolescents were eligible to participate if they were - years of age, englishspeaking, and had a diagnosis of advanced cancer, defined as a -week history of progressive, recurrent, or non-responsive disease or a decision not to pursue curative-focused therapy. a modified version of the memorial symptom assessment scale (msas) was used to measure symptom frequency, severity, and level of distress and was administered to child/parent dyads electronically via smartphones every two weeks. information regarding disease status and cancer treatment was collected concurrently. data was analyzed using descriptive statistics and univariate logistic regression analysis. results: a total of children and adolescents and parents participated in the study. the median age of child participants was years, with half being male. the median age of parents was years. the child participants had a variety of primary diagnosis, including: leukemia/lymphoma (n = , %), solid tumor (n = , %), and brain tumor (n = , %). the most frequently reported symptoms by children with advanced cancer and parents were pain (n = / , . %), lack of energy (n = / , . %), and nausea (n = / , . %). presence of disease (p = < . ), recent disease progression (p = . ), and receiving cancer therapy (p = . ) were significant factors on the presence of pain. high intensity cancer therapy was a significant factor on pain frequency (p = . ) and level of distress (p = . ). it is feasible to collect data prospectively in children with advanced cancer regarding symptom frequency, severity, and level distress. clinicians' increased understanding of the symptom experience may promote communication with children and adolescents and timely intervention. more research is needed to understand symptom clusters in children with advanced cancer. vanderbilt children's hospital, nashville, tennessee, united states background: febrile neutropenia (fn) is a frequent occurrence in children undergoing chemotherapy. though guidelines recommend adding a second antibiotic to broad-spectrum antipseudomonal coverage in specific scenarios, augmenting empiric therapy with a second antibiotic is common practice. additional empiric antibiotic (aea) use increases the risk of antibiotic toxicity and future antimicrobial resistance. data clarifying the indications for aea are limited in pediatric patients. objectives: to identify risk factors for gram-positive (gp) and gram-negative (gn) bacteremia in patients presenting with fn to determine situations in which aea use is warranted. design/method: a retrospective chart review was conducted of pediatric severe fn with absolute neutrophil count < / l occurring at a single institution between and . potential a priori risk factors based on clinical reasons for antibiotic expansion were chills, hypotension, mucositis, skin or soft tissue infections (sstis), recent administration of highdose cytarabine (hdac), and a diagnosis of acute myeloid leukemia (aml). potential factors for gn bacteremia were chills, hypotension, mucositis, and abdominal pain. the association between each potential risk factor and gp or gn s of s bacteremia was identified. logistic regression was used for multi-variable analysis. the review yielded episodes. gp bacteremia was isolated in cases ( . %) and gn bacteremia in episodes ( . %). in multivariable analysis, hypotension (or . ( % ci . , . ), p = . ) and sstis (or . ( . , . ) , p = . ) were independently associated with increased risk of gp bacteremia, while mucositis (p = . ), recent administration of hdac (p = . ) and chills (p = . ) were not. ten patients with aml didn't receive hdac, thus the association between aml and gp bacteremia could not be reliably estimated. hypotension (or . ( . , . ), p< . ) and chills (or . ( . , . ), p< . ) were independently associated with a higher risk of gn bacteremia, while mucositis (p = . ) and abdominal pain (p = . ) were not. of the gn infections, ( %) were resistant to cefepime, the empiric agent of choice at our institution. patients with fn with sstis, hypotension, or recent hdac had increased risk of gp bacteremia indicating potential benefit of empiric vancomycin in these settings, while mucositis and chills were not associated with gp bacteremia. hypotension and chills were associated with gn bacteremia, potentially warranting empiric antibiotic expansion, while mucositis and abdominal pain were not. identifying specific indications for aea use in pediatric severe fn use may improve antimicrobial utilization, decrease unnecessary antibiotic use, and improve patient outcomes. background: for children/young adults with incurable high grade gliomas (hggs), like diffuse intrinsic pontine glioma (dipg) or glioblastoma multiforme (gbm), oncologists endeavor to align therapy with patient/family goals of care, but may be influenced by providers' preferences or limited resources. ethical challenges can arise around the perceived purpose, risks and benefits of therapy options, provider conflicts of interest, access to care, deciding decisional priority between patients and families, and conflicts around end-oflife care. objectives: evaluate factors that play into longitudinal decision making for children and young adults with hggs, their families and oncologists using a qualitative approach with ethnographic elements. design/method: eligible patients were aged - with dipg, gbm, or secondary hgg. patient exclusions included: non-english speaking, in state custody, death prior to diagnosis, seen by oncology once, or an oncologist declined participation. key decision making visits (e.g. mri reviews) were serially audio-recorded, along with subsequent : semistructured interviews with patients and/or parents about the decision making process. field notes from clinician meetings, chart notes, and oncologist questionnaires were obtained. discussions and interviews were transcribed and independently coded by three investigators. inter-rater reliability was assessed during code book development. discrepancies were discussed until consensus met. constant comparison analysis with maxqda software continued until thematic saturation. results: twenty-two of eligible patients were approached; agreed to participate. one withdrew upon transferring care. mean age was . years (sd . ); % male, % caucasian, % african american, % hispanic, and % asian. four encounters, ( . hours), were recorded on average per patient. parent/patient interview themes included: ) hope (for a cure, prolonged life, and quality of life), ) importance of physician recommendations, ) importance of support systems (family, community, social media), ) food (as cancer etiology, intervention) ) finances (personal, research funding), ) communication (with medical providers, family, community), ) death, and ) god (beliefs, prayer, existential questions). oncologists desired prolonged quality of life, while patients/families transitioned to that hope from hope for a cure. decisions made in the setting of hggs are multi-factorial, ultimately reflecting the competing values of decision makers. optimism about treatment efficacy is held in tension with poor prognosis, allowing for functional hope. acknowledging patients' and families' shifting hopes allows for changes in goals of care and shared decision making. future work is needed to ) develop preference tools for pediatric patients and families to inform medical providers and ) provide training in communication and shared decision making with oncologists. emory university, atlanta, georgia, united states background: bone marrow transplantation (bmt) is a potentially curative but underutilized treatment for scd. our previous work has shown that there is variation in physician philosophy and practice in considering bmt as a treatment option for patients with scd, and physicians may not discuss this with patients and families as a potential treatment option. in a randomized clinical trial to test the effectiveness of a decision aid for disease modifying therapies for sickle cell disease, adult patients with scd as well as caregivers of adult/pediatric patients were interviewed about how they seek or have sought information related to scd, made decisions about treatments for scd, and identified a treatment option they were interested in learning more about using the decision aid tool. we performed a secondary analysis of these baseline data to understand patient information needs and attitudes regarding bmt as a treatment option for scd. the goals of this analyses was to understand patient and caregivers' attitudes and perceived information needs regarding bmt as a treatment option for scd. we performed an analysis of baseline interviews from caregivers of patients with scd or adult patients from a randomized control trial for a decision aid tool for scd. of the interviews belonged to caregivers of patients with scd. in addition to reviewing interviews for discussion of bmt, we interrogated for mention of terms such as 'bone marrow transplant' or 'cure' or 'stem cell transplant'. interviews were coded using nvivo and analyzed for emerging themes. results: of the baseline interviews, interviews met selection criteria. thirteen of the interviews were with caregivers of pediatric patients, and the remainder were with adult patients, including young adult patients with scd. the majority of participants want to learn about bmt or curative options. in many participants, this was expressed despite knowledge that they were not a likely candidate for transplant. desired information about bmt included eligibility, benefits, risks, long-term effects, quality of life and financial aspects related to bmt. of the patients who discussed how they learnt about bmt, approximately half mentioned that their healthcare provider had not previously mentioned this to them. we then examined knowledge of bmt and attitudes with demographic and clinical variables. patients and caregivers of pediatric patients with scd want to learn about bmt as a treatment option. healthcare providers should consider discussing bmt with their patients with scd. natasha frederick, anna revette, alexis michaud, jennifer mack, sharon bober dana-farber cancer institute, boston, massachusetts, united states background: adolescents and young adults (ayas) consistently identify the need for improved patient-clinician communication on sexual and reproductive health (srh) issues. however, oncology clinicians do not routinely integrate srh conversations with ayas through disease treatment and survivorship. little is known about why these conversations do not take place. objectives: explore aya perceptions of and receptiveness to srh communication with oncology clinicians and to identify barriers and facilitators to these conversations. design/method: semi-structured interviews were held with aya cancer patients and survivors (ages - years, men, women). twelve participants were on active treatment and were within years of treatment completion. interviews were conducted in english by phone or in person. the interview transcript underwent pre-testing with ayas. all interviews were audio-recorded and transcribed verbatim. transcripts were analyzed and summarized by two trained qualitative researchers according to standard comprehensive thematic qualitative analysis methods. analyses were aided by nvivo software. results: ayas perceived existing srh communication between ayas and oncology providers as inadequate. all ayas reported a need for improved srh communication with oncology providers, and three key areas of need emerged: ) general education; ) addressing specific srh issues experienced during treatment and survivorship; and ) understanding the long-term impact of cancer and treatment on srh. ayas felt that current srh discussions are limited and too narrow in scope and scale. ayas reported that most srh conversations focus exclusively on fertility (n = ), usually taking place at the start of treatment. other additional yet limited communication reported was about sexual activity (n = ), contraception (n = ), sexual function (n = ). no ayas reported conversations about potential treatment complications related to sexuality other than infertility. key barriers to srh conversations include patient discomfort initiating conversation (n = ) and presence of family members (n = ), with additional reported barriers including perceived provider discomfort (n = ), lack of rapport with provider (n = ), and age/gender differences (n = ). ayas felt that s of s communication tools such as handouts, brochures, and websites would be helpful facilitators to direct communication from the oncology clinician, and wanted conversations to start before treatment initiation and to continue through treatment and survivorship conclusion: ayas identify a key role for pediatric oncology providers in srh care from diagnosis through survivorship, however multiple barriers interfere with discussions about srh on a regular basis. identified barriers suggest that future efforts should focus on provider education and training in srh and srh-related communication in order to optimize care provided to this unique patient population. background: peripherally inserted central venous catheters (picc) provide secure vascular access in pediatric patients for the delivery of necessary therapies. the ease of placement in the inpatient and outpatient settings has expanded their utilization. however, recent data analyses show a significant increase in venous thromboembolism (vte) risk with the use of picc lines. with its rising use, modifiable risk factors need to be understood for preventative measures. objectives: in this study we aim to understand patient and catheter specific characteristics in relation to the development of vte. design/method: with irb approval, a retrospective interrogation of the electronic medical record and a picc database, at rainbow babies and children's hospital, was completed. the study cohort contained patients < years of age who had a picc line placed between january of and december of . data collected included indication for line placement, line dwell time, location of insertion including blood vessel and extremity, number of attempts at line placement, lumen size and indwelling line length. in addition, we collected number of days to vte formation, associated symptoms and location of vte. chi-squared analyses and fischer's exact test were used where appropriate for statistical analysis. we analyzed ( neonatal) newly placed picc lines. fifty line-associated vte events were found, for an incidence of . %. all vte occurred with the placement of the first picc line. intravenous therapies were the most common reason for line placement. no statistical significance was found between various indications for placement. the most common symptom of vte manifestation was extremity swelling, follow by extremity pain. right extremity picc was found to have a higher incidence of vte. larger catheter lumen sizes (> french) had a higher incidence of vte. we found a mean time of . days to vte detection. we were unable to find any clinical, patient or line specific factors leading to increased vte formation after statistical analysis. special consideration should be given to the duration of picc line use as this may reduce the incidence and comorbities associated with vte. there is still much to be understood about catheter associated vte formation as our analyses indicates the need for prospective data collection on a larger scale in hopes to create guidelines related to catheter use in pediatrics. background: the decision to transfuse a patient is a complex one and is never based solely on a number; however, certain hemoglobin or platelet count thresholds have been proposed in aiding physicians make transfusion decisions. in our hospital, the thresholds for packed red blood cell (prbc) and platelet transfusion in pediatric oncology patients are hemoglobin levels below . g/dl and platelet counts below , /mm (< , for brain tumors), respectively. recently, these thresholds have been questioned and we were asked whether we could safely lower the thresholds to < . g/dl of hemoglobin and < , /mm platelet count objectives: to investigate platelet and hemoglobin transfusion thresholds for oncology patients at children hospital of michigan design/method: retrospective chart review over a -month period, examining platelet and hemoglobin pretransfusion levels for each prbc and platelet transfusion given to oncology patients results: over the course of months, eligible oncology patients (median age years) received transfusions ( prbc transfusions and platelet transfusions). the mean pretransfusion hemoglobin level was . ± . g/dl (range . - . ) (n = ) for total prbc transfusions and this was not different among disease categories (p = . ). patients who had anemia symptoms and signs (n = ) had a slightly lower hemoglobin level compared to those who did not (n = ): . ± . vs . ± . g/dl (p = . ). the mean pretransfusion platelet count was , ± , /mm (range , - , ) for total platelet transfusions (n = ); , ± , /mm in patients with brain tumors (n = ); , ± , in patients with leukemia (n = ); and , ± , in patients with solid tumors (n = ). the mean pretransfusion platelet count was significantly higher in transfusions for brain tumors compared to that in the other disease groups (p< . for both). the mean pretransfusion platelet count was not different among those patients who had bleeding/bruising symptoms ( , ± , , n = ) versus those who did not ( , ± , , n = ) (p = . ). the bleeding/bruising rate was slightly but insignificantly higher in those who had platelet counts < , vs those who had ≥ , ( . % vs . %, p = . ). since most patients develop symptoms of anemia at hemoglobin above g/dl and about / of patients develop bleeding/bruising symptoms at platelet counts above , /mm , our current policy so far reflects a safe threshold for transfusion, and further lowering of the thresholds should be investigated in prospective studies. background: renal impairment is an important complication of childhood cancer and its treatment. serum creatinine level is frequently used as a screening test to monitor renal function; however, patients can have significantly decreased glomerular filtration rate (gfr) with normal serum creatinine. to determine the prevalence of chronic kidney disease (ckd) among children with cancer diagnosis, based on calculated gfr. to compare the difference between using serum creatinine value alone versus gfr in detecting ckd. design/method: retrospective review of medical records of patients, age - years, diagnosed between / - / with solid tumors were analyzed. serum creatinine and calculated gfr using schwartz formula were recorded. ckd as classified by the foundation of kidney disease and outcome quality initiative was used: ckd stage : gfr ( to ml/min per . m ) ckd stage : gfr ( to ml/min per . m ) statistical analysis using spss software v. . chi-squared test for proportions within group, and pearson chi-squared and fisher exact tests for statistical differences between groups. p-value < . was considered to indicate significance results: out of the records reviewed, ( %) were males and ( %) females, with mean age of . ± . years. ( . %) patients received one or more of nephrotoxic chemotherapy drugs; cisplatinum, carboplatinum, or ifosphamide mainly in the non-wilms solid tumors group ( . %) compared to ( . %) in the wilms tumor (wt) group. based on calculated gfr (by schwartz formula) ckd stage /or was diagnosed in ( %) patients with overwhelming majority ( %) were in the mild stage ckd, only ( . %) of those patients had abnormally high serum creatinine levels (p = . ). . % of patients who received nephrotoxic chemotherapy developed ckd, compared to . % in those who did not receive it, (p = . ). despite that only / ( %) of wt group patients received nephrotoxic chemotherapy, yet this group had higher percentage of ckd ( . %) compared to non-wt group ( . %) p = . . significantly lower mean gfr . ± was noticed in the wt group compared to . ± in non-wt group (p = . ) conclusion: high prevalence of mild ckd was found among solid tumor patients. using serum creatinine alone as measure of renal function significantly under estimates renal impairment in those patients. early identification of ckd is easily achieved by using calculated gfr, which can helps providers and care givers to avoid potential nephrotoxic antibiotics, contrast media, nsaids and dehydration that may further deteriorate renal function the university of texas southwestern medical center, dallas, texas, united states background: children with down syndrome (ds) have increased risk of developing leukemia. pediatric patients with ds-associated acute lymphoblastic leukemia (ds-all) are known to have significant toxicities with reinduction chemotherapy and historically poor outcomes with stem cell transplant (sct). anti-cd chimeric antigen receptor (car) t-cell therapy, tisagenlecleucel, demonstrated high rates of durable complete remission (cr) and a manageable safety profile in children with r/r b-cell acute lymphoblastic leukemia (b-all). objectives: characterize the efficacy and safety of tisagenlecleucel in pediatric/young adults with ds-all. design/method: pooled data from single-arm, multicenter, phase trials of tisagenlecleucel in pediatric/young-adult patients with r/r b-all (eliana, nct ; ensign, nct ) were analyzed. eight patients with ds-all were enrolled (data cutoff: eliana, november ; ensign, february ). seven were infused with tisagenlecleucel; patient died from all progression and intracranial hemorrhage before infusion. no manufacturing issues occurred during production. / infused patients were male, / had prior sct (age range, - years). / patients achieved cr or cr with incomplete blood count recovery (cri) by day (d) (cr+cri, %); died before d and was not evaluable. analysis of minimal residual disease was negative in bone marrow in responding patients. two patients had cd negative relapses at and months. ongoing remissions in patients without relapse ranged from to months. the safety profile (n = ) appears similar to that in patients without ds in the same trials (n = ). grade (g) / cytokine release syndrome occurred in % ( / ) of patients with ds and in % without ds. rates of other g / adverse events of special interest did not appear to favor a consistent trend between patients with/without ds (febrile neutropenia: % vs %; neurological events: % vs %; tumor lysis syndrome: % vs %). g / infections were not observed in patients with ds ( % vs %). one patient died after infusion due to intracranial parenchymal hemorrhage on d associated with ongoing coagulopathy. time and extent of tisagenlecleucel expansion and long-term persistence were similar between groups. conclusion: this is the first analysis of car t-cell therapy in pediatric patients with r/r b-all and ds. these data suggest that toxicities appear similar to those in patients with b-all without ds, remission rates in ds-all are high, and longterm outcomes with sustained persistence appear promising. further exploration of tisagenlecleucel as an alternative to sct in children with r/r ds-all is warranted. sponsored by novartis. background: hispanic adolescence and young adults are twice as likely to develop acute lymphoblastic leukemia (all) with high risk features as non-hispanic whites. they also have poor prognosis and % higher death rate. b-all with crlf overexpression caused by genetic alteration of the cytokine receptor, crlf is five times more common in this subgroup. approximately % of crlf b-all cases also have ikzf genetic alterations. ikaros is involved in transcriptional regulation of several important genes involved in leukemogenesis. overexpressed casein kinase ii (ck ) impairs functions of ikaros. objectives: understand the molecular mechanisms that regulate crlf expression in crlf b-all. here we present evidence that ikaros-mediated repression of crlf transcription in b-all in hispanic children is regulated by ck . design/method: primary b-all patient samples from hispanic children were used. ikaros retroviral transduction, ikaros shrna transfection, real time-pcr, luciferace assay, quantitative chromatin immunoprecipitation (qchip) coupled with the next-generation sequencing (chip-seq), cytotoxicity assay and western blot. results: ikaros binding to promoter of crlf was confirmed using quantitative chip. functional experiments such as overexpression of ikaros in b-all primary cells results in transcriptional repression of crlf whereas ikaros silencing using shrna resulted in increased transcription. these results suggest that ikaros negatively regulates crlf expression. molecular inhibition of ck with shrna targeting the ck catalytic subunit, as well as pharmacological targeting of ck with cx resulted in transcriptional repression of crlf . ck inhibition was associated with increased ikaros dnabinding to the promoter of crlf . however, the ability of cx to repress crlf is lost or severely reduced, in cells with shrna silencing of ikaros, as compared to cells with intact ikaros. moreover, similar results were noted following treatment with cx in leukemia cells obtained from high risk b-all patients with deletion of one ikzf allele. ikaros binds poorly to promoters of crlf gene in these cells. treatment with cx restores ikaros dnabinding to the promoters of crlf , which is associated with its strong repression. serial qchip analysis of the epigenetic signature at the crlf promoter showed that increased ikaros binding to the crlf promoter, following ck inhibition, is associated with enrichment for the h k me histone modification, which is a marker of repressive chromatin. results demonstrate that crlf expression is epigenetically regulated by the ck -ikaros axis .cx show antileukemic effect via restoration of ikaros tumor suppressor function, resulting in crlf repression suggesting advantage of using ck inhibitors as potential therapeutic approach in crlf altered b-all. results: hypodiploid all (modal chromosome number < and/or di < . ) was identified in patients ( . % of all patients; . % of nci standard risk (sr) and . % of nci high risk (hr)), who were removed from frontline protocol therapy post-induction. overall -year efs and os were . %± . % and . %± . %. transplant status was retrospectively available for / ( %), of whom underwent hsct in cr . five-year efs with hsct was . %± . % vs. . %± . % without (p = . ). -year os with and without hsct was . %± . % vs. . %± . % (p = . ). when corrected for the median time to hsct ( days), there were no significant differences in -year efs or os rates with and without hsct: . %± . % and . %± . % vs. . %± . % and . %± . %. no nci risk group or mrd subset benefitted significantly from cr hsct. sr patients (n = ) had -year efs and os of . ± . % and . %± . % with hsct (n = ) vs. . %± . % and . %± . % without. hr patients (n = ) had -year efs and os of . %± . % and . %± . % with hsct (n = ) vs. . %± . % and . %± . % without. for those with end-induction mrd < . % (n = ), -year efs and os were . %± . % and . %± . % with hsct (n = ) vs. . %± . % and . %± . % without. end-induction mrd-positive patients (n = ) fared poorly with both year efs and os of . %± . % with hsct (n = ) vs. . %± . % and . %± . % without. multivariate regression analysis including nci risk group, mrd, and cr hsct, showed only mrd negativity was significantly associated with efs (hr . , p< . ) and os (hr . , p< . ). patients with hypodiploid all fare poorly, particularly those with end-induction mrd ≥ . %. while cr hsct is a standard treatment approach, it does not confer significant benefit. we were unable to assess bridging therapy prior to hsct, and comparator groups are small. taken together, however, new strategies are urgently needed for these patients. background: ras-pathway mutations are known to play a pivotal role in a significant proportion of myeloid malignancies, including upwards of % of pediatric aml cases. ras-pathway mutations in myeloid malignancy commonly co-occur with mutations of epigenetic regulators, suggesting cooperative leukemogenesis. among the epigenetic modifiers most frequently mutated in myeloid malignancy are regulators of dna methylation. this indicates that the alteration of dna methylation contributes to leukemogenesis. the ten-eleven translocation (tet ) is an epigenetic regulator that plays an important role in regulation of dna methylation through its action of hydroxylation of -methylcytosine, which ultimately leads to passive de-methylation of dna cytosines. in myeloid malignancy, loss of function tet mutation is one of the most frequently co-occurring lesions in ras mutated malignancy. how specifically the altered methylation patterns in ras-pathway driven diseases promotes leukemogenesis is unclear. objectives: we hypothesize in mice with a ras-pathway mutation, that when an epigenetic modifier co-occurs, such as loss of function of tet , this primes stem cells and/or early differentiating progenitors for transformation by preventing the repression of stem cell self-renewal genes, inhibiting differentiation, enhancing ras signaling and leading to leukemogenesis. we have generated a novel murine model with constitutive deletion of tet (tet -/-) combined with an inducible activating krasg d mutation (krasg d/wt). mice have been tracked for evidence of hematologic malignancies and compared to mice with corresponding single genetic lesions. cooperative leukemogenesis will be demonstrated by decreased latency to disease onset, impact on malignancy lineage, in addition to investigating mechanistically through which pathways leukemogenesis may be promoted. results: krasg d/wt/ tet -/-mice demonstrate statistically significant differences in peripheral white blood cell count, hemoglobin, and platelet levels as early as -weeks post ras-pathway activation. peripheral cell lineage analysis demonstrates early skewing toward myeloid differentiation and marked splenomegaly in mice harboring both genetic lesions compared to wild type or mice with single genetic lesions. phospho-flow cytometric analysis reveals increased perk and ps activation in krasg d/wt/ tet -/-sca- enriched bone marrow cells compared to either genetic lesion alone. our study utilizing a murine model to examine how in ras-pathway mutations the addition of a co-occurring epigenetic lesion demonstrates that these lesions appear to cooperate to promote early myeloid differentiation with attendant changes in signaling pathways. this exploration to elucidate the mechanics of ras-pathway mediated disease lay the foundation for identification of patients who may benefit from existing therapies, such as dmtis, or identify new signaling targets for therapeutic exploration. background: the humoral immunogenicity of car , a chimeric antigen receptor (car) with a murine scfv domain developed for treatment with tisagenlecleucel in relapsed/refractory (r/r) pediatric/young-adult acute lymphoblastic leukemia (all), was evaluated in studies. little is known about the presence/impact of preexisting/treatmentinduced anti-murine car (mcar ) antibodies in patients treated with car therapy. objectives: patients from eliana (nct ; n = ) and ensign (nct ; n = ) were evaluated before and after tisagenlecleucel infusion to determine the impact of anti-mcar antibodies on cellular kinetics, efficacy, and safety. design/method: anti-mcar antibodies were determined by flow cytometry and reported as median fluorescence intensity. assay validation included evaluation of the interferences of intravenous immunoglobulin (ivig) treatment with the anti-mcar antibody assay. impact of preexisting and treatment-induced immunogenicity on cellular kinetics, efficacy, and safety was determined. treatment-induced immunogenicity was defined by a positive increase in anti-mcar antibody levels over baseline and was assessed by calculating the fold-change between preexisting (ie, baseline) and postinfusion levels. results: % of patients displayed preexisting anti-mcar antibodies; a similar incidence was detected in healthy volunteer samples during method validation. % of patients developed treatment-induced anti-mcar antibodies. no relationship was identified between tisagenlecleucel expansion (auc - d) and preexisting/treatment-induced anti-mcar antibodies (r < . and r = . , respectively); similar results were seen for cmax. presence of treatment-induced anti-mcar antibodies did not appear to impact transgene persistence or response. kaplan-meier estimates showed that preexisting/treatment-induced anti-mcar antibodies did not appear to impact duration of response or event-free survival. strip plots showed consistent levels of preexisting/treatment-induced anti-mcar antibodies across patients with safety events, including cytokine release syndrome, neutropenia, thrombocytopenia, and neurological events. there was no apparent relationship between treatment-induced anti-mcar antibodies and b-cell recovery categories (≤ months, > and ≤ months, > months, and ongoing sustained aplasia). no association existed between time of b-cell recovery and presence of treatment-induced anti-mcar antibodies. b-cell aplasia requiring ivig occurred following tisagenlecleucel in the majority of patients. the tisagenlecleucel concentration-time profiles in patients with treatment-induced anti-mcar antibodies were categorized by time following ivig administration. time of ivig administration had no impact on in vivo transgene expansion and persistence. we report the first comprehensive assessment of the impact of anti-mcar antibodies on clinical endpoints with car therapy. pediatric/young-adult patients with r/r all had a high frequency of baseline anti-mcar antibodies, and preexisting/treatment-induced anti-mcar antibodies did not impact the cellular kinetics, safety, and efficacy of tisagenlecleucel. cell-mediated immunity studies are ongoing. sponsored by novartis. background: adoptive immunotherapy, using cd engager (cd -eng) t-cells, has shown success in preclinical studies, recognizing and killing acute myeloid leukemia (aml) blasts in vitro and in vivo. cd -eng t-cells secrete bispecific molecules that recognize cd (t-cells) and cd (aml blasts), and are able to direct transduced t-cells and recruit bystander t-cells to kill cd -positive blasts. however, cd -engs do not provide costimulation and have not shown the capability for sequential killing of targets in vitro. we are seeking to improve the expansion, persistence and sequential killing capabilities of cd -engs by genetically modifying these cells with an inducible costimulatory molecule, which can be activated by a chemical inducer of dimerization (cid). we generated a retroviral vector encoding cd -eng and the inducible costimulatory molecule myd .cd linked by a a sequence (cd -eng. a.imc). cd -eng and cd -eng.imc t-cells were generated by retroviral transduction, and their effector function was compared with and without cid. we used flow cytometric analysis to assess transduction efficiency, chromium release assays to evaluate cytolytic activity, and elisa to determine cytokine production. we successfully generated cd -eng.imc tcells and achieved a mean initial transduction efficiency of % that was maintained above % throughout our study period. cd -eng.imc t-cells +/-cid and cd -eng t-cells readily killed cd -positive aml blasts (molm and kg a) in cytotoxicity assays when compared to the cd -negative control (k ). in co-culture assays, cd -eng.imc t-cells secreted increased il- and ifn-gamma in the presence of cid and cd -positive targets (kg a and molm ) when compared to co-culture with cd -positive targets in the absence of cid. in addition, cd -eng.imc t-cells displayed enhanced sequential killing capabilities and ifn-gamma secretion when stimulated weekly with cid and tumor cells at a : ratio when compared to cd -eng t-cells. conclusion: cd -eng.imc t-cells are able to recognize and kill cd -positive aml blasts in an antigen dependent manner. cd -eng.imc t-cells have improved effector function in the presence of cid as judged by cytokine production and their ability to sequentially kill cd -positive target cells. thus, inducible myd and cd costimulation is a promising strategy to improve the effector function of cd -eng t-cells, and warrants further active exploration in preclinical studies. background: eliana (nct ; n = ) is a pivotal multicenter study testing the efficacy of tisagenlecleucel, anti-cd car-t, in children/young adults with r/r b-all. tocilizumab (toci) has been used for management of moderate/severe (grade / ) crs in ≈ % of patients treated with tisagenlecleucel at equivalent doses used in approved nononcological pediatric indications (< kg received mg/kg; ≥ kg received mg/kg [ mg max dose]).( ) crs onset, as graded by the penn grading scale, generally occurred at a median of days (range, - ) after infusion, requiring administration of - toci doses in some patients via a protocol-specific treatment algorithm. toci is a humanized monoclonal antibody that inhibits il- receptor (il- r) signaling. the pharmacokinetics (pk) and pharmacodynamics (pd) of toci in pediatric patients with b-all with carassociated crs have not previously been described. objectives: characterize toci pk/pd for crs management following tisagenlecleucel infusion and describe its impact on cellular kinetics. design/method: toci pk and levels of soluble il- r (sil- r) were determined from serum and quantified using validated assays. maximum toci concentration (cmax) was derived using noncompartmental methods. sil- r, proinflammatory cytokines, and crs resolution time were characterized to describe toci pd. summary statistics and graphical analyses of tisagenlecleucel exposure by number of doses were performed to describe the impact of toci on tisagenlecleucel kinetics in patients responding to tisagenlecleucel infusion. : / patients with crs received the first toci dose at a median of days (range, - ) after crs onset. seventeen patients received dose (range, . - mg/kg); received doses ( - mg/kg); received doses ( - mg/kg), per the crs treatment algorithm. first-dose mean cmax (sd) was ≈ ( . ) g/ml; second dose, ≈ ( ) g/ml. individual patient pd concentration-time profiles showed increased sil- r levels after the first toci dose which remained elevated following the second dose. following toci administration, median time to crs resolution (including fever resolution) was days (range, - ). crs onset coincided with tisagenlecleucel expansion, followed by a peak in serum cytokines, including il- . the geometric mean auc - day and cmax of tisagenlecleucel transgene (by pcr) were % and % higher in tisagenlecleucel-responding toci-treated patients. conclusion: crs symptoms resolved within a median of days after toci administration. toci levels achieved in patients with b-all were similar to reported pediatric nononcological indications (tocilizumab label) and resulted in concentration/time-dependent sil- r increases. transgene continued to expand and persist following toci administration. these data support treatment with toci for crs management. ( ) buechner, eha, . sponsored by novartis. background: in acute myeloid leukemia (aml), mesenchymal stem and stromal cells (mscs) in the bone marrow microenvironment contribute to extrinsically mediated chemo-resistance and are therefore important potential therapeutic targets. the study of patient-derived mscs is at a competitive disadvantage, however, because traditional means of isolating mscs from a bone marrow aspirate interferes with isolating the more highly prioritized leukemic cells. many opportunities to study mscs are therefore missed. objectives: to develop a novel method of isolating mscs using the otherwise discarded portion of a bone marrow aspirate, thereby de-coupling the isolation of primary mscs from the isolation of leukemia cells. design/method: aml patient bone marrow aspirates were obtained prospectively from the children's oncology group. healthy patient marrow was purchased. experimental mscs were isolated from the bottom-most layer (rbc-layer) produced by density-gradient separation of a bone marrow aspirate, which is typically discarded. control mscs were isolated from the buffy coat (mnc layer). non-adherent cells were removed after hours, and adherent cells were cultured at % co with mem-alpha containing % fbs. growth curves were obtained by seeding -well plates with , cells per well. cells were stained using oil red o to observe adipocyte differentiation. results: rbc-layer mscs grow successfully following overnight shipment of the aspirate. identical to mnc-layer mscs, rbc-layer mscs exhibit a fibroblastic morphology and are adherent to plastic. rbc-layer mscs persist in culture up to passages before senescence. they exhibit a slower growth curve relative to mnc-layer mscs, but their overall doubling time is similar at approximately hours. surprisingly, mscs from the rbc-layer exhibit adipocyte differentiation on stimulation, revealing their stem-cell like qualities. we present a method of isolating mscs from the discarded portion of a bone marrow aspirate that does not interfere with the isolation of leukemia cells from the same patient. this portion of the aspirate can be shipped, or can sit for at least hours, without sacrificing its mscs. rbclayer mscs are nearly identical to mscs obtained conventionally. perhaps most importantly, rbc-layer mscs retain a stem-cell like capacity, showing them to be a highly valuable cell population in aml research. future plans include investigating potential selective enrichment of stem-cell mscs in the rbc-layer, which could explain the unexpected difference in growth kinetics. aml researchers now have the opportunity to study this exciting component of the bone marrow microenvironment without sacrificing valuable leukemic cells in the process. background: neutropenia is one of the most frequent side effect of chemotherapy associated with an increase in the risk of infection, especially in the cases when the depth and duration of neutropenia are extended. some genes, as variations of darc, gsdma and cxcl are known to influence white blood cell and neutrophil counts. our previous study conducted in children with acute lymphoblastic leukemia (all), showed that polymorphisms in these genes might play a role in the onset of chemotherapy complications during consolidation and maintenance treatment. objectives: in order to support our previous finding, we have expanded the study to the induction period in a cohort of all children treated at the sainte-justine university health center between july and july . design/method: previous associated single nucleotide polymorphisms (snps) in darc, gsdma and cxcl genes were analyzed for an association with the complications occurring during induction including the duration of low neutrophil count (pnn) and low absolute phagocyte count (apc), proven infections and delay between induction and consolidation phases. results: significant effect was found for all studied polymorphims. minor alleles of darc rs , cxcl rs and gsdma rs were all associated with higher risk of complications during induction treatment, whereas that of darc rs (particularly gg genotype) had a protective effect. the gg genotype of rs was associated with a lower risk of post-induction delay (p = . or = . , %ci . - . ), less frequent febrile episodes (p = . ) and lower number of days with apc/pnn count reduction (p = . for apc< . and p = . for pnn< . ). in contrast, the minor t allele of another darc polymorphism (rs ), was associated with longer apc/pnn count reduction (p = . for apc< . and p = . for pnn < . ), as it was the tt genotype of gsdma rs (p = . for apc< . and p = . for pnn< . ). the patients with the gsdma rs had also a higher risk of documented febrile episodes (p = . or = . %ci - . ). the aa genotype of rs cxcl was associated with a higher risk of post-induction delay due to infection (p = . , or = . , % ci . - . ). conclusion: this complementary study confirmed our previous results, showing overall that variations in darc, gsdma and cxcl genes influence the onset of chemotherapy complications in pediatric all, regardless of treatment phases. these polymorphisms might be useful pharmacogenetics markers possibly guiding an adjustment of chemotherapy intensity. background: pediatric acute myeloid leukemia (aml) has a poor survival rate of about % and there is an urgent need for newer targeted therapies. car t-cell based therapies are effective against all but similar therapies against aml are still under development. recent clinical trials have highlighted the concerns about toxicity and therapy related deaths from car t-cells. antigen selection is the key factor determining the specificity, efficacy and toxicity of car t-cells. while contemporary adoptive t-cell therapies use monoclonal antibodies against tumor associated antigens we employed the naturally occurring flt ligand (fl) to target aml cells expressing flt receptors. flt receptor is expressed on multipotent and myelomonocytic progenitors as well as myeloid leukemia cells. to generate fl containing chimeric tlymphocytes designated flcar t-cells and to evaluate their efficacy against aml cells. design/method: flcar was constructed by fusing the coding sequences of the human fl, cd costimulatory domain, and cd -zeta chain (intracellular region) in series. it was then cloned into the phiv-egfp lentiviral vector for expression in cell lines and primary t cells obtained from healthy donors. the empty phiv-egfp vector was used as a negative control. flcar was expressed on both cd + and cd + t-lymphocytes, confirmed by western blot. cell cytoxicity was evaluated by co-culturing flcar t-cells and aml cells followed by flow cytometric analyses. cytokine production was assessed by analyzing expression of interleukin- using quantitative rt-pcr. results: flcar t-cells were generated from cd + jurkat and cd + tk- cell lines with up to % lentiviral transduction efficiency. the efficiency for primary t cells was lower ( - %). flcar was expressed as a ∼ kda protein in cells and was partially phosphorylated on tyrosine. the expression of flcar on lymphocytes lead to increased basal il- expression in the cells. this was further augmented (by > folds) upon co-incubating flcar t-cells with flt expressing target cells. jurkat cells, tk- cells and primary human t cells expressing flcar suppressed the growth of flt -expressing aml cell lines and primary aml cells in vitro. notably, flcar t-cells generated from healthy donors caused strong inhibition of aml cells even at a lower transduction efficiency. in vivo experiments using nsg-sgm mice xenografted with human aml cells are underway. our data demonstrate that flcar can be effectively expressed on t-lymphocytes and mediate potent cytotoxicity against flt -expressing aml cells in vitro. being a completely human derived chimeric protein, it represents a promising candidate for further therapeutic development. holly pacenta, kelly sullivan, ahwan pandey, kelly maloney, joaquin espinosa children's hospital colorado, denver, colorado, united states background: individuals with down syndrome (ds) have a -fold higher risk of developing acute lymphoblastic leukemia (all) than the typical population. there are several important differences between all in individuals with ds (ds-all) and all in individuals without ds (nds-all): first, patients with ds-all have a lower percentage of favorable cytogenetic features compared to nds-all. second, patients with ds-all are more likely to have activating mutations in jak , crlf overexpression, and ikzf deletions. despite these clear genotypic differences, this knowledge has not yet been exploited for therapeutic purposes in ds-all. when outcomes for ds-all are compared to nds-all with similar cytogenetic features, the survival rates are similar. however, individuals with ds-all have an increased risk of treatment-related mortality (trm). current therapy for ds-all is similar to that for nds-all, with the exception of small changes to decrease toxicities that are more prevalent in ds-all. it was recently identified that interferon signaling is constitutively activated in healthy individuals with t . we hypothesize that aberrant interferon signaling could play a role in the unique leukemias observed in ds patients. objectives: to identify differences in gene expression and intracellular signaling cascades that are unique to individuals with ds-all, relative to both nds-all and healthy individuals with ds that can be exploited for therapeutic use. design/method: bone marrow samples were obtained from ds-all patients and matched nds-all patients based on clinical characteristics and genetic features. rna sequencing of these samples was performed and a total of samples were used for the transcriptome analysis ( ds-all vs. nds-all). the differential expression data was generated by deseq and analyzed using ingenuity pathway analysis. the analysis revealed that the chromosome genes that have been implicated in leukemogenesis are not differentially expressed in the ds-all samples, relative to nds-all. an inflammatory signature was identified, which included interferon gamma as an upstream regulator with predicted activation in ds-all. this finding is consistent with prior observations from healthy individuals with ds. other examples of results with potentially actionable targets include the upregulation of several genes in the ras pathway and genes involved in histone methylation. the increased interferon signaling seen in healthy individuals with ds was also identified in ds-all. this may contribute to the development of mutations in inflammatory pathways such as jak and crlf in ds-all. targeting these common pathways with small molecule inhibitors may have a therapeutic benefit in ds-all. cincinnati children's hospital medical center, cincinnati, ohio, united states background: next-generation sequencing (ngs) guides precision medicine approaches in oncology using therapies targeting molecular alterations found within an individual cancer. increased availability of ngs coupled with a proliferation of targeted drugs in development heightens the need for reliable pre-clinical animal models. here we report a patientderived xenograft (pdx) system with integrated molecular profiling for pre-clinical testing of conventional cytotoxic and novel targeted agents. objectives: to utilize ngs from patients with pediatric leukemia to guide rational pre-clinical trials in pdx leukemia avatars, and to determine pdx mice tolerance of and response to cytotoxic and targeted therapies. pediatric acute lymphoblastic leukemia (all) samples were obtained in adherence to an irb-approved protocol and xenografted into nod/rag/interleukin- (il- )rg (nrg) mice. ngs was performed clinically using the foundationone® heme panel. a de novo all sample bearing mutations involving jak , crlf , ntrk , cdkn a/b, ptpn and wt was used for pre-clinical testing. thirty-seven nrg mice were transplanted with million patient cells/mouse via iv injection. standard -drug induction chemotherapy was administered consisting of vincristine, dexamethasone, pegaspargase, and daunorubicin [vxpd, n = mice], in comparison to vehicle control [n = ]. parallel pdx cohorts were treated with single agent targeted therapies based on ngs findings, including ruxolitinib [n = ], crizotinib [n = ] and loxo- [n = ]. the four-week treatment period began on day + from transplant after confirmation of engraftment. following completion of therapy, residual disease burden was analyzed by flow cytometry (hcd +, mcd -cells) in the bone marrow [bm] . to date, pdx models have been established using over thirty ngs-profiled pediatric all samples, including six samples bearing philadelphia (ph) chromosome or phlike mutations. pre-clinical testing was performed in a repre- conclusion: ngs reveals concomitant mutations in ph-like all that may represent additional targets for therapy, or predict tyrosine kinase inhibitor (tki) resistance. we show that all xenograft nrg mice can tolerate a -week multi-agent cytotoxic chemotherapy induction regimen, as well as rational targeted agents, and serve as a robust pre-clinical model for precision medicine trials. background: osteonecrosis is a well-characterized all therapeutic toxicity attributed to glucocorticoids, asparaginase, and methotrexate that disproportionately affects adolescents. in ccg- , alternate-week dexamethasone during double delayed intensification (di) reduced osteonecrosis vs continuous dexamethasone with single di in rapid early responders (rer) ≥ y. to compare efs and os between hr-all patients with vs without osteonecrosis. design/method: hr-all patients - y on aall ( - ) received cog augmented therapy with a × randomization to: ( ) induction dexamethasone ( mg/m d - ) vs prednisone ( mg/m d - ), and ( ) interim maintenance (im) high-dose methotrexate (hdm) vs escalating-dose methotrexate/pegaspargase (ema). rer received single, and slow early responders (ser) double, im/di. initially, all received monthly dexamethasone maintenance pulses, patients ≥ y received di alternate-week dexamethasone, and patients ≤ y received di continuous s of s dexamethasone. there were osteonecrosis-related amendments: after / all patients ≥ y received di alternateweek dexamethasone; after / all patients ≥ y were assigned to induction prednisone, and all patients received di alternate-week dexamethasone and maintenance prednisone pulses. results: osteonecrosis was confirmed in / patients. the y cumulative incidence (ci) was . % overall and increased with age: - y . %, - y . % (alternateweek dexamethasone . % vs continuous dexamethasone . %; p< . ), ≥ y . % (p< . ). among randomized rer patients ≥ y, ci differed by glucocorticoid (dexamethasone . % vs prednisone . %; p = . ) but not methotrexate assignment (hdm . % vs ema . %; p = . ). among randomized ser patients ≥ y, ci was . % with no difference by regimen. results were similar for patients ≥ y. in the entire study population, patients with osteonecrosis had superior y efs ( . % vs . %; p< . ) and os ( . % vs . %; p< . ) than those without osteonecrosis. y efs was significantly higher among randomized patients ≥ y with vs without osteonecrosis ( . % vs . %; p< . ); this finding was present in different age ranges (≥ y, ≥ y, ≥ y) and rer/ser subsets within each, especially in the ≥ y rer ( . % vs . %; p = . ) and ser ( . % vs . %; p< . ) cohorts. across groups, asparaginase allergy was significantly associated with reduced osteonecrosis risk (≥ y: hr . ; p = . ). patients who develop osteonecrosis have significantly increased efs and os, suggesting host differences that increase sensitivity to develop osteonecrosis and render all cells more chemo-responsive. pennsylvania state university, hershey, pennsylvania, united states background: cdc (cell division cycle protein ) belongs to rho family of small gtpases in ras-oncogene superfamily. pro-oncogenic role of overexpressed cdc in ras driven solid tumors are well known. however, role of cdc in leukemia is yet to be established. ikzf encodes ikaros protein which has important role in regulation of lymphoid development and tumor suppression in leukemia. casein kinase ii (ck ) oncogene is overexpressed in leukemia. ck impairs ikaros function which can be restored by using ck inhibitors. objectives: to investigate role of cdc in leukemia and regulation of cdc by ikaros and ck in b-cell acute lymphoblastic leukemia (b-all). shrna transfection, real time-pcr, luciferace assay, quantitative chromatin immunoprecipitation (qchip) coupled with the next-generation sequencing (chip-seq), cytotoxicity assay and western blot. results: cdc is identified as one of the ikaros target genes by analysis of genome-wide dna binding of ikaros using chip-seq and qchip in b-all primary cells. expression of cdc was also noted to be higher in all patient samples compared to normal bone marrow. functional experiments showed that ikaros overexpression via retroviral transduction results in transcriptional repression of cdc . ikaros silencing using shrna resulted in increased expression of cdc . these data suggest that ikaros negatively regulates transcription and expression of cdc . ck directly phosphorylates ikaros and impairs its function as transcription factor. we noted that molecular inhibition of ck via sirna as well as treatment with specific ck inhibitor, cx also decreases expression of cdc . treatment with cx of primary b-all with ikaros haploinsufficiency restores ikaros binding to cdc promoter and represses cdc expression. however, this effect is evident only in presence of ikaros. treatment with cx in ikaros silenced (ikaros shrna) cells showed no change in expression of cdc . these results emphasizes the importance of ikaros in regulating cdc expression. furthermore, we analyzed the changes in epigenetic signature at the cdc promoter following treatment with cx . results show that loss of histone marker of open chromatin (h k ac) and increased histone marker for repressive chromatin (h k me ), at the cdc promoter. these data suggest that ikaros transcriptionally represses cdc via chromatin remodeling. a specific cdc inhibitor, ml showed cytotoxic effects on primary b-all cells. conclusion: cdc may have important role in hematologic malignancies. expression of cdc in b cell all is regulated by ikaros and ck . these results suggest that targeting cdc could be a potential therapeutic strategy in leukemia. caitlyn duffy, laura hall, justin godown, koyama tatsuki, scott borinstein monroe carell jr. children's hospital at vanderbilt, nashville, tennessee, united states background: systemic corticosteroids are widely used as treatment of acute lymphoblastic leukemia (all) and lymphoblastic lymphoma. there are anecdotal reports of bradycardia in pediatric patients receiving corticosteroids, but a more extensive analysis of this effect is needed. objectives: the aim of this study was to describe the incidence, severity, and timing of steroid-induced bradycardia and document any adverse events associated with bradycardia. design/method: we performed a retrospective review of all newly diagnosed patients at our center ( - ) with all/lymphoblastic lymphoma who received corticosteroids (dexamethasone - mg/m /dose or prednisone mg/m /dose) during induction chemotherapy. patients were excluded if they had a pre-existing cardiac abnormality or if they received prior corticosteroids. the average hour heart rate (hr) was assessed for the period prior to initiating steroid therapy and for the hour period surrounding the nadir following steroid administration. the degree and time of steroid induced bradycardia was assessed. adverse patient events and concomitant medication use was documented to identify other contributing factors to bradycardia. a total of children ( females, males, months- years) were included in the analysis with demonstrating a decrease in mean hr following steroid administration. median hr decrease was . beats per minute (quartiles . - ) from prior to initiating steroids to surrounding nadir. sixty one percent developed bradycardia less than or equal to the st percentile for their age range. nadir occurred doses (range - ) into treatment, which corresponded to hours ( - ) after initiation of therapy. of patients who experienced bradycardia, % were associated with dexamethasone rather than prednisone. hr nadir was not associated with other vital sign abnormalities. after completion of induction chemotherapy, % of patients had documented resolution of bradycardia with hr greater than the th percentile for age. it was observed that the children who continued to have relatively low hr were often younger ( months- years old). examination of nadir hr during subsequent hospitalizations in which steroids were not being administered (excluding hr during procedural sedation) did not demonstrate a significant incidence of bradycardia. concomitant opioid, beta-blocker, or other medication exposure did not contribute to the incidence of bradycardia. corticosteroid-induced bradycardia is extremely common in children, teenagers, and young adults with all receiving induction chemotherapy. bradycardia was not associated with clinical adverse events and resolved after completion of corticosteroid treatment. therefore, further cardiac assessment may not be warranted in the presence of bradycardia suspected to be secondary to steroid administration. baylor college of medicine, houston, texas, united states background: survival in newly diagnosed pediatric acute myeloid leukemia (aml) is approximately %; however survival falls dramatically if a patient relapses. currently, approximately one-third of patients with pediatric aml relapse on standard chemotherapy regimens. aml cells are exposed to proteotoxic stress at baseline due to their rapid and inefficient metabolism; proteotoxic stress increases after chemotherapy due to accumulation of reactive oxygen species resulting in misfolded proteins. this leads to activation of cell stress pathways, such as the unfolded protein response (upr) in the endoplasmic reticulum. because an activated upr can make cells more sensitive to proteotoxic stress, we hypothesize that upr activation correlates with response to chemotherapy. objectives: determine the status of upr in pediatric aml and its correlation with chemosensitivity; design/method: peripheral blood samples from pediatric patients with aml were collected at the start of induction chemotherapy, - hours (h) and h post initiation of systemic chemotherapy. tumor cells were sorted from peripheral blood mononuclear cells. expression of upr proteins was determined by chemiluminescence using an automated capillary electrophoresis system. clinical correlations were performed using an annotated database. we measured five upr proteins: grp (glucose regulated protein kda), phospho-eif , inositol-requiring enzyme (ire ) and activating transcription factor (atf ). patients with aml had - times higher expression of upr proteins (except atf ) at baseline than normal controls. grp -the key upr driver-had the highest level of protein expression in myeloid blasts. there was a wide variability in the level of baseline upr expression. eight out of samples expressed > fold increase in grp above those with the lowest grp levels. similarly, and patients respectively, had a > fold increase in peif and ire , compared to patients with low basal expression of these upr proteins. in our limited sample set, there was a trend towards lower overall survival (os) and event-free survival in patients with low baseline grp and ire . conclusion: upr has a variable expression at baseline in pediatric aml, with a trend towards lower os in patients with a low basal grp and low ire expression, suggesting less chemosensitivity in this subgroup. conversely, it is possible that blasts with an upregulated upr prior to chemotherapy manage proteotoxic stress less effectively, having faster apoptosis and hence a better response to chemotherapy in patients with a high basal upr. we are currently expanding our findings in a larger cohort of patients enrolled in the children's oncology group aaml protocol. background: children with newly diagnosed acute lymphoblastic leukemia (all) undergo chest x-ray (cxr) evaluation during initial diagnostic workup to ensure safe airway management. however, to our knowledge, no systematic assessment of cxr findings has been reported. objectives: to evaluate cxr findings at diagnosis of all and their associations with clinical characteristics. we reviewed the cxr findings at diagnosis of all in patients treated on the total xv and xvi protocols at st. jude children's research hospital. findings were evaluated for associations with clinical characteristics at presentation, and the clinical management of mediastinal masses was reviewed. mediastinal masses were seen in ( . %) of patients evaluated and were more common in older patients (mean age, . years) than in younger patients (mean age, . years) (p = . ), in males than in females (p = . ), and in patients with t-all than in those with b-all (p< . ). also associated with mediastinal masses were a higher white blood cell count (wbc) at diagnosis (mean, . × /l) (vs. a lower wbc; mean, . × /l) (p< . ), cns involvement (vs. no involvement) (p = . ), and standard/high-risk disease (vs. low-risk disease) (p< . ). other cxr findings included pulmonary opacity ( patients [ . %]), bronchial/perihilar thickening ( patients [ . %]), cardiomegaly ( patients [ . %]), and osteopenia/fracture/periosteal lesions ( patients [ . %]). pulmonary opacity was more common in younger patients (mean age, . years) than in older patients (mean age, . years) (p = . ) and in those with t-all (vs. b-all) (p = . ). bronchial/perihilar thickening, cardiomegaly, and osteopenia/fracture/periosteal lesions were also more common in younger patients than in older ones (p< . , p = . , and p< . , respectively) and in those with low-risk disease (versus standard/high-risk disease) (p< . , p = . , and p = . , respectively). of the patients with a mediastinal mass on cxr, underwent a confirmatory chest ct scan, and ( . %) were confirmed to have a mediastinal mass. notably, patients ( . %) had airway compression, and compression of venous structures was identified in of patients ( . %) who received iv contrast. the clinical course was evaluated for patients with mediastinal masses detected by cxr. fifty patients ( . %) required icu admission (mean stay, . days). general anesthesia was used for only patients ( . %), and patients ( . %) had a less invasive peripherally inserted central catheter. no deaths occurred in the acute phase. conclusion: cxr at the time of all diagnosis can detect various intrathoracic lesions and is helpful in planning initial diagnostic workup and management. background: mertk is a receptor tyrosine kinase that is aberrantly expressed in % of pediatric primary aml samples. mertk inhibition with the small molecule tyrosine kinase inhibitor (tki) mrx- decreases tumor burden and prolongs survival in aml xenografts. while treatment with mrx- reduces leukemia in the peripheral blood, it is less effective in the bone marrow, suggesting a role for the marrow microenvironment in therapeutic resistance. the jak/stat pathway has been implicated as a mediator of bone marrow derived resistance to tkis and inhibitors of this pathway are in clinical development for the treatment of aml. to determine the role of the bone marrow stromal niche in mediating resistance to mertk inhibition and to evaluate the efficacy of combined mertk and jak/stat inhibition. design/method: aml cell lines were cultured with or without the hs stromal cell line or hs conditioned medium, then treated with mrx- +/-the jak/stat inhibitor ruxolitinib, or control. induction of apoptosis and cell cycle arrest in aml cells was measured by flow cytometry. expression of h ax and total and phosphorylated stat were determined by immunoblot. results: co-culture with stromal cells significantly reduced aml cell death and g /m phase arrest in response to treatment with nm mrx- compared to no co-culture (cell death: . % versus . %, p< . ; g /m arrest: . % versus . %, p< . ). g /m arrest was accompanied by an increase in h ax expression which was similarly abrogated in co-culture. conditioned medium did not provide protection from mrx- induced apoptosis, g /m arrest, or h ax induction. mrx- inhibited stat phosphorylation but direct co-culture and conditioned medium potently increased basal stat phosphorylation which was not inhibited by mrx- . to determine whether the observed induction of stat phosphorylation was functionally relevant, cocultures were treated with both mrx- and ruxolitinib. while ruxolitinib potently inhibited the phosphorylation of stat in the presence of co-culture, combination treatment did not overcome stromal mediated protection from mrx- induced apoptosis. similarly, the addition of exogenous gm-csf induced stat phosphorylation but did not yield protection from mrx- functional effects in the absence of co-culture. together these data support a model whereby direct cell-cell contact with stromal cells in the bone marrow niche protects leukemia cells from mrx- induced apoptosis, cell cycle alterations, and dna damage. while co-culture potently induces phosphorylation of stat in leukemia cells, this is neither necessary nor sufficient for stromal-cell mediated protection from mertk inhibition and combined treatment with jak/stat inhibitors is unlikely to be therapeutically efficacious. background: mercaptopurine ( -mp) is an immunosuppressive thiopurine drug that is a key component of acute lymphoblastic leukemia (all) treatment. -mp is metabolized into -thioguanine ( -tgn), which is responsible for anti-leukemic effects, as well as -methylmercaptopurine nucleotides ( -mmpn/ -mmp), which are associated with hepatotoxicity. some patients preferentially metabolize -mp to -mmpn/ -mmp, increasing their risk for hepatotoxicity and potentially reducing anti-leukemic effects. hepatotoxicity can cause interruptions or delays in therapy that may jeopardize cure rates. allopurinol has been increasingly used in patients with inflammatory bowel disease (ibd) to shunt -mp metabolism toward -tgn and away from -mmpn to minimize hepatotoxicity and preserve therapeutic effects. objectives: this retrospective chart review expands upon our previously published case series of three patients with all in whom allopurinol was successfully used to redirect -mp metabolism. twelve additional patients have subsequently received allopurinol and -mp combination therapy at texas children's hospital. data from this larger patient sample, with longer follow up, is being analyzed to increase knowledge of the effectiveness and longitudinal effects of adding allopurinol to -mp to reduce risk of hepatotoxicity. design/method: data were abstracted from the electronic medical records of patients with all treated at texas children's hospital from to present, who had been found to have evidence of altered -mp metabolism and in whom allopurinol was added to -mp therapy due to concern for risk or recurrence of hepatotoxicity. metabolite levels, -mp dose, and alanine transaminase (alt) prior to initiation of allopurinol and approximately weeks later were compared. wilcoxon signed-rank test was applied for statistical analysis. : after the addition of allopurinol, patients experienced a significant decrease in mean levels of -mmpn (p = . ), correlating with a significant decrease in mean alt (p = . ). with the initiation of allopurinol, the mean -mp dose was decreased from to mg/m /day over an -week period. mean -tgn levels increased (p = . ). in follow up beyond weeks, no patients had further holds in -mp due to hepatotoxicity. addition of allopurinol appears to shift metabolism from -mmpn toward -tgn, with increases in mean -tgn levels despite a decrease in mean -mp dose. this may limit negative side effects, thus resulting in fewer gaps in therapy and possible improved outcomes. further analysis of -mp dose titration and effects on anc over time as well as effects on overall survival is ongoing. prospective background: alterations in epigenetic patterning are a fundamental feature in acute myeloid leukemia (aml). treatment with dna methyltransferase inhibitors (dnmti) yields responses in aml, but the molecular mechanisms underlying this effect are poorly understood. in prior work, we demonstrated induction of genes involved in the pirna rna (piwi) silencing pathway as a common gene feature of aml cell lines treated with decitabine. the piwi pathway is an rna silencing system, distinct from classical small rna transcriptional silencing, responsible for transposon-silencing in gametogenesis; emerging data suggest a role for this system in somatic cells. based on these data, we postulate that piwi induction plays a crucial role in aml recovery following demethylation and that disruption of this pathway would modulate response and/or recovery from decitabine treatment. to assess the effect contribution of the pirna pathway response following dnmti treatment in aml. design/method: to choose target genes in the pirna pathway for disruption, molm cells were first treated with escalating doses of decitabine. using quantitative rt-pcr, the dose-dependent expression of several pirna-associated genes were analyzed. two genes, mael and piwil , were selected for disruption experiments based on preliminary data suggesting decitabine dose-dependent responses. molm cells were transduced with shrna targeting these genes using a lentivirus delivery system with selection in puromycin. knockdown efficiency was assessed by rt-qpcr. to determine how gene disruption affected cell growth, knockdown cells were treated with decitabine nm. proliferation was assessed by celltiter glo assay following decitabine treatment. clonogenic potential was assessed by colony forming assays of transduced cells after treatment with decitabine at nm and nm. results: following decitabine exposure in molm , there was a markedly increased expression of mael and piwil compared to untreated cells ( : and : , respectively) . thus, these were the candidate genes chosen for disruption. of mael shrna constructs, two resulted in a % relative expression of mael compared to controls. of the piwil shrna constructs, the best knockdown showed % relative expression. there were no significant differences in proliferation or clonogenicity of stably selected mael or piwil knock-down molm cells following decitabine treatment. using gene knockdown procedures, mael and piwl do not appear to have a marked effect on growth and response to decitabine treatment in molm . however, these results may be limited by inefficient knockdown using shrna targeting methods. further work using a cas /crispr based inactivation of these genes is ongoing. children cancer hospital cairo, egypt background: hypodiploidy < chromosomes is very uncommon and have particularly poor outcomes in childhood acute lymphoblastic leukemia (all). it is subdivided into: near-haploid ( - chromosomes), lowhypodiploid ( - chromosomes) and high-hypodiploid ( - chromosomes). to determine if minimal residual disease (mrd) can identify a group of patients with better prognosis in the hypodploid population who can be treated with intensive chemotherapy alone. design/method: a retrospective study that included all patients under age of diagnosed as hypodiploid b-precursor all during the period between january -december and treated at children's cancer hospital egypt on sjcrh total study-xv for ir/hr all. sixteen patients had < chromosomes ( nearhaploid and low-hypodiploid), constituting % of all pediatric patients with b-precursor all during the study period. patients with near-haploid all had a median age of years (range - ), initial leukocyte count (wbc) median of . × /l (range . - . ), ( . %) were males and / ( . %) had hr-nci criteria. four patients ( . %) are alive in complete remission(cr) (range - months, median ), one died in induction and ( . %) had hematological relapse (range . - months, median ). patients with low-hypodiploid all had significantly older age (median years, range - ), median wbc . × /l (range . - . ), / ( . %) were males. one patient ( . %) is alive in cr, one died in induction, one failed to achieve cr post-induction and patients( %) had hematological relapse (range . - . months, median . ). mrd< . % by flow-cytometry on day- and end of induction was achieved in / ( . %) and / patients( %) with near-haploid, compared with / ( . %) and / patients( %) with low-hypodiploid; respectively (p = . , p = . ;respectively). allogeneic transplantation was performed during initial remission only in mrd negative patients (one relapsed and one is in cr) and in the patient with induction failure (relapsed post-transplant). five of the total six patients who had negative mrd on day- and end of induction are alive in cr ( / with chemotherapy alone). all patients with negative mrd at end of induction but with mrd levels≥ . % on day- (range . - . %) relapsed as well as all patients with detectable mrd at the end of induction. the difference in relapse was statistically significant in relation to negative-mrd on day- (p = . ), but not at end of induction(p = . ). conclusion: children with hypodiploid all and negative mrd on day- of induction are highly curable with intensive chemotherapy alone, while patients with negative mrd at the end of induction and detectable mrd on day- had dismal outcome. background: overall survival in pediatric acute myeloid leukemia (aml) has plateaued between - %, with death during induction chemotherapy seen in - % of patients. respiratory complications contribute to morbidity and mortality in pediatric aml induction, however the incidence, patterns, and predictors of respiratory adverse events (aes) during this period are unknown. to estimate the incidence of respiratory aes during induction therapy for de novo pediatric aml, to characterize and grade these respiratory aes, and to identify predictors of respiratory ae development. we conducted a retrospective longitudinal study from presentation to day in institutional de novo pediatric aml patients (≤ years) between march and december . outcomes included any nci ctcae grade - respiratory ae or death from another cause. demographic, disease, and treatment-related data were abstracted. the most specific, best-fitting ctcae category and grade for each ae was determined. descriptive statistics, survival analysis, multivariable logistic regression analysis, and time-toevent distributions were performed (sas v . , cary, nc) . among eligible patients, . % (n = ) experienced discrete respiratory aes. incidence of grade - aes was . % (n = ). a bimodal time-to-event distribution demonstrated peaks at treatment days and . induction death occurred in . % (n = ) including deaths from respiratory failure associated with disseminated fungal disease. in univariate analysis, those experiencing aes differed significantly in regards to older age at diagnosis (p< . ), higher initial wbc (p = . ), higher initial peripheral blast percentage (p = . ), coagulopathy at diagnosis (pt (p = . ), d-dimer (p = . )), fluid overload status (p< . ), occurrence of infection (p = . ), and occurrence of tumor lysis syndrome (tls) (p = . ). patients with hyperleukocytosis (p = . ), fluid overload (p< . ), and fab m morphology (p = . ) each had a significantly decreased probability of completing the follow up period without experiencing a respiratory ae. on multivariable analysis, fluid overload (aor . [ % ci: . - . ) and older age (aor . [ % ci: . - . ) were significantly associated with ae occurrence when gender, hyperleukocytosis, tls, and infection status were held constant. we describe a high incidence of respiratory aes during pediatric aml induction. fluid overload and older age at diagnosis are independently associated with ae development when controlling for other proposed risk factors. interventions focused on conservative fluid management and offset of fluid overload should be explored in newly diagnosed pediatric aml in an effort to reduce respiratory complications during induction. overall, all survival rates are outstanding and have continued to improve with risk-adapted therapy. the most striking improvement occurred in t-all where -year os rates now exceed % and parallel b-all. survival improvements, however, have not been observed uniformly across all subgroups. while the gap in outcome differences narrowed among blacks, outcomes for hispanics have remained static. further, no improvements in survival were observed in infants or ayas and new treatment approaches have been implemented for these populations. background: acute myeloid leukemia (aml) accounts for approximately % of new childhood leukemia cases. chest x-ray (cxr) is performed in all newly diagnosed aml cases to evaluate the safety of airway management for anesthesia during diagnostic procedures; however, cxr results in pediatric patients with aml have not been described. objectives: the primary objective was to evaluate cxr findings at diagnosis in patients with aml. the secondary objectives included assessing associations between cxr findings and clinical characteristics, with the overall goal of aiding in the evaluation of the use of cxrs as an initial diagnostic study in pediatric patients with aml. design/method: cxr findings and clinical characteristics were evaluated in patients with newly diagnosed aml who were enrolled in one of three protocols at st. jude children's research hospital (aml , aml , and aml ). the findings were categorized based on radiologic reports. further, the associations of these findings and clinical characteristics were evaluated. we evaluated cxr findings in a total of patients: from aml ; from aml ; and from aml . common cxr findings were pulmonary opacity (n = , . %), bronchial/perihilar thickening (n = , . %), splenomegaly (n = , . %), mediastinal mass and lymph nodes (n = , . %), pleural effusion/thickening (n = , . %), demineralization/fracture/periosteal lesions (n = , . %), scoliosis (n = , . %), and granulomatous disease (n = , . %). three cxr findings were associated with younger age at diagnosis: pulmonary opacity (median age, . years in patients with positive findings vs. . years in those with negative findings, p< . ), bronchial/perihilar thickening (median age, . years vs. . years, p< . ), and demineralization/fracture/periosteal lesions (median age; . years vs. . years, p = . ). two cxr findings were associated with older age at diagnosis: scoliosis (median age, . years vs. . years, p< . ) and granulomatous disease (median age, . years vs. . years, p = . ). higher white blood cell counts (wbcs) at diagnosis were associated with cxrs showing pulmonary opacity (median wbc; . × ^ /l vs. . × ^ /l, p = . ) or splenomegaly (median wbc; . × ^ /l vs. . × ^ /l, p = . ). french-american-british (fab) m /m subtypes were more frequently associated with pulmonary opacity compared with others (p< . ). we did not find significant differences between female and male patients. conclusion: cxr in patients with newly diagnosed aml showed a variety of thoracic, abdominal, and bony lesions that are important for the initial evaluation and management. pulmonary opacity was the most common finding and was frequently seen in patients who were younger or had higher wbcs at diagnosis or fab m /m . background: children diagnosed with acute lymphoblastic leukemia (all) require a central venous catheter (cvc) to administer chemotherapy safely. both external and internal cvcs carry risks of complications including thrombosis, infection, and possible replacement. internal catheters, such as a port, are generally used for the majority of patients for the duration of treatment since therapy lasts for several years. many institutions place a port at the time of diagnosis. other institutions prefer to start induction therapy via placement of a peripherally inserted central catheter (picc) and defer port placement until the completion of induction therapy due to concerns of increased risk of infectious complications with port placement. objectives: to compare rates of common cvc associated complications by type of cvc placed at start of induction therapy in children treated for newly diagnosed all at the jimmy everest center (jec) at the university of oklahoma health sciences center. design/method: a retrospective chart review analyzed data from newly diagnosed all patients treated at the jec between - . data was collected on complications including thrombosis, bacteremia, insertion site infection, cvc malfunction and need for removal. data collection began at the start of induction and was completed at the end of induction therapy. statistical analysis used a univariate and multivariate logistic regression model to compare complication rates between those who had a port versus those who had a picc placed at start of induction. results: data was collected on patients. fifty-six patients had a port placed at start of therapy while had a picc placed. fourteen percent of patients had a cvc associated complication. univariate analysis showed no statistically significant difference in rates of cvc associated complications between the groups (port %, picc . % p = . ). the rates of hospitalization for cvc associated complications were similar between both groups (port %, picc % p = . ). rates of cvc removal were also similar between both groups (port %, picc % p = . ). multivariate model that included baseline patient characteristics including type of all, patient body surface area, gender, ethnicity and age continued to demonstrate no significant difference in cvc associated complications between both groups. conclusion: this single institution study showed that there was no significant difference in cvc associated complications between port and picc line placement at the start of childhood all induction therapy. port placement can be considered as a safe option at the start of induction therapy. complete remission [cr] or cr with incomplete blood count recovery [cri]) within treatment cycles - . interim data are reported (nct ). results: seventeen patients were enrolled and received ≥ dose of lenalidomide; median age was years (range - ); patients were female. patients received median prior regimens (range - ). nine patients had previously undergone bone marrow transplantation (bmt). four patients had relapsed aml and were refractory to immediate prior treatment. median duration of study treatment was weeks (range - ); patients completed a median of treatment cycle (range - ). all patients were evaluable for primary outcome; achieved morphologic cri after cycles (no patients achieved cr). the responder was a -year-old male with history of r/r aml after first-and second-line treatment, bmt, and salvage chemotherapy. at baseline, he had a complex cytogenetic karyotype (monoallelic − q . , − q, − q . , − p ) with no identifiable molecular mutation; he was also positive for del( q) (− q , − q ). his post-treatment karyotype showed no abnormalities. sixteen patients experienced treatment failure; due to resistant disease, of indeterminate cause, and had treatment failure before a post-baseline assessment was performed. all patients experienced ≥ grade - treatment-emergent adverse event (teae). the most commonly reported were thrombocytopenia (n = ), anemia (n = ), febrile neutropenia (n = ), and hypokalemia (n = ). fifteen patients experienced ≥ teae related to lenalidomide. all patients discontinued treatment; remain in follow-up. the study is now closed to enrollment. ten patients died on study: during treatment, during follow-up. all deaths were attributed to aml or complications due to aml. conclusion: third-line lenalidomide monotherapy was associated with clinical response in of pediatric patients with r/r aml; however, treatment exposure was limited. safety data are consistent with the known profile of lenalidomide. lenalidomide was not an efficacious treatment for r/r pediatric aml. funding: celgene corporation, summit, nj, usa. cook children's medical center, fort worth, texas, united states background: it is well documented that pediatric patients with acute lymphoblastic leukemia (all) often experience significant weight gain during induction therapy and later struggle with obesity. however, some patients experience unintended weight loss during induction therapy; since this issue is not well reported, it often goes undertreated. although malnutrition is reported to be associated with decreased survival, increased risk of infection, and loss of lean body mass, there remains a scarcity of in-depth analysis of prevalence and risk factors that contribute to this problem. our study attempts to address this critical yet unmet need. objectives: our aim was to identify the clinical risk factors and outcomes associated with weight loss during induction therapy for pediatric all. design/method: this was a retrospective chart review of patients between and years of age diagnosed with all at cook children's medical center from / / to / / . for each patient, we collected height, weight, age, body mass index (bmi) z-scores at diagnosis and end of induction therapy, risk stratification, and whether consolidation was delayed. patients with a bmi > th percentile at diagnosis were categorized as being overweight or obese. using logistic regression analyses, we examined which variables predicted whether the patient had an increase or decrease in bmi z-score throughout induction. a critical alpha level of . indicated statistical significance. results: ninety-six patients met our inclusion criteria. of these, % experienced a decrease in bmi during induction therapy. compared to patients whose bmi increased during induction, patients with a decrease in bmi were more likely to be overweight or obese at diagnosis ( % vs. %; p< . ), to be ≥ years of age ( % vs. %; p< . ), to have a high-or very-high-risk stratification ( % vs. %; p< . ), and to experience a delay in the start of consolidation therapy ( % vs. %; p< . ). conclusion: this research highlights a risk not previously identified in the literature that may impact outcomes. patients treated on high-or very-high-risk protocols, who are overweight or obese at diagnosis, and who are ≥ years of age at diagnosis should be monitored closely for weight loss during induction therapy. patients who experience weight loss should receive prompt intervention. it is our hope that this information can be used for future prospective studies and to help develop evidence-based guidelines. background: p abnormalities have been observed in some patients with hematologic malignancies. loss of p function as a tumor suppressor gene in the chromosome plays an important role for development of leukemia. these patients usually have poor outcome due to the chemotherapy and are associated with poor prognosis. objectives: this study aimed to identify frequency of p abnormalities between iranian children and adult patients with aml (acute myeloid leukemia) malignancy. design/method: the p abnormalities were analyzed via bone marrow karyotyping and fish method in acute myeloid leukemia patients. in this study, p abnormalities were observed in ( %) patients out of diagnosed cases. a significant strong correlation between p abnormalities and other high risk factors (poor risk cytogenetic) were observed. from patients with aml malignancy ( p abnormalities), ( %) patients have complex karyotype, ( %) patients monosomal karyotype and ( %) patients have monosomal karyotype accompanied with a complex karyotype. overall, p abnormalities are independent risk factor in acute myeloid leukemia and evaluation of these abnormalities by fish or other complementary techniques prior to treatment, might help for better risk stratification of high risk aml patients. background: hepatotoxicity in treatment of acute lymphoblastic leukemia (all) is well studied and transiently affects most patients receiving antimetabolite therapy. rarely, patients develop liver injury severe or prolonged enough to undergo a liver biopsy. little is known about how these patients differ from patients that develop transient hepatotoxicity. we sought to describe disease and treatment characteristics for all patients that developed hepatotoxicity severe enough to undergo liver biopsy. we also looked for pre-dictive factors for liver biopsy, including signs of early hepatic injury from the initial treatment protocol. design/method: pathology reports of all patients from the liver biopsy database at children's healthcare of atlanta were collected. controls were matched : for age, all subtype, and treatment protocol. demographics, treatment protocols, and overall outcomes were collected through the electronic health record. hepatic lab results for transaminases, coagulation, and albumin were collected for induction, consolidation, interim maintenance, delayed intensification, and maintenance. results: sixteen patients diagnosed between - (median age at diagnosis years, range - ; % male; % pre-b all) were included in the case series. the median time from diagnosis to liver biopsy was . years (range - ). eight patients ( %) were in maintenance at the time of biopsy; none had active disease. eight ( %) were postbone marrow transplant. biopsy results included: steatosis ( ), acute inflammatory/infectious ( ), liver infiltration ( ), fibrosis ( ) and graft-vs-host disease (gvhd) ( ). six patients were deceased; -year all-cause mortality from diagnosis was %. thiopurine methyltransferase (tpmt) status was known in % cases and % controls. all cases had intermediate or wildtype status, which did not differ from controls (p > . ). patients requiring liver biopsy did not have evidence of acute hepatotoxicity (ast/alt > × normal values) during their initial treatment protocol. hepatotoxicity requiring liver biopsy is a rare outcome of all treatment. these patients had elevated rates of relapse, bmt, and -year all-cause mortality, suggestive of a more severe disease process. however, it is difficult to sort out the temporality of relapse, bmt, and hepatoxicity requiring biopsy in this limited sample. additionally, patients with bmt preceding liver biopsy have other confounding factors that makes them difficult to include in the analysis. finally, our limited descriptive data show no notable correlation between early hepatotoxicity and later indication for liver biopsy. future cohort or case-control studies with larger sample sizes are required to further explore early predictive factors for severe hepatotoxicity requiring liver biopsy. nathan gossai, joanna perkins, michael richards, yoav messinger, bruce bostrom background: the majority of chemotherapeutic agents used to treat hodgkin lymphoma are teratogenic. pregnancy screening prior to the start of chemotherapy is supported by clinical guidelines and baseline testing is a standard component in therapeutic trials. there is limited data available on the incidence of pregnancy screening prior to the start of hodgkin therapy but previous studies suggest that pregnancy screening, especially at pediatric institutions, is not consistently completed. objectives: the objective of this study is to evaluate the incidence of pregnancy screening and contraceptive counseling prior to the start of therapy in females diagnosed with hodgkin lymphoma. design/method: a retrospective chart review was performed for all female patients newly diagnosed with hodgkin lymphoma from to at the hospital for sick children in toronto, ontario. all patients who were intended to receive multi-agent chemotherapy were included, regardless of age. data collected included demographic and disease information, chemotherapy regimen and enrollment on clinical trial. all pregnancy testing within two weeks prior to the start of therapy was captured, as well as type of pregnancy test performed, documentation of menstrual status, contraceptive counseling and contraceptive provision. univariate and multivariate analyses were used to describe factors influencing the incidence of pregnancy testing. results: a total of female patients with newly diagnosed hodgkin lymphoma between the ages of and years were identified. sixty patients ( %) had pregnancy testing done prior to the start of therapy. testing modalities included serum and urine screens as well as quantitative beta-hcg measures. older age (p = . ), documentation of menstrual status at diagnosis (p = . ) and diagnosis between and (p = . ) were associated with higher incidence of screening. enrollment on a therapeutic trial was not associated with a higher incidence of screening (p = . ). contraceptive counseling was documented for patients ( %) and patients ( %) were prescribed contraceptive medications during therapy. pre-chemotherapy pregnancy testing was completed on % of females with newly diagnosed hodgkin lymphoma. improvement is required and interventions, including clarification of institutional standards, modification of chemotherapy order sets and staff education, are planned. (rao et al., cancer, ) . university of louisville, louisville, kentucky, united states background: granulocytic sarcomas (also known as chloromas or leukemia cutis) were first described by a. burns in . they are solid tumors comprised of immature granulocytic cells and represent extramedullary manifestations of underlying leukemia. chloromas are most commonly associated with acute myeloid leukemia. they may arise in other myeloproliferative disorders but are rarely seen in b or t cell acute lymphoblastic leukemia (all). objectives: although patients with all rarely have chloromas, it should remain on the differential for patients with unusual swelling or masses. design/method: we present a case series of two patients from our institution diagnosed with b cell all who had a chloroma as the presenting symptom. the first patient is a yo who presented to his primary provider with nasal congestion and a one-week history of bilateral eye swelling and was referred to an allergist when the symptoms did not resolve with anti-histamines. his review of systems was otherwise negative. he was referred urgently to ent two months later for a × cm mass palpated along the medial border of the left eye. an mri showed a left facial mass surrounding the zygoma and extending into the anterior inferior left orbit. biopsy revealed b cell acute lymphoblastic lymphoma, and bone marrow aspirate and biopsy confirmed the diagnosis as b cell all. the second patient is a yo who presented to his primary doctor for rapid growth of a scalp nodule that had been present for about months. he was referred to dermatology and treated for a supposed kerion from tinea capitis. the lesion continued to grow and became more irritated with this treatment. punch biopsy revealed a complicated phenotype of lymphoblastic lymphoma. however, after a lymph node biopsy and bone marrow aspirate and biopsy, the diagnosis was confirmed as b all. his only other positive point on review of systems was a questionably pathologic -pound weight loss and an area of matted cervical lymph nodes. for both of our patients, the chloromas completely disappeared during induction therapy. it is worth noting that both of these patients presented with the chloroma as the only symptom of the underlying leukemia. this led to initial misdiagnosis and delay in identifying their leukemia. therefore, while it is very rare for a patient with b all to present with a chloroma, our experience shows that all should be on the differential for patients presenting with unusual swelling or masses. background: hodgkin lymphoma (hl) is a lymphoproliferative neoplasm that commonly presents with history of adenopathy and a predictable pattern of disease involvement with or without systemic symptoms of fever and/or weight loss. in the hands of an experienced oncologist the diagnosis of hl is usually not a challenge. occasionally a diagnostic challenge is presented by a patient who has an atypical presentation which is suggestive of an alternative diagnosis. we describe a case series of patients diagnosed with hl whose initial clinical presentations lead to a diagnosis different form hl. honduras, nicaragua and the united states. results: six pediatric oncology centers from the american continent conducted a retrospective review of patients diagnosed with hl since . patients that had an initial presentation not suggestive of hl or who were initially diagnosed with a disease other that hl were included for a total of patients. argentina n = , guatemala n = , honduras n = , nicaragua n = , united states n = . five patients were female and male. patient's ages ranged from to years. most patients (n = ) were older than years. three patients ( %) presented with non-immune cytopenias without overt lymphadenopathy, of those one had active hemophagocytic syndrome. five patients ( %) were suspected to have localized solid tumors: ewing sarcoma n = , rhabdomyosarcoma n = , hepatocellular carcinoma n = , and soft tissue tumor of the cheek n = . two ( %) metastatic solid malignancy as they presented with disseminated pulmonary nodules. five ( %) with autoimmune disorders: hashimoto thyroiditis n = , autoimmune hemolytic anemia n = , nephrotic syndrome n = . ten ( %) with chronic infectious processes: brucella n = , tonsillar abscess n = , splenic abscess n = , and tuberculosis (tb) n = . patients with suspected tuberculosis were diagnosed outside of the united states. six of patients were ultimately diagnosed as having both tb and hl. seventeen patients had ann-arbor stage iii or iv, seven patient had stage ii with either b symptoms or bulky disease. patients were treated with various chemotherapy regimens according to the treating center: abvd, abve-pc oepa-copdac, avpc, beacopp. two patients had recurrent disease, one died of disease progression and one died from causes not related to hl conclusion: a small proportion of hl patients have atypical or unusual presentations. hl should be included in the differential diagnosis of solid tumors, autoimmune disorders, infections or cytopenias. the most common atypical presentation is an infectious process. background: acute lymphoblastic leukemia (all) represents the largest group of pediatric malignancies. the high cure rate of childhood all represents one of the most remarkable success stories in the war on cancer. in a lower middle income country (lmic) like the philippines, we reviewed the five year survival in a tertiary referral center. objectives: this retrospective cohort study aims to determine the survival of children - years old with all treated at a tertiary referral center for childhood cancer in the philippines from january to december . design/method: this is a retrospective cohort study that reviewed medical charts of newly diagnosed all ages to years old from january to december . a total of subjects were included in the study. the year overall survival (os) and event free survival (efs) were . % and . %, respectively. the year os for standard risk all was . % and for high risk patients was %. the year os for the patients on remission was . % and for those who relapsed was . %. univariate and multivariate by cox proportional hazards regression revealed wbc count at diagnosis, risk classification, immunophenotyping, and development of relapse showed significant prognostic impact for mortality. age and gender were reported with no prognostic significance. the -year os and efs were lower compared to developed countries but are comparable with other lmics. the prognostic factors for relapse and mortality were compatible with the literature. overall, the adopted treatment protocols for childhood all in this institution showed acceptable results. relapse has a significant prognostic impact for mortality. development of accessibility to care, increase awareness, early detection and resources at hand should be achieved. improvement in the follow up protocol to prevent delays in the treatment, patient education to prevent non-compliance and psychosocial support, to developed better supportive care, and expand facilities should be given emphasis to further improve survival and prevent relapse. objectives: here, we seek to further characterize this entity by describing the pathologic and clinical features of pediatric cases of burkitt-like lymphoma with q aberration. we collected pathologic and clinical data from the medical record on all pediatric high grade b-cell lymphoma (hgbcl) cases diagnosed at our institution over a -year period ( - ) . for those cases classified as neither burkitt lymphoma nor diffuse large b-cell lymphoma (dlbcl), fish for myc, bcl- and bcl- , as well as array comparative genomic hybridization (acgh), were performed. we identified cases of hgbcl, including cases of burkitt lymphoma presenting as purely leukemic phase. of the hgbcl cases, had burkitt lymphoma as defined by myc rearrangements, and had dlbcl. collectively, the majority of these patients had primary disease outside of the head/neck, and most patients presented with advanced stage (iii-iv) disease. of the remaining cases, q aberration was identified in cases using acgh. all cases histologically and immunophenotypically resembled burkitt lymphoma but lacked myc rearrangement, instead showing proximal gains in q -q and telomeric losses in q . qter. all cases involved primary disease in the cervical lymph node and/or tonsil. three of these cases were localized (stage ii), and the fourth case involved a few metabolically active but non-enlarged lymph nodes in the chest and abdomen (stage iii). all patients achieved complete remission with standard therapy for mature b-cell lymphoma, and were alive with no clinical evidence of disease at a median follow-up of months. although the number is small, our results suggest that the majority of non-burkitt, non-dlbcl cases of pediatric hgbcl carry q aberrations. in addition, patients with q aberrations appear to be more likely to present with lower stage disease, thus requiring less intensive therapy, and also tend to have primary disease in the head/neck. these findings further support the classification of burkitt-like lymphoma with q aberration as a distinct pathologic and clinical entity, and we propose that all pediatric non-burkitt, non-dlbcl cases of hgbcl regularly undergo further workup for possible q aberrations. marie claire milady auguste, joseph bernard st damien hospital, port-au-prince, port-au-prince, haiti background: hodgkin lymphoma (hl) and non-hodgkin lymphoma (nhl) account for % of cancers in the united states pediatric population ( , ). in central america and the caribbean, they are in second position among all types of pediatric cancers ( ). a previous study on pediatric cancers in haiti showed that the lymphomas were in fifth place after the leukemias, wilms tumor, retinoblastoma and the sarcomas ( ). the main objective of this study is to present the epidemiological profile of lymphomas managed at a haitian pediatric hospital. design/method: this is a retrospective study conducted on the cases of lymphoma diagnosed and managed at st damien hospital from january to december . key variables such as age, gender, stage at diagnosis, histopathological types and outcome were collected to present the characteristics of this retrospective cohort. of the cases of cancer diagnosed during the study period, ( . %) had the diagnosis of lymphoma. the sex ratio was . ( males for females) and the average age was . years [ - years]. there were cases of hl ( . %) and cases of nhl ( . %). . % of the patients were diagnosed at stages iii and iv. among the hl cases, ( . %) were nodular sclerosis lymphoma, ( . %) with mixed cellularity and ( . %) with lymphocytic predominance. for the nhl cases, ( . %) were burkitt's lymphoma and ( . %) lymphoblastic t-cell lymphoma. among the patients for who immunohistochemistry was found, the cases of hl were cd -positive and out of cases of nhl were cd -negative. only patient was hiv-positive, and patients had a confirmed exposure to epstein-barr virus. patients ( . %) were lost to follow-up, ( . %) were in remission, ( %) relapsed, ( . %) were still in treatment and ( %) were deceased. university of chicago, chicago, illinois, united states background: due to the adoption of risk-adapted therapy, pediatric and adolescent acute lymphoblastic leukemia (all) is associated with high cure rates. despite excellent outcomes in most children, patients with certain blast cytogenetic features do not fare as well. furthermore, african american, native american, and hispanic patients have worse outcomes than caucasian patients. while the outcome discrepancies are certainly multifactorial, and blast cytogenetics are related to age, it remains unclear whether ethnicity and blast cytogenetics correlate. the diverse patient population at the university of chicago provides an opportunity to evaluate for such a correlation. objectives: to describe cytogenetic findings in a racially and ethnically diverse population of patients of all age groups diagnosed with all at university of chicago from to and determine if there is a correlation between race/ethnicity and blast cytogenetics. results: a total of newly diagnosed patients with all between the ages of - from - were included in this study. of those, patients ( . %) had b-all, had t-all ( . %), one had early t-cell precursor all and one had mixed phenotype all (b/t). caucasians accounted for % of patients, african americans (aa) %, hispanics . %, asians . %, and % were of other races. age distribution had a bimodal pattern, with a peak in incidence at and another at years of age, consistent with published data. cytogenetic categories included: t( ; )(p ;q ), q rearrangements (kmt a), iamp , t( ; )(q ;p . ), t( ; ) (q ;q ), hypodiploidy, hyperdiploidy and double trisomy of chromosomes and . aa and hispanic patients with b-all presented more frequently between the ages of - years compared to caucasians (p = . and . , respectively). in aa patients, t( ; ) (q ;p . ) was overrepresented (p = . when compared to caucasians), and was mainly observed in patients between - years. caucasian patients were more likely than non-caucasians to have hyperdiploidy (p = . ), especially in patients aged - years. the rate of t( ; )(q ;p . ) was significantly higher in aa patients in our cohort, in particular in patients between the ages of - years. hyperdiploidy was more likely in caucasians aged - years. these findings may suggest that varying blast cytogenetics could contribute to outcome differences between races. ahmed elgammal, yasser elborai, mohamed fawzy, asmaa salama, eman d el-desouky, lobna shalaby national cancer institute, cairo, cairo, egypt background: hodgkin lymphoma (hl) in children is one of the malignancies that have a high chance of cure. stage iv hl remains a challenge for getting good clinical outcome as in other stages. many treatment protocols used to give combination chemotherapy while combined modality treatment is the mainstay in other treatment protocols. objectives: we aimed in to assess the outcome using consolidation radiotherapy to chemotherapy (combined modality treatment) versus combination chemotherapy alone in treatment of stage iv hl. design/method: we included patients with stage iv hl and whose data were retrieved from the medical records of the pediatric oncology department, national cancer institute, cairo university, egypt from till june and were followed till august . treatment was either to give cycles of abvd (adriamycin, bleomycin, vinblastine, dacarbazine) only or to give cycles of abvd followed by consolidation radiotherapy. the study included cases; were males and were females. mean age was . years ranging from to years. the histopathology subtype was nodular sclerosis in the majority of cases ( cases) followed by mixed cellularity ( cases) then only one case of lymphocyte rich. nine cases were initially bulky while cases were not. constitutional manifestations were present in cases while it was absent in cases. bone marrow was involved in only cases. radiotherapy was given after completion of chemotherapy to cases while cases received chemotherapy only. the -year overall survival for patients who received radiotherapy was superior to those who received chemotherapy alone; % versus . % respectively with statistical significance (p = . ). the -year progression free survival was also higher with radiotherapy than others; % versus . % (p = . ). patients with stage iv hl who received consolidation radiotherapy apparently had a better outcome than those who received chemotherapy only. this suggests that radiotherapy contributes significantly with chemotherapy to the cure rate for those patients. the feinstein institute for medical research, manhasset, new york, united states background: microrna (mirnas) are short non-coding rnas that play a decisive role in cancer biology, including leukemia. exosomes are microvesicles ( - nm) produced by most cells in biological fluids. exosomes represent the fingerprint of the parental tumor and are loaded with bioactive markers such as mirnas, which may regulate tumor growth. exosomal cargo can be transferred into target cells changing their biological properties. our study investigates a functional role for exosomal mir- a in pediatric acute lymphoid leukemia (p-all). objectives: / to demonstrate that p-all exosomes induce cell proliferation / to confirm that exosome-induced cell proliferation is disease-stage specific / to analyze exosomal mir- a expression profiles in p-all / to authenticate that inhibition of exosomal mir- a reduces leukemia proliferation design/method: exosomes were isolated by ultracentrifugation from healthy donors (hd) & p-all serum and conditioned medium (cm) of sup-b , jm , and cl- (control) human cell lines. cell lines were exposed to different sources of leukemia-derived exosomes in a paracrine or autocrine fashion for hrs in triplicates. proliferation was assessed by microscopic cell counting and confirmed by gene expression for proliferation, pro-survival and pro-apoptotic genes. mirna profiling was performed with the human cancer pathway finder microarray (qiagen). silencing of exosomal mir a was carried out by a mir- a inhibitor (qiagen), utilizing exo-fecttm exosome transfection reagent (sbi, system biosciences). further, exosomal mir- a silencing was confirmed by q-pcr. cellular uptake of texred-sirna (sbi, system biosciences) was confirmed by flow cytometry. transfer of exosomal mir a to the target cells was evaluated by q-pcr. we elucidated that cm-derived exosomes from sup-b and jm cell lines induce cell proliferation in sup-b , jm (autocrine and paracrine) and cl- cells (paracrine) (p< . ). serum p-all exosomes promote paracrine cell proliferation in all cell lines compared to hdderived exosomes (p< . ). heatmap analysis of mirna profiles of leukemia exosomes (all cell lines and p-all) identified mir- a significantly upregulated in leukemia exosomes compared to controls. mir- a was also upregulated in all cell lines after exposure to leukemia exosomes that induced proliferation. moreover, exosomal mir- a inhibition reduces leukemic proliferation in pediatric all. our data suggest that all exosomes induce cell proliferation of leukemic cell lines in both paracrine and autocrine fashion. exosomes regulate these phenomena in a highly orchestrated way, by transfer of functional exosomal mirnas such as mir- a. the results of this study suggest s of s that exosomal mir- a inhibition can act as a novel way for growth-suppression of pediatric leukemia. results: a total of disease sites were detected at pet/ct, while sites were detected at contrast-enhanced ct and bone marrow biopsy (bmb). pet/ct showed improved detection of nodal lesions (p < . ) (kappa value = . ), extranodal lesions (p < . ) (kappa value = . ) and bone marrow (p < . ) (kappa value = . ) compared to contrast enhanced ct and bmb. pet/ct had upstaged cases ( %) and down-staged cases ( . %) (p < . ) (kappa value = . ). among the upstaged cases, patients ( . %) were upstaged from stage ii to iii, based on residual in pet/ct not seen in contrast enhanced ct after abdominal mass excision. four patients ( . %) were upstaged from stage iii to iv based on bone marrow uptake in fdg-pet without positivity in bma or bmb.regarding response assessment, sensitivity was % for pet and % for contrastenhanced ct (p = . ). specificity was % for pet and % for ct (p< . ). positive predictive value for pet was %, while was % for ct scan (p< . ). negative predictive value for both pet and ct was % (p = . ). five patients had nd biopsy to confirm viability of the residual lesions, lesions were negative in pathological examination (all of them were metabolic negative in pet/ct; deauville score below ). one lesion was positive in pathological examination (was positive in pet/ct; deauville score of ). conclusion: pet/ct detected additional sites compared with contrast-enhanced ct and resulted in changing stage of disease. pet scan is significantly more specific than ct in the management of children with burkitt lymphoma. background: deep sequencing of the immunoglobulin heavy chain (igh) locus indicates that each b all is composed of innumerable subclones. in many cases, subclones exhibit differing phenotypic qualities. however, it remains unclear whether subclones demonstrate distinct tissue distribution within a patient. objectives: . to quantify the extent of clonal heterogeneity in diagnostic b all specimens; . to identify variability in clonal composition between bone marrow (bm) and peripheral blood (pb) disease sites. design/method: igh sequencing was performed on purified dna from pairs of matched bm and pb patient specimens. multiplex pcr was used to globally amplify the igh locus; next generation sequencing (ngs) was performed using illu-mina® miseq. index clones (defined as ≥ % of all sequence reads in a specimen) and their subclone progeny (defined by shared nucleotide bases immediately upstream of a common jh, or n_jx) were identified using igblast-determined vh and jh alignments (http://www.ncbi.nlm.nih.gov/igblast/) and an established in-house computational pipeline. results: up to index clones per specimen were discovered in of the samples. in the remaining ( bm/pb pairs), pair did not reveal a clonal igh and was eliminated from analysis; in the other, clone frequency did not reach the % index threshold, but predominant clonal precursors were inferred by the prevalence of their subclone progeny. subclone counts ranged from to , per index clone. a combined , subclones derived from pb index clones were observed; in contrast, bm index clones gave rise to only subclones. subclone heterogeneity was observed between all paired specimens. in bm/pb pairs, index clones existed in equivalent proportions between disease sites. in contrast, bm/pb pair demonstrated high-frequency index clones in the bm ( . % & . %) with limited representation of these clones in the pb ( . % & . %, respectively); in this case, the most prevalent clone in the pb ( . %) matched the least frequent index clone in the bm ( . %). similarly, another pair showed a predominant index clone in the pb ( . %) which was below index threshold ( . %) in the bm. in paired patient specimens, index clone predominance was discovered to be overtly distinct between bm and pb. among all pairs, the extent of subclone progeny derived from each index clone showed marked variability, with far higher subclone frequency in the pb than in the bm. our data indicate that b all clonal composition differs between disease sites. valley children's healthcare, madera, california, united states background: tuberculosis (tb) presenting with hodgkin lymphoma (hl) is rare. their coexistence could lead to delay in diagnosis of both tb and hodgkin lymphoma due to the similarities in signs and symptoms of presentation. most cases have been reported in the adult literature. we describe a case series of children that were suspected to have tb and were found to have coexisting tb and hl. results: a retrospective review of hl patients in guatemala and argentina over six years, uncovered patients with simultaneous diagnosis of tb and hl. eight patients were from guatemala (incidence of . %) and from argentina (incidence of . %). there were females and males. age ranged from - years (mean . years, media years). nine patients were suspected to have tb at presentation by the referring physician. two patients were found to have tb at the time of relapse through routine tissue culture. initial systemic symptoms included fever (n = ), weight loss (n = ), and night sweats (n = ). six patients had a second systemic symptom in addition to fever. time for referral to oncology center ranged from weeks to months. nine patients were diagnosed with tb and hl through a tissue cultures and with serum quantiferon. one patient was found to have hl without tb. two patients had no systemic symptoms and the diagnosis of tb came to light through routine tissue culture. five patients had stage iiib and ivb, two stage iia and one iib at diagnosis. hl treatment was given according to the insti-tutional standards depending on stage and risk with abvd, oepa/copdac +/-radiation therapy, and ice for relapse. five patients started anti tb treatment (isoniazid, rifampin, pyrazinamide +/-ethambutol for months followed by isoniazid and rifampin for - weeks) simultaneously with chemotherapy, and three others after completing cycles. the two relapsed patients started tb treatment after cycles of chemotherapy. seven patients are alive and have been followed for months - years. one patient died during therapy, another died for causes not related to tb or hl and one is currently receiving treatment. conclusion: tuberculosis can coexist with hl. in areas were the prevalence of tb is high, microbiology investigations of biopsy specimen should be strongly considered. therapy for tb can be given simultaneously with chemotherapy. coexistence of tb and hl does not appear to affect outcomes. the children's hospital affiliated to the capital institute of pediatrics, united states background: the pi k/akt signaling pathway plays a central role in cell growth, proliferation and survival in physiological conditions. this signal pathway is considered to be an innovative targeted therapy of cancer, and its abnormal activation has been proved to be related to t-cell acute lymphoblastic leukemia (t-all). despite improved treatment strategies, such as multi-drug combination, high-dose chemotherapy and all kinds of application and popularization of hematopoietic stem cell transplantation, children with drug resistance or relapse t-all are still rather worse and its overall outcome and prognosis are much poorer than the more common b-lineage all. objectives: to explore the relationship between the pi k/akt pathway and the pediatric t-all, so as to probe the exact molecular mechanisms of t-all and provide more directions for its treatment. design/method: cases of new or recurrent acute t lymphocyte leukemia children with clinical information were collected in the children's hospital affiliated to the capital institute of pediatrics from dec. to oct. , with age and gender matched healty children as control (all was informed consent). the expressions of key genes in pi k pathway were s of s analyzed by western blot rt-pcr analysis, the pi k enzyme activities were detected by elisa,and the ccrf -cem's proliferation and its apoptosis were tested by mtt and flow cytometry technology on t-all cell lines ccrf-cem in different treatment group. the results of t-all children in clinical showed that pi k protein and gene expression level were higher apparent than the control group (p< . ), and pi k enzyme activity increased as well (p< . ); pi k inhibitor ly made a significant inhibition of cell proliferation and promoted cell apoptosis. ly also enhanced the effectiveness of clinical commonly used chemotherapeutic drug dnr. in combination ly and dnr treatment group cell viability dramatically declined, apoptosis and the apoptosis relation protein casepase expression in t-all patients was obviously higher than the control and the single drug group; pi k/akt signaling pathway related proteins and gene expression level, pi k, akt, gsk transcription in ccrf-cem were significantly higher than the control (p< . ), while pten transcription was significantly lower than the control (p< . ). the abnormal activation of pi k/akt signaling pathway might play an important role in pediatric t-all patients, especially in the cell proliferation or apoptosis. the results might provide new train of thought and direction in targeted suppress this signal pathway or in combination with other chemotherapy drugs therapy in looking for the more effective and less cytotoxic treatment of pediatric t-all. cleveland clinic children's hospital, cleveland, ohio, united states background: non-hodgkin lymphomas (nhls) are a heterogeneous group of lymphoproliferative diseases which comprise % of all childhood malignancies. nhls can be divided in to b cell lymphomas and t cell/natural killer (nk) cell lymphomas depending on immunophenotype, molecular biology, and clinical response to treatment. although nk/t cell lymphomas occurring in childhood and adolescence comprise a small portion of all lymphomas, they present many diagnostic and therapeutic challenges. the role of angiogenesis in lymphoma pathogenesis is becoming more evident. high molecular weight kininogen (hk) is a central compo-nent of the kallikrein-kinin system. it has been previously reported that cleaved hk (hka) induces apoptosis of proliferating endothelial cells and inhibits angiogenesis in matrigel plug and corneal angiogenesis models. however, the role of endogenous kininogen in regulation of angiogenesis is in tumor microenvironment is unknown. objectives: to elaborate the role of hk in lymphoma angiogenesis, we used a murine t-cell lymphoma model and compared angiogenesis and tumor growth between wild-type and kininogen deficient (mkng -/-) mice. we also evaluated the effect of hka on lymphoma cell proliferation. design/method: el- murine t-cell lymphoma cells ( × ^ ) were implanted into wild-type and mkng -/-mice. tumor size was measured using calipers and tumor volume was calculated using the formula volume = length × width^ × . . seventeen days after cell implantation, tumors were harvested and processed by immunoblotting and immunofluorescent staining. cell proliferation assays (mts) were performed to investigate any possible inhibitory effect of hka on el- cell growth, with human umbilical vein endothelial cells (huvec) were used as a positive control. results: el- lymphomas grew more rapidly and to larger sizes in mkng -/-mice compared to wild-type mice, with significant differences apparent by day after tumor implantation (p< . ). by day , the volume of tumors in mkng -/-mice was approximately . -fold larger than in wild-type mice (mean volume ± standard deviation; ± vs. ± mm , respectively, p< . ). mts assays showed that hka does not directly inhibit the proliferation of el- cells in vitro, though it does significantly impair the viability of ecs studied simultaneously. conclusion: these findings suggest that hk is an important endogenous regulator of angiogenesis and tumor growth in this t-cell lymphoma model, and suggests that hka specifically modulates endothelial proliferation in tumor microenvironment. further work is needed to understand the mechanisms underlying these findings and provide future anti-angiogenic approaches to increase the therapeutic options for patients with nhl. bruce bostrom, jack knudson, nathan gossai, joanna perkins, michael richards, jawhar rawwas, susan sencer, julie chu, nancy mcallister, yoav messinger children's minnesota, minneapolis, minnesota, united states background: osteonecrosis causes significant pain and morbidity in older patients treated for acute lymphoblastic leukemia. besides altering the schedule of dexamethasone in delayed intensification there is no other intervention known to reduce the incidence of symptomatic osteonecrosis. pamidronate has been shown to reduce bone pain from osteonecrosis but not to prevent joint collapse when advanced. objectives: to compare the incidence of symptomatic osteonecrosis in patients who received prophylactic pamidronate compared with concurrent controls. to describe any increase in side effects from the use of pamidronate. design/method: patients age to years at time of all diagnosis were given intravenous pamidronate monthly for one year at the discretion of the primary oncologist starting in the first year of therapy. concurrent controls were patients age to who did not receive pamidronate. all patients were treated according to the concurrent cog protocols and received intermittent dexamethasone during delayed intensification. patients with bcr-abl all were excluded as the use of imatinib may increase the risk of osteonecrosis. imaging was only done if osteonecrosis was suspect based on clinical symptoms. patients were censored at the time of relapse. data were analyzed as of / / . this retrospective study was approved by the children's minnesota irb. of the patients evaluated % were male and % female, % had b-cell and % t-cell. the median followup is . years with a range of . to years. pamidronate was given to patients with developing symptomatic osteonecrosis. there were concurrent controls with developing osteonecrosis. there was no significant difference in the leukemia lineage, gender distribution or body mass index (bmi) at diagnosis between groups. for all patients the median bmi was with a range of to . the age at diagnosis was significantly higher in the pamidronate group with a median of . years vs. . in the controls (p = . ). by kaplan-meier analyses the incidence of symptomatic osteonecrosis was significantly lower in the pamidronate group at % vs. % in controls. the log-rank p-value was . and the breslow p-value, which is more sensitive to early events, was . . there were no untoward side-effects from pamidronate. pamidronate infusions significantly reduced the incidence of symptomatic osteonecrosis in patients over the age of compared to concurrent controls who did not receive pamidronate. arahana awasthi, dina edani, janet ayello, christian klein, mitchell cairo new york medical college, valhalla, new york, united states background: mature b-nhl, including bl and pmbl express cd +/cd b+ and have an excellent prognosis, however, subset of patients relapse secondary to chemoimmunotherapy resistant disease and have a dismal prognosis (≤ % yr. efs, cairo et al. blood. ; gerrard/cairo et al., blood, , goldman/cairo et al. leukemia, . pv has been demonstrated to possess significant preclinical activity against indolent cd b+nhl (polson et al. can. res. ). we previously observed that obinutuzumab (anti-cd mab) significantly enhanced cell death and increased overall survival against bl (awasthi/cairo et al., bjh ) in xenografted nsg mice. however, additive/synergistic effects of pv with obinutuzumab against mature pmbl/bl are unknown. to determine the efficacy of the pv or obinutuzumab/rtx alone or in combination against pmbl and rituximab (rtx) sensitive/resistant bl cell lines. design/method: raji rh (provided by m. barth, md, roswell park cancer institute) and raji/ karpas p (atcc, usa) were cultured in rpmi. tumor cells were incubated with pv, and/or anti-cd b, mmae (generously supplied by genentech inc.) with obinutuzumab /rituximab ( ug/ml) for hr with nk cells at : e: t ratio and cytotoxicity was determined by delfia cytotoxicity assay. six to week old female nsg (nod.cg-prkdcscid il rgtm wjl/szj), were divided into groups: pbs, isotype control, pv, anticd b mab and mmae ( mg/kg). mice were xenografted with intravenous injections of luc+ bl and pmbl cells and tumor burden was monitored by ivis spectrum system. results: os of mice receiving pv alone was significantly increased compared to anticd b ab or isotype control in raji ( . vs. vs. . our preliminary data indicates that pv significantly increased survival in bl and pmbl nsg xenografts compared to anti-cd b ab alone. furthermore, pv in combination with obinutuzumab significantly enhances in-vitro cytotoxicity in bl and pmbl compared to obinutuzumab or pv alone. results: maximal grades (g) / , , and crs occurred in , , and patients, respectively. median lowest fibrinogen levels were . , . , and . g/l in patients with maximal g - , , and crs, respectively. %, %, and % of patients with maximal g - , , and crs had lowest reported fibrinogen levels of ≥ to < . g/l. eight patients (all with g crs) had very low fibrinogen levels (< g/l), which occurred before (n = ) or during (n = ) maximal crs grade or at time of improvement (n = ). no patients with maximal g - crs had < g/l fibrinogen levels. at the onset of < g/l fibrinogen levels, patient had concurrent g , and had g - increased international normalized ratio and activated partial thromboplastin. cryoprecipitate was the primary treatment in the us, and fibrinogen concentrate (fc) guidelines for tisagenlecleucel-associated coagulopathy were developed for other countries because administration of fresh frozen plasma can be problematic. fc was available at / sites for infused patients: / (g crs) and / (g - crs). cryoprecipitate was available at / sites for infused patients: / (g crs), / (g crs), and / (g - crs). risk of bleeding increases in pediatric patients with comorbid thrombocytopenia and anticoagulant treatments. / patients had g / decreased platelets within day of < g/l fibrinogen levels. fatal case of intraparenchymal cranial hemorrhage occurred during resolving crs with g hypofibrinogenemia, ongoing thrombocytopenia, and continuous veno-venous hemofiltration with citrate. hypofibrinogenemia was observed more frequently in patients with higher crs grades during/when crs was improving or resolving. fc and cryoprecipitate treatment guidelines were developed. frequent monitoring and fibrinogen replacement are needed in patients with g / crs. sponsored by novartis. its prolonged cns half-life, may allow a reduction in the number of intrathecal injections. objectives: to safely reduce the burden of therapy by reducing the number of it injections and reducing the total dose of doxorubicin with the addition of liposomal cytarabine and rituximab. design/method: patients ( - years) with cd + b-nhl with fab group b good risk (=stage i/ii and stage iii with ldh < xuln), fab group b intermediate risk (=stage iii ldh ≥ xuln and stage iv {bm blasts < %}) and fab group c high risk were eligible. patients received fab backbone therapy with the addition of six rituximab ( mg/m ) doses; two doses prior to each of two induction courses and one dose prior to each of two consolidation courses. cumulative doxorubicin was reduced from to mg/m in gr patients. after systemic methotrexate clearance, patients received age based dosing of it liposomal cytarabine. it injections were reduced from nine to five. the primary outcome is safety and toxic deaths among evaluable patients with an estimated -year survival above %, monitored by an independent dsmb. results: to date, evaluable patients, fab group b and group c ( cns positive), median age years (range - ), males, burkitt/ dlbcl with gr, ir and hr have enrolled. there has been one grade anaphylactic reaction to rituximab and one grade facial nerve palsy. no other serious adverse events were attributable to protocol therapy. there has been death from progressive disease and relapse at a median follow up of months. efs and os are % and %, respectively. our initial results show excellent efs and os, consistent with published standard of care outcomes, with the addition of rituximab and intrathecal liposomal cytarabine despite the reductions in therapy. further enrollment is ongoing and continued long term outcomes are needed to confirm early results. future randomized studies are needed to examine both short term (mucositis, infections, hospitalization days) and long term (late cardiac toxicity) endpoints. . goldman etal, leukemia, . cairo etal, jco st. jude children's research hospital, memphis, tennessee, united states background: bereaved parents identify significant spiritual needs around time of death and throughout their bereavement journeys. spirituality has been identified as a primary means by which bereaved parents can find meaning in their losses, and this ability to find meaning is associated with lower maladaptive grief symptoms. the use of spiritual coping strategies has been associated with improved coping and mental health outcomes among bereaved parents. objectives: to better understand how bereaved parents' experiences with spirituality throughout bereavement effects objective measures of grief, depression, and meaning-making. design/method: thirty participants whose children died of progressive cancer or related complications one to three years prior to participation completed an in-depth semi-structured telephone interview about their experiences with grief. participants were prompted to describe the impact of their spirituality on their bereavement processes. additionally, participants completed surveys related to grief (prolonged grief disorder questionnaire, pg- ), depression (beck depression inventory, bdi), and meaning-making (integration of stressful life experiences scale, isles). results were analyzed using a mixed methods approach including semantic content analysis of qualitative content and kruskal-wallis h test and post-hoc analyses of quantitative data. results: correlation analyses demonstrated significant differences between participants with positive and negative spiritual experiences of bereavement. participants with negative experiences of bereavement had a statistically significant increase in scores on the pg- compared to those with positive spiritual experiences signifying greater symptoms of prolonged grief. participants with negative spiritual experiences with grief had significantly lower scores on the isles, suggesting a lesser degree of adaptive integration of their losses. there were no significant differences in depression scores between groups. conclusion: bereaved parents that have a negative spiritual experience of bereavement are at increased risk for prolonged grief symptoms and are less likely to find meaning in their children's deaths than bereaved parents that describe a positive spiritual experience of bereavement. providers should consider exploration of spiritual beliefs and provision of spiritual care for parents of children facing life-limiting illnesses during treatment and bereavement. background: langerhans cell histiocytosis (lch) is an inflammatory myeloid neoplasia characterized by frequent relapse, with treatment failure associated with higher risk of death and neurodegenerative disease (lch-nd). activating somatic mutations in mapk pathway genes have been identified in almost all cases, with braf-v e in approximately % of lesions. targeted therapies have been successful in treating other refractory cancers with braf v e mutations (such as melanoma). given the central role of mapk pathway activation in lch, mapk pathway inhibition may be an effective therapeutic strategy for children with lch. objectives: the purpose of this study was to report the efficacy and toxicity profile of a retrospective cohort of patients with lch treated with mapk pathway inhibitors. design/method: medical records from pediatric patients with lch (systemic and/or lch-associated neurodegeneration) who were treated with a mapk pathway inhibitor were retrospectively reviewed from five institutions. all patients had failed at least one prior systemic therapy and had a proven mapk pathway mutation. results: all patients in this series were less than years old (median = . years; range: - years) with a median of three prior treatments (range: - ). at the time of initial mapk inhibitor use, nine of the patients had lch-nd diagnosed clinically and/or by radiographic imaging; the remaining three patients had systemic disease. patients were treated for a median of months (range: - months) with various reasons for discontinuation. three patients received combination mapk inhibitor therapies and three patients received other concurrent lch-directed therapies. four of the twelve patients had a grade or toxicity reported and three of these patients required dose reduction in order to be able to successfully resume therapy. overall survival was % with median month follow-up (range: - months) with only one patient achieving transient complete response. the remaining ten patients had partial response or stable disease and four of these patients developed progressive disease while on therapy. conclusion: mapk pathway inhibitors may be a relatively safe salvage therapy for refractory systemic lch and lch-nd but the efficacy and durability of this strategy remains to be defined. combination with cytotoxic chemotherapies may be required in order to eradicate the disease-causing cell. future prospective trials of mapk pathway inhibitors for patients with refractory lch are needed in order to directly compare their efficacy and toxicity relative to other current salvage strategies. cincinnati children's hospital medical center, cincinnati, ohio, united states background: medication adherence during maintenance therapy has been shown to have a direct relationship with disease relapse in pediatric leukemia. previous research determined that patients who are ≤ % adherent to mercaptopurine ( mp) have a greater risk for relapse. the primary aim of the present study is to examine the relationship between metabolite profiles of mp with behavioral adherence rates obtained via electronic monitoring at , , and days. it is hypothesized that patients demonstrating low levels of thioguanine (tgn) and methylated mercaptopurine (mmp) will have lower behavioral adherence rates prior to the blood draw. design/method: in a multisite, prospective study of patients ages - years diagnosed with acute lymphoblastic leukemia (all) or lymphoblastic lymphoma (lbl), mp adherence was measured across months of maintenance therapy using behavioral adherence (electronic monitoring) and pharmacological (metabolites) measures of mp. mp is metabolized into mmp and tgn. cluster analysis was used to generate three mutually-exclusive profiles of mp adherence. behavioral adherence rates were calculated for , , and days prior to the blood draw. results: this study identified three metabolite profiles of mp across months. previous research indicated that low levels of both metabolites suggest nonadherence to medication. low levels of one metabolite with high levels of another metabolite indicate adherence to mp. in this study, . % of the low tgn-low mmp group had -day behavioral adherence rates ≥ % (mean = %); . % had adherence rates < % (mean = . %). in the high tgn-low mmp group, . % had a mean -day adherence of %; . % had adherence rates < % (mean = . %). the low tgn-high mmp group had % of patients with a mean -day adherence level of %; % had adherence rates < % (m = . %). at and -days, to % of patients in the low tgn-low mmp group had adherence rates < %. conclusion: these findings suggest that electronic monitoring and metabolite concentrations can be used to monitor mp medication adherence during maintenance therapy. it is notable that there is a sub-sample of pediatric patients who are identified as being nonadherent to mp based on electronic monitoring, however, metabolite levels indicate adherence to mp. similarly, a sub-sample of patients were identified as being adherent based on electronic monitoring, but metabolite profiles indicated sub-therapeutic levels of mp. our findings underscore the clinical significance of using both objective measures of medication adherence to inform clinical decision making. cincinnati children's hospital medical center, cincinnati, ohio, united states background: hemophagocytic lymphohistiocytosis (hlh) is a life-threatening hyperinflammatory syndrome characterized by non-remitting fevers, rash, hepatosplenomegaly, cytopenias, liver dysfunction and coagulopathy, and can include central nervous system involvement. several genetic diseases cause hlh by impairing normal lymphocyte or macrophage function. the hlh panel at the cincinnati children's genetics laboratories includes genes associated with hlh and other lymphoproliferative diseases, including the genes that cause primary hlh (prf , unc d, stxbp , stx , rab a), x-linked lymphoproliferative diseases (sh d a, xiap), itk deficiency (itk), hermansky-pudlak syndrome types and (ap b and bloc s ), chediak-higashi syndrome (lyst), cd deficiency (cd ), xmen syndrome (magt ) and lysinuric protein intolerance (slc a ). deletion/duplication analysis is available as a reflex test for all genes, as copy number variations (cnvs) are not directly assessed by sequencing. objectives: the prevalence of cnvs among large groups of patients with hlh in north america is unknown. we assessed the frequency of cnvs in the genes on the hlh panel through a retrospective review of orders for deletion/duplication analysis performed after next-generation or sanger sequencing: orders for all genes on the panel, and orders of - genes from the panel. deletion/duplication analysis was performed on a custom × k microarray annotated against ncbi build (ucsc hg , march ). deletion/duplication analysis resulted in a confirmatory diagnosis in of cases ( . %). pathogenic or likely pathogenic cnvs were most common in the three x-linked genes: sh d a ( deletions), xiap ( deletions, duplication), and magt ( deletions). hemizygous deletions in xlinked genes in male patients were typically suspected after amplification failure during previous sequencing. of the autosomal recessive genes, pathogenic cnvs were observed once in each of three genes: rab a (heterozygous), lyst (heterozygous), and stxbp (homozygous). in the two heterozygous cases, a second change was not identified by sequencing, so deletion/duplication analysis did not offer a confirmatory diagnosis. in patients, deletion/duplication analysis was performed after a pathogenic or likely pathogenic variant was identified in an autosomal recessive gene during sequencing; however, in no case was a second mutation uncovered by cnv analysis. we recommend that deletion/duplication analysis be routinely performed in all male patients with hlh who lack a genetic diagnosis after sequencing of hlh-associated genes, especially if any regions failed to amplify. deletion/duplication analysis may be performed in female patients after sequencing if a genetic form of hlh is highly suspected, but the yield is expected to be low. cleveland clinic children's hospital, cleveland, ohio, united states background: the development of post-transplant neoplasia, typically from lymphoproliferative disease (ptld), is a severe complication in transplant recipients and affects approximately % of pediatric solid organ recipients. rates of lymphoma in adult heart transplantation patients are comparatively low, at less two percent at ten years. there are few published reports of the long-term outcomes of neoplasia after pediatric heart transplantation. we aimed to identify the subsequent malignancies that occurred in pediatric heart transplantation patients in a large single institution, and describe their treatment and subsequent clinical course. we performed a retrospective chart review of all pediatric heart transplant recipients followed at the cleveland clinic children's hospital from january to october . we excluded patients who died within days of heart transplantation. we reviewed in depth the history and clinical course of subjects who developed neoplasms. results: between and , patients underwent heart transplantation and survived at least days post transplantation. nine patients ( . %) developed a subsequent malignancy. in this case series, the median age at heart transplant was years old and the median time to develop neoplasia was . months. primary neoplasia included monomorphic ptld ( ), polymorphic ptld ( ), burkitt lymphoma ( ), hodgkin's lymphoma ( ), plasmacytoma-like lymphoma ( ) and epstein-barr virus-associated smooth muscle tumor (ebv-smt) ( ). one patient with hodgkin lymphoma subsequently developed monomorphic ptld, one patient with polymorphic ptld subsequently developed ebv-smt and later, an undifferentiated gastric cancer. one patient with monomorphic ptld developed an ebv-smt. evidence of epstein-barr virus was present in six of nine patients at diagnosis of first malignancy. four of nine patients received reduction in immunosuppression as a primary intervention for the initial malignancy, with two complete responses (cr), one partial response, and one with progressive disease. five patients were treated with chemotherapy, with four cr and one with progressive disease. three patients died of malignancy (recurrent ebv-smt, undifferentiated gastric cancer, and monomorphic ptld post-hodgkin disease) and two patients died of other transplant related complications. conclusion: secondary malignancies represent a significant disease burden to survivors of cardiac transplantation. as expected, much of the malignancy burden is driven by ebv. despite aggressive histology, many malignancies can be successfully cured in this setting with a multidisciplinary approach. stanford university school of medicine, palo alto, california, united states background: current treatment of langerhans cell histiocytosis (lch) is based on extent of organ system involvement and if high risk systems are affected. gastrointestinal (gi) involvement is diagnosed in about % of lch patients, and classically presents in children under years of age with malabsorption, failure to thrive, bloody diarrhea and anemia. although the gi system is considered standard risk, a mortality rate over % occurring within years of diagnosis has been reported. this study was performed due to this discrepancy and the limited number of published cases. objectives: to review the clinical course and outcomes of patients diagnosed with gi lch. design/method: a retrospective chart review of patients with histologically confirmed gi lch diagnosed in the last years identified from the bass center histiocytosis clinical database was performed. two other pediatric hematology/oncology centers (ucsf benioff children's hospital oakland and san francisco) were queried for additional cases. results: four patients with biopsy proven gi lch [ subjects ( . %) from database records and l from center queries] were identified. failure to thrive, hypoalbuminemia, bloody diarrhea and rash were the most common presenting symptoms. lch of the skin was found in all patients. risk organ systems were involved in patients. of note, subjects were of african racial background. the median age at diagnosis was . months ( . months to years), mean albumin . g/dl ( . - . g/dl), mean esr of mm/hr ( - mm/hr). all patients initially received combination therapy per lchiii protocol (vinblastine, prednisone, and mercaptopurine). two patients had recurrent disease and received second line therapy (cytarabine, cda, and local radiation therapy). all patients are alive without active disease at last follow-up ( to months after completion of therapy). a systematic approach to evaluate gi involvement should be performed in children diagnosed with lch. from our experience, combination chemotherapy for patients with lch involving the gi tract is an effective intervention for active disease. cincinnati children's hospital medical center, cincinnati, ohio, united states background: bhatia indicated that rates of mp adherence ≥ % have better clinical outcomes. those with adherence rates ≤ % have an increased risk for disease relapse. the present study investigated patterns of mp medication adherence using group-based trajectory modeling in a large sample of pediatric patients. to describe patterns of behavioral adherence during the maintenance phase of therapy for a cohort of pediatric patients ages - years who were diagnosed with acute lymphoblastic leukemia or lymphoblastic lymphoma (n = ). previous research has documented the relationship between optimal levels of medication adherence with positive health outcomes. it was hypothesized that three groups would be identified: optimal adherence, deteriorating adherence, and chronic nonadherence. it was hypothesized that patients in the optimal adherence group would have adherence rates ≥ %. those with poor adherence would have adherence rates ≤ %. design/method: the present study was a longitudinal, multisite study investigating adherence to -mercaptopurine in a pediatric cohort of patients using electronic monitoring devices. daily adherence rates (electronic monitoring of mp) were examined across -months. health outcomes were measured at quarterly intervals through medical chart reviews. results: unconditional growth curve modeling indicated that the mean percentage of behavioral adherence was . % at baseline and declined to . % at -months. three trajectories of mp behavioral adherence were identified: ) optimal adherence ( % of patients): averaging % behavioral adherence across months; ) moderate adherence ( %): relatively stable nonadherence with rates of % across months; and, ) chronically nonadherent ( %): adherence decreased from % to %. with respect to patterns of medication adherence and relationship to clinically-relevant health outcomes, there were no significant differences in health outcomes between patients in the adherent versus nonadherent trajectories, including mean absolute neutrophil counts (anc), risk for infection as measured by anc, healthcare utilization, or risk for disease relapse. although longitudinal patterns of mp behavioral adherence were not related to health outcomes, it is notable that only % of the current sample had adherence rates ≥ %. in fact, % of the current sample demonstrated adherence rates ≤ %. our findings are important for development of future adherence promotion studies in pediatric cancer. our findings underscore the relative significance of tailoring adherence promotion interventions to subgroups of patients, including those with problematic patterns of adherence. patients who demonstrate adequate levels of adherence could still benefit from less intensive, preventative interventions to sustain and improve adherence. sophie gatineau-sailliant, pascale grimard, marie-claude miron, guy grimard, anne-sophie carret, jean-marie leclerc chu sainte-justine, montreal, quebec, canada background: vertebral involvement in langerhans cell histiocytosis (lch) is still a subject of interest, due to its low frequency and the absence of management's guidelines. objectives: to provide additional information on presentation, treatment and morbidity of pediatric lch vertebral lesions, we report cases of children with vertebral lesion of biopsy-proven lch, between january st and december st , at sainte-justine university health center (montreal, quebec, canada). we conducted a retrospective study by reviewing charts and imaging of vertebral lch in a population of children (median age of . years at lch diagnosis), followed for a median duration of months. symptoms at presentation, treatment modalities and morbidities were collected. results: vertebral lesions were present at lch diagnosis in of cases. they were usually diagnosed secondary to back pain in of cases and were asymptomatic in only one case. despite an epidural extension in of cases, no child developed neurological symptoms. lesions frequently involved vertebral body ( of cases) and were rarely unstable ( of cases). out of vertebral lesions, most of them had a dorsal localization ( of lesions) and of patients had lch in multiple vertebrae. at diagnosis, median vertebral height loss was . % compared to % at last imaging control. most used imaging modalities were pet-scan and plain x-rays. treatments were diverse and consisted in chemotherapy in all children but three and bisphosphonates in only cases. radiation therapy was not used in any patient. six out patients did benefit of an orthosis. a lch recurrence was observed in patients and involved vertebrae in cases. one patient with treatment-resistant lch disease had relapses, and required multiple lines of treatment. all children were alive and disease-free at their last follow-up, patients having radiological vertebral sequelae and only had clinical sequelae. our study is consistent with the epidemiological data described in larger cohorts of children with vertebral lesions of lch and the favorable prognosis associated with such lesions. nevertheless, aggressive treatment and long term follow-up seemed to be essential as recurrences are s of s not rare and spontaneous bone regeneration often incomplete. plain x-rays appears to be a good follow-up tool for vertebral lesions as it allows reliable measures, less exposure to radiation at lower cost. national cancer institue, giza, giza, egypt background: acute lymphoblastic leukemia (all) is the most common type of childhood cancer and also the most complicated in the treatment, so it requires many interventions for both treatment and to alleviate suffer form side effects. pancreatitis is one of the toxicities, which is more common in all as it appears in about % of the patients. it occurs in many drug combinations which induce pre-pancreatitis and even direct destruction of pancreatic tissues. pancreatitis can be induced by many drugs used in the treatment such as chemotherapeutic agents or supportive treatment. lasparaginase is the backbone drug of the treatment of all in which to doses are required to achieve complete remission status in the induction phase of treatment and to doses in the maintenance phase.it is an enzyme that destructs the l-asparagine amino acid into aspartic acid and ammonia thus deplete the asparagine from the extracellular matrix . many drugs are investigated for their effect on treatment of induced pancreatitis such as interleukin- , nsaid as antiinflammatory, glycerin tri nitrates as improvement of microcirculation, tnf-alpha antibody, paf inhibitor as specific anti-inflammatory and low molecular weight heparin .none of the drugs was investigated for their ability to prevent the occurrence of pancreatitis. objectives: this study was designed to evaluate the protective effect of enoxaparin and diclofenac against l-asparaginase induced pancreatitis design/method: acute pancreatitis was induced in rats by intra-muscular injection of l-asparaginase ( i.u/kg) given daily for five days. enoxaparin was given subcutaneous ( i.u/kg) and diclofenac was given intra-peritoneal ( mg/kg) daily for five days. then, markers of pancreatic injury, lipids, immune cell infiltration and oxidative stress were analyzed with histo-pathological examination of the pancreatic tissue results: during acute pancreatitis, oxidative stress markers were significantly changed as indicated by reduced tis-sue glutathione and increased malondialdehyde levels. this was accompanied with significant increase in immune cells infiltration as indicated by high levels of myeloperoxidase and pro-inflammatory cytokine tnf-alpha. triglyceride only showed increase level. treatment with enoxaparin and/or diclofenac restored levels of biochemical markers including serum alpha-amylase, reduced glutathione, malondialdehyde, pro-inflammatory cytokine tnf-alpha, myeloperoxidase and triglyceride. histological injuries of pancreatic tissues as vacuolation and necrosis of epithelial lining pancreatic acini, inflammatory cells infiltration and focal pancreatic hemorrhage were also reduced by treatment with enoxaparin and/or diclofenac. the present study emphasizes the potential protective effect of enoxaparin and diclofenac against l-asparaginase induced pancreatitis background: rosai dorfman disease (rdd), or sinus histiocytosis with massive lymphadenopathy (shml), is a rare condition of immune dysregulation of unknown etiology arising from the massive accumulation of non-langerhans type histiocytic cells inside lymph nodes. the disease classically presents as bulky, painless lymphadenopathy often associated with infection showing distension of lymph node sinuses by abundant histiocytic cells (cd a(-), s- (+)/cd (+)). in some cases, the disease can be self-limiting, but in cases with a prolonged chronic course of exacerbations and remissions, those with extranodal involvement, or disease that threatens vitals structures, treatment may be necessary. there is no treatment consensus. to describe a case of life-threatening, unresectable, recurrent rdd successfully treated with langerhans cell histiocytosis (lch) -inspired therapy. design/method: we compared this case to the current literature on chemotherapeutic treatments for rdd. we searched pubmed, ovid, and google scholar for similar cases. we believe this to be the first reported case of using lch therapy to successfully treat rdd. an -year-old male presented to an outside hospital with two years of massive neck swelling causing torticollis. biopsy confirmed rdd. he was intermittently treated with courses of antibiotics with partial response. surgical removal of the affected lymph nodes was unsuccessful due to proximity to the spinal cord. two years later, the patient presented to our institution. he was initially treated with prednisone with a fast tapering dose, but after a second relapse the decision was made to try chemotherapy following the lch- protocol of weekly vinblastine ( mg/m ), -mp ( mg/m ), and high dose steroid bursts. he experienced two additional relapses off therapy at ages and years old, including cmv(+) associated septic shock and cytokine storm requiring rapid response, picu admission, and ionotropic support. this last episode was treated with a more prolonged induction and maintenance therapy. an extended and slowly tapered maintenance therapy regimen of . years of daily -mp, monthly vinblastine and steroids with a slowly tapered dose during his fourth remission has resulted in -months of continuous complete remission-the longest stretch of his life. no similar cases were found. literature search demonstrated no consensus regarding the most effective treatment of rdd, with no previous cases being successfully treated following lch chemotherapy protocols. we hypothesize that the multi-agent relatively mild lch- therapy mitigates the immune dysregulation of rdd. this case suggests that lch- therapy can be used to treat cases of rdd that is not amendable to surgery or observation. nicklaus children's hospital, miami, florida, united states background: central venous catheters (cvc) are necessary in the management of patients with malignancies, especially children. patients with acute leukemia (al) have higher rates of central line associated complications such as bloodstream infections compared with other malignancies. objectives: to examine the choice of placement of cvc and the differences in outcome between peripherally inserted central catheters (picc) and ports in patients with leukemia during induction. design/method: retrospective chart review of patients with newly diagnosed leukemia at nicklaus children's hospital between and . results: ninety four patients with a new diagnosis of leukemia undergoing induction chemotherapy were identified. the average age was . years. overall, ( . %) patients had a port placed and ( . %) had a picc placed. the decision for picc or port was subjective and physician based. the main outcome measures were local inflammation/infection, bacteremia, thrombophlebitis, blocked catheter and premature removal. the most common complication was bacteremia ( . %). in a multiple logistic regression analysis for predicting whether patients had at least one complication, results showed that having at least one complication is . times the odds in patients with aml compared to patients with all (p = . ). when comparing picc vs. ports, patients with picc had more frequent episodes of blocked catheters ( . %) and premature removal ( . %) compared to the patients with ports ( . % and . %) (p = . and p = . respectively) during induction. local inflammation, bacteremia and thrombophlebitis were not statistically different (p = . , p = . and p = . respectively). the most common place for port placement was the right subclavian vein ( %). there was no significant association between port location and having at least one complication (p = . ). acute lymphocytic leukemia subgroup analysis: fourteen patients ( %) in the picc group had at least one complication and ( %) in the port group but that was not statistically significant (p = . ). our series showed a higher incidence of blocked catheters and premature removals with picc compared to ports in patients with leukemia during induction. the choice of placement of picc vs port was subjective and physician based. patients with all, despite receiving steroids and asparaginase during induction, did not show a statistically significant increase risk in thrombosis or infection but larger numbers may be needed in future studies. university of california, san francisco, san francisco, california, united states background: hemophagocytic lymphohistiocytosis (hlh) is classically a disorder of young children meeting systemic hyperinflammation criteria. presentation in late adolescence is uncommon. furthermore, though cns signs occur in - % of cases, initial isolated neurologic presentation is rare, frequently resembling encephalitis or demyelinating disorders. these cns signs can be isolated or precede systemic disease, delaying hlh diagnosis. hlh declaring in adolescence with predominant psychiatric features has not been well documented. objectives: to describe a case of cns hlh presenting with neuropsychiatric features in absence of classic hlh criteria. design/method: retrospective review of clinical, radiologic, histologic, immunophenotypic, and molecular features of a patient with cns hlh. a -year-old female presented with acute-onset headaches following nine months of progressive anxiety, short-term memory loss, emotional lability, perceptual disturbances, and hypomania. brain mri demonstrated numerous enhancing t hyperintense supratentorial and infratentorial white matter lesions in the left thalamus and caudate head. brain biopsy showed histiocyte-rich inflammation and associated demyelination. extensive evaluation including universal microbial pcr failed to reveal underlying infection or malignancy. past medical history was notable for presumptive pulmonary sarcoidosis diagnosed months prior with progressive respiratory failure with associated granulomatous pulmonary nodules which responded to systemic immunosuppression. at presentation of her neuropsychiatric symptoms, she had normal sil- r, ferritin, fibrinogen, and triglycerides. there was no pancytopenia, coagulopathy, bone marrow hemophagocytosis, fevers, or splenomegaly. given the possibility of partial immune suppression of systemic symptoms and the prominent neurologic symptoms, hlh screening labs were sent and notable for decreased natural killer and cytotoxic t lymphocyte function, normal granzyme expression and cd a mobilization, and absent perforin expression. genetic testing confirmed compound heterozygous mutations in prf (c. g>a, c. a>c) and familial hlh type . she was treated with low-dose dexamethasone and intrathecal chemotherapy per hlh- . due to lack of evidence of systemic inflammation, vp- and high-dose steroids were held. within one week of initiating therapy, she had decreased anxiety and improved cognition, with sustained, incremental neuropsychiatric improvement with additional intrathecal treatments. she tolerated dexamethasone tapering without symptom flare. mri also demonstrated parenchymal lesion improvement. for definitive treatment, she underwent unrelated allogeneic hematopoietic cell transplantation and remains at neurologic baseline as of eight months post-transplant with ongoing imaging improvement. conclusion: this case of familial hlh with compound heterozygous perforin mutations in an adolescent with isolated neuropsychiatric symptoms illustrates that cns hlh may be an underrecognized phenomenon in absence of systemic signs. standard hlh therapy may effectively reverse these symptoms with associated radiologic responses. rush university children's hospital, chicago, illinois, united states background: posterior reversible encephalopathy syndrome (pres), a recognized complication of pediatric leukemia treatment has been reported in up to % patients in various series. hypertension, chemotherapy and cortical spreading depression have been implicated in the pathophysiology. due to the combinations used, it is difficult to identify the offending drug, several have been implicated. since delay of chemotherapeutic treatment in children with high risk leukemia is unfavorable, it is important to recognize the characteristic radiologic findings, manage appropriately and reintroduce the treatment as soon as possible. pharmacoethnicity is now recognized as an important factor for variation in neurotoxicity in children with all. ethnic differences in reported pres events in pediatric patients with all has not been well described in literature. to describe the factors associated with pres in a cohort of high risk pediatric all patients at a single institution. design/method: a total of children with an average age of years ( - years) diagnosed with all between - were retrospectively reviewed for the occurrence of pres. various demographic factors, therapy received, clinical features, radiology related findings and management were reviewed. a search for all published articles on pres in leukemia was conducted using pubmed databases. results: five ( %) children (average age . years) developed pres during days - of induction. % of the patients that developed and % of those that did not develop pres were hispanic. all the patients that developed pres and % of those that did not were diagnosed with high risk all. all patients received vincristine, % received daunomycin and intrathecal methotrexate and % received asparaginase in the week prior to the event. mri findings confirmed pres in all patients with no evidence of methotrexate related leukoencephalopathy or leukemia. at the time of pres all patients were in remission based on mrd and spinal fluid cytology. two-thirds of the patients had seizures and hypertension at the time of the event with no prior history of either. all patients had complete recovery of normal mental status after resolution of pres. a higher incidence of pres than previously reported was noted in our series. hispanic ethnicity, high-risk all and exposure to vincristine, daunomycin and intrathecal methotrexate in induction were associated with pres in our cohort. a new association that emerged was that of hispanic ethnicity with pres .larger studies to understand the importance of pharmacoethnicity in pres may help in individualization of chemotherapy based on ethnic differences. children's hospital of illinois, peoria, illinois, united states background: hyper ige syndrome is a primary immunodeficiency characterized by susceptibility to skin and lung infections as well as increased propensity for malignancy. hemophagocytic lymphohistiocytosis (hlh) is a syndrome characterized by overwhelming activation of t lymphocytes and macrophages occurring as either primary hlh caused by genetic abnormalities or secondary hlh associated with infectious, malignant, metabolic, or immunodeficiency causes. we describe the first case to our knowledge of hlh in a patient with hyper ige syndrome. to describe a case of hlh in a pediatric patient with hyper ige syndrome. results: a -year old caucasian male with known autosomal dominant hyper ige syndrome (stat mutation) was transferred to the pediatric intensive care unit secondary to concern for septic shock. the patient had persistent slow bleeding from oral lesions and central catheter sites despite the addition of aminocaproic acid and recombinant factor viia. he also required numerous blood product transfusions sec-ondary to anemia and thrombocytopenia. clinical suspicion was high for hlh and the patient met criteria for diagnosis of hlh with the following: ferritin > , ng/ml, triglycerides mg/dl, decreased nk cell function with the sample only containing % nk cells, elevated soluble il- receptor at u/ml, splenomegaly, and fever. infectious workup was remarkable for a positive ebv qpcr with , copies/ml suggestive of ebv driven secondary hlh. familial hlh testing was unable to be completed. therapy was initiated based upon the hlh- study. the addition of ruxolitinib and anakinra were considered but the patient declined rapidly prior to treatment. ct of the head was concerning for a stroke with signs of edema and increased intracranial pressure likely leading to the development of symptoms consistent with brain stem herniation. the decision was then made to withdraw care. conclusion: to our knowledge, this is the first report of hlh in a patient with hyper ige syndrome. diagnosing hlh requires a high index of suspicion in critically ill patients, and prompt initiation of therapy is essential. this challenging case of hlh in a patient with hyper ige syndrome highlights the diagnostic challenge, variable presentation, and need for effective therapy in this vulnerable patient population. background: adolescents and young adults (ayas) with cancer are at risk for psycho-social as well as physical symptom burden during cancer therapy. the purpose of this study is to explore psychological and physical symptoms endorsed by aya while receiving therapy for cancer design/method: surveys were given in both inpatient and outpatient settings during cancer therapy. symptom screening in pediatrics tool (sspedi) and memorial symptom assessment scale (msas). symptoms severity was rated by teens on a point likert scale. spss , used for statistical analysis. results: : a total of aya on cancer therapy (age range - . years) % female, % male, . % acute leukemia, . % solid tumors, and . % diagnosis was not reported. % of aya on cancer therapy reported at least or more symptoms, % reported > symptoms cluster. of the physical symptoms that were reported as most distressing to the teens, mouth sores and headaches were the top causes. of the physical symptoms that were most frequently endorsed; fatigue was on the top ( %), followed by change in appetite %, vomiting %, and pain %., the least was bowel habit changes. aya rated sadness as the most frequent psychological symptom %, followed by feeling angry %, and scared %. statistically significant difference was noticed based on gender difference with more females reported symptoms (p = . ), while type of cancer (acute leukemia versus solid tumors) was not statistically different. conclusion: aya with cancer reported multiple physical and psychological symptoms with significant distress. females seem to report more symptoms compared to males. screening aya for cancer therapy related symptoms is feasible during routine visits and adds important information about the aya well-being. background: sinus histiocytosis with massive lymphadenopathy (shml), also known as rosai-dorfman disease, is a rare histiocytic proliferative disorder of unknown etiology. many treatment modalities have been employed; however, no uniform guidelines exist. objectives: literature review of treatment options for shml. design/method: chart review was performed on pediatric patients diagnosed with shml at the children's hospital at montefiore between and after irb approval. inclusion criteria included children between the ages of and years with shml. exclusion criteria included children with cutaneous shml. four cases of shml seen at montefiore are described. a comprehensive review of the literature identified additional cases published between and . manuscripts that did not include the treatment modality or outcome were excluded. results: many of the patients with shml responded to observation alone. of patients, patients were observed, with ( %) having resolution of disease, five having stable disease, and five being lost to follow-up. one patient received subsequent systemic therapy. surgical management was con-ducted upfront in patients. of those, ( %) had resolution of disease, one had stable disease, and one had recurrence with no further therapy noted. of the remaining nine patients, % were successfully treated with systemic therapy, consisting of either steroids ( ) or steroids and chemotherapy ( ). systemic therapy was used as first-line therapy in patients. steroids alone or in conjunction with chemotherapy resulted in resolution of disease in / and / patients ( / , %), respectively, with four patients having stable and three with progressive disease. chemotherapy without steroids resulted in resolution of or stable disease in / patients. radiation was ineffective. conclusion: shml is a rare disease with no published guidelines for treatment. from the results of the cases and a detailed review of the literature, it can be suggested that observation may be considered as first line management in patients providing there are no significant symptoms. for patients who are symptomatic or have significant progression, surgery may be considered. in patients with recurrence or refractory disease, steroids and/or chemotherapy may be used. the presence of nodal or extra-nodal disease did not seem to have a significant impact on the course of treatment. given the rarity of the disease, it is difficult to conduct a randomized control trial. further work, involving collaboration between centers and cooperation with the international rare histiocytic disorders registry would be helpful. boston children's hospital, boston, massechusettes, united states background: increasing census and intensified work compression on the inpatient oncology service at our institution was identified as leading to resident dissatisfaction, impaired resident learning and decreased perceived quality of patient care. objectives: to evaluate the impact of a redesign of a pediatric inpatient hematologic malignancy (ihm) service on resident perceptions of the educational value of the rotation and safety of patient care. design/method: during the - academic year, we initiated a bundled intervention on the ihm service. modifications included ) decreased patient volume: the ihm service was divided into two teams, utilizing an extra attending -a teaching service consisting of residents and fellows and a team comprised of nurse practitioners. ) intentional patient team assignment: patients were deliberately assigned to a care team based on educational opportunities and provider skill sets. ) intentional attending faculty selection: attending faculty with deeper clinical and teaching experience were selected to supervise on the teaching team. ) increased weekend staffing. after completing the service, junior residents completed an electronic survey to evaluate their perceptions of the educational value of the rotation, as well as their ability to deliver safe care while on the rotation. fisher's exact tests were used to compare responses from residents in who experienced the redesign to residents in , whose experience results: survey completion rates were % ( / ) in and % ( / ) in . intervention residents were significantly more likely than comparison group residents to choose the answers "very good" or "excellent" to describe both the overall quality of the rotation ( % intervention vs. % comparison, p< . ) and the educational experience on rounds ( % intervention vs. % comparison, p< . ). intervention residents also reported caring for fewer average primary patients daily on weekdays as compared to comparison residents ( . vs . patients, p< . , % ci - . to - . ). furthermore, intervention residents were more likely than comparison residents to "agree" or "strongly agree" that they could provide safe patient care on weekend days ( % intervention vs. % comparison, p< . ) and on nights ( % intervention vs. % comparison, p< . ) while on the oncology service. a redesign initiative of an oncology service with the development of a new teaching service led to improved resident perceptions of the educational value of the rotation and ability to provide safe care to patients. this approach could be useful to other services and institutions to promote similar outcomes in resident education and patient care. background: alk-positive histiocytosis is a rare histiocytic proliferative disorder that has been reported in three infants presenting primarily with hepatosplenomegaly, anemia, and thrombocytopenia. given the rarity of this disease, there are no standard treatment algorithms for this diagnosis and the disease course and outcomes remain largely unknown. the published series describes treatment ranging from monitoring alone to multi-drug chemotherapy regimens. there was ulti-mately resolution of presenting symptoms in all three cases despite varying treatment strategies. objectives: to report a newly diagnosed case of alkpositive histiocytosis that was treated with a novel approach using cytarabine monotherapy. results: a full term male infant presented at birth with difficulty feeding and hyperbilirubinemia. over the first few weeks of his life, he subsequently developed thrombocytopenia, transaminitis, and profound hypoalbuminemia. by six weeks of life, he was experiencing significant abdominal ascites requiring repeat paracenteses, massive hepatosplenomegaly, respiratory distress secondary to abdominal distension, anemia, and coagulopathy. he underwent numerous diagnostic tests, including a liver biopsy followed by a bone marrow biopsy that showed alk-positive histiocytic infiltrates in both sites. treatment was initiated with cytarabine mg/kg/day x days, repeating every weeks. throughout his course of five cycles of treatment, he experienced intermittent fevers and mild nausea with no other adverse events. by the end of five cycles, his hepatosplenomegaly resolved, his blood counts normalized, he demonstrated weight gain on oral feeds, and his liver enzymes normalized. he is currently months post completion of therapy and remains well with a normal physical exam and laboratory values. conclusion: treatment of alk-positive histiocytosis with lose dose cytarabine resulted in complete resolution of our patient's symptoms with minimal treatment related adverse effects, and few long-term treatment related risks. given the rarity of the diagnosis, the reporting of effective novel treatment options is important for future patient care. background: adult patients with melanoma or lung cancer harboring braf v e have benefitted from the development and subsequent approval of specific braf inhibitors. as such, delineating the subset of similarly targetable pediatric oncology patients may spur development and rational use of these inhibitors in children. importantly, other point mutations and fusions of braf may also be targetable in s of s children analogous to recent emerging data in adult cancer patients. objectives: to define the genomic landscape of known and novel braf alterations and raf fusions in pediatric malignancies and report index cases with clinical response to braf or mek inhibitors. design/method: dna was extracted from microns of ffpe sections of , tumors from pediatric (< years of age) oncology patients, and cgp was performed on hybridization-captured, adaptor ligation based libraries to a mean coverage depth of x for up to cancer-related genes plus introns from genes frequently rearranged in cancer. genomic alterations (ga) included base substitutions, indels, copy number alterations and fusions/rearrangements. a total of ( . %) braf-altered pediatric malignancies were identified. ( . %) harbored a single kinaseactivating braf short variant, indel, or fusion. an alteration resulting in reduced braf kinase activity was identified in ( . %) tumors while ( . %) tumors harbored multiple braf alterations, of which contained at least a single activating short variant. the remaining tumors ( . %) contained functionally uncharacterized variants. kinaseactivating braf alterations were identified in diverse tumor spectra comprised of brain tumors ( . %; subtypes), carcinomas ( . %; subtypes, with melanoma constituting % of cases), hematological malignancies ( . %; subtypes), sarcomas ( . %; subtypes), and extracranial embryonal tumors ( . %; subtypes). seventy-two ( . % of braf-altered cases) braf fusions were identified, ( . %) of which were kiaa -braf; involved the novel fusion partners: stard nl and khdrbs . seven ( . %) raf fusionpositive cases, predominantly brain tumors ( ), were identified; involved the novel fusion partners: tmf and sox . index cases of response to therapy of intracranial tumors will be presented. we describe a population of pediatric patients with targetable braf alterations predominantly enriched in primary intracranial tumors, but spanning diverse solid tumor types and hematologic malignancies. we additionally report a cohort of raf fusion-positive patients. an index case and multiple previous reports suggest raf or mek inhibitors may benefit pediatric patients with either intracranial or extracranial disease, and development of such drugs in pediatric indications is strongly warranted. background: diffuse midline gliomas (dmg) with h k m mutation, including diffuse intrinsic pontine glioma (dipg), are the leading cause of brain tumor-related deaths in children. there are no effective therapeutic strategies and the median survival remains dismal. genomic studies have identified a recurrent mutation in the majority of dmgs involving a lysine to methionine substitution (k m) in histones . and . , resulting in changes in the epigenetic landscape that dysregulate gene expression and promote gliomagenesis. panobinostat, a multiple histone deacetylase (hdac) inhibitor, was found to be one of the most effective agents against dipg patient-derived cell cultures and xenograft models in previous studies and is presently in clinical trial for dipg. hdac inhibition with panobinostat may also exhibit activity against h k m+ diffuse midline gliomas of the thalamus and spinal cord. to evaluate the effect of panobinostat as a single agent against patient-derived thalamic and spinal cord h k m+ diffuse midline glioma cell cultures and in an orthotopic xenograft murine model of h k m+ spinal cord glioma. design/method: patient-derived thalamic and spinal cord h k m+ diffuse midline glioma cell cultures were treated with single agent panobinostat at a range of concentrations. cell viability was evaluated using the celltiter-glo assay. panobinostat was systemically administered to orthotopic xenograft murine models of luciferase-expressing spinal cord h k m+ diffuse midline glioma. response to panobinostat was evaluated with ivis in vivo imaging. results: hdac inhibition with panobinostat significantly decreases cell proliferation with an ic of nm and nm in the spinal cord and thalamic glioma patient-derived cell cultures respectively. panobinostat slowed tumor growth in murine models of spinal cord glioma by . -fold in the brain (p = . , n = ) and -fold in the spinal cord (p = . , n = ) when compared to vehicle controls after week of administration. panobinostat is in clinical trials for dipg. this study suggests that hdac inhibition with panobinostat may also be beneficial for patients with thalamic and spinal cord diffuse midline glioma h k m mutants. background: brain tumors are the most common solid tumor of childhood and the leading cause of childhood cancer deaths. while medulloblastoma is the most common malignant brain tumor of childhood with a -year survival - %, children with high-grade gliomas (hggs) such as glioblastoma multiforme (gbm) fare much worse with a -year survival of - %. implicated in this poor outcome is the presence of treatment resistant brain tumor stem-like cells. gbm stem-like cells (gscs) have been implicated in tumor growth, treatment resistance and patient relapse, making them a key therapeutic priority. antipsychotic drugs (apds) have been used for decades in various psychiatric clinical settings and are associated with a lower incidence of cancer, including malignant brain tumors. currently, atypical apds are being evaluated for their potential to alleviate cancer and treatment induced side effects. furthermore these drugs may have direct anti-tumor effects, potentially via inhibition of dopamine d receptors (drd ). objectives: determine the anti-cancer effects of atypical apds on gbm stem-like cells design/method: the anti-cancer effects of apds (quetiapine and risperidone) were evaluated on gbm stem-like cell lines developed in our laboratory (glio and ) and the group medulloblastoma cell line hdmbo . cell proliferation/viability was determined using trypan blue exclusion and mts assays. the effect of apds on cancer stem cell self-renewal was determined by neurosphere assay. receptor expression and apds effect on cell cycle proteins were examined by western blot analysis. results: western blot analysis of gscs and hdmbo demonstrated robust drd expression indicating a viable therapeutic target. both apds induced dose dependent cell death of all cell lines tested. treatment with only um of either apd for days significantly reduced cell proliferation by % (hdmbo ) and - % (gscs). consistent with these findings, we observed an increase in cell cycle inhibitors p and p . furthermore at day both apds induced a robust increase in gsc death, approximately % compared to only % in non-treated controls. lastly, um apds significantly reduced gsc neurosphere formation compared to untreated controls by up to % suggesting inhibition of gbm stem cell self-renewal. our data indicates that clinically relevant concentrations (low micromolar) of these apds induce anticancer effects in both gscs, which are enriched with tumor initiation/propagation properties, and in the group (myc amplified) medulloblastoma cell line. these apds represent strong candidates as potential adjuvant therapies for the treatment of these brain tumors. background: while the poor prognosis for high risk neuroblastoma (hrnb) underscores the need for new treatment strategies, the elucidation of specific biologic subsets of neuroblastoma suggests a way to improve disease management. the identification of agents that target specific molecular pathways associated with the development or progression of diseases holds promise. dfmo, an inhibitor of odc, has been shown to decrease lin and mycn and target cancer stem cells in preclinical studies. currently % of patients undergoing immunotherapy relapse. dfmo is in studies to prevent relapse after immunotherapy and may be helpful during immunotherapy as well. the hypotheses for this study were that: ) the incorporation of a targeted therapy, selected based upon upfront tumor genomic interrogation, into standard induction chemotherapy for hrnb is safe, feasible and may increase the pr/cr/vgpr response rate at the end of induction therapy; and ) the addition of dfmo as maintenance during immunotherapy is safe and feasible and may decrease the relapse rate for hrnb. a multicenter feasibility pilot trial in subjects with newly diagnosed hrnb within the beat childhood cancer consortium. at diagnosis, patients' tumors underwent dna exome and rna sequencing which were analyzed within a molecular tumor board to identify the single best drug of targeted agents to be added to cycles - of induction chemotherapy. after consolidation with asct and radiation, the patients received dfmo along with standard dinutuximab and retinoic acid and dfmo for years after immunotherapy. patients were evaluated for additional toxicities with the addition of targeted agents and dfmo in addition to induction response. results: the pilot study of eligible patients has shown this process to be feasible. all patients have completed induction portions of the study. the combination of targeted agent with chemotherapy was shown to be safe without any unexpected toxicities. delays between induction cycles were < weeks and related to surgery, infection, or thrombocytopenia. the induction response demonstrated % cr/vgpr/pr rate, which suggests improvement over historical %. in addition, patients were eligible for the combination of dfmo with dinutuximab and retinoic acid was well tolerated and safe without additional toxicities due to dfmo. the pilot study of patients has shown the process of genomic sequencing and addition of a targeted agent to upfront chemotherapy and addition of dfmo to dinutuximab and retinoic acid maintenance therapy in newly diagnosed hrnb patients and is feasible and safe without any unexpected toxicities. background: identifying sub-populations of medulloblastoma tumors with stem cell-like properties holds promise for reducing disease recurrence, but there is no known unifying marker of medulloblastoma cancer stem cells. the granulocyte stimulating factor receptor (gcsf-r or cd ) is well understood in the context of hematopoiesis, but its role in solid tumor pathogenesis is less clear. neuroblastoma and melanoma subpopulations expressing gcsf-r have cancer stem cell properties of chemoresistance and increased tumorigenicity, and are enriched in tumors after chemotherapy. gcsf-r activation leads to signaling through the jak-stat pathway, suggesting a potential therapeutic target. we hypothesized that a subpopulation of medulloblastoma cells would express the gcsf-r and that this subpopulation would demonstrate chemoresistance and response to inhibitors of the jak/stat pathway. objectives: our objective was to identify a subpopulation of medulloblastoma cells expressing the gcsf-r and determine their relative growth rates, tumorigenicity, and responses to chemotherapy and jak/stat inhibition. design/method: medulloblastoma cell lines were sorted via flow cytometry for gcsf-r surface expression. subpopulations of gcsf-r-positive and -negative medulloblastoma cells were then monitored for growth by continuous live cell imaging. responses to chemotherapy were measured in subpopulations of gcsf-r-positive and -negative medulloblastoma cells using continuous live cell imaging to measure percent cell confluence and cell viability assays. ic values were calculated for each cell line and each agent. parental medulloblastoma cell lines and isolated gcsf-r-positive and -negative subpopulations were also treated with the jak / inhibitor ruxolitinib and growth rates, viability, and ic values were calculated. results: gcsf-r surface expression was identified on . - . % of medulloblastoma cell lines. isolated gcsf-r positive cells demonstrate a slower growth rate compared to gcsf-rnegative or parental unsorted medulloblastoma cells. gcsf-r positive cells are more resistant in vitro to vincristine, etoposide, and carboplatin, when compared to the gcsf-r negative population and an unsorted population of the same cell line. ruxolitinib is cytotoxic to medulloblastoma cells in vitro, with higher ic values noted in gcsf-r positive cells compared to unsorted and gcsf-r negative cells. we show that a subpopulation of gcsf-r positive cells are present in multiple medulloblastoma cell lines via flow cytometry, and that isolated gcsf-r-positive cells have a slower growth rate than gcsf-r-negative or unsorted populations. we also show that ruxolitinib has in vitro activity against medulloblastoma cell lines. we propose that jak inhibition may represent an adjunct therapy targeting overall tumor burden and specifically targeting the gcsf-r-positive subpopulation of medulloblastoma cells that may drive tumor recurrence. we investigated the efficacy of intensified adjuvant chemotherapy in osteosarcoma patients. design/method: we retrospectively analyzed the medical records of children with osteosarcoma treated at asan medical center between and . all patients received a -drug induction consisting of cycles of cisplatin and doxorubicin along with cycles of methotrexate (map), and proceeded to surgical resection. adjuvant ct was map or map with the additional ifosfamide and etoposide (mapie), and mapie was mainly considered for poor responders (tumor necrosis below %) or patients with metastases. results: among patients, patients had metastases at diagnosis. surgery was conducted in patients who responded to induction ct, and showed over % tumor necrosis. among patients who proceeded to adjuvant ct, and patients received to map and mapie protocols. with a median follow-up of months, the -year overall survival (os) and event-free survival (efs) rates of all patients were % and . %. of those patients, patients recurred, and of them died of disease progression. relapsed patients received salvage ct and/or surgery, and were rescued after autologous stem cell transplantation (sct). three patients developed treatment-related acute myeloid leukemia, and they are alive after allogeneic sct. according to the response to neoadjuvant ct, the os rates of good responders (n = ) and poor responders (n = ) were % and . % (p = . ), and efs rates were . % and . % (p = . ). of the poor responders, patients received map as adjuvant ct, and the other received mapie. the os rates of map and mapie group were . % and . % (p = . ), and efs rates were . % and . % (p = . ), respectively. when patients were classified into three groups: . localized disease & necrosis ≥ % (n = ), . localized disease & necrosis < % (n = ), . metastatic disease (n = ), survival rates were in the order of group > > (os = %: . %: . %, efs = . %: . %: %). in each group, intensified adjuvant ct by mapie did not improve survival outcomes. conclusion: initial metastatic disease and poor histological response to neoadjuvant ct were major risk factors for poor survival in osteosarcoma patients. we found that adding ifosfamide and etoposide to map did not improve survival outcomes of patients with adverse risk factors. more effective adjuvant therapy for these patients is needed. background: circulating cell-free dna (cfdna) that shed from tumors into circulation have been used for noninvasive molecular profiling in adult cancers but little is known about its utility in pediatric cancers. pediatric patients with metastatic and refractory solid tumors are known to have poor survival rates, and a key challenge in their management is obtaining biopsy samples especially at times when disease is widely spread or the patient is physically unfit for sampling. the development of a noninvasive profiling strategy is critical for optimizing molecularly guided therapy and assessing response to treatment. in this study, we want to determine the utility of cfdna to noninvasively analyze the molecular profiles of pediatric solid tumors such as neuroblastoma (nb), osteosarcoma (os), and wilms tumor (wt). design/method: tumor, plasma, and matched controls were collected from patients with nb, wt, and os, at diagnosis or time of disease progression. cfdna was extracted from the plasma and analyzed through multiple methodologies including a targeted next generation sequencing panels and shallow whole genome sequencing (swgs). results: fifteen nb patients, os patients, and wt patients had tumor molecular profiles known from different targeted next-generation sequencing platforms. in the cfdna of / nb patients, somatic mutations and copy number alterations previously reported in the tumors were detected, including recurrent nb drivers such as mycn amplification, alk, and atrx mutations. mutations not detected in the original tumor were also found in / nb patients including nras, mll , arid b, some of which are potentially actionable. in os, mutations known from the tumor were found in the cfdna of of patients, including atrx and notch mutations, as well as copy number alterations such as cdk amplification, which has targetable therapeutics available. of the two wt patients analyzed, cfdna revealed the same mutations as tumor in one patient, however in a cohort of patients where tumor was not available, cfdna revealed recurrent driver mutations such as amer , dicer . it is feasible to noninvasively identify somatic mutations and copy number alterations in cfdna of patients with pediatric solid tumors. establishing a platform using cfdna to identify molecular profiles of these tumors can serve as a powerful tool for guiding treatment and monitoring response to treatment. background: despite multi-modality therapy, the prognosis for patients with metastatic osteosarcoma remains poor necessitating development of novel targeted therapies. immunotherapy can be exploited to target osteosarcoma with exquisite specificity but remains limited by insufficient tumor specific targets. objectives: to overcome the dearth in tumor specific antigens, we have explored the use of tumor derived mrna (representing a tumor specific transcriptome) for development of personalized nanoparticle vaccines. design/method: rna-nanoparticles (rna-nps) can be amplified from limited amounts of biopsied tissue for induction of tumor specific t cells against osteosarcoma. since local vaccination strategies are mired by poor overall immunogenicity, we assessed the feasibility, immunogenicity and antitumor activity of intravenously administered rna-nps (tumor mrna complexed to dotap nanoliposomes) in pre-clinical murine and canine tumor models. we identified a clinically translatable np formulation for the delivery of rna to antigen presenting cells (apcs) that induces in vivo gene expression and preserves rna stability over time. tumor derived rna-nps induced antigen specific t cell immunity and mediated anti-tumor efficacy in several pre-clinical solid tumor models (i.e. b f , kr b). when administered intravenously, rna-nps increased expression of co-stimulatory molecules (i.e. cd , cd , cd , ccr ) and pd-l on cd c+ cells throughout reticuloendothelial organs (i.e. spleen, liver, bone marrow) and within the tumor microenvironment; this phenotype was strictly dependent on type i interferon. targeted inhibition of type i interferon signaling (via infar mabs) abrogated anti-tumor efficacy mediated by rna-nps. we enhanced the immunogenicity of this platform by simply combining mrnas encoding for immunomodulatory molecules (i.e. hcv-pamps, gm-csf) or by combining rna-nps with immune checkpoint inhibitors. addition of checkpoint inhibitors (pd-l mabs) to rna-nps increased tumor infiltrating lymphocytes, and intratumoral mhc class i/ii expression, and mediated synergistic anti-tumor activity in settings where pd- or pd-l inhibition alone did not confer therapeutic benefit. we then explored the feasibility of rna-nps in a large animal osteosarcoma model. in ongoing studies for canines with osteosarcomas, we have shown that sufficient amounts of rna can be extracted, amplified, and manufactured into personalized rna-np vaccines. conclusion: rna-nps reprogram systemic immunity and mediate anti-tumor activity providing near immediate immune induction without the complexity of cellular immunotherapy. the immune correlate of preclinical response to rna-nps is hallmarked by interferon dependent pd-l expression on activated apcs (cd c+ mhcii+ cd + cells). based on these findings, we are exploring the preclinical safety, efficacy and immunologic effects of rna-nps targeting canine osteosarcoma before first in-human evaluation. background: ewing sarcoma is an aggressive bone tumor affecting mainly adolescent and young adults. treatments are based on compressed schedule chemotherapy combined with local control (surgery and/or radiation). prognosis is poorer for patients with metastatic disease, older age and central primaries. survival when disease recurs within two years of diagnosis is < %. the ews-fli fusion gene t( ; ) (q ; q ) has been well characterized as a dominant ews driver-gene. the most common variation is ews exon with fli exon ( % of fusion positive patients). we designed a novel pbi-shrna tm ews/fli type lpx which has demonstrated, safety and efficacy in animal model (rao et all). the pbi-shrna strategy silences target gene expression by concurrently inducing translational repression and p-body sequestration as well as post-transcriptional mrna cleavage. to determine the safety and maximum tolerated dose of intravenous administration of pbi-shrna tm ews/fli type lipoplex in patients advanced ews. design/method: phase i study × escalation cohort. testing pbi-shrna tm ews/fli type lpx (starting iv dose of . mg/kg) on patients (≥ age ) with advanced ewing's sarcoma, all with a type translocation. intravenous infusion was given twice a week for weeks with the following escalation schema: % → % → % → % → %. required kps > % and adequate organ function. cytokines induction pre and post-infusion was analyzed (il- , il- , tnf-alpha, il ra). first cohort of patients has been enrolled (ages between - years). three relapsed patients had > lines of therapy and patient had refractory disease, patients received a complete cycle of pbi-shrna tm ews/fli type lpx with twice a week infusions. a total of doses were given. the most prominent related toxicity has been hematological, patient developed transient g neutropenia, another patient developed g anemia that required prbc transfusion, and of note this patient had significant bone and bone marrow involvement. one patient only received two lpx infusions; she developed a fatal rsv pneumonia. other reported grade toxicity includes fatigue and headache. evaluable patients (n ) had stable disease between and months before progression. one patient had sustained response for month before progression, two patients are still alive. our preliminary experience supports the safety and potential efficacy of pbi-shrna tm ews/fli type lpx as novel treatment for advanced ews with limited toxicity. il- increase correlates with higher bi-shrnai ews/fli lpx infusion rate and clinical symptoms. further clinical testing is indicated. background: as more children with cns malignancies (bt) are surviving, the late effects of the therapies they receive are better described. studies show that radiation therapy is particularly harmful to neurocognitive functioning, specifically processing speed, working memory, and attention span. these deficits have negative effects on quality of life, especially in academic and professional settings. a large proportion of s of s adult survivors of bt are unable to reach adult milestones such as living on their own, holding a steady job, and getting married. proton beam radiation therapy (pbrt), is touted for the potential to have fewer and less severe side effects than traditional photon radiation therapy (xrt). because of the properties of protons, the amount of damaging energy released in non-target healthy tissue is reduced when compared to xrt. although a study comparing iq testing between pbrt and xrt found no difference between the two therapies, no studies have compared the specific neurocognitive domains. it would be valuable to evaluate full neurocognitive testing scores (nct) since the specific domains, particularly processing speed (psi), appear to be most vulnerable to radiation therapy. objectives: our primary aim was to assess differences in psi for patients with bt who underwent pbrt versus xrt. a secondary aim was to assess differences in iq (fsiq) and working memory (wmi). we retrospectively evaluated all patients treated for bt at the jimmy everest cancer center within the past years who received rt and had nct post radiation. we examined the full nct results for both subsets of participants to evaluate differences in the specific domains of processing speed, working memory, and iq by measuring percentiles scored in these domains. objectives: we report our experience on imaging children with mm treated uniformly on an institutional melanoma trial. we retrospectively reviewed the clinical and imaging findings of patients with ajcc stage iic-iv cutaneous mm treated on our institutional mel protocol. brain mri/ct, pet/ct, ct chest, abdomen, and pelvis (ctcap) were performed at diagnosis in all patients. on treatment, stratum a patients (peg-interferon; ajcc iic, iiia, iiib) (n = ) had the same imaging repeated every months; stratum b (peg-interferon and temozolomide; unresectable measurable disease metastatic, or recurrent) (n = ) had pet scans every months and brain imaging every months; those in stratum b (peg-interferon and temozolomide; unresectable non-measurable, metastatic, or recurrent) (n = ) had the same imaging performed every months. off therapy all patients continued same imaging every months for years. results: there were patients ( female; median age years). eleven had spitzoid and conventional melanoma. primary sites included head/neck (n = ), trunk (n = ), and extremities (n = ). patients with spitzoid melanoma had imaging studies ( pet, ctcap, ct chest, ct brain, and mri brain) with a median of , , , and studies/patient respectively. median cost per patient was $ , . thirteen studies ( . %) showed suspicious lesions with additional scans and diagnostic biopsies of which one only was positive stratum a with tert promoter mutation and died from disease). for conventional mm, studies ( pet, ctcap, ct chest, ct brain, and mri brain) were performed with a median of , . , , , studies/patient respectively. median cost per patient was $ , . twenty ( %) showed suspicious lesions with additional scans and diagnostic biopsies; four were positive (two at diagnosis); both died of disease; the other two recurred locoregionally and were detected clinically; both are alive and disease free; one patient had diffuse metastases and died shortly after enrollment. after a median follow up of . years (range . - . ) patients are alive and disease free. children with spitzoid melanoma should have minimal imaging at diagnosis and follow-up given the low risk of recurrence and low yield and high cost of aggressive imaging protocols. patients with conventional mm should be imaged according to the adult guidelines. nationwide children's hospital, columbus, ohio, united states background: the role of infections in the long term outcome of patients with bone tumors is controversial. two retrospective studies have shown increased survival in osteosarcoma patients who had a post-operative wound infection, while another showed no changes in overall survival. to determine the relationship between wound infections and/or bloodstream infection (bsi) on survival in pediatric and young adult patients with osteosarcoma and ewing sarcoma treated at a tertiary children's hospital. design/method: a retrospective chart review was performed for patients with diagnosis of osteosarcoma or ewing sarcoma from - . patients received standard chemotherapy regimens for their disease type and stage. local control included surgical resection and/or radiation therapy. presence of infection was determined by bsi or wound cultures while receiving treatment for primary tumor. the median age of patients was (range - years) at diagnosis. % had a diagnosis of osteosarcoma and % had ewing sarcoma. of these, % of patients developed an infection during treatment; % had bsi, % had wound infections, and % had both. patients with bsi had a year os of . %, compared to % in those without bsi (p = . ). those with both bsi and wound infections had the poorest overall survival of %, compared to . % for patients without any infection. patients with wound infections alone had a year os of . %, compared to % of patients without a wound infection. our analysis revealed decreased os in patients with bsi; however, this could be due to other confounding factors in the presence of bsi. those with bsi or bsi and wound infections had the poorest survival. wound infections without bsi were associated with a slight increase in survival; however, this study was limited by the number of patients that had local wound infections. with the use of newer surgical techniques, availability of antimicrobials and routine use of prophylactic antibiotics, the incidence of infections while undergoing treatment is low. however, the importance of this clinical observation indicates a likely enhanced immune system associated with infection, supporting the role of immunotherapy for treatment of these aggressive tumors. background: hypoalbuminemia is a well-recognized effect of cancer and other chronic illnesses and is often regarded as a marker of malnutrition. in adults, hypoalbuminemia has been associated with adverse outcomes in patients with cancers of the lung, pelvis, head and neck, gastrointestinal tract, and bone marrow, as well as in some pediatric patients with ewing sarcoma and hodgkin lymphoma. hypoalbuminemia has not been well studied in children with cancer. to determine the incidence of hypoalbuminemia (using age-specific references) in children with cancer receiving chemotherapy at baseline (prior to starting chemotherapy) and to determine whether hypoalbuminemia is associated with inferior -year overall survival. design/method: we performed a single institution, irbapproved, retrospective review of pediatric oncology patients diagnosed between and . five-year survival was estimated using the kaplan-meier method; groups were compared using cox regression. we identified pediatric patients with a first diagnosis of cancer, brain tumor, or other condition possibly requiring chemotherapy. of these patients, were excluded for reasons including not receiving chemotherapy and missing data, leaving patients who had a serum albumin level within days prior to starting chemotherapy. the mean age was . years (sd . years); % were male; % were non-hispanic. the most common diagnosis was acute lymphoblastic leukemia ( of ; %). one hundred thirty nine of ( %) had hypoalbuminemia prior to starting chemotherapy. there was no statistically significant difference in -year overall survival between those with and without hypoalbuminemia ( % vs. %, respectively; hazard ratio . , % c.i. . - . ). conclusion: hypoalbuminemia at baseline in pediatric oncology patients requiring chemotherapy is common (one in five), and was not associated with inferior -year overall survival in this cohort. leptomeningeal metastases at diagnosis. standard treatment for completely resected, non-anaplastic supratentorial ependymomas is close observation. treatment for anaplastic or incompletely resected non-anaplastic ependymomas is maximal safe surgical resection followed by focal radiation. however, up to % of localized ependymomas recur. the role of chemotherapy in treating ependymomas is under investigation. extraneural metastases of anaplastic ependymomas have rarely been reported and the outcome is dismal. objectives: to report extraneural cervical node metastases of a non-anaplastic ependymoma and successful treatment with surgical resection, radiation, and systemic chemotherapy. design/method: retrospective review of patient medical records, including radiographic imaging and tumor tissue pathology, and comprehensive literature review. results: a previously healthy -year-old girl underwent gross total resection (gtr) of an isolated right parietal lobe ependymoma (who grade ii). at age years, magnetic resonance imaging (mri) revealed an isolated localized recurrence. she underwent gtr followed by observation. at age years, she again experienced isolated localized recurrence and underwent gtr followed by . gy focal conformal photon radiation. at each recurrence, pathology revealed a non-anaplastic ependymoma, and cerebral spinal fluid (csf) cytopathology and spine mri were negative. at age years, she developed an enlarged right posterior cervical chain lymph node. subsequent mri revealed a large rim-enhancing, t hyperintense lymph node and multiple abnormally enhancing regional nodes consistent with metastases. biopsy revealed a non-anaplastic ependymoma. mri of the brain and spine, computed tomography of the chest, abdomen, and pelvis, and csf and marrow evaluations were unremarkable. chemotherapy according to acns was initiated. mri after course demonstrated significant node size reduction. she underwent right neck node dissection. only one right level ii lymph node showed metastases. she was treated with . gy irradiation to the neck and additional courses of chemotherapy. she remains in remission months and months after diagnosis of metastatic disease and end of therapy, respectively. literature review reveals rare reports of extraneural metastatic disease of anaplastic ependymomas to bone, lung, or liver, and only involving lymph nodes, all associated with a poor outcome despite multimodal therapy. to our knowledge, this is the first report of extraneural metastases of a non-anaplastic ependymoma. extraneural metastases should be considered in children previously treated for non-anaplastic ependymomas who experience systemic symptoms, even in absence of cns relapse. multimodal treatment offers potential long-term disease control with acceptable toxicity. arun gurunathan, joel sorger, andrew trout, joseph pressey, rajaram nagarajan, brian turpin cincinnati children's hospital medical center, cincinnati, ohio, united states background: pigmented villonodular synovitis (pvns) is a benign neoplasm of the synovium. standard treatment is surgery, but post-operative recurrence rate is as high as %. radiation therapy can be used for local control, but is associated with late effects. while pvns is rarely fatal, aggressive disease and/or extensive surgery can result in substantial functional impairment. colony stimulating factor- (csf ) overexpression, often due to chromosomal translocation involving csf , drives pvns through recruitment of synovial-like mononuclear cells expressing the csf -receptor. tyrosine kinase inhibitors such as imatinib are active against the csf -receptor, and have shown benefit in the post-surgical relapse setting. however, questions remain regarding the broader application of imatinib and regarding optimal response assessment. to present three patients with pvns, each with different clinical scenarios, who demonstrate clinical response to imatinib monitored by changes in metabolic activity (maximum suv) on pet/ct. results: three patients with pvns demonstrate pet/ct response to imatinib, guiding management of their challenging clinical scenarios. patient is a year-old female with left hip pvns and high grade articular cartilage loss, with decrease in metabolic activity (suvmax . to . in months) on neoadjuvant imatinib, enabling total hip replacement surgery planning. patient is a year-old female with left knee pvns with recurrences after synovectomies, spared subsequent surgical control attempts after clinical improvement correlating with pet/ct response to imatinib (suvmax . to . in months). patient is a year-old male with right knee pvns that recurred after total knee replacement, now with clinical improvement correlating with pet/ct response to imatinib (suvmax . to . in months). all patients would have been characterized as stable disease by response evaluation criteria in solid tumors (recist). in each of these patients, imatinib has been tolerated well, with no therapy interruptions and absent or easily managed side effects (one patient takes dronabinol for decreased appetite, one patient takes prn immodium for diarrhea). all patients are currently still taking imatinib, with therapy length ranging from five to eleven months. in our series of three patients with pvns, imatinib shows promise for disease management in neoadjuvant and adjuvant settings with a tolerable side effect profile. imatinib should be considered in the treatment of pvns to spare surgical and radiotherapy related morbidity, and treatment effect can be monitored by pet/ct. background: metastatic rhabdomyosarcoma (rms) carries a poor prognosis with three-year event free survival rates ranging between %- % (depending on oberlin risk factors) due to the lack of significantly effective breakthroughs in the recent past. there is an urgent and unmet need for new treatment strategies against this disease. metastatic rms cell lines exhibit increased expression of the erm family membrane-cytoskeleton linker protein ezrin. knockdown of ezrin expression using sirnas decreases the metastatic potential of these cells, whereas forced expression of ezrin results in increased degree of metastasis. the activity of ezrin is controlled by its phosphorylation at the threonine (thr ) residue at the c-terminus of the protein, suggesting that alteration of ezrin phosphorylation may control rms growth and metastasis. our goal was to determine if pharmacological inhibition of thr phosphorylation in ezrin affects the growth, survival and metastasis in rms in vitro as well as in vivo. design/method: rms cell lines representative of the alveolar and embryonal histological subtypes were used. rms cells were treated with a small molecule inhibitor of ezrin, nsc , which specifically dephosphorylates ezrin at the thr residue. baseline expression of ezrin and perm levels as well as the effect of nsc on perm levels in the rms cell lines was determined by western blotting of cell lysates. viability of cells was assessed by trypan blue exclusion, and morphology visualized by bright field microscopy. the extent of apoptosis was detected by imaging caspase / activation using fluorescent microscopy. motility of rms cells was examined by performing a wound-healing assay. subcutaneous and orthotopic xenografts were established in nsg mice using rd cells (embryonal rms). mice harbor-ing xenografts were treated with intraperitoneal injections of nsc or dmso. results: ezrin is constitutively phosphorylated at the thr residue in a majority of the rms cell lines examined. nsc dephosphorylates ezrin at the thr residue in these cell lines. treatment with nsc inhibits growth, induces apoptosis and inhibits the migration of rms cell lines in vitro. further, treatment of nsg mice bearing subcutaneous or orthotopic embryonal rhabdomyosarcoma xenografts with nsc significantly impedes tumor progression without any obvious adverse effects. our findings suggest that dephosphorylation of ezrin at the threonine residue may have the potential to be a novel therapeutic strategy for rms patients. all india institute of medical sciences, new delhi, new delhi, delhi, india background: the role of laparoscopy in the management of pediatric intra-abdominal solid tumors is yet to be established. the safety of laparoscopic management of pediatric intra-abdominal tumors is still questionable. we study the results of the initial case series of pediatric intraabdominal tumors managed laparoscopically at our institute from july onwards. design/method: total children ( -males, females) who presented to us with pediatric intra-abdominal tumors were included. the tumors included wilms tumor (n = ), neuroblastoma(n = ), adrenal cortical tumor(n = ), ovarian teratoma(n = ).children were between months - years and received neo-adjuvant chemotherapy. a -port laparoscopic nephrectomy and lymph node sampling for wilms tumor and adrenalectomy for adrenal tumors was performed. the tumors were removed in-toto with no rupture (except in one). specimens were retrieved through a lumbar incision (n = ) or an inguinal incision(n = ). all the children are under regular follow up. two children with wilms tumor had recurrence. the neuroblastoma child underwent open surgery for recurrence later. conclusion: laparoscopy/laparoscopic assisted removal of pediatric intra abdominal tumor is a feasible and safe option. it has the advantage of less postoperative pain, shorter hospital stay and a better cosmetic result. proper patient selection, port placement and laparoscopic experience are contributory. background: targeting of proteins and cell surface antigens specific to cancer cells with monoclonal antibodies has proven to be an effective form of treatment in many forms of cancer. gd is a cell surface disialoganglioside that is expressed on the cell surface of some normal tissues including nerve cells, melanocytes, and mesenchymal stromal cells and is overexpressed in some pediatric cancers like neuroblastoma and osteosarcoma. dinutuiximab is a chimeric monoclonal antibody that is fda approved for the treatment of patients with high risk neuroblastoma and under investigation for the treatment of relapsed osteosarcoma. little is known about the patterns of gd expression in other pediatric malignancies. objectives: we sought to describe the patterns of gd expression in the following pediatric sarcomas: synovial sarcoma, rhabdomyosarcoma and ewing sarcoma. design/method: synovial sarcoma (n = ), rhabdomyosarcoma (n = ) and ewing's sarcomas (n = ) formalin fixed, paraffin embedded cores were obtained from the seattle children's research institute tissue microarray (tma) biorepository. tma blocks consisting of melanoma cores stained with and without gd antibody were used as positive and negative controls, respectively. slides were incubated with anti-ganglioside gd antibody clone q (ab from abcam) diluted : in % normal goat serum and % bsa in tbs overnight at ˚c. the negative control of human melanoma section was incubated in % normal goat serum and % bsa in tbs without primary antibody. the expression of gd was indicated by characteristic brown diaminobenzidine staining. the intensity and location of tissue staining were assessed and compared to positive and negative controls. staining was considered positive (+++) if the intensity of the staining was consistent with that of the positive control with - % of cells staining positive. classification of intermediate gd expression (++) was assigned to slides in which - % of cells stained positive. slides were classified as sporadic staining (+) if - % of cells stained positive. tissue was considered (-) if there was complete absence of staining, similar to the negative control. objectives: to evaluate the clinical presentation, management and treatment outcomes of children with malignant germ cell tumor at our institute design/method: a prospective study was conducted from june to dec in the department of pediatric surgery in a tertiary care institute in a developing country. all patients were evaluated for local disease and metastatic disease by imaging and tumor markers. risk stratified chemotherapy was used with low risk tumor receiving no chemotherapy, intermediate risk: courses of peb chemotherapy and high risk: courses of peb + courses of pe. upfront resection of the primary or the residual disease after neoadjuvant chemotherapy if feasible was performed. follow up was done with monthly tumor markers for months and imaging studies every - months for initial years. five year overall survival and disease free survival was calculated. results: during the study we treated children who formed the study group. of these ( %) were gonadal ( ; % testicular and ; % ovarian) and the remaining ( %) were extragonadal with sacrococcygeal (sct) being the most common site ( %). one hundred and thirteen children ( %) presented to us primarily while the remaining had received treatment elsewhere. stage or stage disease at presentation was present in ( %) children. recurrence was noted in ( %) patients. respectively. patients with testicular mgct and children with age - years and males had significantly poor rfs rates. conclusion: patients with mgct should be staged correctly and adjuvant chemotherapy is advisable to all patients except stage i endermal sinus tumor of testis. awareness regarding the same is still lacking in our country. meticulous follow up is needed as more than % of will recur. cure rates are dismal in children with recurrent mgct especially those who are not chemotherapy naïve. nemours children's specialty care, jacksonville, florida, united states background: radiotherapy for pediatric head and neck tumors often results in mucositis, limiting oral intake and compromising patients' nutritional status. this may be reduced through the improved conformality offered by proton therapy. despite widespread use of enteral tube feeding through a percutaneous gastrostomy (peg) or nasogastric tube (ngt), there is little data available regarding overall incidence of ngt/peg placement and perspectives of pediatric patients and caregivers. objectives: to (a) estimate the need for ngt/peg support and (b) characterize patient and caregiver perceptions surrounding enteral feeding in children with head and neck tumors undergoing proton therapy. design/method: dependent on development stage, patient (n = ) or parents (n = ) filled out a series of customized surveys according to a prospective irb approved study. seventythree percent of patients also received concurrent chemotherapy. questions addressed their current feeding route and perception, for example, "what aspect(s) of tube feedings are beneficial to you?" and "what aspect(s) of tube feeding worry or scare you?" fifty-five surveys were distributed before and after radiation, and with any change in feeding route. results: at the start of proton therapy, patient had a ngt and patients had peg. of these, patients ( %) had a ngt/peg in place exclusively for the administration of medication; only patient ( %) needed a ngt/peg for nutrition. in those patients without ngt/peg, % would "consider" enteral feeds. in patients without ngt/peg, the most commonly cited benefit was "maximizing my nutrition" ( %) and the most common negative aspect was "fear" of tube placement ( % of patients). all sub-populations ( % of patients) cited change in appearance as a negative aspect. in patients without ngt/peg at the start of proton therapy, % of patients/caregivers felt enteral feeding to be "unnecessary," and % of these patients would not "consider" ngt/peg even if their "physician advised it." over the course of proton therapy, the patients/caregivers who deemed enteral feeding "unnecessary" decreased from % to %. at completion of treatment, patients ( %) were using a ngt/peg tube for nutritional support but only one ( %) patient relied exclusively on their enteral feeds. two patients (without ngt/peg) ( %) required parenteral support. our data does not support prophylactic placement of ngt/peg in of children with head and neck tumors undergoing proton therapy. ongoing research is needed to identify which patients will need ngt or peg to supplement their diet. in this cohort, anticipatory counseling should focus on pain, cosmesis, and utility. children's national medical center, washington, district of columbia, united states background: ovarian sex cord-stromal tumors (osct) are rare neoplasms that typically present with signs/symptoms of an adnexal mass and signs of hormonal production approximately % of ovarian sex cord-stromal tumors in children are sertoli-leydig cell tumors (slct) with median age of presentation years overall. to our knowledge the youngest reported case in the literature describes a -month old female in china with a slct that was treated with oophorectomy alone. some studies have found an association in families between pleuopulmonary blastoma and osct with a germline mutation leading to dicer syndrome, which has been associated with a younger age at diagnosis. , objectives: to describe an unusual case presentation of slct in an infant results: -month old, twin female, ex- week premature infant presented to the emergency department on multiple occasions for abdominal distention and feeding intolerance initially thought to be related to previous omphalocele repair and umbilical hernia. an ultrasound demonstrated an × cm mass arising from the right ovary with large volume ascites. she required admission to the intensive care unit due to s of s respiratory distress from her significant ascites. serum tumor marker including hcg, afp and ldh were negative. patient underwent right oophorectomy with tumor capsule noted to be open at time of surgery. further imaging post operatively demonstrated no other sites of disease. the patient was classified as figo stage ic due to the presence of her significant abdominal ascites that was presumed to be malignant pre-operative tumor rupture. the pathological diagnosis was challenging and eventually resulted as a mixed germ cell sex cord stromal tumor with pattern of sertoli cell tumor with neuroendocrine differentiation. based on the staging of figo ic with pre-operative rupture, the decision was made to treat with a standard platinum based regimen as there is a higher incidence of relapse in stage ic patients when compared to ia treated with observation alone. our patient tolerated four cycles of chemotherapy well and end of therapy scans showed no evidence of disease. interestingly, her dicer mutation genetics performed by ion torrent tm next generation sequencing was negative in germline and tumor studies. to our knowledge, our patient is the youngest described with slct. she will continue to be followed with serial imaging alone as she had no evidence of elevated tumor markers at diagnosis. , due to young age and unusual diagnosis, she was referred to cancer genetics team. background: approximately % of patients with wilms tumor (wt) have metastatic disease at diagnosis and often have a grave prognosis. limited cell lines are available for the study of metastatic wt and long-term passaged cell lines do not always recapitulate the human condition. focal adhesion kinase (fak) is a non-receptor tyrosine kinase that controls cellular pathways involved in the tumorigenesis of pediatric renal tumors. using a novel patient-derived xenograft (pdx) model from a patient's primary wt (coa ) and matched isogenic metastatic wt (coa ), we previously demonstrated that fak is expressed and its inhibition led to decreased tumorigenicity of both the primary and metastatic pdxs. kinomic profiling is an innovative, high-throughput method used to investigate kinase signaling to identify potential therapeutic targets. to date, the kinomic profile of primary and metastatic wt has not been examined. objectives: investigate baseline kinomic differences between primary and metastatic wt and evaluate kinases upstream and downstream of fak as potential targetable therapies. design/method: cells from coa and coa were treated with pf- , (pf), a small molecule fak inhibitor. protein from cell lysates of treated and untreated coa and coa were combined with kinase buffer, atp, and fluorescently labeled antibodies and loaded into a phosphotyrosine kinase or serine-threonine kinase pam-chip® per the uab kinome core protocol. phosphopeptide substrate analysis with the pamstation® kinomics workstation (pamgene® international), pamchip® protocol using evolve software, and bionavigator v. . were used to analyze kinases upstream and downstream of fak. the primary wt had increased epha , ror sgk and decreased pdgfrb relative to the paired metastatic wt at baseline. treatment with pf increased ron, pdgfrb, p s kb, mak, camk g, vacamkl, camk d, ck a and pskh in the primary wt. treatment with pf decreased tnk , lmr , cck , epha , pdk , sgk , lkb and increased pskh in the paired metastatic wt. primary wt displayed a different kinomic profile compared to metastatic wt in a matched isogenic pdx model. these data reveal that alternative therapies to specifically target metastases are needed. furthermore, fak inhibition resulted in diverse kinomic alterations between primary and metastatic wt. inhibitors targeting many of these pathways, such as pdgfrb inhibitors, are currently available and potentially could be combined with fak inhibitors in the treatment of wt. the results of the current study indicate that kinases upstream and downstream of fak in primary and metastatic wt warrant further investigation. background: use of high-dose methotrexate (hd-mtx, g/m^ ) is a mainstay of standard therapy for pediatric osteosarcoma (os) in north america. in pediatric os, there is a narrow therapeutic window for hd-mtx, with decreased tumor response rate with mtx concentrations < m and decreased survival due to severe toxicity with concentrations > m. risk factors for hd-mtx toxicity have been defined in adults, including body mass index (bmi) and male gender, but such studies have not been conducted in children. we sought to examine the relationship between mtx levels and toxicities during hd-mtx infusion for pedi-atric os, thereby identifying risk factors for increased toxicity and providing a framework for therapeutic drug monitoring. design/method: this retrospective chart review included patients treated at texas children's hospital with hd-mtx as first-line therapy for os from - . data abstracted from electronic records included patient characteristics, bmi and body surface area (bsa), baseline and post-treatment laboratory values, mtx levels and hours after dose given ( h, h), hour mtx cleared (mtx < . um), grade / mucositis, myleosuppression, persistent lft elevation (ctace v . ), and % tumor necrosis. correlation between h mtx level and other covariates was summarized using descriptive statistics. we reviewed hd-mtx infusions corresponding to patients. bmi was found to significantly impact h mtx level (p< . ). female gender was also significantly associated with higher h mtx level (p< . ). percent necrosis (available in patients) was associated with h mtx levels at near-statistical significance (p = . ). h mtx level was not found to contribute to toxicities or associate significantly with mtx clearance. analysis in a larger cohort is ongoing. we have identified at least one patient factor (bmi) that significantly impacts h mtx levels and is of potential use for future modeling, as current models incorporate bsa only. our findings concord with studies in adult os in that bmi significantly impacts h mtx level but diverge in that female gender is associated with higher h levels. importantly, these data support targeting h mtx levels to ensure that minimum concentration for adequate tumor necrosis is reached. these results do not suggest that monitoring h levels would prevent toxicities, thus necessitating further characterization of any intrinsic patient factors that associate with toxicity. overall, our definition of the clinical factors that associate with h mtx levels contributes to a framework for therapeutic drug monitoring in pediatric os. children 's mercy hospital kansas city, kansas city, missouri, united states background: post consolidation immunotherapy with dinutuximab, aldesleukin (il- ), granulocyte macrophage colony stimulating factor (gmcsf) and isotretinoin is standard of care for children with high risk neuroblastoma. dinutuximab is combined in alternating cycles with s of s gmcsf or il , followed by a th cycle with isotretinoin alone. il- is administered as a hour continuous infusion on days - at miu/m /day followed by a higher infusion dose, . miu/m /day, in combination with dinutuximab on days - of cycles and . the miu/m /day dose may be administered inpatient or in the ambulatory setting. objectives: to retrospectively compare the incidence of inpatient and outpatient side effects and complications associated with low dose ( miu) il to provide the tolerability data necessary to evaluate these venues for future administration options. design/method: this study was a descriptive, singlecentered definitive study utilizing a retrospective convenience sample population of children with high risk neuroblastoma who received low dose il either as an inpatient or an outpatient without exclusion from may to june . subjects were identified by a tumor registry query post irb approval. electronic and paper medical records were reviewed for the dates and location of the infusions, the home health company used if applicable and all documentation regarding clinical status, side effects and toxicity. demographics was limited to age and gender. results: infusion venue was chosen by provider preference. twenty-six infusions, inpatient and outpatient via separate home health companies were all administered in entirety and without interruption. there were males and females ranging from - years of age. two children received a single outpatient infusion due to intolerance of il when combined with dinutuximab and received therapy in both settings. fever, inpatient and outpatient was the only common side effect. no source of infection was ever identified. there was one incidence of diarrhea and one patient with pruritus in both the outpatient and inpatient settings respectively. no planned outpatient infusions required subsequent admission however the outpatient fever did necessitate an er evaluation. conclusion: low dose il can successfully be administered outpatient. the medication has minimal side effects with fever occurring in %, none of which were associated with infection. no outpatient infusion required a subsequent admission. no patients who received cycle infusions outpatient opted to receive the next cycle inpatient. baylor college of medicine, houston, texas, united states background: metastatic ewing sarcoma (es) has an extremely poor overall survival, necessitating investigations into molecular mechanisms to identify novel targets and develop new therapies. we previously performed an in vivo study, using our mouse model, designed to provide insights into transcriptomic and proteomic signatures for metastatic es to identify potential therapeutic targets. comparing profiles of primary tumors to corresponding metastatic lesions, we identified aberrant expression of integrin ß (itgb ) and downstream activation of integrin-linked kinase (ilk) in metastatic lesions compared to primary tumors, implicating this pathway as a key regulator in the ability of es to establish and enhance metastasis. our hypothesis is that upregulation of itgb and its downstream signaling events play a key role in es metastasis and are viable therapeutic targets. objectives: to investigate the role of itgb and its downstream signaling pathways in driving the establishment and enhancement of metastasis in es and to investigate this pathway as a potential therapeutic target. to investigate the role of itgb and ilk in es metastasis, we used sirna to knock down itgb and ilk expression in established es cell lines and then performed functional assays in vitro, including cell proliferation and invasion/migration assays. we also tested inhibition of this itgb signaling pathway using available small molecule inhibitors targeting itgb , ilk and the downstream target ap- , using cilengitide, compound and sr , respectively. we are currently using these small molecule inhibitors as treatment in vivo and assessing rates of metastatic tumor formation. we generated stable itgb and ilk overexpression and knockdown cell lines, which we are using for similar in vitro and in vivo investigations. knockdown of itgb and ilk in our sirna cell lines resulted in decreased cell proliferation and decreased invasion and migration compared to controls. we also found significantly decreased cell proliferation using each of the small molecule inhibitors in vitro. our preliminary studies using compound in vivo established a safety profile and dose escalation is underway to assess the effectiveness of inhibiting es metastasis. these results support our hypothesis that itgb and its downstream signaling events play a key role in the ability of es to establish metastatic foci and may serve as a potential therapeutic target. we continue to investigate this pathway in vitro. we are also using our small molecule inhibitors and itgb and ilk overexpression and knockdown approaches to study these effects on metastatic tumor development in vivo using our mouse model. background: neuroblastoma (nbl) is characterized by phenotypic heterogeneity. outcome is excellent for patients with low-(lr) and intermediate-risk (ir) disease, whereas only % of high-risk (hr) patients will survive. -hydroxymethylcytosine ( hmc) is an epigenetic marker of active gene transcription, and hmc profiles are prognostic in many types of adult cancers. we hypothesized that hmc profiles will serve as robust biomarkers in children with nbl tumors, refining current risk stratification. objectives: analyze genome-wide hmc in nbl tumors and correlate hmc deposition with chromosomal copy number and gene expression. design/method: hmc was quantified by nano-hmc-seal-seq from the dna extracted from hr, ir and lr nbl tumors. read counts and clinical data were analyzed with deseq to identify genes with differential hmc patterns between risk groups. chromosomal copy number was assessed by chromosomal microarray analysis (cma) in a subset of samples ( lr and hr). expression of genes located on chromosome p was evaluated using publically available microarrays (e-mtab- ) of hr nbl tumors with known p loh status. results: globally, lr tumors had more hmc peaks ( , ) than ir ( , , p = . ) tumors, or hr tumors ( , , p = . ). , genes had different patterns of hmc deposition in hr versus lr tumors. ( %) of these genes mapped to chromosome p and had decreased hmc in hr versus lr tumors (padj < . ). in the cma analysis p deletion was detected in of the tumors tested. in the tumors with p loss, genes that map to p showed decreased hmc deposition compared to the hr tumors without p loss (p< . ). further, compared to the tumors without p loss, the expression of of the p genes was decreased (p< × - ), including chd , camta , and arid a, known and proposed tumor suppressor genes in nbl. conclusion: different patterns of hmc accumulation are associated with neuroblastoma risk classification. nano-hmc-seal-seq is sensitive to copy number variations and has the potential to identify these changes in patient tumors. our results suggest that hmc deposition contributes to the silencing of tumor suppressor genes in p and may also regulate the transcription of other genes that drive tumor phenotype. background: metastatic osteosarcoma has a -year survival rate of - %. pulmonary metastases remain a major treatment challenge in osteosarcoma. current treatment with conventional chemotherapy shows inadequate activity towards metastases and has toxic systemic side effects. chloroquine is a widely used anti-malarial drug and has been shown to have promising anti-cancer and anti-metastatic activity. polymeric drugs have been shown to have multiple advantages over their small molecular parent drugs, including enhancing the therapeutic efficacy, an improved pharmacokinetics profile and decreased systemic toxicity. we hypothesized that by developing chloroquine into a polymeric drug and combining it with conventional chemotherapy it will improve the treatment of metastatic osteosarcoma. objectives: to identify the optimal combination of polymeric chloroquine (pcq) with conventional chemotherapy active in osteosarcoma as a new means of treating metastatic disease in a murine osteosarcoma model. we synthesized and developed pcq and evaluated its anti-invasive activity using an osteosarcoma cell migration and invasion assay. we evaluated the efficacy of cell killing using combination drug therapies with pcq and a panel of conventional chemotherapy agents (doxorubicin, docetaxel, cisplatin and paclitaxel) using celltiter blue cell viability assay. to develop the murine osteosarcoma model, we intravenously injected luciferase-expressing human osteosarcoma cells b into nsg mice. we administered the drug combination that showed the strongest in vitro synergy to the mice and evaluated their anti-cancer and anti-metastatic effects in vivo. tumor growth and suppression were evaluated using whole body bioluminescence imaging. results: we successfully synthesized pcq that contains . % chloroquine with a molecular weight of . kd. pcq was also found to decrease the toxicity of the parent chloroquine. pcq showed strong inhibition of osteosarcoma cell migration with % inhibition compared to % by chloroquine. we screened the combination drug therapies and found the combination of pcq and doxorubicin to show the strongest synergism. the pcq/doxorubicin combination is currently being evaluated in the murine model. combination drug therapy using pcq and doxorubicin showed synergistic cell killing and inhibition of cell migration in vitro. the combination represents a promising treatment strategy for pulmonary metastatic osteosarcoma. emory university/children's healthcare of atlanta, atlanta, georgia, united states background: survival for relapsed high-risk neuroblastoma (rnb) is < %, underscoring the critical need for novel therapies. rnbs have increased ras/raf/mapk mutations and increased yes-associated protein (yap) transcriptional activity. yap is a transcriptional co-activator that binds with tea-domain (tead) transcription factors to regulate cellular proliferation, self-renewal, and survival. we found that shrna inhibition of yap decreases nb cell proliferation and sensitizes ras-mutated nbs to mek inhibitors, supporting yap as a tractable therapeutic target. verteporfin (vp), a photodynamic drug used for macular degeneration, is the only drug found to inhibit yap expression or yap:tead binding to kill tumor-derived cells. peptide is a mer yap peptidomimetic that also disrupts yap:tead interactions. we sought to determine whether these compounds are potent in nb via yap direct effects. design/method: yap expressing (nlf, sk-n-as) or yap null (ngp, lan , sk-n-as-shyap) human-derived nbs were incubated with vp, with and without direct light exposure, or with peptide . celltiter-glo and immunoblots were used to assess for cell death and yap-downstream protein expression, respectively. results: without direct light exposure, vp inhibits yap expression at nm dosing, yet no nb cell death was observed at equal or higher concentrations. egfr and erk / were inhibited along with yap, confirming yap/ras pathway coregulation. when vp was exposed to direct incandescent light for minutes, > % nb cell death occurred in all nbs tested, even those lacking yap. peptide caused no cell death or yap inhibition up to um. neuroblastomas are resistant to vp at doses sufficient to inhibit yap expression. in macular degeneration, light-activated vp produces reactive oxygen species, which we hypothesize is the off target mechanism killing nbs independent of yap. given the off target effects and the need for light activation, vp is not an ideal preclinical or clinical yap inhibitor. accordingly, peptide has poor cell permeability and low tead affinity, leading to its lack of efficacy. given the relevance of yap in rnb and other cancers, we are chemically optimizing a yap peptidomimetic with enhanced permeability, nuclear localization, and tead affinity to create a bonafide yap inhibitor for preclinical and clinical application. kayeleigh higgerson, aaron sugalski, rajiv rajani, josefine heim-hall, jaclyn hung, anne-marie langevin ut health san antonio, san antonio, texas, united states background: osteosarcoma is the most common bone malignancy in children, adolescents, and young adults. most study cohorts have to % hispanic patients that encompass many different hispanic backgrounds. the university of texas health science center at san antonio (uthscsa) sarcoma team serves a latino population that is predominantly mexican american, thus providing a unique opportunity for evaluation this population. this study expands on previous data collected from january to december from the same institution, providing increased insight into outcomes of mexican american children, adolescents, and young adults with osteosarcoma. objectives: to further understanding of osteosarcoma in latino children, adolescents and young adults. design/method: a retrospective analysis of demographics, tumor characteristics, response to treatment, and survival outcome of all localized osteosarcoma of the extremity patients below years of age diagnosed and treated by the uthscsa sarcoma team between january and june was performed. results: in our original cohort from january to december , we observed a significantly decreased -year eventfree survival (efs) in patients diagnosed before age (preadolescent) relative to patients diagnosed between ages and ( % vs. %, p< . ). patients had a -year overall survival (os) and event-free survival of % and % respectively. in our expanded cohort from january to june we evaluated sixty-six patients with a median age of (range, to y) with localized high-grade osteosarcoma of the extremity. the expanded cohort was % mexican american, with a median follow-up of months (range, to ). the analysis of our expanded cohort is ongoing and we postulate that the findings will hold true, as we increase the cohort size and length of follow-up. conclusion: analysis of our previous cohort, predominantly of mexican american ethnicity, showed that preadolescent patients had an increased rate of relapse when compared with previous large studies. we also showed a trend towards decreased efs for the entire cohort. we hypothesize that we will further validate these findings with this expanded cohort and this will support further investigation into potential causes of poor outcome in this vulnerable latino population. background: neuroblastoma in infants has the potential to regress or mature spontaneously. growing literature showed that some cases subjected to initial observation didn't show inferior outcome compared to actively treated similar categories. objectives: we investigated whether early active treatment can be safely avoided/deferred in selected favorable cases at the children's cancer hospital-egypt (cche). design/method: patients enrolled on the watch and see strategy (w&s) at cche had small primary tumor; inss stage - , uncomplicated stage s or stage infants (< days). tissue biopsy was not mandatory for infants below months of age with localized adrenal mass (stage - ). on progression, immediate intervention took place according to stage and risk of disease after biological characterization. results: thirty four nbl patients were enrolled on w&s strategy; m/f: . / . eighteen patients had stage s disease, patients had stage - and were stage . primary adrenal site was reported in patients ( . %), patients ( . %) had small mass measuring ≤ cm in its largest diameter. the -year os & efs were . ± . % and . ± %, respectively, with months median follow-up (range: - months). spontaneous total/near total resolution of mass occurred in / patients ( %). median time to eliciting regression was . months (range: . - . months), and . months (range: - months) till complete resolution. only / patients ( . %) witnessed progression ( local, distant and combined local and distant progression); median time to progression was months (range: - months) with / deaths after starting chemotherapy. watch and see strategy is a safe approach in localized and uncomplicated stage s neuroblastoma. progressive cases could be rescued. baylor college of medicine, houston, texas, united states background: ga- dotatate binds to somatostatin receptor expressed in neuroendocrine tumors (nets). it was approved by fda in for use with pet/ct scan for localization of somatostatin receptor positive nets in adult and pediatric patients. pediatric approval was based mainly on extrapolation of data from adults. objectives: to describe the use of ga- dotatate pet/ct scan in children with neuroendocrine tumors and compare with other imaging modalities. design/method: patients with nets enrolled in texas children's rare tumor registry between february and october were reviewed and those patients who underwent ga- scan were included. results: four patients with nets underwent ga- scans without any adverse reactions. first patient was a -yearold female with small bowel net with multiple liver metastases. mri abdomen and fdg pet at diagnosis showed s of s multiple liver metastases but could not identify the primary lesion. ga- scan was able to accurately identify the enlarged lymph nodes in the small bowel and was better than fdg pet in delineating the liver metastases. second patient was a -year-old female with recurrent small bowel net with liver, lung and paraspinal metastases. the lesions were initially detected by ct scan. octreotide scan failed to show any uptake in the identified lesions while ga- was taken up by the liver lesions, lung lesions > cm in size and the paraspinal lesion. third patient is an year-old male with pancreatic net with peripancreatic lymphadenopathy, multiple liver metastases and cardiophrenic lymph node involvement. the primary lesion in the pancreas could not be identified by ct scan, ct angiogram, mibg scan, or octreotide scan. in addition, there was uncertainty about involvement of the enlarged cardiophrenic lymph node. in addition to clearly identifying the primary lesion, ga- scan was able to detect multiple peripancreatic lymph nodes not detected by other scans and revealed uptake in the cardiophrenic lymph node confirming its involvement by the tumor. fourth patient is a -year-old female with malignant abdominal paraganglioma with solitary lung metastasis. both mibg scan and ga- scan were able to identify the primary lesion. ga- scan was performed after the lung metastasis was removed and thus its ability to detect it could not be confirmed. background: neuroblastoma is the most common extracranial solid tumor of childhood, with overall survival for high-risk patients (hrnbl) near %. the outcomes of hrnbl have improved with high dose chemotherapy followed by autologous stem cell rescue (abmt). data about factors influencing the rate of hematopoietic recovery following abmt in hrnbl is lacking in the literature. our objective was to identify factors influencing the rate of hematopoietic recovery following abmt in hrnbl. design/method: this was a retrospective chart review of patients with hrnbl treated at texas children's hospital from to . neutrophil engraftment was considered the first of three consecutive days with post-transplant neutrophil count greater than cells/ul. red blood cell and platelet engraftment were considered at a hemoglobin greater than g/dl and platelets greater than , /ul three days after the last transfusion. race and conditioning regimen were analyzed using one-way anova; amount of infused cells was analyzed using pearson correlation coefficients; chemotherapy delay and bone marrow (bm) involvement after cycle of induction chemotherapy were analyzed using independent sample t-tests. the study included males and females with a median age at diagnosis of . years. thirtyeight patients were caucasian, african-american, hispanic, asian, and did not have race documented. the mean dose of infused cd + cells was . × ^ cells/kg. forty-five patients received conditioning therapy with carboplatin/etoposide/melphalan (cem), received busulfan/melphalan (bu/mel), and received thiotepa/cyclophosphamide (thiotepa/cpm). the conditioning regimen administered was significant (p = . ) for time to engraftment of neutrophils, with bu/mel at . days, cem at . days, and thiotepa/cpm at days. a delay of chemotherapy during induction (n = ) was significant (p = . ) for time to platelet engraftment of greater than , /ul and trended towards significance (p = . ) for time to neutrophil engraftment. bm involvement at diagnosis and after cycle of induction was not significant for time to engraftment. dose of stem cells infused was the only variable significant for hemoglobin engraftment. background: osteosarcoma (os) is the most prevalent aggressive primary malignancy of the bone affecting children and young adults. approximately % to % of patients have metastatic disease at initial presentation, and % of those patients have isolated pulmonary metastases. although overall survival in patients with os has improved with advances in therapy, there have been no significant improvements in survival outcome in patients with metastatic disease. recent studies suggest that tumor-associated vascular cell adhesion molecule (tvcam- or cd ) plays a critical role in the metastatic progression of various tumors. indirect evidence from these studies suggest that vcam- / integrin signaling promotes tumor survival and metastatic progression by changing the tumor niche and associated immune response. to determine if interfering vcam- / signaling between pulmonary metastatic osteosarcoma (pos) and macrophages (macs) by down-regulating vcam- , depleting macs or blocking vcam- / signaling will reduce pos and improve overall disease-free survival. design/method: we used a pair of spontaneous, high-grade murine os cell lines from balb/c mouse (h- d), k and k m (derived from in vivo k metastasis). we used lentiviral shrnas to knockdown vcam- mrna and protein expression in k m (vcam- kd). we introduced luciferase into k , k m and various k m shrna cell lines to follow lung metastasis by bioluminescence (bli). we depleted macs by intranasal administration of liposomal clodronate formulation. we tested the ability of k and k m supernatants to polarize m macs into m or m phenotype in vitro. we also administered anti- monoclonal antibody (anti- mab) intranasally to assess the outcome of functional blockade of vcam- / signaling. results: k m over-expressed vcam- compared to k . mac depletion in k m -bearing animals exhibited reduced pos. weekly administration of anti- mab resulted in % tumor-free rescue among mice with established k m pos. interestingly, supernatant from k m but not k preferentially induced m -like macs, suggesting a novel integrin-mediated mechanism of m differentiation. validation data with additional os cell lines will be presented. despite aggressive multimodal therapy, overall outcome for patients with pos remains dismal at - %. for this reason, novel and directed therapy approaches are desperately needed. molecular targeted approaches for therapy are challenging, due to the complex genetic heterogeneity of os. immune-modifying therapy is a promising new alternative approach for pos. university of chicago, chicago, illinois, united states background: only half of all patients diagnosed with high-risk neuroblastoma achieve long-term survival. imetaiodobenzylguanidine (mibg) scans are routinely used to evaluate disease at diagnosis and following treatment, and the extent of disease is quantified using the curie scoring system. a previous study by yanik et al., has shown that for high-risk patients with mycn non-ampliified tumors, scores less than versus greater than following cycles chemotherapy are associated of superior survival, whereas scores less than versus greater than were prognostic in patients with mycn-amplified tumors. however, the prognostic significance of specific sites of metastatic disease at diagnosis is not known. to determine if site of metastatic disease determined by i-metaiodobenzylguanidine (mibg) imaging in high-risk patients at the time of diagnosis was associated with outcome design/method: we performed a retrospective chart review of high-risk neuroblastoma patients treated at comer children's hospital and lurie children's hospital in chicago between and with positive mibg scans at the time of diagnosis. we collected imaging data as well as other clinical data including bone marrow status. sites of disease were defined as curie regions with any positive value. kaplan-meier analysis was performed to evaluate the association with disease sites and survival. pearson correlation coefficients were calculated to compare bone marrow disease to sites of positivity on mibg scan. the cohort consisted of high-risk patients. had skull disease, and had pelvic disease. the presence of mibg positive disease in the skull and in the pelvis trended toward worse efs. efs at years for patients with disease in the skull at diagnosis was ± % and for patients without skull disease was ± % (p = . ). efs at years for patients with and without pelvic disease was ± % and ± % (p = . ). consistent with prior data, we found that the presence of bone marrow disease was associated with worse survival with year efs of ± % and ± % with and without marrow disease at diagnosis (p = . ). there is the highest correlation between pelvic disease on mibg scan and bone marrow disease with pearson coefficient . . pelvic disease noted on mibg scan likely reflects underlying bone marrow disease. in patients with high-risk neuroblastoma, skull disease and pelvic disease on mibg scan at diagnosis may predict worse event free survival. background: osteosarcoma is one of the deadliest cancers in the pediatric population with little progress in morbidity and recurrence rates since the 's. oncolytic herpes simplex- virus (ohsv) is an attenuated virus that has shown encouraging results against certain solid tumors. programmed cell death protein (pd)- -mediated t cell suppression via engagement of its ligand, pd-l , is also of particular interest due to recent successes in selected cancers, especially those with high genetic mutational loads. most pediatric cancers do not have a wide variety of mutations; however, osteosarcoma has a chaotic genome, prone to genetic mutations. it has been shown through numerous other studies that pd- inhibition alone is not sufficient to result in statistically significant tumor growth delays in osteosarcoma models and patients. we hypothesize the addition of ohsv therapy as an immunologic stimulus to pd- inhibition is efficacious for osteosarcoma. ( ) to determine whether ohsv therapy enhances response to pd- inhibition in immunocompetent murine models of osteosarcoma and ( ) to quantify and characterize the anti-tumor t-cells infiltration after treatment with ohsv and pd- inhibition individually and in combination. we utilized an immunocompetent transplantable murine model using a cell line derived from a spontaneous metastatic osteosarcoma (k m , balb/c background). we transplanted established tumor wedges subcutaneously and monitored tumor volume by caliper measurement. once tumors reached - mm , we administered intratumoral injections of hsv ( × plaque-forming units) every other day for a total of injections. we then gave intraperitoneal injections of ug anti-pd- or control antibody twice weekly, up to weeks, starting from the last dose of virus treatment. we monitored tumor growth via calipers twice weekly until tumors reached mm or cm diameter. we quantified and characterized innate and adaptive immune cell infiltrates in tumors using flow analysis. we found significantly prolonged survival with our combination therapy group compared to all other groups. we found that anti-pd- by itself had little impact on t cell recruitment while the combination group had higher influx of cd + cells with a reduced amount of t-regulatory cells (cd +foxp +cd +). we also found an increase in cd + effector memory cells. osteosarcoma is a deadly cancer with therapeutics remaining unchanged for the last years. here, we describe prolonged murine survival after treatment with combination of pd- inhibition and ohsv injection. the combination treatment changed the microenvironment to be more inflammatory. our data support further preclinical and clinical studies. background: neuroblastoma is the second most common cause of cancer related death in children. treatment for high-risk neuroblastoma has improved significantly over the past twenty years, however cure rates remain below %. immunotherapy has emerged as an effective therapy for neuroblastoma, however new modalities and targets are needed to improve outcomes. objectives: our lab has developed a chimeric antigen receptor (car) that targets b -h (cd ), an immune checkpoint molecule overexpressed on many cancers, including neuroblastoma. we hypothesized that b -h would be a good target for car based immunotherapy for neuroblastoma. design/method: neuroblastoma tissue microarrays of primary patient samples were screened for b -h expression by immunohistochemistry and cell lines were screened using flow cytometry. b -h car t cells were tested in vitro by measuring tumor cell killing and cytokine production after coculture with tumor cell lines and in vivo in an orthotopic model of neuroblastoma. results: b -h expression was detected by ihc on % of the screened neuroblastoma patient samples. b -h was expressed at high levels ( + or +) in more than half of these samples ( %). almost all cell lines screened were homogeneously positive for b -h by flow cytometry. retrovirally transduced b -h . - bb. car t cells were cocultured with three b -h positive neuroblastoma cell lines (sk-n-be , kcnr, and chla ) and robust tumor cell killing was demonstrated using an incucyte assay. supernatant from the co-cultures was harvested after hours and both interferon gamma and il- production were detected by elisa.in an orthotopic subrenal capsule xenograft model of neuroblastoma, mice treated with b -h car t cells show significant reductions in tumor growth and prolonged survival compared to those treated with untransduced control t cells. however, the treatment is not always curative.b -h car t cells express high levels of exhaustion markers (pd , tim , and lag ) when compared to cd car controls. in order to overcome inhibition from exhaustion, b -h car t cells were co-cultured with neuroblastoma cell lines and pd- blocking antibody. nivolumab significantly increased the production of il- and interferon-gamma by b -h car t cells. further studies are underway to determine if b -h car t cell activity is enhanced in vivo by treating animals with pd- blockade along with car t cells. conclusion: b -h is expressed on a majority of neuroblastoma samples and appears to be a promising candidate for car t cell therapy. b -h car t cells demonstrate activity against neuroblastoma xenografts that may be enhanced by the addition of pd inhibitors. helen devos children's hospital, michigan state university, grand rapids, michigan, united states background: osteosarcoma is the most common bone tumor in children. it is often metastatic at diagnosis and in this scenario less than % of children survive. polyamines, small molecules found in all cells, are involved in many cell processes including cell cycle regulation, immune modulation, cell signaling and apoptosis. they are also involved in tumor development, invasion and metastasis. in neuroblastoma, inhibition of the polyamine biosynthesis pathway with odc inhibitor alpha-difluoromethylornithine (dfmo) results in decreased cell proliferation and differentiation. these finding have led to multiple phase i and phase ii multicenter clinical trials in pediatric neuroblastoma patients. dfmo is an attractive drug as it is oral, well-tolerated, can be given for prolonged periods and is already used in pediatric patients. the polyamine pathway has not been evaluated in osteosarcoma. objectives: evaluate effect of inhibition of polyamine biosynthesis with dfmo on osteosarcoma proliferation and cell differentiation. design/method: up to three osteosarcoma cell lines were used: mg- , u- os and saos- . cells were exposed to mm dfmo for days with replacement of media and dfmo on day . intracellular polyamine levels were measured by high performance liquid chromatography (hplc). cell numbers were obtained with a hemocytometer using trypan blue. flow cytometry cell cycle distribution (facs) and propidium iodide were used to evaluate for cell cycle arrest. the protein expression of several osteosarcoma differentiation markers was measured by sds-page and western blot using differentiation specific antibodies. a bioluminescent cell viability assay was used to measure cell recovery over several days after dfmo was removed and replaced with standard media. results: dfmo exposure resulted in significantly decreased cell proliferation in all cell lines. after treatment, intracellular spermidine levels were nearly eliminated in all cells. cell cycle arrest at g was observed in u- os. cell differentiation was most pronounced in mg- and u- os cells as determined by increased osteopontin levels. remarkably, cell proliferation continued to be suppressed for several days after removal of dfmo. conclusion: based on our findings dfmo is a promising new adjunct to the current osteosarcoma therapy for high risk patients. it is a well-tolerated oral drug that is currently in phase ii clinical trials in pediatric neuroblastoma patients as a maintenance therapy. the same type of regimen may also improve outcomes in metastatic or recurrent osteosarcoma patients for whom there have been essentially no medical advances in the last years. background: recent studies demonstrate that lower levels of the ews-fli fusion oncoprotein are associated with enhanced metastatic capability in ewing sarcoma. the nf-kb transcription factor is a critical mediator of cxcr and cxcr -driven metastasis in multiple cancers, and increased cxcr and cxcr expression have each been associated with increased metastasis and poor prognosis in ewing sarcoma. we thus sought to investigate the impact of ews-fli on cxcr /cxcr -dependent nf-kb signaling in ewing sarcoma. objectives: the goals of this study are ) to determine the impact of cxcr /cxcr signaling on metastasis-associated nf-kb target gene expression in ewing sarcoma and then ) to investigate how the ews-fli fusion oncoprotein modulates this response . design/method: we utilized multiple ewing sarcoma cells lines including a , chla , chla , tc and tc . cxcr /cxcr cell surface expression was determined by flow cytometry. ews-fli level was modulated using sirna and expression levels were confirmed by western blot and rt-pcr. p dna binding was measured via elisa. nf-kb target gene expression was assessed via rt-pcr. results: consistent with ihc analysis of primary and metastatic patient tumor samples, the paired primary and metastatic ewing sarcoma cell lines chla and chla showed dramatic differences in cxcr and cxcr expression, with the metastatic chla line demonstrating much higher expression of both receptors. other cell lines (nonpaired) showed variable cxcr /cxcr expression. genetic knock-out of cxcr lead to significant decrease in expression of both cxcl /sdf- and il- , two nf-kb transcriptional targets known to play a key role in tumor metastasis. knock-out of cxcr did not alter endogenous ews-fli mrna levels. conversely, lowering the level of ews-fli using sirna lead to enhanced nf-kb signaling, indicated by an increase in p dna binding. consistent with this observation, treating ewing cell lines with ews-fli sirna also resulted in significantly increased nf-kb target gene expression compared to control cells and target gene expression was then further enhanced upon cxcr /cxcr receptor stimulation with the receptor ligand cxcl /sdf- . our findings indicate that the ews-fli oncoprotein negatively modulates cxcr /cxcr -dependent nf-kb signaling. this suggests that ews-fli low, cxcr /cxcr high cells, which are associated with enhanced metastasis and poor prognosis, would be anticipated to exhibit enhanced expression of key nf-kb target genes. importantly, the nf-kb pathway is a druggable target that could potentially serve as an "achilles heel" in this subset of high risk tumors. current work is evaluating nf-kb inhibition as an approach to treating metastatic and refractory ewing sarcoma. background: acute graft versus host disease (agvhd) is a major cause of morbidity and mortality following allogeneic bone marrow transplant (bmt) in pediatric patients. gastrointestinal (gi) agvhd is the most serious manifestation. recently, decreased paneth cell (pc) in a predominantly adult cohort was shown to correlate with agvhd clinical grading and response to treatment. we aim to demonstrate the relationship between pc counts and gi agvhd stage and response to therapy. design/method: charts of patients who underwent endoscopy following bmt between - were reviewed. for repeated biopsies during the course of agvhd, only the first was included for analysis. one pathologist retrospectively reviewed the biopsies and counted pcs in high powered fields; the average pc count was analyzed. twenty-six percent of biopsies were reviewed by a second blinded pathologist. statistical associations between pc counts and day (d ) response, agvhd stage, and other study covariates of interest were gauged using general linear regression. agreement in pathologist pc counts was quantified by intraclass correlation (icc). the research was approved by the children's healthcare of atlanta irb. results: seventy-eight biopsies were included in the analysis. mean age at transplant was . years ± . (range: months - years). most patients underwent transplant for hematologic malignancies ( , %). the majority of transplants used a matched unrelated donor graft -including cords ( , %) and myeloablative conditioning regimens ( , %) - % received total body irradiation. of these, % were diagnosed clinically with gi agvhd (stage , %; stage , %; stage , %; stage , %). icc showed good agreement ( . ) between the pathologists. mean pc was . for patients with no gut agvhd, . for stage , . for stage , . for stage and . for stage (p = . ). on multivariate analysis pc was strongly associated with gi agvhd stage (p< . ) after controlling for age, preparative regimen intensity, and diagnosis (malignant vs. non-malignant). mean pc counts were significantly lower in patients with no response to steroid therapy at d (complete response (mean . ) vs. persistent disease ( . ) vs. partial response ( . ) (p = . )). patients diagnosed with gi agvhd with pc counts less than had a higher risk of mortality (hr . , % ci: . , . ; p = . ). lower pc count correlated with stage gi agvhd, refractory disease at d , and mortality. incorporating pc count in pathology review during gi agvhd work-up may help in agvhd risk stratification. background: there have been increasing discussions pressuring health care teams and institutions for potentially bearing the cost of clostridium difficile infections (cdi) as a health care-associated infection in the recent years. the pediatric oncology patient population, though small, accounts for significant portion of all cdi with - -fold increased risk. hematopoietic stem cell transplant (hsct) recipients constitute a unique subset with distinct risk factors, such as severe immune deficiency state and graft versus host disease (gvhd). although there is ample data on cdi in adult hsct recipients, reports on pediatric experience are limited. objectives: to evaluate the incidence and patterns of cdi among pediatric hematology, oncology and hsct inpatients at our institution. a retrospective review of all clostridium difficile (cd) stool tests performed using toxin enzyme immunoassay and later, polymerase chain reaction targeting toxin genes between and in a large, urban academic children's hospital was performed. the data were analyzed for hematology, oncology, hsct inpatient population and all the other cases separately and statistical comparisons were performed. results: a total of samples were submitted to the microbiology laboratory for cd testing during the study period. while hematology patients constituted . %, oncology . %, hsct . % and others . % of the cases on whom cd testing was done; per patient average test number was . , . , . , and . , respectively. of all the cd tests per-formed, . % were positive. test positivity was higher in hsct ( . %) and oncology ( . %) cases tested compared with hematology ( . %) and other cases ( . %) with statistical significance (p< . ). overall recurrence rate was . %; hsct patients had the highest recurrence with a rate of % followed by oncology ( . %), hematology ( . %) and other ( . %) cases, again reaching statistical significance (p< . ). again, hsct patients had the highest average number of recurrences at . ( - ) followed by oncology . ( - ), general . ( - ) and hematology . ( - ) groups. there was no seasonal variability in the incidence of cdi among populations analyzed. prolonged hospital stay/antibiotic use and persistent diarrhea due to gvhd are the likely reasons for higher rate of cd testing in hsct as a result of increased monitoring and thus might have even caused underrepresentation of positive cd test frequency. higher incidence and frequencies of recurrence underscores the inevitable nature of cdi in hsct population as a consequence of the current therapies and may lead to future radical treatment approaches like fecal implantation. background: viral infections remain a challenge to treat post hct in children, and significantly contribute to morbidity and mortality. virus specific t cells (vsts) have shown tremendous clinical efficacy in treating viral infections post-hct, with minimal toxicity and long term efficacy. we have used donor-derived vsts in individual patients, however not all donors are agreeable to the process, and numerous patients may benefit from vsts who do not have an identified donor/have other disease indications objectives: we sought to actively build a third-party vst bank, for "off the shelf" use in eligible patients. design/method: vsts targeting cmv, adenovirus and ebv were manufactured using one of techniques. initially ebv transformed b cells were genetically modified with an ad f pp vector and used as antigen presenting cells (apc) to stimulate and expand ebv, ad and cmvpp specific t cells. more recently, vsts were expanded using s of s apc pulsed with commercially available peptide pools (pep-mixes) to expand ebv/cmv/ad specific t cells. products were entered into the "bank" via two mechanisms: a) left over products from our "donor-derived" protocol when patients no longer required vsts or were not at risk of developing viral infections, or b) by targeting regular blood donors based on their hla typing to ensure an appropriate mix of high frequency hla types for optimal patient matching and antigen presentation based on current knowledge of antigen presentation. results: a total of products are currently in the thirdparty vst bank ready for use. twenty seven of these are from our donor derived protocol, and three from targeted donors. all vst products met safety and in vitro efficacy testing. thirteen vst infusions have been given to patients. eleven infusions have been given for cmv and two for adenovirus. five out of seven patients responded to thirdparty vst infusions, with a median of vst infusions per patient (range - ). the median hla matching was out of per patient (range to ) no patients experienced adverse reactions, gvhd or other toxicity related to the vst infusion. a third-party vst bank is feasible and produces clinically appropriate vsts for use in patients with viral infections. hla typing and matching of vst products is essential to reduce toxicity and promote appropriate antigen presentation and expansion of vsts in vivo. further work is underway to further characterize the vsts using epitope mapping to better define the hla restriction and immunogenicity of each vst product. akron children's hospital, akron, ohio, united states background: acute graft-versus-host disease (agvhd) is a well-known complication of hematopoietic stem cell transplant (hsct) and a major cause of post-transplant related morbidity and mortality. first line therapy of agvhd involves corticosteroids and calcineurin inhibition. in patients with severe refractory gvhd, mortality can reach up to %. currently, there is no standard of care for the treatment of steroid refractory agvhd. many centers have looked at the use of antibody mediated control of agvhd to competitively inhibit the inflammatory cascade. basiliximab, a chimeric monoclonal antibody against the t-cell il- receptor, has been used in adults with steroid refractory agvhd. patients receiving this medication have demonstrated complete and partial responses to therapy with minimal toxicities. objectives: report the successful use of basiliximab in the treatment of agvhd in a -year-old following matched unrelated (mud) hsct. design/method: a -year-old male underwent mud transplant for high risk aml with monosomy . conditioning regimen included busulfan, fludarabine and equine atg. his clinical course was complicated by fever, mucositis and agvhd (stage skin; stage gi-biopsy proven). gvhd prophylaxis included tacrolimus and methotrexate, however with progressive skin rash, diarrhea, and early satiety, gvhd treatment with corticosteroids was initiated. as the patient continued to have worsening symptoms, basiliximab therapy was started. the patient received doses ( mg) iv basiliximab on two consecutive days and then received weekly therapy for a total of doses leading to initial improvement. the patient further developed acute on chronic gvhd on day + , and subsequently received a second course of basiliximab. after initial administration of basiliximab, the patient had near complete resolution of symptoms. however, with a small wean in his tacrolimus dose, the patient experienced another skin gvhd flare prompting the second basiliximab course. the patient was subsequently weaned off all immunosuppression by day + . the only acute complication the patient experienced while receiving basiliximab was right toe paronychia and asymptomatic low ebv titer. the patient is currently off all immunosuppression at the time of report without evidence of cgvhd. conclusion: this single case report, in a young pediatric patient, demonstrates the use of basiliximab may be a safe and efficacious treatment for pediatric patients with agvhd. university of california, san diego, la jolla, california, united states background: clinical outcomes after allogeneic hematopoietic stem cell transplantation (hsct) depend on restoration of t lymphocyte populations. association between recovery of cd +foxp + regulatory t cells (tregs) and protection from chronic graft versus host disease (cgvhd) has been described in adult hsct. in adults, t cell recovery is driven by expansion of donor t cells and treg reconstitution is hypothesized to result from peripheral conversion. restoration of t cells in pediatric patients has a larger contribution from thymopoiesis, however, the relationship between thymopoiesis and treg recovery is undefined. objectives: we hypothesized that effective thymopoiesis is important for restoration of treg populations and protection from cgvhd in pediatric hsct patients. design/method: we performed longitudinal flow cytometry of peripheral blood t cells from pediatric hsct patients and age-matched healthy donors. laboratory data were correlated with clinical outcomes to evaluate impact. recovery of tregs occurred in / ( . %) patients by post-transplant day . day treg frequency in patients that developed cgvhd ( . ± . % of cd + t cells) was reduced compared to cgvhd-free patients ( . ± . %). failure to restore tregs to > . % of cd + cells by day was associated with increased risk of cgvhd in the first year post-hsct (rr = . , p = . ). a majority ( . ± . %) of tregs from patients recovering the peripheral treg compartment expressed helios, a marker of thymic-derived tregs; only . ± . % of tregs expressed helios in patients failing to restore adequate tregs. this prompted examining the relationship between defects in thymopoiesis and inability to restore tregs. we evaluated thymic function by flow cytometry quantification of cd ra+cd +ptk + recent thymic emigrant (rte) cd + cells (confirmed by qpcr for trec content). most ( / , . %) hsct patients had detectable rtes by day post-hsct. thymic production of rtes was persistently absent in patients that developed cgvhd (< / ^ cd + cells in / patients), compared to cgvhd-free patients ( / patients > rte/ ^ cd + cells by day , average . ± . / ^ cd + cells). post-hsct thymic activity as measured by rte enumeration correlated with treg restoration; / ( %) rte+ patients restored tregs, compared to / ( %) of rte-patients. conclusion: failure to restore tregs after allogeneic hsct results in increased risk for cgvhd. in pediatric patients thymic generation of new t cells is an important contributor to restoration of the treg compartment. this data supports further investigation into mechanisms impairing post-hsct thymopoiesis and suggests peripheral blood tregs may be a prognostic biomarker for cgvhd. background: haploidentical stem cell transplantation (haplo sct) is riddled with unique challenges. objectives: we present our experience in the use of haplo sct with post-transplant cyclophosphamide (ptcy) and the adaptations required for each disorder for optimal outcome. design/method: we performed a retrospective study at the pediatric blood and marrow transplant unit, apollo cancer institutes, chennai, india. children up to years of age, diagnosed to have benign disorders and underwent haplo sct with ptcy from to july were included. results: ptcy was used in i.e. % haplo transplants for children with benign disorders. the underlying conditions included fanconi anemia , severe aplastic anemia , mds , jmml , hemoglobinopathy , prca , xld and primary immunedeficiency disorders (pid) . source of stem cells was peripheral blood in %, bone marrow in %. conditioning included fludarabine with treosulphan or cyclophosphamide for pids and aplastic anemia respectively. neutrophil engraftment by day+ - with a durable graft was noted in % transplants with graft versus host disease in %, cmv reactivation in %. mortality rate was % with infants less than months of age developing severe fatal cytokine release syndrome. the median follow up is year with years being the longest. no significant late effects have been noted with chronic skin gvhd in children. survival rate was superior among children with pids with survival of % in this group. haplo sct with ptcy is a feasible and costeffective option for cure in children with life-threatening benign disorders with no compatible family or matched unrelated donor. careful patient selection, reducing cyclophosphamide related free radical toxicity with the use of n acetylcysteine, limiting t cell numbers by capping cd at × /kg, post-transplant viral monitoring protocols are required to reduce morbidity and mortality. we have been working on universal access to care for children from s of s all socioeconomic background and incorporating innovations to reduce the cost of hsct without compromising outcomes. haploidentical hsct using tcr / depletion costs usd as compared to ptcy priced at usd. children with severe aplastic anemia and pids can be transplanted using reduced intensity conditioning and ptcy. in hemoglobinopathies, pretransplant immunosuppression is required to prevent graft rejection. graft versus host disease remains the main cause of mortality in children with fanconi anemia. mortality in infants less than months after ptcy has been high, tcr / depletion would be superior in this cohort. cincinnati children's hospital medical center, cincinnati, ohio, united states background: fanconi anemia (fa) is a congenital bone marrow failure syndrome with hsct the only curative option for associated bone marrow failure. patients with fa undergoing hsct may experience increased toxicity related to either their underlying disease, or the effects of medications, resulting in the inability to tolerate prophylactic medications or sideeffects from anti-microbial therapy. objectives: we postulated that increased cd cell dose would be associated with a rapid immune reconstitution and therefore early withdrawal of anti-infective prophylactic medications. design/method: patients with fa transplanted at cchmc from an unrelated donor had peripheral blood stem cell grafts collected and cd selection performed. where possible, patients had serial measurements of their immune system performed at varying intervals post hsct. we defined immune reconstitution as normalization of lymphocyte subsets-cd , cd , cd and cd cells, as well as a normal response to mitogen stimulation including phytohemagglutinin, concanavalin a and pokeweed. the first measurement of either normal cell number or mitogen response was recorded for each patient. results: a total of patients underwent hsct for fa at cchmc between and . patient demographics included a median age of years at hsct, the vast majority of patients having a fully matched or one anti-gen mismatched donor, and the majority of patients transplanted for bone marrow failure. there was a statistically significantly decreased time post-transplant to immune cell recovery in patients receiving > × /kg cd cells (median . ) compared to those receiving < × /kg cd cells (median . ). the median time to normalization of cd count was days (cd count > /kg) versus days (cd count < /kg), cd count days (cd count > /kg) versus days (cd count < /kg), cd count days (cd count > /kg) versus days (cd count < /kg) and cd count days (cd count > /kg) versus days (cd count < /kg). time to normalization of mitogen response was decreased posttransplant in those patients receiving increased cd cell dose at time of transplant, though this was not significant, reflecting low number of patients with evaluable responses. no patients in either group experienced gvhd or graft failure. patients with fa who are transplanted with higher cd cell doses have quicker immune reconstitution than those who receive lower cell doses. along with benefit to patients including less risk of infection and early termination of immune-prophylaxis medications, this supports the use of high dose cd selected grafts in this vulnerable population. background: parvovirus b (pvb ) infection after transplantation was first reported in . since then, numerous cases of pvb infections after hematopoietic stem cell transplantation (hsct) and solid organ transplantation (sot) have been reported. most report anemia as the predominant clinical manifestation. however, pvb has been associated with pancytopenia, hepatitis, myocarditis, and allograft rejection. we present a patient with acute lymphoblastic leukemia who developed bone pain and pancytopenia following hsct in the setting of pvb infection. to describe an unusual presentation of pvb in a patient with acute lymphoblastic leukemia following hsct. design/method: a search of the english-language medical literature was performed using pubmed and medline databases. a review of the patient's medical history was performed. a year old male with relapsed b-cell all and history of "fifth disease" in infancy presented four months after hsct with focal left arm pain and difficulties fully extending the arm. bone mri showed enhancement of the medullary space centered within incomplete transverse cortical fracture interpreted as pathologic fracture due to neoplastic involvement of the ulna with no history of inciting injury. subsequently, peripheral blood counts decreased from low normal values to wbc . k/microl, anc /microl, plt k/microl, and hemoglobin . g/dl. the patient's chimerism remained % donor. a bone marrow biopsy and aspirate were performed to assess for recurrent leukemia given persistence of bone pain and developing pancytopenia. marrow findings included morphologic cytopathic effects with erythroid precursors and strong parvovirus staining with no signs of red cell aplasia or recurrent b-cell disease by morphology or flow cytometry. pvb was detected in blood by pcr and immunoglobulins with resolution of cytopenia and bone pain. this case highlights an unusual constellation of symptoms following hsct in a child with all. unexplained bone pain and medullary infiltrates with pancytopenia suggestive of recurrent leukemia were likely triggered by pvb infection. the question remains if he had reactivation of pvb , a primary infection by a new strain, or the virus was aquired through stem cells. bone biopsy could not be justified in light of clinical improvement. so far, bone lesions have only been described with congenital pvb infection. pvb appears to be uncommon after hsct, with a review of literature yielding pediatric cases. however, it may be underestimated due to lack of routine screening. our patient's presentation supports that evaluating for pvb may be warranted in hsct patients presenting with symptoms suggestive of relapsed leukemia. background: cardiac injury may occur during hematopoietic stem cell transplant (hsct) in pediatric patients and can be asymptomatic for many years. recommendations for screening are available for patients who received anthracyclines or chest irradiation, but no guidelines exist for unexposed longterm survivors. we sought to define the prevalence of echocardiographic abnormalities in long-term survivors of pediatric hsct and determine the need for screening in asymptomatic patients. design/method: we analyzed echocardiograms performed on long-term survivors (≥ five years) who underwent hsct at cincinnati children's hospital between and . we analyzed echocardiograms for left ventricular ejection fraction (ef), end-diastolic dimension (lvedd), septal thickness, posterior wall thickness, and global longitudinal strain (gls). we normalized linear measurements for age and patient body surface area. we included for further analysis patients who had echocardiogram obtained for routine surveillance. results: a total of patients underwent hsct and were alive more than years after transplant in , with having an echocardiogram obtained ≥ five years postinfusion. those with an echocardiogram were transplanted more recently (median vs. ). however, no difference between screened and unscreened individuals was noted for age at transplant, sex, transplant indication, anthracycline exposure, chest irradiation, or cyclophosphamide based preparative regimen. indications for echocardiograms included: cardiac symptoms ( . %), congenital cardiac anomalies ( . %), hypertension ( . %), known cardiac or pulmonary disease ( . %), routine post-hsct surveillance ( . %), and unknown ( . %). the mean time post-hsct was . years. among routine surveillance echocardiograms, the mean ef z-score was - . . mean lvedd zscore was - . , mean septal thickness z-score - . , mean posterior wall thickness z-score - . , and mean gls - . %. for patients that had echocardiogram performed for routine surveillance, / patients ( . %) had ef measured, and / ( . %) had ef z-scores ≤ - . (abnormally low). patients exposed to anthracyclines had a mean z-score ef of - . vs. unexposed patients - . (p = . ). among individuals who received neither anthracyclines nor tbi only / ( . %) was found to have an abnormal ef, . % (z-score - . ) or gls (- . %). only one patient who had a normal ejection fraction (z-score - . , ef . %) had an abnormal gls, - . % (normal ≤ - . ). long-term survivors of pediatric hsct who are asymptomatic and did not receive radiation or anthracyclines likely do not require surveillance echocardiograms, unless indicated by clinical symptoms. patients exposed to anthracyclines or tbi require close echocardiographic s of s screening and clinical monitoring for the development of cardiac complications. duke children's hospital, durham, north carolina, united states background: children undergoing pediatric blood and marrow transplants (pbmt) experience significant symptom distress. mobile health (mhealth) technologies can be leveraged to collect and monitor patient generated health data, and subsequently enhance our understanding of pbmt symptom clusters, patterns, and trajectories. better understanding of symptom complexity can foster development of precision health strategies to improve patient outcomes. however, limited research exists in integrating mhealth technology into pbmt management. we aimed to explore the feasibility, acceptability, and usability of using a pbmt specific mobile application to collect and monitor symptoms and wearable technology (apple watch) to measure objective data such as heart rate (hr) and activity. design/method: an exploratory mixed method design began in october to monitor pbmt symptoms for patients using real-time data from: ) a self-developed mhealth application (app) to collect subjective symptom data; and ) apple watch to collect physiologic measures such as heart rate and number of daily steps. data is collected pre-transplant through days. acceptability will be assessed through satisfaction surveys at study completion. we have enrolled patients to date who are all currently using the app and watch. patients' average frequency of daily charting in the app %. the wearable average daily recorded measurements are for hr and for step count. most common symptoms recorded within the app include fatigue and pain. we have noted trends in data including a decrease in activity following transplant and gvhd and an increase following engraftment. patients have stated "the app is helpful to keep track of how my pain is doing day to day" and "i try to take more steps each day than the day before". patients often remove the watch for charging, then forget to put it back on, but consistently put it on upon reminder. finally, parents often were required to make app entries with patients too sick to record. we continue to enroll patients with enthusiasm from both patients and parents to use mhealth during pbmt. preliminary findings suggest feasibility of using the mhealth devices is strongly correlated to the patient's post-transplant stage and is facilitated by caregiver participation with device management (charging devices, reminders to wear watch and record in app). patients reported satisfaction and ease of use with devices, but found it difficult to keep up with charging and charting. these findings indicate using mobile devices may be useful methods to collect patient generated health data. cincinnati children's hospital medical center, cincinnati, ohio, united states background: bacterial bloodstream infections (bsi) are a common complication following hematopoietic stem cell transplantation (hsct) in both pediatric and adult populations, and are associated with poor outcomes. there is limited data describing the outcomes and characteristics of patients who develop three or more bsi after hsct. objectives: to describe the characteristics and outcomes of pediatric patients who develop three or more blood stream infections in the first-year post hsct. design/method: we performed a retrospective chart review of consecutive patients who underwent hsct at our institution from through to compile this case series. data were collected through the first year post-hsct including: patient demographics, underlying disease and therapy characteristics; and transplant complications such as thrombotic microangiopathy (tma), graft versus host disease (gvhd) and overall survival. bsis were classified according to current center of disease control guidelines. results: of patients, ( %) developed or more bsi in the first-year post transplant (total bsi cases = including all patients). of the cases, the majority underwent allogeneic hsct (n = / ; %). most cases were from unrelated donor (n = / , %). more than half of patients had grade - gvhd (n = / , %). sixteen ( %) had tma. of these cases, tma preceded the first bsi in n = / ( %). the majority of bsis were classified as central line-associated bloodstream infections (clabsis, n = / , %), followed by mucosal barrier injury laboratory-confirmed bloodstream infections (n = / , %) and secondary bsi (n = / , %). the majority of isolated organisms ( %) were associated with mucosal barrier injury pathogens. one-year overall survival in the cohort was % (n = / ). pediatric patients undergoing hsct who develop or more bsis in the first-year post transplant demonstrated an increased rate of tma compared to the overall institutional incidence of roughly %. tma diagnosis preceded the first bsi in over half of patients, suggesting that tma may predispose to recurrent bsi. improved strategies for early detection and treatment of tma as well as prevention of clabsis may help reduce the number of bsis ultimately leading to decreased morbidity and mortality in this patient population. background: in neutropenic pediatric patients, infection remains a significant cause of morbidity and mortality. while granulocyte transfusions have been utilized for decades to treat infections, including in the pediatric population, the efficacy of this intervention remains poorly described. previous guidelines have primarily utilized information from adult populations. furthermore, recruitment of donors typically involves friends or relatives of the patient with periodic involvement of community donors. the use of a readily available local donor population to improve availability has yet to be well described. as the immunocompromised population is particularly susceptible to worsening infection and clinical deterioration, the ability to rapidly harvest and deliver granulocytes warrants further investigation. to investigate the efficacy, safety, and outcomes of severely immunocompromised patients receiving granulocyte transfusions from a local altruistic granulocyte program in a pediatric tertiary care center. design/method: a retrospective review was performed to evaluate the context for receiving a transfusion as well as primary outcomes including infection clearance, survival to discharge, and overall mortality. the indiana blood bank assisted with timing the interval from initial order placement to onset of first granulocyte infusion. results: among the patient population reviewed, patients received separate granulocyte regimens. ages ranged from - years with a mean neutrophil count of at time of first transfusion. indications for transfusions included bacteremia (n = ), fungal pneumonia (n = ), and fungemia (n = ). primary outcomes included clearing infection ( %) and surviving to discharge ( %). the median time from initial order placement to infusion was hours, although there was no significant difference between responders who cleared the infection and non-responders who did not. however, additional investigation found that ward patients had a % chance of surviving to discharge while patients in the icu at time of initial transfusion had a % chance of survival to discharge. the readily available granulocyte transfusion program allows patients to quickly receive therapy in neutropenic settings. this is beneficial for patients as transfusion prior to clinical decompensation correlates with increased likelihood of infection clearance, and subsequently improved mortality. further investigation is needed, likely as a prospective study, to better explore circumstances that are beneficial for granulocyte transfusions. background: donor lymphocyte infusions (dli) are composed of immune cells to treat relapse after hematopoietic cell transplantation (hct). to date, data regarding its efficacy is limited in pediatric populations. furthermore, while outcomes related to cd content have been characterized, to our knowledge, the relationship between outcomes and other cellular content in dli has never been reported. objectives: determine whether the primary hematological malignancy, presence/absence of graft-versus-host disease s of s (gvhd), and unique phenotypic content of each dli impact overall survival (os) in pediatric patients with hematological malignancies. design/method: irb-approved, retrospective study investigating all consecutive dlis given to patients at the children's hospital of wisconsin. analyses were conducted using mann-whitney, fisher's exact, and chi-square. from from - patients ≤ years old with hematologic malignancies [myeloid (aml/ mds/cml/jmml),n = ; lymphoid (all),n = ] underwent dlis ( %% ≥ dlis). the median time between hct and dli was . (range, . - . ) years. there were significant differences between the lymphoid and myeloid groups, respectively, in regard to median age at hct ( . vs . yrs, p = . ) and at first dli ( vs years, p = . ). ultimately, there were no statistically significant differences in gvhd or os in products with either higher or lower cd , cd , cd , cd , or cd cellular content. however, the median cd /kg content was more than double in the patients who developed gvhd as compared to patients who exhibited no gvhd after dli ( . × vs . × , p = . ). patients receiving one dli had a -year os of ± % vs those receiving + dli of ± % (p = . ). with a median follow-up of . (range, . - . ) years, the year estimated os of patients in the lymphoid group was higher at ± % vs ± % in the myeloid group, although not significant (p = . ). our results indicate a survival benefit when using dli in a subset of patients who relapse after hct. unlike adult studies demonstrating little effect of dli in lymphoid diseases, many children with all achieved durable remission. while our analysis did not demonstrate that dli cellular content had a statistically significant effect on gvhd or os, it is possible that differences could be found if a larger population and more targeted cell doses were studied. more data will be needed to further define these relationships and identify patients who stand to benefit most. cincinnati children's hospital medical center, cincinnati, ohio, united states background: many arabic speaking muslim parents of children requiring bone marrow transplantation (bmt) receive medical care in the united states. providers may not understand the impact of islamic parents' religious beliefs and practices on their health care experience. objectives: to explore how islamic parents used religion in decision making and to understand the impact of their religious beliefs and practices on their overall health care experience. design/method: we used grounded theory, an inductive method gathering data from interviews and analyzing text, to identify core themes. ten caregivers of bmt children from middle eastern countries were interviewed by an arabicspeaking provider; interviews were coded by an interdisciplinary team. we identified key themes: . patience is a core belief in islam. patience results from the acceptance of allah's will. behaviors showing patience include praying rather than questioning and crying. . al qur'an provides comfort, healing, and protection. families listen to recitations of al qur'an in the patient's room because they feel that this practice not only comforts them but promotes healing as well. for some, certain portions of the qur'an were especially meaningful such as surat al-baqara, which explains that while we may think something is bad for us, allah will know it is good for us. . religious care in the medical center helped families feel respected. religious care in the medical center included interactions with chaplains, who were understood to be "religion experts," and provision of space for prayer and religious resources. . seeking religious consultation. religious consultation from imams or religious scholars (muftis or sheikhs) provides interpretations of the qur'an applied to the family's specific situation helps families make difficult decisions and follow allah's plan. . muslim beliefs guided decision making; muslim practices brought comfort, strength, and peace. drawn from the parents' understanding of islam. parents who addressed this topic said they would only do what islam allowed. they did indicate that most aspects of healthcare were understood to be allowed within islam. additionally, muslim practices of prayer, reading/listening to qur'an, and giving alms all provided comfort, strength and peace. we identified several recurring themes through our interviews that allowed us to understand how families use their muslim faith to deal with their children's illnesses and how it influences their decision making. we believe this better understanding will allow for more informed conversations about patients' health care and decision making, and shows respect for religious beliefs and practices. nemours/dupont hospital for children, wilmington, delaware, united states background: virtually all children will be infected with human herpesvirus (hhv- ) by the age of two. hhv- reactivation after stem cell transplantation causes multiorgan toxicities, including encephalitis, with inflammation and destruction of the temporal lobes and hippocampi, memory loss, and seizures. catatonia is characterized by posturing, immobility, mutism, and autonomic instability, and it's associated with various psychiatric and medical conditions. we describe a patient with hhv- encephalitis and unusual neurologic sequelae, including cognitive and neurobehavioral dysfunction and catatonia, which may impact our understanding of the pathophysiology of hhv- reactivation encephalitis. objectives: describe a case of hhv encephalitis with practice implications for stem cell transplantation. results: our patient was diagnosed with acute myeloid leukemia at age . within years, he relapsed and received two stem cell transplants. on the th day after his second transplant, he developed hyponatremia and refractory seizures. brain mri showed edema in the medial right temporal lobe with linear ischemic change. eeg showed diffuse encephalopathy. cerebrospinal fluid (csf) demonstrated white blood cells, red blood cells, and hhv- by pcr. his prophylactic antiviral was switched to foscarnet and ganciclovir. repeat mri showed abnormal signals in bilateral medial temporal lobes and the right insula. three months later he developed episodes of diaphoresis, hypothermia, agitation, mutism, and unusual posturing, recurring almost daily, recognized as catatonia. mri showed improvement of the abnormalities in the bilateral medial temporal lobes and hippocampi. eegs showed diffuse slowing. after months of antiviral therapy, csf was negative for hhv- . over the ensuing years, he had numerous episodes of diaphoresis, hypertension, hypothermia, pruritis, confusion, agitation, cogwheel rigidity, and bizarre posturing. dopamine blocking agents did not help. clonazepam helped reduce their frequency, and hot showers helped break acute episodes. further mris showed generalized cortical volume loss. he suffered from depression and severely impaired sleep and cognitive function. we describe a novel, debilitating outcome of hhv encephalitis which may provide diagnostic considerations as we continue to improve our understanding of the breadth of possible neurologic sequelae in transplant patients. hhv- is understood to infect and destroy the temporal lobes and hippocampi, but our patient's autonomic dysfunction indicate involvement of the hypothalamus and basal ganglia. antidopaminergic agents may worsen catatonia, and they were not effective for our patient. treatment of catatonia includes benzodiazepines; electroconvulsive therapy was not attempted in this case but may also be useful. background: epstein-barr virus (ebv)-related posttransplant lymphoproliferative disorder (ptld) is a lifethreatening complication in patients following hematopoietic stem cell transplantation, with a frequency estimated at . % and a cumulative incidence of mortality estimated as high as %. studies of ebv have hypothesized that the tonsils are critical for propagating this infection, as tonsillar epithelial cells have been shown to be the site of primary viral infection and continued viral shedding; however, to date no studies have been performed assessing the role of tonsillectomy in patients with ebv ptld. objectives: identify patients with localized ebv ptld treated with tonsillectomy to identify prognostic factors that may be able to help guide future treatment decisions. design/method: patients treated at memorial sloan kettering cancer center who had received hematopoietic stem cell transplantation and had billing codes for both ebv and tonsillectomy were eligible for inclusion in this study. a retrospective chart review was performed, assessing patient demographics, transplant characteristics, laboratory values, tonsillar pathology, and clinical course. any patient who did not have unilateral or bilateral tonsillectomy performed or who had non-localized disease (defined as disease involvement outside of the oropharynx and neck) was subsequently immunodeficiency; % (n = / ) fanconi anemia (fa); % (n = / ) hemoglobinopathy; % (n = / ) non-fa marrow failure and % (n = / ) a metabolic disorder. seventy one percent (n = / ) had normal amh for age pre-transplant, % (n = / ) had low amh for age pre-transplant; of these, % (n = / ) had an oncologic diagnosis; % (n = / ) had fa; % (n = / ) had previously treated hlh; % (n = / ) had non-fa marrow failure; one had a metabolic disorder and one a hemoglobinopathy. of the patients with post-transplant amh measurement % (n = / ) had low levels. of the patients with previously normal pre-transplant amh % (n = / ) underwent myeloablative conditioning (mac) regimen with a % (n = / ) having low amh levels post-transplant compared to %(n = / ) who underwent reduced intensity conditioning (ric) regimen with % (n = / ) having low amh levels post-transplant (p . ). fifteen percent (n = / ) had low levels pre-transplant and underwent mac regimen with % (n = / ) remaining low; % of these patients (n = / ) had fa. nine percent (n = / ) had low levels and underwent a ric regimen with % (n = / ) of amh levels remaining low; % (n = / ) of these patients had hlh treated prior to transplant. conclusion: amh levels can be used for detection of premature ovarian failure and fertility counseling. there is a higher risk of premature ovarian failure with mac regimens and prior chemotherapy vs ric regimens. follow up of this cohort will provide more information to understand the effects of hsct in ovarian function and the usefulness of amh as a predictor of fertility potential. background: there are no proven strategies to prevent blood stream infections (bsi) secondary to oral mucosal barrier injury after hematopoietic stem cell transplant (hsct). additionally, we recently reported progressive gingivitis and dental plaque accumulation in hsct recipients despite our current oral standard of care (three times daily oral rinse). xylitol is a non-fermentable sugar alcohol that reduces dental caries, plaque accumulation, and oral disease progression by inhibiting bacterial growth. we hypothesized that the addition of xylitol to standard oral care will decrease dental plaque accumulation, gingivitis and bacteremia from oral flora. objectives: identify a clinically effective strategy to improve oral health and prevent bsi secondary to bacterial translocation through the oral mucosa in patients undergoing hsct. we are conducting a prospective randomized control study to test our hypothesis. those in the intervention arm receive our current standard of care (three times daily oral rinse) in addition to daily xylitol wipes; controls receive oral standard of care alone. oral exams are performed at baseline and weekly for the first days post hsct. metagenomic shotgun sequencing (mss) of gingival samples is performed at all time points to evaluate microbiome diversity and pathogenic bacterial load. finally, we performed whole genome sequencing of pathogenic bacterial isolates causing bacteremia to assess for genetic relatedness to corresponding strains present within the patient's oral microbiome preceding the infection. : preliminary interim analysis of patients demonstrates improved oral health in patients receiving xylitol (n = ) over those receiving standard of care (n = ), measured by the oral hygiene index (p = . ) and gingivitis index (p = . ). in the nine patients having complete oral mss analysis, xylitol appeared to be associated with decreased streptococcus mitis/oralis domination in the oral microbiome. finally, patients receiving xylitol had no incidence of streptococcus mitis/oralis bacteremia through the first days compared to three patients ( %) in standard of care arm. interestingly, streptococcus mitis/oralis comprised % of the oral microbiome in one child who subsequently developed a streptococcus mitis/oralis bsi. we expect to complete this study in the next months (n = ). the addition of xylitol to oral standard care appears to decrease dental plaque and gingivitis in patients undergoing hsct. xylitol may also impede streptococcus mitis/oralis dominance in the oral microbiome with potential reduction in blood stream infections. (range: - days). twenty-one mdli ( %) were administered because of lymphopenia, fourteen of them ( %) in patients with concomitant viral/opportunistic infections. mixed chimerism/graft failure was the motive of % of the mdli (n = ) and six ( %) were administered to accelerate immune reconstitution. all infusions were well tolerated without appearance or worsening of gvhd. an increase in t-cell counts was observed following six mdli ( . %), although it was a transitory response ( - weeks) in five cases. viral/opportunistic infections were controlled in five cases ( . %), requiring a median of mdli to achieve this response. none of the mdli administered in cases of mixed chimerism/graft failure were effective in reverting this situation. our preliminary data suggests that mdli, is a safe adoptive immunotherapy strategy even with high dose of t-cells without infusion side effects or gvhd complications. some efficacy has been observed in patients with lymphopenia and opportunistic infections, with no positive results in patients with mixed chimerism/graft failure, up to date. however, to determine the real efficacy of this strategy, prospective studies are required. jun zhao, kristen beebe, lucia mirea, alexandra walsh, shane lipskind, alexander, ngwube phoenix children's hospital, phoenix, arizona, united states background: male adolescents undergoing myeloablative hematopoietic stem cell transplantation (hsct) develop infertility with impaired spermatogenesis with reported rates ranging from % to %. in nonmalignant diseases, myeloablative regimens have been replaced with reduced intensity conditioning (ric) with the hopes of better survival rate, less organ toxicity and improved quality of life. despite the increased use of ric regimens for hsct, the effects of ric on fertility remain unknown. objectives: to assess fertility following ric hsct in young adult males. we assessed gonadal function and semen characteristics in adolescent males (> years) who received a single ric hsct at phoenix children's hospital for nonmalignant diseases during - . male patients who were a minimum of year from ric hsct and had postpubertal development at tanner stage iii or above were eligible for this study. gonadal status was assessed by measuring fsh, lh, testosterone, and inhibin b levels, and semen anal-yses assessed fertility indicators (semen volume, sperm concentration, motility, viability, forward progression, morphology, and total count). results: hormone levels and semen analysis have been obtained for patients thus far. the median time between transplant and semen analysis was years. post hsct, ( %) patients showed abnormally elevated lh levels, but fsh, testosterone (total and free), and inhibin b levels were within normal range for all patients. sperm morphology and viability testing were not able to be performed due to low concentrations and volumes. as a result, the total motile sperm count, the most useful estimate for fertile potential, is essentially for all patients. conclusion: recruitment is ongoing, but so far our limited results suggest that ric hsct may have detrimental longterm effects on male fertility. a multi-institutional trial may be appropriate due to small patient numbers at each institution. we are currently exploring options to expand to other centers. further consideration is warranted regarding decisions made by providers, ways to improve anticipatory counseling provided to patients and their families prior to transplant, and how to augment the preventive care of these patients in longterm follow-up. currently all male patients being considered for ric transplant should be counseled to sperm bank prior to transplant. background: a previous systematic literature review identified all published studies of defibrotide treatment for patients of all ages with vod/sos. to assess day+ survival for defibrotidetreated pediatric patients (≤ or ≤ years, per study) all patients exhibited infectious complications with at least viral infection. four patients also had bacterial infections. of note, no patient developed evidence of fungal infections. conclusion: early institution of ecp in patients with high risk acute gvhd (grade - ) was very effective at treating agvhd, allowed for an aggressive steroid taper and contributed to excellent overall survival rates ( %). infectious complications were primarily viral and bacterial, with no fungal infections in this very high risk population. background: vod/sos is a life-threatening complication of hsct conditioning. vod/sos with multi-organ dysfunction (mod) may be associated with > % mortality. defibrotide is approved to treat hepatic vod/sos with renal/pulmonary dysfunction post-hsct in the us and severe hepatic vod/sos post-hsct patients aged > month in the eu. there are few published data on survival of neuroblastoma patients with vod/sos post-hsct. objectives: to report day+ survival and safety post hoc for patients with neuroblastoma and vod/sos post-hsct in the defibrotide t-ind trial. design/method: vod/sos was diagnosed by baltimore or modified seattle criteria or biopsy, with/without mod, after hsct or chemotherapy. defibrotide treatment ( mg/kg/day) was recommended for ≥ days. this post hoc analysis is based on adult and pediatric patients receiving ≥ dose of defibrotide, including with mod. results: among patients with neuroblastoma, developed vod/sos after hsct. for these post-hsct patients, . % were male and . % were female, median age was years (range - years): . % aged - months, . % - years, . % - years, and patient > years. day+ survival data were available for / of these neuroblastoma patients ( with mod and without mod); had autologous and had allogeneic transplants. kaplan-meier estimated day+ survival for the neuroblastoma group was . % ( % confidence interval [ci] , . %- . %). for the mod and no mod subgroups, kaplan-meier estimated day+ survival was . % ( % ci, . %- . %) and . % ( % ci, . %- . %), respectively. in the overall t-ind hsct population aged ≤ years (n = ) and pediatric autologous hsct subgroup (n = ), kaplan-meier estimated day+ survival was . % and . %, respectively. treatment emergent adverse events (teaes) occurred in . % (n = / ), with serious teaes in . % ( / ; most common: multi-organ failure, . % [ / ]). teaes lead to treatment discontinuation in . % (n = ; most common: pulmonary hemorrhage, n = ); death occurred in . % (n = ; > %: multi-organ failure, . %; vod/sos, . %). treatment-related adverse events, as assessed by investigators, occurred in . % (n = ; most common: pulmonary hemorrhage, . %). this post hoc analysis found kaplan-meier estimated day+ survival of . % in patients with neuroblastoma and vod/sos post-hsct, which was consistent with outcomes in pediatric patients after autologous hsct. the safety profile of defibrotide in neuroblastoma patients was consistent with the overall hsct population in this study and other defibrotide studies in pediatric patients. cincinnati children's hospital medical center, cincinnati, ohio, united states background: blood stream infections occur in nearly % of patients undergoing hematopoietic stem cell transplant (hsct) and fever is often the first symptom. timely administration of antibiotics is associated with improved outcomes, thus, early recognition of fever is paramount. current standard of care (soc) includes episodic monitoring of temperature in hospitalized patients, which may delay fever detection. therefore, continuous real-time body temperature measurement may detect fever prior to the current soc. temptraq is a food and drug administration cleared class ii medical device and consists of a soft, comfortable, disposable patch that results: of patients, were started on a pca in the days post hct. % were male with median age of y. % had all, and % aml. matched related donors were used in % and % received tbi. pca was initiated median d+ . oral mucositis alone was the most common indication ( %). a majority of patients were started on hydromorphone ( %); % started on morphine and % started on fentanyl. % started on continuous infusion. pca was used for a median of days (range - days). median pain score was highest d+ of pca use, however, there was inconsistency in charting of numerical pain scores. on d+ , patients had insufficient data to determine efficacy of pain control; of the remaining patients, % had good pain control while % had moderate and % had poor pain control using our devised scale. the most common toxicity observed was respiratory depression (∼ %), however, etiology was often multifactorial and not due to opiates alone. analysis is ongoing to assess variables predicting pca use as well as efficacy of pain control and correlation between current reporting scales and patient perception. conclusion: pca use is common in pediatric hct yet pain control remains inadequate. there's a need for better evaluation of pca management, especially uniform assessment of pain, thereby improving quality of life post hct. children's national health system, washington, district of columbia, united states background: actinomycosis is a rare invasive anaerobic gram-positive bacterial disease caused by actinomyces spp. that may colonize the oropathynx, gastrointestinal tract and urogenitial tract and can lead to abscesses. respiratory tract actinomycosis is characterized by pulmonary cavities, nodules, consolidations and pleural effusions. although actinomyces are nearly always sensitive to penicillin they are frequently resistant to cephalosporins and variable sensitives to fluoroquinolones. although rare in children, immunosuppressed patients are at increased risk for actinomycosis. to describe a case of next-generation sequencing identification of actinomycosis. a -year-old male with a history of very high risk b-cell acute lymphoblastic leukemia who was months status post a / matched unrelated donor bone marrow transplant complicated by prolonged fevers, persistent weight loss, and splenic lesions, treated with posaconazole and levofloxacin developed fever and cough in the setting of neutropenia. blood cultures demonstrated staphylococcus epidermidis. ct showed micronodules and effusion not consistent with s. epi, prompting bronchoscopy. all bacterial cultures were negative. patient was prescribed a three-week course of vancomycin with rapid improvement. design/method: s next generation sequencing (ngs) from bronchoalveolar levage sample was performed at the university of washington laboratory results: ngs assay from bronchoalveolar lavage showed major abundance of actinomyces most closely related to meyeri or oodontolyticus. demonstrated actinomyces. the patient was started on a six month course of amoxicillin with continued clinical improvement. in retrospect, the splenic nodules that were presumed fungal disease were likely actinomycosis, partially treated with levofloxacin. this case highlights the potential utility of ngs in the diagnosis of rare diseases in immunocompromised patients. actinomycosis was only demonstrated through ngs and led to a change in treatment regimen and durable clinical improvement. because actinomyces often mimics malignancy, tuberculosis or nocardiosis, the use of this novel test both targeted appropriate therapy and reduced the exposure to unnecessary medications to treat the differential diagnosis. finally, we highlight that actinomyces should be considered in patients who present with unexplained fevers, weight loss, and night sweats. haneen shalabi, cynthia delbrook, maryalice stetler-stevenson, constance yuan, bonnie yates, terry j. fry, nirali n. shah center for cancer research, national cancer institute, national institute of health, bethesda, maryland, united states background: car-t therapy, while effective, may not be durable for all, and antigen negative escape is a growing problem. hct, in relapsed/refractory all, can be curative, particularly for those in an mrd negative remission. we demonstrated that cd directed car-t therapy effectively rendered patients into mrd negative remissions (by flow cytometry) and the leukemia free survival post-hct was high . in pastorek, jesssica bruce, michael a. pulsipher, chloe anthias, peter bader, andre willasch, jennifer sees, jennifer hoag, wendy pelletier, brent logan, pintip chitphakdithai, lori wiener university of pittsburgh, pittsburgh, pennsylvania, united states background: more than , pediatric hscts are performed in north american and europe each year. the ethics of exposing a healthy child to donation procedures which have some risks and no direct medical benefits continue to be a topic of debate. pediatric donors may experience psychological distress and poorer quality-of-life during and after donation compared to healthy controls. although there are fact/jacie requirements related to the management of pediatric donors, it is unclear what standardized practices exist for psychosocial assessment/management of this group. objectives: to describe transplant center practices for psychosocial evaluation/ management of pediatric donors (< years) and to examine differences in practices by location (cibmtr/ebmt) and number of harvests (volume). design/method: data were collected via a single crosssectional survey distributed electronically to cibmtr and ebmt centers between / / and / / . : / ( %) of cibmtr and / ( %) of ebmt centers completed the survey. most centers had written eligibility guidelines for pediatric donors ( %). most also had a process for ensuring that donors were freely assenting to donate ( %), managed by a transplant physician ( %). a single physician often jointly managed donor/recipient care ( %). half of centers had a pediatric donor advocate ( %), who was most often a physician ( %) or social worker ( %). cost was the largest barrier to having a donor advocate ( %). most centers performed psychosocial screening of donors ( %) but rarely declined donors based on psychosocial concerns ( %). less than half of centers provided post-donation psychosocial follow-up ( %). comparisons by center location indicated that ebmt centers were more likely to have a physician doing joint donor/recipient care ( % vs. %; p = . ), less likely to have a psychosocial assessment policy ( % vs. %; p = . ), less likely to have a donor advocate ( % vs. %; p = . ), but marginally more likely to do post-donation psychosocial follow-up ( % vs. %; p = . ). large volume centers were more likely to have a psychosocial assessment policy than their medium/smaller counterparts ( % vs. %, %; p = . ) â€"there were no other differences on key psychosocial management variables by volume. although most centers have written guidelines for pediatric donor eligibility and mechanisms for ensuring assent, substantial numbers of donors do not undergo psychosocial assessment, are jointly managed with the recipient by a single physician without an assigned donor advocate, and do not receive psychosocial follow-up. the field would benefit from guideline development for the psychosocial management of pediatric donors. background: germline mutations in samd and samd l genes cause mirage (myelodysplasia, infection, restriction of growth, adrenal hypoplasia, genital phenotypes and enteropathy) and ataxia-pancytopenia syndromes, respectively, and are associated with chromosome deletions, mds and bone marrow failure (bmf). there are limited data on outcomes of hct in these patients. to describe outcomes of allogeneic hct in patients with hematologic disorders associated with samd /samd l mutations. results: seven patients underwent allogeneic hct for primary mds (n = ), congenital amegakaryocytic thrombocytopenia (camt)(n = ), and dyskeratosis congenita (n = ). retrospective exome sequencing revealed gain-of-function mutations in samd (n = ) or samd l (n = ) genes. constitutional mosaic monosomy was present in cases. two samd patients had features of mirage syndrome. unusual findings of panhypopituitarism, laryngeal cleft, and glomerulosclerosis were noted in one case. in another case with a samd mutation hypospadias & bifid scrotum were the only findings. the remaining patients had no phenotypic abnormalities. median age at hct was y (range: . - . ). patients received transplants from bone marrow (matched unrelated (n = ) & hla identical sibling (n = )), or unrelated cord blood (ucb) (n = ). five mds patients received myeloablative s of s conditioning (busulfan-based (n = ) or tbi-based (n = )); patients (mds (n = ); camt (n = )) received reducedintensity conditioning (ric) (fludarabine, cyclophosphamide, with ratg or alemtuzumab). syndrome-related comorbidities (diarrhea, infections, malnutrition, electrolyte imbalance, lung disease and hypoxia) were present in both patients with mirage syndrome. one patient with a familial samd l mutation, mds and morbid obesity failed to engraft following ric double ucbt. she died one year later from refractory aml. all other patients achieved neutrophil and platelet engraftment, at a median (range) of ( - ) and ( - ) days, respectively. posttransplant complications included severe hypertension (n = ), pericardial effusions (n = ), veno-occlusive disease of liver (n = ), and recurrent aspiration pneumonias (n = ). one patient developed grade iii agvhd which resolved with treatment. one patient developed mild skin cgvhd and suffers from chronic lung disease. all surviving patients had resolution of hematological disorder and sustained peripheral blood donor chimerism ( - %). overall survival was % with a median follow-up of years (range: . - . y). patients with hematological disorders associated with germline samd /samd l mutations tolerated transplant conditioning without unusual, or unexpectedly severe toxicities. allogeneic hct led to successful resolution of mds or bmf, with excellent overall survival. more data is needed to refine transplant approaches in samd /samd l patients with significant comorbidities, and develop guidelines for their long-term follow-up. shyamli singla, tiffany simms-waldrip, andrew y. koh, victor m. aquino background: steroid-refractory acute graft versus host disease (agvhd) is a potentially fatal complication of allogeneic hematopoietic stem cell transplantation (hsct). basiliximab (anti-il -r monoclonal antibody) as a single agent or in combination infliximab (anti-tnf-monoclonal antibody) has demonstrated efficacy in adult cohorts with steroid-refractory agvhd, but has not been well studied in the pediatric population. we adopted the use of basiliximab and infliximab as our institutional standard of care for steroid-refractory agvhd in pediatric hsct patients. to determine the response and survival of hsct children who received basiliximab and infliximab for the treatment of steroid-refractory agvhd. design/method: we retrospectively reviewed children who received basiliximab and infliximab for steroid-refractory agvhd refractory between september and december . complete response (cr) was defined as resolution of all clinical signs of agvhd. partial response (pr) was defined as at least one grade reduction in one target organ (e.g. skin, gut or liver) without increased grade in another target organ. no response was defined as either no improvement or progressive worsening of agvhd in at least one organ. baseline demographics, transplant details, laboratory findings, and treatment outcomes were also evaluated. results: of the evaluable hsct patients, children (median age yrs, range mo- yrs) with steroid-refractory agvhd received combination monoclonal antibody (mab) therapy. the median time from the start of steroid therapy to initiation of mab was days. the overall glucksberg grade of agvhd at the time of initiating mab therapy was grade i (n = , . %) ii (n = ; %), iii (n = ; %) or iv (n = ; %). the overall response rate was %, with ( %) patients achieving cr, ( . %) patients achieving pr, and ( . %) patients with no response at days following the start of mab therapy. the median overall survival was , , and days for patients who exhibited cr, pr, and no response, respectively. the overall survival at year following start of mab therapy was %. background: the role of high dose chemotherapy (hdc) and autologous stem cell rescue (ascr) in patients with high risk (advanced metastatic or relapsed) soft tissue sarcomas is controversial. despite multimodal chemotherapy, radiotherapy, and local control measure advancements, prognosis of patients with advanced metastatic or unresectable and relapsed sarcomas remains poor, with less than % years disease free survival. objectives: to determine if consolidation with myeloablative hdc and ascr improves relapse free (rfs) and overall survival (os) outcomes in a high risk patient subgroup. we performed retrospective review of all high risk soft tissue sarcoma patients who underwent hdc and ascr at the children's hospital at montefiore, bronx, ny between october and january . the protocol was approved by albert einstein college of medicine institutional review board. results: patients ( primary metastatic high risk disease, relapsed or recurrent disease) received hdc with ascr. primary diagnoses were rhabdomyosarcoma (rms) (n = , alveolar histology), primary site nasopharynx (n = ) and lower extremity (n = ). ewing's sarcoma (ews) (n = ), axial site (pelvic) in patients ( %). median age years (range - years), ( %) were male. all patients were in complete metabolic remission before transplant. median pre transplant comorbidity index was (range - ). patients ( rms and ews) received conditioning with carboplatin, etoposide and melphalan. remaining patients with ews received conditioning with busulfan, melphalan and topotecan. all patients received peripheral blood mobilized hematopoietic stem cell transplantation. stem cell mobilization achieved with high dose filgrastim in all patients except one who required addition of plerixafor. median cd +/kg s of s recipient body weight cell dose infused was . × ^ (range . - . × ^ ). median times to neutrophil and platelet (> , / l) engraftment were (range - ) and ( - ) days respectively. patients ( %) developed bk viuria (one with grade iii hemorrhagic cystitis); ( %) developed cmv viremia; and one patient ( %) had asymptomatic ebv viremia. there was no graft failure, sinusoidal obstruction syndrome or transplant related mortality. median follow up post-transplant was days (range - days). year probability of os and rfs were % and % respectively. hdc with ascr is a promising therapeutic strategy to consolidate remission and improve survival in select high risk soft tissue sarcoma patient subgroups. prospective clinical trials will inform the impact of disease status prior to hdc and ascr on outcome, optimal conditioning and long term relapse free and overall survival. background: absence of minimal residual disease is paramount for cure of pediatric acute lymphoblastic leukemia (all). the testis may harbor occult leukemia and this disease may result in treatment failure. objectives: the purpose of this study was to assess the longterm outcomes of boys with or without testicular leukemia pre-hematopoietic stem cell transplantation (hsct). design/method: retrospective analysis of boys with high-risk de novo ( with hypodiploidy all) or recurrent/refractory all was conducted. flow cytometry of bone marrow mononuclear cells was used to determine remission status. testicular evaluations were performed by physical examination and wedge biopsy pre-hsct. the median age at time of transplant was . years. all patients were in remission by flow cytometry of bone marrow mononuclear cells at the time of transplant and none had evidence of clinically apparent testicular disease. testicular leukemia was detected in patient and he underwent bilateral orchiectomy. he developed acute graft versus host disease (gvhd) of the duodenum and sigmoid colon which resolved, and the leukemia remains in second complete remission and he is free of hsct-related morbidity . months post-hsct. of the patients without testicular leukemia died a median of . months (range, . to . ) post-hsct ( with adenovirus infection and each with thrombotic microangiopathy and aspergillus pneumonia); experienced infection (staphylococcus species, corynebacterium, enterococcus, klebsiella, citrobacter, e. coli, epstein barr virus, adenovirus, bk virus, human herpesvirus- , candida albicans, fusarium, aspergillus, yeast, and other fungus); experienced gvhd ( of the gi tract, of the skin, of the liver, of the eyes, of the mouth, and of the lungs); and developed a second neoplasia (right lower leg leiomyosarcoma). one patient developed bone marrow minimal residual disease ( . % phenotypically abnormal cells detected months after / matched sibling hsct). reinduction therapy comprised weekly doses of rituxan, courses of blinatumomab and donor lymphocyte infusions with il- . two subsequent bone marrow evaluations were minimal residual disease negative. thirteen months post-hsct residual disease recurred ( . %) and he will receive inotumumab. overall median survival post-transplant of the boys is . months (range, . to . ) and of the surviving boys is . months (range, to . ). conclusion: testicular biopsy can detect occult leukemia pre-hsct. testicular leukemia pre-hsct does not appear to increase the risk of subsequent relapse or other hsct-related adverse events compared to those without it. yaya chu, nang kham su, sarah alter, emily k. jeng, peter r. rhode, mathew barth, dean a. lee, hing c. wong, mitchell s. cairo new york medical college, valhalla, new york, united states background: rituximab has been widely used in frontline treatment of b-nhl including burkitt lymphoma (bl), however, some patients retreated with rituximab relapse, which limit patient treatment options. novel therapies are desperately needed for relapsed/refractory b-nhl patients. several strategies for overcoming rituximab-resistance are currently being evaluated, including engineering immune cells with chimeric antigen receptors (car), as well as second-generation anti-cd antibodies. nature killer (nk) cells play important roles in the rejection of tumors. however, nk therapy is limited by small numbers of active nk cells in unmodified peripheral blood, lack of tumor targeting specificity, and multiple mechanisms of tumor escape of nk cell immunosurveillance. our group has successfully expanded functional and active peripheral blood nk cells (expbnk). b t m was generated by fusing alt- , an il- superagonist, to four single-chains of rituximab. b t m displayed tri-specific binding activity through its recognition of the cd molecule on tumor cells, activated nk cells to enhance adcc, and induced apoptosis of b-lymphoma cells. objectives: to examine if b t m significantly enhances the cytotoxicity of expbnk against rituximab-sensitive and -resistant bl cells. design/method: expbnks were expanded with lethally irradiated k -mbil - bbl and isolated using miltenyi nk cell isolation kit. alt- and b t m were generously provided by altor bioscience. nk receptors expression and cytotoxicity were examined as we previous described. ifng and granzyme b levels were examined by elisa assays. equal doses of rituximab, alt- , rituximab+alt- , obinutuzumab (obinu) were used for comparison. igg was used as controls. anti-cd car expbnk cells were generated as we previously described by mrna electroporation. rituximab-sensitive raji andresistant bl cells raji- r and raji- rh, were used as target cells. results: b t m significantly enhanced expbnk cytotoxicity against rituximab-sensitive raji cells, rituximab-resistant raji- r cells and resistant raji- rh cells compared to the controls igg, rituximab, alt- , rituximab+alt- , obinu (p< . , e:t = : ). furthermore, we confirmed the enhanced cytotoxicity by measuring ifn-g and granzyme b production. b t m significantly enhanced ifn-g and granzyme b production from expbnk against raji, raji- r and raji- rh compared to igg (p< . ), rituximab (p< . ), alt- (p< . ), rituximab+alt- (p< . ), and obinutuzumab (p< . ). when compared to anti-cd car expbnk cells, b t m + expbnk had the similar cytotoxicity against raji, raji- r and raji- rh as anti-cd car expbnk cells did (p> . ). conclusion: b t m significantly enhanced expbnk activating receptor expression and in vitro cytotoxicity against rituximab-sensitive and -resistant bl cells. the in vivo functions of b t m with expbnk against rituximab-sensitive and -resistant bl cells using humanized nsg models are under investigation. background: cardiac dysfunction, including left ventricular systolic dysfunction (lvsd), is a known complication in stem cell transplant (sct) survivors. while detection of lvsd by echocardiography is important in this population, there has been minimal research to determine if subclinical cardiac dysfunction exists in sct patients. cardiopulmonary exercise testing (cpet) is a valuable tool to assess cardiac function, and to determine how the heart responds to the stress of exercise. no studies have been performed to determine if sct patients with normal lvsd on standard echocardiography may have abnormal cpet. to determine the feasibility of cpet, as well as additional echocardiographic parameters, to detect dysfunction in sct patients with a normal ejection fraction on echocardiogram. design/method: we performed a cross-sectional analysis of sct survivors who were at least years post sct, years of age or older and with an ejection fraction > % (low end of normal range) on echocardiogram. we assessed the exercise capacity of all patients with cpet, and sub-clinical cardiac dysfunction through tissue doppler and strain analysis from the echocardiogram. results: seven patients ( male) have qualified and completed this study so far with an average age of . ± . years. the median time from transplant is . ± . years. all seven patients had a normal ejection fraction, however four patients had abnormalities on their cpet. these abnormalities included abnormal predicted peak oxygen consumption (vo ) ( %± . , normal > %) (the best predictor of functional capacity), predicted oxygen pulse ( %± . , normal > %) (measure of cardiac stroke volume) and ventilatory efficiency (ve/vco slope) ( ± . , normal < ). submaximal exercise data, used when patients are unable to complete a maximal effort test, demonstrated low-normal predicted vo at anaerobic threshold ( . %± . %, normal > % of was . days while patients who received autologous infusions had a mean number of days to engraftment of . . engraftment after hsct needs to be prompt to minimize duration of neutropenia and maximize survival rates . our data demonstrates that the infusion of hematopoietic stem cell products with a syringe or iv pump is an effective method of delivery for stem cell products and does not delay the time to engraftment. the median days to neutrophil engraftment was . days. this is comparable to data from the nmdp, which reports engraftment occurs within - days. the main limitation to this study was its small sample size due to the number of transplants done at our center. however, it does provide evidence to support that infusion of stem cell products via pump mechanism is a safe alternative to the infusion by gravity method in the process of the hematopoietic stem cell administration. johns hopkins all children 's hospital, st. petersburg, florida, united states background: leukemic relapse remains the most common cause of treatment failure after allogeneic hematopoietic cell transplant (allohct) for myeloid malignancies. most children who relapse post-allohct will die of their disease, making interventions to minimize this risk a high priority. objectives: to evaluate the safety and efficacy of posttransplant azacitidine for relapse prevention in children undergoing allohct for myeloid malignancy. design/method: we retrospectively reviewed the charts of children undergoing allohct for myeloid malignancies between february and november at johns hopkins all children's hospital. results: during the study period, children (ages to years, median ) underwent allohct for myeloid malignancies: de novo acute myeloid leukemia (aml), ; mixed phenotype acute leukemia, ; treatment-related aml, ; juvenile myelomonocytic leukemia with aml transformation, ; and myelodysplasia/aml, . thirteen were in first complete remission, were in cr or greater. most patients ( / ) received fludarabine/melphalan/thiotepa conditioning; received hla-identical related or unrelated donors, and received haploidentical bone marrow grafts with post-transplant cyclophosphamide. three patients never received planned azacitidine ( early relapse; early trm), leaving evaluable patients. azacitidine ( mg/m /dose for days, in -day cycles for up to cycles) was started at a median of days post-transplant (range - ). two-thirds ( / ) of patients received eight or more cycles. of five patients who stopped therapy early, only one was due to toxicity; other reasons included severe gvhd ( ), parental preference ( ), and relapse ( ). cycle delays occurred in patients, with a median cycles delayed per patient, mostly for mild myelosuppression with early cycles. no patient required blood product transfusion during therapy, but g-csf was used in three patients to maintain anc> / l. dose-modifications were made in patients (renal tubular acidosis, acute kidney injury, and myelosuppression). there were relapses ( %), two of which occurred in patients in cr , for a relapse incidence of % in patients in cr , with a median follow-up of months (range . to ). no patients who received azacitidine died of transplant-related mortality. conclusion: administration of azacitidine in children undergoing allohct for myeloid malignancies is safe and feasible, with most patients successfully receiving all planned cycles. toxicity was acceptable and there was no trm or secondary graft failure. despite the limitations of a small cohort, relapse incidence-particularly in patients transplanted in cr suggests a potential benefit in disease control that warrants investigation in follow-up studies. background: despite significant improvements in the success rate of hematopoietic cell transplantation (hct), graft failure remains an important complication in patients transplanted for severe aplastic anemia (saa). second allogeneic hct can salvage patients, but -year overall survival (os) rates have been reported as low as % . objectives: identify patients who developed dropping donor chimerism, graft rejection, and/or graft failure after first hct for saa, necessitating additional hcts or cellular boosts (defined as stem cell products infused without preceding chemotherapy), and evaluate treatment-related complications and os. with vod/sos with and without multi-organ dysfunction (mod) pubmed and embase databases were searched for "defibrotide and retrospective chart reviews; excluded publication types were: case reports (< cases); meta-analyses; reviews; animal, modeling, pharmacokinetic, chromatography, and adult-only studies; guidelines; articles; and letters. resulting reports were screened for exclusion criteria. full-text articles were then reviewed for eligibility. study characteristics of selected publications were summarized, and publications were categorized by patients' mod status. when necessary, additional data tables were requested. a random effects model was used for pooling data for efficacy. interstudy heterogeneity was assessed with cochran's q-test. percentage of total variation across studies due to heterogeneity (i ) was evaluated we quantified ∼ proteins in each sample. reproducibility for one donor at different time points children 's minnesota, minneapolis, minnesota, united states background: pediatric and young adult hodgkin lymphoma (hl) has five-year survival rates > %. chemotherapy required to achieve this rate is associated with a lifetime risk of cardiac deaths, second malignancies, pulmonary disease and infertility. as effective salvage therapy exists, outcomes may be improved by de-intensifying initial therapy to lessen toxicity.objectives: we piloted a regimen in low and intermediate risk hl patients using agents without known association to significant late effects. this retrospective chart review was approved by children's minnesota irb.design/method: the bvg(p) regimen incorporated bortezomib ( . mg/m day , , , ); vinorelbine ( mg/m day , ); gemcitabine ( mg/m day , ) every days and prednisone ( mg/m /dose bid x days). we treated newly diagnosed patients, ages - years, with non-bulk stage iia (n = ) or iib (n = ) hl. two patients received bvg and received bvgp with the addition of prednisone.results: newly diagnosed patients were all pet negative after the first or second cycle and remained pet negative at end of therapy, cycles. nausea was well controlled with -ht antagonists and scopolamine. pegfilgrastim was not necessary due to the high absolute neutrophil count nadir [median . and minimum . × /l]. there were no episodes of febrile neutropenia, infection or transfusion need. no patients experienced alopecia. one patient developed sensory neuropathy after the eighth dose of bortezomib that was controlled with gabapentin and a switch to subcutaneous bortezomib administration. of the five newly diagnosed patients, four remain in remission at , , , days; relapsed at previous disease sites at days and subsequently achieved remission with bvgp with the addition of brentuximab. this series provides early evidence to stimulate expansion of this pilot experience and subsequent multiinstitutional study leading to a randomized trial of bvgp and current chemotherapy for low and intermediate hl. st jude affiliate clinic at st francis hospital, tulsa, oklahoma, united states background: symptoms suggestive of morning hypoglycemia has been noticed in children receiving all chemotherapy. only few small studies looked at this therapy related complication. factors increase risk of hypoglycemia in all patients include accelerated starvation, steroid induced adrenal suppression, mercaptopurine therapy and prolonged fasting for procedures.objectives: to study the prevalence and risk factors for hypoglycemia during all therapy design/method: medical records of of children (up to years old) treated for all between - ( patients) were studied for evidence of morning hypoglycemia defined as blood sugar (bs) < mg/dl. statistical mean differences between the subgroups were analyzed with spss using a nonparametric mann-whitney u test.results: fifty two percent ( %) of patients developed hypoglycemia during all treatment, with an average of . episodes/patient. % were males and % females. almost / ( %) of patients with hypoglycemia were in maintenance phase of therapy. % of hypoglycemic episodes occurred in % of patients. majority of hypoglycemic episodes ( . %) occurred on the day of procedure when patients were fasting overnight. . % of hypoglycemic episodes occurred in children ≤ years, with . % in ≤ years. patients who developed hypoglycemia were significantly younger (mean age at time of diagnosis of all was . ± . at the hypoglycemia group versus the non-hypoglycemia ( . ± . ) p< . . no statistically significant difference was found regarding sex, or tpmt genotype. % of hypoglycemic children-all < years of age-presented with life threatening hypoglycemia symptoms including seizure and loss of consciousness. this study showed high prevalence of hypoglycemia during childhood all therapy. younger age, especially ≤ years, is associated with higher risk of hypoglycemia as well as life-threatening episodes. to decrease fasting hypoglycemia during therapy for childhood all, we recommend that children under the age of years receive bed time snack high in proteins and complex carbohydrates, and to get them up early the day of procedure to take clear sugary drink. hospital for sick children, toronto, ontario, canada ann & robert h. lurie children's hospital of chicago, chicago, illinois, united states background: childhood brain tumors are the most common solid malignancy and the leading cause of cancer-related mortality in children. the most aggressive type of pediatric central nervous system (cns) tumors is diffuse intrinsic pontine glioma (dipg). despite decades of clinical trials, there has been no substantial improvement with respect to therapeutic outcomes with most children eventually succumbing to the disease. research on adult high-grade gliomas has shown a targetable pathway through the inflammationinduced expression of indoleamine , dioxygenase (ido ) and its recognized ability to suppress the anti-tumor immune response. a limited understanding into the role of ido in pediatric central nervous system tumors serves as the foundation of this research project. furthermore, the integration of nanotechnology is a fundamental step for the investigation and targeting of ido . spherical nucleic acids (snas) composed of nanoparticles have been shown to transverse cellular membranes, exhibit stability in physiological environments, escape from degradation, and create precise targeting in brain tumors.objectives: the purpose of our project is to delineate the role of ido in pediatric dipg, and develop small inhibitory (si)rna oligonucleotides and snas aimed at therapeutically inhibiting the gene expression of immunosuppressive ido . our specific aims are to: ( ) confirm the gene expression ido in different human dipg cell lines; ( ) generate and characterize sirna oligonucleotides targeting human ido in vitro; and ( ) generate and characterize gold nanoparticles for targeted inhibition of ido .design/method: unique patient-derived dipg cell lines were grown in culture, stimulated with increasing concentrations of the proinflammatory cytokine, ifn , and analyzed for mrna levels. sirna specific to ido was transfected into cells. sna generation is in progress.results: ido is expressed in multiple human pediatric dipg cell lines. sirna targeting ido among exons and results in a significant decrease in overall ido expression by dipg cells. sna generation for targeting ido with improved penetration & stability is ongoing, with preliminary results demonstrating a robust ability to inhibit ido expression. the grim prognosis of children with dipg, the lack of effective therapies, and the expression of ido by human dipg cells emphasize the importance of developing the treatment capability to inhibit ido gene expression, as a excluded from this study. the remaining patients were analyzed using descriptive statistics.results: a total of patients meeting inclusion criteria were identified. of these, patients ( . %) received tonsillectomy alone, ( . %) underwent tonsillectomy and decreased immunosuppression, ( . %) received tonsillectomy and rituximab, and another ( . %) received tonsillectomy with additional therapy (including ebv-specific cytotoxic tlymphocytes, donor leukocyte infusion, and chemotherapy). of the patients who received tonsillectomy with or without a decrease in immunosuppression, all were diagnosed with high-grade lymphoma and achieved clinical remission following tonsillectomy with no evidence of relapse to date. on further analysis looking at ptld risk factors, all patients were under years of age, all received t-cell depleted grafts, and none had significant graft-versus-host disease (gvhd) at the time of ptld diagnosis. we have identified a population of patients with localized ebv ptld that achieved clinical remission with no evidence of recurrence following tonsillectomy, suggesting that tonsillectomy alone may be an adequate treatment for localized ebv ptld in a specific subgroup of patients. further analysis is needed to identify characteristics of this subgroup to determine which patients would be most likely to respond to this treatment. university of rochester, rochester, new york, united states background: malignant central nervous system (cns) tumors in young children have a poor prognosis and pose a significant therapeutic challenge. consolidation therapy with carboplatin and thiotepa was piloted in ccg- , cog acns , and cog acns with the goals of intensifying therapy and omitting or delaying radiation.objectives: to document outcomes for patients undergoing carboplatin/thiotepa consolidation with autologous stem cell rescue (ascr) and to demonstrate the feasibility and toxicity of this regimen.design/method: patients up to years old (median age: months) with malignant cns tumors treated at the university of rochester from - with at least one cycle of carboplatin ( mg/kg/day x days) and thiotepa ( mg/kg x days) followed by peripheral blood ascr were included in retrospective analysis. data were recorded on time to engraftment (defined by absolute neutrophil count (anc) recovery to > . × ^ /l), length of hospitalization, toxicity with each consolidation cycle, progression free survival (pfs) and overall survival (os). stem cell harvest data were also collected.results: eleven patients with malignant cns tumors ( atypical teratoid/rhabdoid tumor, primitive neuroectodermal tumor, glioblastoma multiforme, and pineoblastoma) received a total of cycles of carboplatin/thiotepa. of these, underwent stem cell harvest at our institution, with complications limited to procedure-related hypotension for patient with known autonomic instability, and catheter-associated deep vein thrombosis (dvt) for patient. four patients were in complete remission (cr) /status-post gross total resection, was in cr , and had residual tumor at the time of consolidation. nine patients received planned consolidation cycles, patient (of ) planned cycles, and patient of an anticipated cycles thus far. average time to engraftment for these cycles was . (+/- . ) days, with a mean hospital length of stay of (+/- . ) days. fever occurred in of cycles ( %); infectious toxicity included documented bacterial infection in cases (enterococcus faecalis bacteremia in , klebsiella pneumoniae in ). there were no regimenrelated deaths. with a mean follow-up of months, survivors have not yet completed all therapies, and patients have relapsed ( have died of disease). of the survivors, have been disease-free for > months. background: autologous hematopoietic stem cell transplantation (auto-hsct) has resulted in improved survival for patients with high-risk neuroblastoma. treatment intensification is however associated with greater complications. data on early infectious complications in low-and-middle income countries are limited.objectives: to review the early infectious complications following auto-hsct in patients with high-risk neuroblastoma.design/method: a retrospective chart review of pediatric patients with high-risk neuroblastoma who underwent auto-hsct at the american university of beirut medical center between and was conducted. infectious complications during the first days post-transplant were reviewed.results: forty-three patients ( males and females) with a median age at diagnosis of . years [range: . - . ] years underwent auto-hsct during the above-mentioned period. conditioning regimen consisted of melphalan, etoposide and carboplatin. all patients received antiviral and antifungal prophylaxis. median time for neutrophil engraftment was days [range: - ]. bacteremia and clostridium difficile infections occurred in ( %) and ( %) patients respectively. seven ( %) patients developed enterocolitis diagnosed by imaging, were adenovirus induced. cmv viremia was diagnosed in ( %) patients, of whom required treatment. varicella zoster reactivation, parvovirus viremia, toxoplasmosis encephalitis, bk virus cystitis ( patients) and central nervous system ebv related post-transplant lymphoproliferative disorder were diagnosed in different patients. there was no invasive fungal infection. sixteen ( %) patients have died, of whom died in the early post-transplant period, due to disease progression and ( . %) due to infectious complications. among the patients who died due to infection, developed toxoplasmosis encephalitis, developed severe enterocolitis, of which were adenovirus related. the mean igg level within one week post-transplant was lower in patients with clinically significant viral infection compared to others ( vs . mg/dl, p: . ). the mean igg level at the time of clinically significant bacterial infection was lower in infected patients compared to others ( . vs . mg/dl, p: . ). neither absolute lymphocyte count nor absolute neutrophil count at day post-transplant affected the incidence of clinically significant infections. our results show that the rate of infections during the early post auto-hsct period is higher than what has been described in developed countries and has a significant impact on mortality. prevention, early detection and improvement in the treatment is required to improve outcome. university of miami, miami, florida, united states background: allogeneic hematopoietic stem cell transplantation (allo-hsct) is a curative treatment for many malignant and non-malignant (bone marrow failure, immunodeficiency, or metabolic diseases) in pediatrics. despite advances in medicine, graft-versus-host-disease (gvhd) remains a significant cause of non-relapsed morbidity and mortality, specifically in those with malignant diseases.objectives: to highlight the complexity to acute gvhd management and seldom-described treatment approach. a year male with a history of high risk acute myeloid leukemia (aml) due to failed induction therapy. he received a matched ( / ) unrelated donor hsctmarrow product-conditioned with busulfan, fludarabine, and anti-thymoglobulin (atg). his post-transplant course was complicated by hhv- viremia, pres (prompting a change from prograf to cyclosporine), mucositis, and grade iii acute gvhd (skin s , gut s , liver s ) around post transplant day , which later morphed to ocular involvement by d+ . he was started on mg/kg steroids with good response but flared up with each attempt to taper steroid dose. a course of rituximab and later atg were tried without success in weaning off steroids. switching cyclosporine to sirolimus did not provide any additional benefit either. extracorporeal photopheresis (ecp) was started times a week. he initially responded well, yet was not able to wean off steroids. in addition, he developed a flare when ecp session was reduced to days per week. ecp was therefore increased to days per week, which appeared to stabilize skin lesions. a trial of weekly methotrexate was attempted to wean off steroids and photopheresis, which provided no response. finally, a trial of bortezomib on days , , , and of a day cycle as published in a case series of multiple myeloma patients who developed post hsct gvhd. skin lesions improved remarkably however dose had to be reduced due to related pancytopenia. given the response to therapy, he was continued on a weekly dose of bortezomib, receiving a total doses, which has permitted the slow taper of prednisone that has since been discontinued without a major flare. he however is currently maintained on ecp times per week, which is now been slowly withdrawn.conclusion: management of acute gvhd in pediatric patients after hsct can be challenging with no definite options for those who fail steroids or become steroid dependent after initial response. in these situations, bortezomib could be a valid therapeutic option. background: neuroblastoma (nbl) is the second most common solid tumor in children and despite recent treatment advances, overall survival for high risk nbl remains < %. the addition of immunotherapy has improved survival and includes anti-gd antibody therapy. the success of antibody therapy in neuroblastoma is primarily due to natural killer (nk) cell mediated antibody dependent cellular cytotoxicity. we previously demonstrated that nk cells from patients with high risk nbl can be successfully isolated and expanded to large numbers and exhibit potent anti-tumor effects against nbl ( ). thus, infusions of autologous expanded nk cells in high risk nbl in combination with anti-gd antibody are being studied in clinical trials. toll-like receptors (tlr) present on the surface of leukocytes are responsible for pathogen recognition, and activation of these receptors stimulate the production of cytokines that critically link innate and adaptive immune responses. the tlr agonist, poly(ic) is a synthetic analog of dsrna that has previously been shown to directly stimulate cytokine production and improve cytotoxicity in primary nk cells through activation of genes regulated by interferon-response elements (ire) ( ). we hypothesized that ex vivo activation of tlr pathways in nk cells during our normal -day expansion using k feeder cells expressing membrane bound il- would enhance their function.design/method: nk cells were isolated from peripheral blood mononuclear cells and expanded with our previously described expansion protocol in media containing il- and ug/ml poly(ic) ( ). at the end of the -day expansion, nk cells expanded with poly(ic) were compared to controls using a calcein cytotoxicity assay to measure cytotoxicity against high risk neuroblastoma and cytometric bead array to measure cytokine production. : surprisingly, the addition of poly(ic) during nk cell expansion did not improve proliferation, cytokine production or cytotoxicity compared to our standard expansion method. rnaseq demonstrated that our standard expansion method results in a modest decrease in tlr expression at the transcriptional level, but significant upregulation of several ireregulated genes. we conclude that either our standard approach interferes with tlr signaling or saturates the innate immune response pathway such that co-stimulation with poly ic does not produce an additive effect. we are performing expression analysis on nk cells receiving poly(ic) during expansion to further explore this hypothesis. background: gonadal dysfunction leading to infertility is a complication after hematopoietic stem cell transplant (hsct). anti-müllerian hormone (amh) is a marker of ovarian reserve; it is not controlled by gonadotropins and has minimal inter-cycle variations, therefore, it can be used as a marker of ovarian reserve and aid in fertility counseling.objectives: assess ovarian reserve in hsct patients utilizing amh levels. background: tgf beta is an immune suppressive cytokine frequently elevated in the tumor microenvironment causing tumor immune evasion. acute tgf beta treatment potently inhibits nk cell cytotoxicity, cytokine secretion, and proliferation. however, tumor infiltrating nk cells receive chronic inhibitory tgf beta signals in conjunction with activating signals from tumor cells. objectives: to this end, we hypothesized that long-term tgf beta-cultured nk cells would induce functional and phenotypical changes on nk cells that differ from short-term tgf beta treatment.design/method: to explore this, primary human nk cells were cultured with the leukemia cell line, k , alone or with exogenous tgf beta for weeks. : surprisingly, nk cells cultured in tgf beta proliferated faster, and upon challenge with a variety of cell line targets they secreted much greater quantities of ifnÎ ( -to -fold increase against / cell lines) and tnf ( -to -fold increase against / cell lines). further, the high cytokine secretion induced in these nk cells was no longer inhibited by adding additional tgf beta. degranulation was also increased ( / cell lines), however cytotoxicity was not enhanced in a -hour cytotoxicity assay. after resting in il- , the cytokine hypersecretion of tgf betacultured nk cells was maintained for several weeks suggesting this functional change might involve cellular reprogramming. we investigated the mechanism behind these functional changes and profiled genes involved in tgf beta signaling. we found significant reduction of smad transcription which corresponded to a striking decrease in smad chromatin accessibility. we also found significantly increased smad and decreased tgfbr expression. phenotypic analysis revealed that tgf beta also induced remodeling of the nk receptor repertoire with decreased nkp , cd , and klrg and upregulation of trail. the functional consequences of these tgf beta-induced changes on in vitro and in vivo nk cell function are currently under investigation. background: the use of t-cell depleted grafts in haploidentical stem cell transplantation (hsct) has been associated with a delay in early t-cell recovery which increases the risk of viral infections, relapse or graft rejection. conventional donor lymphocyte infusion (dli) after hsct transplantation is effective but conditioned because of a high prevalence of gvhd. the infusion of selected lymphocyte subpopulations with low aloreactivity is emerging as an effective strategy to rectify this issue. the depletion of cd ra+ naive lymphocytes, preserving cd ro+ memory t-cells, could provide a safe source of functional lymphocytes with anti-infection, antileukemic and anti-rejection properties, and lower rates of adverse effects. our objective is to present data of patients that have received cd ro+ memory t-cells dli (mdli) and assess its safety and outcome. we present data of mdli performed after hsct in cases of mixed chimerism, persistent lymphopenia, viral/opportunistic infections or as a strategy to accelerate immune reconstitution.results: fifteen patients with diagnosis of all (n = ), aml (n = ), mds (n = ), saa (n = ), sideroblastic anemia (n = ) and cgd (n = ), received mdli after hsct. a total of forty-three mdli were infused. the median dose of cd ro+ memory t-cells infused was . × /kg (range: . × - . × /kg), with a median dose of cd ra+ naive t-cells of . × /kg (range: - . × /kg). the mdli were infused at a median of seventy-seven days after hsct (range: - days), with a median interval between mdli of thirty-four days results: eight published studies reported survival outcomes for pediatric vod/sos patients (n = ), across all defibrotide doses. estimated day+ survival ( % confidence interval) was % ( %- %). for vod/sos with mod, studies were identified (n = ) with pooled estimated day+ survival of % ( %- %). only one openlabel expanded-access study, the treatment-ind, reported outcomes separately for pediatric vod/sos patients without mod (n = patients aged ≤ years). the day+ kaplan-meier estimated survival for those patients was % ( %- %). safety results were not pooled due to differences in reporting methodology; however, study results were consistent with the safety profile of the phase historicallycontrolled trial in vod/sos patients with mod ( % pediatric), in which / defibrotide-treated patients and all controls experienced ≥ ae. hypotension was the most frequent ae ( %, defibrotide; %, controls); common hemorrhagic aes (ie, pulmonary alveolar and gastrointestinal hemorrhage) occurred in % of defibrotide-treated patients and % of controls. in this pooled analysis of studies with defibrotide-treated pediatric patients with vod/sos, estimated day + survival was % (without mod, %; with mod, %). safety results in individual studies were generally consistent with the known safety profile of defibrotide. taken together, these results show a largely consistent defibrotide treatment effect in pediatric patients treated with defibrotide for vod/sos, with or without mod. results: six patients met inclusion/exclusion criteria. all patients were started on ecp while concurrently receiving . to mg/kg steroid therapy for agvhd plus a calcineurin inhibitor. patients had initiation of ecp within a maximum of weeks from initial diagnosis of agvhd (range - days). patients had grade - agvhd ( / patients with grade ) with skin, liver, and gi gvhd represented. patients received ei-ecp - times per week for the first weeks and then had ei-ecp frequency tapered based on initial response.after weeks of therapy patient had a decrease in overall gvhd grade by grade. all patients were able to have steroids tapered, with doses decreased by an average of % ( % - % decrease).at weeks of therapy, one patient with grade agvhd died of mof associated with infections. three patients had complete resolution of agvhd and patients decreased by grade. steroid doses were decreased by an average of % ( % - % decrease). continuously measures axillary temperature and wirelessly transmits real time-time data. the primary aim of the study was to evaluate the feasibility, safety and tolerability of continuous temperature monitoring in hsct patients using temptraq. we are performing a prospective observational study of pediatric patients ( - years of age) undergoing hsct at cincinnati children's hospital in cincinnati, ohio. enrolled patients wore a temptraq patch for days. a - rating scale survey was completed by the parent/guardian at the end of the study to determine tolerability, ease of use, satisfaction and desire for future use in the inpatient and outpatient setting. temperature data from the temptraq patch was compared to the standard episodic temperature monitoring to determine detection of febrile episodes. seven of ten patients have completed screening. we anticipate completion of the study in early february. the temptraq patch was well tolerated by study subjects (mean tolerability rating of . / ). one patient developed skin breakdown at the site of the temptraq patch attributed to recent thiotepa. the patch was easy to apply with an easy of application rating of . / . parents were overall satisfied (rating . / ) and would like to use the temptraq patches in future hospitalizations (rating . / ) and at home (rating . / ). temptraq patch identified fever (≥ . • f) in patients. the fever was never detected by episodic monitoring (soc) in patients and significantly delayed in the other patients (> hours). temptraq was well tolerated in pediatric hsct patients. timely fever detection was improved in temptraq over the current soc. background: serotherapy is commonly used in patients undergoing hematopoietic stem cell transplant (hsct) to reduce the incidences of engraftment failure and graft versus host disease. however, one well-known side effect is fever. as children undergoing hsct have compromised immune defenses, fever may also be an early indicator of bloodstream infection, which would warrant prompt use of broad-spectrum antibiotics. in a subset of patients with serotherapy-associated fever, antibiotics, which may induce antibiotic resistance and increase costs, may be unnecessary. we aimed to determine the incidence and characteristics of serotherapy-related fever, as well as the likelihood of concomitant bacteremia, in our institutional experience. a -year retrospective chart review was conducted of pediatric patients who received serotherapy as part of hsct conditioning at the university of minnesota. one-hundred sixty eight consecutive hsct patients who received serotherapy -either atg (n = ) or alentuzumab (n = ) -were identified. the median age at hsct was -years (range, . - years). a total of patients ( %) developed fever while on serotherapy (atg = , alentuzumab = ). one-hundred sixteen patients presented fever following the first infusion, and the median onset of fever was hours after commencing infusion (range, . - hours). fever resolved at a median hours (range, - hours). one hundred and fourteen patients ( %) underwent blood cultures. only seven patient were not started on ( %) empiric antibiotics, while % (n = ) were on antibiotic treatment prior to serotherapy for previously known or suspected infections. nine patients ( % of febrile patients, % of all patients) had positive blood cultures (atg = ; alentuzumab = ). no infection-associated deaths were observed.conclusion: while fever is common during serotherapy conditioning in children undergoing sct, episodes of concomitant bloodstream infection are rare. ongoing analysis identified potential risk factors for bacteremia as recent history of infection, first episode of fever following second or subsequent infusions, and previous central line placement. further analysis is being conducted to identify subgroups of patients for whom close monitoring alone may be safe. background: hsct is potentially curative for caya with high-risk leukemias; however, most lack an hla-matched aspho abstracts related donor. the risk of gvhd is increased with unrelated (urd) or partially matched related (pmrd) donors. selective t-cell depletion based on the elimination of t cells carrying and chains of the t-cell receptor may greatly reduce the gvhd risks, while allowing the maintenance of mature donor-derived alloreactive nk cells and / (+) t cells, which may augment the anti-leukemia effect.objectives: this is a prospective study of caya with acute leukemia who underwent hsct with mmrd or urds and tcr / /cd depletion. outcomes included engraftment, toxicities, viral reactivation, and relapse.design/method: this study included caya with acute leukemia transplanted between october and may . all received a myeloablative preparative regimen with targeted busulfan (n = ) or tbi ( cgy/ fractions) (n = ), with thiotepa ( mg/kg) and cyclophosphamide ( mg/kg). atg ( mg/kg x ) was given to those receiving haploidentical grafts and to the first who received urd grafts. immune suppression was not given post-hsct. the stem cell source was mobilized peripheral blood stem cells (pscs), which then underwent tcr / /cd depletion utilizing the clinimacs device under gmp conditions in the chop cellular immunotherapy lab.results: median age was (range . - . ). diagnoses included all ( -b-cell, -t-cell) and aml ( ; -secondary aml). urd were used for ; were / allele matched and were / matched. haploidentical donors were used for . median cd (+) dose - . × , / (+) cd (+) cells - . × , and b cells - . × . all patients achieved an anc at a median of d+ ( - ), and % had platelet engraftment at median d+ ( - ). nine patients ( %) developed acute gvhd (all skin, grades i-iv). five developed chronic gvhd (skin, gut, lung): limited in , extensive in . viral reactivations included: adenovirus ( , %), bk virus ( , %), cmv ( , %), and hhv ( , %). nine ( %) patients relapsed at a median of days (range - ) post-hsct, including aml patients ( . %) and all patients ( . %). transplant-related mortality was %; causes included sepsis ( ) and ards ( ). os was %; efs was % (gvhd-free efs %, lfs %). hsct with tcr / /cd depletion demonstrates excellent engraftment kinetics with limited gvhd without immune suppression. elimination of post-hsct immunosuppression may offer an excellent platform to augment anti-leukemic immune therapy or to enhance immune reconstitution. background: hematopoietic cell transplantation (hct) is the only curative treatment available for patients with sickle cell disease (scd). low bone mineral density (bmd) has been described in scd, but little is known about the impact of curative hct on this outcome. to determine the prevalence of low bmd and variables associated with low bmd in scd patients after hct. we conducted a retrospective chart review of scd patients who underwent hct at children's healthcare of atlanta (choa) between / and / and survived ≥ year post-hct. transplant characteristics, post-hct dual-energy x-ray absorptiometry (dexa) scan results, vitamin d levels, graft-versus-host-disease (gvhd) status, and fsh levels were reviewed. for patients - years of age, height corrected z-scores were calculated using a nihvalidated calculator, with t-scores used for older patients. bmd was categorized as low if between - and - sd below the mean and clinically significantly low if >- sd, in accordance with the children's oncology group long-term follow-up guidelines. vitamin d levels < ng/mol were considered deficient, and fsh levels > miu/ml suggestive of premature ovarian failure. fisher's exact test was used to compare variables in those with normal versus abnormal dexa scan results, with p< . considered significant.results: hct was performed on patients with scd, with surviving ≥ year post-hct. dexa scans were obtained in patients ( % female), with mean time from hct to dexa scan being years ( . - . years) and mean age at time of dexa . years ( . - . years). patients with and without dexa scans did not differ by sex, donor source, age at transplant, or vitamin d status. low bmd was noted in patients ( . %), with these patients more likely to be > years (pubertal; . versus . %, p = . ). acute gvhd was more common in patients with low bmd ( . versus . %), but not statistically significant (p = . ). clinically significant low bmd was noted in patients ( . % of those with dexa scans). these patients were older ( . years at testing), were more likely to be male ( . %), and all had acute and chronic gvhd, while none had evidence of gonadal failure.conclusion: clinically significant low bmd is uncommon after hsct for scd. patients at risk for low bmd include older patients and likely those with gvhd. this preliminary data suggests routine dexas may not be indicated for all patients who undergo hct for scd, but further data is needed. background: causes of renal dysfunction after hematopoietic cell transplantation (hct) include damage from radiation, nephrotoxic medications, graft vs. host disease (gvhd), hepatorenal syndrome, viral infections, or transplant associated microangiopathy. we sought to investigate the incidence of, and risk factors for, acute kidney injury in pediatric hct patients and associated risk with mortality.design/method: data from patients who underwent hct between and at a single institution were sequentially retrospectively captured on irb approved protocol. acute kidney injury (aki) was defined at multiple time points post-hct using the standardized criteria: kidney disease: improving global outcomes (kdigo). interval differences between values were analyzed using wilcoxon rank sum testing and categorical variables were analyzed using chi-square analysis.results: ninety-eight patients were included in the study: allogeneic (n = ) and autologous (n = ), mean age . years, of whom % were african american, % asian, % caucasian, % latino, and % mixed race. forty-seven percent of patients developed aki within the first years of hct. increased risk for aki was associated with a lower pre-transplant creatinine level (p = . ), abnormal pretransplant bun (p = . ) and an unrelated donor (p = . ) while preparative regimen intensity, race, or primary disease were not. twenty-six percent of patients developed aki within days of hct. of those with aki, % were exposed to either cidofovir, aminoglycosides, and/or ambisome for at least days versus % without aki and % were exposed to vancomycin compared to % without aki. evaluating outcomes at year after hct, of those with stage aki: % had reduced gfr and % died, while % had reduced gfr and % had died for patients with aki stage or . the absence of aki by day was associated with % reduced gfr and % death at -year after hct. overall, those with aki at any time in the first year post-hct had a . fold increased risk of death compared to those without. for patients who required renal replacement therapy (rrt, n = ), the risk of death was . fold greater compared to those who did not. in the % of patients who survived rrt, both recovered renal function within years.conclusion: acute kidney injury is common after pediatric hct, and may be associated with low creatinine, abnormal bun, unrelated donor pre-hct, and renal toxic medications. early-onset aki post hct is associated with an increased risk of mortality. these data should be validated in a larger prospective study but may offer opportunities to intervene and enhance outcomes. background: myeloablative hematopoietic stem cell transplant (hct) for pediatric malignant disease is associated with significant morbidity with % patients experiencing mucositis. patient controlled analgesia (pca) utilizing opioids is an effective strategy for pain management. we sought to describe and analyze pca use in d+ days post myeloablative hct for malignancies at lurie children's hospital of chicago from - .design/method: utilizing retrospective chart review, pca details were collected: indication, initiation day, pca duration, team managing pca (anesthesia or palliative), medication and dose in morphine equivalents, and pca toxicities. efficacy of pca was evaluated on pca day + , + , + , + using demands %, maximum pain score (rflacc, faces, vas) and subjective patient, parent and/or pain team perception of pain control. we devised a scale based on the above to designate pain control as good, moderate or poor. variables being analyzed include recipient age, sex, donor type, source, diagnosis, tbi use, gvhd/trm. this analysis, we analyze the depth of remission, car-t persistence, and post-transplant gvhd on our phase i anti-cd car-t protocol (nct ) to better understand the role of car-t in the peri-hct setting.design/method: children and young adults with relapsed/refractory cd + all treated on our phase i anti-cd car-t protocol were analyzed. mrd was assessed by flow cytometry (fc) in all, with pcr-based mrd analysis using igh or tcr testing assessed in select patients. hcts were performed at each patient's local institution based on standard of care and included varying conditioning regimens, donor types, stem cell source, and gvhd prophylaxis.results: on our cd car trial, patients were treated, the majority of patients (n = ) having relapsed following a prior hct. / patients ( %) attained a cr, of whom were mrd negative by fc. concurrent pcr based mrd analysis available in patients demonstrated that all patients achieved pcr based negativity. in , this was simultaneous with the month mrd negative fc, and in , pcr negativity was achieved over time (fc remained negative). patients proceeded to hct at a median time of days (range: - days) post-car-t, which was a first hct in . these two patients remain in an mrd negative cr, year post-car-t. no patients developed acute or chronic gvhd. car persistence was seen in patients who had detectable car-t cells on the pre-hct marrow suggesting the possibility of ongoing anti-leukemia surveillance prior to initiation of the conditioning regimen.conclusion: by inducing pcr negativity, car-t therapy may have a synergistic role with hct to improve leukemia free survival, prior to emergence of antigen negative leukemia, without an increased risk of gvhd. while the sample size is small, car-t therapy may offer an effective bridge to hct, particularly for those who are pcr negative, and those who have not had a previous transplant. given the underlying risk of hct related trm, pre-hct car may potentially allow for hct conditioning de-intensification as it may not be needed to eradicate residual disease. lee dw, ash abstract , background: post-transplant lymphoproliferative disease (ptld) is a complication after solid organ transplantation (sot) that is frequently due to epstein -barr virus (ebv) as a decrease in ebv-specific t cell immunity due to immune suppression allows for uncontrolled proliferation of ebv-infected b cells. outcomes for ptld are suboptimal with relapse rates approaching %. however, ebv-infected b cells in ptld express the ebv antigens lmp and lmp that can be targeted with immune therapy.objectives: we hypothesize that third party "off the shelf" lmp-specific t cell products may improve outcomes and decrease associated co-morbidities for patients with ptld by not only target the lymphoproliferating ebv-infected b cells but also restoring ebv-specific immunity.design/method: lmp-specific t cells (lmp-tcs) are manufactured from eligible donors with a broad range of hla types in our gmp facility to be used in a children's oncology group (cog) trial (anhl ) for patients with ptld after sot. lmp-tc products are manufactured from healthy donors using autologous monocytes and lymphoblastoid cell lines (lcl) transduced with an adenoviral vector expressing Δlmp and lmp as antigen presenting cells. lmp-tc products undergo comprehensive characterization by ifn-elispot assay to determine lmpspecific epitopes, class i and/or ii response, and hla restriction to guide selection of lmp-tc product for each patient.results: thus far, lmp-tc products have been manufactured. lmp-tcs were active against lmp (mean: sfu/ × ^ cells; range: - ), lmp ( ; - ), and lcl ( ; - ) as determined by ifn-elispot assay. at the time of cryopreservation, the lmp-tc products comprised a mean of % cd + t-cells, % cd + t-cells, and % nk cells. no b cells or monocytes were detected in the final products. thus far, we have identified novel lmp epitopes (lmp specific: n = ; lmp specific: n = ). approximately % of the lmp-tc products have lmp-specific activity through multiple hla alleles, and % have a mixed class i and class ii response. conclusion: thus, lmp-specific t cell products can be expanded from healthy donors to creat a third party bank, and identifying epitopes and hla alleles with lmp activity will facilitate selecting the most appropriate product for patients. while lmp-specific t cells have previously demonstrated safety and efficacy in phase i studies, anhl is the first trial using cellular therapy within a cooperative group setting. children's cancer hospital at the university of texas md anderson cancer center, houston, texas, united states background: in , the united states food and drug administration (fda) approved the first chimeric antigen receptor t cell (car-t) therapy; tisagenlecleucel. this cd -directed genetically modified autologous t cell immunotherapy has shown response rates of almost % among children and young adults with b-cell precursor acute lymphoblastic leukemia (all) that are refractory or in second or later relapse. cytokine release syndrome (crs) and car-t cell related encephalopathy syndrome (cres) are well described toxicities associated with car-t therapy. crs is a systemic inflammatory response and is typically characterized by fever, hypoxia, tachycardia, hypotension and multi-organ toxicity. cres may occur concurrently or following crs, or without any associated crs symptoms and is characterized by encephalopathy, delirium, seizures and rarely cerebral edema. almost half of patients who receive tisagenlecleucel may require pediatric intensive care unit (picu) support. crs and cres are generally reversible but may be associated with fatal outcomes. pediatric specific management guidelines, comprehensive training of multidisciplinary staff, effective communication and phased infrastructure ensure that adequate resources are available to facilitate early diagnosis and appropriate management of pediatric patients with crs and cres and allow for optimal patient outcomes and accreditation by the foundation for accreditation of cellular therapy (fact).objectives: develop a comprehensive program to ensure safe administration of immune effector cell (iec) therapy to pediatric patients.design/method: an inter-disciplinary pediatric cartox (car t cell therapy associated toxicity) committee consisting of cell therapy and picu physicians, neurologists, fellows, nursing leadership, advanced practice practitioners, pharmacists, registered nurses and social workers was created to monitor patient toxicity and establish specific clinical guidelines and diagnostic and treatments algorithms for pediatric patients receiving iec therapy. educational modules were developed as (i) live in-services and (ii) an online module with a competency based assessment. electronic medical record (emr) order sets and documentation and warning systems were also developed by the committee. the pediatric cartox committee developed a diagnostic and treatment algorithm for patients receiving iec therapy. emr orders and flowsheets were developed to support adherence to the algorithm. inter-disciplinary staff training and competency assessments were closely tracked. almost % of identified staff have completed training and achieved competency including, pediatric cell therapy staff, emergency center, picu, outpatient clinic/triage, neurology and sub-specialty staff and nocturnalists.conclusion: an inter-disciplinary approach can assist in institutional readiness for an iec program, promote quality assurance and perhaps fact iec accreditation. future directions include a program for ongoing staff competency assessments. predicted peak vo ) and abnormal oxygen uptake efficiency slope at the anaerobic threshold ( . ± . . , normal ± ). additionally, on echocardiogram three patients had evidence of diastolic dysfunction as evidenced by an elevated e/a ratio ( . ± . ) on tissue doppler. three patients demonstrated depressed longitudinal peak systolic strain (- . ± . ), indicating dysfunction not captured by ejection fraction. in this feasibility study, sct patients without evidence of lvsd on standard measures by resting echocardiogram can demonstrate abnormal exercise capacity. additionally, they can demonstrate systolic and diastolic dysfunction by measures not always included in standard echocardiography. these data suggest the need for a more thorough screening of survivors, and will be further validated as additional patients are recruited for this study. background: in hematopoietic transplantation, the t lymphocytes of the inoculum play a determining role in promoting hematopoiesis, transferring immunity to pathogens and acting as mediators of the graft-versus-leukemia effect (gvl). however, they are also responsible for graft-versus-host disease (gvhd), the main cause of post-transplant morbidity and mortality. the depletion of cd ra lymphocytes, by eliminating naive t lymphocytes from the inoculum, aims to conserve the gvl without producing gvhd.design/method: since april , patients ( boys and girls), with a median age of years, have undergone an allogeneic hematopoietic transplant from an hla donor identical with cd ra/cd depletion. the indication for transplant was: acute lymphoblastic leukemia ( ), acute myeloblastic leukemia ( ), myelodysplasia ( ) and medullary aplasia ( ). the donor was familiar in cases and unrelated in . the conditioning regimen was with fludarabine, busulfan and thiotepa. the median of cd + cells infused was . × / kg. on the day , + and + a programmed infusion of × / kg lymphocytes cd ra-was performed.results: all the patients grafted with a median leukocyte (> . × / l) and platelet (> × / l) engraftment time of and days, respectively. only one patient has developed acute gvhd grade i and no patient has developed chronic gvhd. immune reconstitution was early and rapid in all t cell subsets no patient has relapsed so far and only patient with myelodysplasia has developed an aml. she has received a nd transplant and has died of relapse. there was no case of toxic mortality. the event-free survival (sle) was ± % with a median follow-up of months. at present, patients are alive, out of immunosuppressive treatment and doing well. allogeneic transplantation with cd lymphocytes ra depletion resulted on very encouraging results, with a very low incidence of acute and chronic gvhd, but preserving the gvl effect by infusing cd ra-donor lymphocytes. miami children's health system, miami, florida, united states background: hematopoietic stem cell transplantation (hsct) using autologous or allogeneic progenitor cells is a potentially curative treatment for patients with high-risk malignancies and nonmalignant conditions. the american society for blood and marrow transplantation developed a task force to establish consensus guidelines for defining patient care in hsct and advocated for further studies to delineate safe procedural steps as an increasing amount of hsct are being offered to patients. there is limited evidence to support engraftment in recipients who receive their infusions via iv or syringe pump. we present novel data from patients who achieved neutrophil engraftment following hsct by a pump mechanism.objectives: to provide evidence supporting the use of pump (intravenous or syringe) infusion method in hematopoietic stem cell transplantations.design/method: a retrospective review was completed for patients who underwent hsct between and . inclusion criteria included patients who had received hematopoietic stem cell transplants between and and who were ages months to years old. the main outcome measure was days to neutrophil engraftment (defined as the first of three consecutive days with an anc > × /l).results: among patients who received infusion of hematopoietic stem cell products via pump mechanism, patients ( . %) received autologous products and ( . %) received stem cells from allogeneic donors. neutrophil engraftment (anc > × /l) occurred in a median of . days after stem cell infusion. the mean number of days to engraftment for patients who received allogeneic infusions s of s design/method: a retrospective chart review was performed at the children's hospital of wisconsin. statistical analyses included kaplan-meier estimate for os, mann-whitney test for comparing outcomes between subjects, and descriptive analyses.results: from - , patients with a median age of . ( . - . ) years at st hct were identified. patients were conditioned with cy/atg (n = ), cy/flu/atg (n = ), or cy/flu/atg/tbi (n = ) and received marrow (n = ) or cord blood (n = ) with median cd /kg dose of . ( . - . ) × . two patients developed grade i acute graftversus-host disease (gvhd); none developed chronic gvhd. due to dropping chimerism, graft rejection, or graft failure, nd hct (n = ) or boost (n = ) was offered. the median cd chimerism prior to hct/boost was ( - )%. median time between st hct and nd hct or boost was days ( days- . years). in patients receiving nd hct, used the same donor, of which used the same stem cell source (marrow) and switched to peripheral blood stem cells (pbsc). in patients who switched donors, used pbsc and used cord blood. most patients receiving nd hct underwent a uniform conditioning regimen of cy /flu /equine atg/ gy tli (n = ) or cy /flu /rabbit atg/ gy tbi (n = ); one received cy/atg. acute and chronic gvhd (limited seen in %) developed in % and % of patients, respectively. four patients required additional boosts and additional hct. after final intervention, cd and whole-blood chimerism at last follow-up was between - % (n = ) and - % (n = ), respectively. with a median follow-up of . ( . - . ) years, of patients are alive with an estimated -year os of . ± . %, having performance status ≥ % (n = ) or % (n = ). one patient developed chronic extensive gvhd and died of fungal infection . years after nd hct. our single-center experience demonstrates excellent ability to salvage patients who develop graft failure after initial hct. transplant-related complications such as gvhd and infections remain significant concerns. key: cord- -ezrn cva authors: nan title: physicians – poster session date: - - journal: bone marrow transplant doi: . /bmt. . sha: doc_id: cord_uid: ezrn cva nan hematopoietic stem cell transplant unit, hematology department, hospital universitario de donostia, donostia/san sebastián and informatics and automatics department, university of salamanca, spain the immature platelet fraction (% ipf) is a relatively new parameter that measures young (reticulated) platelets in peripheral blood (pb). ips rise as bone-marrow (bm) production of platelets increases. several clinical utilities of the %ipf have been already proved, as the treatment response monitoring in aplastic anemia or immune thrombocytopenic purpura. in this study, we aimed to found if ip measurement might be useful during the grafting phase of hsct. this study includes patients who underwent allo-hsct in our center during the last . years. were male ( %) and female ( %). median age was years (range: - ). baseline diseases were: acute leukemias ( ), lymphoproliferative disorders ( ), myelodysplastic syndromes ( ), chronic myeloproliferative diseases ( ), multiple myeloma ( ) and bone marrow failures ( ) . donor was unrelated in cases, and related in (including haplo-identical). conditioning regimen was: busulphan-based ( ), melphalan-based ( ), tbi-based ( ) and others ( ) . progenitors source was pb in , and bm in . platelet count, %ipf and absolute ip count (aipc) from day + to the day of stable graft were analyzed. . % patients reached plat ⩾ /mcl at day + , . % at day + and . % at day + . median first day of plat ⩾ /mcl was day + (range: . median %ipf was . % (range: - . ), . % (range: - . ) and . % (range: - . ) at days + , + and + , respectively. median aipc was /mcl (range: - ), (range: - ) and (range: - ) at days + , + and + , respectively. among the time points analyzed, aipc at day + showed the best positive correlation with platelets counts at day + (r = . ). interestingly, patients with lower aipc at day + showed a delayed platelet graft (see table ). contrarily, patients with higher aipc at day + had an earlier platelet graft. absolute immature platelet count before the graft seems to predict the precocity of the platelet graft for the majority of patients undergoing allo-hsct. this finding might help physicians for the patient management (anticipation of hospital discharge and so on). disclosure of conflict of interest: none. [p ] p analysis of genetic polymorphism for cardiovascular diseases (cvd) in placental and maternal blood in hypertension and hypercholesterolemia c khalil , a azar and a ibrahim , reviva stem cell research and application center, lebanese university, middle east institute of health hospital and faculty of medical sciences, lebanese university, lebanon cardiovascular diseases are the world's leading cause of death representing % of the total global mortality. the genetic polymorphism of the cvd genes, especially the ace: angiotensin converting enzyme gene risky alleles (ins/del) which are associated with a high and inappropriate level of ace can be considered as a genetic model in the development of hypertension and its complications in cvd. we evaluated the mutation impact of the cvd genes in the lebanese population, based on samples derived from placental blood (pb) and samples derived from peripheral blood of postpartum mothers. adult females (age ⩽ years) were divided (n = per group) into group (normotensive, normocholesterolemia: nn), and group (hypertension, hypercholesterolemia: hh). buffy coat were extracted from the pb. all tests on pb and maternal blood were done by using the test strip assay to identify the most relevant genetic variations to estimate the risk for cvd. the presence of a double mutation (ins+/del+) related to the ace gene in the hh group was %. the presence of a single mutation (ins − /del+) was only associated to the hh by %. (ins − /del − ) was absent in % of the pb and nn. despite the presence of double mutation ins/del for cvd in maternal blood, pb was free of this mutation. therefore, beyond genetic mutations, other factors can play a major role in the occurrence of cvd. disclosure of conflict of interest: none. s b e mt automated red blood cell depletion in abo incompatible grafts in the pediatric setting c del fante, l scudeller, s recupero, g viarengo, f compagno, m zecca and c perotti fondazione irccs policlinico san matteo red blood cell (rbc) depletion by apheresis is employed to reduce the rbc content from abo major or bidirectional mismatch bone marrow (bm) grafts mainly to avoid severe haemolysis . rbc depletion results in a significant volume reduction (due to both rbc and plasma depletion) and buffy coat concentration . . in pediatric setting, both rbc depletion and volume reduction before transplantation or cryopreservation can avoid fluid overload and renal impairment, especially in low/very low body weight recipients. the aim of this study was to evaluate the quality of the graft and immediate post infusion complications in rbc depleted bm in major and minor abo mismatch recipients using an automated device. patients and methods: bm aspirates for transplantation in pediatric setting were processed at our centre using the spectra optia (terumo bct) automated device. the initial collection preference was set at level and then was adjusted in order to maintain a haematocrit of % (colorgram) in the collection bag. flow speed was set at ml/min for cycles. mean recipients' body weight was kg (range: - ). pre and post procedure bm bag volume, hct%, mononuclear cells (mncs) count, (including b and t lymphocytes), cd + cell and cell viability were calculated. moreover, post procedure rbc volume and procedure time were registered. on the patient's side, post infusion complications (renal impairment, fluid overload, fever and haemolitic reactions) and time to engraftment were evaluated. results: a total of rbc depletion procedures were consecutively performed on bm grafts ( major and minor abo incompatibility, mud and related donors). data about pre and post procedure graft composition are reported in table . mean time to engraftment for pmn was . days (range: - ) and for plt was . (range: - ). pre and post-procedure cell viability were always %. mean procedure time was . minutes (range: - ). no bacterial or fungal contamination was detected. no infusion complications were recorded. one graft failure was observed. conclusions the spectra optia automated system is efficient in rbc depletion of abo mismatched grafts, permitting an effective volume reduction and an excellent mncs and cd + cell recovery in pediatric setting. automated rbc depletion may be proposed in low/very low body weight recipients both in abo major and minor incompatibility setting to minimize graft infusion side effects. building up a stem cell transplantation program in an emergent country, in the public setting, with limited economic resources, is not an easy work to do. international cooperation may be essential for the development of the program, in training, technological support and implementation of international guidelines. after years, we show an experience of international cooperation between a highly developed center in france (institut paoli calmettes, marseille) and the stem cell transplantation department of hospital maciel, a public assistance service in montevideo, uruguay. fourteen persons between doctors and nurses have been trained in france in stem cell collection and processing, patient's clinical handling, nursing, outpatients care and quality management. french missions of experts have been also received in hospital maciel every year since for in situ human resources training. in last years we developed a program for optimizing transplant results and reducing transplant related mortality (trm), based on several measures: improvement of patients selection, applying the sorror comorbidy index; adjustment of conditioning regimen doses, in order to reduce toxicity; development of a program to improve interaction with the intensive care unit; protocolization of the standard proceedings treatments; and initiating a program of quality and safety at the national institute of quality of uruguay inacal. adult patients have been treated with autologous (asct) ( ) or allogeneic (allosct) ( ) sct, with hematological malignancies. different modalities of allosct have been included progressively, becoming the only center accredited by the national regulation authorities (fnr) to perform unrelated donor sct and the haploidentical donor sct. this increased the proportion of allogeneic transplants from the historical % until % in last years. regarding patients health coverage, % comes from the private assistance system and % from the public health system. the major indications are lymphoid malignancies and acute leukemia, for asct and allosct, respectively, showing the same trend than cibmtr. three-year overall survival (os) for acute myeloid leukemia after allosct is %. considering asct for diffuse large b cell lymphoma, years os after autologous sct is % and % for chemo sensitive and resistant disease, respectively. threeyears os after asct for hodgkin disease is and % for sensitive and resistant disease, respectively. asct in multiple myeloma shows an os of and % at and years, respectively. in trm, results during the last years (after the described strategy) are shown in figure . the development of the-program of continuous improvement in quality-and the impact of results was locally recognized by two annual prices from inacal in (bronze) and (silver) in the category ‛commitment to public service.' a successful mirna- and the level of proangiogenic cytokines: angiopoietin- (angpt ), matrix metalloproteinase- (mmp- ) and vascular endothelial growth factor (vegf) in patients with lymphoproliferative malignancies prior to autologous hematopoietic stem cell transplantation (hsct) and in early posttransplant period. twenty-four patients were enrolled to the study ( f, m). the median (me) age was years. the investigated group consisted of multiple myeloma and lymphoma patients. the plasma samples were collected on time points: before chemotherapy-‛bc', on the day of hsct -‛ ', days after hsct-‛+ ' and days after hsct-‛+ . ' the cytokines were evaluated using elisa method, while mirna levels were estimated by qpcr method. the wilcoxon matched-pairs test was used to compare groups of dependent continuous variables: mirna's relative quantification (rq) levels or cytokines expression at two different time points. spearman rank correlation coefficient (r) was used to compare independent variables. we observed continuous decline of cytokines and mirnas level after conditioning treatment. the deepest decrease of expression was marked on ‛+ ' day ( table ) . we noticed a positive correlation between mirna- , mirna- cells in pbsc product. among the autologous transplanted patients between march and october , we have selected according to diagnosis, conditioning regimen and number of infused bags of cryopreserved pbsc. this group included females and males with median age of (range: . most of them, ( . %), had multiple myeloma (mm), ( . %) had non-hodgkin's lymphoma (nhl) and ( . %) had hodgkin's disease (hd). after harvesting, cd + cell and leukocyte number in pbsc product were enumerated on flow cytometer and blood cell counter, respectively. pbsc were cryopreserved with % dymethil sulfoxide (dmso) and cell viability was measured with trypan blue exclusion test before and after adding dmso, and as well after thawing in water bath on °c. as a conditioning regimen for the mm patients, melphalan was used and for the nhl and hd patients we used beam regimen. all received one bag of cryopreserved pbsc and pegfilgrastim mg on the first or the second post-transplant day. time to hematopoietic recovery was measured; for neutrophils . × /l, leukocytes × /l and platelets × /l with at least days without platelet transfusion. the median number of total leucocytes infused was . × /l (range: . - . × /l) of which cd + cells were - . × /kg of patient's body mass (median . × /kg). pre-freezing cell viability before and after adding dmso was with a median of % ( . - ) and , % ( . - ), respectively, and post-thaw viability . % ( . - ) . the average time to engraftment was . days ( - ) for neutrophils, days ( - ) for leucocytes and . days ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) for platelets. our results confirmed the known correlation between the number of infused cd + cells and engraftment of neutrophils (po . ), leukocytes (p o . ) and platelets (p = . ). we found inverse correlation between the infused leukocytes and cell viability with dmso (p = . ) and after thawing (p = . ). no correlation was found between prefreezing and post-thaw viability with hematopoietic recovery, and also between the cd + number and these viabilities. no differences were found considering patients' age, gender, diagnosis, conditioning regimen or day of applying pegfilgrastim. we can indirectly infer good survival of cd + cells and higher sensitivity of other nucleated cells to preparation of pbsc product. trypan blue exclusion assay, due to its inability to distinguish type of stained cells, is not relevant for cd + cells survival determination. disclosure of conflict of interest: none. chronic granulomatous disease (cgd) is a kind of primary immunodeficiency disorder of phagocytic cells which resulting in failure to kill a defined spectrum of bacteria and fungi and in concomitant chronic granulomatous inflammation. allogeneic hematopoietic stem cell transplantation is the only treatment proved to be potentially curative in cgd. unrelated umbilical cord blood (ucb) is increasingly used as an alternative to bone marrow. methods: unrelated ucbt was performed consecutive cgd children at our center between and . median age was . months (range: - months), median body weight was . kg (range: - kg). all patients received myeloablative conditioning regimen consisting of busulfan, fludarabine, cytarabine, cyclophosphamide and g-csf. all patients received tacrolimus as prophylaxis for graft-versus-host disease (gvhd). median nucleated cells were . × /kg (range: . - . × /kg), and median cd + cells were . × /kg (range: . - . × /kg). median follow-up time was . months (range: - months) results: of patients engrafted. median time to neutrophil engraftment was days, and median time to platelet engraftment was . days. / patients were alive, and / had full donor engraftment. overall survival rate was . %. disease-free survival was . %. of patients had grades iii-iv acute gvhd. no patients developed chronic gvhd. only one patient died from multi-organ failure related to adenovirus infection. conclusion: unrelated ucbt should be considered as potential curative methods in children with cgd. myeloablative conditioning regimen has improved the engraftments of the ucb. disclosure of conflict of interest: none. reduced muscular mass and excess visceral fat in patients undergoing hsct are associated with higher mortality, longer hospitalization, longer use of immunosuppressive drugs, graftversus-host disease (gvhd) and comorbidities leading to shorter survival time. a recent study of patients undergoing allogeneic hsct showed that occurrence of enlarged areas of visceral and peripheral fat is inversely associated with the disease-free interval after the transplant. reduced muscle mass has also been associated with higher prevalence of chronic gvhd and low rates of success following allogeneic hsct. objectives: to investigate whether amount muscle mass and muscle strength (ms) as well as the amount of visceral fat (vf) of patients undergoing hsct would influence the duration of the engraftment time (en). we evaluated hsct patients (⩾ years) at hospital israelita albert einstein, são paulo, brazil, on their first day of hospitalization, before hsct. the thickness of the right femoral quadriceps muscle (rfq), measured at cm from the top edge of the patella was measured using ultrasound (us) in b-mode. the dominant upper limb strength of the patients was evaluated by the hand grip test. the vf was measured in the abdominal region, by the thickness of the fat layer between the linea alba and the anterior wall of the aorta. most patients were women ( %) with a mean age of years (± years) and % of our patients were elderly (⩾ years). the haploidentical ( %) was the predominant hsct, autologous ( %) and allogeneic ( %). most patients were overweight, with body mass index (bmi) of kg/m (± kg/m ). the average time en was days (± days). rfq was . cm (± . cm), ms was kgf (± . kgf) and the vf was . cm (± . cm). patients with lower rfq had a longer engraftment time that was statistically significant as the negative correlation between rfq and en was rs = . , p o . ), independent of the age and the hsct type as analyzed by linear regression. no significant correlation between vf or ms with en was found. in this cohort of patients we found that longer engraftment times were significantly correlated to reduced muscle mass but no positive or negative correlation was found with superior limb muscular force or with the amount of visceral fat. disclosure of conflict of interest: none. hematopoietic stem cell transplant unit, hematology department and pharmacy department. university hospital of donostia. donostia/san sebastiań introduction: lymphocytes are the cells responsible for the cellular and humoral immunity and, consequently, critical for hematological patients. the aim of this study was to analyze the eventual conexion between lymphocyte recovery and survival (srv) after allogeneic hematopoietic stem cell transplantation (allo-hsct). patients and methods: we retrospectively analyzed data from consecutive patients who underwent allo-transplants in our unit. in total, patients were male ( . %) and female ( . %). median age was years old (range: . baseline disease was: acute leukemia ( . %), lymphoma ( . %), myelodysplastic syndrome ( . %), chronic myelogenous leukemia ( . %), multiple myeloma ( %), aplastic anemia ( . %), chronic lymphocytic leukemia ( . %) and others ( . %). . % of allo-hscts were from an unrelated donor, and . % from a family donor ( % of them haplo-identical). the sc source was pbsc in . %, and bm in , %. a variety of conditioning regimens were employed, including: busulphan-based ( . %), melphalan based ( . %), tbi-based ( . %) and others ( . %). evolution of absolute lymphocyte counts (alc) and subpopulations during the first year after allo-hsct were analyzed. results: as shown in table , alc decreased abruptly during conditioning therapy and recovered up to baseline at days + and + ; at day + median alc had clearly improved compared with admission values. median cd + cells were lower than /mcl in two thirds of pts at day + and in only one third at day + . as shown in table , we found a significant link between alc at day + and srv, as well as between cd + cells at day + and srv. in our series, immunity recovery was a late event for the majority of patients undergoing allo-hsct. in addition, in our experience, the precocity and quality of the alc and cd + recovery was clearly linked with long-term survival. disclosure of conflict of interest: none. although there is experimental evidence suggesting the presence of a common mesoderm cell as origin of both hematopoietic (hsc) and mesenchymal progenitor cells (msc) in an animal model, it is still controversial if durable engraftment of native donor-derived mscs without ex vivo treatment can occur in the recipient of allogeneic hsct. to assess the presence of donor-derived msc following hsct. between july and july , a total of recipients of hsct were analyzed for hsc and msc chimerism. eighteen patients received bm grafts ( %), patients had peripheral blood as stem cell rescue ( %) and finally patients had a cord blood transplantation ( %). patients received myeloablative ( %) or reduced intensity conditioning ( %) for malignant ( %) or nonmalignant disease ( %). bm aspirate cells were plated and expanded in α-mem with % human platelet lysate at cells/cm . after - days, nonadherent cells were removed, while the adherent cells were expanded until they reached confluence. after weeks we quantified msc precursors as colony forming unit fibroblast (cfu-f). finally the amplified sequences were resolved by capillary electrophoresis ( ruo genetic analyzer, applied biosystems) and analyzed by comparing genotypes of bmt recipell detachment, nuclear dna was extracted (dneasy blood and tissue kit-applied biosystems) and specific polymorphic tandemly repeated regions (strs) were amplified by means of the polymerase chain reaction(pcr) following the specific manufacturers' instructions. (ampfℓstr identifile kit, applied biosystems following hsct (hsc and msc) to those of donors. we cultured whole bm aspirates from patients following hsct with a median time of day (range: - ). cfu-f/ × growth was observed in a majority of bm the prevalence of human pegivirus in recipients of allogeneic hematopoietic stem cell olga koroleva , e parovichnikova , l kuzmina , m drokov , v vasilyeva , z konova , ekaterina mikhalcova , d dubnyak , n popova , tamara romanova , d tikhomirov , t tupoleva and v savchenko bone marrow transplant department, national research center for hematology and virology department, national research center for hematology human pegivirus (hpgv; previously named as gb virus c/hepatitis g virus) was discovered more than years ago. it is an rna virus referred within the genus pegivirus of the family flaviviridae. hpgv rna is found in liver, spleen, bone marrow and peripheral blood mononuclear cell, including t-and b-lymphocytes, nk-cells and monocytes. despite of the fact that it is a molecular structure, mechanism of replication and transmission routes are very well understood but the clinical significance of hpgv is still not determined. recipients of allogeneic hematopoietic stem cell have a high risk infection of hpgv. it is known, that hpgv is a nonpathogenic virus, however, it may play a role in immunocompromised individuals. to investigate the frequency of occurrence of hpgv and its clinical significance in recipients of allogeneic hematopoietic stem cell. blood samples were obtained from patients who underwent allogeneic hematopoietic stem cell transplantation (allo-hsct): all n = , aml n = , mpn n = , cll n = , mm n = , lpd n = , aa n = , mds n = . a median of age was years ( - years) . forty five patients were males and patients were females. conditioning regimen was ric in cases, mac in . bone marrow as a graft source was used in , pbsc- . all patients received multiple transfusions of blood components at the previous stages of treatment. hpgv rna had been assayed by polymerase chain reaction real time (rt-pcr) on plasma samples before started pre-transplantation conditioning. despite the diagnosis incidence of hpgv was high . % (rna-hpgv was positively in patients). patients with piercings and tattoos had incidence of hpgv in % that was not statistically significant (p- . ). hpgv is known as nonhepatotropic virus. in our study there was also no statistical reliability of specific changes in liver function test such as elevating the levels of alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase and total bilirubin due to the rna-hpgv. liver enlargement was also not statistically significant according to ultrasound scan results in patients infected with hpgv. we also analyzed the co-infection with hepatitis b and c virus. results are presented in table . coinfection was not statistically significant. however, only one patient with hepatitis c was coinfected hpgv. leukocytes recovery median was days ( - ). thrombocytes recovery median was ( - ). the presence of rna-hpgv did not affect the recovery of peripheral blood cells in patients after allo-hsct. according to our study the frequency of hpgv infection in recipients of allogeneic bone marrow was quite high ( . %), and it did not depend on the presence of any other hepatotropic viruses. clinical significance of hpgv infection in recipients of allogeneic hematopoietic stem cell has not been revealed, it is possible due to the short follow-up. it needs further clinical research. disclosure of conflict of interest: none. quantification of cd + recent thymic emigrants and t cell receptor excision circles (trecs) in umbilical cord blood transplanted patients v devlia , , j gridlestone , m raymond , s tulpule , d tewari , , r hough , c navarrete , a madrigal , , b shaw , , r danby and a saudemont , anthony nolan research institute, london, uk; ucl cancer institute, london, uk; nhsbt colindale, london, uk and ucl, london, uk reconstitution of t lymphocytes is a limiting factor in the regeneration of an effective immune system in adult patients following hematopoietic stem cell transplantation. cd (pecam- ) is a transmembrane glycoprotein expressed on naive t-cells that have recently emigrated from the thymus into the periphery. in peripheral blood, cd + t lymphocytes also contain high numbers of t-cell receptor excision circles (trecs); excision loops of dna excised during t-cell receptor gene rearrangement during t cell maturation within the thymus ( ) ( ) ( ) . however, quantification and correlation of cd and trec has not been formally investigated in patients following umbilical cord blood (cb) transplantation. quantification of cd and trecs post cb transplant will provide an insight into the immune reconstitution of t cells from the thymus. we therefore sought to measure cd and trecs in patients after cb transplant and assess whether these markers provided evidence of thymic recovery. we followed adult patients (median age . years) who underwent cb transplant in the uk. patient samples were collected , , , , and days post transplant. using flow cytometry, we determined absolute counts of cd +cd +cd ra+ and cd +cd +cd ra+, and quantified the copy numbers of trec genes in peripheral blood mononuclear cells (pbmcs) via real time pcr. results: at the six time points, the number of samples collected were the following: , , , , and . in all of the samples, the overall median number of cd +cd +cd ra+ was cells/μl (range: - cells/μl). the median level of cd +cd +cd ra+ cells increases from to cells/μl from day to day . absolute counts of cd +cd +cd ra+ at all of the six time points is -fold lower compared to healthy controls (median: cells/μl, range: - cells/μl). the overall median number of cd +cd +cd ra+ cells is cells/μl (range: - cells/μl). there is an increase in the median number of cd +cd +cd ra+ cells between days and posttransplant from to cells/μl. however, the absolute median counts of cd +cd +cd ra+ cells in patients are twofold lower, years post transplant, compared to healthy controls (median: cells/μl, range: - cells/μl). in the majority of the patient samples throughout all time points the trec gene copy numbers were undetected (n = ). in a few patient samples (n = ) trec gene copy numbers were quantified but with this limited sample size no correlations can be made between the absolute counts and trec gene copy numbers. our data suggests that cord blood transplant patients within the uk have reduced levels of cd +cd introduction: common variable immunodeficiency (cvid) is a highly heterogeneous group of primary immunodeficiency characterized by defective antibody production, recurrent infections, lymphoproliferation and autoimmunity. autosomal recessive mutations in lrba, encoding lps-responsive beigelike anchor protein were first described as a cause of cvid-like disease in . although hsct is accepted as a standard treatment modality for long-term resolution of severe primary immunodeficiencies, its role is less established in patients with lrba deficiency. patients and methods: whole exome sequencing of patient's genomic dna obtained prior to the hsct revealed a homozygous deletion in lrba (c. delt:p. c fs). immunological analyses including serum immunoglobulin levels, flow cytometry analyses of lymphocyte subsets, cytotoxicity/proliferation assays, vaccine responses were studied at several time points throughout the disease course, prior to and after hsct. a -year-old boy, born to consanguineous healthy parents of turkish origin became symptomatic at the age of months. he hospitalized several times due to recurrent pulmonary infections. he developed pancytopenia, lymphadenopathy, hepatosplenomegaly and autoimmunity (autoimmune hemolytic anemia and thyroiditis) with low serum immunoglobulin levels at the age of . as a result, he received several courses of steroid and prophylactic immunoglobulin and wide-spectrum antibiotics. over time he manifested growth failure and diagnosed with ibd-like colitis. due to the cumulating severe cvid-related complications, a hsct was performed at the age of years with the bone marrow stem cells from his hla identical brother after a conditioning regimen including fludarabine, busulfan and atg. severe intractable colitis with hypoalbuminemia continued till the engraftment despite vigorous fluid-electrolyte replacement therapy and accompanied with severe episodes of acute gastrointestinal bleeding. after the achievement of full donor chimerism, diarrhea episodes resolved. he received three doses of abatasept because of persistent cytopenia thinking about unresolved immune dysregulation. he is in complete remission at -year post-hsct with no signs of graft versus host disease. allogeneic hsct should be considered in patients with lrba deficiency prior to the development of disease-related severe cumulative manifestations. disclosure of conflict of interest: none. inflammatory bowel disease (ibd) is a chronic disorder of the gastrointestinal tract. very early onset ibd (veo-ibd) represents those severe children with disease onset occurring before -years-old. interleukin- receptors (il- ra, il- rb) mutation are considered to be one of the very important genes for veo-ibd. currently variant treatment, such as steroid medication, immunosuppressive agents and biological agents could not get complete remission. allogeneic hematopoietic stem cell transplantation (allo-hsct) was reported to induce remission in those with veo-ibd. we performed unrelated umbilical cord blood transplantation (ucbt) in five consecutive children with veo-ibd due to il- receptor mutation between and . median age of five children was months (range: - months), and median body weight was kg (range: . - . kg). all patients received reduced intensity conditioning (ric) regimen consisting of busulfan, fludarabine and cytarabine. prophylaxis for graft-versus-host disease (gvhd) was tacrolimus. most patients ( %) received a or hla alleles-mismatched cord unit. median nucleated cells of the cord blood were . × /kg (range: . - . × /kg), and median cd + cells were . × /kg (range: . - . × /kg). median follow-up time was months (range: - months). all patients engrafted, median time of neutrophil engraftment was days, and median time of platelet engraftment was days. four of five patients were alive with continuous donor engraftment, and achieved complete clinical remissions. colonoscopy at months after transplantation in two children revealed the mucosa healing. two children had grade iii acute graft-versushost disease (gvhd). one child developed severe chronic gvhd of both lungs and died of ards at months after transplantation. it is the first clinical trial that unrelated ucbt was performed in veo-ibd children in china. our data should unrelated ucbt with ric should be considered as a potentially curative therapeutic option in children with veo-ibd. disclosure of conflict of interest: none. patients with refractory primary induction failure and resistant relapse are poor candidates for hematopoetic stem cell transplantation (hsct) . additional attempts at remission induction with various combinations of chemotherapy will unlikely improve the outcome and will contribute to excess toxicity. a major goal of sct has been to develop strategies to reduce the risk of gvhd while maintaining or enhancing gvl. tcrαβ+/cd +lymphocytes depletion is a technology of graft manipulation with a potential to increase gvl effect and improve gvhd control and immune reconstitution in this group of patients. a total of pts with refractory aml (primary induction failure (n = ), refractory relapse (n = )), female/ male, median age . years ( . [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] , underwent allogeneic sct between may and august , median fu . years ( . - . ). pts were transplanted from haploidentical donors and from mud. all pts had active disease (ad) at the moment of sct and received treosulfanbased high-intensity conditioning regimen. three regimens of gvhd prophylaxis were used. regimen (n = ): atgam mg/kg with (n = ) or without (n = ) post-transplant tacro/ mtx; regimen (n = ): thymoglobulin mg/kg, rituximab mg/m and post-transplant bortezomib on day+ ,+ (n = ); regimen (n = ): tocilizumab mg/kg on day- and post-transplant bortezomib (n = ), pts receive additional abatacept mg/kg on day+ , + , + , + . tcrαβ+/cd +-depletion of sct with clinimacs technology was implemented in all cases. the median dose of infused cd + cells was × /kg (range: . - ), tcra/b- × /kg (range: - ). all engrafted pts received additional post-transplant courses of low-dose chemotherapy, including hypomethylating agents and dli. primary engraftment was achieved in of pts(three pts had disease progression, one died at the moment of engraftment), the median time to neutrophil and platelet recovery was days ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) . early mortality within days was . % (one pt with aml had acute lung injury after engraftment on day + ), . -years ptrm- . % ( %ci: . - ) . there were no allergic or infusion-related adverse events associated with tocilizumab or abatacept. ci of gvhd grades ii-iv and iii-iv was . % ( % ci: - ), and . % ( %ci: . - ), respectively. ci of cgvhd was % ( % ci: - ). ci of acute gvhd was lower in a group with prophylaxis regimen without serotherapy: % ( % ci: - ) vs . % ( % ci: - ) in atg group. no correlation between graft composition, donor type with the incidence of agvhd and cgvhd was noted at . years ppfs (event = death or relapse or progression) was % ( % ci: - ), . -years pos - % ( % ci: - ). median time of fu for survivors is . years (range: . [ ] [ ] . we confirm that the depletion of tcrαβ +/cd +lymphocytes from the graft ensures high engraftment rate and low transplant-related mortality in pediatric pts with refractory aml. we suggest that tocilizumab and abatacept can be safety administered to children with acute leukemia in the context of treosulfan-based conditioning regimens. long-term follow-up will demonstrate if the gvhd prophylaxis without serotherapy and combined administration of tocilizumab, abatacept and bortezomib post-tcrαβ+/cd +depleted grafting will improve gvl effects without extensive gvhd-related morbidity and mortality in pts with refractory aml. disclosure of conflict of interest: none. the jacie experience at the university hospital of amiens l marie-noelle , w brigitte , f isabelle , g bérengère , h muriel , h anne , v elsa , m jean-pierre and c amandine lacassagne; oncopôle, chu amiens picardie; hématologie clinique et thérapie cellulaire-chu amiens picardie; oncopôle-chu amiens picardie and the jacie (joint accreditation committee of isct and ebmt) accreditation aims to improve the management of patients benefiting from autologous or allogenic hematopoietic stem cell (hsc) transplants. usually, candidates' centers for jacie accreditation have already existing clinical activity when they have willingness to comply with jacie standards. here, we present our new experience in the implementation of jacie quality process, at the same time as allograft clinical activity. implementing process autograft clinical activity existed at amiens hospital, since , but in lack of center cellular therapy laboratory and hsc collection that were outsourced. the collection activity for autologous transplant was set up in , the cellular therapy laboratory in november and then the allogenic transplant was started in july . as early as march , we set up a steering committee with hematological clinicians, managers of each sector, a transplant coordinator nurse, the head of the processing laboratory and a part-time quality engineer recruited part-time. each actor had to become familiar with standards to obtain information from accredited centers in order to evaluate objectives and their prioritization. steering committee decided on deadlines and established a roadmap including the following: the list of jacie required standard operating procedures and their writing; assignment of the tasks for each actors in order to evaluate, writing and approval each document; organization of documents diffusion; information to all staff on the approach; creation of feedback committee for adverse events management; establishment of morbidity and mortality review; formalization of initial and continuing training for medical and paramedical staff; and organization of cross audits with external teams. at the same time, we assessed requirements for starting activity: training for medical and paramedical staff; training for the transplant coordinator nurse; circuits for taking care of donors; the organization of the in-patient department; the organization of follow-up of post-transplant patients; and authorizations of the national regulation agencies for processing facility on manipulations and cellular qualifications and for regulatory collection and transplant. allogenic transplant clinical activity started in july . accreditations jacie visit occurred on and june and our center has been officially accredited since march . neither quality approach, nor clinical activities were easy to implement. medical and paramedical staff had to get acquainted with a new organization and restrictions. despite difficulties, implementing jacie quality process, concomitantly with allograft activity allowed to create a true team dynamics with a common reflection on the means to be implemented. moreover, quality approach has assured us best ensure to care graft patients. the result is true satisfaction, which be credited to all. disclosure of conflict of interest: none. previously published p three-dimensional co-culture of peripheral blood monocytes supports and expands functional hematopoietic stem/progenitor cell without immobilization y xu , x li , b wang , w shan , h chen , s liu , r tie , y long , s cai , h xu , x yu and h huang bone marrow transplantation center, the first affiliated hospital, school of medicine, zhejiang university very low numbers of circulating hematopoietic stem/progenitor cells (chspcs) are found in normal human peripheral blood (pb) without mobilization. here, we developed a three dimension co-culture system to seize and expansion chspcs from pb monocytes without mobilization. flow cytometry analysis was carried out to identify chspc phenotypes. multipotential properties of chspcs were determined using colonyforming unit assay in methylcellulose and reconstitution ability in the compromised animals. the critical regulation mechanism underlying chspcs was identified with transcriptome analysis based on next-generation sequencing technology at total or single cell levels. loose cobble stone colonies (lcs), round or vessel-like compact colonies (rccs or vccs) were presented in three dimension co-culture system after about weeks. the colonies lasted for at least six passages with no obvious apoptosis sign, and expanded more than~ fold during the period. we studied the niche-mediated regulation mechanism of chspc fate at molecular level compared to the conventional method of two dimension culture. furthermore, chspcs were capable of forming all types of hematopoietic colonies, including cfu-gemm, and especially held short term engraftment capacity for compromised nogs by radiotherapy. transcriptome analysis by deepsage identified genes significantly associated with regulating the function of chspcs. figure : the cellular morphology in three dimension culture system for peripheral blood monocytes without mobilization during the culture for - weeks. figure : short transplantable potential analysis of chspcs. figure : (a) static of differentially expressed genes between three-and two-dimensional culture systems for peripheral blood cells. (b) go functional analysis classifies those genes by biological process, cellular component and molecular function. (c) the significant differences between the molecular phenotypes of three-and two-dimension chspcs indicating that chspcs from three dimension culture hold stem properties. our system may provide a more ideal and balanced approach which not only seizes circulating chspcs, promotes selfrenewal and expansion of chspcs, but also holds phenotypic and functional attributes of chspcs. . to wash or not to wash? comparison of neutrophil and platelet engraftment after infusion of cryopreserved autologous stem cells before and after the implementation of bedside thawing am halldorsdottir , s atladottir , m thorsteinsdottir, na arnason , g runarsson , t jonsson , oe sigurjonsson , and s reykdal the blood bank, landspítali, the national university hospital of iceland; department of hematology, landspítali, the national university hospital of iceland and school of science and engineering, reykjavik university cryopreserved autologous peripheral blood stem cell (pbsc) grafts are widely used after high-dose chemotherapy in the treatment of patients with myeloma or lymphoma. prior to infusion, cryopreserved grafts can be thawed at the bedside, or thawed and washed at the cell therapy laboratory. at our institution the practice of routine washing of stem cell grafts in the laboratory was discontinued in april and bedside thawing implemented instead. this was done to minimize the time thawed cells are exposed to toxic dmso. this study was performed at a single center, at landspítali-the national university hospital of iceland, which is the only transplant center in iceland. autologous pbsc transplants have been performed in iceland since . the study compares outcome for two groups of patients, who received either; (a) thawed and washed autologous pbsc cell grafts from january to [p ] april , or (b) autologous pbsc grafts thawed at the bedside from april to november . the following outcomes were compared; days to neutrophil engraftment (absolute neutrophil count (anc) . per μl), and platelet engraftment ( and × e /l). data on mean cd + cell content/kg of the infused grafts, measured prior to cryopreservation, were also compared. all patients have received premedication with solucortef, clemastine and ondansetron prior to infusion of the graft. from january to april a total of patients received thawed and washed autologous pbsc grafts, and between april and november patients received autologous pbsc grafts thawed at the bedside. majority of the patients were diagnosed with either multiple myeloma or related disorders (n = ) or lymphoma (n = ) whereas the remaining patients (n = ) had miscellaneous diagnoses. days to engraftment and the dose of cd + cells infused are compared in table . there was no significant difference in the mean cd content of infused autologous stem cells in the two groups ( . vs . × e cd +cells/kg, p = . ). there was also no difference in the mean number of days to engraftment of neutrophils ( . vs . days, p = . ), platelets at days ( . vs . days, p = . ) or platelets at days ( . vs . days, p = . ) after transplant. one hundred day mortality was comparable in the two groups or . %. additional data on transfusion requirements, infections and use of granulocyte-colony stimulating factor will be presented. [p ] there was no difference in neutrophil or platelet engraftment after changing the autologous stem cell graft thawing procedure from post-thaw washing in the laboratory to bedside thawing. bedside thawing of stem cells is a safe procedure that results in acceptable cellular engraftment. disclosure of conflict of interest: none. the procedure of autologous hematopoietic stem cell (hsc) transplantation requires cryopreservation of hscs. addition of dmso (dimethyl sulfoxide) is necessary to secure the viability of such cells, but this cryoprotectant causes adverse reaction during infusion into patient. the concentrations of dmso in cryopreservation mixture vary strongly between different transplant centers. usually, the hscs are stored in mixtures containing % dmso, however, many centers successfully use lower concentrations. the main aim of the study was to evaluate the clinical impact of different dmso concentrations in cryopreservation mixture ( %, . %, %) on reconstitution of hematopoiesis after autologous hsc transplantation. the project was approved by the local bioethics committee. written informed consent obtained from all of patients. the study is registered to clinicaltrials.gov (identifier: nct ). between january and july , consecutive patients with hematological malignancies or solid tumors, referred for autologous hsc transplantation, were recruited in the study. the patients were randomly assigned to one of three study arms ( patients each). hscs obtained by leukapheresis were cryopreserved in three concentrations of dmso: %, . %, %, respectively. study groups did not differ significantly with regard to the diagnosis (mostly mm, nhl or hl), age or conditioning regimen (chemo-or radiotherapybased). all patients received granulocyte-colony stimulating factor (g-csf, filgrastim) starting from day + after transplantation to support neutrophil recovery. in case of patients, the transplantation was cancelled due to progression or other medical reasons. four patients died shortly after transplantation, due to refractory infections. data for patients were subjected to statistical analysis. the viability of nucleated cells on the day of transplantation was similar in all groups (median %, range: - % for % dmso group; %, range: - % for . % dmso; %, range: - % for % dmso; p = . ). the dose of transplanted cd + cells was comparable in all group: (median . × /kg of recipient body weight for % dmso, . × /kg for . % dmso and . × for % dmso, p = . ). the median time to leukocyte recovery, defined as the first day with wbc count exceeding . × /l was days in all groups (ranges: - for % dmso; - for . % dmso; and - for % dmso; p = . ). similar results were obtained in case of neutrophil recovery-the median day, when the anc exceeded . × /l, was in all arms (ranges: - ; - and - , respectively; p = . ). the day when the platelets level were greater than or equal to × /l (sustained without transfusion within days) was similar in all groups: medians were days in %, . % and % dmso (ranges: - ; - ; - ; p = . ). no serious adverse effects were observed during hscs infusion and during h after transplantation. reduction of dmso concentration from in cryoprotective mixture % to . % and % has no negative impact on cell viability during cryopreservation and engraftment after auto-hsc transplantation. disclosure of conflict of interest: none. a real-world cost-effectiveness analysis demonstrates that introducing plerixafor to improve mobilization in multiple myeloma patients who behave as poor mobilizers is cost-effective considering the whole mobilization and transplant procedure r touzani , , a-m stoppa plerixafor, a cxcr -antagonist, is efficient to improve cd + cell mobilization and collection in candidates for autologous transplantation who behave as poor-mobilizers. the cost of the drug is however of concern. published medico-economics studies were mostly conducted in the us, and few including detailed and comprehensive micro-costing of the collection and transplantation process; conclusions may thus not apply to european countries where cost structures are different. to compare costs and effectiveness of plerixafor-free and plerixafor-replete management strategies for multiple myeloma patients who behaved as poor-mobilizers after adequate administration of a standard rhg-csf mobilization regimen. sixty patients diagnosed with multiple myeloma were consecutively identified during years - , immediately before and after ema granted marketing authorization for plerixafor. poor-mobilizers were defined as having circulating cd + cell counts below /μl. plerixafor was introduced or not as a result of the attending physician's decision, reflecting progressive changes in medical practices over this transitional period. the historical and study groups were matched over four criteria: disease stage at diagnosis, age, gender and number of chemotherapy treatments received before mobilization. two cost-effectiveness analyses (cea) were conducted; the primary cea looked at the criterion ‛collecting at least × cd + cells'; a secondary cea looked at the criterion ‛successful autologous transplant administered'. detailed micro-costing evaluations ( figures) did not or did include transplantation costs for the first and second cea, respectively. the two groups were similar in terms of age, sex distribution, disease characteristics or previous treatments. / and / patients proceeded to high-dose melphalan and autologous transplantation in the study and historical groups, respectively. there was a trend to a higher number of collected cd + cells in the control group; however, the proportion of patients who met the minimal target number of × collected cd + cells/kg was identical ( / ). length of hospitalization, times to neutrophil and platelet recoveries, numbers of prbc and platelet transfusions were identical in the two groups. mobilization and collection costs per patients were more important in the plerixafor group that in the historical group ( . vs . €, p o . ), and proportionally higher in patients who received plerixafor as part of a remobilization treatment rather than pre-emptively ( . vs . €, respectively). the main cea concluded to a . € increase in costs for the same number of patients achieving a minimal target number of × collected cd + cells/kg. the second cea found a decrease in the cost of transplant, with . € in the study group vs . € in the historical group (ns). in total, the . € increase for the complete procedure cost ( . € per successfully autografted patient in the study group vs . € in the historical group) was not statistically different. cost-effectiveness arguments should not been used against the administration of plerixafor in multiple myeloma patients in the european context. future prospective researches looking at patients reported outcome criteria and labour organization in apheresis facilities are needed. disclosure of conflict of interest: this work was supported by a grant from sanofi s.a.; cc: research support, honorarium & hospitality from sanofi s.a. administration of plerixafor for peripheral blood cd + stem cell content of o × /l for autologous stem cell mobilization leads to decreased apheresis days and increased total yield m kamdar , s abebe , gr gonzalez fontal, l gates , a hammes , d abbott , j gutman , b haverkos , d sherbenou and c smith division of hematology and transplantation and department of biostatistics and informatics, university of colorado, denver, colorado, usa autologous stem cell transplantation (asct) is an effective treatment for lymphoma and plasma cell neoplasm (pcn) (multiple myeloma and amyloidosis). granulocyte-colony stimulating factor (g-csf) is the most commonly used upfront mobilizing agent with plerixafor-based higher cost approaches reserved for poor/unsuccessful mobilizers. several mobilization algorithms utilizing g-csf and plerixafor have been published however the most efficient and cost effective strategy is yet to be determined. most transplant centers administer plerixafor for peripheral blood (pb) cd + stem cell content of o × /l on day (d) of g-csf mobilization. at the university of colorado (uch) we changed our programmatic approach in and administered plerixafor for pb cd + count of o × /l on d of g-csf mobilization. in this study we evaluate the impact of this novel mobilization algorithm on apheresis days and total stem cell yield. patients (pts) with lymphoma and pcn who underwent asct at uch until / received plerixafor if pb cd + cells on d of g-csf mobilization was o × /l. based on our institutional review of poor/unsuccessful mobilizers and using logistic regression analysis this algorithm was revised in / . in the new algorithm all pts received plerixafor if pb cd + cells on d of gcsf mobilization was o × /l. demographics were compared between pts with lymphoma and pcn before (group : / - / ) and after (group : / - / ) the new algorithm was implemented. the primary goal of this analysis was to assess the total days of apheresis and total stem cell yield between the two groups. we also sought to analyze days to wbc engraftment and platelet engraftment. a total of pts were included in this analysis. group consisted of pts ( pts had lymphoma and pts had pcn). group consisted of pts ( pts had lymphoma and pts had pcn). we found that there was a significant increase in total yield (p = . ) in group as compared to group . on further disease subtype assessment we noted that pts with pcn in group had a significant increase in total yield (p = . ). in lymphoma pts on univariate analysis group showed a significant decrease in apheresis days ( . days, p = . , % ci: (− . , − . )). on multivariate analysis there was still a marginally significant decrease in group ( . days, p = . , % ci: (− . , − . )) compared to group . in pcn pts on univariate analysis group showed a significant decrease in apheresis days ( . days, p = . , % ci: (− . , − . )). on multivariate analysis group continued to show a significant decrease in apheresis days ( . days, p = . , % ci: (− . , − . ) compared to group . we found no significant difference between the two groups in days to neutrophil engraftment and platelet engraftment. our analysis showed that a mobilization algorithm of administering plerixafor for a pb cd + stem cell count of o × /l on d of g-csf mobilization led to a decrease of roughly . days in the lymphoma cohort and a significant decrease of . days in the plasma cell neoplasm cohort. we also noted a significantly increased total yield of stem cell collection in group . overall our programmatic approach led to decreased chair-time for apheresis and better resource utilization. pharmacoeconomic impact of this approach will be updated at the meeting. disclosure of conflict of interest: mk: speakers bureau, seattle genetics; remaining authors declare no conflict of interest. administration of stem cell boosts (scbs) from the original donor offers a therapeutic option. we report on pediatric patients with pgf who received a total of boosts with cd + selected peripheral blood stem cells (pbsc) after transplantation from matched unrelated (n = ) or mismatched related (n = ) donors. median time between hsct and infusion of the scbs was days ( - ). boosts contained a median number of . × cd + progenitor cells/kg body weight (range: . - . × ) with a median number of /kg (range: - ) residual cd + t cells. within weeks after application, a significant increase in median neutrophil counts ( vs /mm , po . ) and a decrease in erythrocytes and thrombocytes transfusion requirement (median frequencies and vs , p o . and o . ), were observed, and . % of the patients resolved one or two of their initial cytopenias whereas . % had a complete hematological response. additionally median lymphocyte counts for cd +, cd +cd +, cd + and cd + increased . fold, . fold, . fold and . fold, respectively. the rate of de novo acute gvhd grade i-iii was only % and resolved completely after treatment. no gvhd iv or chronic gvhd occurred. patients who showed a response to scb displayed a trend toward better overall survival (os) (p = . ). administration of cd + selected scbs from alternative donors is a safe and effective procedure. we hypothesize that the cd + progenitor boosts may have an enhancing effect on maturation of committed lymphoid precursors already present in the host or generate another wave of thymic seeding with accelerated t-cell differentiation process in the absence of any immune suppression. further studies are warranted to better define the impact on immune reconstitution and survival. disclosure of conflict of interest: none. plerixafor plus granulocyte-colony stimulating factor (g-csf) has been shown to mobilize more cd + cells than g-csf alone for autologous hematopoietic stem cell transplantation (hsct). however, there are few studies that analyze the impact of this strategy in engraftment. the aim of our study is to compare mobilization and engraftment between patients who received a combination of plerixafor plus g-csf and patients (pts) who mobilized with g-csf alone. a retrospective casecontrol analysis was performed in pts with myeloma who mobilized with plerixafor plus g-csf (group p/g-csf) and was compared with matched for sex and age controls who mobilized with g-csf alone (group g-csf). all pts underwent hsct between and . mobilization with g-csf at dose of μg/kg/day was used in all pts. the aphaeresis was scheduled on day + . plerixafor ( . mg/kg) was added if the number of cd +cells on day + was o /μl for × cd +/kg requested (or o /μl for × cd +/kg), or if the number of cd +cells collected in the first apheresis was o % of cd + requested. conditioning and supportive care were similar in both groups. in p/g-csf group, were male and female. median age was . years (range: - ). in group g-csf, were men and female. median age was . years (range: - years). there were no differences between both groups. disease status at time of mobilization was different between groups (p = . ). in p/g-csf group: ( . %) pts were in complete remission (cr), ( . %) very good partial responses (vgpr), ( . %) partial response (pr) and ( . %) had no response to treatment. in g-csf group: ( . %) pts had reached cr, ( . %) vgpr and the remainder in pr. sixteen ( . %) pts in p/g-csf group had received ⩾ lines of treatment vs ( . %) pts in g-csf group (p = . ). no difference was seen on mean day-dose of g-csf ( μg/kg/ h in p/g-csf group vs μg/kg/ h) (p = . ). there was no difference on cd +/kg requested ( / pts in p/g-csf were requested × /kg vs / in g-csf group) (p = . ). p/g-csf group needed more apheresis sessions, ( . %) pts required ⩾ sessions against ( . %) pts in group g-csf (p o . ). we obtained enough cd + cells to carry out hsct in all patients, although mean number of cd + cells obtained in p/g-csf group was lower than in g-csf group ( . × /kg vs . × /kg, respectively) (p o . ). also, mean number of cd + infused in p/g-csf group was lower ( . × /kg vs . × /kg) (po . ). however, engraftment results were similar in both groups, as represented in table . patients who required mobilization with plerixafor plus g-csf got an engraftment as good as patients who do not require the combination despite of worse baseline parameters. given that the number of cd + infused in the p/g-csf group has been lower than g-csf group, these results might suggest that the different composition of graft cell with plerixafor plus g-csf mobilization, described in some studies, could impact on engraftment outcomes. high-dose chemotherapy following autologous hematopoietic stem-cell transplantation (autohsct) is an effective method of treatment both recurrent and primary refractory lymphoma patients. however, some patients have mobilization failure (‛poor mobilizers') with inadequate collection of peripheral blood stem cell (pbsc). aim: to evaluate the efficacy and factors influencing pbsc mobilization and collection for the autohsct in patients with lymphomas. thirty patients were included in this study: -with hodgkin lymphoma, -with non-hodgkin lymphoma, -with multiple myeloma; women and men of them. the median age of patients was years ( - years). the mobilization of pbsc with only colony-stimulating factors (csf) was carried out for patients, chemotherapy (cyclophosphamide, etoposide) in combination with csf-for patients. only one patient had plerixafor mobilization. the concentration of cd + in peripheral blood (pb) was studied on the day of the intended cytapheresis. cytapheresis was commenced when cd + concentration had been greater than . × cells/ml. twenty-four patients ( %) from had collection of pbsc. the collection was not performed in six patients ( %) because the concentration of cd + in pb on the day of the intended cytapheresis was lower than . × cells/ml. there was no possibility to use plerixafor in these cases for economic reasons. the median concentration of cd + in pb on the first day of the intended cytapheresis in the group of patients that had cytapheresis was . × cells/ml whereas in the group of failed- . × cells/ml (p o . ). fifty-nine tests of cd + in pb were done. distribution and test results by days from the first day of the intended cytapheresis are presented in table . the total number of the cytapheresis was . the majority of patients had procedure of pbsc collection (n = ), patients had procedures and only had . the last patient had had two previous failed cytapheresis procedures and the adding of plerixafor helped him to collect necessary number of cells. the median of cd + cells on patient's kilo was . × cells/kg. sex, age, mobilizing regimen, previous radiation therapy, the count of lines of chemotherapy before autohsct were not significantly associated with poor pbsc mobilization and collection. only tumor response before autohsct (complete/ partial response or stabilization) was significantly associated with cd + cell count in the product of cytapheresis. patients with complete or partial response had significantly better cd + count. [p ] disclosure of conflict of interest: none. factors associated with failure in mobilization of peripheral blood hematopoietic progenitor cells in autologous transplantation je dulon-tarqui, bl acosta-maldonado, l rivera-fong, sa sánchez-guerrero, jf zazueta-pozos, ja padilla-ortega, wj ladines-castro and lm valero-saldaña high dose therapy followed by autologous stem cell transplantation (asct) obtained from peripheral blood is currently the standard model for treatment consolidation in various hematologic malignancies. a global incidence of - % of failure to mobilization is reported, and some factors associated with poor mobilizers in hodgkin's lymphoma (hl), non-hodgkin's lymphoma (nhl) and multiple myeloma (mm) when the yield in peripheral blood stem cells (pbsc) collection is unsatisfactory, the effects for the recipient can be serious. the donor's age, gender, body surface area (bsa), processed blood volume and the method of g-csf dose calculation may affect the cd + yield. as g-csf has a low distribution volume in the peripheral blood (pb), it might be appropriate to calculate the doses by using the bsa instead of per kg body weight. consecutive allogeneic pbsc donations performed in healthy donors at the karolinska university hospital in stockholm were included. a complete medical history, physical examination, electrocardiogram, chest x-ray and laboratory testing were done before pbsc donation. relevant data for analysis were collected from the institutional quality database for a retrospective review. the total blood volume was calculated using the formula by nadler et al. the bsa was calculated using the formula by du bois and du bois. the concentration of cd + cells in the pb and the processed volume of blood were significantly correlated to cd + cells yield (po . and po . , respectively, see table ). the g-csf dose per m was significantly correlated to the concentration of cd + cells in the pb (p = . ) and in the product (p = . , see table ). smaller bsa (p o . ) and less processed volume (p o . ) were found among female donors, who were given lesser g-csf dose per m (p o . ) and showed lower yield compared to men (po . ). however, multivariate analysis of the yield showed that only the concentration of cd + cells in the pb and the processed volume remained independent significant (see table ). [p ] in this study, we found the concentration of cd + cells in the pb and the processed volume of blood to be independent predictors of yield. we recommend to get a high concentration of cd + cells in the pb, and to process adjusted volumes of blood when needed. an evaluation if the calculation of g-csf dose per m is more appropriate than per kg body weight should be done in future studies. autologous stem cell transplantation (asct) has been widely used in the treatment of hematological malignancies over the last two decades. despite its broad use, some characteristics that might influence engraftment have not been exhaustively investigated, particularly graft purity with respect to contamination by platelets (plts) and white blood cells (wbc). here we report collection characteristics and engraftment kinetics of a single center consecutive series of asct. we retrospectively collected clinical records of patients who underwent leucapheresis procedures (la; followed or not by asct) and data on asct at our institution over years ( - ) ( table ). the impact on engraftment kinetics of conditioning chemotherapies, amount of infused cd + cells and wbc/plts graft contamination were analyzed. absolute neutrophil count (anc) engraftment was defined as the duration of neutropenia (from day to the first of consecutive days of anc /μl post asct). regarding cd + cell collection, no impact of mobilizing regimens and wbc count during la was observed. on the other hand, we observed a difference in the number of total cd + cells collected among different diagnoses: the median overall collection was . ( . - . ) × /kg cd + cells for nhl patients, . ( . - . ) × /kg for mm patients, . ( . - . ) × /kg for hl patients and . ( . - . ) × /kg for aml patients) (p = . ). considering cd + cells/kg harvested on the first day of la, . % of nhl and hl, . % of mm patients and % of aml patients harvested ⩾ × /kg cd + cells. of note, among aml patients, . % collected o . × /kg. the differences were statistically significant (p = . ). moreover, an inverse correlation between collected cd + cells and age was shown (p = . ). anc recovery after asct was not influenced by conditioning regimen whereas diagnosis impacted on the duration of neutropenia (aml patients displayed a longer aplasia, po . ). we observed that the median days with anco /μl were , and in patients who received . × /kg, . - . × /kg and o . × /kg cd + cells, respectively (po . ). furthermore, the same finding was observed considering the duration of thrombocytopenia (median number of days with plts o /μl: , and in patients who received . × /kg, . - . × /kg and o . × cd + cells, p o . ). looking at the apheresis product, we analyzed the impact of harvest contaminating wbc and plts on engraftment kinetics. notably, when the asct collection contained × /μl wbc, anc engraftment (days with anc o /μl) lasted longer (median days ) compared to patients who received a graft with lower wbc count (po . ). a faster anc engraftment was also observed in patients receiving harvests with plts levels × /μl compared to those who infused a collection bag with plts o × /μl (p = . ). herein, we confirmed that the disease and the amount of infused cd + cells significantly influence time of anc and plts engraftment; furthermore, we observed for the first time that quality and purity of the graft have a substantial impact on engraftment kinetics. a combination of chemotherapy with growth factor is a commonly used strategy for hematopoietic stem cell (hsc) mobilization. the collection of timely and adequate numbers of hscs is a prerequisite for proceeding to transplantation. a variety of mobilization strategies are currently used. the knowledge of efficacy, safety and predictability of different hsc mobilization strategies might help blood and marrow transplantation (bmt) programs to effectively schedule patients for mobilization. given the many variables associated with the mobilization of hsc, collecting an adequate stem cell dose in a timely and effective manner is an art and science. factors that might affect the process includes type of disease and mobilization protocol, financial clearance, availability of chemotherapy beds, scheduling various diagnostic procedures and transplant urgency. to evaluate the effectiveness and related coordination efforts of ‛just-in-time' strategy of hsc mobilization and collection, we performed a retrospective study comparing all patients in whom peripheral hsc mobilization was attempted at khcc from january through november . data collected included the disease type, mobilization protocol, days to and number of collections, cd + cell dose, calendar of the mobilization and collection. the records of a total of mobilizations were reviewed. were of healthy allogeneic donors, and the remaining were of patients undergoing autologous transplantation. table depicts the overall summary of number of days and collection procedures per each protocol. detailed mobilization kinetics per disease type and mobilization protocol were also captured and evaluated. [p ] s [p ] detailed analysis of mobilization kinetics comparing different mobilization strategies aids in prediction of number of days of mobilization and anticipated number of collections. this helps in proactively scheduling patients based on collection predictability. a seamless communication through a shared calendar between key parties, primarily bmt physicians and nurse coordinators, bmt and flow cytometry laboratories and chemotherapy unit can be achieved. autologous stem cell transplantation is still a standard of care in the treatment of multiple myeloma. lenalidomide-based regimens are commonly used in both transplant-ineligible as well as -eligible patients. prolonged lenalidomide-exposure is known to affect mobilization of cd + cells, although the basic mechanisms are poorly understood. limited prospectively collected data is available on the effect of lenalidomide in the capacity to mobilize cd + cells for transplantation as well as graft cellular composition and post-transplant hematological recovery compared to the lenalidomide-naive patients. this prospective study included newly diagnosed myeloma patients who received mobilization with low-dose cyclophosphamide + g-csf, were successfully apheresed and transplanted before the end of . twenty-six patients had received a median of three cycles of lenalidomide-based induction ( %), whereas patients were lenalidomide-naive and served as the control group. both baseline characteristics and collection targets were similar between the groups. cd + mobilization and apheresis yields were analyzed and compared between the groups. blood graft cellular composition was analyzed from the thawed cryopreserved samples with a flow cytometry. graft function was evaluated by collecting engraftment data as well as by total blood counts at day + and at , , and months after post-transplant. the patients in the lenalidomide group had both lower median peak b-cd + counts and about % lower cd + yields of the first apheresis but without statistical significance ( table ). the median number apheresis was significantly higher in the lenalidomide arm ( . vs . , p = . ). the number of cd +cd +cd -, cd +cd +, cd +cd + cells and nk cells in the cryopreserved grafts were comparable between the arms. time to neutrophil engraftment was days in the both groups. the median time to platelet engraftment was d in the lenalidomide group and d in the control group. hematological recovery was comparable between the groups within months post-transplant. lenalidomide-based induction therapy seems to have an impact in the number of apheresis needed, but not in the total yield of cd + cells in the graft. neither the graft cellular composition nor posttransplant recovery in myeloma patients was affected by the limited duration of lenalidomide used before mobilization and collection of blood grafts. between september and november . siemens hematek system was used for luc count. luc numbers and percentage was measured before leukapheresis. we used pearson test for the correlation and roc curve for cut off value. patients' characteristics were shown in table- . there was not a correlation between luc number and mobilized cd positive stem cell number. but luc percentage was positively correlated with mobilized stem cell number (p: . ). a count of × /kg collected stem cells are optimal for autologous stem cell transplantation. we found % luc percentage as a cut-off value for prediction of collecting optimal number of stem cells with % sensitivity and % specificity. as expected luc percentage was negatively correlated with white blood cell count. there was no correlation between mobilized cd positive stem cell number and age. both luc percentage and mobilized cd positive stem cell number did not differ with underlying disease. we found only one study in the literature that evaluated luc percentage as a tool for the prediction of successful stem cell collection. they found that baseline luc numbers negatively correlated with stem cell mobilization in healthy donors ( ) . but we measure luc on apheresis day and found a positive correlation between luc percentage and stem cell mobilization. and we found a cut-off value for optimal stem cell mobilization with acceptable sensitivity and specificity. in our study we demonstrate that luc percentage measurement on apheresis day may be a very simple and cheap tool for the prediction of optimal stem cell mobilization. the spectra optia (so) apheresis system performs a wide range of therapeutic procedures, including peripheral blood stem cell (pbsc) collection in mobilized donors and patients (pts). the device was studied to evaluate the cellular composition of pbscs harvested in pts with multiple myeloma (mm), non hodgkin's lymphoma (nhl) and hodgkin's lymphoma (hl) planed for autologous peripheral stem cell transplantation (apbsct), and to optimize the collection of pbscs using the cd + precount and collection efficiency (ce ) of apheresis device which is calculated as follows: ce = total cd + cells collected × /kg; cd + precount/ μl × blood processed (liters). the blood volume processed is calculated as follows: desired cd + × /kg × recipient weight (kg): ce × cd + precount/μl in our study enrolled pts undergoing pbsc mobilization and planed for apbsct. we evaluated so system's mononuclear cell (mnc) collection performance, with respect to cd + cells and mnc collection efficiency, platelet reduction pre to post apheresis, and product purity in view of using prediction algorithms to optimize the procedure and predict the cd + yield, blood volume processed and platelets loss. we also evaluated neutrophil and platelet recovery in pts who underwent apbsct. results: between / / and / / , pts underwent pbsc harvesting by so device. median age was years ( - ). there were females and males. diagnosis was mm in pts, hl in pts and nhl in pts. the number of ahereses procedures was . mobilization consisted in g-csf alone in pts, chemotherapy and g-csf in pts, and g-csf + cxcr inhibitor in one patient. median count of cd + cells pre-collection was /μl ( . - ) . median total blood volume processed was . l ( . - . ). median count of cd + cells collected was . × /kg ( - . ). median mnc collection efficacy was % .median cd + cell collection efficacy was . % ( - %). median platelet reduction pre to post apheresis was % ( - %). median product hematocrit and granulocytes product was % ( - ) and % ( - ), respectively. twenty-six of the pts underwent myeloablative high dose chemotherapy followed by apbsct which was performed for mm in pts, hl in pts, and nhl in pts. the median count of cd + cells infused was . × / kg ( . - . ). all the pts received g-csf post-apsct until neutrophil recovery. the median day for neutrophil recovery was ( ) ( ) ( ) ( ) ( ) ( ) ( ) . median duration of severe neutropenia (anc o . × /l) was days ( - ). the median day for platelet recovery was ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) . median duration of severe thrombocytopenia (platelets o × /l) was . days ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) . conclusion: the study results confirm that the so apheresis system's mnc collection protocol is safe and effective. the neutrophils and platelets recovery in pts auto-transplanted was not inferior compared to historical controls. in addition, this system help to use prediction algorithms for whole blood processing to achieve a desirable and optimal yield based on cd + precounts and ce of the apheresis device. disclosure of conflict of interest: none. peripheral blood stem cell apheresis in small children is difficult! aa hedayati-asl , m emam-jome, p dinarooni, v fallah, a mehrvar and r zangooei in low-weight children with cancer and healthy donor children, peripheral blood progenitor cells (pbpcs) have largely replaced bone marrow as source of autologous and allogeneic stem cells in part because of their relatively easy collection. however, there is a concern regarding medical, psychosocial and technical difficulties in small children. we retrospectively analyzed peripheral blood stem cell apheresis in collections. patients were with cancer ( patients = neuroblastoma, patients = retinoblastoma, patients = germ cell tumor, patient = hepatoblastoma, patient = wilm's tumor) and healthy children donors. the study was conducted between and . peripheral stem cell apheresis was performed in the mahak cancer children's hospital in a nice room for children where the patients stayed with their families. patients s were not routinely sedated. pbpc were collected by a cobe spectra cell separator (cobe, denver, co, usa). harvesting was performed after days mobilization. mean body weight was . kg (range: - kg) for a median age of years (range: months- years). mean duration of harvesting was min (range: - min). mean volume of stem cell collection was ml (range: - ml). the mean number of total nucleated cells collected was . × /kg (range: . - . × /kg recipients). no side effects occurred. children didn't require an additional haematopoietic progenitor mobilization or additional apheresis in other day. pbsc collection was without transfusion in healthy donor children. pbsc collection may be difficult in small children owing to the large volume apheresis compared to the child's weight. various problems, such as metabolic or haemodynamic disorders may be were seen. peripheral stem cell harvest can be performed in low-weight children under safe and effective conditions even when systematic priming by blood is avoided. processing with increase of blood volume may to increase in the yield by recruiting progenitor cells. disclosure of conflict of interest: none. peripheral blood stem cell collection in low body weight children: a single centre experience g del principe*, g leone, s lazzaro, a meschini, k feri, p marchitelli, d carasso, f locatelli and m montanari department of pediatric hematology/oncology and transfusion medicine, irccs bambino gesù children's hospital, rome, italy pbsc became preferred source for autologous transplantation because of easier collection and faster engraftment. however apheresis for low body weight children ( o . kg) is affected by some issues: venous access, extracorporeal volume, metabolic and hemodynamic complications, citrate toxicity, so is crucial to standardize harvesting procedure both maximizing stem cells collection and reducing adverse events. a dual lumen central venous catheter was used to obtain a minimal blood flow of - ml /min and pbsc collection was performed with spectra optia mnc v . apheresis system, starting with cd + cell ⩾ μl in peripheral blood. the priming of extracorporeal circuit was made with compatible, irradiated, leucodepleted packed red cell to avoid hypovolemic state. citrate dextrose solution a(acd-a), with a ratio of : to whole blood, and a bolus of heparin ui/kg were used as anticoagulants. all patients, treated without sedation, were monitored by ecg, pulse oximetry and non invasive blood pressure; electrolytes panel (na, k, ca) and act (activated coagulation time) were assessed at the beginning, minutes after and then every hour during apheresis. hypocalcemia was managed by mg calcium gluconate slow infusion. we report our experience of pbsc collection in low body weight children ( o . kg) treated in our apheresis department between january and november . a total of pbsc collections were performed in children ( m/ f, median age months, median weight . kg) affected by medulloblastoma (n = ), germ cell tumor (n = ), neuroblastoma (n = ), retinoblastoma (n = ), brain cancer (n = ). total blood volume processed ranged from . to . tbv (median . ) and median count of cd + collected was . × /kg(range: . - ). all procedures were performed with a median duration time of minutes (range: - min) and no serious adverse events occurred. in our experience pbsc collection is safe and feasible also in low body weight children using a tailored apheresis procedure. disclosure of conflict of interest: none. plerixafor on demand in the first or in the second attempt of cd mobilization j romejko-jarosinska, e paszkiewicz-kozik, l targonski, m szymanski, z pojda and j walewski high-dose chemotherapy and autologous hematopoietic cell transplantation (auto-hct) is a recommended strategy for patients with relapsed, refractory or high risk lymphoma. mobilization failure of cd + cells after granulocyte colonystimulating factor (g-scf) with or without chemotherapy is a factor limiting patient access to this potentially curative procedure. the use of plerixafor with g-csf may improve cd + cell harvest in poor mobilizing patients. we evaluated the clinical effectiveness of plerixafor and g-csf ± chemotherapy administered on demand in the first and second attempt of mobilization in lymphoma or myeloma patients who were eligible for auto-hct. we evaluated data on consecutive patients with hodgkin lymphoma ( ), dlbcl ( ), mantle cell lymphoma ( ) , myeloma ( ) and other lymphoma subtypes ( ) who were mobilized with plerixafor between january and october . median (range) age of patients was ( - ). patients received a median of ( - ) chemotherapy lines. radiotherapy was applied in patients. all patients received g-csf ( μg/kg/day) ± chemotherapy and plerixafor ( μg/kg/day) on demand in the absence of increase in the number of cd + cells in peripheral blood above /μl on the day of the scheduled apheresis (within days following the chemotherapy and after at least days of g-csf). plerixafor was given to patients in the first attempt of mobilization and to patients during the second mobilization. the mobilization was considered effective if the harvest cell dose was × /kg cd or more. after plerixafor administration circulating cd + cells increased to /μl in patients ( %) and in patients ( %) in the first and in the second mobilization, respectively (p = . ). the cd + cell collection was performed in / patients ( %): in / ( %) patients in the first and in / patients ( %) during the second mobilization cycle. the median number of apheresis was (range: - ), for both mobilizing cycles. the median (range) cd cell dose collected in the first and second cycle was . (range: . - . ) × /kg and . (range: - . ) × /kg, respectively (p = . ). the harvest was successful in / patients ( %) in the first and in / patients ( %) in the second cycle (p = . ). three patients ( %) who failed the collection with plerixafor in the first attempt, succeeded in the second cycle. additional second mobilization with plerixafor was successful in five patients ( %) who failed the first mobilization. in total, / ( %) and / ( %) of patients given plerixafor in the first or in the second mobilizing cycle harvested at least a minimum cd cell dose for auto-hct (p = . ). these results show that plerixafor administered on demand is an effective rescue strategy for poor mobilizing patients. each mobilization cycle with plerixafor resulted in the increase of circulating cd cell count. successful harvest is more frequent if plerixafor is administered in the first than in the second mobilization attempt. the evaluation of the prognostic factors for mobilizing failure with plerixafor is necessary to identify the poor mobilizers precisely. disclosure of conflict of interest: jr-j, ep-k, lt and jw: sanofi (travel grants); ms and zp: none; jw: lecture, honoraria cryopreserved stem cell grafts are still widely used both in the autologous or allogeneic settings. cryopreserved grafts can be thawed at the bedside or thawed and washed at the cell therapy laboratory. we recently reported that post-thaw washing did not impair hematopoietic engraftment, in a cohort of autologous transplanted patients receiving either unwashed or washed grafts (calmels b et al, bone marrow transplant. ). post-thaw washing can be implemented using various methods such as manual centrifugation, automated centrifuge-based (sepax , biosafe) or spinningmembrane devices such as lovo (fresenius kabi). we here report a step by step implementation of the lovo biomedical device (bmd) for washing thawed stem cell grafts. having defined a washing program, we aim to compare this protocol to our routine process, using the sepax bmd. we took advantage of apheresis products intended for destruction and cryopreserved in identical bags; after dry-thawing (plasmatherm, barkey), bags were connected to the sepax or to the lovo bmd, diluted volume to volume with + - °c % hydroxyethylstarch / . (voluven, fresenius kabi) and processed using the smartwash program (sepax ) or a cycles standard wash protocol on lovo (a cycle referring to one pass through the spinning membrane). the lovo settings were customized for this application: reduction retentate pump rate ml/min, desired inlet pcv %, and automated volume to volume dilution. after processing, cd and cd absolute counts and viability were evaluated by single platform flow cytometry (stem-kit, beckman coulter) and dmso was quantified by capillary zone electrophoresis (p/ace, beckman coulter). post-wash data show comparable cd + cell recovery, viability and effective dmso depletion. we conclude that lovo enables high efficiency dmso depletion while preserving optimal cd viability and recovery. comparison with sepax , a widely used automated centrifugebased device, reveals comparable efficiency. moreover, the length of the procedure when using the lovo does not significantly delay the process as compared to bedside thawing. we are currently evaluating lovo for the processing of multiple bags and higher cell contents, due to its ability to concentrate large volumes of cells suspension. post-thaw washing using automated cell processing systems have thus to be preferred over bedside thawing, since they provide multiple benefits including a short processing time, efficient dmso and cell debris removal, precise determination of infused cd + cell dose, and improved cellular stability. [p ] disclosure of conflict of interest: none. using bone marrow (bm) as the graft source results in lower graft-versus-host disease incidence, which is particularity important in haploidentical (haplo) stem cell transplantations (sct). nonetheless achieving adequate cd + cell count might be complicated in cases of donor-recipient weight differences. priming with g-csf may partly solve this problem. also there are reports of immunomodulatory effect of bm priming. in the retrospective study we have evaluate the effect of priming on stem cell yield and the outcomes of sct. patients and methods: patients with primed bm graft were matched in the ratio : to non-primed grafts. the criteria for matching were type of the donor, age of the recipient, underlying disease and disease status at the time of sct. priming was performed with three injections of filgrastim - mcg/kg daily for days prior to bm harvesting. median recipient age was years (range: - ). % of patients received the graft from haplo donor, % from matched related donor (mrd). % had acute lymphoblastic leukemia, % had acute myeloid leukemia, % had aplastic anemia, % had other malignancies. % were classified as salvage patients. % received myeloablative conditioning, % received reduced intensity. post-transplantation cyclophosphamide (ptcy) was used as graft-versus-host disease prophylaxis in % of patients. results: the yield of cd + × cells /kg of recipient weight was only non-significantly higher in the priming group: . ± . vs . ± . , p = . . the yield of cd + cells per kg of donor weight was also not different: . ± . vs . ± . , p = . . there was no difference in the incidence of primary graft failure ( % vs %, p = . ). median time to neutrophil ( vs days, p = . ) and platelet ( vs days, p = . ) engraftment was shorter in nonpriming group. there was no differences between priming and non-priming groups in the incidence of acute grade ii-iv gvhd ( % vs %, p = . ), moderate and severe chronic gvhd ( % vs %, p = . ), -year non-relapse mortality ( % vs %, p = . ), relapse incidence ( % vs %, p = . ), overall survival ( % vs %, p = . ), event-free survival ( % vs %, p = . ) and gvhd-relapsefree survival ( % vs %, p = . ). conclusions: priming of the bone marrow with reported schedule did not result in higher cd + cell yield and was not associated with any differences in the outcomes of sct. nonetheless, these results should be interpreted with caution, because our study included large proportion of pediatric patients, patients with active disease and ptcy as gvhd prophylaxis, and they may not translate to the other groups of patients. disclosure of conflict of interest: none. priming with granulocyte-colony stimulating factor preserves the contents and abundant ifn-γ production capacity of γδ t cells z bian , q fu , m huo , xj huang and j liu peking university people's hospital the increasing evidences indicate that removal of αβ t-cell and b-cell from grafts was efficient and reproducible in allogeneic hematopoietic stem cell transplantation (allohsct). γδ t cell is one of the functional subpopulations preserved by this graft manipulation and supposed to play a role in improving the transplant outcomes. thus, comprehensive understanding the subsets and functional capacities of γδ t cells in graft becomes important. although there is increased attention paid on this special t-lymphocyte subpopulation, the contents and cytokine production capacities of peripheral γδ t cells before and after granulocyte-colony stimulating factor (g-csf) mobilization for allohsct have not been reported. peripheral blood (pb) before g-csf treatment, g-csf-primed pb and bone marrow (bm) grafts were obtained from healthy donors. the proportions of total γδ t cells and various γδ t-cell subsets were detected by flow cytometry. furthermore, effects of g-csf on the contents and cytokines production by γδ t-cell subsets were also determined. the percentages of most γδ t-cell subsets including cd +, cd -, vδ +, vδ +cd +, vδ +cd -, vδ +, vδ +cd +, vδ +cd -, and non-vδ /δ were preserved in the g-csf-primed pb grafts compared with those before g-csf mobilization. interestingly, we found that peripheral γδ t cells and various subsets all predominantly expressed ifn-γ in response to stimulation. this abundant ifn-γ production capacity of peripheral γδ t cells were maintained after g-csf treatment. in contrast, production of il- by γδ t cell and its subsets were decreased in the same context. priming with g-csf preserved the contents and abundant ifn-γ production capacity of γδ t cells. our data suggests a reasonable role of γδ t cells in preventing from allohsct associated complications and may help establish an effective γδ t cell-based immunotherapeutic approach to improve the overall survival of allohsct. disclosure of conflict of interest: none. processing of hematopoietic stem cells grafts: towards automation of cryopreservation/thawing steps a-l chateau , j gaude , c malenfant , a autret , c lemarie , c chabannon and b calmels centre de thérapie cellulaire-institut paoli-calmettes and unité de biostatistiques-institut paoli-calmettes autologous hematopoietic stem cells (hsc) support is still widely used to allow for high-dose chemotherapy in the context of myeloma and lymphoma treatment. in the autologous setting, mobilized aphereses are systematically cryopreserved. currently, cryopreservation and subsequent thawing rely on manual and largely operator-dependent processes such as manual addition of dmso for cryopreservation or thawing in standard water baths. these operations are thus hampered by significant intra-and inter-facility variability and have to be replaced whenever possible with automated and harmonized processes. the aim of our study was to evaluate a recent, versatile device: smartmax (biosafe, eysins, switzerland), based on the peltier-seebeck effect, for its ability to automatically add the dmso-containing solution to the cell product and to thaw hsc bags. we thus compared three different cryopreservation/thawing protocols ( figure ). we first evaluated the use of the smartmax at the thawing step by comparing cryopreserved apheresis products thawed using our routine device: the plasmatherm (barkey), an automated dry-thawing device that contains water (protocol a), with products thawed with the smartmax (protocol b); after thawing, all products were washed using the smartwash program of the sepax (biosafe). we then evaluated the smartmax for its ability to automatically add the dmso solution: autologous grafts were processed with the smartmax, both for cryopreservation and thawing (protocol c); we compared these ‛fully automated processes' to apheresis processed with protocol b. absolute cd + and cd + cell counts and viability were measured before cryopreservation and after washing using single platform flow cytometry. for all three protocols, the quality of the collected product was comparable in terms of median cd + cell and neutrophil contents. when comparing protocols a and b, viable cd + cell recovery after thawing and washing was slightly lower in the smartmax group ( %) as compared to the plasmatherm group ( %, p = . ). when comparing protocols b and c, viable cd + recovery was comparable (p = . ) when the cryopreservation solution was automatically added by the smartmax ( %), as compared to the manual technique ( %). these preliminary data need to be validated on larger numbers of procedures, however suggest that smartmax use can safely be substituted both to the manual addition of the cryoprotectant and to the traditional thawing step in water baths; potential advantages include complete water removal from sensitive clean rooms and gmp environments. full automation of previously manual and operatordependent technical processes will ultimately allow for improved standardization and reproducibility across cell processing facilities. [p ] disclosure of conflict of interest: none. reduced efficacy of mobilisation using gdp compared to ive a hunter, w merrison, am martin, k hodgson, f miall, r moore and r lewin university hospitals of leicester, nhs trust the use of ive ± rituximab for relapsed/refractory disease in lymphoma is well established. stem cell mobilisation using g-csf post ive administration has been the standard of care in our unit for years. recent interest in cisplatnin-based treatments has seen a change in practice with the use of gdp ± rituximab increasingly common. we have assessed the success of stem cell mobilisation post gdp and compared it to ive using g-csf. patients were eligible for augmentation with plerixafor if their peripheral blood cd levels were between - × cells/l at the time of collection. from sept to oct patients with progressive or relapsed lymphoma underwent stem cell collection. patients characteristics: dlbcl, follicular and t-cell nhl. had a median age ( - years). received gdp, ive. overall % patients failed to mobilise a sufficient cd cell dose to proceed to hdt. all the patients who received ive mobilised successfully but / ( %) patients receiving gdp failed to mobilise. of the patients who did mobilise the average cd collection was higher in the patients who received ive . ( . - . ) and the number of apheresis procedures was lower, median ( - ) compared to . ( . - . ) and ( ), respectively, in the gdp group. patients in the gdp group who failed to mobilise were not eligible for plerixafor because cd levels were below × /l. taking age into account the median age in the ive group was higher ( - ) than the gdp group ( - ) and the lines of previous therapy were not different. patients who had successful stem cell collections went on to receive hdt with leam and all patients engrafted. in this small collection of patients we have experienced a higher failure of mobilisation post a cisplatnin-based protocol compared both to our historical controls pre plerixafor usage (data not shown) but also to current patients. further investigation is needed to ascertain the impact of cisplatnin on stem cell mobilisation and its impact of treatment strategies. disclosure of conflict of interest: none. single centre experience of zarziotm biosimilar granulocyte-colony stimulating factor (gcsf) for the mobilisation of healthy donors demonstrates good leukapheresis yields and safety profile at month median follow-up jg taylor , , t seddon , k alizadeh , c agrawal , l kempster , jg gribben , and sg agrawal , centre for haemato-oncology, barts cancer institute, dept. haemato-oncology, st bartholomew's hospital, london, uk and experimental pathology, blizard institute, queen mary university of london, uk biosimilars have led to significant improvements in the affordability of growth factors such as granulocyte-colony stimulating factor (gcsf). data has shown similar performance and efficiency to parent drugs but concern has been raised about their use in healthy donors due to lack of data examining adverse effects in this setting. we conducted a retrospective analysis investigating mobilisation and adverse effects in healthy sibling donors of adults undergoing an allogeneic haematopoietic stem cell transplant at st bartholomew's hospital from to . harvest data were gathered from hospital records. adverse effects data were gathered from hospital records and telephone follow up. % of donors were male with a median age at harvest of ( - ). all donors were mobilised using zarziotm biosimilar gcsf at a dose of μg/kg/day. median number of apheresis required was ( ) ( ) ( ) . median cd + cell count was . × /kg bodyweight ( . - . ) with × cd +/μl ( - ) in peripheral blood. the target cd + count ( × /kg) was achieved in % of donors and an adequate yield ( - × /kg) in %. in four donors ( %), the harvest was deemed to have been unsuccessful as the cd + count was o × /kg. the patients with donor harvest yields o × /kg proceeded to transplant; all four patients engrafted and one patient had mixed chimerism at day but was fully donor by day . median cd + cell count was . × /kg bodyweight ( . - . ) . median days to neutrophil engraftment ( . × /l) was . median days to platelet engraftment ( × /l) was ( - ) with one patient never engrafting. forty ( %) of donors were contacted at a median of months ( - ) post mobilisation to establish incidence of adverse effects. three donors were uncontactable as they had moved overseas. eight donors were not contacted to avoid distress as their sibling had died since transplant. among contacted donors . % reported side effects including bone and lower back pain controlled with analgesia, constipation and low mood. other side effects included chest pain which was considered to be musculoskeletal in origin on day of gcsf administration associated with taking an increased dose due to patient error (n = ) and abdominal contractions like labour while receiving gcsf (n = ). three ( . %) reported side effects lasting beyond one month post mobilisation: lower back pain lasting months (n = ), fatigue of months duration (n = ), and cough of months duration (n = ). our data demonstrates good mobilisation using μg/kg/day zarziotm biosimilar gcsf without significant adverse effects at years median follow up. this supports its ongoing use for the mobilisation of healthy donors. disclosure of conflict of interest: sga has received honoraria from sandoz and grant support from sandoz and amgen. stem cell mobilization in poor mobilizers with multiple myeloma (mm) or non-hodgkin lymphoma (nhl) before and after introduction of plerixafor: single center comparative analysis using a cost-efficient single fixed-dose schedule r wäsch , c greil , c kiote-schmidt , s hildenbeutel , k kühbach , r bosse , j duyster and m engelhardt department of hematology, oncology and stem cell transplantation, university medical center, freiburg, germany collection of hematopoietic stem cells (hsc) from the peripheral blood (pb) is routinely conducted prior to highdose chemotherapy and autologous transplantation. despite safety and efficiency of current apheresis procedures including mobilizing chemotherapy and granulocyte colony-stimulating factor (g-csf), there is a significant rate of mobilization failures due to different patient-dependent factors necessitating additional agents like plerixafor. while plerixafor is approved for patients with mm or nhl based on prospective studies using steady state mobilization with g-csf − /+ plerixafor, prospective studies using chemo-mobilization are lacking. here we compared the outcome of poor mobilizer from the pre-plerixafor era with poor mobilizers who received additional plerixafor in a real world analysis. we analyzed consecutive patients with mm or nhl who were mobilized at our academic center between and and received plerixafor, because they were expected to be poor mobilizers, due to . low counts of cd + cells in pb samples prior to apheresis, . after a first apheresis day with insufficient yield or . as a rescue strategy after insufficient harvest with previous mobilizing chemotherapy (greil c,…engelhardt m, wäsch r. leukemia & lymphoma , in press). we examined cd + cell counts in pb and in apheresis products to identify those patients who were able to collect a sufficient cd + cell count for transplantation after application of plerixafor. we compared these data with consecutive poor mobilizers from the pre-plerixafor era, who were mobilized between and without plerixafor. the median pb cd +/μl count at first apheresis was significantly higher after the first dose of plerixafor when compared to the pre-plerixafor group with . vs . (p o . ). accordingly, the median collected cd + cells/d (× /kg bw) and total cd + cells (× /kg bw) were significantly increased with . vs . (p o . ) and . vs . (p o . ), respectively. the rate of × cd + cells/kg bw in first apheresis (%) increased from % in the pre-plerixafor era group to % after the first dose of plerixafor in the plerixafor group. consistently, the successful transplantation rate increased from % in the preplerixafor group to % in the plerixafor group. successful stem cell mobilization could be achieved with only a single fixed-dose of plerixafor in % of poor mobilizers as previously reported by our group. the addition of plerixafor to chemomobilization in poor mobilizers with mm or nhl significantly increased pb cd +/μl counts, apheresis yields and transplantation rates when compared to poor mobilizers from the pre-plerixafor era. these favorable apheresis results can be obtained using our cost-efficient, single fixed-dose plerixafor schedule in the majority of the patients leading to a % transplantation rate in poor mobilizer. disclosure of conflict of interest: rw received research funding, advisory and speaker's honoraria from sanofi-aventis. high-dose chemotherapy followed by autologous peripheral blood stem cells transplantation (pbsct) is the standard of treatment for patients with hematological malignancies. recombinant granulocyte colony-stimulating factors (g-csfs) are widely used alone or in combination with chemotherapy, in order to mobilize patient's stem cells (cd +) for autologous and allogeneic peripheral blood stem cells transplantation. aim: the aim of our study was to compare effectiveness and safety of different biosimilar products of filgrastim used in autologous pbsc mobilization in patients with hematological malignancies. our retrospective analysis included patients ( women and men) with median age years ( range: ,who underwent the procedure of autologous pbsct in years - in the haematology, blood neoplasms, and bone marrow transplantation clinic of medical university in wrocław. there were three different biosimilar products of filgrastim used: tevagrastim (teva) in patients, nivestim (hospira) in patients and zarzio (sandoz) in patients. ( %) patients were diagnosed with plasma cell neoplasms, ( %) with hodgkin's and non-hodgkin's lymphomas, ( %) patients had acute myeloid leukemia and ( %) had other hematological malignancies. statistical analysis was conducted using statistica (statsoft polska) statistical software. for quantitative variables arithmetic means and standard deviations were calculated for the estimated parameters in the studied groups. distribution of variables was tested using w-shapiro-wilk test. p . ). there were also small variations in the mean number of leukapheresis necessary to obtain the minimum cd + cell count: . in zarzio group, . in nivestim group and . in tevagrastim group. however, there were no difference between biosimilar g-csfs. the highest rate of successful mobilizations (defined as × /kg cd + cells collected) was observed in . % patients received zarzio, in . % received nivestim and in . % patients received tevagrastim. the safety profile was comparable between the biosimilar g-csf and included bone pain in ( %) patients and headache in ( %) patients. the results are shown in table . all three used biosimilar g-csfs demonstrated similar efficacy and safety in stem cell mobilization in patients with hematological malignancies. therefore, it seems that all the analyzed products can be used interchangeably. presented observations should be verified with wider prospective research. [p ] disclosure of conflict of interest: none. use of g-csf stimulation of bmt donors might prove to be beneficial in many respects, improving tnc yield but also through immunomodulatory effect on donor t cell function and apcs . we analyzed outcomes of consecutive patients receiving bone marrow transplants from hla-haploidentical related donors that received g-csf stimulation prior to harvest. fourteen patients received hla-haploidentical bmt with pt-cy between / and / . five donors were siblings, children, mothers and father. donors received g-csf at the dose of mcg/kg bw sc. on days − , − and before bm collection. twelve patients received nonmyeloablative conditioning according to baltimore protocol , while two patients received myeloablative conditioning (bucy). along with post-transplantation cyclophosphamide, all patients received tacrolimus and mmf form day + , as described earlier . median age was years (range: - ), female and male patients. eight patients had aml, cml, mh and one all. ten of them were in remission, while mh patients were in pr, and aml patients had residual disease as evident by immunophenotyping. median number of infused tnc was . × /kg bw (range: . - . ); cd + cells . × /kg bw (range: - . ) and cd + cells . × /kg bw (range: . - . ). median follow up was days (range: - ). eleven patients engrafted ( %), one patient had primary rejection, one had overt disease relapse at day + and one patient died in aplasia due to sepsis. median day to neutrophil recovery (anc . × /l) was (range: - ), median days to platelet recovery (plt × /l) was (range: - ). in all patients mmf was discontinued at d + . two patients developed acute gvhd in our cohort ( %), one after receiving dli for falling chimerism at day + . one patient ( %) developed chronic gvhd, after having received dli due to disease progression. at the time of analysis patients are evaluable; patients had disease relapse/progression ( %), patients are alive and in remission. one patient died due to sepsis in aplasia (accounting for % non-relapse mortality). one patient that rejected the graft was transplanted again from the same donor, using myeloablative conditioning and peripheral stem cells as graft source and engrafted. overall survival median is . years, with significantly shorter survival if patient was not in complete remission at time of transplant (p o . ). even though the experience with g-csf mobilized bm graft in the hlahaploidentical setting with pt-cy is relatively small, in our series it has been beneficial in terms of tnc yield. also, the incidence of acute and chronic gvhd in our patients has been low, particularly agvhd with one case developing only after dli. whether the observation is the result of limited number of patients, or it reflects the immunomodulatory effect of g-csf on bm graft as previously suggested remains to be seen as further studies are warranted. autologous transplantation of haematopoietic stem cells (ahsct) is usually perceived as a fully standardized and safe procedure; however, a minority of patients experience a delayed engraftment and seldom even an engraftment failure, possibly related to a poor quality of the graft. therefore the current policy in many centers is aimed to increase the target dose of collected cd + cells up to an ‛optimal' level of × /kg. plerixafor was introduced in the clinical practice to maximize the mobilization of hsc, in order to collect an optimal number of cd + cells in a limited number of collections also in poor and slow mobilisers. we carried out a retrospective analysis of our case series aimed to individuate mobilization predictors optimize the ‛on demand' use of plerixafor. we analyzed patients who underwent mobilization with cyclophophamide ( g/sqm) and filgrastim mcg/kg from + in our unit from and . diagnosis were multiple myeloma (mm) ( . %), non-hodgkin lymphoma (nhl) ( . %), hodgkin lymphoma (hdg) ( . %) and ( . %) autoimmune disease (ms . %; ssc . %). median age (range) was years ; male/female ratio / . circulating cd + cell count was started at white blood cells (wbc) recovery, which was defined as the first day when their count exceeded × /l. the primary goal was to identify at wbc recovery one or more factors predicting a suboptimal mobilization, which was defined as the failure to exceed cd +/mcl circulating cells in the day after the wbc recovery. patients were excluded from this analysis if ) showed a cd + count /mcl at wbc recovery (very good mobilizers) and/or ) had received plerixafor and/or ) did not proceed to another cd + count the day after wbc recovery. binary logistic regression was used to obtain the factors that increased the odds for an optimal mobilization. overall out ( . %) patients were shown as very good mobilisers as their cd + count exceeded /mcl at wbc recovery. on the remaining , were excluded for the lack of a second assessment and for the lack of data. among the remaining patients, the threshold of cd +/mcl cells on the second day was reached by ( . %) of patients (group a) while the remaining ( . %) failed the goal (group b). median (range) wbc × /l and cd +/mcl counts in group a and b at wbc recovery were . ( - . ) and . ( . - . ) and . ( - ) and . ( - ), respectively, with a statistically significant differences among group (mann-whitney u test with p = . and p = . , respectively). wbc (or = . ; % ci: . - . ) and cd +/mcl (or = . ; % ci: . - . ) in first day count, but not gender, disease category and time from mobilization chemotherapy to first cd + count, were predictors of optimal mobilization. combining these two predictors we found that wbc/cd + ratio has a sensitivity of . % with an auc . in roc analysis. assessment of circulating wbc, cd + and their ratio at wbc recovery in a chemo-based mobilization strategy can predict sub-optimal mobilization of hsc and support the decision of adding plerixafor. these data will be prospectively validated in a broader set of patients. disclosure of conflict of interest: none. human platelet lysate (hpl) is rich in growth factors (gf) and nutritive elements and represents a powerful xeno-free alternative to fetal bovine serum (fbs) notably for mesenchymal stem cell (hmsc) proliferation. however, there is a large variability in hpl preparations (various sources, use of different and non-standardized production protocols, with variable and limited number of donors), resulting in discrepancies in product quality, low management of product safety and poor batch-to-batch standardization. we describe here the development and the characterization of a standardized hpl prepared from outdated transfusional grade screened normal human donor platelet concentrates (pcs), manufactured on an industrial scale (batch size of donors) and following a highly qualified process (clean room, trained operators, validated aseptic filtration). pcs were frozen at − °c and thawed at + °c to lyse platelets. cell debris were removed by centrifugation and the supernatant (hpl) was recovered. clinical grade l batches of aseptic filtered hpl were characterized. first, we showed that hpl prepared from a limited number of donors displayed a variability in terms of gf contents. on the contrary, we observed a robust standardization between industrial batches of hpl ( donors) in terms of gf contents (bfgf, egf, vegf, pdgf-ab, tgf-beta and igf- ), biochemical analyses (total proteins, albumin, fibrinogen, vitamins and iron) and efficacy on bone marrow (bm)-hmsc proliferation. secondly, we compared expansion and functional characteristics of bm-hmscs grown in clinical grade hpl vs msc-screened fbs batches. we showed a reproducible increase in cell growth kinetics using hpl, a maintenance of bm-hmsc clonogenic potential and membrane marker expression (with however a strong overexpression of cd ). we observed a similar adipogenic and osteogenic differentiation potential and finally that immunosuppressive properties of bm-hmscs (inhibition of t-cell proliferation) cultivated in parallel in both conditions also remained identical. finally, we demonstrated the stability over time of hpl stored at − °c and − °c. in conclusion, we demonstrated the feasibility to use a standardized, characterized, efficient and clinical grade hpl for research and cell therapy applications. disclosure of conflict of interest: sv, se, lc, pb, tb, al, fg and bd are employees of macopharma. previously published p alpha/beta t cell depleted donor lymphocyte infusion m karakukcu, e Ünal, l kaynar, s Özcan, g tezcan karasu and mb acar the main objective of this project is to improve a safe and efficient new donor lymphocyte infusion (dli) with depletion of αβ+ t cells which cause graft versus host disease (gvhd), and enrichment of anti-leukemic γδ+ t cells, nk cells and dendritic cells to build an effective and permanent anti-tumor effects for patients relapsed hematological cancers after allogeneic hematopoietic stem cell (hsc) transplantation who have blasts and mixed chimerism. this study is conducted with collaboration of erciyes university pediatric and adult hsct units, and bahcesehir university, medical park hospital pediatric hsct unit. the tcr αβ+ t cell depleted dli product that is used in the study was collected and separated at erciyes university apheresis unit. the cell contents obtained for tcr αβ+ t cell depleted dli used for patients were cd cells were reduced to . - . × cells/kg, γδ+ t cells were reduced to . - . × cells/kg, αβ+ t cells were reduced by . %, and were obtained at - cells/kg. a total of patients ( female, male) were included in the study, consisting of an adult and children. nine patients had hematological malignancies. five patients were referred for all, three for aml, one for mds and one for griscelli syndrome. efficiency: the clinical response to the αβ+ t cell depleted dli treatment was achieved in / patients ( %). in these patients, although the increase of chimerism was limited in patients, no recurrence was occurred. one of the two patients who previously responded to the treatment but experience of decreasing chimerism had relapsed after months, and months later. one of these two patients died after relapse. the other was managed by the second transplant. the most important objective of this study was to show that αβ + t cell depleted dli treatment is reliable. none of the patient showed severe gvhd except one patient with mild grade ii gvhd. despite the presence of severe gvhd after hsct in two patients, reactivation for gvhd was not observed after treatment with αβ+ t cell depleted dli. none of the patients had a bone marrow aplasia. as a result, αβ + t cell depleted dli treatment seems to be highly safe, and effective in selected patients. disclosure of conflict of interest: none. hematopoetic stem cell transplantation (hsct) is associated with several potentially lethal complications; for example, relapse of the malignant disease, graft rejection, infectious complications and graft versus host disease (gvhd). higher levels of cd + cells in the graft have clearly been associated with increased risk of gvhd, but also superior gvl effect and less infectious complications. to tackle post-transplant complications such as graft failure and relapse, donor lymphocyte infusion (dli) have successfully been used for decades but with an associated risk of gvhd. to decrease the risk of gvhd but still use facilitating cells in the cell product we performed αβ depletion of grafts for use as stem cell booster after allogeneic hsct to treat infections or poor immune reconstitution. in this study, patients were infused post-hsct with αβ t-cell depleted grafts. the indication for infusion of αβ t-cell depleted graft in all patients was poor immune reconstitution with associated infectious complications. for all patients, the original hsct donor was used for the αβ t-cell depleted boost. to characterize the αβ-depleted stem cell grafts, samples were stained for various cellular subsets and analyzed by flow cytometry. we could show a median log depletion of αβ cells of . and a median yield of γδ t-cells (%) of . . the median cd + cell dose (× /kg) was . . all patients were alive months after infusion. after year only one patient succumbed. despite that the majority of patients suffered from agvhd grade or before infusion of αβ t-cell depleted graft none showed increased symptoms afterwards. in more than % of the patients there was a increase in granulocytes, thrombocytes and white blood cells months after infusion. in conclusion, we describe the use of αβ t-cell depleted grafts as stem cell booster in patients suffering infectious complications due to graft failure after hsct with encouraging results. disclosure of conflict of interest: none. delayed engraftment or graft failure still remains a concern in bone marrow transplantation (bmt). graft composition may predict engraftment after infusion. this study aims to determine which quality control parameters used for the characterization of bone marrow grafts are the most predictive in order to minimize the risk of engraftment delay or graft failure. we conducted a multicenter retrospective study in pediatric patients who underwent first allogenic bmt at two centers in barcelona (catalonia, spain) between and . quantitative variables considered for the study were: total nucleated cells (tnc), mononucleated cells (mnc), cd + cells, cd + cells and granulocyte-monocyte (gm) colonies enumeration. qualitative variables considered for the study were viability assessed by flow cytometry and clonogenic efficiency of the cd + cells (clonegm) which is the ratio between gm colonies and cd + cells. patients were included (median age (range) were years old ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ). the median tnc(range) was . e /kg ( . - . e /kg) and . e /kg ( . - . e /kg) for mononuclear cells (mnc). on the other hand, the median (range) cd + cell dose was . e /kg ( . - . e /kg) and t-cell dose (cd +) was . e /kg ( . - . e /kg). the median (range) colonyforming unit granulocyte macrophage (gm/kg) dose was . e /kg ( . - . e /kg). the median (range) of cd + cell viability, was % ( - %) and the median(range) of the clonogenic potential of cd + cells (clonegm) was . % ( . - . %). the median (range) of engraftment was days for neutrophils and ( - ) days for platelets. patient was considered as primary graft failure. in the univariate analysis, cd + (p = . ) and mnc (p = . ) cell dose predicted a faster neutrophil engraftment and female donor a slower neutrophil and platelet engraftment (p = . and p = . ). cell viability also correlated to a better platelet engraftment (p = . ). in the multivariate analysis we observed a trend for a faster neutrophil recovery according cd + cells infused. again, female donor was associated with slower engraftment. in order to establish a safety threshold, we did a quartile analysis of cd dose and found . e /kg (quartile ) discriminates a faster neutrophil engraftment [median days vs days for those with higher cd + cells (p = . )]. in conclusion, we found an association between mnc and cd + cell dose and time to engraft, and established a safety threshold of . e cd +/kg. also, bm grafts from female donors were associated with slower engraftment. no other qualitative parameters were predictive of engraftment. disclosure of conflict of interest: none. plasma cell myeloma (pcm) is currently treated with chemotherapy and autologous stem cell transplantation (asct) but relapse rates remain high. adoptive transfer of mature haploidentical natural killer (nk) cells is a promising approach to provide pcm patients with highly immunocompetent effector cells with anti-myeloma function early post transplantation. here we report on the current clinical phase i/ii trial of multiple preemptive infusions of good manufacturing practice (gmp) expanded nk cells to pcm patients (clinicaltrials.gov nct ). ten pcm patients were recruited (seven males, three females, median age: y). all patients received four cycles of vtd chemotherapy (reaching cr: × , vgpr: × and pr: × ) before high dose therapy with melphalan mg/m and asct. after successful stem cell mobilization and cryopreservation of patients' stem cells after the third vtd cycle, nk cells from haploidentical family donors were purified from unstimulated leukapheresis by t cell depletion and nk cell selection using clinimacs. highly pure nk cells (mean: . × cells) were obtained with a minimal t cell contamination corresponding to a . log t cell depletion. nk cells were expanded ex vivo for days in gmp-medium containing autologous irradiated feeder and interleukin- and - . nk cell numbers increased -fold (range: - -fold). in three nk cell products t cell contents were × cells/kg body weight (bw: × above limit of clinical trial) and were successfully reduced by °cd -depletion to . × cells/kg bw. nk cell products were cryopreserved in escalating doses ( . × , . × and rest as multiple doses of maximal . × cells/ kg bw). the pcm patients received - × expanded nk cells (median: . × cells/kg bw, range: . - . × cells/ kg bw) as - infusions (median. dlis). the nk-dlis were administered between day and after asct and were well tolerated without any acute adverse events. no signs of acute or chronic graft-versus-host disease were observed in any of the patients after a total of nk-dlis. engraftment occurred between days - (median: days). infused donor nk cells were monitored by short-tandem repeats pcr. donor nk cells were detected in peripheral blood one and h post infusion (% donor nk of enriched blood nk cells: mean: %, range: - %, and mean: %, range: - %, respectively) indicating significant nk cell survival in recipients in the absence of il- support in vivo. clinical responses at last follow-up compared to a retrospective cohort of matched control patients will be presented. these results demonstrate the feasibility of large-scale gmp expansion and safety and immunotherapy with third-party leukemia-specific t cells (leuk-sts) represents an attractive approach for acute leukemia (al) patients lacking a fully matched donor or relapsing after allogeneic hematopoietic cell transplantation (hct). its application however, is limited by the demand for high numbers of antigen presenting cells (apcs), capable to produce clinically relevant numbers of leuk-sts. low volume, non-transplantable cord blood units (cbus) could theoretically serve as an easily accessible source to generate high numbers of dendritic cells (dcs) and subsequently leuk-sts, providing also the advantage of reduced alloreactivity, even in cases of partial matching. our goal was to generate clinically relevant doses of leuk-sts targeting al-related antigens, the wilms tumor protein (wt ) and the preferentially expressed antigen in melanoma (prame), through the exploitation of non-transplantable cbus. to generate dcs, immunomagnetically enriched cd + cells from cbus ⩽ ml were cultured in g-rex devices in the presence of scf, gm-csf and il- . dcs matured by toll-like receptor ligand and / were immunophenotypically characterized by flow cytometry (fcm). secreted cytokines were measured with elisa. matured dcs were activated with a peptide-mix of wt and prame and used as apcs to repeatedly stimulate naive t-cells (derived from the cd fraction of the same cbu). the phenotype and the specificity of generated leuk-sts were determined by fcm and ifn-γ/ elispot, respectively. starting from mean . × ± . × cd + cells, from non-usable cbus, we generated . × (range: . - . × ) myeloid dcs (cd +/cd c+: . ± . %) in days (fold change~ . ). the produced cells highly expressed maturation markers (cd +/cd +: ± %; cd c+/hla-dr+: ± %) and secreted high levels of th cytokines (Ιl- : ± pg/ml; il- : . ± . × pg/ml, tnf-α: ± pg/ml) and low levels of the th -cytokine, il- . the average number of cd -cell-derived leuk-sts after week-culture was . ± . × (~ logs above clinical doses). the produced cells were enriched in cd + polyclonal cells ( ± %), comprising of cd + ( ± %) and predominantly cd + cells ( ± %), expressing effector memory (cd ra − /cd l − : . ± %) and effector memory ra markers (temra: cd ra+/cd l − : ± %), while containing insignificant numbers of cd +/cd +cells ( ± . %). specificity was seen after the second stimulation at the earliest and was increasing after each stimulation [mean spot forming cells (sfc)/ × cells at second, third, fourth stimulation: ± ; ± ; ± ; respectively]. in particular, produced cells were highly specific for both targeted antigens (prame: ± , wt : ± ), while they expressed low the programmed cell death protein- (cd +/pd- +: ± %), implicating absence of cell exhaustion after repeated stimulations. we report a paradigm of ‛circular economy' in science, by the exploitation of non-usable cbus, towards scalable generation of cb-cd +-cell-derived dcs and cb-cd -cellderived leuk-sts from the same cbu and establishment of leuk-sts banks. whether similarly produced leuk-sts could significantly advance the treatment of al or leukemic relapse after hct, will be ultimately determined in vivo. disclosure of conflict of interest: none. comparison of two different methods to generate antifungal-specific t-cells under pre-clinical-scale conditions r geyeregger , s tischer , invasive infections with aspergillus fumigatus constitute a major cause of morbidity and mortality in immunocompromised patients after haematopoietic stem cell transplantation. although adoptive immunotherapies against viral pathogens are already in phase i/ii trials, clinical-grade methods for the generation of aspergillus-specific t-cells (asp-t-cells) from healthy transplant donors or even related or unrelated thirdparty donors are still under development. in this study, two different strategies interferon-gamma (ifn-g) cytokine capture system (ccs) vs short-term in vitro expansion (ste) were performed from the same healthy volunteers in order to evaluate the most suitable approaches for the in-time generation of clinical applicable asp-t-cells. pbmcs from leukapheresis of healthy donors (n = ) were first prepared in hannover for the ifn-g-ccs and then sent to vienna to prepare the ste. all donors belong to the allocell registry (www.allocell.com) of hannover medical school and the frequency of asp-t-cells was pretested by high-throughput ifn-g elispot assay. for the ifn-g-ccs, × cells were stimulated for h with gmp-conform aspergillus lysate followed by magnetic selection of ifn-g-producing t cells. cells were characterized for phenotype and function by multicolour flow cytometry. for the ste, × cells were cultured in g-rex devices and stimulated for days with either the aspergillus lysate alone or with pooled overlapping pepmixes (catb, crf , f , gel , pmp , shmt and sot) and il- . to further characterize the final cell products, multicolour flow cytometry, ifn-g elispot and ifn-g/granzyme b flurospot analyses were performed. ifn-g-ccs: frequency of ifn-g positive asp-t-cells pre-magnetic enrichment ranged between . and . %. recently we defined t-cell donors as eligible if ⩾ . % specific ifn-g+ t cells are detectable. the purity of asp-t-cells among cd + cells, obtained from three donors after magnetic selection was in mean % ± (range: - %). the absolute number of selected ifn-g+ cd + t-cells was ± . this could be approximately multiplied by a factor of , if × pbmcs are used for the generation of clinically applicable t cells using the ccs and the prodigy device. ste: after days, asp-t-cells (n = ) showed highly specific activity against the lysate (in mean ± spot forming colonies (sfc)/ cells) and pooled pepmixes (in mean ± sfc/ cells). in both methods (lysate vs pooled pepmixes), predominantly cd + t-cells were expanded ( % ± . vs % ± . of cd +) compared to cd + t-cells ( . % ± , vs . % ± . ). interestingly, whereas after ste, cd + t-cells include mainly central memory t-cells (mean %; cd l+cd ra − ), cd + t-cells include mainly effector memory t-cells ( %; cd l − cd ra − ). generated t cells were highly functional and cytotoxic as determined by the secretion of effector molecules granzyme b and ifn-g. based on the purity of up to % after the ifn-g-ccs and the high number of sfc received after ste with lysate and pepmixes, both methods seem to be suitable for clinicalscale productions. for patients who are in need for high asp-tcell numbers the application of first in-time ccs-purified asp-t-cells followed by the administration of ste cells might be a promising way to boost antigen-specific t-cell response. disclosure of conflict of interest: none. complete computerization of cell therapy product files (‛zero paper') in the qap software o christéle , r catherine , r aline , k mathias , m lavinia , d vincent , m jean-pierre and l marie-noelle , hematologie clinique et thérapie cellulaire-chu amiens picardie, simedia-ver, hématologie clinique et thérapie cellulaire-chu amiens picardie, simédia-ver, direction système informatique-chu amiens picardie, hématologie clinique et thérapie cellulaire, lacassagne and the computerized management of cell therapy products (ctp) is an obligation for processing laboratories to meet regulatory requirements. the software used is often independent of institutional systems in view of the specificity of cellular therapies and do not always allow the implementation of the ‛zero paper' policies that are being put in place. we report here our experience with the qap software (quality assurance partner) developed by the company simédia (www. qap .com) in open source (mit license) allowing the management of fully computerized ctp files. the qap software has been developed to ensure the traceability of ctp for both preparation and quality control by combining the product preparation environment (personnel, premises, reagents, consumables, equipment). initially, with the help of the company simédia, we parameterized the software in accordance with our procedures for the preparation and quality control of ctp. we built a file that we printed out for archiving on paper. it soon seemed necessary to reverse this mode of operation to add to the software the documents papers to obtain a file completely computerized and to avoid paper archiving. the close collaboration between the cell therapy laboratory staff, the software referent within the information system department of the amiens hospital and the company simédia enabled: set up a document backup server sufficiently proportionate in memory. have simedia carry out the necessary developments so that all documents can be integrated into the software, set up a coherent working circuit, organize the registration of documents, put in place a rigorous verification of the mandatory elements of the file. the reflection on the computer file made it possible to evolve the software to widen its use to all documents of management of the laboratory: maintenance of equipment, control of premises, housekeeping, staff training, quality control of automatons, reagents and consumables, process, reception, distribution. rigorous formalization was mandatory to ensure that the record was organized in a uniform manner. an intermediate paper record is still necessary for a period of about month: from the programming of the graft to the final validity of the injected product. this folder consists only of transient elements that cannot be integrated into the qap software immediately. the transition from the paper file to a computer file took place in several stages, calling into question our functioning. the difficulties of this implementation are of several natures: the heterogeneity of the documents components a cell therapy product file, the impossibility of benefiting from an interface between all computer software used on the hospital, the psychological barrier prompting us to keep a paper copy, work habits, the guarantee of computer backup quality as well as its verification. but the complete computerization of the ctp file has the following advantages: easy and secure accessibility of information, resolution problems archiving paper files, a single backup media folder. disclosure of conflict of interest: none. conditioned media from allogenic mesenchymal stem cell culture (msc-cm) enhances wound healing in an allogenic d skin model moyasasr al-shaibani, x wang , p lovat, a tulah and a dickinson newcastle university migration of the epidermal layer towards the wound centre is an important step in the healing process. full thickness in vitro skin models can be used to investigate epidermal migration towards an injury site. since wound healing therapies often require allogenic transplantation of primary keratinocytes, an allogenic d skin model was developed to investigate epidermal migration. the effect of mesenchymal stem cell conditioned media (msc-cm) was assessed for wound healing using this in vitro human d skin model. human mscs were derived from human hip joints, and characterised using standard protocols. at % confluence, msc secretions were collected in serum free medium and referred to as msc-cm which were then analysed for protein content using elisa. fully humanised allogenic d skin models were developed (n = ) and a mm punch was induced into each model followed by daily treatment with msc-cm to investigate the migration of the epidermal layer towards the punch centre over the dermal layer at different time points ( week, weeks, and weeks). intact and wounded models were characterised structurally by haematoxylin/eosin (h&e) staining and immunofluorescence (if) was used to validate the dermal and epidermal biomarkers such as collagen (col ), cytokeratin (k ), keratin (k ), loricrin and involucrin. mscs were characterised as stipulated by the international society for cell therapy, that is, fibroblast like cells with the ability to differentiate into tri-lineages (adipocyte, chondroblast and osteoblast). phenotypically, over % of the cells were able to express phenotypic markers for variant stem cells such as cd , cd and cd . over % of the cells were negative for the expression of cd , cd , cd , cd and hla-dr (p = . ). msc-cm contained different concentrations of a variety of growth factors such as keratinocyte growth factor (kgf), hepatocyte growth factor (hgf), platelet-derived growth factor (pdgf), stromal-derived factor- (sdf- ) and macrophage stimulating protein- (msp- ). h&e staining showed that the models had distinct dermal and epidermal layers similar to that of real skin. additionally, if showed that the models expressed dermal and epidermal biomarkers, for example, col , k , k , loricrin and involucrin. after treatment with msc-cm, the epidermal multilayers of the punched models started to migrate towards the punch centre and covered the whole punched area after weeks of treatment with recovered expression of the epidermal biomarkers, for example, k , k , loricrin and involucrin. a fully humanised allogenic d skin model is a useful tool to mimic the in vivo environment and evaluate the wound healing process. it could also be used as a screening method to test candidate wound healing drugs. allogenic keratinocytes could be used as a cellular sheet to cover the wound area with the ability to migrate towards the wound centre and promote wound healing. a possible explanation for promoting epidermal migration at the injury site is that msc-cm contains cytokines which accelerate cell migration such as kfg, sdf- and msp- , in addition to other cytokines which promote both migration and proliferation of epidermal cells, for example, hgf and pdgf. disclosure of conflict of interest: none. before each freezing and after each thawing, a quality control is performed including a minima: (i) cd + quantification; (ii) estimation of the percentage of hsc cd + viability, via aminoactinomycin-d ( -aad) staining and (iii) evaluation of hsc functional ability to form colony ‛cfu-gm' (colony forming unit-granulocyte macrophage). apoptosis, or programmed cell death, involves complex pathways in part the path fas-fas ligand (fasl), mitochondrial components and caspase enzymes. the involvement of apoptosis dependent on caspases activation pathway in hsc cd + after thawing remains unknown. here, we assess the extent of apoptosis caspase-dependent before and after cryoconservation of hsc cd +, using a fluorescent labeled inhibitor of caspases ‛flica. ' we tested the induction of apoptosis caspasedependent, before and after hsc cd + cryoconservation from patients with different hematological malignances: multiple myeloma (n = ), lymphoma (n = ). caspases pathway activation status was evaluated by flow cytometry, using a fluorescent labelled inhibitor of caspases ‛flica' staining test, in hsc cd +, lymphocytes cd +, monocytes cd + and natural killer cells cd +. in order to assess cell viability, cells were stained in parallel with -aad. we determined positive cells %, that is, showing caspase activation in viable cells (flica+ cells), before and after cryoconservation. caspase pathway activation level was then correlated with hsc functional ability to form colony ‛cfu-gm,' and day's number of clinical aplasia. in our cohort, we showed a significant caspases pathway activation, with . % cd + flica+ cells after thawing, compared with the . % described in fresh cd + cells (p o . ). moreover, caspases pathway was significantly activated in thawing cd +, cd + and cd + cells: flica+ cells % in thawing cells were, respectively, . %, . % and . % vs %, . % and o % in fresh cells. we also report a significant increase of apoptosis caspasedependent in lymphoma patients ( . % of cd + flica+) in comparison to myeloma patients studied ( . % of cd + flica+) (po . ). in contrast, no correlation has been established between observed caspases pathway activation and hsc cd + capacity to form cfu-gm, or still day's number of clinical aplasia. our results show substantial cell death, induced by the increase in caspases pathway activation, secondary to the thawing process, and across all study cell types. this advance of apoptosis caspase-dependent may affect the immune response quality during recipient aplasia, without detecting a clinical impact. moreover, caspases pathway activation through cd + and cd + subpopulations could modify the therapeutic result of donor lymphocytes infusion dli, though yet untested. thawing process in autologous graft induces apoptosis caspase-dependent in all apheresis product cells, particularly in hsc cd +, without clinical impact in graft fate. disclosure of conflict of interest: none. donor-derived nk cell infusion combined with hla halpoidentcial blood stem cell transplantation to decrease leukemia relapse for high risk acute myeloid leukemia patients b wu, y huang, j xu, y he, jxm zhang*, z wu* hematology department, zhujiang hospital of southern medical university, guangzhou, china *shenzheng hank biologoical engineering co.ltd. hla halpoidentcial blood stem cell transplantation have solved the donor deficiency for patient who need to treat by transplantation. the high relapse of leukemia especially for high risk patient post transplantation affect the outcome of haploidentical stem cell transplantation. natural killer (nk) cells are part of the innate immune system and play a scavenger role to detect targets marked by ‛missing self' induced by viral infection or malignant transformation. infusion nk cells into receipt prior to stem cell transplantation could decrease the gvhd in mouse bone marrow transplantation model. in an effort to decrease the leukemia relapse and gvhd after halpoidentical stem cell transplantation for high risk acute myeloid leukemia patients, we evaluated the addition of donor-derived nk killer cells before halpoidentical stem cell transplantation in high risk acute myeloid leukemia patient. here we report interim results for five patients enrolled last year. five high risk acute patients received halpoidentcial stem cell transplantation combined with donor-derived nk cells infusion. all patients received an fbca conditioning regimen, which consisted offludarabine ( mg/m /day, intravenous) on days − to − , busulfan ( . mg/kg/day, intravenous) on days − to − , cyclophosphamide( mg/kg/day, intravenous) on days − to − and rabbit antilymphocyte globulin (atg . mg/kg/day, intravenous) on days − to − . donor-derived nk cells were infused into patient prior to stem cell transplantation. gvhd prophylaxis was a combination of cyclosporine a (csa) and short term methotrexate. five high risk patients ( patients with aml m cr , patient with aml m nr, patient with aml m cr and patient with aml m cr ) enrolled from jan to nov. ; the donors are parents and sibling. hla were mismatched between donor and patients. median cd + dose infused was . /kg (range: . - /kg) and the nk cell dose infused was × /kg ( . - . × /kg). all five patients got hematology recovery and achieved hematology cr. only one patient occurred grade ii agvhd post transplantation and controlled by methylprednisolone. at a median time of months (range: - months) post peripheral blood stem cell transplantation, the incidence of acute gvhd grade ii is % ( / ) . no chronic gvhd observed. four patients are still cr and survival with event free survival with median year follow up. one patient with aml m who had not achieved remission before transplant relapsed after months and got cr with second nk infusion and still survival. nk infusion prior to transplantation was found to be safe and feasible. there was no increase acute gvhd or chronic gvhd risk. there was a trend towards increased -year survival for high risk leukemia patient. the potential benefit on overall survival remains to be further evaluated with additional patient enrollment and longer follow up. however, given the favorable safety profile of nk cells, future strategies to enhance efficacy such as repeat dosing or modification of nk cells are worth potential exploration. disclosure of conflict of interest: none. donor lymphocyte infusion (dli) is a therapeutic option in the treatment or prevention of relapse after allogeneic stem cell transplantation (allohsct).of note, the risk of graft-versus-host disease (gvhd) associated with the graft-versus-tumor (gvt) effect may be influenced by the level of hla disparity between donor and recipient. data on use of dli after unmanipulated haploidentical hsct (haplohsct) with post-transplant cyclophosphamide (pt-cy) are still currently limited. we report patients (pts) receiving dli between and for prevention or treatment of relapse after haplohsct. seven pts were given haplodli doses, as treatment for relapsed disease (n = ) or as preventive therapy of relapse for high risk disease (n = ). four pts had acute myeloid leukemia (aml), had acute lymphoblastic leukemia and lymphomas - hodgkin (hl) and non-hodgkin dlbcl. pt had intermediate risk disease features, adverse risk and pts had refractory disease at time of haplohsct. pts had a previous hsct ( allogeneic and autologous). of the pts received a ric regimen and the source of stem cells was peripheral blood s (n = ) and bone marrow (n = ). gvhd prophylaxis was cyclosporine and mycophenolate mofetil (mmf), atg and pt-cy. median follow-up after haplohsct was (range: months. median time to neutrophil and platelet ( g/l) recovery were and days, respectively. after haplohsct, pts developed acute gvhd (agvhd) of grade i (n = ) or ii (n = ), at a median of days after haplohsct. the median time from haplohsct to first dli was days (range: - ). all pts had full donor chimerism at time of dli. before dli pts relapsed at a median time of days (range: - ), of whom pts had aml and received salvage chemotherapy and pt with hl being treated by dli alone. of the relapsed pts, showed progressive disease after first dli dose and achieved a sustained cr (with duration of cr of and months at last follow-up). the remaining pts were given dli in cr, in case (of aml) associated with azacytidine. pts received dli dose and pts were given dli injections with escalating doses. the first dose of dli was × cd /kg in pts, × in pt and × in pts. the pts who received dli doses (lymphomas) were given: ( ) × - × - × ; ( ) × for doses followed by dose of × . four pts developed chronic gvhd (cgvhd, %) in a median time of days (range: - ) after dli ( of them had presented previously agvhd grade i-ii). cgvhd was limited in case, moderate in pt and severe in pts. of these pts presented features of an overlap syndrome (acute/chronic gvhd) with signs of agvhd de grade i,ii and iii in pt each. involved organs were skin/mucosal (n = ), liver (n = ), gastrointestinal tract (n = ), lung (n = ) and joints (n = ). all patients experiencing gvhd after dli were treated by systemic corticotherapy, extracorporeal photopheresis and cyclosporine or weekly low dose methotrexate. median follow-up after first dli was months (range: . none of the pts receiving prophylactic dli relapsed during the follow-up period. pts died, of relapse and of severe cgvhd. pts were in cr at last follow-up, with no signs of gvhd and with limited cgvhd. despite the limited cohort, dli after haplohsct appears to be a therapeutic option in high risk pts allowing enhancement of gvt in the setting of haplohsct with post-cy infusion. disclosure of conflict of interest: none. previously published p early and sequential ctla ig primed donor lymphocyte infusions (dli) following post-transplantation cyclophosphamide (ptcy)-based haploidentical pbsc transplantation for advanced hematological malignancies promote proliferation of mature natural killer (nk) cells with cytotoxic potential and markedly reduces relapse-risk without increase in gvhd sr jaiswal, s zaman, p bhakuni, s bansal, s deb, s bhargava and s chakrbarti we have earlier shown that cd enriched cell infusion following ptcy resulted in rapid proliferation of mature nk cells with attenuation of gvhd and early use of prophylactic g-csf mobilized dli resulted in improved disease-free survival. ctla ig has been shown to be effective in attenuating t cell activation and induce transplantation tolerance in preclinical models. it has recently been employed to induce transplantation tolerance and reduce early alloreactivity in patients with nonmalignant disorders undergoing ptcy-based haploidentical hsct. nk cells on the other hand are resistant to ctla ig and in fact might demonstrate better anti-tumour effect in presence of ctla ig as cd is a putative activation receptor. to explore this phenomenon, we employed sequential ctla ig primed dli following ptcy-based haploidentical hsct in patients with relapsed/refractory hematological malignancies. patients ( - years; aml- , all- , nhl- ) received abatacept (ctla ig) as a part of gvhd prophylaxis at mg/kg on day − followed by pbsc and sequentially on days + , + and + followed h later by dli of × cd cells/kg containing . - × /kg cd + cells. ptcy was administered on days + and + with cyclosporine from day + to day + and subsequent rapid tapering. the immune reconstitution of the study group (ctla ig-dli) was compared with the cohort of patients with both malignant and nonmalignant diseases who received abatacept but not dli (n = ; ctla ig group) and those receiving cd enriched donor cell infusion on day + (n = ; nki group). results: there were no acute infusion related toxicities. all patients engrafted at a median of days ( - days). the incidences of acute and chronic gvhd (all mild) were % and %, respectively. three patients reactivated cmv and there was only one non-relapse mortality ( . %). only patients relapsed ( . %) with a disease-free survival of . % at year. these cells had greater expression of cd a compared to normal healthy donors. the recovery of cd +, cd + + and cd + − cells were similar in the ctla ig-dli and nki groups at days , and post-transplant and this was significantly higher than the ctla ig group ( figure ). in contrast to ctla ig group, nk cells recovered at day + with predominantly cd dim cd + phenotype with significant population of cells expressing kir+nkg a phenotype in both ctla ig and nki groups with higher expression of cd a. interestingly, the patients who relapsed had attenuated recovery of cd + + cells at and days( /μl and cells/μl) without cd a expression, in contrast to the rapid and sustained recovery of this population of nk cells in those not experiencing relapse (cd + + cells /μl and /μl). however, the recovery of tregs was prompt and sustained in the comparator groups, which remained low in the ctla ig-dli group until day + . there were no differences in the recovery of other t cell subsets between the three groups. the study demonstrates the unique ability of ctla ig to augment nk cell proliferation, maturation and cytotoxicity and reduce relapse with attenuation of t cell activation and gvhd in the context of the early use of ctla ig primed dli following ptcy-based haploidentical hsct without ex vivo selection or expansion. we hope this novel strategy might offer less expensive and yet a viable alternative in the field of nk cell therapy. [p ] disclosure of conflict of interest: none. enhanced cytotoxicity of γδ-cytokine induced killer cells against hematologic malignancies n bloom, s eldror, s caspi, s teihuman,h vernitsky, e jacoby, b bielorai and a toren cik cells are ex vivo expanded by scheduled addition of anti-cd mabs and a cytokine cocktail that contains ifn-γ, il- or il- . cells represent an in vitro generated heterogeneous population consisting of different effector cells-cd poscd pos, cd negcd pos and cd poscd neg-t cells that mainly ( %) express α/β t-cell receptor (tcr) s and to a lesser extent (o %), tcr γδ phenotype. these nklike t cells product show a dual functional activity, retaining their original t cell specificity and nk cytotoxic capacity via marked up regulation of the nk cell receptor, nkg d. pre and clinical studies showed that the optimal cytotoxic effect of cik cells against different malignancies (target cells) is achieved at : e:t ratio, which means high numbers of αβ t-cells that might increase the risk of gvhd. here we produced ciks from αβ tcr depleted cellular products (defined as γδcik) and tested their phenotype expression and in vitro cytotoxic activity against hematological malignancies. fresh apheresis products were processed using the clinimacs depletion reagent, according to manufacturer instructions. target product was cultured with rpmi supplemented with % fcs and ex vivo expanded by scheduled addition of cytokine cocktail that contains ifn-γ ( iu/ml), anti-cd mabs ( ng/ml) and iu/ml il- . the cells were cultured for days. cytotoxic activity of the γδcik was evaluated against various target hematological malignant cell lines (k , reh, jurkat, and u ). after days, the αβ depleted cik cultures resulted in . % γδ t-cells ( folds expansion) compared to . % of γδ t-cells immediately after depletion, and compared to only . % in non-selected cik cells. the percentage of αβ t cells in γδcik cell cultures started from . % (immediately after depletion) to . % compared to . % αβ t cells were found in non-selected cik cells cultures. γδcik cells produced robust cytotoxic activity at a : e:t ratio against reh cells ( . ± . %), jurkat cells ( ± . %); u ( . ± . %) and k ( . ± . %), compared to nonmanipulated cik cell activity against the same targets ( ± . %; . ± . %; . ± . %; . ± . %, respectively). we found higher degranulation capacity of γδcik cells compared to non-selected cik cells against reh ( . ± . % vs . ± . %), jurkat ( . ± . % vs . ± . %), u ( . ± . % vs . ± . %) and k ( . ± . centre de thérapie cellulaire, institut paoli-calmettes; unité de transplantation et de thérapie cellulaire, institut paoli-calmettes; centre d'immunologie de marseille-luminy and laboratoire d'immunomonitoring, institut paoli-calmettes during the past years, the major improvements in the field of allogeneic hematopoietic stem cell transplantation (hsct) (reduced intensity conditioning regimen, high level hla typing, alternative donors, gvhd prophylaxis…) significantly extended the feasibility of this procedure. in contrast, disease recurrence after hsct remains a main issue. thus, many post-hsct prophylactic interventions are under investigation. unmanipulated donor lymphocyte infusion (dli) remains one of the most frequently used post-hsct treatment, but its potential benefit in increasing gvl effect may be counterbalanced by the induction of gvhd. in this setting, the use of adoptive transfer of ex vivo enriched and activated nk cell infusions from the same donor (dli-nk) may induce gvl effect without causing gvhd. we therefore report on a single-center phase clinical trial (nct ) evaluating the safety of ex vivo activated allogeneic nk cells infused between days and after hsct. the aim was to determine the maximum tolerated dose (mtd) of ex vivo highly purified and activated dli-nk after matched related donor hsct. the schedule plan a first phase of + dose escalation method using dose levels ( . e /kg, . e /kg and . e /kg). grade - secondary adverse events according to nctci classification and severe gvhd occurring within days after dli-nk were considered as dose-limiting toxicities (dlt). a second step allowed enrolling patients at the mtd. over a period of . years, patients with various hematological malignancies (aml, all, hl, nhl, mds) were infused with activated nk cells at a median time of days (range: - ) post-hsct. apheresis products were collected from the hsc donor, cd -depleted and cd -selected by immunomagnetic separation using clinimacs. selected nk cells were cultured for days in medium supplemented with % fetal calf serum in the presence of u/ml of il- in air-permeable cell culture bags. after immunomagnetic separation, cd enriched products had a median cd + cell purity of % (range: - ) and viability of % ( - ). after il- activation, the median cd + cell dose was . × e /kg ( . - . ) , with a viability of % ( - ) and a residual cd + cell content of . × /kg ( - . × /kg). all release criteria to be met were fulfilled for the preparations infused : viability %, negative microbiological testing, cd + cell count ⩾ × /kg, and cd + cell content o × /kg. standardized quality controls were employed at all steps of the manufacturing process, adding a level of consistency to the product testing before release. activated-nk cells were well tolerated in all patients, with no occurrence of dlt. thus, mtd was not reached. two patients presented with a moderate chronic gvhd, both of them during cyclosporine a dose reduction. relapse occurred in patients with aml. one patient died from idiopathic pneumoniae, without evidence of relapse, gvhd or infectious disease. with a median follow up of months ( - ), year os was % ( figure ). therefore, infusion of highly purified, activated-nk cells of donor origin as a substitute to standard dli does not induce gvhd nor other side effects after hsct: the demonstration that modulation of nk cell activity can achieve disease control after hsct deserves to be investigated in larger trials. [p ] disclosure of conflict of interest: none. feasibility, safety, rapid production and efficacy of institution-produced cd car staff were trained on site for collection, processing and cryopreservation by regional experts. a total of units were collected and processed as part of the initial validation of the project. ucb units were processed on either axp or sepax systems, and all cryopreserved in bioarchive (an automated, robotic cryopreservation system that can archive up to units). the characteristics of which as well as the post processing data are depicted in table . [p ] we shared a successful story of establishing the first public cord blood bank in jordan. the first units collected showed excellent sterility, viability, collection volume and total nucleated cells. a very good recovery of both nucleated and cd + cells were obtained using axp and sepax cell separation systems. the process of validation of equipments and methodology is complete. we anticipate moving to permeant facility of the cord blood bank in the new expansion in early . we look forward for steady progress in ucb recruitments, hla typing, cryopreservation and adherence to netcord-fact standards as well as participation in international registries. functionally active ifn-gamma secreting cmv pp specific t cell therapy as an alternative for clinically urgent cmv related diseases n kim, y-s nam , k-i im , j-y lim, y-w jeon , y song and s-g cho the catholic university of korea, seoul cytomegalovirus (cmv) related diseases are a serious cause of morbidity and mortality following hematopoietic stem cell transplantation (hsct). it has been reported over the last two decades that cmv-specific cytotoxic lymphocytes (cmv-ctls) can provide long-term cmv-specific immunity without major side effects as an alternative to antiviral drugs. however, its application has been limited by prolonged manufacturing process of cell therapy. in this study, we apply the ifn-γ cytokine capture system (ccs) using the fully automated clinimacs prodigy device to rapidly produce cmv-ctls that may be applicable in clinically urgent cmv-related diseases. five validation runs were performed using apheresis samples from randomly selected cmv-seropositive healthy blood donors. then, clinimacs prodigy automatically performed successive processes including antigen stimulation, anti-ifn-γ labelling, magnetic enrichment, and elution which took~ h. the original apheresis samples consisted of . % ifn-γ secreting cd + t cells in response to cmv pp antigen (cd +ifn-γ+ cells) which were mainly cd ra+cd l+ naive t cells. following ifn-γ enrichment, the target fraction contained . % cd +ifn-γ+ cells with reduction in naive t cells and the selection of cd ra − cd l − and cd ra +cd l − memory t cells. furthermore, extended culture of these isolated cells revealed functional activity including efficient proliferation, sustained antigen-specific ifn-γ secretion and cytotoxicity effect against pp pulsed target cells. therefore, we suggest ifn-γ ccs by clinimacs prodigy as a simple and robust approach to produce cmv-ctls, which may be highly feasible and applicable in clinically urgent cmvrelated diseases. disclosure of conflict of interest: none. in vitro generation of tumor antigen-specific t cells from patient and healthy donor stem cells s bonte , s snauwaert , g goetgeluk , b vandekerckhove and t kerre hematology, ghent university hospital, ghent, belgium and department of clinical chemistry, microbiology and immunology, ghent university, ghent, belgium acute myeloid leukemia remains a therapeutical challenge, as many patients relapse after chemotherapy. allogeneic stem cell transplantation is in most of these patients the only option for cure, but carries a high risk of morbidity and mortality and a suitable donor may be lacking. recently, advances are being made in the field of t cell immunotherapy. the classical protocol, in which peripheral blood lymphocytes (pbl) are transduced with a tumor antigen-specific t cell receptor (tcr), can generate t cells with low and possibly hazardous specificities (due to mispairing of the endogenous and introduced tcr α and β chains). therefore, we have developed a novel protocol in which we generate tumor antigen-specific t cells from cd + hematopoietic stem cells. we have already succeeded in generating large numbers of tumor-specific, naive and resting t cells that only carry the introduced tcr, starting from postnatal thymus and cord blood cd + cells. now we are optimizing this protocol for clinically more relevant samples, such as mobilized peripheral blood from healthy stem cell donors and from patients in remission after chemotherapy and/ or other treatments, and leukapheresis samples from patients at diagnosis. in our protocol, cd + cells were isolated from hla-a + fresh patient and healthy donor samples and cultured on op -dl in the presence of scf, flt l and il- , until t cell commitment. subsequently, the cells were transduced with a tumor antigen-specific tcr and again co-cultured until cd + cd + double positive cells were abundantly present. at that point, agonist peptide was added, which induces maturation. finally, cells were polyclonally expanded on feeder cells. for hla-a negative samples, cd + cd + double positive cells were co-cultured with a cell line (t pulsed with the agonist peptide or a cell line with endogenous expression of the agonist peptide) which can present the agonist peptide to the maturing t cells. using the above protocol, we were able to generate tumor antigen-specific t cells from out of healthy donor samples, / sample from a patient in remission and / samples from patients at diagnosis, who were all hla-a +. for most samples, multiple rounds of agonist peptide stimulation were necessary to obtain further maturation. in contrast, generation of mature t cells from cd + cd + double positive cells in postnatal thymus or cord blood co-cultures, requires only round of agonist peptide stimulation. for the hla-a negative samples, we were able to generate an adequate cd + cd + double positive population from / healthy donor sample, / samples from patients in remission and / sample from a patient at diagnosis. agonist selection using a cell line seems inefficient as cd is not upregulated and cells did not mature to cd + or cd + single positive mature t cells. we are currently co-culturing more samples using our protocol. furthermore, we are investigating the effect of freezing and thawing on the in vitro t cell generation process (cell numbers and efficiency). finally, we are also working on optimizing the protocol for generation of tumor antigen-specific t cells from hla-a negative patient and healthy donor samples. disclosure of conflict of interest: none. increase of polyspecific immune responses against leukemia-associated-antigens (laa) and reduction of regulatory cytotoxic t-cell (ctl) responses against malignant cells play a major role in maintaining remission and prolonging overall survival in patients with hematologic malignancies after allogeneic stem cell transplantation (allo-sct) and/or donor lymphocyte infusions (dli). graft versus leukemia (gvl) effects after allogeneic stem cell transplantation and/or dli are considered to be t cell-mediated. many groups described specific t-cell responses against several leukemia associated antigens (laa) in different hematological malignancies. however, t cell responses after allo-sct and dli are not well characterized. in this study, we analyzed laa-specific t cell responses after allo-sct and dli. to this end, we assessed the frequency and diversity of laa-specific cd + t cells using elispot analysis and tetramer assays in patients ( patients (pts) with acute myeloid leukemia, pts with chronic myeloid leukemia, pts with multiple myeloma and pts with chronic lymphatic leukemia) before and after dli. epitopes derived from prame, npm mut, rhamm, wt- and other laa were tested. moreover, the frequency of regulatory t (treg) cells was measured and the course of cytokine profiles before and after dli was analyzed. these immunological findings were correlated to the clinical course in the respective patients. in elispot and tetramer assays, an increase in frequency and diversity of laaspecific t cells was observed in all patients. importantly, there was a significant increase from a median of to laa-derived t cell epitopes (p = . ) in clinical responders (r) when compared to non-responders (nr). these positive results in r vs nr where s confirmed by tetramer-based flow cytometry assays, where an increase in frequency from . to . % in the r group of laaspecific t cell/all cd + t cells was observed. interestingly, the frequency of tregs in clinical responders decreased significantly from a median . % to . % (p = . ) while the frequency of tregs kept stable over time in non-responding patients. t cell subset analysis did not reveal significant differences before vs after dli administration. in cytokine assays using elisa for the detection of more than cytokines before and after dli we found a shift towards proinflammatory and t cell stimulating cytokines. taken immunologic surveillance of leukemia is employed for the prevention and treatment of relapse post allohsct. to augment this effect donor lymphocytes are infused (dli) in patients at risk. this procedure is associated with a high risk of agvhd and we believe that this route of administration may not make the direct contact between infused cells and blasts the optimal one. to address these issues, we started delivering donor lymphocytes directly to the bone marrow cavity (ib-dli) in patients post allohsct at relapse. three with aml and one with cll, all relapsed post allohsct: allosib: -year-old female aml patient (relapsed years post hsct), -year-old aml male q del (relapsed in years, traumatic brain injury), -year-old male aml flt itd+ received mud hsct (relapsed months) and -year-old cll male, tp del, ebv reactivation (progressed years). two patients ( % and % blasts in the marrow) received ib-dli up-front and two others due to higher proportions of leukemic cells received either flag (aml case) or anti-cd moab (cll case) followed by ib-dli. tcr clonotyping revealed in all patients the presence of the prevailing oligoclonal response on the polyclonal background (characteristic for each individual) which was identified in the marrow and in the blood. however, in two out of patients a distinct oligoclonal peak was seen at first in the marrow and then in the blood. microarray analysis of the transcriptome in the marrows of patients who received three ib-dli courses revealed in all patients preferential use of genes associated with lymphocyte or lymphocyte activation pathways. the patients who responded favorably (cr or pr) clustered with the transcriptomes of normal individuals, but those who failed to respond clustered separately. ib-dli was safe and not associated with gvhd. selective accumulation of cd +cd + as well as the presence of a distinct oligoclonal peak in the marrow suggest that tcrbeta clonotypes may be private to leukemia cells recognition. the response may result in cr or pr and the patients were in a good physical shape during the treatment, which makes it possible to deliver the salvage chemotherapy if required. broad spectrum antibiotics were started. after the orthopedic consultation, the fourth finger was amputated and amputation from the left ankle was recommended. a stem cell transplantation option was offered to patients and their relatives as one of the therapeutic approaches. upon acceptance by patient, μgr/kg of colony stimulating agent was started to patient. when the stem cell was /μl, the stem cells were collected. the obtained stem cell product was injected intra-lesionally (picture b). granulation tissue began to develop from the second week in the foot floor of the patient. after from th week, the necrotic tissue was disappeared and the granulation tissue was appeared. at weeks, % of the lesion healed. at th week, there was normal tissue instead of necrotic tissue on plantar surface at left leg (picture c). this case report suggests that diabetic foot/ulcer can be healed with intralesional application of stem cells in patients with diabetes mellitus. [p ] disclosure of conflict of interest: none. and third (n = ) cell infusions were cryopreserved. cells were infused following conventional chemotherapy (ia, mec, hdac) in cases ( %), chemotherapy plus hypomethylating agents in cases ( %) and hypomethylating agents alone in cases ( azacytidine, decytabine; %). the procedure was well tolerated, with mild and transient ‛haploimmunostorm syndrome' (fever %, rash %, diarrhea %). only the two patients with cmml received corticosteroid. one patient suffered early infusional reaction that was resolved with support treatment. none of the patients showed acute or chronic gvhd or persistent donor engraftment in chimerism tests. four patients had bacterial infections, but no other significant invasive fungal or viral infections were observed. all aml/raeb patients treated achieved complete remission with microhct treatment ( ; %). only one patient, with cmml, died during microhct induction ( %). four patients relapsed at , , and months after the infusion; two of them achieved a second sustained complete remission with another micro-hct from a different donor (one of them had developed anti-hla antibodies). as described in figure , median overall survival is months and overall survival at years is %. microhct is a well tolerated procedure in elderly aml/mds patients who are not candidates to allogeneic hct. infectious complications are insignificant and the remission rates are very encouraging in very high risk cases, with no evidence of gvhd. patients can undergo a second microhct from a different donor. in addition to the experience by ai et al, we have also shown that microhct can be safely administered following a hypomethylant agent course instead of conventional chemotherapy. a large, international, randomized clinical trial will address the safety and efficacy of microhct for elderly aml/ mds patients (nct ). [p ] disclosure of conflict of interest: none. wilms tumor protein (wt ) is expressed in a variety of solid tumors and is found in more than % of patients with acute myeloid leukemia which makes it an attractive target for immunotherapy. previously it was shown that t cells recognizing wt are suitable for adoptive t-cell therapy by increasing the graft versus leukemia effect. however, the efficiency of this therapeutic strategy is still limited due to the low precursor frequency and specificity of wt -specific t cells in the peripheral blood of healthy donors. the ubiquitous antioxidant inducible enzyme heme oxygenase- (ho- ) and its products have immunomodulatory effects, which render it as a potential target for the modification of t-cell responses. recently, we found that inhibition of ho- enzyme activity via tin-mesoporphyrin (snmp) results in activation and proliferation of antiviral t cells from healthy donors. in this study we aimed ( ) to identify the mechanism of ho- modification in the generation of wt -specific t cells and ( ) to develop strategies for the sufficient generation of wt specific t cells from healthy donors to augment effective t-cell immunity in leukemia patients and to broaden the applicability of adoptive t-cell therapy to the majority of patients. the frequency of wt -specific t cells in peripheral blood of healthy donors (n = ) was examined before and after snmp treatment via ifn-γ elispot using the wt -overlapping peptide pool (ppwt ). enrichment efficiency of wt -specific t cells after ho- inhibition was verified in response to ppwt and the hla-a* : -restricted wt peptides (vldfappga, wt ) and (rmfpnapyl, wt ) by ifn-g secretion assay and expression analysis of the t-cell activation marker cd . phenotypic and functional characterization of wt specific t cells were further assessed by multicolor flow cytometry, luminex assays and elisa with respect to t-cell subsets, cytotoxicity, proliferative capacity and secretion of effector molecules. in % of donors we found specific t cells against ppwt by ifn-γ elispot ( spots/ . pbmcs). the frequency of wt -specific t cells in these donors could be increased fivefold after inhibition of the enzymatic activity of ho- via snmp. to assess the possibility that ho- modulation might be clinically applicable in conformity with good manufacturing practice, enrichment of snmp-treated wt -s specific t cells was evaluated based on ifn-g secretion and cd expression. compared to snmp-untreated cells there was a . -fold higher response of ho- modified wt -specific t cells pre-enrichment and an up to -fold higher enrichment efficacy, while snmp treatment did not affected the t-cell functionality. in conclusion, modification of the enzymatic activity of ho- resulted in a more effective generation of functionally active wt -specific t cells suitable for adoptive t-cell therapy. this makes ho- a promising therapeutic target to boost antigen-specific t-cell responses for treatment we recently developed and characterized a standardized and clinical grade human platelet lysate (hpl) that constitutes an advantageous substitute for fetal bovine serum (fbs) for human mesenchymal stem cell (hmsc) expansion required in cell therapy procedures, avoiding xenogenic risks (virological and immunological) and ethical issue. because of the progressive use of pathogen reduced (pr) labile blood components, we evaluated the impact of the novel procedure theraflex uv-platelets for pathogen reduction on hpl quality (growth factors content) and efficacy (as a medium supplement for hmsc expansion). this technology is based on short-wave ultraviolet light (uv-c) and has the main advantage not to need the addition of any photosensitizing additive compounds (that might secondary interfere with hmscs). we applied theraflex uv-platelets procedure on fresh platelet concentrates (pcs) suspended in platelet additive solution and prepared hpl from these treated pcs. we compared the quality of pr-hpl with the corresponding non-pr ones, in terms of growth factor contents. then, we evaluated the efficacy of pr-hpl, in comparison with hpl and msc-screened fbs. we performed large scale culture of hmscs during passages and evaluated the proliferation of cells and the maintenance of their properties: profile of membrane marker expression, clonogenic potential, immunosuppressive properties (inhibition of t-cell proliferation) and potential to differentiate in adipocytes and osteoblasts. we showed no impact on the content in growth factors tested (egf, bfgf, pdgf-ab, vegf and igf) and a significant decrease in tgf-b (− %, n = , p o . ). a large scale culture of hmscs during passages showed that hpl or pr-hpl at % triggered comparable hmsc proliferation than fbs at % plus bfgf (n = ). moreover, after proliferation of hmscs in hpl or pr-hpl containing medium, their profile of membrane marker expression, their clonogenic potential and immunosuppressive properties (inhibition of t-cell proliferation) were maintained, in comparison with hmscs cultured in fbs conditions. the potential to differentiate in adipogenic lineage of hmscs cultured in parallel in the conditions, evaluated using oil red o and nile red stainings and the measurement of triglyceride accumulation, remained quantitatively identical. we also showed that the potential to differentiate in osteoblasts (quantified using alizarin red s and von kossa stainings and alp activity measurement) of hmscs grown in hpl or pr-hpl was not impaired, in comparison with fbs. in conclusion, we demonstrated the feasibility to use uv-c treatment to subsequently obtain pathogen reduced hpl, while preserving its optimal quality and efficacy for hmsc expansion for cell therapy applications. although it is still not used widely in clinical practice. in this paper, we demonstrated a case of ada-scid who received hsct as an adolescent from matched unrelated donor (mudd) after termination of her peg-ada treatment due to severe intractable thrombocytopenia induced by peg-ada. patient showed good engraftment and incremental clinical improvement. her post transplantation course was complicated with multiple complications including: grade i gut gvhd as well as hemorrhagic cystitis (btk related) and ebv infection, additionally, she developed several cns complaints like headache, vomiting and dizziness which were found to be due to increased intracranial pressure with multiple enhancing cerebral lesions found on brain imaging. further investigations for the brain lesions confirmed the diagnosis of malignant diffuse large b cell lymphoma (dlbcl) involving the brain. the lymphoma was highly suggested to be originated from donor cells giving the timing relationship between transplant and establishment of the diagnosis. this lymphoma was successfully treated with full recovery and good final immune reconstitution but with lack of b cell engraftment and need for monthly ivig. we conclude that, peg-ada can rarely induce thrombocytopenia in an autoimmune manner by forming antibodies against platelets and good recovery of thrombocytopenia can be achieved after discontinuation of peg-ada. hsct can be considered as modality of treatment even in older patients with scid due to ada deficiency keeping in mind high possibility of complications including, autoimmunity and malignancy. disclosure of conflict of interest: none. ( ) ( ) ( ) ( ) . based on the pre and post-apheresis cd + cell counts, the collection efficiency of the apheresis amicus device was median . % ( - ) and of the comtec median % ( - ). in mm the apheretic collections were started on median day ( - ), while in lymphoma patients, due to chemotherapy, the day of apheresis start was ( - ). after cryopreservation and thawing, viability ( -aad, bd) was median . % ( - ). with these cell products, up to now we engrafted patients following high-dose chemotherapy ( mm autografted after mel , hl and nhl autografted after beam). engraftment was prompt and stable in all with anc . and . × /l on median day ( - ) and . ( - ), respectively, and with platelet count and × /l on median day ( - ) and . ( - ), respectively. these results are similar to those obtained by most experienced centers in europe and us, and confirm the fact that autologous transplantation may be implemented also in developing countries when appropriate technology and application of standard procedures are employed. with this experience our center is also developing allogeneic transplantation, and the initial results in thalassemia will be reported in a separate abstract. disclosure of conflict of interest: none. extracorporeal photopheresis (ecp) is a safe and effective immunoregulatory therapy for steroid-refractory graft-versushost disease (gvhd) but its mechanism of action is poorly understood. ecp is a non-immunosupressive therapy whose modulating mechanism is thought to result in an increase in t-regs in the patient and in inversion of the cd /cd ratio at the end of treatment. in this study, we evaluated the effect of ecp on t cell response in a cohort of steroid-refractory gvhd patients. from november to november , patients ( con acute gvhd and with chronic gvhd) treated with ecp in our unit, were retrospectively evaluated. patient characteristics are shown in table . we performed an ‛off-line' system ecp using a cell separator (spectra optia, teruno bct) for the cmn apheresis; after -methoxypsoralen was added, the product was photoinactivated in the ultraviolet a irradiator (uvamatic-g , macopharma). ecp procedures were performed for two consecutive days, initially weekly (agvhd), or every two weeks (cgvhd) and afterwards monthly according to clinical response. anthracycline-induced cardiotoxicity (aic) is irreversible, which has limited the use of this anthracycline in cancer chemotherapy. to explore the therapeutic effect and its possible mechanism of bone marrow derived mesenchymal stem cells (bmscs) on cardiac damage induced by anthracyclines in a rat model. study selects sd rats aged - weeks to isolate and culture bmscs, and flow cytometry was used for phenotypic identification of bmscs. female sd rats were first randomly divided into groups: the sham control, bmscs control, . mg/kg daunorubicin (dnr), dnr with bmscs, dnr with dexrazoxane (dzr), dnr with bmscs and dzr. left ventricular (lv) function before, during and after chemotherapy were assessed by echocardiography. at the end of weeks, animals were euthanized and organs were collected in % buffered formalin for histopathology using hematoxylin and eosin staining and immunohistochemical analysis was used to identify the cellular subpopulations that infiltrate the cardiac tissues. after the construction of microrna- (mir- )modified bmscs with lentiviral vector, sd rats were randomly assigned into groups: the normal control, the empty vector control, dnr, dnr with bmscs, dnr with mir- -modified bmscs. the density of new blood vessels of rats in each group was detected by immunohistochemical method. mir- , bcl- , bax and vegf mrna expressions were detected by qrt-pcr. bcl- , bax and vegf, cx , troponin t and bnp protein expressions were detected by western blotting. all procedures performed in studies involving animals were in accordance with the ethical standards of the institutional. an animal model of drug-induced cardiomyopathy was built in the dnr treated rats.lv ejection fraction (lvef) and lv fractional shortening (lvfs) were significantly decreased compared to that of the sham control (p o . ), and the signs of the myocyte injury (myocytolysis, vacuolization and disruption) in paralleled with the inflammatory infiltrates, marked by cd and hla-dr, were observed in the dnr group, while bmscs alone or synergistic with dzr facilitate the anthracycline-induced lv dysfunction returning to the baseline values and the recovery of myocarditis (p o . ). in the mir- -modified bmscs transplant group, mir- expression, cell migration and proliferation ability were higher than that in the bmscs and empty vector groups (p o . ). the cardiac regenerative capacity of bmscs following significant myocardial injury were further enhanced by mir- compared to that of the dnr group and the control groups (all po . ), revealed by the significantly higher density of new blood vessels and upregulation of vegf expressions, during which the pro-apoptotic protein bax were down-regulated and the anti-apoptotic protein bcl- function were upregulated in the mir- overexpression group compared to that with the bmscs, dnr group and the control groups (all po . ). western blotting demonstrated that the expression of c × were significantly decreased, while expressions of troponin t and bnp were significantly increased in the mir- overexpression group in contrast to that with the dnr group (all p o . ). these results showed that bmscs could reverse cardiac damage induced by anthracycline, and the cardioprotective efficacy was further enhanced by mirna- -mediated regulation of apoptosis and angiogenesis. disclosure of conflict of interest: none. effective adoptive t cell therapy against cancer is dependent on long-lived tumor-specific stem cell-like t cells with the ability to self-renew and differentiate into potent effector cells. however, current protocols for ex vivo generation of tumorspecific cd + t cells result in terminally differentiated effector t cells. it was found that minor histocompatability antigen (miha)-specific cd + t cells with an early memory-like phenotype and long-lived memory transcription profile could be expanded from naive precursors using akt-inhibitor viii . importantly, these akt-inhibited tumor-specific cd + t cells showed a superior expansion capacity and anti-tumor effect multiple myeloma bearing mice. for the clinical exploitation of ex vivo generated akt-inhibited tumor-specific cd + t cells, we tested the effect of potential clinical grade akt-inhibitors azd , gdc , gsk , gsk , mk and triciribine in polyclonal stimulations, allogeneic mixed lymphocyte reactions (mlr), and antigen-specific t cell assays. polyclonal stimulation with anti-cd /cd beads on cd naive t cells was used for a first screening of the akt-inhibitors. for all inhibitors, a dose dependent effect on the naiveassociated receptors ccr , cd l and cxcr was observed. this had limited effect on viability, activation and proliferation except for triciribine, which was therefore excluded for further assays. moreover, in the mlr, treatment of naive cd + t cells with remaining akt-inhibitors resulted in a dose dependent effect associated with higher ccr , cd l, cxcr and cd expression. furthermore, the akt-inhibited cd + t cell products showed a - fold increased expansion capacity upon restimulation in vitro. when expanding miha-specific cd + t cells from the naive repertoire in the presence of one of the akt-inhibitors, the miha-specific cd + t cells showed a more early memory phenotype compared to controls. this was displayed in higher levels of the naive-associated receptor cd l ( figure ). in addition, these miha-specific cd + t cells were shown to be functional, as antigen-specific restimultation resulted in degranulation (cd a) and ifn-γ production. based on this ifn-γ production, the akt-inhibited antigenspecific cd + t cells can be selected using the cytokine capture assay (miltenyi, for enriched infusion in patients suffering from hematological malignancies. using aktinhibition in the generation of tumor-reactive t cells results in a more early memory tumor-specific cd + t cell product. this adoptive immunotherapy product retains superior proliferation capacity upon infusion, and its potential selfrenewal capacity could result in a long-term anti-tumor effect in patients suffering from a hematological malignancy. chimeric antigen receptors (cars) are composed of an extracellular domain-derived from a tumour-reactive monoclonal antibody, linked to one or more signalling endodomains. in early clinical trials, cd car-t cells have demonstrated impressive anti-tumour activity against different b-cell malignancies, including chronic lymphocytic leukaemia, acute lymphoblastic leukaemia (all) and non-hodgkin lymphoma. conventional alpha-beta car-t cells are however hla-restricted and could cause graft-versus-host disease (gvhd) when used across major mismatches, as expected in the highly anticipated setting of off-the-shelf car-t cells from third-party donors. besides being non-hla restricted, gammadelta t cells possess intrinsic anti-tumour reactivity, making them attractive effectors for next-generation car-t cell therapies. so far, however, attempts at exploiting gammadelta t cells in patients have been largely disappointing, possibly because of sub-optimal ex vivo culture conditions. the aim of our study was to optimise the generation of gammadelta car-t cells and to test their anti-tumour potency both in vitro and in vivo. starting from peripheral blood mononuclear cells of healthy donors, we stimulated gamma-delta t cells with zoledronate and il- /il- , and transduced them with retroviral vectors encoding for cd cars carrying either cd .z or - bb.z signalling endodomains. we assessed antitumour activity in vitro by measuring killing, secondary expansion and cytokine production after co-culturing gamma-delta car-t cells with different cd + all cell lines, and in vivo in nsg previously engrafted with a b-all semi-cell line. although allogeneic hematopoietic stem cell transplantation (allosct) is a curative option to treat hematologic malignancies, disease recurrence remains a concern in the setting of high risk diseases. thus, post allosct therapeutic strategies are needed to treat and/or prevent disease progression. in this setting, donor lymphocytes infusion (dli) is an option as post allosct immunotherapy aiming to enhance graft versus leukemia (gvl) effect. although dli may induce persistent remission, graft versus host disease (gvhd) is a potential complication following dli. because of the suspected higher incidence of gvhd in the presence of hla mismatches, few series focused on dli following haploidentical stem cell transplantation (haplosct) so far. we therefore report our experience of dli following haplosct using post-transplantation cyclophosphamide (pt-cy) platform. we included in this single center study all consecutive adult patients with hematological malignancies who received dli after haplosct with pt-cy as part of gvhd prophylaxis from to (n = ). conditioning regimens were non-myeloablative (low dose tbi-based) or with reduced toxicity (various dose of busulfan according to disease and patient characteristics). ciclosporine a and mycophenolate mofetil were given as additional gvhd prophylaxis in all cases. dli were given at escalating doses, expressed as cd +cells/kg, without gvhd prophylaxis, and ranged from × our study suggests that dli following haplosct with pt-cy is feasible. gvhd is frequent but with a relatively low incidence of severe forms. no response rate was achieved in the context of hematological relapse, underlining that preemptive or prophylactic strategy might be preferred. indeed, the overall good outcome in patients receiving prophylactic dli is promising taking into account the poor prognostic of the diseases indicated for alternative donor transplantation. further prospective studies are needed in specific disease settings to assess the benefit for using such post allohsct immune-intervention. [p ] disclosure of conflict of interest: none. dual specific cytokine-induced killer cell therapy as a treatment option for life-threatening ptld-a case report of the frankfurt experience l-m pfeffermann infection with epstein-barr virus (ebv) is a frequent complication after allogeneic hematopoietic stem cell transplantation (hsct) and besides relapse remains a significant cause of morbidity. prolonged immunosuppression or delayed t-cell recovery may favor ebv reactivation after transplantation, which under these circumstances can lead to life-threatening lymphoproliferative disease (ptld). consensus is lacking on the optimal treatment of ptld. adoptive immunotherapies with both anti-tumor capacity and restored virus-specific cellular immunity may represent optimal treatment options especially when considered in the context of ptld. in this case report we applied in vitro activated t-cells namely cytokineinduced killer (cik) cells with dual specific cytotoxic capacities transferring both anti-cancer potential and donor t-cell memory against ebv infection for the treatment of ebvassociated ptld which progressed to highly proliferative large b cell lymphoma during delayed t-cell recovery after allogeneic hsct. the reported patient had received an allogeneic hsct for secondary myelodysplastic syndrome following acute myeloid leukemia, and due to delayed t-cell recovery had developed ebv-related ptld two months after transplantation. treatment with rituximab, conventional ebvspecific t-cells and wildtype cik cells failed, therefore the patient was offered ebv-specific cik cells on a compassionate use basis. ebv-specific cik cells were generated from peripheral blood mononuclear donor cells. cells were activated and expanded in the presence of ifn-γ, il- , anti-cd antibody and il- . on day and of culture an ebv peptide pool was added for additional priming. follow-up analysis included in vitro and in vivo monitoring of ebv-specific cik cells. with above mentioned protocol we were able to generate cik cells containing × cd + ebv-specific t-cells/kg body weight of the patient. infusion of ebv-specific cik cells resulted in rapid clearance of plasma ebv dna level and sustained disappearance of large (vol. cm ) ptld-malignant lymphoma. during one-year follow-up analysis we were able to detect ebv-specific cik cells (cd + and cd + ) in vivo by flow cytometry using specific mhcdextramers. facs-monitoring of the patient´s blood revealed besides cd bright t-cells also an increasing cd dim t-cell population with a remarkable percentage of t emra cells within this compartment (up to %) indicating virus-specific t-cells. no cytokine release syndrome appeared after ebv-specific cik cell treatment, but cytokine secretion patterns, analyzing serum of the patient, reflected cytotoxic and anti-virus capacity provided by this treatment. cytotoxic potential, as well as t h cell differentiation and function offered by ebvspecific cik cell treatment were further confirmed by in vitro analysis. ebv-specific cik cells revealed an . s hematologic disease. the global survival ratio in the follow-up was . % (with . %, . % and . % survivals in , and months, respectively). the variables significantly associated with greater survival were: type of gvhd (cgvhd), number of affected organs (an organ had to be moderately or severely affected to be included in this category) and steroid dependence as the main reason to initiate ecp (see figure ). there was a trend towards significance for the degree of gvhd and cutaneous involvement to be factors associated to enhanced survival ratios. extracorporeal photopheresis is a safe treatment option for patients with gvhd, generating a response and decreasing immunosuppression in an important percentage of them. the presence of cgvhd rather than agvhd, a lower severity degree of the condition, having a lower number of affected organs, skin as main affected organ and steroid dependence as the reason to start the ecp treatment were all factors associated with greater survival in our sample. disclosure of conflict of interest: none. tx) . we report the case of a patient refractory to chemotherapy treated with ibrutinib as debulking therapy before allo-tx. in june , a years old woman was diagnosed with mcl. the staging performed by whole body ct scan, colonoscopy, egds and bone marrow (bm) biopsy was conclusive for stage iva with bulky lymph node over and below the diaphragm, bm, enteric and peripheral blood (pb) localization. the planned treatment included cycles of r-chop, cycle of high dose (hd) cy, cycles of hd arac and autologous sct. after completion of hd cy, the restaging showed progressive disease, with a thyroid involvement and histologic switch in a blastoid variant. disease continued to progress even after cycles hd arac, so we tried to control the disease with r-bendamustine ( mg/mq on days - of -d cycle), but after the first cycle the neck circumference increased. we shift to lenalidomide ( mg on days - of -d cycle) without any response after two cycles. we excluded patient from autologous sct programme because of chemo-refractoriness and we searched matched unrelated donor because no hla identical sibling was available. we started a therapy with ibrutinib ( mg/die on days - of d cycle). after the first cycle we observed a rapid response with decrease of neck size and the disappearance of superficial lymphnode; we performed cycles of ibrutinib, and we reached a good partial remission with lymphnode of max cm, and a bm and pb involvement of %. meantime an unrelated donor with / hla matching was identified, so in december we performed allo-tx with reduced-intensity conditioning (thiotepa mg/kg-fludarabine mg/m -melphalan mg/m ) and cyclosporine and short term methotrexate as gvhd prophylaxis. engraftment was at day + . in the first days after allo tx she experienced a clostridium enteritis, transient cmv reactivation and acute gastrointestinal gvhd on day + with rapid response to steroid therapy. main complication happened on day , when sudden fever and stupor, progressive to coma, occurred; subsequently pneumococcal encephalitis was diagnosed, with positive csf microbiological exam and two signal alteration in the right cerebral hemisphere at mri. the patient was treated with ampicillin and ceftriaxone with a favourable outcome. the mcl revaluations performed at , and months showed complete remission with disappearance of all pathological lymph node and pb involvement. currently the patient is at year post asct, she is enrolled in a rehabilitation program and mcl is in complete remission. our experience seems to indicate that ibrutinib is safe and can be used as bridge to allo-tx therapy in refractory mcl. we will investigate side effects of this platform of therapy, and, given the early occurrence of pneumococcal infection, we will consider to perform capsulated bacteria vaccination before allo-tx. disclosure of conflict of interest: none. [p ] pt was diagnosed with c-all in and received a mud-hsct (tbi gy, cyclosphosphamide) in / due to persistend mrd. following early rel / , cycles of blina led to mrd+ cr, for which a nd hsct from a haploidentical family donor (busulfan, thiotepa, fludarabine) was performed in / . molcr lasted months and rel was treated with cycles of weekly io followed by one dli ( x - cd + cells/kg), resulting in mrd+ cr and complete donor chimerism. five weeks after the last io cycle, the pt was admitted with ascites, hyperbilirubinemia and reduced general condition. vod was suspected, but diagnostic paracentesis revealed malignant ascites demonstrating fatal progressive disease. conclusion. our data suggest that the sequential use of io and dli is feasible even for heavily pretreated patients with r/r all after hsct and can induce molecular remissions. we observed an unusual case of late onset, severe vod responding to defibrotide and one all relapse manifesting itself with ascites in our patients. we therefore suggest close monitoring of liver function tests in the setting of this therapy and extensive diagnostic work-up for any developing liver abnormalities or ascites. disclosure of conflict of interest: ng: advisory board (pfizer, amgen); research support (amgen); gb: honoraria (amgen). [p ] p while allogeneic hematopoietic stem cell transplantation from matched related and unrelated donors has become a standard of care treatment for patients with hematological malignancies, transplantations from mismatched or haploidentical family donors remain challenging. currently t-replete and t-deplete transplantation strategies are applied aiming to improve the outcome after haploidentical transplantation. despite high rates of relapse many centers regard post-transplant cyclophosphamide, a t-replete strategy, as a standard of care approach. we have developed a t-depleted transplant approach where donor lymphocytes selectively depleted of alloreactive t-cells (atir ) using th , arhodamide-like dye, are infused after cd -selected haploidentical hsct, to overcome the challenges of infectious complications, gvhd and relapse. in phase i (cr-air- ) we have demonstrated safe infusion of these lymphocytes at doses up to × viable t-cells/kg. recently, we reported a promising -year grfs was % from a phase ii trial (ash ),that is awaiting final results soon. here, we introduce a randomized, multicenter phase study (cr-air- ), where patients with acute myeloid leukemia (aml), acute lymphoblastic leukemia (all) or myelodysplastic syndrome (mds) are planned to undergo a haploidentical hsct with either a t-cell depleted graft and adjunctive treatment with atir , or with a t-cell repleted graft and use of posttransplant cyclophosphamide. inclusion and exclusion criteria are listed in table . all patients will undergo myeloablative conditioning consisting of either tbi ( gy) or melphalan/ busulfan, in combination with thiotepa and fludarabine. patients in the atir study group will receive atg ( . mg/ kg once daily for days) during conditioning and atir infusion at a dose of × viable t-cells/kg is given between and days after the hsct. patients in the ptcy group will receive cyclophosphamide ( /mg/kg) on day and (or ) with subsequent use of immune suppression up to months post-hsct. the primary endpoint of the study is gvhd-free, relapse-free survival (grfs). grfs is defined as time from randomization until grade iii/iv acute graft-versus-host disease (gvhd), chronic gvhd requiring systemic immunosuppressive treatment, disease relapse, or death, whichever occurs first. this endpoint captures both safety and efficacy. additional secondary endpoints are overall survival (os), progression-free survival (pfs), relapse-related mortality (rrm) and transplantrelated mortality (trm). patients are planned to be randomized in study centers in europe and north america. a number of - sites are planned to participate in the study. enrolment is expected to continue until mid- with initial results being available first half . results of this study will determine which transplant regimen provides most clinical benefit in haploidentical donor transplantation, with the promise of an effective regimen without the use of post-transplant immune suppression. disclosure of conflict of interest: jr is an employee of kiadis pharma. multipotent mesenchymal stromal cells (mscs) are used for prevention and treatment of graft versus host disease after allogeneic hematopoietic stem cells transplantation due to their immunomodulatory properties. mscs fate in vivo after infusion is unknown. the aim of this study was to analyze the changes in mscs and allogeneic lymphocytes properties when co-cultured in vitro to simulate their interactions in vivo. the bone marrow from donors ( male and female aged - years, median years) was used. mscs were cocultured with allogeneic lymphocytes in a ratio of about : for days and their basic properties were analyzed over time. lymphocytes were activated by adding to the culture medium mg/ml of pha (pha-lymphocytes). some mscs were treated for h with u/ml ifnγ (γmscs). determination of gene expression levels was performed by reverse transcription polymerase chain reaction in real time (modification of the taq-man) and of antigen expression on mscs and lymphocytes by flow cytometry. significant reduction in the proportion of viable cells was observed in mscs co-cultured with pha-lymphocytes. in γmsc co-cultured with pha-lymphocytes no reduction in the proportion of living cells was revealed. this indicates the sensitization of mscs by ifnγ to factors secreted by pha-lymphocytes. in mscs co-cultured with pha-lymphocytes and lymphocytes mean fluorescent signal intensity level (mfi) of cd gradually decreased. ifnγ treatment and co-cultivation with lymphocytes led to significant increase of hla-dr mfi on mscs. co-cultivation with lymphocytes increase the hla-dr mfi on mscs much stronger than ifnγ treatment. relative expression level (rel) of ido gene increased dramatically in both mscs and γmscs when co-cultured with lymphocytes. at a day in γmscs rel of ido increased fold, and then gradually declined. in mscs cocultured with lymphocytes il- rel increased almost -fold and then decreased -fold at the fourth day. the csf rel in γmscs showed twofold increases, upon incubation mscs and γmscs with lymphocytes csf rel increased fourfold and sevenfold, respectively. co-culture of msc and γmscs with lymphocytes led to decrease in the proportion of cd , cd , cd , hla-dr, and pd- positive cells (both cd and cd ) after one day, compared with pha-lymphocytes without mscs. proportion of cd +, hla-dr+ and pd- + cells also decreased after days of co-culturing with msc or γmscs (compared with pha-lymphocytes without mscs), but anyway number of activated cells increased . - folds compared with first day. number of cd + lymphocytes after days of co-culturing with mscs or γmscs did not vary significantly from control and decreased in comparison with first day. main inhibition of activated lymphocytes by co-culturing with mscs occurs during the first day of their interaction, and then the inhibition became less effective, moreover in mscs decreased the rel of the main immunomodulating factors, and most of them were eliminated. mscs treatment with ifnγ resulted in improved survival and resistance of these cells to lymphocytes action. the results indicate that the effect of mscs injected intravenously to patients is limited to several days. disclosure of conflict of interest: none. autologous adipose-derived mesenchymal stem cells (admscs) embedded in platelet-rich fibrin (prf) promote healing in different types of wounds. by avoiding the needlerelated complications, prf-embedded autologous admscs graft provides a new effective stem cell-based therapeutic strategy for wound healing. adult male (age ⩽ yo) were equally divided (n = per group) into group (prf only), and group (prf+admscs). regular dressing (without any agent) was used for both groups with a frequency of changing every days. rpf with or without admscs was patched on the wound (maximum surface area cm ). all patients were followed up until complete healing. a complete healing was noted in both groups; however, the healing in group was very slow (after weeks), compared to a quicker one in group (after weeks). control of the moisture was very well noted in group , less in group . group showed a lot of exudates on the wound; less exudate in group were noted. infections were absent. both groups had a colonized wound. signs of inflammation were very well noted in group ; no signs of inflammation in group . admscs embedded in prf offered rapid wound healing responses then prf alone. keywords: mesenchymal stem cells, platelet-rich fibrin, engineered tissue wound healing disclosure of conflict of interest: none. stem cell source p additionally cryopreserved g-csf primed pbsc can substitute the second transplantation for the patients with acute leukemia who lately relapsed after hematopoietic stem cell transplantation y lee , j moon , ih lee and s sohn department of hematology, kyungpook national university hospital; department of hematology,kyungpook national university hospital and kyungpook national university hospital although allogeneic hematopoietic cell transplantation (allo-hct) is a potentially curative therapy for acute leukemia, survival outcomes of the patients relapsed after transplantation remains poor with high early mortality and a small percentage of second remission. this study evaluated the efficacy of dli using g-csf primed pbsc additionally cryopreserved for the patients who relapsed after allo-hct. we retrospectively reviewed the medical records of the patients who received allo-hct for acute myelogenous leukemia (aml), myelodysplastic syndrome (mds), and acute lymphoblastic leukemia (all) between and in kyungpook national university hospital. among the patients who had relapsed after allo-hct, the patients received dli using the additionally harvested cells. at the time of harvest for the first hct, collecting targeted pbscs (greater than × /kg cd + cells) allowed us to cryopreserve surplus pbscs, including cd + cells with dimethylsulfoxide in a nitrogen tank. then, we analyzed the efficacy of dli for the patients who were classified into early relapse or late relapse group by the median time of relapse after transplant. the median age at transplant was . years (range - years) and male was patients ( . %). primary diseases for allo-hct were aml/mds (n = , %) and all (n = , %). one hundred forty three patients ( . %) were in cr (complete remission), ( %) in further cr, and ( . %) in relapsed and refractory status. one hundred seventy one patients ( . %) received myeloablative conditioning regimen. the median dose of cd + cell was . × /kg (range: ~ . × /kg). almost % of patients achieved the neutrophil engraftment with a median time of days (range: - days). the -year overall survival (os), relapse free survival (rfs), non-relapse mortality (nrm) and graft-versus-host disease (gvhd)-free/relapse-free survival (grfs) since hct was . ± . %, . ± . %, . ± . %, and . ± . %, respectively. there was no significant difference according to s the infused cd + cell dose (lower o × /kg vs higher ⩾ × /kg). the incidence of chronic gvhd was more frequent in higher cd + group ( . % vs . %, p = . ). median time from hct to relapse was days (range: ~ days). after relapse, patients ( . %) were treated with salvage chemotherapy, patients ( . %) with second allo-hct, and patients ( . %) with dli. the median number of cd +t cell was . × /kg (range: . ~ . × /kg). fourteen patients ( . %) achieved dli induced cr, patients progressed, and patients were not evaluable for response. dli induced acute gvhd was observed in patients ( . %) and chronic gvhd developed in patients ( . %). in late relapse group, the -year os since post-transplant relapse was significantly higher in dli group than non-dli group (figure , . ± . % and . ± . %, p = . ) but, early relapse group had no difference. the patients treated with dli showed significantly survival benefit in late relapse group (median days vs days, p = . ). the incidence of dli-induced gvhd does not differ between two groups. dli for the patients who lately relapsed after allo-hct can be a feasible and an effective option in terms of response, donor convenience and it's cost. in the late relapse group, g-csf primed dli may replace second transplantation. disclosure of conflict of interest: none. cord blood transplantation- years of experience c alves , f amado , f bordalo , s ferreira , s lopes , c pinho , t rodrigues , l antunes and s roncon serviço de imuno-hemoterapia; serviço de terapia celular and registo oncológico do norte, instituto português de oncologia do porto francisco gentil, epe allogeneic haematopoietic stem cell transplantation (hsct) is a well-established treatment for patients with malignant and non-malignant haematological disorders. cord blood transplantation (cbt) has extended the availability of hsct to patients that would not otherwise be eligible for this curative approach because of the lack of human leucocyte antigen (hla) identical donor. the aim of this retrospective study was to analyse and characterize years of cbt activity in our institution ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) . we examined patient electronic files and created a database in excel to register cord blood unit (cbu) parameters and patient characteristics. after thawing, cbu was washed/diluted with validated procedures; the cellular content was evaluated by immunophenotyping and followed ishage recommendations; sterility was assessed by bacterial/fungal cultures, viability by trypan blue exclusion assay and functionality by clonogenic capacity. the transfusion of blood products after transplant was quantified and the hematological recovery (hr) established using cibmtr criteria. correlation between continuous variables was assessed with spearman coefficient. overall survival (os) was determined according to cellular content and hla-disparity by the kaplan-meier estimator. survival between groups was compared using the log-rank test. a total of cbu were administered to patients (table ) : / female, the main diagnosis was acute leukaemia ( / ). a sibling cbu was used for patients; the unrelated were imported from europe ( %), usa ( %) and oceania ( %). hla-matching was / , / , / and / for , , and patients, respectively; % were abo-identical. after thawing, % were washed and presented no microbial growth. the majority of patients were submitted to a bussulfan-based myeloablative conditioning regimen; graft versus host disease prophylaxis was performed with a calmodulin inhibitor+mycophenolate mofetil. complete and mixed chimerism was verified in % and % of patients; % had graft failure; the rest were unknown results ( %). at the moment, we reported patients alive ( in complete remission, with evidence of disease relapse) and dead at a median of ( . - ) months after cbt; the most frequent cause ( %) was recurrence of the initial clinical condition. the correlation between nucleated cells (nc) and cd + cells per kg/hr (p = . ; p = . ) and number of cd + cells per kg/os (p = . ) was not statistically significant. however, the engraftment and os was associated with hla-mismatch (p = . ; p = . ) and os was related to nc per kg (p = . ). our clinical results suggest that despite increased hla disparity, ucb offers promising results. ucbt is feasible in patients when the unit contains a high number of cells. there are several strategies for the future, related to cbu expansion and homing techniques, nurturing procedures, selection of optimal cbu unit and enhancement of immune recovery, in order to improve the application of cbu. s received sirolimus and mmf. median time to neutrophil and platelet engraftment was days ( - ) and days ( - ), respectively. one patient died from parainfluenza pneumonia (d ), one patient from ptld (d ), one patient from late pulmonary vod (d ), and one patients from relapse (d ). with a median follow up for survivors of months, one year survival is %. three patients had grade - gvhd and none of the survivors have chronic gvhd. though unrelated donor chimerism was dominant early after transplant and contributed to early count recovery, definitive engraftment was dominated by ucb chimerism in all but one patient. conclusion: among older adult patients with hematological malignancies,~ % lack haplo-identical relatives. for these patients, double or single ucb transplant is challenging because of delayed engraftment. cd selected partially matched grafts from unrelated donors hasten hematopoietic recovery and are over time outcompeted by ucb grafts which provide robust hematopoiesis with low risk of chronic gvhd. the combination of mismatched unrelated hematopoietic progenitors and ucb grafts provides an attractive alternative for older patients lacking hla-idental donors or haploidentical relatives. in planned trials, mismatched donors may be selected based on kir type to further enhance gvl effects. disclosure of conflict of interest: partially supported by miltenyi biotec. haploidentical stem cell transplantation (haplo-sct) is an attractive option for patients who do not have an hlamatched donor. historically it has been associated with high rates of graft rejection, relapse and low incidence of graft versus host disease (gvhd). to decrease these issues we have considered the use of primed bone marrow as stem cell source, early withdrawal of immunosuppressive therapy and the use of donor lymphocytes infusions (dli) in haplo-sct with high-dose post-transplantation cyclophosphamide (ptcy) as main gvhd prophylaxis. to analyse our incidence of acute and chronic gvhd and overall survival (os) in patients with haplo-sct with short course of inmunosupressive therapy. we retrospectively analyzed a cohort of patients who underwent haplo-sct with primed bone marrow as stem cell source, between years and in our centre. gvhd prophylaxis consisted in ptcy ( mgr/kg on days + and + ) and tacrolimus plus mycophenolate (mmf) from day + as recommended by baltimore group. mmf was stopped on day + . tacrolimus was tapered off from day + with withdrawal on day + in patients without gvhd or with active disease. dli were considered if mixed chimerism, relapse or disease progression appeared. the characteristics of the patients are shown in table . results: there was no primary graft failure. eight of patients ( . %) developed agvhd (grade ii-iv) and it was severe (grade iii-iv) in patients ( . %). cutaneous agvhd location was the most common presentation ( patients ( %)) and it was associated with intestinal gvhd in patients. twenty two patients were evaluable for cgvhd. thirteen patients ( %) developed chronic gvhd that was mild, moderate and severe in: ( . %), ( . %) and ( %) patients, respectively. the median time of onset cgvhd was months (range: - ) and it was related with previous withdrawal of the immunosuppression in ( . %) patients, tapered off immunosuppression in ( . %) patients and dli in ( %) patients. systemic treatment was required in / patients but only patients were treated with high doses of steroids ( mg/kg/day). the median days of is therapy in patients who developed gvhd was days (range: - ). dli were used in ( %) patients because of: relapse/disease progression in ( . %) and secondary graft failure in ( . %). two patients achieved complete remission and patients developed cghvd. the median number of dli per patients were ( - ) with a median cd +cell of × /kg (range: × - × /kg). with a median follow-up of months (range: - ), the estimated os at and years after haplo-sct were % and %, respectively. at the moment of this study patients ( . %) were alive, patients in complete remission, in partial response and in progression. eleven ( . %) patients died due to: disease ( ), infections ( ), pleuropericarditis ( ), hepatic veno-occlusive disease ( ) and refractory gvhd. five patients ( . %) are without is therapy and without gvhd symptoms. in our experience, early withdrawal of immunosuppression following haplo-sct with primed bone marrow and posttransplantation cyclophosphamide facilitates the development of chronic gvhd and can decrease the relapses in patients with high-risk hematological malignancies. it is necessary more follow up and more studies to confirm this preliminary results. [p ] disclosure of conflict of interest: none. s myeloid malignancy (n = except patient with saa) received fludarabine (flu)/busulfan ± tbi gy, while lymphoid malignancies (n = ) received flu/tbi gy or cy/ tbi gy. all patients received g-csf-mobilized t-cell replete pbsc from a haplo donor. gvhd prophylaxis was ptcy mg/ kg on day + /+ , tacrolimus (d+ to + ), and mycophenolate (d+ to ). the median duration of follow up of surviving patients is months. median age was ( - ) years, patients ( %) were male, ( %) were african american, and patients ( %) had comorbidity index (hct-ci) ⩾ . all patients had hematological malignancy (except patient with saa) including patients ( %) not in cr. disease risk index was high/very high in ( %) and intermediate in ( %). on the day of transplant, patients ( %) did not receive steroid premedication ( = no-steroid group), while patients ( %) received mg of methylprednisolone minutes prior to pbsc product infusion ( = yes-steroid group). all the following outcomes are described in the ‛no-steroid' vs ‛yessteroid' group, respectively. cumulative rate (ci) of anc engraftment (⩾ /μl) on day + was % ( % ci - %) and % ( % ci - ) (p = . ). ci of platelet engraftment (⩾ /μl) on day + was % ( % ci - %) and % ( % ci - %) (p = . ). primary engraftment failure was observed in patients; in yes-steroid and in no-steroid. no primary engraftment failure was observed with myeloablative tbi ( - gy) (n = ). ci of agvhd gii-iv (day+ ) was % ( % ci: - %) and % ( % ci: - %) (p = . ). ci cgvhd ( year) was % ( % ci - %) and % ( % ci - %) (p = . ). ci of relapse ( year) was % ( % ci - %) and % ( % ci - %) (p = . ). ci of non-relapse mortality (nrm) ( year) was % ( % ci - %) and % ( % ci - %) (p = . ). post-infusion noninfectious fever (d to + ) was observed in / ( %) and / patients ( %). median tmax was f and f (p = . ). only patient in the no-steroid group developed life-threatening cytokine release syndrome and survived. no difference of viral reactivation was noted between groups. cmv reactivation occurred in ( %) and patients ( %), bk reactivation in % (in both groups), hhv in % and %, ebv in % and %. the -month overall survival was % ( % ci - %) and % ( % ci - %) (p = . ). the -month disease-free survival was % ( % ci - %) and % ( - %) (p = . ). t-cell replete haplo pbsc transplant is effective therapy for patients with high-risk hematological malignancies. high-dose steroid premedication with pbsc infusion neither influences transplant outcome nor prevents post-infusion febrile reaction. disclosure of conflict of interest: as discloses grant support (american porphyria foundation), consultation (medpace inc), research support (astellas and fate therapeutics), honoraria (alxion, and spectrum), and royalty for licensing of intellectual property (incysus biomedical). intrabone transplant of unwashed cb in hematological malignancies: engraftment and safety f giglio , s marktel , r greco , m morelli , mt lupo-stanghellini , e xue , l lazzari , m marcatti , m zambelli , c parisi , r milani , s piemontese , a assanelli , c corti , m bernardi , f ciceri and j peccatori unit of hematology and bone marrow transplantation, irccs san raffaele scientific institute, milano, italy and immunohematology and transfusion medicine unit, irccs san raffaele scientific institute, milano, italy cord blood transplant (cbt) in adult patients (pts) is limited by the risk of graft failure or delayed engraftment due to low cell counts. to improve the capacity and speed to engraft, intrabone (ib) cbt technique has been investigated, showing high rate of engraftment and low acute gvhd, also when compared with double cb transplant. cb units washing procedure has been suggested to remove dmso toxic potential effect. we report our experience in adult pts with hematological malignancies receiving ib unwashed cb in an attempt to reduce the loss of progenitor cells and the risks associated with cell-washing procedure. between and we performed allogeneic hematopoietic stem cell transplant (hsct) using unwashed cbu as a source and infusing them ib. all pts were adult and suffering from hematological malignancies. this population was characterized by very high-risk and advanced phase disease. all pts received a cb hsct because of unavailability of sibling or matched unrelated donors. eleven pts received a treosulfanbased myeloablative conditioning regimen and a sirolimusbased ghvd prophylaxis; four pts received busulfan-based myeloablative conditioning and a cyclosporine-based ghvd prophylaxis. cb units were thawed and diluted with albumindextran solution immediately before the transplant. this ‛nowash' dilution was implemented to reduce product manipulation that may results in cell loss. furthermore, graft manipulation risks potential contamination, requires increased technologist time, and delays time to infusion. the ib infusion was performed under local anesthesia and with short conscious sedation, at bedsite in the bmt ward. the infusion was preformed monolaterally or bilaterally according to the volume to be infused. starting from a % dmso concentration in the cb units before the dilution, the graft products contained a median of . % dmso (range: . - . ) at ib-hsct. the median cd + cells infused were . × /kg b.w. (range: . - . ). the median mono-nucleated cells were . × /kg b.w. (range: . - . ). the median cd + t-cells were . × /kg b.w. (range: - . ). the median infused volume was ml (range: - ). no procedure-related adverse events were observed, nor related to the ib technique, neither to the sedation. of the transplanted pts, were evaluable for engraftment ( patient rejected the graft and patients died before day + because of severe infections). all achieved anc . × /l after a median of days (range: - ) and achieved plt × /l at a median of days (range: - ). three patients developed grade iii-iv acute gvhd grade. according to extreme heterogeneity of the population no correlations with relapse incidence and diseasefree survival could be evaluated. ib infusion of unwashed cb is feasible, safe, easy to perform. no adverse events related to the procedure were documented. no dmso toxicity was documented. engraftment was obtained in all evaluable pts. our data confirm that direct ib cbt overcomes the problem of graft failure even when low numbers of cb cells were transplanted, thus leading to the possibility of using of this technique in a large number of adult pts, for whom this approach represents the sole possibility of long-term survival. the ‛no wash' cb dilution can also help the implementation of ib transplant thanks to the easier graft manipulation. [p ] disclosure of conflict of interest: none. lower incidence of cgvhd after cord blood transplantation for hematological malignancies in comparison with hematopoietic stem cell transplantation from other donors: years' experience in a single institute m yoshino, m obiki, m osakie, s ikeno, t sato, m nasashima, y kagaya, n kawashima, t morishita, y ozawa and k miyamura department of hematology, japanese red cross nagoya first hospital the outcome of cord blood transplantation (cbt) for hematologic malignancy was investigated. however the incidence of gvhd is not accurately known. the goal of this study was to compare the outcome of cbt with allogenic hematopoietic stem cell transplantation (allo-hsct) from other sources, mainly unrelated bone marrow (urbm). patients' characteristics: patients who underwent allo-hsct, between and in our hospital were retrospectively analyzed. donor sources were cord blood cell (n = ), urbm (n = ), hla matched sibling bone marrow (sibling bm) (n = ), and hla matched sibling peripheral blood stem cell (sibling pbsc) (n = ). in cbt, the median age was . ( - ), and the diagnosis included aml ( ), all ( ), mds ( ), cml ( ) and other ( ). the disease risk was good in and poor in . disease risk was slightly higher in comparison with other sources. prophylaxis of acute gvhd was tacrolimus, short-term methotrexate ( ), cyclosporine, short-term methotrexate ( ) and others ( ) . the -year overall survival (os) rate after cbt ( cbt ( - . %, engraftment failure . %, acute gvhd . %), relapse . % and other . %. in cbt cases, engraftment failure after allo-sct was observed in cases ( . %) which is higher than that among urbmt ( . %), out of . cbt cases underwent the second allo-hsct and patients achieved engraftment and patients were alive at days after allo-hsct. of them survived at years after allo-hsct. our results suggest that the outcome of cbt has improved in recent years. moreover, cbt has an advantage in the least cumulative incidence of acute/chronic gvhd. cbt may well create the best outcome in the future. disclosure of conflict of interest: none. chronic active epstein-barr virus (ebv) infection is a major type of ebv-associated t/nk-cell lymphoproliferative disorders (lpd) in childhood. however, young adults rarely develop chronic active ebv infection (caebv), and shows more aggressive features than that of childhood. umbilical cord blood transplant (ucbt) is a possible treatment option for caebv patients who have no hla-matched donor, but there is little information available about the efficacy and safety of ucbt for adult patient with caebv. we analyzed six adult patients with caebv who underwent a single-unit ucbt between and at our institute (including a case reported in [ ] ). the diagnosis of caebv was made according to the criteria proposed in [ ] ; persistent infectious mononucleosis (im)-like symptom and detection of increased ebv genomes in peripheral blood mononuclear cells (pbmc). ebv-dna load was measured using real-time quantitative pcr. median patient age at diagnosis was ( - ) years. target cells of ebv-infection were cd +t cells (n = ) or nk cells (n = ). median ebv-dna load was . × copies per microgram of dna in pbmc (range: . × - . × ) at the diagnosis. all patients were given prednisolone and cyclosporine, and then etoposide (n = ) or combination chemotherapy (n = ) before transplant. ebv load has slightly decreased to a median of . × copies (range: . × - . × ), but disease status was active at ucbt in all. median time from the diagnosis to ucbt was days (range: - ). one patient received total body irradiation (tbi) gy + cyclophosphamide (cy) mg/kg + cytosine arabinoside g/m , and the other five patients received fludarabine (flu) + melphalan (lpam) - mg/m or cy mg/kg with tbi gy before ucbt. umbilical cord blood (ucb) was / hla-matched to the recipients. median number of infused ucb cd + cells was . × /kg (range: . - . × ). gvhd prophylaxis was consisted of tacrolimus + methotrexate or mycophenolate mofetil. neutrophil engraftment and complete donor chimerism were achieved in four patients, but two of them developed secondary graft failure (gf) early after engraftment. the other two patients developed primary gf. second ucbt was successfully performed in the patients with gf a median of . days (range: - ) after the first ucbt. ebv genomes in pbmc became undetectable immediately after ucbt. at a median follow-up of days (range: - ), ebv-dna was undetectable or very low, and im-like symptoms were resolved in all cases. however, at - months after ucbt, two patients developed ebv+ b-cell lpd derived from donor cells, that was successfully treated with rituximab therapy. this study suggested that ucbt could eradicate ebv-infected cd + t cell-or nk cell-clones. ucbt can be a treatment option for adults with caebv. rituximab monotherapy was effective for post-transplant lpd from donor b cells. however, a high incidence of gf was observed in patients receiving reduced-s intensity conditioning of flu/lpam or cy /low-dose tbi. further studies are needed to find more optimal regimens for stable engraftment of ucb in adult patients with caebv. there is an increased incidence of ab incompatibility- - %, in allogeneic hematopoietic stem cell transplantation (allohsct) in patients who are russian citizens as a result of the variability of genetic polymorphism in the multi-ethnic population and a significant number of unrelated donors from international bone marrow registries. ab incompatibility in different types of allohsct may be an additional aggravating factor for the development of immunological complications and decrease effectiveness of treatment, but the data is still controversial [ ] . from may to december in raisa gorbacheva memorial institute for children oncology, hematology and transplantation patients with leukemia, malignancies and hereditary diseases were included to the study, who were performed hsct: allogeneic unrelated - ( %); allogeneic related- ( . %); haploidentical - ( . %); umbilical cord blood in patients ( . %). age was - , median- years. patients were predominantly with acute myeloid leukemia- % (n = ), acute lymphoblastic leukemia- % (n = ) and chronic myeloid leukemia - % (n = ). results: in . % of cases (n = ) АВ incompatibility was determined: major- . % (n = ); minor- . % (n = ); combined- . % (n = ). АВ incompatibility in allohsct did not influence overall survival (p = . ) and frequency of acute graft versus host disease (gvhd) (p = . ). also there was no difference in overall survival depending on combination of condition regimen and ab incompatibility: reduced intensity (ric) or myeloablative (mac) (p = . ). an increased frequency of acute gvhd was observed in ric and АВ incompatibility ( . %) compared to mac ( . %, p = . ). ab incompatibility was not a major factor (log worth . ) which influenced the fact and speed of donor's transplant engraftment in comparison to level of hlacompatibility ( . ), hematopoietic stem cell source ( . ) and type of hsct. but the presence of major ab incompatibility increase the period of erythroid recovery (p = . ) as reflected in the higher amount of blood transfusions. complications caused by ab incompatibility were identified in . % of all cases (n = ) including acute and delayed hemolysis, partial red cell aplasia and immune thrombocytopenia. conclusion. the presence of АВ incompatibility is not a limiting factor to perform allohsct, however, it demands high quality prophylaxis and accurate transfusion therapy depending on ab incompatibility type to prevent immune complications. keywords: allogeneic hsct, ab -incompatibility. poor graft function or graft failure have become common indications for infusion of immune-selected cd + cells (‛boost') or second unprocessed allo-hsct, creating the need for remobilization of the same related or unrelated. we retrospectively compared the results of two consecutive cycles of rh-g-csf treatment and peripheral blood progenitor cell collections in related donors cared for at our institution between and . mobilization consisted of the administration of rh-g-csf at a dose of μg/kg per day injected in the evening, and apheresis was started in the morning of the fifth day after the fourth dose of rh-g-csf. collection was performed with a spectra or spectra optia cell separator (terumo bct). eleven out of were haplo-mismatched donors and were hla matched donors. four donors were re-collected because of recipient graft failure and because of poor graft function; in the latter situation, immunomagnetic selection of cd + cells was performed on the collected cell product prior to infusion into the recipient, using the clinimacs medical device, as previously published. median donor age was years (range: - ) at time of first donation, median weight kg ( - ) and bmi ( - ). median delay between mobilizations and was days ( - ). interestingly, the median delay between collections was days ( - ) in the haplomismatched setting and ( - ) in the matched setting. median number of circulating cd + cells/μl after the first injections of rhg-csf was vs at the first and second mobilization cycles (po . , table ). seven out of donors ( %) requested more than one apheresis session to obtain the target number of collected cd + cells during the first cycle, as compared to out of ( %) for the second cycle: this is largely due to the higher target of cd + cells for the second collection, expecting that the median cd recovery after immunomagnetic selection is % in our experience. our study shows that a second cycle of mobilized peripheral blood progenitor cell collection from related donors is associated with a significant reduction in response to hematopoietic growth factors and mobilization capacity. this information allows planning the number of aphereses at the second cycle-and subsequently the number of immunoselection procedures to be carried out-taking into account the higher cd + cell dose target needed for subsequent immunomagnetic selection. cmv reactivation remains one of the main complications after allogeneic stem cell transplantation (hsct), requiring antiviral therapy, causing myelosuppression, prolonged hospitalization, higher treatment costs and mortality. cmv seronegative donors are recommended for cmv seronegative recipients. however data about donor selection for cmv positive (cmv-pos) recipients is not conclusive. some studies showed that selecting cmv-pos donors for cmv-pos patients might be beneficial. cmv-seropositivity is very high in lithuania among healthy blood and bone marrow donors ( %) and even higher among hsct recipients (up to %), so donor selection for cmv-pos recipients is an object of interest. retrospective analysis of cmv reactivations in cmv-pos allogeneic hsct recipients (transplanted during - year in vilnius university hospital) who survived at least months post hsct was performed. data about cmv reactivation frequency, time post hsct, duration, maximal cmv dna copy number at each reactivation collected. cmv reactivation was considered when cmv dna copies detected /ml in patient's blood. statistical analysis conducted using sas . ; student's tests for statistical significance; kaplan-meier methods for overall survival. among allogeneic hsct recipients ( . %) were cmv-pos. cmv-pos allo-hsct recipients were further analysed. of them received graft from cmv-pos (pos/pos group) and -from cmv-seronegative donors (pos/neg group). more patients in pos/neg group experienced cmv reactivation in first months post hsct in comparison to pos/pos group ( . % vs . %, p o . ). pos/neg group patients had more cmv reactivations ( . vs times in months post transplant period, po . ), reactivations were diagnosed earlier post transplant ( . vs days post hsct, p o . ), had longer duration ( . vs . days, po . ) and larger maximal cmv dna copy number ( , vs . copy/ml, p o . ) in comparison to pos/pos group patients. pos/pos group patients showed tendency for better survival than pos/neg group patients, however did not reach statistical significance. in a univariate analysis only hla mis-match and donor cmv seronegativity were factors statistically significantly associated with cmv reactivation. donor cmv serostatus is significant factor selecting donor for allo-hsct recipients. according to our findings, selecting cmv-pos donor for cmv-pos recipient may reduce cmv reactivation frequency and duration. disclosure of conflict of interest: none. selection of the best hsc donor when a matched donor is not available is still a matter of debate, and reports in pediatric population are scarce. this is a retrospective study conducted by brazilian society of hsct (sbtmo), including centers, aimed to compare matched unrelated (matched-urd), mismatched unrelated (mm-urd) and unrelated cord blood (ucb) hsct. all or aml/mds patients o y/o who have received first unrelated hsct between - were included. hla -digit typing was available for urd; for ucb, hla class-i -digit typing. overall survival (os), and cumulative incidence (ci) of agvhd, cgvhd, nrm and relapse were analyzed. on an unplanned analysis, we fitted a lognormal bayesian survival model with random effects, imputing the probabilities of ucb matching at loci. a total of patients were included ( matched-urd, mm-urd and ucb). median age was . y/o. most patients had all ( %). proportion of early disease in matched-urd was higher ( %, against and %). matched-urd were / -digit hla matched (except one, for whom dq was not available), while most of mm-urd ( %) were / hla matched. ucb were loci ( %) or -loci typed ( %). based on previous report on extending -loci hla typing to -loci ucb, we estimated that ucb were - mismatched at loci, had mismatches and had or more mismatches. conditioning regimens were mainly myeloablative, tbi-( %) or bu-( %) based. grafts were in vivo t-cell depleted in % of the patients, not balanced between groups (p = . ). with median follow-up of . years, y os was %, % and % for matched-urd, mm-urd and ucb, respectively (p = . , log rank test). for matched-urd, mm-urd and ucb, ci of grades iii-iv agvhd at months were %, % and % (p = ns); moderate/severe y-cgvhd, %, % and % (po . ); y-relapse, %, % and % (p = ns); and y-nrm, %, % and % (p = . ). we found out that primary graft failure occurred in ( %) of ucb, compared to % in mm-urd and % in matched-urd. when ucb matching probabilities at loci were imputed and analyzed in a bayesian model (controlled for age, gender, disease status and diagnosis, and t-cell depletion), survival was inferior in ucb with + mismatches ( . -times lower median survival, ci . - . ), but not with up to mismatches ( . -time higher median survival, ci . - . ). of note, in vivo t-cell depletion marginally impaired survival ( . -times decrease, ci . - . ). discussion: in our population, overall survival achieved with mm-urd was not different to / matched-urd, despite higher incidence of moderate/ severe cgvhd. on the other hand, survival with ucb was significantly lower. recent report have shown excellent os with ucb compared to / hla-matched urd. ucb cohort inferior results may have been due to hla disparity degree, since survival in ucb with up to mismatches (out of ) was not worse. one limitation of our study is that tnc and cd from ucb units were not available, impairing primary graft failure analysis. we have also found that in vivo t-cell depletion might have a detrimental effect on survival and should be studied further in prospective trials. in conclusion, mm-urd, especially hla / , is a suitable option when a fully hla / matched-urd is not available. ucb matched at least / may also be a good option. disclosure of conflict of interest: none. allogeneic hematopoietic stem cell transplantation (hsct) is a proven treatment for patients with high risk or relapsed hematological malignancy. the probability of having a hla matched family donor is about %. in populations with high consanguinity rates, the probability of having non-sibling matched family donor(mfd) is much higher. to explore the impact of msd vs non-sibling mfd on outcome of hsct recipients, we undertook a single center retrospective analysis of pediatric patients transplanted with the diagnosis of hematological malignancy at our center in the last five years. a retrospective cohort from to current included pediatric patients with hematological malignancies transplanted from family donors, of which were from msd and from non-sibling mfd. hla matched family donors were identified by high resolution allelic typing and were matched of hla loci. diseases were all (n = ), aml (n = ), mds (n = ), jmml (n = ), kml (n = ), nhl (n = ) and hodgkin's disease (n = ). conditioning regimens were tbi or busulphan-based myeloablative in all patients. the median age of the patients was . years (range: month- . years). although peripheral stem cell seemed to be used more commonly in non-sibling mfd recipients ( % vs %), the difference was not statistically significant. the median follow up time for alive patients months ( - months) . two year overall survival and leukemia free survival did not differ between patients with transplantations from msd or nonsibling mfd ( % ± . vs ± . , p = . ) similarly, leukemia free survival was not different between msd and non-sibling mfd transplants ( . % ± . vs . % ± . , respectively). the incidences of grade ii-iv acute gvhd in msd and nonsibling mfd transplants were % and % , respectively. the incidences of relapses were % in msd transplants and % in mfd transplants and the difference was not significant (p = . ). these data show that the results of hsct from nonsibling mfd is comparable to hsct of msd in children with hematological malignancy. our data emphasize the need for extended high resolution family typing for patients in regions where there is high rate of consanguinity. disclosure of conflict of interest: none. [p ] p donor-recipient rh incompatibility is a risk factor for mortality after pediatric matched related allogeneic hematopoietic stem cell transplantation k ghanem , n hariss , z merabi, n kreidieh , n tarek, r saab , s muwakkit , h el-solh and m abboud american university of beirut, department of pediatrics and adolescent medicine and american university of beirut, optimal donor selection is critical to achieve the best outcome after allogeneic hematopoietic stem cell transplantation (allo-hsct). there is no consensus regarding the effect of donor-recipient rh incompatibility on survival after matchedrelated donor (mrd) allo-hsct in children and adolescents. this abstract aims to study this effect in a single-institution cohort over a period of years. this is a retrospective chart review for all patients aged o years who underwent allo-hsct at the american university of beirut medical center between august and june . a total of patients with a median age of years (range: . - years) underwent allo-hsct from mrd for the following diseases: leukemia (n = ), bone marrow failure (n = ), thalassemia (n = ), scid (n = ), metabolic diseases (n = ) and lymphoma (n = ). the stem cell source was bone marrow for patients ( %) and mobilized peripheral blood stem cells for patients ( %). the grafts contained a median of . × /kg total cd cells. tbi was used in patients ( %). all but patients achieved sustained neutrophil and platelet engraftment. after a median follow-up of months (range: - months), the -year overall survival rate was % ( % ci: - %). by multivariate analysis using cox proportional hazard regression model looking at the following factors for overall mortality: diagnosis, recipient's age, donor's age, the use of tbi, stem cell source, cd count, donor-recipient abo incompatibility, donorrecipient rh incompatibility, and donor-recipient sex-mismatch, the only statistically significant risk factor for mortality was donor-recipient rh incompatibility (hr: . , p: . ). this risk was not statistically significant when looking at transplantrelated mortality (hr: . , p: ) and relapse-related mortality for malignant diseases (hr: , p: . ). there was no association between the incidence of acute or chronic gvhd and rh incompatibility. donor-recipient rh incompatibility was associated with an increased risk of mortality in children and adolescents undergoing mrd allo-hsct. further studies with larger number of patients are needed to confirm this finding. disclosure of conflict of interest: none. effect of iron or vitamin b deficiencies on in vitro colony forming capacity of peripheral blood-derived hematopoietic stem cells in children ny Özbek, mm zabun, y köksal and m Özgüner iron deficiency (id), id anemia (ida)and vitamin b deficiency (vit-b d) are common disorders in developing countries. in urgent situations, children with these disorders could be donors before treatment. in this study, we investigated capacity of peripheral blood-derived hematopoietic stem cells to develop colony-forming units (cfu) in children with id and vit-b , in vitro. patients and methods: we included children (age months- years) in the study in groups: children with id (n = ); children with ida (n = ); children with vit-b d (n = ); children with both id and vit-b d (i/ vit-b d; n = ); and control children (n = ) who has normal peripheral blood findings, and normal ferritin and vit-b levels. from each child complete blood counts (cbc), and levels of ferritin, vit-b , and cero-reactive protein (crp) have been obtained. who criteria, adjusted for age and sex, have been used for definition of anemia, id, ida and vit-b d. four ml peripheral blood drawn into tubes with edta has been used for cfu analysis. mononuclear cell suspension ( . × cell/ml), obtained from peripheral blood by ficoll-hypaque density gradient separation method, has been cultured in dishes containing semi-solid agar culture medium (methocult, h classic, stem cell technologies, canada) in appropriate conditions. after weeks, number of cfu colonies [burst forming erythroid (bfu-e); colony forming unitgranulocyte macrophage (cfu-gm); colony forming unitgranulocyte-erythrocyte-monocyte-megakaryocyte (cfu-gemm)] have been investigated by an inverted microscope. results: statistical analysis showed no difference between groups for age, sex, crp levels, and cfu-e, cfu-gm and cfu-gemm numbers. however, expected differences between groups were present concerning mean values of hemoglobin, ferritin and vit-b levels, mean corpuscular volume (mcv), and red cell distribution width (rdw) ( table ) . discussion: this study shows in vitro proliferation capacity of peripheral stem cells has not been influenced by id, ida, vit-b d, or i/vit-b d. our results may indicate normal grafting ability of peripheral stem cells obtained from donors with iron, vit-b or i/vit-b deficiencies for hematopoietic stem cell transplantation. however, in vivo analysis should also be performed in order to reach a definite conclusion. [p ] disclosure of conflict of interest: none. efficiency of day compared to day stem cell mobilization in allogeneic donors h al-gaithi , s al-mamar , m al-huneini , d dennison , s al-kindi , k al-farsi and m al-khabori hematology residency training program, oman medical specialty board; hematology department, sultan qaboos university hospital granulocyte colony stimulating factor (g-csf) given for - days is commonly used for mobilization of allogeneic stem cell donors. the optimal days of g-csf administration is still debatable. the primary objective of this study is to compare the yield of stem cell mobilization, assessed using cd + cell count, between day and day . secondary objectives include the assessment of the impact of donor's age, weight, mean corpuscular volume and blood group on the difference in the cd + cell count. in this retrospective study we included all allogeneic stem cell donors mobilized with g-csf for days from january till october in the bone marrow transplantation unit at sultan qaboos university hospital. of donor records reviewed, were with available data and selected for the study. descriptive and analytical statistics were performed using stata . . we included donors with median age and weight of years and kg, respectively. the median day wbc and cd + cell count were . × /l and × /l respectively; while the median day wbc and cd + cell count were . × /l and × /l, respectively, (figure) with a statistically significant difference from day (p o . ). in the multivariable model, there were no significant impact of donor's age (p = . ), weight (p = . ), height (p = . ) and mean corpuscular volume (p = . ) on the difference in cd + cell yield. however, donor's blood group ab predicated a significantly higher difference (p = . ). six days of g-csf mobilization achieves higher cd + cell count than days in allogeneic stem cell donors especially in donors with blood group ab. however, cd + cell count on day is high enough to allow for successful mobilization. appropriately designed prospective trial is needed to confirm these results. disclosure of conflict of interest: none. there are known differences between individuals on an unrelated hsc donor register who decide to proceed with verification typing (vt) vs those who choose not to. in the anthony nolan registry, white british donors are more than twice as likely as other ethnic groups to continue with testing at vt (or . ; po . (unpublished data)). the purpose of this study was to explore differences in key characteristics between white british donors and british donors from other ethnic groups with a view to developing interventions to reduce vt stage attrition. study recruitment occurred april -may . all donors not proceeding at vt were invited to participate, and a stratified random sample of those proceeding at vt were recruited to meet pre-determined targets for each ethnic group. data were collected via structured interview (telephone or online). broad categories of participant characteristics were assessed: demographic, culturally related, psychosocial, and donation-related. measures were previously validated scales with established psychometric properties either created for, or used in other donation-related settings. for analyses donors were divided into two groups based on ethnicity: white british (wb), and non-white british (nwb). results: wb donors and nwb donors completed interviews donors proceeding at vt were more likely than their counterparts to participate in the study ( % vs %, p o . ). mean donor age was . with no difference between ethnic groups and % of donors in both groups were female. nwb were statistically more likely to have completed higher education, and have a stronger religious affiliation. in contrast they were less likely to be blood or organ donors. nwb also described greater mistrust of the medical system and of hsc allocation. nwb donors were more likely to have joined the register at a recruitment event (p= . ) or a place of worship (p= . ), while wb donors were more likely to have joined online (p= . ). wb donors reported significantly higher scores regarding feeling well informed about donation both at the point of joining, and at the point of vt and were more likely to remember joining the register and the two donation methods. this study highlights important differences in demographics, culturally related variables and donor interaction with the register between white british donors and donors from other ethnic backgrounds. given the higher rate of vt attrition in nwb donors, these findings could be used to tailor interactions/information given to donors on the register to ensure their priorities are addressed. disclosure of conflict of interest: none. data on mismatched family donor transplants for myelofibrosis are scarce due to the risk of poor engraftment, gvhd and exclusion from trials. outcomes from such transplants performed between and reported to the ebmt are presented. sixty-nine patients, median age ( - ) years; ( %) male, ( %) had primary, ( %) had secondary myelofibrosis ( from et, from prv and others) and unknown ( %). jak v f was mutated in / . karnofsky performance status was % in %; median time from diagnosis to allograft was . (range: . - ) months. the donors were predominantly male ( %), median age ( - ) years, hla mismatched at locus in ( %) and or more loci in ( %). donor-recipient serology was cmv − / − in ( %) ± in ( %), − /+ in ( %) and +/+ in ( %) missing ( %) . bone marrow was used in ( %) and peripheral blood in ( %). the median total nucleated cell count (tnc) was . × /kg (range: . - × /kg) (n = ). the median cd + cell dose was . × /kg [p ] s (range: . - . × /kg)(n = ). patients. conditioning was myeloablative in ( %) and ric in ( %). predominant conditioning regimes were fludarabine, busulphan, atg (fbatg) and thiotepa, busulphan, fludarabine (tbf n = ). tbi was administered in ( %) and t cell depletion in vivo in ( %) and ex vivo in ( %) patients. gvhd prophylaxis varied with post transplant cyclophosphamide administered in / ( %) and atg in / patients ( %).neutrophil engraftment occurred in ( %) patients at a median of days (range: - ). primary graft failure ensued in ( %) and secondary graft failure in ( %) patients at a median of (range: . - ) months. eleven patients had a second allograft at a median interval of ( - ) months. responses to the first allograft censoring for a second allograft, data available in patients, showed that complete remission was achieved in patients ( %), ( %) were never in cr and ( %) were not evaluable. relapse occurred in ( %) of patients at a median interval of ( . - . ) months. the cumulative incidence (ci) of grade ii-iv acute gvhd (agvhd)was % ( % ci - %) and for grade iii-iv agvhd at was % ( % ci - %). data for chronic gvhd (cgvhd) was valid in patients of whom % developed cgvhd. the ci of cgvhd at years was % ( % ci - %):ci of limited cgvhd was % ( % ci - %) whereas the ci of extensive cgvhd was % ( % ci - %). median follow-up was ( % ci - ) months. the and year os was % ( % ci - %) and was % ( % ci - %). the and year rfs was % ( % ci - %) and % ( % ci - %). the -year ci of relapse was % ( % ci - %). the year nrm was % ( % ci - %), which increased to % ( % ci - %) at years. thirty patients died due to infection ( , %), gvhd ( , %), organ damage or failure ( , %), relapse/disease progression ( , %) and secondary malignancy or ptld ( , %) unknown . there was no significant effect (univariate analysis) of recipient or donor gender, degree of hla mismatch, cmv matching, primary or secondary mf, chronic vs advanced disease at transplant, conditioning intensity or regimen, gvhd prophylaxis with atg or post transplant cyclophosphamide or stem cell source on overall survival. the data are encouraging for patients with myelofibrosis, with engraftment, pfs and os being attained with limited severe chronic gvhd from family mismatched donors. disclosure of conflict of interest: none for all other authors, fc consulting with molmed. feasibility of salvage second allogeneic stem cell transplantation for disease relapse or graft failure: a single centre experience g battipaglia , , d salvatore , , r dulery , f giannotti , f malard , e brissot , s sestili , f isnard , s lapusan , a-c mamez despite high rates of toxicity and mortality, a second salvage allogeneic stem cell transplantation (second allohsct) might be an option to consider in patients experiencing disease relapse or graft failure after first allohsct. we retrospectively analyzed outcomes after second allohsct in a cohort of patients ( males and females) transplanted either for disease relapse (group , n = ) or graft failure (group , n = ) between and in a single centre in france. median age at second allohsct was (range: - ) years. diagnoses were acute myeloid leukemia (group : n = ; group : n = ), acute lymphoblastic leukemia (group : n = ; group : n = ), myelodysplastic syndrome (group : n = ; group : n = ), myeloproliferative neoplasm (group : n = ; group : n = ), bone marrow failure (group : n = ; group : n = ). median time from first allohsct to second allohsct was (range: . - ) months in group and . (range: - ) months in group . graft source for the second allohsct were: haploidentical bone marrow (group : n = ; group : n = ), haploidentical pbscs (group : n = ; group : n = ), cord blood (group : n = ; group : n = ), matched unrelated pbsc (group : n = ; group : n = ). at time of second allohsct, patients were in cr and presented active disease in group . conditioning regimen was myeloablative in patients (group : n = ; group : n = ), reduced intensity (ric) in cases (group : n = ; group : n = ). a sequential schema consisting of a combination of thiotepa, etoposide and cyclophosphamide followed by a fludarabine and busulfanbased ric was used in out of patients with active disease in group . sixteen patients received atg as part of the conditioning regimen for second allohsct (group : n = ; group : n = ). all but one patient engrafted, at a median time of (range: - ) days. cumulative incidence of acute and chronic gvhd were ± % and ± %, respectively, - ) months, non-relapse-mortality (nrm) and relapse incidence (ri) were ± % and ± %, respectively, while disease-free (dfs) and overall survival (os) were ± % and ± %, respectively, for the entire cohort. in all, patients died of infections (n = ), hematological disease (n = ), gvhd (n = ), hemorrhage (n = ) and for unknown causes (n = ). main outcomes of patients in group were: ri ± %, nrm ± %, agvhd ± %, cgvhd ± %, dfs ± %, os ± %, respectively. main outcomes of patients in group were: ri ± %, nrm ± %, dfs ± %, os ± %, agvhd ± %, cgvhd ± %. historically, a second allohsct was hampered by significant morbidity and mortality. however, the advent of reduced-toxicity conditioning regimens and improved supportive care allowed to significantly improve the results of patients receiving a second allohsct as suggested from the above results. therefore, a second allohsct could be considered as an option to rescue a certain number of patients experiencing disease relapse or graft failure, for which prognosis is very poor. decision is to be discussed on a case-by-case basis. disclosure of conflict of interest: none. haploidentical hematopoietic stem cell transplantation with post-transplant cyclophosphamide for patients with high-risk hematologic malignancies am carella department of oncology and hematology, irccs casa sollievo della sofferenza, san giovanni rotondo allogenic hematopoietic stem cell transplantation (sct) has been increasingly used for treatment of adult with high risk hematologic malignancies. for patients lacking an hlamatched related or unrelated donor, unmanipulated haploidentical (haplo)-sct is a potential alternative. haploidentical transplantation performed with post-transplantation cyclophosphamide (ptcy)-based graft-versus-host disease (gvhd) prophylaxis has been associated with favorable outcomes for patients with acute leukemia and lymphomas we analyzed outcomes of patients with hematologic malignancies who received t-cell-replete haematopoietic stem cells and posttransplantation cyclophosphamide after myeloablative or nonmyeloablative hla-haploidentical donor transplantation. the median age was years ( - ); twelve patients were in first remission (cr ), in second remission (cr ) and had an active disease. ). the diagnosis was acute leukemia (n = ), myelodisplastic syndrome (n = ), hodgkin disease (n = ) non hodgkin lymphoma (n = ) and multiple myeloma (n = ). median follow-up was days. stem cell source was bone marrow (bm) for patients, and peripheral blood (pb) for . myeloablative conditioning (mac) was used in patients and reduced intensity regimen (ric) in patients. thirty one patients were first grafts, the others underwent previous autologous sct (n = ) or mud (n = ). gvhd prophylaxis s consisted in pt-cy on days + and + , cyclosporine (from day + ), and mycophenolate (from day + ). the median day for neutrophil engraftment was day + ( - ). no graft failure was observed. chimerism was evaluable in patient; on day + all patients had % donor chimerism on marrow cells median follow-up was days. the cumulative incidence of acute gvhd grade ii-iv was %, grade iii-iv % and chronic gvhd %. one-and -years os was . % and . %, respectively. with a median follow-up for the surviving patients of days ( - ), the cumulative incidence of transplant-related mortality (trm) is %, and the relapserelated death is %. thus, we demonstrate excellent rates of engraftment, gvhd, and trm in adult patients treated with haploidentical hematopoietic stem cell transplantation with post-transplant cyclophosphamide. this approach is a widely available, safe, and feasible option for adult patients with high risk hematologic malignancies, including those with a prior history of myeloablative bmt and/or those with co-morbidities or organ dysfunction, also for patients with active disease at the time of transplant. disclosure of conflict of interest: none. it has recently been shown that t-replete allogeneichematopoietic stem-cell transplantation (allo-hsct) from a haploidentical donor (haplo-id) could be a valid option when a matched donor is not available. unfortunately, the worldwide donor registries comprise mainly donors of caucasian origin and patients of non-caucasian origin have a much lower chance of finding a matched unrelated donor (mud). the lengthy period of international search when required and the financial burden of this process are considered as additional significant limitations. at the american university of beirut medical center (aubmc) in lebanon, we started the mud program in and haplo-id hsct program in . we report here our experience in this two groups of patients. patients and methods: we have transplanted patients from a haplo-id donor since and compare their outcome with the patients transplanted from a mud since . the patients and transplant characteristics are listed in the table . the groups were comparable except for conditioning. patients in haplo-id group received two days of posttransplant high-dose cyclophosphamide (pt-hdcy) followed by cyclosporine a (csa) and mycophenolate-mofetil while patients in the mud group received pre-transplant antithymocyte-globulins and csa starting on day- . all patients engrafted in the mud group, while one patient did not engraft in the haplo-id group, the patient had refractory all transplanted with progressive disease, and died on day + . the median of anc /mm was days ( - ) vs days ( - ) in the haplo-id and mud groups, respectively. fourteen patients from the haplo-id group developed grade acute graft-versus-host disease (agvhd) vs one after mud-hsct. two patients haplo-id group developed limited cgvhd and none after mud grafts. six patients relapsed in the haplo-id group vs three patients in the mud group. two and three patients died from non-relapse mortality in the haplo-id and mud group, respectively. at the last follow-up, patients are still alive in the haplo group vs patients in mud group and all of them are in cr. we conclude that t-replete haplo-id hsct followed by pt-hd cy is associated with promising results or at least comparable to patients transplanted from mud. haplo-id hsct seemed to be safe and feasible in patients with high risk hematological malignancies. finally, because of the obvious advantage in rapidly finding a donor ( haplo transplants in three years vs mud transplants in years), development of haplo-id hsct is warranted to satisfy the regional needs. [p ] disclosure of conflict of interest: none. haploidentical hematopoietic stem cell transplantation (haplo-hsct) using t-cell-replete (tcr) grafts and posttransplantation cyclophosphamide (ptcy) provides a curative approach for patients with high-risk mds/aml lacking a conventional hla-matched donor. in children and adults haplo-hsct using ptcy as gvhd prophylaxis seems to be safe with low treatment related morbidity and mortality (trm). however, few data are available for elderly patients with advanced disease. we retrospectively analyzed the outcome of patients with mds (n = )/aml (n = ) age - years (median age years; patients - years, patients ⩾ years; male), who underwent tcr haplo-hsct with high-dose ptcy at our institution between january and november . disease was active in patients while had achieved cr. patients failed previous allo-hsct. pretransplantation risk factors were scored using the hematopoietic cell transplantation-specific comorbidity index (hct-ci) which was ⩾ in patients (median hct-ci = , range: - ). a sequential therapeutic concept using either flamsa (n = ) or clofarabine (n = ) as cytoreduction was used prior to reduced intensity conditioning (ric) in all but patients. ric consisted of fludarabine/cyclophosphamide combined with either melphalan (n = ), busulfan (n = ) or gy tbi (n = ). post-grafting immunosuppression consisted of cyclophosphamide, tacrolimus and mmf in all patients. % received a bone marrow graft. one graft rejection occurred. neutrophil and platelet engraftment was achieved in % and % of evaluable patients, respectively at a median of ( - ) and ( - ) days. acute gvhd grade i-iii occurred in % of the patients whereas no grade iv agvhd was observed. chronic gvhd presented in %. it was most frequently assessed as mild to moderate ( pts). only patients developed severe cgvhd; no gvhd related death was observed. cmv reactivated in of patients at risk, one patient developed cmv disease (pneumonia). no ebv reactivation or ptld occurred. one-year trm was %. / ( %) patients relapsed, three within the first days after haplo-hsct. at a median follow up of months (range: - months) estimated one-and two-year overall survival (os) was / %, respectively. when stratified by age, estimated one-and two-year os was / % in patients o years and / % in patients ⩾ years (p = . /p = . ). one-and two-year progression-free survival (pfs) was / %, respectively. stratified by age estimated one-and two-year pfs was / % in patients o years and / % in the elderly (p = . /p = . ). unmanipulated haploidentical allografting using ptcy-based gvhd prophylaxis in high-risk mds and aml patients aged over years is safe and well tolerated resulting in acceptable trm. a remarkable survival outcome can be achieved in elderly high-risk aml/mds patients with significant comorbidities. disclosure of conflict of interest: none. key performance indicators to assess the quality of a collection facility: experience of a single center s roncon*, c pinho , f bordalo , s lopes , s ferreira and f amado allogeneic hematopoietic stem cell transplantation (allo-hsct) has evolved into an effective immunotherapy for the treatment of a variety of disorders. when patients do not have a familiar matching donor, transplant centers (tc) search for an unrelated and volunteer donor. this one must be previously evaluated by the collection center (cc) to donate peripheral blood stem cells (pbsc) or bone marrow (bm); lymphocytes can also be asked after allo-hsct. this work aims to evaluate our performance as cc, ensuring donor safety, quality of cell therapy products (ctp) and the accomplishment of tc requirements. we retrospectively analyzed all the requests of ctp collections sent by the portuguese registry from to . countries of destination, number and type of ctp were determined. we established eight key performance indicators (kpi) classified into four categories: response time; product quality; satisfaction of patients and donors; and on-site donor motivation. the intended target was defined by the mean result obtained in the first half of (excluding kpi- ). written comments from donor center (dc) and tc were received by email or written in the local notebook. the donor's answers were obtained through a survey given on the collection day. a total of requests were assessed: pbsc, bm, lymphocytes and cancellations; % were sent to europe ( / to portugal), % to america and % to oceania; / were withdrawn by tc ( patients died, presented progressive disease and had a better hlamatched donor) and / by dc ( donors not cleared and refused). the results obtained with kpi- , - , - and - exceeded the intended target (table ) . after the first kpi- results, we verified a positive evolution. we took an average of days of delay in sending donor clearance. however, there is no holdup in the ctp delivery, as demonstrated by kpi- . regarding kpi- it is important to notice that % of ctp with a cell number less than requested were bm and lymphocytes; when pbsc was considered separately, the result increased ( % vs %). analyzing kpi- , % (n = / ) of the contaminated ctp were bm. concerning kpi- , - acknowledgments and - commitment, we recognize that our initial targets were too ambitious ( %). the kpi- shows a low number of complaints (n = ): one due to a misreading of the request and three to communication failures; all were properly examined and rectified. a good general status was guaranteed in almost all the donors (kpi- ). the decrease of kpi- is due to the fact that one donor refused to proceed after three postponements of the collection date by tc. table -key performance indicators of the quality of our activity as a cc. the overall good level of our results reflects an extremely professional performance as a cc. we consider that these kpi should be continuously monitored with the purpose of earlier detect any deviation of the stated goals and assess the progress against settled strategies. we further suggest the establishment of universal indicators in order to standardize [p ] practices, share expertise and improve the quality of services and products provided to patients and donors. two year later, the patient had a genoidentical allogenic stem cell transplant (from the bone marrow stem cell of his sister, who was years old, hla compatible). he had a reduced intensity conditioning, associating busulfan, fludarbine and antilymphocyte serum. months from the asct, he was in complete remission with % donor chimerism. years after the asct, the patient presented a progressive thrombocytopenia without any other peripheral causes. the bone marrow aspiration initially showed a refactory cytopenia with multilineage dysplasia. the patient was followed up during months, and then a second bone marrow aspiration has shown a refractory anemia with excess blasts raeb . a cytogenetic study has every time demonstrated a female karyotype ( ,xx) on mitoses out is , and chimerism was % donor. the diagnosis of the myelodsplastic syndrome of the donor cells was approved. the patient was treated by azacitidine ( mg/m , from j to j , j = j ). after cycles, the patient was in complete hematologic response (normalization of the platelet count) and a partial bone marrow response (normalization of the blasts rate but persistence of the signs of dysplasia). he received more cycles, and presented hematologic relapse (reemerging of thrombopenia). a phenoidentical allogenic stem cell transplantation was suggested. conclusion the occurrence of mds on the donor cells is rare. these anomalies are secondary to intrinsic factors (of donor) or extrinsic factors )of the transplant recipient). the treatment is not definitely determined. disclosure of conflict of interest: none. nk-cell alloreactivity based on kir/ligand mismatch in the donor vs recipient direction provides better graft-versustumor effect in patients with active hematological malignancies undergoing allogeneic t-replete haploidentical transplantation followed by post-transplant cyclophosphamide a wanquet , , s bramanti , , s harbi , s fürst , f legrand , c faucher , a granata , p-j weiller , , c picard , b calmels , c lemarie , c chabannon , , , l castagna , , d blaise , , and r devillier , haplo-sct have been developed in the past years with very interesting results in high risk patients. gvhd prophylaxis using post-transplant cyclophosphamide (pt-cy) recently allowed extending the use of unmanipulated haplo-sct. it was shown that nk alloreactivity, triggered by donor-recipient inhibitory kir gene-gene mismatches, could lead to better outcomes and survival in the setting of in t-cell-depleted haplo-sct. however, few data is available on the impact of kir-ligand mismatch on the outcome after t-replete haplo-sct with pt-cy. we thus assessed the impact of nk alloreactivity on the outcome of patients who received haplo-sct followed by pt-cy. we retrospectively collected the data from patients from two centers who were treated for various high risk hematological diseases and underwent a haplo-sct with pt-cy from december to december . we assessed the kir-binding epitope in hla-c and hla-b molecules for all patients, and we predicted nk cell alloreactivity in the donor vs recipient direction via the immune polymorphism database kir ligand calculator, based on the kir-ligand mismatch between donors and patients. because disease status at the time of haplo-sct is one of the most important predictor of outcome, we separately analyzed two cohorts of patients: those transplanted in complete remission (cr group) and those transplanted with active disease (no cr group). using a multivariate cox model (adjusted by disease type, age and conditioning), we therefore evaluated the impact of nk alloreactivity on outcome in both cr and no cr groups. we analyzed patients with a median age of y ( - ). they were mostly transplanted for lymphoma (n = , %) or aml/ mds (n = , %). patients mostly received a tbi-based nonmyeloablative conditioning regimen (n = , %) and pbsc as graft source (n = , %). eighty one and patients were transplanted in cr and in no cr, respectively. nk alloreactivity was found in / cr patients ( %) and / no cr patients ( %). with a median follow up of months ( - ), cr patients had a significantly better outcome than those in the no cr group ( -year pfs % vs %, respectively, p o . ). in no cr patients, multivariate analysis showed that nk alloreactivity was significantly associated with reduced the risk of relapse (hr = . , p = . , figure a ) with no increase of both acute (hr = . , p = . ) and chronic gvhd (hr = . , p = . ), and nrm (hr = . , p = . ). this led to significantly better pfs (hr = . , p = . , figure b ) and a trend for better os (hr = . , p = . ). in contrast, in cr patients, we found no difference in outcome according to nk alloreactivity for all end points (acute gvhd: hr = . , p = . ; chronic gvhd: hr = . , p = . , nrm: hr = . , p = . , relapse: hr = . , p = . , figure c ; pfs: hr = . , p = . , figure d ; os: hr = . , p = . ). our results suggest that nk alloreactivity provides better disease control with no increase of gvhd, especially in patients transplanted with active disease. thus, donor selection should rely on the prediction of nk alloreactivity. this may contribute to improve outcome of these patients with high risk of relapse after transplantation, underlining the need of a specific strategy of donor search, and the promising perspective of early post-transplant nk-cell-based immunotherapy. haploidentical bone marrow transplantation (haplo-bmt) with post-transplant cyclophosphamide (pt-cy) is being increasingly used, in the last five years, for patients lacking a suitable hla-matched donor. genoa study (eudract number: - - ) provides for a modified gvhd prophylaxis platform compared to the original baltimora protocol. aim of the study: in this study we assessed outcomes in consecutive patients transplanted from a haploidentical donor for haematological malignancies. all patients received a uniform gvhd prophylaxis: cyclosporine (csa) starting on day , mycophenolate (mmf) starting on day + , and post transplant cyclophosphamide (pt-cy) mg/kg, on days + and + . all patients received a myeloablative conditioning consisting of thiotepa, fludarabine, busulfan (three doses n = or two doses n = ), or tbi, fludarabine (n = ). the median age was years ( - ); at transplant ( %) patients were in remission of disease (cr and cr ), and had an active disease ( %); all patients were first grafts. the diagnosis were acute myeloid leukemia (n = ), myelodisplastic syndrome (n = ), acute lymphoblastic leukemia (n = ), myelofibrosis and myeloproliferative diseases (n = ), non hodgkin lymphoma (n = ), chronic lymphocytic leukemia (n = ) and multiple myeloma (n = ). the median follow up was days (range: - days). the median infused mononucleated cells was . × e /kg (range: . - . ). seven patients died before engraftment, and ( %) had autologous recovery: ( %) after conditioning with doses of busulfan. full-donor chimerism on day + was reached in ( %) patients. the median day for neutrophil engraftment was day + (range: - days). the cumulative incidence of grade ii-iv and iii-iv acute gvhd (agvhd) was % (n = ) and % (n = ), respectively. two years cumulative incidence of moderate-severe chronic gvhd (cgvhd) was % (n = ).sixty one ( %) patients experienced haemorragic cystitis. at years the cumulative incidence of non relapse mortality (nrm), relapse and relapse related death was % (n = ), % (n = ) and % (n = ), respectively. causes of death were infections (n = ), hemorrhage (n = ), gvhd (n = ), secondary neoplasia (n = ) and relapse (n = ). at years of follow up overall survival and disease free survival was . % and %, respectively. at the same time overall survival rate was % for patients in remission and % for patients with active disease at transplant(p o . ). in conclusion, a modified pt-cy as gvhd prophylaxis and ma conditioning regimen followed by haploidentical bmt results in a low risk of agvhd and cgvhd and encouraging rates of trm and dfs. disclosure of conflict of interest: none. the italian bone marrow donor registry (ibmdr), in collaboration with admo (associazione donatori di midollo osseo) since has implemented, as part of the donor enrollment strategy, public enrollment events (pe). our donor center (dc) has taken part to those events since the first years. one or more clinician (or trained biologist) has been present to pe to inform the potential donors, evaluate the candidates and supervise the collection of biological fluids. all the local permission where obtained. aim: aim of this study was to compare the compliance of the donor enrolled in pe with donors enrolled at our dc institutional site. we prospectively evaluated all the donors recalled for further evaluation and/or for requalification in the years and at our dc itmi . we defined possible results for the call: ‛success' (the donor was eligible and accepted to be evaluated, or only temporarily ineligible) ‛not eligible' (the donor was definitively ineligible) and ‛consent denied'. results: a total of donors were called back in the years and ( not found). eightyfour recalled donors had been enrolled after . among them ( . %) had been enrolled at the dc and during pe ( . %). the two populations were not different for age at the call, age at enrollment and gender (table ) . [p ] when evaluating the probability of obtaining a "success", no significant difference was found between the two populations: . % vs. . % (chi square p= . ). no significant difference was also found for the "not eligible" and the "consent denied" categories. of note, when we turned to the whole donor population we had called back (median age , range - ), the probability of "success" and "consent denied" were not related to donor age, and time from enrollment to recall, whereas donor ineligibility was (spearman test p= . and . ). public events with the presence of an adequate trained medical team represent a valid option for the enrollment of new unrelated donors. disclosure of conflict of interest: none. the search for hematopoietic stem cell unrelated donors in patients with malignant hemopathies with not-sibling matched family donor: the experience of a center a pérez , r goterris , m gómez , s blanco , a segado , c arbona , jch boluda , m poch and c solano hematology department, hospital clínico universitario, valencia unfortunately, as few as - % of patients will have an hlaidentical matched sibling donor available for hematopoietic stem cell (hst) donation. the search for an unrelated donor (urd) (adult or cord blood) is often the best option for those patients lacking a suitable matched donor. below we describe the experience with the search for an unrelated donor in our center. between september and march the search for urd was activated for patients. the median age of the patients was years (range: . - ), % were under years and % were males. acute myeloid leukemia (n = ), acute lymphoblastic leukemia (n = ), non-hodgkin's lymphoma (n = ), chronic/prolymphocytic lymphocytic leukemia (n = ), hodgkin's lymphoma ((n = ), multiple myeloma (n = ), chronic myeloid leukemia (n = ), philadelphianegative myeloproliferative neoplasms (n = ), myelodysplastic syndrome (n = ), aplastic anemia/paroxysmal nocturnal hemoglobinuria (n = ), others (n = ). the disease status in hematological malignancies was: first cr (n = ), second cr (n = ), pr (n = ) and refractoriness (n = ). the donor type requested at the activation of the search was an adult (n = ), umbilical cord blood (n = ) and two options (n = ). results: a compatible donor was found in patients ( % of the series) after a median of days (range: - ) from the activation of the search. the degree of adult donor compatibility (not available in cases) was: complete hla identity ( / : n = , / : n = ); an hla difference ( / : n = , / : n = ); lower degree of compatibility (n = ). the degree of umbilical cord blood compatibility: identity ⩾ / (n = ). a total of patients ( %) were transplanted, from adult donor and from umbilical cord blood. the median time between the activation of the search and the hst transplantation was months (range: . - ), being . months for acute leukemia and . months for other pathologies, and between the location of the donor and the hst transplantation days (range: - ), being days for umbilical cord blood and days for an adult donor. there were cancellations of the urd search ( % of the total) for the following reasons: clinical status of the patient (n = ), performing a haploidentical transplant (n = ), transplant center does not consider (n = ), norms of the registry (n = ) and loss of indication of transplantation (n = ). the median time from the beginning of the search to its cancellation was . months (range: . - ). at the time of analysis, the median follow-up of the patients is months. the survival of the series in the years is % and % for patients transplanted from urd. % of the searches activated in our center allowed the localization of a urd with an adequate degree of hla compatibility. however, only % of the patients for whom the search was activated were finally transplanted. the most frequent cause of cancellation of the procedure was the clinical deterioration of the patient. disclosure of conflict of interest: none. the leukemic transformation of otherwise healthy donor stem cells provides a useful in vivo model to study the mechanisms involved in leukemogenesis. we report two cases of donor cell-derived haematological malignancy in which wholeexome sequencing (wes) was performed in bone marrow (bm) samples from recipient at different times after allogeneic hematopoietic stem cell transplantation (allo-hsct) in order to study the dynamics of emergence of mutations that precede the development of donor cell leukemia (dcl) and donor cell myelodysplastic syndrome (dc-mds). case : a -year-old female diagnosed with lymphoblastic leukemia-b t( ; ), who developed acute myeloid leukemia (aml) with normal karyotype, npm +of donor origin months after unrelated cord blood transplantation (ucbt). case : a -year-old male diagnosed with mantle cell lymphoma, who developed mds ,xx,- ,del( )(p ) of donor origin, months after allogeneic bm transplantation from his hla-identical brother. the donor also developed mds several months later. wes (sureselect-xt human-exon mb) was performed by next generation sequencing (hiseq) on donor stem cells (scs) infused as well as on bm samples from recipient after allo-hsct. the exome of donor scs and bm samples, from case , were aligned to the human reference genome (grch /hg ) and donor scs and bm samples were aligned to grch / hg in the second case. in both cases non-synonymous variants in the coding regions or synonymous variants in splice regions of genes related to leukemia were selected. in addition, bm samples were matched to their scs and to prior bm samples to identify the acquired variants. variants meeting such criteria were evaluated with functional predictor software's (sift, polyphen and mutation taster). wes analysis revealed progressive emergence of multiple somatic mutations probably related to the development of leukemia in bone marrow samples post allo-hsct ( figure ). both scs showed alterations that may be involved in leukemogenesis. (case : sh b and case : kmt c, kmt a, arhgap and monosomy ). somatic mutations, acquired over time, fall into genes that play well-established roles in signalling pathways (ras-mapk, pre-mrna splicing factor, apoptosis, dna doublestrand break repair, dna replication and so on). mutations in leukemic subclones that disappear after chemotherapy were indentified, as well as the acquisition of new mutations in resistant subclones. we propose a possible model of leukemogenesis in these cases ( figure ). the present study reveals a process of sequential clonal expansions, promoted by the acquisition of additional somatic mutations in donor hematopoietic cells. detection of heritable or acquired gene mutations in donor associated with predisposition to haematological malignancies could have clinical implications for the patients undergoing to allo-hsct. although the cause of donor cell-derived haematological malignancy onset seems to be multifactorial, the infusion of a scu with pre-leukemic potential in a context of residual toxicity in recipient as a result of pre-transplant chemotherapy, a post-transplant environment characterized by a decreased immune surveillance may well have played role in these cases. the study of a greater number of dcl cases by next generation sequencing could help to understand this process and to detect new mutations involved in the emergence of aml. disclosure of conflict of interest: none. the impact of donor and recipient sex in allogeneic stem cell transplantation-single center experience (cic ) y petrov , p ganeva , g arnaudov , s lozenov , y davidkova , v stoeva , i tonev , m guenova and g mihaylov national hospital for active treatment of hematological diseases allogeneic hematopoietic stem cell transplantation (hsct) has been one of the most effective therapeutic modalities for patients with hematological malignancies and bone marrow failure syndromes. optimal donor selection is one of the key factors to enhance the success rate of this procedure. we [p ] s retrospectively investigated whether and how donor-recipient sex affects transplantation outcomes of patients transplanted between and in our center. the median age of the patients was years (range: - ). thirty-nine of the patients ( %) received a pbsc from a hla-identical sibling, and patients ( . %) received pbsc from matched unrelated donor. forty-six percent were male recipients with male donors (m-m), . % were female recipients with male donors (m-f), . % male recipients with female donors (f-m), and . % female recipients with female donors (f-f). we performed a crosstab analysis and χ tests to observe whether the donor sex affects our study population. patients with male donor had superior overall survival and progression-free survival compared to those with female donor ( . % vs . % p = . for os, and . % vs . % p = . for pfs; cramer`s v = . ). we further investigated how the disparity of the donor in the four groups (m-m, m-f, f-m and f-f) affects the os, pfs and nrm. the f-m group had a worse overall and progression-free survival comparing the other groups ( % -year os and % pfs; p o . ).this group had % relative increase in the non-relapse mortality compared with m-m group (p = . ). for m-m group there was a % relative increase in the subdistribution hazard of nrm compared with m-f group (p = . ). the f-f group and m-f group had similar subdistribution hazard of nrm ( % vs % p = . ). the incidence of acute gvhd and chronic gvhd for the groups was: % and % (m-m), % and % (m-f), % and % (f-m), % and % for the (f-f) group. the appearance of either acute or chronic gvhd did not show statistical significance regarding the os and pfs in the groups (p = . ). we examined the effect of donor-recipient sex incompatibility on the outcome of hsct in out center. our results showed inferior os and pfs for f-m group and a higher incidence of nrm compared with other groups. these effects might be associated with allogeneic immune responses against h-y antigens. key words: stem cell transplantation, donor sex, recipient sex, overall and progression-free survival [p ] disclosure of conflict of interest: none. from to , % of the patients affected by hematological malignancy searching for an unrelated donor through the italian registry successfully identified a suitable donor. this proportion increases up to % when searching for a cord blood unit was considered, corresponding to total transplant efficiency of %. from april , the rome transplant network adopted a unique policy for the identification of a potential alternative donor, following a hierarchical selection that considered as first choice a volunteer unrelated donor, secondly a cord blood unit and last a haploidentical related donor. before starting the unrelated donor search, a preliminary query through the bone marrow donor worldwide database was performed for all the patients referred to the rome transplant network. based on the low resolution hla typing (a, b and drb ) it was possible to arbitrary assign a good or poor score that might predict the identification of a full matched ( / a, b, c and drb ) donor. therefore, aims of the present study were to assess the utility of the preliminary query and the impact of the use of high resolution hla typing since the starting of donor search on the timing for the unrelated donor identification. moreover, the final aim was of comparing donor identification and transplant efficiency between the national registry, that considers only the unrelated donor and the rome transplant network, whose policy includes also haploidentical donor as third choice in the donor search process. at rome transplant network % out of adult patients met criteria of a good preliminary query corresponding to a matched unrelated donor identification in % of cases vs only . % for patients with poor preliminary query. our policy led to % and %, respectively, of alternative donor identification and transplant efficiency, significantly higher than the corresponding data of % (p = . ) and % (p o . ) reported by the national registry. moreover, the median duration of search process for mud identification has been significantly reduced by the use of hr hla typing patient at the start of the formal search activation from (range: - ) to (range: - ) days at ibmdr (po . ) and from to days ( - ) at rtn (po . ). in conclusion, the preliminary query represents a useful tool to address the search towards the best donor choice and to perform transplant in adequate time. moreover, the timing of donor identification has been significantly reduced with the use of high resolution typing at the start of donor search. a search and selection donor policy should be basically established and should include the haploidentical donor to improve the transplant efficiency. disclosure of conflict of interest: none. the long term prognosis of elderly acute myeloid leukemia (aml) patients remains poor. advances in the uses of alternative donors and reduced intensity conditioning regimens have extended the use of allogeneic hematopoietic stem cell transplantation (hsct) to a wider number of patients. however, few studies have reported data on the efficacy of hsct from alternative donors in elderly aml patients. we retrospectively analyzed the transplantation outcome in consecutive elderly aml patients aged years who received hsct ( hsct ( - at the catholic blood and marrow transplantation center. donor types were autologous (n = ) or hla matched related (mrd, n = ), unrelated (mud, n = ), or haploidentical (n = ). for graft-versus-host disease (gvhd) prophylaxis, methotrexate and cyclosporine (mrd) or tacrolimus (mud/haploidentical donor) were used. mud and haploidentical donors were given antithymocyte globulin. the median age was years, with patients ( %) years. intermediate-or adverse cytogenetic risk was observed in % of patients. with a median follow-up of . months, overall survival (os) and disease-free survival (dfs) at years after transplantation were % and % for autologous, % and % for mrd, % and % for mud, and % and % for haploidentical hsct, respectively. the -year relapse was significantly higher for autologous hsct compared to allogeneic hsct ( % vs %, p = . ), while it was similar among allogeneic donors: mrd, %; mud, %; haploidentical, % (p = . ). the -year non-relapse mortality (nrm) for mud ( %) or haploidentical donor ( %) hsct was comparable to that of autologous hsct ( %), while it was relatively higher for mrd hsct ( %, p = . ). of the patients receiving allogeneic hsct, the -year cumulative incidence of moderate to severe chronic gvhd was significantly increased for mrd ( %) compared to alternative donor hsct ( %, p = . ). in multivariate analysis, patient age (hr . , % ci . - . , p = . ) and donor type (hr . % ci . - . , p = . for mud; hr . , % ci . - . , p = . for mrd compared to haploidentical donor) were significantly associated with the cumulative incidence of moderate to severe chronic gvhd, while female-to-male hsct showed a borderline significance (hr . , % ci . - . , p = . ). incidence of acute gvhd was similar according to donor type. in the multivariate analysis for nrm, patient age (hr . , % ci . - . , p = . ), mrd (hr . , % ci . - . , p = . ), and hematopoietic cell transplantation-comorbidity index high risk (hr . , % ci . - . , p = . ) were significantly associated. in conclusion, our results showed significantly higher relapse rate for elderly aml patients receiving autologous hsct compared to allogeneic hsct, responsible for the lower survival rate in autologous hsct. we observed that nrm rate for mud and haploidentical donors for elderly aml patients were lower than expected and similar to autologous hsct. relatively higher incidence of nrm for mrd hsct seemed responsible for the low long term dfs. these results suggest a need for strengthening of gvhd prophylaxis in mrd hsct for elderly aml patients. our results suggest a potential role of alternative donor hsct to improve long term survival rates in elderly patients with aml. disclosure of conflict of interest: none. for patients with saa, transplantation from an unrelated donor (ud) is usually considered after failure of at least one course of immunosuppression. this strategy is based on a relatively high risk of complications for ud transplant recipients, such as graft rejection, graft-versus-host disease (gvhd) and infections. however, the outcome of unrelated donor transplants has significantly improved in recent years, due to better donor selection, conditioning regimen optimization and better supportive care. the authors describe results from patients with saa who receive unrelated allogeneic transplants in a single reference institution from to . data was retrieved from the center databasis and there were females and males. median age was years old . median total number of cells infused was . × /kg. % of the patients have received more than transfusions previously. conditioning regimen were: cy + tbi ± atg in ( %) patients, bu mg/kg+ cy + atg in ( %), and fludarabine + cy+atg in ( %), fludarabine, cy+tbi in ( %) patients. stem cell source was marrow in %, cord blood in % and peripheral blood in % of patients. transplants were full matched in ( %) patients, had one mismatch (out of ) in ( %) and mismatches in ( %) patients. engraftment was complete as evaluated by donor chimerism at day and post transplant in patients ( %), partial in ( %) and graft failure was observed in ( %) patients. acute gvhd grade ii-iv was seen in patients ( %) and nih moderate to severe chronic gvhd was seen in ( %) patients. median overall survival was days ( - ) and estimated years overall survival was %. risk factors for survival identified were: hla mismatch and stem cell sources other than marrow. unrelated transplants are a feasible salvage therapy for patients with saa refractory to immunosuppression, being hla compatibility and marrow stem cell source factors with a positive impact on survival. disclosure of conflict of interest: none. use of haploidentical stem cell transplantation continues to increase, the european society for blood and marrow transplant activity survey report jr passweg , h baldomero , p bader c bonini , rf duarte , c dufour , , a gennery , n kröger , j kuball , f lanza , s montoto , a nagler , ja snowden , j styczynski and m mohty for the european society for blood and marrow transplantation (ebmt) hematopoietic stem cell transplantation (hsct) is an established procedure for many acquired and congenital disorders of the hematopoietic system, including disorders of the immune system, and as enzyme replacement in metabolic disorders. the annual activity survey of the ebmt describes the status of hsct in europe and affiliated countries and has become an instrument used to observe trends and to monitor changes in technology use. teams were invited to report their transplant activity for by indication, stem cell source and donor type using a single paged survey. a record number of ' hsct in ' patients ( ' allogeneic ( %), ' autologous ( %)) were reported by centers in countries in . trends include continued growth in transplant activity during the period and , with the highest percentage increase seen in middle income countries (allo %, auto %), and the lowest in very high income countries (allo %, auto %), for both allogeneic and autologous hsct. in contrast the absolute growth is highest in the very high income countries (growth allo rates transplants per × inhabitants, auto rates for very high income countries; allo rates , auto rates for middle income). main indications for hsct were myeloid malignancies ' ( %; % allogeneic); lymphoid malignancies ' ( %; % allogeneic); solid tumors; ' ( %; % allogeneic); and non-malignant disorders; ' ( %; % allogeneic). remarkable is a decreasing use of allogeneic hsct in cll from patients in to in and is most likely due to the development of potentially very effective cll drugs. use of haploidentical donors for allogeneic hsct continues to increase ' in ; a % increase since . the highest growth is seen in myeloid malignancies ' , with lymphoid malignancies , nonmalignant disorders and others. in aml, haploidentical hsct increases similarly for patients with both advanced disease and those in cr . both marrow and peripheral blood is used as stem cell source for haploidentical hsct with higher numbers reported for the latter. this year's activity survey shows continued increase in the use of haploidentical hsct across europe within the main indication groups and cell source. it reflects in a timely manner current trends in stem cell transplantation and is an essential tool for health care planning and health policy makers. human bone marrow mesenchymal stromal cells derived exosomes (hbmmdes) are small membrane vesicles secreted from mesenchymal stromal cells that may serve as a vehicle for protein, mrna and microrna (mirna) transfer to distant cells; affecting gene expression, proliferation, and differentiation of the recipient cells. therefore, mdes may possess some of the immunoregulatory properties of their parental cells. in the present study we aim to explore the immunomodulatory function of mdes and understand the molecular mechanisms enabling it. for this purpose, we co-cultured hbmmdes with activated human lymphocytes. using ultracentrifugation, hbmmdes were isolated from expanded human bone marrow derived mesenchymal stromal cells (hbmmscs). using em and zeta sizer, particles were shown to be in the range of - nm. pha activated human peripheral blood lymphocytes (pbls), r- /il activated b cells and anti cd /cd activated t cells were co cultured with purified mdes. cell proliferation was tested using thymidine incorporation assay. we found that exosomes derived from × to × mscs exhibited a dose-dependent inhibition of lymphocyte proliferation. exosomes derived from × mesenchymal stromal cells co cultured with pha activated pbls, activated b cells and activated t cells showed proliferation inhibition of %( p ⩽ . ), . % (p ⩽ . ) and . % (p ⩽ . ), respectively. in order to understand the molecular mechanism behind the immunomodulatory effect of mdes, we have profiled mde's mir content using illumina hiseq platform and we are currently profiling co cultured activated lymphocytes mrna content using next-generation sequencing system, illumina. preliminary results demonstrate some higher abundance of specific mscs derived mirs in the mdes. hbmmscs have been shown to serve as immune modulators in patients with acute and chronic graft versus host (gvhd). in the future, mdes may provide an alternative therapy for gvhd. compared with bmmscs, mdes are more stable, have no risk of aneuploidity or ectopic proliferation and have less probability of immune rejection. additional studies are needed to explore the applicability of mdes to serve as modulators of the immune response. disclosure of conflict of interest: none. graft-versus-host disease (gvhd) is the major complication after allogenic haematopoietic stem-cell transplantation s (hsct). extra virgin olive oil (evoo) is a source of phenolic compounds such as glycoside oleuropein, hydroxytyrosol and tyrosol. olive oil polyphenols have shown antioxidant, immunomodulatory, antiproliferative, anti-apoptotic and antiinflammatory properties that might be useful in the prophylaxis and treatment of gvhd. polyphenolic extract (pe) of evoo was obtained by the method described by vazquez roncero et al. with some modifications. briefly, fifty grams of evoo (oleoestepa, seville, spain) was extracted with methanol/water ( : , vol/vol, ml ). the mixture of evoo, methanol and water was decanted and the methanolic extract was concentrated and lyophilized. then, the effect of pe in cell viability and activation of t lymphocytes from healthy donor's buffy coats either resting or activated with anticd plus anticd was analyzed by flow cytometry after staining with aad, anexin-v and cd . proliferation assays were performed with pkh and the quantification of il- , il- , il- , il- , tnf-α and ifn-γ cytokines in cell culture supernants with bd cytometric bead array (cba). signaling pathways were analyzed by western blot. finally, in a mouse model of acute gvhd (c bl/ in balb/c), mice were randomized into two experimental diet groups: standard diet ( s harlan laboratories) and standard diet ( s harlan laboratories) supplemented with ppm of pe obtained of evoo. the severity of gvhd was assessed by a scoring system described by cooke et al. that incorporates five clinical parameters: weight loss, posture (hunching), activity, fur texture, and skin integrity. pe did not affect t cell viability. by contrast, pe decreased t-cell activation and proliferation of t-lymphocytes stimulated with anticd plus anticd . in addition, there was a decreased production of th (ifnγ, il- and tnf) and th cytokines (il- , il- and il- ) in the presence of pe. regarding the signaling pathways analyzed, pe inhibited phosphorylation of akt and nuclear translocation of nfkb in activated t cells. in the mouse model of acute gvhd, animals which received the pe supplemented diet had an increased survival as compared to mice receiving a standard diet. also, gvhd incidence was significantly lower among mice receiving the pe supplemented diet as assessed by both the presence of gvhd signs as well as pathological examination. polyphenols obtained from evoo are an important immunomodulatory agent capable to reduce the proliferation and activation of activated t cells and the production of proinflammatory cytokines. in a mouse model of acute gvhd, pe supplemented diet reduced the incidence and severity of the disease and increased the survival of mice. disclosure of conflict of interest: none. graft-versus-host disease (gvhd) is a leading cause of postallogeneic haematopoietic stem cell transplantation (hsct) morbidity and mortality ( ) . extracorporeal photopheresis (ecp) is an alternative therapeutic strategy that appears to act in an immunomodulatory fashion, potentially involving regulatory t lymphocytes and dendritic cells in patients who are refractory to steroids. dendritic cells (dcs) are the most important antigen-presenting cells, playing a pivotal role in t-cell function and in the link between innate and adaptive immunity. moreover, dcs are also critical mediators of immune tolerance and energy. they can be divided into two major subsets, plasmacytoid dcs (pdcs) and myeloid dcs (mdcs) which have distinct functions. pdcs play a pivotal role in peripheral tolerance through generation of regulatory t (treg). on the other side mdcs promote, as well as pdcs, th and th /tr responses ( - ). our study was performed to understand the mechanism of action involved in immunomodulatory effect of ecp. as the modulation of dcs and tregs number and function ( , ) may be a central mechanism of ecp in maintaining self-tolerance, down-regulating immune responses, and limiting inflammation ( ). eight patients affected by gvhd were included in this pilot study. in ecp apheresed mononuclear cells are exposed to methoxypsoralen and uva radiation. after this photoactivation, which induces dna damage and apoptosis, the cells exposed are re-infused into the patient inducing an immunomodulatory effect. all patients or their legal guardians gave their consent for this study. a sample of peripheral blood (pb) (basal condition), a sample of apheresis pre-uva photoactivation (pre-pa) and a sample of photoactivated apheresis (pa) were collected at the first day of ecp and every week for the first month of treatment. circulating dcs, mdcs (cd / -cd +cd +), pdcs (cd / -cd +cd +) and tregs (cd +cd +foxp +) were directly enumerated and phenotypically characterized. the assays were performed at day+ ,+ , + ,+ ,+ data are expressed as mean ± s.d. of absolute number of cells/μl. at day + there were no differences in the absolute number of both mdcs and pdcs between pre-pa and pa. consequently there were no differences between pb and pa. from day + till + we observed an increase of these two cellular populations at every date of treatment. comparing the basal pb of day + vs day + we observed an increment of % and %, respectively for mdcs and pdcs (mdc from cell/μl to cell/μl; pdc from cell/μl to cell/μl). comparing the basal pb of day + vs day + we observed an increment of % of tregs (from cell/μl to cell/μl) while we observed a median increment of % calculated between pre-pa and pa of each day of treatment from day to day + . no firm conclusions can be drawn from a clinical point of view, however a biological effect has certainly highlighted. in particular no substantial differences in basal pb mdc or pdc emerged during the first month of treatment while a significant increase of mdc and pdc can be observed since day + following uva photoactivation. regarding tregs we observed an increment of % of tregs between pb from day + to day+ and a median increment of % calculated between pre-pa and pa of each day of treatment. disclosure of conflict of interest: none. [p ] p impact of th cells on xenogeneic graft-versus-host disease l delens, s servais , g ehx , l vrancken , g fransolet , c gregoire , m hannon , s dubois , c daulne , f baron and y beguin giga i : hematology, university of liege acute graft-versus-host disease (gvhd) is a severe complication of allogeneic hematopoietic stem cell transplantation. its pathophysiology is complex and not yet fully understood. in particular, the impact of th cells on murine acute gvhd has yielded conflicting results, while demonstration of increased levels of th cells at the site of acute gvhd provided only indirect evidence of their involvement in humans. here, we assessed the potential implication of th cells in a humanized mouse model of xenogeneic gvhd (x-gvhd). methods: x-gvhd was induced by infusing human peripheral blood mononuclear cells (pbmcs) into nod-scid il- rγnull (nsg) mice given . gy total body irradiation day prior transplantation. th cells were generated by culturing naive cd + t cells with anti-cd /anti-cd coated beads under th -skewing cytokines (tgf-β , il -β, il- , il- , il- , neutralizing anti-il- and anti-ifnγ antibodies) in hypernatremic conditions (nacl mm). results: after days of culture, a median of . % of il- a+ cells was obtained. we confirmed the expression of il- a, rorc and il- r by these cells by rt-qpcr. we next assessed the co-injection of human pbmcs ( . ) with in vitro differentiated cells under th skewing conditions ( × ) (co-injection group, n = ), in comparison with the injection of pbmcs alone ( × cells, pbmcs group, n = ). we observed higher x-gvhd score (p %) of cells expressing both il- a+ and ifnγ+ cells (th / th -like phenotype) among cd + il- a+ cells while coinjected mice had higher blood concentration of il- a (p = . ) than pbmc mice. these results demonstrate that addition of th cells worsened x-gvhd confirming their role in acute gvhd pathogenesis. disclosure of conflict of interest: none. although survival from allogeneic stem cell transplantation (hsct) has significantly improved, acute graft-versus-host disease (gvhd) remains a major cause of death. intestinal dysbiosis has been associated with acute gastrointestinal gvhd and poor outcome after hsct. we reported a correlation between microbiota (gm) composition and short chain fatty acid (scfa) production and gvhd in transplanted children. to assess how the metabolic pathways of gm change during transplantation and identify modulators of immune response, we perform first longitudinal metagenomic analysis in children undergoing hsct. patients (pts) ( male; mean age: y) with hematologic malignancies ( all, aml), who received busulphan-based myeloablative conditioning and t-cell replete bone marrow graft were enrolled. pts were prospectively enrolled in a protocol with at least specimens fecal samples collected: one before and two after hsct, in order to build a proper trajectory. gvhd prophylaxis was cyclosporine for pts receiving a matched related donor and cyclosporine, short-term mtx and atg for pts receiving a matched unrelated donor. non-gvhd and gvhd patients had similar exposures to antibiotics during the stool collection. of these pts, % developed gvhd within the first days. we applied shotgun metagenome sequencing to total fecal dna from samples collected. functionalities were assigned by reads mapping at different levels of the kegg database. relative abundance was calculated and statistical analysis was performed. according to our findings, core functional profiles were overall conserved through the time-points in all patients ( figure a ), in contrast to the phylogenetic profiles behavior, this finding confirming the overall redundancy of gut microbiome core functionalities. analyzing the single metabolic pathways in subjects who developed gvhd, we found in the pre-hsct period a higher relative abundance of nucleobasis (purine and pyrimidine) metabolism (p o . ) and branched-chain amino acids biosynthesis (p o . ). functions related to the production of branched-chain amino acids are involved in the biosynthesis of the cell wall of gram-negative bacteria, microorganisms including subgroups with well know opportunistic pro-inflammatory. in addition, post-hsct samples of gvhd patients showed a lower abundance of genes involved in polysaccharides metabolism, as glycan biosynthesis and glycosaminoglycan degradation (p o . ) ( figure b ). glycosaminoglycan degradation activity gets bacteria able to survive during extreme situations, as fasting using mucus polysaccharides as energy source, contributing to maintain a mutualistic composition of gm and scfa production by the saccharolytic functions of the endogenous mucus polysaccharides. this study detects functional peculiarities in the gm of non-gvhd pts. the gut metagenome configuration of non-gvhd patients is structured to derive scfa after hsct. the production of these metabolites promotes peripheral regulatory t-cell generation , potentially explaining the protective role of gm from gvhd. although intestinal epithelial cells (iecs) are crucial regulators of barrier function and immune homeostasis, they also facilitate inflammation in exaggerate responses to proinflammatory mediators by pretransplant conditioning regimen, which plays a critical role in amplifying graft-versus-host disease (gvhd). thus inhibition of the converting to pathogenic iecs by conditioning may represent a novel approach to inhibit gvhd. aryl hydrocarbon receptor (ahr) is the ligandactivated transcription factor which has the ability to mediate the biochemical, metabolic, and toxic effects of environmental chemicals. recently, it has been demonstrated that ahr is an important regulator of cell development, differentiation, and function of both innate and adaptive immune cells. the ability of ahr is induced by respond to endogenous ligands generated from the host cell, diet, and microbiota. here, we investigated the regulatory role of ahr in iecs under inflammatory responses and its therapeutic activity for modulation of gvhd. ahr and cyp a expression in mouse iecs were determined by real-time pcr. mouse iecs were pretreated with endogenous ahr ligands l-kynurenine (l-kyn, mm) or pbs for h and then stimulated with lps or il- b for h. cytokine levels were measured using the mouse flex-set cytokine bead array or real-time pcr. b d f (h- b/d) recipients were administrated l-kyn daily by i.p. injection for days. then the recipients were lethally irradiated and transplanted with × tcd-bm plus × t cells from b (h- b) donor. mice were monitored every other day for survival and clinical score. colons were collected and stained with hematoxylin and eosin (h&e) for histopathological scoring. we found that ahr was constitutively expressed in the mouse iecs. cyp a (an ahr target gene) was significantly increased by treatment of l-kyn under un-stimulatory condition. we further observed that l-kyn completely abrogated il- β-mediated il- or lps-mediated tnf-a expression in iecs. administration of bdf recipient mice with l-kyn before transplantation significantly reduced the lethality and severity of gvhd. histopathology clearly revealed that treatment of l-kyn inhibited intestinal gvhd. our results demonstrate that ) ahr is constitutively expressed in iecs, ) treatment of endogenous ligand l-kyn induce ahr activation in the steady status, ) ahr activation blocks conversation of the epithelial cells into pathogenic cell type, and ) pre-administration of ahr ligand reduces gvhd. our study suggests that activation of ahr pathway in iecs before allogeneic hematopoietic stem cell transplantation (hsct) is a possible strategy to reduce intestinal gvhd. disclosure of conflict of interest: none. [p ] s relating with acute graft-versus-host disease (gvhd) after allogeneic hematopoietic stem cell transplantation (allo-hsct), protecting endothelial cells (ecs) from damage may be a potent prophylaxis and therapeutic strategy of acute gvhd (agvhd). conventional agvhd therapies may cause many adverse side effects because of their multiple targets. therefore, we explored the therapeutic efficacy of simvastatin, a lipid-lowering drug, which has been demonstrated endothelial protection. our previous clinical observation has found patients with agvhd had lower angiopoietin- (ang- ) level at day but higher ang- level at day than those without agvhd. in this study, we explored changes in ang- and ang- expression in an agvhd mouse model and determined whether simvastatin prevents gvhd through regulating ang- and ang- expression. we preincubated ea.hy ecs with simvastatin ( mmol/l) h before stimulated with tnf-a, then ang- and ang- concentration in the cell supernatant was measured by elisa. ang- and ang- mrna and protein level of treated and untreated cells were examined simultaneously. in vitro simvastatin increased ang- production and release but conversely inhibited ang- release from ea.hy ecs. donor mice spleen cells were injected along with bone marrow cells into recipient mice after lethal irradiation to induce agvhd. simvastatin was administered orally once daily to mice ( mg/kg) for days after allo-hsct and started − day after allo-hsct. then mice survival time was monitored and organ damage was evaluated. the plasma level of ang- and ang- was measured by elisa, expressions of ang- and ang- in aortic endothelium were assessed by immunohistochemistry. simvastatin improved the survival and attenuated the histopathological gvhd grades of agvhd mice. plasma levels of ang- were significantly decreased, while plasma levels of ang- obviously increased in agvhd mice after transplantation. simvastatin reduced plasma levels of ang- , elevated the plasma levels of ang- as well as the aortic endothelial levels of ang- and ang- . in summary, simvastatin represents a novel approach to combat gvhd by increasing ang- production while suppressing ang- release to stabilize endothelial cells. there is a growing evidence of safety and efficacy of posttransplantation cyclophosphamide (ptcy) in stem cell transplantations (sct) from different donors and graft sources. still the optimal combination of immunosuppressive agents with ptcy should be elucidated for different types of scts. we report the -year update of the prospective nct single-center trial that evaluated risk-adapted graft-versushost disease (gvhd) prophylaxis with ptcy in related, unrelated and haploidentical scts. adult patients (median age y.o., range: - ) with hematologic malignancies, including aml ( . %), all ( . %), cml ( . %), mds ( %), and lymphomas ( . %), were enrolled in the study. % of patients were classified as salvage. % received the graft from matched related (mrd), % from matched/mismatched unrelated (mud/mmud), and % from haploidedntical (haplo) donor. % received bone marrow graft (bm) and %peripheral blood stem cell (pbsc) graft. . % had myeloablative conditioning and . %-reduced-intensity conditioning. gvhd prophylaxis for matched bm grafts consisted of single-agent ptcy mg/kg days+ ,+ , for matched pbsc graft-ptcy+ tacrolimus+ mycophenolate mofetil (mmf) mg/kg days - , and for any mismatched graft-ptcy+ tacrolimus+ mmf mg/kg days - . median follow-up was months (range: - ). grade ii-iv ( % vs % vs %, p = . ) and grade iii-iv acute gvhd ( % vs % vs %, [p ] p = . ) were not different in mrd, mud/mmud and haplo groups, respectively. moderate and severe chronic gvhd was infrequent in all groups with slightly lower incidence after mud/mmud graft: and % vs % vs %, p = . . nonrelapse mortality (nrm) was not different after mrd, mud/ mmud and haplo sct ( % vs % vs %, respectively, p = . ), while relapse incidence was higher after mrd and haplo grafts: ( % vs % vs %, p = . ). -year overall survival (os), event-free-survival (efs), and gvhd-relapse free survival (gfrs) were % vs % vs % (p = . ); % vs % vs % (p = . ); % vs % vs % (p = . ) for mrd, mud/mmud and haplo groups, respectively. in the multivariate analysis only disease risk index (hr . %ci . - . , p = . ), severe sepsis (hr . %ci . - . , p = . ) and chronic gvhd (hr . %ci . - . , p = . ) were predictive for efs, while type of donor was not a significant factor (hr . %ci . - . , p = . ) (figure ). the incidences of complications were: hemorrhagic cystitis- %, sepsis- %, severe sepsis- %, invasive mycosis- %, cmv reactivation- %, veno-occlusive disease- . %, transplant-associated microangiopathy- . %, grade - liver toxicity- %, grade - kidney toxicity- %. more than one third of patients experienced poor graft function during days after sct, and in % of them cmv, hhv and bk virus reactivations were identified as the cause. the reported risk adapted strategy alleviates the risk of gvhd and nrm after mmud and haplo grafts. the observed differences in the relapse incidence, os and efs were predominantly due to unbalanced disease risks in the groups. the relapse of underlying malignancy with this prophylaxis still significantly influences the outcome. substantial number of patients experience poor graft function, which doesn't translate into nrm. disclosure of conflict of interest: none. a high migratory capacity of donor t-cells in response to the lymph node homing receptor ccr increases the incidence and severity of graft-versus-host disease vg garcía de soria , ip sainz , e jiménez , a arriero , c fernández-arandojo , c cuesta , b colom , a marcos , a rosendo and cecilia muñoz calleja department of hematology hospital universitario de la princesa and department of inmunology. hospital universitario de la princesa graft-versus-host disease (gvhd) pathogenesis involves migration of the donor t-cells into the secondary lymphoid organs (slo) in the recipient, which is steered by two homing molecules: cd l and ccr . therefore we investigated whether the migratory capacity of donor t-cells is associated with gvhd. this single center prospective study included donor-recipient pairs. in vitro chemotaxis assays of the lymphocytes of the apheresis product were performed in parallel with the analysis of cd l and ccr by flow cytometry. the potential of activation of ccr + t-cells was assessed through ex vivo activation assays with peripheral blood monuclear cells (pbmc) from healthy donors using anti-cd and anti-cd mabs. the migratory index to the ccr ligands, ccl and ccl , was higher in t-cells from donors whose recipients will develop gvhd. these data indicated that the migratory capacity of the donor t-cells is clearly related to the development of gvhd. this prompted us to study the relationship between gvhd and the expression of two of the most relevant molecules in the trafficking of lymphocytes towards slo, cd l and ccr ,as a subrogate index of the migratory potential of t-cells. consequently, we quantified the numbers of cd l+ and ccr + t-cells in the graft. the initial transversal analysis of our data revealed that the percentage of cd l+ lymphocytes in the apheresis product was very low compared to healthy lymphocytes. the analysis also confirmed that cd l undergoes plasma membrane shedding after g-csf mobilization thus making it a non-valid biomarker. the analysis of ccr molecule revealed that the acute gvhd group received higher percentage of cd +ccr + t-cells, whereas chronic gvhd patients were transplanted with higher percentage of cd +ccr + t-cells compared to the non gvhd group. these results were confirmed when patients were subdivided into degrees of severity. a multivariate analysis was performed to investigate the real value of ccr to predict the development and severity of gvhd, and confirmed that ccr expression is a risk factor for the development of gvhd. thus, the percentage of ccr +cd + t-cells increases the probability of developing acute gvhd (or = . , c.i ( %) = . - . , p = . ) and suffering a higher degree (or = . , c.i ( %) = . - . , p = . ). similarly, the or of the percentage of ccr +cd + t-cells was . (c.i ( %) = . - . , p = . ) and . (c.i ( %) = . - . , p = . ) for the development of chronic gvhd and its degrees, respectively. finally, to study the potential of activation of ccr + t-cells, we carried out ex vivo activation assays with pbmc from healthy donors using anti-cd and anti-cd mabs and the expression of cd l on cd + t-cells and of cd on cd + t-cells as markers of activation, demonstrating that ccr + t-cells exhibited higher potential of activation than ccr -t-cells. to our knowledge this is the first analysis of the influence of the migratory capacity of the donor t-cells on clinical outcome following allogeneic hsct. our data show that ccr could be considered a subrogate biomarker of the migratory capacity of the donor lymphocytes for predicting the risk of suffering gvhd. based on the previous findings, we propose that the selective depletion of ccr expressing cells could be an effective preventive therapy for gvhd. disclosure of conflict of interest: none. previously published p a single center research for outcome in patients receiving imatinib for steroid-refractory chronic gvhd after allogeneic stem cell transplantation l ni, y luo, y tan, y hu, y zhao, j shi and h huang despite of major progress in allogeneic stem cell transplantation over the last decades, steroid-refractory chronic graftversus-host disease (sr-cgvhd) remains a leading cause of late morbidity and mortality. pre-clinical evidence confirms cgvhd has antibodies activating the platelet-derived growth factor receptor (pdgf-r) pathway. since this pathway can be inhibited by imatinib, we performed a study including patients with sr-cgvhd given imatinib at a dose of mg per day. all patients with a median age of years (range: - ) underwent allogeneic hematopoietic stem cell transplantation in our single center between and , and chronic gvhd occurred at a median time of months (range: - ) after transplantation. patients had active cgvhd with measurable involvement of skin, lung or other districts and had previously failed in first-line immunosuppressive therapy. the major organs involved were lung (n = ), skin (n = ) and mouth (n = ), including cases involving both lung and skin, cases involving or more organs. according to the national institutes of health (nih) criteria and nih global severity, patients were evaluated as severe cgvhd, and the other three were moderate. meanwhile, the nih working group had updated its recommendations for overall responses, consisting of complete remission (cr), partial remission (pr), and lack of response (unchanged, mixed response, progression). cr was defined as resolution of all manifestations in each organ or site, and pr was defined as improvement in at least organ or site without progression in any other organ. after months treatment, patients receiving sufficient dose of imatinib revealed overall response rate (orr) at . %, and orr remained unchanged at months assessment, but with cr rate increased to . %. two patients couldn't meet the response of cr or pr were considered as a lack of response, including one evaluated as unchanged and one mixed response because of pr in lung accompanied by progression in eyes. with a median follow-up of months, patients were alive, with a year estimated overall survival was . %. patients eventually died of pneumonia. except patient discontinued imatinib because of grade toxicity as gastrointestinal discomfort at the first month, no one had imatinib-related grade to toxicity. this study suggests that imatinib is a promising and better tolerated treatment for patients with sr-cgvhd. disclosure of conflict of interest: none. acute graft-versus-host-disease (agvhd) is a major complication after allogenic hematopoietic transplantation (allo-sct). in recent years, a number of tissue-specific proteins have been described as biomarkers that could contribute to anticipate and/or diagnose this complication earlier and more accurately. reg α (regenerating-islet-derived- -alpha) has been directly related to gastrointestinal (gi) agvhd. our objective was to analyze plasma levels of reg α at days + and + in patients who underwent unmanipulated haploidentical transplantation with reduced conditioning regimen (haplo-ric), and to correlate the results with the development of agvhd. we retrospectively analyzed consecutive patients ( - ) who underwent haplo-ric with post-transplant cyclophosfamide (days + , + ), mmf and csa as gvhd prophylaxis. seven cases were excluded due to early death (before day + ) and cases due lack plasma sample. characteristics of the patients included in the analysis are described in table . reg α detection was performed by elisa (mbl international corp, woburn, ma) according to manufacturer's instructions on μl of plasma obtained at day + and + . the association of the incidence of agvhd with known clinical variables and plasma reg a levels were performed by cox regression and mann-whitney u-test, respectively. the determination of the best cut-off of reg α levels to stratify patients with gi agvhd was performed with roc curves. the stadistical program used was r v . . . the cumulative incidence of grade ii-iv and grade iii-iv agvhd was % and %, respectively. characteristics of agvhd are shown in table . no association was found between agvhd and usual clinical variables (stem cells source, age, sex, conditioning regimen, donor/recipient sex and number of infused cd + cells), and with plasma reg α levels at day + .plasma reg α levels at day + were higher in patients who devolved gi agvhd compared to patients who did not showed gi agvhd (median [p ] s and range: ( - ) vs ( - ) pg/ml, p = . , figure ).the best cut-off selected on day + was pg/ml (s %, e %). patients with levels higher than pg/ml at day + had a significantly higher incidence of gi agvhd grade ii-iv (hr . , p = . , figure ). plasma levels of reg α at day + after haplo-ric correlated with the occurrence of gi agvhd grade ii-iv. therefore, plasma levels of reg α could be use for the prediction and/or diagnosis of gi agvhd. disclosure of conflict of interest: none. anti-fibrotic treatment with pirfenidone in patients with gvhd-associated bronchiolitis obliterans syndrome ke hostettler, s gerull, g nair , j passweg , m tamm and j halter hematology, university hospital basel, switzerland and pneumology, university hospital basel, switzerland prognosis of lung gvhd remains poor due to progressive decrease of lung function and repeated infections. pirfenidone exhibits anti-fibrotic effects and has been shown to reduce disease progression in patients with idiopathic pulmonary fibrosis. five patients with established bos (nih criteria ) and stable or deteriorating lung function under standard immunosuppressive treatment without active infection were treated with pirfenidone ( mg/d) in addition to their current therapy. clinical assessments and pulmonary function tests were performed every three months. five patients ( m, f), median age y (range: - y) that were diagnosed with bos at a median time of . months post-transplant started pirfenidone at a median time of months ( - ) after diagnosis of bos. two patients are currently still under treatment after and days. two patients had to stop treatment due to financial reasons after and days of therapy. one patient never reached more than % of the planned dose due to gastro-intestinal symptoms and was excluded from further analysis. at the start of treatment median fev was . l ( . - . ); . % predicted (range: - %) and median fvc . l ( . - . ); - % predicted. median fev trajectory was − . % predicted/ month during median months before start of pirfenidone (median − ml/month) and + . % predicted/month (+ . ml/month) during treatment with pirfenidon. the treatment was well tolerated except in one patient with gastrointestinal complaints, no phototoxic reactions or serious drugrelated adverse events occurred. in our small number of patients pirfenidone was rather well tolerated and generally safe. the observed, albeit small trend in change of fev trajectory justifies further studies of anti-fibrotic therapy as a new therapeutic option in bos after allogeneic hsct. disclosure of conflict of interest: none. anti-thymocyte globulin has been widely used for the prevention of severe graft versus host disease in patients undergoing hsct from unrelated donor. however, the optimal dose remains to be defined. in samsung medical center (seoul, korea), institutional strategy for the atg use has been changed since april , and we hypothesized that the incidence of chronic gvhd may differ by atg strategy. before april , atg . mg/kg was routinely used in allogeneic hsct from unrelated donor, whereas, the dose of atg was escalated to . mg/kg since april . in this study, a total of patients who underwent allogeneic hsct from matched or unmatched unrelated donor between jan and dec were retrospectively analyzed. peripheral blood was used as the source of stem cells in all patients. after a median follow up of . months, the cumulative incidence of moderate to severe chronic gvhd was . % ( % confidential interval [ci], . to . ) in the low-atg group and . % ( % ci, . to . ) in the non-atg group (p = . ). the rate of year overall survival (os) was not significantly different between the groups ( . % in low-atg group vs . % in high-atg group, p = . ), as was the rate of disease free survival (dfs) ( . % in non-atg group vs . % in atg group, p = . ) and cumulative incidence of relapse (cir) ( . % in non-atg group vs . % in atg group, p = . ). in allogeneic hsct from unrelated donor, larger atg dose ( . mg/kg) did not reduce the incidence of chronic gvhd when compared to lower atg dose ( . mg/kg). disclosure of conflict of interest: none. allogeneic hsct provides a curative chance for patients with hematological fatal disease. however, substantial risks remain for morbidity and mortality caused by disease relapse and graft-versus-host disease. in samsung medical center (seoul, korea), institutional strategy for the atg use has been changed since april , and we hypothesized that the incidence of chronic gvhd may differ by atg strategy. before april , atg was not routinely used in matched sibling donor (msd) transplantation, whereas, atg mg/kg has incorporated into hsct process in transplantation from msd thereafter. in this study, a total of patients who underwent allogeneic hsct from msd between jan and dec were retrospectively analyzed. peripheral blood was used as the source of stem cells in all patients. after a median follow up of . months, the cumulative incidence of moderate to severe chronic gvhd was . % ( % confidential interval [ci], . to . ) in the atg group and . % ( % ci, . to . ) in the non-atg group (p = . ). the rate of -year overall survival (os) was not significantly different between the groups ( . % in non-atg group vs . % in atg group, p = . ), as was the rate of disease free survival (dfs) ( . % in non-atg group vs . % in atg group, p = . ) and cumulative incidence of relapse (cir) ( . % in non-atg group vs . % in atg group, p = . ). in allogeneic hsct from msd, atg use was significantly associated with less occurrence of chronic gvhd, but not linked to increasing risk of relapse, with showing similar os and dfs between atg and non-atg group. disclosure of conflict of interest: none. long-term follow-up from the prospective randomized phase iii multicenter trial comparing a standard gvhd prophylaxis with cyclosporine a and methotrexate with or without additional pretransplant atlg (grafalon, previously atg-fresenius s) (given mg/kg/day, days − to − ) in unrelated donor hematopoietic cell transplantation after myeloablative conditioning resulted in a significant reduction of acute and chronic gvhd without compromising relapse rate and survival [ , , ] . here we report on a subsequent prospective non interventional observational study evaluating the outcome of patients receiving atlg in unrelated donor transplantation in day to day clinical practice without the selective measures of a clinical trial (german clinical trials register drks ). thirteen transplant centers included patients with haematological malignancies (median age years, iqr - years, range: - years) in early (n = , %), intermediate (n = ; %) or advanced (n = ; %) disease status receiving marrow (n = ) or pbsc (n = ) from / matched ( ; %) or mismatched ( ; %) unrelated donors (n = related) after myeloablative (n = , %) or ric (n = , %) conditioning. gvhd prophylaxis consisted of calcineurin inhibitors, mainly csa (n = , %) with mtx or mmf and atlg. different dosing regimens were allowed according to current practise of centers. median total atlg dose was mg/kg (iqr - mg/kg, range: - mg/kg). median follow-up was months (range: - months). as compared to patients in our randomized phase iii multicenter trial [ , , ] , patients in this study were older; advanced disease status, / match, pbsc transplantation were more frequent, and given median atlg dose was lower. acute and chronic gvhd, nrm, relapse risk, dfs and os at one year were similar to the results obtained in our randomized trial: incidence of°ii-iv agvhd: %, iii-iv agvhd: %; moderate/severe cgvhd: %; nrm: %; risk of relapse: %; relapse mortality: %; os: %. the experience in day to day clinical practice confirms the results shown in our randomized trial, namely the gvhd protective effect of atlg without compromising nrm or relapse rates. baseline calprotectin as a predictor for acute gastrointestinal graft versus host disease (gvhd)-a prospective study n schmidlin , a holbro , , jp halter , d heim , l infanti , , a plattner , r plattner , c rothen , a buser , , c bucher and jr passweg division of hematology, university hospital basel, switzerland; blood transfusion center, swiss red cross, basel, switzerland and rothen medical laboratories, basel, switzerland graft versus host disease (gvhd) is a major complication after allogeneic stem cell transplantation. so far there is no good validated predictor for the incidence and severity of gvhd. fecal calprotectin (cpt) is a protein in leukocytes with antibacterial properties. it has been shown to be elevated in acute gastrointestinal gvhd. additionally, cpt may be predictive for treatment response. the aim of the current prospective study was to investigate the role of baseline cpt in predicting incidence and severity of intestinal gvhd. in this prospective study conducted at the university hospital basel, switzerland, we included all adult patients undergoing hsct. the institutional review board approved the study. data were collected prospectively. cpt was measured twice before conditioning and at transplantation. fecal samples for cpt were obtained before conditioning and on the day of transplantation and assessed twice by standard elisa. between march and april a total of patients ( % males, patients with both baseline and transplant cpt values) were included. patient, disease and transplant characteristics are described in table . median age at transplant was years (range: - years). most patients had myeloid neoplasia and % received myeloablative conditioning. gvhd prophylaxis consisted mainly of cyclosporine containing regimens ( %). cpt levels ranged from to μg/g both at baseline (median: μg/g) and at transplantation (median: μg/g), with a good consistency between the two measurements performed (internal quality control). on the other hand, cpt did not correlate with c-reactive protein. the two measurements were taken in median days apart, depending on the conditioning regimen. eighty-five patients had an increase of at least μg/g between baseline and transplantation. overall ( . %) patients developed acute intestinal gvhd (grade : ; grade : ; grade : , and grade : patients, respectively). cpt both at baseline and at transplantation was not predictive for the incidence of gvhd, acute intestinal gvhd, and for acute intestinal gvhd grade - ( figure) . additionally, we did not find a significant association between cpt levels and the above mentioned endpoints for patients showing an increase of cpt of at least μg/g between baseline and transplantation. in the current prospective study, we didn't find any correlation between baseline cpt values and the incidence and severity of gvhd and intestinal gvhd. further studies identifying early markers and predictors of gvhd are urgently needed. [p ] disclosure of conflict of interest: none. calcinuerin inhibitor (ci) free graft-versus-host disease (gvhd) prophylaxis: its effects on resource utilization, renal function, and the cost of care m muilenburg , k cole, m abidi , , s williams and as al-homsi , spectrum health blood and marrow transplantation and michigan state university, college of human medicine effective gvhd prevention following allogeneic hematopoietic stem cell transplantation (ahsct) is vital to reducing transplant morbidity and mortality and improving overall outcomes. several strategies are currently utilized for gvhd prophylaxis including mtx, mmf, cis, post-transplant cyclophosphamide (cy), and proteasome inhibitors. recently, we described the results of a phase i-ii trial of cyclophosphamide (cy) and bortezomib (bor) where patients (pts) received cy ( mg/kg) on days (d) + & + and bor on d & + . the incidences of grade ii-iv and grade iii-iv acute gvhd were % and %. the incidence of chronic gvhd was %. in addition to gvhd, there are other factors that affect patients' quality of life and cost of care and that should be considered. it is well documented that cis have an unfavorable toxicity profile. this includes nephrotoxicity and electrolyte disturbances. furthermore, the cis need serial level monitoring. thus, we endeavored to compare the effects of cybor combination against ci-based regimens by focusing on electrolyte requirements, specifically mg, and renal function. we also sought to better understand financial considerations surrounding the need for ci drug level monitoring. sixteen pts were randomly selected from the cybor group and patients from an internal control group of patients who received mmf and cyclosporine or tacrolimus following reduced-intensity ahsct. the groups were well matched in regards to age, sex, disease status, pam score, and baseline renal function. on each pt, mg results from d to + were compiled. based on institutional protocol, a mg replacement value was assigned as well as the corresponding drug and infusion charges. next, the number of immunosuppressant (is) trough levels from d to + was tallied and the internal lab charges calculated. to compare renal function, gfr was calculated at baseline, d , and d + . χ tests and wilcoxon rank square tests were used to analyze the data. for the cybor group, median mg value was . mg/dl (iqr . ) vs . ( . ) in the control group (po . ). cybor pts required a median of grams ( ) vs grams ( ) in the control group (p = . ). the cost of mg replacement and infusion was significant (p = . ) ( table ) . for is checks, drug levels were checked a mean of . times per patient in the cybor group compared to . times in the control group (po . ), which also translates to significant savings ( table ) . considering these costs, the cybor group saved~$ . for gfr, cybor pts and control pt had reduced gfr at baseline. on d + , cybor pts had better renal function in comparison to the control group (p = . ) ( figure ). in summary, cybor significantly reduced the use of resources post-transplant and thereby the associated cost related to mg replacement and need for drug level monitoring. furthermore, cybor preserved renal function at d + . these findings could also impact patient's quality of life. although our cost analysis was restricted to certain aspects of care and did not take into account other factors, it highlights specific important benefits of ci-free gvhd prophylaxis and supplicates further study. a formal prospective comparison of cost and qol is warranted. . related donor n = and unrelated donor n = (compatibility / n = ), with conditioning regimen: myeloablative n = and non-myeloablative n = . median interval between transplantation and diagnosis of cgvhd of months( . - . ); and between cgvhd diagnosis and sativex months ( . - . ), with a median of prior treatment lines ( - ). at the time of beginning, the cgvhd was extensive in all patients, severe cgvhd n = and moderate cghvd n = . all patients except one had cutaneous involvement (n = with sclerodermal features). in addition, other organs were affected: digestive n = , pulmonary n = , hepatic n = , ocular n = , oral n = , genital n = and muscular n = . drug was started because of pulmonary affectation in patients and due to sclerodermal/muscular involvement in patients. concomitant therapies during treatment were: topical cutaneous treatment n = , topical ocular treatment n = , pulmonary n = , sirolimus n = , tacrolimus n = , oral corticosteroids n = , extracorporeal photopheresis s n = , ruxolitinib n = , imatinib n = , mesenchymal stem cells n = . the mean dose were three puff/day ( ) ( ) ( ) ( ) , with good tolerance, only two discontinuations of treatment because of adverse effects. median time of treatment days ( to ). at the time of the analysis patients were still under treatment. responses mainly occurred within the first days, s with a median time of duration of days ( in ). responses after two months of treatment were: partial organ response, mixed responses, unchanged and organ progressions; at th day ( / ) only two patients maintained their responses (one pr and one mixed response). it must be pointed out that one patient who reached pr with sativex in monotherapy maintain response after months of treatment. in addition, cramps were resolved in patients. sativex appears to be an effective treatment option in patients with chronic gvhd, particularly in those having cramps, sclerodermal features and pulmonary affectation. as seen in multiple sclerosis context, the main issue with its use is the loss of response in the long-term follow up. the median dose is inferior to the one described in ms, leaving the question if higher doses can deepen the response. these results should be confirmed in prospective trials. disclosure of conflict of interest: none. several risk factors associated with acute and chronic graftversus-host-disease (gvhd) have been identified in multiple studies. most commonly associated factors are human leukocyte antigen (hla), mismatch between recipient and donor, as well as several other characteristics such as age, conditioning regimen and prior acute gvhd. objective: the aim of this study was to evaluate the characteristics of acute and chronic gvhd in patients who underwent an allogenic hematopoetic stem cell transplantation (hsct), identify differences in the profile risk factors for acute and chronic gvhd and their impact in post-transplant morbidity and mortality. this retrospective study included mexican adult patients who received an allogenic hsct between january and march , at instituto nacional de cancerologia. we analyzed patients with a median age of years ( - ), from which, % were male patients. among the participants with hematologic malignancies, were previously diagnosed with acute lymphoblastic leukemia, acute myeloid leukemia, chronic myeloid leukemia, lymphoblastic lymphoma and with myelodysplastic syndrome. because bone marrow transplants are not performed at this institution, all transplants were from peripheral blood stem cell harvest. acute gvhd prophylaxis consisted in a triple immunosuppressive drug regimen for all patients. . % of the patients had high risk disease prior to hsct. myeloablative conditioning represented % of the applied regimens, which consisted of iv busulfan in . % of the cases. . % of patients, were transplanted within months from diagnosis. the cumulative incidence of acute gvhd at days was . % ( patients). patients with acute gvhd had % grade a, % grade b and % grade c, according to the ibmtr grading system. patients had skin involvement, with grade - acute gvhd in % of the cases, patients developed liver involvement and patients had gastrointestinal tract disease. % of the patients developed chronic gvhd, from which, % were classified as severe, . % as moderate and . % as mild. % of the patients who developed chronic gvhd had a single organ involvement, while . % had or more organs/sites. prior acute gvhd was associated with de development of chronic gvhd. the multivariate analysis identified hla unrelated donor as the only risk factor associated with the development of acute gvhd (hr, . ; % ci, . - . , p = . ). the overall survival at years was of % poor patients who developed acute gvhd and of % for those who didn´t (p = . ). our analysis showed that the incidence of acute and chronic gvhd at our center is lower than the reported at other centers, but we were not able to identify risk factors usually associated with the development of gvhd, perhaps due to the small population that we evaluated. graft versus host disease (gvhd) remains one of the main obstacles to broader application of allogeneic stem cell transplantation (sct). despite the routine use of prophylactic therapies, chronic gvhd (cgvhd) occurs in to % of patients undergoing allogeneic sct. ciclosporin a (csa) remains the backbone for gvhd prophylaxis in both myeloablative (mac) and reduced intensity conditioning (ric) sct. however, in a significant proportion of patients, csa causes important side effects and needs to be discontinued. in this study we have evaluated the impact of substituting csa for mycophenolate mofetil (mmf) as immunosuppression (is), on the incidence of cgvhd. we have compared the outcome of consecutive patients that underwent allogeneic sct from march to november at the bmt unit of the hammersmith hospital and received csa as part of the gvhd prophylaxis. of them, patients ( %) remained on csa prophylaxis for the duration of the planned post sct immunosuppression period and patients ( %) required a switch to mmf before day + . the reason for changing the is was nephrotoxicity in the majority of cases (n = , %), neurological toxicity (n = , %), disease relapse (n = , %), intolerance (n = , %) or not determined (n = , %) . we excluded from the analysis those patients whose is was changed due to the presence of acute gvhd. both groups had similar patient and transplant characteristics (see table ). [p ] however, distribution according to diagnosis showed a predominance of aml ( %) in patients that remained on csa and mds ( %) for those that switched to mmf. the mean survival rate of the entire cohort was . days (± ) . the mean survival of each group was: csa . days (± . ) and mmf . (± . ). this difference in survival reached statistical significance (p: . ). we graded cgvhd using the nih scoring system as mild, moderate and severe. out the patients that continued with csa, . % (n = ) had no cgvhd; . % (n = ) had mild cgvhd; . % (n = ) had moderate and . % (n = ) had severe cgvhd. in patients that switched to mmf . % (n = ) did not develop any cgvhd; . % (n = ) developed mild; . % (n = ) moderate cgvhd and . % (n = ) developed severe cgvhd. (p: . ). the cumulative incidence of any cgvhd at years post sct was % for the csa/mmf group and % for the csa only group (p = . ). csa is one of the standards of care for gvhd prophylaxis in both ric and mac sct. in our cohort of patients, those who remained on csa had a better overall survival and a reduced incidence of chronic gvhd compared with those patients that stopped csa and replaced it by mmf. csa toxicity should be prevented to avoid gvhd-related complications. disclosure of conflict of interest: none. although the outcome of allogeneic stem cell transplantation (sct) from an unrelated donor (ud) has considerably improved over the recent years, graft versus host disease (gvhd) still represents a severe and potentially lethal complication. in vivo t-cell depletion with anti-thymocyte globulin (atg) has been shown to significantly decrease the risk of both acute and chronic gvhd without compromising survival, however the optimal dose has not been defined yet. aim of present retrospective study was to evaluate the impact of two different doses of rabbit atg (thymoglobulin) on gvhd incidence, infectious complications and outcome of patients undergoing sct from ud. between february and september , patients received thymoglobulin mg/kg (atg- group) and received thymoglobulin mg/kg (atg- group) in addition to cyclosporin and short course mtx as gvhd prophylaxis. the two groups were comparable regarding sex, age, diagnosis and disease phase at transplant, comorbidity index, stem cell source and antimicrobial prophylaxis. conditioning treatment was myeloablative in % of atg- group patients and in % of atg- group patients. donor and recipient pairs were / hla matched in % of the cases of the atg- group and in % of the cases of the atg- group (p . ). netrophil engraftment occurred in ( %) patients at a median of days post transplant (range: - days); six patients ( in the atg- group and in the atg- group) died before engraftment. overall, patients ( %) developed grade ii-iv acute gvhd, without significant differences between the two groups (atg- % and atg- %, p . ). similarly, chronic gvhd was not significantly different between the two groups: moderate to severe chronic gvhd occurred in % of the patients in the atg- group and in % of the patients in the atg- group (p . ). univariate logistic regression analysis didn't show any significant differences between the two groups respect the incidence of bacteremia, invasive fungal infections acute and chronic gvhd. with a median follow-up of . months, patients ( %) are alive, in complete remission and after disease relapse. transplant related mortality was superimposable in the two groups (atg- % vs atg- %). kaplan-meier estimates of overall survival and event free survival were % and %, respectively, without statistically significant differences between the two groups and between hla matched and hla mismatched sct. the results of our study suggest that different doses of atg tailored on hla compatibility might be effective for preventing gvhd with any detrimental effect on overall survival and incidence of infectious complications. a prospective randomized study is mandatory to confirm our preliminary results. disclosure of conflict of interest: none. c-reactive protein levels at acute gvhd diagnosis predict steroid-resistance, treatment related mortality and overall survival after allogeneic hematopoietic stem cell transplantation l minculescu , ls friis , bt kornblit , sl petersen , i schjødt , ns andersen and h sengeløv department of hematology, rigshospitalet, copenhagen, denmark acute graft versus host disease (agvhd) remains an excessive cause of morbidity and mortality after allogeneic hematopoietic stem cell transplantation (hsct). primary treatment consists of high-dose corticosteroids, but a small group of patients are steroid-resistant and their prognosis is especially poor. a predictor of patients at risk of steroid-failure would aid the decision of additional immunosuppressive treatment at an early stage. there is experimental evidence that co-existing inflammation aggravates agvhd. since c-reactive protein (crp) is a systemic inflammatory marker, we aimed to investigate whether crp levels at agvhd diagnosis could predict the risk of failing first-line therapy and developing steroid-resistance. we retrospectively studied patients transplanted between and , table . acute gvhd was diagnosed in patients, of whom had grade ii-iv. crp, total white blood cell-, lymphocyte-and neutrophil counts were available for all patients at the time of agvhd diagnosis. according to local protocol, patients with failed response to high-dose steroid ( mg/kg) were treated with the tumor necrosis factor (tnf) alpha inhibitor infliximab and categorized as steroid-resistant. of grade ii-iv agvhd patients ( %) developed steroid resistant disease. crp levels at diagnosis among these where between o and mg/l. crp levels where significantly higher in patients who developed steroid resistance compared to patients responding to high-dose corticosteroids, p = . , hr . ( % ci . , . ). this translated into significantly increased transplant-related mortality (trm) and decreased overall survival in patients with high crp levels, figure . total white blood cell-, lymphocyte-and neutrophil counts were not associated with steroid resistance in agvhd patients. cxcr is chemokine receptor expressed on activated t lymphocytes, in particular on th cells, nk cells, dendritic cells, and subsets of epithelial and endothelial cells. cxcr ligands attract th cells into inflamed tissues and concomitantly block the migration of th cells. furthermore, inhibitory functional autoantibodies against cxcr occur in humans which play an important role in cxcr -dependent immune regulation. in addition, cxcr regulates endothelial cell homeostasis. there are two variants of cxcr : cxcr -a and cxcr -b. overexpression of cxcr -a on endothelial cells is associated with an increase in cell survival, whereas overexpression of cxcr -b dramatically reduced dna synthesis and up-regulated apoptotic endothelial death. here we have studied if a dysfunctional cxcr axis might be involved in gvhd pathogenesis and could link endothelial and t cell pathology in acute gvhd. we assessed concentrations of the cxcr ligands cxcl , cxcl and cxcl as well as anti-cxcr autoantibodies in patients with high grade ( ) ( ) acute intestinal gvhd for whom serum was available at gvhd onset. furthermore, anti-cxcr autoantibodies and cxcl levels were measured in sera stored before conditioning therapy. all variables were tested for influence on post-gvhd survival using cause-specific cox regression analysis. at gvhd onset, we observed a strong inter-correlation of cxcr ligands, but no correlation with anti-cxcr auto-antibodies. compared with pre-conditioning probes, cxcl levels strongly increased (median to pg/ml, p o . ), whereas anti- these results suggest crp levels at diagnosis as a valid predictor of developing steroid resistant disease in agvhd grade ii-iv and survival in allogeneic hematopoietic transplant recipients. [p ] s cxcr decreased (median . to . u/ml, po . ). anti-cxcr levels before conditioning and at gvhd onset correlated (coeff. . , p o . ), whereas cxcl levels did not. in multivariable analyses, low anti-cxcr and high cxcl measured at disease onset were strongest predictors of survival after acute gvhd. notably, high levels of the proinflammatory chemokine cxcl were particularly prognostic of an adverse outcome of gvhd in the presence of a high endothelial risk as assessed by the previously published easix score, while high anti-cxcr levels were most protective in patients with low easix score (that is, low endothelial risk). a score based on cxcl , anti-cxcr , and easix allowed an effective prediction of acute gvhd outcome ranging from mortality % (high cxcl + high easix) to mortality o % (low cxcl , low easix, high anti-cxcr . our data suggest a strong role for the cxcr axis in the pathology of acute high grade gvhd. the opposing effects of cxcl and anti-cxcr indicate a functional, attenuating role for these auto-antibodies. the overall prognostic impact of the immunemodulating cxcr axis appears to depend on the underlying integrity of the patients' endothelial homeostasis. despite some progress in acute lymphoblastic leukemia (all) treatment including modern chemotherapy modalities, monoclonal antibodies and newer tyrosine kinase inhibitors (tki) for ph positive cases, the final success is still difficult to reach. allogeneic hematopoietic stem cells transplantation (allohsct) has remained an essential approach in attempts to cure all. tki routinely used for all ph(+) pre-and post-transplant treatment are also described as an alternative and adjunctive approach for chronic gvhd especially with fibrotic features due to their antifibrotic activity targeting the platelet-derived growth factor receptor (pdgfr) pathways. in this study we have tried to estimate the potential influence of pretransplant tki treatment on gvhd occurrence comparing all ph(+) and all ph(− ) cases treated with allohsct. a cohort of all patients consisted of all ph( − ) and all ph(+) cases treated with allohsct was retrospectively analyzed. all patients were transplanted from sibling or unrelated donor (no haploidentical procedures were included). all ph(+) patients achieved pretransplant treatment with imatinib and chemotherapy, and ph( − ) patients with chemotherapy alone. the median age in ph( − ) and ph(+) group was vs (p = . ), the percentage of hla mismatched transplantations - ,. vs . (p = . ), the percentage of acute gvhd cases- . vs . (p = . ) and extensive chronic gvhd cases- . vs . , respectively. there were no significant difference between groups in patients sex (f/m- / vs / respectively), ric/mac conditioning, unrelated/sibling donor, donors age, bm/pbpc transplantation, number of cd cells and chronic gvhd incidence. all patients received cyclosporine-and methotrexate-based gvhd prophylaxis. gvhd occurrence was analyzed in subgroups as previously described: all ph( − ) and all ph(+). as mentioned above the incidence of acute gvhd was higher in ph(+) group (higher number of hla mismatched transplantations in this group) but the incidence of extensive chronic gvhd was higher in ph (-) group. cox proportional hazard model analysis revealed death risk caused by gvhd higher in ph negative group (hazard ratio = . ; ci % = . - . ; p = . ). the analysis of competing events was performed to estimate the probability of death caused by gvhd vs other complications (transplant related mortality, infections and relapse). the impact of conditioning was not significant on gvhd related deaths vs other complications (p = . vs . , respectively- figure ). the same results were achieved with donor cmv status (p = . vs . - figure ). we have not found any significant difference either in gvhd or other complications related deaths taking into account patient s sex/age, donor sex/age, patients cmv status, number of cd cells transplanted. on the other hand, the influence of agvhd and chgvhd on deaths related to other complications was not significant (p = . vs . ). cumulative probability of overall survival was higher in ph(+) group but the difference was not significant. the impact of pretransplant treatment with imatinib on gvhd occurrence has not been estimated so far. we are aware of our results to be preliminary and variety of data is still to be evaluated. however our results, if confirmed, may suggest the influence of imatinib on decreasing the extensiveness of chronic gvhd. disclosure of conflict of interest: none. seta-tsukinowa, otsu, - , japan; department of pathology and laboratory medicine, emory university hospital, atlanta, ga, usa; department of biostatistics and bioinformatics, rollins school of public health, emory university, atlanta, ga, usa; pathology and pediatrics, emory university school of medicine, atlanta, ga, usa, aflac cancer center and blood disorders service, children's healthcare of atlanta, atlanta, ga, usa and bloodworks northwest research institute, seattle, wa, usa more than % of allogeneic hematopoietic stem cell transplant (allo-hsct) patients receive red blood cell (rbc) and platelet (plt) transfusions in the peritransplant period. preclinical models indicate that rbc and plt transfusions trigger inflammation, raising the question of whether such transfusions are associated with development of severe acute graft-versus-host disease (grade iii/iv agvhd) and mortality in allo-hsct recipients. we conducted a retrospective analysis of rbc and plt transfusions, agvhd incidence, and mortality among consecutive adult patients receiving non-t celldepleted allogeneic bone marrow ( %) or g-csf-mobilized blood stem cell grafts ( %). common underlying diseases were acute myeloblastic leukemia ( %), myelodysplastic syndrome ( %), and acute lymphoblastic leukemia ( %) . underlying disease risk was ranked as low ( %), intermediate ( %) or high ( %). allografts were obtained from / hlamatched sibling donors ( %), unrelated donors ( %), or from donors mismatched at - hla alleles ( %). graft sources were bone marrow ( %) or mobilized pbsc ( %). the cumulative incidences of grade iii-iv agvhd and mortality prior to day without developing grade iii/iv agvhd were estimated using the cumulative incidence function and a cox proportional hazards regression model. covariates included in multivariable analysis was limited to baseline covariates associated with grade iii/iv agvhd at the p median number of rbc or plt transfusions ( figure ). univariate analysis showed a lower hematocrit on admission (median of rbc units transfused (p = . ) were significantly associated with the risk of developing grade iii/iv agvhd, while a longer time to neutrophil engraftment was inversely associated with grade iii/iv agvhd ( ⩾ median of days, hr . , p = . ). multivariate cox regression analysis showed only larger numbers of rbc units transfused and hla mismatch independently associated with severe agvhd (p = . and p = . , respectively), while underlying disease risk and larger numbers of transfused rbc units were independently associated with overall survival in a multivariate analysis that excluded agvhd grade. overall mortality rate was lowest for the group with fewer rbc and plt transfusions ( %), and greatest for the group with more rbc and plt transfusions ( %). groups that received more rbc units had higher rates of mortality due to gvhd, while patients who received more plt transfusions and fewer rbc transfusions had greater mortality from relapse ( figure ). these data support the hypothesis that peritransplant rbc transfusions are associated with the risk of developing severe agvhd and worse overall survival following allo-hsct. prospective studies are warranted to whether rbc transfusions promote t-cell activation and inflammation in allo-hsct recipients, leading to increased severe agvhd. disclosure of conflict of interest: none. early high umbilical cord blood cd chimerism associated with acute gvhd at time of onset in haplo-cord transplantation h choe , j hsu , s mayer , u gergis , a phillips , t shore and k van besien department of hematology/oncology, weill cornell medicine, new york, ny , usa introduction: haplo-cord transplantation is a combined haploidentical and cord blood transplant that allows for more rapid engraftment by the haplo with eventual loss of the haplo graft upon engraftment of the cord. haplo-cord transplants are associated with an approximate - % incidence of agvhd. reported, using chimerism assessments at approximately day after transplant for aml and mds, that lower umbilical cord blood (ucb) chimerism in the cd or cd lineages were associated with increased rates of relapse. we did not find a statistically significant association between day chimerism and risk for acute gvhd.( ) here we report our analysis of chimerisms at the onset of agvhd. patients and methods we retrospectively reviewed all patients who underwent haplocord sct for all hematologic malignancies between july and march . ucb for haplo-cord transplants were selected based on hla-typing and cell count. grafts were matched for at least of hla loci by the standard criteria and contained a minimum cell count of × nucleated cells per kilogram (kg) of the recipient's body weight before freezing. the haploidentical donor was a relative in the large majority of cases. we identified patients evaluable for agvhd (onset before day ) without preceding relapse or early death. of the total patients, patients were diagnosed with agvhd of any stage and grade. fractionated chimerisms including cd and cd components were routinely sent to evaluate for engraftment of the recipient vs haplo vs ucb. chimerism data was collected for both agvhd and no agvhd patients. the two-sided student's t-test was used to compare the agvhd cohort to the no agvhd cohort. chimerisms collected on patients with agvhd were within median ± days of onset of agvhd. the median time to onset of agvhd was days (range: - days). the median post-transplant chimerism recorded for comparison with the no agvhd patients was days. the agvhd cohort had significantly lower cd recipient (p = . ) and higher cd ucb engraftment (p = . ). all other fractions, including the cd chimerisms, were not significantly different between the two cohorts. the agvhd vs no agvhd cohorts were further compared by degree of hla mismatch ( - out of vs - out of ). the frequency of agvhd was similar in the - out of ( / , %) and the - out of ( / , %) groups. within these subgroups, cd ucb chimerism was higher for those with agvhd (p = . and p = . , respectively). conclusion the onset of agvhd in haplo-cord transplantation is associated with a significantly higher cd ucb chimerism and lower cd recipient chimerism. higher ucb chimerism may indicate that full ucb chimerism poses a higher risk of agvhd development. vice-versa persistent recipient chimerism may protect from acute gvhd. il- is a pleiotropic cytokine with both pro-and antiinflammatory properties (scheller ). the proinflammatory properties are mediated through trans-signaling that depends on the soluble il- receptor. il- trans-signaling is involved in several autoimmune diseases and in regulation of tissue regeneration of the gi-tract. specific snps in the il- receptor have been associated with increased baseline crp levels, severity of autoimmune diseases and response to interleukin- inhibition in rheumatoid arthritis. so fare little is known about the role of trans-signaling in graft-versus-host-disease (gvhd). in this study we investigated how specific snps in the il- receptor influence pretransplant levels of crp, il- sil- r and the risk of grade ii-iv acute gvhd in allogeneic stem cell transplantation (asct) in patients with family donor. dna was available for patients ( male, female median age , range: - ) and donors, that underwent asct with a matched related donor ( sibling) at haukeland university hospital in the period - . the majority received conditioning with either bycy ( ) or flubu ( ) and only patients were transplanted with tbi-based conditioning. four different snps in the il- r gene (rs , rs , rs , rs ) were chosen on the basis of (i) documented or suspected roles in autoimmune disorders; and (ii) allele frequency between . - . and r o . between the different snps. genotyping was done using kaspar assays with viia instrument (life technologies). the overall genotype call rate was %. no departures (p-values o . ) from hardy-weinberg equilibrium were observed. pretransplant serum levels of il- , sil- r and were analyzed with bio-plex kits (bio-rad, hercules, usa). both serum and dna analyses were performed in duplicates. patients being homozygous for the rs minor allele had significantly higher pretransplant serum sil r levels but lower crp levels compared with patients homozygous for the major allele. the overall incidence of agvhd requiring high-dose steroid treatment (grade ii gastrointestinal, grade iii-iv liver and skin) in the cohort was %. when analyzing the conventional clinical and laboratory parameters only transplantation with a non-sibling donor was associated with increased risk of agvhd (p-value o . hr , confidence interval . - . ). the presence of the rs in donor or recipient was associate with a significant increase in the rate of aghvd (p-value . hr . confidence interval . - . ). the snp rs (p-value . hr . , confidence interval . - . ) was also significant in an adjusted model including both donor type and rs . none of the evaluated snps were associated with an increase in early or late trm and did not influence os either. this study suggests that snps in the il- r influence pretransplant biochemical characteristics and clinical outcomes after asct. future studies investigating the effect of il- inhibition as gvhd prophylaxis or treatment should include analyses of il- receptor snps to investigate their possible influence on treatment outcomes. graft-versus-host disease (gvhd) continues to be the major cause of morbidity and mortality in allogeneic hematopoietic stem cell transplant (allo-hsct). the prophylaxis scheme varies according to the center and the country. in ours institution we use triple-prophylaxis based on cyclosporin a (cya), metrotexate (mtx), and mycophenolate mofetil (mmf). this scheme has been used for more than one decade in asian centers where it has proven adequate effective and safe to prevent gvhd. we evaluated patients undergoing allogeneic hematopoietic stem cell transplantation treated at the national cancer institute from january to december . the triple-prophylaxis scheme consists in cya (adjusted serum levels, mtx ( mg/m days + , + , + , + ) and we evaluated different doses of mmf, one of them includes mg bid × days and the other has high doses ( mg/kg bid × days), as gvhd prophylaxis. the response characteristic was analyzed using the pearson test, fisher's exact test on categorical variables and student's t-test, mann-whitney u on continuous tests. kaplan-meier method was used to estimate the probabilities of os, sle with the differences compared by the log-rank test. we analyzed patients with median of age of years (range: - ), % male gender, all were transplant with peripheral blood progenitor cells as a source. . % were acute lymphoid leukemia and . % acute myeloid leukemia, . % chronic myeloid leukemia, . % myelodysplastic syndromes, . % aplastic anemias, . % non-hodgkin's lymphomas and . % hodgkin's lymphomas. myeloablative conditioning was used in % (bucy, cfm-gat) and % reduce intense conditioning (flubu, flucy, flucy-gat), . % related hla compatibility. mmf mg twice daily (bid) for days (group ) and of mmf mg/kg bid for days (group ), in the group the . % developed febrile neutropenia vs . % in group (p = . ). the frequency of gvhd was . % group vs . % group (p = . ), chronic gvhd was . % vs . % respectively (p = . ). at the moment of analysis . % vs . % were free of disease (p = . ). there no difference at -year overall survival was % (group ) vs % (group ) (p = . ), neither free-survival disease (p = . ). the mmf regimen shows noninferiority scheme for gvhd. the low doses and for shorter administration did not show differences in the incidence and severity of acute or chronic gvhd, os, dfs compared to the mmf regimen at days with high doses. the high doses shows higher incidence of febrile neutropenia, but there were no differences in documented infections. disclosure of conflict of interest: none. a protein-losing enteropathy can develop due to conditioning regimen related gut toxicity and can cause albumin decline during peritransplant period in allogeneic stem cell transplantation (allohct). damaged intestinal mucosal barrier results in alloactivation of donor t cells and this situation is considered a primary event in the pathogenesis of acute graft-versus-host disease (agvhd). peritransplant albumin decline, as a result of conditioning regimen related protein-losing enteropathy, may predict agvhd ( ) . in this retrospective study we tested this hypothesis. we evaluated patients who received allohct between and . albumin decline from the day of conditioning initiation until its nadir in the first weeks of post-transplant period was calculated as delta albumin. acute gvhd was proven by biopsy in all patients. chi square and mann-whitney test were used for statistical analysis. patients' characteristics were shown in table- . acute gvhd was developed in patients and severe agvhd was developed in patients. delta albumin was not different between agvhd patients and no agvhd patients. delta albumin was not related with severe agvhd. delta albumin was not different between patients who received myeloablative and reduced intensity conditioning regimens. when we used a cutoff value of . gr/dl for delta albumin, we could not find a relation between delta albumin and development of both agvhd and severe agvhd. we repeated the analysis for acute myeloid leukemia (aml) and myelodysplastic syndrome (mds) patients who receive myeloablative conditioning regimen and we found the same results, there was no difference between agvhd patients and no agvhd patients in terms of delta albumin. there was a number of studies that used albumin as a predictive and prognostic marker in the setting of agvhd. but albumin may decrease in patients due to many reasons like malnutrition, proteinuria, enteropathy, liver disease or being negative acute phase reactant. because of albumin value can show variability between patients, albumin decline may be a more objective criterion. rashidi et al. showed that . gr/dl decline in albumin may be a predictor of severe agvhd in patients who was diagnosed with aml and mds and received myeloablative conditioning regimen. we repeat this analysis in our mds and aml (n = ) patients but we couldn't find this relation. when we evaluated all our patients, again there was no relation between delta albumin and development of both agvhd and severe agvhd. in conclusion, our study did not support rashidi et al.'s findings. because serum albumin level shows variability due to many reasons, it is hard to use albumin as a predictor of agvhd. sclerodermatous chronic graft-versus-host disease (scl-cgvhd) in its severe manifestation affects the patient quality of life and, due to complex pathomechanism, does not respond to standard immunosuppressive therapy-calcineurin inhibitors (cni) with corticosteroid. methotrexate (mtx) and rituximab appeared to be effective in some patients but the novel strategies, including extracorporeal photopheresis (ecp), imatinib, m-tor inhibitors (for example, sirolimus) and ruxolitinib seem to become the real breakthrough. we retrospectively analysed data of patients with scl-cgvhd, who underwent allogeneic hematopoietic cell transplantation (hct) between - in transplant centres. the study group consisted of patients with haematological malignancies and one with aplastic anaemia, female and male, with the median age ( - ). donors' median age was , with predominance of matched sibling donors ( donors) and even distribution of the donors' gender. in patients ( %) acute gvhd (agvhd) was diagnosed with skin involvement observed in ones. acute gvhd directly progressed to cgvhd in cases. in patients ( %) cgvhd developed ‛de novo' and in cases cgvhd was induced by dli. median time from hct to cgvhd diagnosis was months and to scl-cgvhd diagnosis- months. seven patients ( %) were scored as moderate cgvhd and patients ( %) as severe cgvhd according to nih- cgvhd activity classification. in patients sclerotic features had superficial form and in ones deep sclerosis was observed. chronic gvhd manifestation in other organs includes: mouth ( %), joints and fascia ( %), liver ( %), eyes ( %), gi tract ( %) and lungs ( %). patients were treated with ecp and/or sirolimus and /or imatinib with % response rate (complete-cr, partial-pr or minimal-mr). median duration of ecp therapy, sirolimus and imatinib treatment was months ( - ), months ( - ), and months ( - ), respectively. sirolimus was added more likely ( patients) as the first in case of suboptimal response to ecp after median weeks and in patients was subsequently replaced by imatinib with no favourable outcome in cases. in patients imatinib was initially used in combination with ecp therapy, leading to pr or mr. mtx without novel therapies was effective in patients with limited skin involvement, patients responded to mtx plus imatinib and patient to mtx plus sirolimus. two patients, after failure of other therapies, have been receiving ruxolitinib with improvement. only patient ( %) were nonresponsive to ecp (progressive or stable disease), patients ( %) to sirolimus and patients ( %) to imatinib. toxicity incidence was equally observed in case of sirolimus and imatinib and lead to the therapy discontinuation in altogether patients. infectious complications were observed in patients ( %). ecp confirms to be the most effective therapeutic strategy in severe forms of scl-cgvhd with favourable safety profile. imatinib and sirolimus, targeting different fibrotic pathways, both play important role in nonresponsive patients, improving the outcome in ecp and non-ecp group. in case of limited access to ecp, mtx remains to be beneficial in combination therapy of moderate scl-cgvhd and an alternative to cni. disclosure of conflict of interest: none. post-transplant morbidity and mortality are majorly determined by gvl effect counter-balanced by gvhd. treatment with systemic steroids represents the first-line therapy for gvhd, but is associated with increased incidence of infection and relapse. ecp can reduce the extent of gvhd while preserving anti-virus/-tumor activity. to elucidate this clinical phenomenon on an immunological level, we correlated clinical data with immunological findings in patients under ecp treatment. nine patients with acute gvhd (agvhd) of the gut ii-iv suffering from severe diarrhea were treated by ecp in addition to triple-drug immunosuppressive therapy. furthermore, patients with chronic gvhd (cgvhd) of the skin or lung despite triple-drug received ecp treatment. patients were evaluated according to their individual response and clinical condition. phenotypical analysis of different cellular subsets of patients and healthy donors was performed by multicolor flow cytometry. functional properties of virus-specific cd + t and nk cells were evaluated by inf-γ-elispot and cr-release assay. about patients were treated by ecp in this study. however, two agvhd and two cgvhd patients had to be withdrawn from ecp treatment after a few ecp cycles due to pancytopenia or poor clinical condition. for patients with agvhd up to ecp cycles were needed for response. all patients achieving a complete response (cr) were still alive year after initiating ecp therapy. overall response, that is, cr or partial response (pr) according to nih criteria, was obtained in of patients with agvhd ( . %) including cr in of ( . %). out of cgvhd patients ( . %) reached pr, and ( . %) remained stable under ecp treatment. after year, overall survival (os) was % for agvhd patients responding to ecp, while only % for non-responders. os for cgvhd patients was %. during intensive ecp treatment for patients with agvhd of the gut, the average stool volume and frequency decreased and consistency changed from loose to formed stool. steroids could be tapered down to a mean of % of the initial dosage. cgvhd patients were stabilized under ecp treatment and steroid dosage could be reduced to a mean of %. clinically responding patients showed increased numbers of regulatory cells including mdscs, foxp +cd + and foxp +cd +cd + tregs, as well as cd − cd − cd + t, vδ + t cells and regulatory b lymphocytes. furthermore, loss of cd l expression on effector cells like cd + te, cd + te, nk and nkt cells was observed under ecp treatment. interestingly, ecp treatment did not dramatically influence the frequency of cd +cd +cd + t, γδ t cells and nkt cells, which possess anti-virus/-tumor function. elispot and cr-release assays revealed stable anti-viral activity of cd + t cells as well as functional cytotoxicity of nk cells. moreover, cd + t, cd + tem, cd l+cd + temra, cd +cd − nk and cd brightcd − nk cells could serve as reliable biomarkers for prediction of response to ecp. conclusion: ecp treatment might stabilize or even improve clinical situation of patients suffering from gvhd. in clinically responding patients an immunomodulation was observed in terms of increasing numbers of regulatory cells with loss of migratory capacity of effector cells while anti-virus/-leukemia t-cell function was preserved. disclosure of conflict of interest: none. extracorporeal photopheresis affects dendritic cells by reducing total numbers and blunting cytokine production in patients with graft versus host disease tj altmann , , m bickerton , am flinn , u cytlak , p milne , s pagan , m collin , , v bigley , and ar gennery , institute of cellular medicine, newcastle university, newcastle upon tyne, united kingdom and newcastle upon tyne hospitals nhs foundation trust, newcastle upon tyne, uk graft versus host disease (gvhd) and concomitant immunosuppression is a leading cause of morbidity and mortality post hematopoietic stem cell transplantation (hsct). the pathophysiology of gvhd is complex, involving presentation of histo-incompatible antigen by activated recipient dendritic cells (dcs), activation and proliferation of donor t cells and resultant tissue damage. extracorporeal photopheresis (ecp) is a second-line treatment for steroid refractory or dependent gvhd that facilitates the reduction of immunosuppression. ecp's mechanism of action is unclear and is likely to be multifaceted. apoptosis of lymphocytes, induction of a th favoured environment and increased numbers of regulatory lymphocytes have been implicated . although ecp has been shown to modify the function of in vitro monocyte-derived dcs , its effect on primary (non monocyte-derived) dcs has not been studied. our aim was to determine whether ecp had immediate or long-term affects on primary dc numbers or function. we enumerated monocyte and dc subsets (cdc , cdc myeloid dcs and plasmacytoid dcs) in whole blood before, during and after ecp cycles, and developed a novel dc function assay, suitable for use on clinical samples. four adults with immunosuppression withdrawal gvhd and four children with acute gvhd, received ecp during the study. all received ciclosporin gvhd prophylaxis and corticosteroid treatment at onset of gvhd. children received ⩾ dose of infliximab prior to starting ecp. adults received two ecp treatments (one cycle) every weeks and children received two ecp treatments (one cycle) weekly. whole blood was taken before and after each cycle of ecp. trucount flow cytometry analysis of whole blood was used to enumerate mononuclear leukocytes. to assess function, peripheral blood mononuclear cells were isolated by density centrifugation and stimulated with toll-like receptor agonists. cell-specific cytokine production was then analyzed by flow cytometry. samples were compared to healthy controls and pre-ecp samples. median time to first ecp treatment from gvhd diagnosis was . days. no gvhd flares were experienced during study period. ( ) adult had a cycle of treatment delayed due to intercurrent pneumonia. numbers of cdc , pdcs and classical monocytes were significantly reduced after each ecp treatment in the adult group. dc numbers followed the same trend after ecp in the paediatric group but were not significantly different before and after ecp. this is perhaps due to initial lower dc count compared to adults in the children before the first ecp cycle. functional analysis showed a reduction in cytokine production in dcs and monocytes in both groups over the course of ecp treatment. our data support a cell-intrinsic effect of ecp on monocytes and dcs, with numerical and functional consequences. this may contribute to the beneficial effect of ecp both through reduction of inflammatory effector function and through modulation of interactions with other immune cells. correlation with immunosuppression withdrawal and clinical events during treatment may provide further insight into the role of monocytes and dcs in gvhd and ecp, which may aid in the development of novel targeted therapies for gvhd. extracorporeal photopheresis as early second-line treatment for patients with steroid-dependent or refractory acute graft-versus-host disease: a single-centre experience i sakellari , i batsis , e gavriilaki, a-k panteliadou, a lazaridou, k leontopoulos, d mallouri, a bouinta, v constantinou, e yannaki, c smias and a anagnostopoulos department of hematology bmt unit, g. papanicolaou hospital, thessaloniki, greece acute graft-versus-host disease (agvhd) remains a severe complication of allogeneic haematopoietic cell transplantation (allohct). corticosteroids as the backbone of initial therapy for agvhd result in varied complete responses ( - %). traditional secondary treatments lead to profound immunosuppression without improved survival. on the basis of our experience in chronic gvhd, we aimed to prospectively assess the role of extracorporeal photopheresis (ecp) as early secondline treatment in steroid-dependent and refractory agvhd. we enrolled consecutive patients with steroid-dependent or refractory grade (gr) ii-iv agvhd post allohct from january to august . all patients with unrelated or haploidentical donors received thymoglobulin (atg) mg/kg as prophylaxis. post-transplant gvhd prophylaxis included cyclosporine-methotrexate in myeloablative and cyclosporine-mycophenolate mofetil in reduced toxicity or intensity regimens. ecp was commenced after assessment of response to days of steroid treatment according to our protocol: two sessions per week for month, one session per weeks for months, evaluation of response and one session per month for months. we studied patients, aged ( - ), post allohct with myeloablative ( ), reduced toxicity ( ) and intensity ( ) conditioning, from sibling ( ), matched ( ) or one locus mismatched ( ) volunteer unrelated and haploidentical ( ) donors. disease risk index was high ( ), intermediate ( ) and low ( ). acute gvhd was observed at day + ( - ) in patients, late onset at + ( - ) in patients and induced at + post donor lymphocyte infusion in a relapsed aml patient. skin, intestine and liver involvement was evident in patients, skin and intestine in and skin only in patients. nine patients ( with grii, with griii agvhd) were steroid-dependent and ( with griii, with griv) steroidrefractory. atg was administered simultaneously with ecp initiation in six refractory patients that further developed ebv reactivation (p = . ) treated pre-emptively with rituximab. ecp was commenced at day + for ( - ) sessions. the majority of patients ( / ) presented partial ( ), very good ( ) or complete ( ) response to ecp. with . ( . - ) months of follow-up, immunosuppression was reduced in / and ceased in patient. clinically significant bacterial infections were found in patients, fungal in , cmv and ebv reactivation in and , respectively, and other viral in patients. cumulative incidence (ci) of chronic gvhd was . at year. one-year ci of agvhd-related mortality was %. one-year overall survival (os) was % and significantly increased in steroid-dependent vs refractory patients ( % vs %, p = . ). reduction of immunosuppression (p = . ) and response to ecp (p = . ) were associated with improved os, irrespectively of other factors. our data indicate that ecp should be considered early in the course of steroiddependent or refractory agvhd, before irreversible end organ damage has been established. optimal timing of intervention, frequency, duration and tapering schedule of ecp remain important unanswered questions. [p ] disclosure of conflict of interest: none. extracorporeal photopheresis for treatment of chronic graft versus host disease m lanska, a zavrelova, j radocha and p zak faculty of medicine, th department of internal medicine-hematology, university hospital hradec kralove, czech republic allogeneic stem cell transplantation represents a curative approach to many hematologic disorders. graft versus host disease (gvhd) is a complication with significant morbidity, mortality and decreased quality of life. extracorporeal photopheresis (ecp) represents possible treatment approach. mononuclear cells (mnc) collected by apheresis are photosensibilized with -methoxypsoralenem ex vivo, irradiated with uva and transfused back to the patient. aim of the study: evaluation of patients treated with ecp for gvhd at our center. thirteen patients ( females and males, median age years) were treated with ecp. about patients (pts) had matched sibling donor and patient had unrelated donor. about pts had sclerodermic form of gvhd, had concomitant pulmonary gvhd, had pulmonary gvhd alone. one pts had mild, seven moderate and five severe gvhd according to nih. about patients were treated with steroids. mnc separation was prepared on cobe spectra and spectra optia (terumo bct, usa). -methoxypsoralen was added, irradiation was done on macogenic g (macopharma, mouvaux, france). about procedures in pts were performed (median procedures, - procedures). the schema was as follows: ecp on consecutive days every - weeks first months with subsequent increase in interval. median follow up was months. in sclerosing form two pts reached cr, six pts pr, one is stable and one patients progressed. in pulmonary gvhd one reached cr, two partial improvement, one is stable. seven pts are still alive, six died (two due to relapse, one secondary malignancy and three infections). it was possible to withdraw steroids in pts. adverse events were clinically negligible. ecp is an effective treatment for chronic gvhd. especially sclerodermic form responds to ecp very well. it is safe and well tolerated procedure with minimal toxicity. supported by prvouk p- . disclosure of conflict of interest: none. allogeneic haematopoietic stem cell transplantation (hsct) is a potentially curative treatment option for children with a variety of haematological, oncological and immunological diseases. graft versus host disease (gvhd) represents a major cause of post-transplantation mortality and morbidity affecting multiple organs including skin, gut, liver and lungs. gvhd is considered a succession of inflammation and donor t-cell activation initiated by translocation of gastro-intestinal microorganisms through impaired mucosal barriers after chemotherapeutic conditioning and/or infection. diagnosis of gvhd is based on clinical symptoms and histological findings, necessitating invasive and potentially harmful procedures including endoscopy and biopsy. as yet, no non-invasive markers are available for diagnosis or treatment monitoring in children with gvhd. faecal calprotectin (fc) reflects intestinal mucosal inflammation of any origin. in the setting of allogeneic hsct in adults, fc has shown to be a marker for acute (steroid-resistant) gvhd. we aimed to evaluate the feasibility of prospective fc measurement as a non-invasive marker for diagnosis and treatment in children with gvhd. a prospective, observational, single centre study was started in july . by december , paediatric allogeneic hsct patients (age - years) were included after informed consent. faecal samples were collected from weeks before to months after hsct. fc levels were measured by elia, according to manufacturer's instructions. clinical symptoms were prospectively evaluated and managed according to local guidelines. if gvhd was suspected on clinical grounds, histological confirmation was obtained. first-line therapy for gvhd consisted of corticosteroids. in case of steroid-resistant disease, more advanced immune modulation was applied. a total of five patients developed histologically confirmed gvhd: acute gvhd of skin and gut (n = , one patient with steroidresistant disease), acute gvhd of skin only (n = ); chronic gvhd of lung only (n = ) and acute gvhd of skin followed by chronic oromucosal gvhd (n = ). without exception and regardless of gut involvement, gvhd occurrence was accompanied by rises in fc levels to values μg/g (range: - μg/g). fc levels correlated with clinical and histological grading. moreover, adequate response to therapy was consistently reflected by return of fc levels to values o μg/g. sensitivity of fc levels to diagnose gvhd was poor due to increased fc levels in patients with posttransplant complications other than gvhd such as viral reactivation and pulmonary or gastro-intestinal infections. fc levels reflect gvhd occurrence and correlate with clinical and histological grading in paediatric allogeneic hsct patients. fc levels increase in case of gvhd regardless of gut involvement, supporting a central role for (subclinical) intestinal inflammation in gvhd initiation. although, in this interim analysis, fc lacks sensitivity to diagnose gvhd, fc may serve as a noninvasive marker for monitoring therapy response and, thereby, reduce the need for repeated invasive procedures including endoscopy and biopsy. acute graft-versus-host disease (agvhd) is a major complication of allogeneic hematopoietic cell transplantation (hct), and glucocorticoids are typically used as first-line treatment. the aim of our study was to evaluate the effect of first-line ecp +/ − steroid therapy in order to reduce the incidence of infections and toxicity. from december to january , of pts ( %), were diagnosed with agvhd grade ⩾ following allosct. pts were treated with ecp +/ − steroid as first-line therapy. about ( %) pts were treated with ecp only and ( %) with ecp + steroid - mg/kg/day. we compare this cohort with an historical group of patients, transplanted between and , who were treated with steroid only for grade - agvhd (n = out of ). the two cohorts were well balanced in terms of median age (p = . ), disease type (p = . ), disease status (p = . ), graft source (p = . ), conditioning regimen (p = . ) and hct-ci (p = . ). there were more female patients (p = . ) and more haploidentical transplant (haplo-sct) (p = . ) in the cohort treated with ecp+/ − steroid. ecp was performed using the offline technique, and was started as soon as possible with a treatment schedule consisting of four rounds of two procedures per week, three rounds of two procedures every other week and finally two procedures every month. steroid was tapered as soon as possible after starting ecp. the clinical response was evaluated at day + . median follow-up for alive patients was months for ecp group and months for control group. there was no difference in terms of median time of agvhd onset ( vs days) and number of pts with grade or - agvhd ( figure ). ecp was started after a median of ( - ) days from agvhd diagnosis. every patient underwent a median of ( - ) ecp procedures, during a median time of months. on day after starting agvhd treatment with ecp+/ − steroid, pts ( %) achieved cr or pr, pts did not respond and experienced agvhd relapse to front-line therapy. one year cumulative incidence (ci) of agvhd relapsed/refractory was . % for ecp+/ − steroid. these percentages were not different from the cohort receiving steroid alone. ci of moderate-severe cgvhd was lower in the ecp group, probably due to the higher frequency of haplo-sct with pt-cy in the ecp group. about days after agvhd onset, ci of infection ( % vs %), especially cmv reactivation ( % vs %), was lower in the ecp group, but was not statistical significant. ecp allowed a faster taper of steroid: ( - ) vs days ( - ) (p o . ). overall survival, progression-free survival, non-relapse mortality and ci of relapse rates did not differ in the two groups. in multivariate analysis, visceral involvement by agvhd was associated with an increased risk of failure to front-line therapy (hr: . ; range: . - ; p = . ). this observational study suggests that the overall response rate of ecp +/ − steroids is similar to steroid alone for front-line treatment of grade - agvhd, but is potentially associated with lower incidence of infection, and in particular of cmv reactivation. a prospective phase clinical trial is warranted to address whether augmentation with ecp may be beneficial for agvhd frontline treatment. [p ] a -year-old girl with neuroblastoma received autologous stem cell transplant (asct), followed by antibiotic prophylaxis and filgrastim. her transplant preparative regimen consisted of busulfan and melphalan. engraftment of neutrophil took place on day after asct. twentieth day after asct, she experienced nausea and diarrhea. there was neither skin rash nor elevation in liver enzymes. the diarrhea continued to worsen day by day and reached to a daily volume of ml/ m . infectious studies for stool and blood including testing for influenza a and b, parainfluenza, adenovirus, epstein-barr virus, amebiasis, cryptosporidium parvum, cytomegalovirus, clostridium difficile, salmonella, campylobacter, yersinia and shigella were all negative. colonoscopy and endoscopy were performed by an experienced pediatric gastroenterologist and findings were suspicious for severe graft versus host disease (gvhd). colonoscopy and rectoscopy revealed severe inflammatory changes, friability and patchy dark exudates on the mucosa of rectum. endoscopy revealed erosions, ulcers in the esophagus and a pale mucosal surface with reticulated submucosal vessels accompanied with erosion and erythema in the antrum of stomach. grade gvhd was confirmed by pathologic analysis that revealed diffuse crypt dropout and mucosal erosion on rectal mucosal biopsy. mucosal erosions, apoptosis of epithelial cells and small lymphocytic infiltration of the lamina propria were found on duodenal biopsy. after these results, we started methylprednisolone intravenously at a dosage of mg/kg/day. on the fourth day of treatment we increased the dosage to mg/kg/day and added cyclosporine to treatment. because of unresponsiveness to treatment we decided to administer thirdparty mesenchymal stem cells (msc) ( × cd +/cd + cells per kg). these were given intravenously at day + asct as single infusion. the second dose was given at day + . within days after first application of mscs, the frequency of diarrhea decreased to one-third. at day + after second dose of mscs, the patient's stool became nearly normal. we tapered the steroids first and stopped cyclosporine at + th days after asct. discussion and conclusion: to our knowledge, this is the second case report of spontaneous severe autologous gvhd in a child with a solid tumor malignancy. regarding the pathogenesis of autologous graft-versus-host disease, there may have multiple causes for the loss of tolerance to self because of disrupted immune system. alteration of t regulatory cells by previous chemotherapy may be key point. endogenous cells that survive conditioning and assist in post-transplant maintenance of self-tolerance may be affected. microchimerism due to maternal cells transmitted during fetal development and persisting throughout adult life has also been postulated as a cause. however it is not very clear for factors that may contribute to the pathogenesis of this rare disease. autologous gvhd has the potential to cause critical illness in the hematopoietic stem cell transplantation patient population. in patients with multiple myeloma some experts report pathologically verified gastrointestinal gvhd as high as %. responses to steroids are variable. however, a significant proportion improve dramatically after early therapeutic intervention. so clinicians and pathologists should be aware in suspecting and recognizing gvhd in patients with diarrhea to guide therapy as soon as possible. disclosure of conflict of interest: none. haplosct patients did not receive additional gvhd prophylaxis aside from the ex vivo t-cell depletion (tcd) with clinimacs system. of the bud bmts out of patients engrafted, % of which had gvhd. the % who did not have any gvhd, relapsed. eighty one percent had agvhd, of which majority ( %) were grade ( table ) three patients did not have agvhd ( / mud and two cord blood grafts), but developed cgvhd. one of the patients who had grade agvhd died and one still has intermittent cgvhd years post bmt. there were % who had cgvhd. currently, all of our haplosct receive a cd /cd ra tcd grafts (n = ). the depletion techniques for the others were either cd tcd, cd / cd ra /tcrab tcd+cd +/cd ra tcd; tcrab tcd. all patients who received haplosct engrafted and % had agvhd ( % grade ) ( table ). we noted that some of the patients presented with nonclassical agvhd signs (upper gut gvhd, oral gvhd, blood). there were no patients who presented with grade agvhd. cgvhd in the cohort was %. of note, % of patients did not have any gvhd and did not receive any form of immunosuppression post bmt. only eight patients received further immunosuppression for agvhd, median duration (range: - ) days. at the time of this report, there are four patients with agvhd still receiving immunosuppression all o days and all on tapering doses. haplosct using ex vivo tcd techniques has a lower risk of gvhd with comparable, if not superior outcome to bud. the degree and duration of immunosuppression is also much less. this may translate to earlier immune reconstitution and less viral reactivation. [p ] disclosure of conflict of interest: none. allogeneic hematopoietic stem cell transplantation (allo-hsct) offers a potential cure for several hematological diseases, but it is burdened by severe life-threatening complications, being gvhd the major cause of morbility and mortality. recently, more have been understood of the physio-pathologic relationships between endothelium and graft-versus-host disease (gvhd), showing that vascular endothelium is an early phase target of gvhd. in recent years, the direct count of circulating endothelial cells (cec) has emerged as a valuable biomarker of endothelial damage in a variety of disorders. however, due to their rareness and complex phenotype, different published techniques have showed variable degrees of uncertainty, reporting a wide range of cec values in healthy subjects. by means of the commercially available rare cell isolation platform cellsearch system, for cec identification and count, we correlated cec count changes to gvhd onset and response to treatment in allo-hsct patients. cec were analysed in allo-hsct patients ( aml, all, hd, nhl, cll, mds, cms, mm, saa) at the following time points: t (pre-conditioning), t (pre-transplant), t (engraftment), t (day+ or onset of gvhd), t ( week after steroid treatment). the median cec/ml at t was (range: - ), in comparison to a value of (range: - ) in healthy controls (p %: or . , % ci . - . ; p = . ). we confirm that cec count represent a valid biomarker to monitor endothelial damage in patients undergoing allo-hsct and can be a valuable tool in supporting the diagnostic definition of gvhd and in monitoring responsiveness to treatment. moreover, the use of the cellsearch system can be crucial in order to move routinely cec monitoring into clinical practice of allo-hsct. reference clinicaltrials.gov nct . disclosure of conflict of interest: this research was conducted with the support of the investigator-initiated study program of janssen diagnostics, llc to ca. kb is employee of janssen diagnostics. results of hla mismatched unrelated donor (mmud) hematopoietic cell transplants (hct) are worse than results of fully matched hct due to higher risk of gvhd, infection and graft failure. atg during conditioning reduces incidence of gvhd but can increase risk of infection and relapse. high doses posttransplant cyclophosphamide ( × mg/kg) prevent gvhd in haploidentical hct. we initiated this approach instead of atg in hct from one alelle or antigen mismatched unrelated ( / )mmud-hct in . here we present outcome of patients (cy-group) transplanted between and , comparing it with outcome of patients transplanted between and from / mmud with atg-f (fresenius) mg/kg given during conditioning. patients in cy-group ( males, females) were transplanted from mmud mismatched for hla ( a- , b- , c- , dr- ). about patients had aml, mds, cll, cml, all and mps. med. age of patients was years ( - ). about patients received myeloablative (flu- mg/m + iv bu . mg/kg) and nonmyeloablative(flu- mg/m+mel - mg/kg +-tt mg/kg) conditioning. about patients received pbpc and bm as a graft. graft versus host prophylaxis consisted of cyclophosphamide ( mg/kg aibw) on d+ and + , cyclosporine a from d and mmf from d+ . all patients received antibacterial, antifungal, hsv and pcp prophylaxis. historical control (atg) group consisted of patients ( males, females), med. age y ( - ) who had mmud-hct for aml- , mds- , nhl- , mf- , all- , cll- , cml- , h.d- , saa- , mps- and mm- . there were mismatches for a, for b, for c and for dr. myeloblative conditioning was used in and nonmyeloablative in patients. all patients received atg-f mg/kg × given d- and- . cyclosporin was initiated d- and mmf d- . all patients received anti-infectious prophylaxis as described previously. three of patients from cy group died so far. two of them due to relapse and one due to toxicity and infection during aplasia. five patients relapsed . two achieved cr after dli and one is alive in relapse expecting second hct. about patients are alive, of them in cr. eight patients experienced agvhd (gr.i- , gr.ii- ,gr.iii- , gr. iv- ) and eight developed clinically mild cgvhd). about patients from atg group are alive - m (med. m) posthct. about patients died - m postransplant (med. m) due to vod, gvhd, infections and relapse. -day mortality is % ( / ) in cy group and % ( / ) in atg group. one year mortality is % ( / ) in cy and % ( / ) in atg group. patients from cy group have % probability of os at months posthct vs % from atg group. cyclophosphamide × mg/kg instead of atg fresenius( mg/kg) for gvhd prophylaxis reduced -day and -year mortality, and improved probability of m os significantly in our cohort of patients. this approach seems to be safe and effective in / mmud-hct. disclosure of conflict of interest: none. high transplanted cd + cells are not associated with beneficial effect on graft-versus-host disease-free, relapse-free survival (grfs) after allogeneic hematopoietic cell transplantation y lee and ih lee department of hematology, kyungpook national university hospital and inhee lee the success of allogeneic hematopoietic cell transplantation (allo-hct) is comprehensively assessed by individual comorbidity, relapse, graft-versus-host disease (gvhd) and death. besides, inconsistent results have been reported regarding the dose of cd + cells. in the current study we have addressed the issue of the potential effect of stem cell dose on the of gvhd-free/relapse-free (grfs) associated with cd + cells doses. we retrospectively reviewed the medical records of the patients who received allo-hct for acute myelogenous leukemia (aml), myelodysplastic syndrome (mds) and acute lymphoblastic leukemia (all) between and in kyungpook national university hospital. the grfs included grade - acute gvhd, systemic therapy-requiring chronic gvhd, relapse or death. the patients were reclassified into two groups according to the targeted cd + cell doses ( × per kg) by knuh protocol. a lower cd + group (n = , . %), patients who underwent allo-hct with cd + cell dose o × per kg; and a higher cd + group (n = , . %) patients who underwent allo-hct with cd + cell ⩾ × per kg. the median age at transplant was . years (range: - years) and male was patients ( . %). primary diseases for allo-hct were aml/mds (n = , %) and all (n = , %). one hundred forty-three patients ( . %) were in cr (complete remission), ( %) in further cr and ( . %) in relapsed and refractory status. one hundred seventy-one patients ( . %) received myeloablative conditioning regimen. gvhd prophylaxis consisted of methotrexate and cyclosporine a or mtx and tacrolimus. the median dose of cd + cell was . × per kg (range: . - × per kg) in lower cd + group and . × per kg (range: . - . × per kg) in higher cd + group. there was no significant difference in neutrophil, platelet engraftment between two groups. the incidence of chronic gvhd was more frequent in higher cd + group ( . % vs . %, p = . ). the median follow-up duration was . months, with a range of . - . months. the -year overall survival (os), relapse free survival (rfs), nonrelapse mortality (nrm) and graft-versus-host disease (gvhd)free/relapse-free survival (grfs) since hct was . ± . %, . ± . %, . ± . % and . ± . %, respectively. there was no significant difference according to the infused cd + cell dose (figure ). the relapse rate was not proportionally affected by the cell dose ( . % vs . %, p = . ). and there was no significant correlation between the number of cd + and cd + cells infused (spearman correlation coefficient: p = . ). in a univariate analysis, patients transplanted with the higher cd + cell doses and higher cd + cell doses had no increased grfs (p = . and p = . ). an independent factor associated with worse grfs was risk status at transplant (hr = . , % ci: . - . , p = . ). these results suggest that careful assessing the cd + and cd + graft content and tailoring the cell dose infused may help in reducing cgvhd risk without negative impact on grfs. a large and prospective study in a homogenous population will be needed to confirm the effect of stem cell dose. disclosure of conflict of interest: none. imatinib associated with extracorporeal photopheresis can fully reverse severe sclerotic-type lesions in patients with chronic graft-versus-host disease: the lille university hospital experience l magro , j gauthier, b catteau , l mannone , a lionet , v coiteux and i yakoub-agha lille university hospital and nice university hospital severe sclerotic-type chronic graft-versus-host disease (cgvhd) is difficult to reverse and can dramatically alter the quality of life of patients after allogeneic hematopoietic cell transplantation (allo-hct). imatinib or extracorporeal photopheresis (ecp) used separately yield sustained responses in s only about % of patients with steroid-refractory cgvhd. given their respective modest efficacy we hypothesized that the combination of imatinib with ecp could lead to higher response rates. we are reporting here on seven patients with severe steroid-refractory sclerotic-type cgvhd treated at our institution using this combination. we retrospectively analysed all patients treated at our institution (n = ) with the combination of imatinib with ecp for severe steroidrefractory scgvhd. imatinib was started at mg/day and increased to mg/day if well tolerated. the cellex closed system was used for ecp. ecp was initiated twice weekly during weeks. after this « induction » period, ecp sessions were scheduled less frequently according to the response to treatment. additional immunosuppressants were tapered gradually in responding patients. initial grading and response evaluation was determined according to the nih criteria. steroid-refractoriness was defined as progression of gvhd on high-dose steroids (⩾ mg/kg) or progression during corticosteroid tapering. patient characteristics are displayed in table . patients received an allo-hct between may and april . median age at allo-hct was (range: - ). a variety of myeloablative (n = ) and non-myeloablative conditioning regimens were used (n = ). antithymocyte globulin was used before allo-hct in one patient. gvhd prophylaxis consisted of ciclosporine and methotrexate in six patients. one patient received tacrolimus and methotrexate. five patients had prior history of acute gvhd. nih global severity grade was severe in all patients (n = ) due to severe sclerotic features. the median number of previous therapies was (range: - ). all patients were steroid-refractory. after a median follow-up of months (range: - months) the overall response rate was %. the complete response rate was %. median time on ecp associated with imatinib was months (range: - months). median time to best response was months (range: - months). corticosteroids could be discontinued in all patients after a median time of months (range: - months). patients # , # and # received maintenance therapy with ecp upon discontinuation of imatinib. in four patients, both ecp and imatinib led to complete response and could be discontinued after , , and months for patients # , # , # and # , respectively. patient # and # passed away after due to a myocardial infarction and the development of a solid tumour, respectively. patient # was off therapy while patient # remained on maintenance with ecp. both remained in complete response. patient # remained in response during months before progression of cgvhd while on imatinib and ecp. none of our patients experienced adverse events related to either imatinib or ecp. despite the limited number of patients in this report, we observed that the combination of imatinib and ecp can lead to complete and sustained reversal of severe steroid-refractory sclerotic-type cgvhd. these encouraging results should be confirmed in a larger cohort. disclosure of conflict of interest: lm: therakos (honorarium). allogeneic hematopoietic cell transplantation (allo-hsct) is an established treatment modality that is potentially curative for many patients (pts) with acute myeloid leukemia (aml). aml itself is the most common indication for pts undergoing hsct nowadays. for pts with high-risk disease, allo-hsct is, perhaps, the most effective curative treatment and is considered the standard post-remission therapy in first complete remission (first cr). this is a retrospective study to analyze those variables which were associated with patients' overall survival (os) after allo-hsct. the study population consisted of pts who were diagnosed of aml from january to july at the hospital universitario central asturias, and submitted to allo-hsct in first cr. risk status based on validated cytogenetics and molecular abnormalities following recommendations of european leukemianet was performed. sixteen ( . %) were male. median age was years old (range: - ). clinical characteristics at transplantation are represented in table . median follow-up was months ( - ). considering the donor type, os at year was higher in pts receiving sd ( . %) compared to % in those who received urd (p = . ). regarding graft source, os at year was . % who received pbsc compared to % in pts receiving bmsc (p = . ). gender also showed significant association with os, which was higher among men, os at year was %, compared to . % for women) (p = . ). the presence of minimal residual disease (mrd) detected using multiparametric flow cytometry was performed prospectively after induction and consolidation, and before transplantation. thirteen pts had negative mrd before transplantation. median os was greater in pts with negative mrd before transplantation compared to the group with positive mrd ( vs months, respectively) (p = . ). this difference did not reach statistical significance probably because the low number of the sample. thirteen pts developed agvhd. only ( . %) pts receiving sd developed agvhd compared to ( %) pts among those who had an urd; however this association was not statistically significant (p = . ). also, we observed higher incidence of agvhd in bmsc group ( pts; %) whereas only ( . %) in pbsc group developed agvhd. this tender did not reach significant association (p = . ). one year os was . % in pts who developed agvhd and . % who did not (p = . ). all factors that had a significant influence on pts survival were included in a multivariate analysis (cox regression model): graft source, donor type, pts gender and agvhd development. developing agvhd kept an independent association with mortality (or . , % ci . - . , po . ) and male gender also persisted as an independent protective factor (or . , % ci . - . , p = . ). in our series, agvhd has shown a significant and independent association with os over other parameters such as graft source, type of donor or mrd before transplantation. identifying reliable predictors for agvhd development, controlling well known risk factors for this disease, as well as improving management of immunosupressors should still be the key to potentiate longer os in our patients. larger studies are needed to confirm our results. acute and chronic graft-versus-host diseases (gvhd) are associated with increased morbidity and mortality after allogeneic hematopoietic stem cell transplantation (allo-hsct). older patients undergoing allo-hsct may experience a high degree of transplant-related complications and this concern has historically limited the use of allo-hsct for some older patients. in many studies, age has been shown to be a negative prognostic factor for survival and associated with higher transplant-related mortality (trm). however, in others, age was not shown to be a significant factor if appropriate adjustments for other comorbidities are incorporated in the analyses. there are very few studies that evaluated the relationship between patient's age, the presence of gvhd and long-term transplantation outcome. the aim of this study is to evaluate the impact of age in patients who develop acute and/or chronic gvhd after allo-hsct for hematological malignancies on the trm incidence. we included in the study patients with hematological malignancies who received allo-hsct and were followed in our center between january and january . for the purpose of this study, only patients who developed grade ii-iv acute gvhd and/or limited or extensive chronic gvhd where considered for analysis (n = ). patients were split into three homogeneous groups according to age at transplantation taking into consideration the underlying disease, type of conditioning and disease response at transplantation. group (younger) included patients aged o years (n = ), group (intermediate) included patients aged between and years (n = ) and group (older) included patients older than years (n = ). gvhd evolution over time was followed as well as the cumulative incidence of trm was calculated in case of acute or chronic gvhd in each group. thirty seven percent of grade ii gvhd occurred in the younger group (n = ), % (n = ) in the intermediate group and % (n = ) in the older group; majority ( %) resolved in the younger group as well as in and % in the latter two groups, respectively, while trm rates at year were %, % and %, respectively, sdhr = . , p = . . among patients who had acute gvhd grade ii, , and % in the three respective groups developed chronic gvhd later. grade iii-iv gvhd occurred in % (n = ) in the younger group, % (n = ) in the intermediate group and % (n = ) in the older group; with a respective resolution in %, % and % of patients and were associated with comparable trm rates at year of %, % and %, respectively, p = . . among patients who had acute gvhd grade iii-iv, , and % in the three respective groups developed chronic gvhd later. de novo chronic gvhd was observed with a higher rate in the intermediate and in the older group (table) while patients with extensive chronic gvhd older than years had significantly higher trm at years ( %) compared to % in those younger than years, sdhr = . , p = . . patients who develop acute gvhd grade iii-iv could incur over % of trm at year independently of age. resolution of acute gvhd grade ii was significantly better in younger patients while older patients with grade ii acute gvhd or with extensive gvhd had higher mortality compared to younger ones. in addition to an adapted prophylaxis, a better preemptive gvhd strategy should be warranted in older patients. [p ] disclosure of conflict of interest: none. in vivo effects of nilotinib on lymphocyte subpopulation and function following allogeneic stem cell transplantation e marinelli busilacchi , a costantini , j olivieri , n viola , s coluzzi , e pirro , g mancini chronic graft versus host disease (cgvhd) is a major complication of allogeneic stem cell transplantation and is characterized by frequent multiorgan involvement resembling autoimmune diseases; its pathogenesis is still incompletely defined and a standard treatment is lacking. donor-derived cd + and cd + t lymphocytes have been considered the main effector cells mediating cgvhd pathogenesis; however, recent studies suggest that b cells might also play an important role. in vitro data indicate that tyrosine kinase inhibitors (tkis) such as imatinib and nilotinib affects both innate and adaptive immune response by interacting with different cell populations (t cells, b cells, dendritic cells, mast cells and macrophages). we sought to evaluate the impact of different doses of nilotinib on the distribution and function of lymphocyte subpopulations. we analyzed samples obtained from patient with steroiddependent/refractory cgvhd enrolled in a phase - study with nilotinib in steroid-refractory cgvhd (nct ): triplets of patient were treated with escalating doses starting from mg/die ( ), mg/die ( ), up to mg/die ( ) . blood and plasma were collected at baseline and at day and of therapy. trough plasma nilotinib concentrations had been previously determined by hplc (abstract c , haematologica: evaluation of nilotinib safety in patients with steroid-refractory chronic graft-versus-host disease: a phase i-ii gitmo study). peripheral blood mononuclear cells were isolated by density gradient centrifugation using ficoll biocoll. six color flow cytometry analysis (facs canto ii) was performed using conjugated antibodies (anti-cd , cd , cd , cd , cd , cd ). inflammatory cytokine analysis was performed on plasma samples according to the instruction of bioplex pro human cytokine plex assay (bio-rad). statistical analysis was performed by -tailed student's t-test; differences were considered statistically significant for po . . flow cytometry analysis showed that nilotinib did not exert any significant impact neither on the proportion of t lymphocytes subpopulation (cd +cd + t helper, cd +cd +cd + t regulatory, cd +cd − t cytotoxic), nor on b lymphocytes and nk cells. on the contrary, a statistically significant and dose-independent decrease of pro-inflammatory and th- cytokine production was observed ( figure ): reduction of il (po . ), il (po . ) and ifnγ (po . ) were already significant after days; decreases of il (po . ) and tnfα (po . ) become significant after days. interestingly, after days of therapy, among the patients enrolled (according to the itt criteria) ten patients showed cgvhd improvement and the other five remained stable. this study shows that therapeutic doses of nilotinib can reduce plasma levels of inflammatory cytokines without affecting the proportions of lymphocyte subpopulations. these findings correlate with clinical response and suggest that besides the previously demonstrated anti-fibrotic effects, nilotinib has also potent anti-inflammatory and immune regulatory properties, supporting its role in patients with cgvhd. disclosure of conflict of interest: none. [p ] previously published p infectious gastro-enteritis after allogeneic hematopoietic transplantation after reduced intensity conditioning (allo-ric): incidence and possible role in gastro-intestinal acute gvhd i garcía-cadenas , r martino , a esquirol , a bosch , n rabella , s saavedra , c muñoz , j briones , s brunet and j sierra hematology and microbiology departments and hospital de la santa creu i sant pau, autonomous university of barcelona, spain enterotoxigenic c. difficile-associated associated disease or infection (cdi) is a common cause of diarrhea after hematopoietic stem cell transplantation (sct). recent studies have suggested the relationship of cdi with gastro-intestinal (gi) graft-versus-host disease (gvhd). the possible role of other types of infectious gastro-enterocolitis (g-ec) in gvhd development has not been studied. as a prior investigation to a national prospective observational study on this issue, we conducted a single-center retrospective analysis including all adult patients who received an allo-ric sct between january and march . the aim was describing the cause(s) (if known), timing and outcomes of recipients with possible g-ec (defined as new onset acute diarrhea grade ⩾ ) in the first year after sct. of the patients studied (median age: years, % male, % aml or mds as underlying disease), ( %) had a total of episodes of acute diarrhea, with ( %) developing more than one event. these acute diarrheas occurred at a median of days (range: - ) after sct. overall, a g-ec causing pathogen was identified in of stool specimens ( %) and included: cdi ( ), c. jejuni ( ), rotavirus ( ), adenovirus ( ), norovirus ( ), b. hominis ( ), s. stercoralis, g. lamblia, a. caviae, salmonella enterica and cryptococcus (one in each case). most postransplant diarrheas ( / ; %) occurred during the weeks after infusion and were attributable to mucosal damage caused by the ric (negative microbial screening and no evidence of gvhd).the rate of infectious g-ec among the diarrheas occurring after day + was % ( / ). the overall incidences of enteric infection were . % ( % ci: . - . ) and . % ( % ci: . - . ) at + and + months after sct, respectively. all the infected patients had mild to moderate disease, and no deaths were attributable to this complication. there were no differences in year-os and nrm between the infected and uninfected patients ( % vs %, p = . and % vs %, p = . , respectively). in univariate analysis age o years, prior sct, donor type, atg administration and prior grade - agvhd were associated with development of infectious gastro-enteritis. in multivariate analysis, unrelated donor and grade - agvhd were the only factors significantly associated with gastrointestinal infection (hr . ; % ci: . - . , p = . and hr . , % ci . - . , p = . ; respectively). acute gvhd occurred in % of patients (n = ), with a median onset of days (range: - ). the cumulative incidences of - acute gvhd at days and months post-sct were % ( % ci: . - %) and . % ( % ci: . - %), respectively, and there was a trend toward a higher risk of - gvhd in the group of patients with an enteric pathogen ( . % vs % at year, p = . ). more importantly, an enteric infection occurred just before the onset or aggravation of gvhd in / infected patients in our study ( %) at a median interval of days after the infection (range: - ). in summary, our results confirm that enteric infections are a common complication after allo-ric, representing at least % of the episodes of acute diarrhea during the first year post-sct. a possible interplay between infectious g-ec and gvhd was observed in this study. disclosure of conflict of interest: none. long-term efficacy of extracorporeal photopheresis in chronic graft versus host disease m nygaard , t karlsmark , n smedegaard andersen , i schjødt , s lykke petersen , l smidstrup friis , b kornblit and h sengeløv department of dermatology, bispebjerg hospital, copenhagen, denmark and department of hematology, rigshospitalet, copenhagen, denmark chronic graft versus host disease (cgvhd) activity is known to fluctuate over time, so we evaluated cgvhd continuously throughout the extracorporeal photopheresis (ecp) treatment course and after stopping ecp. patients with at least year follow-up, who were treated with ecp at department of dermatology, bispebjerg hospital between and were evaluated. a single investigator retrospectively evaluated response to ecp monthly for months, every months until years and every months until years. prednisolone doses were recorded every months. responses were defined as complete remission (cr) if no symptoms of cgvhd were present, partial remission (pr) as improvement in cgvhd or stationary cgvhd with more than % reduction in prednisolone, no change (nc) as no difference in symptom burden and o % reduction in prednisolone. progressive disease (pd) was defined as worsening of symptoms with unchanged or intensified immunosuppressive medication. ecp was performed with therakos uvar xts or cellex. there were evaluable patients with moderate (n = ) or severe (n = ) steroid-refractory, dependent or -intolerant cgvhd. the median age was years (range: - ) and there were females and males. conditioning regimen was myeloablative (n = ) and non-myeloablative (n = ). seventeen had related donors and had unrelated. stem cell source was peripheral blood (n = ), bone marrow (n = ) or umbilical cord blood (n = ). number of organs affected by cgvhd was one (n = ), two (n = ), three (n = ), four (n = ) or five (n = ) and involved organs were skin (n = ), eyes (n = ), mouth (n = ), lungs (n = ), genitals (n = ), liver (n = ), musculoskeletal system (n = ) or gastrointestinal tract (n = ). time from diagnosis of cgvhd to first ecp was median days (range: - ) and time from referral to ecp and the first ecp procedure was median days (range: - ). at the time of the first ecp procedure patients were also treated with prednisolone (n = ), sirolimus (n = ), calcineurininhibitor (n = ), mycophenolate mofetil (n = ), imatinib (n = ), methotrexate (n = ) or rituximab (n = ). one patient received no immunosuppression. total number of ecp cycles was median (range: - ). responses over time are shown in figure . overall response to ecp was seen in ( %) of the patients. most responses were seen after more than months ecp treatment. in univariate analysis of possible baseline predictors of response, no significant associations were found. prednisolone dose was significantly reduced at every months after start of ecp (p o . ). additional cgvhd treatment was administered to ( %) patients during ecp treatment (sirolimus n = , calcineurininhibitor n = , uva n = , methotrexate n = , rituximab n = , mycophenolate mofetil n = ). about ( %) patients had more than additional treatment. prednisolone dose was increased at least once in ( %) patients during ecp treatment. overall survival at years was %. follow up was median days (range: - ). more than half the patients with cgvhd ( %) improve overall after treatment with ecp, but flares in cgvhd activity still occur. prednisolone dose is significantly reduced at all time points after starting ecp, but short term increased doses or additional immunosuppression was necessary in more than one-third of the patients. larger prospective studies with long-term end points are warranted. disclosure of conflict of interest: marietta nygaard has received a travel grant and speaker's fee from therakos/ malinckrodt. chronic graft versus host disease (cgvhd) remains a major cause of morbidity and mortality after hematopoietic stem cells transplantation despite the improvement of the immunosuppressive prophylaxis. skin, buccal, lacrymal and hepatic disorders are the most frequent. sclerotic gvhd remains a severe form and often refractory to standard treatment lines such as corticosteroids and calcineurin inhibitors. the antifibrotic activity of imatinib by the inhibition of pdgf-r and tgfb-β pathways has been used in the treatment of refractory gvhc with sclerotic features and systemic scleroderma. here, we report the results of imatinib treatment in patients (pts) with refractory cgvhd. over a period of years (january -december ), pts received allogeneic stem cells transplantation from related donors, of whom were treated with imatinib for refractory cgvhd: pts for malignant diseases ( cml, aml, nhl) and pts for aplastic anemia. the median age is years ( - ), the sex ratio m/f: . . conditioning regimen used with chemotherapy alone: myeloablative ( pts with gvhd prophylaxis combining ciclosporin and methotrexate), reduced intensity ( pts with prophylaxis combining ciclosporin-mycophenolate mofetil). all pts received peripheral blood stem cell transplant with an average of cd cell count: . × /kg ( . - . ). the median duration of the cgvhd is months ( - ). the firstline treatment consisted of the combination of steroidsciclosporin with or without mycophelonate mofetil. imatinib was administered to these pts after median treatment duration of months ( - ) for moderate ( pts) and severe ( pts) cgvhd according to the nih classification. treatment with imatinib, at doses ranging from to mg/d, was introduced in the second line for all pts. the evaluation is conducted in october after a median follow-up of months ( - ). tolerance was good except in a one pt with severe thrombocytopenia that led to a transient cessation of treatment. after months, analysis of pts who received imatinib according to couriel criteria and nih criteria: complete remission (cr): pt ( %), partial remission (pr): pts ( %), stable disease (sd): pts, failure: pts ( %). a long-term evaluation performed after a mean duration of treatment months ( - ) finds similar results with a cr: pts ( %), pr: pts ( %), sd: pts ( %) and failure: pts ( %). corticosteroids were tapered or discontinued in pts (cr or pr). at october , pts ( %) were alive and pts ( %) died of severe infections. treatment with imatinib seems to be a good therapeutic option in the treatment of cgvhd in its moderate or severe form refractory to a minimum of two immunosuppressive agents according to the nih criteria as shown by our results in terms of response and survival with good tolerance. disclosure of conflict of interest: none. atg significantly reduces the risk of cgvhd both in unrelated and in hla identical sibling. the finke's study randomised pts undergoing an allogeneic unrelated stem cell transplant (sct) after a myeloablative regimen to receive or not mg/kg atg-grafalon reporting a significant reduction of cgvhd without increase of relapse and no os and dfs effect. however a successive study didn't confirm those results (significant reduction of acute and chronic gvhd but poorer survival mainly due to higher relapse probability in the atg arm). the conflicting data reported on urd sct have several explanations, one is about the dose and the timing of atg. the timing of atg infusion has been demonstrated to be crucial for cb transplant : an earlier administration is still active in preventing gvhd while ensuring engraftment and low hampering of immune reconstitution. here we report a large ( sct) retrospective monocentric analysis on low atg doses (and - mg/kg for bm according to the degree of hla matching and mg/kg for all pbsc sct) given early (from day − to − ). pts in the study were aml (n = , %), all (n = , %), hr mds (n = , %), cr (n = , %) cr or (n = , %), active disease (n = , %) for al; median age was (range: - ). myeloablative conditioning were bu-cy (n = , %), bu-flu (n = , %), edx-tbi (n = , %), other (n = , %); pbsc was used in % (n = ); sct were performed between and at the bologna transplant center. sct were performed from hla / identical urd (n = , %), or from / (n = , %), / (n = , %) and o / (n = , %). median follow up was months. overall, grade - agvhd was %, grade - agvhd %; cumulative incidence (ci) of cgvhd of any severity was %, for moderatesevere cgvhd %. ci of relapse and nrm was % and %, respectively. the -year overall and disease-free survival were % ( % ci: - %) and % ( % ci: - %). the gvhd (agvhd grade - and moderate-severe cgvhd) and relapse free survival (grfs) of the entire population ( figure ) was % at years ( % ci: - %). restricting the analysis to patients in cr - , we found that cgvhd (any severity), gfrs and os at years were %, % and %, respectively. comparing transplants with / urd to mismatched ones ( / or less) we found a trend for increased mod/sev cgvhd in pts undergoing transplant with mismatched urd (shr . , % ci: . - . , p = . ); agvhd grade - and cgvhd overall were not significantly increased; relapse incidence according to hla mismatches resulted % and % in / and ⩽ / , respectively; grfs was % in / and % in ⩽ / . the data reported show that low and early administration of atg is able to effectively prevent acute and chronic gvhd without increasing relapse thus ensuring really convincing grfs, even for o / matched urd transplants. graft versus host disease (gvhd) is one of serious complications in patients after allogeneic hematopoietic stem cell transplantation. the application of mesenchymal stem cells (msc) represents a promising method for the treatment of severe steroid refractory gvhd. we present the data from an interim analysis of clinical trial, within which we applied msc in patients with acute or chronic gvhd after allogeneic transplantation. the diagnoses included aml ( pts), mds ( pts), all ( pts), cll/nhl ( pts), mpn ( pts). the patients underwent sibling hla-compatible ( ), haploidentical ( ), unrelated hla-compatible ( ) or hla-mismatched ( ) transplants. the median interval between the transplantation and msc was months ( - ). the indications for msc infusion were steroid resistant acute gvhd ( pts), steroid-dependent gvhd (agvhd pts, cgvhd pts) or chronic gvhd with the need for long-term immunosuppression and corticosteroid intolerance ( pts). msc were applied as a single infusion at a median dose of . ( . - . ) × /kg. response to treatment was assessed on day , , and . the severity of gvhd prior to msc was graded as clinical stage ( - ) in acute and stage ( - ) in chronic gvhd, respectively. the median dose of corticosteroids was . ( . - . ) mg/kg/day in agvhd and s . ( . - . ) mg/kg/day in cgvhd patients. on day + the partial response (pr) was achieved in % of patients with agvhd, the stabilisation of gvhd (sd) was found in % of patients with cgvhd. the dose of corticosteroids was reduced in most patients with agvhd (to % of the starting dose; - %), while the early reduction was possible only in % of cgvhd patients. on day+ only patients were evaluable. the agvhd patients ( pts) achieved a significant clinical response: pr, cr and dose of corticosteroids was reduced in all of them (to %; - %). the minor responses were achieved in cgvhd patient ( pts.) with pr and sd. however the dose of corticosteroid was reduced in % of these cases (to % of the initial dose; - %). a total of patients died because of infectious complications. most of them ( / ) were agvhd patients who expired early up to day + . there were observed no side effects of msc application neither during the infusion nor later during the follow-up of ( - ) months. the analysis of lymphocyte reconstitution revealed the changes of kinetics of some subsets as compared to the day + benchmark. the b-lymphocyte count tended to decrease in % of patients from chronic gvhd subgroup (vs % in agvhd). conversely nk cells declined in most agvhd patients ( % vs % in cgvhd). also the pro-inflammatory th cell was affected especially in agvhd (decrease in % pts vs % pts in cgvhd). the counts of myeloid/plasmocytoid dendritic cell increased in %/ % agvhd and %/ % ccvhd patients. the screening testing of cytokines (raybiotech, cytokines, pts, day + to + ) revealed changes of some analytes after msc infusion, including a decrease of proinflammatory cytokines such as ifn-γ, tnf-α, il- . our experience with the treatment of gvhd using msc confirmed the safety of this immunotherapy. the favourable clinical effect with reduction of severity of gvhd and steroids dose was observed, especially in patients with acute form of gvhd. methotrexate day + omission is not associated with higher incidence of acute graft-versus-host-disease mm rivera franco, e leon rodriguez and a campos castro allogeneic hematopoietic stem cell transplantation (allo-hsct) remains a high-risk procedure due to its related morbimortality, limiting the broader application of this important treatment modality. despite extensive research over the years, acute graft-versus-host-disease (agvhd) affects the majority of patients undergoing allo-hsct, and up to % will develop clinically significant grades (ii). over the years, several methods for gvhd prophylaxis have been implemented, including immunosuppresive agents. methotrexate (mtx) is one of the earliest drugs used for gvhd prophylaxis. frequently, a short course of intravenous methotrexate (given on days + , + , + and + after hsct) is combined with a -month tapered course of cyclosporine. there is no consensus on which drugs and schedules for prevention of gvhd are best and clinical practice varies by institution. further, it is not clear whether omission of the day + dose of mtx has a negative effect on outcome in terms of morbidity. to describe the frequency of acute and chronic gvhd, mucositis and engraftment in patients receiving methotrexate (plus csa) as prophylaxis, omitting day + . ninety-five consecutive patients who underwent allo-hsct from to september , and received mtx as immunosuppressive prophylaxis were included. all patients received three doses of mtx, always excluding day + . mtx was administered iv, either mg/m day + , + , + or mg/m day + , and mg/m during days + and + . we included patients ( % male). the most frequent underlying diseases were aplastic anemia ( %) and acute lymphocytic leukemia ( %). ninety-nine percent of patients had a matched related donor. forty patients ( %) had gender disparity with their donor, and % presented abo incompatibility (major in %). most of the patients received myeloablative conditioning regimens (n = , %). the median of cd + infused cells were × (range: . - . ). the median neutrophil and platelet engraftment was ( - ) and (range: - ) days, respectively. from all the cohort, only patients ( %) developed acute gvhd ( % grades i-ii) ( figure ). thirty patients ( %) developed chronic gvhd, which was limited in %. most of the patients, % (n = ), presented acute toxicity after the conditioning regimen, from which % (n = ) corresponded to superior mucositis ( % grade i-ii and % grade iii-iv. the -year overall survival was % and the -year relapse free survival was %. our results showed a low incidence of acute gvhd, mostly grades i-ii, and similar survivals compared to previously reported studies, proposing that the administration of day + mtx as gvhd prophylaxis is not mandatory, however, prospective studies might be necessary to test our results. [p ] disclosure of conflict of interest: none. outcome of refractory graft versus host disease (gvhd) treated with extracorporeal photopheresis (ecp) as second line: a single-center experience j cornago navascues, b aguado bueno, av arriero garcía, edc jimenez barral, i vicuña andres and a alegre amor hematology department, university hospital la princesa, madrid, spain gvhd is a common and, sometimes, life-threatening entity related to hematopoietic stem cell transplantation (hsct). steroids remain the first-line therapy but they are not always enough to control it, or their side effects are simply unacceptable. both acute and chronic gvhd are responsible of impairment occurred in different organs that can lead to increase morbidity and mortality in our patients. different options are available as second line, but it is a well known fact that ecp, due to its inmunomodulatory mechanism, yields satisfactory response rates and presents excellent safety profile. from may to october , patients with steroid-dependent or refractory gvhd have been treated in our centre with ecp. we have performed ecp procedures with the therakos cellex device, an integrated 'on line' system. the transplant was from a sibling donor in cases and from an unrelated donor. the median of cd + infused was . × cd /kg. eight patients ( . %) presented agvhd, ( . %) cgvhd and finally, ( . %) had an overlap gvhd syndrome. most of patients ( . %) with agvhd had a severe intestinal involvement as the main manifestation of the disease. however, all patients with cgvhd had a multiorgan involvement with a median of four organs affected, being skin, mouth, eyes and lungs the most common implicated. ten patients in our series have died, for gvhd complications or infections and due to relapse of aml. as firstline treatment they all received steroids and cyclosporine or mycophenolate mofetil. median ecp per patient has been ( - ). ecp procedures were performed for consecutive days, in initial phase weekly (in those with agvhd), or every weeks (cgvhd) and then monthly according to clinical response, evaluated by clinical assessment and reduction in immunosuppression. about % of patients with agvhd had a significant clinical response to ecp so that steroid doses could be tapered and even in . % of them withdrawal was possible. in the cgvhd group overall response rate (orr) to ecp was . %. in % of these patients steroids could be suspended after a median of . ecp procedures. all patients who responded to ecp in cgvhd are still alive. independently of gvhd type, . % of patients responded to ecp and % of them even could stop steroid therapy. those who had no response are dead. in cgvhd, . % of patients remain alive, in contrast with agvhd or overlap syndrome patients whom survival is around %. about adverse events, % of patients did not present any complication associated with ecp. complications were mostly related to central venous catheter, with cases of bacteremia and thrombosis, easily recovered. in our experience, ecp is effective as second line treatment in gvhd, obtaining the best results in the chronic gvhd group. in fact, cgvhd patients with a good clinical response to ecp, specially when steroid doses can be tapered, have the better outcomes and longer survival. the tolerance to the procedure is excellent without severe adverse events. more experience is required to determine the best scheme of ecp and its role as prophylactic treatment. mesenchymal stromal cells (mscs) possess immunomodulatory properties and may play important roles in graft-versushost disease (gvhd) and engraftment. this study examined co-transplantation of mscs and hscs (hematopoietic stem cells). we investigated co-administration of ex vivo expanded mscs along with hla-identical sibling-matched hscs in β thalassemia major patients. we recruited patients from january to january in our study. all participants received cyclophosphamide-based or fludarabine-based conditioning regimens and short-course methotrexate and cyclosporine as gvhd prophylaxis. mscs were administered intravenously ( . - . × /kg) into patients (n = ) h before infusion of hscs. the outcomes were then compared to those of patients transplanted with hscs alone. the median follow-up in the msc and non-msc group was . and . years, respectively. median time to wbc engraftment . × /l was . days (range: - days) in both groups (p-value = . ) and median time to platelet engraftment × /l was . days (range: - days) in the msc group, while it was . days (range: - days) in the non-msc group (p-value = . ). fifty-six percent of patients had acute gvhd in the msc group compared to the non-msc group where . % developed acute gvhd (p-value = . ). meanwhile, chronic gvhd was % in the msc group and % in the non-msc group (p-value = . ). although the incidence of acute and chronic gvhd was lower in co-transplantation of hscs and mscs, no statistically significant difference was noted between the two groups. three-year overall survival rate was % and % in the msc and non-msc group, respectively (p-value = . ). three-year thalassemia-free survival rate was % in the msc group and % in the non-mscs group, showing no statistically significant difference (p-value = . ). the -year rejection incidence in the msc and non-msc group was % and %, respectively (p-value = . ). there was no statistically significant difference between the two groups in terms of -year transplant-related mortality (pvalue = . ). this study indicates that co-transplantation of hla-identical sibling hscs with mscs does not inflict harm on bone marrow transplantation procedure and seems to be safe and secure. on the other hand, differences between the two groups in acute and chronic gvhd, engraftment, overall survival, thalassemia-free survival and rejection incidence did not reach statistical significance. therefore, despite the immunomodulatory activity of mscs and their role in gvhd amelioration and engraftment improvement resulted from in vitro studies, their efficacy in the clinical setting has not been conclusively proven which indicates further multicenter randomized clinical trials are required. keywords: β-thalassemia major, co-transplantation of mesenchymal and hematopoietic stem cells, engraftment, graft-versus-host disease. hematology-oncology and stem cell transplantation research center, tehran university of medical sciences, shariati hospital, tehran, iran. disclosure of conflict of interest: none. a number of studies were published with contradictory results comparing tacrolimus (tac) and cyclosporine a (csa) for graftversus-host disease (gvhd) prophylaxis, but there are only few that accounted for pharmacokinetic (pk) parameters. in this retrospective study we have identified pk parameters that affected gvhd incidence and incorporated them in the s multivariate comparison of tac-and csa-based prophylaxis. the retrospective study included consecutive patients with csa and consecutive patients with tac prophylaxis. % were grafted from matched related donor (mrd) and % from unrelated donor (ud). about % received busulfanbased myeloablative conditioning (mac) and % reducedintensity conditioning (ric). second agent for gvhd prophylaxis was short-course methotrexate (mtx) - mg/m on days + , , , in % of patients and mycophenolate mofetil mg/kg days − to + in %. unrelated graft recipients also received antithymocyte globulin (atgam, pfizer, ny, usa) mg/kg. the pk parameters analyzed were mean and median concentrations, pk variability parameters and number of concentrations below the targeted limit (nlow) within , and days after hsct. for tac the highest predictive value for acute gvhd was observed for median concentration during first days (auc = . ), and for absolute skewness (auc = . ) of concentration data. for csa parameters with highest predictive value were median concentration (auc = . ) and variability coefficient (auc = . ) during first days. nlow was also a significant parameter for tac current gvhd prevention regimens are partially effective, delay immune reconstitution, impair graft versus tumor effect and are cumbersome to use. therefore, there is a pressing need to develop innovative approaches for the prevention of gvhd. we completed a phase i-ii study employing a calcineurin and mtor inhibitor-free regimen based on a combination of post-transplant cyclophosphamide and bortezomib (cybor) in patients receiving fludarabine and busulfanbased reduced-intensity conditioning followed by peripheral blood, matched related or unrelated transplant. patients receiving grafts from unrelated donors also received ratg. we reported that the regimen was feasible and safe and yielded promising outcomes. ( , ) herein, we compare the results to those of a quasi-contemporaneous matched group of patients receiving a calcineurin-based gvhd prophylaxis. the experimental and control groups were well-matched in terms of age, sex, donor type, disease status, renal function and pam score. the cybor group (n = ) was treated during a timeframe spanning from to and the control group (n = ) from to . gvhd prophylaxis for the control group was mmf and csa (n = ) or tacrolimus (n = ). both groups received supportive care according to standard institutional protocols. median follow-up for the cybor group was . months as opposed to . for the control group. median times to neutrophil engraftment for the cybor and control groups were days ( - ) and ( - ), respectively (p = . ). two patients from the cybor arm died before achieving platelet engraftment. for the remaining s patients, median time to platelet engraftment was days ( - ). for the control group, five patients never dropped their platelet count below × /l. for the remaining patients, median time to platelet engraftment was days ( - ) (p = . ). there was no primary or secondary graft failure in either of the two groups. the incidences of acute grade ii-iv and grade iii-iv for the cybor group were . and . %. for the control group, the incidences were (p = . ) and % (p = . ). the incidence of chronic gvhd for the cybor and control groups were % and . %, respectively (p = . ). treatment-related mortality was . % and % for the cybor and control groups, respectively (p = . ). the incidences of cmv, ebv and bk reactivation for the cybor group were . %, . % and . %, respectively. for the control group, the incidences were . % (p = . ), . % (p = . ), % (p = . ). the -year progression free survival and overall survival were . % and . % for cybor group and . % and . % for the control group ( figure ). the year gvhd and disease-free survival (grfs) were . % and %, respectively. despite the limitations of our study that include its size and its design and the delayed neutrophil and platelet engraftment associated with the cybor regimen in comparison to calcineurin-based prophylaxis, our data confirm the promising outcomes previously reported with the cybor combination and reaffirm the need for a large randomized study comparing cybor to a standard calcineurin-based regimen. clostridium difficile infection (cdi) causing enterocolitis may represent a serious clinical problem in patients undergoing allogeneic hematopoietic cell transplantation (allo hct). the reported prevalence varies substantially among heterogeneous patient cohorts. although cdi has been proposed as a risk factor for the development of gastrointestinal (gi) acute graft-versus-host-disease (agvhd), limited knowledge on the prevalence of cdi, occurrence of gi agvhd in cdi patients, relapse incidence and mortality of cdi patients in large patient cohorts is available. the aim of this analysis was to study the implications of cdi in a homogenous cohort of patients with either aml or mds undergoing allo hct. at our center all patients undergo stool test once a week for clostridium difficile antigen while in aplasia until discharge, irrespective of clinical symptoms for enterocolitis. patients with positive stool antigen tests (that is, toxin test) in the absence of clinical symptoms were referred to as cd+, in contrast to patients without a positive test and without clinical symptoms which were referred to as cd − . we retrospectively analyzed the data of a total of n = patients with either aml or mds undergoing allo hct in our institution between and . overall survival (os) was measured from allo hct to the date of death or last follow-up. after hsct, relapse and nonrelapse mortality were considered as competing events. eventprobabilities were calculated according to kaplan-meier for os and using competing event statistics for the cumulative incidence of relapse (cir), non-relapse mortality (nrm) and agvhd. % confidence intervals (ci) were provided for major endpoints. statistical analyses were performed using the r environment for statistical computing version . . (r core team , vienna, austria, www.r-project.org). from a total of n = patients with either aml or mds who underwent allo hct, we identified n = ( %) who were cd − , n = ( %) who were cd+, and n = ( %) who had cdi. interestingly, n = ( %) of patients with cdi were diagnosed having gi agvhd as compared to n = ( %) of patients who were cd+ and compared to n = ( %) of patients who were cd − , p = . . the three groups harbored no differences when comparing incidences of liver and skin agvhd or chronic gvhd, respectively. when dissecting gi agvhd according to ctcae criteria, only n = ( %) of cdi patients vs n = ( %) of cd+ patients, and n = ( %) of cd − patients had grade - gi agvhd, p = . . with regard to os and trm, no statistical differences were observed between the three groups. the cir was % for patients with cdi, % for cd+ patients and % for cd − patients, p = . , respectively. this analysis represents the largest published analysis of clostridium difficile in patients with aml or mds who underwent allo hct. the prevalence of cdi in this patient cohort was %. patients with cdi developed significantly more often gi agvhd as compared to patients who were either cd+ or cd − , respectively. however, this did not translate into differences in os or trm. disclosure of conflict of interest: friedrich stölzel has received research funding from astellas. the majority of studies on cytokines in allogeneic hsct were performed with classical gvhd prophylaxis, consisting of nonspecific immunosuppressive agents. with this type of prophylaxis almost in all studies published, higher levels of proinflammatory cytokines are associated with development of acute gvhd, while lower levels indicate the success of immunosuppressive agents in abrogation of alloreactive response. currently, there is no data, whether the dynamics of cytokines after ptcy is similar to the situation of classical gvhd prophylaxis. out of adult patients transplanted at first state medical university with ptcy between and we have identified cases with acute gvhd and plasma samples available. these patients were matched in the ratio : to patients who did not develop acute gvhd. the study group was comprised of adult patients with hematological malignancies who underwent hsct. all patients received ptcy-based gvhd prophylaxis. five plasma biomarkers were studied by elisa: il- a, il- , il- , tnf-α and ifn-γ. blood samples were obtained from patients on days − , , + , + . the fifth time point varied between day + and + to represent the sample after engraftment, but before onset of acute gvhd. about ( %) out of gvhd patients had a grade i, ( %) grade ii, ( %) grade iii agvhd, ( %) patients developed multiorgan agvhd. about patients ( . %) had chronic gvhd. there was no difference between gvhd + and gvhd − groups in any of the clinical parameters. the median of engraftment for all patients was ( - : range). the median agvhd was days ( - : range). neither of the cytokine levels was significantly different in patients with agvhd grades i − iv and without gvhd. however, for patients with agvhd grade ii − iv we found that low levels of il- on day + ( . ± . vs . ± . pg/ml, p = . ) and ifn-γ on day + - ( . ± . vs . ± . pg/ml, p = . ) were associated with increased risk of gvhd. the roc analysis was performed to determine the cut off values for il- - . pg/ml (auc = . ) and ifn-γ - . pg/ml (auc = . ). the incidence of agvhd grade ii-iv was significantly higher in patients with levels of cytokines lower than cut off ( % vs . %, p = . and . %, p = . for il- and ifn-γ, respectively). the same pattern was observed for patients with agvhd grade iii-iv. low levels of il- ( . ± . vs . ± . pg/ml, p = . ) and ifn-γ ( . ± . vs . ± . pg/ml, p = . ) on day + were especially predictive. the cut off values for il- was . pg/ml (auc = . ) and for ifn-γ- . pg/ml (auc = . ). the incidence of agvhd grade ii-iv was also significantly higher in patients with levels of cytokines lower than cut off (p = . and p = . for il- and ifn-γ, respectively). for chronic gvhd only higher level of il- at day + ( . ± . vs . ± . pg/ml, p = . for patient with and without gvhd, respectively) was significantly predictive. in this pilot trial we have demonstrated that dynamics of cytokines after gvhd prophylaxis with ptcy may be different from conventional one, and well-known predictive biomarkers might not work after ptcy. further large prospective trials are warranted to elucidate reliable biomarkers for gvhd after this type of prophylaxis. disclosure of conflict of interest: none. graft versus host disease (gvhd) remains one of the main obstacles to broader application of allogeneic transplantation. gvhd prevention and treatment techniques are poorly standardized. the st-line treatment of newly diagnosed chronic (c) gvhd is corticosteroid. there is no standard ndline treatment for cgvhd. approximately - % of patients (pts) with cgvhd require secondary treatment within y after initial systemic treatment. recently the jak / inhibitor ruxolitinib emerged as an efficacious treatment for corticosteroid-refractory (sr) acute and c-gvhd with a % of sr-cgvhd patients reporting a long lasting immunosuppression-free complete response. the current study seeks to analyse the efficacy and safety of ruxolitinib in highly pre-treated sr-cgvhd pts in our centre. ruxolitinib treatment was given off label after provision of an informed signed consent and in the absence of alternative therapeutic options including clinical trials. we analysed data prospectively collected at our long-term follow-up clinic between and . a written consent was given by pts allowing the use of medical records for research in accordance with the declaration of helsinki. overall pts (median age y-range: - years; mean karnofsky score %) with sr-cgvhd were treated s with ruxolitinib. median time from transplant was months (range: - ). ruxolitinib was initiated at a starting dose of mg twice daily-median time on ruxolitinib months (range: - )- / pts increased the dose up to mg twice daily. four pts had a classic and an overlap sr-cgvhd. all of them had skin sclerodermatous involvement and / joint and fascia involvement with significant decrease of range of motion and limitation of adl. all pts were previously treated with several lines of immunosuppression ( - ) including high-dose prednisone in st line ( / ), rapamycin ( / ), tk-inhibitor imatinib ( / ), extracorporeal photopheresis ( / ). all pts were pre-screened for risk of infection and regularly checked on a fortnightly basis. all pts were under active prophylaxis according to recommendation for gvhd pts and ruxolitinib therapy. after a cumulative follow-up of days we reported only one serious adverse event represented by a cmv pneumonia requiring hospitalization with complete recovery. early time point evaluation ( / pts evaluable) at + month underlined how all pts were reporting subjective improvement at the patient global ratings according to nih . data were confirmed at the health care provider global ratings. month evaluation ( / ) confirmed meaningful responses (partial responses / ) according to nih , with both patient and health care provider global ratings improvement and concomitant enhancement in lee skin symptoms score and sf- health-related qol. at last followup no evidence of myelosuppression, infections, pml, nonmelanoma skin cancer was registered. considering the concomitant treatment (with reference to azoles and rapamycin or cyclosporine) no cases of toxicity due to drug-druginteraction was reported. ruxolitinib is well tolerated in highly pre-treated sr-cgvhd. its safety profile seems to be reassuring. the efficacy data observed also at this early time point is preliminary but promising in this subset of pts with a long history (⩾ lines) of treatment for cgvhd. confirmatory study in a larger number of patients is underway on a multicentre basis. disclosure of conflict of interest: none. severe acute enteral graft-versus-host-disease (gvhd) is a lifethreatening complication of allogeneic bone marrow transplantation. in case of resistance to corticosteroids as the firstline treatment severe enteral gvhd harbors a high morbidity and mortality. retrospective analyses indicate efficacy of the jak / -inhibitor ruxolitinib in the treatment of acute or chronic gvhd in adults, but experience in paediatric patients is limited. here, we report a small cohort of paediatric patients with stage steroid-refractory gvhd of the gut who received ruxolitinib as salvage therapy within a multimodal immunosuppressive regimen. we retrospectively analysed four patients aged - years with severe, steroid-resistant acute gvhd of the gut who were treated with ruxolitinib in our institution. all patients were transplanted for non-malignant haematologic disorders, graft source was × mmud, × mud, × msd. the conditioning regimen consisted of treosulfan, fludarabine and thiotepa. serotherapy with thymoglobuline was administered in all patients transplanted from unrelated donors. all patients received mtx and cyclosporine as gvhdprophylaxis. gvhd was staged according to the glucksberg-scale. ruxolitinib was added to the immunosuppressive regimen when acute stage gvhd was reached and became resistant to treatment with methylprednisolone ( mg/kg/day) as well as infliximab and mycophenolate (mmf) as second-line immunosuppressants. acute stage enteral gvhd developed at a median of days after transplant ( - days) and ruxolitinib was started at a median of days post-transplant ( - days). the starting dose varied between mg/day and mg/day, that is, . - . mg/kg/day, taking into account the expectedly low bioavailability of the oral drug during severe diarrhea. upon improvement of gvhd symptoms and/ or increasing side effects the dose was gradually tapered and ruxolitinib was discontinued after a median of treatmentdays ( - days) . after addition of ruxolitinib to the immunosuppressive regimen, the symptoms of acute gut gvhd gradually improved in all four patients with decreasing abdominal pain and stool volumes. immunosuppression with steroids and mmf could slowly be tapered. all patients are alive after a median follow-up of days ( - days) from diagnosis of acute stage gut gvhd. the most prominent side effect attributable to ruxolitinib was thrombocytopenia with a nadir in platelet counts after days of ruxolitinib treatment in / patients. platelets recovered within weeks after ruxolitinib was discontinued. neutropenia was observed in one patient with anc dropping o . /nl after days of ruxolitinib treatment. mild to moderate elevation of liver transaminases was observed in all four patients during ruxolitinib treatment. one patient developed imminent acute renal failure, another patient showed symptoms of hemolytic uraemic syndrome. however, due to the multimodal treatment of these critically ill patients, these complications could not clearly be attributed to ruxolitinib. ruxolitinib is potentially beneficial in severe acute enteral gvhd in children refractory to corticosteroids as well as second-line immunosuppressants. however, randomized trials are warranted to verify safety and efficacy of ruxolitinib in this patient cohort. disclosure of conflict of interest: none. steroid refractory acute gvhd is a major cause of mortality after allogeneic stem cell transplantation. until date, no agent or treatment strategy has demonstrated superior efficacy in this patient group. the dose and duration of steroid treatment is associated with several short and long-term side effects, therefore concepts facilitating rapid steroid taper may be beneficial. both ruxolitinib and ecp have been reported to be effective in treatment of steroid refractory (sr) agvhd. we analyzed data from consecutive adult patients who received ruxolitinib for sr agvhd between march and august in our institution overall, patients (male n = ; female n = ) with a median age of years (range: - ) were included. donors for allogeneic sct were msd (n = ), mud (n = ) and mmud (n = ). median time to gvhd onset after stem cell transplantation was days (range: - days). about patients had agvhd grade iii or iv (all with gi involvement), while patients had skin grade involvement. sr agvhd was diagnosed if agvhd manifestations were progressive after days or persistent and without improvement after days or no partial remission after days of treatment with mg/kg bw of systemic steroids. patients received additional ecp (n = ), if response to ruxolitinib was lacking or slow (n = ) or instead of ruxolitinib due to cytopenias (n = ). ruxolitinib was first-line treatment for sragvhd in patients ( %). median initial dose of ruxolitinib was mg (range: - mg) twice daily. steroids were tapered and stopped, even if agvhd was still active. primary end point was non-relapse mortality at months. secondary end point was response on day after initiation of ruxolitinib. response occurred relatively slowly, resulting in a day overall response rate of % (cr = , pr = ). however, a total nine patients ( %) attained a complete response (cr), five with ruxolitinib alone and four others in combination with ecp. about patients ( %) required dose reduction or interruption of ruxolitinib mainly due to cytopenias. after a median follow-up of days, patients are alive. causes of death were relapse of malignant disease (n = ), gvhd (n = ), infections (n = ) and other (n = ). median survival from diagnosis of sr agvhd was days for non-responders and days for responders ( figure , p = . ). in univariate analysis, non-response was associated with higher risk of nonrelapse mortality (rr; . , % ci: . - . , p = . ). ruxolitinib and ecp are two effective promising treatment options, which may be complementary in patients with sr agvhd. cytopenia is the most frequent side effect of ruxolitinib while infections remain the major cause of death. [p ] disclosure of conflict of interest: ayuk-therakos: honoraria; kröger: novartis: honoraria, research funding. steroid-refractory acute graft-versus-host disease (sr-agvhd) is associated with a dismal outcome. janus kinase (jak) / signaling has been shown to be instrumental in multiple steps leading to inflammation and tissue damage in gvhd. jak / inhibitor ruxolitinib was studied in the treatment of sr-gvhd by zeiser et al. (leukemia ) , and the overall response rate was reported to be . %. we have now studied ruxolitinib in the treatment of six adult patients with steroid-refractory, grade iii-iv, intestinal agvhd. all the patients were male. the median age of the patients was (range: - ) years. three of the patients were transplanted for aml, one for all, mds and mm each. all the patients had been given a myeloablative conditioning treatment (cytbi , treosulfan+fludarabine ). two patients had a sibling donor and four a matched unrelated donor. the graft was from peripheral blood in all the patients. gvhd prophylaxis consisted of cyclosporine and a short course of methotrexate, and in addition antithymocyte globulin in the unrelated donor setting and methylprednisolone in one sibling recipient. agvhd of the intestine manifested on days + , + , + , + , + and + with diarrhea. in two patients it was preceded by agvhd of the skin by and days, respectively. gi-biopsy showed acute gvhd of grade iii and of grade iv in three patients each. treatment of intestinal gvhd was started with methylprednisolone mg/ kg/day, tapering the dose to and mg/kg after doses each. gastroduodenoscopy and colonoscopy were performed at the onset of symptoms indicating intestinal gvhd. biopsy confirmed the diagnosis in all cases. because the diarrhea continued in spite of methylprednisolone treatment, ruxolitinib was started , , , , and days from the first day of diarrhea. the dose of ruxolitinib was mg × per day orally. four patients showed a clear response to ruxolinitib, normalization of bowel function, after , , and days from the start of ruxolitinib treatment. the healing of the intestinal lesions was verified by biopsy. two of these patients had received extracorporeal photopheresis simultaneously. two patients did not benefit from ruxolinitib treatment. one of them had continuous infectious complications and therefore ruxolitinib was only started after days from the start of diarrhea. the other patient died of fulminant diarrhea after weeks of ruxolitinib treatment. cmv reactivation was detected in three of the responders, and two of them had also polyoma virus cystitis. one patient developed a pulmonary aspergilloma, which is under control with drugs. corticosteroid-resistant gastrointestinal acute gvhd was treated in six patients, out of whom four showed a good response. disclosure of conflict of interest: none. although methotrexate (mtx) is commonly used in the prophylaxis of graft-versus-host disease (gvhd) after allogeneic hematopoietic stem cell transplantation (allo-hsct), some small studies have also reported its use in the treatment of chronic gvhd. the aim of this study was to evaluate the efficacy and safety profile of low-dose mtx for treatment of sclerodermatous chronic gvhd (sgvhd) after the failure of first and second line treatments. we retrospectively evaluated adult patients who received low-dose mtx as salvage treatment of sgvhd during the period elapsed between june and june in a tertiary referral university hospital in spain. there were ( %) males and ( %) females. the median age was years (range: - ). all had received an allo-hsct for hematologic malignancies. the median time from allo-hsct to sgvhd was days (range: - ). thirteen patients ( %) had presented previous acute skin gvdh. superficial skin lesions mimicking lichen planus (lichenoid gvhd) were diagnosed in ( %) patients, while lesions resembling lichen sclerosus, morphea or fasciitis (sgvhd) where seen in all ( %) patients. the total body surface area was affected by more than % in patients ( %). besides the skin, other organs/tissues involved were the eyes ( %), mouth ( %), nails ( %), lungs ( %), liver ( %) and gastrointestinal tract ( %). treatment lines prior to mtx administration were: prednisone (pdn) in patients ( %), phototherapy (pht) in ( %), cyclosporine (cya) in ( %), mycophenolate mofetil (mfm) in ( %), pht + pdn + cya in ( %), pdn + mfm + cya in ( %), extracorporeal photopheresis + pdn in ( %). the median time from sgvhd onset to mtx treatment was days (range: - days). mtx was administered subcutaneously in patients ( %) and orally in patients ( %). median dose of mtx was . mg/week (range: . - . mg/week) and median length of treatment was weeks (range: - weeks). in two patients ( %) early withdrawal of mtx occurred (one due to early death secondary to septic shock and other due to rapid disease progression). mtx-related toxicity occurred in three patients ( %): megaloblastic anemia, asymptomatic increase of liver enzymes and mucositis, respectively. response to mtx was evaluated in the patients ( %) s who did not suffer early mtx discontinuation. seventeen patients ( %) presented a partial response; of them, two are still under mtx treatment for and weeks, respectively. fourteen patients ( %) received pdn concomitantly to mtx (median dose mg/day, range: - ); year after mtx treatment, only four patients were receiving pdn (median dose mg/day, range - ). seven patients have finished mtx treatment without reappearance of symptoms, receiving only topical treatment with emollients, tacrolimus or corticoids for short periods. in four patients ( %) sgvhd progressed despite mtx administration. our data suggest that mtx is a safe, inexpensive and effective alternative for refractory sgvhd. its potential used in earlier phases of sgvhd deserves further investigation. disclosure of conflict of interest: none. severe chronic gvhd has a major influence on late morbidity and mortality after hematopoietic stem cells transplantation (hsct). ecp is a good approach to treat refractory-gvhd: leucocytes are obtained from peripheral blood by apheresis, incubated with -mop, irradiated and then infused to the patient where they undergo apoptosis and induce tolerance. it is a promising alternative that reduces doses of immunosuppressive therapy and their side effects in the treatment of gvhd. this study shows its efficacy in persistent refractory cgvhd. the procedure was applied to three patients (pts) aged , and years (two aml and one cml), sex ratio ( m/ f) who underwent allogeneic-hsct with myeloablative conditioning regimen based on chemotherapy alone from a peripheral blood stem cells with cd levels: . , . and . × /kg respectively. prophylaxis of gvhd combined ciclosporin and methotrexate in short cycle. severe extensive cgvhd (according to nih criteria) was observed in the three cases after an average delay of . months ( - ) with involvement of - organs (mouth, eyes, skin, liver, joints and lungs). all pts are refractory to three lines of immunosuppressive agents (ciclosporin-corticosteroids, mmf and imatinib), with an average of thrusts/pt ( - ) over an average period of months ( - ). ecp was performed under the open system or dissociated system (macopharma) for two sessions per week for weeks, one session per week for weeks, one session every weeks for weeks and one session per month for months. after a median period of months ( - ), an average of sessions/pt ( - ) was performed. in terms of tolerance, a red blood cell transfusion was required in one pt, spontaneously resolved lymphopenia was observed in another pt, and a poor venous approach led to the pause of a central catheterization in one pt. the month and -month evaluation according to the couriel response criteria shows a partial response observed as of the first month with net improvement especially on skin sclerotic features and joints retractions initially refractory to all therapeutic lines. this allowed gradual reduction doses of corticosteroids. pce is recommended in the curative treatment for refractory chronic gvhd from the second line. this encouraging study on a small series shows its efficacy in persistent and late refractory forms. it is nevertheless necessary to evaluate it on a larger number of pts. disclosure of conflict of interest: none. successful treatment of steroid-refractory acute gastrointestinal graft-versus-host-disease by fecal microbiota transplantation p neumeister steroid-refractory acute gastrointestinal (gi) graft-versus-host disease (agvhd) is a severe complication of allogeneic hematopoietic stem cell transplantation (allo-hsct) associated with a high mortality rate. loss of intestinal bacterial diversity is thought to be associated with severity of gi-agvhd and an impaired intestinal microbiota with reduced diversity is an independent predictive factor for mortality. fecal microbiota transplantation (fmt) is the application of a fecal suspension derived from a healthy donor into a patient's gi tract. it has been successfully applied in recurrent clostridium difficile associated diarrhea including patients who underwent allo-hsct. we report the complete resolution of lower gi-agvhd following colonoscopic fmts in three patients that had been refractory to - lines of immunosuppressive therapies. microbiota analysis by s rdna before fmts revealed a severely depleted microbiota in all patients. donors (different persons for each patient) were healthy adult subjects. repetitive ( - ) colonoscopic fmts were necessary to permanently establish the donor's microbiome. all patients responded clinically by reduction/normalisation of stoll volumes, stopping total parenteral nutrition and tapering of steroids. a possible causative relationship of fmt in the reversal of severe intestinal dysbiosis and subsequent resolution of gi-agvhd can therefore be hypothesized. the establishment of donors' microbiota and increase in bacterial richness was associated with disease control. no immediate procedure-related infections or other side effects were observed. besides restoration of an initially severely reduced microbial richness by fmts, response of gi-agvhd was sustained despite reduction and discontinuation of concomitant immunosuppressive treatments. restoration of dysbiosis by fmt might represent a promising novel therapeutic approach for refractory lower gi-agvhd. disclosure of conflict of interest: none. tear film proteomics reveals important differences between patients with chronic ocular gvhd and healthy controls k plattner , n gerber-hollbach , s moes , p jenoe , d goldblum and j halter ophthalmology, university hospital basel, ch- basel and proteomics core facility of the biozentrum, university of basel, ch- , basel chronic gvhd frequently involves the eyes, leads to important decrease of quality of life and may threaten visual capacity. sicca syndrome is one of the hallmark of ocular cgvhd. analysis of tear protein composition may help to identify biomarkers for early diagnosis and prognosis of ocular cgvhd. tear fluid of patients with ocular cgvhd were compared with healthy individuals in this single center study. results of the first patients are reported here. tryptic digests from schirmer strips were analyzed on an orbitrap mass analyzer. clinical examinations included slit lamp examination, fluorescein staining, schirmer test, break-up-time (but) and a quality of life questionnaire (osdi). outcome measures were differences and consistency of proteins in human tear fluid s between patients with ocular gvhd and healthy controls. statistical analysis was performed by one sample wilcoxon-tests, p-values o . was considered significant. ten patients (eight males, two females) with a median age of years (range: - ) were analyzed. all underwent pbsct, eight from an unrelated donor. cgvhd overall score was moderate in three and severe in seven. eye organ score was in six and in four patients. all patients had more than one organ manifestation of cgvhd. eight were under systemic immunosuppressive therapy at the time of analysis, two had topical treatments only. in total different proteins were detected in tears analyzed. compared to controls, were differentially expressed in ocular cgvhd. expression was highly significantly different in proteins. compared to controls, expression of proteins was at least -fold increased, representing different categories. among them, more than % of all proteins belong to one of three categories: cytoskeletal proteins, nucleic acid binding or structural proteins. albumin, cluster or keratin (keratin type i-iii) and cluster of pyruvate kinase were most highly overexpressed. expression of proteins was decreased to o to %, belonging to different protein classes. half of them belong to defense/immunity proteins, enzyme modulators, hydrolases, nucleic acid binding and carrier proteins. expression of lactotransferrin, proline-rich protein and prolactin-inducible protein was most profoundly decreased. compared to healthy controls, a high number of protein is found to be differentially expressed in tears in ocular cgvhd. among them high expressions are observed for proteins that may indicate disturbed integrity of ocular surface and leakage of conjunctival capillaries. most profoundly decreased proteins include proteins with important functions in host defense and immunomodulation. more detailed pathway analysis is necessary to identify biomarkers for ocular cgvhd. disclosure of conflict of interest: none. steroid-dependent chronic gvhd after allogeneic peripheral blood stem cell transplantation is a great problem. nonresponders to corticosteroid therapy are at high risk of mortality. we hypothesized that such patients could benefit from treatment strategy using in patients with primary severe autoimmune diseases like multiple sclerosis and crohn's disease. patient z., y.o. was diagnosed in july with myelomonocytic leukemia (jmml). initially he was treated with low-dose of cytarabine and epigenetic agents. in september , jmml progression was observed with leukocytosis, thrombocytopenia, splenomegaly. bone marrow aspirate showed . % monocytes and . % blast cells. splenectomy was performed in november due to refractoriness to blood components transfusions. in december unmanipulated haploidentical peripheral blood stem cell transplantation from mother with . × /kg cd + and . × /kg cd + was performed. the conditioning regimen was myeloablative including melphalan mg/m day − and treosulfan mg/m days − , − , − . no organ toxicity grade was observed. gvhd prophylaxis consisted of hatg mg/kg on days − , − , − , + , i.v. tacrolimus from d − and mmf mg/kg from d . engrafted was fast and prompt ( % donor) with wbc . × /l on d + , plts × /l on d + . acute gvhd of stage ii was observed in early posttransplant period and treated with steroids and tnf-α inhibitor (infliximab). patient also received five procedures of ecp. all attempts of immunosupression tapering failed and the patient was staying on high dose of tacrolimus, mmf and courses of steroids till october . in october , gvhd stage ii flare with blood eosinophilia occurred after another attempt of steroids withdrawal. clinical examination showed that the patient was in complete remission with full donor chimerism. mild response of gvhd to steroids was observed. in april and may patient received two doses of rituximab mg/m with no significant response. in order to restore naive immune system first course of chemotherapy with cyclophosphamide mg/m was performed in the end of may . no toxicity grade was observed. the patient recovered wbc . × /l on d + , plts × /l on d + . in the phase of hematological recovery he was mobilized with g-csf and two leukaphereses of pbscs were performed. in june our patient was transplanted with previously collected . × cd +/kg following nonmyeloablative regimen including cyclophosphamide mg/m on day − and fludarabine mg/kg, on days − , − , − . second dose of cyclophosphamide was not administered because of severe hyponatriemia with seizures due to the cpm administration. no other significant toxicity was observed. the patient did not require either blood product or i.v. antibiotics. doses of tacrolimus and mmf were picked on months late and no more steroids were given. the patient is well in cr with no signs of gvhd for months. we speculate that pbsc collection from patients under massive immunosupression underwent allogeneic transplant is difficult but feasible. the nonmyeloablative regimens in such group of patients could be well tolerated and ensure the restoration of naive recipient immune system. this option could be discussed as an attractive alternative for treating resistant gvhd in steroid resistant patients. disclosure of conflict of interest: none. severe acute gi-gvhd is a serious early complication of allotransplants, and still remains a clinical diagnosis.( ) endoscopic biopsies provide the best supportive evidence, but are invasive and morbid in patients who are already medically compromised. f-fdg pet/ct may be able to stratify patients who require endoscopy and biopsy. to evaluate the performance of fdg pet/ct in differentiating moderate to severe gi-gvhd from no or mild disease in pediatric patients with suspected gi-gvhd. retrospective chart review of all paediatric allo-transplant patients referred for f-fdg pet/ct with suspected gi-gvhd from to . clinical follow-up, endoscopy and biopsy findings were correlated with f-fdg pet/ct. regional suv parameters were extracted by placing rois around stomach, duodenum, distal ilium, caecum, ascending, transverse, descending colon, recto-sigmoid colon and rectum. regional, and average large and small bowel suv data were statistically compared between patients with no or mild git-gvhd vs moderate to severe disease. the clinical and biopsy-supported diagnosis of acute gi-gvhd was taken as the true positive diagnosis for acute gi-gvhd. roc curves were generated for whole bowel suvmax values. about scans in patients, median age of years ( mths to y), were performed at a median of days post bmt. there were stage , stage - and with no acute gi-gvhd. transverse colon suvmax was significantly higher in the stage - gi-gvhd compared to no or stage disease (mann-whitney-u-test p o . ). there was a non-significant trend for average large bowel suvmax to be higher in the stage - group than the no or stage disease group (mean suvmax . compared to . , p = . ). a cut off whole bowel suvmax . had a sensitivity of % and specificity of % for detecting moderate to severe gi-gvhd. f-fdg pet/ct is a feasible and potentially useful non-invasive tool in the diagnosis and monitoring of therapeutic efficacy in acute gi-gvhd ( ) . large bowel suvmax may be higher in patients with stage - gi-gvhd, and transverse colon suvmax could have the ability to differentiate children with no or stage gi-gvhd from those with stage - disease. a negative fdg-pet/ct could serve as a criteria to avoid invasive endoscopic procedures and observe for the persistence of gastrointestinal symptoms before subjecting these patients to an imageguided biopsy. in patients too unwell for endoscopy, suvmax (roc curve specificity %) and a high suvmax in the transverse colon could serve as supportive evidence for moderate to severe acute gvhd, in the absence of biopsy findings. a major advantage of a pre-endoscopic f-fdg pet/ ct is to guide the procedurals to sample areas with the best diagnostic yield. prospective controlled studies are needed. oral mucosal progenitor cells (omlp-pcs) possess immunomodulatory and antibacterial properties, suggestive of their in vivo function in healthy tissue and their potential contribution to scarless wound healing in the buccal mucosa ( , ). our aim was to establish whether the function of oral stromal progenitors is impaired in chronic graft versus host disease (cgvhd) and restored with response to treatment. a patient with grade oral cgvhd was treated with systemic thalidomide for weeks ( mg/day). punch biopsies of buccal mucosa were taken before and after treatment. oral progenitor cells were isolated and expanded in vitro. numbers of progenitors was assessed using colony forming unitfibroblast (cfu-f) assays. stem cell markers (cd , cd , cd , cd , cd , cd , hla i and ii) were evaluated by flow cytometry. wound healing and antibacterial potential were assessed using a collagen gel lattice assay and bacterial cocultures as previously described ( , ) . secreted levels of relevant cyto-and chemokines associated with wound healing were assessed by elisa. significant clinical improvement with reduced inflammation in the oral mucosa and healing of ulcers was seen after weeks of thalidomide treatment, with continued improvement after weeks. cell surface expression of cd and cd on omlp-pcs was elevated postthalidomide; markers correlated with stemness and angiogenesis in mesenchymal stromal cells. this correlated with a restoration of wound healing potential and antibacterial function after thalidomide treatment ( figure ). figure : antibacterial testing demonstrated a loss of antibacterial function against (a) gram positive and (b) gram negative micro-organisms in the cgvhd omlp-pcs that could be completely or partially restored to levels comparable with healthy controls after thalidomide treatment. *p ⩽ . , **p ⩽ . , ***p ⩽ . . we demonstrate, for the first time a correlation between clinical improvement of oral cgvhd with thalidomide treatment and restoration of endogenous progenitor cell function. this study highlights the importance of a dysfunctional oral mucosal stroma in the pathogenesis of cgvhd. further studies should focus on the role of the stroma in promoting cgvhd and the precise mechanisms by which thalidomide is able to restore its functions. chronic graft-versus-host disease (cgvhd) is a major cause of late morbidity and treatment-related mortality in patients undergoing allogeneic hematopoietic stem cell transplantation (hsct). cgvhd is driven by a th biased t-cell mediated alloreactive immune response that leads to chronic inflammation and fibrosis in various organs. thymic stromal lymphopoietin (tslp) is an epithelial cell-derived cytokine that mainly affects myeloid cells. upon stimulation with tslp, dendritic cells are polarized towards a dc phenotype driving th biased immune response. we hypothesized that tslp is involved in the pathogenesis of cgvhd and that elevated levels of tslp post-transplant may lead to an increased risk of cgvhd. in the present study, we measured plasma tslp levels during hsct to study associations with clinical outcomes including cgvhd. about adult patients undergoing myeloablative hsct at rigshospitalet, denmark, from to were included. diagnoses included aml (n = ), all (n = ), myelodysplastic syndrome (n = ), other malignancies (n = ) and anemias (n = ). donors were either hla matching siblings (n = ) or mud (n = ). grafts were either bmsc (n = ) or pbsc (n = ). conditioning included tbi (n = ) or high-dose chemotherapy alone (n = ). plasma tslp was measured by elisa (abcam) before transplantation, at the day of transplantation and at day + , + , + and + post-hsct. monocytes were counted daily, and t, b and nk cells were measured at day + and + using flow cytometry. about ( %) patients engrafted; acute gvhd grade - was seen in ( %) patients, and ( %) patients developed severe cgvhd. oas was . %, trm . % and relapse rate . %. median plasma tslp levels increased from before conditioning ( pg/ml) to reach a peak at day + ( pg/ ml, p = . ), followed by a gradual decline. the plasma levels of tslp at day + were positively correlated with same-day monocyte counts (ρ = . , p = . ). approximately half of the patients (n = ) experienced an overall rise in tslp from baseline ( pg/ml) to day + ( pg/ml). this increase in tslp was not significantly associated with any transplantrelated baseline characteristics. however, patients, who had an increase in tslp levels from baseline to day + , had a significantly higher risk of extensive cgvhd compared to those in whom tslp levels at day + were similar or below baseline levels (cumulative incidence of cgvhd: % (increased tslp at day + ) vs % (normal/low tslp at day + ), p = . ). development of cgvhd was also associated with the nucleated cell dose infused (p = . ) and transplant using pbsc (p = . ). tslp plasma levels were not associated with acute gvhd, oas, trm, relapse rate or numbers of t cell, b cell or nk cells posttransplant. we have found that increased levels of tslp from baseline to day + were associated with an increased risk of extensive cgvhd. this association may be due to the ability of tslp to polarize the immune system toward a th response. importantly, the increase in plasma tslp levels was not associated with any transplant-related characteristics suggesting that tslp may be an independent predictor of cgvhd. these findings indicate that anti-tslp treatment may be a new approach to fight severe cgvhd. disclosure of conflict of interest: none. thymopoiesis following hct: a retrospective review comparing interventions for agvhd in a paediatric cohort c roberts , am flinn , ma slatter , , r skinner , h robson , j lawrence , j guest and ar gennery , institute of cellular medicine, newcastle university and great north children's hospital, newcastle-upon-tyne, uk acute graft-versus-host disease (agvhd) is a life threatening complication of allogeneic haematopoietic cell transplantation (hct), treated with topical and/or systemic corticosteroids. in steroid-refractory agvhd extracorporeal photopheresis (ecp) can be effective. ecp exposes apheresed mononuclear cells to -methoxypsoralen and ultra-violet radiation. systemic corticosteroids and agvhd are damaging to thymic tissue. delayed immune reconstitution, especially of the t lymphocyte compartment, is associated with increased morbidity and mortality. therefore, management strategies must be effective in treating agvhd but endeavour to minimise resulting thymic damage. we compare the effect of topical steroid therapy, corticosteroids and ecp on thymic reconstitution following hct in paediatric patients. statistical analysis was performed using the kruskal-wallis test. about paediatric allogeneic hcts were performed between june and april , at the great north children's hospital, newcastle for malignant and non-malignant disease. we reviewed computerised records to categorise patients into four groups: no agvhd, mild agvhd treated with topical steroid, agvhd treated with systemic steroid, agvhd treated with ecp. laboratory data were reviewed to provide values of naive (cd + and cd − )cd ra+cd + t-lymphocytes at , , and months post-hct. values for thymic output for the ecp group were additionally recorded at , , and months during ecp. excluded were patients with no available data, those with o months follow-up, those with chronic gvhd, recipient of hct or received dli post-hct. about patients were included, ( . %) had no agvhd, ( . %) had agvhd treated topically or systemically, ( . %) had agvhd and received ecp. for analysis, the group treated with steroids were divided into those treated with topical therapy and those given systemic steroids. the median values of all groups at each time point ( , , and months) are shown ( figure ). there was a significant difference between the rate of thymopoiesis (measured by the addition of cd + and cd − cd +cd + cells) between all groups (no agvhd, agvhd treated with topical or systemic steroids, and agvhd treated with ecp) at , , and months post-transplant (p = . , p o . , p o . , p = . respectively). further analysis excluded those treated with ecp (so including the no gvhd (n = ), topical treatment (n = ) and systemic steroid treatment group (n = )). at each time point p = . , p = . , p = . and p = . , respectively, demonstrating a statistically significant difference in time to thymopoiesis between those that had developed agvhd and those that had not. acute graft-versus-host disease (agvhd) is a significant cause of morbidity and mortality following allogeneic hematopoietic stem cell transplantation. which despite first line treatment is well-established (esteroids), second line is not well defined. evaluate the results with different second line treatment used and the risk factors associated with of sr-agvhd. we review the clinical records of consecutive patients undergoing allogenic hsct from to in our hospital. about % presented agvhd. sr-agvhd was defined as progression after days, no clinical change in days or incomplete response after days of treatment. about patients ( %) met s criteria for sr-agvhd. there were no significant differences between both groups (sr-agvhd vs no sr-agvhd) respect to age (recipient/donor), unrelated donor, prophylaxis of gvhd, cd lymphocyte and cd cell. the median time between transplantation and agvhd diagnosis was days ( - ). patients who did not respond on fifth day of steroid treatment have an % rate mortality vs % on no sr-agvhd group (p = . ). sr-agvhd: patients presented sr-agvhd and this was related to: hla mismatch ( % sr-agvhd vs % no sr-agvhd, p = . ), male recipient/female donor ( % sr-agvhd vs % no sr-agvhd, p = . ) and advanced underlying disease ( % sr-agvhd vs % no sr-agvhd, p = . ). second line: basiliximab ( . %); extracorporeal photopheresis (ep) ( . %), timoglobulin ( . %) and others therapies ( . %). two patients ( %) obtained complete response (cr) and patients ( %) partial response (pr). global response (cr, pr) after second line (mainly basiliximab) showed better overall survival (p = . ). third line: basiliximab ( . %); ep ( . %), mesenchymal cells (msc) ( . %), ruxolitinib ( , %) and others ( . %). ruxolitinib improve gvhd cutaneous and hepatic but not intestinal. the best results were achieved with ep ( cr, pr) and basiliximab/msc ( pr, respectively). only patients who achieved cr survived. fourth line: msc ( %)/ ruxolitinib ( %) does not improve the prognosis. no serious adverse effects were observed with msc therapy, basiliximab and ep. about % of patients showed cmv reactivation with basiliximab. about patients died ( %), patients with early mortality ( o months) due to refractory agvhd ( %) or secondary infections ( %). overall survival at months and year was ± % and %, respectively. in multivariate analysis the main factor for trm was the steroid-refractory vs steroidsensitive (hr . , % ci . - . ; p = . ) and was unfavorable the association of hepatic and intestinal agvhd (hr . , % ci . - . ; p = . ) no sr-agvhd: patients. trm- was % (n = ), mainly due to infection ( %). trm- year was % (n = ), mainly by gvhd ( %) and infections ( %). median follow-up of months, os- months and year were ± %/ ± %, respectively. trm was associated with not obtained cr/pr after second line (p = . ), no cr after third line (p = . ) and relapse of gvhd despite achieving cr initially (p = . ). in our series only the patients that obtained cr/pr after second-line or cr after third-line improved os. the best results in sr-agvhd were obtained with basiliximab and extracorporeal photopheresis. trm was associated with relapse of gvhd and advanced disease to the transplant. randomized clinical trials are needed to assess different treatment modalities for sr-agvhd. [p ] disclosure of conflict of interest: none. extracorporeal photopheresis (ecp) is a therapy for steroidrefractory chronic graft versus host disease (cgvhd). therapeutic response to ecp has been linked with a progressive increase in circulating granulocytic myeloid-derived suppressor cells (g-mdsc) in acute gvhd, but not in cgvhd . low density neutrophils (ldn) phenotypically resembling g-mdsc (putative g-mdsc) show marked flux in cgvhd patients receiving ecp, and a reduction in their frequency is associated with a sustained therapeutic response to ecp . recent data has identified lectin-type oxidized ldl receptor- (lox- ) as a specific marker of ldn with t-cell (tc) suppressive activity . using this marker we have conducted a cross-sectional study to assess whether putative g-mdscs in this patient cohort have suppressive activity. about patients with steroid refractory or steroid-dependent cgvhd (mean treatment duration of months) receiving ecp and healthy controls were recruited. patients had gvhd affecting skin ( / ), liver ( / ) and gut ( / ). pbmc were isolated and immunophenotyped by flow cytometry for markers of g-mdscs (cd − ve , cd , cd b, hla-dr − ve , cd int ) and lox- expression. suppressive function was determined by measuring the inhibition of proliferation of anti-cd /cd -activated purified cd tc from healthy donors by -day co-culture with g-mdscs from patients. statistical analysis was conducted using graphpad . ecp patients had substantially greater frequencies of circulating putative g-mdsc than healthy controls (p o . ; median: % and iqr %- % vs . % and iqr . %- . %, respectively). while there were substantially greater frequencies of circulating lox- + cells in pbmc from ecp patients than healthy controls (p o . ; median: . % and iqr . %- % vs . % and iqr . %- . %, respectively), these were mainly the minority population within the putative g-mdsc fraction with no significant difference between ecp patients and healthy controls in the proportion of lox- + cells ( % ± % vs % ± %, respectively). ecp had no significant effect on circulating putative g-mdsc frequency measured before and the day after treatment (median: . % and iqr %- % vs % and iqr %- %; n = , respectively) nor on lox- frequency (median: % and iqr . %- % vs . % and iqr . %- . %; n = , respectively). at a tc:g-mdsc ratio of : , isolated g-mdscs from ecp patients suppressed cd tc proliferation (mean ± sd: % ± %; n = ). however, the potency of suppression was highly variable (min-max: - %). the pattern of lox- expression suggests that only a subset of putative g-mdscs in ecp patients are suppressive and may explain why suppressive function in this cell fraction is so highly variable. however, the relatively high frequency of lox- cells in this patient cohort might contribute to immunosuppression resulting in increased susceptibility to opportunistic infections. according to the revised eortc/msg criterion, patients were diagnosed with cns-ifd. among those patients without cns-ifd ( patients), cns-ifd were matched in a : ratio for analyzing the risk factors of cns-ifd. and among ( . %) patients who occurred pulmonary ifd without cns involvement, patients were selected as control group for analyzing the risk factors associated with involvement of cns in pulmonary ifd. we selected the control group using a : ratio matched-pair method with the variates of ( ) age; ( ) sex; ( ) underlying disease. we retrospectively reviewed patients complicated with cns-ifd after hsct in our single center during a years period. most patients received haploidentical stem cell transplantation. the median onset time of cns-ifd was ( - ) after hsct, and most ( . %) of them have prior pulmonary ifd. the most frequent pathogen was aspergillus, while no crypoccosis and candidas were found. the most common clinical presentation was space-occupying symptoms and signs. brain abscess were the most common imaging finding. prior pulmonary ifd (po . , hr . ( % ci, . - . )) was the only risk factors associated with occurrence of cns-ifd. while poor response at weeks (p = . , hr . ( % confidence interval: . - . )) was the only risk factor predicting the involvement of cns in pulmonary ifd. the response (complete and partial response) at weeks and last follow-up was . % and . %, respectively. the overall survival was . % at the last follow-up with a median ( - ) days after transplantation. in conclusion, patients with pulmonary ifd had higher risk of cns-ifd, especially in those with poor response after weeks of treatment. and the prognosis of cns-ifd was very poor after hsct. disclosure of conflict of interest: none. adv may cause severe infections in hsct recipients, especially from unrelated donors or cord blood particularly in pediatrics. disseminated infections usually occur after digestive reactivations. at mo.post-hsct, the incidence of adv digestive infection and viremia in pediatric hsct is about % and %, respectively. therapeutic strategies to control adv infections are limited to the use of infusion of cidofovir (cdv) or ex vivo anti-adv selected cytotoxic lymphocytes (ctl). however cdv is not labeled for adv treatment, presents a renal toxicity and has shown limited efficacy. specific-ctl remain difficult to produce. brincidofovir (cmx , bcv, chimerix, usa) is an orally-available lipid conjugate of the nucleotide analog cdv that has demonstrated broad clinical antiviral activity against double-stranded dna viruses (that is, herpes-, adeno-, orthopox-and polyomaviruses. the drug has an increased bioavailability compared to cdv and has shown encouraging results. we report here the results obtained with this compound in patients treated in six centers from january . there were pts ( m/ f), median age at hsct: mo. ( - ). hsct indication was all in nine, pid in six, aml in two, fa in two and ibmf in one. donor was / or / mud in four and six pts, respectively; haplo-identical familial donor in ; / or / unrelated cb in two and three pts, respectively; msd in one. stem cell source was bm for pts, cb in and pbsc in . two pts underwent a second hsct. cond' regimen were mac in pts. all pts received either ex vivo or in vivo t-cell depletion. three pts presented with adv-disseminated disease, seven pts with blood + other site (throat, urine or stools) adv infection, three with adv-related gut disease, three with blood infection and three with gut infection. the remaining patient received bcv for jc viremia with fever. median time for virus infection diagnosis was d post-hsct (range: d- to d+ ). about pts experienced other viral infection episodes after hsct (cmv: ; ebv: ; bk: ; hsv: ; hhv : ; influenzae: ( ). about pts received - injections of cdv prior to bcv treatment. one pt received specific-adv ctl before bcv without efficacy. the reason to switch from cdv to bcv was uncontrolled adv infection (n = ) or cdvinduced renal failure (n = ). two additional pts experienced renal impairment after cdv. about pts received - lines of immunosuppressive therapies (including ecp) in addition to calcineurin inhibitor at time of bcv therapy due to grade iii and iv acute gvhd in seven and seven pts, respectively. median adv load at time of bcv initiation was . log copies/ ml (range: - ) in blood and log copies/ml ( . - . ) in stools. median duration for bcv therapy was weeks (range: - ). about seven pts with blood adv infection or disseminated disease experienced adv disappearance as well as four pts with gut disease or infection. three of them experienced adv infection relapse and received thereafter cdv, bcv or advspecific ctl. five pts presented with grade - diarrhea during bcv treatment. about were alive at end point where seven died from septis (n = ), multi-organ failure (n = ), gvhd (n = ) and adv disseminated infection (n = ). adenovirus infections occur often in immunocompromized pts receiving concomitant nephrotoxic drugs that may avoid cdv use.bcv appears as efficient therapy against adenovirus infection in such pediatric pts since here out of pts where alive after adv infection and bcv treatment. in this study we retrospectively analyzed cmv reactivation determined by pcr and response to pre-emptive therapy in patients receiving an haplo (n = , %) comparing them with a control group of non haplohsct ( mrd and mud) (n = , %). median age was years (range: - ), for haplohsct and for control group. conditioning regimen was myeloabaltive (mac) in . % and reduced intensity (ric) in . %. haplohsct characteristics: haplo conditioning was fludarabine ( mg/m or mg/m × days in ric or ma regimen) and busulfan ( . mg/kg × in ric or days in ma) ( . % mac, and . % ric). cyclophosphamide-post was used for gvhd prophylaxis in %. median of days to reach more than × granulocytes and more than × platelets were ( - ) and ( - ), respectively. incidence of acute gvhd was % (grade i-ii . %, and iii-iv . %), with two steroid-refractory cases. cmv reactivation: . % of haplohsct patients presented cmv reactivation, vs . % in control group (p = . ). median number of cmv reactivation episodes was in both groups. median time to cmv pcr detection was days ( - ) and ( - ) in haplohsct and control group respectively (p = . ). average maximum cmv iu by pcr was . in haplo vs . in the control group (p = . ). first antiviral pre-emptive therapy (valganciclovir in . %) was effective in % in haplohsct vs % in control group (p = . ). main reason for antiviral treatment switch was failure in cmv iu reduction, and foscarnet was the most used therapy in refractory cases. twenty patients developed cmv disease ( in haplo and in control group) (gi disease % and pulmonary disease % in both groups). in a multivariate cox-regression model, receiving an haplohsct, serological cmv status (positive patient/negative donor), mac regimen and development of acute gvhd grade i/ii or grade ii/iv were variables associated with a higher risk of cmv reactivation. based on these results, haplohsct is associated with a higher cmv reactivation compared to non-haplohsct, despite a lower incidence of all other risk factors as agvhd or mac in the haplo group. although it is not statistically significant, response to pre-emptive therapy is higher in haplohcst and no differences in cmv disease were observed. disclosure of conflict of interest: none. although a number of patients with hiv infection and hematological disease have successfully undergone allogeneic hsct together with combination anti-retroviral therapy (cart), short and long-term outcomes remain not well known. we report the spanish experience treating hiv-infected adult patients with high-risk hematological malignancies with allogeneic hsct. we retrospectively reviewed hiv-positive patients who received allogeneic hsct in three institutions in spain within geth (grupo español de trasplante hematopoyético y terapia cellular). seventeen patients have been transplanted between and . median age was ( - ), % male. diagnosis and transplant characteristics are summarized in table . cumulative incidence of neutrophil and platelet engraftment were % at days (median days), and % at days (median days), respectively. with a median follow-up of months ( - ), os and efs were %. trm was % at months and % at months. grade ii-iv agvhd rate was %, and moderate/severe cgvhd rate was %. all patients received cart. two patients showed severe toxicity related to interaction of immunosuppressive s drugs and protease inhibitors. about % of patients showed infectious complications. viral infections were the most frequent cause: cmv ( ), bk ( ), adv ( ), hhv- ( ), hcv ( ), hhv- ( ), parainfluenza ( ). two patients had invasive aspergillosis and one patient presented disseminated tuberculosis. causes of death were: relapse ( ), infection ( ), gvhd ( ) and toxicity ( ). all surviving patients showed undetectable hiv load after hsct. allogeneic hsct is an effective therapy for high-risk haematological malignancies in patients with hiv infection, and long-term hiv suppression with cart is feasible. however, interactions between immunosuppressive agents and anti-retroviral drugs, high rates of significant gvhd, and frequent infectious complications account for a complex procedure in this population. selected hiv-infected patients with hematologic malignancies should be considered for allo-hsct when indicated, in experienced centers. disclosure of conflict of interest: none. clostridium difficile infection (cdi) is one of the most common causes of nosocomial infectious diarrhea in europe and usa, which results in high morbidity and mortality among hospitalized patients. allogenic hematopoietic stem cell transplant (hsct) recipients remain at high risk for cdi. incidence rate ranges from to %. numerous risk factors including acute graft-versus-host disease (agvhd), hla matching status, conditioning-intensity, use of total body irradiation (tbi) may play an important role in the course of cdi in these patients. the aim of this study was to evaluate the prevalence of cdi in children, and to assess the influence of such factors as gender, age, diagnosis, hla matching status, conditioning-intensity, use tbi-containing regimen, source of graft (bone marrow/bm/ vs peripheral blood/pb/)or agvhd on course, duration of treatment and outcome in children undergoing hsct. between and a total of hscts were performed in five polish pediatric transplant centers, including allogeneic and autologous. all patients were followed up to months post hsct. we analyzed retrospectively episodes of cdi infection in the group of children. twenty-one of children were diagnosed with hematological malignancies: acute lymphoblastic leukemia (all), acute myeloblastic leukemia (aml), myelodysplastic syndrome (mds), two were diagnosed with severe aplastic anemia (saa), one with chronic granulomatous disease (cgd) and of them-with solid tumors. the median age was . years (range: . - . years). majority of patients underwent myeloablative conditioning protocol ( / ). in allogeneic setting / patients underwent mud-hsct, / pts msd-hsct and one patient was given a haploidentical pbsct. in this series, in out of cases bm was a transplant source, and pb in out of . cdi was defined as having diarrhea that tested positive for c. difficile via pcr, cytotoxin assay, or dual enzyme immunoassays. kruskal-wallis test, wilcoxon test and χ -test were used to estimate the influence of risk factors on severity of disease, duration of treatment and outcome. we observed episodes of cdi ( . %) in hsct recipients: in allotransplant recipients ( . % of all transplants) and in autotransplant recipients ( . % of all auto-hsct). nine patients responded to therapy with metronidazole, seven patients responded to vancomycin alone, and in two patients rifaximine was administered. six children required adding second drug: vancomycin or metronidazole, five patients were not given any medications. there was no significant correlation between such factors as diagnosis, gender, age, conditioning regimen, hla matching, agvhd and severity of disease, and duration of treatment. recurrence rate was difficult to assess due to lack of data. we observed three deaths. one of them was connected with cdi. there was one -year-old boy with saa (mud-pbsct, hla / ) with no agvhd. the other two deaths were due to progression of s disease. cdi occurred in nearly % of pediatric patients undergoing hsct, surprisingly often in autologous hsct too ( . %). almost all patients experienced mild cdi with adequate response to antibiotic therapy. cdi is a rare cause of death among transplant recipients. disclosure of conflict of interest: none. antifungal prophylaxis in high-risk paediatric patients with haematological malignancies: a monocentric experience k perruccio, i capolsini , a carotti , n albi , l pitzurra , a velardi and m caniglia pediatric haematology oncology section; haematology and clinical immunology section; transfusion medicine and the choice of antifungal prophylaxis in high risk paediatric haematological patients (according to the latest ecil /seifem guidelines) remains an open question. a recent retrospective survey from associazione italiana ematologia oncologia pediatrica (aieop) showed that, in these patient categories, the only variable which significantly impacted on invasive fungal infection (ifi) occurrence was the presence or not of antifungal prophylaxis at the ifi onset (unpublished data). from january , in our pediatric hematology oncology unit, allogeneic hematopoietic stem cell transplantations (hsct) were performed (median age: years; range: months- years), mainly for acute leukaemia (median follow-up: months; range: - ). patients received liposomal amphotericine b (n = ), micafungin (n = ), or fluconazole (n = ) as primary antifungal prophylaxis until neutrophil recovery ( × /l). seven patients developed acute gvhd ( %) which evolved in gvhd in ( %). as outpatients, they continued with posaconazole (n = ), voriconazole (n = ), or micafungin (n = ) until cd +t-cell recovery ( /cmm) or gvhd immune suppressive prophylaxis/treatment withdrawn. during the last year, according to ecil /seifem guidelines , we administered primary antifungal prophylaxis also to / high risk (hr) acute leukaemia patients. two patients with aml were treated with posaconazole, four patients with hr-all received micafungin, four relapsed all patients received micafungin (n = ), or liposomal amphotericine b (n = ), or posaconazole (n = ). one aml patient was then transplanted; all relapsed patients are waiting for transplant. no differences were observed in terms of breakthrough proven/probable (pp)-ifi incidence, according to antifungal prophylaxis in the various patient groups. in particular, in the early phase, we observed a pp-ifi incidence of % in both treatment arms (micafungin vs liposomal amphotericine b, p = ns). in the late phase, we observed case of pp-ifi who were receiving posaconazole as prophylaxis. overall survival (os) was %, with % mortality rate. in hr leukaemia patient group, we observed pp-ifi in the only two patients who were not receiving any antifungal prophylaxis at the ifi onset. antifungal prophylaxis is strongly recommended in paediatric patients with haematological malignancies who are at high risk of ifi. the choice of antifungal drug depends on the treatment phase, drug interactions (particularly for azoles), patient compliance and clinical conditions which interfere with intestinal absorption. in our experience, as no differences were observed in term of efficacy, micafungin resulted the best choice in terms of tolerability, toxicity, compliance and cost saving. antifungal prophylaxis with micafungin and bridging to inhaled liposomal amphotericin b after engraftment in patients undergoing allogeneic hematopoietic stem cell transplantation d rivera* , , c de ramon , a avendaño , j carrillo , d caballero , l lopez , i ormazabal , a navarro , a martin micafungin is an effective antifungal for prophylaxis, active against candida spp. including those resistant to other antifungals (c. glabrata and c. krusei) and also active against aspergillus spp. guidelines focused on antifungal prophylaxis, recommend its use during preengraftment and early postengraftment period in allogeneic hematopoietic stem cell transplant (allo-hsct) recipients. moreover, its profile of low drug interactions and side effects, makes it a suitable alternative for patients who need concomitant treatments, present hepatic insufficiency and in those who do not tolerate oral drug administration. the addition of inhaled liposomal amphotericin b (lamb) after engraftment, provides an alternative way to effectively prevent mold infections, that are acquired mainly by inhalation. inhaled lamb has good tolerance with absence of drug interactions and low toxicity. the aim of this study was to describe the experience in the hsct unit of the university hospital of salamanca with micafungin and lamb as primary prophylaxis in patients undergoing allo-hsct with reduced intensity conditioning (ric) and graft-versus-host disease (gvhd) prophylaxis with tacrolimus and sirolimus. thus evaluating efficacy and tolerability in our population. retrospective observational study from january to august , including all adult patients undergoing allo-hsct with ric and gvhd prophylaxis with tacrolimus and sirolimus, in whom an azole derivative is not indicated, due to drug interactions. therefore received prophylaxis with micafungin during the preengraftment period and bridging to lamb after engraftment at discharge, and continuing it during the first days post-transplant. data from patients from our hsct unit. ten ( . %) patients who had invasive fungal infection before undergoing allo-hsct, and ( . %) patients who received prophylaxis with drugs other than micafungin-lamb were excluded. underlying disease was grouped by leukemia in ( . %) patients, lymphoma in ( . %), myelodysplastic syndromes in ( %), multiple myeloma in ( . %) and other diseases in ( . %) patients. eighty patients underwent peripheral blood allo-hsct, of whom were related donor in ( %) patients and unrelated donor in ( %). prophylaxis with micafungin in ( . %) patients, dose of mg per day, with a mean of days (± days) with postengraftment bridging with lamb mg weekly, continuing it during the days posttransplant. days of neutropenia during preengraftment, o days in ( . %) patients, - days in ( . %), more than days in ( . %). during follow-up there were three cases ( . %), two catheter related candida infection, and one esophageal candidiasis. there were no reported aspergillosis cases (possible, probable or proven), according to the european organization for research and treatment of cancer (eortc) criteria. finally, prophylaxis with micafungin and inhaled lamb, was considered an effective and safe strategy in ( . %) patients, with no side effects reported. according to our experience with micafungin and the addition of inhaled liposomal amphotericin b, the results indicate that, this is an appropriate alternative for antifungal prophylaxis, in patients undergoing allo-hsct, because of their efficacy, few side effects and drug interactions. disclosure of conflict of interest: none. recipients of allogeneic hematopoietic cell transplantation (allohct) are at high risk of developing invasive fungal infections (ifi). in the early phase (o days) after allohct, the use of antimold prophylaxis has been generalized, although there is no consensus on the best therapeutic strategy. the use of nebulized liposomal amphotericin b and fluconazole has been shown to be effective, safe and associated with low economic costs in lung transplantation . however, the use of this prophylactic strategy in the early phase of allohct setting has not been evaluated. we included all consecutive patients who received their first allohct in our center from january to august and who underwent antifungal prophylaxis according to the prospective ambineb protocol (nebulized liposomal amphotericin b mg administered three times per week as loading dose and once per week and fluconazole mg per day until day + ). patients with a previous ifi were excluded. patients with graft-versus-host disease (gvhd) receiving high dose corticosteroids were allowed to be changed to voriconazole or posaconazole at physician's discretion. the primary objective of the study was the incidence of ifi at day + . the secondary objectives were to assess adherence and toxicity of the ambineb protocol. only cases with proven or probable ifi according to eortc-msg criteria were considered. a multidisciplinary team of experts in hematology, infectious diseases, microbiology and radiology prospectively evaluated and categorized each case. we included patients with a median age of years (range: - ) and a median follow-up for survivor of months (range: - ). patients received allohct mainly for acute leukemia ( %), non-hodgkin lymphoma ( %) and myelodysplastic syndrome ( %) . patients received allohct from hla unrelated ( %) or related donors ( %) mostly using a reduced intensity conditioning ( %). graft-versushost disease (gvhd) prophylaxis was performed with calcineurin inhibitors mainly in combination with sirolimus ( %) or methotrexate ( %). after the comprehensive review, only one case of proven or probable ifi at day + was diagnosed. prophylaxis with ambineb was completed in patients ( %) while ( %) stopped the treatment. the most frequent causes of discontinuation were possible ifi ( %), gvhd ( %), admission in intensive care unit ( %) and toxicity ( %) (figure) . ninety-four patients ( %) did not have adverse advents associated with ambineb. eight patients presented organ toxicity which was at least partially attributed to ambineb, including gastrointestinal symptoms (n = ), liver function test abnormalities related to fluconazole (n = ) and cough (n = ). of the patients who discontinued ambineb, ( %) were switched to other antifungal drugs including (echinocandins ( , %) posaconazole ( , %), voriconazole ( , %) or others ( , %)). overall survival and non-relapse mortality of all patients at median follow-up were % ( % ci - ) and % ( % ci - ), respectively. the combination of nebulized ambisome and fluconazole is effective in preventing ifi in the early phase of allo-hct and is associated with high adherence and low toxicity. neutropenia-related infections is a common complication of apsct in patients (pts) with mm and dlbcl. our study aims are to: ( ) assess antibacterial susceptibility patterns of isolated organisms, to guide antibiotic prophylaxis ( ) identify the epidemiology of bacteremia with susceptibility patterns to direct empiric therapy of febrile neutropenia ( ) assess the interval between the occurrence of neutropenia and the isolation of resistant bacteremia to identify the best timing to start prophylaxis ( ) identify contributing factors for the development of bacteremia and mortality. our retrospective study included adult pts who underwent apsct for mm and dlbcl between and . we recorded the following: age, gender, basic illness, comorbidities, number of cd + cells infused, a central venous catheter, duration of neutropenia, diarrhea and mucositis, mechanical ventilation, positive bacterial cultures with susceptibility profiles and history of broad-spectrum antibiotic intake for more than h for the past months on hospital setting, and mortality. statistical analysis was carried out using spss (version ). about isolates were obtained: urine ( . %), blood ( . %), sputum ( %), wound ( %), venous catheter ( %) and stool ( %). gram-negative (gn) species were predominant ( . %) with e. coli ( . %), klebsiella (k) ( %) and pseudomonas (pseudo) ( %). isolates sensitive to third generation cephalosporins ( gc) represented % of the enterobacteriaceae (entero) including % in e. coli and % in k. all entero isolated were susceptible to carbapenems (carba), pipercillin/ tazobactam (pip/taz), amikacin and ciprofloxacin (cipro). all pseudo (n = ) and acinetobacter (n = ) isolates were susceptible to carba, pip/taz, amikacin, cipro, colistin and tigecycline. as for gram-positive (gp) bacteria ( . %), coagulase negative staphylococci (cns) were predominant ( %) . oxacillin susceptibility reached % and two isolates methicillin resistant s. aureus were identified. all gp were susceptible to glycopeptides. a total of bacterial isolates were identified ( episodes of bacteremia) from pts. gn were predominant ( . %) with e. coli being most common ( %). all gn were susceptible to gc, carba, pip/taz, amikacin and cipro. as for gp ( . %), cns predominated ( %) including % oxacillin-susceptible causing seven episodes of bacteremia with six central line-associated. no glycopeptide resistance was identified. none of the clinical features and pts' characteristics reached statistical significance as risk factor for bacteremia. however, the need for mechanical ventilation and mortality were higher in bacteremic vs non-bacteremic pts ( . % vs %, p = . , and . % vs %, p = . , respectively). all bacteremic episodes occurred after developing neutropenia (median = . days, range: - ) except for one case of clabsi caused by e. coli occurring day before neutropenia. pip/taz was prescribed in % of bacteremia episodes followed by quinolones ( %) and carbapenems ( %). no previous use of third and fourth generation cephalosporins was observed. we recommend quinolone prophylaxis in apsct pts. for empiric therapy, antibiotics recommended by international guidelines including, cefepime and pip/taz still fit. thus, we could spare the use of carba and other last-resort antibiotics to other conditions. we also recommend continuous surveillance of resistance. disclosure of conflict of interest: none. fever is an almost universal complication in patients undergoing autologous stem cell transplantation (asct), however, microbiological documentation is only achieved in - % of such febrile episodes (fe). this low diagnostic efficiency makes epidemiological assessment in transplant units difficult, and may lead to a suboptimal empirical treatment. we have studied the utility of strict blood culture (bc) extraction as a mechanism to improve microbiological documentation of fe in these patients. we conducted a retrospective study over consecutive asct performed in our centre between june and may ( year). about patients were male and female, with age between and years (mean . ). diagnosis was hodgkin lymphoma, non hodgkin lymphoma and multiple myeloma. ascts were performed in reverse isolation conditions, in rooms equipped with hepa and pall filters. prophylaxis against herpes virus and p. jirovecii with acyclovir and pentamidine was used. no prophylaxis against gram negative bacteria or filamentous fungi was performed. bc were extracted at the beginning of every fe and every - h if fever persisted (or more frequently, following clinical criteria). blood samples from intravascular devices and peripheral blood were collected in two bactec bottles each (for aerobic and anaerobic microorganisms). complementary diagnostic techniques and empiric antibiotic therapy were performed following our institution's guidelines. fe were classified in microbiologically documented infections (mdi), clinically documented infection (cdi) and fever of unknown origin (fuo) following his criteria. fe were studied (average . fe per patient, . days of fever duration per fe). about % of fe were classified as mdi, % as cdi and % as fuo. mdis were cause by gram positive bacteria ( %), gram negative bacteria ( %) and polymicrobial infections ( %). no viral or fungal infections were observed. an average of . bc per fe and . per patient were extracted. the proportion of fe classified as mdi was related to the number of blood cultures extracted during the episode. only % of fe with three or less bc extracted were classified as mdi, % if - bc were extracted, and % if - bc were extracted (p o . ). no significant difference in proportion of mdi classified fe between the extraction of or more bc and - . all patients were discarded in good clinical conditions. according to our experience, a strict - blood culture extraction is related to a high rate of microbiological documentation of febrile episodes. moreover, we have not observed the rise in gram negative bacteria reported by other studies and gram positive cocci persist as the main infection cause in our centre. candida which has been traditionally related to duration of neutropenia, emerges as a pathogen beyond the aplastic period in allogeneic haematopoietic cell transplantation (allohct). in the setting of alternative transplants and aggressive immunosuppressive therapy, these infections are a challenging problem. there is scarcity of data regarding the significance of breakthrough candidaemia in allohct. to that end, we aimed to determine the incidence, clinical and microbiological characteristics and outcome of candidaemia in allohct recipients. we studied consecutive allohct recipients from january to june . blood cultures were obtained from peripheral vein or central venous catheters (cvcs) routinely and on febrile patients. well-known risk factors for candidaemia were studied: neutropenia, type of transplant, moderate to severe graft-versus-host disease (gvhd) and coexisting infections. among allohct recipients, we identified seven patients with candidaemia: five post matched unrelated (four myeloablative and one reduced intensity conditioning) and two post haploidentical transplant. in median time of . ( . - ) months, episodes of candidaemia were noted, despite antifungal prophylaxis with echinocandins or azoles. infections with non-albicans candida spp. occurred more frequently ( / ) and c. parapsilosis was the predominant microorganism ( / ). other species were isolated: c. famata ( ), c. krusei ( ) and c. haemulonii ( ). all candida spp. isolates were phenotypically susceptible to antifungal agents already administered to patients. there was no resistance to echinocandins indicated by minimum inhibitory concentrations (mics). all patients had severe acute or late-onset gvhd with intestinal involvement and cvcs prior to candidaemia. although cvcs were removed in / and patients were treated with echinocandins, new cvcs were re-contaminated in / with the same or other species. all patients presented well known risk factors for candidaemia (use of broad spectrum antibiotics due to severe bacterial infections, total parenteral nutrition due malnutrition, long-term high-dose corticosteroids and other immunosuppression), but no neutropenia. one patient survived, whereas five patients succumbed to gvhd and multi-organ failure and one patient to sepsis due to bacteremia. candidaemia was observed in non-neutropenic patients with agvhd and cvcs on antifungal prophylaxis, despite difficulties in diagnosis due to poor sensitivity of blood cultures. the epidemiology of candidaemia has changed in the last decade and its risk is more diverse and complex. the irreversible intestinal gvhd lesions might be the main source of candida in patients receiving antifungal prophylaxis. our data show that candidemia remains an important issue in profoundly immunosuppressed patients contributing to excessive morbidity. our aim was to compare the rate of neutropenic sepsis, defined as fever of o c and a neutrophil count of o . × /l, before and after the introduction of ciprofloxacin prophylaxis. one hundred and eight adult patients, of which had acute myeloid leukaemia, had acute lymphoblastic leukaemia, had lymphoma and had other haematological malignancies, were identified through our admission database. of these patients, received oral ciprofloxacin during their neutropenic phase. the median duration of neutropenia was days in both the no-prophylaxis and ciprofloxacin groups. there was a significant reduction in the rate of neutropenic sepsis from . % ( / ) in the no-prophylaxis group to . % ( / ) in the ciprofloxacin group (p = . ). prolonged infection, suggested by the use of broad-spectrum antibiotic treatment for more than days, was more common in the group which did not receive prior ciprofloxacin prophylaxis ( . % vs . %, p = . ). rate of intensive care admission ( . % vs . %, p = . ) was also reduced by the use of ciprofloxacin. however, there was no significant difference in the length of stay (mean of vs days, p = . ), or in the -day infection-related readmission rate ( . % vs . %, p = . ) between the two groups. in terms of the cause of neutropenic sepsis, escherichia coli, klebsiella pneumoniae and pseudomonas aeruginosa were the most common bacteria isolated from cultures in the no-prophylaxis group. eighty percent of these organisms showed sensitivity to ciprofloxacin. in the ciprofloxacin group, staphylococcus epidermidis was the most frequently found bacteria. with regards to treatment related adverse effects, none of the patients who received ciprofloxacin prophylaxis developed clostridium difficile diarrhoea. in conclusion, ciprofloxacin is still an effective antibacterial prophylaxis during neutropenia following allogeneic stem cell transplantation. clinicians should have a high suspicion of a gram-positive infection in patients who develop neutropenic sepsis on ciprofloxacin prophylaxis. disclosure of conflict of interest: none. s hematopoietic stem cell transplantation (haplohsct) to cure leukemia, malignancy and some inherited diseases, different additional reasons interfere microbiota metabolism and integrity. among them are radiation and chemotherapy, mucositis, infection and graft versus host disease (gvhd). the curative mechanism of fmt is based on the ability of donor intestinal microbiota to substitute and to provide all necessary functions of altered patient's microbiota. three patients ( , and years old) after haplohsct, who observed pseudomembranous colitis (toxin b-positive) as gvhd of intestine outcome, were enrolled to the study and performed fmt. relatives (mother, father and brother) were used as microbiota donors. donor and patient examination have included routine clinical and biochemistry laboratory data, microbiota cultural methods, pcr of most common intestinal microorganisms. additional for patient-level of fecal calprotectin by elisa was tested, identification of drug resistant bacteria and histology of intestine were made. patient's preparation for fmt included-probiotic (inulin) administration h prior procedure, discontinuation of all antibiotics h prior procedure and antiemetics ( ht agonist), prokinetics and proton pump inhibitor. delivery of donor's microbiota was performed in two consecutive steps under total intravenous anesthesia: with esophagogastroduodenoscopy-to the duodenum; with colonoscopy-to the caecum. all patients observed complete clinical response in - days after fmt (table ). in days we have revealed significant quantitative and qualitative changes in microbiota composition, which was matched to donor's microbiota. in days after fmt we identify microbiota changes in oropharynx and urogenital tract similar to donor microbiota. this leads to substitution of multidrug resistant klebsiella pneumoniae strains by drug sensitive microorganisms and helps to treat severe infection complications after haplohsct. platelet aphereses were carried out in donors ( males and females with median age . years) using haemonetics instrument with simultaneous leucoreduction. quantitative detection of cmv, ebv and hhv- dna was performed by multiplex real-time quantitative pcr kit (interlabservice, russia) in donors' whole pb, plasma and platelet aphereses at the time of platelet collections. viral load in hsct recipients was monitored weekly after hsct and days before hsct by the same pcr kit. lower limit of detection (llod) of the applied kit for all viruses was copies/ml. in specimens of platelet donors we additionally performed ultra-sensitive pcr with llod copies/ml. only one patient ( . %) was cmv-positive by pcr prior to hsct. cmv reactivation after hsct ⩾ copies/ml was noted in whole pb of patients ( . %) with median time of days (range: - ). donor source in cmv-reactivated patients was as follows ( mud, msd, haplo). cmv viremia ⩾ copies per ml was detected in seven patients ( . %). cmv disease was found in five cases ( . %). none of patients were positive by pcr for ebv or hhv- prior to hsct. ebv reactivation ⩾ copies/ml was found in six cases ( . %), ⩾ copies/ml in ( . %) with median time of days (range: - ). no signs of ptld or other ebv-dependent clinical symptoms were observed. hhv- level after hsct ⩾ copies/ml was detected in patients ( . %), ⩾ copies/ml in ones ( . %) with median time of reactivation days (range: - ). hhv- disease was observed in one patient. none of platelet donors were cmv-positive in plasma, whole blood or platelet aphereses products. ebv ⩾ copies/ml was detected in whole pb specimens of five platelet donors ( . %). application of ultra-sensitive pcr revealed low level of ebv-viremia in additional pb cases with median ebv level copies/ml (range: - ). none of platelet donors have any clinical signs of ebv disease. there is no any ebv-positive case among platelet concentrate specimens. in two cases low levels of hhv- was found in a whole pb ( and copies/ ml). none of hhv- -positive case was observed among plasma and platelet concentrate specimens. despite high incidence of cmv, ebv and hhv- reactivation after hsct in pediatric patients we could not show that source of viral reactivation was contamination of platelet apheresis products by donorderived herpes viruses. disclosure of conflict of interest: none. conventional respiratory virus (crv) infections are known to be major causes of morbi-mortality after stem cell transplantation (sct) due to the increased risk of progression to lower respiratory tract infection (lrti) in this setting. risk of developing severe lrti is mostly related to factors specific to the patient and the underlying disease, although the intrinsic virulence of crvs may also determine their outcomes. we conducted a single-center retrospective study including all adult sct recipients who had crv disease (defined as patients with symptomatic respiratory disease and crv identification) during a -year period ( - ) with the main objective of evaluating epidemiological changes over time and their association with infection outcomes. during the study period episodes of crv disease were diagnosed in patients (median age: years, % male, % aml or mds as baseline disease). patients ( %) received an allogeneic-sct (allo-sct) ( % had a prior sct) and ( %) an autologous sct. crv disease was diagnosed at a median of days after sct (range: - ), with cases ( %) occurring before day + . during the infectious episode of allo-sct recipients ( %) had active gvhd and ( %) were under s prednisone (pdn) mg/kg. most of the patients ( %) had symptoms compatible with an upper respiratory tract infection (urti), with of them ( %) progressing to a lrti, while ( %) had a lrti only. hospital admission was required in episodes ( %) with a median duration of hospitalization of days (range: - ), % required supplemental oxygen, % were transferred to the intensive care unit and % required mechanical ventilation. the most commonly identified pathogens over time are shown in figure . twenty-four cases ( %) had concomitant bacterial or fungal infections. influenza a virus was the most frequent crv detected ( episodes, %) followed by human respiratory syncytial virus ( episodes ( %) and human parainfluenza virus type ( episodes, %). during the flu pandemic, only of the crv infections diagnosed in sct recipients ( %) were associated to influenza a virus h n . antiviral treatment was started in episodes ( %), antibiotics in % and combined therapy in % during a median of days (range: - ). the rv resolved in cases ( %) at a median of days ( - ) from onset, with crv being considered the leading cause of death in only patients ( % of all cases and / ( %) in those with a lrti). predictors of severe crv infection (including icu admission, need for supplemental oxygen, need for mechanical ventilation requirement or death) in multivariate analysis were lymphocyte count o cells/μl (hr: . , % ci: - , p = . ) and co-infection with other pathogens (hr: . , ci %: . - , p = . ). no specific crv nor period post-sct of the infection influenced the risk of severe infection. our results confirm that crv infections are a frequent cause of morbidity after sct with a high need for hospital-based care. temporal changes in the principal circulating crvs has been identified during the -year study period, with influenza a virus being the most common. profound lymphocytopenia and presence of co-pathogens are associated with infection severity. [p ] disclosure of conflict of interest: none. the consecutive hsct performed from to are being analysed retrospectively. out of them ( %) performed in hepa filter room and ( %) in non-hepa filter room, criterion was purely financial to make this decision. / ( %) were allogeneic and / ( %) were autologous hscts. blood cultures both bacterial and fungal were taken at onset of fever and with every change of antibiotics till patient became afebrile. chest x-ray and if required hrct chest was done for all patients who had respiratory complaints. we did not use antibacterial prophylaxis; however, antifungal prophylaxis was administered along with conditioning; and at the onset of fever systemic antibiotics were started. antifungal agents were added if fever persisted for days pre empatively. extremely well trained nurses were looking after both the groups. all treatment protocols, antibiotic/antifungal policies were same in both the groups. median time for neutrophil engraftment was days in hepa filter room and days in non-hepa filter room. total / ( %) patients did not engraft till days. out of them / ( %) were in hepa filter room and / ( . %) in non hepa filter room. blood cultures were positive in total / ( %) patients, were positive for bacterial and for fungal organisms. in hepa filter hsct / ( %) were positive and in non-hepa filter hsct were / ( %) positive. total / ( %) patients developed pneumonia, out of them / ( %) were in hepa filter and / ( %) hsct were in non-hepa filter room. statistically not significant. no central venous access cather issues or infections were documented in any groups gr - agvhd : hepa rooms / ( . %),non hepa rooms / ( . %) :was not statistically significant the -day mortality was / ( %), / ( %) patients were from hepa filter rooms and / ( %) were from non-hepa filter rooms. cost : average cost of allogeneic hsct in hepa room : usd . average cost of allogeneic hsct in non hepa room: usd . average cost of auto hsct in hepa room: usd . average cost of auto hsct in non hepa room : usd . incidence of blood culture positivity & incidence of pneumonia was not different. these are two very important issues in outcome of hsct. agvhd incidence did not depend on the room type. these are significant findings from this study. results were slightly better in hepa filter rooms compare to non-hepa filter rooms, which was statistically insignificant. our study had few confounding factors hence we could not be concluded that hepa-filtered rooms are not necessary. nevertheless, our experience suggests that availability of dedicated hepa units with special air-handling equipment should not be considered a critical and essential precondition for providing allogeneic hsct to patients even in developing world with financial constraints. these would otherwise succumb to potentially curable hematological illnesses with background of financial constraints and wait list of hepa filter rooms. early hsct in a clean patient in non hepa rooms is extremely cost effective with comparable outcomes. nursing care, experience of the team, experience in hsct program & well established protocols are more important in outcome of hscts. disclosure of conflict of interest: none. we present five cases of cytomegalovirus (cmv) pneumonitis occurring in patients after recent allogeneic stem cell transplantation (allohsct). these cases were complicated by an organising pneumonia (during the recovery period) with a predominantly central peribronchial pattern. all patients presented with evidence of active cmv pneumonitis which was treated successfully with anti-viral therapy but was followed by persistent severe dyspnoea, cough and hypoxia. high resolution computed tomography (hrct) imaging showed widespread central peribronchial consolidation with traction bronchiectasis. in most cases there was a marked clinical and physiological improvement after treatment with systemic corticosteroids. however, in all patients the lung function remained abnormal and in some cases imaging revealed a fibrosing lung disease. these cases represent a previously undescribed central peribronchial pattern of organising pneumonia complicating cmv pneumonitis that can result in chronic lung damage. disclosure of conflict of interest: none. cytomegalovirus reactivation in pediatric acute leukemia after stem cell transplantation has an effect on relapse and survival in aml but not in b-precursor all j-s kühl , l sparkuhl and s voigt department of pediatric oncology/hematology/sct, charité universitätsmedizin berlin, berlin, germany several studies have indicated better survival after stem cell transplantation (sct) for acute leukemias, especially acute myeloid leukemia (aml), in case of cytomegalovirus (cmv) reactivation. here, we investigated if cmv reactivation had an impact on survival after sct for aml or acute lymphoid leukemia (all) in children. pediatric allogeneic stem cell transplant recipients from our institution who received myeloablative conditioning were included. transplant indications included aml, t-all and b-precursor all. cmv reactivation was correlated with relapse, mortality as well as acute graft-versus-host disease (gvhd) and was analyzed by fisher's exact test or χ -test (if n ). from the patients included, were transplanted for aml ( %), for t-all ( %), and for b-precursor all ( %). mortality and relapse rates ( - % and - %, respectively), cmv reactivation rates ( - %) as well as numbers of negative cmv serology status ( - %) of donor and recipient were comparable between different acute leukemias. when patients were analyzed altogether, cmv reactivation had no effect on relapse rates or mortality. however, a tendency towards fewer relapses after cmv reactivation was observed in aml patients (no relapse ( %) with cmv reactivation vs relapse cases ( %) without cmv reactivation; p = . ). in those leukemia patients capable of reactivating cmv (that is, donor or recipient cmv seropositive prior to sct), cmv reactivation had a protective effect on relapse rates in aml (no relapse ( %) with cmv reactivation vs relapse cases ( %) without cmv reactivation; p = . ). a similar tendency could be seen in t-all whereas no effect in patients with b-precursor all was documented. numbers of acute gvhd cases grade i between aml and t-all with or without cmv reactivation were similar. different effects of cmv on relapse rates and mortality in aml vs b-precursor all were noticed in patients who were either not capable of cmv reactivation or who did reactivate cmv post sct. in aml patients, there were no relapses ( %) and deaths ( %) in contrast to relapse cases ( %) and deaths ( %) in children with b-precursor all (p = . and p = . , resp.). latently cmv infected aml patients without documented cmv reactivation after sct have a significant worse prognosis compared with all other aml patients. this is also likely to be the case in patients with t-all, however, patient numbers in our cohort were too few. the protective effect of cmv reactivation in aml and possibly t-all does not appear to be gvh-related since the rate of relevant acute gvhd cases was comparable. cmv reactivation after sct for b-precursor all lacks significance. disclosure of conflict of interest: none. infections are among the most frequent and relevant complications of hematopoietic stem cell transplantation (hsct). little is known about the role of dental foci for the prevalence of infections in hsct. dental status was prospectively evaluated in all patients at our center before undergoing hsct. a total of different patients before undergoing hsct ( allogeneic and autologous), with a median age of years (range: - years) were evaluated. for evaluation a panoramic x-ray evaluation was performed. dental findings included the status of third molars and root fillings as well as caries, periodontitis, destructed teeth and apical bone loss. as non-dental parameters we used age, sex, type and status of central venous line, mucositis, and type of transplantation. these were correlated with neutropenic fever, bacteremia and pneumonia in a bivariate manner before a multivariate analysis was performed. no correlation of initial dental status to neutropenic fever, bacteremia or pneumonia was found. however, bacteremia and suspected infection of central venous lines was a significant predictor of neutropenic fever. in conclusion, dental surgery should only be performed prior to hsct if urgently required and limited to those individuals with overt infection. disclosure of conflict of interest: none. [p ] early experience with clinimacs prodigy ccs method in generation of virus-specific t-cells for pediatric patients with severe viral infections after hematopoietic stem cell transplantation k kallay viral reactivation especially in children is a frequent complication of allogeneic hematopoietic stem cell transplantation. most of these episodes can effectively be controlled by an antiviral or antibody therapy; in refractory cases a novel virusspecific t-cell therapy could be a promising management option. in our pediatric cohort of allogeneic transplantation during year patients fulfilled criteria for virus-specific t-cell therapy ( boys, girls, median age of ( . - ) years). six patients were transplanted because of hematological malignancies and for inborn errors. donor distribution was the following: matched unrelated, sibling and haploidentical donor. in cases bone marrow, cases peripheral blood and case cord blood was used as a stem cell source. the underlying viral illness was cmv in , ebv in and adenovirus in case, while more than one virus was detected in cases (cmv+adenovirus cases, cmv+ebv cases). viral diseases necessitating a t-cell therapy were cmv pneumonitis and colitis, adenovirus enteritis and cystitis and ptld. patients initially received cidofovir for adenovirus, rituximab for ebv and a combination of gancyclovir and foscarnet for cmv infections. the indication for t-cell therapy was progressive viral disease in of the cases and uncontrollable viral load in case. the procedure was performed on a median of ( - ) day post transplant. donors were st degree relatives in cases, nd degree relatives in cases and an unrelated person in case, the best hla match was haploidentical. the median age of the donors was ( - ) years. cells were produced by the clinimacs prodigy cytokine capture system (ccs) method after mononuclear leukapheresis. the system produced a median of . ( . - ) times /kg cd + and a median of . ( - . ) times /kg cd + interferon producing cells while the non-interferon producing cells were far below gvhd limit with a median of . ( . - . ) times /kg cd + and a median of . ( . - . ) times /kg cd + cells. the t-cell products were administered uneventfully in all but one case. we observed a manageable cytokine storm in one patient. glucocorticoid treatment was ongoing due to acute gvhd in children; however we could manage to keep the steroid dose below mg/kg in all cases. eight patients became completely asymptomatic, while also cleared the virus. we experienced decreasing viral load in all cases, the first negative viral results were achieved on a median day of ( - ). six patients are alive without viral illness or sequale, and complete viral dna clearance in peripheral blood with a median follow up of ( - ) days. one patient with cmv pneumonitis improved during the first week but deteriorated on the second week and died of respiratory insufficiency despite of mechanical ventilation. in cases the viral illness improved or cleared, but the patients died of invasive aspergillosis. no cases of gvhd, rejection, organ toxicity or recurrent infection were noticed. virus-specific t-cell therapy produced by the clinimacs prodigy ccs is a feasible, fast, safe and effective way to control resistant viral diseases after pediatric hematopoietic stem cell transplantation. this treatment can be implemented within a week in most cases. in order to define the appropriate place of this approach for patients with viral reactivations more data should be collected. disclosure of conflict of interest: none. central venous catheter (cvc) is essential for the treatment of recipients of stem-cell transplant. it is usually placed for the administration of conditioning regimen, stem cell infusion, intravenous antibiotics, immunoglobulins, electrolyte and nutritional support and blood concentrates. this patient group is at high risk for catheter-related bloodstream infections that can result in substantial morbidity and mortality. the neutropenia secondary to the conditioning regimen determines the risk of catheter-related infections, which may serve as an entry into the blood circulation, leading to bacteremia, fungemia, and consequently to septic shock and death. the risks of infection and the spectrum of infectious syndromes differ according to the type of transplant, conditioning regimen, type of implant of stem cells and therapies used after the procedure. gram-positive bacteria, particularly coagulase-negative staphylococcus spp, remain the leading cause of catheter-related bloodstream infection, although an increase in gram-negative bacteria as the causative agent has been noted. aim of the study: to evaluate the impact of the early cvc removal on the frequency of febrile episodes and infections in our group of patients. during a years period we have treated patients with hematologic neoplasm with high-dose chemotherapy and stem cells transplantation. patients were treated in sterile room conditioned with hepa filtration. in every patient was introduced double-lumen cvc ( subclavia, jugular, and femoral). % were febrile ( % fuo), catheter-related infection was present in %, while positive culture from cvc was present in %. the most frequent isolated bacteria from cvc were gram positive-staphylococcus coagulasa negative. the catheter was removed on the day of discharge. trm is . %. from january to november we have transplanted additional patients. to aim to decrease infection related mortality we perform strategy to remove cvc on day + after stem cell transplantation. the febrile episodes decreased on % ( / ), there were no early post-transplant mortality due to infection. early removal of the cvc and adequate handling from the nursing staff is essential for outcome of this patient population in regard of infective complications efficient prevention, early diagnosis, and effective treatment of catheter related infection are essential to providing the best care to these patients and can minimized morbidity and mortality. disclosure of conflict of interest: none. fever in patients with agranulocytosis during autologous hematopoietic stem cell transplantation (autohsct) can be associated with non-infectious causes due to g-csf, vancomycin, engraftment syndrome. in this case biochemical markers, such as presepsin (psp), procalcitonin (pct) and c-reactive protein (c-rp), can help in differential diagnosis of fever of infectious and non-infectious genesis. psp, pct and c-rp were assessed on the day of admission to the hospital (da), on d+ , on d+ , on d+ and on the day of discharge (dd). if patients developed neutropenic fever (nf), the markers were assessed at the beginning of the fever, h after, then on the second, third, fourth days after. if patients developed nf immediate empirical antibiotic therapy (at) was implemented with meropenem. in cases of ineffective st line ab, nd line at was added or totally changed. there were patients included in the study: patients with hodgkin lymphoma, with non hodgkin's lymphoma, with multiple myeloma, out of patients there were women and men. the median age was years ( - years). the conditioning regimens were cbv, beeac or hd melphalan. patients developed infectious complications (ic): of them had sepsis and others-nf. the median of nf development was . days. depending on the efficiency of at therapy patients were divided into two groups: group : patients that have had effective at (they 've had fast clinical response and they haven't needed to change medicine (n = )); group : patients that have had ineffective st line at, they haven't had response to st line at and they've needed to change another at (n = )). there were significant differences in psp levels on the third day after ab had been admitted: . pg/ml in group and . pg/ml in group (p = . ). similar differences between the analyzed groups were observed on the fourth day: . and . pg/ml, respectively (p = . ). pct and c-rp didn't show any significant changes between group and on each day of the study (table ) . disclosure of conflict of interest: none. enterovirus related immune reconstitution inflammatory syndrome (iris) following haploidentical stem cell transplantation in an mhc class ii deficient child r shah , s waugh , k foong ng , z nademi , t flood , m abinun , s hambleton , a gennery , m slatter and a cant paediatric immunology and bmt, great north children's hospital, newcastle upon tyne, uk and department of virology, great north children's hospital, newcastle upon tyne, uk immune reconstitution inflammatory syndrome (iris) has been described after hsct in association with fungal, viral and bcg infections. we describe a case of post-hsct iris associated with enterovirus infection. case: a girl with mhc ii deficiency (rfxp c. mutation) underwent treosulfan/fludarabine/ thiotepa/atg conditioned tcrαβcd + depleted haploidentical hsct at . years of age. pre-transplant work up did not reveal any viral or fungal infections except norovirus in stool. cyclosporine (csa) was given as gvhd prophylaxis. neutrophil and platelet engraftment occurred on d+ and d+ , respectively. on d+ , her stool was tested positive for enterovirus (taqman pcr), however; she was asymptomatic. the child started having fevers and irritability from d+ which persisted despite the use of antimicrobials. no evidence of fungal or bacterial infection was found. enterovirus pcr in blood was found positive on d+ (cycle threshold value, ct . ) and further typing showed it to be echovirus . at this time, symptoms progressed with diarrhoea, developmental regression and signs of radiculopathy. mri (brain and spine) was normal and csf showed pleocytosis ( wbc/mcl- % lymphocytes, protein . g/l) with positive enterovirus pcr (ct ). subsequently, immunoglobulin prophylaxis was increased to . g/kg bi-weekly, and with supportive measures, the patient slowly recovered. blood enterovirus pcr remained positive. with no evidence of gvhd, csa was tapered off by day+ and child was discharged on d+ on a bi-weekly ivig replacement. she presented days later with signs of raised intracranial pressure. mri showed hydrocephalus, and vp shunt was placed and broad spectrum antibiotics administered. csf showed wbc o /mcl, protein . g/l and enterovirus positive. methylprednisolone mg/kg/day was started suspecting iris. in subsequent csf testing days later, enterovirus was negative. enterovirus pcr remained positive in blood during this period. patient's clinical deterioration correlated with a rise in cd /cd counts and c reactive protein with clearance of enterovirus from csf, blood and stool ( figure ). subsequently, the child showed gradual but marked improvement and discharged home. discussion: the clinical features of index case fits into criteria for iris . markedly raised crp suggests high il- levels without any bacterial or fungal pathogens being isolated. in addition, iris occurs at the site of prior active infection (brain in index case) and viral clearance and clinical recovery demonstrated with the continuation of steroids. the incidence of enterovirus infection in hsct recipients is around % . iris, in this case, had a temporal correlation with discontinuation of csa, and it has been shown that discontinuation of immunosuppression is associated with higher risk of iris. a high index of suspicion for iris is necessary during immune recovery post-hsct especially when immunosuppression is being tapered in a patient with pre-existing infection. aggressive antiviral treatment (when available) and judicious immunosuppression are the keys to managing iris complications. posttransplantation lymphoproliferative disease (ptld) is a significant cause of morbidity and mortality in allogeneic stem cell transplant patients. identifying high risk patients, routine pcr screening, early diagnosis and therapy are crucial for successful management. patients and methods primary objectives of this study were to describe epidemiology of ebv associated ptld and to assess risk factors in our paediatric cohort. additionally, role of immunoglobulin (ig) levels as a possible diagnostic/prognostic marker was analyzed. between january and june , allogeneic transplantations were performed in pediatric patients ( boys and girls) at our center. median age was . years ( . - ). underlying diseases were hematological malignancies ( %), nonmalignant hematological conditions ( %), immunodeficiencies ( %) and others ( %). stem cell source was bone marrow ( %), peripheral blood ( %) and cord blood ( %). donors were unrelated ( %), sibling ( %), haploidentical ( %) or other matched family donors ( %). routine ebv pcr screening and ig level detection were performed weekly. rituximab prophylaxis was given only in nine cases. results ebv dnaemia was found in / patients ( . %), while ptld was diagnosed in / patients ( . %). all ptld cases were related to ebv infection, median of highest viral load was copies/ml ( - ). diagnosis was confirmed by biopsy in / cases, further five fulfilled criteria of probable ptld (positive pcr with appropriate clinical symptoms). ptlds occurred at a median of day + ( - ) after transplantation. all patients received rituximab treatment along with a reduction of immunosuppressive therapy. four patients died of ptld (mortality %), all confirmed by autopsy. a higher incidence of male gender ( / ; . % vs . %), bone marrow graft ( / ; . % vs . %), hematological malignancy ( / ; . % vs . %) and second transplantation ( / ; . % vs %) could be detected among ptld patients when compared to the non-ptld group. elevated igg, iga or igm levels were observed in / patients. nine out of had positive ebv pcr testing, eight of them developed ptld. five of the ptld patients had monoclonal or biclonal immunoglobulin elevation, two of them died. in cases, elevated ig level preceded the positive ebv pcr results by at least week. conclusion: at our centre incidence and mortality of ptld was similar to published data. we observed a tendency that a higher representation of male gender, hematological malignancy, bone marrow graft and second transplantation could be confirmed in the ptld group however due to small number of patients, a correlation and statistical significance could not be calculated. elevation of immunoglobulin levels do not seem to be specific for ptld but in selected cases it could predict ebv disease earlier than pcr testing. disclosure of conflict of interest: none. autologous peripheral hematopoietic stem-cell transplantation is a procedure of a stem cell rescue with patients' own previously collected hematopoietic stem cells, after myelotoxic therapy. the purpose of stem cell reinfusion is to ensure adequate recovery of hematopoiesis, shorten the period of profound neutropenia and to reduce the risk of infections. the transplantation itself carries a moderate risk for infection but some patients have higher risk due to the nature of underlying disease, earlier treatment and in case of severe mucositis. for these reasons, all treated patients are in isolated clean rooms and receive ciprofloxacin, fluconazole and acyclovir prophylaxis. in the . -year period, autologous transplantations were performed. the patients were - years old, with median of . years. of all transplanted patients, or . % had multiple myeloma, or . % had lymphoma and or . % had acute myeloid leukemia. all of the patients received pegfilgrastim mg on the first or the second posttransplant day. febrile neutropenia (ne o . × /l) was reported if patient's temperature was above . °c in one measurement or above °c in two consecutive measurements. these patients were treated empirically with piperacillin/tazobactam . g four times a day with the addition of vancomycin in the case of severe mucositis or pulmonary infiltrates. in all cases blood and urine cultures were performed, as well as testing for seasonal flu. time to neutrophil recovery (ne . × /l) was - days, with a median of days, and average of . days. febrile neutropenia was reported in patients ( . %) and in ( . %) patient's samples pathogen was isolated. gramm negative bacteria caused sepsis in . % of patients. we had to change empirical therapy according to antibiogram in . % patients. in month follow-up period, there were two ( . %) infection related deaths. our data on incidence of infections is consistent with literature data but large number of papers show satisfactory results of safety of patients discharged from hospital immediately after the autologous stem cell transplantation and who were treated at home during the phase of profound neutropenia. there is still an ongoing debate whether it is possible to conduct this procedure in such manner in our health system. disclosure of conflict of interest: none. fluconazole was equal to mold-active drugs in preventing early invasive fungal disease after allogeneic stem cell transplantation regardless of transplantation type y sun, j hu, h huang, j chen, j-y li and x-j huang there are still controversies that whether mold-active drugs is better than fluconazole in preventing invasive fungal disease (ifd) after allogeneic stem cell transplantation (hsct). we hypothesis that the optimal prophylaxis might be different in patients with different risk profile, such as in different time period after hsct or received alternative donor transplantation. in the prospective china assessment of antifungal therapy in haematological disease (caesar) study database, out of patients received primary antifungal prophylaxis were analyzed. the ifd incidence of different time period after transplantation (early, late and very late) and survival were compared among different drug groups. in patients with fluconazole, itraconazole, voriconazole or micafungin prophylaxis, the overall incidence of ifd after transplantation were . %, . %, . % and . %, respectively (p = . ). however, there is no difference in early ifd (o days post hsct) among groups of patients. the risk factors associated with occurrence of ifd were neutropenia duration days (po . , or . ( . - . )), adult (p = . , or . ( . - . )) and alternative donor (unrelated donor or haploidentical donor) transplantation (p = . , or . ( . - . )). in the sub-group analysis with only alternative donor (unrelated donor and haploidentical donor), it also demonstrated that fluconazole is equal to other mold-active drugs in preventing early ifd. patients received fluconazole prophylaxis has even better overall survival. the overall survival in patients with fluconazole, itraconazole, voriconazole or micafungin prophylaxis were . %, . %, . % and . %, respectively (p = . ). our current [p ] study suggests that fluconazole is equal to mold-active drugs to prevent early ifd in hsct patients, even in high-risk patients received transplantation from alternative donors. however, further prospective randomized study was warranted to confirm this conclusion. disclosure of conflict of interest: none. ( . %) received autologous hsct and ( . %) allogeneic hsct. sixty-five out of patients ( . %) were affected by different haematological diseases: by lymphoma, by multiple myeloma, by chronic lymphocytic leukaemia and by others diseases including mastocytosis, amyloidosis and essential thrombocythemia. hbv reactivation prophylaxis prescribed was entecavir for hbsag+ inactive carrier patients and prolonged lamivudine (lmv) course for ( %) patients. in patients ( . %) lmv prophylaxis was withdrawn - months after the end of immunosuppressive therapy. eight out of patients ( . %) experienced hbv reactivation: of them during lmv treatment and then they were switched to entecavir or tenofovir therapy, patients reactivate hbv after lmv interruption ( . %). in these patients reactivation was observed after an average time of months (range: - ) after discontinuation of lmv prophylaxis. median duration of prophylaxis was months (range: - ) after the end of immuno-suppression. three out of patients ( %) underwent allogeneic hsct and patients ( %) received rituximab. one out of ( %) seroreverted in hbsag positive and hbsab negative status, with hbv-dna ui/ml (table ) . two patients out of ( %) experienced hbv-dna detection below ui/ml. disclosure of conflict of interest: none. [p ] [p ] the severity is measured on grades (grade : microscopic hematuria to grado : clots cause urinary tract obstruction). the treatment is based on support measures: hyperhydration, continuous bladder irrigation, instillation of topical agents and in severe cases must be performed a cystoscopy for clot evacuation. in the case of the presence of poliomavirus virus (bk virus) the use of cidofovir had been demonstrated in vitro studies to have activity against bk virus. we performed allogenic transplants of which are haploidentical from to october . we realized a retrospective case study to analyses the experienced in the management of hc. results: of a total of allogenic transplants realized, developed a hc: patient received an identical hla transplant and the patients remaining haploidentical allotrasplant. all cases were male, with an age range of - years. the status of the disease was: were in complete remission and had visible disease. of the patients received cyclophosphamide as immunosuppressive therapy and all patients received cyclosporine and mofetil micofenolate also. the onset of the symptomatology was between day and day post transplant and the range of duration was from to days. the four patients precised continuous bladder irrigation but because of the poor response they received instillation of hialuronic acid ( doses). two patients required the use of cidofovir ( doses). one of the four patients required urinary tract catheterization because of hydronephrosis and renal impairment. in our review we confirmed that this entity is more frequent in the haploidentical transplant and bkv is the most prevalent cause in the late hc. -the three patients received doses of cidofovir ( mg/kg) without probenecid and had a good response. -three patients present acute renal failure associated to hc. the four patients needed bladder instillations with saline but they had poor response and received at least doses of hyaluronic acid. disclosure of conflict of interest: none. hsv infection in allo-hsct setting is mostly reactivation of latent virus. hsv disease commonly presents as mucocutaneous lesions of the oral cavity. however some patients develop serious fatal visceral dissemination. prophylactic use of acyclovir has markedly reduced the incidence of hsv disease during the period of neutropenia after allo-hsct. in this study, our aim is to demonstrate the incidence, clinical outcome and risk factors for hsv disease in adult allo-hsct. between and , patients who underwent allo-hsct in our center were included to the study. all hsct candidates and donors were tested for hsv- / immunoglobulin g (igg) antibodies prior to transplantation. all patients received acyclovir prophylaxis (related transplants mg tid, unrelated transplants mg tid) during conditioning and after allo-hsct up to months. chlorhexidine oral solution as well as bioadherent oral protective gels was used for oral hygiene. all patients were followed for symptoms of reactivation. hsv / igg seropositivity was detected in recipients ( %) and donors ( %). the distribution of hsv status was as follows: recipient and donor seropositive in ( %), recipient and donor seronegative in ( %), recipient seropositive and donor seronegative in ( %), recipient seronegative and donor seropositive in ( %) transplants. the median age of the patients was (range: - ), patients were male ( %) and ( %) had malign disease. the stem cell source was peripheral blood in ( %) patients and ( %) received grafts from related donors. sixty four patients ( %) received myeloablative conditioning regimen. the most common graft-vs-host disease (gvhd) prophylaxis administered was cyclosporine (csa) and methotrexate (mtx) in patients ( %). acute graft vs host disease was detected in patients ( %).four patients from seropositive patients ( %) had hsv reactivation, the patient characteristics are given in the table. all patients had hsv reactivation within month of allo-hsct except one patient had symptoms at sixth month posttransplant when he suffered from oral gvhd. all patients s and donors were seropositive prior to allo-hsct and responded well to antiviral treatment. the incidence of hsv reactivation in allo-hsct was detected as % which is lower to previous studies. successful primary prophylaxis and oral hygiene might reduce the incidence. all patients were responded to antiviral treatment and no visceral dissemination was detected. disclosure of conflict of interest: none. patients who have received hematopoietic stem cell transplantation (hsct) may suffer, to some extent, losses in humoral and cell immunity against antigens to which they had been previously exposed naturally (infection caused by wild microorganisms) or artificially (through vaccination). the conditioning regimen for hsct replaces the patient's immune system and involves the loss of previous immunity. this study analyzed patients included in the vaccination program for hsct recipients in the salamanca health care complex during the period - . we assessed the serological status prior to hsct for the following immunopreventable diseases (hepatitis b, hepatitis a, varicella), and the study after hsct also included measles, rubella and parotitis, prior to their inclusion in the hsct vaccination program. the study included patients, . % of which (n = ) were men. . % of the patients (n = ) were allogeneic hsct recipients with an average age of ± years, and . % ( ) were autologous hsct recipients with an average age of ± years. prior to hsct, % of the patients showed immunity against hepatitis b (hbv antibodies ui/l), . % against hepatitis a (positive for hav igg) and % against varicella (positive for varicella igg). no statistically significant differences were observed regarding this variable hepatitis b anti-hbs ui/l, hepatitis a, igg positive, varicella igg positive, measles igg positive, rubella igg positive, parotitis igg positive. table compares the serological status before and after transplantation. in the pre-transplant serological study we observed that less than half of the patients are protected against hepatitis b, while over % of them are protected against hepatitis a and varicella. regarding the diseases in which we know the serological status before and after transplantation (hepatitis a, hepatitis b and varicella), we observed that most patients maintain immunity. in the case of rubella, measles and parotitis we only have access to the serological status after transplantation, and we observed that parotitis is the disease with the lowest seroprotection. therefore, vaccination would be indicated, just as in the case of hepatitis b. the clinical results support the need to adapt the vaccination schedule to the immunological status of the patients after hsct individually. disclosure of conflict of interest: none. impact of cumulative steroid dose on infectious diseases after allogeneic hematopoietic cell transplantation m watanabe , j kanda, t kitano, t kondou, k yamashita and a takaori-kondo after allogenic hematopoietic cell transplantation (hct), highdose steroids are used to treat transplantation-related complications such as graft-versus-host disease (gvhd). however, the use of high-dose steroids is associated with an elevated risk of infectious diseases. information on the association between cumulative steroid dose and infectious diseases after hct is scarce. a total of patients who underwent their first hct in kyoto university hospital from to and survived at least days after transplantation were included in this study. we analyzed the association between cumulative steroid dose used within days after transplantation and the occurrence of infectious diseases, including invasive fungal infection (possible/probable/proven cases), cytomegalovirus (cmv) antigenemia, and bacteremia through days after transplantation. sixty-three patients received transplantation from a related donor, received unrelated bone marrow grafts, and received unrelated cord blood units. their median age was (range: - ) years and median day of neutrophil engraftment after transplantation was . patients were categorized into groups according to their cumulative steroid dose within days: no steroid administration (n = ), low-dose cumulative steroid administration under mg of prednisolone in total (n = ), and high-dose cumulative steroid administration over mg of prednisolone in total (n = ). reasons for steroid administration were treatment for gvhd in patients, engraftment syndrome in , and other reasons including lung complications in . the rate of invasive fungal infection was % ( possible cases with pneumonia and proven case of candida blood stream infection) and we found no apparent association between fungal infection and steroid use regardless of dose. cmv antigenemia was diagnosed in %, % and % of patients in the groups respectively, and both low-dose and high-dose steroid groups were significantly associated with a high risk of cmv antigenemia (low-dose group, adjusted hazard ratio (ahr), . , p = . ; high-dose group, ahr . , p = . ). bacteremia was diagnosed in . %, % and % of patients in the groups, respectively. high-dose steroid use was a risk factor for bacteremia (ahr . , p = . ). seven patients died from infection (fungal, ; viral, ; bacterial, ). two of three bacterial infection-related deaths occurred in the highdose steroid group, although the number of events was too small to analyze. our data confirmed that steroid administration is itself a risk factor for cmv antigenemia and close observation to detect cmv antigenemia is mandatory for patients using steroids regardless of its cumulative dose. high-dose cumulative steroid use is a risk factor for bacteremia. contrary to our expectations, steroid administration showed no apparent association with invasive fungal infection in our study, perhaps because of its generally low incidence in our hospital. disclosure of conflict of interest: none. impact of different t-cell depletion techniques on the incidence of infectious complications after allogenic hematopoietic stem cell transplantation k aikaterini , s federico , d-l vu , e boely , c dantin , a pradier , y tirefort , a-c mamez , o tsopra , c stephan , y beauverd , e roosnek , s masouridi-levrat , c van delden , y chalandon department of oncology, hematology unit, university hospital of geneva and department of medicine specializations, infectious diseases, university hospital of geneva t-cell depletion (tcd), obtained by either in vivo antithymocyte globulin (atg) administration or ex vivo depletion, is a well-established strategy for graft-versus-host-disease (gvhd) prevention after allogeneic hematopoietic stem cell transplantation (hsct) - . however, the prolonged lymphopenia associated with tcd can result in increased incidence of disease relapse and infections. although many studies investigated the impact of tcd on disease relapse - , little is known about the impact of tcd strategies on the incidence of infectious complications after allogeneic hsct. we retrospectively evaluated the incidence of infectious complications in consecutive patients who underwent allogeneic hsct at our center from september to december . patients received tcd grafts obtained by in vivo atg administration as part of the conditioning regimen (atg group). patients received partially tcd grafts obtained through incubation with alemtuzumab in vitro washed before infusion followed on day + by an add-back of donor t cd + cells (ptcd group). patients received grafts tcd by both methods combined. patients did not receive any form of tcd (no-tcd group). cumulative incidence estimates of infectious complications were calculated and compared using the gray test. given the increased risk of infection associated with gvhd and its treatments, gvhd or death from other causes were defined as competitive events in the analysis. we didn't observe any significant difference in the -year cumulative incidence of bacterial infections in patients receiving tcd by atg ( % ( % ci - . %)) ptcd ( . % ( % ci . - . %)) or both ( % ( % ci . - . %)) compared with patients receiving no tcd ( . % ( % ci . - . %)). similarly, the -year cumulative incidence of viral infections or reactivations was comparable in patients receiving no-tcd grafts ( . % ( % ci . - . %)) compared with patients receiving tcd grafts (atg: . % ( % ci . - . %); ptcd: . % ( % ci . - . %); atg+ptcd: . % ( % ci . - . %)). finally, no significant impact of tcd was observed on -year cumulative incidence of fungal (no-tcd: . % ( % ci . - . %); atg: . % ( % ci . - . %); ptcd: . % ( % ci . - . %); atg+ptcd: . % ( % ci . - . %)) and parasitic (no-tcd: . % ( % ci . - %); atg: % ( % ci . - . %); ptcd: . % ( % ci . - . %); atg +ptcd: . % ( . - . %)) infections. image/graph: -year cumulative incidence estimates of infectious complications depending on the tcd strategy employed. the results of our retrospective analysis indicate that the cumulative incidence of bacterial, viral, fungal and parasitic infectious diseases are similar in patients receiving tcd grafts compared to those receiving no-tcd graft, suggesting a favorable toxicity profile of different tcd strategy in respect of infections. these results should be confirmed by similar analysis in large scale, prospective clinical trials assessing the potential benefits of tcd on transplantation outcomes. patients with aml were considered eligible for hsct, died before transplantation. patients ( %) underwent transplantation from hla-identical sibling, ( %) from haploidentical family donor and ( %) from matched unrelated donor, while patients ( %) received unrelated cord blood cells. twenty ( %) out of eligible patients have had an ifi episode before transplant: were proven, probable and possible; ( ( %) pneumonia, ( %) gastroenteritis, ( %) sinusitis, ( %) candida sepsis, ( %) meningitis and ( %) cutaneous abscess were registered). five ( %) out of patients with a previous ifi and ( %) out of without previous ifi did not receive hsct (or . % ci . - . , fisher test p: . ). the majority ( %) of patients with a previous ifi waited hsct more than months from the date of eligibility in comparison with those without a previous ifi ( % vs %; or . , % ci . - . , p-value . fisher test) overall a post transplant ifi episode was diagnosed in ( %) of transplanted patient; ( %) had a relapse of a past ifi vs ( %) of the patients without a previous ifi who had a new episode. (or . , % ci . - . , p-value . yates test).a higher number of patients with ifi ( out of , %) respect to those without a previous ifi ( out of , %) died in a median time of days(range: - ) after hsct. furthermore, those who had a previous ifi had a lower median survival ( days (range: - )) compared to patients without a previous ifi ( days period (range: - )) (student's t-test p: . )). a previous ifi episode in the pre transplant period slows and limits the accessibility to hsct, and is significantly associated with an increased mortality. disclosure of conflict of interest: none. s delayed immune reconstitution has been described for haploidentical hematopoietic stem cell transplantation (hsct) compared to conventional hsct, nevertheless the incidence of invasive aspergillosis infections (iai) in haploidentical sct and the efficacy of primary prophylaxis are not well defined. our objective is to describe the incidence, risk factors and mortality of iai in our patients, using as prophylaxis micafungin during the conditioning and neutropenia period, switched to posaconazole or voriconazole when oral intake is feasible. we retrospectively analyzed consecutive patients from to who received haploidentical grafts: unmanipulated for adults, tcrab depleted in children and cd ra depleted in children. the stem cell source was peripheral blood in all cases. adults ( - yo) were treated for aml/mds (n = ), all (n = ) and lymphoma (n = ). children ( mo- yo) were treated for aml (n = ), all (n = ), aplastic anemia (n = ) and immunodeficiencies (n = ). conditioning regimen was bu-flu-cy (n = , adults), thio-bu-flu (n = , adults), flu-mel-thio for all pediatric patients; atg was used in children and tli in children. median follow up was months ( - ) for adults and months ( - ) for children. we used eortc criteria for iai and analyzed probable or definite as cases. there were events of iai, with a bimodal presentation: events ( . %) during neutropenia period and ( . %) after months of hsct ( figure ). five of them were probable and one definite (aspergillus niger). site of infection was mainly pulmonar; cns was suspected in two adult patients and skin was proven in one adult patient. all patients at the late period had chronic gvhd at diagnosis. one patient had primary graft failure. severe cmv disease (hepatitis and colitis) was present in one adult. mortality related to iai was high ( / ), patients died at a median of days. figure . iai patients characteristics the global incidence of iai in haploidentical hsct is similar to conventional hcst. primary prophylaxis with micafungin switching to oral triazole is successful ( . %) during the early period. late cases ( . %) had clearly known risk factors (chronic gvhd, steroids and cmv), and primary prophylaxis had been modified due to toxicity or interactions. iai mortality in our patients is very high ( %) despite effort in prophylaxis, diagnosis and treatment. visceral intractable abdominal pain prior to skin lesions from herpes zoster can be misdiagnosed as gvhd post stem cell transplantation which may lead to initial increase in immunosuppression and hence high mortality if we don't suspect. case report and literature review through pubmed results: year-old male with relapsed all post mud pbsct ( / ) transplant in following cy tbi atg conditioning presented at day + with intractable diffuse abdominal pain with constipation. no history of nausea, vomiting or skin rash. on physical examination his abdomen was soft, diffuse tenderness but no rigidity, muscle guarding and rebound tenderness. laboratory tests including liver function test, amylase, lipase were normal. usg abdomen and mri abdomen showed no abnormalities, except for presence of fecolith. during the stay his pain worsened needing morphine infusion, pca and later ketamine. he had previous history of acute gut gvhd controlled on budesonide and cyclosporine which was later being weaned once his symptoms were controlled. in view of previous history of gvhd, gi consultation was sought and he underwent ugi endoscopy and biopsy which was non-significant. on day of his admission he developed a pustular skin lesion on thigh and scrapping from that showed vzv and his blood pcr was also positive, he was started on intravenous acyclovir. his lesions improved and crusted and his abdominal pain subsided after h of acyclovir and was discharged on oral acyclovir after days of intravenous therapy. review of literature illustrated in table . severe abdominal pain in patients who received an allogeneic stem cell transplant has a broad differential. here we describe a case of vzv presenting with intractable abdominal pain needing opioids. because of the poor prognosis and life-threatening nature of disseminated vzv disease, it should be considered and included in the patient's workup. intravesical cidofovir ( mg/kg, diluted in ml sterile water) was once weekly applied until symptom control for min. via a transurethral catheter, i.v. cidofovir was initiated if no symptom control was achieved after local applications. in patients with hc or a lavage catheter was added. bkv cystitis (dysuria (n = ) or dysuria combined with hematuria (n = )) developed in out of transplants ( %). median age was years, % were female and % received a mismatch transplantation after mac or ric conditioning regimens. in % of bkv cystitis cases also cmv reactivation within the first days could be detected. % had acute gvhd ii°-iv°at the onset of bkv cystitis and % received steroid medication. the median time to symptom occurrence was day + after hsct (iqr - : - ). patients ( with dysuria and one either hc °and °) didn´t require therapy due to self limiting symptoms. ( %) of treated patients showed only dysuria, ( %) hc °, ( %) hc °, ( %) hc °and ( %) hc °. the first patient was treated with i.v. cidofovir twice and symptoms relieved. all the following patients were exposed to intravesical cidofovir as st line therapy. patients ( %) achieved a complete remission with a median of intravesical procedures (range: - ). patient showed symptom improvement and all patients didn´t require further therapy. patients had to be switched to i.v. application due to bladder spasms during intravesical application (n = ) or to insufficient symptom control (n = ). out of these responded to i.v. treatment, whereas patient receiving nd transplant didn´t respond at all. in patients with spontaneous symptom relieve the median bkv concentration at the time of symptom onset was log lower compared to those requiring antiviral therapy. local therapy reduced bkv viruria by log. pain during cidofovir instillation in % of patients was the only significant side effect of local therapy compared to creatinine increases by % in . % of i.v. treated patients. intravesical treatment of symptomatic bkv cystitis with cidofovir ( mg/kg) is safe and effective with an % symptom improvement rate and no systemic side effects. in patients without sufficient symptom or bleeding control i.v. cidofovir is still an option, which however induces significant renal toxicity. we therefore recommend intravesical cidofovir as st line therapy in case of dysuria or hematuria induced by bkv after hsct. disclosure of conflict of interest: none. haemorrhagic cystitis is a recognised complication of stem cell transplant (sct), with a reported incidence of - % of cases ( ) . the majority of cases are associated with bk polyomavirus (bkv), and less often adenovirus and cytomegalovirus. there are a lack of high quality studies on the optimal prevention and management of haemorrhagic cystitis. treatment options are restricted by conditioning toxicity, immunosuppression and other co-morbidities such as renal impairment. cidofovir has an inhibitory effect on bkv replication and has been used extensively in the treatment of haemorrhagic cystitis. however, severe nephrotoxicity limits routine intravenous use in sct patients. alternative options include using low dose intravenous cidofovir or intravesical administration. we conducted a retrospective case review of post sct patients presenting with bk virus associated haemorrhagic cystitis in our institution between january and november . we identified patients in total ( male, female). the indications for stem cell transplant were as follows: severe aplastic anaemia high risk aml relapsed aml relapsed all onset of symptoms (haematuria and painful micturition) ranged from day − to day + , and the time to resolution of symptoms varied from days to days. four of the patients were treated with intravesical cidofovir only, with the number of doses required varying from to . one patient received combination treatment with both intravenous ( doses), and intravesical cidofovir ( doses). all patients had a good clinical response with complete resolution of symptoms and no major complications. however, the level of bk virus in the urine did not always correlate with clinical response. some of the patients did not tolerate urethral catheterisation and required a general anaesthetic for the placement of the urethral catheter; patient required a supra-pubic catheter. currently out of patients are alive and well; patients died from causes not related to bk virus associated haemorrhagic cystitis. our experience shows that intravesical administration of cidofovir is a safe and effective option for the treatment of bk virus associated haemorrhagic cystitis. an allogeneic stem cell graft from a cytomegalovirus (cmv) seronegative donor puts recipients at high risk of cmv reactivation which can lead to cmv disease and mortality. based on the immunogenicity of cmv phosphoprotein (cmvpp ) we initiated a clinical phase i trial with a novel vaccine designed by our group: a cmvpp -derived peptide in water-in-oil emulsion (montanide) plus administration of granulocyte-macrophage colony stimulating factor. ten patients received four vaccines s.c. at a biweekly interval after allogeneic stem cell transplantation. we monitored the patients for their clinical outcome and cmvpp antigenemia. multi-color flow cytometry test were performed to assess cmvspecific cd + and gamma-delta t cells. novel neutralizing anti-cmv antibody assays were established and correlated to clinical parameters. findings: in general, patients tolerated the peptide vaccination well, no drug-related adverse events others than rash or induration at the site of injection were detected. seven of nine patients with cmvpp antigenemia cleared the cmv after four vaccinations and were hitherto free from antigenemia. two patients with cmv reactivation showed persisting cmv antigenemia. one of these two refractory patients received additional four injections and remained hitherto free from cmv antigen. another patient obtained a prophylactic vaccination and did not develop antigenemia. an up to six-fold increase in frequency of both cmv-specific cd + t cells or vdelta -gamma-delta t cells was detected in five patients. moreover, titers of neutralizing antibodies increased in four patients up to -fold over the time of vaccination. humoral and cellular immune responses correlated with clearance of the cmv load. cmvpp peptide vaccination was safe and well tolerated in patients after allogeneic stem cell transplantation at high risk for cmv reactivation. the vaccine showed encouraging immunological and clinical results. a prophylaxis study using the vaccine in solid-organ transplant patients is ongoing. disclosure of conflict of interest: none. sporopachydermia cereana is a rare yeast found in necrotic cactus tissue, predominantly in the americas. infection in humans has only been reported in neutropenic patients with fatal course, either directly from the pathogen or other complications of immunosuppression. treatment is complicated by difficulties in pathogen-identification with conventional diagnostic techniques and by resistance to echinocandins. here we present a patient with acute myeloid leukemia (aml) and s. cereana infection. this is the first patient who was successfully treated with antifungal therapy and who survived s. cereana infection. case presentation we present the case of a -year-old female patient who was diagnosed with normal karyotype aml with dnmt a and idh mutations in december . she achieved complete remission after two cycles induction chemotherapy. during the nd induction cycle the patient developed persistent fever in neutropenia despite broad-spectrum antibiotics and the replacement of prophylactic fluconazole to caspofungin. blood cultures showed growth of s. cereana, shown to be sensitive to azoles (mic fluconazole o mg/l, mic voriconazole o . mg/l) as well as amphotericin b (mic o . mg/l), but resistant to caspofungin (mic mg/l). following the susceptibility profile the treatment was changed first to liposomal amphotericin b, and with the availability of mic results to voriconazole. metastatic fungal infection (that is, endocarditis, endophthalmitis, hepatosplenic candidiasis) was excluded. after regeneration of peripheral blood values the treatment was switched to oral voriconazole. a ct scan of the chest and abdomen prior to allo-hsct after weeks of treatment with voriconazole revealed new multiple necrotic mesenteric lymph nodes. an ultrasound-guided biopsy of a node revealed no growth on fungal cultures, a grocott stain revealed no hyphae or spores. a panfungal pcr of an its (internal transcribed spacer) fragment revealed fungal dna, which could be confirmed as s. cereana. at this time the level of voriconazole in serum was found to be sub-therapeutic ( . mg/l), and the dosage was increased accordingly. subsequent ct scans and weeks later revealed a regression of the affected abdominal lymph nodes. in the further course non-myeloablative conditioning with fludarabine and busulfan prior to allo-hsct using pbsc from her hla-matched brother was performed. under prophylaxis with cyclosporine, methotrexate and antithymocyte globulins (atg) graft-versus-host disease (gvhd) remained absent. the allo-hsct was performed under voriconazole treatment with no further complications and the patient engrafted at day . the treatment was changed to fluconazole mg daily before discharge. due to the complete radiological regression of the infection in follow-up scans and excellent general condition of the patient months after hsct, fluconazole was discontinued. the patient remains in morphological complete remission months after hsct and has a % donor chimerism. the first published case of survival of infection with s. cereana exemplifies the continual progress made in treating infections in the severely immunocompromised patient. diagnosis via its sequence-analysis seems reliable but a high index of suspicion is required for neutropenic patients who do not respond well to standard antimycotic therapy. the increased availability of the technology may lead to more frequent diagnoses in the future. disclosure of conflict of interest: none. neutropenic enterocolitis (ne) is a clinical syndrome characterized by fever and abdominal pain in patients who received chemotherapy for hematological malignancies and who treated with stem cell transplantation (sct). the aim of this study was to determine the incidence, risk factors and outcome of ne after autologous sct (auto-sct). we retrospectively evaluated patients with non-hodgkin lymphoma (nhl), hodgkin lymphoma (hl) and multiple myeloma (mm) who underwent auto-sct between january and december in our center. patients with lymphoma were conditioned with carmustine, etoposide, cytarabine, melphalan (beam) or thiotepa, etoposide, cytarabine, cyclophosphamide, melphalan (tecam). patients with multiple myeloma were treated with melphalan as conditioning. diagnosis of ne was established in case of neutropenic fever, abdominal pain or diarrhea, and bowel wall thickening mm on abdominal ultrasonography. febrile neutropenia was seen in ( %) patients of all. the median time from transplantation to neutropenia was . days (range: - days). ne occurred in ( . %) in all neutropenic patients. the median time to ne after auto-sct was days (range: - days). the median neutrophil engraftment time was . days (range: - days). abdominal pain was seen in all patients with ne. twenty one patients ( %) had diarrhea. ileus was seen in ( . %) patient and septic shock was developed in ( . %) patients. five ( . %) of patients had bloodstream infection. klebsiella pneumoniae in , pseudomonas aeruginosa in , escherichia coli in , staphylococcus aureus in and coagulasenegative staphylococcus in patient were documented in patient's blood stream. early diagnosis was made by abdominal ultrasonography in all patients at a day of median days (range: - ). twenty ( %) patients were resolved completely with good supportive care and proper antibiotherapy. two ( %) patients died of septic shock and ileus. ne is a rare but serious complication in patients underwent high dose chemotherapy followed by auto-sct. gramnegative bacteria are the main causative pathogens. abdominal ultrasonography is the simple, cheap, fast diagnostic and noninvasive procedure that allows the early diagnosis and effective treatment. disclosure of conflict of interest: none. [p ] neutrophil transfusions in the treatment of neutropenic patients submitted to allogeneic hsct: possible role on graft failure s giammarco, p chiusolo , l laurenti , f sorà , n piccirillo , l teofili and s sica hematology department, università cattolica del sacro cuore and hematology departement, università cattolica del sacro cuore granulocyte transfusions (gtx) from g-csf-stimulated donors have been shown to increase the absolute neutrophil count (anc) before expected haematopoietic recovery in neutropenic patients after chemotherapy or haemopoietic sct. thus gt offers a therapeutic option along with antimicrobial agents and growth factors to improve clinical outcome of neutropenic patients with severe infections. the primary limitations of gt include low component cell dose and leukocyte incompatibility. the transfusion of g-csf-mobilized, hla-matched granulocyte components resulted in sustained anc increments, but the efficacy of this procedure has not been established by convincing randomized control trials. aim: we focused our attention on gt in the setting of allogeneic hsct, in particular on the feasibility and safety of this procedure on the rate of engraftment. between and our centre performed allogeneic hsct. we analyze data from transplanted patients receiving gt at some point during their disease. indication for gt was severe sepsis mainly due to mdr gram-bacteria. patients received a median of gt ( - ), in different phase: patients during induction therapy, during hsct, at diagnosis and during hsct and after hsct. patients' characteristics are summarized in table . median cd + cells dose was . × /kg (range: . - ). donor source was in patients g-csf mobilized peripheral blood, bone marrow and cord blood. median neutrophil recovery ( /mmc) was days and platelet recovery ( / mmc) was days. sepsis were documented in pts and pts developed fuo. relapse was documented in pts ( %). twenty-two pts are still alive and in complete remission ( %), death occurred in pts: due to trm and the remaining for disease relapse. graft failure occurred in of the pts submitted to hsct. among the patients ( %) who experienced graft failure, six ( %) received gt before hsct, because of sepsis during the induction therapy, and the remaining after hsct, during aplasia period. in the remaining group ( pts) not receiving gt, only ( %) graft failure were observed. thus a statistically difference (p = . fisher's exact test) increase in the rate of graft failure was detected in patients receiving gt. the role of gt in the treatment of infections in neutropenic patients remain still unclear for several reasons including the lack of clinical trials convincingly and consistently demonstrating efficacy, by availability of gt donors and by center's experience. gt has been successfully used in our center in patients with severe sepsis from mdr gram-bacteria during severe neutropenia but an increase number of graft failure has been registered in patients subsequently receiving hsct. alloimmunization to hla antigens in patients receiving gt might lead to an excess of graft failure requiring hla antibodies detection and attempt to reduce titer prior to hcst and maximizing stem cell dose. disclosure of conflict of interest: none. viridans streptococci are microorganisms frequently isolated from blood cultures of patients undergoing myeloablative allogeneic hematopoietic cell transplantation (allohct). poor dentition status has been associated with an increased risk of streptococcal bacteremia in the immediate post-allohct neutropenic period. the objective of this study was to evaluate the impact of oral health status on bacteremia risk in a cohort of patients undergoing therapy for acute myeloid leukemia (aml). a retrospective study was conducted in patients with aml treated at dana-farber/brigham and women's cancer center (df/bwcc) from to . there was no formal dental assessment prior to aml induction therapy. all patients underwent protocol directed pre-allohct dental evaluation that included a standardized examination, comprehensive dental radiographs, and detailed treatment planning guidelines. poor oral health status was defined as presence of acute or chronic odontogenic infection, and it was assumed that oral health status at the time of induction therapy was the same as the pre-allohct evaluation findings. oral health status at the time of allohct was determined by the completion of required dental treatment. positive blood cultures were recorded from aml induction to day + post allohct. organisms that caused bacteremia were classified as 'of possible oral source' by a blinded microbiologist. two-sided fisher's exact test was used to compare the oral health status of the entire cohort to patients with blood cultures of potential oral source. from january to january , patients with aml underwent myeloablative allohct at df/bwcc and were s followed through today + , and of these, patients met the inclusion criteria and were included in the cohort. the median age was years (range: - ) and there was similar distribution of genders. the most common aml induction regimen was daunorubicin and cytarabine ( / ; %) and of those that received consolidation therapy ( / ; %), almost all patients were treated with cytarabine. nearly all patients ( / ; %) received cyclophosphamide and total body irradiation for allohct conditioning and the majority of patients ( / , %) received tacrolimus/methotrexate (n = ) or tacrolimus/sirolimus (n = ) for gvhd prophylaxis. over half of patients ( / , %) experienced mucositis during their course of therapy for aml. pre-allohct dental evaluations were completed in / ( %) of patients. of the / ( %) patients identified as having poor oral health status, / ( %) completed all required dental treatment prior to allohct. bacteremias occurred in / ( %) patients, and / ( %) had positive blood cultures of potential oral source. of the patients with positive blood cultures of potential oral source, / ( %) patient developed bacteremia during induction and / ( %) patients developed bacteremia during allohct. of the / ( %) patients identified as having poor oral health status, one patient ( / ; %) had a positive blood culture with a bacteria of potential oral source during induction/consolidation (p = . ). oral health status was not associated with risk of bacteremia of potential oral source at either aml induction/consolidation or allohct. risk of such bacteremia in the setting of myeloablative allohct may be related more to overall gastrointestinal translocation. disclosure of conflict of interest: none. is one of the main alternatives to trimethoprimsulfamethoxazole (tmp-smx) for prophylaxis of pneumocystis pneumonia (pcp)(maertens et al. jac ). ato is less effective than tmp-smx to prevent pcp but the reasons of this lower efficacy are not well understood. ato acts on pneumocystis, plasmodia and toxoplasma species by inhibiting mitochondrial pyrimidine biosynthesis. ato is highly lipophilic and its absorption in volunteers is improved by a fatty meal. there is a wide inter-individual variability in bioavailability and many drug interferences. the aim of this study was to assess the plasma concentrations of ato in patients under pcp prophylaxis with ato oral suspension and explore the factors which might impact its bioavailability. all adult patients receiving ato for pcp prophylaxis in the hematology and clinical immunology wards between may and september were included in the study. the prescribed dose was mg of oral suspension twice a day. blood samples were collected around h after the evening dose (cmin) and - h after the morning dose (cmax). plasma was immediately separated after each sample and frozen at − °c until proceeding to the assay. ato plasma levels were measured by uv-high-performance liquid chromatography. clinical and biological data, exact timing and modalities of intake (during a meal or not), and concomitant medications were collected. cmin and cmax results are presented as median (iqr - %) and compared by mann-whitney u-test or signed rank test when appropriate. patients: a total of measurements were performed in patients (allogeneic hsct patients: ; hematology non-transplanted patients: ; hiv-infected patients: ). the mean age (range) was years ( - ), the m/f ratio was / . only two patients were neutropenic. the median cmin was . μg/ml ( . - . ) and the median cmax was . μg/ml ( . - . ). thirteen of the ( %) patients had a cmin. disclosure of conflict of interest: none. presepsin as a marker of infectious complications during high-dose chemotherapy following autologous hematopoietic stem cell transplantation in lymphoma patients y dubinina, v sarzhevskiy and v melnichenko national pirogov medical surgical center lymphoma patients, who undergo high-dose chemotherapy following autologous hematopoietic stem cell transplantation (autohsct), are at high risk of developing infectious complica- tions (ic). mortality from ic during the transplantation, according to various data ranges from to %. thus the development of models of early prognosis of ic during autohsct has become more urgent. it's reasonable to include the dynamics of biochemical markers of inflammation in these models. presepsin (psp), procalcitonin (pct) and c-reactive protein (c-rp) were assessed on the day of admission to the hospital (da), on d+ , d+ , d+ and on the day of discharge (dd). if patients developed neutropenic fever (nf), the markers were assessed at the beginning of the fever, h after, then on the second, third, fourth days after. there were patients included in the study: patients with hodgkin lymphoma, with non-hodgkin's lymphoma, with multiple myeloma, out of patients there were women and . the median age was years ( - ). the conditioning regimens were cbv, beeac or hd melphalan. depending on the presence of ic, the patients were divided into groups: group patients without infectious complications (n = ), group patients with the development of infectious complications (n = ). the median of the nf development was . days. patients from group had no microorganism growth in blood stream, either in repeated studies. gram+ flora was detected in patients, patient had gram-, patients had mixed flora and patient had pneumocystis jirovecii infection with respiratory insufficiency grade . significant differences in psp level between groups and were determined on d+ , on d+ and the dd after autohsct. considering the median day of the nf appearance ( . days), it's supposed both the prognostic value (differences on d+ , that is, days before the clinical manifestation of infection) and the diagnostic value of psp (differences on d+ and on the dd) ( table , graph ). [p ] disclosure of conflict of interest: none. hc is often a serious complication and occurs in % of allo-hsct recipients. early bleeding is usually the result of chemotherapy toxicity however late occurring hc is multifactorial. bk virus infection has been shown to be related with hc. most studies demonstrate bk virus at the time of bleeding therefore not allowing the risk imposed by asymptomatic infection to be estimated. in this study, our aim is to show the effect of risk factors as well as pre-transplant bk viral load in asymptomatic recipients on development of hc in allo-hsct. between and , we prospectively evaluated allo-hsct. in order to detect the bk viral load, we performed quantitative bk virus pcr (altona diagnostics, germany) from blood samples at days , , and after allo-hsct. informed consents were obtained from all participants. bk virus pcr was considered positive if any number of copies were detected above the analytical sensitivity of the tests. the patients were monitored for signs and symptoms of hs. the risk factors for the development of hs were evaluated by univariate and multivariate analysis. p o . was considered statistically significant. the median age of the group was (range: - ), of the patients ( %) were aged . male to female ratio was . ( / ). fifty two patients ( %) had diagnosis of malign hematological disease. stem cell source was peripheral blood in ( %), bone marrow in ( %) allo-hsct. patients received stem cells from related donors ( %) vs ( %) unrelated or haplo donors. myeloablative conditioning was administered in patients ( %). forty-four of the conditioning regimens ( %) included cyclophosphamide. hc was diagnosed in patients ( %) at a mean of days (range: - ), early hc was detected in of patients ( %). the frequency of bk viremia and number of viral copies are given in detail in table. the frequency of bk viremia increases during transplantation in relation to clinical hc ( %, %, %, %; p = . ). acute graft vs host disease (agvhd) was diagnosed in patients ( %) at a median time of posttransplant day : grade i-ii gastrointestinal/skin/liver in ( %), grade iii-iv gastrointestinal/ skin/liver in patients ( %). the most common gvhd prophylaxis preferred was cyclosporine and methotrexate in patients ( %). in univariate and multivariate analysis (age , sex, diagnosis, stem cell source, donor type, conditioning regimen, agvhd, cy administration, bk virus pcr at days , , ) bk virus titer positivity at day , , (p = . , p o . , p o . ), myeloablative conditioning (p = . ), the presence of agvhd after day (p = . ) and conditioning regimen that includes cyclophosphamide (p = . ) are found to be related with increased risk of hs. patients with hc and clots were treated with continuous bladder irrigation as well as of patients with bk viremia received cidofovir and six of them responded to treatment ( %). our study showed that, bk titer positivity, myeloablative conditioning, presence of agvhd, cyclophosphamide containing conditioning are associated with hc. detection of bk viremia in later transplant period is more sensitive for clinically proven hc. prophylactic treatment might be considered in patients with asymptomatic bk viremia in pretransplant period. [p ] disclosure of conflict of interest: none. this project has been granted by ankara university scientific research committee numbered as b . high-dose chemotherapy with peripheral blood progenitor cell (pbpc) collection followed by a myeloablative conditioning and autologous stem cell transplantation (asct) is considered the standard of care of relapsed/refractory non hodgkin/hodgkin lymphoma (nhl/hl). a widely adopted conditioning regimen is the combination of carmustine etoposide cytarabine and melphalan (beam), whose feasibility and efficacy has been largely demonstrated. high dose fotemustine plus etoposide, cytarabine and melphalan (feam) has in some cases replaced beam conditioning. neutropenic enterocolitis (nec) is a life threatening complication of patients (pts) treated with chemotherapy (cht) with mortality rate up to %. it's a clinical syndrome in neutropenic patients (pts) characterized by abdominal pain (ap), fever (f) and diarrhoea (d). ultrasound (us) was used to evaluate bowel-wall thickening (bwt), and mm is considered diagnostic of nec. early diagnosis is crucial to start conservative medical management (cmm), which appears the optimal strategy for most cases. objective: . to evaluate if nec incidence and outcome differs in beam vs feam and . to evaluate prospectively if bed-side-us (bus) can detect early signs of nec and guide a prompt treatment (cmm or surgical) in order to reduce mortality. in the last years all pts with nhl/hl admitted in our bmt unit wards at university of pisa (italy), undergoing asct were prospectively enrolled. abdominal us was performed, baseline before treatment, and as only one symptom (or a combination) appeared within h from onset: f and/or d and/or ap in cht-related neutropenic pts. pts were conditioned with beam and pts with feam. nec was diagnosed in n = / feam and in n = / beam patients. incidence was % and % respectively, without a statistically significant difference (p = . ). two pts died/ in feam arm ( . %) and pts/ in beam arm ( . %), without a statistically significant difference (p = . ). at time of diagnosis (dx) symptoms were: f+ap+d %, f+d %, f+ap %, ap+d %,d %,ap %. f alone was never present at diagnosis of nec. at dx, f was absent in / nec episodes ( %). all pts were treated promptly as bus allowed diagnosis with cmm except one pts who underwent surgery, guided by us features, during neutropenia. the likelihood of nec dx in a discriminant st model (bayes theorem) for pts with bwt and ap = . %, ap+d = . %, ap+d+f = %, ap+f = . %, d+f = %. bus allowed to detect early signs of nec and to start prompt treatment in this life threatening complication, of nhl/hl pts undergoing asct. this is a prospective study thus the true incidence of nec in nhl/hl undergoing asct should not be underestimated. there is not a statistically significant difference in incidence and outcome of nec in pts conditioned with beam in respect to feam. with bus pts do not live the isolation room. fever is not a condition sine qua non for nec diagnosis. early diagnosis allows most of pts to be treated with cmm. images of bus and ct were superimposable with lower costs, and less radiation exposure. a low mortality rate in pts with a - % chance of developing this life threatening complication suggests that a prompt bus in neutropenic patients as just one symptom presents allows to make early diagnosis of this life threatening complication and guide prompt treatment (conservative or surgical), reducing mortality. disclosure of conflict of interest: none. quantiferon-cmv in the evaluation of cmv-specific immunity after autologous and allogeneic hsct j moreno , ltesta , l zanetti , l serra , b pereira , m souza , a carolina souza , mp souza , vr colturato and cm machado , hsct program, amaral carvalho foundation and virology laboratory, institute of tropical medicine, university of são paulo cytomegalovirus (cmv) is a major cause of morbidity and mortality after allogeneic hsct. the same is not observed in autologous hsct recipients who do not need to receive immunosuppression after transplantation. in the present study, we compared the reconstitution of cmv-specific immunity in autologous and allogeneic hsct recipients. patients were invited to participate in the study and signed the informed consent. cmv surveillance with the antigenemia (ag) test (cmv brite, biotest, germany) was done weekly in the first months of transplant in allogeneic hsct recipients. preemptive ganciclovir therapy was initiated whenever a positive antigenemia was detected. the presence or absence of cmvimmunity was determined by a commercial interferon (inf) gamma release assay (quantiferon cmv, qiagen) before hsct and monthly thereafter up to d+ . from january to october , hsct recipients ( auto and allo) were included in the study. ag was positive in ( %) of the allohsct recipients at a median of (range: - ) days. ag recurrences occurred at a median of . ( - ) days, in of the pts ( . %) who had at least one episode of positive ag. hsct recipients were included in the analysis of qtf-cmv. in the pre-hsct sample, qtf-cmv was reactive in of the allohsct ( . %) and in of the autohsct ( . %). significantly less allo hsct recipients recovered cmvimmunity at day + ( . %) and day+ ( . %) in comparison with autohsct ( % and %, respectively, p o . ). up to day + , all autohsct have recovered cmvimmunity, in comparison to % of the allohsct recipients (p = . , figure ). the qtf-cmv test performed at d+ , d + and d+ did not predict the risk of cmv reactivation in the following month. similarly, the test did not anticipate the risk of ag recurrences: % of the hsct recipients with undetermined or non-reactive qtf-cmv test at d+ had ag recurrence after this period, in comparison with % of the patients with a reactive result (p = . ). in the present study, the qtf-cmv test alone could not predict the risk of cmv reactivation or recurrences. [p ] disclosure of conflict of interest: qiagen. recovery of vδ + γδ t cells is critical to epstein-barr virus reactivation after haploidentical hematopoietic stem cell transplantation j liu, z bian, q fu, l xu, x zhang, y wang and x-j huang peking university people's hospital, peking university institute of hematology, beijing, china epstein-barr virus (ebv) reactivation and its related disease are life-threatening complications in patients undergone haploidentical hematopoietic stem cell transplantation (haplohsct). our previous studies found that impaired cd − cd − t-cell recovery correlated to the increased occurrence of ebv infection after haplohsct. γδt cells make up - % of cd − cd − t cells in the peripheral blood of healthy donors. expansion of vδ + γδt t cells after hsct has been reported and this subset could respond against autologous ebv-lcl in vitro. selective activation and expansion of vγ vδ -t cell could inhibit ebv-lpd development in humanized mice. however, the association of γδ t-cell recovery with ebv reactivation after allohsct remains unknown. this is a prospective cohort study including consecutive patients who were diagnosed as hematological malignancy and underwent haplohsct. recovery of t lymphocyte and a panel of subsets, including cd +, cd +, cd +, cd -cd -, tcrαβ+, tcrγδ+, vδ +, and vδ + t cells, were determined by flowcytometry at , , , days after haplohsct. all recipients and donors were tested negative for ebv dna in the peripheral blood before transplantation. recipients were monitored weekly for ebv dna load until day after transplantation. recipients with peripheral blood plasma ebv dna load copies/ml at least on two consecutive occasions were diagnosed as ebv reactivation (ebv +). ebv − cohort generally represents patients whose ebv dna loado copies/ml in peripheral blood. within days after haplohsct, of ( . %) recipients were diagnosed as ebv reactivation. compared to recipients with negative ebv dna load, the counts of cd +, cd +, and tcrαβ+ t cells were not statistically different in the ebv+ cohort from to days after haplohsct. in contrast, recoveries of cd + and cd -cd -t cells in ebv+ patients were significantly hampered at days after transplantation (p = . and p = . , respectively). although the tcrγδ+ t-cell counts were also decreased at and days in the ebv+ cohort, the comparisons did not reach the statistical significance (p = . and p = . , respectively). notably, recoveries of vδ + γδ t cells at , and days were continuously delayed in recipients with ebv reactivation (p = . , p = . and p = . , respectively). whereas the counts of vδ + γδ t cells were similar between the two groups from to days in this context. in this prospective and large cohort study, we showed that the occurrence of epstein-barr virus (ebv) reactivation was associated with the hampered recovery of vδ + rather than vδ + γδ t cells after haplohsct. our findings will help explore γδt subset-dependent therapeutic strategies to control the serious complications due to ebv infection post transplantation and improve the overall survival of haplohsct recipients. disclosure of conflict of interest: none. in particular, bloodstream infection (bsi)is a frequent complication in the pre-engraftment phase with an impact on the morbidity and mortality of these patients. objectives: to analyze the incidence of bsi in patients undergoing hsct in our center, and to identify predisposing factors for the development of bsi in pre-engraftment phase patients after hsct. fifty-one consecutive patients undergoing hsct were analyzed retrospectively in our center during the period of july and june . the characteristics of the sample are shown in table . we have reported all the bsi between day and day after stem cell infusion. . % ( patients) received antibacterial prophylaxis with ciprofloxacin, five patients with broad spectrum antibiotics and five did not received any drug. the average days of fever have been . days ( - days). a total of blood cultures has been collected ( . per patient). there have been bsi ( . % of the patients) with ( . %) of cases caused by gram-negative organism ( escherichia coli, klebsiella pneumoniae, acinetobacter baumanii, proteus vulgaris and delftia acidovorans) and ( . %) by gram-positives ( enterococcus faecium, enterococcus faecalis, staphylococcus epidermidis, streptococcus mitis and streptococcus viridians group). one patient presented different episodes of bsi, two patients independent episodes and the rest eight, only one microorganism isolated. we have identified two bsi by extended-spectrum betalactamases (esbl-producing organism) and one isolation of carbapenem-resistant gram-negative bacteria. the rate of quinolone-resistant is % in all the sample. in univariate analysis, several factors like presence of comorbidities, presence of severe mucosits, type of catheter and antibacterial prophylaxis modality don't increased the risk to develop bsi (p . ). the place where the procedure is performed does not influence the development of bsi. although the presence of previous infections is not a risk factor, hospitalization for infection in the days before hsct does influence the development of bsi with statistical significance (po . ). the crude mortality rate of the sample has been very low ( %), with only one death related to bloodstream infection. bsi are a common relative complication in the patient undergoing hsct but with an extremely low mortality in our sample. hospitalization for infection in the days before hsct does influence the development of bsi. it is important to note that outpatient model and conventional rooms don't increased the incidence of bsi. although the use of quinolones in prophylaxis does not result in an increase in infections caused by multiresistant micro-organisms (esbl and carbapenemias) with acceptable resistance rates ( %), it also does not reduce the incidence of bsi in our sample. according to our analysis, his routine employment still throws light and shadows. [p ] disclosure of conflict of interest: none. septic episodes with multiple bacterial strains during antithymocyte globulin (atg) therapy for conditioning for allogeneic stem cell transplantation under rifaximin gut decontamination d markel , c schultze-florey , t brockmeyer , v panagiota , c lück , a schwarzer , m beck , e dammann , a ganser , g beutel and m eder recent evidence demonstrates the importance of the enteric microbiome for the development of gastrointestinal graftversus-host disease (gvhd) and mortality after allogeneic stem cell transplantation (sct) ( , ) . accordingly, the usage of the non-absorbed rifamycin derivate rifaximin for gut decontamination has been reported to preserve the intestinal microbiota composition with a positive effect on overall survival in a single centre retrospective analysis ( ). we here report severe septicaemia requiring therapy at the intensive care unit (icu) during atg application for conditioning in three patients with rifaximin used as single agent for gut decontamination within months. after changing our gut decontamination from a chinolon-metronidazole regimen to rifaximin, three cases of severe septicaemia by gram-negative and gram-positive bacteria during atg treatment occurred within months. patient # was a -year-old woman with tmds/aml after breast cancer conditioned according to the flamsa-bu protocol. the second (# ) and third (# ) patient were and -year-old males with a complex karyotype secondary aml after omf and relapsed inv( ) aml with meningeosis leucaemica, respectively. patients # and # were treated with flamsa-bu and flamsa-tbi, respectively. all patients received rabbit atg (atg fresenius/grafalon) at a dose of × mg/kg body weight and rifaximin ( × mg) for gut decontamination. patient # developed severe escherichia coli and pseudomonas aeruginosa septicaemia on day − of the conditioning regimen and had to be transferred to the icu with septic cardiomyopathy for therapy with vasopressants and levosimendan. in patient # escherichia coli, klebsiella oxytoca, staphylococcus hemolyticus and staphylococcus epidermidis were simultaneously detected in blood cultures at day − . the patient was transferred to the icu and treated with vasopressants for septic shock. patient # developed septic shock due to klebsiella pneumoniae and enterobacter cloacae on day − under atg therapy. mechanical ventilation and vasopressor therapy were required. fortunately, all three patients survived and completely recovered without any sepsis related disabilities under escalated anti-infective and intensive care therapy. all were discharged from the hospital in the outpatient clinics. interestingly, all isolated gram-negative pathogens were found to be sensible for a chinolon based gut decontamination. the reasons for these septic complications under atg therapy are not exactly understood but raise a note of caution on the use of rifaximin as single agent gut decontaminant during atg application in conditioning for allogeneic sct. infections with mycobacterium genavense were described for the first time in . since then, several cases have been reported, but almost exclusively in patients with aids. most patients who underwent hsct have insufficient cellular immunity. here we report a mycobacterium genavense infection in a patient mimicking a lymphoma-relapse after hsct. a year-old female patient was diagnosed in july with stage ivb alk-negative anaplastic t-cell-lymphoma with cervical, retro-/supraclavicular, mediastinal, axillary and retroperitoneal lymphadenopathy as well as pulmonary manifestation. two chemotherapy treatment lines and autologous stem cell transplantation resulted in a partial remission. to improve remission prior to hsct the patient received courses of brentuximab-vedotin. after conditioning therapy with fludarabine, busulfan, cyclophosphamide and atg, hsct from a hla compatible unrelated donor was performed in april . a pet-ct-scan in november confirmed complete remission. after hsct the patient remained lymphocytopenic with cell count of cd + cellso /μl. after acute stage iii gastrointestinal graft-versus-host disease (gvhd) low dose immunosuppressive therapy was maintained due to mild chronic gvhd of the liver and the upper gastrointestinal tract. beginning in june the patient experienced increasing fatigue, general weakness, loss of appetite, nausea, night sweating and fever. abdominal ultrasound, urine and blood culture as well as ct scans revealed no focus of infection. different lines of empirical antibiotic therapy resulted only in short term improvement. several blood culture tests remained sterile. a fdg-pet-ct scan showed a paraaortal and parailiacal lymphadenopathy with a high fdg uptake (suv between . and . ), highly suggestive of lymphoma relapse. endoscopic evaluations revealed two polypoid lesions in the bulbus duodeni. histology of duodenal biopsies revealed a massive accumulation of weakly pas-positive bacilli. pcr analysis confirmed an infection with mycobacterium genavense. despite several attempts mycobacteria were not recoverable on solid media even by long term culture. treatment was started with rifampicin, ethambutol, ciprofloxacin and clarithromycin. lymph node manifestation responded to therapy with decreasing fdg-uptake (suv . ) in a control fdg-pet-ct scan months later. after months treatment was terminated due to therapy refractory nausea. lymphocytopenia was persisting with cd + cellso /μl. six weeks after stopping the antibiotic therapy, symptoms as fever and weakness reappeared. duodenal biopsy could not confirm persistent mycobacterial infection. fdg-positive intraabdominal lymph nodes (suv . ) and spleen (suv . ) were detected in a control fdg-pet-ct-scan. five lymphnodes were surgically removed. immunohistology detected histiocytic cell proliferation with no sign of lymphoma relapse. pcr confirmed the presence of mycobacteria-dna. consequently, antibiotic treatment was resumed. mycobacterium genavense can present with all the symptoms of a lymphoma relapse and should be considered in immune compromised patients. reliable diagnosis can only be obtained from lymph node biopsies and/or endoscopic evaluation. treatment has to be accompanied by restoring cellular immunity and should only be stopped after pcr-negative biopsies. disclosure of conflict of interest: none. stratification of patients with multiple myeloma and lymphoma undergoing autologous hematopoietic stem cell transplantation in term of antifungal prophylaxis r moghnieh, s khaldieh, l awad, d abdallah, n droubi, a youssef, a mougharbel, t jisr and a ibrahiim makassed university hospital, beirut, lebanon autologous hematopoietic stem cell transplantation (ahsct) is at intermediate risk for invasive fungal infections (ifi). the recommendations of international scientific societies are not homologous regarding prophylaxis against ifi in patients (pts) undergoing ahsct. the primary end point was to assess risk factors for the need of empiric/preemptive antifungal therapy in ahsct recipients, and to extrapolate to the subgroup of pts that requires antifungal prophylaxis in our population of ahsct pts. the secondary endpoint was to determine the fungal species distribution infecting or colonizing the pts. our study included adult pts ( yo) who underwent ahsct for lymphoma and multiple myeloma (mm) between and . all febrile neutropenic pts are being managed according to the infectious diseases society of america (idsa) guidelines regarding the use of antimicrobial agents in neutropenic pts with cancer. eligible pts were divided into two groups: those who received empirical antifungal therapy and those who did not need it. we recorded demographic and baseline clinical characteristics including: age, gender, comorbidities, stage, disease status at ahsct, high-dose therapy regimen, the presence of mucositis and its grade, the number of cd + cells transfused, the presence of central line or portacath, the need for mechanical ventilation, the presence of diarrhea, the duration of neutropenia, and the presence of bloodstream infections. pts who had lung infiltrates suggestive of ifi were analyzed separately. the causative fungal pathogens and colonizers were analyzed. univariate and multivariate analysis of potential risk factors to assess further significance was performed using spss. patients were included. pts ( %) had lymphoma and pts ( %) had mm. the need of empiric antifungal therapy was statistically more significant in lymphoma than mm pts (po . ).the presence of mucositis grade ⩾ showed a statistical significance for the need of antifungal therapy (p = . ). in the lymphoma group, remission status (pr vs cr) was not a significant factor for the need of empiric antifungal therapy (p = . ).the presence of mucositis grade ⩾ was at the limit of significance ( p = . ). in the mm group, remission status (pr vs cr) did not affect the need of empiric antifungal therapy (p = ). however, mucositis grade ⩾ was found to be a significant risk factor for the need of empiric antifungal therapy (p = . ). following factors: the number of cd + cells transfused, the presence of central line and portacath, the need for mechanical ventilation, the presence of diarrhea, the duration of neutropenia, and bloodstream infections did not show any significance for the need of antifungal prophylaxis in both groups. all recovered fungal isolates (n = ) were not from deep seated tissues biopsies or blood, and were identified as candida albicans in with lymphoma, and in with mm. they reflected the candida ecology in this pts series rather than deep seated fungal infections. we suggest to give antifungal prophylaxis to all lymphoma pts because of the higher need of empirical antifungal therapy, and give antifungal prophylaxis to mm pts having a predisposition for severe mucositis. fluconazole is the antifungal of choice for prophylaxis since all the fungal isolates were candida albicans. keywords: autologous hematopoietic stem cell transplantation, antifungal prophylaxis. disclosure of conflict of interest: none. a -year-old previously fit woman from a rural area of eastern europe was admitted to the hospital for severe aplastic anemia. steroids, csa, antinfective prophylaxis and supportive therapy were administered without response; therefore rabbit atg was then administered, with minor response; the year later, she underwent allogeneic-hsct (mud / , ric: tbi, cyclophosphamide and fludarabine; gvhd prophylaxis: atg, csa, mtx). several days after transplantation she developed left migraine with ipsilateral back-eye pain. brain mri and ct showed a diffuse opacification of paranasal sinuses, mainly in the sphenoid sinus. the symptoms gradually improved with a specific treatment. the patient achieved a quick and complete haematological recovery and she was discharged. at follow-up visits she complained a flare of the migraine, with a left-sided headache that did not improve with nsaids. the headache gradually intensified until vision in the left eye became blurred with conjunctival injection. after consultation with ophthalmologist, for suspected toxoplasma retinitis, administration of intravitreal steroids and clindamicine was begun with partial benefit. however days after (d + ) she was admitted in hospital because of worsening headache, irradiated in the occipital area, and weakness in the right hemibody. tests on csf were negative for neurotropic pathogens. an mri showed a complete occlusion of the intracranial tract of left internal carotid artery, with likely infectious material localized in the left lateral cerebral fissure. a chest tc showed a nodule with initial excavation in the right superior pulmonary lobe. for suspected tuberculosis she started antitubercular therapy. despite a second lumbar puncture confirmed pleocytosis compatible with acute purulent meningitis, microbiological research for bacteria, fungi and bk were negative. so antitubercular and antitoxoplasma therapy were stopped and the patient underwent surgical biopsy within the sphenoid sinus. pathological examination of the biopsy specimens showed acute and chronic inflammation of the respiratory mucosa, periodic acid. schiff and grocott staining ( figure ) highlighted several septate fungal hyphae. cultural analysis revealed colonies of scedosporium apiospermum so the patient started targeted voriconazole intravenous therapy. nevertheless, days later, she developed aphasia and right hemiparesis. a brain angio-mri confirmed the appearance of new lesions compatible with infectious localizations associated to an increased defect of left internal carotid artery vascularisation and complete left choroid detachment. after weeks of voriconazole a significant clinical improvement have been observed and she was discharged, continuing oral antifungal therapy with voriconazole. at the last follow-up she achieved a complete resolution of neurologic symptoms, with permanent left eye blindness. months later (d + ) she was asymptomatic, with normal haematological and neurological conditions and was able to stop the antifungal therapy. this case-report confirms that the risk of invasive fungal infection (ifi) is relevant in patients receiving hsct for aa, probably due to the prolonged neutropenia and association of other risk factors such as the immunosuppressive therapy and the iron overload. in this very poor prognosis infection, the early diagnosis of cns ifi remains challenging, but the administration of voriconazole was extremely effective. disclosure of conflict of interest: none. in this study, we aim to present the seroprevalence of ebv and incidence of posttranplant lymphoproliferative disease as well as to evaluate the relation with gvhd. between and , the ebv serology of patients that underwent allogeneic hematopoietic stem cell transplantation and their donors were evaluated in the study. ebv ig g (vca-igg, ebna ig g, ea-igg) and igm (vca-igm) antibodies were detected by chemolluminesance method (abbott, abd). all patients were followed for reactivation. ebv igg seropositivity was detected in patients ( %) and donors ( . %). there was no statistically difference in related vs unrelated transplants in seropositivity. the median age of the patients was (range: , patients were male ( %) and ( %) had malign disease. the stem cell source was peripheral blood in ( %) patients and ( %) received grafts from related donors. myeloablative conditioning regimen was received by of patients ( %) (table) . all patients received acyclovir prophylaxis (related transplants mg tid, unrelated transplants mg tid) during and after allo-hsct up to months. twenty six-yearold pretransplant ebv seropositive aplastic anemia patient had ebv ig m positivity after months of allo-hsct and developed lymphoproliferative disease. he was in complete remission after courses of rituximab and methylprednisolone. three patients were ebv igm seropositive in th, th and th months of allo-hsct and received symptomatic treatment. acute gvhd was detected in patients ( %) whereas patients ( %) had chronic gvhd. acute gvhd and chronic gvhd incidences were similar in comparison of donor ebv seropositive vs seronegative status ( % vs %, p = . ; % vs %, p = . ). ebv seropositivity was detected in . % of patients. the donor ebv serology was not related with acute or chronic gvhd. [p ] disclosure of conflict of interest: none. the umc utrecht pediatric experience with brincidofovir after allo hsct ca lindemans, m bierings and jj boelens pediatric blood and marrow program, dept. of pediatrics, university medical center utrecht, the netherlands viral reactivation with dna viruses form a considerable complication of allogeneic hematopoietic stem cell transplantation (hsct). there are little effective antiviral therapies and most have considerable toxicity. especially for adenovirus, there is no satisfactory therapeutic option. recently a new oral antiviral agent, the cidofovir prodrug brincidofovir became available to european patients only on the basis of urgent medical need and after a case by case approval by the health authorities. the aim was to describe our single center experience with brincidofovir in the pediatric allogeneic hsct setting. in the umc utrecht, pediatric patients receive t-replete bone marrow or unrelated cord blood (ucb) as the donor source after mostly myeloablative conditioning regimens (+ serotherapy in unrelated-hct). as gvhd prophylaxis patients receive cyclosporine a (csa) and mtx for bone marrow, csa and prednisone for ucb. patients are by standard weekly monitored for the presence of adenovirus, ebv, cmv en hhv viremia by rt pcrs in the plasma. extensive immune reconstitution measurements are performed every weeks. since , patients that developed viral reactivation with adenovirus, or a combination of other dna viruses (cmv, bk or hhv ) were offered brincidofovir if the viremia was progressive or in the context of poor immune reconstitution. brincidofovir was given in suspension ( mg/ml) at the dose of mg/kg biw, or mg biw for larger children. de drug was discontinued when the viral load was below detection level. in total, six pediatric patients (age range: - ) received brincidofovir ( patients tablets, the suspension). four received it for adenovirus reactivation, a th patient for cmv and bk and a th patient for cmv en hhv . the median day post-hsct of the first administration was days post hsct (range: − to ), the median day post detection of viral reactivation days . the median duration of administration was days ( - ) with two patients being discontinued because of death. in no patient the drug was discontinued due to toxicity issues. the patients that died had multi-organ failure due to a combination of severe agvhd and multiple infectious issues. the patients were discontinued when the viral load was low and when they had cd counts of at least /μl. none of the four alive patients reactivated after the drug was discontinued. urgent medical need administration of brincidofovir is feasible. in our limited series we found the drug was well tolerated. disclosure of conflict of interest: i am a medical consultant for brincidofovir (chimerix). reactivation of herpes simplex virus (hsv- ) or varicellazoster virus (vzv) occurs frequently after allogeneic stem cell transplantation (asct). here, we report three unusual cases, two with reactivation of hsv- and one with vzv. patients and methods: patient (pt) ( -year-old, male) was allografted for high risk acute lymphoblastic leukemia in first complete remission after conditioning with total body irradiation ( gy) and etoposide ( mg/kg). graft-versus-host disease (gvhd) prophylaxis was performed using cyclosporine a, short course methotrexate and anti t-lymphocyte globulin (atg). pts ( year-old, female) and ( -year-old, male) were allografted for acute myeloid leukemia in second and first complete remission, respectively. conditioning regimens used were flamsa-ric in pt and fludarabine/busulfan in pt . in both cases, gvhd prophylaxis consisted of cyclosporine a, mycophenolate mofetil, and atg. pts and had already experienced hsv- -positive oral mucositis following induction chemotherapy and had successfully been treated with acyclovir. both developed hsv- -positive oral mucositis again after asct. in both cases, initial therapy with acyclovir i.v. at a dose of up to mg/kg t.i.d. was ineffective. to explore the mechanism leading to clinical acyclovir resistance, the thymidine kinase genes of both viral strains were sequenced. pt presented with severe abdominal pain and nausea months after asct. in this case, acyclovir prophylaxis post asct had been stopped months before due to side effects. moreover, low dose prednisolone therapy was necessary for chronic gvhd. the hsv- -strain from pt showed a single base pair deletion in the region from nucleotide position to of the thymidine kinase gene (which consists of a guanosine repeat). in pt a single base pair insertion in the same region was found. both genetic alterations lead to a loss of enzyme activity and acyclovir resistance. in both pts treatment was changed to foscarnet which led to rapid improvement. in the case of pt , multiple mucosal erosions were found on endoscopy of the esophagus. in these vzv dna was detected by polymerase chain reaction (pcr). only days later, a vesicular skin eruption developed, which did not follow a dermatomal distribution. again, in the vesicular fluid vzv dna was detected by pcr. in this patient, acyclovir ( mg/kg i.v., t.i.d.) resulted in rapid improvement. reactivation of hsv- and vzv after asct is a frequent finding. usually, hsv- strains respond well to acyclovir. in some cases, resistance can develop, especially in patients that had been treated with acyclovir before. acyclovir resistance of hsv- caused by mutations in the thymidine kinase gene can be overcome by treatment with foscarnet which directly inhibits the viral dna polymerase. disseminated vzv reactivations after asct have been described. clinical presentation can be misleading, for example, beginning with severe abdominal pain that precedes the vesicular eruption by several days. disclosure of conflict of interest: none. toxoplasmosis is a rare but severe complication after hematopoietic stem cell transplantation (hsct) ( ) . it can involve the central nervous system alone or can manifest as a disseminated disease. in the paediatric population the mortality rate is high and sequelae are often severe. new diagnostic tools, such as the pcr assay, may allow for rapid diagnosis and preemptive therapy ( , ) . we retrospectively analysed all children who underwent allogeneic hsct in our centre between january and december . patients lost to follow up before day + were excluded. patients and donors were tested before transplant in order to assess their immunological status against t. gondii. a total of allo-hsct were analysed. before transplant, . % of recipients (r) were toxo-igg positive and . % were toxo-igg negative. among donors (d), serology was available only for / : % were toxo-igg positive, % were toxo-igg negative. we found a high number of not tested donors ( . %, / ) which included, in most cases, mud from foreign registries. the group at higher risk for toxoplasmosis, d − /r+, included . % pairs, whereas d − /r − were . %, d+/r-were . % and d +/r+ were . %. in our series the cumulative incidence of toxoplasmosis disease was . %, with cases out of transplants. two of them (case and ) had cerebral toxoplasmosis, one (case ) had disseminated toxoplasmosis and case had toxoplasmic chorioretinitis. mortality rate was %: two patients died because of multiorgan failure and disseminated toxoplasmosis respectively. in no case localized cerebral toxoplasmosis was the main cause of death. no complications were seen in surviving patients. all patients who developed toxoplasmosis were toxo-igg positive before hsct and three of them were transplanted from a toxoplasma igg negative donor (fourth donor not tested). in the two fatal cases the interferon-gamma releasing assay (igra) never became positive, confirming the absence of specific cellular immunity. toxoplasmosis disease can affect hsct outcome in paediatric recipients and pre-hsct seropositivity is the most important risk factor for toxoplasma disease in the post transplant period. in our cohort seroprevalence was higher than expected, probably due to the high number of patients coming from eastern europe. in order to reduce the burden of toxoplasmosis disease in our population we decided to implement a real-time pcr screening protocol for d − /r+ pairs, to provide rapid diagnosis and early therapy. all positive recipients with a seronegative donor will undergo real-time pcr screening starting on the day of stem cells infusion, and regularly until cd + t cell recovery. in the future we will analyse the impact of this strategy in this particular subset of immunocompromised patients. treatment with brincidofovir for adenovirus disease in pediatric hematopoietic transplants introduction adenovirus may cause serious morbidity and mortality after allogeneic hematopoietic transplants in children. severe lymphopenia is the main risk factor associated with progression to disseminated and often fatal disease. treatment with unlicensed cidofovir is based on monitoring of plasma viral load by pcr. however, cidofovir is only moderately effective at controlling adenovirus and it is associated with significant renal toxicity. brincidofovir is a lipid conjugate of cidofovir. it has a good oral bioavailability and achieves higher intracellular levels of active drug than cidofovir with a better safety profile. it is a potent inhibitor of viral dna synthesis so it could be indicated in immunocompromised patients with adenovirus disease. patients and methods we present three children of , and years old diagnosed of acute lymphoblastic leukemia (all) in nd complete remission (the first two patients) and severe aplastic anemia the last one. there were girls and boy. they underwent a peripheral blood hematopoietic stem cell transplantation using αβ/cd depletion with a haploidentical donor in the two patients with all and cd ra depletion with a matched unrelated donor in the other patient. patients that underwent haploidentical transplants developed early acute graft versus host disease grade iii with gut and skin involvement so immunosuppressive treatment with corticoids was started. they developed severe lymphopenia ( o / mm ). in the first month after transplant an adenovirus disease was diagnosed in the three patients from the weekly monitoring of plasma viral load by pcr. adenovirus was also tested in stools, urine and respiratory sample. in all patients adenovirus was also detected in urine sample. in one of them adenovirus was detected in nasal exudate too and in the other the virus was isolated in stools and in a skin biopsy. results: all of them were initially treated with cidofovir with poor results. foscarnet and gancyclovir was also used without improvement. finally they started a treatment by compassionate use with oral brincidofovir twice a week. with the first dose of brincidofovir plasma viral load started to go down until its complete disappearance. brincidofovir tolerance was good with only mild and limited diarrhea in two cases in the day they were taking brincidofovir. two of the three patients were alive without signs of adenovirus disease. in the other patient blood adenovirus load by pcr decreased below /ml, but remain high in urine. she died of respiratory failure due to pulmonary graft versus host disease. conclusion brincidofovir may be a promising therapeutic option for the treatment of severe adenovirus disease in immunocompromised patients with a good toxicity profile. disclosure of conflict of interest: none. table . all patients were transplanted with pbsc for haematological malignancy, and s received reduced intensity conditioning (ric) regimens with in vivo t-cell depletion. the proportion of patients with baseline and post-vaccination hi titres ⩾ : were . and % for a(h n )pdm , . % at both time points for a (h n ), and . and % for b/phuket. pre and postvaccination geometric mean titres gmt) were higher by mn than hi for a(h n )pdm and a(h n ), but lower for b/ phuket (p = . ). no post-vaccination seroconversions were detected by hi, while a single seroconversion to a(h n ) pdm was detected by mn in a patient vaccinated at - months. the mn assay did not detect any additional low-titre seroresponses (negative to detectable titre) below hi threshold. none of patient age, lymphocyte count, days from transplant to vaccination, donor type, and gvhd or ist at vaccination correlated with baseline or post-vaccination titres by either assay. response to iiv was virtually absent throughout the first year post-hsct, with a single seroconversion to a(h n )pdm detected by mn but not hi, although the sample size was small and half of patients were vaccinated at - months. there is a clear need for a novel, immunogenic seasonal iiv and/or novel vaccination regimens in this population. vaccination of recipients' relatives and close contacts, and hsct healthcare workers should be strongly encouraged. pre-and post-transplant iron overload (io) has been associated with considerable long-term morbidity and mortality in pts undergoing transplantation. classically, management of io in the post-allo-hsct setting has been based in the performance of therapeutic phlebotomies (tp), which are inconvenient for the patient and are often not feasible due to ongoing anemia. we recently published the first prospective study of deferasirox in adult allo-hsct pts with io (vallejo, et al. haematologica ). in this retrospective analysis, we analyzed the real-life management of io in the post-allotransplant setting. this study includes the last pts with a minimum follow-up of weeks, who underwent allo-hsct in our center (october -october ). pts were male ( . %) and female ( . %). median age was years (range: - ). baseline diseases were: aml ( . %), lymphoproliferative disorders ( . %), mds ( . %), all ( . %), chronic myeloproliferative diseases ( . %), mm ( . %), and bm failures ( . %). donor was unrelated in cases ( %; of them hla mismatched), and related in ( %; of them haplo-identical). conditioning regimen was: busulphan-based ( . %), melphalan-based ( . %), tbi-based ( . %), and others ( . %). progenitors source was pb in ( . %), and bm in ( . %). pre-hsct: pts had been transfused with a median of prbc (range: - ), and their median serum ferritin (sf) was ng/ml (range: - ). day + post-hsct: pts had died, and pts had not reached that day yet, so pts were evaluable. they had been transfused with a median of prbc (range: - ), and their median serum ferritin (sf) was ng/ml (range: - ). % pts had sf superior to ng/ml. liver mri (by sir method) to assess liver iron concentration (lic) was performed in pts at day + . seven pts ( . %) had no io (lic - mg/g), pts ( . %) had moderate io (lic . - . mg/g), and pts ( . %) had severe io (lic superior to . mg/g). median lic was . mg/g (range: . - . ). among the cases with history of more than prbc transfused and sf higher than ng/ml at day + , ( . %) were proved to have liver io by mri; the other pt had io in spleen. pts started some kind of therapy to treat the io: pts with severe io initiated a tp program and pts ( out of with moderate io, and out of with severe io) initiated chelation therapy with deferasirox. the drug was started at low dose ( . - mg/kg/ day), and was increased if tolerated up to a maximum of mg/kg/day. of note, the majority of pts were also taken a number of medications (immunosuppressants, statins, antimicrobials, etc). of those pts ( . %) did not tolerate the drug, and were changed to tp. for more details, see the table. ( ) the combination of the history of prbc transfusions and serum ferritin levels was, in the majority of cases, enough to assess the io in the post-allo-hsct setting. ( ) liver mri (by sir method) helped to assess io in doubtful cases. ( ) deferasirox, initiated at low doses and increased if tolerated, was safe and its use helped to avoid the need of therapeutic phlebotomies for the majority of patients. this study reproduces, in a real-life setting, our previous findings in a prospective clinical assay. [p ] disclosure of conflict of interest: none. a case-control study of risk factors of primary graft failure with a focus on associated early-onset severe infections v alcazer , a conrad , f-e nicolini , s ducastelle-lepretre , f barraco , x thomas graft failure (gf) is a rare but devastating event after allogeneic haematopoietic stem cell transplantation (ahsct), exposing the recipient to disease relapse, drawbacks of marrow aplasia, infections and death. the aim of this study was to analyse the risk factors associated with graft failure after ahsct, with a specific focus on early-onset severe infections (esi). we conducted a retrospective, observational, single-centre, matched case-control ( : ) study among adult s ahsct recipients transplanted at the haematology department of our institution between and , with a subsequent follow-up of months. engraftment was assessed at day+ post-ahsct. gf cases were classified as primary gf (pgf), defined as failure to achieve donor-derived absolute neutrophil count (anc) ⩾ . × /l or lasting more than consecutive days without evidence of disease relapse and early-secondary gf (esgf), referring to the loss by day post-ahsct of a previously functioning graft associated without evidence of disease relapse. each case was matched with two controls according to underlying haematological disease, hla matching, stem cell source, intensity of conditioning and temporal proximity of ahsct. demographics, haematological and graft characteristics as well as esi report were retrieved. esi were classified in invasive fungal infections, viral infections (cmv, ebv, hhv- , other viruses), toxoplasmosis and severe sepsis of bacterial origin. during the study period, ahsct were performed at our center. seventeen ( . %) gf cases were identified, of which pgf and esgf, and were matched with controls. in the descriptive analysis, gf and control populations did not significantly differ when considering demographics, haematological characteristics and hematopoietic stem cell source. regarding pretransplantation status and graft characteristics, only disease status (progressive disease) and cell dose (both cd + and cd + cells number/ kg) were associated with graft failure. the proportion of patients with ⩾ esi before day was significantly higher in cases than in controls ( / vs / , p = . ), with an overall number of esi events of and among cases and controls, respectively. five cases had ⩾ concurrent esi. the median time from ahsct to the first esi event for gf cases was days (interquartile range (iqr), - ) vs (iqr, - ) days for controls (p = . ). in the gf setting, the most prevalent infections were herpesviridae infections (n = including hhv- n = , ebv n = , cmv n = ), probable ifi (n = ), severe sepsis of documented bacterial origin (n = ), toxoplasmosis (n = ) among whom one patient developed haemophagocytic syndrome. when further analysing subsets of esi using logistic regression, only toxoplasmosis was a significant risk factor for gf (p = . ). death related to an infection was proven for gf patients vs control patients (p = . ). the overall survival probability at months was significantly lower in the gf setting than in control patients (hr = . ( % ci . − . ), p = . ). the survival rates at months were . % and . % for gf and control patients, respectively. at our center, graft failure is statistically associated with early-onset severe infections, and already known graft characteristics such as cell dose and disease status. however, our study would need more power to increase its significance. disclosure of conflict of interest: none. allogeneic stem cell transplantation (asct) is a curative option for hematological disorders, especially malignancies. in immunosuppressed women after asct, the progression from cervical dysplasia to invasive carcinoma is accelerated, and cervical cancer is likely a more aggressive disease. therefore, follow-up protocols after asct should include regular gynecologic evaluation with papanicolaou (pap) smears. we retrospectively evaluated pap smears in women who underwent asct and searched the risk factors for abnormal cervical cytology. the median age at transplantation was . years (range: - years). the most frequent indication for asct was leukemia ( %), and % of the patients received a transplant from a sibling hla-matched donor. stem cell source was peripheral blood in all patients. myeloablative conditioning regimen was used in % of patients. cyclophosphamide, busulfan and fludarabin were used in ( %), ( %) and ( %) patients, respectively. acute graft versus host disease (gvhd) occurred in patients ( %) and chronic gvhd in patients ( %). secondary cancer ( breast cancer) was reported in only one patient at months after asct. the follow-up time was months (range: - months). after asct, benign and abnormal pap smears were found in ( %) and ( %) women, respectively. the median time between asct and development of abnormal cytology was months (range: - months). four ( %) women had at least one smear with atypical squamous cells of unknown significance (asc-us), one ( %) had a low-grade squamous intraepithelial lesion (lsil), one ( %) had atypical squamous cells/high-grade lesion (asc-h) and one ( %) had asc-us and asc-h. one ( %) patient had malign smear. two patients with asc-h showed high-grade atypia mimicking cancer but had a negative follow-up. patient who had malign smear died because of aorta dissection. cervical biopsy showed cervical intraepithelial neoplasia (cin) i in ( %) women who had asc-us or asc-h. one patient was hpv-positive. we did not find any relationship between cervical cytological abnormality and clinical factors. after asct, patients are high risk for abnormal cervical cytology and secondary gynecological cancer. regular surveillance of patients is the most important factor for decreasing the risk of developing cervical and other secondary cancers. gynecologic examinations and cervical cytological testing after asct allows early diagnosis and effective management of cervical abnormalities. disclosure of conflict of interest: none. kidney dysfunction is a frequent complication of allogeneic stem cell transplantation (sct) and contributes to the morbidity and mortality of the procedure. incidence of severe acute kidney injury (aki) in patients undergoing nonmyeloablative allogeneic sct for malignant diseases ranges from to %. lymphoma patients are often heavily pretreated through both chemotherapy and autologous sct and may be at increased risk of developing kidney injury. we performed a retrospective analysis of consecutive patients with lymphoma undergoing nonmyeloablative allogeneic sct between and (table ) . acute kidney injury (aki) within days of allogeneic sct was diagnosed and staged according to rifle-criteria, and severe aki was defined as rifle stage i-e ( doubling of creatinine or % decrease of egfr). chronic kidney disease was defined as an estimated glomerular filtration rate (egfr) o ml/min/ . m year after allogeneic sct. we performed multivariate logistic regression to evaluate potential risk factors for severe aki. severe aki developed in patients ( . %). reduced overall survival was observed in these patients, although not statistically significant. no significant associations were seen with age at transplantation, baseline kidney function or prior autologous sct. severe aki was associated with acute graft versus host disease (gvhd) (or . , p = . ) and the use of an unrelated donor (or . , p = . ). chronic kidney disease was observed in ( . %) of patients alive after year. we report a substantially higher incidence of severe aki after nonmyeloablative allogeneic sct for lymphoma than has been reported for other malignancies. acute gvhd and unrelated donor stem cell s source were associated with severe aki, while prior autologous sct, age and baseline kidney function were not. [p ] disclosure of conflict of interest: none. patients with acute myeloid leukemia who are treated with conventional chemotherapy still have a substantial risk of relapse. we, therefore, retrospectively analyzed data to investigate the effects and some risk factors of allogeneic hematopoietic stem cell transplantation in relapsed and refractory acute myeloid leukemia patients, and to provide some suggestion for the clinical treatment. a total of refractory and relapsed acute myeloid leukemia patients receiving allogeneic hematopoietic stem cell transplantation in our center between february and december were retrospectively analyzed, including patients in no-remission (nr) and patients in second complete remission (cr ) at the time of transplant. the median age was years (range: - ). conditioning was myeloablative using cyclophosphamide, busulfan and total-body irradiation (bu/cy, n = ; tbi/cy, n = ), and others were underwent nonmyeloablative stem cell transplantation. patients had successful engraftment. acute-gvhd and chronic-gvhd appeared in and patients. the year overall survival (os), relapse rate and disease-free survival (dfs) of the cases was ± . %, . ± . % and . ± . %, respectively. the -year dfs were higher for patients in cr patients ( . ± . %) than in nr patients ( . ± . %), and the relapse rate in nr group and cr group were . ± . % and . ± . % respectively. there was no significant difference in treatment-related mortality compared cr group with nr group. sex, age, related-donor graft were not independent factors affecting os, dfs and relapse rate. it is concluded that allo-hsct is an effective salvage therapy for patients with refractory and relapsed aml. non-remission before transplant and severe agvhd are high risk factors of poor prognosis for allo-hsct. patients in cr group who accept reinduction chemotherapy before transplantation have better prognosis than those in nr. the overall outcome seems related to the disease status. hsct during refractory and relapsed can achieve long-term survival in selected patients with individual therapy. disclosure of conflict of interest: none. the incidence of most hematologic malignancies increases with age. aging is related with a greater prevalence of impaired functional status and comorbidities. although cure of malignant and non-malignant hematological diseases is potentially possible with allo-hsct, it could lead to significant transplant-related mortality. decision making about referral to allo-hsct in older adults is a challenging task. in this study we aim to present our geriatric allo-hscts. from to , [p ] patients (age ) underwent allo-hsct in our center included to this retrospective study. pre-transplant status as well as posttransplant toxicities, complications and outcomes were determined. the age distribution of the group: patients was aged and o , patients was aged and o , patient was years old. the median age of donors was (range: - ). the pre-transplant patients' characteristics are given in the table. remission was achieved in twenty-three ( %) patients. twenty-six patients ( %) had neutrophil engraftment ( . × /l) at a median day of (range: - ) and platelet engraftment ( × /l) at a median day of (range: - ). post-transplant complications are detailed in the table. acute graft vs host disease (gvhd) was occurred in patients ( %) and chronic gvhd in patients ( %). eight patients ( %) were diagnosed with a relapse and year relapse-free survival was %. the -year and -year os were detected as % and %. the most common reason for mortality was sepsis. the -year os was higher in patients who had reduced intensity conditioning regimen and remission status pre-transplant however they were not statistically significant ( % vs %, p = . ; % vs %, p = . ) (figure) . since increasing number of older patients being diagnosed with hematologic malignancies, this trend of increasing number of allo-hsct will continue. tolerability and effectiveness are lesser, toxicity is higher in older adults. although study population is relatively small, reduced-intensity conditioning and pre-transplant remission status may be related to better survival. comprehensive geriatric assessment may be considered prior to allo-hsct for global evaluation. disclosure of conflict of interest: none. allogeneic hematopoietic stem cell transplantation (asct) is a procedure with high morbidity and mortality ( - %) requiring a complex hospital infrastructure. improved support measures and development of homecare units has allowed that asct at-home programs may be possible. our center has launched a pioneering program in our country in patients with asct to perform at home the following of aplasia, control of immunosuppressive therapy (ist) and intravenous support from the d+ of asct until the engraftment and independent ambulatory patient. to evaluate the patient safety, we compared the group of patients at-home (asct-op) with a cohort of asct 'in patient' with similar characteristics (asct-ip). asct patients between january and october at the hospital clinic of barcelona. patients performed asct-op and had an asct-ip. all patients received conditioning (myeloablative-mac-or reduce intensity-ric-) in the hospital with fludarabine mg/m (d - ) and busulphan . mg/kg ( - doses), prophylaxis of gvhd was performed with tacrolimus/mycophenolate (mmf) in asct-op group and cyclosporine(csa) and methotrexate (mtx) or mmf in asct-ip group. in all patients, the infectious prophylaxis was conventional (levofloxacin, fluconazole and acyclovir). moreover, the asct-op group received prophylaxis with ceftriaxone g intravenous (iv) once daily and liposomal amphotericin b inhaled mg twice a week during neutropenia. the asct-op group from d+ received a nurse visit once daily and physician visits twice a week in the hospital. baseline characteristics were analyzed those related to toxicity and patient outcomes. the median age (range) was years ( - ), male/female / ; ( % male). the source of the progenitors was peripheral blood in all cases and analysis of the results detailed in the table: disclosure of conflict of interest: none. an increase in rdw-sd after allogeneic hematopoietic transplantation is associated with a poor prognosis s leotta, a cupri, a di marco, a spadaro, l scalise, g sapienza, mg camuglia, g avola, g moschetti and g milone istituto oncologico del mediterraneo red cell distribution width (rdw), is an erythrocyte index influenced by stress erythropoiesis, inflammation and antioxidants. rdw predict mortality in sepsis, chronic kidney diseases and in cardiovascular disease. no data are available on rdw after hematopoietic transplantation. in a retrospective study we collected data on changes of rdw-sd in a group of patients who received allogeneic hematopoietic transplantation. fortyeight patients were affected by acute leukemia, by lymphoma, by mm, and by other diagnosis. rdw was studied at baseline and monthly for the first months. a subset of patients were studied prospectively for clinical and laboratory signs of microangiopathy. at baseline before the transplant a rdw-sd higher than normal upper limit was observed in % of allogeneic candidates. a high co-morbidity score (htc-ci score - ) at the pre-transplant screening was a factor associated to high rdw-sd (χ p = . ). a value of rdw-sd higher than normal range, at baseline, was not associated to any other factors, such as age, diagnosis, phase of the disease, previous transplantation, c-reactive protein, bilirubin, creatinine and arterial hypertension. early after allogeneic transplant we noticed at day + a significant reduction of rdw-sd but subsequently (at day + ) the proportion of patients showing an abnormal rdw-sd increased to %. an abnormal rdw-sd at s day + was registered in % of allogeneic transplant patients who presented an acute gvhd while in only % of patients who did not presented during the first months an acute gvhd (χ p = . ). in allogeneic transplantation group, patient who, at day + , had a rdw-sd higher than normal value had a inferior outcome in respect to patients having a rdw-sd within normal ranges (os was % vs %; logrank: p = . ;), (ci of trm: % vs %).these two groups were not significantly different for pretransplant features in the subset of patients studied prospectively, abnormal rdw-sd was associated to presence of schystocytes in pb (chi test: . ) and patients having ⩾ % schystocytes had a median rdw-sd of (iqr ) vs a median rdw-sd of (iqr . ) in patients who did not show schystocytes in pb (mann-whitney u-test p = . ). rdw-sd was significantly correlated also to serum triglycerides (r = + . , p = . ) and to red blood cell mean corpuscular volume (r = + . , p = . ). abnormal rdw-sd is frequent after allogeneic transplantation. abnormal rdw-sd is associated to acute gvhd and its value obtained at day + marks a group of patients with poor prognosis because of high trm. this simple parameter warrant further studies to determine its clinical usefulness in monitoring of patients suffering acute-gvhd and in diagnosis and monitoring transplant associated microangiopathy. [p ] disclosure of conflict of interest: none. sickle cell disease (scd) poses a lot of psychological burden for the patient and the caregiver. it also poses a significant financial burden over the family. ohaeri et al. developed a point questionnaire to asses sickle cell disease burden called as sickle cell disease burden index (scdbi) and its impact on caregiver's quality of life (qol). we used this questionnaire to assess the impact of hematopoietic stem cell transplant (hsct) on caregiver's qol. point questionnaire was sent to set of parents whose child underwent hsct between january and june . scdbi contained questions in various domains ( :family finances, :family interactions, :routine family activity and :parental coping ability). answers were graded on a score of - ( :never occurred and :occurred regularly or had a severe impact on the family). the results were interpreted in two headings a. family finances and interactions ( : no impact; - : insignificant impact; - : moderate impact; - : severe impact) and b. routine family activity and parental coping ability ( : no impact; - : insignificant impact; - : moderate impact; - : severe impact). all these domains were assessed before and after hsct. ten parents replied with duly filled questionnaire. mean age at hsct was . years (range: - ), m/f: / . all were symptomatic for months before hsct with % having more than hospital admissions. majority of parents were from middle class with median family income of usd per annum (range - usd). median score for family finances and interactions (a) before hsct was (range: [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] which decreased to (range: - ) after hsct. median score for routine family activities and parental coping ability (b) before hsct was (range: which decreased to (range: - ) after hsct. our results suggest that before hsct there was a moderate impact on family finances and interactions which reduced to no impact after hsct. similarly there was severe impact on family activities and parental coping ability before hsct which changed to no impact after hsct. our study suggests that hsct not only improves the qol of the child but also of the caregivers. chronic graft versus host disease (cgvhd) is a late complication of allogenic hematopoietic stem cell transplantation (hsct) that affects many tissues and organs and manifests with polymorphic clinical features similar to autoimmune diseases. poorly understood pathophysiological mechanisms are implicated in inflammation and tissue fibrosis which is a hallmark of cgvhd. the affection of lachrymal glands is frequent and contributes to ocular manifestations presenting as dry eye syndrome. autologous serum eye drops (aesds) are used topically to facilitate tissue healing and ease the symptoms in a variety of ocular diagnosis. it is unclear if the serum of a patient with cgvhd is suitable for remedy preparation and if the transplanted patient himself can meet the criteria for autologous donation. aim is to show the safety, feasibility and efficacy of autologous serum preparations in ocular lesions after allogenic hsct. donors should meet criteria for autologous blood donation (infectious disease status, complete blood count hgb g/l, hct %, adequate venous access). aesds are prepared from ml of autologous blood left to clot, irradiated and centrifuged to separate serum which is diluted with saline in : ratio or : if requested. product is dispensed into . ml ampules, stored at − °c and a -month supply is released to the patient after receiving negative results of sterility testing. in period from to . in the aesds program patients ( female, male) with ocular symptoms were included. all met required predonation criteria. of collections performed, one failed due to venous access problem and one product had to be discarded due to hemolysis. cgvdh global nih score of the patients at start of the program was: severe, moderate, mild and not scored. all patients presented with moderate to severe dry eye symptoms. in ( %) patients aesds alleviated dry eye symptoms. in ( %) out of patients referred to aesd program, more than autologous blood collections were performed (range: - ) and aesds were used regularly through period of - months, which points to the beneficial effect of the long-term use of the serum. three patients dropped out because aesds showed no advantage compared to commercial lubricant eye drops preparations. one patient dropped out because of a venous access problem, patients had disease s progression and needed other therapies: cases of amniotic membrane application of which continued with aesds to facilitate the healing effect. one patient was recently included and the effect of aesd is still evaluated. autologous donations in cgvhd patients are feasible, safe and autologous serum preparations can help relieve symptoms of dry eyes. it needs to be further elucidated specifically in which patients and at what point of the disease course the effect of the aesds is the most beneficial to make optimal use of these preparations. disclosure of conflict of interest: none. idiopathic pneumonia syndrome (ips) is a non-infectious pulmonary complication with diffuse lung injury that develops in - % of patients who undergo hematopoietic cell transplantation (hct) and the mortality rate remains high at % . the major aim of this study was to identify prognostic biomarkers for ips and establish positive and negative predictive values (ppv and npv) of ips. in a case-control study, we compared patients with ips with available samples (transplanted between and at fhcrc) with hct control recipients who did not require bronchoscopic examination and who did not grow any bacterial or fungal blood cultures. for each subject, plasma samples at day post hct and onset of ips or matched time points for controls were analyzed. the 'onset sample' for controls was the sample closest to day (median day of onset for patients with ips). we measured six proteins by elisa: suppressor of tumorigenicity (st ), tumor necrosis factor receptor (tnfr ), interleukin- (il- ), lymphocyte vessel endothelial receptor (lyve)- , endothelial protein c receptor (epcr), and herpes virus entry mediator (hvem). multivariable logistic regression models were used to evaluate the association of each protein with ips vs controls. cytokine cutoff values that maximized discrimination between ips and controls were identified using receiver operating characteristic (roc) analysis. ppv and npv of ips were calculated using the identified cytokine cutoffs across a range of hypothetical ips prevalence values ( - %) day weighted kaplan-meier survival curves were estimated for high/low cytokine subgroups. similarly, a weighted log-rank test was used to evaluate p-values. a multivariable logistic regression model including six cytokines showed that st and il- were significantly important markers to identify ips at the onset (table ) . st value at day post hct was significantly associated with occurrence of ips and il- had a marginal association. predictive values for ips by a plausible percentage of the actual hct population (up to %) are shown in figure . of the six proteins, st showed the highest ppv both at onset and day post hct followed by tnfr , and il- . npv were high in all the markers. to analyze whether st and il- at day after hct can predict survival following ips, we dichotomized the patients into cytokine high and low groups (cutoff level: st , ng/ml; il- , pg/ml) and compared survival after downweighting the observations to represent a plausible percentage of the actual population (ips prevalence, %). day survival rate were significantly lower in st high value group than in st low value group ( % vs %, p = . ). similarly, il- high value was associated with high mortality (day survival rate, % vs %, p = . ). st , il- , and tnfr were good prognostic markers for occurrence ips. especially, st and il- at day after hct can be a predictor for both ips occurrence and survival following ips. these results require validation in an independent prospective hct population. body composition parameters are sensitive nutritional indicators that influence response to treatment and mortality in cancer patients. research is not conclusive on the changes in muscle attenuation and adipose tissue areas in the stem cell transplantation (sct) phases. objective is to assess the changes in adipose tissues, skeletal muscle index (smi) and waist circumference (wc) among stem cell recipients in the peri-transplantation phase. study design: institutional review board approved this retrospective study with adult patients (age years) having b and t lymphoma who underwent sct. each patient was imaged by pet/ct scan pre-sct and months post transplantation. a cross sectional image was analyzed at the level of the l to calculate total adipose tissue (tat), visceral adipose tissue (vat), intra-muscular fat (imf), smi and wc. data was analyzed by gender since body composition parameters differed significantly between the two categories in the literature. the study sample consisted of patients (mean age: . ± . years, ( %) males, ( . %) autologous sct, median overall survival in months: . in males and . in females). death was observed in ( . %) males and ( . %) female. patient characteristics were similar for males and females except for weights (kg) and body mass index (kg/m ): . and . vs . and . in males and females respectively. changes from pre-sct to months post sct revealed that tat, vat, smi and wc decreased with mean differences of ± . cm , . ± . , . ± . cm /m and . ± . cm, respectively in males (po . ). in females, tat and wc significantly decreased with mean differences of . ± cm and . ± . cm, respectively (po . ). in females, vat and smi decreased clinically but did not reach clinical significance. in multivariate analysis, no significant associations were shown with mortality and progression rates. this study fills a research gap by providing data on the evolution of body composition parameters in the peri-transplantation phase. tat, vat, smi and wc decrease months post transplantation. future studies should evaluate the associations of these parameters with major outcomes on larger sample sizes. [p ] disclosure of conflict of interest: none. . patients received tbicontaining preparative regimen. all these patients were exposed to calcineurin inhibitors for prevention and treatment of gvhd. patients suffered from cgvhd-grade moderate or severe. all patients required systemic corticosteroids, because of gvhd ( pts) or during basic treatment of lymphoma ( pts). all patients had deficient states of vitamin d initially and required replacement. all of them, except for patients, had balanced adrenal insufficiencies and patients had balanced hypothyroidism. all women had premature ovarian failure ( received hrt). according to measurements of bone mineral density (bmd), low bone mass was detected in patients; osteopenia ( pts), osteoporosis ( pts). bone loss of femoral neck ( -osteopenia, -osteoporosis) occurred more often than lumbar vertebral ( -osteopenia, -osteoporosis) or radius ( osteopenia, -osteoporosis). presence of avascular necrosis of bone (avn), confirmed by mri, was detected in patients and the most common site of involvement was the femoral head(all patients), knee( pts) and shoulder( pts). one of the first symptoms of avn was pain and functional limitation. all patients required intensive analgesic treatment, usually nsaids and patientsfentanyl. fractures occurred in patients. the femoral neck ( pts) and thoracic or lumbar vertebral ( pts) were two most common fracture sites. all patients were qualified for surgery; patients required hip replacement, patients still awaited to perform surgery or were disqualified because of severe, skin cgvhd. bone complications may occur in about % of allo-hct survivors (including % patients with gvhd, and up to % patients with severe or moderate cgvhd) within first years after allotransplantation. bone loss, particularly at the femoral head, is the most common complication. avascular necrosis usually requires surgical intervention because of fractures. exposure to higher doses of corticosteroids (during treatment of gvhd) increases risk of bone complications. early diagnosis by mri and dxa may help to detect bone complications. ( %) and (e) ( %). a total of ( %) pts failed to meet these criteria but remained alive on day and ( %) died before day . the overall survival (figure ) for the pts was % at years with an overall mortality in icu of % ( / ) compared to % ( / ) for those who did not meet our criteria. the overall survival for pts that met our criteria at fifth day and were discharged to the haematology ward (n = ), was % at years. in this study, % of patients survived their icu admission. patients could be stratified according to the reason for admission and given an individualized -day trial: those who met our criteria for successful icu trial ( %) had a low icu mortality ( %) and those who were subsequently discharged home had a overall survival of % at years. this study raises the possibility of offering a short-term icu stay to oncohematologic patients and perhaps allows for the ceiling of intensive care for those who fail these criteria. [p ] disclosure of conflict of interest: none. in contrast, only . % of patients without cgvhd showed a thromboembolic complication in the later time course, with one patient showing an additional thrombotic risk factor. in multivariate analysis cgvhd was an independent risk factor for thromboembolic complications after hsct. . % of patients with thrombosis before hsct showed one afterwards. thrombosis before hsct was not found as risk factor for thromboembolic complication after hsct. our retrospective analysis showed an increased risk for thromboembolic complications after allogeneic hsct, with substantial higher risk in patients with chronic gvhd ( . %). in ongoing studies we currently investigate a vascular screening procedure with additional biomarkers according to inflammation and endothelial damage in patients with cgvhd prospectively. we hope to identify patients at risk for thromboembolism and prevent future complications on an individualized basis. disclosure of conflict of interest: none. ( ). pts with cns involvement received intrathecal therapy with cytarabine and in one case additional cns irradiation was applied. / pts died after a median time of mts (range: - mts) due to resistant systemic relapse, infectious complications or extensive graft-versus-host disease following allohsct. patient remains alive and disease-free at + mts following secondary allohsct. conclusions: our data indicate that em disease following allohsct affects a significant proportion of pts with aml. sites of em relapses vary widely among the pts with skin and cns being frequently involved. an aggressive approach combined of local and systemic therapy including secondary allohsct may produce favorable response in a small proportion of pts, however, overall prognosis for pts with isolated em relapses still remains poor. due to the lack of effective treatment strategies, there is a need for novel approaches to manage isolated em relapses after allohsct. disclosure of conflict of interest: none. hematopoietic stem cell transplantation (hsct)-associated thrombotic microangiopathy (tma) is a multifactorial complication, and has variable incidence in study populations due to different diagnostic criteria. aim: our aim was to identify pediatric patients with hsct associated tma using different diagnostic tma criteria published in literature and to compare the various groups for tma parameters and outcomes. we enrolled pediatric patients who underwent allogeneic hsct using treosulfan based or reduced intensity conditioning therapy. different tma diagnostic criteria, the bmt ctn toxicity committee consensus definition ( ), the overall thrombotic microangiopathy grouping ( ), the diagnostic criteria created by city of hope ( ) and the criteria proposed by jodele et al. ( ) were used to startify the patients. we determined and registered the following tma activity markers: presence or development of increased ldh and decreased haptoglobin levels, new onset anemia, thrombocytopenia, fragmentocytes, coombs test, kidney function, proteinuria, hypertension and terminal complement complex (sc b- ). complement pathway activities, components and sc b- were measured during early hsct period. two/ ( ), / ( ), / ( ) and / ( ) subjects met the different tma diagnostic criteria according to the four different systems on day and ( ) and on median ( ), ( ), ( ) post-hsct days. all of the / patients who were defined with the first three criteria, met the forth definition. due to normal haptoglobin levels and kidney function, / patients fulfilled only the forth criteria. tma coexisted with acute graft-versus-host disease in / cases ( / vs / ; p o . ). patients who met any of the different tma diagnostic criteria had higher sc b- level on day ( vs ng/ml; p = . ) compared to those without. all of the / subjects defined with tma had elevated sc b- ( ng/ml) level during the early hsct period. two patient died before day after hsct, out of which one patient met all of the four tma diagnostic criteria. after a median . ( . - . ) year follow-up time, overall survival was / . / patients with tma survived, compared to / patients without tma. relapse related mortality was the most common cause of death (n = / , po . ), while tma was not a significant cause of mortality after reduced toxicity conditioning therapy. hsct-associated tma has a variable and complex pathophysiology. using the different diagnostic criteria may influence the incidence and the time of diagnosis of this transplant-related complication. monitoring all of the published tma activity parameters, including complement terminal pathway activation marker, may help to guide physicians to recognise tma after hsct. disclosure of conflict of interest: none. hematopoietic stem cell transplantation (hsct) is associated with a risk of non-relapse mortality (nrm). it's important to assess the risk of complications and mortality before the hsct. some indexes quantify the impact of patients' comorbidities on hsct outcome. the most frequency used is the hct comorbidity index (hct-ci) and the european group for blood and marrow transplantation score (ebmts). this study tried to determine which of the two indexes best predicts the outcome in a series of patients submitted to hsct in a single center. between and , hsct were performed in our center. a total of hsct have been analyzed (we excluded patients o years (yr), nd hsct, haploidentical donors and hsct for specific diseases with very low number ( o %) of hsct performed: aplastic anemia, cll, prolymphocytic leukemia, mycosis fungoides, sezary syndrome, dendritic cell neoplasia, plasma cell leukemia and poems syndrome). the hct-ci and ebmts were calculated retrospectively (yr - ) and prospectively (yr - ). overall survival (os), relapse incidence (ri) and nrm were analyzed in the overall series and separately according to the type of hsct: autologous hsct (auto-hsct) or allogeneic hsct (allo-hsct). male: ( %) patients. median age: yr (range: - ). diseases: aml ( %), all ( %), mm ( %), nhl ( %), hl ( %), mds ( %), cmpd ( %). disease status: st complete remission (cr) or st chronic phase ( %), ⩾ second cr ( %), first partial remission (pr) ( %), ⩾ second pr ( %), no response ( %) and without previous treatment ( %). auto-hsct in patients ( %) and allo-hsct in ( %) patients. related and unrelated donor were ( %) and ( %), respectively. the conditioning regimen was standard in ( %) cases and reduced intensity in ( %). hct-ci and ebmts grouped - , and ⩾ were ( %), ( %), ( %) and ( %), ( %), ( %) in auto-hcct and ( %), ( %), ( %) and ( %), ( %), ( %) in allo-hsct, respectively. median follow-up was . yr ( . ; . ) for the overall series, . yr ( . ; . ) for auto-hsct and . yr ( . ; . ) for allo-hsct. significant differences in os and nrm were found according to the ebmts in patients submitted to auto-hsct. one-yr-os and -yr-os were % ( % ci: %; %) and % ( % ci: %; %), respectively, in patients with ebmts - , vs % ( % ci: %; %) and % ( % ci: %; %), respectively, in patients with ebmts ⩾ (p = . ). one-yr-nrm and -yr-nrm were % ( % ci: %; %) in patients with ebmts - , vs % ( % ci: %; %) in patients with ebmts ⩾ (p = . ). no significant differences were observed for ri according to ebmts in patients submitted to auto-hsct. no significant differences in os, ri and nrm were observed according to ebmts in patients submitted to allo-hsct. no significant differences regarding os, ri or nrm were found when the hct-ci was assessed. in our series, only the ebmts was predictive of os and nrm in patients submitted to auto-hsct. failure to find statistically significant differences for the hct-ci and for ebmts in allo-hsct recipients could be due to an insufficient number of patients or to a partial retrospective collection of data. infertility is common after hct predominantly as a result of the chemoradiotherapy used in conditioning. nonetheless, some patients do retain or recover fertility. newer reduced intensity regimens may be less gonadatoxic. in addition, patients are increasingly encouraged to store gametes, or embryos before transplant. we sent questionnaires to ebmt centers requesting retrospective details of number of pregnancies and pregnancy outcome for all patients treated between - . centers responded from countries detailing patients who became pregnant/partners conceived. the most frequent underlying diagnoses were acquired bone marrow failure (n = , f) aml (n = , f), hd (n = , f), cml (n = , f), all (n = , f) and b nhl (n = , f). other diagnoses included mds, mps, solid tumours, autoimmune disease, cll, t-nhl, haemoglobinopathy. of females (f), ( %) involved assisted reproductive techniques (art). f had tbi ( seven o gy) of which ( %) had art. f had reduced intensity conditioning of whom ( %) had art. f were specified as having standard conditioning of whom ( %) had art. f had allogeneic ( art, %) and f had autologous transplants ( art, %). of men (m) whose partners conceived, ( %) had art. m received tbi of which ( %) had art. where specified, had reduced intensity hct ( art, %) and had standard conditioning ( art, %) . had had allogeneic hct ( art) and autologous ( art). men had reduced intensity transplants. men received tbi (two o gy) of whom ( %) had art compared to men without tbi, ( %)of whom had art. data on return of menstruation was available for . indicated yes and ( %) had art. indicated amenorrhoea of whom ( %) had art. specifying number of children had live births (lb) and ( %) patients had more than one child after hct. lb occurred in female patients ( art, %) and lb were in partners of male patients ( art, %). the median gestational age for female patients was weeks (range: - ) and the median birth weight was kg (range: . - . ). there were / congenital anomalies. the median follow up of the offspring was y (range: - ). developmental problems were indicated for / (fine motor skills) and learning difficulties in / (adhd). in partners of male patients the median gestational age for offspring was weeks (range: - ). the median birth weight for offspring was kg (range: . - . ). congenital malformations occurred in / . one infant died of pulmonary infection. in women, several methods of assisted conception were used including hormone stimulation, ivf, cryopreserved embryos, donor embryos and cryopreserved ovarian tissue. the most frequent method was use of donor embryos ( / ) in which a minimum of attempts led to lb. the median number of attempts was (range: [ ] [ ] [ ] [ ] [ ] . art were frequently used in this group of posttransplant patients particularly in male patients vs female, tbi vs non-tbi, amenorrhoeic vs menstruating women, standard conditioning vs ric. in patients who conceive after hct, successful pregnancy leading to healthy offspring is the likely outcome. disclosure of conflict of interest: none. reduction of trm after sct was observed over the transplant periods and supportive care with danaparoid was found to be significantly effective to reduce trm. therefore, prophylactic administration of danaparoid is considered to be a reasonable option to improve the transplant outcomes for children. [p ] disclosure of conflict of interest: none. the attainment of transfusion independence after transplant is sometimes hampered by a combination of factors, ranging from infections to the need of combined therapy for clinical complications, as well as control of gvhd. iron overload is frequently observed in hematological patients before and after hematopoietic stem cell transplantation (hsct). whereas several reports have focused on iron overload before transplant, up to now, this is the only report that show full recovery of hematopoiesis and correlate this to deferasirox chelation performed on this particular subset of patients. we report on patients, transplanted for hematological diseases ( acute leukemia, aplastic anemia, multiple myeloma) heavily transfused before transplant that, considering the iron overload, were treated with deferasirox after hsct. before starting deferasirox, the patients were fully engrafted and in complete remission, although transfusion dependent, and with incomplete hematological reconstitution after allogeneic hsct. patients were selected according to the following inclusion criteria: ( ) transfused pre-transplant with more than rbc units; ( ) incomplete hematological recovery; ( ) transfusion-dependence; ( ) serum ferritin ng/ml; ( ) normal creatinine value. the workup for other aetiologies resulted negative. all patients received an initial dose of deferasirox mg/kg/day, later adjusted according to side effects. all patients experienced an increase in hemoglobin levels, with a reduction in the frequency of rbc transfusions, followed by transfusion independence (median time: days from the first dose of deferasirox). in addition, it was promptly (median time: days) associated with hematological improvement, with sustained values and no further platelet support or growth factors administration. no relevant modifications with immunosuppressive or myelosuppressive drugs were made during deferasirox treatment. deferasirox was well tolerated. basing on our results, we think that deferasirox determined stimulatory, and/or depressive effects on hematopoiesis after allo-hsct. this clinical experience raises the possibility of a potential additive benefit on hematopoiesis after transplant following iron chelation therapy with oral deferasirox. further long term studies, in larger cohorts of patients are needed to confirm these data and to design an efficient strategy to reduce iron loading after transplant. disclosure of conflict of interest: none. supported in part by ail pesaro onlus. ( ) hypertension (n = ). all five patients had normal adamts levels and negative testing for shiga toxin. complement mutation genetic studies were obtained for four patients including genes (n = ) and genes (n = ) and were all negative. testing for complement pathway including c b- were obtained for patients and were normal. all five patients were treated with eculizumab with induction treatment at mg weekly × doses, followed by one dose of mg on the fifth week, and mg every weeks thereafter. patients had a recovery of hemoglobin and platelets and a rise in haptoglobin and a normalization of ldh within - weeks from the start of eculizumab. eculizumab was discontinued for of the patients without recurrence of their tma; they are now - months since the discontinuation of eculizumab. in summary, there is a subacute syndrome of thrombotic s microangiopathy that can occur late post transplant. this syndrome appears to be complement mediated as shown by its response to a terminal complement inhibitor. it also appears to be transient without recurrence following treatment discontinuation. disclosure of conflict of interest: none. transplant associated microangiopathy (tam) is a very severe complication occurring after allogeneic bone marrow transplantation (bmt), burdened by a high case-fatality rate. it is characterized by abnormal complement activation, triggered by various agents (calcineurin inhibitors, acute gvhd, infections) with subsequent endothelial damage. in the literature, cases of mutations in recipient complement genes are described, but none in donor dna. here we describe for the first time patients affected by tam, carrying mutations in donor complement genes. in our lab, we studied patients affected by tam; they were screened for cfh autoantibodies, adamts function and variants and macro-rearrangements in cfh (and related), cfi, cfb, cd , c , dgke, thbd genes and at-risk haplotype (cfh-h and mcpggaac) by means of next-generation sequencing (ngs) and multiplex ligationdependent probe amplification analysis (salsa mlpa p armd mix- ; mrc holland). ngs was used to sequence dna by haloplex kit (agilent) on a miseq (illumina) platform with -fold coverage of every target base. the bioinformatic analysis was performed using sophia genetics and the pathogenicity was assessed by means of in silico predictions (polyphen , sift, mutationtaster, aligngvgd). all of the predicted pathogenetic variants were confirmed using sanger sequencing. the same genetic screening was extended also to donor dna in all cases. the screening for known causes of tam revealed mutations in recipient complement genes in one case; no mutations were found neither in recipient nor in donor dna in two cases; instead, donor genetic alterations were found in patients whose characteristics are summarized in table s donor hematopoietic cells. in the three cases presented, tam was relatively delayed with respect to hsct, in particular in two cases ( months) and this timing is compatible with the concept of reticulo-endothelial 're-population' by donor cells of monocytic lineage, responsible for the production of regulatory proteins of the alternative pathway of the complement. we also underline the response to anti-c inhibition in the patients who were treated with eculizumab; this fact further supports the hypothesis that the disease was related to complement dysregulation. we therefore suggest that both the recipient and the donor should be screened for complement gene mutations, so that more cases could be identified and the pathogenesis of tam could be further clarified. among these we observed one autologous engraftment, one death due to septic shock before engraftment and two primary gf. we used a desensitization treatment based on plasma exchange procedures, intravenous immunoglobulin ( g/kg) and rituximab ( mg/sm) in patients. one of these patients (aml, haploidentical donor) had dsa against hla-b (mfi ). she experienced primary gf with increasing titles of dsa (maximum mfi ); so, on day , a second transplant from the same donor was performed after a desensitization treatment. a progressive decrease in dsa was documented (up to mfi ⩽ ). on day patient achieved neutrophil count over /μl and on day platelet count over /μl. the second patient (mds, haploidentical donor), instead, received a desensitization procedure before the first transplant. she had dsa against hla-a (mfi ), and after desensitization dsa levels decreased and reached . on day patient achieved neutrophil count over /μl and on day platelet count over /μl. dsa were detected in / of usct candidates ( %) and / of haplosct candidates ( %) and they were associated with failure to obtain allogeneic engrafment in cases. desensitization treatment achieved dsa clearance and engraftment in the patients in which it was performed, underlining the potential benefit of this procedure in the setting of hsct with dsa that has to be validated by prospective and controlled studies. disclosure of conflict of interest: none. early complications and late effects and quality of life at myeloma multiplex patients z trajkovska-anchevska, a pivkova, s genadieva-stavrich, l chadievski, z stojanoski, l chevreska and b georgievski university hematology clinic, skopje, macedonia the subject of this research is the quality of life at patients with myeloma multiplex at diagnosis and during therapy within - months. the research aims to analyze patients to be able to continue activities which will contribute for improving their quality of life as a priority task placed before the patient, his family, health institutions and social environment. this research was conducted at the university clinic for hematology skopje in the period from june to march . it covers patients infected with multiple myeloma, diagnosed and treated during this period. a total of patients analyzed, using the eortc qlq c ver. . standardized questionnaires for hr quality of life that analyzed the physical, cognitive, emotional, personal and social functions related to the patients. it also analyzed and general health and quality of life. analysis of physical functioning at diagnosis is . during treatment . , significantly improved. personal functioning at patients at the diagnosis is . , during therapy − . . analyzing emotional functioning in patients at diagnosis is . , during the therapy over . significantly improved. in examining the cognitive functioning is also a significant difference at diagnosis . , during treatment . . social functioning of the patients was . at the diagnosis; during the treatment grow to . . significant improvement was notices in these patients' symptoms like fatigue, nausea and vomiting, pain, dyspnea, insomnia, loss of appetite, constipation and diarrhea. the analysis of the financial difficulties of patients at diagnosis is . and . during treatment, meaning no significant difference in the time given. the analysis of the overall health and quality of life at patient has a value of . , and during therapy . . quality of life at patients with myeloma multiplex that makes the research group was significantly improved as a result of on time diagnosis and treatment with modern medicaments and the role of social worker with the application of certain social skills, continuous counseling, guidance and education for their reintegration in the community. installing the quality of life as a separate category and investigating the factors that affect its expression in the daily functioning of the patients within the changed framework of action, as like this example for malignant disease. the needs of clearly defined interactions patient illness and treatment, quality of life and specifying the segments where it can effectively act and improve in order to achieve positive progression towards improving the qualitative features of this category is a clear and primary objective that must be inserted into the current approaches to monitoring patients with malignant hematological diseases. acute graft-versus-host disease (agvhd) is a common and severe complication after allogeneic stem cell transplantation. since the current first-line treatment is based on treatment with systemic glucocorticoids (gc), steroid-induced hyperglycaemia develops frequently in patients with (agvhd) potentially impacting on their outcome. we performed a retrospective analysis on patients who received systemic gc for agvhd and thoroughly investigated the consequences of aberrant glucose metabolism. in particular, we focused on glucose parameters early after initiation of gc. with a median of (range: - ) blood glucose measurements during gc treatment, increasing mean, median and maximum glucose levels as well as the need for insulin treatment were associated with decreased overall survival (os) in simple and multiple survival analysis. early hyperglycaemia, as defined by mean blood glucose levels mg/dl during the first days of gc therapy, was also found to be highly associated with adverse outcome: in multivariate analysis, the hazard ratio (hr) for death was . ( % ci . - . , p = . ) in patients with early hyperglycaemia. while the risk of death due to relapse was not increased, the hr for death due to non-relapse mortality was . ( % ci . - . , p = . ) in a competing risk analysis. a score based on early hyperglycaemia and non-response to gc within days allowed the identification of three risk groups: patients with both risk factors had an inferior os at years of . % as compared to . % in patients with none. patients with one risk factor had a -year os rate of . % (p = . for trend). in this retrospective study, we identified early hyperglycaemia after gc initiation as a prominent factor predicting increased nonrelapse mortality in agvhd patients. in addition, a score based on early hyperglycaemia and lack of response to gc was highly predictive for overall survival in these patients. disclosure of conflict of interest: none. early toxicity because of infectious complications not relapse is the main cause of death after allogeneic transplantation in aplasia for patients with refractory or relapsed acute myeloid leukemia high-dose cytarabin was given in / pts with induction failure. the search for a stem cell donor was started immediately after results of high-risk cytogenetic, no achievement of bone marrow aplasia on day of induction therapy, or immediately after diagnosis of relapse. four patients had a related / donor, for patients a / matched unrelated donor was identified and patients received a transplant from a / unrelated donor. the interval between diagnosis of primary disease or relapse and tx was ( - ) months (mo) for both groups . in patients melphalan ( - mg/m ) was used to induce an aplasia before starting conditioning therapy. the interval between melphalan and conditioning therapy was ( - ) days. three pts started the conditioning therapy while in aplasia after previous chemotherapy. the conditioning therapy was of reduced intensity in all pts. and consisted of treosulfan ( g/m )/fludarabin(flu) in pts, tbi( gy)/flu in pts and busulfan( m/(kg)/flu in pts, respectively.atg was given to all pts with an unrelated donor. most pts ( / ) had a severe neutropenia with a median of . /nl ( . - . ) before starting melphalan because of refractory leukemia. after a median follow-up of ( - ) mo pts ( %) were alive without relapse. ( %) pts died because of a relapse after a median of ( - ) mo. the nonrelapse mortality was % ( / pts). most of these pts ( / , %) died because of infectious complications early after transplantation (med ; - mo). in pts graft versus host disease was the main cause of mortality. in this retrospective 'real-life' analysis, we showed that an early allogeneic transplantation is feasible for patients with primary refractory or relapsed aml. a reduced intensity conditioning after induction of aplasia with melphalan offers a chance of long-term relapse-free survival for about % of patients with an otherwise dismal prognosis. nrm is high, especially because of infectious complications early after transplantation, probably related to the long period of severe neutropenia. therefore, the focus has to be set on early recognition and intervention of infectious complications. disclosure of conflict of interest: none. recent evidence supports the effector role of complement activation in several types of thrombotic microangiopathythe atypical hemolytic uremic syndrome (ahus) as well as the transplantation-associated thrombotic microangiopathy (ta-tma). the blockade of the terminal complement complex formation by anti-c monoclonal antibody eculizumab provides an effective treatment option in severe and devastating cases of ta-tma. the experience with the use of eculizumab in this indication is slowly accumulating in the hsct community, however the published data originate from small case series or uncontrolled trials and sharing of emerging real-life observations may be valued. on case reports of two pediatric patients treated with eculizumab for ta-tma with very detailed followup of multiple complement parameters, including terminal complex sc b- and eculizumab drug levels we would like to demonstrate: ( ) achieving therapeutic levels of eculizumab ( μg/ml) may be unsuccessful even with initially intensified dosing interval. furthermore, we documented rapid eculizumab clearance from circulation which allowed only for short periods ( o h) of efficient drug levels during the weekly dosing. ( ) we did not observe tightly correlated sc b- and eculizumab levels within the dosing intervals; however the long-term sc b- formation suppression was achieved concomitantly with improved eculizumab levels and slowed drug clearance. ( ) classical complement pathway activity assay (ch ) may not reliably substitute for therapeutic efficiency monitoring in case of hypocomplementaemia due to protein losses (profound diarrhea, proteinuria, gi bleeding, catabolism). this holds true also for the alternative pathway activity which remained low during treatment in both patients. ( ) mycotic infections may represent serious therapy related risks in eculizumab treatment after hsct (both patients achieved control of complement activation after multiple doses of eculizumab, however suffered fatal infections subsequently). besides, we observed a significant increase in c a concentrations correlated with clinical onset of infection which invites for further investigation of this complement cascade product as early indicator of mycotic infection. in conclusion, we would like to highlight the great added value of timely available complement assay results, including sc b- and especially eculizumab drug level values-to be used together with detailed clinical parameters for directing effectively these highly personalized (and also costly) treatments. [p ] disclosure of conflict of interest: none. table . no difference in terms of drug-related adverse events was observed in the three patient cohorts with no reported serious adverse events. similar results were obtained performing two separate sub-analysis only for lymphoma or myeloma patients. despite the limitations due to the non-randomized nature of the study, from our data on a large cohort of patients s with a long-term follow-up biosimilar filgrastim (zarzio®) could be considered substantially equivalent in terms of efficacy and safety to lenograstim (myelostim®) and peg-filgrastim (neulasta®), when used for hematological recovery and febrile neutropenia prophylaxis after asct in adult patients with hematologic malignancies. disclosure of conflict of interest: none. we studied all adults who underwent allo-hsct during a -month period ( january to november ) in our center. a total of pts ( . %) received epag for pfg with thrombocytopenia. three pts were male, and three female. median age was years ( - ). the baseline diagnoses were: alm ( ), mds-raeb ( ), idiopathic myelofibrosis ( ), aa ( ), and cll ( ). three transplants were from family donor (all of them haplo-identical), and from unrelated donor (the three of them hla / ). sc source was pb in cases, and bm in . epag was started at mg/day and escalated each weeks to , and mg if platelet count was o × /l. we analysed the platelets, anc, and hgb at epag initiation and days after being with the maximum dose. median time between allo-hsct and eltrombopag initiation was days ( - ). median maximum dose used of epag was mg/day ( - ). median platelets, anc and hgb before starting treatment were × /l ( - ), × /l ( . - . ) and . g/dl ( . - . ), respectively. five patients ( %) were severely thrombocytopenic (platelet count ⩽ × /l), ( %) were anemic (hbg o g/dl), and ( %) were neutropenic (anc o . × /l). median platelets, anc and hgb at day + of maximum dose were: × /l ( - ), . × /l ( . - . ) and . g/dl ( . - . ), respectively. the thrombocytopenic pts ( %) responded to epag, with increases of , , , and × /l in the platelet count. three anemic pts ( %) responded and achieved increases of hgb of . , . and . g/dl. finally, the neutropenic pts ( . %) responded and achieved increases of anc of and × /l. at the moment of the analysis close, pts are at a median of + . months post-hsct ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) , and all but one (who died from a septic shock) are alive and outpatient. this survival is striking for subjects who develop a complication with such a high expected mortality as pfg. pgf is a life-threatening complication, relatively frequent after alternative donor hsct, whose treatment has been very disappointed. we report our experience in pts who developed pgf during the last years. epag induced responses in platelets in all pts of the studied group. bilineal and trilineal responses were also seen. in our opinion, prospective studies are warranted in order to confirm epag as a new efficient treatment of post-hsct poor graft function. disclosure of conflict of interest: none. s ing patients who developed es with an equal number of patients who did not between january and november . we analyzed variables such as cd + cells per kg infused, use of granulocyte colony-stimulating factor (csf-g) and engraftment day. analytical data, including baseline and maximum determination of serum glutamic oxaloacetic transaminase (got) and glutamic pyruvic transaminase (gpt), c-reactive protein (crp) and procalcitonin (pct), as well as clinical data fever, weight gain, digestive and respiratory symptoms, pulmonary infiltrates were analyzed. sixty-eight patients were women. median age was years old (range: - ). patients were conditioned with beam ( %), melphalan mg/m ( %) and bcnu-tt ( %). nineteen patients developed es in our series, which correspond to eight percent of all asct. case and control groups were matched according to age, sex, diagnosis and conditioning regimen. the most prevalent baseline disease in the group with es was myeloma ( . %), followed by mantle cell lymphoma ( . %). all patients who developed es had fever, % skin rash, % respiratory symptoms, % pulmonary infiltrate an % digestive symptoms. a summary of the comparison of data analyzed in subgroups is shown in table . we found significant difference in the percentage of weight gain (p = . ), increase of tgo (p = . ), increase of tgp (p = . ) and increase the number of cd + cells per kg infused (p = . ), we found an inverse correlation between the number of cd + cells per kg infused and incidence of es. however, in terms of post-transplant csf therapy (p = . ) and crp and pct valor (p = . and p = . , respectively) we did not find significantly difference to develop es. in our series, weight gain and tgo and tgp rise were risk factors for es development. therefore, we should be aware of es in patients who develop fever, elevated liver enzymes and weight gain during graft phase. we did not find a significant difference in crp and pct suggested in other studies. further studies are required to better characterize risk factors of es development. busulphan-based ( ), melphalan-based ( ), beam ( ), tbi-based ( ), and others ( ). weight at hospital discharge was significantly lower than at admission ( . % in allo-hsct, and . % in auto-hsct). weight at day + was also significantly decreased compared with the admission ( . % in allo-hsct, and . % in auto-hsct). weight at day + was lower than the ideal for their sex and height in the allo-hsct setting. contrarily, among the patients undergoing auto-hsct, the weight at day + remained higher than the ideal for their sex and height in a high proportion of cases. regarding serum albumin, it was significantly decreased at discharge ( % in allo-hsct, and . % in auto-hsct), but recovered values similar to admission at day + . in the auto-hsct setting, prealbumin levels were significantly reduced at discharge ( %), and in lower proportion at day + ( %), compared with admission values. in the allo-hsct patients, prealbumin levels were significantly reduced at discharge ( %), but had been recovered at day + , compared with admission values. disclosure of conflict of interest: none. recently, blood and marrow transplant clinical trials network has proposed a composite endpoint: gvhd-free, relapse-free survival (grfs) for hsct outcomes. this endpoint includes as event: iii-iv acute gvhd (agvhd), relapse, death or chronic gvhd (cgvhd) requiring systemic treatment. in the last embt annual meeting a redefinition of this endpoint was proposed changing cgvhd event from those patients with cgvhd requiring systemic treatment (the original one) to those with just severe cgvhd (the redefined one). we retrospectively analysed patients consecutively transplanted ( - ) excluding non-malignant diseases, second allo-sct and those o years old age. we had generated two composite endpoints: in both iii-iv agvhd, relapse or death were considerated events but we defined grfs as the one with cgvhd event including those who required systemic treatment (as the original one) and in grfs just those with severe cgvhd (the ebmt redefined one). the median age was years ( - ) and % ( ) were males. other characteristics of patients are resumed in table . with a median follow up for patients alive of months , the median estimated survival in months and the % at + year and + years was: months, % and % overall survival (os); months, % and % event free survival (efs); months, % and % grfs ; months, % and % grfs . ( %) and ( %) hadn't any event in grfs and in grfs , respectively. in grfs , event's incidence was: ( %) for iii-iv agvhd, ( %) for cgvhd, ( %) for relapse and ( %) for death; in grfs was ( %), ( %), ( %) and ( %), respectively. considering those patients with cgvhd as event in grfs , of them hadn't the event as cgvhd at the same time in grfs (since they had cgvhd requiring systemic treatment but not severe cgvhd). for these patients, the alternative event in grfs was: without any event, relapsed and died. in the multivariate, the factors associated with better outcomes were: in grfs early ebmt stage (p o . with early as reference; intermediate p factor with more impact in both, but it is interesting to point it out that haploidentical donor had an advantage in grfs . these results are being validated in a large series and the definitive results will be available at the moment of the meeting congress. [p ] disclosure of conflict of interest: none. steroid refractory acute graft-versus-host disease (gvhd) remains a major complication of allogeneic hematopoietic stem cell transplantation (allo-hsct). affected patients have a very poor prognosis. gvhd has been associated with transplant-associated thrombotic microangiopathy (ta-tam). endothelial damage mediated by radiation, viral reactivation, drug exposure or alloreactivity results in exposure of subendothelial collagen, activation of coagulation and small vessel occlusion to a degree that results in organ failure. complement is thought to be a major mediator of endothelial damage. although a consensus exists about the exceedingly high morbidity and mortality of ta-tam and diagnostic criteria have been converging to a consensus, no biomarkers to diagnose tam and predict outcome have been established. we hypothesize that a ta-tma, related to dysregulation of the alternative complement pathway correlates with organ damage. a retrospective analysis of consecutive patients with hematological malignancies receiving an allo-hsct at the university hospital basel in the period from to was performed. data on the occurrence, risk factors and outcome of patients with ta-tma and the correlation with acute gvhd was collected. available biopsies of organs suspected to be affected by tam and/or gvhd will be performed. routine bone marrow biopsies for histological, immunohistochemical signs of ta-tam and complement activation will be analyzed. serum samples will be used to characterize markers of complement activation using plasma levels of c b- and c b- deposition in tissues biopsies. patients (aml n = ; all n = ; mds/mpn n = ; lymphoid neoplasm n = ; plasma cell disorder n = ; bone marrow failure n = ) underwent myeloablative (n = ) and non-myeloablative (n = ) allo-hsct at a median age of years (range: - years). forty-eight ( . %) patients matched the established diagnostic criteria for tam (increased ldh, platelet count o g/l or o % of normal baseline, schistocytes per high power field, creatinine increase). the median time to onset of tam was days post-transplant (range: - days). subjects with ta-tam had significantly higher -year nonrelapse mortality compared to those without ( . % vs . %, p o . ). grades - agvhd and cytomegalovirus viremia were independent risk factors for ta-tam, and serum ldh level u/l as well as arterial hypertension were early signs of ta-tma occurrence. patients with clinically relevant agvhd (⩾ grade ) had more ta-tam than patients without agvhd ( % vs %; po . ). tam correlated with agvhd severity; the higher the agvhd grade, the more the patients who suffered from tam. allo-hsct recipients with grades - agvhd or cytomegalovirus viremia should be closely monitored for the presence of ta-tma. at the meeting first results of histological, immunohistochemical and complement activation analyses will be presented. disclosure of conflict of interest: none. hemorrhagic cystitis (hc) after stem cell transplantation (sct) can cause significant morbidity and prolonged hospitalization. early bleeding occurs almost exclusively when using cyclophosphamide (cy) ( - % of cases), while late onset hc are classically attributed to bkv infection, and occurs up to % of patients (pts) receiving myeloablative haplo-sct who had positive bk viruria ( , ). we retrospectively studied hc cases among pts submitted to haplo-hsct in our department. thirty-eight pts receiving an haplo-sct with post-transplant cy (pt-cy) were included (table ) . prophylaxis for cy included hyperhydratation ( l/m of . % saline) and mesna administration ( mg for each mg of cy/daily divided into three doses). hematuria was graded as follows: grade i, microscopic; grade ii, macroscopic; grade iii, with clots; and grade iv, leading to urinary retention or requiring surgical intervention ( ). pts with hc and clots were treated with continuous bladder irrigation. twenty-three pts ( . %) developed hc at a median of . days post-sct (range: - ). clinical severity was grade i in cases ( . %), grade ii in cases ( . %), grade iii in cases ( . %) and grade iv in cases ( . %). at the onset of hc diagnosis, bk viruria was investigated in / pts. five pts ( . %) had bkv negative (bkv − ) hc and pts ( . %) bkv positive (bkv+) hc. bkv-hc occurred after a median of days (range: - ) while bkv+ hc after . days (range: - ), respectively (p = . ). among bkv+ pts, received iv cidofovir mg/kg once a week for weeks and then once every weeks. median number of administrations was (range: - ). oral probenecid was given at the dose of g h before and g and h after cidofovir administration. two pts obtained a complete response (cr) after and days, respectively, one patient reached a partial response after days and one pt failed to obtain a response. no pts developed renal toxicity during treatment. one pt received ganciclovir for concurrent cmv viremia and bkv+ hc resolved in days. three patients did not receive any treatment for mild or asymptomatic cystitis. all of them achieved remission after a median of days from the onset (range: - .) among bkv-hc, pts obtained spontaneous resolution after a median of days (range: - ), while two pts died early after sct. finally, among pts for whom bk viruria was not available, a remission was reached in of them after a median of . days (range: - ), while pts died early after sct. in our cohort of pts, hc occurrence was of . % and bkv was responsible for the . % of cases. contrary to its high incidence, hc showed a relative benign course, with an overall remission rate of . %, regardless of treatment. finally, we found a trend for a longer interval between sct and hc onset in pts with bkv+ hc, as compared to cy-related hc (p = . ). sos is a rare and serious complication of hematopoietic stem cell transplantation (hsct). it is diagnosed using the modified baltimore criteria of hyperbilirubinemia, weight gain or ascites % over baseline, hepatomegaly or right upper quadrant pain of liver origin. only defibrotide has been approved for the treatment of veno-oclusive disease. hdmp has been described as effective sos therapy in a few case series ( , ). we describe our experience of treating adult sos using hdmp. objective is to retrospectively analyze the treatment efficacy and overall survival of patients diagnosed with sos after hsct and treated with hdmp. we used vilnius university hospital data base to identify patients diagnosed with sos under baltimore criteria and treated with hdmp over - period. patient demographics, transplant and clinical data, response, survival (kaplan-meier survival analysis) and hdmp infusion related complications were analyzed. we identified patients ( males) of whom had had allogeneic hsct ( reduced intensity conditioning) and one had received a double autologous hsct. sos was diagnosed on the median day + (+ to + days). the median bilirubin value was . μmol/l ( . - μmol/l). all patients had liver enlargement of median mm ( - mm) on ultrasound. two patients had normal bilirubin values but displayed the remaining signs and symptoms of sos at diagnosis. patients received intravenous methylprednisolone mg/m every h for days. none received defibrotide. seven ( %) patients responded on median day + (+ to + days) after the start of hdmp. four responded by decrease in serum bilirubin by % and resolution of symptoms without the need of further treatment. the remaining three responders received maintenance treatment after one course of hdmp with reduced doses of methylprednisolone until resolution of symptoms. four patients failed to respond and died of multiorgan failure on median day + (+ to + ). the median observation time was months ( - months). the median overall survival for the sos group was months (range: - ) and it was months among the responders. no adverse reactions related to hdmp infusion were observed. hdmp therapy in adult sos results in clinically relevant response rate. further prospective trials are required to assess hdmp efficacy in comparison to defibrotide or as add on therapy. prevalence of hypertension (ht) in general pediatric population is~ %, while in children treated with hematopoietic stem cell transplantation (hsct) it is up to %. we assessed factors contributing to the development of ht in children treated with hsct and usefulness of ambulatory blood pressure monitoring (abpm) in this population of patients. the study included children ( boys, girls; mean age . years) treated with hsct for neoplasms (n = ; %) or non-neoplastic disorders (n = ; %). control group included children ( boys, girls; mean age years). abpm measurements (spacelab device) were performed before hsct and after a mean of months after hsct (in of the children). blood samples were collected from children treated with hsct and all controls. total rna extraction was performed and microarray analysis was conducted using genechip human gene . st arrays (affymetrix). in patients after hsct no antihypertensive treatment was used. mean systolic blood pressures (sbp) before and after hsct did not differ significantly from the control group. mean diastolic blood pressures (dbp) before and after hsct were . ± . mm hg and . ± . mm hg, respectively, and mean dbp percentiles were . ± . and . ± . , respectively; the differences between the study group and the control group were significantly higher before hsct. mean -hour arterial pressure (map) percentiles were . ± . and . ± . , respectively; the differences between the study group and the control group were significantly higher before hsct. before hsct and after the procedure, the european society of hypertension criteria for high normal blood pressure (bp) and ht were fulfilled in %/ % patients and %/ % patients, respectively. nocturnal bp decrease o % was found in %/ % patients and % nocturnal bp decrease in %/ % patients, respectively. in the control group o % nocturnal bp decrease was found in % of children and % nocturnal bp decrease in % of children. when the groups of patients before and after hsct were compared, highly significant differences were found in gene expression levels for mthfr ( in children referred for hsct a trend towards higher bp values was seen. in children assessed months after hsct more abnormalities in nocturnal bp measurements were seen, which may be a predictor of ht. in children treated with hsct significant differences in the expression of ht-related genes were found. abpm was useful in bp monitoring in children treated with hsct. hypothyroidism may complicate of allogeneic hematopoietic stem cell transplantation (allo-hsct); risk factors are analysed. we studied patients with aml who underwent an allo-hsct between and with different conditioning regimens (myeloablative, reduced-intensity, chemotherapybased, total body irradiation-based). thyroid stimulating hormone (tsh) and free thyroxin levels (ft ) were available in patients before and after allo-hsct. median age at transplantation (n = ) was years (iqr - ), ( . %) were female and overall mortality was . % (n = ) ( table ) ursodeoxycholic acid (udca) has been shown to have a protective effect in the liver complications after allogeneic stem cell transplantation (allo-sct), but it also has other immunomodulatory effects; it has been described also a potential benefice as graft-versus-host disease (gvhd) protection. we retrospectively analysed patients consecutively transplanted between - excluding second allo-sct and those o years old. we analysed the differences between those with and without prophylactic udca using spps v . results: the median age was years ( - ) and % ( ) were males. other patient characteristics are resumed in table objective of study was to evaluate the impact of disease status on the outcomes of allogeneic hematopoietic stem cell transplantation (allo-hsct) in the treatment of patients with refractory and relapsed acute lymphoblastic leukemia(all). patients with refractory and relapsed all, including cases in advanced stage (nonremission, nr) and cases in more than or equal to second complete remission(⩾ cr ), received allo-hsct after myeloablative conditioning regimen in our department. results: patients engrafted successfully. the transplantation-related mortality (trm) rate of nr and ⩾ cr was . % vs . % (p = . ). the incidence of agvhd was . % vs . % (p = . ), including . % vs . % (p = . ) with mild (grade i-ii) and . % vs . % (p = . ) with severe (grade iii-iv) agvhd. the incidence of cgvhd was similar also( . % vs . %, p = . ). with a median follow-up of ( . - . ) months, the cumulative relapse rate of nr and ⩾ cr was % vs . %(p = . ), respectively. the estimated year overall survival (os) and year leukemia-free survival (lfs) rate were . % vs . % (p = . ) and . % vs . % (p = . ), respectively. multivariate analysis results showed that cgvhd was independent favorable risk factor for os and lfs of r/r all. for relapsed patients, os was significantly better with first cr duration months and time to transplant ⩽ months. alio-hsct is an effective salvage treatment option for patients with refractory and relapsed all. our retrospective analysis showed that r/r all with different status prior transplant had similar outcome post transplantation. disclosure of conflict of interest: none. the deleterious effect of intensive care unit (icu) admission during hematopoietic stem cell transplantation (sct) on patient survival is well established. however, it is unknown whether admission into the icu during the chemotherapy for the underlying disease has any impact on survival after sct. we reviewed patients who had received a first sct between the years and in our institution, and we compared the overall survival (os), relapse incidence (ri) and non-relapse mortality (nrm) between patients who required icu admission during the chemotherapy prior to the sct (icu group) with matched patients ( : ) who did not need it (no-icu group). sixty-six patients were included, of them in the icu group and in the no-icu group. as shown in table , the main clinic-biologic variables and the sct procedure were comparable between the patient groups. the causes of icu admission for the icu group patients were: ( %) respiratory failure, ( %) septic shock, ( %) neurological disturbance, ( %) post-surgery and ( %) tumor lysis syndrome. seventeen patients ( %) needed mechanical ventilation. the median time between icu admission and the sct procedure was days (range: - ), and the median days of icu stay were . . with a median follow-up after sct of . years ( . - . ) for the icu group and . ( . - . ) for the no-icu group, the year os (ic %) probabilities were % ( - %) and % ( - %) in the icu and no-icu patients (p = . ), the -yr probabilities of relapse were % ( - %) and % ( - %)(p = . ) and the -yr probabilities of nrm were % ( - %) and % ( - %)(p = . ), respectively. there were no differences in either os, ri or nrm between icu and no-icu in the allogeneic or autologous subgroups considered separately. at the moment of the study, s ( %) of icu and ( %) of no-icu group had died. the causes of death in the icu group were: relapse in ( %), infection in ( %), gvhd in ( %) and gvhd plus infection in ( %). the causes of death in the no-icu group were: relapse in ( . %), infection in ( . %), gvhd in ( . %), gvhd plus infection in ( . %) and veno-occlusive disease and secondary malignancy, one each ( . %). in this series, admission to the icu before sct did not have an impact on outcomes after sct. these results suggest that these patients benefit from this treatment as much as the other patients, without expecting worse outcomes as a result of a previous icu admission. supported in part by grants rd / / (rticc, fejer), pi / , instituto carlos iii, and sgr (gre), spain. disclosure of conflict of interest: none. autologous stem cell transplant (asct) is a well established treatment for several haematologic and non haematologic malignancies, either as front-line or rescue therapy. however it is associated with toxicity and complications which might lead to treatment-related mortality (trm). although decrease in trm has been reported, data about the precise reduction and detailed analysis of causes of mortality throughout years are scanty. the aim of this study was to evaluate early trm and its causes in patients who underwent an asct in a single center along the last three decades. data of all consecutive adults ( years old) asct recipients were prospectively collected at a single center from december to august and then retrospectively analysed. trm was defined as mortality happened into the days post asct or during conditioning treatment due to any cause except relapse or progression of main diagnosis. demographic characteristics, diagnosis, conditioning regimen and cause of death were analysed. data were compared for two periods: from december to december and from january to august . a total of patients were included, median age was years ( - ) and . % were male. diagnoses were: lymphoma (n = ), multiple myeloma (mm) (n = ), acute myeloid leukaemia (aml) (n = ), amyloidosis (al) (n = ), acute lymphoblastic leukaemia (all) (n = ), solid tumours (including breast cancer and germ-cell tumours) (n = ), chronic myeloid leukemia (cml) (n = ), thrombotic thrombocytopenic purpura (ttp) (n = ) and autoimmune disease (n = ). the most frequent indication for asct was lymphoma ( . %) and mm ( . %). twenty patients died within d from asct (trm). demographic characteristics and causes of death for this patients are shown in table . the cumulative incidence of trm at day + was . % ( % ci . - . ). comparing both periods, trm cumulative incidence was . % ( % ci . - . ) in first period ( - ) vs . % ( % ci . - . ) in last period ( - ) po . . (figure ). according to main diagnosis trm cumulative incidence was higher in patients diagnosed with solid tumour . % ( % ci . - . ) and al . % ( % ci . - . ) followed by acute leukaemia (aml/all) . % ( % ci . - . ), mm . % ( % ci . - . ) and lymphoma . % ( % ci . - . ) p o . (figure ). sepsis ( %) was the main cause of death in both periods of time, and the only one cause of death in the last period. the second cause was sinusoidal obstruction syndrome (sos/vod) ( %), which only appeared in the first period. this study shows a low incidence of trm in asct recipients, with a significant decrease in the last period ( - ), despite the higher risk in some groups of patients such as those with amyloidosis and solid tumours. in our experience, infection is the main cause of early death in asct recipients and sos/vod has disappeared in last years as a cause of early transplant related mortality. disclosure of conflict of interest: none. incidence and risk factors for hepatic sinusoidal obstruction syndrome after allogeneic hematopoietic stem cell transplantation: a retrospective multicenter study of turkish hematology research and education group (threg) hepatic sinusoidal obstruction syndrome (hsos) is a potentially life-threatening complication of allogeneic hematopoietic stem cell transplantation (allo-hsct). the mean incidence of hsos was found to be . % ( - %) in the literature. we examined the incidence and risk factors for hsos after allo-sct. eight centers from turkey were enrolled in the study. we retrospectively evaluated the medical records of patients who were treated with allo-sct between january and december . a baltimore criterion was used for assessment of hsos. two hundred eighty three ( %) of patients who were treated with prophylaxis with defibrotide alone or one or more of the n-acetylcysteine, diuretics and heparin used defibrotide ( - mg/kg/day). the study included patients ( males/ females) with median age of ( - ) years. the demographic and clinical characteristics of patients were summarized in table . the incidence of hsos was . % ( ). prophylaxis for hsos was used in ( . %) of patients, who developed hsos. defibrotide as prophylaxis was received by of ( %) of patients. hsos developed in a median of ( - ) days after stem cell infusion. seventy-five ( . %) of patients who developed hsos had infection at the time of diagnosis. forty-five of them had ascites, had hepatomegaly and, had weight gain. seventy-two ( . %) of patients with hsos were treated with defibrotide after diagnosis. the median time of starting defibrotide in these patients was ( - ) days. thirty-six ( . %) of patients with hsos recovered completely and forty-seven ( . %) of them died as a result of multi organ failure. the incidence of hsosrelated mortality in allo-hsct cohort was found to be . %. in univariate analysis, statistically significant associations were not found between hsos incidence and age/sex of recipient, type of conditioning regimen, stem cell source and type of gvhd prophylaxis. on the other hand primary diagnosis of myelofibrosis, donor type, engraftment status and prophylaxis for hsos were significantly associated with hsos development. hsos prophylaxis was significantly decreased hsosassociated mortality (p = . ). hsos still remains a serious life-threatening complication of allo-sct. although the incidence is low, hsos is associated with increased -day nonrelapse mortality. hsos prophylaxis especially with defibrotide, seems to reduce hsos associated mortality in high risk patients. [p ] disclosure of conflict of interest: none. hemorrhagic cystitis (hc) is a serious complication occurring after allogenic hematopoietic stem cell transplantation (hsct) more frequent on haploidentical (haplo) hsct, with an incidence of - % associated mainly with the effect of cytotoxic agents such as cyclophosphamyde (cy). the conditioning regimen, bkpyv infection and graft versus host disease have an implication in the incidence. other authors related the reactivation of cmv and a previous transplantation as risk factors to hc development . with this study we aim to describe the hc incidence and risk factors in all haplo-hsct performed in the canary islands. we analyzed all consecutive haplo-hsct from family donors performed at our hospital between and . the conditioning regimen used for the transplant was the hopkins haplo protocol with high dose cy ( mg/kg on days and ) posttransplantation (ptcy). we used as hc prophylaxis intense hydratation on the cy administration day and the following h (using bladder wash only in patient with cardiac dysfunction) and perfused mesna at % of cy dose beginning min before the cy administration on pts and at % of the last dose at , and h on all pts. we used spss v. to determine the cumulative incidence (ci) of hc. we performed haplo-hsct, of which were males ( was transplanted times) and were women. the mean age was (range: - ). the pts presented the following diagnosis: aml ( ), all ( ), eh ( ), nhl ( ), am ( ). % of pts received the haplo-hsct in remission, % with refractory disease and % of pts did not receive previous treatment. pts developed hc ( . % ci at day + ) (figure a ) with a median time from haplo-hsct to onset of days (range: - ), ( %) was grade i, ( %) grade ii and ( %) grade iv. the grade i case did not received the mesna infusion like most of the other pts. no pts died due to hc and all cases resolved without sequelae. pts received cy pre-and post-transplant and only pts received ptcy. the ci at day + for the pts with ptcy was . % and for cy preand post-transplant . % (figure b) . we found no statistically significant difference on the ci of hc between these two groups. the development of hc was related to cy in patient, who suffered from this complication on the second and third haplo-hsct. for the rest of the pts (after day + ) the hc was related to bkpyv infection, as a consequence of the immunosuppression state of the patient, we also observed all these pts had positive serum viral load for cmv. the incidence of hc associated to post-hsct high cy dose in our series is % lower than other ones. most of them on grade or and without mortality associated. the risk of hc is high, particularly in the setting of highly pre-treated patients (especially those undergoing a nd transplant). the development of hc after day + is evidently associated to bkpyv as a contributing factor for continuous inflammation and cmv reactivation (as an immunosuppression marker). in our study, hc did not have an impact on mortality of high-risk patients after haplo-hsct. the hc remains frequent with a high morbidity in particular when it is severe, often causing prolonged hospitalization and resource use. we need further studies to recognize the at-risk population early. [p ] ta-tam is not a rare post-transplant complication and it is potentially fatal. in survivors, it was often associated with longterm morbidity and chronic organ damage, mostly to the kidney with poor renal prognosis. our retrospective study showed ta-tam associated risk factors included t reg haplo hsct as the incidence was highest in this group, tbi-based conditioning or tbi based conditioning plus cyclophosphamide. although acute gvhd and infection were associated with ta-tam in retrospective studies, no association emerged between acute gvhd or infection preceding diagnosis in our series of patients. in order to prevent ta-tam we need to understand its underlying biological mechanism so we are investigating its pathogenesis by means of cytokine assays, histology and murine models. disclosure of conflict of interest: none. mortality in children requiring invasive mechanical ventilation (imv) after allogeneic hematopoietic stem cell transplantation (hsct) is known to be high. little is known about the longterm outcome of those who survive imv. we therefore reviewed the medical records of children who survived s imv after they had received a hsct between and in the two pediatric hsct centers in the netherlands. retrospective multi-center cohort study in two university hospitals that perform all pediatric hscts in the netherlands. long-term survival of hsct recipients who had received imv was assessed. health status was reviewed more in detail for those who were still alive years after discharge from the pediatric intensive care unit (picu). in the absence of standardized use of quality of life questionnaires, health status was expressed as the number of affected domains (cardio-respiratory, motor and miscellaneous, regardless of the degree of dysfunction) and level of education. health status was categorized as follows: no health problems when all four domains were normal; mild health problem when there was an abnormal score in one of the four domains; moderate health problems when there was an abnormal score in two domains; severe health problem when there were abnormal scores in three or all four domains. between january and december , patients underwent a hsct in the two study centers together. a total of hsct recipients received imv within year after hsct ( % of all hsct recipients). median time between hsct and picu admission was days (iqr - days). the most common indication to start imv was respiratory failure ( %). median duration of imv was days (iqr - days). patients ( %) died during their picu admission. of the patients who were discharged alive from picu, patients were still alive years after picu discharge ( % of those who survived picu admission). health status of these long-term survivors was assessed in december by hospital database review, using the most recent hospital contact. follow-up time varied from to years (median . years) after picu discharge. two patients ( %) had no health impairment, eight patients ( %) had mild health problems, five patients ( %) had moderate health problems, and nine patients ( %) had severe health related problems. very little is known about long-term mortality and morbidity of hsct recipients who survived imv. survival of picu treatment in pediatric hsct recipients is limited. however, long-term outcome of those who survive picu treatment seems promising: a considerable proportion of them still is alive years later without obvious sequelae. this is the first study which assessed long-term outcome of imv after hsct. further studies in this population are urgently required to counsel parents and to optimize quality of life outcomes in these children. disclosure of conflict of interest: none. long-term surveillance data support lack of increase in mortality or cancer risk in brincidofovir clinical trial participants m morrison, k fitzgerald , t brundage, a harrison and wg nichols chimerix brincidofovir (bcv) is an orally bioavailable lipid conjugate of cidofovir (cdv), with broad-spectrum activity against doublestranded dna viruses, including cytomegalovirus (cmv), adenoviruses (adv), polyomaviruses (bk and jc viruses), and orthopoxviruses. bcv is being evaluated for prevention of cmv and other dna viruses in high-risk hematopoietic cell transplant (hct) recipients, and for the treatment of serious adv infection. bcv is also being developed for the treatment of smallpox under the us fda's animal efficacy rule. because bcv, cdv, and other marketed nucleoside analogs are reported to be carcinogenic in rodents, a registry was established to evaluate the long-term safety of bcv in subjects who have participated in bcv clinical studies. to date, the registry includes prior participants from study (a placebo (pbo)-controlled study of bcv for cmv prevention) and study (a single-arm study of bcv for adv treatment). subjects are encouraged to consent for long-term follow-up in the registry following participation in bcv clinical studies. registry participants are followed at -month intervals for a minimum of years from the time of completion of the parent study. development of malignancies (new or relapsed), lifethreatening or fatal adverse events (aes) assessed as potentially related to bcv, and subjects' vital status are collected. a total of subjects were enrolled in the parent studies ( bcv and pbo from study , bcv from study ). of these, are enrolled in the registry as of october ( bcv and pbo from study , bcv from study ). bcv recipients in the registry are % male, % white, with a median age of ( o - ) years, similar to the bcv recipients in the parent studies. the median duration of follow-up is ( - ) months, with % of subjects continuing in follow-up at the time of analysis. all-cause mortality from the time of first dose in the parent study through current registry follow-up is % for bcv vs % for pbo (p = . ) in study , and % for bcv in study . all-cause mortality in the registry since completion of the parent study is % bcv vs % pbo for subjects from study (p = . ) and % bcv for subjects from study (figure ) . the incidence of a new malignancy in registry subjects from study is % bcv vs % pbo (p = . ), and the incidence of relapsed primary malignancy is % bcv vs % pbo (p = . ). in registry subjects from study , % developed a new malignancy and % had a relapse of the primary malignancy. no bcv-related life-threatening or fatal aes have been reported to date in the registry. registry data collected to date support no increase in late mortality or increased risk for carcinogenicity in patients treated with bcv. long-term surveillance for cancer risk and other drivers of mortality is important for novel compounds undergoing development in hct and other immunocompromised patient populations, with high background risk for these outcomes. [p ] disclosure of conflict of interest: all authors of this abstract are employees and stockholders of chimerix. hematopoietic stem cell transplantation (hsct) is a medical procedure that allows the cure of many paediatric diseases. it has been described an increased risk of new malignancies in this population and it represents an important cause of late mortality. we analyzed the late evolution of patients submitted at pediatric age to hematopoietic transplantation (hsct) (allogeneic or autologous) in santa creu i sant pau hospital between and . a total of hsct was analyzed. it has been calculated the cumulative incidence of secondary malignancies at years of follow-up. it has been done univariate and multivariate analysis of risk factors through χ -test and binary logistic regression method (odds s ratio, or). it has been studied the relative risk (rr) for new malignancies through comparison of observed cases in our cohort with the expected cases in the general population. we observed cases of secondary malignancies with a cumulative incidence of % at years, % at years and % at years of follow-up. the risk was higher of expected in general population for each tumor type and in the different range of age, being the rr for malignancies in our cohort of . at years of follow-up. solid tumors were the most prevalent malignancies ( out of cases). the median time of latency from hsct to diagnosis of malignancy was years ( - years) . the thyroid tumors were the later ones and hematologic malignancies the earliest to be developed. chronic graft versus host disease was a statistically significant risk factor in univariate (or = ; p = . ) and multivariate analysis (or = . ; p = . ). total body irradiation of conditioning was a significant risk factor only in multivariate analysis (or = . ; p = . ). previous radiotherapy was a significant risk factor only in univariate analysis (or = . ; p = . ). mortality was % ( out of ) between patients with a new malignancy and it was the cause of death for all the cases. we observed an incidence of secondary malignancies after hsct of . % that is significantly higher compared to the expected in the general population (p = . ). the factors that have been related to an increased risk were chronic gvhd, tbi and previous radiotherapy. microalbuminuria defined, as urinary albumin: creatinine ratio (acr) of - mg albumin/g creatinine is a marker of endothelial dysfunction and inflammation. in general populations albuminuria predicts development of chronic kidney disease (ckd) and cardiovascular disease ( ). in the general population microalbuminuria is associated with the metabolic syndrome ( ) . in patients with hypertension, diabetes and the critically ill, it is a marker of adverse events and poor outcomes. following hematopoietic cell transplant (hct), microalbuminuria at day was associated with a four-fold increased risk of chronic kidney disease (ckd) at year in a single centre study; macroalbuminuria at day was associated with a . -fold increased risk of non-relapse mortality ( ) . international guidelines for adult and children survivors of hct recommend that proteinuria is assessed at least annually ( , ) . there is a paucity of data on the prevalence and implications of micro and macroalbuminuria in long-term survivors ( years) of adult hct, however. this was a single-centre retrospective study conducted in patients attending a dedicated clinic for long-term (minimum years) survivors of hct. we investigated prevalence of albuminuria and its association with renal disease, cardiac disease and the metabolic syndrome. of patients, were treated for acute leukemia, for aplastic anaemia and for cml. the median follow up time was years (range: - years) and the median age at follow up was years (range: - years). for / urinalysis was normal (group a) and ( %) had microalbuminuria (group b). none had macroalbuminuria. group b were significantly more likely to have ckd grade - (egfr o ) compared to those in group a (p = . ). group b patients were significantly more likely to have diabetes or impaired glucose tolerance / ( %) vs / ( %) in group a (p = . ). group b patients were also significantly more likely to have dyslipidaemia (p = . ) with / ( %) affected vs / ( %) in group a. cardiac disease and hypertension were more frequent in group b, / ( %) and / ( %), respectively vs group a / ( %) and / ( %) but these data were not statistically significant. the more features of the metabolic syndrome present, however, (elevated hba c, /glucose, dyslipidaemia, hypertension) the more likely a patient was to have microalbuminuria (p = . ). our data demonstrates that microalbuminuria is a frequent finding in long term survivors of hct. patients with microalbuminuria are more likely to have ckd grade or below. they are also more likely to have diabetes and dyslipidaemia. as this was a retrospective study we are not in a position to comment on whether microalbuminuria is predictive of the development of renal disease, metabolic syndrome or cardiovascular disease in this group of patients. this warrants further study as intervention, for example with ace inhibitors, may have the potential to reduce morbidity. the purpose of the study is the improvement of transplantation techniques and supportive care lead to an increasing number of long-term survivors after allogeneic hematopoietic stem cell transplantation (ahsct). recipients of ahsct have a higher prevalence of cardiovascular risk factors. ambulatory blood pressure measurement (abpm) is the 'gold standard' to diagnose arterial hypertension (ht). the prevalence and treatment control of ht by abpm is unknown in ahsct patients (pts). this prospective single center study at university hospital basel included all pts ⩾ year after ahsct in complete hematological remission during annual follow-up consultation. office blood pressure (obp) was measured on both arms after minutes rest. abpm by noninvasive continuous bp monitoring (pulse transit time method) was performed on the same day. ht was defined as obp ⩾ / mm hg, mean systolic bp ⩾ mm hg on abpm s (bp ) and/or current use of antihypertensive drugs. pts ( % female) were included with median age of years (range: - ) and years (range: - ) after transplantation. ( %) pts received total body irradiation-based conditioning, ( %) pts had chronic graft-versus-host disease, and ( %) required immunosuppression. mean bmi (kg/m ) (± sd) was ± , with ( %) pts . twenty-seven ( . %) pts were current smokers. fourty-three ( %) pts had chronic kidney disease (egfr o ml/min/ . m ) and ( %) diabetes. ( %) pts were on antihypertensive drugs consisting of ace/at-ii-inhibitors in ( %), calciumchannel blockers in ( %), beta-blockers in ( %) and diuretics in ( %) pts. thirty-nine ( %) pts were on ⩾ drugs. among our cohort ( %) pts were normotensive without antihypertensive treatment (mean age ± years, % female and mean bp (systolic/ diastolic bp) ± / ± mm hg). ( %) pts were hypertensive and/or on antihypertensive treatment. untreated ht was diagnosed in ( %) pts (mean age ± years, % female and mean bp of ± / ± mm hg), including ( %) with white-coat hypertension and ( %) masked hypertension (normal obp, high abpm). in the group of pts with current antihypertensive medication / ( %) were controlled (mean age ± years, % female, and mean bp ± / ± mm hg) whereas / ( %) were hypertensive on abpm (mean age ± years, % female, mean bp ± / ± mm hg). thirty-four ( %) pts with uncontrolled ht were already hypertensive at obp. although long-term survivors after ahsct are known to be at elevated cardiovascular risk, diagnosis of arterial hypertension was missed in every fifth patient. the proportion of controlled hypertension is poor with only %. disclosure of conflict of interest: none. myasthenia gravis (mg) is a rare complication of allogeneic stem cell transplantation (sct) and is often associated with graft-versus-host disease (gvhd). we report a -year-old man who presented with oculobulbar and neck weakness months after an unrelated donor, allogeneic sct for chronic myeloid leukaemia (cml). he was diagnosed in with chronic phase cml. this responded poorly to tyrosine kinase inhibitors (tkis) and he was found to carry the t i mutation with additional monosomy . he underwent a fully hla matched unrelated donor sct with y -anti cd targeted radiotherapy, fludarabine, melphalan and alemtuzumab conditioning. he had grade cutaneous gvhd on ciclosporin withdrawal but no other significant gvhd. he has an immune mediated neutropenia since months post sct and has reduced immune reconstitution as demonstrated by a sub-normal absolute cd level. he remains on pneumocystis prophylaxis and has not experienced increased infection. chemotherapeutic agents have a cytotoxic effect on the oral mucosa and is a major problem following cancer treatment. cooling the oral mucosa in conjunction with chemotherapy infusion, using ice chips, is known to reduce the severity of oral mucositis ( , ) . although effective, ice chips are perceived as uncomfortable. the aim of the present study was to determine the optimal cooling temperature to prevent adverse effect of chemotherapeutic agents using tissue engineered oral mucosal models (teom). teom were incubated at °c, °c, °c or °c for min followed by exposure to μg/ml of -fu for h (control models were incubated at °c). teom were then washed and further incubated for h at °c co . cell viability and inflammatory cytokine production (il- and tnf-α) were measured using (prestoblue) and (elisa), respectively this study demonstrates an increased capacity to restore cell viability with decreasing temperature (figure a ). teom treated with -fu further showed an increased secretion of the pro-inflammatory cytokines tnf-α and il- at all temperatures compared to un-treated controls. for il- , secretion increased markedly when cells were incubated with μg/ml -fu at °c and °c compared to cells incubated with medium alone at °c (figure b) . for tnf-α, secretion was significantly higher (p o . ) in cells treated with μg/ml fu at °c compared to untreated mucosal models and mucosal models treated with μg/ml fu but incubated at °c (figure c ). teom models incubated at °c has an increased capacity to restore cell viability following exposure to -fu. incubation at °c further reduces the release of pro-inflammatory cytokine compared to those incubated at °c. ( ) and one received fludarabine and cyclophosphamide. all patients received campath- h as part of the conditioning regimen. stem cell source: peripheral blood stem cells patients and bm. comorbidity was assessed using the haematopoietic cell transplantation co-morbidity index (hct-ci), with patients ( %) having no co-morbidities, ( %) a co-morbidity index of - and ( %) had a score ⩾ . follow up of survivors ranged from to months (median: months). at the specified end point patients had relapsed ( %) with an actuarial -year relapse rate of %. there were deaths ( %). relapse ( ) was the main cause of death with transplant related mortality of % ( ) at day , % ( ) at months and % ( ) at year. the actuarial os at years was %, with a -year dfs of %. of the surviving relapsed patients all received chemotherapy and donor lymphocyte infusions resulting in effective recovery of remission, showing the utility of this approach. in terms of co-morbidity, actuarial survival rates were % in those with an hct-ci index of , % with an index of - and % with an index ⩾ . the results of this retrospective study indicate that allosct using reduced intensity conditioning regimens can be an effective treatment strategy for older patients with high risk myeloid malignancies including those with significant co-morbidities. relapse remains the main cause of treatment failure and strategies to reduce relapse risk are required. patients that relapse post allosct may respond to further treatment such as azacytidine or intensive chemotherapy and donor lymphocyte infusions. ( ) whether patient-related variables were associated with disagreement. this is a secondary analysis of a cross-sectional multicenter study where patients and clinicians completed an identical qol questionnaire (fact-bmt) at day . clinical and demographic variables as well as anxiety and depression (hads) were collected. agreement was analyzed with the intraclass coefficient correlation (icc). rates of under-and over-estimation were calculated. logistic regression models identified predictors of disagreement. we analyzed pairs of questionnaires, filled in by patients and clinicians. patients' median age was years, ( %) were men, and ( %) received an allogeneic hct. clinicians' median age was years, were men and had worked on the transplant field for a median of years (range: - ). agreement on qol was moderate (icc = ). exploratory analyses revealed that agreement for emotional (icc = ) and social (icc = ) wellbeing was poor, whereas it was moderate for physical (icc = ), functional (icc = ) and bmt concerns (icc = ). patients' wellbeing was overestimated in - % of the categories of wellbeing parameters, and underestimated in - %. patient-related variables explained - % of the variance on disagreement across scales. specifically, anxiety contributed to disagreement in all subscales, except in social wellbeing, where non-significant univariate associations were observed (p . ). type of transplant (allogeneic vs autologous), performance status, and graft-versus-host disease were not associated with disagreement (p . ). patients and clinicians agreement on qol is suboptimal, particularly on emotional and social wellbeing. patients' wellbeing cannot be estimated from other sources than themselves. these results highlight the unmet needs of hct recipients with respect to qol-related issues; an outcome that must be addressed by hct programs since their wellbeing is as important as survival endpoints. disclosure of conflict of interest: none. . we wanted to test the function and the safety of picc device as alternative to standard cvc in patients submitted to autologous stem cell transplantation (abmt). the primary end point of the study was to individuate the cause leading to the failure of picc (its removal or the needing of another cvc during the abmt procedure). secondary end points were the correct function of the device and its praticity. twenty patients submitted to abmt for multiple myeloma ( ) or lymphoma ( ) experienced a double lumen picc device ( ) or a single lumen ( ) if the patient already carried a permanent single lumen cvc such as hickman or port-a-cath. we excluded from this experience patients with high risk of life-threatening situation or high risk of intensive care already before abmt. picc devices were placed from a specialistic nurse team by ultrasound identification of a deep venous vessel in upper arms. melphalan or ceam were the standard conditioning regimens employed in myeloma and lymphoma abmt respectively. we considered a failure all the causes leading to the removal of picc or requiring another cvc before the end of the transplant procedure. at last we collected nurses and clinicians opinions about the picc functionality. no complication has been recorded in positioning phase. / patients maintained the picc device for all the time of transplant procedure. only one patient needed to remove the device for infection. the opinion of nurses and clinicians about the picc device was a significatively slower speed of infusion and resistance to the flow; in fact, / patients needed an infusional pump. the idraulic resistance of the catheter was particular evident against cellular fluids (stem cells suspension, transfusions of blood and platelets). for this reason picc seems to be less indicated in patients requiring many endovenous infusions (nurses' opinion). the rate of infection of picc devices seems to be lower compared to cvc, but the number of cases tested in this experience is too limited for definitive conclusions about it. for other aspects picc is similar to other cvcs. picc seems to be a valid alternative to standard cvc in patients who do not require intensive care, and in particular in patients with low intensity abmt who do not present a high number of endovenous infusions. maybe picc is less burdened of infections respect to normal cvc. this fact, summed to the lower risk during the positioning of the device, leads to consider the use of this device in abmt setting for standard risk patients. disclosure of conflict of interest: none. there are only few algorithms for the selection of hlamismatched unrelated donors, when no fully matched donor is available. indirect recognition of hla-mismatches can be predicted using the model of 'predicted indirectly recognizable hla epitopes' (pirche). the pirche model is a recently developed computer-based strategy, which classifies hladerived epitopes that are potentially presented by patientdonor shared hla-molecules. we performed a multicenter retrospective study evaluating the impact of pirche on outcome after allogeneic stem cell transplantation from hla / matched unrelated donors. the study cohort included adult patients who had undergone allogeneic stem cell transplantation for aml or mds. pirche scores were computed for recipients of hla / matched unrelated donor transplants ( / mud) using a web-based tool. primary endpoint was overall survival at years. patients with a / mud were divided into groups according to the sum of pirche i+ii values (pirche score). eighty-five ( ) patients had a pirche score of (no pirche detected), a pirche score . km estimate of year os was higher for / mud with pirche score = compared to pirche score : % ( % ci: - %) vs % ( % ci - %), p = . . os was similar for / mud with pirche score = and / mud ( % vs %). cox regression analysis revealed poorer os for pirche scores (rr . , % ci: . - . , p = . ). cumulative incidence of nrm at years was lower for / mud with pirche score = compared to pirche score ( % vs %, p = . ). multivariate cox regression analysis revealed poorer nrm for pirche score (rr . , % ci: . - . , p = . ). cumulative incidence of agvhd grade - at months was not significantly different for / mud with pirche score compared to pirche score ( % vs %, p = . ). cumulative incidence of cgvhd at years was lower for / mud with pirche score compared to pirche score ( % vs %, p = . . our findings require confirmation, ideally in a large prospective cohort study. if validated, the pirche model would allow selection of permissible hla-mismatches that may be associated with an improved transplant outcome in terms of reduced nrm and better os. [p ] disclosure of conflict of interest: none. this study was supported by a research grant from pirche-ag to the university medical center, hamburg-eppendorf. pretransplant liver dysfunction has been recognized as a risk factor for complications and mortality after allogeneic hematopoietic cell transplantation (allo-hct). however, there is no consensus on the optimal way to evaluate liver function in hct candidates. transient elastography (te) is a noninvasive method for diagnosing liver damage and cirrhosis. while elastography is widely used in the setting of viral hepatitis, its possible role in allo-hct recipients has not been deeply evaluated. patients receiving allo-hct in our center from may are scheduled to receive pretransplant evaluation by a hepatologist under a prospective protocol. the evaluation includes a hepatologist consultation, liver function and infectious serology tests and te. all patients receive ursodiol from hct admission to day + . this study constitutes the first evaluation of the ongoing protocol for patients receiving their first allo-hct from may to august . sixty patients received a first allo-hct during the study period. sixteen patients did not undergo hepatologist evaluation due to timing issues (n = ), unstable medical condition (n = ) or other reasons (n = ). finally, patients received pretransplant evaluation by a hepatologist under the current protocol and constitute the study population. median age at transplantation was years (range: - ). most patients received a transplant for acute leukemia (n = , %) or non-hodgkin's lymphoma (n = , %) mainly from hla matched unrelated donors (n = , %). thirty-two patients received reduced-toxicity regimens ( %). graft-versus-host disease (gvhd) prophylaxis consisted of tacrolimus in combination with another agent. median follow-up for survivors of months (range: - ). median elastography was . kpa (range: . - . ). considering the hct-ci categories on hepatic dysfunction, , and patients scored , and points, respectively. there were two cases of veno-oclusive disease (vod). overall survival and non-relapse mortality of all patients at median follow-up were % ( % ci - ) and % ( % ci - ), respectively. in the univariate analysis, median elastography was not associated with a higher risk of nrm (p-value = . ), os (p-value = . ) or hepatic chronic gvhd (p-value = . ). the two patients with vod had normal pre-hct transaminase levels and te. this first analysis of an ongoing protocol with universal pre-hct evaluation of hepatic function indicates that increased values of transient elastography are not associated with higher nrm or lower os after the procedure. further studies including a larger number of patients are needed in order to clarify the possible role of elastography in the hct setting. disclosure of conflict of interest: none. allogeneic hematopoetic stem cell transplantation (hsct) remains associated with a high morbidity and mortality in spite of advances in hsct management. specifically, pulmonary complications account for a substantial proportion of deaths within the first days after hsct. therefore, identification of lung dysfunction and additional comorbidities are crucial for preventive strategies in hsct. given the inconsistent association of pretransplant lung function s parameters on mortality after hsct and the significant changes in hsct care over the last decades, the aim of our study was to assess the effect of pulmonary function and comorbid conditions on mortality in patients undergoing hsct for hematological disorders. we retrieved relevant clinical data of all consecutive patients at the hematology division of the basel university hospital with a transplant for hematological disorders between and . we examined the lung function at baseline and , and months after hsct-including the s forced expiratory volume (fev % of predicted), fev /vcmax and diffusing capacity for carbon monoxide (dlco, adjusted for hemoglobin concentration). in addition, we assessed pretransplant conditions such as age, sex, karnofsky performance status (kps), donor type and various risk scores in hsct (hematopoietic cell transplantation comorbidity index (hct-ci), european society for blood and marrow transplantation (ebmt), revised pretransplant assessment of mortality score (pam)). using uni-and multivariate cox proportional-hazards regression analysis, we evaluated patient-and transplant-related risk factors for all-cause mortality by including the following categorical candidate variables: fev (⩾ % vs - % vs o % of predicted), kps ( o % vs ⩾ %), age ( o vs ⩾ years), conditioning intensity and donor type (matched-related vs mismatchedrelated vs matched-unrelated vs mismatched-unrelated). within the study period, patients with predominantly acute leukemia ( %) or lymphoproliferative disorders ( %) underwent myeloablative (n = ) and non-myeloablative hepatic veno-occlusive disease/sinusoidal obstruction syndrome (vod/sos) is a potentially life-threatening complication of conditioning for hematopoietic stem cell transplantations (hsct). recombinant thrombomodulin (rtm) is a new drug for treating disseminated intravascular coagulation (dic) and is an endothelial anticoagulant cofactor that promotes the thrombin-mediated formation of activated protein c (apc). rtm has been used to treat vod/sos, but its ability to prevent vod/sos has not been established. we evaluated the cases of pediatric hematology and oncology patients ( ( %) acute myeloid leukemia, ( %) acute lymphoblastic leukemia, and ( %) neuroblastoma patients, and ( %) patient each with myelodysplastic syndrome, rhabdomyosarcoma, hemophagocytic syndrome (hlh), and wiskott-aldrich syndrome) who underwent hsct at our institution between and and had ≧ risk factors for vod/sos. these risk factors included previous treatment with gemtuzumab ozogamicin (go), receiving hsct, undergoing conditioning with busulfan (bu), and being diagnosed with hlh. the patients who received hsct after (n = ; rtm group) were treated with rtm as a prophylaxis against vod/sos ( u/kg per day for days; from days to ) together with ursodeoxycholic acid (urso) and low-molecular-weight heparin (lmwh), and the others (n = ; control group) were only treated with urso and lmwh. the incidence of vod/sos was evaluated, and various coagulation parameters and markers of endothelial injury (plasminogen activator inhibitor type (pai- ) and apc) were measured in both groups. the patients' median age was (range: - ) years, and ( %) were male. clinical characteristics, including vod/sos risk factors, were wellmatched in both groups. the risk factors possessed by the patients included receiving hsct ( / , %), previous go treatment ( / , %), conditioning with bu ( / , %), and a diagnosis of hlh ( / , %). although vod/sos occurred by post-hsct day + in ( %) patients in the control group, vod/sos was not seen in the rtm group. two of the former patients ( : previous treatment with go, : a diagnosis of hlh) suffered severe vod/sos, and (a diagnosis of hlh) died of the condition. no grade / adverse events involving bleeding or severe organ damage were reported in the rtm group. interestingly, the mean peak value of pai- and apc (markers of endothelial injury) were significantly lower in the rtm group (table ) . [p ] disclosure of conflict of interest: none. protective effect of early human cytomegalovirus reactivation on relapse of myeloproliferative disorders after allogeneic hematopoietic stem cell transplantation z peric , , j wilson , n durakovic , , l desnica , a ostojic , vv rezo , v marekovic , , r serventi-seiwerth and r vrhovac , school of medicine, university of zagreb, zagreb, croatia and university hospital centre, zagreb, zagreb, croatia there have been conflicting results regarding the association between early cytomegalovirus (cmv) reactivation and decreased incidence of relapse after allogeneic hematopoietic stem cell transplantation (allo-hsct). this prompted us to retrospectively evaluate the potential impact of cmv reactivation on transplantation outcomes in a study population of consecutive adult patients who underwent allo-hsct in our institution and were treated and followed in a homogenous manner. patients were monitored for cmv reactivation once weekly for the first days after allo-hsct. monitoring was done with a real time qpcr with lower limit of detection of genome copies per ml of blood. when cmv viremia was detected, all patients were treated with intravenous ganciclovir or oral valganciclovir untill two consecutive negative qpcr assays. univariate and multivariable proportional hazards models using the fine and gray approach were considered to evaluate the variables for relapse, treating death as competing event. between and , male and female patients underwent allo-hsct at a median age of years (range: - ). among them, most patients were treated for myeloid malignancies ( aml, mds and mpn with cml, mf and cmml), while the rest had lymphoproliferative disorders ( all, nhl, mm, mh and cll) and one patient had aplastic anemia. the donors were unrelated in cases, related in patients and haploidentical in patients. most of the patients ( %) received peripheral blood stem cells after a reduced-intensity conditioning regimen ( %). with a median follow-up of months, early cmv reactivation occured in % patients at a median of days after transplantation and did not affect relapse incidence in patients with lymphoproliferative disorders. on the contrary, the cumulative incidence (ci) of hematologic relapse in patients with myeloproliferative disorders (aml and mpn) at months after allo-hsct was % ( % ci, - %) in patients without, opposed to % ( % ci, - %) in patients with cmv reactivation (p = . ). however, cmv reactivation did not significantly affect (p = . ) overall survival between patients with ( %; % ci - %) and without cmv reactivation ( %, % ci - %). a striking and previously unreported correlation between cmv reactivation and relapse was found in patients with mpn; the ci of relapse was % ( % ci, - %) in patients without, opposed to only % ( % ci, - %) in patients with cmv reactivation (p = . ). a substantial and independent reduction of the relapse risk in myeloproliferative disorders (aml+mpn) associated with early cmv reactivation was confirmed by multivariate analysis using time-dependent covariate functions for high-risk disease, use of atg, chronic graft-versus-host disease (hazard ratio . ; % ci, . - . , p = . ), and cmv reactivation (hazard ratio . ; % ci, . - . , p = . ). in summary, this report supports an independent role of cmv reactivation on relapse in patients with myeloproliferative disorders. to our knowledge, we are the first to show a significant reduction of relapse incidence in patients with mpn, even though our findings are based on a relatively small number of patients. however, this putative virus-versus-myeloproliferation effect definitely warrants further research. [p ] disclosure of conflict of interest: none. final result of fact-bmt is score ranged - point (the higher the score, the better qol). for qualitative assessment of donor-recipient relationship, the adult sibling relationship questionnaire (asqr) in polish version was used. the asrq-s consists of items which are spread over eight scales designed to investigate three factors: warmth, conflict and rivalry. the questionnaires were given to both subgroups, donors and recipients of msd-hsct and the results were compared to each other. the overall result of the fact-bmt questionnaire was . ± . points, which means that the examined group generally described their qol as 'quite good'. the best results were found in functional well-being ( . ± . ), while the worst in emotional well-being ( . ± . ) dimension. statistically, the qol score was not influenced by age at hsct (p = . ), current age (p = . ) or gender (p = . ) of the respondents. the recipients scored highest on warm factor ( . ± . ), while donor respondents scored slightly higher rivarly ( . ± . ) than warm ( . ± . ). the second dimension scored by recipients was rivarly ( . ± . ). conflict scores were lowest, although donor respondents scored higher on these than recipient respondents ( . ± . in donors vs . ± . in recipients). statistical analysis revealed that the being a donor or recipient of msd-hsct determines the level of rivarly in the sibling relationship (p = . ) with no impact on warm and conflict dimension. health-related qol in transplanted patients is quite good. sibling donor-recipient relationship is unbalanced with recipient respondents being more likely to assess a warm relationship, while rivalry was more likely to be present among donor. further multicenter studies based on larger cohort of patients are necessary to assess sibling relationship after transplantation life experience. disclosure of conflict of interest: the authors have nothing to disclose. this work was supported by grant from poznan university of medical sciences ( - - - ). rate of re-admission in patients undergoing allogeneic transplants from identical siblings, unrelated donors or haploidentical donors f sora , s sica, l laurenti, p chiusolo, s giammarco, i innocenti, e metafuni, a corbingi and a bacigalupo department of hematology, catholic university of rome hla identical siblings (sib), unrelated (ud) and family hla haploidentical donors (haplo) are currently being used for patients undergoing an allogeneic transplant (hsct) for hematologic disorders. gvhd prophylaxis is usually different, and is commonly based on a calcineurin inhibitor (cni) and methotrexate (mtx) with or without atg for sibs and uds, wheres post-transplant cyclophosphamide (pt-cy)+a cni and mycophenolate (mmf) is used for haplos. we will refer as sib, ud, haplo platform, the combination of a given donor and a given gvhd prophylaxis. the outcome of these three different platforms is usually measured in terms of gvhd, non relapse mortality (nrm) and survival. days of admission and readmissions are important in terms of morbidity, but also of costs, and are usually not reported. aim of the study: assess the duration of the first admission and the incidence of a new re-admissions, in the first days after the transplant. we retrospectively analyzed patients from to . sixtyone received peripheral blood stem cell graft from an ud, and gvhd prophylaxis with cya+mtx+atg; received a peripheral stem cell graft from a sib and gvhd prophylaxis with cya +mtx; patients received bone marrow hsct from haplorelated donor and pt-cy+cya+mtmf. patients characteristics are shown in table . relapses were excluded from the readmission analysis. the median time from the transplantation to discharge was days for ud, for haplo and days for sib: there was no significant difference between haplo vs ud (p = . ), whereas the admission of both haplo and ud was longer than sibs (po . ). first readmission. fiftyone patient out of required of a new admission for complications after tranplant ( out of after mud ( %), out of ( %) using a sibling donor and out of using an haploidentical donor ( %)). there were significantly more re-admissions in the ud vs sib group ( . ) and a trend for more ud readmissions vs haplo (p = . ); siblings had the lowest number of readmissions. time to neutrophil engraftment was comparable in haplo vs ud patients (p = . ) and in sib vs ud (p = . ); the time was longer in haplo vs sibs (p o . ). the reason to re-admitted the patients in the hospital after tranplantation was fever in out of ( %) new admissions in ud setting, out of ( %) in sib and out of ( %) in haplo; acute gvhd was the cause for re-admission in out of ( %) ud, out of ( %) sib and none in haplo. the other causes for re-admission in the hospital were hemorragic cistitis, thoracic or abdominal pain. second re-admission. of hospitalization is registered in out of patients in ud ( for aghvd and fever), out of ( %) in sib ( episodes of fever) and out of ( %) patients in haplo ( for fever and progressive disease). also for second episodes, ud grafts had significantly more admissions compared to haplo and sibs. third re-admission was recorded only in ud patients ( out of - %). this study shows a comparable duration of admission for transplant for haplo and ud patients, both significantly longer than sib grafts. the number of re-admissions is comparable in haplo vs sibs and there is a trend for lower number of re-admission as compared to uds. we interpret this outcomes with caution given the relatively small sample size and heterogeneous disease population included. future studies need to confirme our results. disclosure of conflict of interest: none. prolonged thrombocytopenia (pt) is frequent event after allogeneic haematopoietic stem cell transplantation (hsct), especically in haploidentical transplantation, which could be up to % according to our previous report. pt has significant negative impact on long-term outcomes, mainly due to increased non-relapse mortality. however, there are no efficious treatment. in this study, we report the preliminary results of recombinant human thrombopoietin (rhtpo) in treating this kind of patients. from . to . , patients were enrolled under the following inclusion criterion: ( ) diagnosed with dpe or sfpr after allogeneic stem cell transplantion; ( ) no sign of minimal residual disease or recurrence of hematological malignancy; ( ) not using other tpo receptor agonist or il- within month of enrollment. pt include delayled platelet engraftment (dpe) and secondary failure of platelet recovery (sfpr). the former was defined as failure to achieve platelet counts ⩾ /μl for consecutive without transfusion until days after transplantation, while the latter was defined as a decline in platelet counts below /μl for consecutive days, or requiring transfusion support after achieving sustained counts without transfusions for consecutive days after hsct. the prescription of rhtpo was iu once daily for days, or if patients achieve platelet ⩾ /μl for consecutive days with a duration o days. response was defined as success of achieve platelet counts ⩾ /μl for consecutive days. the response time was defined as the first day achieve response from the start of prescription. the primary end point was response rate, and the secondary end point was reponse time. a total of patients were enrolled, including males and females. the median age was ( - ) years. all patients received haploidentical transplantation. among these patients, patients were dpe and were sfpr. all patients received a -day prescription. the overall response rate was % ( out of ) in the overall population, while % ( out of ) in dpe and . % ( out of ) in sfpr, respectively. among the patients with response, the median response time was ( - ) days from the first dose of rhtpo. after weeks of the last dose of rhtpo, none of the responsed patient lose response. since the followup time is too short, the impact of relapse, gvhd were not reported. this single-arm preliminary result suggest that rhtpo could be a efficious method to manage pt after stem cell transplantation. however, these result need further confirmation. disclosure of conflict of interest: none. reproductive health in long-term female survivors after allogeneic hematopoietic stem cell transplantation z peric , , a samardzic , n durakovic , , d tina , , l desnica , r serventi-seiwerth and r vrhovac , school of medicine, university of zagreb, zagreb, croatia and university hospital centre zagreb, zagreb, croatia most female recipients of allogeneic hematopoietic stem cell transplantation (allo-hsct) suffer from premature menopause, infertility and endocrine imbalance owing to gonadal damage from myeloablative conditioning. in order to evaluate ovarian recovery and long-term endocrine complications in our institution, we performed a retrospective study of female patients who received a myeloablative allo-hsct during their reproductive age. we identified female patients who underwent myeloablative allo-hsct in our institution between and and were still alive with available follow-up at the time of this study. among them, patients accepted to participate and responded to a query designed for this s purpose. the median age of our patients at transplantation was years (range: - years). they were interviewed at a median of years (range: - years) post allo-hsct. the majority of patients were transplanted for a myeloid malignancy ( acute myeloid leukemia, chronic myeloid leukemia, myelodysplastic syndromes and chronic myelofibrosis), while patients had aplastic anemia and had acute lymphoblastic leukemia. all patients received bone marrow transplant from a hla-matched related donor after a myeloablative conditioning. conditioning regimen consisted of cyclophosphamide with or without total body irradiation (tbi) or in combination with busulfan. only patients ( %) resumed a normal menstrual cycle after allo-hsct, without the need for hormonal replacement therapy (hrt). all these patients were transplanted for aplastic anemia and none of them received tbi in the conditioning regimen. eight patients ( %) remained amenorrheic indefinitely and never started hrt, even though most of these women were transplanted under the age of years. % of these patients were diagnosed with osteoporosis later in life. the remaining patients ( %) started hrt at a median of months after allo-hsct (range: - months). however, only seven patients on hrt ( %) resumed regular menstrual cycle. a median duration of hrt therapy was years (range: - years). none of the women receiving long-term hrt had severe cardiovascular complications or breast cancer. finally, five women gave birth to eight healthy children in our study population. three unassisted pregnancies were observed in two female patients after spontaneous recovery of ovarian function (both patients with aplastic anemia). the remaining two patients restored ovarian function with the use of hrt and gave birth after an assisted pregnancy (one woman gave birth to triplets after an in vitro fertilization (ivf), while other became pregnant with a donated oocyte). in spite of the fact that almost all women who undergo allo-hsct develop an ovarian failure, spontaneous recovery is sometimes possible, particularly following conditioning regimen without tbi. in patients without spontaneous recovery, hrt should be initiated promptly to prevent the early and late unwanted effects related to estrogen deficiency. moreover, recovery of normal ovarian function and even a viable pregnancy is a realistic possibility in patients placed on hrt, particularly with the use of potential therapeutic interventions as ivf or oocyte cryopreservation. it is therefore crucial to provide adapted pre-transplant counselling and recommendations for regular post-transplant follow-up in female patients who undergo allo-hsct. disclosure of conflict of interest: none. transplant-associated thrombotic microangiopathy (ta-tma) is a multifactorial disorder caused by systemic vascular endothelial injury leading to end-organ damage often involving the kidney. ta-tma occurs in up to % of patients undergoing hsct, and may be associated with poor outcome. although pathogenesis has not been fully clarified, activation of the complement system has been suggested to play a central role, and eculizumab, a monoclonal antibody (mab) that mediates terminal complement blockade, has shown therapeutic benefit in cases unresponsive to immunosuppression modulation. we report the case of a pediatric allogeneic hsct recipient with severe ta-tma, who did not tolerate treatment with eculizumab, now successfully treated with oms , a novel human mab targeted to the mannan-binding lectin-associated serine protease- (masp- ), a molecule central to the activation of the lectin pathway of complement. a -year-old girl received an allogeneic hsct from a hla-compatible unrelated donor for the treatment of diamond-blackfan anemia. at month + of the posttransplant course, she developed progressive deterioration of renal function, microhematuria and serositis, that prompted the cyclosporine discontinuation. from month + , the patient experienced progressive trilinear cytopenia, elevated ldh, schistocytes, undetectable haptoglobin, hypertension, increased serum creatinine, nephrotic range proteinuria, and serositis, and a diagnosis of ta-tma was established. laboratory investigations documented no abnormalities in the patient but identified a stop-codon heterozygous variant in cfhr c. _ dupaa (p.glu lysfs* ) in the donor's dna. the patient was initially treated with eculizumab, but she developed acute pulmonary edema soon after eculizumab administration as the consequence of a possible reaction to the drug which had to be discontinued. the patient was subsequently treated with plasma exchange, with only limited benefit. upon ta-tma relapse at month + , eculizumab was re-administered at lower doses, but she developed a new episode of acute pulmonary edema, preventing further eculizumab continuation. renal function progressively deteriorated and she was started on hemodialysis, reaching a times weekly regimen. the patient received oms , kindly provided on a compassionate use basis by omeros corporation, seattle, usa, starting with an iv dosing schedule. she did not experience any adverse events, and was able to tolerate the treatment well. at months from oms initiation, she has shown improvement in ldh and haptoglobin levels, and, more importantly, her creatinine levels have normalized, allowing for complete discontinuation of hemodialysis and partial outpatient management. anti-masp- mab oms is a promising new option for the treatment of ta-tma occurring after hsct, and seems to have a safe profile also in the pediatric/adolescent setting. disclosure of conflict of interest: none. severe cytokine release syndrome after t-cell replete haploidentical transplantation with post-transplant cyclophosphamide is associated with increased death rate d taurino , j mariotti , b sarina , l morabito , s bramanti , c carlo-stella , a santoro and l castagna bone marrow unit, humanitas cancer center, istituto clinico humanitas, rozzano, italy and hematology department, humanitas cancer center, istituto clinico humanitas, rozzano, italy haploidentical stem cell transplant (haplo-sct) represents a potential curative strategy for several hematological malignancies. haplo-sct may represent an alternative option when a hla matched-identical sibling (hlaid) or a matched unrelated donor (mud) is not available. the syndrome of systemic inflammation, characterized by fevers, vascular leak, hypotension, and respiratory and renal insufficiency, in the context of elevated inflammatory markers and cytokine levels was previously described as cytokine-release syndrome (crs) . recent publications have elicited the occurrence of crs after haploidentical transplant, especially after peripheral blood stem cell graft, and its high-related mortality - . here we report the experience of our institution with crs after haplo-sct. between march and october , we treated patients with haplo-sct with a graft source represented by peripheral blood stem cells. we monitored the occurrence of crs symptoms and utilize a previously described grading system , starting from day , up to day after transplant. severe crs is defined as grade or higher because it requires aggressive interventions and is characterized by oxygen requirement ⩾ %, l nasal cannula, hypotension requiring high dose or multiple vasopressors, grade renal toxicity or grade transaminitis. other characteristics comprise newonset altered mental status without other explanation and new cardiomyopathy without wall motion abnormality. results: out of patients experienced fever between day and day post transplant with most episodes ( patients) occurring between day and day . on day after transplant, patients had grade , grade and grade crs, respectively. by day post haplo-sct, patients had crs grade , grade and grade . overall, the incidence of crs any grade was % ( % ci - %). year after transplant patients died because of non-relapse related side effects. with a median follow-up for alive subjects of months, -year overall survival (os) was % ( % ci: - %). -year os was % for patients with a crs on day (p = . ). conclusions: crs represent an important complication after haplo-sct. crs score on day after hst apparently correlates with long-term survival. better strategies need to be implemented for an early detection of severe crs in order to develop effective treatments, such as tocilzumab, for this important side effect. further studies are ongoing at our institution in order to correlate post-haplo crs with graft composition, laboratory parameters and immunereconstitution. hematopoietic cell transplantation (hct) is associated with significant morbidity that impairs survivor's sexual functioning. however, few studies have specifically addressed it. thus, we examined ( ) sexual functioning during the first year post hct, ( ) differences between allogeneic and autologous hct, and ( ) whether demographic, clinical and psychological variables were associated with sexual functioning. this is a prospective multicenter study assessing patients before hct, at day , and . sexual functioning was assessed with the changes in sexual functioning questionnaire, which yields a total score, along with scores for the dimensions of frequency, pleasure, orgasm, desire and arousal. anxiety and depression (hads) were also collected. we included consecutive hct recipients: ( %) were men, with a median age of years (range: - ), ( %) received an allogeneic hct and ( %) an autologous hct. sexual functioning was significantly affected: % of the sample reported impairment at pre-hct, % at day , % at day and % at day . mixed model analysis indicated that sexual functioning was not associated with time from hct (p = . ) or hct type (p = . ). however, there was an interaction between these two variables (p = . ), particularly at day , since sexual functioning had improved among autologous survivors and worsened among allogeneic survivors leading to nonsignificant differences between hct type (p = . ). frequency of sexual functioning improved during the study period (po . ), and no differences were observed between hct type (p = . ). again, there was a borderline interaction between post-hct time and hct type (p = . ), since autologous survivors reached higher frequencies than allogeneic survivors, with significant differences at day (p = . ). pleasure significantly improved during the study period (p = . ), without observing differences between hct groups (p = . ). again, however, autologous survivors reported significant improvements in pleasure at day (p o . ) and a trend at day (p = . ) when compared with allogeneic survivors. orgasm did not improve during the study period (p = . ), and no differences were obtained between hct groups (p = . ). allogeneic survivors had higher orgasm scores at pre-hct (p = . ), which worsened during the study period, particularly at day (p = . ). in contrast, autologous survivors reported improvements in orgasm by day . non significant results were obtained in the sphere of sexual desire and arousal (p . ). bivariate analyses indicated that women, older age and depression were associated with impaired sexual functioning at all assessed time-points (p o . ). chronic graft-versus-host disease (gvhd) was associated with worse sexual functioning at day (p = . ) and (p = . ). no differences were obtained when considering diagnosis, having received previous hct, intensity of the conditioning regimen and whether patients lived with a partner (p . ). stepwise multivariate regression analyses indicated that gender (p = . ) and extensive chronic gvhd (p = . ) predicted for worse sexual functioning at day . sexual functioning should be routinely assessed and considered for eventual targeted intervention in both hct populations, particularly during the first year post transplant. additional clinical efforts should focus on patients more vulnerable to impaired sexual functioning. disclosure of conflict of interest: none. significant improvement of qol by using atg as part of the conditioning regimen followed by hla-identical peripheral stem cell transplantation in acute leukemia patients. results from a prospective, randomized phase iii study (atg family study) b francesca , s carlos , w christine , s mariarosaria , p massimo , s carmine , m giuseppe , b wolfgang , cm angelo , p francesca , m nicola cgvhd is a major complication after allogeneic sct. we previously demonstrated that the addition of anti-tlymphocyte globulin (atlg neovii, formerly atg-fresenius) to a myeloablative preparing regimen followed by peripheralblood sct from an hla-identical sibling for pts with acute leukemia resulted in a significant reduction of cgvhd, without increasing the risk of relapse or infection. the study protocol included quality of life (qol) questionnaires (eortc qlq- and hdc ) before and after sct (day+ , , and mos). the qlq-c includes a global qol scale, five functional scales (physical, role, emotional, cognitive and social function) and nine symptom scales (fatigue, nausea-vomiting, pain, dyspnea, insomnia, appetite loss, constipation, diarrhea and financial problems). the qlq-hdc includes six multi-item scales and eight single items that describe impairment through highdose treatment. mixed models for repeated measures (mmrm) and linear mixed models (lmm) were used to analyze the time courses and the slopes of the outcomes depending on treatment arm (atg vs non atg), age, country, sex, and cgvhd. (clinicaltrials.gov: nct ). pts with a qol form returned decreased by visit ( % pre-sct, % at days and % at mos after sct). forty-nine percent in the atg and % in non atg arm provided any qol forms after sct. return of any post-sct qol forms by country was % for germany, % for italy and % for spain. pts with cgvhd were more likely to return qol questionnaires ( % vs % w/out cgvhd) while neither age nor sex were closely associated with qol form return. the majority of subscales of the qlq- indicated an average improvement of qol and reduction of symptoms over time, notably in the atg group. in an mmrm model controlling for country, age, sex and cgvhd, pts treated with atg showed significantly more pronounced improvement of global health status/qol over time compared to non-atlg (p = . ), with a treatment group difference of . ± . points (marginal mean ± sem) at day and increasing to . ± . points at month favoring atg. significant superiority of atg (po . ) was also observed for four of the five functional scales as well as for several symptom scales scores including appetite loss, insomnia, nausea-vomiting and dyspnea. for the qlq-hdc , significant treatment effects favoring atg were observed for gi side effects and impact on family. lmm analyses of qol by country indicate that patients from italy generally gave more favorable ratings for all functional scales and lower scores for most symptom scales than those from germany while the time courses and slopes were similar for most scales. these results underline the importance of the habits and cultural environment which are distinctive of each country. males and females showed similar qol ratings at pre-and post-sct. patients up to years tended to provide more favorable functional ratings and less severe symptom scores than older patients and also showed more pronounced improvements of qol. pts receiving atg in a randomized study have significantly less cgvhd and improved grfs, resulting in an improved qol regarding global health status and most functional scales. notably, we also observed a significant difference in qol assessment between pts from germany and italy. oral mucositis (om) is a well-known side effect of high-dose chemotherapy and radiotherapy in hematological patients, which influences the health-related quality of life (hrqol) of the affected patients. the purpose of this study is to demonstrate the impact of om on hrqol in stem cell transplanted patients in routine care. prospective, noninterventional single-center observational study was performed at a german university hospital. inpatient allogenic and autologous stem cell transplant patients ⩾ years with high-dose chemotherapy. om was assessed with the who oral toxicity scale, pain using the numeric rating scale (nrs) and the performance status with the ecog score. hrqol was captured with the eortc qlq-c and the qlq-oh questionnaires ( days before hematopoietic stem cell transplantation (hsct); days after hsct; days after hsct). statistical significance was assumed p o . . a total of patients ( autologous and allogenic) was included from august to december . a total of ( %) patients developed om. of these patients, suffered from grade , from grade , from grade and from grade om. three days before hsct, the mean qol of all patients was %, the mean qlq-c summary score . % and the mean oral health related quality of life . %. most of the patients suffered from om around day after hsct. after days, quality of life (qol) was higher in patients with no om ( . %) than in patients with om ( . %). the qlq-c summary score was significantly (p = . ) lower in patients affected by om ( . %) than in patients who did not develop an om ( . %). om affected patients had significantly more limitations in emotional (no om . %; om . %; p = . ) and cognitive functioning (no om . %; om . %; p = . ) and in fatigue (no om . %; om %; p = . ), pain (no om . %; om %; p = . ) and insomnia (no om . %; om %; p = . ), they had a significantly higher rate of problems. oral health-related quality of life was significantly (p = . ) lower in patients who were affected by om ( . %) compared to patients who did not develop an om ( . %) and patients with an om had significantly more problems with a sore mouth (no om . %; om . %; p = . ), sticky saliva (no om . %; om . %; p = . ) and sensitive mouth (no om . %; om . %; p = . ). after days, qol was higher in patients with no om ( . %) compared to patients with om ( . %). patients with no development of om had a higher but not significant physical functioning, cognitive functioning and social functioning. patients affected by om had higher levels of fatigue and pain and more often suffered from a sore mouth. oral health-related quality of life was higher in patients without om ( %) compared to patients with om ( . %). comparing all assessed days patients with om had higher scores on the nrs increasing with a higher grade of om (mean nrs score grade ; - . , grade ; - . ), the ecog index was higher in om affected patients during episodes with om (mean ecog score- . ) compared to episodes without om (mean ecog score- . ). om has a major impact on the hrqol, health-related symptoms and functionality. in the future, there has to be a higher awareness from clinicians and patients of the prevention, assessment und causes of om. more research has to be initiated to ease the symptomatology and to improve patients' quality of life. disclosure of conflict of interest: none. according to ebmt data, chronic gvhd (cgvhd) occurs in - % of all patients after allogenic hematopoietic stem cell transplantation (allo-hsct). pulmonary cgvhd is the most severe form. but it is very unpredictable to use due to the fact that many factors can affect it (breath-dependent; need experience not only from physician but from patients also and so on). here we show that routine software-based image analysis algorithm can provide data that highly correlated with pft results and have excellent sensitivity and specificity in pulmonary cgvhd diagnosis. we blindly analyzed ct scans (made without additional expiration) in allo-hsct patients at different time points. all scans were performed on ct scanner aquilion , toshiba, japan. according to hounsfield units (hu) definition, − hu ('air') have approximate density at g/ml; hu ('water') have approximate density at g/ml. the analysis of ct scans (heart, vessels and bronchi were excluded from analysis) was based on automated software conversion (image-analysis algorithm providing by multivox software, msu, moscow, russia) of each ct-image pixel from hu to density units (g/ml). pft were performed using standard procedures at same as ct scans time points (spirolab iii, italy). all patients with hematological malignancies (acute leukemia- , aplastic anemia- , chronic myeloid leukemia- , t-cell lymphoma- , chronic myeloproliferative disorder- , myelodysplastic syndrome- ) were transplanted in national research center for hematology between and . median of age was . years (range: - years). eight patients were males, -females. seventeen received reduced-intensity and -myeloablative conditioning regimen. graft from match unrelated donor (mud) were used in cases, 'mismatch' mud- , match related donor (mrd)- , 'mismatch' mrd- . median follow-up is . months. we analyzed lung tissue experimental density in patients before and after allo-hsct at different time points. median of lung tissue experimental density were . (interquartile range (iqr), . - . ), . (iqr, . - . ) and . (iqr, . - . ) for patients before allo-hsct, after allo-hsct with cgvhd (except pulmonary cgvhd) and with pulmonary cgvhd, respectively. mann-whitney u test was used to reveal significant differences between these groups (see figure ). also, we found strong correlation between pft and experimental density (spearman's correlation coefficient r = . ) (see figure ). forty-five ct scans of patients with pulmonary cgvhd and ct scans of patients without pulmonary cgvhd at the time of ct scan as control subjects were included in roc analysis to assess the clinical values of our model. we generated an roc curve and found that the area under the curve (auc) was . ( % ci, . - . ) (p o . ) (figure ). standard ct scan is presented as easy to perform, breath-independent, standardized and wide spread method for every patient after allo-hsct. it can be performed many times during all their post-hsct life. ct scan with a simple software analysis allows to select a group with high probability of pulmonary cgvhd and who can be suspected of cgvhd development by this method with sensitivity- % and specificity- . %. disclosure of conflict of interest: none. the choice of effectiveness criteria affects conclusions of economic evaluation of newer allogeneic bone marrow transplantation modalities :example based on a randomised multicenter trial comparing two reduced intensity conditioning regimen (flu-bu-atg) vs (flu-tbi) for matched related allo-sct s le corroller*, anne-gaelle , c siani , , r tabrizi a re-evaluation of the per-diem hospitalization cost was performed in and included the utilization of hospital technical facilities and a more precise estimation of overheads costs. we performed three separated cost-effectiveness analysis, using, respectively, pfs, os and qaly as end point. when using pfs as effectiveness, relapse costs were not included. weighting coefficients for the cost per qaly analysis came from the literature. at years, os and pfs were % and %, respectively, and did not statistically differ between groups. the mean total cost per patient was not statistically different between groups ( € for fba vs € for ftbi, ns). using pfs as end point, the icer of fba compared to ftbi is € per year of pfs gained. using os, the icer became non-statistically significant, signifying that when handling uncertainty, no difference in term of cost-effectiveness was observed between fba and ftbi with os as end point. using s qaly, the icer was statistically ns again, showing no advantage in terms of cost per qaly of one conditioning regimen over the other. this result was obtained both considering three weighted health states (dfs, progression and death) and four weighted health states (dfs without gvhd, dfs with gvhd, progression and death) for the qaly calculation. using os and qaly, the two conditioning regimens were not different in terms of cost-effectiveness, while fba may be considered as more cost-effective using pfs as effectiveness criterion. using intermediary end points allows economic evaluation to be available earlier in the life cycle of an innovation. however, it implies strong hypotheses about the predictive value of the pfs over the os. longer period evaluation and qaly may reverse preliminary results. this situation is likely to exist in the hematology setting where alternatives between chances of cure and toxicities of treatment are often observed. research about allogeneic sct modalities is archetypical of such situations and decisions makers should be aware of the necessity of further economic re-evaluation along the development and diffusion process of innovative treatments. disclosure of conflict of interest: none. the impact of corticosteroids prophylaxis for the engraftment syndrome incidence during autologous stem cell transplantation in multiple myeloma and amyloidosis the es is a complication of asct characterized by an inflammatory response during peripheral blood recovery. the standard treatment is based on corticosteroid therapy. the incidence of es after asct increases in chemotherapy lowtreated patients such as those with multiple myeloma (mm) and amyloidosis (al).moreover, the es is associated with the use of g-csf after infusion of stem cells. therefore, our bmt team does not use g-csf since in this population reducing the incidence and severity of es. therefore, it makes sense to use low-dose prednisone to prevent this complication. in this study, we compared two consecutive cohorts of patients with mm/al that performed an asct while evaluating the corticosteroids prophylaxis (cp) in the es incidence and its effect on other clinical variables. we included patients with mm (n = ; %) and al (n = ; %) that performed an asct between january and november in a single institution. the median age (range) was . ( . - . ) years. during the procedure, all patients received melphalan as conditioning chemotherapy and none received g-csf. fortyseven patients ( %) received intravenous methylprednisolone or oral prednisone . mg/kg/day from day + until reaching a neutrophil count ⩾ per mm for consecutive days (cs group), and ( %) patients did not receive corticosteroids (noncs group). the characteristics of patients in both groups (age, gender, status performance and previous treatment were similar (p . )). the cs group, received higher doses of cd + than the noncs group ( . × /kg vs . × /kg, respectively, p = . ). the median (range) days of neutropenia ( o per mm ) was ( - ) days. es was diagnosed in ( %) patients. fifty-seven ( %) patients had fever, showing infectious focus or microbiological isolation in ( %) cases, whereas the incidence of grade iii-iv oral mucositis and relevant gastrointestinal toxicity was % and . %, respectively. the complete analysis between groups (cs versus noncs) for the whole series and in the mm/al subgroups is detailed in table . the administration of corticosteroids as prophylaxis seems to reduce the incidence of es in the overall series or in the analysis for the subgroups (mm and al) without increasing infection. [p ] disclosure of conflict of interest: none. chronic gvhd is a condition that might occur after allo-hsct and has been proved to impair long-term survival and quality of life of patients. graft failure is also a major potential complication for patients undergoing transplant for an aplastic anemia/bone marrow failure (bmf). partial in vivo t-cell depletion, employing anti-thymocyte globulin (atg) during conditioning, has been proved to successfully prevent the mentioned potentially life-threatening complications in highrisk patients. however, the possibility of developing epstein-barr virus (ebv)-induced post-transplant lymphoproliferative disorders (ptlp) has been a limiting factor to use atg. this study includes the last pts with a minimum follow-up of days, who underwent allo-hsct in our center (november -august ). a total of pts were male and female. median age was years (range: - ). baseline diseases were: acute leukemias ( ), lymphoproliferative disorders ( ), myelodysplastic syndromes ( ), chronic myeloproliferative diseases ( ), multiple myeloma ( ) and bone marrow failures ( ) . donor was unrelated in cases, and related in (including haplo-identical). conditioning regimen was: busulphan-based ( ), melphalan-based ( ), tbi-based ( ) and others ( ). progenitors source was pb in and bm in . patient/donor ebv pre-transplant serology was: +/+ in cases, +/ − in and − /+ in . rabbit atg (thymoglobuline) was employed in cases: at . - mg/kg (urd transplants) (low dose), and cases at . mg/kg (all of them pts with bmf) (intermediate dose). family donor (including haplo-identical) transplants of those pts with diagnosis different from bmf ( cases) did not receive atg. systematic monitoring of ebv using quantitative pcr was employed. ebv reactivation was considered when dnaemia was superior to copies per ml. a total of pts presented ebv reactivation: / ( %) in cases without atg, / ( . %) in cases with low-dose atg and / ( %) in cases with intermediate-dose atg. median time of reactivation was the day + (range: + to + ). there was one single case of ebv-induced ptld which belonged to the intermediate-dose atg group. all cases (including the one with ptlp) were successfully treated with rituximab at mg/m /week. median number of doses employed were (range: [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] . mortality due to ebv was % in our series. limited donor availability in the form of either matchedrelated or unrelated donors drew attention to haplo-hct. donors of haplo-hct shares an exact haplotype with the recipient but is mismatched for hla genes on the unshared haplotype. most studies have shown promising results in terms of graft success and survival. in this study our aim is to present the early and late outcome of our haplo-hct patients. between and , we retrospectively evaluated haplo-hct in terms of post-transplant outcome, survival and complications who diagnosed and followed in our center. the median age of patients was (range: - ), ( %) of them were male recipients. the patient characteristics were given in table . thirteen patients ( %) had pre-transplant active disease. neutrophil and platelet engraftment was achieved in patients ( %) at a median day of (range: - ) and (range: - ). eight of patients ( %) died within month after transplant because of sepsis without achieving engraftment. haplo-hct is the second transplant in four of patients ( %): patient relapsed after full-matched related transplant, patient relapsed after / matched unrelated transplant, patient had engraftment failure after full-matched unrelated transplant, patient underwent haplo-hct in another center, followed in remission for years and relapsed. acute graft vs host disase (agvhd) was diagnosed in patients ( %), whereas chronic gvhd in patients ( %). four patients were relapsed ( %) during follow-up with median rfs of months. three patient had bk virus-positive hemorrhagic cystitis ( %). the distribution of infections is shown in figure, viral infections were detected later than fungal and bacterial infections. previous history of invasive pulmoner aspergillosis was detected in of the patients ( %) ( of them were re-transplanted) and received secondary prophylaxis. overall survival (os) of months and year were % and %, respectively. the choice between alternative graft sources depends on the urgency of the transplant on each institutional preference. higher complication and infection rates in addition to decreased survival compared with previous studies since our patient population consisted of refractory patients with comorbidities. preferable patient profiles undergoing haplo-hct may have better outcomes. disclosure of conflict of interest: none. the third month risk factor score: detection of disease at day + of allogeneic stem cell transplantation is the most important risk factor of worse prognosis m celis , c fernández , l yáñez , , a bermúdez , , a insunza , m colorado , m lópez-duarte , i romón , s garcía-Ávila , a cabero , a casado , m sánchez-escamilla , c richard and e conde , hematology department, hospital universitario marqués de valdecilla and university of cantabria before allogeneic stem cell transplant (sct), several index can provide prognostic information (ebmt risk score and hcti score). however, there is scarce data for the impact of the procedure during the first days of transplant, in which opportunistic infections and the acute graft versus host disease (gvhd) can induce harmful effects. our purpose is to create a risk factor score, measured at day + post sct, to give information about the prognosis of the patient. we retrospectively analyzed seven clinical (disease, fungal and cmv infection, acute gvhd, treatment with corticosteroids, karnosfsky status and length of hospitalization) and eight analytical (related to immune status, liver and lung function, nutritional status, iron overload and platelet count) risk factors in patients who underwent sct in our center between and and were alive at day + . data were collected as categorical variables and compared by χ -test. significant variables (p o . ) were evaluated in a multivariate logistic regression model. those who maintained statistical significance were then assigned a point value calculated with their β-coefficient. summation of the points resulted in a weighted risk score. median age was years (range: - ) and were males ( . %). the most frequent disease was aml, patients ( . %). the conditioning regimen was myeloablative in patients ( %) and bone marrow was the principal stem cell source ( %). donor was mrd in ( . %), mud in ( . %) and mmd in ( . %). the median followup was months (range: - ). the univariant model identified five prognostic variables: detection of disease by molecular, cytogenetic or flow cytometry asses in leukemias, myelodisplastic syndrome and multiple myeloma or image (ct scan ± pet) in lymphoma, dose of corticosteroids ⩾ . mg/kg/ day, ferritin ng/ml, albumin o . g/dl and platelet o per mm . table shows variables evaluated. in the multivariate model, the detection of disease (hr . , % ci . - . , p ng/ml (hr . , % ci . - . , p = . ), and platelet o per mm (hr . , % ci . - . , p = . ) were associated with higher risk of death and according with their-coefficient , and points were, respectively, assigned. the third month risk score (tmrs) was calculated in all patients and they were stratified into three groups: low risk of death (a, - points), intermediate risk (b, points) and high risk (c, ⩾ points). at years post sct, the estimated overall survival according with the tmrs was . % ± . in group a, . % ± . in group b and . % ± . in group c, po . . although the harmful effect of the first months of transplant can impact in the survival, the detection of disease at day + is the most determinant risk factor of death. this fact gives us the need of transplant in the best response and, in those who cannot, to plan promptly rescue strategies. the next objective is to confirm our risk score in a validation group. disclosure of conflict of interest: none. recombinant human soluble thrombomoduline alpha (rhtm) is a novel anticoagulant agent and approved for disseminated intravascular coagulation in japan. the aim of the study is to evaluate the therapeutic potential of rhtm for sinusoidal obstructive syndrome/hepatic veno-occlusive disease (sos/ vod). we retrospectively studied times of allogeneic hematopoietic cell transplantation in toranomon hospital from june to june . we extracted the patients who used rhtm for dic and satisfied the diagnostic criteria of sos/ vod around the same time, because the use of rhtm for sos/ vod alone is off-label. data on the patients who used rhtm for days within days after transplantation were analyzed. the patients who were already treated with rhtm before the emergence of the first symptom or sign of sos/vod, and who started rhtm over days after the emergence of the first symptom or sign of sos/vod, were excluded from the [p ] analysis. to diagnose classical sos/vod (⩽ days after transplantation), we used two classical criteria of the modified seattle and the baltimore. for late-onset sos/vod ( day of transplantation), we used the criteria of ebmt. we defined as severe sos/vod, if the patients had renal (cr ⩾ times of baseline), respiratory (spo ⩽ % or the need for positive pressure) or central nervous system failure until weeks after the diagnosis of sos/vod. complete response (cr) was defined as the resolution of all the symptoms and the signs in sos/vod diagnostic criteria. a total of patients were extracted. the median age was years (range: - ) and patients ( %) was male. donor cell sources were ucb (n = ) and ubm (n = ). most of the prophylaxis regimen was the combination of ursodeoxycholic acid and dalteparin in patients ( %). classical sos/vod was diagnosed in ( %) and patients ( %) by the criteria of the modified seattle and the baltimore at the median day of (range: - ) and (range: - ), respectively. twenty-eight patients ( %) were diagnosed as late-onset sos/vod at the median day of (range: - ). severe sos/vod developed in patients ( %) (renal, n = ; respiratory, n = ; central nervous system, n = ). the elevation of transaminase was observed in patients ( %). the median interval from the emergence of the first symptom or signs of sos/vod to rhtm administration was days (range: - ). the median duration of rhtm use was days (range: - ). rhtm was used alone in patients ( %), in combination with dalteparin in ( %), with atiii in ( %), with dalteparin and atiii in ( %), with atiii and pge in ( %), and with pge in ( %). corticosteroid was used concomitantly in patients ( %). finally, patients achieved cr of sos/vod. the cumulative incidence of cr of sos/vod was . % at year after the administration of rhtm ( % confidence interval, . - . %). the median interval from the administration of rhtm to cr of sos/vod was days (range: - ). at year after transplantation, overall survival was . % ( % confidence interval, . - . %). from the administration of rhtm to weeks after the cessation of rhtm, hemorrhagic adverse events were observed. seven out of events were at grade - , and out of events were fatal (intra-abdominal in , gastrointestinal in , lung in and brain in ). we concluded that rhtm had a therapeutic potential for sos/vod. disclosure of conflict of interest: none. thrombopoietin receptor agonists for delayed and prolonged clinically-relevant severe thrombocytopenia after allogeneic hematopoietic stem cell transplantation v bosch vilaseca , i garcía cadenas , e roldán , s novelli , r martino , p barba , a esquirol, l díaz polo , g orti , d valcárcel and j sierra hematology department, hospital de sant pau, barcelona, spain and hematology department, hospital de la vall d'hebron, barcelona, spain persistent thrombocytopenia is a common complication after allogeneic stem cell transplantation (allosct), which dramatically increases the patients' dependence on hospital-based healthcare. thrombopoietin receptor agonists (tpoa) increase platelet counts in other clinical settings; however, the experience regarding their use after allosct is limited. we retrospectively evaluated tpoa efficacy in consecutive adult allosct recipients who received tpoa as a compassionate use for severe thrombocytopenia post-engraftment. five patients ( %) had primary and prolonged failure of platelet recovery, while had secondary thrombocytopenia: in seven of these cases, gvhd and/or a viral infection were the 'attributed' cause, while three were classified as post-allosct itp. all patients were dependent on platelet transfusions (median: times per week, range - ), with severe bleeding episodes in nine cases ( %) before tpoa onset. tpoa was started at a median of days after allosct (range: - ). romiplostim was used in ( %) cases. the median starting dose was μg/kg once a week (range: - μg/kg), while the final dose identified as most beneficial was μg/kg (range - μg/kg). eltrombopag was used in cases ( %), with an initial dose of mg daily; while the final doses were and mg daily. overall, / patients responded to tpoa therapy (defined as a stable platelet recovery to ⩾ /μl without transfusion support). the -day cumulative incidence of successful platelet recovery to ⩾ /μl and ⩾ /μl was % ( % ci, - %) and % ( % ci, - %), respectively, which were reached at a median of and days from start of therapy. five of the patients ( %) with severe bleeding at onset responded to tpoa ( of them without further hemorrhages) at a median of days (range: - ). at a median follow-up of days from start of therapy, three patients who responded continue tpoa treatment, while four other responders were able to discontinue it without recurrence of thrombocytopenia. among these patients, s the median total duration of treatment was days (range: - ). one patient lost his response within months after tpoa onset when he developed thrombotic microangiopathy associated with progressive gvhd. the remaining responder experienced disease relapse on day + after allosct. among the non-responders, had leukemia relapse during tpoa treatment, switched from romiplostim to eltrombopag without success and the remaining cases had active severe infections at tpoa onset ( hemorrhagic cystitis and cmv colitis) or non-controllable intestinal bleeding due to progressive gvhd. tpoa were well tolerated, with only patients showing adverse events (grade liver toxicity and grade fatigue), which did not lead to any change in therapy. six patients ( %) underwent follow-up bone marrow biopsies that did not display any increase in marrow fibrosis, including the patient who had myelofibrosis prior to allosct. although six patients in the study had active gvhd when tpoa was started, no patients showed worsening of gvhd. our results support the safety and efficacy of tpoa for the treatment of persistent thrombocytopenia in allosct recipients. further studies should compare the efficacy of romiplostim and eltrombopag and identify surrogate clinical and laboratory variables that are predictive of response to one (or both) of these tpoa. disclosure of conflict of interest: none. . clinical response in both groups was defined as improvement of organ function (no neurological residues; normalization of kidney function) and independence of red blood cell and platelet transfusions. results: the median time of ta-tma onset was . months ( . - . ) after hsct. thirty-five of patients ( %) were under treatment with calcineurin-inhibitors or sirolimus at the time the ta-tma occurred. in all cases, the immunosuppressive drug was stopped promptly. in patients, classical treatment was the primary therapy with a response rate of % (including four patients who switched to ec), whereas the response rate to ec treatment was significantly higher with % (p = . ). all patients receiving ec showed sufficient blockade of the terminal complement pathway after the second ec application (ch o %). despite the increased response rate for ec therapy, there was no difference seen between these two groups according to overall survival in weeks: classical treatment ( % ci - . ) vs ec treatment . ( % ci . - . ) p = . . the main cause of death differed significantly between this two treatment approaches with a therapy-related mortality due to infection with % in the ec group during tma therapy and none seen in the classical treatment group (p = . ). progressive gvhd was identified as an adverse prognostic factor in both groups (p = . and p = . ). conclusion: in our analysis, we show that ec shows a significant higher response rate in severe ta-tma patients compared to the classical treatment approach. however, in both groups the outcome remained very poor. since most patients presented with advanced, severe ta-tma, especially in the ec group, we hypothesize that earlier diagnosis and treatment of ta-tma and more effective prevention and treatment of infections will improve the outcome of patients with this complication. however, randomized studies are essential for comparison of these two treatment strategies to identify patient groups that benefit from a treatment with ec. disclosure of conflict of interest: none. tocilizumab as an effective treatment in cytokine release syndrome as an early peri-transplant complications in patients subjected to allogeneic stem cell transplantationproinfammatory/autoimmune patient/donor hla haplotype life-threatening early allogeneic hsct complication risk factor hypothesis m-g patrycja , , p-j beata , , s marcin , k ksenia , s-k agnieszka and sb aleksander , bone marrow transplantation unit, department of haematology, krakow university hospital and jagiellonian university collegium medicum cytokine release syndrome (crs) is classical complication of car t cells therapy, but also can be connected with early peritransplant complications in patients subjected to allogeneic stem cell transplantation. it can be connected with atg infusion, but also with inflammatory response during periengraftmetnt period (pre-engraftment syndrome and engraftment syndrome) and septic infections. severity of these complication can differ depending on patient's performance status and therapeutic options from just observation and vigilance to mechanical ventilation need. we would like to present small patient series (n = ) subjected to msd (n = ) and mud (n = ) with early transplant-related complications treated with combination of steroids (dexamethason) and tocilizumab. in two of them, tocilizumab was used after second dose of atg. both patients present hypotonia with decreased urine output, prompt increase of creatinine level and presence of acute inflammatory parameters crp, beta microglonulin and procalcitonin level, fluid retention and decreased oxygen saturation. in another one patient, these symptoms were connected with pbsc infusion from unrelated donor. in later two patients, we observe almost the same clinical presentation in preengraftment phase. in every of patients infection was ruled out-blood cultures were negative. all these patients were treated with tocilizumab in a single dose of mg/kg. in all patients, we observed prompt response-normalization of clinical state, renal function, oxygen saturation and decrease of inflammatory factors-crp, procalcitonin and beta microglobuline. discussion: crs is a rare complication connected with early phase of allogeneic stem cell transplantation. there were no results of treatment with steroids, reduction of a dose of cyclosporine a according to decreased renal function, but all patient completely/fully recovered after single dose tocilizumab treatment. all our patients were subjected to reduced intensity protocols, what might be a risk factor to develop crs because non complete depletion of the patient origin monocytes/macrophages active population. we also analyzed other factor connected with crs in early peritransplant period finding possible connection with s proinflammatory hla phenotype. it was obvious in the patient one our patients with peri-engraftment phase crs-he was diagnosed previously with rheumatoid arthritis b pos, dr . in three of five, we have found sle predisposition in hla phenotype (drb * /dqb * or drb * / dqb * ), in later one-ra associated hla antigen drb .these patients were analyzed correlating with historical cohort of additional five patients with mortal and another three with very severe early peri-transplant complications and in all we have found the same 'sle or ra hla phenotype'. because small number of analyzed patients and documented high frequency of these haplotype in population, this is still an opened question is proinflammatory/autoimmune hla phenotype connected pathogenically with predisposition to develop severe transplant complications and are we able to treat all these patients with combination of steroids with tocilizumab. further analysis is needed. disclosure of conflict of interest: none. transplant-associated thrombotic microangiopathy (ta-tma) is a severe complication post haematopoietic cell transplantation (hct) leading to high mortality rates. however, outstanding questions regarding its diagnosis, pathophysiology and treatment remain in the literature. recent studies suggest evidence of complement activation, implicating that complement inhibition may be an effective alternative treatment strategy in refractory patients. therefore, we hypothesized that increased complement activation can be detected in ta-tma patients using a functional assay, the modified ham test. we enrolled consecutive patients with ta-tma according to the international working group criteria from january to june . as controls, we studied patients with graft-versushost-disease (gvhd). complement activation was detected using the modified ham test, a cell proliferation assay based on the susceptibility of a pnh-like cell line to complement activated serum. normal human serum was used as a negative control and lipopolysaccharides(lps)-incubated normal serum as a positive control. all samples were tested in triplicates and twice. we studied ta-tma patients transplanted from unrelated / matched ( ) or / mis-matched ( ) donors, identical ( ) and haploidentical ( ) siblings. all patients presented severe acute and/or chronic gvhd. ta-tma presented at median + ( - ) day post-transplant. in the control group, we studied two patients with steroidsensitive grii and two with steroid-refractory griv acute gvhd. we were able to detect significantly increased complement activation in the serum of ta-tma compared to gvhd patients (p = . ). based on previous studies and present controls, percentage of non-viable cells higher than % was considered a positive modified ham test, indicating increased complement activation in four ta-tma patients. regarding treatment outcomes, two patients with a negative modified ham test responded to cyclosporine cessation and steroid administration. plasma infusion with/without plasma exchange was initiated in seven patients. however, only three of them responded to second-line treatment. the modified ham test result was significantly increased in refractory patients (p = . ). the terminal complement inhibitor eculizumab was administered in one refractory patient with a positive modified ham test and renal failure at presentation. despite delayed initiation ( days post ta-tma diagnosis), response was observed after three doses of eculizumab including evidence of reduced hemolysis, schistocytosis and transfusion needs. however, the patient succumbed to complications of end-stage renal disease ( days post ta-tma diagnosis). among ta-tma patients, succumbed at a median + ( - ) day to transplant-associated complications, related to gvhd and infections from multi-resistant pathogens. ta-tma is associated with increased morbidity, mortality and severe complications, including gvhd. unlike gvhd, increased complement activation was observed in a significant portion of ta-tma patients. complement inhibition seems an encouraging therapeutic option in these patients. given the lack of robust functional assays for complement activation, the modified ham test may be useful for early recognition of patients that would benefit from complement inhibition. . this proposal includes, along with the 'classical sos' (cases diagnosed before day + ), the new type 'late onset sos' (cases diagnosed afterwards). new ebmt criteria for severy grading classify cases of sos into four grades (mild, moderate, severe, and very severe). the aim of this retrospective study is to analyze the cases of severe/very severe, both classical and late onset sos, occurred in our unit during the most recent period of time. we studied the last pts, with a minimum follow-up of days, who underwent allo-hsct in our center (november -august ). pts were male and female. median age was years (range: - ). baseline diseases were: acute leukemias ( ), lymphoproliferative disorders ( ), myelodysplastic syndromes ( ), chronic myeloproliferative diseases ( ), multiple myeloma ( ), and bone marrow failures ( ) . donor was unrelated in cases, and related in (including haplo-identical). conditioning regimen was: busulphan-based ( ), melphalan-based ( ), tbi-based ( ) , and others ( ). all patient received prophylactic [p ] s ursodeoxycholic acid. progenitors source was pb in , and bm in . five patients developed severe/very severe sos ( % incidence); were classical (at days + , + and + ), and were late onset (at days + and + ) (see table ). four cases had received conditioning with a busulphan (iv)-based regimen (doses from . to . mg/kg), and one case with tbi plus cyclofosfamide at high doses. all cases presented with right upper quadrant pain, jaundice, ascites, weight gain, hiperbilirrubinemia, and renal function impairment. all but one had increased transaminases. the five cases were treated with defibrotide, in spite of which all of them died. considering that overall day + mortality was %, severe/ very severe sos was the most important cause of death of the series. [p ] although milder forms of sos might resolves within weeks, the most severe forms are still associated with a very high mortality rate. prophylaxis with defibrotide (the drug currently licensed for treatment) for high-risk patients has not been sufficiently studied yet. therefore, a high index of suspicion, early detection and early therapy are the only ways to try to reduce mortality due to sos in the hsct setting. disclosure of conflict of interest: this research has been performed entirely with public financial support. the royal marsden hospital, sutton, uk; anthony nolan research institute, london, uk and university college london, london, uk secondary poor graft function (spgf) complicates up to % allogeneic hcts, and is associated with increased mortality and poor quality of life due to recurrent infections and the need for ongoing blood product support. potential interventions include a second allograft using further conditioning, however many patients with spgf have a reduced performance status and are at an increased risk of complications from this procedure. unconditioned haematopoeitic progenitor cell (hpc) top-ups are associated with a high risk of gvhd if unmanipulated cellular products are used. cd + selection offers an attractive alternative, but incurs a loss of up to % hpcs and is an expensive procedure, unavailable to many centers internationally. alemtuzumab, a monoclonal anti-cd antibody, is routinely used in allogeneic transplant conditioning in the uk to prevent gvhd. we report the results of a retrospective study examining the efficacy of alemtuzumab conditioned hpc top-ups for spgf. data pertaining to patients who had undergone a second infusion of hpcs from their original donor were identified from our hospital-specific promise database. those who met the criteria of spgf defined as ⩾ of hemoglobin × /l without support. patients ( pediatric, adult) who underwent initial allogeneic transplants for malignancy ( ) or bone marrow failure ( ) received an alemtuzumab conditioned hpc top-up for spgf at our center - . the diagnosis of spgf was made at a median . months post allograft (range - ) with trilineage cytopenias in patients and bilineage cytopenias in patients. all patients had received transplants from / ( patients) or / ( patients) matched unrelated donors. the median interval between initial transplant and top-up was days (range - ), and a median cd dose of . × /kg recipient weight (range . - . ) was infused. % patients achieved haematological improvement (hi) at a median days post-top-up (range - ), with the only failure to achieve hi seen in the patient who had received the lowest cd dose ( . × /kg). one patient developed grade i agvhd post top-up but no grade ii-iv agvhd was observed. year os was % and year os % following hpc top-up. deaths occurred due to infection at , and months post top-up, and one due to relapse of a prior non-haematological malignancy. patients had an aplastic or hypocellular bm trephine pre-top up, which was repeated at days post topup in patients, of whom had a normocellular bm trephine, while remained hypocellular. alemtuzumab conditioned hpc top-up appears an effective intervention for spgf with results comparable to those of cd selected top-ups, and therefore represents a feasible alternative. larger studies are needed to exclude complications including viral reactivation and to investigate immune reconstitution following this procedure. disclosure of conflict of interest: none. high dose chemotherapy (hdt) followed by autologous stem cell transplantation (asct) has shown to improve outcome in patients with relapsed/refractory diffuse large b cell lymphoma (dlbcl). in the rituximab era, the benefit of asct has been debatable as prior study (coral study) has shown that patients who received r-chop as induction chemotherapy & responded to salvage chemotherapy had a poorer outcome following asct compared with those who received chop alone. in addition, it remains unclear whether addition of rituximab to standard high dose beam regimen provides any additional benefit. we retrospectively analyzed dlbcl patients receiving high dose beam (n = ) or rituximab +beam (r-beam) (n = ) followed by asct for relapsed/ refractory dlbcl since . all patients who received chop (n = ) ± rituximab (n = ) as first line therapy and who received ⩽ lines of salvage chemotherapy before asct were analyzed. rituximab was given at the dose of mg/m on day + and + of asct. twenty-two ( %) patients in beam group and all the patients ( %) in r-beam group received rituximab-based salvage chemotherapy prior to asct. the year overall survival (os) was % and event-free survival (os) was % for the whole cohort. r-chop induced patients did not fare any worst after asct than chop induced patients ( year os vs %; p = . ). there was a trend towards better survival in patients with pre-transplant disease free interval (dfi) months compared to those with dfi /μl) time was days and days, respectively. median platelet recovery ( /μl) time was days and days, respectively (p = . ). ten year os ( % r-beam vs % s beam, p = . ) and efs ( % r-beam vs % beam, p = . ) were also comparable between both groups. hdt with beam and asct remains beneficial for patients with relapsed/ refractory dlbcl. it should be offered to all patients who respond to salvage chemotherapy with the expectation that they fare no worse than patients who do not receive rituximab in the induction chemotherapy. addition of rituximab following the standard beam for hdt and asct does not compromise haematopoietic recovery, but does not result in improved outcome in our study. prior use of rituximab during first-line or salvage therapy in most of the patients of r-beam group might have negated the beneficial effect of r-beam over beam. ( ) . in this study, we aimed to develop a cns targeted chemotherapy regimen, which has lower toxicity and higher complete remission rates, in combination therapy. eight patients with secondary cns lymphoma (scnsl) and two with primary cns lymphoma (pcnsl), followed between the years and , were included in the study, retrospectively. the patients were histologically diagnosed with biopsy and underwent autologous stem cell transplantation (apkht). all patients were treated with r-idaram/ rt (radiotherapy)/subsequently autologous stem cell transplantation (apsct) with r-beam protocol. the r-idaram regime consists of the following substances: rituximab mg/m , cc/h infusion, day ; cytosine arabinoside . gr/m i.v., h infusion, days and ; dexamethasone mg, h infusion, days , and ; idarubicin mg/m i.v., min infusion, days and ; methotrexate gr/m ( gr/m at years old-patients), h infusion, day ; and cytosine arabinoside mg plus methotrexate mg, intrathecally, days and . the patients included seven males and three females. the median age was years (range: - ). six scnsl patients were diagnosed in the application and two of them were diagnosed during r-chop chemotherapy (ct) protocol. five patients ( %) were stage ivb, and the others ( %) were stage iiib at diagnosis. after two or three chemotherapy cycles, patients were mobilized with growth factor support and median . cells per kg (range: - ) stem cells were collected. then, at a dose of - cgy cranial rt was administered for days. after the third cycle of r/idaram, the state of remission was evaluated by cranial mri and lumbar puncture (lp). all patients achieved complete remission. neutrophil engraftment occurred at a median of days (range: - ) and platelet engraftment occurred at a median days (range: - ). after apkht, three patients relapsed and died at the fourth, ninth, and thirteenth months. grade i-ii manageable neurological toxicity occurred in two patients. the median follow-up time was (range: - ) months. the five-year overallsurvival (os) was %. serious signs of infection were not observed in patients during transplantation. in pcnsl and scnsl, a standard treatment regimen has not yet been found. apsct with r-beam following modified r/idaram/rt is a curative and applicable therapeutic regimen with low toxicity, which can provide high rates of long-term survival and disease-free survival. despite the advent of novel therapies, autologous hematopoietic stem cell transplantation (ahsct) following melphalan (m)-based conditioning remains the standard of care for patients with multiple myeloma who are eligible. still, the majority of patients experience disease progression and ultimately succumb to their disease. we hypothesize that integrating novel agents in the conditioning is feasible and safe and may increase complete remission rates and overall survival. we completed a phase i, dose escalation study of carfilzomib (c) added to a backbone of bendamustine (b) and melphalan. all patients received a fixed dose ( mg/m ) of c on days (d) − , − , − , − , − and − . in addition, patients were conditioned as described in table . due to dose-limiting toxicity in cohort , the study was amended after the first patients. subsequently, the dose of m was reduced to mg/m and the d + dose of c was omitted, per oversight of a data safety monitoring board. fifteen patients were enrolled, males and females. median age was years ( - ). performance status was ⩾ % (kps) in all patients. per the international staging system (iss), patients had stage i disease, had stage ii, had stage iii, and had unknown staging. three patients had high-risk cytogenetics: with t( ; ) and with deletion p. four patients had undergone a prior ahsct. disease status at enrollment was stable disease (sd) (n = ), partial response (pr) (n = ), or very good partial response (vgpr) (n = ). median cd + cell dose infused was . × /kg ( . − . × ). median follow-up was . months ( . - . ). all fifteen patients are evaluable s for engraftment. median time to neutrophil engraftment was d ( - ). one patient died before achieving platelet engraftment. for the remaining patients, median time to platelet engraftment was d ( - ) . non-hematologic toxicities included grade acute mucositis (n = ), lower gi complications (n = ), electrolyte disturbances (n = ), transaminase elevation (n = ) renal insufficiency (n = ), atrial fibrillation (n = ), hypoxia (n = ), prolongation of the qtc interval (n = ), and grade acute sepsis (n = ), including death (cohort ) on d + . eight patients went on to receive maintenance therapy: with bortezomib, with lenalidomide, and with lenalidomide, dexamethasone, and c. posttransplant disease status was assessed per protocol by spep, spif, serum free light chains, and light chain ratio. twelve patients were evaluable on d + . two patients had sd, had vgpr, and had complete response (cr). eight patients were evaluable on d + . two patients had progressive disease, had pr, had vgpr, and had cr. the combination of cbm prior to ahsct appears feasible, with manageable toxicities, at the doses described in cohort b. a prolonged follow-up and a phase ii study are warranted to determine response rates and long-term outcomes. disclosure of conflict of interest: none. beam (carmustine, etoposide, cytarabine, melphalan) is the most frequently used high-dose chemotherapy regimen for patients with lymphoma referred for autologous hematopoietic cell transplantation (autohct). in recent years a novel conditioning protocol containing bendamustine instead of carmustine (beeam) has been proposed in order to potentially increase the efficacy. so far, however data on its safety are limited. the aim of this study was to retrospectively compare the safety profile of beam and beeam based on single center experience. consecutive patients with lymphoma treated with beam and patients treated with beeam between year and were included in the analysis. the median age was ( - ) years and ( - ) years, respectively (p = ns). clinical characteristics of both groups were comparable. patients with hodgkin's lymphoma constituted % in the beam group and % in the beeam. among those with non-hodgkin lymphoma the diagnosis of dlbcl predominated. beam treatment consisted of carmustine mg/m on day − , etoposide mg/m /d on days − to − , cytarabine mg/m /d on days − to − , and melphalan mg/m on day − . in the beeam regimen carmustine was substituted by bendamustine administered on days − , − at the total dose of mg/m i.v. peripheral blood was used as a source of stem cells. cd + cell dose was . ( . - . ) × /kg in the beam group and . ( − . ) × /kg in the beeam group (p = ns). time to engraftment and the rates of adverse events up to day + after autohct were the study endpoints. all patients engrafted in both study groups. median time to neutrophil . × recovery was ( - ) days after beam and ( - ) days after beeam (p = . ). median time to achieve platelet count × was ( - ) days and ( - ) days, respectively (p = . ). two patients died without progression before day + in the beam group, both due to bacterial infections. no early deaths were reported in the beeam group. the rates of grade or adverse events were comparable (see: table ). administration of bendamustine instead of carmustin as part of conditioning does not affect engraftment as well as toxicity profile of the regimen. therefore beeam may be safely used in patients with lymphoma undergoing autohct. its efficacy requires evaluation in prospective studies focused on homogenous patient populations. [p ] disclosure of conflict of interest: none. the baltimore group reported a low dose tbi-based nonmyeloablative conditioning regimen followed by t cell replete bone marrow, with post-transplantation cyclophosphamide (pt-cy) to control gvhd and graft rejection. based on the fact that in our facility conventional low dose tbi was not available, we wanted to explore whether tmi/tli could be a potential substitute the aims of our study was to explore if tmi/tli can be considered an effective substitute of tbi in terms of os, pfs and nrm. retrospective analysis was applied in cases of haploidentical hsct from april to october . all patients underwent baltimore conditioning associating fludarabine ( mg/m /day) day − to − , cy ( . mg/kg/day) on days − and − , and tbi gy in patients and tmi/tli gy in patient at day − . unmanipulated bone marrow graft was infused at day . postgrafting immunosuppression consisted of cy ( mg/kg/day) on day + and + , and mycophenolate mofetil for days, and tacrolimus or cyclosporine. no differences between the two groups was observed in term of age, gender diagnosis, disease status and donor type. % of patients engrafted in both arm ( / and / ). in tbi cohort vs tmi/tli cohort, the median time to anc /μl and platelet recovery /μl was not different ( and days vs and . days, p = . and . , respectively). in all tmi/tli evaluable patients, full chimerism was observed at days + . after a median followup of months in tmi/tli cohort and months in tbi arm, -year nrm was . % and . % (p = . ), respectively. the years os and pfs were not statistically different in the two groups % vs . %, p = . and . % vs . %, p . , respectively). the -year relapse incidence was % in tmi/tli group and . % in tbi group, p = . . no difference in incidence of both agvhd and cgvhd was observed between the two groups. this retrospective analysis suggests that tmi/ tli could be considered an effective substitute of low dose tbi, with a sufficient degree of immunesuppression of recipient, allowing engraftment and full chimerism. the gvhd both acute and chronic as well as the -y nrm were not different. disclosure of conflict of interest: none. comparison of the beeam conditioning regimen and the beam conditioning regimen in the autologous transplantation for hl and nhl s lozenov , p ganeva , y petrov , g arnaudov and g mihaylov the beam has established itself as a standard of care conditioning regimen in the autologous lymphoma hsct setting for most transplant centres in europe. yet however various other regimens are being compared with it in order to achieved better safety profile, better os and dfs, in order to improve results with chemoresistant and unfavourable patients. one such regimen is beeam (bendamustine, etoposide, cytarabine, melphalan).we aimed to compare the efficacy of the beam and beeam conditioning regimens and to compare their myelotoxicity profile. we evaluated retrospectively adult patients (mean age . with sd . ), receiving auto-hsct at the national specialized hospital for active treatment of hematological diseases in sofia, bulgaria for relapsed/refractory hl or nhl (of them mh - , dlbcl - , pmbcl - , fl - , lbl - , ptcl-nos - , aitl - , alcl - , mcl - , mzl - ) for the period from . . to . . with a follow-up of patients up to . . . ninety-two of the patients received the beam (as previously described -bcnu mg/m i.v. day − , etoposide mg/m i.v. days − to − , cytarabine mg/m i.v. days − to − , and melphalan mg/m i.v. day − ) regimen and received beeam regimen (bendamustine on days − and − ( mg/m ); cytarabine, mg/m intravenously daily, from day − to day − ; etoposide, mg/m intravenously daily, from day − to day − ; and melphalan, mg/m intravenously on day − ). the overall survival at the second and third years of follow-up (os- , os- ) and dfs at the third year, the cr rates and the average time periods to hematological recovery, were compared. the os at and years, respectively, was . % and . %, for beeam and . % and % for beam, the dfs at years was . % for beeam and . % for beam, provided that the differences did not have statistical significance (p . for os and p . for the dfs). the cr rate was . % in the beeam group versus % in the beam group. from the patients who received autologous hsct in stable disease or progression pre-transplant status (chemoresistnat patients), . % of the patients receiving beeam achieved cr at the first post-transplant evaluation versus . % respectively for the beam group. the mean time to hematological recovery for neutrophils was . ± . days (beeam) versus . ± . days (beam) and . ± . days (beeam) versus . ± . days (beam) for platelets. beeam appears to be a non-inferior alternative conditioning regimen to the standard beam, it shows a trend towards higher myelotoxicity, but also a trend towards better response rates in chemoresistant patients. [p ] disclosure of conflict of interest: none. autologous hematopoietic stem cell transplantation (asct) is widely used as a consolidation therapy in aggressive non-hodgkin's lymphoma (nhl) and recurrent or refractory classic hodgkin's lymphoma (hl). in mexico, the use of carmustine (bcnu) in the conditioning regimen of these patients is limited due to the lack of access to the drug and its high costs. this study aims to compare results in terms of toxicity, disease-free and overall survival between a group of patients treated with the standard regimen beam and another group treated with a scheme in which carmustine was replaced by cisplatin (peam regimen). a comparison of two groups with lymphoma was performed and the clinical aspects of cisplatin mg/m d . the characteristics were well balanced between the two groups. the mean time for neutrophil grafting ( per mm ) was significantly slower with beam than with peam ( vs days, p = . ), hospitalization time was longer with beam compared to peam ( vs , p = . ). on the other hand, proportion of patients who require red blood cell s transfusion was significantly higher in beam group ( %) versus peam group ( %) (po . ), but total amount of platelet transfusion did not differ between groups. about the toxicity, beam patients had significantly more frequent incidence and severity of nausea/vomiting ( % vs . %) and diarrhea ( . % vs %) compared to peam (p o . ). no significantly differences were observed in incidence of mucositis (p = . ). at the moment of the analyses, % of patient of the peam group were in complete response versus % of the patients treated with beam, but it did not represent a significant difference. disease-free survival and -year overall survival in the peam vs beam scheme were similar with % vs % (p = . ) and % vs % (p = . ) respectively but with less toxicity using the peam scheme. peam regiment is not inferior scheme compared with beam, because it shows similar outcomes in disease-free survival and overall survival. additionally, peam is a well-tolerated regime and beam scheme was associated with greater gastrointestinal toxicity such as nausea, vomiting and diarrhea, also greater hematology toxicity such as more requirement of red blood cell transfusion. [p ] disclosure of conflict of interest: none. cumulative busulfan exposure is associated with relapse following busulfan and cyclophosphamide myeloablative allogeneic stem cell transplantation for acute myeloid leukaemia e wong, d kliman , m chau , j szer , c nath , p shaw , d ritchie , d gottlieb and a bajel westmead hospital, new south wales, australia and royal melbourne hospital, victoria, australia the optimal busulfan exposure to reduce disease relapse in adult patients with acute myeloid leukaemia (aml) undergoing busulfan/cyclophosphamide myeloablative allogeneic stem cell transplant (allosct) is poorly defined. we retrospectively analysed busulphan pharmacokinetics (pk) and outcomes of patients who underwent busulfan/cyclophosphamide conditioned allosct for aml from to . busulfan was administered intravenously over days ( . mg/ kg/d for days followed by . mg/kg for days). peripheral blood was obtained for busulfan pk after the first dose. subsequent doses of busulfan were decreased if daily busulfan exposure (area under the curve; auc) was anticipated to exceed μm per min/day. cyclophosphamide was dosed at mg/kg. the primary outcome was the cumulative incidence of relapse (cir) accounting for non-relapse mortality (nrm) as a competing risk. independent variables analysed included age, sex, cytogenetic risk group, disease risk index (dri), donor type, stem cell source, t-cell depletion, and cumulative busulfan auc (cumauc) calculated as previously described. (figure ) . t-cell depletion was also associated with increased cir (hr . ; p = . ). patient age, sex, cytogenetic risk, dri and graft type were not significantly associated with cir. on multivariate analysis, cumauc μm per min remained significantly and independently associated with lower cir (hr . ; p = . ). cumauc was not associated with nrm, rfs, os, or the incidence of acute or chronic gvhd. figure . cumulative incidence of relapse in patients stratified by total busulfan exposure. [p ] cumulative busulfan exposure μm per min is independently associated with reduced relapse following busulfan/cyclophosphamide allosct for adults with aml. these findings support further evaluation of the optimal busulfan exposure to reduce aml relapse in a prospective clinical trial, whereby patients could be randomised to target cumauc μmol per min versus standard practice. hematopoietic stem cell transplant with busulfan and cyclophosphamide (bucy) based conditioning has a relatively high incidence of liver toxicity and sinusoidal obstruction syndrome (sos). busulfan and cyclophosphamide metabolites share the same glutathione conjugation in the liver metabolism. a small number of studies addressed different sequence of both drugs bucy vs cybu during conditioning. differences in liver toxicity, sos, transplant related mortality (trm), relapse incidence (ri) and overall survival (os) were reported favoring cybu conditioning. we decided to address the above issues at the umc ljubljana, slovenia. this was a retrospective study following patients with myeloid malignancies (aml, mds, mpn) with bucy (n = ) and cybu (n = ) conditioning through a three year period in a single institution. primary endpoint was detecting difference in liver toxicity by measuring levels of liver enzymes. secondary endpoints were incidence of sos, difference in trm, ri and os. patients characteristics between groups at the time of the transplant did not differ significantly. we observed significantly higher liver toxicity through elevated bilirubin and alt in the bucy . % than cybu . % patient group (picture ). the highest probability of liver toxicity was around d in the bucy group and in the second week after the transplant in the cybu group. the incidence of sos, trm and ri were comparable between the groups. there was no difference in os between the patient groups during the -month follow-up. bucy conditioning for hematopoietic stem cell transplant causes higher incidence of liver toxicity compared to cybu conditioning. there is no difference in sos frequency, trm, ri and os between bucy and cybu conditioning. prospective controlled comparison would be needed for further study of the subject. disclosure of conflict of interest: none. early monocyte recovery is associated with better overall survival after busulfan containing myeloablative conditioning allogeneic hematopoietic cell transplantation in patients with acute myeloid leukemia a lojko-dankowska the outcomes of allogeneic hematopoietic cell transplantation (allohct) in acute myeloid leukemia (aml) depend on different patient-, disease-and transplant related factors, including the dose and combination of agents used for conditioning. the aim of the study was to analyze the outcomes of allohct in patients with intermediate or high risk aml according to disease risk index (dri) who received myeloablative conditioning consisted of intravenous busulfan ( . - . mg/kg) combined with cyclophosphamide ( mg/ kg) or fludarabine ( mg/m ) between and in our institution. the published data indicate that the combination of busulfan (bu) and fludarabine (flu) seems to have more favorable toxicity profile than combination of bu and cyclophosphamide (cy), so bucy regimen has been substituted with buflu as the myeloablative conditioning for aml patients in our institution practice since . we evaluated the influence of type of regimen on transplant outcomes along with the impact of other potential prognostic factors, including age of patient, dri, donor type, hla and gender mismatches, stem cells source, and lymphocyte and monocyte recovery. the study group consisted of aml patients, median age years (range: - ), classified as intermediate (n = ) or high (n = ) risk according to the dri, who were conditioned with bucy (n = ) or buflu (n = ) followed by allohct from hla identical sibling (n = ) or - / matched unrelated donor (n = ). the stem cell were collected from peripheral blood (n = ) or bone marrow (n = ). gvhd prophylaxis consisted of calcineurin inhibitor combined with mtx plus atg in allohct from unrelated donors. engraftment was observed in all patients. the median time to neutrophil count ( . g/l) and platelet count ( g/l) recovery was shorter after buflu in comparison with bucy ( days vs days; p = . and days vs days; p o . ), however peripheral blood stem cells were used more often after buflu regimen than after bucy ( % vs %, p /mm on+ day after transplant ( -year os % vs %, p = . ) and intermediate vs high dri ( -year os % vs %, p = . ). in multivariate analysis higher amc after allohct remained the only independent favorable prognostic factor for os (rr . ( % ci . - . ), p = . ). our results suggest that early monocyte recovery after myeloablative bu containing conditioning allohct is significant favorable predictor of outcome. in our experience both bucy and buflu myeloablative regimens result in similar long-term survival after allohct in aml patients. [p ] disclosure of conflict of interest: none. the use of t-cell depletion as part of the conditioning protocol has the potential to improve the tolerability of allogeneic stem cell transplantation (hsct) through the reduction in graft versus host disease (gvhd). despite the wide spread adoption of this practice in many parts of the uk and europe, definitive recommendations regarding the most appropriate dose remain elusive. previous experience by our group with mg of alemtuzumab combined with fludarabine and busulfan based conditioning demonstrated good long-term outcomes with low rates of gvhd. however, due to concerns of high relapse risk especially in patients with high-risk myelodsypastic syndrome and acute myeloid leukaemia, we instituted a policy change in to reduce the dose of alemtuzumab in the conditioning protocol from a total of - mg. we conducted a retrospective analysis of all consecutive patients undergoing reduced intensity unrelated allogenic stem cell transplantation with fludarabine ( mg/m ), busulfan ( . mg/kg iv or . mg/kg iv) and alemtuzumab (fb c or fb c, respectively) conditioning for neoplastic myeloid disorders between and . patients were subsequently analysed in two cohorts; those receiving mg of alemtuzumab (n = ) and those receiving mg of alemtuzumab (n = ). apart from a decreased proportion of females in the mg alemtuzumab group, the cohort was balanced across the different dose levels ( table ). the longterm overall survival (os) of the entire cohort was good with a year os of %. no significant differences in overall outcomes across the two groups were observed with a year os of % in the mg group vs % in the mg group (p = . ). cumulative incidence of relapse (cir) and nonrelapse mortality (nrm) was % and % and % and % in the mg and mg groups, respectively. interestingly, age had a significant effect on nrm in the mg ( % age o , % age - and % age p = . ), but not in the mg group ( % age o , % age - and % age p = . ). the effect on relapse rate was not significant in either group (p = . and p = . , respectively). this retrospective analysis did not demonstrate an overall improvement in transplant outcomes with dose de-escalation of alemtuzumab from to mg. in particular, we did not see the anticipated improvement in relapse rate in this cohort. notably older patients seem to tolerate the mg dose better due to the lower nrm. prospective trials with accompanying translational work are required to determine the optimal dosing and schedule for this group of patients. disclosure of conflict of interest: none. bendamustine was given at mg/m /d for the first pts then mg/m /d for the subsequent pts and finally at mg/m /d for the remaining pts ( pts). among the beam group, % had non-hodgkin's lymphoma (nhl) and % hodgkin's lymphoma (hl) compared to % and %, respectively, in the beeam group (p = . ). hhv- detection was performed by pcr for symptomatic pts (fever, rash or prolonged cytopenia). patients were housed in single bedrooms with air filtration and received the same supportive care. median age was ( - ) and ( - ) in the beam and beeam groups respectively and median of previous chemotherapy regimens was (range: - ). fifty two out of patients were male ( / in the beam group and / in the beeam group). pts were in cr ( . % vs . %) or pr ( . % vs . %) at time of transplant. there was no difference in terms of hematologic recovery (median = days (range: - )), blood and platelets transfusion, mucositis toxicity. there was no statistical difference in the incidence of acute renal failure when comparing the two groups. however, there was a very striking difference when considering the highest dose of bendamustine when compared as well to the two others doses of bendamustine (po . ) as to the beam group (p = . ). additionally, we also observed a high incidence of symptomatic hhv- infections ( . % vs . %, p o . ), digestive toxicity ( . % vs %, p = . ) and a longer hospitalization duration ( days (range: - ) vs days (range: - ), p = . ) for patients in the beeam group overall. with a median follow up of . and . months for beam and beeam respectively, overall survival ( % vs %), transplant related mortality ( % vs %) and event free survival ( % vs %) were comparable. overall, beeam regimen was associated with longer duration of hospitalization, higher rate of digestive toxicity and increased risk of symptomatic hhv- infection as compared to the beam regimen. in addition, higher doses of bendamustine ( mg/m /d for two consecutive days) were associated with unacceptable high rate of acute renal toxicity. with a still short follow-up, the absence of benefit on disease control together with higher short term toxicity does not allow to recommend the use of beam instead of classical beam. should it be used, we suggest that pts should be carefully monitored for renal toxicity and for hhv- infection in case of symptoms. disclosure of conflict of interest: none. high-dose treosulfan and melphalan for consolidation therapy in high-risk ewing sarcoma me abate, a paioli, a longhi, m cesari, e palmerini and s ferrari musculoskeletal department, rizzoli orthopaedic institutes, bologna, italy common toxicities observed after high dose chemotherapy with busulfan and melphalan for high risk ewing sarcoma (es) are generally well managed by current supportive care but some patients can develop severe complications. treosulfan is an alkylating agent that has recently been used as a substitute of busulfan to prevent potential serious complications related to busulfan. medical records of es patients undergoing autologous peripheral blood stem cell (apbsc) transplantation after intravenous treosulfan (treo) and melphalan (mel) from / / to / / were analyzed with regard to toxicity and outcome. patients were included into the study if they were eligible for the protocols activated in our institution for es and presented reasons that did predict potential complications related to busulfan, such as previous radiotherapy on axial skeleton/pelvis or coexistence of high risk of epilepsy. as consolidation treatment patients received intravenous treo g/m over days and mel mg/sqm with support of apbsc transplant and use of granulocyte colony stimulating factor. in those patients with lung metastases total lung irradiation was performed at least months after treomel. frequency of toxicity for treomel was recorded with at least months of follow-up and was evaluated according to nci ctg common toxicity criteria. the median age at diagnosis of patients receiving treomel was years (range - years), males and females. patients had localized disease at diagnosis with poor radiological or histological response to standard chemotherapy; one patient had lung metastases at diagnosis and one patient had relapsed disease with lung metastases. before receiving treomel the primitive tumour underwent radiation therapy in cases ( pelvis, cervical vertebra, sacrum), surgical resection in one case(tibia) and surgical resection plus radiation therapy in one case (fibula). patients showed eeg abnormalities at high risk of developing epilepsy. the median number of infused cd + cells was . × /kg (range . - . ). febrile neutropenia occurred in / patients and lasted one day in patients and days in patients. median time to granulocyte engraftment was days (range - days); median time to platelet engraftment was days (range - days). only one patient needed red blood cells transfusions; patients needed platelet transfusion and patients needed platelet transfusions. none developed grade - stomatitis or grade - [p ] hematuria or grade - liver toxicity. surprisingly, a patient became pregnant after year and months from transplantation. with a median follow-up of months (range - months) patients are alive in complete remission, one patient is alive with relapsed disease and one patient died for disease progression. these results, related to a limited cohort of patients, confirm the lower toxicity observed for treosulfan with respect to busulfan. although more data are needed to clarify the role of treosulfan in es, the impact of potential severe complications observed with busulfan, including infertility, should suggest its replacement with treosulfan in selected cases. disclosure of conflict of interest: none. immunoadsorption procedures prior to haploidentical allogeneic pbsct could prevent graft failure in patients with hematological malignancies displaying anti-donorspecific hla antibodies donor-specific anti-hla antibodies (dsa) have been shown to be associated with a high risk of rejection in solid organ transplantation and with graft failure (gf) in allogeneic hematopoietic stem cell transplantation. a combination of anti-cd (rituximab), plasma exchange (pe), and ivig in patients with additional buffy coat infusion in among them prevented graft loss in all patients that became c q negative before sct. we addressed the question whether immunoadsorption in combination with rituximab can also be applied in patients with dsa to prevent graft failure in haploidentical pbsct. four patients with acute myelocytic leukemia or myeloma in second complete remission were enrolled. the presence of dsa was determined by luminex at pre bmt checking. immunoadsorption was performed with polyclonal sheep anti-human igg adsorbers (miltenyi biotec gmbh, germany) on life apheresis system. in addition all patients received rituximab mg/kg bw in a single dose. patients were conditioned with a reduced intensity regimen comprising tbi gy, cyclophosphamide mg/kg, and fludarabin mg/m . all patients received cyclophosphamide post bmt (ptcy) mg/kg. non-t-cell depleted pbsct were transfused in a sequential manner in doses each. the data of the patients and treatments is summarized in table . two patients had a normal hematopoietic reconstitution and are alive at + and + months post-transplantation, one with hepatic gvhd; chimerism was % in peripheral blood on last follow up. one patient died following a graft failure. by a combination of rituximab and repeated immunoadsorption prior to allogeneic pbsct the titer of dsa could be lowered sufficiently to enable engraftment. ia turned out to be a safe procedure without relevant clinical side effects. hematopoietic reconstitution was in the normal range in of evaluable patients. disclosure of conflict of interest: none. allogeneic hematopoietic stem cell transplantation (hsct) is the only curative option for patients with beta thalassemia major. however, the availability of hla-matched related donor remains the main obstacle for allogenic hsct. although, a few studies have been reported, experience with hla matched unrelated donors is limited. we present the result of children with beta thalassemia major who received allogeneic hsct from hlamatched unrelated donors with using a novel conditioning regimen. we retrospectively assessed unrelated hsct in children with beta thalassemia major. all patients received busulphan (bu) based myeloablative conditioning regimen. busulphan was used according to weight adjusted doses. in addition, all patients received fludarabine mg/m in days, cylophosphamide mg/kg in days, thiotepa mg/kg in one day and atg mg/kg in days. cyclosporin-a and mtx were used for graft versus host disease (gvhd) prophylaxis. donor chimerism was evaluated in the peripheral blood on days + , + and + . the median age of the patients was . years (range month- year). two of the patients were grouped in class i and rest of them were class ii. the median serum ferritin level was . ng/ml (range, - ). all of s the donors were matched / with high-resolution hla typing in gvhd direction but three of them / with graft failure direction. twenty-three of them received bm (median tnc: . x /kg) and pbsc (median mnc: . x /kg) with median cd + cell number . x /kg. the median neutrophil and platelet engraftment days were and days in pbsc and and days in bm group, respectively. grade i-iv acute gvhd was observed in patients ( %) and only one experienced limited chronic gvhd with only skin involvement. mild to moderate vod was seen in patients ( %) and treated with defibrotide successfully. all patients except one are alive with full donor chimerism (between - %) with a median months (range - months) follow-up. one patient died because of cmv pneumonia. these data show that the results of hsct from unrelated donors in selected low risk thalassemia patients may be comparable to hsct of matched sibling donors. however, it needs further studies with long term follow up and larger study population. disclosure of conflict of interest: none. table . data about cytogenetic risk of group patients were available only in individuals. differences between groups were analyzed by t-student and chi square tests. survival was analyzed by kaplan-meier method and differences in survival between groups were evaluated by log rank test. no differences were found between groups regarding gender, sc source, disease status at sct, type of donor and number of cd + cells infused. patients in group were significantly older (median age for groups and : vs , p = . ). gvhd prophylaxis protocols included atg in a higher frequency in group . no differences between groups and were observed in neutrophils recovery (median days to anc /μl: vs respectively, p = . ) and platelets recovery (median days to platelets /μl: vs respectively, p = . ). patients in group required more red cell transfusions (median packed rbc: vs , p = . ). no differences were observed regarding platelets transfusion requirements or length of hospitalization. post-sct os was significantly better in group ( years-os group : %; group : %; p = . ) (figure ). there were no significantly differences between groups regarding frequency of mucositis, diffuse alveolar haemorrage, sepsis, acute and chronic gvhd. vod was more frequent in group ( / vs / , p = . ). trm mortality was higher in group ( / vs / ), being this difference no statistically significant (p = . ). as it was reported by others, the use of fludarabine-based conditioning regimen was associated with a significantly better post-sct os and a reduced frequency of vod in aml patients. reduction in trm and differences in the frequency of described complications are not statistically significant probably due to the small size of this sample. since march , given the limited availability of melphalan, we administer the beac regimen (carmustine, etoposide, cytarabine, and cyclophosphamide), instead of the gold standard conditioning regimen beam, followed by autologous haematopoietic cell transplantation (ahct) in relapsed or refractory hodgkin (hl) and non-hodgkin (nhl) lymphoma patients. the primary goal of this analysis was to assess the immediate related toxicity of this alternative regimen. we used beac (carmustine mg/m , etoposide mg/m , cytarabine mg/m , and cyclophosphamide mg/kg) in consecutive lymphoma patients ( hl, nhl) who underwent ahct for relapsed or refractory disease. the median age of the patients was . years ( - ). they all received peripheral stem cell grafts with a median cd + cell dose of . × /kg cells ( . - . ). disease status post salvage treatment (at ahct) was complete remission (cr) in , partial remission (pr) in and progressive disease in . the disease was chemosensitive to salvage therapy in / patients. median follow up was days ( - ). toxicity was assessed according to the who toxicity scale grading. all patients engrafted successfully. median time for engraftment was day + (d+ ) for neutrophils ( /mm ) and d+ for platelets ( /mm , without transfusion within the previous days). patients were hospitalized for a median of days ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) . no treatment-related mortality occurred. two patients died due to disease progression (both nhl patients, on d+ and d+ ). toxicity assessment until d+ is presented in table : moreover, no hemorrhagic cystitis or macroscopic hematuria, and no cardiac events were encountered. febrile neutropenia was recorded in and bacteremia in patients ( gram+, gram-, / related to central venous catheter), with fever ≤ grade in all cases. during d+ - two patients presented fever of unknown origin, and patients had upper or lower respiratory infections, with no other adverse events being recorded. in terms of disease best response within months post ahct ( / patients evaluated), patients achieved or sustained cr, pr ( of these patients eventually died due to disease progression), relapsed and succumbed due to disease progression (no response). according to our preliminary results, the early toxicity profile of beac is very low, the regimen is easily tolerable for the patients, and without any treatment-related mortality. its use as an alternative conditioning regimen in ahct for lymphoma patients seems feasible. further investigation including more patients and comparative analysis to other conditioning regimens are warranted for reliable conclusions on the toxicity and efficacy of beac. disclosure of conflict of interest: none. veino-occlusive disease (vod) is a potentially fatal adverse event caused by intravenous (iv) busulfan used in bone marrow transplantation (bmt) conditioning. the objective of this study was to identify determinants of vod in children treated by iv busulfan. this was a retrospective analysis of data collected in children from two bmt centers over years. vod was diagnosed according to modified seattle criteria. individual pharmacokinetic data, including busulfan area under the concentration-time curve (auc) and maximal concentration (cmax) were estimated in all children by using a validated bayesian approach. we examined the relationships between the occurrence of vod and available data in a learning (n = patients) and validation set (n = patients) obtained by random splitting. logistic regression was used as a continuous statistical model. in addition, we used classification and regression tree (cart) analysis, a machine learning and binary partitioning technique, to identify determinants of vod and their optimal cut-off values. the predictive performance of variables within both models was assessed by these results are compared with historical data from our service using beam as conditioning followed by auto-sct in lymphoma patients. nine patients were enrolled to receive neam: mitoxantrone mg/m day - to - , etoposide mg/m every hours and cytarabine mg/m every hours day - to - , and melphalan mg/m day - , followed by auto-sct. the median age was years ( - ); five non-hodgkin lymphomas (nhl) and four hodgkin lymphomas (lh). six patients were in partial remission (pr), two in complete remission (cr), and one with progressive disease at time of auto-sct. neam patients were compared with a historical control group of patients receiving beam regimen (n = ). differences between groups were analyzed by t-student and χ -tests. median cd + cells infused in neam and beam groups was . × ⁶/kg ( . - . ) and . × ⁶/ kg ( . - . ), respectively (p = . ). the median time to neutrophil recovery ( /μl) was days ( - ) and days ( - ) (p = . ) and median time for platelets recovery ( /μl) was and days ( - ) and days ( - ) (p = . ) respectively, for neam and beam patients. median duration of hospitalization was days ( - ) with neam and days ( - ) with beam (p = . ). among neam patients, % had one or more febrile episodes during neutropenia. no case of grade iii or iv mucositis was described. there was no transplant-related mortality (trm: %) associated with the use of neam regimen. at the present, all neam patients are alive, two of them in relapse ( %). due the difficulties in obtaining carmustine in our region, neam can be considered as a feasible alternative to beam. however, despite the sample was small enough to draw conclusions, we find that neam presents prolonged aplasia of significant value, we are currently exploring conditioning regimens followed by auto-sct in hodgkin's and non-hodgkin's lymphomas based on bendamustine, etoposide, cytarabine and melphalan. disclosure of conflict of interest: none. at present, decision-making about conditioning regimens for allogeneic hsct is based on patient's and donor's features, and disease characteristics. during the last years, terms as 'myeloablative/non-myeloablative/reduced-intensity' have been frequently employed in a confusing and unequal way among the different centers. knowing the expected intensity and myeloablative effect from each regimen is very useful and constitutes the aim of this analysis. we have analysed the severe neutropenia (anc o per mcl), and thrombocytopenia durations (platelets o per mcl), the need for platelet concentrates transfusion and the duration of the inpatient period of the allo-hsct carried out during the last four years in our centre. these data are reported according to the conditioning regimen used and to the type of transplant performed. then, they are compared among them in order to stablish intensity ranks. results: population characteristics are described in table : conventional intensive regimens (bu-cy , cy-bu , tbi-cy) reported more days of severe neutropenia and greater need of platelet concentrates transfusion. the regimens with less days of severe neutropenia, less need of platelet concentrates transfusion and fewer days of admission were flu-bu and flu-bu . allotransplants carried out with stem cells from bm presented more days of severe neutropenia and longer hospital stay. similar platelet transfusion need was reported. haplo-identical allotransplants reported more days of severe thrombocytopenia, but were not asociated to longer neutropenia or longer hospital stay than the others. these data are described in detail in table . flu-bu: the number ( , or ) expresses the doses of busulphan administered at . mg/kg/day. pc: platelet concretates data is expressed in medians. within the analysed conditioning regimens, an intensity rank is stablished regarding the myelosupression induced (in descending order): conventional intensive regimens (cy-bu , bu-cy , tbi-cy), flu-mel, flu-bu , flu-bu and flu-bu . all of them induced severe neutropenia and thrombocytopenia, and for that reason they must be considered myeloablative regimens. disclosure of conflict of interest: none. myeloablative allogeneic stem cell transplant for aml and mds: the impact of advanced age in the outcome m sánchez-escamilla* , , s garcía-Ávila , l yáñez san segundo , , ma bermudez rodriguez , , mm colorado araujo , , a casado diez , m celis alvarez , a cabero martinez , c fernandez martinez , a insunza garminde , , c richard espiga , and e conde garcía , allogeneic hematopoietic stem cell transplantation (allo-hsct) is the only curative option in high risk myeloid hematological malignancies. myeloablative conditioning (mac) regimen has been proven to be effective in the control of high risk diseases in advanced age patients. objective: the aim of this study was to analyze the efficacy of myeloablative allo-hsct in two cohorts of patients considering their age at transplant. we also analyzed the incidence of acute and chronic graft versus host disease (gvhd) and procedure related outcomes who underwent to myeloablative allo-hsct were retrospectively analyzed. the median age was years (iqr - ). both groups were divided regarding their age at allo-hsct [group , age ⩾ years (n = ) and group , age o years (n = )]. patient´s characteristics are shown in picture . data were collected as either continuous data and compared by two-tailed unpaired t-test or mann-whitney test, or as categorical variables and compared by chi-square. the procedure related outcomes were analyzed with the kaplan-meier test. the incidence of acute gvhd grade ii-iv was similar in both groups ( . % in group and . % in group , p = . ). the mean day to acute gvhd (grade ii-iv) development was days in group and days in group . the most involved organs in both groups were skin (group : . % and group : . % [p = . ]) and gut (group : . % and group : . % [p = . ]). at day + post-transplant patients were alive and evaluable for chronic gvhd. the incidence of cgvhd development was similar between group and ( . % versus . %, respectively, p = . ). however, severe grade s of cgvhd was high in group patients ( . % versus . %). with a median follow up of months (iqr, - ) the probability of os was significantly low (p = . ) in group ( . % +- . ) compared with group ( . % ± . ). pfs was also significantly low (p = . ) in group ( . % +- . ) compared with group ( . % ± . ). trm at months was higher in group compared with group ( . % versus . %). mortality due to relapses was also higher in group ( . % versus . %). most of the patients died during the first month. comparing both groups at this time ( months post-transplant), trm was higher in group compared with group ( . % versus . %). deaths due to relapse were also higher in this group ( . % versus . %). in our series, myeloablative conditioning regimen provides good survival rates and disease control in high risk hematopoietic diseases, however in patients aged ⩾ years confers high toxicity. it may be necessary to evaluate other strategies in this group of patients. disclosure of conflict of interest: none. allogenic hematopoietic cell transplantation (hct) is reserved for a group of high risk multiple myeloma (mm) patients having relapsed after high-dose melphalan and autologous transplantation. in general, reduced-intensity conditioning (ric) regimen are applied in this patient group with the aim of reducing transplant related mortality (trm). however, relapse of disease remains a major challenge after allogeneic hct. to address this issue, we added radioimmunotherapy (rit) to a conventional ric regimen. we have used a rhenium anti cd antibody in combination with a ric conditioning regimen. this ß-emitter leads to a so called 'cross-fire' effect allowing for bone marrow doses of gy and in parallel may target cd on myeloma cells. we hypothesized that this strategy may decrease the incidence of relapse. so far, we have treated nine patients with high risk relapsed multiple myeloma. all patients had received one (n = ), two (n = ) or three (n = ) high-dose regimens and autologous hct. conditioning therapy was flu/mel (n = ), flu/ bu (n = ) or flu/treo (n = ). flu/cy and gy tbi were applied before haploidentical transplantation. patients received g-csf mobilized pbsc from unrelated (n = ) or haploidentical (n = ) s donors. either tacrolimus/ methotrexate/ bortezomib or cyclosporine a/ methotrexate were used for gvhd prophylaxis. early extramedullary toxicity was limited. neutrophil and platelet engraftment was timely and complete in time in seven of nine cases. all patients achieved full donor chimerism around day fifteen after hct. severe acute graft-versus-host-disease (gvhd grade iii-iv) occurred in two patients and was lethal in both cases. two patients have experienced extramedullary relapse, one of them in the central nervous system and the other in the soft tissue. in two patients, a transplantation-associated thrombotic microangiopathy (ta-tma) was diagnosed. four patients are alive and in complete remission. we conclude that the combination of a ric regimen with a rhenium anti-cd radioimmunotherapy is save and feasible. the incidence of gvhd, ta-tma and extramedullary relapse will be monitored closely and will be presented in a larger patient cohort. disclosure of conflict of interest: none. the sirolimus/tacrolimus (sir/tac) combination has been associated with a better outcome after allogeneic hematopoietic stem cell transplantation (allo-hsct) when compared with conventional prophylaxis for graft vs host disease (gvhd) as cyclosporine/methotrexate in the true reduced-intensity conditioning (ric) setting but not in the myeloablative setting. in moderate-intensity regimens as thiotepa/busulphan/fludarabine (tbf), the sir/tac combination has not been evaluated. from january to december , all consecutive ric-allo-hsct recipients who received sir/tac combination to prevent gvhd in three spanish institutions were included in the study. the reduced-toxicity regimens used in this study where: (a) intravenous busulphan ( . mg/kg) and fludarabine mg/ m (bf), or (b) thiotepa days - and - and mg/kg if yrs old or mg/kg if o yrs old) on days - and - added to the bf regimen (tbf). the gvhd prophylaxis with sir/tac was given as detailed elsewhere (cutler c blood ) and was consistent within the center. the outcomes of the procedure according to the conditioning regimen were analyzed. overall, patients were included: tbf and patients in the bf group, with a median follow-up of months (range - ) and no difference in the median age ( vs years old). there were more males ( % vs %, p = . ) and more female donors to male recipients ( % vs %, p = . ) and more patients with lymphoid diseases and previous asct in the tbf group ( % vs %, p = . ), whereas there were more unrelated donors in the bf group ( % vs %, p = . ). other baseline characteristics were balanced between the groups (table ) . sir/tac prophylaxis had to be discontinued in % and % patients in the tbf and bf groups, respectively. toxicity was the main reason for discontinuation in the tbf group. the most frequent toxicities were renal injury (tbf % and bf %) and neurologic impairments (tbf %, bf %). in the bf group, the main reason of discontinuation was relapse or a mixed chimera. patients who received tbf presented higher incidence of extensive chronic gvhd ( % vs %, p = . ), higher nrm at days ( % vs %) and at years ( % vs %, p = . ). there were no differences in os ( years) between both groups ( ± . % vs ± . %, p = . ) (figure) . there were no differences regarding to acute gvhd - ( % vs %, p = . ), acute gvhd - ( % vs %, p = . ), or relapse (up to years, % vs %, p = . ) between the groups, either. the combination of sir/tac as gvhd prophylaxis was associated with higher incidence of chronic gvhd and nrm in patients receiving conditioning regimen with tbf compared to those receiving bf. there were no differences in os between both groups. [p ] cr complete remission, pr partial remission, nr non remission, hct-ci hematopoyetic cell transplant comorbility index. disclosure of conflict of interest: none. reduced-intensity conditioning regimen with fractionated total body irradiation of gy and cyclophosphamide mg/kg for allogeneic hematopoietic stem cell transplant is well tolerated and offers a potential disease control as treatment of acute leukemia and lymphoproliferative disorders m adler, t girinsky , s koscielny, g ferini, s wittnebel, s mayeur, c chahine, m vanghele, s pilorge, c castilla-llorente and j-h bourhis the use of reduced-intensity conditioning regimens (ric) before allo-hsct is widely extended since it preserves the graft-versus-leukemia effect but reduces treatment related mortality. however, there exist different ric regimens with diverse outcomes and the choice of the ric regimen relies on the type of disease treated, experience of the center and previous therapies. this is a retrospective study of patients treated in our institution within / and / . the ric regimen consisted of fractionated total body irradiation (ftbi) of gy administered in consecutive days ( gy/day) and cyclophosphamide mg/kg given in days ( mg/kg/day). post-transplant immunosuppression consisted of csa started the day before allo-hsct and short mtx on days , and after transplantation. for patients receiving transplant from unrelated donors, anti-thymocyte globulin at a dose of mg/ kg ( . mg/kg/day for days at day - and - ) was used as part of the immunosuppressant therapy. patients (median age: years: range: - years) were included. the median hct-ci was . (range: - ). primary disease was multiple myeloma (mm) in ( %), al/mds in ( %), cll in ( %), nhl in cases ( %) . patients ( %) received transplant from matched related donors, ( %) from matched unrelated donors and ( %) from mismatched unrelated donors. female to male mismatch incidence was % (n = ). most of the patients (n = ) received a peripheral blood graft. patient received a second allogeneic transplant. mm patients were transplanted in a "tandem" autologous-allogeneic hsct program in cases. the median number of chemotherapy lines prior to transplant was . in cll, . in mm and . in nhl. patients ( %) engrafted by day post transplant. neutrophil engraftment occured at a median of days (range: - days) and platelet engraftment at a median of . days (range: - days). full donor chimerism was observed in out of patients ( %) having survived by day . primary graft rejection was observed in patients. treatment related toxicities consisted of grade / mucositis in patients ( %), grade (range: - ) cardiac toxicity in patients ( %), grade (range: - ) hemorrhagic complications in patients ( %) including cases of hemorrhagic cystitis and secondary malignancies in patients, this within a median follow-up of . years. infectious complications during aplasia included fever of unknown origin (n = ), bacteremia (n = ) with cases of bacteremia with severe sepsis and cases of infections defined by bacterial foci. incidence of agvhd was % with cases of grade / refractory agvhd. cgvhd occurred in pts ( %). the non-relapse mortality (nrm) at days was % including cases of septic shock, case of acute cardiac toxicity and case of agvhd. the nrm at year was %. -year survival rates were % in al, % in cll and % in nhl with extended survival benefit. in al patients, the relapse incidence was % comprising patients who progressed during conditioning. the -year survival rate in mm patients was %. in mm patients who were in complete response prior to transplant, median overall survival was . years. the used ric regimen resulted in durable donor engraftment with an acceptable toxicity profile permitting efficient disease control in the described cohort. disclosure of conflict of interest: none. graft manipulation using selective depletion of αβ-t cells provides a source for haploidentical hematopoietic stem cell transplantation (haplo-hsct) enriched in effector cells. initial reports demonstrated safety and rapid immune reconstitution using this method, in malignant and non-malignant disorders using several proposed conditioning regimens. no specific considerations were given to hematologic malignancies. we reviewed a total of twenty seven pediatric patients who underwent haplo-hsct using αβ-t cell depletion between - in a single tertiary referral center. we report the results of procedures performed in eighteen patients transplanted for malignancies. twenty two haplo-hsct were performed in eighteen patients. the indication for hsct was acute leukemia in sixteen (all = , aml = ) and neuroblastoma in two. median age at hsct was . years. six patients had failed a prior hsct, and the remainder had no matched donor. the initial reduced-toxicity conditioning regimen consisted of melphalan, fludarabine, thiotepa and atg, and resulted in a high rate of graft rejections ( of ). thus, a totalbody irradiation (tbi)-based regimen was implemented, with prompt engraftment in all the patients. we observed rapid neutrophil and platelet engraftment kinetics (median time to engraft, days and . days, respectively). significant treatment related complications were all due to graft failure in patients receiving reduced-toxicity conditioning, with two infection-related mortalities in the presence of prolonged neutropenia. none of the patients developed hepatic sinusoidal-obstruction syndrome, and no grade - acute graft-versus-host disease (gvhd) or chronic gvhd were observed with either regimen. importantly, the majority of patients with acute leukemia were free of immunosuppression in the first days post hsct. the -year actuarial event-free and overall survival of the entire cohort were % and % respectively, with results for tbi-based conditioned patients being % and %. overall, we demonstrated that a tbibased conditioning for haplo-hsct using αβ-t cell depletion for malignant diseases resulted in acceptable outcomes in these high-risk patients without increased toxicity. disclosure of conflict of interest: none. high-dose chemotherapy conditioning regimens followed by autologous stem cell transplantation (auto-sct) generally provide good results in relapsed and refractory lymphomas. we evaluated the efficacy and safety of tecam regimen as conditioning with autologous stem cell support in patients with relapsed/refractory lymphomas. thirty-two ( patients were refractory, patients were relapse and one frontline treated) patients ( m, f) with lymphoma at various stages (stage ii, %; stage iii, %; stage iv, %) who underwent asct were included in this retrospective study. the median age at transplantation was . years (range, - years). the diagnosis were as follows: diffuse large b-cell non-hodgkin lyphoma (nhl), hodgkin lymphoma (hl), mantle cell lymphoma, follicular lymphoma, marginal zone lymphoma and t-cell nhl. all patients received tecam as conditioning regimen that consist of thiotepa ( mg/m × days), etoposide ( mg/m × days), cyclophosphamide ( mg/ kg × day), ara-c ( mg/m × days) and melphalan ( mg/m × days). median cd (+) cells were . × /kg (range; . - . × /kg) which were infused at day , followed by recombinant human granulocyte colonystimulating factor (rhug-csf) at a dose of μg/kg/day. the median time between mobilization and auto-sct was months (range; - months). the median time to recovery of absolute neutrophil and platelet counts independent of transfusion were (range; - ) and (range; - ) days, respectively. the median stay in hospital was days (range, - days). bacterial, sitomegalovirus and invasive fungal infection were detected in ( %), ( %) and ( %) patients, respectively in first days of auto-sct. three s patients ( . %) died from transplant-related complications. the overall response rate was % ( cr, . %; pr, . %) after auto-sct. relapse developed in patients during median follow-up period of . months (range; - months) after auto-sct. the -year estimated dfs ( figure ) and os were % and %, respectively. no statistical significance was observed for os and pfs in terms of gender, patient age ( o and ⩾ years) and nhl and hl lymphoma group (p ⩾ . ). the tecam regimen for auto-sct in lymphoma seems to provide encouraging results in terms of response and its good tolerance with acceptable toxicity. [p ] disclosure of conflict of interest: none. allogeneic hematopoietic cell transplantation (allosct) is the only curative treatment for myelofibrosis. however, its widespread use is limited by early non-relapse mortality (nrm). the optimal modalities of the conditioning regimen are a major unmet clinical need. in an attempt to reduce early nrm, we used a tbf conditioning regimen (thiotepa, busulfan (bu), fludarabine (flu) and antithymocyte globulin (atg)). our aim was to reduce nrm and improve engraftment by using such tbf conditioning. thirty consecutive patients with a median age of years (range, - ) who underwent allosct for primary (n = ) or secondary (n = ) myelofibrosis were included. according to the refined dynamic international prognostic scoring system (dipss-plus), patients were stratified as intermediate- (n = ), intermediate- (n = ), and high (n = ) risk. five patients had blast transformation. ruxolitinib was given to patients ( %) prior to allosct. graft source was pbscs in patients ( %) and bm in patients ( %). donors were matched related (mrd, n = ), unrelated (n = ) and haploidentical (n = ). conditioning regimen was tbf in patients ( %). in our historical cohort patients ( %) received fb (flu, bu, atg). in addition, patients received a 'tec-ric' sequential conditioning (thiotepa, etoposide, cyclophosphamide, and after days rest, flu, bu and atg) for blast transformation (n = ) or refractory proliferative myelofibrosis (n = ). gvhd prophylaxis consisted of cyclosporine (csa) and mycophenolate mofetil in patients ( %), csa and short course methotrexate in patients ( %) with abo mismatch and csa alone in patient ( %) with mrd. high dose posttransplant cy (pt-cy) was added in haplo cases. no significant difference was observed between tbf, fb and tec ric patients in terms of age, gender, karnofsky score, comorbidity index, number of previous treatment line, history of ruxolitinib administration and source of stem cells. median follow-up was months (range, - ). two tbf patients died of septic shock before engraftment at day + and + after allosct, respectively. one fb patient died of graft failure at day + post allosct. median time to neutrophils and platelets ( g/ l) recovery was days (range, - ) and days (range, - ) with tbf, days (range, - ) and days (range, - ) with fb, and days (range, - ) and days (range, - ) with tec ric. grade ii-iv acute gvhd occurred in . % of tbf patients, . % of fb patients, and % of tec ric patients (p = . ). moderate chronic gvhd developed in / evaluable tbf, / fb and / tec ric patients. no severe forms of chronic gvhd were observed. at last follow-up, patient relapsed, died and are still alive. main causes of death were disease progression (n = ), infection (n = ) and gvhd (n = ). nrm at years was . % in tbf patients, % in fb patients, and % in tec ric patients. the -year os were . % in tbf patients, . % in fb patients, % in tec ric patients, respectively. cd +-selected stem cell boost without further conditioning allowed to patients for poor graft function, with significant hematological improvement in patients. tbf conditioning regimen seems to be efficient in allosct for patients with myelofibrosis and compares favorably with previously published fb regimens. these preliminary results give a rationale to support a prospective evaluation of this platform. disclosure of conflict of interest: none. we proposed here to compare the outcome of patients receiving either thymoglobuline (atg), a rabbit anti-human thymocyte immunoglobulin or campath, a recombinant dna-derived humanized monoclonal antibody directed against cd . campath and atg are both commonly used as in vivo tcd before hsct, respectively in united kingdom and france but very few comparing data are available. all consecutive patients with acute myeloblastic leukemia (aml), myelodysplastic syndrome (mds) or myeloproliferative neoplasia (mpn) who received a reduced intensity hsct from an unrelated donor transplanted between and were included in this study. a propensity score was used to identify and control potential confounding to relate the treatment group to the outcomes. in the matched sample, cox regression model was used to describe the association between treatment and outcomes. patients have been included. all patients received fludarabine and busulfan with either atg (n = ) or campath (n = ). patients treated by atg received cyclosporine plus mycophenolate mofetil or methotrexate and patients treated by campath received cyclosporine alone as gvhd prophylaxis. comparing patient and transplant characteristics, atg patients were older ( vs years), had less often aml ( vs %), had higher disease risk (adverse dri: vs %; poor cytogenetics: vs %; high cibmtr score: vs %), were less often in complete remission at time of transplant ( vs %) and were transplanted less often from a mismatched hla donor ( vs %). cumulative incidence of sustained engraftment was in % and % campath and atg patients. time to neutrophil engraftment was longer in atg patients ( vs days). acute gvhd ii to iv rate were higher after atg ( % vs %) as well as chronic extensive gvhd ( % vs %). relapse rate was higher after campath ( % vs %). disease-free survival (dfs) was higher after atg ( vs %) and the gvhd-free relapse free survival (grfs) was similar ( % vs %). according to the prognostic factors for outcome, a propensity score was developed selecting patients from the original cohort. the estimation of tcd effect was than studied. relapse risk was higher in patients treated by campath while there is a non-significant advantage for atg in dfs (table ) . [p ] tcd with atg or campath gives similar os, dfs and grfs. severe acute or chronic gvhd is lowered by campath but the higher relapse risk counterbalances the potential benefit of campath finally given similar os. nevertheless, lower risk disease patient might benefit from campath while higher risk patients might benefit from atg. disclosure of conflict of interest: none. high-dose chemotherapy (hdt) with autologous stem cell transplantation is the standard of care for relapsed/refractory (rr) or high grade non-hodgkin-lymphoma (nhl) and hodgkin-lymphoma (hl) . the standard hdt in autologous stem cell transplantation (asct) for lymphoma is carmustinebased hdt using a combination of carmustine, etoposide, cytarabine and melphalan (beam); this standard conditioning programme is used by most groups worldwide . we have designed novels hdt regimens in which carmustine was substituited by an equal dose of fotemustine (feam) or thiotepa (team) and we compared these two hdt regimens in terms of engraftment times, toxicity, tolerability and frequency of relapse after asct. from february to september we consider a total of relapsed/refractory patients affected by hl and nhl respectively hl and nhl with different grade of initial disease (grade i-iv) and different response to prior treatments. the all other drugs were administered according to a standard beam regimen . after a day of rest, autologous peripheral blood progenitor cells were infused on day , followed by s.c. g-csf ( mg/kg) from day of asct until consecutive days when the ancs were × /l . the primary objectives of the study were to assess the feasibility and safety of the feam and team regimens in terms of acute toxicity, grade of mucositis, hemopoietic engraftment and relapse after asct. acute toxicity include chemotherapy-induced nausea and vomiting, diarrhea, hepatotoxicity, nephrotoxicity and infection complication. in all patients cd + cells were collected from peripheral blood and the median number of infused cells per patient was . × e /kg. the median time of engraftment was days for neutrophil recovery (n × /l) and days for plt recovery ( × /l). acute toxicity occurred in total patients ( . %), mucositis grade - occurred in patients ( % of cases). frequency of relapse in all cases was . %. feam conditioning regimen was used in cases showing a median time of neytrophil recovery of days and a median time of plt recovery of days. acute toxicity occurred in of these cases ( . %), mucositis grade - occurred in patients ( . % of cases). frequency of relapse in feam group of patients was . %. team conditioning regimen was used in cases showing a median time of neytrophil recovery of days and a median time of plt recovery of days. acute toxicity occurred in of these cases ( . %), mucositis grade - occurred in patients ( . % of cases). frequency of relapse in team group of patients was %. relapse/progression of lymphoma and conditioning regimen toxicities remain limitations to treatment success. the two novels hdt regimens feam and team are safe and feasible and show similar engraftment times, tolerability and frequency of relapse. maybe the team regimen shows toxicity slightly higher than feam regimen but longer follow-up is needed to evaluate fully its efficacy and long-term safety. disclosure of conflict of interest: none. treosulfan is a prodrug of a bifunctional alkylating cytotoxic agent. there are few reports regarding toxicological side effects of treosulfan-based conditioning prior to hsct. here we report on incidence of early potential treosulfan-related toxicity in patients treated with treosulfan-based conditioning before hsct. treosulfan was given at a dose of g/m /d for days in combination with fludarabin mg/m /d for days prior to hsct. most patients (n = ) had a haematological malignancy, while patients had a non-malignant disorder as hsct indication. an hla-a, -b and -dr matched unrelated donor (mud) was used in cases, patients had a hla-identical sibling donor and received an hla-a, -b or -dr allele mismatched unrelated donor. as graft versus host-disease (gvhd) prophylaxis, most patients (n = ) received cyclosporine and methotrexate. patients medical records were scrutinized retrospectively to collect laboratory tests (aspartate aminotransferase (ast), alanine aminotransferase (alt), creatinine) before hsct and then weekly until weeks after hsct. levels of ast and alt were significantly increased week after hsct compared to before hsct. however, only a few patients had transaminase levels over or times the upper normal level (unl) levels decreased sharply after the first week. most of the cases with high levels of ast/alt at one week had normal or close to normal levels before hsct. creatinine levels increased after week but no patient had levels ⩾ × unl. clinical features of all oral mucositis (om) were recorded using the world health organization (who) scoring system. most patients ( %) had no or very limited (grade i) om, % had grade ii and % had grade iii or iv of om. according to our toxicological results this is low-toxic protocol. however, all patients became neutropenic, % already at the time of graft infusion, indicating that the protocol has a myelo-toxic effect comparable to conventional mac protocols. all patients engrafted, except three patients who died very early. median time to neutrophil and platelet engraftment was (range - ) and days ( - ), retrospectively, which is significantly later when compared to engraftment data for other ric protocols used at our centre (data not shown). median duration of neutropenia (o . × /l) was days , comparable to what is expected after conventional mac conditioning. secondary graft failure (gf) occurred in ( . %) patients, all having a nonmalignant disorder and / having a urd. non-relapse mortality (nrm) was . % ( % ci . - . %) at days and . % ( . - . %) at one year after hsct. causes of death within one year after hsct were: relapse , epstein-barr virus associated posttransplant lymphoproliferative disease (ptld) , other infections , organ failure , gvhd , hemophagocytic lymphohistiocytosis (hlh) . other infections occurring within days after hsct were cytomegalovirus (cmv) reactivation ( %), invasive fungal infection ( . %) and blood stream infection ( %). veno-occlusive disease of the liver or sinusoidal obstruction syndrome (vod/sos) occurred in one patient and haemorrhagic cystitis in two patients. this study shows that early regimen-related toxicity after hsct was low despite similar marrow toxicities compared to mac regimens. disclosure of conflict of interest: none. allogeneic stem cell transplantation from haploidentical donors (haplosct) is an increasingly adopted option for patients (pts) with high-risk hematological malignancies. in our institution, we previously described a platform for unmanipulated peripheral blood stem cell (pbsc) haplosct using a calcineurin-free gvhd prophylaxis with sirolimus, micophenolate and anti-human t-lymphocyte immunoglobulin (atg) after conditioning with treosulfan and fludarabine (trramm; peccatori et al., leukemia ) . as an attempt to decrease relapse rate, especially in advanced-phase diseases, we designed a new phase ii prospective clinical trial intensifying conditioning regimen with the addition of gy total-body irradiation (tbi) (trramm gy; eudract# - - ). we report results on pts. pts affected by aml (n = ), other myeloid (n = ) and lymphoid (n = ) malignancies were prospectively enrolled from may to june . median pts age was y (range - ). revised disease risk index (r-dri) was low or intermediate in pts, high in pts and very-high in pts. twenty-five pts had previously received an allogeneic stem-cell transplantation with a median time from st to nd sct of months . median hct-comorbidity index by sorror et al. was ( ) ( ) ( ) ( ) ( ) ( ) . pts received a myeloablative conditioning regimen consisting of treosulfan ( g/m /d from - to - ), fludarabine ( mg/m /d from − to − ) and tbi gy (fractionated in doses, from − to ). source of stem cells were unmanipulated g-csf-mobilized pbsc from haploidentical donors. gvhd prophylaxis consisted of atg-fresenius (grafalon, neovii) mg/kg/d from − to − , rituximab mg/m on − , mycophenolate mofetil mg/ kg from − to + and sirolimus (target concentration - ng/ ml) from − . median infused cd + and cd + cell doses were . × ⁶/kg and . × ⁸/kg, respectively. median follow-up for survivors was months ( - ). neutrophil engraftment occurred in % of pts with a median of d ( - ), platelet engraftment was reached in % of pts with a median of d ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) . the -d cumulative incidence (ci) of grade ⩾ acute gvhd (agvhd) was ± % and of grade ⩾ agvhd ± %; the years ci of chronic gvhd was ± %. the ci of transplant-related mortality (trm) at y and y were ± % and ± %, respectively. the ci of relapse at y and y were ± % and ± %, respectively, with a median time to relapse of d . interestingly, we did not observe any extramedullary relapse; loss of mismatched hla-haplotype occurred in % of relapses. among pts who were in active disease at time of haplosct and who were evaluable, % achieved complete remission (cr) and full donor chimerism at day+ . the y and y probabilities of disease-free survival (dfs) were ± % and ± %, respectively. at y, % of pts are alive, disease-free and immunosuppression-free minimal residual disease, tolerance, chimerism and immune reconstitution the number of human leukocyte antigen (hla)-mismatched hematopoietic cell transplantation (hct), including cord blood transplantation, has been increasing. hla-flow method can discriminate mismatched hla antigens between donor and recipient by using flow cytometry, and can evaluate minimal residual disease (mrd) or chimerism after hla-mismatched hct. by developing more simple methodology, hla-flow might be more widely applicable. we have developed modified -colorbased hla-flow method. the aim of this study is to evaluate the utility of the -color-based hla-flow for monitoring of mrd and chimerism after hla-mismatched hct in children. from june to november , serial monitoring of mrd or chimerism by the -color-based hla-flow was performed in twelve patients undergoing hla-mismatched hct ( tests). nucleated cells obtained from bone marrow were stained by immunofluorescent antibodies against hla antigens mismatched between donors and recipients. these cells were also stained by immunofluorescent antibodies against surface antigens such as cd , cd , cd , cd and cd /cd for determining lineage of the cells. these surface antigens were also used as a marker of leukemic blasts in the mrd study. we used -color-based flow cytometry (facs-navious) and the data were analyzed with flow jo. erythroblasts and dead cells were excluded from the analysis. in each study, at least cells were analyzed. for mrd analysis, we concurrently tested real-time quantitative polymerase chain reaction (pcr) of peripheral wt mrna or leukemia-specific fusion genes. pcr of polymorphic short tandem repeats or fluorescent in situ hybridization of x/y chromosomes was concurrently tested for chimerism study. age of patients ranged from to years. donor sources included bone marrow (n = ) and cord blood (n = ). for mrd monitoring of acute leukemia (n = ), the -color-based hla-flow could detect mrd in three patients. five patients have not experienced relapse. no discordance with other mrd markers was observed in these patients. hla-flow could not separate donor-derived cells from recipient-derived ones in one patient receiving bone marrow transplantation. as for chimerism testing (n = ), the -colorbased hla-flow could successfully evaluate quantitative lineagespecific chimerism in all patients. there is no discrepancy between hla-flow and other methods. we could complete evaluation of the -color-based hla-flow within two days in all tests. the -color-based hla-flow is a simple, quick and useful method for the quantitative evaluation of mrd and lineagespecific chimerism after hla-mismatched hct in children, irrespective of donor sources. it is thought that our method is applicable in all institutions owing -color-based flow cytometry. acknowledgement: we thank drs. nobukazu watanabe, eri watanabe and natsuko sato (university of tokyo) for their technical advices. we also thank drs. tomoko okunushi (chiba university), hidefumi hiramatsu and katsutsugu umeda (kyoto university) for their care of patients involved in this study. disclosure of conflict of interest: none. survival (pfs), cumulative incidence of relapse (cir) and acute/ chronic gvhd incidence in aml patients (pts) submitted to allo-hsct at our institution between january and december . this retrospective study evaluated aml pts submitted to allo-hsct from matched sibling donors (msd) and matched unrelated donors (mud) who provided bone marrow (bm) or peripheral blood as stem cell grafts.ir was evaluated at , and days post-transplant in all pts. cmv-dna copies were determined in peripheral blood by quantitative pcr twice weekly in the first days post-transplant and subsequently once weekly. cmvi/r was analyzed as a timedependent covariate.effect of cmvi/r on os and pfs was estimated by cox proportional hazard model. cir and gvhd incidence were analized with a competing risk approach, considering death from any cause as a competing event. effect of cmvi/r on cir and gvhd were evaluated by fine & gray model. median age at allo-hsct was . years ( . - . ). in our population % of donors were seropositive for a previous cmv infection and pts transplanted from these donors showed a significantly lower cumulative incidence of cmvi/r than pts transplanted with seronegative ones (shr = . , %,ci: . our study demonstrates that cmvi/r influences the success of allo-hsct by determining a better ir characterized by a higher cd + cell number that might exert an immune protective control on disease outcome by improving os,pfs and cir with no effect on gvhd. another factor of utmost importance to achieve this same goal might be constituted by the significantly increased nk cell number six months after allo-hsct. to assess the dynamics of molecular response to treatment in aml adult patients with concomitant flt and npm mutations. this retrospective single center studystudy was approved by the institutional review board of american university of beirut medical center. twelve consecutive newly diagnosed (n = ) or relapsed (n = ) aml patients received idarubicin/cytarabine induction and one or two consolidation (s) ( table ) . seven patients received allogeneic stem cell transplant (allo-sct) and had haploidentical-sct (hap-lo_sct); all followed by post-transplant sorafenib maintenance. median follow-up was . ( - ) months. all transplanted patients remain alive and disease free.flt mutation was tested on dna using a qualitative method with a sensitivity of . %. npm- mutation was tested on cdna using a qualitative or a quantitative rt-pcr with a sensitivity of . % and . ncn respectively. patients were tested at diagnosis, after induction, after each consolidation, before and s at days , and after allo-sct for kinetics of npm and flt molecular response. after induction, flt became negative in all tested patients (n = ). after first consolidation, flt- was not tested in patients who had a negative result after induction, was negative in patients including the patients who were not tested after induction, whereas a molecular relapse was noted in one patient who developed a hematological relapse and rapidly died. another patient died after the third consolidation, in complete remission, due to septic shock. no molecular positivity for flt- was noted later on, whether after second consolidation or post-transplant. conversely, npm- mutation became negative in out of tested patients after induction, in additional patient after first consolidation and in additional patients after sct, mostly after starting sorafenib. npm- mrd value remained elevated in out of patients with quantitative assessment at diagnosis and post induction (figure ). flt become negative early after induction while npm negativity lags behind. persistent npm- mrd does not seem to predict post-transplant outcome and may indeed become negative after sorafenib. these results need confirmation in larger studies. disclosure of conflict of interest: none. in allogeneic stem cell transplantation (allo-sct), an early detection of the transplant outcomes such as overall survival (os), event-free survival (efs), cumulative incidence of relapse (cir) and non-relapse mortality (nrm) is fundamental regarding the use in time of additional therapy after sct. therefore, we investigated the association between early immune reconstitution (ir) on day + after allo-sct and outcomes in children suffering from acute leukemia or myelodysplastic syndrome (mds). this study collected data from allo-sct from january until december in our institution. the median survival follow-up was months. indications of allo-sct were all (n = , %), aml (n = , %) and mds (n = , %). the median age was years (range, . - ). patients were transplanted in cr (n = , %) and pr/nr (n = , %). patients included in the study received st sct (n = , %), nd sct (n = , %) or sct (n = , %). grafts were from sibling (msd; n = , %), matched unrelated (mud; n = , %), haploidentical (n = , %) or mismatched unrelated (mmud; n = , %). conditioning regimens were tbi-based (n = , %) or chemo-based (n = , %). stem cells were from bone marrow (n = , %) or peripheral blood (n = , %). we analyzed the absolute count of lymphocytes (alc), monocytes, cd + t cells, cd +cd + t-helper cells, cd +cd + cytotoxic t-cells, cd -cd + natural killer (nk) cells and cd + b cells assessed on day ± after sct. we used the percentiles of the lymphocyte subsets of the same cohort to categorize the samples throughout the study. patients with alc over the th percentile of alc (alc o cells/μl) had a . -fold increased hazard ratio (hr) to develop relapse (p = . ). nk cell counts on day after sct were strong associated with os, efs, cir and nrm. patients with nk cell count over the th percentile (nk . cells/μl) had increased hr for os (hr = . , p = . ) and efs (hr = . , p = . ) compared to patients with nk count under the th percentile. patients with nk cells over the th percentile (nk o . cells/μl) had a hr = . (p = . ) for relapse and hr = . (p = . ) for nrm compared to patients with nk cell count under the th percentile. monocyte cell count on day correlated with os, efs and cir. patients with cd + cells count under the th percentile of cd + (cd + o cells/μl) has an increased hazard ratio for os, efs and relapse compared to patients with cd + cell counts over the th percentile. no association between absolute cell count of cd +, cd +, cd + and cd + on day + after allo-sct and any outcomes either os, efs, cir or nrm was found. the study confirms the strong association between early ir and outcomes after allo-sct in children. our study suggests that especially nk cell and monocyte cell count on day + may have significant prognostic implications. our findings suggest that the cells count of alc, nk cells and monocytes on day + after allo-sct could be useful to predict outcomes after allo-sct and should be taken into account in considering alternative treatment. disclosure of conflict of interest: none. early immune reconstitution (eir) has proven to be a significant determinant for the outcome of allogeneic hematopoietic stem cell transplantation. in the setting of unmanipulated haploidentical transplantation (haplo-hsct), some groups have identified the absolute leukocyte count on day + (alc ) as an independent prognostic factor in terms of overall survival (os), disease free survival (dfs) and infectious mortality (im). the aim of this study was to evaluate the impact of eir on os, dfs and im among patients who underwent haplo-hsct with postransplant cyclophosphamide (ptcy) at our institution. from july to april , haplo-sct were performed at our institution. threedied before day after haplo-sct, and patients had missing data. conditioning regimen consisted of fludarabine, cyclophosphamide and busulfan. twenty-nine patients received a reduced intensity conditioning regimen ( - days of busulfan) while a myeloablative regimen ( - days of busulfan). gvhd prophylaxis comprised ptcy, cyclosporine and mycophenolate mofetil. patients were assessed for eir by means of alc , cd + t lymphocyte count on + (cd ), nk lymphocyte count on + (nk) and nk cd bright percentage on + (cd br ). we analyzed pts, with a median follow-up of months ( - ). the median age of the pts was (range - ), %men. diagnosis were: aml( %), hl ( %)non-hl ( %), all ( %),mds( %), cml( %), others( %). % were in complete remission at the time of transplant, % in partial remission and % had overt disease. in terms of infectious complications, cmv reactivation was documented in % of the pts, % developed a proven invasive fungal infection and % suffered from bk+hemorrhagic cystitis. median os and dfs were ( - ) and months ( - ), respectively. im rate was % at the end of follow up. median follow-up was months ( - ). roc curves were used to determine the optimal cut-off values for each of the studied parameters: cells/μl for alc , cells/μl for cd , cells/μl for nk and % for cd br were chosen. pts with alc ⩾ /μl had better os (p = . ) and dfs (p = . ), than those with alc o /μl. median os and dfs were months vs not reached (nr) and months vs nr, respectively. pts with cd br ⩾ % had better os (p = . ) than those with lower values. median os was months vs nr; however no difference was seen in terms of dfs. we didn´t observe statistically significant differences in os or dfs, among pts with different levels of cd and nk on + . cumulative incidence of im was significantly lower in pts with an alc ⩾ (p = . ), pts with cd ⩾ /μl (p = . ) and pts with nk ⩾ (p = . ); patients with cd br ⩾ % showed tendency to have lower cumulative incidence of infectious mortality (p = . , non-significant). cumulative incidence of relapse was not affected by alc , cd , nk or cd br . our study supports the independent prognostic value of early immune reconstitution after unmanipulated haploidentical transplantation with ptcy, especially in terms of lower infectious mortality. os and dfs were better among patients with alc ⩾ cells/μl. pts with cd br ⩾ % also showed better os. no correlation was found between cd or nk on + with os or dfs. cumulative incidence of infectious mortality was affected by alc , cd and nk on + ; while cd br seems to have less impact. [p ] disclosure of conflict of interest: none. an early absolute lymphocyte count (alc) recovery after autologous stem cell transplantation (asct) for hematologic malignancies has been related with an improved transplant outcome due to a faster autologous immune restoration. in this retrospective study we analyze post-transplant survival of non hodgkin lymphoma (nhl) patients and its relation with alc at day + post-asct. we analyzed consecutive adult nhl patients who underwent asct at the hematology and sct department of hospital maciel (montevideo, uruguay). only individuals with at least months post-transplant follow up were included. all patients received beam (bcnu, etoposide, cytarabine and melphalan) conditioning regimen followed by peripheral blood stem cells previously collected by apheresis. median cd + cell dose was . × e /kg ( . - . ). median alc at day + was /μl. patients were divided into two groups: alc at day + inferior than /ul (group ) and alc at day + superior or equal than /ul (group ). differences between groups were analyzed using t-student and chi-square tests, with statistical significance determined at p o . . disease free survival (dfs) and overall survival (os) were analyzed by kaplan meier method. differences in survival between groups were determined by log-rank test. no differences were observed between groups regarding gender, histology, disease status at transplant and cell dose. patients in group were older and more heavily pretreated. neutrophils and platelets engraftment were significantly faster in group (table ) . after a median follow up of months, disease-free survival (dfs) and overall survival (os) were superior in group . median dfs was months and not reached (p = . ) and os was months and not reached (p = . ) in groups and respectively (figure ). an early alc recovery after asct was associated with a superior dfs and os in nhl patients. individuals with alc major or equal than /ul had also a shorter time to neutrophils and platelets recovery and a shorter hospital stay. in this study, cd + cell dose does not seems to be a determinant factor for lymphocyte recovery. the load of previous treatment may influence lymphocyte recovery after asct. these results support the association between early post-asct lymphocyte recovery and improved survival in nlh patients. [p ] disclosure of conflict of interest: none. t cell depletion (tcd) reduces the risk of graft versus host disease (gvhd) but also the graft versus leukaemia (gvl) effect, thus increasing the risk of relapse. donor lymphocyte infusions (dli) can be given to boost donor chimerism, with the intention of enhancing the gvl effect. it is not currently known whether giving dli based on bone marrow chimerism (bmc) influences survival, or whether certain groups of patients benefit more from dli than other groups. in addition, it is not known whether the overall aim of achieving % bmc associates with improved survival. we investigated whether day (d ) bmc was predictive of survival, and whether giving dli based on this result was associated with improved overall survival. data were retrospectively collected from case notes and laboratory reports for patients who underwent allogeneic stem cell transplant (allosct) for aml or mds at the northern centre for bone marrow transplantation between and . patients who died prior to d were excluded from the analysis. of the patients analysed (aml , mds ), % were male and % female. the median age was years (range - ). conditioning was with flu/bu/ alemtuzumab ( ), flu/mel/alemtuzumab ( ), cy/tbi alemtuzumab ( ), flamsa tbi/bu atg/alemtuzumab ( ), other ( ) . ( %) received a graft from an unrelated donor, ( %) a matched sibling donor and ( %) another source. ( %) received mobilised pbscs, ( %) bone marrow and ( %) cord blood. statistics were performed using graphpad prism. p values were calculated using the chi square test and taking po . to determine significance. bmc was divided into groups %, - % and o %. % bmc at d was associated with a significant increase in year overall survival (os) ( . % vs . % and . % for - % and o %, respectively, p o . ). patients with a d bmc o % had a year os of o % (with relapse the cause of death in %). in patients whose d bmc was o %, there was a significant improvement in year os seen in those who received dli ( . % survival at years vs % with no dli, po . ) (figure : os by d bm chimerism (with and without dli). attainment of % bmc at a subsequent time point also significantly improved survival in those with a d bmc of - % ( . % year survival vs . % who never attained %, p o . ) and o % ( % year survival vs . %, p o . ). we found d bmc to be predictive of os in this population. in addition, dli was associated with an improvement in os, especially in patients whose bmc at d was o %. there was also a statistically significant improvement in os seen in patients who subsequently attained a % bmc, where it was o % at d . the objectives of this analysis were to examine the optimal alc recovery cutoff utilizing receiver operator characteristics (roc) analysis and to examine infused allograft characteristics associated with early alc recovery. after due irb approval, patients (pts) with aml and all who underwent hct at our institution between - were identified. pts with t-cell depletion or maintenance post hct were excluded. data were collected retrospectively from the patient's records. cellular contents of infused products (cd , cd , tnc, mnc, alc and amc) in addition to alc post hct were analyzed and optimal cutoff, if present, was established using roc analysis for the end point of relapse. time to end point analysis was computed using the kaplan-meier with log ranks. for competing events, cumulative incidence was computed using grey's model. univariable and multivariable analyses were performed using cox proportional hazard regression. a total of pts met the inclusion criteria and were analyzed. optimal alc cutoff by roc analysis was established to be on day + (d ) with alc . × /l and was subsequently defined as early lymphocyte recovery (erl). pts with alc ⩽ . × /l were deemed to have delayed lymphocyte recovery (dlr). patients were subsequently stratified accordingly and patient, disease and transplant related factors were well balanced between the groups. median follow up of the entire cohort was ( - . ) months. graft characteristics: roc analysis established optimal cellular cutoff, if present to predict elr. pts in the elr group were more likely to receive cd × /kg o ( . ), cd × /kg ( . ) and alc . × /kg (p = . ). we did not find a significant threshold for other allograft variables i.e. (tnc, mnc or amc). post hct outcomes: at years, corresponding cumulative incidence of relapse (cir) and non-relapse mortality (nrm) was . % vs . % (p = . ) and . % vs . % (p = . ), for elr and dlr cohorts, respectively. there was a trend towards improved progression free survival (pfs) and overall survival (os) in favor of elr vs dlr at . % vs . % (p = . ) and . % vs . % (p = . ), respectively. median time to neutrophil and platelet engraftment was and days, respectively for both groups. incidence of acute graft vs host disease (agvhd) was similar (p = . ); however, chronic gvhd (cgvhd) was more prevalent in the elr group at % vs %, respectively (p = . ). on s multivariable analysis for relapse, elr retained its prognostic significance with hr . ( . - . ; p = . ). cgvhd and first complete remission (cr ) at the time of hct were also protective factors from relapse in multivariable analysis. we observed that elr is an independent predictor for relapse in patients receiving allogeneic hct for acute leukemia with a trend towards improved os. this is possibly related to higher incidence of cgvhd. elr was influenced by infused allograft characteristics particularly cd count. given the sample size and retrospective nature of the analysis, these important observations should be examined prospectively. disclosure of conflict of interest: none. allosct is the only curative option for the treatment of hematological disorders with depression of hematopoiesis and primary immunodeficiencies.non-myeloablative conditioning (mac) regimens lead to long persistence of mixed chimaerism (mc) in the majority of patients. purpose: to estimate the relationship between type of hematopoietic chimaerism and appearance of gvhd in patients with non-malignant diseases after allosct eleven patients ( boys and girls) with median age of years (range - ) were included in the current study. among them there were patients with severe aplastic anemia (saa), with fancony anemia (fa), with thalassemia, with nijmengen syndrome, treated in our center from to . donors' sources were as follows: siblings in cases, mud ( / ) in ones. in cases bone marrow aspirate were used, in mobilized peripheral blood hematopoietic stem cells. conditioning regimens included fludarabin, cyclophosphamide and horse atg for saa patients, in fa and nijmengen syndrome patients this scheme was augmented by low-dose busulfan. in thalassemia patient we used mac with busulfan, fludarabin and horse atg. in majority of case gvhd prophylaxis consisted of tacrolimus and methotrexate combination. when allosct was performed form mud patients were additionally administered with mycophenolate mofetil. median of follow-up period was mo (range - ). quantitative evaluation of chimerism was done by multiplex amplification of str loci with subsequent fragment analysis using «cordis plus» kit («gordiz llc», russia). we analyzed whole bone marrow and peripheral blood together with cd + and cd + lymphocyte subpopulations. presence of ⩾ % donors' hematopoietic cells was considered as complete donor chimerism (cc), less than % was considered as mc. all patients engrafted in time and all of them are alive at the time of current analysis. there were no severe life-threatening complications, infections or graft rejections. only patients achieved cc at day + . at day + only these patients stayed in cc. at this time point mc was mainly revealed in cd + lymphocytes. in year after hsct proportion of cc patients enlarged to % ( patients did not achieved this time point). there is no any correlation between time of engraftment and chimerism value at day + , either between the dose of transplanted cd + cells and chimerism level ((p . in both cases). severe gvhd was noted only in female patients with cc at day + . in the first case it was acute gvhd grade iii after hsct from mud, in the second case extensive form of chronic gvhd in year after hsct from sibling was observed. there are no other cases of grade iii-iv acute gvhd in the observed cohort of patients. localized form of chronic gvhd [p ] was revealed in ( %) patients. in other patients there were no signs of chronic gvhd. despite limited number of observations we assumed that fast achievement of cc corresponds to severe gvhd. and vice versa, long persistence of mc prevented emergence of gvhd. however our findings need to be confirmed in a larger group of patients and preferably in a multicenter setting. disclosure of conflict of interest: none. interest of quantitative assessment of hematopoietic chimerism by real-time quantitative polymerase chain reaction after hematopoietic cell transplantation for hematological malignancies: a retrospective analysis on adult patients from rennes university hospital g laure , c mathilde , b marc , n stanilsas , d charles , l christine , a mehdi , lb laetitia, s gilbert , l thierry and r virginie department of hematology, chu pontchaillou, rennes; immunogenetics and histocompatibility laboratory, etablissement français du sang, rennes and etablissement français du sang-bretagne chimerism (percentage of recipient versus donor-derived blood cells) is used to document engraftment after hematopoietic stem cells transplantation (hsct). detection of persistant host cells, - as well as an increase in recipient cells chimerism has been associated with impaired dfs and os. quantitative real time pcr (qrt-pcr) is a highly sensitive, reproducible method, which can detect very low levels of recipient cells. the aim of this study was to evaluate the prognostic impact of early chimerism days (d ) and days (d )) after hsct and the meaning of detection of an increase of chimerism, even at low levels, during follow-up. adult patients who underwent hsct in rennes between and were included in this retrospective study. all chimerism analyses were done with qrt-pcr using whole blood sample. complete chimerism (cc) was defined by less than . % recipient cells detected. with a median follow-up was days, patients relapsed with a median time of . days after hsct. both d and d mixed chimerism (mc) ( . % recipient cells detected) were associated with an increased relapse risk (p = . and p o . respectively) compared to patients with cc in univariate analysis. however, when looking at subgroups analysis, d and d mc vs cc was significantly associated with increased relapse risk in this cohort for myeloid diseases (p = . and p o . ) but not for lymphoid diseases (p = . and p = . ). no difference in os was observed (p = . and p = . ). more important, detection of an increased of mc (imc) was associated with an increased relapse risk in univariate and multivariate analysis (or = . [ . ; . ], or = . [ . ; . ]), (po . ), as well as impaired os (p = . ) and dfs (p o . ). among the patients with aml and at least chimerism analysis available, only relapsed without imc detected but the patients' last available chimerism analysis was , and days before relapse respectively. median levels of recipient cells detected was . %. altogether, these results indicate that serial analyses of chimerism with qrt-pcr are a useful tool for post-transplant monitoring and might help identify patients at highest risk to relapse after transplant, especially in myeloid disease. monitoring frequency is critical in order to obtain the highest clinical impact, and the timing of monitoring as well as the safety and type of pre-emptive interventions still need to be explored. considering the kinetics of the disease, frequent analysis in myeloid pathology might improve the detection of impending relapse. although an approximately -log higher sensitivity of quantitative pcr (qpcr) compared to short tandem repeat (str) has been documented in different studies, the latter remains the standard procedure for hc assessment to date. we hypothesized that qpcr could be superior to str for monitoring the molecular kinetics of donor cell engraftment, response to donor lymphocyte infusions (dli) and development of relapse post-hct. we analyzed patients (pts) who underwent mainly / hla-matched unrelated hct mostly for acute myeloid or lymphatic leukemia at the university hospital essen between and . transplant conditioning was mostly myeloablative and gvhd prophylaxis was by cyclosporin a and methotrexate without anti-thymocyte globulin (atg). median follow-up of pts was days (d) ( - ). cytomegalovirus (cmv) reactivation in the first d posttransplant was measured by pp (n = ) or pcr (n = ). a total of retrospective genomic dna samples from peripheral blood (pb; n = ) or bone marrow (n = ) collected between d and d post-transplant were available for hc analysis in parallel by str (mentype chimera, biotype) and qpcr (alleleseq, abbott). threshold for hc positivity in qpcr was set at . % following published protocols. concordance in hc analysis between qpcr and str was found in / ( . %) samples, with all discordant cases positive in qpcr but negative in str. engraftment could be assessed in samples drawn at d -d from pts without relapse in the first months post-hct. these samples showed concordant negative or positive qpcr and str results reflective of full donor engraftment or persistent mixed chimerism (pmc) in and pts, respectively. in pts, qpcr but not str documented delayed conversion to full donor chimerism until d . in the remaining pts, positive results in qpcr but not str during early engraftment were observed exclusively in bm, in particular those drawn before d post-hct. qpcr but not str was also able to document the kinetics of conversion to full donor chimerism which took d and d in pts receiving dli for treatment of pmc and relapse, respectively. informative relapses could be assessed in samples drawn at least d before onset. / bm and / pb were positive in qpcr, compared to / bm and / pb in str, with a sensitivity of / ( %) and / ( . %) relapses, respectively. consistent with previous reports on a protective effect of early cmv reactivation on relapse in gvhd prophylaxis regimens without atg, relapse occurred in / ( %) pts who experienced cmv reactivation in the first d post-transplant, compared to / ( %) pts who did not. no apparent associations were observed between early cmv reactivation and engraftment kinetics post-hct. hc assessment by qpcr is highly concordant with str, but markedly superior for molecular monitoring of engraftment kinetics and relapse. positive qpcr results in bm should be interpreted with caution during early engraftment, while both bm and pb were highly informative for relapse in our series. these results advocate the feasibility and clinical utility of qpcr for post-hct hc monitoring in routine use. disclosure of conflict of interest: the commercial assays for qpcr chimerism analysis were provided by abbott molecular free of charge. tyrosine kinase inhibitor (tki) has become the standard of care in patients (pts) with chronic myeloid leukemia (cml) and an unavoidable tool in the combined therapy for pts with philadelphia chromosome positive (ph+) acute lymphoblastic leukemia (all). nevertheless, allogeneic stem cell transplantation (hsct) remains the standard therapy of all ph+ and of cml pts failing st line therapy with tki, with failure or insufficient response or intolerance or mutations resistant to nd generation tki, or in the advanced phase at diagnosis. in the past decade the feasibility and safety of post-hsct imatinib administration as prophylactic or therapeutic strategy was confirmed. second and rd generation tki administration after hsct is under investigation. here we are reporting our experience in post-hsct treatment with the rd generation tki ponatinib in pts treated between and at our institution. pts data and information were collected from institutional database and chapters revision. a written consent was given by pts allowing the use of medical records for research in accordance with the declaration of helsinki. pts and diseases features are reported in table . pre-transplant treatment for the all ph+ patient consisted of chemotherapy combined with dasatinib, followed by a st mud hsct and dasatinib in maintenance. the patient relapsed year after hsct with documentation of mutation v l. ponatinib was introduced as salvage treatment to bridge nd haplo hsct. pre-transplant treatment for the cml patients consisted of tki therapy with combination of chemotherapy in case of uncontrolled progression of disease. two pts received a st mud hsct but relapsed respectively months and years later. ponatinib was introduced as salvage treatment to bridge nd haplo hsct. four pts received ponatinib mg daily before the last hsct: one patient achieved sustained major molecular response, pts obtained transient response. all pts were presenting nd generation tki resistant mutations (table ) . ponatinib was started at a median of days after hsct (range, - ) as salvage treatment in overt relapse ( cases), prophylaxis ( case) or preemptive therapy ( case). acute gvhd was diagnosed in pts before ponatinib administration, of them also experienced chronic gvhd. no new cases of gvhd were observed after initiation of ponatinib. immunosuppressive treatment and azoles treatment were discontinued before ponatinib in all but one patient who was under combined treatment for chronic gvhd: therapeutic drug monitoring was closely performed without evidence of drug-drug interaction. pts were regularly evaluated for toxicities and monitored for cardiovascular events. no serious adverse events were reported in our experience: we administered ponatinib at a median maximum dosage of mg daily (range, - mg), for a median of weeks (range, - weeks). at last evaluation one patient maintained the status of molecularly undetectable leukemia (follow-up post hsct: months) and one major molecular response (follow-up post hsct months). three patients who received therapeutic ponatinib in overt relapse did not respond and died for progressive disease. ponatinib is safe and well tolerated as bridge to hsct and to maintain disease control after transplant. prophylaxis targeted therapy and preemptive therapy with ponatinib may lead to the reduction of disease relapse for high-risk ph+ leukemia. disclosure of conflict of interest: none. at nuh singapore we have adopted the cd ra depletion to ameliorate graft versus host disease. materials and methods: we have transplanted leukemia patients with cd depleted hsct followed by cd ra depleted donor lymphocyte infusion. no additional gvhd prophylaxis or gcsf was used. results: % patients achieved primary engraftment. median time for neutrophil engraftment ( /μl without gcsf) was days (range - days), platelet engraftment ( ) was days (range to days). immune reconstitution was rapid with median cd and cd cell counts /μl at day . by day median cd count was /μl (range - /μl). no patient developed grade iv acute gvhd. there was a significantly reduced incidence of invasive viral infections as compared to conventional transplants. importantly, all patients achieved complete remission (cr) on day + and remained in cr for longer time as compared to conventional transplants. conclusion: our preliminary data suggests that rapid immune-reconstitution of nk cells and t cells with this strategy correlates with reduced infection related mortality without loss of graft versus leukemia effects. disclosure of conflict of interest: none. the use of post-transplantation high-dose cyclophosphamide (pt-cy) has overcome the need for extensive depletion of t lymphocytes from haploidentical donor grafts, which traditionally resulted in severe and prolonged immunosuppression. it is therefore relevant to investigate the degree and the tempo of immune reconstitution after t cell replete haploidentical stem cell transplantation (haplo-sct) by use of pt-cy. we prospectively monitored cellular immunity in consecutive adult patients (male/female: / ), who underwent haplo-sct with pt-cy for myeloid (n = ) or lymphoid (n = ) malignancies. the median age at transplant was (range, - ) years. the conditioning regimen was myeloablative in , reduced-intensity in , and non-myeloablative in case. the source of the graft was peripheral blood (n = ) or bone marrow (n = ). in addition to pt-cy, graft-versus-host disease (gvhd) prophylaxis included tacrolimus and mycophenolate mofetil. absolute counts of cd +cd +, cd +cd +, cd + and cd +cd + cells were measured by flow cytometry at , , , , , and months post transplant. the median doses of infused cd + and cd + cells were . (range, . - . ) × /kg and . (range, . - ) × /kg, respectively. neutrophil engraftment ( /ul) was achieved at a median of (range, - ) days, whereas platelet engraftment ( /ul) was observed at a median of (range, - ) days. seven patients developed acute gvhd (grade i/ii: , grade iii: ). chronic gvhd occurred in patients, and was extensive in the of them. cytomegalovirus infection was detected in / cases at a median interval of (range, - ) days post transplant. two patients were administered rituximab for epstein-barr virus reactivation at months, whereas one patient developed bk virus-associated hemorrhagic cystitis at . months following haplo-sct. there was death due to gvhd and infection at months post transplant. at a median follow-up of (range, - ) months, / patients remain alive and disease-free. the absolute counts of t and b cells were extremely low early post transplant, while nk cells recovered from the first month (mean count, /μl). the number of cd + t cells started to increase beyond the first month, and exceeded lower normal limit at months (mean count, /μl). cd + t cells remained in general low ( o /μl) for the first months, increased moderately by months (mean count, /μl), and approached lower normal values at months (mean count, /μl) [ figure ]. of note, cd +cd ra+ naïve t cells remained significantly impaired (absolute count range, - /μl) in all patients in which they were assessed beyond the st year from transplant. cd + b cells were suppressed for the entire first trimester (mean count at months, /μl), but increased rapidly between and months (mean counts, /μl and /μl, respectively). in haplo-sct with pt-cy, reconstitution of cellular immunity can be achieved at adequate levels by - months following transplant. the observed deficit in the recovery of naïve t-helper cells may be related to a possible effect of pt-cy on thymopoiesis and warrants further investigation. [p ] disclosure of conflict of interest: none. hematopoietic stem cell transplantation (hsct) is a curative therapy for patients with sickle cell disease (scd). hemoglobin a in scd ameliorates the manifestations of the disease and this could be achieved with stable mixed chimerism after a reduced intensity hsct. this study aims to estimate the proportion of patients who develop mixed chimerism after hsct for scd and to characterize its progression in patients who develop it. this is a retrospective cohort study conducted at sultan qaboos university hospital (squh) bone marrow transplant unit in oman. we included all patients with scd who received hsct over the course of years between may to may . patients who received second hsct were excluded. short tandem repeat polymerase chain reaction was used for chimerism assessment. mixed chimerism was defined as − % chimerism at months from hsct. the data was analyzed by r program . . . χ or student t-test were used to assess the impact of acute graft versus host disease (agvhd) prophylaxis, age at transplantation, gender, red blood cell antigen alloimmunization, preparative regimen, and ferritin on the development of mixed chimerism. we included eligible patients. the median follow-up time after hsct was months (interquartile range: . − . months). the mean age at transplant was . years (standard deviation: . ). fifty-nine percent of patients were male. most patients had s/s genotype ( %), followed by s/beta-thalassemia mutation ( %). the indications for bmt were: stroke in %, acute chest syndrome (acs) in %, recurrent vaso-occlusive crisis (voc) in %, stroke and acs in %, acs and voc in %, orbital compression syndrome in %, stroke and moyamoya disease in %, and moyamoya disease in %. the two most frequently used preparative regimens were busulfan/fludarabine/atg in % and thiotepa/treosulfan/fludarabine in %. twenty-five percent of patients developed mixed chimerism at six months after hsct. on follow up of patients with mixed chimerism, % rejected the graft, % developed complete chimerism, and % continued to be in mixed chimerism. preparative regimen and the development of agvhd were statistically significant predictors of mixed chimerism at months (p values: . and . respectively). age at transplant, gender, red blood cell antigen alloimmunization, and ferritin were not statistically significant predictors of the mixed chimerism (p . ). the study confirmed that mixed chimerism can commonly be achieved in patients with scd after hsct and in majority, it remains stable on long-term follow-up. reduced intensity preparative regimen and lack of agvhd predicts the development of mixed chimerism. larger prospective studies are needed to confirm these results. disclosure of conflict of interest: none. there was a majority of male patients ( %). the median hbf level was % ( - ), median monocyte count was . /l (range: - . ), median platelet count was . /l ( - ), median marrow blasts was % ( - ) above patients who were explored by marrow karyotype, % of them had a monosomy . mutations in ptpn were detected in patients. fifty patients ( %) were treated with the bu/cy/mel regimen, whereas patients ( %) received the bu/flu/mel regimen. at years, the overall survival (os) was % ( % ci: - ). nineteen and patients developed vod after bu/cy/ mel and bu/flu/mel conditioning regimen, respectively. the cumulative incidence of agvhd - was % ( % ci: - ). the -year cumulative incidence of relapse and non-relapse mortality was % ( % ci: - ) and % ( % ci: - ), respectively. the median delay of relapse was days (range - ). among relapsing patients, were transplanted twice and one underwent hsct. in multivariate analysis, female donor to male recipient sex-mismatch, cmv status, total body irradiation and ras-double mutation/other additional mutation predicted poorer outcomes. the bu/flu/mel conditioning regimen was associated with a decreased risk of relapse. however, there was no statistical difference for os between the two main preparative regimens, bu/cy/mel vs bu/ flu/mel. our results show that allogeneic hsct may cure approximately % of patients with jmml and are similar to the best results published by other groups. relapse represents the main cause of treatment failure and a second hsct should be proposed. despite a decreased risk of relapse with the bu/ flu/mel regimen, there was no statistical difference in terms of os between the two main conditioning regimens, bu/flu/mel vs bu/cy/mel. disclosure of conflict of interest: none. allogeneic hematopoietic stem cell transplantation (ahsct) is the only curative treatment modality for the majority of pediatric patients with myelodysplastic syndrome (mds) and myeloproliferative neoplasms (mpn). the purpose of this study is to evaluate overall (os) and failure (relapse or death from any cause) free survivals (ffs), non-relapse mortality (nrm) and relapse incidence in children who underwent ahsct for mds or mpn in a single center from turkey. we retrospectively analyzed ahsct carried out in patients (median age: . years; range: . - ; males). thirty four had primary mds and had secondary mds. according to the modified who mds and mpn classification, had refractory cytopenia (rc), , refractory anemia with excess blast (raeb), , refractory anemia with excess blast in transformation (raeb-t) and , juvenile myelomonocytic leukemia (jmml). amongst patients with secondary mds, had been treated for acute myeloid leukemia, had been treated for non-hodgkin's lymphoma and had been treated for acute lymphoblastic leukemia, retinoblastoma and osteosarcoma, each, previously. donors were related in transplantation ( haploidentical transplantation) and the stem cell resources were bone marrow (n = ), peripheral blood (n = ), cord blood (n = ) and bone marrow +peripheral blood (n = ). three-year ffs and os for patients with mds were % and . %, respectively; and for patients with jmml, % and %, respectively. crude incidence of nrm and relapse for entire group were % and %, respectively. ahsct offers durable ffs and os for a significant group of pediatric patients with mds and mpn. less toxic conditioning regimens could result in better results in some patients. disclosure of conflict of interest: none. allogeneic stem cell transplantation in children with autism z antonella, g alessandra, ma beatriz and r vanderson bone marrow transplantation unit, hospital sirio-libanes, são paulo, brazil autism spectrum disorders (asd) are severe heterogeneous neurodevelopmental abnormalities characterized by dysfunctions in social interactions and communication skills, restricted interests, repetitive, and stereotypic verbal and non-verbal behaviors. the etiology of asd remains unknown, but recent studies suggest a possible association with altered immune responses and asd. inflammation in the brain and central nervous system has been reported with microglia activation and increased cytokine production in postmortem brain specimens of individuals with asd. other studies have established a correlation between asd and family history of autoimmune diseases, associations with mhc complex haplotypes, and abnormal levels of various inflammatory cytokines and immunological markers in the blood. the paracrine, secretome, and immunomodulatory effects of stem cells would appear to be the likely mechanisms of application for asd therapeutics. we describe two cases of patients with asd who underwent hsct for acute lymphoblastic leukemia (all) and whose symptoms were markedly decreased like an improvement of social interaction, communication, and behaviors. the first patient is an -year-old girl with asd who was diagnosis with ph-positive all in october (at the end of treatment, bcr-abl remained positive). she underwent a matched sibling hsct in march . the conditioning regimen was total body irradiation (tbi) and cyclophosphamide. during the -month follow-up period, we observed improvement in social interaction, communication, and behaviors (according to the childhood autism rating scale-cars). the second case is a -year-old boy with asd, asperger syndrome, who was diagnosis with all in september . he presented with bone marrow and testicular relapse in may and underwent a matched unrelated hsct in november . the conditioning regimen used was etoposide, atg and tbi. during the -month follow-up period, we observed improvement in social interaction, communication, and behaviors (according to cars). there is no treatment for asd thus every effort to minimize the symptoms are valuable. in both cases, social interaction was significantly increased, and the aggressive behaviors decreased. clinical cases have reported responses in autistic children receiving cord blood cd + cells. several incurable neurological disorders have shown benefits with cellular therapy. thus, autism should be explored as an indication. clinical studies are an immediate need to fully explore its potential in autism. disclosure of conflict of interest: none. conditioning is the initial phase of hematopoietic stem cell transplantation, based on high dose chemotherapy, combined or not with total body irradiation, aiming to eradicate the disease and prepare the environment of the bone marrow for the new cells. conditioning regimens can be characterized as myeloablative or non-myeloablative. during the period of conditioning and immunological reconstitution, signs and symptoms of the gastrointestinal tract are frequent, negatively influencing oral food intake, and may require the use of complementary nutritional therapies, aiming at an adequate caloric intake with the objective of avoiding decreasing in the nutritional status. the study aims to describe the association between the regiment intensity and the nutritional aspects during hospitalization of children and adolescents undergoing allogeneic hematopoietic stem cell transplantation (hsct) at a tertiary hospital. a retrospective study with medical records of patients undergoing allogeneic hsct, aged between and years of age (incomplete) between january and december . data were collected (regimen intensity, clinical signs of mucositis and nutritional therapies used) during the hospitalization and analyzed by the relative risk (rr). sixty-three patients were evaluated, being % male, with a median age of years. nineteen types of conditioning protocols were used. of these, % were high intensive regimen and % were low intensive regimen. the four most applied ( % of cases) were bucy (busulfan + cyclophosphamide) with and without atg (thymoglobulin), as well as cytb (cyclophosphamide+total body irradiation), also with and without atg. mucositis were observed in % of patients, being % grade and grade . the association was positive when analyzed the regimen intensity (myeloablative) with mucositis (rr = . ( . - . )) as well with the use of parenteral nutrition (rr = . ( . - . )). patients showed high prevalence of mucositis during hospitalization decreasing food intake, being necessary to use the parenteral nutrition. myeloablative regimen needed more nutritional therapy intervention when compared to non-mieloablative regiment. results demonstrate that an appropriate nutritional screening tool considering these aspects could help to intervene earlier maintaining an adequate nutritional status. autoimmune cytopenia (aic) is a potentially serious complication of hematopoietic stem cell transplantation (sct). autoimmune hemolytic anemia (aiha) is the most common aic, followed by immune thrombocytopenic purpura and autoimmune neutropenia. aic after sct is considered difficult to treat and associated with high morbidity and mortality. the aim of this cohort study is to evaluate incidence, outcome, potential risk factors and current treatment strategies and to explore the immune dysregulation predisposing to aic. the ebmt-promise database was accessed to identify all pediatric scts between and complicated by aic at our center. potential risk factors (i.e., age, gender, diagnosis, donor type, stem cell source, conditioning regimen) for aic after sct were assessed using univariate and multivariate cox regression analysis. in addition, we summarized treatment decisions of all aic patients. a nested matched case-control study was performed to search for possible biomarkers for aic. of consecutive scts, were complicated by the development of aic (cumulative incidence . %) at a median of months post-sct (figure) . aiha was the most common aic ( %), followed by combinations of two or more aics (evans syndrome, %). non-malignant disease, young age, alemtuzumab serotherapy pre-sct, non-tbi based conditioning regimen and cmv reactivation were associated with aic in univariate analyses. using multivariate cox regression analysis, non-malignant disease (hr . , p = . ), alemtuzumab use (hr . , p = . ) and cmv reactivation (hr . , p = . ) were independently associated with aic (figure) . for patients with cmv reactivation, diagnosis of aic was made at a median of months (iqr [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] ) after detection of maximum viral load. in our nested case-control analysis, serum levels of individual anti-and proinflammatory, and regulatory cytokines did not differ significantly between patients and controls. however, the cytokine profile of aic patients appeared to skew towards a more pronounced th response, compared to controls. firstline treatment, usually with prednisone and/or ivig, or a waitand-see approach led to resolution of aic in ( %) cases. second and subsequent-line therapies, often in combination with continuation of other treatments, consisted of rituximab (n = ), bortezomib (n = ) or sirolimus (n = ) and eventually led to resolution of aic in %, % and % of cases, respectively. overall survival of aic patients was %. in this retrospective cohort study, we identified cmv reactivation post-sct, alemtuzumab use and non-malignant disease as independently associated clinical risk factors for the development of aic. treatment with first-line therapy was mostly insufficient. for patients with severe aic, rituximab, bortezomib or sirolimus can be regarded as promising step-up therapies. figure (bkv) may cause polyomavirus-associated nephropathy or polyoma virus-associated hemorrhagic cystitis in bone marrow-transplant patients.we present patients with bk polyoma virus (bkv) ascociated hemorrhagic cystitis and patients with bk polyoma virus associated hemorrhagic cystitis and nephritis. between and , patients received an allogeneic bmt at acıbadem adana hospital pediatric bone marrow transplantation unit. patients occurred bkv associated hemorrhagic cystitis and nephritis. bkv was detected in the urine analysis and blood by pcr (polymerase chain reaction) in all patients. we presented patients with bkv infection, age ranging from to with a average of . years. they underwent allogeneic bmt due to thalassemia major ( patients), aplastic anemia ( patients) and acute lymphoblastic leukemia ( patients). the patients were treated with hydration, continuous bladder irrigation, ciprofloxacin, and weekly intravesical hyaluronic acid instillation for four weeks, and cidofovir. fourteen patients showed complete resolution of hematuria. one patient with refractory above these therapy also received hyperbaric oxygen and recombinant factor viia (rfviia, novoseven; novo nordisk,bagsvaerd, denmark). hemodialysis was performed in two patients who developed renal failure due to nephritis. bkv is ubiquitously present in the general population. reactivation of infection occurs under conditions of immunosuppression, particularly hsct or renal transplantation, and causes late-onset hc. bkv the management of bkv cystitis and nephritis sometimes may be very difficult and refractory all treatments, we presented our experience of bkv infection and management in transplanted patients in our center. patients with high-risk hematologic malignancies (hrhm) are among those in the highest risk group for developing invasive fungal disease (ifd), especially mold infections. allogenic hematopoietic stem cell transplantation (alsct), acute myeloid leukemia (aml), refractory and relapsed acute lymphoblastic leukemia (all), myelodysplastic syndromes and chronic extensive graft-versus-host disease are considered hrhm. ifd are a major cause of morbidity and mortality in these patients, however, the optimal strategy for antifungal p s prophylaxis in this population is not well defined yet. we performed a retrospective, observational study to investigate documented ifd during antifungal prophylaxis in children with hrhm who were admitted in our unit between and . demographic and clinical data were collected from patient's electronic medical records. all patients were treated with prophylactic voriconazole (vcz) according to our local practice. oral administration was preferred when available. vcz therapeutic drug monitoring (tdm) was not available in our center until june . breakthrough ifd was defined as occurrence of a proven or probable ifd according to eortc/ msg criteria while on vcz prophylaxis (⩾ days of treatment) or within days after discontinuation of prophylaxis. during the study period, hrhm patients were treated with prophylactic vcz in our unit. patients out of developed a breakthrough ifd. patient's demographic characteristics, main diagnosis and treatment are collected in table . initial and maintenance vcz doses are adjusted by weight in all patients except in patient- (adjusted according to vcz plasma level). adherence and tolerance to treatment was excellent in all patients. disclosure of conflict of interest: none. ( ) stable mixed chimerism (smc) when fluctuations of ac were o %; and ( ) mixed progressive chimerism (pmc) when ac were ⩾ %. - hscts performed in patients (pts) were included: children with a median age of . yrs (iqr . - . yrs) at diagnosis and . yrs (iqr - yrs) at hsct received one hsct ( . %), pts two hsct ( . %), and pts three hscts. primary diagnosis were bone marrow failures in pts ( . %), primary immunodeficiencies in ( . %), inborn errors of metabolism in ( . %) and haemoglobinopathies in ( . %). the donor was match related in ( %) procedures, match unrelated in ( %), and haploidentical in ( %); stem cell source was bone marrow in ( %), peripheral blood in ( %) and cord blood in ( %). conditioning regimen (cr) included busulfan in hscts ( . %), treosulfan in ( . %), while hscts ( . %) were conditioned with reduced intensity crs (including low dose of tbi in ); pt did not received cr. gvhd prophylaxis was based on csa/mtx (or mmf) in association with atg ( ) or alentuzumab ( ) ; recipients of tcrαβ/cd depleted haploidentical graft did not received post transplant immunosuppression. engraftment was observed in hscts ( after st , after nd and after rd hsct) after a median of day (iqr - days). acute gvhd occurred in hscts at risk ( %), and it was severe (gr. iii-iv) in ( %), chronic gvhd in ( %). at last follow-up (median . yrs), ( %) pts were alive, while pts are dead for infections (n = ), vod (n = ), c-gvhd (n = ) and vascular event (n = ). figure reported the evolution of chimerism over time. in our experience in children with non malignant disease, cc increased from % to % at subsequent analyses. % of pts with mc at st evaluation became cc, % remained smc, % evolved in pmc, and % rejected. only pts with cc at first time point rejected the graft. this study highlight the extreme variability of chimerism in the early post transplant course of children with non malignant disease and confirmed the relevance of performing serial analysis to monitor and, if necessary, improve graft function. naive t-cells identified by cd ra expression are believed to cause graft-versus-host-disease (gvhd), while cd ra-t-cells are memory cells that provide anti-infection and anti-tumoral effects. depleting cd ra+ naïve cells and retaining memory t-cells in the graft is a novel approach to haploidentical hsct for children. children with high risk leukemia ( aml, all) received cd ra-depleted haploidentical hsct following non-myeloablative conditioning. cell-selection performed on g-csf-mobilized peripheral blood. two cellular products obtained using clinimacs device, infused to each patient: a cd selection and a cd ra depletion from the cd negative fraction. product infused contained a median of . (range . - . ) x /kg cd + cells and a median of . (range . - ) × /kg of cd + cells in the cd -selected s graft. the second product was the cd ra depletion, cd ra +/kg was a median of . × /kg (range - × /kg) and a median . (range . - . ) depletion log of cd ra + cells. median dose of cd ro+ cells (memory t-cells) infused was (range . - ) × /kg. seventeen patients achieved neutrophil engraftment at median of days (range - ) post-transplant. one patient could not achieve engraftment, died at day + due to sepsis. two patients presented secondary graft failure (day + and + ), both received a second hsct. three patients developed agvhd grade ii with gastrointestinal tract involvement, all steroids responsive. three patients presented clinical features of cgvhd. patients have an extensive skin involvement, with hepatic findings in one and pulmonary affection in other, at day + , + and + post. ten of patients ( . %) remain alive in remission with median follow-up (range - ) days post-transplant. eight patients died, due progression at day + , + , + ( presented positive minimal residual disease at hsct), due to infectious complications (days + , + , + , + ) and due to cardiogenic shock at day + . four patients relapsed, of them died afterward with progressive disease. t-cells led immune recovery, achieved values higher than , , and cells/mcl at day , , and respectively. most of t cells were cd +cd ra-(median of , and × /mm respectively on day + , + and + ) and cd +cd ra-t cells (median of , and × /mm respectively on day + , + and + ), while cd + ra+ and cd + ra+ cells remained low recapitulating the cd ra depleted graft composition. six patients presented cytomegalovirus reactivation, one progressed to cmv disease. five patients with hhv- encephalitis. probable aspergillosis in patient (aml-m with secondary graft failure) at day + after second hcst. two cases of toxoplasmosis ( cns, pulmonary). cd ra-depleted haplo-hct showed acceptable tolerability with rapid and sustained engraftment as well as a full donor chimerism, minimal risk of acute gvhd and accelerated inmunologic reconstitution. to note the high incidence of hhv- encephalitis seen. disclosure of conflict of interest: none. collection of peripheral blood stem cells in teenager sibling donors: a single center experience c carvalho , f amado, f bordalo, s ferreira, s lopes, c pinho and s roncon serviço de terapia celular, instituto português de oncologia do porto francisco gentil, epe human leukocyte antigen (hla) compatibility is important in allogeneic haematopoietic stem cell transplantation in order to reduce post-transplant complications; however, siblings only present a % chance of hla-match with the patient. the well-known advantages and the low risk of complications associated to peripheral blood stem cells (pbsc) collected by apheresis made this procedure the first option in teenagers. the aim of this retrospective study was to analyse and characterize the paediatric sibling pbsc donor population assuring safety during the collection procedure, providing a high-quality product and accomplishing patient needs. we consulted the clinical files of donors under years old since - ; a database in excel ® was created to register population characteristics, collection parameters and graft requirements. the informed consent was obtained from parents before procedure. the leukapheresis were performed with a cobe spectra system; since , we use a spectra optia apheresis system. the donor/patient weight ratio (proposed by styczynski et al.) was determined for each pair. the collection was programed based on clinical and analytical donor's features as well as transplant requirements. the analytical assays were done by a certified laboratory. we performed pbsc apheresis in healthy donors, females and males ( table ) . all of them started on the th day after mobilization with granulocyte colony-stimulating factor (g-csf) administered subcutaneously, bidaily. the weight ratio was o in eight situations. most of donations were performed by peripheral vein; a central venous catheter (cvc) was placed into a femoral vein in six adolescents. a median of ( ) ( ) ( ) blood volemias were processed during ( - ) minutes; the anticoagulation used was citrate+heparin (ratio : ). in general, one-collection day was enough to obtain the number of cd + cells required; six donors had to perform a nd collection. in cases, we cryopreserved the exceeding cells after graft infusion. the procedure was well tolerated, with only adverse reactions registered (one hematoma in the puncture local; one paraesthesia due to hypocalcaemia induced by citrate). no blood products were used after the procedure or needed for the priming of the extracorporeal circuit. so far, no serious long-term adverse events were observed. table . median (range) of donors and leukapheresis products data. our long lasting experience shows that pbsc collection in the teenage population is safe and feasible, allowing us to obtain a high-quality product for the patients. there were no adverse events associated with the g-csf mobilization or cvc placement which is different from the experience of other groups. we recognize that leukapheresis by peripheral vein is a lengthy procedure but no complaint was reported to the collection team. [p ] disclosure of conflict of interest: none. correspondence between clinical and hystological grading of gastro-intestinal grading acute graft versus host disease in children m faraci , a rizzo , p gandullia , s arrigo , , a barabino , e lanino , s giardino and c coccia hematopoetic stem cell transplant unit, institute g gaslini, genoa, italy; pediatric department, institute g gaslini, genoa, italy; gastroenterology and digestive endoscopy unit, institute g gaslini, genoa, italy; gastroenterology and digestive endoscopy unit, institute g gaslini, genova and department of pathology, institute g gaslini, genoa, italy diagnosis of gastro-intestinal acute graft versus host disease (gi-agvhd) is frequently confirmed by apoptosis findings on mucosal biopsies. aims of this single center retrospective study is to evaluate the correlation between clinical and histological grading of gi-agvhd in children undergoing allogeneic haematopoietic stem cell transplantation (allo-hsct), and to describe histological findings obtained by gi endoscopies in order to evaluate usefulness in the diagnosis of gi-agvhd. allo-hscts were performed in our department between january and december . gi biopsies were performed in pts ( . %) because of suspected gi-agvhd. pts were transplanted for malignant ( . %) and for not malignant diseases. the median age at hsct was . years ( . - . ). pts ( %) received myeloablative and ( %) reduced intensity conditioning regimen. pts ( . %) received an unrelated donor (ud), pts a related donor (rd) ( . %), and an haploidentical donor ( . %). at onset of diarrhea, microbiological examinations of stool were performed and pcr research for cmv, adenovirus, hhv , ebv were evaluated in blood and in mucosal biopsies. mucosal biopsies were obtained with esophago-gastro-duodenoscopy in pts ( . %),esophago-gastro-duodeno-colonscopy in ( . %), pancolonscopy in ( . %), flexible sigmoidoscopy in ( . %), and rectal suction biopsy in pts ( . %). all mucosal biopsies, except in case of rectal suction, were obtained under sedation. the interval between mucosal biopsies and onset of gi acute symptoms was days (from − to days). biopsies were taken from different sites in the gi tract, were stained using hematoxylin-eosin and evaluated using histological grading of agvhd. in these pts the maximum grade of agvhd was: grade in one ( %), grade in ( %), and grade in pts ( %). at time of histological evaluation, diarrhea was the most common gi symptom ( . %); children had also cutaneous agvhd and hepatic agvhd. pcr-cmv was positive in mucosal biopsies obtained with pancolonscopy, pcr-adenovirus in other obtained with upper and pancolonscopy, pcr-hhv in rectal biopsies, and pcr-ebv in one with upper and pancoloscopy. the comparison between clinical and histology grading of gi-agvhd is shown in table . mucosal biopsies were positive in / pts evaluated with esophago-gastroduodenoscopy ( %) (grade agvhd), in / pts undergone esophago-gastro-duodeno-colonscopy (grade in and grade in ), in / ( %) who received a pancolonscopy (grade in , grade in , grade in ), and / ( %) of rectal biopsy obtained by sigmoidoscopy or rectal suction biopsy (grade in , grade in , grade in , and grade in ). one patient developed duodenal intraparietal hematoma after upper endoscopy. in our experience, we did not demonstrated a overall correlation between clinical and histological grading of agvhd showing that hystological examinations underestimated the grade mild or moderate of agvhd. we confirmed , that rectal biopsies represent to be more effective and safe diagnostic method for the confirm of diagnosis of gi-agi. during the past few decades, hematopoietic cell transplantation (hct) as a treatment modality for primary immunodeficiencies (pid) has undergone remarkable advancement mainly due to better availability of alternate donors resulting in increase in not just matched unrelated donor (mud) but also increased haploidentical (haplo) and cord blood transplants (cbt). additionally, refinement of the conditioning regimens and better graft versus host prophylaxis have presumably led to better survival outcomes. however, a literature gap is identified in evaluation of these outcomes in general with respect to donor and conditioning regimens. we conducted a systematic review by performing a comprehensive search of the pubmed and ovid library from its inception to august . mesh terms included 'primary immunodeficiency (immunodeficiencies)', 'stem cell transplant', 'bone marrow transplant' and 'hematopoietic cell transplant'. all pid studies which used hct as a treatment modality were included. experimental cellular therapies were excluded. both cellular immunodeficiencies (e.g. scid, was, a-t), and innate immunity disorders (e.g. ifngr, cgd) were included in the search. reviews, case reports, meta-analysis and non-english language articles were excluded from our electronic search. publication bias was excluded by performing a methodological search of unpublished conference abstracts from the annual meetings of cis, aspho, asbmt, ebmt, and siop from to . the data were analyzed considering the outcomes -overall survival and gvhd. studies fulfilled the strict selection criteria for the electronic search comprising of pid patients. in majority of the hcts, a myeloablative conditioning regimen (mac) was utilized ( % of the evaluable) but a shift towards more reduced intensity conditioning (ric) was observed in the later years. cbts were identified. % of patients developed some degree of acute gvhd, whereas chronic gvhd was identified in % of the patients. total number of haplos was . overall survival was found to be % post-hct. a meta-analysis could not be performed due to the heterogeneity of both the predictor variable data (combined stem cell sources were also used for hct) and due to the extremely small number of the patients when categorized in subgroups (e.g. for omenn syndrome, rag deficiencies). this is the largest study of hcts in pid, and we observe that alternate donor hcts have increased significantly over the past decade for the treatment of pid. while the incidence of chronic gvhd was low, acute gvhd still remains a problem in about a third of the pid patients transplanted. disclosure of conflict of interest: none. hepatic veno-occlusive disease (vod) is a common and serious complication of hemotopoietic stem cell transplantation (hsct) in children. we aimed to assess prospectively the use of prophylactic defibrotide in pediatric patients undergoing hsct. in this study, patients who underwent hsct were given defibrotide prophylaxis as mg/kg per day in four divided intravenous infusions over h, starting on the same day as the pretransplantation conditioning regimen. the mean duration of use of defibrotide is days as a prophylaxis. in this study, patients were recruited, male patients and female patients, with the average of . years, range - ; % infants, % children and % adolescent. there were patients with thalassemia major, patients with leukemia, patients with aplastic anemia, one patient with diamond blackfan anemia, two patients with congenitale dyserythropoetic anemia, one patient with osteopetrosis, four patients with famial hemophagocytic lymphohistiocytosis, two patienrs with severe immune deficiency and one patient with kostman syndrome. all transplants were allogeneic. no serious side effects were seen. in eight patients developed clinical vod (seattle criteria). in these patients, defibrotide dose was increased to a treatment dose of - mg/kg per day. one infant patient with kostman syndrome died due to hepatic and pulmonary veno-occlusive disease. after months of follow up, patients who developed vod are being well and no patient have transplant related complications. hepatic veno-occlusive disease, which is caused by hepatocyte and sinusoidal vessel endothelium damage, can ocur early after hsct, and in its severe form, may lead to liver faillure, hepatorenal syndrome, portal hypertension, and eventually death from multiorgan faillure. in this prospective study, we demonstrated that the use of defibrotide is safe and effective in preventing and treating vod in pediatric patients at high risk. immune reconstitution (ir) is critical for clinical outcome after allogeneic hematopoietic stem cell transplantation (hsct). host and proceeding-related factors affect the ir dynamics and survival. isolated ir parameters are commonly correlated and proposed to predict clinical outcomes after hsct, but these approaches only confer prognostic value at single time points or for single markers. we aim to demonstrate an appropriate methodology to assess the capability of combined serial measurements of lymphocyte subsets to reflect the impact of infections on ir after paediatric hsct. retrospective data of patients receiving a first hsct for any indication with any cell source in the paediatric hsct program from to were included. to characterize the kinetics of immune reconstitution, cd +, cd +, cd + t-cells, b-cells, nk-cells and their naive and memory subsets were measured and analysed at various time points at years post-hsct to stablish a joint model for the evolution of cell subpopulations. slope per month (cellular increase or decrease) of each lymphocyte subsets were calculated and compared with clinical outcomes and cumulative risk of infections. a total of children (range from - y.o. median y.o.) were included, with cb (n = ) pb (n = ) and bm (n = ) as cell sources. the cumulative incidences after early period were % for viral infections (ebv %, cmv %, bk %, adv %) and % for bacterial infections. data on ir were available for %, of the diseasefree survivors. in a exploratory multivariate analysis we detected mainly differences in cd +, cd +cd ro+ memory and nk cells at year after hsct, with dependent tendency according on the cell source and hla compatibility. analysis of the slope tendency patterns were stablished for the analysis of the impact of infections in the ir. delay in cd + and cd +ra+ appearance (mean slope/m = − . % and − . % respectively) remarks the ir profile for bacterial infections, and delayed in nk, cd and cd ro+ (− . %, − . %, − % respectively) for overall viral infections. additional correlations allow differences in ebv (cd +ra+ high mean slope/m = . %), cmv (delayed in cd ro+ slope/m = %), and bk infection (cd +ra + plus cd ro+ and nk high mean slope/m = . %, % %). understanding the dynamics of reconstitution by integrating information from the monitoring of lymphocyte subpopulations allows the establishment of kinetic profiles that may help to evaluate the risk of infections and adjust infection prophylaxis in the follow-up of transplanted patients. mortality rate in hsct patients admitted to intensive care unit (icu) is still as high as % to %. this rate increases when respiratory complications progress to acute respiratory failure (arf) requiring mechanical ventilation (mv). the aim of this study was to determine the feasibility and effectiveness of early continuous positive airway pressure (cpap) delivered in a pediatric hematology-oncology ward to prevent occurrence of arf requiring mv. we retrospectively analysed children treated with cpap in our pediatric hematology-oncology ward between october and october . thirty-two patients received cpap delivered with helmet during the study period. data were available for patients, males and females, median age years [range - ]. eighteen patients underwent allogenic hsct: from sibling donor, from matched unrelated donor, from haploidentical family donor, from cord blood unit. seven patients had a malignant disease: all, aml, ewing sarcoma. infectious pneumonia was the main cause of arf in / patients ( . %): viral pneumonitis ( rhinovirus, parainfluenzae virus, respiratory syncitial virus and cmv). five patients had proven/ probable invasive fungal infection according to eortc criteria ( aspergillosis and mucormycoses). other causative agents were pneumocystis jiroveci ( ), bacillus of calmette and guerein ( ), toxoplasma gondii ( ) and st. mitis ( ) . non infective causes of arf were acute transfusion related lung injury ( ), hemorragic alveolitis ( ), cryptogenic organizing pneumonia ( ), tumor lysis syndrome ( ), and alveolar oedema due to renal failure ( ). according to chest imaging, / patients ( %) presented with pulmonary consolidations, while % had both interstitial infiltrates and pulmonary consolidations. at baseline median neutrophil count was . × /μl (range - . × /μl), mean heart rate bpm, pulsiossimetry saturation in room air %. h-cpap was applied in / patient with a curative aim, in / patients as palliative support to reduce respiratory distress. median positive pressure delivered was cmh o ( - cmh o), median fio was % ( - %). h-cpap was applied for a median of days . no patient failed h-cpap because of agitation or adverse events (skin breakdown, conjunctivitis, gastric distension or epistaxis). ten patients were transferred to icu ( . %), / because of hsct complications. median icu stay was . days ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) . only patients required mechanical ventilation, in cases associated to ecmo. nether psao in room air (p . ci %) nor cpap level (p . ci %) correlated with the need of icu admission. patients requiring higher fio during cpap demonstrated a not statistically significant trend to higher icu admission rate (p = . ).there was a higher rate of mv in patients with higher cpap fio level (p = . ). mv prolonged icu stay (p . ). cumulative mortality was . % ( / ); only patient died in icu ( %), because of post hsct parainfluenza virus pneumonitis requiring ecmo. helmet cpap delivery in pediatric hsct ward is feasible and safe, both for curative and palliative aim. if applied early, cpap could reduce icu admission rate for mv and icu mortality. veno occlusive disease (vod) and graft versus host disease (gvhd) are both dreadful complications of hematopoietic stem cell transplantation (hsct). although they have different clinical signs, it is suggested that they share similar pathophysiological pathway. defibrotide is used in the treatment of vod for a long time but it is very less known about its effect on gvhd. in this study, we analyzed a 'high risk' patient population in pediatric hsct to show the effect of defibrotide on acute gvhd. between june -august totally 'high risk' pediatric allogenic hsct procedures were enrolled in this study. 'high risk' definition involves busulphan/ melphalan usage in conditioning regimen, second myeloablative hsct, pre-existing liver disease, allogenic hsct for leukemia with second relapse and diagnosis of hemophagocytic lymphohistiocytosis (hlh) or osteopetrosis. defibrotide prophylaxis group (n = ) received mg/kg/day per day and continued for at least days after transplantation. the control group (n = ) received only continuous infusion of low-dose heparin until days after transplantation. for the comparison between groups, the fisher's exact test was used. all analyses were performed using spss and p-value of . was considered statistically significant. we analyzed totally hsct procedures with different diagnosis; beta thalassemia major, leukemia, hlh, primary immunodeficiencies, osteopetrosis, fanconi aplastic anemia (faa), myelodysplastic syndrome, neuroblastoma, congenital amegakaryocytic thrombocytopenia, krabbe disease, aplastic anemia, hypereosinophilic syndrome and sickle cell disease. all the procedures meet the 'high risk' definition; most of them (n = ) have busulphan for conditioning, also there are hlh and osteopetrosis patients, neuroblastoma patients had the second myeloablative regimen, faa and aplastic anemia patients had pre-existing liver disease, and of the leukemias had beyond second relapse. the mean age was . years old ( . - . ), hsct performed from match sibling donor (msd) ( %), hsct from match family donor (mfd) ( %), hsct from match unrelated donor (mud) ( %) and hsct from haploidentical mother ( %). we especially focused on gvhd and vod. totally vod cases ( %) in these hsct procedures were detected. only two of them detected in the prophlaxis group ( %) and cases in the control group ( %). there were cmv reactivation cases detected in hsct procedures ( %). in the prophlaxis group there were cases ( %) and in the control group cases ( %). we detected acute grade i-iv gvhd cases in hsct procedures ( %). only of them were in the prophlaxis group ( %) and cases were in control group ( %). the prophlaxis group's agvhd ratio was significantly lower than the control group (p = . ). defibrotide for vod prophylaxis is confirmed by several studies, but its benefits for agvhd is not clear. in this study, we show the significant effect of defibrotide on agvhd. we speculate that the protective effect of defibrotide on both vod and agvhd could be explained by the similar pathophysiology of these complications. we need larger studies on the pathophysiological pathways, then we could invent more effective interventions. disclosure of conflict of interest: none. conventional extracorporeal photopheresis (ecp) has proven efficient for the treatment of graft-versus-host-disease (gvhd) but is limited to patients with sufficient body weight. a mini buffy coat ecp (mini-ecp) 'off line' technique that allows treatment of small children has been developed, using various methods for mononuclear cell (mnc) separation from whole blood. we present treatment of low body weight child with mini-ecp 'off line' technique using sepax system for mononuclear cell (mnc) selection and macogenic irradiator. a toddler with juvenile myelomonocytic leukemia (jmml) developed acute gvhd, after a matched unrelated stem cell transplantation (sct) performed at the age of months. acute gvhd of the skin occurred three months after sct and responded to high dose steroids, but recurred six months after sct (biopsy of the skin confirmed acute gvhd) together with gvhd of the liver. because of the resistance to steroids and cyclosporine, mini-ecp was introduced as therapy. the patient weighed kilograms. blood was collected from tunneled central venous catheter, and collected volume was replaced with saline infusion. the cord blood collection bag (macopharma, france), which contains ml citrate phosphate dextrose (cpd) anticoagulant solution was used for whole blood collection. whole blood was processed using sepax system separator (biosafe, switzerland), and final volume of buffy coat was set on ml. extracted buffy coat was transferred into the uv-a illumination eva bag (macopharma, france) and diluted with saline solution up to ml. methoxypsoralen (gerot, austria) was injected directly into the uv-a illumination bag, and cells were irradiated by the uv-a illumination device macogenic (macopharma, france). irradiated cells and autologous residual blood were reinfused back to the patient. during the whole procedure patient's vital signs were monitored. ecp procedures were performed times per week for weeks, followed by times per week at weeks intervals for months. in month period mini-ecp procedures were performed. median of collected whole blood was ml (range - ). median of total nucleated cell (tnc), and mononuclear cell recovery after sepax separation were . % (range . - ), and . % (range - ), respectively. median of hematocrit in final irradiated product was % (range . - %). patient was reinfused with median of . (range . - . ) tnc × /kg bw, and median of . (range . - . ) lymphocyte × /kg bw. after one month of ecp together with steroids and cyclosporine, gvhd of the skin improved, and the steroids were gradually weaned, with no recurrence. gvhd of the liver showed no improvement, and other therapies had to be introduced, but without steroids. for the first time in croatia, mini-ecp was performed in a child with gvhd, in whom conventional ecp could not be used because of insufficient body weight. mnc separation using automated closed system sepax separator has proven efficient and safe. mini-ecp treatment was continued for three months, without technical difficulties. positive effect was experienced concerning the skin gvhd, but not the liver gvhd. after the first experience in our country, in future we plan to use this technique for low-weight patients or patients with contraindications for apheresis, which are in need of second-or third-line therapy for gvhd. disclosure of conflict of interest: none. gonadal failure represents one of the late effects of haematopoietic cell transplantation (hsct) with a negative impact on quality of life in young patients (pts) undergoing hsct , . the aim of this retrospective multicentre ebmt study was to assess gonadal function in untreated pts undergoing allogeneic hsct between to years (yrs) of age, after a preparative regimen with busulphan (bu) or treosulfan (treo). eighty-seven pts ( females, males) were reported from out of contacted ebmt centers: / ( %) received allogeneic hsct during pre-pubertal and / ( %) in pubertal phase. of the pts, ( . %) received bu in myeloablative dose [ pre-pubertal, (median age of . yrs) and pubertal, (median age of . yrs)] and pts ( . %) received treo ( in pre-pubertal and in pubertal period). underlying diseases were primary immunodeficiency ( . %), chronic myeloid leukemia ( . %), myelodisplastic syndrome ( . %), familial haemophagocytic lymphohistiocytosis ( . %) and shwachman-diamond syndrome ( . %). / of prepubertal pts ( %) developed spontaneous puberty ( . % in the bu group and % in treo group). / ( %) females undergoing hsct during puberty completed their pubertal development ( . % in bu group and % in treo group). none of females ( / ) with bu during pre-pubertal phase developed spontaneous menarche (sm), while . %( / ) of females who received bu in pubertal period had sm. all females (n = ) treated with treo during pubertal phase had sm ( %). for both conditioning regimens, the . % ( / ) s of females treated during the puberty experienced sm. among the remaining females (for pts the information is missing) who did not developed sm, received hrt . yrs after hsct and of them had ovarian recovery after a median of . yrs from hsct ( . - . ). the median age at last follow up was . and . yrs in bu and treo pre-pubertal group, and . and . yrs in bu and treo pubertal group respectively. in the pubertal group, females ( . %) are still receiving hormonal replacement therapy (hrt) ( in the bu group and in treo group). pts ( . %) had spontaneous pregnancy. no problems in newborns are reported. sperm analysis was performed in . % of pubertal pts ( / ) of males, and % (n = / treated with bu) were azoospermic (data regarding pts were missing). the sperm analysis was repeated in half of the males. until now no paternity was reported. in this experience, the pubertal development in pts who received treo (n = ) was normal, and in the bu group the majority of females ( %) had normal puberty. the rate of sm is higher ( %) in females after treo than bu ( %). the hrt is ongoing at last follow-up in % of females treated with bu and in % of those who received treo. our data suggests that treo may have a better outcome than bu in young girls receiving allogeneic hsct and larger studies are warranted. male patients require longer follow-up. prevention in patients transplanted from partially matched donors. we report the single centre experience in haploidentical sct. in years - in the department of pediatric bmt, oncology and hematology at wroclaw medical university, children underwent sct from partially matched, haploidentical parental donors. graft manipulation in patients consisted of cd sel, in patients the cd immunomagnetic depletion (tcd-cd ) was performed, and in -tcr alpha-beta depletion (tcd-ab). we analysed the impact of type of manipulation procedure, conditioning regimen, demographic factors, and kir genotype on survival and probability of neutrophil recovery. the probability of engraftment and neutrophil recovery was % vs % in cd sel group (p = ns). probability of year overall survival in the tcd group was similar to the cd sel group ( % vs %, p = ns). in the tcd patients, neither use of busulfan vs treosulfan, nor kir genotype, nor donor sex had noticeable impact on sct result and survival. patients transplanted after tcd due to non-malignant disease had higher survival probability, than those with malignancies ( % vs %, p = . ). the trm in tcd patients was reduced in comparison to cd sel ( vs %, p = . ). the trm after tcd resulted mostly from severe viral infections in tcd-cd patients. in / tcd patients spontaneous acute, skin (stage ) gvhd was diagnosed and successfully treated. two patients received unmanipulated donor lymphocyte infusions (dli) and developed severe acute steroid-resistant (grade ) gvhd, in one of them with fatal outcome. tcd methods are superior to cd sel due to significant reduction in treatment related mortality. haploidentical sct after tcd can result in durable engraftment, but warrants intensive post-transplant monitoring for infectious complications and cautious approach to dli therapy. disclosure of conflict of interest: none. median of days for neutrophils in both groups, for platelets ( in ptcy, in αβ+cd +/cd +depleted, p . ). donor chimerism was complete in patients ( . %). in αβ +cd +/cd +depleted group, patient rejected ( . %: primary and secondary reject, , , and days after haplo, respectively) and were rescued with a second transplant. seven patients ( %) developed acute (a-) gvhd in ptcy group (grade - in ; grade - in ), compared to one ( . %: grade ) in αβ+cd +depleted group (p . ). among patients at risk, out of in ptcy group developed chronic (c-) gvhd ( . %: score- , overlap, score- ), compared to / patients in αβ+cd +/cd +depleted group (p . ). the cumulative risk of cmv-reactivation was % and % in ptcy and αβ+cd +/cd +depleted groups, respectively (p . ). t-cell reconstitution was significantly different in the two groups,with a median absolute number of cd + +cd -and γδ+cd + higher in αβ+cd +/cd +depleted group on day + (p . ) and a median number of cd +cd + higher in ptcy group on day+ (p . ). length of hospitalization was shorter in the αβ+cd +/cd +depleted group, with a median time from haplo to discharge of days compared to days in the ptcy group (p . ). some children have not donor and an urgent need to proceed to transplantation because of disease status. we reviewed the role of haploidentical transplantation in children and report our single center experience. ten children were transplanted from haploidentical family members donors (median age: . years). we performed alfa beta t cell depleted transplantation in three patients and unmanipulated bone marrow transplantation with posttransplant cyclophosphamide in seven patients. the diagnosis were eight high risk leukemias (three all and five aml) and two severe aplastic anemia. patients were myeloablative conditioned with cyclophosphamide, fludarabine and total body irradiation in aplastic anemia received alfa beta t cell depleted grafts with a median cd cell dose of . × /kg (range: . - . ) and busulphan, cyclophosphamide in high risk leukemias received unmanipulated bone marrow grafts with posttransplant cyclophosphamide in rd and th day of posttransplant with a median cd cell dose of . × /kg (range: . - . ). median follow up of our patients months. six of patients are alive and in disease free follow up. four patients were relapsed and dead median . months of transplantation. the rate of relapse was % for leukemia patients in remission and % for patients with active disease. myeloablative conditioning regimen followed by haploidentical bone marrow transplantation with posttransplant cyclophosphamide may be an option in high risk leukemia patients need urgent transplantation because of desease status who have not donor. table . all patients received hd-cy mg/kg on d+ and d + . cyclosporine a mg/kg/d i.v., then mg/kg/d p.o. adjusting for blood levels - ng/ml and mycophenolate mofetil mg/kg times daily po were started on d+ . mmf was discontinuated on d+ , csa-after d+ . all pts received anti-microbial prophylaxis for bacteria, fungal, herpes infection and pneumocystis according to institutional practices. analysis for donor chimerism and mrd were performed at d+ , + , + , + . pts, donors and stem-cell harvest characteristics are described in table . pts had high risk hematological malignancies, and relapsed after auto-sct neuroblastoma (hr-nb). pt was transplanted in st cr (aml m ) and others in nd cr. pts had full engraftment (neutrophil engraftment at , and days). pt (hr-nb) was concerned as a primary failure for achieving neutrophil and platelet engraftment by d+ , despite of complete donor chimerism in bone marrow. he was transplanted additionally with the same donor at d+ after st transplant. pt died before engraftment at d+ (fulminant ps. aeruginosa-sepsis). pts remain alive in cr ( ndcr-aml and st cr-m aml) with follow-up of and days ( / / ) without cgvhd with complete donor chimerism. pts relapsed after d+ ( were transplanted in nd cr-flt aml and nd cr-nb) and died. pt died because of infectious complication at d+ (transplanted in d cr-all). / pts had grade acute gvhd. hla-haploidentical hsct with post-transplant t-cell in vivo repletion grafts by using hd-cy is feasible and effective in children with hr-haematological malignancies. [p ] who were match unrelated donor. thalassemic reconstitution occurred in three patients. acute graft-versus-host disease (gvhd) of grade ii-iv occurred in % and chronic gvhd in %. acute and chronic gvhd were seen more frequently in patients with class - compared to class . mortality rate was also higher in these groups. seven patients died. one patient died on post-transplant day due to intracranial bleeding. the other patients with chronic gvhd died between and days, on average days post-transplant. these data suggest that allogeneic bmt remains an important treatment option for children with beta-thalassaemia major, particularly when compliance with iron chelation is poor. the society to support children suffering from cancer, also known as mahak, was set up in as a non-governmental and non-profit organization. in the past two decades, the organization has attracted a vast public support and fulfilled a great part of its mission which is to support children with cancer, reduce the child mortality rate and create an appropriate environment that empowers families who have children with cancer. pediatric stem cell transplant also is used to treat many types of conditions affecting children and adolescent, including cancer and certain hematologic, immune reconstitution inflammatory syndrome (iris) is a clinical condition emerging after immune recovery of an immunocompromised status, mostly after the initiation antiretroviral therapy (art) in human immunodeficiency virus (hiv) infected patients, but also in several other settings, such as the recovery from the severe combined immunodeficiency (scid) status after hematopoietic stem cell transplantation (hsct). herein, we report a patient transplanted for scid who developed iris for two times, namely shortly after transplantation and after donor lymphocyte infusion (dli) ( table ) in our patient, t cells passing from the donor probably contributed to the immediate post-transplant increase in the size of granulomas. this inflammatory response waned after the institution of immunosuppressive and methylprednisolone therapy. however, immunosuppressives were stopped due to lowered chimerism at follow-up, and the inflammatory response re-appeared after administering stem cell support containing a large amount of t cell from the donor for dli purpose. although the mechanism by which dli results in clinical responses is unclear, it is presumed to be a t cellmediated process. several studies have been performed to strengthen our understanding of the immunopathogenesis of iris. while some of those studies put forth t cell-associated causes, others implicated cytokines and non-t cells. the reaction that developed in our patient is suggestive of t cellassociated causes. immune reconstitution inflammatory syndrome remains a poorly understood entity. the dli procedure in our case provides a unique clue supporting a t cell mediated process. pediatric transplant teams need to be s aware of the previous iris phenomenon of bcg-adenitis while making the decision of dlis. [p ] disclosure of conflict of interest: none. pediatric patients treated with a hematopoietic stem cell transplantation (hsct) often suffer from late side effects caused by the treatment. the aim of this study is to investigate the late effects of a hsct on dental development. in addition, patients and parents awareness on this topic was investigated. young adults treated and followed at the ghent university hospital who were under the age of y at the time of hsct were examined clinically and radiographically (planmeca promax d). transplants ( autologous/ allogeneic) were done for malignant disease in pts. eight patients received a hsct for a non-malignant disease. twelve patients underwent a conditioning regimen with total body irradiation (tbi), patients with busulfan and patients with other chemotherapeutic agents. patients were o y, patients were - years and patients were years at hsct. every patient was evaluated on dental agenesis, microdontia and rootcrown ratio. patients and their parents were asked about their knowledge and interest for dental screening at the follow-up clinic using a questionnaire. overall, the prevalence of agenesis and microdontia of one or more dental elements is respectively . % and . % in our study population. . % of patients have a strongly aberrant root-crown ratio of at least one element. patients treated o years of age show significantly more microdontia ( . %; po . ) as well as agenesis ( . %; p o . ) compared to patients treated at an older age. the first premolar of the mandible is the most vulnerable element for agenesis as well as for microdontia. more microdontia is found in patients treated with a busulfan conditioning regimen compared to the other conditioning regimens ( . % versus %). patients older than years, treated with busulfan have statististically more microdontia compared to patients y treated with tbi conditioning regimen (p = . ). there was no difference of the conditioning regimens on agenesis nor on root-crown ratio. almost all patients/parents find it important to receive information about the dental late effects of a hsct and are interested in dental screening at the follow-up clinic. treatment with hsct has an explicit negative impact on dental development. the degree of this effect depends on age at hsct and used conditioningregimen. dental follow-up of these patients is essential and should be incorporated in the follow-up program. disclosure of conflict of interest: none. importance of body composition in the outcome of hematopoietic stem cell transplantation in elderly patients l koch , n hamerschlak , r garcia , c prado , c silva and a pereira hospital israelita albert einstein the loss of muscular mass is a well recognized cause of the decline in muscle strength and functionality that accompany the aging process. in , irwin rosenberg proposed the term 'sarcopenia' to describe the decline in muscle mass associated with aging. changes in body composition after hematopoietic stem cell transplantation (hsct) have been the subject of previous studies. immunosuppressive therapy and corticosteroids are known to alter skeletal muscle metabolism. infections and graft-versus-host disease (gvhd) that can occur after hsct may also affect body weight and composition. therefore, both the treatment and complications after hsct exert large negative effects on lean muscle mass, especially in elderly patients. patients with hematologic malignancies are usually well nourished before undergoing hsct. objective: the aim of this study is to determine in an elderly population whether parameters of body composition could be correlated to outcomes after hsct. we performed a retrospective longitudinal study through review of medical records of patients ⩾ years old undergoing hsct at hospital israelita albert s einstein, from to , that were subject to tomography scans (cts) in a period ranging from days before and days after hsct. table . there were no differences between groups with respect to age, gender, diagnosis, stage of disease, and source of stem cells. in ly patients, the quantity of peripheral cd + cell dose (× /kg) infused was different between groups (group ly-ct the incidence of nf was significantly higher in group mm-g ( ( . %) vs ( . %); p = . ). no differences were observed in the incidence and severity of mucositis, first day and duration of fever, documented bacterial infections or readmission rate between mm patients groups. this study suggests that in at home asct, the use of piperacillintazobactam prophylaxis significantly reduces the incidence of neutropenic fever and hospital readmission in patients with ly, and also that no administration of g-csf in mm patients reduces significantly the incidence of neutropenic fever. disclosure of conflict of interest: none. [p ] allogeneic stem cell transplantation (sct) has been recognized as a curative treatment for patients with wiskott-aldrich syndrome (was). in sct for was, myeloaberative conditioning (mac) has been indicated to avoid a mixed chimera. however, risk factors for a mixed chimera in patients with was who have received sct have not been evaluated. here, we analyzed the outcomes of sct and risk factors for a mixed chimera in patients with was who underwent sct in japan since . we reviewed medical records of consecutive was patients who received sct since january who were registered with the japan society for hematopoietic cell transplantation. the age of the patients at transplantation ranged from months to years, and the mean age was . years. the origin of the stem cells was related bone marrow (bm) or peripheral blood stem cells (pbsc), unrelated bm or pbsc, and unrelated cord blood (cb) for , and patients, respectively. a preparative conditioning regimen consisting of mac was provided to patients, and reduced-intensity conditioning was provided to patients. fifty-one patients received prophylaxis against graft-versus-host disease (gvhd) with cyclosporine in combination with methotrexate (mtx) or a steroid, and patients received tacrolimus (tac) with mtx or a steroid. chimerism analysis had been performed in patients. neutrophil engraftment was achieved in patients ( . %). the engraftment rate was significantly higher in patients who received tac for gvhd prophylaxis, (p = . ) overall survival rate was significantly higher in patients with complete chimerism than in patients with mixed chimerism ( . ± . % and . ± . %, respectively, p = . ), though there was no significant difference in stem cell sources. using multivariate analysis, the rate of complete chimerism in patients who received mac including cyclophosphamide (cy) at more than mg/kg was significantly higher (p = . ) than the other conditioning. not only patients with mixed chimerism but also patients with complete chimerism were complicated with auto-immune diseases. in this study, achievement of complete chimerism after sct was important for survival in patients with was. we found that patients who underwent mac including cy at more than mg/kg had a higher rate of complete chimerism. we also found a higher neutrophil engraftment rate in patients who received tac for gvhd prophylaxis. thus, mac including cy at more than mg/kg and tac for gvhd prophylaxis are optimal conditions of sct for patients with was. disclosure of conflict of interest: none. adenosine deaminase (ada) deficiency is an inherited autosomal recessive immunodeficiency which represents about - % of scid. since we diagnosed patients affected by ada-scid: underwent hematopoietic stem cell transplantation (hsct), were treated with replacement therapy with peg-ada and received gene therapy; patients died before or after treatment. maternal t lymphocyte engraftment is frequently detected in scid patients, but this is never been found in ada deficient patients. a -months-old italian girl, from non-consanguineous parents, presented to our hospital with a history of frequent bronchiolitis associated with dermatitis, mycosis, hypogammaglobulinaemia, marked lymphopenia (t cells cd , /mmc; cd /cd , /mmc; cd / cd , /mmc, b cells . /mmc, and nk cells, /mmc) and in vitro absence of proliferative response to pha. level of immunoglobulins was almost normal (igg mg/dl, iga mg/dl, igm mg/dl). high levels of toxic metabolites were found: axp, . micromol/ml rbc; daxp, . micromol/ml rbc; %daxp, . . ada activity in rbc lysates was abnormally high for scid-ada ( . u/g hb). molecular analysis confirmed diagnosis: the sequencing of exon revealed two mutations: a missense mutation previously reported called p.ser leu (c. c t) and a new missense mutation defined p. leu pro (c. t c). t-cells str analysis of patient showed . % maternal t lymphocytes engraftment never reported before in ada-scid patients. the girl was transferred to the isolated bmt unit and the respiratory symptoms progressively improvement. replacement therapy with peg-ada was started immediately at a dose of u/kg/twice per week. ig therapy was started at a dose of mg/kg every two weeks. after three months of treatment patient showed an increase in t cells count (cd , /mmc), and a decrease of toxic metabolites: axp, . micromol/ml rbc; daxp, . micromol/ml rbc; %daxp, . maternal t-cell engraftment persists, despite a good response to the peg-ada therapy. the last examination before hsct reveals maternal t-cell engraftment of . %. patient underwent hsct from mud hlaidentical donor after a myelo-ablative reduced intensity conditioning regimen protocol d ebmt/esid guidelines. the number of infused cd + cells was . × /kg and . × cd /kg. she is actually at day+ post hsct, is doing well and shows % engraftment of donor cells. disclosure of conflict of interest: none. graft versus host disease (gvhd) is a frequent complication in patients undergoing haematopoietic stem cell transplantation. while the exact pathophysiology of gvhd is not known, the gut microbiome has been implicated in its development since it was shown that total gut decontamination (tgd) decreases the incidence of gvhd. with this study we aim to get insight into the diversity of the gut microbiota before, during and after total gut decontamination in comparison with selective gut decontamination (sgd). secondly, we want to identify changes in microbiota composition that relate to the occurrence of graft-versus-host disease. for this prospective cohort study we recruited children (o y) that were eligible for a stem cell transplantation at the leiden university medical center between january and december . of these, % (n = ) received tgd (consisting of piperacillin/ tazobactam and oral amphotericin b), whereas the other % (n = ) received selective gut decontamination with polymyxin /neomycin and oral amphotericin b. in total, fecal samples were collected, weekly during admission for the stem cell transplantation and monthly thereafter up to months after transplantation. also samples were collected from family stem cell donors as healthy controls. samples were processed within hours and stored in the - freezer, after which s v amplicon sequencing (illumina hiseq, rapid mode, bp read length) was applied. data analysis (taxonomy and shannon diversity) was primarily done using qiime (ref). compared to microbiota diversity in stem cell donors (mean shannon index (si) . ), we observed an overall lower mean si during tgd ( . ) and slightly higher mean si during sgd ( . ). microbiota diversity months after sgd ( . ) was similar to diversity during sgd ( . ), while diversity months after tgd ( . ) was higher than during tgd ( . ). further analysis of repopulation dynamics demonstrated no differences in repopulation duration after both decontamination strategies. however, we did observe differences in the type of bacteria that repopulated, with bacteroidales being more prominent in sgd and lactobacillales more prominent in tgd patients. actinomycetales (genus rothia) was exclusively present in tgd patients during decontamination. also, the clostridiales (blautia, lachnospiraceae and peptostreptococcaceae) were bacteria that appeared after the decontamination period. four patients ( %) in this cohort developed gvhd grade or more. in these patients we did observe individual compositional changes of the gut microbiota at the time of ghvd diagnosis, e.g very low diversity or dominance of enterobacteriales. considerable microbiota diversity is observed in patients that received tgd. different repopulation dynamics were observed between tgd and sgd. no common pattern was found in the gvhd cases. disclosure of conflict of interest: none. minimal residual disease (mrd) pre-and post-hct for children with aml is highly predictive of event-free survival: a pediatric blood and marrow transplant consortium study d jacobsohn johns hopkins all children's hospital, children's hospital los angeles, usc keck school of medicine multicenter data regarding the significance of mrd in children with aml pre-and early post-hct are lacking. we hypothesized that pre-and post-hct mrd assessments using wt pcr combined with multi-dimensional flow cytometry (mdf) would be predictive of disease relapse and event-free survival (efs) at -yrs post-hct. subjects were o yrs with aml in morphologic cr undergoing ma allogeneic hct. stem cell sources included bm, pbsc, or cb. bm and pb samples were collected at time points: baseline ( o weeks prior to preparative regimen); day+ (± days); and day+ (± days). bm samples were analyzed for both wt expression and mdf mrd (single reference lab using a 'difference from normal' approach without access to diagnostic phenotype); pb samples were analyzed for wt only. mdf detection limit was . %; however, we required that independent analysts certify that the abnormal population was aml. in addition, sorted mrd+ cells were tested for chimerism. wt positivity was defined as ⩾ copies for bm and ⩾ copies for pb. results were not available to the treating clinician. subjects were enrolled at centers in us and canada. enrolled subjects did not undergo hct and were excluded for progression prior to hct or other ineligibility. in eligible subjects, -yr efs and os were % and %, respectively. the -yr ci of relapse and trm were % and %, respectively. mdf identified subjects pre-hct having . - % residual disease. the -yr relapse rate in subjects with +mrd by mdf pre-hct was % vs % ( - %) in those who were negative. -yr dfs and os were % and % ( - %) for positive mdf pre-hct, and % ( - %) and % ( - %) for negative mdf. pre-hct mdf sensitivity for -yr dfs was %; specificity was %. mdf mrd at days and were similarly predictive of outcome. sorted mrd+ cells from post-hct samples were all noted to be of recipient origin. pb wt had no correlation with dfs or relapse; bm wt at day+ correlated with -yr os ( % ( - %) low/neg vs % ( - %) high). other wt cutoffs studied demonstrated no correlation with outcomes. figure : relapse probability by flow cytometry mrd at time points. mdf mrd pre-hct and at days + and + was significantly associated with lower efs and os in children with aml undergoing hct. mdf is specific but not sensitive, as many negative mdf patients relapsed. our goal was to define a reproducible assay that did not depend on having the initial aml profile. this would facilitate multi-institutional studies aimed at decreasing relapse. given that constraint, we were able to detect clear mdf mrd in a small percentage of patients that was highly predictive and can be used in trials. wt level was not predictive in this multi-institutional trial. the sensitivity of flow was significantly affected by not having the initial flow available. future attempts to improve sensitivity should include initial flow and/or test higher channel flow or molecular pcr techniques. in addition, we confirmed that mrd + cells obtained by cell sorting post-hct were of recipient origin. future testing of 'suspicious' sorted cells by fish, molecular, or comparative genomic hybridization could possibly increase mfd sensitivity. novel cellular or targeted therapies should be tested in clinical trials to improve outcomes in patients with mfd mrd noted either pre-or post-hct. [p ] disclosure of conflict of interest: none. novel mutations were identified with ngs and low intensity of conditional regimen succeeded in children with fanconi anemia who received allo-hsct s hu , h hou, j lu, p xiao, x bian, h liu, y hu, j ling, l li, l kong, z zhai and y yao children's hospital of soochow university to explore the possiblity of applying next-generation sequencing (ngs) to diagnose the disease of fanconi anemia (fa) and evaluate the efficiency and safety of low intensity conditional regimen on children with fa receiving allogenic hematopoietic stem cells transplantation (allo-hsct). five patients initially suspected as severe aplastic anemia were diagnosed as fa by the method of next-generation sequencing (ngs)-based genetic diagnosis panel. one patient received hla-identical sibling donor hematopoietic stem cell transplantation (mrd), three patients underwent unrelated donor matched (ud) hsct, and one patient received unrelated cord blood transplantation (ucb). the conditional regimen consisted of either cgy tbi or . - . mg/kg of busulfan with - mg/kg of cyclophosphamide. meanwhile, atg at mg/kg and fludarabin at - mg/m were included as well. cyclosporin or tacrolimus as well as mycophenolate mofetil (mmf) were used for the prophylaxis of graft versus host disease (gvhd). engraftment of neutrophil and platelet and complications followed transplantation such as infection, gvhd, and hemorrhagic cystitis (hc) were observed. of cases diagnosed as fa by ngs, only case showed the abnormality of chromosome fragility test which has been regarded as golden criteria in the diagnosis of fa. meanwhile, we found novel mutations in cases of fa which enriched chinese national database with data of rare diseases by ngs. the counts of mononuclear cells (mnc) were ( . - . ) × /kg for non-ucb and . × /kg for ucb. the counts of cd + were ( . - . ) × /kg for non-ucb and . × /kg for ucb. all cases succeeded in allo-hsct with the low intensity of conditional regimen. the median time for neutrophils engraftment was days (range ~ days), median time to platelets (plt) engraftment was days (range ~ days). one case occurred with grade i of agvhd, cases with hemorrhagic cystitis. after transplantation, all patients were monitored the copies of ebv-dna and cmv-dna of whole blood, and five case with ebv positive and cases with cmv positive. no patient suffered of ebv or cmv disease. the hepatic veno-occlusive disease (vod) and hc were observed in fa patients after transplantation. ngs showed much more specific and facilitated for the diagnosis of fa. low intensity of conditional regimen is efficient and safe which should be recommended for the treatment of fa patients. disclosure of conflict of interest: none. outcome of alternate donor stem cell transplantation in children: an indian experience sp yadav , , n rastogi , , d thakkar , , s kohli , , s nivargi , , r misra and s katewa in india due to lack of donor registries and cord blood banks very few alternate donor stem cell transplants (sct) are performed. haploidentical sct has become feasible with availability of post-transplant cyclophosphamide (ptcy) technique. here we present our experience of setting up alternate donor program for sct for children in india and report the outcomes of the same. we collected data retrospectively of all children who underwent alternate donor sct during jan to dec in two centres. a total of sct were performed for children; median age years ( - years) and were boys and girls. of these, underwent haploidentical ( ptcy and tcr alpha-beta/cd depleted), matched unrelated donors (mud) and unrelated cord blood (ucb) sct. the diagnosis was: primary immunodeficiency- , thalassemia major- , sickle cell disease- , inherited bone marrow failure- , acquired aplastic anemia- , acute lymphoblastic leukemia- , acute myeloid leukemia- , neuroblastoma- , ewings sarcoma- & leukodystrophy- . the conditioning was with fludarabine, cyclophosphamide and total body irradiation backbone in children (thiotepa added in ), fludarabine and treosulfan in , fludarabine and busulfan in , busulfan and cyclophosphamide in . serotherapy was part of conditioning, rabbit anti-thymoglobulin . mg/kg in and campath mg/kg in . graft vs host disease (gvhd) prophylaxis was ptcy along with tacrolimus and mycophenolate mofetil in patients ( haploidentical, mud & ucb) and ex-vivo tcr alpha-beta depletion in and cyclosporine and methotrexate in . all were transplanted after a signed informed consent. a median of million of cd cells/kg was infused (range - million/kg).graft source was peripheral blood in and bone marrow in and ucb in . five children died before engraftment. the remaining had neutrophil engraftment by median of days (range - ) and platelet engraftment by median of days (range - ). chimerism at day+ was available in cases; of them had full donor hematopoiesis. one had mixed chimerism and fully recipient. four children underwent a second haploidentical sct after rejection of which are alive and disease free. the median follow-up of remaining patients is months (range - ); the cumulative incidence of graft versus host disease (gvhd) acute and chronic extensive was % and % respectively. grade-iii-iv acute gvhd was seen in patients. a total of patients have died (sepsis- , stroke- , gvhd- , vod- , encephalitis- and progressive disease- ). among encephalitis deaths, one child had undergone ucb with ptcy and another tcr alpha-beta depleted second sct.; both had bk virus in the csf.there were / deaths in haploidentical (ptcy- / & tcr alpha-beta- / ), / in mud and / in ucb sct. overall survival is % and disease free survival is % at last follow up. alternate donor sct is an acceptable curative option for children lacking a matched sibling donor. haploidentical donor sct is more feasible in the setting of lack of donor registries having indian ethnicity donor. disclosure of conflict of interest: none. hematopoietic stem cell transplantation (hsct) from an unrelated donor (ud) is largely used for pediatric patients with all in second complete remission (cr) lacking an hlaidentical sibling. in this study, we retrospectively analyzed outcome of patients (pts) given ud-hsct in centers affiliated to the associazione italiana di ematologia ed oncologia pediatrica (aieop) network between and . three hundred fifty-six pts with all in second cr experiencing either bone marrow (bm), isolated extramedullary or combined relapse were included in the study; were males and females, median age at hsct being . years (range . - ). bm, peripheral blood (pb) and cord blood (cb) were the stem cell source in %, % and % pts, respectively. all children received a myeloablative conditioning regimen, either tbi-( pts) or chemotherapy-based ( pts). as gvhd prophylaxis, the combination of cyclosporine a, short-term mtx and atg was employed in most pts. according to the berlin-frankfurt-munster (bfm) classification of first leukemia recurrence, % and % of pts were assigned to the s +s and s +s groups, respectively. level of pre-hsct minimal residual disease (mrd), measured within days before hsct through flow cytometry (fcm) in the laboratory of padova, is available in children; more data will be presented during the s meeting. with a medium follow-up of . years (range . - ), the overall survival (os) was %, while the event-free survival (efs) was %. the cumulative incidence of transplant-related mortality (trm) and leukemia recurrence were % and %, respectively. the efs probability for children transplanted in the time period [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] and - was %, % and %, respectively (p = ns). patients who received a tbi-based conditioning regimen had a significantly better outcome in comparison to children who received chemotherapy-based treatment, efs being % and %, respectively (p = . ). efs of pts belonging to s +s and s +s groups was % and % respectively (p = . ). the difference in efs is largely explained by an increased incidence of leukemia recurrence in s +s ( %) compared to s +s pts ( %) (p = . ). efs of pts who experienced grade ii acute gvhd was %, while that of pts with either absent/grade i acute gvhd or grade iii-iv acute gvhd was % and %, respectively (p = . ). pts with limited chronic gvhd had a better efs as compared to those with either extensive or absent chronic gvhd ( %, % and %, respectively; p = . ). the choice of stem cell source (bm, pbsc, cb) did not influence the probability of efs, which was %, %, % respectively (p = ns). importantly, among pts with evaluable mrd before hsct (n ), the group with detectable levels . % (n ), respectively % and % (p = . ). conclusions. outcome of children with nd cr all who underwent transplant from an ud is significantly influenced by the presence of tbi in the conditioning regimen, limited severity of acute and chronic gvhd and bfm classification at time of st relapse. notably, mrd level before transplant, namely with a cut-off of . %, influences efs. disclosure of conflict of interest: none. the median mononuclear cell dose was . × /kg. the median time to reach absolute neutrophil count . × /l was days, and the median time to platelet count × was days. grade and grade mucositis was seen in % of our patients. transplant-related mortality at days not occurred. only three patients relapsed , and months after transplant (mean . m.). with a median follow-up of months ( - months) after transplant the event free survival were %. only one patient had death, two years after transplantation. no significant different between cbv group vs ceam group in engraftment day. high-dose therapy with stem cell rescue can lead to durable remissions in children and adolescents with advanced hd. future investigations should focus on strategies designed to decrease relapse after auto-transplantation, particularly in patients at high risk for relapse. our analysis suggests that these regimens (ceam, cbv) are feasible in pediatric patients with acceptable engraftment and toxicity. pbsc collection may be difficult in small children owing to the large volume apheresis compared to the child's weight. various problems, such as metabolic or haemodynamic disorders may be were seen. peripheral stem cell harvest can be performed in lowweight children under safe and effective conditions even when systematic priming by blood is avoided. processing with increase of blood volume may to increase in the yield by recruiting progenitor cells. disclosure of conflict of interest: none. outcomes of children with hemophagocytic lymphohistiocytosis given allogeneic hematopoietic stem cell transplantation in italy allogeneic hematopoietic stem cell transplantation (hsct) is the only curative treatment for patients with familial hemophagocytic lymphohistiocytiosis (hlh) or relapsed/ refractory hlh. we analyzed outcomes of a cohort of patients ( m, f) with hlh given hsct between and . median age at hsct was years (range . - ). genetic testing was performed for / patients ( %). mutation of prf was found in patients ( %), of unc d in ( %), of stxbp in ( %), of rab a in ( %), of sh d a in ( %), of birc in ( %) and of lyst in patient ( %). no known gene abnormality was found in patients who had recurrent/ refractory hlh. central nervous system (cns) involvement at diagnosis was recorded for patients ( %) and was present in of them ( %). the primary endpoint was event-free survival (efs), defined as the probability of being alive and in continuous complete remission (cr) at last follow-up. in order to determine efs, death from any cause, relapse or graft failure were considered events. ninety-five patients received one transplant, while received more than one hsct, because of rejection in patients or disease relapse in (preceded by rejection in case): hsct were performed in cases, while and hsct were performed in case each. donor for first transplant was an hla-matched sibling for patients ( %), an unrelated donor for patients ( %) and a partially matched family donor for patients ( %). conditioning regimen was busulfan-based for patients ( %), treosulfan-based for patients ( %) and fludarabine-based for patients ( %). the -year probability of overall and efs were % and % respectively (fig. ) . twenty-six ( %) patients died due to transplant-related causes, while ( %) and ( %) patients experienced graft rejection and/or relapse, respectively (see also fig. ). twelve out of children ( %) given a nd hsct after graft failure/relapse are alive and disease-free. active disease at hsct was not statistically associated with adverse outcomes, while patients had a trend for a worse outcome if the interval between diagnosis and hsct was months. patients transplanted from partiallymatched family donors (pmfd) had a significantly worse efs ( %) than recipients of a matched family donor transplant ( %) or a matched unrelated donor allograft ( %, po . ). the main reason for the dismal efs of pmfd recipients was graft rejection, which, however, was largely rescued by a nd hsct. patients given peripheral blood stem cell transplantation had a lower efs probability ( %) as compared to bone marrow ( %) or cord blood recipients ( %, p = . ). children given hsct o o/u months from diagnosis had a better efs as compared to those transplanted months from diagnosis ( % vs %, p = . ). in multivariate analysis, only the use of a pmfd predicted a worse efs probability (relative risk: . , p = . ). these data suggest that in patients with hlh allogeneic hsct is able to cure / of patients. haploidentical hsct in patients with hlh is currently associated with unsatisfactory rate of engraftment, new approaches being needed to ameliorate this outcome. active disease does not preclude the chance of benefiting from transplantation, which should be ideally performed within months from diagnosis. [p ] defibrotide shows efficacy in the prevention of sinusoidal obstruction syndrome (sos) after allogeneic hematopoietic stem cell transplantation: a retrospective study on patients. disclosure of conflict of interest: none. standard gvhd prophylaxis regimens impair the graft-versustumor (gvt) effect, delay immune reconstitution and are associated with high rate of infections. high-dose posttransplantation cyclophosphamide (ptcy) targets alloreactive donor t cells proliferating early after bmt, promotes regulatory t cell and permits rapid immune reconstitution. in this pilot trial we evaluate the safety and effects of ptcy in unmanipulated haploidentical and matched unrelated transplantation (mud) in pediatric patients with all. fifteen pediatric patients with high risk all underwent unmanipulated allogeneic bone marrow (bm) (n = ) or peripheral blood stem cell (pbsc) (n = ) transplantation followed by ptcy between april and march with a median follow-up of months ( - ). eight patients were transplanted from haploidentical donors and from mud. the median age was . years (range . - ) and were in complete remission (cr) at the moment of bmt. in patients this was a second bmt. all pts. received myeloablative conditioning regimen (treosulfan-based n = , tbi based n = ) and ptcy on day + , + , posttransplant prophylaxis consisted of tacrolimus from day + (n = ), tacrolimus/mmf (n = ), atg (rabbit, thymoglobuline) at mg/kg without other posttransplant prophylaxis(n = , both from mud). primary engraftment was achieved in % of pts., the median time to neutrophil recovery was days and to platelet recovery was ( - ) days. all pts. had full donor chimerism on day + . causes of death included viral infections (n = ); gvhd and viral infection (n = ). cumulative incidence (ci) of acute gvhd grade ⩾ ii was % ( % ci: - ), grade iii-iv- . % ( % ci: . - ) and chronic gvhd- . % ( % ci: . - . ). two-year event-free survival (efs) and overall survival (os) were . % ( % ci: . - ) and were equal. median time of follow-up for survivors is years (range . - . ). we demonstrate that unmanipulated hsct and posttransplantation cyclophosphamide allows for high rate of engraftment with acceptable transplant-related mortality in pediatric patients with all. all major outcomes were equivalent between transplantation from unrelated and haploidentical donor. gvhd prophylaxis including ptcy was effective. event-free survival was high despite chemotherapybased conditioning in most patients. disclosure of conflict of interest: none. serotherapy with atg is frequently used in allogeneic hsct to prevent gvhd and rejection. however, the choice of the two most frequently used rabbit atg brands depends on country, disease protocol, national recommendations and/or physician's preference. atg-genzyme (atg-g, thymoglobulin) is prepared by immunizing rabbits with human thymocytes, whereas rabbits are immunized with a jurkat cell line for production of atg-fresenius (atg-f, recently named as antihuman t-lymphocyte immunoglobulin atlg, grafalon, noveii biotech). the recommended dose of both brands differs a factor - . we have previously reported the pharmacokinetics/ pharmacodynamics (pkpd) of atg-g in a large cohort of pediatric hsct recipients and concluded that the clearance of the active component of atg, which is the portion of atg binding to lymphocytes, had a major impact on immune recovery post-hsct, while total atg did not. both atg brands have frequently been compared according to disease outcome, without detailed analysis of composition and clearance of the active components. in the present study, we compared clearance of the active component and immune recovery after atg-g and atg-f, respectively. the serum concentrations of total and active atg were measured longitudinally after hsct in children ( atg-g, atg-f), transplanted with bm or pbsc of unrelated donors for all or aml between january and june in leiden (n = ) or copenhagen (n = ). atg-g treated patients received a total dose of - mg/kg and atg-f was given at a total dose of - mg/kg in both cohorts administration was divided over - days. serum samples (pre-conditioning, day of hsct, + ; + ; + ; + and + weeks and + and + months after hsct) were analyzed by elisa for total atg and by quantitative flow cytometry on hut cells for active (lymphocyte binding) atg. lymphocyte (sub-)populations were analyzed at + , + and + months post-hsct by flow cytometry. as reference group for immune recovery, children transplanted for all or aml with an hla-identical donor and not receiving serotherapy were included. the median serum concentration of total atg at the day of hsct was times higher for atg-f (atg-g μg/ml, atg-f μg/ ml; figure a) as the result of the higher dose of atg-f given. the active atg concentration was twice as high for atg-f (atg-g . au/ml, atg-f . au/ml figure b ). three weeks later at the expected time of engraftment, the total atg concentration was decreased with the same factor for both atg brands (atg-g from to μg/ml, factor . ; atg-f from to μg/ml, factor . ). however, the active atg concentration showed a much faster decline for atg-f (atg-g from . to . iu/ml, factor . ; atg-f from . to . iu/ml, factor ). correspondingly, the number of cd t-cells at month post-hsct was higher after atg-f than after atg-g (atg-g, atg-f and no-serotherapy , and cells/μl, respectively. figure c) . this is the first study to compare the pkpd of total and active atg-genzyme and atg-fresenius. active atg-f showed a much faster clearance than atg-g, which was associated with a significantly faster cd t-cell recovery at month post hsct. thus, atg-f is not only quantitatively but also qualitatively very different from atg-g, which will clearly impact hsct outcomes. reduced toxicity myeloablative conditioning regimen in pediatric hematologic malignancies not associated with improved outcomes s chaudhury , , i helenowski , r duerst , , wt tse , , m kletzel , , j schneiderman , and d jacobsohn ann and robert h. lurie children's hospital of chicago; northwestern university feinberg school of medicine, chicago and children's national health system, washington dc allogeneic (allo) hematopoietic cell transplantation (hct) is the only curative potential therapy in refractory and relapsed pediatric leukemias. poor outcomes in allo hct are associated with treatment-related mortality (trm), mostly due to regimen-related toxicities (rrt) and graft-versus-host disease (gvhd) after myeloablative conditionings (mac), but high relapse rate with reduced-intensity or nonmyeloablative regimens. to improve trm, without compromising conditioning intensity, we prospectively explored the feasibility and efficacy of a mac but reduced-toxicity conditioning (rtc) regimen, consisting of fludarabine mg/m /d (given first) × d, daily busulfan dosed to target an auc of microm*min/d × , ratg . mg/kg/d × and cgy of total body irradiation in patients (table ) with hematologic malignancies. gvhd prophylaxis was cyclosporine and mmf. all patients tolerated the rtc well, with no graft failures. rrt included moderate mucositis ( %), infections (bacterial %, viral reactivation %, fungal %) and cases of venoocclusive disease (vod). cumulative incidence d ⩾ gr acute gvhd was % ( % confidence interval [ci], - ), extensive chronic gvhd was . % ( % ci, . - ). mortality at days was . % ( % ci - ), due to infections with agvhd and vod. with a median follow-up of . y (range, . - ), the cumulative incidences of relapse at years was % ( % ci, - ). mortality due to severe agvhd was %. overall survival (os) and progression-free survivals (pfs) for year was % ( % ci, - ), and % ( % ci, - ) respectively. on univariate analysis there was no association of outcomes with donor type, graft source, disease or busulfan exposure except significantly higher cgvhd in unrelated donors, agvhd severity with peripheral blood. in summary, the use of the myeloablative rtc resulted in comparable trm, with high relapse rate was high, including in those developing chronic gvhd. this suggested a less robust graft-versusleukemia effect resulting in poor pfs and os. nonetheless, this regimen may be used as a lower-trm platform to combine with other strategies, intensive disease monitoring pre and post hct, addition of post hct maintenance therapy in combination with marrow as the stem cell source to decrease relapse or gvhd. specific immune response to vaccinations decline after hematopoetic stem cell transplantion (hsct). re-vaccination of all hsct recipients is recommended in all guidelines but bcg vaccination is not recommended due to safety concerns after hsct. mycobacterium tuberculosis can cause severe disease in children including meningitis and milliary tuberculosis (tb). the bacille-carmette-guerin (bcg) is a liveattenuated vaccine with documented efficacy against milliary disease and meningitis. routine vaccination of all infants residing in countries with high tb incidence is recommended by world health organization. however, there is no data in literature regarding its safety in post hsct setting. here, we report children who underwent matched related allogeneic hsct at ankara university pediatric bone marrow transplant (bmt) unit and received bcg -months post-transplant. all patients were free of graft versus host disease (gvhd) and immunosuppressive therapy (ist) and had negative ppd skin test prior to vaccination. none of the recipients developed local or disseminated tuberculosis as a complication of bcg with a median follow up of years. we conclude that the bcg vaccine is safe in the post hsct period when administered at least months out of transplant to a selected group of patients who are free of gvhd and ist. disclosure of conflict of interest: none. single centre experience of harvesting bone marrow from donors o years of age r raj, r uppuluri , d subburaj , d jayaraman , k mullanfiroze , v swaminathan and l vaidhyanathan department of paediatric blood and marrow transplantation, apollo speciality hospital harvesting bone marrow for allogeneic marrow transplantation from donors o kg presents special challenges. we present data on sibling donors from our institution between and . the mean age was months with a range between months to months. children less than one year accounted for % of our donors with the youngest being months of age and the smallest donor weighed . kg. all aspirations were performed from iliac crests and all donors were given general anaesthesia by a paediatric anaesthetist. irradiated blood was transfused in % of the donors during the procedure. the volume of marrow obtained ranged from to a maximum of ml/kg donor weight. the product contained an average cd count of . × /kg recipient weight with a range from . to × /kg. only on one occasion was a second harvest needed, where the donor weighed kg and recipient kg with major blood group incompatibility requiring red cell reduction. the yield of cd cells per ml of bone marrow was on average % higher than children above years of age. all recipients showed brisk engraftment in weeks. none of these donors experienced major difficulties following the aspiration procedure. thus, very young children may safely donate marrow for allogeneic transplantation and the yield of stem cells obtained is substantial. this data is particularly relevant in transplantation for haemoglobinopathies like thalassaemia major and sickle cell anaemia, where families are being counselled about a target of kg for the donor in order to plan transplantation. disclosure of conflict of interest: none. sinusoidal obstruction syndrome-veno-occlusive disease in pediatric patients given either autologous or allogeneic hematopoietic stem cell transplantation (hsct). a retrospective study of the aieop-sct (italian haematology-oncology association-stem cell transplantation) group m faraci , r luksch, e calore , f saglio , a prete , mc menconi , v trevisan , g de simone , v tintori , s cesaro , s santarone , mg orofino , e lanino , m zecca and a bertaina sinusoidal obstruction syndrome (sos), known as venoocclusive disease (vod), is a potentially life threatening complication that can develop after hsct. although sos progressively resolves within few weeks in most patients, the severe forms result associated with multi-organ dysfunction and high mortality rate ( %). aim of this survey is to evaluate incidence and management of sos in a large cohort of children receiving either allogeneic or autologous hsct. we retrospectively reviewed pediatric hscts performed in ( %) out of aieop affiliated centers, between january and april . new ebmt criteria have been used for the diagnosis of sos (serum total bilirubin ⩾ mg/dl and of the following criteria: painful hepatomegaly, weight gain %, and ascites) and for the classification of severity grading. , among a total number of hsct procedures ( autologous and allogeneic), we identified ( . %) patients with sos. this complication occurred in and cases after autologous and allogeneic hsct, respectively. fiftytwo pts ( %) received iv busulphan (bu) at myeloablative dose, ( %) oral bu, while ( %) were treated with different conditioning regimen. the median time of sos occurrence was days after hsct. details about prophylaxis and therapy are reported in figure . out of the children, ( %) fulfilled all sos-ebmt criteria. bilirubin ⩾ mg/dl, gain of weight %, ascites, and painful hepatomegaly did not occurred in , , and patients, respectively. thrombocytopenia was present in pts ( %), thickening of gallbladder in ( %) and abnormalities of coagulation parameters in ( %). according to sos ebmt severity grading, levels of transaminases were mild in pts ( %), moderate in ( %), severe in ( . %), and very severe in ( . %). notably, creatinine was mild in pts ( %), while ( . %), ( . %), and ( %) children showed moderate, severe and very severe grade of renal failure. thirty-three pts ( %) had respiratory failure, and ( %) of them experienced right pleural effusion. six out of the patients who developed acute kidney injury, required dialysis. severe encephalopathy occurred in pts ( . %) and ( %) out of the pts evaluated, were admitted in intensive care unit. as therapy of sos, pts received defibrotideâ (df); the dosage was mg/ kg/day in % of them. the median duration of df treatment was . days (range - ). thirty-three ( %) pts received methylprednisolone (median dose of mg/kg). fifteen pts ( . %) died due to mof ( in moderate, in severe, and in very severe group) at a median time of days from sos diagnosis (range - gg). our multicenter survey showed that, at least in our experience, there is a significant variability in the management approaches to sos/vod in children, while, diagnostic evaluations are more homogeneous. interestingly, in our cohort, the increase of bilirubin may be an absent criteria, while thrombocytopenia and abnormalities of coagulation parameters are more frequent. as expected, mof occurred mostly in patients experiencing severe sos. df represents first strategy to treat sos in the majority of patients, even if steroids and ursodeoxycholic acid are still used. the hyper-ige syndromes are characterized by marked elevations in plasma ige levels and eosinophilia with impairment in t cells which clinically results in combined immune deficiency. dock deficiency, the autosomal recessive form, brings about allergic/atopic manifestations and unusual susceptibility to infections with herpesvirus family members (herpes simplex virus, human papilloma virus) and molluscum contagiosum. symptoms in patients with dock deficiency typically emerge during childhood, and the majority results in death because of infections and malignancy by the third decade. hematopoietic stem cell transplantation (hsct) is now considered a standard of care for dock deficiency when an appropriate donor is available. in this study, we present the unrelated hsct results of children with dock mutation. the demographic and clinical data of the patients with transplantations studied are shown in table . hsct was administered between august and august at bahçeşehir university medical park antalya hospital and the clinical data of the hscts are presented in table . all patients were transplanted from unrelated donors with bone marrow, except one with cord blood. the cord blood transplantation´s regimen was non-myeloablative which resulted with rejection. despite existence of serious morbid problems before transplantation, all the patients engrafted successfully. majority of the complications mentioned in the table were improved and they are in the follow-up in an outpatient basis. discussion dock deficiency has high mortality, and hsct should be considered as early as possible before development of significant organ damage. despite myeloablative conditioning and high morbidity before the transplantation, survival was very good in our patients. myeloablative and nonmyeloablative transplants have been performed from related and unrelated donors and have reported successful results even without the preparative regimen. in our center, all transplants performed from unrelated donors by myeloablative regimen have been successful but have resulted in transplant rejection with cord blood transplantation after nonmyeloablative regimen. in all of our patients, stable full chimerism has been detected, however mixed chimerism have also been shown to be useful in several reports. whether hsct also cures the autoimmune complications and reduces the risk of cancers is as yet undetermined. however, a myeloablative conditioning regimen followed by allogeneic hematopoietic stem cell transplantation from unrelated donors in dock deficiency results in improvement of the clinical phenotype with a low incidence of regimen-related toxicity. disclosure of conflict of interest: none. successful bone marrow transplantation after myeloablative conditioning in a child with ipex syndrome b kuşkonmaz , d ayvaz , mh abur , fv okur , g karagüzel , f orhan , İ tezcan and du Çetinkaya immune dysregulation, polyendocrinopathy, enteropathy, x-linked (ipex) syndrome is a rare disorder. although most patients present in infancy with a clinical triad of intractable diarrhea, insulin-dependent diabetes, and eczematous dermatitis, some patients present with severe food allergies and other autoimmune manifestations. the disease is caused by mutations in the forkhead box p (foxp ) gene, a transcription factor that is essential for the development and function of regulatory t (treg) cells. this cells plays an essential role in controlling immune responses and preventing autoimmunity. patients usually die in the first years of life without treatment. the only effective cure is hematopoietic stem cell transplantation (hsct). here we report a patient with ipex syndrome who underwent hsct after myeloablative conditioning. months of age boy with the history of diarrhea, insulin-dependent diabetes, eczematous dermatitis, pneumonia, coombs positive hemolytic anemia, referred to our hospital for investigation of immunodeficiency. on admission physical examination showed eczematous skin rash, submandibular lymphadenopathy, hepatosplenomegaly. before hsct the patients treated with immunosuppressive agents including methylprednisolone, mycophenolate mofetil and monthly intravenous immunoglobulin. complete blood count revealed anemia (hb: . g/dl), and eosinophilia ( /mm ). serum immunoglobulins were: ig g: mg/dl ( - ), igm: mg/dl ( - ), iga: . mg/dl ( - ), ige : iu/ml. lymphocyte subset analysis showed cd %, cd %, cd %, cd + %, cd %. foxp gene analysis showed c. _ delaag mutation. at the age of year, patient underwent hsct from his hla matched sibling. myeloablative conditioning regimen including busulfan ( . mg/kg) and fludarabine ( mg/m ) was given to the patient. cyclosporine a and methotrexate (day + , day + , day + ) were used as graft versus host disease prophylaxis. bone marrow was used as the stem cell source and the number of cd + cells was . × /kg. neutrophil and platelet engraftment were achieved on day + and + [p ] s respectively. acute and chronic gvhd were not observed, but patient developed veno-occlusive disease treated with defibrotide, sepsis treated with broad spectrum antibiotics. chimerism analysis showed % donor profile at the third month of hsct. after hsct, autoimmune hemolytic anemia, eczematous dermatitis, food allergies, diarrhea and type diabetes resolved completely within two months after hsct. now the patient is in good clinical condition without any symptoms months after hsct. early hsct provides better outcome in patients with ipex, before the organ damage due to autoimmunity and/or adverse effects of immunosuppressive therapy. myeloablative conditioning is associated with substantial transplantation-related mortality whereas nonmyeloablative conditioning carries an increased risk of rejection because of dysregulated effector t-cell function. in this patients, myeloablative conditioning was preferred because of the risk of rejection. although the required levels of donor chimerism and conditioning intensity are unknown, engraftment of donor treg cells seems to be sufficient to control the disease. the patient is well without any symptoms of ipex after hsct with full donor chimerism. disclosure of conflict of interest: none. interferon gamma receptor deficiency (ifnr ) is a rare autosomal recessive immune deficiency disorder associated with very poor outcome secondary to severe and disseminated mycobacterial infections. hematopoietic stem cell transplantation (hsct) has been proposed as a curative option. however, hsct for these patients is particularly difficult owing to a high rate of graft rejection. the use of a non t-cell depleted transplant from an hla-identical sibling and fully myeloablative conditioning regimen has been shown to have improved outcomes. we report the first successful hsct with a t-depleted haplo-identical donor, performed in a girl with severe ifnr deficiency. we reviewed the medical chart of a -year-old hispanic girl with ifnr deficiency who was diagnosed at birth, since her brother had previously been diagnosed with the same complete ifnr deficiency. they were found to have a novel mutation variant detected at c. - g t. as expected with this disorder, she developed disseminated infection with mycobacterium abscessus infection at months of age and was subsequently found to have mycobacterium abscessus osteomyelitis. she was treated with multiple antibiotics including: amikacin, linezolid, meropenem and clarithromycin while tigecycline was added a few weeks prior to admission for hsct. she was continued on this therapy until day + following which antimicrobials were gradually weaned off. she was enrolled on the bp- trial, a multicenter, prospective phase i-ii trial (enrolling both malignant and non-malignant diseases) evaluating αβtcr +/cd + depleted haplo-transplantation followed by administration of bpx- t cells containing the ic suicide gene, (clinicaltrials.gov nct ). her conditioning regimen included busulfan ( mg/kg/day for days) and cyclophosphamide ( mg/kg/day for days). fludaragbne, tli ( cgy) . gvh prophylaxis with atg/rituximab. the patient received a graft with: tnc- . × cells/kg, cd + cells- × cells/kg, and αβtcr+ t cell content of . × cells/kg. as per protocol, since the αβ tcr+ t cells in the product was below threshold of × cells/kg, she did not receive any post-transplant immune suppression. bone marrow recovery occurred at day + with anc /mm and platelet recovery at day + . full engraftment with % donor chimerism based on cytogenetic analysis was observed at day + after transplantation and has remained stable. she is currently months post-transplant, and has done well without major complications and or signs of mycobacterial infection. there is limited data in patients receiving hsct for ifnr deficiency with very poor outcomes either relating to graft failure, transplant complications and progressive mycobacterial infection. to our knowledge, this is the first patient with ifnr deficiency transplanted successfully with a haploidentical donor and alive without any active mycobacterial infection. this report suggests that using a highly immunopotent graft depleted of only αβtcr+ t cells while retaining other immune effectors might offer a potential strategy to engraft these high risk patients using haplo-identical donors thereby allowing access to virtually all patients in need. disclosure of conflict of interest: none. tandem autologous stem cell transplantations for high risk pediatric embryonal central nervous system tumors: a single center experience k rosenfeld , r dvir , s constantini, j roth , s edelman , a tal , d levin , m manisterski , s achituv and r elhasid , department of pediatric hematology-oncology, tel aviv medical center; department of pediatric neurosurgery, tel aviv medical center and sackler faculty of medicine, tel aviv university pediatric embryonal central nervous system tumors are highly malignant tumors, which tend to disseminate through the cerebrospinal fluid to the brain and spinal cord and include: medulloblastoma, pinealoblastoma and primitive neuroectodermal tumors (pnets). the recommended treatment for these tumors is a complete surgical excision, craniospinal radiation and chemotherapy. the use of high dose chemotherapy with tandem autologous hematopoietic stem cell transplantation (hsct) has been advocated for high risk patients, and infants who could not be irradiated. between july and november , pediatric patients ( males, females) suffering from high risk medulloblastoma, pnet or pinealoblastoma underwent tandem autologous hsct. they were treated according to two protocols: group a consisted of ten patients with median age of . years (range . - . years) received the st jude sjmb protocol, while group b consisted of six patients with median age of . years (range . - . years) who received the children's oncology group -acns protocol. all patients engrafted with median time for neutrophil engraftment of days (range - days) and for platelets engraftment ( ) of days (range - days). median follow-up was . years (range week- years). neurological toxicity: two group a patients had convulsions episodes, one occurred during infusion of cryopreserved stem cells, and the other was a result of progressive disease during the last course of hsct. gastrointestinal toxicity: seven patients required total parenteral nutrition due to mucositis. diarrhea occurred in seven patients, two of them were diagnosed with rota virus and two with clostridium difficile. infectious complications: all patients suffered from at least one episode of neutropenic fever which was treated with broad spectrum antibiotics. there were documented bacteremia in patients. ( klebsiella pneumonia, proteus mirabilis, staphylococcus aureus, streptococcus viridans and staphylococcus epidermidis). metabolic complications: four patients in group a developed reversible syndrome of inappropriate anti-diuretic hormone secretion (siadh) during chemotherapy, and all group a patients developed hypomagnezemia. four patients died, one due to progressive disease, one due to early relapse months post treatment, one due to late relapse years post treatment and one due to sepsis months post treatment. another patient relapsed . years s post treatment, underwent surgery and radiotherapy and is now years post therapy. late effects: four group a patients developed endocrinological sequelae at a median of months (range - months) and require hormone replacement therapy. tandem autologous hsct is a feasible treatment for pediatric high risk embryonal tumors, with good engraftment and acceptable toxicities using sjmb and acns protocols, with overall survival of %. long follow-up is needed in order to diagnose and treat late effects. disclosure of conflict of interest: none. the diagnostic role of liver stiffness measurement in predicting hepatic veno-occlusive disease (vod) in pediatric hematopoietic stem cell transplantation (hsct) k kleinschmidt , f ravaioli , r rondelli , g marasco , r masetti , a prete , a colecchia , d festi and a pession pediatric oncology and hematology unit, department of pediatrics, university of bologna, sant 'orsola-malpighi hospital and department of medical and surgical sciences, university of bologna vod is a potentially life-threatening complication associated with hsct in which immediate therapeutic action is crucial for patients' outcome. liver stiffness measurement (lsm) using fibroscan represents a non-invasive method to detect the grade of liver fibrosis and portal hypertension as in case of vod. to evaluate the predictive potential of lsm in pediatric patients (pts) at risk for developing vod, a prospective, ongoing, single-center study has been performed at the university hospital of bologna. lsm was performed by using the fibroscan device, which consists of a . mhz ultrasound transducer probe that transmits low-frequency vibrations ( hz) to the liver tissue. the propagation velocity is proportional to the stiffness (elasticity) of tissue. lsm will obtain pathological high values ( . kpa) when the tissue is altered like in liver fibrosis, or post-sinusoidal portal hypertension. from november -september , pediatric pts ( male, female), aged - years (mean . ), affected by hemato-oncologic disease, eligible to allogeneic ( ) or autologous ( ) sct conditioned with busulfan-based chemotherapy, were enrolled. pts were scheduled for study examinations with lsm: at t (baseline) before chemotherapy, t (day - after sct), t (day - ) and t (day - ). the diagnosis of vod was defined according to modified seattle/baltimore criteria. twenty-five pts were enrolled in the protocol, of which were evaluable for the study (pts characteristics table ). out of pts ( %) developed vod. the cumulative incidence (se) of vod in our setting was % ( . ). baseline lsm values on t of all pts were normal ( . kpa at t (p = . ) and t (p = . ). from our observations, an anticipating pattern of pathological lsm in presence of clinical and laboratory parameters within normal ranges in patients who develop vod can be derived. preliminary data indicate a high predictive potential of lsm in the diagnosis of vod, however the number of cases is not sufficiently representative to draw definitive conclusions. to optimize the predictive potential of the method, more frequent (daily) measurement in the critical time frame are currently investigated. [p ] all= acute lymphoblastic leukemia, aml= acute myeloid leukemia, bu= busulfan, treo=treosulfan, fluda= fludarabine. disclosure of conflict of interest: none. [p ] the exact role of extra-corporeal photopheresis in children with gvhd: an unanswered question ss anak, h bilgen , , , , , , , y yaman, et saribeyoglu, k ozdilli, v hazar, m elli, am kokrek, h hizli and k payalan ecp continues to be a controversial treatment, probably due to the mechanism of action not being identified, the varying photopheresis procedures and treatment schedules, and the difficulty of conducting trials on relatively rare diseases with involvement of clinically heterogeneous organs. ecp was performed in our pediatric transplant center to patients mean age of years ( - ) diagnosed to have all ( pts), thalassemia ( pts), aplastic anemia ( ), blacfan diamond ( ), refractory hodgkin disease ( ) following our internal protocol for ecp sessions. five of the patients had mud, had hla id sibling transplants. chronic gvhd was diagnosed in of the patients had acute gvhd. skin was involved in all the patients, liver in of the patients, lung in , gut in and mucous membranes in patients. the ecp treatment consisted essentially of three steps: ( ) collection of mncs from the patient, ( ) processing of mnc buffy coat, and ( ) return of mncs to the patient. collection was performed using a cell separator (haemonetics mcs plus), processing two blood volumes. our protocol provides for a maximum final mnc volume to be collected at ml, with a hematocrit (hct) value below %. the maximum procedure time was set at min. the mncs collected were adjusted to a constant volume of ml by the addition of saline and ml of -mop in aqueous solution, to always obtain a final concentration of the drug of ng/ml. the diluted buffy coat was transferred into a special uv-a-permeable bag (pit-kit medtech solutions), and uv-a radiation at j/cm was performed (uva-pit irradiator). the photoactivated mncs were returned to the patient within minutes using a blood transfusion set. during ecp procedure, patients' vital signs were monitored. anticoagulation consisted in acidcitrate-dextrose formula a set at a variable ratio ( : - : ) according to the patient's characteristics (clinical conditions, body weight, coagulation values) and platelet (plt) count. prophylaxis of hypocalcemia consisted of the administration of calcium gluconate ( ml diluted in - ml saline) every to minutes. all procedure related side effects were recorded. during the reinfusion and postreinfusion phases, the patients were monitored for fever, chills, headache, rash, erythema, urticaria, itching and edema. no serious complication was detected. all the patients had also steroids, had concurrent mesenchimal stem cells. ecp was applied on consecutive days every - weeks which is continued for approximately months followed by a maintenance schedule tapered to an every -to -weeks. the mean session cycle was ( - ) between february to november . the most commonly involved organ was the skin which demonstrated a response rate of %, followed by liver ( %), lung ( %), gut ( %) and mucous membranes ( %) the concurrent immunosuppression could be reduced during ecp therapy, and no increase in opportunistic infections was detected. / patient died after a relapse, / are alive with chronic mild gvhd. however, despite our good response rates, our understanding of ecp remains limited. patients who suffer from acute and chronic gvhd have limited treatment options. ecp remains an important therapeutic option. future basic, translational, and clinical research studies will provide a better understanding of its mechanism of action and optimize its therapeutic potential. disclosure of conflict of interest: none. tolerability and responses to ex vivo il activated nk cells from haploidentical parental donors in paediatric patients with refractory leukaemia/lymphoma pl tan prognosis for patients with refractory leukaemia/lymphoma ineligible for transplants and those who relapse posttransplant is poor. in adult settings, adoptive transfers of ex vivo il activated natural killer ('ank') cells from nk alloreactive donors, especially for nk sensitive cancers, has been successful in bridging patients to curative transplants.( ) this approach has not been reported in paediatric patients. we report our experience in consecutive patients, of median age (range, - ) years, with refractory leukaemia/ lymphoma (aml, ; all, ; mixed phenotype acute leukaemia, ; lymphoma, ) who received treatments with 'ank' from haploidentical parental donors on institutional protocol, between aug and . parents/legal guardians/patients provided informed consents as per institutional guidelines for donors and patients procedures. donor lymphocytes harvested at steady state were cd depleted followed by overnight culture in il before being infused into patients lymphodepleted with fludarabine and cyclophosphamide. additional rituximab were given to patients and another received tbi gy. subcutaneous il injections at doses - mu/m /dose started on d- and were planned for doses, as tolerated. nk alloreactive donors (kir-ligand mismatch) and kir b/x genotype were available to all except patients. two patients were treated for post-transplant relapse; of whom also received 'ank' pre-transplant; other patients had failed best conventional therapy including cd /cd bispecific t cell engager (blinatumomab) in . lymphodepletion was well tolerated. a median tnc and cd + dose of . (range, . to ) × /kg and . (range, . - ) × /kg, respectively were administered. cytokine release syndrome (crs) was observed in of treatments ( grade , grade , grade ). the patient with dock deficiency, disseminated ebv+ cerebral lymphoma had grade crs and robust tumour lysis syndrome but succumbed to neurotoxicity. of the treatments, there were responses, including the given posttransplant. excluding the treatments given post-transplant and non-responders, median peak donor chimerism was % (range, - %) occurring at a median of (range, - ) days. five patients ( responders, non-responder) proceeded to transplants at a median of (range, - ) days after 'ank.' responders had longer survival time compared to nonresponders (median vs days). two responders ( %) achieved sustained minimal residual disease (mrd) remission after transplants and are alive and days from 'ank.' five eventually died of their primary leukaemia/lymphoma; from crs. our preliminary experience in a small cohort of paediatric patients with refractory leukaemia/ lymphoma showed that adoptive transfers of ex vivo il activated nk cells from haploidentical parental donors were tolerable; with responses seen in % of patients; and % achieving prolonged mrd remissions after transplants. patients with cerebral diseases might be at increased risks of neurotoxicity with this approach; and care must be taken in patient selection and the design of the lymphodepletion therapy. alternative donor choices are limited in multi-racial, multiethnic societies with small families such as singapore. unrelated cord blood transplant provides a feasible alternative to patients lacking adult stem cell donors in children with primary immunodeficiency diseases. method: we describe our experience using unrelated cord blood transplant (ucbt) for children with pid from august to november . during this period we performed hsct for children with pid: with unrelated cord blood ( %); with msd and i mud. out of cases of ucbt there were severe combined immunodeficency (scid), chronic granulomatous disease (gcd), hyperigm syndrome and wiskott aldrich syndrome (was). the median age of transplant was . months (range . to . months). all presented with multiple infections ranging from disseminated bcg infection to parainfluenza /rsv /rotavirus infection to pseudomonas sepsis, staphylococcal endocarditis to pulmonary aspergillosis for scid. hyper igm presented with pnemocystitis carini pneumonia while cgd conditions presented with perianal abscess and fungal pneumonia. the child with was had life threatening git bleeding and a hemorrrhage trachaebronchial cast removed after a failed initial extubation for gastroscopy. conditioning regimes consisted of reduced intensive (fludarabine based) conditioning regime for scid and myeloablative regime for the rest. the median tnc dose was . × ( )/kg (range . to . ) and median cd + cells dose was . × ( )/kg (range to . ). results: all engrafted well except for one graft failure in cgd. he refused nd transplant and died . years post transplant from fungal pneumonia. median engraftment time for neutrophil was days (range to ) and platelet was days (range to days). grade skin aghvd occurred in one patient while another patient died of agvhd of liver and lungs. chronic gvhd was found skin and liver in one patient. trm was % (due to agvhd). median follow up was days (ranged to ). overall years survival was %. post-transplant complication with life threatening puemonitis was not uncommon. one patient developed biopsy -proven idiopathic interstitial pneumonitis and required ecmo for one month. he received immunosupressive drugs including methylprednisolone, infliximab, oral imatinib (tk inhibitor), azithromycin and nebulised becotide. he was weaned off oxygen after - months. conclusion: our limited experience showed unrelated cord blood is good source of stem cell for transplant in pid in a multiracial population. one case of graft failure was likely due too low cell dose cd +cells dose × ( )/kg. the expertise in icu has enabled us to support several patients who presented with infective pneumonia pre-transplant and post -transplant. with better technology like alpha/beta depletion haploidentical transplant may be a better option to achieve engraftment earlier so as to avoid stormy post-transplant infections seen in unrelated cord blood setting. disclosure of conflict of interest: none. in spite of these recommendations, literature from developing countries suggest that pbscs are used more and more frequently without compromising the transplant results, as they seem to be preferred graft source for donors in many countries incl. poland. therefore we analyzed the efficacy of mud-hsct in children with saa transplanted in our centre. clinical data of saa and pnh children and adolescents ( boys and girls), who underwent mud-hsct between october and july were retrospectively analyzed. the median age was . years (range . - years) according to the graft source, the patients were divided into pbsct group ( patients) and bm group ( patients). four patients required second mud transplant due to graft rejection. overall survival for all patients was %. estimated -year overall survival (os) was not statistically different between pbsct group and bm group [( % vs % ) p = . ]. there was no significant difference in os between group who had ist before transplant and the group, who had an upfront transplant as a first line of therapy [ % vs %, p = . ].the time to neutrophil and platelet engraftment was statistically longer in bm group than in pbsc group [(anc vs days, plt vs days, respectively) p = . ]. the incidence of grade iii-iv acute graft-versus-host disease (gvhd) in pbsct group was similar to that in bm group [ % ( / ) vs % ( / )]. the incidence of chronic gvhd in pbsct group was similar to that in bmt group [ % ( / ) vs % ( / )]. other transplantrelated complications like heart failure, central nervous bleeding, incidence of infections were comparable within the two regimens. there were deaths in the whole group. the main reason of death were infectious complications or multiorgan failure (mof) in severely pretransfused patients in this historical cohort of patients. unrelated donor pbsct in children and adolescents with saa seems to be not inferior to unrelated donor bmt. the incidence of chronic gvhd was surprisingly low in saa recipients of mud pbsc. increased morbidity and mortality due to infections was due to individual poor clinical situation of patients before transplant (i.e. fungal infections, contamination with resistant bacteria, prolonged neutropenia). disclosure of conflict of interest: none. dyskeratosis congenita (dc) is characterized by the clinical triad of reticular skin pigmentation, nail dystrophy, and oral leukoplakia. the majority of patients with dc develop bone marrow failure (bmf), which is the main cause of death in dc patients. allogeneic hematopoietic stem cell transplantation (hsct) is the only curative treatment for bmf associated with dc. transplant-related morbidity/mortality is common, especially after myeloablative conditioning regimens. hsct has been introduced into the management of dc, which has had remarkable clinical results. we report our experience in children with dc who underwent allogeneic transplantation at a single medical center. patients received a fludarabine-based reduced intensity conditioning (ric), and the graft source was unrelated peripheral blood stem cells. median age at the time of hsct was . years (range, - years). the numbers of infused mononuclear cells and cd + cells were . ± . × /kg and . ± . × /kg, respectively. the median time of neutrophil and platelet recovery were . days (range, - days) and . days (range, - days). two patients experienced grade ii-iii acute graftversus-host disease (gvhd), and chronic gvhd was only observed in one patient. all four patients remained alive and transfusion independent at the median follow-up of . months (range, - months). correction of previously existing physical defects was observed in two patients. unrelated peripheral blood hsct can be a curative option for dc. ric based on the type of disease is important to s achieve successful hsct. a larger sample size and extended follow-up of this rare patient population are needed to determine whether the changes in therapy will improve longterm survival. disclosure of conflict of interest: none. autosomal recessive hyper-ige syndrome due to dock mutation is a combined primary immunodeficiency, characterized by severe eczema, recurrent infections, and susceptibility to autoimmunity, malignancy, and multiple allergies, in addition to unusual high serum ige level. dock patients tend to have a progressive severe clinical course with mostly fatal outcome during second to third decade of life without hematopoietic stem cell transplantation (hsct). in our center we have a large number of dock patients. during a period of years ( - ), we transplanted patients with documented dock- mutation confirmed by molecular genetics. one patient did not receive any conditioning because of poor clinical condition and he died from severe cutaneous and gut gvhd and another patient received cbt with bu/flu with zero engraftment. the rest of the patients received hsct from hla full matched donor with chemoablation with bu/cy for all with % lymphoid and myeloid engraftment (str). among those patients who received chemoablation, gvhd developed in patients mostly grade i and ii. in addition patients died: one died of severe gvhd and the other two died of sepsis. for dock patients we highly recommend early hsct if fully matched donor is available to prevent the high mortality associated with the disease. alloreactivity triggered by interactions between killer cell immunoglobulin-like receptors (kir) and natural killer (nk) cells plays a role in graft-versus-tumor (gvt) effects after hematopoietic stem cell transplantations (sct). in particular, kir-ligand mismatching between the donor and recipient might promote nk cell alloreactivity after unrelated cord blood transplantations (ucbt) in adult patients with acute myeloid leukemia (aml). recently, it has been suggested that allogeneic nk cells could be the effector cells that mediate gvt effects after mismatched allogeneic transplants for refractory childhood solid tumors. however, there are few reports about the efficacy of kir-ligand mismatched sct in pediatric cases. here, we report the excellent outcomes of kirligand mismatched cbt (kir-cbt) in pediatric patients with refractory malignant disease. we evaluated the cases of pediatric hematology and oncology patients [ ( %) aml, ( %) myelodysplastic syndrome [mds] , and ( %) neuroblastoma [nbl] patients] who underwent kir-cbt between and at our institution. among the aml cases, one involved refractory disease (induction failure), and the other three involved relapsed aml (one patient relapsed after the st sct because of q-). all nbl patients underwent kir-cbt followed by auto-peripheral blood stem cell transplantation (pbsct) because of stage disease. the mds patient underwent kir-cbt because of refractory anemia with excess blasts. kir mismatching was defined as incompatibility between the donor kir and recipient kir ligand, and only inhibitory kir that interacted with human leukocyte antigen (hla)-bw , -c , or -c group ligands were considered. the median age of the patients was (range - ) years. all of the aml patients were in complete remission (cr) at the time of the hsct (cr = one case, cr = cases). the mds patient was in a non-cr state, and all of the nbl patients were in their st cr at the time of the hsct. the aml patients received total body irradiation (tbi)-based conditioning ( gy tbi and mg/kg cyclophosphamide [cy]), and the mds patient received busulfan (bu)-based conditioning ( . mg/kg bu and mg/kg cy). the nbl patients received reducedintensity conditioning regimens ( mg/m fludarabine, mg/m l-pam, and gy tbi). the cb exhibited hla - locus mismatches (dna typing), including at least one inhibitory kir gene mismatch. the prophylaxis for graftversus-host disease (gvhd) consisted of tacrolimus and shortterm methotrexate. anti-thymocyte globulin (atg) was not used as a gvhd prophylaxis in any case. after the median follow-up period of months (range: - months), all patients were alive, and none of them had relapsed after the kir-cbt. although grade ii-iv gvhd was observed in patients ( %), it was controlled with prednisolone. chronic gvhd was not seen in any case. the present findings suggested that nk cell alloreactivity plays a role in preventing childhood myeloid leukemia and nbl relapse after kir-cbt. although our results are limited, this report provides novel data to support further investigations into the use of kir-cbt for the treatment of pediatric refractory malignant disease. disclosure of conflict of interest: none. impact of fcm-based minimal residual disease on transplant outcomes in patients with aml in hematological complete remission t oka, j kanda , k ohmori , m hishizawa , t kitano , t kondo , k yamashita it is reported that the presence of minimal residual disease (mrd) before hematopoietic stem cell transplantation (hsct) is associated with poor overall survival in patients with acute myelogenous leukemia (aml) in hematological complete remission (cr). we retrospectively analyzed the association between flowcytometry (fcm)-based detection of mrd and transplant outcomes. we included adult patients with aml in hematological cr, who underwent their first allogeneic hsct between april and may at kyoto university hospital. mrd of bone marrow before hsct was measured using fcm. to search for target antigens to detect mrd, threecolor fcm analyses were performed using a differential panel for every disease and patient, which allowed us to detect ⩾ . % of mrd. of the patients (median age: . , range: - ), patients were included in the mrd-negative group (mrd o . %), whereas were included in the mrd-positive group (mrd ⩾ . %). in the latter group, patients were included in the mrd-low group (mrd o . %), and were included in the mrd-high group (mrd ⩾ . %). there was no significant difference in the patient background between the mrd-negative and mrd-positive groups. the -year overall survival rates for the mrd-negative, mrd-low, and mrd-high groups were %, %, and %, respectively (p = . , figure ). in a multiple regression analysis, the mrd-high group was significantly associated with higher overall mortality than the mrd-negative group (mrd-low vs mrd-negative, hazard ration [hr] . , p = . ; mrd-high vs mrd-negative, hr . , po . ). the -year relapse rates for the mrdnegative, mrd-low, and mrd-high groups were %, , and %, respectively (p o . ). there were no significant differences in non-relapse mortality among the three groups. the analysis of fcm-based detection of mrd revealed that an mrd positivity of ⩾ . % was significantly associated with high risk of relapse and death even in patients with aml with hematological cr. the stronger consolidation or conditioning therapy before hsct based on mrd could improve transplant outcomes in these patients. [p ] disclosure of conflict of interest: none. post-transplant cyclophosphamide (ptcy) and megadose t cell depleted (tcd) haplohsct for tolerance induction f aversa , e bachar-lustig , n or-geva , y zlotnikov klionsky , l prezioso , s bonomini , a monti , i manfra , c schifano , s pratissoli , f lohr , r lamanna , v sgobba , n giuliani and y reisner hematology and bmt unit, university hospital of parma, italy; department of immunology, weizmann institute of science, rehovot, israel; neurology department, stanford school of medicine, stanford, california; radiotherapy unit, university hospital of modena, italy; radiotherapy unit, university hospital of modena, italy; genetic unit, university hospital of parma, italy and hematology and bmt unit, university hospital of parma, italy the use of ptcy is associated with reduced risk for gvhd in t cell replete nma haplo-hsct; however, this intervention is still not sufficiently safe to justify treatment of non-malignant diseases or as a platform for organ transplantation. experimental data: in a total of mice, we showed that combining the power of megadose tcd hsct with high dose ptcy (fig. a) , enables marked and durable chimerism following nma conditioning, while each modality alone was ineffective (figure a) . chimerism included all myeloid and lymphoid lineages, and lda analysis of alloreactive t cells revealed specific immune tolerance towards donor stimulators (fig. b) , also associated with acceptance of donor but not rd party skin. clinical trial: a similar protocol was developed for clinical use. the first patient, a yr old male with high-risk multiple myeloma in cr after autohsct, received megadose ( . x cd + cells/kg) cd /cd depleted ( . x cd +t cells/kg) haploidentical pbpcs after atg, fludarabine and gy single frcation tbi. ptcy was given to control both hvg and gvh reactions (fig. c) . hematopoietic engraftment was achieved at day + with over % donor type chimerism during the first months in the myeloid and b cell lineages. t cells during this period were predominantly of host type ( - % donor type), gradually increasing to - % at - months post transplant (fig. d) . the patient overcame cmv and subsequently ebv reactivation without any treatment (fig. e- g) . dextramer facs analysis revealed that cmv and ebv specific cd t cells were exclusively of host origin (fig. f- h) . at + months, cr and normal free light chain ratio were confirmed. the second patient, a year-old male with high risk heavily pretreated multiple myeloma (tandem auto-hsct, yr maintenance with lenalidomide, salvage therapy with vd) received a similar hsct ( . x cd + cells/kg, . x cd +t cells/kg). despite transient engraftment ( % donor cell on day + ), graft failure with autologous recovery ( . % donortype chimerism) was documented on day + . this may be due to the extended treatment ( yrs) with lenalidomide, but rejection cannot be excluded. after months, this patient tolerated a second haplo-hsct (different donor) after myeloablative conditioning (atg, treosulfan, thiotepa and fludarabine) and alfa/beta tcr/cd -depleted pbpcs. at month follow up, he shows no sign of gvhd, good immunological reconstitution, excellent quality of life, and remains in complete remission. collectively, our murine proof of concept data supported by clinical experience in the first high risk mm patient. the marked level of host t cells persisting over the first year after hsct can provide anti-viral immune protection until thymus-derived donor t cells are generated. avoiding additional post transplant immune suppression ensures a robust anti-viral immunity and a graft vs tumor effect. the rejection experienced by the nd patient, although corrected by a nd myeloablative tcd hsct, indicates that the conditioning must be fine-tuned to optimize engraftment in every patient. we are therefore testing, increasing tbi from gy to gy. further studies will determine the efficacy of this approach in elderly mm patients, in non-malignant hematopoietic diseases, or as a prelude for organ transplantation and cell therapy. over the last decade the addition of alemtuzumab to fludarabine-based reduced intensity conditioning regimen is common practice in the unrelated donor allograft setting. in recent years, however, its use has extended to reduced intensity hla-identical sibling donor allografts with the aim of providing an additional prophylaxis against gvhd. it is difficult to assess though whether this practice has any negative influence in the relapse rate or whether it has any net benefit or disadvantage in terms of overall survival. in this retrospective study we have analysed a historical cohort of patients [ males, females, mean age . ( - )] who s received a ric fully matched unrelated donor ( patients) or sibling donor ( ) hsct as consolidation treatment for hr aml in transplant centres in uk and greece. the conditioning regimen included fludarabine in all cases, together with melphalan and alemtuzumab( patients), busulphan and campath ( patients), busulphan and thiotepa ( patient), melphalan ( patients), busulphan with and without atg ( patients) total body irradiation ( cgy, patients). in total, patients received alemtuzumab ( mud mg alemtuzumab and sibling donor hsct recipients mg alemtuzumab) and patients ( mud and sibling donor hsct recipient) received atg with patients receiving t replete allografts. gvhd prophylaxis was ciclosporin for patients receiving alemtuzumab based or atg based regimen and ciclosporin with low dose methotrexate for t-replete allografts. the median follow up was . months (range - months).all but four patients were transplanted in cr overall, patients receiving conditioning without alemtuzumab suffered more frequent (po . ) and more severe (po . ) acute gvhd. this group, however, had a significantly (po . ) lower relapse rate. the overall survival remained unaffected. the subgroup of patients receiving allografts from mud had a clear benefit in terms of a lower incidence (p o . ) and severity (p o . ) of acute gvhd: none of the patientsreceiving alemtuzumab experienced grade iv agvhd, but up to / patients not receiving alemtuzumab suffered severe grade iv gvhd. however, the use of campath was associated with a significantly higher rate of relapse or progression of the aml (po . ), so that none of the mud recipients not having campath relapsed, while / patients having alemtuzumab relapsed. although none of these factors had a net impact on survival, there was a nonsignificant (p = . ) trend towards a higher survival in patients who received alemtuzumab. in the sibling donor allograft setting, alemtuzumab had no significant impact on the incidence of acute gvhd, relapse or survival. finally, in diseases where cytogenetic or molecular markers of high risk were available, our results showed a better overall survival (po . ) in ric alemtuzumab conditioning undergoing fully matched unrelated donor hsct, probably as a result of the protection against graft versus host disease while maintaining graft versus leukaemia effect. overall, alemtuzumab is a highly protective agent against agvhd in mud hsct recipients while it maintains the graft versus leukaemia effect.however it did not show any clear benefit of its use in the identical sibling donor setting. larger prospective studies are required in order to determine the need for this agent in this particular setting. disclosure of conflict of interest: none. blastic plasmacytoid dendritic cell neoplasm (bpdcn) is a rare disease which constitutes o % of all hematologic neoplasms annually. majority of bpdcn present with diverse skin involvement prior to leukemic dissemination, whereas a minority (~ %) have systemic involvement at diagnosis. there are no established therapies for bpdcn and most pts receive acute leukemia, myeloid or lymphoblastic, induction regimens; but responses are short-lived and prognosis is poor upon relapse. allogeneic hematopoietic cell transplantation (allo-hct) is offered to bpdcn cases based on small retrospective or registry case series. we retrospectively analyzed outcomes of bpdcn pts who received an allo-hct at transplant centers in the usa. a total of pts were eligible for analysis ( table ). the primary endpoint was overall survival (os). twenty patients (m = , %), median age of ( - ) yrs, received an allo-hct from a matched related (n = , %), matched unrelated (n = , %), mismatched-unrelated (n = , %), umbilical cord (n = , %) or haploidentical (n = , %) donor using myeloablative (mac) (n = , %) or reducedintensity (ric) (n = , %) conditioning. fifteen pts received hyper-cvad as pre-allograft therapy (front-line = , salvage = ). the majority (n = , %) were allografted in cr . median f/u for survivors was . ( . - . ) months. median time-to-neutrophil and platelet engraftments were ( - ) days and ( - ) days, respectively. five pts never dropped s platelet counts below /μl. three pts (mac = , ric = ) relapsed at , , and months, respectively. all relapsed with marrow involvement ( had also skin involved). mean os was . ( . - . ) months. one-year and -year os were % ( % ci = - %) and % ( % ci = - %), respectively. there was no difference in -year os when comparing mac versus. ric (hr = . ( % ci = . , . ), p = . ). median time to onset of acute gvhd was ( - ) days; grade ii-iv acute gvhd occured in cases. chronic gvhd was seen in cases (mild = , mod/severe = ). allo-hct is an effective therapy for bpdcn resulting in durable remissions. encouraging outcomes observed in this analysis may be explained by offering allo-hct early in the disease course and in the setting of complete remission. larger studies are needed to better understand risk factors for relapse to develop post-transplant strategies to improve outcomes. disclosure of conflict of interest: none. a risk-factor analysis for overall survival in patients with acute leukemia that relapse following t-replete haploidentical transplantation: on behalf of the acute leukemia working party of the european society for blood and marrow transplantation s piemontese , , m labopin , , f ciceri , , c schmid , , a ruggeri , , w arcese , z gulbas , y koc , j tischer , b bruno , w depei , d blaise , d beelen , g ehninger , a boumendil , , m houhou , , m mohty , and a nagler , relapse of acute leukemia is the leading cause of transplantation failure with devastating results. relapse post t-replete haploidentical transplantations (haplo-sct) is not well characterized. the objective of this study was to identify riskfactors for overall survival in patients with al that relapsed after a haplo-sct. from to , haplo-sct were performed in ebmt centers as first allogeneic transplantations for adults with acute leukemia. out of patients for whom we were able to receive updated data, relapsed and were included in this analysis. median follow-up among survivors was months after haplo-sct ( - ) and . months ( - ) after relapse. median time from haplo-sct to relapse was months ( d- m). diagnosis was acute myeloid leukemia (aml) in % and acute lymphoblastic leukemia (all) in % of the patients, respectively .fifty-two ( %) patients were transplanted in first complete remission (cr ), ( %) in cr or cr , while ( %) were transplanted in active disease. ric regimen was used in ( %) patients and ( %) received bone marrow as stem cell source. post-transplant cyclophosphamide (pt-cy) was used for graft-versus-host disease (gvhd) prophylaxis in patients ( %). fifty-two ( %) of the patients who relapsed post haplo-sct experienced previously acute gvhd and ( %) chronic gvhd post transplantation. treatment of relapse varied and included: none in ( %), ist withdrawal only in ( %), chemotherapy (ct) only in ( %), tyrosine-kinase inhibitor (tki) only in'( %), tki and ct in ( %), dli only in ( %), subsequent transplant in ( %), ct and dli in ( %), ct and subsequent transplant in ( %), tki ct and subsequent transplant in ( . %), dli and subsequent transplant in ( . %) patients. donors for second allogeneic transplant were unrelated (n = ), haploidentical (n = ) and cord blood (n = ). second transplant was performed in cr for patients and in relapse for patients. only patients who received a second haplo were alive at and months post second transplant. the majority of patients who received dli were in relapse at time of dli ( %), and % achieved cr after dli. os y after dli was %, patients being alive at a median time of mo ( - ) post dli. overall, the one-year overall survival (os) following relapse was % ( % ci: . - . ). in univariate analysis disease status at haplo-sct (cr vs active disease), cytogenetics (good/intermediate vs poor) and median time from haplo-sct to relapse ( or o . months) were associated to a higher os at one year after relapse: % (p = . ), % (p = . ) and % (p o - ), respectively. in multivariate analysis complete remission at haplo-sct (p = . ; hr . ; ci: . - . ) and time from haplo-sct to relapse higher than . months (p = . ; hr . ; ci: . - . ) were risk factors for a higher os after relapse. in the patients transplanted in cr and relapsing more than month after haplo, and y os were respectively % and %. these findings suggest that similar to other transplantation setting os for acute leukemia that relapse post haplo-sct is dismal. disease status at transplant and time from transplant to relapse are the two important prognostic factors that can predict somewhat better survival. indication for second transplant should be carefully evaluated. integrations with novel therapies are in unmet need to prevent and treat relapse post haplo-sct. disclosure of conflict of interest: none. patients (pts) with aml who relapse after autologous stem cell transplantation (asct) have a dismal outcome but some can be rescued with an allogeneic transplantation (allohsct). yet, available evidence presently stems from analyses of limited patient numbers. we decided to analyze the ebmt registry to evaluate the outcome and determine the prognostic factors in a large series of such pts. the ebmt registry was screened for adult pts with de novo aml (non-apl) who received an allograft in cr or first relapse ( - ) after being autografted in cr . pts receiving ex vivo t cell depletion (tcd) were included only if they received a haploidentical allohsct. inclusion criteria were met by pts ( % female, median age [range - ] years). median time from asct to relapse was (range . - , iqr . - . ) months. at allohsct, pts were in st relapse ( %) or cr ( %). donors were matched sibling ( %), unrelated ( %), haploidentical ( %), or cord blood ( %), respectively. conditioning was myeloablative in % and reduced intensity in % of the pts, respectively. the median follow up was months (range o - months). at years post allograft (figure), leukemia free survival ( - ] of the pts. all factors significantly associated with ⩾ endpoint in univariate analysis were entered in a multivariate cox regression model (table ) . ri was lower in pts transplanted in cr rather than in relapse ( . % vs . %; hr . , p = . ) and in pts who relapsed later ( months, median value) as opposed to those who relapsed early post asct ( . % vs . %; hr (per month) . , p o - ). ri was lower in pts transplanted with an unrelated donor (ud) in comparison to those transplanted from a matched sibling donor ( . % vs . %; msd, hr: . , p o - ). patient age, poor cytogenetics, transplantation in relapse, previous tbi for asct, myeloablative conditioning (mac) vs reduced intensity (ric) and ud, haplo or cbt vs msd all significantly increased nrm. lfs was significantly better in pts with good risk ( . %) than in pts with intermediate risk or poor risk cytogenetics ( %; hr . , p = . ) or in pts who relapsed late (per month: hr . , p = . ) post asct. lfs was worse in pts who previously had received tbi ( % vs %; hr = . ; po - ). the same prognostic factors were significant for os. haploidentical (hr . , p = - ) and cord blood (hr . , p = . ) transplants resulted in lower os than those from msd. finally, date of transplant significantly influenced os which was higher in pts transplanted after january vs those allografted before; . % vs . %, hr (per year) . , p = . ). about one third of adult patients with aml who relapse post asct can be rescued with an allogeneic transplantation, especially if the duration of persisting cr post asct is long and no tbi was received in the past. transplantation from an msd while in cr rather than at relapse offers the best outcome. disclosure of conflict of interest: none. high incidences of graft-versus-host disease (gvhd) and relapse have seriously impeded the widespread application of haploidentical hematopoietic stem cell transplantation (haplo-hsct) for high-risk acute leukemia lacking conventional hla-matched donors. one hundred and ten high-risk acute leukemia patients underwent haplo-hsct with idarubicin (ida) intensified conditioning regimen (ida intensified bucy for acute myelocytic leukemia (aml) and ida intensified tbi-cy for acute lymphoblastic leukemia (all)). for donor-recipient hla / or / transplant, we separately administered a total of mg/kg or mg/kg antithymocyte globulin (atg) and basiliximab for gvhd prophylaxis. all enrolled patients were observed longitudinally until death or lost to follow-up. the -day cumulative incidences of Ⅱ-Ⅳ and Ⅲ-Ⅳ agvhd for all patients were . %, . %, respectively. the -year cumulative incidence of extensive cgvhd was . %. the relapse rate was . %. the -year probability of overall survival (os) reached . %. the patients in non-complete remission (nr) showed significantly higher relapse and worse survival than complete remission (cr) minimal residual disease (mrd) (-) and cr mrd (+) patients. however, the relapse, y-os and disease-free survival (dfs) of cr mrd (-) did not differ from cr mrd (+) patients, indicating our intensified transplant technique could overcome the poor prognosis of mrd. for whatever aml or all patients, the relapse rates, agvhd, cgvhd and the estimated -year os and dfs between two atg group were equivalent, except that all patients in atg mg/kg experienced higher relapse ( . % vs . %, p = . ). although the incidence of cytomegalovirus (cmv) reactivation in atg mg/kg and mg/kg was . %, . %, the average episodes of cmv reactivation were remarkably [p ] higher in mg/kg. our ida intensified haplo-hsct technique could improve the outcome of high-risk acute leukemia and could be recommended as a good alternate for patients lacking hla-matched sibling donors. diagnosed secondary aml were randomized : to cpx- or standard + therapy. cpx- induction was units/m on days , , (first induction) and days , (reinduction); + first induction was cytarabine mg/m /day × days and daunorubicin mg/m on days , , , and reinduction was cytarabine mg/m /day × days and daunorubicin mg/ m on days , . a dynamic allocation procedure stratified patients by age group ( - or - years) for each study arm. patients with complete response (cr) or cr with incomplete platelet or neutrophil recovery were considered for allogeneic hct, based on institutional criteria. overall survival (os) landmarked at the time of hct was assessed. a total of patients were enrolled on the induction trial. . patient and aml characteristics in the hct age subgroups were generally similar between arms. in both age subgroups of patients receiving hct, median os was longer in the cpx- arm than in the + arm (table ). in the - group, serious adverse events (saes) prior to hct in the cpx- and + arms occurred in % and % of patients, respectively; in the - group, in % and %, respectively. the most common sae was febrile neutropenia (cpx- , . %; + , . %), occurring in all age groups. relapse after allogeneic haematopoietic stem cell transplant (allo-hsct) for acute myeloid leukaemia (aml) and myelodysplastic syndrome (mds) remains the main cause of treatment failure. it is associated with dismal prognosis and short survival. proposed salvage strategies are tapering of immunosuppressive therapy, re-induction with chemotherapy and consolidation with donor lymphocyte infusion (dli) or second allo-hsct, although, results remain disappointing. azacitidine (aza) and dli has proved to be an effective and well-tolerated outpatient approach in this setting, and results in at least temporary disease control in the majority of patients, thus, representing a valuable alternative to current treatments. between january and november , patients with relapsed aml or mds after allo-hsct were treated with subcutaneous aza mg/m days - every days and escalating doses of dli if feasible at manchester royal infirmary, uk. aza was continued until cr or disease progression. patients characteristics: median age (range - ) years, % males, diagnoses were aml (n = ) and mds (n = ). five ( %) patients had either monosomal or complex karyotype. fifty percent of patients were in cr before transplant, . % in cr , . % had a partial response and % did not receive any chemotherapy before the transplant. fifteen out of received fludarabine-base reduced intensity conditioning regimen and all but one had a t-cell depleted graft. at relapse % had mixed donor chimerism. median time to relapse was . (range - ) months after allo-hsct. with a median follow up of . (range - ) months a median of (range - ) courses of aza were administered and median of (range - ) dli were infused. doses of dli were administered starting at . x /kg and escalating by log . aza and dli infusions were well tolerated; only two patients withdrew due to intolerance. seven patients were admitted at least once due to infections ( %) or progressive disease. only two patients developed mild gvhd grade . complete remission was achieved in . % patients and stable disease in %. patients in cr had full donor chimerism. median overall survival for patients in cr was months compared to months for those who did not respond (p = . ). patients with more than % blasts on bone marrow at time of relapse after allo-hsct had a worse outcome than those with less than % blasts ( months and months respectively, p = . ). no differences were seen when compared time to relapse ( o months vs ⩾ months) s and outcome, or disease and overall response, although numbers in this series are small. image/graph: overall survival following azacitidine and dli, patients in complete remission, stable disease and disease progression. azacitidine and dli can provide long term remissions in patients with relapsed aml/mds post allo-hsct with low toxicity. lower disease burden at relapse carries better outcomes. low rates of gvhd are seen following azacitidine and dli most likely showing the immunomodulatory effect of azacitidine described by other groups. acute myeloid leukemia (aml) is a frequent complication in patients affected by telomere maintenance disorders ('telomeropathies') such as dyskeratosis congenita (dkc). treatment of aml in dkc patients by chemotherapy and hematopoietic stem cell transplantation is characterized by frequent remission failure, high organ toxicity and poor outcome. a -yearold patient with aml was admitted to our hospital in december . he had been treated with cycles beacopp for hodgkin´s lymphoma (hl) in . on admission, the patient presented clinical signs of premature aging with hair greying and lack of fully recovered hair growth after chemotherapy (cx) for hl. flow-fish analysis revealed tl below the % percentile within leucocytes in line with the suspected diagnosis of telomeropathy. retrospective tl analysis by confocal q-fish from bm at hl diagnosis confirmed short tl before the start of any chemotherapy. he received standard aml induction cx ( + ), but follow-up revealed persistence of aml. salvage cx with flag-ida was applied resulting in partial remission with only weak regeneration of normal hematopoiesis. the patient received an allogeneic stem cell transplantation (asct) after conditioning with mg/m melphalan and fludarabin from his hlamatched brother whose tl was found to be normal. after asct, he developed sinusoidal obstructive syndrome and progressive liver failure treated with defibrotide and he was admitted to icu for sepsis. leucocyte count showed sufficient engraftment on day ; however, liver function recovered only partially. during critical care treatment, the patient showed cardiomyopathy, renal failure and extensive wound healing problems without epithelial proliferation indicative of severe replicative exhaustion. finally, he died due to sepsis with acute liver failure on day after asct. aml arising from dkc is a rare event with substantial impact on patients´prognosis. therapy remains challenging due to poor bm function and high risk of organ toxicity, especially liver failure and lung fibrosis. dose reduction of alkylating agents and avoidance of total body irradiation are necessary in conditioning prior to asct in patients with dkc and aml, however no clear data or recommendations exist for the management of these patients. tl screening can help to identify patients with suspected dkc related bm failure or aml and to identify family donors without telomeropathy. physicians should be aware of possible dkc related aml, especially in familial cases of aml or bone marrow failure, impaired or prolonged recovery following cytoreductive treatment or coincidence of solid (e.g. oral cavity carcinomas) and hematological malignancies. disclosure of conflict of interest: none. chronic graft-versus-host disease and donor lymphocyte infusions in patients with non-de novo acute myeloid leukemia or advanced myelodysplastic syndromes after allogeneic stem cell transplantation pg hemmati , k pfeifer , lg vuong , cf jehn , p le coutre , b dörken and r arnold medizinische klinik mit schwerpunkt hämatologie, onkologie und tumorimmunologie, charité-universitätsmedizin berlin, campus virchow-klinikum, berlin, deutschland aml with myelodysplasia-related changes and therapy-related aml (taml), collectively termed secondary aml (saml) in daily clinical routine, represent distinct subgroups in the revised who classification of myeloid neoplasm and leukemias. as compared to de novo-aml, saml is associated with a poor survival when using conventional chemotherapy approaches. this is mainly due to unfavorable cytogenetics, older age and/or the presence of comorbidities as well as poor response to induction therapy. furthermore, cumulative organ toxicity resulting from treatment of the antecedent solid malignancy in patients with therapy-related disease has to be taken into account. allogeneic stem cell transplantation (allosct) represents the only option to achieve long-term disease control and definitive cure. we retrospectively analyzed patients with saml or advanced mds (eb- according to who) transplanted at our center between and . at the time of allosct, patients ( %) were in complete hematologic remission (chr), whereas patients ( %) had active disease. cytogenetic risk was categorized according to the swog/ecog classification and was favorable (n = ; %), intermediate (n = ; %), unfavorable (n = ; %), or unknown/undetermined (n = ; %). standard myeloablative conditioning (mac) using gy total body irradiation (tbi) and cyclophosphamide was used in patients ( %), whereas fludarabin/busulfan/atg-based reduced intensity conditioning (ric) was applied in patients ( %). grafts were from related (n = ; %) or unrelated (matched: n = ; % or mismatched: n = ; %) donors. the median follow-up of the surviving patients was ( - ) months. a graft failure occurred in / patients ( %). at last day of follow-up / patients ( %) were alive and in chr. relapse occurred in / patients ( %) after a median interval of . (range: . - ) months. cause of death were either relapse or nrm (gvhd and/or infections) in / patients ( %) or / patients ( %). at , , , and years after allosct overall survival (os) or disease-free survival (dfs) of the entire cohort was %, %, %, and % or %, %, %, and %, respectively. at the same time points, the cumulative incidence of relapse (ci-r) or non-relapse mortality (ci-nrm) was %, %, %, and % or %, %, %, and %, respectively. extensive uni-und multivariate analyses revealed a number of factors associated with inferior outcome, e.g. poor-risk cytogenetics, the presence of taml, advanced age, reduced physical performance, and comorbidities, whereas donor type (unrelated versus unrelated), and remission status had no significant impact on overall outcome. furthermore, the development of gvhd, especially the presence of cgvhd, and the use of donor-lymphocyte infusions (dli), either in a prophylactic or pre-emptive setting, were identified as independent predictors for a reduced relapse incidence, which in turn, led to an improved os and dfs. our results indicate that allosct represents an important treatment option for patients with saml. however, a relapse rate of % at months prompts the development of novel approaches to prevent early disease recurrence. strategies to augment the graft-versus-leukemia (gvl) effect of allosct may help to improve the results. disclosure of conflict of interest: none. myeloid sarcoma (ms) is a rare hematologic myeloid neoplasm that can involve any site of the body. it can occur as an exclusively extramedullary form or it can be associated with an acute myeloid leukemia (aml), a chronic myeloproliferative neoplasm (mpn) or a myelodysplastic syndrome (mds) at onset or at relapse. the rarity of ms does not enable prospective clinical trials and therefore a specific multicenter register can be useful for the clinical and biological studies of this rare disease. we report the clinical characteristics and outcome of histologically confirmed ms, diagnosed and treated in italian hematological centers in the last years. the patient's median age was years. there were / de novo extramedullary ms, / de novo aml-related ms and / were secondary aml-related ms. the most common extramedullary anatomic sites of disease were: skin, lymph nodes and soft tissues. forty-three patients ( %) underwent a program of intensive chemotherapy including flai, hdac-ida, hypercvad and mec schemes, with a cr rate of % ( / ). twenty-two ( %) patients underwent allogeneic sct, from a mud, from an hla-identical sibling donor and from an haploidentical donor. the median os of the whole population ( pts) was . months. the os probability at , and years was %, % and %, respectively. the os was better in patients that underwent an intensive therapeutic program (median os: months vs months). among the intensively treated patients, in univariate analysis, the os was better in young patients (p = . ), in patients that underwent allo-sct (p = . ) and in patients that achieved a cr during treatment (p = . ), and was worse in pts with secondary aml-related ms (p = . ). age, response to intensive chemotherapy and allo-sct were the only three variables that significantly influenced dfs (p = . , p = . and p = . , respectively). in multivariable analysis, allo-sct and response to intensive chemotherapy remained significant in predicting a better os (p = . and p = . , respectively), and response to intensive chemotherapy was the only significant variable in predicting dfs (p = . ). after allo-sct we observe a survival advantage in patients who achieved a pre-transplant cr (p = . ) and in those who developed a chronic gvhd (p = . ). patients with ms, both with de novo and secondary forms, still have a very unfavorable outcome and require an intensive therapeutic program, that includes allo-sct, whenever possible. the outcome after allo-sct is positively influenced by the development of chronic gvhd suggesting a graft versus ms effect. disclosure of conflict of interest: none. relapse of acute lymphoblastic leukemia (all) after allogeneic stem cell transplantation (sct) is associated with poor prognosis. blinatumomab may enhance the efficacy of donor lymphocyte infusions (dli) in this specific situation but data on the concurrent use of dli and blinatumomab are sparse. the patient presented here was diagnosed with standard risk pre-b-all (presence of t( ; ); bcr-abl and cd negative) at the age of . during treatment according to the german multicenter all-study group (gmall) protocol he presented with molecular relapse and months after initial diagnosis he received a tbi-based myeloablative sct from an unrelated hla-identical ( / ) donor. post sct he was negative for minimal residual disease (mrd) with % donor engraftment. given the high relapse risk he received prophylactic dli without occurrence of graft-versus-host disease (gvhd). one year after st sct he presented with an extramedullary (testes) and molecular relapse. after remission induction resulting in negative mrd he received a nd sct from an alternative, hlaidentical ( / ) donor after reduced intensity conditioning. this again resulted in negative mrd with % donor chimerism without any gvhd. six months after nd sct he presented with bone marrow relapse. we decided on the concurrent use of blinatumomab and dli. the first cycle of blinatumomab was initiated at standard dose including dose escalation without relevant toxicities. on day of the nd cycle, i. e. in the infusion-free interval before the rd cycle the patient received the first dli at x cd /kg. no toxicities or gvhd occurred. the rd cycle of blinatumomab was initiated and a second dli at . x cd /kg was applied on day of the rd cycle. on day of the rd cycle, i. e. day after nd dli the patient presented with signs of overlap gvhd (mouth, skin) and topical steroids were started. upon progression of clinical gvhd systemic steroids were initiated with immediate response. steroids were rapidly tapered and a th cycle of blinatumomab was started. gvhd did not recur. current staging after the th cycle blinatumomab, i.e. on day + after nd sct and months after initiation of blinatumomab treatment revealed complete remission with negative mrd, % donor chimerism and no signs of extramedullary relapse. counts of cd -cells at that time point were /μl. no relevant infections or relevant blinatumomab-associated toxicities were present during the entire course after the nd sct. in this case concurrent treatment of blinatumomab and dli resulted in the longest disease-free interval for our patient compared to preceding chemotherapy or dli alone. together with the small number of reported cases (ueda et al.) this supports the concept of concurrent blinatumomab and dli as an effective post sct treatment. the objective of the study is to evaluate the clinical efficacy and safety of decitabine (dac) in combination with haag regimen [homoharringtonine (hht), cytarabine (ara-c), doxorubicin (acla) and recombinant human granulocyte colony stimulating factor (g-csf)] for advanced patients with acute myeloid leukemia (aml). thirty-six patients with advanced aml receiving dac combined with haag chemotherapy in our center from december to august were enrolled in this study. eighteen of them were refractory or relapsed aml, and another patients were those who didn't achieve complete remission (cr) after a course of induction chemotherapy. the therapeutic responses, side effects and longtime survival were retrospectively analyzed. after a course of treatment, the rate of cr and partial response (pr) was . % ( / ) and . % ( / ) respectively, while the overall response rate (orr) was . % ( / ) in the cohort. for the patients with refractory or relapse aml, cr was . % ( / ), pr was . % ( / ), and orr was . % ( / ). while for the other not getting cr after a course of induction chemotherapy, cr was . % ( / ), pr was . % ( / ), and orr was . % ( / ). grade hematological toxicities were observed in all patients, and . % cases experienced infection. and all non hematological side effects were mild and well-tolerated. with a median follow-up of . ( . ~ . ) months, the -year overall survival (os) rate was . %, . % for the refractory or relapsed aml patients, and . % for those not achieving cr after a course of induction chemotherapy. the difference was significantly (p = . ). conclusion dac combined with haag regimen is safe and effective salvage treatment for advanced stage aml patients. disclosure of conflict of interest: none. aml patients harboring flt -itd mutation are associated with decreased survival compared to patients without flt -itd mutation. nevertheless, whether flt-itd mutation also has negative impact on the post-transplant survival is less clear. for flt -itd mutated aml, a decreased leukemia-free survival (lfs) after allogeneic hsct was observed in ebmt analysis but not cibmtr. in this study, unlike studies of ebmt or cibmtr which only pre-specified populations of patients were analyzed (cr in ebmt, cr +cr in cibmtr), we examined the prognostic impact of flt -itd mutation on post-transplant outcome of "all" the adult aml patients reported to taiwan bone marrow transplant registry (tbmtr). tbmtr is a research collaboration affiliated to the taiwan society of blood and bone marrow transplantation. it comprises all the transplantation centers in taiwan that contribute detailed data on hsct. adults aged ⩾ years with a diagnosis of aml and with known flt-itd mutation status in the registry were included. patient characteristics and transplant outcome following allogeneic hsct for flt -itd mutated and nonmutated aml were compared. kaplan-meier estimates were used to calculate the probability of lfs and overall survival (os). multivariable analyses for lfs and os were performed using cox proportional hazards model. patients who met the eligibility criteria were enrolled for analysis. the median follow-up of survivors was months. of the patients, ( . %) were positive and ( . %) were negative for flt -itd mutation. flt -mutated patients had significantly more transplantation at cr ( . %), shorter time interval between diagnosis and hsct ( . months), and higher wbc count at diagnosis ( . × /l) comparing to patients without flt mutation ( . % at cr , . months from diagnosis to hsct, and . × /l wbc count at diagnosis). significant more flt mutated patients had intermediate-risk ( . %) and normal ( . %) karyotype at diagnosis. the age, donor type, stem cell source, conditioning regimen, and atg use were not significant different between flt -mutated and non-mutated patients. of the whole population, flt mutation status did not negatively impact the transplant outcome ( years os for flt mutated and non-mutated patients: . % vs %, log rank p = . ; years lfs for flt mutated and non-mutated patients: . % vs . %, log rank p = . ). when different pre-transplant conditions (cr , subsequent cr, and no cr) were analyzed separately, flt -itd mutation status is still not a significant prognostic factor of os and lfs for patients in cr (equally good) and no cr (equally bad). however, for patients in subsequent cr, flt -itd mutation is the only significant factor predicting poor os and lfs in multi-variable analysis (median os and lfs for flt mutated and nonmutated patients: vs days, log rank p = . ; vs days, log rank po . respectively). the incidence of non-relapse mortality, grade / acute gvhd and extensive chronic gvhd is comparable between flt -mutated and nonmutated patients. flt -itd mutation is a significant and strong predictor of poor survival for aml patients in subsequent cr at hsct. for flt -itd non-mutated aml, a sizable portion of patients can have disease free survival after allogeneic hsct at subsequent cr. however, allogeneic hsct at cr should be strongly recommended for flt -itd mutated aml. [p ] disclosure of conflict of interest: none. allogeneic stem cell transplantation (asct) is a curative strategy in acute myeloblastic leukemia (aml) and myelodysplastic syndrome (mds). however, relapse keeps being the main cause of treatment failure. extramedullary relapse (er) is a rare event and its management is not well standardized. we retrospectively analyzed patients who received asct from to and developed er in our centre. we performed a descriptive study to analyze characteristic of these patients, post-relapse treatment and survival. statistic analysis was performed using spss v. . we found a total of patients with er, one of them with er after consecutive asct, so we analyzed cases of er. patient and transplant characteristics are summarized in table . at day + , % of patients were in complete response (cr). er occurred after a median of ( - ) months post-asct. eleven patients ( %) presented with a bone marrow relapse concomitant with the er. er affected central nervous system (cns) in patients ( . %), bone in patients ( %), skin or soft tissue in patients ( . %), mama in patients ( . %), ocular globe in patients ( . %) and teste in patients ( . %). two of them presented with multiple sites affected. between the patients who developed cns relapse, of them had received intrathecal prophylaxis. regarding post-er management, immune modulation was conducted in patients (immunosupression tapering in , donor lymphocyte infusions in and both strategies in ). all patients except one received systemic treatment (salvage chemotherapy in , azacitidine in , low dose arac in and atra in patient with a promyelocytic leukemia). together with systemic treatment, received radiotherapy and intrathecal therapy was used in all patients with cns involvement. response: out patients treated, ( . %) achieved cr and ( . %) progressed. two responding patients received a nd asct. after a median follow-up of months ( - ), patients are alive and disease free, with an estimated overall survival of % at years. patients receiving salvage chemotherapy followed or not by a nd asct experienced a significantly better os than those receiving other therapies (median os vs months; p = . ). patients with bone marrow involvement at relapse show a worse prognosis (median os vs months; p = . ) although not statistically significant due to small number of patients (image ). ten patients died due to disease progression. er must be considered in patients receiving an asct in case of organ symptoms. patients can be rescued with salvage chemotherapy followed or not by a nd asct achieving good results in terms of long term os. it seems that involvement of bone marrow at relapse confers a worse prognosis, what should be confirmed in a larger series of patients. [p ] disclosure of conflict of interest: none. flag-ida regimen as bridge therapy to allotransplant in refractory/relapsed aml patients: a single-center experience c pasciolla, m delia, d pastore, p carluccio, a ricco, a russo rossi, a mestice, f albano and g specchia university of bari, italy although treatment outcome in acute myeloid leukemia (aml) adult patient has improved over the past decade, relapse still occurs in up to - % of cases. furthermore, - % of patients fail to achieve complete remission (cr) because of treatment-resistance. the management of primary refractory and/or relapsed disease remainschallenging for clinicians. in our study, we reviewed the outcome of refractory and/or relapsed aml patients who underwent salvage therapy with the flag-ida regimen between and at our institution. the study aim was to determine the efficacy of the flag-ida regimen in order to clarify which variables (who ps, ldh, bone marrow, peripheral blood blasts and platelets counts, white blood cells (wbc), pmn, molecular-cytogentic risk, duration of response and relapsed or refractory disease), present before starting flag-ida treatment, might have an impact both on cr and on os. we analyzed consecutive adult patients ( males, females; median age years, range - ) with newly diagnosed acute myeloid leukemia refractory to standard induction regimens or relapsed after cr, who received the flag-ida protocol as salvage therapy between january and december . sixty-eight of the patients ( %) were in first relapse, forty-seven patients ( %) were refractory to conventional chemotherapy. median wbc count before salvage therapy was . x /l (range . - ). median bone marrow and peripheral blasts counts were and %, respectively; median platelets count was x e /μl. according to the fab classification, patients had m , m , m , m , m , m , had biphenotype acute leukemia. according to molecular-cytogenetic risk stratification ( %), ( %) and ( %) patients belonged to poor, intermediate and good risk group, respectively. sixty-nine of patients ( %) achieved complete remission (cr); forty-seven %) patients were refractory to the salvage therapy. in multivariable analysis, variables with positive impact on response rate were lower wbc counts (o e /μl, p = . ), higher platlets counts ( x e /μl, p = . ), molecular-cytogenetic risk (p = . ), duration of response in relapsed aml (p = . ) and relapsed rather than primary refractory disease (p = . ), respectively. median os was months (m). cox regression analysis confirmed that both higher platlets counts, p = . ( ( x e /μl) vs m (o x e ul), log rank, p = . ) and relapsed disease, p = . ( (relapsed) vs m (refractory), gehan-breslow, p = . ) correlated with better survival. of note, molecular-cytogenetic risk evaluated before starting treatment was associated with cr, while no correlation was found with os. our data seem to confirm the value of flag-ida in relapsed amland may suggest its best usage as bridge-therapy in patients awaiting allotransplantation. disclosure of conflict of interest: none. s leukemia relapse is the major cause of death in patients received allogeneic hematopoietic stem cell transplantation (allo-hsct). the precise etiological mechanisms of leukemia relapse remain unclear. both leukemia cells themselves and hematogenesis micro-environment are involved in the relapse event. in our previous study, we reported a case of donor derived relapse of acute myeloid leukemia (aml) after allo-hsct. the patient and his donor-sister both harbored a germline mutation(c. - dup) in cebpa gene. donor hematopoietic cells transformed to aml by developing two somatic cebpa mutations ( dupc and - dup) in the patient's microenvironment. hence we suspect that - dup mutation of cebpa gene may altered hematopoiesis microenvironment and increased the survival of aml cells. to conform our hypothesis, we transfected mesenchyme stem cells (mscs) with cebpa - dup or wide type and took vector as control. aml cell line hl cells were co-cultured with transfected mscs and then treated with ng/ml doxorubicin. apoptosis and cell cycle were detected at day . mscs protected hl cells from toxicity of doxorubicin. this protection was enhanced by overexpression of cebpa - dup . apoptosis rates of hl cells in group of msc-vector and msc-cebpa - dup were . ± . % vs . ± . % (p＜ . ). a larger part of hl cells remains quiescent with s higher rate of g /g phase in msc-cebpa - dup group, which may reduce the sensibility of hl cells to doxorubicin. to explore mechanisms involved in the alteration of microenvironment, we performed rna sequence with each group of mscs. we found that col a , col a and col a were upregulated in msc-cebpa - dup group compared with msc-cebpa wt group (col a :cebpa wt vs cebpa - dup was . vs . , p = . e- ; col a :cebpa wt vs cebpa - dup was . vs . , p = . e- ; col a : cebpa wt vs cebpa - dup was . vs . , p = . e- ). furthermore, we found that ddit and herpud genes, which were important factors in cellular unfolded protein response(upr) and to topologically incorrect protein, failed to augment in cebpa - dup group (ddit : vector vs cebpa wt the cure rate of childhood acute lymphoblastic leukemia (all) has improved considerably and approaches % today. however, the outcomes of patients who suffer from leukemic relapse remain unsatisfactory. despite the high cure rate of children and adolescents with all a subgroup of patients benefit from allogeneic hsct. allo hsct remains the standard treatment for intermediate/high risk aml patients. patients, all = and aml = age to years with median age years, m/f = / (m/f all = / , aml = / ) underwent sct in our hospital (from to ). fifty-eight patients transplanted allo hsct and pt aml auto hsct. conditioning regimens consisted of busulfan (iv) +cyclophosphamide for allo and cyclophosphamide + vp +cytarabine for auto hsct. peripheral blood (pb) was the source of progenitor cells in patients, bone marrow (bm) in patients and cord blood in one patient. in allo hsct, patient transplanted / matched and patients / matched. gvhd prophylaxis regimen was cyclosporine + mtx. all patients engrafted. in allogeneic pbsct all patients' median time to absolute neutrophil count (anc) . × /l was days, and the median time to platelet count × was days vs and days in allo bm all patients. in allogeneic pbsct aml patients median time to anc . × /l was days, and the median time to platelet count × was days. (all patients with aml transplanted with pb). at present pts are alive ( all, aml) and pts died due to ards, vod, hemorrhagic stroke, sepsis and relapse. trm was % at days. median time of death after transplantation was days in all and in aml. in allo pbsct all patients hospitalization period were days vs in allo bm all patients. acute gvhd appeared in % pts. chronic gvhd appeared in % pts. with a median follow-up of months ( - months) after transplant the event-free survival were % and four years overall survival % in all patients. a median follow-up of . months ( - months) after transplant the event-free survival were % and three years overall survival % in aml patients. hematopoietic stem cell transplantation can lead to durable remissions in children and adolescents with leukemia and increase in survival of children. pbsct in childhood all was consistent with significant faster anc and platelet recovery in allogeneic pbsct, hospitalization was shorter. longer follow-up is required to evaluate fully efficacy and long-term results. disclosure of conflict of interest: none. group hla-c /c subtypes were defined as previously practiced. median age of patients was , % of them were male. allo-hscts were performed from % unrelated donors vs % related donors. remission status was detected in % of patients whereas % had active disease pre-transplant. stem cell sources were as follows: % peripheral blood, % bone marrow, % cord blood, % bone marrow plus peripheral blood. the most frequent fab subtype was aml-m . patients were grouped by hla-c status: % c /c homozygote, % c /c heterozygote and % c /c homozygote. the frequency of hla c donor/recipient mismatch allo-hscts was %. relapse was detected in % of patients. the relapse risk was significantly lower in c /c homozygote patients compared to c /c homozygotes ( % vs %, p = . ). lfs was similar between c /c homozygote group and c /c homozygote group (p = . ) (figure ). in multivariate analysis (age, sex, remission status, related/ unrelated transplant, aml subtype), lfs was increased by pre-transplant remission status (p year). there was no difference detected between -years os in c /c homozygote group and other groups ( % vs %, p = . ) (figure ) . when similar analysis were repeated with donor hla type results were not significant. our results confirm two earlier published reports on aml and all. even in the absence of kir genotyping, hla group c has a protective effect. if hla matched donor is not possible a donor-recipient hla-c mismatch favoring c to c may be preferable. disclosure of conflict of interest: none. immunomodulatory kits do not induce aml-blasts' proliferation ex vivo: ipo- is an appropriate and reliable marker to detect and quantify proliferating blasts c plett, dc amberger, a rabe, d deen, z stankova, a hirn, y vokac, j-o werner, j schmohl, d krämer, a rank, c schmid and h schmetzer aml-blasts can be converted to dcleu by immunomodulatory 'kits' (ex vivo). t-cells' energy can be overcome after stimulation with dc/dcleu and results in antileukemic reactivity. a potential induction of blast-proliferation (e.g. by immunomodulatory kit-application in vivo) in aml-pts has to be excluded. kits containing combinations of gm-csf with - additional factors (pge- / , picibanil, ifnα, tnfα, calciumionophore) were studied with respect to the generation of dc/dcleu from blasts, mediation of antileukemic reactivity (after dc/dcleu-stimulation) and with respect to their potential to induce blast-proliferation in a whole blood (wb) culture-system. we studied different markers (ipo- , ki- , cd ) and quantified blast proliferation before/after culture. we correlated findings with (ex vivo) antileukemic functionality, with disease-entities and the course of the disease. dc-generation: we could generate dc/dcleu regularly from wb culture from aml-pts. detection of blast proliferation: Ø . % (range - %) of uncultured blasts expressed ipo- , . % ( - %) cd , . % ( - %) ki- . induction of blast proliferation: pooling all results we found lowest amounts of proliferating blasts after culture with kit i (gm-csf+picibanil, % ± . ), kit k (gm-csf+pge , . % ± . ), kit m (gm-csf+pge , . % ± . ). amounts of proliferating blasts were lower compared to uncultured cells. highest expression of proliferating blasts was found with ipo- followed by cd and ki- .we found few individual aml-samples with increased blast-proliferation after ex vivo kit-culture. antileukemic activity: t-cells stimulated with dcleu (generated with kits) improved antileukemic activity. correlations between blast-proliferation and antileukemic activity will be presented. clinical correlations: pts with bad (vs good) cytogenetic risk were characterized by higher proportions of proliferating blasts in uncultured blasts; in some pts with iron-deficiency anemia (ida) proportions of cd +unculured blasts were lower than of ipo- /ki- + blasts. ipo- is a stable marker to be used to quantify proliferating blasts in aml-pts. cd is also a good marker, although not suitable for some pts with ida, ki- is no reliable marker for every given pt. subtypes of pts correlated with proportions of proliferating blasts. in general kit treatment of blasts did only exceptionally induce blast proliferation ex vivo. in general lowest risk for blast proliferation was seen after culture with kit i, k and m. t-cellstimulation with dc/dcleu generated after kit-treatment resulted regularly in antileukemic reactivity. we conclude that an in vivo treatment of aml-pts with kits i, k or m might be safe (no induction of blast proliferation). disclosure of conflict of interest: none. the occurrence of additional cytogenetic abnormalities (acas) is common in philadelphia chromosome-positive acute lymphoblastic leukemia (ph+ all), but is of unknown significance in the tyrosine kinase inhibitor era. recent study [aldoss et al., ] has revealed the acas appear to have a significant deleterious effect on outcomes post-hsct in adult ph+ all patients only. we retrospectively analyzed data from adult and pediatric patients with ph+ all who had undergone allogeneic hematopoietic stem cell transplantation (allo-hsct) at our university between and . among patients with ph+ all, patients had available data on conventional cytogenetics before allo-hsct. all patients and transplant characteristics are listed in table i . thirty-three of patients ( %) had isolated t( ; ). acas were revealed in / ( %) pts, including / ( %) pts with ⩾ cytogenetic abnormalities (with complex karyotype, ck). the median follow-up was ( - ) days. overall survival (os) and event free survival (efs) were % ( % ci - ) and % ( % ci - ) at years, respectively. in univariate analysis, prognostic factors associated with increased os and efs were donor type (match related/match unrelated vs haploidentical; p = . for both), the disease status at transplant (cr vs beyond cr ; p = . , only for efs), acas (aca-vs aca+; p = . , only for os) and, especially, the complex karyotype (ck-vs ck+; p = . , only for os) (figure ). multivariate analysis showed that the independent prognostic factors for os and efs remained the complex karyotype (hr- . , % ci, . - . ; p = . ) and disease status at transplant (hr- . , % ci, . - . ; p = . ), respectively. the study demonstrates the acas and disease status at allo-hsct to be independent prognostic factors not only for adult, but for pediatric ph+ all patients too. up to % of newly diagnosed acute myeloid leukemia (aml) patients (pts) present initially with hyperleukocytosis consequently placing them at increased risk for morbidity and mortality during induction. , whereas early publications have indicated that hyperleukocytosis is an adverse prognostic factor associated with poor long term outcome, it is currently unknown whether hyperleukocytosis still retains prognostic value for aml patients undergoing allogeneic stem cell transplantation. furthermore, it is unknown whether hyperleukocytosis retains prognostic value when modern molecular markers such as flt and npm are accounted for. we hypothesized that hyperleukocytosis at initial diagnosis is still an independent adverse prognostic factor influencing long term outcome in aml pts undergoing allogeneic stem cell transplantation. we performed a retrospective analysis using the multicenter registry of the acute leukemia working party (alwp) of the european society for blood and marrow transplantation (ebmt). pts included in the analysis were over years of age, with de-novo non-m aml, a presenting white blood cell count of over k, with an hla matched related or unrelated donor, transplanted between and . clinical outcome indices of hyperleukocyotosis pts namely, non-relapse mortality (nrm), graft versus host disease (gvhd), relapse incidence (ri), leukemia free survival (lfs), overall survival (os) and gvhd-free/relapse-free survival (grfs) were compared to a cohort of pts without presenting leukocytosis. multivariate analyses were used to assess whether hyperleukocytosis was independently associated with ri, nrm, os, lfs, and grfs. age, gender, number of chemotherapy inductions, cytogenetics, donor type, fms-like tyrosine kinase- (flt ) status, nucleophosmin (npm ) status, and conditioning intensity were covariates for regression modeling. a cohort of pts with hyperleukocytosis ( patients with wbc over k and less than k, and patients with wbc over k) was compared to pts without hyperleukocytosis. pts with hyperleukocytosis were younger, had an increased rate of favorable risk cytogenetics, were more likely to be flt and npm mutated, and had an increased rate of myeloablative conditioning. on univariate analysis pts with hyperleukocytosis had an increased rate of ri ( % vs . %, p = . ), and decreased incidence of grfs ( . % vs . %, p = . ). in multivariate regression analysis, hyperleukocytosis was significantly associated with increased ri (hazard ratio [hr] of . , % confidence interval [ci], . - . ; p = . ), s poorer lfs (hr of . , % ci, . - . ; p = . ), decreased grfs (hr of . , % ci, . - . ; p = . ), and poorer os (figure ) (hr of . , % ci, . - . ; p = . ). image/graph hyperleukocytosis at initial presentation retains a significant prognostic role for aml patients undergoing allogeneic stem cell transplantation even in the current era of advanced molecular prognostication. outcome after hematopoietic stem cell transplantation for philadelphia-positive aml: relatively favorable outcome in patients allografted in first complete response; a survey from the acute leukemia working party of the european society for blood and marrow transplantation (ebmt) v lazarevic , m labopin , w deipei , i yakoub-agha , a huynh , p ljungman , n schaap , d blaise , jj cornelissen , n maillard , p pioltelli , t gedde-dahl , s lenhoff , m houhou , j esteve , m mohty and a nagler , aml with t( ; ) and bcr-abl rearrangement (ph-pos aml) is a very rare aml subtype, recognized as a new provisional entity in the recent who classification. the role of stem cell transplantation (sct) in the era of abl tyrosine-kinase inhibitors (tkis) is mostly unknown. we analyzed long-term outcome in patients ⩾ years after allogeneic or autologous sct performed between - in ebmt centers responding to a designated survey. patients with blast crisis cml and philadelphia-positive all were excluded. primary end-point was os. secondary end-points were nrm, acute gvhd, chronic gvhd, lfs, ri, and the effect of tki on outcome. patients (median age, years, range: - ; males and females) with ph-pos aml undergoing sct (allogeneic, ; autologous, ) were identified. median wbc count at diagnosis was x /l ( . - ) and % had splenomegaly (data missing on patients). translocation t( ; ) was the sole abnormality in patients ( . %). the majority of the patients received one or two courses of chemotherapy before transplant and % attained cr after one course. the majority ( %) received a tki (mostly imatinib, / ) before transplant, with a median period of exposition of days (iqr - ), while ( %) received tki after the transplant either as maintenance (n = ) or treatment for relapse (n = ). at time of transplant, patients were in complete response (cr - , including all autosct; cr - ) and the remaining patients were allografted in advanced phase. among patients with available information, achieved a mrd negative status at transplant (ratio bcr-abl/abl o ).regarding allosct, conditioning regimen was myeloablative (mac) in / ( %) patients o years, while in patients years received a reduced intensity regimen (ric) and mac. cell source was peripheral blood stem cells in and bone marrow in allogeneic transplants. the donor was a hla matched sibling (msd) in cases and unrelated (ud) in , amongst whom were / and were / hla matched, respectively. in the patients undergoing autologous sct the majority received busulfanbased conditioning (n = ) and peripheral stem cells (n = ). median follow-up was . . %, . %, . %, and . %, respectively. by the univariate analysis, age ( o vs ⩾ ) was associated with ri ( -yr: . vs %), lfs ( -yr: . vs . %, and grfs ( -yr: . vs . %), whereas mrd-negative status before allosct was associated with an improved grfs ( . vs . %). in the patients autografted, ri, nrm, lfs and os at -year was % ( . - . ), %, % ( . - . ), and . % ( - ), respectively. outcome of patients with ph-pos aml who received allosct in cr in recent years was relatively favorable, especially among younger patients, probably reflecting the beneficial effect of tki. disclosure of conflict of interest: none. outcome of allografting for aml-cr is equivalent across the bsbmt and ebmt and is associated with encouraging os and dfs across all age groups j byrne, j perry , k kirkland , r pearce and g jackson bsbmt and newcastle university and freeman hospital, newcastle relapsed aml has a very poor prognosis with a high mortality, even if a second cr is achieved. the only curative treatment is with an allogeneic hsct but allografts for aml in cr are considered to have a worse outcome compared to those performed in cr , especially in older patients for whom this therapy may not be considered. the bsbmt undertook a bench-marking study analysing the outcomes for all patients allografted for aml-cr from to . the uk outcomes from paediatric and adult patients were compared to non-uk patients transplanted for the same indication reported to the ebmt during the same period. allogeneic transplants for aml-cr represent an important part of any allograft program and numbers referred for allograft were stable between - in both programs. the median age of patients was . yrs and . years in the bsbmt and ebmt cohorts respectively, with % and % of patients aged o years and % and % aged years in the groups. the length of first remission was missing in many of the ebmt registrations so time from diagnosis to transplant was used as a surrogate for this and was similar in both cohorts ( m and m respectively). similarly the presence of comorbidities was poorly reported in both databases but was similar. the bsbmt cohort included fewer patients undergoing ric conditioning protocols ( % vs %), fewer sibling transplants ( % vs %) and more pbsc allografts ( % vs %). transplant related morbidity and mortality were similar across the two cohorts (bsbmt v ebmt) with rates of severe acute gvhd (grade iii and iv) % v %, limited chronic gvhd % v %, relapse at year % v % and death in cr at year % v %. chronic gvhd (both limited and extensive) appeared more common in the bsbmt cohort ( % v %) although reporting of cgvhd was more comprehensive and complete within the bsbmt registry and may be under-reported in the ebmt registry. outcomes were excellent in both cohorts with outstanding rates of leukaemia free survival in this high risk cohort at day (bsbmt v ebmt) % v %, at year % v %, at years % v % and at years % v %. although os and lfs was significantly shorter in patients aged years, at % and % the results in this high risk age group are acceptable and warrant its continued use. multivariate analysis of the combined cohorts showed that age at transplant ( o yrs/ - / yrs), time from diagnosis to transplant and the presence of agvhd were important factors affecting dfs. risk factors for relapse included the type of conditioning used, presence of agvhd and time from diagnosis to transplant, whereas those for trm included age, agvhd, source of stem cells and time to transplant. there was no significant difference in outcomes between the bsbmt and ebmt for this indication. outcomes for patients allografted for aml-cr are excellent and appear superior to those reported for patients not undergoing an allograft in both the bsbmt and ebmt cohorts. the os and dfs observed are comparable to those reported for allografts in aml-cr and, although this study has not considered outcomes for patients who did not achieve a nd cr, it nevertheless supports the practice of risk stratification of aml patients such that only high risk patients are offered an allograft in cr with the remainder being offered an allograft as salvage after relapse. disclosure of conflict of interest: none. to evaluate the efficacy and safety of intensive chemotherapy followed by allogeneic hematopoietic stem cell transplantation (allo-hsct) for atl. a total of evaluable atl patients treated at our center from to are retrospectively analyzed. the htlv-i proviral dna load in peripheral blood mononuclear cells using pcr assay was developed, in comparison with htlv- tax protein expression measured by western-blot, to confirm the diagnoses and monitor the disease control. patients were male and patients were female. median age was . (range - ) years. all obtained patient samples were subjected to flow cytometric examination and karyotype analysis. patients received chemotherapy as the induction therapy while quit at the time of diagnosis, with da-edoch regimen while with other regimens such as chop, vcap and amp. da-edoch regimen is a variation of dose-adjusted epoch regimen with the replacement of prednisone ( mg/m per day) by dexamethasone ( mg/m per day). before the conventional regimens bucy followed by prophylaxis donor lymphocyte infusion, both received a course of salvage chemotherapy including fludarabine and cytarabine for days registered on http://clinicaltrials.gov (nct ). the gvhd prophylaxis consisted of atg, csa and mtx. the patient characteristics, therapeutic effect and survival data were collected. all patients came from the coastal area in the south-east of china. subtype classification of these atll were acute, lymphoma and chronic type. the main manifestations were characterized with cutaneous and respiratory involvement, hepatosplenomegaly, lymphadenopathy and the laboratory abnormalities as leukocytosis with atl cells, hypercalcemia and elevated serum ldh. typical morphological characterisitic of "flower cells" were observed in . % cases and most of the atll cells are cd +cd -. chromosomal abnormalities were detected in cases. all patients who didn't receive da-edoch regimen died of disease progress, while among patients with da-edoch regimen, achieved cr, pr and died. with a median follow-up of . ( - . ) months, patients respond to da-edoch are still alive. patients in cr achieved successful engraftment with complete donor chimerism in one month post haplo-identical transplant. both were received prophylaxis donor lymphocyte infusion and the immunosuppressive agents were abruptly discontinued for induction of a graft-verus-atl (gvl) effect. they keep remain alive and disease free longer than years so far without severe graftversus-host (gvhd), and the htlv- proviral dna became undetectable after allo-hsct. conclusion: it shows great promise of da-edoch regimen followed by allo-hsct to the long-term cure of atl with apparent clearance of the virus. haplo-identical transplantation can be an alternative option for the atl patients without increasing non-relapse mortality. [p ] disclosure of conflict of interest: none. in the absence of an hla-matched related donor or a good matched unrelated donor in time, haploidentical stem cell transplantation (haplo-sct) is an option for patients requiring an allogeneic hematopoietic stem cell transplant. substantial progress has been made in the last two decades with a dramatic improvement in patient outcomes, with some groups reporting preliminary beneficial effects similar to the ones in hla matched unrelated donor and cord blood transplant. several strategies have been adopted through the years for the procedure. the two strategies used in haplo-sct are ex vivo t-cell depletion and t-cell replete transplantation. the latter can be done with a combination of immunosupressive drugs ( beijing approach) or with post-transplantation highdose cyclophosphamide (post-cy). due to of its lower cost, feasibility and practicality, post-cy has become the most often used platform for haplo-sct in the majority of allogeneic transplantation units worldwide. we analyzed our experience in haplo-sct, since the first one in march to the last one that has just been done in october . we collected all complications reported, also mortality related to treatment and to the disease. we analyzed the overall survival (os). transplants were treated, with different sct indications, the most common being acute myeloblastic leukemia (n = , %), the rest of indications are exposed in figure . all our patients, independent of the conditioning receive post-cy as t cell depletion measure and stem cells were collected from peripheral blood. age at the time of transplant was . years, % were males, % females, the rest of patient characteristics are listed in table . in our series the treatment related mortality (trm) was low with only patient ( %) that died before the day + . as complications, we reported % of hemorrhagic cystitis, % of sinusoidal obstruction syndrome, % reports systemic inflammatory response syndrome, % of citomegalovirus (cmv) reactivation. neutrophils graft is . days (r = - ) and the platelets graft is . days (r = - ). in our series we haven't reported any case of graft failure, one of the transplantes the patient had antihla antibodies, this was treated with a plasmapheresis previous the stem cell infusion, and was infused with a high number of cd + cells ( × /kg), no graft problems, and has had no complications since then with the graft. the os for the whole group is months, with a median not reach at months, with patient's dead at time of analysis. two patients had a relapse after the haplo-sct ( %), both of them received lymphocyte donor infusion, sadly, neither of them responded. the haplo-sct procedure is been adopted by many centers for high risk hematology malignancies, mainly because the fast availability of donors, and because of the preliminary results that have been reported place it as good as the unrelated donor or cord unit transplant. our center is getting experience in these types of transplants, and our results reflect similar outcomes as larger studies. with longer follow ups we will be able to keep the trend of good results both in procedure safety and disease efficacy. os, toxicity and trm are expected for these high risk malignancies. in overall, the haplo-sct seems a reasonable technique that is reflecting in our short series, the results being reported in studies worldwide at bigger scale. disclosure of conflict of interest: none. complex karyotype, the presence of flt- itd and losses of genetic material in chromosome , are all considered high risk markers in aml. patients bearing these abnormalities could undergo a myeloablative allogeneic transplantation in cr whenever this is possible, although this could significatly reduce the chances for cure in older patients due to increased transplant related mortality. ric allografts could be performed in older patients in order to overcome the deleterious effects of these individual abnormalities but its effect still remains controversial in the high risk group. between and , a total patients [ males, females, mean age . ( - )] received a ric allograft ( from fully matched unrelated donors, and from an identical sibling) for high risk aml in transplant centres. high risk disease was classified according to their response to treatment, the presence of complex karyotype, the presence of individual cytogenetic/molecular abnormalities or a combination of these. in particular, patients presented one single karyotype abnormality and presented three or more. ten patients presented alterations of chromosome and patients presented flt- itr. the conditioning regimes included fludarabine in all cases, together with melphalan and campath ( patients), busulphan and campath ( patients), busulphan and thiotepa ( patient), melphalan ( patients), busulphan with and without atg ( patients) total body irradiation ( cgy, patients). graft versus host disease prophylaxis was ciclosporin for patients receiving alemtuzumab or atg but for patients who had a t-replete allograft ciclosporin and low dose methotrexate was the preferred prophylaxis. all but four patients were transplanted in cr patients were followed-up for a mean . months (range - ). thirty-one ( %) patients remain alive. the causes of death ( cases) include relapse or progression of the original disease ( ), transplant-related causes ( ) and unknown in cases. the influence of the genetic abnormalities on survival was analysed, showing there were no significant differences between patients with normal karyotype, single chromosomal abnormalities and two or more abnormalities. likewise, the kaplan-meier survival analysis of patients bearing flt- itd was not significantly different to the rest of the cohort (p = . ; / died, with only one case being related to relapse). patients bearing chromosome abnormalities (with or without other chromosomal aberrations) had a comparable, not significantly (p = . ) different survival to the rest of the cohort: / patients bearing this abnormality died although, most interestingly, none of these deaths were related to relapse. our results indicate ric allografts can provide an adequate consolidating effect in hr aml with complex karyotype, alteration of chromosome or flt- itr, yielding clinical results that are comparable to those obtained in patients with aml allografted for other indications. this is particularly important as these alterations are more frequent in patients whose age prevents them from having myeloablative grafts. disclosure of conflict of interest: none. early detection of inapparent replicative human cytomegalovirus (hcmv) infection together with its preemptive antiviral treatment has led to a marked reduction of life-threatening hcmv disease after allogeneic hematopoietic stem cell transplantation (allosct). we first reported in a retrospectively performed study that hcmv reactivation is associated with a reduced risk for relapse in patients with aml after transplant. now, we evaluate the impact of early hcmv replication on the risk for leukemic relapse in patients with aml after t cell depleted transplantation in a prospectively performed observational study (registration trial drks ). between january and march we enrolled patients with aml in this trial who were consecutively transplanted at the university hospital of essen. patients received a myeloablative regimen (tbi based conditioning n = , chemotherapy based conditioning n = ) and patients a ric (n = chemotherapy based regimen). patients were transplanted in .cr (n = ), .cr (n = ) or more progressive disease stages (n = ) from hla-identical sibling donors (n = ) or hlaidentical unrelated donors (urd) (n = ) or mismatched unrelated donors (n = ). patients who received a second transplant were excluded from the study. the median age of patients was years (range - ) and that of the donors years (range - ). gvhd prophylaxis was performed with mtx and csa, or csa and mmf with (n = ) or without atg (n = ) ( - mg total dose). the incidence of acute gvhd grade - was statistically not different in both groups ( % versus %). hcmv-reactivation (hcmv-r) detected by pcr occurred between and days (median days) after allo sct. only patients surviving day after transplant were included in the study for estimation of relapse incidence (cir) or overall survival (os). hcmv status of recipients (r) or donors (d) were in % r-/d-, % r-/ d+; % d+/r-and % r+/d+. patients with a documented hcmv-r had a cir at -year after transplant of only % as compared to % in patients without a hcmv-r (p = . ). estimates for -year os were in favor for patients with hcmv-r ( % for patients with hcmv-r versus only % for patients without hcmv-r), but this did not achieve statistical significance. non-relapse mortality was greater in patients with hcmv reactivation ( % versus %, ns) a substantial and independent reduction of relapse risk associated with early hcmv replication was confirmed by multivariate analysis including competitive factors as unfavorable cytogenetics according to eln, advanced disease stages of aml, hla-identical donor versus mismatched donor, sibling versus unrelated donor, presence of acute gvhd grades ii-iv, chronic gvhd, and hcmv-r [(hazard ratio: . , % ci: . - . , p o . ) for occurrence of hcmv-r]. the final result of this first prospective performed study confirms an independent advantageous effect of early hcmv replication on the leukemic relapse risk in patients with aml after transplant. disclosure of conflict of interest: none. . primary engraftment of wbc and platelets was achieved in pts, one patient (pt) died at day + after hsct, and one had a secondary graft failure. a median time to wbc engraftment was days ( - ), to platelets - days ( - ). cumulative incidence (ci) of acute gvhd (agvhd) grade ⩾ ii was % ( % ci: - ): from haplo- %( - ), from mud % ( - ),p = . . ci of agvhd iii %( % ci: - ): haplo vs mud - % ( % ci: - ) vs %( % ci: - ), respectively,p = . . ci of chronic gvhd (cgvhd)- % ( % ci: - ). ci of agvhd was significantly lower in a group with regimen ( - ) of gvhd prophylaxis: % ( % ci: - ) vs %( % ci: - ), po . . regimen was also effective in prevention of cgvhd: ci at year after hsct was % vs %, p = . . Сi of trm was %( % ci: - ): haplo- %( - ), mud - % ( - ). early mortality (before + -day) was relatively low (n = ): pt died from bacterial sepsis; two pts died due to adv infection. thirteen pts died after days: pts relapsed and died due to complications of second hsct; bacterial sepsis in pt and viral infection (adv and cmv) in pts ( with extensive chronic gvhd). ci of relapse was % ( % ci: - ) at year: from haplo- % ( % ci: - ), from mud- % ( % ci: - ),p = . . event-free survival (efs) at years was % ( % ci: - ): haplo - %( % ci: - ), mud - % ( % ci: - ), p = . . os was % ( % ci: - ) at years: haplo- % ( % ci: - ), mud - % ( % ci: - ), p = . . relapse-gvhd-free survival at years was different among recipients of haplo and mud hsct: %( % ci: - ) vs % ( % ci: , p = . . we confirm that the depletion of tcrαβ+/cd + depletion from the graft ensures high engraftment rate and acceptable transplant-related mortality in pediatric aml pts. there is a trend towards better efs for haploidentical transplantation. gvhd prophylaxis including ratg/rituximab/bortezomib improves gvhd control s in recipients of tcrαβ+/cd +depleted grafts in comparison to hatg/tacro/mtx apparently without loss of anti-leukemic activity. disclosure of conflict of interest: none. results of t-cell depleted haploidentical stem cell transplantation in adults with acute leukemia improve with time: a study from the acute leukemia working party of the european society of blood and marrow transplantation (ebmt) s simona , , m labopin , a ruggeri , , , a velardi , f ciceri , j maertens , l kanz , f aversa , d bron , d bunjes , m mohty , and a nagler , t cell-depleted (tcd) transplants from haploidentical donors are increasingly used in the absence of a hla full-matched donor for patients (pts) with high risk acute leukemia (al). progress has been made in optimization of conditioning regimens and post-transplant cellular therapy to potentiate the graft-versus-leukemia effect with no graft-versus-host disease (gvhd). however, relapse incidence (ri) and non relapse mortality (nrm) remain the main obstacles for pts outcomes. we report adults with de novo al, given a tcd haplo from to . to analyze the effect of transplant period on tcd haplo outcomes, pts were analyzed in two separate groups: - (n = ) and - (n = ). tcd haplo were performed in ebmt centers. median follow-up was months with no difference according to transplant periods. median age was different between groups, being and years respectively (p = . ). the majority of pts had acute myeloid leukemia (aml) ( % vs %, p = . ) and disease status at haplo was first complete remission (cr ) in % and % of pts respectively (p = . ). pts transplanted before had more frequently a karnofsky performance status o % ( % vs %, p = . ). conditioning was myeloablative in % and % of tcd haplo before and after (p = . ), mainly based on fludarabine(flu)-tbi, flu-melphalan-thiotepa or cyclophosphamide-tbi. as for ric it was flu-melphalan-thiotepa, flu-tbi or cyclophosphamide-tbi. the cumulative incidence (ci) of neutrophil engraftment, grade ii-iv acute gvhd and chronic gvhd were not different according to transplant period, being % and %, p = . ; % and %, p = . ; % and %, p = . , respectively. in the whole population -year ri and nrm were % and %, with no difference before and after ( % vs %, p = . ; % vs %, p = . , respectively). ri was % before versus % after . the main cause of nrm was infection, with no difference over time ( % vs %, p = . ). in multivariate analysis, disease status was the only factor associated with ri (hr . , % ci . - . , p = . ). tcd haplo after (hr . , % ci . - . , p = . ), younger age (hr . , % ci . - . , p = . ) and ric (hr . , % ci . - . , p = . ) were independently associated with lower nrm. -year os was % with a marked improvement for tcd haplo performed after ( % vs %, p = . ), while lfs and grfs were % and %, respectively. according to disease status, -year lfs, os and grfs were higher for pts transplanted in cr ( % vs %, p o . ; % vs %, p = . ; % vs %, respectively p = . ). in multivariate analysis, tcd to further establish the role of haplosct in high-risk aml, we performed a retrospective matched-pair comparison of hlamatched sct vs haplosct/ptcy in two german centers. highrisk aml was defined by any of the following criteria: refractory or relapsed aml, secondary aml, or genetic aberrations classified as intermediate-high or adverse accordingly to the eln classification. all consecutive adults, who fulfilled ⩾ of these criteria before either hla-matched or haplosct/ptcy were included (n = ). recipients of haplosct were pair-matched with patients receiving a matched donor sct. matching variables were ( ) stage at sct, ( ) genetic subgroups accordingly to eln, and ( ) age (± y). patients (pts) undergoing haplosct/ptcy could be successfully pairmatched (p = . for stage and genetic subgroup, . for age) with recipients of matched sct ( family, unrelated sct). within the entire cohort, median patient age was y ( - ). at start of conditioning, % of patients were in cr, % had refractory, % had relapsed, and % had untreated disease. genetics were favorable ( %), intermediate i ( %), intermediate ii ( %) and unfavorable ( %). hla-matched sct was uniformly performed following flamsa-ric. recipients of haplosct/ptcy ( %) received cytoreductive chemotherapy with flamsa (n = ) or clofarabine (n = ) before ric was started. median follow-up among survivors was months. overall cr rate at d+ was %, patients suffered from refractory disease or early death, (n = each). overall-survival (os) for the entire cohort was %/ . % at y/ y from sct. the corresponding y/ y leukemia-free survival (lfs) was . %/ . %. median time to engraftment was . and . days after matched and haplosct, respectively (p = . ). with respect to outcome, no difference was observed between the two groups: os at y/ y was . %/ . % after matched sct, and . %/ . % after haplosct/ptcy (p = , , figure ). lfs at y/ y was . %/ . % within the hla-matched group and . %/ . % within the haploidentical group (p = , ). recipients of haplosct showed a higher incidence of agvhd ⩾ ii°( % vs %, p = . ), as well as a trend towards increased y-nrm ( % vs %, p = . ), whereas y-relapse rates were comparable ( % after haplosct/ptcy vs % after matched sct, p = . ). relapse was the most frequent cause of death in both cohorts, main causes of nrm were gvhd and infections (no difference between the two groups). allogeneic sct following sequential conditioning can achieve excellent results in high-risk aml. in our study, results after haploidentical transplantation were comparable to those obtained after hla-matched sct. hence, haplosct/ptcy following sequential conditioning can be considered as a reasonable option for patients with high-riaml. [p ] disclosure of conflict of interest: none. a significant proportion of patients with acute myeloid leukemia ( aml) will either be refractory to initial chemotherapy or will suffer refractory relapse. the role of allogeneic transplantation (hct) in active disease is contentious. there is a growing body of literature that sequential chemotherapy, pioneered by the german flamsa regimen, followed by ric hct is a safe and efficacious modality in these patients, and there have been numerous modifications of this regimen, especially as amsacrine is not widely available. fludarabine, cytarabine and etoposide (vp ) (flav) have been reported as an an effective salvage regimen. here we report on single center outcomes of a variation of the flamsa regimen, substituting amsacrine for etoposide with mainly myeloablative conditioning. patients were consented for a clinical protocol if they had aml that was refractory to cycles of chemotherapy, or cycle and considered to be at risk of complications of a second cycle, and if they had a matched related donor. patients with myelodysplasia received flav if they had high or very high risk cytogenetics. cytoreductive chemotherapy consisted of fludarabine mg/m / day × days, cytarabine g/m /day × days, etoposide mg/m /day × days, commenced simultaneously. after days of rest, conditioning chemotherapy consisted of fludarabine mg/m × days and and iv busulfan . mg/ m q hours; the number of busulfan doses varied between - , depending on patient comorbidity. ten patients ( %) received myeloablative doses of busulfan. patient received doses of atg at . mg/m /day on day - and - . patients received gcsf mobilized peripheral blood hematopoietic cells. post-transplant gvhd prophylaxis was csa and mmf. csa was tapered from day+ and stopped at day + in the absence of gvhd. mmf was discontinued between day + and day + . donor lymphocyte infusions were collected for planned prophylactic dli. thirteen patients received a flav-sct between march and october . the median age was ( - ); male:female ratio was( : ). patients ( %) had aml and ( %) pts had mds. all patients had detectable disease prior to flav. the median time for plt engraftment was days ( - ). the median time for anc engraftment was days ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) . cytogenetic cr rate on a day - bone marrow was %, and morphological cr was %. patients ( %) developed veno-occlusive disease. acute gvhd grade ii-iv occured in pts ( %). ( %) patients developed chronic gvhd. death was due to disease relapse in ( %) and nrm in ( . %) patients, resulting from h n pneumonia. patients ( %) received dli for post transplant relapse, and one of these is in molecular remission. at a median follow up of . months post transplant ( . - m), year dfs was %. the year and year os was % (± %) (figure ). our experience, consistent with published data, demonstrates that for patients with active aml refractory to chemotherapy, transplantation is an effective modality of disease control and may be the only curative therapy in a significant proportion. etoposide appears to be a suitable alternative to amsacrine. our patients tolerated busulfan at myeloablative doses, and this may be required for adequate disease control. our report is limited by small numbers and relatively short follow-up, but is encouraging enough for us to continue offering flav hct for these high-risk patients. [p ] disclosure of conflict of interest: none. sequential high-dose chemotherapy reinduction followed by haploidentical transplantation in acute leukemias l brunello , , cm dellacasa , l giaccone , , e audisio , d ferrero , , s d'ardia , b allione , s aydin outcomes after t-cell replete haploidentical stem cell transplantation (haplo hsct) have been encouraging and haplo hsct has become an alternative option for patients without a hla-identical related or unrelated donor. , as previously published, the sequential use of intensive chemotherapy and allogeneic transplantation represents a possible approach to the treatment of high-risk acute myeloid leukemia (aml). , between and , acute leukemia (al) patients received sequential therapy (s.t.) consisting of high-dose chemotherapy reinduction and haplo hsct during the chemotherapy-induced neutropenia. median age at transplant was years (range: - ); median number of previous therapy lines was (range: - ) and / ( %) patients had received a previous allogeneic hsct. twelve and out of patients had de-novo aml and secondary aml, respectively; furthermore two patients presented blastic crises of chronic myeloid leukemia. all patients had active disease at the time of s.t. and median marrow blast count before reinduction was % (range: - %). hematopoietic cell transplantation comorbidity index was ≥ in / patients ( %). all patients received high-dose cytarabine (≳ g/sqm) containing regimens as reinduction therapy. the conditioning regimen was started after a median of days from the end of reinduction (range: - ). sixty-eight percent of patients ( / ) were conditioned with a myeloablative regimen (thiotepa tot. mg/kg, busulfan tot. . mg/kg, fludarabine tot. mg/ sqm), while / ( %) patients received a non-myeloablative conditioning. bone marrow was used as stem cell source in / ( %) patients. graft-versus-host disease (gvhd) prophylaxis consisted of post-transplant cyclophosphamide with calcineurine inhibitors and mycophenolic acid. all patients engrafted but one, who was rescued with a second haplo-hsct with peripheral blood stem cells from the same donor. median day of neutrophil recovery was day + (range: - ). median follow-up of survivors was . years (range: . - . ); -year overall and event-free survivals were % and %, while -year relapse incidence and non relapse mortality were % and %, respectively. overall cumulative incidences of acute and chronic gvhd were % and % at day + and + . among patients who developed gvhd, grade iii-iv acute gvhd and moderate-severe chronic gvhd were observed. at . years post haplo-hsct, % of patients are alive and disease free. in our cohort of heavily pre-treated and high-risk patients, s.t. with a myeloablative conditioning was safely used to reduce leukemic burden pre-transplant and enhance graft-versus-leukemia effects. only the prompt availability of a haploidentical donor allowed to implement this treatment modality. though small the patient cohort, our findings suggest that transplant-related toxicity was acceptable and early relapse was the major treatment-failure. however, long-term survival and disease-free rates of % in these very poor prognosis patients are highly encouraging. in elderly patients with acute lymphoblastic leukemia (all) and kinase activating lesions allogeneic stem cell transplantation (allo-sct) is considered to be the only curative option. however, high risk of relapse and non-relapse mortality (nrm) often withholds elderly patients with existing comorbidities from definitive therapy. even though, age alone as the most important eligibility criterion for allo-sct has become less important. a -year old patient with a medical history of adipositas, hypothyreosis, arterial hypertension, hypercholesterolemia and coronary artery disease with stent implantation was diagnosed with common-b-cell-all based on immunophenotype. cytogenetic analysis showed two coexisting clones with an abnormal karyotype of ,xx; t( ; )(q ;q ), + [ ] and ,xx; t( ; )(q ;q ),der( )( ; )(p ;q ) [ ] and no evidence of bcr-abl positive disease using fluorescence in situ hybridization (fish) technique. rt-pcr and sequencing of the fusion transcript was performed to validate the rcsd -abl t( ; )(q ;q ) fusion between exon of rcsd and exon of abl . western analysis of phosphorylated abl and its downstream target crkl was performed to investigate the in vivo activity of dasatinib. clinical monitoring of minimal residual disease (mrd) levels has been performed via rt-pcr of the rcsd -abl fusion transcript followed by nested pcr of the amplicon to detect early relapse or mrd. as positive control the plasmid pcr . -topo_rcsd -abl ( bp) was synthesized encoding rcsd -abl amplicon with the fusion site. our patient was enrolled on gmall elderly ( / ) and treated accordingly. thereby, no sustained remission could be achieved. flag-ida re-induction and study treatment with an oral pi k/mtor inhibitor remained futile. cytogenetics and verification of the rcsd -abl fusion gene prompted salvage treatment with the tyrosine kinase inhibitor (tki) dasatinib as single agent. the in vivo activity of dasatinib was highlighted by a decrease of rcsd -abl amplicon and inhibition of phosphorylated abl and its downstream target crkl was shown. clofarabine and cyclophosphamide complemented treatment and mrd negative remission was achieved due to administration of the bi-specific t-cell engager blinatumomab. consolidation with allo-sct was performed. ongoing remission has been achieved for more than months now. we demonstrate that monotherapy with tki like dasatinib is effective in refractory rcsd -abl positive all. to the best of our knowledge, this is the first elderly patient with rcsd -abl positive all with a sustained and ongoing complete remission. thereby, we suggest allo-sct after successful tki even for elderly patients with existing comorbidities and uncommon cytogenetics. relapse is the most important cause of failure of allogeneic hematopoietic stem cell transplantation (hsct) for flt -itdpositive acute myeloid leukemia (aml). in those cases, treatment with flt tyrosine kinase inhibitors (tki) constitutes a promising clinical approach to induce remission without conventional chemotherapy. forty-eight-year-old woman was diagnosed of aml secondary to myelodisplastic syndrome with npm mutation and internal tandem duplications of the flt gene (flt -itd). after achieving complete remission (cr), she received a sibling allogenic-hsct. four months later, aml relapsed at the medullary level, without central nervous system (cns) involvement, treated with conventional chemotherapy and donor lymphocytes infusions (dli). she achieved second cr and developed chronic graft-versus-host disease (cgvhd). nine months later, she suffered the first extramedullary relapse, at the mammary, cutaneous and probably pericardial levels. there was not medullary involvement. disappearance of the lesions at all levels was achieved with conventional chemotherapy and radiotherapy, and full donor chimerism. eight months later, she referred atypical precordial pain irradiated to the back. cardiac mri was performed in which several masses were visualized in a pericardial sac up to cm in diameter (dec- ). bm was maintained in cr. in study of pericardial fluid, infiltration by leukemic flt positive cells was observed. the patient was not considered candidate for further systemic chemotherapy nor radiotherapy treatment. then, treatment with the flt sorafenib inhibitor was started, by compassionate use, at dosage of mg/ h, which maintains after one year. after first month with sorafenib, pericardial lesions decreased considerably, ranging from cm in diameter to . cm (jan- ). in the subsequent ct controls, progressive decrease of the lesions has been observed and no new lesions have appeared in other locations. in the last ct (oct- ) pericardial thickening is almost non-existent, without new lesions. after one year of treatment, she maintains cr at medullary and extramedullary levels images. in our patient, treatment with sorafenib has achieved sustained control of extramedullary disease, which had escaped the mechanisms of action of conventional chemotherapy, allotransplant and dli. further studies are needed to corroborate the efficacy of flt inhibitors in the control of aml extramedullary disease and in the treatment of relapses after allo-hsct. all pts with tbi-based regimen received rabbit atg. tcrαβ+/cd + depletion of hsct with clinimacs technology was implemented in all cases according to manufacturer's recommendations. the median dose of cd + cells in transplant was × /kg (range: . - . ), tcrα/β- × /kg (range: . - ). primary engraftment was achieved in of pts. ( pts died before engraftment, one received second hsct), the median time to neutrophil and platelet recovery was and days, respectively. early ( day) mortality was %( % ci: - ), -year ptrm- % ( % ci: - ). the three early deaths were due to bacterial sepsis (n = ) and viral infections(n = ), seven late: viral infection in pts (adv = , adv+cmv = , cmv = ), bacterial sepsis in pts and rhinocerebral mucormycosis in pt, all late deaths were associated with cgvhd and prolonged corticosteroid therapy. ci of acute gvhd grades ii-iv and iii-iv was . % ( % ci: . - ), and % ( % ci: . - ), respectively. ci of cgvhd was . % ( % ci: . - ). regimen was more effective in prevention of agvhd ii-iv: ci at year after hsct was % vs , % in regimen , p = . and in cgvhd % vs . %, p = . . ci of relapse at years was % ( %ci: . - . ). two-year pefs(event = death or relapse) was . % ( % ci: - ), -year pos- % ( % ci: - ). in patients, who received tbi-based conditioning pefs was % ( % ci: - ), as compared with treosulfan-based % ( % ci: - ), p = . . median time of follow-up for survivors was . years (range: . - . ). we confirm that the depletion of tcr-alpha/ beta and cd lymphocytes from the graft ensures high engraftment rate and acceptable transplant-related mortality in pediatric all patients. viral infections and leukemia relapse await further improvement of control. all major outcomes were equivalent between transplantation from unrelated and haplo donor. disclosure of conflict of interest: none. the prognosis of patients (pts) with relapsed/refractory acute lymphoblastic leukemia (all), especially after allogeneic hematopoietic stem cell transplantation (allo-hsct), is very poor. therapeutic options for these pts are limited. blinatumomab is a bispecific t-cell engager (bite) antibody construct with dual specificity for cd and cd . bite therapy may help to overcome the resistance to chemotherapy (ct) with minimal toxicity, and may be a bridge to allo-hsct. we analyzed data of pts from hematologic centers in russian federation with relapsed cd positive all, who received bite. the median age was (range: - ) years, ( %) pts o yrs, ( %) pts ≥ yrs. the diagnosis was all b-i (egil) subtype in pts, b-ii-in , b-iii-in pts, and patient had mixed phenotype leukemia (m (fab) and b-all). three ( %) pts had philadelphia positive (ph+) all. in pts it was the first relapse of all, in -second, in -third. thirty pts had isolated bone marrow relapse, pts-combined relapse (bone marrow and extramedullary sites). the bone marrow blast infiltration was o % in pts, % in pts. disease relapse was revealed after ct in pts ( ( %) pts received allo-hsct after the therapy of relapse), after allo-hsct ( -from related, -from unrelated, -from haploidentical donors)-in pts ( pts received second allo-hsct after the therapy). in pts with posttransplant relapse donor lymphocyte infusion (dli) was used in combination with bite. every patient received from to cycles (median ) of bite. complete remission (cr) was achieved in ( %) pts (in ( %) pts it was minimal residual negative remission): in ( %) pts with all relapse after ct, in ( %) pts-after allo-hsct. pts with less than % blasts in bone-marrow at baseline experienced substantially higher response rates compared with patients with % blasts or higher ( % ( / ) vs % ( / )). response rates were similar although the number of relapse- % ( / ) in first relapse, % ( / ) in second relapse and % ( / ) in third relapse. pts with posttransplant all relapse who received bite in combination with dli had higher response rate than pts, who received bite as monotherapy: . % ( / ) vs % ( / ), respectively. one-year os was % ( % ci - %). one-year dfs was % ( % ci - %). grade ≥ hematological toxicity (neutropenia, thrombocytopenia) was observed in ( %) pts, grade ≥ liver toxicityin ( %) pts. five patients ( %) developed toxic neuropathy during the therapy. cytokine release syndrome occurred in ( %) pts. one patient after allo-hsct (but not after dli) developed grade i agvhd. there were no fatal treatment related toxicity. tree ( %) pts who responded to bite had relapse. eighteen ( %) pts died: pts-of disease relapse/ progression. the treatment of relapsed/refractory all with bite is effective and has acceptable toxicity. we demonstrated high efficacy in therapy of posttransplant all relapses, especially when bite was combined with dli, perhaps, due to additional immunological effect of the transplant. disclosure of conflict of interest: none. the mechanism of sorafenib anti-leukaemic effect seen in aml patients relapsing post allohsct involves the augmentation of alloreactivity which includes infiltration of the affected marrow by cd + cells having pd- receptor which presence characterize lymphocytes with antitumour potential multikinase inhibitor (mki) sorafenib is clinically active in acute myeloid leukaemia (aml) patients, especially in those with flt itd who receive allohsct as a part of their primary treatment. to throw some light on the mechanism of this antileukemic post-transplant sorafenib effect we studied the fate of two patients (flt itd-positive, npm -positive) who relapsed ( -year-old male) and ( -year-old female) days post-transplant and their salvage treatment included sorafenib. the multikinase inhibitor ( mg twice daily) was given either together with flag or da + . the response was prompt. the patients became, after completion of the chemotherapy, leukaemia free. both patients continued the sorafenib ( mg twice daily) treatment together with the aml standard maintenance chemotherapy (female case) or without any chemotherapy (male patient, substantial comorbidity and liver toxicity). ( ) . the patients responded well to the therapy and were free of leukaemia for and + months after initiation of the mki treatment (flt itd negative, % chimerism documented in the blood and in the marrow). ( ) . in both patients, and months on sorafenib, skin lesions appeared either in the context of cgvhd, which progressed to a life-threatening level in a male patient or as a photodermatitis-like cheek eruption. histopathology revealed the presence of severe cd + cells infiltration in affected tissues in both patients. ( ) . cd positive lymphocytes colonized the marrow of both patients. these marrows infiltrating cells co-expressed cd (pd- receptor) in proportions which were higher than those seen in the blood ( . % ± . % vs. . % ± . %, p = . ). a similar observation was made for cd +cd + cells ( . % ± . % vs. . ± . %, po . ). . transcriptome analysis of the marrow cells, which addressed the genes involved in lymphocyte activation, revealed the presence of proinflammatory profile which included a higher expression of tlr and il- . ( ) sorafenib given with or without moderate chemotherapy was effective in two patients in maintaining the anti-leukaemic effect of salvage chemotherapy. ( ) this was associated with the presence of alloreactivity (affected tissues infiltration with cd + cells) clinically seen as a severe fatal cgvhd aggravated by sorafenib treatment associated unwanted effects in one cases and with rather mild skin lesions appearing later during the treatment. the outcome of elder patients with acute myeloid leukemia or high risk myelodysplastic syndrome treated with allogeneic hematopoietic stem cell transplantation ch tsai , , division of hematology, department of internal medicine, national taiwan university hospital; tai-cheng stem cell therapy center, national taiwan university and genome and systems biology degree program, national taiwan university allogeneic hematopoietic stem cell transplantation (hsct) is a curative-intent treatment for patients with high-risk hematologic diseases, including acute myeloid leukemia (aml) and myelodysplastic syndrome (mds). both the incidences of aml and mds increase with age and patients elder than years of age were traditionally excluded from hsct because of high possibilities of therapy related morbidity/mortality. recently, with the introduction of reduced intensity conditioning regimens and the improvement of hsct care, more and more elder patients could undergo hsct for consolidation or salvage purposes. however, literature regarding the treatment outcome of elder patients receiving hsct is scarce. patients diagnosed as aml or high risk mds aged equal or more than years were recruited consecutively at national taiwan university hospital. the high risk mds was defined to include myelodysplastic syndrome with excess of blasts- and according to the world health organization (who) criteria. the cytogenetic risk stratification was based on original medical research council classification. from to , a total of patients were enrolled consecutively. the median age was . (range: - ) years and the gender distribution was even. among them, ( . %) patients had high risk mds, ( %) had de novo aml, ( . %) had secondary aml, and three ( . %) had therapy related aml. at diagnosis, four ( . %) patients had extramedullary disease. nine ( . %) had unfavorable-risk cytogenetics, ( . %) had either unfavorable-risk cytogenetics or intermediate-risk cytogenetics but with flt -itd mutations. regarding the pre-hsct disease status, nine patients had the first complete remission (cr), had the second cr, and patients were treatment naive or had refractory disease. the graft-versushost-disease(gvhd)-free/relapse-free survival (grfs) in which events include grade - acute gvhd, chronic gvhd with moderate severity according to cibmtr criteria, relapse, or death of any cause. with median follow-up of months (range: . - . ), the median overall survival (os) for all patients was . months, the relapse-free survival (rfs) was . months, and the grfs was . months. in univariate analysis for os and rfs, high-risk mds was a favorable prognostic factor but secondary or therapy related disease (p = . for os and . for rfs, respectively), unfavorablerisk cytogenetics or intermediate-risk cytogenetics but with s flt -itd mutations (p = . and . , respectively), pre-hsct refractory disease (p = . and . , respectively), and grade - acute gvhd (p = . and . , respectively) were unfavorable prognostic factors. however, for grfs, only unfavorable-risk cytogenetics or intermediate-risk cytogenetics but with flt -itd (p = . ) and pre-hsct refractory disease (p = . ) were unfavorable prognostic factors. in multivariate cox proportional hazards regression analysis for os and rfs, grade - acute gvhd was a significant unfavorable risk factor; for gfrs, pre-hsct refractory disease status was a significant unfavorable risk factor. our results showed that the choice of hsct should not solely based on the age factor and pre-hsct disease status. incorporating cytogenetics and genetic mutation status could risk-stratify elder patients with hsct. further prospective trials are warranted to validate these findings. disclosure of conflict of interest: none. children affected with acute lymphoblastic t cell leukaemia (t-all) and relapse after allogeneic stem cell transplantation (sct) have limited treatment options and a poor prognosis. immune checkpoint inhibitors targeting the programmed death (pd- ) receptor pathway may enhance the graftversus-leukaemia (gvl) effect by blockade of inhibitory signals to t cells mediated by its ligand pd-l . we report a -year old girl with refractory t-all after allogeneic sct, who was treated off-label with the pd- inhibitor pembrolizumab. the girl was diagnosed with t-all ( . g/l wbc, % bone marrow infiltration, cns negative, t ( ; )) and underwent hlahaploidentical bone marrow transplantation from her mother with post-transplant cyclophosphamide since she failed to achieve molecular remission despite an intensified chemotherapeutic regimen. on day post sct, she had a % donor chimerism and decreasing minimal residual disease (mrd) marker (minimal × − ). days post sct she had a molecular relapse with an mrd of × − and a subsequent morphological relapse as well as mixed donor chimerism. further treatment regimens included chemotherapy, intrathecal therapy and four donor lymphocyte infusions (dlis). initially, she displayed a good morphological response to dlis but the leukaemic burden eventually remained stable with an mrd of × − . considering . % pd- expression on cd + t cells in the patient's bone marrow and the encouraging data in other hematologic malignancies an off-label therapy with the pd- inhibitor pembrolizumab - was initiated. the patient and her parents gave informed consent and she received a single dose of pembrolizumab at . mg/kg days after sct. one week after administration of pembrolizumab, the patient developed acute gvhd grade iv of the skin, mucosa, liver, lung, central nervous system and eyes. she had a severe generalized inflammatory reaction with high inflammatory markers, increased hepatic transaminases and lymphocytic infiltration of the liver, cerebrospinal fluid and bronchoalveolar lavage fluid. magnetic resonance imaging (mri) of the brain revealed periventricular white matter lesions and hyperintensities of basal ganglia and bilateral temporal lobe consistent with autoimmune encephalitis. treatment with high-dose corticosteroids, cyclosporine and the anti-interleukin receptor antibody tocilizumab slightly improved her clinical condition. her mrd value significantly decreased to × − two weeks after administration and she achieved a % donor chimerism in bone marrow. despite this promising response her medical condition deteriorated and the severe inflammatory reaction caused fatal multi-organ failure. this is to our knowledge the first report on a remarkable and fast response to pd- inhibition in a patient with pediatric t-all refractory to multiple lines of therapy including allogeneic sct. this case illustrates the potential risk of checkpoint inhibitors to trigger severe gvhd that is not responsive to steroids. induction of inflammatory gvl responses without causing severe gvhd by therapeutic checkpoint inhibition needs to be addressed in future clinical trials. in recent years, there is a remarkable trend in the use of haploidentical-related hematopoietic cell transplantations (haplo-hct) in patients who do not have a hla matched related or unrelated donor. here, we report our single-center experience, in patients who underwent haplo-hct for acute leukemia. between and seventeen consecutive adult patients, seven males and ten females, median age years (range: - years) with high-risk acute leukemia underwent unmanipulated, bm or pbsc transplantation from an haploidentical family donor. eleven patients transplanted for acute myeloid leukemia ( in cr , in cr , in minimal active disease after cr , second trasplant in cr , transformed mds in cr , aml secondary to myelofibrosis in cr ), for acute lymphoblastic leukemia ( in cr , in active disease) and mastcell leukemia (secondary to aml) in active disease. sixteen patients received myeloablative conditioning, and reduced intensity, respectively. in five patients stem cells source was bm, in were g-csf mobilized pbsc. the median infused cd + cell dose was . × (range: . × - . × ). conditioning regimens were: bu-flu-mac (n = ), tbf-mac (n = ), tbf-ric (n = ) the regimens for gvhd prophylaxis were: ptcy as sole gvhd prophylaxis (n = ), mtx-csa-atg (n = ), methylpred-atg-tacrolimus (n = ), atg-csa-mtx-mmf (n = ). sustained trilineage engraftment occurred in patients ( %), two patients died of transplantation-related complications before day after transplantation without myeloid recovery. for patients receiving bm or pbsc grafts, the median time to neutrophils recovery was days (range: - ), and , platelets recovery was days (range: - ). / patients ( . %) and / ( . %) had ii-iv and iii-iv grade of acute gvhd, respectively.the incidence of grade ii-iv cgvhd was %. after a median follow-up of months, / patients ( . %), out patients transplanted in cr , are alive and disease free at , , , months (inclusing the patient transplanted for aml after imf). the -year probability of overall and progression-free survival was % ( % ci, . - . %) and . % ( % ci, . - . %), respectively. causes of death were: sepsis (n = ), fatal agvhd (n = ), pneumonia (n = ), toxicity (n = ), progression (n = ) and relapse (n = ). in our experience unmanipulated bm or pbsc transplantation from haploidentical family donor is feasible approach with high engraftment rates and acceptable trm ( %) and rate of grade iii-iv agvhd, associated with durable remission in a proportion of patients with high-risk acute leukemia, specially in patients with aml transplanted in first remission. it is generally recognized that allogeneic hematopoietic stem cell transplantation (allo-hsct) should not be administrated to patients with severe aplastic anemia (saa) or very severe aplastic anemia (vsaa), when they got active infection. however, without neutrophil, severe infection is usually difficult to control and even fatal. under these circumstances, rapid recovery of neutrophil by allo-hsct might be an alternative to control infection. from january to december , there were young patients received allo-hsct for saa or vsaa at shanghai children's medical center in china. among them, patients ( males and females) with a median age of . years (range: . - . years) received allo-hsct with refractory active infections. refractory active infection was defined as persistent neutropenic fever with nonresponse to standard doses of broad-spectrum antibacterial agents and antifungal agents for more than three weeks, with or without definite focus of infection. prior to allo-hsct, four patients had persistent fever of unknown origin, patients with singlesite infection, and patients with multiple-site infections. sites of infection included lung, sinus, cellular tissue, peritoneum, liver, spleen and skin. the conditioning regimen consisted of fludarabine, cyclophosphamide and rabbit-antithymocyte globulin with or without total body irradiation (tbi) ( - gy). twelve patients were transplanted from mismatched unrelated donors, from matched sibling donors, and from haploidentical donors. sixteen patients received g-csf mobilized peripheral blood stem cells, three patients g-csf mobilized peripheral blood stem cells plus g-csf primed bone marrow stem cells, two patients bone marrow stem cells, and patient umbilical cord blood stem cells. a median of . × /kg mononuclear cells with . × /kg cd + cells were transfused, except the patient who underwent ucbt with a total of . × /kg mononuclear cells and . × /kg cd + cells transfused. eighteen patients achieved recovery of neutrophil and finally control of infections, including one patient who suffered primary graft failure and had autologous marrow recovery. three patients died of infection and one patients died of acute renal failure before recovery of neutrophil. one patient died of pneumonia months after allo-hsct. one patient become thrombocytopenia after allo-hsct. the other patients are all disease-free. there were five patients developing grade i-ii acute gvhd, and patient grade iii-iv acute gvhd. all were cured at last. three patients had localized chronic gvhd and one patient had extensive chronic gvhd. with a median of years follow-up, the overall survival rate and disease-free survival rate are . % ± . % and . % ± %, respectively. allo-hsct could be a feasible way to control infection for children with saa or vsaa in the present of refractory active infections. disclosure of conflict of interest: none. paroxysmal nocturnal hemoglobinuria (pnh) may present hemolysis isolated (classical pnh) or associated with aplastic anemia (aa; aa/pnh syndrome). while classical pnh patients require anti-complement treatment (eculizumab), the treatment of aa/pnh patients should target their underlying aa by immunosuppression (ist), or even bone marrow transplantation (bmt). however, in a few patients clinically meaningful aa and hemolysis may be concomitant, eventually justifying both ist and eculizumab. to date there is no standard treatment for s this rare condition. amongst a large cohort of pnh patients (between and ) at our reference centers, st. louis hospital (paris) and federico ii university (naples), we retrospectively assessed characteristics and outcomes of patients diagnosed with aa/pnh who received intensive ist during or immediately before ( - months) eculizumab treatment. nine patients were identified. eight patients fulfilled the criteria of severe aa, and one had an immunemediated isolated agranulocytosis. since no patient had a hla-matched related donor for bmt, all patients received intensive ist according to institutional guidelines. six out of patients were already on eculizumab treatment at the moment of starting intensive ist (concomitant treatment) whereas patients received ist in the - months (median time of months) before the introduction of anti-complement therapy (sequential treatment). for all patients already on treatment, eculizumab was not discontinued to minimize the risk of rebound intravascular hemolysis and thrombotic complications. eculizumab was administered at the standard dose of mg fortnightly in all but one patient, who needed an increased dose ( mg) because of pharmacokinetic breakthrough. six patients ( aa and agranulocytosis), including the three undergoing a sequential treatment, received standard ist with horse-antithymocyte globulin (h-atg, mg/kg for four consecutive days) combined with cyclosporine a (csa). the remaining three aa patients received alemtuzumab ( - - - mg subcutaneously in four consecutive days) and csa within the prospective trial nct ; one of these patients a few months later also received a second ist course with rabbit-atg ( . mg/kg for five consecutive days) and csa. all the patients completed the scheduled treatment without any side effect, including infusion-related reactions. lymphocyte depletion (o /μl) was observed in all patients irrespective of sustained therapeutic complement blockade. all the patients were available for response assessment at months. among the six patients receiving a concomitant treatment we observed one partial response (pr) and two complete responses (cr), whereas the three remain patients were non-responders (nr). of them one was rescued with an unrelated bmt, while two remained on eculizumab treatment. one of the cr relapsed at years showing clonal evolution and finally died. all the other patients are alive, keeping their hematological response. patients receiving a sequential therapy were one in pr and two in cr months after introduction of eculizumab. in conclusion, for patients diagnosed with severe aa/pnh syndrome intensive ist and eculizumab treatment, can be safely delivered either concurrently or sequentially, with an overall response rate of nearly %. this is the first systematic description of the management of severe aa in hemolytic pnh patients receiving eculizumab treatment. disclosure of conflict of interest: none. ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) in aa, ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) in rcc, and ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) years in rcmd. sixty-five patients underwent bmt from hla-matched (related , unrelated ) and from hla-mismatched (related , unrelated ) donors. conditioning regimens were used as follows; cyclophosphamide (cy)+antithymocyte globulin (atg) ± total body irradiation (tbi) (n = ), fludarabine (flu)+cy ± atg ± tbi (n = ), and flu +melphalan (mel) ± atg ± tbi (n = ). all patients got engraftment after bmt. however, late graft failure was found in patients with rcc, and with rcmd, but none with aa. out of patients who developed late graft failure, patients used flu+cy ± atg ± tbi, used cy ± atg ± tbi, and used flu +mel ± atg ± tbi for conditioning regimens. five-year cumulative incidence (ci) of graft failure was higher in rcmd ( ± . %) than in aa ( %) and rcc ( ± . %), significantly (po . ). five-year ci of graft failure tended to be higher in flu regimen ( ± . %) than in cy+atg ± tbi regimen ( ± . %), but not significant (p = . ). five-year ci of graft failure did not differ between with ( ± . %) or without tbi ( ± . %) (p = . ). multivariate analysis revealed that the morphological classification was a significant risk factor for graft failure (po . ). five-year failure free survival rate ( ± %) in rcmd was significantly lower than in aa ( ± . %) and rcc ( ± . %) (p = . ). graft failure, second malignancy, and death were considered as failure events. one patient with aa died of infection, four with rcc died of infection (n = ), bleeding (n = ) and myocarditis (n = ), and one with rcmd died of infection. five-year overall survival rates were not different among groups (aa, ± . %; rcc, ± . %; rcmd, ± . %) (p = . ). high incidence of graft failure in rcmd may be due to higher bm cellularity than in aa and rcc. the optimal conditioning regimen of bmt should be established for children with abmf based on the bm cellularity and morphological classification. disclosure of conflict of interest: none. recent studies have suggested inferior outcome of patients treated with rabbit atg (thymoglobulin, sanofi) as compared to horse atg (atgam,pfizer or lymphoglobulin, genzyme), and a higher early mortality with rabbit atg has been suggested to explain this difference. aim: to assess early mortality, response rates at , and months and long term outcome, in a large cohort of aa pts, treated in europe or asia with rabbit atg and cyclosporin, as first line treatment. eligible for this study were pts with aa, treated with thymoglobulin between and in europe (n = ) or asia (n = ). median year of treatment, was : characteristics were comparable : median age and years, interval diagnosis treatment ( and days) and severity of the disease ( % and % with vsaa). early mortality was analyzed for all patients.long term outcome was also analyzed for pts for who response data (no, pr, cr) were available. mortality o days was . % and . %, respectively, in the time period - and - (p = . ). in these time periods, early mortality for patients aged - , was reduced from . % to . % and for patients over , from % to %. overall response was recorded in patients. at months the cumulative incidence of response was comparable in the time periods: % vs %, and at year, % vs % (p = . ). response rates at months were age dependent: %, %, %, % respectively in patients aged - , - , - , (p = . ). when non responders at months were reevaluated at year, % had responded, % were non responders, % had died, and % had received other treatment. responses at months, were %, %, %, in pts with very severe, severe and non severe aa (p = . ). the actuarial year survival for the entire population was %, and %, when pts were censored as surviving at transplant. actuarial year survival in univariate analysis was as follows: % vs % for day responders vs non responders (p o . ), % vs % for males versus females (p = . ); %, %, %, % in pts aged - , - , - , years (p o . ); %, %, % in pts with neutrophils o . × /l, - × /l and . × /l (p o . ); %, %, % for pts with an interval diagnosis-treatment of o days, - days or days (p = . ). finally pts treated had a year survival superior to pts treated before ( % vs %, p = . ). survival at years, in the recent period ( - ), was % for pts aged - and % for pts over years. in multivariate cox analysis the following variables remained independent predictors of survival: patient age, year of treatment, severity of the disease, interval diagnosis treatment, and gender. thymoglobulin +csa is effective and safe in patients with aa. the outcome is mainly age dependent. the inrerval between diagnosis and treatment remains a strong predictor: the earlier the better. for pts o years old current early mortality. . for pts years of age, current early mortality is higher ( %), response rate ( %) and year survival ( %) are lower. . the actuarial year survival for the enire population was %. survival at years has improved from % (o o/u ) to % ( - ), especially for pts over years ( % vs %, p = . ). [p ] s disclosure of conflict of interest: we thank centers for providing up date follow up of their patients . this study was supported by a grant of sanofi-genzyme. both of the patients have extremely low anc o . × /l, reto . %, plt o × /l. both was given antibiotic treatment with carbapenem, vancomycin/linezolid, voriconazole or amphotericin b liposome and got no response. no pathogenic bacteria or fungus was found from either of the patients. both of them had no full sibling or matched unrelated donor and had their father as their haploidentical donor. bone marrow combined with peripheral blood stem cell (pbsc) was adopted. conditioning: fludarabine days − through − , ( mg/m × ), intravenous busulfan ( . mg/kg q h) on days − to − . gvhd prophylaxis: high-dose cyclophosphamide mg/kg on days + and + , mmf and tacrolimus since days + . rabbit anti-thymocyte globulin (thymoglobulin) . mg/kg on days − to − . stable neutrophil engraftment (anc . × /l) occurred on day + and day+ respectively. platelet achieved × /l on day + and day + , respectively. both transplant course was complicated by febrile neutropenia without detected etiology, while both children have no fever since the first day anc . × /l.the facial swelling was resolved in both patients except for palatal fistula and fistula of maxillary sinus as the sequela of severe nasosinusitis. no acute or chronic gvhd. case had hemorrhagic cystitis on day + which last for about days, and suspected thrombotic microangiopathy (tma) with hypertension, thrombocytopenia, elevated ldh and creatine on day + which was resolved soon after discontinue of tacrolimus. case had delayed engraftment of platelets and herpes simplex virus encephalitis on days + which was cured by ganciclovir and high dose intravenous immunoglobulin. now they are and months post-hsct respectively and are doing well with % chimerism and no gvhd. alternative donor hsct may be considered as the first line salvage therapy for patients of vsaa with extremely low anc and active infection. fast reconstruction of neutrophil helped to control the infection. hallo-identical hsct make sure nearly every patients can find a donor. ptcy is proved to be efficient and safety in gvhd prophylaxis and facilitating engraftment in these two challenging cases. disclosure of conflict of interest: none. long-term outcome of patients with severe aplastic anemia receiving allogeneic hematopoietic cell transplantation using nonmyeloablative conditioning with fludarabine and low dose total-body irradiation l cheryl xiu qi , l yeh ching , p michelle li mei , t lip kun , h william ying khee , g yeow tee , t patrick huat chye and k liang piu department of haematology-oncology, national university cancer institute, singapore and department of haematology, singapore general hospital, singapore allogenic haematopoeitic stem cell transplant (ahct) offers the best prospect of cure in patients with severe aplastic anaemia (saa). the use of myeloablative hct is however limited by the toxicity of preparative regimens, the lack of matched sibling donors, transplant related mortality and graft rejection. the introduction of non-myeloablative (nm) conditioning offers the possibility of extending this potentially curative treatment to patients in whom ahct was previously contradindicated. in , we reported the outcome of patients with saa who have received ahct using nonmyeloablative conditioning comprising of days of fludarabine at mg/m and total body irradiation at gy (flu + tbi gy). here, we report a longer follow-up, with additional patients who had recevied ahct with this regimen. fourteen patients with a median age of years old (range: - years old) received filgastrim-mobilised peripheral blood stem cell transplant from either hla identical sibilings (n = ) or matched unrelated donor (n = ) after receiving nm conditioning consisting of flu + tbi gy. the first two patients received cyclosporine (cya) and mycophenolate mofetil (mmf) for the post-transplant immunosuppessive therapy. the remainining patients received cya, mmf and a short course of methotrexate (mtx) for additional graft-versus-host dsease (gvhd) prophylaxis. results all patients achieved prompt engraftment. the median time for engraftment of neutrophils ( . × /l) and platelets ( × ) were days (range: - days) and days (range: - days), respectively. chimerism analysis on day and subsequently showed % donor cells in all patients except , who developed secondary graft failure at months and required salvage hct. none of the patients experienced grade and above regimenrelated toxicity. five patients developed grade ii-iv acute gvhd and patients developed limited chronic gvhd. with a median follow-up of . years (range: . - . years), the estimated overall survival and event free survival were % and % respectively. the two patients who did not receive mtx developed acute gvhd of the liver and succumbed to infective complications. the remaining patients who had received triple immunosuppressive therapy were well, with limited chronic gvhd seen only in . our results suggest that the nm conditioning regimen comprising of flu + tbi gy provides sufficient immunosuppression to allow prompt and stable engraftment with minimal regimen-related toxicity. it is an attractive option for patients with saa who require ahct but are at increased risk of regimen-related complications from more intensive cyclophosphamide-based regimens. disclosure of conflict of interest: none. paroxysmal nocturnal hemoglobinuria (pnh) is a rare acquired clonal disorder of hematopoietic cells characterized by the triad of hemolytic anemia, cytopenias and high risk of venous thrombosis. due to the rarity of the disease, most reported data derive from multicenter studies. we describe the natural history of the disease in a -year (yrs) long single center series of pnh patients (pts). we performed a retrospective analysis of consecutive pts followed at our center from to . since , the diagnosis was made by ham test; starting from , flow cytometry (fc) analysis was used to diagnose new pts and to confirm pnh in pts previously diagnosed by the ham test. at diagnosis, pts had classic pnh, aplastic pnh and intermediate form. the cumulative incidences of thrombosis, cytopenia and clonal neoplasm were %, % and %, respectively. except for pt with aplasia, no severe infections were diagnosed, nor renal failures or pulmonary hypertention. the yrs overall survival (os) was %. a nonsignificant better os was associated to the absence of thrombotic events ( % vs %) and to a diagnosis made during the last decade ( % vs % vs %).up to the treatment options were supportive care or allogenic bone marrow transplantation. since , eculizumab was used in transfusion-dependent patients and/or in case of a thrombotic history. overall, pts were transfusion-independent for the entire period of the illness, were transfusion-dependent and/or had thrombotic events( pts). six of the latter pts never received eculizumab but only transfusion support ( pts) or allogeneic bone marrow transplant ( pts), while pts received eculizumab (the first pts were included in the phase iii triumph and shepherd trials).considering the increased risk of meningococcus infection for pts on eculizumab, vaccination with conjugated anti-meningococcus serotypes acwy was employed and, since , conjugated antimeningococcus serotype b was added. overall, pts treated with eculizumab became transfusion-independent and four remained transfusion-dependent. no thrombotic event was observed after eculizumab, even if pts had recurrent thromboembolisms prior to receiving the drug. no severe infection was documented. one patient developed extravascular hemolysis and receive a successfully selective splenic artery embolization. the yrs os in the eculizumab group was %.no pnh-associated death occurred. our study confirms that thrombosis is a major complication in pnh pts not receiving eculizumab, influencing os. the better os in the last decade is probably due to the use of eculizumab and to lack of thrombotic events. in particular, for pts on eculizumab the year os was %, even though half of the pts had thromboembolism and diagnosis made prior to the last decade. although kidney failure and lung hypertension have been reported, we did not observe these complications in our long follow-up case series. we can assume that the availability of a dedicated emergency room at our center allows to perform, promptly, hyper-hydration or transfusion support in case of hemoglobinuria crisis, reducing the risk or organ damage. no infections have been observed after eculizumab, probably due to the vaccination program schedule recommended in the literature, plus the addition of conjugated anti-meningococcus serotype b. however, shared guidelines are needed. disclosure of conflict of interest: none. mortality following hsct in saa pts over the age of is reported to be in the order of %, without taking in to account long term sequelae such as chronic gvhd, known to be more frequent in older patients. this has prompted international guidelines to recommend first line immunosuppressive therapy above years of age. the question is whether this is still true in . the aim of the study is to assess whether trm in saa patients grafted - is reduced,as compared to the era - . we used the wpsaa ebmt registry, and identified pts aged years or more, with acquired saa, grafted between and . we divided pts in transplant eras: - (n = ) and - (n = ). in the more recent period ( - ) pts were older ( vs year, p o . ), were more often grafted from alternative donors (alt) ( % vs %, po . ), with a greater use of bm ( % vs %, p o . ), and with a longer interval dx-tx ( vs days, p . ), and more often received a fludarabine containing regimen ( % vs %, p o . ). the os year of pts grafted in - was %, compared with % for pts grafted - (p = . ). in multivariate analysis, including the interval diagnosis transplant, patient's age, donor type, stem cell source and conditioning regimen, the lack of improved survival in - was confirmed (p = . ). a very strong age effect was shown both in univariate and multivariate analysis: survival of pts aged - years, - years and years, was respectively %, %, % (p o . ) and this was confirmed in multivariate analysis. the conditioning regimen, also proved to be a significant predictor, with improved survival for alt transplants receiving flu containing regimens ( % vs %, po . ). in general pts receiving either cy or a flu containing regimen, did significantly better than pts receiving other preparative regimens ( % vs %, p = . ). the use of a sibling donor (sib) did not prove to predict survival in multivariate analysis. pts receiving campath in the conditioning,did significantly better than pts not receiving campath ( % vs % po . ); similarly survival of patients with atg was superior % vs % compared to patients not receiving atg (p o . ). when pts receiving either campath or atg (n = ) were compared to patients not receiving either (n = ), the difference in survival was % vs % (p o . ), and this was significant also in multivariate analysis. combined primary and secondary graft failure was reduced from % to % in the two time periods (p = . ), acute gvhd grade ii-iv was reduced from % to % (p = . ) and chronic gvhd was also reduced from % to % (p = . ) infections remain the leading cause of death in both transplant eras ( % and % respectively), followed by gvhd ( % and %) and graft failure ( % and %), whereas ptld have been reduced from % to . %. hsct in pts with acquired saa aged and over, continues to carry a significant risk of trm also in - , ranging from % in younger pts ( - ) to % in older pts ( years). survival is predicted in multivariate analysis, by two crucial predictors: patients' age and the use of either campath or atg,the latter giving a % survival advantage over no campath/atg. alt and sib donors produce similar survival. this study gives further support to current guidelines, suggesting first line therapy with atg+csa, in pts over the age of . [p ] disclosure of conflict of interest: none. autoimmune diseases p allogeneic haematopoetic stem cell transplantation as curative therapy for early-onset, refractory crohn's disease e groene , p bufler , k krohn , s immler , g marckmann , t feuchtinger , s koletzko and m albert dr. von hauner university children's hospital and institute of ethics, history and theory of medicine, lmu results of a recent randomized trial suggest that autologous hsct is an option in adult patients with severe, therapyrefractory crohn's disease (cd) with an associated mortality risk of %. however, relapse of the disease is frequent ( ). in contrast allogeneic hsct has resulted in long-term cure of cd in affected patients transplanted because of haematological malignancy ( ). we report a year old girl who was diagnosed with severe cd at age seven (paris classification l , l a, b ). neither next generation sequencing nor immunological work up identified a monogenetic cause of cd. progressive chronic inflammation manifesting ubiquitously in the gastrointestinal tract resulted in severe complications, such as perianal fistulas with rectal stenosis, intestinal abscesses, dysphagia, severe weight loss, failure to thrive, delayed puberty and the need for ileostomy and long-term exclusive enteral nutrition via tube feeding. despite multiple lines of therapy, including repeated nutritional therapy, steroids, immunosuppressants (methotrexate, azathioprine) and biologicals (infliximab, adalimumab, certolizumab) a lasting remission could not be achieved resulting in poor quality of life. after careful risk/benefit assessment including ethical counselling allogeneic hsct was offered. she underwent allogeneic hsct from a matched ( / ) unrelated bone marrow donor ( . × /kg total nuclear cells). conditioning was performed according to a protocol successfully applied in adolescents with chronic granulomatous disease ( ) with alemtuzumab ( × . mg/kg/d), targeted busulfan (tauc ng × h/ml) and fludarabine ( × mg/m ). cyclosporine a and mycophenolate mofetil were used as gvhd prophylaxis. neutrophil and platelet engraftment were observed on days + and + , respectively. the post hsct course was complicated by grade i acute skin gvhd treated with topical steroids and toxic megacolon secondary to scarring stenosis on both ends of the unused colon on day + requiring surgery and a colostomy. at months post hsct the patient is well, off immunosuppressive medication, without gvhd and exhibiting % donor chimerism. the cd is in complete clinical and histological remission as proven by endoscopy and biopsies. stoma reversal with restitution of intestinal continuity is planned for the next months. refractory cd can lead to life-threatening complications and severely reduced quality of life. although long-term outcome in our patient will need to be carefully assessed, allogeneic hsct may offer a curative therapy in children and young adults with severe cd, even in the absence of an identified monogenetic cause. current ebmt recommendations include consideration of ahsct in exceptional circumstances for patients with severe refractory cd. the only randomised trial of ahsct in cd (astic) confirmed substantial short-term benefits but failed to meet its primary year endpoint. to further clarify the longterm safety and efficacy of ahsct in cd we performed a retrospective analysis of patients undergoing ahsct for cd outside the astic trial using the ebmt registry. patients were identified from the ebmt registry. all adult patients undergoing ahsct for a primary diagnosis of cd from to were eligible for inclusion. patients who were enrolled in the astic trial were excluded. from a total of patients (across centres) on the ebmt registry, data were obtained from patients transplanted in centres in countries. median patient age was yrs (range: - ) and % were female. median age at first diagnosis of cd was yrs (range: - ). patients were heavily pre-treated, having failed or been intolerant to a median of previous lines of therapy (range: - ). % had received experimental therapy prior to auto-hsct. % of patients had undergone at least operation. the median time from first diagnosis of cd to auto-hsct was . years (range: . - . ). all patients received peripheral blood stem cells following conditioning with cyclophosphamide mg/kg and % received anti-thymocyte globulin (atg). the median cd + dose infused was . (range: . - . ) × /kg. twelve percent of patients underwent cd + selection. neutrophil and platelet engraftment occurred at a median of day (range: - ) and day (range: - ), respectively. sixety-one percent received post transplant g-csf. median length of follow-up following auto-hsct was months (range: - ). at days post auto-hsct, % of patients were in clinical remission (cr), defined as no abdominal pain and normal stool frequency. a further % experienced significant improvement, defined as improvement in abdominal pain and stool frequency. for % there was no appreciable change in disease and in % the disease worsened compared to baseline. at year post auto-hsct, % were in cr, % were improved, % were unchanged and % had worsened. at last follow-up, % were in cr, % were improved, % were unchanged and % had worsened. overall % restarted medical therapy post auto-hsct and % required further surgery. overall % developed an infection requiring treatment post auto-hsct ( % bacterial, % viral). ebv and cmv reactivation occurred in % and % respectively and herpes zoster occurred in %. a secondary autoimmune disease developed in %, most commonly thyroid disease ( %). malignancy developed in %, of which skin cancer accounted for % of cases. one patient died at days post auto-hsct due to cmv infection, sepsis and multiorgan failure. this large retrospective series further supports the safety and efficacy of ahsct in a population with severe and treatment-refractory crohn's disease, % of patients experienced complete remission or significant improvement in cd symptoms with long-term follow-up. trm observed was similar to ahsct for other indications. in summary, ahsct appears to be an extremely promising therapy for severe refractory cd. further follow up of astic patients and future randomised trials are warranted. disclosure of conflict of interest: none. memory stem t cells (tscm) are long living self-renewing memory t cells with long-term persistence capacity, which play a relevant role in immunological memory and protection against infectious diseases and cancer , , , , , . the aim of this work is to investigate the potential role of tscm as a reservoir of arthritogenic t cells in rheumatoid arthritis (ra). we analysed the dynamics of circulating tscm (here identified as cd ra+ cd l+ cd + t cells) and other memory t-cell subpopulations by multiparametric -color flow cytometry in patients with active ra and in of them also during treatment with anti-tnfα biological agents (etanercept). to analyse cytokine productions, functional assays were performed stimulating peripheral blood mononuclear cells (pbmcs) with pma/ionomycin and brefeldin a. after the stimulation, cells were stained for surface markers, fixed and stained for intracellular cytokines. we traced circulating antigen specific cd + t cells for the vimentine-derived citrullinated peptide (vimcit) savracitssvpgvr , in hla-drb × : ra patients before and during the anti-tnfα treatment using custom mhc class ii tetramers. viral antigen specific cd + t cells were traced using mhc class i dextramers. age-matched healthy donors (hds) were used as control for all the experiments. we found a significant expansion of cd + tscm in patients with active ra both in terms of frequency and absolute counts. notably, cd + tscm significantly contracted upon anti-tnfα treatment, suggesting a role of tnfα in tscm accumulation. in contrast to cd +t cells, cd compartment did not show significative alterations compared to (hds). furthermore, cd +tscm in ra patients displayed an enrichment in the th phenotype, largely implied in autoimmune disorders, while the other t cell subpopulation were not enriched in the th phenotype. at the antigen specific level, we were able to trace in hla-drb × : patients antigen specific cd + t cells, comprising tscm, specific for the vimentin-derived citrullinated peptide. of notice, citrullinated vimentin specific cd + t cells, including tscm, contracted during anti-tnfα administration, while viral-specific cd + t cells (ebvbhrf- ) and antiviral cd specificities (cmvpp , flump, ebvbmlf- ) were not affected by etanercept administration, thus suggesting a possible role of cd + tscm as reservoir of arthritogenic autoreactive t cells. overall, our results suggest that tscm, by representing a long-term reservoir of undesired specificities, might play a non redundant role in sustaining ra and possibly other t cell mediated disorders, thus representing novel biomarkers as well as therapeutic targets. ongoing experiments will characterize the tcr repertoire on sorted tscm and cd + memory subsets in order to identify a possible oligoclonality in tscm repertoire. in conclusion, the analysis of tscm dynamics in autoimmune disorders could have relevant clinical implications as new biomarkers and for devising innovative therapeutic strategies. ebv and cmv reactivation following autologous haematopoietic stem cell transplantation (hsct) for autoimmune neurological diseases resolves spontaneously and rarely requires treatment c mapplebeck , b sharrack , h kaur , y ezaydi , h jessop , l pickersgill , l scott , m raza and ja snowden departments of haematology, neurology and virology, sheffield teaching hospitals nhs foundation trust, sheffield, uk autologous haematopoietic stem cell transplantation (hsct) for severe autoimmune diseases involves immunosuppressive conditioning regimens and current guidelines recommend monitoring for viral reactivation of cytomegalovirus (cmv) and epstein barr virus (ebv) (snowden et al ) . however, the incidence, degree and management of viral reactivation are not established. we performed a retrospective observational service evaluation study of all patients receiving cyclophosphamide mg/kg + rabbit anti-thymocyte globulin mg/kg (atg, thymoglobulin) followed by autologous hsct for various autoimmune neurological diseases between and at our centre. data collected included the baseline serological status of the patient prior to transplant and serial blood pcr quantitation (copies/ml). if ebv and cmv reactivation occurred details of further management was collected and descriptive statistics were used to summarise outcomes. twenty-three patients received autologous hsct between january and october ; patients with multiple sclerosis (ms), with chronic inflammatory demyelinating polyneuropathy (cidp) and with stiff person syndrome. twenty-two patients had positive ebv igg serology prior to transplant and patient had an equivocal result. seventeen patients had evidence of ebv reactivation and a further patient had ebv dna detected post-transplant but with less than copies/ml. the average time to peak ebv pcr was . (range: - ) days post-transplant and a range: in ebv pcr peak level from to copies/ml. the patients who had ebv pcr results of over copies/ml had ct scans of chest, abdomen and pelvis performed which did not demonstrate significant lymphadenopathy or hepatosplenomegaly. in all patients monitored for a detectable ebv reactivation, the ebv pcr spontaneously began to fall within months (average days, range: - days) post-transplant and no specific treatment was required. one patient had late ebv reactivation of copies/ml at months post-hsct associated with chronic tonsillitis and tonsillectomy specimens showed follicular hyperplasia without evidence of post-transplant lymphoproliferative disorder (ptld) and ebv pcr levels normalised without other treatment. ( %) patients had positive cmv igg serology prior to transplant and one patient had an equivocal result. only of patients had a significant reactivation of cmv with copies/ml at days post-transplant, successfully treated with intravenous immunoglobulins and valganciclovir. two other patients had low level cmv reactivation with and copies/ml, respectively which resolved spontaneously without treatment. ebv reactivation in patients with neurological autoimmune disease undergoing autologous hsct is common and usually resolves spontaneously without treatment. asymptomatic cmv reactivation occurs in approximately % of patients in this setting and may require treatment. autologous hematopoietic stem cell transplantation (hsct) has been utilised for the treatment of severe multiple sclerosis (ms). it results in significant improvement of neurological function, although patients can experience exacerbations of ms-related symptoms during the procedure. we reviewed patients with ms who underwent stem cell mobilisation and collection from march to november . the median age was years ( - ). nine patients ( %) were male. the interval from diagnosis to hsct was . months (range: . - . ). patients ( %) had relapsing-remitting (rrms), six patients ( %) secondaryprogressive (spms) and two patients ( %) primary-progressive (ppms) multiple sclerosis. only patients ( %) had not received any prior treatment, whereas patients ( %) received two prior treatments, three patients ( %) received three treatments and two patients ( %) received four treatments. the median expanded disability status scale (edss) score was (range: - ). peripheral blood stem cells were mobilised with cyclophosphamide (cy) g/m on day + and daily gcsf ( μg/kg subcutaneously) from day + until the completion of the harvest. hsct was performed at a median of days after mobilisation (range: - ). the conditioning regimen consisted of cy ( mg/kg/day from day − to − ) and atg ( mg/kg/day from day − to − ). exacerbation of ms symptoms was defined as the appearance of new or worsening of old symptoms for at least h duration in a previously stable ( weeks) patient. of the total cohort, patients ( %) underwent mobilisation with cy+gcsf uneventfully. only two patients ( %) had an exacerbation of ms requiring hospital admission after collection (one with fatigue and increase of spasticity, other with worsening weakness). no patient required hospital admission during the mobilisation procedure. the median cd + cell dose was . × /kg (range: . - ). the median number of apheresis was ( - ). a total of patients have undergone hsct at the time of this analysis. during transplant a total of patients ( %) experienced an exacerbation of ms. of these, % (n = ) before day and % (n = ) between day + and + . symptoms of exacerbation were: muscle spasms in patients ( %), weakness and reduced power of limbs in patients ( %), increase instability and tremor in two patients ( %) and one patient ( %) with worsening of neuropathic pain. only three patients ( %) received treatment with methylprednisolone for ms exacerbation and symptoms had fully resolved by discharge in all patients. other transplant complications included neutropenic fever in all, invasive fungal infection in , fluid overload in ( %) and atg related complications in ( %) such as fever (n = ) and pericarditis/serositis (n = ). the median time to neutrophil engraftment was days ( - ) and the median duration of hospital admission was days ( - ). exacerbation of ms symptoms is common during hsct and can also occur during mobilisation. in our hands, after cy and gcsf mobilisation only two patients ( %) developed an exacerbation of ms symptoms compared with patients ( %) after ct and atg conditioned hsct. it is possible that the addition of atg to cy triggers an immunological response involved in this transient deterioration of the ms symptoms. further studies are required to confirm this hypothesis. disclosure of conflict of interest: none. inflammatory immune response syndrome (iris) is a noninfectious worsening of neurological condition during immune recovery and has been documented to occur in hiv and in multiple sclerosis following alemtuzumab. the manifestation of iris includes headache, nausea, weakness, neurologic deficits, and mri enhancing lesion. we report three cases of iris after autologous non-myeloablative hematopoietic stem cell transplantation (hsct) in patients for which the transplant indication was an inflammatory neurologic disease: neuromyelitis optica (nmo), chronic relapsing inflammatory optic neuritis (crion), and multiple sclerosis (ms). mobilization was with cyclophosphamide gr/m and gcsf. conditioned regimen was mg/kg cyclophosphamide ( mg/kg/d) and . mg/kg ratg (thymoglobulin). the conditioning regimen for nmo and crion also included mg rituximab. case . a years old african-american female with systemic lupus erythematosus (sle) and nmo was discharged day and readmitted on day for fever, headache, progressive altered mental status with dysarthria and legs. brain mri had numerous t /flair hyperintense and enhancing lesions in the subcortical and periventricular white matter. a lumbar puncture was negative for infection including jcv. complete recovery occurred after treatment included high dose of steroids and plasmapheresis. case . a years old female with crion experienced blindness, weakness and slurring of speech three months post hsct. mri showed a large enhancing brain stem lesion. lumbar puncture was jcv negative. complete recovery occurred after solumedrol and rituximab. mri months later demonstrated complete resolution of the enhancement with return of vision to baseline. case . a ten year-old boy diagnosed with paediatric ms developed hemichorea seven days after hsc reinfusion. brain mri revealed a gadolinium-enhancing lesion in the contralateral basal ganglia. lumbar puncture was negative for infection including jc virus. symptoms resolved spontaneously after seventeen days. the appearance of new neurologic symptoms and mri enhancing lesions early after autologous hsct is unexpected and may be related to lymphocytes in the graft, immune recovery post engraftment, and or persistent auto-antibodies. it is mandatory to perform a lumbar puncture to exclude the possibility of infections including progressive multifocal leukoencelopathy (pml) due to jcv. the timing of presentation, the negativity of jc viral load, and the complete recovery with or without immune suppression suggest the hypothesis of iris, as an epiphenomenon of the immune reconstitution following autologous hsct for neurologic diseases disclosure of conflict of interest: none. hematopoietic stem cell transplantation is the effective method of therapy for cns autoimmune disorders in children. long-term outcomes and late effects estimation required. the aim of the study is to estimate long-term outcomes and late effects at children underwent auto-hsct for multiple sclerosis (ms) and allo-hsct for neuromyelitis optica (nmo). twelve pts. with ms and pts. with nmo were included to the analysis. ms pts. gender: female - % (n = ), male - % (n = allogeneic haematopoietic stem cell transplantation (hsct) remains the sole curative option for patients with myelofibrosis (mf). although a spectrum of conditioning regimens has been used, the optimal preparative treatment before hsct remains to be defined. we did a phase ii randomized study at transplant centers in italy with the aim of comparing the reduced-intensity conditioning (ric) fludarabine-busulfan (fb) (conventional arm), that had been already tested in the prospective ebmt study ( ) with the ric fludarabine-thiotepa (ft) (experimental arm), that has been widely used in italy in the last two decades ( ) . eligible to this study were patients with primary mf or a mf subsequent to a previous essential thrombocytemia or polycyhemia vera, an age ≥ ≤ years, a karnofsky performance status , a comorbidity index o o/u and with at least one of the following unfavorable prognostic factors: anemia (hb o g/dl), leukocitosis ( × /l), circulating blasts % or constitutional symptoms. patients were randomized to receive intravenous busulfan . mg/kg for doses or thiotepa mg/kg for two doses associated to fludarabine mg/m for impact of pre-transplant ruxolitinib in myelofibrosis patients on outcome after allogeneic stem cell transplantation syed abd kadir, sharifah shahnaz, author , , zabelina, tatjana, co-author , christopeit, maximilian, co-author , wulf, gerald, co-author , wagner, eva, co-author , bornhaeuser, martin, co-author , schroeder, thomas, co-author , crysandt, martina, co-author , mayer, karin tina, co-author , stelljes, matthias, co-author , badbaran, anita, co-author , wolschke, christine, co-author , ayuk ayuketang, francis, co-author , triviai, ioanna, co-author , wolf, dominik, co-author ruxolitinib (rux) is the first approved drug for treatment of myelofibrosis. because spleen size and constitutional symptoms may influence outcome after allogeneic stem cell transplantation (asct), rux is recommended before stem cell transplantation in order to reduce therapy-related morbidity and mortality and improve outcome (ebmt/eln recommendation, leukemia ) the aim of this retrospective study was to evaluate the impact of pretreatment with rux in comparison to transplantation of rux-naïve mf patients with regard to outcome after asct. we included myelofibrosis patients (pts) with a median age of years (range: - ) who received asct between and from related (n = ), matched (n = ) or mismatched (n = ) unrelated donor. all patients received busulfan-based reduced intensity conditioning. while pts ( %) did not receive rux, pts ( %) received rux at any time point prior to asct. the median daily dose of rux was mg (range: - mg) and the median duration of treatment was days (range: - days). in pts rux was stopped before stem cell transplantation because of no response or loss of response, while in pts rux was given until start of conditioning. gvhd prophylaxis consisted of cni plus short course mtx or mmf and anti-lymphocyte globulin. according to dynamic ipss (dipss) (n = ) the patients were either low (n = ), intermediate- (n = ), intermediat- (n = ), or high risk (n = ). as the median follow up was shorter for patients treated with rux ( vs months, po . ). primary graft failure was seen in pts in the rux and three in the non-rux group. the median leukocyte engraftment was days (range: - ) in the ruxolitinib and days (range: - ) in the non rux group (p = . ). the incidence of acute gvhd grade i to iv was significantly lower in the rux group ( % vs %, p = . ), while agvhd grade ii-iv ( % vs %, p = . ) and grade iii/iv ( % vs %, p = . ), did not differ significantly. the ci of nrm at year was % ( % ci: - %) for the rux group and % ( % ci: - %) for the non-rux group (p = . ), and the ci of relapse at years was % ( % ci: - %) vs % ( %ci: - %, p = . ). the years rfs and os was % ( %ci: - %) and % ( %ci: - ) for the rux group and % ( % ci: - %) and % ( % ci: - %) for the non-rux group (p = . and p = . , respectively). within the rux group (n = ), pts responded to rux (more than % spleen size reduction), while pts did not respond or lost response prior to stem cell transplantation. here, no significant difference could be seen between the responding and non-responding group for nrm ( % vs %, p = . ), relapse ( % vs %, p = . ), rfs ( % vs %, p = . ) and os ( % vs %, p = . ). in a multivariate analysis including rux treatment as variable there was a non-significant trend in favor for in the tyrosine kinase inhibitor (tki) era, allogeneic haemopoietic stem cell transplantation (allo-hsct) has become the later-line therapy but still remains the only known curative treatment for chronic myeloid leukemia (cml). since the introduction of tki in our centre in , the trend of allo-hsct among our cml cohort has changed over time. the purpose of this study is to examine hsct outcomes of our cml cohort who was either tki naïve or has received tki therapy prior hsct. between may and december , cml patients in our center received allo-hsct with % were tki naïve. the time of diagnosis to transplant was significant shorter among the tki naive group as compared to those received tki prior hsct ( . ± . months versus . ± . months, respectively). there were no gender different ( % males) but the median age at hsct was younger among tki naïve group, . years (range: - years) versus . years (range: - years) respectively. malays remained majority ethnic group but the percentage was reduced among patients received tki prior hsct ( . % versus . % respectively). the disease phase at hsct was significant different whereby majority of tki naïve group was in first chronic phase (cp ) ( . %) as compare to patients with prior tki exposure ( . %). all the patients in the tki naïve group received hla-matched related siblings donor (mrd) with . % marrow stem cell source whereas only . % of patients who have prior tki exposure received mrd with . % were from peripheral blood stem cell (pbsc). all patients in the tki naïve group but only . % among patients who have prior tki exposure received full myeloablative conditioning regimen. there was slower neutrophil and platelet engraftment ( . ± . days versus . ± . days and . ± . days versus . ± . days respectively) among tki naive group. at june , the -year overall survival (os) of cml at all disease status was % in tki naive group versus % for patients who have prior tki exposure and transplanted in more advance disease stage. in general, patients in cp have the best os. there was higher incident of grade to acute graft-versus-host-disease (gvhd) among the tki naïve group ( . % versus . %, respectively) likely due to intensity of conditioning regimen with no significant different in chronic gvhd incident. similarly, there was higher relapse rate among tki naive patients ( . % versus . %, respectively) as upfront post transplant tki was not routinely given to this group of patients in the past. further multivariate analysis to ascertain predictors of transplant outcome among this cohort of patients included disease status, donor-recipient gender combination, ethnic difference will be presented. in conclusion, despite emergent of effective and potent next generation tki, hsct still has it role as curative modality for patients who failed tki. as showed in our data, the transplant outcome is excellent for patients who remain in cp at the time of hsct and it is important to identify patients earlier, before disease progression, especially young patients, in order to optimize transplantation outcomes. disclosure of conflict of interest: none. the purpose of this analysis was to provide -year follow-up of the gcllsg cll x trial which aimed at evaluating reducedintensity conditioning (ric) hsct in patients with poor-risk cll. the cll x trial included patients (median age ( - ) years), of whom patients were allografted with blood stem cells from related ( %) or unrelated donors ( %) using fludarabine-alkylator-based ric regimens. % had refractory cll at hsct, and % had a tp deletion and/or mutation. the -year follow-up of the trial including the observation that genetic risk factors such as tp lesions and sf b and notch mutations had no prognostic impact has been previously reported. survival and relapse information was requested for all patients who underwent hsct within the cll x trial in german centres (the canadian centre was unavailable for follow-up) and were alive at the -year followup. results: follow-up information was received for / patients ( %) alive at the -year follow-up. of these, patients had died ( cll, chronic gvhd, secondary cancer), and had experienced disease recurrence. with a median follow-up of survivors of . ( . - . ) years, -year nrm, relapse incidence (rel), event-free survival (efs), and overall survival (os) of all patients allografted was %, %, % and %, respectively, without significant effects of tp lesions on outcome. absence of minimal residual disease (mrd) at the -month landmark post hsct was highly predictive for a reduced relapse risk, in particular if mrd eradication occurred only after immunosuppression withdrawal, suggesting of effective graft-versus-leukemia activity (gvl; -year rel %). in the patients who were alive and event-free years post allohct, nrm, rel, efs, and os years after this landmark (or years after transplant) was . %, %, %, and % with a median follow-up of . years ( . - . ) after the -year landmark. notably, no relapse event occurred beyond years post hsct. of those who remained event-free beyond years, all patients who were available for mrd assessment at their most recent follow-up were mrdnegative. altogether of the allografted patients had cll recurrence after transplant; between and , and from onwards. whilst the median survival of those patients who relapsed during the earlier period was months, all patients with late relapse are currently alive - (median ) months after the event. conclusions: long-term observation of patients allografted in the cll x trial confirms that ric hsct can provide gvl-mediated sustained disease control in a sizable proportion of patients with poor-risk cll independent of the tp status. patients who are in mrdnegative remission one year after hsct have an % probability of remaining disease-free at least for years. however, late relapses do occur but may benefit from strategies involving innovative pathway inhibitors. hallek: consultancy and speakers bureau for pharmacyclics, llc and an abbvie company; speakers bureau for janssen; m. kneba: consultancy, honoraria, travel grants and research funding from gilead and roche; consultancy, honoraria and travel grants from abbvie and janssen-cilag; research funding from amgen; travel grants from glaxo-smithkline; p.dreger: consultancy for roche and janssen; consultancy and speakers bureau for novartis and gilead. no evidence for an increased gvhd risk associated with post-transplant idelalisib given for relapse of chronic lymphocytic leukemia or lymphoma: first results of a survey by the ebmt chronic malignancy and lymphoma working parties p dreger , , a boumendil, l koster , c scheid idelalisib is a kinase inhibitor (ki) approved for the treatment of cll and follicular lymphoma (fl). idelalisib has a specific adverse effect profile including immune-mediated inflammatory conditions such as colitis and pneumonitis, raising concern about the safety of this ki if administered for treatment of malignancy recurrence after allogeneic hematopoietic cell transplantation (allohct). the purpose of this ongoing study is to provide information on the safety and efficacy of idelalisib in this setting. we included in this study adult patients who had been registered with the ebmt for an allohct for cll or lymphoma and who received idelalisib for treating disease relapse or persistence at any time after transplant as indicated by participating investigators upon request by the ebmt study office in leiden. baseline patient, disease, and transplant data were collected from med-a forms. centers were requested to provide additional treatment and follow-up information. as of november , , a total of patients have been registered, of whom a full dataset as required for this analysis was available for patients (cll , fl , diffuse large b-cell lymphoma (dlbcl) , peripheral t-cell lymphoma , unspecified ) who had undergone allohct between july and april . all patients except one were male. median age at transplantation was ( - ) years and the median interval from diagnosis to allohct was . ( . - . ) years. prior to allohct, patients ( cll and lymphoma) had received an autohct and two other patients had been exposed to ki (idelalsib , ibrutinib ). disease status at allohct was sensitive in % of the patients. conditioning was reduced-intensity in % of the transplants and included in vivo t cell depletion in the majority of cases ( %). donors were identical siblings in % with pbsc being the stem cell source in all cases. the interval between hct and idelalisib commencement was ( - ) months in the cll group but only ( - ) months in the lymphoma group. prior to idelalisib, grade ii-iv acute gvhd and chronic gvhd had been observed in % and % of the patients, but was still active at the time of idelalisib commencement in only two cases ( %) . four patients with cll had already failed ibrutinib given for post-hct relapse prior to idelalisib. the median time on idelalisib until documented withdrawal or event (progression, retreatment, death) was ( - ) days. after start of idelalisib, one patient developed grade acute gvhd and subsequently chronic gvhd, however, in this patient idelalisib was started as early as days after transplant. efficacy of idelalisib in this high-risk patient sample was limited with only one pr in the cll group (stable disease , progressive disease , not available ; lymphoma not available), translating into a median event-free survival after start of idelalisib of days. five patients with cll underwent a subsequent treatment with an alternate ki (ibrutinib , venetoclax ). altogether, there were five deaths, all due to diease progression (cll , lymphoma ). median overall survival was days for the whole sample and not reached for cll. this preliminary data does not support concerns about the safety of idelalisib in the post allohct setting. updated results of this ongoing study will be presented at the meeting. tested patients ( %) achieved a ccyr and at least a mmolr, respectively. the response to transplant by day assessment correlated significantly with the disease status before transplant. a higher percentage of patients who experienced cytogenetic response before transplant experienced molecular response post-transplant ( %) compared with those who did not ( %; p = . ). for the entire group, the -year cumulative incidence (ci) of acute gvhd grade ii-iv and grade iii-iv were % and %, respectively; -year ci of extensive chronic gvhd was %. there was no significant difference in the ci of severe acute or chronic gvhd between donor types. the ci of nrm at days and year was % and %, respectively. the ci of cytogenetic and molecular relapse at years was % and %, respectively. overall the -year os, pfs and gvhd-free, relapse-free survival (grfs) were %, %, and %, respectively. in multivariable analysis for grfs, transplant in cp and the use of haploidentical donor significantly associated with better grfs. the -year grfs of patients in cp , ap and bp before transplant was %, % and %, respectively (p = . ). ( figure a ) patients receiving a haploidentical donor had a better -year grfs when compared with hla matched transplants ( % vs %, p = . ). ( figure b ) for pfs, transplantation in cp , using a haploidentical donor and mac regimen associated with better pfs while age, cytogenetic and molecular response before transplant did not predict survival. ahsct is curative for a proportion of patients with advanced cml. pfs and grfs are favorably influenced by percentage of bm blasts and donor type, with haploidentical donor having at least as good outcomes as hla matched donors, while molecular and cytogenetic response before transplant do not appear to correlate with survival posttransplant disclosure of conflict of interest: none. allogeneic stem cell transplantation (sct) has been considered as the treatment of choice for younger patients (pts) with high-risk chronic lymphocytic leukemia (cll). role of allogeneic sct in era of novel drugs is widely discussed. here we present our results after sequential use of chemotherapy and reduced-intensity conditioning (ric) in cohort of high-risk cll pts. high-risk cll was defined by one of the following: disease refractory to purine analogs, short response or early relapse (within months) after purine analog combination treatment, and/or progressive disease with unfavorable genetic abnormalities (del [ p]/tp mutation). we analyzed pts with high-risk cll undergoing chemotherapy and ric sct in our centre from august to june . the median of pretransplant lines were (range: - ), novel drugs (idelalisib, ibrutinib) were used in % of pts ( / ). fludarabine ( mg/m ) and cytarabine ( g/m ) for days (fc) were used for cytoreduction in all pts. after days of rest, ric consisting of gy tbi, anti-thymocyte globulin - mg/ kg/day for days, and cyclophosphamide - mg/kg/day for days followed. median age of pts was years (range: five-year overall (os) and relapse-free survival (rfs) was % and %. ci for cgvhd in pts surviving more than months post-hct was % after years and % after years. in a multivariate cox-regression model the occurrence of cgvhd independently improved os (p = . , hr . ; % ci . - . %) as well as rfs (po . , hr . ; % ci . - . ). high risk dipss plus score demonstrated significant inferior survival compared to intermediate- (os p = . ; rfs p = . ), int- (os p = . ; rfs p = . ) and low risk (os p = . ; rfs p = . ) which could be confirmed in multivariate analysis for os (p = . , hr . ; % ci . - . ) and rfs (p o . , hr . ; % ci . - . ). rfs additionally was improved by splenomegaly (n = ) vs. normal spleen size (n = ) at time of hct (p = . , hr . ; % ci . - . ). ruxolitinib (n = ) or none (n = ) pre-treatment compared to other drug therapy (n = ) resulted in improved os (p = . ) and rfs (p = . ) and was an independent factor for rfs in multivariate analysis (p = . , hr . ; % ci . - . ). non-relapse mortality (nmr) was significantly determined by high-risk dipss plus score (p = . ) or dipss high and int- (p = . ). relapse incidence was significantly lower in pts with splenomegaly compared to asplenic or normalspleensized pts prior to hct (p = . ). our data point out that pre-therapy and dipss or dipss plus score are relevant pre-transplant outcome factors while chronic gvhd is the most important independent hct-related factor. furthermore, splenomegaly at hct reduces risk of relapse and therefore improves rfs. [p ] disclosure of conflict of interest: none. thalassaemia major affects new babies in india each year and haematopoietic stem cell transplantation offers the only chance of cure. we present data on children with thalassaemia major aged between months and years using a uniform conditioning regimen consisting of thiotepa mg/kg, treosulphan gm/m and fludarabine mg/m . equine antithymocyte globulin at a dose of mg/kg was added to children who were undergoing transplantation from an unrelated donor source. there were eight deaths before engraftment due to sepsis or bleeding and two related to graft versus host disease. all patients showed complete chimerism on day . however, in children there was an acute drop in donor chimerism between day and post transplantation. immunosuppression was abruptly stopped when donor chimerism dropped below % in all children. seven children responded well and re-established complete chimerism with this measure. seven children progressed to develop complete graft loss. donor lymphocyte infusion (dli) in the form of small aliquots of peripheral whole blood from the donor was administered in seven children. dli was used in a graded fashion every weeks starting from × /kg of cd , followed by × /kg and × /kg. all of them continued to maintain their graft with these interventions. drop in chimerism was seen particularly in children less than years at the time of transplantation comprising out of children. older children with lucarelli class iii were also prone to rejection in our earlier series and this complication has now been eliminated with pre-transplant immunosuppression and hypertransfusion. children above the age of years were more prone to graft versus host disease and required on average months of immunosuppression. treosulphan based protocol has been equally well tolerated by all age groups, all lucarelli classes of children with thalassaemia major and different donor sources. the transplant related mortality and graft rejection rates have been low at . % and . %, respectively. however, children less than years need to be monitored carefully during the first months of transplantation as early withdrawal of immunosuppression can prevent graft rejection resulting in excellent outcomes. disclosure of conflict of interest: none. institute of cellular medicine, newcastle university, newcastle-upon-tyne, uk hemophagocytic lymphohistiocytosis (hlh; hemophagocytic syndrome) is a rare syndrome of potentially fatal, uncontrolled hyperinflammation. allogeneic stem cell transplantation (allosct) is indicated in familial, recurrent or progressive hlh. additional recommendations include central nervous system involvement and unknown triggering factor. while data for allosct outcome are available for the pediatric setting, information for adults is very limited. the aim of this study was to retrospectively analyze the information from the ebmt databases about adult hlh patients who underwent allogeneic stem cell transplantation. we obtained data of adult (≥ years of age) patients transplanted due to hlh. additionally, an hlh-oriented questionnaire was sent to the clinical centers, with responses received so far. median age at transplantation was (range: - ). there was a slight male predominance / ( %). the majority of patients were reported with secondary hlh / ( %), the familial disease was reported in / ( %) patients. in two patients triggering factor was attributed to malignancy. the majority of patients received stem cells obtained from the peripheral blood ( / ; %) while for the remaining ones it was bone marrow. reduced intensity conditioning was used since in / ( %) of patients. thirteen ( %) patients received tbi. donor choice was: matched unrelated ( %), mismatched unrelated ( %), identical sibling ( %). engraftment was observed in / ( %). the cumulative incidence of acute graft versus host disease (gvhd) at days was % ( % ci - %). the cumulative incidence of chronic gvhd at year after allosct was % ( % ci - %) and increased to % at years ( % ci - %). the -year probability of overall survival is shown in fig. . the median survival time was months. the -year os was % ( % ci - %). for patients who survived until months, this proportion was more favorable with an os of % ( % ci - %) at years after transplantation. among patients with observation times longer than months, only one patient died (in the th month after allosct due to relapse which occurred in the th month. after months no more relapses of hlh were recorded-the cumulative incidence reached %. the non-relapse mortality reached % after months. the familial disease was associated with a better prognosis than secondary hlh (p = . ). unlike the pediatric population, where reduced intensity conditioning (ric) was associated with higher survival, in adult patients there was no difference between the conditioning types. data form the questionnaires confirm clinical picture typical for hlh at the diagnosis: fever in / ( %), splenomegaly in / ( %), hemophagocytosis in / ( %) and hyperferritinemia with median concentration of ng/ml (range: - ). image fig. overall survival after allogeneic stem cell transplantation for adult hlh patients until months ( % confidence intervals are shown in grey). the number of patients at risk is indicated below the time axis at the corresponding time points. to our knowledge, this is the largest group of adult patients with hlh who underwent allogeneic stem cell transplantation. relatively low relapse incidence shows that allosct can effectively cure hlh. patients who survive the first period after this procedure can expect a long disease-free survival. disclosure of conflict of interest: none. allogeneic hematopoietic stem cell transplantation (hsct) is the only curative option for children suffering from various life-threatening inherited non-malignant diseases with best results using hla-identical family donor. hsct from unrelated or mismatched family donors is associated with increased risk of agvhd and graft rejection.use of post-transplantation cyclophosphamide (ptcy) with or without additional immunosuppression has been shown to be effective prophylaxis against gvhd in patients with hematological malignancies. there are limited reports of hsct using pt cy for patients with non-malignant disorders. we retrospectively analyzed results of hsct in patients with life-threatening non-malignant diseases using ptcy-based gvhd prophylaxis. patients characteristics are presented in table . thirteen patients ( . %) were transplanted upfront, patients ( , %) were rescued after primary or secondary graft failure after first hsct. donors were hla-matched (n = ) or mismatched ( - / ) (n = ) unrelated, haploidentical (n = ) or hla-identical family (n = ). bone marrow was used as graft source in ( . %) patients and peripheral blood stem cell in ( . %). median cd +/kg recipient weight- . × ( . - . ), cd +/kg- . × ( . - . ). the conditioning regimen was myeloablative in patients (conventional- , reduced toxicity- ), reduced intensity- . the gvhd prophylaxis consisted of a combination of ptcy at dose of mg/kg on days + and + with calcineurin inhibitors (tacrolimus- pts, cyclosporine a- pts) or sirolimus ( pt) and mmf ( pts) starting on day + . all but one patients received also serotherapy with rabbit ( pts) or horse atg ( pst) and rituximab ( pts). with a median follow-up of months (range: - ), the kaplan-meier estimates of os − . %. one patient with thalassemia died before engraftment on day+ from severe vod. / pts ( %) achieved engraftment. the median time for neutrophil and platelet engraftment was ( - ) and days ( - ), respectively. primary graft failure was observed in patients ( was successfully retransplanted from another haploidenticle donor, was not eligible for a second transplantation, but alive). at last follow up, ( %) patients had full donor chimerism, ( %) had stable mixed chimerism without signs of disease progression. one patien with wiscott-aldrich syndrome had secondary graft failure with progressive loss of donor chimerism and were successfully rescued with second haploidentical transplant from the same donor. of engrafted patients, agvhd ii-iv was seen in ( . %) patients. one patient developed grade ii (gut stage ii) agvhd, which resolved with systemic steroids. severe (griii-iv) agvhd was observed in pts after second hsct, both had calcineurin and mtor-inhibitors induced toxicity leading to discontinuation of this drugs, but responded on combined (steroids and ruxolitinib) therapy. one patient with was developed grade iii gvhd (gut stage ) after severe cmv-colitis and died on day from multiple organ failure (suspected tma). one patient developed extensive chronic gvhd of kidney (minimal change [p ] disease) after tapering of immunosupression. one patient with hurler syndrome had seizures, died on day+ from multiple organ dysfunction syndrome. conclusion: ptcy is a promising option for agvhd prophylaxis in patient with non-malignant disease, lacking an hla-matched family donor. disclosure of conflict of interest: none. an exploratory, open-label study to evaluate the safety and feasibility of atir , a t-lymphocyte enriched leukocyte preparation depleted ex vivo of host alloreactive t-cells (using photodynamic treatment), as adjuvant treatment to a t-cell depleted haploidentical hematopoietic stem cell transplantation in patients with beta-thalassemia major c selim , w rob , l sarah , f josu de la previous studies demonstrated that donor lymphocytes, selectively depleted of alloreactive t-cells (atir), could be given safely in adult patients receiving a haploidentical hsct. in patients a single dose of atir, at doses up to × viable t-cells/kg, was given and no grade iii/iv acute gvhd has been reported. this confirms the efficacy of the elimination method of allo-reactive t-cells and attributes to its beneficial safety profile. in an ongoing phase study, cr-air- (nct ), infusion of atir at days post-hsct results in a reduction of transplant-related mortality (trm) and improvement of overall survival and event-free survival. adjunctive treatment with donor lymphocytes in patients receiving a t-cell depleted, haploidentical hsct for nonmalignant diseases such as beta thalassemia major, could provide early immunological support and better immune reconstitution in the absence of gvhd. in an open-label, multicenter phase study (cr-bd- ; eudract - - ), patients age ≥ years and ≤ years with beta thalassemia major will undergo a haploidentical hsct with adjunctive administration of atir . patients will receive a t-cell depleted graft (cd -selected, or cd /cd depleted, or tcr-αβ depleted, depending on the experience of the study center) from a related, haploidentical donor, patient conditioning will be myeloablative following standard practices at the study center. atir infusion at a dose of × viable t-cells/kg is given between and days after the hsct. to assess safety, patients will be evaluated for the occurrence of dose limiting toxicity (dlt), defined as acute gvhd grade iii/iv within days post hsct. efficacy will be primarily evaluated by transfusion-free survival (tfs), occurrence of severe infections, and time to t-cell reconstitution, taking into account hematologic and sustained engraftment. all patients will be closely monitored for cmv, ebv and adenovirus titers, with initiation of pre-emptive treatment upon rising blood titers. regulatory authorities in the united kingdom and germany have approved this clinical study protocol. enrolment of the study is expected to continue during , with first report of safety of atir to be expected first half . disclosure of conflict of interest: j. rovers is employee of kiadis pharma, sponsor of the study. sickle cell disease (scd) can be cured with haematopoietic cell transplantation (hct), yet progress in the practice and research of hct for scd has not come without risks and uncertainty. the information and decisions that families and physicians encounter in this field are complex and hanging. in this hermeneutic study, we analyze the case of one family who advocated for hcts for two of their four children knowing the potential risks. these experiences have had a profound impact on both the family and the medical team. this study was conducted through the research method of hermeneutic phenomenology. hermeneutic inquiry is described as the practice and theory of interpretation and understanding in human contexts and aims to make sense of the particulars of these contexts and arrive at deeper understandings. data collection: in-depth interviews were conducted with the mother of the family, the hct nurse coordinator, and the hct physician. the interviews were audiotaped and transcribed verbatim. the transcribed interviews were later reviewed by the physician, who then wrote an additional reflection. this work culminated in approximately pages of single spaced data in textual form. in hermeneutics, interpretation takes place through a careful reading and re-reading of the data, looking for statements and instances that resonate with the researcher. initial individual interpretations of researchers are then raised to another level of interpretive analysis in the research team's communal attention to the data. particular criteria guide the analysis: agreement, coherence, comprehensiveness, potential, and penetration. the following excerpts and interpretations are provided as examples of the analysis, with names changed for confidentiality. "being heard" arose repeatedly in this family's experience, including at the time of their request for a transplant without meeting the traditional criteria for hct. they persisted in their belief that their children would benefit from hct. "they gave us options to see if there was a chance for a transplant...how life would look…. and then we figured…a transplant for him was better at the time…worth the risks…. and you wouldn't even know. he plays basketball now, he plays sports, he's active and he can exercise and run. i never had any regrets because i felt it was better and the most important thing is his organs were really intact; none of the organs were destroyed…so i think it's the right decision we made" (mother). "this family has changed my career, and my life as a result. they challenged my practice and way of thinking. they did so in a considerate way, out of a duty to advocate for their children. we worked through the tension of different viewpoints with respect and all of us grew in the process. at least i can say our team did. i certainly did... i am humbled by their trust and respect…i am grateful to them" (physician). patients and providers are deeply impacted by their interactions. dr. robert buckman stated that it was the individual case that changed his practice always. he claimed he could not walk into a new patient's room without his practice being forever changed. in presenting this hermaneutic analysis, we aim to remind ourselves of the opportunity for growth that can result from reflection on this sacred patientprovider relationship. disclosure of conflict of interest: none. defibrotide (df) prophylaxis and adjustment of busulfan schedule to prevent veno-occlusive disease and thrombotic microangiopathy in an infant with a membrane cofactor protein (mcp) gene mutation and metachromatic leukodystrophy undergoing hematopoietic stem cell gene therapy (hsc-gt) v calbi , , f fumagalli , , , r penati , g consiglieri , m migliavacca , , d redaelli , s acquati , v attanasio , r chiesa , f ferrua , , f barzaghi , , m cicalese , , a assanelli , , p silvani , s tedeschi, r arora , a soman , f ciotti , m sarzana , g antonioli , , c baldoli , s martino , gl ardissino , mg natali sora , l naldini , , f ciceri , , a aiuti , , and me bernardo hepatic veno-occlusive disease (vod) and thrombotic microangiopathy (tma) are life-threatening complications that can occur after hsc transplantation. expert consensus guidelines support use of df for treatment and prophylaxis of vod due to its ability to restore thrombo-fibrinolytic balance and protect endothelial cells. presymptomatic monozygous twins affected by late infantile metachromatic leukodystrophy (mld) underwent investigational hsc-gt after conditioning with busulfan. no comorbidities were evident at baseline. the dose of transduced cd + cells was similar in both patients ( . × cd +/kg for patient and . × cd +/kg for patient ). patient (p ): at months of age, received conditioning with iv busulfan mg/m /dose for doses (target auc mg × h/l). on day (d) + after gt, he developed severe vod and was treated with diuretics, fresh frozen plasma, paracentesis and df. on d+ schistocytes in peripheral blood, marked proteinuria, complement factor consumption, and increases ldh and bilirubin were observed. the patient's condition worsened, with reduced urine output and generalized oedema with pleural effusion. stool, urine and blood cultures were negative and adamts activity was %; anti-complement factor h (cfh) antibodies (ab) were positive ( ui/ml). these findings led to the diagnosis of atypical hemolytic uremic syndrome (ahus; a form of tma) and eculizumab ( mg/weekly dose) was started on d + . patient subsequently developed pulmonary oedema and needed non-invasive ventilation. molecular analysis revealed a heterozygous deletion of cfhr -r and ala val mutation in the mcp gene, a defect previously shown to be associated with ahus. due to the presence of ab anti-cfh and antiplatelet, weekly doses of rituximab ( mg/m ) were administered. after treatment, p progressively improved although he showed prolonged severe anaemia and thrombocytopenia and bone marrow (bm) hypoplasia, secondary bleeding which required reinfusion of unmanipulated autologous bm cells on d + . nine months after hsct-gt p has shown good hematopoietic recovery, stable engraftment of the transduced hscs, no signs of renal damage or complement activation, albeit with neurodevelopmental delay. patient (p ): given his twin history and genetics, this month old infant was considered at increased risk of vod, so prophylaxis with df ( mg/kg/d) was administered from d- to d+ and the busulfan conditioning was modified by adjusting the auc to a lower target ( mg/kg/dose for doses; target auc . mg × h/l). the child had a good clinical recovery and didn't develop signs of vod or tma after hsc-gt. on d+ and + , respectively, anti-cfh and anti-platelet ab were positive. considering the history of the twin, weekly doses of rituximab were administered. p is currently months after gt with persistent engraftment of transduced hscs and no signs of tma. data from this case-control report of monozygous twins diagnosed with mld, and subsequently shown to also harbor mutations in complement regulator gene, suggest that df prophylaxis and busulfan adjustment may have helped prevent systemic microangiopathic damage in the second twin. patients with rare disease may have mutations in genes in addition to those that cause their disease. patients at risk of post-transplant tma following hsc-gt for genetic diseases may require tailored df prophylaxis and treatment. disclosure of conflict of interest: a. aiuti is the principal investigator of the tiget-mld clinical trial of gene therapy. the mld gene therapy was licensed to glaxosmithkline (gsk) in and gsk became the financial sponsor of the trial. all authors declare no other competing interests. hematopoietic stem cell transplantation (hsct) using an optimized conditioning regimen is essential for the longterm survival of patients with inherited bone marrow failure syndromes (ibmfs). we report hsct in children with fanconi anemia (fa, n = ), diamond-blackfan anemia (dba, n = ), dyskeratosis congenita (dc, n = ) and shwachman-diamond syndrome (sds, n = ) from a single hsct center. the graft source was peripheral blood stem cells (n = ) or cord blood stem cells (n = ). fa, dc and sds patients received reduced-intensity conditioning, while dba patients had myeloablative conditioning. the median numbers of infused mononuclear cells and cd + cells were . × /kg and . × /kg, respectively. the median time for neutrophil and platelet recovery was and days, respectively. there was one primary graft failure. after median follow up years the overall survival was %. the incidence of grade ii-iii acute and chronic graft versus host disease (gvhd) was % and % respectively. in a multivariate analysis, the type of conditioning regimen was the only factor identified as significantly associated with grade ii-iii acute gvhd (p = . ). we conclude that hsct can be a curative option for patients with ibmfs. disease specific conditioning regimen was important to disease the transplant-related mortality. [p ] disclosure of conflict of interest: none. homozygous sickle cell disease (scd) patients suffering from end-stage renal disease (esrd) show a variable outcome after kidney transplantation as underlying disease can cause poor allograft survival and disease-specific problems. we present a case of a -year old patient with severe scd and esrd who underwent haploidentical bone marrow transplantation (bmt) with consecutive living kidney transplantion (lkt). the patient suffered from multiple complications of scd including stroke with secondary hemorrhage, symptomatic epilepsy, esrd and uncontrolled hypertension. the patient had been on hydroxyurea without success and required regular blood transfusion due to severe renal anemia. the rationale for bmt was uncontrollable iron overload. a reduced intensity conditioning regimen was used with (fludarabine, cyclophosphamide and gy of tbi, dose-adjusted to esrd). graft-versus-host disease (gvhd) prophylaxis consisted of post-transplant high-dose cyclophosphamide, cyclosporine a (cya) and mycophenolate mofetil (mmf). the donor was her -year old mother with hbs trait, the stem cell source was bone marrow, the cell dose . × nucleated cells/kg. during conditioning daily hemodialysis was performed to keep drug levels stable. neutrophil engraftment occurred on day + , chimerism at day + was %. hbs increased from . % pre-hsct to . % months after hsct. hemoglobin values increased from g/l pre-hsct to g/l post-hsct and reticulocytes from g/l to g/l. erythropoietin levels increased from . iu/l pre-hsct to iu/l months after hsct. during the follow-up, the patient did not show any sign of acute gvhd or vaso-occlusive crisis, hemolysis or sickling. relevant complications were disease-related (therapy resistant hypertension and epileptic seizure due to former brain damage). on day + a lkt from the same donor was performed. the initial immunosuppressive treatment with mmf was continued, cya was switched to tacrolimus and steroids were added for months. the post-transplant period was uneventful. currently, months after haploidentical bmt and months after lkt there are no signs of gvhd, the blood chimerism is %, the kidney allograft function is very good (gfr ml/min/ . m ) and immunosuppression is withdrawn. iron overload is being corrected by regular phlebotomies. the patient no longer requires antihypertensive medication and there is evidence of vascular remodeling. this is the first report of a successful haploidentical bmt followed by kidney transplantation from the same donor in a patient with scd. disclosure of conflict of interest: none. allogeneic hematopoietic stem cell transplantation (hsct) can cure non-malignant diseases, such as primary immune deficiency (pid), severe aplastic anemia (saa) and osteopetrosis (op). in the absence of a well-matched donor, transplantation from a haplo-identical donor maybe considered. post-transplant cyclophosphamide (ptcy) is a new strategy derived from the treatment of malignant diseases in adults that has been little studied in high-risk pediatric nonmalignant diseases. fifteen children ( . years, range: . - . ) underwent hsct in the pediatric immunology and hematology unit of necker hospital, paris, between december and september . these children were suffering from op (n = ), saa (n = ), hemophagocytic lymphohistiocytosis (hlh) (n = ), immunodysregulation polyendocrinopathy enteropathy x-linked (ipex) syndrome (n = ), combined immune deficiency (n = ) and leukocyte adhesion molecule deficiency (n = ). three patients received a low-intensity conditioning regimen (cr) (based on fludarabine, cyclophosphamide, and total body irradiation) whereas the other received myeloablative cr (based on busulfan auc targeted and fludarabine). fourteen patients received serotherapy before hsct. post-transplant cyclophosphamide ( mg/kg/ day) was given on d and d and graft versus host disease (gvhd) prophylaxis with cyclosporine and mycophenolate mofetil was initiated on d . the transplanted stem cells were obtained from bone marrow in all cases. engraftment with full donor chimerism was observed in patients. the median cd + cell dose was . × cells/kg body weight (range: . - . × ). neutrophils recovered after a median of days (range: - ), and overall survival (os) was % after a median follow-up of year (range: . - . ). three patients died due to graft failure (n = ) or infectious complications related to gvhd (n = ). grade ii acute gvhd occurred in of the patients displaying engraftment ( %), and chronic gvhd and/or autoimmune complications were observed in four patients ( %). viral complications were frequent, occurring in patients ( %) with cmv infection (n = ) /disease (n = ), adenovirus disease (n = ) and bk virus cystitis (n = ). haploidentical transplant with ptcy is feasible in high-risk patients with non-malignant diseases. chronic gvhd and autoimmunity were more frequent than for more conventional approaches in such patients. infection rates were high. disclosure of conflict of interest: none. sickle cell disease (scd) remains associated with high risks of morbidity and early death. even best of supportive care fails to improve quality of life. hematopoietic stem cell transplant (hsct) can be considered for selected group of patients. in long run it is not just economical but also substantially improves quality of life (qol). we report our experience with hsct for scd from india. seventy three consecutive patients suffering from scd who underwent hsct between january and november were included in the study. fifty two underwent matched sibling donor (msd), matched family donor (mfd), matched unrelated ( / or / ), cord blood transplant cbt ( matched sibling cord blood and matched unrelated) and patient underwent haploidentical transplant. different conditioning regimens were used and so was the graft versus host disease prophylaxis depending on institutional protocols as depicted in table . a total of patients underwent sct. the median age was years ( months- years). m/f ratio was / . majority of patients were either from african union or oman. all patients suffered from one or other severe symptoms making them eligible for sct. graft source was bone marrow (bm) in with median cd count of . x /kg ( . - . ), peripheral blood (pb) in with median cd count of . x /kg ( . - . ), cord blood in with median cd count of . x /kg ( . - . ) and combined bm & pb in with median cd count of . x /kg ( . - . ). of the patients, are alive and disease free with lansky/karnofsky scores of . there were deaths ( msd/mfd/mud; haploidentical and matched unrelated cbt). four patients rejected the graft ( haploidentical and msd/mfd/mud). at the last follow up, the probabilities of survival, scd-free survival, and transplantrelated mortality were %, . %, and %, respectively. outcome of hsct in scd has improved significantly. with better conditioning regimens, improved supportive care, the outcome of alternative donor transplant and adult scd has improved and matches sibling donor transplant. hsct should be strongly considered as a curative modality for selected patients suffering from scd. disclosure of conflict of interest: none. s staff jointly defined more than local standard operating procedures. patients with low-risk characteristics (age ≤ years, liver size ≤ cm below costal margin) and a hla matched sibling donor were considered eligible in this initial phase of activity. a downstaging protocol with hydroxyurea and deferoxamine or deferasirox was adopted. conditioning regimen included iv busulfan and cyclophosphamide. gvhd and rejection prophylaxis included atg from day − to − and csa, mtx and methylprednisolone. gcs-f primed bone marrow was chosen as stem cell source. the first allogeneic hsct of the whole iraq was performed in a child with thalassemia at hiwa hospital in october . up to now, patients ( females, male) underwent hsct; median age at transplantation was years; median infused tnc . × /kg, cd + . × /kg. all of them engrafted. no major complication were observed. one of them developed grade ii agvhd (skin only) which resolved after increasing the dose of steroids. a huge number of patients with low-risk thalassemia are now in the waiting list and some of them have already started downstaging having planned hsct in a short time. a matched sibling transplant program in children with thalassemia is feasible and safe in kurdistan. such a program can provide many advantages: far less psychosocial and financial burden for the families and significant saving for the government. the estimated costs of performing locally hsct are much less than in the countries where patients were previously referred. the continuation of cooperation is of paramount relevance for further implementing the activity and extending the transplant accessibility to patients with other hemato-oncological disorders of childhood. disclosure of conflict of interest: none. long term follow-up after reduced-intensity conditioning and stem cell transplantation for thalassemia major r rihani, a natsheh, sm abu, e khattab, r najjar, f sheab, s sharma, n hussein, a tbakhi and m sarhan bone marrow and stem transplantation program-king hussein cancer center, amman, jordan hematopoietic stem cell transplantation (hsct) is the only curative treatment for thalassemia major (tm). reducedintensity conditioning (ric) before hsct for high risk tm patients results in fewer complications, when compared with myeloablative regimens. one hundred and three tm patients received hscts from an hla-identical related donor at king hussein cancer center, between january and november . of those, were high risk tm ( %) who received ric hscts. in this report, we describe follow-up beyond years (median, ; . - months) post ric hscts. forty-four class ii-iii patients ( %) were identified ( % with hepatitis c); with a median age of ( . - ) years. females accounted for % (n = ). conditioning regimen consisted of oral busulfan mg/kg, fludarabine mg/m ,tli cgy and atg followed by pbsct. gvhd prophylaxis consisted of mmf and csa. median infused stem cell dose was . × /kg. all patients attained neutrophil and platelet engraftment (median, . and . days, respectively). persistent mixed donor or full donor chimerism were observed in . % (n = ) and . % (n = ), respectively. immune-suppressive therapy for gvhd treatment was required in ( . %) patients (agvhd, n = ; cgvhd, n = ). moreover, veno-occlusive disease occurred in patients ( %) that resolved completely. secondary graft failure was noted in ( %) patients. the -year overall survival was %, while the -year probability of thalassemia-free survival was . %. other factors evaluated include: growth parameters, endocrine and other organ functions, in addition to functional status. this report confirms the safety and efficacy of ric regimens in hscts for high risk tm patients. those regimens are associated with excellent engraftment and sustained mixed donor chimerism; and lead to excellent thalassemia-free and overall survival rates. [p ] disclosure of conflict of interest: none. in-time hsct for pts. with hurler syndrome (hs) can significantly improve the results. long-term follow-up and late effects estimation required to prepare a special observation and rehabilitation programs. aim. to analyze our experience with hsct for hs in the field of special observation and rehabilitation programs. forty hsct during the - were performed for pts. with hs. median age at the diagnosis was months ( - months), at hsct- months ( - months). bm used in . % (n = ), pbsc- . % (n = ), cb- . % (n = ). mac conditioning was used for hsct, ric-for . ric regimen: flu+mel+atg, mac: bu/treo +flu+thio/mel (bu was used in early ) and atg +rituximab (in case of mud hsct). all pts. with ric received mud hsct, pts. with mac mud- pts., mrd- pts. pts. received csa/tacro-based gvhd prophylaxis. mmf/mtx was additionally added in all cases. in ric hsct immunomagnetic Сd /Сd + depletion of pbsc (by clinimacs) was used. a special observation protocol including somatic and neurocognitive estimation was developed. all pts. engrafted with full donor chimerism on d+ . median of engraftment day- ( - days). thirty three pts. survived. reasons of death-mac: infections- pts., ric: trali- pt., agvhd- pt. trm improved, over the years, with improving of supportive care and donor selection as well as pre-transplant screening. no early severe toxicity revealed. pulmonary infection episodes was registered in % of pts. in our study. gvhd: grade iideveloped pts., grade iii-iv- pts. (after ric), local cgvhd- pts. (ric). no extensive cgvhd. pts. rejected (mac and ric rejection rate was same). at median follow up of months ( - months), the estimated years pos was %. best response correlated with early hsct (and better status before hsct) and higher level of aidu after. late effects estimation showed that . % (n = ) of patients experienced late effects: cardio-vascular- pts., skeletal- pts., endocrine- pts. all pts. with cardio-vascular effects received mac. skeletal effects affected patients of older age, pts. transplanted in younger age do not have such effects. median period of late effects arising after hsct was month ( - months). only pts. experienced serious pulmonary late effects (infections), all episodes was before . no pts. in our study have progressive retinal degeneration. % of pts. improved in the neurosensory component and all pts. improved in neurocognitive status and development after hsct. best response correlated with neurocognitive rehabilitation based on unique computer model used by our group in russian national rehabilitation center "russkoe pole." in-time hsct is an effective and safe way to stop neurodegenerative process for pts. with hs. both mac and ric regimens can be used with the same effectiveness. mac regimens associated with bigger number of cardiovascular late effects. long-time follow-up showed that these patients require the special observational protocol including estimation of cardio-vascular, skeletal, endocrine and neurocognitive risks. better neurocognitive response correlated with intensive rehabilitation using computer model. russian joint study showed effective cooperation for treatment pts. with hs in the national setting. disclosure of conflict of interest: none. little is known about pathogenesis of solid tumors after hsct but, intensive cytotoxic conditioning therapy with defective dna repair of persisting stem cells/stromal cells, viral infection, and immunosuppression may play a role. / patients with solid tumors had a melphalan-based conditioning. melphalan was linked to sarcoma and lung cancer in animal model. there are few data linking parotid mec to infection by cmv and hhv which can remain dormant in the salivary glands. both affected patients had hhv during the transplant period. p and p had a family history of solid tumor pointing to a possible genetic factor. whilst secondary malignancy post-hsct for patients with malignant disorders is well recognised, non-ptld malignancy post-hsct for pid has not previously been reported. a larger study is needed to evaluate incidence and risks. allogeneic hsct is a treatment of choice for the bone marrow failure in patients with sds. hsct from unrelated or mismatched family donors is associated with higher morbidity and mortality compared with matched sibling. combined pgd and hla antigen testing is a possible option to preselect a compatible donor for an affected sibling requiring hsct. we describe a case report demonstrating first successful hsct for years girl with sds by using preimplantation genetic diagnosis and hla matching. diagnosis of sds was suspected at m.o., based on clinical features, family history, laboratory studies. at m.o., bone marrow (bm) aspiration revealed hypocellular marrow with signs of dysplasia and expansion of blasts ( . % blasts). the sanger sequencing of sbds gene showed c. - ta ct and c. + t c mutations. the patient had recurrent infections, including bilateral pneumonia caused by phaeohyphomyces, bloodstream infection, cmvdisease. due to the lack of matched related or unrelated donors, hsct with ric (flu, mel, atg) from haploidentical father was performed at months of age. after the st allo-hsct, engraftment was achieved on d+ , initial str study showed full donor chimerism. post-transplant period was complicated with severe cmv-infection and signs of secondary hlh. at d+ , graft rejection was registered. the girl became dependent on regular rbc and platelet transfusions, bm examination revealed hypocellularity with moderate signs of myelodysplasia without elevated blast count. due to lack of available hla-compatible donors, an option of in vitro fertilization (ivf) with preimplantation selection of a normal hla-matched embryo was considered. after controlled ovarian hyperstimulation embryos were hla-compatible and healthy (first, wild-type; second, heterozygous for sbds gene mutation c. - ta ct). hence, the only unaffected hla-identical embryo was transferred resulting into full-term pregnancy. at the age of . years after st hsct, the nd s transplant was performed with a combination of cb and bm as a source of hematopoietic stem cells. the donors' age was years a reduced toxicity conditioning regimen (rtc) based on flu mg/m , treo g/m , thiotepa mg/kg with serotherapy (thymoglobuline . mg/kg) was used. because of neurotoxicity, arterial hypertension, since d+ csa was changed to sirolimus +mmf for gvhd prophylaxis. the total number of infused nc was . × /kg; cd +, . × /kg; cd +, . × /kg. engraftment was achieved on d+ . any signs of gvhd, severe infectious or toxic complications were not observed. eight months later, the patient is alive, has full donor chimerism in bm and is not transfusion-dependent. in the absence of hla-identical donor, ivf with preliminary pgd and hla-typing could be a chance for matched donor to cure patients with non-malignant genetic diseases. in case of low cord blood cellularity, a combination of cb and bm from the same sibling could be used. our experience showed a successful engraftment of sds patient and stable donor chimerism after second hsct of cb and bm from pgd-selected sibling with rtc. disclosure of conflict of interest: none. the safety and efficacy of familial haploidentical (fhi) stem cell transplantation utilizing cd enrichment and cd addback in patients with high risk sickle cell disease (scd) ( figure a ). probability of yr efs is . % (ci : - %) ( figure b ). immune cell reconstitution has been robust and similar to rtc and msd allosct in scd (table ). there have been deaths, vod, steroid refractory agvhd and cgvhd. mac followed by fhi utilizing cd enrichment and t-cell addback in patients with high-risk scd is safe, tolerable and results in long-term donor chimerism and absence of scd symptoms or complications. a larger cohort and follow-up will be required to confirm these preliminary findings. disclosure of conflict of interest: none. supported by r fd - a . [p ] s lymphoma p a clinical prognostic index for assessing patients aged being considered for high-dose therapy and autologous stem-cell transplant in relapsed or refractory high-grade non-hodgkin lymphoma d edwards , k kirkland , r pearce, s robinson and g cook bsbmt patients with relapsed high-grade nhl or disease refractory to first-line therapy can still be cured with high-dose therapy and autologous stem cell transplant if they respond to salvage chemotherapy. this aggressive algorithm is accepted in younger patients but is less well established in the elderly. age has a negative predictive score in the international prognostic index (ipi) and there are concerns that the outcomes of hdt in these patients are significantly worse. deciding which older patients will benefit from hdt is challenging and there are no established predictive tools to guide physicians. we present a clinical prognostic index derived from information readily available at the time a patient is being assessed for asct the bsbmt audited the outcomes for uk patients aged transplanted between - (n = ) and benchmarked against the european bone marrow transplant (ebmt) database for the same period (n = ). the primary outcome was progression-free survival (pfs) but data was also analysed for overall survival (os), relapse rate (rr) and non-relapse mortality (nrm). we included all patients with a diagnosis of high grade nhl and the following demographic features were also analysed: age at diagnosis; age at transplant; m/f; year of transplant; cr/not cr at transplant; no. of prior therapies; no. of cells infused; clinical staging; karnofsky status at transplant; histology; ipi at diagnosis; mobilising regime and conditioning regime. candidate prognostic indices were factors achieving significance in univariate and multivariate analyses of the main outcomes by regression analysis. the best prognostic index was selected based on the bsbmt dataset and then applied to the rest of the ebmt dataset (the validation dataset). there were no significant differences in patient characteristics between the uk and non-uk groups nor in outcomes of pfs, os, rr or nrm. (figure ). in both univariate and multivariate analysis the following features were associated with a significantly worse outcome for pfs, os, rr and nrm : age , karnofsky score. disclosure of conflict of interest: none. underwent an allo-sct at our center after a treosulfan-based conditioning regimen. eleven pts received a mrd, pts a mud, and pts a haplo unmanipulated pbsc allo-sct. at allo-sct pts were in cr, pts were in pr, and pts had sd/pd. hct-ci was evaluable for pts, had a score ≥ . the backbone conditioning regimen consisted of treosulfan g/m from day − to − , and fludarabine mg/m from day − to − ; twenty-five pts were treated with this reduced toxicity conditioning (rtc) regimen. intensification with other alkylating agent (melphalan, thiotepa, or cyclophosphamide) or radiotherapy ( gy total dose) was applied on the remaining pts (myeloablative conditioning, mac). gvhd prophylaxis was based on cyclosporine a and methotrexate ( pts) or rapamycin and mycophenolate mofetil ( pts), plus anti-thymocyte globulin or post-transplant cyclophosphamide accordingly to donor type. median numbers of infused cd +/kg and cd +/kg were . × (range: . - . ) and . × (range: . - . ), respectively. median follow-up was months (range: - ). thirty-nine pts were evaluable for engraftment; median time to neutrophil ≥ . × /l was days (range: - ), and days (range: - ) to platelet ≥ × /l. treosulfan conditioning provided a cr in and pts respectively in pr and sd/pd at transplant. no graft failure was observed. one and years overall survival (os) was . % and . %, respectively. progression free survival (pfs) and gvhd-free/relapse-free survival (grfs) were respectively . % and . % at year, . % and % at years. one and years relapse/progression incidence (ri) was . % and . %, respectively. transplant related mortality (trm) was . % at days, . % at year and for the entire follow-up. the -day cumulative incidence (ci) of agvhd grade ≥ was . %; ci of moderate to severe cgvhd was . % at years. the outcome of pts in cr at years was significantly better compared to that of pts with active disease in terms of both os ( . % vs . %, p o . ), pfs ( . % vs %, p o . ), grfs ( . % vs . %, p o . ), and ri ( . % vs %, p o . ). no statistical differences in os, pfs, and ri were found when pts were stratified according to donor type and [p ] the use of rtc or mac regimen. at last follow-up, patients are alive and disease free; of them obtained a durable cr using chemotherapy and/or dli for disease progression after allo-sct. treosulfan-based conditioning regimen is effective and well-tolerated in patients with advanced b-nhl undetgoing allo-sct. disclosure of conflict of interest: none. systemic anaplastic large cell lymphoma (salcl) is a very infrequent well-defined histological entity that comprises around % of all t-cell non-hodgkin lymphoma. in the absence of prospective clinical trials, autologous stem cell transplantation (autosct) is considered the standard of care as consolidation therapy after first line therapy for those patients not expressing the alk protein (alk neg salcl) and for patients with relapsed disease. the objective of this retrospective analysis was to analyse the long-term outcome of patients diagnosed with salcl and being treated with autosct during the course of the disease, making special emphasis on the potential impact of the administration of brentuximab vedotin (bv). eligible for this study were patients years or above with salcl who underwent autosct between to and were reported to the ebmt. baseline patient, disease, and transplant data were collected from ebmt med-a standard forms. centers with potentially eligible patients were contacted to provide additional treatment and follow-up information including a written histopathology report for central review. seventy-nine patients ( males) with a median age at diagnosis of years (range: - ) and at transplantation of years were included in the final analysis. thirty-nine patients were alk negative, alk positive and in patients expression of alk protein was unknown. at diagnosis, patients ( %) presented with advanced stage and ( %), with b symptoms. sixty-three patients ( %) received - lines of therapy before autosct. ten patients were treated with bv at some point before autosct; two patients as second line therapy, three as third line, one as fourth line and four as fifth line therapy. the median number of bv doses was (range: [ ] [ ] [ ] [ ] [ ] [ ] . the median time between diagnosis and transplantation was months (range: . most patients had chemosensitive disease at autosct [ patients ( %)] and in all but patients peripheral blood was used as the source of stem cells. conditioning regimen consisted on beam / beam-like protocols in patients ( %). all patients engrafted. with a median follow up for surviving patients of months (range: - ), patients are alive ( %), patients died ( %) and patients ( %) are lost for follow up. disease relapse after transplantation was the most frequent cause of death after the procedure. cumulative incidence of non-relapse mortality for the whole series was % ( % ci, . - ) at days, year and years. cumulative incidence of relapse was % ( % ci - ) and % ( %ci - ) at and years, respectively. and years progression free survival (pfs) was % ( % ci - ) and % ( % ci - ), respectively and and -years overall survival (os) was % ( % ci - ) and % ( % ci - ), respectively. there were no significant differences in any of the outcomes between bv treated and non-treated patients. autosct results in a promising pfs and os in patients with salcl. the potential impact of the administration of bv as salvage strategy before the procedure needs to be further elucidated. disclosure of conflict of interest: none. coeliac disease (cd) is a t-cell immune-mediated enteropathy to dietary gluten, characterized by small bowel villous atrophy resulting in malabsortion. the enteropathy is reversible with a gluten-free diet (gfd), however symptoms and signs which persist year are defined as refractory coeliac disease (rcd). rcd is divided into type i and ii, depending on absence/ presence respectively of clonal intra-epithelial t-lymphocytes (iels) with an aberrant phenotype (cytcd pos, membranous cd , cd and cd neg). rcdii patients have a year survival of . , plts ) was successful at a median of . (range: - ) days and no transplant-related mortality occurred. all patients achieved a clinical complete remission, with normalization of nutritional indices at days, but persistently abnormal iels and clonal t-cells on duodenal biopsy. with a median follow-up of . (range: - ) months, patients remain in clinical remission, patient relapsed with rcd and no patient progressed to eatl. chemotherapy and asct is a safe and effective strategy for the treatment of rcd offering the possibility of sustained clinical responses. clonal tcr in duodenal biopsy/blood and iel flow cytometry form part of the patient evaluation prior to the chemotherapy/asct program. most patients with hodgkin lymphoma (hl) are cured with conventional chemotherapy. however, approximately % of patients relapse after primary treatment. for those, high-dose chemotherapy (hdc) followed by autologous stem cell transplantation (asct) is the standard of care. fifty seven adult patients with relapsed or refractory hl submitted to asct between and were reviewed. variables examined were sex, age, ann arbor stage (i-ii vs iii-iv), b symptoms, bulky disease, extranodal involvement, nodal areas involved (≥ vs vs ≤ months) and response to the treatment prior to asct. log-rank test was used to compare differences in survival for each factor. patients median age was ( - ) years at diagnosis. ann arbor stage iii-iv in ( %) patients, b symptoms in ( %), extranodal involvement in ( %) and bulky disease in ( %). all patients were treated according to the abvd protocol in first line. indications for asct were relapsed disease (n = , . %) and lack of complete response (cr) or progressive disease with st line treatment (n = , . %). there were a median of ( - ) treatment-lines before asct (protocols eshap, ice, beacoop, gvd and others). the disease was chemosensitive in % cases: cr in and partial response (pr) in patients prior to asct. refractory disease (rd) in % (n = ). in . % patients, the hematopoietic cells mobilization was performed under stimulation with granulocyte-colony stimulating factor in hematologic recovery after the cycle of nd line chemotherapy, and most of which required ( - ) apheresis. conditioning regimens were beam ( %) and gmb ( %). the median time to hematologic recovery was days ( - ) for neutrophils /ul and days ( - ) for platelets , / ul. three months after asct, thirty-nine ( . %) patients had cr, one ( . %) patient maintained pr and ( . %) patients had disease progression. status unknown in patients and four ( %) patients died. relapse rate % (n = / ). with a median follow-up time after asct of ( - ) months, median disease-free survival (dfs) was ( - ) months and overall survival (os) was ( - ) months. there were deaths ( . %), four ( %) related to early infectious complications of asct, two ( . %) due to late infectious complications, eleven ( . %) due to disease progression and ( . %) in context of secondary acute myeloid leukemia. response to the treatment prior to asct was the only factor with survival influence. the dfs and os differed significantly in chemosensitive disease compared with rd (dfs mean: vs months,p = . , os mean: vs months, p = . ). the response to salvage treatment prior to asct is the main prognostic factor for survival after asct. prognosis remains poor in patients with rd or early and disseminated relapses. for these patients, the therapeutic approach should include intensive treatment with tandem hdc and stem cell transplantation, allogeneic transplant or early consolidation with brentuximab-vedotin after asct. hodgkin's lymphoma (hl), although considered a curable neoplasm in adults, could be associated with a very poor prognosis when refractory to primary induction therapy or when it relapses within months from an autologous stem cells transplant (auto-sct). the optimal treatment of patients with heavily pretreated/refractory hl is controversial. brentuximab vedotin (bv) is an active single agent in this context; unfortunately, there are no well established therapies when patients fail to respond or progress after bv. encouraging results were recently described with checkpoint inhibitors. similarly, data pertaining to efficacy of bendamustine (benda) shows encouraging activity in various refractory lymphomas. we included in this study adult patients with hl who relapsed post auto-sct and were refractory to or progressed after salvage bv and were treated with benda as salvage therapy with an intention to proceed with an allo-sct. this study was [p ] conducted in two major centers in lebanon, the american university of beirut medical center (aubmc) and makassed university hospital. we identified eligible cases. the primary study endpoint was objective response rate (orr). the secondary endpoint evaluated successful rate of bridging into an allo-sct. the median follow-up times from auto-sct and from benda salvage were ( - ) and ( - ) months, respectively. the median age of patients was years ( - ). all patients had bv as salvage therapy post auto-sct, and all of them progressed after a median of ( - ) cycles. clinical characteristics are outlined in table . patients received a median of cycles ( - ) of benda. the treatment was well tolerated, with rather infrequent adverse events and transient and manageable toxicities. the orr was %, in of patients, with % obtaining a complete response. eventually, of proceeded to allo-sct using a matched related donor, and the remaining patients are planned for allo-sct. only one patient died from disease progression after months post allo-sct. two of patients who progressed following benda received salvage therapy with nivolumab and are being planned for haplo-identical transplant while the third one is being planned for therapy with nivolumab. from the initiation of benda, the median duration of response for the patients was months ( - ); all these patients had maintained a continuous response at the last follow-up examination. conclusion: notwithstanding the limitations associated with our analysis, namely a small sample size and its retrospective nature, these results suggest a role for bendamustine in post bv failures. these findings also provide the basis to evaluate the concept of benda as a bridge to allo-sct in a large prospective study. [p ] disclosure of conflict of interest: none. brentuximab vedotin for relapsed or refractory hodgkin lymphoma, single center experience king faisal specialist hospital and research center, riyadh, kingdom of saudi arabia ms rauf , i maghfoor , a badran , mn zahir and s akhtar hodgkin lymphoma (hl) patients with relapsed or progressive disease after high dose chemotherapy (hdc) and auto-sct have limited curative options. fda granted approval of brentuximab vedotin (bv) for the treatment of hl and anaplastic large cell lymphoma (alcl) patients who fail auto-sct or have had at least prior multiagent chemotherapy regimens and are not candidates for auto-sct. we are reporting single center experience of bv usage in this "approved" setting. medical records were reviewed to collect required data. kaplan-meier (km) method was used to calculate overall survival (os) and progression free survival (pfs) from date of first dose of bv. from - , patients received bv. / had hl ( classic hl-nodular sclerosis, hl-mixed cellularity) and alcl. / ( %) pts were primary refractory or had early relapse after initial treatment. / ( %) pts received bv were refractory to the last treatment. all the baseline characteristics of patients are mentioned in table . median bv cycles administered were ( - ). overall response rate (orr) was % ( patients): cr in ( %), pr in ( %) ( / were primary refractory or early relapsed). median pfs for whole group was months ( % ci, . - . ). km estimated -year os was % and year was %, median os has not been reached yet. for patients who responded, pfs at months was % ( % ci, %- %), median pfs not reached. for / patients with progressive disease (pd) or non responders after bv, median pfs was only months ( % ci, . - . ). there was no difference in os between patients with responders and non responders. median os has not yet been reached in either group as mentioned in survival curves. at the median follow up of months (range: - months) patients are alive, patients are alive without disease, patients received consolidation bone morrow transplant ( auto-sct and allo-sct). patients completed courses and achieved cr. rest of patients who are alive without disease; they had pd on bv but achieved cr with other treatments. patients are alive with disease; patient is on bv and are on another treatment. patients have died, because of pneumonia while being on bv and due to pd. / patients who received bv, achieved cr after failing all previous treatments and are in cr. peripheral sensory neuropathy developed in patients; one required dose reduction. patient stopped treatment due to pulmonary toxicity. we are reporting largest single center data from middle east which confirms that bv as a single agent is effective and safe. overall response rate is lower as compare to pivotal trial but cr rate is comparable to other reported case series. this analysis also concludes that bv can be used as bridge to transplant in patients who don't respond salvage chemotherapy. disclosure of conflict of interest: none. was used to diagnose hiv infections. cox proportional hazards models were used to evaluate risk factors of overall mortality. fifty-six patients with nhl ( . %) and patients with mm ( . %) were positive for hiv antibody. in patients with nhl, overall survival was significantly lower in the hiv-infected patients than in the hiv-negative patients [ - year overall survival: hiv-infected patients, % ( % confidence interval, %- %) vs. hiv-negative patients, % ( % confidence interval, %- %), p o . )]. in a multivariate analysis, hiv infection was significantly associated with an increased risk of mortality (hazard ratio . , p o . ), and this effect was consistent regardless of transplant year. on the other hand, overall survival in patients with mm was similar between the groups [ % ( % confidence interval, %- %) vs. % ( % confidence interval, %- %), p = . ]. previous studies in europe and the united states showed comparable survival rates between hiv-infected and hiv-negative patients with nhl. however, our study showed that hiv infection was associated with a higher risk of mortality in patients with nhl in japan. suppression of t cell-mediated immunity or hiv related diseases might affect transplant outcomes in japanese patients. [p ] disclosure of conflict of interest: none. while beam and beac regimens (bcnu, etoposide, cytosar in both regimens and melphalan or cytoxan, respectively) are commonly used as conditioning high-dose therapy (hdt) in patients with non-hodgkin lymphoma (nhl), there have been few reports comparing these regimens. a retrospective analysis found the superiority of beam over beac in terms of overall survival (os) and event-free survival (efs). toxicities were similar, except that beam was associated with more frequent lower gastrointestinal (gi) mucositis. other studies reported that these regimens had similar efficacy and outcome. recently, a concern regarding cardiotoxicity of beac has risen. the current study aimed to compare efficacy and toxicity of beac and beam as consolidation hdt in young patients with mantle cell lymphoma (mcl) undergoing autologous stem cell transplantation (asct). this is a retrospective analysis of outcomes in mcl patients who received hdt with beam or beac followed by asct at bone marrow transplant centers in israel. os, disease-(dfs) and progressionfree survival (pfs) and regimen toxicity were compared. seventy seven mcl patients who were diagnosed between - / and received consolidation with beac or beam were included in the analysis. forty nine patients were treated with beam and patients-with beac. no significant differences between the groups were revealed in terms of age, sex, the mantle cell lymphoma international prognostic index (mipi) risk score, induction protocol and% of patients transplanted in first complete response (cr ) (mean age yrs in beam vs yrs in beac group; % of patients in beam group had mipi risk score - vs % in beac group; % of patients in beam group were transplanted in cr vs % in beac group). the amount of infused cd cells was significantly higher in the beam group (median cd cells/ kg: . in beam vs . in beac groups; p = . ); the number of days to platelet engraftment was significantly greater in the beac group (median days in beam vs days in beac group; p = . ). there were no differences in the number of blood transfusions or hospitalization days between the groups. the rate of grade - upper mucositis was significantly higher in the beam group ( % in beam vs % in beac group; p = . ); no other differences in toxicity (grade - lower mucositis, pulmonary congestion, infections) were observed between the regimens. non-relapse mortality by day posttransplant was % in both groups. a median follow-up was (range: - ) months. the -yr dfs in beam and beac groups was % and %, respectively (p = . ). there was no difference in the -yr os between the groups ( % in beam and % in beac group; p = . ). there was a trend to improved dfs and os in patients transplanted in cr receiving beam (p = . , figure) . in multivariate analysis, low-to-intermediate mipi and transplant in cr were found to significantly increase pfs (p = . and. , respectively), while the hdt regimen did not affect pfs. beac and beam hdt regimens followed by asct had similar efficacy in mcl patients. there was a trend to improved dfs and os in patients transplanted in cr and treated with beam vs beac. the toxicity profile was similar in both groups, except a significantly higher rate of grade - upper gi mucositis. [p ] disclosure of conflict of interest: none. early or refractory relapsed ( o year) diffuse large b-cell lymphoma has a very poor prognosis especially for those not responding to salvage chemotherapy. allogeneic stem cell transplantation is potentially curative. even though this is less likely in those not responding or having frank progression pretransplantation. methods: at our institution we identified all patients with aggressive b-cell lymphoma (diffuse-large b-cell lymphoma and blastoid mantle cell lymphoma) who were refractory or progressive to salvage chemotherapy with r-dhap and who had peripheral blood stem cells ( × cd +/kg body weight) collected after the st or nd cycle. after high-dose melphalane and autologous stem cell transplantation patients had a partial and a complete remission. patient died due to neutropenic infection, patients died due to progressive disease leading to a transplant related mortality of . %. median progression-free survival after autologous transplantation was . months. proceeded to allogeneic stem cell transplantation. patients had a matched related sibling, had a matched unrelated donor and had a mismatched unrelated donor. transplant related mortality was % in this heavily pretreated population. -year overall survival of all patients intended for treatment is %. one of these patients with relapsed mediastinal lymphoma after allogeneic transplantation was cured by salvage radiotherapy and is in long-term remission ( years). conclusions: salvage high-dose melphalane and autologous peripheral blood stem cell transplantation for diffuse large b-cell lymphoma as a bridge to allogeneic transplantation is potentially curative for a minor fraction of these patients. however, the remission rate of % ( % pr, % cr) and progression-free survival of . months after high-dose melphalane and autologous stem cell transplantation provides a window of opportunity to use new drugs and cellular therapies in these poor prognosis patients. high dose chemotherapy and autologous stem cell transplantation is the treatment of choice for patients with relapsed refractory hodgkin lymphoma. several factors including number of chemotherapy lines received before conditioning, time of relapse and remission status before transplantation can predict survival and pfs in patients undergoing autologous stem cell transplantation. in , we reported on a patients who underwent high dose chemotherapy followed by autologous stem cell transplantation from to . all patients with relapsed or refractory hodgkin lymphoma in the period of - , who underwent high dose chemotherapy followed by autologous transplantation were retrospectively analyzed. the main outcomes of the study were complete remission (cr) at day , overall survival (os) and relapse-free survival (rfs). the impact of the following variables on os and rfs: (a) disease status at the time of transplant, (b) number of chemotherapy lines prior to conditioning and (c) time of relapse months and (d) age. a total of patients were identified. the median age was year. there were . % females and . % males. complete remission (cr) was achieved in . % of patients and . % with chemotherapy sensitive disease at the time of transplantation. prior to conditioning regimen, . % received two chemotherapy lines, and . % received more than two lines. % relapsed in less than months and % relapsed more than months after completion of therapy cr at day was . %. the median os for the whole group was . months; the median rfs was , months. the number of chemotherapy lines significantly impacted os and efs. cr status before conditioning, favorably influenced os and efs with a trend toward better os in favor of those who underwent abmt while in complete remission. the time of relapse and the age did not affect survival outcomes. [p ] the outcome of patients with relapsed or refractory hodgkin lymphoma is favorable and the number of chemotherapy lines received before conditioning is the only factor that had a statistically significant impact on os and efs. since the identification of human immunodeficiency virus (hiv), a clear association between hiv and specific malignancies has been recognized. high-grade b cell lymphomas are the most common malignancy complicating hiv infection and one of three aids defining malignancies. diffuse large b cell lymphoma (dlbcl) accounts for % of cases. before , lymphomas were the cause of % of all deaths attributable to aids. after the introduction of highly active antiretroviral therapy (haart) overall incidence of adm declined, however longer survival and exposure to environmental risk factors have increased the incidence of non adm (adm) such as hodgkin's lymphoma (hl). since haart has improved overall survival substantially, the aim of chemotherapy should be complete remission rather than palliation with careful consideration of drug interactions and side of haart. between and a total of patients were detected hiv positive. twenty-one of these patients were diagnosed with a malignancy and patients referred to our department with a hematologic malignancy were evaluated retrospectively. diagnosis, stage, treatment, survival data were recorded. haart during chemotherapy, nadir cd count and cd count at diagnosis of malignancy was evaluated. four patients were diagnosed with high grade b cell lymphoma, patients with primary central nervous system lymphoma (pcnsl), patient with hl and patient with multiple myeloma (mm). all patients were male and median age at diagnosis was . ( - ). hiv seropositivity was identified during evaluation of malignancy in both pcnsl patients. median duration of hiv seropositivity before diagnosis of malignancy was months for the remaining patients. patient characteristics, treatment modification and cd counts are summarized in tables and . lymphoma was fatal in and the cause of death was identified as lymphoma progression in all patients including one patient diagnosed with hodgkin's lymphoma. a patient presented with multiple plazmositomas was diagnosed with multiple myeloma is currently receiving induction treatment together with haart. hiv related lymphoma patients frequently present with extra nodal disease, incidence of central nervous system involvement is also higher and prognostic score tends to be in the intermediate or high-risk groups. prognosis is also worse than hiv negative population. degree of immunosuppression is implicated and the duration of immunosuppression is directly correlated with the risk of developing lymphoma rather than hiv itself. haart allowed the use of aggressive chemotherapy since it improved immune system and decreased infectious complications. multiple myeloma is a rare neoplasm observed in hiv infection and the treatment is based on data obtained from hiv negative patients. treatment of such patients as well as lymphomas should take into consideration the toxic effects of haart combined with chemotherapy. since hiv positive [p ] patients are excluded from most studies, there are no guidelines to direct treatment and avoid toxicities. drug interactions should be monitored closely and modifications should be made accordingly. interruption of haart may not be mandatory since studies have shown safety of continuation of haart during chemotherapy. for newly diagnosed hiv and malignancy, careful clinical and laboratory evaluation should be made before postponing haart until after chemotherapy. disclosure of conflict of interest: none. the outcome of hdct and asct in refractory hodgkin lymphoma (r-hl) is not as encouraging as in relapsed hl. ten years ago we analyzed and reported outcomes of asct in our r-hl patients, however the follow-up was short. now we a reporting long term outcomes in r-hl after asct in one of the largest numbers reported to date. between and , patients with hl who underwent hdc and asct for r-hl in adult medical oncology (age years) were identified. r-hl is defined as partial response (pr), no response (nr), stable disease (sd), progressive disease (pd), relapsing within months (relapse o m) of finishing the planned (chemotherapy + radiation therapy (xrt)) treatment or refractory to salvage chemotherapy. kaplan-meier (km) method was used to estimate progression free survival (pfs) and overall survival (os) from the day of asct while progression is defined as progression of disease, relapse and death from any cause. all percentages are rounded to nearest. patients underwent hdc and asct during - and of them met the criteria of r-hl. male ( %), female ( %), median age at diagnosis was . years ( - years) and at asct was years ( - years). initial therapy was abvd in ( . %), mopp/copp alternating with abv or abvd in ( %) and others in ( %). ( %) had xrt after initial chemotherapy. response to initial chemo + xrt was pr in ( %), pd in ( %), cr in ( %) ( / relapsed within months and others have refractory relapse) and no response in ( %) and others in ( %). prior to salvage chemotherapy, ( %) had stage iii-iv, ( %) extra-nodal involvement, ( %) bulky disease and ( %) had b symptoms, spleen involvement in ( %), performance status , in ( %). eshap was used as first line salvage in ( %) or rd line ( %). post salvage / prior to hdc and asct disease status was pr in ( %), cr in ( %) and nr/sd in ( %). ( %) patients had a fdg-pet scan prior to asct, ( %) were in cr. beam was used as conditioning regimen. median follow-up for all alive patients is months ( - ) from asct. response rate post asct: cr in ( . %), pr in ( %), nr/sd in ( . %) and pd in ( . %) patients, others /unknown in ( . %). ( %) patients had xrt post auto-sct. type of first post hdc auto-sct event was no event in ( %), persistent disease in ( %), pd in ( %), relapsed disease in ( %), treatment related mortality in ( %) and died of other cause ( %). at last follow-up in november , patients ( %) are alive with no disease, ( %) alive with disease, ( %) died of disease and ( %) died of treatment related mortality or other causes. for entire group, km estimated median os is months, , , , and year survival is %: %: %: %: % respectively. median pfs is . month, , , , and year pfs is . %: . %: %: %: % respectively. we are reporting a very high risk group of patients with a very long follow-up. in patients with r-hl, eshap + beam combination resulted in high response rate ( . %). these remissions are durable. a year os survival of greater than % in our population is higher than most reports with similar numbers. although our cohort has a year os survival of %, % patients have either relapsed or died underscoring need for improvements in the management refractory hl. [p ] disclosure of conflict of interest: none. here we update the previously reported results of our reduced-intensity conditioning (ric) allo-hsct experimental program, initiated in . as of november , in our centre patients underwent ric allo-hsct. donors were hla-identical sibling in , fullymatched unrelated in , or -mismatch-unrelated in and haploidentical relative in . median age was years (range: - ). all patients ( m and f) had stage iib/iv refractory mf (n = ) or refractory ss (n = ). median number of previous treatment lines was (range: - ). source of stem cells was peripheral blood in patients and bone marrow in . median time from diagnosis to hsct was months (range: - ). conditioning included flu/ctx/tbi , pentostatin +tbi and flu/mel in case of hla-identical or unrelated donor, whereas the tt/flu/ctx/tbi regimen was used in the haplo setting. gvhd prophylaxis included csa/mmf in all patients, with the addition of atg in cases with unrelated donor and post-transplant ctx ( mg/kg giorni + e + ) in cases with haploidentical donor. full donor chimerism was obtained in / of the evaluable patients, in a median time of months (range: - ). grade ii-iv acute gvhd occurred in patients ( %), while grade iii-iv was observed in patients ( %). chronic gvhd occurred in patients ( %), being extensive in ( %), all transplanted from hla-identical sibling (no atg). following transplantation, a complete remission (cr) was achieved in out of the evaluable patients ( %), of whom experienced relapse at + and + months, respectively. transplant-related death occurred in patients ( %), of whom were in cr. out of the patients who did not achieve cr, died from progressive disease (median follow-up of months, range: - ), from a secondary malignancy and is still alive with disease months after transplant. of note, all pts who died in progression had chemoresistant disease at time of transplant. at the last follow-up, patients were alive and ( %) maintained cr after a median time of months (range: - ). in the whole population, the -year overall survival was % ( % ci - ) and the -year disease-free survival (dfs) was % ( % ci - ). however, when mf and ss were analysed separately, -yrs dfs were % ( % ci - ) and % ( % ci - ), respectively (figure) . apart from diagnosis, outcome appeared to be primarily associated with the disease status at transplantation, with a -yr dfs of % in the group of patients (n = ) who were in cr before starting the conditioning. after a median follow-up longer than years, we confirm the efficacy of ric allo-hsct as a powerful therapeutic strategy in inducing and maintaining remission in selected patients with chemosensitive advanced-stage ctcl, with results particularly encouraging in ss. [p ] disclosure of conflict of interest: none. outcomes of allogeneic hematopoietic stem cell transplantation for hodgkin lymphomas: a retrospective multicenter experience of the rete ematologica pugliese (rep) f gaudio , p mazza , am carella , d pastore , g pisapia , a mele , p galieni , n cascavilla , g specchia and v pavone hematology, university of bari, bari, italy; hematology, ospedale "san giuseppe moscati", taranto, italy; hematology, ospedale "casa sollievo della sofferenza", san giovanni rotondo, fg, italy and hematology, ospedale "cardinale panico", tricase, le, italy; hematology, ospedale "c. g. mazzoni", ascoli piceno, italy hodgkin's lymphoma (hl) is a potentially curable disease, and modern therapy is expected to successfully cure more than % of the patients. second-line salvage high-dose chemotherapy and autologous stem cell transplantation (sct) have an established role in the management of refractory and relapsed hl, leading to long-lasting responses in approximately % of relapsed patients and a minority of refractory patients. patients progressing after intensive treatments, such as autologous sct, have a very poor outcome. allogeneic sct represents the only strategy with a curative potential for these patients; this study reports a retrospective multicenter experience of the rete ematologica pugliese (rep) over the past years aiming to define the impact of patient, disease, and transplant-related characteristics on outcomes. patients with histologically confirmed diagnosis of hl who received allogeneic sct from to were retrospectively studied. the median age was years (range: - years) and ( %) were male. the majority of patients ( %) had had a prior autologous sct. at the time of allogeneic sct, ( %) patients had a chemosensitive disease and ( %) were chemorefractory. most ( %) patients received reduced-intensity conditioning, % received matched sibling donor and % matched-unrelated donor grafts. the disease status at day post-transplant was reported in out of evaluable patients. of the patients with chemosensitive disease, ( %) achieved a cr, ( %) had a pr or stable disease and ( %) had progressive disease. of the patients with chemorefractory disease achieved a cr ( %), had a pr or stable disease ( %) and ( %) had progressive disease. although the overall survival has improved significantly in mantle cell lymphoma (mcl) according to advanced treatment options, relapsed or refractory disease remains a challenge. recently, lots of targeted agents actively have been tried clinical studies and adapted to clinical practice in indolent lymphoma. however, the role of frontline autologous hematopoietic stem cell transplantation (auto-hsct) has not been fully understood in patients with mcl, compared with a few impressive published data about auto-hsct as salvage treatment option for patients with relapsed mcl. so, we retrospectively evaluated consecutive patients diagnosed mcl, and compared the clinical outcomes of high-dose chemotherapy followed by auto-hsct and conventional chemotherapy alone. between january and december , consecutive patients with newly diagnosed with mcl at catholic blood and marrow transplantation center in south korea were included in this study. all of the patients received high-dose cytarabine-containing regimen or chop with/without rituximab regimen for induction therapy regardless of transplant eligibility. the treatment approach in our institution for patients was based on the physician discretion for transplant eligibility or ineligibility that depend on patient age, comorbidities, and disease status. seventy patients were included in the analysis. initial chemotherapy regimens were consisted of chop (n = , %), r-chop (n = , %), r-hypercvad (n = , %), and hypercavd (n = , %). demographics and disease characteristics of both groups are shown in table . patients received auto-hsct were superior s overall survival (os; p = . ) and progression-free survival (pfs; po . ). the subgroup analysis according to high-risk of mcl international prognostic index (mipi) or bone marrow involvement was performed. between the two treatment arms among the high-risk mcl group, the clinical parameters were not different. the high-risk mcl patients with frontline auto-hsct showed superior os (p = . ) and pfs (po . ) compared with conventional chemotherapy alone. although mcl is classified within indolent lymphoma, frontline auto-hsct can be considered for patients diagnosed with mcl in the group of high-risk mipi or bm involvement with the favorable survival outcomes. disclosure of conflict of interest: none. nasal type extranodal nk/t-cell lymphoma (enktl) is a very rare and agressive malignancy characterized by a poor outcome. current standard therapy is not yet established. the role of high dose therapy followed by haematopoietic stem cell transplantation (hsct) is still controversial. we evaluated the outcomes of all the enktl patients undergoing hsct in a multicenter analysis on patients registered by the société francophone de greffe de moelle et de thérapie cellulaire (sfgm-tc) and compared them with a population of french patients who received chemotherapy alone. sixty four enktl ( males and females) received hsct, including allogeneic (allosct) and autologous transplantations (autosct). median age at the time of hsct was years (range: to years). overall, % of the patients presented with disseminated disease ( % and % in the allosct and autosct, respectively), % were in complete response (cr) at the time of hsct ( % and % in allosct and autosct groups, respectively) and % had received l-asparaginase regimen prior to hsct ( % and % in allosct and autosct groups, respectively). five ( %) and ( %) patients of the allosct and autosct groups underwent upfront hsct therapy, respectively. four patients received tandem autologous/ allogeneic transplants. in allosct, stem cell source was a matched related donor in patients, an unrelated donor in patients and an umbilical cord blood in patients. reduced intensity conditioning regimens (based on fludarabine-busulfan combination) and beam regimen were used in % and % of patients from the allosct and autosct groups, respectively. median overall survival for the whole cohort was . months (range: to months). the -year non-relapse mortality was . % and . % in the allosct and autosct groups, respectively (p = . ). the -year overall survival (os) and progression free survival (pfs) were . % and . % in the autosct and . % and . % in the allosct group, s respectively ( figure a) . the absence of cr prior to hsct was associated with a poor prognosis (p = . ). as compared to allosct, autosct resulted in a better outcome in patients who didn't achieve cr before transplant (p = . ) and tended to have better outcome in high pink risk score (figure b-c) . finally, at years pfs and os of patients who have been treated by chemotherapy alone (ct) (n = ) or followed by allosct (n = ) or autosct (n = ) in cr were %, % and %, % and %, %, respectively ( figure d ). in this french cohort, more patients received autologous hsct in upfront therapy than allogeneic hsct. in cr , there is no evidence suggesting that transplantation is associated to a better outcome than chemotherapy alone. however, a precise matching based on the pink score will be evaluated to ensure that patients who were intensified were not of worst prognosis. in refractory patients there is also no clear advantage to perform allosct when compared to autosct. however, in relapsing disease after ct or autosct allosct, allowed to obtained durable control of the disease. disclosure of conflict of interest: none. high relapse rate is one of concerns for allo-sct in pts with relapsed/refractory aggressive lymphoma. an optimal conditioning regimen designed for aggressive lymphoma may reduce relapse, especially during early post-transplantation period. however, it is not established yet. results of a german phase study of allo-sct with conditioning regimen of fludarabine, busulfan ( mg/kg po or . mg/kg iv), and cyclophosphamide with or without post-transplantation rituximab for relapsed/refractory aggressive lymphoma suggested the role of myeloablative busulfan-containing regimen in reducing relapse rate in pts with aggressive lymphoma. based on these results, we conducted a single institution prospective study to explore feasibility of the bmf regimen consisted of full-dose busulfan, melphalan, and fludarabine in pts with relapsed/refractory aggressive lymphoma (umin ). patients with aggressive lymphoma who achieved at least sd with salvage chemotherapy after experiencing either pd during first-line therapy, early relapse ( o mo) after firstline therapy, late relapse (≥ mo) but refractory to salvage therapy, relapse after auto-sct,; age - ; ecog ps of - ; and without severe organ dysfunction were eligible. donor source could be / matched related or unrelated donor pb/bm or cb with ≤ antigen mismatch; the bfm regimen was consisted of busulfan . mg/kg iv, fludarabine mg/m , and melphalan mg/m (yamamoto h. bbmt ). gvhd prophylaxis was csa + mtx (related pb), tac + mtx (unrelated bm), and tac + mmf (cb). primary end point of the study was survival with engraftment at day , and secondary end points were engraftment rate at day ; nrm and relapse rate at day and y; progression free survival (pfs), overall survival (os), and gvhd at y. protocol was approved by irb and written ic was obtained from all pts. twelve pts (male , female ) with a median age of y ( - ) were enrolled. ps was - in pts. diagnosis were dlbcl (n = ), transformed fl (n = ), enktcl (n = ), ptcl (n = ), and aitl (n = ). median number of previous line of therapy was . ( - ) and pts had failed previous auto-sct. diseases status at transplantation was cr (n = ), pr (n = ), and sd (n = ). donor source was cb (n = ), unrelated bm (n = ), and related pb (n = ). survival with engraftment at day , primary endpoint of the study, was achieved in %. neutrophil engraftment was achieved at a median of day ( - ). full donor chimerism at day was achieved in all of the pts evaluated. two pts developed vod which was manageable. with a median follow-up of mo, pts had progression of lymphoma at , , mo. five pts died and cause of death were progression of lymphoma in , interstitial pneumonitis in (at mo), systemic adenovirus infection in (at mo), and agvhd in (at mo). os and pfs at y were % and %, respectively. relapse and nrm rates were % and % (day ), and % and % ( y), respectively. agvhd of grade ii-iv was observed in / pts and pts developed limited cgvhd. this prospective study shows that allo-sct using myeloablative conditioning regimen with full-dose busulfan, melphalan, and fludarabine for relapsed/refractory aggressive lymphoma is feasible and deserves further evaluation. disclosure of conflict of interest: none. for patients with advanced ctcl, the allogeneic hsct seems to be curative with graft versus lymphoma effect playing a major therapeutical role. in this retrospective study, patients with a median age of years (range: - ) affected by ctcl underwent allogeneic hsct after a median of (range: - ) lines of chemotherapy, including autologous transplant for of them. the median time from diagnosis to hsct was months (range: - ). the diagnoses were: sezary syndrome (ss, n = ). mycosis fungoides (mf, n = ), primary cutaneous cd + lymphoma (n = ), panniculitis-like t-cell lymphoma (n = ), nk t cell lymphoma (n = ). at time of hsct, patients ( . %) were in complete remission (cr), ( . %) in partial remission (pr) and ( . %) had active disease. the patients were transplanted from an hla-identical (n = ), mismatched (n = ) or haploidentical (n = ) sibling, from matched unrelated donor (n = ) or from a single cord blood unit (n = ). different pre-transplant regimens were used as myeloablative (mac) in (th-bu-flu, n = ; bu-cy, n = ) or as reduced intensity (ric) in (th-flu-cy, n = ; th-bu-flu, n = ). al patients engrafted for neutrophils at a median of days (range: - ) and patients engrafted for platelets at a median of days (range: - ). acute gvhd was of grade -i in patients and ii-iv in ( . %). skin was the most common organ involved. five of evaluable patients experienced chronic gvhd which was mild in and severe in . at a median of months (range: - ), patients died ( mac and ric) because of gvhd (n = ), vod (n = ), pneumonia (n = ) or multiorgan failure (n = ). all patients surviving at months from transplant were in cr. only patients prepared with a ric (n = ) relapsed respectively at , , and months from hsct. these patients received dli associated or not to chemotherapy. three achieved cr, which remained stable in , while one patient died in cr from post dli acute gvhd. one patient (nk-t cell) not achieving cr is still alive with active disease. for all patients the median survival was months (range - ). with a median follow up of months (range: - ), patients ( mac, ric) are alive, in cr and with active disease. at years, the os was ± %; at years dfs was ± %. according with the median time ( months) from diagnosis to transplant, the -year os was ± % for patients transplanted early and ± % for the others (p o . ), while dfs was respectively s ± % and ± % (p o . ). despite the small number of patients, our results confirm the high susceptibility of ctcl to the graft versus lymphoma effect and point out the time to transplant as a crucial prognostic factors for the outcome. finally, the long-term follow up of our series strongly supports hsct for the cure of ctcl. disclosure of conflict of interest: none. recently, a new prognostic score, the nccn-international prognostic index (ipi) has been developed to stratify patients affected by diffuse large b cell lymphoma (dlbcl), and in high-intermediate and high risk groups the survival was equal or less than %. the aim of this analysis was to evaluate the outcome of a cohort of dlbcl patients undergoing high dose chemotherapy (hdc) as consolidation following first line chemo-immunotherapy, after their re-classification according to the nccn-ipi. we performed a retrospective study on patients diagnosed with dlbcl, with a high/intermediate or high-risk disease according to the ipi ( - ), who received upfront hdc with asct, in institutions. the patients were then re-stratified according to the nccn-ipi and arbitrarily classified in groups: low risk (nccn-ipi ≤ ) and high risk (nccn-ipi ≥ ). the pre-transplantation disease status was assessed by positron emission tomography (pet) or computed tomography (ct). the primary endpoints were non-relapse mortality, progression-free survival (pfs), overall survival (os) and relapse risk. the estimated -year pfs for all patients was . % ( % confidence interval [ci] . - . ) and the -year os was . % ( % ci . - . ). of these patients, had a low risk ipi score (ipi = ) and were considered high risk (ipi ≥ ). subsequently, the whole cohort was re-stratified according to the nccn-ipi: patients were allocated to the high-risk (nccn-ipi ≥ ) group, and to the low-risk group (nccn-ipi ≤ ). the analyses were then carried out for both groups. the -year pfs was . % ( % ci . - . ) in the low-risk group and . % ( % ci, . - . ) in the highrisk group (po . ), whereas the -year os was . % ( % ci . - ) in the low-risk group and . % ( % ci . - . ) in the high-risk group (p = . ). the significant difference in os and pfs between the two groups was mainly due to the cumulative incidence of relapse at years (graph ): . % ( % ci . - . ) in the low-risk group and . % ( % ci . - . ) in the high-risk group (po . ). non-relapse mortality was comparable in both cohorts: % ( % ci . - . ) for all patients. figure : cumulative incidence of relapse following hdc and according to nccn-ipi. patients affected by high-risk dlbcl still have an unsatisfactory prognosis after treatment with conventional therapy regimens, even in the rituximab era. the -year os and pfs in patients with nccn-ipi score ≥ range: from % to % and from % to % respectively . although this is a retrospective analysis subject to all related biases, our results suggest that upfront intensive therapy with autologous stem cell transplantation may significantly improve the outcome of these patients compared to conventional chemotherapy. the role of hdc in the treatment of dlbcl is controversial. however, new entities or new risk stratifications, as the one reported here, could allow to identify high risk subpopulations that could benefit from this approach. enteropathy-associated t-cell lymphoma (eatl) is an exceedingly rare and often rapidly fatal subtype of peripheral t-cell lymphoma, arising from intraepithelial lymphocytes. eatl type i is associated with celiac disease; type ii occurs in patients without inflammatory pre-conditions (according to who classification now called monomorphic epitheliotropic intestinal t-cell lymphoma (meitl)). surgical debulking and anthracyclinebased chemotherapy (ctx) followed by high-dose chemotherapy (hdctx) and autologous cell rescue (asct) are pursued when possible in this often malnourished and frail patient cohort. yet, even with intensive consolidation relapse occurs in - % of patients. the value of allogeneic hematopoietic cell transplantation (hct) is not clarified as of today due to limited reports. here, we report on a patient with meitl who was rescued with an allo-hct for his nd relapse following prior asct. moreover, we summarize the available literature on the use and outcomes of allo-hct for eatl and meitl. a y old man with spontaneous intestinal perforation was diagnosed with meitl following emergency partial resection of the small intestine. histology revealed infiltration by monotonous medium-sized lymphocytes with abnormal immunophenotype (cd +, cd +, cd +, cd -, cd -, tia- +) consistent with type ii eatl. post-surgical f-fdg pet-ct scan showed abnormal uptake in gastric antrum and pyloric region but no other manifestations. ctx with cho(e)p ( × ) followed by beam hdctx and asct was performed and achieved a complete remission (cr ). however, m post asct disease relapsed and was treated with × dhap, and × dhaox. cr was achieved after the rd cycle of salvage therapy. due to anthracyclineinduced cardiopathy allo-hct could not be performed at that time. m after completion of salvage therapy, disease relapsed again, and was progressive under pralatrexat treatment ( cycle, infusions). by then cardiac function had recovered and therapy was switched to dose-reduced mini-beam ( × ). in cr reduced intensity conditioning (ric; fludarabine, busulfan, atg) and allogeneic hct from a matched sibling donor was performed. ciclosporin a (csa) and mycophenolate mofetil (mmf) were given as gvhd prophylaxis. prompt engraftment in blood (day+ ) and full donor chimerism in the marrow (d+ ) were achieved. immunosuppression was tapered and discontinued on d+ (mmf) and d+ (csa), respectively. f-fdg pet-ct scan at m post-hct showed cr, but at m relapse was suspected (under work-up). only few cases of patients with eatl/meitl treated with allo-hct are reported in the literature (n = , table ), and the value of this highly aggressive therapy is not clear at this point. of note, the patients listed in table were given allo-hct instead of asct. long-term complete remission (cr) could be achieved in / patients, while patients suffered from early relapse and died of the disease (n = before d+ post allo-hct). asct following surgery and ctx appears to cure - % of patients in available series. no treatment concept is available for relapse following asct, and no published data are available for allo-hct for relapse post asct. the disease is exceedingly rare and is afflicted with very poor outcomes. therefore, patients given this aggressive treatment should be reported, even when treatment outcomes are not positive. disclosure of conflict of interest: none. strong graft versus lymphoma effects with low toxicicty of haploidentical hematopoietic stem cell transplantation comparing with hla-identical in t cell lymphomas: a retrospective multicenter study s bramanti, r devillier, s fuerst, b reda, a granata, s harbi, c faucher, i legrand, a santoro, d blaise and l castagna istituto clinico humanitas rozzano consolidation treatment of relapsed/ refractory t-nhl with allogeneic stem cell transplant (sct) is considered a curative options but few patients manage to undergo this procedure, due to the highly refractory nature of the disease. the primary aim of this work is to evaluate the gvl effects among t-nhl with both hla identical and haploidentical donors. we have retrospectively analized the long term outcome of consecutive patients affected by t-nhl, received hlaidentical or t-cell replete haplo-sct with pt-cy, in european centers, between february and october . the patients received nonmyeloablative (nmac) or reduced intensity (ric) conditioning regimen. gvhd prophylaxis consisted of mg/kg of pt-cy (day + and + ) in haplo setting and atg plus cyclosporine a in the hla identical setting. patients characteristics were reported in the table . no differences were founded in the two groups . most of the patients were transplanted in complete remissions but only as consolidation of first line. no graft failure occurred. the cumulative incidence of acute gvhd grade - was % in the haplo setting vs % in the hla id .extensive chronic gvhd was seen in % of haplo, and in % in the hla id . patients had cmv reactivation, hemorrhagic cystitis, and ebv reactivation. after a median follow-up of years os was % and % and pfs was % and % in the haplo vs hla id group see figure . nrm was % in haplo setting and % in hla identical one. the years cir is % and % in haplo and hla id setting respectively. this study confirm a strong anti-lymphoma effect of allo hsct without prohibitive toxicities. haplo-hsct with pt-cy shows low rate of cgvhd in a contest of poor prognosis t-nhl patients. [p ] disclosure of conflict of interest: none. ibrutinib is the first-in class bruton tyrosine kinase inhibitor that has been approved for the treatment of relapsed mantle cell lymphoma. however, despite the high response rate of % including % of complete response, the median duration of response is relatively short with an overall survival of % at months (wang ml, et al, n engl j. med ). we report a single experience of patients with relapsed mcl who underwent allogeneic stem cell transplant (allosct) after ibrutinib monotherapy salvage. all patients had previous autologous stem-cell transplantation (asct) before and were given ibrutinib at a dose of mg daily after the second or subsequent relapse. all patients had to be at least in pr according to cheson criteria before allosct. patients had an unrelated / ( ) or / ( ) allo-sct from peripheral hematopoietic stem cells after a reduced conditioning regimen with busilvex, fludarabine and antithymocyte globulin in association with zevalin according to our recent published phase study protocol (bouabdallah k, et al. ann oncol ). graft versus host disease (gvhd) prophylaxis consisted on ciclosporine and methotrexate. patients ( m/ f) were aged from to years and received between and previous chemotherapy regimens including asct in their last treatment strategy before introduction of ibrutinib. all patients had extensive disease with gastric involvement in patients and pulmonary localization in patient. median time between diagnosis and ibrutinib introduction was years ( - ) and the median time between asct and allosct was years ( ) ( ) . median duration of ibrutinib treatment was months ( - ) and it was stopped one week before proceeding to allo-sct. it was not planned to restart btk inhibitor after transplant. patients were assessed for response after at least months of treatment with ibrutinib. at time of evaluation, all patients were in complete ( ) or very good partial response ( ) before allosct. the patient in partial response had % tumor reduction with persistent gastric ulcer where histology examination shows cd + but negative cycline d lymphoid cells. all patients engrafted (median duration of pnn o g/l = days ( - ) and median duration of platelets o g/l = days ( - )) with fulldonor chimerism at month. one patient had a grade ii cutaneous chronic gvhd (cgvhd) with favorable outcome and developed months later a bronchopulmonary obstruction syndrome related to cgvhd. with a median follow-up of months ( - ) after allo-sct, all patients are alive in cr. one patient, in complete metabolic response before transplant had a gastric relapse months later but achieved again a cr months after reintroduction of ibrutinib. after the first case reported by furtado m et al (leuk lymphoma ), we report here additional cases with longer follow-up after allogeneic transplantation. the excellent tumor control after treatment with ibrutinib together with a very good outcome after allosct should drive to consider this approach in young patients with mcl relapse after asct. disclosure of conflict of interest: none. autologous hematopoietic stem cell transplantation (autosct) is considered the standard approach for high risk or relapsed/ refractory non-hodgkin and hodgkin lymphoma. although a large variety of conditioning regimens are available, including the widely used beam (carmustine, etoposide, cytarabine, melphalan), there is no consensus regarding a standard approach. in the context of carmustine shortage, we have chosen to replace it by thiotepa. however, clinical data about thiotepa-based autosct conditioning are still sparse, except some retrospective data for primary central nervous system lymphoma. thus, we designed a multicenter prospective study (nct ) to assess the efficacy and toxicity of a team (thiotepa, etoposide, cytarabine, melphalan) conditioning regimen. team regimen consisted in total dose thiotepa of mg/kg on day- ; etoposide mg/m / h and cytarabine mg/m / h (day- to - ); melphalan mg/m on day- . patients underwent autosct with team conditioning, and were included in this analysis if they have fullfilled the following criteria: age older than years, biopsy-proven hodgkin or non-hodgkin lymphoma, hiv seronegative, and first autosct. thirty-three male and nine female with a median age of years (range: - ) were analyzed thus far. karnofsky score was g/l was days (range: - ). of note, patients received thrombopoietic agents after engraftment because of persisting thrombocytopenia. the most significant regimen-related toxicities were mucositis in % of patients (median grade = , range: - ) and diarrhea in % of patients (median grade = , range: - ). other non-hematologic grade adverse events occurred in patients ( %) and no grade adverse events were observed. central line-associated bloodstream infection occurred in patients ( %). surprisingly, / evaluable patients ( %) developed human herpesvirus reactivation. only patients required intensive care unit transfer. the median duration of hospital stay was days (range: - ). after a median follow-up of months (range: - ), the non-relapse mortality (nrm) was %. only one patient relapsed of refractory aitl months after autosct and died month after. the estimated -year overall survival and progression-free survival were % and %, respectively. a team conditioning regimen seems to be a safe and valid platform in autosct for patients with high-risk or relapsed/ refractory lymphoma. although mucositis and diarrhea were frequent, there were no grade adverse events and no deaths related to the treatment. updated results with updated followup will be presented. disclosure of conflict of interest: none. the cell of origin has no prognostic impact on high-dose chemotherapy with r-beam and autologous stem cell transplant for diffuse large b cell lymphoma s lozano cerrada, r saliba, s srour, s ahmed, c hosing, r champlin and y nieto university of texas, md anderson cancer center, department of stem cell transplantation and cellular therapy diffuse large b-cell lymphoma (dlbcl) is a biologically heterogeneous disease that can be classified according to its cell-of-origin (coo). the germinal center b-cell (gcb) subtype has better outcome with frontline r-chop than the activated b cell (abc) subtype. however, the prognosis of these two types of dlbcl after high-dose chemotherapy and autologous stem cell transplant (asct) is less clear. the purpose of our study was to evaluate progression-free survival (pfs), event-free survival (efs) and overall survival (os) in a cohort of dlbcl patients treated with r-beam (rituximab, carmustine, etoposide, cytarabine, melphalan) and asct according to coo. we have the dicep regimen effectively reverses the poor outcome for lymphoma patients with suboptimal response or failure post st salvage treatment p kaloyannidis the outcome of patients (pts) with refractory hodgkin's (hl) and non-hodgkin lymphomas (nhl) post st salvage treatment (salv ) is considered poor. the published data, have shown extremely low survival rates ( - %) even after nd salvage treatment (salv ) followed by autologous stem cell transplantation (asct), due to the low response rates post salv and the high relapse rates post asct, confirming that the management of these pts remains a major challenge. we herein evaluated the dicep regimen [dose intesified cyclophoshamide ( gr/m ), etoposide ( mg/m ) and cisplatin ( mg/m ), days - ] as a salv treatment, in terms of safety and efficacy regarding disease response and stem cell mobilization/collection. moreover, we evaluated pts' long term outcome post asct. we retrospectively analyzed the data of ( hl, nhl) pts, with a median age of ( - ) yrs. twenty-one had suboptimal response ( % reduction): and minor response (≤ % reduction): ). three pts had stable disease while experienced progression. overall / pts underwent asct after a median of days (range: - ) post dicep. no pt was considered ineligible for the asct due to unacceptable toxicity post dicep; did not undergo asct because of progressive (n = ) or stable (n = ) disease. the -yr overall survival (os) was % for the whole cohort of pts ( % for hl and % for nhl, p = ns) while the -yr progression free survival (pfs) from dicep administration (± asct) was % ( % for hl and % for nhl p = ns). in particular, for the autografted pts, the -yr os was % ( % for hl, % nhl p = ns) and the -yr pfs was similar, % ( % for hl, % for nhl, p = ns) our data demonstrate that dicep is an effective salvage regimen with acceptable toxicity and no negative impact on the cd + collection. the promising response rates post dicep in combination with the very encouraging pfs rates achieved post asct, in this unfavorable and heavily pretreated group of patients, strongly support the rationale for using dicep as st line salvage regimen in selected pts in order to proceed to a successful asct. for the treatment of aggressive lymphoma, high dose chemotherapy followed by autologous stem cell transplant (asct) is an important component. however, the role of upfront asct in patients with diffuse large b cell lymphoma (dlbcl) is still controversial. furthermore, there is currently no consensus on a single best conditioning regimen for asct in patients with dlbcl. we retrospectively analyzed the records of patients with dlbcl who underwent upfront asct in state of complete remission (cr) or partial remission (pr) from institutions in korea. we evaluated the outcomes and prognostic factors of upfront asct in patients with dlbcl. we compared the outcomes of most widely used two conditioning regimens for asct; carmustine based regimens and busulfan containing regimens. total patients ( . %) achieved cr after asct and overall response rate (orr) was . %. with median follow up of months, patients ( . %) had progression or relapse. the -year overall survival (os) rates and progression free survival (pfs) rates were % and %, respectively. infection events were found in patients ( . %) and treatment related mortality was . %. these outcomes were comparable with the results of previous other studies. cox multivariate analysis for os showed that eastern cooperative oncology group performance status (ecog ps) ≥ (p = . ) and rituximab based induction therapy (p = . ) were significant prognostic factors. in addition, the following factors were significantly associated with pfs in multivariate analysis; female (p = . ), ps ≥ (p = . ) elevated β -microglobulin (p = . ), failure to achieve cr with induction chemotherapy (p = . ), carmustine based conditioning regimen (p = . ) and melphalan based conditioning regimen (p = . ). there were no significant differences in os and pfs according to stage, b symptom, bulky disease, high lactate dehydrogenase, bone marrow involvement, high or high-intermediate international prognostic index (ipi), absolute lymphocyte count and absolute monocyte count. therefore, it is considered that upfront asct can overcome the poor prognosis in patients with advanced stage or high risk ipi. in the analysis with conditioning regimen, neutrophil and platelet engraftment were slower in the carmustine group compared to the busulfan group. there were no significant differences in os between busulfan group and carmustine groups with -year os rates of . % and . %, respectively (p = . ). pfs at years was . % in busulfan group versus . % in carmustine group (p = . ). however, carmustine based conditioning regimen was poor prognostic factors for pfs in multivariate [p ] s analysis (p = . ). in subgroup analysis, busulfan group had significantly higher pfs compared to the carmustine group especially in female patients ( . months vs. months, p = . ), with b symptom ( . months vs. . months, p = . ) and abnormal serum ldh level ( . months vs. . months, p = . ). the outcomes of upfront asct in patients with dlbcl after induction therapy were acceptable. it is considerable in selected high risk patients who achieve cr with induction treatment, and have good performance status at diagnosis. in cases of conditioning regimen, busulfan based regimen resulted in improved outcomes compared with carmustine based regimen especially in patients with disseminated disease or female patients. disclosure of conflict of interest: none. no heavy chain was present in %). the predominant light chain was kappa ( %). patients had bence-jones positive myeloma. received bortezomib as induction therapy before transplant. we analyzed overall survival (os) and progression-free survival (pfs) in groups of patients. we separated the groups according to improvement in grade of response from preasct to postasct. the post-asct grade of response was measured months after asct. the os and pfs were estimated by the kaplan-meier method. pfs was measured from diagnosis to disease relapse and os was measured from diagnosis to death by any cause. results by subgroups of patients are detailed in table . median os and pfs of the whole group was years and months, respectively. if we analyze groups only by their grade of response before asct we find the following results: rc ( years os rate . %, median pfs months); pr/vgpr (median os . years and pfs months); sd/progression (median os . years and pfs months). according grade of response after asct, instead: rc ( years os rate %, median pfs: months); pr/vgpr (median os , years, and pfs months); sd/progression (median os . years and pfs months). in our experience, the grade of response before asct is a capital predicting factor for patients os and pfs. patient in cr before asct that preserve it after transplant, have a median pfs of months, the years os rate being . %. patients in situation of progression after asct have a very dismal prognosis (median os . years, pfs: months), however, patients who change from sd/progression to pr after asct have a median pfs of months and a os of . years. comparing these results we observe that this second group is particularly benefited by transplant. autologous peripheral blood hematopoietic stem cell transplantation in elderly patients with multiple myeloma as a standard therapeutic procedure. is it feasible? a single-center experience l cadievski, s genadieva stavric, z stojanoski, a pivkova veljanovska, d miloska, b kocoski, l cevreska and b georgievski university clinic of hematology, department for hematopoietic stem cell transplantation, university ss. cyril and methodius, skoje, republic of macedonia autologous peripheral blood stem cell transplantation (pbsct) represents a standard therapeutic approach in the treatment protocol of myeloma patients. it is known that multiple myeloma is a hematological disease that is a characteristic for the older population. autologous pbsct ideally should be performed in every myeloma patient, but with the elderly myeloma patients the procedure might be risky if know the possible comorbidities, or the possibility of the body to fully compensate the side effects of the conditioning regimen, the procedure or its possible complications. we present our experience in using high dose conditioning with melphalan mg/m followed by autologous pbsct for elderly myeloma patients, using the age limit od years. our retrospective analysis of our data during years of experience, shows that we have performed autologous pbsct on patients with myeloma at the age of or older. males ( . %), and female ( . %). patients ( %), were diagnosed with igg type myeloma, patients ( %) with iga myeloma, and patient ( %) with light chain myeloma. median age of the patients was . years ( - ). all patients were initially treated with cy-thal-dex regimen. in ( %) patients complete response (cr) was achieved, in ( %) very good partial response (vgpr), and in ( %), partial response (pr). in all patients the mobilisation of hematopoietic stem cells was performed with g-csf, and a median of apheresis procedures were performed, and the average number of collected cells was . × /kg tt mononuclear cells (range: . - . ). days to confirmed engraftment in our group of patients was . (range: - ). the number of blood transfusions was on average . (range: - ), and the number of transfusion of thrombocytes . units (range: - ). in the majority of patients, mainly after the year (that represents patients of the whole group), we used noncryopreserved hematopoietic stem cells, kept under the temperature of c, for median of days, thus avoiding the toxicity of dmso. additionally, we used central venous catheter inserted in the femoral vein for apheresis and application of the stem cells afterwards. the day after, the catheter was removed, thus avoiding catheter associated infections. all patients received standard infectious prophylaxis with fluconasole mg/daily, ciprofloxacin mg/ two times daily, acyclovir mg/ three times daily, cefixime mg/once daily, and ursodeoxycholic acid for vod prevention. no serious infectious complications were reported. our transplant related mortality was %. in the group with noncryopreserved stem cells no graft failure was reported. in two patients we even performed tandem autologous pbsct with no major complications. of the group of patients, the majority, patients ( %), had hta as comorbidity, ( %) with cardiomyopathy, and ( %) with inserted prosthetic aortic valves. three patients ( %) have died because of relapse of the disease. our oldest patents were and years old, and are still alive year posttransplant, in cr. we can conclude the performing autologous pbsct in elderly myeloma patients can be safe and effective therapeutic option, but with careful selection of the patients, balancing the risk profile of the patient and the benefit, or the risk of the procedure. affective supportive care, monitoring and reducing the risk of complications is an imperative to a good result. disclosure of conflict of interest: none. autologous stem cell transplantation program for patients with multiple myeloma in an outpatient setting k lisenko , s sauer , g egerer , j schmier , m witzens-harig , a schmitt , ad ho , h goldschmidt , , j hillengass and p wuchter , department of medicine v, heidelberg university hospital, heidelberg, germany; national center for tumor diseases heidelberg (nct), heidelberg university, heidelberg, germany and institute of transfusion medicine and immunology, mannheim, german red cross blood service baden-württemberg-hessen, medical faculty mannheim, heidelberg university, germany the first and second authors contributed equally. high-dose chemotherapy with melphalan and autologous blood stem cell transplantation (absct) for treatment of symptomatic multiple myeloma (mm) is performed in the usa and canada mostly on an outpatient basis, whereas in germany and western europe an inpatient setting is the standard. we report on a german single-centre program to offer the procedure on an outpatient basis to selected patients. major inclusion and exclusion criteria for eligibility were defined as follows: patients had to be able to reach the hospital within minutes, had reliable support from their family at home, had an ecog performance score of - and were willing and able to comply with the demands of the program. patients with severe co-morbidities were not included. all patients were treated on our outpatients' clinic and examined on daily visits by a team of physicians. feedback from patients was obtained by means of a questionnaire. from september to september , patients with mm stage iiia were enrolled. all engrafted within the expected time range: (median time to leukocyte , /μl and neutrophil recovery μ/l: days; median time to platelet recovery /nl: days, /nl: days). twenty patients ( %) had an episode of neutropenic fever but only in patients ( %) blood cultures were found to be positive. there occurred no cases of infection with multiresistent bacteria. although rather liberal criteria for hospital admission were applied, of patients ( %) could be treated entirely on an outpatient basis. eight patients ( %) were temporarily admitted for inpatient treatment with a median duration of . days (range: - days), mainly because of neutropenic fever. no severe adverse events occurred. feedback from patients revealed a high level of satisfaction with the outpatient setting. high-dose chemotherapy and absct on an outpatient basis is safe and feasible if conducted in a comprehensive surveillance program. the feedback from patients was very positive, thus encouraging further continuation and expansion of the program. disclosure of conflict of interest: none. high dose of melphalan (bor-mel). we retrospectively analyzed patients with mm who underwent asct between january and march . in these patients, conditioning regimen consisted of a high dose of melphalan ( - mg/ m ) intravenously on day - and two doses of intravenous bortezomib at . mg/m administered on days − and + . this cohort was compared with patients underwent asct between and , conditioned with high dose of melphalan alone. response rate was evaluated according to imwg criteria. all patients were evaluated after induction therapy and months after asct. all patients were followed until death or reference date (november, ). results: patients' demographics and baseline disease-related characteristics are shown in table . [p ] no difference was found in terms of neutrophil and platelet engrafment, hospitalization days (p= . ) and use of mechanical invasive ventilation (p= . ). bor-mel regimen did not enhance severity of preexisting peripheral neuropathy (pn) in any patients, and only one presented de novo grade pn. non relapsed mortality was . % and % in the bor-mel and mel cohorts, respectively (p= . ). complete response rate after transplant was significantly better in the bor-mel cohort than in the mel cohort ( . % vs. . %; p= . ) ( figure b) . when the analysis was restricted to patients who received bortezomib-based therapy, this difference was also statistically significant ( . % vs. . %; p= . ) ( figure d ). median of follow-up was months in the bor-mel vs. months in the mel cohort. no difference was found in terms of overall survival (os) and progression free survival (pfs) between both groups. for all patients, a post-transplant deeper response was associated with better os and pfs (p= . and p o . , respectively). our results are in line with previous studies demonstrating that bortezomib combined with melphalan is a well tolerated conditioning regimen and may enhance the response rate after transplant, even in patients receiving bortezomib in the induction therapy. these results should be confirmed in a randomized trial. for newly diagnosed patients (pts) with multiple myeloma (mm), the triple-agent induction treatment based on bortezomib plus dexamethasone in combination with cyclophosphamide (vcd) or lenalidomide (vrd) represent extremely reliable regimens, which in combination with early autologous stem cell transplantation (asct) result in high response rates and prolonged long-term outcomes. however, though both regimens are widely used, there are extremely limited studies that compare the vrd vs. vcd in terms of safety and efficacy. in the present study we compared the outcomes of newly diagnosed mm pts who received induction treatment vrd (n = ) or vcd (n = ) and proceeded early to asct. the vrd and vcd pts groups were similar regarding age at diagnosis ( vs. ys, p = ns), interval between diagnosis-asct ( , vs. , months, p = ns) and maintenance treatment post asct ( vs. pts, p = ns). per revised international scoring system (riss), the vrd-group had slightly more advanced disease (stage i: , stage ii: and stage iii: ), compared to vcd-group (stage i: , stage ii: and stage iii: ), however this difference was not statistical significant. the conditioning regimen consisted of single agent melphalan: mg/m . the t-test, kaplan-meir and cox regression were utilized for the statistical analysis. following a median of cycles of treatment (range: - for vrd vs. - for vcd, p = ns), in the vrd-group pts achieved complete remission (cr), pts very good partial remission (vgpr ≥ % reduction of m-band) and pt partial remission (pr: - % reduction of m-band) while in the vcd-group cr: , vgpr: and pr: pts (p = ns). the toxicities in terms of peripheral neuropathy, myelosuppression, liver and renal function were well tolerated and no patient discontinued treatment due to severe side effects. the -yr overall survival (os) was % for the vrdgroup vs. % for the vcd-group; nevertheless, the difference was not significant due to the size sample of the pt groups. the stage at diagnosis, the disease status pre-asct and the maintenance post-asct did not influence the os. interestingly, the -yr progression free survival (pfs) was significantly superior for patients who had been induced with the vrd regimen ( % vs. % p = . ) and for patients who achieved cr or vgpr before asct (pfs: %) while no pts with pr pre-asct was progression-free yrs post asct (p = . ). in multivariate analysis, only the cr or vgpr status before asct favorably affected the long term pfs. our results are in line with the limited published data from other studies with larger series of patients. in our study, very low disease burden before asct proven to be an independent factor for prolonged pfs. taking into consideration that vrd resulted in more cr or vgpr status, it is reasonable to conclude that vrd is a highly effective regimen and could be first treatment choice for newly diagnosed mm patients who are fit for early asct post induction. lenalidomide cohort and in the maintenance bortezomib cohort. baseline characteristics and outcome data were obtained via chart review. the primary outcome was pfs. the secondary outcomes were overall survival (os) and treatment-related toxicities. the median follow-up time was months. median time to death ( . years vs . , p = . ) and median time to progression ( . years vs . , p = . ) were not significantly different in the maintenance lenalidomide cohort compared to the maintenance bortezomib cohort. in the multivariate analysis, pfs was worse in patients at international staging system (iss) stage at diagnosis compared to those at iss stages and (hr, . ; % ci, . to . ; p = . ) and worse in patients with less than very good partial response (vgpr) to last prior therapy compared to those with a response to prior therapy of at least vgpr (hr, . ; % ci, . to . ; p = . ) [see figure ]. pfs was improved in patients with more than two years of maintenance therapy compared to those with less than two s years of maintenance therapy (hr, . ; % ci, . - . ; po . ), but this result does not account for patients who ended maintenance therapy due to disease progression. os was worse in patients at iss stage at diagnosis compared to those at iss stages and (hr, . ; % ci, . to . ; p = . ). peripheral neuropathy was more common in the bortezomib cohort ( % vs %, p o . ), while cytopenias were more common in the lenalidomide cohort ( % vs %, po . ). figure kaplan-meier curve for pfs for the maintenance lenalidomide group versus the maintenance bortezomib group by log-rank test (p = . ). lenalidomide and bortezomib maintenance after transplantation have equal efficacy in prolonging progression-free and overall survival in patients with multiple myeloma. iss stage significantly affects time to progression and overall survival, and response to last prior therapy affects time to progression. length of maintenance therapy may be a significant predictor and warrants further analysis. these findings suggest that both lenalidomide and bortezomib are acceptable maintenance therapy options for post-transplantation multiple myeloma patients. autologous stem cells transplantation (auto-hct) is an accepted method in multiple myeloma (mm) patients, but usually it is not curative. the issue of allogeneic hematopoietic stem cells transplantation (allo-hct) is challenging yet for myeloma. we investigated allo-hct in mm and compared with auto-hct. in this retrospective study, we recruited patients from january to january ( ( . %) patients in autologous group and ( . %) in allogeneic group). we performed allogeneic hct with peripheral blood stem cells source in our center for patients who are relatively young (less than years old) with good performance, have match sibling donor and accepted allogeneic hct. the conditioning regimens in autologous group was melphalan mg/m only and in allogeneic groups was fludarabine mg/m plus melphalan mg/m in consequent days. gvhd prophylaxis consisted of methotrexate and cyclosporine. the outcomes then compared between two groups using log-rank and gray tests and cox proportional hazard regression. the median follow-up in the autologous and allogeneic group was . months. three years disease-free survival of auto-hct was . % (ci: . %, . %) and . % (ci: . %, . %) for allo-hct patients (p value = . ). three years overall survival of auto-hct was . % (ci: . %, . %) and . % (ci: . %, . %) for allo-hct patients (p value = . ) showing no significant statistical difference between two groups. mortality rate was . % for auto-hct and for allo-hct was . %. the most common cause of death between two groups was relapse of primary disease. three year relapse incidence was . % (ci: . %, . %) for allo-hct and . % (ci: . %, . %) for auto-hct (gray's test p value = . ). the three year trm incidence was . % (ci: . %, . %) and . % (ci: . %, . %) in allogeneic and autologous patients respectively (gray's test p value = . ). despite there was no statistically significant difference between two groups in terms of os but dfs and relapse incidence was meaningfully better in allogeneic group. so, perhaps the reason of non-significant os improvement in allogeneic group is higher early death due to higher trm. we suggest that this study needs longer follow up to see whether allo-hct resulted in os improvement. disclosure of conflict of interest: none. myeloablative allogeneic hematopoietic stem cell transplantation from unrelated donors for patients with relapsed or refractory multiple myeloma n tsukada , s shingaki, m ikeda, t ishida and k suzuki division of hematology, japanese red cross medical center allogeneic hematopoietic stem cell transplantation (allo-sct) for patients with multiple myeloma (mm) is increasing in number despite in the era of novel agents, especially as a second line treatment and beyond. it has been reported that allo-sct for patients with mm resulted in high incidence of treatment related mortality (trm). high incidence of disease relapse is also a major problem especially after reducedintensity stem cell transplantation (rist). it is an important issue to reduce the incidence of trm while preventing disease relapse. the use of stem cells from unrelated donors is required for those without hla-matched sibling donors. the purpose of this study is to evaluate the feasibility of an intensified conditioning regimen incorporating both mg/ m of melphalan and gy of total body irradiation (tbi), followed by allo-sct from unrelated donors for patients with relapsed or refractory mm. we retrospectively analyzed eight consecutive patients who received allo-sct from unrelated donors with the conditioning regimen including gy of tbi, fludarabine mg/m for five days, and melphalan mg/m between april and july at the japanese red cross medical center. six patients received unrelated bone marrow transplantation (bmt) and two patients received cord blood transplantation (cbt). graft-versus-host disease (gvhd) prophylaxis was consisted of tacrolimus and short term methotrexate. the median age at allo-sct, the time from diagnosis of myeloma to allo-sct, and the numbers of prior treatment lines were . years (range: - years), . months (range: - months), and . lines (range: - lines), respectively. five patients are female. no episode of either grade ≥ iii toxicity or non-relapsed mortality was documented during the median follow-up period of over two years. cumulative incidence of grade ≥ ii acute and severe chronic graftversus-host disease were . % ( % confidence interval [ci] . %- . %) at days and . % ( % ci . %- . %) at days, respectively. probabilities of progression-free survival and overall survival were . % ( % ci . %- . %) and . % ( % ci . %- . %), at years, respectively. the results suggest that allo-sct conditioned with this intensified regimen may be tolerable for patients with relapsed or refractory mm. disclosure of conflict of interest: none. the role of allogeneic stem cell transplantation (allosct) in the era of novel myeloma drugs remains controversial. it is the only curative treatment option but non-relapse mortality makes the decision making difficult as opposed to achievements with autologous sct and new mm drugs by which the median survival is nowadays nearing years. aim of this study was retrospectively evaluate the outcome of allosct for mm performed at our institute, including evaluation of factors affecting survival. all consecutive patients allotransplanted for mm between and were included. the data were collected from our transplant registry. frequencies and medians were produced as appropriate. kaplan-meier method was used to calculate os and pfs and log rank test for comparisons. univariate analysis for factors affecting survival was performed with cox proportional hazard model. median age of all patients was ( - ) years. half of the patients had igg myeloma, % had iss score (score available for patients), and % had high-risk cytogenetics (data available for patients). response to treatment at sct was at least vgpr in % of patients, and transplant timing was early (within months from dg) in % of patients. sibling donors were used in % and muds in % of transplants, and conditioning was ma for % and ric for another % of patients. acgvhd grade - occurred in % and grade - in % of patients; % of patients had extensive chrgvhd. posttransplant cr rate was %. % of pattients have relapsed after allosct, and % are alive with the median follow-up of , years. non-relapse mortality has been % ( % until , % since then). the median survival of patients up to age of years is . years vs . years for patients years (n = ) with survival plateau after years at % level. transplant period, cytogenetics, donor type, conditioning intensity or occurrence of chrgvhd had no statistical impact on survival. significant differences in os were observed between disease status at scr vgpr vs o months from dg vs later), grade of acgvhd - vs - , and best resonse post-transplant cr vs not less than cr. the respective differences for pfs were in sct timing, grade of chrgvhd, and best post-transplant response. in univariate cox regression analysis the only significant factors for os were severity of acgvhd and cr vs other responses after sct, and for pfs allosct timing, severity of chrgvhd, and best response to sct. with allosct ca. % of mm patients can be cured but at the cost of high non-relapse mortality. the occurrence of grade - acgvhd and less than cr response to sct predict poor survival. considering the increasing survival expectations with modern standard therapy for mm, allosct may be recommended for younger patients with high-risk features, and allosct should be done early in the disease course. disclosure of conflict of interest: none. angiogenesis plays an important role in the pathophysiology of hematological malignancies including plasma cell myeloma (pcm). microrna- (mir- ) is overexpressed and displays oncogenic activity in cancers. however, little is known about the role of mir- in pcm. the aim of the present study is to examine the expression level of peripheral mir- in pcm patients and to determine its role in angiogenesis. vegf serum levels and mir- in pbmcs was measured in patients with pcm directly before melphalan mg/m followed by autologous hematopoietic stem cell transplantation (auto-hsct) and months after hsct; and healthy controls. the study population was divided into two groups after therapy: responders (stringent complete response, complete response, very good partial response, partial response) and nonresponders (stable disease, progressive disease). gene expression of mir- was quantified by sybr green real-time fluorescent quantitative pcr. further tube formation of huvecs and vegf secretion was measured in mir- mimic or inhibitor transfected human plasma cell myeloma cell lines h and rpmi- . the expression level of mir- was significantly increased ( . ± . versus . ± . ; p o . ) in pbmcs of pcm patients compared with healthy controls. further, serum vegf levels were increased in pcm patients ( ± pg/ml versus ± pg/ml in normal controls; p o . ). after auto-hsct, the expression level of mir- was significantly different in responders compared to nonresponders. responders had a lower expression of mir- compared to non-responders. further, serum vegf levels decreased in responders to auto-hsct compared to nonresponders. vegf expression was increased in the supernatant from mir- mimic transfected human pcm cell lines h and rpmi- compared with the negative control, while s vegf was decreased in the mir- inhibitor transfected cell lines. the angiogenic ability of huvecs was increased under pretreatment with the supernatant from h and rpmi- cells transfected with mir- mimic compared with negative controls and decreased when pretreated with mir- inhibitor transfected cells (fig. ) . this study demonstrated that mir- was upregulated in pcm patients. responders to auto-hsct had a decrease of mir- expression and vegf levels. further, mir- regulated angiogenesis. therefore inactivation of mir- or activation of its target gene may be a potential therapeutic approach in pcm. fig. : in vitro matrigel tube formation assay. (i), normal control (ii, iii), mir- mimic transfected h cells (iv), mir- mimic transfected rpmi- cells (v), mir- inhibitor transfected h cells. original magnification × . disclosure of conflict of interest: none. [p ] pilot study of busulfan/thiotepa as conditioning regimen followed by allografting and post transplantation cyclophosphamide in advanced relapsed myeloma patients c wolschke, e klyuchnikov, d janson, m heinzelmann, m christopeit, f ayuk and n kröger university medical center hamburg-eppendorf despite the significant improvement in outcomes has been observed for myeloma patients, the disease still remains incurable. due to limitations, such as trm and gvhd, the role of allogeneic stem cell transplantation as salvage therapy in this setting remains unclear. in present pilot study we provide data on the use of post cyclophosphamide (ptcy) as gvhd prophylaxis after a busulfan/thiotepa based conditioning regimen in patients who relapsed after autologous stem cell transplantation. between / and / myeloma patients (male n = , female n = ) with a median age of years (range: - ) (pts), who relapsed after autologous stem cell transplantation received allogeneic stem transplantation with with ptcy as gvhd prophylaxis after busulfan ( . mg/kg for age y and . mg/kg for age years) and thiotepa ( mg/ m )and for haploidentical and mmud additional fludarabin ( mg/m ). all pts. were relapsed after one or two autologous stem cell transplantations. donors were haploidentical (n = ), mmud (n = ), mud (n = ) and hla-identical sibling (n = ). stem cell source was pbsc (n = ) or bm (n = ). all patients received cyclophosphamide mg/kg of body weight on day + and + , which was in pts (n = mrd, n = mud) the only gvhd prophylaxis, while patients with mmud and haploidentical donor received also cyclosporine a from day + and mmf (until day ) and patients (mrd and mud) received additional cyclosporine. we observed no primary or secondary graft failure. the median time for neutrophil and platelet engraftment was (range: - ) and days (range: - ), respectively. major toxicities grade and were: renal (n = ) and mucositis (n = ). major infectious complications were: cmv: n = cmv-reactivations (n = ), sepsis (n = ), pneumonia (n = ) rsv-(n = ) and hsv (n = ). acute gvhd grade ii to iv and ii/iv was noted in % and %, respectively and mainly seen in patients with cyclophosphamide as single gvhd prophylaxis. remission rate were n = complete remission, n = vgpr, n = partial remission, n = n.a. after a median follow up of months pts progressed and patients (n = relapse, n = trm) died. the year pfs was % (n = ). busulfan/thiotepa is an active conditioning regimen for advanced relapsed myeloma patients. post cyclophosphamide might increase anti-myeloma activity, but as single gvhd prophylaxis it causes significant agvhd in mrd and mud and additional immunosuppressive agents such as cyclosporine should be added. disclosure of conflict of interest: none. magnetic resonance imaging (mri) for multiple myeloma (mm) is a sensitive, non-invasive and non-toxic method for detecting myeloma lesions. the goal of the study was to assess whether quantitative mri metrics can detect treatment response and replacement of neoplastic cells by fat marrow. the study was hipaa-compliant and irb-approved. we retrospectively identified all patients who achieved a complete response (cr) after induction therapy between and . inclusion criteria for the study was total spine mri imaging at diagnosis and after achieving cr. cr was determined using the imwg criteria. spinal vertebrae t through l were outlined with imagej software. fractures and lesions were excluded. images were analyzed using histogram-based (entropy, skewness, kurtosis) and texture-based statistics. a two-sided t-test was used to compare quantitative mri metrics from before therapy and after achieving cr. cox regression was used to explore the association between progression free survival (pfs) and change in each quantitative mri metric based on a median split. pfs was defined as the time from the second mri to death or progression of disease. nineteen patients met the above criteria. median age was . years (range: . - . ). majority of patients ( %) were male. majority of patients had iss stage disease ( . %) and standard-risk cytogenetics ( . %). an induction regimen containing an imid and/or a proteasome inhibitor was commonly used ( . %). all patients received an autologous stem cell transplant (asct) consisting of high dose melphalan followed by autologous stem cell rescue. three patients received a planned second asct. seven patients ( . %) were in cr before asct. nine patients ( . %) were treated with imid maintenance after planned initial therapy. median time to repeat mri imaging after cr was months (range: . - . ). mean change in measurements of kurtosis, skewness, entropy and texture analyses are shown in table . no significant change was detected between preand post-cr mri. furthermore, no significant association was seen between the change in any quantitative metric and pfs. [p ] despite promising results by other groups, we could not find a significant association between quantitative t image analysis and cr or pfs. there was heterogeneity in the time of repeat mri imaging which may have limited our ability to study interval change. although no definitive conclusions can be made from this small sample, correlation between pfs and kurtosis or texture d may be promising and should be investigated in a larger group prospectively. multiple myeloma (mm) treatment (tx) has evolved in recent years. solid data on the impact of new tx on patient (pt) outcomes outside clinical trials, however, is lacking. this study aimed at investigating tx practices, pt journeys, and outcomes in the real-world in countries with different access to new tx. the study was conducted between / and / in bulgaria, croatia, czech republic, poland, romania, and slovakia. it consisted of a cross-sectional (x) and a retrospective (r) phase. for the x-phase, investigators included all symptomatic mm pts seen during a -week counting phase to provide a snapshot of where in the pathway pts were at a given moment. for r-phase, investigators collected data on current and past tx, including symptoms, dosages, administration schedule, tx durations, tx interruption, reasons for change/discontinuation, and tx response. pts were selected in reverse chronological order with a quota of a maximum of pts who completed first-line ( l) tx within the past months (mo), pts in second-line ( l) and pts in third or higher lines ( +l). pts included in the x-phase could also be included in the r-phase, if they met the respective inclusion criteria. in total, physicians included pts in the x-and physicians included pts in the r-phase. in the x-phase, % of pts were o , % were - , and % were years; the median time since diagnosis was mo. % of pts were currently undergoing tx, % were previously treated and % had never been treated. of currently-treated pts, % received l, % l, % l and % +l. in the r-phase, % of pts were o years. of pts receiving l, % continued to l, % to l, % to l and % to l. of the % of pts eligible for stem cell transplantation (sct), % ( = % of all pts) received sct at l; these proportions were similar across countries. the most frequently-used regimens in l and l were bortezomib-based ( % and %, respectively), in l and +l lenalidomide-based ( % and %, respectively). median duration of l was mo, followed by a median disease-free interval (dfi) of mo. median dfi was longer in pts with sct than in those without ( . mo vs . mo). time to progression (ttp) decreased with later tx lines, from median mo at l to mo at l. depth of response, as assessed by the treating physician, decreased with each additional line of tx: % of pts achieved at least very good partial response (≥ vgpr) in l, while only % achieved ≥ vgpr at +l. ttp was longer in pts with better response levels: in l, median ttp for pts with ≥ vgpr was mo versus mo for pts with mo for pts with ovgpr. the most common ( ≥ %, all grades) adverse events (aes) and co-morbidities in l were anemia ( %), thrombocytopenia ( %), neutropenia ( %), neuropathy ( %), and fatigue ( %). these aes disrupted treatment in % in l, % in l and % in +l. the study found that of sct eligible pts, only slightly more than half were transplanted. poorer outcomes and increasing ae incidence with each tx line highlight the challenges of mm tx. information on real-world pt management may be valuable for physicians to plan their tx strategies and can provide input for health economic evaluations of existing and new tx. disclosure of conflict of interest: daniel coriu declares to have received consulting fees or other remuneration (payment) from novartis, amgen, pfizer, takeda, janssen. ivan spicka declares to have received research grants from celgene, consulting fees or other remuneration (payment) from bms, takeda, celgene, janssen-cilag, and amgen, and to be a member of the speakers bureau of celgene, janssen-cilag, amgen, and bms. zdenka stefanikova declares to have received consulting fees or other remuneration (payment) from amgen, celgene, and takeda and be a member of the speakers bureau of amgen, celgene, and takeda. daniela niepel, krisztian szabolcs toka, and paul schoen are amgen employees and hold amgen stock. dominik dytfeld, and georgi mihaylov have nothing to declare. safety and efficacy of autologous stem cell transplantation in elderly patients with multiple myeloma t maia , c marini , p medeiros , e aguiar , j cancela pires , r bergantim , f trigo and je guimarães , hematology department, centro hospitalar de são joão and faculty of medicine, university of porto autologous stem cell transplantation (asct) is considered standard treatment for multiple myeloma (mm) patients under the age of years, but its safety and efficacy still uncertain for patients over this age. retrospective analysis from one single centre concerning mm patients under, equal or over years who underwent asct between january/ and july/ . it was also compared to - years old mm patients diagnosed in this period of time who were not transplanted. we analysed a total of patients, of which underwent asct. onehundred-and-six of the transplanted patients were aged years or less (median , iqr years), patients were aged more than years (median , iqr years) and patients were non transplanted (median , iqr ). the conditioning regimen for younger patients who underwent asct consisted mainly of melphalan mg/m (mel ) while half of the elder patients received melphalan mg/m (mel ). regarding transplant-related myelotoxicity there were no statistical differences between patients aged years or less and over years old, however the first group needed less days of g-csf (p = . ). non-hematopoietic toxicity measured by infections and mucositis was not influenced by age. patients years conditioned with mel had more days of aplasia (p = . ), greater need of g-csf (p = . ) and transfusional support (p = . ) than patients ≤ years. there were no differences on non-hematopoietic toxicity. in the elderly group, patients conditioned with mel presented more aplasia days (po . ), higher grade of mucositis (p = . ) and more days of iv antibiotics (p = . ) than those transplanted with reduced dose of melphalan. comorbidities had no effect on transplant-related toxicity, either by age or by dose of melphalan. days of hospitalization and post-transplant complications did not differ according to age group. transplant related mortality was % at day posttransplant. survival after transplant in patients years old or under vs older patients (median follow-up time, months), was not influenced by age (os, mo vs mo, p = . ; pfs, mo vs mo, p = . ). regarding the non-transplanted elderly group, these are patients with more renal disease (p = . ) and poorer performance status (p = . ) than the transplanted cohort. there is also higher cytogenetic risk (p = . ). induction regimens were similar in transplanted group and non-transplanted group years old, and response to first line therapy (before asct of transplanted group) revealed no differences. infections were the most common complication in both groups. transplanted patients needed less days of hospitalization (p = . ). comparing the long term outcome of these two groups, survival curves of the elderly patients transplanted were clearly superior to the nontransplanted (os, mo vs mo, p o . ; pfs, mo vs mo, p o . ) although one has to consider that the non-transplant group has worse features than the elderly transplant group. transplantation in the elderly still debatable but this study shows that it might bring benefit. globally, transplant related toxicity is not influenced by age. regarding dose of melphalan, higher dose in elderly patients has higher toxicity, without apparent benefit in survival. therefore, age should not restrict the access to asct, but instead selection must be based on individual clinical and functional status. disclosure of conflict of interest: none. second autologous stem cell transplantation for relapse after allografting in multiple myeloma using cd + selected donor cells without immunosuppression p novak number of patients receiving a second allogeneic stem cell transplantation (sct) in europe is increasing despite high treatment related mortality (trm). in multiple myeloma only very few reports of second allogeneic sct exist with limited number of patients and substantial mortality. while in most hematological malignancies, the donor cell chimerism is dropping down if patients are relapsing, in myeloma donor cell chimerism remains complete despite relapse. to reduce trm we thought that full donor cell chimerism may allow us to perform a second high dose busulfan based chemotherapy followed by "autologous transplantation" after stem cell mobilization and collection. however, because two consecutive patients failed to collect sufficient cd + cells for an autologous transplantation even with plerixafor, we used donor t cell depleted cd + selected cells and transplanted those patients in an "autologous" fashion without any immunosuppression. to enhance graft-versus-myeloma effect, we added donor lymphocyte infusion (dli) at day . we report here on myeloma patients with a median age of years (range: - ) who relapsed after allogeneic sct and underwent a second "autologous" sct with cd + selected donor cells. all patients had received one (n = ) or two (n = ) autologous sct before . allografting. patients received an upfront auto-allo protocol and patients received . allogeneic sct as a salvage therapy. % of patients received a reduced intensity melphalan based conditioning regimen for . allogeneic sct and the median pfs was months (range: - ). before . allograft patients had received overall a median of (range: - ) treatment lines. at the time of . allogeneic sct all patients had a full or nearly full donor cell chimerism and remission status was very good partial remission (n = ), partial remission (n = ), stable disease (n = ), progressive disease (n = ). % of patients received a myeloablative busulfan based conditioning regimen and all received cd + selected stem cells with a median number of . × /kg cd + cells (range: . - . ) and × /kg cd + cells (range: . - ). engraftment was noted in % at a median of days (range: - ). no further graft-versus-host disease (gvhd) prophylaxis was performed and no acute gvhd (agvhd) was observed. according to treatment plan, patients received escalating dli around day + , starting with a median dose of × /kg (range: . - ) in combination with lenalidomide maintenance in patients. patients experienced agvhd ii-iv after dli. two patients had a severe gvhd (grade iii) which resolved completely after steroid therapy. no nonrelapse mortality after sct and dli was observed. after a median follow up of months (range: - ) the median pfs was months (range: - ) which translates into a pfs for all patients of % at year and % at years. median os was months (range: - ) and an os of % at years and % at years was observed. for patients with advanced multiple myeloma relapsing after allografting, a second "autologous" sct with cd + selected donor cells without immunosuppression followed by dli is an encouraging treatment option with low toxicity. disclosure of conflict of interest: none. second autologous stem cell transplantation as treatment option for relapsed patients with multiple myeloma: a single center experience (cic ) p ganeva, y petrov , m mincheff , i tonev , m guenova, l gartcheva , a michova , g arnaudov and g mihaylov ya. petrov; m. mincheff; i. tonev; l. garcheva; a. michova; g. arnaudov and g. mihaylov the use of modern therapies such as bortezomib, lenalidomide, thalidomide coupled with upfront high-dose therapy and autologous stem cell transplantation (asct) has resulted in improved survival in patients with newly diagnosed multiple myeloma (mm). the role of second asct as salvage therapy for relapse is unclear because of the availability of new agents to treat progression in multiple myeloma (mm). as the treatment options for management of patients with relapsed mm has become increasingly complex, physicians must consider both disease-and patient-related factors when choosing the appropriate therapeutic approach, with the goal of improving efficacy while minimizing toxicity. we retrospectively reviewed all mm patients who received a second asct as salvage therapy at our center from to december . for this period we performed transplants for mm patients. twenty five ( . %) patients received second asct ( patients were relapsed) and for patients asct was performed as tandem transplant. we analyzed only second asct for relapse. the median time to relapse after first transplant was . months (range: - months). all patients received reinduction therapy before the second asct. conditioning was performed with melphalan with two different doses ( mg/m and mg/m ). the median age at second transplant was . years (range: - years), and female/man ratio was / . median interval between first and second asct was . months (range: - months). we have no observed early deaths. until now ( %) patients are dead because of progression disease. response rate was assessed three months after asct, nine ( %) patients achieved vgpr, three ( . %) patients achieved at least a partial response, three ( . %) had sd and three ( . %) progressed despite salvage asct.median overall survival (os) was . months ( relapse ≥ months = . ; ≤ months = ). second asct is a feasible and safe option for salvage therapy in mm, especially in bulgaria where the possibility of using novel agents such as carfilzomib, lenalidomide, daratumomab for relapsed patients is limited to clinical trials, because of no reimbursement. the best results were observed in patients whose time to progression was more than months after first asct. advances in treatment of multiple myeloma (mm) has improved overall survival in these patients (pts). a steady increase in the risk of secondary malignancies has been reported over the last decades in mm survivors. estimated incidence of secondary acute myelogenous leukemia or myelodysplastic syndrome (t-mds/aml) after treatment with alkylating agents is %- . % per year - years after primary chemotherapy. no specific risk factor has been recognized, but genetic instability, natural history of the disease as well as induction therapy and autologous stem cell transplantation (hct) have been implicated. recently, novel anti-myeloma treatments have been linked with an increase in secondary malignancies, but no solid relationship has been established yet. in a retrospective study, we analyzed the incidence of secondary malignancies (t-aml/mds and solid tumors) in patients suffering from mm who had undergone autologous hct using high-dose melphalan conditioning regimen in our bmt unit. study population consisted of consecutive pts with median age of years ( - ), . % of them being male, who were transplanted during a period of years . type of myeloma was igg/a/d in %, . % and . % respectively, while . % was light chain and % nonsecretory. the majority of pts presented with k light chain myeloma ( . %). there was almost equal distribution between iss stages i and ii ( %/ . %) and only . % were diagnosed with advanced stage myeloma. most pts received two lines of chemotherapy ( %) and all of them more than one. treatment regimens before autologous hct included vad ( pts), bortezomib-based ( pts), dcep ( pts) and rd ( pts) and pts received radiotherapy. chemotherapy administration for mobilization was used in pts ( . %). conditioning regimen before autologous hct consisted of high-dose melphalan ( mg/m ) and in case of renal insufficiency mg/m . incidence of a secondary malignancy was . % after a median follow up period of months. t-aml /mds was diagnosed in ( . %) pts and ( . %) were diagnosed with breast and lung cancer respectively. pts diagnosed with secondary malignancy were previously exposed in induction therapy to melphalan ( ), vad ( ), bortezomib ( ), high-dose cyclophosphamide as mobilization treatment ( ) and radiotherapy ( ) . cytogenetic analysis was available in patients diagnosed with t-mds/aml and the majority ( / ) presented complex karyotype. abnormalities mainly observed were deletions and insertions in chromosomes , , . pts with secondary malignancies had an overall survival of months ( - ), however, after malignancy diagnosis, survival was very poor, four months only . secondary malignancies in pts with multiple myeloma after autologous hct occur with a substantial frequency and have a dismal prognosis. the role of novel treatment agents has to be elucidated. further studies are needed to identify new risk factors and develop better surveillance strategies. [p ] disclosure of conflict of interest: none. survival analysis after allogeneic hematopoietic stem cell transplantation in patients diagnosed with multiple myeloma: a single center experience p patricia hernandez* , r maria calbacho , a laura posada , g fabio augusto ruiz , r anabelle chinea hospital universitario nuestra señora de candelaria, santa cruz de tenerife (spain); hospital universitario ramón y cajal, madrid (spain) and hospital universitario cruces, barakaldo (spain) allogeneic hematopoietic stem cell transplantation (allohsct) may provide long term remission cures for patients diagnosed with high-risk multiple myeloma. however, its use is limited since it has a high rate of treatment-related mortality (trm), and because its efficacy compared to autologous hsct is not fully established. we studied patients that underwent allohsct between - . population characteristics are in table . all patients were treated at least with one prior therapy lines ( ) ( ) ( ) ( ) ( ) , all including autohsct ( . % underwent prior autohsct). % had or prior therapy lines. of them received bortezomib as part of treatment regimens. donor characteristics: non-related; hla-identical. gvhd prophylaxis: methotrexate plus a calcineurine-inhibitor: cyclosporine and tacrolimus. median follow-up . months ( . - . ), average was . months. seven patients died ( . %); because of progression ( . %), and ( . %) due to trm, including infections and immediate complications of transplantation, such as toxicities, icu admission and agvhd: infections: cmv reactivations, invasive fungal and bacterial infections. disease status: patients were in cr prior to allohsct. of them maintained it after. remaining patients died before disease was evaluated. seven patients were in pr prior to transplant, and reached cr after allohsct. one had progressive disease and reached cr after the procedure. two had stable disease and progressed after allo; one of them is in cr after additional therapy lines, and the other one died months after due to it. donor characteristics: hla-identical sibling donors: . % ( hla-mismatch, passed away . months after allo due to trm). one of the nonrelated donors, had an hla-mismatch, and died months after allohsct due to trm, the other one is alive after months. gvhd: ( . %) developed agvhd and of them maintained it chronically. two suffer from cgvhd, plus that initiated it as agvhd. were refractory to steroids. longterm survivors: patients had overcome three years after allohsct. they were among and years old at the time transplant was performed. none of them received bortezomib as part of therapy protocols for the disease. all had therapy lines prior allograft. were submitted to prior autologous hsct. relapse: patients relapsed after allohsct ( %, median time to relapse . months), being alive % at the end of the study. allogeneic hsct is associated with a high incidence of nrm and a low incidence of relapse. rates of acute and chronic gvhd are high. in our cohort, besides that more than % are alive until now, they suffer from extensive chronic gvhd and are in need of treatment. long-term survival may be related with patient factors such as young age, but also low tumor burden, or less prior therapy lines; in our group there are no differences in this aspect. studies including high-risk abnormal cytogenetics should help to define which patients are best candidates to allohsct. high-dose melphalan followed by autologous haematopoietic cell transplantation (ahct) remains the standard of care for eligible multiple myeloma (mm) patients. the majority of patients in clinical practice and trials receive a melphalan dose of mg/m (mel ), but a reduced dose of mg/m (mel ) is often used in patients perceived to be unable to tolerate mel . it remains to be determined whether this considerable dose difference results in different clinical outcomes. we therefore analysed patients with mm who underwent a first single mel or mel -conditioned ahct between january and december . all patients were included in the calm study, an analysis of a prospectively defined cohort of patients with data reported retrospectively to the ebmt, covering ahcts for mm and lymphoma. patients in the mel group were older than patients in the mel group at the time of ahct (median years [range: - ] vs years ; p o. ). compared to the mel group (n = , . %), fewer patients in the mel group (n = , . %) were overweight or obese ( . % vs . %; p = . ). compared to the mel group, more patients in the mel group had received proteasome inhibitor-containing induction therapy ( . % vs . %; p = . ), had a karnofsky score of ≤ ( . % vs . %; p = . ), and were transplanted in less than pr ( . % vs . %; p = . ). overall survival (os) from the time of ahct was not significantly different between the mel and mel group ( - significantly different between mel and mel ( days in both groups for neutrophil recovery; vs days for platelet recovery). however, late neutrophil recovery was noted in a small proportion of patients in the mel group. neutrophil recovery days post ahct was not observed in any engrafting patient in the mel group, but occurred in ( . %) engrafting patients in the mel group (p = . ). a cox proportional hazards model that included melphalan dose, age, and remission status at ahct showed that melphalan dose had no effect on os, pfs and relapse risk. the findings suggest that mel is not inferior to mel in younger and older mm patients and may reduce the risk of delayed haematological recovery in some patients. further analyses in relevant subgroups such as patients with high-risk features or renal impairment are required. disclosure of conflict of interest: none. high-dose therapy (hdt) followed by autologous stem cell transplantation (asct) remains the standard of care for patients younger than years of age with multiple myeloma (mm). different agents are being used to control the disease before asct, including the older thalidomide based combination or the newer bortezomib and lenalidomide based combination. the relation between the initial induction regimen and outcomes after asct is not completely clear. to evaluate the effect of different induction regimens on asct outcome, we retropsectively evaluated the outcomes of a low cost older regimen of thalidomide based combination in doublets or triplets with newer novel agents like bortizomib or lenalidomide based combination in a low resources country in transplant-elegible patients with multiple myeloma who underwent autologous stem cell transplantation at king hussein cancer center bmt program we retrospectively reviewed the files of patients diagnosed with mm from january till december , who received induction treatment followed by hdt and asct and followed up in a single institution. we compared the effects of different induction regimens, disease stage, and remission status before transplantation on over-all survival (os), event free survival (efs) and progression free survival (pfs) using kaplan meier curves. a total of patients were included, ( . %) of them received thalidomide based induction (group ) and ( . %) received bortezomib and lenalidomide based induction (group ). patients also offered no consolidation nor maintenance therapy. ( . %) patients were stage i, ( . %) stage ii and ( %) were stage iii. stage was not documented for ( , %) of cases. ( . %) were in complete remission (cr) and ( . %) were in partial remission (pr). the estimated -year os for the whole group was . %. there was no statistically significant difference between both groups in regards to initial iss stage of disease (p = . ) or cr status before asct. patients ( . %) in group achieved complete remission ( cr ) or very good partial response (vgpr), while ( . %) patient in group achieved cr or vgpr. there was no statistically significant difference between group and group in -years os ( -year os was % vs %, p = . ), efs ( . % vs . %, p = . ) or pfs ( . % vs . , p = . ). the use of an old, low-cost, thalidomide-based regimen in a low-resources country achieved a favorable transplantation outcomes in patients with multiple myeloma who received hdt and asct. double autologous stem cell transplantation (asct) is a useful treatment for multiple myeloma (mm) patients. we can make the second asct ( asct) without reinduction treatment (tandem regimen) or after a reinduction treatment after first asct ( asct) relapse (salvage regimen). we have conducted a retrospective study over mm patients undergoing a double asct performed in our centre from to . we have compared the different conditioning regimens used, and if there are any difference between tandem or salvage asct. we do not use maintenance treatment systematically. characteristics of patients and conditioning regimens in table . the overall survivals (os) of our patients are months (m) from treatment start till last control. the most important prognostic factors are the duration of the progression free survival (pfs) after asct (hr: . ( . - . ); p = . ), and the use of bumel like conditioning regimen at the asct or at the asct vs another conditioning regimens (hr: . ( . - . ); p = . ). today there are patients alive ( %), but only ( %) are free of mm now. the patients who were treated with tandem have a little better os than salvage patients ( m vs m; p = . . not significative). patients at tandem group who received different conditioning regimen at the asct and at the asct live more time than patients treated only with melphalan (mel ) at both asct. at salvage group the duration of pfs after asct is better than the pfs after asct ( m vs m). the use of the same conditioning regimen at the both asct has worse results than if we use different treatment. patients who were treated with s bumel at the asct or asct have better os than patients treated with cbv or mel . patients who not responded to reinduction treatment before asct have worst pfs after asct (rc: m, response; m and not response; only m). attention is drawn to the fact that patients who received bumel at asct have large os, but they are very few ( ) patients. only one patient has died during the asct, and was a patient of salvage group treated with bumel. double autologous transplantation continues to be a useful treatment despite the new mm treatments, and allows to prolonged the os. tandem asct probably is a useful treatment in high risk mm patients. salvage treatment is most useful in patients with a large pfs after asct, and good response to reinduction treatment. although mel continue to be the standard conditioning regimen for asct in mm patients, we have observed that patients treated with different conditioning regimen at asct and asct have better prognostic, and bumel has the best results in our serie. disclosure of conflict of interest: none. allogeneic haematopoietic stem cell transplantation (allo-hsct) is an effective treatment for myelodysplastic syndrome (mds) and acute myeloid leukemia (aml). the prognosis of elderly patients with mds and aml after chemotherapy is poor. allo-hsct is feasible in these patients; however the management of elderly patients with mds and aml for allo-hsct is difficult. we performed a retrospective survey of allo-hsct for elderly patients with mds and aml in our institution. we retrospectively analyzed the records of elderly patients ≥ years with mds and aml who underwent allo-hsct in our hospital between january and december . in this study, we assessed the ipss-r (revised international prognosis scoring system) cytogenetic score and the ipss-r score against the outcome of elderly mds and aml patients who treated with allo-hsct. fifty-one elderly patients with mds and aml were treated with allo-hsct in our institution, patients with mds ( with mds overt aml) and with de novo aml. ages ranged from to years (median ), patients were female and were male. there was a history of malignant disease in patients. according to the ipss-r cytogenetic scores of mds patients, patients fell in the good risk group, were in the intermediate risk group, were in the poor risk group, and were in the very poor risk group. regarding the ipss-r score, patient fell in the low risk group, in the intermediate risk group, in the high risk group, and in the very high risk group. sixteen patients were in st complete remission (cr), patient was in nd cr, patients were in partial remission, and patients were not in remission (nr) upon administration of allo-hsct. all patients received a reduced intensity conditioning regimen. patients [p ] were treated with fludarabine (flu), melphalan and low dose tbi-containing regimens; patients were treated with flu, intravenous busulfan and low dose tbi; and one patient was treated with flu, cyclophosphamide and low dose tli. graftversus-host disease (gvhd) prophylaxis consisted of tacrolimus plus methotrexate in patients, and tacrolimus, methotrexate and mycophenolate mofetil in patients. thirty-four patients received anti-thymocyte globulin (atg). the donor source was sibling bone marrow (bm) in patient, sibling peripheral blood stem cell in , unrelated bm in and unrelated cord blood in . relapse-free survival (rfs) and overall survival (os) were . % ( % confidence interval (ci): . - . %) and . % ( % ci: . - . %) at year, . % ( % ci: . - . %) and . % ( % ci: . - . %) at years, respectively (figure .) . in this study, patients died before engraftment. non-relapse mortality (nrm) was . % at day . twenty-five patients developed chronic gvhd ( patients limited and extensive). the causes of death were disease progression ( patients), treatment-related mortality ( patients), infection ( patients) and other causes ( patients). we suggest that many elderly allo-hsct patients with mds and aml were in the very poor risk group when the ipss-r cytogenetics score was assigned, in the very high risk group when the ipss-r score was assigned and nr upon administration of allo-hsct. rfs and os were . % and . % at years, respectively. there is a need for novel treatment strategies to manage elderly mds and aml patients for allo-hsct. [p ] disclosure of conflict of interest: none. counting bone marrow blasts as a percentage of nonerythroid cells provides superior risk stratification for mds patients with erythroid predominance a sun , y yu , t zhang , q wang , d liu and s chen the first affiliated hospital of soochow university, jiangsu institute of hematology, suzhou, china patients with erythroid predominance (≥ % erythroblasts, mds-erythroid) compose a significant proportion of patients with mds. the erythroid/myeloid subtype was divided from the aml category into mds-erythroid by the who classification of myeloid neoplasms. at that time, there was no consensus on a more appropriate way of enumerating bone marrow (bm) blasts from tncs or necs in mds-erythroid patients. to clarify these questions, mds patients were retrospectively analyzed in our center. mds-erythoid was observed in . % of patients ( / ), and these patients had similar clinico-pathological features and overall survival, with cases of mds with o % encs. by calculating the percentage of bm blasts from necs, of patients ( . %) with mds-erythroid who were diagnosed within who subtypes without excess blasts (eb) were moved into higherrisk categories and showed shorter os than those who remained in the initial categories (p = . ). recalculating the international prognostic scoring system-revised (ipss-r) by enumerating blasts from necs, of ( . %) mdserythroid patients with relatively lower risk were re-classified as higher-risk and had significantly poorer survival than those who remained in the lower-risk category (p = . ). this was especially true for the intermediate risk group that was stratified by ipss-r (unchanged patients vs. shifted patients, p = . ). however, the impact of enumerating bm blasts from necs on classification and prognostication was not evident in all mds patients. in conclusion, our results suggested that enumerating the percentage of bm blasts from necs significantly improved the prognostic assessment of mds-erythroid, especially for patients within the intermediate risk group stratified by ipss-r. disclosure of conflict of interest: none. myelodisplastic syndrome (mds) is a group of clonal and heterogeneous diseases, characterized by ineffective hematopoesis. the incidence of mds is about % of all blood disorders in children, approximately % of them develops acute leukemia. allogeneic hematopoietic stem cell transplantation (allo-hsct) is effective curative treatment of mds in children, but depends on disease status, type of clonal chromosomal abnormalities presented at the time of allo-hsct and graft quality. the aim of this study: to analyze the influence of graft quality on the outcome of childhood mds after allo-hsct. allo-hsct were performed in patients (pts) p hypomethylating agents vs. allogeneic sct in elderly patients with advanced myelodysplastic syndromes: a single center study j cermak, a vitek, m markova-Šťastná, j soukupova-maaloufova and p cetkovsky institute of hematology and blood transfusion, prague, czech republic a group of patients older than years of age with myelodysplastic syndrome (mds) raeb ii or with acute myeloid leukemia with multilineage dysplasia with less than % of bone marrow blasts (mds raeb-t according to the fab classification) was treated with hypomethylating agents (hma) and the results were compared to those obtained in an age and diagnosis matched group of patients who underwent allogeneic stem cell transplantation (sct). in the hma group, patients received azacytidine (vidaza) in the dose of mg/ m × every days and patients were treated with decitabine (dacogen) in the dose of mg/m × every days. median number of cycles administered was . (range: - ). in the transplanted group, patients were transplanted upfront and patients were pretretated with combination chemotherapy, patients received myeloablative conditioning and patients were transplanted after reduced conditioning regimen. a hematologic response to hma (cr,pr, hematologic improvement) was observed in patients ( . %), cr was achieved in patients ( . %). in sct group, engraftment was achieved in out of patients, patients died after sct ( on complications related to sct, patients relapsed). no difference in year survival was observed between both the groups ( . % for hma vs. . % for sct), however, median overall survival (os) was . months in hma treated group compared to . months in sct group (p = . ). in a recent analysis performed at months after starting the treatment, patients treated with hma ( . %) and transplanted patients ( . %) were alive, patients in hma group and patients in sct group relapsed. estimated years survival was . % in sct group and only . % in hma group (p = . ). no significant differences in results and adverse effects of treatment were observed between patients aged - years and those older than years in both hma and sct groups. our results confirm previous observations showing that despite a promising effect of hma resulting in hematologic response in more than % of elderly patients with advanced mds, allogeneic sct still represents the only potentially curative treatment connected with long-term survival in a significant number of patients even in this mds patients subgroup. disclosure of conflict of interest: none. immunophenotypic assessment of erythroid dysplasia by a simplified cocktail in myelodysplastic syndromes in taiwan c-c li , p-f weng , c-t lin , j-l tang hypomethylating agent (hma) is commonly used as a bridge therapy to prevent leukemic transformation prior to selection of a donor for allogeneic stem cell transplantation (sct) in patients with myelodysplastic syndrome (mds), and showed low toxicity. although its roles are known, the underlying genetics and clonal dynamics upon hma treatment has not been systematically examined using serial samples, especially in allogeneic stem cell transplantation (sct) setting. in this study, we performed targeted serial sequencing on bone marrow samples from mds patients treated with hma for bridging of allogeneic sct. to perform targeted deep sequencing, bm mononuclear cells before hma treatment and, and fractionated t-cell samples (cd + fraction) as controls were taken before hma treatment. analysis of genetic mutations were performed using targeted resequencing by illumina hiseq (sureselect custom probe set targeting [p ] entire exon regions of a myeloid panel consisting of genes). all patients received hma (decitabine: , azacitidine: ), and the median number of cycles was four (range: - ). the overall response rate for hma pre-treatment was %: there were four cases of complete remission (cr) ( %), six cases of marrow cr ( %), and two cases of hematologic improvement ( %). targeted sequencing revealed mutations in patients ( / , %) with median of mutations per patient (range: - ). mutated genes were then grouped into biological pathways, defined in the cancer genome atlas (tcga) aml study. the most frequent biological pathway at diagnosis was dna methylation ( %), followed by activated signaling ( %), chromatin modifiers ( %), tumor suppressors ( %), spliceosome ( %), cohesin complex ( %), npm ( %), and myeloid transcription factors (tfs) ( %). when assessing the difference in pattern of variant allele frequency (vaf), we found the significant reduction of vafs in four ( %) patients after hma. with a median follow-up of . months, -year overall survival (os) were . % ( % ci, . - . ). there was no significant difference in os according to the presence of mutations in each biological mutational pathway (all, p . ). to identify prognostic value of mutational dynamics, we subclassified the change of variant allele frequencies (vafs) after median fourth cycles of hma [no mutated or reduction of vafs ( patients) vs. stable or increased ( patients)]. however, there was no significant association between the dynamic of mutation and os (p = . ). these data show that hma using as bridge therapy for allogeneic sct in mds patients is insufficient to achieve the sufficient molecular responses and, mutational pathway and dynamics may not prognostic in this clinical setting. to clearly demonstrate the role of hma pre-treatment in mds, systematic assessment on a larger cohort is necessary. disclosure of conflict of interest: none. any role of high-dose chemotherapy in mediastinal nonseminoma germ cell tumors? p pedrazzoli, s secondino, a necchi , f lanza and g rosti istituto nazionale tumori, milano and ospedale santa maria delle croci, ravenna among germ cell tumors, primary mediastinal nonseminoma germ-cell tumors (pmnsgcts) have the poorest outcome with -year overall survival ranging from to %. indeed, the presence of mediastinal location defines per se a "poor prognosis" category according to the igcccg classification. this clinically and biologically distinct disease entity is associated with lower complete response rates to chemotherapy (ct), high rates of relapse and disappointing results from salvage ct. current standard first line treatment for patients with mediastinal primary location is still four cycles of peb, as for all igcccg poor-prognosis patients. we have reviewed available data present in the literature, including recommendations and expert opinions, on the use of high-dose chemotherapy (hdc) with autologous stem cell support in pmnsgcts. the use of hdc as both early intensification (that is, first-line setting) and at disease recurrence (salvage setting) have been reported in small cohorts of patients. according to the largest retrospective comparison, it has been suggested that hdc, given up-front, may produce a % to % absolute improvement in survival compared with standard dose ct. studies of the ebmt suggest that responsive disease after induction therapy may have a better outcome. mediastinal primary had salvage rates by hdct of less than % based on an international multicenter analysis and an ebmt study. the use of hdct in pmnsgcts warrants further investigation, preferably with the use of modern hdct strategies (that is, multiple carboplatin and etoposide courses). while hdc cannot be routinely recommended in pmnsgcts, selected patients with chemosensitive disease may benefit from early intensification. a retrospective analysis evaluating the large ebmt database is ongoing; results will be presented at the meeting. disclosure of conflict of interest: none. high dose therapy and autologous stem cell transplantation in gynaecological malignancies: a monocentric retrospective study m nderlita , i vergote and d dierickx university hospitals leuven, department of gynaecology; university hospitals leuven and department of hematology high-dose chemotherapy (hdt) followed by autologous stem cell transplantation (asct) has been established as a treatment option in many relapsed hematologic malignancies. however, in spite of many small trials, there still is no proven role for this treatment in solid tumors including most gynaecological epithelial carcinomas. however, in some recurrent non-epithelial ovarian cancers, such as sex cord stromal tumors, germ cell tumors, neuroendocrine gynaecological tumors and gestational trophoblastic disease, some studies suggest a possible role for hdt followed by asct. we performed a monocentric retrospective descriptive analysis of all patients diagnosed with gynaecological malignancies and treated with hdct followed by asct in our center. clinical, laboratory, pathological and imaging data were collected and analysed, together with information on treatment and outcome. eleven patients were included in this analysis, with a median age of years (range: - ) at time of diagnosis. eight patients suffered from ovarian neoplasia. at time of diagnoses patients showed metastatic disease. first line therapy consisted of surgery (n = ), chemotherapy (n = ) or a combination of both (n = ). median time to progression after first line therapy was . months (range: - ) with a median time between primary diagnosis and start hdt of . months (range: - ). three patients underwent single ast, whereas the other patients had a tandem ast, with a median time of months between first and second hdt (range: - ). treatment related toxicity was manageable, although there was treatment-related death. at last follow up patients ( %) were still alive with a median follow up of . years (range: . - . ) after last asct for all patients. of the deceased patients died with progressive disease. although the number of patients is very small, this retrospective study shows that hdt and asct is feasible in heavily pretreated patients with relapsed/refractory gynecological malignancies, although further studies are mandatory for optimal selection of patients, histological subtype and timing of hdt during the disease course. disclosure of conflict of interest: none. the human endogenous retroviruses (hervs) are remnants of ancient exogenous retroviral infections of the humans: they represent about % of the human genome . the basic genes of hervs are group-specific antigen (gag), polymerase (pol) and envelope (env); there are also two regulatory regions, long terminal repeat (ltr), located at ' and ' ends. several reports have shown that hervs may play a role in the development of autoimmune diseases, such as multiple sclerosis . additionally the existence of a strong relationship between hervs expression and cancer, based on the mrna expression profile of hervs in normal and cancer tissues has been suggested . the increased level of expression level of herv-h in colorectal cancer (crc), a major cause of cancer death worldwide has been already shown. the aim of the study was to analyse the expressions of env genes of herv-r, herv-h, herv-k and herv-p in the peripheral blood mononuclear cells (pbmcs), in the tumor and in the adjacent normal tissues of colorectal cancer patients. a group of control composed by pbmcs from healthy subjects was also included. rna was isolated from the biological samples and a reverse transcription assay was conducted. quantitative real time pcr was performed to evaluate the expression of the hervs env gene. all the env genes were related to the expression of an housekeeping gene, gapdh. the quantification was carried out using comparative ct method and the difference between the levels of env gene expression in pbmcs, cancer and adjacent normal tissue was given by fold difference. fold difference values were relative to a calibrator: first the pbmcs of patients and then pbmcs of control healthy group. Δct values were analysed using the paired sample t-test, followed by a bonferroni correction. higher levels of expression of herv-h, herv-k and in particular herv-p were found in tumor tissues, as compared to pbmcs and to adjacent normal tissues of patients, with an increase of -, -and folds, respectively. the Δct distribution of herv-h, herv-k and herv-p in cancer tissues were statistically significant (po . ) ( table ). the expression of herv-h, herv-k and herv-p env gene resulted increased in the colorectal tumor tissues also when compared with the pbmcs of the healthy controls ( -, -and -folds, respectively). the Δct distribution of herv-h, herv-k and herv-p in tumor tissues were statistically significant ( ρ < . ). no difference of expression was observed between pbmcs of healthy controls, pbmcs and normal adjacent tissues of patients (figure ). hervs env gene expression cannot be used as a diagnostic biomarker, but it is conceivable that hervs are directly involved in the pathogenic process of cell transformation and, if the protein expression will be demonstrated, the protein of hervs env gene could be the target for new immunotherapy strategies against colorectal cancer. [p ] disclosure of conflict of interest: none. a biosimiliar g-csf filgrastim is as effective as a reference drug however itis not as cost effectiveas it supposed to be and by the way no impact on the health care system m kurt yüksel, g pekcan, u sahin, s bozdag, s toprak, p topcuoglu, o arslan, g gurman and m beksac ankara university school of medicine biosimiliars are up to times the size of small molecule generic drugs and far more structurally complex. additionally biosimiliars are manufactured in living cell lines using processes that cannot be exactly replicated from one manufacturer to the next. a biosimiliar cannot be identical to its reference biologic drug. with billion dollars in global sales of biologic medicines anticipated to go off patent by .this lead to fast production of biosimiliar drugs. besides, it is expected that biosimiliar drugs will be more cost effective than the reference drugs and will have a meaningful impact on health care systems around the world. aim: to compare biosimiliarfilgrastim (leucostim) with two reference g-csf filgrastim (neupogen) and lenograstim(granocyte) in the context of safety, efficacy and cost effectivity. records of patients with multiplmyeloma(mm) whom underwent autologous stem cell transplantation(asct) and received g-csf sc mikrogram/kg/day from +day until engraftment were [p ] retrospectively evaluated mm patients were treated with high dose melphalan and asct at the ankara university school of medicine bone marrow transplantation unit between and . the median age was ( - years) with % male. patients were divided into three groups (n = ) whom received reference filgrastim (neupogen), lenograstim (granocyte) and biosimiliarfilgrastim (leucostim): groups a, b and c respectively. the total cost of each g-csf in dollars was calculated by one package of g-csf multiplied by total used days . chi-square, mann-whitney u and kruskal-wallis tests were used for analyses of variance. the percentage of patients who received melphalan mg/m were% , , in groups a, b, c respectively (p = . ).there was no statistically significant difference between the engraftment day of neutrophil and ; platelet and in the groups. (p = . , p = . , p = . , p = . respectively) themedian numbers of g-csf administered days were ( - ), ( - ), ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) in groups a, b, c, respectively .eventhough there was no statistical difference between the numbers of days( p = . ), the total cost in dollars was statistically difference between a vs b and c vs b (both p o . ) and there was no statistical difference between a vs c (p = . ), total cost in dollars as follows: $ ( - $), $( - $) and $( - $) for the group a, b and c respectively. our results demonstrate that biosimiliar gcsf leucostim is highly similar to existing licenced biologic products in turkey with no clinically meaningful difference interms of safety and efficacy. on the other hand it as a biosimiliardoes not have a meaningful impact on the cost savings to the health care system as expected when compared with reference filgrastim. disclosure of conflict of interest: none. in this study, we investigated the roles of prx ii, one of critical peroxidases besides catalase and gpxs, in cml primary cells at diagnosis and remission while patients were treated with sti (signal transduction inhibitor) and tested the same roles in imatinib(im) sensitive ph+ cell lines and resistant cell lines as well. newly diagnosed cml cells, im resistant k cells and parental k cells were treated stis and analyzed western blot assay to detect bcr-abl, phosphorylated bcr-abl and prx protein expression level. we added n-acetylcysteine ( - mm, hr) to k cells to show antioxidant effect of imatinib and analyzed dcf-da detection for intracellular ros level and western blot for prx protein level. mtt assay was performed to detect cell death by nac time-dependent treatment of mm nac( , , , hr). imatinib resistant k cells were established by treatment of gradual increment of imatinib. we also repeatedly investigated the effects of im therapy using prxii overexpressed k cells by transfection. at diagnosis of cml, ros level was elevated and prx ii was either absent or significantly suppressed. as ph chromosomes were decreased with stis, suppressed or absent prxs levels were restored to the level of normal individuals. these findings were also inversely correlated with the level of ph chromosomes in the cases of disease progression and re-remission with further treatment. when sti were treated in ph positive cell line, we found deceased cell survival and ros level by mtt assay and dcf-da methods respectively, but elevated prx ii by western blot. by the treatment of nac into ph+ cell lines, the level of dcf-da was decreased and mtt level was down, but prx ii level was elevated. interestingly, the level of bcr-abl oncogene were decreased in prx ii tranfected cells. meanwhile, we observed that prx ii restoration was mild or weak in imatinib resistant k- , which we established in our lab. the importance of the roles of ros and its prx ii, antioxydant enzymes in cml is further established by our work. our finding may contribute to find a new pathway on which tkis are working besides the mechanisms of atp binding competitively, blocking the binding of abl-bcr kinase to the substrate resulting apoptosis of ph+ cells. furthermore, our finding may be useful to overcome the stis resistant cml in the clinics in the future. disclosure of conflict of interest: none. allogeneic hematopoietic stem cell transplantation for the treatment of mucopolysaccharidosis f shunqiao, s xiaodong and l junhui department of hematology, capital institute of pediatrics, china mucopolysaccharidosis (mps) is a lysosomal storage disorder caused by deficient activity of the iduronate-sulfatase.this leads to accumulation of glycosaminoglycans(gags) in the lysosomes of various cells,which causes progressive multisystem involvement with ensuing death.the aim of this study was to exploit the effect of treatment with allogenic hematopoietic stem cell transplantation and administration of high doses of cyclophosphamide early after haplobmt in these cases. we retrospectively reviewed data from mps patients ( cases mps ii, and case mps i). the two mps ii patients were -month-old and -month-old boy and the mps i patient is a -month-old girl at the time of transplantation. the reduced-intensity of bu+flu conditioning regimen in allo-hsct for these patients was as follows: busulfan mg/kg at - days before transplantion,fludarabine mg/m at - days before transplantion.graft-versus-host disease(gvhd) prophylaxis:rabbit antithymocyte globulin . mg/kg daily at - days before transplantation,shortcourse methotrexate,posttransplantation high-dose cyclophosphamide on days + and + was followed by mycophenolate mofetil and cyclosporine.the donors all were their hlahaploidentical father. these three patients' neutrophil engraftment occurred on + d, + d and + d after transplantation respectively, platelet engraftment occurred on day + d, + d and + d after transplantation respectively.complete donor type engraftment was confirmed by short tandem repeat-polymerase chain reaction(str-pcr) on day after transplantation. no regimen-related toxicity occurred,gvhd and graft failure were not observed. month after transplantation, the activity of the iduronate-sulfatase was increased to normal. the motion of metacarpophalangeal joints ameliorated, regression of hepatosplenomegaly, the neurocognitive function improved. allogeneic hematopoietic stem cell transplantation is an effective measure to treat patient with mps at least mps ii and mps i. the reduced-intensity conditioning regimen was helpful to decrease the regimen-related toxicity. posttransplant cyclophosphamide approach successfully used and reduced the incidence of gvhd. this study aimed to evaluate the feasibility of alternative donor hematopoietic stem cell transplantation (hsct) using busulfan, fludarabine, and thymoglobulin conditioning for patients with chronic granulomatous disease (cgd) who lack an hla-matched familial donor. medical records of consecutive patients who received alternative donor hsct between may and may were reviewed, and the transplant-related outcome measures were analyzed retrospectively. the donor source was unrelated peripheral blood (pb) in , unrelated cord blood (cb) in , and haploidentical father in patients. only transplants ( / allele-matched unrelated pb) were hla-matched according to current standards relevant to the donor type. the conditioning regimen was uniform; fludarabine mg/m on days - to - , busulfan . mg/kg/d (or mg/m /d) on days - to - , and thymoglobulin . mg/kg/d on days - to - (or on days - to - in cb recipients). all but one patient were male and their median age at transplantation was . y (range: . - . ). one patient who received a cord blood graft suffered from primary engraftment failure, while the other patients were successfully engrafted with their chimerism levels ranging from % to % (median %) at month post-transplant. the median days to neutrophil and platelet engraftment were . (range: - ) and (range: - ), respectively. among the patients engrafted, one patient experienced secondary graft failure which was rescued by a second transplantation. the remaining one patient who failed to engraft was also rescued with a haploidentical graft from his mother. eight patients ( %) developed cmv antigenemia, and one of those patients developed cmv hepatitis. three patients developed grade acute gvhd which were manageable. one patient who developed grade hepatic gvhd eventually died. two patients developed extensive chronic gvhd, but became free of immunosuppressants after a complete resolution in one and with remaining stable mild joint contractures in the other. including patients who were rescued by additional transplantation, patients are alive with their latest chimerism levels ranging from . % to % (median %). the estimated -y overall survival rate was . % with a median follow-up of months (range: - ). even though the majority of our cohort underwent a mismatched transplantation, the survival rate was excellent. while conditioning with busulfan, fludarabine, and thymoglobulin seems feasible for alternative donor hsct in patients with cgd, special attention needs to be payed on cmv infection and severe gvhd which might offset the high survival rate. disclosure of conflict of interest: none. diarrhea is a common infectious complication in patients who had hematopoetic stem cell transplantation so, we aimed to detect entamoeba histolytica ratio before engraftment, amoung patients who had diarrhea after periferic hematopoetic stem cell transplantation (phsct) in our clinic. allogenic phsct patients had a median age of (range: - ) and autolog phsct patients had a median age of (range: - ). diarrhea is described as an abnormal increase in the frequency ( or more times per day), volume or liquidity of stools. we based upon this description in this study. we made stool examination in the first day of diarrhea. as stool examination, we used direct microscopic evaluation and adhesin antigen test specific for e.hystolytica with enzyme linked immunosorbent assay (elisa), e. histolytica ii, techlab, blacksburg, usa). we accepted e.hystolytica positivity as detecting cyst or/and trophosoit in stool and antigen test positivity at the same time. in our study, of patients had diarrhea in the first days of phsct. diarrhea was found in of in autologous phsct patients (% ), of patients in allogenic phsct with non-myeloablative conditioning regiment (% ) and of patients in allo phsct with myeloablative conditioning regiment (% ). diarrhea occured at + th day of transplantation and the median duration of diarrhea was days. e. histolytica positivity was found of patients ( %) who underwent phsct within first days of transplantation. infection is an important mortality and morbidity factor for patients who had hematopoetic stem cell transplantation, when especially before engrafment (between - days). autologous phsct patients were elderly, with poor self-care and low socioeconomic status individuals. e. hystolytica is a frequent pathogen in posttransplant diarrhea at endemic regions. prophylactic metronidasole treatment should be used routinely for autologous phsct as in allogenic phsct. patients and companions sholud be tested for e.hystolytica before autologus/allogenic phsct in endemic regions. prophylactic treatment for amebiasis and scanning patient/companions could be a part of solution for post phsct diarrhea. despite the emergence of disease modifying therapies (dmts) for multiple sclerosis (ms) a cohort of patients with aggressive disease have ongoing progression/relapse, associated with progressive disability. autologous haematopoietic stem cell transplantation (ahsct) has been used worldwide for aggressive ms with inflammatory changes on mri. we update on a uk single centre experience of ahsct in ms. a retrospective audit of ahsct performed for ms from to at uk centre (king's college hospital) was undertaken. patients were selected for transplantation based on persistent clinical relapses (relapsing-remitting ms) or secondary progressive neurological disability with mri lesion activity despite use of at least dmt. primary progressive patients were also eligible if new/active mri lesions were demonstrable. followup included clinical evaluation, edss assessment and mri scanning. we report our preliminary findings. as of november , patients ( female, male, rrms, spms, ppms) had received ahsct. mean age at transplant was . years (range: - ). the mean baseline edss was . (range: . - . ). patients underwent cyclophosphamide/atg conditioning, while received beam/atg. whilst conditioning and stem cell infusion were well tolerated there was a high rate of infections, with / patients developing a culture confirmed infection. reactivation of ebv and cmv were observed in a number of patients ( and , respectively) while a number of delayed herpes zoster infections were also seen ( cases of shingles and of disseminated varicella infection in patients who had previously experienced it in childhood). median follow-up was for days ( - ). of patients with a formal month assessment (n = ), had a stable edss, had an improved score (median improvement . , range: . - . ) and had a deterioration in their score (median . , range: . - . ). at months (n = ), had a stable edss, had an improved score (median . , range: . - ) and had a deterioration in score (median . , range: . - . ). at months, two patients assessed both had improvements in edss scores (median , range: . - . ). for patients who underwent mri at month follow-up (n = ), had a stable lesion load, demonstrated improvement in lesions and had a new lesion (the remaining mri was difficult to read due to a high baseline lesion load in this patient). patients had mri's at months; were stable and demonstrated a reduction in lesion load. to date, no patients have developed secondary malignancies or autoimmune diseases. of patients with followup data, / rrms patients experienced suspected clinical relapses following hsct-only one had a new lesion on mri (with no gadolinium enhancement). of the received steroids to treat these relapses (it is unclear if the remaining patient received treatment). patient tried a new disease modifying therapy ( dose of rituximab) following hsct. ahsct in this cohort was feasible with universal mobilisation and harvest. whilst conditioning and stem cell infusion were well tolerated, there was a high rate of infectious complications in the neutropenic phase. however, the transplant related mortality was % despite significant levels of disability amongst this patient cohort. ahsct remains a treatment option to be further investigated in this difficult cohort of patients. disclosure of conflict of interest: none. peripheral blood (pb) stem cells (scs) mobilized with g-csf are the first-choice source for allogeneic transplantation. we carried out a prospective study on healthy donors (hds), to identify donor characteristics that could influence the effectiveness of mobilization with special focus on the value of the basal cd + cell count. sibling hds were analyzed in a prospective study. we tested somatic variables (sex, age, weight, height, volemia) and, basal blood counts (white blood cell, peripheral blood mononuclear cell, platelets, hematocrit, hemoglobin, cd + cell). hds received g-csf subcutaneously at a dose of μg/kg day. two different determinations of cd + cells were done in each donor: baseline (before g-csf administration) and in pb on the morning of the fifth day (after g-csf administration). consecutive hds ( males) with a median age of years were enrolled. the mean value of cd +on day was . cells/μl, while the median value was . cells/μl. we performed two multivariate analyses either by using median regression (to predict the median value of cd +on day ) according to the values of cd + at baseline, the first adjusted by gender, age and blood volume and the second by gender, age and bmi. results of both models indicate that from basal cd + values o = to values ranging between and cells/μl, predicted median values of cd + on day significantly increase, from . to . cells/μl for model adjusted by blood volume, and from . to cells/μl for model adjusted by bmi. baseline, pb cd + cell count correlated with the effectiveness of allogeneic pbscs mobilization and could be useful to plan the collection. disclosure of conflict of interest: none. comparison of efficacy between chemotherapy plus granulocyte colony stimulating factor (g-csf) and chemotherapy plus g-csf and granulocyte-macrophage colony stimulating factor (gm-csf) for mobilization of peripheral blood stem cells (pbsc) and hematological recovery post-transplantation in patients with multiple myeloma (mm). a retrospective study of autologous peripheral blood stem cell (apbsc) mobilization data of mm patients who treated with chemotherapy plus g-csf or chemotherapy plus g-csf and gm-csf from may to july . the mobilization efficacy and hematopoietic recovery were analyzed. a total of stem cell mobilizations were performed in mm patients. in the univariate analysis, successful collection rate of single harvest in female and in patients at iss stage iii, r-iss ii/iii and chemotherapy plus g-csf was lower(po . ). however, age(≦ yrs vs yrs), subtype, d-s staging (i+ii vs iii), cycles of chemotherapy before mobilization (≦ cycles vs cycles), disease phase before mobilization (pr vs cr) and interval diagnosismobilization (≦ months vs months) were not correlated with the cd + cell collection and successful mobilization rate(p＞ . ). in the multivariate model, rate of successful mobilization in patients who received chemotherapy plus g-csf+gm-csf mobilization regimen was high (or = . , %ci . - . ). the effect of mobilization regimen remained significantly (p = . ). all patients successfully underwent hematopoietic reconstruction without transplantation-related mortality. chemotherapy plus g-csf +gm-csf mobilization regimen can significantly increase the effect of apbsc mobilization and ensure the reconstruction of hematopoietic function after transplantation. this mobilization regimen is a safe and effective method of mobilizing apbsc. disclosure of conflict of interest: none. clinical efficacy of bk virus specific t-cells in treatment of severe refractory hemorrhagic cystitis after hla haploidentical transplantation om pello , a bradshaw , a innes , s-a finn , s uddin, e bray , e olavarria, jf apperley and j pavlu centre for haematology, imperial college at hammersmith hospital, london, uk and department of clinical haematology, hammersmith hospital, imperial college healthcare nhs trust, london, united kingdom hemorrhagic cystitis caused by bk virus (bkv) is a significant complication of allogeneic hematopoietic cell transplantation (hct). it is particularly common in the setting of hla haploidentical transplantation and can be challenging to manage. here we present a post haploidentical hct patient who developed severe bkv haemorrhagic cystitis resistant to standard therapy and who responded to adoptive transfer of donor t cells enriched with anti-bkv specific cells. a year old man underwent hct for acute myeloid leukaemia with inversion of chromosome and monosomy of chromosome while in first complete remission. as he had no related or unrelated hla identical donor, cells from his hla haploidentical sister were used. on day + of this procedure he developed haemorrhagic cystitis. supportive treatment was initiated and cystoscopy showed diffuse bleeding from his urinary bladder with blood clots. urine pcr for bkv showed . billion copies/ml. despite bladder irrigation, local therapy to s bladder mucosa and intravenous hydration, he failed to improve, so treatment with weekly intravenous cidofovir was initiated on day + . his symptoms improved, but on day + he again deteriorated on weekly infusions of cidofovir. his immunosuppression was slowly tapered off without any graft versus host disease (gvhd) but without any significant effect on his hemorrhagic cystitis. he underwent bladder diathermy, was treated with intravesicular hyaluronate and with intravenous cidofovir, but continued to have frank haematuria with blood clots and significant lower abdominal pain. although there was no evidence of obstruction his renal function deteriorated on cidofovir therapy. hence we elected to trial adoptive anti bkv therapy. a leukoapheretic lymphocyte collection was used to prepare an anti-bkv t cell enriched product using the clinimacs prodigy and the cytokine capture system from miltenyi biotec. the eluted product contained % and % of cd + and cd + lymphocytes expressing ifng+ respectively and the cd +/cd + dose adoptively transferred on day + of transplantation was . × /kg. in vivo expansion of anti-bkv t cells in the patient was analysed weekly for the first month using the research grade peptivators bkv lt and bkv vp and the rapid cytokine inspector (cd /cd t cell) kit. bk viral load was monitored by pcr in urine samples twice weekly. ifng+ anti-bkv reactive t cells were undetectable in the patient for the first two weeks after adoptively transfer of donor t cells. twenty days after the adoptive transfer an increase in the cd + ifng+ population was observed, in response to the bkv vp peptivator. this observation correlated in time with a substantial decrease of the urine bkv viremia from . million copies/ml to copies/ml and a complete resolution of patient's symptoms. no gvhd, recurrence of urinary symptoms or any other problems have been observed to date (day + of transplantation, + days after the adoptive transfer). we are not aware of any other reports of successful adoptive anti bkv cellular therapy. our experience suggests safety and efficiency of the use of anti-bkv t cell enriched products using the clinimacs prodigy and the ifng capture system in hla haploidentical hct where bkv cystitis constitutes a significant complication. this opens the possibility of further clinical trials. disclosure of conflict of interest: none. haploidentical donor (hd) has been used as an alternative stem cell source when patients do not have a hla-matched related or unrelated donor. to overcome the hla barrier, haploidentical stem cell transplantation (haplosct) using post-transplantation cyclophosphamide (ptcy) has been conducted. here, we compared the clinical outcomes of haplosct using ptcy with those of unrelated donor transplantation. eighty-two patients ( from hd and from unrelated donor [ud]) who underwent allogeneic hematopoietic stem cell transplantation (hsct) in seoul national university children's hospital from january to june , were analyzed. there were no significant differences between hd and ud patients with respect to median age of patients, sex distribution, and diagnosis [ . %], p = . ). the conditioning regimen of haplosct included targeted busulfan, fludarabine and cyclophosphamide using ptcy, tacrolimus and mycophenolate mofetil for graftversus-host disease (gvhd) prophylaxis. all patients showed engraftment except for a patient who underwent unrelated hsct. neutrophil engraftment of ud was faster than hd (median days versus . days, respectively, po . ). however, there was no significant difference of platelet engraftment. incidences of complications, such as hepatic venoocclusive disease, cmv infection, and hepatic dysfunction, between both groups, were comparable, except hemorrhagic cystitis (hd: . %, ud: . %, p = . ). moreover, cumulative incidence of acute gvhd (hd: . %, ud: . %, p = . ), severe chronic gvhd (hd: . %, ud: . %, p = . ), relapse (hd: . %, ud: . %, p = . ) and non-relapse mortality (hd: %, ud: . %, p = . ) were not significantly different. the overall and event-free survival of hd and ud were . % vs . % (p = . ) and . % vs . % (p = . ), respectively. the clinical outcomes of haplosct using ptcy were comparable with those of ud, and a trend of lower cumulative incidence of severe chronic gvhd and non-relapse mortality was encouraging. it could be a promising alternative therapeutic option in pediatric hsct. disclosure of conflict of interest: none. , median number of apheresis procedures was , ( - ), median amount od dmso infused ml ( - ). time to engraftment was median days ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) . statistical comparison between cryopreserved pbsc grafts and bm showed benefit for pbsc in the terms (po . ) of faster engraftment, less infective complications, less transfusion support and less hospital stay. in patients ( %) dmso related events were not registered during graft administration. in patients ( %) mild to moderate dmso related events were registered, as nausea in patients ( . %), vomitus in patients ( %), tachycardia in ( . %), hematuria in patients ( %) and patients ( . %) with bradycardia, hypotension, fever and high temperature during graft infusion. cryopreservation of stem cells is a feasible procedure at our institution. there are some issues that have to be improved. the process is standardized with achieved engraftment in all transplanted patients. disclosure of conflict of interest: none. effectivity of a fludarabine based conditioning regimen in allogenic hematopoietic stem cell tranplantation for patients with severe aplastic anemia and over twenty years old p mustafa , , k melya pelin , s handan haydaroglu and g ilknur gaziantep university, faculty of medicine, department of internal medicine; hematology and bone marrow tranplantation unit, gaziantep, turkey severe aplastic anemia (saa) is an anemia with bone marrow hypocellularity and caused by hematopoietic stem cell failure ( ) . allogenic periferic hematopoietic stem cell transplantation (aphsct) is a curative treatment choice ( ) . although cyclophosphamide (cyc) and anti thymocyte globulin(atg) is accepted as standart conditioning regimen, especially for patients with high rejection risk, using fludarabin (fu) based regimens show increased successful engraftment ratio with minimal toxic side effects ( ) . to the study, saa patiens who were transplanted from hla matched sibling donors between the years - were included. the patients comprised of male (% ) and female (% ). median age was (range: - ). the median time from diagnosis to transplantation was (range: - ) months. conditioning regimen consisted of cyc ( mg/m ), fu ( mg/m ), atg (fresenius rabbit, mg/kg). the median dose of stem cells was × stem cell/kg (range: - ). methotrexate ( mg/ m given four days) and cyclosporine (cyca) ( - mg/kg given months) were applied for graft versus host disease (gvhd) prophylaxis. all patients ecog performance status were good ( - ). prior to transplantation only one of the patients received atg-csa, the others received only supportive treatment. after aphsct, neutrophil engraftment was occured at a median of days (range: - ) and thrombocyte engraftment was occured at a median of days (range: - ). posttransplant graft failure was observed in only one patient at tenth month and this patient had aphsct again from the same donor with the same conditioning regimen. acute gvhd didnot occur in any patient. the (% ) of patients had common chronic skin/oral mucosa gvhd. these patients received methylprednisolone (mp) and/or mycophenolate mofetil (mmf) in addition to the cyclosporine treatment. extracorporal photopheresis was applied to the two patients with chronic gvhd. all chronic gvhd patients had complete response to the immunsupresive treatment with a median follow up time months (range: - ). one patient died from sepsis. at year overall-survival rate was %. fu based conditioning regimen in aphsct with young saa patients has favorable results. fu based regimen might be a gold-standard treatment in the future. cgvhd; tgfb-induced factor homeobox , interleukin receptor gamma, tetra trico peptide repeat domain , carbonic anhydrase i, serpin peptidase inhibitor clade a and myod family inhibitor. we established a -gene model (myod family inhibitor, tgfb-induced factor homeobox , tetra trico peptide repeat domain ) to diagnose cgvhd. our -gene model showed . % sensitivity, . % specificity, . % precision, . % accuracy and . % roc area in diagnosing cgvhd. tgfb-induced factor homeobox increased in expression after rituximab treatment in responders. myod family inhibitor was found to be able to predict rituximab responses in steroid-refractory cgvhd patients. we could demonstrate that gene expression studies were useful in the diagnosis of cgvhd after allo-hsct. we developed a -gene model to diagnose cgvhd. hematopoietic stem cell transplantation (sct) is physically and psychosocially demanding. however, exercise interventions may have positive impact on sentiment and psychological well-being in patients undergoing sct. we report on a prospective, randomized study comparing the influence of a multimedia sensor-based practice with classical physiotherapeutic treatment (pt) on psychological aspects and quality of life (qol). patients undergoing sct were randomized into the control group (n = ) receiving pt or the experimental group exercising on the nintendo-wii (n = ). patients of both groups performed the exercises under the supervision of a physiotherapist and completed the functional assessment of cancer therapy -bone marrow transplantation (fact-bmt), hospital anxiety and depression scale (hads-d) and distress thermometer at the date of hospital admission (t ) and on day (t ), (t ) and (t ) after sct. questionnaires were completed by the participants independently and without supervision. groups were compared using the mann-whitney u-test. a p value o . was considered statistically significant. the median age of patients was years in the control group and years in the experimental group. results of fact-bmt generally showed a decline of the qol domains measured on t and t and a raise at t in both groups. physical well-being (pwb) showed the strongest fluctuation of all domains. it declined significantly between t -t in both groups (pt p = . , wii p = . ), followed by a significant increase between t -t (both groups p = . ). however, only in wii-group results of pwb at t ranked significantly above t (p = . ). highest scores were proved for social and emotional well-being (swb/ewb) in both groups. in wii-group ewb increased significantly between t -t (p = . ) and ranked above pt-group at all times. functional well-being (fwb) scored lowest in both groups at all times. the score of bone marrow transplant scale (bmts), the second lowest score in both groups, was always higher in wii-group. the level of distress was comparable between both groups. however, at t distress increased above the cut-off level of in both groups (wii-group p = . , pt-group p = . ). this was accompanied by an increase of anxiety (p = . ) and depression (p = . ) in the pt-group, while both parameters decreased in the wii-group (p = . and p = . ), respectively. anxiety in intervention group , / , / , /, at t /t /t /t stayed below standard group , / , / , / , at all times. depression averaged out at , / , / , / , in physiotherapy group and , / , / , / , in wii-group. to the best of our knowledge, this is the first study to show that exergaming using the nintendo-wii is feasible in the immediate phase after hsct. exergaming may be regarded as beneficial since our data indicate less psychological distress and higher qol in sct recipients exercising with nintendo-wii. therefore, it may be used in addition to pt. disclosure of conflict of interest: none. acute graft versus host disease (agvhd) is the most frequent and serious complication following haematopoietic stem cell transplantation (hsct), with a high mortality rate. a clearer understanding of the molecular pathogenesis may allow for improved therapeutic options or guide personalised prophylactic protocols. circulating micrornas are expressed in body fluids and have recently been associated with the etiology of agvhd, but global expression profiling in a hsct setting is lacking. this study profiled expression of n = mature micrornas in patient serum, using the nanostring platform, to identify micrornas that were dysregulated at agvhd diagnosis. selected micrornas (n = ) were replicated in independent cohorts of serum samples taken at agvhd diagnosis (n = ) and prior to disease onset (day post-hsct, n = ) to assess their prognostic potential. sera from patients without agvhd were used as controls. dysregulated micrornas were investigated in silico for predicted networks and mrna targets. profiling identified micrornas that were differentially expressed at agvhd diagnosis. mir- a (p = . ), mir- b- p (p = . ), mir- - p (p = . ), mir- a (p = . ), mir- a (p = . ) and mir- a (p = . ) were significantly verified in an independent cohort (n = ). mir- a (p = . ), mir- a (p = . ), mir- (p = . ), mir- a (p = ), mir- b ( . ) and mir- (p = . ) were differentially expressed days post-hsct in patients who later developed agvhd (n = ). high mir- b expression was associated with improved overall survival (os) (p = . ), while high mir- a and mir- b- p were associated with lower rates of non-relapse mortality (p = . and p = . ) and improved os (p = . and p = . ). pathway analysis associated the candidate micrornas with haematological and inflammatory disease. circulating biofluid micrornas are dysregulated at agvhd onset and have the capacity to act as prognostic and diagnostic biomarkers. their differential expression in serum suggests a role for circulatory micrornas in agvhd pathology, which warrants further investigation. disclosure of conflict of interest: none. factors associated with medication adherence amongst allogeneic hematopoietic stem cell transplantation recipients j lehrer , e brissot , , a ruggeri , , r dulery , a vekhoff , g battipaglia , f giannotti , c fernandez , , , m mohty , and m antignac ap-hp, hôpital saint-antoine, service de pharmacie, paris, f- ; service d'hématologie et de thérapie cellulaire, hôpital saint antoine, assistance publique-hôpitaux de paris; sorbonne patients with median ages of years (range: - years) between december and march , which including case of primary hlh (homozygous missense mutation in unc d: n = ; homozygous missense mutation in prf : n = ; heterozygous missense mutation in prf in the combination with hemizygous missense mutation in sh d a: n = ; mutation in rab a: n = ; mutation in itk: n = ). cases of unknown causes hlh, cases of lympgoma -hlh (nk/t-cell lymphoma: n = , primary γδt cell lymphoma in skin: n = ; subcutaneous panniculitis-like t cell lymphoma: n = ; primary t cell lymphoma in skin: n = ) and cases of ebv associated hlh. patients achieved cr+pr before hsct, and patients nr. patients were transplanted from hla-haploidentical family donors, from hla-identical sibling donors, and from a matched unrelated donor. conditioning regimen include tbi and non-tbi. the median overall survival rate was . % with a median survival time of months (range: - months). os of primary hlh is . %, os of unknown causes hlh is %, os of lymphoma-associated hlh is %, os of ebv-hlh is . %. os of cr+pr is . %, os of nr is . % patients without engraftment died because of graft failure and toxicity of conditioning regimen. patients with engraftment died. of those, patient died of hsct-associated tma, patient died of grade iv agvdh, patients died of relapsed hlh or organ failure as results from unsuccessful treatment of the progressively elevated ebv-dna load. patient died of tumor relapse, and patient died of infection. acute gvdh occurred in patients with grade i-ii agvdh in patients and grade iii-iv agvdh in patients; chronic gvdh occurred in patients. patients achieved completed chimerism, patients appeared with mixed chimerism,and patient presented with graft failure. of ebv-hlh with engraftment, reactivated ebv infection was found in ( %) with the whole blood ebv-dna load at - copy numbers per ml. ptld occurred in patients confirmed by pathology. after reduced immunosuppressors, negative result of ebv infection was obtained while patients developed gvdh. for ebv-hlh, patients who carry with ebv loading ebvdna ≤ copies/ml before transplantation, overall survival rate was significantly higher than that of ebvdna copies/ml (po . ); who achieved cr+pr os was significantly higher than that of nr (po . ); who range: from diagnosis to transplantion ≤ months os was significantly higher than that of months (po . ). allogeneic hematopoietic stem cell is an effective method for primary hlh and lymphoma-hlh, ebv-hlh,even haploid transplantation. the remission status before transplantation is decisive for the prognosis. disclosure of conflict of interest: none. hepatic veno-occlusive disease after allogeneic hematopoietic stem cell transplantation in a single centre: revised diagnosis and incidence according to new ebmt classification s santarone, a natale, p olioso, g papalinetti, t bonfini, p accorsi, s angelini, g iannetti, m di ianni and p di bartolomeo dipartimento di ematologia, medicina trasfusionale e biotecnologie, bmt center, ospedale civile, pescara, italy sinusoidal obstruction syndrome, also known as venoocclusive disease (sos/vod), is a potentially life threatening complication that can develop early after hematopoietic cell transplantation (hct). in this study we retrospectively investigated the incidence, risk factors and outcomes of sos/vod in transplants, performed in patients between march and may , on the basis of the new diagnostic criteria and classification of the ebmt. the patient's median age was years ( to ). of them, were males and females. patients received one transplant and two transplants. a diagnosis of hematological malignant and nonmalignant disease was present in and cases, respectively. the disease risk at hct was standard in cases, intermediate in and high in . an hla identical sibling donor was used in in cases, an unrelated donor in and a haploidentical family donor in . conditioning was myeloablative in transplants and at reduced intensity in . source of hematopoietic stem cells was bone marrow in transplants and peripheral blood in . we did not limit the diagnosis of sos/vod to the classical days after hct, but all suspicious cases appearing in the first days were evaluated. sos/vod was diagnosed in cases, of which in the first days after transplant and between day and (median day ). their main clinical characteristics are shown in table . the severity of sos/vod was mild in patients ( %), moderate in ( %), severe in ( %) and very severe in ( %). the cumulative incidence (ci) of sos/vod was . + . %. among the most relevant variables studied in univariate analysis (recipient age and gender, ferritin level at hct, type of hematological disease, disease risk at hct, type of donor, number of transplants, time of transplant, drugs used in the conditioning regimen, intensity of the conditioning regimen, source of stem cells), there was no factor with an adverse impact on sos/vod incidence. of patients with diagnosis of sos/vod, ( %) died. sos/vod was the main cause of death in patients and a relevant contributing cause of death in . of relevance, of patients ( %) with severe sos/vod and of patients ( %) with very severe sos/vod died, whereas only one patient with moderate sos/vod died and no patient with mild sos/vod died. among patients with sos/vod, received defibrotide therapy and the best supportive available therapy. defibrotide was given for a median of consecutive days (range: to ), starting at day post-hct (range: to ) with a median total bilirubin level of , mg/dl (range: . - . ). the -year overall survival (os) of patients treated with defibrotide was better as compared to that of patients who received the supportive therapy ( % versus %) although the difference doesn't reach the significance (p = . ). the occurrence of sos/vod does influence significantly the -yr os considering that it was + . % for patients without sos/vod and + % for patients with sos/vod (p = . ). in conclusion, the new ebmt diagnostic and severity criteria for sos/vod has been very useful in identifying patients with severe and very severe forms of this complication. if validated in prospective studies, these criteria will allow an earlier selection of patients requiring immediate therapeutic intervention. [p ] disclosure of conflict of interest: none. the prognosis of patients with newly diagnosed ewing's sarcoma family of tumors (esft) has improved significantly over the last few decades. nonetheless, the long-term survival is still below % patients with high risk features.the role of s high dose chemotherapy and autologous stem cell transplantation (hdct and asct) for high risk and relapsed esft was analyzed. a retrospective medical chart review was done on patients with efst who underwent hdct and asct between september and january at seoul national university children's hospital. indications for hdct and asct included metastasis at diagnosis, bulky primary tumor ( ml), axial/ central primary site, and relapsed disease. single hdct and asct was performed in the earlier period, and the regimen was changed from mec (melphalan, etoposide, carboplatin), to topothiocarbo (topotecan, thiotepa, carboplatin), and to bumel (busulfan, melphalan). tandem hdct and asct was performed in the recent period, st hdct with bumel and nd hdct with modified mec (melphalan, etoposide, carboplatin). twenty-one patients who were diagnosed with esft at a median age of . years old underwent conventional chemotherapy, radiation therapy and/or surgery and received hdct and asct in complete response (n = ) or partial response (n = ). the overall survival of the patients was . % at median . years and the event free survival (efs) of the patients was . % at median . years from the last asct. the efs of the patients who underwent single hdct and asct with mec (n = ), topothiocarbo (n = ), and bumel (n = ) was . %, . % and % respectively. the efs of the patients who underwent tandem hdct and asct (n = ) was . %. seven patients relapsed at median . months from the last asct. despite further treatment, patients died of disease and patients are currently alive without disease. one patient developed acute myeloid leukemia at . months from the last asct and is currently alive without disease after additional chemotherapy, hla-haploidentical stem cell transplantation and donor lymphocyte infusions. one patient died of transplantation-related mortality due to septic shock and lung infection. hdct and asct may be a promising treatment option for patients with high risk or relapsed esft. further refinements may be needed to identify the optimal regimen and number of hdct and asct. disclosure of conflict of interest: none. post transplant cyclophosphamide (pt-cy) has expanded the use of unmanipulated haploidentical grafts which have a high hla disparity between host and donor. one of the consequences of hla disparity is the development of engraftment syndrome (es). this is an immunological reaction characterized by non-infectious fever and skin erythema that develops after neutrophil engraftment. es resembles an infectious process but treatment involves the use of high dose steroids. our hypothesis is that pts undergoing haploidentical transplants (haplo) with pt-cy should have a high rate of es given the high hla disparity between donor and recipient. objectives: to determine the incidence, symptoms, morbidity and mortality of es in patients undergoing haplo with pt-cy at our institution. retrospective analysis of patients with highrisk hematological diseases undergoing haplo with pt-cy at clinica santa maria between november and august . es was diagnosed using the spitzer criteria ( ). es was diagnosed if pts met major criteria or major plus minor criterion. symptoms could occur prior to or after neutrophil engraftment (neutrophils over cells / ul). all patients signed informed consent and the study was reviewed by our institutional review board. patients received haploidentical grafts ( table ) . all patients had neutrophil engraftment at a median of days. / patients ( %) had symptoms that met criteria for es ( table ). / were transferred to icu due to hypoxemia and patient died after diagnosis of es. / pts were treated steroids. all patients received broad spectrum antibiotics during the febrile period and neutropenia. blood cultures, ebv and cmv pcr were negative in all es pts. there were no significant differences in hospital stay or one-year overall survival (os) between patients who developed and pts who did not develop es (median vs. days respectively, p = . ; one-year os % vs. %, p = . , respectively). es is a frequent complication in patients undergoing hsct haplo with pt-cy. the incidence of es in our study was higher when compared to historical full match related donors series and lower when compared to cord blood transplant studies ( ) there was no increased morbidity and mortality associated with es diagnosis. prompt institution of steroids is recommended in es patients after ruling out an underlying infectious process to avoid further complications. haploidentical allogenic hematopoietic stem cell transplants (haplo-hsct) is an alternative transplant procedure for patients with hematologic malignancies that are in need of transplant and do not have a compatible donor. due to the broad hla disparity, the haplo-hsct can be performed with t-cell depletion and megadose of cd +. alternatively haplo-hsct can be performed with non t-cell depleted transplants (t-replete) either in combination with anti-thymoglobuline serum (atg) or post-transplant cyclophosphamide (pt-cy) as gvhd prophylaxis strategy. center effect is a known risk factor for outcomes of haplo-hsct in both t-cell depleted (tcd) and t-replete settings. however, many centers tend to specialize in one gvhd prophylaxis strategy making it difficult to differentiate the treatment effect from the center effect. the objective was to investigate the role of center effects in gvhd prevention strategy, on leukemia-free survival (lfs) and overall survival (os) in a population of adult patients with acute leukemia receiving haplo-hsct. a retrospective multicenter study was conducted on patients reported to the ebmt registry. inclusion criteria were: age years, lymphoblastic or myeloid acute leukemia (all or aml) in first or second complete remission (cr or cr ), receiving a haplo-hsct between and . in this population (n = ), in order to assess the interaction between center and gvhd prevention treatment, we then included in the study selected patients from the centers that had performed more than % of both tcd and t-replete haplo-hsct during the study period. center effects on the outcomes consisted of ) center effect on the baseline risk of event and ) interaction between center and strategy of gvhd prevention. these center effects were estimated using cox mixed-effects models and tested using permutation tests. all models were adjusted on age, cmv statuses, disease (all or aml), secondary leukemia, previous autologous transplant, disease status (cr or cr ), peripheral blood vs. bone marrow transplant, conditioning regimen. after selection, patients were available across centers in europe. one hundred and one ( %) patients received tcd, t-replete haplo-hsct ( ( %) using atg and ( %) using pt-cy). overall, ( %) patients had aml. there were ( %) peripheral blood transplants in the tcd group and ( %) in t-replete. median follow-up was . years. in adjusted analyses, without accounting for center effect, t-replete tended to be associated with better lfs (hazard ratio (hr): . ( %ci . - . ), p = . ) and os (hr = . ( %ci . - . ), p = . ). when center effects were included, there was significant heterogeneity across centers on the baseline risk of both outcomes (lfs: p = . and os: p = . ). when accounting for interaction between center by strategy for gvhd prevention, the effect of t-replete vs. tcd on the outcomes did vary across centers (p = . and p = . for interactions in lfs and os, respectively) ( figure ). we found an interaction between center and strategy for gvhd prevention on outcomes of patients who received a haplo-hsct. the difference between the strategies (tcd or t-replete) varied across centers, in size and direction. this could be in part related to the increase in expertise with each technique in some centers and with the different management of complications, such as infections-related and relapse. disclosure of conflict of interest: none. adherence included recognition of spuriously high levels (typically from contaminated lines) and delayed dose adjustment due to late reporting of levels by the laboratory. the most common cause of unjustifiable non-adherence was failure to increase the dose in response to a low level. inadequate or excessive dose adjustments may be due to lack of experience or unfamiliarity with the sop. two interventions were launched with the aim of improving adherence to the sop for therapeutic tacrolimus dosing. firstly, to provide a rapid and user-friendly calculation method, we developed a mobile phone application (tacrocalc, a dose calculator based upon the sop algorithm) for android and ios devices using python and swift, respectively. secondly, to reduce the number of spuriously high levels, all nurses responsible for specimen collection participated in an educational module delivered by medical and senior nursing staff. key messages included the need to: use only the dedicated colour-coded tacrolimus lumen to infuse iv tacrolimus; avoid sampling from this lumen; sample peripherally when other lumens are known to be contaminated (reasons for this are being explored); suspend infusion of iv tacrolimus minutes before taking a level; send only immediately pre-dose levels for oral tacrolimus. initial re-audit of episodes post intervention (data collection is ongoing) demonstrated a % increase in sop adherence (p = . ; fisher's exact test), with no cases of unjustifiable non-adherence and a significant reduction in spuriously high levels. in conclusion, the use of tacrocalc by doctors and the implementation of targeted teaching for nurses dramatically improved adherence to the tacrolimus sop. this should ultimately improve therapeutic dosing whilst avoiding toxicity, which may result in better transplant outcomes. tacrocalc is now being adapted to include an option for paediatric dosing, with the potential to dose related medications such as cyclosporine. disclosure of conflict of interest: none. king's college hospital, imperial healthcare, charing cross hospital and imperial healthcare, hammersmith hospital managed with calcium and vitamin d alone in / cases ( %) and together with bisphosphonates in / ( %). osteoporosis was managed with bisphosphonates ± calcium/ vit d in / and with calcium/vit d alone in / . / indicated that they would give bisphosphonates in the absence of osteoporosis, if a patient with osteopenia was receiving long term steroids. dissemination and implementation of existing guidance on investigation and managing low bmd post hct appeared to be poor amongst respondents to our survey. routine dxa scanning was underused; the trigger for dxa in the context of steroids is inappropriately high at many centres at mg/kg daily for months; in established osteoporosis, bisphosphonates were used less frequently than would be anticipated. these findings may reflect the limited data on which current recommendations have been made, or the large number of non-transplant guidelines for investigating and managing low bmd which confound management of this post-hct patient group. hematopoietic stem cell transplantation (hsct) still remains as the most efficient therapy for adult patients with acute myeloid leukemia. for older patients and those lacking a hlacompatible donor, autologous hematopoietic stem cell transplantation (auto-hsct) is a valid therapeutic option. authors aimed for determining the effect of auto-hsct for acute myeloid leukemia patients and analyze group of patients who underwent auto-hsct. the study has been set as a retrospective single center study. clinical information included age, gender, aml type and cytogenetic risk. pretransplantation treatment, mobilization and conditioning were analyzed and thus subsequently authors used kaplan and meier method to calculate the actuarial overall survival rate. table describes patients' characteristics. majority of patients received similar induction therapy based on combination of cytarabine and anthracycline. timespan from the diagnosis to auto-hsct varied from days to days, median was days. seventy ( , %) patients received a preparative regimen consisting of busulfan at mg/kg orally, four times daily for days for a total dose of mg/kg administered on day - through day - and melphalan - mg/m intravenously for over hours on day - . patients achieved an absolute neutrophile count (anc) of ≥ . × /l in between to days; median was days. patients achieved not transfused platelet count ≥ × /l in between to days; median was days. median of patients' discharge from hospital was days (range: from to days) since auto-hsct. hundred day mortality after autologous transplant was at . % ( / ). on the date of our evaluation (april , ), patients were alive and in continued cr. the relapse rate was . % ( patients) and patients ( . %) were lost from follow-up. the -year overall survival (os) was . %, so the target median of overall survival has not been reached. [p ] the development of dyslipidaemia is commonly observed after haematopoietic stem cell transplantation (hsct). few data are available concerning lipid profiles over a long followup period or with regard to the different transplantation types (autologous vs. allogeneic) or the effect of multiple transplantations on the development of dyslipidaemia. a retrospective, single center cohort study including adult patients ( years) who underwent hsct at the university hospital basel s - and who survived ≥ days was performed. patients with at least a baseline lipid measurement were included (n = ) and grouped according to the type of their first hsct (autologous or allogeneic). for the examination of the effect of subsequent hscts, patients with consecutive transplantations of the same type were included and other patients were censored when a different transplantation type was performed. serial lipid profiles (total-, ldl-and hdl-cholesterol and triglycerides) before and after transplantation were examined. of the patients, underwent a first, and of these at least one subsequent autologous hsct. underwent a first, and of these at least one subsequent allogeneic hsct. median age of patients at autologous hsct was y (iqr - ) and y ( - ) at allogeneic hsct. % and % were males, median bmi pre-transplant was ( - ) and ( - ). the majority of patients underwent s intensive conditioning before hsct. median follow-up time was . years in the autologous and . years in the allogeneic group, with a maximum follow up time of . and . years, respectively. table shows the number and percentage of patients with dyslipidaemia ( st autologous and allogeneic transplants). the distribution of exact total cholesterol values along with comparisons with baseline measurements according to group are presented in the figure . *% based on number of measurements available total, ldl-and hdl-cholesterol and tg increased within months of transplantation, regardless whether autologous or allogenic transplantation or a first or a subsequent transplantation was performed. the percentage of patients with dyslipidaemia accordingly rose significantly within months of transplantation and persisted throughout follow-up. although patients undergoing an autologous hsct presented with higher baseline values of total cholesterol, a significantly greater increase post-transplant was observed after allogeneic hsct. first and subsequent transplantations seem to behave similarly with respect to changes in lipid profiles. disclosure of conflict of interest: none. nuremberg, erlangen, germany; department of cancer immunology, institute for cancer research, oslo university hospital, radiumhospital, oslo, norway; kg jebsen center for cancer immunotherapy, institute of clinical medicine, university of oslo, oslo, norway; department of haematology and oncology, university hospital of the goethe university, frankfurt, germany and childrens hospital, goethe university, frankfurt, germany natural killer (nk) cells are lymphocytes of the innate immunity with a potent anti-tumor capacity. in tumor patients, such as multiple myeloma (mm) patients, an elevated number of nk cells correlates with a higher overall survival (os) rate. our study adressed nk cells characteristics and anti-tumor ability in mm patients. especially cytotoxicity of patientderived, cytokine-stimulated nk cells against mm cells has been analyzed at various time points (at diagnosis, before/ after chemotherapy and/or auto-sct). nk cells from patients were analyzed by facs after pbmcs isolation via ficoll separation at different time points: tp , before the start of high dose chemotherapy (hdc)/auto-sct; tp , after early leukocyte recovery (leukocytes /μl) and tp : at least weeks after tp . for testing nk cell cytotoxicity against mm cells, nk cells were purified via negative selection and expanded in vitro for - weeks in low doses il- and il- . nk cells were divided into the cd ++ cd − or cd + and cd + cd ++ subsets. while the major nk cell subset at tp was the cd + cd ++ nk cell subpopulation ( . %), after leukocyte recovery at tp cd ++ cd − /+ nk cells were the main subsets (cd − : . %; cd + : . %). we further evaluated the nk cell function upon tumor interaction at the defined time points. cd ++ cd − nk cells were the main subset to produce ifn-γ upon interaction with k cells at all different time points. the percentage of ifn-γ-positive cd + + cd − nk cells was slightly decreased at tp compared to tp but significantly increased from tp to tp (p-value: . ). similarly, mip- β-and cd a-positive cd ++ cd − cells remained constant between tp and tp , whereas their percentages increased from tp to tp [p-values: . (mip β) and . (cd a)]. moreover, in a small group of mm patients, we isolated nk cells and expanded them for - weeks prior to the functional assays. as expected, the expansion rate was reduced after chemotherapy compared to nk cells from healthy controls, but the patients nk cells increased their ability to kill mm cells due to the ex vivo cytokine expansion. conclusion: our data demonstrate that nk cells have an altered phenotype and function after hdc/auto-sct. remarkably, these nk cells were able to secrete cytokines and still displayed cytotoxic capacity against different types of tumor cells. however, as the proliferative capacity of nk cells seemed to be reduced following chemotherapy, innovative nk cell therapeutic approaches further improve the patients nk cell activity by an ex vivo cytokine stimulation procedure. finally, we suggest that an additive cell therapy with cytokinestimulated autologeous nk cells might improve the outcome of mm patients. lymphoid and myeloid acute leukemia are the most frequent type of cancer and the most frequent cause of cancer related death in children. relapse and refractory disease are the main clinical problems that current therapies are still unable to solve. one of the main nk cell activating receptors is nk cell group d (nkg d). nkg d receptor recognizes human mica/ulbp - ligands. these nkg d ligands are expressed in leukemia cells and constitute suitable targets for immunotherapy. the expression of nkg d ligands was analyzed in peripheral blood mononuclear cells from pediatric patients suffering from acute leukemia ( acute myeloid leukemia, b cell acute lymphoid leukemia and t cell acute lymphoid leukemia), as well as in leukemia cell lines (k , rs - , jurkat, nalm- , molt- , reh and cem), by flow cytometry using specific monoclonal antibodies directed against mica, micab, ulbp- , ulbp- , ulbp- and ulbp- , and by quantitative pcr using taqman probes. peripheral blood mononuclear cells from healthy donors were labeled with cd ra microbeads and depleted using automacs device. the hl i r-mndanticd bbz lentiviral vector was derived from the clinical vector cl i r-ef a-hgcopt but contained the extracellular domain of nkg d, the hinge region of cd a and the signaling domains of - bb and cd -z. the cassette was driven by mnd promoter. viral supernatant was produced by transient transfection of hek t cells with the vector genome plasmid and lentiviral packaging helper plasmids pcagg-hivgpco, pcagg-vsvg and pcag -rtr . cytogenetic studies and array comparative genomic hybridization were performed to analyze the genetic stability of lentiviral-transduced memory t cells. the in vitro cytotoxicity of cd ra − t cells against leukemia cells, healthy pbmc and mesenchymal stem cells (msc) was evaluated by performing conventional -hour europium-tda release assays or by flow cytometry using cfse and aad labeling of target cells. nkg dl were heterogeneously expressed in leukemia primary cells and cell lines. for b cell all primary samples, we found expression of mica/b, mica and ulbp decreased in refractory disease compared to remission (p = . , p = . and p = . , respectively). lentiviral transduction of nkg d- - bb-cd z markedly increased nkg d surface expression in cd ra − memory t cells, which became consistently more cytotoxic than untransduced cells against leukemia cells. additionally, no chromosomal aberrations nor cytotoxic activity against healthy pbmc or mesenchymal stem cells was observed in nkg d car expressing t cells. our results demonstrate nkg d-car redirected cd ramemory t cells target nkg dl expressing leukemia cells in vitro and could be a promising and safe immunotherapeutic approach for acute leukemia patients. peripheral blood stem cell mobilization and collection from elderly patients (≥ years) with multiple myeloma: a single center experience g cengiz seval , sk toprak , s civriz bozdag , m kurt yuksel , p topcuoglu , o arslan , m ozcan , t demirer , g gurman , h akan , m beksac and o ilhan clinic of hematology, yildirim beyazit university yenimahalle education and research hospital and department of hematology, ankara university school of medicine high-dose melphalan followed by autologous hematopoietic cell transplantation (auto hsct) has become the standard procedure for patients with symptomatic multiple myeloma (mm). the ability to mobilize stem cells from healthy donors shows little deterioration with age, the influence of patients' age on auto hsct is uncertain and studies in patients' ≥ years are scarce. severe studies specific to mm have failed to show an independent effect of patient age on cd + mobilization. we retrospectively compared myeloma patients below the age of with patients above years of age, analyzing cd mobilization into peripheral blood and the number of leukapheresis needed to collect at least one single stem cell graft. material and methods: from february through april , data from myeloma patients below the age of were compared to myeloma patients above years of age. all these data were obtained from the ankara university faculty of medicine center for therapeutic apheresis and written informed consent was signed according to our institution regulations. most of the patients received only gcsf at a dose of μg/kg bw twice-daily s.c. until stem cell procurement. patients underwent further pbsc collections until we obtained the target dose cd + cells/μl blood. a maximum of collections were performed in the first mobilization; if the cell dose was not achieved, we submitted patients to a second mobilization. fifty two of patients were above years of age (median age , range: - ) and patients were below the age of (median age , range: - ). baseline characteristics of the older and younger patient cohorts are summarized in table . mobilization regimens for the younger patient population were cyclophosphamide based (n: ), g-csf only (n: ) and +plerixafor (n: ). mobilization in the older population was with cyclophosphamide based (n: ), g-csf only (n: ) and +plerixafor (n = ). the chemotherapy regimens were not statistically different between both age groups. there were no significant statistical differences in time from diagnosis to mobilization, number of prior therapies or disease status between both patient groups. the number of cd + circulating cells before scheduled leukapheresis was mean . cells/μl (median cells/ μl, range: - ; sem ± . ) in all patients (including patients who failed mobilization). our data support the observation that after a standard mobilization regimen with anti-myeloma chemotherapy and once-daily growth factor support, patients above years of age show an impaired cd mobilization into peripheral blood compared to a younger population. this can be overcome by an increased number of leukaphereses. still the number of progenitor cells in the actual graft is inferior compared to the younger population. [p ] disclosure of conflict of interest: none. donor and/or recipient citomegalovirus (cmv) seropositivity has been associated with a poor overall survival (os) in patients who have received an allogeneic hematopoietic stem cell transplantation (allohsct). in comparison with seronegative donors, hsct from seropositive donors has been associated with decreased disease-free survival (dfs) and increased non-relapse-related mortality (nrm). we analyzed the prognostic impact of cmv serology status (donor/ recipient) in patients diagnosed with acute leukemia (al) [p ] s who had received an allohsct in our institution. retrospective unicentric study of patients diagnosed with al between and who received allohsct.the following outcomes were studies: os, dfs, and cumulated incidences of relapse (ri), nrm, acute graft-versus host disease (agvhd) and chronic gvhd (cgvhd). the series included patients ( males, females), median age of years . al type: ( %) all, ( %) aml. type of transplant: ( %) related donor, ( %) unrelated donor and ( %) unrelated umbilical cord blood. the majority, ( %), received myeloablative conditioning. stem cells source: peripheral blood ( %), cord blood ( %) and bone marrow ( %). cmv serology status: positive receptor ( %), negative receptor cases ( %); positive donor ( %), negative donor ( %). serology status combinations (d/r): +/+ ( %), +/ − ( %), − / − ( %), − /+ ( %). patients developed agvhd and ( %) cgvhd. the impact of donor/recipient cmv serology status on os, dfs, ri, nrm and incidence of agvhd and cgvhd for the overall series is reported in table . no statistically significant differences were detected in any of the analyzed variables. in this study, donor/recipient cmv serology showed no influence on the analyzed variables os, dfs, al relapse, nrm, acute and chronic gvhd. however, the sample size limits the validity of the results. disclosure of conflict of interest: none. supported in part with the grants pi / from fondo de investigaciones sanitarias and rd / / from rticc, instituto carlos iii and sgr (gre), generalitat de catalunya, spain. petersburg, russia during the last two decades ahsct has been used as a treatment option for ms with promising outcomes. qol is an important outcome of ms treatment. its assessment gives the patient's perspective on the overall effect of treatment. we aimed to study qol in ms patients before and after ahsct and search the value of the data obtained for decision-making. a total of patients with different types of ms were enrolled in the study: mean age- (range- - ) years old; male/ female- / ; mean edss- . (range: . - . ). all patients were treated by ahsct. reduced-intensity beam-like conditioning was used (bcnu mg/m , etoposide mg/m , ara-c mg/m and melphalan mg/m ). mean follow-up was months (range: - months). qol was assessed using generic questionnaire sf- . for comparisons t-test for independent samples or mann-whitney test was used. qol parameters in ms patients at months after ahsct improved in comparison to base-line: physical functioning- . vs . , role-physical functioning- . . further qol improvement was registered at long-term follow-up: integral qol index exhibited . at long-term follow-up as compared to . at base-line. qol improvement was more dramatic in relapsing-remitting ms than in progressive ms. we found a significant increase of all eight sf- scales in a year posttransplant as compared with base-line in relapsing-remitting ms patients (po . ). in progressive ms patients statistically significant improvement was registered for six out of eight sf- scales (except bodily pain and role-emotional functioning) (p o . ). improved qol parameters were preserved over the entire study period in all the patients who did not have disease progression or relapse. in conclusion, qol monitoring in ms patients after ahsct provides clinicians with the unique information regarding the changes in physical, psychological and social well-being of patients who have been treated with this new treatment modality. it allows to evaluate risks/ benefits of ms patients undergoing ahsct and might influence decision-making. further studies are needed to examine the trajectory of qol changes in this patient population to better define treatment outcomes after ahsct. disclosure of conflict of interest: none. pediatric patients with leukocyte adhesion deficiency type-i (lad-i), a rare autosomal recessive primary immunodeficiency disorder, experience severe and recurrent lifethreatening bacterial infections. allogeneic haematopoietic stem cell transplantation (hsct) offers the possibility of curative therapy although the conditioning regimen used for hsct in lad-i is still a controversial issue. this study provides evaluation of outcome of the lad-i pediatric patients who underwent reduced-intensity conditioning (ric) hsct. twenty four patients ( female) with severe lad-i who received hscts between februay and september at our center were enrolled. the median age at hsct was months (range: months- years). patients received bone marrow (n = ), peripheral blood progenitor cells (n = ) or umbilical cord blood grafts (n = ) from hla-matched related donors (n = ), mismatched related or unrelated donors (n = ), unrelated fully matched donors (n = ) and haploidentical relative donors (n = ). ric regimen was provided with fludarabine, melphalan and anti-thymocyte immunoglobulin. cyclosporine a and prednisolon were used as graft-versus-host disease (gvhd) prophylaxis. engraftment occurred in / , of which one patient experienced graft rejection.the median times to neutrophil and platelet engraftments were days (range: - days) and days (range: - days), respectively. with a median follow-up of months (range: - months), overall survival (os) was . %.the main causes of death were gvhd and infection. acute gvhd occurred in ten patients ( grade i-ii, grade iii-iv) and patients also developed chronic gvhd. there were no significant differences in acute gvhd occurence and also os regarding to the stem cell sources. at this time, patients with full chimerism and patients with mixed chimerism are alive and disease free. conclusion: hsct offers long term benefit in lad- and should be considered as an early therapeutic option if a suitable hla-matched stem cell donation is available. as pretransplant infections in primary immunodeficient patients especially those affected by lad- lead to rise in mortality rate, ric regimen is found to be safe and mixed donor chimersim appears sufficient to prevent significant symptoms. disclosure of conflict of interest: none. tregs based immunotherapy may be beneficial in several immune mediated diseases including graft versus host disease (gvhd). the possibility of cryopreserving tregs might lead to the administration of multiple doses, thus potentially increasing their efficacy in chronic diseases. however, there are few and controversial data on the functionality of tregs after cryopreservation. here, we evaluated the phenotype and the inhibitory capacity of thawed tregs. tregs were purified from leukapheresis of normal donors (n = ) by double immunomagnetic depletion (cd and cd ) followed by cd enrichment using the clinimacs system (miltenyi biotec) under gmp condition. the cells were cryopreserved in saline solution containing % human serum albumin (hsa) and % dmso with a controlled-rate freezing. cell viability was assessed by -aad staining. number/phenotype and function were evaluated on fresh and thawed tregs. cryopreserved autologous t effector (teff) cells were used in mlr assays. before cryopreservation the tregs enriched product mean viability was ± % and the mean percentage of cd +cd +cd +cd low and cd +cd +cd +cd lowfoxp + cells was ± % and ± %, respectively. we then analysed the tregs enriched product after thawing. mean viability of thawed tregs, by -aad staining, was ± %. the viable tregs were almost totally cd +cd + ( ± %). the mean percentage of cd +cd +cd low and cd +cd +cd lowfoxp + thawed cells was ± % and ± % respectively. the contaminant cells present in the treg enriched product were mostly cd +cd +cd + (around %). we further characterized the phenotype of the cd +cd +cd low population. this population was almost totally foxp + ( ± %) and expressed selected markers at various degree (cd l ( ± %), cd s ( ± %), cd ra+ ( ± %), hla-dr+ ( ± %), ccr + ( ± %), cd d ( ± %), cd + ( ± . %), cd +cd + ( ± %). notably, viable thawed tregs were able to induce inhibition of autologous teff cells in a : tregs:teff ratio as freshly isolated tregs: ± % (thawed) vs ± % (fresh) of inhibiton (p . ). in conclusion, here we demonstrated that thawed tregs from healthy donors mantain a stable phenotype. in addition, in our hands tregs show good suppressive ability after thawing despite lower expression of cd l and cd s relapsed and refractory malignant b cell diseases: evidence for therapeutic efficacy via subcutaneous administration of anti-cd × anti-cd antibody lymphomun r buhmann, p ruf, j hess, h lindhofer, u jacob and m dreyling the trifunctional antibody anti-cd × anti-cd lymphomun represents a chimeric immunoglobulin scaffold (mouse igg a/ rat igg b) with promising treatment outcome in patients suffering from malignant b cell diseases. by changing the lymphomun administration route from intravenous (i.v.) to subcutaneous (s.c.) the proinflammatory cytokine-mediated side effects were considerably slighter and generally welltolerated. most importantly, s.c. lymphomun showed outstanding responses in b cell depletion even in the absence of elevated cytokine levels (e.g. il- ) that are required for cytotoxic t cell activation. in summary, the clinical tolerability of s.c. lymphomun may result in a considerable improvement of the subjective well-being and in enhanced mobility due to decreased pain symptomatology. intestinal microbiota disruption is associated with acute gastrointestinal (gi) gvhd and inferior outcome in patients after allogeneic stem cell transplantation (asct). the wide use of systemic broad spectrum antibiotics adds a further risk factor contributing to major microbiota shifts. here, in a retrospective analysis of patients undergoing asct at the regensburg university medical center we assessed the relative expression of paneth cell antimicrobial peptides (amps) in human intestinal biopsies in relation to acute gi gvhd and systemic antibiotic treatment. the relative expression of paneth cell amps was significantly higher in biopsies of the upper gi tract than in the lower gi tract for reg α (p ≤ . ), human defensin (hd) (p ≤ . ) and hd (p ≤ . ). regarding the distribution of paneth cell amps in the gi tract we observed significantly higher expressions of all three paneth cell amps in the duodenum, jejunum and ileum compared to the stomach, colon and rectum (po . , figure ). in the presence of acute gi gvhd, paneth cell amps reacted contrarily in the upper and lower gi tract: we observed a decrease of hd , hd and reg α in the upper gi tract (p ≤ . ), similarly paneth cell count dropped in case of severe gi gvhd stage - (po . ). however in the lower gi tract severe acute gi gvhd was associated with an increase of paneth cell amps (p ≤ . ). initiation of additional systemic antibiotic treatment prior to day after asct correlated with a significantly higher expression of hd (p = . ) and reg α (p = . ) in intestinal biopsies compared to patients without or with initiation of systemic antibiosis after day . however, no significant differences were found in terms of hd expression in intestinal biopsies and start of systemic antibiotic therapy. the expressions of hd , hd and reg α in intestinal biopsies seem to respond to major microbiota disruptions caused by acute gi gvhd or systemic antibiotic treatment. while observations in the upper gi tract seem to reflect paneth cell damage, the relative increase in the lower gi tract may indicate inflammatory induction of amps in colonic epithelial cells in the course of gvhd. [p ] disclosure of conflict of interest: none. patients ( %) were in complete remission at the time of pcy haplo-sct. hematopoietic cell transplantation-comorbidity index was ≥ in patients ( %). thirteen patients ( %) received non-myeloablative conditioning regimen (as baltimore schema, luznik et al. bbmt ) prior to haplosct while remaining patients received busulfan-based regimen. all patients were given pcy and both csa and mmf as gvhd prophylaxis. day+ cumulative incidence of grade to and to acute gvhd was % and %. -year cumulative incidence of chronic gvhd was %. the cumulative incidence of non-relapse mortality and relapse at years were % and %, respectively. with a median follow up of months (range: - ), -year progression-free and overall survivals were % and %, respectively. disease status at the time of haplosct was a major determinant for outcome. indeed, year nrm and os were % and % in patients transplanted with active disease, respectively, while corresponding values in patients transplanted in cr were % (p = . ) and % (p = . ), respectively ( figure a and b) . we can conclude that in selected patients who could be candidate for second transplantation, haplosct is feasible and may represent a curative option. the overall incidence of relapse of % is promising in this situation for which no alternative for cure is available, with relatively good survival in patients transplanted in cr. however, the very high nrm ( %) in refractory patients should make us consider second transplant with caution in this setting. for these patients, specific developments are needed to avoid procedure-related toxicity. [p ] disclosure of conflict of interest: none. secondary solid tumors following hematopoietic cell transplantation for thalassemia major a natale, s santarone, a meloni, a pepe, m di ianni, s angelini, p di bartolomeo dipartimento di ematologia, medicina trasfusionale e biotecnologie-ospedale civile, pescara, italy secondary solid tumors (sst) have been described after hct, in particular for patients affected by hematologic malignancies. there is limited information about the incidence of sst following hct for thalassemia major (tm). the aim of this study was to determine the incidence of sst in patients with tm who received hct in our center between and . patients survived more than years after hct and were enrolled in the study. of them, were males and females. their median age at time of hct was years ( - ). as conditioning regimen, they received busulfan ( mg/kg) and cyclophosphamide ( mg/kg). the gvhd prophylaxis included cyclosporine and methotrexate. all patients received bone marrow cells from an hla identical donor. at time of this report, patients were cured, whereas patients rejected their graft and are now under regular transfusion treatment. overall, the median follow-up after hct was years ( - ). seven patients developed a malignancy . to years (median . years) after hct including carcinomas of the tongue, oral squamous cell carcinoma, colorectal cancer, thyroid carcinoma, carcinoma of the uterine cervix, and parotid carcinoma. the -yr cumulative incidence (ci) of developing sst was + . %. all patients underwent surgical resection of the tumor and in addition of them received chemotherapy and/or radiotherapy. of relevance, the patients with cancer of the oral cavity were affected by severe chronic gvhd with buccal cavity involvement. patients ( with parotid and with tongue carcinoma) died of tumor progression and are living. we compared these results with case control populations. first of all, we investigated the occurrence of solid tumors in the individuals ( males, median age years at time of marrow donation), who served as stem cell donors for hct. one donor developed breast cancer years after marrow donation at age of . the -yr ci of developing solid tumor for donors was . + . % with a statistically significant difference (p = . ) as compared to that of transplanted patients. the second case control population consisted of patients affected by tm treated with transfusions and iron chelation. the matching technique applied was based on the variables age and sex. one control per case (transplanted patient) was randomly selected from the miot (myocardial iron overload in thalassemia) registry and matched by sex and age with the transplanted patient population. patients developed an hepatocellular carcinoma (hcc) at age of and years, respectively. one patient died and one is living. using the event rate measure, we observed an event rate of . at years for the transplant group and . for the nontransplant group (p = . ). this study shows that the magnitude of increased risk of sst is twofold to threefold for patients treated with hct as compared with an age-and sex matched nontransplant tm patients or with stem cell donors. notably, among the transplanted patients we didn't observe any case of hcc, which is one of the most frequent solid tumor in nontransplant tm patients, whereas we observed cases of head/neck cancers. in our series, cgvhd seems to be a strong risk factor in the development of new solid tumors. patients with cgvhd, especially those with involvement of the oral cavity, must receive a very long careful monitoring and surveillance in order to prevent the development of secondary cancers. disclosure of conflict of interest: none. sequential treatment with bortezomib plus thalidomide plus dexamethasone followed by autologous hematopoietic stem cell transplantation (hsct); consolidation and maintenance therapy in patients with multiple myeloma a bachiri , ma bekadja , s talhi , s abderrahmani , h ouldjeriouat and r bouhass department of hematology, hmru oran, oran, algeria; department of hematology and cell therapy, ehu st november, oran, algeria and department of hematology and cell therapy, oran, algeria the management of multiple myeloma (mm) has been significantly improved in recent years in young patients, where ahsct and advent of new molecules was introduced as first line treatment. the sequential treatment (induction followed by autologous hematopoietic stem cell transplantation; consolidation and maintenance therapy) has increased rates response (cr and vgpr) as well as the overall survival. our purpose was to assess the efficacy and adverse effects of sequential treatment with vtd chemotherapy and autologous hsct followed by consolidation and maintenance therapy. in a prospective multicenter study, we evaluated this mm management strategy at oran, in two hematology centers. patients aged under years with de novo mm, were treated with induction included: bortezomib ( . mg/m , d -d -d -d ), thalidomide ( mg/ m d -d ) and dexamethasone ( mg, d -d ; d -d ). a total of to cycles where delivered every days. autologous stem cell was mobilized using g-csf alone ( μg/kg/day for days). leukapheresis to harvest stem cells were performed on day - and - . the conditioning regimen consisted of melphalan mg/m . a consolidation phase was initiated two months later with the same protocol (vtd), followed by a maintenance treatment with thalidomide mg/day given orally for months. this study was done over a -years period (january -december ). fifty patients were included. they include women and men (sex ratio = . ). the median age at diagnosis was years ( - ). according to durie salmon staging, % of patients were in stage iii, while % were in stages iii according to iss staging. the monoclonal component was igg in % of patients. postinduction overall response rate in the eligible patients was %, including % vgpr and % cr/ and % pr rates. the median of cd + rate was . x /kg ( . to ). all patients had engraftment on the median of day (range; to ) and platelet transfusion independence on the median of day (range; to ). there was no graft failure. one patient died following the procedure (trm). posttransplantation on day , cr and at least vgpr remained significantly higher ( %). in the evaluable patients, the estimated os at months was %, the estimated dfs at months was % and the pfs at months was . %. at the / / , ( %) patients are alive and ( %) without disease activity after a median follow-up of months (range; - ). the main hematological toxicities post transpland (grade / ) were thrombocytopenia ( %), neutropenia ( %), and anemia ( %). the most frequently observed nonhematological toxicities (all grades) included peripheral neuropathy ( %). our experience suggests that the sequential protocol used in first line produce a better outcome with fewer adverse events and is an interesting therapeutic option in term of efficacy and tolerance. disclosure of conflict of interest: none. micrornas are small, non-coding single-stranded rnas and regulate approximately % of all genes by repressing translation. they are present in bodily fluids, where they are protected from rnase-mediated degradation by encapsulation into extracellular vesicles (evs) and demonstrate a novel capacity to regulate the cellular differentiation of blood cells and immune function. candidate micrornas mir- , mir-- , mir- * and mir- have previously been associated with acute graft versus host disease (agvhd) in posthematopoietic stem cell transplant (hsct) patient plasma. however, validation in independent cohorts is necessary, and their presence within extracellular vesicles (evs) has not been explored. microrna expression was evaluated in a prognostic cohort (n = ) of day (d ) post-hsct patient serum samples by taqman qrt-pcr. further assessment in an independent cohort of serum samples taken at the time of agvhd diagnosis was also performed. expression was also assessed in serum evs at sequential time points (pre-hsct, d , d and d ) and an independent verification cohort of d serum samples by ev isolation, rna extraction and taqman qrt-pcr analysis. this study replicated elevated serum expression of mir- (po . ), mir- (p = . ), mir- * (p o . ) and mir- (p = . ) in agvhd, in a prognostic cohort of d post-hsct patient samples (n = ). expression was also associated with disease severity. further analysis at agvhd diagnosis in an independent cohort (n = ) confirmed high expression of mir- (p = . ), mir- (p = . ) and mir- * (p = . ) at disease onset. investigation of microrna expression patterns during early hsct at sequential time points (pre-hsct to d ) identified elevated micrornas at d post-hsct in all transplant patients. in a novel investigation of microrna expression in serum evs (n = ), mir- (p = . ), mir- (p = . ) and mir- * (p = . ) levels were lower at d in patients who later developed agvhd, and this was replicated for mir- (p = . ) and mir- (p = . ) (n = ). comparing serum to circulating evs, at d patients remaining agvhd-free had significantly higher expression of mir- (p = . ), mir- (p o . ) and mir- * (p = . ) in the ev fraction. results validate the capacity for circulating serum mir- , mir- and mir- * to act as diagnostic and prognostic biomarkers for agvhd. novel findings of differential expression between whole serum and the ev compartment prior to disease onset suggest a role for ev micrornas in the biology of agvhd, which warrants further investigation. disclosure of conflict of interest: none. prior data indicate similar outcomes after transplants from hla-haplotype-matched relatives, hla-idntical siblings and hla-matched unrelated donors. we used our dataset to answer a clinically important question: who is the best donor for a person with acute leukemia. we analyzed data from persons with acute leukaemia in st complete remission treated in a prospective, multi-centre study. patients were randomly divided into training (n = ) and validation (n = ) sets. consecutive subjects received a transplant from an hla-haplotype-matched relative (n = ) or an hlaidentical sibling (n = ). -year leukaemia-free survivals (lfss) were % ( % confidence interval [ci], , %) and % ( , %; p = . ). the multivariate model identified major risk factors for transplant-related-mortality (trm): older donor/recipient age (donor years/recipient years; hazard ratio [hr] = . ; [ . , . ]; p = . ), female-to-male transplants (hr = . ; [ . , . ; p = . ) and donor-recipient abo major-mismatch transplants (hr = . [ . , . ; p = . ). a risk score was developed based on these three features. trms were % ( , %), % ( , %) and % ( , %) for subjects with scores of - , and (p o . ). year lfs were % ( , %), % ( , %), and % ( , %; p = . ). the risk score was validated in an independent cohort. in recipients years, lfss were % and % (p = . ) after transplants from identical-sibling or children. our data confirm donor source or degree of hla-disparity is not significantly correlated with transplant outcomes. selection of the best donor needs to consider donor-recipient age, sex-matching and abo-incompatibility amongst persons with acute leukemia receiving transplants from family members. [p ] disclosure of conflict of interest: none. synergistic effect of kir ligands missing and cytomegalovirus reactivation in improving outcomes of haematopoietic stem cell transplantation for treatment of myeloid malignancies d cardozo, a marangon, r da silva, fj aranha, j visentainer, s bonon, s costa, e miranda, c souza and f guimarães. the lack of one or more hla class i alleles, whose protein products are the ligands for kir receptors, has been exploited as a prognostic factor for the outcome of patients with haematological malignancies treated by haematopoietic stem cell transplantation (hsct). although it has been accepted that kir-hla interactions may influence the outcome of the hlamismatched hsct, there is no consensus regarding the settings of hla-matched transplantation. there are studies that have reported either benefits, or no effects, under the influence of inhibitory kir-hla interactions. additionally, certain activating kirs and/or reactivation of cytomegalovirus (cmv) infection have been reported to affect the outcome of hla-matched transplantation. the goal of this study was to evaluate the influence of kir-hla genotypes on the outcome of patients undergoing treatment for haematological malignancies by non-t-depleted lymphocyte haematopoietic stem cell transplantation (hsct) from hla-matched sibling donors. the prospective study was conducted at the center of hematology, university of campinas, and patients and their donors were followed up from to . kir and hla class i genes were genotyped and patients grouped based on the presence of kir ligands combined with kir genotype of their respective donors. patients with all kir ligands present (n = ) had a significantly higher (p = . ) incidence of acute graft-versus-host-disease (gvhd) than patients with one or more kir ligands missing (n = ). the overall survival following transplantation of patients with myeloid malignancies (n = ) was significantly higher (p = . ) in the group with one or more kir ligands missing (n = ) than in the group with all ligands present (n = ). presence of kir ds was associated with a worsening of hsct outcome while reactivation of cytomegalovirus (cmv) infection improved the outcome of patients with one or more kir ligands missing. our results indicate that kir-hla interactions affect the outcome of the hla-matched transplantation, particularly in patients with myeloid malignancies. disclosure of conflict of interest: none. p = . ), lower disease-free survival (p = . and p = . ) and lower overall survival (p = . and p = . ). one-year cir of the above two groups were . ± . % vs. . ± . % in mrd negative and positive patients, respectively (p = . ). in addition, those who had consistent positive mrd prior to hlamatched sibling hsct showed even worse outcomes compared to patients without pre-mrd. unmanipulated haploidentical hsct might have the stronger graft-versus-leukemia effect compared to hla-matched sibling hsct. it suggested that those who received unmanipulated haploidentical hsct with pre-mrd might not need more intensive relapse intervention after transplantation. disclosure of conflict of interest: none. the retrospective study of allogeneic hematopoietic cell transplantation for patients with mixed-phenotype acute leukemia in toranomon hospital, japan in the real clinical setting, however, there are substantial number of patients who can not achieve cr after chemotherapy. we conducted a retrospective study including such patients to elucidate the outcome of allogeneic hct in toranomon hospital, japan. we studied the patients with mpal diagnosed from july to september . mpal was diagnosed according to who classification in . from june , we examined cytoplasmic myelo-peroxydase (cmpo) routinely for flowcytometric analysis in all the patients, to distinguish mpal from acute lymphoblastic leukemia (all). we included the patients who were diagnosed as mpal in toranomon hospital, regardless of their diagnosis or clinical course in the previous hospitals. a total of mpal patients underwent their first allogeneic hct with related bone marrow or peripheral blood stem cells (r-bm/pb) (n = ), unrelated bone marrow (u-bm) (n = ), and unrelated umbilical cord blood (u-cb)(n = ). the median patient age was years (range: - ). the immunophenotype of leukemia cells included cases of b and myeloid (b/my) ( %) and cases of t and myeloid (t/my) cell lineage( %).eleven patients( %) harbored philadelphia chromosome. the remission induction chemotherapy was performed with all-type regimens in patients, and acute myeloid leukemia (aml)type regimens in of patients, patients( %) were not in cr at the time of transplantation. myeloablative conditioning (mac) regimens were used in pantients( %). the -year overall survival (os) rate was . % ( % confidence interval (ci), . - . %). to identify the factors that influenced os, we performed univariate analysis and compared the following pre-transplantation factors: age at the time of transplantation ( o vs. = years), committed immunophenotype (b/my vs.t/my), karyotype (philadelphia chromosome (ph vs.non-ph), disease status at the time of transplantation (cr vs.non-cr), donor cell source (r-bm/pb vs.u-bm vs.u-cb, cb vs.non-cb), and conditioning regimen (mac vs.reduced intensity conditioning). cr at the time of transplantation was extracted as a significant predictive factor for the better os( -year os; cr vs. non-cr, . % ( % ci, . - . %) vs. . % ( % ci, . - . %), p = . ). the cumulative incidence of relapse rate (rr) at years after transplantation was . % ( % ci, . - . %). to identify the factors that influenced relapse rate, we performed univariate analysis and compared pretransplantation factors same as above. harboring philadelphia chromosome was extracted as a significant predictive factor for lower relapse rate ( -year rr; ph vs.non-ph, . %( % ci, . - . %) vs. . %( % ci, . - . %), p = . ). the older patients(p = . ) and the patients in cr (p = . ) also showed a trend towards lower relapse rate. allogeneic hct provided . % of -year os for mpal patients in cr at the time of transplantation. on the other hand, for patients not in cr, year os was approximately %. the use of tyrosine kinase inhibitors along with chemotherapy before transplantation might prevent relapse after transplantation in mpal patients with ph chromosome. disclosure of conflict of interest: none. allogeneic hematopoietic stem cell transplantation (allo-hsct) is a standard of treatment for many patients with hematological malignancies. however, the disease relapse and graft failure after first allo-hsct ( st allo-hsct) lead to poor outcomes almost in all cases. second allo-hsct ( nd allo-hsct) is one of primary options that can decrease the mortality in this group of patients. here we report our experience of patients who underwent nd allo-hsct. the aim of the study was to estimate a clinical efficiency and practicability of nd allo-hsct. we included patients ( males/ females) with acute myeloid leukemia (aml, n = ), acute lymphoblastic leukemia (all, n = ) and myeloproliferative disease (mpd, n = ) who underwent nd allo-hsct for relapse ( , %) or graft failure ( , %) from the same (n = ) or another donor (n = ) between november and october . median age was years (range: - years). three ( %) patients had a matched related donor (mrd), nine ( %) patients had a matched unrelated donor (mud) and three ( %) patients had a mismatched unrelated (mmud) at the second transplant. to evaluate time gap affecting outcomes all patients were divided into two groups: who underwent nd allo-hsct in more/less than months after st allo-hsct. in "less than months" group three patients were re-transplanted for relapse and one-for graft failure, in other group there were seven patients who received nd allo-hsct for disease relapse and four-for graft failure. fisher's exact test were performed to exclude probability of imbalance between groups (p . ). median of overall survival (os) and disease-free survival (dfs) after nd allo-hsct was . months and . months respectively. (see figure a , c) two patients ( . %) developed graft failure and three relapsed ( %). acute graft-versus host disease (agvhd) incidence was extremely low as . % (n = ) even despite use of mud/mmud in % of cases. mortality rate were . % in a group of nd allo-hsct. it should be noted that only ( %) patients died because of disease progression. five patients ( . %) died in complete remission due to severe infections or previous toxicity (e.g. heart failure). the effect of donor change on dfs was not significant (p = , ). our statistical analysis reveal significantly differences in os in patient with long-term interval ( months) between st and nd allo-hsct. median of os in patients who underwent nd allo-hsct in more/less than months after st allo-hsct was , vs , months respectively. (see figure b , d) for hazard ratio (hr) estimation mantel-haneszel approach were used hr for group who were transplanted in less than months from st allo-hsct was . , ( % ci, . s to . , p = . ). as for dfs difference was not significant (p = . ). according to our analysis, performing nd allo-hsct in a period less than months after st allo-hsct seemed not very reasonable due to extremely high mortality even in young patients (hr- . , p = . ). as for "more than months" group it can be considered even despite hla-disparity between donor-recipient pair due to extremely low agvhd rate ( . %). donor change was not associated with better outcome (p = . ) disclosure of conflict of interest: none. hemopoietic stem cell transplantation (hsct) is an effective treatment for many hematologic disorders, and globally over procedures/year are performed in more than countries. however, not all the countries have enough resources and expertise to establish an hsct program, and patients are often forced to emigrate for transplantation, with heavy social and economic consequences. in the year the iuc (an italian ngo) identified the hiwa cancer hospital (hch) in sulaymaniya (iraqi kurdistan) as a possible site for the establishment of a new hsct transplant center. a hsct expert from italy (mi) following a visit to the hch, reported a positive conclusion on the feasibility of an hsct project. this was mainly due to the fact that many of the required technologies were already available at hch, including a -bed positivepressure, hepa-filtered-air clinical unit, last-generation cell separators and a well equipped hla laboratory. following this preliminary survey, a capacity building project was rapidly made and submitted to the italian agency for development cooperation, that approved and funded it in march with the specific aim to cure thalassemia patients either of kurdistan and of the refugees population from syria and other parts of iraq. in april , the joint italian and kurdish team started the project. a first autologous transplant was done in june followed by more autografts (overall, myeloma and lymphoma patients). in october, following appropriate downstaging, a first low-risk thalassemia patient was allografted from her hla-id sibling, followed by more patients. all the patients engrafted promptly, with one death occurring on day + with acute cardiac failure and a major toxicity recorded in a single patient (nhl, severe enterocolitis with perforation) that was successfully treated. the full process for the start-up included the following activities developed during -month time: ( ) s of transplants, the hch group also submitted to ebmt an application for full membership, that was promptly approved. in all this project, the italian counterpart provided over highly-experienced volunteer specialists (physicians, nurses, technicians and one physicist), each with a specific mission plan. despite the many difficulties and obstacles encountered, the clinical results obtained so far appear encouraging, though there is still need to furtherly support the hch in order to make it totally independent. following this intervention, the hch is the only one center performing both auto and allo hsct not only in the iraqi kurdistan region, but also in all the iraqi nation. we conclude that international cooperation may be fruitful also in the field of high-technology medicine, and may contribute to improve the capabilities of centers even in critical geographic areas, representing a valuable instrument also to implement nation-to-nation scientific exchanges. disclosure of conflict of interest: none. the use of plerixafor with g-csf in conditioning regimen for hematopoietic stem cell transplantations with tcr alpha/beta and cd depletion of graft in wiscott-aldrich syndrome patients: a single-center experience b dmitry , l alexandra , s larisa , g elena , s irina , t pavel , k rimma , n galina , m michael and m alexei grade acute gvhd (agvhd) was % ( pts). no pt experienced a grade agvhd. three patients presented a limited form of chronic gvhd ( %). incidence of oral mucositis and gastrointestinal/liver toxicity has been extremely low in this population of patients, even in those with active disease and heavily treated at the time of transplant. eight out of fifteen pts ( %) are alive with a median follow-up of months (range: - m). seven ( %) are in cytogenetic/molecular remission. six out the eight patients who were transplanted in cr or cr are alive ( %), while two out the seven patients who were transplanted in advanced phase are alive ( %). in this preliminary clinical experience, we find that unmanipulated haploidentical transplants with post-transplant cyclophosphamide are a valid alternative and have outcome comparable to unrelated and match sibling transplants, in pts with hematologic malignancies. advanced disease is the only adverse factor for diseasefree survival. we therefore consider this therapeutic option when a match sibling or a / ag mud donor is not immediately available. disclosure of conflict of interest: none. autism spectrum disorder (asd) is a group of neurodevelopmental disorders characterized by impaired social communication and interactions with restricted and repetitive behaviors. although asd is suspected to have either heritable or sporadic genetic basis, its fundamental etiology and pathogenesis are poorly understood. recently researchers have suggested that stem cells have therapeutic potential for asd. wharton's jelly-derived msc (wj-msc) from third-party donors (tpd) have high proliferation and differentiation potential. this cell population has also non-immunogenic and immunomodulatory properties, thus seem to be a promising treatment stem cell source. the polish stem cell bank (pbkm) has provided wj-msc for clinical application in a medical therapeutic experiment for children with asd. twenty-three patients (pts) with asd aged from to . / (median age: years and months), after bioethical committee approval, received intravenous injections of wj-msc, obtained from tpd. the cells were previously collected from healthy newborns, then processed, screened for bacterial contamination as well as endotoxin content, and frozen in liquid nitrogen vapour. wj-msc immunophenotype was confirmed using flow cytometry assay. the pts received from to injections in intervals from to weeks. the average cell dose per infusion was . × ^ /kg of body weight (bw). each pt was examined by the same neurologist at the day of infusion. comorbidities present in some patients: unspecified speech disturbances, flaccid paralysis, flaccid tetraplegia, unspecified encephalopathy, epilepsy, sensorineural hearing loss. one patient was diagnosed with comorbidities: conductive hearing loss and intellectual disability. almost % of pts, after their treatment with wj-msc, revealed positive changes in neurological examination. an improvement in speech was observed in pts and improvement of cognitive functions ensued in pts. what is more, % of children showed progress in self-reliance, social interactions and improved their ability to concentrate. there was a reduction of aggressive behavior in pts and pts have experienced better quality of sleep. there was only one adverse event after wj-msc infusions -psychomotor agitation occurred in hours after the administration. five follow-ups have not yet been completed. the administration of thirdparty donor wj-msc seems to be safe and efficient procedure with promising preliminary results in patients with asd. hematopoietic stem cell transplantation between red cell incompatible donor-recipient pairs red blood cell depletion from bone marrow and peripheral blood buffy coat: a comparison of two new and three established technologies human bone marrow processing using a new continuous-flow cell separation device disclosure of conflict of interest: none. references . zama d, et al. gut microbiota and hematopoietic stem cell transplantation: where do we stand? bmt the kyoto encyclopedia of genes and genomes-kegg metabolites produced by commensal bacteria promote peripheral regulatory t-cell generation disclosure of conflict of interest: none antifungal prophylaxis in hematopoietic stem cell transplant recipients: the unfinished tale of imperfect success guidelines for preventing infectious complications among hematopoietic cell transplantation recipients: a global perspective differences in aspergillus-specific immune recovery between t-cell-replete and t-cell-depleted hematopoietic transplants toxoplasmosis following allogeneic hematopoietic stem cell transplantation diagnosis of toxoplasmosis after allogeneic stem cell transplantation: results of dna detection and serological techniques implementation of molecular surveillance after a cluster of fatal toxoplasmosis at neighboring transplant centers management of high blood pressure genes for blood pressure a prospective studyon the predictive value of plasma bk virus-dna load for hemorrhagic cystitis in pediatric patients after stem cell translantation cidofovir for bk virusassociated hemorrhagic cystitis:a retrospective study hemorrhagic cystitis after bone marrow transplantation bcsh/bsbmt guideline: diagnosis and management of veno-occlusive disease (sinusoidal obstruction syndrome) following haematopoietic stem cell transplantation drug safety evaluation of defibrotide defibrotide for prophylaxis of hepatic veno-occlusive disease in paediatric haemopoietic stem-cell transplantation: an open-label, phase , randomised controlled trial safety and effects of prophylactic defibrotide for sinusoidal obstruction syndrome in hematopoietic stem cell transplantation disclosure of conflict of interest: none university children's hospital basel, division of paediatric oncology/haematology late complications subcommittee of translated related complications and quality of life wp; clinic of paediatric haemato-oncology, department of women's and children's health, university of padova, italy; department of surgery, division of transplantation division of blood and marrow transplantation, the children's hospital at westmead ovarian function after bone marrow transplantation during childhood pregnancies following high-dose cyclophosphamide with or without high-dose busulfan or total-body irradiation and bone marrow transplantation unmanipulated haploidentical bone marrow transplantation and posttransplantation cyclophosphamide for hematologic malignancies after myeloablative conditioning haploidentical hematopoietic transplantation:current status and future perspectives t-cell replete haploidentical donor transplantation using post-transplant cy: an emerging standard-of-care option for patients who lack an hla-identical sibling donor hematopoietic stem cell transplantation in thalassemia major and sickle cell disease: indications and management recommendations from an international expert panel allogeneic stem cell transplantation for thalassemia major killer-cell immunoglobulin-like receptors reactivity and outcome of stem cell transplant kir b haplotype donors confer a reduced risk for relapse after haploidentical transplantation in children with all kir/hla interactions negatively affect rituximab-but not ga (obinutuzumab)-induced antibody-dependent cellular cytotoxicity reduction of minimal residual disease in pediatric b-lineage acute lymphoblastic leukemia by an fcoptimized cd antibody diagnoses: hodgkin's lymphoma(hl)- pts (refractory ; relapsed ); non-hodgkin's lymphoma(nhl disease status before asct: st (after refractority prior radiotherapy to the mediastinum - / ( . %); heavily pretreated patients with advanced disease (x lines previous treatment) / ( . %). grafts: pbsc - / pts with median of cd +cells- ccnu dose: / pts mg/m ; pt mg/m . engraftment: anc> cells/mkl: median=d+ ( ÷ ), / pts. plt> cells/ mkl: median=d+ ( ÷ ), / pts. full engrafted / pts / pt required a short-term mechanical ventilation ( of them died because of lung infection ad d+ and d+ ) aeruginosa associated sepsis on a background of graft failure); pts ( . %)-d+ and d+ (pulmonary toxicities +infection; both had prior mediastinal radiotherapy). relapse/ progression after asct- / pts ( . %), of them died. pt achieved secondary mds (diagnosed . mo after asct). for this group of pts with relapsed/refractory lymphomas (n= ) -year os= for nhl(n= ) efs= . (se ± . ) lomustine-containing conditioning regimen cem (lomustine, etoposide, melphalan) is effective and feasible in autologous stem cell transplant efficacy and toxicity of a ccnu-containing high-dose chemotherapy regimen followed by autologous hematopoietic cell transplantation in relapsed or refractory hodgkin's disease champlin re reduced-toxicity conditioning therapy with allogeneic stem cell transplantation for acute leukemia idarubicin-intensified bucy conditioning regimen improved survival in high-risk acute myeloid, but not lymphocytic leukemia patients undergoing allogeneic hematopoietic stem cell transplantation: a retrospective comparative study comparison of outcomes of idarubicin intensified tbi-cy and traditional tbi-cy conditioning regimen for high-risk acute lymphoblastic leukemia undergoing allogeneic hematopoietic stem cell transplantation: a single center experience inhibition of cd (il- r alpha) expression and t-cell proliferation by polyclonal anti-thymocyte globulins csf-primed bone marrow transplantation for patients with high-risk hematologic malignancies in an exploratory analysis, os after hct appeared to be longer in the cpx- arm in both age groups. these results suggest that cpx- may provide an effective bridge to successful transplant for a high-risk subgroup of aml patients. support: celator pharmaceuticals, inc., a subsidiary of jazz pharmaceuticals plc consulting ambit biosciences, amphivena therapeutics, ariad, astellas pharma sunesis, tolero; institutional research funding abbvie chiarella and louie: employment celator/jazz; stock jazz pharmaceuticals plc. hoering disclosure of conflict of interest: none. inkt-/nk-/cik-cell (subsets) are important for immunesurveillance. antibody b targets the vα -jα -invariant-t-cell-receptor (tcr) in the cdr -region, which is semiinvariantly rearranged in inkt-cells. we characterized: i.) inkt-/nk-/cik-subsets in pb-samples from healthy donors (n = subsets under stimulation with dendritic-cells of leukemic origin (dc leu ), generated from aml-blasts in mononuclear cells(mnc) and whole-blood (wb, containing soluble/cellular components of pts' pb) with 'cocktails' (dc-generating-methods/kits). . ) compared to healthy mnc (significantly) lower proportions of inkt-cells comparable correlations were seen in adultall-and cll-pts. . ) we quantified inkt-/nk-/cik-subsets before/after mixed-lymphocytecultures (mlc) of t-cell-enriched immune-reactive cells stimulated with mnc/wb (with or without pretreatment 'cocktails' inducing blasts' conversion to dc leu ) from aml-pts. our findings show, that )inkt-/nk-/cik-cells increase after mlc independent of the stimulator-cells-suspension (under the influence of il- ); ) pretreatment of mnc/wb-blasts with 'cocktails' increases inkt-counts and induces a shift in the composition of inkt-/nk-/cik-subsets after mlc, that might correlate with an improved antileukemic potential; ) individual samples showed varying, however higher inkt-, cik-cell-counts after pretreatment with different (especially prostaglandin-containing) 'cocktails'; ) dc-/inkt-/nk-/cikcells-values after mlc were comparable in physiological hypoxia vs normoxia; ) in cases with antileukemic blast-lytic activity after mlc t-/inkt-/nk-/cik-cells were significantly increased-pointing to an involvement of these cells in antileukemic reactions. in summary: ( ) healthy mnc present with significantly higher inkt-/nk-/cik-cells compared to aml/all/cll-leukemic mnc. ( ) subtypes of inkt-cells differ in healthy vs leukemic samples, resembling a shift in the composition of inkt-cells. ( ) amounts of inkt-/ nk-/cik-cells in aml/all/cll-mnc-samples correlate with prognosis. ( ) 'cocktail'-treated aml-blasts (resulting in dc leu ) lead to a shift in t-,inkt-/nk-/cik-cell-counts/compositions, what correlates with improved antileukemic activity against aml-blastspointing to a cross-talk of these cells. proportions of inkt-/ nk-/cik-cells management of philadelphia chromosome-positive acute lymphoblastic leukemia (ph+ all) outcome of allogeneic stem cell transplantation for aml and myelodysplastic syndrome in elderly patients (⩾ years) comorbidity-age index: a clinical measure of biologic age before allogeneic hematopoietic cell transplantation high rate of hematological responses to sorafenib in flt -itd acute myeloid leukemia relapsed after allogeneic hematopoietic stem cell transplantation phase i trial of maintenance sorafenib after allogeneic hematopoietic stem cell transplantation for fms-like tyrosine kinase internal tandem duplication acute myeloid leukemia haematopoietic cell transplantation with and without sorafenib maintenance for patients with flt -itd acute myeloid leukaemia in first complete remission quantitative monitoring of minimal residual disease (mrd) after sct was performed by four-colour flow cytometry and/or real time pcr. the median time of neutrophil engraftment (above . × e /l) was days, % of pts ( / ) engrafted, one patient died in aplazia. non-relapse mortality (nrm) after year and years was % ( / ) and % ( / ). causes of death were refractory gvhd (n = ), infection (n = ) and multiorgan failure (n = ). incidence of acute gvhd was evaluated in pts: % ( / ) of pts had gvhd (grade i+ii in pts, grade iii in pts). incidence of chronic gvhd was evaluated in pts, % ( / ) of pts had gvhd with median follow-up from sct months (range: - ), % of all pts ( / ) were alive ( in remission of cll with mrd negativity, with relapse), pts died ( from nrm, from cll relapse/progression), relapses ( %; / ) occurred. sequential use of chemotherapy and ric regimen with allogeneic sct is safety and effective treatment of high-risk cll with reponse rate % and low nrm. progression-free survival and overall survival at years from sct were % and % department of hematology, hemostasis, oncology and stem cell transplantation hannover deutsche klinik für diagnostik helios klinik wiesbaden, germany; imperial college london at hammersmith hospital du cane road centre for haematology london disclosure of conflict of interest: none. references . sibon d, brice p. optimal treatment for relapsing patients with at our institution, pr-hl is defined as partial response (pr), no response (nr), stable disease (sd), progressive disease (pd), relapsing within months of finishing the planned treatment. progression free (pfs) and overall survival (os) from the day of auto-sct was estimated by kaplan-meier (km) method. from to , patients with aethera trial criteria were identified. male ( %), female ( %), median age at diagnosis: yrs ( - ), at auto-sct: . yrs ( . - )( % o yrs). initial chemo: abvd in ( %). ( %) had radiation therapy (xrt) after initial chemo. response to initial chemo + xrt was refractory disease: ( %), relapse between - months: ( %) and relapse after months: ( %) aethera had % stable disease before sct vs we have only %. aethera months pfs ( % control arm, % brentuximab arm, investigator assessment) and our . % is not much different. despite having similar selection criteria, our median pfs is higher than both aethera trial placebo and experimental arm. clinically, rate of progression in both studies are very high and comparable at months. given the very high cost of this drug and while waiting for survival fifty-nine ( %) and ( %) patients had relapsed and primary refractory chemosensitive dlbcl, respectively. secondary ipi was - in ( %) patients, in ( %) patients and - in ( %) patients. fifty-one ( %) and ( % patients had gcb and abc tumors, respectively. abc patients received more prior lines of chemotherapy than gcb patients ( % vs % received lines of chemotherapy, p = . ). the rest of characteristics were equally distributed between both groups (table ) disclosure of conflict of interest: none disclosure of conflict of interest: none. p upfront autologous stem cell transplantation in patients with diffuse large b cell lymphoma: focused on risk factors for survival and conditioning regimens ds kim association between complete response and outcomes in transplant-eligible myeloma patients in the era of novel agents e jantunen and v varmavuo department of medicine disclosure of conflict of interest: none. and hematology department lenalidomide after stem-cell transplantation for multiple myeloma bortezomib induction and maintenance treatment in patients with newly diagnosed multiple myeloma: results of the randomized phase iii hovon- /gmmg-hd trial disclosure of conflict of interest: none we performed a retrospective study to investigate survival outcomes and toxicities of l maintenance therapy compared with b maintenance in mm patients post-ahct. this study included patients who received ahct for mm between and after induction with l-or b-based therapy. all patients received ahct within months of mm diagnosis and received melphalan mg/m conditioning. patients who received tandem transplantations (autologous or allogeneic) were excluded. only patients initiating maintenance therapy within months post-ahct were included. maintenance therapy was defined as monotherapy with either l or b. the primary outcome was pfs. secondary outcomes were overall survival (os) and treatment-related toxicities. patients received l maintenance and b maintenance post-ahct. at baseline there were no differences in iss stage, ds stage or cytogenetic risk between maintenance cohorts. at time of analysis, % (n = ) receiving l maintenance and % (n = ) on b maintenance experienced disease progression. median time to progression ( . vs . yrs, p = . ) was not significantly different between cohorts. by multivariable analysis, choice of maintenance (l vs b) was not significant for pfs or os. variables significant for improved pfs were iss stage i disease response improved while on maintenance in % (n = ) with l and % (n = ) with b. median os was not statistically different between maintenance cohorts ( . vs . yrs, p = . ). iss stage i/ii vs iii while cytopenias were more common in the l cohort ( % vs %, p o . ). the median follow-up time for survivors was months. these findings suggest that both lenalidomide and bortezomib are equivocal maintenance therapy options for post-transplantation mm patients. choice of maintenance therapy post-ahct for mm did not demonstrate a difference in survival outcomes. based on these data, maintenance choice should be guided by patient specific anticipated tolerance rather than drug type alone. iss stage and post-ahct disease response continue to be significant predictors for outcomes. toxicities recorded on maintenance were as anticipated. length of maintenance therapy may be a significant predictor and warrants further analysis. the analysis was underpowered to disclosure of conflict of interest: none. p real-world multiple myeloma management practice patterns and outcomes in six central and eastern european countries d coriu , d dytfeld , d niepel , i spicka second autologous stem cell transplantation as salvage therapy for multiple myeloma: impact on progression free survival and overall survival second autologous stem cell transplantation: an effective therapy for relapsed multiple myeloma second auto asct for treatment of relapsed multiple myeloma the role of second autografts in the management of myeloma at first relapse moving beyond autologous transplantation in multiple myeloma ebmt data office bortezomib-based versus nonbortezomib-based induction treatment before autologous stem-cell transplantation in patients with previously untreated multiple myeloma: a meta-analysis of phase iii randomized, controlled trials first-line therapy with thalidomide and dexamethasone in preparation for autologous stem cell transplantation for multiple myeloma mechanism of action of bortezomib and the new proteasome inhibitors on myeloma cells and the bone microenvironment: impact on myeloma-induced alterations of bone remodeling boys; girls) with following mds types: refractory cytopenia of childhood- ( %), refractory anemia with excess blasts - pts ( %), refractory anemia with excess blasts in transformation- pts ( %), juvenile myelomonocytic leukemia in pts ( %). the median of age was years ( - years) mac consisted busulfan (bu) mg/kg + cyclophosphamide mg/kg. ric included fludarabine (flu) mg/m + melphalan (mel) mg/m , flu - mg/m + bu mg/ kg. the bone marrow (bm) was used in pts ( %), peripheral blood stem cells (pbsc) in pts ( %), combination of bm and pbsc in pts ( %). -years overall survival (os) was % os was in pbsc group - %; bm group- %, combination of bm and pbsc- % there were two cases of mds, eb- , although erythroid aberrancy can not be found, fc did disclose significant aberrancy on myelomonocytic lineages. on the other hand, all the normal control bm samples revealed no any erythoid phenotypic abnormality. our study suggests this simplified cocktail of -tube, -color, fc is very sensitive and useful in the assessment of erythroid phenotypic abnormalities in mds we analyzed consecutive patients ( % were female, median age of (range: - ) allografted for mds (median ebmt risk score of , median disease risk index of intermediate risk) over a -year period ( - ) with mac conditioning for % and ric for % pfs ± %, grfs ± %, ri ± % and trm similarly, there was not difference between tdep and non tdep patients for -years pfs ( ± % and ± %, p value . ), -years gfrs ( ± vs ± , p value . ) (graph), -years ri ( ± % and ± %, p value . ) and -years trm ( ± % and ± %, p value . ). finally, tdep had no significant impact on -years grade - agvhd when compared to the non tdep ( ± % and ± %, p value . ). it had not either on -years cgvhd ( ± % and ± %, p value . ). our study shows that tdep is feasible on patients undergoing hsct for mds disclosure of conflict of interest: none. p mutational pathway and dynamics may not be prognostic in patients with myelodysplastic syndrome receiving hypomethylating agent pre-treatment for allogeneic stem cell transplantation republic of korea; department of computer science; the donnelly center for cellular and biomolecular research amebiazis after bone marrow transplantation use of a five-agent gvhd prevention regimen in recipients of unrelated donor marrow impact of age on outcomes after bone marrow transplantation for acquired aplastic anemia using hla-matched sibling donors treatment of acquired severe aplastic anemia: bone marrow transplantation compared with immunosuppressive therapy-the european group for blood and marrow transplantation experience disclosure of conflict of interest: none. leukemia, myelodisplastic syndrome, juvenile myelomonocytic leukemia and chronic myelomonocytic leukemia s bondarenko hla-mismatched unrelated (n = , %), and haploidentical (n = , %) donors. response was achieved in % (n = ) of pts after - (median ) courses of hma therapy: complete remission (cr) in ( %), partial remission (pr) in ( %) of pts. stabilization (s) was documented in ( %) pts, in ( %) pts there was disease progression (p) after beginning of hma therapy mismanaging the gift of life: noncompliance in the context of adult stem cell transplantation l'adhésion thérapeutique et at. des lieux en allogreffe de cellules souches hématopoïétiques (csh) dans des services de pédiatrie et d'adulte. rapport de la sfgm-tc predictive validity of a medication adherence measure in an outpatient setting data is limited to small case series, transplant registries and a single prospective multicenter observational study. here we report our institutional experience with auto-hct in patients with hrl. twenty patients with hrl [non-hodgkin = ( %), hodgkin = ( %)] and treatable hiv infection underwent hdt consisting of carmustine, etoposide, cytarabine and melphalan (beam) followed by peripheral blood auto-hct from / to / . in cases rituximab was administered as part of the preparative regimen. patient-, disease-, and transplant-related characteristics are summarized in table . median age was years (range: - ). the median follow-up for surviving patients was months (range: - ) abbreviation: n: number of patients; m: male gender; auto-hct: autologous hematopoietic cell transplant; nos: not otherwise specified; dlbc: diffuse large b-cell lymphoma ara-c), melphalan; cr : first complete remission; cr :second complete remission disclosure of conflict of interest: none. p incidence of secondary primary malignancies (spm) in patients with multiple myeloma m curly , g laurent and k nicolaus city of hope igm ( . %), lines of induction regimens prior to hsct one in pts ( %), two in pts ( . %), in pts ( . %), and missing in pts ( %). induction regimens included imids and proteasome inhibitor (pi)s with alkylating agents in pts ( . %), imids and pis with no alkylating agents in ( . %), and alkylating agents with no imids or pis in ( . %) and missing data in ( %). radiotherapy was used pre hsct in pts ( . %), no radiation in pts ( %) and missing data in ( . %). plerixafor (p) was administered mostly for poor hsc mobilization as defined by the centers number of hsc collected o × in pts ( . %), - in pts ( . %), × in pts ( %), and data missing in ( %). the number of cd + hsc infused o × in pts ( %), - × in pts ( % , × in pts ( %), and missing in ( %). a total of pts developed spm with cumulative incidence of . % ( %ci . , . ) at mo. data are missing in pts ( %) use of radiotherapy, type of induction, hsc cell dose did not influence the cumulative conflict of interest: f. sahebi, none declared, s. iacobelli, none declared, l. koster none declared l. gardaret none declared, n. kroger received research fund from sanofi, curly morris, none declared p interaction between center effect and strategy for gvhd prophylaxis on outcome of t-cell depleted and t-cell replete haploidentical transplant inserm u ecstra team expanding transplant options to patients over years-improved outcome after reduced intensity conditioning mismatched-unrelated donor transplantation for patients with acute myeloid leukemia: a report from the acute leukemia working party of the ebmt nkg d ligands in tumor immunity comprehensive analysis of nkg d ligand expression and release in leukemia: implications for nkg d-mediated nk cell responses nkg d cars as therapy for cancer russian federation high incidence of mixed chimerism with impaired graft function remains a significant issue in patients with wiskott-aldrich syndrome (was) after hsct. simultaneous use of plerixafor with g-scf is efficient in inducing stem cell release and opening of bone marrow niches. the use of plerixafor/g-csf in conditioning demonstrates better levels of donor chimerism in patients with acute myeloid leukemia. we report our experience of plerixafor/g-csf usage in patients with was as an addition to myeloablateive conditioning to improve stem cell engraftment p = , . events were considered: death in patients, graft rejection in patients, mixed myeloid chimerism (less than % donor) in patients. median time of event was , months after hsct ( . - . ) all patients are alive, median fu is months, range: . - . . patients had acute gvhd: -grade (gut), -grade (skin), in both cases resolved after a short course of steroids. all patients had more than % donor chimerism monthly till the time of last fu. the comparison of peripheral blood chimerism (% of donor cells) in was patients transplanted with and without plerixafor/g-csf in conditioning is shown (figure ). the additional use of plerixafor with g-csf references . moratto et al disclosure of conflict of interest: none. is undesirable. fifteen pts ( males, females, median age , range: - years) with high risk hematologic malignancies ( acute myeloid leukemia n. , %; acute lymphoblastic leukemia n , % pretransplant conditioning regimen consisted of thiotepa mg/kg in two days, busulfan . mg/kg in three days, and fludarabine. source of stem cells was g-csf stimulated bone marrow in all. dose of marrow nucleated cells and cd + were . (range: . - . ) × /kg and . (range: . - . ) × /kg respectively. post-transplant cyclophosphamide at mg/kg/ day was given on days and after transplantation, together with cyclosporine (starting at day − until day posttransplant) and mycofenolate (from day + to day + ) modeling autism spectrum disorders with human neurons autism spectrum disorders neurobiology and genetics of autism: a developmental perspective. the development of autism: perspectives from theory and research wharton's jelly-derived mesenchymal stem cells treatment in children with cerebral palsy: our second preliminary results of the clinical application in poland a mucha , k kosterna , m chroscinska-krawczyk , m kotarska , k mitosek-szewczyk , m murzyn the polish stem cell bank cases application potential of bone marrow mesenchymal stem cell (bmscs) based tissueengineering for spinal cord defect repair in rat fetuses with spina bifida aperta sensory neuron differentiation potential of in utero mesenchymal stem cell transplantation in rat fetuses with spina bifida aperta: sensory neuron differentiation of in utero mscs analysis of post allo-hct relapse in acute leukaemia patients, a comparative on second allo-hct and donor lymphocyte infusions g orti , j sanz , i garcia-cadenas , i sanchez-ortega , mj jimenez , p barba , c ferra , r parody , j sierra , ma sanz , s querol and d valcarcel hospital universitari vall d´hebron; hospital universitario la fe; hospital de sant pau i la santa creu; hospital duran i reynals ico, hospital germans trias i pujol ico; hospital germans trias i pujol and banc de sang i teixits acute leukaemia relapse after allogeneic hematopoietic cell transplantation (allo-hct) associates poor prognosis. in this scenario, lowering the tumour burden prior to a second allo-hct ( nd allo-hct) or donor lymphocyte infusions (dli) is essential to improve survival. thus, patients that respond to chemotherapy and subsequently receive a dli or nd allo-hct appear to associate better outcomes compared to patients receiving only chemotherapy, but data regarding this particular group of patients is lacking. we retrospectively analysed a cohort of post allo-hct relapsed acute leukaemia patients, who, after tumour reduction, were treated with either a nd allo-hct or dli. data was collected from centers, patients were consecutively included from to . patients were treated to reduce the tumour burden and received the nd allo-hct or dli on morphological remission or postchemotherapy aplasia. patients ( %) were diagnosed with aml and ( %) with all. patients ( %) underwent nd allo-hct and ( %) received dli. median patient age was ( - ) years. the median follow-up was ( - ) days. since data regarding time from first allo-hct to relapse was unavailable, we calculated the time from allo-hct to nd allo-hct or dli (time to nd allo-hct or dli). median time to nd allo-hct/dli was ( - ) days, and was days and days for nd allo-hct and dli respectively (p = . ). regarding the dli group, the median dli dose was . x / cd + ( . - x ) cells and the mean number of infused dli was . /patient. one-year os was % (se ± %). in os univariate analysis, longer time to nd allo-hct/dli associated better survival rates (p = . ). the -year dfs was % (se ± %). a longer time to nd allo-hct/dli (p = . ) and nd allo-hct compared to dli (p = . ) (figure ) associated better dfs. the -year nrm was % (se ± %). univariate analysis identified pb as stem cell source as linked to better nrm (p = . ). the -year relapse incidence (ri) was % (se ± %). ri univariate analysis related longer time to nd allo-hct/dli (p = . ) to lower ri. on os multivariate analysis, longer time to nd allo-hct/dli was associated to better survival (p = . ). this association was also observed on dfs multivariate analysis (p = . ). table summarizes nd allo-hct and dli univariate analysis. grade ii-iv acute gvhd was diagnosed in ( %) and ( %) patients post nd allo-hct and dli, respectively. chronic gvhd was diagnosed in ( extensive) and patients after a nd allo-hct and dli, respectively. in this study, longer time to nd allo-hct/dli associated better dfs. nd allo-hct (compared to dli) associated better dfs on univariate analysis, but this association was not observed on multivariate analysis. of note, the nd allo-hct group included more patients with longer time to nd allo-hct/dli. this might be explained by nd allo-hct patients relapsing later or by the fact that the preparation of a nd allo-hct might require longer time than dli. results of this analysis warrant further study with larger number of patients.advancing age is associated with worse prognosis in acute myeloid leukemia (aml). intensive induction chemotherapy in patients aged ⩾ years results in lower aml remission rates with increased induction mortality vs younger patients. cpx- is a liposomal formulation of cytarabine and daunorubicin encapsulated at a : molar ratio. a phase iii, randomized, open-label study of cpx- vs + (cytarabine and daunorubicin) in newly diagnosed older patients with high-risk secondary aml showed superior survival in the cpx- arm (hazard ratio . ; p = . ). in that trial, eligible patients went on to allogeneic hematopoietic cell transplantation (hct). an exploratory analysis of those patients by age strata is reported here. patients aged to years with newly p number, composition and/or antileukemic activity of (dc-stimulated) invariant nkt-, nk-and cik-cells is predictive for outcome of patients with aml, all and cll cl boeck # , dc amberger # , f doraneh-gard , w sutanto , t guenther , j schmohl , f schuster , h salih , f babor , a borkhardt myelofibrosis (mf) is a hematolgic malignancy which is characterised by extramedullary hematopoiesis due to bone marrow fibrosis resulting in spleno-and/or hepatomegaly. allogeneic stem cell transplantation (allo-hsct) is the only curative treatment for mf but is associated with therapy related morbidity and mortality. retrospective studies suggested an increase of liver toxcicity in mf patients in comparison to other diseases following allo-hsct. the aim of this prospective study was to evaluate the impact of liver stiffness measured by transient elastography (fibroscan) on liver toxicity after allo-hsct. between and we included patients (male %, female %) who underwent allo-hsct due to primary mf( %), postpv/et-mf ( %) or mf in transformation ( %). the median age of the patients was y@@@ears (range: - ). conditioning regimen was mainly busulfan based reduced intensity. all patients received atg. gvhd prophylaxis was csa/mmf in all patients. stem cell source was peripheral blood in % and bone marrow in % of the patients. donor sources were as follows: mrd ( %), mud ( %) and haploidentical relative ( %). fibroscan was performed prior to conditioning. elevated liver enzymes, bilirubin above the normal value or the onset of veno-occlusive disesae (vod) from the time of conditioning start and within the first post-transplant days were considered as indicators for liver toxicity. the median stiffness of the liver measured by fibroscan on the day before conditioning treatment start was . kpa (range: . - . ). six patients ( %) had prior liver diseases such as cirrhosis (n = ), viral hepatitis (n = ), steatosis (n = ), or vod (n = ). the median onset of liver toxicity was day (range: − until + ). the median bilirubin level of all patients was mg/dl (range: - ). the median ap level was u/l (range: - ), the median ggt level was u/l (range: - ), the median alt level was u/l (range: and the median ast level was u/l (range: - ). the pearson-test revealed a positive correlation between liver stiffness and the elevation of the ap (r = . , p = . ) and ggt levels (r = . , p = . ). the comparison of the median maximum enzyme and bilirubin levels is shown in table . in two patients who developed severe vod requiering defibrotide, the liver stiffness level was . kpa and . kpa, respectively. the patient with the highest stiffness level ( . kpa) developed acute gvhd of the liver, which completely resolved after steroid treatment. only one of those five patients who had stiffness levels kpa died due to liver toxcity and concurrent septic shock, he suffered from viral hepatitis prior to transplantation. liver stiffness measured by transient elastography (fibroscan) positively correlates with the elevation of the cholestatic enzymes ap and ggt in myelofibrosis patients after allo-hsct and may predict liver toxicity. disclosure of conflict of interest: none.[p ]in the era of tyrosine kinase inhibitors (tki) as superior first line treatment in the therapy of cml, the concept of allogeneic hsct has been pushed to the role of salvage therapy. to date, data on allogeneic hsct after tki-therapy are scarcely available. in this study, we report single center data on the outcome of cml patients, for the most part pretreated with tki, who underwent allogeneic hsct between and with a follow-up of months to years. upon obtaining written informed consent patients diagnosed with bcr-abl-positive cml and patients with bcr-abl-negative atypical cml were included in this analysis. the majority of patients underwent myeloablative conditioning regimen. the median age at time of hsct was years with a range: from to years. twenty-one patients were transplanted from a matched related donor, and received stem cell grafts from an unrelated hla-compatible donor. / patients received tki-therapy before transplantation, patients received more than tki prior to hsct. / patients were treated with interferon prior to hsct. twenty-two patients were transplanted due to acceleration or blast crisis. twenty-six patients received an allogeneic hsct in chronic phase (cp, n = ) or complete hematologic (chr, n = ) or cytogenetic remission (ccyr, n = ). kinase domain mutations could be identified in seven patients including t i-mutation in four patients. seven patients showed "major route" cytogenetic aberrations. next to advanced disease status, tki intolerance (n = ) and tki resistance (n = ) were the main indications for hsct after . after a median follow up of years and months, those patients transplanted in cp, chr or ccyr showed an overall survival (os) of %. / patients died in remission and two patients died after cml relapse. after none of the patients transplanted in cp, chr or ccyr died or relapsed so far, with a median follow-up of days. all of these patients received tki therapy prior to transplant. twenty-two patients transplanted in advanced stage cml (bc and ap) had after a median follow up of years an os of %. the difference between survival curves is significant (log rank test p = . ; hr . , % ci of ratio . - . ). prior to transplantation of these patients received a tki-therapy. in this group, four patients died due to cml relapse, one died after development of donor cell leukemia and five patients died in remission. one patient with atypical cml was transplanted in bc and died of progressive disease shortly after transplantation. the other three patients with atypical cml were transplanted in cp-phase. with a median follow-up of days these patients are in ongoing remission. even in times of tkitherapy allogeneic hsct remains a successful and safe therapy option for cml patients with tki intolerance or resistance. patients transplanted in cp or complete remission had an excellent long-term outcome. allogeneic hsct should be considered in tki resistance or intolerance before the development of blast crisis. despite tki therapy, overall survival deteriorates in patients with advanced disease. however, this treatment modality can improve survival rates substantially compared to other available therapies. tkimaintenance therapy could be a possible strategy to prevent cml relapse, although randomized data on tki-maintenance therapy after allogeneic hsct are still lacking.[p ]disclosure of conflict of interest: none. use of first or second generation tki for cml after allogeneic hematopoietic stem cell transplantation: a study by the cmwp of the ebmt y chalandon, s iacobelli , j hoek , l koster , l volin , j finke , jj cornelissen , i yakoub-agha patients (pts) relapsing with cml after allogeneic hematopoietic stem cell transplantation (allohsct) may be treated with tki and/or dli. as nowadays the majority of cml pts would have received at least imatinib prior to transplantation, we were interested in analizing (a) the type of tki used after allohsct, (b) the indication for tki treatment, (c) the outcome of this treatment and d) the temporal relationship with dli if given. pts received tki after first allogeneic hsct for cml. transplants had been performed in cp , n = , ap, n = or for more advanced disease (bc/ cp , n = ) from hla identical siblings (n = ) or ud (n = ) between and . tki given prior to transplant was imatinib (n = ), dasatinib (n = ), nilotinib (n = ), bosutinib (n = ) and ponatinib (n = ). median age at transplant was ( . - ) years, pts ( %) were male. tki post allohsct were given between and . first tki given was either imatinib (n = ), dasatinib (n = ), nilotinib (n = ), bosutinib (n = ) or ponatinib ( ). the indications for tki therapy were the same as for transplantation (n = ), for relapse/progression/ persistent disease (n = ), for prophylaxis/pre-emptive (n = ), planned (n = ), others (n = ) and missing (n = ). median follow-up from start of tki was ( - ) months. the median time interval from transplant to tki was ( . - ) months. it was longer for tki given for relapse/progression with ( - ) months and shorter for tki given for prophylaxis/pre-emptive with . ( . haematopoietic cell transplant (hct) is the only curative approach for scd. due to concerns regarding the toxicities associated with myeloablative conditioning regimens in adults, a non-myeloablative protocol was developed by hsieh et al. (national institutes of health, nih protocol). the use of this novel regimen was able to achieve a curative degree of mixed donor chimerisms with minimal transplant-related complications. the alberta children's hospital (ach) has adopted this conditioning regimen in children due to the efficacy and low rates of toxicities published by the nih group. with generally lower rates of gvhd in younger recipients, our group had no reason to believe rates of toxicities would be greater in a younger population with fewer comorbidities secondary to scd than those described in the nih cohort. to our knowledge, there is no published literature describing the utilization of the nih protocol in a paediatric population. we describe our experience in children with scd who underwent matched sibling donor (msd) peripheral blood hct using nih protocol. this retrospective cohort describes outcomes of msd hct in children with scd who underwent hct with the nih conditioning regimen between - . a total of potential subjects were identified. eight subjects have consented to the analysis to date. msds with either normal haemoglobin or sickle cell trait were considered appropriate for donation. the transplant procedure: the conditioning regimen consisted of alemtuzumab . mg/kg/dose administered subcutaneously daily for five days (days − to day − ). patients received a tbi dose of cgy on day - , with testicular shielding for male recipients. gvhd prophylaxis consisted of a sirolimus load of mg/ m /dose (po) on day − , followed by mg/m /dose once a day starting on day . unmanipulated peripheral blood stem cells were infused on day . sirolimus was used for gvhd prophylaxis post-hct and continued until at least one year. weaning of sirolimus was initiated no earlier than year post-hct and if donor t-cell chimerisms were greater than %. institutional supportive care protocols for scd hct were followed. patients were eligible for early discharge post-hct even prior to neutrophil engraftment. eight patients ( f, m) have been registered on this retrospective study. follow-up ranges from to months post-hct. there were no failed stem cell mobilizations. all patients had donor neutrophil engraftment at a median of days. all patients are currently alive. there have been no cases of graft failure to date and no sickling crises post-hct. one patient has dropping myeloid chimerisms but still % donor. no cases of veno-occlusive disease, idiopathic pneumonia syndrome, cerebral hemorrhage, pres, or posttransplant lymphoproliferative disease were observed. three cases of cytomegalovirus (cmv) reactivation required pre-emptive therapy. only one patient did not initiate sirolimus weaning at year post-hct due to donor t-cell chimerisms of %; this patient is months post-hct and is likely to start weaning sirolimus soon. there have been no cases of acute or chronic graft-versus-host disease. nonmyeloablative conditioning regimen is safe and effective as curative therapy for scd. long-term follow-up of these children to assess organ function post-hct is underway. disclosure of conflict of interest: none.the number of new hiv/aids cases has been declining in developed countries, whereas it is still increasing in japan, with the cumulative number reaching , as of june , . hiv infection is associated with an increased risk of hematological malignancies such as non-hodgkin lymphoma (nhl). autologous hematopoietic cell transplantation (auto-hct) is a treatment option for hiv-infected patients with nhl and multiple myeloma (mm). however, the prognosis after auto-hct in hiv-infected japanese patients remains unclear. the aim of this study is to evaluate the effect of hiv infection on transplant outcomes after auto-hct in japan. using the national database of the japan society for hematopoietic cell transplantation, we retrospectively evaluated patients with nhl (n = ) and mm (n = ) who underwent their first auto-hct between and . presence of hiv antibodyperipheral t-cell lymphomas (ptcl) comprise a heterogeneous group of diseases among which ptcl-not otherwise specified (ptcl-nos) represents the most common histology. patients with ptcl are typically offered high-dose chemotherapy followed by autologous hematopoietic cell transplantation (auto-hct) as front-line consolidation. allogeneic hct (allo-hct) is generally offered in the relapsed setting; however, in selected cases it is also offered as front-line consolidation. no randomized controlled trial (rct) have been performed to date comparing offering an allo-hct versus other treatment modalities either in the front-line or in the relapsed setting. thus, we performed this systematic review/meta-analysis to assess the totality of evidence pertaining to the role of allo-hct in ptcl. search of the literature was undertaken via pubmed and web of science from inception until september , . no search limits were applied but studies presented only in abstract form were excluded. data were collected on treatment benefits (complete remission (cr), progression-free survival (pfs), overall survival (os)) and harms (non-relapse mortality (nrm), grade ii-iv acute graft-versus-host disease (gvhd), and chronic gvhd). the search identified references; however, only studies ( in front-line (n = pts), in relapsed/refractory setting (n = pts)) were eligible based on our inclusion criteria and had extractable data. three studies included both frontline and relapsed/ refractory cases but data for certain outcomes were reported separately. the median follow-up time for studies evaluating allo-hct in the front-line or relapsed/refractory setting ranged from - months and - months, respectively. in the front-line setting, allo-hct resulted in cr rates of % (( %ci = - %), studies, n = pts), pfs rate of % (( % ci = - %), studies, n = pts), and os rate of % (( % ci = - %), studies, n = pts). nrm rate was % (( % ci = - %), studies, n = pts). acute (grade ii-iv) and chronic gvhd rates were % ( % ci = - %), studies, n = pts) and % ( % ci = - %), studies, n = pts), respectively. in the relapsed/refractory setting, allo-hct resulted in cr rates of % (( % ci = - %), studies, n = pts), pfs rate of % (( % ci = - %), studies, n = pts), and os rate of % (( % ci = - %), studies, n = pts). nrm rate was % (( % ci = - %), studies, n = pts). acute (grade ii-iv) and chronic gvhd rates were % ( % ci = - %), studies, n = pts) and % ( % ci = - %), studies, n = pts), respectively. notwithstanding the need to perform a rct to compare the efficacy of allo-hct versus auto-hct as front-line consolidation in ptcl, the results of this systematic review/meta-analysis show very encouraging os rates of % following allo-hct. moreover, allo-hct also offers an encouraging os rate of % in patients with ptcl in the relapsed/refractory setting. the higher nrm rate in the relapsed/refractory setting probably reflects the adverse effect of a higher number of prior prescribed therapies. one of the limitations of our analysis is the inability to analyze outcomes for individual histologic subtypes due to the aggregate nature of the published data. disclosure of conflict of interest: none. high-risk patients with relapse or refractory hodgkin lymphoma do significantly better after hdc auto-sct compared to control arm of aethera trial. mature results from a cohort of patients s akhtar, s rauf, tam elhassan and i maghfoor king faisal specialist hospital and research center, riyadh, kingdom of saudi arabia brentuximab vedotin use in hodgkin lymphoma (hl) patients who had hdc auto-sct has been reported to improve progression free survival (pfs) but not the overall survival (os) in a phase trial (lancet ; : - ). in this trial, after hdc auto-sct, high risk hl patients were randomized to receive placebo (control gp) vs brentuximab (experimental gp) as consolidation therapy. we are reporting our experience of patients with similar selection criteria as control gp. hl patients z yrs who received hdc auto-sct with similar selection criteria as defined in aethera trial were identified that is, patients had at least one of the following risk advanced lymphomas still represent a therapeutic challenge and allo-hsct is among treatment options. between march and august , seventy-three patients (pts) affected by r/r lymphomas ( nhl and hl) underwent an allo-hsct after a treosulfan-based conditioning regimen and sirolimus as calcineurin-inhibitor-free prophylaxis of gvhd. six pts received a mrd, pts a mud, and pts a haplo unmanipulated pbsc. at allo-sct pts were in cr, pts were in pr, and pts had sd/pd; sixty patients underwent autologous sct before allo-hsct. hct-ci was evaluable for pts, had a score ≥ . thirty-three pts received treosulfan and fludarabine reduced toxicity conditioning regimen (rtc) and intensification with other alkylating agent or with gy total body irradiation was added on the remaining pts (myeloablative conditioning, mac). all pts received a backbone gvhd prophylaxis with sirolimus and mycophenolate mofetil; atg or pt-cy or both were added in , , and pts respectively. median numbers of infused cd +/kg and cd +/kg were . × (range: . - . ) and . × (range: . - . ), respectively. median follow-up was months (range: - ); median time to neutrophil ≥ . × /l was days, and days to platelet ≥ × /l. sixteen out of patients with pre-transplant active disease obtained a cr after treosulfan conditioning; nine of them ( hl and b-nhl) achieved durable cr without post transplant treatment. oneand -years os was % and %, pfs was % and % at and years respectively; cumulative incidence of relapse/ progression was % and % at and years. grfs was % and % at and years, respectively. transplant related mortality (trm) was % at days, % at year and for the entire follow-up. the -day cumulative incidence (ci) of agvhd grade ≥ was % and ci of agvhd grade ≥ was %; ci of moderate to severe cgvhd was % at years and for the entire follow-up. no differences in ci of agvhd or cgvhd were found if pts were stratified according to donor type, but ci of moderate-severe cgvhd was significantly higher in pts after mac regimens (p o . ). as expected, the outcome of pts in cr was significantly better compared with active disease, in terms of os (p = . ), pfs (p = . ), ri (p = . ). in multivariate analysis, intensity of conditioning regimen (rtc vs mac), gvhd prophylaxis (use of atg, pt-cy or none), donor sex and age at allo-sct did not impact the transplant outcomes; both os and pfs were reduced by active disease at allo-hsct (hr = . , ci % . - . , p = . and hr = . , ci % . - . , p = . , respectively) and by nhl histology (hr = . , ci % . - . , p = . and hr = . , ci % . - . , p = . , respectively); grfs and ri were impacted only by active disease (hr = . , ci % . - . and hr = . , ci % . - . , p = . , respectively). allo-hsct after treosulfan conditioning and sirolimus gvhd prophylaxis is feasible even in heavily pretreated pts affected by lymphomas. complete remission status before transplant remains crucial for better outcomes and in the era of new targeted treatments should be pursued. disclosure of conflict of interest: none.university of eastern finland, kuopio, finland and department of medicine, kymenlaakso central hospital, kotka, finland autologous stem cell transplantation continues to have an important role in the treatment of patients with multiple myeloma (mm). in mm patients the most commonly used mobilization method is granulocyte-colony stimulating factor (g-csf) ± cyclophosphamide (cy). generally, up to - % patients mobilize poorly with these methods and plerixafor may be used to enhance mobilization. the most important parameter of graft quality has usually been the number of cd + cells, but there are also significant numbers of other cell subsets in the grafts and they may also be of special interest in regard to post-transplant recovery and outcome. for example, a higher number of lymphocytes and nk cells in the grafts has been associated with improved lymphocyte as well as nk cell recovery, respectively. the mobilization methods used seem to affect the graft composition. however, there is currently no prospective data on the effects of plerixafor on the graft composition, post-transplant hematological and immune recovery or outcome in patients with mm. altogether eighty-seven patients with mm were included into this prospective study. seventy-seven patients were mobilized with g-csf ± cy (control group) and ten patients received also plerixafor due to poor mobilization (plerixafor group). in the control group / ( %) and in the plerixafor group / ( %) of patients were mobilized with g-csf+cy (p = . ). there were no statistically significant differences between the groups according to age, gender, paraprotein type, initial iss, induction therapy used or disease status at the time of mobilization. by imwg risk stratification, there were more high risk patients in the plerixafor group ( / vs. / , p = . ). cryopreserved graft samples were analyzed with flow cytometry for t and b cells (cd /cd /cd /cd ) as well as for nk cells (cd /cd +cd ). also, cd + cell subclasses were analyzed (cd /cd /cd ). complete blood counts were evaluated at + days, , , and months posttransplant. to evaluate immune reconstitution, flow cytometry of lymphocyte subsets (t, b, nk) was performed in a subset of patients at , and months after the graft infusion using the same antibody panel as for graft analysis. there were no significant differences between the groups in the number of cd + cells in the grafts. also, the median number of aphereses was two in the both groups (p = . ). the proportion of the more primitive cd + cells (cd + cd + cd -) was significantly higher in the plerixafor group (p = . ). in addition, the number of various lymphocyte subsets analysed was significantly higher in plerixafor group table ). there were no statistically significant differences in the course of hematological recovery. the recovery of blood cd +cd + t cells was significantly faster in the plerixafor group at one at three moths post-transplant. there was no significant difference in the progression-free survival (pfs) (log rank, p = . ) with the median follow-up time of days in the plerixafor group and days in the control group ( . ). in the present study plerixafor added to g-csf ± cy seemed to significantly alter the cellular composition of autologous blood grafts in poorly mobilizing mm patients. hematological recovery was comparable but the cd +cd + t lymphocyte recovery was faster in the plerixafor group. the pfs was comparable between the groups. disclosure of conflict of interest: dr. valtola has received honoraria from sanofi and jansen-cilag. dr. silvennoinen has received a research grant from celgene and janssen, honoraria from genzyme and sanofi and participated in advisory board organized by amgen, janssen and takeda. dr. siitonen has received honoraria from amgen and celgene. dr. jantunen has received honoraria from genzyme, amgen and sanofi and has participated in eu leadership meeting organized by genzyme as well as medical advisory board meeting organized by genzyme and amgen. dr. varmavuo has received consultancy fees from abbvie, roche, celgene, amgen and sanofi. the other authors declare no conflicts of interest. bortezomib after high-dose melphalan as conditioning regimen before autologous stem cell transplantation in patients with multiple myeloma: a comparison with the historical conditioning regimen with melphalan alone ga ferini; ja arbelbide; al basquiera; e nucifora; n schutz; v otero; d fantl hospital italiano d buenos aires, buenos aires, argentinahigh dose of melphalan followed by autologous stem cell transplantation (asct) is the standard of care for younger patients with multiple myeloma (mm). to enhance the efficacy of the conditioning regimen, the intergroupe francophone du myelome added bortezomib to melphalan showing improved response rates, without significant toxicity. bortezomib has shown synergistic effects with melphalan, mainly if the bortezomib is administered hours after the melphalan. since , we have changed our conditioning regimen for patients with mm undergoing asct by adding bortezomib toallogeneic stem cell transplantation (allosct) is a potencially curative option for patients with multiple myeloma (mm). despite the improvement of reduced-intensity-conditioning (ric), transplant-related mortality (trm) remains high. there is no consensus on which graft versus host disease (gvhd) prophylaxis regimen is superior. some studies have suggested that tacrolimus-based prophylaxis is more effective than cyclosporine (csa) in terms of lower incidence of severe acute gvhd (agvhd), with no impact on overall survival (os). herein, efficacy and toxicity between two gvhd prophylaxis regimens is analyzed. we retrospectively analyzed patients (pts) with relapsed mm who received allosct ric in the period from to in a single centre (table ) . population: age, years ( - ); median follow-up: months ( - ). conditioning regimen: allo-ric (fludarabine + busulfan or melphalan regimens) and % was bortezomib-based in the tacrolimus group. donor: matched related ( pts), unrelated ( ), mismatch unrelated ( ) and haploidentical ( ) donor. gvhd prophylaxis: all patients received a short course of methotrexate + csa ( pts, %) or tacrolimus ( pts, %). complete response at transplant was % at csa group and % at tacrolimus group. all pts underwent toxicity related to chemotherapy (mainly mucositis and neutropenic fever) with organ impairment (renal or liver) in % tacrolimus arm as well as pts in csa group. the incidence of agvhd was % and . % in tacrolimus and cyclosporine groups, respectively (p = . ). grade iii-iv agvhd were reported in pts ( %,tacrolimus) and pts ( %, cyclosporine), with severe gastrointestinal and liver involvement. glucocorticoid resistance was observed in % in both groups. patients with refractory agvhd received other immunosuppressive therapies: more than second-lines agents ( ) ( ) ( ) ( ) were necessary in fifty percent of pts in both groups to control gvhd. two patients had to interrupt tacrolimus due to neurological toxicity and suspected thrombotic thrombocytopenic purpura. no patients had to discontinue treatment in the csa arm because of toxicity. the -months os was . % ( % in tacrolimus vs . % in csa (p = . )) and the -months was . %. a total of pts died because of gvhd. during follow-up, only patients relapsed ( and months after allosct, respectively) in csa group. no relapse were seen in tacro group. in our experience, no significant differences were observed between both calcineurin inhibitor in terms of os, toxicity and gvhd incidence. an explanation could be our small number of patients. allosct is an effective therapy for selected patients but it is associated with high rates of gvhd and trm. a long-termsafety and effective prophylactic regimen is necessary as main objective.[p ]disclosure of conflict of interest: none. several parameters, including early lymphocyte, neutrophil, platelet recovery, and infused dose of cd + cells, have been associated with clinical outcome of patients with haematological malignancies. however, their prognostic significance remains uncertain. the aim of current study was to evaluate prognostic significance of clinical and laboratory parameters that might influence survival after autologous stem cell transplantation (asct) in hodgkin lymphoma (hl) and multiple myeloma (mm). this retrospective study included a total of with hl and mm patients (median age years, years, respectively) who underwent asct between november and june . hl patients were conditioned with beam ( . %) and cbv ( . %) regimen, while mm patients received conditioning with high dose of melphalan. high ips (international prognostic score) at diagnosis had . % hl patients and high iss (international scoring system) had . % of mm patients, of which . % had renal impairment. the average of transplanted cd + cells in hl patients was . × /kg (range: - . × /kg), and . × /kg (range: - . × / kg) in mm patients. after asct, favourable treatment response (partial/complete remission) achieved . % hl patients, of whom . % had infused o × /kg cd + cells. median time to recovery of absolute lymphocyte count × /l or greater (alc ) was days (range: - days), recovery of absolute neutrophil count ≥ × /l (anc ) was (range: - days), and platelet recovery ≥ × /l (plt ) was days (range: - days). after asct, . % mm patients achieved favourable treatment response, of whom . % had infused cd + cell dose . × /kg. median time to alc was days (range: - days), anc was (range: - days), and plt was days (range: - days). median follow up of patients with hl was months, while after asct, median event free survival (efs) was months, and overall survival (os) was months. treatment response after asct strongly influenced both efs and os after asct (po . ). in patients who achieved favourable treatment response, os and efs after asct were influenced by infused cd + cell dose (o × /kg vs. ≥ × /kg), prolonged recovery of alc by day+ , plt by day + , and achieving of anc by day + (po . ). multivariate analysis among significant variables showed that infused cd + cell dose was the most important parameter that influenced os and efs (po . ). median follow up of mm patients was months, while after asct, median efs was months and os was months. regarding patients who achieved favourable treatment response, os and efs after asct were influenced by the presence of renal impairment, infused cd + cell dose (≤ . × /kg vs. . × /kg) and plt recovery by day + (po . ). among these significant parameters, multivariate analysis pointed out infused cd + cell dose as the most important parameter that influenced both os and efs (po . ). these data suggest that number of infused cd + cells is an independent factor that may contribute to outcome of patients with hl and mm. disclosure of conflict of interest: none. high-dose therapy with autologous stem cell transplantation (asct) has become the treatment of choice for symptomatic eligible patients with multiple myeloma (mm). we studied an induction regimen of cyclophosphamide, bortezomib and dexamethasone (cybord) and showed rapid and deep responses after cycles in patients with newly diagnosed mm and we subsequently done asct with melphalan (mel) conditioning. cost is the major limiting factor in developing world.all the drugs used are generic brands manufactured in india. a total of mm patients (median age: . years, % male and % female) were transplanted between and . in all, patients had igg kappa- ( %), igg lambda- ( %), iga lambda- ( %), iga kappa- ( %), kappa light chain ( %), lambda light chain ( %) patients. prior to autograft, all cases had received cybord with generic medicines. median time diagnosis to asct was . months ( to months). stem cell mobilization was done with g-csf alone in ( %), g-csf plus plerixafor in ( %) and chemo mobilization in ( %) patients. all patients received asct support after conditioning with mg/m generic melphalan alone (dose adjustment was done according to renal status). all patients received thalidomide maintenance from march . bortezomib used was manufactured by dr. reddy's lab, hyderabad and melphalan used was manufactured by emcure pharmaceuticals, pune, india. patients from to received cyclophosphamide, vincristine, adriamycin and dexamethasone (cvad) protocol of originator medicines followed by originator melphalan conditioning and asct (cvad-mel-asct). at the time of autograft, ( %) of patients were in complete remission, ( %) in partial remission, ( %) very good pr. median day of engraftment was for neutrophils and for platelet. transplant related mortality was % ( / ) out of which died of infection and deaths of cardiac events. the pfs and os rates were % and % at median follow up of . months. patients who were treated with cvad-mel-asct had efs of % at yrs and % at yrs. cost of bortezomib showed significant difference, generic was usd where as for originator drug was usd for cycles of chemotherapy. cost of melphalan also showed difference with usd for generic and usd for originator drug. generic cybord showed excellent response rate and allows excellent stem cell collection and transplantation which can further consolidate response and improve outcome. cybord induction and melphalan conditioning with generic medicines can be considered a standard regimen for transplant-eligible patients with newly diagnosed mm in resource constraint situation. generic cybord-mel-asct is more cost effective than originator cvad-mel-asct. generic medicines produced in india are of good quality and cost effective. this study needs long term follow up to assess survival parameters at a median. disclosure of conflict of interest: none. and an extra copy of one or more odd-numbered chromosomes and as intermediate risk(ir) if they had t( ; ) or del( ) (q).overall survival (os) and relapse-free survival (rfs) were calculated from the time of allo hsct and auto hsct on day , from diagnosis to death or disease progression. the median age at presentation was . (range: - ) years, and ( . %) were men. at a median follow-up time of months, % were alive. of the patients with available fish samples underwent auto hsct. patients ( . %) achieved cr and patients ( . %) relapsed. of the patients who had received allo hsct, five patients ( . %) achieved cr and five patients ( . %) remained alive. in patients who received auto hsct, the risk of relapse was % less than those never transplanted (p = . ), but the difference was not significant in patients who received allo hsct. the relapse-free survival in hr patients was months (po . ), in ir was months (p o . ) and in sr was . months (p o . ). in transplant patients, rfs in hr patients was . times more than sr group (po . ) and in ir group was . times more than sr (p o . ). the survival time in transplant patients was significantly better than non-transplanted patients (p o . ). the median overall survival (os) in hr patients was . months, in standard risk group months and in sr patients was months. cytogenetic abnormalities detected by fish are of significant value in classification, risk stratification and management of patients with mm. we can use cytogenetic data to provide prognostic information and also to guide management and clinical practice. these data indicate that autologous stem cell transplantation could potentially be of benefit to myeloma patients. disclosure of conflict of interest: none. chronic graft-vs-host disease (cgvhd) is the most troublesome complication developing after allogeneic hematopoietic stem cell transplantation (allo-hsct). diagnosis of cgvhd has largely been based on clinical features only. we previously reported gene expression profiles in patients with cgvhd after allo-hsct. we extended our study to develop a molecular diagnostic method of cgvhd. we selected six most commonly expressed genes from the former dna expression study. and, a home-made -gene pcr array were used to evaluate gene expression profiles in the peripheral blood mononuclear cells of patients given allo-hsct ( cgvhd patients, non-cgvhd patients) and normal controls. the gene expressions of the allo-transplanted patients were compared to those of the stem cell donors. sybr green qpcr and multiplexqpcr were performed to confirm the usefulness of the selected genes in the diagnosis of cgvhd. infogainattributeeal and ranker were used to develop a gene model to diagnose cgvhd. k-nearest neighbor model and weka classifiers lazy ibk module were applied to evaluate the performance of the gene model. in another steroid-refractory cgvhd patients ( responders, non-responders), the gene expression changes were analysed using our -gene pcr array before and days after rituximab treatment. we identified six genes most accurately delineating cgvhd patients from those without treatments after allogeneic hematopoietic stem cell transplantation (hsct) are long and constraining for patients. medical adherence in hsct patients is of major concern in daily practice but it has been not yet described. , the aims of our study were to evaluate treatment adherence and to identify factors associated with adherence behaviors. an observational single-center study was based on self-reported questionnaires completed by patients in a hematology day hospital between november and july . the patientreported adherence was evaluated using the eight-item morisky medication adherence scale (mmas- ). , individual item scores were summed: patients with a score of / were considered as good adherents to medication whereas a poor adherence referred to a score under . among the latter, medium adherence ranged to a score of - , while a score of o was considered low adherence. socio-demographic and medical characteristics were collected by health records. a univariate model was used to evaluate if some of patients' characteristics were associated with adherence. statistical analysis was performed using r software (version . . - - rstudio, inc). fifty-six patients were included in the current study. median age at transplantation was years (range: - years). diagnosis were aml (n = ), all (n = ), myelofibrosis (n = ) and other hematological diseases (n = ). patients received a hsct from a related donor ( haploidentical). myeloablative conditioning was used in patients and reduced intensity regimen in patients. a total of . % ( / ) of the patients were poor adherent according to mmas- . among these patients, / were low adherent and / were medium adherent. the results of univariate analysis showed that a poor adherence was associated with a longer time since hsct and discharge at home. however elderly patients, patients treated with cyclosporine and patients with daily hydration at home were associated with a better adherence (po . ). our study presents the first data on adherence among patients undergoing hsct. risk factors associated with a poor adherence have been identified in order to determine patients' profiles that will benefit more from interventions to improve adherence. particular attention has to be paid to younger patients. efforts to establish a regular follow-up of these patients are needed in order to sustain patients in the treatment adherence to prevent the occurrence of severe complications. we studied the effect of basic fibroblast growth factor (fgfb) and dexamethason on expansion and immune modulation of mscs in patients with lymphomas. mscs were generated from bone marrow aspirates obtained from the patients with hodgkin's lymphoma (hl; n = ) and non-hodgkin's lymphoma (nhl; n = ). the adherent fraction of marrow aspirate was cultivated with/without the basic fibroblast growth factor (fgf-b, ng/ml) or dexamethason ( − М or − М) to reach - % confluence. then mscs were passaged with accutase and used for experiments after - passages. the number of msc precursors (cfu-f) in bone marrow of lymphoma patients was found to be significantly decreased both in patients with nhl ( ± , p o . ) or hl ( ± , p o . ). the time until - % confluence was significantly increased and took on average ± days (vs . ± . in donors). finally, the immunosuppressive ability of patient msc was significantly lowered and was only registered at the high concentrations of mscs ( : and : ). the expansion of patient mscs was significantly promoted with fgfb resulting in a significant decrease of primary cell cultivation (from . ± . to . ± . days; p = . ) and a statistically significant twofold increase in the number of cells received at the first passaging. in addition, in cultures with fgfb there was a decrease in the relative amount of resting mscs and a threefold increase of cycled cells in cd + mscs. dexamethasone has also provided a moderate stimulating effect on the msc growth. in fact, the use of − М of dexamethasone resulted in the increase of the cell yield by . times and of − М-by . times. however, fgfb and dexamethasone differed in their effect on the msc ability to inhibit the proliferative response of t lymphocytes upon stimulation with mitogens or alloantigens. indeed, fgfb failed to correct the impaired immunosuppressive activity of patient mscs, and median percentage of suppression still remained lowered- % vs % without fgfb. in contrast to fgfb, dexamethason could increase the immunosuppressive activity of patient mscs by . times (in dose of − М) and by . times (for − М). our data indicate that fgfb and dexamethasone used during the generation of mscs exert a stimulating effect on the msc expansion. in contrast to fgfb, dexamethasone, in the broad range: of doses, was able to enhance the suppressive properties of mscs that are initially reduced in patients with lymphoma. these findings suggest the existence of at least two mechanisms of impairments in immunoregulatory function of mscs in lymphomas-dependent and independent of the msc proliferation. disclosure of conflict of interest: none. free nonabsorbable antibacterial digestive decontamination is associated with a low incidence of gastrointestinal acute gvhd and better gvhd-free/ relapse-free survival (grfs) in the atg-based conditioning regimens nabil yafour commonly antibacterial prophylaxis based of oral no absorbable antibiotic such as (neomycin colistin, gentamicin, vancomycin) used before and after engraftment, other fluoroquinolone such as levofloxacine were recently used to prevent invasive infection. however the exact interaction with gastrointestinal acute graft versus host disease (gi-agvhd) remains unclear. the objective of this study was to evaluate a novel composite endpoint of gvhd-free/relapse-free survival (grfs), in which events include grades - gi agvhd, chronic gvhd requiring systemic therapy, relapse, or death in atg based-conditioning regimens, with free no absorbable antibacterial digestive decontamination prophylaxis. a total of evaluable consecutive patients with hematological disease were included in period of february to mai . patients with malignancies disease (n = ) received myeloablative conditioning regimens plus atg ( mg/kg); including once daily busulfan ( mg/m ,- d to - d, iv ) + fludarabine ( mg/m /d, - d to - d, iv) (aml = , all = , cml = ) or melphalan ( mg/m , d- , iv) (all = ). six patients received cy/atg for saa. gvh prophylaxis consisted to; ciclosporine a (csa) + mtx. csa was maintain levels between - ng/ml and tapered at the discretion of the treating physician. all patients were received peripheral blood stem cells (pbsc) graft from a matched related donor. since december levofloxacine and voriconazole was administered as antibacterial and antifungal prophylaxis. diagnostic, clinical grading and treatment of gi-agvhd and gi-cgvhd were performed according to established criteria and nih recommendations. probability of grfs was estimate by kaplan-meier method. median age was years (range: - ). median dose of cd + and cd + cell doses were . × (range: . - ) and . × (range: , - , ). the median time to neutrophil and platelet recovery were days (range: - ) and days (range: - ) respectively. at time of transplant / ( %) had an intestinal colonization with extended-spectrum betalactamase (esbl) producing bacteria. only / ( %) developed infectious diarrhea during the period of transplant. incidence of grade iii/iv gi-agvhd and gi-cgvhd requiring systemic therapy were % and % respectively. for patients with malignancies diseases (n = ), ( %) were alive at a median follow up of months (range: - ). incidence of relapse, disease free survival rates were %, % respectively. the grfs rate as defined previously was % at months. these results confirm that free no absorbable antibacterial digestive decontamination and atg-based conditioning regimens were associated with very low incidence of gi-gvhd and better grfs in patients with malignancies diseases. diverse bacterial populations of the gastrointestinal tract remain important factors to promote immune tolerance after allogeneic sct. disclosure of conflict of interest: none. g-csf primed hla haploidentical transplantation from maternal or collateral donor using atg plus reduced dose of posttransplantation cyclophosphamide: results of a phase ii prospective trial y wang , x-j huang peking university people's hospital, peking university institute of hematologythe transplantation milieu using granulocyte colony-stimulating factor (g-csf), and anti-thymocyte globulin (atg) for hlahaplotype-mismatched transplants from related donors has resulted in favourable outcomes with low transplant-related mortality (trm), without increased relapse rate. however, in this transplant modality, the poorer outcome owing to high incidence of graft-versus host disease (gvhd) related to maternal donor or collateral donor remains a concern. meanwhile the use of post-transplant cyclophosphamide (pt/cy) in recent years appears to be protective against severe acute and chronic gvhd. we performed a prospective pilot study of hla haploidentical stem cell transplantation (sct) from maternal or collateral donors with intensified conditioning including g-csf and atg, followed by two lower doses of pt/cy ( . mg/kg × doses). outcomes were compared with those of controls from matched-pair analysis who undergone haploidentical sct from other donors than mother or collateral relatives at the same time period. a total of patients with myelodysplastic syndrome (mds) or leukaemia undergoing haploidenticla sct from maternal or collateral donors were enrolled in the study. incidence of grade ii-iv and grade iii-iv acute gvhd at day were comparable between the study group and the control group ( . % vs. . %, p = . ; . % vs. . %, p = . ). incidence of cmv and ebv reactivation at day were also comparable between the study group and the control group ( . % vs. . %, p = . ; . % vs. . %, p = . ). after a median follow-up of days and days, the incidence of trm and relapse at year were comparable between the study group and the control group ( . % vs. . %, p = . ; . % vs. . %, p = . ); the probability of overall survival and lfs at year were comparable between the study group and the control group ( . % vs. . %; p = . ; . % vs. . %, p = . ). in conclusion, conditioning with atg and low-dose pt/ cy might be a feasible option for patients undergoing hla haploidentical, t-cell replete sct from maternal or collateral donors. trial registration: the study is registered at www. clinicaltrial.gov as nct . disclosure of conflict of interest: none. hematopoetic stem cell transplantation (hct) is a lifesaving treatment option for eligible patients with hematological malignancies. hct is inherently associated with a risk of nonrelapse mortality that varies greatly depending on transplant and patient characteristics. the assessment of the risk of complications and mortality before the procedure is extremely important. the hct comorbidity index (hct-ci) introduced by sorror m. is one of the tools proved to predict hct outcomes and was shown to be significant in various disease and hct settings. the objective is to evaluate hct-ci index of hct recipients, determine impact of different variables on ci score, particularly those, showing pulmonary and cardiac function. data of hct-ci of autologous (auto) and allogeneic (allo) hct recipients, transplanted during period january -october were analyzed. impact of pulmonary and cardiac function values on ci score was evaluated: dlco (diffusing capacity of the lung for carbon monoxide), fev (forced expiratory volume) and ef (cardiac ejection fraction) are parameters, reflecting pulmonary and cardiac function, which values are included into hct-ci score. the statistical data analysis was conducted using spss program. the differences were considered statistically significant at p ≤ . . records of allo and auto hct recipients, transplanted during . - . in vilnius university hospital were revised. median age of allo hct and auto hsc recipients was ( - ) and ( - ) years respectively. main indication for allo hct was acute myeloid leukemia ( %) patients and for auto hct -multiple myeloma ( . %) patients. hct-ci was completely calculated (no values missing) in allo and auto hct recipients. only patients with available complete hct-ci data were further analyzed. hct-ci in hct recipients was as shown in table . hct-ci score o was calculated in ( . %) and ≥ in ( . %) allo hct recipients. hct-ci score o was calculated in ( . %) and ≥ in ( . %) auto hct recipients. hct-ci score did not differ statistically significant between male and female recipients in both hct categories as well as in different age groups of patients (below and above years in allo and below and above years in auto hct). dlco was found to be below normal values (o %) in ( . %) allo hct and in ( . %) auto hct recipients. fev was less affected and found to be lower % in ( . %) allo hct and in ( . %) auto hct recipients. ef below % detected in ( , %) allo hct and in ( . %) auto hct recipients. low dlco was found to cause the greatest impact on hct-ci score and was statistically significantly associated with higher hct-ci (po . ). the most common hct-ci in both hct groups was score . dlco was found to be below normal ranges in relatively large patient group and had the greatest impact on hct-ci score. further studies on reasons of pulmonary function impairment and it's impact on hct outcomes are warranted.[p ]disclosure of conflict of interest: none. haemophagocytic lymphohistiocytosis (hlh), a life-threatening hyper-inflammation syndrome, is classified into primary and secondary forms. primary hlh is caused by gene mutations resulting in impaired cytotoxicity of natural killer (nk) cells and cytotoxic t lymphocytes (ctls). secondary hlh arises in the setting of autoimmunity, infection, malignancy, or less commonly, may be idiopathic. treatment of hlh has two major goals: halting the triggering event and controlling the overactive immune system. however, patients with primary or recurrent secondary hlh should subsequently undergo allogeneic hct for long lasting disease remission. we retrospectively evaluated hematopoietic stem cell transplantation (hsct) might be a valid treatment option for adults suffering from aggressive t-cell malignancies providing long term disease control. since a suitable hla-matched donor cannot be identified for all patients (pts) in need for transplantation, alternative donors graft sources such as related hla-haploidentical donors are considered. through introduction of t-cell-replete (tcr) hlahaploidentical transplantation (haplo-hsct) using post transplantation cyclophosphamide (ptcy) successful treatment with low non-relapse mortality rate (nrm) has been observed in lymphoma patients (luznik et al., bmt, ) . however, less data are available on the outcome of this haplo-approach in the treatment of t-cell malignancies, in particular when disease is refractory. we retrospectively evaluated the outcome of haplo-hsct using tcr grafts and ptcy in pts with peripheral t-cell lymphoma treated between and at our institution (t-nhl = , t-all = ; male n = ; median age: years). disease was refractory/active at time of transplantation in pts, while one had achieved second cr. all patients received at least prior treatment lines and one patient failed previous allogeneic transplantation. while fludarabine and cyclophosphamide served as backbone for conditioning, pts received a tbi-based and a drug-based conditioning regimen which was myeloablative in %. if disease was active at time of haplo-hsct, a sequential therapeutic concept was performed involving intensive chemotherapy (clofarabine n = ) shortly preceding conditioning (zoellner ak et al., bmt, ) . post-grafting immunosuppression consisted of cyclophosphamide, tacrolimus and mycophenolate mofetil in all patients. graft source was bone marrow in pts. no primary graft rejection occurred; / pts engrafted, one died early in aplasia. neutrophil/platelet engraftment was achieved at a median of (range: - ) and (range: - ) days, respectively. acute gvhd grade ii-iii was observed in pts, whereas no patient developed grade iv agvhd. mild chronic gvhd occurred in one patient. % of the pts developed grade ii-iii treatment-related toxicities most commonly diarrhea ( %) and mucositis ( %); grade iv toxicity (mucositis) was observed in one patient only. no vod occurred. cmv reactivated in / pts at risk, whereas no ptld was seen. proven invasive aspergillosis was diagnosed in one patient. at day + seven pts achieved cr. pts relapsed and died (relapse n = , infection n = ). -year nrm was %. at a median follow up of months (range: - ) the estimated -year and -year overall survival (os) and progression-free survival (pfs) were %/ % and %/ %, respectively. three pts received haploidentical dlt pre-emptively (n = ) and therapeutic (n = ), leading to sustained cr in two, while no severe gvhd occurred. sequential therapy in the setting of tcr haplo-hsct using ptcy as gvhd prophylaxis is feasible, well tolerated and shows low rates of gvhd and acceptable nrm in patients with relapsed/refractory t-cell lymphoma/ leukemia providing an effective anti-lymphoma/leukemic activity. thus, we suggest that intensified tcr hapo-hsct using ptcy should be considered as an alternative for patients suffering from aggressive t-cell malignancies, lacking hlamatched donors. disclosure of conflict of interest: none. tacrolimus is a calcineurin inhibitor increasingly used as immunosuppression following allogeneic stem cell transplantation; maintenance of therapeutic serum levels is essential to reduce the risk of graft rejection and graft versus host disease. however, tacrolimus can be associated with serious side effects and potential drug interactions. regular monitoring of serum levels and appropriate dose adjustment is essential to ensure therapeutic levels and to avoid toxicity. in our adult bmt unit, an established standard operating procedure (sop) provides a prescriptive dosing algorithm for: (i) initiation of tacrolimus therapy; (ii) conversion between iv and oral routes; (iii) dose adjustment based upon tacrolimus serum level and interacting medications. we performed an audit assessing adherence to the sop dosing algorithm. inpatient tacrolimus dosing episodes from five consecutive haploidentical transplants were retrospectively analysed. episodes were excluded due to insufficient records. for the remaining episodes, tacrolimus serum levels and corresponding doses were identified. the response of the medical team to each serum level was compared with the sop dosing recommendation. to account for sensible rounding of doses, a margin of error of ± % was permitted. adherence to sop dosing was %. non-adherence to the sop ( %) was subcategorised as justifiable ( %) or unjustifiable ( %). justifiable non-autologous hsct is currently being explored for its efficacy and safety in the treatment of multiple sclerosis (ms). as more experience is gained in treating this cohort, treatment related mortality has steadily improved although the procedure still carries a degree of risk. ebv reactivation is well described in allogenic stem cell transplants although less so in autologous transplantation. we investigated the frequency of ebv reactivation in patients with ms undergoing autologous hsct at a single uk site. patients underwent autologous hsct for treatment of ms at king's college hospital between feb and aug . all were mobilised with cyclophosphamide g/ m and g-csf. were conditioned with cyclophosphamide and atg, and one with beam/atg. previous exposure to ebv (ebv igg) was assessed prior to transplant and local posttransplant ebv monitoring was performed on whole blood samples by means of quantitative pcr in patients. data was collected retrospectively. all ( %) patients were positive for ebv igg pre-transplant. overall, samples were tested for monitoring post-transplant. ( . %) patients demonstrated positive pcr post-transplant on local testing with one further patient being negative on local tests but later becoming positive on testing in their parent hospital (full results unavailable). of these , the median time to positive testing post-transplant was days ( - ). maximal ebv dna titre was reached at a median time of days post-transplant ( - ) with a mean maximum titre of . log ( . - . ). patients experienced symptomatic reactivation with an associated large paraproteinemia. one of these developed hyper-viscosity requiring plasma exchange and developed neurological symptoms mimicking an ms relapse (max ebv titre of . log). this patient received rituximab and ebv level is declining, the other was observed carefully but developed right leg weakness which is slowly improving. the patient with raised ebv at their parent hospital also received rituximab (unclear if this reactivation was symptomatic). we have developed a protocol to pre-emptively treat ebv reactivation with rituximab once a log titre is reached and one patient has so far been treated according to this. of the patients with locally confirmed reactivation who did not receive rituximab, ( %) self-resolved at a median time of days ( - ), ( . %) have ongoing re-activation ( with improving, with stable titres) and ( . %) have not had any local bloods performed ≥ months. patient with selflimiting reactivation later had a further positive titre ( days post-transplant and days post initial resolution). ebv reactivation appears to be common in patients with ms in the first months post autologous hsct. unlike in other patient groups such as aplastic anaemia patients receiving allogeneic transplants it can cause significant neurological symptoms which may be confused with ms relapse. the mechanism of this reactivation is probably related to atg administration but may be exacerbated by prior immune suppression in this heavily pre-treated group, the majority of whom have received highly active disease modifying therapies in the past. these results demonstrate the importance of monitoring for ebv reactivation following autologous hsct and the consideration of pre-emptive therapy. disclosure of conflict of interest: none. reduced bone mineral density (bmd) is a well recognised complication of hct. guidelines recommend scanning by dual energy x-ray absorptiometry (dxa) one year after transplant in all hct patients or else specific groups of high risk patients. , it is recognised that both dose and duration of steroids are risk factors for low bmd and it is recommended that prednisolone doses greater than or equal to mg/day for more than months should prompt a dxa scan. for patients with osteopenia it is recommended that calcium/vitamin d supplements are given together with lifestyle advice including diet, smoking cessation and weight bearing exercise. in this survey we have investigated the current practise in investigating and managing bone health in the context of hct. a survey was sent to all centres including countries registered with ebmt as of november . centres replied from countries. response numbers to each question were variable and are indicated by the denominators. / used a national guideline to guide their practise, and / used an international guideline. no single guideline was quoted more than once. low testosterone has been demonstrated to be an independent determinant of endothelial (dys)function in men. graftversus-host disease (gvhd) is a major contributor to nonrelapse mortality (nrm) after allogeneic stem cell transplantation (allosct). vulnerability of the recipients' endothelial cell system is a novel concept to explain why a proportion of patients with acute gvhd fail to respond to escalating immunosuppressive therapy and ultimately succumb to gvhd and related complications. this retrospective study investigated the prognostic impact of pre-transplant testosterone levels on nrm after allosct in male patients. between and , a total of male patients undergoing allosct at heidelberg university (median age years) provided informed consent to participate in this observational study (training cohort). a total of patients ( %) received transplants from related donors (rd). diagnoses were aml ( %), mds ( %), lymphoid malignancies ( %) and multiple myeloma ( %) . a total of patients ( %) received statin treatment post allosct as per institutional standard policy. for validation, an independent patient cohort of men allografted for aml and mds (median age years, % rd, no statin treatment) at essen university was analysed. pretransplant serum samples were collected between and months before allosct and cryopreserved at − °c. testosterone and suppressor of tumorigenicity- (st ) levels were measured by radioimmunoassay and elisa, respectively. median pre-transplant testosterone level in the training and validation cohort was . nmol/l (range: . - . nmol/l) and . nmol/l ( . - . nmol/l), respectively. in the training cohort, lower pre-transplant testosterone as continuous variable was associated with shorter os (p = . ). lower testosterone levels showed a trend towards higher nrm (p = . ) and a significant association with nrm after onset of acute gvhd (p = . ). multivariate analysis confirmed lower pre-transplant testosterone levels as a significant predictor of an increased nrm risk after gvhd onset (p = . ). in the subgroup of patients not receiving statins post-transplant, lower testosterone levels were associated with increased incidence of transplant-associated microangiopathy (p = . ), and, in addition, with higher pre-transplant st levels indicating endothelial vulnerability. in the validation cohort, similar results with regard to overall survival (os, p = . ), nrm (p = . ), nrm after acute gvhd onset (p = . ) in univariate analysis, and to nrm after gvhd onset (p = . ) in multivariable analysis could be observed. the association of pre-transplant testosterone levels (in quartiles) and incidence of nrm after gvhd onset in the training and validation cohort is depicted in figure a and b, respectively. our study suggests that low pre-transplant testosterone is associated with serological and clinical evidence for endothelial damage and is an independent risk factor for a fatal outcome of gvhd. prospective studies in the allosct setting investigating testosterone and testosterone supplementation in deficient patients are highly warranted. disclosure of conflict of interest: none. nk cells anti-tumor ability in multiple myeloma patients s tognarelli , , b jacobs , , , i von metzler , h serve , p bader, t klingebiel , a mackensen and e ullrich , department of pediatric stem cell transplantation and immunology, childrens hospital, goethe university, frankfurt, germany; cellular immunology, loewe centre for cell and gene therapy, goethe university, frankfurt, germany; department of hematology and oncology, university hospital erlangen-busulfan is one of essential drugs for hematopoietic stem cell transplantation (hsct). because of its narrow therapeutic range: targeted busulfan using therapeutic drug monitoring (tdm) has been used. generally, the initial dose of busulfan is determined by patients' body surface area as mg/m except for infants ( mg/m ). however, pharmacokinetic evidence of these initial doses is scarce. therefore, we investigated the full pharmacokinetics of busulfan in infant and child, and attempted to validate that these initial doses are acceptable. one hundred ninety-five pediatric patients undergoing hsct using four-day targeted busulfan were enrolled. of them, patients received hsct when their age was ≤ year old (infant group [ig]), and patients received when - years old (toddler group [tg]). the remaining patients were defined as a child group (cg). busulfan was administered intravenously once daily for consecutive days. tg and cg received mg/m as the first dose, and ig received mg/m . using daily tdm, we adjusted the next dose of busulfan. target daily and total area under the curve (auc) were μg*h/l/day and - μg × h/l, respectively. median first-day busulfan auc of ig, tg, and cg were , and μg × h/l, respectively, which was significantly different (p = . ). however, there was no significant difference in median total busulfan auc (ig; , tg; , and cg; μg × h/l, respectively, p = . ). the coefficient of variance (cv) of four-day busulfan aucs in ig and cg was similar (median cv: . % and . %, respectively), whereas cv of tg was . %. in sub-analysis of tg and cg who received equally mg/m as the first dose, there was an inverse correlation between age and first-day busulfan auc (r = − . , p = . ), as well as between age and cv of four-day busulfan aucs (r = − . , p = . ). initial busulfan dose as mg/m for infant could be acceptable in aspect of first-day auc and cv of four-day busulfan aucs. however, higher first-day auc and cv were shown in tg. although target total busulfan auc could be achieved safely by tdm, we suggest that reduction of initial dose less than mg/m is also necessary to patients with - years old to lower the relatively higher first-day auc. taken together, tdm is highly recommended to reduce busulfan toxicity, especially in younger children. disclosure of conflict of interest: none. post-induction treatment strategy of acute myeloid leukemia (aml) is currently driven by european leukemia net (eln) risk assessment at diagnosis. if it is well established that patients belonging to favourable-risk group can be treated with chemotherapy and/or autologous stem cell transplantation (sct) and that those belonging to the unfavourable-risk group should be addressed to allogeneic (allo) sct, for patients included in the intermediate-risk groups the best post-induction treatment has not been established yet. we report here a years ( - ) allo-sct single center experience in aml patients. median age was years (range: - ), %, %, % and % were grouped in the eln favourable, intermediate-i, intermediate-ii and unfavourable risk category, respectively and % of the patients were allotransplanted in advanced disease-phase ( nd complete remission). half of the patients received a sibling hla compatible donor, % of the cases received peripheral blood stem cells and half of the patients received a myeloablative conditioning regimen. graft versus host disease prophylaxis was conventionally based on cyclosporine and shor-course methotrexate, with the addition of antilymphocyte immunoglobulin in case of matched unrelated donor. the clinical and transplant characteristics of the patients according to the eln-risk group were well balanced. with a median follow up of months (range: - months), the projected years overall survival (os) and disease free survival (dfs) is % ( % ci: - %) and % ( % ci: - %). the median os and dfs in favourable/intermediate-i vs intermediate-ii/unfavourable is . and . months ( figure a ; p = , ) vs and , months ( figure b ; p = . ). the relapse rate (rr) and the non relapse mortality (nrm) at two years are % ( % ci: - %), and % ( % ci: - %), respectively. non differences were observed comparing the years rr and the years nrm of patients in the favourable/intermediate-i vs intermediate-ii/unfavourable eln risk group ( % vs %; p = . and % vs %; p = . ). interestingly, the percentage of patients allotransplanted in advanced phase of the disease was higher in those included in low/intermediate-i with respect to intermediate-ii/unfavourable eln-risk group ( % vs %; p = . ). our data suggest that allo-sct can cure approximately - % of aml patients, with no difference within the eln risk groups. disease recurrence remains the major problem and this is highly correlated to the percentage of patients in advanced phase of the disease at transplant, particularly in eln favourable/intermediate-i patients. we are currently collecting the data on minimal residual disease (mrd) status of these patients during chemotherapy and before transplant using moelcular biology on target genes and/or multiparametric flow cytometry on leukemia associated immunophenotype, in order to assess if the prognosis of these patients may be refine by the prospective application of mrd data.[p ]disclosure of conflict of interest: none. outcome of allogeneic stem cell transplantation for patients with high-risk acute leukemia according to donor type and graft-versus-host disease prophylaxis s lindner, t berg , j riemann , s ajib , z jedlickova , s gueller , f lang , a sackmann , , n goekbuget , , h martin , a bacigalupo , h serve , and g bug department of medicine ii, hematology and oncology, university hospital frankfurt, goethe university, germany; s german cancer consortium (dktk), german cancer research center, heidelberg, germany and università cattolica del sacro cuore, fondazione policlinico universitario gemelli, roma, italyin high-risk acute leukemia (hr-al), allogeneic hematopoietic stem cell transplantation (hsct) is the only potentially curative treatment. increasingly, hsct is being performed utilizing alternative donors. we retrospectively analyzed the outcome of consecutive patients (pts) with hr-al (aml/all, n = / ) undergoing first allogeneic hsct in our transplant unit between / and / according to donor type and graft-versus-host disease (gvhd) prophylaxis: in the matched related donor group (mrd, n = ), hsct was performed with standard immunosuppression (is), that is, calcineurin inhibitor (cni) plus methotrexate or mycophenolate mofetil (mmf). for / hla-allele matched unrelated donors ( / mud, n = ) or / hla-allele mud ( / mud, n = ) we used is and anti-thymocyte globulin (atg fresenius/neovii). hsct with a haploidentical family donor or an / hla-allele mismatched unrelated donor was performed using is with cni plus mmf and post-transplant cyclophosphamide (pt-cy, n = ). a myeloablative (n = ) or reduced-intensity (n = ) conditioning regimen was applied in complete remission (cr, n = ) or active disease (n = ). pts had a median age of years (range: - ) and hematopoietic cell transplantation comorbidity index of (range: - ). patient and treatment characteristics were well balanced between the groups except for a higher percentage of pts transplanted in cr in the pt-cy group ( % vs. - %, p = . ). peripheral blood stem cells were preferred for mrd, / mud and / mud ( %, % and %, respectively) and bone marrow for % of pt-cy based hsct. all pts engrafted. with a median follow-up of months (range: - ), probability of overall survival (os) at years was ± % for the mrd, ± % for the / mud, ± % for the / mud and ± % for the pt-cy group, without significant differences (p = . ). however, the probability of achieving the combined endpoint gvhd-and relapse-free survival (grfs) at years varied significantly between the groups (mrd ± %, / mud ± %, / mud ± % and pt-cy ± %, po . ), reflecting the high cumulative incidence (ci) of chronic moderate and severe gvhd at year in the mrd ( ± %) as opposed to the other groups ( / mud ± %, / mud ± % and pt-cy ± %, p o . ). of note, donor type had no impact on ci of transplant-related mortality (trm) at years ( ± %), acute gvhd g - at day + ( ± %) or leukemic relapse at years ( ± %). overall, aml pts years of age had a significantly inferior relapse-free survival compared to younger pts ( ± % vs. ± %, respectively, p o . ) without a higher ci of trm (p = . ). median time to aml relapse was months. our results suggest that pt-cy-based alternative donor hsct is safe in hr-al pts and provides a solid basis for a randomized clinical trial comparing hsct from haploidentical family donors and / mud, currently in preparation. while os did not vary between groups, grfs was dismal after mrd transplants without atg, due to high rates of severe chronic gvhd, consistent with published data. as leukemic relapse remains the major cause for treatment failure especially in elderly pts, maintenance strategies using novel drugs or cellular therapies are warranted. disclosure of conflict of interest: none. relapse following hematopoietic stem cell transplant (hsct) is the leading indication for a second transplant in patients with malignant disease. hsct has been shown to be superior to chemotherapy alone or palliative measures in these patients. for non-malignant disease a second transplant may be considered for graft failure after first transplant. data regarding the outcome of a second hsct for non-malignant disease is scarce. we retrospectively analyzed patients who underwent a second hsct, for survival and toxicity data. twentynine patients (age - years) who received a second hsct at our institution during - were included in the analysis. thirteen patients had an underlying malignancy and patients were transplanted for non-malignant indications, including inborn errors of metabolism, non-malignant hematologic diseases and immune deficiency. median follow up was months (range: - ). there were deaths ( %) in the malignant group, ( %) were due to disease relapse and ( %) were transplant related. fifty percent of deaths occurred within the first year following the second hsct. in the non-malignant group there were deaths ( %), of which ( %) were attributed to the underlying disease and ( %) were transplant related. all deaths but one occurred within the first year post hsct. treatment related mortality following second hsct is higher compared to first transplant. the higher survival rate in the non-malignant group suggests that transplant following graft failure should be considered ins patients with otherwise incurable underlying disease. though the outcome for patients with relapse of malignant disease following hsct is poor, a second transplant may benefit a subset of these patients. attempts to achieve complete remission prior to transplant should be made to improve outcome. due to the small number of patients in our cohort, further multi-center trials are needed. disclosure of conflict of interest: none. disseminated bcg infection (bcg-osis) is a rare but most serious complication in vaccinized especially immunocompromised children. severe combined immunodeficiency disorder (scid) is probably the commonest primary immunodeficiency associated with bcg-osis, though there is no such definitive data as most of the cases described in literature are in the form of reports. hematopoietic stem cell transplantation (hsct) is a life-saving treatment for patients with scid, especially if therapy is instituted early, prior to onset of infections.as bcg vaccine is routinely given to all iranian children at birth, the likelihood of having an active infection at the time of transplant would be significantly high. the main objective of this study was to evaluate the outcomes of hsct in scid patients with disseminated bcg infection . sixteen scid patients underwent hsct in our center since to , of which nine patients ( male, female) were enrolled in this analysis. all the patients had received bcg vaccination according to the national vaccination protocol, and had undergone anti-tuberculosis (tb) treatment prior to transplant due to disseminated bcg infection. the mean age at hsct was . months (range: - months). patients received bone marrow (n = ), peripheral blood progenitor cells (n = ) or umbilical cord blood grafts (n = ) from hla-matched related donors (n = ) and mismatched unrelated donors (n = ). three patients received unconditioned matched sibling donor transplants and ric regimen was provided with fludarabine, melphalan and rabbit anti-thymocyte immunoglobulin (thymoglobulin) in others . cyclosporine a and prednisolon were used as graft-versus-host disease (gvhd) prophylaxis. they also continued to receive anti-tb treatment. all patients but one engrafted. the median times to neutrophil and platelet engraftments were days (range: - ), and days (range: - ), respectively. engraftment with full chimerism ( %) occurred in patients and the other patients had mixed chimerism. with a median follow-up of months (range: - months), overall survival was . %. the main cause of death was disseminated bcg infection.three out of patients who achieved engraftment, developed acute gvhd (grade i-ii), while one patient developed extensive chronic gvhd. although anti-tb treatment continued, tuberculous dactylitis occurred in patients post-hsct that were successfully treated. on last post-hsct follow-up, patients with full chimerism and with mixed chimerism are alive and disease free. scid is called as a pediatric emergency as it invariably leads to fatality in infancy without early aggressive therapy and hsct. in hsct recipients, the impaired cellular immunity renders these patients more susceptible to infection. as previous reports suggest, our study demonstrates that with appropriate anti-tb cover, immunological reconstitution with complete recovery from bcg infection can be achieved by early hsct. disclosure of conflict of interest: none. paraproteinemia occurrence after allogeneic hematopoietic stem cell transplant as a possible marker for chronic gvhd onset f monaco , s tamiazzo , f dallavalle , l calcagno , m pini and m ladetto hematology and transfusion medicine, azienda ospedaliera ss. antonio e biagio e cesare arrigo, alessandria, italy transient monoclonal gammopathy is commonly reported after solid organ or stem cells transplant (sct) for hematologic malignancies. however the clinical significance of a paraproteinemia appearance is not fully understood, because the attempts to correlate its effect on survival rates, graft versus host disease (gvhd) occurrence and viral reactivations have led to controversial results. starting from these reports we decided to evaluate among our allogeneic transplanted patients the incidence of m-component and its possible relationship with chronic gvhd. one-hundred and one patients undergoing allosct at the hematology unit of alessandria (italy) between and were evaluated. % of patients were male and % were females. pretransplantation diagnosis included: acute myeloid leukaemia/ high-risk myelodisplastic sindromes ( %), acute lymphoblastic leukaemia ( %), lymphoproliferative disorders ( %) and other less common malignancies ( %) . patients with multiple myeloma were excluded from the study. all patients had, at least, two pre-transplantation serum electrophoresis with no evidence of pre-existing monoclonal component. serum electrophoresis was scheduled to be performed at , and days and years after transplantation. forty-nine patients were submitted to allo-sct from a sibling donor and from a matched related donor (mud); in vivo t-cell depletion with anti-thymocyte globulin was used in patients. thirty-four patients relapsed after allosct, ( %) developed chronic gvhd and patients ( %) are currently alive at the last follow-up. posttransplantation follow up ranged from to days with a median of days. paraproteins were detected in out of patients ( %), being monoclonal in patients, and bi or tri-clonal in the remaining cases; the immunoglobulin subclass most commonly observed was igg. ten-year overall survival of the whole population was %; splitting the population in two cohorts (with or without paraproteinemia) we did not detect any statistical differences in overall survival, gvhd development and relapse incidence at + and + days posttransplant; viceversa, after days, a statistically significant difference was observed in chronic gvhd occurrence in patients with or without paraproteinemia ( % vs %, respectively, po . ). ten-year overall survival curves were significantly better in patients with paraproteinemia as compared with the paraprotein-free group ( % vs %, p = . ), and an even more evident significance was seen in ten-year relapse free survival curves ( % for patients with paraprotein vs % for patients without paraprotein, p = . ). monoclonal gammopathy, also in our experience, is frequent following allo-sct. we observed a strong correlation between the occurrence of paraproteinemia, chronic gvhd and a significantly better overall and relapse-free survival. recently many evidences showed that b cells are involved in the pathogenesis of chronic gvhd (cgvhd) and anti-b-cell therapy has been suggested for the treatment of cgvhd. we speculate that the presence of a monoclonal gammopathy after allogeneic transplant is expression of the activation of the b-cell compartment. a prospective study with a larger population should be considered, in order to confirm our results and assay post-transplantation monoclonal gammopathy as an early marker for gvhd development. disclosure of conflict of interest: none.inherited bone marrow failure (ibmf) syndromes are rare pediatric disorders that characteristically associate physical abnormalities, progressive bone marrow failure and predisposition to cancer. the most common of these disorders is fanconi anemia (fa). stem cell transplantation (sct) using related or unrelated donors are the only curative therapeutically approach when severe marrow failure is established. the aim of the study was to analyze the results of sct for patients with ibmfs in a single center. we performed a retrospective study in pediatric patients with ibmf admitted in pediatric hematology and bone marrow transplant department, fundeni clinical institute between january and september . diagnosis and severityof ibmfs were established based on hematological results, bone marrow biopsy and clinical findings. genetic testing for ibmfs is not currently available in our country. indication for sct was established when patients developed moderate/severe aplastic anemia and became transfusion dependent.in case of dba, sct indication was established for steroid resistant disease. the donors were selected from family members or unrelated donors, / matched.the conditioning regimens used were reduced intensity (fludarabine - mg/m , cy mg/kg, f-atg mg/kg) for af, dc and myeloablative (busulfan i.v., fludarabine mg/m , thiotepa mg/kg, f-atg mg/kg) for dba. gvhd prophylaxis consisted of standard methotrexate and csa/tacrolimus. all parents signed informed consent forms. in our center, between and , patients with ibmf were diagnosed: ( %) patients with fa, ( %) patients with diamond blackfan anemia (dba), ( %) patients with diskeratosis congenita (dc), and ( %) patients with not classifiable ibmfs. the patient data is available in table . seven out of patients ( %) performed sct procedures: sibling patients ( patients with af, patient with dc), mud patients ( patients with af, patient with dba). all patients ( %) engrafted for pmn (median = , range: - days) and platelet (median , range: - days). / ( %) presented reactivation of cmv and received valganciclovir, / developed cmv disease (encephalitis and pneumonia), / ( %) developed bkv cystitis and required extensive hydration and levofloxacin. / ( %) developed grade i-ii skin acute gvhd day + , which responded to topical treatment and low dose of corticosteroids. / ( %) developed grade iii intestinal acute gvhd, which responded to high-dose corticosteroids. / ( %) developed grade iv intestinal chronic gvhd (day + ), without response to high-dose corticosteroids, mmf and later died on day + , due to infectious complications (severe pulmonary and cerebral aspergillosis). / patients ( %) are alive, with % donor chimerism / ( %) or stable mixed chimerism / ( %). median follow-up for sct patients was days ( days- y mo). conclusions in our study we observed a low incidence of severe complications associated with low mortality rate ( %) . sct is a procedure that associates multiple risk situations, but it remains the only curative cytomegalovirus (cmv) infections remain a significant cause of morbidity and mortality in patients whose immune systems are compromised, including hematopoietic stem cell transplant (hsct) recipients. although the adoptive transfer of third party cmv-specific t cells has proven both safe and clinically beneficial in treating even drug-refractory infections/disease, broader implementation and commercialization of this strategy has been hampered by (i) the postulated need for extensive cell banks generated from donors representing diverse hla profiles, and (ii) lack of large scale t cell manufacturing processes. to address these limitations we have developed a proprietary decision tool (cytomatch™) to identify a small panel of healthy donors who should provide almost universal hla coverage; and optimized a simple, scalable manufacturing process to generate large numbers of cmv-specific t cells. to assess the robustness of our strategy we generated a bank of cmv-specific t cells (viralym-c™) from carefully selected healthy donors. the lines were polyclonal, comprising both cd + ( . ± . %) and cd + ( . ± . %) t cells, expressed central cd ro+/cd l+ ( . ± . %) and effector memory markers cd ro+/cd l-( . ± . %), and were specific for the immunodominant cmv antigens ie and pp (ie : ± ; pp ± sfc/ × , n = ). a fixed-dose ( × cells/m ) phase i clinical trial was subsequently initiated to test the safety and efficacy of these "ready to administer" t cells in pediatric and adult hsct recipients with drug-refractory cmv infections. using our bank of just lines, we have identified a suitable line for of patients screened. of these, patients have been treated with viralym-c cells; received a single infusion and patient required infusions for sustained benefit. there were no immediate infusion-related toxicities; and despite the hla disparity between the viralym-c™ lines and the patients infused, there were no cases of de novo or recurrent graft versus host disease (gvhd). based on viral load (measured by quantitative pcr) and/or symptom resolution, viralym-c cells controlled infections in all patients with complete (cr) and partial responses (pr) achieved within weeks of infusion. one patient with cmv retinitis had complete resolution of symptoms following viralym-c™ infusion. our results demonstrate the feasibility, preliminary safety and efficacy of "ready to administer" viralym-c™ cells that have been generated from a small panel of healthy, eligible cmv seropositive donors identified by our decision support tool. these data suggest that cost-effective, broadly applicable t cell anti-viral therapy may be feasible for patients following hsct and potentially other conditions. disclosure of conflict of interest: drs. juan vera, ann leen and brett giroir hold equity and drs. ifigeneia tzannou, sunitha kakarla are employed by viracyte. haploidentical stem cell transplantation (hsct) protocols utilizing ex vivo t-cell depleted grafts have been proven efficient in preventing graft versus host disease (gvhd), but cause a delay in early t-cell recovery that increases the risk of graft rejection, leukemia relapse and viral infections. conventional donor lymphocyte infusion (dli) after hsct transplantation is conditioned because of the high prevalence of gvhd even with low dose of t cells. here we present preliminary data of escalating cd ro+ memory t cells as dli in three patients that received a selective graft depleted of naïve (cd ra+) t-cells. three children that were transplanted following nonmyeloablative conditioning regimen with a graft consisting of cd + and cd ra-cells, with mixed chimerism, lymphopenic and viral/opportunistic infections and minimal residual disease positive before hsct received dose scalating cryopreserved haploidentical cd ra-memory t cell starting with a initial dose of × /kg, until a maximal dose of × /kg with a days interval. we infused products with a naïve ( ra+) t-cell dose less than × /kg with . % purity of cd + cd ro+ memory t-cells in all cases. all infusions were well tolerated without any side effect during infusions neither gvhd. following the dli, a progressive increase in t cell counts was observed. our preliminary data suggest that dose escalating of haploidentical memory t cells ( ro+) as dli provides a safety platform, even with high dose of t cells ( × /kg), for adoptive immunotherapy in haploidentical ra+ depleted grafts with no gvhd complications, and allows an increase in t cell reconstitution. however, efficacy of this strategy requires longer studies. relapse after allogeneic hematopoietic stem cell transplantation (allohsct) remains a major therapeutic challenge: outcome is very poor, without curative option in most cases. second allohsct may be considered in few selected patients because of anticipated limitations: ( ) donor availability; ( ) high toxicity due to previous treatments; ( ) low efficacy considering the very advanced disease situation. we hypothesized that the use of post transplantation cyclophosphamide (pcy) haplo-sct after relapse following allohsct may deal in part with these limitations. in particular, the presence of full haplotype hla mismatch could provide a decisive antileukemic effect relative to alloreactivity. in absence of large series in this setting, we report here the outcome after haplosct for patients who relapse after a first allohsct. we retrospectively studied adult patients, who received a second pcy haplo-sct for hematological malignancies. patients were treated between and . the objective was to assess both the feasibility and the efficacy of haplosct in this setting. twenty seven patients were included: median time between first allohsct and relapse was months (range: - ). median age at second transplantation was years old (range: - ). most of patients had acute myeloid leukemia (n = , %) or hodgkin lymphoma (n = patients, %). fifteen the impact of minimal residual disease and its kinetics prior to different types of allogeneic hematopoietic stem cell transplantation on clinical outcomes in patients with acute myeloid leukemia z xiao-su , , l yan-rong , yan-hong , p xu-ying , qian-jiang , hao-jiang , lan-ping, xu , xiao-hui zhang , , yu-wang , , h xiao-jun , and c ying-jun this study investigated the impact of minimal residual disease (mrd) and its kinetics prior to different types of allogeneic hematopoietic stem cell transplantation (hsct) on clinical outcomes in patients with acute myeloid leukemia (aml) in complete remission (n = ). patients who received unmanipulated haploidentical hsct and patients who received hla-matched sibling hsct were enrolled. mrd measured using -color flow cytometry (fcm) at fixed time points before transplantation was retrospectively analyzed. the patients were divided into four groups based on mrd kinetics before transplantation: consistent negative, positive to negative, negative to positive and consistent positive. during the follow-up, total twenty ( . %) patients underwent relapse. through unique variate analysis, none of mrd status at various time points before unmaipulated haploidentical transplantation was associated with clinical outcomes, as well as the dynamic change of mrd before hsct (p . ), although the patients with consistent positive mrd before hsct seemed to have a relatively higher incidence of relapse (p = . ). one-year cumulative incidence of relapse (cir) were . ± . % vs. . ± . % in mrd consistent negative and consistent positive groups, respectively (p = . ). however, patients with positive mrd after the second chemotherapy or pre-mrd before hla-matched sibling hsct showed a significant poor outcomes including higher cir (p = . and both neuroblastoma (nrb) and rhabdomyosarcoma (rms) in childhood are the aggressive malignant disease with higher mortality. this paper aims to study the efficacy of autologous peripheral blood stem cell transplantation (apbsct) in the treatment of high risk advanced nrb and rms. patients with high-risk stage iv nrb and patients with advanced childhood rms were treated by apbsct in our hospital from october of to may of . in the subgroup of nrb patients, patients got complete remission (cr) and patient got cru while patients had tumor residual disease after intensive induction therapy before asct. the median age was . ( - ) years old. primary sites of the tumors included submaxilla (n = ), cervical (n = ), adrenal gland (n = )and retroperitoneal (n = ). the conditioning regimen consisted of busulfan and melphalan (busulfan mg/kg × d, melphalan mg/m x d) or cem regimen (carboplatin mg/m × d, etoposide mg/m × d, cyclophosphamide mg/ m × d); the pathology of stage iii childhood rms patients was embryonal rhabdomyosarcoma. there were cases in cr and case in partial remission (pr). the median age was . ( - ) years old. primary sites of the tumors included bladder (n = ), left forearm (n = ), retroperitoneal (n = ), pelvic (n = ) s and talus (n = ). the conditioning regimen consisted of melphalan, cyclophosphamide and dactinomycin (melphalan mg/m × d, cyclophosphamide mg/m × d, dactinomycin . mg/kg × d).there were double apbsct cases (nrb n = , rms n = ). all the relapse patients were treated with chemotherapy and radiation therapy. all the patients successfully underwent mobilization, collection and reinfusion. the time of hematopoietic reconstitution was ( . ± ) days, no severe toxicity was observed, no transplant-related death was found. with a median follow-up of . ( - ) months, one of the patients was lost to follow-up. in the subgroup of nrb patients (n = ): the -year event-free survival and total survival rate of all patients were . % and . %, respectively. the survival time of no recurrence was significantly different between the double transplantation group and single transplantation group (p o . ). in the subgroup of rms patients (n = ), patient died, patients live without pd( patients had double apbsct), patients suffered recurrence but still alive. apbsct achieved good outcome in patients with high risk advanced nrb and rms. transplantation-related toxicities were tolerable. double apbsct significantly improved the depth of remission. disclosure of conflict of interest: none. transplantation outcomes of a once-daily intravenous busulfan and fludarabine conditioning for allogeneic hematopoietic stem cell transplantation in pediatric aml and high risk mds: single center experience in korea y-t lim, e-j yang and k-m park department of pediatrics, pusan national university children's hospital, yangsan, koreathere have recently been some reports suggesting that oncedaily intravenous busulfan as a conditioning regimen for hematopoietic stem cell transplantation (hsct) possibly reduces the toxicities without influencing the clinical outcome as compared with the traditional times daily dosage schedule. but until recently there has been little research and limited data available on the safety and efficacy of oncedaily intravenous busulfan and fludarabine in pediatric allogeneic hsct. we report the outcomes for allogeneic hsc recipients, evaluating engraftment status, regimen related toxicities (rrt), and event free survivals (efs) after use of oncedaily intravenous busulfan and fludarabine conditioning for allogeneic hsct in children with aml and high risk mds in a single pediatric center of korea. from january to december , aml and high risk mds children who received once daily iv busulfan/fludarabine based conditioning regimen for allogeneic hsct were reviewed, bu/flu ± atg consist of intravenous fludarabine ( mg/m ) and busulfan ( ~ mg/m , once daily iv) on days - to - , and antithymocyte globulin (atg) ( mg/kg) on days - to - . all patients received tacrolimus and mini-dose methotrexate ( mg/m( )) for graft versus host disease (gvhd) prophylaxis. boys and girls were enrolled with median age of . years (range: . - . years). the median period from diagnosis to transplantation was months (range: - months). more than half of the patients had a matched sibling donor (n = , %), % patients (n = ) had a matched unrelated donor, % patients (n = ) had a mismatched unrelated donor, and the remaining patient had a mismatched family donor. as a stem cell source, peripheral blood stem cells (pbsc) were cases ( %), bone marrow and cord blood were cases in each. the median follow-up for patients was months. the median number of infused total nucleated cells and cd + cells except cord blood transplantation were . × ( )/kg and . × ( )/kg. all patients including who received cord blood were successfully engrafted. the median time to absolute neutrophil count (anc) recovery (anc × ( )/l) and platelet recovery (platelet , × ( )/l) were days, days in each. the incidence of acute gvhd was . %, while severe grade iii/iv gvhd was observed in only patient ( . %). there were only two cases ( . %) of extensive chronic gvhd in this study. transplant-related toxicities were acceptable, there was no case with cns toxicity, eleven patients ( . %) developed grade ii,iii mucositis and grade i-iii hepatic toxicity in twenty four ( . %), but transient. there was clinically diagnosed veno-occlusive disease (vod), but most recovered by fluid restriction and diuretics. nine patients ( %) showed positive cytomegalovirus (cmv) antigen/pcr but only one patient developed cmv colitis. eight patients died: due to relapse/disease progression, due to extensive chronic gvhd. the -year efs and overall survival were . % and . % respectively. at year, the cumulative incidence of relapse was . %. overall, once-daily intravenous busulfan and fludarabine was less toxic and effective as conditioning regimen in aml and high risk mds patients undergoing allogeneic transplantation in children. disclosure of conflict of interest: none. haploidentical stem cell transplantation from unmanipulated graft has becoming a practiced option for high risk hematological malignancies who lack a matched related or unrelated donor. lack of a matched sibling or unrelated donor (mud) can be a significant barrier to allogeneic transplantation in patients who stand to benefit from this procedure. hlahaploidentical donors are readily available for nearly all such patients. haplo transplantation has inherent advantages over mud transplantation including the lower cost of graft acquisition, greater availability of donors for ethnic minorities, and immediate access to the donor in patients in whom delay cerebral palsy (cp) is a heterogeneous group of conditions that result in permanent motor disability. it may occur due to perinatal hypoxic insults, developmental brain abnormalities, genetic diseases, traumatic or infectious causes. in general the condition is non-progressive, but improvement over time is rarely seen. various treatment methods have been used for the management of this disorder. however, there has been no absolute cure for cp. the ultimate goal of stem cells therapy is to use the regenerative capacity of the stem cells causing a formation of new tissues to replace the damaged tissue. the polish stem cell bank (pbkm) has provided wharton's jellyderived msc (wj-msc) for medical therapeutic experiment application in children with cp. wj-msc from third party donors were administered to patients (pts) with cp aged from . / to . / (median age: years and month). twenty two pts have received infusions intravenously (i.v.), pt intrathecally (i.t.), and pts via both routes (first i.v., next i.t.). the cells were previously collected from healthy newborns, processed, screened for bacterial contamination as well as endotoxin content, and frozen in liquid nitrogen vapour. msc immunophenotype was confirmed using flow cytometry assay. the pts have received from to infusions in intervals from weeks to months. median i.t. dose was × cells per infusion, while median i.v. dose was × cells/kg of body weight per infusion. each patient has been examined by the same neurologist at the day of each infusion and the result of examination has been described in a follow-up. twelve patients were diagnosed with epilepsy as comorbidity. eighteen pts ( %) showed positive changes in neurological examination after their treatment with wj-msc. almost half of the children experienced improvement of cognitive functions ( out of pts). muscle tension was reduced in pts. improvement in the ability to concentrate, better contact with others and improved social interactions were observed in % of pts. correction of motility was noticed in pts, pts have experienced better quality of sleep. in cases there has been a reduction in the number of epileptic seizures ( pt even discontinued some of his medicines). there were no s noticeable changes in neurological examination of patients. seven follow-up forms have been not received yet. the experiment data provide evidence that third-party donor wj-msc are suitable and efficient stem cells for treatment in patients with cp. however further and more extensive examination, with a greater number of patients is needed, which will be beneficial for far-reaching results. spina bifida (sb) is a congenital malformation resulting from failure of fusion in the spinal neural tube during embryogenesis. despite surgical repair of the defect, most patients who survive with spina bifida have multiple system damage due to neuron deficiency in the spinal cord. it has been confirmed that the mesenchymal stem cells (mscs) have the ability to survive, migrate and differentiate into cells of a neural lineage. wharton's jelly-derived mscs (wj-mscs) from third-party donors have high proliferation and differentiation potential along with non-immunogenic features, thus seem to be a promising stem cell source. the polish stem cell bank (pbkm) has provided wj-msc for clinical application in a medical therapeutic experiment for children with sb. eleven patients (pts) were qualified for administration of wj-mscs. three pts have been waiting so far for their therapy after bioethical committee approval. seven pts were in the middle of stem cell therapy (after or injections), pt had finished one cycle of stem cell therapy ( injections -ijs) and resumed therapy by administering a first dose of wj-mscs. the cells were previously collected from healthy newborns, processed, screened for bacterial contamination as well as endotoxin content, and frozen in liquid nitrogen vapors. six pts have received infusions intravenously (median dose: . × /kg body weight per infusion), and pt was given injection of × cells intrathecally. each patient has been examined by the same neurologist at the day of each infusion and the result of examination has been written in a follow-up. there were pts, who received at least doses of wj-msc, and all of them showed positive changes in neurological examination. the important improvement, declared by pts, was in areas: pronunciation and/or self-reliance ( pts), movement of arms and/or legs ( pts), quality of life ( pts), core stabilization ( pt). only one adverse event occurred after third injection of wj-msc: pt had nausea and a fever. in case of other pts it was too early to provide reliable feedback. the transplantation of wj-mscs could stimulate the mscs to differentiate towards sensory neurons. this could be one of the reasons of observed improvement of many vital functions in patients, after mscs treatment. this approach might have value in the experimental treatment of sensory neuron deficiency in spina bifida. key: cord- - qib authors: nan title: the th annual meeting of the european society for blood and marrow transplantation: physicians – poster session date: - - journal: bone marrow transplant doi: . /s - - - sha: doc_id: cord_uid: qib nan background: allogeneic hematopoietic stem cell transplantation is routinely offered to patients with high-risk or advanced all in the hopes of improving outcomes. use of truly non-myeloablative (nma) conditioning reduces toxicity in other contexts but outcome data for all patients after nma transplants is lacking. we report the outcomes of patients with all transplanted using a nma conditioning without t cell depletion. methods: first transplant patients between october and june were reviewed. these were consecutive patients until then only those considered unfit for fmc conditioning as per the ukall protocol. all patients were conditioned with fludarabine mg/m /day for days and cyclophosphamide g/m /day for days. short course mtx and ciclosporin were used for gvhd prophylaxis. standard supportive care was employed. thirty-one patients with a median age of ( - ) met the criteria for this case review. had b-all and were philadelphia chromosome positive. patients ( %) had high risk disease by standard diagnostic criteria. ( %) were in first complete remission (cr ). matched sibling donors were used in instances with the remaining being fully matched unrelated donors. % of patients had a hct-ci score of , % a score of or with patients having a score of or higher. median cd dose was . x /kg ( . - . ) with a median cd dose of . x / kg ( . - . ) results: trm was low at % at year and % at and years respectively. no factors included in a univariate analysis (which included age, diagnosis, disease status, hct-ci, donor type, cmv risk and cell dose) significantly impacted trm. the incidence of classical acute (a) gvhd grade - and - was % and % by day and % and % by day if late onset agvhd is included. out of eligible patients developed chronic gvhd of any stage. relapse incidence was low ( % at years in all patients, % in cr patients) and was not impacted by any pre-transplant factors including positive mrd post phase induction (present in patients). notably, in univariate analysis relapse was significantly lower in patients who developed chronic gvhd. background: allogeneic stem cell transplantation (allosct) is the treatment of choice for many patients (pts) suffering from acute myeloid leukemia (aml). the graft vs. leukemia effect (gvl), applied by immunocompetent cells of donor origin, is the most important effector mechanism for the eradication of leukemia, the presentation of leukemic or allospecific antigens by malignant blasts is regarded as a crucial trigger for an effective allogeneic immune response. conversely, insufficient stimulatory capacity by the leukemic blasts is thought to be a relevant escape mechanism from cellular immunotherapy (allosct or donor-lymphocyte infusion (dli)). the purpose was to test, whether the ability of malignant blasts to differentiate in vitro towards dendritic cells of leukemic origin (dc leu ) is associated with response to allosct or outcome after immunotherapy (second allosct or dli) for post-transplant relapse in aml. methods: leukemic blasts were isolated from peripheral blood (pb) or bone marrow (bm) samples of aml patients before allosct (n= ) or at relapse after allosct (n= ). a panel of different assays was used to generate dc leu in vitro ( of them containing gm-csf). finally, in vitro results were correlated with clinical characteristics and outcome of patients treated with donor lymphocyte infusion and/or allosct. results: dc leu could be generated in vitro from all samples. when correlating proportions of dc-subtypes generated ex vivo with clinical data, significantly higher mean proportions of dc leu in the dc-fraction were found in responders vs. non-responders to immunotherapy ( . % vs . %,p= . , range: %- %). vice versa, the chance for response to immunotherapy was significantly higher, if a dc leu /dc ratio of >= % could be reached in vivo (p= . ). those patientswere characterized by a longer time to relapse (p= . ) and by a higher probability for leukemia-free survival (p= . ). similarly, generation of higher amounts (> %, p= . ) of dc leu in the mnc-fraction, and generation of more mature dc (> % cd +, p= . using the best gm-csf containing assay) were associated with a longer time to relapse in the respective patients. moreover, overall survival was improved, if > % dc leu /dc could be generated with the best gm-csf containing assay (p= . ). conclusions: in vitro generation of dc/dc leu from leukemic blasts obtained in active stages of aml before allosct or at relapse post transplant were associated with clinical outcome. this observation supports a role of antigen presentation by leukemic cells for an allogeneic immune response in aml. disclosure: nothing to declare background: the role of autologous hematopoetic cell transplantation (hct) in the treatment of aml is not clear. trials in the past have shown that autologous hct consolidation lowers the risk of relapse, however the magnitude of this effect is limited . autologous hct is advocated in patients with aml with lower genetic risk in cr .many of these patients will eventually relapse and will undergo reinduction followed by allogeneic hct in cr . methods: the aims of this study is to analyze outcome of allogeneic hct performed in cr comparing patients with prior consolidation by autologous hct vs. patients with chemotherapy consolidation. primary outcome is non relapse mortality (nrm) of allogeneic hct in cr in patients with, or without prior autologous hct in cr . secondary outcomes include leukemia free survival (lfs), relapse rate (ri), graft versus host disease free relapse free survival (grfs), overall survival (os), and treatment related toxicities. results: adult patients reigstered with the alwp of the ebmt with de novo aml were included, receiving a first allogeneic hct in cr , in cr , in - or without (n= ) prior autologous hct. patient and transplant characteristics are shown in the table. patient groups were not entirely comparable, patients with prior autologous hct were younger, had less often a favorable cytogenetic profile, had more commonly donors other than matched siblings and more often received reduced intensity conditioning (ric) as compared to mac conditioning. univariate outcomes are shown in the table with slightly higher nrm risks in patients with prior autologous hct consolidation. in multivariate analysis nrm risks in patients with prior autologous hct were . ( . - . ), p= . after adjustment for patient age, cytogenetic risk category, year of transplant, donor type, conditioning intensity, sex matching, time from diagnosis to relapse and time from relapse to allogeneic hct as compared to patients with chemotherapy consolidation. similarly, risks of events in lfs and grfs were higher with prior autologous hct, . ( . - . ), p= . and . ( . - . ) p= . , respectively, risk of death was also higher . ( . - . ) p= . but this was not statistically significant. conclusions: we may conclude that some of the advantages of potentially higher anti-leukemic activity of high dose chemotherapy and autologous hct when given to patients with aml in cr (as was shown in a randomized trial by vellenga e et al with lower relapse and higher lfs by approximately % but no significant differences in overall survival) may be lost by higher toxicity of allogeneic hct in cr in case of subsequent relapse. background: although relapse is a major cause of mortality in patients receiving allogeneic hematopoietic cell transplantation (hct) for acute leukemia, limited and conflicting data exist on extramedullary relapse (emr). we aimed to describe the incidence, risk factors, outcomes and prognosis in relapsed hct recipients. methods: we retrospectively reviewed charts of consecutive allogeneic hct recipients transplanted in our center with the indication of acute leukemia ( / - / ). we recorded: age, gender, disease, previous extramedullary involvement, phase at transplant, type of transplant, donor, conditioning, graft-versus-host-disease (gvhd), infections, treatment-related mortality and relapse mortality. in patients with extramedullary relapses, additional data on clinical manifestations, imaging, cerebrospinal fluid testing, histopathology and management were additionally documented. incidence of isolated emr (iemr) and bone marrow relapse (bmr) was calculated using cumulative incidence (ci) analysis, with each and treatment-related mortality considered a competing risk. results: among allohct recipients followed for . ( . - . ) years, ( %) patients presented with emr. the majority of emrs involved the central nervous system (cns, %). isolated emr was observed in patients at . ( . - . ) months. -year cumulative incidence (ci) of . % for iemr was associated only with pre-transplant advanced disease phase (p< . ). bmr was observed in patients at ( . - months), with a -year ci of . %. in the multivariate analysis, bmr ci was independently associated with fungal infections (p< . ), pre-transplant disease phase (p< . ) and lines of treatment (p= . ). -year trm of our whole cohort was . %. the majority of iemr and bmr ( % and %, respectively) patients received systemic treatment combined with local radiation for iemr ( %) and donor lymphocyte infusions (dlis, % and % respectively) when feasible. extensive chronic gvhd was recorded in % of iemr and % of bmr patients. outcomes were poor in iemr, with -year overall survival (os) of . %. favorable os in iemr was associated only with sibling donors (p= . ) and not with other factors, such as treatment with dlis or presence of chronic gvhd. similarly poor outcomes ( year os of . %) were observed in bmr. favorable os was independently associated only with the diagnosis of aml (p= . ) and absence of bacterial infections (p= . ). in the whole cohort, both iemr and bmr were independent unfavorable predictors of os (p< . ) along with extensive chronic gvhd (p= . ). conclusions: in a large population with long-term follow-up, incidence of iemr was relatively high, developed at the late post-transplant period and associated only with disease phase at transplant. furthermore, iemr and bmr conferred similarly poor outcomes despite systemic treatment or extensive chronic gvhd. these independent predictors of survival highlight the unmet clinical need of novel approaches either as maintenance or treatment to reduce extramedullary or systemic relapse post allohct for acute leukemia. disclosure: no competing financial interest. impact of t-cell depletion on outcome in patients undergoing allogeneic hematopoietic stem cell transplantation for acute myeloid leukemia background: after a diagnosis of acute myeloid leukemia (aml) the majority of patients (pts) who achieve complete remission (cr) eventually relapse, with only approximately % of pts maintaining cr for years or longer. late relapses (after years in cr) occur rarely ( - %) in pts receiving hsct in cr and late effects are followed up by routine surveillance as well as preventative measures. the purpose of this study was to investigate long-term outcomes in pts with diagnosis of aml undergoing hsct at our institution in cr . methods: a standardized follow-up of hsct-survivors is applied at our center. we analyzed adult pts with aml in cr consecutively transplanted between january and december at our institution. a written consent was given for the use of medical records for research. a landmark analysis was adopted for patients in cr at -y after hsct (ltcr -long-term cr). results: ltcr was achieved after hsct in / patients (male , female ) transplanted in cr . the median follow-up was years and the median age at transplant years (r - ). the selected donor was a family haploidentical relative in cases, an hla identical relative in , a match unrelated donor in and a cordblood in . in this cohort of ltcr, the -year overall survival was % ( % ci - ). cumulative incidence of relapseevaluated in competing risk with transplant related mortality (trm) -and trm -evaluated in competing risk with relapse -were respectively % ( % ci - ) and % for the cr cohort. the event-free-survival (efs) was % ( % ci - ). the causes of death were relapse ( / pts), second cancer ( / pts) and sepsis ( / pts). the -year incidence of dyslipidemia -defined as cholesterol >/= mg/dl, and/or ldl >/= mg/dl, and/or triglycerides >/= mg/dl or need for specific treatment -was %. the -year incidence of osteopenia / osteoporosisdefined as t-score lower than - and greater than - . and t-score lower than . respectively -was %. the -year incidence of second cancer was %: nonmelanoma skin cancer, lung carcinoma, cervical intraepithelial neoplasm, thyroid cancer, gastric cancer and colon cancer. the -year incidence of chronic moderate-severe gvhd was % ( % ci - ), with the latest diagnosis performed on day . of note, / pts are still on active treatment at last follow-up. conclusions: relapse incidence is low for patient that reached ltcr: patients in cr at transplant can obtain excellent os and efs once reached the target of ltcr. a proactive long-term follow-up and strategy of counseling are essential to keep at best quality the survival advantage offered by hsct in patients with aml in cr . disclosure: chiara bonini has research contract with intellia therapeutics. the other authors declare that they have no conflicts of interest. background: relapse, graft-versus-host disease (gvhd) and gvhd-associated mortality are major obstacles to success of transplantation from unrelated (mud) donors in children with acute leukemia (al). negative depletion of αβ t cells and cd + b lymphocytes, conserves the mature donor-derived natural killer cells and γδ t cells in the graft, may improve gvhd control, immune reconstitution and prevent the relapse. we present a retrospect analyses of a cohort of pts with al in cr transplanted from mud with depletion. methods: a total of children with acute leukemia ( aml, all, female, male, median age , y) underwent allo hsct from matched unrelated donor between june and july . all pts were in complete remission (cr = , cr = , cr> = ). all pts, except one, received treosulfan-based conditioning. either melphalan (n= ) or thiophosphamide (n= ) or etoposide (n= ) were added as a second agent. fludarabine was used in all pts. two types of gvhd prophylaxis were used: type (n= ): hatg mg/kg and post-hsct tacro/mtx (n= ) or without prophylaxis (n= ); type (n= ): thymoglobulin(ratg) mg/kg, rituximab mg/ m with either bortezomib on days + , + (n= ) or tacro/ mtx (n= ) . aβ t cell depletion with clinimacs was used in all cases. the median dose of cd + cells was x / kg, aβ t cells - x /kg. median time of follow-up for survivors was , years (range, , - , ) . results: primary engraftment was achieved in % pts., the median time to neutrophil and platelet recovery was and days, respectively. all evaluable pts achieved sustained complete donor chimerism by day + . early ( day) mortality was , % ( pt -bacterial sepsis, pt -adv fulminant hepatitis), -years overall ptrm at years was , % ( %ci: - ). six late trm events were due to: viral infection in pts (cmv= , adv+cmv= ), bacterial sepsis in pts and pts had bacterial and viral infection, all late deaths were associated with cgvhd and prolonged corticosteroid therapy. ci of acute gvhd grades ii-iv was % ( % ci: - ), acute gvhd grades iii-iv , % ( % ci : , - , ) . ci of cgvhd was %( %ci: - ). regimen was more effective in prevention of agvhd ii-iv in comparison with regimen : % ( % ci: , - ) vs , %, respectively, p= , . all events with acute gvhd grades iii-iv had pts with regimen . ratg was also effective in prevention of cgvhd: ci at years after hsct was , % vs. %, respectively, p= , . cumulative incidence of relapse was % ( %ci: - ) without difference between ratg and hatg. event-free survival (efs) (event=death or relapse) at years was % ( %ci: - ), overall survival %( % ci: - ), there were no difference between age and diagnosis. conclusions: we confirm that the depletion of tcrαβ +/cd + t lymphocytes from the graft ensures high engraftment rate. transplant-related mortality is caused by infections, mostly associated with cases of chronic gvhd. gvhd prophylaxis including ratg/rituximab/ bortezomib improves gvhd control in recipients of tcrαβ+/cd +depleted grafts in comparison to hatg/ tacro/mtx apparently without loss of anti-leukemic activity. disclosure results: at baseline, r/r all with emd and lbl were diagnosed in and ino patients and and sc patients. median (range) age of the ino and sc patients was . ( - ) and . ( - ) years, with / ( . %) and / ( . %) males, respectively. the rate of cr/cri was significantly higher in the ino group ( / [ . %] , % confidence interval [ci] : . - . ) compared with sc ( / [ . %], % ci: . - . ; p= . ) (table) . allogeneic hematopoietic stem cell transplantation was carried out in / ( . %) ino and / ( . %) sc patients prior to any post-study induction therapy. the pfs hazard ratio [hr] was . ( . % ci: . - . ; p= . ), with median pfs of . ( % ci: . - . ) months among ino and . ( % ci: . - . ) months in sc patients. the os hr was . ( . % ci: . - . ; p= . ), with median os of . ( % ci: . - . ) months in ino versus . ( % ci: . - . ) months in sc patients (figure) . all patients had adverse events (aes). serious aes occurred in / ( . %) ino and / ( . %) sc patients; ( . %) ino and sc patients had grade ae. one ( / , . %) patient in the ino group died from veno occlusive disease. conclusions: among r/r all patients with emd and lbl, improvement in remission rates, transplant rates, and progression free survival was shown in the ino group versus the sc group. although patient numbers were small and limited the ability for a robust comparison, these results support the use of ino in patients in this difficult to treat population with r/r all and emd or lbl. background: bcr-abl-targeted tyrosine kinase inhibitors (tki) revolutionized the outcome of patients inflicted with ph+ b-all. moreover, addition of tki may be relevant strategy for ph-like all patients. methods: we hypothesized that overcoming the bm microenvironment-mediated protection of all cells from tki-mediated apoptosis may further enhance the responsiveness to tki therapy. results: in vitro treatment of bcr-abl-positive all cell lines nalm and nalm ) with dasatinib resulted in significant dose-dependent cell growth inhibition, with ic of - nm (p< . ). furthermore, dasatinib exhibited significant growth suppression of bcr-abl -negative all cells (nalm and reh), with ic of nm and nm, respectively. however, when cocultured with bone marrow stromal cells (bmscs), dasatinib-mediated effect was abrogated in both ph-and ph+ all cells. furthermore, dasatinib treatment promoted significant upregulation of chemokine receptor cxcr , on both mrna and cell surface levels. elevated cxcr expression was accompanied by increased responsiveness of all cells to cxcl stimulation, resulting in strong and sustained phosphorylation of erk / and akt and increased adhesion capacity to bmscs. therefore, dasatinib-induced upregulation of cxcr promotes stroma-mediated survival advantage of all cells upon tki therapy. next, in order to overcome the cxcr -mediated stromal protection, we choose to combine dasatinib with the histone deacetylase inhibitor panobinostat, for its known ability to deplete cxcr in aml cells. single-agent treatment with panobinostat demonstrated significant inhibition of ph-and ph+ all cell growth at low nanomolar concentrations (p< . ). importantly, combination of panobinostat with dasatinib synergized (ci< . ), effectively overcoming the protection provided by bmscs and inducing the apoptosis of ph-and ph+ all cells, as demonstrated by phosphatidylserine externalization, mitochondrial depolarization and dna fragmentation. furthermore, combining panobinostat with dasatinib significantly reduced cxcr surface levels in ph-and ph+ all cells. accordingly, cxcl mediated responses, including erk / and akt activation and adhesion to bmscs were significantly reduced upon combined panobinostat/dasatinib treatment. these data indicate that panobinostat effectively suppresses both basal and dasatinib-induced cxcr expression and function in all cells overcoming stroma-mediated resistance to dasatinib. to determine the molecular mechanism, we performed gene and protein expression analysis. panobinostat, alone or in combination with dasatinib, significantly down-regulated the protein levels of calcineurin, a serine-threonine protein phosphatase previously implicated in t-all and b-all pathogenesis, as well as of nfatc , a critical effector of the calcineurin signaling cascade, and nfatc -regulated target genes. it was previously found that calcineurin signaling positively regulates cxcr expression in t lymphocytes. additionally, cyclosporin a (csa) decreased both basal and dasatinib-induced cxcr surface levels in all cells, overcoming the protection of the bmscs which result in potentiation of the cytotoxic effect of dasatininb and panobinostat. combining csa with panobinostat resulted in deeper suppression of nfatc -regulated target genes. we thus link the effect of panobinostat with calcineurin-dependent downregulation of cxcr , blocking the ability of the leukemic cells to respond to cxcl mediated stromal support. conclusions: taken together, our results identify calcineurin signaling pathway as a novel target of panobinostat in all cells and indicate that hdac inhibition with panobinostat may be effective strategy for facilitating the anti-leukemic activity of tki therapy. disclosure: nothing to disclose background: the treatment of relapsed/refractory acute lymphoblastic leukemia (rr-all) remains a clinical challenge with a generally dismal prognosis. allo-sct using a sequential conditioning ("flamsa"-like regimen) has shown promising results in relapsed/refractory aml, but little is known about the efficacy of this procedure in rr-all. methods: we identified adult patients ( % females; median age: y; range, - ) with all in primary refractory phase ( %) or in relapse ( %), allografted between and from a matched sibling ( %), matched unrelated ( %) or haploidentical donor ( %) at ebmt participating centers. almost half ( %) of the patients had t-all and % had a positive philadelphia chromosome. six patients ( %) underwent a previous autotransplant. karnofsky score was above in % of patients. conditioning was myeloablative (mac) with high dose tbi in % of patients, reduced intensity (ric) including low dose tbi in %, or with chemotherapy alone in %. in vivo t cell depletion was performed in cases ( %). most patients ( %) and about half of the donors ( %) were cmv positive. % of patients were males who received a graft from a female donor. the median follow-up was (range, - ) months. results: overall, patients ( %) failed to engraft, ( %) died within days after allo-sct without relapse, and ( %) could achieve complete remission. at day , the cumulative incidences of grade ii-iv and grade iii-iv acute gvhd were % and %, respectively. the year cumulative incidences of chronic and extensive chronic gvhd were % and %, respectively. the -year relapse incidence (ri) and non-relapse mortality (nrm) were % and %, respectively. the -year leukemia free survival (lfs), overall survival (os) and gvhd relapsefree survival (grfs) were %, % and %, respectively. in a multivariable cox analysis, karnofosky score below negatively affected ri, lfs, os and grfs. also, conditioning with chemotherapy alone, compared to tbibased conditioning, negatively affected relapse rates (hr= . ; p= . ), lfs (hr= . ; p= . ) and os (hr= . ; p= . ). conclusions: allo-sct using a sequential conditioning regimen is proposed by different teams in rr-all, and could be an option, especially when considering a tbibased regimen. however, the overall -year lfs of % suggests that these patients still face extremely dismal outcomes, highlighting that other therapies (e.g. bite antibodies, inotuzumab, car t cells) need to be combined prior and/or after allo-sct in order to further improve outcome. disclosure: no conflict of interest, no funding received chemotherapy courses, only pts were not treated: pts for the worsening of the general status and the other for invasive fungal infection. results: forty-three pts ( %) were in complete remission (cr) and negative minimal residual disease (mrd) at the time of hsct; pts were in active disease ( %), and ( %) showed a morphological cr with positive mrd. pts ( %) developed chronic graft versus-host disease (cgvhd) as followed: pts ( %) mild, pts ( %) moderate, and only sever grade respectfully. only patient developed cgvhd after dli. the overall leukemia free survival (lfs) time was months, the absence of cgvhd (hazard ratio -hr: , ; p = , ) and the pre-hsct disease status (hr , ; p = , ) were the most important factors on lfs. all pts treated with chemo-based regimens died due to progression or infective complications. patient of aza/dli group is still alive with a extramedullary relapse; pts treated with bl/dli are in cr. os was better for the dli group compared to the chemotherapy group ( vs months respectfully; p < , ). conclusions: dli after allo-hsct has exhibited definite anti-leukemic effects in post-transplant patients. bl and aza were reported to increase dli's graft vs-leukemia (gvl) effect. although cgvhd could be the most important protective factor against the relapse but it remains the main cause of morbidity. maximising the gvl effect without putting the patient at risk of gvhd still represents an unmet need. our data show that the combination of either bl or aza with dli infusion is safe and might represent an improvement in disease control in the early phase of relapse. disclosure: nothing to declare p increased detection of (leukemiaspecific) adaptive and innate immune-reactive cells under treatment of amldiseased rats and one therapy-refractory aml-patient with blastmodulating, clinically approved response modifiers (pg-e ,kit-k) or +pge (kit-m),patent ) convert myeloid blasts into dendritic cells of leukemic origin (dc leu ). after stimulation with dc leu , antileukemic tcells can be generated ex vivo. the compounds are approved for clinical use and are therefore attractive tools for immunotherapy in myeloid leukemia. methods: dc/dcleu-culture from rats'/patients' wholeblood (wb) with kits, mixed lymphocyte culture (mlc) of tcells with kit-treated blood, functional blast-cytotoxicity and leukemia-specificity assays (csa/elispot/degranulation/intracellular cytokine-assays). in addition flowcytometric evaluations of cellular and (leukemia-specific) lymphocyte compositions were performed from rats'/pts' blood in the course of the disease. results: ) aml-diseased rats: each rats were treated with "i", "k" or "m" or were untreated (controls). a significant increase of dcleu could be detected in spleen/pb in kit-(esp. m) treated compared to untreated animals without induction of blasts' proliferation (ki positivity): a significant reduction of blasts was seen with "m" (p= . / . in spleen/pb) and "i", but not "k". successful treatment correlated with an increase of cd l+tcells, most likely representing tmem-cells, (p= . ) and a reduction of cd +treg (p= . ). ) therapy-refractory aml-patients (during the course of decitabine/ld-aractreatment): kit-m was shown to ex vivo generate dcleu, activate immunereactive cells and mediate leukemia-specific/antileukemic response. activated or leukemia-specific lymphocytes were monitored in low proportions in active stages of the disease as well as of two patients during the further course of persisting disease. one of these patients ( yo male), was offered an individual systemic salvage-treatment (kit-m, applied as continuous infusions) for refractory leukemia. after approval from the local ethical commitee,extensive information of the patient about the experimental nature of the treatment and obtaining his written informed consent. clinically the treatment was well tolerated and the patient improved clinically. neutrophils in wbc increased from % to %, thrombocytes reached g/l after days. after weeks of treatment, the patient was discharged in good clinical conditions. days later, progression of aml was seen with high blast counts in pb and bm. the patient developed severe sepsis and died few days later. immune monitoring showed (other than before treatment and in the patients without kit-m-treatment) a continuous increase of proliferating and non-naïve tcells, nk, cikand nkt-, th cells, bmem-cells and dc in pb. the production of ifnƔ producing t-, cik and nkt-cells was demonstrated, suggesting an in vivo production/activation of (potentially leukemia-specific) cells. immune stimulatory effects decreased after discontinuation of therapy. conclusions: treatment of wb as well as leukemically diseased organisms with blast-modulating kits (especially gm-csf and pge ) was well tolerated and induced clinical and immunological improvement (adaptive and innate immune system), whereas low counts of (leukemiaspecific) activated immune-reactive cells were found in non-kit-treated organisms. disclosure: nothing to declare p long-term outcomes after allogeneic hematopoietic stem cell transplantation for acute myeloid leukemia with non-myeloablative and myeloablative conditioning: a single-center cohort study of consecutive patients lars klingen gjaerde, niels smedegaard andersen , lone smidstrup friis , brian thomas kornblit , søren lykke petersen , ida schjødt , henrik sengeløv rigshospitalet, copenhagen, denmark background: since , we have at our institution used a non-myeloablative (nma) conditioning regimen for older (> years) or significantly comorbid younger patients undergoing allogeneic hematopoietic stem cell transplantation (allo-hsct) for acute myeloid leukemia (aml). we aimed to compare the long-term outcomes of nma conditioned patients with myeloablative (ma) conditioned patients. methods: we studied nma and ma conditioned adult (> years) consecutive patients receiving their first allo-hsct for aml from to at rigshospitalet. nma conditioning consisted mainly of gy total body irradiation (tbi) and fludarabine mg/m ( % of cases). ma conditioning consisted mainly of cyclophosphamide mg/ kg and either gy tbi ( % of cases) or busulfan . mg/ kg ( % of cases), or fludarabine mg/m and treosulfan mg/m ( % of cases). five percent and % of nma and ma conditioned patients, respectively, received anti-thymocyte globulin. patients were followed until death or end-of-followup on october st , . cumulative incidences with % confidence intervals (ci) of acute graft-versus-host disease (agvhd) grade ii-iv, chronic graft-versus-host disease (cgvhd), relapse and non-relapse mortality (nrm) were calculated and compared between nma and ma conditioned patients using gray's test with death as a competing risk (or relapse when comparing nrm). overall survival (os) was estimated by the kaplan-meier method. results: nma and ma conditioned patients were comparable when regarding sex ( % and % female, respectively) and donor (matched related donor in % and %, respectively), but differed, as expected by indication, with regards to age (median of versus years, respectively) and karnofsky score (< in % and %, respectively). nma conditioned patients had generally a lower aml stage at transplant ( st complete remission in % versus % of ma conditioned patients) and a lower aml cytogenetic risk (adverse risk in % versus % of ma conditioned patients). patients were followed for a total of person-years (median follow-up in surviving patients was . years). agvhd grade ii-iv occurred less frequently in nma conditioned patients ( % [ci: %- %] versus % [ci: %- %] in ma conditioned patients, p < . ), while cgvhd occurred in similar rates ( % [ci: %- %] in nma conditioned patients and % [ci: %- %] in ma conditioned patients, p = . ). there was a trend towards a higher relapse rate in nma conditioned patients ( % [ci: %- %] versus % [ci: %- %] in ma conditioned patients, p = . ), and nma conditioned patients had, however not with statistical significance, lower nrm ( % [ci: %- %] versus % in ma conditioned patients, p = . ). os ( figure) was comparable, with -year os rates of % (ci: %- %) in nma conditioned patients and % (ci: %- %) in ma conditioned patients. conclusions: patients with aml undergoing allo-hsct with nma conditioning at our institution were older and frailer than ma conditioned patients, but their overall survival after transplantation was comparable. this might be explained by a generally lower aml stage and cytogenetic risk at transplant in nma conditioned patients. jedlickova , saskia güller , rosa toenges , juliane steinmann , hans martin , hubert serve , gesine bug background: allogeneic hsct is urgently indicated in patients with aml in first complete hematologic remission (chr) after intensive chemotherapy with increasing or recurrent minimal residual disease (mrd). these patients are at high risk of hematologic relapse (hr) during preparation of their transplant and hsct with active aml was found associated with poor outcome. azacitidine has recently been shown to substantially delay or even prevent hr in > % of patients (relaza trial, platzbecker et al., lancet oncology ) . we here present the outcome of a small cohort of consecutive patients with mrd-positive aml who received low dose cytarabine (ldarac) as bridging therapy prior to hsct. methods: mrd was assessed by quantitative polymerase chain reaction (qpcr) using mutated npm (n= ), runx -runx t (n= ), cbfb-myh (n= ) or kmt a-ptd (n= ). mrd negativity was defined as ratio of oncogene to control gene (abl ) ≤ , % while increased or recurrent mrd required a ratio > % (shayegi et al., blood ) . primary endpoint of our retrospective analysis was progression to hr (≥ % bone marrow blasts or extramedullary disease); secondary endpoints were achievement of molecular remission prior to hsct, neutropenia g according to ctcae, thrombocytopenia g , anemia ≥g , admission to hospital, os and rfs. os and rfs were calculated from the first dose of ldarac. ldarac was self-administered subcutaneously by the patients at home at a flat dose of mg bid over days and repeated after weeks if necessary. results: between / and / , nine patients (median age , range, - years) with low (n= ), intermediate (n= ) or high-risk cytogenetics (n= ) according to eln criteria were treated in continuous chr for increasing (n= ) or recurrent mrd (n= ) starting at a median of (range, - ) days after the last consolidation therapy, i.e., duration of chr was > months in all pts. patients received one (n= ), two (n= ) or three cycles (n= ) of ldarac prior to hsct. in three patients, neutropenia g occurred and one patient needed platelet transfusion. all patients were managed in the outpatient setting. in eight out of nine patients ( %), hr was successfully prevented and patients ( %) even became mrd negative prior to hsct. one patient (runx -runx t positive aml) progressed to hr after one cycle of ldarac and received salvage therapy with high-dose arac and mitoxantrone (ham) prior to hsct. all patients proceeded to hsct from a matched related (n= ), unrelated (n= ) or haploidentical donor (n= ) and are still alive (median follow-up of days). conditioning regimens included fludarabine (flu)/melphalan (mel)/tbi (n= ), flu/mel (n= ), flu/tbi (n= ), flu/busulfan (bu) (n= ) and thiotepa/bu /flu (n= ). after hsct, only the ldarac-refractory patient relapsed, resulting in a probability of rfs of % at years. conclusions: our data suggest that a bridging therapy with up to three cycles of ldarac prior to hsct is feasible and was associated with favorable outcomes in patients with npm -mutated or core binding factor aml and molecular relapse > months after achieving a first chr. the treatment has low costs, can be administered on an outpatient basis and is very well tolerated. clinical background: allogenic hematopoietic stem cell transplant (hsct) is the only curative treatment for all the patients with aml. high risk disease qualifies for upfront hsct irrespective of the presence of matched sibling donor (msd). in the absence of msd, haploidentical stem cell transplant is easier option with success rates as high as msd in a high volume transplant centre. we present our experience from a single centre. methods: we analyzed retrospective data of aml patients who have undergone hsct at our centre between january- and august- . for msd transplant we used fludarabine + busulfan or fludarabine + melphalan conditioning regimen, in matched unrelated donor transplant (mud) regime used was fludarabine + busulfan + atg. we followed john hopkins's protocol for haploidentical hsct. cyclosporine + methotrexate was used as gvhd prophylaxis in msd and unrelated donor group and cyclophosphamide + tacrolimus + mycophenolate was used for haploidentical post-transplant. day survival, overall survival (os), incidence of gvhd and cmv reactivation was computed. results: a total of aml patients underwent hsct during the study period, the basic and clinical characteristics of the study patients are presented in table . conditioning regime did not have significant impact on os. survival at day was %. the os function and relapse free survival (rfs) function did not significantly differ between msd and haploidentical transplantation ( . % vs . %; p= . ) and ( . % vs . %; p= . ) (graph ). disease status at latest follow up showed that % were in remission and % had relapsed. overall one year survival and five year survival in the entire cohort was % and % respectively. the average cost of msd transplant at our centre is inr , , (€ - ), haploidentical transplant is inr , , (€ - ) and mud transplant is inr , , (€ , + for stem cell procurement). conclusions: our study showed comparable outcomes in msd and haploidentical transplant with respect to day survival, os, and rate of gvhd. in a developing country like india where patients are not covered under state health insurance, the additional cost of procurement of stem cells in a mud transplant would add to the financial burden to the patients. haploidentical transplant is a feasible option in case of non-availability of msd, due to ease of donor availability and strong motivation from the family donor to donate the stem cells. background: allogeneic stem cell transplantation (allo-hsct) is not indicated as consolidation of first complete remission (cr ) in favorable-risk acute myeloid leukemia (aml) bearing mutations in nucleophosmin (npm ) in the absence of flt internal tandem duplication (flt -itd). nevertheless, a substantial proportion of patients eventually proceed to allo-hsct beyond cr or for chemoresistant minimal residual disease (mrd) while in cr , which might compromise transplantation outcomes. the study aimed at examining the characteristics and results of allo-hsct in aml cases with mutated npm and wild-type flt (npm mut/flt wt), with special focus on molecular monitoring of mrd following transplantation. methods: from / until / , patients (women/men, / ) underwent allo-hsct for npm mut/ flt wt aml. at transplant, median age of patients was . years (range, - ) , and disease phase was cr (n= ), cr (n= ), or primary refractory (n= ). among the patients who were transplanted in cr and had available molecular mrd assessments, had detectable mutant npm transcripts by real-time quantitative pcr (rq-rcr). also, patients fulfilled criteria of molecular relapse (increasing levels of npm -mutated transcripts in two successive bone marrow samples), with mutant npm load of - , transcripts/ , abl transcripts). the conditioning regimen was myeloablative in the majority of cases (n= ) or reduced-intensity (n= ). the type of donor varied, namely hla-identical sibling (n= ), matched unrelated (n= ), haploidentical relative (n= ), or double umbilical cord blood (n= ). results: engraftment was achieved in all cases, with a median time to absolute neutrophil count > /ul of days (range, - ) . among the patients with posttransplant monitoring of mrd by rq-pcr, exhibited a stable molecular remission whereas a rising level of npm mutated transcripts was observed in cases due to either hematologic (n= ) or molecular (n= ) relapse of disease. the cumulative incidences (cin) of hematologic relapse and non-relapse mortality (nrm) were . % and % at months, respectively. no events of relapse or nrm were encountered beyond months from allo-hsct. out of patients with hematologic relapse post transplant, died of disease whereas one achieved a stable complete remission after withdrawal of immunosuppression. at a median follow-up time of months (range, - ), / patients continue to be alive in cr. the estimated disease-free background: cpx- (vyxeos®) is an advanced liposomal encapsulation of cytarabine/daunorubicin at a synergistic : molar ratio. cpx- is approved by the us fda and ema for the treatment of adults with newly diagnosed, therapy-related aml or aml with myelodysplasia-related changes. methods: safety data were pooled from studies of cpx- in adults aged - years with newly diagnosed or relapsed/refractory aml. cpx- induction consisted of units/m (cytarabine mg/m + daunorubicin mg/m ) on days , , and (second induction: days and ) . cpx- consolidation consisted of or units/m (varying by study) on days and . cpx- was evaluated against standard-of-care controls. results: baseline characteristics were generally balanced between cpx- (n= ) and controls (n= ); the majority of patients were aged ≥ years ( %; %) and had secondary aml ( %; %). controls included + (n= ) and salvage therapy with mitoxantrone/etoposide/ cytarabine (n= ), idarubicin/cytarabine (n= ), other cytarabine-based chemotherapy (n= ), and mitoxantrone/ etoposide (n= ). the treatment-emergent adverse event (teae) profile of cpx- units/m was comparable to induction controls, but associated with a greater proportion of patients with teaes, grade ≥ teaes, and serious teaes during consolidation (table) . therefore, the cpx- consolidation dose was reduced to units/m in latter studies; this dose demonstrated an improved teae profile similar to consolidation controls. the most frequent system organ class was gastrointestinal disorders for both cpx- and controls; a lower incidence was reported for cpx- ( %) versus controls ( %), with this difference driven by the lower incidence of diarrhea for cpx- ( %) versus controls ( %). the most frequently reported grade ≥ teaes were febrile neutropenia (cpx- : %; controls: %), pneumonia ( %; %), hypoxia ( %; %), and bacteremia ( %; %). early mortality rates, both overall and by treatment period, appeared lower with cpx- versus controls at day and day ( table) ; the majority of early deaths were attributable to teaes. conclusions: across the studies comprising the cpx- clinical development program, cpx- demonstrated a safety profile comparable to conventional chemotherapy in adults with newly diagnosed or relapsed/refractory aml. background: haploidentical hematopoietic stem cell transplantation (hsct) with post-transplantation cyclophosphamide (pgcy) marked improved clinical outcome. recent studies comparing allogeneic hsct using unrelated donors versus haplo donors in patients with acute leukemia have suggested equivalent outcomes. the depletion of tcells with pgcy was subsequently applied for unrelated hsct setting for patients with unrelated donor. methods: we performed a retrospective study on patients with acute leukemia in order to compare the outcome after hla haploidentical (n= ) and unrelated hsct (n= ) with pgcy. the main characteristics of patients were similar in both groups. baseline disease were: aml ( %) and all ( %) for haplo group and aml ( %) and all ( %) for unrelated group. disease state at time of haplo and unrelated-hsct were following: and patients in cr ( % and %) and and non cr ( % and %). for aml recipients mainly received thiotepa, busulfan and fludarabine and for all recipients received tbi and etoposide conditioning. all patients who received pbsc graft were treated with rabbit antithymocyte globulin (atg) on days - and - . results: at the time of analysis, the os and dfs did not differ between the haplo and unrelated groups ( % vs %, and % vs %). incidence of severe (grade [ ] [ ] acute gvhd was the same in two groups ( % versus %). recipients of haplo-hsct transplant were statistical significance less likely to experience disease relapse ( % vs %) and chronic gvhd ( % vs , %). however, gvhd free relapse free survival (grfs) rate was slightly higher after haplo-hsct ( % vs %). addition, cumulative incidence of trm rate was higher after haplo-hsct ( % vs %).for haplo and unrelated groups who underwent hsct in cr , the os were % and % versus % and % for those in non cr . for aml, the os was same in two groups (haplo % versus unrelated %). however patients with all, the os was higher in haplo group compared with unrelated group ( % versus %). the impact of pretransplant disease state have a more powerfull effect on survival in the haplo-hsct setting (for aml cr % versus non cr % and for all cr % versus non cr %). viral reactivations were significant concern in both groups. conclusions: our retrospective analysis suggests largerly similar os and dfs with haplo versus unrelated transplants with pgcy for acute leukemia. our data indicate that haplo-hsct results in a lower incidence relapse and of chronic gvhd and higher grfs compared with unrelated hsct. in addtion, the pretransplant disease state have the important effect on the outcomes in both groups. allo-pbsc with atg can be used safely and effective as graft source in haplo-hsct with acceptable post-transplant outcomes and replaced bm in this settings. more statistical data for transplant related characteristics will be provided at the presentation.we emphasize that use the same pgcy gvhd prophylaxis for all types of allogeneic transplant. based on our results, we recommend haplo-hsct with pgcy against unrelated transplant for patients with acute leukemia. disclosure: disclosure of conflict of interest: none. excellent efficacy and tolerability of inotuzumab ozogamicin in b-cell all relapsed after allo-hsct background: donor lymphocyte infusion (dli) could be used prophylactically to reduce relapse after allogeneic hematopoietic stem cell transplantation for very high-risk leukemia/lymphoma without effective targeted therapy. to compare the safety and efficacy of prophylactic dli for prevention of relapse after allogeneic peripheral blood stem cell transplantation from haploidentical donors (hid-sct) and matched-sibling donors (msd-sct) in patients with very high-risk acute myeloid leukemia (aml), we performed a retrospective, observational cohort study enrolled in hid-sct and msd-sct recipients. methods: the very high-risk features were defined as: (i) in the non-remission (nr) state prior to transplantation, including primary induction failure, relapse untreated or refractory to reinduction chemotherapy, or untreated aml evolution from mds; (ii) achieving complete remission with ≥ cycles of induction of chemotherapy; (iii) carrying tp , dnmt a, tet or flt -itd gene mutation. the scheduled time of the prophylactic dli was + - days after transplantation for msd-sct recipients and + - days for hid-sct recipients. the g-csfmobilized peripheral blood stem cells were infused to the recipient at a dose of × cd + cells/kg. csa was given at mg/kg b.i.d from day - to day + (hid-sct) or to day + (msd-sct), and then tapered at % per month to be discontinued on day + - (hid-sct) or on day + - (msd-sct) unless graft-versus-host disease (gvhd) developed. if the patients received dli before day + (hid-sct) or day + (msd-sct), csa was given weeks after dli in hid group and weeks in msd group at a though concentration of - ng/ml for dliassociated gvhd prophylaxis, and then tapered and discontinued within weeks unless gvhd developed. if gvhd occurred before the scheduled time of prophylactic dli, it would be delayed for weeks when gvhd was well controlled. results: prophylactic dli was administered at a median of ( - ) days for hid-sct recipients and ( - ) days for msd-sct recipients (p= . ), and both groups displayed similar baseline characteristics except for donor's gender distribution (table ) . grade - acute graft-versushost disease (gvhd) at -day post-dli was higher in hid-sct group than that in msd-sct group ( . % vs. . %, p= . ). grade - acute gvhd ( . % vs. . %), -year chronic gvhd ( . % vs. . %) and severe chronic gvhd ( . % vs. . %) were similar between two groups (p> . ). one-year non-relapse mortality was higher in hid-sct group than that in msd-sct group with marginal significance ( . % vs. . %, p= . ). one-year relapse rate was similar between hid-sct group and msd-sct group ( . % vs. . %, p> . ). estimated -year overall survival (os, . % vs. . %) and relapsefree survival (rfs, . % vs. . %) rates were both similar between hid-sct group and msd-sct group (p> . ). in multivariate analyses, non-remission status prior to transplant, poor-risk gene mutations and donor's age ≥ years predicted a higher risk of relapse after dli. nonremission status prior to transplant predicted inferior os and rfs. patient's age ≥ years also predicted an inferior os. conclusions: prophylactic dli after hid-sct demonstrated similar tolerance and efficacy for reducing relapse compared to that after msd-sct for very high-risk aml. disclosure: the authors declare no conflict of interest. prognostic impact of pre-transplant tim levels on transplant outcome in acute leukemia patients background: t cell immunoglobulin and mucin domaincontaining protein- (tim ), a negative regulator of t cells, is expressed on a variety of tumors including hematological malignancies like acute myeloid leukemia (aml) and some lymphoma types in which it was shown to be associated with an adverse prognosis. the aim of this study is to identify the prognostic impact of pre-transplant tim levels on early and late transplant related complications as well as post-transplant relapse and survival methods: a total of hematopoietic stem cell transplantation (hsct) recipients with an initial diagnosis of acute leukemia [median age: ( - ) years; male/ female: / ] were included in the study. aml was the initial diagnosis in patients ( . %), acute lymphoblastic leukemia (all) in patients ( . %), mixed phenotype acute leukemia in patients ( . %) and blastic plasmacytoid dendritic cell neoplasm in patient ( . %). soluble tim- levels in pre-transplant serum samples were measured with enzyme linked immunosorbent assay (elisa). results: median pre-transplant tim level was . ( . - . ) pg/ml in the whole cohort. pre-transplant tim levels were significantly higher in aml patients when compared to all [ . ( . - . ) vs . ( . - . ); p= . ]. tim levels were significantly lower in patients with abnormal cytogenetics when compared to normal karyotype (p= . ). cytogenetic abnormalities, including mainly a complex karyotype or chromosome abnormalities, were more frequent in patients with low tim levels (p= . ). pre-transplant tim levels were significantly higher in patients who developed post-transplant viral hemorrhagic cystitis (p= . ). a positive correlation was demonstrated between tim levels and acute graft versus host disease (gvhd) grade (p= . ; r= . ). at a median follow-up of . ( . - . ) months, overall survival (os) was found to be better in low-tim group when compared to high-tim group, without statistical significance (% . vs % . ; p> . ) ( figure ). probability of os was relatively better in both aml ( . % vs . %; p> . ) and all patients ( . % vs %; p> . ) representing low pretransplant tim levels in the subgroup analysis conclusions: in this study, elevated levels of pretransplant tim levels in aml patients were compatible with the previous reports which had underlined an increased tim expression on aml stem cells. the possible association of tim expression with cytogenetic features should be confirmed with further studies as there is no adequate data except its relationship with flt -itd mutational status. tim- is also expressed on exhausted t cells in patients with viral infections, including human immunodeficiency virus, hepatitis b and hepatitis c virus. it plays an essential role in the regulation of antiviral and antitumor immune responses which may be an explanation for the increased frequency of hemorrhagic cystitis in patients with higher tim- levels. the adverse prognostic impact of tim on gvhd and os was confirmed without statistical significance which may be related to small sample size. as tim has a wide spectrum of action in the tumor microenvironment including stimulatory and inhibitory activities, further clues are required to define the exact role of this molecule in the clinical course of allogeneic hsct in order to develop targeted therapeutic strategies clinical trial registry: n/a disclosure: nothing to declare p homozygous hla-c is associated with increased risk of relapse after hla-matched transplantation in recipients with acute lymphoid leukemia: a japanese national registry study background: after hematopoietic stem cell transplantation (hsct), the role of natural killer (nk) cells which express killer-cells immunoglobulin-like receptors (kirs) and recognize hla-class ligands is important. kir dl recognizes not hla-c asp (c ), but hla-c lys (c ) and has polymorphism based on the th amino acid of the transmembrane domain. low frequency of c and high frequency of strong kir dl are characteristics observed in japanese. by using large transplant database, we reported that homozygous hla-c (c /c ) recipients displayed lower relapse rates than did c /c recipients after hla-matched hsct for acute myeloid leukemia (aml; hr = . , p = . ) or chronic myeloid leukemia (cml; hr = . , p = . ). this effect seemed to be independent of acute graft-versus-host disease (agvhd) or cytomegalovirus reactivation occurrence (arima n et al bbmt ) . methods: relapse rates of japanese recipients who first underwent hla-matched hsct between and for the treatment of acute lymphoid leukemia (all) were compared between c /c pairs and c /c pairs, using data from japanese data center for hematopoietic cell transplantation and adjusting for transplant characteristics. cord blood transplantation was excluded. multivariable competing risk regression analyses were performed to evaluate relapses and relapse-free survival (rfs) was estimated using kaplan-meier method. results: after recipients who did not achieve remission or experienced graft failure and recipients not-expressing c were excluded, resting recipients aged - years (median, . years) were analyzed. the median follow-up period for survivors was . years. there were recipients expressing c /c and recipients expressing c /c , respectively. after hla-matched hsct, c /c recipients had higher relapse rates than c / c recipients (hr = . , p = . ), resulting in worse rfs among c /c recipients (hr = . , p = . ). the frequent relapse in c /c recipients than in c /c was noticeable among recipients with agvhd (hr = . , p = . ), those without cytomegalovirus reactivation (hr = . , p = . ), and those with ph-negative all (hr = . , p = . ). conclusions: kir dl -positive nk cells may promote graft-versus-leukemia (gvl) in c /c recipients with aml or cml but suppress gvl in c /c recipients with all. one interpretation is that transplant-activated nk cells impair antigen-presenting cells or deprive cytotoxic tlymphocytes of their gvl effects on all cells. this hypothesis may be explained by the fact that agvhd was necessary for the recessive relapse in c /c recipients with all. furthermore, ph-positive all cells sometimes mimic aml cells in terms of their frequent myeloid antigen expression and might be directly targeted by nk cells. it would be necessary to further clarify in vitro the character of nk cell-affecting in the transplant immunity against residual leukemia cells. disclosure: authors have nothing to declare. hematopoietic stem cell transplantation with sequential conditioning for children with relapsed/refractory acute leukemia nao yoshida , kazuki matsumoto , daiki yamashita , yiqing zhu , daichi sajiki , ryo maemura , hirotoshi sakaguchi , asahito hama children's medical center, japanese red cross nagoya first hospital, nagoya, japan background: patients with acute leukemia who fail to achieve complete remission show a dismal prognosis even with allogeneic hematopoietic stem cell transplantation (hsct). this study evaluated whether sequential conditioning approach that is cytoreductive chemotherapy applied shortly prior to the main conditioning followed by hsct can improve prognosis in such high-risk patients. methods: we retrospectively analyzed the outcomes of children (median , range - years old) with primary refractory (n = ) or refractory relapsed (n = ) acute leukemia (aml n = , all n = ) who received hsct in our department between and . the stem cell source was related peripheral blood (pb) in patients, related bone marrow in , unrelated bone marrow in , or unrelated cord blood in . the grafts were hla serologically matched (n = ) or mismatched (n = ) with the recipient. in total, patients received the sequential conditioning approach. as cytoreductive chemotherapy, fludarabine/cytarabine/idarubicin/g-csf (flag-ida) was used in patients, mitoxantrone or daunorubicin/cytarabin in , or other regimens in , and of them were combined with gemtuzumab ozogamicin. without waiting for hematological recovery, the patients promptly underwent hsct; therefore, the median interval between cytoreductive chemotherapy and main conditioning was days. the main conditioning regimens were total body irradiation-based myeloablative (n = ), busulfan-based myeloablative (n = ), or reduced intensity (n = ). results: in children with relapsed/refractory acute leukemia, the -year overall survival (os), leukemia-free survival (lfs), cumulative incidence of relapse (ri), and transplantation-related mortality (trm) were %, %, %, and %, respectively. in multivariate analysis, the use of sequential conditioning was identified as the most favorable factor for lfs (hazard ratio [hr] . ; p = . ), although there were no differences in the outcomes according to the types of cytoreductive chemotherapy or the main conditioning regimen. hla-matched donor (hr . ; p = . ) and pb blasts-negative at the beginning of conditioning (hr . ; p = . ) were also independently associated with better lfs. with sequential conditioning, leukemia burden prior to the hsct was significantly reduced; pb blasts became undetectable at the beginning of conditioning in % patients given the approach, while in % patients without the approach (p = . ). notably, the outcomes in the patients without pb blasts at the beginning of conditioning who received sequential conditioning were promising; the -year os and lfs reached % and % and the -year ri and trm were % and %, respectively. conclusions: our study reveals that hsct with sequential conditioning can be an effective and tolerable treatment option for children with relapsed/refractory acute leukemia. the treatment strategies that focus on the reduction of leukemia burden immediately prior to hsct may contribute to the induction of long-term remissions in patients with high-risk acute leukemia. disclosure: this research was funded by japanese red cross, nagoya st. hospital research grant nfrch - . use of blinatumomab to achieve remission and consolidation with haploidentical transplant with cyclophosphamide post for the treatment of children with refractory acute lymphoblastic leukemia (all) background: most of patients with all in relapse or refractory to conventional treatment have only % possibilities to achieve long term remission. this report refers to the therapeutic efficacy and adverse events from the blinatumomab to achieve molecular remission in patients with pre-b cd + which lead to haploidentical with cyclophosphamide post transplant as a consolidation. methods: a pilot study was conducted in children with refractory all preb-cd +. as a strategy to achieved remission blinatumomab was used at a dose μg/m for continous infusion of hours, increasing the dose to μg/m during days, patients with a mrd of < . log, after cycles received an haploidentical bone marrow transplant as a consolidation, the conditioning regimen was with total body irradiation scheme at cgy/day/ days, cyclophosphamide and etoposide. receiving prophylaxis for gvhd with cyclophosphamide. results: a total of patients were included, seven of them achieved complete remission after cycles of blinatumomab, one with partial remission (table ) , these seven patients, six received an haploidentic transplant achieving graft in of the transplanted patients. one patient had a bone marrow relapse in the first months of the follow-up and patients are free of disease with a follow-up to months (figure ). as a acute complication the patients presented cytokine release syndrome, during the infusion of blinatumumab patients presented tachycardia (table ) and the patients presented agvhd after hsct ( grade i-ii and i grade iv). conclusions: allogeneic bone marrow transplant constitutes a treatment option on those patients that relapse or become refractory to treatment, one of the major problem is basically to identify a hla-identical donor, the alternative is an haploidentical donor. the most important factor to get these results is the disease status before transplant. the use of blinatumomab has proven to be effective in achieving remission in relapse acute linfoblastic leukemia pre-b cd + or refractory to treatment. characteristics nº % male % median age at diagnosis, (range), years . ( - ) status of disease o + relapse % refractory to primary or salvage therapy % complete remission after blinatumomab % partial remission after blinatumomab % active disease % [[p table] . table n° . demographic characteristics of patients undergoing blinatumomab (n= )] disclosure: a. olaya-vargas, r. rivera-luna, y. melchor-vidal, h. salazar-rosales, g. lopez-hernandez, n. ramirez-uribe. we wish to confirm that there are no known conflicts of interest associated with this abstact, the only financial support was provided by mexican associations that helping children wiht cancer in a few patients. sequential high-dose chemotherapy reinduction followed by myeloablative allogeneic transplant for active acute myeloid leukemias methods: at our center, relapsed/refractory aml patients were transplanted during chemo-induced neutropenia after high-dose salvage chemotherapy. median age at transplant was years (range - ). patients suffered from de novo (n= / , %) or secondary aml (n= / , %). genetic risk stratification was reported using stardardized groups proposed by the european leukemia net (eln) in . favorable, intermediate i and ii and adverse risk category at diagnosis was observed in / ( %), / ( %), / patients ( %) respectively. all patients had active disease at the time of sequential therapy and median marrow blast count was % (range - %). patients received a high-dose cytarabine based (mec in / , %) regimen as salvage therapy. donors were haploidentical relatives for / ( %) patients, identical siblings and matched-unrelated for / patients ( %) and / ( %), respectively. a myeloablative conditioning was used to further implement anti-leukemic effects. conditioning, thiotepa-busulfan-fludarabine in % patients, was started at a median of days (range [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] after the last day of chemotherapy. bone marrow and peripheral blood stem cells were used as graft source in / ( %) and / ( %) patients. graft-versus-host disease (gvhd) prophylaxis and supportive care were administered accordingly to each hsct platform. results: all patients engrafted. median day of neutrophil recovery was day + (range - ). median follow-up of survivors was months (range - ). non relapse mortality and relapse incidences (nrm, ri) were % and % at year and % and % at years, respectively. overall cumulative incidences of acute and chronic gvhd were % and % at day + and + . one and year overall survival (os) were % and %, while and year event-free survival (efs) were % and %. significant better os and efs were observed in patients with favorable-intermediate i-ii versus adverse risk score ( - years os % and % vs % and % p= . ; - years efs % and % vs % and % p= . ). adverse risk had a significant impact on os (hr . , p= . ) and efs (hr . , p= . ) by univariate analysis and on ri (sdhr . , p= . ) by fine and gray test. conclusions: though small the patient cohort, our findings suggest that sequential therapy with a myeloablative hsct is feasible in treating relapsed/refractory aml. transplant-related toxicity was low ( %) and relapse was the major treatment-failure. however, even with this approach, patients with adverse cytogenetic features have a very dismal prognosis. for these patients, the use of new drugs before hsct and/or maintenance therapy after transplant is highly encouraged to improve outcomes. disclosure: alessandro busca: honoraria from gilead sciences, merck, pfizer pharmaceuticals and jazz background: in spite of satisfactory results of overall survival (os) after allohsct in st and nd cr aml, relapse free survival (rfs) and graft-versus-host-disease free/relapse free survival (grfs) require further improvement. the detection of mrd is one of the factors which influence on the outcome of allohsct in aml is unclear but identification is important to improve risk-adapted relapse prophylactic treatment after allohsct. aim. to evaluate outcomes of allohcst in st and nd cr pediatric aml depending on the level of mrd status before myeloablative (mac) or reduced intensity conditioning regimens (ric). methods: the data of children with aml in st and nd cr underwent allohsct between and were analyzed. median age at the moment of allohcst was years old ( - ). mrd negative status had ( %) patients, ( %) were mrd positive by flow cytometry. mac based on busulfan ( mg/b.w.) received ( %) patients, on treosulfan - ( %) patients. ric based on melphalan received ( %) patients, based on busulfan ( mg/b.w.) - ( %) patients. patients received prophylaxis of agvhd by atg ( %) or ptcy - ( %) patients plus csa - ( %) or tacrolimus ± sirolimus - ( %) patients that depended on source of transplant (related, unrelated or haplo donor) . results: at the median follow up years in the cohort of mrd positive patients os is % vs % in mrd negative (p> , ). rfs is % vs % accordingly (p= , ). graft-versus-host-disease free/relapse free survival (grfs) in mrd positive patients is % vs % in mrd negative (p> , ). os, rfs, grfs in mrd positive patients after mac is %, %, % vs %, %, % after ric accordingly (p> , ). os, rfs, grfs in mrd negative patients after mac is %, %, % vs %, %, % after ric accordingly (p> , ). os, rfs in mrd negative patients with/without ptcy is %, % vs %, % (p> , ); grfs is % vs % accordingly (p= , ). os, rfs, grfs in mrd positive patients with/without ptcy is %, %, % vs %, %, % (p> , ). conclusions: mrd status does not statistically significant affect on os that can be related to different approaches to the treatment of relapse after allohsct. mrd positive status statistically significant decreases rfs that underline the necessity of posttransplant therapy improvement. ric vs mac in all patients in first and second remission do not show statistically significant impact on os, rfs, grfs. ptcy significantly improves grfs in mrd negative patients. disclosure: none of the authors has anything to disclose. background: with increasing overall-survival (os) of lymphoma patients, higher incidences of therapy-related clonal bone marrow diseases, such as acute myeloid leukemia (aml) and myelodysplastic syndrome (mds) are occuring. generally, the outcome is considered poor. allogeneic hematopoietic stem cell transplantation (allo hsct) often remains the only potentially curative treatment option. nonetheless, there is only little data available concerning this patient group. methods: we retrospectively collected data from patients with therapy-related aml (taml) and mds (tmds) after treatment for hodgkin's lymphoma (hl; n= and n= ) or non-hodgkin's lymphoma (nhl; n= and n= ), who received an allo hsct between and . median follow-up of surviving patients was . years (range . months- . years). background: the prognosis of relapsed/refractory acute leukemia (r/r al) is poor and the treatment is challenging. in this setting, allogeneic stem cell transplantation (allo-sct) constitutes the only curative option although the high relapse rate and non-relapse mortality (nrm). the sequential conditioning regimen followed by allo-sct has been used for persistent disease and aims to improve disease control by intensified chemotherapy, thus conceding more time for the presumed graft-versus-leukemia effect to occur. methods: the clinical outcome of r/r al with the sequential conditioning regimen combining a chemotherapy rescue followed by ric allo-sct in our center is described. patients who underwent a sequential allo-sct from to are included. the primary endpoint was progression free survival (pfs) and overall survival (os) that were estimated by the kaplan-meier method. secondary endpoints were non-relapse mortality (nrm). background: recommendations of the european leukemia net (eln) for favorable-risk genetics (frg) acute myeloid leukemia (aml) favor consolidation over transplantation, although reviews suggest advantage of autologous stem cell transplant (asct) in event free survival. our objective was to compare the progression free survival (pfs) and overall survival (os) of normal karyotype npm mutated without flt itd or allelic ratio < . (npm +) aml patients treated with consolidation chemotherapy alone (cc), asct or allogeneic stem cell transplant (allosct). methods: retrospective review of npm + frg-aml patients, treated in one institution ( to ) with the following induction regimens: cytarabine (ara-c) and vp- with daunorubicin (ade) or mitoxantrone (mice). consolidation regimens were ara-c with daunorubicin (ac-d), idarubicin, vp- and ara-c (mini-ice) or highdose ara-c (hidac). in asct, conditioning regimens were bucy or bvac and in allosct were bucy or flubu. pfs and os were calculated from the start of the last consolidation or stem cell infusion. results: a total of patients were evaluated, with a median age of years (y) ( - y), % female, % with ecog performance status (ps) - and % with ageadjusted charlson comorbidity index (aacii) ≥ at diagnosis. patients were treated with cc in % (n= ), asct in % (n= ) and allosct in % (n= ) of cases. there were no differences between groups for age, aacii, ps, leucocytes at diagnosis or extra-medullary disease. flt -itd was more frequent in allosct group ( %) than cc ( %) or asct ( %; p= . ). at induction, ade was used in % and mice in % of patients, with a complete remission (cr) rate of %. there were no differences between groups for induction regimen or cr. in cc group, consolidation regimens were cycle ( %) and cycles ac-d ( %) and cycles mini-ice ( %). asct patients received consolidation with - cycles ac-d ( %) and cycle mini-ice ( %), while allosct patients received - cycles ac-d ( %), cycles hidac ( %) and no consolidation in %. [[p image] . figure . pfs at y in cc, asct and allosct groups.] median follow-up was months, pfs at y was % and os at y was %. pfs at y for asct group was superior then cc and allosct groups ( %, % and %, respectively; figure ), although not statistically significant. os at y was statistically similar between groups, although inferior in allosct comparing to cc and asct ( %, % and %, respectively). conclusions: in this historical cohort review, although there was no advantage in os for asct in npm + frg aml, our data suggests that there might be a pfs improvement in asct over cc, which needs to be further addressed in prospective studies. disclosure: nothing to declare background: acute myeloid leukemia is a hematological malignant disease that motivates the persistent struggle in the scientific world to provide effective cure that can establish acceptable survival rates in this group of patients. autologous stem cell transplantation with myeloablative conditioning is still a powerful weapon that can be used against this entity methods: we have evaluated retrospectively patients with aml where autologous stem cell transplantation was performed in the period from till . our group consisted of patients; male patients ( . %), female patients ( . %). median age at diagnosis was years ( - ). the average period from time of diagnosis to autologous sct was . months. results: in the majority of our group, we used myeloablative conditioning regimen with busulphan-cyclophosphamide, patients ( . %), in patients ( . %) we have added melphalan to bu-cy conditioning, in ( . %) patients we used beam conditioning and in the rest, patients ( . %) we used bam conditioning regimen. as auto graft we used peripheral blood stem cells (pbsc) in patients ( . %), and in patients ( . %) we used bone marrow. the main mobilising regimen for pbsc was g-csf + etoposide and it was performed in patients ( . %), and in the remaining patients ( . %) mobilising of pbsc was performed only with g-csf. the mean number od apheresis procedures done in our group was . , and the mean number of collected mononuclear cells was . x /kg tt. the mean time to engraftment was . days ( - ). the transplant related mortality (trm) was . %. the year overall survival of our patients was . patients. the main reason for death was relapse of the primary disease( %). patients ( %)were treated with salvage chemotherapy regimen (flag-ida) because with the standard induction regimen + there was absence of adequate therapeutic response, or predominantly no complete remission was achieved. all patients were transplanted in complete remission conclusions: autologous stem cell transplantation could be an acceptable therapeutic solution for patients with aml as a consolidation therapy, where neither suitable compatible donor is available nor allogeneic stem cell transplantation could not be performed from various reasons depending on the bone marrow transplant unit disclosure: nothing to declare p prophylaxis dli alone may not prevent relapse of flt -itd positive aml after allogeneic hct background: one of the most potent prognostic factors affecting outcomes in aml is the presence of cytogenetic and molecular markers which can guide the selection of post-remission therapies. recently, favorable outcomes of npm wt /flt -itd neg /non-cebpa dm group after allogeneic hematopoietic cell transplantation (allo-hct) have been reported, that is similar to those of favorable risk by the eln risk classification. however, the role of allo-hct compared to consolidation chemotherapy has not yet been elucidated. methods: the data of patients who were diagnosed with aml and received intensive induction therapy from march to july were included in the current study. to address the time dependence of the allo-hct, the simon and makuch method was used in the graphical representation and the mantel-byar test and andersen and gill methods for identifying risk factors for long-term survival. results: median age of the patients were years (range - ), and patients ( %) were male. npm mutation was detected in patients ( %), and flt -itd were none, low, and high ratio in patients ( %), ( %), and ( %), respectively. the eln risk classification divided the patients into favorable, intermediate, and adverse risk group in patients ( %), ( %), and ( %), respectively. npn and flt -itd both negative group included patients ( %). allo-hct was performed in patients ( %). overall, complete response (cr) after induction therapy achieved in patients ( %), and patients ( %) were primary refractory disease. cr rates did not differ between npm wt /flt -itd negative group (n= / , . %) and other intermediate risk group (n= / , . %; p= . ) . with median follow-up duration of . months (range . - . months), one-year os rate were %, . ± . %, . ± . % in favorable, intermediate, and adverse risk group (p < . ). among intermediate risk group, os rate of npm wt /flt -itd negative group was similar to other intermediate risk (p= . ). allo-hct was performed in patients of npm wt /flt -itd negative group. one-year os rates did not differ between npm wt /flt -itd negative and other for allogenic hematopoietic stem cell transplant (allo-hsct), as a strategy to prolong survival. methods: data from aml pts over years, who underwent ric allo-hsct in our institution between september and september , was retrospectively collected from clinical files to evaluate the overall survival (os) up to november . we calculated the os using kaplan-meyer curves. results: we identified pts, median age y.o. ( - ) and median htc-i score . the median follow-up was months. one patient (pt) had cml blast crisis and was on first major molecular remission. of the remaining aml pts, were in st complete remission (cr), in nd cr and with progressive disease (pd); the other pts could be classified as mds according to who diagnostic criteria and were in cr . donors (d) were: matched unrelated (mud), mismatched unrelated (mmud - / ), matched siblings and haploidentical. thirteen pts were infused with peripheral blood hsc and with bone marrow. conditionings were: flubcnumel in unrelated donor (ud) pts and siblings, flumel in ud pt and sibling, flubu in ud pts and flutbi gy in sibling and in the haploidentical. graft versus host disease (gvhd) prophylaxis was tacrolimus (tac) + mmf in ud pts and sibling, tac + mtx in ud pts and cyclosporine (cya) + mmf in ud pt and siblings. all mmud pts had atg. ptcy was done in the haploidentical setting with tac + mmf. the median time to neutrophil and platelet engraftment for the whole cohort was and days, respectively. one pt with secondary engraftment failure required re-infusion of selected cd + cells. ten pts presented with mild acute skin gvhd. eleven pts had chronic gvhd, classified as severe; required systemic therapy, of those beyond year. the median time on immunosuppressants was days. at years the os was . %. there were deaths: disease-related ( relapses at and months and pd at d+ ), infection complications ( septic shock) and to secondary neoplasia. other relevant complications were hypoxemic pneumonia in pts, urinary sepsis, cmv and ebv reactivation respectively in and pts; pulmonary and renal toxicity either in pts. at end of follow-up, pts were in remission, without negative measurable residual disease (mrd), the other mrd negative pt died of septic shock and severe intestinal gvhd. conclusions: in this small cohort, chronic gvhd and infectious complications were major causes of morbidity but there were no treatment related deaths before d+ . pts maintaining or achieving mrd negativity after transplant had better survival. although with only pts, these results suggest that allo-hsct is feasible as consolidation strategy in selected aml pts over years. [[p image] . overall survival] disclosure: nothing to declare. background: hematopoietic stem cell transplantation (hsct) is the only curative option for fanconi anaemia (fa); an inherited disorder characterized by congenital anomalies, progressive bone marrow failure (bmf) and a predisposition to develop malignancies. methods: we retrospectively analysed the data of consecutive patients that underwent hsct at this centre from till june . the data was analysed for variables affecting the outcome in terms of overall survival (os). results: median age at diagnosis was years ( - years). median age at transplant was . years ( - yrs). all patients at transplant were in aplastic phase. male to female ratio was . : . twenty-four ( . %) patients had congenital anomalies along with bmf while were phenotypically normal. twenty-three ( . %) patients were / hla matched with siblings, with parents and with cousins. eleven ( . %) patients had gender mismatch transplant. three patients had major and had minor abo mismatch. background: paroxysmal nocturnal hemoglobinuria (pnh) is a rare clonal non-neoplastic hematopoietic stem cell disease whose incidence is . - . cases/million of individuals worldwide. disease characteristics and natural history have been mostly analyzed by multicenter, retrospective studies, with the limit of heterogeneous approaches. herein we report the incidence of severe complications and outcome in a real life setting scenario of pnh patients consecutively diagnosed and managed at our pnh referral center between january and june . methods: patients received a homogeneous diagnostic and treatment approach according to the period of observation (availability of diagnostic tests and eculizumab). all patients treated with eculizumab received vaccination with conjugated anti-meningococcus acwyserotypes and, since , conjugated anti-meningococcus b-serotype. in the event of any complication, patients could refer to dedicated hematology emergency rooms (er) hours daily. the occurrence of renal failure and pulmonary hypertension was specifically evaluated. the renal function was studied according to the cockcroft-gault formula and the lung function was prospectively monitored by daytime-on exertion, nocturnal pulsoximetric profiles and complete spirometric tests, with dlco measurement. results: overall, pnh patients, median age years (range - ), were analyzed. at diagnosis, patients had classic pnh, aplastic pnh and an intermediate form. the cumulative incidences (ci) of thrombosis, and clonal hematologic neoplasm were %, and %, respectively. ci of pancytopenia in the patients with classic pnh was %. one patient showed a spontaneous disappearance of the pnh clone. since , eculizumab was administered in patients. after eculizumab treatment % and % of patients reached hemoglobin level > g/dl and > < g/ dl without transfusion, respectively, while % were nonresponsive. during eculizumab treatment no thrombotic event was observed while two severe infectious episodes (respiratory tract and urinary tract infection) were observed in only one of the patients. extravascular hemolysis was demonstrated in % of patients. no patient showed a significant reduction of the renal function.out of patients prospectively monitored for lung function no pathological alteration in any diurnal or nocturnal pulseoximetric test was observed. no patient showed obstructive or restrictive ventilatory deficiency, nor reduced dlco values. -years overall survival (os) was % ( patients who died for non-pnh related reasons were censored at the last follow-up).a better os, even if not statistically significant,was associated to the absence of thrombotic events ( %vs %), and the period of diagnosis ( % in - , % in - , % in - ) . conclusions: our study reports a better os and lower rate of severe complications in pnh compared to previous experiences. although renal failure and lung hypertension have been reported by other groups, we did not observe these complications along a prolonged follow-up. we can assume that the availability of a dedicated er service enabled an early diagnosis and prompt treatment in case of hemoglobinuria crises (reducing the risk or organ damage) or other complications. the use of eculizumab, together with improved supportive approaches, presumably accounts for the trend towards a better survival witnessed over the last decade in the management of pnh patients. disclosure: nothing to declare haploidentical and unrelated allogeneic stem cell transplantation in aplastic anemia:single center experience zafer gulbas , elif birtas atesoglu , meral sengezer , İmran dora , cigdem eren , suat celik , demet cekdemir background: aplastic anemia is a syndrome of bone marrow failure characterized by peripheral pancytopenia and marrow hypoplasia. allogeneic stem cell transplantation from hlamatched sibling is performed in the firstline setting in young aplastic anemia patients and in elderly patients who are refractory to immunosuppressive treatment. but if the patient does not have a hla-matched sibling, allogeneic stem cell transplantation is performed from unrelated and haploidentical donors. in this study, we analyzaed and compared the results of aplastic anemia patients who had undergone allogeneic stem cell transplantation either from matched unrelated or haploidentical donors. methods: we collected and analyzed data of aplastic anemia patients who had undergone allogeneic stem cell transplantation from matched unrelated or haploidentical donor between and . results: there were patients who had received allogeneic stem cell transplantation from unrelated donors and there were patients who had undergone haploidentical transplantation. but in patients who had undergone haploidentical transplantation, engraftment failure had occurred and they were transplanted from different haploidentical donors fort he second time. so a total of unrelated and haploidentical transplants were performed. the median age of patients who had undergone unrelated transplantation was ( - ) and the median age of patients who had undergone unrelated transplantation was ( - ) . the results of the haploidentical and unrelated transplantations are shown in table . the neutrophil and platelet engraftment times were significantly longer in haploidentical transplantations (p= , and p= , , respectively). however, vod, gvhd and day mortality rates were not different in the groups. similarly overall survival (os) of the patients who had undergone haploidentical or unrelated transplantation were not significantly different (p= , ) ( figure ) . conclusions: although the number of patients are low in this study, we can conclude that urelated and haploidentical transplantation in aplastic anemia have comparable toxicity and efficacy. background: autologous stem cells transplantation (ahsct) is an effective treatment for very aggressive autoimmune diseases such as multiple sclerosis (ms). however, toxicity remains the major concern to a wide application of this approach. post transplant viral reactivations may induce severe complications and a rigorous monitoring of peripheral blood viral load for a prompt and effective therapy is required. a higher rate of ebv and cmv reactivation has been observed in patients affected by ms and conditioned with beam plus atg compared with a controlled group of lymphoma patients without atg in the conditioning regimen [ ] . we report here the policy of our center about both monitoring and treatment of such side effect. methods: a series of consecutive patients with ms, transplanted between and is included in this analysis. all patients were mobilized with cyclophosphamide g/sqm + g-csf and conditioned with beam plus rabbit atg (thymoglobulin©, . mg/kg). monitoring of cmv/ebv dna on whole blood by quantitative pcr was performed after the engraftment, weekly for at least one month, then at longer intervals. pre-emptive treatment with valgancyclovir was started in case of cmv viral load ³ x ^ copies/ml. in case of ebv assay between x ^ and x ^ copies/ml further determinations were performed and rituximab-based treatment was started if the viral copies exceeded x copies. patients received treatment in case of symptomatic disease for any value of the pcr of both viruses or ebv titer ³ x ^ copies/ml. results: detectable dna for cmv was observed in / ( , %) patients at a median time from transplant of days (range - ) and / ( %) required pre-emptive treatment. all patients promptly responded to treatment within weeks. ebv viral load was detectable in / patients ( , %) at a median time of days (range - ). one patient out of started the treatment on first determination for high viral load (> x ^ /ml); nine presented an ebv viral load over x ^ copies/ml, three of them were treated thereafter for the persistent increase of the viral load (> /ml). six patients spontaneously recovered the ebv reactivation. previous treatments were not predictive of any higher risk of viral reactivation. no impact on engraftment related to the reactivation was observed. conclusions: this policy shows that, despite a high rate of cmv and ebv reactivation, no grade iii-iv adverse events were observed, suggesting the key role of viral monitoring in these patients and the efficacy of the preemptive treatment. ebv reactivation at low titers should be monitored to identify those cases that could achieve a spontaneous resolution and avoid the induction of further immunosuppression by rituximab. these data confirm that patients diagnosed with ad undergoing autologous hsct need a more intense pattern of care than hematological patients. background: autoimmune diseases are chronic serious conditions that are often refractory to standard therapies. since , autologous haematopoietic stem cell transplantation (hsct) has been a very promising alternative that has shown satisfactory long-term results. the aim of this study is to evaluate immune reconstitution and mortality following hsct in patients with autoimmune disease. methods: a retrospective study was conducted on patients with diagnosis of autoimmune diseases that underwent autologous hsct between july and january at a tertiary referral center in colombia, south america. descriptive statistics were used to analyze patient's demographic and clinic characteristics. results: seven patients were included, with a mean age of years (range - ). five patients were female ( %). the indications for hsct were systemic sclerosis (n= ) , multiple sclerosis (n= ), and myasthenia gravis (n= ). the conditioning regimen administered in patients with systemic sclerosis was cyclophosphamide + human anti-t lymphocyte immunoglobulin, beam (carmustine, etoposide, cytarabine (ara-c), and melphalan) + human anti-t lymphocyte immunoglobulin in patients with multiple sclerosis, and cyclophosphamide + human anti-t lymphocyte immunoglobulin in myasthenia gravis. median time to myeloid engraftment (neutrophils> . × /l) was days post-transplantation, and platelet engraftment, defined as > , platelets/mm untransfused, was days post-hsct. median time of hospitalization was days (range - ); longer in-patient management was due to infectious complications. infectious complications included bacteremia caused by e. coli and pneumocystis pneumonia that resulted in septic shock and acute respiratory failure, respectively. evaluating t-cell immune reconstitution, none of the patients had reached normal cd + cell value after one year of hsct follow-up. four patients ( . %) reached normal cd + cells value at months post-hsct. regarding bcell immune reconstitution, . % of the patients ( / ) had reached both igg and iga normal levels at one-month post-hsct, and four patients ( . %) had achieved normal igm background: multiple sclerosis(ms) is an inflammatory disease caused by autoimmune reactivity of t cells against myelin. there is accumulating evidence of the efficacy of highdose chemotherapy followed by autologous haematopoietic stem cell transplantation(ahsct) in ms patients who failed response to standard immunotherapy, despite a variability in eligibility criteria, conditioning regimens and outcome. methods: we retrospectively reviewed ms patients submitted to ahsct in our centre ( ) ( ) ( ) ( ) ( ) . patient eligibility criteria were active relapsing remitting(rrms) or secondary-progressive ms (spms), with prior failure to treatment with disease-modifying therapies and evidence of disease activity (clinical relapse or new active lesions in magnetic resonance [mr] ). mobilization of cd +cells to peripheral blood was performed with granulocyte colony-stimulating factor(g-csf μg/kg/day) and conditioning regimen according to beam protocol. the severity of ms disability was classified according to the expanded disability-status scale (edss) and ahsct toxicity was evaluated by ctcaev . . results: seven ms patients had undergone ahsct ( female/ male), with a median age at ahsct of . years ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) . median age at ms diagnosis was . years( - ) and median time from diagnosis until ahsct was . years ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) . four patients ( . %) had spms and ( . %) had rrms, with ( . %) having mr active lesions. pre-ahsct relapse rate per year was ( ) ( ) ( ) ( ) . median baseline edss was . ( . - . ). median number of previous dmts was ( ) ( ) ( ) ( ) ( ) ( ) ( ) . all patients had been treated with corticosteroids and copaxone, ( . %) received rituximab, ( . %) natalizumab, ( . %) alemtuzumab, ( . %) fampridine and ( . %) fingolimod. all patients collected enough cd + cells in a single apheresis session. median number of cd + cells infused was . ± . x ^ /kg, for a mean dmso volume of . ± . ml. median inpatient stay during ahsct was days( - ). all patients developed febrile neutropenia, one was admitted to intensive care unit due to sepsis. one patient developed an anaphylactic reaction to transfusion and another a self-limited encephalopathy. two ( . %) patients had grade≥ oral mucositis, and all had gastrointestinal toxicity (grade - ). median time until neutrophils> /μl was ( - ) days, to platelets> , /μl was days( - ), and to engraftment was days ( ) ( ) ( ) . patients were transfused with a median of ( - ) unit for erythrocytes and ( - ) for platelets. there was no treatment-related mortality and no long term side effects have been observed so far. for a median post-ahsct follow-up of . months, no patient developed new lesions in mr, but ( . %) experienced symptoms consistent with a clinical relapse, at a median time of . ( - ) months, effectively rescued with corticosteroids. the absence of evidence of disease activity at -months was . %. although there was no variation concerning edss punctuation, ( . %) patients self-reported significant benefits, especially concerning limb strength and sphincter continence improving. conclusions: our real life results claim a stabilization effect of ms patients with highly active/progressive disease after ahsct, with no significant toxicity. the failure in reporting benefits in edss punctuation is probably due to a small sample size and short follow-up. more studies are needed to establish the best patient selection criteria and define the ideal time to include these patients in transplantation programs, as well as to evaluate its long term outcome. disclosure: nothing to declare. elena poponina , elena butina , anna yovdiy , galina zaytseva , natalia minaeva , igor paramonov background: reactivation of epstein-barr virus (ebv) represents a potentially life-threatening condition in approximately % of patients after allogeneic stem cell transplantation, with no specific treatment available. methods: we have previously developed a manufacturing protocol for the expansion of cytomegalovirus (cmv) and ebv-specific t cells by stimulation of g-csfmobilized stem cell grafts with defined peptide pools (gary, ) . this advanced therapeutic medicinal product is currently investigated in an ongoing phase i/iia trial (eudract number: - - ) . however, the expansion of virus-specific t cells relies on a pre-existing virusspecific memory compartment in the stem cell donor. in virus-seronegative donors, no expansion can be achieved. we therefore aim to identify ebv peptide-specific t cell receptors (tcrs) that can be translated into off-the-shelf cell products for the treatment and prophylaxis of ebv infection and ebv-associated malignancies. leftover cells from five allogeneic stem cell grafts were expanded in vitro in the presence of hla-b * -restricted peptides (hpvgeadyfey from ebna , eplpqgql-tay from bzlf ) associated with latent and lytic ebv infection. after expansion, single ebv-specific t cells were facs sorted using peptide-mhc (pmhc) tetramers, and individual tcr αand β-chain pairs were determined with single cell sequencing (han , penter . to confirm peptide specificity, dominant tcr pairs were transfected into a murine αβreporter t hybridoma cell line with nfat-driven gfp expression (siewert, ) . functional tcrαβ candidates were transduced into human peripheral background: lymphoid and myeloid acute leukemia are the most frequent cause of cancer related death in children. interactions between nkg d receptor, expressed in cytotoxic immune cells, and its ligands (nkg dl), that are upregulated in many types of tumor cells including leukemic blasts, are important for anti-tumor immune surveillance. nevertheless, tumor cells may develop immune scape strategies like ligand shedding, which reduces nkg dl expression and may cause nkg d receptor downregulation. engineering t lymphocytes with nkg d car may overcome immune evasion and become an effective therapeutic strategy. methods: cd ra -t cells were obtained by depletion of non-mobilized apheresis with cd ra magnetic beads using clinimacs. nkg d-car t cells were generated by lentiviral (nkg d- bb-cd z) transduction of cd ra -t cells with moi= . the expression of nkg d ligands was analyzed in peripheral blood or bone marrow samples from a total of leukemia patients (aml= , b-all= and t-all= ), at different status of the disease (diagnosis, remission, relapse/refractory), and in different leukemia cell lines by qpcr and flow cytometry. cytotoxicity of nkg d-car t cells against leukemia cells was evaluated by performing conventional- hours europium-tda assays. soluble nkg dl (snkg dl) concentration was measured in the sera of leukemia patients by elisa. to evaluate the effect of snkg dl on nkg d-car t cells, those were cultured in the presence or absence of different concentrations of snkg dl for days. one week later, cell proliferation and car downregulation were measured by flow cytometry using cell trace violet and nkg d labeling, respectively. the production of ifn-g and tnf-a was measured in the supernatants by elisa. the effect on cytotoxicity was evaluated in a hoursdegranulation assay by co-culturing snkg dl pretreated nkg d-car t cells against k cell line. results: nkg d ligands were expressed in leukemia cell lines and leukemic blasts. nkg dl expression changed with disease status with a trend to decrease at diagnosis and relapse/refractory compared to remission. nkg d-car t cells were cytotoxic against / leukemia cell lines with a percentage of specific lysis over %. myeloid and t-all cell lines were more susceptible to nkg d-car t cells (specific lysis ranging from - %) compared to b-all cell lines ( - %). physiological concentrations of snkg dl caused an increase in nkg d-car expression. however, supra-physiological levels of snkg dl decreased nkg d-car expression up to times and increased cell proliferation up to times. cd + subpopulation was more affected by downregulation, while proliferation had more impact on cd + subset. the effects of snkg dl were dose-dependent and attenuated by il- . conclusions: nkg d-car t cells are cytotoxic against leukemia cells, specially aml and t-all, and thus could be a novel therapeutic approach for non-b leukemia, or those b-all that relapse with undetectable cd after cd -car treatment. nkg d-car expression may be downregulated only by supra-physiological levels of snkg dl, although antitumor activity is not affected. il- softens the negative effects of snkg dl inducing nkg d expression, cell proliferation and cytokines production. the changes observed in nkg dl surface expression at the different stages of the disease could be related to ligands release and immune escape. disclosure: nothing to declare denis-claude roy , , ines adassi , , céline leboeuf , , vibhuti p. dave , hôpital maisonneuve-rosemont research center, montréal, canada, university of montréal, montréal, canada background: for patients with high-risk leukaemia, allogeneic haematopoietic stem cell transplantation is the only curative treatment. the presence of alloreactive t cells in the donor graft, however, leads to a high probability of developing graft-versus-host disease (gvhd) . t-cell depletion minimises the presence of gvhd-causing alloreactive cells, but often results in an increased incidence of infections and disease relapse. photodepletion treatment (pdt) can specifically deplete activated alloreactive t cells while conserving resting t cells. pdt-treated cells have been utilised after t-cell-depleted haploidentical transplant to help reduce infection and relapse. the efficacy and safety of such pdt-treated cells is currently under clinical investigation in a phase iii trial (hatcy, nct ; kiadis pharma). here the reactivity of pdt-treated donor t cells was assessed toward tumour-associated and viral antigenic peptides derived from wilm's tumour protein (wt p), preferentially expressed antigen in melanoma (pramep), and from cytomegalovirus and epstein-barr virus (cmv/ ebvp). methods: healthy donor (hla-a* ) peripheral blood mononuclear cells (pbmcs) were co-cultured with irradiated pbmcs from another mismatched donor ( : ) in a -day mixed lymphocyte reaction. th , a photoactive rhodamine derivative, was added and cells were exposed to visible light to deplete the th -containing activated alloreactive cells. elimination of alloreactive cells post-pdt was assessed using cd and hla-dr as activation markers. an ex vivo expansion protocol was exploited to evaluate the impact of pdt on reactivity to tumour and viral antigenic peptides. post-pdt t cells were co-cultured with irradiated autologous monocyte-derived dendritic cells ( : ) pulsed with wt p, pramep or cmv/ebvp. antigen-specific t cells were re-stimulated on days and with wt p-or pramep-pulsed autologous pbmcs or with cmv/ebvp added directly to the culture. mhctetramer staining was performed on days and ; ifn-γ elispot was conducted on day . [[p image] . functional wt -specific and viralspecific t cells can be expanded post-pdt] results: pdt resulted in a drastic decrease of cd and/ or hla-dr activation marker-expressing cd + and cd + t cells. pdt-treated cells showed a significant increase in background: acute lymphoblastic leukemia (all) is the most common childhood cancer and relapsed or refractory all is still difficult to treat. engineered t cells equipped with a synthetic chimeric antigen receptor (car) targeting cd have demonstrated remarkable efficacy to treat all. however natural killer (nk) cells are known for their target-independent cytotoxic potential without induction of cytokine release syndrome (crs) or graft-versus-hostdisease (gvhd), car-nk cells can overcome the persisting problem in the therapy with car t cells. as the use of viral vector generated car nk cells is limited by theire genotoxicity, cost and regulatory demands, we are developing an innovative protocol using non-viral sleeping beauty (sb) transposition of third party nk cells as a source to produce 'off the shelf' car-engineered cell products. methods: nk cells are isolated from peripheral blood mononuclear cells (pbmcs) using cd selection kits. they are successfully expanded ex vivo with il- cytokine stimulation under feeder-cell free conditions. after few days of expansion nk cells are electroporated using pmaxgfp. transfection efficiency and percent of living cells after electroporation is analyzed by flow cytometry. transposition based nucleofection using an sb x mrna and a minicircles (mc) dna vector is performed at different time points after nk cell isolation. the transient mc-venus longtime expansion and the viability after sb x based nucleofection is measured over two weeks. furthermore, α-retroviral (α-rv) cd -car transduction of nk cells with different viral amounts (moi) is conducted and the cytotoxicity of the engineered cd -car-nk cells against the cd positiv cell line supb is addressed. results: for an α-rv cd -car transduction of maximal x nk cells we could show transduction efficiency of , % for moi . the α-rv cd -car modified nk cells had a high killing activity against cd positiv supb cells (e:t ration : , %) compared to cd negativ k cell lines (e:t ration : , %) and the non-transduced nk cells (e:t ratio : , %). in first experiments with pmaxgfp vector based nucleofection, we could show an increasing efficiency of , % h post electroporation with a only slightly decreas of living cells ( , %) comparing to the non-electroporated nk cell viability. using sb x mrna with mc-venus dna we electrotransfected x nk cells after fews days of cultivation and we reached , % of transfected nk cells h post electroporation and a transient expression of mc-venus positive nk cells up to , % efficiency with an increasing rate of live cells over days after electroporation. conclusions: the sleeping beauty based nucleofection of nk cells is a very promising non-viral method to generate more easy, safer and higher amounts of genetically modified third party nk cells for therapy of all and has also a broad range of clinical applications. disclosure: winfried s. wels is an inventor on a patent describing chimeric antigen receptors with an optimized hinge region. axel schambach is an inventor on a patent describing alpharetroviral sin vectors. michael hudecek and zoltan ivics are inventors on patents related to sleeping beaut gene transfer technology. the remaining authors have nothing to disclose. background: mature immune cells from the stem cell graft are essential for the graft-versus-tumor (gvt) effect to eliminate residual malignant cells after hematopoietic stem cell transplantation (hsct), but donor cells are also involved in complications such as graft-versus-host disease (gvhd). methods: we performed a prospective study of the detailed graft composition in recipients of peripheral blood stem cells (pbsc) or bone marrow (bm) in order to identify correlations to clinical outcomes, table . grafts were characterized with concentrations of t cells and nk cells together with a multi-color flow cytometry panel with staining for tcrαβ, tcrγδ, vδ , vδ , cd , cd , cd , hla-dr, cd , cd ro, cd ra, cd , cd , cd and cd for detailed immune phenotyping. cell contents in stem cell grafts were analyzed both as fractions of cd positive lymphocytes and as absolute concentrations converted to transplanted cells/ kg. fractions were evaluated in patients receiving both bm and pbsc (n= ), while concentrations (cells/kg) were only analyzed in patients transplanted with pbsc (n= ). table] . table ] [[p image] . figure ] results: we found, that patients transplanted with graft nk cell doses above the median of x /kg and fractions of nk cells out of lymphocytes above the median of . % had significantly increased relapse-free-survival compared to patients transplanted with grafts containing nk cell doses below these values, figure ; results stayed significant in multivariate analyses. relapse incidence was significantly lower in uni-and multivariate analyses in patients receiving grafts with high nk cell fractions compared with low fractions, p= . , with -year relapse rates of % versus % in patients transplanted with high versus low fractions of nk cells, p= . . peripheral blood concentrations of nk cells obtained from samples from pbsc donors before g-csf mobilization were significantly correlated to graft concentrations-and fractions of nk cells.. analyses of graft contents of nkt cells showed that the incidence of grade ii-iv acute gvhd were significantly lower in patients background: extracorporeal photopheresis (ecp) is an immunomodulatory treatment that has shown efficacy in steroid refractory acute gvhd, but the mechanism of action is only partially understood. there is no clear relationship between the ecp-treated mononuclear cells (mnc) or lymphocyte numbers and response to ecp. the objective of the study was to analyse the relationship between the infused subpopulation cellularity and response. methods: patients from different centers with a total of ecp procedures were retrospectively analized. ecp procedures were performed from january- to june- . all ecp procedures were performed with the off-line system. the response was defined as responder (complete and partial response) and non-responder. infused cell numbers for lymphocytes, monocytes and mononuclear cells (lym+mon, mnc) were calculated. for analytic purposes, the median number of cells infused per procedure until response (or until the median number of procedures until response for non-responding patients) and the cumulative number of cells infused until response (or until the median in non-responders) were calculated for all the subgroups. same procedures were performed with the number cells infused until day of ecp. finally, the response and survival impact of infusing a number of cells above or below the median and in different tertiles was assessed until the median number of procedures needed to achieve a response. [[p image] . results: the median number of procedures until response was . we observed a trend towards a higher median number of monocytes per procedure and cumulative infused monocytes in responding patients (median number infused . vs . x /kg p= . , cumulative infused median number . vs x /kg p= . ) that was lost in the day of treatment. there was also a trend toward higher median infused mnc until response for responders ( vs x /kg p= . ). we observed no differences in the number of lymphocytes infused, but patients who received a number of lymphocytes per procedure over the first tertile ( x /kg) presented higher response rates ( % vs %, p= . ). none of the other analysed parameters showed a significant impact in overall survival. conclusions: patients with acute gvhd who responded to ecp received higher numbers of monocytes and mnc in the early phase of the treatment (a median of the first processes). also the patients who received higher numbers of lymphocytes in the first procedures achieved a higher response rate. these findings suggest the possibility that higher number of treated and infused cells could influence the response to ecp, but specifically designed prospective studies are need to asses this possibility. disclosure: nothing to declare background: the field of kidney transplantation has made enormous progress over the last decades towards being a standard treatment for patients with end-stage renal disease. however, administration of immunosuppressive drugs is still one of the major limitations of long-term allograft survival. therefore, strategies for induction of donorspecific tolerance are highly desirable. to this aim, a clinical phase i study with donor-derived modulated immune cells (mics) was conducted. methods: donor-derived mics were manufactured under gmp conditions. potency of mics was tested by different in vitro bio-assays. mics were administered to patients with an escalation from . x mics/kg on day - (n= , group a), to . x mics/kg on day - (n= , group b) or on day - (n= , group c) before kidney transplantation accompanied by standard immunosuppressive medication post-transplantation. frequency of adverse events (ae) was assessed from day until day post-transplant. dynamic changes of various lymphocyte subsets in patients after mic therapy were detected by multicolor flow cytometry. donor-specific immunosuppression was assessed by measuring anti-donor antibodies and mixed lymphocyte reaction (mlr) against donor and thirdparty cells. results: in all kidney transplant recipients, we observed a median serum creatinine of . mg/dl at day which remained stable until day (median creatinine of . mg/ dl) without significant proteinuria. none of patients experienced rejection episode. aes were observed while three aes being severe. most importantly, none of them was associated with mics transfusion. besides two infectious complications, no post-transplant positive cross match results against the donor or titers of de novo donorspecific antibodies were recorded. notably, immunosuppressive therapy could be reduced without signs of rejection in group c. after infusion, we observed a dramatic increase of cd + b cells up to a median of cells/μl until day , followed by a reduction to cells/μl on day in group c. notably, regulatory b cells significantly increased from a median of % on day to % on day . in parallel, the plasma il- /tnf-α ratio increased from a median of . before cell therapy to . on day . after mic cell therapy recipient lymphocytes showed no or only minimal reactivity against irradiated donor pbmcs in vitro, while reactivity against rd -party-donor pbmcs was not impaired. moreover, in vitro mics product demonstrated their immunomodulatory potency by inducing tolerogenic dendritic cells (tdcs) characterized by low expression of costimulatory (cd , cd ) and maturation (cd ) as well as high expression of inhibitory marker cd . functionally, tdcs could inhibit not only the release of ifn-γ but also the proliferation of cmv specific cd + t cells. moreover, mic-induced tdcs showed the capacity to inhibit donor-specific allo-reactive cd + and cd + t cell proliferation. conclusions: mic cell therapy modulates the immune system of kidney transplant recipients by increasing the ratio of regulatory b cells and facilitates the reduction of conventional immunosuppressive therapy without allograft injury or rejection episodes. therefore, mic cell therapy represents a promising strategy in transplantation medicine. we currently prepare a phase ii trial with mic cell therapy. disclosure: nothing to declare p genome editing of graft-derived t cells for post- background: immunotherapy using car t cells has shown promising results to fight cancer. however, car-t cell production requires specialized infrastructure and operators, which implies high cost and centralized production. automated production of car-t cells in clinimacs prodigy device allows clinical-grade manufacturing of car t cells. methods: million cd ramemory t cells from healthy donors were cultured in texmacs supplemented with iu/ml il- . at day cells were activated with t cell transact for h. at day one, activated cd ramemory t cells were transduced with nkg d-cd tm- bb-cd z lentiviral vector at moi = . then, nkg d-car t cells were expanded for - days. nkg d-car t cell products were next harvested, counted and analyzed for viability, nkg d-car expression and anti-tumor cytotoxicity. different quality tests including sterility, vector copy number, genetic stability, quantification of viral particles in the supernatant, myc/tert expression and endotoxin detection were performed. spare cells were cryopreserved either in autologous plasma and % dmso, m % albumin and %dmso or hypothermosol. after months, cryopreserved nkg d-car t cell products were analyzed for viability, nkg d-car expression and cytotoxicity. results: nkg d-car memory t cells expanded up to ± million with , ± % nkg d-car expression and ± % viability. harvested car t cells showed ± % of specific lysis against jurkat cells and ± % against mii osteosarcoma cell line. no microbiological contamination was observed in final car t cells products. vector copy number was ≤ in all validations except for one. cgh and karyotype showed no genetic alterations. free viral particles were undetectable in the supernatants. no overexpression of myc/tert was found except for one validation. endotoxins were ≤ . eu/ml. all cryopreserved nkg d-car t cell products kept nkg d-car expression one year after freezing. however, viability and cytotoxicity was best preserved using autologous plasma %dmso. conclusions: automated production of large-scale clinical-grade nkg d-car t cells using clinimacs prodigy is feasible and reproducible, allowing a decentralized protocol to generate car t cells for clinical use. background: immune reconstitution (ir) is essential to control severe infections after hematopoietic stem cell transplantation (hsct). reconstitution of adaptive immunity may take up to years to recover t-lymphocytes (lt). delay in early lt recovery increases the risk of relapse, viral infections and transplant related mortality. adoptive transfer of selected t cell subset with low alloreactivity potential is emerging as a strategy to improve ir. methods: depletion of cd ra+ naive t cells, preserving cd ro+ memory t cells could provide functional lymphocytes to protect against infection and leukemia relapse with low risk of graft versus host disease (gvhd). we present our experience with high-dose donor cd ro + memory t cell as donor lymphocyte infusions (dli) to assess safety and outcome. a total of dli of cd ro+ after hsct was performed in cases of cmv/ebv reactivation ( %), mixed chimerism ( %), persistent lymphopenia ( , %), graft rejection ( , %) , relapse ( %) or to boost ir ( %). dli product was obtained performing a cd ra depletion on donor leukapheresis product using the clinimacs® device. results: twenty-two pediatric patients, median age years (range - ), with malignant (n= ) and nonmalignant diseases ( ), received cd ra+ (n= ), tcr alpha/beta (n= ) depleted grafts from haploidentical and cd ra+ depleted grafts from match unrelated (n= ) and match related (n= ) donors. at a median of days (range - ) after transplantation, patients received a total of dli of cd ro+ cells, median (range - ), containing a median of . x /kg (range . x - x /kg), cd +cd ro+ . x /kg (range . x - . x /kg) and cd ra+ cells x /kg (range - . x /kg). all infusions were well-tolerated and did not develop or worsen gvhd. a total of / episodes of cmv/ebv viral reactivations decreased viral load, / cleared viral load and / showed a clinical improvement. a total of / patients with persistent lymphopenia there was a slightly increase in total lymphocyte count, but not to normal levels. prophylactic dli of cd ro+ to boost ir increased lymphocyte count in of cases. none of the dli administered in cases of mixed chimerism, graft failure or relapse were effective in reverting those situations. conclusions: our preliminary data suggest that infusions of high dose cd ro+ memory t cells are a safe adoptive immunotherapy strategy. efficacy has been observed in patients with lymphopenia and cmv/ebv reactivation, with no positive results in patients with mixed chimerism, graft failure and relapse. however, to determine the real efficacy of this strategy, prospective studies are required. disclosure: nothing to declare. background: increasing clinical trials have confirmed that chimeric antigen receptor t cells (car-t) targeting cd antigen (car-t- ) is a promising effective approach for the treatment of relapsed/refractory(r/r) b-cell lineage malignancies. considering cd is frequently expressed in large part of t( ; ) acute myeloid leukemia (aml) cells, we suppose that car-t- may be used as an approach to rescuing r/r t( ; ) aml patients. methods: both patients received lymphodepletion chemotherapy with decitabine mg/m × d, fludarabine mg/m × d and cyclophosphamide mg/m × d (dac +fc). two days after chemotherapy, autologous/allogeneic cart- cells provided by the unicar-therapy biomedicine technology co.(shanghai, china) at a total dose of - × cells per kilogram(kg) were infused dose escalation within to days. the research protocol was approved by the institutional review boards of the first affiliated hospital of soochow university and both patients gave written informed consent. results: both cases responded well with transient and reversible toxicities. case presented with grade cytokine release syndrome (crs), manifested by intermittent fever and chill from day after car-t- infusion for half months associated with neutropenia. car-t cells expansion were observed in blood without obvious increase of cytokines. after infusion, case achieved and maintained molecular complete remission (cr) for more than months. case presented with grade crs manifested by continuous high fever, hypotension and grade liver disfunction from day after car-t- cell infusion for week. obvious cytokines releasing (peak il- serum concentration . pg/ml, peak crp serum concentration pg/ml) were detected which were associated with car-t- cell expansion in blood and no severe off-tumor effect was observed. after infusion, case achieved hematological cr and cytogenetic cr and got months disease free survival. conclusions: our report implicates that car-t- is a safe and promising approach to managing r/r t( ; )aml with cd expression, and may provide a salvage treatment approach for all aml patients with cd expression and benefit a certain population with aml besides b-linage malignancies. clinical trial registry: na disclosure: nothing to declare. this work was supported by research grants from the national key r&d program of china ( yfc ), national natural science foundation of china ( , , , ) background: chimeric antigen receptor engineered t (car-t) cells have emerged as a powerful cellular therapy to treat malignant disease, which is currently revolutionizing field of cancer immunotherapy. a cryopreservation step postmanufacture is not only a logistical necessity for large scale cell manufacturing processes but also a mandatory request by regulatory authorities. in case relapse after st car-t cell transplantation, a second application, maybe at a higher dose constitutes a therapeutical option. however, data concerning clinical grade car-t cell stability and functionality after months of cryopreservation have not been released by companies so far. to investigate the effect of cryopreservation on car-t cells, we performed this study. methods: different batches of cd car-t cells were manufactured according to gmp requirements at our institution. final car-t products were frozen at concentrations of x cells/ml (high batch) and x cells/ml (low batch) by a controlled freezing process with the biofreeze bv device and stored in liquid nitrogen tanks below - °c until release. quality control tests for sterility, endotoxin and mycoplasma were performed for each batch according to european pharmacopoeia and united states pharmacopoeia guidelines. stability of cd car-t cells in terms of viability, recovery, transduction efficiency and functional capacity were determined by microscopy, multi-parametric flow cytometry as well as chromium- release tests following our sops. results: all the results of quality controls fully met the requirements of the regulatory authorities. stability results were highly robust and reproducible over time for all our gmp car-t batches. duration of cryopreservation (up to days) had no negative influence on cell viability, recovery of viable cd car-t cells and transduction efficiency. however, the cell concentration for cryopreservation has a significant impact on the post-thawing viability (low batches vs. high batches: . ± . vs. . ± . , p < . ) and recovery (low batches vs. high batches: . ± . vs. . ± . , p < . ) of cryopreserved cd car-t cells, but not the transduction efficiency. moreover, we observed four transient side-effects of cryopreservation on the amount of cytokines released by car-t cells, the cytokine release on a per-cell basis, the multifunctionality of car-t cells and the killing capacity. of note, functional capacity of cryopreserved car-t cells after overnight resting was comparable or even enhanced for inf-γ and tnf-α release by cd + and cd + cd car-t cells when compared to fresh car-t cells. the multi-functionality of car-t cells could be preserved. furthermore, the killing capacity of cryopreserved cd car-t cells after overnight resting could reach the level of non-cryopreserved/fresh car-t cells. conclusions: cryopreservation up to days has no harmful effect on transduction efficiency and functionalities of car-t cells. however, the cell number per milliliter freezing medium matters. dose over x cells/ml should be avoided. for the conduction of in vitro bio-assays to determine the function of car-t cells, an overnight resting process could mimic the situation after clinical application and eliminate the transient side-effects of cryopreservation to fully regain the functional potency of car-t cells. disclosure: nothing to declare background: dc and specific t-cells are important mediators of ctl-responses. we could already show that allogeneic donor-or autologous t-cells obtained from amlpatients can be stimulated by dc leu , resulting in a very efficient lysis of naive blasts. methods: chemokine-release (cxcl , - , - , ccl , - , and il- ) was analysed by cytometric bead array in serum of aml/mds-pts as well as in supernatants from different dc-generating-methods and correlated with pts' clinical course, dc-and t-cell-interactions as well as specific t-cell-reactions. the lytic activity of dcleu/blast -stimulated t-cells in mlc against naive blasts was quantified in a cytotoxicity assay. results: minimal differences in median chemokine-levels in pts' serum subdivided in subtypes were seen, but higher release of cxcl , - , - and lower release of ccl and - tendentially correlated with more favourable subtypes (< years of age, < % blasts in pb). in persisting disease, a higher serum-release of ccl and at relapse a significantly higher ccl -release were found compared to first diagnosis -pointing to a change of 'disease activity' on a chemokine level. whereas chemokine-levels in dc-culture supernatants compared to serum were variable, clear correlations with lateron (after stimulating t-cells with dcleu in mlc) improved antileukemic t-cell activity were seen: higher values of all chemokines in dc-culture supernatants always correlated with improved t-cells' antileukemic activity (compared to stimulation with blast-containing mnc as control) -whereas with respect to the corresponding serum values higher release of cxcl , - , and - but lower values of ccl and - correlated with higher probabilities to improve antileukemic activity of dcleu-stimulated (vs. blaststimulated) t-cells. predictive significant cut-off-values could be evaluated separating the groups compared. moreover, correlations with lateron achieved response to immunotherapy and occurrence of gvhd were seen: higher serum values of cxcl , - , - and ccl and lower values of ccl correlated with achieved response to immunotherapy. predictive cut-off-values could be evaluated separating the groups compared in 'responders' and 'non-responders'. higher levels of ccl and - but lower levels of cxcl , - , - correlated with occurrence of gvhd. conclusions: we conclude, that in aml-pts' serum higher values of cxcl , - , - and lower values of ccl and in part of ccl correlate with more favorable subtypes and improved antitumor'-reactive function. since in dcculture supernatants higher values of all chemokines correlated with improved antileukemic t-cell reactivity we conclude a change of functionality of ccl and - from an 'inflammatory' or 'tumor-promoting' to an 'antitumor'reactive function. this knowledge can contribute to develop immune-modifying strategies that promote antileukemic adaptive immune-responses. disclosure: nothing to declare background: allogeneic hematopoietic cell transplantation (allo-hct) is a curative treatment option for patients suffering from hematologic malignancies. infusion of donor lymphocytes (dlis) can induce sustained remission in case of minimal residual disease or relapse through potent graftversus-leukemia (gvl) effects, although graft-versus-host disease (gvhd) represents a common dose-limiting toxicity. as invariant natural killer t (inkt) cells are known to prevent gvhd while promoting beneficial anti-tumor effects, we investigated the role of inkt cells for successful dlis. methods: we analyzed dli samples by flow cytometry. inkt cells were identified by staining with pbs -loaded cd d tetramers. culture-expanded and purified dli-inkts were then tested against tumor cell lines and primary leukemia cells in an ex vivo tumor control model. tumor cell viability after coincubation with dli-inkts was measured by flow cytometry using -aad. results: inkt cells represent . % (range . - . %) of donor lymphocytes and can be expanded fold following a two-week protocol with a preferential expansion of cd + inkt cells. tumor cell lines such as jurkat were efficiently lysed after coincubation with dli-inkts. cd a as a marker of degranulation was significantly upregulated on dli-inkts after stimulation by jurkat. in addition, increased concentrations of tnfα, ifn-γ, sfasl and perforin were measured after coincubation of dli-inkts with jurkat. we observed that tumor cell lysis correlated with the expression of the mhc-i-like molecule cd d. consequently, adding a cd d antibody to the coculture abrogated the dli-inktmediated kill of tumor cells. dli-inkts also efficiently lysed primary leukemia cells such as aml blasts: expression of cd d on these aml blasts significantly correlated with dli-inkt-mediated tumor cell lysis (r = . , p= . ). conclusions: ex vivo expansion of dli-inkts and subsequent dli enrichment is an immunotherapeutic approach that could improve leukemia control and thus, prevent relapse after allo-hct without exacerbating gvhd. disclosure: nothing to declare. generation of antigen-specific cytotoxic t lymphocytes targeting wt using activated b cells sun ok yun , kyung won baek , hee young shin , hyoung jin kang seoul national university, seoul, korea, republic of background: the wilms tumor antigen (wt ) is highly expressed in many malignancies including leukemia and targeting wt as a tumor associated antigen (taa) in cancer immunotherapy is attractive. in this study, we generated wt -specific cytotoxic t lymphocytes to confirm if activated b cells can act as a cancer antigen presenting cell and induce ctls. methods: for the induction of ctls against wt , activated b cells were used as an antigen presenting cells. b cells were isolated from pbmcs of normal healthy donors and activated with α-galactosylceramide (α-galcer) and nucleofected with wt -coding plasmid dna. activated b cells were the cultured with pbmcs for days in vitro and harvested for assay. results: cells expanded about times after days of culture. we examined characteristic of wt -specific ctls by their surface markers. wt -specific ctls had more than % cd + marker, and ratio of cd to cd was . - . . we also examined nkt cell markers to see if nkt cells were activated by il- , a cytokine used in the induction of ctls, and the portion of nkt cells was about %. the ctls showed a decrease in naïve cell (cd l+cd ra +) and an increase in effector memory (cd l+cd ra-) and central memory (cd l-cd ra-) compared with non-stimulated pbmcs. subsequently, the ifn-γ elispot (enzyme-linked immunospot) assay was performed to confirm the response of the induced wt -specific ctls to the wt antigen. when wt -specific ctls encounters a target that does not have a wt antigen, it did not produce ifn-γ, but when it encounters a target cells loaded wt antigen, it responded to secrete ifn-γ. killing assays were also performed to determine the immunogenicity of induced ctls. the induced wt ctls was found to be killing more than % when the e:t ratio was : when the autologous pbmc met the target with wt pepmix. in addition, we found that wt ctls has killing activity when it encounters leukemia cell lines that express wt and matched hla-a* . conclusions: in this study, we can induce antigenspecific ctls that specifically react to wt using activated b cells as antigen-presenting cells. these observations confirmed that b cells activated by α-galcer can act as a taa presenting cell to induce taa specific ctls as viral antigen, such as pp and ie , and consequently wt specific ctls could be induced. moreover, ctls induced activated b cells had ability to recognize and kill the target cells expressing wt correctly. our results demonstrate that these in vitro expanded wt -specific ctls using activated b cells can be a promising candidate for adoptive immunotherapy against cancer. disclosure: nothing to declare judith böhringer , michael schumm , christiane braun , marina schmidt , patrick schlegel , christian seitz , murat aktas , georg rauser , sandra karitzky , peter lang , rupert handgretinger university children's hospital tübingen, tübingen, germany, miltenyi biotec gmbh, bergisch gladbach, germany background: t cells with chimeric antigen receptors (cars) on their surface facilitate to target specific surface expressed antigens. research and clinical trials with cd -car t cells show impressive remission induction rates and increased survival in heavily pretreated patients. therefore, car t cells are introduced as new potent cellular therapeutics in the clinical routine. in order to establish the manufacture of cd -car t cells, validation runs with the fully automated clinimacs prodigy t cell transduction process have been performed using the miltenyi anti-cd -car lentiviral vector. methods: unmobilized leukaphereses from donors ( x healthy, x all) were used for the clinimacs prodigy t cell transduction process. leukocytes undergo a cd + / cd + t cell enrichment via magnetic beads, followed by stimulation with macs ® gmp t cell transact™, transduction with an anti-cd -car lentiviral vector, expansion with il and il , and final formulation to the cellular product. during and after the manufacture, facs analyses were performed as well as cytotoxicity assays after cd -car t cell production. results: total volumes of leukaphereses were between and ml with . - . x total mononuclear cells. after enrichment x cd + / cd + t cells were transduced with anti-cd -car lentiviral vector and were further expanded. cells were harvested on day . the final cell counts of the cellular products were . , . and . x mononuclear cells from two healthy volunteers and the all-patient, respectively. the transduction efficiency of the cd -car t cells was . %, . % and . % among viable cd + cells. the final count of car t cells was therefore . , . and . x cells. the final products exerted excellent cytolytic activity against cd + bcp-all cell line nalm- . importantly, cd -car t cells generated from the all patient demonstrated complete eradication of autologous blasts at . to e:t ratio after hours incubation. conclusions: the clinimacs prodigy t cell transduction process has been shown to run a fully-automated manufacturing process over days without any deviations in a clean room environment on a single device. the user interaction was reduced to activities at only days to set up the system and provide fresh medium and reagents. the transduction process yielded a high number of t cells with a high frequency of cd -car t transduced cells. the results were comparable for both unmobilized leukaphereses from healthy donors and showed expected slightly lower results in the patient. finally, these results demonstrate that the clinimacs prodigy t cell transduction process is well suited to provide the clinical mb-cart . r/r cd + bcm study with appropriate investigational medical products. disclosure background: allogeneic hematopoietic stem cell transplantation (allohct) is an effective strategy in the long term control of several hematologic diseases, however, patients could experience complications, as graft versus host disease (gvhd) and disease relapse. recently, the introduction of post-transplant cyclophosphamide (ptcy) allowed to significantly reduce gvhd, but disease relapse remains an important issue. donor-lymphocyte infusion (dli) is an established adoptive cell therapy for disease relapse after allohsct, but, in order to be efficient and safe, patients have to be off immunosuppression treatments and gvdh-free. here we report our data about efficacy and safety of dli infusion as treatment for disease relapse in patients who received peripheral blood stem cell transplantation (allopbsct) from hla-matched unrelated/related plus ptcy as gvdh prophylaxis in our clinical trial (nct ). methods: we collected data from patients, treated with ptcy ( mg/kg/die, days + + ), mofetil mycophenolate (mmf) and tacrolimus (t) as gvhd prophylaxis after allo-pbsct, who received dli infusions. they were treated between january and october . we report data about overall response rate (orr), disease control rate (dcr), and dli-related mortality and morbidity. diagnosis were as follow: had multiple myeloma, had acute myeloid leukemia, had acute lymphoblastic leukemia and had lymphomas. all patients but one, who had chimerism loss, received dli because of disease relapse. results: median time between transplant and dli was (range - ) months. median number of dli infusions was (range - ). patients ( %) received cyclophosphamide mg/m preparative regimen the day before the cryopreserved dli infusions, while in the other cases dli were associated with lenalidomide, ponatinib and -azacitidine. the overall response rate (orr) was %, while disease control rate (dcr) was achieved in %. the patient who received dli because of loss of chimerism converted it in full donor after infusions. after dli treatment the incidence of acute gvhd grade i-iii was %, while was % for grade ii-iii and patients were started on short course of systemic immunosuppression treatments . none of these patients died because of dli adverse events. estimated -year overall survival was % with a limited follow-up length ( months). conclusions: the infusion of non-manipulated lymphocytes from allogeneic donors is a valuable and safe strategy of treatment for patients relapsing after allopbsct with ptcy. ptcy showed high efficacy in gvhd prevention, allowing early discontinuation of immunosuppression drugs. because of this, we can reach the goal to transform transplant in a platform where we could add early dli infusions as a new strategy for disease control. clinical trial registry: nct disclosure: nothing to declare p extracorporeal photopheresis in the treatment of refractory chronic gvhd: analysis of mononuclear cell infusion gillen oarbeascoa , maria luisa lozano , , , luisa maria guerra , cristina amunarriz , nuria revilla , , , pastora iniesta , , , cynthia acosta fleitas , jose luis arroyo , eva martinez revuelta , andrea galego , dolores hernandez-maraver , mi kwon , , aurora viejo , jose maria garcia gala , concepcion andon saavedra , jose luis diez-martin , , background: extracorporeal photopheresis (ecp) is an immunomodulatory treatment that has shown efficacy in steroid refractory chronic gvhd, but the mechanism of action is only partially understood. in some studies, a correlation has been suggested between treated mononuclear cells (mnc) or lymphocytes and response to ecp. the objective of the study was to analyze the relationship between the infused cellularity and response in chronic gvhd. methods: patients from different centers with a total of ecp procedures were retrospectively analyzed. ecp procedures were performed from january- to june- . all ecp procedures were performed with the off-line system. the response was defined as responder (complete and partial response) and non-responder. infused cell numbers for lymphocytes, monocytes and mononuclear cells (lym+mon, mnc) were calculated. for analytic purposes, the median number of cells infused per procedure until response (or until the median number of procedures until response for non-responding patients) and the cumulative number of cells infused until response (or until the median in non-responders) were calculated for all the subgroups. same procedures were performed with the number cells infused until day of ecp. finally, the response and survival impact of infusing a number of cells over or below the median and in different tertiles (t , t and t ) was assessed until the median number of procedures needed to achieve a response. results: the median number of procedures until response was . we observed no differences in the median number of lymphocytes, monocytes or mncs infused until response or until day between responding and non-responding patients. there were no differences in response if patients received lymphocytes or monocytes above or below the median number. nevertheless, patients that received a total absolute number of mncs above the median ( x cells) showed a trend towards a higher response rate ( % vs %, p= . ). the patients that received a cumulative number of lymphocytes in the first ecp procedures above the median showed improved overall survival (os) ( y os % vs %, p= . ). patients that received a number of monocytes above the median showed a trend towards better survival (p= . ), that was significant when the number of monocytes infused surpassed the first tertile ( y os % for t , % for t , % for t , p= . ). finally, the patients that received a cumulative number of mncs above the first tertile also showed improved survival ( y os % for t , % for t , % for t , p= . ). conclusions: there were no differences in the infused cellularity between responding and non-responding patients with chronic gvhd. at the same time, we found that except for a trend toward better response with higher mncs infused, there was no relationship between lymphocytes and monocytes with the response rate as other previous studies have suggested. however, even if there is no relationship with the response rate, the patients receiving the highest numbers of lymphocytes, monocytes and mncs in the cohort showed an improved survival, suggesting that larger quantities of cells could exhibit a protective effect. nevertheless, prospective studies that address this relationship are needed. disclosure: nothing to declare comparative analysis of the cytotoxic potential of cytokine-induced killer and natural killer cells for neuroblastoma therapy annekathrin heinze , beatrice grebe , eva mudry , jochen früh , bushra rais , claudia cappel , sabine hünecke , eva rettinger , thomas klingebiel , peter bader , evelyn ullrich , university hospital frankfurt, frankfurt, germany, german cancer consortium (dktk) partner site:, frankfurt, germany background: neuroblastoma (nb) is the most common solid extracranial tumor in childhood. despite therapeutic progress, prognosis for high-risk nb is poor and innovative therapies are of medical need. therefore, we investigated the cytotoxic potential of interleukin (il)-activated natural killer (nk) cells compared to activated cytokine-induced killer (cik) cells against different human nb cell lines in vitro. methods: nk cells were isolated from peripheral blood mononuclear cells (pbmcs) using cd enrichment or cd /cd depletion kits. they were successfully expanded ex vivo with different cytokine combinations such as il- , il- , il- and/or il- under feeder-cell free conditions. in contrast, cik cells were generated from pbmcs by ex vivo stimulation with interferon-γ, il- , okt- and il- . a comparative analysis of expansion rate, purity, phenotype and cytotoxic activity against different nb cell lines following different culturing protocols was performed. results: cd enriched nk cells showed a median expansion rate of . -fold after to days in culture with a final frequency up to . % nk cells and a median frequency of . % cd + cd -t cells. in contrast, the starting cell product after cd /cd depletion consisted of a median frequency of . % nk cells that expanded significantly faster with . -fold and also reached up to . % purity without any relevant t cell contamination. cik cells expanded with a median rate of . -fold and contained . % nk, . % t and . % nk-like t cells. interestingly, nk cells, particularly after cd /cd , showed a significantly higher median cytotoxic capacity against nb cells depletion ( . % for cd enrichment, . % for cd /cd depletion) compared to cik cells that induced . % killing of nb cells with e:t ratio : in a hours' co-incubation assay. interestingly, prolonging the ex vivo stimulation after cd /cd depletion to days enhanced the median expansion rate to . -fold with a slightly reduced cytotoxic potential ( . % for days' ex vivo expansion, . % for days' ex vivo expansion, comparison of the same donors). the addition of an il boost prior harvesting increased the expansion rate to median . -fold (compared to . -fold for the same donors) with an improved cytotoxicity of . % (compared to . %) . fortunately, all nk cell products showed a high viability and no relevant t or b cell contamination (median < . %). interestingly, further optimization of the culturing procedure with use of another cell culture medium led to an improved median . -fold (compared to . -fold) nk cell expansion rate in days, also resulting in comparable cytotoxicity of . %. conclusions: nk and cik cell products may offer an innovative immune therapeutic option for patients with high-risk nb after allogenic stem cell transplantation. our study revealed that nk cells have a significantly higher cytotoxic potential to combat nb. interestingly, the use of il- expanded and il- activated nk cells developed from a cd / depleted apheresis product is highly promising as additional immunotherapy in combination with haploidentical stem cell transplantation of children with nb. disclosure: nothing to declare. quantitative determination of donor allo-reactive t-cells in haploidentical donor-recipient pairs by enzymelinked immunospot (elispot) and mixed lymphocyte culture (mlc) assays background: t-cell alloreactivity is responsible not only for graft versus host disease and morbidity, associated with hematopoietic stem cell transplantation (hsct) but also for graft-versus-leukemia (gvl) activity. in this regard, monitoring and quantitation of alloreactive t-cells (allo-t) may potentially provide valuable information for individualized clinical management of transplant recipients. the aim of this study was the optimization of allo-Т detection and comparison of the elispot and mlc assays. methods: allo-t were determined in haploidentical donor-recipient pairs before hsct. donor mononuclear cells (mnc) served as effector cells (ec) . patient cd depleted mnc were used as stimulatory cells (sc).the ratio ec:sc were : and : . the frequency of allo-t in donor peripheral blood was tested in elispot assay and mlc. elispot provides the detection and quantitation of activated t-cells on the basis of cytokine secreted by each cell. the co-incubation time was h for ifn-gamma and h for il- detection. in mcl assay donor mnc were labeled with cfse and allo-t, proliferating in response to stimulation with alloantigens, were determined by flow cytometry on day . results: the median number of ifn-gamma producing allo-t per donor mnc was , ( - ; ec:sc ratio - : ) and , ( - ; ec:sc ratio - : ). the median frequency of allo-t was , % ( , - , ; ec:sc ratio - : ) and , % ( , - , ; ec:sc ratio - : ) among lymphocytes. il- -producing allo-t were less frequent in donor mnc in comparison with ifngamma-producing allo-t. the median number per mnc was , ( , - ; ec:sc ratio - : ) and , ( - ; ec:sc ratio - : ). the median frequency of il- -allo-t was , % ( , - , ; ec:sc ratio - : ) and , % ( - , ; ec:sc ratio - : ) among lymphocytes. the ec:sc ratio : is enough for stimulation of il- producing by mnc in elispot assay, but for optimal stimulation of ifn-gamma producing cells ec:sc ratio : is preferable. this suggests that allo-t are predominantly ifn-gamma producing cells. alloreactive proliferating t-clones were detected in mlc in of donor-recipient pairs on day of cocultivation. median percentage of proliferating t-clones were , % ( , - , ; ec:sc ratio - : ) and , % ( , - , ; ec:sc ratio - : ) among lymphocytes. however, mlc assay only permit a qualitative analysis that confirmed the presence of alloreactive t-clones, giving no information on their frequency within the culture. results of elispot and mcl assay directly correlated. conclusions: allo-t were detected in , % of assayed haploidentical donor-recipient pairs by elispot and only in , % by mlc. this difference in detection is due to the fact that elispot allows to detect single cytokine secreting cell whereas mlc can reveal proliferating аllo-t clones. the analysis of allo-t in haploidentical donor-recipient pairs may provide rationale to manipulate the allo-immune response and to exploit the powerful ability of allo-t to control hematologic malignancies. disclosure: nothing to declare allogeneic mesenchymal stromal cell as rescue therapy in an infant with life-threatening respiratory syndrome due to a filamin a mutation background: cell-based therapy has gained attention in the respiratory system diseases and encouraging results are reported following mesenchymal stromal cells (mscs) administration. due to their capacity to produce and secrete a variety of paracrine factors and bioactive macromolecules, mscs became a key player in lung tissue injuries and function, reducing fibrosis, promoting the normal development of alveoli and pulmonary vessels. for the first time we used the msc infusions as rescue therapy in a pediatric patient with flna gene mutation and life-threatening respiratory syndrome. methods: a child with a new pathogenic variant of the flna gene c. _ del; (p.val alafster ) at the age of months, due to the serious and irreversible chronic respiratory failure and dismal prognosis, was treated with intravenous infusions of allogeneic bone marrow (bm)-mscs at the dose of × mscs/kg body weight. bm-mscs were produced at "cell factory", fondazione irccs policlinico s. matteo, pavia,isolated and expanded ex vivo from healthy donor bm, following a previously reported protocol. premedication with antistaminic drug, min before every infusion to avoid any potential reaction was performed. the evolution of the respiratory condition was detected. peripheral blood were collected before each msc treatment for treg and th monitoring. treg, defined as cd + cd neg cd + cells expressing the forkhead box p (foxp ) transcription factor, and th , defined as cd + cells expressing intracellular il- , evaluation was performed by flow cytometry (facscanto; bd biosciences, san diego, ca) as previously reported, following standard procedures. results: no acute adverse events related to mscs infusion was recorded. during follow-up, patient maintained a good general condition and showed a regular growth. no systemic or respiratory infections occurred. after the second infusion, the child experienced a progressive improvement of his clinical respiratory condition, with a good adaptation to mechanical ventilation, in the absence of episodes of respiratory exacerbations. the baby maintains adequate volumes of exchange with substantial reduction of the inspiratory support. a reduction of trigger sensitivity was also obtained. thorax ct scan showed a recovery of the basal parenchyma bilaterally and the improvement of the anatomical-functional alignment and aerial penetration. after the first msc administration, an enrichment of treg and th percentage in peripheral blood, was observed. while, after the second msc infusion a significant increase in treg/th ratio was noted. conclusions: this report suggest that msc serial infusions are a promising therapy in aiding the respiratory failure, even in a pediatric patient with flna mutation. intravenous administrations of allogeneic mscs are feasible and safe without toxicity. our results suggest that to mitigate lung injury, mscs may act as regulators of treg and th balance. further investigations are upcoming to establish the useful of this therapeutic proposal in interstitial lung diseases in children. disclosure: nothig to declare p feasibility of il- stimulated donor nk cells manufacturing for early infusion in patients with high risk acute myeloid leukemia undergoing haploidentical transplantation background: nk cells provide a potent antitumor effect in the setting of manipulated haploidentical hematopoietic stem cell transplant (haplo-hsct). we propose a novel strategy to enhance the antitumor effect of allogeneic transplant through the infusion of nk cells stimulated with il- exvivo in adult high-risk acute myeloid leukemia (aml) patients undergoing unmanipulated haplo-hsct. the objective of this study was to provide efficiency and productivity data obtained in the manufactured cellular products infused. methods: selection criteria included patients with highrisk aml undergoing unmanipulated haplo-hsct. lymphoapheresis of the haploidentical donor was performed using spectra optia (terumo® bct) on days + and + after transplant. from the obtained product a double immunomagnetic cellular selection with clinimacs system (miltenyi biotec®) was performed in two steps: cd + depletion followed by positive cd + selection. the obtained an enriched cellular product of cd -cd + nk cells was incubated with il- ( ng/ml) between and hours at ºc and % co in gmp conditions. quality and microbiological controls were performed at the end of each manufacturing step. dxh cellular counters (beckman coulter®) and multiparametric flow cytometry were used for lymphocyte subpopulations and viability analysis (navios cytometer; beckman coulter®, conjugated monoclonal antibodies; miltenyi biotec®). the final product was infused intravenously to the patient on days + and + if manufacturing conditions were met (range of . - x nk/kg, purity ≥ %, viability≥ % and < x cd + cells/kg). if not, it was discarded. nk cell activation in the product was measured by the expression of cd and cd . results: between november and april , patients were included in this ongoing trial. two products were manufactured for of the patients, and only one for the first patient, due to transplant complications between first and second infusion. one product did not meet minimum viability criteria and was discarded. in the infused final products mean and sem of nk cell purity, recovery and viability were . %± . , . %± and . %± . , respectively. log cd + depletion ranged between - . and - . . median infused doses of nk cells and cd + cells per kg were . x ( . x - . x ) and ( - ). complete manufacturing data of all procedures are shown in table background: cytokine-induced killer (cik) cells are a promising immunotherapeutic approach to combat relapse following allogeneic hematopoietic stem cell transplantation (hsct) for acute leukemia or myelodysplastic syndrome. to show safety and efficacy, a multicenter clinical study with pediatric and adult patients including up to eight cik cell applications with escalating doses is ongoing. methods: we favor single large scale cik cell generation with the aim to apply fresh cik cells and cryopreserve ready-for-use doses according to the study protocol in contrast to recurrent manufacturing. therefore cryopreserved cik cells were tested against freshly generated cik cells to approve equivalence. furthermore, an alternative medium supplement for cik cell culturing was investigated to avoid supply bottlenecks in ab-serum. results: fresh frozen plasma (ffp), platelet lysate (pl) and ab-serum in cik cell culture showed median expansion rates of -fold, whereas cultivation without medium additive resulted in significantly lower proliferation (p< . ). cik cell composition including t cells, nk like t cells and a minor part of nk cells was not significantly influenced by changing the medium additive. moreover, neither cytotoxicity against thp- cells nor cd expression on nk like t cells were significantly influenced by the different medium additives. for cik cell generation either ficollized peripheral blood (pb) or unstimulated leukapheresis (lp) products were utilized. with regard to repeated manufacturing within the clinical study, also cryopreserved lp and pbsc as starting material came into the focus of interest. comparing cik cell expansion rates, no significant differences for the entire cik cells and the subgroup of t cells were detected between the four starting materials. cryopreservation of cik cells had no significant effect on cik cell composition, cytotoxicity and cd expression on nk like t cells. a small, albeit not significant effect of cryopreservation on viability was detected, which was . % before and . % after freezing and thawing. conclusions: the challenge was an efficient time-, personal-and cost saving production of cik cells within the clinical study. introducing ffp enabled cik cell manufacturing for an increased patient cohort by avoiding supply bottlenecks in ab-serum. furthermore, cryopreservation allows the storage of ready-for-use cik cell doses fulfilling the demands of the clinical study. clinical trial registry: eudract number - - disclosure: nothing to declare. automated generation of cd ra depleted donor lymphocyte infusion (dli) with the clinimacs prodigy® cd ra system , methods: the current clinimacs cd ra system was developed for graft engineering. up to x e magnetically labeled cd ra+ cells from leukapheresis products can be depleted from up to x e white blood cells (wbc). we developed a new clinimacs prodigy® process in order to ease the procedure for routine-use, to reduce the specifications according to reported cell numbers for dli applications, and to enable the use of peripheral blood products with high amounts of red blood cells (rbc). the new system was tested by performance runs. an new fluorescent flow analysis protocol was developed. results: the resulting clinimacs prodigy pb- ra system is an automated procedure with integrated labeling and washing steps. the new application software pb- ra depletion enables to deplete up to . x e cd ra + cells from up to x e total wbc from peripheral blood products. a major difference of this process is the rbc removal option based on an integrated camera for cell pellet detection. the final cell product is provided in physiologic saline. verification runs with peripheral blood products (n= in total, n= with whole blood, n= with leukapheresis products) resulted in a mean depletion of . log (range . - . ) for cd ra + t cells in the cd ra depleted product. viability of the target products was always above %, and mean wbc recovery was %. the mean process time was h min (range h to h min) without including the manual steps, i.e. tubing set installation and downstream analysis of blood products by flow cytometry. this data were in line with preceding evaluation runs (n= ), and results obtained in cooperation with an external beta test site. the performance results were furthermore in line with results obtained on clinimacs plus instrument runs. for quality control of cd ra depleted products we developed a flow cytometric analysis strategy for fast, accurate, and convenient analysis of even rare counts of remaining unwanted cells. it allows to determine naïve t cells at two different levels of subset staining. the minimum requirement for the flow cytometric analysis includes colors to define viable cd +cd ra+ cells. for further evaluation of the naïve t cell subsets additional colors are used to define viable cd +cd ro-cd -cd l+cd + cells. conclusions: the automated clinimacs prodigy pb- ra system process is capable to deplete cd ra+ cells efficiently from peripheral blood products within hours. the new process is a fast, convenient, and regulatory compliant method for the preparation of ready-to-use cd ra-depleted cell products for clinical applications. the submission to an european notified body for ce certification is an important next step. myeloablative conditioning regimen was preferred for patients out of . gvhd prophylaxis regimens are csa +mtx: (% ), csa+mmf: (% ), csa only: (% ). atg was given patients. despite been given gvhd prophylaxis (% , ) patients out of transplanted patients had gvhd features. of patients, had experienced steroid resistant gvhd after transplantation, including (% ) grade and (% ) grade . ecp treatment was started mean days after diagnosis of steroid resistant gvhd and (% ) patients had complete response while (% ) patients had partial and (% ) patients had no response to ecp treatment on day . sixteen out of patients had also received mesenchymal stem cell therapy as salvage therapy. only one patient had experienced hypocalcemic tetany, a complication of ecp procedure. thirteen patients had died and were directly related with steroid resistant gvhd. other conditions like relapse of primary disease or pres syndrome also played role in death. conclusions: extracorporeal photopheresis is a reliable and effective second line treatment modality in steroid resistant gvhd. starting ecp sessions as soon as gvhd symptoms occur increases its effectivity. mesenchymal stem cell administration with ecp for (% ) patients limits our study to reach o conclusion for efficacy of ecp itself. need for hemodialysis catheters, the prolonged sessions while adequate flow is not possible and catheter related infections are the lmitations for feasibility of ecp. disclosure: nothing to declare donor lymphocyte infusion administrations after allogeneic stem cell transplantations in pediatrics: a single center experience background: loss of chimerism is one of the major problems after allogeneic stem cell transplantation(sct). donor-lymphocyte infusions(dli) are used as a treatment after taper or stopping immunosuppression. in this study, dli experience in patients with loss of chimerism after sct due to various benign and malign hematological diseases was presented. methods: between july -august , twenty patients, detected chimerism loss and received dli after sct were evaluated retrospectively. patients received myeloablative or reduced intensity conditioning, atg, cyclosporine a and methotrexate for gvhd prophylaxis. chimerism analyses were performed with short tandem repeat(str) method from peripheral blood. results below % were considered as mixed chimeric and below % were nonchimeric. when patients considered as mixed chimeric, immunosuppression therapy was ceased immediately and treated with dli. donor lymphocyte infusions were performed at two-week intervals with chimerism follow-up. student t, mann whitney u, ki kare tests and kaplan-meier analysis were used. results: between - ages (median ), female, male patients were evaluated. the initial diagnoses were thalassemia major( ), aplastic anemia ( ),all( ), aml ( ) . dli initiation time was . +- . days after sct, total number of dli administrations were . +- . . dose of dli was x - . x /kg (mean . x /kg). nine patients' chimerism out of , fell below % at first month after transplant; patients were nonchimeric, of them were complet chimeric and were mixed chimeric. eleven patients´chimerism were below % between - months after sct, patients were nonchimeric and were mixed chimeric. early mixed chimerism was found relevant with graft rejections (p= . ). patients were followed up for - days. eight patients' chimerism increased after dli infusion and continued to decrease in patients. after dli, acute gvhd has been seen in both group.the group with decreased chimerism after dli, dose was mean x ± x /kg while the group with increased chimerism had dli dose mean x ± x /kg. although the difference was not statistically significant, numerical value revealed significantly different. eventually patients out of were mixed chimeric, patients were complete chimeric and were none. in thalassemic patients, patients with thalassemia-trait donor were mixed chimeric, in patients whose donors were normal, of them were complete chimeric and one of them was nonchimeric.the difference was significant (p= . ). the cd infusion doses revealed mean . ± . x /kg in mixed chimeric patients, . ± . x /kg in complete chimeric patients and . ± . x /kg in the patients with loss of chimerism. cd amount was seen high as numerical value in complete chimerics but no statistical significance was found. overall survival was %, disease-free survival was %. conclusions: we evaluated the efficacy of dl for patients with mixed chimerism in our patient group. we concluded that chimerism loss in patients with early decreased chimerism is similar to those in literature in spite if dli practices. dose and application frequency were greater in patients with increased chimerism. the small number and the heterogeneity of the patients limited our study. in this regard, studies with larger series and homogeneous groups are acquired. disclosure: nothing to declare phase i clinical trial of repeated administrations of bone-marrow derived mesenchymal stem cells in steroid-refractory chronic graft-versus-host disease patients nayoun kim , young-woo jeon , jae-deog jang , keon-il im , nak-gyun chung , young-sun nam , yunejin song , jun-seok lee , seok-goo cho (cghvd) is the most common long-term complication of allogenic hematopoietic stem cell transplantation which is associated with poor quality of life and increased risk of morbidity and mortality. currently, there is no standardized treatment available for patients who do not respond to steroids. as an alternative to immunosuppressive drugs, mesenchymal stem cells (mscs) have been used to treat and prevent steroidrefractory acute gvhd patients. these studies and reports have also provided a basis for using mscs in steroid refractory cgvhd patients. methods: to evaluate the safety and efficacy of repeatedinfusions of mscs, we enrolled ten severe steroid-refractory cgvhds patients. steroid refractory was defined as either no response to steroids lasting at least weeks or progression of disease during treatment or tapering lasting at least weeks. patients were intravenously administered with mscs produced from third-party bone marrow donors at a -week interval for a total of four doses. each dose contained x cells per kg body weight and all four doses consisted of mscs from the same donor and same passage. results: we enrolled ten patients ( female/ male, with a median age of . (range - ). median of cgvhd affected organs was (range - ) including the skin (n= ), eyes (n= ), oral cavity (n= ), lung (n= ), liver (n= ) and joints (n= ). all ten patients received their planned four doses of mscs, administering a total of infusions. median time from initial cgvhd diagnosis to first msc treatment was days (range - ). msc infusions were well tolerated with no immediate or delayed toxicities. after weeks of the first msc infusion, all ten patients showed partial response showing alleviation in clinical symptoms and increased quality of life. organ responses were seen in skin (n= ), eyes (n= ), oral cavity (n= ), liver (n= ), and joint(n= ). however, one patient died of progressive gvhd and one patient relapsed from primary disease. conclusions: repeated infusions of mscs was feasible and safe and may be an effective salvage therapy in patients with steroid-refractory cgvhd. further large-scale clinical studies with long-term follow up is needed in the future to determine the role of mscs in cgvhd. background: the majority of pregnant polish women ( %) have heard of cord blood banking. however, most doctors do not have sufficient knowledge about the possibility of using cord blood in order to respond to their potential concerns. only . % of healthcare professionals were aware that cord blood could be used to treat haematological diseases. in order to make doctors aware of this issue and provide patients with the information they expect, we would like to present data on the use of cord blood stored in our blood bank for haematological and nonhaematological therapies. methods: the table presented below has been created using data from the general database of the polish stem cell bank, warsaw, poland. no data regarding umbilical cord blood data have been excluded. all patients were planned to be assessed on day , day , on discharge, days after transplantation and , , , , and years after transplantation, but in some cases, patients were lost from follow-up due to a persistent lack of reports from transplantation centres. results: in cases, the therapeutic use of cb was transplantation (replacement of patients' own tissue); in cases it was administration (infusion without destruction of patients' own tissue). thirty-three were administered as standard therapy and as experimental therapy. conclusions: the survey study cited above, indicated low awareness of cord blood use among healthcare professionals. on the other hand, one study indicated that the expectations placed in cord blood banking may be unreasonable. as a private cord blood bank, we support recommendations which underline the importance of patient education. in poland, cord blood has been approved as standard therapy in approx. diseases; most of them are rare, but polish law allows the use of cord blood for the siblings, parents and grandparents of a donor. additionally, active and planned clinical trials throughout the world evaluate the therapeutic efficacy of cord blood in such areas as haematology, neurology, cardiology, diabetology, congenital paediatric disorders, ophthalmology, dermatology, gastroenterology, hiv infection, and the quality of life during aging. therefore, further indications may be expected in the future. background: cytokine release syndrome (crs) has been identified as a clinically significant, on target, off tumour side effect of the chimeric antigen receptor (car) t-cell therapies. it is clinically increased in interkeukin and elevations in other cytokines, lactate dehydrogenase (ldh), c-reactive protein (crp), and ferritin. these side effects can include fever, fatigue, headache, encephalopathy, hypertension, tachycardia, coagulopathy, nausea, capillary leak and multi organ dysfunction. crs symptoms can appear as early as one day after infusion and can resolve quickly or last for weeks. it´s severity to be related to the disease burden prior to car t-cell therapy. methods: the bristol oncology and haematology centre will be providing car t-cell therapy to patients in early . on collating from the leading consultant on the ward and fellow nursing team members it was apparent that an effective way at managing patients post car t-cell therapy side effects is to provide an educational and informative poster depicting a flow chart that will aid the practitioner to recognise and effectively treat/manage a patient with crs symptoms. results: none as of yet as this is a prospective tool ready for our first patient in early conclusions: through continuing reading and study days prior to the ward receiving its first car t-cell patient it is increasingly important that the entire multi disciplinary team recognise crs and understand the importance of early detection, careful monitoring and early intervention. background: allogeneic stem cell transplantation (allosct) is the only curative procedure for primary and secondary myelofibrosis (pmf, smf). elderly people are mainly affected, limiting the feasibility of intensive myeloablative chemotherapy regimens. the introduction of reduced-intensity conditioning (ric) made allosct feasible and effective for old patients. nevertheless, the incidence of pmf and smf is not negligible in young patients, theoretically able to tolerate also high-intensity therapy. very few data are available about the efficacy of ric-allosct in the particular setting of young-aged mf patients. methods: this study includes myelofibrosis young patients (age < y) who received allosct between and at the university hospital hamburg/germany. four patients were previously splenectomized. patients mostly fall into intermediate risk groups according to dipss. four patients belonged to the high-risk triple-negative category (jak /calr/mpl-). asxl -mut was tested in patients (pos: ). in % graft source was pbsc, patients received bmsc. only % of patients had a / hla-matched sibling, the others were transplanted from fully-matched ( %) or partially-matched ( %) unrelated donor. all transplants were conditioned according the ebmt protocol with busulfan ( mg/kg po or mg/kg iv), fludarabine ( mg/m ), atlg (grafalon® neovii, germany) administered in days at a dose of mg/kg die for mud, mg/kg die for mrd transplants, followed by cylosporina, and mycophenolate in the first days. results: engraftment rate was %, median neutrophil engraftment time days. platelet engraftment was reached by patients ( %, median days). four patients ( %) developed poor graft function, successfully treated with cd + selected pbsc-boost. after a median follow up of . years, estimated y-pfs and os were % and % respectively. dipss-risk and donor hla-matching resulted the only significant impacting factors on os. neither cytogenetic nor molecular abnormalities were significantly related to os. twenty-five patients ( %) experienced agvhd grade > . c-gvhd was observed in patients ( %), mostly ( %) beginning in the first days after transplantation. cumulative incidence of trm was % at year, with a plateau after the first year ( y trm = %). trm was observed only in patients with maximal grade ( ) of marrow fibrosis. furthermore, trm never occurred in previously splenectomized patients (p= . ), but no significant impact from splenectomy on os was observed (p= . ). after transplant, patients ( %) relapsed: died without any treatment because of infection, received dli ( durable cr), patients ( after dli) underwent a second allosct, with long-term survival in cases. conclusions: ric followed by allogeneic sct is a curative treatment approach for younger patients with myelofibrosis with a low nrm. the most important outcome-determining factor is donor hla-matching. interestingly, marrow high grade fibrosis showed to significantly impact trm. biological markers such as asxl mutation and cytogenetics, largely known as highly predictive for poor prognosis in the disease natural course, did not show any impact on survival, suggesting that patients harboring these abnormalities could get the greatest benefit from allosct. further data collection, and a prospective randomized trial are needed to confirm our conclusion. disclosure: nothing to declare p abstract already published. splenectomy as a risk factor for relapse after allogeneic hematopoietic stem cell transplantation in patients with myelofibrosis -retrospective cohort study background: splenectomy is a common procedure in patients (pts) with myelofibrosis (mf) performed to achieve improvement in blood cell counts and reduce b-symptoms. however, it has also been shown that splenectomy may adversely predispose to leukemic transformation in this setting. aim: to evaluate in a single-center retrospective analysis the long-term impact of pre-or post-transplant splenectomy on transplant outcome regarding overall survival and relapse risk. methods: this retrospective analysis comprises the data of pts ( male and female) with primary (n= ) or secondary (n= ) mf after allo-hsct from hla-matched sibling (n= ) or unrelated (n= ) donors in our center between and . the median age was years (range, to years). a myeloablative conditioning regimen was performed in pts, while pts where treated with a reduced intensity conditioning. peripheral blood stem cells (n= ) or bone marrow (n= ) with a median of . x cd + cells/kg bodyweight (bw) (range, . to ) were transplanted. splenectomy was performed in of pts: pts were splenectomized prior to and pts after allo-hsct. relapse was diagnosed in ( %) of pts. the median duration to relapse after transplantion was months (range, - months). results: the median duration of follow-up of this cohort was months (range, - months), the -year overall survival (os) was %. pts died, including pts who relapsed and pts who died of treatment related causes. the observed probability of relapse was significantly higher in splenectomized pts than in non-splenectomized pts: % versus % (relative risk (rr) . , % ci, . - . , p= . ). at years, the os was % in nonsplenectomised and % in splenectomised pts (p= . ) (fig. ) . the relapse rate in splenectomised pts was independent of pre-( of pts, %) or post-transplant ( of pts, %) treatment (rr . , % ci, p= . ) . conclusions: on the basis of our cohort, we could assert that pre-and post-allo-hsct splenectomy was equally and significantly associated with an increased relapse ratio in patients with mf, which also tends to negatively affect overall survival. [[p image] . figure : the overall survival after allo-hsct in patients with myelofibrosis.] background: b-cell prolymphocytic leukemia (b-pll) is a very rare lymphoproliferative disorder. although allogeneic stem cell transplantation (allosct) could be a curative option, patients often do not qualify for this consolidation treatment due to an aggressive course of disease. in this case study, we report on three patients who failed ibrutinib therapy but achieved complete remission and even mrd negativity after treatment with the chimeric cd -antibody rituximab, enabling them to undergo allosct. methods: clinical data and follow-up data were collected by chart review. results: all three patients (pt# : male, years; pt# : female, years; pt# : female, years) were referred with b-pll harboring highly complex aberrant karyotypes, including p abnormalities in pt# and pt# . a tp mutation could be detected in pt# and pt# . all three patients had symptomatic disease with rapidly increasing hyperleukocytosis and massive splenomegaly. two of them were treatment-naive and one relapsed after chemoimmunotherapy. all patients were put on ibrutinib mg. despite initial response to treatment, two patients developed progressive disease after (pt# ) and months (pt# ) on ibrutinib, whereas pt# remained in partial remission with persisting leukocytosis, precluding consolidating allosct as originally intended. in pt# , ibrutinib was replaced by venetoclax, but without response. in order to control rapid lymphoproliferation, rituximab was added to venetoclax. grade infusion reaction / tumor lysis syndrome (tls) (fever, tachycardia and hypotonia requiring intravenous vasopressors) followed each rituximab administration despite fractionating rituximab to small doses. however, continuation of rituximab ( mg/d over d) led to complete and durable clearance of hyperleukocytosis (from /nl to mrd negativity) despite venetoclax cessation. a similar pattern was observed in pt# , who received rituximab while showing rapidly increasing leukocytosis upon ibrutinib. again, complete clearance of b-pll cells in the peripheral blood (from /nl to mrd negativity) occurred after initial grade tls despite only modest cd expression on tumor cells in this patient. also, pt# achieved profound b-pll cell depletion (from /nl to a mrd rate of . %) upon addition of rituximab to ibrutinib (without tls in this case). subsequently, all three patients were able to undergo allosct after conditioning with fludarabine and total body irradiation with gy. pt# received stem cells from a hla-ident sibling donor, whereas pt# and pt# had unrelated donors (hla-ident and hla-matched respectively). with follow-up times of and months post-transplant, pt# and pt# are currently in ongoing mrd-negative remission. pt# developed an acute graft-versus-host disease (gvhd) of the liver (grade ), nevertheless the clinical course was well controlled by immunosuppression. in pt# a chronic gvhd of the skin occurred. pt# , who achieved mrd negativity after allosct, developed acute and chronic steroidrefractory gvhd of the skin and gastrointestinal tract. nine months post-transplant, gvhd deteriorated and after further complications the patient died of pneumonia months post-transplant. conclusions: supplementary treatment with rituximab can induce deep remissions in patients with ibrutinibresistant, genetically poor-risk b-pll, thereby enabling them to undergo successful consolidation with allosct. a high risk of life-threatening infusion reactions / tls associated with the addition of rituximab has to be taken into account. background: there is little experience on the use of the newer targeted therapies in cll patients relapsed after allogeneic stem cell transplantation (allo-sct). against this background, we evaluate the safety and efficacy of the bcr inhibitors (bcri), ibrutinib and idelalisib, administered after allo-sct for the purpose of treating the cll relapse. methods: data from cll pts who relapsed after sct, and were subsequently treated with ibrutinib (n= ), idelalisib (n= ) or both (n= ),were retrospectively collected in collaboration with the spanish group of cll (gellc) and the spanish group of stem cell transplantation (geth). results: transplant characteristics are summarized in table . eight patients received the bcri as the first salvage treatment after sct relapse, whereas patients had received ≥ prior lines of treatment. at the time of the onset of the bcri, patients had rai stage and patients had a lymph node size ≥ cm. del p was present in patients and del q and complex karyotype in patients, respectively. tp gene mutation was detected in patients (all with del p ). median time from sct to bcri therapy was . months, being shorter in patients treated with ibrutinib (n= , median months) than in those treated with idealisib (n= , median months). median time on ibrutinib and on idelalisib was . months ( . - . ) and months ( . - . ), respectively. the best overall response rate (orr) obtained with ibrutinib was % ( cr, pr, pr+l) whereas it was of % for patients receiving idelalisib ( cr, pr+l). among the patients treated with ibrutinib, ( . %) presented an adverse event (ae), being diarrhea (n= ), asthenia (n= ) and infections (n= ) the most frequent. hypertension was seen in patient and none patient developed atrial fibrillation. five patients stopped ibrutinib treatment, due to toxicity (n= ) or progression (n= ). after ibrutinib discontinuation, patients were newly treated with idelalisib (n= ) or venetoclax (n= ). all patients treated with idelalisib developed at least one ae, being diarrhea (n= ), pneumonitis (n= ) and neutropenia (n= ) the most common. four patients discontinued idelalisib because progression (n= ) or toxicity (n= ). venetoclax was given after idelalisib in patients. although acute and/or chronic gvhd before bcri was documented in ( . %) and ( . %) patients, respectively, only one patient (treated with idelalisib) reactivated a mild chronic gvhd. none patient received infusion lymphocyte from donor after bcri and one patient underwent a second sct. with a median follow-up of . months ( . - . ) after the onset of the bcri treatment, patients had died, all of them due to cll progression ( richter´s transformation), whereas patients remained in response ( cr, pr). the overall survival probability of the whole series at months was . % ± . %. conclusions: in our study, ibrutinib and idelalisib, administered in cll patients relapsed after sct did not increase the risk of gvhd reactivation but they show high incidence of adverse events. nevertheless, bcri offers a possibility of disease control in these patients with poor prognosis. further studies are needed to confirm these data. background: prior to the introduction of tyrosine kinase inhibitors (tki), median survival of chronic phase chronic myeloid leukemia (cp-cml) patients was approximately months and the standard treatment with interferon-alpha resulted in complete cytogenetic responses in about % of the patients. autologous stem cell transplantation (auto-sct) was first attempted for patients in transformation in order to restore a second cp and was introduced secondarily in cp to try to prolong the response. the main rational for autografting in cp resides on the reduction of the tumor burden and the number of leukemic cells at risk of developing blastic transformation. nevertheless, auto-sct alone was not able to maintain a long-term remission. nowadays, tkis represent the state-of-the-art therapy for cml and the concept of auto-sct has only little interest while long-term follow-up and outcome in this setting are currently unknown. the aim of our study is to evaluate at a first time the longterm outcome of cml patients who received auto-sct in chronic phase, and to evaluate at a second time in a subgroup analysis, the outcome of those who received tki after having been auto-transplanted, mainly for disease progression/loss of response and/or to enhance disease response. methods: we found a total of patients who received auto-sct for cp-cml in europe between years and , ( %) were males, median age at auto-sct was years (range: - ), the median time between diagnosis and auto-sct was months, stem cells source was peripheral blood in % of patients, most frequent conditioning regimen was busulfan mg/kg/day days + day of melphalan mg/m² one day prior to the cells reinfusion. information about receiving tki post auto-sct was available only for patients, first tki was imatinib for ( %) patients, dasatinib for ( %), nilotinib for ( %) and ponatinib for one ( %) patient. results: after a median follow-up of . years (range: - ) from time of auto-sct for the whole population, the probability of overall survival (os) at years was % ( % ci: - ); there was ( %) patients who relapsed after a median time of months after auto-sct. there was a total of patients transplanted before the tki era and survived until the availability of tkis. when we performed a landmark analysis evaluating the outcome of patients who received auto-sct, survived until the tki era and received tki (n= ), the years os probability of these patients from tki treatment was % ( % ci: - ). additional data requests will be sent to centers querying about prognosis, molecular responses, treatment and disease details. conclusions: we demonstrate here with these preliminary results that the introduction of tki has improved survival of cml patients. in addition, patients who received auto-sct, survived until the tki era and also received tki, had encouraging rates of long-term survival. an extensive analysis will be performed when additional data will be available and the study will be updated with more results. disclosure: nothing to declare a year single center transplant experience in chronic myeloid leukemia background: allogeneic hematopoietic stem cell transplantation (hsct) has been considered for decades the only curative approach for patients with chronic myeloid leukemia (cml). in the tyrosine kinase inhibitors (tkis) era, hsct for cml has been reserved only to patients not achieving a cytogenetic remission or showing progressive disease after multiple tki treatment lines. however, a progressive improvement in the long-term survival has been obtained in the overall hsct population. the present study aimed at evaluating whether in cml patients transplanted at our center over a long time period -from to -the outcome improved over time. methods: consecutive patients who underwent a transplant between and were compared to patients who received the transplant between and . overall survival (os), leukemia-free survival (lfs) and graft-leukemia-free survival (glfs) were estimated using the kaplan-meier method and the log-rank test was used to compare risk factors categories. results: of the patients [median age years (range - )], ( . %) were in st or nd chronic phase, ( . %) in accelerated phase and six ( . %) in blast crisis. matched related donors and alternative donors (matched unrelated donors, cord blood or mismatched related donors) were used in and cases, respectively. as stem cell source, bone marrow was used in patients, peripheral blood in and umbilical cord blood in . tbibased conditioning regimens were used in patients, while in the other cases irradiation-free conditioning regimens were used. both in univariate and multivariate analysis, irradiationfree conditioning regimens (hr . ; %ci . - . , p=. ) and transplants performed in st chronic phase (accelerate phase hr . ; %ci . - . , p=. - nd chronic phase hr . ; %ci . - . , p=. -blast crisis hr . ; %ci . - . , p< . ) were associated with a better os. patients transplanted before had a worse os (hr . ; %ci . - . , p < . ) and dfs (hr . ; %ci . - . , p=. ). a trend for a worse glfs was observed in univariate analysis (hr . ; %ci . - . , p= . ), in the first period of observation. conclusions: our single center experience confirms that higher os, dfs and glfs are observed in cml patients allografted in more recent years. improvement of conditioning regimens, use of tbi-free conditioning regimens and supportive therapy, have presumably contributed to these results, together with the more recent strategy of close monitoring of minimal residual disease, and prompt use of tki or donor lymphocyte infusion in case of relapse. hsct is nowadays a safer therapeutic procedure in cml patients that should be considered timely in tki-resistant patients to avoid progression into a more advanced disease phase. disclosure: the authors declare no conflict of interest. reduced-intensity transplantation (rit) in patients with high-risk or advanced chronic lymphocytic leukemia in last years: improvement of transplant outcomessingle centre experience . hct-ci ≥ was in % of pts. source of stem cells was peripheral blood in % and bone marrow in % of pts. the median of infused cd + cells was , x ^ / kg. the conditioning regimen consisted of fludarabine and melphalan (+atg in unrelated donor). gvhd prophylaxis were cyclosporine and methotrexate. results: all pts engrafted. none of pts in cr before rit progressed at day + after rit and among pts beyond cr before rit all of them achieved at least pr at day + after rit. pts ( %) developed acute gvhd ( pts grade iii-iv) and among evaluable pts ( %) of them developed chronic gvhd ( mild, moderate, severe). with median follow-up months (range - months) pts ( %) are alive in cr. pts ( %) relapsed or progressed , and months after rit and of them died. last relapsed patient achieved next cr after ibrutinib. pts ( %) died due to nrm. nrm till day + after rit was %. the estimated probabilities of -years cgrfs, pfs and os are %, % and %. conclusions: in spite of relatively small number of evaluated pts and retrospective type of analysis our data show that rit in pts with high-risk or advanced cll has achieved promising results ( -and -years pfs and os % and % resp. and %) in recent years and these results are better than outcomes of our historical patient cohort from period - ( -and -years pfs % and % resp. os % and %, p= . ) or ebmt published data of pts transplanted for cll in period - ( and -years pfs % and % resp. os % and %). current results of transplantation should be taken into account in our future decision-making process on indications for transplantation in pts with high-risk cll, of course also in the context of new or updated results of targeted cll treatment and its complications. disclosure methods: retrospective data and treatment outcomes were collected from the singapore childhood cancer registry (sccr). most children with cancer in singapore receive therapy at one of the two public paediatric cancer centers (kkh or nuh). a total of thirty two cases were diagnosed with cml and received treatment in either of these centers over a twenty year period ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) . results: the age at diagnosis of the thirty two children ranged from to years (median . years). six patients in the pre-tki era were treated with an upfront hsct. the remainder twenty six patients were initially started on a tki. of these / ( %) had a hsct at a median period of . months from diagnosis (range - months). the reason for hsct in ten out of the twelve children was due to high risk features i.e. accelerated/blastic phase/ no ccr/no cmr. the remaining two patients had a hsct due to parent and patient preference for attempt at upfront cure rather than the use of life-long and expensive tki therapy. non-compliance to tki therapy was a major finding in our teenage cohort. eleven of the eighteen transplants used a matched sibling donor. three patients had cord blood as their stem cell source. one patient had a single antigen mismatched related donor and three patients had a mismatched unrelated donor for their hsct. all patients except one had myeloablative conditioning with busulfan and cyclophosphamide. atg was added according to physician preference. one patient had cy/tbi conditioning because of pre-transplant lymphoid blast crisis. anti gvhd medications included cyclosporine/ methotrexate or tacrolimus and methylprednisolone in the cord transplant patients. six of the eighteen ( %) patients who had a hsct died. four died due to treatment related mortality ( infections, acute gvhd and pulmonary fibrosis). one patient died due to an early relapse and one had a late relapse related mortality. for the pre-tki era, hsct related and year os was % and % respectively. post-tki era and year os was %. for the entire cohort, the year os was %. conclusions: the post-tki era transplant outcomes from our two centers is comparable to leading centers in the world. outcomes for patients with mismatched unrelated donors was poor in our cohort. we recommend a haploidentical related donor transplant or an unrelated cord blood stem cell source for patients when a matched sibling or unrelated donor is not available. clinical trial registry: na disclosure: we have nothing to disclose. fludarabine, busulfan, and thiotepa may be a promising conditioning regimen for myelofibrosis patients undergoing allogeneic stem cell transplantation background: allogeneic stem-cell transplantation (sct) is a curative therapy for patients with myelofibrosis. however, recurrent disease and non-relapse mortality (nrm) are frequent causes of treatment failure. the optimal conditioning regimen for sct in this disease has not been defined. methods: we retrospectively analyzed transplantation outcomes of all adult patients given sct for myelofibrosis between and at a single large academic medical center. patients (n= ) were treated with several conditioning regimens that were grouped according to conditioning intensity. myeloablative conditioning (mac) included busulfan . mg/kg and cyclophosphamide mg/kg (bucy, n= ), fludarabine and busulfan . mg/kg (flu/ bu , n= ) and fludarabine and treosulfan - g/m (flu/ treo, n= ). reduced-intensity conditioning included fludarabine and busulfan . - . mg/kg (flu/bu , n= ). more recently we adopted the tbf regimen including fludarabine, busulfan . - . mg/kg and thiotepa - mg/ m (n= ). all patients were also given anti-thymocyte globulin during conditioning, irrespective of donor source. results: the median age was years (interquartile range [iqr] - ). the majority of patients had documented splenomegaly ( %) and were not previously exposed to ruxolitinib ( %). donor type was an hla-matched sibling ( %), / ( %) or / ( %) matched unrelated donor. the dipps+ score distribution was intermediate- ( %), intermediate- ( %), or high (n= %). the median followup was . years since the success of tyrosine kinase inhibitors (tkis), transplant-related mortality is considered too high to justify allogeneic hematopoietic stem cell transplantation (allohsct) as first-line treatment for chronic myeloid leukemia (cml) patients in chronic phase (cp). allohsct is currently considered for patients failing to at least tkis or with disease in advanced phase. nevertheless, the optimal timing for transplant referral is still not well defined. methods: we performed a retrospective analysis on consecutive patients with cml in cp receiving first transplants from an hla-identical sibling donor with partially t-cell depleted grafts from to at our center. partial t-cell depletion (ptd) consisted of in vitro alemtuzumab incubation of a part of the graft for infusion at day while the rest, containing x cd + cells/kg was given as a t-cell add-back at day . donor lymphocyte infusions (dlis) were provided, in the absence of gvhd, in case of disease relapse or mixed chimerism. molecular monitoring was performed by -month bcr-abl rt-qpcr testing in peripheral blood during at least a -year period after hsct. thereafter, -month testing schedule was maintained where possible, or followed by a -month one. kaplan-meier method was employed to determine the probability of overall survival (os) and leukemia free survival (lfs) since allohsct. results: median age at hsct was years (range, - ). all patients were in first cp but one who was in second cp. twelve patients were tki-naïve at hsct ( hsct ( - , patients had presented suboptimal response or/ and intolerance to imatinib ( - period) , while the last seven patients had presented suboptimal response or/ and intolerance to imatinib, dasatinib and nilotinib ( - period) . the time interval from diagnosis to transplant was < months in / ( %) patients. ( %) patients had an ebmt risk score of - , while ( %) patients of - . the conditioning regimen was myeloablative for all but one patients. the stem cell source was pbsc for patients and bone marrow for one. all patients engrafted. patients presented molecular relapse and one patient hematological relapse with a median interval from transplant to relapse of months (range, - ) . patients received dlis ( for relapse and for mixed chimerism), while patients in relapse also received tki. without prior administration of dli, ( %) patients presented grade ii agvhd and patients moderate cgvhd. after dli, agvhd occurred in and cgvhd in patients. one patient died of disease progression years after hsct and one of myocardial infarction years after hsct. with a median follow-up of . years (range . - . ), -year os and lfs were %. at the time of the analysis / patients were alive and in major molecular response. conclusions: these results of excellent long-term survival and no transplant-related mortality suggest that ptcd improves the outcome of cp-cml patients transplanted from an identical sibling donor and they can be useful for deciding risk-adapted strategies. we believe that ptcd could allow earlier transplant referral of patients failing tkis and having an identical sibling donor. disclosure: nothing to declare single tertiary centre experience in allogeneic haematopoietic stem cell transplantation (allo-hsct) for primary and secondary myelofibrosis (mf) the only curative option for fit patients is allo-hsct. novel therapy is emerging but current recommendation is that eligible patients with life expectancy less than years should be considered for allografting. methods: we retrospectively looked at the clinical features and outcomes of all allo-hsct for mf performed in our centre since . results: patients ( male, female) aged between - years old (median age ) with intermediate- or high-risk mf as per the international prognostic scoring system (ipss) or dynamic ipps (dipps) were transplanted in our centre since . of them ( %) were diagnosed with pmf and the remaining % with secondary mf; post-et and post-pv mf. / of our patient group received a sibling allograft and / a matched unrelated donor allograft ( % received a / human leukocyte antigen (hla)-matched transplant, % a / hlamatched and % a / hla-matched graft). all patients received a reduced intensity conditioning (ric); / patients with fludarabine/ melphalan/ campath (fmc), / fludarabine/ busulphan/ atg (fbatg) and fludarabine/ ara-c/ campath (flag/ campath); all received peripheral blood as source of hsc. engraftment occurred between day - , with a median of d+ . one late graft rejection occurred. all patients were alive at d+ . patients are currently alive; overall survival (os) is %. transplant related mortality (trm) was . % at year, % at years. patient died of graft versus host disease (gvhd) and patients of septicaemia leading to multiorgan failure. acute gvhd grade ii skin occurred in patients, grade iii and above in patients. patients have limited chronic gvhd. / patients received donor lymphocyte infusion (dli) for mixed chimerism (one of which had nd graft failure). out of these patients developed acute grade gvhd and died. response rate: / alive patients i.e . % exhibit no fibrosis in trephine biopsies, / alive patients had residual fibrosis but % donor chimerism, / alive patients had residual fibrosis with mixed donor chimerism, other patient non-assessable. conclusions: allo-hsct remains the only potentially curable option for myelofibrosis. in our centre which serves . million population, with new cases per year, patients were transplanted since . our data suggest that close collaboration between mpn-treating haematologists and transplant physicians is required so that all suitable patients have a transplant assessment early in their disease course. novel molecular prognostic systems are likely to identify those best placed to benefit in future but this series currently supports allo-hsct survival and cure. (range, - ) . dynamic international prognostic scoring system (dipss) score at the time of hct was intermediate- or high risk in patients ( %), intermedate- in patient. molecular evaluation was available in out of : jak v f mutation was detectable in patients, mpl-w k in patient, carl in patient. patients were "triple negative" for driver mutations. cytogenetics information was available for out of ; among which patients had complex karyotype, trisomy and trisomy . patients underwent splenectomy before hct. ruxolitinib was administered in patients before hct. ( %) patients received stem cells from an hla identical sibling, ( %) from a matched unrelated donor and ( %) from an haploidentical sibling. graft source was bone marrow in patients ( %) and peripheral blood in ( %). conditioning was myeloablative in patients ( %), reduced intensity in ( %). all patients engrafted. acute graft versus host disease was absent in patients ( %), grade i-ii in ( %), grade iii-iv in ( %). in evaluable patients chronic graft versus host disease was limited in ( %), extensive in ( %) and absent in ( %). transplantrelated mortality at days was %. main causes of death were: acute gvhd in patients, chronic gvhd in , pancreatitis in , pulmonary aspergillosis in . relapse occurred in patients and was the main cause of death in of them. notably, patients experienced late relapse after . and . years after hct. both of them are living while receiving ruxolitinib therapy. after a median follow up of days (range, - ), out of patients are alive. of them ( %) are disease-free and are living. the kaplan-meyer overall survival and disease-free survival at years was % and %, respectively. conclusions: our experience confirms that hct is a valid option to achieve cure in one third of mf patients. two patients experienced very late (> years) recurrence of mf. the rarity of this condition limits the amount of data and cases available for evaluation and study. life-long follow-up of all mf transplanted patients is warranted to better understand this rare event. disclosure: nothing to declare methods: А -years old female was diagnosed with jak v f-positive pmf, xx, ipss low risk, dipssplus intermediate - risk, subacute budd-chiari syndrome and portal vein thrombosis four years before allohsct. to reduce the splenomegaly and constitutional symptoms we performed pre-transplant ruxolitinib therapy mg daily. after three months of therapy the patient achieved clinical improvement (eln criteria). contrast-enhanced computer tomography and magnetic resonance imaging showed enlarged intrahepatic collateral vessels and signs of portal vein thrombosis with cavernous transformation and multiple dilated collateral veins. gastroscopy documented enlarged esophageal veins. allogeneic stem cell transplantation was performed from / -hla matched unrelated donor with peripheral stem cells ( . x Сd + cells/kg). conditioning regimen consisted of fludarabine ( mg/ m ), busulfan ( mg/kg p.o.). post-transplant cyclophosphamide was administered at mg/kg at day + , + , and ruxolitinib mg was used from d+ till d+ as graft versus host disease prophylaxis. results: starting d+ the patient experienced eight episodes of ebv some of them with severe blood loss. to treat the bleeding episodes blackmore tube was placed six times with temporary effect. to place blackmore tube the patient was two times intubated and required mechanical ventilation. at d+ leukocyte and neutrophil engraftment, full donor chimerism and molecular remission were achieved. platelet engraftment was documented only at d + and poor graft function was present due to cytomegalovirus reactivation (d+ ) and parvovirus b reactivation (d+ ). evb was stopped at d+ only after two esophageal veins ligations, and two procedures of gastric veins sclerotherapy. soon (d+ ) the patient achieved complete platelet recovery (more than x /l) and became red blood cells transfusion independent. at day + complete remission was confirmed by splenomegaly resolution, regression of bone marrow fibrosis, full donor chimerism, jak v f-negative molecular status. cbc showed hb g/l, platelets x /l, leucocytes , x /l. ultrasound examination after transplant documented portal vein thrombosis recanalization. at day + she developed mild (nih) chronic graft versus host disease with eyes and mouth involvement, which was managed with topical steroids. at d+ after transplant the patient is alive in complete remission and has no recurrent bleedings. conclusions: splanchnic vein thrombosis can significantly complicate the course of allohsct in pmf. easy access to surgical, intensive care unit and endoscopic teams is required to make allohsct more feasible in this group of patients. disclosure all patients received treosulfan-based mac regimens, treosulfan(total dose, - gms/m ) was given in combination with different conditioning drugs. the most commonly used regimen was treosulfan, fludarabine ( mgs/m ) and thiotepa( mgs/kg) referred to as ftt that was used in %(n= ). serotherapy was given in % of patients(n= ), as either alemtuzumab or antithymocyte globulin in %(n= ) and %(n= ), respectively. post-transplant graft-versus-host disease (gvhd) prophylaxis was given in all patients, based mostly on ciclosporin. patients( %) received the transplant from identicalrelated donors, patients( %) received the transplant from matched-unrelated donors, and two patients( %) had haploidentical transplants. % of the patients(n= ) were fully hla-matched. all stem cell sources were used as bone marrow in %(n= ), peripheral blood stem cells in %(n= ), and umbilical cord blood in %(n= ). this treosulfan-based conditioning was given as the st transplant in %(n= ), and as the nd transplant after the failure of a first procedure in %(n= ). two patients received treosulfan-based conditioned transplant twice. results: neutrophil engraftment and platelet engraftment occurred at a median of days and days respectively. chimerism was full donor in %(n= ), high donor in %(n= ), and mixed donor in %(n= ). gvhd developed in % of patients(n= ), with acute gvhd grade i/ii and grade iii/iv developed in %(n= ) and %(n= ), respectively. chronic gvhd grade i/ii and grade iii/iv developed in %(n= ) and %(n= ), respectively. all chronic gvhd were mild, limited, non-extensive, and resolved completely. none of our patients had persistent gvhd necessitating long-term systemic immunosuppression. mild vod occurred in %(n= ), and severe vod occurred in %(n= ). one of them died but was believed to be related to the underlying disease (wolman syndrome). viral reactivation occurred in % of patients(n= ), with cmv, ebv, and adenovirus reactivation was found in %, %, and %, respectively. five patients had invasive adenoviraemia that contributed to death in two of them. primary graft failure happened in two patients( %) due to adenoviraemia. seven patients( %) had secondary graft failure with autologous reconstitution. graft failure was significantly lower (p . ) in the ftt group than other conditioning groups. at a median follow-up of months (range, two- months), eleven patients( . %) died, with overall survival of . %, and event-free survival of . %. five patients died due to complications related to their original disease, while six patients died due to transplant-related causes (transplant-related mortality . %). immune reconstitution in alive patients was achieved at a median of eight months. this time was significantly longer (p . ) in ftt group. conclusions: this study demonstrates that treosulfan is a safe and effective conditioning drug that can achieve engraftment, with low rates of graft failure, transplantrelated mortality and morbidity, even if it is used twice in the same patient. disclosure: nothing to declare background: high-dose chemotherapy (hdc) followed by autologous stem cell transplantation (asct) is the treatment of choice for the patients with relapsed or high risk nhl. although the high-dose conditioning regimens commonly used in patients with non-hodgkin lymphoma (nhl) are beam (bcnu, etoposide, cytarabine, and melphalan), beac (bcnu, etoposide, cytarabine, and cyclophosphamide), survival of patients with nhl received above high-dose chemotherapy followed by asct was still unsatisfactory. methods: we prospectively evaluated the efficacy and toxicity of busulfan, etoposide, cytarabine and melphalan (bueam) including iv busulfan instead of bcnu of standard beam as a conditioning for asct in patients with nhl. the high-dose chemotherapy consisted of bu ( . mg/kg i.v. q.d. from day - to day - ), e ( mg/m i.v. b.i.d. on day - and day - ) a ( g/m i.v. q.d. on day - and day - ) and m ( mg/m i.v. q.d. on day - ) at centers in korea. results: two hundred five patients were enrolled onto the study. main subgroup was diffuse large b cell lymphoma (n= , . %), t cell lymphomas (n= , . %), and nk/t cell lymphoma (n= , . %). upfront asct was performed in patients ( . %), and salvage asct in patients ( . %). the disease status of the patients before hdt/asct consisted of patients ( . %) with complete response and patients ( . %) with partial response. treatment related toxicities included nausea in patients ( . %), diarrhea in patients ( . %), anorexia in patients ( . %) and stomatitis in patients ( . %), which were grade i or ii in the majority of cases. the common grade iii toxicities were stomatitis ( . %), diarrhea ( . %), and anorexia ( . %). there were no vod, and transplant-related mortality occurred in patients ( . %), due to infection. one hundred fifty three patients ( . %) achieved a complete response and patients ( . %) after asct, while patients ( . %) showed progressive disease. at a median follow-up duration of . months, the estimated -year overall survival and progression free survival for all patients was . % and . %, respectively. conclusions: the conditioning regimen of bueam for asct was well tolerated and seemed to be effective in patients with relapsed or high risk nhl. disclosure: none of declare background: allogeneic hematopoietic cell transplantation (hct) is potentially curative for high risk acute myeloid leukemia (aml) and myelodysplastic syndrome (mds), however both gvhd and disease relapse remain major challenges. we recently introduced a combination of posttransplant cyclophosphamide (ptcy) and atg ( . mg/kg) as graft-versus-host disease (gvhd) prophylaxis. the purpose of our study was to compare outcomes between ptcy/ atg and other gvhd prophylaxis regimens for high risk aml and mds. methods: we retrospectively investigated outcomes of patients that underwent allogeneic hct between january and july for high risk aml (n= , %) and mds (n= , %). gvhd prophylaxis regimens were compared for overall survival (os), cumulative incidence of relapse (cir) and non-relapse mortality (nrm) in univariate and multivariable analysis. high risk aml was defined as secondary aml, therapy related aml, high risk cytogenetics (eln criteria) in cr , good/ intermediate cytogenetic risk aml in cr and primary induction failure; high risk mds was defined as high/very high risk wpss score. results: median age of patients was years (range - years). donors were matched related in ( %) patients, matched unrelated in ( %) patients and haploidentical in ( %) patients. graft source was peripheral blood stem cells in patients ( %). myeloablative conditioning was used in patients ( %), reduced intensity regimens in ( %) patients. ptcy combined with atg was used in ( %) patients, other gvhd prophylaxis regimens were used in ( %) patients. both donor and recipient were cmv negative in ( %) patients. median follow-up of survivors was months (range - months). univariate analysis demonstrated os of the entire cohort at years was % ( %ci - %), cir at years was % ( %ci - %) and nrm at years was % ( %ci - %). concerning gvhd prophylaxis regimen, -year os for ptcy/atg versus others was % ( %ci - %) versus % ( %ci - %) (p= . , figure) , -year cir for ptcy/atg versus other was % ( %ci - %) versus % ( %ci - %) (p= . ) and -year nrm for ptcy versus other was % ( %ci - %) versus % ( %ci - %) (p= . ). grade ii-iv acute gvhd was seen in % of ptcy/atg patients versus % using other regimens (p< . ). chronic gvhd was observed in % of ptcy/atg patients versus % using other regimens (p= . ). multivariable analysis for os confirmed that the gvhd prophylaxis regimen has no influence (p= . ), while the predominant predictor of survival was age at hct (hr . , %ci . - . , p= . ). for cir, the ptcy/atg combination had no influence compared to other gvhd prophylaxis regimens (p= . ), while ric conditioning was the predominant predictor of relapse (hr . for ric, % p= . ) . for nrm, the atg with ptcy combination demonstrated no significant difference (p= . ), while age at hct was the predominant predictor (hr= . , %ci . - . , p= . ). conclusions: the ptcy/atg combination for gvhd prophylaxis has demonstrated on multivariable analysis similar os, cir and nrm with other previously used regimens at our center. a decrease in atg dose may potentially decrease the relapse rate while retaining the advantage of decreased gvhd. [ background: the combination of fludarabine with myeloablative doses of busulfan (fb ) represents a standard of care conditioning regimen before allogeneic transplantation in patients with myeloid malignancies (giralt, s.: the lancet oncology ). fb has potent antileukemic activity and is associated with low transplantrelated mortality and acute gvhd. however, early after transplantation (days - ), a proportion of patients may not convert to a full donor haemopoietic chimerism, particularly if anti-t lymphocyte globulin (atg) is used as gvhd prophylaxis (rambaldi a, et al.: the lancet oncology ) methods: we retrospective analyzed patients who underwent an allogeneic stem cell transplantation after fb conditioning regimen at our hospital, from november to august . the median age was years (range - ) and diagnoses were aml %, mds % cml % mfi %). the disease status at transplantation was: cr in %, cr in % and active disease in % of patients. the stem cell source was represented by pbsc in more than % of cases and anti-t lymphocyte globulin (atg) was part of the conditioning regimen in more than % of cases at a dose of mg/kg. the donor was a hla identical sibling ( %), a matched unrelated ( %) or mismatched (one allele or one antigen mismatched) unrelated, %. hematopoietic chimerism was molecularly evaluated by variable number of tandem repeats (vntr) on bone marrow (bm) mononuclear cells or peripheral blood (pb) t lymphocytes, purified by immunomagnetic positive selection (miltenyi, biotec). the analysis was performed at day , , , and after transplantation results: after , and days from transplantation, the proportion of patients with a full bm chimerism was %, % and %, respectively. at the same time points, the pb t cell chimerism was %, % and %. before day , patients required the infusion of dli to treat a pending or overt hematologic relapse and patients to convert the lymphoid chimerism from mixed (median %, range - ), to complete (successfully in cases). after day , additional patients required dli to treat disease relapse or progression and patients to improve the chimeric status or the immune reconstitution. at years, the overall survival is %, with a relapse and non-relapse mortality of % and %, respectively ( figure ). by uni and multivariable analysis, aml diagnosis and a mixed bm chimerism before day were associated with relapse and overall survival while age > was the only factor significantly associated with nrm. a mixed pb t-lymphoid chimerism before day does not adversely impact on non-relapse mortality, cumulative incidence of relapse, leukemia-free and overall survival. conclusions: after fb and atg, a progressive increase of pb lymphoid donor chimerism develops gradually after transplantation, in most of cases without the need of dli. early mixed lymphoid chimerism does not compromise the main long-term clinical outcomes and may at least partially explain the low non-relapse mortality. an incomplete bm chimerism within the first months strongly correlates with early disease progression or relapse. background: busulfan (bu) is widely used as a component of myeloablative conditioning regimen before hematopoietic stem cell transplantation (hsct) in children. bu has a narrow cumulative exposure window. the relation of bu exposure with toxicity is well established, but the link between the exposure and efs is not clear due to conflicting reports especially in pediatric patients. obtaining the ratio of bu to its metabolite i.e. metabolic ratio (mr) may serve as an indicator of bu gsh conjugating capacity of an individual, thus cumulative exposure of bu for a particular day that could be used along with auc as a marker to predict efs. the present investigation is aimed at evaluating the utility of bu mr to predict efs in children undergoing allogeneic hsct. methods: two different cohorts with children receiving bu in four times daily (qid, n= ) and once daily doses (qd, n= ) at st. justine's hospital, montreal were studied. bu and su levels were measured on day of the conditioning regimen at the end of infusion (dose in qid or dose in qd dosing). efs was defined from the time of transplant until death, relapse, or rejection, whichever occurred first. a receiver-operator characteristic curve (roc) for bu mrs measured was plotted to show the trade-off in sensitivity vs. -specificity rates for efs, as the cut-off of the test was shifted from low to high. cutoff values were defined based on the youden´s j statistic (i.e. sensitivity+specificty- ). results: twenty-two males and females aged from . to . years (mean±sd: . ± . ) from bu qid cohort had the mean mr of . (sd: . ). a cut off value of . in mr was chosen in roc analysis in this cohort, with better sensitivity ( %) and specificity ( %) for efs prediction (p= . , auc= . ( % ci= . - . ). in qd cohort nine females, and four males aged between . and . years ( . ± . ) had the mean mr of . (sd: . ). in roc analysis, a cut off value of . was chosen with better sensitivity ( %) and specificity ( %) for efs prediction (p= . ; auc= . ). conclusions background: treosulfan is an alkylating agent increasingly used prior to hematopoietic stem cell transplantation (hsct). the main objective of this study was to develop a population pharmacokinetic model of treosulfan in pediatric hsct recipients and to explore the effect of different covariates on treosulfan pharmacokinetics (pk). also, a limited sampling model (lsm) was developed. methods: in this multicentre study, patients, receiving a dose of , or g/m treosulfan a day, administered during consecutive days, were enrolled. a population pharmacokinetic model was developed using nonlinear mixed effect modelling (nonmem version . . , using psn toolkit . . and piraña version . . as modelling environment). demographic factors, as well as laboratory parameters, were included as covariates. results: treosulfan pk was best described by a twocompartment model. a bodyweight-based allometric model improved the model more than a model incorporating body surface area (bsa). clearance (cl) and intercompartmental clearance parameters were . l/h/ . kg ( %ci . - . ) and . l/h ( %ci . - . ). typical volumes of distribution of the central and peripheral compartments were . l/ . kg ( %ci . - . ) and . l ( %ci . - . ). a model-based dosing table based on bodyweight is created to achieve a target exposure of mg*hr/l (table ) , which was the median exposure of our population. estimated glomerular filtration rate (egfr) was shown to be the only parameter that significantly reduced interpatient variability in cl from . % to . %. a limited sampling model with samples (taken at . , and hours after start of infusion) accurately estimated pharmacokinetic parameters of treosulfan. conclusions: to the best of our knowledge, this is the largest cohort of pediatric patients treated with treosulfan used for a population pharmacokinetic study. we developed a two-compartment model with weight and egfr as covariates influencing treosulfan pk. recently we showed a relationship between treosulfan exposure and early toxicity. patients with an exposure > mg*hr/l have an increased risk of developing grade or higher mucositis and skin toxicity. another study in pediatric patients with thalassemia major reported an association between treosulfan clearance (< . l/h/m ) and poor overall survival. our model, together with the limited sampling strategy, can be used to adjust the dose, prior to or during treosulfan administration. ongoing studies conducted in different disease settings will determine if treosulfan exposure can influence patient outcome. subsequently, the optimal target exposure can then be established. background: autologous stem cell transplant (asct) is an effective treatment method for non-hodgkin lymphoma (nhl). until recently, carmustine, etoposide, cytarabine and melphalan (beam) was the most commonly used conditioning regimen. despite acceptable efficacy with beam, carmustine is associated with major pulmonary toxicity. for this reason, the aim of this study was to investigate the safety and efficacy of beb conditioning regimen for asct in nhl. methods: we conducted a prospective, multicenter, phase ii study for beb conditioning regimen for asct in nhl patients. a total of patients were enrolled from centers. they underwent asct with beb conditioning regimen (busulfan . mg/kg for days, etoposide mg/ m for days, bendamustine mg/m for days) between and . [[p image] . two year progression-free survival and overall survival.] results: the median age was years (range - ) and patients ( . %) were men. the most common type was diffuse large b cell lymphoma (n= , . %) and more than half of patients (n= , . %) were classified as ipi score or . eight patients ( . %) had a history of relapse and patients ( . %) received more than lines of chemotherapy before asct. most patients (n= , . %) were complete remission (cr) state at asct. a median number of . x /kg cd cells were infused (range . - . ). all patients engrafted after a median time of days (range - ). twelve patients ( . %) experienced neutropenic fever and patients ( . %) had grade toxicities during asct. however, no one had a documented infection, veno-occlusive disease, or treatment-related death. three months cr rate was . %. during a median follow-up period of . month, patients ( . %) exhibited relapse or progression, while patient ( . %) died of the disease. the estimated -year pfs and os rate were . % and . %, respectively ( figure ). conclusions: the beb conditioning regimens for asct is a feasible with tolerable toxicity in patients with nhl. disclosure: nothing to declare long-term report of total marrow or total lymphoid imrt in advanced leukemia, myeloma and lymphoma background: during the last three decades, total body irradiation (tbi) continues to play an important role in the conditioning regimens for patients undergoing stem-cell transplant (sct) for a wide variety of advanced hematological malignancies. however, tbi showed boundaries in dose limits for toxicity in allogenic and moreover in autologous stem cell transplantation. currently, the choice of conditioning regimen is based on the use of the least-toxic regimen to achieve the optimal therapeutic result. this report aims to assess the feasibility of a conditioning strategy based on high dose chemotherapy and whole-body radiotherapy focused on selective extensive tumor burden irradiation, both in allogeneic and autologous stem cell transplantation. methods: since december , sixty-two patients (pts) have been irradiated by helical tomotherapy (ht) to extensive target before allogeneic or autologous transplantation. selected total marrow irradiation (tmi) schedules were planned to treat patients with high risk acute leukemia (all or aml) or multiple myeloma (mm) as a part of conditioning regimen. total lymphoid irradiation (tli) was planned for patients with refractory or relapsed (r/r) hodgkin (hd) or non-hodgkin lymphomas (nhl). results: tmi and tli allowed delivering therapeutic dose over extensive selected targets with wide reduction of toxicity to all the organs at risk (oars). the higher radiation doses rate to the oars is reduced from % to %. allogenic conditioning regimen was tli ( gy x fx) than fludarabine + endoxan for patients with hd ( pts). tmi ( gy x fx) + fludarabine + melphalan for patients with mm ( pts). tmi ( gy x fx) + thiotepa + fludarabine + busulfan for advanced lam patients ( pts). tmi as the boost ( - gy) after conventional tbi was ( gy in bi-fractionated doses) by cyclophosphamide ( pts). autologous preparation to sct consisted of tli ( gyx fx) followed by high-dose bendamustine and melphalan for patients older than years and conventional feam (fotemustine, etoposide, cytarabine, and melphalan) for younger patients, in hd e nhl ( pts). while tmi ( gy x fx) plus melphalan was delivered for autologous sct in mm and lam ( pts). no unexpected acute toxicity was found. in the allogenic setting, all the patients' engraftment was achieved in all patients. no acute graft versus host disease increasing was detected. within the autologous setting, only % developed grade / mucositis. none experienced grade / extra-hematological toxicity. outcomes of the specific disease will be reported. conclusions: the current report describes the clinical feasibility of using ht to deliver tmi or tli in the setting of autologous transplantation or during allogenic stem cell conditioning regimen, to allow all patients (old, fragile or with high tumor burden) to achieve an ablative regimen before sct. to our knowledge, this single institution experience describes data from one of the largest cohort of patients treated in europe since the development of this irradiation techniques. disclosure induction therapy in both groups of patients was based in polychemotherapy without the use of new drugs. case matching was performed according to age, clinical stage at diagnosis, and response to induction therapy. conditioning regimen consisted of iv bu at a dose of . mg/ kg once a day on days - to - followed by mel at a dose of - mg/m on day - in the bumel group versus mel in the control group. maintenance therapy after transplant consisted of interferon and steroids in the majority of patients. results: the cut-off date for this update was june , . after a median follow-up of and months in the bumel and mel groups respectively, patients had relapsed in the bumel group and patients in the control group. median pfs was ( % ci, . - . ) months in the bumel and ( % ci, . - . ) months in the mel group (p = . ) ( figure ). in this update, patients in the bumel group are in maintained response and of them are in continuous cr (two with negative status for minimal residual disease) between and years after transplantation. ten-year os was not significantly different between both groups, being ( % ci - ) months in the bumel and ( % ci - ) months in the control group.transplant-related mortality was similar in both groups of patients ( % in the bumel and % in the mel group). regarding toxicity, bumel was associated with a higher incidence of mucositis and liver toxicity than the melphalan-only approach but no patient in our series developed sinusoidal occlusive syndrome and the hepatic toxicity observed was only grade i/ii. finally, no long-term side effects have been reported among bumel recipients. conclusions: this long-term follow-up analysis confirms that a therapeutic strategy including bumel as conditioning regimen beforeasct in patients with newly diagnosed mm is highly active and safe in these patients. [[p image] . figure . progression free survival in the bumel (____) and control group (…… frequency of acute gvhd grade iii-iv [cc: %; ct: %; tt: %, p= . ], and transplant-related mortality was higher in tt-carriers (cc: %; ct: %; tt: %, p= . cc&ct vs tt) . ta-tma, cmv infection/reactivation and cgvhd were also not different according to donor genotypes. fungal infections occurred more frequently as causes of death in carriers (cc: . % vs. ct: . % vs tt: . %, p= . ). conclusions: our results suggest that donor tgfb - c>t may exert an adverse influence on the outcome of myeloablative conditioning. our finding might be explained by the combination therapy of calcineurin and mtor inhibition in gvhd prophylaxis in myeloablative conditioning. disclosure: nothing to declare. treosulfan-based reduced intensity conditioning in hla-haploidentical transplantation using ptcy as gvhd prophylaxis in high-risk mds /aml of the elderly background: standard conditioning regimens prior to allogeneic hematopoietic stem cell transplantation (allo-hsct) are often associated with a considerable risk of severe adverse events, especially in elderly patients suffering from high-risk (hr) mds/aml. previous clinical studies have demonstrated feasibility of treosulfan-based reduced-intensity conditioning (ric) by stable engraftment, low non-relapse mortality (nrm), and favorable survival in elderly patients undergoing hla-matched related or unrelated allo-hsct (beelen et al, ash # ). however, data for treosulfan-based conditioning in the t-cell-replete hlahaploidentical (haplo-hsct) setting in high-risk aml/mds patients are rare. here we report on the outcome of eleven patients treated with a treosulfan-based conditioning undergoing haplo-hsct using exclusively post-transplantation cyclophosphamide (ptcy) as gvhd prophylaxis. methods: eleven patients with high-risk (hr) aml (n= )/mds (n= ) who underwent haplo-hsct using treosulfan for reduced intensity conditioning (ric) and ptcy as gvhd prophylaxis were retrospectively analyzed with respect to outcome and toxicity. all patients were > years old and transplanted between january and february at our institution. the majority of the patients ( / ) suffered from active disease at time of treatment initiation, only two patients presented in cr. all but one received sequential conditioning with cytoreductive chemotherapy using flamsa applied shortly prior to treosulfan-based ric ( g/m over days). a bone marrow graft was used in / patients. post-grafting immunosuppression consisted of cyclophosphamide, tacrolimus and mmf. national cancer institute common terminology criteria for adverse events version . were used for nonhematologic toxicity assessment starting from sequential therapy initiation or conditioning until day + . results: median age of the entire cohort was years (range: - ). the hct-ci was ≥ in eight pts (median hct-ci= , range: - ). no graft rejection occurred. neutrophil and platelet engraftment were achieved in % and % of the patients at a median of ( - ) and . ( - ) days, respectively. acute gvhd grade ii-iv occurred in % of the patients, exclusively involving the skin. no one developed severe (°iii-iv) acute gvhd. no patient died prior to haplo-hsct. severe nonhematologic regimen-related toxicities (°iii-iv) occurred in / patients, predominately affecting the gastrointestinal tract. no patient suffered from ≥two iii-iv°toxicities. all patients developed fever during treatment course, four with positive blood cultures. cmv reactivated in / patients at risk. no ebv reactivation or ptld occurred. six patients had clinical and radiological signs of pneumonia (probable invasive aspergillosis) without detection of aspergillus/antigen in the bronchoalveolar lavage. ci of nrm at day + was %. four patients relapsed within the first year after haplo-hsct, with two of them dying due to relapse. at last follow-up (dec ) / patients were alive. with a median follow-up of months ( - ) estimated -year os and dfs were % and %, respectively. conclusions: treosulfan-based unmanipulated hlahaploidentical allo-grafting using ptcy as gvhd prophylaxis in hr mds and aml patients aged over years is safe and well tolerated resulting in stable engraftment and a favorable toxicity profile. our preliminary data further show promising outcome with low nrm, no severe acute gvhd and favorable survival offering an attractive alternative in ric for haplo-hsct of the elderly. disclosure: nothing to declare comparison of outcomes of total body irradiation (tbi) vs non-tbi conditioning regimens in acute lymphoblastic leukemia for allogeneic transplantation background: in adult patients diagnosed acute lymphoblastic leukemia (all) long-term results are poor with intensive chemotherapy. allogeneic stem stem cell transplantation is the potential treatment that provides cure for these patients. myeloablative preparation regimens include total body irradiation (tbİ)+ cyclophosphamide(cy) and busulfan + cyclophosphamide.in adult all patients wbi/cy widely used, but the toxicity rate is higher. the aim of this study is to compare the result and effect of the tbi/cy and busulfan/cy regimens in allogenic bone marrow transplantation in all patients. methods: between - there were all patients who underwent transplantation using myeloablative preparation regimen with or without addition tbi in the adult bone marrow transplantation units of medipol medical faculty, istanbul university istanbul medical faculty, sisli florence nightingale hospital, atakent acıbadem hospital adult bone marrow units . we analyzed overall survival (os), progression free survival (pfs), veno occlusive disease, acute and chronic graft versus disease development rates in these patients. results: demographic characteristics of patients summarized in table - there was no significant difference between groups in donor age, gender, stem cell source. it was observed that the relapse rate was not statistically significant in both group.there was no statistically significant difference between the patients who underwent myeloablative regimen and myeloablative regimen with tbi in relaps,death, os, pfs. (figure- ) [[p image] . figure ] in terms of transplant complications there was also no respectable difference in development of vod and acute and chronic graft versus disease but vod was more common in the group that did not use tbi (p: . ) ( conclusions: although there are contradictory data in the literature, in our multicentre study, it was revealed that the addition of tbi in the myeloablative preparation regimen compared with myeloablative preparation regimen alone did not have a positive or negative effect on overall survival.we think that if we can prepare a good vod prophylaxis approches, we can give up tbi in future. disclosure (n= ) . for gvhd prophylaxis, cyclosporine a was given either alone (n= ), with mmf (n= ) or with methotrexate (n= ). the graft source was bone marrow (bm) in most cases (n= ), pbsc in seven cases, matched sibling cord +bm in two cases and one matched related cord. twentyfive of the donors were family donors and ten were unrelated. twenty-nine of the donors were / hla matched, six were / mismatched and one haploidentical. four patients had engraftment failure and required a second transplant, two of them were re-transplanted with cyclophosphamide and tbi, one with fludarabine, busulfan and campath, and one with no conditioning. thirty of the patients are alive ( %). four patients died of transplant complications and one died of metastatic squamous cell carcinoma. eight survivors are mixed chimeras ( %- % donor) and are all doing well, none of them developed any gvhd. nine patients developed acute gvhd, four of them with grade - . seven of these patients later developed chronic gvhd, two of them have extensive disease. conclusions: our results show a high survival rate of %, with a low rate of engraftment failure and reasonable rates of gvhd. only one of our patients died of late effects of hsct for fa. mixed chimerism does not seem to present a problem. we conclude that reduced intensity fludarabine based conditioning regimens are a good treatment option for patients with fanconi anemia undergoing hsct. disclosure: nothing to declare total marrow irradiation + bendamustine as reducedtoxicity myeloablative conditioning prior to allohsct for younger patients with multiple myeloma background: the prognosis of patients with multiple myeloma (mm) has improved markedly over the last two decades. despite that, allohsct remains the only treatment option with curative potential. however, its use is limited due to high incidence of non-relapse mortality (nrm) after myeloablative conditioning while insufficient efficacy of reduced-intensity regimens. we developed a new protocol characterized by reduced toxicity while preserved myeloablative potential, based on the use of total marrow irradiation (tmi) in combination with bendamustine. the aim of this study was to evaluate its safety and efficacy in a singlecenter experience. methods: between years - , mm patients below years old were offered tandem auto-allohsct as part of first-line therapy. the decision was based on individual patient preferences after detailed description of potential risks. autohsct was preceded by melphalan mg/m iv. the conditioning prior to allohsct consisted of tmi performed using helical tomotherapy at the dose of gy/d on days - , - , - (total gy) and bendamustine - mg/m /d iv. on days - , - (total - mg/m ). the immunosuppressive therapy consisted of cyclosporine + methotrexate +/-atg. peripheral blood was used as a source of stem cells. results: the analysis included patients (women - , men - ). the median follow-up was ( - ) months. the median age at allohsct was ( - ) years. the disease stage before allohsct was as follows: cr- , vgpr- , pr- . patients were treated with hsct from either hlamatched siblings (n= ) or unrelated donors (n= ). the interval between autohsct and allohsct was ( - ) months. all patients engrafted after allohsct with median time of neutrophil and platelet recovery of and days, respectively. one patient ( %) experienced grade acute gvhd, while there were no cases of grade - acute gvhd. the incidence of mild, moderate and severe chronic gvhd was %, % and %, respectively. the rate of grade non-hematological toxicities was %. one patient died of late bacterial infection. the incidence of trm was %. grade adverse events were not reported. disease status months after allohsct was: cr- , vgpr- , pr- . the probability of os and pfs after months was % (+/- %) and % (+/- %), respectively. the incidence of progression and trm was % and %, respectively. conclusions: allohsct using tmi gy + bendamustine conditioning protocol is characterized by good tolerance and low risk of gvhd. it may be used for younger patients with mm as part of tandem auto-allohsct strategy. encouraging results reported in this study should be confirmed in prospective clinical trials. disclosure: nothing to declare p comparison between two reduced intensity conditioning regimens in patients with a myeloid malignancy: a single center experience comparing fb with flumel background: hematopoietic stem cell transplantation (hsct) remains the only curative option for high-risk myeloid neoplasms. the optimal reduced-intensity conditioning (ric) is still debated. methods: a single-center retrospective analysis was conducted at our institution to compare two different ric regimens in adult patients transplanted for myeloid malignancy from to . a total of patients were analysed, of them treated with busulfan-based (fludarabine mg/m , busulfan . mg/kg, fb ) and with melphalan-based conditioning regimen (fludarabine mg/m ,melphalan mg/m , flumel). antithymocyte globulin (atg) was administered in all patients while no one received tbi. partial in vitro t-cell depletion was performed using alemtuzumab for low risk patients. results: the two groups were well balanced with a median age of and years in the fb and flumel group, respectively, and a median follow up of months. the most frequent indication for transplant in both groups was aml ( . and . % for fb group and flumel group, respectively) and the stem cell source was peripheral blood in . and . % of patients. more patients in the first group had near to significant worst karnosfky status (< ) at transplant compared to second ( . vs %, p= . ) and more patients received a t-partial depleted graft ( . vs . %, p= . ). the neutrophil engraftment was significantly shorter after flumel ( vs days, p < . ). the -year overall survival (os) and disease-free survival (dfs) were of . and . %, respectively, after fb and . and . % after flumel, respectively, and were not significantly different (p=. for os and . for dfs), with a karnofsky >= being the only factor significantly associated in univariate analysis with better os and dfs (p=. for both). the cumulative incidence (ci) of grade to acute graft-versus-host disease (agvhd) was . % after fb and . % after flumel (p< . ) and was associated in multivariate analysis with both t depletion and ric type (p< . and . , respectively). the ci of chronic gvhd at years was . % in fb and . % in flumel group (p=. ) . the ci of non-relapse mortality at years was . % after fb and . % after flumel (p=. ). the ci of relapse at years was . % for the first and . % for the second group (p< . ) and was associated with conditioning regimen in multivariate analysis (p=. ). no difference in -years gvhd-free/ relapse-free survival (grfs) was observed between the two group ( . % for fb and . % for flumel, p=. ). conclusions: when comparing two ric regimens for myeloid neoplasms, we observed a higher incidence of agvhd after flumel whereas no statistical difference was noted for the cgvhd occurrence. while the toxicity appears to be higher after flumel, this result is counterbalanced by a higher proportion of relapse after fb , accounting for no difference in os, dfs and grfs between the two groups. these findings could be partially explained by a larger proportion of patients receiving a partial t-depletion after fb ric, but a larger trial is needed to clarify this issue. disclosure: nothing to declare. once-daily vs -times daily intravenous busilvex in conditioning regimen before allogeneic stem cell transplantation for patients with myeloid malignancies: safety and efficacy background: busilvex (bu) is part of standard conditioning regimen before allogeneic stem cell transplantation (asct) for patients with myeloid malignancies and usually administered as an intravenous (iv) infusion -times daily. this study aimed to compare the saftey and efficacy of this schedule to a once-daily iv bu. we conducted a retrospective study in adult patients (≥ years) with myeloid malignancies who received asct from hla-identical sibling donors between january and june following iv bu-based preparative regimens. graft-versus host disease (gvhd) prophylaxis consited of cyclosporine and short course of methotrexate. intravenous bu was administered -times daily ( . mg/kg every hours x to doses) or oncedaily in a -hour infusion ( . mg/kg x to days) since june . results: ninty-nine patients were enrolled ( men and women). median age was years (range, - y). the median time from diagnosis to asct was months (range, days - years). diagnosis were acute myeloid leukemia (n= , %), chronic myeloid leukemia (n= , %), myelodysplasic syndrome (n= , %), primitive myelofibrosis (n= , %) and chronic myelomonocytic leukemia (n= , %). thirty-seven ( . %) patients had ebmt-score ≥ . sixty-five ( . %) patients were transplanted in cr , ( %) beyond cr and ( . %) had active disease. conditioning regimens consisted of bu/cyclophosphamide in patients ( . %), bu/fludarabine in patients ( . %). four-times daily bu was given to patients ( . %, groupe ) and once-daily bu to patients ( . %, groupe ). stem cell source were bm in patients ( . %) and pbsc in patients ( . %). globally, patients characteristics were well balanced between the two groups. the rates of severe complications were similar between the two groups with no statistically significant differences except oral mucositis (table ). non-relapse mortality (nrm) was comparable in the two groups ( % and % in groups and , respectively, p= . ). the relapse rate was % and %, respectively (p= . ). after a median follow-up of years (range, days - years), the os was not significantly different between groups and : % vs % (p= . ). however, the rfs was significantly better in the groupe : % vs % (p= . ). conclusions: once-daily iv bu regimen seems to be an efficient and safe alternative to the -times daily protocol. however, results should be interpreted with caution because the historical comparison and lack of bu pharmacokinetics studies. disclosure background: standard therapy of the most patients with juvenile myelomonocytic leukemia (jmml) is allogeneic hematopoietic stem cell transplantation (ahsct). the choice of optimal conditioning regimen for patients with jmml is crucial as well as long-term observation. we aimed to estimate the long-term follow-up and survival rates of patients with jmml after ahsct with the help of busulfan or treosulfan-based conditioning regimens. methods: thirty eight patients with jmml underwent ahsct in - . we compared equal groups of patients received busulfan (n= ) and treosulfan-based (n= ) conditioning regimen. m:f= : . median of age at hsct was . ( . - ). donor type: hla-related / - % (n= ), hla-related / - . % (n= ), hlaunrelated / - . % (n= ), hla-unrelated / - . % (n= ), and haploidentical - . % (n= ). stem cell source: bm - . % (n= ), pbsc - . % (n= ), ucb - , % (n= ), and ucb+bm - . % (n= ) . disease status on hsct: cr - . % (n= ), refractory - . % (n= ). results: median follow-up . months ( - months) . the estimated -year overall survival (os) probability in patients received busulfan-based conditioning was , ± , % in comparison with , ± , % in patients with treosulfan-based regimen (р= , ). event-free survival (efs) was , ± , % in group with busulfan-based regimen and , ± , % in patients with treosulfanbased conditioning (р= , ). background: post-transplant relapse remains the leading cause of treatment failure in high risk (hr) acute myeloid leukemia (aml), myelodysplastic syndrome (mds), myeloproliferative neoplasia (mpns) receiving allogeneic hematopoietic cell transplantation (allo-hct), especially for patients with relapsed or refractory aml. recently, a sequential transplant approach, as developed by the munich group, comprising of intensive cytoreductive chemotherapy flamsa (fludarabine/amsacrine/cytarabine) to decrease leukemia cell burden shortly prior to conditioning regimen, has been successfully used for high-risk (hr) aml/mds with promising results. methods: we studied patients (median age years, range - ) with hr aml (n= ), as defined by refractory, relapsed disease, secondary leukemia, or high/ very risk disease risk index risk, and hr mds (n= ) according to ipss-r, undergoing allo-hct using the sequential transplant approach in institutions between january and october . the sequential transplant approach combined a cytoreductive chemotherapy, which consisted of either flamsa (n= ), flag +/-ida (fludarabine/cytarabine/granulocyte colony stimulating factor /idarubicin) (n= ), or clo-arac (clofarabine/cytarabine) (n= ), followed by reduced (ric) (n= ) or myeloablative (mac) (n= ) conditioning regimen. all patients received peripheral blood stem cell from matched related donors (n= ) matched unrelated donors (n= ), or mismatched unrelated donors (n= ). post-grafting immunosuppression consisted of calcineurin inhibitor and mycophenolate mofetil in all patients. thymoglobulin was added for gvhd prophylaxis for unrelated donor transplant. results: the median time to neutrophil > /μl was days (range, - ) . with a median follow-up of . months (range, . to . months), the kaplan-meier estimate of leukemia-free (lfs) and overall survival (os) at years were % ( % ci, - ), % ( % ci, - ), respectively. patients receiving flag or clo-arac based sequential regimen showed a trend towards more favourable overall survival (os) as compared to patients given flamsa ( year os: % vs %; p= . ). at years, the cumulative incidences of relapse and non-relapse mortality (nrm) were % ( % ci, - %) and % ( % ci, - %), respectively. in multivariate analysis, the type of sequential conditioning regimen did not show any significant impact on lfs, os, nrm or relapse. conclusions: sequential transplant conditioning with flamsa, flag or clo-arac followed by allo-hct is an effective strategy in overcoming the dismal prognosis of hr aml and mds, and enabling long-term disease free survival. more studies on effective strategies such as posttransplant maintenance therapy of prophylactic donor lymphocyte infusion, are needed to further eliminate the risk of relapse, without increasing risk of treatment related toxicity. disclosure: nothing to declare optimization of the blood sampling procedure for busulfan therapeutic drug monitoring (tdm) to optimize our sampling scheme ( minutes, , , , , , and hours after the end of a -hour infusion), we reduced the number of blood samples collected, reducing nursing and laboratory staff time and increasing patient convenience. this study aims to show the performance of a simplified sampling protocol which includes the first samples from the original protocol. methods: individual pk parameters were retrospectively estimated using samples (simplified protocol) and were compared with those obtained after samples (original protocol). individual pk parameter values for a one compartment model were estimated using a maximum likelihood estimation modelling algorithm (adapt . ) and the statistical analysis of the results was performed (statgraphics centurion xv). results based on the approved dosage recommendations, mean (sd) initial dose was . ( . ) mg. after tdm, mean (sd) calculated dose at day for the remaining days (to achieve the defined target cumulative auc) was , ( , ) mg obtained from the original protocol. according to the simplified protocol the result would be , ( , ) mg. the median and the mean variation of the calculated dose were % and % ( - %) between protocols. a strong relationship between the cl of the day - obtained from the original protocol and the simplified protocol is observed (r = . ). this high correlation is also observed for patients with busulfan t / > h (r = . ), a population were the reduction of sampling could be more problematic. anova test for the log cl with the factors: patient, day of busulfan and type of sampling protocol was performed. sampling protocol was determined as non-statistically significant (p = . ). conclusions: results suggest that both protocols are equivalent concerning to the busulfan cl estimation and calculated auc. variation between protocols regarding the calculated dose at day for the remaining days to achieve the defined target cumulative auc is considered acceptable. we verified a strong relationship between busulfan cl obtained from both protocols and sampling protocol doesn't influence cl statistically. a reduced sampling collection of determinations until h after the end of the infusion is shown to be sufficient for the tdm of busulfan, so this was implemented in our centre in line with published data. disclosure: nothing to declare p impact of anti-thymocyte globulin doses in unrelated hematopoietic stem cell transplantation for patients with myeloid neoplasm background: anti-thymocyte globulin (atg) is widely used for the prophylaxis of graft-versus-host disease (gvhd) in hematopoietic stem cell transplantation (hsct). however, there is still controversy regarding the optimal dose of atg. therefore, we analyzed the impact of atg doses in unrelated hsct for patients with myeloid neoplasm. methods: this was a retrospective multi-center study that assessed the impact of atg doses on clinical outcomes in patients with acute myeloid leukemia (aml) or myelodysplastic syndrome (mds) undergoing an unrelated hsct. the patients who received peripheral blood stem cells (pbsc) transplantation after conditioning regimens containing i.v. busulfan (bu), fludarabine and rabbit atg between and were included in this study. results: a total of patents, median age years, with aml (n= ) or mds (n= ) were included in our analyses. patients ( %) received a myeloablative regimen (i.v. bu> . mg/kg). high-atg (atg mg/kg), intermediate-atg (atg . - mg/kg) and low-atg (atg mg/kg) were given in , and patients, respectively. after a median follow-up of months, the cumulative incidence of extensive chronic gvhd was . % in the high-atg group, . % in the intermediate-atg group and . % in the low-atg group (p= . ). conclusions: our study shows that the incidence of extensive chronic gvhd was similar regardless of the doses of atg after transplantation of pbsc from unrelated donor for patients with aml or mds. however, the rate of relapsefree survival and the rate of a composite end point chronic gvhd-free and relapse-free survival were significantly higher in the intermediate dose ( methods: we retrospectively retrieved data from the electronic medical records for consecutive patients aged and older, who underwent an asct for lymphoma over the last years at our institution. results: forty four patients ≥ years old underwent asct between aug and aug . twenty eight of them received a reduced-dose conditioning (median %, range %- % dose reduction). the dose was reduced for % of patients ≥ years old and for % of patients aged - . the outcomes of the following three groups of patients were compared: a) age ≥ ; without dose reduction, b) age - ; with dose reduction and c) age ≥ ; with dose reduction (table ). only one patient aged received full-dose conditioning. there was no significant difference between the groups in the number of previous chemotherapy cycles (median , range - ). however, significantly more patients at the age of - were in complete remission (cr) pre-transplant in both full and reduced-dose conditioning groups (a and b). no significant intergroup differences were observed in the occurrence of complications (mucositis and infections), day transplantrelated mortality (trm) or engraftment day. similarly, no significant differences were found either in the -year progression-free survival (pfs), which was %, % and %, or -year non-relapse mortality (nrm), which was %, % and %, respectively for groups a, b and c. the -year overall survival (os) tended to be higher in group b ( %), compared to groups a ( %) and c ( %). conclusions: beam/beac conditioning dose reduction was not found to adversely affect -year pfs and os rates. despite the fact that / of the patients in the age group ≥ underwent asct in partial remission and had dose reduction, theier achieved trm, pfs and os rates were similar to those of patients aged - . beam/beac conditioning at a %dose may be a suitable option for patients in their seventh decade requiring asct. this strategy should be further evaluated in prospective clinical trials. background: the transplant related mortality in autologous transplants for lymphoma and multiple myeloma, reported worldwide ranges from - %. from - , the trm at our center for these two diseases was approximately %. we introduced changes in mobilization schedule, conditioning regimens and drug dosages to determine whether these changes affected the transplant related mortality and overall survival. methods: from april -december , we used beam (bcnu: mg/m on day - ; etoposide mg/ m on days - to - , cytarabine mg/m on days - to - and melphalan mg/m on day - as conditioning chemotherapy for patients admitted in transplant unit for autologous transplants in hodgkin's and non-hodgkin's lymphoma. in patients with multiple myeloma high dose melphalan ( mg/m ) was used. the mobilization protocol consisted of cyclophosphamide . gm/m followed by gcsf μgm/kg twice daily till stem cell collection was completed. from january , we changed the beam protocol to bendaeam with dose modifications that included: bendamustine mg/m on days - and - , cytarabine mg/m on days - to - , etoposide mg/m on days - to - and melphalan mg/m on day - . for multiple myeloma melphalan was reduced to mg/m . we used only gcsf for mobilization of stem cells, which was continued till stem cell harvest was complete. response to treatment was evaluated by comparing trm and overall survival for two time periods: - and from till date. results: from april till december , n= autologous transplants were performed. the male:female ratio was . : . fifty seven patients underwent transplant for lymphomas, n= for multiple myeloma and n= for other diagnosis. median age was ± . ( - years). the mean mnc was . × ± . /kg. engraftment was achieved in % of patients. the transplant related mortality was . % and overall survival was % (follow up: months). since january till march we have performed n= autologous transplants of which n= were males. fifteen transplants were performed for lymphomas (nhl: , hd: ) and n= for multiple myeloma. median age was ± ( - years). the mean mononuclear cell count was . x /kg and the mean cd count was . x /kg. engraftment was achieved in all patients. the transplant related mortality was % and the overall survival was % (follow up months). conclusions: we were able to reduce the autologous transplant related mortality to % by decreasing dosages of conditioning chemotherapy and changing the mobilization protocol. long follow-up is needed to determine late mortality and late relapse in comparison to standard chemotherapy dosages disclosure: nothing to declare risk and benefit of thiotepa based conditioning followed by autologous stem cell transplantation in high risk lymphomas years ( - ) . stage (ann-arbor) at diagnosis of hd/dlbcl/pcnsl: stage ie n= / / ( / / %), stage ii n= / / ( / / %), stage iii n= / / ( / / %), stage iv n= / / ( / / %). median time from diagnosis to asct hd/dlbcl/pcnsl: / / month ( - ). induction treatment in hd patients was abvd, in most dlbcl patients r-chop and in pcnsl patients high dose methotrexate and cytosin arabinoside. tumor status at asct hd/dlbcl/pcnsl: complete metabolic remission (cmr) n= / / ( / / %) and from pcnsl patient's n= ( %) were in first complete remission (cr ). type of stem cell graft was periferial blood stem cell in all case. conditioning: thiotepa ( mg/ m on days - to - , busulphan , mg/kg on days - to - and cyclophosphamide mg/kg on days - to - plus rituximab mg/m on day - in dlbcl and pcnsl. median follow up from asct days . tumor stage at asct was defined with computer tomography with positron emission tomography (pet-ct). results: median time of engraftment was days ( - ). thiotepa caused toxicoderma appeared at ( %) patients. cytomegalovirus (cmv) reactivation was seen in ( %) cases with low dna content ( , , copies/ml) and responded completely to oral valgancyclovir therapy. transplantation related mortality hd/dlbcl/pcnsl n= / / ( / / %), in cases bacterial sepsis and one systemic mycoses and one pulmonary fibrosis. incidence of long-lasting grade iii-iv thrombocytopenia and anaemia: n= ( , %) and n= ( %), median time of duration from transplantation days ( - ) and days ( background: this study evaluated the efficacy and toxicity of intravenous busulfan and thiotepa as a conditioning regimen for autologous stem cell transplantation (asct) in patients with multiple myeloma (mm). methods: we retrospectively analyzed the data of patients with mm who received the intravenous busulfan and thiotepa conditioning for asct between november and april in korea. results: the median time to transplant was . months, and patients ( . %) underwent asct within months of the diagnosis. the overall response rate after asct was . %, including . % with complete response, . % with very good partial response, and . % with partial response. the most common severe non-hematologic toxicity (grade - ) was infection ( . %). three patients ( . %) developed venous-occlusive disease. one patient ( . %) died due to severe pneumonia after asct. after a median follow-up of . months, the median progression-free survival (pfs) and overall survival (os) were not reached. conclusions: in conclusion, a conditioning regimen of intravenous busulfan and thiotepa was effective and tolerable. clinical trial registry: not applicable disclosure: the authors have declared no conflicts of interest. myeloablative haploidentical bone marrow transplantation with post-transplant cyclophosphamide in paediatric patients with haematological malignancies santanu sen , sameer tulpule background: haploidentical transplants have been shown to be safe and effective in treating haematological malignancies in the paediatric population. we have previously reported on our experience of using reduced intensity conditioning with post transplant cyclophosphamide in haploidentical patients. we herein report our experience of using a tbi based myeloablative conditioning to treat our first patients with haematological malignancies. methods: patients were enrolled in the study, with relapsed acute lymphoblastic leukemia (all) and with relapsed/resistant acute myeloid leukemia (aml). all aml patients had genetic markers of high risk disease and all all patients had very early relapses (either on therapy or within months of stopping therapy). all patients were conditioned with an identical protocol using tbi-based myeloablative preparative regimen (fludarabine mg/m /d × d and tbi cgy bid on d − to − [total dose cgy]) followed by an infusion of unmanipulated peripheral blood stem cells from a haploidentical family donor. postgraft immunosuppression consisted of cyclophosphamide mg/kg/day on days and , mycophenolate mofetil through day , and tacrolimus through day . results: median time of neutrophil and platelet engraftment was and days, respectively. all patients achieved sustained complete donor chimerism by day + . acute gvhd, grades ii-iv and iii-iv, was seen in % and %, respectively. disease progression occurred in patients: & months after transplant and there was one death due to severe fungal infection. estimated twoyear survival and relapse were % and %, respectively. patients had severe bk viremia and cmv reactivation occurred in patients. all patients were successfully managed with appropriate supportive and antiviral therapy. conclusions: we report good outcome with a myeloablative conditioning in haploidentical transplants with excellent engraftment and hopefully a longer life expectancy. with small number of patients, it is difficult to state whether using a myeloablative conditioning would lead to better long term outcomes in this cohort of patients with very haematological malignancies, but we certainly showed that it is possible to achieve excellent early results. disclosure: nothing to declare fludarabine in combination with melphalan and atg can be the best conditioning for hematopoietic stem cell transplant of children with hemophagocytic lymphohistiocytosis methods: in this prospective study, we analyzed the outcome of two pediatric patients with hlh who had received hsct, using reduced-intensity conditioning (ric) regimen. they received the same ric regimen based on the use of fludarabine ( mg/m /day for days) in combination with melphalan ( mg/m /day for days) and horse antithymocyte globulin (atg mg/kg/d for days). cyclosporine and methotrexate were used as graft-vs.-host disease (gvhd) prophylaxis. results: a months boy with primary hlh (fhl ) was transplanted from his mother and a years girl with secondary hlh was transplanted from her brother. both of donors were hla match with their recipients. they were received x /kg and x /kg cd + cells from the harvested peripheral blood stem cells, respectively. they achieved full neutrophil and platelet recovery. the time to neutrophil recovery was and days, respectively. full chimerism was achieved for both of them. in addition, they was developed grade and of acute gvhd, respectively. gvhd was completely controlled with prednisolone. they are alive and in complete remission without any significant complications after and months, respectively. conclusions: it appears that fludarabine in combination with melphalan and atg may be the best conditioning regimen for hematopoietic stem cell transplant of children with hlh. due to a few number cases of this study, a study with sufficient sample size is required. disclosure background: hematopoietic cell transplant (hct) recipients often report depression and impaired quality of life (qol) before transplant. mixed evidence suggests depression may be a risk factor for greater mortality and worse qol. inconsistent findings may be due to the fact that previous studies have not evaluated antidepressant use. the aim of the study was to compare pre-transplant patientreported physical functioning and post-transplant overall survival (os) between four groups of hct recipients: ) non-depressed/taking antidepressant (treated depression), ) depressed/taking antidepressant (undertreated depression), ) depressed/not taking an antidepressant (untreated depression), and ) not depressed/not taking an antidepressant (control). it was hypothesized that physical functioning and os would be worse among patients with untreated and undertreated depression relative to those with treated depression and controls. methods: this retrospective case-control study included patients completing depression (phq- ) and quality of life (sf- ) questionnaires at pre-transplant. analyses were conducted separately for allogeneic and autologous recipients. results: participants (n= , ) were % men, mean age years ( - ), % allogeneic recipients. regarding depression and antidepressant use, ( %) allogeneic patients were characterized as having treated depression, ( %) as untreated depression, ( %) as undertreated depression, and ( %) as controls. hierarchical linear regression models indicated that after adjusting for significant univariate factors (performance status, disease status, and regimen intensity), allogeneic patients with treated depression (b=- . , % ci=- . , - . ) reported better physical functioning than patients with undertreated depression (b=- . , % ci=- . , - . ) and untreated depression ) but worse physical functioning than controls (p values < . ). cox regression models indicated depression/antidepressant usage was not associated with os among allogeneic patients (p values> . ).among autologous patients, ( . %) were characterized as having treated depression, ( . %) as untreated depression, ( . %) as undertreated depression, and ( . %) as controls. hierarchical linear regression models indicated that after controlling for significant univariate factors (gender, performance status, diagnosis, and disease status), autologous patients with treated depression (b=- . , % ci=- . , - . ) reported better physical functioning than patients with undertreated depression (b=- . , % ci=- . , - . ) and untreated depression (b=- . , % ci= . , - . ), but worse physical functioning than controls (p values < . ). cox regression models showed depression/antidepressant usage was associated with os (p values < . ), with patients with treated depression demonstrating significantly worse os than other groups (p= . ), but this association was no longer significant in multivariate analyses controlling for diagnosis and disease status (p= . ). conclusions: patients with untreated or undertreated depression pre-transplant may benefit from depression screening and treatment to improve physical functioning. disclosure: hslj: consultant for redhill biopharma and janssen scientific affairs p eltrombopag (epag) induces a high percentage of responses in patients with post allo-hsct poor graft function (pgf) and no active gvhd lourdes aguirre , aitziber lizardi , pilar bachiller , brigida esteban , carmen gonzález , nagore argoitia , maría araiz , aranzazu aguirre , anunciación urquía , carlos vallejo background: persistent cytopenia is a life-threating complication after hsct. several causes can lead to this situation (viruses, gvhd, drugs, etc) . a specific entity is the one called "poor graft function (pgf)", which is diagnosed in pts with ≥ cytopenias after day + , in the presence of donor chimerism and the absence of gvhd or relapse. pgf is more frequent after alternative allo-hscts, such a haplo-identical, mismatched, or ucb. several therapeutic approaches for pgf, with poor results, have been tested. recently, epag has been shown to improve platelet counts in the post-allo-hsct setting. in this study, we analysed the efficacy of epag in pts with post-transplant persistent cytopenias. methods: the population analyzed includes all pts who underwent allo-hsct from june through may in our unit. median age was years ( - ). were male ( . %) and female ( . %). baseline diseases were: aml, lpd, all, mds, mpd, mm, and bmf. donor was unrelated in ( . %) and was family in ( . %) (including haplo-identical). conditioning was ric in ( . %) and intensive in ( . %). sc source was pb in ( . %) and bm in ( . %). median followup was months ( - ). epag was initiated at some point during the first -month post-hsct period in pts ( . % of the series) due to thrombocytopenia (< /mcl) plus, at least, one other cytopenia. patients characteristics shown in table . epag was started at mg/day and escalated each weeks to , and mg/day if platelet count was < /mcl. global response was considered when, after epag, the patient needed no transfusions and reached the three of the following: platelets > /mcl, hgb > g/dl, and anc > /mcl. epag was tapered off in responders and discontinued if no response was reached after weeks. results: at epag initiation, all the pts had thrombocytopenia (< /mcl), had anemia (hgb < g/dl), and had neutropenia (anc < /mcl). counts pre and post and response to epag are shown in table . among the responders, all but one (who relapsed from thrombocytopenia and died from bleeding) were alive at analysis close ( . %). among the non-responders, three pts had gvhd-associated cytopenias, and finally died from infectious complications; the other patient relapsed from her aml, reached a new cr after treatment, and is alive and well months afterwards. epag was tapered off and discontinued in / pts who responded; / responders are still on epag. epag was discontinued in the / pts who did not respond. rest of treatment details shown in table . conclusions: ) epag worked striking well in subjects with pgf, an otherwise a life-threatening situation for patients. ) epag induced impressive responses in platelets, but strong bilinear and trilinear responses were also seen. ) epag did not improve gvhd-associated cytopenias. ) to confirm these innovative and transcendent results, we have just initiated a multicenter prospective study on the role of epag for treatment of post-hsct pgf. * five out of the six urdt were mismatched ** the donor was a woman in the six cases: three sisters and three daughters background: hepatic vod/sos with multi-organ dysfunction (mod; typically, renal or pulmonary) may be associated with > % mortality. defibrotide is approved for treating severe hepatic vod/sos post-hsct in patients aged > month in the eu, and for hepatic vod/sos with renal or pulmonary dysfunction post-hsct in the us. this analysis provides an overview of the safety results from studies of patients with vod/sos, with or without mod, who received defibrotide mg/kg/day. methods: safety data were pooled from patients with vod/sos post-hsct treated with defibrotide in a phase trial (n= ) and a phase , randomized dose-finding trial (n= receiving mg/kg/day). safety data for historical controls (hc) from the phase study (n= ) also are provided. reported separately, due to differences in patient population and data monitoring protocol, are aes from the expanded-access program (t-ind) in patients with vod/ sos with and without mod (n= post-hsct). vod/ sos was diagnosed by baltimore criteria/biopsy for the phase / studies; diagnosis by baltimore or modified seattle criteria was permitted in the t-ind. results: median patient age at hsct for the phase / studies was . years, . years for the hc, and . years for the t-ind. in the phase / studies defibrotide-treated group (n= ), ( . %) experienced aes; most common (> %) were hypotension ( . %), diarrhea ( . %), and multi-organ failure ( . %). treatment-related aes were at least possibly related to defibrotide (table) . any hemorrhage (an ae of special interest) occurred in patients ( . %); most commonly epistaxis ( . %), gastrointestinal and pulmonary alveolar hemorrhage and hematuria ( . % each), and conjunctival hemorrhage ( . %). all hc experienced an ae; most common (> %) were hypotension ( . %), tachycardia ( . %), diarrhea ( . %), nausea ( . %), and pyrexia, agitation, and petechiae ( . % each). any hemorrhage occurred in patients ( . %): most common (> %) were petechiae ( . %); hematuria, epistaxis, and pulmonary alveolar hemorrhage ( . % each); and lip hemorrhage ( . %). in the t-ind (n= ), / patients with mod ( . %) and / patients without mod ( . %) had an ae; other than vod/sos and mod, most commonly (> % in either subgroup) hypotension ( . % and . %, respectively). traes occurred in patients ( . %) ( table) . any treatment-emergent hemorrhage occurred in patients with mod ( . %) and patients without mod ( . %); most commonly (> % in either subgroup) pulmonary hemorrhage ( . % and . %, respectively) and gastrointestinal hemorrhage ( . % and . %, respectively). conclusions: the incidence and type of aes were as expected in these critically ill patients. of the pooled patients, % had aes; . % had a hemorrhage. all hcs had an ae, with . % having a hemorrhage. in the t-ind, patients with mod had higher rates of aes. support: jazz pharmaceuticals event, n(%) phase / studies (n= ) disclosure: paul g. richardson has served on advisory committees and as a consultant, and has received research funding from jazz pharmaceuticals. angela r. smith and leslie lehmann have nothing to disclose. nancy a. kernan received grants from gentium during the conduct of the study, and her research was supported by the national cancer institute of the national institutes of health under award number p ca ; the content is solely the responsibility of the author and does not necessarily represent the official views of the national institutes of health. she has a research grant from jazz pharmaceuticals. robert ryan and william tappe are employees of jazz pharmaceuticals and hold stock and/or stock options in jazz pharmaceuticals plc. stephan a. grupp has served on a steering committee and as a consultant to jazz pharmaceuticals. defibrotide for treatment of adults with hepatic vod/ sos with or without multiorgan failure after hematopoietic cell transplantation: results of a systematic review/meta-analysis background: although hematopoietic cell transplantation (hct), autologous or allogeneic, is potentially curable in various hematologic malignancies, the procedure is associated with serious and potentially life-threatening complications, among them veno-occlusive disease/sinusoidal obstructive syndrome (vod/sos) of the liver. several studies, prospective or retrospective, have reported outcomes of defibrotide, when used as prophylaxis or treatment, in a mixed population of adult and pediatric patients. in this systematic review/meta-analysis, we analyze outcomes of defibrotide when specifically used for treatment of adult patients with hepatic vod/sos with or without multiorgan failure. methods: a comprehensive search of large databases (medline/pubmed, cochrane and embase) on november , identified publications. analysis was restricted only to adult patients (defined as median age older than years) who received defibrotide for treatment of vod/sos and were reported in prospective or retrospective (which included ≥ patients) studies published in full manuscript form. there were no limitations based on language. data were extracted in relation to benefits [complete remission (cr) rate and overall survival (os)] and harms (hemorrhage, any site or organ-specific). a total of studies (prospective= ; retrospective= ) with patients met inclusion criteria. results: the median year of publication of prospective studies was ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) and for retrospective ones ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) . the prescribed starting dose of defibrotide varied among studies ranging from . mg/kg/day to mg/kg/day, mostly for a -day course. the pooled cr rate was % ( %ci= - %) for prospective and % ( %ci= - %) for retrospective studies. the pooled day + os rates were % ( %ci= - %) and % ( % ci= - %) for prospective and retrospective studies, respectively. the pooled rates of hemorrhage (any site) were % ( %ci= - %) for prospective and % ( % ci= - %) for retrospective studies. when analyzing organ-specific hemorrhage, prospective studies (n= patients) reported pooled rates of pulmonary alveolar (pa) hemorrhage of % ( %ci= - %) and of % ( %ci= - %) for gastrointestinal (gi) hemorrhage. only one retrospective study (n= patients) reported an incidence of pa hemorrhage of % ( %ci= - %) and a different study (n= patients) reported an incidence of gi hemorrhage of % ( %ci= - %). none of the studies reported cerebral hemorrhage as a complication of defibrotide therapy. conclusions: this systematic review/meta-analysis confirms the efficacy of defibrotide for treatment of vod/sos with or without multiorgan failure, yielding cr rates of - % and day + os rates of - %. the purportedly higher pooled cr and os rates observed with retrospective (vs. prospective) studies are likely due to assignment-bias inherent to observational studies. moreover, although the pooled hemorrhage (any site) rates of - % is considered proportionally significant, the pooled rates of pa and gi hemorrhage were ≤ %, in prospective studies. clinical trial registry: not applicable disclosure: m.a.k-d: consultancy for pharmacyclics m.m: received lectures honoraria and research support from jazz pharma efficacy and safety of defibrotide in the treatment of hepatic veno-occlusive disease/sinusoidal obstruction syndrome following hematopoietic stem cell transplantation: interim results from the defifrance study background: hepatic veno-occlusive disease/sinusoidal obstruction syndrome (vod/sos) is a potentially lifethreatening complication of conditioning for hematopoietic stem cell transplant (hsct) but may occur after nontransplant chemotherapy alone. vod/sos with multi-organ dysfunction (mod) may be associated with > % mortality with supportive care alone. diagnosis of vod/sos was traditionally based on baltimore or modified seattle criteria; however, the ebmt recently published separate diagnostic criteria for adults and children. defibrotide is approved for treating severe hepatic vod/sos post-hsct in patients aged > month in the eu, and for hepatic vod/sos with renal or pulmonary dysfunction post-hsct in the usa. the goal of the defifrance study, requested by the french health authorities, is to collect real-world data on safety and efficacy in a broader patient population in france, including all indications. this is the first interim analysis of the largest current evaluation of defibrotide for the treatment of vod/sos in europe. methods: defifrance is an observational, multicenter, post-marketing study that includes any patient treated with defibrotide from hsct centers in france. this interim analysis is based on all patients treated with defibrotide, including those with severe and very severe post-hsct vod/sos. vod/sos was diagnosed using traditional criteria. day+ survival, complete remission (cr; total serum bilirubin < mg/dl and resolution of mod), and safety profile are reported. results: a total of patients treated with defibrotide were included retrospectively and prospectively between july and october from table] disclosure: mohamad mohty: has received honoraria and research funding from jazz pharmaceuticals, delphine lebon: nothing to disclose, ann berceanu: none, charlotte jubert: has received funding from jazz pharmaceuticals, ibrahim yakoub-agha: has received honoraria from jazz pharmaceuticals, stéphane girault: none, marie detrait: has received research funding from jazz pharmaceuticals, cécile pochon: none, fanny rialland: none, virginie gandemer: none, jean-hugues dalle: has received honoraria from jazz pharmaceuticals, régis peffault de latour: has received research grant / honoraria / board from pfizer, novartis, alexion; research grant amgen; and honoraria from jazz pharmaceuticals, david michonneau: has received honoraria from jazz pharmaceuticals, myriam labopin: has received honoraria from jazz pharmaceuticals, floriane delaval: employee of jazz pharmaceuticals and holds stock and/or stock options in jazz pharmaceuticals plc, gerard michel: none, anne sirvent: none, laurence clement: none anne-lise menard: none, anne huynh: has received honoraria from jazz pharmaceuticals, virginie bouvatier: employee of jazz pharmaceuticals and holds stock and/or stock options in jazz pharmaceuticals plc, raj hanvesakul: employee of jazz pharmaceuticals and holds stock and/ or stock options in jazz pharmaceuticals plc, zakaria medeghri: employee of jazz pharmaceuticals and holds stock and/or stock options in jazz pharmaceuticals plc p incidence and predictors of severe cardiotoxicity in patients with severe aplastic anemia after haploidentical hematopoietic stem cell transplantation zheng-li xu , lan-ping xu , yuan-yuan zhang , yi-fei cheng , xiao-dong mo , feng-rong wang , yu-hong chen , wei han , chen-hua yan , yu-qian sun , ting-ting han , yu wang , xiao-hui zhang , xiao-jun huang peking university institute of hematology, peking university people's hospital, beijing, china background: severe cardiotoxicity after hematopoietic stem cell transplantation (hsct) is a rare but fatal complication. the aim of this study was to evaluate the frequency of severe cardiac complications and to assess the ability of various factors to predict these complications in patients with aplastic anemia after haploidentical transplantation., this is the first study evaluating the values of both clinical and imaging factors in the prediction of severe cardiotoxicity among saa patients after haploidentical transplantation. methods: a retrospective study was conducted in consecutive aplastic anemia patients who received haploidentical transplantation from to . all patients received a unified regimen including busulfan, cyclophosphamide (ctx) and antithymocyte globulin at our single center. results: a total of ( . %) patients developed grade iii or iv cardiac toxicity. patients with cardiotoxicity had significantly poorer overall survival (os) than those without cardiotoxicity ( . % vs. . %, p< . ). our multivariable model identified four independent adverse predictors of severe cardiotoxicity, including pre-transplant ecog score (≥ ), abnormal st-t wave on -lead electrocardiogram (ecg), hyperlipemia and recalculated ctx dose (≥ . g/m /d). a predictive risk model was refined as low risk ( - factor), intermediate risk ( factors) and high risk ( - factors) . the respective incidences of severe cardiotoxicity were . %, . %, and . % in the high-, intermediate-and low-risk groups (p< . ). the corresponding os rates were . %, . %, and . % in the three groups (p< . ) at the last follow-up. conclusions: patients with high risk scores had the poorest outcomes and should be monitored closely. a reduced intensity conditioning might be recommended for these patients. disclosure: there are no conflicts of interest to declare. background: allogeneic stem-cell transplantation (allo-sct) is associated with significant transplant-related mortality (trm). acute renal failure (arf) is a frequent complication and usually presents early after the procedure, compromising its feasibility. the aim of this study is to analyse the incidence of arf, its risk factors and its potential impact on trm after allo-sct. methods: patients were included ( males [ %]; median age years, range - ) treated with allo-sct consecutively between january and april in a single institution. patient characteristics are detailed in table . median follow-up was . years (range, . - . ). renal function was evaluated using creatinine and data was collected pre-transplant (baseline) and at the point when arf was developed after allo-sct. arf was evaluated using akin criteria, being akin- an increase . -to . -fold from baseline, akin- an increase . -to . -fold and akin- an increase ≥ -fold. chronic renal disease was evaluated one year after the date of arf using kdigo criteria. results: cumulative incidence of arf at year was % (akin- , %; akin- , %; akin- , %). in the multivariate analysis, arf (akin- / ) was associated with: non-use of antithymocyte globulin in conditioning chemotherapy, p= . (hr . , . to . ) and development of severe agvhd, p= . (hr= . , to . ). in patients with arf akin- , the most important variables in the multivariate analysis were: use of methotrexate (mtx) plus cyclosporine vs mycophenolate mofetil plus cyclosporine as gvhd prophylaxis, p= . (hr= . , . to . ); myeloablative conditioning vs reduced intensity, p= . (hr= . , to . ) and use of total irradiation therapy in conditioning, p= . (hr= . , . to . ). trm at year increased significantly according to akin: akin- , %; akin , %; akin , %; p= , ; hr= . . overall survival at years according to akin was: akin , %, akin , % and akin , %; p= , (figure ). the incidence of chronic renal disease at year after allo-sct according to arf was: no arf ( %), akin- ( %), akin- ( %) and akin- ( %); p= . . conclusions: arf is a frequent complication during the first year after allo-sct and is associated with several factors. arf akin- was associated with more intensive strategies received during conditioning, meanwhile akin- / were related to development of gvhd. there is an association of arf (akin- , or ) with development of chronic renal disease. background: the introduction of cellular therapies such as car-t and modalities of gvhd-prophylaxis with posttransplant/cyclophosphamide (ptcy) that increase the number of admission days have boosted the pressure of available beds in the bm-units. in this sense, our centre started an at-home allogeneic stem cell transplantation (allo-sct) program to follow aplasia from the d+ until independent ambulatory patient. to evaluate the feasibility and safety of allosct, we compared two groups: allohsct/athome (ah-group) vs. allohsct/in-patient (ip-group). methods: we included patients receiving allosct (january -november ) in a single centre: patients, ah-group and , ip-group. all patients received conditioning at the hospital. gvhd-prophylaxis consisted in tacrolimus (tk) plus mycophenolate (mpm) or methotrexate, or ptcy (d+ , d+ ) plus tk (d+ ). all patients received prophylaxis with levofloxacin, fluconazole and acyclovir. besides that, ah-group patients received prophylaxis with ceftriaxone g/ h iv or ertapenem g/ h iv, and aspergillus-prophylaxis with inhaled liposomal amphotericin-b or posaconazole during neutropenia. patients of ah-group since d+ or d+ (in ptcyprophylaxis) received a nurse visit at-home once daily. the visits by the physician were performed at the hospital and only during complication events. first-line therapy of neutropenic fever was meropenem g/ h in both groups, using a portable infusion pump in ah-group. in this group, the absence of focal infection or signs of severe sepsis allowed returning home after the initiation of antibiotics. the platelets support was performed at-home and the red blood support at hospital. results: the median (range) age (years) of the series was . the median follow-up of the series has been not achieved. the source of the sct was peripheral blood in all cases. we didn't find statistical differences between two groups (ah vs ip) in terms of age, diagnosis, type of donor, intensity of conditioning, gvhd-prophylaxis, toxicity (mucositis, acute renal injury, neutropenia and thrombocytopenia), agvhd, aspergilosis and trm. interestingly, a significant reduction of neutropenic fever was observed resulting the lower use of meropenem in the ah-group than ip-group. the admission median days were similar in the both groups and it represented - days the reduction in the total economic cost of the ah-group. the whole analysis of the results are detailed in table: in-patient group, conclusions: in our experience, at home allosct, including ptcy-gvhd prophylaxis, is a feasible and safe procedure reflected in similar trm and aspergillosis incidence. at-home allo-sct is associated with a significant lower risk of neutropenic fever than in-patient group, as well as a very low readmission rate. disclosure: gonzalo gutiérrez-garcía: honoraria from gilead. grant from jazz pharmaceutical and janssen. laura rosiñol: honoraria from takeda, janssen, amgen and celgene. the others author do not have any disclosures to declare. background: renal complications in sickle cell disease (scd) include episodes of acute kidney injury (aki), progressive chronic kidney disease (ckd) and hyperfiltration, defined by abnormally high glomerular filtration rates (gfrs). hematopoietic stem cell transplant (hsct) from an hla identical sibling donor is a well-established curative treatment for scd, but traditional myeloablative conditioning (mac) regimens pose risks of kidney injury due to intensive use of chemotherapeutic agents, infectious risks, and use of calcineurin inhibitors (cnis). aki and subsequent fluid overload (fo) are common in pediatric hsct with reported aki incidence of %- % (kyung-nam koh et. al., ). we report renal outcomes in pediatric patients with scd who received hsct following a non-myeloablative conditioning (nma) regimen without cni exposure. methods: retrospective chart review describing renal outcomes in pediatric patients ( years of age or younger) with scd (hbss) who underwent nma hsct in alberta, canada from july to february . the nma regimen is illustrated in figure . reported renal outcomes: ) measured gfr (dtpa) pre-hsct, ) aki (kdigo definition) post-hsct by reviewing all serum creatinine levels from pre-hsct to one month post-hsct, ) %fo calculated: (max post hsct weight -baseline weight)/ baseline weight x for the two first weeks post-hsct, and ) estimated gfr (egfr) using the pediatric schwartz formula at last follow-up post-hsct, ckd defined as egfr < ml/min/ . m , mildly reduced gfr: - ml/min/ . m , and hyperfiltration: gfr ≥ ml/ min/ . m . [[p image] . results: eighteen patients ( % male, - years old at transplant) were included. most common pre-morbid events: vaso-occlusive crisis (n= ), acute chest syndrome (n= ), splenic sequestration (n= ), and cholelithiasis (n= ). median follow-up time: months (range: - months). all patients engrafted successfully with no acute or chronic gvhd. baseline measured gfrs were all > ml/min/ . m (range: - ) with mildly reduced gfr and hyperfiltration seen in one ( . %) and ( . %) patients respectively. at baseline (pre-hsct), the only aki event was one transplant related aki secondary to delayed hemolytic reaction after exchange transfusion in preparation for transplant. post-hsct, there were no aki events. additionally, no substantial %fo post-hsct was observed. average %fo week one post-hsct: + . % (range: - . % -+ . %) and week two post-hsct: + . % (range: - . % -+ . %). post-hsct egfr remained > ml/min/ . m at last follow-up in all patients. hyperfiltration was present in ( . %) of the patients. conclusions: this is the first study describing stable kidney function in children with scd after the present nma hsct regimen with alemtuzumab/ cgy total body irradiation (tbi) with prolonged post-hsct sirolimus. no episodes of aki or significant fluid overload were observed during the first month post-hsct, and no patient developed ckd during follow-up. further prospective studies are needed to confirm our findings and to determine if stable renal function persists during longer-term followup. disclosure: nothing to declare. lung microbiota in patients with idiopathic pneumonia syndrome (ips) after hct background: idiopathic pneumonia syndrome (ips) is a non-infectious pulmonary complication after hematopoietic cell transplantation (hct) and the etiology remains unknown. recent studies have reported that various diseases are associated with changes of microbiota. the aim of this study was to evaluate the lung microbiota in hct recipients with ips and identify microorganisms potentially associated with ips. methods: frozen bronchoalveolar lavage (bal) samples from hct recipients with ips (n= ) and research bal samples from asymptomatic hct recipients as controls (n= ) were retrospectively analyzed. all samples were negative for common viruses by quantitative pcr. sequencing libraries were made with ng of input dna per sample (nextera xt, illumina). samples were pooled and sequenced by hiseq to obtain -bp paired end data. sequence data analysis and read classification were performed with sunbeam and the quality control and read classification were performed using komplexity and kraken, which classifies bacterial, archeal, and viral genomes. we used sequence data of bronchoscope prewashes from a separate cohort as controls for environmental sources (n= ). bray-curtiss dissimilarity among samples was calculated using the vegan r packages. permanova and a two-sided wilcoxon rank sum test were used to compare between the study groups. results: bal samples started at a median of x raw read pairs per sample and reduced to x reads assignable to microbial taxa following quality control. the bacterial phyla proteobacteria and firmicutes were most abundant followed by bacteroidetes and actinobacteria in both bal and bronchoscope prewash samples. separation of bal and prewash microbiota using bray-curtiss dissimilarity plots showed that bal samples were distinguished by sequences assigned to staphylococcus, acidovorax, and bradyrhizobium species, while prewash samples were distinguished mostly by pseudomonas and elizabethkingia species, consistent with environmental sources (figure) . within bal samples, staphylococcus species were the main drivers of separation between ips cases and the controls (p= . , permanova, figure) . consistent with this, a linear discriminant analysis to identify taxa best distinguishing cases and controls identified staphylococcus, especially s. epidermidis, in ips cases with lactobacillus and streptococcus species in controls. we then compared relative abundances of s. epidermidis between all study groups. ips case samples were significantly enriched in s. epidermidis compared to control (p< . , two-sided wilcoxon rank sum test) and prewash samples (p< . ). viruses were classified by category as human pathogens, non-human pathogens, and bacteriophages. torque teno viruses (ttv) was the most commonly detected virus among viruses that replicate on human cells, and there was a trend towards higher abundance in ips case samples than controls. conclusions: lung microbial sequences in hct recipients predominantly consisted of proteobacteria and firmicutes, and had considerable overlap with environmental background. patients with ips had significantly more staphylococcus sequences detected than asymptomatic hct patients. these results suggest that patients with acute lung injury post-hct show distinct patterns of lung microbiota, although heterogeneity of sample collection and processing cannot be excluded and no singular organism was uniquely associated with ips. a prospective study is required to confirm these findings and define the clinical significance of differences in abundance patterns. disclosure: nothing to declare p abstract withdrawn. romiplostim for the treatment of thrombocytopenia after allogeneic stem cell transplantation background: thrombocytopenia is a common complication after allogeneic stem cell transplantation (allo-hct). with variable possible causes, such as drug side effects, infections, poor graft function, graft vs host disease (gvhd) and immune mediated. the purpose of this study was to evaluate the efficacy of romiplostim, a thrombopoietin receptor agonist, in patients with prolonged thrombocytopenia with no obvious cause after allogeneic transplantation. methods: retrospective analysis of allo-hct patients who received romiplostim at a single bmt unit between november and november . romiplostim was given because of prolonged (> weeks) thrombocytopenia (< , μl) that couldn't be explained by obvious causes such as administration of drugs (antibiotics/antivirals), infection or gvhd. all patients were in complete remission and had complete chimerism. response to romiplostim treatment was considered transfusion independence or plt> . /μl. results: in total, patients (median years, - ) received romiplostim. patients ( male, females) had aml ( pts), all ( ), mds ( ) or hodgkin ( ), received a myeloblative (busiphex-based: , tbi-based: ) or ric ( ) conditioning and were transplanted from a sibling ( ), vud ( ) or haploidentical ( ) donor with pbsc ( ) or bm ( ) . all patients revealed primary neutrophil (median days, range - ) and > . /μl platelet ( days, - ) engraftment. romiplostim was started at median day + (range - ) with a median dose μg/kg ( ) ( ) ( ) ( ) ( ) . the median platelet count before commencement of treatment with romiplostim was . /μl (range . - . ) and them ( %) were transfusion-dependent. in total / ( %) patients responded to romiplostim treatment. eight out of the ( %) transfusion dependent patients responded to the administration of romiplostim. six out of the patients ( %) who were transfusion independent at romiplostin initiation (plt median . /μl, range . - . ) responded. the median duration of treatment was days ( - ) and the median follow up from the commencement of romiplostim was days ( - ). three out of ( %) patients experienced relapse of thrombocytopenia after discontinuation of romiplostim and re-initiation of romiplostim was commenced in all of them, of which responded and didn't. the administration of romiplostim was done on an external basis and was well tolerated by the patients. two patients experienced gvhd during romiplostim treatment (both patients transplanted from / unrelated donor, and days after initiation treatment with romiplostim). / patients interrupted romiplostim due to disease relapse. / patients receiving romiplostim are alive in complete remission and died ( due to relapse, and due to trm). conclusions: we present high response rates to romiplostim in patients with prolonged thrombocytopenia after allogeneic transplantation. in this retrospective study there were no side effects from the administration of romiplostim. however, the administration of romiplostim after allo-hct should be controlled in prospective trials. disclosure we report a single-center analysis of adult patients (median age years, range - , m/f / ), receiving tpo agonists for isolated severe thrombocytopenia (n= ) and spgf (n= ) after allo-hsct. primary diagnoses were aml ( ), all ( ), mds ( ), pmf ( ), mds/mpn ( ), saa ( ), cml ( ), nhl ( ) . severe pgf was defined as cytopenia in ≥ lineages (platelet < × /l, anc < . × /l, hemoglobin < g/l any time after sustained engraftment), full or stable mixed donor chimerism > % and no signs of relapse. median dose of romiplostim was (range, - ) mcg/kg weekly, eltrombopag - (range, - ) mg/day. overall response (or) included cr (platelet ≥ × /l, anc ≥ . × /l, and hemoglobin ≥ g/l) and pr (platelet > × /l, anc ≥ , × /l, hemoglobin > g/l). results: median time from pgf diagnosis to treatment with tpo agonists was days ( - ), median treatment duration was weeks ( - ). tpo agonists were well tolerated with no cases of grade iii-iv toxicity. tpo agonists were combined with rituximab (n= ), rituximab and dli (n= ) and hsc boost (n= ) in ( %) patients. a total of ( %) patients met criteria of response (cr: n= , %; pr: n= , %). combination therapy showed no difference in or compared to tpo agonists alone. or was not depended on the tpo agonist used nor the time to therapy initiation. median increase in anc in responders was . × /l ( . - . ), in platelet count - × /l ( - ). a total of patients died due to relapse (n= ), gvhd iii-iv grade (n= ) and infection (n= ). two-year os from the start of tpo agonist therapy was % ( % ci, - ) with a significant difference between responders and non-responders: % ( % ci, - ) vs. % ( % ci, - ) (p= , ). conclusions: this study showed promising results of tpo agonists for management of spgf. further studies are warranted to specify optimal timing and dosing regimen, predictors of response. [[p image] . two-year os in responders and nonresponders to tpo agonist therapy] disclosure: there are conflicts of interest to disclose p cytomegalovirus reactivation kinetics and peak titers as novel predictors of survival and relapse after allogeneic cell transplantation for hematologic malignancies saskia leserer , evren bayraktar , nikolaos tsachakis-mück , michael koldehoff , lara kasperidus , esteban arrieta-bolanos , mirko trilling , katharina fleischhauer , dietrich w. beelen , amin t. turki university hospital essen, essen, germany, background: after allogeneic hematopoietic cell transplantation (hct), human cytomegalovirus (cmv) reactivation associates with non-relapse mortality (nrm) but also with reduced relapse in patients with leukemia, as shown by numerous studies that evaluated cmv reactivation as a qualitative yes/no parameter in the first months posttransplant. we hypothesized that longitudinal quantitative assessment of cmv reactivation kinetics and virus loads might improve patient-specific clinical outcome associations. methods: this retrospective study included patients with hct for hematologic malignancies treated between / and / at university hospital essen, germany. cmv titers were monitored weekly by quantitative pcr (qpcr); cmv reactivation was defined by a cutoff of > genome copies per ml. patients were included for analysis, if at least measurements were available during the first days after hct. in total, , samples were analyzed. subgroup analyses were performed according to the time of cmv reactivation (before/after + d) or the cmv viremia titer (> , , , - , and - , copies/ml). results: cmv reactivation was detected in (median age years; range - years) out of patients. baseline characteristics (age, gender, underlying disease, transplant) of patients without cmv reactivation were comparable. cmv reactivation kinetics followed a gaussian normal distribution with a median first reactivation at + d and peak titers at + d. all except patient reactivated before d, % before + d. overall survival (os) of the cmv reactivation group as a whole did not significantly differ from the non-reactivation group ( vs. months). however, in subgroup analyses os was significantly reduced in patients with very early (< + d) compared to later reactivation ( vs. months, p= . ). moreover and importantly, os was significantly reduced in patients with cmv reactivation at high titers of > , copies/ml compared to those with lower titers (( vs. months) p< . ).cox regression analyses confirmed significantly reduced os for patients with cmv reactivation > , copies/ml and < day + as compared to the other cohorts (hr . , %ci . - . , p< . and hr . , % ci . - . , p= . ) respectively). the nrm was consistently higher (hr . ; %ci, . - . , p< . ) for patients with cmv copies > , /ml. the risk of hematologic relapse was exclusively reduced in patients with a peak cmv viremia between , and , copies/ml (hr . , % ci . - . ; p= . ) as compared to patients without cmv reactivation. for other levels of cmv reactivation this effect was not observed. conclusions: our data showed that cmv reactivations before + d or with high titers of > , copies/ml associated with significantly reduced os, while cmv reactivations at intermediate titers between , and , copies/ml had a positive impact on relapse incidence. these findings underline the complexity of cmv reactivations after hct outcome, and support longitudinal evaluation of cmv titers and individualized quantitative kinetics models for risk assessment after hct to distinguish the advantageous from the detrimental aspects of cmv reactivation. disclosure: att has received lecture fees from jazz pharmaceuticals and travel subsidies from neovii biotech outside the submitted work. the other authors declare no competing financial interests within the submitted work. association of serum ferritin levels before start of conditioning with mortality after allosct -a prospective, non-interventional study of the ebmt transplant complication working party background: elevated serum ferritin levels occur due to iron overload or during inflammation and macrophage activation. a correlation of high serum ferritin levels with increased mortality after allosct has been suggested by several retrospective analyses as well as by two smaller prospective studies. methods: this international multicentric study aimed to study the association of ferritin serum levels before start of conditioning with allosct outcome. patients with acute leukemia, lymphoma or mds receiving a matched sibling allosct for the first time were considered for inclusion, regardless of conditioning. data were prospectively collected between / and / . a comparison of outcomes between patients with high and low ferritin level was performed using univariate analysis and multivariate analysis using cause-specific cox model. variables included in the multivariate analyses were age, sex mismatch, diagnosis, disease status, karnofsky score, number of cd cells given, intensity of conditioning, type of gvhd prophylaxis, atg use, time from diagnosis to transplant, year of transplant and cmv status. results: twenty centers from european countries reported data on allosct recipients. patient characteristics are given in table . the ferritin cut off point was determined at μg/l (median of measured ferritin levels). overall survival of allosct recipients with ferritin levels above cut off measured before start of conditioning was significantly shorter ( figure a , univariate hr= . ci= . - . p= . ; multivariate hr= . , ci= . - . , p= . ). progression-free survival was also shorter ( figure b , univariate hr= . ci= . - . p= . ; multivariate hr= . , ci= . - . , p< . ). excess mortality in the high ferritin group was due to both higher relapse incidence (univariate hr= . ci= - . p= . ; multivariate hr= . , ci= . - . , p= . ) and increased non-relapse mortality (univariate hr= . ci= . - . p= . ; multivariate hr= . , ci= . - . , p= . ). non-relapse mortality was driven by significantly higher infection-related mortality in the high ferritin group (univariate hr= . ci= . - . p = . ; multivariate hr = . , ci = . - . p = . ). acute and chronic gvhd incidence or severity were not associated to serum ferritin levels. conclusions: ferritin levels before start of conditioning can serve as routine laboratory biomarker to predict mortality after allosct. disclosure: the authors declare no confict of interest related to this study p prediction of reduced lung function and acute gvhd by surfactant protein d in allogeneic stem cell transplantation transplantation (hsct) and may progress to bronchiolitis obliterans that has a high mortality rate. surfactant protein d (sp-d) is an innate defense molecule involved in immune regulation at the epithelial surfaces, particularly in the lungs, and elevated levels have been associated with exacerbation of chronic obstructive pulmonary disease (copd). the aim of this study was to investigate, whether sp-d plasma levels and variants in the gene encoding sp-d may predict the development of reduced lung function after allogenic hsct. methods: we performed a population-based, singlecenter study of children (aged - years) treated with allogeneic hsct. the study consisted of ) a prospective study of serial plasma sp-d levels and rs genotypes in patients during the first months after hsct, and ) a retrospective study of rs genotypes within the sp-d gene in patients transplanted between - . pulmonary function tests were performed regularly as part of the clinical monitoring. results: at the day of graft infusion (day ) sp-d levels were reduced compared to levels before start of treatment with conditioning chemotherapy, defined as baseline ( ng/ml (quartiles - ) at day vs ng/ml ( - ) at baseline, p< . ). from day + sp-d levels increased and remained increased during the whole study period ( ng/ml ( - ) at baseline vs ng/ml ( - ) at months, p< . ). acute gvhd (agvhd) occurred in patients, of those patients with grade - . high sp-d levels at day + were associated with the development of agvhd ( ng/ml ( - ) vs ng/ml ( - ), p< . ) ( fig. ). the c/c genotype was associated with generally low sp-d levels and low fev /fvc at all time intervals compared to the other genotypes, significantly - months post-hsct (p= . ). there was no overall correlation between sp-d levels and lung function, but stratifying for genotype, high baseline sp-d levels were predictive for reduced fev /fvc at - months in cc and tt homozygous individuals. conclusions: patients with a genotype causing low capacity for sp-d production are at increased risk of developing pulmonary impairment after hsct. in addition, our data lend support to other studies indicating that spd production may increase during inflammatory pulmonary disease, acting as a reactive, protective mechanism. further research is warranted to define the role of sp-d levels and genotypes as a prognostic tool for lung function and agvhd. [[p image] . background: allogeneic hematopoietic cell transplantation (allohct) means a long period of restricted mobility and a range of therapy related side effects on muscle function. in this context patients demonstrated a huge decline of physical capacity and muscle mass in particular, accompanied with a decrease of quality of life (qol). resistance training could maintain muscle mass but is limited by patientsb lood values (platelet-count) and well-being. whole body vibration (wbv) was shown to maintain muscle mass during bed rest and has less impact on blood pressure than conventional resistance exercises. furthermore it was also shown to be feasible in patients during high dose chemotherapy. therefore the aim of our study was to examine the effects of wbv during allohct on patients physical and functional performance as well as qol. methods: patients receiving allohct were randomly allocated to either a wbv exercise group (ig) or an active control group (cg) doing stretching and mobilization. both groups exercised during the whole time of hospitalization for times per week and underwent pre-, post-and followup-assessment. physical capacity was determined by maximum oxygen consumption (vo peak ) and maximum power (p max ) during cardiorespiratory exercise test and by maximum strength of the knee extensors and flexors (ex max , flex max ) during isokinetic strength test. functional performance was assessed by jumping height during counter movement jump (cmj) and time of chair rising test (crt) as well as power output during both tests. qol was assessed by questionnaires of the eortc. results: during allohsct vo peak and p max decreased in both groups but till follow-up an increase is seen in the ig (p= . ; p= . ). at day + /follow-up a vo peak group difference is seen (p= . ). ex max (p= . ) and flex max (p= . ) were only reduced in the cg during hospitalization. jumping height and power output decreased in the cg during hospitalization (p= . , p= . ) and a difference between groups were seen in changes of jumping height from pre-to follow-up-assessment (p= . ): increase in the ig and decrease in the cg. the ig showed a decrease in time from baseline to follow-up (p= . ) in the crt and an increase of power output (p= . ). qol decreased only in the cg during hospitalization (p= . ) while during follow-up qol increased in both groups (ig: p= . ; cg: p= . ). in the cg physical functioning decreased during intervention (p= . ) whereas an increase was seen in the ig from pre-to follow-upassessment (p= . ). body image was significant worse in the cg compared to the ig at hospital discharge (p= . ) as well as at follow-up measurement (p= . ) where it got worse over time (p= . ). conclusions: wbv was shown to maintain maximum strength, jumping performance and qol during allohct. although cardiorespiratory fitness could not be maintained by wbv during hospitalization, it seems in the follow up period till day + that recovery of the cardiorespiratory system is enhanced by wbv carried out during allohst. nevertheless reasons for this changes in recovery have to be analyzed in further studies as well as treatment effects of wbv compared to conventional resistance training. disclosure: supported by a grant of the faculty of medicine and comprehensive cancer center freiburg respiratory virus infection within year after of allo-sct is the significant risk factor of obstructive ventilatory disturbance kosei kageyama , michiho ebihara , mitsuhiro yuasa , daisuke kaji , aya nishida , shinsuke takagi , hisashi yamamoto , go yamamoto , yuki asano-mori , naoyuki uchida , atsushi wake , akiko yoneyama , shigeyoshi makino , shuichi taniguchi toranomon hospital, hematology, tokyo, japan, background: obstructive ventilatory disturbance (ovd) is one of the major life-threading complication at the chronic phase of allogeneic stem cell transplantation (allo-sct). bronchiolitis obliterans has been the most established etiology as a part of chronic graft-versus-host disease and major cause of late non-relapse mortality of allo-sct. but other etiologies impact on respiratory function after allo-sct and risk factor of ovd have not been well understood. methods: to address these issues, we retrospectively reviewed the medical record of consecutive patients who first allo-sct at toranomon hospital between and . to detect ovd, forced expiratory volume in second (fev . ) showed less than % of predicted in spirometry test was defined as positive. in the recipients who showed fev . less than % in pre-transplant test, more than % reduction of fev . was regarded as positive. nasopharyngeal swab of those who had upper respiratory tract symptoms were tested for the presence of respiratory viral antigens (adv, piv, and rsv). patients with ecog performance status of , had active infection at transplant were excluded from this analysis. the cases of early death or relapse before days post-transplant, and the cases of graft failure were also excluded. results: the median age was years (range, - ). underlying diseases were aml in , mds/mpd in , cml in , all in , atl in , hl in , nhl in , and others in . five hundred twenty-nine ( %) were not in remission at the time of transplant. five hundred eightythree patients ( %) were conditioned with myeloablative regimens, whereas patients received reduced-intensity regimens. donor sources consisted of related peripheral blood /bone marrow (bm) (n= ), unrelated bm ( ) forty-six developed ovd on median of ( - ) days post-transplant. cumulative incidence of ovd was . % in total population. in recipients those who could spirometry, overall survival at years was . % in patients who developed ovd and was comparable with those who did not develop it ( . %, p= . ). in univariate analysis, disease status (cr/aa or noncr), recipient age (age< or ≥ ), prior autologous stem cell transplantation (yes or no), intensity of conditioning regimen (mac or ric), tbi dose (< gy or ≥ gy), busulfan dose (< . mg/kg or ≥ . mg/kg), donor source (cord blood or non-cord) had no impact on the incidence of ovd. patients who developed respiratory virus infection showed significantly higher incidence of ovd compared to those who did not developed it ( . % vs . %, p< . ). in multivariate analysis, respiratory virus infection was the only significant risk factor for the development of ovd (hr= . , % ci . - . , p< . ). conclusions: respiratory virus infection within year after allo-sct is the significant risk factor of ovd. disclosure: nothing to declare. background: metabolic syndrome (mets) is related to increased risk of cardiovascular disease and type- diabetes (dm- ) and usually seen in overweight individuals in the general population. we investigated mets and clinical risk factors two decades after hsct. methods: all male survivors treated with myeloablative allo-hsct during childhood (< years) between - in denmark were invited to a follow-up study. mets was defined as the presence of at least three ncep atp iii criteria: fasting plasma triglyceride (tg) ≥ . mmol/l, high density lipoprotein (hdl) < . mmol/l or medical treatment of hyperlipidemia; fasting plasma glucose (fpg) ≥ . mmol/l; abdominal circumference (ac) > cm; bp ≥ mmhg (systolic) / ≥ mmhg (diastolic) or medical treatment for hypertension. patients with overt dm- were included into the mets group. furthermore, patients were examined for chronic graft-versus-host disease (cgvhd) by the nih-criteria at the time of follow-up and high sensitivity c-reactive protein (hscrp) was measured. the prevalence of mets was compared to a nordic reference group (hildrum et al. ) . results: we included out of eligible males (participation rate %) aged - years, median years. median (range) follow-up was ( - ) years. of these males, % had a malignant diagnosis and % were treated with tbi-based conditioning. donors were matched siblings (n= ), matched relatives (n= ) or matched unrelated donors (n= ). mets was more prevalent ( %) in the young adult survivors compared to the prevalence reported for - year-olds in the nordic reference ( %). instead the prevalence was comparable to that reported for the - year-olds ( %). of the components of mets, elevated tg ( %), hypertension ( %), and decreased hdl ( %) were frequent, while fpg was elevated in %. importantly, only % of those with mets had increased ac and mean bmi ( . kg/m ) of the hsct survivors was within normal range in contrast to features of mets observed in the background population. having mets was significantly associated with tbi (rr = . , %ci ( . - . ), p= . ) as was the following single components of mets (mean in tbi group vs. mean in non-tbi group): elevated tg ( . mmol/l vs. . mmol/ l, p= . ), lower hdl ( . mmol/l vs. . mmol/l, p= . ) and higher diastolic bp ( mmhg vs. mmhg, p= . ). mets was only demonstrated in one patient who received non-tbi based conditioning. sixteen of patients had cgvhd of which nine were moderate to severe cases, but cgvhd was not associated with mets. however, low-grade inflammation measured by hscrp was related to increased ac (rho= . , p= . ) and tg (rho= . , p= . ). conclusions: our results indicate that male long-term survivors of allo-hsct during childhood have a high risk of mets at an earlier age than the general population. the presence of mets despite normal bmi in several patients suggests unconventional etiologies like the effect of tbi and low-grade inflammation. disclosure: nothing to declare. results: this survey was completed by transplant directors ( %), transplant consultants ( %), nonconsultant grade physicians ( %), hsct clinical nurses specialists (cns) ( %) and other ( %) from centres in countries. % of the centres are adult-only, % paediatric-only and % treat adult and paediatric patients (mixed centres). % are located higher than degrees latitude (northern countries) and % lower than this latitude (southern countries). at the time of the survey % were members of the european union (eu). measurement of serum vd is routinely performed in % of the centres prior and in % after allogeneic hsct. the main clinical indications are known osteopaenia/osteoporosis ( %), previous fracture ( %), treatment with steroids ( %), premature menopause ( %) and established menopause ( %). monitoring occurs every months ( %), every months ( %), once a year ( %) or at other time-points ( %). in this regard, seasonality is not taken into account in the majority of the centres ( %). local and national/international guidelines (nice) are only followed by % and % of the centres, respectively. the most common cut-off value of serum vd for commencing on replacement is nmol/l ( %). northern countries tend to use values of ≥ nmol/l whereas southern countries ≤ nmol/l. % do not use cut-off values. following hsct, % of centres prescribe vd supplements to maintain calcium metabolism and bone health ( %), enhance immune reconstitution post-hsct ( %), gvhd prevention ( %), enhance immune-suppression to treat gvhd ( %), treat depression/fatigue ( %) and reduce relapse risk %. a "loading" dose is administrated in % ( % adult, % mixed and % paediatric), with a mean duration of weeks . the median daily loading dose is , iu ( - , ). the median "maintenance" daily dose is iu ( - , ). there are not remarkable differences between adult and paediatric centres or northern and southern countries. vd replacement is prescribed by transplant physicians ( %), family physicians ( %), endocrinologists ( %), cns ( %), others ( %) and in % of the centres, patients are advised to buy it over-the-counter. vd is prescribed combined with calcium carbonate in % and alone in % of the centres. it is eventually discontinued by % of the centres when therapeutical levels of vd are reached ( %), dexa scan returns to normal ( %) and symptomatic improvement ( %). conclusions: this survey has demonstrated discrepancies in monitoring and replacement of vd across ebmt allogeneic hsct programmes. although awareness has arisen over the last decade, there is still lack of evidence about the optimal levels of vd required for immunemodulation post-hsct. this survey emphasises the need for specific guidelines to harmonise the current management of vd deficiency in adult and paediatric hsct setting. disclosure background: the use of unmanipulated haploidentical sct (haplo-sct) with post-transplant cyclophosphamide (pt-cy) as gvhd prophylaxis has widely extended. primary and secondary graft failure are relatively uncommon complications. however, poor graft function (pgf) after haplo-sct with pt-cy has not been described thoroughly. the objective of this study is to describe characteristics, treatments and outcomes of patients with pgf after haplo-sct with pt-cy. methods: we retrospectively analyzed haplo-sct with pt-cy consecutively performed between and in our centre. pgf was defined as either occurring after initial engraftment: persistent neutropenia (anc < /ul) with the need of at least doses of g-csf and/or thrombocytopenia (platelets < . /ul) with platelet transfusion dependence, with complete donor chimerism and without concurrent severe gvhd or disease relapse. results: nineteen patients were excluded from the analysis due to early mortality (death before day + ), primary graft failure (absence of neutrophil engraftment by day + , with mixed chimerism) or secondary graft failure (development of severe cytopenias and mixed chimerism after initial achievement of neutrophil engraftment). thirty one patients ( , %) were diagnosed with pgf. main characteristics of these patients are summarized in table . twenty six patients ( %) presented with neutropenia and were treated with g-csf, while patients ( %) only developed severe thrombocytopenia without neutropenia, and were treated only with platelet transfusion. twenty four patients ( , %) had at least cmv reactivation, patients ( %) had or more cmv reactivations and patients ( %) received valganciclovir for cmv reactivation treatment. although most patients achieved adequate peripheral blood counts (pbc) with initial salvage therapy, patients ( %) had persistent cytopenias in spite of g-csf, platelet transfusion, cmv reactivation resolution and myelotoxic drugs withdrawal. four of them were treated with a boost of cd + selected peripheral blood donor cells at a median of days after . median cd + cells infused was , x /kg. these patients achieved adequate pbc after salvage therapy and two developed gvhd. the other patients were treated with increasing doses of thrombopoietin (tpo) receptor agonist (tra) eltrombopag. one patient started treatment days after hsct with mg daily and increased dose to mg daily, with complete recovery of pbc months after initiating tra. the second patient started treatment days after hsct with mg daily and increased dose to mg daily, with complete recovery of pbc months after initiating tra. twenty one patients ( %) with pgf diagnosis had long term survival. conclusions: poor graft function is a frequent complication after haplo-sct with cy-post. cmv reactivation and myelotoxic drugs could be the most relevant factors associated with development of this entity. although most patients recover pbc without specific therapies beyond g-csf and platelets transfusion, there is a small group of patients with persistent cytopenias. boost of cd + selected cells is effective in reverting this condition, with gvhd as main complication of this procedure. use of tra seems to be an interesting option for these patients, although more experience is needed to draw definitive conclusions. disclosure: nothing to declare. were also frequently observed. the high risk patients for anxiety (hads-a score ≥ ) and depression (hads-d score ≥ ) was found in . % and . %, respectively. . % of patients was in high distress status (nccn dt score ≥ ). we found that younger age (< years) was significantly associated with poor quality of life score (fact-bmt) (p= . ) and high risk of fatigue (p= . ), anxiety (hads-a) (p= . ), and depression (hads-d) (p= . ). female sex was significantly related to lower physical well-being score and higher distress score (p= . and p= . , respectively). acute lymphoblast leukemia (all) survivors after allo-hct showed significantly worse quality of life score (fact-bmt) (p= . ) and higher depression score (hads-d) (p= . ) compared to those with other disease. chronic graft versus host disease (gvhd) and continuous immunosuppressant usage also have significant adverse impact on lower fact-bmt score (p= . and p= . , respectively) and higher hads-d score (p= . and p= . , respectively). but there was no significant difference in fact bmt, hads-a, hads-d, nccn dt according to donor type, conditioning intensity, anti-thymocyte globulin use, acute gvhd. smoking and alcohol drinking was continued in . % and . % of allo-hct survivors. . % of survivors did not exercise regularly. regular health screening tests have been done only in patients ( . %). conclusions: allo-hct survivors over years following allo-hct still have many physical and psychological symptoms. younger patients (< years), female, all, chronic gvhd, and sustained use of immunosuppressant were significant risk factors for poor quality of life and anxiety. we need to build more active survivorship care plan after allo-hct especially for those patients. disclosure: all authors have nothing to declare. evaluation of the new ebmt criteria for the diagnosis of vod/sos in consecutive transplant patients using an electronic patient record analysis system asha aggarwal , nicola gray , oliver lomas , katalin balassa , nadjoua maouche , robert danby , , andy peniket , grant vallance background: veno-occlusive disease (vod), or sinusoidal obstruction syndrome (sos), is a recognised complication of haematopoietic stem cell transplantation. hepatic vasculature endothelial cells are damaged by conditioning chemotherapy, leading to venous occlusion and centrilobar necrosis. the ebmt criteria for diagnosis of vod are bilirubin >= with two of painful hepatomegaly, > % weight gain and ascites. vod is often under-diagnosed, and as a result, treatment may be delayed. integrated electronic patient record (epr) systems are now widely used, and provide an opportunity to retrospectively audit practice to identify patients in whom vod may have been un-diagnosed or in whom treatment was delayed. in addition these systems have potential for alerting clinicians to the potential diagnosis of vod. methods: we have developed software to analyse the data downloaded from epr to identify patients in whom vod was a possible diagnosis according to the new ebmt criteria. in order to identify patients who may have had vod we first screened for patients with a bilirubin of >= mmol/l (which is an absolute requirement for the clinical diagnosis of vod) within the first days of transplantation. epr data was then used to assess whether patients had > % weight gain. radiology reports were reviewed for patients who had bilirubin >= mmol/l to ascertain if they revealed ascites or painful hepatomegaly. results: patients underwent transplant procedures (january st to july st ). of all transplant patients ( . %) were found to have a bilirubin of >= mmol/l. of ( . %) autograft patients and of ( . %) allograft patients had an elevated bilirubin at this level. these patients were assessed for evidence of % weight gain. this was the case in patients overall- % of autograft patients, . % of allograft patients. seven patients ( autograft and allograft) had radiological evidence of ascites. two patients had a recording of painful hepatomegaly (both post allograft). overall our analysis identified patients ( . % overall) fulfilling the ebmt diagnostic criteria for classic early vod all of whom received defibrotide. all patients had received allogeneic transplants. we failed to identify any cases of late onset vod or any undiagnosed patients over this period. conclusions: this analysis enabled us to efficiently perform a complete audit of our practice to identify patients with vod. we would recommend using electronic patient records to retrospectively audit practice in this way. the tool that we have created for this analysis will be made freely available for public use and the details will be presented at the ebmt meeting. we now plan to extend the function of our epr system to provide alerts to clinicians when vod is a possible diagnosis and may lead to more rapid treatment of these patients. our data suggests that elevation of bilirubin and weight gain of > % will be the most frequently occurring criteria on which to base these alerts. disclosure: g.vallance has performed consultancy work for jazz pharmaceuticals. endothelial activation and stress index in predicting outcome of allogeneic stem cell transplantation-a retrospective cohort analysis zinaida peric , tomislav taborsak , nadira durakovic , lana desnica , alen ostojic , ranka serventi-seiwerth , radovan vrhovac university hospital centre zagreb, zagreb, croatia background: endothelial dysfunction is a common pathophysiology of major complications after allo-sct, such as graft-versus-host disease, veno-occlusive disease, thrombotic microangiopathy and sepsis. endothelial activation and stress index (easix) is a simple score comprised of standard laboratory parameters (creatinine, ldh and thrombocytes) developed as a potential tool to predict allo-sct mortality by luft and colleagues. a recent validation of easix included three retrospective cohorts and showed that easix taken before start of conditioning can be used as an independent predictor of survival after allo-sct. methods: the aim of our study was to retrospectively evaluate pre-transplant easix in our cohort of consecutive patients who underwent allo-sct in the university hospital centre zagreb between and . with the use of a cut-off used in the validation cohorts, we compared two groups of patients for overall survival (os) and transplantrelated mortality (trm). group comparisons were done using the log-rank test or gray test for competing risks outcomes. a multivariate analysis evaluated the association of os with relevant variables by using a cox's proportionalhazard regression model. results: our study group included patients and comprised males ( %) and females ( %, with a median age of years (range, to years) at the time of transplantation. the most frequent malignancies in our population were acute leukemia ( patients, %) and myelodysplastic/myeloproliferative neoplasm ( patients; %). the donor was an identical sibling for patients ( %), matched unrelated donor for patients ( %) and haploidentical for patients ( %). patients ( %) received a myeloablative conditioning regimen while patients ( %) received a reduced-intensity conditioning regimen. with a median follow-up of months (range, - ) for the whole study group, the os at months was %, ( %ci - ) in the group of patients with low easix score and % ( % ci - ) in the group of patients with high easix score (p= . ). this difference was mainly attributed to higher trm in the group with high easix score ( %, %ci - at months) compared to the group with low easix score ( %, %ci - at months) (p= . ). in the multivariate analysis which included easix, patients' age, intensity of conditioning, diagnosis (lymphoid vs myeloid), status of the disease at transplant and type of the donor, worse os was independently associated only with older age of patients (hr . ; % ci, . - . , p= . ) and high easix score (hr . ; % ci, . - . , p= . ). conclusions: our retrospective data support previous data and suggest that easix could potentially serve as a valid tool for prediction of allo-sct outcomes. as a simple biomarker panel, easix could easily be implemented in clinical decision making in the field of allo-sct. these retrospective data need validation in a prospective study which is currently being conducted. clinical background: veno-occlusive disease (vod) is a potentially devastating complication that can occur after hematopoietic stem cell transplant (hsct) and in severe cases can lead to multi-organ failure. (mohty ) defibrotide has been proven to be effective to prevent and treat vod, and it is critical that clinicians are aware of how to diagnose and treat this serious complication of hsct. this study was conducted to determine if an online, simulation-based continuing medical education (cme) intervention could improve performance of hematologists/oncologists (hem/ onc) and advanced practice providers (nurse practitioners and physician assistants, apps) in the diagnosis and treatment of patients with vod. ( methods: a cme certified virtual patient simulation (vps) was made available via a website dedicated to continuous professional development. the vps consisted of cases presented in a platform that allows clinicians to assess the patients and make diagnostic and therapeutic decisions supported by an extensive database of diagnostic and treatment possibilities, matching the scope and depth of actual practice. clinical decisions were analyzed using a sophisticated decision engine, and tailored clinical guidance (cg) employing up-to-date evidence-based and faculty recommendations was provided after each decision. one case was about vod and the other case was about acute myeloid leukemia (aml). decisions were collected post-cg and compared with each user's baseline (pre-cg) decisions using a -tailed paired t-test to determine p-values (p < . indicates significance). data were collected between / / and / / . results: at the time of assessment, hem/oncs and apps had fulfilled the participation criteria for completing the vod case simulation. conclusions: this study demonstrates that vps that immersed and engaged clinicians in an authentic and practical learning experience improved evidence-based clinical decisions related to the management of vod. this vps increased the percentage of clinicians who utilized standardized criteria to diagnose vod and who ordered defibrotide and iv fluids for vod management. however, further education is needed to increase the competence and performance of clinicians, particularly apps, in these areas in order to positively impact patients. disclosure: nothing to declare. a nationwide retrospective study of hematopoietic stem cell transplantation in solid organ transplant recipients: on behalf of jshct, transplant complications working group background: the outcome of hematopoietic stem cell transplantation (hsct) in solid organ transplant remain unclear. to address this issue, we conducted a retrospective survey of the japan society for hematopoietic stem cell transplantation centers. methods: to address this issue, we conducted a nationwide retrospective survey of the japan society for hematopoietic stem cell transplantation (jshct) centers. a first questionnaire was emailed to jshct centers requesting information on cases of hsct in sot recipient. patients' data about sot were collected by sending a second questionnaire to the centers with the patient. based on these reports, patients' data about hsct was identified in the japan transplant outcomes registry database by the transplant registry unified management program (trump), confirmed in . results: of the jshct centers, responded to the survey ( . %). of the responding centers reported a total of patients who had undergone sot from living donor, and subsequent hsct. they consist of three autologous hsct (auto-hsct) and allogeneic hsct (allo-hsct). in auto-hsct, all patients had received liver transplant for hapatoblastoma. they achieved neutrophil engraftment at days after hsct, and two of three patients were alive at one year after hsct. in allo-hsct (n= ), seven patients had received liver transplants, and nine patients had received kidney transplants. five patients received hsct from unrelated donor, and patients received hsct from related donor; two donors were identical in sot. their stem cell sources were seven peripheral blood stem cell, six bone marrow, and three cord blood. all but one patients achieved neutrophil engraftment at days after hsct. five-year overall survival ( yos) was . %. while yos in patients with bone marrow failure (n= ) was %, that in patients with malignant disease (n= ) was . %; all but one patients with malignant disease received allo-hsct in non-remission. seven of nine kidney-transplant recipients experienced dialysis, and three patients experienced renal rejection after hsct. on the contrary, no liver-transplant recipient experienced hepatic rejection. conclusions: in sot recipients, the outcome of allo-hsct for malignant disease was poor, partly due to disease status before allo-hsct. severe renal complications were common in kidney-transplant recipients, suggesting renal care with caution during and after allo-hsct. disclosure: this work was supported in part by the practical research project for allergic diseases and immunology (research technology of medical transplantation) from japan agency for medical research and development, amed. high incidence but low mortality of ebv related ptld after t-cell replete allo-peripheral blood hct with aggressive monitoring and without pre-emptive rituximab background: the aim of the study is to report the incidence and outcome of post-transplant lymphoproliferative disorder (ptld) in the setting of allogeneic peripheral blood hematopoietic stem cell transplantation (allo-hsct) combining post-transplant cyclophosphamide (ptcy) and anti-thymocyte globulin (atg) as graft versus host disease (gvhd) prophylaxis. methods: between october and may , adult patients diagnosed with hematological malignancies underwent a first t-cell replete allo-hsct in our center. all patients received a reduced intensity conditioning regimen with fludarabine, busulfan, and cgy of total body irradiation, combined with rabbit-atg, ptcy and cyclosporine (csa). ebv titres were monitored weekly by quantitative pcr in plasma samples. the cut-off value for test positivity was > copies of ebv dna/ml of plasma. last follow up was november . median follow up for patients known to be alive was months (range - ). results: patient information is summarized in table . ebv reactivation was documented in ( %) patients. median time to ebv reactivation and the diagnosis of presumed/proven (p/p)-ptld were ( - ) days and ( - ) days [ ( - ) months], respectively. median time between first ebv reactivation to p/p-ptld was ( - ) days. seventeen ( %) of the patients developed p/p-ptld. median age was years . two ( %) received mrd, ( %) / mud, ( %) / mud, and ( %) haploidentical donor grafts. twelve ( %) were on therapeutic cyclosporine at diagnosis. pre-emptive therapy was not given to any case and only probable or proven ptld were given rituximab. treatment was based on reduction of the immunosuppression in patients and with the addition of weekly rituximab mg/ m in cases. fifteen ( %) achieved complete clinical responses with pcr negativity. two ( %) patients died secondary to ptld. conclusions: atg based conditioning is associated with increased viral reactivations. frequent ebv monitoring and pre-emptive treatment may lead to rapid disease control. further research is required to optimize monitoring and management strategies in allo-hsct recipients. disclosure: nothing to declare p acoustically enriched extracellular vesicles as potential markers for allogeneic hematopoietic stem cell transplantation complications hooi-ching lim , robert palmason , stig lenhoff , thomas laurell , stefan scheding , background: extracellular vesicles (evs) contain a number of condition-specific proteins, dna and rna types and might therefore be used for the early detection of posttransplant complications. however, traditional ev isolation (ultracentrifugation) is time consuming and requires large sample volumes thus making it difficult to perform longitudinal studies on larger patient cohorts. we therefore investigated whether recently-developed acoustic trapping could be applied to isolate evs from patient plasma for biomarker development. methods: plasma samples were collected from consecutive patients before and up to months after allogeneic hematopoietic stem cell transplantation. patients (age: - years) with high-risk or refractory/relapsed diseases were transplanted with mobilized pbsc from related (n= ) and unrelated donors (n= ) after standard conditioning. gvhd prophylaxis was cyclosporine and methotrexate. plasma samples were frozen and thawed for ev enrichment using a novel acoustofluidic-based technology (acoustic trapping). acoustic trapping uses ultrasound as a local λ/ acoustic standing wave produced by a piezoelectric transducer over a capillary. first, μm polystyrene beads are captured which serve as seeding particles. after washing, target particles (evs) are then captured ("trapped") in the acoustic field. a semi-automatic trapping device (acoutrap) was used to isolate evs from diluted plasma ( : in pbs). the number of evs and size distribution were analyzed by nanoparticle tracking analysis. mirna analysis was performed by qpcr.evs were enriched in duplicate from μl and μl of diluted plasma for nanoparticle tracking analysis and qpcr analysis, respectively. results: evs were successfully isolated from all plasma samples. a total of plasma samples were processed. numbers of trapped evs ranged from . x - . x before conditioning to . x - . x per μl diluted plasma after transplantation. the maximum change in ev numbers in individual patients compared to pretransplantation values ranged from -fold to -fold. most patients showed slight increases in ev size after transplantation. eight of the patients showed signs of infection and received i.v. antibiotics. increased levels of evs (> -fold) were recorded in three patients during these episodes. furthermore, increased ev numbers were observed in a patient who required i.v. antiviral therapy for cmv reactivation. acute grade i gvhd was observed in five patients of which two had increased ev numbers (> -fold). one patient developed grade iv gvhd which was accompanied by a -fold increase in ev numbers. interestingly, progressively increasing ev numbers preceded the detection of early relapse in a pre-b all patient by three weeks. rna isolation from trapped evs yielded sufficient material for mirna profiling. here, first mirna profiling data demonstrated that mirnas were detected in ev samples (mir- a, - a, - c, - and - a) , and that acoustically enriched evs were not affected by hemolysis in contrast to the corresponding whole plasma samples (dcq of mir- a and mir- a). conclusions: acoustic trapping allows for efficient and rapid enrichment of evs from small volume plasma samples. trapped ev samples contain sufficient amounts of mirna for downstream analysis and are thus promising candidates for biomarker development in transplantation. disclosure: laurell and scheding are founders and board members of acousort ab, a lund-based biotech sme that develops particle and cell sorting methods based on ultrasound. the incidence, risk factors and outcomes of primary poor graft function after allogeneic hematopoietic stem cell transplantation fei gao , , jimin shi , , yi luo , , yamin tan , , xiaoyu lai , , jian yu , , he huang , , yanmin zhao , background: allogeneic hematopoietic stem cell transplantation (allo-hsct) is a curative therapy for both hematologic malignancy and many other blood disease. while, primary poor graft function (pgf) is still a severe complication following hsct which lead to poor prognosis. up to now, the incidence and risk factors of pgf have not been totally revealed. methods: from january to december , a total of patients who received allo-hsct in our center were analyzed retrospectively. there were males ( . %) and females ( . %) with a median age of . years ( - years) . pgf was defined as persistent neutropenia (≤ . × /l), thrombocytopenia (platelets≤ × /l), and/ or hemoglobin≤ g/l after engraftment with hypocellular bone marrow and full donor chimerism, without concurrent graft-versus-host disease or disease relapse. incidence was calculated from all patients. of the total patients, nineteen ( . %) developed primary pgf. a : ratio of nested case control study using the good graft function (ggf) subjects transplanted in the same year with the same sex and age of ± years was carried out. results: data was analyzed by univariate and multivariate logistic regression, and univariate analysis identified disease species, the time from diagnosis to transplantation, disease states, myelofibrosis, splenomegaly, serum ferritin (sf) level, cmv infection, mononuclear and cd + cells in graft as potential risk factors (p < . ) for pgf. multivariate analysis identified elements as the independent risk factors (p < . ), including cd + cells < × / background: transplant survivors affected by cgvhd usually take one or more immunosuppressants, as well as prophylactic antimicrobials; use of multiple medication classes concurrently poses a risk for drug-drug interactions or amplified side-effects. the use of medications other than cgvhd-direct immunosuppressive therapies has not been well-characterized. this study aims to evaluate patterns of opioid analgesic use in a cohort of patients severely affected by cgvhd. methods: patients (n= ) with cgvhd were consecutively enrolled in a cross-sectional natural history study (nct ) from / - / at the nih. participants underwent a comprehensive evaluation including a detailed history and physical examination (including current medications), multidisciplinary evaluations, and laboratory and diagnostic testing. for this analysis, respondents were classified as receiving or not receiving an opioid analgesic. following the initial screening by univariate methods (n= ), multivariable logistic regression analysis (mlr) was used to identify a set of factors which could jointly impact opioid use. for mlr data were divided into a training (n= patients) and a validation set (n= ). results: study participants´median age was . years ( - ), % were female, % had severe cgvhd per nih scoring criteria, and % were currently receiving high or moderate levels of systemic immunosuppression. approximately one third ( %) were taking opioid analgesics (oa). based on the univariate screening results (p< = . ), a set of parameters was evaluated by univariate logistic regression in the -patient training set, and the following parameters retained their significance and were included in the mlr model: nih average score per organ, total lss, patient impression of severity, nih cgvhd severity, presence of skin erythema, karnofsky performance score (kps,) clinician's therapeutic intent, nih joint score, and with the presence of several cgvhd symptoms including rashes, mouth sores, avoidance of food, vomiting, weight loss, joint and muscle aches, joint limitation, energy loss, need for naps, fevers, anxiety. multivariable logistic regression identified kps < % as predictive of oa use, or . , % ci . - . . in the training set . % of pts using opioids were correctly identified, . % of those not taking opioids were identified, an overall fraction of correctly identified pts was . % ( % ci . - . %), while in the testing set, . % of those using opioids were correctly identified, and . % of those not taking opioids were correctly identified, with overall . % ( % ci: . - . %) classification accuracy. conclusions: this study showed the burden of oa in this cgvhd cohort. lower kps was significantly associated with oa use, as well as self-reported symptoms and a more severe cgvhd disease, which could be of interest in the development of non-pharmaceutical interventions in this patient population. additional, prospective studies are needed to explore the indications for and effectiveness of oa in this population of survivors. disclosure: no conflict of interest to declare. rcts that tested an internet-based program and patientcentered survivorship care plans for hct survivors. patient and caregiver input is essential to inform the design and features for the mobile app platform so that it is usable and engaging for those it targets. methods: using a qualitative research design, we conducted telephone focus groups of adult patients and caregivers in the united states. adult (age > years at the time of study entry) hct recipients had to be at least oneyear post-hct to participate. participants had to be able to communicate in english, and could have received a hct for any diagnosis, and from any donor source or stem cell type. those who had multiple transplants were included. participants were asked to review printed and online visual presentations of the mobile app before the focus groups so they were prepared to discuss their responses to the materials during the call. focus groups were conducted to saturation, when no new qualitative content was offered. results: three focus groups were conducted with total participants ( patients, two caregivers/patient advocates). all patients received an allogeneic hct; average time since hct was years (range: - years).the majority of participants were female ( . %). participants had differing perspectives on the usefulness of the app to track follow-up appointments, lab values, and other health care plans. there was high interest in having the app tailored to meet specific needs of patients, including tracking information over time (e.g. test results, medications), and having health information available specific to their needs. to minimize duplication of information and data entry, participants recommended syncing the app with their calendars and online patient portals they already use. reasons provided for not using the app included perception that the materials repeated information already received, side effects such as graft-versus-host disease that restricted vision or motor skills, and lack of comfort with apps for some older participants. conclusions: many health technology and mobile apps are being created to improve patients' health and survivorship care. in this study, hct survivors and caregivers identified a variety of features that they would want in an app or website, in particular, features tailored to individual needs. health technologies provide an opportunity to improve survivorship care, but patients and caregivers should be engaged in the process of developing these tools to assure the technology fits their needs and will be used. given the effort required to maintain these technologies, they require testing for health benefits in rigorous clinical trials. clinical background: thanks to allogeneic stem cell transplantation (allo-hsct) patients suffering from hematologic malignancies have seen an increase in there life duration expectancy, but they are many side effets including decreasing in physical performance and in quality of life. the intensity of physical performance decrease is variable between patients, and today we did not know why. the aim of our study was first to characterize the physical performance of subjects less than year following allo-hsct by the use of a cardiopulmonary exercise testing (cpet), and then to determine the predictive factors of exercise performance. methods: we did a retrospective analysis from patients who had an allo-hsct at hematology department of toulouse-oncopole and cpet from / to / . the cpet was performed using a cycle ergometer with o and co analyzer breath by breath, (masterscreen cpx carefusion, san diego, usa), a continuous -lead electrocardiogram, and a blood pressure monitoring. the protocol included a -min rest period, a -min warm-up of w pedaling followed by a w/min incremental phase, up to exhaustion, then a -min active recovery of w pedaling, then a -min passive recovery. three exercise markers were analysed: the peak of oxygen uptake (peak vo ), the ve/vco slope and the first ventilatory threshold (vt ). data relative to conditioning regimen, short-term complications, impairment at cpet day, and physical activity since allo-hsct were gathered. results: after allo-hsct, nearly over patients reported fatigue, a half reported dyspnea, and over or more reported pain, muscular, neurological or psychological impairment. more than % of patients suffer from moderate or severe physical intolerance, particularly when myeloablative conditioning regimen was used. only % of patients followed rehabilitation sessions supervised by a physiotherapist, and non-supervised physical activity has been performed by % of patients. despite normal lung function tests and echocardiography findings in most patients, % had exercise intolerance (ei), % exercise deconditioning, and % had abnormal ventilatory efficiency. patients with moderate and severe impaired exercise capacity were significantly younger at diagnosis and at allo-hsct, such as patients with severe deconditioning conclusions: based on a retrospective study, we reported for the first time complete results from cpet and detailed clinical evaluation concerning deficiency and disability following first year after allo-hsct. these results confirm that exercise impairment is very frequent with more than a half of patients suffering from alterations of one or more of the three performance markers, despite being active. disclosure: nothing to declare demyelinating disorders: a paradigm of immunity disorders after hematopoietic stem cell transplantation background: neurologic complications are a major problem in patients who undergone hematopoietic stem cell transplantation (hsct). given the higher survival of transplanted patients, the burden of neurological complications is increasing in the last years. a significant reduction in overall survival was demonstrated in patients who developed neurological complication after hsct, irrespectively of the hematopoietic stem cell (hsc) source. neurologic disorders in transplanting setting comprise a wide variety of ethiologies including demyelinating disease, which are caused by immune and non-immune mechanisms. here, we analyzed the clinical presentation and the underlie ethiologies of patients developing hsct-related demyelinating disorders in order to give diagnostic and prognostic clues useful to manage these severe but treatable complications in the transplant setting. methods: a total of patients of our department which developed neurological complications after hsct were consecutively collected and ( %) of them, namely those having a diagnosis of a demyelinating disorder, were grouped and described according to the ethiologies of their neurological disorder. results: in / ( %) patients, an immune-mediated process was found, while / ( %) were diagnosed as having an infective etiology and / ( %) were supposed to have a demyelinating disorder caused by toxic exposition. a definitive etiologic diagnosis was not formulated in the remaining / ( %) patients. when patients who developed an immune-mediated demyelinating disorder ( / ) were compared to those in which a clear immune pathogenic mechanism was not detected ( / ), a higher incidence of acute graft-versus-host disease (agvhd) was detected in the former than in the latter ( % vs %). moreover, comparison of these two groups revealed that those with no evidence of immune-mediated process have a slight higher prevalence of t-cell depleted hsct thanthose with an immune-mediated demyelinating disorder ( % vs %). finally, a lymphoproliferative disorder pre-existing the hsct was detected in / ( %) patients with immune-mediated demyelinating disorder but only in / ( %) of those without evidence of immune-mediated processes. conclusions: demyelinating disorders may be responsible of near % of neurologic complications in the posttransplant setting and, among them, an immune-mediated process is likely to be involved in more than % of cases. our results suggest that the immune mechanism that underliesthe agvhd may also be involved in developing demyelinating disease in transplanted patients. it also may be possible that the lymphoproliferative disorder preexisting the hsct is a risk factor able to increase the risk to develop an immune-mediated demyelinating disorder in the post-transplanting setting. using a t-cell depleted hsct can increase the risk of immune-mediated disorders in at least a small fraction of transplanted patients. despite our results should be validated on a larger cohort of patients, we can speculate on the possible connections between the wide range of complex and still poorly defined immunity disorders which can influence the prognosis and course of transplanted patients. disclosure background: injury to the mucosal barrier and subsequent development of oral mucositis (om) is among the most common toxicities of allogeneic stem cell transplantation (sct). despite the high prevalence of om and its debilitating nature, prospective studies evaluating determinants of om are scarce. we therefore prospectively evaluated the occurrence of om following sct. risk factors for om and its implications short and long-term outcomes were assessed. methods: om was prospectively evaluated on a weekly basis in patients undergoing allogeneic hsct. the grade of om was determined based on the national cancer institute common toxicity criteria for adverse events (ctcae) scale (v. . ). severe om was defined as grade ii to iv. conditioning regimens were evaluated individually and according to intensity; myeloablative (mac), reduced intensity (ric) or reduced toxicity (rtc). the latter category included only patients receiving fludarabine and treosulfan at dose of - g/m (flu/treo). risk factors for the development of severe om were initially identified by a univariate analysis and then analyzed in a multivariate logistic regression model. association of om with peritransplant infectious complications, iv morphine consumption, hospitalization length, neutrophil engraftment, acute and chronic graft-versus-host disease (gvhd), non-relapse mortality (nrm) and overall survival were assessed in a univariate analysis. competing events were considered in analyzing engraftment, gvhd, and nrm. results: patients who underwent an allogeneic sct between and were included. median follow-up was days. leading indications for transplantation were acute myeloid leukemia ( %), lymphoma ( %), and myelodysplastic syndrome ( %). the majority of patients received an allograft from a matched sibling or unrelated donor ( %) and methotrexate gvhd prophylaxis ( %). the median time to om onset was (interquartile range [iqr] - ) days. prevalence of grade ii-iv om was %. the median duration om was [ - ] days, and iv morphine was administrated for a median of [ - ] days for patients with grades iii-iv om ( %). in a univariate analysis a younger age (p= . ), lower bmi (p= . ), recent smoking history (p= . ), recent antibiotics exposure (p= . ), mac (p< . ), and use of methotrexate (p= . ) were associated with an increased risk for grade ii-iv om. in a multivariable model the risk for grade ii-iv om was lower with rtc (i.e., flu/treo) vs. mac (odds ratio [or] . ; p< . ) and rtc vs ric (or . ; p= . ), mycophenolate mofetil vs. methotrexate (or . ; p= . ) and recent smoking (or . ; p= . ). compared to lower grades, grade ii-iv om was associated with a longer hospitalization duration (median days vs. days; p= . ), delayed neutrophil engraftment (median vs. days; p=- . ), and more gastrointestinal related infections ( % vs. %; p= . ). grade ii-iv om was not associated with increased risk of bloodstream infections, acute or chronic gvhd, non-relapse mortality, and increased mortality. conclusions: oral mucositis is prevalent among allogeneic-sct recipients. importantly, fludarabine-treosulfan, which is considered a myeloablative is associated with a markedly reduced risk for om. consequences of om include prolongation of hospitalization, delay in neutrophil engraftment, and a tendency for gastrointestinal infections, but does not increase the risk for gvhd and mortality. disclosure: nothing to declare background: the advent of recent diagnostic techniques for the assessment of iron overload (t *-mri) and their systematic use as screening tools in the setting of secondary hemochromatosis have led to an increased awareness that focal nodular hyperplasia (fnh) represents a possible incidental finding after hematopoietic stem cell transplantation (hsct). methods: clinical and radiological features of patients undergoing hsct in a single pediatric institution have been retrospectively reviewed for fnh. in order to provide an estimate of the prevalence of fnh after hsct, we analysed all the t *-mri scans performed during the last years in our centre and recorded the number of patients with fnh (group a). in addition, data about patients incidentally diagnosed with fnh at abdominal imaging performed for different clinical indications have been collected (group b). results: eight out of ( %) transplanted patients who underwent at least one t *-mri scan from september to september were incidentally diagnosed with fnh. group b included subjects with fnh incidentally found at ultrasound or non-t * mri scans performed before . overall, transplanted patients ( males, %), transplanted for al ( cases) or bone marrow failure ( cases) at a median age of . ± . years, were diagnosed with fnh between . and . years after hsct, namely . ± . years in group a and . ± . years in group b. a variable degree of iron overload was demonstrated in all patient (lic: - ± microg/g; baseline serum ferritin: - ng/ml). the potential risk factors for fnh are reported in table . in / patients, the radiological finding was pathognomonic; in / the diagnosis of fnh was confirmed histologically, while / subjects were labelled as "fnhlike", although a potential diagnosis of hepatic adenoma could not be ruled out. in / patients, fnh presented with an isolated lesion, while / had to more than hepatic nodules at diagnosis. the size of nodules at diagnosis ranged from to mm. in unenhanced mri scans, lesions were predominantly hyperintense on both t -and t weighted sequences. in dynamic studies with contrast medium, all lesions strongly enhanced during the arterial phase, with a variable degree of wash-out in the late venous scans. hepatic function tests were normal in all the enrolled patients at diagnosis of fnh. among the / patients for whom at least a follow-up scan was available, presented a complete regression, a reduction and an increase in the size and/or number of lesions, while in patients the nodules remained substantially unchanged after a mean radiological follow-up of . ± . years. no malignant transformations were observed. conclusions: fnh represents a relatively frequent incidental finding after hsct. although a malignant transformation is rare, given the demonstrated variable evolution of the hepatic nodules, a radiological follow-up is highly recommended. disclosure: nothing to disclose. incidence and risk factors for hepatic sinusoidal obstruction syndrome after allogeneic transplantation: retrospective multicenter study of turkish hematology research and education group (threg), updated data methods: ten centers from turkey were enrolled in the study. we retrospectively evaluated the medical records of patients who were treated with allo-sct between january and december . a baltimore criterion was used for assessment of hsos. four hundred twenty six ( . %) of patients who were treated with prophylaxis with defibrotide alone or one or more of the n-acetylcysteine, diuretics and heparin used defibrotide ( - mg/kg/day). results: the study included patients ( males/ females) with median age of ( - ) years. the demographic and clinical characteristics of patients were summarized in table seventy-three ( . %) of patients with hsos were treated with defibrotide after diagnosis. the median time of starting defibrotide in these patients was . ( - ) days. thirty-seven ( %) of patients with hsos recovered completely and forty-nine ( %) of them died as a result of multi organ failure. the incidence of hsos-related mortality in allo-hsct cohort was found to be . %. in univariate analysis, statistically significant associations were not found between hsos incidence and age/sex of recipient, type of conditioning regimen, stem cell source and type of gvhd prophylaxis. on the other hand donor type, engraftment status and prophylaxis for hsos were significantly associated with hsos development. hsos prophylaxis was significantly decreased hsos-associated mortality (p= . ). conclusions: hsos still remains a serious lifethreatening complication of allo-sct. although the incidence is low, hsos is associated with increased -day non-relapse mortality. hsos prophylaxis especially with defibrotide, seems to reduce hsos associated mortality in high risk patients. disclosure: nothing to declare prophylaxis with defibrotide in adults at very high risk of veno-occlusive disease: results in patients background: hepatic sinusoidal obstruction syndrome/ veno-occlusive disease (sos/vod) is a life threatening complication that can occur after hematopoietic stem cell transplantation (hsct). severe sos/vod rapidly evolves in multiple organ dysfunction syndrome (mods), associated with a mortality rate exceding %. precocity of defibrotide (df) treatment is the leading factor for efficacy. prophylactic use of df is recommended in children, but its value has not been validated in the adult population, although factors for individual risk assessment for vod are debated. we here present a real-world experience of df prophylaxis in adult patients at very high risk of sos/vod receiving allogeneic hsct. methods: from to we treated with prophylactic defibrotide and ursodeoxycholic acid (udca) patients, median age years (range - ). nine patients received allogeneic hsct for acute lymphoblastic leukemia ( b-all and t-all), one patient for severe aplastic anemia, one patient for primary myelofibrosis. they were all at high risk for sos/vod because of previous hepatotoxicity ( patients), previous hsct ( patients), double alkylating agent ( patients) or previous treatment with inotuzomab ozogamicin (io; patients). of the patients treated with io, received cycles of io, and received cycle, with the last io dose administered a median . days before hsct (range - d). defibrotide was administered in daily doses for a total dose of mg/ kg per day and udca at the dose of mg twice per day, starting from day - prior transplant. all patients received treosulfan-fludarabine based conditioning. in patients thiotepa was added to the conditioning and in patients a low dose gy tbi. gvhd prophylaxis included posttransplant cyclophosphamide, rapamycin and mycophenolate in all patients, except one patient with aplastic anemia receiving atg, rapamycin and mycophenolate. donor source was pbsc in all cases. seven patients received family haploidentical (mmrd) transplant, patient a mrd transplant and patients a mud transplant. results: the median duration of defibrotide therapy was days (range - days). documented non-severe gastrointestinal bleeding occurred in patients requiring defibrotide temporarily discontinuation, no other significant bleedings were experienced. four patients developed grade ii-iv acute gvhd and no transplant-associated thrombotic microangiopathy were diagnosed. overall, sos/vod occurred in / cases within days after hsct (days , and ) and no late-onset sos/vod were diagnosed. sos/vod was very severe, causing mods and death in all cases. all patients were characterized by a common pattern of very high risk factors for sos/vod by prior hsct and salvage treatment for b-all with cycles of io close to hsct. furthermore, they all received a fully myeloablative conditioning regimen with treosulfan and thiotepa and a mmrd transplant. conclusions: defibrotide prophylaxis was safe and well tolerated with no severe related complications. sos/vod occurred despite continuous df prophylaxis in / patients treated with inotuzomab ozogamicin close before undergoing nd transplant. to reduce the incidence of severe vod, pre allo-hsct treatment with inotuzomab ozogamicin should prompt avoidance of other cumulative risk factors for vod, such as use of double alkylating agents. disclosure background: busulfan is the backbone of many preparative regimens administered to children undergoing allogeneic and autologous hematopoietic stem cell transplantation (hsct). among its many long-term adverse effects, busulfan can cause various degrees of pulmonary injury. although well described in adults, there are few large series exploring pulmonary toxicity of busulfan in children. we describe long-term pulmonary follow-up in a large group of children treated at a single center who had received high-dose busulfan and examine the relationship of systemic drug exposure and lung function over time. methods: all surviving children who had received highdose busulfan between - in the context of hsct at the schneider children´s medical center, were referred for serial pulmonary function monitoring (including spirometry, plethysmography and diffusing capacity for carbon monoxide [dlco] . pre-transplant testing was available for children who were old enough to perform the procedure. spirometry results were adjusted according to the revised global lung initiative formulas for age, gender, and height. pulmonary injury was defined as a z score below - . for spirometry, or < % of predicted for the other parameters. busulfan levels were monitored following the second drug dose. all patients received busulfan in four daily doses. area under the curve (auc) calculations were performed by bayesian calculations. results: between - , patients aged - years were diagnosed with malignant or non-malignant diseases and treated with high-dose busulfan. of shortterm survivors, had at least one post-transplant pulmonary function evaluation. the mean age at treatment with busulfan was . years (range, . - years). of these children, children had undergone autologous transplantation and children had an allogeneic transplant. of these patients eventually relapsed and died. children had one or more pulmonary risk factors before hsct -chest or upper abdomen radiation ( ), chest wall tumors or lung metastasis ( ), chest surgery ( ), prior administration of pulmonary-toxic drug ( ) or asthma ( ). during follow-up (up to years, median . years), fev and fvc spirometry tests both decreased significantly (p= . and . , respectively), while the decrease in dlco was not statistically significant. % of patients had abnormal pulmonary function tests and seven children had symptomatic disease which in two may have been manifestations of gvhd. interestingly, no correlation was found between busulfan auc, busulfan peak levels, the number of busulfan doses administered, the type of transplantation (autologous vs. allogeneic) or primary disease to pulmonary injury. even after censoring of children with pre-transplant pulmonary risk factors we noted a decrease of fev and fvc. conclusions: as in adults, pulmonary injury is observed in children treated with high-dose busulfan prior to hsct. no correlation was observed between busulfan auc and pulmonary injury. follow-up of children who receive this drug should include regular pulmonary monitoring, referral to a pulmonologist when subclinical pulmonary compromise is found, and counseling regarding measures that might prevent or ameliorate pulmonary damage. continued follow-up of this cohort of patients should inform our pretransplant patient information sessions, and the future use of busulfan in children. disclosure: nothing to declare background: transplant-associated thrombotic microangiopathy (ta-tma) is a specific complication of allogeneic hematopoietic stem cell transplantation (hsct). post-hsct tma has been attributed to the vascular endothelial damage caused by high-dose chemotherapy, calcineurin inhibitors (cnis), graft-versus-host disease (gvhd), infections. there is a little evidence published regarding the efficacy and factors influencing the outcome of withdrawal of cnis. methods: the analysis comprised a total of patients, with diagnosed hematologic malignancy (aml ( ), all ( ), mds ( ), hodgkin lymphoma ( ), cml ( ) and neuroblastoma ( ) received allo-sct, from a matched related, unrelated or haploidentical donor between and . patients were diagnosed with ta-tma based on cho criteria. the median age of patients was ( adults, children). gvhd prophylaxis was performed with tacrolimus (tac) in , cyclosporine a(csa) in , combination tacrolimus+sirolimus (sir) in . patients received atg and ptcy. withdrawal of cnis was accompanied by administration of systemic steroids ( patients) or substitution with sir after reaching levels of csa< ng/ml or tac < ng/ml in . the target concentration of sir was - ng/ml. in pediatric patients who received combination tac+sir, the tac was discontinued in one step while sir continued. median time to development tma was , days after allo-sct (range - ). median follow-up of surviving patients was days. the primary outcome was overall survival (os) up to years after development of ta-tma. results: the following significant predictors of -year os were identified: tac replacement with sir (p< , ), ptcy in prophylaxis (p< , ), acute gvhd (agvhd) grade - (p= , ), previous sepsis (p= , ), level of ldh in debut (p= , ), combination sir+tac in prophylaxis (p= , ), major ab -mismatch (p= , ), severity of cns symptoms (p< , ). there was no significant difference in os according to patients' age, sex, "salvage" disease status at transplantation, previous vod, viral (hhv , , , cmv, ebv) reactivations, count of cd + cells transfused, ldh level, shizocytes and creatinine in the debut of ta-tma. in the multivariate analysis replacement of cnis with sir (hr . , %ci . - . , p= . ) and baseline ldh level (hr . , %hr . - . , p= . ) were associated with survival differences. the cut off for ldh was xunl. agvhd grade - (hr . , p= , ) and use of ptcy (hr . , p= . ) were not significant in the multivariate analysis (figure ). ta-tma cases after ptcy were significantly less frequently associated with clinically significant agvhd ( % vs %, p< , ). the survival was higher after ptcy ( % vs %), but not significant due to sample size and other ta-tma factors. leading causes of death were: gvhd progression ( %), bacterial infection ( %), tma ( %) and other ( %) . conclusions: replacing tac by sir is an effective therapeutic strategy in a group of patients with debut of ta-tma at least after ptcy, where it is less likely to be associated with agvhd. there is a significant overlap of populations with ptcy prophylaxis and substitution with sir, thus the study is not powered to provide guidance for patients on conventional prophylaxis with ta-tma. [[p image] . disclosure: none of the authors has anything to disclose. donor-recipient ab mismatch effect on the allogeneic hematopoietic stem cell transplantation outcome: a single-center retrospective study background: because transmission of major histocompatibility complex and blood group system genes is independent from each other, approximately - % of all allogeneic hematopoietic stem cell transplantations (allo-hsct) are realized crosswise the ab -blood group boundary. however, due to the widespread expression of ab antigens on a variety of human tissues other than erythrocytes, ab incompatibility may have an impact on the outcome of allogeneic hsct that goes beyond the wellknown immune-hematological complications such as immediate hemolysis due to the presence of isoagglutinins and delayed hemolysis due to passenger b lymphocytes. here we aimed to assess the donor-recipient ab mismatch effect on the allo-hsct outcome, comprising non-relapse mortality (nrm), overall and relapse-free survival, posttransplant prc transfusion requirement, as well as relapse rate, incidence of graft-failure and acute gvhd. methods: clinical and laboratory data from consecutive patients undergoing allogeneic hsct between / and / at the fondazione irccs ca' granda maggiore policlinico hospital in milan, italy, were retrospectively collected. kaplan meier estimates were used for the analysis of survival outcomes while nrm, relapse and acute gvhd cumulative incidences were investigated by competing risk analysis. results: the patient series included ab -match, major ab -mismatch, minor ab -mismatched and bidirectionally ab -mismatch transplants. indication for allo-hsct were mainly aml/mds ( pts), all ( pts) and t-nhl/ctcl ( pts). mean overall survival for groups of patients undergoing ab -identical, major ab mismatch and minor ab mismatch hsct were months ( % ci [ ; ]), months ( % ci [ ; ) and months ( % ci [ ; ]), respectively. nrm in the three groups were significantly different, with point estimates of %, % and % at years, respectively, whereas no significant differences were observed for relapse rate and graft failure incidence. although not statistically different, incidence of acute grade iii-iv gvhd was twice as high in patients transplanted from minor ab mismatched donors than in the ab identical group ( % vs %). following transplantation, prc transfusion requirement was significantly higher in the major ab mismatch then in the ab -match transplanted patients (median vs , p= . ), with a marginal positive correlation between the anti-donor a/b igg titers measured prior hsct and the total number of prc transfused during the first year following transplantation. we observed only one case of prca occurring in a year-old + woman who was transplanted from a -yearold male a+ hla-identical sibling using peripheral blood as the stem cell source following a myeloablative conditioning for aml in first complete remission. anti-a igg isoagglutinin titers prior to transplantation were : . during the first year post transplantation, the patient required a total of prc transfusions, with gradual resolution occurring only after introduction of danazole treatment. conclusions: in our patient cohort, both major and minor ab mismatch associated to a significantly higher nrm. major ab mismatch associated to a higher prc transfusion requirement. a more frequently occurring severe acute gvhd was also suggested in minor ab -mismatch transplants. altogether, our results suggest that allo-hsct outcome may be significantly affected by ab blood group mismatch. disclosure: nothing to declare background: at-tma is a severe endothelial injury complication and it may involve the intestinal vasculature. intestinal tma could be fatal and missdiagnosed. clinical and pathological criteria to differentiate from intestinal gvhd are needed. the aim of this study was to analyze the incidence and histological characteristics of intestinal tma in patients diagnosed of systemic tma. methods: we analyzed the incidence of tma in patients who underwent allo-hsct in our institution between january -august . tma diagnosis was based on ho criteria. we do a pathological review in biopsies from out of patients in whom an endoscopy have been performed days before and days after the diagnosis of tma for suspicious of gvhd. review was performed by a pathologist expert in gvhd, who examined the biopsies in search of hystopathological features of gvhd, tma or viral infection. diagnosis of gastrointestinal gvhd was stablished according to mcdonald and sales criteria, while intestinal tma diagnosis was stablished by warren et al criteria. results: out of patients ( , %) were diagnosed of tma. transplant characteristics and tma data of patients with systemic tma are shown in image. out of patients with tma ( %) had been diagnosed with prior/ simultaneous acute gvhd, of them grade iii-iv, and % with gastrointestinal gvhd. intestinal tma have been reported only in out of patients ( %) at diagnosis, whereas when review based on warren criteria was performed, in patients ( %) the pathologist found at least of the criteria of endothelial damage and % of the patients or more warren criteria were founded. the most frequent features were endothelial cell swelling (n= , %) and perivascular mucosal hemorrhage (n= , %). review hystological features of biopsies are shown in table of the image. regarding gvhd, it was found in patients ( %) at diagnosis and in ( %) at pathological review. with a median follow-up of months ( - ) patients of the with systemic tma ( %) are dead. of the deaths ( %) were related to tma ( tma, tma +gvhd, and tma+infection). patients with or more warren criteria in pathological review had poor outcome compared with patients less than criteria ( % alive vs % at months, p= . ). conclusions: intestinal tma is a life-threatening underdiagnosed entity. only patients of patients were diagnosed of intestinal tma. we found that most of our patients had endothelial damage in the gastrointestinal biopsy pathological reviews. gvhd histological criteria were present in most of the patients, mainly histological grade i-ii. prognosis of these patients is poor and pathologist effords in diagnosed the entity is guarranted. disclosure: nothing to disclosure p strategies to reduce neutropenic fever and hospital readmission in multiple myeloma patients managed at home after autologous stem cell transplantation background: neutropenic fever (nf) is the most frequent cause of readmission in the outpatient autologous stem cell transplantation (asct) programs. in our at home model for multiple myeloma patients, we added primary prophylaxis with ceftriaxone, decreasing the incidence of fever during aplasia phase from % to . %. the aim of this study was to analyze the addition of two strategies to reduce the non-infectious nf: withdrawal of g-csf and the addition of primary prophylaxis for engraftment syndrome with corticosteroids after asct. methods: between january and august myeloma patients were managed at-home since day + of asct. all were conditioned with mel . all patients received prophylaxis with quinolone, fluconazole, aerolized pentamidine, low-dose acyclovir (hvs+), and ceftriaxone (since day + ). the patients were classified into groups: group a (n= ; g-csf since day + without corticosteroid), group b (n= ; no g-csf and no corticosteroid), group c (n= ; no g-csf with prednisone . mg/kg/day since day + until granulocyte recovery). first-line therapy at home of nf was piperacillin-tazobactam . g/ h i.v. using a portable intermittent infusion pump. fever was an indication of immediate medical consultation and those patients presenting signs of focal infection or severe sepsis were admitted. other indications for readmission were: willingness of the patient or caregiver, uncontrolled nausea, vomiting or diarrhoea, and mucositis requiring total parenteral nutrition or i.v. morphics. results: the main characteristics of the patients and outcomes are shown in table . there were no differences between groups regarding age, gender, immunological subtype, response before asct, hct-ci, and cd cell dose infused. there were more patients with advanced disease (iss iii) in group c compared to group a ( . % vs. . %; p= . ). the duration of neutropenia was longer in those groups that did not receive g-csf (a: days, b: days, c: days; p< . ). comparing group a with group c, we observed that the incidence of nf and the readmissions rates were lower in group c (nf: . % vs. . %; p= . ; relative risk reduction: . , and number needed to treat . ; readmissions: . % vs. . %; p= . , respectively). the -day cumulative incidence of nf were . % in group a, . % in group b, and . % in group c; p= . . the non-administration of g-csf with the addition of prophylactic corticosteroid did not modify the incidence and grade of mucositis, the first day and duration of fever, nor the number of bacterial infections documented. in the multivariate analysis, this combination (no g-csf with corticosteroid) maintained its protective effect for the development of nf and hospital readmission (or . ; p= . and or . ; p= . , respectively). conclusions: the non-use of g-csf and the addition of prophylactic corticosteroid in mm patients managed at home after asct minimize the incidence of non-infectious fever and optimize hospital resources by reducing hospital readmissions. disclosure: nothing to declare. background: antibody titers to vaccine-preventable diseases decline during the - years after allogeneic hematopoietic stem cell transplantation (hsct) if the recipient is not revaccinated. it is therefore considered best practice to try to offer hsct recipients the same level of protection against all vaccine preventable diseases as the general population. few data in the literature are available concerning vaccine-related problems in hsct recipients. we performed a farmacovigilance evaluation in a cohort of allotransplanted patients followed in our clinic during a year period. methods: from october to november we administered a list of recommended vaccines to hsct recipients attending our routine out patient clinic who fulfilled the following criteria: cd t cells> /μl, cd b cells> /μl, anti-cd antibody infusion> months, ivig therapy> months, no active and severe graft-versus-host-disease (gvhd), no chemotherapy or biological therapeutic agents on going. vaccines suggested were influenza, pneumococcal conjugate (pcv ), polio (inactivated polio vaccine), diphteria, tetanus, acellular pertussis, hepatitis b, hepatitis a, haemophilus influenzae type b, meningococcal quadrivalent (mcv ), human papillomavirus, meningococcal b, measles-mumps-rubella (mmr), varicella. live vaccines (mmr and varicella) were not recommended before years after hsct and in patients with chronic gvhd. all the patients were asked to take the list to the local health facilities in order to have the vaccines injected and a vaccination table arranged with the doses already received and those to receive. we checked the vaccination tables at each visit and monitored potential side effects and gvhd status at , , and months after the first vaccine injection. results: twenty-nine out of patients were evaluable (table ), without gvhd and with chronic gvhd ( mild, moderate, severe). median time after hsct was months ( - ). median number of vaccines received was ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) . as regards patients without chronic gvhd, out of experienced fever after vaccine injections; out of developed transient mild reduction of platelet count; patient reported headache and otalgia after vaccine injection, while another one transient joint pain; out of patients presented signs of mouth chronic gvhd (score nih) and transaminase increase (grade according to world health organization toxicity scale) months after the first vaccine dose, so that cyclosporine dose had to be augmented. as regards patients with chronic gvhd, out of experienced fever after vaccine injections; patients with mild chronic gvhd of the mouth presented hepatic flare two and three months after the first vaccine dose, respectively. in both cases a new increase of cyclosporin and methylprednisolone doses determined progressive normalization of liver enzymes. conclusions: these data show that vaccines were globally well tolerated in hsct recipients, even when they suffered from chronic gvhd. however, close monitoring is warranted in order to better evaluate possible vaccine side effects in this setting of patients. background: allogeneic hsct improves survival for aml patients over the age of years of age when compared to chemotherapy alone. the haematopoietic stem cell transplantation comorbidity index (hct-ci) and ebmt score predict for non-relapse mortality and overall survival, yet little is known about whether qol is preserved in this patient group and whether hct-ci and other performance scores pre-bmt correlate with qol post allo-hsct. methods: we conducted a retrospective analysis of patients years and older who underwent ric allo-hsct at the university hospital of wales, cardiff between september and december (n= ). hct-ci, karnofsky performance score (kps) and ebmt scores were calculated prior to transplant and qol measured using the fact bmt (version ) questionnaire, which was completed at , and months post transplant. patients were grouped at the -, -and -month time points for each of the different performance indices, allowing group comparison against compound sub scores using the mann-whitney u test. results: patients were included in this study, with median age years (range - ). patient characteristics, including conditioning, donor type, pre-transplant hct-ci and kps scores are summarised in table . the year and year overall survival (os) for the patient cohort was . % and . % respectively. hct-ci of ≥ vs was significantly associated with poorer bmt-related qol domains at months (p= . ) and general qol domains at months (p= . ) post-transplant. while ebmt score showed no correlation with qol parameters, patients with kps of vs ≤ showed significant differences in both general (p= . ) and bmt-related qol (p= . ) at months and in all qol domains at months (symptomrelated qol p= . , general qol p= . , bmt-related qol p= . ). importantly neither the hct-ci nor the kps pre-transplant predicted for qol at months post transplant. conclusions: patient selection is key to ensuring maximum benefit from allo-hsct both in terms of overall survival but also with regards to qol and survivorship. we note that while patients with hct-ci ³ or kps ≤ had significantly poorer qol at months post allo-hsct, qol was recovered by months post transplant, with this significant difference no longer seen. our data shows that in selected aml patients over the age of years with good performance status and low comorbidity index, a favourable outcome can be achieved with good qol maintained throughout the post transplant period. background: advances in allosct technology, supportive care, and use of reduced intensity conditioning regimens for older patients have led to significant improvements in longterm survival after transplant. the survivors have an elevated probability of late morbidity and mortality, including abnormalities in phosphocalcic metabolism and bone disease. rapid and progressive bone loss occurs within the first - months after transplant, and this is followed by a slow process of recovery, with bone loss persisting for to months. bone fractures can worsen the quality of life of allosct survivors, but the real burden of the disease is unknown. the objective of the study is to ascertain the prevalence of bone pathology and vertebral fractures early after transplant in our center. methods: this is a retrospective and observational study. forty-nine patients ( male/ female, median age y, range - ) that underwent allosct were included in the study in the period of to months after transplant (may -december ). pre-and post-transplant risk factors associated with bone disease were recorded: age > years, female sex, menopause, hormone replacement therapy, previous treatment with steroids, previous fractures, weight < kg, bmi < - , low physical activity, low calcium intake, smoking, alcohol intake, and history of femoral fractures in parents. in all patients laboratory data (including serum calcium, -hydroxyvitamin d, and pth), lumbar and femoral bmd (dxa), and spinal x-ray were also evaluated. a vertebral fracture was defined as a reduction of > % in the anterior, middle or posterior high of the vertebral body. results: we identified vertebral fractures in ( %) patients. five patients had fractures prior to transplantation, and patients presented "de novo" vertebral fractures following transplantation; therefore, the prevalence of "de novo" postransplant fractures was / ( %). most ( %) of these fractures were asymptomatic at the time of diagnosis. most patients ( %) with vertebral fractures had > pre-sct risk factors (median risk factors pre-sct , range - ), the most frequent being low calcium intake, steroid exposure, presence of previous fractures, and menopause. those patients with fractures and less than risk factors pre-tph, added new risk factors after transplant, mainly steroid treatment. forty-four patients ( %) had vitamin d insufficiency (< ng/ml), ( %) had osteopenia and ( %) had osteoporosis. vitamin d insufficiency and bone disease were more frequent in women than in men ( % vs. % for vitamin d, % vs. % for osteopenia, % vs. % for osteoporosis, and % vs. % for vertebral fractures, respectively). conclusions: the prevalence of post-transplant bone disease and vertebral fractures in our series is high. most fractures appearing "de novo" after allosct were asymptomatic and were diagnosed by x-ray. patients who presented vertebral fractures frequently had more than risk factors identified pre-sct. patients undergoing allosct should have their bone health assessed early in their treatment and, if indicated, should start preventative therapy to avoid bone loss and fractures. other measures such as physical exercise, vitamin d and calcium supplementation, and dxa and spinal x-ray at baseline and following transplantation are also highly recommended. disclosure: maría suárez-lledó received a grant from dkms-spain foundation. other authors have nothing to declare the use of g-csf in selected patients after autologous stem cell transplantation is associated with low incidence of engraftment syndrome background: the use of g-csf after autologous stem cell transplantation (asct) accelerates neutrophil recovery, however it has been related to an increased risk of engraftment syndrome (es) development in some studies. for this reason, we do not routinely prescribe g-csf after asct and we only use it in patients with significant complications (enterocolitis, severe sepsis, atrial fibrillation) after stem cell infusion. the main objective of this study is to evaluate the incidence of es in patients who receive asct for monoclonal gammopathies (mg), non-hodgkin lymphoma (nhl) and hodgkin lymphoma (hl) and receive g-csf only if needed. as secondary objectives we evaluate differences in the engraftment day as well as the length of inpatient stay. methods: we retrospectively analyzed patients with mg or lymphoma, who underwent asct conditioned with high dose melphalan ( - mg/m ) or beam, respectively, between and in our center. specific clinical features for es according to spitzer and maiolino criteria were evaluated between days before and days after the engraftment. statistical analysis was performed with spss v. . . results: thirty-one patients with mg and patients with lymphoma were analyzed. median age at transplant was . years ( . - . ) and patients ( . %) were male. median prior lines of treatment in patients with gm or lymphoma were ( - ) and ( - ), respectively. table shows patients´characteristics. mobilization with g-csf ± plerixafor was performed in patients ( %) and chemotherapy + g-csf ± plerixafor in patients ( %). median cd x /kg cells infused was . ( . - . ). eleven patients ( . %) received g-csf, due to infection ( enterocolitis, listeriosis, acute hepatitis, septic shock) and because of atrial fibrillation or fibrilloflutter. median time from sct to first day of g-csf was days ( - ) and median time on g-csf treatment was days ( - ). patients who received g-csf showed a short time to neutrophil engraftment (≥ . x /l), days vs. days, p< . but longer duration of hospitalization, days vs. days, p = . . non-relapse mortality at day + , + and + was %. es was diagnosed in ( . %) patients, amyloidosis, multiple myeloma and plasmablastic lymphoma. there was not statistical difference in the incidence of es between patients who received g-csf ( . %) and patients who did not ( . %), p= . . analyzed by disease, es appeared in of patients who received g-csf in the lymphoma group ( . %) but none of the patients with mg that received g-csf developed it. we did not find statistical differences between patients who developed es and those who did not in age ( years vs. years, p= . ), length of hospitalization ( days vs. days, p= . ) and the number of cd x /kg cells infused ( . vs. . , p= . ) . conclusions: the use of g-csf in selected patients is associated with low incidence of es. our study confirms that the use of g-csf accelerates neutrophil recovery but it is unclear if it can increase the incidence of es, especially in patients with lymphoma. [[p image] . background: graft failure is one of the top- problems of allo-hsct (after gvhd and relapse). the problem of graft failure becomes more significant due to increasing number of allo-hsct with ric conditioning regimen from haploidentical and hla-mismatched unrelated donors. role of t cells in graft failure is well known. here we report an impact of t-memory cell subsets count before antithymocyte globulin (atg) administration on primary graft failure after allo-hsct. methods: sixteen patients with acute leukemia transplanted in national research center for hematology were included on this prospective study. all patients received horse atg at dose mg/kg/day from day - to - before allo-hsct as gvhd prophylaxis and were balanced by other factors that could affect engraftment. detailed patients characteristics are listed in table . peripheral blood samples were collected on day - before allo-hsct (before atg injection) in edta-tubes. flow cytometry analysis was performed on bd facs canto ii (becton dickinson, usa) to define t-memory subsets: t-naive and t-stem cell memory (tnv+scm) -cd r -ccr +cd +; t-central memory (tcm) -cd r +ccr +cd +; t-transitional memory (ttm) -cd r +ccr -cd +; t-effector memory (tem) -cd r +ccr -cd -; t-terminal effector (tte) -cd r -ccr -cd -, among cd + and cd + t-cells . sysmex xe- was used to calculate absolute count of different t-cell subsets. mann-whitney u test was used for nonparametric data analysis between two groups. fisher's exact test was used for x tables. p-value less than . was considered statistically significant. results: an influence of t-memory cell subsets count before atg administration on primary graft failure is shown in figure . according to our data high absolute number of cd +ttm and cd +tte is associated with primary graft failure. conclusions: based on these findings high absolute number of cd +ttm and cd +tte could be one of the prognostic factors of primary graft failure after allo-hsct. optimizing atg dose due to recipient absolute t-memory cell subsets count before atg administering may prevent graft failure and improve posttransplant results. background: upper gastrointestinal graft-versus-host disease (gi gvhd) has been an increasingly recognised entity following allogeneic stem cell transplantation (sct). budesonide, widely used in inflammatory bowel conditions, has also been found beneficial in gi gvhd. the major benefit of budesonide is attributable to its poor absorption and extensive first-pass metabolism via cytochrome p (cyp) a , which translates to less systemic steroid-related effects. however, transplant patients are often exposed to multiple drugs, among which some agents act as cyp a inhibitors and therefore can increase budesonide bioavailability and might lead to systemic toxicity. azole antifungal drugs are probably the most common concomitantly used cyp a inhibitors in transplant recipients. methods: we reviewed allogeneic sct records for patients treated with oral budesonide for gi gvhd at our transplant centre between and retrospectively. the aim of the work was to assess the development of adrenal suppression with or without clinical features of iatrogenic cushing`s syndrome. the standard dose of budesonide was mg three times a day. patients receiving prednisolone or other glucocorticosteroids and those with no available serum cortisol level measurements were excluded. results: our analyses identified four allogeneic sct patients in whom adrenal suppression was diagnosed with undetectable serum cortisol levels during oral budesonide treatment. of these patients two developed iatrogenic cushing`s syndrome and both patients were treated with cyp a inhibitors concomitantly: . clarithromycin and fluconazole; . clarithomycin and voriconazole. the development was rapid (within and weeks). symptoms included morphological features such as moon face, high blood pressure, weight gain, peripheral oedema and proximal myopathy. symptoms resolved gradually following cessation of azole antifungal agents and on gradual weaning of budesonide. conclusions: although single agent budesonide treatment given for gi gvhd is rarely associated with systemic side effects, patients on azole antifungal drugs and macrolide antibiotics are at higher risk of systemic toxicity due to drug interactions. patients who are allergic to penicillin and receive macrolide-based prophylaxis can be especially vulnerable. to our knowledge the number of cases reported in literature about systemic effects of oral budesonide in transplant recipients is less than . our observation supports previous reports on the potential of oral budesonide to induce systemic effects. we therefore advise careful monitoring of patients treated with budesonide in combination with cyp a inhibitors, including antimicrobial agents routinely used in sct. disclosure: none implemented strategies to overcome barriers in the establishment of a consolidated hematopoietic stem cell transplantation program in a developing country background: the national institute of medical sciences and nutrition "salvador zubiran" is a national health institute located in mexico city. although mexico is considered an upper-middle income country, more than % of the population lives in poverty without health care coverage and patients within this social stratum are referred to our institution. the first hematopoietic stem cell transplantation (hsct) in mexico was performed at our institution in . from this year until , hsct were sporadically performed (n= ), showing a poor overall survival (os) and high non-relapse mortality (nrm). these outcomes resulted from an unstructured hsct program, limitedresources, patient low socioeconomic status, and paucity of population-adapted procedures. in , according to these results, a decision to establish a hsct program was made. therefore, in order to set up a successful hsct program, implementation of financial and medical strategies were necessary. the objectives of this study were to describe the barriers and implemented strategies for the establishment of a hsct program in mexico along with the outcomes of patients undergoing this procedure throughout the reorganization of the program. methods: this study is a health services research. barriers were detected based on the results of the hsct program from - (not shown). table shows the financial, medical, and research strategies that were implemented for each barrier. results: from november to november , hsct have been performed in patients at our institution. most hsct were autologous (n= , %). forty one patients underwent hsct. from the patients, most were males (n= , %) and the median age was . years (range, - ). the most frequent underlying diseases for auto-hsct were lymphomas (n= , %), non-seminomatous germ cell tumors (n= , %), and multiple myeloma (n= , %). acute leukemias (n= , %), aplastic anemia (n= , %), and myelodysplastic syndromes (n= , %) were the most frequent diagnosis for patients undergoing allo-hsct; and acute leukemia was the most frequent diagnosis for patients undergoing haploidentical hsct (n= , %). acute and chronic gvhd were present in % (grades i-ii %) and % (limited %), respectively. for allo-hsct, , day, and -year nrm was . %, %, and %, respectively; and -day nrm in auto-hsct was . %; year os was % and % for auto and allo-hsct, respectively. conclusions: future perspectives of the hsct program include the acquisition of funds for unrelated donors; to improve outcomes of patients undergoing haploidentical hsct, and to increase the number of in-patient rooms. we conclude that despite paucity of resources and other limitations, the implementation of financial, medical, and research strategies have shown that barriers can be effectively overcome in a developing country in order to establish a consolidated and nationally renowned hsct program, providing good outcomes for patients. disclosure: none of the authors have any conflict of interest to disclose. the effect of protective buffering on daily stress and relationship quality in dyads following hematopoietic stem cell transplantation: results from daily process methodology malgorzata sobczyk-kruszelnicka , aleksandra kroemeke , zuzanna kwissa-gajewska , sebastian giebel background: cancer-related support communication (e.g., protective buffering) may impact the risk for psychological and relationship distress in patients following hematopoietic stem cell transplantation (hsct) and their caregivers. previous studies have revealed that protective buffering (i.e., hiding one's concerns and denying one's worries) has mixed effects: is beneficial (for "protected" person), costly (especially for the person using it), or unrelated to dyadic wellbeing. there has been, however, little evidence linking dyadic protective buffering with distress using daily process methodology. we assessed ( ) the relationship between daily protective buffering, and same-and next-day stress and relationship quality in patient-caregiver dyads following hsct and ( ) whether similarity or complementarity in protective buffering between dyads is adaptive. methods: two hundred patients (after first autologous or allogeneic hsct) and their caregivers (spouse or another relative) independently completed measures of daily protective buffering, daily relationship quality, and daily stress for consecutive evenings after patients´hospital discharge. actor-partner-interdependence model (i.e., both partners' and caregivers' reports regarding support communication and distress were studied) was used to test study hypotheses. results: for both patients and caregivers, multilevel structural equation modeling showed a significant positive relationship between daily protective buffering and sameday relationship quality. association of protective buffering with same-day stress level was negative. in next-day analyses, patient-reported protective buffering was related to patient's higher relationship quality, whereas caregiverreported protective buffering increased patient's daily stress. complementarity in protective buffering was related to higher immediate same-day relationship quality for both patients and caregivers, while benefits from similarity have delayed effects, although only in patients. conclusions: contrary to previous studies, protective buffering rather has a beneficial effect in dyads following hsct. protection of the partner and relationship against revealing negative emotions and powerlessness was not related to costs in both parties. the findings suggest that the effect of daily protective buffering in dyads following hsct depends on support timing (same-or next-day effect) and differs for both parties. patients seem to benefit the most from the similarity in protective buffering, while caregivers from complementarity. the "fit" between patient and caregiver in support communication ought to be taken into consideration in the practical approach. disclosure: nothing to declare. virus reactivation and low dose of cd + cell were associatied with secondary poor graft function within the first days after allogeneic stem cell transplantation yuqian sun , xiao-jun huang background: secondary poof graft function (spgf) was defined as the secondary cytopenia after initial engraftment of hsct. it was shown to be associated with poor prognosis, however there are very few reports on the incidence, risk factors and outcomes of spgf. methods: patients who received transplantation from peking university people's hospitial during january, to december, were retrospectively reviewed if they fulfilled the following conditions: ( ) diagnosed with acute leukemia or myelodysplastic syndrome; ( ) received allo-sct from either matched sibling donor (msd) or haploidentical related donor (hid). pgf was defined as persistent neutropenia (≤ . × /l), thrombocytopenia (platelets ≤ × /l), and/or hemoglobin ≤ g/l for at least consecutive days, transfusion-dependence, associated with hypoplastic-aplastic bone marrow (bm), and complete donor chimerism without concurrent graftversus-host disease (gvhd) or disease relapse. primary pgf was defined as the failure to achieve initial engraftment by days after transplantation, while secondary pgf was defined as the fulfillment of the criteria after initial engraftment hsct. results: during january, to december, , patients who received transplantation from peking university people's hospitial were retrospectively reviewed. among the patients who achieved initial engraftment, patients developed spgf. the cumulative incidence of spgf on day was . %. the median time of secondary pgf was . ( - ) days after transplantation. low (< median) cd + cell dose (p= . , hr . ( %ci, . - . )), ebv reactivation (p= . , hr . ( %ci, . - . )) and cmv reactivation (p= . , hr . ( %ci, . - . )) were identified as independent risk factors with spgf. there is no significant difference of pgf incidence in msd group and hid patients (p= . ). the overall survival of patients with spgf at year after transplantation was significantly poor than patients with ggf ( . % versus . %, p< . ). conclusions: in conclusion, spgf develop in . % patients after allo-sct, especially in patients with cmv, ebv reactivation or infused with low dose of cd + cell. the prognosis of spgf is still poor due to lack of standard treatment. disclosure: there is no conflict of interet thiotepa with treosulfan and busulfan based conditioning are significantly more gonadotoxic than treosulfan previous studies suggest that busulfan results in long-term gonadal toxicity. no previous studies have compared gonadal toxicity outcomes after treatment with busulfan with treosulfan, a newer agent with similar marrow toxicity to busulfan but with reduced non-marrow toxcitiy. our aim was to determine whether there are differences in pubertal and fertility outcomes in paediatric patients treated with treosulfan compared with busulfan. methods: inclusion criteria were patients who had received either busulfan or treosulfan or treosulfan with thiotepa, only one hct and were aged years and above in august . eligible patients were reviewed in clinic as part of their routine follow-up, thus research ethical approval was not required. follice stimulating hormone, luteinising hormone, oestradiol, and pubertal history were noted. ovarian reserve was estimated in female patients by measuring serum anti-mullerian hormone (amh). male patients had serum testosterone measured and were also offered semen analysis. results: thirty-five patients met the inclusion criteria, of which twenty-five wanted to be reviewed ( %); seventeen females and eight males. mean age at hct was years, mean age at review was years and mean years since hct was years. female patients treated with busulfan or treosulfan with thiotepa (n= ) had minimal amh and none of these patients were having regular periods. females treated with treosulfan (n= ) had normal amh and regular periods without needing hormone replacement. only four male patients opted for a semen analysis and all had significantly reduced sperm counts. conclusions: our results suggest that females treated with treosulfan have minimal (if any) reduction in ovarian reserve compared to other conditioning regimens which casue significant compromise. although this was a small study, and thus not suitable for statistical analysis, the clinical findings are marked. future studies should further investigate optimal doses of treosulfan that could be used to achieve bone marrow engraftment and limit long-term effects on fertility. disclosure background: autologous and allogenic hematopoietic stem cell transplantation (hsct) are potentially curative treatments for hematological malignancies. patients with related complications may need admission to the intensive care unit (icu) for specific therapy and organ support. mortality risk factors, supportive care and principal causes of admission in icu are described in our cohort of patients (pts). methods: we retrospectively studied pts, male, with a median age of , years (range: - ) who underwent allo-hsct in our center between july and october . two hundred and twenty-seven( , %) pts received autologous hsct (auto-hct) and ( , %) allogenic hsct (allo-hsct); from unrelated donor, from identical sibling, and the remainder, mismatched related donor . twenty-three ( , %) out of pts were admitted in the icu in the transplant procedure admission. results: fifteen ( , %) out of pts were male with a median age of years (range: - ). patients' baseline diseases were: multiple myeloma ( , %), non-hodgkin´s lymphoma ( , %), hodgkin´s lymphoma ( , %), acute lymphoblastic leukemia ( , %), myelodisplasic syndrome ( , %), solid tumor ( , ) and acute myeloblastic leukemia ( , %). fifteen ( , %) pts received auto-hsct, ( , %) allo-hsct from unrelated donor, ( , %) allo-hsct from identical sibling, and the remainder haploidentical hsct ( ) ( , %). so, , % of auto-hsct pts and % of allo-hsct were admitted in the icu. the median stay in the icu was days (range: - ) and reasons for admission were: respiratory insufficiency ( , %), septic shock ( , %), renal insufficiency ( , %) and multi-organic failure ( , %). twenty-one ( . %) pts required respiratory support with: nasal cannula or oxygen mask (c/m) ( %), non-invasive mechanical ventilation (nimv) ( , %) and invasive mechanical ventilation (imv) ( , %). fourteen ( %) pts needed inotropic agents for shock treatment. finally, ( , %) pts required substitutive renal therapy with hemodialysis or haemofiltration (hd/hf). eleven ( , %) out of pts died, ( , %) were male with a median age of years (range: - ). ten of them ( , %) needed imv and were treated with inotropic agents. all patients who required hd/hf (n= ) died. imv and treatment with inotropic agents were associated with icu mortality (or , ; p= , , or ; p= , ; respectively). conclusions: in our series of pts, , % needed admission in the icu, presenting a mortality rate of % approximately. there were no differences in the prevalence of icu admission regarding hsct donor. main reason for admission was respiratory failure with imv requirement in , % of pts. imv and treatment with inotropic agents were associated with icu mortality. an early identification of pts at risk of icu admission could have a beneficial impact on survival improvement disclosure: nothing to declare is there any association between thrombotic risk factors and veno-oclusive disease in childhood allogeneic hematopoietic stem cell transplantation? background: veno-oclusive disease (vod) is a major complication of hematopoietic stem cell transplantation (hsct). in some studies levels of fibrinolytic factors especially plasminogen activator inhibitor- (pai- ) level were found associated with vod. however, little is known about the relationship between thrombophilia risk factors and vod. in this study we aimed to investigate association of major thrombophilic gene mutations on vod in pediatric hsct patients. methods: we reviewed retrospectively patients with vod who underwent hsct between - in ankara pediatrics and pediatric hematology-oncology training and education hospital, bone marrow transplantation unit, turkey. fifty-one patients who did not develop vod and transplanted during the study period were accepted as control group. we evaluated plasma homocysteine and lipoprotein a level, protein s and c activity and antigen levels and factor v g a mutation, prothrombin g a mutation, methylenetetrahydrofolatereductase (mthfr) c t and a c mutations, plasminogen activator inhibitor- - g/ g polymorphism before hsct. we also evaluated the patients' hospital files and noted the demographic values and complications of hsct. statistical investigations were done with spss statistics . for windows and p< . has been accepted as significant. results: there was no difference between control and vod groups as regard to age, sex, diagnosis, donor type, conditioning regimen, hsc source, and hla typing . there was no difference between the groups according to homocysteine, lipoprotein a, protein s and c activity and antigen levels. we did not find any relation between the genetic variations of thrombophilia and vod (table ). in vod group there were patients ( . %) with acute graft versus host disease (agvhd) and in control group there were ( . %) patients with agvhd (p= . ). febrile episodes were more frequent in vod group compared to the controls (respectively; n= , . % vs. n= , . %, p= . ). -year overall survival was % . in vod group and % in control group (p= . ). disease free survival was also different between vod and control groups (respectively; . % vs. . %, p= . ). conclusions: in literature there are recent studies showing higher pai- levels in patients with vod. however, in our study we did not find any relationship between congenital thrombophilia factors and vod. new studies with larger sample groups is necessary to better evaluate the association of congenital thrombophilia factors and vod. disclosure: nothing to declare p different strategies of chemotherapy-induced nausea and vomiting (cinv) prevention in hematological patients receiving an autologous hematopoietic stem cell transplantation: a single center experience ilaria cutini , riccardo boncompagni , chiara nozzoli , antonella gozzini , stefano guidi , chiara innocenti , massimo di gioia , lorenzo tofani , riccardo saccardi background: despite the improvements of pharmacological control, cinv still represents a major problem in patient undergoing hematopoietic stem cell transplantation (hsct). we present here a comparison of two pharmacological strategies for preventing cinv in multiple myeloma (mm), hodgkin (hl), and non-hodgkin lymphoma (nhl) patients who received an autologous hsct in our institution. methods: from january to july , we retrospectively analyzed consecutive patients, median age years ( - yo) , diagnosed with mm, hl, and nhl, who underwent an autologous hsct following a melphalan mg/sqm and beam/feam condition regimens, respectively. the first patients received cinv prophylaxis with palonosetron i.v and dexamethasone mg die (regimen a), whilst the following were administered with fosaprepitant iv, ondansetron iv and dexamethasone mg die (regimen b) both cinv prophylaxis was administered the day of melphalan infusion (day - form transplant). emesis breakthroughs were treated with alizapride and metoclopramide. nausea and vomiting were assessed through the ctcae . score system. categorical variables were compared with pearson chi-square test. results: the overall incidence of nausea was %, ( % grade , % grade , and % grade , respectively). in regimen a was shown to be %, ( % grade , % grade , and % grade , respectively) while in regimen b was % ( % grade , % grade , and % grade , respectively). pearson chi-square test did not show any differences between the groups (p= . ). the overall observed vomit was % ( % grade , % grade , and % grade ). in regimen a it was ( % ( % grade % grade , and % grade ), and % in regimen b ( % grade and % grade ). conditioning regimens didn't' have any significant impact on either nausea or vomit. patientsyounger then median ( yrs), were reported to have higher incidence of both nausea, (p= . ) not related to cinv treatment, and vomit ( % vs %, p= . ). in multivariate analysis the overall incidence of nausea is related to age (younger patients have higher probability to develop nausea (or , ; p= , ) whilst the higher incidence of vomit is related to: regimen a (or . ; p< , ), previously reported nausea (or , ; p< , ), and no smoking habits (or , ; p= , ). conclusions: both regimens are equally effective for nausea control however regimen b evidenced a better vomiting control. this finding is particularly relevant when the center policies include an early discharge program, therefore improving both patient's quality of life and procedure cost-effectiveness. clinical background: patients who underwent an allogeneic hematopoietic cell transplantation (hct) are challenged by medical, psychological and social complications. support groups might help hct-survivors to cope with these challenges. however, the existing literature about post-hct support groups is scarce. moreover, data on professionallyfacilitated support groups do not exist. the aim of this project was ( ) to establish a professionally-facilitated support group and ( ) to assess the discussed topics. methods: from / until / all patients who received an allogeneic hct at the adult stem cell transplantation program of the university clinic mannheim were invited to participate in a professionally-facilitated support group. additionally, spouses and life partners were invited. a theologian who is also a physician served as facilitator. he had no further function within the transplant team. the format of the group was unstructured without any rules regarding regular attendance. the facilitator did not provide topics or a curriculum. during the first year the group met every days followed by a monthly schedule. from the fifth until the th meeting the attendance and the discussed topics were minuted by the facilitator. the content of the minutes was analysed by a combination of an inductive and a deductive approach. all participants provided their informed consent for the study. results: altogether patients (female: n= ; male: n= ) and spouses/life partners (female: n= ; male: n= ) participated. patients ( %) and spouses ( %) attended more than one meeting. among those who participated in ≥ meetings the median time of participation was months. the median count of participations was eight. % of the participants attended the meetings longer than one year, % longer than three years. there was no sex difference with respect to the frequency and the duration of participation. however, the frequency of participation decreased significantly the longer a participant was attending the meetings. during group meetings the facilitator recorded thematically different contributions to the discussions divided in distinct topics. these topics were grouped into main categories [(a) medical topics, (b) private life and environment, (c) human relationships, (d) physical and mental condition and (e) the support group itself] and eight further categories [( ) compliance, ( ) economic issues, ( ) religion, ( ) sexuality, ( ) death and dying, ( ) support and coping, ( ) objectives and needs and ( ) not otherwise specified issues] which could not be grouped in one of the main categories. the most frequent issues were medical topics ( %), human relationships ( %), physical and mental condition ( %), private life and environment ( %), financial issues ( %), the support group itself ( %), support and coping ( %) and objectives and needs ( %). noteworthy, death and dying ( . %) were rare topics and sexuality was never mentioned. conclusions: to our knowledge, this is the first prospective and systematic analysis of a professionallyfacilitated support group for hct-survivors. these data might help to establish support groups and to identify psychosocial needs of patients and targets for specific support. disclosure: nothing to declare background: endothelial damage is associated with inflammatory complications that appear early after hsct, such as sinusoidal obstruction syndrome or acute gvhd. engraftment syndrome (es) is an inflammatory condition diagnosed by maiolino clinical score. potentially, es can exhibit high morbidity and mortality, especially after autologous-hsct in multiple myeloma (mm) patients since the introduction of new drugs such proteasome inhibitors and immunomodulatory drugs (imids). the objective of the present study was to evaluate if es is associated with endothelial dysfunction in patients with mm who underwent auto-hsct. methods: we included six patients with mm who received induction treatment including new drugs and consolidated their response with an autologous-hsct. we analysed comparatively the effect of incubating endothelial cells in vitro with serum samples from patients with es vs. no es. serum samples were collected before (pre), and after , , and days from the transplant. an additional sample was collected at the es onset and at the discharge day (no es group). endothelial cells (hmec) in culture were exposed to media containing % of serum from each patient for h. cell growth was controlled morphologically. expression of the adhesion receptor icam- on the cell surface was analysed by immunofluorescence, and activation of the inflammation related p- mapk signalling pathway was evaluated by sds-page and western blot. results: exposure of hmec monolayers to sera from patients who developed es (onset day, n= ) resulted in an increased icam- expression on the cell surface, higher that the observed with sera from patients who did not develop es (discharge day, n= ) ( . % of labelled area vs. . %, respectively). in addition, in experiments with sera from patients not developing es, icam- expression on cells exposed to sera from day + was reduced with respect to the observed with sera from day + , probably due to the corticosteroid used as a prophylaxis in our centre. this reduction was not observed in es patients. regarding phosphorylation of p- , it was significantly higher in cells exposed to sera from es patients than in response to sera from patients who did not develop es. conclusions: the increase in the expression of the adhesion receptor icam- on the surface and the intracellular activation of p mapk in endothelial cells exposed to sera from patients developing es indicates the existence of endothelial activation in association with es. interestingly, the prophylaxis of es with corticosteroid seems to be less effective in patients who developed es than in patients who did not develop this complication. these results need to be validated in a higher number of patients and modifications in additional markers of endothelial dysfunction should be investigated. disclosure: gonzalo gutiérrez-garcía: honoraria from gilead. grant from jazz pharmaceutical the other authors do not have any disclosure to comment. p association between uric acid levels before and after allogeneic haematopoetic stem cell transplant and transplant outcomes: a single centre experience background: uric acid (ua) is a known endogenous danger signal which activates the nod-like receptor protein (nlrp) inflasome.ua is released from injured cells during conditioning in allogeneic stem cell transplantation (hsct). a pre-clinical study has demonstrated that nlrp inflasome-mediated il- production regulates graft-versushost disease (gvhd). the ua role in inflammation and gvhd is unclear. there are discordant reports in the literature about a potential protective role of ua on gvhd after a hsct. methods: we performed a retrospective study to assess the association between serum ua levels pre-and post- table] . table ] results: the characteristics of the patients are shown in table . median age was years (range - ), and patients ( %) were male. twenty-seven patients ( %) received low doses atg as part of gvhd prophylaxis. allopurinol was from the day before start of conditioning therapy until day . the median levels of ua were , mg/ dl before conditioning, , mg/dl at day , , mg/dl at day + and , mg/dl at day + . there was no impact between the ua levels and os at any time of the hsct. ua levels at day + were associated with a higher ci relapse at years ( % [ % ci, - %] for ua level > , mg/dl, and % [ % ci, %- %] for ua level ≤ , mg/dl [p= , ]). there was a trend for a higher ci of grade ii-iv agvhd for the subgroup of patients not treated with atg with ua < , mg/dl ( % vs %; p= , ) on day - and a higher nrm with ua < , on day ( % vs %; p= , ). conclusions: in our study the ua levels showed no impact on os, and only a tendency for ci of grades ii-iv agvhd grades ii-iv and nrm for the subgroup of patients not treated with atg. surprisingly, high levels of ua at day + of hsct were associated with a significant higher incidence of relapse. disclosure: dkms foundation, pi / (instituto carlos iii) and sgr (grc), generalitat de catalunya. background: veno-occlusive disease (vod) is an early, uncommon but serious complication of stem cell transplantation (sct) that is associated with high morbidity and mortality. defibrotide is the only licensed treatment for vod, and time to start of treatment (tst) affects outcomes. minor differences exist between the seattle, baltimore and classical ebmt ( ) criteria, which may trigger different start points for treatment. late onset vod (> days) is less recognised and we hypothesize, may have worse outcomes with longer time to diagnosis, and more limited treatment options across different healthcare systems. methods: electronic patient records from sept. -oct. at king´s bmt centre and pharmacy databases were reviewed, timepoint to clinical and bio-chemical manifestation of vod, diagnosis, tst, survival and longterm outcomes were analysed. results: of the patients( . %) who underwent an allogeneic sct, developed vod, including paediatric cases. none of the autologous sct patients developed vod. the paediatric and autologous sct patients were not analysed any further. adult patients (male= ; . %) developed vod at a median age of years(range - ), of whom developed < days and patients had late-onset vod as per ebmt criteria(range - days). cases classed as severe and as moderate vod. patients received defibrotide at diagnosis, patients within days, patients between - days, and patients received treatment after days. overall mortality for this cohort was %( / ). / ( . %) of patients with early onset vod and / ( . %) patients with late-onset vod died. of the deaths, died of liver failure and a further patients had vod as a likely contributing factor in their deaths. patient died with subarachnoid haemorrhage and with relapsed disease. patients that received defibrotide after days, / patients( . %) died, as compared to / ( %) for treatments between - days, / ( . %) for treatments within days. the lone surviving patient who received treatment after days has severe chronic liver disease and it's complications. of the patients who fit seattle criteria for early-onset vod, only fit the baltimore or ebmt criteria for classical vod. of these patients met the baltimore criteria later than the seattle criteria were met(range = - days). conclusions: vod carries high morbidity and mortality, and beyond the known risk factors and with the caveat of limited numbers in this study, we strongly suspect this is further increased when time to definitive treatment with defibrotide is delayed, particularly beyond days. nearly a quarter of cases with vod are late-onset as per classical ebmt criteria. however contrary to our hypothesis, their overall outcomes and mortality do not appear worse, with time to treatment again emerging as a strong predictive factor. conditioning treatment related factors, which play a stronger role in endothelial dysfunction in the hepato-portal circulation, may not be as much at play, perhaps for late-onset disease. uniformity in the use of diagnostic criteria, and high degree of vigilance, even beyond days, leading to early treatments may improve outcomes in vod. disclosure: nothing to declare background: hsct-associated thrombotic microangiopathy (ta-tma) affects - % of patients receiving an allogenic sct, with a high mortality up to - % in severe cases. endothelial injury mediated by complement activation has been atribuited a major role in the pathogenesis, and blockade of c with eculizumab offers promising results. methods: we present our experience with pediatric cases of ta-tma treated with eculizumab. the diagnosis of ta-tma was stablished attending to jodele et al criteria. clinical data were collected retrospectively from medical records. results: all cases were diagnosed between august and april , with a median age of years ( . - ) at time of diagnosis. primary disease was acute leukemia in cases ( all and aml), severe aplastic anemia in , and primary immunodeficiency in . they received their first sct in all cases, from mud and from mmrd (cd ra+ depleted haploidentical grafts), with mac regimen in cases, and ric in cases. of them received calcineurin inhibitors (cyclosporine) as gvhd prophylaxis. all patients developed agvhd (grade or higher in cases). and patients presented viral reactivation. hypertension was present in cases at tma diagnosis, requiring or more antihypertensive drugs in of them. all patients had renal injury consisting of less-than-normal glomerular filtration rate (median of ( - )) and proteinuria, with urine protein-to-creatinine ratio higher tan mg/mg in cases (data not available in patients). serum haptoglobin was decreased in just cases at diagnosis, and schistocytes were detected in patients. cutaneos signs were present in all cases, digestive symptoms in , neurological affection in , and notoriously all of them developed polyserositis. c and c were normal in all cases, with sc b higher than ng/ml in patients and lower in (data not available in cases). all patients received defibrotide as treatment, and cases received also rituximab, associated to therapeutical plasma exchange in . all of them received eculizumab, as first line in cases (median of days between diagnosis and eculizumab start). treatment was correctly monitorized with ch levels in cases (not available quick enough in other ). median number of doses needed in induction therapy was , and median interval between doses was days. patients required reduced interval and higher doses to maintain ch supressed. patients did not respond, and died because of tma. patients had hematological response, with chronic renal injury in of them and resolution of acute renal failure in case. nevertheless patient responding to eculizumab died because of tma related complications, and because of an invasive fungal infection. patients are alive, with a median follow up of months from treatment start. conclusions: our experience supports promising results of eculizumab based treatment for ta-tma, highlighting the importance of an early treatment and a careful therapy monitoring by ch supression. prospective studies are needed to achieve a better knowledge of this pathology and its treatment. disclosure: nothing to declare background: approximately - % of allogeneic hematopoietic stem cell transplant (allo-hsct) are made with some sort of abo blood group system incompatibility. an hsct abo donor-recipient incompatibility implies risks of complications during the process of infusion as acute hemolytic anemia (ah), delayed graft and other later complications due to the presence of isohemaglutinins (pure red cell aplasia or passenger lymphocyte syndrome). also, abo incompatibility could impact on graft versus host disease (gvhd) incidence, and could be associated with not relapse mortality (nrm) and overall survival (os). there are not concluded evidence about the abo incompatibility impact, so the aim of this study was to identify complications and response associated with abo incompatibility in patients undergoing allogeneic hematopoietic stem cell transplantation. methods: a retrospective study was performed on patients who receive an allo-hsct between january and august . two groups were performed according to the presences or not of abo incompatibility. demographic and clinical information was collected from physical and electronic medical records, and information was analyzed in spss v results: sixty-eight patients were enrolled in the study, % male, the median age was years ( - ) with the following diagnoses: acute lymphoblastic leukemia %, acute myeloblastic leukemia . %, granulocytic chronic leukemia . %, myelodysplastic syndrome . %, dendritic cell neoplasm . %, aplastic anemia . %. ninety-one percent of the patients received a transplant from an identical hla donor and . % received a haploidentical transplant. fifty-two patients ( %) were abocompatibility (g ) and patients ( %) had aboincompatibility (g ). none patient with aboincompatibility received a haploidentical transplant. the contrast between groups didn't show differences in fever, infections, bacterial isolation, presence and degree of acute or chronic gvhd and relapse of the disease. graft failure was %(g ) vs %(g ) (p= . ), intermediate risk cmv serostatus %(g ) vs (g ) (p= . ). the most relevant characteristics and complications are described in table . contrast analysis between g vs g showed that within the whole group there were deaths ( % vs % respectively) (p= . ), the overall survival -year was % vs % (p= . ) with a median of vs months respectively; mortality associated with relapse was % vs % respectively, and mortality related with transplantation was % vs % respectively. conclusions: abo incompatibility did not show association with complications related with the infusion, but there was a higher tendency of graft failure in the abo incompatibility group. it has no statistical significance, but it is important to expand its study. disclosure: none declared methods: retrospective data for nhl patients who underwent asct between and was analysed. patients were identified using the swbmt database and data on mets was collected using paper and electronic hospital records. forty-eight patients were excluded due to loss of follow-up, inaccessible/incomplete records, or death. cause of death was not determined. the ncep-atpiii definition of mets was used. this requires ≥ of criteria to be met. a bmi of ≥ kg/m and hba c of ≥ mmol/l were used to replace central obesity and impaired fasting glucose, respectively. other criteria include triglycerides (tgs) ≥ . mmol/l or treatment, high density lipoprotein cholesterol (hdl-c) < . mmol/l (male), < . mmol/l (female) or treatment and blood pressure > mmhg systolic or > mmhg diastolic, or treatment. results: the prevalence of mets in the cohort was % (n= ). eighty-two percent of patients (n= ) met one or more criterion for mets. twenty-seven percent (n= ) fulfilled only one criterion, % (n= ) fulfilled two criteria, % (n= ) three criteria, % (n= ) four criteria, and % (n= ) five criteria. the greatest prevalence of mets was in the + age group, accounting for out of ( %) patients with mets. overall prevalence decreased with declining age ( table ). the number of patients aged < years was too small to make any judgement on risk. raised triglycerides was the criterion most frequently met ( / patients), followed by hypertension ( ), raised bmi ( ), low hdl-c ( ) and an increased hba c ( ). conclusions: the prevalence of mets in our cohort ( %) was higher than the estimated worldwide prevalence of %, with the majority in the + age category. this is in keeping with other post-transplant studies, which show an increase in prevalence of mets after transplantation. moreover, the overall prevalence of mets was greater in the older population, which could be associated with the cumulative effect of ageing on the decline of normal metabolic homeostatic mechanisms. background: acute renal failure (arf) is a frequent complication in the early post-allogeneic hematopoietic stem cell transplant (allohsct) period with either myeloablative (ma) or non-myeloablative (nma) conditioning regimens. the aim of this study was to compare the incidence of arf in both types of hsct and to evaluate its impact on overall survival (os) and non-relapse mortality (nrm). methods: all allosct performed in one center between and were included in this study. allohsct from cord blood and from haploidentical donors were excluded. the renal function and the incidence of the main complications after allosct from day to day + were evaluated. arf was defined according to kdigo (kidney disease improving global outcomes) classification; the relative increase of serum creatinine levels was considered a marker of kidney damage. results: seventy-seven patients received a ma allohsct and a nma allohsct. recipients of nma allohsct had a higher median age ( years [range: - ] vs. years , p< . ), higher frequency of arterial hypertension ( % vs. %, p< . ) and showed most frequently active disease at allosct ( % vs. %, p= . ). in both groups the most frequent graft-versushost disease (gvhd) prophylaxis regimen was cyclosporine a and methotrexate. the median follow-up time was . years for the nma group and . years for the ma group. patients from the ma group had higher incidence of grade - mucositis ( % vs. %, p< . ) and acute gvhd of any grade ( % vs. %, p= . ) than patients from the nma allohsct. the incidence of arf was similar in both groups ( % in nma and % in ma). in the nma group arterial hypertension (hr . , p= . ), obesity (hr . , p< . ) and prior pneumonia (hr . , < . ) were predisposing factors for arf by multivariate analysis, whereas any factor was identified in the ma group. arf had no impact on -year os in both groups ( % vs. % p= . for the nma group and % vs. % p= . for the ma group). however, worse os were observed in patients with grade - arf in the nma group ( % vs. %, p= . ) and in patients with grade arf in the ma group ( % vs. %, p= . ). in turn, arf had no influence on nrm in the ma group but was associated with a trend for higher nrm in the nma group ( % vs. %, p= . ). conclusions: arf is a frequent complication in patients receiving allohsct irrespective of the intensity of the conditioning regimen. moderate and severe arf had negative impact on os. disclosure: supported by grants from: asociación española contra el cáncer, aecc (gc biga), instituto carlos iii (pi / fi), -sgr (grc), cerca program from generalitat de catalunya, and "la caixa" foundation. treatment and risk factors of hepatic veno-occlusive disease after pediatric hematopoietic stem cell transplantation: a single-center experience barbaros sahin karagün , ilgen sasmaz , ali bülent antmen background: defibrotide emerged as a promising treatment option for hepatic veno-occlusive disease, a significant cause of mortality in recipients of hsct. as vod diagnosis is quite difficult even with the recently introduced ebmt criteria, studies which report treatment outcomes and response to prophylaxis are required. our aim was to evaluate the efficacy of defibrotide prophylaxis in hsct recipients at our center. methods: a total of transplants in patients from january to july were included in this study. all patients had factors that increased the risk of vod and all received mg/kg/day prophylaxis. patients' coagulation, renal and liver function test were monitored daily and all clinical findings and complaints were recorded. diagnoses were made via the ebmt vod criteria and patients who developed vod received treatment with increased df dose ( mg/kg/day) and supportive interventions. after complete remission of vod findings, patients were returned to the prophylaxis dose. close follow-up of patients was performed until days. results: in total, patients developed vod ( . %), none of the cases were severe ( mild, moderate). median age was . years and the most common clinical findings were weight increase, hepatomegaly, right upper quadrant pain and ascites development. in those with vod, treatment with mg/kg/day df was initiated and average duration of treatment with this dosage was . ( - ) days. no adverse events were reported in any of the patients. conclusions: our findings are consistent with previous studies on this topic, and we believe that the use of df as a prophylactic agent for vod is beneficial for pediatric patients with risk factors. disclosure: the authors report no conflicts of interest in this work. background: several factors might influence outcome of allo-hsct. analysis of the impact of donor-receptor blood group-incompatibility have been performed in different series not always showing the same results. as a consequence, its clinical impact remains controversial. minormismatch is characterized by the ability of donor b lymphocytes to produce anti-recipient antibodies. in majormismatch cases, antibodies against donor antigens are present in the recipient. methods: pts underwent allo-hsct between may and august in our center. median age was years (range: - ). pts were male ( . %) and female ( . %). baseline diseases were: aml, lpd, mds, all, mpd, mm, and bmf. donor was unrelated in , and related in cases (including haplo-identical). donor-recipient abo compatibility was as follows: ( . %) majormismatched (including bidirectional), and ( . %) nonmajor-mismatched (including minormismatched and matched). donor-recipient rh compatibility was as follows: ( . %) major-mismatched, and ( . %) nonmajor-mismatched (including minor-mismatched and matched). the impact of donor-recipient abo and rh compatibility on transfusion needs (prbc and platelet concentrates) and survival by day + was analyzed. results: for the global series the median number transfusions by day + was: ( - ) prbc and ( - ) platelets concentrates. day + overall mortality was . %. rh-incompatible and nonmajor abo incompatible cases showed no different results. however, major abomismatched cases needed more prbc transfusions (median: ; range: - ) and more platelet transfusions (median: ; range: - ), and had higher day + mortality ( . %) (p < . ) (see table) . conclusions: our analysis showed: ) donor-recipient rh-incompatibility, as well as minor aboincompatibility had no impact on prbc and platelet concentrates transfusion needs nor on -day mortality; ) contrarily, donor-recipient major abo-incompatibility had a significant adverse impact on prbc and platelet concentrates transfusion needs and -day mortality. ) donor-recipient rh-incompatibility and minor aboincompatibility.might be considered of marginal importance at the time to choose a potential donor. ) donorrecipient major abo-incompatibility should probably be a factor to be considered, along with other features, to choose the best donor background: survivors of haematopoietic stem cell transplantation (hsct) are at significant risk of developing treatment-related complications, including cardiovascular risk factors such as arterial hypertension, that could eventually lead to cardiovascular disease. the aim of this study is to evaluate the incidence and risk factors of hypertension following hsct in a colombian population. methods: a retrospective cohort study was conducted to assess the incidence and risk factors of hypertension in consecutive adult hsct recipients who underwent transplantation between and at a tertiary referral center in colombia, south america. blood pressure data, from two different measures, were collected at time points: day of mobilization for autologous hsct and day before infusion for allogeneic transplantation, day , and months , , and post-transplantation. hypertension was defined as having a systolic blood pressure >= mmhg and/or a diastolic blood pressure >= mmhg. patients with history of arterial hypertension were excluded. results: one hundred and seventy-five patients were included, with a mean age of years (range - ). ninety-one patients ( %) were male. one hundred and sixteen patients ( . %) underwent autologous hsct and ( . %) allogeneic hsct. the most common indication for hsct was acute leukemia ( . %), followed by non-hodgkin lymphoma ( . %) and multiple myeloma ( . %). twelve patients ( . %) had medical history of type diabetes mellitus (dm), ( . %) dyslipidemia, ( . %) alcohol consumption, and ( . %) tobacco smoking. only two of the patients with history of tobacco smoking were active smokers at time of transplantation. twenty-four patients ( . %) had developed hypertension by the end of the first year post-hsct follow-up. two patients ( . %) had systolic and diastolic, ( %) had only systolic, and ( . %) had only diastolic hypertension. only one patient was hypertensive at more than two time points. the incidences of hypertension at each time point were . % on day post-hsct, . % at first month, . % at three months, . % at months, and . % at one-year post-transplantation. allogeneic hsct (p< . ), therapy with calcineurin inhibitors (p< . ), pre-hsct fasting glucose levels (p< . ), acute gvhd (p< . ), chronic gvhd (p< . ), and media of diastolic blood pressure (p< . ) were significantly associated with the development of arterial hypertension. however, age, history of type dm, history of tobacco consumption, volume of infusion, prophylactic treatment for gvhd with mycophenolate, chronic gvhd, serum creatinine level on day of hsct, and being overweight or obese at time of transplantation were not significantly associated with the development of hypertension. conclusions: arterial hypertension is a fairly common complication in hsct recipients. similar to findings reported in previous studies, association between allogeneic stem cell transplantation, therapy with calcineurin inhibitors, and acute and chronic gvhd, and post-hsct hypertension was found in the present cohort. further studies are needed to assess the link between hsct and developing long-term cardiovascular complications. disclosure: nothing to declare tramadol-based pain management of oral and esophageal mucositis in pediatric hsct recipients background: mucositis is one of the most common early hsct complications seen in about % transplant recipients with % of patients developing gr iii-iv mucositis. mucositis is characterized by painful gastrointestinal mucosa lesions impairing the solid and liquid foods intake and increased risk of infections, bleeding, and intestinal paresis. thus, it greatly decreases the quality of life of a transplant recipient. according to who recommendations, the moderate pain control in pediatric patient is based on the use of low-dose morphine. however, there are some factors such as genetic polymorphisms causing variable morphine pharmacokinetics in children, side effects, and social factors (caregivers' general unwillingness to use narcotic analgesics), which cause the need for alternative pain relief options in pediatric practice. tramadol, which has both opioid and non-opioid mechanisms of action, may be a feasible option in mild to moderate pain. it may be delivered via patient-controlled analgesia (pca), although there is no consensus on its optimal parameters in pediatric practice. methods: a total of pediatric patients with a median age of (range to ) years receiving an autologous or allogeneic hsct in our clinic as part of the treatment regimen for solid tumor (n= ), leukemia (n= ), acquired aplastic anemia (n= ) or inherited condition (n= ) were included. conditioning regimens were myeloablative (mac) in and reduced-intensity (ric) in patients. all patients had oral and/or esophageal mucositis accompanied by moderate pain. the pain severity was assessed using the scales corresponding to patient's age and varied from to points. the pain control was based on intravenous tramadol administration using patientcontrolled analgesia (pca) approach. the following pca parameters were used: loading dose of . mg / kg (not exceeding mg), basal infusion rate of . mg / kg (not exceeding . mg), a bolus of . mg / kg (not exceeding . mg), lockout interval of min. the maximal daily dose was mg/kg/day. the pain control was considered adequate if a patient was satisfied or the basic and breakthrough pain score values were not higher than and, accordingly. in case of inadequate pain control nsaids were added. non-responders were switched to morphine. all patients were divided into groups based on conditioning regimen intensity. results: as a whole, % of patients did not require pain control measures escalation. the tramadol pain control rate was slightly higher for ric (n= , %) compared to mac (n= , %) recipients. in most cases the inadequate pain control was due to progressive mucosal lesions. the pca regimen used was characterized by very few complications. drowsiness was observed in ( %) of patients, in all cases the patients also had anemia. there was only ( %) patients with severe nausea requiring switching to morphine. conclusions: tramadol is an effective pain control option in transplant recipients with mild to moderate pain due to oral and esophageal mucositis without progressive mucosal lesions. the pca allows achieving a very low complication rate. therefore, this option may be considered for both mac and ric recipients. disclosure: no immune reconstitution of lymphocyte subsets after allogenic stem cell transplant (sct) and vaccination background: infectious diseases are a major cause of morbidity and mortality after allogenic stem cell transplant (sct). vaccines constitute an effective strategy to prevent infections but the optimal timing to start vaccinating is not well stablished. in order to individualize the early vaccination schedule, we studied the lymphocyte subsets involved in generating enough response to produce protective serological levels. methods: we studied retrospectively patients that had undergone allogenic sct at our hospital. patient distribution -age range: - years-old; diagnosis: acute leukaemia/myelodysplastic syndrome/ chronic myeloid leukemia ( patients), lymphoma ( patients). analytic parameters: tcd +, tcd +, nk, total b and functional b lymphocyte subsets (naïve igd+cd -, memory igd+cd + and igd-cd +, and effectors cd ++cd ++). immunoglobulin levels (igg, iga, igm) and specific igg for pneumococcus, tetanus, hbv, chickenpox, measles, rubella and mumps. clinical parameters were collected from medical records. results: we distributed patients in two groups, based on the timing of lymphocyte analyses: -less than months since sct ( patients) no patient showed complete immune reconstitution, although had enough t and functional b lymphocytes to generate response to vaccination. in these patients, vaccination for pneumococcus was completed and they generated sufficient protection antibody levels, despite being under immunosuppressive treatment. -more than months since sct ( patients) before the beginning of vaccination, we collected specific antibodies of patients. we compared the serological status before and after sct and observed that protection against tetanus was the most frequently preserved ( patients) and hbv the least frequent ( patients). other than one patient treated with alemtuzumab, all patients in this group had minimum absolute count of tcd + (> cells/microl), tcd + (> cells/microl), nk (> cells/microl) and b cells (> cells/microl). we also observed presence of b effector and b memory cells, with predominance of igd-cd + memory cells. immunoglobulin levels were within the normal range. in this group, we registered vaccination in patients. all of them were vaccinated against flu, and against pneumococcus and hbv. the rest of vaccines administered were heterogeneous in type and timing. patients were under immunosuppressive treatment at the time of vaccination and were able to generate enough specific antibodies for pneumococcus. conclusions: immune reconstitution was not completed months after sct, although minimal immunological reconstitution was observed tcd + and no-switching memory b lymphocytes were the last ones to reach minimum normal values according to patient age. some patients maintain serological protection after allogenic sct. immunoglobulin levels were normal, suggesting no need for immunoglobulin administration to prevent infections. flu, pneumococcus and hbv vaccines were the most frequently administered. pneumococcus vaccination generated a much larger serological response than hbv. this seroconversion occurred in patients under immunosuppressive treatment. the analysis of lymphocyte t, nk, b total and b functional subsets could be useful when programming an early vaccination schedule after sct. completion of the vaccination schedule was heterogeneous despite giving specific indications. therefore a more rigorous supervision of the process may be required. background: the significant advances that have been achieved in the allogeneic transplantation (allohct) field, have resulted in better post-transplant outcome and therefore complications other than the graft vs. host disease (gvhd) or disease recurrence become increasingly important. the post transplant metabolic syndrome (pt-ms), which caused by several factors (i.e. immunosuppressive agents, chemo-radiotherapy, anti-viral, and biologic therapies) is a well known post transplant complication in pediatric allografted long-term survivors however, only few studies have evaluated the prevalence of the pt-ms in adults. in this retrospective study, we sought to evaluate the incidence, the risk factors and the impact of the pt-ms on the allosct outcome. methods: since , patients ( males and females) with adequate clinical and laboratory data and a minimum follow-up of months were included in the study. their median age was . ( - ) years and after a myeloablative (n= ) or a reduced intensity (n= ) regimen they received either mobilized peripheral blood stem cells (n= ) or marrow graft (n= ), originated from full-matched siblings (n= ) or haploidentical donors (n= ). calcineurin inhibitors plus either short-term methotrexate or mycophenolate mofetil were given as gvhd prophylaxis. the diagnosis of pt-ms was based on the ncep-atpiii criteria; for patients with unknown data for abdominal circumference the body mass index (bmi) ≥ kg/m was consider as a criterion for pt-ms diagnosis. the independent t-test, logistic regression analysis and logrank tests were used for the statistical analysis. results: twenty ( . %) patients ( males, females) assessed to have pt-ms within the first months following the allograft. seventeen diagnosed after the st trimester post allosct and additional patients after nd trimester. sixteen out of patients had elevated glucose and bmi> kg/m , / elevated triglycerides levels, / low hdl levels and / hypertension. four ( %) had already known history of ms before allosct (for patients no data were available for ms diagnosis before allosct). interestingly, for / ( %) patients who had diagnosed with pt-ms either in the st or in the nd trimester the syndrome was reversible and did not fulfill the criteria for pt-ms beyond months post allosct. patients' gender, age, bmi, the type of conditioning regimen and gvhd co-existence evaluated as potential predisposing factors for pt-ms diagnosis. in univariate and multivariate analysis only the: bmi> kg/m and age> years were detected as significant risk factors (p< . ). the pt-ms did not affected negatively the survival or the nrm incidence post allosct conclusions: in our study, in agreement with other publications, we demonstrated that the pt-ms is not an uncommon complication post in the early post transplant period however, for a significant number of patients the syndrome was a reversible. for patients with high risk features (bmi> kg/m , age> years, known history of diabetes-mellitus, dyslipidemia, hypertension) apart of close monitoring, specific diet and encouragement for adequate exercise might help to reduce the incidence and the severity of pt-ms. nevertheless, prospective and well design trials are warranted to determine the accurate incidence, severity and the impact of pt-ms on the allosct outcome. disclosure: no conflict ofinterest experience of a single center in the humanization of the hospitalization process: technology and team training impact on the qol of the patient and family maria claudia moreira , marcia rejane , marcia garnica , andrea ribeiro , paulo cesar dias , ilza fellows background: hematopoietic stem cell transplantation (hsct) is one of the most aggressive therapeutic modalities of internal medicine, making it a highly stressful experience for the patient and his family. the duration of hospitalization can be prolonged by several intercurrences, frequently generating anxiety in the patient and their caregiver, which may lead to confinement and reactive depression. interventions in the hospital environment, in addition to the continuous training of the multidisciplinary team, can have a positive impact in this process with improvement in the process of discharge and quality of life of the patient and his / her family. methods: the objective of this research was to evaluate the impact of a reformulation in the unit, completed in may , which modified the facilities with availability of hermetic balconies in each room, with a view of an internal garden. there was also the addition of a screen in the corridor of the floor with images -technology known as videoowall, interconnected to motion sensors (kinects) that allow interaction between patients and families, besides facilitating physiotherapy and physical exercise. there was re-training of the multidisciplinary team with emphasis on the practice of humanization. the methodology consisted in the application of questionnaires of satisfaction to patients and their families during the period of hospitalization in a bone marrow transplant unit in the third quarter of . the items evaluated ranged from the quality of the information provided by the medical team and nursing, to the cordiality and agility with which the patient and his patient were treated by the global team. the results were compared with a similar period of the same unit in the previous year and with the indices collected simultaneously in another unit of the same hospital (cardio-intensive). results: overall and segmental satisfaction scores in the various items surveyed were higher when compared to the previous period of the same unit and were also higher in those obtained in a high complexity unit of the same hospital, composed of patients submitted to mental and psychological stress similar to onco-hematologicos.a reports of "free speech" were also obtained anonymously, in order to guarantee the authenticity and free expression of the subjects analyzed. conclusions: the results obtained allowed the validation of the technical and professional team initiatives, bringing indicators that will allow better monitoring and support of these patients and their relatives in this difficult time of treatment. they served as an initial tool in the continuous process of humanization and stimulated the multidisciplinary team to continuously improve this process. disclosure background: pure red cell anemia (prca) is a rare complication of abo-incopatible hematopoetic stem cell transplantation characterized by anemia, reticulocytopenia and absence of erythroid precursors in patient's bone marrow. most patients with prca resolve spontaneously within months, however a small number of patients requires continued red blood cell (rbc) transfusions. the treatment of this complication is difficult and not standardized. different approaches has been used such as rituximab, donor lymphocytes, plasma exchange with different outcome. recently, a remarkable response to treatment with bortezomib has been described in a case of prca. methods: we reviewed patients who received an allogeneic hematopoetic stem cell transplant (hct) between januar and august at our institution. sixty eight patients received a major abo-mismached hct. prca was defined as a completely absence of erythroid precursors on day + bone marrow puncture, with absence of donor red cells and the recipient requiring rbc transfusion. results: only one patient developed prca ( . %). a years old male received a myeloablative hla-matched abomismatched sibling donor transplant (brother, years) for acute myeloid leukemia (aml), with t( ; ) cr ,mrd positive (runx -runx t ). the donor was blood type a rh positive and the patient rh positive. the patient had no complication after transplant. the day + bone marrow puncture has shown only few erythroid precursors and day + puncture and biopsy no erythroid precursors, he had transfusion dependent anemia requiring a rbc transfusion every two weeks and retukulocytopenia. parvo virus and cytomegalovirus were negative. due to very high ferritin level (> . u/l) and increased luiver enzymes without signs of gvhd, the treatment with deferasirox has been started. the patient has achived cr , mrd negative, and has evidence of complete chimerism. high titers of anti-a and anti-b issohemagglutinin was present.we started the treatment with rituximab mg/m weekly, weeks, however without response. the pathogenesis of the prca is thought to be due to the recipients plasma cells, bortezomib, a proteasome inhibitor inducing apoptosis of plasma cells has been given s. c. , mg/m two times weekly, for two weeks. the patient responded to the treatment two weeks later with increase in hb, which was , g/dl and increase in retikulocyte number. the patient has continued to be well at the last control. conclusions: prca aplasia is a rare but serious complication after abo-incompatible hct. bortezomib is an effective treatment for this complication if mediated by residual host isohemeagglutinins after hct and should be recommended as standard of care. clinical methods: this work is retrospective, observational, cross-sectional and analytical. it included all patients who received hsct at stem cell transplantation unit (utmo, by its spanish acronym) at solca-guayaquil, between the years - .we use the kaplan-meier method to analyze the survival rate between the autologous and allogeneic transplant. the information collected for this study was obtained from the database of the solcay institute and the review of the files of the patients included. results: at least, patients have been undergoing to hsct between - years. according to the type of hsct, . % received an autologous transplant and . % received an allogeneic transplant, from which . % were from a related donor. the main source of transplant was peripheral blood in . %, followed by % obtained from umbilical cord blood and . % by bone marrow aspiration. the most frequently reported pathologies were acute lymphoblastic leukemia (all) ( %), multiple myeloma (mm) ( %) and acute myeloid leukemia (aml) ( . %). the overall survival was % (ic: %). the . % of patients that were undergoing to autologous transplant have survive, meanwhile the patients that were undergoing allogeneic transplant only the . % have survived (p< . ). the highest death rate occurred during the first year after hsct, and decreased considerably after that period. the main cause of mortality related to transplant (mrt) was the graft-versus-host disease (gvhd) ( %); however, the main cause of mortality in the study population (n= ) was relapse in . % of the patients, presented more frequently in all. conclusions: the results showed that % of patients undergoing to hsct have survived. a high rate of deceased patients in this study, have died in the first year before the transplant ( . %%), due to relapse. the main cause of deceased in the study is not related to hsct, and was the relapse in % of patients, in compare the gvhd was the main cause of mrt ( %). we consider that hsct is a technique that is still under development in ecuador, but despite the short time it has been taking and the institutional and medical limitations present in the health field, has presented excellent results comparable to studies conducted in developed countries. [ background: pigmented epithelioid melanocytoma (pem, early known as 'Аnimal type' melanoma) is a rare tumor with unpredictable clinical behavior and metastatic potential. pem generally has favorable prognosis. involvement of regional lymph nodes is not rare. extranodal and distant nodal metastases are extremely rare. we report about patient with fanconi anemia (fa) and pem with developed distant metastases in the early term after allogeneic hematopoietic stem cell transplantation (hsct). methods: -years old boy with fa was hospitalized for hsct. the blue-black painless nodulus х mm was noted on the left cheek. this lesion was observed from early childhood and during life only slightly increased in size. there were no distant and regional metastases on computerized tomography (ct) and scintigraphy with m tc. the nodulus and regional lymph nodes were radically removed before hsct. the resection margin was within the normal tissue. microscopically the derma and subcutaneous fat were infiltrated with epithelioid and spindle cells with total expression of s , melana, mhb , cyclind . ki- expression level was - %. histological structure was specific for pem. hsct with tcrαβ+/cd + graft depletion from match unrelated donor was performed. the conditioning regimen included total lymphoid irradiation gy, fludarabin mg/ m , cyclophosphamide mg/kg, rabbit atg mg/kg and rituximab mg/m . results: at + day after hsct was detected the tumor on the left cheek and parotid region with a histological structure identical to the primary lesion. on ct in s segment of the left lung was detected focus x mm with a cavity. invasive aspergillosis was suspected and empirical antifungal treatment was started. but in days the lung lesion increased in size to x x mm and penetrated in the bronchus. after bronchoscopy with biopsy, pem metastasis was histologically confirmed. moreover, the tumor on the face continued to grow. therapy with cobimetinib and vemurafenib was not effective and patient died from progression of pem on + day after hsct. conclusions: pem was early described as indolent tumor with rare distant metastasis and favorable prognosis. we suspect that pem may acquire an aggressive course in the absence of immunological control, especially in high immunocompromised patients after hsct. disclosure: nothing to declare p abstract withdrawn lidia gartcheva , antoaneta mihova , penka ganeva , margarita guenova , branimir spassov background: the main objective of the study is to assess the dynamics of quantitative and qualitative changes in the parameters of the b cell population and the production of immunoglobulins in patients after autologous transplantation of hematopoietic stem cells in the course of recovery of the immune system. methods: patients with hematological neoplasms undergoing autologous transplantation were included in the study: women and men, with an average age of years. patients were diagnosed with lymphoma (n = ), multiple myeloma (n= ), leukemia (n = ) and solid tumors (n = ). at the time of transplantation, patients were in complete clinical remission or at least with very good partial response, patients were in partial remission and patients -with progression. all patients were evaluated in nine time points through examinations by clinical-laboratory, flow cytometric and immunochemical methods. results: the percentage of cd (+) b cells reached the minimum values one month after transplantation then began to increase in the second month reaching a plateau around the mean values in the period - months after transplantation. the absolute number remained low during the entire period of observation. the amounts of igg and igm serum immunoglobulins gradually increased within the reference range throughout the entire period, while the iga level varied around the lower reference range. conclusions: implementation of an adequate humoral immune response is hampered by the reduction of circulating b cells, suppressed proliferative potential and functional deficits. restoration of b-cell function occurs over a period of months to years after autologous transplantation. clinical trial registry: no clinical trials disclosure: nothing to declare justyna background: allogeneic hematopoietic stem cells transplantation (allo-hsct) is a life-saving and well established therapy for wide range of diseases. however, it is still uncommon treatment for infants less than months of age. the data about indications and outcome of allo-hsct in the youngest group of patients is sparse. the primary objective of this study was to assess the incidence, indications, post-hsct complications and general outcome of allo-hsct among infants not older than months. latter sequelae of hsct such as physical and cognitive development were secondary aim of this study. methods: we retrospectively analyzed data of patients who underwent allo-hsct before year of age in department of pediatric hematology, oncology and bone marrow transplantation in wrocław during years - . clinical and epidemiological features as well as indications for transplantation, early and late complications and general outcome were assessed. results: infants who underwent hsct in our department comprise . % of all patients undergoing hsct in analyzed period of time. thirty-one ( . %) patients received stem cells from matched unrelated donor (mud), ( . %) from mismatched (haploidentical) related donor (mmrd) and ( . %) from a sibling donor (msd). non-malignant disorders were indication for transplant in ( . %) patients and malignant diseases in ( . %) . acute graft versus host disease (agvhd) occurred in ( %) infants, chronic graft versus host disease (cgvhd) in ( %). majority of graft rejections were seen in infants transplanted from mmrd ( . %), whereas the rest ( . %) was associated with mud. median follow-up in study cohort was days, days for alive patients (range days- . yrs) and days for those deceased (range days- days). overall survival (os) in study cohort was . and transplant related mortality (trm) was . . in children with malignancy ( . %) patients died comparing to ( . %) patients in non-malignant group respectively. main cause of death in analyzed group of infants was infection ( %). conclusions: . allo-hsct is rarely performed in children less than months of age. . majority of those patients receive stem cells due to non-malignant disorder. . among youngest hsct recipients, haploidentical transplant are more common than in general pediatric transplant population. . graft rejection is a significant problem in infants transplanted from mmrd. disclosure: nothing to declare unusual non-infectious lung complication after allogeneic haematopoietic stem cell transplantation claudia lucia sossa melo , , manuel rosales , francisco fernando naranjo junoy , , sara inés jiménez , , luis antonio salazar , , angela maría peña , , maría angélica chacón manosalva , maria luna-gonzález , claudia marcela chalela , manuel ardila-báez jirovecii infections, viral infections or nocardia. we describe the case of a patient with acute lymphoblastic leukemia (all) diagnosis with pap associated to a hsct and pulmonary pneumocystis. methods: a -year-old colombian female patient diagnosed with b-precursor all of high-risk in january , positive philadelphia chromosome, positive bcr / abl in february , infiltration to the central nervous system (cns), . % of lymphoblasts, and karyotype without legible metaphases. refractory to induction according to the pethema protocol (vincristine, daunorubicin, prednisone, l-asparaginase) with presence of . % blasts at the end of the induction. re-induction was performed with the flag-ida protocol (idarubicin, fludarabine, cytarabine) achieving complete remission, obtaining minimum residual disease (mrd) < . . dasatinib was initiated by bcr / abl expression and cns involvement at the time of diagnosis. an allogeneic hsct was performed, from a male brother donor, with low intensity conditioning tt buflu and prophylaxis of graft-versus-host disease (gvhd) with tacrolimus and sirolimus. patient showed early posttransplant complications, given the reactivation of cytomegalovirus and hemorrhagic cystitis grade i due to adenovirus. late complications such us gvhd at the cutaneous level and subsequent hepatic and gastrointestinal involvement were seen too, for which immunosuppressive therapy was administered with high doses of systemic corticosteroid. results: patient was hospitalized on day + posttransplantation due to febrile neutropenia and respiratory symptoms, with normal chest ct, and ct of paranasal sinuses with acute pansinusitis, for which she received meropenem gr intravenously every hours plus vancomycin gr intravenously every hours during days with symptom resolution. she remained hospitalized for cytopenias with normal bone marrow and % chimerism. on day + posttransplant she presented fever and leukocytosis, with acute respiratory failure with chest ct that showed bilateral alveolar occupation, "crazy-paving" pattern and frosted glass (see image), so diagnostic fibro-bronchoscopy was performed, reporting postoperatively for pneumocystis jirovecii. she received days of trimethoprim-sulfamethoxazole, with a torpid evolution requiring mechanical ventilation and tracheostomy, persisting with hypoxemia. the report of cultures for fungi, mycobacteria, and respiratory panel of filmarray were negative. a pathology report was obtained with % neutrophils, as well as pas staining with acellular pink material and elevated serum ldh, with a diagnosis of secondary pap. the patient continued with poor general condition, refractory hypoxemia, high ventilatory parameters and hemodynamic instability, due to which she was not able to be a candidate for treatment with total pulmonary lavage; leading to multi-organ failure and later death. [[p image] . high resolution chest ct. sample opacification in frosted glass (a) and pattern ''crazypaving'' (b)] conclusions: the importance of considering the diagnosis of pap as a noninfectious pulmonary complication in patients with allogenic hsct despite its low incidence is recognized. disclosure: nothing to declare methods: once the project was approved by the clinical trials and ethics committee, pairs of blood samples were drawn ( from picc line and from venepuncture) from voluntary allo-hsct recipients who were receiving continuous infusion tacrolimus from february through august . the pts had inserted a double-lumen polyurethane picc. tacrolimus was always administered through the red line, and the blood draw always performed through the purple line. all of the patients signed the informed consent. were male and women. median age was years ( - ). of the venepunctures were carried out in the arm where the picc was set, and the other from the contralateral arm. a limited group of nurses performed the extractions of the samples. results: as shown in the table, tacrolimus trough levels determined in blood from venepuncture were similar to those in blood drawn through the picc (median: . vs . ng/ml). when comparing one by one in the individual patients, the differences were not significant, and changed the dosing prescription in no cases. conclusions: in our experience, there are not significant differences in tacrolimus levels draw from the picc line, compared with a peripheral site. so, in our opinion, if the line for tacrolimus infusion is properly identified and the one used for the sample draw is the alternative one, venepunctures to obtain sample from peripheral sites are not justified for tacrolimus levels measurements. background: patients undergoing a hsct may require icu admission due to transplant-related toxicities. the aim of this study was to analyse a single centre experience with hsct patients requiring icu admission and the factors affecting outcome. methods: we included all adult patients (age >= ) who had an allogeneic or autologous hsct during (d between - - to - - ) at st. george's hospital. data was retrospectively collected from patients' notes. icu outcome and -day survival were analysed. for those patients who were admitted to icu more than once, outcome was analysed from their last icu admission. results: allograft patients were included. were male, with a median age years (range - years). were female, with median age years (range - years). diagnosis n (%) includes all ( %), aml ( %), acml ( %), cmml ( %), hl ( %), mds ( %), mds/mpn ( %), fl ( %), scd ( %). sixteen ( %) patients received their first transplant, ( %) received second transplant. eight ( %) patients had sibling donor, patients ( %) had unrelated donor. sixteen ( %) patients had / matched donor, ( %) patients had / matched donor, ( %) patients had / matched donor. nineteen ( %) received reduced intensity conditioning (ric), one ( %) received myeloablative (ma) conditioning. majority of ric allo-hsct patients were conditioned with fludarabine, mephalan, campath (fmc). a small number were conditioned with busulfan, fludarabine and atg. the ma allo-hsct patient was conditioned with tbi, cyclophosphamide. gvhd prophylaxis was ciclosporin alone starting on day - with a target level of - ug/l for all ric and ciclosporin and methotrexate for the ma patients. two ( %) allograft patients were admitted to icu on three occasions. both patients were male, and years old. one had mmud allograft for mds/mpn. the other had nd mud allograft for relapsed aml. the reasons for icu admission include sepsis, cardiac arrest and respiratory failure. the median duration of icu admission was days (range - ). there were deaths within days of transplant. one patient died on day + during his second icu admission with multi organ failure (mof). one patient died after icu discharge on day + with relapsed disease, bronchopneumonia with disseminated fungal infection. icu mortality rate was %, and -day mortality rate was %. nineteen autologous patients were included (median age (range - years)), ( %) were myeloma patients who were conditioned with melphalan, ( %) were lymphoma patients who were conditioned with beam. the icu admission was %. the -day mortality rate was %. conclusions: our centre's icu admission rate, icu mortality rate, cause of icu admission in allo-hsct patients and autologous patients is comparable to literature reports. autologous transplant is safe with no deaths and icu admissions despite an older age. the mortality rate for allo-hsct patients requiring icu admission remain high. all patients were appropriately referred to icu and there was no one who was denied icu admission. this analysis is being extended to preceding years. disclosure: nothing to declare liposomal doxorubicin for the treatment of iatrogenic kaposi sarcoma following hematopoietic stem cell transplantation background: iatrogenic kaposi's sarcoma (iks) represent a rare complication after hematopoietic stem cell transplantation (hsct), related to hhv- infection in hivnegative immunocompromised patients (pts). methods: we describe a case of iks occurred after an allogeneic hsct and we provide a review of the literature using pub med. results: a -year-old man, hiv-negative, received full hla-matched related hsct after a reduced intensity conditioning regimen for relapsed aml. gvhd prophylaxis was based on atg (fresenius mg/kg), cyclosporine (cya) and methotrexate. no severe complication occurred in the first days after transplant. shortly after cya withdrawal, he developed grade i acute gvhd. gvhd resolved after restarting cya. at fifth month after transplant, the patient developed several red and purple angiomatous plaque and nodules involving the skin of both lower limbs, right arm and the nose (figure ). skin biopsy revealed multiple localizations of iks and positive hhv- viremia was detected in the peripheral blood. a visceral involvement was excluded. patient was treated with cya tapering and nine courses of liposomal doxorubicin mg/m every days, obtaining a negativity of hhv- viremia and partial response of the skin lesions. at last follow up, at months after transplant, the patient was in complete remission (cr) for aml, cya-free without signs of gvhd recurrence and with his single stable residual iks lesion on his left limb, currently waiting for local radiotherapy. we found additional iks published cases after hsct. most of post-hsct iks were secondary to an allogeneic-hsct ( out of , . %) and occurred in adult ( , %) and male ( , %) pts. median age at the time of iks diagnosis was . years (range - ). thirteen pts ( . %) had mediterranean origin. the most frequent underlying disease was aml ( . %). gvhd prophylaxis was primary based on calcineurin inhibitor. half of the pts developed gvhd and were treated with steroid and other immune suppressive drugs. median time between the hsct and the occurrence of iks was . months (range . cutaneous iks was the prevalent form of manifestation, however visceral involvement was reported in pts ( . %). in four cases ( . %) an hhv- associated bm failure was report. immune suppression drugs tapering ( . %) and chemotherapy ( . %) were the most frequent actions taken after the diagnosis of iks. in most cases, liposomal doxorubicin was used as chemotherapy. cr rate was high, . %, whereas progression disease occurred in out pts ( . %), all of which had visceral involvement. in pts ( . %), iks was the cause of death. conclusions: withdrawn of immune suppression drugs and anthracycline based chemotherapy can represent a feasible treatment option for pts with iks after hsct. clinical background: acquired haemophilia a (aha) is an autoimmune disease caused by the spontaneous production of neutralizing immunoglobulin g (igg) autoantibodies (inhibitors) targeting endogenous fviii. treatment of these inhibitors presents additional challenges in a hematopoietic stem cell transplantation (hsct) recipient, because preservation of the graft that restores a normal hematopoiesis is critical. here we describe the management of a case of aha in an acute myeloid leukemia patient following hsct. methods: the clinical, laboratory and molecular aspects of a -year-old italian male who developed aha after allogenic bone marrow transplantation were collected and presented in order to show how we diagnose and manage this severe but rare complication within the special setting of hsct. results: a -years-old man with a flt- itd, npm- , runx -runx t and cbfb-myh negative, not differentiated, chromosomally normal acute myeloid leukemia (aml) in third complete remission (cr) was submitted to a hematopoietic stem cell transplantation (hsct) from his haploidentical son. the conditioning regimen consisted of oncothiotepa, busulfan and fludarabine and was followed by the infusion of a t-cell depleted bone marrow graft. gvhd prophylaxis consisted of cyclosporine a (csa) and mycophenolate mophetyl (mmf). neutrophil engraftment occurred on day + . recipient's autoimmunity was negative. at months post-transplantation the patient received an antipneumococcal vaccination. fifteen days post-vaccination the patient was admitted to our in-patient ward due to general malaise, diffuse muscle and joint pains, cutaneous bleedings, oedemas, hyperchromic urines and constipation. physical examination revealed diffuse ecchymosis, swelling of deep muscles with a progressive functional disability due to hematomas and hemorrhagic suffusions of the tongue frenulum. and anti-factor viii inhibitors . bu/ml (high titers > bu/ml). thus, a diagnosis of acquired autoimmune haemophilia a was made and treatment with feiba combined with prednisone was started. patient's clinical conditions dramatically improved as he referred an improvement of movements and the resolution of joint and muscle pains despite the persistence of deep hematomas just after one day of treatment that had determined an increase of fviii:c value to . % and an improvement of aptt to . seconds. on the following medical checks physical examination showed the progressive disappearance of deep muscle hematomas, and normal values of fviii:c. conclusions: aha is a rare but severe complication following hsct and it could appear years afterengraftment. a prompt diagnosis and an early treatment with feiba and corticosteroid are necessary to avoid life-threatening sequelae. the inclusion of the coagulation panel in the laboratory exams performed during the follow-up is advisable in order to early detect this life-threatening complication. disclosure: nothing to declare background: splanchnic thrombosis is an uncommon complication of myelofibrosis and a controindication to proceed to hematopoietic stem cell transplantation (hsct) due to the risk of additional vascular and endothelial complications. we present a patient with myelofibrosis (mf) that proceeded to hsct from an unrelated donor, despite splanchnic thrombosis unresolved after heparin treatment and unable to proceed to surgical treatment due to severe thrombocytemia. methods: a -year woman with mf secondary to essential thrombocythemia, with intermediate- score according dynamic international prognostic staging system (dipss) and with extreme splenomegaly (maximum diameter cm), refractory to ruxolitinib, showed an extensive thrombosis of the portal and splenic veins, unresolved after -week heparin therapy, at the time of availability of an hla ( / ) and abo matched unrelated donor. she received a conditioning regimen including fludarabine and thiotepa and a gvhd prophylaxis with atg thymoglobuline, cyclosporine and methotrexate, followed by the reinfusion of . x /kg cd + pbsc. at the time of transplant we were aware of an high risk of developing sos, on the basis of the older age of the recipient, the unrelated donor, the advanced stage of myelofibrosis and the ferritin serum level of . ng/mg. results: on day + after hsct sos complicated the aplasia phase, characterized by jaundice, ascites, weight gain, progressive increase in creatinine and bilirubin serum levels. an ultrasound of abdomen confirmed an unchanged thrombosis extension and the development of ascites. on day + the patient was categorized as very severe sos stage, according to ebmt severity criteria, because of doubling of bilirubin serum level in hours and a % increase in comparison with her baseline weight. therefore, defibrotide was promptly started in association with diuretic therapy. the treatment was continued for weeks and allowed gradual restoration of the water balance and normalization of bilirubin serum level. at the last follow-up, months after hsct, the patient shows the persistence of a non-transfusion dependent anemia, platelets . x ^ /ul, palpable spleen cm below the rib, > % allogeneic chimerism in the granulocytic compartment and % in the t lymphocyte compartment. splanchnic thrombosis is partially recanalized and replaced by collateral circles with cavernous aspects. the patient is on treatment with fondaparinux and has shown neither significant infectious episodes or acute or chronic gvhd. conclusions: we conclude that defibrotide treatment allowed to perform a successfull allogeneic transplant in a patient with mf associated with an overt picture of splanchnic thrombosis. background: hematopoietic stem cell transplantation (hsct) is associated with an increased incidence of secondary malignancies including skin cancer. squamous cell carcinoma (scc) is the most common type in patients who are receiving immunosuppressive therapy and chronic graft-versus-host disease (cgvhd) appears to be an important risk factor for its development. recent studies describe voriconazole exposure as an independent factor that may contribute to this increased risk as well. in our best knowledge, no cases of scc have been reported in pediatric allogeneic hsct to date. methods: we present a case report of a year-old boy who developed a scc with high-risk features six years after undergoing hematopopoietic stem cell transplant. results: a year-old boy with acute lymphoblastic leukemia (all) underwent a matched unrelated bone marrow transplant years ago. he developed grade iv agvhd followed by extensive cgvhd with generalized scleroderma. he required intensive and continued immunosuppressive therapy and was on prolonged antifungal prophylaxis with voriconazole. in march , he developed scc involving left temporal region that was completely excised. two months later, more lesions in scalp and nose were noted and intralesion treatment with methotrexate was started. however, an unfavorable evolution was noted and he was put on systemic treatment including cisplatin and cetuximab receiving the whole scheme from january to march and continuing only with cetuximab, ten doses in total, until may, for unaceptable and severe tubulopathy that required admission at the hospital in several ocassions. he achieved a very good partial response but progression was noted shortly in follow up. at this point, non curative therapeutic options were found and he was put on intralesion methotrexate and photodynamic theraphy in a weekly basis with palliative intention. unfortunately, tumor growth was fast and patient passed away in august , fifteen months after squamous cell carcinoma diagnosis, due to tumoral progression. conclusions: ) scc is a rare, non-previously described, secondary malignancy in children undergoing hsct. ) high-risk features scc constitutes an aggresive disease with a median overall survival below year. ) cgvhd appears to be an important risk factor for its development. ) voriconazole induced-photosensitivity might have played a role. ) cisplatin based regimens +/-cetuximab are a therapeutic option in disseminated and/or high risk cases. as outcomes are unsatisfactory in these cases, alternative therapeutic options need to be explored. disclosure background: pregnancy is a rare event after allogeneic stem cell transplantation (sct) for acute leukemia. here we report, to the best of our knowledge, for the first time on a successful pregnancy after treosulfan-based conditioning. methods: a -year old woman was diagnosed with acute myeloid leukemia (aml) secondary to chronic myelomonocytic leukemia in july . ovarian preservation was performed by leuprolide acetate depot injection prior to cytostatic chemotherapy. of note, no cryopreservation of oocytes or ovarian tissue was conducted. she received two cycles of chemotherapy consisting of idarubicine ( mg/m² on day - ) and cytarabine ( mg/m² b.i.d. on days , , and ). due to secondary origin of aml sct was performed in first complete remission of aml after conditioning with treosulfan ( g/ m² days - ) and fludarabine ( mg/m² days - ). she received . × cd -positive cells per kilogram body weight from a hla-matched unrelated donor. results: follow-up bone marrow aspirates showed continuous complete remission of aml. seven months after sct she became pregnant, but decided for induced abortion. in january , months after hsct she became pregnant again and desired the child. medical examinations were performed monthly on an outpatient basis in stringent cooperation with the maternity clinic. the course of pregnancy was unremarkable, although she was hospitalized due to premature labor in the th week of pregnancy. however, gynecological examination showed no clinical significant findings, so that section was planned and she could be discharged again. in the th week of pregnancy she gave birth to a healthy girl ( cm, g) by cesarean section. peripartum she developed hypoethesia of the left body half. neurological examination showed no abnormalities and she recovered immediately. there were no other postpartum complications. breastfeeding was established but additional food was necessary for a sufficient nutrition of the child. conclusions: this case of successful pregnancy following sct demonstrates that fertility can recover after treosulfan-based conditioning. however, detailed studies of ovarian function and fertility are necessary to gain more insight into the risk of premature ovarian failure. disclosure: nothing to declare. experimental stem cell transplantation p cd -cart therapy before allo-hsct in children and adolescents patients who diagnosed r/r b-all with e a-pbx background: b-all with e a-pbx in children and adolescents is described with favourable prognosis. but there are more than % patients with e a-pbx diagnosed as relapsed or refractory. the results of allo-hsct in children and adolescents with this group leukemia in our center was analyzed in order to understand the therapeutic effect of cd -cart on the patients. methods: retrospective analysis, from june st, to july , , all children and adolescents diagnosed relapse or refractory b-all with e a-pbx who received allo-hsct, total cases. all patients was divided into two groups depending on whether or not accepted cd -cart before allo-hsct. according to fcm-mrd and e a-pbx level before allo-hsct, os lfs and cumulative recurrence rate were analyzed. r . . was used as statistical analysis software. results conclusions: . for r/r b-all with e a-pbx in children and adolescents, fcm-mrd pre-transplant hasn't obvious effect on the outcome of allo-hsct, while the level of e a-pbx has obvious effect. the out come of e a-pbx negative group was obviously better than positive group. . cd -cart can obviously improve the os and lfs, it is mainly because of cd -cart can makes more patients fusion to zero. . for r/r b-all with e a-pbx in children and adolescents, if chemotherapy can't make the fusion to zero. it is suggested to accept cd -cart therapy to make the fusion zero. it can improve the outcome of os and lfs. disclosure background: currently, hematopoietic stem cell transplantation (hsct) represents the only curative treatment for numerous hematopoietic malignancies like leukemias, immune deficiencies or metabolic diseases. cd serves a quality marker for stem cell grafts, which is not solely expressed on stem cells but also on a variety of progenitors. the role and the impact of these subpopulations remains unknown. we made use of our genetic barcode system to analyze the influence and contribution during reconstitution on a clonal level. methods: fluorescence activated cell sorting (facs) was used to sort hematopoietic stem and progenitor populations, namely hscs, mpps, cmps and clps, which were lentivirally transduced with our previously established bc barcoding system. after mixing the marked cells with bone marrow support, lethally irradiated recipient animals were and transplanted and monitored over weeks. we focused on bone marrow, blood, spleen and thymus, on chosen endpoints ( w, w, w, w) and samples were used to analyze the contribution of the subpopulations during the reconstitution process based on fluorescent protein (fp) expression. to investigate the clonal contribution in different organs, we performed next generation sequencing (ngs) and frequencies of unique barcodes in a sample were analyzed by bioinformatical approaches. results: a maximum of % of cells expressed the encoded fps, which were mostly derived from the hscs and mpps. cmp-derived cells were only detected week after transplantation in the myeloid compartment. cells derived from the clps were not detected at any time point. we analyzed the barcode content of the differently marked cells after next-generation-sequencing. in accordance with the facs data, the majority of the clones during the weeks of observation are derived from hscs and mpps. cmp-derived clones were only contributing during the first weeks and clp-derived clones are barely detectable. we did not observe any major differences with regard to age of donor or recipient, despite the total number of clones is higher in the group, which received the "aged" graft, independently from the transduced cell population. conclusions: here we show the suitability of our highly complex multi-color barcode system to study the clonal contribution of hscs and three progenitor populations after hsct. our results will contribute to a better understanding how these different populations interact to support the establishment of a new hematopoietic system. emphasized by the variability in data of graft and recipient age, this comprehensive analysis gives rise to an impression to the necessity of personalized graft composition, by which treatment success could be influenced. disclosure: nothing to declare survival and fate of adipose derived mesenchymal stem cells in a rat brain injury model background: mesenchymal stem cells have been identified as promising candidates in the treatment of central nervous system (cns) injury through neurotrophic support and immunomodulation. adipose tissue is an attractive source of mesenchymal stromal/stem cells (ascs) for regenerative therapeutic applications because they can be harvested from autologous donors with minimally invasive methods, can be rapidly expanded ex vivo, show low immunogenicity if allogeneic, and can be used in autologous or heterologous settings. the present study examines the fate and effects of intracerebroventricularly (icv) transplanted ascs in a traumatic brain injury (tbi) model. methods: ascs were isolated from inguinal fat pad of adult wistar rats under sterile conditions and cultured according to standard procedures. ascs at passage ( x cells) were seeded and transfected with sleeping beauty transposase and pt venus-neo r plasmids. selection with g antibiotic resulted in the generation of a homogeneous asc population which expressed fluorescent venus protein for several passages, phenotypic characterization showed that these cells were . % double positive for cd and cd stem cell markers, verifying their mesenchymal origin. tbi was induced by stereotactic surgery under deep anaesthesia and subsequently icv transplantation of venus+ ascs was performed on adult wistar rats. normal ascs-transplanted and tbi-saline transplanted rats were used as controls. the proliferation, migration, survival and fate of transplanted ascs and their effect on injury restoration were examined six weeks post transplantation (pt). results: six weeks pt ascs expressed the fluorescence venus protein and therefore were identified in brain parenchyma. their presence into brain was also confirmed by masson trichrome staining, which revealed their collagen depositions. ascs were found in lesser numbers compared to those transplanted and exhibited no proliferative activity. ascs were found scattered distributed in brain as individual cells, and there were no aggregates of ascs or mass formation into lateral ventricles. extensive migration of ascs was mainly performed through white matter tracks in the corpus callosum and fimbria of hippocampus. six weeks pt ascs retained the characteristics of mesenchymal cells and did not differentiate into cells of neural lineage. ascs exhibited limited long-term survival, which is restricted in perivascular areas probably contributing to vascular formation. homing of ascs into peri-injured area was detected in half of the animals and achieved through the corpus callosum, as revealed by the collagen depositions, in this white matter track. transplanted ascs reduced the area of tbi cavity and did not enhance the astroglial scarring in peri-injured area. in tbi +ascs transplanted animals, the cortical injury site, showed a significantly smaller volume and lower % tissue loss compared to that of tbi+vehicle animals ( . ± . mm and . ± . % respectively, versus . ± . mm and . ± . %, p= . and p= . respectively). conclusions: considering the effects of ascs on inflammation and regeneration, we suggest that their transplantation after brain injury may promote host brain repair mechanisms. ascs transplantation may be beneficial in tbi, however some of its effects need careful and indepth evaluation. disclosure: nothing to declare xie-na cao , yuan kong , zhong-shi lyu , , qi wen , min-min shi , , qian-yu sun , yu-hong chen , yu wang , lan-ping xu , xiao-hui zhang , xiao-jun huang , background: poor graft function (pgf) remains a serious complication after allogeneic hematopoietic stem cell transplantation (allo-hsct). our previous work reported that abnormal bone marrow (bm) endothelial cells (ecs) were involved in the pathogenesis of pgf patients after allo-hsct (bbmt ; bmt ; blood ), but the explicit mechanism requires further clarification. autophagy is a self-degradative process responsible for the elimination of cytosolic components including proteins and damaged organelles. recent findings demonstrated that stimulation of autophagy could reduce oxidative status and angiogenic potential in ecsafter high-glucose exposure, from diabetic patients.however, little is known regarding the autophagy of bm ecs in pgf patients. therefore, the current study was performed to evaluate whether autophagy in bm ecs play a role in the pathogenesis of pgf. moreover, to investigate the effects of autophagic regulation on ecs and thereby regulating hematopoietic stem cell (hscs). methods: in the prospective case-control study, the autophagy levels were compared in bm ecs from pgf patients, and their matched good graft function (ggf) patients.the expression levels of autophagy-related markers (lc , beclin , and p ), and intracellular autophagosomes were detected by immunohistochemical staining, flow cytometry, western blot and transmission electron microscopy. subsequently, rapamycin (the autophagy activators) or hydroxychloroquine (hcq, the autophagy inhibitor) were administrated tothe -day cultivated bm ecs and human umbilical vein endothelial cells (huvecs), respectively.the autophagic vacuoleswere detected by monodansylcadaverine (mdc) staining assay. the bm ecsand huvecs were evaluated by cell counting, dii-ac-ldl and fitc-lectin-uea- double staining, migration, cell proliferation, and levels of reactive oxygen species (ros). to explore whether autophagy would affect the ability of bm ecs to support hscs in vitro, bm cd + cells from healthy donors were co-cultured with cultivated bm ecs and huvecs. colony-forming unit (cfu) and the apoptosis of co-cultured hscs were analyzed. results: the defective autophagy in bm ecs, characterized by decreased intracellular autophagosomes and autophagic vacuoles, decreased expression of lc -ii and beclin , and high level of p , were observed in pgf patients compared with ggf patients. moreover, the coculture of bm cd + cells with bm ecs showed significant deficient cfu plating efficiency, and increased apoptosis of cd + cells in pgf patients. in vitro upregulation of autophagy by rapamycin quantitatively and functionally improved bm ecsand huvecs, which manifested as more dii-ac-ldl and fitc-lectin-uea- double stained cells, increased capacities of migration, lower levels of ros and apoptosis via regulating beclin pathway, whereas inhibition of autophagy by hcq aggravated the huvecs and bm ecs from pgf patients. furthermore, in vitro upregulation of autophagy by rapamycin significant improved cfu plating efficiency, and decreased apoptosis in bm hscs co-cultured with huvecs and bm ecs from pgf patients. conclusions: these findings suggest that defective autophagy in bm ecs may be involved in the pathogenesis of pgf. the effect of rapamycin in pgfpatients is potentially mediated by improving the dysfunctional bm ecsto support hscs. therefore, it would be of value to investigate whether upregulating of cytoprotective autophagy of bm ecs may ameliorate pgf, thereby providing a novel clinical intervention for pgf in the future. clinical background: heparanase (hpse) in an endoβ-glucuronidase that specifically cleaves the saccaride chains of heparan sulphate proteoglycans (hs), leading to a loss of integrity of the extracellular matrix and to release of hs-bound cytokines, chemokines, angiogenic and growth factors. hpse gene is polymorphic and includes approximately snps. the combination of two snps, rs and rs , are involved in the regulation of hpse expression with an inverse correlation between mrna expression and protein levels: gg-cc, gg-ct, gg-tt, ga-cc (low group) expressed high hpse concentration; ga-ct and ga-tt (median group) expressed intermediate hpse levels; aa-tt and aa-ct expressed low hpse concentration (high group). we studied hpse snps in the allogeneic stem cell transplantation (hsct) setting to evaluate a possible association with post-hsct outcomes. methods: we enrolled patients submitted to hsct in our department since to . for each couple recipient-donor, rs snp was genotyped using restriction fragment lenght polymorphism assay, whereas for rs snp an allele-specific polimerase chain reaction was applied. hpse genotype distribution was compared in different groups according to post-hsct outcome: graft-versus-host disease (gvhd), transplantrelated mortality (trm), overall survival (os), infectious complication and disease-free survival (dfs). statistical analysis was performed using ncss . results: distribution of rs snp was as follows: gg . %, ga . % and aa . % among recipients and . %, . % and . % among donors, respectively. hardy-weinberg equilibrium (hwe) was respected. distribution of rs snp was as follows: cc . %, ct . % and tt . % among recipients and . %, % and . % among donors, respectively. rs snp distribution did not respect the hwe. an association was found between recipient rs snp and the cumulative incidence of agvhd among patients submitted to a reduced intensity conditioning (ric): . % for tt genotype and % for ct or cc genotype (p= . ). on the other hand, an association was identified between donor rs /rs snps combination and the cumulative incidence of agvhd: . % for low group donor, % for median group donor and . % for high group donor (p= . ). conversely, aa genotype for donor rs resulted independent risk factor for cgvhd de novo development (p= . , od . ) together to donor-recipient sex mismatch (female donor to male recipient vs. others: p= . , od . ) . considering cmv reactivation rate after hsct, an association was observed according to recipient rs snp: % for cc genotype, . % for ct genotype and . % for tt genotype (p= . ). multivariate analysis confirmed recipient rs snp as independent risk factor for cmv reactivation after hsct (p= . , od . ) together with recipient cmv serostatus at transplant (positive vs. negative: p< . , od . ). conclusions: hpse role was widely studied in the setting of inflammation, autoimmune diseases, hematological disease and tumor. however, it still remains debated the inducing or protective activity of hpse in the setting of gvhd. obviously, our results need to be confirmed in a validation cohort. clinical trial registry: na disclosure: nothing to declare novel protocol for autologous hsct in patients with high risk of complications: ambulatory chemomobilization and transplantation of fresh hematopoietic stem cells with backup storage background: autologous hematopoietic stem cell transplantation (ahsct) is standard of treatment in many patients with high risk of complications: dialysed patients, patients with heart and kidney amyloidosis or patients with systemic sclerosis. we introduced recently a novel protocol for ahsct: combination of ambulatory mobilization with very low doses of ara-c and g-csf connected with direct ahsct with fresh cells. this protocol allowed us to reduce the transplant risk in various patient groups traditionally connected with high risk of complications. in this work we summarize the experience in such high risk patients. methods: the prospectively collected database of patients after ahsct was searched for patients who underwent ahsct after chemomobilization with ara-c and transplantation with fresh cells and who fulfilled at least one study inclusion criteria: a) dependence on dialysis b) amyloidosis c) systemic sclerosis d) disqualification from transplantation at other centre due to the high risk of complications. there were together patients selected for this analysis - with amyloidosis ( with ≥ organs involved), dialysed, with systemic sclerosis, unfit at other centre. the database included prospectively recorded serious adverse events during the mobilization and transplantation. results: there were transplantations performed in this group of patients. mortality was % at days. all patients underwent successful ambulatory mobilization. all patients received mephalan conditioning with single infusion with median dose of mg/m (min , max ). mean engraftment was . days for white blood cells and . days for plt over g/l. the rate of complications was low with cases of neutropenic fewer, single bacterial culture with staphylococcus epidermidis without clinical signs of infection, median mucositis grade of . and without patients on parenteral nutrition. the median time of hospitalization was days (min , max ). conclusions: we present here novel protocol of transplantation combining chemomobilization and ahsct with fresh cells with excellent safety profile among most severely ill patients allowing for safe and efficient transplants. with this protocol we were able to overcome multiple risk factors and perform full intensity transplantation in very fragile patients. disclosure: nothing to declare single umbilical cord blood transplantation provides durable disease remission of advanced hematological malignancies in elderly patients background: although allogeneic hematopoietic stem cell transplantation (allo hsct) is potentially curative therapy in a variety of hematological malignancies, little has been reported of the outcome for elderly patients who are not in remission at transplantation. but it has been pointed out that recipient age alone can not be regarded as contraindication for allo hsct in the literature recently, supported by suitable donor, conditioning regimens and appropriate management of complications. we conducted a retrospective study of elderly patients who had advanced hematologic malignancies to elucidate the outcome of single umbilical cord blood transplantation (sucbt) in toranomon hospital kajigaya, japan. methods: we retrospectively investigated the outcomes of patients aged over who underwent their first ucbt from june to december in our medical center. results: diseases included acute myelogenous leukemia (n= ), myelodysplastic syndrome (n= ), adult t-cell leukemia/lymphoma (n= ), myelofibrosis (n= ) and chronic lymphocytic leukemia (n= ). the median age at transplantation was years (range, - ) and follow-up for survivor post transplantation was day (range, - ). all patients were not in complete remission (cr) at the time of transplantation. reduced intensity conditioning (ric) regimens were used in patients. all patients received tacrolimus and mycophenolate mofetil as graftversus-host disease (gvhd) prophylaxis. all cases except early death achieved neutrophil recovery at median days (range, - ). at year, overall survival (os) rate and disease free survival (dfs) were , % ( % confidence interval (ci), . - . ). we performed univariate analysis to identify the factor that influenced os at year, but no statistical significance was demonstrated at the age of transplantation (aged to vs. ≧ , . % ( % ci, . - . ) vs. . % ( % ci, . - . ), p= . ). the cumulative incidence of non-relapse mortality (nrm) at days was . % ( % ci, . - . %) and relapse at year was . % ( % ci, . - . %). only two patients developed acute gvhd(ii-iv) and one developed severe gvhd at days after transplantation. the main causes of death was infection (n= ), including sepsis (n= ) and viral encephalitis (n= ), followed by idiopathic pneumonia syndrome (n= ) and thrombotic microangiopathy (n= ) during the early phase of transplantation. in contrast, no patients died of recurrence. conclusions: although our report consisted relapsed/ refractory disease of elderly patients at the time of sucbt, durable remission and lower incidence of gvhd could be noteworthy compared with previous reports. further strategies to reduce the rate of nrm and longer duration of follow up would be warranted. disclosure background: pearson syndrome and kearns-sayre syndrome are metabolic disorders caused by a de-novo deletion in the mitochondrial dna (mtdna). allogeneic stem cell transplantation has shown to improve metabolic function in distal organs in several metabolic disorders, but bears significant morbidity and mortality, especially for patients with mitochondrial disorders. novel gene therapies may correct diseases rising from genomic dna mutations, but targeting the mitochondrial dna is complex. mitochondria are able to transfer into cells and between cells, as seen in preclinical models of mitochondrial and other metabolic disorders. here, we introduce a novel concept of mitochondrial augmentation therapy (mat) of autologous cd + cells in children with mitochondrial deletion syndromes. methods: patients were treated under a compassionateuse program, approved by the sheba medical center irb and the israeli ministry of health. briefly, mobilization was performed using gcsf alone (n= ) or in addition to plerixafor (n= ) . cd + cells were isolated via miltenyi clinimacs system and co-cultured with maternal mitochondria, drawn from peripheral blood and confirmed nondeleted, for hours, and re-infused to the patient without any conditioning. patients were followed for clinical and metabolic parameters. results: all four patients presented with different deletions in mitochondrial dna, and different baseline characteristics, and were treated at the age of . , , and years. despite normal cbc, significant bone marrow hypocellularity was seen in evaluated patients ( %, % and % cellularity at age , and ), which correlated with low colony forming unit capacity of patients and low yield of cd + mobilization in the leukapheresis product. patients received on average x enriched cells/kg (range, . - . ), and the median enrichment of cd + cells was % (range, - %). no infusion reactions occurred, and the only severe adverse events of this cellular therapy were leukapheresis-related anemia, hypokalemia, hypocalcemia and alkalosis, all resolved promptly with proper supplementation. follow-up duration is variable, ranging - months. we were able to show improvement in mitochondrial heteroplasmy (proportion of deleted mtdna of total mitochondrial dna) and in normal mtdna content, starting - months from cell therapy, which correlated with improved atp production in peripheral blood derived mononuclear cells. clinically, patients showed improvement in aerobic function and endurance (measured by the half-bruce protocol, sit-to-stand test and -minute walk test), muscle strength (hand-held dynamometry), and in quality of life, measured by the international pediatric metabolic disability scale. no metabolic crises occurred following cell infusion. conclusions: patients with deletion in mtdna have metabolic dysfunction, including poor bone marrow cellularity and function. hematopoietic stem cells in patients with mtdna deletions can be enriched with normal mitochondria, via mat, as first shown in our patients. this novel process is safe and results in increase in the normal mtdna in peripheral blood of patients, and in improved metabolic and clinical function. clinical trial registry: clinicaltrials.gov nct disclosure: moria blumkin, noa sher and natalie yivgi ohana -minovia therapeutics, employment p high cytotoxic efficiency of alpharetrovirally engineered cd -specific chimeric antigen receptor natural killer cells for treatment of acute lymphoblastic leukemia stephan müller , tobias bexte , annekathrin heinze , franziska schenk , axel schambach , winfried s. wels , , ute modlich , evelyn ullrich , background: autologous chimeric antigen receptormodified (car) t cells with specificity for cd showed potent antitumor efficacy in clinical trials regarding relapsed and refractory acute lymphoblastic leukemia (all). natural killer (nk) cells are cytotoxic lymphocytes that are capable to kill their targets in a non-specific manner and additionally do not cause gvhd. therefore, using cd -car-nk cells exhibits several advantages, such as safety in clinical use, possible allogenic settings and the potential to also attack heterologous leukemia cells which lost cd . previous approaches used cd -car-nk cells pre-stimulated by feeder cells, bearing potential risks. thus, we focused on the optimization of generating cd -car-nk cells by viral transduction under feeder-cell free conditions. methods: human nk cells were isolated from healthy donor peripheral blood mononuclear cells via cd negative selection. after a feeder-cell free expansion phase with interleukin , transductions were performed with an egfp or a cd -car encoding vector at different multiplicities of infection (moi). to optimize gene modification different transduction enhancers (retronectin and vectofusin- ) and viral vector systems (lentiviral and alpharetroviral) were compared. finally, generated cd -car-nk cells were tested in their ability to kill cd positive and cd -negative cell lines. results: nk cells transduced with a lentiviral egfp encoding vector or a lentiviral cd -car vector using retronectin and vectofusin- showed similar transduction efficiencies for both transduction enhancers (egfp: retronectin moi : . %; vectofusin- moi : . %; cd -car: retronectin moi : . %, moi : . %; vectofusin- moi : . %, moi : . %). the generated cd -car-nk cells showed increased cytotoxic capacity against cd -positive cells compared to nontransduced (nt) nk cells ( . % vs. . %, effector to target (e:t) ratio : ). both nk cell populations were equally efficient in killing cd -negative cells ( . % vs. . %). alpharetroviral transduction of nk cells with an egfp encoding vector showed higher transduction rates with vectofusin- than with retronectin (retronectin moi : . %, moi : . %; vectofusin- moi : . %, moi : . %). further using vectofusin- , similar transduction efficiencies could be achieved with an alpharetroviral cd -car encoding vector (moi : . %, moi : . %, moi : . %), outperforming the efficiencies of lentivirally generated cd -car-nk cells in the same experiments (moi : . %, moi : . %, moi : . %). additionally, alpharetroviral cd -car-nk cells showed a higher cell killing activity against cd -positive cells than lentiviral cd -car-nk cells or nt-nk cells ( . % vs. . % vs. %, e:t ratio : ). interestingly, similar killing activities were achieved with an e:t ratio of . : ( . % vs. . % vs. . %) and alpharetroviral cd -car-nk cells remained a stable cytotoxicity level at lower cell concentrations down to an e:t ratio of . : . all three nk cell populations were equally efficient in killing cd negative cells ( . % vs. . % vs. . %, e:t ratio : ). conclusions: cd -car-nk cells can be successfully generated under feeder-cell free conditions using different transduction enhancers and viral vector systems. these data suggest the usage of vectofusin- in combination with alpharetroviral vectors to genetically modify nk cells to achieve sufficient amounts of transduced cells. these cd -car-nk cells mediate high cytotoxicity and therefore may offer a new therapeutic option in the treatment of all. disclosure: axel schambach is an inventor on a patent describing alpharetroviral sin vectors. winfried s. wels is an inventor on a patent describing chimeric antigen receptors with an optimized hinge region. the remaining authors have nothing to disclose. graft-versus-host diseaseclinical walter spindelböck , bianca huber-krassnitzer , barbara uhl , gregor gorkiewicz , hildegard greinix , christoph högenauer , peter neumeister background: steroid-refractory acute gastrointestinal (gi) graft-versus-host disease (agvhd) is a severe complication of allogeneic hematopoietic stem cell transplantation (allo-hsct) associated with a high mortality rate. loss of intestinal bacterial diversity is thought to be associated with severity of gi-agvhd and an impaired intestinal microbiota with reduced diversity is an independent predictor of mortality. methods: the fecal microbiota transplantation (fmt) procedures were performed according to a protocol approved by the local ethical committee ( - ex / ) after obtaining informed consent. donors were healthy adult subjects screened for potential infections by serologic and microbiologic tests according to local standards. donor stool was diluted with saline and homogenized to a volume of~ ml fecal solution for instillation into the terminal ileum and caecum via colonoscope. microbiota sequencing analysis of s rdna was performed before fmts and afterwards at predefined timepoints. results: we report the outcome of nine patients refractory to - lines of immunosuppressive therapies with lower gi-stage iii (n= ) or iv (n= ) agvhd following repetitive fmts from a single donor. all patients had received an allo-hsct for mds (n= ) , aml (n= ), pmf (n= ) and mm (n= ) following a reduced intensity (n= ) or mac (n= ) conditioning regimen using pbsc as stem cell source. after an onset of lower gi agvhd between - days after allo-hsct, nine patients refractory to several lines of immunosuppressive therapies received - fmts ( patients were treated with more than fmts, in patients fmt was only administered once or twice) mostly in weekly intervals. five patients achieved a clinical complete response with resolved diarrhea and no gastrointestinal complaints, and four of these could be discharged without gvhd symptoms. two patients (pr, nc) were discontinued after or fmts in pr or nc due to concomitant infections (metapneumoviral pneumonia, cmv gastroenteritis), the other non-responders succumbed to gvhdrelated infectious complications. the establishment of donors' microbiota with the emergence of new taxa, an increase in bacterial richness/diversity, and the disappearance of the "enterococcus signature" were associated with disease control and response to fmt. except the possible transmission of adenovirus by fmt in one patient, no other immediate procedure-related infections or other side effects were observed. conclusions: restoration of dysbiosis by fmt might represent a promising novel therapeutic approach for a subset of patients with refractory lower gi-agvhd. vigorous donor screening for infectious disease is mandatory. clinical background: migration of allo-activated donor effector tcells from lymphoid tissues to target organs is an important step in acute graft versus host disease (gvhd). the sphingosine- -phosphate- (s p ) receptor plays a crucial role in lymphocyte trafficking. data from animal models suggest that pharmacological modulation of the s p receptor reduces gvhd and improves mortality. we investigated this mode of action by using the secondgeneration s p modulator krp for the prophylaxis of gvhd in a pilot clinical trial in patients undergoing allogeneic hsct. methods: a multi-centric, phase b, prospective, open label, two-part study was conducted to evaluate the safety, tolerability and pharmacokinetics of krp in patients undergoing allogeneic hsct for hematological malignancies. primary endpoint was safety. initial efficacy was explored based on the incidence of gvhd, mortality and relapse. part was a single arm open label study to investigate the safety of mg/day krp added to standard of care gvhd prophylaxis (csa/mtx) in patients. part was a randomized two-arm open label study to compare the safety, efficacy and pk of mg/day of krp in combination with tacrolimus/mtx to mg/day of krp in combination with csa/mtx in patients. in both parts, treatment with krp was initiated days before hsct and continued for an additional days. patients were followed up for up to years. results: patients were included in the study. of patients completed the -day treatment with krp at the assigned doses. median duration of follow-up was days (range to days). krp was safe and well tolerated. serious adverse events (saes) suspected to be related to krp were observed. macular edema (n= ) and peripheral edema (n= ) as s p related adverse events occurred and resolved without sequelae. of note, the incidence of macular edema in hsct recipients is unknown. neutrophil engraftment was confirmed in all patients with a median of days (range to days). of patients presented with grade iii or iv acute gvhd (on days , , , and ) . no gvhd or infection related death occurred during the first days. -day survival was %, with no death occurring during krp treatment. death occurred on study day due to lymphoma relapse. a second death occurred on study day due to liver gvhd. four patients died in the follow-up period due to gastrointestinal gvhd (day ), aspiration pneumonia (day ) and relapse (day and day ). the kaplan-meier estimate of overall survival at year was . . when comparing the data from the two dose groups ( and mg krp ), no major differences in safety, engraftment, gvhd rate or mortality were observed. conclusions: this clinical trial was the first to test s p modulation in this population. our data suggest that krp had no negative impact on engraftment and overall, was safe, and well tolerated. based on exploratory data, when comparing to matched historical mortality data, krp may have favorable effects on overall survival ( figure ). background: uric acid is a danger signal contributing to inflammation. relevance to allosct has been demonstrated in preclinical models: the depletion of uric acid led to improved survival and reduced gvhd (j exp med. sep ; ( ): - ). results of a clinical pilot trial suggested that peri-transplant uric acid depletion reduce acute gvhd incidence (bbmt may; ( ): - ). methods: this international multicentric study aimed to study the association of uric acid serum levels before start of conditioning with allosct outcome. patients with acute leukemia, lymphoma or mds receiving a matched sibling allosct for the first time were considered for inclusion, regardless of conditionning. data were prospectively collected between / and / . a comparison of outcomes between patients with high and low uric acid level was performed using univariate analysis and multivariate analysis using cause-specific cox model. variables included in the multivariate analyses were age, sex mismatch, diagnosis, disease status, karnofsky score, number of cd cells given, intensity of conditioning, type of gvhd prophylaxis, atg use, time from diagnosis to transplant, year of transplant and cmv status. results: twenty centers from european countries reported data on allosct recipients. patient characteristics are given in table . the uric acid cut off point was determined at . mg/dl (median of measured uric acid levels). overall survival (os) and progression free survival (pfs) of allosct recipients with uric acid levels above cut off measured before start of conditioning were significantly shorter ( figure a , os univariate hr= . ci= . - . p< . ; multivariate hr= . , ci= . - . , p< . ) ( figure b , pfs univariate hr= ci= . - . p= . ; multivariate hr= . , ci= . - , p= . ). nonrelapse mortality was significantly increased in allosct recipients with high uric acid levels prior to start of conditioning (univariate hr= ci= . - . p= . ; multivariate hr= . , ci= . - . , p= . ). in addition, there was a non-significant trend towards higher acute gvhd incidence (gvhd grades ii-iv univariate hr= . ci= . - . p= . ; multivariate hr= . ci= - . , p= . ) in allosct recipients with uric acid levels above cut off before transplantation. finally, the incidence of relapse after allosct was moderately increased in the cohort with higher uric acid levels (univariate hr= . ci= - . p= . ; multivariate hr= . , ci= . - . , p= . ). conclusions: high uric acid levels before start of conditioning correlate with high mortality after allosct. our results can serve as rationale for clinical trials on depletion of uric acid during allosct. results: we found significant correlation between donors' ctla- + a>g polymorphism and hsct outcome. genotype aa was present in donors, ag in donors and donors was homozygous for g allele. recipients who received graft from g allele carrier donors showed significantly increased cumulative incidence of relapse (at months aa: . %, ag: . % and gg: . %; p= . ). on contrary, the frequency of the acute gvhd grades iii-iv and cytomegalovirus (cmv) reactivation/disease decreased according to the presence of the g allele in the donor ctla- genotype [agvhd: aa: %, ag: %, gg: %; p= . ; cmv: aa: %, ag: %, gg: %; p= . ]. cumulative incidence of agvhd was also markedly decreased among patients with g allele carrier donors (at days aa: . %, ag: . %, gg: . %; p= . ). donor genotype similarly influenced hsct outcome in mud donor and mac conditioning subgroups. overall survival (os) was not different in patient subgroups according to donor genotypes [os at months: aa: . ± . %, ag: . ± . %, gg: . ± . %; p= . ]. we did not find any correlation between recipients' ctla- + a>g polymorphism and hsct outcome. conclusions: several ctla- snps have previously been described to be associated with relapse rate, incidence of agvhd and os, but results are often contradictory in the publications. in our study, ctla- + a>g polymorphism of hsct donors influenced risk of relapse, agvhd, cmv and cause of death, but not overall survival. the genotyping of ctla- + a>g polymorphism in donors may help in the risk assessment process and the choice of personalised therapy. disclosure: nothing to declare. background: although steroids remain first-line therapy for the treatment of acute graft versus host disease (agvhd), response rates in patients with grade iii-iv disease are poor, with no apparent improvement in survival over the past years. we performed a prospective, multicenter trial to assess the efficacy and safety of the combination of ruxolitinib and etanercept as a novel approach to treat grades iii-iv sr-agvhd . methods: forty malignant hematologic disease patients with grades iii-iv sr-agvhd after allo-sct from three centers in east china were enrolled from january to june . ruxolitinib was initiated at a dose of - mg bid for months, and then tapered gradually for another one month. etanercept was administrated at mg biw for - weeks. results: the median age of patients was (range - ) years. at day after the combination treatment, the overall response rate (orr) was % including crs ( %) and prs ( %). the median time to the optimal response was (range - ) days. the incidences of cr per organ were . %, . %, and % for skin, liver, and gut, respectively. the agvhd relapse rate was analyzed for the patients who had achieved cr or pr and survived beyond days. relapses in agvhd occurred in . % ( / ) of responsive patients. the patients who received ruxolitinib within days after agvhd onset have a significant higher cr rate that those with delayed ruxolitinib therapy ( . % vs. . %, p= . ). and the patients without gut infections have a significant higher cr rate than infected cohort ( . % vs. . %, p= . ). by logistic regression analysis, the time from agvhd to ruxolitinib (rr= . , p= . ) and gut infection (rr= . , p= . ) were independent predictors for incomplete response. thirteen patients ( / , . %) suffered from at least infectious episode after the start of the combination therapy, and pulmonary infectious diseases was a frequent complication ( / , . %). iii-iv cytopenia and cmvreactivation were observed in % and . % of patients. the -year overall survival (os) after initiation of the combination therapy were . %. the -year nrm and relapse incidence was . % and . %, respectively. patients with complete response on day had significantly higher os probability than non-cr patients ( -year os: . % vs . %, p= . ). compared with the historical cohort of basiliximab and etanercept for sr-agvhd in our center (n= ), no significant difference was found on the baseline. although the orr in patients treated with ruxolitinib and etanercept is identical with the historical cohort, ruxolitinib group achieved rapider remissions in liver agvhd and gut agvhd than the historical cohort (gut agvhd: days vs. days, p= . ; liver agvhd: days vs. days, p= . ), thus, with regard to hospital stay after agvhd onset, the ruxolitinib cohort stayed shorter (median: days vs. days, p= . ) than basiliximab cohort. conclusions: combined treatment with ruxolitinib and etanercept resulted in a rapid cr to visceral agvhd and meanwhile reserve graft anti-leukemia (gvl) effect as the relapse rate of primary disease is relatively lower. the various infection complications associated with ruxolitinib merit more attention. disclosure: nothing to declare background: graft-versus-host disease (gvhd) remains one of the main life-threatening complications after allo-hsct, especially in patients with non-malignant diseases. the standard gvhd prophylaxis strategy is mostly based on the use of calcineurin inhibitors alone or in combination with other immunosuppressive (is) post-transplant cyclophosphamide (ptcy) is effective gvhd prophylaxis optiont for adult patients (pts), but has limited data in children. methods: the study aim was to evaluate ptcy as gvhd prophylaxis in pediatric pts with inherited disorders undergoing allo-hsct. pts, the most of them are pediatric age (median age - y.o., range month - y.o.) with different types inherited disorders (β-thalassemia - , bone marrow failure syndromes - , storage diseases - , primary immunodeficiencydisorders - ) were inrolled in retrospective study. donor type was: matched/mismatched unrelated (mud/mmud) - , matched related donor (mrd)- , haploidentical (haplo) - . conditioning regimen was: myeloablative (mac) - , reduce-intensity (ric) - . graft sourse was: bone marrow (bm) - , peripheral blood stem cells (pbsc) - , combintions bm +pbsc/bm+cord blood - . ptcy mg/kg days + , + based gvhd prophylaxis recived pts., standart gvhd prophylaxis based on calcineurin inhibitors - pts. results: cumulative incidence (ci) of agvhd was %. grade - , - agvhd were % and % respectively. ptcy based gvhd prophylaxis reduced ci of agvhd ( % vs %, p= , ). another reduce ci of agvhd factors were mac ( % vs % in ric pts group, p= , ), mrd ( % vs % in haplo group vs % in mud/mmud group, p= , ), bm as a transplant source ( % vs % in pbsc group, p= , ). in a multivariate analysis mac (hr , %ci , - ,, p= , ), time from diagnosis to allo-hsct less then month (hr , %ci , - , , p= , ) were predictive for reducing ci agvhd. for agvhd - st. significant factor increase ci was female donor both in univariate ( % vs %, p= , ) and multivariate analysis (hr , %ci , - , , p= , ). years overall survival (os) was %. improving os factors were: transplant age younger then y.o. ( % vs %, p= , ), time from diagnosis to allo-hsct less then month ( % vs %, p= , ), engraftment ( % vs %, p= , ). in a multivariate analysis only transplant age younger then y.o. (hr , %ci , ( ) ( ) ( ) ( ) ( ) p= , ) and engraftment (hr , %ci , - , , p= , ) were predictive for os. conclusions: ptcy-based gvhd prophylaxis can be effective options for reduce risk of acute gvhd. using unrelated donors, bone marrow as transplant source and mac can reduce ci of gvhd. performing allo-hscr earlier from diagnos and in earlier age can improve os patients with inherited disorders background: diarrhea is a frequent complication after allo-sct. at onset it is often difficult to differentiate gi gvhd from other causes of enterocolitis. recently, non-invasive tests, such as fecal calprotectin (fc), have been validated as markers of gut inflammation in patients with inflammatory bowel disease, but only a few studies have been published regarding its use as a diagnostic marker in gi gvhd. methods: our aim in this study was to explore the levels of fc in allo-sct recipients with new-onset diarrhea. so far we have included allo-sct recipients who developed acute diarrhea ≥ stage - at a median of days (range: - ) post allo-sct. stool samples were analyzed as soon as possible after the onset of diarrhea. fc levels were determined in addition to an extensive microbiological panel for infectious enterocolitis (including norovirus pcr and c. difficile associated diarrhea). endoscopies for histologic analysis were performed according to the treating physicians' discretion (n= ). results: patients characteristics are summarized in table . median follow-up for survivors was days (range: - ). twenty-eight patients ( %) were diagnosed of gi-gvhd. the additional causes of diarrhea were: drug-related enterotoxicity (n= ), viral enteritis (n= ), food intolerance (n= ), c.jejuni-enteritis (n= ), and non-specific causes (n= ). the concentration of fc was higher in patients with gi gvhd vs. other causes of diarrhea ( μg/g +/- vs. μg/g +/- , p= . ). patients who did not develop severe enterocolitis had normal to slightly raised calprotectin at the onset of diarrhea [< - in out of ( %) cases], including % ( / ) of patients with enterotoxic drug-related diarrhea. among the patients with gi-gvhd, ( . %) were later found to be steroid-resistant. as shown in figure , we found a significant association between high fc (≥ μg/g) and severe-refractory gvhd (hr . , p= . ). of note, high values of fc were also found in patients with severe infectious enteritis (norovirus, adenovirus and c.jejuni infections), with baseline fc> μg/g, respectively. overall survival was % (ic %: - ) at months. hypoalbuminemia and thrombocytopenia were the only variables linked to -yr os in univariate analysis, regardless of the cause of enterocolitis. conclusions: in the absence of standarized (and expensive) biomarker panels for analyzing and predicting gvhd onset and outcomes, the fc test may be an useful tool in the allo-sct setting. our initial results show that fc is helpful in predicting mild causes of diarrhea and to identify patients with a high probability of developing severe (and potentially steroid-refractory) gi gvhd, although high levels are also found in severe infectious enteritis. background: there is an urgent need for effective therapy for severe acute gvhd. results of gvhd therapies beyond months are rarely reported. we here report a median follow-up of years. we introduced mesenchymal stromal cells as therapy for severe acute gvhd, with a dramatic response in some, but not all patients. the placenta protects the fetus from the mothers haploidentical immune system during pregnancy. we found that maternal stromal cells from the fetal membrane, so called decidua stromal cells (dscs) were more immunosuppressive than other sources of stromal cells. methods: we treated patients, median years of age (range . - ) for severe acute gvhd. all had biopsy proven gastro-intestinal gvhd. all were steroid refractory, after > days or with progression and after > days. we used an improved protocol where dscs were thawed and infused in a buffer with % albumin. dscs were given at a median dose of . ( . - . ) x cells/kg and ( - ) doses, given one week apart. viability of frozen and thawed dscs was % ( - ) and cell passage was ( - ). results: complete resolution of gvhd was seen in patients and had a partial response. the cumulative incidence of chronic gvhd was %. six had mild, moderate and one severe nih overall gvhd severity scoring. nine patients died, from relapse, acute gvhd and septicemia, zygomycetes infection, liver insufficiency, cerebral hemorrhage, multiorgan failure and chronic gvhd with obstructive bronchiolitis. four years transplant related mortalliy was . % and overall survival was %. survival was not significantly worse (p= . ) than % for all patients undergoing allogeneic hematopoietic cell transplantation during the same period - . conclusions: to conclude, dscs seems to be a promising therapy for severe acute gvhd. randomized trials are under way. disclosure: nothing to declare p anti-apoptotic protein bcl- is upregulated in graftversus-host disease stem cell transplantation (allo-hsct) with - % developing either acute or chronic gvhd. recently, bcl- inhibitor venetoclax was approved for treatment of chronic lymphocytic leukemia. induction of apoptosis and depletion of lymphocyte subpopulations e.g. follicular b-cells or cd + and cd + t-cells led to further exploration in autoimmune disease. methods: to establish expression levels of genes in the bcl- pathway, low-input rna sequencing was performed on t cells isolated from non-inflamed skin and peripheral blood of hsct recipients at different time points before until year after transplantation. furthermore, we analyzed blood, lung, gut and skin samples of patients post allo-hsct with and without previously untreated acute or chronic gvhd by rt-pcr, flow cytometry and tissue immunofluorescence. [[p image] . bcl- is up-regulated in t and b lymphocytes of acute and chronic gvhd lesions.] results: rna-sequencing revealed that t cells upregulated bcl- upon conditioning treatment (day ) and cells of patients who later developed gvhd failed to downregulate bcl- after transplantation (day+ , day+ ). bcl- protein levels were elevated in overall leukocytes and pathogenic cell subsets including monocytes, cd + t lymphocytes and nkt cells showed significantly higher expression of bcl- in peripheral blood of gvhd patients as compared to healthy controls. these results could be recapitulated in tissue samples, where disease-promoting lymphocytes (t, b, nk, nkt) were numerically expanded and expressed bcl- in acute and chronic gvhd skin lesions. notably, non-pathogenic cell types such as keratinocytes did not exhibit increased bcl- expression compared to control samples from hsct recipients and healthy donors. while bcl- rna expression did not depend on type of conditioning (mac vs. ric) or gvhd grade, it correlated to disease severity and was significantly elevated in biopsies of patients with steroidrefractory gvhd. conclusions: we could show exclusive upregulation of bcl- in gvhd-mediating cell types in peripheral blood and tissue samples affected by gvhd, correlating to gvhd severity and response to first-line therapy. thus, bcl- inhibition may present a novel and urgently needed targeted therapy in treatment of steroid-refractory acute and chronic gvhd. disclosure: supported by a docmed fellowship od the austrain academy of sciences background: graft-versus-host disease (gvhd) represents a major contributor to morbidity and mortality in recipients of allogeneic hematopoietic cell transplants (hct). several therapeutic strategies exist for gvhd prophylaxis and include post-transplant cyclophosphamide (ptcy) and antithymocyte globulin (atg). while several groups have described the use of ptcy in younger patients, there is a paucity of data about the efficacy of ptcy in older individuals, particularly when combined with atg. we investigated the combined effect of ptcy with atg on transplant outcomes in older patients at princess margaret cancer centre, toronto, canada. methods: this retrospective study included all patients age ≥ who underwent allogeneic hct for any indication at our centre between december and july . overall survival (os) was calculated using kaplan-meier analysis and multivariable cox proportional hazards regression. cumulative incidence of relapse (cir) and non-relapse mortality (nrm) were calculated using competing risk regression (fine and gray method). incidences of acute (agvhd) and chronic (cgvhd) were compared using the fisher's exact test. results: of patients, ( %) were male. median age was (range - ) and median follow-up among survivors was months (range - ). acute myeloid leukaemia (aml) was the most common indication for hct ( patients, %), followed by myelodysplastic syndrome ( patients, %) and myelofibrosis ( patients, %). eightyfour ( %) patients had a matched unrelated donor, ( %) had a matched related donor and ( %) had a haploidentical donor. one hundred twenty-five ( %) patients received reduced intensity conditioning. sixty-two ( %) patients received ptcy combined with atg ( . mg/kg) while ( %) received other forms of gvhd prophylaxis. os at years was % ( % confidence interval (ci) - ) in the entire cohort. patients who received ptcy with atg had a superior -year os compared with other gvhd prophylaxis regimens ( figure a ): % ( % ci - ) vs. % ( % ci - ), respectively (hr= . , % ci . - . , p= . ). the -year nrm for the entire cohort was % ( % ci, - ). patients who received ptcy with atg had a lower -year nrm compared to those who did not ( figure b ): % ( % ci - ) vs. % ( % ci - ), respectively (hr= . , % ci . - . , p= . ). the -year cir in the whole group was % ( % ci - ). use of ptcy with atg was associated with a modest increase in cir at two years ( figure c ): % ( % ci - ) vs. % ( % - ), respectively (hr= . , % ci . - . , p= . ). there was a trend toward lower incidence of grade ii-iv agvhd among patients who received ptcy with atg compared to those who did not: % vs. % (p= . ). the incidence of grade ii-iv cgvhd was lower in individuals who received ptcy with atg compared to those who did not: % vs. % (p= . ). conclusions: in older hct recipients, use of ptcy combined with atg is associated with improved os, lower nrm, decreased risk of both agvhd and cgvhd and a modest increase in relapse risk. therefore the ptcy with atg combination represents an effective strategy for gvhd prophylaxis in older allogeneic hct recipients. disclosure: the authors have no conflict of interest to declare. outcome of severe graft versus host disease in pediatric patients with nonmalignant diseases after allogeneic bone marrow transplantation. a single center experience irina zaidman , sigal grisariu , batia avni , ehud even-or , bella shadur , adeeb nasereddin , polina stepensky background: hematopoietic stem cell transplantation (hsct) remained the only curative option for many nonmalignant diseases in pediatric patients. survival after hsct has improved the last few years due to significant advancement in human leukocyte antigens (hla) typing techniques, less toxic conditioning regimens and better supportive care and resulted to % survival and cure in some non malignant diseases. graft-versus-host disease (gvhd) remains a major complication of hsct and leading cause of morbidity and mortality. prognosis of patients with high grade gvhd is dismal and survival rate varies between % to % in pediatric patients. methods: the retrospective study included patients with non malignant diseases who underwent allogeneic hsct at hadassah medical center from to . the collected data included patient´s clinical data and transplant characteristics. the study was approved by the institutional helsinki committee. results: children with nonmalignant diseases underwent allogeneic bone marrow transplantations in hadassah university hospital during ten years period. fifty seven patients ( %) developed agvhd grade - , twenty five of them ( . %) grade - . median age was . (range . - . ), most patients were males ( males, females). patients underwent bmt from fully matched family members, children were transplanted from matched unrelated donors and from mismatched donors. twenty one of patients with severe gvhd ( %) survived. four patients ( %) died from severe gvhd and complications of immunosuppressive treatment. of deceased patients were transplanted from mismatched donor, in of cases the age of donor was advanced, of patients developed severe gvhd and died after second hsct. all patients were refractory to different treatment modalities. three of patients died in and one in , it was no death from severe gvhd in patients that were transplanted and developed high grade gvhd after . conclusions: the results of this study show a high survival rate of % in pediatric patients with non malignant diseases and severe gvhd. significant risk factors for mortality in our group included mismatched donor, advanced age of donor and second transplant. trend to better survival was observed after . additional multicentral studies analyzed the outcomes of agvhd in pediatric patients with nonmalignant diseases are urgently required. background: chronic graft-versus-host disease (cgvhd) is a serious late complication after allogeneic hematopoietic stem cell transplantation (allohsct) with heterogeneous presentation and still poorly understood pathophysiology including inflammation and endothelial dysfunction. factor viii (fviii) and von willebrand factor (vwf) are coagulation factors but also known indicators of endothelial dysfunction and inflammation in different settings, and therefore could serve as interesting candidate biomarkers of cgvhd. methods: since patients after allohsct were assessed by the multidisciplinary cgvhd team at the university hospital center zagreb, croatia, using established nih cgvhd-related measurements. an extensive history, physical and laboratory evaluations were performed, including fviii, vwf:ag and vwf:ac analysis. descriptive statistic and non-parametric analyses were performed. variables that showed significant univariate correlations were used in multivariate logistic regression (mlr) to identify the most predictive for fviii, vwf:ag and vwf:ac in cgvhd patients. results: cgvhd patients and controls (subjects after allohsct without cgvhd) were analysed. median age of cgvhd patients was ( - ) years, % females, . % underwent allohsct for hematologic malignancies, . % had myeloablative conditioning and . % matched related donor. median time from hsct to study was . days and from cgvhd diagnosis to study days. there were no demographic neither transplant related significant differences between cgvhd patients and controls beside stem cell source (peripheral blood . % vs . %, p= . ) and history of acute gvhd ( . % vs . %, p< . ). majority of patients had moderate ( . %) or severe ( . %) nih global cgvhd score, . % active cgvhd by clinician´s impression. median number of organs involved by cgvhd was ( - ), and the most frequently involved organs were mouth, skin and eyes ( . % each). cgvhd patients compared to controls had higher fviii levels (median ( - )% vs ( - )%, p= . , reference range - %) and higher vwf:ag (median . ( . - )% vs . ( . - )%, p= . , reference range - %), while vwf: ac showed a trend toward higher levels among patients (median . ( - )% vs ( . - )%, p= . , reference range - %). patients had higher ggt (p= . ), lower anticardiolipin igg (p= . ) and igm (p= . ), and lower albumin (p= . ) than controls, without differences between other laboratory parameters. univariate analysis showed that among cgvhd patients higher fviii was associated with worse karnofsky score (ks) (p= . ) and performance score (ps) (p= . ), higher leukocytes (p= . ), cholesterol (p= . ), triglycerides, ast, alt, ggt, ldh, and lower albumin. higher vwf:ag and vwf:ac in cgvhd patients were associated with worse ks and ps (p< . ), with more active cgvhd (p< . ), worse nih cgvhd liver (p= . ; p= . ) and nih cgvhd mouth (p= . ; p= . ), higher total nih score (p= . ; p= . ), higher number organs involved (p= . ; p= . ), higher esr, monocytes, ddimers, ast, alt, ggt, ldh, triglycerides, β- -microglobulin, ferritin, total proteins, iga and lower albumin. mlr analysis showed leukocytes (p= . ) and cholesterol (p= . ) as the strongest predictor of fviii (r = . %; p< . ), while strongest predictor of vwf: ac was number of organs involved by cgvhd (r = . %; p= . ). conclusions: results of this study detected high fviii and vwf levels in cgvhd patients with possible reflections to cgvhd manifestations, what needs to be further confirmed in larger longitudinal studies. disclosure: this work was supported, in part, by the unity through knowledge fund project entitled "clinical and biological factors determining severity and activity of chronic graft-versus-host disease after allogeneic hematopoietic stem cell transplantation", and also, in part, by the croatian science foundation project entitled "new biomarkers for chronic graft-versus-host disease". antonela samardzic -work financed by the croatian science fondations`"young researchers`career development project -training of doctoral students" background: thrombotic microangiopathy (tma) is a severe complication of allogeneic hematopoietic cell transplantation (hct) with multisystem involvement. a few recent reports have recognized evidence of tma in the intestinal vasculature (intestinal tma/itma) of patients with graft-versus-host disease (gvhd) with or without tma. we aimed to identify patients with itma and describe histological, clinical and prognostic features. methods: we prospectively evaluated available endoscopic samples (stomach and/or colon) from consecutive adult hct recipients for previously described histopathologic signs of itma (january -september ). systemic tma was diagnosed according to the international working group criteria. we compared findings among clinical groups: gvhd/systemic tma, gvhd/no systemic tma and no gvhd/no tma. results: we studied patients, classified as gvhd/ systemic tma, gvhd/no systemic tma and no gvhd/no tma. baseline transplant characteristics (age, donor, hla matching, conditioning) did not differ significantly among groups. histological features of itma, including loss of glands, total denudation of mucosa, apoptosis and detachment of endothelial cells, intraluminal fibrin, intraluminal microthrombi and mucosal hemorrhage were found in patients. previously described features of intraluminal schistocytes were not observed in our patients. interestingly, loss of glands, total denudation of mucosa, apoptosis and detachment of endothelial cells were also found in patients with gvhd and no itma, suggesting that these features are not pathognomonic of itma. among itma patients, two patients were classified in the clinical group of acute gvhd/systemic tma, while the other patients had clinical and histopathological features of itma and severe grade iii-iv steroid-refractory acute gvhd ( patients) or extensive chronic gvhd ( patient) but no evidence of systemic tma. in the majority of patients ( / ), itma occurred during the early posttransplant period at . ( . - ) months. clinical features (gastrointestinal bleeding, diarrhea, pain, nausea) presented no differences between patients with or without itma. prognosis was poor for patients with itma who suffered from a significantly higher mortality rate of % compared to the rest patient population (p= . ). with a median follow-up of . ( . - . ) months, year overall survival probability (os) was . for itma, % for gvhd and % for systemic tma. unfavorable predictive factors for os were itma (p= . ), hla mismatched donors (p= . ) and gastro-intestinal bleeding (p= . ). conclusions: intestinal tma has emerged as a novel distinct entity in patients with gvhd and/or systemic tma. distinct histological features may be useful in differential diagnosis of these severe hct complications. mortality rates higher than those of systemic tma highlight the need of proper recognition of itma that needs to be further studied in terms of diagnostic and therapeutic potential. disclosure: e.g. was supported by the european hematology association clinical research grant. the remaining authors declare no competing financial interest. the beneficial effects of thrombomodulin gene polymorphisms after hematopoietic stem cell transplantation background: chronic graft-versus-host disease (cgvhd) remains the major cause of late morbidity and mortality after allogeneic blood and marrow transplantation. treatment options for cgvhd, particularly its sclerotic forms remain limited. active hedgehog (hh) signaling was shown as a therapeutic target in both mouse and human cgvhd, with limited efficacy and significant toxicities described in a published clinical trial (defilipp, ). methods: adult patients with steroid refractory sclerodermatous cgvhd, defined as requiring > . mg/kg/day of prednisone dose equivalent (pde), or need for second-or third-line therapy beyond corticosteroids and calcineurin inhibitors or sirolimus were eligible for this open label study of vismodegib, a first generation hh pathway inhibitor. primary endpoint was failure free survival, defined as absence of non-relapse mortality, no recurrent malignancy, steroid dose at months =< . mg/kg/day of pde, and no addition of new systemic treatment. vismodegib was administered orally for - months, with dose reductions at development of toxicities. peripheral blood mononuclear cells were isolated from samples collected at treatment initiation and every three months thereafter. the immune profile of circulating b cells was analyzed by flow cytometry and t helper polarization by qrt-pcr of sortpurified cd + t cells. results: at the time of interim analysis, patients were evaluated. patients completed months of treatment and five patients completed months of treatment. therapy was discontinued in patients prior to months due to treatment-related (n= ) and unrelated (n= ) side effects. most patients experienced grade toxicities (muscle cramps and dysgeusia), with only a single grade toxicity (weight loss). patients who completed months of therapy demonstrated partial response, and overall, the primary endpoint was reached in % ( / ) of patients. in patients who discontinued vismodegib, cgvhd worsened acutely after discontinuation. correlative analysis of immune cellular subsets in peripheral blood in paired samples (pre-treatment and month of therapy) documented modulation of b cell subsets pathogenic in cgvhd (pregerminal center and plasmablast-like b cells) and diminished t helper polarization in cd t cells. conclusions: overall, use of vismodegib was associated with potential clinical efficacy in sclerodermatous cgvhd with possible mechanistic evidence arising in correlative studies. while side effects were common, further studies of hh inhibition in cgvhd are warranted. future studies should employ adjusted dosing regimens, along with supportive care interventions to offset side effects, and testing of novel hh inhibitors with enhanced safety profiles. clinical background: graft-versus-host disease (gvhd) results from recognition of host antigens by donor t cells following allogeneic hematopoietic stem cell transplantation (sct). we tested the hypothesis that somatic neomutations occurring after sct from donor and/or recipient dna may trigger gvhd. methods: we longitudinally analyzed both constitutive and somatic mutations by whole exome sequencing (wes) in patients who received sct from a sex-matched hlaidentical sibling for npm mutated acute myeloid leukemia (pt# ) and jak v f mutated primary myelofibrosis (pt# ). both patients were initially refractory to alloreactivity, i.e. not displaying any signs of gvhd, even after several donor lymphocyte infusions. acute gut gvhd finally occurred after a further dli preceded by a lymphodepleting chemotherapy. in pt# , gvhd correlated with a graft-versus-tumor effect. wes was performed on dna from recipient saliva and donor pbmcs (germline samples) and from sequential post-sct pbmcs samples on a hiseq illumina with x bp paired-end reads at a mean depth of coverage of - x. germline and somatic mutations were determined using in-house bioinformatic pipelines (named ewok from the curie institute and smaug from the henri mondor hospital), using briefly gatk as variant caller for germline samples, and a combination of variant callers for matched normaltumor pairs. we adjusted parameters to detect somatic mutations at a minimal variant allelic frequency (vaf) of % compared to recipient and donor germline for all variations (minimal coverage = x for germline and x for tumor sample). results: wes allowed detecting somatic driver mutations explaining aml and pmf for both patients in the initial timepoint and all these driver mutations disappeared at the following timepoints. as expected, the somatic variant rate was x higher in pt# with aml than in pt# with pmf at each timepoint, except for the final gvhd timepoint. indeed, at this final point, the somatic variant rate dramatically decreased by % as compared to previous timepoints. by subtracting variants detected pre-and post-sct from those identified at the ultimate time-point of gvhd occurrence, we created sets of and variants respectively for each patient (keeping only variants with at least reads of mutated dna). these variants can be classified in categories: (i) those with only with a slight increase at time of gvhd, i.e. ≤ -fold compared to highest previous vaf (lrrc , or u , or g , alpp, frg , frg b and lilrb genes), and (ii) those with a significant increase at that time, i.e. > -fold compared to highest previous vaf (phf , smpd , ercc and krtap - genes). none of the variants or genes involved was common between the patients. ontology classification of mutated genes showed the implication of some of them in cell death, regulation of map kinase activity, mrna splicing and immune system process, making them good candidates for further studies. identification of variants appearing pre-gvh and turning off at time of gvhd is ongoing to unveil putative neoantigens that could trigger the alloreactive response. conclusions: using a comprehensive, pre-and post-sct, wes of donor/recipient pairs, we identified several neomutations from donor and/or recipient dna correlating with gvh/gvt effect development. disclosure results: a total of patients experienced cgvhd, and mild, moderate, and severe cgvhd were observed in , , and patients, respectively. the -year cumulative incidence of total cgvhd was . % ( % ci, . - . %), and the -year cumulative incidence of moderate to severe and severe cgvhd was . % ( % ci, . - . %) and . % ( % ci, . - . %), respectively. the patients who had loci mismatched had a higher -year cumulative incidence of total cgvhd ( . % vs. . %, p= . ) and moderate to severe cgvhd ( . % vs. . %, p= . ) compared to those of the patients who had - loci mismatched. the patients who had maternal donors had a higher -year cumulative incidence of moderate to severe cgvhd ( . % vs. . %, p= . ) compared to that of the patients who had other donors. the patients who had grade iii to iv acute graft-versus-host (agvhd) had a higher -year cumulative incidence of total cgvhd ( . % vs. . %, p< . ) and moderate to severe cgvhd ( . % vs. . %, p< . ) compared to those of the patients without agvhd. in multivariate analysis, grade iii to iv agvhd was the only independent risk factor for total cgvhd (hr= . , %ci, . - . ; p< . ) and moderate to severe cgvhd (hr= . , %ci, . - . ; p< . ). in the model excluding agvhd, maternal donor was the risk factor for moderate to severe cgvhd (hr= . , %ci, . - . ; p= . ). conclusions: we observe that severe agvhd was the most important risk factors for cgvhd after haplo-hsct, and further interventions should be considered in these patients to prevent severe cgvhd. disclosure: none of the authors have any potential financial conflict of interest related to this manuscript. background: extracorporeal photopheresis (ecp) has been successfully used for the treatment of graft-versus-host disease (gvhd). ecp therapy might restore the balance between effector and regulatory cells which is severely impaired in gvhd. nk cells are the first lymphocyte subset to be reconstituted after allogeneic hematopoietic stem cell transplantation (allo-hsct). as an important innate immune cell population, nk cells can temporally bridge the transient period of t-cell deficiency post allo-hsct, by protection from opportunistic infections and prevention of leukemic relapse by graft-versus-leukemia (gvl) effect. nk cells not only preserve homeostasis through targeted killing of allo-reactive t cells and thereby control gvhd but also enhance inflammation by secretion of tnf-α and ifn-γ and thereby promote gvhd. therefore, we investigated here the role of nk cells in gvhd patients under ecp therapy. methods: thirty four patients with steroid-refractory/ resistant agvhd ≥ ii°and moderate to severe cgvhd received ecp therapy which performed according to the guidelines. glucksberg and nih criteria were used for clinical staging of agvhd and cgvhd under ecp therapy, respectively. the comprehensive phenotypical analysis of nk cells was evaluated by multicolor flow cytometry. nk activity in terms of killing function, cytokine release capacity and proliferation function was monitored by chromium- release assay, intracellular cytokine staining and cfse staining, respectively. results: five different nk cell subsets were defined based on cd and cd expression. cd bri nk cells displayed an immature and activation profile with high expression of cd l and nkg d. agvhd patients had a higher frequency of cd bri nk cells when compared with hds and cgvhd patients, who were characterized by significant increase of the cd dim cd + and cd -cd + nk cell subsets with high expression of differentiation markers cd b and cd . of note, cd bri cd -nk cells could serve as a novel predictive biomarker for the response of agvhd patients to ecp treatment. in responding agvhd patients, an increase of cd bri nk cells was observed already during the early ecp treatment phase, suggesting immune reconstitution. after priming of the progenitors, ecp could differentiate immature cd bri nk cells into mature cd dim nk cells with reduction of cd l on cd bri nk cells. moreover, cd dim nk cells could further be matured through upregulation of cd expression by ecp. notably, ecp therapy could shift the nk cells from a cytotoxic to a regulatory phenotype within the cd bri nk cells. in spite the immunomodulatory effect of ecp on nk cells, nk activity could be kept intact under ecp therapy. the killing activity of nk cells was stable as confirmed by a cr release assay. ecp therapy had no negative effect on the quantity and quality of cytokine release by nk cells upon k stimulation. especially, the polyfunctionality of nk cells was not altered significantly by the ecp therapy. conclusions: nk cells play an important role in gvhd and could serve as a predictive cell population for the clinical response to ecp therapy. in the current study, ecp influenced the differentiation, maturation and education of nk cells ameliorating gvhd without comprising the antiviral immune defense and gvl effect. disclosure: the authors declare no competing financial interests, except the following: therakos mallinckrodt gave a financial support to as and ms for the documentation of the clinical course and for the analysis of immune cells of the patients, pw has honoraria and membership on advisory boards for sanofi-aventis. abstract withdrawn. cyclosporine levels > µg/l on day post-transplant was associated with significantly reduced acute graftversus-host disease following allogeneic hematopoietic stem cell transplantation monica bianchi , dominik heim , claudia lengerke , martina kleber , dimitrios tsakiris , jakob passweg , alexandar tzankov , michael medinger background: acute graft-versus-host disease (agvhd) remains a major complication of allogeneic hematopoietic stem cell transplantation (allo-hsct). affected patients, especially with steroid-refractory agvhd, have a very poor prognosis. prophylaxis with cyclosporine a (csa) is the backbone of gvhd prevention in most conditioning regimens. methods: in a retrospective analysis of patients treated with allo-hsct, we correlated csa levels at the day of transplantation (day ) and day + with the incidence of acute and chronic gvhd. we postulate that higher target csa levels > μg/l will result in a lower incidence rate especially of agvhd after allo-hsct. results: we assessed patients with either aml n= , lymphoma/myeloma n= , mds/mpn n= , all n= , cll n= , cml n= , or bone marrow failure n= . in patients with clinically relevant agvhd grade ≥ , mean csa levels was lower on day and day + ( ± μg/l; and ± μg/l; respectively) compared to patients without agvhd ( ± μg/l; and ± μg/l; respectively; day : p= . ; day + : p= . x - ). in patients with csa level < μg/l, the incidence of agvhd was significantly more frequent compared to patients with csa levels > μg/l [( / ; %) versus / ( %); p= . x - ]. in patients with cgvhd, there was no significant difference between csa levels < μg/l ( / ) compared to csa levels > μg/l ( / ; p= . ). the optimal csa cut-off level for the prevention (i.e. roughly % incidence reduction) of agvhd was > μg/l at day and > μg/l at day + ( figure ) in a competing risk analysis, time to agvhd grade ≥ (using death of other causes as competing risk) was associated with csa levels > μg/l on day and on day , unrelated donors, myeloablative conditioning (mac), and for the diagnosis lymphoma/myeloma. conclusions: our data support close monitoring with active adjustments of csa dosing to maintain therapeutic csa levels above μg/l in the first days of allo-hcst to reduce agvhd. disclosure: noting to declare. liposomal cyclosporine a for inhalation (l-csa-i) to treat bronchiolitis obliterans syndrome: novel formulation with therapeutic potential for patients with bos following allo-hsct noreen roth henig , emilie hofstetter , dominik kappeler , gerhard boerner background: bronchiolitis obliterans syndrome (bos) is a rapidly progressive lung disease caused by t-cell mediated inflammation that leads to blockage of bronchioles, leading to respiratory failure and death shortly after diagnosis. approximately % to % of patients who undergo allogeneic hematopoietic stem cell transplant (allo-hsct) will develop bos, with - % developing bos as a respiratory form chronic graft-vs-host disease (cgvhd) in addition to other signs of cgvhd. mean time to bos diagnosis ranges from to days post-transplant. the histopathology of bos after allo-hsct and lung transplantation is identical. early studies of l-csa-i for the prevention of bos in lung transplant recipients demonstrated therapeutic benefit. l-csa-i is a novel, liposomal formulation of cyclosporine administered via a pari investigational eflow â nebulizer which delivers a potent immunosuppressant to the site of disease. pharmacokinetics and tolerability of l-csa-i is presented. methods: retrospective review of two clinical studies of l-csa-i (isotonic, mg/ml) for bos associated with lung transplantation. both studies had a control arm and results reported here are for patients who received l-csa-i. subjects received mg (single lung transplant) and mg (double lung transplant) bid via inhalation. blood samples for pharmacokinetic analysis of cyclosporine a concentrations were collected before inhalation, immediately after inhalation, and thereafter in intervals of , , min and , , and hours. local and general tolerability of l-csa-i was investigated. results: between the two studies, subjects received either or mg bid of l-csa-i. pharmacokinetic models predict a constant drug level in the lung. maximum serum cyclosporine a concentration after inhalation was . ± . ng/ml. trough levels for up to -years of daily administration was - ng/ml with no evidence of accumulation following repeated exposure. tolerability data was assessed from patient-month exposure to l-csa-i. reported symptoms were: pharyngeal soreness %; cough %; dyspnoea %; and wheezing %. no subject discontinued due to intolerability. inhalation time is on average - min. conclusions: l-csa-i provides high and constant concentrations to the airways of the lungs and the site of bos. l-csa-i is well tolerated in lung transplant patients. use of l-csa-i instead of augmentation of systemic csa reduces the total drug exposure. a multicentre phase safety and exploratory efficacy trial for the treatment of bos in allo-hsct recipients is underway. disclosure background: there are a number of biomarkers that predict non-relapse mortality (nrm), graft-versus-host disease (gvhd) and relapse incidence (ri) after conventional gvhd prophylaxis based on calcineurin inhibitors with or without antithymocyte globulin. currently there is limited data whether the conventional predictive biomarkers work with posttransplantation cyclophosphamide (ptcy) prophylaxis. methods: prospective single-center study in - enrolled adult patients with acute leukemia in cr ( % with all, % with aml). received matched related bone marrow (bm) graft with single-agent ptcy and received unrelated peripheral blood stem cell graft (pbsc) with ptcy, tacrolimus and mmf. the grafts were studied by flow cytometry (facs aria ii, antibodies by miltenyi biotec). the following populations were analyzed: cd , cd , cd , cd cd , nkt, inkt, treg, double-positive t-cells, double-negative t-cells, tcralpha/beta, tcr v memory cells. the crypreserved plasma from were analysed by elisa (commercial kits by ebioscience and critical diagnostics) for vegf a soluble tnf receptor (stnfr), il- , il- , soluble il- receptor, st , il- and stnfr. the above mentioned biomarkers were tested in logistic regression with roc analysis, assays with auc> . were selected for analysis in fyne-gray regression with competing risks. cut off levels were determined for significant parameters. results: median follow-up was months (range - ). in the whole group overall survival (os) was %, eventfree survival (efs) %, grade ii-iv acute gvhd %, moderate and severe (m&s) chronic gvhd %, nrm %, mortality in patients with gvhd %, ri %. there was no difference between bm/related and pbsc/unrelated grafts in the incidence of gvhd, nrm and ri (p> . ). the only significant predictor of acute gvhd were low levels of il- level on day+ (p= . , % vs % with the cut off pg/ml). m&s chronic gvhd was predicted only by the high percentage of inkt cells in the graft (p= . , % vs % with the cut off . %). there was a correlation between il- levels and number of nk cells in the graft (p= . ). nrm was related to infectious complications, nonetheless high levels of vegf a on day (p= . , % vs % with the cut off ng/ml), st on day+ (p= . , % vs % % with the cut off ng/ml) and low percentage of cd +cd -cells in the graft (p= . , % vs % with the cut off . %). the identified biomarkers of nrm had no association with the pre-transplant crp and ferritin levels (p> . ). the only significant parameter for ri was the level of cd cells in the graft (p= . ). none of the identified biomarkers significantly predicted overall survival (p> . ). conclusions: in the related and unrelated grafts with ptcy the study of biomarkers has low clinical utility due to very low gvhd-related mortality. however st and vegf a can predict infection-related mortality. also the study verified previous observations that high level of il- is associated with reduced gvhd incidence after ptcy and identified the importance of nk and inkt cells in the induction of tolerance with ptcy. references background: hematopoietic cell transplantation (hct) is the only curative approach for many hematological malignancies but life-threatening toxicities, such as graft-versushost disease (gvhd) and infections, still limit its fullpotential impact on the disease. strategies for keeping allohsct more effective and safe are needed in order to reduce morbidity while improving its immunological effect to control disease relapse. post-transplant cyclophosphamide (ptcy) has been demonstrated to improve acute gvhd (agvhd) and chronic gvhd (cgvhd) control in allogeneic bone-marrow hct from identical and haploidentical donor. the use of ptcy, after peripheral blood stem cell transplantation (allopbsct) from hla-matched unrelated/related donors, has been investigated by our group in a clinical trial (nct ) and preliminary results were published last year. here we report updated efficacy and safety data about the expanded cohort of patients treated with ptcy followed by tacrolimus and mycophenolate mofetil (t/mmf). methods: we analysed data about consecutive patients with high-risk hematologic malignancies received allopbsct from hla-matched unrelated/related donors between march and august . gvhd prophylaxis was ptcy mg/kg (days + + ), tacrolimus from day + and mmf from day + to day + . primary objectives were cumulative incidence of agvhd and cgvhd. secondary objectives were event-free survival (efs), cgvhd-efs, overall survival (os) and non-relapse mortality (nrm). results: patients median age at transplant was (range - ) years. ( %) patients were transplanted in first complete response (cr), ( %) patients in second/third cr, the others in disease control. a median dose of . (range - ) x ^ cd /kg was infused. primary graft failure was observed in one patient. all patients were off mmf on day + , the median day of tacrolimus discontinuation was (range - ). eight out of ( %) patients developed agvhd, ( %) of them were grade ii-iii; median day of onset was day (range . no grade iv was observed. no cases of late-onset agvhd were reported. cumulative incidence of cgvhd was % ( / ), median day of onset was (range - ). systemic treatments were required, but all patients were able to discontinue immunosuppression (is). with a median follow-up of (range - ) months, efs was %, cgvhd-efs was % and os was %. non-relapse mortality (nrm) was % ( / ): patients died because of multidrug resistant bacteria septicemia. nowadays patients are alive with no evidence of disease, being continuously off is and completely reintegrated in their normal daily life activities. conclusions: the updated reported results confirm, in a larger cohort of patients with a longer follow-up, that ptcy after pbsc-hct is highly active in agvhd and cgvhd prevention with extremely limited nrm. this strategy, not only allowed earlier discontinuation of immunosuppression, but also reduced the overall time of exposure to is for most of the patients. all these features might contribute, in the future, to transform hct into a safe immunologic platform that may be combined with advanced form of cellular therapies (car-tcells), aiming to increase safely the graftversus-tumor effect. clinical methods: pediatric patients ( - years) with nmd undergoing unrelated hct were eligible for this single center, phase i trial. following reduced intensity conditioning, abatacept ( mg/kg iv on days - , + , + , + ) was added to standard gvhd prophylaxis (cyclosporine, mycophenolate mofetil [mmf]). patients were followed for years for standard hct outcomes. [[p image] . figure results: since june , patients have been enrolled and transplanted (table , excluding # ). donor source was bone marrow in all. with median follow-up of . years, of patients survive without disease. initial engraftment was successful in , at a median of and days, for neutrophils and platelets respectively. one patient ( ) had secondary graft rejection in the setting of viral reactivation (cmv/ebv), with successful engraftment following a nd unrelated hct. in engrafted patients, myeloid (cd ) chimerism was % at all timepoints; t-lymphoid (cd ) chimerism was mixed but reached >/= % (figure ). one patient ( ) with saa had primary graft rejection in the setting of inadequate tnc dose ( . x /kg) and died from marrow aplasia/infection despite nd hct. a second death from wilms' tumor occurred months post successful hct, in a patient ( ) with dba and constitutional chromosome abnormality. except patient , all patients received doses of abatacept, which was well tolerated, with all severe adverse events expected for a hct population. cmv and ebv reactivation occurred in patients each, with resolution using standard anti-viral therapy. one patient ( ) was diagnosed with ebv-driven post-transplant lymphoproliferative disease, which responded to rituximab and immune suppression withdrawal. no patients developed severe acute (grade iii-iv) or chronic gvhd (table ) , and no patients required systemic immune suppression at > year. conclusions: these preliminary data suggest that abatacept can be safely added to cyclosporine and mmf gvhd prophylaxis in pediatric patients with bone marrow failure undergoing unrelated donor hct, with encouraging rates of gvhd despite half of patients having a mismatched ( / ) donor. given the higher risk of graft rejection in this non-malignant cohort, rejection (in addition to gvhd) will be a primary focus in our subsequent multi-center, phase trial. clinical trial registry: clinicaltrials. gvhd and may to have be separated from from toxicity to infectious complications in the early phase after allohsct. methods: from our files we identified patients which had upper gastrointestinal tract endoscopy after allohsct in with biopsies were taken from the esophagus, stomach and duodenum simultaneously. of these patients were excluded because of infection, reflux disease or drug toxity and the remaining patients were included in our study. we evaluated the routine stained esophageal biopsies, applied a grading scheme and compared the histological findings with those within the stomach and duodenum, the endoscopic findings and the clinical course. results: in of biopsy samples of the esophagus, we identified histological features of acute gvhd, ranging from vacuolar degeneration (grade ) and single-cell apoptosis (grade ) to the formation of clefts (grade ) and mucosa denudation in advanced cases (grade ), resembling epithelial lesions in acute gvhd of the skin. these findings correlated with gvhd involving the stomach and duodenum and the clinical manifestations of gvhd in other organs. endoscopically patients with gvhd revealed signs of inflammation, ranging from erythema to ulceration in the more advanced cases, sometimes reminiscent of reflux or infection. clinically these patients had abdominal discomfort ranging from inappetence to nausea, accompanied by emesis or diarrhea and weight loss. conclusions: we have shown that acute esophageal gvhd occurs after allohsct and is correlated with acute gvhd in stomach and duodenum. it could be diagnosed and graded histologically. the endoscopic findings are signs of inflammation. our results may help to establish the histological diagnosis of acute gvhd using endoscopic biopsies from the esophagus and to explain the alterations observed in the esophageal mucosa in patients after allohsct. [ background: intestinal acute graft versus host disease (agvhd) is a major thread after allogenic hematological stem cell transplantation (allohsct), with a high mortality in patients which were refractory to steroid treatment in particular. recent papers point to a correlation of histological grading of intestinal gvhd and prognosis in patient after allohsct. however a comparison with clinical scores has not been performed so far. methods: in this analysis, retrospective data from patients who underwent endoscopy due to clinical signs of agvhd (day + to + after allohsct) were evaluated. of each patient least biopsies from different sites of the colon which were taken simultaneously. of each biopsy series the maximum histological grad of agvhd according to the lerner scheme was obtained and compared with the glucksberg stage of the lower gastro intestinal tract (gslgi) and the overall glucksberg grade (ogg). these three grades were compared for non-relaps related mortality using the log-rank test and for sensitivity to steroid treatment applying the receiver operating characteristic for the patients who received steroid treatment. for these patients the non-relaps related mortality for responder and non-responder were calculated using also log-rank test. results: the histological grade strongly correlated with the survival (p= . ). a statistical significant correlation was also found for the gslgi (p= . ), whereas the ogg revealed no significant correlation (p= . ). non-relaps related mortality was mainly related to infection or sepsis (in / patients who died). -eighty-one of the patients received steroid therapy. the sensitivity to the steroid therapy correlated with each of the three scores (p< . ) but was the strongest for gslgi (area under the curve (auc) . ), compared to ogg (auc . ) and the histological score (auc . ). the survival of the patients, which were sensitive to steroid treatment was significantly better than those of steroid refractory patients (p= . ). conclusions: we found that histological and clinical grading in patients after allohsct with intestinal gvhd was correlated with survival and respond to steroid treatment. histological scoring may predict survival more precisely than ogg and gslgi but did not add substantial information to the prediction of treatment response. [ emerging evidences suggest that regulatory b cells (bregs) play essential roles in inflammation, autoimmune diseases and tumors. few data exist about the role of bregs in the contest of hematopoietic allogeneic stem cell transplantation (hsct). some authors have observed that bregs from patients with chronic graft-versus-host disease (cgvhd) were less frequent and less likely to produce il- than bregs as compared to healthy donors or patients without cgvhd. these findings suggest that bregs may be involved in cgvhd pathogenesis. the purpose of our study was to evaluate a possible role of b cell subsets on gvhd occurrence. methods: lymphocyte subset enumeration was performed by aquios cl flow cytometer (beckman coulter), a quantitative automated analyzer that performs two diagnostic panels: tetra- cd -fitc/cd -rd / cd -ecd/cd -pc and tetra- cd -fitc/cd +cd -rd /cd -ecd/cd -pc . b cell subsets (memory, mature and transitional b cells) on peripheral blood samples were analyzed by aquios designer software, a tool for the creation of user-defined applications. panel- cd -fitc/cd -pe/cd -ecd/cd -pc /cd -pc and panel- cd -fitc/cd -pe/cd -ecd/cd -pc / cd -pc were specifically designed by beckman coulter for our center. the flow cytometric analysis was performed as follows: in donors and patients at basal level; on graft products and in patients at days + , + , + , + after hsct. statistical significance was assessed with prism software (graphpad) by mann withney test. p < . was considered statistically significant. results: actually we enrolled patients submitted to hsct in our center from november . a preliminary statistical analysis was performed on patients. stem cells source was peripheral blood (pb) in cases and bone marrow (bm) in the others . the conditioning regimen was myeloablative in patients and ric in patients. agvhd was diagnosed in patients ( %). no associations were found between b cell subsets in donors and patients at baseline and the occurrence of agvhd. however we found a higher median percentage of transitional b cells in graft products in patients without agvhd ( . %, . - . ) compared to patients with agvhd ( . %, . - . ) (p= . , fig a) . in addition, patients without agvhd showed a lower median percentage of memory b cells ( . %, range . - ) in graft product as compared to patients with agvhd ( . %, range . - . ) (p = . , fig. b ). finally in the subgroup of patients receiving pb as stem cell source we observed a higher percentage of cd + lymphocytes in graft product in patients with agvhd ( %; range - ) compared to patients without agvhd ( %; range - ) (p= . ). in the monitoring of b cells reconstitution we observed that cd + events did not appear before day + after hsct and these were b transitional immature events predominantly. conclusions: our data suggest a possible protective link between transitional b cells and agvhd development. these results data need to be confirmed in a larger cohort of patients. moreover, it will be interesting to evaluate the relationship between transitional b cells at day + and the occurrence of cgvhd. clinical background: agvhd is a major complication of allogeneic hematopoietic stem cell transplant (hsct) and a risk factor for post-hsct mortality. the objective of this analysis is to describe patients with agvhd who had a suboptimal response to corticosteroids. methods: patients who developed ibmtr severity index ii-iv agvhd after first hsct between / / to / / were included in an ongoing chart review at centers in the united states. patients who had ever participated in a gvhd prophylaxis trial or used jak inhibitors were excluded from the study. suboptimal response to corticosteroids was defined as use of additional systemic anti-gvhd therapy, inability to taper high-dose steroids (≥ mg/ kg) by ≥ %, or tapered corticosteroids by ≥ % but not to < mg/day. results: the analysis included patients with suboptimal response to corticosteroids. mean age was years; % were male. median time from transplant to agvhd diagnosis was days. at the time of maximum agvhd grade, % of patients were grade ii and % were grade iii-iv; % had lower gi involvement, and % had ≥ organs involved. from time of diagnosis to maximum agvhd grade, % of patients had new organ involvement or an increase in agvhd grade. median time from diagnosis to maximum grade was . days, and was . days for patients with lower gi involvement. systemic corticosteroids were initiated on the day of diagnosis for % of patients. average starting daily dose was mg ( . mg/kg) for prednisone and mg ( . mg/kg) for methylprednisolone. steroid dose was increased for % of patients during follow-up; % were unable to taper below mg/day. among patients who received additional systemic anti-gvhd therapy (n= ), % increased their corticosteroid dose before initiation of additional anti-gvhd therapy. median time from initiation of corticosteroids to additional therapy was . days. frequently used therapies were mycophenyalate mofetil ( %), atg ( %), extracorporeal photophoresis ( %), tocilizumab ( %), etanercept ( %), and sirolimus ( %). agvhd recurred in % of patients and was managed by increasing corticosteroid dose in % of patients. % had any infection within first days post-hsct. forty patients ( %) required hospital readmission(s); % had ≥ readmissions within days post-hsct, with a mean inpatient lengthof-stay of days. relapse of underlying malignancy was reported for ( %) patients. two-thirds ( %, n= ) patients died at a median of (interquartile range (iqr): - ) days from agvhd diagnosis; a higher proportion ( %) of patients with maximum grade iii-iv agvhd died at a median of (iqr: . - . ) days; majority ( %) of patients with lower gi agvhd died at a median of . (iqr: - ) days. conclusions: a majority of patients with agvhd who had suboptimal response to systemic corticosteroids had severe and rapidly progressing disease and resulted in a high mortality rate ( %); progression was more rapid and mortality increased for patients with lower gi involvement. most patients required readmission to the hospital with extended length-of-stay. an urgent need exists for effective and tolerable therapies that quickly resolve life-threatening agvhd in early stages of disease. disclosure results: median time to onset of bo from allohct was . months (range . - . ). previous acute gvhd in . % (n = ) [grades iii-iv . % (n = )]. in . % (n = ) cgvhd had exclusive lung involvement, while the other patients ( %) had other organs affected. at diagnosis of bo, . % (n = ) were under immunosuppressive treatment. . % (n= ) of patients with bo received ecp as second-line treatment. median duration of treatment was months ( . - . ) and time to response . months ( . - . ). median of sessions was ( - ). evaluation of response was based on the evolution of fev measurement: . % (n = ) complete response; % (n = ) partial response and % (n = ) stable disease. one patient did not get any response and another was not evaluable. . % of patients (n = ) could reduce immunosuppression, and in one case it was completely discontinued. there is a trend for early separation between survival curves in favor of ecp ( figure ). one patient had sepsis secondary to central venous catheter infection as complication related to ecp. conclusions: ecp has emerged as a promising treatment for bo after allohct. in our experience, ecp was effective to stabilize or improve the disease in many patients and allowed to taper esteroids with minimal associated complications. however, prospective studies and longer follow-up are needed to support these findings. disclosure: nothing to declare background: the key role of il- signaling in acute graft vs. host disease (agvhd) and cytokine release syndrome (crs) has evoked growing use of tocilizumab, an anti-il receptor (il -r) antibody, in these settings. apart from regulation of t-and b-cell differentiation, immune cells migration to inflammatory sites and t-cell recruitment, il- complex with il -r through gp upregulates production of fibrinogen (fg) and other acute phase proteins, including c-reactive protein (crp). methods: we retrospectively analyzed data of patients treated with tocilizumab ( mg/kg) due to steroid-refractory (sr) agvhd and patients because of crs. median age was and years, respectively. seven patients were transplanted from unrelated donors (mud/mmud) and from sibling donors. eight patients received myeloablative and reduced intensity conditioning regimen. analyzed data included concentrations of fg, crp, an incidence of infections at tocilizumab administration and in weeks following the infusion. results: stage ii agvhd was diagnosed in patient, stage iii in , and stage iv in patients. involvement of the gastrointestinal tract (gi) was observed in % of cases. the median fg concentration before tocilizumab administration was . g/l (range, . - ) and crp mg/dl (range, - ) and % of patients had an active infection. after infusion of the antibody, we observed a decline of fg and crp levels. the median level of fg was . g/l (range, . - . ) - days after the tocilizumab infusion with no severe bleeding complications. a median crp value was . mg/dl (range, . - ) despite confirmed infectious complications. three weeks after infusion of tocilizumab fg raised to the normal range in % of patients (fig ) . five patients with sr agvhd achieved a complete response, and had a partial response after tocilizumab therapy. [[p image] . fibrinogen levels in gvhd patients following tocilizumab infusion.] a group treated with tocilizumab due to crs had higher initial levels of fg . g/l (range, . - . ) and crp mg/ dl (range, - ) before administration of the drug. reduced fg and crp levels from a baseline value were also observed in this group. however, concentrations were higher than in gvhd patients: fg . g/l (range . - . ) and crp . mg/dl (range . - ). in all patients, a differential diagnosis of disseminated intravascular coagulation was excluded. conclusions: . fibrinogen declines after tocilizumab therapy due to its cytokine-regulated production in the liver. coagulation monitoring should be performed during the first weeks after administration of the antibody to avoid serious bleeding complications. . crp concentrations remain low despite the presence of active infections following infusion of tocilizumab. crp fails as a marker of infection during weeks following the therapy. . tocilizumab is an effective therapy in patients with agvhd, especially with the gi involvement. disclosure: nothing to declare vanishing bile ducts after allogenic hsct: is it really gvhd? antonio grasso , lorenzo d'antiga , aurelio sonzogni , massimo gregori , alessandra maestro , roberto simeone , natalia maximova background: evaluation of liver gvhd was historically based by elevation of bilirubin levels and by reduction and degeneration of small bile ducts on histological samples of post-transplant liver biopsy. however, there is a lack of studies that compared histological finding of ductopenia between post-autologous hsct and post-allogenic hsct. studying severity of ductopenia following allogenic hsct, we aimed to demonstrate lack of correlation between ductopenia and clinical signs of liver gvhd. methods: we retrospectively collected a series of allogeneic hsct performed from to in the institute burlo garofolo. all patients undergo percutaneous liver biopsy in most cases at three months, one year and three or more years after hsct. indications for biopsy were alteration noted at weekly follow-up assessments of at least one clinical or laboratory marker of liver impairment or cholestasis. ductopenia was defined by number of portal tracts with no interlobular bile duct divided by the total number (severe if the ratio was less than . ). clinical gvhd was defined by nih consensus criteria results: our population involved % males and % females with oncological ( %) and non-oncological underlying disease ( %). clinical signs of liver gvhd were present in % of the patients (n= ), % with contextual intestinal involvement, % with cutaneous and intestinal involvement. patients underwent biopsy at a mean time of +/- days after hsct, patients underwent a biopsy at months after hsct and patients after three or more years from hsct. results of biopsies are showed in table . no difference in incidence of ductopenia were found between liver gvhd group and no gvhd. table] . table : incidence of ductopenia - months, months and or more years after hsct in total population, gvhd group and no-gvhd group] the group that not received chemotherapy prior the hsct had an overall incidence of ductopenia of % (severe ductopenia of %) statistically significative in comparison with the oncological underlying disease group ( % of ductopenia and % of severe ductopenia). furthermore, a little sub-group of patients extrapolated from our population received liver biopsy before hsct for diagnostic assessments: of the with an oncological underlying disease % already showed ductopenia, while no signs of ductopenia were found in the others with a nononcological disease. conclusions: there is no correlation between incidence of gvhd and histologically finding of ductopenia on liver biopsy. ductopenia may be caused in the first place by chemotherapy treatment received before hsct and myeloablative conditioning for hsct and it's not related with gvhd. this hypothesis is strengthened by the subgroup analysis of pre-hsct biopsy. background: second and third line therapies for steroid refractory acute graft versus host disease (agvhd) after allogeneic stem cell transplantation (asct) are still lacking. ruxolitinib, a selective januskinase / inhibitor could show high efficacy in agvhd, as well as extracorporeal photopheresis (ecp). here we report a single center experience of combining both therapeutic approaches in severe steroid refractory agvhd with additional analysis of immune status of these patients to elucidate direct effects of this treatment on immune response. methods: from june to february , patients ( . % male, . % female, median age: . years, r: - ) with steroid refractory agvhd of lower gi-tract after asct were treated with ruxolitinib and extracorporeal photopheresis as third, fourth or fifth line therapy. some patients showed additional agvhd of skin (n= ), liver (n= ) or upper gi-tract (n= ). all patients had an overall grade iii ( %) or iv agvhd ( %). steroid refractoriness was defined as no improvement in days or aggravation after days of steroid treatment.median start of ruxolitinib or ecp was day . after asct (r: - ). medianduration of ruxolitinib therapy was . days (r: - ) with a median start dosage of mg per day ( x mg; r: - mg). all patients started with ecp treatments per week with an individual reduction of treatment frequency. median number of ecp treatments was . (r: - ) with a median frequency of ecp therapy once a week (r: . - . ). cytomegalovirus (cmv) status of all patients and immune status of ten patients (lymphocyte count with cd + t helper lymphocyte and regulatory t cell count) were collected previously, after four weeks of starting combined treatment and four weeks after stopping the treatment. results: one-year estimated overall survival (os) of all patients was % with a median estimated os of days. patients died because of relapse of underlying disease, one of severe therapy refractory agvhd of lower gi tract and due to infection complications in agvhd refractory setting. overall response was . % (complete remission rate: . %, partial remission rate: . %). . % (n= ) of the patients had cytopenia ctc i-iii during the treatment, no grade iv cytopenia was reported. cmv reactivation during ruxolitinib occured in . % of cases (n= ). tapering of steroids could be performed rapidly with a medium reduction time of . days for reducing to half of the dosage.remarkably, regulatory t cells significantly increased during combined ruxolitinib/ecp treatment compared to regulatory t cell count before treatment (p= . ) and after stopping treatment, regulatory t cell count decreased again (p= . , see figure) . significant changes in whole lymphocyte count or in cd + t helper cell count were not observed. conclusions: treatment of severe steroid refractory agvhd with ruxolitinib plus ecp could show a high complete remission rate of . % with an one year os of %. detecting increased regulatory t cell count during the treatment underlines its direct effects on immune response and encourages to pursue this promising therapeutic approach. [ background: due to increased immunosuppression infections remain the main cause of death followed by higher risk of relapse in patients treated for acute graft versus host disease (agvhd) after allogeneic stem cell transplant (sct). here we report a single-centre experience with extracorporeal photopheresis (ecp) for acute gvhd that was introduced in order to reduce steroid treatment. comparison of overall survival (os) for patients on ecp and patients that received standard first line therapy for agvhd was performed. methods: we retrospectively analysed patients ( %) with acute gvhd grade ii-iv treated from january to october out of total allogeneic sct in that period. all patients received calcineurin inhibitors or sirolimus while receiving steroid treatment for agvhd. twenty-five patients ( %) received ecp with steroid lowering intent. we defined response as ( ) reduction of steroid dose for at least % from baseline while not adding another immunosuppresive agent and ( ) not repeating second steroid treatment if the ecp was started after lowering of steroids to prevent agvhd flare. we checked separately patient responsive and refractory/dependent to steroids. on average patients received ecp procedure once weekly. results: tapering of immunosuppressive therapy as defined was successful in ( %) out of patients in ecp group. in a group of patients without ecp ( %) patients had steroid refractory or steroid dependent agvhd compared to ( %) patients in ecp group. four ( %) patients with steroid refractory or dependent agvhd showed improvement in ecp group compared to only one ( , %) in non ecp group. twenty ( %) patients died due to infectious complication and ( %) due to relapse in non ecp cohort. in ecp cohort ( %) patients died due to infection and ( %) due to relapse. median os was months in non ecp group (r., - ) compared to months (r., - ) in ecp group and os of % at years in non ecp compared to % in ecp cohort was observed. patients with agvhd treated with ecp and faster steroid tapering had longer os compared to patients without ecp (p= , ). conclusions: ecp enables successful tapering or withdrawal of steroid therapy in many patients, even in those who are steroid refractory or steroid dependent. reduction of immunosuppression leads to reduced incidence of infection and relapse which translates into a better overall survival. background: the curative potential of allogenic stem cell transplantation is hampered by graft-versus-host disease (gvhd). pre-clinical study showed an efficacity of jak / inhibitor, ruxolitinib, in treatment of steroidrefractory gvhd. methods: we reported in this monocentric retrospective study, ruxolitinib response and follow up of cases of chronic gvhd (cgvhd) not improved with standard immunosuppressive therapy. complete organ response (cr) was defined as the resolution of clinical manifestations of cgvhd in a specific organ. very good response partial (vgpr) was defined as an improvement of clinical manifestations of cgvhd with more than % decrease of corticosteroid, while a partial response (pr) was associated with less than % decrease of corticosteroid. treatment failure was defined by the absence of improvement of cgvhd, deterioration of cgvhd in any organ by at least one stage, the development of cgvhd manifestations in a previously unaffected organ, and the use of any additional agents to control the disease. results: median age at transplant was years (range, - ). % of patients presented an acute myeloid leukemia. donor type was sibling (n= ), unrelated (n= ) or haploidentical (n= ).two patients benefited a cord blood transplant. patients received either myeloablative ( %) or reduced intensity ( %) conditioning regimens. stem cell source was peripheral blood for % of patients. patients presented mild (n= ), moderate (n= ) or severe (n= ) cgvhd according to nih score. median number of regimens prior to ruxolitinib was (range, - ), among those corticosteroids (n= ). median follow-up after ruxolitinib was months (range, - ). overall responses rate (orr) at month was % with % cr, % vgpr and % pr ( figure ). % of patients failed at month after introduction of ruxolitinib. the rate of cr increased with time : % at months (n= ), % at months (n= ) and % at months (n= ). but vgpr rate was rather stable at months ( %), at months ( %) and at months ( %) vs % at month. among the patients under steroids, ( %) patients discontinued steroids. the -months overall survival (os) and diseasefree survival (dfs) after ruxolitinib was % ( %- %, % ci) and % ( %- %, % ci), respectively. severe cytopenia (grade and ) was observed in patients. after introduction of ruxolitinib, patients presented bacterial infections, patients presented an invasive pulmonary aspergillosis and patient developped a pneumocystis. cytomegalovirus reactivation requiring preemptive treatment was observed in patients. no toxicities required withdrawal of ruxolitinib. [[p image] . figure and partial response to mesenchymal stem cells (msc), as second-line therapy, varies from % to % in acute gvhd patients. we report our experience using mscs to treat refractory agvhd. methods: the study was a retrospective single center study. all data were collected from patients' files. twenty patients were enrolled (age ranging from months to years) between april and april . results: five of these patients received reduced intensity conditioning and patients received myeloablative regimens before hsct. one haploidentical, autologous, cord blood, mud, msd transplantations were performed. the patients were eligible if they developed grades ii-iv agvhd. all patients were treated with standard first-line treatment with corticosteroids and at least one second-line therapy. the definition of steroid resistant agvhd considered as either no response to steroid treatment lasting at least days or progression during treatment of at least one grade within the first hours. prophylactic treatment with calcineurin inhibitors continued at therapeutic dose level. totally, doses of mscs were infused. the median dose of msc was . × cells per kg body weight. the median duration between the diagnosis of agvhd and initiation of mscs therapy was days (range: - ). the received msc doses ranged from one to seven. none of our patients had severe side-effects during infusions of mscs. overall, complete response (ocr) was obtained in patients, partial response in patients and no response (nr) was documented in patients. in our study group, the complete response rates in liver, gastrointestinal, skin agvhd were %, %, % respectively. four patients ( %) died in days after using mscs from complications of agvhd. eleven of patients ( %) were still alive with a median follow-up of days (range: - days) after first mscs infusion. one year estimated probability of overall survival for patients achieving ocr and partial remission/no remission in th day of mscs were . % and %, respectively. conclusions: in conclusion, mscs appears to be a safe and effective treatment option for pediatric patients with steroid refractory agvhd. disclosure: nothing to declare effect of extracorporeal photopheresis on production of serum elafin in chronic graft versus host disease arun alfred , charlotte burton , kathryn goddard , nichloas matthews background: extracorporeal photopheresis (ecp) is a second line therapy for steroid refractory, dependent or intolerant chronic gvhd (cgvhd). in order to guide ecp there is an unmet need for predictive and diagnostic biomarkers. elafin is a serine-protease inhibitor primarily produced by epithelial cells, particularly keratinocytes in inflammatory skin diseases and plasma and epidermal elafin have been identified as biomarkers of skin gvhd ( , ) . since skin cgvhd is noted for a particularly high response rate to ecp, we conducted a study to investigate whether ecp affects the production of elafin. methods: serum samples were collected from cgvhd patients ( male / female; age range: - ) and age-matched healthy controls ( male / female) before ecp and at month intervals up to year. patients had gvhd affecting skin ( / ), mucosal membranes ( / ), liver ( / ), joints ( / ), gut ( / ), eye ( / ), genital ( / ), and respiratory involvement ( / ). serum elafin was assessed by elisa (r&d systems). data were analysed using graphpad prism . statistical tests performed include -tailed mann-whitney, pearson's correlation test, and -way anova with repeat measures, as appropriate. results: chronic gvhd patients presenting for ecp had significantly elevated serum levels of elafin (p= . ; median of ng/ml, iqr - ng/ml) compared to healthy controls (median of ng/ml, iqr . - ng/ml).while % of patients had skin involvement, only % had elafin levels above the iqr of healthy controls. where disease scores were available (n= ) there were no significant correlations with modified rodnans (r= . ) or nih bsa scores (r= . ).sub-analysis was performed by grouping cgvhd patients according to quartiles of serum elafin at pre-ecp baseline. retrospective analysis of patients after months of ecp (n= ) revealed that those with serum elafin levels in the upper quartile (elafin hi ) pre-ecp (min-max: - ng/ml), showed a significant reduction after months of therapy (p< . ; mean +/-sd : ng/ml +/- ng/ml vs ng/ml +/- ng/ml, respectively), which was sustained up to months of ecp (p< . ; mean +/-sd: ng/ ml +/- ng/ml). in contrast, patients with elafin levels below the upper quartile (elafin lo ) showed no significant change (mean +/-sd: ng/ml +/- ng/ml vs ng/ml +/- ng/ml, respectively). of note, pre-ecp patients with elafin below the median received significantly more corticosteroid (cs) than those above, (p< . ; mean +-sd: +/- mg/d vs +/- mg/d, respectively), which was significantly reduced after months of ecp (p< . ; to +/- mg/d), while cs dose was not significantly changed in elafin hi patients until months (p< . ; mean +/-sd: mg/d +/- mg/d vs mg/d +/- . mg/d, respectively). conclusions: consistent with recent data, we found that serum elafin is significantly elevated in a subset of cgvhd patients compared to healthy controls, but did not correlate with skin disease scores. ecp administration was associated with a reduction in serum elafin in the elafin hi subset. further, elafin lo and elafin hi patients tolerated different rates of ecp-mediated tapering of cs immunosuppression suggests pre-ecp elafin measurements may have predictive value. references : background: allogenic hematopoietic stem cell transplantation (hsct) is a potential curative treatment for many malignant and no malignant hematologic diseases, primary immunodeficiencies and some metabolic and deposit diseases in children. graft versus host disease (gvhd) is a major cause of morbidity and a leading cause of non-relapse mortality. corticosteroids are the standard first-line systemic treatment for both acute and chronic gvhd, whereas no second line option for corticosteroid-refractory patients is standardised. ruxolitinib is a potent inhibitor of jak / showing significant responses in refractory gvhd patients in recent reports. methods: we present two centres experience with ruxolitinib for gvhd treatment in pediatric patients. the study was conducted in two spanish pediatric hsct centres, hospital vall d'hebron (barcelona) and hospital universitario la paz (madrid). all patients receiving ruxolitinib since the drug was available were included for retrospective analysis. results: between march and december pediatric patients with acute or chronic gvhd with refractoriness to corticosteroids were treated with ruxolitinib, in different episodes (one patient received it in different moments, and one patient received it in ). patient's sex at birth was female in and male in cases. median age at hsct was , years ( , - , ). primary disease was malignant in patients and non malignant in . median time of gvhd diagnosis was , days ( - ). all gvhd episodes were treated with corticosteroids as first line, with maximum doses between - mg/kg/day (the main dose used was mg/kg/day, / episodes). patients received a median number of , ( - ) previous lines of treatment including steroids before starting ruxolinib; they were extracorporeal photopheresis ( / episodes), sirolimus ( / ), mesenchymal cells ( / ) ruxolitinib initiation was indicated for acute gut refractory/steroid dependant gvhd in episodes and chronic multisystemic in episodes. other indications were chronic lung ( / episodes), chronic skin ( / ) and acute skin gvhd ( / ) . median post-hsct time of ruxolitinib start was days. doses ranged between , - mg/ h depending on age, weight, and tolerance (hematologic and liver toxicities). average duration of treatment was days ( - ). complete response (cr) rate was , %, global partial response (pr) , %, and no response (nr) % (progression in one patient and recent treatment start in other patient). mean time to maximum response was weeks. treatment stop cause was cr in cases, infection in , liver toxicity in . no severe side effects directly related to ruxolitinib treatment were described. conclusions: ruxolitinib has been recently introduced as second line strategy for rescuing corticosteroid-refractory gvhd in pediatric patients. while results of randomized trials are lacking, we present our experience (two centres). the main indications for starting treatment were acute gut and chronic multisystemic gvhd. most patients achieved some grade of response (partial or complete), allowing stopping or tapering corticosteroids. toxicity profile appears to be acceptable. disclosure: nothing to declare. stability of tacrolimus concentration early after allogeneic hematopoietic stem cell transplantation reduces the risk of acute gvhd background: tacrolimus is used as an immunosuppressive drug after allogeneic hematopoietic stem cell transplantation (allo-hsct). it is well known that early concentration level of tacrolimus is correlated with the risk of acute graft versus host disease (agvhd), however, whether range of standard derivation (sd) of early tacrolimus concentration after allo-hsct also affect to the risk of agvhd still remains unknown. here, we investigate the correlation between the range of sd of early tacrolimus concentration after donor hematopoietic cells engraftment and the development of agvhd. methods: we retrospectively assessed patients who underwent allo-hsct in our hospital from - . all patients received standard gvhd prophylaxis by continuous intravenous (iv) tac with starting dose of . mg/ kg/day from day before allo-hsct (day - ) and iv methotrexate on day , , at dose of mg/m , mg/m , mg/m , respectively. tac dosage was adjusted to target the serum concentration of - ng/ml until at least day and then tapered. to evaluate the sd of weekly tacrolimus concentration, the range of sd of tacrolimus concentration at day - (week- ), day - (week- ), day - (week- ) and day - (week- ) were calculated. the difference of the range of sd between the groups that develop or did not develop agvhd was compared by using mann-whitney u test. multivariate analysis was performed by using multiple logistic regression analysis. patients had given written consent allowing the use of medical records for research, in accordance with the declaration of helsinki, and the institutional review board approved the study. results: there were males and females and the median age was years (range, to years). the risks of disease were low-standard in and high in pts. the number of donors were in hla-identical sibling, in hla-mismatched related donor, in hla-matched unrelated donor and in hla-mismatched unrelated donor. thirty-seven patients developed agvhd (grade i-ii; , gradeiii-iv; patients). as a result, the wide range of sd at week- significantly increased the risk of agvhd (agvhd-group; . ± . ng/ml, agvhd+ group; . ± . ng/ml, p= . ). multivariate analysis demonstrated that narrow range of sd of tacrolimus concentration at week- reduce the risk of agvhd (or= . ; % ci: . - . ; p= . ). there were no correlation between gender, age, disease status, hla with the development of agvhd. conclusions: the range of sd at week- , an engraftment phase of donor hematopoietic cells, was significantly correlated with the development of agvhd. fine tuning of early tacrolimus concentration with narrow range of sd reduces the risk of agvhd, resulting in improvement of the overall survival after allo-hsct. disclosure: nothing to declair p ruxolitinib treatment for steroid-refractory graftversus-host disease han-seung park , je-hwan lee , jung-hee lee , eun-ji choi , miee seol , young-shin lee , young-ah kang , mijin jeon , kyoo-hyung lee background: steroid-refractory graft versus-host disease (gvhd) is one of the most lethal complications after allogeneic hematopoietic cell transplantation. recent studies have shown that ruxolitinib, a janus kinase / inhibitor, is effective in patients suffering from gvhd. here, we report a retrospective result of ruxolitinib treatment for steroid-refractory gvhd. methods: all patients had received cyclosporine and a short course of methotrexate as gvhd prophylaxis. antithymocyte globulin was added for unrelated or mismatched familial donor hct. ruxolitinib mg twice daily was added to immunosuppressive treatment in patients with steroid-refractory gvhd. results: a total of patients with gvhd (acute, , including patients with donor lymphocyte infusion [dli]related; and chronic, ) were included in the analysis. all patients had grade / acute gvhd or severe chronic gvhd at the time of ruxolitinib treatment. six ( . %) of patients with acute gvhd responded to ruxolitinib, including with complete response (cr). the median time to response was . days (range, - ) . nineteen patients received ruxolitinib for severe chronic gvhd, with the median of involved organs (range - ). fourteen patients ( . %) showed response to ruxolitinib, including crs. the median time to response was days (range, - ). five responders discontinued ruxolitinib and patients are still on the agent. after a median follow-up duration of . months, died ( from relapse of disease, from infection). the -year survival probability was . %. eleven of responders discontinued ruxolitinib. gvhd relapsed in of patients at , , and days after ruxolitinib discontinuation. thrombocytopenia ( / , grade / ; ) was the most common adverse event of ruxolitinib. during treatment, with grade / infectious adverse events occurred; pneumonias, brain abscess, and liver abscess. conclusions: ruxolitinib treatment seems to be effective for the treatment of steroid-refractory gvhd including long-standing chronic gvhd. the agent was well tolerated and relatively safe. disclosure: nothing to declare. il -receptor antibody tocilizumab as salvage therapy in the treatment of severe chronic gvhd after stem cell transplantation: a retrospective analysis background: severe chronic graft-versus-host disease (cgvhd) remains the most relevant factor affecting survival and long-term quality of life after allogeneic hematopoietic stem cell transplantation (hct). besides corticosteroids there is no established therapy for cgvhd and many of the used immunosuppressive agents may lead to significant toxicity incl. infectious complications. tocilizumab (an il -receptor antibody) has shown efficacy in acute gvhd and cgvhd. we retrospectively analyzed the efficacy and safety of patients having received tocilizumab for treatment of advanced cgvhd at our center between the years and . methods: patients with severe steroid refractory cgvhd and a median age of years (range: - yrs) having received at least two prior lines of therapy for cgvhd (range: - regimens) were treated with tocilizumab for at least one cycle (q w, dosage: mg/kg iv, maximum: mg) with a median number of cycles (range: . nih consensus criteria grading for cgvhd and the immunosuppressive regimen were noted at the time of the first tocilizumab administration and after , and months of therapy. all patients received additional concomitant immunosuppressive agents already given at least weeks without response before start of tocilizumab. no new immunosuppression (is) was added in parallel to tocilizumab and response assessment was stopped at start of any additional new is. all patients had received peripheral stem cell allografts. gvhd prophylaxis consisted of a calcineurin inhibitor in combination with methotrexate or mycophenolate and in case of unrelated donors atg was added. / patients had quiescent onset of cgvhd, one patient developed de novo cgvhd. the median number of days between hct and onset of cgvhd was ( - ). the median number of days between hct and initiation of tocilizumab therapy was ( - ) days. at cgvhd onset, / patients had mild cgvhd and / patients had moderate cgvhd. the thrombocyte count was < /nl in / patients. organs involved at initiation of tocilizumab therapy were skin ( %, all grade ), eyes ( %), mouth ( %), fascia ( %), lungs ( %) and genitals ( %). / patients are still receiving tocilizumab at the time of analysis. results: as tocilizumab was given fairly recently in most patients, -and -month follow-up was only reached in / patients ( %). at three-month follow-up after initiation of tocilizumab therapy, / patients ( %) showed partial remission, / patients stable disease ( %), and / patients progressive disease ( %) of cgvhd. maximal response was partial remission ( %), stable disease ( %) and progressive disease ( %). patients required subsequent new immunosuppressive treatment. one patient has not yet reached -month follow-up. during tocilizumab therapy none of the patients suffered recurrence of underlying malignancy. two patients developed significant respiratory infection and one patient developed soft tissue infection, all requiring antibiotic treatment and pausing of tocilizumab administration, hospital admission was not required. the os and rfs was % with median follow up of . months (range - months). conclusions: tocilizumab appears to be a promising treatment option in advanced cgvhd but further evaluation within a phase ii trial is required. disclosure: nothing to declare background: allogeneic hematopoietic stem cell transplantation (hsct) is for many patients suffering from aml the only curative treatment option. one major complication is graft versus host disease (gvhd), caused by donor immune cells attacking healthy tissue. regulatory t cells (treg) have been getting huge attention during the past years because of their important role in maintaining immune balance. here we collected peripheral blood samples from patients at different time points after hsct to investigate immune-reconstitution of treg as predictive marker for the development of gvhd. methods: we collected blood samples from patients in the course of allogeneic hsct prospectively once a week from d+ up to d+ . all patients received conditioning regimen with fludarabine and melphalan, combined with alemtuzumab for t cell depletion. patients developed acute gvhd in the later course. after isolation of pbmc`s we performed facs multicolor staining of t cell and nk cells. treg were identified as cd + cd + cd ++ foxp + , nk cells were characterized as cd neg cd + cd + and divided in nk cell subpopulation due to their expression of cd dim or cd high . results: . cd neg t cells: all patients developing acute gvhd in the later course showed significant elevated levels of cd + cd neg t cells, especially cd neg treg at d+ . . cd neg treg / cd + cd + t cells: one patient not developing acute gvhd showed lots of cd neg treg but missed cd + cd + effector t cells. we recently showed that cd + cd neg effector t cells are of impaired effector function. these data suggest that cd neg treg are only of relevance combined with functional cd + cd + effector t cells in the development of agvhd. . t cell marker: patients without agvhd showed elevated expression of garp on treg. garp was significantly higher expressed on cd + treg, indicating a better suppressive capacity of cd + treg. this was detected throughout from d+ until d+ . tigit and ilt showed a heterogeneous expression profile without significant differences between the two groups. . nk cells: we detected a higher ratio of cd ++ /cd dim nk-cell population in patients without. we could also show that tigit is mainly expressed on cd dim nk cells. conclusions: we and others showed reconstitution of cd neg t cell subsets after alemtuzumab mediated t cell depletion -our data on effector t cells showed an impaired effector function for cd neg cd and cd t cells. recently we presented data on impaired suppressive capacity of cd neg treg and the association with acute gvhd retrospectively (wölfinger ebmt , ash ). here we provide prospective data on patients after the use of alemtuzumab in the context of hsct: our preliminary data suggest that the total amount of cd neg-treg and the ratio of cd neg treg to cd + cd + treg on d+ after allogenic hsct could predict agvhd. this data may be a basis for immune monitoring of patients at d+ to evaluate their risk for agvhd and could lead to the use of prophylactic treg dli in the context of alemtuzumab mediated t cell depletion. disclosure: medac -travel support, novartis -consultancy fee, pfizer -consultancy fee, shireconsultancy fee background: multiple factors such as disease activity and severity, therapy and/or dietary habits can cause changes in nutritional status independently or by interacting with each other. presence of malnutrition or significant weight loss in chronic gvhd (cgvhd) patients was reported in literature up to %. the aim of this cross-sectional study was to identify factors that affect nutritional status in cgvhd patients. methods: nutritional status in patients with cgvhd treated at the university hospital center zagreb, croatia from to was assessed. anthropometric measurements (height, body weight (bw), body mass index (bmi)) and clinical validated tool patient-generated subjective global assessment (pg-sga) (where patients were categorized as well-nourished (pg-sga a), moderately malnourished (pg-sga b) or severely malnourished (pg-sga c)) were used. all patients were evaluated according to nih criteria for cgvhd diagnosis. descriptive and correlation analysis were preformed. results: in total, adult cgvhd patients were included in the study, women ( . %), median age ( - ) years, with mild cgvhd in ( . %), moderate in ( . %) and severe in ( . %) patients. according to the pg-sga rating ( . %) patients had pg-sga a, ( . %) pg-sga b and ( . %) had pg-sga c, giving a total malnutrition or risk of malnutrition prevalence of . %. the mean bmi was . ± . kg/m with correlation to pg-sga rating (r= . , p= . ). malnutrition according to the bmi (defined as bmi< kg/m ) was found in patients ( . %). bw changes ( % or more in months) were significant in patients ( . %). according to the pg-sga assessment tool, oral symptoms reported by patient ( . %) and decreased appetite reported by patients ( . %) were associated with oral cgvhd nih score (r= . , p= . ; r= . , p= . ) but not with bw or bmi. gastrointestinal (gi) symptoms assessed with sga, were generally mild with no correlation to gi cgvhd nih score. no significant association was found between nutritional status and other nih cgvhd scores. corticosteroid therapy present in ( . %) correlated with pg-sga rating (r= . , p= . ) but not with bw, bmi or appetite changes. in patients ( . %) with altered pg-sga rating, bmi, appetite and body weight changes, dietary counseling and oral nutritional supplementation were initiated. conclusions: oral symptoms, decreased appetite and corticosteroid therapy in our cgvhd patients were associated with altered nutritional status according to the pg-sga, but not with bmi. therefore, pg-sga might be a more sensitive tool in assessment of changes of the nutritional status and detection of patients at risk of malnutrition than bmi since it includes different factors like physical examination, presence of gi symptoms and corticosteroid therapy in its scoring system. nutritional counseling and support are important in cgvhd patients especially in presence of oral symptoms. disclosure: nothing to declare. background: sclerotic skin changes are common features in chronic graft versus host disease (cgvhd). one of the most challenging aspects in the diagnosis and management of sclerodermoid cgvhd (scgvhd) is the differentiation between reversible symptoms related to active cgvhd and nonreversible symptoms related to residual permanent damage such as long-standing fibrosis. although several candidate biomarkers of cgvhd inflammatory activity have been proposed, none of them are currently validated. therefore, there is a need for the development of more quantifiable and reproducible measurements tools to guide clinical decisions. we report our experience evaluating the usefulness of high-frequency ultrasonography (hfus) plus doppler ultrasound (doppler-us) and serum fibrosis biomarkers to determine the inflammatory activity of scgvhd. methods: we report patients with scgvhd. hfus plus doppler-us were performed at diagnosis of scgvhd and at different time-points after treatment initiation. serum hyaluronic acid and pro-colagen-iii were measured as fibrosis biomarkers simultaneously with hfus and doppler-us. nih cgvhd consensus conference diagnosis criteria, scoring system, and response criteria were used to assess global and organ-specific cgvhd, and to measure overall response to therapy. abnormal ultrasound findings were defined as the presence of ≥ of the following: hypoechogenic dermis, dermo-epidermal junction effacement, hypoechogenicity of septa and/or hyperechogenicity of lobules in hypodermis, hypoechogenic fascia, or myositis, for hfus; and, vessels thicker than mm in dermis and/or hypodermis, systolic pressure > cm/sec, and index of vascular resistance > . , for doppler-us. inflammatory activity was classified as mild, moderate and severe according to the severity of doppler-us findings. results: hfsu showed abnormal findings in all patients at diagnosis with no changes except in two patients along the treatment follow-up. inflammatory activity by doppler-us was observed in / patients at diagnosis ( mild, moderate, severe). four patients responded to treatment ( complete responses, cr, and partial responses, pr), one presented clinical improvement less than pr, and one, progressive disease. all patients with clinical response had also a p-rom improvement or normalization. all patients achieving a response showed normalization (n= ) or improvement (n= ) of doppler-us findings. the patient with clinical improvement less than pr and the patient with progressive disease showed persistence of inflammatory doppler-us findings. most patients had normal or light increase of pro-collagen levels at diagnosis and no significant changes were observed during follow-up. levels of hyaluronic acid tended to be very high in patients with progressive scgvhd (patients and ) and tended to decrease or normalize in those who responded to therapy (patients , , and ). conclusions: in this exploratory study, hfsu was a reliable method for evaluating sclerotic skin changes in scgvhd. doppler-us showed a good correlation with disease activity and response to treatment. serum hyaluronic acid levels might be a biomarker of disease activity that deserves further investigation. hfsu plus doppler-us is a useful, non-invasive, repeatable device in monitoring patients suffering from scgvhd. according to our results, doppler-us may be a more sensitive parameter than hfsu in assessment inflammatory activity of scgvhd. disclosure: maría suárez-lledó received a grant from dkms-spain foundation. other authors have nothing to declare post-transplant cyclophosphamide versus antithymocyte-globulin in hla-matched unrelated and haploidentical transplantation for hematologic malignancies background: post-transplant cyclophosphamide(ptcy) and antithymocyte-globulin(atg) are the most commonly used regimens for the prophylaxis of graft-versus host disease(gvhd). we compared these two regimens in hlamatched unrelated (mud) and haploidentical transplantation for hematologic malignancies. methods: we retrospectively analyzed the consecutive adult patients with hematologic malignancies who received mud and haploidentical transplantation at chungnam national university hospital between january and january . patients who received second transplantation and had refractory disease were excluded. results: this study included patients with median age of (range, - ) years: ( . %) patients received mud transplant ( and patients in ptcy and atg group, respectively), and ( . %) patients received haploidentical transplant ( and patients in ptcy and atg group). graft source was peripheral blood stem cell in all patients. median follow-up duration was . months (range, . - . ). in mud transplant, the estimated -months survival rate were . % in ptcy vs. . % in atg (p= . ), the -months relapse rate were . % in ptcy vs. . % in atg (p= . ), the cumulative incidence of grade to acute gvhd were . % in ptcy vs. . % in atg (p= . ), and the estimated -month extensive chronic gvhd rate were . % in ptcy vs. . % in atg (p= . ). in haploidentical transplant, the estimated -months survival rate were . % in ptcy vs. . % in atg (p= . ), the -months relapse rate were . % in ptcy vs. . % in atg (p= . ), the cumulative incidence of grade to acute gvhd rate were . % in ptcy vs. . % in atg (p= . ), and the estimated month extensive chronic gvhd rate were . % in ptcy vs. . % in atg (p= . ). patients receiving ptcy had significantly longer neutrophil engraftment time than those receiving atg in haploidentical transplant [median(range); . ( . - . ) days vs. . ( . - . ) days, p= . ]. conclusions: ptcy might be a good option for the prophylaxis of gvhd in hla-matched unrelated transplant as well as haplo-identical transplant. disclosure: nothing to declare early fam therapy for post allo-hsct bronchiolitis obliterans syndrome background: bronchiolitis obliterans syndrome (bos) is a potential major complication after allogeneic hematopoietic stem cell transplantation (hsct). attributed to an allo-immune reaction against the small airways, bo is considered a pulmonary manifestation of chronic gvhd. reported incidence of bos ranges from to %, and bos-attributed mortality as high as %- %. a few years ago, a new therapeutic approach with fluticasone, azithromycin, and montelukast (fam) was described (norman bc, et al. bmt ) . our aim was to analyze the outcomes of pts who developed bos and were precociously treated with the fam scheme. methods: all the allo-hsct performed in our center from january and july were included in the analysis. baseline diseases were: aml, lpd, mds, all, mpd, mm, and bmf. day + and day + overall mortality were , % and , %, respectively. rest of characteristics of the series are shown in table. fam therapy was systematically started when any patient was first diagnosed with bo. results: eleven patients ( , %) were diagnosed with bos. at diagnosis of bos, the pts exhibited a fev % of predicted (median fev : %; range; - %) and/or a decline > % from pre-hsct . at day + , pts had already the syndrome. two of them died before the end of the first year: one due to invasive zygomycosis (cns plus pulmonary) and the other to baseline disease progression. at day + , more pts had bos. two more pts with bos died at and months post-hsct due to baseline disease progression. at the close of the analysis, of the pts were alive. so, with a median follow-up of months (range: - ), mortality and bos-attributable mortality of the pts with the complication were , % and , %, respectively. conclusions: ) bos is an infrequent but very severe complication of allo-hsct; ) bos seems to be less frequent in pts with prophylactic pre-transplant ratg or post-transplant cyclophosphamide, as well as in pts undergoing transplantation with bm (compared to pbsc). ) early diagnosis and therapy are critical to minimize the bos-attributable mortality. disclosure background: donor lymphocyte infusion (dli) is an established treatment for patients with hematological malignancies relapsed after allogeneic hematopoietic stem cell transplantation (hsct). however, it is associated with an increased risk of graft-versus-host disease (gvhd) and modest anti-tumor activity. compared to the infusion of nonmobilized lymphocytes, granulocyte colony-stimulating factor (g-csf)-primed dli might induce a stronger anti-tumor effect and reduce the risk of infusion-induced gvhd. due to the limited experience of g-csf primed dli in patients relapsed after haploidentical hsct, we conducted a retrospective study of all patients at our hospital who received dli for the relapsed hematological diseases following related hla-matched or hla-haploidentical hsct. methods: the institutional research board approved the study. we identified patients with hematological malignancies receiving dli following related allo-hsct at national taiwan university hospital between and aug. the infusate was obtained from the cryopreserved specimen, which had been collected and stored at multiple aliquots at the same time as the initial haploidentical peripheral stem cell graft. patients received dli for either hematological relapse, preemptive or prophylactic treatment. univariate and multivariate analysis was performed using cox proportional hazard regression model. results: for the patients following related hlamatched and the patients following hla-haploidentical hsct received and doses of dli, respectively. in comparison, the median cd + cell dosage of haplo-dli is significantly lower (p = . ) than that of dli from sibling donors, with median cell dosage . × /kg (range, . - × /kg) and . × /kg (range, . - . × /kg), respectively. the median time to dli from initial sibling hsct and haplo-hsct was days (range, - days) and days (range, - days), respectively. overall, ( %) of the patients following sibling hsct developed grade - acute gvhd after dli, whereas ( %) of the patients receiving haplo-hsct developed grade - acute gvhd after dli (p= . ). importantly, for patients receiving dli with cd + cell dosage less than × /kg, there is no difference in the risk of developing grade - acute gvhd between patients receiving dli from sibling or haplo donors ( figure a) . interestingly, for patients receiving dli with cd + cell dosage more than or equal to × /kg, ( %) of the patients following haplo-hsct developed grade - acute gvhd after dli, significantly more than ( %) of the patients following sibling hsct developed grade - acute gvhd after dli ( figure b) . the cumulative incidence of grade - acute gvhd at day after haplo-hsct and sibling hsct were % ( % ci: . - . ) and . % ( % ci: . - . ), respectively ( figure b , p = . ). [[p image] . conclusions: our study shows that the administration of g-csf mobilized dli is feasible after haploidentical hsct for relapsed hematological malignancies. however, dli with cd + cell dosage more than or equal to × /kg in patients receiving haplo-hsct is associated with significantly higher risk of developing acute gvhd than dli from the sibling donors. disclosure: the authors declare no competing financial interests. background: the fresenius phelix is a uva irradiation device used to photoactivate mnc collected on the amicus. the system is closed, utilizing a special mnc kit and modified instrument software. the preliminary results of a phase i safety trial involving three patients ( treatments) with chronic graft vs. host disease are presented. methods: reasons for transplantation for the patients ages , and years were: acute myelogenous leukemia, myelodysplastic syndrome, and myelodysplastic syndrome with pnh. stem cell source was peripheral blood with a / match for all. each developed chronic skin gvhd. inclusion criteria included wbc and plt counts > and x /l, gfr > ml/min/bsa, and ast - unit/l. exclusion criteria included active gi bleeding, nyha cardiac disease greater than grade iii, and the presence of light-sensitive diseases. amicus software . and phelix software . were used. settings included: ml/min max draw rate, ml fixed cycle volume, . mg/kg/min citrate infusion rate, and : acd-a ratio. venous access was peripheral or subcutaneous port. target uva dose was . j/cm and -methoxypsoralen dose was . ml. results: the following mean + sd procedure results were obtained: , + ml whole blood with acd-a drawn, + ml acd-a used, + ml saline used, + minutes procedure time, and , + ml total blood volume. minor alarms (n= ) on the amicus and no alarms on the phelix were encountered. all -day aerobic and anaerobic cultures were negative and mean endotoxin levels were . + . eu/ml. mean pre/post cbc and plasma hemoglobin levels were: . / . wbc, . / . neutrophils, . / . basophils, . / . eosinophils, . / . lymphocytes, . / . monocytes, / platelets x /l, / % hct, . / . g/dl hgb, and . / . mg/dl plasma hemoglobin. plasma hemoglobin delta in the product was . + . grams and the subject was - . + . grams. collected product hct. mean . + . %. yields are in the table. adverse events included one each: acute respiratory failure, respiratory failure, muscular weakness, musculoskeletal discomfort, and peripheral swelling. three of four events occurred in one patient two weeks after the study procedure. none of the adverse events were considered related to the procedure or investigational product. the patient who experienced acute respiratory failure was removed from the study because of death due to pneumonia, felt to be unrelated to the procedure. conclusions: results indicate the new closed photopheresis system is capable of collecting sufficient mnc and irradiating the cells producing high lymphocyte apoptosis, with minimal alarms and adverse reactions. ( . %)). ( . %) of the patients also had acute gvhd of the skin or liver. patients ( . %) could be treated and controlled with methyl-prednislone monotherapy, patients had steroid refractory gvhd of whom patients ( . %) could be salvaged with additional drugs (infliximab: ; tacrolimus: ); patients ( . %) had refractory acute gut gvhd and could not be salvaged despite more than three lines of therapy. at the time of reporting, patients ( . %) of the are alive. patients died due to transplant related mortality, while patients developed relapsed disease. on binary logistic regression analysis, no baseline clinical or treatment related predictor (disease indication, disease status at transplant, transplant type, graft source, type of conditioning) could be identified for developing acute gvhd of the gastrointestinal system. conclusions: acute gvhd of the gastro-intestinal system is a significant cause for morbidity in allo-hct patients at our centre. further studies are warranted in our cohort, and a prospective analysis of gut microbiome analysis, faecal multi-drug resistance organism surveillance, conditioning related toxicity and antibiotic usage is ongoing. clinical trial registry: not applicable disclosure: the authors declare no potential conflicts of interest benefits and precautions of ruxolitinib in steroidrefractory acute gvhd background: corticosteroids are the standard first-line treatment option for patients with acute graft-versus host disease (gvhd), but approximately half of patients become refractory to steroids and require second-line treatment. ruxolitinib has the potential to treat gvhd in steroidrefractory (sr) patients based on retrospective clinical data. the ongoing prospective trials are currently enrolling patients to evaluate the therapeutic potential of ruxolitinib for gvhd. methods: we analyzed retrospectively clinical experience with ruxolitinib in patients (n= ) with grade ~ steroid-refractory acute gvhd patients compared with the control group not receiving ruxolitinib. in addition, immune status was evaluated about weeks~ weeks after the administration of ruxolitinib using flow-cytometry. ruxolitinib was used as a third option for sr gvhd, combined with previously used immunosuppressive drugs. and steroids were gradually decreased according to the symptoms and discontinued. patients received ruxolitinib mg twice daily (bid), with increase to mg bid if hematologic parameters are stable and no treatmentrelated toxicities. results: fifteen patients all were assessable for response. seven patients achieved a complete response, had a partial response, and had no response at weeks after the first ruxolitinib dose. overall response rate was %. three were treatment failures. most adverse effects were manageable, except infectious complications. infectious complications were occurred in about % patients (n = ), resulting in two deaths. common cause of infectious events included cytomegalovirus (n = ), herpes-zoster (n= ), epstein-barr virus (n= ), fungal infection (n = ), pneumocystis jiroveci (n = ), bacterial infections (n = ), and pneumonia of unknown origin (n = ). t cell counts tended to decreased in the group with ruxolitinib compared with the control group, especially cd cell counts. conclusions: ruxolitinib is effective in controlling sr gvhd and can lead to clinical benefits. however, we need to be aware of the infectious complications because ruxolitinib may lead to increased risk of opportunistic infections or reactivation of latent infections. in addition, common infectious complications are presumed to involve t cell dysfunction. clinical background: graft versus host disease (gvhd), being one of most common life-threatening complication post hsct, contributes significantly to morbidity and mortality. when affecting gastrointestinal tract (gi) it is the major cause of death in early period post hsct. due to widespread tissue involvement in most patients diagnosed with gi gvhd, surgical treatment is rarely considered. methods: among allo-hsct performed in department of pediatric hematology, oncology and bone marrow transplantation in wroclaw, poland during years - , ( , %) cases were diagnosed with gi gvhd. in this study we present cases ( %) which were referred to and benefit from surgical approach. results: . male, years old underwent hsct from matched unrelated donor (mud) due to chronic myelogenous leukemia (cml) and subsequent molecular relapse succesfully treated with donor lymphocyte infusion, followed by agvhd (skin and gut involvement, grade iv). extensive immunosuppression (steroids, mycofenolate mofetile, atg, okt ) resulted in significant resolution of agvhd symptoms. however aggravating severe abdominal pain and lack of gut movement suggesting bowel obstruction. due to presence of acute abdomen patient was immediately directed for laparotomy. resection of constricted bowel segment followed by subsequent laparotomies for secondary obstruction provided complete resolution of abdominal symptoms. after years of follow-up patient is alive and well. . eleven years old male was diagnosed with skin and gut grade iv agvhd on day + post mud-hsct performed due to acute myelogenous leukemia (aml). he received pronlonged immunosuppressive treatment including steroids, antibodies, msc and ecp which led to resolving of skin leasions and diarhoea. nevertheless patient was suffering from severe paroxysmal abdominal pain and incidentally vomiting. ct enterography showed partial small bowel constriction. after numerous surgical consultations, eventually on day+ patient underwent laparotomy with constricted bowel resection. histopatological examination of resected tissue revealed moderate gvhd. immunosuppersion was tapered to low dose of steroids with ecp. for now, years post hsct patient is alive, rarely experiencing mild abdominal cramps . fourteen years old female developed severe abdominal pain and high volume diarhoea on day + post mud-hsct performed for severe anaplastic anemia (saa). despite extensive immunosuppression (steroids, anti-tnf, anti-il antibodies) patient condition did not improved. through consistent stomach pain, suspected subileus confirmed by ct enterography, laparotomy was performed (day+ ). resection of inflamated and obstructed bowel was made. microscopic evaluation confirmed prior gvhd diagnosis therefore immunosuppression including csa and tapered doses of steroids was continued. complete resolution of abdominal symptoms was almost immediately achieved post-surgery, however months after recurrent abdominal cramps were observed and are now well controlled by pain killers. conclusions: commonly gi gvhd is diffused inflammatory process. however in some cases it may be localized and may lead to partial bowel constriction. in case of severe and prolonged stomach pain, despite of partial resolving of other gvhd symptoms, ileus should be considered. ct enterography may be useful for diagnosis confirmation. in those patients, surgical intervention may improve quality of life or even be a salvage approach. disclosure: nothing to disclose is there any impact of the uric acid levels during the preand early post-graft infusion period, on the gvhd occurence and allotransplant outcome? . ( . - . ) years, who underwent allogeneic stem cell transplantation (allosct) from full-matched sibling donors for acute leukemia (n= ), very severe aplastic anemia/pnh (n= ), lymphoma (n= ), myelodysplastic/ myeloproliferative syndrome (n= ) . thirty-two patients were in remission at the time of allosct (cr : , cr : , beyond cr : ). for a better and more accurate assessment of the ua levels on the agvhd incidence, unlike to the other published studies which evaluated the ua levels only at day , we evaluated the ua levels in different time points during the the peri-transplant period (at the conditioning regimen initiation, and at days , + and + ). because the majority of our patients developed agvhd within the - days post-transplant, we did not incorporated in the study the of ua levels beyond the + day. we also investigated the effect of the ua on survival and the non-relapse mortality (nrm). the vast majority of patients received allopurinol from the st day of conditioning regimen till day - . the independent t-test, kaplan-meir method and logrank test were used in the statistical analysis. results: the median ua levels were . , . , . and . mg/dl at days - , , + and + respectively. for the statistical analysis purposes, we grouped our patients as low-ua if they had values < mg/dl or high-ua if they had > mg/dl. this threshold was chosen based on the ua values from all the collected samples (n= ). finally / ( %) patients developed agvhd; ( %) were assessed as gr ≥ii, while ( %) as gr iii-iv. the incidence of the agvhd gr ≥ii was similar (ranged from - %) in both groups of patients (low-ua and high-ua) and for all the estimated time points (days - , , + , + ). we noticed a better -years overall survival for patients with low-ua ( % vs. %) however without any statistical significance. ten patients succumbed to nrm causes; / deaths attributed to gvhd complications. the nrm was assessed higher in the high-ua group ( % vs. %) but also this difference was not statistically significant. conclusions: though our study bears the limitations of the small number of patients and the retrospective origin, at least to our knowledge is the first which evaluates the impact of ua levels at different time points in the peritransplant period, on the agvhd incidence. in our study the ua levels did not influence the incidence or the severity of agvhd. the higher nrm rates for patients with ua> . mg/dl merits further evaluation. definitely, the role of ua on the allosct outcome will be clarified through well designed prospective trials. disclosure results: five male patients ( %) had genital cgvhd manifestations presented by urethral stricture in / patients and phimosis requiring surgical treatment in one patient. all five patients had simultaneously cutaneous, oral, and/or ocular cgvhd manifestations. the first patient underwent urethroplasty of bulbomembranous part of urethra with termino-terminal anastomosis and urethroplasty of penile part of urethra with buccal mucosa autograft -bmg (dorsal onlay) that resulted in significant improvement of symptoms and normal miction afterwards. biopsy of the urethra showed mononuclear infiltration in lamina propria consistent with cgvhd. biopsy of the buccal mucosa was done prior to surgery and was negative for cgvhd involvement. the second patient underwent urethrotomy due to circular strictures, but symptoms reappeared again and he is now candidate for bmg. in two patients urethral dilatation was done, and the fifth patient presented with phimosis requiring circumcision, resulted in significant improvement of symptoms. conclusions: male genital cgvhd is an underrecognized and under-reported manifestation. patients after allo-hsct need to be actively asked about their genital symptoms and sexual function, especially if they are diagnosed with other mucocutaneous or ocular cgvhd. multidisciplinary approach, early recognition and frequent follow-up is necessary for timely start of treatment. new methods, such as bmg for cgvhd patients with urethral stricture seem promising and should be further investigated. disclosure: nothing to declare. p abstract withdrawn. heracles: a phase ii single-arm prospective study to assess the efficacy of fecal microbiota transfer in the treatment of steroid refractory gastro-intestinal agvhd post allo-hsct background: steroid-refractory acute graft-versus-host disease (sr-agvhd) is associated with an % mortality rate and reduced quality of life (qol). so far, there is no approved standard of care for agvhd second-line treatment. there is an urgent need to identify effective therapy for sr-agvhd to improve patients' outcomes. fecal microbiota transfer (fmt) might be beneficial to substantially improve the prognosis. higher gut microbial diversity is strongly associated with increased survival in gvhd patients. recent studies reported promising results of sr-agvhd patients treated with fmt. further evaluation to confirm the efficacy and safety of fmt for agvhd is warranted. the ongoing phase study (heracles) investigates the efficacy of allogeneic fmt in the treatment of patients with sr-agvhd. heracles was launched after the odyssee study showed promising results in the reconstruction of gut microbiota diversity after induction chemotherapy with fmt in acute myeloid leukemia patients. we expect that fmt-based biotherapeutic drugs could be effective treatments to contain sr-agvhd, and thereby reduce the risk of life-threatening complications after allogeneic hsct. methods: heracles is a single-arm, multicenter prospective trial in european countries. patients aged ≥ years-old, who underwent allogeneic hematopoietic stem cell transplantation (allo-hsct) and developed a first episode of stage or agvhd with gut predominance resistant to a first-line steroid therapy are eligible for inclusion. main exclusion criteria comprise the use of other second-line gvhd therapy, patients with grade iv hyperacute gvhd, late onset agvhd, and overlap chronic gvhd and agvhd after donor lymphocyte infusion. patients receive a first maat enema within days after sr diagnosis (v ) and additional ones week apart (v / v ) from each other. maat is a highly-diverse, microbiome-rich enema formulation obtained from pooled, rigorously screened faeces from healthy donors, manufactured with a standardized process using the signature maat microbiome restoration biotherapeutic (mmrb) platform. at inclusion (v ), before each dosing (v , ), and days post inclusion (v ), patients' faeces and blood are collected. safety monitoring will be performed with corresponding blood analyses. exploratory measures on faeces include characterization of gut microbiota composition and evolution, impact of maat on metabolism, and gut inflammation. immune system phenotyping will be performed by flow cytometry on peripheral blood mononuclear cells, and by elisa assay on plasma. patients' qol will be assessed using a standard, eq- d- l questionnaire. the primary objective is to assess the efficacy of maat by evaluating complete response (cr, according to modified glucksberg criteria) and very good partial response (vgpr, defined by martin et al., bbmt, ) days post-inclusion (primary follow-up). secondary objectives include fmt safety assessment and evaluation of fmt impact on several endpoints, such as overall, relapsefree or gvhd-free survival and chronic gvhd evaluation, as well as multi-drug resistant bacteria carriage. patients will be followed-up until year after inclusion. overall, patients are planned to be enrolled and treated, to assess overall response rates and maat 's safety profile. results background: anti-programmed cell death protein (pd ) monoclonal antibodies can be used as "bridge to" a subsequent allogeneic hematopoietic stem cell transplantation (hsct) in patients with relapsed/refractory hodgkin´s lymphoma (hl). this strategy has been reported to be effective, but a frequent onset of steroid-refractory graft versus host disease (gvhd) was also reported. we report clinical cases of patients affected by hl undergoing allogeneic hsct after having been treated with nivolumab. methods: the patients of , and years respectively had advanced hl and had relapsed after a previous autologous ( ) or allogeneic ( ) hsct. they underwent a rescue therapy with , , nivolumab cycles respectively, depending on the time of partial response achievement and the availability of a donor. two patients received a thiotepa-fludarabinecyclophosphamide conditioning, atg-based prophylaxis and pb cells from unrelated donors. the third patient received bm cells from an haploidentical donor using the "baltimora" nonmieloablative platform. results: at a follow-up of , , months after hsct, respectively, all patients achieved and maintained a complete remission by pet-ct scans. all the patients developed acute gvhd on day + , + and + , respectively. patient progressed to grade iv acute gvhd with hepatic and intestinal involvement unresponsive to first line mg/kg steroid therapy and second line etanercept plus extracorporeal photopheresis (ecp). third line therapy with ruxolitinib partially controlled the gvhd. gvhd onset in patients and was preceeded by a prolonged fever without microbiological findings. patient developed hepatic grade ii gvhd with high transaminase levels, initially responsive to steroid therapy, then it progressed to gut requiring second line therapy with etanercept. patient progressed to severe chronic gvhd with skin involvement and resulted unresponsive to steroids and ecp and it was partially controlled by ruxolitinib. immune reconstitution was delayed in all patients: at months post transplantation cd levels were /μl, / μl and /μl and cd levels were /μl, /μl and / μl respectively. only patient , that underwent haploidentical transplant and received post-trasplant cyclophosphamide (pt-cy), is off of immunosuppressive treatment at months after hsct, without evidence of gvhd and no history of infections. out of the patients receiving pbsc from unrelated donors and atg prophylaxis, patient developed a disseminated fusariosis on day + and died of cns fusarium localization year after hsct, despite targeted antifungal therapy. patient had pulmonary aspergillosis, sepsis by multidrug resistant psuedomonas aeruginosa and otomastoiditis: at + months after hsct, he is on ruxolitinib treatment with skin clinical partial response. conclusions: this case series confirms that nivolumab as "bridge to transplant" is effective in appropriately selected patients. however, risk of acute gvhd and delayed immune reconstitution may require a careful consideration at the moment of planning the transplant. a possible advantage of pt-cy gvhd platform and haploidentical donors should be addressed in larger studies. background: acute graft-versus-host disease (agvhd) is the most important complication after an allogeneic hematopoietic stem cell transplant (hsct). no standard secondline treatment has been established for the corticosteroid refractory agvhd. the anti-tnfα agents are a good option of treatment for these patients, especially when lower gi tract is involved. methods: from april to july we reviewed the outcome of patients with steroid-refractory (sr) agvhd treated with etanercept as at least, second line treatment. etanercept dose was mg twice a week for the first weeks, followed by weekly doses. results: median age was years (range - years), and patients ( %) were male. fourteen patients ( %) had a non-advanced disease status at hstc. eleven patients ( %) received a myeloablative conditioning, and the stem cell source was peripheral blood in patients ( %). sixteen patients ( %) were / hla matched. the characteristics of the patients, their agvhd stage previous to rescue treatment with etanercept and their outcome are shown in table . seventeen patients ( %) had a classic agvhd while had a late-onset agvhd. etanercept was given as a nd , rd and th line in ( %), ( %) and ( %) patients respectively. the median doses of etanercept administered were (range - ), and just patients ( %) completed the doses planned treatment, of whom were alive at , and months from the onset of rescue treatment. complications during etanercept treatment were: infection (n= [ %]: gram negative bacilli [n= ]), grade - neutropenia (n= ) and grade - thrombocytopenia (n= ). etanercept was indicated as a rescue treatment due to: progression after days of agvhd treatment (n= ), no response after days of treatment (n= ), no complete remission after days of treatment (n= ) and relapse due to decrease corticosteroid doses (n= ). at the end of treatment patient achieved a complete response and patients a partial response, all of them are alive. these patients received etanercept as a nd (n= ) and th line (n= ) treatment, all of them had lower gi agvhd without any other organ significantly involved. causes of death were: agvhd with or without infection in patients ( %) and leukemia relapse in patient. conclusions: although if etanercept is an option for treatment of sr agvhd in some patients, their prognostic remains poor and more effective alternative strategies are needed. a prompt initiation of etanercept as a rescue treatment for sr agvhd is crucial to improve the prognosis. ( ) ( ) background: although both cyclosporine (csa) and tacrolimus are calcineurin inhibitors, csa is more widely used in pediatric hematopoetic stem cell transplantation (hsct) as a prophylactic drug for acute graft versus host disease (agvhd). there are some clinical experience but very few data about the clinical efficacy of conversion to tacrolimus. here, we present our single center data on this arguable topic. methods: this study involves the data of pediatric hsct patients in medical park göztepe hospital between - . all patients had prophylactic csa therapy and for various reasons csa was converted to tacrolimus therapy. most of the patients had this conversion due to agvhd. as steroid is the first line therapy for agvhd, conversion to tacrolimus is done concurrently at the start of steroid therapy (within hours after the start of steroid). and also, patients who had any other immunosupressive therapy for agvhd are excluded. response is defined as resolution of symptoms within days after conversion. results: mean age of the study population is months ( - months), male/female ratio is , ( / ), donor types are mud patients ( %), mfd patients ( %), haplo patients ( %) and mean conversion time is days ( - days) . the rationales for conversion are agvhd for patients, unproper csa plasma levels for patients, allergic reaction for patients, nephrotoxicity for patients, hepatotoxicity for patients, severe headache for patients, high arterial blood pressure for patients and one each for refractory vomiting, autoimmune thyroiditis and visual disturbance. the subgroup analysis of agvhd patients reveals that mean conversion time for agvhd is days ( - days) and there are only responders whose agvhd resolve completely (% ) after conversion. all of the patients had proper tacrolimus levels after conversion due to unproper csa levels and also patients in allegic reaction, severe headache, visiual disturbance and refractory vomiting group responded to conversion completely but only one of the patients in nephrotoxicity group responded and also of the patients in hepatoxicity group responded. the only one patient suffered from autoimmune thyroiditis did not respond to conversion. conclusions: in this study, it is obvious that there are response to conversion for some specific adverse effects of csa and tacrolimus is a good alternative for the patients who have unproper csa levels. conversely, the high percentage (% ) of non-responders shows that it is not feasible to make a conversion to tacrolimus for acute gvhd. disclosure: nothing to declare background: capillary leak syndrome is caused by the dysfunction of the vascular endothelial cells,and is characterized by weight gain,generalized edemas,unresponsive to diuretic treatment,and hypotension.it usually develops in the first days post hsct.and it is of great difficuty to distinguish from other complications which are occured post the allo-hsct. to diagnose this complication at the early stage,it is very difficulty. methods: a -year-old man was admitted to ningbo first hospital for its abnormal in the peripheral blood .he was diagnosed with aml-m by the classical morphology and immunophenotype.cytogenetic evaluation showed a normal , xy( ).the patient achieved cr with induction therapy including idarubicin, cytarabine and etoposide. after consolidation therapy,an allo-hsct from hla identical related dornor( -year-old male, donorrecipient matched by high resolution hla typing at hlaa, -b, -c, drb , and dqb , / matches) was performed.the recipient received conditioning with busulfan, mg/kg/day injection for days; cyclophosphamide, mg/kg/day injection for days; cytarabine, g/m /day injection for day; semustine, mg/m /day orally for day; donor peripheral blood stem cells (pbsc:mnc: . × /l, cd +: . × /l) were mobilized, pheresed and administered to the recipient. gvhd prophylaxis consisted of traditional cyclosporine, short-course methotrexate ( mg/ m at day + , mg/m at days + , + , and + ) and cyclosporin a injection mg/kg qer day was mot reduced untill the hematopoietic reconstitute sucessfully . on day + , complete donor chimerism was acheieved. the csa was gradually reduced and tapered.on day + ,the patients was manifested with increasing in the time and volume of the faeces, he was diagnosed with ii°gvhd (gut).the standarded does of immunosuppressive drug including methylprednisolone and cyclosporin a was administrated. the immunosuppressive drug was gradually reduced when the gvhd was controlled.on day+ ,the patient felt distress and the distress was not related to with the exercise,the temperature was normal,and he did not gain weight.there was no edema in the body.laboratory test including routine blood test,c-reactive protein,procalcitonin,blood gas analysis,cmvdna,ebvdna was normal. the ct scan shows that the pleural is filled up with water, and could not be enlarged promptly, there is pericardial effusion in the body.pulmonary function test shows that reduced function in ventilation and diffusion fuction.the laboratory test of the pleural effusion was normal,the blast cell was not detected in the pleural effusion,the cd + cell count was below the dectable level,the next generation sequencing for minimal residual disease shows that there was no gene mutation .thus, post capillary leak syndrome was considered .sirolimus was adopted and taken the place of cyclosporin a,immunoglobulin was adminstrated to reduce the edema. results: taking together comprehensively,the effusion in the pleural and cardiac was absorbed well. conclusions: occurance of capillary leak syndrome is rare,there is limited data about capillary leak syndrome. comprehensively,the mechanism of cls has not been totally identified.and there is no standard treatment to treat the complication.at present,the cls of this patient was absorbed well by administrating sirolimus,closely followup is needed. disclosure: nothing to declare graft-versus-host diseasepreclinical and animal models p short-term krp and posttransplant cyclophosphamide for graft-versus-host disease prophylaxis emi yokoyama , daigo hashimoto , takahide ara , eko hayase , takanori teshima hokkaido university faculty of medicine, hematology, sapporo, japan background: post-transplant high-dose cyclophosphamide (ptcy) in combination with other immunosuppressants such as calcineurin-inhibitors (cis) has been increasingly used as gvhd prophylaxis after hla-haploidentical or matched hematopoietic stem cell transplantation (hsct). however,cis could hamper reconstitution of regulatory t cells (tregs) and tolerance induction after hsct, facilitating us to develop novel ci-free/ptcy-based gvhd prophylaxis. in the current study, we developed a novel gvhd prophylaxis in which ptcy was combined with short-term administration of krp , a selective agonist of sphingosine- -phosphate receptor type (s pr ), using murine models of mhc haploidentical bone marrow transplantation (bmt). methods: b d f (h- b/d ) recipients were lethally irradiated and transplanted with bone marrow cells and splenocytes from allogeneic b (h- b ) donors. cy at a dose of mg/kg was intraperitoneally injected into the recipients on day + , and krp at a dose of . mg/kg was orally administrated daily from day to day + after bmt. donor t cells in the target organs and secondary lymphoid organs were evaluated by flow cytometric analysis. plasma levels of tnf-α were determined using cytometric beads array. to evaluate graft-versus-leukemia (gvl) effects, recipient mice were intravenously injected with luciferase-transduced p cells (p -luc) on day , and in vivo bioluminescence imagingwas conducted weekly after bmt. results: severe gvhd was developed in allogeneic recipients and all mice died by day after bmt.ptcy alone at a dose of mg/kg significantly ameliorated gvhd and % of ptcy-treated allogeneic recipients survived. oral administration of krp alone enhanced contraction of donor t cells in the lymph nodes and also ameliorated gvhd as has been previously shown with multi-s pr agonist, fingolimod. next, we tested if shortterm krp on days to + added to ptcy enhances anti-gvhd effects of ptcy. we found that survivals of ptcy+krp group were significantly prolonged compared to those of ptcy-alone group ( figure a) . plasma levels of tnf-a, clinical gvhd scores ( figure b) , and donor t-cell infiltration into the target organs such as the gut and skin were also significantly reduced in ptcy +krp group compared to ptcy-alone group (figure c and d) . unlike cis, addition of krp to ptcy promoted treg reconstitution after bmt. finally, bioluminescence imaging demonstrated that proliferation of p -luc injected on day was significantly delayed in ptcy +krp -treated allogeneic recipients compared to control mice transplanted only with t-cell depleted bone marrow cells, suggesting that significant gvl effects persisted in ptcy+krp -treated recipients. conclusions: a combination of short-term krp and ptcy is a promising novel calcineurin-free gvhd prophylaxis in mhc-haploidentical sct. we recently showed that donor inkt cells can be expanded ex vivo and that they are able to prevent activation and proliferation of alloreactive donor t cells while promoting efficient graft-versus-leukemia effects (schmid et al. ). however, the underlying mechanisms how human inkt cells induce immune tolerance after allogeneic hct are not fully understood. methods: monocyte-derived dendritic cells (dcs) were cultured in a mixed lymphocyte reaction with mhcmismatched t cells and culture-expanded inkt cells. tcell activation and proliferation was analyzed by multiparametric flow cytometry and released cytokines were measured via multiplex analysis. transwell assays and imaging flow cytometry were performed to elucidate cellcell interactions. bead-controlled flow cytometry-based cytotoxicity assays were used to evaluate dc apoptosis. apoptotic dcs were then purified by fluorescence-activated cell sorting to investigate their tolerogenic potential to prime regulatory t cells (tregs). results: the addition of inkt cells to mixed lymphocyte reactions resulted in a significantly reduced activation and proliferation of mhc-mismatched t cells. transwell assays and imaging flow cytometry revealed a cell contactdependent mechanism between inkt cells and dcs leading to apoptosis with increasing dna fragmentation of dcs over time. interestingly, various fluorescence-activated single cell sorted inkt-cell subsets were all able to induce apoptosis of host dcs. multiplex analysis revealed that dcs triggered inkt-cell release of cytotoxic factors like perforin, granzyme b and granulysin. blocking the inktcell receptor engagement with a cd d antibody prevented inkt-cell degranulation as well as the subsequent induction of host dc apoptosis. inhibition of cytotoxic factors also abrogated apoptosis of dcs. in turn, sorted apoptotic dcs induced tolerogenic dcs characterized by a high expression of pd-l in mixed lymphocyte reactions. such tolerogenic dcs promoted the expansion of cd + cd + foxp + tregs and prevented activation and proliferation of mhcmismatched t cells. conclusions: we propose a novel mechanism how culture-expanded human inkt cells prevent gvhd after allogeneic hct. host dc apoptosis through donor inkt cells induces a tolerogenic immunoenvironment characterized by pd-l high dcs and expanding donor tregs inhibiting activation and expansion of alloreactive donor t cells. our findings pave the avenue for clinical translation of adoptively transferred culture-expanded inkt cells in humans. disclosure: nothing to declare results: vip-ko mice transplanted with allogeneic tcd bm alone had increased graft rejection with lower levels of donor chimerism and % day survival compared with % survival of wt recipients. transplanting tcd bm plus × e donor t cells from b .br or balb/c donors in vip kio mice led to > % donor chimerism and significantly increased gvhd-mortality compared with wt recipients, with % vs % survival in the b .br-->b model (p< . ), and % vs % survival in the balb/c-->b model (p< . ). donor-derived t cells in vip-ko recipients had significantly higher th and th polarization, with higher rorγt in both cd + (p< . ) and cd + (p< . ) t cells, and higher frequencies of ifn-γ (p< . ), tnf-α (p< . ), and il (p< . ) in cd + and cd + t cells compared to wt recipients. b .br-->b second allogeneic transplantation of radiation chimeras caused lethal gvhd mortality in vip-ko-->vip-ko and wt-->vip-ko mice, but not in wt-->wt or vip-ko-->wt b mice, demonstrating the protective effect of vip was due to synthesis by non-hematopoietic recipient cells. immunofluorescent imaging of allo-bmt recipients showed marked up-regulation of vip in lungs post-transplant and high vip production within neurons innervating the lungs. finally, we demonstrated that short-term administration of vip ( mcg/day) from day to day prevented gvhdmortality in vip-ko recipients transplanted with b .br-->b mhc donor bm & t cells. conclusions: the absence of vip in recipient cells led to increased graft rejection in the absence of donor t cells and increased lethal gvhd when donor t cells were transplanted, indicating vip induced post-transplant regulates allo-reactivity of host graft-rejecting lymphocytes and donor gvhd-causing t cells. the protective effect of parenteral vip administration suggests vip-mimetics represent a novel approach to prevent and treat gvhd. these data also suggest a mechanism of action for the mitigation of gvhd by alpha- anti-trypsin (aat) whereby aat inhibits the proteolytic inactivation of endogenous vip. disclosure: dr. waller reports personal fees and other support from cambium medical technologies, grants from celldex, personal fees from kalytera, grants and personal fees from novartis, grants and non-financial support from pharmacyclics, and equity ownership in cerus corporation and chimerix outside the submitted work. in addition, dr. waller has intellectual property related to vip signaling that has been licensed to cambium oncology in which he holds equity. low-density neutrophils expansion is associated with acute graft versus host disease in allogeneic hematopoietic stem cell transplant patients background: low-density neutrophils (ldns) are distinguished from normal-density neutrophils (ndns) by their anomalous sedimentation within the mononuclear cell fraction after density gradient centrifugation of peripheral blood (pb). by analysing ldns and ndns from g-csfstimulated donors or lymphoma patients, we have previously demonstrated that, depending on physiopathological conditions, immature cd b + cd -ldns can promote t cell survival and ifn-γ production, while mature cd b + cd + ldns can exert immunosuppressive proprieties. aim of this study was to establish the frequency of cd b + cd and/or cd b + cd + ldns in pb of allogeneic hematopoietic stem cell transplant (hsct) patients throughout immune reconstitution, and verify their potential correlation with acute graft versus host disease (agvhd). methods: patients undergoing hsct in our institution between december and june were prospectively enrolled in the study upon informed consent and after institutional board approval. criteria of inclusion were age ≥ years and absence of rheumatologic or viral diseases. pb samples were collected at day + , + , + , + and + after hsct and any time within day + in case of gvhd, before first-line therapy. eight healthy donors (hds) were enrolled as control. mononuclear, polymorphonuclear, and whole blood cells were analysed by flow cytometry after cd vioblue, cd apc-cy , cd b pe-cy , cd pe, cd b fitc staining. cd b + ldns were expressed as percentage of cd + pb mononuclear cells (pbmcs) or cd + whole blood cells and were further characterized based on cd expression. cd b + ndns, expressed as percentage of cd + whole blood cells, were also analysed for cd staining. results: patients (m/f / , median age ) were enrolled in the study. patients received hsct from hlaidentical ( ) or haploidentical ( ) related and from hlaidentical unrelated ( ) donors. after a median time of ( - ) days, patients developed grade ii-iv agvhd. no patients were receiving g-csf at agvhd onset. the scheduled assessments were interrupted in agvhd patients at the beginning of first-line treatment and in patients relapsed of their primary malignancy. no patients developed de novo late-acute or chronic gvhd. starting from day + the frequency of ldns within cd + pbmcs was higher in all patients as compared to hds. the patients that did not develop agvhd showed a decreasing frequency of cd b + cd -ldns, with a progressive increase of cd b + cd + ldns, from day + to + . interestingly, patients with agvhd showed a significantly higher frequency of cd b + cd -ldns as compared to patients without agvhd throughout the same time lapse (i.e. from day + to + ) ( . vs . , p= . ). consistently, patients with agvhd had a significantly lower frequency of cd b + cd + ldns ( . vs . , p= . ). the frequency of mature cd b + cd + ndns was normal in all patients since day + . conclusions: ldns are more represented in hsct patients than in hds, with a significant expansion of the cd b + cd subpopulation (with a parallel decrease of the cd b + cd + subpopulation) in patients with agvhd as compared to those without agvhd. according to the previously demonstrated t cell activating function of cd b + cd -ldns, it is tempting to speculate that the expansion of this subpopulation may contribute to agvhd development. disclosure: nothing to declare background: acute graft-versus-host disease (agvhd) is a major complication after allogeneic hematopoietic stem cell transplantation (allo-hsct) which has negative impact on the morbidity and mortality of the patients. accumulating evidences suggest that abnormalities of foxp + regulatory t (treg) cells contributed to the pathogenesis of gvhd, but the underlying molecular mechanisms still remain largely unknown. methods: in this study, we enrolled all the patients treated with allogeneic hsct at the institute of hematology, chinese academy of medical sciences between and ,as well as age-matched healthy adults as control samples. the ratio of tregs in pb and bm of healthy controls (hcs) and patients with and without agvhd was determined by flow cytometry. the transcription profile between tregs from patients with or without acute gvhd was measured,the pathway enrichment analyses were performed by the kyoto encyclopedia of genes and genomes (kegg) pathway database and geneset enrichment analysis (gsea).the expression of lkb at transcript levels and protein levels was measured by realtime pcr and analyzed by the nanopro tm system. a series of functional assays in vitro were performed to assess the function and stability of tregs from patients with and without agvhd.meanwhile, to assume the affect of lkb on gvhd outcome, we established a murine transplant model,which recipient balb/c animals were transplanted with the same amount of mixture made by bm, cd +cd -tcon cells from c bl/ and cd + foxp yfp+ tregs from either foxp crelkb f/f or foxp cre mice. results: in this study, we demonstrated that bm had decreased frequencies of tregs, accompanied with a reversed lower ratio of tregs frequencies between bm and pb in agvhd patients. meanwhile, the number and function of tregs in bone marrow also affected hematopoietic reconstitution. futhermore,to elucidate these mechanisms which regulate tregs homeostasis, we examined the role of lkb on tregs in patients with agvhd and in agvhd murine model. studies demonstrated that lkb deficient tregs lost foxp expression and weaken suppressor function during agvhd. transcriptional profiling and pathway analysis revealed that nf-kb signaling activation and the impairment of a wide spectrum of immunosuppressive genes in agvhd tregs. further mice experiments suggested that cns methylation might lead to the instability of tregs in agvhd group. transplantation with marrow grafts from foxp crelkb f/fmice exacerbates gvhd lethality. conclusions: these studies indicate that lkb is a critical homeostatic regulator for tregs during agvhd. targeting of lkb therefore represents a novel therapeutic strategy that promote immune tolerance to mitigates the severity of agvhd. disclosure: national program on key basic research project ( program) role of aryl hydrocarbon receptor in intestine after allogeneic hsct in mice won-sik lee , soung-min lee , sj-kil seo inje university, busan paik hospital, hemato-oncology, busan, korea, republic of background: aryl hydrocarbon receptor (ahr) is a ligandactivated transcription factor that is activated by various small molecules from the diet, microorganisms, host metabolism, and xenobiotic toxic chemicals. the function of ahr has been demonstrated as a crucial regulator in intestinal homeostasis. here, we investigated the regulatory role of ahr in intestine of recipients after allogeneic hematopoietic cell transplantation in mice. methods: wild-type (wt) b (h- b ), ido -/-(h- b ) and ahr -/-(h- b ) mice were lethally irradiated and transplanted with x tcd-bm plus x t cells from balb/c donor mice. ahr activation in colon tissue of recipients was determined by the ahr target genes cyp a and cyp b expression using real-time pcr. the recipient mice were monitored every other day for survival and clinical score. histopathology and pathogenic effector cytokine levels in colon tissue were analyzed for evaluating ahr function. results: we observed that cyp a was constitutively expressed in the colon tissue of naïve recipient mice. although the expression levels were increased by tbi conditioning, the additive up-regulation of its levels with donor t cell alloreactivity was not observed. in contrast, cyp b expression was markedly induced in the colon tissue by donor t cell alloreactivity. we further observed that the cyp b expression was significantly decreased in the colon of ido-/-recipients with donor t cell alloreactivity, but cyp a was not changed. ido-/-and ahr-/recipient mice showed higher histopathological score for intestinal gvhd and increasing pathogenic cytokine levels in the colon compared with wt mice. conclusions: our results demonstrate that ahr-induced target gene profiles might be differently induced in intestine by ligand dependent manner after hsct, which affect intestinal gvhd. disclosure: nothing to declare. abstract already published. abstract withdrawn. in vitro platelet activation evaluation in allogeneic hematopoetic stem cell transplanted patients in response to haemostatic stimulation and cytomegalovirus stimulation (gvhd), complication of which one of the risk factor is cmv reactivation. the resultant inflammatory platelet response during the high-risk period of gvhd after allogeneic hsct remains unknown. our study aimed to characterize spontaneous platelet activation during the d and d months after allogeneic hsct, and in response to haemostatic stimulation and cmv stimulation. methods: we compared a group of healthy volunteers to a group of allogeneic hsct patients followed between the th and the th days after hsct. platelet activation was determined by the platelet surface expression of cd p and cd using flow cytometer after stimulation by an haemostatic agent, thrombin-receptor activating peptid (trap) and after stimulation by cmv glycoprotein b. the inflammatory response was determined by the detection of immune mediators, rantes, cd ps, pf , cd l and ccl , using the elisa technique in the stimulated platelet supernatants. results: no platelet activation or molecules release were observed after stimulation by cmv glycoprotein b in both groups. rantes and cd ps baseline levels are spontaneously higher in allogeneic hsc patients than in healthy volunteers. platelets from allogeneic hsct patients can be activated after haemostatic stimulation and release cd ps and rantes. in this situation, platelets release more cd ps, rantes and pf than platelets from healthy volunteers. conclusions: although no platelet activation was detected in response to cmv glycoprotein b stimulation, our study revealed a chronic platelet activation condition during the d and d months after allogeneic hsct with an haemostatic inducible hyper-responsiveness. this leads to the release of molecules with immune-modulating properties involved in the pathophysiology of gvhd. as we move further away from the hsct, that phenomenon seems to gradually weaken. clinical trial registry: nct , fipalloc https://clinicaltrials.gov/ct /show/nct disclosure: nothing to declare p efficient process and characteristics of umbilical cordderived mesenchymal stromal cells as a feasible source for anti-inflammatory therapy background: recently, umbilical cord (uc) has become attracted source of mesenchymal stromal cells (msc), because of abundant sources and ease of collection of fetal origin without invasive process for the donor and low immunogenicity with immunosuppressive ability and tissue repair potency. objectives of this study were to explorer the efficient and safe products and to evaluate the antiinflammatory potency of uc-mscs for the application of acute graft versus host disease (gvhd). methods: informed consent was obtained from mothers planning to have cesarean sections. uc tissue was cut and once cryopreserved. the safety assessment including infections and baby's health and development were done after months of birth, and performed small-scale quality test of the frozen uc. then we initiated to isolate master uc-mscs from frozen-thawed uc by an improved explant method, which was passed for quality test. the master uc-mscs were cryopreserved once and thawed and expanded until p . product cells were cryopreserved in original serum-free cryoprotectant dba-d solution. mixed lymphocyte reaction (mlr) assay co-cultured with uc-mscs was carried out using responder mononuclear cells (mnc) stained with cfse, and proliferation and cytokine secretion were analyzed by flowcytometry. results: uc-msc cultured showed significantly higher proliferation ability compared with those from bone marrow-derived mscs, and positive for cd , cd , cd , and negative for cd , hla-dr. cd , and cd were negative even in the high concentration of ifn-γ, while bm-mscs became positive for hla-dr. pd-l was constitutively expressed in uc-msc, while pd-l was induced by the addition of ifn-γ. in mlr, responder t cell proliferation triggered by allogeneic dendritic cells was inhibited efficiently by rd party derived uc-mscs, in which was induced ido, pge , hgf, and tgf-β analyzed by rt-pcr, and inhibited ifn-γ and tnf-α in the supernatant by cytokine beads array. uc-mscs migrated toward the tnf-α treated mnc and increased regulatory t cells incidence in peripheral mononuclear cells by the coculture. conclusions: these results demonstrated that cryopreserved uc are feasible and efficient source of mscs and frozen-thawed uc-mscs have high anti-inflammatory background: a new protocol is under development on the amicus separator that enables the device to perform ecp procedures. the amicus separator is used with a photoactivation device, disposable kit and -mop to provide ecp therapy in a closed system. the objective of this study was to evaluate the safety and performance of the investigational amicus ecp system in healthy human subjects. methods: an irb-approved written informed consent was obtained from subjects ( male, female). the amicus ecp system processed either , or ml whole blood (n ≥ per arm) using double-needle access and acd-a anticoagulation at a : wb:ac ratio. after mnc collection was completed, the subject was disconnected from the device. -mop ( . ml, μg/ml) was injected directly into the collected mnc product and saline (approximately ml total), which was photoactivated with - j/cm uva light. post photoactivation, the amicus separator reinfused the treated mncs into a transfer pack. subject laboratory and safety parameters were evaluated; in vitro evaluations were performed on subject whole blood, collected mncs, treated mncs, and reinfused cells. lymphocyte and monocyte analysis were performed on samples purified using density gradient separation and cultured for up to days post treatment. results: in procedures, median (range) wb processed was . ( - ) ml using . ( - ) ml of acd-a. procedure time was . ( - ) minutes, including photoactivation. no adverse events were reported. subjects' vital signs and hematology values were unremarkable and within expected values. the wbc count of the collected mncs was . ( . - . ) x /μl, comprised of . ( . - . ) % lymphocytes, . ( . - . ) % monocytes and . ( . - . ) % granulocytes and platelet count was . ( - ) background: transfusion of white blood cells (wbc) causes a number of transfusion reactions and complications, for example transfusion-associated graft versus host disease (tagvht), which still does not have effective treatment and is a fatal complication of transfusions. the only effective method of preventing tagvht is irradiation of blood components with ionizing radiation (x-ray or gamma radiation). but the use of ionizing radiation sources has a number of technical and material difficulties. the emergence of pathogen reduction technologies (prt) in blood components targeted by nucleic acids has opened the possibility of using these technologies as an alternative to irradiating of blood components. several prt demonstrated effective inactivation of wbc in platelet concentrates and blood plasma. so, determination of the influence of prt based on the combined effect of riboflavin (rf) and ultraviolet (uv) on the viability and proliferating potential of lymphocytes in whole blood is important. methods: samples of whole blood were obtained in healthy volunteers. each sample was divided into three unequal parts: untreated control, gamma irradiated, and treated by rf and uv prt (mirasol, terumo bct inc.). mononuclear cells (mnc) were cfse stained, viability and proliferating activity were tested at intervals of hours for consecutive days by flow cytometry. statistical analysis was performed with xlstat . . levels of significance were calculated by mann-whitney test, expressed as p-values (p< , ). results: the median viability of mnc after application of both methods of treatment was over , % on day and decreased to day -median percentage of viable mnc were , % (control group), , % (after gamma irradiation) and , % (rf/uv prt). the median of spontaneous proliferative activity on day of untreated and gamma irradiated mnc did not differ ( , % and , % respectively, p< , ). phytohemaglutenin (pha) induced proliferation on day in gamma-irradiated samples was significantly lower in comparison with control group ( , % and , % respectively, p< , ). in samples treated with rf/uv, spontaneous and stimulated proliferating cells was not detected. median percentage of proliferating mnc was less than , %. the use of this prt on whole blood, as well as gamma irradiation, significantly reduces the viability of lymphocytes during storage for days. conclusions: inactivation of wbc using rf/uv prt is a useful and very necessary bonus for a number of reasons. in one procedure two effects are achieved: infectious and immunological safety. the use of prt on whole blood gives the potential for obtaining pathogen-reduced and immunological safety components of blood, which reduces their material cost and staff loading. the use of rf/uv system does not have such complex security requirements and difficulties in servicing as the use of sources of ionizing radiation. the results demonstrate a promising potential for using this technology as an alternative to irradiation disclosure: nothing to declare p influence of patients´serum after allogeneic stem cell transplantation on t cell proliferation and treg function background: acute or chronic graft versus host disease (a/ cgvhd) is one of the major complications after allogeneic hematopoietic stem cell transplantation (ahsct). application of regulatory t cells (treg) as "immunosuppressive dli" to prevent or treat gvhd is investigated in clinical trials. here we ask the question, if there could be clinical conditions (e.g. cytokines or drug effects) limiting the efficacy of this approach. to face this problem we tested the influence of patients´serum on t cell proliferation and treg function. methods: lymphocytes from healthy donors were incubated with t cell medium ( % aim v + % serum + il /okt ) containing serum from healthy donors or serum derived from patients after ahsct with or without gvhd (n= ). next we evaluated the suppressive function of treg by performing treg suppression assays, also comparing serum from patients suffering from gvhd versus serum obtained from healthy donors (n= ). proliferation of cfse stained t cells was measured after days. to test the effect of immunosuppressive drugs on treg we performed treg suppression assays after incubation of treg with corticosteroids or tacrolimus or the combination of both drugs. results: serum of patients with acute or chronic gvhd had a negative effect on t cell proliferation. to avoid bias tests were performed with samples from patients without or only with low levels of immunosuppressive drugs. incubation with serum of patients without gvhd or with serum of healthy individuals showed no differences in t cell proliferation. treg from healthy donors showed a stronger antiproliferative capacity when incubated with serum derived from patients with gvhd. treg previously incubated with immunosuppressive drugs showed no decreased suppressive capacity. conclusions: components of serum from gvhd patients seem to have an antiproliferative effect on t lymphocytes itself. this fact might influence the clinical course of gvhd, but should not be a limiting factor for therapeutic application of treg dli. even the systemic treatment with immunosuppressive drugs e.g. corticosteroids or calcineurin-inhibitors should not diminish the treg application. in a next step we will analyze serum components responsible for this immunosuppressive effect with multi cytokine assays and proteomic analysis. the aim of our project is to develop new strategies to avoid gvhd and to optimize clinical settings for treg dli. disclosure background: hypercalcaemia can be very severe following stem cell transplant (sct) in some osteopetrosis patients. denosumab is a fully human monoclonal antibody that binds the cytokine rankl (receptor activator of nfκb ligand), an essential factor initiating bone turnover. rankl inhibition blocks osteoclast maturation, function and survival, thus reducing bone resorption. we describe the effective management of hypercalcaemia in a patient with rank mutation osteopetrosis who received a haploidentical sct. methods: our patient was diagnosed with osteopetrosis at year of age with a defect in the tnfrsf a gene which codes for rank and received a maternal haploidentical sct aged years. the patients calcium levels were monitored regularly post sct. denosumab was administered for hypercalcaemia as per laboratory reports or clinical symptoms. the drug was diluted with water for injection to make mg/ml solution to facilitate subcutaneous administration. results: significant hypercalcaemia emerged on day + with a level of mmol/l and treated with hyper-hydration and diuretics. this was ineffective in reducing the hypercalcaemia; therefore denosumab was initiated on day + post-transplant. initial dosing was determined using the only available paediatric case report at . mg/kg. a repeated larger dose of . mg/kg was given days later due to an inadequate response (calcium decreased from . mmol/l to . mmol/l). the calcium decreased to . mmol/l after this dose. four weeks later a third dose was required at . mg/kg as the calcium level had increased to . mmol/l. the dose was further increased to . mg/kg for another four doses and then further increased to . mg/kg for another doses and repeated every weeks. normalisation, but not excessive drop in calcium was achieved with these larger doses. over the month follow up post-transplant there were three admissions lasting less than hours for symptoms of hypercalcaemia. these were managed with denosumab administration and hyper-hydration. the remaining doses were given in an outpatient setting. conclusions: denosumab can be safely used as a first line agent in treating post stem cell transplant hypercalcemia in patients with osteopetrosis. a dose of . mg/kg is required as an initial starting dose in order to control hypercalcemia. this is a new higher dose than previously suggested by the original report. denosumab can be effective even after dilution and safely given in children weighing less than kg. disclosure: nothing to declare methods: the clinical, laboratory and molecular aspects of this italian male patient who developed such a complication were collected and presented in order to discuss the origin, clinical outcome and management of this very rare post-transplant event. results: a -years-old man affected by a high-risk chromosomally abnormal, ph -, mll-pro-b (egil b-i) all relapsed during maintenance treatment, nonresponsive to re-induction chemotherapy, in second complete remission (ii cr) after blinatumumab treatment received a female cb transplant. according to sorror's and ebmt scores he was considered a high-risk transplant. the patient and the cb unit were sex-mismatched, shared the same blood groups and were both cmv+/ebv+. he received a tbf conditioning regimen that was followed by the infusion of . x /kg cd + cb cells. gvhd prophylaxis consisted of rabbit atg, cyclosporine a (csa) and mycophenolate mophetyl (mmf). neutrophil engraftment occurred on day + , whereas platelets were never > . /μl. on day + a cm bulged area became apparent on the left parietal region of the skull. an echotomography showed that the lesion adhered to the bone without infiltrating it and lacked blood vessels and suggested that it may be either a site of disease relapse or an area of infection. at the same time a bone marrow (bm) aspiration showed morphological cr confirmed by immune-phenotypic studies and x-y fish a complete chimera. since the patient was still febrile no biopsy was performed, but on day + the axial diameter of the lesion that on a ct scan showed the same appearance revealed by the previous echo-tomography increased to cm. thus, the lesion was surgically removed and histological examination showed cd +, cd +/-, cd +/-, cd /lca+/-, cd -, cd -, cd -, cd -, and s -neoplastic cells whose phenotype suggested a granulocytic sarcoma rather than a histiocytic sarcoma. immuno-chemistry confirmed this suggestion by showing a nuclear npm positivity. fish studies demonstrated that these neoplastic cells were of recipient's origin. a novel bm aspiration showed cr confirmed by immune-phenotypic studies and fish revealed a complete chimera. since the patient was still pancytopenic due to anti-cmv treatment, radiotherapy with gy in nine fractions were given and the lesion completely resolved. conclusions: a granulocytic sarcoma of recipient's origin occurring three months after a cb transplant is a very rare and unusual event. in order to explain such a complication we suggest that granulocytic sarcoma cells were dormant but already present at the time of pro-b all diagnosis and survived not only the initial all treatment but also the cb transplant conditioning regimen. we can't exclude that immune-suppressive treatments given early post-transplant might have promoted the outgrowth of these neoplastic cell population. disclosure: nothing to declare haemoglobinopathy and inborn errors of metabolism p abstract already published. addition of fludarabine on to anti-thymocyte globulin, busulfan and cyclophosphamide conditioning improves outcomes in low-risk matched-related bone marrow transplantation in children with severe thalassaemia flu-atg-bucy. atg dose was mg/kg in all patients except patients with splenomegaly > cm from costal margin and/or sex-mismatched/maternal donor in whom atg was increased to mg/kg. all patients were younger than years and had no hepatomegaly (liver ≤ cm from costal margin) at bmt. results: actuarial overall survival (os) in the atg-bucy and flu-atg-bucy groups is % and %, thalassemia-free survival (tfs) % and %, gvhdfree and thalassaemia-free survival (gtfs) at a median follow up of . and . months was . % and . % months respectively, which is a significantly improved outcome by log-rank statistics (p= . ) in the flu-atg-bucy group. there was no significant difference between the groups in pre-transplant characteristics and posttransplant complications except for the following: median cell dose more in nd group with total nucleated cell dose of . vs . x cell/kg with p< . ; csa taper started later in the new protocol ( day vs. p= . ); median age at bmt ( . vs. . years, p= . ); number of pre-bmt transfusions (p= . ) and ferritin at bmt ( . vs. . ng/ml, p= . ) were higher in the second group; day and chimerisms were also significantly higher in new protocol (p= . and . respectively). there was a trend towards increased incidence of veno-occulsive disease (vod) and posterior reversible encephalopathy syndrome (pres) on the second group but this difference did not reach statistical significance. conclusions: adding fludarabine and targeted dose increase of atg in the standard bucy context seems to significantly improve outcomes of thalassaemia transplants without contributing to excessive gvhd or infectious complications. this protocol can be easily administered in low resource setting without major additional costs. clinical is an acquired clonal disorder of the hemopoietic stem cells for which the only curative treatment is allogeneic hematopoietic stem cell transplantation. however, there are still few reports on the outcomes of allogeneic hematopoietic stem cell transplantation (allo-hsct) in patients with pnh compared to paroxysmal nocturnal hemoglobinuria-aplastic anemia (pnh-aa) syndrome. our study aimed to compare the outcomes of allo-hsct for pnh with pnh-aa syndrome. methods: the clinical data of pnh patients received allo-hsct (pnh = , pnh-aa = ) in our center from july to june were analyzed retrospectively to compare the outcomes of pnh group with pnh-aa group. the clinical data including male patients and female patients, the median age was years (range - ). all patients had received various treatments before transplantation such as steroids, androgens, cyclosporine (csa), antithymocyte globulin, and growth factors. the median interval from pnh diagnosis to hsct was months (range - ). the conditioning regimen was modified bu/cybased regimen in haploidentical donors and unrelated donors, csa, mycophenolate mofetil (mmf) and shortterm methotrexate (mtx) were administered for graftversus host disease (gvhd) prophylaxis. patients with matched sibling donors were treated with the flu/cybased regimen and csa were administered for gvhd prophylaxis. results: there were no differences of baseline between the groups (p> . ) except gender and haploidentical donors. the median values of absolute nucleated cell counts were . ( . - . ) × /kg in the pnh group and . ( . - . ) × /kg in the pnh-aa group (p = . ). the median doses of cd + cells infused were . ( . - . )× /kg and . ( . - . )× /kg (p = . ), respectively. all patients attained complete engraftment, no patient occurred graft failure. the median time for myeloid engraftment were (range, - ) days in the pnh group and (range, - ) days in the pnh-aa group (p = . ). the median time for platelet engraftment were (range, - ) days and (range, - ) days (p = . ), respectively. with a median follow-up of ( - ) months in the pnh group and ( - ) months in the pnh-aa group (p = . ). in pnh and pnh-aa groups the incidences of grade i-iv acute graft-versus-host disease (agvhd) were . % and . % (p = . ), grade ii-iv agvhd were . % and . % (p = . ); chronic gvhd were . % and . % (p = . ), moderatesevere chronic gvhd were . % and . % (p = . ). in haplo-hsct and msd groups the incidences of infection were . % ( / ) and . % ( / ) (p = . ). no patient occurred early death and relapse. -year estimated overall survival (os) of pnh and pnh-aa groups were . % ± . % and . % ± . % (p = . ), gvhd-free and failure-free survival (gffs) were . % ± . %、 . % ± . % (p = . ). conclusions: the preliminary results indicated that allo-hsct is a feasible choice for pnh with favorable outcomes, time for myeloid and platelet engraftment in pnh group were faster than pnh-aa group. there were no differences in os and gffs between pnh group and pnh-aa group. disclosure: no disclosure pattern of calcineurin inhibitor-associated neurotoxicity in sickle cell disease patients receiving a stem cell transplantation background: allogeneic hsct with a msd represents currently the only curative option for sickle cell disease (scd), limited by a donor availability < %. neurotoxicity (nt) contributes significantly to hsct-associated morbidity and mortality. calcineurin-inhibitor (cni) associated nt ranges from . %- . % (severe nt %- %). the elevated incidence of nt in scd (around %) might be triggered by the systemic vasculopathy of scd, with the brain being the primary target. although both cyclosporine a (csa) and tacrolimus (fk ) have a proinflammatory effect, it is more pronounced in csa. infusion modalities also might impact ( . % after bolus injections versus . % after continuous infusion). methods: in a pilot study, we compared t-cell depleted haploidentical hsct (t-haplo hsct) with msd hsct in patients (pts) with advanced stage scd, using almost identical conditioning regimens. pts ( - years; yrs) with homozygous scd or hbs /+ ß-thal were treated between and . nine pts received a msd bone marrow graft, pts received t-haplo-hsct ( second t-haplo due to graft rejection). immunosuppression consisted of either csa ( msd, t-haplo) or fk ( msd, , in combination with mycophenolate mofetil (mmf). fk was administered as a -hours continuous infusion, csa as -hours bolus injections; both target level adjusted (csa: - ng/ml; fk : - ng/ ml). duration of immunosuppression was > months in thaplo-sct and < months in msd, depending on chimerism. results: cni-related nt was observed in . %, severe nt (pres, visual disturbance, aphasia) in . %. nt was more prevalent in msd (n= , . %) than in t-haplo (n= , . %). the incidence of nt was identical under csa ( / ; . %) and fk ( / ; . %), however the majority of severe nt (all pres) occurred with csa. complete recovery of nt was achieved in all pts either spontaneously or after switching to fk /everolimus or withdrawal of fk . moreover, . % of pts with nt were > yrs, and . % > yrs, suggesting an increased risk with age. only . % of pts with pre-existing cerebrovascular disease experienced post-hsct nt. of note, . % of pts with severe nt also developed mild acute gvhd. the overall (os) and disease-free survival (dfs) with a median follow-up of months in t-haplo-hsct and months in msd hsct was % vs. %, respectively. conclusions: our data confirm an elevated nt risk in scd pts following allo-hsct. importantly, the incidence of nt seems to be related to age ( % of pts with nt were > yrs), donor source (msd . % vs. t-haplo . %) and type of cni inhibitor where almost all severe nt ( . %, particularly all pres) was observed under csa. continuous infusion of fk vs. bolus injections of csa might have levelled concentration peaks. the nt observed with csa could be the consequence of predominantly csarelated vascular toxicity inflicting pre-damaged vessels in scd. the mechanism of action could be related to other systemic endotheliopathies such as vod, tam and agvhd, which was observed in . % of pts with severe nt, compared to an overall agvhd rate of %. disclosure: nothing to declare background: matched-related bone marrow transplantation (bmt) may cure over % of low-riskchildren with severe thalassemia (st) defined as a thalassemia syndrome with inability to keep a spontaneous hemoglobin > g/dl. it is well known that patient status at the time of transplant is critical in predicting transplant outcome. liver size > cm is an established adverse prognostic factor in terms of transplant-related mortality and, in our own experience,a spleen size > cm from costal marginis associated with increase rejection rates (blood vol. no. suppl ) . optimising liver and spleen size prior to transplant is likely to improve transplant outcomes. methods: we retrospectively reviewed the effectiveness of our strategy to reduce liver and spleen size pre-transplant using hydroxyurea, super-transfusion and intensive iron chelation. we considered liver size < cm and spleen size less than cm below costal margins as good risk features. liver biopsies were not performed thus pesaro risk classification could not be assigned. all transplant candidates were started on hydroxyurea for a minimum of months and pre-transfusion haemoglobin was maintained > gm/dl while on hydroxyurea. if the child had hepatospenomegaly at enrollmentand no improvement in liver and spleen size after an adequate trial of hydroxyurea (minimum of months of treatment achieving maximum dose of mg/kg day or tolerable haematological toxicity, i.e. neutrophil count between and /μl and/or platelet count between . and . /μl) patients were given a trial of supertransfusion maintaining haemoglobin above g/dl) for a minimum of months prior to declaring the patient as having failed downstaging. results: out of transplants across collaborating centers in india, patients had no hepatosplenomegaly at enrolment and hence were not actively downstaged. twelve patients were excluded due to inadequate information on their records. all of the remaining patients with enlarged liver and/or spleen were downstaged to low-risk features. all patients received adequate hydroxyurea trial among which seven ( %) patients required super transfusion in addition to maximal hydroxyurea. out of the patients ( %) were successfully down-staged with the above strategy and proceeded to transplant as low-risk patients. among the remaining ( %) patients had liver > cm and one had a spleen > cm only. there was significant improvement in liver and spleen size from the time of enrollment to transplant (p value . and . respectively by wilcoxon test for paired samples -two tailed) with median duration of downstaging of months (range - months). there was no significant difference in overall survival (os) and disease-free survival (dfs) by log rank test between the downstaged group and those who did not have hepatosplenomegaly at enrollment (p value . . respectively). conclusions: in the majority of children with thalassaemia and high transplant risk features liver and spleen size can be reduced pre-transplant using hydroxyurea and supertransfusions thereby decreasing transplant risk. disclosure: nothing to declare p abstract already published. abstract withdrawn. longitudinal analysis of the effect of hematopoietic cell transplantation on ocular disease in children with mucopolysaccharidosis i shows ongoing disease progression background: corneal clouding is seen in nearly all patients with mucopolysaccharidosis- (mps- ) causing visual impairment. hematopoietic cell transplantation (hct) is able to stabilize disease in many organs including the brain. however, residual disease in peripheral tissues is often described. therefore, the aim of this study was to determine the long-term effect of hct on ocular disease in mps- patients. methods: corneal clouding (grade - ) and visual acuity (decimal scale) were prospectively collected from all consecutive mps- patients treated with hct between and at the umc utrecht. the primary outcomes of interest, the effect of time on corneal clouding and visual acuity, were analyzed using a linear mixed model. the correlation between corneal clouding and visual acuity was analyzed with pearson's rho. other parameters studied were clinical phenotype, age at time of transplantation and hematological enzyme level after transplantation. other outcomes of interest analyzed included intra-ocular pressure, refraction, and macula and lens abnormalities. [[p image] . results: successfully engrafted mps- patients were included ( % with > % chimerism and normal enzyme levels after hct). corneal clouding stabilized during the first years after hct, but increased rapidly beyond three years (figure ). other predictors for increased corneal clouding were age at time of transplantation ( . , %ci . : . ; p= . ) and clinical phenotype (- . , %ci - . :- . ; p= . ). visual acuity also worsened significantly over time (- . , %ci - . :- . ; p= . ). corneal clouding was strongly negatively correlated with visual acuity (ρ - . , p = . e- ). conclusions: after initial stabilization, ongoing ocular disease is seen in mps- patients despite successful hct. this hallmarks the shortcomings of current standard therapies. new therapies that overcome the weak spots of current therapies are necessary to improve the late outcomes of these patients. clinical trial registry: n.a. disclosure: b.t.a.v.d.b. was supported by a research grant from the sylvia toth charity foundation, the hague, the netherlands, while working on this study. the sponsors of this study are public or nonprofit organizations that support science in general. they had no role in gathering, analyzing, or interpreting the data. all authors would like to thank all parents and patients for participating in this study. all authors state they have no competitive (financial) interests in this study. background: paroxysmal nocturnal hemoglobinuria (pnh) is an acquired clonal disorder of the hemopoietic stem cells for which the only curative treatment is allogeneic hematopoietic stem cell transplantation. haploidentical donor hematopoietic stem cell transplantation (haplo-hsct) is now increasingly applied as a curative therapy for patients with hematologic diseases. however, there are still few reports on the use of haplo-hsct for the treatment of pnh. our study aimed to compare the outcomes of haplo-hsct with matched-sibling donor transplantation (msd-hsct) for pnh. methods: the clinical data of pnh patients received hsct (haplo-hsct = , msd-hsct = ) in our center from july to may were analyzed retrospectively to compare the outcomes of haplo-hsct group with msd-hsct group. the clinical data including male patients and female patients, classical pnh and pnh-aa syndrome, the median age was years (range - ). all patients had received various treatments before transplantation such as steroids, androgens, cyclosporine (csa), antithymocyte globulin, and growth factors. the median interval from pnh diagnosis to sct was months (range - ). the conditioning regimen was modified bucybased regimen in haplo-hsct group, csa, mycophenolate mofetil (mmf) and short-term methotrexate (mtx) were administered for graft-versus host disease (gvhd) prophylaxis. patients with msd-hsct were treated with the flucy-based regimen and csa were administered for gvhd prophylaxis. results: there were no differences of gender, age, patients of pnh-aa and median time from diagnosis to transplantation between the groups (på . ). the median values of absolute nucleated cell counts were . ( . - . ) × /kg in the haplo-hsct group and . ( . - . ) × /kg in the msd-hsct group (p = . ). the median doses of cd + cells infused were . ( . - . ) × /kg and . ( . - . ) × /kg (p = . ), respectively. all patients attained complete engraftment, no patient occurred graft failure. the median time for myeloid engraftment were (range, - ) days in the haplo-hsct group and (range, - ) days in the msd-hsct group (p = . ). the median time for platelet engraftment were (range, - ) days and (range, - ) days (p = . ), respectively. with a median followup of ( - ) months in the haplo-hsct group and ( - ) months in the msd-hsct group (p = . ). in haplo-hsct and msd-hsct groups the incidences of grade i-iv acute graft-versus-host disease (agvhd) were . % and . % (p = . ), grade ii-iv agvhd were . %、 . % (p = . ). chronic gvhd were . % and . % (p = . ), moderate-severe chronic gvhd were . % and . % (p = . ). in haplo-hsct and msd groups the incidences of infection were . % ( / ) and . % ( / ) (p = . ). no patient occurred early death and relapse. -year estimated overall survival (os) of haplo-hsct and msd-hsct groups were . % ± . % and . % ± . % (p = . ), gvhd-free and failure-free survival (gffs) were . % ± . % and . % ± . % (p = . ). conclusions: the preliminary results indicated that haplo-hsct is a feasible choice for pnh with favorable outcomes, haplo-hsct and msd-hsct had similar therapeutic efficacy. disclosure: no disclosure p pres in bmt for thalassemia major in india: lower incidence and limited impact background: posterior reversible encephalopathy syndrome (pres) is a relatively common complication seen after blood or marrow transplantation (bmt) for hemoglobinopathies with a reported frequency of - %. pres has also been associated with poorer survival rates. severe hemoglobinopathies are one of the most frequent indications for bmt in the developing world, particularly in india. given the risk of rejection in multiply transfused patients and the need to minimize gvhd risk, immunosuppression post-bmt for these non-malignant conditions can be particularly intense and prolonged. we sought to measure the incidence and impact of pres in developing countries. methods: we analysed successive transplants for thalassemia using protocol (atg-bucy+csa/mmf or csa/mtx) maintaining cyclosporine a (csa) blood levels - ng/ml for patients and protocol (flu-atg-bucy+csa/mtx) maintaining higher csa levels post, i.e. - ng/ml for patients from fully matched donors with g-csf-primed bone marrow. for patients this was the second transplant from a different matched related donor. pres was confirmed with brain ct/mri for all patients. results: all recipients who had pres had sibling donors, males and females. age median . (iqr . - years). the frequency of pres was . %; disease free survival for patients who had pres was %. pres resolved completely in all. csa was switched to mmf in patients who had received mtx and were on csa only at the time of pres occurrence, while csa was stopped but mmf continued in patients taking csa/mmf combination and csa was continued for patient. three patients with pres had grade acute gvhd, had grade gvhd and none developed chronic gvhd. csa levels at the time of pres were a median of ng/ml (iqr: to ) with patient having ng/ml. three patients had pres while they were thrombocytopenic. hypertension stage was observed in four patients, stage in one patient, one patient was not hypertensive and in one patient blood pressure values were not available. two patients were on methylprednisolone and . mg/kg/day and one was on dexamethasone mg/m /day. one patient was started on csa again after the pres episode and within weeks had another one while on csa (level ng(ml), methylprednisolone . mg/kg/day and ruxolitinib for gvhd. protocol had statistically significant improvement in disease free survival from % to % (p< . ) with probability of occurrence of pres increasing from . % to . % (p = . , see figure ), yet had a benign course in all patients. conclusions: not stopping immunosuppression may have been the key factor which could explain why we have better outcomes with pres than what is reported. intensifying immunosuppression pre-bmt did lead to more pres, albeit not significantly, and yet it was quite manageable. even with addition of fludarabine our pres incidence is lower than previously reported. [[p image] . background: sickle-cell diseases (scd) are a group of genetic hemoglobin disorders marked by brain vasculopathy. allogeneic hematopoietic stem cell transplantation (hsct) is a curative option able to stop vascular disease progression. diffusion-tensor imaging (dti) is a magnetic resonance imaging (mri) technique sensitive to the brownian motion of water molecules and cellular environment. this microscopic quantitative technique is able to detect white matter (wm) alterations before a conventional mri. the aim of this study was to use dti to evaluate axonal damage and structural connectivity in the brain of patients with scd submitted to hla-identical sibling allogeneic hsct. methods: sixteen scd patients with no extensive vasculopathy detected by conventional mri ( male, age range: - years) and age-matched healthy controls ( male, age range: - years) participated in this prospective study. mri acquisitions were performed in a t scanner two times for patients (before and - years after hsct) and at a single moment for controls. from dti acquisitions, fractional anisotropy (fa), mean (md), radial (rd) and axial diffusibility (ad) were calculated in the wm of the whole brain. structural connectivity was also analyzed, based on graph theory, obtaining efficiency, length path and clustering coefficients of the brain network. an anova test was applied to analyze fa differences among controls and patients, before and after hsct. a paired two-tailed t-test was used to determine statistical significance of changes in the fa, diffusivity mean values and network parameters before and after hsct. results: mean fa was lower in patients before hsct than controls (p = , ) and increased after hsct being not statistically different when compared to controls (controls = , ; patients before hsct = , ; patients after hsct = , ; post hoc dunnett's test -error , ; anova test). when patients were compared before and after hsct, md and rd decrease after hsct (p = , and , , respectively). on the other hand, fa increased (p = , ). after hsct, efficiency was higher (p = , ) and path length index was lower (p= , ) than at study entry (table ) . conclusions: this study indicates that, before hsct, patients with scd present axonal damage not detectable by conventional mri, when compared to healthy controls. we also suggest that hsct is able to promote axonal recovery and reorganization. partial diffusivity recovery could be associate to a still unidentified mechanism of myelin regeneration. in the future, longer follow up and comparisons with other forms of treatment are required. background: bmt is a well-established treatment modality for haemoglobinopathies, limited by the availability of related donors. unrelated transplantation has historically shown variable outcomes driven by gvhd and toxicity, and usually restricted to / matches, but the impact of reduced toxicity conditioning regimens is yet to be known. methods: from to twenty-five consecutive unrelated bone marrow transplants were conditioned with fludarabine mg/m , treosulfan g/m , thiotepa mg/ kg and atg (thymoglobulin) . mg/kg if the source of stem cells was marrow (n = ) or ptcy if pbsc (n = ). endogenous haemopoiesis was suppressed pretransplantation for a minimum of weeks. gvhd prophylaxis was provided with ciclosporin/sirolimus and mmf. thirteen patients were transplanted for b thalassaemia major, one of a thalassaemia major and sickle cell disease. the median age was years ( - ). ten patients were / matched ( thalassemia and sickle) and patients had a / match ( thalassaemia and sickle). the median cell dose was . x tnc/kg (range . - . ) and . x cd +/kg (range . - . ). the median survival was . months ( . - . ). patients with thalassaemia were pesaro class i or ii (pesaro class iii patients were intensively chelated pretransplantation to return to class i or ii). patients with sickle cell disease were transplanted for stroke or recurrent vaso-oclusive crises and/or acute chest syndrome not responding to hydroxycarbamide. results: all patients engrafted and achieved evidence of donor haemopoiesis on day + and achieved transfusionindependence and donor haematological values, but subsequently one / patient with thalassaemia suffered secondary graft failure on day + after macrophage activation syndrome. median neutrophil engraftment was days (range to ) and days ( - ) for / and / patients respectively. patient with sickle cell disease had the platelet count maintained > x /l at all times. the median platelet engraftment > x /l was days (range to ) and days (range to ) / and / patients respectively. there were three deaths, all in the / matched group: two with thalassaemia (day + due to idiopathic pneumonia syndrome and day + due to mas) and one with scd (day + due to ips). there were different trends of complications seen by degree of matching that did not segregate otherwise by disease. conclusions: in conclusion, unrelated bmt for haemoglobinopathies with reduced toxicity regimens is feasible. whilst gvhd caused significant morbidity during the transplant period, other alloreactive/endothelial complications (vod, macrophage activation syndrome, idiopathic pneumonia syndrome) were only seen in the / transplants. disease-free survival, dependent on transplantrelated mortality, and lack of long-term toxicity, including chronic gvhd, are determined by the degree of matching. / matched transplants have excellent long-term outcomes with no chronic gvhd > months and can be considered for patients without a related donor; whereas / transplant have significant toxicity and mortality, warranting a haploidentical approach. disclosure: no conflict. long-term safety and efficacy of lentiglobin gene therapy in patients with transfusion-dependent β-thalassemia following completion of the phase / northstar study patients with transfusion-dependent β-thalassemia (tdt) may benefit from gene therapy involving β-globin gene addition to hematopoietic stem cells (hscs) enabling production of functional hemoglobin (hb). lentiglobin gene therapy contains autologous cd + hscs transduced ex vivo with the bb lentiviral vector encoding β-globin with a t q substitution under transcriptional control of the encoding β-globin locus control region. the safety and efficacy of lentiglobin was evaluated in adults and adolescents with tdt in the -year phase / northstar study (hgb- ; nct ). methods: patients with tdt (≥ ml/kg/year of red blood cells [rbcs] or ≥ rbc transfusions/year) received g-csf and plerixafor for hsc mobilization. to generate drug product (dp), cd + hscs were transduced with the bb lentiviral vector. patients underwent single-agent, myeloablative busulfan conditioning, were infused with the dp, and were followed for safety and efficacy. results: eighteen patients have been treated in the completed northstar study. as of september , patients had a median follow-up of . (min -max: . - . ) months. the median age at consent was (min -max: - ) years including patients ≥ years old. patients received a median cell dose of . (min -max: . - . ) cd + cells x /kg with a median dp vector copy number (vcn) of . (min -max: . - . ) vector copies/ diploid genome. the median liver iron content (lic) at baseline was . (min -max: . - . ) mg fe/g dw. outcomes by age and baseline iron status will be presented. the median time to neutrophil and platelet engraftment was . (min -max: - ) and . (min -max: - ) days, respectively. four patients had platelet engraftment ≥ day and four patients had platelet counts of ≤ x /l at month . none of these patients had ≥ grade bleeding events post-lentiglobin infusion. transfusion independence (ti, defined as weighted average hb ≥ g/dl without rbc transfusions for ≥ months) was achieved in / patients with non-β /β genotypes and / patients with β /β genotypes. in patients who achieved ti, total hb at last visit was . - . g/dl. lic increased from baseline in patients who achieved ti by a median of . % and . % at month and then decreased from baseline by a median of . % and . % at month and , respectively. non-hematologic grade ≥ adverse events post-infusion in ≥ patients included stomatitis, febrile neutropenia, pharyngeal inflammation, and irregular menstruation. there was no transplant-related mortality, vector-mediated replication competent lentivirus, or clonal dominance. two patients experienced grade serious veno-occlusive liver disease (table ) . events resolved following treatment with defibrotide and were attributed to myeloablative conditioning. conclusions: in the northstar study, % of patients with tdt and non-β /β genotypes and % of patients with β / β genotypes achieved transfusion independence. the safety profile of lentiglobin remains consistent with myeloablative busulfan conditioning. longer time to platelet engraftment was observed in some patients, but no graft failure was reported. clinical background: sickle cell disease (scd) is an inherited hemoglobin disorder associated with high morbidity and mortality. currently, allogeneic hematopoietic stem cell transplantation (hsct) is the only curative therapy for scd. transplant outcomes with thiotepa, treosulfan and fludarabine (ttf) preparative regimen are encouraging but this regimen has not been directly compared to other preparative regimens in scd. we therefore planned to compare the event free probability for death, rejection and high grade acute graft versus host disease (agvhd) between ttf and busulfan and fludarabine (bf) regimens. methods: in this retrospectively cohort study, we included all patients with scd who received allogeneic hsct at our center or who were transplanted in other centers and referred to ours for follow up before day . patients were transplanted between july and december . we used kaplan-meier curve to estimate the event free probability for death, rejection and high grade agvhd (grades - ). cox regression was used to assess the impact of the preparative regimen on these outcomes. results: a total of patients were included with a median age of years (interquartile range [iqr]: - ) and a median hemoglobin of g/dl (iqr: - ). sixtytwo percent were males. the proportion of patients who had splenectomy, stroke and acute chest syndrome was %, % and % respectively. all patients received peripherally collected hematopoietic stem cells from a matched sibling donor with a median stem cell dose of x /kg (iqr: . - . ). most patients, %, received cyclosporine or tacrolimus based agvhd prophylaxis. most patients received ttf ( %) or bf ( %) preparative regimens. all patients in the bf group received atg. the median follow-up time was months (range: - ). four patients died during the follow-up period with an os of % ( % confidence interval [ci]: %- %) at years. the os was not different (hr . , p = . ) between the ttf ( %) and the bf ( %) regimens. the probability of high grade agvhd free survival at day was % ( % ci: - ) for all patients. this probability was % in the ttf group and % in the bf group and the difference was not statistically significant (hr . , p = . ). the rejection free survival at months was % ( % ci: - ) for all patients. no patients in the ttf group rejected while the rejection free survival at months for the bf group was %. this was not statistically significant (p = . ). conclusions: in patients with scd undergoing allogeneic hsct from a matched sibling donor, the ttf preparative regimen is not associated with improved os, rejection free or high grade free agvhd survival when compared to the bf preparative regimen. larger studies are needed to confirm these findings. disclosure: nothing to declare. novel strategy for haploidentical hematopoietic stem cell transplant in sickle cell disease methods: consecutive patients suffering from scd who underwent hhsct between jan till date were enrolled in the study. all underwent autologous backup (target dose> x /kg) followed by pre-transplant immune suppression (ptis) cycles at weekly intervals using fludarabine @ mg/m /day(d -d ) + cyclophosphami-de@ mg/m /day(d ) + dexamethasone@ mg/m / day(d -d ) along with hypertransfusion (target hb - gm/dl), hydroxyurea ( mg/kg/day) and azathioprine ( mg/kg/day) from day - . the graft was mobilized using gcsf@ mcg/kg/day(d -d ) + plerixafor@ . mg/kg s/ c on d - hours before the pbsch. conditioning included thiotepa mg/kg in two divided doses (d- ), fludarabine mg/m (d- to d- ), cyclophosphamide . mg/kg (d- , d- ), tbi gy with thymic shielding (d- ), ratg (genzyme thymoglobulin . mg/kg (d- to d- ). gvhd prophylaxis included ptcy mg/kg/day on d and , sirolimus (target levels - ng/ml) (till - months post hsct) and mmf (till d ) starting from d . results: the median age of patient's was years (range - years). before transplantation all patients had repeated episodes of one or other complication warranting a transplant, non-responsive to hydroxyurea. six had maternal donors, paternal and sibling. median age of the donor was years (range - years). all were dsa negative with a cutoff mfi of > iu. all patients received x /kg cd cells irrespective of harvested dose which ranged from ( . - . x /kg). median cd dose was . x /kg (range . - . x /kg). all patients engrafted with median time to neutrophil engraftment days (range - days) and median time to platelet engraftment days (range - days). median duration of hospital stay was days (range - days). one patient had cytokine release syndrome needing tocilizumab. five had engraftment syndrome treated with short course of steroids. two had cmv reactivation needing treatment with ganciclovir/valganciclovir. acute gvhd grade ii was seen in one patient. till date of analysis none had features compatible with chronic gvhd. of the patients, are alive without sickle cell disease with lansky/ karnofsky scores of . at median follow up of days (range - ) the probabilities of survival, sca-free survival, and transplant-related mortality after transplant were . %, . %, and . %, respectively. one patient died due to mdr klebsiella sepsis after being discharged initially while he was receiving iv ganciclovir on day care basis. he had full donor chimerism. none of the patient had primary or secondary graft failure. conclusions: pre-transplant immune suppression and upfront use of plerixafor for graft mobilization decreases the risk of graft failure and graft versus host disease leading to overall better survival in hhsct for sickle cell disease. disclosure: none. combined haematopoietic stem cell transplant and enzyme replacement therapy in wolman disease: outcomes and challenges jane kinsella , denise bonney , helen campbell , robert wynn , simon jones background: infantile lysosomal acid lipase deficiencymore commonly known as wolman disease -is an autosomal recessive lysosomal storage disease, characterised by storage of cholesterol esters in the liver, spleen and gastrointestinal tract. these children present under the age of months and traditionally had a poor prognosis, with almost all being dead by the age of months. bone marrow transplant has been used to correct disease manifestations, but limited by high procedure-related mortality with the significant co-morbidities. the survival has changed over the past few years due to pharmacological enzyme replacement therapy but still presents challenges for these patients and their clinicians. in these children haematopoietic stem cell transplant we have offered bmt with enzyme replacement therapy, in certain specific circumstances. methods: four children with wolman disease being treated with enzyme replacement therapy, limited by alloantibody, or poor venous access, received treosulfan-based, myeloablative conditioning with serotherapy followed by a matched haematopoietic stem cell transplant: two family donors, one sibling donor and one unrelated donor. results: three of the four children survived transplant. they have continued to receive enzyme replacement therapy but at reduced dose and frequency with improved tolerability. they have continues to grow and develop. growth and gastrointestinal histology is improved for children having received transplant compared to those receiving enzyme replacement alone. monitoring of peripheral blood chimerism has shown a disease-associated engraftment defect, with mixed chimerism in the surviving patient. conclusions: haemopoietic stem cell transplant is a suitable treatment option in children with wolman disease in whom receiving enzyme replacement therapy is not possible because of venous access, sensitisation or cost reasons. it improves their tolerability of the enzyme treatment and allows for a reduction in enzyme dose and frequency. however, the results of engraftment are not as good as expected for a transplant with myeloablative conditioning and a matched donor. an engraftment defect has been observed in lysosomal acid lipase deficient animal models. a further understanding of this poor engraftment in children with wolman disease is required as to determine whether the risks of transplant is beneficial in these patients and for the consideration of future treatment options including gene therapy. background: thalassemia major is the most common transfusion dependent hemolytic anemia in the world. the absent or reduced production of the β-chain of hemoglobin causes severe ineffective erythropoiesis, massive erythroid hyperplasia in the bone marrow and extramedullary hematopoesis occurs. patients require regular transfusion therapy lifelong. currently, the only proven curative treatment of thalassemia is allogeneic stem cell transplantation (sct). methods: we evaluated the immune reconstitution results of patients at year after hematopoetic stem cell transplantation at our pediatric bone marrow transplantation center between january and december . all patients were not receiving any immunosuppressive treatment at least for months and they have normal lymphocyte counts, immunoglobulin levels and transfusion independent. lymphocyte subtypes and chimerism percentages and the relationship with the donor type were evaluated at year of transplantation. results: ages of transplantation was ranged between - years (median: years). seven ( %) of them was male. matched unrelated donor type was chosen in patients while others ( patients) were transplanted from family matched donor (matched sibling: patients, matched family: patients). all patients received myeloablative conditioning regimen containing busulfan/treosulfan, cyclophosphamide, thiotepa and fludarabine. follow up time was between - months (mean: ± months). in patients, whole bone marrow product was used while peripheral stem cell harvest in remaining patients. cd levels were found low in only patients, in normal patients mean was % ± %. cd levels were severely low in patients while cd in only patient. cd levels were increased in total patients in as compensatory. cd /cd ratios were very low in all patients (range: . - . ). b cells (cd +) were low in patients while immunoglobulin levels were normal. chimerism values between - % (mean: ± %). donor and product types did not differ in cd + lymphocyte reconstitution at year (p= . , p= respectively). all patients were alive and well at year after transplantation. conclusions: after year of transplantation, although patients are in well condition regarding to infection frequency and transfussion dependency, it was seen that their lymphocyte subtypes reconstitution could not be achieved enough as in normal children. we can conclude that low cd + cell levels were an expected finding in almost all patients. so, these patients may have a tendency to suffer serious bacterial and viral infections, and close follow up be required in terms of infections as long as cd levels continue to be low. immunoglobulin replacement therapy did not required even in patients with low b cell levels. disclosure: nothing to declare p phase international, multicentre trial to assess haploidentical aß t-cell depleted stem cell transplantation in patients with sickle cell disease with no available sibling donor background: sickle cell disease (scd) is an inherited disorder with an estimate of , affected newborns per year worldwide. allogeneic hematopoietic stem cell transplantation (hsct) with a matched sibling donor (msd) is currently the curative standard of care for scd patients (pts). however, msd availability is < %. a t-cell depleted haploidentical hsct (t-haplo-hsct) from a relative, mostly a parent, expands the donor availability while exhibiting low gvhd rates and thus could offer cure to the remaining % of scd patients. in a pilot study, comparing t-haplo-hsct with msd hsct in advanced stage scd, using almost identical transplant regimens for both. the overall (os) and disease-free survival (dfs) was % vs. %, respectively. methods: these results led to the design of a clinical trial to assess tcd-haplo-hsct prospectively which aims to demonstrate that a hsct from a haploidentical relative is not inferior to a msd hsct with regard to major outcome parameter. this phase , prospective, stratified, open-label study is targeting enrollment of patients aged - years with homozygous hbs disease or heterozygous hbsc or hbs /+ ß-thal suffering from severe or moderate scd related complications. inclusion criteria are clinically significant scd related complications such as stroke, silent crisis, pathological angio-mri, transcranial doppler (tcd) velocity > cm/s, or more episodes of acute chest syndrome (acs) in a lifetime, chronic transfusion dependency, transfusion-refractory allo-immunization and others. pts fulfilling inclusion criteria will be stratified according to donor availability. pts with a msd will receive a bone marrow graft, pts requiring an alternative donor will be transplanted with an aß/cd depleted graft from a haploidentical family donor. the conditioning regimen for both groups will be identical with the exception that antithymoglobulin (atg-neovii ® ) is given upfront in thaplo-hsct versus day - to - in msd. chemotherapy consists of thiotepa, fludarabine and treosulfan. posttransplant immunosuppression will consist of mofetil mycophenolate and tacrolimus for a duration > months in t-haplo-hsct and < months in msd, depending on chimerism. (eudract number: - - ) results: primary efficacy endpoint: event free survival (efs). event is defined as incidence of acute gvhd, grade iii -iv, chronic gvhd, rejection (graft failure) or death (for any reason). key secondary endpoint(s) are os, dfs, graft failure, hematological and immunological reconstitution, quality of life (qol) assessment and fertility. the primary null hypothesis is: efs of scd patients treated with t-haplo-hsct is non-relevantly inferior to efs in the msd arm. conclusions: results will help to determine if an a/ß depleted t-haplo-hsct can be considered equivalent to msd hsct with regard to dfs, adverse events and safety, in order to offer this form of cure to the majority of patients with scd. disclosure: nothing to declare hit three birds with one stone: successful stem cell transplantation from one family donor to three siblings methods: in august , three thalassemic siblings were admitted to hospital for stem cell transplantation from a full match donor, their years old sister. the patients' general health conditions and specific health issues due to thalassemia were checked extensively. it was decided to perform first transplant to older sister whom the disease and transplant complications are expected more intense due to prolonged transfusion and chelation therapy. the oldest daughter of family, healthy, was planned to accompany her sisters in transplantation unit so parents can take care the others and organize this period for whole family. results: the years old sibling was first admitted to bone marrow transplantation unit in july . the conditioning regimen was busulfan, fludarabine, cyclophosphamide and thiotepa with antithymocyteglobulin(atg) and defibrotide prophylaxis was given. the healthy donor was admitted to hospital and received g-csf for continuous days before harvesting. stem cells were collected peripherally on day and viable cd + cells were /ul. patient received , x e /kg stem cell and the other cell products were divided into parts according to other recipients´weight. no infusion problems were recorded in stem cell transfusion. gvhd prophylaxis was given with cyclosporin and methotrexate. severe sinusoidal obstruction syndrome was observed and successfully managed with supportive therapy. neutrophils were engrafted + . day, and platelets were on day . full blood chimerism results were % in day , % in day and % in day consecutively. after months from first transplant the years old sister was admitted to hospital on october . same conditioning with defibrotide prophylaxis and gvhd prophylaxis were given and , x e /kg peripherally derived and previously frost stem cell was infused without any complications. mild sinusoidal obstruction syndrome was observed and managed with supportive therapy successfully. neutrophils were engrafted + . day, and platelets were on day . full blood chimerism results were % in day , % in day and % in day consecutively. the third transplant was performed on january with the same conditioning and prophylaxis regimen. although defibrotide was used mild sos was observed and treated with supportive therapy with success. neutrophils were engrafted + . day, and platelets were on day . full blood chimerism results were % in day , % in day and % in day consecutively. conclusions: the patients are being followed for over a year after first transplat, neither adverse nor gvhd symptoms were observed. we presented this case for being a unique example for match family donor transplant and the first successful example from one donor to three recipients. disclosure: nothing to declare results: our female patient admitted for anemia at rd month of birth and was transfused every - months from th months to . years of age. since investigations directed towards hemoglobinopathies or membrane defects like hereditary spherocytosis were unremarkable, she was not transfused for years after the age of . -years because hemoglobin level was constant over g/dl. her bm examination showed erythroid hyperplasia and feature of dyserythropoiesis with a few binucleated erythroblasts. it was decided to follow-up the patient with a diagnosis of cda ii. after the age of -years, the need for transfusion started again for every to months which led the parents of our patient to request for bone marrow transplantation, however, the diagnosis was not definite, and because of the insufficient data for the transplantations for cda ii patients, it was decided to go on to follow-up. nevertheless, after years, the frequency of transfusion gradually increased to every - weeks, and bone marrow transplantation was brought into question again. at that time, genetic examination was started and sec b gene was analyzed by direct sequencing. hsct decision from her hla / matched brother, carrying sec b mutation in heterozygous state, was taken. in the preparation regimen, busulfan (bu) at a myeloablative weight adjusted dose ( days), mg/kg cyclophosphamide (cy) ( days), and mg/kg antithymocyte globulin (atg fresenius) were used. graftversus-host disease (gvhd) prophylaxis was with cyclosporin a started on day - and short-term methotrexate on day + ,+ and + . she was transplanted with bm with a dose of total nucleated cells= . x /kg and cd = x / kg. neutrophile and platelet engraftment were achieved at + and + , respectively. indeed, grade hemorrhagic cystitis due to bk virus and a moderate veno-occlusive disease prolonged platelet transfusion days which concealed the exact engraftment day of platelet. the patient was discharged on day with no more need for any transfusion and followed up as a complete chimeric with no type of gvhd since then. now, she is years old, under regular surveillance at our transplant centre without any symptoms. conclusions: hsct data in cda ii patients is still insufficient, however based on data from tm patients with similar treatment approaches in td cda ii patients, it is seen that the hsct is reliable and effective. disclosure: nothing to declare hematopoietic stem cells background: the prognosis after frontline therapy in b-all patients have improved due to monoclonal antibodies (cd , cd , cd ) and approximately % of patients achieve complete remission. in relapsed and refractory (r/ r) b-all and also in mrd + outcomes are relatively poor. disease-free survival (dfs) in this cohort is - %. in this cohort allo-hsct is indicated and complete remission before transplantation is crucial for prognosis. conventional chemotherapy is associated with high failure rate and significant toxicity. immunotherapy with monoclonal antibodies and car-t are more promising approaches. the aim was to evaluate the efficacy (frequency of responses, os, dfs) and toxicity, especially neurotoxicity and cytokinerelease syndrome, of a bispecific monoclonal antibody blinatumomab in patients both children and adults with persistence of minimal residual disease (mrd + ) or r/r b-all as a bridge to allo-hsct. methods: this study included patients with high risk b-all blinatumomab treated in - , among them pts ( %) with t( ; ), ( %) with t( ; ), with mll ( %), pts ( %) who were refractory to previous chemotherapy, ( %) after allo-hsct from deferent type of donors. median age was y.o. (range m- y.o), children - y.o. ( %) and adults > y.o. ( %). r/r all had pts ( %), mrd + - pts ( %), median days of follow up were ( - ). blinatumomab was applied as -day cycles followed by a -day off-period before the start of the following cycle. majority pts received one cycle (n= , %). in r/r all group dose was of mcg/d during the first days and afterwards mcg/d. patients with weight less than kg received mcg/m /d and mkg/m /d accordingly. in mrd group dose was mcg/m /d. results: the frequency of responses to blinatumomab was higher in mrd + pts in comparison r/r all pts ( % vs % p= . ). in mrd + pts cr mrdwas achieved in pts ( . %), pts ( . %) were mrd+ after blinatumomab. two-year os in this group was %. twenty pts ( %) received allo-hsct. in rr all pts cr mrdwas achieved in pts ( %), pts ( %) were mrd+ after blinatumomab, pts ( %) had no hematological response . two-year os in r/r all was %. fifteen pts ( %) received allo-hsct. os in cr mrdpatients who received allo-hsct was not significantly different in comparison with patients who received blinatumomab as a monotherapy ( % vs %, p= . ). no significant differences in dfs were observed at two years in cr mrdpts depending status of the disease before therapy-mrd vs r/r ( % vs %). of the reported adverse events, febrile fever was the most common pts ( %), neutropenia ( %), thrombocytopenia ( %), infection ( %), neurotoxicity ( %), cytokine-release syndrome ( %). all complications were reversible. conclusions: blinatumomab is effective option in patients with high risk b-all especially in the group with mrd persistence after previous chemotherapy and facilitates effective bridging to hsct. blinatumomab therapy is generally well tolerated. disclosure here we address the transcriptional regulation of differentiated cells from human embryonic stem cells (escs) using self-assembling peptide hydrogel without stromal cells, and compare with embryoid body (eb) culture system. methods: esc differentiation was induced in eb culture system or three-dimensional ( d) hydrogel culture system. the engraftment potential of differentiated cells was evaluated by flow cytometry. cd + cells from mobilized peripheral mononuclear blood cells (mpbmcs) or differentiated from escs at different times (day , day , day ) were purified by fluorescent-activated cell sorting. sorted cells were captured on medium-sized microfluidic chips using the fluidigm c single cell auto prep system. sequencing was performed by hiseq x ten. results: self-assembling peptide hydrogel formed a d scaffold for cell culture, the pore diameter of which ranged from to nm. compared to eb culture system, escs in d culture system differentiated more potently. the differentiated cells from d system were short-term engrafted in the nog mice, and myeloid cells, b cells and t cells could all be detected in peripheral blood after transplantation. however, the engraftment was not obtained in differentiated cells from eb culture system. we obtained and analyzed escs, cd + cells from eb culture system, cd + cells from d culture system, and cd + cells from mpbmcs. the cells were divided into cluters ( figure a ). in both differentiation systems, the cd + cells from day were more heterogeneous than cd + cells from day and day ( figure b) . however, cd + cells from mpbmcs were more homogeneous, probably because the differentiated cd + cells contained several cell lineages, including hematopoietic cells, endothelial cells and mesenchymal cells. there is transcriptional overlap between individual cd + cells from eb and d culture systems. however, we found that cluster , which is composed mainly of cd + cells from d at day and day , expressed similar level of several hematopoietic regulator as hsc, such as tal , lmo , erg ( figure c ). the cluster , which is almost the cd + cells from d at day , also expressed the highest gata among the clusters from differentiated cells ( figure c) . conclusions: our study demonstrates that d hydrogel culture system facilitates hematopoietic specification of escs. disclosure: nothing to declare higher cd + cell dose increases overall survival in the setting of dual t-lymphocyte suppression with atg and ptcy in matched related and unrelated donor allosct background: there is no consensus on the cd + donor cell numbers required for optimal outcomes in allogeneic stem cell transplant (allosct). there is controversy on the benefits or harm in higher cell dose for allosct. this study aims to evaluate the impact of cd + cell dose in allosct patients receiving reduced intensity conditioning (ric) combined with anti-thymoglobulin (atg) and posttransplant cyclophosphamide (ptcy) using related (mrd) and / and / matched unrelated donors (mud). methods: this is a single-centre retrospective analysis of adult patients who received allosct for hematologic malignancies between october and may . all received ric using fludarabine ( mg/m /day: day - to - ), busulfan ( . kg/m /day: day - and - ) and total body irradiation ( cgy: day - ). all patients also received rabbit-atg ( . mg/kg: day - to - ), ptcy ( mg/kg/day: day + ,+ ) and cyclosporine (from day + ). unmanipulated peripheral blood stem cells were infused on day . analyses were done using thresholds: ( ) an arbitrary cd + cell dose of x /kg (as this was our target dose) and ( ) cell dose according to quartiles (< . , . - . , . - . and ≥ . x /kg). results: median cd + cell dose was . x /kg. median follow up was months (range - ). median neutrophil engraftment was (range - ) days and platelet engraftment was (range - ) days. a cell dose greater than x /kg was associated with an increased overall survival (os) at year ( . %; % ci, . - . vs . %; % ci . - . ; p= . , figure ). the higher dose was also associated with shorter platelet engraftment time (p= . , figure ). there was no significant difference in neutrophil engraftment, nonrelapse mortality (nrm), relapse free survival (rfs), grade ii-iv acute graft versus host disease (agvhd) and moderate to severe chronic graft versus host disease (cgvhd), (table ) . analyses using quartile cell dose thresholds showed a trend towards decreased os with a cell dose of < . x ^ /kg, however this was not statistically significant ( figure ). higher cd + cell doses were associated with shorter platelet engraftment time (p= . , figure ). there was no significant difference in neutrophil engraftment, nrm, rfs, agvhd and cgvhd (table ) . conclusions: cd + cell dose greater than x /kg significantly increases overall survival in the setting of ric and dual t-lymphocyte suppression with atg and ptcy in mrd and mud allohsct. further studies in a larger number of patients and longer follow up are recommended to validate these findings. disclosure methods: fifty two adult patients were included. median cd + cells requested for infusion were x ^ /kg. all patients received the same ric regimen including fludarabine ( mg/m /day day - to - ), busulfan ( . kg/m /day day - and - ), and total body irradiation ( cgy) (day - ) combined with rabbit-atg ( . mg/kg: day - to - ), ptcy ( mg/kg/day: day + ,+ ), and cyclosporine. unmanipulated peripheral blood stem cells were infused. last followup was november . median follow-up was months (range - ). median cell dose count infused was . cd +/kg. we arbitrarily divided the cohort in two groups with cd + dose of > x ^ cd /kg as cut-off point. results: findings are summarized in figure . the infusion of more than x ^ cd /kg dose had a significant worse impact on overall survival (os) (p= . ), relapse-free survival (rfs) (p= . ) and cumulative incidence of acute gvhd (p= . ). chronic gvhd could not be compared between the two cohorts due to the different median follow-up. conclusions: the infusion of a cd + cell dose count higher than x ^ cells/kg had a significant adverse impact in overall survival and grade ii-iv acute gvhd in the setting of ric and dual t-lymphocyte suppression with atg and ptcy for haplohsct. disclosure: nothing to declare p long-term thymic activity and immune-reconstitution after haplo-identical allografting with post-transplant cyclophosphamide background: the use of post-transplant cyclophosphamide (ptcy) has expanded the application of t repleted haploidentical stem cell transplantation (haplo-hsct). in this setting, to investigate thymus role in longterm clinical outcomes, evaluation of immune reconstitution kinetics was performed. methods: twenty-nine patients (median age ) were enrolled. blood samples were collected before conditioning and at , , , , , months after haplo-hsct. analyses of cd and cd t-cell subsets by flow-cytometry were correlated by generalized linear models with real-time pcr (rt-pcr) quantification of signal joint t-cell receptor excision dna circles (sjtrecs), specific marker of naive t-cells thymopoiesis. a) naive; b) central; c) memory; and d) revertant cd and cd t-cells were defined as follows: a) cd ra+cd l+; b) cd ro +cd l+; c) cd ro+cd -; and d) cd ra+/ ro +, respectively. sjtrecs rt.pcr was performed on genomic dna ( ng) extracted from sorted cd and cd t-cells. results: a gradual increase in absolute numbers of all cd and cd t cell subsets and of sjtrecs copies from the first month up to years post-transplant was observed ( figure ) . however, at years, cd and cd t-cell levels and sjtrecs levels were lower than those observed in healthy donors. sjtrecs kinetics was associated with the increase in cd naive t-cells (overall, p < . ). this correlation suggests that most of cd naive t-cells derives from thymic re-education of donor precursor stem cells, whereas cd naive t-cells undergo peripheral expansion after thymic production. furthermore, an increase in cd revertant memory t-cells was also significantly correlated with sjtrecs kinetic (p , ). central and effector memory t-cells showed a faster thymic-independent expansion in both cd and cd tcells. interestingly, sjtrecs levels and thymic dependent immune-reconstitution were higher in a cohort of patients undergoing hsct from hla identical donors (manuscript in preparation). clinical outcomes and thymic function were correlated starting at months after hsct. lower thymic output was significantly associated by multivariate analysis with low pre-transplant trecs values (p , and p < , in cd and cd , respectively), moderate-severe chronic graft-versus-host disease (gvhd; p < , in cd ), and age (≥ years, p , in cd ). conclusions: the thymus, despite age-dependent involution, substantially contributes to t-cell reconstitution after haplo-hsct. chronic gvhd and older age were significantly correlated with reduced thymic function. overall, lower production of sjtrecs after haplo-hsct as compared after hla identical sibling hsct may partly be due to a higher degree of "mismatching" of mhc molecules during thymic re-education. [[p image] . figure ] background: the use of allogenic hematopoietic stem cell transplantation (hsct) in the treatment of adolescents and young adults (aya) with philadelphia negative all is decreasing with the adoption of pediatric inspired protocols to treat this age group and the incorporation of minimal residual disease assessment in the routine care of all patients. previously, its use was defined mainly by disease risk features at presentation. methods: a study on aya (age - years), who underwent allogenic hsct at our institute for philadelphia negative all, between february and december . all the studied patients received calgb based adult chemotherapy protocol for induction, and underwent a matched related donor (mrd) transplant with cy/tbi conditioning and mtx/csa as gvhd prophylaxis. the patients were eligible for allogeneic hsct, if they have a mrd plus one or more of the following risk factors: ( ) age ˃ years, ( ) high presenting wbc count (> for b-all, > for t-all), ( ) high risk immuno-phenotyping (pro-b, pro-t, early t, and mature t), ( ) bulky splenomegaly or bulky lymphadenopathy, ( ) high risk cytogenetics ( ; , ; , low hypodiploidy/near triploidy, complex), ( ) cns involvement, ( ) relapsed or refractory disease at d of induction. in this study, we investigated the impact of those different risk factors on the long term outcome of allogeneic hsct. results: the median os of our studied patients was not reached at . years, with a median dfs of . years (figure ). in a univariate analysis, relapsed or refractory disease prior to transplant was the only independent risk factor for os and dfs (p-value= . , and . respectively) (figure ). in addition, patients who had or more risk factors ( , . %) prior to transplant had a significantly lower long term outcome compared to patients, who had one ( , . %) or two risk factors ( , . %) with a median os of months, and a median dfs of only months (p-value= . , and . respectively) ( figure ) . conclusions: our results show that the long term outcomes of hsct in aya with philadelphia negative all treated on an adult type chemotherapy regimen, were significantly better in patients who showed a good response to initial therapy and a limited poor prognostic factors at presentation, with worsening of dfs as the number of poor prognostic features increase. we can conclude that, using this risk score can be helpful in predicting the outcome of allogenic hsct in aya with philadelphia negative all treated with adult type chemotherapy protocol. disclosure: no conflict of interest a prospective single center survey on donor-specific anti-hla antibodies and desensitization strategy in patients undergoing an allogeneic stem cell transplant background: in the setting of hematopoietic stem cell transplantation (hsct), considering the risk of poor engraftment or graft failure (gf), the detection of antibodies (ab) directed against donor specific hla loci (dsa) represents a contraindication to proceed with the same donor, suggesting the search of other donors. in many cases, there is not sufficient time to search for alternative donors and it is necessary to plan an immunosuppressive strategy to decrease the dsa level, thus reducing the risk of gf. to date, there is no consensus on desensitization standards to manage dsas in hsct. the aim of this study was to determine the incidence of anti-hla ab and dsas in hematologic patients candidate to an allogeneic hsct, and the efficacy of our desensitization protocol. here, we present an update of the results obtained with our strategy. methods: between august and september , we prospectively screened for dsa consecutive patients candidates to an allogeneic hsct. anti-hla ab research was carried out using the luminex bead assay (lifecode screen and lsa i/ii-immucor). the results were expressed as mean fluorescence intensity (mfi); mfi > was considered positive. in case of a mismatched related donor, a flow cytometric crossmatch test (fcxm) was performed. if the patient had dsas and only one available donor, a desensitization strategy was employed, scheduled with rituximab on day - , single-volume plasmapheresis procedures (pp), usually on day - and - , intravenous immunoglobulins on day - , infusion of hla selected platelets for dsa absorption in case of persistent antibodies directed against class i hla antigens. the aim of this schedule was to avoid interferences with chemotherapy and anti-t-cell globulins, infused during condition regimen results: since august , patients have been prospectively screened. thirty-three patients ( . %) showed anti-hla ab and of them ( . %) had dsas: were treated with the desensitization strategy, applied according to the mfi score and the fcxm result, and all of them obtained an engraftment; in cases, an alternative donor was selected and in case the research for an alternative donor is still underway. dsa detection was performed every days after hsct for the first month and , and days following hsct. neither a dsa rebound nor other complications were observed during the follow-up. conclusions: our prospective analysis underlines the high frequency of anti-hla antibodies detection in hematologic patients, confirming the necessity to routinely evaluate the presence of dsas before an allogeneic mismatched hsct. our desensitization schedules based on the combination of pp, rituximab, ivig and platelet absorption proved successful in reducing dsas. we confirm the necessity of a prospective multicenter collaboration to better define the role of dsas against each hla locus and the critical mfi cut-off level associated with a higher risk of gf. transplant and transfusion specialists should joint to define a consensus for a standard desensitization strategy. disclosure the most frequent technique used for counterbalance partial incompatible hsct is cd + selection that is associated with sustained engraftment and effective reduction of t cells that minimizes gvhd. on the other hand, this approach could delay immune reconstitution and increase risk of viral and fungal infection. in mud setting the use of pbsc is the procedure that most centers have recently adopted. this implies the infusion of a relevant higher number of t cells to times more as compared with bone marrow (bm). since in our centre most part of our patients are primary immunodeficiencies, we applied a procedure to minimize the risk of severe gvhd infusing a controlled number of cd positive cells. methods: we report data about paediatric patients who received mud hsct ( patient received hsct) between and in the bmt unit of the children's hospital of brescia. patients received conditioning, according to the european group for bone marrow transplantation (ebmt) and the european society for immunodeficiencies (esid) guidelines. cd + selection has been realized by a milteny column with an ideal addback of cd positive cells of x /kg. stem cell source was bm in cases and pbsc in cases. results: median patients age at transplant was years (range . months- years). the mean number of infused cells were: x /kg cd + and x /kg cd + in bm product, while x /kg cd + and x /kg cd + in pbsc. mean time for engraftment was day post-hsct. as concerns acute gvhd overall incidence . % ( / ) of the children presented this complication, but only % ( / ) presented gvhd grade iii and none gvhd grade iv, while chronic gvhd presented in . % ( limited, extensive/ ). while acute gvhd incidence and severity weren't significantly different between bm recipients and pbsc recipients, the cases of chronic gvhd were prevalently in the latters. no major infections presented in the post-transplant period and immunological reconstitution both cellular and humoral was completed by months. overall survival at years is % ( / ). the results obtained show how it is possible control severity of gvhd if an addback of a controlled number of cd + lymphocytes. acute gvhd wasn't severe and only few children presented with limited chronic gvhd. the method allows to graft primary immunodeficiencies patients even with pbsc without infusing too many t cells. in fact, especially in very young children, the number could be excessive and risky. nevertheless in case of an oncohaematological patient, gvl effect is preserved. disclosure background: dc is a rare genetic disorder that results from a defective telomere length maintenance and is characterized by mucocutaneous features, bone marrow failure (bmf) and a high predisposition to cancer and pulmonary fibrosis. bmf remains the major cause of mortality and the hsct is the only definitive treatment to restore hematopoiesis but is limited by a high incidence of treatmentrelated mortality. methods: a retrospective analysis of patients (pts) with dc who underwent hsct at the bone marrow transplantation unit in the clinical hospital of federal university of paraná, brazil, between july- and november- . results: boys and girls, with a median age of y ( - y) received a hsct from a mds (n= ), mud (n= ) or mmrd (haploidentical, n= ). pts received bone marrow (bm) and pt received a cord blood unit (cbu). the median of tnc infused was , x /kg (range , - , x /kg) and in the cbu was , x / kg. two pts received a myeloablative preparatory regimen with busulfan (bu) mg/kg + cyclophosphamide (cy) mg/kg or fludarabine (flu) mg/m + antithymocyte globulin (atg). the remaining pts received a ric regimen with cy mg/kg (n= ), flu mg/m + cy mg/kg + atg mg/kg (n= ), and flu mg/m + cy + tbi rads (n= , haplo). graft versus host disease (gvhd) prophylaxis consisted of cyclosporin (csa) and methotrexate or steroids (cbu) and post-transplant cy + csa + mycophenolate mofetil in the haploidentical transplants. of evaluable pts engrafted with a median time to neutrophil recovery of days (range: - days). one patient experienced primary graft failure (haplo) while second graft failure occurred in other pts. all these pts went a second hsct and survived. acute gvhd grade ii-iv occurred in of pts at risk. moderate to severe chronic gvhd occurred in pts with cases occurring in pts who had previously presented acute gvhd. overall survival (os) was , % at a median follow-up of y. the y os was slightly better in msd transplants compared to the others ( , % x , % p= , ). causes of early death include adenovirus sepsis (n= ), toxicity to preparatory regimen and sepsis (n= ), primary graft failure (n= ). pts remain alive between - y after hsct with a median fu of y. among them only pt has developed organ involvement by the underlying disease: hepatopulmonary syndrome (hps). pts died due to pulmonary fibrosis (n= ), liver fibrosis(n= ), gi bleeding(n= ), hps (n= ); cgvhd and sepsis(n= ), infection (n= ), and pts were lost to fu. conclusions: early mortality from bmf can be reduced by hsct, but late outcomes remain a consequence of the underlying disease. long term fu is essential in order to detect late complications related to the hsct procedure or the underlying disease. disclosure: nothing to declare single intra-bone cord-blood transplantation with a treosulfan-based regimen, atg-free and sirolimusbased gvhd prophylaxis: fast hematopoietic engraftment and immune-reconstitution in patients background: cord blood transplants (cbt) require less stringent hla-matching, compared to peripheral blood stem cell or bone marrow. however, cbt has been associated with delayed engraftment and immune reconstitution, especially if in vivo t-cell depletion, such as antithymoglobulin (atg), is used. methods: from to , patients with high-risk diseases received intra-bone infusion of unwashed single cb unit with an atg-free gvhd prophylaxis; were in active disease at cbt and had received prior allogeneic stem cell transplantation. median age was y [range (r) . conditioning regimen was myeloablative, with treosulfan and fludarabine in all, intensified with melphalan in or with gy tbi in . hla matches was / , / , / in , and cases, respectively. gvhd prophylaxis included sirolimus and mycophenolic acid (mmf). results: after thawing, median cd + cells was . x /kg [r . - . ], median cd + cells . x /kg [r . - . ], and median cd + cells . x /kg [r . - . ]. of the evaluable patients all engrafted with a sustained full donor chimerism at day . median time to neutrophils ( / , anc> /μl for consecutive days) and platelet engraftment ( / immune-reconstitution of cbt patients (tables a-b ) was compared with two cohorts of patients transplanted at our center from any adult donor with ( ) or without ( patients, including post-transplant cyclophosphamide cohort) atg in association with sirolimus and mmf. profiles of immune-reconstitution at day - - showed a better cd + recovery at any time-point in both cbt and no-atg versus atg cohort, with no statistic significant difference in the first cohorts. moreover, cd +/cd + ratio at any time point was better in the cbt cohort vs the no-atg cohort. b cell recovery was faster in the cbt cohort; immunoglobulin recovery was superimposable across different platforms. focusing on late events (> days from cbt), / pts experienced ebv reactivation, median time days [ - ] treated with rituximab, and experienced late hhv and cmv reactivation, both solved at last visit. sirolimus was withdrawn after a median of days [r - ]. only patient developed severe chronic gvhd, solved at last visit. overall, after a median follow-up of days [r - ], pts are alive and well. conclusions: our data confirm that intra-bone cbt without in-vivo t-cell depletion is associated with fast hematopoietic engraftment and immune-reconstitution, with very low rate of chronic gvhd and late infective events. background: a promising improvement of hematopoietic stem cell transplantation (hsct) may lie in the transplantation of high numbers of pluripotent stem cells to minimize the time span between transplantation and immunological reconstitution. hence, an ex vivo platform is needed that supports hsc proliferation before application and, at the same time, the maintenance of pluripotency by diminishing hsc differentiation into lineage-specific progenitor cells. methods: to artificially model the natural hsc niche in vitro, we used d bone marrow (bm)-like scaffolds made of polydimethylsiloxane (pdms). these structures are based on a human long bone cross section as a representative of the bm. human cryoconserved hscs were cultured in distinct cultivation systems for days under different conditions. cell counting and facs analyses at day were conducted. for characterization of the cultivated hscs, we used antibodies against cd alone or in combination with antibodies against cd , cd , cd ra and cd f. results: for optimization of culture conditions for human hscs, a commercially available medium was supplemented with a panel of cytokines and valproic acid. we found a significant increase in the number of cd + hscs by simultaneously increasing their vitality using the d system compared with conventional d culturing. a further improvement was achieved by introducing a silicon oxidecovering of the d pdms structures, suggesting that hydrophilic surface properties offer superior attachment for semi-adherent hscs. for a more precise characterization of the cultivated hscs, we introduced a panel of facs markers reflecting the immaturity of the amplified hsc. surprisingly, with increasing immaturity of the cultivated hsc, non-covered d pdms revealed to be best suited for amplification: cell number of vital immature hscs was increased after cultivation on non-covered d pdms compared with silicon oxide-covered d pdms and the d system. conclusions: by establishing a d scaffold according to the human bm, we found a platform mimicking the natural niche of human hsc which is suitable to amplify human hscs in vitro and support their vitality, pluripotency and ability for self-renewal. [[p image] . with the introduction of sion covering of d pdms structures the maintenance of cd + hscs could be further improved. despite the better conservation of cd + hscs by using silicon oxide-covered d pdms, we found that immature human hscs obviously prefer more hydrophobic conditions found on non-covered d pdms. disclosure: nothing to declare. abstract already published. improvements in neutrophil engraftment following changes in freezing method background: in the setting of autologous haematopoietic progenitor cell (hpc) transplants for haematological disorders, peripheral blood stem cells are routinely collected via apheresis and cryopreserved. leicester royal infirmary had been using a controlled rate freezer (crf) to cryopreserve cellular therapy products up until . in a literature review of cryopreservation techniques was undertaken, since the crf required replacement. this review found consistent evidence that cryopreservation using minus o c is comparable to crf, and engraftment times should not be negatively affected by changing to a more simplified method of freezing. there would also be cost saving benefits from switching from a crf to minus o c freezers. methods: as a result two minus o c freezers were purchased following the acceptance of a preparation process dosier (ppd) which was prepared for the human tissue authority. validation was carried out, and from january stem cell laboratory at the lri switched from the crf method to minus o c for cryopreservation of cellular therapy products. briefly, cells are frozen using % dmso (wak-chemie) in g/l human albumin solution (grifols) in cyrobags (origen biomedical, inc). the cells are transferred on cold packs to the minus o c freezer. cells are packaged in between stainless steel heattransfer plates. the plates are placed within a bubble wrap bag, and are placed in a rack within the minus o c freezer, which allows air to circulate freely around each bag. in one plate a el-usb-tc thermocouple data logger (thermosense) is inserted between the bag and stainless steel plate, to record the freezing profile. this data is downloaded after each run. after an overnight freeze at minus o c, the cells are subsequently removed from the bubble wrap and plates, and are transferred to minus o c freezers the following morning. results: a total of patients have had autologous stem cells frozen using this method so far this year. in addition to engraftment data for neutrophils, post-freeze trypan blue viabilities were also compared to the previous year. during a total of patients, who had all their cells cryopreserved, underwent collections. the post freeze median viability was % ( - %). a total of median neutrophil engraftment was . days, with a median cd dose infused of . x /kg. during so far, patients have undergone collections. median viability is . % ( - %). subsequent median neutrophil engraftment is days, with a median cd dose infused of . x /kg. conclusions: ongoing savings of approximately £ per annum have been made by changing our procedure. the benefit of changing to a simplified method of freezing has also resulted in a reduction in staff working overtime. more importantly, this simplified cryopreservation method has resulted in an improvement in neutrophil engraftment times since changing the cryopreservation technique from the previous method using crf to mechanical freezing using a minus o c freezer. disclosure: nothing to declare low doses of granulocytes in the apheresis product predict a better outcome of autologous hematopoietic stem cell transplantation in multiple myeloma patients background: high-dose chemotherapy with autologous stem-cell transplantation (asct) remains the standard consolidation therapy for multiple myeloma (mm). peripheral blood stem cell collection may be contaminated with large quantities of granulocytes and its consequences on the outcome of asct are still unclear. on the other hand, the effect of performing apheresis with high levels of monoclonal component (mc) on outcome is unknown. the objective of this study was to analyze the effect of total nucleated cells (tnc) and granulocytes count (considered as contaminating components of apheresis products) as well as the influence of mc in the apheresis product on outcome of asct in mm. methods: eighty-two patients diagnosed with mm were mobilized with filgrastim μg/kg/day (plus plerixafor if insufficient mobilization of cd + cells on day ). apheresis collection was performed with cmnc program by spectra optia cell separator. cd + count was carried out according to ishage protocol (target: ≥ x e cd +/kg). subsequent cryopreservation were performed according to the local protocol. results: the medians (range) of collected cd +, tnc and granulocytes were . x /kg ( . - . ), . x / kg ( . - . ) and . x /kg ( . - . ), respectively. the medians (range) of infused cd +, tnc and granulocytes were . x /kg ( . - . ), . x /kg ( . - . ) and . x /kg ( . - . ), respectively. a successful collection after first line therapy was performed in % of patients. treatment for mm was continued after carrying out apheresis in % of patients, as per protocol. a significant reduction of mc was observed prior to asct, indicating a further improvement in responses after apheresis (p= . ). an optimal response (cr or vgpr) at the time of apheresis was achieved in % of patients and a suboptimal response (partial or minimal response) was observed in the remaining %. undergoing apheresis in optimal response did not result in lower number of tnc or granulocytes in the harvest. the subtype of mc did not influence on the number of tnc and granulocytes in the product of apheresis. no differences in collected tnc and granulocytes were observed when plerixafor was used as mobilizing agent. the type of chemotherapy given prior the apheresis did not have influence on the characteristics of the apheresis product. a significant improvement in overall survival (os) probability ( %ci) was observed when low tnc (< . x figure a ). lower incidence of relapse (p= . ) ( figure b ) and non-relapse mortality (p= . ) was observed in patients who received low granulocyte count in the graft. no significant correlation was observed between the time of engraftment and the number of tnc or granulocytes infused. similarly, no increase in the frequency of the engraftment syndrome was observed when higher number of tnc or granulocytes were infused. conclusions: in our series, low doses of granulocytes in the product of apheresis predicted a better outcome of asct in mm patients. the amount of mc at the time of apheresis did not have influence in the characteristics of the harvest. efforts for avoiding contamination in grafts are important for its impact on outcome of asct in mm patients. disclosure: dkms foundation, pi / (instituto carlos iii) and sgr (grc), generalitat de catalunya. background: our initial experience (from to years) with hematopoietic stem cell transplantation (hsct) from mud with tcrαβ+/cd + graft depletion in patients with cgd showed high rate of secondary graft dysfunction, the incident of graft rejection was %. to improve the outcome, we hypothesized that the use of plerixafor and g-csf as additional agents in conditioning regimen would offers advantages and better outcomes. this trial was registered at www.clinicaltrials.gov (nct ) methods: between april and september , patients with cgd underwent allogeneic hscts from mached unrelated donors with tcrαβ + / cd + graft. the conditioning regimen included -treosulfan g / m , fludarabine mg / m , thiotepa mg / kg, g-csf mcg / kg and plerixafor mcg / kg. all patients received rabbit atg -timoglobulin® ("genzyme europe b.v.", the netherlands) mg/kg for serotherapy. in all cases, tcrab +/cd + graft depletion was used with the immunemagnetic method (miltenyi biotec, bergisch gladbach, germany) according to the manufacturer's instructions. stem cell source was g-csf mobilized peripheral blood stem cells in all cases. posttransplant gvhd prophylaxis was not performed. minimal follow up was days after hsct. results: neutrophil and platelet engraftment occurred at - days post-hsct in all patients. patients had full donor chimerism in whole blood and in cases we observed predominantly donor mixed chimerism at last follow up. all patients had full donor chimerism in cd + compartment and mixed chimerism in cd + lineage, but it stable without any sign of graft dysfunction. acute gvhd is verified in of cases and was limited to skin grade . all patients are alive for periods from to months post-hsct with good graft function without severe clinical problems. conclusions: we presented first experience of g-csf and plerixafor addition to conditioning regimen before hsct with tcrαβ+/cd + graft depletion in cgd patients . we suppose, the preliminary results are encouraging, as the frequency of primary and secondary graft dysfunction in patients from this group is not observed today, there are no significant toxic complications, as well as clinically severe manifestations of gvhd. currently, the recruitment of patients is continuing, and estimation of the rates of immune reconstitution and a more detail analyses will be evaluated later. background: introduction -it is believed that aboincompatibility is of minor importance in allogeneic stem cell transplantation (allo-sct) and that the clinical outcome is equivalent to abo-compatible sct. therefore, we performed a single center retrospective study to characterize the impact of abo-incompatibility on the outcome of haploidentical stem cell transplantation (haplo-sct). methods: this analysis included consecutive patients who underwent haplo-sct for various hematological malignancies at our center between october and may . we used our institutional database to evaluate details and characteristics of patients and transplant outcomes. results: demographic features of the patients and donors have been summarized in table . all of the patients had advanced hematological disease with a high risk of relapse ( patients with acute leukemia). out of patients, early transplant-related mortality was seen in this cohort of patients. the remaining patients were followed in and months. donor type abo group switch was observed in a median of days ( - days) after transplant. we were not able to show any statistical difference in terms of blood group switch between minor and major abo incompatible transplant. the median red blood cell (rbc) transfusions in the first days for the abo compatible and incompatible transplants were median units (range, - ) and median units (range, - ) (p= . ). no statistical difference was also encountered for the rbc transfusion need for stem cell source, peripheral blood vs bone marrow. a total of patients were followed up for reticulocyte engraftment. the median time for reticulocyte engraftment was days (range, - ) for all patients. reticulocyte engraftment was tended to be faster in minor abo-mismatched group (p= . ) than major or abo-compatible ones. nineteen patients achieved independence from rbc support after a median time of days (range, - days) in abocompatible patients, days (range, - days) in minor abo-incompatibilityand days (range, - days) in major abo-incompatibilitygroup, respectively (p> . ). the engraftman kinetics due to major and minor aboincompatibilitytransplants were presented in table- . pure red cell aplasia was not developed in our cohort. conclusions: the present single center study provides new evidence for the importance of the abo system for erythrocyte recovery in haploidentic stem cell transplantation. it's important to note that, randomize prospective and larger studies are warranted. disclosure: nothing to declare low dose of anti-t lymphocyte globulin protects against severe forms of graft versus host disease in patients undergoing allogenic stem cell transplantation background: graft versus host disease (gvhd) is the most important complication after allogeneic stem cell transplantation (allosct). optimal dose of different anti-tlymphocyte globulin (atg) formulations in this setting has not been established yet. the aim of this study was to analyze the impact of a low dose of atg-fresenius (atg-f) in allosct outcomes. methods: we analyzed adult patients who received an allosct for hematologic malignancies from october to march . the gvhd prophylaxis included a total dose of mg/kg ( mg/kg on day - , - and - ) of atg-f for patients who received a graft from peripheral blood, an unrelated donor; with a mismatch, and/or were older than years; associated to a calcineurin inhibitor and mycophenolate mofetil/short course-methotrexate. statistical analysis was performed using spss v. and r software. results: median age was years ( - ) and . % of patients were males. seventy-four percent of patients underwent myeloablative conditioning. the stem cell source was peripheral blood in patients ( . %), % were from unrelated donors ( % mismatched). seventeen ( . %) patients had high risk cmv status (d-/r+) (see image b). engraftment was observed in patients ( . %). primary graft failure occurred in patients ( myelofibrosis, aml). twenty ( . %) out of evaluable patients developed grade - acute gvhd. the cumulative incidence of severe agvhd and moderatesevere chronic gvhd were . % ( % ci, . - . %) and . % ( % ci . - . %), only ( . %) patients developed severe cgvhd. twenty-nine patients ( . %) discontinued immunosuppression before the first year of transplant. the median duration of immunosuppression for patients with moderate-severe cgvhd was days ( - ). at years non-relapse mortality (nrm) was . % ( % ci, . - . %). thirty-nine ( %) patients developed relevant infectious complications. two ( %) patients died within the first days due to gram negative blood stream infection. eleven ( . %) had at least two episodes of cytomegalovirus (cmv) reactivation between day and . three ( %) patients developed cmv gastrointestinal disease, ( %) had probable invasive fungal infection and ( . %), post-transplant lymphoproliferative disorder associated to epstein barr virus. with a median follow up of months for alive patients , the gvhd and relapse free survival (grfs) at one year, overall survival (os) and progression free survival (pfs) at two years were . % ( % ci, . - . %), % ( % ci, . - . %) and % ( % ci, . - . %), respectively. the relapse incidence at two years was . % ( % ci . - . %). complete remission at transplant was associated with better long term survival ( % at years, p < . ). hla disparity did not affect os (see image a). conclusions: the use of low doses of atg-f is protective against severe forms of acute and chronic gvhd in a cohort with high prevalence of unrelated donors and a high median age. this strategy showed good results in grfs, os and pfs in a population at high risk for developing gvhd or relapse. disclosure: nothing to declare performance parameters of a ngs-product for chimerism monitoring -applicable in patients after hematopoietic stem cell transplantation methods: for this purpose, samples from patients with mixed chimerism (mc) with increasing amounts of recipient dna were analyzed and compared using realtime pcr of insertions/deletions (indels), fragment analysis of short-tandem repeats (str) and ngs of indels. results: whereas real-time pcr displayed excellent sensitivity down to , % mc, but poor precision above %, fragment analysis exhibited good precision with limited sensitivity (> , %). in contrast, ngs chimerism demonstrated good sensitivity, with a limit of detection (lod) of , % mc, and precision throughout the whole spectrum of patient/donor mixed chimerism. the ngs chimerism product (devyser chimerism) exhibited at least three (average eight) and at least two (average ) informative genetic markers (indels), suitable for monitoring mixed chimerism of patients with their corresponding matched unrelated ( ) or related ( ) donor samples. in order to establish the performance of the separate techniques for determination of mixed chimerism on retrospective patient samples, a cohort of patient monitoring samples ( - weeks post-hsct) with low (< %), intermediate or high mixed chimerism (> %) were included and analyzed. dna from all monitoring samples was extracted from sorted cell fractions. the results show that although all evaluated techniques are suitable for monitoring patient/donor chimerism after allogeneic hematopoietic stem cell transplantation (hsct), only the ngs chimerism product exhibits high sensitivity (lod , %) and a broad dynamic range (detection range , - %) with good precision and accuracy throughout the whole spectrum of mixed chimerism (% patient/donor). in addition, the ngs chimerism product employ non-population dependent highly informative genetic markers providing stable resolution power and thus suitable for monitoring mixed chimerism. disclosure: dan hauzenberger is medical adviser at devyser ab and shareholder in devyser holding gender distribution: male - % (n= ), female - % (n= ). age median - , years old ( months - ). stem cell source: bone marrow - % (n= ), peripheral blood stem cells - % (n= ). patients ( %) received / matched unrelated donors hematopoetic stem cells transplantation and patients ( %) - / matched unrelated donors hematopoetic stem cells transplantation. differences in the antigen blood system: single group % (n= ), minor % (n= ), major % (n= ), mixed % (n= ). age of donor: - years old - % (n= ), - - % (n= ), - - % (n= ), and more % (n= ). gender differences in donor/recipient: male/female % (n= ), female/male % (n= ), one sex % (n= ). we also took into account the impact of gender difference and cytomegalovirus serostatus in the donor/recipient pair. results: in / group the estimated probability of overall -years survive was % and in the / group -years survive was %. the increase in donor age of years reduces the -years survive by - % (p= , ), however, the -years survive from donors over years old was %. we have found no difference between -years survive in transplants from donors that are compatible/ incompatible with the antigen blood system, cytomegalovirus serostatus, or the gender differences in donor/ recipient. in the study of donor-related factors, we found the negative impact of an human leucincompatibility ( / ) on the incidence of chronic gvhd - % (p = . ). the combination of cytomegalovirus positive serostatus of the donor and the negative status of the recipient increases the risk of primary graft rejection up to %, in comparison with others (p = . ). conclusions: our study showed the role of genetic matching on the hla system between the patient and the unrelated donor, and the donors age value. / transplants have better outcome and lower incidence of severe a. gvhd and ch. gvhd. younger donor increases -years survive, but there is a significant increase in -years survive if the donor is over years old. disclosure: nothing to declare allogeneic stem cell transplantation in chronic myelomonocytic leukemia. a single center experience nine patients ( %) relapsed with a median of , months ( - ) with different strategies at this point: in all cases we modulated immunosuppression, in cases as the unique strategy, in cases with donor lymphocyte infusion (dli), in cases we employed hypometilating agents (hma) and in cases with intensive chemotherapy, reaching cr only in two patients, one of them after dli and the other one after hma and consolidation with a second asct. eleven patients ( %) died being the relapse the main cause ( %). transplant related mortality (trm) at + were % and global trm were %. in the last follow-up, patients ( %) are still alive, ( %) in cr and ( %) in relapse situation. with a median follow-up of months ( - ), the event free survival (efs) were months ( - ) and the overall survival (os) were months ( - ). we observed advantage in terms of os in those patients that reach cr at + post asct and in those who develop chronic gvchd (p= . and p= . respectively) conclusions: asct is still the only curative option despite the high relapse rates. to reach cr at + post asct and the development of chronic gvhd seems that they confer advantage in terms of os. the importance of knowing the molecular profile of the entities that we consider for asct. disclosure this study documents a first experience of a cell processing lab seeking to integrate process automation technology to wash and volume reduce products which can account for the initial material source volume variability, product characteristics, and number of bags. methods: here we report the pre-clinical assessment of the lab's initial work with the lovo cell processing system for a product experience over days with machine. this study used products intended for destruction. the workflow used parallel and sequential processing schedule. after water-bath thawing, bags were sampled, weighed to determine volume, and subsequently connected to lovo or pooled into a transfer pack and then connected to lovo. the bags were then diluted : at ml/min with lovo at + - c using % hydroxyethylstarch / . (voluven, fresenius kabi) and processed using a cycle procedure. after processing the bags were weighed for volume, sampled, and stored in a - c refrigerator in their lovo final product bags. samples were assessed from t= to t= hours. results: cd + viability and absolute counts were determined using flow cytometry. processing duration and solution volume consumed was determined by the lovo's sensors and confirmed by the operator. data is presented as a percentage relative to the post-thaw values. note, the values presented are not total process yields. the results focus on the lovo processing step. conclusions: the operators easily integrated into the software to drive the machine. the machine demonstrated it's flexibility with a wide-range of volumes, cell-inputs, and number of bags. the lovo produced products which meet our specifications in a quick and reliable manner. further work on this platform will be performed to validate and qualify this system for production use. properties. the aim of this study was to evaluate prospectively the efficacy of a fbm protocol for the prevention of om in patients undergoing a hsct. methods: all patients consecutively who underwent a hsct at our center from x onwards received five weekly fbm sessions with a bidiodic laser (lumix ®, prodent, italy), which simultaneously emitted at nm and nm with a power of mw and energy of j per point. the procedure started the day before the beginning of the conditioning regimen up to the tenth day post-transplant. the laser was applied in a defocused mode on each of the mucosal surfaces ( areas). at each session, the morphine dose, the om level (according to the who scale) and pain through a numerical rating scale (nrs) were recorded. results: consecutive patients ( male/ female) submitted to a hsct were analyzed. the median age was years (range - ). eighteen patients had acute leukemia, myelodysplastic syndromes, lymphoproliferative diseases. the median number of treatment lines before hsct was (range - ). at transplant, patients had advanced disease. the myeloablative conditioning regimen mac (thyotepa, busulphan, fludarabine) was employed in patients; the same conditioning, with a reduced dose of busulphan (ric), was infused in patients. seven patients ( %) had no evidence of om. the incidence of grade ii-iv om was % in the group of patients receiving mac and the median duration days (range - ); grade om was observed, for day, in patient. in the ric group the incidence of om was %, the median duration days (range - ); no patient had evidence of grade iv om. in the whole population, the maximum nrs value was . morphine administration was required in patients, due to the occurrence of non-oral complications. conclusions: in our experience, prophylaxis with fbm to prevent or reduce om was safe and effective, compared to results of previous experiences reported in the literature, which used no prevention against this complication that negatively affects the quality of life of transplanted patients. further studies on a large series of are necessary to confirm our results. disclosure: nothing to declare background: cytogenetic abnormalities are an essential part of prognostic systems in myeloid malignancies before hematopoietic stem cell transplantation (hsct), however, their role in posttransplantation prognosis is unknown. the aim of this study was to assess the prognostic impact of genetic risk stratification of aml and mds patients on posttransplantation course, which could be an additional tool in making decisions regarding preemptive therapy. methods: a retrospective analysis covering patients treated with allo-hsct between and . cytogenetic studies included karyotyping (c-and gbanding) and fluorescence in situ hybridization (fish). the number of analyzed cells exceeded european cytogenetics association guidelines (for each fish at least interphase nuclei were analyzed). cytogenetic risk group in aml was assessed based on the eln criteria. patients with mds were stratified into three groups; favorable (good and very good prognostic score), intermediate, and adverse (poor and very poor) prognostic score according to ipss-r . interestingly, the poorest survival was in patients with monosomy of chromosome , which was present in patients of whom succumbed to refractory disease, while all patients who had deletion of long arm of chromosome (del q)-are alive at the time of writing of this report after a median follow-up of months ( - ). relapse was diagnosed in patients ( %), including; ( %) with adverse, ( %) with intermediate and ( %) with favorable cytogenetic risk. among patients with a complex karyotype and/or cytogenetic evolution prior hct: patients ( %)relapsed, including ( %)-who died. follow-up cytogenetic studies in relapse after transplantation were performed for patients; of them ( %) had clonal evolutions of the original karyotype with additional abnormalities-( % died) and ( %) had new aberrations in cells without primary changes (all died). in patients ( %) ( unfavorable, intermediate group) cytogenetic relapse was diagnosed by fish analysis and they were treated with azacitidine (+/-dli) achieving cr (n= , %), stabilization-(n-= , %), transient response (n= ), while deceased). conclusions: cytogenetic studies in patients after transplantation may facilitate assessment of mortality. karyotype may undergo cytogenetic evolution after allo-hsct. patients with monosomy of chromosome seem to have a particularly poor prognosis. transplanted patients are vulnerable to new cytogenetic alterations. disclosure: nothing to declare methods: the primary end-points were the rate of complete response (cr), defined as no emesis and no nausea without rescue medications, for both acute (cr- ) and delayed (cr - ) cinv and rate of post-transplant complications until discharge. we prospectively analyzed patients undergoing autologous ( %) and allogeneic ( %) stem cell transplantation and receiving cinv prophylaxis with nepa and dexamethasone (schedules shown in fig. ). in our series, patients ( %) were female. patients median age was years ( - ). the most frequent diagnosis were myeloma ( %) and lymphoma ( %), while % of patients were diagnosed with aml or mds. myeloma patients received one day hd-ct with melphalan ( % mel / % mel ). lymphoma patients were conditioned with feam ( , %) or tt_flu_edx ( , %) hd-ct. busulfan-based mds-ct regimen was offered to aml/mds patients. results: the incidence of cr- and cr - observed was % ( / ) and , % ( / ), respectively. more than grade nausea and vomiting (according to ctae- ), was reported in % ( / ) and % ( / ) of patients, respectively. female sex was associated with an increased risk of acute (hr , ; p , % ci . - . ) but not delayed (hr , ; p , % ci - . ) cinv. similar rate of cr and cr - was observed in one-day hd-ct ( % and %) compared to mds-ct ( % and , %) group (pns). median lenght of stay was days ( - ). no case of cardiotoxicity and no exitus was observed. the incidence of febrile neutropenia was % ( % fuo; % sepsis; % pneumonia). only one patient experienced an agvhd on day + . neutrophil (> /mcl) and platelet (> /mcl) recovery occurred in median on day ( - ) and on day ( - ) respectively. conclusions: nepa seems to be safe and effective in preventing acute and delayed cinv in patients receiving both one day hd-ct and mds-ct as conditioning regimen for hsct. more studies are needed to define the better -ht ra and nk- ra combination and the better schedule in transplant setting. disclosure: "nothing to declare background: vod and ta-tma represent two early endothelial complications occurring after allogeneic stem cell transplantation (sct) sharing many pre-transplant risk factors. the aim of our study is to evaluate the impact of donor graft composition, engraftment kinetics and infections on the development of these endothelial complications (ec). methods: we retrospectively reviewed consecutive sct recipients at our institu-tion between january and june . the median age was years (range - ). acute leukemia was diagnosed in patients ( %). complete remission was documented in % of patients at transplant. donor source was from hla mismatched donor in % and from unrelated donor in % of the patients. hbv positive patients were % of the sample. conditioning regimen was busulfan based in % of patients. ursodeoxycholic acid and unfractionated heparin were given to all patients as vod prophylaxis. cyclosporine was used as gvhd prophylaxis. lymphocytic subpopulation analysis (cd +, cd +, cd + and cd +/cd +) and cd + cells count on the donor graft were performed using bd facs cantoll. all patients had routine monitoring for ebv and cmv pcr, hemolysis tests, creatinine and electrolyte panels, proteinuria, complete blood count, blood pressure and schistocytes by direct examination until day + . the fisher's exact test was used to compare categorical variables, while continuous variables were analyzed with anova test. diagnosis of vod and ta-tma were carried out by using ebmt and cho criteria respectively. results: the incidence of very severe vod and tma was % ( / ) and , % ( / ) respectively. cmv reactivations with viral load over . cv/ ml was % and % in patients with and without ec, respectively (p , ; hr , p , %ci , - , ). the median day to neutrophils (ns) engraftment ( /ml and /ml) was vs and , vs in vod/tma group vs control group (p , and , , respectively). more rapid neutrophils engraftment (ns > /ml and ns> /ml within days) was related to a higher risk of ec with a hr of , (p , ; %ci , - , ) and , (p , ; % ci , - , ). patients with ec received a donor graft with a higher median numbers (x e /kg) of cd + and cd + (p> , ) and a lower numbers (x e /kg) of nk cells (p> , ). patients who received a cd + cells count > x e /kg and nk cells count < , x e /kg presented a relative risk of ec of , (p , ; % ci , - , ) and , (p , ; % ci , - , ), respectively. there were no differences with respect to the other analyzed variables between patients who developed vod/tma compared with those who did not. (pic_ ) conclusions: cmv viremia, early neutrophils engraftment and donor nk and cd + cells infused are associated with the risk of vod and tma. very few studies evaluated the link between these variables and the risk of developing such two complications. it could be interesting to investigate these relationships on larger series. clinical trial registry: not applicable disclosure: nothing to declare p when the last hope turns out to be just as good as best: haplosct following tbf conditioning, pt cyclophosphamide and tacrolimus as gvhd prophylaxis background: hematopoietic stem cell transplantation is an effective therapy for a variety of severe hematological diseases. in last decades, haploidentical sct (haplosct) followed by ptcy as gvhd prophylaxis has been reported as a valid alternative for patients who lack an hla matched donor. we therefore analysed outcomes with this. methods: patients without hla-matched donor received a haplosct between / and / . thiotepa mg/kg ( days), fludarabine mg/m ( days) and oral busulfan mg/kg/ h ( days,with pkd dose adjustments) was used as conditioning regiment; in patients > years busulfan administration was limited to two days. gvhd prophylaxis consisted of cyclophosphamide mg/kg on day + and + , and tacrolimus as a continuous iv infusion from day + . all patients received pbsc as the stem cell source. outcomes analysed were overall survival (os), progression free survival (pfs); cumulative incidences (ci) of gvhd, relapse and non-relapsed mortality (nrm). results: median ages was (range - ). % male and % female. diagnoses were: aml ( %), mds ( %), all ( %), hl and nhl ( %), and mm ( %). % (n ) were transplanted in early disease status, while most cases ( %) were in advanced status, including, second/third cr ( %, n ), one ( %) in aplasia without progressive disease and % (n ) had active/progression disease, % (n ) had stable disease; four cases were second alosct. thus, % of patients had a high or very high rdri and % had intermediate. ebmt score was ≤ was in % of patients (n ) . the donor was as son/ daughter in %, % a sibling and % a patient's mother. median time to neutrophil ( . x e /l) and platelet (> x e /l) recoveries were + and + days, respectively (g-csf was not used). only one patient had a primary graft failure attributed to anti-hla donor specific antibodies. median follow up in survivor is months (range - ). overall survival is ± . % (at months) and ± % (at months). pfs is ± % and ± % respectively. the cumulative incidence of relapse was % and %, respectively, while nrm is % at months. day + , grade - acute gvhd were %, while mild/ moderate and severe chronic gvhd were % and % respectively. ebmt ≤ and first alosct were the only variables to clearly impact -months os in univariate analysis. a combined covariate of ebmt ≤ -no prior alosct vs other patients showed a month os ± % vs ± % (p . ), pfs ± % vs ± % (p . ) and nrm % vs % (p . ) but without impact on relapse % vs % (p . ). conclusions: haplosct with an age-adapted tbf conditioning regimen, pbsc and ptcy followed by tacrolimus, led to very encouraging results, mainly in patients with a low ebmt score and as a first alosct. although formerly considered as a last alosct strategy, we now agree that the time has come to compare this strategy with hla mud (and even elderly sibling donors) in ongoing prospective randomized multinational trials. disclosure: nothing to disclosure background: autologous hematopoietic stem cell transplantation (autosct) for acute myeloid leukaemia (aml) is increasing becoming a viable option for an increasing number of patients due to limited availability of matched sibling or unrelated donor for allogeneic hematopoietic stem cell transplantation (allosct). we examined the relevant long-term outcomes in our local patient cohort. methods: we retrospectively reviewed the data for all autosct done for aml in our centre over a -years period between st january until st dec from our electronic record. patients with acute promyelocytic leukaemia (apml) were excluded from this analysis. patients were further stratified based on the number of high risk features present; not achieving complete remission (cr) following induction chemotherapy, high presenting total white cell count (wbc > x /ml, adverse cytogenetics (example: complex cytogenetics) and adverse molecular mutations (example: flt -itd & mll gene arrangement). outcome data including mortality (overall survival (os) and non-relapse mortality (nrm)) and morbidity (leukaemia free survival (lfs)) were recorded and analysed. results: a total of patients were identified. median age at diagnosis is -years old. the cohort comprised of males and females. the overall median os and median lfs is . years and . years respectively. the nrm is . % ( / ). there was no difference in the median os and median lfs for the patients achieving cr following induction chemotherapy and those not in cr following induction chemotherapy; . years versus . years (log-rank, p= . ) and . years versus . years (log-rank, p= . ) respectively. the median os were statistically significant for patients with zero versus one and two and more high risk features present; . years versus . years versus . years (log-rank, p= . ) respectively. however, the median lfs were not statistically significant for these three patient cohorts; . years versus . years versus . years (log-rank, p= . ) respectively. conclusions: in our patient cohort, autosct appeared to be a feasible option for patents with aml without matched sibling or unrelated donor available. disclosure: none to declare methods: between - , thalassemic patientsunderwent hisct. the median age of patients was ( - )years with male preponderance (n= , . %). across the gender and abo mismatch transplants were done in . % and % of patients. stem cell source was bone marrow in ( %) while peripheral blood in ( %) of patients. mean stem cell dose was . ± . x cells / kg and mean volume of product was ± . ml. preparative regimen included anti-thymocyte globulin, busulfan, fludarabine and cyclophosphamide.graft versus host disease (gvhd) prophylaxis comprised of posttransplant cyclophosphamide on day + & + followed by tacrolimus and mycophenolate mofetil. patients were observed for hematopoietic recovery (neutrophil and platelet engraftment) and transplant related mortality including acute and chronic gvhd for skin, gut, liverand lungs, primary and secondary graft failure and infectious complications. results: nine( . %) of sixteenpatients were engrafted with full donor chimerism. twelve ( %) patients belonged to pesaro class i and ( %) toclass ii patients. median time to neutrophil and plateletengraftment were ( - ) and ( - )days respectively. average number of packed red cell and platelet transfusions were . ± . and . ± . respectively.primary graft failure was observed in ( %) and secondary graft failure was observed in ( %) patients. two patients received a second dose of stem cells and they engrafted at and days of infusion respectively. of patients with primary graft failure died, one with sepsis (day + ) and the other because of intracranial bleeding (day + ).acute gvhdof gut and skin (grade ii-iii) was observed in patients each, within first days post-transplant. none of the patients had grade iv gvhd. cytomegalovirus reactivation occurred in % of patients, all of them received pre-emptive therapy with intravenous ganciclovir. none of them developed cmv disease. invasive fungal infection was not observed in any of the patient. culture proven bacterialinfection was documented in % of patients requiring intravenous antibiotics during first days post-transplant.overall survival and relapse free survival were . % and . % over a median follow-up of ( - ) days. conclusions: haploidentical transplant is a suitable modality for thalassemic patients lacking a full matched donor in pakistan. in view of our results, we suggest that thalassemia patients should be offered hisctas an option for cure. clinical background: gemtuzumab ozogamicin (go) is an anti-cd monoclonal antibody with significant activity in de novo and relapsed/refractory (r/r) acute myeloid leukemia (aml). a relevant side effect consists of hepatotoxicity and especially sinusoidal obstruction syndrome (sos). the objective of this study was to analyze tolerability of go during the induction and reinduction therapy in patients with aml, and its possible impact on subsequent hematopoietic stem cell transplantation (hsct). methods: from to , patients who had received go in three hospitals were collected and their medical records were retrospectively reviewed. results: fourteen patients diagnosed with de novo aml received go ( mg/m ) on day + in combination with standard chemotherapy (idarubicin and cytarabine, x schedule) as induction therapy. hyperbilirubinemia (bilirubin > . unl) was detected in patients and increase of aspartate aminotransferase (ast) (> . unl) in . twelve patients achieved complete remission (cr) and one was refractory ( not evaluated). in the r/r setting, patients diagnosed with aml (n= ), biphenotypic acute leukemia (n= ) and acute promyelocytic leukemia (n= ) received go as rd or subsequent rescue therapy either as monotherapy (n= ) or in combination with cytotoxic chemotherapy (n= ). prior hsct was performed in patients (autologous [n= ], allogeneic [n= ] ). rescue therapy was indicated for refractoriness (n= ), relapse (n= ), partial response (n= ) or absence of donor (n= ). four patients received doses of go ( mg/m ) and patients, one dose. hyperbilirubinemia (> . unl) was observed in patient and increase in ast (> . unl) in patients. seven patients achieved cr, was refractory, obtained partial response and died early during induction). thirteen patients received subsequent hsct (autologous [n= ], allogeneic [n= ]) after go therapy ( in the de novo aml and in the r/r group). the reasons for not performing hsct in the remaining patients were: low cytogenetic risk (n= ), active chronic graft versus host disease (gvhd) in previous hsct (n= ), early death during treatment (n= ), relapse (n= ), severe complications in rescue treatments (n= ), and unknown (n= ). the conditioning regimen was myeloablative (n= ), non-myeloablative (n= ) and sequential (n= ), and the donors were matched sibling (n= ) or unrelated (n= ) . cyclosporine, methotrexate and thymoglobulin were administered as gvhd prophylaxis in patients and cyclosporine, mycophenolate and thymoglobulin in . hyperbilirubinemia was observed in patients belonging to the de novo aml group. death after hsct occurred in patients due to infection (n= ), relapse (n= ), gvhd (n= ) and traffic accident (n= ). three patients are currently alive in remission. no sos was observed in any patient. conclusions: in both de novo and r/r aml the administration of low dose go is feasible and does not have impact on subsequent hsct outcome. although some degree of hepatotoxicity was observed, no cases of sos were observed, either before or after hsct. disclosure: supported by grants from: asociación española contra el cáncer, aecc (gc biga), instituto carlos iii (pi / fi), -sgr (grc), cerca program from generalitat de catalunya, and "la caixa" foundation. outcome of allogeneic hematopoietic stem cell transplantation in patients with benign hematological disorders saqib ansari , tahir shamsi , uzma zaidi , saima siddiqui , tasneem farzana background: allogeneic hematopoietic stem cell transplantation is a potentially curative treatment modality for hematological disorders. we evaluated the outcome of patients suffering from benign hematological disorders, including aplastic anemia, fanconi's anemia and thalassemia after matched related allogeneic transplantation. methods: all patients having hematological disorders including aplastic anemia (aa), beta thalassemia (btm), fanconi's anemia (fa) and severe combined immune deficiency disorder (scid) with hla identical related donors who underwent allogeneic transplantation were included. donors were given g-csf at a dose of μg/ kg/day daily for four days prior to harvest. the conditioning regimens for thalassemia included cyclophosphamide (cy) + busulfan (bu) in ( %), bu + cy + thiotepa in ( %) and bu + cy + antithymocyte globulin (atg) in ( %). conditioning regimens for aplastic anemia included, fludarabine (flu) + cy in ( %), flu + atg in ( %) and cy in ( %), cy+ atg in ( %) patients. for fanconi's anemia flu + atg in ( %), flu in ( %), cy + atg in ( %) and flu+ cy + atg in ( %). flu + atg in ( %) and cy given in ( %) and no conditioning regimen was offered to ( %) patients with scid. results: a total of allogeneic transplants were performed for benign hematological disorders including aa (n= ), btm (n= ), fa (n= ) and scid (n= ) from to july . median age was . years (range . - ). across the gender and abo blood group transplants were ( . %) and ( %). the median time to neutrophil and platelet recovery was days (range: - ) and (range: - ). primary and secondary graft failure was observed in ( . %) and ( . %). overall survival in aplastic anemia ( / , %), beta thalassemia ( / , %),fanconi's anemia ( / , %) and severe combined immune deficiency disorder ( / , %). eighty four patients expired ( . %) among them patients expired within days post transplant main cause of deaths included sepsis ( %), multi organ failure ( %) and gut gvhd ( %) conclusions: in developing world scenario where non malignant disorders are leading cause of morbidity and mortality. bone marrow transplantation has been successfully implemented with better long term diseases free survival and quality of life. clinical background: hematopoietic cell transplantation (hct) remains the only curative therapy for many diseases, yet transplant survivors carry an unusually high burden of morbidities, primarily because of exposure to intense chemotherapy, radiation and /or gvhd. this study aimed to evaluate the burden of chronic diseases at the end of life after allogeneic-hct and to identify the disability-adjusted life years (daly). methods: the pubmed, medline, and ovid databases were queried utilizing specific mesh terminology (post, allo stem cell, hematopoietic, bone marrow, transplantation).we collected data on the impact of the hct on the variables affecting survivor's health in all aspects .the rates of late complications were compared to the risks in the general population (united states). results: a total of studies fulfilled the selection criteria totaling to patients (table ) . median os at -year mark varied widely between studies from % to %. majority of the patients at -year mark were found to have new comorbidities thereby indicating a huge burden of late effects at the end of life, though exact dalys could not be calculated due to incomplete data. hct survivors were found to have higher risk of premature arterial disease (pad) at . % compared to the general population, however gvhd or the addition of tbi to the conditioning regimen were not found to be significantly associated with pad. regarding the risk of new cancers, the cumulative incidence of their development at and years was . , and . , respectively. increased risks compared with the general population were seen for some solid cancer including cancers of the lip: p= . , tonsils: p= . , oropharynx: p< . , bone: p< . , soft tissue: p< . , and vulva: p= . . with respect to mental health, depression was prevalent in ( . %) survivors, in whom ( . %) were still on antidepressants at the last follow-up. cognitive impairment and other psychiatric disorders were found in ( . %) and ( . %) survivors, respectively. the most common cause of nrm in the first years was gvhd. however, after years, the leading cause of death in those conditioned with mac regimen was secondary cancer, but in the ric group, new cancers and gvhd contributed equally. conclusions: hct survivors remain at risk of significant complications which lead to premature death and their burden of comorbidities at the end of life is significantly more than that of general population. background: late onset hemorrhagic cystitis (hc) is a common complication of hematopoietic stem cell transplantation (hsct) frequently associated with reactivation of bk virus (bk-hc).there is no consensus as to the best therapy for bk-hc, and many different treatments have been reported. hyper baric oxygen therapy (hbot) is used as primary or adjuvant therapy in diverse clinical situations involving hypoxic injury to tissues and has been explored as a useful tool in treating bk -hc. we report our experience with hbot in combination with non-invasive supportive care in children and adolescents suffering from bk-hc following allogeneic hsct. methods: the computerized database of schneider children´s medical center of israel was reviewed for all patients aged to years who underwent hsct between january and june and developed bk-hc. hbot therapy consisted of hours sessions at atmospheres, with patients breathing oxygen by mask. parents accompanied patients during the treatment. results: fourteen patients with a variety of underlying diseases received (hbot) for treatment of bk-hc following hsct. the initial treatment for children with bk-hc at our center prior to included continuous bladder irrigation and intravesicular instillation of various medications. beginning in , we adopted a non-invasive strategy that included the administration of oral anticholinergics (oxybutinin), systemic pain management, hyperhydration and the administration of weekly cidofovir with probenecid. hbot was administered to patients who failed the above regimen. with this protocol, the average time of starting hbot dropped from (prior to ) to . days. the median onset of hc was days post hsct. all patients were receiving immunosuppressive treatment at the onset of bk-hc. all patients suffered from macrohematuria with blood clots (grade iii cystitis), ( . %) experienced severe dysuria and ( . %) urgency. bk viruria was present in all patients, and concurrent bk viremia was detected in % of those who were tested. patients reported symptomatic improvement at a mean of . days following the initiation of hbot. no patient experienced serious adverse effects due to hbot, but two patients required insertion of tympanic ventilation tubes. eleven of our patients ( . %) experienced complete remission of bk-hc following hbot, with an overall response rate of . % ( / patients).eight of our patients ( . %) eventually succumbed due to either hsct complications or disease relapse. conclusions: hyperbaric oxygen therapy is a safe, effective, non-invasive and well tolerated treatment modality for bk -hc and should be considered for first line therapy for this complication of hsct. clinical background: every year, almost one thousand cases of hematological malignancies in pediatric population are reported in peru. allogeneic hematopoietic stem cell transplantation (allo-hsct) is an alternative strategy in many of these cases. only between - % of the pediatric population that requires a hsct has a compatible human leukocyte antigen (hla) donor. the remaining % have to access international donor registries, extending the awaiting time and conditioning the progress of the disease. allo-hsct has the potential to help children with several hematological disorders with non-compatible hla donor. hla genotypically identical sibling donors are the best option when pursuing an hsct. nevertheless, patients' alternative sources of stem cells could be obtained from an haploidentical donor like one of their parents. the haploidentical transplantation program with macs was implemented in peru in to reduce the risk of graft-versus-host disease (gvhd), support the immune system reconstitution and to expand pool of donors. it allows patients to access a treatment that is efficient and safe, as shown in the depletion of positive tcrα/β+ and b cells procedures for allo-hsct. methods: the mobilized leukapheresis products (n= ) of haploidentical healthy donor was washed to remove platelets and preparations were performed according to miltenyi's clinimacs® manual for tcrα/β+ and cd + cell depletion. analysis of the initial leukapheresis product and tcrα/β + and cd + depleted graft (target and non-target product) was performed using flow cytometer. cells were analyzed for cd +, cd +, -aad, cd +, tcrα/β+, tcrγ/δ +, cd + and cd + with fluorochrome-labeled antibodies from miltenyi biotec using a novocyte cytometer. the results obtained with the ex vivo t-cell depleted allo-hsct procedures show an overall survival (os) over % with an ic % at the end of the first year with low incidence of gvhd. macs of tcrα/β+ and cd + cells are effective (logp . and logp . , respectively) obtaining minimum levels of depleted lymphocytes within clinical established parameters for diverse pathologies. in addition, tcrα/β+ and cd + cell depletion does not significantly affect hematopoietic stem cell populations such as cd + and cd + cells or tcrγ/δ+ cell population. cd + and tcr γ/δ cells are highly recovered ( . % and . , respectively), which contributes with a better engraftment after allo-hsct. conclusions: peruvian results oscillated within european ranges with an os over %. our data suggests that the macs method is an efficient, effective and safe strategy for haploidentical hsct which has a remarkable cost-benefit ratio and makes it viable in countries of the latin american region with peruvian socio-economic characteristics. evaluation of genoresistance for viral reactivation treatment has been implemented as a strategy to improve os. better results are achieved in patients after allo-hsct with the validation of these tests in peru. preliminary pharmacoeconomic evaluations allow us to establish magnetic activated cell sorting (macs) as a promissory strategy compared to other alternatives for haploidentical hsct. it is necessary to increase the number of procedures in order to confirm efficacy and safety of macs in a larger population. disclosure: all authors have no conflicts of interest. abstract already published. micro-costing study of hematopoietic stem cell transplantation in two hospital institutions from southern of brazil background: hematopoietic stem cell transplantation (hsct) is a potentially curative treatment indicated for patients with onco-hematological, hereditary and immunological diseases. considering the increase of patients indicated to the hsct and the lack of knowledge about the costs resulting from this treatment, is important to identify and detail the resources consumed in each phase of hsct and provide knowledge to the public health brazilian system. we aimed measure the total cost of related hsct, based on micro-costing study of patients assisted in two hospitals in the south region of brazil. methods: hsct costs were estimated using the timedriven activity based costing method (tdabc), which measured cost of services / products based on actual consumption of resources. we collected data from medical records of patients submitted to allogeneic hsct in from public and private (philanthropic) hospitals. we interviewed professionals involved in the tcth activities, we performed chrono-analysis and, we consulted financial and administrative systems reports of hospitals. in order to compare costs according to clinical complications observed in patients, we grouped into two ranges of complexity: low/ medium and high. the study was divided into stages: hsct processes mapping; costs measurement; and analysis of results. finally, the costs were compared: by activity, by resource and by hospital. this study was financed by psid-uhs, by an agreement signed between ministry of health and moinhos de vento hospital, through adjustment term number: / , and approved by research ethics committees. results: from the hsct processes mapping, the following steps were defined: (i) hospitalization; (ii) conditioning; (iii) transplantation; (iv) period of aplasia; (v) engraftment; (vi) observation; (vii) pre-and medical discharge. seven patients were classified in low / medium complexity level, with hospitalization median time of days and an median cost of usd , . , whereas the other five patients, classified as high complexity, presented median time of days and median cost of usd , . . the hsct costs evaluation identified that steps ii and iv presented greatest cost in high complexity patients. lower complexity patients presented, in steps ii and iv, median costs of usd , . while in higher complexity usd , . . in addition, median costs of materials and drugs were usd , . and usd , . in lower and higher complexity patients. conclusions: tdabc method allowed the identification of the moment when patients consume the most resources. of all the hsct stages, periods of conditioning and aplasia presented higher costs, representing . % of the total hospitalization value. in these stages, higher complexity patients presented three times higher the median cost. the resources that had the greatest impact were medicines and medical materials, costing times more than lower complexity patients. conclusion: this study allowed a detailed identification of the hsct costs in patients with different complexity ranges in two hospitals from southern brazil. therefore, the identification of service demand regarding the clinical complexity, allows the generation of important information for the management of the best care in the health service. disclosure background: immunodeficiency due to lrba deficiency is characterized by hypogammaglobulinemia and autoimmunity. hemolytic anemia, lymphadenopathy, autoimmune hepatitis and, above all, autoimmune enteropathy are the fundamental characteristics of these patients together with the history of recurrent invasive infections.the therapy includes immunosuppressants, endovenous immunoglobulins. bone marrow transplantation is the final therapy of these patients, especially in the most serious cases and in recent years, also on the light of increasingly targeted conditioning regimes, it is associated with ever better prognosis. methods: we present the case of caterina, an -year-old patient with lrba deficiency, diagnosed at years of age for a history of hypogammaglobulinemia, recurring invasive, bacterial and fungal infections and a picture of autoimmunity represented by ahia, myelitis (c -c ), autoimmune hepatitis and enteropathy in treatment with abatacept and sirolimus. it also presents leptin deficiency lipodystrophy.she presented to our observation for ostemielitis in multiple outbreaks (tibia, femur and left knee) secondary to sepsis from mrsa. broad spectrum antibiotic therapy and curettage surgery were performed during hospitalization.after months of broad-spectrum antibiotic therapy (daptomycin, rifampicin, ceftaroline, cotrimoxazole, levofloxacin, dalbavancin) there was resolution of the infections.because of the seriousness of the disease it is therefore decided to subject catherine to hematopoietic stem cell transplantation. results: therefore were performed bone marrow transplant from mud after conditioning at reduced intensity, delayed in days, with treosulfan, fludarabine and thiotepa. prophylaxis for gvhd was performed with atg ( mg / kg / day), sirolimus (already practiced for enteropathy) and mmf. because of the hepatic picture, we also performed prophylaxis for vod with defibrotide for days. transplantation was performed by peripheral stem cells with x ^ cd / kg and x ^ cd / kg.the patient had always presented good general conditions with engrafment of the pmn to the d + and the plt to the d + . the chimerism at d + was % donor both on pmn and on pbl. prophylaxis for gvhd was changed on d + by replacing the sirolimus with tacrolimus for the appearance of grade i cutaneous gvhd . conclusions: we have successfully performed bone marrow transplantation in patients with lrba deficiency. the new antibiotic molecules, used to induce infectious remission, the new low-intensity regimens, the prevention of the most fearful complications (vod) have been the key to success in such complicated case. the high number of cd cells infused with a controlled number of cd were the key then of rapid engraftment with minimal gvhd readily controlled by the immunosuppressant. disclosure background: in the absence of hla-matched related donor, allogeneic stem cell transplantation from haploidentical donors are potential alternatives for patients with hematological malignencies with an indication to allogeneic stem cell transplantation. herein, we retrospectively assessed the outcome of haplo-sct for patients with refractory hematological malignancies. methods: this analysis included consecutive patients who underwent haplo-sct for various hematological malignancies at our center between october and may . we used our institutional database to evaluate details and characteristics of patients and transplant outcomes. results: demographic features of the patients and donors have been summarized in table . all of the patients had advanced disease with a high risk of relapse. the majority of patients underwent haplo-sct from their parents. out of patients, early transplant-related mortality was seen in this cohort of patients. four patients treated with second haplo-sct and recovered hematopoiesis after second transplant. the remaining patients were followed in a median of months. donor type abo group switch was observed in a median of days ( - days) after transplant. the median time for engraftment was days (range, - ) for all patients. after the first transplant, patients developed acute gvhd ( . %) with patients having grade ii-iii acute gvhd. five ( . %) had chronic gvhd, none of them with extensive manifestation. the prepative regimen was relatively well tolerated with limited regimen-related toxicity. cmv reactivation occurred in patients ( . %) during the follow-up of the study. eight patients ( . %) relapsed after a median of days post transplant (range, - days). cr was achieved in ( %) patients after haplo-sct. mean estimated -year os and pfs are . %± . % and . %± . %, respectively. conclusions: given the growing data on the similarity of outcomes after hla-matched and haploidentical sct, further studies are required to determine whether factors may be more important for donor selection than hlamatching. clinical trial registry: -disclosure: nothing to declare outcome of allogeneic stem cell transplantation for hodgkin and non-hodgkin lymphoma: single center experince from turkey ayşe uysal , hale bülbül , nur akad soyer , mahmut tobu , murat tombuloglu , guray saydam , filiz vural background: allogeneic sct (allosct) is generally optionally treatment choice for young and fit patients with relapsed/refractory lymphoma who were heavily pre-treated and after the failure of autologous stem cell transplantation (asct). relapse after asct is associated with a poor prognosis and allosct is a potentially curative therapy for lymphomas which have relapsed after asct. methods: in this study, we evaluated patients with hl and nhl who had treated with allo-hsct between november and december in ege university adult hematology transplantation unit. results: patients, disease and transplant characteristics were illustrated in table. histologic subtype of nhl was evaluated as t cell lymphoma (n= ; , %), mantle cell (n= ; , %), diffuse large b-cell lymphoma (n= ; , %) and b-cell lymphoma, unclassifiable, with features intermediate between dlbcl and classical hodgkin lymphoma (n= ; , %). all histologic subtype of hl was determined as nodular sclerosing. the median number of prior treatments before allo-hsct was (range, - ). twelve ( , %) patients had refractory disease, ( , %) patients were in complete remission and ( , %) patients were in partial remission before allo-hsct. the median time from diagnosis to allosct was (range, - ) months. peripheral stem cell was used for stem cell source in all of them. total body irritation plus fludarabine plus cyclophosphamide and busulfan plus cyclophosphamide were preferred most frequently for conditioning as non-myeloablative and myeloablative, respectively. neutrophil engraftment was occurred median of (range, - ) days. graft versus host disease (gvhd) prophylaxis was applied all of them and cyclosporine plus methotrexate was preferred most frequently (n= ; , %). gvhd was occurred in , % of them ( , % acute gvhd, , % chronic gvhd and , % both). veno-occlusive disease (vod) was occurred in ( , %) patients. transplant related death was observed in ( , %) patients. overall survival (os) and disease-free-survival (dfs) were evaluated as median (range, - ) and (range, - ) months, respectively. analyze of os and dfs was illustrated in figure. six patients are alive without disease. after a rapid tapering of immunosuopressive therapy was underwent a therapy with ponatinibat dose of mg/die results: afther a month of therapy we observed a rapid decrease in minimal residual disease on molecular assessment with an mmr of p -bcr-abl/abl non detectable confirmed by bone marrow revaluations at days + , + nd + after the salvage therapy. the patient has not had experienced of graft-versus-host disease, ponatinib treatment was well tolerated and considered safe with easily manageable side. conclusions: maybe in the era of tyrosine kinase inhibitors (tkis), philadelphia chromosome positive acute lymphoblastic leukemia (ph+ all) it could benefit from a combined treatment between transpalnt and tkis however more studies are needed to confirm these hypotheses. disclosure: nothing to declare immunodeficiency diseases and macrophage background: although a number of patients with hiv infection and hematological disease have successfully undergone allogeneic hsct together with combination anti-retroviral therapy (cart), short and long-term outcomes remain not well known. we report the largest spanish experience of hiv-infected adult patients with high-risk hematological malignancies with allogeneic hsct. methods: we retrospectively reviewed hiv-positive patients who received allogeneic hsct between and in spanish centers within geth (grupo español de trasplante hematopoyético y terapia celular). results: baseline and transplant characteristics of patients are shown in table . median age was years and % of the patients were men. the most frequent underlying malignancies were non-hodgkin lymphoma ( , %) and aml ( , %). in half of the patients an hlaidentical sibling was the donor; and in the other half, an alternative donor was used. peripheral blood was used as graft source in % of the transplants. at the time of hsct, all patients had been receiving suppressive cart for a median of years and only of them showed detectable plasma hiv rna, one of them because of poor adherence to cart together with the accumulation of multiple resistance mutations; and the other patient had detectable hiv rna at low levels (< copies/ml). all patients received cart throughout the transplant procedure, being temporally stopped in two patients due to significant mucositis. after a median follow-up of months ( - ), -year overall survival (os) and event-free survival (efs) were %. nrm was % at months and relapse was % at months. grade ii-iv agvhd rate was %, and moderate/severe cgvhd rate was % at months. a significant proportion of patients ( %) showed infectious complications with viral infections as the most frequent cause. two patients had invasive aspergillosis and one patient presented disseminated tuberculosis. causes of death included infections ( %), relapse ( %) and toxicity ( %). among the patients who died due to infections, had severe chronic gvhd and were under immunosuppressive therapy. two patients showed severe toxicity related to drug interaction with anti-retroviral therapy. all survivors except one showed undetectable hiv load at last follow-up after hsct. conclusions: allogeneic hsct is an effective therapy for high-risk hematological malignancies in patients with hiv infection, providing long-term disease free survival together to long-term hiv suppression with cart. however, drug interactions with anti-retroviral agents, occurrence of gvhd, and frequent infectious complications account for a complex procedure in this population. selected hiv-infected patients with hematological malignancies should be considered for allo-hsct when indicated, in experienced centers, with a multidisciplinary care. disclosure background: primary immunodeficiencies (pid) are rare diseases often associated with genetic defects in the immune system, predisposing individuals to recurrent infections and increased risk of allergy, autoimmunity and malignancy. allogeneic haematopoietic stem cell transplantation (hsct) has been successfully used as a curative therapy for most severe forms of pid. because pid is a genetic disease, < % of these children will have a healthy, human leukocyte antigen (hla) matched sibling donor available, and umbilical cord blood grafts from unrelated donors are a suitable alternative cell source. we report the results of umbilical cord blood transplantation (ucbt) performed in patients with pid between and at children's hospital of fudan university in china. methods: patients included chronic granulomatous disease (cgd, n= ), severe combined immunodeficiency (scid, n= ), interleukin- receptor-a deficiency (il- rad, n= ), wiskott-aldrich syndrome (was, n= ), leukocyte adhesion deficiency (lad, n= ), severe congenital neutropaenia (scn, n= ) and other immunodeficiencies (n= ). all patients were assessed by clinical immunologist to confirm clinical phenotype and genetic diagnosis. median age of patients was months (range, to months), and median body weight was . kg (range, . to kg). all patients received a ≤ / hla alleles-mismatched cord blood unit, were hla fully matched, were / matched, were / matched and were / matched. median nucleated cells of the cord blood were . x /kg (range, . to . x /kg), and median cd + cells were . x /kg (range, . to . x /kg). results: median follow-up time was months (range, to months), the overall survival rate at year for all patients was . %, and was . %, . % and % for cgd, scid and il- rad, respectively. patients died, most deaths ( / , . %) occurred in + days after transplantation, the main cause of death was infection ( / , . %). / ( . %) patients engrafted, median time of neutrophil engraftment was days (range, to d), and median time of platelet engraftment was days (range, to d). the cumulative incidence of grade - acute gvhd was . %, and that of chronic gvhd was . %. conclusions: unrelated ucbt should be considered for pid patients without an hla -matched sibling donor. effective control of infection before and after transplantation is important for improving survival. disclosure background: dedicator of cytokinesis (dock ) deficiency causes a combined immune deficiency characterised by recurrent bacterial infections, susceptibility to viral infection, eczema, food allergies, vasculitis and increased risk of malignancy. due to the high morbidity and mortality of the disease hsct has been increasingly offered to patients as a potentially curative therapy . methods: we retrospectively reviewed the outcomes of hsct for patients with dock deficiency at great north children's hospital newcastle upon tyne between and ( in published reference). results: ten patients with dock deficiency were treated with hsct (median age . y range . - . y). median duration of follow up was . years (range . - . y). there were a range of donor sources ( msd, mud and tcr ab/cd + depleted haploidentical), conditioning regimens ( treo-flu, treo-flu-thiotepa) and serotherapy ( alemtuzumab, atg+rituximab, none). one patient who received a cd + tcr alpha beta/cd + depleted haploidentical transplant received add back t-cells with caspacide molecular safety switch (bellicum pharmaceuticals). skin only agvhd occurred in / patients ( x stage , x stage ). no patients had cgvhd. overall survival was % ( / ). survival was comparable regardless of donor source. all deaths occurred within months of transplant. the patients who died had significant burden of disease pre-transplant: patient had chronic liver failure secondary to cryptosporidial sclerosing cholangitis, had a cirrhotic liver secondary to cryptosporidium, cerebral vasculitis, an axillary aneurysm and aortic vasculitis requiring grafting of an ascending aortic aneurysm, was pn dependent for failure to thrive with a history of cryptosporidium infection and had candida in a bal pre-transplant. causes of death in these patients were: respiratory failure (n= ), progressive encephalopathy (n= ), multi-organ failure with septic shock and encephalopathy (n= ) and multiorgan failure and septic shock after treatment for tma (n= ). two of these patients had reactivation of cryptosporidium prior to their death. pretransplant cryptosporidium was associated with mortality (graph ). one patient who survived had suffered from stroke pretransplant. one suffered from a basilar artery aneurysm years post-transplant at yo. at the time of latest follow up donor chimerism was % in / survivors and high level mixed in the other( % cd , % cd and % cd ). conclusions: this single centre study of hsct for patients is consistent with literature indicating that hsct is a potentially curative therapy for patients with dock deficiency. the increased morbidity associated with cryptosporidial infection is likely to be a consequence of overall disease burden rather than an infection specific effect. this does however highlight the improved outcomes of transplant prior to development of multiple comorbidities and suggests that hsct should be considered early. it is unclear whether the late occurrence of vascular complications after transplant were caused by a manifestation of disease which is not corrected by transplant or a result of vascular injury sustained pre-transplant. reference methods: referred infants underwent testing for: immune phenotype (ab, gd, naïve and memory t cell, b cell and nk cell numbers); functional activity of t and nk cells; maternal engraftment; adenosine deaminase (ada) and purine nucleoside phosphorylase (pnp) enzyme activity; and genetic testing. those with a confirmed diagnosis of scid underwent either allogeneic hematopoietic stem cell transplant (hct) or (if eligible) gene therapy (gt). infants identified as having ada deficiency as the etiology of their scid received enzyme replacement therapy prior to proceeding to definitive therapy. results: twenty-three ( %) infants were confirmed to have scid. three ( %) of these infants had a family history of scid but would not have been identified without nbs. in addition, one infant, born prematurely at weeks, was diagnosed as having pnp deficiency only after developing infections. this infant was identified by nbs but repeat testing at weeks gestation was normal likely due to support of transient t cell production from exogenous enzyme provided by red cell transfusion. twelve infants with confirmed low trecs had a non-scid diagnosis: with transient lymphopenia of infancy who normalized trec, immune phenotype and function, with prenatal exposure to -mecaptopurine ( -mp), genetically confirmed with digeorge syndrome, and with prolonged lymphopenia. of the three with prolonged lymphopenia, two had recurrent infections: one ultimately diagnosed with ataxia telangiectasia and one with absent trec but near normal number of t cells, normal pha but no specific antigen responses, and absent b cells who will be undergoing transplant in the near future. the third continues with absent trec, short telomeres, low numbers of a/b t cells, presence of g/d t cells, vaccine responses and freedom from infection with no identified genetic etiology. in summary, % of the patients referred to msk with confirmed abnormal nbs for scid have a non-scid diagnosis. there is no uniform collection of data for these infants and the threshold trigger for repeat testing varies from state to state, so the incidence of significant non-scid disorders identified will also likely vary from state to state. although our institution specific experience is biased, as most infants had confirmation of a low number of trec prior to referral, the significant number of disorders in the non-scid cohort emphasizes the importance of full evaluations and follow-up for these infants. disclosure: none of these relate to the work being presented. susan prockop -research funding mesoblast and atara biotherapeutics. nancy kernan -research funding jaz pharmaceuticals. richard o´reilly research funding and royalties atara biotherapeutics. kevin curran consulting juno pharmaceuticals, novartis. j.j. boelens avrobio, magenta, chimerix and bluebird bio background: post-transplant autoimmune cytopenia (aic) is challenging and associated with substantial morbidity and mortality. we aimed to study the cumulative incidence (ci) of post-hct autoimmune cytopenia (aic) and its predictors in a cohort of children with primary immunodeficiency (pid). methods: in this retrospective study, we included children with pid who underwent their first hsct with fludarabine(f)-treosulfan(t)±thiotepa(thio) at great north children's hospital from - . main outcomes of interest were the ci of aic and its predictors. fine-and-grey regression models were used to analyse predictors of aic, considering death as a competing event. variables included were age at transplant (< . years vs > . years), gender, diagnosis (scid vs immune-dysregulatory disorders vs other pids), pre-transplant aic, pre-and posttransplant respiratory virus, donor (mfd vs mud vs mmfd/mmud vs haploidentical donor), abo incompatibility, conditioning (ft vs ftthio), serotherapy (none vs alemtuzumab . - . mg/kg vs alemtuzumab . - . mg/kg vs atg), stem cell source (marrow vs pbsc vs cord vs exvivo t depleted pbsc), infused stem cell doses (tnc, cd and cd ), agvhd (none vs any agvhd), cgvhd (none vs any cgvhd), viral infections (cmv/adenovirus/ ebv/hhv viraemia), chimerism (full vs mixed chimerism (wb < %) within first year post-hct). impact of thymopoiesis using naïve t cell recovery was studied. results: median age at transplant was . years (range, . - . years). primary diagnoses were scid ( %), immune-dysregulatory disorders ( %) and other pids ( %). donors were mfd ( %), mud ( %), mmfd/ mmud ( %) and haploidentical parents ( %). stem cell sources were marrow ( %), unmanipulated pbsc ( %), ex-vivo t-depleted pbsc ( %) and cb ( %). % received additional thiotepa and % had csa/mmf as gvhd prophylaxis. median duration of follow-up of survivors was . years (range . to . years). -year os for the entire cohort was %. -month and -year ci of aic were % and %. of developed aic, ( %) had aiha, ( %) had aiha±itp and ( %) had aiha ±itp±ain. median onset of aic was . months post-hct (range . - . months). patients were treated with a median of treatment modalities (range, - ). one ( %) had steroid, ( %) had steroid+high-dose-ivig, ( %) had steroid+high-dose-ivig+rituximab, ( %) had steroid +high-dose-ivig+sirolimus and ( %) had steroid +high-dose-ivig+rituximab+sirolimus. the median time to resolution in ( %) who achieved remission after first aic was . months (range . - . months). had one relapse and had two relapses. died after development of aic ( aspergillus pneumonia; multi-organ failure). of ( %) surviving patients after aic, had on-going aiha at median of follow-up . years post-hct (range . - . years). on univariate completing-risk analysis, age at transplant > . years (p= . ) and pre-transplant aic (p= . ) were associated with higher incidence of aic (figure a -ic). on fine-and-grey models, only age at transplant (hr . , %ci . - . , p= . ) was independently associated with aic. of with complete immune reconstitution data, naïve t cells > cells/ml at months post-hct was associated lower incidence of aic (hr . , %ci . - . , p= . ) (figure d) conclusions: younger age and thymopoiesis were associated lower incidence of aic in children with pid after hct clinical background: human heme-oxygenase- (ho- ) deficiency has been reported to present with tetrad of anemia, nephritis, inflammation and asplenia and is fatal if not treated. its an auto-inflammatory disorder. macrophages/ monocytes express ho- and are engaged in recycling of red cells. human ho- deficiency results in intravascular hemolysis and severe damage to the endothelial system, kidneys, and other organs. transplantation of either healthy wild type macrophages or new macrophages produced by sct from healthy donor has been proven to be curative for ho- deficiency in mice. in , we had reported first successful allogeneic sct for human ho- deficiency. here we report second successful non-myeloablative msd hsct for a child with ho- deficiency. methods: a -yr-old-girl presented with complaints of fever, anemia and severe hypertension. in the past, at the age of years she was admitted for high fever for month and needed blood transfusion for the first time for severe anemia and had high platelets and high ferritin. she was treated as macrophage activation syndrome with prednisolone alone and later cyclosporine was added. she had short stature, abnormal facies but normal development. hemoglobin g/dl, urine for haemoglobin was positive, platelets , /ul, ferritin mcg/l and urine albumin + and urine rbc - /hpf. ultrasound and ct scan abdomen showed asplenia. a diagnosis of ho- deficiency was suspected. mutation analysis showed homozygous missense mutations in exon (r x) on chromosome q , which would result in the absence of the functional ho- protein. both parents were carriers of this mutation. we managed her over next -years with prednisolone, hydroxyurea and mycophenolate mofetil (mmf). however she remained steroid dependent. hla-typing confirmed her healthy unaffected -year-old brother to be a fully matched donor. at the age of years she was taken up for msd sct after taking informed consent. she weighed just kg. we conditioned her with alemtuzumab- . mg/kg, fludarabine- mg/m , cyclophospamide- mg/kg and total body irradiation gray. we infused million/kg peripheral blood stem cells from her brother. graft-vs.-host disease (gvhd) prophylaxis consisted of tacrolimus & mmf. results: she tolerated procedure very well. her entire hospital stay was uneventful and lowest platelet count recorded was , /ul. her neutrophils engrafted on day + and she was discharged on day+ . his urine albumin was nil by day+ . she had no gvhd. her chimerism on day+ showed % donor cells, on day+ was % and on day+ was % donor. now he is day+ post-sct and doing well. she has no evidence of hemolysis, proteinuria, hypertension, fever. she has normal ferritin and platelets. she has gained cm height and kg weight in last months. she had no viral reactivation and her immune recovery at months post sct is good. conclusions: non-myeoablative allogeneic msd sct is a curative treatment option for human ho- deficiency. disclosure: nil two decades of excellent transplant survival in children with chronic granulomatous disease: a report from a supraregional immunology transplant centre in europe background: haematopoietic stem cell transplantation (hsct) confers life-long curative therapy for chronic granulomatous disease (cgd). the ability of donor-derived neutrophils to replace recipient's defective neutrophils makes hsct a superior therapy compared to conventional standard of care using antimicrobial therapy. methods: we examined the outcome of children with cgd who received a first hsct at great north children's hospital from to . outcomes included overall survival (os), event-free survival (es), toxicity endpoints, autoimmune disease, long-term survival and graft function. cox proportional-hazard models were used to analyse predictors of os and es. variables included for predictor analysis were age at transplant, donor, stem cell source, stem cell doses and conditioning. results: = children were included in this analysis. median age of transplant was . years (range, . - . years). ( %) had x-linked and ( %) autosomal recessive cgd. twenty ( %) had matched family donor, ( %) had unrelated donor and ( %) had parental haploidentical donor. prior to , various conditioning regimens were used, with ( %) patients undergoing conditioning with pharmacokinetic guided intravenous (iv) busulfan (bu) and iv cyclophosphamide with or without serotherapy. from , the conditioning regimen was switched to flu-treosulfan-alemtuzumab with gvhd prophylaxis using ciclosporin (csa) and mycophenolate mofetil (mmf) for family and unrelated donors (n= , %). flu-treosulfan-thiotepa-atg-rituximab was used for cd tcr alpha-beta cd depleted haploidentical grafts (n= , %). ten ( %) patients had grade ii-iv acute gvhd while had ( %) had grade iii-iv acute gvhd. none had chronic gvhd. the -year os for the entire cohort was % ( % ci, - %) (figure ). analysis by age at transplant revealed a -year os of % for children transplanted at < = years of age and % ( %ci, - %) for the children > years of age (p< . ) (figure ) . the os was comparable between match family donor ( %, % ci, - %) and unrelated donor transplant ( %, - %) ( figure ). all four haploidentical transplants were successful. the -year es for the entire cohort was % ( % ci - %). none of the variables was associated with es. all seven patients with slipping chimerism received a successful second transplant. the five deaths were all due to transplantrelated complications ( multi-organ failures; pulmonary haemorrhage; graft iv acute gvhd; post-transplant lymphoproliferative disease). the median age at transplant of deceased patients was . years (range . to years). the -year and -year cumulative incidence of autoimmune diseases were % and % respectively. three ( %) had immune cytopenia while ( %) had autoimmune endocrinopathy ( thyroid dysfunction; type diabetes mellitus). the median age of long-term survivors was years (range, to years) with the median duration of follow-up of . years (range, . to . years). there was no late death in the entire cohort. the median donor myeloid chimerism was % (range to %) conclusions: despite the limitations of a single centre study, our findings confirm that hsct is a safe and longlasting curative therapy for children with cgd disclosure: none non -medical challenges in the diagnosis and transplantation of patients with primary immune deficiency: an experience from a tertiary care center in india sagar bhattad , stalin ramprakash , raghuram cp , chetan ginigeri , fulvio porta , background: primary immune deficiencies (pid) are increasingly being recognized in several parts of india. despite being diagnosed, many patients fail to receive optimal care due to financial and social constraints. methods: case records of patients diagnosed and treated (including hematopoietic stem cell transplants) for pid diseases during feb -nov at aster cmi hospital, bangalore, india were analysed. factors leading to deferred or suboptimal care were assessed in detail. results: patients with various pids were diagnosed during the study period (details in table). of them warranted a hematopoietic stem cell transplant (hsct) as definitive curative treatment. a total of children received hsct. of them died while of them are alive and well. children ( with severe combined immune deficiency) died before a hsct could be carried out. of them were critically ill at presentation, while were stable but deferred further treatment citing financial and social constraints. children needing transplant continue to remain on follow-up and have not been transplanted to date ( of them have significant financial constraints, families are not convinced about the need for transplant and of them are being prepared for transplant). table: (scid -severe combined immune deficiency, vodi-veno-occlusive disease with immunodeficiency, cgd -chronic granulomatous disease, hlh -hemophagolymphohistiocytosis, was -wiskott aldrich syndrome, xlt -x linked thrombocytopenia, lad-leukocyte adhesion deficiency, msmd -mendelian susceptibility to mycobacterial disease, xla -x linked agammaglobulinemia, cvid -common variable immune deficiency, apeced -autoimmune polyendocrinopathy candidiasis ectodermal dystrophy, cmcc -chronic mucocutaneous candidiasis, at -ataxia telangiectasia). conclusions: we present our experience from a developing country and discuss non-medical factors leading to suboptimal care in children with pid. only % children warranting hsct could be transplanted in our cohort. among those where hsct is potentially curative % of children died before hsct could be offered. transplants in developing countries pose unique challenges due to the absence of government funding and/or universal insurance coverage. in addition to delay in diagnosis and critical state of patients at admission, financial and social factors significantly contributed to poor outcome. disclosure: none the outcome of hematopoietic stem cell transplantation (hsct) in pediatric patients with hemophagocytic lymphohistiocytosis (hlh) in korea methods: the korea histiocytosis working party retrospectively collected nation-wide data from the patients diagnosed with hlh and underwent allogeneic hsct between and . the clinical characteristics and treatment outcomes of the patients were analyzed. results: a total of patients were enrolled. there were patients with fhl ( fhl , fhl , and fhl ), infection associated hlh, and secondary hlh of unknown cause. all the patients were treated with hlh- protocol, and patients achieved complete response (cr) after treatment for weeks, while did not. the main reasons for receiving transplantation were fhl in , reactivation in , and refractory disease in . the conditioning regimens were busulfan-based in patients, fludarabine-based in , treosulfan-based in , and busulfan/fludarabine-based in . stem cell sources used for hsct were from peripheral blood in patient, cord blood in , and bone marrow in . the donor types of hsct were unrelated donor in patients and related in ( matched sibling donor, haploidentical donor, partially matched donor). the causes of death of patients were disease reactivation/ progression in , acute gvhd with/without vod in , and graft failure in . five year overall survival rates were . %, respectively. the disease status at the time of hsct was cr in patients, and non-cr in . the -year survival rate of patients who received hsct in cr was % and % for patients transplanted while in non-cr status (p= . ). patients who received hsct using peripheral blood stem cells had a better -year survival rate of % compared to % of patients who received cord blood stem cells, significantly. the presence of neurologic symptoms, disease status after intial week therapy, conditioning regimen, and cd positive cell count did not have statistically significant impact on survival. conclusions: hsct improved the survival of patients who had familial, or relapsed, or refractory hlh in the korean nation-wide hlh registry. these results are similar to other reports in the literature. the disease status at the time of hsct and the stem cell source of the transplant were the important prognostic factors that affected the survivals of the hlh patients who underwent hsct. clinical trial registry: no registry number. disclosure: to the best of our knowledge, the named authors have no conflict of interest, financial or otherwise p hematopoietic cell transplantation with reduced intensity conditioning regimen using fludarabine/ busulfan and fludarabine/melphalan for primary immunodeficiency diseases background: primary immunodeficiency disease (pid) is congenital disorders of innate or acquired immune system. hematopoietic cell transplantation (hct) was a treatment option for pids with life-threatening infections or immune dysregulations. reduced intensity conditioning (ric) was increasingly used to prevent complications in hct, but optimized regimens have not been established. we performed hct for pids with ric using fludarabine and busulfan (flubu) or melphalan (flumel) according to the guidelines of european society for immunodeficiencies (esid) / european society for blood and marrow transplantation (ebmt), and assessed the efficacy and safety of these ric. methods: from april to december , pid patients underwent ric-hct using flubu or flumel in tmdu were analyzed retrospectively. the auc of bu was set to mg*hour/l for severe combined immunodeficiency disease (scid) and mg*hour/l for non-scid. overall survival (os) was analyzed. results: the median age at hct was . ( . - ) years old ( male and female patients). flubu was used for patients ( scid, combined immunodeficiency disease (cid), ectodermal dysplasia (eda), and severe congenital neutropenia (scn)) and flumel was used for patients ( scid, cid, xiap deficiency, and eda). anti-thymocyte globulin was used in patients of flubu group and patients of flumel group. cord blood in and bone marrow in was used for donor sources. matched donor was used for and patients in flubu and flumel groups ( . % and . %), respectively. median follow up period was . years. two years-os of all patients, flubu group patients and flumel group patients was . %, . % and . %, respectively. neutrophil engraftment was . %, . % and . % (all patients, flubu group and flumel group). in scid, all patients in flubu group achieved engraftment and survived. seven out of in flumel group achieved engraftment, but patient had secondary graft failure and patients died. in non-scid, out of in flubu group achieved engraftment, but patients had secondary graft failure. all non-scid patients in flumel group were engrafted and survived. two and patients in flubu group and flumel group suffered from severe acute graft-versus-host disease (grade iii-iv). ten patients had hemophagocytic lymphohistiocytosis (hlh). viral reactivation or infection was observed in patients, and resolved in all but one patient. conclusions: the ric-hct using flubu or flumel was advantagous for neutrophil engraftment, and flubu for scid and flumel for cid with immune dysregulation may be an effective opinion. flubu regimen needs to be improved for secondary graft failure in non-scid. prevention of hlh after transplantation using dexamethasone palmitate will be considered. disclosure: nothing to declare. survival after hematopoietic stem cell transplantation with tcrαβ/cd graft depletion in older children with primary immunodeficiencies background: tcrαβ/cd depletion is a graft engineering method that proved valuable in increasing the survival rate after hematopoietic stem cell transplantation (hsct) in patients with primary immunodeficiencies (pid). decreased survival rate in older patients with pid was previously reported after transplantations with nonmanipulated grafts. methods: patients with various pid (excluding classic scid) who received allogenic hsct with tcrαβ/ cd graft depletion from to september in our center were analyzed. the median age at hsct was , years (range , - , ). patients were divided into age groups: - years - patients, - years - , - years - . patients received hsct from matched unrelated, -haploidentical donors, -siblings. conditioning regimens with - alkylating agents and antithymocyte globulin were used in all patients. patient received short courses of various posttransplant immunosuppressants. median follow up after hsct is , years (range , - , years). results: overall survival (os) in patients was , ( % ci , - , ). we observed similar os in the younger age groups: , ( % ci , - , ) in - years and , ( % ci , - , ) in - years of age. seven of patients in older group ( - years of age) died, the os was only , ( % ci , - , ), p= , . all patients from older age group who died had combined pid: -wiscott-aldrich syndrome, -undefined pid, -il rg deficiency, -dclre c deficiency, -nijmegen breakage syndrome (nbs), -kabuki syndrome, -icf syndrome. the median time of death after hsct was , years (range , - , ). six of those had transplant-related mortality (trm). five patients had hsct-associated viral infections: -cmv pneumonias, -adv infections ( -fulminant hepatitis, -multiorgan). interestingly, of them had prolonged history of disseminated viral infections (adv), with the reduction of viral load in blood and other fluids and tissues upon treatment. one patient with kabuki syndrome after hsct developed hhv associated kaposi sarcoma, was successfully treated. eventually all patients with reduction of viral infection and sarcoma symptoms developed multiorgan failure with some clinical and laboratory evidences of endothelium cell damage syndrome. one patient with nbs died of high grade lymphoma progression. conclusions: hsct with tcrαβ/cd depletion demonstrates high survival rate in patients with various pid. in our group patients' age older than years predisposes to decreased posttransplant survival. patients with combined pid are at higher risks of posttransplant mortality. we conclude that at least in some of our patients with prolonged history of viral infections after hsct the cause of death could be multiorgan failure due to endothelium cell damage syndrome resulting from persistent inflammation and drug toxicity effects. disclosure background: chronic granulomatous disease (cgd) is a primary immunodeficiency (pid) caused by a mutation in of the subunits of the nicotinamide dinucleotide phosphate (nadph) oxidase, which leads to a reduction in the microbicidal activity of phagocytic cell. starting at an early age, cgd patients suffer from severe recurrent infections, as well as inflammatory events. allogeneic hematopoietic stem cell transplantation (hsct) is a curative option for cgd in patients with insufficient benefit from supportive care and prophylactic antibiotics. we reported a series of patients with cgd who underwent unrelated umbilical cord blood transplantation (ucbt) at our center. methods: in this retrospective study, we observed a series of consecutive ucbt performed at our center in children with cgd between and .median age at transplantation was months (range, to months), median body weight was kg (range, to kg). / patients received a myeloablative conditioning regimen consisting of busulfan, fludarabine, cytarabine, cyclophosphamide and g-csf, / patients received another myeloablative conditioning regimen consisting of busulfan, fludarabine, cyclophosphamide and atg. prophylaxis for graft-versus-host disease (gvhd) was tacrolimus. results: engraftment occurred in / ( . %) patients. / ( . %) patients occurred graft failure, all of them received a myeloablative conditioning regimen without atg. median time to neutrophil and platelet engraftment were (rang, - ) and . (rang, - ) days. / ( . %) patients developed acute gvhd, with / ( . %) episodes of grade iii-iv agvhd. chronic gvhd occurred in / patients ( . %). at a median follow-up of months (rang, to months), the overall survival rate was . %, and event-free survival rate was . %. conclusions: unrelated ucbt should be considered as potential curative methods in children with cgd. cgd patients who used myeloablative conditioning regimen with atg shows better graft and survival. disclosure: nothing to declare background: invasive fungal infections (ifi) remain a major cause of treatment-related morbidity and mortality in aml patients. although not uncommon, the presentation of unusually severe clinical features might be indicative for an underlying immunodeficiency. caspase-associated recruitment domain (card ) is recognized to have a crucial role in effective antifungal response, leading to th and th differentiation and to the initiation of the inflammatory cytokine cascade. particularly interferon-gamma (ifng) increases macrophage activity. patients with homozygous card mutations are known to have a significantly increased susceptibility to life-threatening systemic candidiasis. however, some sequence variants may lead to increased ifi-susceptibility even in heterozygosity, e.g. under immunosuppression. ifn-γ has been described as an additive treatment option because of its immune stimulating effect on the leukocyte immune response in a situation of immunological "blindness". methods: here, we report the case of an -year old male with aml m with a severe systemic candida tropicalis infection, unresponsive to triple-antimycotic regimen, leading to multi-organ failure. he was discovered to bear a heterozygous card mutation, and ifn-γ immunotherapy leaded to complete response of all disseminated infections. results: the patient developed septic fever immediately after the first chemotherapy cycle. unexpectedly, candida tropicalis was confirmed in the blood culture within hours. liposomal amphotericin b (ambisome ® ) was started immediately, however candida rapidly disseminated to lungs, liver, spleen, kidneys and cns despite extended antimycotic therapy with caspofungin, voriconazole and fluconazole. the patient was splenectomized due to massive infiltration ( figure ). genetic testing for mycosis predisposition revealed a heterozygous mutation in the card gene, inherited from the father (c. a>t(p.glu val)). ifn-γ treatment was started ( μg subcutaneously, times per week), leading to an almost complete response of disseminated infections. due to the severe infection, chemotherapy had to be interrupted after one course. however, bone marrow remained in complete remission for almost one year. the patient experienced altogether two relapses requiring an unrelated allogeneic and a haploidentical hsct. under combined ifn-γ and ambisome/ fluconazole prophylaxis no further mycosis was observed despite extensive and prolonged immunosuppression. conclusions: ifi in aml patients are common, however an unusual presentation in presumably immune competent individuals should raise the suspicion for immunodeficiencies. in our case, an unexpected early candida-sepsis was completely unresponsive to an adequate multi-agent treatment. while ifn-γ is used in adults as an immune stimulatory cytokine, little data are available for children. to our knowledge, this is the first case of successful ifn-γ treatment of a pediatric aml patient with disseminated candida sepsis, bearing a card mutation. given the elevated mortality risk for ifi, and the apparently safe and well-tolerated application of ifn-γ, an adjuvant immunotherapy might be considered. further studies are needed to define the indication and duration of this kind of adjunctive immunotherapy. moreover, considering the wide heterogeneity of genetic mutations involved in ifi-susceptibility, genome-wide expression profiling might be useful for pediatric cancer patients, as the identification of specific immune pathways might help to identify individual ifisusceptibility in order to improve the outcome of those high-risk patients. [[p image] . background: chronic granulomatous disease (cgd) is curable by allogeneic hematopoetic stem cell transplant (hsct). recent reports of haploidentical donor hsct with with post transplant cyclophosphamide (ptcy) from family donors in pediatric primary immune deficiencies have shown encouraging results. however, it has not been reported in cgd. here we describe successful haploidentical hsct in a child with cgd with myeloablative conditioning and ptcy. a year-old, male child diagnosed with cgd showed oxidative activity . % by dihydrorhodamine (dhr) test. he had no matched related or unrelated donor available so underwent haploidentical hsct after taking informed consent of the parents in may . donor was his / hla matched healthy elder sister (oxidative activity % by dhr). he has had multiple admissions for recurrent pneumonia prior to hsct. the conditioning was with rituximab mg/m iv on day - , thiotepa mg/ kg/dose intravenous (iv) for day (day- ), busulfan . mg/kg/dose daily iv for days (day - to - ) and fludarabine mg/m /dose daily iv for days (day - to - ) and rabbit anti-thymoglubulin (thymoglobulin) . mg/ kg/dose daily for days (day- to - ). peripheral blood stem cells ( million/kg cd + cells) were harvested from his sister and transfused to the patient on day . graft vs. host disease (gvhd) prophylaxis was with ptcy mg/ kg on day+ & , intravenous cyclosporine from day- (targeting levels - ng/ml) and mmf from day+ . results: his neutrophils engrafted on day+ and platelets on day+ . chimerism on day+ , and months was fully donor. he developed no acute or chronic gvhd. at months his lymphocyte counts showed cd - /ul, cd - /ul, cd - /ul and cd / - /ul. his had no viral reactivation. he is disease free and gvhd free on day+ post hsct and is on tapering doses of cyclosporine. his dhr test showed oxidative activity of % on day + . background: primary immune deficiencies (pid) are a functional disorder of inheritance immune system that increase predisposition to infectious disease in number and severity. the incidence is : , birth live; its immunological dysregulation may increase the predisposition of autoimmune diseases and malignancy, the latter being more frequent ( - %). at present, the only curative treatment is hematopoietic stem cells transplant (hsct). methods: we describe all patients transplanted with primary immune deficiencies at instituto nacional de pediatria. the conditioning regimen depended on the type donor and pathology: myeloablative ( . %), reduced intensity ( . %) and non myeloablative ( . %) without modification statistically significantly in overall survival. results: a total of patients were included from to january/ . severe combined immunodeficiency (scid) is the pathology most frequently transplanted ( figure ) . seventy three percent have molecular diagnosis, and . % have cases of family pid. the most used sources were umbilical cord blood (ucb) with . % and peripheral blood ( . %), however the trend of the source of obtaining it has been modified a few years ago (figure ) . the median graft was days for ucb, days for bone marrow (bm) and days for peripheral blood (pb) (figure ) the main complications are infectious (bacterial . % and viral . %) and non-infectious as pre-graft syndrome ( . %). conclusions: the overall survival was . % survival according to pathology was: % chediak higashi syndrome, % scid, . % griselli syndrome, . % hyper igm syndrome, % was, % cgd, % hemophagocytic lymphohistiocytosis. disclosure: ramírez-uribe rosa maria nideshda, salazar-rosales haydeé, olaya-vargas alberto, lópez-hernández gerardo, del campo-martínez maria de los Àngeles we wish to confirm that there are no known conflicts of interest associated with this abstact, the only financial support was provided by mexican associations that helping children wiht cancer in a few patients. long-term outcome following hematopoietic stem cell transplantation of wiskott-aldrich syndrome in a single institute mamoru honda , , yukayo terashita , minako sugiyama , yuko cho , akihiro iguchi background: wiskott-aldrich syndrome (was) is an xlinked disorder of hematopoietic cells, characterized by thrombocytopenia with small platelets, eczema, and immunodeficiency. hematopoietic stem cell transplantation (hsct) is the only curative treatment, and it is recommended to be performed as soon as was is diagnosed. myeloablative conditioning before hsct is recommended because there is a high risk of development of autoimmune disease in patients with mixed chimera after hsct. however, there are few reports about late complications such as pubertal development and eruption of teeth in patients with was receiving hsct. thus, we evaluated late complications in patients with was receiving hsct at hokkaido university hospital. methods: we reviewed medical records of male patients with was who received hsct between and . results: mean age at hsct was . (range, . - . ) years, and median follow-up time after hsct was . (range . - . ) years. conditioning regimen in all patients comprised busulfan at mg/kg for days and cyclophosphamide at mg/kg for days or mg/kg for days. additionally, anti-thymocyte globulin at . mg/kg/day for - days was administered in patients. engraftment, normal platelet count, and complete chimera were confirmed in all patients. no patients showed complications such as severe chronic graft-versus-host disease, autoimmune disease, short stature (≤ - . sd) and second malignancy. however, high ige level was observed in patients. pubertal development has been confirmed in patients. lack of complete eruption of permanent teeth has been observed in patients who received hsct at age of < years. conclusions: although this was a small-cohort study in a single institute, complete chimera has been achieved in all patients who received hsct with busulfan-based myeloablative conditioning. however, late complications such as male infertility and incomplete eruption of permanent teeth remain major problems. disclosure: nothing to declare methods: we performed unrelated umbilical cord blood transplantation (ucbt) in consecutive children with lad-i. median age of children was months (range, to months), and median body weight was kg (range, to . kg). all patients received myeloablative conditioning regimen consisting of busulfan, fludarabine and cytarabine. prophylaxis for graft-versus-host disease (gvhd) was tacrolimus. all patients received a ≤ hla alleles-mismatched cord unit, was hla fully matched, were / matched, was / matched. median nucleated cells of the cord blood were . x /kg (range, . to . x /kg), and median cd + cells were . x / kg (range, . to . x /kg). results: all patients engrafted, median time of neutrophil engraftment was days (range, to d), and median time of platelet engraftment was days (range, to d). median follow-up time was months (range, to months), all patients were alive with continuous completely donor engraftment, and achieved complete clinical remissions. / patients developed grade ii/iii acute graft-versus-host disease (gvhd), and / patients developed chronic gvhd with skin. conclusions: it is the first successful unrelated ubct for lad-i children in china. our data shows ucbt provided excellent outcome for patients with lad-i. disclosure: nothing to declare p excellent outcome using 'nktm' enriched hematopoietic stem cell transplants for patients with inborn errors of immunity results: majority of patients in the cohorts had significant infective co-morbidities at the time of hsct with patients in the later cohort entering hsct earlier. patients in the later cohort were sicker at hsct. final engraftment occurred in all except patient who received a hla mis-matched cord blood hsct. graft failures occurred in patients ( in earlier and in later cohort); of these patients received unmanipulated hscts from hla mis-matched unrelated donors ( cb, bm). second hsct were with same donors in and different donors in patients. no grade ii to iv acute gvhd or extensive gvhd occurred. one patient (cbt) died of infections/ non-engraftment. all patients in the later cohort compared to of patients in earlier cohort are alive, engrafted and cured. performance status were % in all alive patients. of the patients in the later cohort, ( hla mis-matched related and hla matched related donors) received 'nktm' enriched hsct. in the hla mis-matched 'nktm' enriched hscts, patients received high cd +, cd +cd ro+ and nk cell doses, with median of . (range, . - . ), . (range, . - . ) and . (range, - ) x /kg, respectively. no invasive infections occurred in these patients and immune reconstitution in t, b, nk compartments were complete at year after hsct with cd > by months and tcrαb > by year after hsct. background: viral infections contribute to significant morbidity and mortality after allogeneic hematopoietic cell transplantation (allo-hsct), increasing both the human and the financial cost. antiviral agents are often ineffective or/ and associated with toxicity. methods: in view of t-cell anti-viral immunotherapy in greece, we evaluated the actual cost of conventional pharmacotherapy for cmv, ebv and bkv reactivations after allo-hsct, by calculating the costs of (i) the antiviral agents, (ii) the treatment (excluding transfusions) of antiviral drug primary toxicity (e.g. graft failure, cytopenias, renal or hepatic dysfunction) and secondary toxicity (e.g. leukopenia-associated bacterial infections), iii) the treatment (excluding transfusions, ie for bk cystitis) of infectionrelated complications, iv) the transfusions due to treatmentrelated toxicity (ie cytopenias) or infection-related complications (ie, bk cystitis), v) the inpatient or outpatient daily care. notwithstanding that blood and its products, as a common natural good, are provided free in our country, the costs related to blood and platelet collection, processing, storage, laboratory testing and infusions were included in our model. results: the treatment cost of cmv, ebv and bkv reactivations/infections for the first six months post allo-hsct was evaluated in / patients who reactivated viruses and were transplanted between / - / from matched related ( / ), matched unrelated ( / ), mismatched unrelated ( / ), haploidentical ( / ) and mismatched related donors ( / ). we detected cmv, ebv and bkv infections/reactivations in , and patients respectively, with a mean of ± . infection per patient from all three viruses ( - /patient). of note, / patients experienced reactivations from more than one virus, requiring repeated treatments with antiviral agents and/or rituximab. the cost of antiviral agents for all cmv, ebv and bkv reactivations/infections was , €, , € and , € respectively ( , €, , € and , €/patient, respectively). the treatment cost of toxicity related to antiviral drugs and infection-related complications was , € ( , €/patient) excluding transfusions and , € ( , €/patient) including transfusions. in particular, the cost of transfusions for bkv hemorrhagic cystitis reached , €/patient. repeated ( - ) and/or prolonged (Δm d, range - d) hospitalizations were needed, up to a total of and days of inpatient hospitalization and short-term outpatient treatment, respectively. hospitalizations further increased the cost of inpatient and outpatient post-transplant care by , € and , € respectively ( , € and €/patient, respectively), onthe basis of a, rather underestimating the true cost, fixed, unified hospitalization fee ( €/day and €/day). conclusions: overall, in a six-month study period, the treatment of cmv, ebv and bkv infections substantially increased the cost of post-transplant care by , € ( , €/patient). the actual cost is undoubtedly higher as the hospitalization fee for transplant recipients is largely underestimated in greece. considering not only the hematopoietic but also the solid organ transplant recipients, the financial burden of antiviral treatment for national economies is enormous. given that antiviral pharmacotherapy is often associated with suboptimal efficacy, toxicity, development of drug resistance, reactivation recurrences and repeated hospitalizations, it is expected that a one-time treatment with multi-virus-specific t cells, able to expand in vivo and provide a long-lasting protection without significant toxicity, will serve as a powerful and costeffective treatment over conventional pharmacotherapy. disclosure: nothing to declare methods: this is a single-centre retrospective analysis of consecutive patients who underwent tcd allo-hscts for myeloid malignancies between january -june . ebv-dna was monitored frequently on whole blood samples with standardised quantitative real-time pcr. serum protein electrophoresis was routinely tested with immunoglobulin subclasses identified by immunofixation electrophoresis. histological confirmation of ptld was based on standard who diagnostic criteria ('proven'), while those without biopsy were classed as 'probable' based on clinical & radiological criteria as defined by ecil- guidelines. results: majority of patients had aml(n- / ) and mds(n- / ) with a median age of years(range . median follow up of survivors was months(range - ). majority of patients(n- / ; %) developed ebv-r with a median time of days[inter quartile range(iqr) - days] &higher cumulative incidence with atg(n- ) versus alemtuzumab(n- )(p< . ). figure- a shows schematic representation of ebv and ptld events (cumulative incidence of . %( %ci- . %- . %) at months). significantly higher peak ebv dna viral load(evl) were noted in patients with ptld(p-< . ). development of post-hsct mg was observed in %(n- / ). roc curve identified peak blood evl> , copies/ml significantly correlated with risk of developing ptld (sensitivity- . %,specificity- . %;auc- . ,p< . ). based on these estimates, subgroup of patients with no ebv-r(n- / ), peak evl < , (< k)copies/ ml(n- / ) & > , (> k)copies/ml(n- / ) were categorised in groups along patients with/without mg accordingly (groups - ;figure- b). patients with ebv-r had significantly better os [ -year os of % vs %(no ebv-r);log-rank p< . ],with this survival benefit mainly driven by subgroup of patients with lower evl(< k)(p< . ). ptld patients had trend towards inferior -year os( % vs %;p- . ). patients with mg had a significantly better os irrespective of degree of evl (group - ,p< . ).we report a 'sweet spot' of low evl & presence of mg in these patients, with a clear survival advantage compared to those with no ebv-r and/or no mprotein (group- -year os % vs % in group- ; hr- . ; %ci: . - . ;p< . ;figure b). overall cumulative incidence of relapse (cir) was %( %ci: - ) and non-relapse-mortality(nrm) of %( %ci: . - ) at years. multivariate analysis(mva) revealed absence of m-protein,high evl (> k copies/ml) or no ebv-r and absence of any gvhd as significant factors for high cir. similarly, high evl or no ebv-r, absence of m-protein and itu admission were significant predictors of high nrm. conclusions: this study adds to our understanding of role of ebv viraemia & associated mg in tcd-hscts while highlighting its significant impact on risk of ptld, os, nrm & cir. low ebv burden and development of mg is protective with significantly better survival outcomes and we recommend pre-emptive approach of using rituximab for ebv-r /ptld is best employed at higher ebv burden (e.g. > k copies/ml dna) in high risk patients and be prospectively evaluated in future studies. clinical trial registry: n/a disclosure: nothing to declare p impact of early candidemia on the long-term outcome of allogeneic hematopoietic stem cell transplant in non leukemic patients: an outcome analysis on behalf of idwp background: to assess the incidence of, and risk factors for, candida infection in the first days post-allogeneic hematopoietic stem cell transplantation (hsct) and the impact on long-term survival. methods: outcome analysis of , patients, % male, median age years (range - ), with diagnosis of hemoglobinopathies in ( . %), bone marrow failure in ( . %), lymphoma in ( . %) and myelodysplastic/myeloproliferative disesases in ( . %) patients who underwent hsct from to : with candidemia by day + , and , without candidemia. the incidence of -day candidemia was estimated by using the cumulative incidence method. the univariate and multivariate risk factor analysis for -day candidemia was performed with the cause-specific cox regression model. the occurrence of candidemia was analyzed as a timedependent covariate. the overall survival and non-relapse mortality after day + were assessed in a land-mark setting, this analysis was restricted to patients surviving to day + post transplant. [[p image] . figure a - b] results: the incidence of candidemia by day + was . % ( % c.i. . - . ) ( / , ) and occurred at a median of days post-hsct (range - - ). considering the candidemia within -day from hsct as a time dependent covariate, a higher -day non-relapsemortality (nrm) (hr . ( . - . ), p < . ), and a lower -day overall-survival (os) (hr . , % ci . - . ), p< . ) were obtained from the cox model for patients with candidemia. factors significantly associated with candidemia occurrence in the multivariate analysis were: gender female, increased age at hsct, bone marrow failure, lymphoma or myelodysplastic/myeloproliferative diagnosis, bone marrow or cord blood stem cell source, t-cell depletion, less recent year of hsct. among patients alive at day + , the -year nrm and os with and without candidemia were . % vs. . %, p < . , and . % vs. . %, p< . , respectively, after a median follow-up of . years ( % ci . - . ) (figure ). in multivariate analysis, the occurrence of a candidemia episode within day + was an independent risk factor for higher nrm, hr . ( . - . ), p= . , and lower os, hr . ( . - . ), p= . . conclusions: despite the general improvements in prophylaxis and treatment, the occurrence of early post-hsct candidemia had a negative impact on transplant outcome as showed previously in leukemic patients. abstract already published. carbapenem-resistant enterobacteriaceae colonizationimportance in the risk of cre bacteremia and mortality in stem cell transplant (hsct) and acute leukemia patients marcia garnica , , marco a f bellizze , priscila g a de jesus , rafaela r c gomes , filipe m akamine , alan j marçal , luzinete co rangel , andreia assis , marcia rejane valentim , angelo maiolino background: spread of infections due to carbapenemresistant enterobacteriaceae (cre) is a worldwide phenomenon and has been associated with high mortality and clinical complications. gut translocation is the most important portal of entry of bacteria during neutropenia, and cre gut colonization is a possible risk factor for bacteremia during neutropenia. goals: in the present study, we describe the frequencies of cre colonization and analyzed its relationship with development of cre bacteremia and mortality in two different scenarios: stem cell transplant patients (hsct) and leukemia patients. methods: prospective cohorts of hsct (from to ) and leukemia patients (from to ). hsct patients were analyzed from conditioning until discharge (pre-engraphment phase) and leukemia patients from first induction chemotherapy until last intensification. if an hsct was performed in leukemia patient the patient was censored in leukemia cohort and included in hsct cohort. all patients had rectal swabs performed weekly during hospitalization for the identification of cre colonization. patients with at least one positive swab (cre colonization group) were compared to patients with no documentation of colonization (controls). the outcomes analyzed were bacteremia due to cre, and overall mortality. results: there were hsct performed during the study ( [ %] autologous and [ %] allogeneic). multiple myeloma and non-hodgkin lymphoma were the most frequent baseline diseases (n= ; %, and n = ; %), respectively. cre colonization was documented in % (n= ), and it was more frequent among allogeneic hsct and leukemia patients (p< . for both). cre colonized patients had longer hospitalization ( vs. days, p< , ), higher frequency of cre bacteremia ( % vs. . %; p= . ), and mortality ( % vs. . %, p< , ) compared to non-colonized hsct. negative and positive predicted values for cre bacteremia were % and %, respectively. thirty-one patients were analyzed in leukemia cohort, accounting to hospitalizations (median hospitalizations per patient, ranging from to ). the median age was years, and % aml vs. % all. cre colonization was documented in eight ( %), with a median time from leukemia diagnosis and colonization of days ( - days). cre bacteremia was documented only in colonized patients ( % vs. zero; p= , ). all eight colonized patients were submitted to other cycles of chemotherapy after colonization, in one of them cre bacteremia relapsed. conclusions: a routine surveillance of cre colonization showed colonization frequencies from % to % in hsct and leukemia patients respectively and was effective to stratify cre bacteremia risk as the predictive negative value was over %. colonization had association with cre bacteremia and overall mortality. efforts to minimize risks for colonization and mortality are necessary. the information of surveillance can be a tool to improve adequacy in empirical febrile neutropenia therapy in hsct and leukemia patients. disclosure: nothing to declare background: the incidence of hepatitis b virus infection is high to . % in asian population, so there is more and more attention to the risk of hepatitis b virus(hbv) reactivation in the hepatitis b core antibody positive patients during chemotherapy, anti-cd monoclonal antibody, hsct, or other intense immunosuppressive drug therapy (isdt). hepatitis b core antibody is associated with a significant risk of hbv reactivation in patients undergoing hsct. however, there are remain uncertain that the effect of anti-hbsag antibodies in hepatitis b virus reactivation among the hepatitis b core antibody positive patients undergo hsct. we aim to investigate the role of the anti-hbs and the necessity of anti virus in hepatitis b surface antigen(hbsag) negative, hepatitis b core antibody positive patients during hsct. methods: we enrolled hematological malignant patients received hsct in our center from to . we classified hbsag negative and undetectable hbv dna patients into groups as anti-hbc(-)anti-hbs(-) (n= ), anti-hbc(-)anti-hbs(+) (n= ), anti-hbc(+) anti-hbs(-) (n= ), and anti-hbc(+)anti-hbs(+) (n= ). results: hbv reactivation was identified in patients ( . %) after hsct. there was a significant difference in hbv reactivation rate in anti-hbc(+)anti-hbs(-) ( . %) vs anti-hbc(+)anti-hbs(+) ( . %) (p= . ) and anti-hbc (+)anti-hbs(-) ( . %) vs anti-hbc(-)anti-hbs(-) ( . %) (p= . ), anti-hbc(+)anti-hbs(-) ( . %) vs anti-hbc(-) anti-hbs(+) ( . %) (p= . ), but not among anti-hbc(+) anti-hbs(+) ( . %) and anti-hbc(-)anti-hbs(-) ( %) and anti-hbc(-)anti-hbs(+) ( . %). whereas there were no difference according to the donor viral profile(p= . ). the median time of hbv reactivation in hbsag negative patients accepted hsct was ( - ) days after hsct. all of the patients with hbv reactivation have been controlled with nucleos(t)ide analogues drugs, and of them achieved reverse seroconversion which detect persistent anti-hbsag antibodies in their bodies. conclusions: the anti-hbsag antibodies negative and anti-hbc positive patients have the highest risk of hbv reactivation after hsct in resolved hbv patients. the anti-hbsag antibodies play a protective role in resolved hbv patients receiving hsct. we recommend not prophylactic anti hepatitis b virus in hbsag negativity and anti-hbsag antibodies positive patients following hematopoietic stem cell transplantation. disclosure: nothing to declare methods: allo-hscts performed between and for acquired bone marrow failure ( . %) or hemoglobinopathies ( . %), with bm±cb ( . %) or pb±cb+bm ( . %) as a stem cell source were included in this retrospective registry megafile idwp ebmt study. results: demographics: the median age of recipient was . years (range: . - ), and . % were children. . % recipients and . % donors were ebv-seropositive. . % had hsct from a matched family donor, . % from a mismatched family donor, and . % from an unrelated donor. t-cell depletion was performed in vivo in . %, and ex vivo in . % patients. conditioning regimen was myeloablative in . %, ric in . %. median follow-up was . years ( % c.i. . - . ). transplant out-comes: ebv-seropositive recipients in comparison to ebv-seronegative recipients had lower os ( . % vs . %, p= . ), and higher nrm ( . % vs . %, p= . ). no other significant differences were found for: ri, rfs, and acute or chronic gvhd with respect to ebv pretransplant serostatus donor and/or recipient. multi-variate analysis: ebv serostatus as a risk factor did not reach significance, while a trend towards higher risk of development of cgvhd (hr= . ; %ci . - . ; p= . ) and better survival (hr= . ; %ci . - . ; p= . ) in allo-hsct from ebv-seropositive donors. allo-hsct in ebv-seropositive recipients had a trend towards lower risk of development of cgvhd (hr= . ; %ci . - . ; p= . ). when subgroups (r-/d-, r-/ d+, r+/d-, r+/d+ ebv serology) were analyzed, the ebv serostatus had no significant impact on os, rfs, ri, trm and development of acute or chronic gvhd. conclusions: allo-hsct from ebv-seropositive vs ebv-seronegative donors are at % higher risk of chronic gvhd in patients with non-malignant hematological disorders undergoing allo-hsct, however this difference is non-significant in multivariate analysis. disclosure: nothing to declare. results: twenty-eight ( %) pts ( male, female) were tested positive (group ) for subtype a (n= , %), b (n= , %) or a (h n ) (n= , %) while ( %) pts ( male, female) were negative (group ). vaccination rate in group ( %) was significantly lower compared to group ( %, p= . ). the median time after transplantation ( vs days), t-cell counts ( vs / μl), bcell counts ( vs / μl), igg-level ( , vs , g/ l), proportion of immunosuppressed pts ( % vs %), male/ female ratio was not significantly different between groups and . within group influenza subtypes were similarly distributed in vaccinated and not vaccinated pts (a % vs %, b % vs %, a (h n ) % vs %). pts. with subtype b infection had higher levels of t-( vs / μl) and b-cells ( vs / μl) and a longer follow up from sct ( vs days) compared to subtype a / a (h n ) infection but differences were not significant. conclusions: influenza could be proven in one third of all tested pts. dominance of b and a (h n ) pdm e subtype occurrence corresponded to the flu epidemic dissemination in the german population. the most important protective factor for outpatient sct recipients was influenza vaccination. disclosure: nothing to declare background: cmv infection is one of the most frequent complications after haplo. some risk factors are well known but the best strategy (prophylactic or preemptive treatment) to mitigate this complication is not still well defined. the primary endpoint in our study is to describe incidence and risk factors to develop cmv infection or disease in haplo. as secondary objective we analyzed efficacytoxicity of treatment and cmv related mortality. methods: we analyzed patients who underwent haplo in our center between may and may . all of them received ptcy (d+ and d+ ), tacrolimus and mycophenolate as graft versus host disease prophylaxis. a preemptive therapy based on viral load was applied. treatment was started when > ui/ml of cmv were detected in one determination or > ui/ml in two consecutive determinations. cmv analyses were made in plasma using cobas pcr technique® and positive viral load cut-off point was ui/ml. the cmv viremia was determined weekly until d+ and then every two weeks until immune reconstitution. results: the cmv infection and disease incidence at d + was . % ( episodes) and . % ( episodes), respectively. cmv disease was digestive (n= ), pulmonar (n= ), neurologic (n= ) and disseminated (n= ). the median time to first cmv infection was . days ( - ). thirty-six patients had at least one episode of cmv infection: of them ( . %) had one episode, ( . %) had two episodes and ( . %) had or more episodes, respectively. only pre-transplantation cmv status was significantly associated with cmv infection (p< . ). risk factors are shown in image . the median viral load in first cmv infection and disease was ui/ml ( - ) and ui/ml ( - ), respectively (p= . ).the median counts of cd lymphocytes at d+ in cmv infection and disease were /mm and /mm , respectively (p= . ). preemptive therapy for the first episodes of cmv infections (n= ) was valganciclovir ( , %), ganciclovir ( . %) or foscarnet ( . %), reaching a complete viral load clearance in %, with a median time to response of . days ( - ) and a median treatment duration of days ( - ). grade iii-iv toxicity (mainly hematologic) was observed in . % (n= ), % (n= ) and . % (n= ), respectively. three patients had an ul mutation, one of them with clinical and microbiological resistance to the mentioned drugs. three patients ( %) had a graft failure secondary to cmv infections. five patients ( . %) died as consequence of cmv infection: before d+ secondary to cmv disease ( pulmonar, disseminated) and after d+ due to graft failure and infectious complications. with a median follow up of . months, overall survival at months for patients who had cmv infection was . % compared to . % for those who had no infection (p= . ). conclusions: a high incidence of cmv infection in haplo with ptcy was shown in our series and it contributed to mortality in . % of patients. only cmv status (d-/r+ and d+/r+) was significantly associated with higher risk of infection. identification of high risk patients and new prophylactic and treatment strategies may improve these results. disclosure: nothing to declare. methods: consecutive patients admitted at the sct unit between january- to november- were reviewed. only first admission was analysed. screening consisted of rectal and perineal swap on admission and weekly until discharge. in case of detection of mdro, patients were isolated and infection control strategies were applied. results: patients were analysed, median age years ( - ). % were male (n= ). median duration of hospitalization was days( - ). swabs were performed, with a median of swaps/patient ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) . patient characteristics are shown in table . patients ( %) spiked fever in a median of days after admission . % (n= ) had previous documented mdro colonization. median neutrophil engraftment was days ( %ci - ), in % patients (n= ) of patients had a positive screen: in ( %) patients at baseline and in ( %) patients were detected for the first time beyond baseline screen. cumulative incidence of colonization at days was . % ( %ci . - . ), at days % ( %ci . - . ), and at days . % ( . - . %) (figure ). mdro identified were: with extended-spectrum beta-lactamases producing e. coli (esbl-ec), multidrug-resistant pseudomonas aeruginosa (mr-ps), vancomycin resistant enterococci (vre) and patient with carbapenemaseproducing (cp) citrobacter freundii. / colonized patients developed mdro infection ( %): patients mr-ps, site of infection was urinary tract infection (uti), urethritis, genital ulcer. two patients were treated with ceftolozane/ tazobactam, with meropenem+amikacin; patients esbl-ec both uti treated with meropenem; patient cpcitrobacter freundii uti treated with ceftazidime/avibactam. in % patients ( / ) antibiotic treatment at febrile episode was guided by positive screening. no mdro related icu admission or mortality was observed. in % patients (n= ) background: hepatitis e virus (hev) can cause chronic infection and liver cirrhosis in immunocompromised individuals. there is limited data on hev infections in patients undergoing allogeneic hematopoietic stem cell transplantation (hsct). the aim of this study was to investigate the frequency and clinical importance of hev in a swedish cohort of hsct recipients. methods: we analyzed serum samples from hsct patients ( adults and children), collected months after hsct. hev igg and igm were detected by elisa (dia.pro®), hev rna by reverse transcriptase pcr, and quantification of hev rna was performed by digital pcr. in all patients, who were positive for hev-rna and/or serology at months, also samples collected at the time of hsct from both the patients and their donors were analyzed. in the hev rna positive patients, additional samples were analyzed to determine the duration of viremia. three hev rna negative controls were selected for each case of hev infection, matched for age, diagnosis, conditioning regimen and donor type. results: hev rna was detected in / ( . %) patients. in three of the patients hev rna was positive during a period of - months, and two of these patients were infected already at the time of hsct. in five patients hev-rna was positive, at a low level, only at months. / ( . %) patients had detectable hev igg and/or igm, whereof eight patients were hev rna negative. in / ( %) patients with hev infection (hev rna positive) alanine aminotransferase (alt) was > upper limit of normal (uln), in / ( . %) patients > . uln, and in / ( . %) patients alt was normal, at months after hsct. bilirubin was elevated > . uln in / ( . %) patients, and > uln in no patient at months after hsct. two patients died with ongoing signs of hepatitis and hev rna detected in blood. one of them developed acute liver failure, at the time interpreted as drug toxicity, and died of multi-organ failure. the other patient died of unrelated causes. the remaining six patients had cleared the infection at - (median . ) months after hsct. active gvhd was present at months after hsct in / ( . %) patients with hev infection, involving the liver in of these patients. corticosteroid treatment was ongoing in / ( %) patients; the mean dose during the preceding days was > . mg/kg in / ( . %) patient, . - . mg/ kg in / ( . %) patients, and < . mg/kg in / ( %) patients. hev infection correlated to elevated alt > . uln, or . p= . ) and > uln, p= . ) at months, but not at months, after hsct, compared to hev rna negative controls. conclusions: hev infection was detected in . % of patients tested at months after hsct and was correlated to abnormal alt. spontaneous clearance was common but one patient died in acute liver failure, where hev may have contributed. hev infection is a differential diagnosis in patients with elevated alt months after hsct. disclosure: nothing to declare monitoring of t-cell responses to viral-coded antigens in pediatric patients receiving tcrαβ-depleted haplo-hsct followed by bpx- cell administration background: αβ t-cell-depleted haplo-hsct is an effective option for children with hematological disorders in need of an allograft. however, recovery of adaptive immunity is impaired in these patients. thus, in order to accelerate immune reconstitution, we developed a novel approach based on post-transplant infusion of a titrated number of donor t cells, transduced with the suicide gene inducible-caspase- , ic (bpx- cells, sponsor bellicum pharmaceuticals®; nct ). we previously reported on immune recovery of children transplanted at our institution, showing that bpx- cells infused after αβ t-cell-depleted haplo-hsct expand in-vivo and persist over time, contributing to fasten adaptive immunity recovery (merli, ash ). here, we report the results of lymphoproliferation assay to viral-encoded antigens to assess tcell function in patients transplanted with this approach. methods: we evaluated children, male and female. median age at transplant was . years (range . [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] . patients had either malignant ( children) and nonmalignant ( ) disorders. no patient was given any posttransplant graft-versus-host disease prophylaxis. nine children were enrolled in the phase i portion of the trial consisting of cohorts receiving escalating doses of bpx- cells. the remaining patients (phase ii portion) received the recommended dose of x bpx- cells/kg identified in phase i. bpx- cells were infused at a median of days post-hsct (range - ). antigendriven activation of peripheral mononuclear cells was evaluated by lymphoproliferation assay with h-thymidine pulsing at d+ and harvesting at d+ . stimuli included pha or cmv, ebv and adv whole viral lysate. results are given as stimulation indexes (si, cpm stimulated sample/cpm unstimulated control). thresholds for positive response were arbitrarily set at si> for viral-encoded antigens and at si> for mitogenic stimulation with pha. fractions of responders are indicated in the figure. results: patients were analyzed from d+ to d+ post-hsct. pha responders (a) increased to %, while cmv (b), ebv (c) and adv (d) responders were %, % and % at years after haplo-hsct. responses to ebv and adv antigens were slightly delayed but improved over time. responses to pha and to cmv (e,f) were analyzed in the cmv-reactivating and cmv-non reactivating groups (cmv-yes/cmv-no). significant differences in pha response were observed at d+ and d+ . moreover, increased cmv responses were observed in cmvreactivators at d+ , d+ and d+ , with approximately % of responders at d+ , as opposed to cmvnon reactivators which comprised % responders. neither primary disease, age nor tbi during the conditioning regimen influenced proliferative capacity of the two subgroups (not shown). conclusions: we showed a rapid recovery over time of t-cell function after αβ t-cell-depleted haplo-hsct followed by bpx- cells administration. when patients were grouped according to cmv reactivation (previously demonstrated as a strong driver of immune reconstitution), a significant difference in the number of responders among the patients experiencing viral reactivation was observed using the cmv lysate only but not the immunodominant pp protein (not shown), suggesting that other viral antigens account for increased t-cell responses. results of t-cell function after bpx administration complements the phenotypic data we already reported. clinical trial registry: nct disclosure: nothing to declare. background: cytomegalovirus (cmv) is associated with significant morbidity and mortality in allogeneic hematopoietic cell transplantation (allo-hct) patients (pts). cumulative incidence of cmv infection in high-risk patients such as cd -selected or haploidentical hct have been reported as high as . - . % and - %, respectively. letermovir (ltv) was approved in / for prophylaxis (ppx) in cmv-seropositive recipients (r+) of allo-hct. since / , ltv ppx was implemented at our center for both primary and secondary ppx. we report our real-world experience. methods: adult cmv r+ allo-hct pts who initiated ltv as primary and/or secondary prophylaxis were identified between / / and / / . cord blood transplants were excluded. the primary outcome was the incidence of clinically significant cmv infection (cmv viremia requiring preemptive treatment or cmv disease). pts were followed through / . results: pts initiated ltv. . % pts were at high risk for cmv reactivation and disease (primarily ex vivo t-cell depleted hct [n = ; %] or haploidentical t-replete hct [n = ; . %]). the most common indication for hct was acute myeloid leukemia (n = ; . %) and the majority of patients received myeloablative conditioning (n = ; . %). pts ( . %) received ltv as primary ppx after hct, with a median day of ltv initiation of d+ (range d+ ─d+ ). at ltv initiation, pts had an undetectable cmv dna, and had cmv < iu/ml. clinically significant cmv infection requiring preemptive treatment occurred in of pts ( . %). one patient was treated with valganciclovir (vgv) for persistent cmv < iu/ml and received ltv as secondary ppx. a nd patient developed persistently detectable cmv (< iu/ ml) and breakthrough cmv viremia with a mutation in ul at site c yltv successfully treated with vgv. the median duration of primary ltv ppx was days ( - ), with primary ppx continuing beyond weeks post hct in pts. the median additional follow-up in patients who discontinued ltv was days ( - ), without clinically significant cmv infection to date. an additional pts ( pts overall; . %) received ltv as secondary ppx after cmv pre-emptive therapy. the median duration of secondary ltv ppx was days ( - ), with no reactivation. ltv was not discontinued due to toxicity or intolerance in any patient. cmv outcomes are summarized in figure . all-cause mortality for the pts over the observational period was . %. conclusions: primary ltv ppx significantly reduced cmv reactivation, and high-risk patients may benefit from extended prophylaxis. in patients who received preemptive therapy for cmv, use of secondary ppx showed no recurrent cmv reactivation. ltv is well tolerated. additional studies are needed to determine optimal ppx duration and to clarify role of secondary cmv ppx in high-risk allo-hct. the future standard of care will likely include extended primary ppx and secondary ppx and result in decreased morbidity and mortality associated with cmv. disclosure: andrew lin -nothing to declare, molly a. maloy -nothing to declare, valkal bhatt -nothing to declare, lauren derespiris -nothing to declare, meagan griffin -nothing to declare carmen lau -nothing to declare, anthony j. proli -nothing to declare, juliet barker -angiocrine bioscience , letermovir primary prophylaxis (pp) has been shown to reduce clinically significant cmv infection with a favorable safety profile. letermovir pp will improve the outcome of seropositive patients. however, patients who did not benefit from pp and experienced > cmv episode (infection or disease) after hct may be candidate to secondary prophylaxis (sp). indeed half of them will have > recurrent episode after pre-emptive treatment (pet). letermovir is available since november as part of the french early access program for pp and sp. we report the outcome of patients who benefited from letermovir sp in the context of this program. methods: letermovir is granted, in a restrictive manner, by the french drug agency (ansm) on a case-by-case basis for prophylaxis of cmv episode, in cmv-seropositive adult allogeneic hct recipients. sp patients should have a negative baseline cmv pcr, have already experienced > cmv episode, in the context of a potentially harmful pet according to physicians. planned letermovir daily dose was mg in case of concomitant cyclosporine and mg otherwise. all patients were routinely screened by blood or plasma cmv pcr. results: between november -july , patients received letermovir in the early access program, for pp, and for sp. among the sp patients, had previous cmv disease (gut: ; cns: ). mean age was ± years, m/f ratio was / . the sp cohort included one cord blood and haplo-identical hct. main diagnoses were acute leukemia ( %) and myelodysplastic syndrome ( %). the conditioning regimen was myeloablative in % and included atg in %. based on available data ( missing data, md), previous gvhd was present in ( %) patients, and active at letermovir initiation in ( %). thirty two ( %) patients were planned to receive immunosuppressants. donor's cmv serology was negative in / ( %) ( md). at baseline, cmv pcr was detectable in / patients. letermovir was initiated a median of days (iqr: - ) after transplant for a mean duration of ± days. only one ( %) patient developed cmv breakthrough. the median follow-up from letermovir initiation was days. among the patients exposed to letermovir prophylaxis, two patients permanently discontinued because of letermovir-related adverse events (acute gvhd and nephropathy for one, loss of appetite, pruritus, diarrhoea and weight loss for the other); two deaths occurred with no causal relationship to letermovir. data were consistent with the known safety profile of letermovir. conclusions: letermovir is or will be soonly available in most european countries for cmv prophylaxis in hct recipients. pending its routine use, letermovir used as sp was well tolerated and effective, with only / patients developing a breakthrough infection. in this high-risk population for cmv recurrence, letermovir may provide a safe bridge between pet and specific immune reconstitution, pending tapering or discontinuation of immunosuppressants. whether sp may improve survival deserves further studies. disclosure: thierry allavoine is a former employee of msd france, nathalie benard and amir guidoum are employees of msd france, marion masure is an employee of icta pm, sophie alain and catherine cordonnier have participated in advisory boards and have been members of the speaker bureau of msd. ibrahim yacoub-agha has received honoraria from msd, other authors: nothing to declare p real-world data on letermovir prophylaxis for cytomegalovirus reactivation after allogeneic hematopoietic cell transplantation: a single center experience patrick derigs , maria-luisa schubert , paul schnitzler , carsten müller-tidow , peter dreger , michael schmitt heidelberg university hospital, heidelberg, germany, background: reactivation of cytomegalovirus (cmv) still contributes substantially to morbidity and mortality after allogeneic hematopoietic cell transplantation (allohct). recently, letermovir became available as the first drug approved in europe for prophylaxis of cmv reactivation in seropositive patients who have undergone allohct. letermovir is neither myelo-nor nephrotoxic, and significantly reduced the incidence of cmv reactivation in a pivotal phase iii trial (nejm ; : ) . therefore we adopted letermovir prophylaxis according to the label as standard policy in our institution: in seropositive recipients letermovir was initiated after engraftment and continued until day + or cmv reactivation. the aim of the present study was to investigate if the favorable trial results could be reproduced under real-world conditions. methods: the study cohort consisted of the first seropositive patients who received letermovir prophylaxis at our institution (between march and august ). these were compared with a control cohort transplanted between august and march before the advent of letermovir. study and control cohorts were matched for cmv donor/recipient sero-status, underlying disease and donor type source of stem cells and application of atg. cmv viremia was monitored by a quantitative pcr twice a week during the inpatient period and weekly thereafter. patients reactivating cmv prior to engraftment were not considered as event in both groups. results: no major side effects of letermovir intake were observed. with altogether reactivation events, the cumulative rate of cmv reactivation on day + was % ( %ci - %) in the letermovir cohort and thus significantly lower than in the control group ( events, % ( %ci - %); hr . ( . - . ); p= . ). the median time to reactivation was days for the control group and not reached for the letermovir group. the cumulative number of days on valganciclovir before d + was d for the letermovir patients vs d for the control patients. there were no hospitalizations for foscavir administration in the letermovir group compared to hospitalizations in the control group. there were deaths before d + in the letermovir group (one pd, one nrm) and deaths in the control group (all pd). conclusions: this observational study confirms the safety and efficacy of letermovir for the prophylaxis of cmv reactivation in seropositive patients after allohct in a real-world setting. our results are in good concordance with the phase iii trial. although letermovir appeared to reduce the need for therapeutic valganciclovir and foscavir tremendously, larger samples with longer follow-up are needed to assess the impact of letermovir prophylaxis on non-relapse and overall mortality as well as on resource consumption. background: cmv viremia occurs in %- % of cmv r + hct recipients. pet use has reduced the risk of cmv end-organ disease (eod) and associated mortality; however, pet use may lead to substantial antiviral use and healthcare resource utilization. limited real-world data are available on the outcomes of pet. therefore, we aimed to examine cmv outcomes (eod, resistance), cmv-related mortality by day (d) and healthcare resource utilization between pet and no-pet groups among cmv r+ recipients undergoing first hct. methods: we conducted a retrospective cohort study of adults, cmv r+ recipients of first peripheral blood or marrow allograft at mskcc identified from march through december . data was extracted from electronic medical records and hct databases. cmv+ recipients were monitored weekly by quantitative pcr assay starting on d through d post hct. use of antiviral therapy for cmv viremia defined pet. high cmv risk (hr) comprised t-cell depleted (tcd) hct by cd +-selection regardless of donor hla match or conventional hct from mismatched or haploidentical donors; low risk (lr) included conventional hct from matched related donors. cmv eod was scored by standard criteria. cmv resistance mutations were confirmed by sequencing (viracor-eurofins). length of stay (los) for hct admissions and readmissions were identified through d . stratified analyses were performed to examine outcomes by pet use and cmv risk. background: in a phase iii randomized, double-blind, placebo-controlled study of cmv-seropositive post-hsct recipients, letermovir prophylaxis significantly reduced the incidence of clinically significant cmv infection through week . the objective of this research was to assess the impact of cmv prophylaxis on rates of rehospitalization in adult cmv seropositive allogeneic hsct recipients from the letermovir phase clinical trial. methods: rehospitalization was recorded as an exploratory endpoint in the clinical trial at end of treatment (week ), time of primary endpoint (week ) and through an extended follow-up period (week ). cmv-related rehospitalization was assessed in the trial. prespecified analyses describe the observed rates of rehospitalization for the letermovir and placebo groups at the specified times. fine-gray cumulative incidence function(cif) regression models were used to explore the rate of all-cause, and cmv-related rehospitalization accounting for the competing risk of mortality. a multiple linear regression model was used to describe the cumulative length of stay (los) for all-cause rehospitalizations that occurred through week (excluding time of initial transplant stay). results: observed rates of all-cause rehospitalization were lower for the letermovir group compared to placebo at end of treatment ( . %vs. conclusions: letermovir was shown to significantly reduce the rate of clinically significant cmv infection in a placebo-controlled randomized clinical trial. these analyses suggest that there is also a reduction in the rate and cumulative days of rehospitalization. this trial was not sufficiently powered to detect differences in this exploratory endpoint. nonetheless, these data provide valuable insights into the economic burden of cmv. real world data and findings from future clinical trials are needed to better understand the nature of the association between cmv and rehospitalizations. clinical methods: all consecutive patients with hematologic disorders who received hsct at our center between january and august were included. among the evaluable patients, received levofloxacin as antibacterial prophylaxis (group a) while did not receive any fq prophylaxis (group b). baseline characteristics were similar in the two groups, except for the number of patients with advanced disease ( % in group a and % in group b, p , ). median duration of neutropenia was days (range - ) in group a and days (range - ) in group b. a positive rectal swab for carbapenem-resistant enterobacteriaceae (cre) was detected in patients in group a and patients in group b. results: overall, bsi was detected in patients ( , %), ( , %) in group a and ( , %) in group b (p= , ). the median onset of bsi was days post transplant (range - ), without significant differences between the two groups. in univariate analysis, fq prophylaxis (or , ; % ic , - , ) and bone marrow stem cell source (or , ; % ic , - , ) were significant factors associated with the risk of bsi. gramnegative bacteria accounted for , % (n= ) of bsi in group a and , % (n= ) in group b, and gram-positive bacteria for , % (n= ) of bsi in group a versus , % (n= ) in group b, without statistically significant differences (p = , ). polymicrobic bsi were , % (n= ) in group a and , % (n= ) in group b. mdrgram negative bsi were detected in patients ( %) in group a and in patients ( , %) in group b (overall, cre, esbl producing enterobacteriaceae and mdr-pseudomonas). death attributable to bsi occurred in of patients ( , %); of these patients did not receive fq prophylaxis, but of them had both a pre transplant kpc colonization and active disease at transplant. neither antibacterial prophylaxis (p = , ) nor bsi (p = , ) had a significant impact on overall survival (os). conclusions: the preliminary data of our study show that fq prophylaxis is associated with a reduced incidence of bsi, in particular gram-negative infections, with no impact on os. the limitations of our study may be the different group sizes and the retrospective nauture of the study. whether antibacterial prophylaxis should be avoided in the pre-engraftment period in still a matter of debate and needs to be evaluated in larger prospective studies. disclosure: nothing to disclose. gillen oarbeascoa , nieves dorado , , laura solan , , rebeca bailen , , pascual balsalobre , , carolina martinez-laperche , , ismael buño , , javier anguita , , jose luis diez-martin , , , mi kwon , hospital general universitario gregorio marañón, hematology, madrid, spain, instituto de investigación sanitaria gregorio marañón, madrid, spain, universidad complutense de madrid, madrid, spain background: incidence and outcome of invasive fungal infection (ifi) are not well characterized in the setting of peripheral blood, non-manipulated haploidentical stem cell transplantation with postransplant cyclophosphamide (haplosct). the aim of the study was to analyze incidence and risk factors of ifi in patients who underwent haplosct at our institution. methods: consecutive patients who underwent peripheral blood haplosct with postransplant cyclophosphamide between and at our centre were reviewed. ifi was classified according to eortc definitions. proven and probable ifi were included. results: patients´characteristics are shown in table . primary antifungal prophylaxis was performed with micafungin from day - until oral intake, followed by posaconazole until day + . patients on steroid treatment for gvhd received prophylaxis with micafungin or posaconazole. % of patients obtained neutrophil engraftment. twenty-two episodes of ifi were observed in patients: proven and probable, with a cumulative incidence of ifi of % at days. most commonly isolated organism was aspergillus spp (n= ), followed by candida spp (n= : c. kruseii and c. parapsilosis), and fusarium spp (n= ). isolated cases of inonotus spp, mucor spp and trichosporon ashii were observed. pulmonary involvement was the most frequent clinical presentation (n= ), followed by fungemia (n= : candidemia, trichosporon ashii) and skin-pulmonary involvement (n= ). among patients with lung involvement, showed probable ifi: with elevated serum galactomannan and positive galactomannan in bronchoalveolar lavage (bal). there were patients without galactomannan, one with a positive bal culture for penicillum spp and the other with an aspergillus spp. median time to ifi diagnosis was days. thirteen cases were diagnosed in the pre-engraftment period, after engraftment and cases after day + . among patients with late ifi, median time to development was days. all of them were associated with gvhd ( grade iii-iv acute gvhd and moderate/severe chronic gvhd). ifi outcome was favorable in out of the ifi. treatment was liposomal amphotericin b in cases, voriconazole in and combined treatment (amphotericin b and azole) in . there were ifi related deaths, with a cumulative incidence of ifi related death of . %. prior transplant (or . , p< . ), particularly allohsct was associated to ifi development (or . , p< . ). patients with previous allohsct presented ifi mainly from molds: aspergillus, fusarium, inonotus, trichosporon and mucor. there were also candidemia episodes. no other factors were significantly associated to ifi occurrence. conclusions: in our experience, cumulative incidence of ifi in the setting of haplosct with posttransplant cyclophosphamide was similar than observed in previous studies in allosct. having received a previous sct, especially allosct, was the most significant factor related to ifi development. this high risk population should be closely monitored and could benefit from prophylaxis with azoles. disclosure: nothing to declare. methods: rsv infection was diagnosed in nasal wash (nw) or bronchoalveolar fluid (bal) by dfa (millipore, usa) or pcr (seeplex, seegene, kor). urti and lrti were defined according to ecil- guidelines. death from all causes was assessed within days after rsv infection and was attributed to rsv if the patient had persistent or progressive rsv infection with respiratory failure at the time of death. neutropenia and lymphocytopenia were defined as an absolute neutrophil count (anc) < /ul and absolute lymphocyte count (alc) < /ul, respectively. results: median number of confirmed rsv infections per year was , ranging from to . an outbreak of rsv was detected in , possibly due to a lack of compliance with contact precautions in the unit. median patients' age was years and time to rsv infection was day (- to ). twenty-three patients (pts) had received an autologous transplantation ( . %) and were allogeneic hsct recipients ( , %). median time to engraftment was days, ranging from to days. at rsv diagnosis, pts presented with urti ( . %) and with lrti ( . ) . surprisingly, around % of the auto hsct recipients had rsv pneumonia at diagnosis. variables significantly associated with lrti at diagnosis were mud hsct (no/ yes, or . ; ci . - . ); anc < /ul (or . ; ci . - . ); alc < /ul (or . ; ci . - . ); and recent or pre-engraftment hsct (no/yes, or . ci . - . ). among the pts with urti at diagnosis, progressed to lrti ( . %). forty-four of the pts died ( . %) and mortality rate was significantly higher in pts with lrti in comparison with pts with urti ( . % versus . %, p= . ). death was attributed to rsv in of the pts who died ( %). conclusions: autologous hsct recipients are also at risk of lrti caused by rsv. risk of rsv lrti is higher in mud hsct, infection acquired pre-engraftment or early after hsct, and low neutrophil and lymphocyte counts. continued education is necessary to sustain compliance to contact precautions in hsct units. disclosure background: measles is a life-threatening infection after allogeneic hct. due to the decreased coverage of vaccination in many countries, the disease reappears, increasing the risk of outbreaks worldwide. allogeneic hct recipients have been shown to be seropositive for measles in roughly - % of the cases years after transplant. however, these data were obtained before the 's from hct populations mainly conditioned with myeloablative (ma) regimens. our aim was to assess measles immunity before considering vaccination in a cohort of hct survivors including patients conditioned with reduced intensity (ric) or non-ma regimens. methods: allogeneic hct adult recipients who had not been vaccinated for measles since hct were routinely screened for measles immunity. measles igg titers were determined with a chemiluminescence immunoassay (liaison measles igg kit, liaison xl analyser, diasorin, italy). patients were considered to be seropositive if the igg titer was > . ua/ml. risk factors for seropositivity were analyzed. qualitative variables were described as numbers (%) and compared using the chi- test or fisher exact test as appropriate. quantitative variables were described as median or mean (range) and compared using the kruskall-wallis test. ors were estimated separately for factor yielding a p-value < . in the univariate analysis using logistic regression models. results: eighty-six patients, transplanted . to years (mean: , years) ago, were included. the mean age was years (range: - ), the sex ratio m/f: , . the underlying diseases were acute leukemia: ( %), myelodysplastic syndrome: ( %), lymphoproliferative diseases: ( . %), myeloproliferative neoplasms: ( %) and non-malignant diseases : ( . %). the hct was performed from an hla-identical donor in , an unrelated donor in , and a cord-blood in . conditioning regimen were ma in ( %), ric in ( %) and non-ma in ( %) patients no patient had experienced measles or had received measles vaccination since transplant. fifty-seven of the ( %) patients were seropositive for measles. measles seropositivity was not associated with conditioning regimen, patient age at transplant, patient age at time of assessment, donor age at transplant, lymphocyte count or gammaglobulin levels, or type of transplant (hlaid. vs others) measles vaccination before transplant or previous measles before transplant. the only parameters significantly associated to seropositivity were absence of previous gvhd (any type or severity, p= , or , [ , - , ]), and absence of previous extensive chronic gvhd (or , [ , ] p , ). conclusions: sixty-seven percent of allogeneic hct are seropositive for measles at a median of years after hct before vaccination. the only risk factor strongly associated with seronegativity is extensive chronic gvhd. in patients background: cytomegalovirus (cmv) reactivation is a frequent complication after hematopoietic stem cell transplantation (hsct). extracellular vesicles (evs) have emerged as a promising new category of biological biomarkers in different scenarios, including inflammation, tissue damage, cancer and viral infections. we recently reported on the potential use of serum evs as biomarkers of agvhd (lia g. et al. leukemia ( ) , ) . here, we investigated the potential correlation of cmv reactivation with plasma evs in post-transplant cyclophosphamide (ptcy) haploidentical-hsct (haplo-hsct). methods: plasma samples were collected after mononuclear cell separation at given time-points (pre-transplant, on day , , , , , , , , , and after haplo-hsct) and evs were extracted by a protamine-based precipitation method and their concentration and dimension were characterized by nano-tracking particle analysis (nanosight). after extraction, evs were analyzed by flowcytometry (guava easycyte flow cytometer) with a panel of antibodies (cd , cd , cd , krt , cd a, cd , cd , cd , cd , cd , cd , cd , cd , and cd a). results: thirty-two patients with hematological malignancies underwent haplo-hsct between and . cmv reactivation was observed in / ( , %) and occurred at a median of (range: - ) days after transplant. preliminary analysis ( / patients) showed that cd a fluorescence (platelet-derived growth factor receptor-α or pdgfr-α), cd fluorescence (ki- antigen) and cd fluorescence (ve-cadherin) were associated with an increased risk of cmv reactivation (or . p= . ; or . p= . ; or . p= . ), whereas cd (platelet endothelial cell adhesion molecule, pecam- ) concentration level was associated with a decreased risk of cmv reactivation (or . , p= . ). all these biomarkers showed a signal change before cmv reactivation (an increase with cd a, cd and cd , a reduction with cd ). (figure ). conclusions: we observed a potential association of evs membrane proteins with cmv reactivation: cd a, cd , cd and cd . these proteins are crucial for endothelium and immune cells interaction. cmv can infect different cell types including endothelial cells (bentz gl. pnas ( ) ). moreover, cd a (pdgfr-α) has been shown to function as an entry receptor for cmv expressing gh/gl/go complex (wu y. et al. plos pathog ( ) ). we plan to implement our analysis characterizing evs contents (mirnas) and will be applied to investigate other viral reactivations (e.g. epstein barr virus and human herpes virus ). [[p image] . methods: to explore the value of cmv dna extracted from gi tissue for the diagnosis of cmv gastroenteritis, we retrospectively evaluated patients, aged - (median . years) who received allo-hct from sibling( ), matched unrelated( ) or haploidentical donors( ), after receiving myeloablative ( ) or reduced intensity conditioning( ). they all underwent endoscopy for gastrointestinal symptoms between - . cmv dna from tissue samples and parallel blood samples were measured by q-pcr. positive cmv dna on the tissue was considered cmv gi infection.cmv gi disease was proven with the identification of cmv inclusion bodies or positive immunehistochemical staining using anti-cmv antibodies. results: overall, endoscopic tests were performed ( gastro-, colonoscopies) at a median of days (iqr: ) post transplantation. symptoms included nausea, vomiting, diarrhea, abdominal pain and weight loss. cmv dna was positive in / tissue samples: median copies/ml, range: - x ^ . only half patients ( / ) had concurrent cmv viremia (plasma viral load> c/ml). cmv gi infection was not correlated to the type of transplant, acute or chronic gvhd. gi cmv disease was documented by biopsy in patients. cmv dna of the tissue, but not the plasma viral load, was a predictor of biopsy positivity (or: . , %ci: . - . , p= . ). thirty-six out of cmv dna positive patients received specific treatment for at least days. symptoms resolved in / patients ( %) and the gi viral load was not a significant factor to predict cure. gi gvhd was diagnosed in / patients, among which %( / ) with cmv dna positivity. median os was days ( %ci: - ) for patients with cmv infection, similar to those without (median os: , %ci: - days, p=ns). we studied separately endoscopies of the upper ( / ) or lower gi tract ( / ) . there was no significant relationship between cmv gastritis proven by biopsy and cmv dna levels in gastric tissue. however, the viral load of the colon was a predictor of cmv enteritis (or: . , %ci: . - , p= . ). the auroc of the q-pcr was . ( %ci: . to ), the sensitivity was . % and the specificity was . % with a cutoff value of copies/ml dna. conclusions: pathognomonic findings in the biopsy remain the gold standard for the diagnosis, especially for the upper gi tract. however, when the lower gi tract is involved, quantification of cmv viral load in the tissue may be a valuable tool to support the diagnosis. positivity of cmv dna of the gi tissue, in linearity to the cmv viremia, may guide to preemptive treatment for prevention of cmv disease . disclosure: nothing to declare background: clostridium difficile infection (cdi) is caused by cd overgrowth in antibiotic-disturbed intestinal microbiota. antibiotics targeting unselectively beneficial for t-regulatory cell formation strains of clostridiales may increase pro-inflammatory processes in the guts promoting or augmenting the development of graft vs. host disease (gvhd). the efficacy of cdi treatment has impact on the persistence of inflammation which might influence the alloreactive reactions. methods: we retrospectively and, from , prospectively analyzed the data from transplant centers concerning cdi occurrence, treatment efficacy, and gvhd development. the study included patients with hematological malignancies who underwent allogeneic hematopoietic cell transplantation (allohct) between - . results: median time to cdi was days post-allohct with detection of both toxins a and b in % of cases. disturbance of intestinal microbiome was confirmed by a % rate of colonization with multidrug-resistant bacteria (mdrb). the cdi symptoms resolved with the negative toxins after the first line treatment in . % of patients. the median time to remission and therapy duration was and days, respectively. fifteen therapeutic failures were observed after treatment with metronidazole ( ), vancomycin ( ) and a combination therapy ( ) . eleven patients responded to second line treatment. thirty-seven ( %) patients died due to infections ( ), relapses ( ) and gvhd/infections ( ). we noted recurrent cdi in cases. eight patients died with active cdi. we observed occurrence or exacerbation of gvhd in ( %) patients following cdi, including cases with gut involvement (gi-gvhd). treatment with metronidazole and failure of the first line therapy increased the development or escalation of gi-gvhd (p= . and p< . , respectively). the duration of cdi exceeding days also had impact on the gi-gvhd incidence (p= . ). conclusions: . patients colonized with mdrb are at high risk of cdi. . high mortality due to infections and/or gvhd in patients with cdi. . due to lower efficacy and harmful immunomodulatory impact, metronidazole should not be the first line treatment in cdi post-hct. . emphasis must be put on fast cdi resolution to interrupt a vicious circle of the intestinal inflammatory processes. disclosure: nothing to declare establishing optimal preemptive cytomegalovirus therapy threshold post allogeneic hct in a patient population with high prevalence of seropositive status background: preemptive therapy (pet) for cytomegalovirus (cmv) reactivation post allogeneic hematopoietic stem cell transplantation (hct) was shown to decrease the incidence of cmv disease. however, the optimal pet threshold is unknown and there are significant toxicities associated with anti-cmv therapy. at our institution, we initiate pet at cmv dna titer above copies/ml ( iu/ml). our aim was to examine the efficacy of this approach including the incidence of spontaneous clearance in a population with high prevalence of cmv seropositive status. methods: after due irb approval, patients that underwent allogeneic hct were identified and records retrospectively extracted.cmv reactivation was defined as the first detectable viral titer post hct from plasma samples whereas clearance of viremia as the first date of two negative pcr values obtained at least week apart. cmv monitoring was initiated post hct performed at least weekly during the first days and every - weeks thereafter. a high sensitivity assay abbott realtime cmv was used with detection threshold of copies/ml ( . iu/ml). analysis was computed using jmp v. . . results: a. baseline characteristics: a total of patients were identified and included with a median follow up of . ( . - . ) months. median age was ( - ) years and % were male. indication for hct was for a malignant disorder in % of cases. the majority had a matched related donor ( %) and cmv igg was positive in both donor and recipient in % of cases. myeloablative conditioning was given to ( %) and ( %) received tbi. in vivo t-cell depletion was given to ( %); atg in ( %) and alemtuzumab in ( %). b. cmv reactivation and pet: a total of ( %) patients had a positive cmv pcr with median days to reactivation post hct of ; ( %) patients had peak cmv titer < copies/ml (low titer) whereas the remaining ( %) had a peak titer ≥ copies/ml (high titer). patients with high titer were more likely to be older (p = . ), have malignant disease (p = . ), haploidentical or unrelated donor (p < . ) and higher incidence of agvhd grade ii-iv (p = . ) as shown in the table. median peak titers for the low and high groups were vs. , respectively (p < . ). ( %) patients with low titers cleared spontaneously with median time to clearance of days ( - ), ( %) received anti cmv therapy and the remaining died with active viremia (range - copies /ml) with active disease. one patient in the high titer group developed cmv disease. -year os and ci-nrm was . % vs. . % (p = . ) and % vs. . % (p = . ) in the low and high titer groups, respectively. conclusions: cmv reactivation was high in this cohort however of low titer viremia in over %. a pet threshold of copies/ml ( iu/ml) appears desirable as it was associated with spontaneous clearance in almost all patients while minimizing treatment related toxicity. validation of these observations is warranted. background: the risk of pneumocystis pneumonia often warrants antifungal prophylaxis for recipients of blood and marrow or solid organ transplantation. however, complications such as myelosuppression, nephrotoxicity, and intolerance with the existing standard, trimethoprim/sulfamethoxazole (tmp/smx), may hinder or interrupt prophylaxis. rezafungin (rzf) is a novel echinocandin in development for prevention of invasive fungal disease caused by candida, aspergillus, and pneumocystis species in blood and marrow transplant patients. rzf has a favorable safety and tolerability profile and a low risk of drug-drug interactions. furthermore, the stability and pharmacokinetics of rzf allow for once-weekly dosing and broad distribution to the lung and other target organs. rzf was shown to prevent in vitro pneumocystis biofilm formation and to reduce the viability of mature biofilms. a previous prophylactic study was conducted using a broader range of rzf doses. in the current study, the efficacy of rzf was evaluated to better understand the minimum doses necessary to prevent pneumocystis growth in a mouse model. methods: c h/hen mice were immunosuppressed (dexamethasone mg/l in acidified drinking water) and then infected intranasally with p. murina ( x / μl). given the slow growth of p. murina, test agents were administered at the same time mice were inoculated to test for prophylactic efficacy. mice received intraperitoneal injections of either vehicle (control/steroid [c/s]), tmp/ smx / mg/kg/ x/week (wk), caspofungin mg/kg once daily, or rzf mg/kg or . mg/kg once daily, x, or x/wk. after a -week dosing period, mice were sacrificed and lung homogenates were processed for analysis to quantify the nuclei (trophic) and asci (cyst) forms of pneumocystis. prophylaxis efficacy was based on reduction of organism burden compared with c/s. nuclei and asci counts were log transformed and analyzed by anova; individual groups were compared by the student-newman-keuls t test. survival rates were compared using graphpad prism v . results: all mice in the rzf groups had significantly reduced nuclei and asci burdens compared with the c/s group, and all but the lowest doses of rzf ( . mg/kg x or x/wk) worked as well as tmp/smx at reducing nuclei levels. similarly, all rzf groups except for the . mg/kg x/wk group showed reductions in asci levels comparable to that of tmp/smx. the survival rates were not statistically different between treatment groups. conclusions: rzf demonstrated potent in vivo efficacy for prophylaxis against pneumocystis in an in vivo mouse infection model at dose regimens much lower than the human equivalent phase regimen. these data support the development of rzf for the prevention of invasive fungal infections including pneumocystis pneumonia. disclosure: melanie t. cushion: research funding (cidara therapeutics) taylor sandison: employee, stockholder (cidara therapeutics) alan ashbaugh: nothing to declare. yuhua ru , , ziling zhu , , yang xu , , suning chen , , xiaowen tang background: immunocompromising period following allogeneic hematopoietic stem cell transplantation (allo-hsct) may allow opportunistic pathogens to thrive and result in fatal complications. epstein-barr virus (ebv) infects more than % of chinese population, and its reactivation after hsct is one of the major concerns due to the increased risk of ebv diseases and post-transplant lymphoproliferative disease. with the development of infection prophylaxis and supportive care after hsct, demographic data on ebv reactivation post-hsct needs to be updated. methods: we retrospectively analyzed the data of patients who received allo-hsct between july and july in the first affiliated hospital of soochow university. quantitative pcr (q-pcr) was used to monitor ebv-dna load in peripheral blood dynamically. ganciclovir (pre-hsct) followed by acyclovir was given as viral prophylaxis. the treatment protocol for ebv reactivation consisted of tapering of immunosuppressive agents, antiviral agents (including ganciclovir and sodium phosphonatel), and rituximab for persistent positive patients. results: totally cases from most of the provinces in china were enrolled (characterized in table ), among whom ebv reactivation developed in recipients. most reactivation events ( . %) occurred in the first year post-hsct, with a peak of . incidence rates per personyears at the second month. besides, more episodes of lateonset reactivation occurred in patients receiving grafts from haploidientical donors ( figure a ) . multivariate analyses revealed that the major impactors of ebv reactivation included atg as gvhd prophylaxis (p< . ), hlamismatched donor (p= . ) and the appearance of chornic gvhd (p= . ). cumulative incidence of ebv reactivation was low ( . %) among patients with no major risk factors, but increased to . %, . % or . % with , , and major risk factors, respectively ( figure b) . there was no statistical difference of overall survival between people with or without ebv reactivation (p= . ). conclusions: we concluded that there are similar ebv reactivation impactors in chinese population compared to literatures, including atg use, hla-mismatched donor and the appearance of chronic gvhd. additionally, incidences of ebv reactivation increased significantly with the accumulation of risk factors. however, ebv reactivation had no impact on overall survival in current virus management protocol. disclosure: nothing to declare background: several studies have shown loss of diversity of the gut microbiome in association with significant gut injury following hematopoietic stem cell transplantation (hsct). prolonged broad spectrum antibiotic use further promotes loss of microbiome diversity and increases the risk of intestinal colonization by multi-drug-resistant (mdr) bacteria. aims of this study were to prospectively evaluate the overall changes in gut microbiome composition after hsct and differences in patients colonized by mdr bacteria and treated with carbapenems. methods: we performed a prospective observational study evaluating the gut microbiota of hematological patients undergoing hsct, from admission (t ) through day + (t ). fecal microbiota was assessed by s amplicon-based sequencing. clinical, and microbiological data as well as fecal samples were collected every th day from admission. results: one-hundred fecal samples were analyzed. overall, we found a progressive decrease of bacterial richness from t to t , with a significant reduction of blautia, ruminococcus and dorea species, which are strictly associated with the production of short chain fatty acids (sca) (fig. ) . moreover, in the % (no. ) of patients who were colonized by esbl bacteria, we observed a significant reduction of clostridium spp and bifidobacterium species. as for antibiotic therapies, carbapenems were used as second line treatment of febrile neutropenia in % (no ) of cases, usually associated with aminoglycosides. in patients treated with meropenem, a strong decline of blautia and ruminococcus species was observed. this finding suggests a correlation between carbapenem regimens and increase of pro-inflammatory bacterial strains in the gut. conclusions: our data support the hypothesis that loss of intestinal commensals that produce short-chain fatty acids may increase dysbiosis. moreover, for the first time we report significant and progressive alterations in the composition of blautia, ruminococcus and bifidobacterium species in patients treated with meropenem and colonized by esbl bacteria, respectively. our findings offer potential modifiable targets to reduce risk of colonization by mdr bacteria and to promote a carbapenem-sparing approach in the hsct setting. clinical background: cmv is associated with significant morbidity after allogeneic hematopoietic stem cell transplantation. strategies to prevent cmv-related complications include universal prophylaxis and preemptive therapy, more widely spread. antivirals used for cmv reactivation (cmv-r) produces major toxicities and costs. rate and characterization of cmv-r after haploidentical transplantation with post-transplant cyclophosphamide (haplo pt-cy) is scarce. our goal was to analyze cmv-r rate after haplo pt-cy, outcome, complications associated to therapy, and to identify risk factors. methods: one hundred haplo pt-cy transplants using peripheral blood as stem cell source performed between and in our center have been retrospectively reviewed. gvhd prophylaxis consisted of pt-cy mg/ kg/day on days + and + , mmf and csa from day + for all cases. cmv pcr was performed in a biweekly basis during admission for transplant and treatment, and weekly thereafter. cmv-r was considered with any cmv dna level by pcr assay above copies/ml. prior four consecutive negative weekly pcrs were needed to consider a new reactivation episode. preemptive strategy was applied in all cases. data collected in relation to cmv-r included: cmv serostatus of donor/recipient (d/r), number of cmv reactivations, length of each reactivation, antiviral treatment used, need for admission to receive treatment and adverse events related to cmv reactivation and/or antiviral treatment. results: patients characteristics are summarized in table . among patients, of them with positive cmv serology, episodes of cmv-r were detected. seventysix patients ( %) had at least one cmv-r in a median of days after transplant. none of them had cmv disease or die as a consequence of cmv-r. median duration was days ( - ). valganciclovir or ganciclovir was used in episodes ( %). foscarnet was used in episodes ( %). six of the episodes occurred after initial discharge, and required re-admission for treatment, with a median length of hospitalization of days ( - ). cytopenias requiring transfusion or g-csf support occurred in episodes ( %) treated with ganciclovir or valganciclovir. three of them needed further cd + cells booster for graft rescue. mild acute renal failure and genital ulcers were found in ( %) and ( , %) events treated with foscarnet, respectively. no cases of severe renal failure were observed. serological status different than negative/negative (n/n) (p . ) and older age ( vs years, p . ) were significantly associated with cmv-r. no relationship was observed with gender, disease, donor relationship, conditioning, gvhd or cells infused. more than reactivations were more frequent among patients with grade ii-iv acute gvhd (agvhd) and moderate-severe chronic gvhd (cgvhd). conclusions: in our experience, rate of cmv-r after unmanipulated haplo pt-cy, using pbsc as stem cell source, is considerably high. a significant proportion of patients presented complications associated with cmv-r and its treatment. cmv serological status other than n/n and older age are associated with high risk of cmv-r. patients with grade ii-iv agvhd are at higher risk of multiple reactivations. this population could be benefited from primary prophylaxis, in order to decrease treatment´s complications, re-admissions and costs. disclosure: nothing to declare. impact of infectious events occurring during the first hundred days after hsct for hematological malignancy: a monocentric retrospective study over a five-year period marie-pierre ledoux , célestine simand , , karin bilger , annegret laplace , bruno lioure background: patients undergoing hematopoietic stem cells transplantation (hsct) for hematological malignancy often present with infectious events in the early stages of the procedure, some of which having a documented impact on the outcome of the graft. for instance, cytomegalovirus (cmv) has been shown by some authors to have a protecting effect against relapse, whose features remain to be elucidated. we conducted a retrospective monocentric study regarding the outcome in terms of graft versus host (gvhd), relapse and survival of consecutive patients over a period of years, whether they presented or not with an infectious event by day among the following: cmv viremia, epstein-barr virus (ebv) viremia, human herpes virus (hhv ) viremia, bk virus (bkv) viruria, bacterial bloodstream infection (bsi) or invasive fungal infection. results: a high proportion of cmv seropositive recipients underwent a viral reactivation of cmv by day of the hsct: % if the donor is seronegative and % if the donor is seropositive. we observed that cmv wasn't associated with a lower relapse rate in our cohort, and data weren't sufficient to conclude firmly, but showed a trend towards a worse acute gvhd (hazard ratio hr . , pvalue . ). no significant correlation was found for ebv viremia. occurring in % of our patients and mostly with an early timing, hhv strongly correlated with worse acute gvhd (hr . , p-value < . ) but its impact on survival was not significant. bkv ( % of our patients) and bsi ( % of our patients) both correlated with poorer outcome in terms of overall survival (logrank < . and . respectively) although not significantly associated with relapse or acute gvhd. fungal infections were too rare events to draw any conclusion. conclusions: thus, contrary to many studies, we found no protection against relapse induced by cmv, although the trend for worse acute gvhd was obvious. the mechanisms behind this discordance could include early treatment, but remain to be studied. whether hhv is a cause rather than a consequence of acute gvhd or its treatment is debated, but the correlation is strong and the sequence of events suggests hhv might act as a trigger for gvhd. the association between bkv viruria and a higher mortality is in contrast with previous observations, and the lack for association with gvhd and relapse could suggest bkv is a surrogate for poor immune recovery and therefore other causes of non-relapse mortality. in addition to the direct lethal risk of bacteriemia, bsi also are a promoter of late non-relapse mortality through indirect toxicity. through the expansion of immune effectors they promote, one could assume that infectious events play a role in gvhd and gvl, and therefore have an interference with relapse. however, the association between each infectious event and outcome remains to be clarified to guide our prophylactic and therapeutic choices by a better understanding of the bright and dark sides of infectious events. disclosure background: rezafungin (rzf) is a novel echinocandin in phase development for treatment of candidaemia and invasive candidiasis and for antifungal prophylaxis against invasive fungal diseases caused by candida, aspergillus, and pneumocystis in blood and marrow transplant patients. rzf is differentiated by stable, prolonged pharmacokinetics (pk) that allow for once-weekly dosing and a pk-pharmacodynamic (pd) profile correlating with efficacy. clinical in vivo evaluations of drug interaction potential were performed proactively to assess the risk of drug-drug interactions (ddis) with respect to the phase dose of mg once weekly and known pk exposure in healthy individuals. methods: this open-label study of healthy inpatients assessed ddis between rzf (as perpetrator) and drugs known to have interactions with cyp enzymes and transporters (probe drugs): repaglinide (cyp c ), metformin (oct/mate), rosuvastatin (bcrp/oatp), pitavastatin (oatp), caffeine (cyp a ), efavirenz (cyp b ), midazolam (cyp a ), and digoxin (p-gp), as well as tacrolimus, a drug likely to be coadministered with rzf. an initial dose of rzf mg was administered on the first dosing day, to approximate a steady state plasma concentration of multiple once-weekly -mg doses, followed by once-weekly -mg doses on days and . probe drug cocktails containing ≥ drugs were administered, once before and once after rzf administration, on a schedule designed to allow for washout between doses and to limit interactions with other probe and test drugs. samples were analysed to determine respective drug concentrations in plasma (except for tacrolimus which was in whole blood) to characterize the pk profile of each analyte. area under curve (auc) and maximum concentration (c max ) were calculated from the plasma/blood concentration-time profiles by noncompartmental analysis. ln-transformed pk parameters were statistically analysed using an analysis of variance model. the ratio of geometric least squared means between each substrate drug when administered with and without rzf and corresponding % confidence intervals (cis) were calculated for lntransformed c max and auc. results: when rzf was given concomitantly with the probe drugs, six of nine substrates (metformin, pitavastatin, caffeine, efavirenz, midazolam, and digoxin) statistically demonstrated the absence of drug-drug interaction, as their % ci were all included within the default - % noeffect boundary. three substrates had the upper (repaglinide and rosuvastatin) or lower (tacrolimus) bounds of their ci falling just outside of this range (figure ), and these changes are considered unlikely to be clinically significant. conclusions: no meaningful pk interactions were observed between rzf and drugs known to have ddis and/or likely to be coadministered with rzf. these findings provide evidence that no dose adjustment is expected when rzf is co-administered with these commonly used drugs, which stand in contrast with the ddi complications widely associated with azole antifungals. disclosure: voon ong: employee, stockholder (cidara therapeutics), michael boily: employee (altasciences), hong wong: employee (altasciences), taylor sandison: employee, stockholder (cidara therapeutics), shawn flanagan: employee, stockholder (cidara therapeutics) abstract withdrawn. background: cytomegalovirus (cmv) continues to cause morbidity following allogeneic hematopoietic stem cell transplantation (hsct). letermovir is a newly approved drug for cmv prophylaxis in cmv-seropositive allogeneic hsct recipients. however, there is a paucity of data for its efficacy in patients receiving in-vivo t-cell depletion (tcd). at weill cornell medical center, we perform in-vivo tcd with alemtuzumab for related and hla-identical unrelated transplants, and anti-thymocyte globulin for umbilical cord blood transplant supported by third party accessory cells (haplo-cord transplant).although these drugs reduce the frequency of graft-versus-host-disease (gvhd), they significantly delay t-cell immune reconstitution post hsct, and may cause higher rates of cmv reactivation. our historical rate of cmv reactivation in cmv seropositive recipients receiving high dose valacyclovir prophylaxis is approximately %. therefore, we implemented letermovir for cmv prophylaxis in february . the primary aim of this study is to determine the incidence of cmv infection (defined as cmv viremia warranting treatment or development of end-organ disease) in tcd cmv seropositive allogeneic hsct patients who received letermovir prophylaxis. methods: this is a single center, retrospective cohort study to determine the incidence of cmv infection in adult, cmv-seropositive recipients receiving letermovir prophylaxis after in vivo tcd hsct with atg or alemtuzumab for gvhd prophylaxis. all included subjects were at least days post-transplant. results: allogeneic hsct transplant recipients met inclusion criteria. median age was years, iqr [ , ] and % were male. eight ( %) had a matched related donor, six ( %) had a matched unrelated donor, and ( %) were haplo-cord transplants. their underlying malignancy and conditioning regimens are summarized in table . ( %) received atg and ( %) received alemtuzumab for gvhd prophylaxis. median follow up time for survivors is days, iqr [ , ] . the incidence of cmv infection in the first days post-transplant was % as only one patient reactivated with detectable cmv viremia. this same patient developed cmv pneumonitis with documented ul resistance, and was successfully treated with ganciclovir. the incidence of cmv infection within the first days post-transplant was % ( / patients) . six patients ( %) developed acute gvhd in the first days, and one ( %) had relapse of their malignancy. five patients ( %) died within days post-transplant, but none of these deaths were cmv related. background: infectious complications caused by endogenous adenovirus (adv) are common and associated with morbidity and mortality rates in patients after hematopoietic stem cell transplantation (hsct). adv infections occur in about % to % of hsct recipients, with significantly higher rates in pediatric patients. a better understanding of adenoviral-specific t-cells (advt) response in donors can serve as a basis to develop more effective strategies for antiviral therapy. methods: frequencies of cytomegalovirus (cmv)-and adv-specific t cells were determined by enzyme-linked immunospot (elispot) assays with adv hexon and cmvpp respectively in health donors. we used x of mononuclear cells (mnc) per well in elispot assays. all donors were divided into groups according to the number of spots per well (spw) as follows: high responders (hr) (≥ spots; n= ), low responders (lr) (> and < spots; n= ), nonresponders (nr) (≤ spots; n= ). the average spot area of adv-and cmv-specific lymphocytes was calculated by immunospot® multiplate autocount™. cd ra+ and cd ro+ t-cells were generated by immunomagnetic negative selection. hla typing for class i and ii was performed by sequence specific oligonucleotides technology. statistical analysis was performed using graphpad prism v . software. levels of significance were calculated by mann-whitney rank-sum test, expressed as p-values (p< . ). results: the median frequency per well of advts were in hr group, in lr group, in nr group. the median spw of cmv-specific t cells in donors mnc were and didn´t differ between groups. antiviral activity may depend not only on the amount of advt but also on their ability to produce ifnγ. the average spot area for advt did not differ between hr, lr and nr groups and were , , , and , mm respectively. the median of the average spot area for anti-cmv t-lymphocytes was equal to , mm . thus, the frequency of advt was lower than cmv-specific t-cells, but advt have the ability to produce more ifnγ per cell (p< . ). in order to evaluate the distribution of the advt between naive and memory t cell compartments, we evaluated response to adv in preselected cd ra+ and cd ro+ fractions of t-cells in a group of donors. the median frequency of advt in unfractionated mnc was ; the median frequency of advt in cd ra and cd ro fractions were and , respectively. the amounts of cd ra and cd ro tcells were normalized to their amounts in mnc. we evaluated the impact of hla-alleles on the anti-adv response of t-cells in different groups and found association: hla-a* with hr group (p-value= , ; rr= , ; % ci: , to , ) and hla-a* with lr group ; rr= , ; % ci: , to , ) . conclusions: in this study the frequency of donors with advt is , % which corresponds to the reported frequency of adv-seropositive people ( %) in population of russia. advt are exclusively cd ro-positive cells. the analysis of advt in potential hsct donors will allow to determine more accurately the amounts and functional activity of specific antiviral t-lymphocytes administered to patient and optimize antiviral therapy. disclosure: nothing to declare p abstract withdrawn. chemotaxis and exhaustion of γδ t cells in the allografts are associated with cmv reactivation after hematopoietic transplantation background: cytomegalovirus (cmv) reactivation and its related diseases remain the most common and serious complications in patients who underwentallogeneic hematopoietic stem cell transplantation (allohsct). we previously reported that the incidences of total and refractory cmv reactivation reached approximately % and % after haploidentical hsct. while majority of studies in the literatures focused on the adaptive cd + αβ t cellsand nk cells in anti-cmv immunity, increasing evidences highlighted the important role of γδt cells in this context. a progressive and prolonged expansion of vδ + t cells in response to cmv reactivation was observed after allohsct. the effect of vδ + t cells associated with cmv clearance has been reported in vitro and in vivo. in contrast to the reconstituted γδt cells post transplantation, whether the phenotypes of γδt subsets in allografts correlate to cmv reactivation in hsct recipients have not been documented. methods: the proportions and phenotypes of γδ t cells were detected inallografts those were unmanipulated g-csf-mobilized bone marrow (bm) and peripheral blood (pb) harvests from donors for haplohsct. bm grafts were collected by aspiration on the fourth day of g-csf treatment (filgrastim, μg/kg/day), and pb grafts were obtained on the fifth day by leukapheresis. immunophenotyping for γδ t-cell subpopulations, including the expression of cd , cd , cd , tcrγδ, tcrvδ , tcrvδ , hla-dr, nkg d, cxcr , ccr , pd , ki , ifnγ, tnfα, and il- , was performed using flow cytometry. for detection of the intracellular cytokines, bm and pb grafts were pre-stimulated with x cell stimulation cocktail ( x, ebioscience). cmv dna in the peripheral blood of recipients was routinely monitored by quantitative pcr. the association of γδ t-cell contents in allografts with cmv reactivation in haplohsct recipients was analyzed using the mann-whitney u test and spearman test. all calculations were performed using spss . statistical software. results: we found that the proportions of total γδ t cells, and vδ and vδ subsets in both bm and pb grafts for cmv+ and cmv-recipients were comparable. neither the expression of hla-dr nor nkg d in the allografts were significantly different in correlation to cmv reactivation after hsct. the productions of intracellular cytokines of γδ t subsets did not varied in bm and pb grafts for cmv+ and cmv-recipients. interestingly, the proportions of cxcr +vδ and ccr +vδ cells in bm grafts for cmv + recipients were significantly higher than those for cmvrecipients (p = . and . , respectively). meanwhile, pma-stimulated ki +vδ cells in bm grafts for cmv+ recipients were less than those for cmv-recipients (p = . ). in parallel, the concentration of pd +vδ cells in pb grafts for cmv+ recipients were significantly higher than those for cmv-recipients (p = . ). conclusions: this study is the first to connect the chemotaxis and exhaustion of γδ t cells in grafts to the risk of cmv infection after allogeneic hsct. future studies should explore how the expressions of chemokines and exhaustion marker on the effector γδ t cells in allografts facilitate cmv replication and/or dissemination in the setting of hematopoietic transplantation. disclosure: all authors do not have conflicts of interest. this study is supported by the national natural science foundation of china (grants no. and no. ) results: incidence of ic was , %: allo-hsct - % (n= ), auto-hsct - , % (n= ). the etiology: c. parapsilosis %, c. albicans %, c. krusei %, candida tropicalis %, candida dubliniensis %. the most frequent underlying diseases was acute leukemia - % (n= ). the median age was y.o. [ month - years] . the median day of onset of ic after allo-hsct was , auto-hsct - [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] . febrile fever was the main clinical symptom; septic syndrome develops in % cases. antifungal therapy was with echinocandins - %, lipid ampho b - %, azoles (fluconazole, voriconazole) - %, without therapy (the early mortality) - %. overall survival (os) at days from diagnosis of invasive candidiasis was %. the central venous catheter (cvc) removal was the only factor significantly improved os ( % vs %, p= , ). conclusions: incidence of invasive candidiasis in children after hematopoietic stem cell transplantation was . %. the main etiology agent was c. parapsilosis. invasive candidiasis infections most often affect leukemia patients, developed later after allo-hsct than auto-hsct. overall survival at days from the diagnosis was %. removing of cvc improved overall survival in children with invasive candida infections after hsct. disclosure: nothing to declare background: graft versus host disease (gvhd) and virusassociated enteropathy in allogeneic hematopoietic stem cell transplantation (allo-hsct) may cause severe quantitative and qualitative composition changes of intestinal microbiota, leading to the development of small intestinal bacterial overgrowth (sibo) on the background of immunodeficiency, which can have a negative impact on treatment effectiveness. the gold standard for diagnosis and the main criterion for sibo is the detection in the jejunum aspirate > /ml bacteria and/or the appearance of colonlike microbiota in small intestine. it is also acceptable to use an alternative non-invasive technique -hydrogen breath test, which could be especially important in patients with severe mucositis, grade iii-iv and thrombocytopenia grade iv. sibo diagnosis in the setting of the gastrointestinal tract damage and dysfunction in patients treated with allo-hsct is insufficiently studied. methods: the study included patients with acute myeloid leukemia (n= ), acute lymphoblastic leukemia (n= ), myelodysplastic syndrome (n= ), non-hodgkin´s lymphoma (n= ), hurler syndrome (n= ), who underwent allo-hsct from an unrelated (n= ) and haploidentical donor (n= ) , and which were complicated by enteropathy development. in cases, the enteropathy reason was a combination of intestinal gvhd and viral colitis (hhv- ), in cases -viral colitis (hhv- ). all patients had esophagogastroduodenoscopy with species aspiration from descending part of the duodenum and feces collection, with further bacteria pcr identification. hydrogen breath test was performed also in which patients were treated with oral lactose g/kg with subsequent hydrogen assessment after and minutes. the study was performed in the period from to days after allo-hsct. results: according to feces analysis data, colon microbiota composition significantly differed from the reference values. at the same time total bacterial mass of the duodenum was less in comparison with colon microbiota: e+ ( e+ / e+ ) and e+ ( e+ / e+ ), respectively (p< . ). quantitative composition of the duodenal microbiota was comparable to that of colon: lactobacillus spp. e+ ( e+ / e+ ) > . e+ ( e+ / e+ ), (p= . ); bifidobacterium spp. e+ ( e+ / e+ ) < . e+ ( e+ / e+ ), (p= . ); escherichia coli e+ ( e+ / e+ ) > e+ ( e+ / e+ ), (p= . ); bacteroides fragilis group e+ ( e+ / e+ ) > e+ ( e+ / e + ), (p= . ); faecalibacterium prausnitzii e+ ( e+ / e+ ) < . e+ ( e+ / e+ ), (p= . ), indicating the presence of sibo. in this case, the hydrogen breath test was completely uninformative: basal values - . ( . / . ) ppm, hydrogen concentration in minutes - . ( . / . ) ppm, in minutes - . ( . / . ) ppm, which is less than in healthy volunteers. conclusions: quantitative composition of the duodenal and colon microbiota is similar in the case of intestinal gvhd and/or virus-associated enteropathy in allo-hsct patients, which may be of diagnostic value for sibo confirmation. the hydrogen breath test is an uninformative method for sibo identification in patients after allo-hsct. disclosure: nothing to declare johannes schulte , patrick hundsdörfer , sebastian voigt background: adenovirus (adv) infections or reactivations frequently occur in the pediatric hematopoietic stem cell transplant (sct) setting and these infections contribute to increased morbidity and mortality. the nucleotide analog cidofovir might be effective in reducing adv load, however, nephrotoxicity is a considerable side effect. the new antiviral compound brincidofovir (bcv, cmx- ), a lipid-conjugate nucleotide analog with broad-spectrum antiviral activity in vitro, has been reported to be effective in cases where cidofovir treatment was unsuccessful. methods: data of eight pediatric patients undergoing sct for malignant and nonmalignant indications were analyzed. all patients were weekly monitored for adv viremia by pcr. in case two consecutive positive adv pcr results indicating a viral copy number > /ml were documented, patients received a weekly dose of cidofovir. if no reduction of adv load was seen within two weeks after the commencement of treatment or side effects demanded cidofovir discontinuation, bcv was obtained through an emergency expanded access programme. results: eight pediatric patients developed adv viremia with maximum viral loads ranging between and copies/ml. six patients had c type adv and two patients had non c type adv infections. five patients had viral co-infections: two had an additional cmv infection, one had an epstein-barr virus (ebv) and herpes simplex virus co-infection, one patient had an ebv co-infection and one patient had a bk virus co-infection. all eight patients initially received cidofovir, however, a substantial decrease in adv load could not be observed in any patient after a two-week administration course. except in one patient who had extensive intestinal graft-versus host disease (gvhd), adv infection was cleared in all patients within three weeks after the beginning of bcv treatment. in addition, all coinfections were cleared. no nephrotoxicity or other side effects were observed. conclusions: bcv was effective in all but one patient. oral bcv might not be effective in advanced upper gut gvhd, especially when applied via a gastric tube, yet this was observed in only one patient. eventually, without nephrotoxic side effects, bcv could be an useful alternative to cidofovir. disclosure: nothing to declare. background: the use of post-transplant high-dose cyclophosphamide (ptcy) has overcome the need for extensive depletion of t lymphocytes from haploidentical donor grafts, which traditionally resulted in severe and prolonged immunosuppression. however, reconstitution of cellular immunity may be delayed even after t cell replete haploidentical stem cell transplantation (haplo-sct) with ptcy. the study of the incidence and severity of viral reactivation is therefore relevant to the outcomes of haplo-sct with ptcy. methods: our study enrolled patients (women/men, / ), who underwent t cell replete haplo-sct from / to / and achieved hematopoietic engraftment. median age at transplant was . years (range, - ) . the underlying disease was aml (n= ), all (n= ), mds (n= ), myelofibrosis (n= ), cml (n= ), or cll (n= ). the conditioning regimen was myeloablative (n= ), reduced-intensity (n= ) or non-myeloablative (n= ). peripheral blood was the graft source in the majority of cases (n= ) and bone marrow in the remaining (n= ). recipient/donor cytomegalovirus (cmv) serostatus was -/-(n= ), -/+ (n= ), +/-(n= ), or +/+ (n= ). the combination of tacrolimus and mycophenolate mofetil was administered in addition to ptcy for prevention of graftversus-host disease. cmv, epstein-barr virus (ebv), and human herpesvirus- (hhv- ) reactivation was monitored by real-time quantitative pcr (rq-pcr) in blood twice weekly post haplo-sct. bk virus (bkv) reactivation was assessed by rq-pcr in urine and/or blood specimens in cases with symptoms suggestive of bkv-associated hemorrhagic cystitis (hc). results: with a median follow-up time of months (range, , the cumulative incidences (cin) of relapse and non-relapse mortality (nrm) were . % ( % ci, . - . %) and . % ( % ci, . - . %) at years, respectively. median disease-free (dfs) and overall survival (os) were . % ( % ci, . - . %) and . % ( % ci, . - . %) at years, respectively. the cin of cmv reactivation/infection (> copies/ml) reached . % ( % ci, . - . %) at months. cmv infection developed in out of patients who were at risk, whereas recurrent cmv reactivation was observed in patients with a median number of episodes (range, - ) per patient. the median total duration of antiviral therapy for cmv infection was days (range, - ) . cmv disease (pneumonia) was documented in patients. the cin of ebv reactivation (> , copies/ml) was . % ( % ci, . - . %) at months. no case of ebv-related post-transplant lymphoproliferative disorder was observed, however preemptive therapy with rituximab was required in patients with rapidly increasing ebv viral load. hhv- reactivation (> , copies/ml) was observed in patients (cin, . % at months; % ci, . - . %), with none of them requiring specific therapy. bkv-related hc occurred at a cin of . % ( % ci, . - . %) at months. cystoscopy for bladder hemostasis was required in / and nephrostomy in / patients with hc. conclusions: despite preservation of non-alloreactive memory t cells, haplo-sct with ptcy is associated with substantial rates of viral reactivation (especially cmv and bkv) resulting in the need for prolonged antiviral therapy and considerable morbidity as well. therefore, strategies to prevent viral reactivation and disease are still warranted in haploidentical stem cell transplantation. disclosure: nothing to declare. background: adenovirus(adv) infections are a wellrecognised cause of morbidity and mortality in children and adults receiving an allogenic stem cell transplant(hsct).the reported incidence of adv infection is higher( %- %) in paediatric hsct than in adults( - %),but we currently lack accurate data of adv infection burden among adults.cidofovir has been extensively used as a pre-emptive anti-adenoviral therapy and is current standard of care.we present our single centre experience of adv incidence and outcomes with pre-emptive approach in adult patients receiving t-cell depleted(tcd) hscts for myeloid disorders. methods: this is a single-centre retrospective analysis of consecutive hsct patients for myeloid disorders including aml, mds, mpn & aplastic anaemia between january -june using atg or alemtuzumab based tcd.adv screening was performed in all patients with standardised real time quantitative pcr on weekly basis during standard risk period. figure a - b] results: baseline characteristics (table ) of patients were similar across both cohorts with or without adv infection. overall . %(n- / ) patients were positive for adv dna on atleast one of the sanctuary sites(upper respiratory airway,blood,faeces,urine) and %(n- / ) of these experienced disseminated infection(defined by adv in ≥ sanctuary sites or rising adv dna copies in blood), while developed typical adenoviral disease (pulmonary). among patients with disseminated infection,majority had adv in gastro-intestinal( %), . % in genitourinary and % as both sanctuary site of infection,in addition to blood viraemia( % of all cases).cumulative incidence of adv infection was . %( %ci: . - . %) at months with median time of days(iqr: - days) to detect adv-dna post hsct.overall survival(os) at years for whole cohort was %( %ci: - ;median os- months) with no statistical difference between patients with disseminated adv infection vs those with none(log rank; p- . ; fig- a ). overall cumulative incidence of non-relapse mortality (nrm) was %( %ci: - %) and relapse(cir) was . %( %ci: - %) at years,but no statistical difference noted between patients with disseminated adv infection & those with none(nrm:p- . ;cir: p- . ;gray test).pre-emptive therapy with cidofovir ( mg/kg weekly iv infusion for weeks and fortnightly thereafter until infection free) was required in %( / ) of symptomatic adv infection patients and %( / ) with disseminated infections.one patient required brincidofovir therapy for refractory disease,but one patient died due to severe sepsis, before adv specific therapy could be given.remaining patients were monitored and all self-recovered on cessation of immunosuppression.all patients treated with cidofovir developed renal impairment(defined by atleast > % increase in baseline creatinine),however majority( %) recovered their renal function near their baseline (fig- b) . conclusions: adv infection remains a significant cause of morbidity in adult hsct patients, however pre-emptive management with cidofovir has improved os and nrm despite use of tcd conditioning.renal toxicity remains common with cidofovir but with use of intermediate doses, majority do recover their renal functions. clinical trial registry: n/a disclosure: nothing to declare background: publications on invasive fungal disease (ifd) in lymphoma patients are limited especially after allo-hsct. there are no data on outcome of allo-hsct in lymphoma patients with prior ifd. this study focuses on epidemiology of ifd before and after allo-hsct in children and adults with hodgkin's lymphoma (hl). methods: single center prospective study included patients with classical r/r hl who received allo-hsct from to . the median age was ( - ) y.o., children (< yo) - %. allo-hsct from mud was performed in , % (n= ), mrd - , % (n= ), mmud - , % (n= ), haplo - , % (n= ), with ric ( %) and predominantly ptcy-based gvhd prophylaxis ( %). primary antifungal prophylaxis was fluconazole in %, secondary -voriconazole ( %). eortc/msg criteria for diagnosis and bronchoscopy before allo-hsct in pts with ct-scan lung lesions were used. "active ifd" means ifd diagnosed just before hsct. median follow-up time was months . results: incidence of ifd before allo-hsct was , % (n= ). ifd prior to hsct were invasive aspergillosis (ia) with lungs involvement. antifungal therapy before allo-hsct was used in , % pts with median duration - months. complete response to antifungal therapy was in , % pts, partial response or stabilization - , %, and , % pts had an "active ifd". after allo-hsct all pts received voriconazole as an antifungal therapy or secondary prophylaxis. cumulative incidence of relapse or progression of ia after allo-hsct was , % with the median day after hsct, which were successfully treated with voriconazole in post hsct period. incidence of ifd after allo-hsct for naïve patients was , % (n= / ). etiology of ifd after allo-hsct was ia - %, invasive candidiasis (ic) - %, mucormycosis - % and % combined ifd caused by aspergillus fumigatus + rhizopus stolonifer. the median day of onset of ifd after allo-hsct was day+ and was associated with post-hsct relapse of hl (p= , ). the main site of infection were lungs ( %), the main clinical symptom -febrile fever ( %). antifungal therapy was used in all patients: voriconazole - %, micafungin - %, posaconazole - %, lipid amphotericin b - % and combination lipid amphotericin b with caspofungin - %. overall survival (os) at weeks from the diagnosis of ifd after allo-hsct was %. the -year os in children and adult with hl after allo-hsct was , %. development of ifd after allo-hsct do not decrease the -year os rate ( , % vs %, p= , ). the impact of prior ifd on -year os in allo-hsct recipients was not statistically significant in all group ( , % vs , %, p= , ) , and separately in children and adults. conclusions: incidence of ifd in children and adults with hodgkin's lymphoma before allo-hsct was , %. incidence of ifd after allo-hsct in patients with hodgkin's lymphoma was , %. the major etiology agents as before as after allo-hsct were aspergillus spp. ifd was a late complication after allo-hsct and associated with post-hsct relapse. despite the high incidence ifd before or after allo-hsct didn't influence the outcome in children and adults with hodgkin lymphoma. disclosure: nothing to declare our community has high cmv positive serostatus, which is a known risk for cmv infection or reactivation. we conducted a study to explore the incidence and outcome of cmv infection among post-hsct children. methods: medical records of pediatric patients (age ≤ years) undergoing single allogeneic hsct from january to december , at king faisal specialist hospital and research centre, riyadh, saudi arabia, were reviewed. all patients with active cmv infection or disease before and during transplant were excluded. a total of patients were included in the study; were female. median age at hsct was years. recipient cmv serostatus was positive in patients before hsct, and donors were cmvpositive. the recipient-donor (r/d) serology was . % r +/d+, . % r+/d-, . % r-/d+, and . % r-/d-. indication for hsct was immune disorders . %, hemoglobinopathies . %, bone marrow failure . %, malignant disorders . %, histiocytic . %, and metabolic disorders . %. source of stem cells was bone marrow in , cord blood in and peripheral blood stem cell in cases. donor was matched related among , unrelated matched/mismatched in , haploidentical , and related with -antigen mismatch in . total body irradiation (tbi) based conditioning was used for patients, while atg was used in patients. results: out of a total of patients, patients developed cmv infection post-hsct ( . %). incidence in female recipients was high ( . % versus . %, p-value . ). both recipient and donor cmv serology positive ( . %) developed cmv infection (p-value < . ). however, no cmv infection in both recipient and donor negative group (r-/d-). the incidence of cmv infection post-hsct was high in patients received tbi based conditioning ( out of , . %, p-value . ), and in haploidentical transplant with . % (p-value . ). source of stem cells, myeloablative versus nonmyeloablative conditioning, atg use in conditioning and agvhd, did not exhibit significant association with cmv infection. in multivariable setting, when adjusted for primary indication for transplant, donor hla type, tbi based conditioning and recipient and donor cmv serology at transplant, haploidentical donor (odds ratio: . , p-value: . ) and donor-recipient cmv sero-positivity (odds ratio: . , p-value: . ) were found to be significant risk factors. cmv infection resolution rate was . % ( ). with a median follow-up time of . ± . months from infusion, five-year overall survival of cmv infected group was lower ( . ± . ) as compared to non-cmv infected ( . ± . , p-value: . ). conclusions: incidence of cmv infection post-hsct in our center is comparable to other centers. our data suggest that donor-recipient cmv positive serostatus, haploidentical donor, and use of tbi based conditioning necessitate close attention and surveillance. background: toxoplasmosis is a rare and underestimated complication following allogeneic stem cell transplantation (allo-sct) with an often fatal course. this is in part due to limited diagnostics relying mainly on imaging and detection of parasite dna by pcr. we present here eleven cases of toxoplasma disease following allo-sct. methods: we retrospectively analyzed consecutive adult patients who received an allo-sct in our bone marrow transplant unit between july and july . eleven ( %) of these patients were diagnosed of toxoplasma disease. the main characteristics of the patients are shown in table . all patients, except two cord blood, have received peripheral blood stem cells. fludarabine-based conditioning regimes were used in all patients. only the two cord blood patients received thymoglobulin in the conditioning. graft-versus-host disease (gvhd) prophylaxis consisted on tacrolimus plus mycophenolate mofetil in ( %) patients and post-transplant cyclophosphamide followed by tacrolimus in ( %). before the allo-sct was performed the igg/igm toxoplasma serology of the recipient and donor. we reviewed the absolute lymphocyte count (alc) and cd + lymphocyte count within four weeks prior to the diagnosis of toxoplasmosis, and if the patients took effective primary prophylaxis for this parasite. toxoplasma disease was defined as the presence of toxoplasma infection plus clinical, radiological or pathological evidence. toxoplasma disease was considered the main cause of death when no other major life-threatening infection or other potential fatal complication occurred immediately before death. results: median (range) age (years) of the eleven patients diagnosed with toxoplasma disease was ( - ). for pancytopenia, no patient received trimethoprimsufmethoxazole (tmp-smz) but pentamidine for pneumocystis jirovecii-pneumonia (pcp) prophylaxis in cases and atovaquone in one. toxoplasma serology pretransplant was positive (igg+/igm-) in ten of the eleven patients. all donors were seronegative (igg-/igm-) except two. toxoplasmosis was diagnosed a median (range) of days ( - ) post allo-stc. the clinical presentations were as cerebral-encephalitis (n= ), chorioretinitis (n= ), pneumonitis (n= ) and disseminated toxoplasmosis (n= ). one case, patient and donor seronegative pre-transplant, was presented as a primary infection in form of chorioretinitis. all three patients with chorioretinitis were diagnosed after day + of allo-sct. at the time of toxoplasma disease, of ( %) of patients had an alc < cells/μl and all of them with immunosuppressive therapy and corticosteroids for acute or chronic gvhd. we had cd + lymphocyte count only in four patients and in three of them was < cells/μl. eight of the eleven ( %) patients died, with a median (range) of days ( - ) since diagnosis of toxoplasmosis, and in of them the toxoplasma disease was the main cause of death. conclusions: in our series, the incidence of toxoplasma disease after allo-sct is low and is related to high mortality, in accordance with what has been reported by other groups. positive pre-transplant serology and gvhd and its treatment were factors strongly related with toxoplasmosis. we encourage the use of tmp-smz instead of pentamidine for pcp-pneumonia prophylaxis in patients seropositive for toxoplasma gondii pre-transplant. clinical trial registry: data about its epidemiology in children are scarce. we retrospectively analyzed the incidence, the severity and the risk factors that contribute to the manifestation of this complication in a pediatric population. methods: during a -year period (january -june ) we performed in our center allogeneic transplantations, for malignant hematological diseases and for non-malignant. the majority of our patients received myeloablative conditioning regimens. diagnostic criteria of hemorrhagic cystitis were the detection of the virus with pcr in urine samples and/or in blood samples, in combination with hematuria and lower urinary tract symptoms (dysuria, urinary frequency, urgency, suprapubic pain) that couldn't be attributed to any other reason. we defined the hemorrhagic cystitis as severe when one of the following factors was present: formation of clots and continuous bladder irrigation, obstructive uropathy with creatinine elevation or need for urological intervention. results: a total of patients with median age , years ( , - ) were studied. children ( %, % ci, , - , ) manifested bk virus associated hemorrhagic cystitis with median age , years ( , [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] ) . onset of cystitis occurred at a median time of days (day +-day + ) after transplantation. in children cystitis was severe. the median duration of symptoms was days ( - ). the median time of hospitalization for children with severe cystitis was days ( - ) whereas for those who didn't manifest cystitis was ( - ) . in of the patients we examined the presence of the virus not only in urine but also in blood samples. in of them the test was positive and almost half of them ( ) manifested severe cystitis. the risk factors that were examined were age, administration of antithymocyte globulin, type of disease, graft source, type of donor and the presence of acute graft versus host disease (agvhd). in multivariable analysis, independent risk factors for the manifestation of hemorrhagic cystitis were age > years old (hr: , , % ci, , - , p< , ), transplantation for malignant disease (hr: , , % ci, , - , , p= , ) and the presence of agvhd (hr: , , % ci, , - , , p< , ). the overall survival of children with hemorrhagic cystitis was , % vs , % of those who didn't manifest this complication, but in multivariable analysis for survival cystitis wasn't a statistically significant risk factor. conclusions: according to our results, stem cell transplantation in children > years old who suffer from a malignant disease and the presence of agvhd consist independent risk factors for the manifestation of bk virus associated hemorrhagic cystitis. the identification of the risk factors of this serious complication will contribute to better management of transplanted patients. further research through prospective trials can contribute to the better understanding of the pathophysiology of hemorrhagic cystitis and to the establishment of appropriate diagnostic and therapeutic guidelines. disclosure: nothing to declare p impact of natural killer cell reconstitution on outcomes in patients with early cytomegalovirus reactivation after allogeneic hematopoietic stem cell transplantation background: cytomegalovirus (cmv) reactivation influences survival after allogeneic hematopoietic stem cell transplantation (sct) and induces natural killer (nk) cell expansion. we evaluated nk cell reconstitution and clinical outcomes following early cmv reactivation after sct. methods: lymphocyte subsets were measured by flow cytometry on day in patients with hematologic malignancies undergoing sct between january and december at kanagawa cancer center, excluding patients with graft failure or death within days. cmv reactivation was defined as initiation of preemptive cmv therapy following pp antigenemia surveillance. results: the subjects were males and females with a median age of years (range: - years). the median follow-up period for survivors was . years (range: . - . years). there were patients with acute myeloid leukemia, with acute lymphoblastic leukemia, with myelodysplastic syndromes, and with other diseases. at transplantation, patients were standard risk and were high risk. myeloablative conditioning and reduced-intensity conditioning were employed in and patients, respectively. bone marrow transplantation, peripheral blood stem cell transplantation, and cord blood transplantation was performed in , , and patients, respectively. cmv reactivation occurred in patients ( %) at a median of days (range: - days) after sct. grade ii-iv acute gvhd and chronic gvhd affected patients ( %) and patients ( %), respectively. among all patients, -year overall survival (os), cumulative nonrelapse mortality (nrm), and cumulative relapse (cir) rates were %, %, and %, respectively. in patients without cmv reactivation (cmvr-) versus patients with cmv reactivation (cmvr+), -year os, nrm, and cir were % vs. % (p < . ), % vs. % (p = . ), and % vs. % (p = . ), respectively. among all patients, the median level of cd -cd + cells, cd +cd cells, and cd +cd + cells on day was /μl (range: - /μl), /μl ( - /μl), and /μl ( - /μl), respectively. nk cell subsets showed no significant differences between cmvr-and cmvr+ patients. when patients were divided into low and high groups at the median level of each nk cell subset, cmvr+ patients with high cd -cd +, cd +cd -, or cd +cd + cells showed significantly better -year os than those with low cells ( % vs. %, p < . ; % vs. %, p < . ; % vs. %, p = . , respectively). high cd -cd + cells were significantly associated with lower nrm ( % vs. %, p = . ), while high cd +cd -cells were significantly associated with lower cir ( % vs. %, p = . ). multivariate analysis confirmed these nk cell subsets as prognostic factors in cmvr+ patients. conclusions: nk cell reconstitution may contribute to improved transplantation outcomes in subgroups of cmvr + patients. disclosure: nothing to declare background: rezafungin (rzf) is a novel echinocandin in development for prevention of invasive fungal infections caused by candida, aspergillus, and pneumocystis spp. in patients at high risk of infection. rzf has demonstrated in vivo prophylaxis efficacy and low risk of drug-drug interactions. furthermore, the stability and pk profile of rzf allow for once-weekly dosing. rzf is also in development for treatment of candidemia and invasive candidiasis using a dosing regimen of rzf mg followed by mg once-weekly, which achieved > % target attainment against candida. while lower doses might be useful to prevent candida and pneumocystis, invasive aspergillosis is a different challenge. we evaluated rzf dosing for prophylaxis against aspergillus fumigatus in blood and marrow transplant (bmt) patients using pk/pd simulations of the treatment dosing regimen. methods: a previous population pk model was refined using data from phase and phase trials of iv rzf (nonmem vers . ). stepwise forward selection (α = . ) and backward elimination (α = . ) were used to assess for relationships between interindividual pk variability and covariates, such as age, sex, bsa, albumin, liver and renal function markers, and infection status. the final model was validated by comparing model-based predictions to observed data. the model and demographic data from bmt recipients at stanford medical center were used for monte carlo simulation (n= , ) of expected rzf concentrationtime profiles in bmt patients receiving iv rzf mg on week followed by mg weekly x . of the patients included in the demographic dataset, were female (mean values at baseline: age, years [ - years]; weight, . kg [ - kg] ). the median (range) bsa in the demographic dataset was . m ( . - . ), and albumin was . g/dl ( . - . g/dl) . free-drug concentration-time profiles were evaluated ( . % human protein-binding) relative to the a. fumigatus minimal effective concentration required to inhibit % of isolates tested (mec ; jmi - sentry international surveillance data). results: the population pk model was a linear, compartment model with zero order iv input. albumin, sex, infection status, and body surface area were statistically significant predictors of interindividual variability; clinical significance of these factors was not determined. the model provided precise, unbiased fits to the observed data (r = . observed vs individual-predicted concentrations). rzf plasma free-drug concentrations at weeks , , and were above the a. fumigatus mec ( . mg/l) for the entire dosing interval in . %, . %, and . % of simulated patients, respectively, and in ≥ . % for all weeks based on the mec ( . mg/l). conclusions: these data modelled from bmt patients support the rzf dosing regimen of mg iv followed by mg once-weekly for prophylaxis against a. fumigatus. current antifungal prophylaxis may be limited by toxicity, ddis, or patient factors such as mucositis. the pk of rzf and its spectrum, safety, tolerability, and lack of ddis may address current unmet needs in ifi prophylaxis for bmt and other immunocompromised patients. disclosure: janice brown: research funding, cidara therapeutics, elizabeth lakota: research funding, cidara therapeutics, shawn flanagan: employment, cidara therapeutics, taylor sandison: employment, cidara therapeutics, voon ong: employment, cidara therapeutics, christopher rubino: research funding, cidara therapeutics p is fungal prophylaxis necessary in non myeloablative peripheral blood stem cell allogeneic transplantation in the pre-engrafment period? julien vaidie , jean-baptiste woillard , stéphane girault , marie-laure dardé , arnaud jaccard , daniel ajzenberg , bernard bouteille , pascal turlure background: non myeloablative peripheral blood stem cell transplantation (pbsc), by limiting toxicity, can be proposed to elderly patients or patients with comorbidities. however, fungal infections remain a key issue that can negatively impact outcome, and increase duration and cost of hospitalization. systematic fungal prophylaxis have demonstrated benefits in outcome in the context of myeloablative conditioning but are not currently in reduced intensity conditioning allograft with pbsc. fluconazole prophylaxis is currently recommended in this situation (ecil). methods: primary objective of this retrospective study was to evaluate fungal infection incidence after allograft procedure in patients who received a non myeloablative allograft with pbsc in limoges university hospital between june and june . patients received fludarabine mg/m /day between d- and d- before allograft and busulfan . mg/kg/day at d- and d- . gvh prophylaxis consisted in rabbit anti-lymphocyte serum at the dose of . mg/kg at d- and d- , and ciclosporin at the beginning dose of mg/kg per os twice a day. mycophenolate mofetil was adding for patients with hla-matched or mismatched unrelated donors. patients did not systematically receive antifungal prophylaxis during the neutropenic pre-engraftment period. when patients had fever during more than hours, an empirical fungal treatment (caspofungine) was added to empirical antibiotics. as soon as neutropenic recovered and in the case of apyrexia without microbiologic documentation, antimicrobial treatments were stopped while in the case of microbiologic documentation, treatments were adjusted to germ in term of dosing and time of administration following recommendations. however, some patients received antifungal azole prophylaxis during the neutropenic pre-engraftment period in case of history of previous invasive aspergillosis (ia), or a nasal colonization by aspergillus. in post-engraftment period, posaconasole prophylaxis was administered for patients with systemic corticotherapy for acute graft-versus-host disease. results: patients were evaluated (median [min-max] age of [ - ] years). % of patients received an hlaidentical related donor, % an hla-matched related donor and % an hla-mismatched unrelated donor. the five years overall survival and survival without relapse or gvhd were % ic [ %- %] and % respectively ic [ %- %]. the median time for neutrophil recovery was days. patients did not receive prophylaxis and only patients received systematic fungal azole prophylaxis in the pre-engraftment period. two patients received an empirical treatment by caspofungine. only ifi was documented during the neutropenic period : candida krusei in blood culture. in the post engraftment period, patients with acute gvhd treated by corticotherapy received an antifungal prophylaxis by posaconazole and only patient had a probable ia at day despite prophylaxis by posaconasole. conclusions: except for patients with previous history of ifi, our results provide additional arguments against systematic fungal prophylaxis after reduced intensity conditioning with pbsc allogenic transplantation in the pre-engraftment period with a very low incidence of invasive fungal infections. in post-engrafment period, posaconazole prophylaxis is required for patient with gvhd treated by corticotherapy. disclosure methods: a simple, rapid and sensitive method using hplc with a diode-array detector (dad) was developed and validated for the quantification of letermovir in human serum using sorafenib as internal standard. after pretreating serum samples by liquid-liquid extraction with tert-butyl methyl ether, separation was achieved on a x-terra rp- column (dimension x . mm, μm) at c using gradient elution with a mobile phase of mm ammonium bicarbonate ph . (mobile phase solvent-a) and acetonitrile: mm ammonium bicarbonate ph . (mobile phase solvent-b). samples were eluted at a flow rate of . ml / min throughout the -minute run. uv wavelength mode was used, detection was at nm. results: the calibration curve was linear (r > . ) in a concentration range of - ng / ml for letermovir. the hplc assay established for letermovir determination showed a high rate of accuracy and precision with an intraday variability of - . to % (accuracy) and . to . % (precision) and an interday variability of - . to . % (accuracy) and . to . % (precision), respectively. letermovir serum concentrations of patients ( male / female, mean age . years) were determined in daily clinical practice. the mean concentration was ng / ml (median ng / ml, standard deviation ng / ml, range - ng / ml). conclusions: the newly developed hplc method is useful for the determination of letermovir concentrations. patient samples analyzed in a routine clinical setting demonstrated considerable interindividual variability. all measured concentrations were above the ec of letermovir. monitoring the concentration of letermovir could help to prevent over-or underexposure, especially in patients with polypharmacy which is frequent in allogeneic hematopoietic stem cell transplant recipients. disclosure background: the use of preemptive strategy (pet) has lowered the incidence of cmv disease in allo-sct to - %. nonetheless the use of this strategy implies that more than % of seropositive patients will replicate cmv. several studies have shown that cmv replication is detrimental for patient survival although the viral load related to this bad outcome variates among studies. objective: to analyse thel impact of cmv replication in overall survival (os) in allo-hct patients. methods: to analyse the impact of cmv replication in os we perform a unicentric, retrospective study on consecutive first allo-hsct patients transplanted between jan- and oct- with a median follow-up of days ( - ). all patients were monitored post-hct with real time pcr cobas-taqman® /cobas ® (rtpcr) in plasma. the cut-off for inception of pet was iu/ml. cmv mutations (ul /ul gene), were studied in plasma samples by sanger sequencing, median cmv viral load iu/ml ( - ). results: patients ( ): women/men ( %/ %), median age was years (range - ). identical allogeneic scts ( %), haploidentical scts ( %). donors were related in cases ( %) and ( %) unrelated. progenitors source was % peripheral blood and % bone marrow. cmv status was (d+/r+) in %(n= ), (d+/r-) in %(n= ), (d-/r+) in % (n= ) and (d-/r-) in % (n= ), unknown cases. positive pcrs were detected in patients: one episode in ( %); episodes in ( %) and to episodes in patients ( %). fifty-five patients ( %) received preemptive therapy. fourteen episodes ( %) were refractory/ probable refractory cmv infections (according to the criteria of chemaly r. cid ). a resistant mutation (ul gene) was detected in one patient with refractory infection patients that developed cmv infection had an inferior non-significant os at years ( , % vs , % log-rank p , ). those patients that received pet for cmv had a significant inferior os compared with those that replicate cmv but didn't receive preemptive therapy ( , % vs %, log rank p= . ). os of patients that received pet was inferior compared with those without pet (with or without infection) ( , % vs , %, logrank p , ). no difference in survival was found for those patients treated pre-emptively that were refractory vs no refractory ( % vs , %, log-rank p , ). conclusions: patients that received preemptive therapy had a significant inferior overall survival compared with those that didn´t replicate and those that replicate cmv but didn't receive preemptive therapy. this reinforce the relevance of prophylactic strategies for cmv with drugs with good safety profile like letermovir that in a randomised trial proved to decrease the need for preemptive therapy. disclosure: rafael, de la camara: has received grants from astellas, gilead, janssen, merck, novartis and pfizer clinical evaluation of stenotrophomonas maltophilia infection in allogeneic hematopoietic stem cell transplant recipients -retrospective single-center data analysis negative bacillus that causes severe infections associated with high morbidity and mortality in immunocompromised patients. the aim of our study was to determine incidence, characteristics and outcome of s. maltophilia infection in patients (pts) who underwent allogeneic hematopoietic stem cell transplantations (allo-hsct) in institute of hematology and transfusion medicine between october and november . methods: we retrospectively evaluated incidence, clinical features and outcome of s. maltophilia infections in consecutive patients with median age- years (range - ), who underwent allo-hsct from unrelated donors - ( . %), matched sibling donors - ( . %) and haploidentical donors - ( . %) in our center. s. maltophilia was detected by culture-based microbiological tests. invasive infection was defined by isolation s. maltophilia from cultures in the presence of both clinical symptoms and signs of infection -blood stream infection (bsi), pneumonia with or without pulmonary haemorrhage. the only colonization status was defined as s. maltophilia culture-positive samples in the absence of infection symptoms. in vitro susceptibility tests to antibiotics were performed. results: pts ( . %) with median age- years (range - ) with s.maltophilia culture positive samples were identified. ( . %) underwent allo-hsct from unrelated donors, -from matched sibling donor and -from haploidentical donor. among them bsi developed in pts ( . %), pneumonia in pts ( %) -with fulminant and fatal pulmonary hemorrhage in pts ( . %). all patients with pneumonia demonstrated bsi. positive sputum cultures were detected in pts, in pts hemoptysis was observed. the rest of isolated strains were identified as colonization (throat -in pts, stool -in pts). all patients with invasive s. maltophilia infection before pathogen identification demonstrated persistent fever despite of the use of broadspectrum antibiotics (carbapenems, glycopeptides, aminoglycosides, colistin), prophylactic antifungals and antivirals. all of them received fluoroquinolone (ciprofloxacin) as a standard antibacterial prophylaxis before neutropenic fever occurred. all patients ( %) with bsi, pneumonia and pulmonary hemorrhage died before engraftment (anc - . g/l) - of them during - hours from the onset of a positive blood culture for s. maltophilia. the c-reactive protein (crp) concentration before identification of s. maltophilia invasive infection was > x- x upper normal limits (unl). susceptibility to antibiotics of isolated strains from blood and sputum was respectively: % and % for ceftazidime, % and % for trimethoprim-sulfamethoxazole, % and % for levofloxacin; while % and % strains were resistant to ciprofloxacin. -year overal survival (os) and -y os for this group was . % and . % respectively compared with . % and . % for group without s. maltophilia infection. conclusions: s. maltophilia invasive infections are associated with high morbidity and mortality in allo-hsct recipients especially in the period from conditioning therapy to engraftment. an exposure to broad-spectrum antibiotics in the treatment of neutropenic fever or confirmed bacteremia of other etiology is one of risk factors of breakthrough s. maltophilia infections. empiric therapy against s. maltophilia in selected patients in risk of such infection before pathogen identification may be lifesaving procedure. disclosure: nothing to declare. role of cmv reactivation following allogeneic stem cell transplantation in preventing relapses in patients with acute myeloid leukemia background: cytomegalovirus(cmv) reactivation is common in patients undergoing allogeneic stem cell transplantation. it has been shown recently that cmv reactivation is associated with reduced risks of relapse in patients undergoing allogeneic stem cell transplantation for aml. however the analysis of cibmtr data did not show any effect of cmv reactivation on relapse. with this background we conducted an analysis of patients suffering from aml who are undergoing allo-sct for their long term disease free survival with respect to cmv reactivation. methods: after obtaining permission from hospital medical records committee, we retrospectively analysed data from electronic medical records of patients undergoing allo-sct for aml at our center between january to august . patients who underwent matched sibling, matched unrelated and partially matched allo sct were included. all patients underwent cmv monitoring with weekly pcr starting from the time of engraftment till d+ following allo sct. value of ≥ copies/mcl was considered as cut off for initiation of treatment in matched sibling donor transplant but in unrelated donor or partially matched donor transplants, ≥ copies/mcl was used as cut off for initiation of pre emptive therapy. results: total of patients were included in study. median age was . ± . years ( - yrs). ( . %), ( . %) and ( . %) patients underwent matched sibling, haplo (partially matched) and mud transplantation respectively. median follow up was months( - months). (table ) acute gvhd (grade - ) was observed in ( . %) of patients. cmv reactivation occurred in ( . %) of patients. overall survival at last follow up was . % ( / patients). ( . %) patients relapsed during follow up. relapse free survival at at last follow up was . %. ( . %) of patients who had cmv reactivation didń t relapse, whereas ( %) of patients who didn´t have cmv reactivation relapsed which was statistically strongly significant p < . . (figure ) similar results were seen in recently published paper from japanese society for hematopoietic cell transplantation (jshct) transplantation-related complication working group. conclusions: . cmv reactivation following allo sct had beneficial effect on preventing relapse in patients with aml. . probable immune activation resulting due to cmv reactivation may result in better graft versus leukemia effect preventing subsequent relapses. [ background: human herpesvirus (hhv- ) causes lifethreating central nervous system disorders such as encephalitis after allogeneic hematopoietic stem cell transplantation (hsct). recent studies showed that cd , a member of the tumor necrosis factor receptor superfamily, has been implicated as a specific receptor of hhv- b, and that its expression levels in cd -positive t cells after hsct could be related to the reactivation of hhv- . real-time quantitative polymerase chain reaction analysis (qpcr) is the most commonly used method for detecting and evaluating hhv- reactivation after hsct, but more sensitive detection method is required. we recently developed a new monitoring method for hhv- reactivation using digital pcr (dpcr) which provides high sensitivity of detecting hhv- dna in clinical samples. in this prospective study, we evaluated the relationship between hhv- reactivation monitored by dpcr and expression of cd on cd + t cells before and after allogeneic hsct. methods: thirty-four patients who underwent allogeneic hsct for hematological diseases at keio university hospital (tokyo, japan) between january and march were consecutively enrolled into this study. peripheral blood samples of the patients were obtained before the conditioning (pre), the day of transplant (day ), and weekly during the first month after transplantation (days , , , and ) . hhv- viral load in plasma was quantitatively measured by dpcr. the primers and a probe of dpcr for hhv- b were selected from immediate-early (ie- ) protein transactivator region (u ). we evaluated the relationship between hhv- reactivation and the serial expression rates of cd in cd + t cells (cd /cd ratio) measured by flow cytometry before and after hsct. results: median age of the patients was . years. onethird of patients received cord blood as a stem cell source. hhv- reactivation was detected in patients ( %) with dpcr. a comparison of cd /cd ratio between the patients with and without hhv- reactivation after hsct revealed that cd /cd ratio was significantly higher in patients with hhv- reactivation than those without before conditioning ( in contrast, there was no such significant difference after transplant (days to ) . in multivariate analysis, higher cd /cd ratio before conditioning (odds ratio (or) = . , % confidence interval (ci): . - . , p = . ) and stem cell source from human leukocyte antigen mismatched donor (including all cord blood transplantation cases) (or = . , %ci: . - . , p = . ) remained to be significantly associated with the incidence of hhv- reactivation. conclusions: higher cd expression rate in cd + t cells before hsct was associated with higher risk of hhv- reactivation, which could be a promising marker for predicting hhv- reactivation after allogeneic hsct. careful observation and monitoring may be needed in cd highly expressed patients. it is a subject of further research to clarify the role of cd + cd + t cell in hhv- reactivation. disclosure: nothing to declare. methods: criteria for the administration of ici (vistide) were grade iii-iv (clinically significant hematuria with clots) bk-related hemorrhagic cystitis after allo-hct which showed no improvement after symptomatic therapy with hyperhydration and bladder irrigation. cidofovir was diluted in ml of normal saline and installed via a foley catheter which was blocked for hour. not knowing the level of absorption of the drug we decided to give probenecid prophylaxis in all patients. ici was repeated weekly according to severity of symptoms. urine and plasma bkv viral loads were quantified by rq-pcr results: six patients (median years, - ) received ici after allo-hct. patients had haematological malignancies (aml , all , mds ), received busilfex-based myeloablative conditioning and a graft (pbsc , bm ) from a / hla-matched ( pts), / ( pt) or haploidentical ( pt) donor. median time for the onset of bkv-hc after allo-hct were . days (range - ). all patients were under standard cyclosporine prophylaxis and none of the patients had any signs of acute gvhd at the time of onset of hc. the median pcr-bkv viral load at the onset of bkv-hc in urine and plasma were . x (range . x - x ) and . (range - ), respectively. the median maximum pcr-bkv viral load in urine and plasma were . x (range . x - x ) and . (range - . ), respectively. five patients had impaired renal function (median egfr ml/min, range - ) at first ici which was probably multifactorial. the median dose of intravesical cidofovir was mg/kg (range . - mg/kg) and a median number of . instillations (range - ) were given. in / cases symptoms of cystitis improved dramatically and hematuria resolved. virological response (at least log reduction) was observed in all cases. two patients experienced relapse of hemorrhagic cystitis and were retreated with ici which resulted in resolution of the symptoms and the hematuria. no deterioration of renal function of other systemic adverse effects were observed. after a median follow up of . days after transplantation (range - ), / patients are alive without cystitis symptomatology and died ( due to relapse and due to trm). conclusions: in this retrospective study we propose that local therapy of bkv-hc with ici is safe and has high clinical and virological response rates. the administration of ici after allo-hct should be controlled in prospective randomized trials. disclosure: nothing to declare background: since cmv-preemptive therapy approach was implemented, cmv disease frequency is very low. however, cmv reactivation and the need of using nephrotoxic plus/less myelotoxic drugs is very frequent. in addition to the toxicity of the medications to avoid cmv disease, other potential adverse effects of cmv have been mentioned in medical literature. in this study, we wanted to estimate how recipient/donor serologic status influences the outcome of allo-hsct in our most recent series of patients. methods: the population analyzed for this report is the all patients who underwent allo-hsct during the -year period from october september in our unit. median age at transplant was years (range: - ). one hundred and thirty were male ( %) and were female ( %). baseline diseases were: aml, lpd, all, mds, mpd, mm, and bmf. donor was unrelated in transplants ( , %) and was family in ( , %) (including haplo-identical). conditioning regimen was ric in procedures ( %) and intensive in ( %). stem cell source was pb in ( , %) and bm in cases ( , %). median follow-up was months (range: - ). patient's and donor's cmv igg were positive in ( , %) and ( , %), respectively. recipient/donor serology was +/-(risk group ) in ( , %), +/+ (risk group ) in ( , %) , -/+ (risk group ) in ( , %) y -/-(risk group ) in ( , %). results: two pts underwent a second transplant before day + due to graft failure. overall mortalities (om) at days + and + of the rest of the series ( pts) are shown in table. the highest risk group (recipient cmv + / donor cmv -) exhibited more than double om at day + and more than four times om at day + , when compared with pts at lowest risk (recipient cmv -). those striking differences were mainly due to nrm. om for risk group ii (recipient cmv + / donor cmv +) was intermediate. conclusions: in our studied population, mainly adult patients, the combination of cmv-seropositive patient with a cmv-seronegative donor had a very clear adverse impact on hsct outcome. as a result, we considered that the election of a cmv-positive donor for a cmv-positive patient continues to be strongly advisable, whenever is possible. on the other hand, once letermovir has proved to be efficient and well-tolerated and has been licensed for prophylaxis of cmv in high risk recipients, this approach appears to be very attractive to try to avoid the adverse impact of recipient cmv-seropositivity, particularly when finally chosen donor is cmv negative. disclosure: nothing to declare an active surveillance and an early and individualized management is critical to avoid mortality from respiratory viral infections in allo-hsct recipients background: respiratory viral infections (rvis) are frequent among the general population. in transplant recipients, rvis are known to cause an important morbidity and potential mortality. for this reason and several others, as the need of preventing other pts from contagious or avoiding misdiagnosis with other infections processes, a high index of suspicion of vris is necessary. during the last few years, we have implemented an active and systematic surveillance policy orientedto early detection and management of rvis in the hsct recipients. methods: the population analyzed for this report is the patients who underwent allo-hsct from january through march in our unit. median age at transplant was years (range: - ). one hundred and four were male ( . %) and were female ( , %). baseline diseases were: aml, lpd, all, mds, mpd, mm, and bmf. donor was unrelated in transplants ( . %) and was family in ( . %) (including haplo-identical). conditioning regimen was reduced in procedures ( %) and intensive in ( %).stem cell source was pb in ( . %) and bm in pts ( . %).median follow-up was months (range: - ); at the close of the analysis, majority of the series ( . %) had a follow-up superior to one year from hsct. a throat swab(ts) was taken from every patient with any, even minor, respiratory symptoms. the respiratorysamples were tested whith a complete pcr panel of human respiratory viruses: rhinovirus (rv), influenza a and b virus (iv-a, iv-b), parainfluenza virus (pivs - ), respiratory syncytial virus (rsv), metapneumovirus (mpv), coronavirus (cov), adenovirus (adv), and bocavirus (bov). results: day + overall mortality of the series was , %. day + overall mortality was , % ( , % nonrelapse mortality -nrm-, and , % progression/relapse mortality). causes of nrm reflected in table . no patients died due to rvis. from st july through th june (a -month period), ts samples were obtained from pts ( , %).the median number of samples/patient was (range: - ).a total of ( - ) rvis episodes were diagnosed in pts ( , %).the median presentation of the first rvi was at the day + ( - ) post-hsct. the viral distribution was: rv ( . %), iv ( . %), piv ( . %), rsv ( . %), mpv ( . %), cov ( . %), adv ( . %), and bov ( . %).there were mixed (two or more viruses) rvi episodes. the temporary distribution of vri episodes is shown in figure . conclusions: ) symptomatic infections due to respiratory viruses are very frequent among the allo-hsct recipients. ) a high level of suspicion, as well as an early and systematic screening and management policy, are critical to avoid potential attributable mortality and the nosocomial spread of rvis among the transplant recipients. ) in our series, rhinovirus, parainfluenza and adenovirus might be detected at any moment of the year; the rest of the viruses showed a clear seasonal pattern (november to april). [[p image] . background: trimethoprim-sulfamethoxazole (tmp-smx) is the most suitable drug for prophylaxis against pneumocystis pneumonia and infections with toxoplasma after allogeneic haematopoietic stem cell transplantation (allo-hsct). allergic reactions or hypersensitivities, mainly exanthemas, occur in about - % of the patients, usually resulting in the use of alternative prophylactic drugs (e.g. pentamidine or atovaquone). it has been hypothesised that allergies might be cured with allo-hsct. methods: we conducted a retrospective chart review of patients with tmp-smx re-exposition after allo-hsct from december to september . follow-up is current as of december . results: six patients (f/m: / , median age: years, range: - years) with a history of tmp-smx hypersensitivity prior to allo-hsct were re-exposed to tmp-smx after engraftment of a matched related (mrd, n= ) or matched unrelated (mud, n= ) donor. median time to re-exposition was . (range: - ) days after allo-hsct with one oral dose of tmp-smx. in four patients, tmp-smx was tolerated without any signs of hypersensitivity reactions and has been continued for a median of days (range - ) until last followup. one patient (mud, re-exposition at d+ ) experienced pruritus and erythema some hours after tablet intake. another patient (mud, re-exposition at d+ ) developed an exanthema one day after re-exposition which was later diagnosed as a cutaneous gvhd. conclusions: re-exposition of tmp-smx in patients with prior hypersensitivity is feasible after allo-hsct. after successful re-exposition, patients can be treated with the best-studied drug for prophylaxis of infections with pneumocystis and toxoplasma. disclosure: nothing to declare brincidofovir for adenoviremia in paediatric hsct for primary immune deficiency background: reactivation of adenovirus is a severe complication of hsct associated with significant morbidity and mortality, particularly for children with primary immune deficiency (pid). the only drug currently licensed to treat adenovirus infection is cidofovir. brincidofovir is a lipidlinked derivative of cidofovir which has been shown to be a safe and effective alternative treatment to cidofovir. there is limited data describing the use of brincidofovir in patients undergoing hsct for primary immune deficiency. we reviewed all patients who received brincidofovir after undergoing hsct for primary immune deficiencies between and at the great north children's hospital, newcastle upon tyne, uk. results: of patients transplanted for pid, developed significant adenoviraemia ( %). all were treated with cidofovir initially but were switched to brincidofovir because of a failure to respond or because of renal toxicity. of these, resolved their adenoviraemia within days of commencing treatment (figure ). donor sources were tcr alpha/beta/cd depleted haplo-identical (n= ), tcr alpha/beta/cd depleted mmud (n= ) and / mud (n= ). patients were conditioned with treosulphan/fludarabine/thiotepa/atg/ rituximab (n= ), treosulphan/fludarabine/atg/rituximab (n= ) or treosulphan/fludarabine/alemtuzumab/ gcsf/plerixafor (n= ). occurrence of agvhd and treatment of agvhd are outlined in table . patient died + days post-transplant of multi-organ failure, severe thrombotic microangiopathy and sepsis. although patient initially responded to brincidofovir, reactivation occurred after cessation of treatment; severe diarrhoea precluded the reintroduction of brincidofovir and the adenoviraemia persisted with poor immune reconstitution. treatment with addback t cells was attempted however the patient died days post-transplant after a cerebral haemorrhage. patient had long-standing chronic diarrhoea which was thought not severe enough to warrant cessation of brincidofovir treatment. conclusions: the complete resolution of adenoviraemia in / patients who had previously failed to respond to prior therapy with cidofovir suggests that brincidofovir may be an effective treatment option for adenoviral reactivation post-hsct for pid. however, resolution of adenoviraemia is influenced by many other factors, including the adequacy of immune reconstitution, the degree of induced immune suppression and the presence of comorbidities such as gvhd. due to the small sample size it was difficult to assess the relative importance of these factors in this cohort. brincidofovir was well tolerated however its effectiveness may have been limited by poor gastrointestinal function in one patient (patient ) and could not be used after a viral reactivation in another for the same reason. further studies of the use of brincidofovir in this specific cohort are needed to clarify the role and effectiveness of this treatment. background: there is a high prevalence of cmv seropositivity in algerian population. because of high morbidity and mortality in pts who underwent allo sct with cmv reactivation, effective surveillance and timely treatment using anti-viral therapy s required. the risk of cmv reactivation depends on the type of stem cell source, immunosuppression (is) and serological status of the donor/ recipient pair. methods: over a months period (from / / to / / ), pts underwent allo-hsct for malignant or non-malignant hematology diseases of which pts are evaluated for this study. cmv reactivation was observed in pts ( . %) (aml: pts, all: pts, cml: pts, multiple myeloma: pt, nhl skin: pt, primary myelofibrosis: pt, aplastic anemia: pts, fanconi anemia: pts, β-thalassemia: pt), with a median age of years ( - ), sex ratio (m/f) of . . allo-hsct done with sibling donors: pts, haplo-identical donors: pts and pheno-identical donor: pt. all pts were treated by chemotherapy alone with myéloablative conditioning (mac) in pts and reduced intensity (ric) in pts. all pts received peripheral blood stem cells with an average rate of cd + cells: , . /kg ( . - . ). additional bone marrow graft was used in pts that received a haploidentical graft without pt-cy. gvh prophylaxis associated cyclosporine (csa) and methotrexate (sibling and phenoidentical); csa-mtx-mmf or csa-mtx-cyclophosphamid (haplo-identical). before transplantation, donor/recipient pair is at high risk reactivation in pts ( . %). detection of cmv reactivation done by antigenaemia pp or by quantitative pcr weekly for the first months and during an is treatment for acute or chronic gvhd. pre-emptive therapy is initiated by ganciclovir as soon as positivity of antigenaemia or increased viral load in pcr. results: a first reactivation occurred on average day ( - ) in pts ( . %) of which pts under corticotherapy for acute gvhd ( pts), thrombotic micro-angiopathy ( pt) and renal failure ( pts) or due to a reinforced is for haplo-identical transplantation ( pts). one pt with chronic gvhd presented a late reactivation months after transplant. twenty pts presented a nd reactivation on average day ( - ) and pts a rd reactivation on average day ( - ). pre-emptive treatment is introduced in the first episode by a viral dna polymerase inhibitor (ganciclovir: pts; valganciclovir: pts, foscarnet: pt). the negativity of antigenaemia is observed on average at days of treatment ( ) ( ) ( ) ( ) ( ) . second line treatment was required in pts ( %) due to resistance ( pts), severe cytopenia ( pt) or renal failure ( pt). the onset of severe cytopenia imposed a dose reduction ( pts) or a therapeutic stop ( pts) before days. two pts received additional maintenance treatment for negativation delay. three pts ( . %) died from cmv infections resistant to antiviral treatment (pneumonia: , colitis: ). conclusions: cmv infection is a serious complication after allo-hsct. in the absence of vaccination, the systematic monitoring for cmv reactivation is strongly recommended for the establishment of a rapid and effective preemptive treatment. disclosure: nothing to declare p abstract withdrawn. results: in transplanted group, episodes of bkv reactivation occurred in patients ( %). in cases only urine colonization (c) found before hsct. in this group in patients ( %) virus was transmitted from urine to the blood (b) . dysuria and/or hc were observed in / ( %) patients . all of them ( %) had urine and serum involvement. in cases bkv replication was found after hsct ( -cases detected in urine, cases-bothserum and urine). dysuric syndromes and/or hc were found in / of cases ( %)-all in patients with serum and urine involvement. urinary tract was always first location of the virus. there was no case of isolated serum reactivation. the incidence of bk infection was higher in patients older than > yrs (p< . ), transplanted from family donor (msd) (p< . ). mud recipients had more often both serum and urine reactivation (p< . ) than isolated urine involvement. sex, day of neutrophil recovery, conditioning regimen, or use of total body irradiation were not significant risk factors for bkv infection, or hc . six patients were treated with cidofovir (range - doses) with good response. there was no death due to evident bkv infection. conclusions: bkv reactivation remains one of the most frequent infectious complication in children undergoing allogeneic hsct. most of patients experienced mild infection and age < years was the positive prognostic factor influencing its incidence. bkv monitoring and prompt treatment of hc resulted in excellent outcome. we observed surprisingly high rate of new bkv replication after hsct. disclosure: nothing to declare background: high-dose chemotherapy (hd-ct) and auto pbsct have been the standard therapy for multiple myeloma (mm) for more than two decades, despite a wide range of new therapeutic options. recurrent/refractory malignant lymphomas and recurrent/metastatic germ cell tumors (gct) also benefit from this intensive therapy. in comparison to allogeneic transplantation, this treatment is known for lower complication rates, e.g. infections. however previous studies have schown that treatment related toxicity may not be underestimated and depending on the conditioning regimen used. methods: we retrospectively analyzed patients ( cases) who underwent hd-ct plus auto pbsct between and in a single-center study. to anlyze the incidence of infections depending on the conditioning regimen, we formed the following categories based on the agiho: no infections, neutropenic fever, sepsis and severe sepsis. results: the median age in this analysis was years; . % were male. the most frequent diagnosis was mm ( . %) receiving high dose melphalan (mel), followed by malignant lymphoma ( . %) receiving beam (bcnu, etoposide cytarabine, melphalan) and relapsed/metastatic germ cell tumours (gct) ( . %) receiving high dose carboplatin/etoposide (ce). % of all patients developed severe sepsis, patients had to be ventilated and patients died. sepsis was documented in . % of all cases ( cases). the majority of patients ( . %, cases) developed neutropenic fever and . % ( cases) didn´t have any infection complications. the beam conditioning regimen showed the highest tendency to result in a septic course ( . %), followed by ce ( . %) and mel ( . %). the most commonly documented pathogen in blood cultures was s. epidermidis ( . %), followed by e. coli ( . %) and s. mitis ( . %). only in one blood culture we detected a multi-resistant pathogen ( mrgn e. coli). p. aeruginosa was detected in blood cultures ( . %), l. monocytogenes in ( . %) and s. aureus in ( %). . % of all patients developed diarrhea, only in . % of these cases we could detect c. difficile. the conditioning regimen shows no significant effect on the incidence of c. difficile. the mean neutropenic period was . days in malignant lymphoma patients, followed by . in mm patients and . days in gct patients. the hospital discharge, calculated from the day of transplantation, was significantly different: for malignant lymphoma the mean was . days, for mm . days and for gct . days. conclusions: our data correspond to former published results by many groups. the beam regimen shows the highest infectious complication rate followed by ce and mel. the duration of neutropenia and hospital stay depends on the conditioning regimen. the type of infectious complication doesn't effect the progression free-and overall survival in our analysis. disclosure: nothing to declare. impact of donor and recipient cytomegalovirus serostatus on outcomes of unrelated allogeneic haematopoietic stem cell transplantation background: cytomegalovirus (cmv) is an important cause of morbidity and mortality in allogeneic haematopoietic stem cell transplant (hsct) patients. the aim of our study is to evaluate the outcomes of our cmv seropositive recipients who received grafts from seropositive unrelated donors (d+r+) compared with grafts from seronegative unrelated donors (d-r+). methods: this is a retrospective single center study on a series of cmv seropositive recipients who underwent hsct from unrelated donors between febuary to july . a total of patients were analyzed. their clinical course and laboratory results were reviewed for evidence of cmv reactivation and/or cmv disease. we defined cmv infection as detection of cmv reactivation or primary infection by antigenaemia or polymerase chain reaction (pcr) assays, but was not accompanied by signs and/or symptoms suggestive of a systemic disease. cmv disease occurred when cmv was isolated from any site in association with organspecific signs and/or symptoms. monitoring for cmv infection commenced upon engraftment (approximately day + ). peripheral blood samples were sent twice a week for cmv antigenaemia or cmv quantitative pcr. the duration of twice weekly monitoring was at least about days. longer monitoring was performed in patients who experienced cmv infection after hsct. results: all patients received graft-versus-host-disease (gvhd) prophylaxis using anti-thymocyte globulin (atg) at . mg/kg in addition to cyclosporin or tacrolimus. among the entire cohort of patients, ( %) had cmv infection, including ( . %) out of patients from the d-r+ group and ( %) out of patients from the d+r + group. patients ( . %) from the d-r+ group and patients ( . %) from the d+r+ group had ≧ cmv reactivation above the threshold for preemptive therapy respectively; p= . . patients developed cmv disease, ( . %) from the d-r+ group and ( . %) from the d +r+ group. cmv resistance to both foscarnet and ganciclovir was detected in patients ( . %) from the d-r+ group but none from the d+r+ group. patients died due to cmv disease, both were from d-r+ group. year overall survival (os) were % versus % for d-r+ group and d+r+ group respectively; p= . . median survival was not reached at years. year non-relapse mortality (nrm) were % for d-r+ group and % for d +r+ group respectively; p= . . conclusions: the incidence of recurrent cmv infection was higher in the d-r+ group compared to the d+r+ group. there were no statistically significant differences between the groups in terms of os and nrm. however, there was a trend towards higher nrm in the d-r+ group compared to d+r+ group. our findings suggest that for matched unrelated hsct, it may still be important to select a seropositive donor for a seropositive recipient. disclosure: none background: it´s known that some patients submitted to allogeneic stem cell transplantation (asct) could present a greater susceptibility to infection even when they are in long term complete remission or potentially cured. this fact is related to the dynamic of immunological recovery that is variable in every single patients and it is dependent from many factors: the haematological disease, the conditioning regimen, the age of patient and donor, the number of stem cell and lymphocytes infused, the anti-gvhd prophylaxis, the use of anti-thimoglobulins and others. in clinical practise we can observe patients who are potentially cured, who tapered and stopped the immunosuppressive treatment months or years ago and who are suddenly graved from opportunistic infections. the largest part of these infections is represented from varicella-zoster virus (vzv) cutaneous eruption. methods: in this report we retrospectively analysed a monocentric cohort of patients submitted to asct for haematological malignancies from a median time of months. all of them were free of disease. they stopped the immuno-suppressive treatment in a median time of days after asct (range: - ) and did not present later chronic gvhd needing treatment neither other moderate or severe chronic post transplant complications nor other diseases. prophylactic treatment with anti viral agents (acyclovir or valacyclovir) has been conducted simultaneously to immuno-suppressive treatment and for a period ranging between to months after its suspension. in this cohort of patients we considered the incidence of vzv eruption occurred after the suspension of the immunosuppressive treatment, and we analysed the immunological recovery in terms of lymphocytes sub-population after , , and months from asct. results: of these patients considered, developed at least one vzv manifestation. all the vzv presentation were cutaneous, we did not observe neurological, ophthalmic or visceral presentation. all the vzv manifestation occurred in patients who ended the anti-viral prophylaxis. median time of presentation was days after asct (range: - ) the remaining patients did not present vzv manifestation nor other kind of opportunistic infection despite the absence of anti-viral prophylaxis. the analysis of lymphocyte sub-population after - - and months did not show a significant difference in b, t, t , t and nk lymphocytes in the different post transplant period. conclusions: vzv reactivation seems not to be correlated with the number of the different lymphocyte subpopulations in the post transplant period. actually it is not possible to distinguish patients more suitable of vzv reactivation on the basis of lymphocyte sub-populations analysis, so anti-viral prophylaxis should be prolonged for a medium period after suspension of immuno-suppressive drugs. in absence of anti viral prophylaxis a careful clinical surveillance should be performed in order to treat early eventual vzv manifestations. disclosure background: infection and disease cytomegalovirus (cmv) are common problems in patients undergoing hematopoietic stem cell transplantation (hsct). cmv infection has a high overall seroprevalence, therefore, during the first days post-hsct, it is important to prevent reactivation of cmv. the international clinical recommendation is the use of ganciclovir as prophylaxis in hsct patients; however, the cost of this treatment is not accessible for our population. in this respect it has been used as an alternative valganciclovir because of its lower cost and oral administration. our study´s aim was to assess the response and safety of valganciclovir in comparison with ganciclovir to prevent viremia and cytomegalovirus disease in patients undergoing allogeneic hsct methods: a retrospective study was performed on patients who receive an hsct-allo between january and august . participants were enrolled in two groups according to prophylaxis treatment: (a) ganciclovir mg/k once daily and (b) valganciclovir mg twice daily for days pretransplant, at day + ; viremia was measured by pcr. demographic and clinical information was collected from medical records and furthermore analyzed in spss v . results: sixty-eight patients were enrolled in the study, % male, the median age was years ( - ) with the following diagnoses: acute lymphoblastic leukemia %, acute myeloblastic leukemia . %, granulocytic chronic leukemia . %, myelodysplastic syndrome . %, dendritic cell neoplasia . %, and aplastic anemia . %. ninety-one percet of the patients received a transplant from an identical hla donor and . % received a haploidentical transplant. thirty-four patients received ganciclovir (g ) and thirtyfour valganciclovir (g ). median age was vs years (p= . ), intermediate risk cmv % vs (p= . ), associated bacterial infections was %vs % (p= . ), and fungal infections % vs % respectively (p= . ). the reactivation by cmv was presented in % vs % respectively (p= . ). there were no significant differences in fever, bacterial isolation, dysfunction or graft failure, presence and degree of acute or chronic gvhd and relapse of the disease. the most relevant characteristics and complications are described in table . within the whole group there were deaths, % in the ganciclovir group and % in valganciclovir group (p= . ), overall survival -year was % vs % (p= . ) respectively; in both groups % was associated with relapse and % associated with transplantation. conclusions: ganciclovir and valganciclovir were effective in preventing the reactivation of cmv, the only statistically significant difference was that the presentation of the disease appeared earlier in the valganciclovir group. no difference in toxicity between the groups was identified. disclosure: none declared background: invasive pulmonary aspergillosis (ipa) is a severe and serious complication that occurs in the immediate post-transplant period due to severe neutropenia or late usually following prolonged corticosteroid therapy during treatment of graft-versus-host disease (gvhd). the objective of this study is to analyze the epidemiological, diagnostic and evolutionary characteristics of this major complication over a period of years. methods: from january to december , patients (pts) received an allogeneic hematopoietic stem cell transplantation (allo hsct) for malignant and nonmalignant haematological diseases. during the transplant procedure, anti-infectious prophylaxis consisted of pts isolation, digestive decontamination, fluconazole and aciclovir. secondary prophylaxis done for pts with prior history aspergillosis. during the follow-up, a standard chest x-ray is performed systematically at each control or in case of clinical signs a thoracic ct scan is requested from suspicion. the diagnosis of ipa is made according to the criteria of the eortc-msg based on the predisposing criteria of the host and clinico-radiological criteria (possible infection). galactomannan antigen and histopathology criteria are not common practice. results: a total of ipa episodes ( %) were identified in pts (aml: , aa: , all , cml , mm ) of median age ( - ) , sex ratio: . . all of them had transplantation from a family donor (geno-identical: , haplo-identical: ) with conditioning by chemotherapy alone and a graft of csp ( pts) and peripheral stem cells-bone marrow ( pts). all pts had at least one predisposing risk factor: antecedent of aspergillosis ( pts), prolonged neutropenia> d ( pts), acute gvhd ( pts), chronic gvhd ( pts), prolonged corticosteroid therapy ≥ , mg /kg/day exceeding days ( pts). the diagnosis of api was possible on average at j ( - ) after appearance of clinical signs (in all cases) and evocative radiological in cases (in cases, the standard chest x-ray was normal). at the time of thoracic ct scan, pts ( %) had characteristic signs: halo sign ( pts), crescent sign ( pt) and cavity ( pts). other minor radiological signs are found in the other pts. empirical first-line antifungal therapy was started as monotherapy in pts (voriconazole: , caspofungin: pts) or in combination in pts. a secondline treatment was required in pts for failure after an average duration of days . three pts presented a second episode after an average delay of months ( ) ( ) ( ) ( ) with a favorable evolution of resumption of thetreatment. fourteen pts ( %) are alive with complete resolution after a median treatment time of months ( - ). twelve pts ( %) died rapidly on average days after diagnosis (ipa , relapse of his disease: ) conclusions: ipa occurring after an allograft of allo-hsct is a severe complication with high mortality. it is essential, in each case, to identify the pts with risk factors, perform a thoracic ct-scan, send serum serology for apergillus galactomannan antigenand start specific treatment as soon as possible while waiting to be able to reinforce the diagnosis by direct examination or sputum or brochoalveolar lavage with aspiration. disclosure: nothing to declare background: cmv (cytomegalovirus) has a prevalence varying between - %. its pathogenicity is relatively low in the general population, usually resulting in a selflimiting viral illness. in an immunosuppressed host, infection can lead to life threatening illness. disseminated cmv infection can manifest in a number of organs and is diagnosed using internationally accepted criteria. in the post solid organ and stem cell transplant (sct) setting, it is postulated that it is viral reactivation, rather than primary reinfection that leads to cmv viraemia. prevention of reactivation requires the presence of a competent immune system, mediated by t-cells. this accounts for the increased incidence in intensive and t-cell depleting sct conditioning regimens. despite improved outcomes following the introduction of cmv monitoring by pcr and pre-emptive treatments (current uk guidance), cmv pneumonitis still carries a high mortality. the use of cmv specific immunoglobulins (cmvig) for the treatment of this complication is generally not recommended post chemotherapy or sct in haematological cancers due to lack of evidence. however, cmvigs are widely used in the setting of cmv reactivation post solid organ transplants. we report the use of cmvig in patients with suspected cmv pneumonitis at a single uk centre. the aims of this retrospective study were to establish safety and review efficacy in this highly immunocompromised group of patients. methods: data was collected retrospectively on the use of cmvig in patients with haematological cancers post sct or chemotherapy alone between and at manchester royal infirmary, uk. all patients included had cmv positive pcr in blood (and or from bronchoscopy), as well as high resolution ct imaging evidence of cmv infection. the data was sourced from pharmacy database and crossreferenced with a departmental list. for each patient identified, case notes and prescriptions were sourced. data collected included patient baseline characteristics, timing of treatment, number of doses of cmvig and outcome. results: eight patients received cmvig for suspected cmv pneumonitis. seven patients were post sct and one patient was severely immunosuppressed with chemotherapy alone. median age was years (range - ). the cmvig regimen used was ml/kg of cytotect ® on days , , and , followed by ml/kg every four days until resolution of symptoms. there were no infusion related reactions observed. patients received a median of doses of cmvig. four out of patients responded to the treatment and showed full recovery but only are alive and well to date. conclusions: this study shows that the use of cmvig is safe in the post-sct setting of acutely unwell patients with multi-organ failure. despite limitations of retrospective studies, there appears to be benefit for the use of cmvig in our patient population, with % of patients showing a full recovery from that episode. allogeneic sct plays a confounding role in the outcome of patients although the numbers in our study are small. there is clearly a need for better treatments of cmv pneumonitis. cmvig is a promising treatment but further studies are needed to identify the optimal dosing regimen and provide evidence of efficacy. disclosure: biotest-honaria p abstract withdrawn. background: the risk of fungal infection related to allogeneic transplantation is a well-known cause of morbidity and mortality. the main agents implicated are yeast during the neutropenic period and filamentous fungi after this period. methods: we decided to evaluate the effectiveness of a prophylactic regimen containing fluconazole since day - . after discharge fluconazole was kept until day or or switched to posaconazole in high-risk patients. patients with gvhd under steroids were kept under prophylaxis.the group of high risk patients was defined by one of the following variables: -non related donors -atg, campath or fludarabine in the conditioning -presence of gvhd with need of steroids above . mg/kg we have analyzed the patients submitted to allobmt during and . all patients were first admitted to an isolation room with hepa filters.patients under secondary prophylaxis were excluded. breakthrough fungal infections during the first year and toxicity leading to discontinuation was evaluated. results: sixty six patients were included with transplants. male/female ratio was / . the age range was . - yo with a median of . malignant ( ) and nonmalignant ( )diagnosis were included. donor type was related ( ) haploidentical ( ) and non-related ( ). the conditioning regimen includes atg in , campath in and fludarabine in . fourteen patients were treated after discharge with fluconazole and with posaconazole. three patients fluconazole were switched to micafungin for hepatic toxicity, two cases to amphotericin due to persistent fever and in one case to caspofungin for a proven fungal infection (candida parapsilosis in blood stream in day + ). after discharge and during the first year of follow-up a single case of possible fungal infection was diagnosed, in a patient with gvhd with a lung nodule. conclusions: during the neutropenic period after transplantation the main risk of fungal infection is associated with candidiasis. the greatest risk of aspergillosis occurs later and have a significant relation with gvhd. except for candida parapsilosis the main source of yeasts are the gi tract. the main source of aspergillus are aerosolized particles retained by hepa filters. in patients without a previous episode of fungal infection the main risk of filamentous fungi occurs only after discharge. we conclude that fluconazole alone or followed by posaconazole in high risk patients is a feasible and effective regimen for primary prophylaxis, in allogeneic transplantation. disclosure background: bk virus-associated hemorrhagic cystitis (bkv-hc) has emerged as a serious infection after hematopoietic stem cell transplantation (hsct). it is characterized by painful hematuria due to hemorrhagic inflammation of the urinary bladder mucosa, this causes significant morbidity, prolonged hospital care with extensive nursing requirements and increases in healthcare costs. the purpose of this study is to determine the incidence, risk factors, and duration of treatment in our center. methods: we performed a retrospective review of hsct patients at luis calvo mackenna children´s hospital in santiago, chile diagnosed with bkv-hc, from st january to th november . we investigated the incidence, risk factors and duration of treatment of bkv-hc in paediatric patients undergoing hsct over a months period. bkv-hc was defined as bk virus (bkv) detection in urine by pcr testing in association with clinical symptoms and hematuria grade or higher. sixty-seven patients were trasplanted during this period. results: eleven patients were diagnosed with bkv-hc at our institution, only one with bk viremia. the cumulative incidence of bkv-hc in our series was %. all of them were treated with cidofovir. the median age at diagnosis was years old (range: - y.o.). the median time from hsct to hemorrhagic cystitis (hc) was days (range: - days), the median length of treatment was weeks (range: - ). all patients received myeloablative conditioning regimens and used cyclophosphamide ( %); ten ( %) were unrelated cord blood transplant recipients and nine ( %) used antithymocyte globulin. a concomitant viral reactivation (cmv/vh ) was demonstrated in six ( %) patients. no patient died due to bkv-hc or its complications, but in the follow up three patients died, one in relapse and two of other post transplant´s complications. conclusions: bkv-hc is the result of a complex interaction between patient characteristics, donor type and conditioning regimen intensity. these patients experienced significant morbidity and prolonged treatment. in our cohort bkv-hc of all patients but one were transplanted with an unrelated umbilical cord blood unit, all of them received myeloablative conditioning regimen with cyclophosphamide and most of them received anti-thymocyte globulin. we also observed frequently co-existence of viral infections from herpes family as cmv and vh . the main limitations of this work are its retrospective nature and it´s from a single center. more studies are necessary to better understand the epidemiology and risk factor associated with bkv-hc and the morbidities associated with its treatment. disclosure: nothing to declare how we manage hhv- reactivation in the posttransplant setting oscar borsani , anna amelia colombo , daniela caldera , paolo bernasconi university of pavia, san matteo hospital, pavia, italy background: hhv- encephalitis is a life-threatening complication in the post-transplant setting and it develops in about % of patients receiving traditional hsct. several risk factors were described. a differential diagnosis between hhv- encephalitis and other neurological complications is extremely important but often not-easy to achieve because of the highly heterogeneous clinical and radiological features and complexity of interpretation, especially in transplanted patients. here we described vignettes that represent and highlight distinct problems in the diagnosis and management of transplanted patients with suspected hhv- reactivation. methods: we collected the clinical, laboratory and radiological (electroencephalogram, brain mri and brain ct) data of transplanted patients who developed a neurological syndrome suspected for hhv- reactivation. hhv- was detected on serum and csf using rt-qpcr. results: ) a -years-old patient developed a diffuse erythema and subsequent encephalitic syndrome following hsct. the brain mri revealed clear signs of limbic encephalitic and searching for hhv- on serum and csf revealed . copies/ml and . copies/ml respectively. an antiviral therapy was started but no clinical benefit was achieved. ) a -years-old patient developed a typical neurological syndrome without brain mri findings of encephalitis and with no evidence of skin involvement. the lumbar puncture and csf analysis showed a total of . hhv- dna copies/ml. antiviral therapy with ganciclovir and foscarnet was promptly started with clinical improvement and a drastically reduction of hhv- dna on both csf and serum. a new brain mri revealed an acute limbic encephalitis. ) a slight neurological syndrome consisting of confusion and amnesia developed in a -years-old-patient. brain mri findings were compatible with a wernicke syndrome, but no improvement of neurologic symptoms were obtained with thiamine supplementation. csf analysis did not revealed hhv- dna, which was detected at low copies number on serum analysis. a second brain mri was conclusive for limbic encephalitis, so an antiviral therapy with foscavir was started and radiological but not clinical improvement was noted. the patient died after few days. ) in the last case we present a -years-old patient who developed a clinical picture of encephalopathy (i.e. amnesia, ataxia, drowsiness, weakness, depression) with rapid progression to coma after seventy-eight days from hsct. a brain mri showed a slight contrast enhancement in parietal-occipital regions. during the recovery phase from conditioning-induced cytopenia, an increasing in serum hhv- dna was detected. searching for hhv- dna on donor's follicles showed a chromosomally integrated hhv- (cihhv- ). cyclosporin a (csa) was interrupted and neurological improvement was observed in the following hours: a diagnosis of pres was made. conclusions: hhv- encephalitis should be suspected in transplanted patients with a clinical syndrome of encephalopathy. pcr detection of hhv- dna in csf associated with either typical brain mri abnormalities or a clinical diagnosis of nonspecific encephalopathy must lead to the urgent initiation of systemic antiviral treatment. if an increase of both serum hhv- dna and wbc is detected, a cihhv- should be confirmed. pres is an important differential diagnosis in transplanted patients which developed an encephalitic syndrome. disclosure: nothing to declare background: cytomegalovirus (cmv) infection is a major cause of morbidity and mortality after hematopoietic stem cell transplantation (hsct). it causes end-organ disease, multi-organ dysfunction syndrome, graft failure, increased susceptibility to infections and gvhd. greatest risk of cmv infection in a seropositive host is the reactivation of latent virus. methods: a prospective descriptive study performed at armed forces bone marrow transplant centre, rawalpindi, pakistan from dec to sep . hundred consecutive patients who underwent hsct were followed with weekly cmv dna quantitative pcr from engraftment till day for cmv reactivation. patients in whom cmv pcr showed more than copies/ml were treated with antiviral therapy. factors associated with cmv reactivation, outcome of antiviral therapy and effect of cmv on transplant outcome is studied. results: out of cases, were hla matched siblings, were matched family donors and were haploidentical transplants there were males and females. mean age was . ± . years. fourty-two transplants were done in thalassemia, in aplasia, in leukemias and in other hematological disorders and immune deficiencies. ninety-eight recipients and all the donors were cmv seropositive before hsct. cmv reactivation was seen in patients and of them had cmv viral load more than copies/ml and patients had cmv viral load less than copies/ml. nineteen patients had no cmv reactivation. mean time to reactivation since transplant was ± days. valganciclovir was given in patients due to ease of administration and six patients were treated with ganciclovir during their hospital stay. only one patient had resistant disease. mean time to clear viremia was ± . days. the patients having viral load less than copies/ml, subsequently cleared cmv without any treatment. antiviral agents; ganciclovir and valganciclovir were equally effective for treating cmv infection with % efficacy, however, more adverse effects were seen with ganciclovir. myelosuppression i-iii was seen in % patients treated with valganciclovir and in % treated with valganciclovir. renal impairment i-ii was seen in % of valganciclovir and % of ganciclovir treated patients. steroid administration was strongly associated with cmv reactivation (p = . ). no statistically significant association was found with the use of atg, gvhd, underlying disease, abo or gender mismatch. os was . % and . % in with and without cmv reactivation (p= . ) and dfs was . % and . % in with and without cmv reactivation (p= . ) conclusions: cmv reactivation was seen in % of the transplant recipients, this is higher compared to the western world due to high cmv seropositivity is this region. steroids administration in post-transplant period significantly increase the risk of cmv reactivation. preemptive therapy with valganciclovir effectively treats cmv reactivation with acceptable side effects. viral threshold for treatment should be decided considering the regional endemicity. cmv adversely affects the transplant outcome in terms of dfs and os. disclosure: no conflict of interest. acute nephritis requiring nephrectomy caused by adenovirus (hadv) and human polyomavirus bk (bkpyv) following allogeneic hematopoietic stem-cell transplantation in a patient with ph+ all background: adenovirus infection represents an important cause of morbidity and mortality after allogeneic hematopoietic stem cell transplantation (allo-hsct), with no established therapy. although different organs may be affected by disseminated hadv infections, kidney involvement has been rarely reported. co-infection of hadv and bkpyv are common complication in patients undergoing allo-hsct, but recent studies demonstrate that bkpyv may facilitate the replication of hadv and lead to elevated viremia with increased virulence and serious clinical consequences. here we report a case of an adult patient who required a monolateral nephrectomy due to hadv pyelonephritis as an early complication of allo-hsct for philadelphia-positive acute lymphoblastic leukemia (ph+ all). methods: in september , an ethiopian gentleman was diagnosed with ph+ all at the age of years. he was treated with polychemotherapy in association with the tyrosin kinase inhibitor imatinib mesylate achieving a complete remission (cr). one year later, due to disease relapse with cns involvement, he was started on vincristine and dexamethasone plus imatinib treatment and in april he was referred to our bmt center from ethiopia. upon confirmation of the p ph+ b-all diagnosis, therapy with the scr/abl dual inhibitor dasatinib associated to intrathecal chemotherapy was started and a salvage treatment with inotuzumab ozogamicin followed by an allogeneic hsct from a hla-identical brother was planned. having achieved a documented molecular cr disease status, in june the patient underwent allo-hsct following the fludarabine-melphalan reduced-intensity conditioning regimen. graft-versus-host prophylaxis included anti-thymocyte globulin, cyclosporine and mycophenolate mofetil results: on day + post-transplantation the patient developed macro-hematuria due to hemorrhagic cystitis and a ct scan unveiled a left pyelonephritis with marked kidney enlargement. kidney microbial investigations were all negative. at the same time, hadv viremia with very high copy number (> cp/ml) was documented and also elevated bkpyv (> cp/ml) viruria and viremia ( cp/ml). the genotyping of hadv evidenced serotype b mainly involved in infections of the urinary tract. treatment with cidofovir was immediately started; nonetheless, due to rapid clinical worsening despite maximal antibiotic therapy, on day + a left nephrectomy was performed, which led to a subsequent progressive resolution of the clinical symptoms and negativization of hadv and bkpyv viremia and viruria. pcr real time performed on the kidney tissue unveiled very high concentration of hadv copy number. conclusions: acute pyelonephritis due to disseminated hadv infection may represent a possible cause of severe complication following allo-hsct. monitoring of hadv copy number is helpful to evaluate infection severity and response to treatment. co-infection of hadv and bkpyv in immunocompromised patients should be always considered likely to worsen clinical course and outcome. disclosure: nothing to declare background: infection is a major cause of morbidity and mortality in patients (pts) receiving an allo-hsct. its severity is related primarily to the depth and duration of neutropenia. febrile neutropenia (fn) is defined as a neutrophil count below cells/mm and a fever ≥ . °c at a single measurement or≥ °c times at one hour intervals. the objective of our study is to analyze the epidemiological, clinical, biological characteristics of febrile episodes (fe) occurred in pts who benefited an allo-csh over a period of years. methods: from january to december , allo-hsct were performed in pts including sibling-hla identical, haplo-identical and phenoidentical for essentially acute leukemia ( pts, %), acquired and congenital aplasia ( pts, %). the median age is years ( - ) and sex-ratio (m/f): . . prophylaxis consisted on isolation sterile room with laminar flow, digestive decontamination, fluconazole and aciclovir. nine pts ( . %) were infected at the time of hospitalization (cellulitis , pneumoniae , bacterial angina , veinitis , bronchial pneumonia , furuncle cutaneous ) requiring treatment with antibiotics. conditioning regimen is myeloablative in all pts. anti-thymocyte globulin is used in pts ( . %). peripheral blood stem cells (pbsc) are used in pts ( %) with an average level of cd + cells: , . /kg ( . - . ) , bone marrow (bm) in pts with a mean level of nucleated cells: . x /kg ( . - . ) and the association of pbsc-bm in pts (haplo-identical). at each fe, are practiced: chest x-ray, procalcitonin test, blood culture, microbiological study of urine and stool (if diarrhea). results: all pts showed aplasia with an average duration of days ( - ), neutrophil engraftment was observed at day ( - ). one hundred and twenty-nine pts ( . %) presented fe with an average of . per pt. eleven pts ( %) had fe or more. forty nine ( , %) fe are clinically documented (digestive: , skin: , pulmonary: , urinary: , oto-rhino-laryngology: ). the blood cultures are made at fe, fe are microbiologically documented ( %): gram-positive bacteremia in % (mainly coagulase negative staphylococci) and gramnegative bacilli in % of cases. procalcitonin test performed during fe: normal ( cases), probable infection ( cases), probable sepsis ( cases), severe sepsis ( cases) and septic shock (one case). empirical double antibiotic therapy is initiated in pts without waiting for the results of the microbiological study. this association was sufficient in pts ( %). the transition to a second line was needed in pts ( . %) and third line in pts ( %). antifungal is added in cases ( %). eight pts benefited from g-csf. the evolution is favorable in fe ( . %), apyrexia obtained after an average of . days . three pts died ( %) by severe sepsis on a durable aplasia, of which had a cellulitis before the conditioning. conclusions: fe increase morbidity and mortality in allo-hsct so prophylactic measures are essential. empirical antibiotics treatment has to be instituted very quickly in the absence of documentation. disclosure: nothing to declare p abstract withdrawn. atsushi satake , masaaki hotta , ryo saito , akiko konishi , hideaki yoshimura , takahisa nakanishi , shinya fujita , tomoki ito , kazuyoshi ishii , shosaku nomura kansai medical university, osaka, japan background: cytomegalovirus (cmv) infection remains a common complication after allogeneic hematopoietic stem cell transplantation (ahsct), which results in increased morbidity and mortality. letermovir is a novel anti cmv drug that inhibits the cmv-terminase complex. the purpose of this retrospective study is to elucidate the efficacy and safety of cmv prophylaxis with letermovir early after ahsct in clinical practice. methods: we retrospectively analyzed the incidence of cmv infection, cmv disease, preemptive therapy, adverse events through week after ahsct, the rates of engraftment and overall survival. all patients underwent ahsct in our institution for hematopoietic malignancies between may and nov . data collected in this study included patient's characteristics such as age, sex, disease status, donor source and cmv disease risk. cmv infection was evaluated by cmv antigenemia. this study was approved by the research ethics committee of the faculty of medicine, kansai medical university. results: thirteen patients (male , female ) underwent ahsct and received cmv prophylaxis with letermovir. the median age was years (range, - years). overall, of patients ( . %) were considered to be at high risk for cmv, including patients ( . %) with haploidentical donors, and ( . %)with mismatched, unrelated donors. all patients began letermovir from day after ahsct, and achieved engraftment (median , - days). no patient developed cmv disease and required preemptive therapy. one patient died of treatment-related mortality, and patients died of acute gvhd. although one patient discontinued letermovir before day after ahsct because letermovir was suspected to be a cause of persistent nausea, severe adverse events were not observed. conclusions: it is still unknown whether cmv prophylaxis with letermovir improves os and reduces trm; however, our data suggests that cmv infection is considerably inhibited by administration of letermovir early after ahsct. clinical trial registry: not applicable. disclosure: the authors declare noconflicts of interest for this study. background: cytomegalovirus (cmv) is cause of increased morbidity and mortality after transplantation of hematopoietic cells. the pathogenesis of cmv disease or infection is complex with multiple interactions with the immune system, mainly in acute and chronic graft-versus-host disease (gvhd). the aim of this study is to analyze the risk factors for the reactivation of cmv in patients undergoing allogeneic hematopoietic cell transplantation (hct). methods: prospective descriptive study of the risk factors for the reactivation of cmv in the described population. univariate and multivariate analysis of the predisposing factors were performed: donor graft, treatment with corticosteroids, use of antithymoglobin, serologic status, conditioning regimen and the presence of gvhd. results: during the period between august until january , patients were evaluated. . % (n: ) had reactivation of cmv. average reactivation was days post transplant. both (recipient and the donor) had positive cmv igg in . %. in the univariate analysis, the reactivation of cmv was associated with haploidentical transplantation (p: < . ), with the use of corticosteroids (p: < . ) and gvhd (p: < . ). in the multivariate analysis, the haploidentical transplant maintained its statistical significance in comparison with the related allogeneic transplant (p: . , or: . ; ic %: . - . ) as well as the use of corticosteroids (p: . , or: . ; ic %: . - . ). % of patients receiving corticosteroid treatment had grade ii / iii gvhd. the serologicac status, myeloablative conditioning regimen and the use of atg showed no statistically significant association. conclusions: in patients undergoing allogeneic transplantation, were found as risk factor to reactivation, those who received haploidentic transplantation and treatment with corticosteroids. another risk factor that showed greater reactivation was the presence of gvhd. disclosure: nothing to declare methods: a y/o male was referred for allogeneic transplant following cycles of induction therapy for aml with complex karyotype and axsl mutation having achieved complete remission following the first cycle of chemotherapy. his first induction cycle was complicated by a perianal myeloid sarcoma which became infected and required surgical drainage and formation of a defunctioning colostomy. results: following allogeneic transplants, the first complicated by secondary graft failure and the second by primary graft failure he presented with two skin lesions, with a third lesion adjacent to his stoma developing shortly after admission. all lesions were erythematous with central necrosis and progressed rapidly in size over hours. biopsy of the skin and para-stomal lesions revealed fungal mycelia, with culture subsequently identifying rhizopus oryzae. initial treatment was with liposomal amphotericin b mg/kg/day followed by dose escalation to mg/kg/day due to the development of new skin lesions. the patient had been taking posaconazole (tablet) prophylaxis since his first allogeneic transplant and peripheral blood drug levels checked at the time of admission were therapeutic confirming that this was a breakthrough fungal infection. consequently posaconazole was stopped and isavuconazole added to the treatment regimen. surgical assessment was undertaken but surgery was deferred on the basis of high risk due to the extent of the infection and the patient´s profound pancytopenia. the organism was tested for in vitro susceptibilitiy and found to be resistant to posaconazole (mic > mg/l), with borderline resistance to isavuconazole (mic mg/l) and sensitive to amphotericin b (mic . mg/l) (phe mycology reference laboratory, england). isavuconazole was therefore stopped and the patient was managed with liposomal amphotericin b along with daily granulocyte infusions. he underwent a third allogeneic transplant using a different unrelated donor and stable engraftment was achieved. post transplant there was initially an increase in the size of the para-stomal lesion, but no new skin lesions developed. following engraftment he underwent resection of the stomal lesion, with primary closure and re-siting of his stoma. amphotericin b was replaced by isavuconazole prophylaxis on discharge and he continues to make an excellent recovery. conclusions: whilst aspergillus species remain the most common cause of invasive fungal infections in allogeneic transplant patients, other species including the mucorales are seen, and generally associated with poorer outcomes. whilst there are standardised methodologies for susceptibility testing, fungi specific cut offs based on clinical outcomes are only available for a limited number of species/ antifungal agents. in this case, susceptibility testing demonstrated resistance to posaconazole which was consistent with the clinical presentation of invasive infection despite therapeutic levels of posaconazole. it is also worth noting that an estimated % of r. oryzae isolates in the uk are resistant to posaconazole. treatment with high dose amphotericin b resulted in improvement in small skin lesions with stabilisation of the larger stomal lesion until count recovery allowed surgical resection. background: total depletion of innate and adaptive immune cell populations occurs after intensive chemotherapy and hematopoietic stem cell transplantation (hsct). both t and b lymphocyte pools are restored slower that myelomonocytic populations. hsct patients are at high risk for bacterial and viral infections at early terms (< days) post-transplant. the reconstitution of the immune system depends on the time required for stem cell recruitment, differentiation, expansion, maturation and release into the bloodstream. restoration terms for myeloid cells after hsct are usually defined as the st day with neutrophil count of ≥ . x ^ /l with mean recovery terms of to days. high occurrence of cytomegalovirus (cmv) in hsct patients mostly result from reactivation of a latent virus acquired in early childhood. however, delayed immune reconstitution and subsequent infections such as cmv, adenovirus (adv) or herpes (hhv- ) diseases are not unusual and still constitute a major cause of death in peru. methods: peruvian pediatric patients (n= ) diagnosed with aplastic anemia, mds, aml or all underwent a haploidentical hsct performed with the clinimacs device. patients treated were separated in two groups. the group of patients who received viral prophylaxis (ganciclovir) was compared to the group that did not receive any prophylaxis treatment. viral reactivation was confirmed by pcr test twice a week and clinical signs within days after hsct. results: in the group that didn´t received prophylactic treatment, engraftment occurred close to day post haplo-hsct and none of the patients developed gvhd (graft versus host disease). nevertheless, incidences of cmv, hhv- and bkv infections before day post haplo-hsct were still high. an overall survival (os) over % with an ic % was reached at the end of the first year. on the other hand, the group of patients that received prophylaxis with ganciclovir did not developed gvhd and reached the engraftment close to day with a very low viremia incidence after the first month post haplo-hsct. all viral reactivations were caused by cmv and the os was over % with an ic % at the end of the first year. previous prophylaxis to both the donor and the receptor with ganciclovir ( mg/kg) every hours before and during the conditioning regimen has allowed a better control of viral reactivation. conclusions: the attempts to improve immune function and reduce nonrelapse mortality from infectious complications without increasing gvhd have focused on a partial t cell depleted graft, such as t cell depletion (tcr α/β). this graft retains a large numbers of effector cells, such as tcr γ/δ and natural killer cells. however, delayed immune reconstitution and subsequent infections are a big issue. a novel partial t cell depletion strategy such as depleted naïve t cells (cd ra+ t cells) could enhance the recovery of immune function after haplo-hsct because donor pathogen memory t cells from the donor are retained. it is necessary to increase the studies and the database to set the scheme of previous prophylaxis to the recipient to contain the viral reactivation and to help a rapid immune reconstitution. disclosure: no conflict of interest is declared information was recovered from the medical records. results: thirty-four patients were included, of them with the following diagnoses: acute leukemia ( ), granulocytic chronic leukemia ( ), dendritic cell neoplasia ( ), aplastic anemia ( ). % of the patients received a transplant from an identical hla donor and % received a haploidentical transplant. mean age's patients was years ( - ). prophylaxis with posaconazole was performed on % of the patients with identical hla and % on haploidentical group; the rest of the patients received fluconazole. the posaconazole group presented: fever %, mucositis gi-ii %, gastrointestinal toxicity gi-ii % (p= . ), hepatic toxicity %, kidney toxicity %, oral candidiasis %. during this period none of the patients presented invasive fungal infection in any group. there were deceases, one on each group and none related to a fungal infection. the overall survival was of the % versus % on the posaconazole group and the fluconazole group respectively. conclusions: the prophylaxis with posaconazole and fluconazole is effective on the prevention of invasive fungal infection on the first days. the toxicity was similar on both groups. posaconazole can be effective on the prevention of the haploidentical type. is necessary to continue following the patients with infection risk on a long-term period associated with the chronic gvhd. disclosure: none declared lymphoma these results open the question whether allo-sct should still be offered to these patients. methods: we aimed to define the role of allo-sct in refractory or relapsing after two lines de novo or transformed dlbcl patients, and its comparison with zuma- car-t cells trial (neelapu et al nejm ). we analyse long-term allo-sct results in de novo (n= ) or transformed dlbcl (n= ) out of the allo-sct performed in our institution between october to october . results: patients and transplant characteristics are summarized in table . complete response (cr) at days was , % and % of them remain in cr at months. with a median follow-up of months, -year progression-free survival (pfs) was % and -year overall survival (os) %, with a -year transplant-related mortality of %. refractoriness at the time of the transplant was associated with a poorer prognosis, with only out of refractory patients being long term survivors (figure ). similar results were reported for zuma- trial, with a best response of % cr retained in % of them at months. with a median follow-up of months, -months pfs was % and -months os %. patients characteristics did not differ in our series and zuma- , except that all the patients in zuma- were refractory prior to therapy (table ) . conclusions: although very few patients with de novo or transformed dlbcl are offering an allo-sct ( % of all allo-sct), this is a curative option in chemosensitive patients and with more mature data and longer follow-up than with car-t therapy; for these reasons, it should still be offer to these poor prognosis patients. moreover, almost all patients have now available donor, better graft-versus-host disease prophylaxis will decrease trm and morbidity, and new therapies will make more patients in sensitive disease before allo-sct. therefore, allo-sct and car-t cells are strategies to be discussed in every young patient with available donor. disclosure: honoraria as advisor or speaker from gilead ( methods: consecutive patients transplanted for hgbl (excluding burkitts lymphoma) between - in the ebmt database were included. data collected included age, sex, pathology subtype (hgbl (including subtypes), tfl, dhl), disease status at sct, conditioning (ma vs beam cam vs flu-mel-cam/atg), engraftment, day outcome, trm, os and pfs and eligibility for emea licensed indication of car-t therapy. results: fifty patients ( m, f) with a median age of at diagnosis and at sct were included. the subtypes included hgbl (n= ), tfl (n= ) and dhl (n= ). indications for sct were: primary refractory (n= ), relapse < months after primary treatment (n= ), previous autologous-sct (n= ) and dhl (n= ). the median lines of therapy was (range to ). conditioning used was cytbi n= , bu/cy n= , etop/tbi n= , flubucy n= , beamcam n= , fmc/t, n= . all patients engrafted with neutrophil > . /l at median days and platelets > /l at median days. the day mortality was % (progressive disease %, nrm %) with a year os of % and mortality due to progressive disease % and nrm %. disease subtype influenced outcome with an os for primary refractory hgbl, relapsed hgbl, tfl and dhl respectively of %, %, % and %. patients were eligible for a licensed car-t product. conclusions: the outcome of these high risk hgbl patients have an acceptable os of %, with relapsed disease being the commonest cause of mortality. patients with dhl have a particularly good outcome in this series; recent evidence indicates that some of these patients with a non-immunoglobulin gene associated myc translocation could be managed more conservatively (ash sehn). the outcomes achieved with allogeneic-sct in this series will provide a baseline for outcome assessment with a cart program. disclosure: nothing to declare background: immune checkpoint inhibitors (ici) allow to achieve a durable remission in patients with resistant or refractory (r/r) classical hodgkin lymphoma. however, the information about optimal duration of therapy and the prognosis of the patients after ici cessation is limited (manson, blood ). therefore, the optimal role of hematopoietic sct (hsct) in this patient group is not defined. our aim was to determine remission duration in patients who discontinued ici monotherapy after achieving complete remission (cr). methods: this analysis included patients ( male/ female) aged to (median years) with r/r classical hodgkin lymphoma who were treated with nivolumab ( mg/kg every days) and achieved cr. response was assessed by positron-emission tomography/computed tomography (pet/ct) using lyric criteria every month. after nivolumab therapy had been stopped the patients received no other treatment and disease was assessed every months by pet/ct. median follow-up after therapy discontinuation was ( - ) months. results: at the moment of therapy initiation ( %) patients had stage disease, ( %) patients had progressive disease (pd), ( %) patients had stable disease, ( %) patients had partial remission and ( %)complete remission; ( %) patients had b-symptoms and ecog score > . the median number of previous therapy lines was ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) . before nivolumab initiation high dose chemotherapy with autologous sct was performed in patients ( %) and ( %) received brentuximab vedotin. the median number of nivolumab cycles was ( - ). cr was achieved after median of ( - ) cycles. the median duration of therapy after achievement of cr was ( - ) months. at the time of analysis, all patients were alive, ( %) out of patients relapsed after therapy discontinuation. the median progression-free survival (pfs) for the total group was not achieved. among patients with relapse, the median time before pd was ( - ) months. after relapse all patients were retreated with nivolumab monotherapy or with chemotherapy combination. one patient achieved complete remission; -partial remission; -indeterminate response type . other patients are continuing the therapy and their response has not yet been evaluated. conclusions: while complete response was maintained in some patients at median follow up of months after nivolumab therapy cessation, the pfs plateau was not reached. we report that patients with relapse after nivolumab discontinuation sustained sensitivity to nivolumab and achieved a response during retreatment with nivolumab monotherapy or with chemotherapy combination. in patients with unsatisfactory response to nivolumab retreatment, hsct option should be considered. disclosure: nothing to declare high dose chemotherapy with autologous stem cell transplantation in primary central nervous system lymphoma: data from the japan society for hematopoietic cell transplantation (jshct) registry center hospital, tokyo, japan, national cancer institute, bethesda, md, united states, okayama university hospital, okayama, japan, kanazawa medical university, uchinada, japan, kyoto university, kyoto, japan, aomori prefectural central hospital, aomori, japan, yamagata unversity school of medicine, yamagata, japan, tenri hospital, tenri, japan, hiroshima university, hiroshima, japan, japanese data center for hematopoietic cell transplantation, nagoya, japan, nagoya university graduate school of medicine, nagoya, japan, shimane university hospital, izumo, japan background: high-dose chemotherapy (hdt) with autologous stem cell transplantation (asct) has been shown to improve prognosis of patients with central nervous system (cns) lymphoma. whereas the common regimen of hdt for pcnsl in the europe and the us is thiotepa-based regimen, e.g. bcnu-thiotepa, tbc (thiotepa-busulfan-cyclophosphamide), thiotepa-based regimen was only available before discontinuation of thiotepa in in japan. we report the results of asct for pcnsl from the japan society for hematopoietic cell transplantation (jshct) registry. methods: data from the jshct registry were retrospectively analyzed. patients with pcnsl who received first hdt/asct between and were evaluated. distribution differences of clinical characteristics between groups were analyzed with fisher´s exact or mann-whitney u tests. overall survival (os) and progression free survival (pfs) were calculated using kaplan-meier method. two-group analysis of the cumulative incidence of relapse was conducted using the grey test. factors were analyzed in univariable analysis, and all factors with p≤. were retained in the multivariable model. all p values were sided, and values were regarded statistically significant if p< . . results: median age was months (range - ) with patients over years of age; males and females. ecog-performance status (ps) at diagnosis was better (ps - ) in patients and poor (ps - ) in patients. serum lactate dehydrogenase (ldh) levels at diagnosis were elevated in patients. karnofsky ps and cerebrospinal fluid (csf) protein concentration at diagnosis were not collected in the registry. patients were in complete remission (cr), patients were in partial response (pr), and patients were stable disease (sd) or progressive disease (pd) at the time of hdt/asct. after hdt/asct, additional patients achieved cr. with median follow-up period of months, the -year os and pfs were . % and . %, respectively. the was no significant difference in os and pfs between upfront and salvage hdt/asct. since thiotepa, a key agent in hdt/asct for pcnsl, has been unavailable after the discontinuation in japan, the hdt regimens used were not uniform. thiotepa-containing hdt was received by out of patients before , but by out of patients after . thiotepa-containing hdt showed improved pfs (p=. ), lower relapse (p=. ) and a trend toward a survival benefit. in the multivariate analysis, non-complete remission at hdt/asct was an independent predictor for os (hr= . , %ci: . - . , p=. ) and thiotepacontaining hdt remained significant for pfs (hr= . , %ci: . - . , p=. ). [[p image] . os(a),pfs(b) in all patients (n= ) and cumulative incidence of relapse in cr patients (c; n= )] conclusions: our results confirm the activity of thiotepacontaining regimen for hdt/asct in pcnsl patients. currently a pharmaceutical company re-develops thiotepa for new approval of hdt/asct in pediatric solid cancer and adult lymphoma in japan (japiccti- ). further evaluation with the thiotepa by prospective clinical trials is warranted. disclosure background: t-cell non-hodgkin lymphomas (t-nhl) are rare diseases and they are associated with worse prognosis when compared to their b-cell counterparts. allogeneic stem cell transplantation (allo-sct) may have a curative potential for these patients due to the graft versus lymphoma effect. however, data is limited on the efficacy of allo-sct for these diseases. methods: we identified patients ( % females; median age: y; range, - ) with t-nhl that underwent allo-sct at university hospital eppendorf between and . twenty-one patients (underwent allo-sct from a matched sibling donor (msd) and ( %) from a matched unrelated donor (mud). sixteen patients had ptcl ( %), n= ( %) anaplastic large-cell lymphoma (alcl), n= ( %) angioimmunoblastic large cell lymphoma, n= ( %) adult t-cell leukemia/lymphoma, n= ( %) hepatosplenic gamma/delta t-cell lymphoma, n= ( %) enteropathy associated t-cell lymphoma, n= ( %) tcell-prolymphocytic leukemia, and n= ( %) each extranodal t/nk-cell lymphoma, cutaneous t-cell lymphoma as underlying diagnosis. the median ann arbour stage at diagnosis was (range, - ). ten patients ( %) had bone marrow involvement at diagnosis. all patients were heavily pretreated, ( %) patients relapsed post autologous stem cell transplant (apsct) and one patient post allo-sct. fifteen patients ( %) were transplanted in complete remission (cr) ( in st cr, in nd cr), n= ( %) in partial remission (pr), and n= ( %) with advanced disease. most of the patients received myeloablative conditioning ( %). thirty-eight ( %) patients received total body irradiation based regimens and ( %) received chemotherapy based regimens. twenty patients ( %) received anti-t-lymphocyte globulin (atlg neovii), and most patients ( %) received g-csf mobilized peripheral stem cells. results: overall, patients ( %) had neutrophil engraftment (median days: ; range, - ) . at day , the cumulative incidences of grade ii-iv and grade iii-iv acute gvhd were % and %, respectively. after a median follow up of months (range, - ) the cumulative incidences of chronic gvhd was % distributed evenly between limited and extensive. twenty nine patients ( %) achieved cr after allo-sct. median overall survival (os) and disease free (pfs) survival were months and months respectively. the year os and pfs were % and % respectively. fourteen ( %; % ci [ . - . ]) deaths were due to non relapse mortality (nrm) and patients ( %; % ci [ . - . ]) died due to disease progression. patients with a male donor had improved os compared to those with a female donor ( year os male %, female %; p= . ). patient gender, disease subtype, bone marrow involvement, type of allo-sct, donor, patient cmv status, abo incompatibility, disease stage at diagnosis, previous transplant, disease status at transplant, conditioning regimen, atg and stem cell source had no effect on os, pfs, nrm, and post transplant complications. conclusions: acknowledging the retrospective nature, our study shows that allo-sct induces high rates of complete remission, and may have a curative potential even in diseases relapsing post asct. however our findings need to be confirmed in larger prospective studies. disclosure: no funding, no conflict of interest p abstract already published. at-home autologous stem cell transplantation in lymphoma patients: clinical impact of non-g-csf administration post-transplant background: severe neutropenia remains the main cause of morbidity and mortality after autologous stem cell transplantation (asct). g-csf administration after asct is a common practice, performed to reduce the duration of neutropenia and its complications. in a previous work in patients with multiple myeloma managed at home after asct, we did not observe a deleterious clinical impact in those patients that did not receive g-csf post-transplant (martinez-cibrian n. et al, bmt ) . despite the fact that lymphoma patients receive a more intensive conditioning regimen that multiple myeloma patients, we hypothesized that the use of g-csf in lymphoma patients managed at home during the aplasia phase of asct does not provide a significant clinical benefit. methods: lymphoma patients were managed at-home since day + of asct. between february and july , patients received at-home g-csf μg/kg per day since day + until their anc reached x /l (g-csf group) and, since august , patients did not receive g-csf (non-g-csf group). all patients were conditioned with beam and received prophylaxis with a quinolone, fluconazole, aerosolized pentamidine and low-dose acyclovir (hvs+). in all cases we added primary prophylaxis with piperacillin-tazobactam . g/ h i.v., using a portable intermittent infusion pump (iip), from an absolute neutrophil count (anc) < . x /l until the first day of fever or until attaining an anc of x /l. first-line therapy at home of neutropenic fever (nf) was refrigerated meropenem g/ h i.v using a portable iip. fever was an indication of immediate visit to the hospital, and those patients presenting with focal infection or signs of severe sepsis were admitted. other indications for readmission were: willingness of the patient or caregiver; uncontrolled nausea, vomiting or diarrhea and mucositis requiring total parenteral nutrition or i.v. morphics. results: the main characteristics of the patients are shown in table . there were no differences between groups with respect to gender, diagnosis, stage of disease, comorbidity index (hct-ci), source of stem cells (peripheral blood) and cd + cell dose infused. the median (range) age (years) was ( - ) in g-csf group and in non-g-csf group (p= . ). duration of neutropenia less than . x /l was significantly longer in non-g-csf group, with a median of days (range - ), compared with (range - ) in g-csf group (p < . ). severe neutropenia, less than . x /l, was also longer in the non-g-csf group ( days ( - ) vs. ( - ); p= . ). no differences were observed in the time to platelet engraftment. g-csf post-transplant avoidance did not influence the incidence of neutropenic fever, the first day and duration of fever, the incidence and severity of oral mucositis, bacterial infections documented and number of readmissions. the median duration of the whole procedure at-home was day shorter in the g-csf group ( vs. days; p= . ). conclusions: the policy of not administering g-csf post-asct in our home-based program for lymphoma patients, that include intensive bacterial prophylaxis, did not have a deleterious impact on the main results reviewed, which suggests that elimination of its use can be achieved. disclosure the aim of this study was to analyze the spanish experience with patients diagnosed of nhl who received haplosct with pt-cy. methods: sixty patients who received haplosct with pt-cy in spanish centers from to were analyzed. patients were followed-up until . gvhd prophylaxis consisted in cyclophosphamide mg/kg/d on days + and + , and mmf and a calcineurin inhibitor from day + . results: patients' characteristics are summarized on table . median age of patients was , % male, and diagnosed from t cell lymphoma ( %). most of them didn´t achieve complete response prior to transplant ( %), but only % with active disease. up to % of patients had received previous transplant, from which % was an allogeneic transplantation. source of stem cells was mainly peripheral blood ( %), and reduced intensity conditioning was the preferred ( %) regimen. donors were % siblings ( ), % offspring ( ), and % parents ( ). median neutrophil and platelet engraftment was ( - ) and ( - ) days, respectively. acute gvhd grade ii-iv rate was %, with only patients developing grade iii-iv ( %). chronic gvhd rate was %, and only in ( %) was extensive. median follow-up was months. the -year overall survival and event free survival was % and %, respectively. the -year cumulative incidence of relapse was % and -year cumulative incidence of nrm was %. conclusions: relapsed/refractory nhl are aggressive entities with a fatal course in a short period of time. haplosct with pt-cy permit a new treatment option among these patients, with acceptable outcomes. more studies are needed with a larger cohort of patients and longer follow-up to confirm these results. disclosure: nothing to disclose. higher suv at pre-transplant and day posttransplant pet scan both independently predict inferior survival in patients with diffuse large b cell lymphoma background: autologous stem cell transplant (auto-hct) can cure some patients with relapsed diffuse large b-cell lymphoma (dlbcl) but relapse occurs in about % of patients. while our center and others utilize routine surveillance imaging post-transplant, the utility in this setting is unclear. imaging is costly and exposes patients to radiation. novel interventions are now available for patients relapsing after auto-hct making early disease recognition crucial to intervene prior to clinical progression. hence, we studied impact of post-auto-hct surveillance ( )f-fdg-pet ct at day on transplant outcomes. methods: we analyzed a cohort of consecutive auto-hct recipients with relapsed/refractory dlbcl who then underwent pre-transplant pet/ct and surveillance pet ct at day (interquartile range (iqr): - days) post-hct at the university of minnesota medical center. univariate analysis was performed to analyze pet parameters including deauville score (d), standardized uptake values (suv), total lesion glycolysis (tlg) and total metabolic tumor volume (tmtv) as predictors of relapse and survival after auto-hct. in addition, we assessed outcomes of patients with clinically versus radiographically detected relapsed dlbcl after auto-hct. other pre-hct factors analyzed included age, gender, conditioning regimen, performance status, consolidation radiation therapy, tmtv, suv, tlg. results: five-year cumulative incidence of relapse after auto-hct was % ( %ci to ) and overall survival (os) was % ( % ci to ). twelve ( %) relapsed prior to day . d-score for patients with d pet/ct were d ( %), d ( %), d ( %), d ( %), d ( %) with median survival in years for d , d , d and d of . , . , . , and . , respectively. mean suv varied from . (d ) to . (d ). suv was predictive of relapse and os. risk of relapse increased with doubling of suv; -fold higher suv increased hr by . ( %ci . - . ; p= . ). mortality increased with doubling of suv in both pre-hct ( -fold increase in suv associated with hr . [ % ci . to . ]; p= . ) as well as post-hct pet (hr . [ % ci . to . ]; p= ) irrespective of the bulk of tumor. in addition, risk of death was times higher in d patients relative to d (hr . [ % ci = . to . ]; p≤ . ). patients with d (n= ) had higher tmtv ( cm ) compared to d (n= , tmtv . cm ). the hazard ratio for death following relapse was -fold higher (hr . [ % ci . to . ]; p= . ) if relapse was detected clinically versus only radiographically over a median follow-up time period of . years. other pretransplant patient and disease characteristics did not significantly influenced the outcomes. conclusions: in patients with r/r dlbcl undergoing auto-hct, surveillance pet/ct at day identified patients with poor survival~ year. higher suv in both pre-transplant as well as post-hct pet was predictive of increased mortality. these patients may benefit from novel treatments. [ there are concerns about the risks of nivolumab treatment before and after allo-hsct, due to the risk of heavy gvhd, thus the place of immune checkpoints inhibitors is not yet defined. this report include analysis of our center experience of nivolumab treatment in patients with r/r hl before and after allohsct. methods: we retrospectively evaluated the results of allohsct in patients with r/r chl who had undergone transplant from to . the analysis included patients received the flube conditioning and ptcy gvhd prophylaxis. in group a patients (n= ) received bridge therapy with nivоlumab and in group b patients (n= ) received bridge therapy with brentuximab vedotin or chemotherapy-based bridges. time from the last nivolumab administration to allohsct was at least months. results: at the time of analysis, median follow-up was ( - ) months for group a, and ( - ) months for group b. there was no difference in two-year os (p= , ) with significantly better efs (p= , ) for group a versus group b: % and % vs , % and % respectively. relapse incidence was % for group a versus , % in group b (p= , ). cumulative incidence of non-relapse mortality at years was , % and , % in group a and group b, respectively (p= , ). there was no difference in grade ii-iv ( % vs %, p= . ) and grade iii-iv ( % vs %, p= . ) agvhd, as well as extensive chronic gvhd ( % vs %, p= , ) in groups a and b, respectively. ten patients with relapse after allohsct were treated with different doses ( , - mg/kg) of nivolumab in cic center. at the median follow up of mo ( , - ) all patients remain alive. the objective response to therapy was assessed in patients noted in all patients ( %), disregard the dose of the nivolumab: cr in %, and pr in %. the response was lost in four patients, which required nivolumab retreatment. none of the patients developed gvhd after nivolumab administration. in this analysis, there was also no correlation between dose of nivolumab and incidence and severity of adverse events. conclusions: allohsct in combination with immune checkpoints inhibitors is a good option for patients with r/r chl. consideration for immune-mediated toxicities and the potential for increased graft-versus-host disease remain important. early data suggest that nivolumab may be an efficient therapy in patients with r/r chl relapse after allo-hsct. further research needed. disclosure: the authors declare no conflicts of interest. background: transformation to diffuse large b-cell lymphoma (dlbcl) is considered to be one of the most unfavourable events of lymphoma natural history with poorer outcome as compared to de novo dlbcl (alonso-Álvarez et al, bjh ). in patients suitable for salvage therapy, hematopoietic stem-cell transplantation (sct) could be an option, although its role is not well stablished. we analyse indication and outcome after transplant in transformed dlbcl at a single reference transplant unit. methods: out of total of transplants performed at our unit between and - autologous and allogeneic- were dlbcl transformed from an indolent nhl. of them, received an autologous sct (asct) and an allogeneic sct (allo-sct). results: median age was years old (range - ) and (range - ) for patients receiving asct and allo-sct, respectively. all asct received beam as a conditioning regimen and most of the patients in the allo-sct group received a fludarabine/melphalan combination ( %). gvhd prophylaxis consisted on tacrolimus/sirolimus combination in % and calcioneurin plus methotrexate in %. regarding transplant disease status, ( %) of the asct patients were transplanted in complete response (cr). in the allo-sct group, ( %) patients had received three or more treatment lines before transplant and patients ( %) had received a previous asct, being ( %) in cr, in partial response (pr) and in progressive disease. transplant related mortality (trm) was . % in the asct and % in the allo-sct group. overall survival (os) and progression-free-survival (pfs) at months were % (os), % (pfs) for patients receiving asct and % (os) and % (pfs) for allo-sct. with a median follow up of months for patients receiving an asct, ( %) remain in cr. in the allo-sct group median follow up is months for the whole group and months for alive patients; patients are alive and disease free and have died, due to trm ( %). regarding progression, ( %) have progressed after autologous transplant and after allo-sct. conclusions: indication for hematopoietic sct in transformed dlbcl is stablished in few patients. only % of the patients in our transplant unit receive a transplant due to transformed lymphoma, corresponding to a . % of autologous activity and . % of allogeneic activity. according to our results transplant should be considered a curative option. most of our patients were transplanted in cr, so new agents trying to reach best response before transplant should be considered. [[p image] . eva konirova , antonin vitek , marta krejci , edgar faber , katerina steinerova , david belada , jan novak , juraj duras , petr sedlacek , veronika valkova , andrea janikova , ludek raida , pavel jindra , pavel zak , tomas kozak , marie trnkova , michal karas , marek trneny management. however, differences in patient's characteristics as well as frequency of hsct indication in different lymphoma subtypes have been observed in the last decade. the aim of this study was retrospective analysis of hsct for lymphomas in czech republic. methods: data of adult patients transplanted between years - were retrospectively analyzed using ebmt database. results: between and , autologous hsct (asct) were performed in patients ( men, %) with different lymphoma subtypes. the median age was years (range - ). out of these, ( %) were patients with non-hodgkin lymphoma (nhl), ( %) with hodgkin lymphoma (hl). the nhl group comprised of diffuse large b-cell lymphoma (dlbcl, %), follicular lymphoma (fl, %), mantle cell lymphoma (mcl, %) and t-nhl ( %). the frequency of asct in lymphomas increased from to and has been constant since ( - transplants per year). differences in frequency of asct were observed among lymphoma subtypes -decreasing numbers of dlbcl and fl and increasing numbers of t-nhl and mcl, with asct as part of the induction therapy. between and a total of allogeneic hsct (allosct) were performed in patients ( men, %). median age was years (range - ). out of these ( %) were patients with nhl, ( %) hl. the most common nhl subtypes were fl ( %), mcl ( %), t-nhl ( %) and dlbcl ( %). in the last years the number of allosct for lymphoma is fluctuating around per year. the median age at asct was significantly higher in the years - vs - [ . ( . - . ) vs. . ( . - . ), p < . , fig ] , while the increase at allosct [ . ( . - . ) vs . ( . - . )] did not reach statistical significance (p= . ). with median follow up for allosct, y probability os for patient transplanted in the later period - was in relapsed dlbcl . %, in fl . %, in hl . % and in mcl . %, y os for asct as part of first line therapy in the same period was in mcl . % and in t-nhl . %. os was significantly better in all patients who underwent asct in the years - vs - ( .% vs. . %, p < . ) and there was a trend towards better os in patients after allosct (with . % vs . %, p= . ) (fig ) . conclusions: hsct remains important treatment modality for lymphomas in the era of targeted antibody and molecular therapy and we can transplant older patients due to better supportive treatment. acknowledgment: progress q - uk from the czech ministry of education youth and sports disclosure: nothing to declare background: disease chemosensitivity to salvage treatment has been proven to be a major predictive factor for a favorable outcome after autologous stem cell transplantation (asct) for patients with refractory lymphomas. therefore the importance of effective and safe salvageregimens is indisputable. methods: we retrospectively compared the outcomes in terms of safety and efficacy, in (hl: , nhl: ) patients, with a median age of . ( - ) years, who received as st salvage either dicep [cyclophoshamide ( mg/m ), etoposide ( mg/m ), cisplatin ( mg/ m ), days - , (n= )] or the widely used regimen eshap (n= ). rituximab was additionally given to all cd- positive lymphoma patients. the statistical analysis based on the independent t-test, kaplan meir method and logrank test. results: the reason for salvage treatment was primary induction failure (pif, n= ), early relapse (< months post induction-remission therapy n= ) and late relapsed disease (n= ). more specifically, / patients ( %) in the dicep-group, and / patients ( %) in the eshapgroup were assessed with pif or early relapsed disease, however this difference was not statistically significant. both regimens were well tolerated and no major organ toxicities were noticed. eleven patients ( %) from the dicep-group, while only ( %) from the eshap-group developed febrile infections. all patients were successfully managed with the appropriate treatment and only one, from the eshap-group, required for short period admission to the intensive care unit. after cycle of dicep and cycles of eshap the disease response was re-assessed by pet/ct scan. the overall response rate (> % tumor reduction) was significantly superior for the dicep-regimen, reaching % ( / patients) vs. % ( / patients) for eshapregimen (p= , ). eleven patients ( %) from the dicep-group and ( %) from the eshap-group achieved complete metabolic remission according to pet/ ct criteria (p=ns). the median hospitalization period was ( - ) days for the dicep-group compared to ( - ) days for the eshap-group. however, for the eshapgroup, an additional median of ( - ) hospitalization days were required, since of the non-responders patients received a nd salvage before asct. the mobilization and stem cell collection was successful for both groups, though significant higher number of cd + cells were collected in the dicep-group ( . x /kg vs. . x /kg, p= , ). all but two patients (due to refractory disease) underwent asct. noticeably, the median period from st salvage treatment to asct was significantly shorter for the dicepgroup ( vs. days, p= , ), apparently because non-responders patients from eshap-group treated with a nd salvage. the -years overall and progression free survival were similar for dicep-and eshap-groups ( % vs. % and % vs % respectively). two heavily pretreated patients from the eshap-group developed secondary myelodysplastic syndrome post asct conclusions: in our series of patients both regimens proved to be safe. interestingly, despite the fact that more patients in dicep-group had poor risk disease the dicepregiment was significantly more effective, resulting thus in an earlier asct, less exposure to chemotherapeutic agents, that might led in less long-term toxicity. nevertheless, prospective trials with large series of patients are needed to define the role of dicep in the salvage treatment setting. disclosure: no conflict of interest background: although autologous hematopoietic stem cell transplantation (auto-hsct) is one of the best curative strategies for patients with chemosensitive t-cell lymphoma, major limitation remains a tumor contaminated graft-related relapse or residual disease after chemotherapy. several purging methods were introduced in auto-hsct for these limitations, however there are few studies of ex vivo purging of the autograft in lymphomas, especially t-cell lymphoma. therefore, we retrospectively analyzed consecutive t-cell lymphoma patients receiving auto-hsct with/without ex vivo purging. methods: among them, patients underwent autograft manipulation with ex vivo purging by cd + cells selection using a clinimacs device. results: with median follow-up duration of months (range, - months), -year overall survival (os; . % vs. . %, p= . ) and -year progression-free survival (pfs; . % vs. . %, p= . ) in a purged and unpurged group, respectively. transplant-related mortality was observed in both groups ( patients of a purged group and patient of an unpurged group). neutrophil ( vs. days, p= . ) and platelet ( vs. days, p= . ) recovery were similar in both group and there was no engraftment failure. on subgroup analysis according to upfront and salvage auto-hsct, while survival outcomes were improved by stem cell purging in the upfront auto-hsct (os with p= . and pfs with p= . ), there were no different survival outcomes in salvage auto-hsct. the unmanageable late-infectious complications were few in both groups except for predominantly cytomegalovirus reactivation in a purged group ( vs. patient). conclusions: although cohort was a small number, ex vivo graft-purging method was feasible and safe in t-cell lymphomas. and this purging strategy observed the more favorable survival outcomes in the upfront auto-hsct than salvage setting. therefore, further randomized studies are needed to determine the firm efficacy of cd + purification with the large number of patients in auto-hsct for t cell-lymphomas. disclosure: nothing to declare nivolumab-based regimens in relapsed or refractory non hodgkin lymphomas: the role of hematopoietic stem cells transplantation methods: we analyzed data of patients with r/r nhl, among them n with diffuse large b-cell lymphoma (dlbcl), n with primary mediastinal b-cell lymphoma (pmbcl), n with gray zone lymphoma (gzl) and n with gamma-delta peripheral t-cell lymphoma (ptcl), who received nivolumab-based regimens. the median age was years (range, - years). most of the patients n ( %) had a primary chemoresistant disease, the rest patients n ( %) had a relapse. the median of lines of prior therapy was lines (range, - lines). all sixteen patients with dlbcl and pmbcl received - cycles of nivolumab in combination with bendamustine, gemcitabine and rituximab (begern). the patient with gzl received cycles of nivolumab in combination with brentuximab vedotin and epoch. and the patient with ptcl received cycles of nivolumab monotherapy. results: at median follow up months ( - ) objective response (or) after nivolumab-based regimens was noted in n ( %) patients, complete response (cr) and partial response (pr) in n ( %) and n ( %) patients, respectively. cr observed in n patients with dlbcl, n with pmbcl, n with gzl, n with ptcl. and pr observed in patient with dlbcl. two responding patients with dlbcl underwent auto-hsct. and four responding patients (n dlbcl, n pmbcl, n gzl, n ptcl) received allogeneic hematopoietic stem cells transplantation (allo-hsct). the median duration of response for all n patients with or was (range: - +) months. among n patients who achieved or without hsct, only n remain in cr. two patients who received auto-hsct had a relapse. one patient with dlbcl improved the response after allo-hsct from pr to cr, and all four patients with allo-hsct remain in cr. the probabilities of -year os and pfs rates were % and %, respectively. conclusions: nivolumab-based regimens can lead to an objective response in % patients with r/r nhl. however, the durability of response to therapy is not long. nivolumab-based regimens can be used as bridge to allo-hsct disclosure: there are no conflicts of interest to disclose background: patients with aggressive non-hodgkin lymphoma (nhl) who relapse after autologous stem cell transplantation have a dismal outcome and could benefit from radiotherapy, allogeneic stem cell transplantation or experimental treatments. systemic inflammatory parameters at diagnosis have demonstrated to be useful to predict lymphoma evolution. methods: we conducted a retrospective review of patients with aggressive nhl who underwent autologous stem cell transplantation (astc) to evaluate the relationship between ldh, β -microglobulin, inflammatory parameters (lymphocyte (alc) and monocyte count (amc), ferritin or c-reactive protein) and imaging techniques before and on day + post-astc and progression free survival (pfs), as well as the role of residual disease directed radiotherapy (rt). results: one hundred and sixty patients with aggressive nhl received asct as consolidation treatment in our center between and . the most common diagnosis was diffuse large b-cell lymphoma (dlbcl). one hundred and nine patients received upfront asct for high risk dlbcl (defined as age-adjusted ipi - )(n= ) or for having received two or more lines to obtain first complete remission (n= ), for t-cell lymphoma (n= ) and for mantle cell lymphoma (n= ). the rest was performed in relapsed lymphomas. forty-seven patients ( %) relapsed and pfs was months. pretransplant response was evaluated with ct scan in patients ( of this with partial remission (ct-pr) and patients were evaluated with fdgpet/ct ( were pretransplant positive (pet ); of these, patients maintained positivity at day after astc (pet ). pfs in patients with ct-pr was months, in pet positive ones months and in pet positive ones months. univariate analysis showed pet positivity as the most accurate predictor of relapse (hr , , p= , ) followed by amc at day + (hr , , p= , ), albumin at day + (hr , , p= , ), ldh at day + (hr , , p< , ) and pretransplant alc/amc ratio (hr , p= , ). multivariate analysis only demonstrated an association with pet positivity (hr , ) p< and ldh in day + (hr , ) p= , with pfs. five and ten years overall survival were % and % in pet negative patients vs and % in pet positive ones (p< , ). eight out of patients with pet positivity did not relapse. salvage radiation therapy was used in patients with positive residual mass and of them did not relapse. two patients relapsed: one patient had residual mass and another had remote affectation from primary site and could be considered as progression before day + . conclusions: post asct fdgpet/ ct is superior to conventional ct in predicting outcome in aggressive lymphoma after astc. pre and post asct systemic inflammatory parameters didn't help to improve the relapse risk prediction. addition of consolidative rt after astc has demonstrated improvement in pfs in patients with pet positivity. it would be neccesary to develop randomized trials to assess the role of rt in residual disease in advanced aggressive nhl with insufficient response to systemic treatment with pet response evaluation. disclosure: nothing to declare long term outcome of patients with lymphoid malignancy who underwent high dose chemotherapy followed autologous hematopoietic cell transplantation at a single institution over years joanna romejko-jarosinska , ewa paszkiewicz-kozik , lukasz targonski , lidia popławska , jan walewski background: high dose chemotherapy (hdt) and autologous hematopoietic cell transplantation (auto-hct) is a standard of care for relapsed/refractory lymphoma patients (pts) or it is used as a consolidation for myeloma and high risk lymphoma patients in first line treatment. we retrospectively evaluated long-term outcome including late effects and risk factors in patients with lymphoid malignancy who underwent auto-hct. methods: we collected data from consecutive patients with hodgkin lymphoma (hl) (n= ), aggressive b lymphoma (dlbcl) (n= ), myeloma (n= ), indolent lymphoma (n= ), mantle cell lymphoma (n= ) and peripheral t cell lymphoma (n= ) who underwent auto-hct at our institution between and . at transplant median (range) age was ( - ) years, clinical stage iii/iv was found in of lymphoma pts, complete remission, partial remission and stable/ progressive disease occurred in ( %), ( %), ( %) pts, respectively. beam regimen was used in pts ( %), mel in pts ( %) and other myeloablative regimens in pts ( %). results %) ], respectively. partial remission or stable disease at transplant, clinical stage iii or iv, and age more than , were identified as risk factors associated with inferior os and pfs in univariate and multivariate analysis. histopathologic diagnosis was not a risk factor for os and pfs (p=ns). the outcome of patients who underwent auto-hct between - was inferior to the outcome of patients treated in - or - . - year os was %, %, %(p< . ) and year pfs was %, %, % (p< . ), respectively. we recorded ( %) cases of second primary cancer ( solid tumors and hematologic cancers). acute cardiotoxicity occurred in patients from to years after transplant, and required heart transplant in patients. patients ( %) died. the main causes of death were progressive disease in pts ( %), second primary malignancy in pts ( . %) treatment related mortality was . % ( pts), and mortality within days was ( , %). [[p image] . pfs and os in patients underwent hdt and auto-hct - , - , - conclusions: more than % of patients who underwent hdt and auto-hct had long term survival without progressive disease. older age, non-complete remission at transplant, advanced stage are associated with poor outcome. patients recently transplanted had a better outcome than patients transplanted before . disclosure: nothing to declare outcomes after haploidentical and matched related hsct in lymphoma do not differ significantly: a single center study nadira durakovic , , zinaida perić , , lana desnica , ranka serventi-seiwerth , sandra bašić kinda , ivo radman-livaja , alen ostojić , ante vulić , dražen pulanić , , pavle rončević , zorana grubić , igor aurer , , radovan vrhovac , university of zagreb, school of medicine, internal medicine, zagreb, croatia, uhc zagreb, zagreb, croatia background: allogeneic hsct still offers patients with relapsed/refractory lymphoma the best chance of long-term survival. in most such patients timing of hsct is crucial, therefore a related donor is preferred. we analyzed acute and chronic gvhd incidence, relapse and overall survival, but also time to immunosuppression (is) discontinuation and hematopoietic recovery comparing transplantation using haploidentical (haplo) and matched related donors (mrd) in single center in this indication. methods: in the time period between / and / at uhc zagreb, croatia, mrd and haplo transplantations in lymphoma were done, for hodgkin and for nhl. all patients transplanted from haploidentical donors received ptcy. data were computed using the r package. the probability of gvhd was calculated using the cumulative incidence method and subgroups were compared using the gray test. results: median age was ( - ) years; ( - ) in haplo and ( - ) in mrd group. four patients were in pr and in cr in haplo group, while in mrd group patients were in cr and in pr. in haplo group patients ( %) received bone marrow (bm) and only ( %) peripheral blood stem cells (pbsc). in mrd group all patients received pbsc. all patients in haplo group received nma ("baltimore") conditioning with ptcy while in mrd group patients ( %) received flu-bu atg, and only one received flutbi as conditioning protocol. in haplo group % patients were previously treated with autologus transplantation, % in mrd group. there was no significant difference in time to is discontinuation, and days in haplo and mrd group, respectively. patients after haplo recovered slower, recovering anc after . days ( % ci, . - . ) and . ( % ci, . - ) (p= . ) and recovering platelets after . days ( % ci, . - . ) and . ( % ci, . - . ) (p< . ) in haplo and mrd group. with a median follow up of days, overall survival was % ( % ci, - ) in haplo and % ( % ci, - ) in mrd group. trm was % in haplo and % in mrd group. cumulative incidence of agvhd ii-iv was % ( % ci, - ) and % ( % ci, in haplo and mrd group, respectively (p= . ). cumulative incidence of cgvhd requiring treatment was % ( % ci, and mrd % ( % ci, - ) in haplo and mrd group, respectively (p= . ). all cases of cgvhd developed after dli. cumulative incidence of relapse was % ( % ci, - ) and % ( % ci, - ) for haplo and mrd group, respectively (p= . ). conclusions: we found no significant difference in overall survival, relapse incidence, agvhd and cgvhd incidence between these two groups. hematopoietic recovery was slower after haploidentical transplantation, but it did not influence trm as it was higher after mrd. even though limited in number, this data contribute to the growing body of evidence that use of haploidentical donors, particularly in lymphoma setting, is as worthy as using matched related donors and should be at least second choice in donor selection, and in older patients (with older donors) probably the first one. disclosure: nothing to declare. adjuvant involved field radiotherapy post autologous stem cell transplantation for refractory/relapsed lymphomas results in favorable outcome with low toxicity: a single center experience background: involved field radiotherapy (ifrt) to previous bulky or localized residual disease, is a widely used treatment approach to minimize the risk of relapse post autologous stem cell transplantation (asct). however, the proper time for irradiation treatment remains controversial. adjuvant ifrt (adj-ifrt) in pre-asct period could cause undesirable toxicity which might delays or even cancel the asct resulting in increased risk of relapse, or could affect the marrow environmental and marrow niche resulting thus in impaired engraftment. on the other hand, the ajd-iftr in the early post-asct period, upon marrow recovery, offers a potential advantage by delivering irradiation after sufficient disease response, without affecting the engraftment. in this retrospective study we evaluated the safety and efficacy of the ifrt as adjuvant treatment in patients who had previously treated with asct for relapsed or refractory lymphomas. methods: twenty-three patients (hodgkin= , non-hodgkin= ), aged of ( - ) years, underwent asct, for primary refractory (n= ) or relapsed (n= ) disease. patients who had bulky disease at the time of relapse or those with residual mass post salvage treatment, were considered as candidates for adj-ifrt, early (within - months) after documentation of autologous stem cells engraftment. all patients proceeded to asct with chemosensitive disease after a median of lines of salvage therapy. at the time of asct patients ( %) had residual disease while ( %) evaluated to be in complete remission. the preparative regimens were: single-agent melphalan (n= ), busulfan-etoposide-melphalan (n= ), beam (n= ) and bendamustin-etoposide-cytarabine-melphalan (n= ). filgrastim was given till neutrophills recovery, while prophylaxis against bacteria, fungus, viruses and pcp were administered till the completion of adj-ifrt. results: all patients engrafted promptly and successfully. no patient experienced any severe toxicity or active infection before adj-iftr. though our plan was to proceed with adj-ifrt within months post asct, finally it was delivered after a median of . ( - ) months; the median radiation dose was ( - ) gy. ten patients received radiotherapy in the mediastinum, in the abdomen/pelvis/ inguinal area in the neck, and in the left leg. the adj-ifrt was well tolerated. no patient experienced toxicity grade > and none required hospitalization. currently, after a median follow-up of ( - ) years, / patients are alive and well; the -years overall and progression free survival rates are % and % respectively. four patients died; due to relapsed disease and heavily pretreated patients due to secondary myelodyspalstic syndrome conclusions: in our study, the adj-ifrt in the early post transplant period demonstrated a safe and well-tolerated profile. taking into consideration the poor risk status of our patients (residual disease post salvage regimen or bulky disease at the time of relapse), the promising overall and progression free survival rates suggested that adj-ifrt post asct is also an effective approach. well designed trials are needed to clarify the role and the appropriate time of radiotherapy in the asct setting disclosure: no conflict of interest adverse prognostic impact of pre-transplant neutrophil/ lymphocyte ratio in lymphoproliferative disorders background: brentuximabvedotin(bv) is a chimeric anti cd igg antibody, conjugated to synthetic antitubulinmomomethylauristatin. bv is approved for the treatment of classical hodgkin lymphoma (hl) in relapse either after autologous stem cell transplantation (asct) or after two lines of combination chemotherapy in transplant ineligible patients. the aethera trial revealed increased pfs when bv is used as maintenance therapy for cycles in high risk patients after asct. however, this schedule is associated with a high cost and significant toxicity particularly in term of peripheral neuropathy. our primary objective is to assess the efficacy of cycles brentuximab as consolidation therapy after asct for relapsed/refractory (r/r) hl. secondary objectives include side effects, progression free survival (pfs), and overall survival (os). methods: this is a retrospective single center analysis approved by the irb of the american university of beirut medical center. we included in this study consecutive patients with r/r hl who underwent asct between and , and received bv consolidation post-asct. results: we identified consecutive adult patients with r/r hl treated with bv . mg/kg iv every weeks as consolidation therapy after asct. the indications for bv consolidation was primary refractory disease in patients ( %), early relapse in patients ( %) (after a median time of months; range, - ) andextranodalinvolvement in one patient ( %). the median number of lines of therapy pre-asct was (range, - ). the median time to bv initiation post-asct was days (range, - ). patients received a median of cycles (range, - ) of bv post-asct. after a median follow up of months (range, - ), five ( %) patients relapsed after asct. the median time to relapse was months (range, - ). median pfs and os were not reached. we did not observe any significant toxicities during or after therapy. conclusions: cycles of bv consolidation after asct seem to be safe and effective in preventing relapse, however our findings need to be confirmed with larger prospective studies. chemotherapy or who progress after autohsct is poor. despite introduction of novel agents like brentuximab vedotin (bv) or nivolumab, allohsct appears the most effective treatment option with curative potential. the goal of this study was to evaluate efficacy of allohsct for hl, including patients pre-treated with novel agents. methods: between years - , patients (including males) with hl were treated with allohsct in msc institute of oncology in gliwice, poland. median age was ( - ) years. median lines of preceding chemotherapy was ( - ); ( %) patients had been pre-treated with autohsct, ( %) with radiotherapy, ( %) with bv, ( %) -with nivolumab. disease status at allohsct was as follows: cr- , pr- , nr- . patients were treated with hsct from either hla-matched sibling donor (msd, n= ), unrelated donor (urd, n= ) or haploidentical donor (n= ). conditioning was myeloablative in ( %) cases. peripheral blood was used as a source of stem cells. results: all but one patient engrafted with median time of neutrophil recovery of ( - ) days. the incidence of grade - and grade - acute gvhd was % and %, respectively, while the incidence of chronic gvhd was %. the probabilities of os and pfs at years were % (+/- %) and % (+/- %), respectively. the incidences of progression and transplant-related mortality were % and %, respectively. the y pfs rates were % for msd, % for urd and % for haploidentical donors. in a univariate analysis pfs was affected by recipient gender (female - %, male - %, p= . ) and disease status at allohsct (cr - %, pr - %, nr - %, p= . ). in a multivariate model the disease status other than cr was the only factor associated with increased risk of treatment failure (reverse pfs) -hr= . , %ci . - . , p= . . neither donor type nor conditioning affected long-term outcome. conclusions: results of allohsct for patients with relapsed/refractory hl are determined by disease status at transplantation. efforts should be done to reduce tumour burden before transplantation, optimally to achieve cr. disclosure: nothing to declare background: brentuximab vedotin (bv), nivolumab and pembrolizumab have been assigned to chemorefractory hodgkin lymphoma treatment. impact of these agents on disease-free-survival after autologous stem cell transplantation (asct) remains under investigation. aim of the study is to compare bv-and nivolumab-treated patients with a control group. methods: clinical characteristics and outcomes of chemo refractory hodgkin lymphoma patients who underwent asct during - . results: a total of patients ( men; women, median age years old, - ) were treated with bv: pre-transplant, post-transplant and pre-and posttransplant. pre-transplant bv patients had primary refractory disease or early relapse in the majority ( %). post-transplant treatment occurred in the context of relapsed/refractory disease in patients; ( %) had an allogeneic stem cell transplant. among them, had additional chemotherapy and nivolumab, gaining a complete metabolic response. in the rest of patients, change of treatment due to eventual bv failure occurred. bv was administered as a maintenance treatment in patients. in six of them bv had already been administered pre-transplant as well. out of maintenance treatment patients, relapsed and subsequently received nivolumab. two patients died due to prior chemotherapy complications, whereas are currently on nivolumab treatment. pet-based response was available in patients, having a complete metabolic response (cmr) and a partial metabolic response. stable disease was achieved by ctbased response in the rest patients. no major toxicities were observed. one patient presented with grade asymptomatic hypothyroidism and one with grade anemia attributed to non-inflammatory upper gastrointestinal blood loss. in total, patients received anti-pd treatment, all post bv failure. with a median follow-up of . ( . - . ) months, -year overall survival (os) was . % in patients treated only with bv compared to . % in patients treated with additional anti-pd treatment (p= . , figure) . median os for patients treated only with bv was . months, whereas median os has not been reached for patients that received anti-pd treatment. conclusions: bv pre or post-transplant and anti-pd treatment post-transplant after bv failure have outstanding results in chemo refractory lymphoma patients. treatment sequence in allogeneic transplantation eligible patients remains to be further studied. disclosure: nothing to declare background: allogeneic hematopoietic cell transplantation (allo-hct) with reduced-intensity conditioning (ric) has been used in heavily pretreated lymphoma patients with the promise of decreased treatment-related mortality. despite improvements in outcomes of patients with lymphoid neoplasms, several new agents emerge as potential therapies. therefore, we aimed to describe our long-term experience in patients with hodgkin (hl), non-hodgkin lymphomas (nhl) and chronic lymphocytic leukemia (cll) post allo-hct. methods: in this retrospective study, we enrolled consecutive patients who underwent allo-hct for lymphoid neoplasms in our institution ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) . results: in total, patients (male:female= : ) aged ( - ) years, underwent allo-hct for hl (n= ), nhl (n= ) and cll (n= ). the majority of patients were diagnosed at stage iv ( %); % had bone marrow involvement and % had undergone autologous hct. most patients were heavily pretreated (median lines= , range - ), of them had received more than treatment lines and at the time of transplantation only had complete response, while had partial response and were refractory. according to disease-risk index (dri), patients were stratified at low (n= , . %), intermediate (n= , . %), high (n= , . %) or very high (n= , . %) category. among patients with hodgkin lymphoma, brentuximab vedotin was administrated in and of them were effectively bridged to allo-hct. all patients received ric, mainly fludarabine ( mg/ m )-cyclophosphamide ( g/ m ) in cll and nhl and thiotepa ( mg/kg)-fludarabine ( mg/m )-cyclophosphamide ( mg/kg) in hl from matched sibling (n= ), matched (n= ) or mismatched unrelated (n= ) donors. graft-versus-host disease (gvhd) prophylaxis consisted of cyclosporine or tacrolimus and mycophenolate mofetil or short-term methotrexate and additional low-dose antithymocyte globulin ( mg/kg) in unrelated donors. peripheral blood was the main cell source ( / ) and median number of cd + cells infused was . x /kg ( . - . ) . two patients succumbed to advanced underlying disease before engraftment; while the other engrafted successfully. median time until neutrophil and platelet engraftment was and days respectively. eighteen patients ( . %) developed acute gvhd (grade iii-iv, n= ), steroid sensitive in ( . %) and relapsed. one-year cumulative incidence (ci) of extensive chronic gvhd was . %, and patients required more than one additional line of immunosuppression (range - ). ten patients presented cmv reactivation successfully treated with antiviral medication and patient died from hsv encephalitis. with a median follow of years ( - years), -year os was . %, -year non-relapse mortality ci . % and year dfs %. there was no difference in survival according to original disease ( -year os for nhl= . %, hl= . %, cll= %%, p= . ). multivariate analysis revealed high and very high dri as the single predicting factor of os (hr . , ci . - . , p= . ), when assessing impact of disease, dri, prior treatment lines, gender and bone marrow infiltration at diagnosis. conclusions: our data suggest that ric allo-hct provides encouraging survival rates, potentially offering the chance of cure, with acceptable long-term mortality in selected high-risk patients with lymphoid neoplasms. dri that is mainly associated with disease stage at transplant independently affects survival. therefore, continued efforts are necessary for clinical application of novel agents aiming to lower disease stage pre-transplant. disclosure: nothing to declare results: six pts were identified, with a median age of years at diagnosis: five with hl nodular sclerosis and with lymphocyte depletion. the median number of therapeutic lines prior to allo-hsct was [ - ]; four pts were previously treated with brentuximaband two pts had been submitted to high dose chemotherapy with autologous bone marrow support. at the time of allo-hsct, pts had progressive disease (dp), was in partial response and in complete response (cr). five allo-hsct were performed with a related donor, of wich were haploidentical ( parents, sibling and descendant) and with an unrelated donor ( / ). prophylaxis for gvhd was performed with tacrolimus and mycophenolate mofetil (with post-transplant cyclophosphamide in haploidentical allo-hsct). on day + evaluations, pts had a cr and patient (pt) had dp. the median time to relapse after allo-hsct was of months. at the time of initiation of nivolumab, pts were under steroid therapy for disease control, without other immunosuppressive therapy. the median time between allo-hsct and the beginning of nivolumab was months. the initial dose was mg / kg (associated with corticosteroid therapy), escalated up to mg/kg biweekly, according to patient's tolerance. after the start of nivolumab, patients, with previous gvhd manifestations, presented a worsening of the cutaneous gvhd, which required an escalation of immunosuppressive therapy. as toxicity, pt had a grade pneumonitis, pt had a grade encephalitis/hypophysitis, pt had a grade pancreatitis, pts had headache (grade and ), pts had a grade - cutaneous reation. with a median follow-up of months since nivolumab treatment, the overall response rate was %: pt obtained cr and pts partial remission. nevertheless, there were deaths after the onset of nivolumab: pt at monts with dp and another one due to acute myocardial infarction at months. at the time of analysis, pts maintained response under nivolumab treatment (median cycles ) and pt had therapy suspended because of toxicity. conclusions: these results demonstrate the high probability of achieving response with nivolumab treatment in patients with rr-hl relapsing after allo-hsct, but adverse events of grade were frequent in this small group, and the treatment toxicity was significant. disclosure: nothing to declare background: intravascular large b-cell lymphoma (ivlbcl) is a rare form of large b-cell lymphoma with pathological findings of intravascular proliferation and/or sinusoidal involvement of lymphoma cells. according to their geographic distribution, ivlbcl could be dichotomized into asian and western variants. compared with the western variant, where skin involvement was common, the asian variant was reported to involve more frequently the liver, spleen and bone marrow, and hemophagocytic lymphohistiocytosis is more common in asian variant. diagnosis of ivlbl is still difficult because of the lack of overt lymphadenopathy and peripheral blood involvement. thus, timely diagnosis and immediate treatment remain as a challenge to improve outcomes for patients with the asian variant. therefore, we analyzed the clinical features and treatment outcomes of patients with the asian variant of ivlbcl. methods: we analyzed patients who were diagnosed with ivlbcl between and . all patients were treated with rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisone (r-chop). results: forty-six patients were diagnosed with ivlbcl, and their median age at diagnosis was years (range: - years). male patients predominated (n= , %), and b symptoms were present in patients ( %). hepatomegaly and/or splenomegaly were observed in patients ( %), whereas lymphadenopathy was less common (n = , %). bone marrow and liver were the most commonly involved extranodal organs ( %, and %, respectively) and were the most common sites of biopsy for diagnosis in this study. all patients received r-chop as a first-line treatment after diagnosis with a median number of six cycles (range one to eight). at the end of treatment, patients achieved a complete response (cr), whereas eight patients showed progression. six patients died after the first or second cycle of r-chop, and the causes of death were treatment-related adverse events including cytopenia, infectious complications, and pulmonary hemorrhage. upfront asct was done for two patients including one patient with cns involvement at diagnosis, and these patients were still alive at the time of analysis without evidence of relapse. on the other hand, the outcome of six patients undergoing salvage asct after relapse was poor; thus, only one patient was alive. likewise, patients with disease progression at the end of treatment with r-chop showed dismal prognoses even after salvage chemotherapy except for one. at a median follow-up of . months ( % confidence interval, ci . - . ), the median overall survival was . months ( % ci . - . ). the treatment outcome of patients with the asian variant of ivlbcl is still not satisfactory. although upfront autologous stem cell transplantation might be effective for selected patients at high-risk of relapse, its role is still not clear, either. thus, further study should be warranted to develop more effective strategies for diagnosis and treatment. clinical trial registry: not applicable disclosure: nothing to disclare background: peripheral t-cell lymphomas (ptcls) are about % of non-hodgkin´s lymphomas usually with an aggressive clinical course and unfavorable prognosis.given their heterogeneity, consensus on the best first-line treatment and the role of autologous/allogeneic (asct/allosct) stem cell transplantation as consolidation is controversial. methods: we evaluated the overall survival (os), progression-free survival (pfs) and toxicities of a cohort of patients with ptcls submitted to asct/allosct intensification at our institution between january and july . os was calculated from the date of diagnosis until death. pfs was measured from transplant until relapse, progressive disease or last follow-up. os and pfs rates were estimated using the kaplan-meier method and compared with the log-rank test. results: twenty-six patients were identified, female ( %), median age was years (range: to ). ninetytwo percent of patients presented with advanced-stage disease at diagnosis (ann arbor stage iii or iv), % with b symptoms. according to the who classification, histologic ptcl subtypes included angioimmunoblastic tcell lymphoma (n = ); ptcl not otherwise specified (n = ); anaplastic large cell lymphoma, alk-negative (n = ); anaplastic large cell lymphoma, alk-positive (n = ); nodal peripheral t-cell lymphoma with tfh phenotype (n = ). extranodal nk/t-cell lymphoma, nasal type and primary cutaneous subtypes were excluded. the ageadjusted ipi (aaipi) was low/intermediate low in patients ( %) and intermediate high/high in patients ( %). twenty-seven transplants were performed ( asct, allosct); were consolidation in st response ( asct and allosct) with in complete remission (cr) and in partial remission (pr). nine transplants were performed as consolidation of nd response ( asct and allosct) with in cr and pr. in patient allosct was performed after asct, due to early relapse (< months). beam regimen was used in asct as conditioning and flumel in allosct. all patients engrafted, the median time to leukocyte recovery > , /μl was days (range, to ). four of the pts ( %), submitted to allosct had chronic graft versus host disease which was the most relevant complication of this analysis. considering the whole cohort, the median follow-up was . months (range, to ). the estimated -year os and pfs were % and %, respectively. seven patients relapsed ( early) all after asct, there were no relapses after allosct, however, the results were not statistically significant between the allosct and asct groups; the -year os rates were % and % (p = , ) and the year pfs rates were % and % (p = , ) respectively. for the all patients treatment-related mortality (trm) was , %; patients died, with progressive disease (asct) and for hepatic toxicity (allosct) before d+ . conclusions: the results of this retrospective study, taking into account the adverse risk profile of the population, suggest that autologous/allogeneic stem cell transplantation as an effective and safe option for the consolidation of patients with ptcls. these results need to be validated in prospective studies, including a larger number of patients. disclosure background: autologous stem cell transplantation is used as consolidation therapy in relapsed lymphoma patients. however, outcome of lymphoma patients relapsing after autologous stem cell transplantation is poor and allogeneic stem cell transplantation which can be curative is used in the transplant eligible patients in this setting. besides, allogeneic stem cell transplantation can be an option before autologous stem cell transplantation in some high risk patients. in this study, we aimed to compare the survival rates of lymphoma patients older than years of age and patients aged - who had undergone allogeneic transplantation in our center. methods: we collected the data of lymphoma patients older than years of age who had undergone allogeneic transplantation in our center and analyzed the results by grouping them into , namely the ones between - years of age and the ones over years of age. [[p image] . figure results: there were patients over the age of who had undergone allogeneic stem cell trasplantation with the diagnosis of lmphoma between and . of these patients were over years of age. patients had non-hodgkin lymphoma and patients had hodgkin lymphoma. the characteristics of the patients are summarized in table . patients' comorbidity indexes were calculated with augmented hct-ci which includes patients' pretransplant ferritin, albümin and thrombocyte counts as a variable. no difference could be found between groups regarding neutrophil and platelet engraftment times and comorbidity indexes. however, acute graft versus rate and documented bacterial infection rate during the hospitalization period were higher in the - years age group (p= , ). day mortality rate and non-relapse mortality rate were not different between groups. more importantly, progression free survival(pfs) and overall survival(os) of patients in the - years age group and over years of age group were not different (p= , ) (figure ) conclusions: in the present study, although the number of patients is low, we showed that lymphoma patients over years of age have similar outcomes and transplant related toxicity as the patients between to years of age. pfs and os were very close in this study. we think that this may be due to low relapse rate in the patients and high mortality rate in relapsing patients. in conclusion, allogeneic stem cell transplantation which has a curative potential may be employed in transplant eligible elderly lymphoma patients disclosure: nothing to declare background: follicular lymphoma (fl) histologic transformation consist on the development of an aggressive lymphoma, usually a diffuse large b cell lymphoma (dlbcl). histological transformation has been considered to have poor prognosis. in pre-rituximab era median os ranged between and years, however, in recent series of patients treated with chemotherapy plus rituximab, the outcome of transformed fl has improved, especially in those that receive autologous stem cell transplantation (asct), who reach -year os up to % in some series. methods: we have retrospectively studied consecutive patients undergoing asct for transformed fl between and in a tertiary center in the basque country, spain. patients were considered to have a transformed fl if they were diagnosed of a dlbcl and they have previous history of fl or histological evidence of a fl in another location. these patients were compared to a retrospective cohort of dlbcl patients with high ipi or stage that received asct in first remission according to our institution strategy. pfs and os were calculated from the time of the asct. in the case of transformed fl, both relapses of the aggressive or indolent lymphoma were considered. survival analysis was performed with kaplan-meyer estimator results: a total of transformed fl and dlbcl patients were studied, with a median follow up of . and . months respectively. patient characteristics are described in table . -year pfs was % in transformed fl and % in dlbcl, and -year os was % and %, respectively (picture ). there were no significant differences in pfs or os between this two groups (p = . ). in both groups all relapses occurred in the first three years after asct. among the patients with transformed fl relapses were observed. five of them ( %) were aggressive relapses, while only one patient presented relapse as an indolent lymphoma (fl histological grade a with an aggressive clinical course). [[p image] . image : transformed fl and dlbcl pfs after asct] conclusions: in our experience, asct in transformed fl offers good results, similar to those in dlbcl. fl presents a natural course akin to that of dlbcl, with relapses occurring early and survival reaching a plateau. this data suggests that some patients with transformed fl can be cured after asct. disclosure: nothing to declare. safety and efficacy of intensive preconditioning regimen containing cladribine in autologous peripheral blood stem cell transplantation of refractory and relapsed young highly invasive lymphoma background: autologous peripheral blood stem cell transplantation (apbsct) is one of the main treatments for patients with non-hodgkin's lymphoma (nhl). effective and safe conditioning regimens can improve the cure rate of nhl. beam is the most common pretreatment scheme, but for refractory and relapsed young highly invasive lymphoma, especially for dual-expression dlbcl, pretreatment needs to be strengthened. studies have shown that the cladribine (clad)+gemcitabine (gem)+busulfan (bu) combination provides synergistic cytotoxicity in lymphoma cell lines.we evaluated the the safety and short-term efficacy of intensive preconditioning regimen containing cladribine (clad+gem+bu) for refractory and relapsed young highly invasive lymphoma undergoing apbsct. methods: ten patients with nhl received apbsct. ca)ctx+ ara-c) therapy followed by g-csf was used for pbsc mobilization. sevenr patients received conditioning regimens of beam(beam group): bcnu mg/ m ·d - × d (- d), vp mg/m · q h× d (- d-- d), ara-c mg/m ·q h× d (- d-- d), mel mg/m ·d - × d (- d). three patients received intensive preconditioning regimen containing cladribine (clgb group): clad mg/m ·d - × d (- d-- d), gem mg/m ·d - × d (- d, - d), bu . mg/kg q h× d (- d-- d). follow-up date expires on december , . results: the age of patients in clgb group was , and years, respectively. two patients were diagnosed as diffuse large b-cell lymphoma with double expression and one was diffuse large b-cell lymphoma with two recurrences. the patients of beam group were all high-risk, relapsed and refractory nhl.all patients were successfully engrafted after infusing apbsc. the average lowest leukocyte in clgb group and beam group were ( . ± . ) × /l vs ( . ± . )× /l, respectively. the average lowest leukocyte in clgb group was lower than that in beam group. the average time to anc < . × /l in clgb group and beam group were . d ± . d vs . d± . d. the average time to anc≥ . × /l in clgb group and beam group were . d± . d vs . d± . d; the average time to plt≥ × /l of clgb group was not different to that of beam group ( . d± . d vs . d± . d) the average time of neutropenia wasn't significantly different in two groups ( . d± . d vs . d± . d). the adverse reactions of gastrointestinal tract and oral mucosa were close in tow groups.vod, hemorrhagic cystitis, pretreatment-related interstitial pneumonia, liver and kidney dysfunction were not happened in tow groups. the rate of infectious fever was close in two groups ( / vs / ). the median followup period in beam group was ( ~ ) months. in the beam group, a patient died days after transplantation, because he was diagnosed with recurrent nkt cell lymphoma and intracranial infection caused by severe sinus infection. another case of beam group was diagnosed as double-expressed dlbcl, which relapsed months after transplantation. the remaining patients in ebmt group survived disease-free. the follow-up time of patients in clgb group were months, months and months respectively. all patients survived without disease.however, the follow-up time is short and needs long-term follow-up. conclusions: the treatment of intensive preconditioning regimen containing cladribine (clgb) for refractory and relapsed young highly invasive lymphoma undergoing apbsct is safe. the time of hematopoietic reconstruction is short, and the adverse effects is tolerable for patients with refractory and relapsed young highly invasive lymphoma. the current short-term outcome is good, but the long-term effect need a longer time to follow-up. disclosure: this work was supported by national nature sciences found of china ( ). there is no disclosure of conflict of interest.the all authors name: xiang-li chen, yu-zhu zang, wen-hui zhang, yin zhang, zhong-wen liu, ping-chong lei, jing yang, yu-qing chen, kai sun. background: small part of children with hodgkin disease (hd) demonstrate initial resistance to the standard and even "salvage" chemotherapy and need innovative drugs for the treatment. methods: a -year girl was diagnosed with classical hd (nodular sclerosis)corresponding to stage ii e b (fever > °c) in april .after two cycles of oepa (vincristine, etoposide, prednisone and doxorubicin) and next two cycles copp (cyclophosphamide, vincristine, prednisone and procarbazine) the patient again had progressive disease. as the patient achieved a partial response (pr) after "salvage"therapy with two cycles of igev (ifosfamide, gemcitabine, vinorelbine, and prednisone), she received auto-sct in february (patient status before auto-sct was pr). we used ccnu-containing conditioning regimen cem: lomustine (ccnu) mg/m + etoposide mg/m + melphalan mg/m . at day + after auto-sct, the patient again demonstrated progression of the disease: pet/ct-examination showed mediastinal tumor mass enlargement with increased left lung nodule simultaneously to short metabolic activity. patient was under observation. at day + the disease had relapsed and progressed -examination by pet/ct demonstrated multifocal progressive disease with multiple pulmonary lesions and increased metabolism in comparison with the previous pet/ct scan. in july-october , the patient had salvage chemotherapy with a combination of brentuximab vedotin (bv) (bv dhap (dexamethasone, cytarabine, cisplatin) + bv (without chemotherapy due to suspected invasive mycosis) + bv dhap), however, only partial pet-positive remission was achieved. because of many times relapsed and progressed disease pembrolizumab therapy was started in october : mg / kg every three weeks four cycles totally. toxic effects and serious complications during and after therapy by pembrolizumab were not observed. in february , after pembrolizumab # , the patient showed complete metabolic remission of the disease by control pet-ct. in april , the patient received haplo-sct with post-transplant hd-cyclophosphamide. we used conditioning modes with reduced toxicity (fludarabin mg/m + treosulfan g/m ), high doses of cy ( mg / kg) on days + and + . tacrolimus and mycophenolate mofetil started on day + . mmf was terminated on day , tacrolimus -on day . patient did not have acute and chronic gvhd. results: at the moment the patient is alive and still in pet-negative cr with duration more than mo. conclusions: pembrolizumab has demonstrated high activity against resistant hd even after post-auto-sct progression with good tolerability for the sick child. disclosure: nothing to declare p high dose chemotherapy followed by autolougous peripheral blood stem cell transplantation (asct) in diffuse large b cell lymphoma (dlbcl) median age is , years ( to ) and sex ratio (m/f) . ; ann arbor stage iii-iv: pts. before hdt induction chemotherapy (chop, c h opa) was instituted and associated with rituximab in pts ( , %), pts ( , %) received more than treatment lines and pts ( , %) received complementary radiotherapy. transplant disease status before hdt was complete remission (cr) in pts, partial remission in pts (rp) and disease progression in pts. the delay from diagnosis to hdt is , months ( - ). the hdt protocols used are: tutshka : pts, tutshka+vp : pts, bam (busulfan +cytarabine+melphalan) : pts et beam : pts. all pts received, after thawing, mobilized pbsc obtained by g-csf mobilization ( μg/kg/d, days) alone and froze in liquid nitrogen. the median rate cd + cells infused is , x /kg ( . - . ) . the median follow-up at / / is months . results: the median time to graft (pnc > . x /l) was days ( - ). ten early deaths were observed including infection (trm: , %) and in disease progression at months. after months of hdt pts are assessable including pts in cr ( , %) and pts in pr ( , %). relapse was observed in pts ( . %) and it was earlier relapse in a period of months in pts ( %). deaths were among / pts ( %). persistent cr was achieved in / pts ( , %) including / ( , %) mlcl and / ( , %) others dlbcl. the overall survival (os) and event free survival (efs) at years are respectively % and %. the os and efs are better in patients who received rituximab in initial therapy : os ( % vs %; p< , ) et efs ( % vs %; p< , ). herein, we present one patient with refractory mcl, who were insensitive to chemotherapy and then experienced a dramatic improvement with ibrutinib mono-therapy but later developed ibrutinib resistance,ultimately resulting in the deterioration of disease and death. methods: we give the patient several examinations including ultrasound, bone marrow biopsy, lymph node biopsy, exome sequencing, sanger sequencing, and so on. for the treatment of lymphoma, the patient received chemotherapy, including course of chop(cyclophosphamide . g day , doxorubicin mg day ,vinorelbine mg day , and dexamethasone mg from day to ) and course of r-dhap (rituximab mg day , cytarabine g q day , cisplatin mg day ,dexamethasone mg from day to )in succession.because of the failure to control disease progression, ibrutinib mg qd was used until the patient died. results: the -year-old man initially referred to our hospital for complaints of abdominal pain and distention over months. ultrasound showed splenomegaly and multiple enlarged retroperitoneal lymph nodes.excisional biopsy conducted on the right neck lymph node revealed the presence of malignant cells.immunohistochemically, the neoplastic cells were positive for bcl , bcl ,cd , cd , cd a, cd , ki- ( %), sox , cd (fdc) and cyclin d and negative for cd , cd and cd ; fluorescence in situ hybridization(fish) showed igh/ ccnd ,t( ; ) %.thus a diagnosis of mcl was confirmed. course of therapeutic chemotherapy were applied to the patient but he did not respond well.he suffered recurrent fever, thrombocytopenia, left abdominal pain, splenomegaly and multiple enlarged lymph nodes. then he received ibrutinib mono-therapy, and experienced a dramatic improvement as his body temperature was controlled, his hemogram became normal and his spleen and lymph node tapered.after about months response of ibrutinib, the disease deteriorated rapidly and he died very soon. exome sequencing from the patient peripheral blood at this time detected one missense mutation in exon of tp at nucleotide g>a, resulting in an argnine to histidine change at amino acid (p.arg his). but sanger sequencing of the patient bone marrow ffpe sample at the time of original diagnosis did not detect this mutation. conclusions: thus, our study reported a tp r h mutation mcl patient who developed ibrutinib resistance and progressed aggresively, which may open new insight for future effort for alternative therapeutic strategies in ibrutinib-refractory mcl. disclosure: nothing to declare. minimal residual disease, tolerance, chimerism and immune reconstitution peripheral blood samples were obtained for routine analysis at several time points after hsct. all available blood samples between . and years were used in the analysis. to assess changes in the cd +b and cd +cd +memory b cell counts over time while accounting for the correlation between the repeated measurements of each patient, we used linear mixed-effects models. wilcoxon rank test, kruskal-wallis test and linear regression were used for univariate analysis. results: at one year after hsct, univariate analysis showed that patients transplanted with a cb graft compared to bm and pbsc had a significantly higher absolute number of b cells (median bm= , median cb= , median pbsc= cells/μl, p= . e- ) and memory b cells (median bm= , median cb= , median pbsc= cells/μl, . recipients with age under years had significantly higher absolute numbers of b (median= , median= cells/μl, p= . e- ) and memory b cells (median= , median= cells/μl, p= . e- ) than above years. increase in donor age was associated with a decreasing effect on b cell (r = . , p= . e- ) and memory b cell (r = . , p= . e- ) reconstitution as determined in regression analysis. following univariate analysis, we analysed these factors in a mixed effects model to assess the relation with differences in b cell or memory b cell numbers . - years after hsct. in our analysis we found significant decreasing b cell and memory b cell numbers with increasing donor age corrected for recipient age and source (both p< . ). increasing recipient age also showed a significant decrease in b cell and memory b cell numbers (both p< . ) but there was no significant influence of donor source ( figure ) . conclusions: b cell and memory b cell numbers after hsct are influenced by donor and recipient age but not by donor source. older donors and recipients show a decrease in b cells and memory b cells numbers . - years after hsct. [[p image] . figure . b cell development and donor age. green shows cb, red bm, blue pbsc at mean donor age.] copenhagen university hospital rigshospitalet, copenhagen, denmark, leiden university medical center, leiden, netherlands background: the outcome of allogeneic hsct is challenged by a delayed and long-lasting imbalanced t-cell reconstitution increasing the risk of acute gvhd, infections and disease progression. although the role of differentially and functionally distinct t-cell subsets in the development of complications has been addressed, little is known about the factors controlling their recovery. in this study, we investigated the impact of immuneregulating and homeostatic cytokines on the reconstitution of functionally distinct t-cell subsets and associated clinical outcomes. methods: we included children undergoing allogeneic hsct for all (n= ) or aml (n= ) with a median age of . years (range: . - . ). donors were either mrd (n= ), mud (n= ) or mmud (n= ). bm (n= ) or pb (n= ) were used as stem cell source. conditioning regimens were based on tbi (n= ) or highdose chemotherapy alone (n= ) and included atg in patients. thirty age-matched healthy children were included as controls. cytokines (il- , il- , il- , scf, il- , il- and tnfα) and active atg in plasma were longitudinally measured from before conditioning until months after hsct (n= ) along with an extended phenotyping of t-cell maturation and differentiation by flow cytometry (n= ). results: the homeostatic cytokines il- and il- increased from pre-conditioning to peak - weeks post-hsct and gradually declined thereafter. il- levels were shortly elevated, while il- and scf remained relatively stable, and il- and tnf-α levels were below threshold of detection at all time points. the early rise of il- and il- was strongly associated with the degree of t-cell depletion by atg, while il- also correlated with markers of systemic inflammation. il- and il- levels were significantly higher in children treated with atg (p< . ) and correlated with both longer exposure to atg (p< . ) and increased levels of active atg (day + : il- : r= . , p< . ; il- : r= . , p< . ), indicating that high levels of these cytokines reflected more pronounced t-cell depletion during lymphopenia. higher circulating levels of il- and il- were associated with a slow recovery of cd +, cd + and cd + t-cell counts at day + and + post-hsct (p< . ), while the remaining cytokines did not correlate with immune reconstitution. looking into t-cell subpopulations, increased levels of il- and il- during the first month post-transplant were associated with lower numbers of naïve t cells and correlated with an increased proportion of cd + and cd + effector memory cells ( figure) . no differential effect of cytokines on polarization of cd + t cells into th , th , th cells or treg cells was found. in atg-treated patients, il- and il- levels at day + were significantly lower in patients developing acute gvhd grade ii-iv (p= . and p= . , respectively). in the total cohort, increased il- levels were associated with a reactivation of ebv (p= . ). conclusions: these findings suggest that quantification of il- and il- can be indicative for the degree of t-cell depletion during the first weeks after hsct and predictive of complications. overall, these results indicate that the lymphopenia-induced elevation of il- and il- is a major driver of the initial expansion of donor t-cells. background: mathematical kinetic models were adopted to study immune cell reconstitution after allogeneic hematopoietic stem cell transplantation (allo-hsct). the associations between acute graft-versus-host disease (agvhd), relapse and the immune cell reconstitution kinetic models were explored. methods: from june , to may , , sixty-five patients with hematological malignancies after allo-hsct were recruited. peripheral blood was collected on + day, + day, + day and in + month, + month, + month, + month, + month, + month, + month. lymphocyte subsets were determined by flow cytometry, including in total t lymphocytes (cd + ), helper t cells (cd + cd + ), cytotoxic t cells (cd + cd + ), cd /cd ratio, nature killer (nk) cells (cd -cd + ), nkt cells (cd + cd + ), b lymphocyte (cd + ), naive t cells (cd + hla-dr + ), static t cells (cd + hla-dr -), and regulatory t cells (cd + cd high foxp + ). mathematical kinetic models were calculated for immune cell reconstitution with spss. results: after allo-hsct, a logarithmic curve model was observed for cd + t cells. cubic curve models were observed for cd + cd + t cells, cd + cd high+ foxp + t cell, cd + hla-dr -t cells, cd + cd + nkt cells, cd + b cells. cd + cd + t cells, cd + hla-dr + t cells, and cd -cd + nk cells showed s type curve models. considering t cells were the major mediators for agvhd and graft-versusleukemia effect after allo-hsct. with established immune cell kinetic models, we found that different curve models were observed between patients with and without agvhd after allo-hsct. although the kinetic models were almost the same for leukemia-free and relapsed patients in the first months after allo-hsct, significantly different kinetic curves could be observed thereafter. conclusions: the immune cell reconstitution showed different mathematical curve models after allo-hsct. kinetic reconstitution model of certain immune cell was associated with agvhd and relapse. hence, mathematical kinetic models of immune cell reconstitution may be potential indictor for predicting agvhd and relapse after allo-hsct. disclosure: nothing to declare lineage specific chimerism analysis in pediatric patients following allogeneic hematopoietic cell transplantation (hct background: the outcome of allogeneic hct is dependent on several variables that include patient age, disease and stage, cytoreduction, graft, graft manipulation, and graft versus host disease (gvhd) prophylaxis. one aspect of hct that remains poorly defined and studied is the donor/ host (d/h) chimerism post hct. since , we followed patients with d/h lineage specific chimerism post hct. analyses were performed by short tandem repeat (str) polymorphism analysis at the american red cross blood services (philadelphia, pa). studies were performed on blood total leukocytes, myeloid/neutrophil cells, t-cells, bcells, and nk-cells. methods: in this retrospective study, the charts of consecutive patients who underwent allogeneic hct between january to june on the pediatric bone marrow transplant service at mskcc were retrospectively reviewed. lineage specific donor chimerism post hct was studied including d/h chimerism trend, and factors with potential impact on chimerism including: age, disease, graft source, and t-cell depletion (tcd). preliminary analyzes performed on this cohort included wilcoxon rank test and cox proportional hazard analyses. results: patients were selected based on the number of analyses. the median age was . years. patients had hematologic malignancies (n= ) or non-malignant hematologic diseases (n= ), or immune disorders (n= ). cytoreduction included tbi-(n= ), or chemotherapybased regiments (n= ). patients were recipients of t-cell depleted marrow or peripheral blood grafts (n= ), unmodified marrow or peripheral blood grafts (n= ) or cord blood grafts (n= ). full donor chimerism of myeloid cells, b-cells and nkcells, but not t-cells occurred early post-transplant. there was no difference in the percentage of total donor leukocytes at months vs. months post hsct (n= ), while the median of donor t-cell chimerism was % at months and % at months post hsct (p< . , n= ). figure shows the impact of different factors including: (a) the use of tbi-or chemotherapy-based regimens, (b) age (< or > years), and (c) type of graft (t-cell depleted vs unmodified vs cord blood). donor total leukocytes chimerism was significantly lower at months as compared to months for patients < years of age (p= . ). for most grafts, full donor chimerism of t-cells occurred early, while for t-cell depleted transplants, it took up to one year to complete. cord blood grafts were associated with high t-cell donor chimerism throughout the post-transplant period. there was a significant difference in the % donor t-cells at and months post hct when comparing t-cell depleted and unmodified grafts (p= . ). conclusions: this preliminary analysis of lineage specific chimerism post-transplant showed that donor tcells may take one year to fully recover post-transplant, mostly following t-cell depleted grafts, without intervention. cord blood grafts were associated with high donor chimerism throughout the post-transplant period. lastly, factors associated with loss of donor chimerism posttransplant were younger age and non-malignant disorders. more in-depth analyses are being performed including the relation of chimerism and hct outcome. disclosure: eileen nicoletti -employee rocket pharmaceuticals, susan prockop -investigator atara biotherapeutics -research funding, susan prockop -mesoblast -research funding, nancy kernan -gentium -support; jazz pharmaceuticals -support, richard o´reilly -atara biotherapeutics -royalty, consultancy and research, jaap jan boelens -bluebird bio -consultancy, avrobio -consultancy; jaap jan boelens -chimerix -consultancy; magenta -consultancy background: the success of hematopoietic stem cell transplantation (hsct) lies with the ability of the engrafted immune system to remove residual leukemia cells via a graft-versus-leukemia effect. despite this, relapse remains the major cause of mortality among patients receiving hsct. one of the immune evasion mechanisms of leukemic cells to escape from donor t cell recognition in haplo-hsct is the genomic loss of the patient specific hla. it has been described in - % of acute myeloid leukemia (aml) and myelodysplastic syndrome (mds) relapses after haplo-hsct. the aim of this study was to analyze hla loss in a large cohort of patients who relapsed after t-cell replete haploidentical transplantation with posttransplant cyclophosphamide. methods: from december to september , patients with hematological malignancies who received a haplo-hsct were recruited. among them, patients presented a relapse after haplo-hsct. hla typing was performed by real-time pcr using hla-kmr kit (gendx, netherlands).nine patients were excluded from the analysis because the kit employed did not include the recipientspecific hla. thus, a total of relapse cases were analyzed. the analysis of chimerism was carried out using short tandem repeat pcr amplification (ampflstr sgm plus, thermo fisher, usa) with a sensitivity of %. results: genomic loss of the patient hla occurred in / patients ( %) ( table ) . these patients presented different hematological neoplasms. interestingly, patients presented lymphoid neoplasm ( acute lymphoblastic leukemia (all-t), dentritic cell leukemia (dcl) and hodking´s lymphoma (hl)). hla loss relapses occurred later than classical relapses ( vs. days). regarding the treatment received (table ) , four patients were studied retrospectively. three of them were treated with donor lymphocyte infusions (dlis) + chemotherapy or other drugs at the time of the relapse. the other patient did not receive any treatment. in the end, all patients died from disease progression. prospectively, we detected hla loss at relapse in other two patients. at the moment of relapse, the first case received brentuximab + haplo-hsct from alternative donor and the other case received daratumumab + haplo-hsct (pending). both patients are alive, the first one in complete remission (cr) and the second one in partial remission (pr). conclusions: the frequency of hla loss at relapse is similar in our cohort to what is described in the literature. hla loss has been identified in patients with lymphoid neoplasms, while this mechanism has not been previously described in such diseases. the analysis of this immune evasion mechanism should be implemented in the routine screening of patients transplanted from haploidentical donors in order to design effective rescue strategies. these treatments should not be based on dlis or second transplantation with the same donor, instead, alternative donors should be used. background: an adequate immune reconstitution (ir) is crucial to reduce transplant toxicity, relapse rate and mortality after allogeneic stem cell transplantation (allohsct). the aim of this, single center retrospective study was to investigate the correlation between the recovery of different lymphocyte subpopulations with the main transplant outcomes, including overall survival (os), disease free survival (dfs) and non-relapse mortality (nrm). methods: we analyzed the ir of adult patients (aml n= , all n= , mds n= , nhl n= , hd n= , cll n= , cml n= , mm, n= , mpn n= ) who underwent (allohsct) between january and march . median age at transplant was years (range . - . ) with male/female ratio of %. donors were hlaidentical siblings (n= , %), family haploidentical (n= , %), matched unrelated ( , %), mismatched unrelated (n= , %) and cord blood units (n= , %). the stem cell source was the bone marrow (bm) in patients ( %), the cord blood in ( %) and g-csf mobilized peripheral blood in ( %). the conditioning regimen was myeloablative in ( %) transplant, reduced intensity in ( %) and immunosuppressive in ( %). gvhd prophylaxis was based on calcineurin inhibitors in combination with methotrexate or mofetil mycophenolate. antilymphocytes immunoglobulins was used in patients ( %) (anti thymocytes globulin, atg sanofi-genzyme in or anti t-lymphocyte globulin, atlg -neovii biotech, in ). the peripheral blood lymphocyte subsets (cd +, cd +cd +, cd +cd +, cd + (b cells) and cd +cd + (nk) were analyzed by flow cytometry at , , , , and months after hsct. post-transplant engraftment was molecularly determined by vntr analysis. results: as detailed in table the proportion of full donor chimerism analyzed in the peripheral blood t lymphocytes improved progressively after transplantation and the same pattern was observed when the chimeric status was measured in bone marrow mononuclear cells. to favor the achievement of a full donor chimerism, dli were performed in patients starting at the median of days after transplant (range: - ). with a median follow-up observation of months (range - ), the one year os and nrm was % and %, respectively. at months after allohsct, the achievement of values higher than , and /μl for cd +, cd + and nk cells, respectively was significantly associated to a better os (figure ), dfs (p = . ), and to a lower nrm (p< . for cd + and cd +, p= . for nk). a better lymphoid reconstitution was observed after the use of either a sibling or a haplo donor than a matched unrelated or cord blood donors. the use of atg was significantly associated with a delayed cd + recovery but with a faster nk cells reconstitution. conclusions: at six months after allohsct, recovery of cd + and nk cells predicts survival. monitoring of immune recovery may help to guide pre and post-transplant treatment strategies. days infections and disease control. several groups have demonstrated the independent prognostic value of different lymphocyte subpopulations in hsct outcomes. posttransplant cyclophosphamide (pt-cy) effectively prevents gvhd after hla-haploidentical hematopoietic stem cell transplantation (haplo). the use of pt-cy in hla matched related (mrd) or unrelated (mud) donors hsct, although less explored, has also been introduced. the aim of this study was to compare the early immune reconstitution after allogeneic hsct from haploidentical and hla-matched donors using pt-cy. methods: one hundred and sixty-four hsct performed in our center were analyzed: haplo performed between and and hsct from hla-identical donors ( consecutive mrd sct performed with pt-cy between and and mud sct with pt-cy between and ). pt-cy was administered at mg/kg/d in days + and + postransplant, followed by mmf mg/kg/ d and a calcineurin inhibitor (ciclosporina a or tacrolimus) from day + ahead. we retrospectively compared early immune reconstitution at day + and day + among these three populations. early ir was assessed through the analysis of lymphocyte subpopulations including total t lymphocytes cd +, cd + and cd +subpopulations, nk cd -cd + cells, cd + bright immature subpopulation and total b cd + lymphocytes.. lymphocytes subpopulations were determined by multiparametric flow cytometry (fc and navios, beckman coulter®). results: all patients, but mud and haplo, received pb as stem cell source. patients received prior transplant in haplo group. patient´s characteristics are shown in table . patients who received hsct from mrd showed the fastest ir, with statistically significant differences compared to haplo in almost all lymphocyte populations at day + (cd +, cd +, cd + and nk cells), and also in cd +, cd + and b lymphocytes at day + . comparison between haplo and mud hsct showed better ir among haplo, demonstrated by higher counts in cd +,cd +, cd + and nk cell counts at day + . no differences were seen at day + . (figure ). percentage of immature cd bright nk cells was higher in mud hsct at + , with no differences between haplo and mrd hsct. conclusions: in our cohort of patients with pt-cy based gvhd prophylaxis regimen, those who received hsct from mrd showed the earliest immune reconstitution compared to haplo and mud at day + and + . haplo showed better ir compared to mud at day + . nk maturation at day + was a little better among haplo and mrd hsct recipients than mud hsct patients. in our experience, using mostly pbsc as graft source, type of donor influenced early ir in pt-cy based hsct, background: cell-free dna (cfdna) isolated from plasma or serum has received increasing interest for diagnostic applications. however, the reported clinical usefulness of cfdna in patients undergoing allogeneic cell transplantation (hsct) is scarce. methods: the chimeric status both in peripheral blood and in cfdna obtained from plasma was investigated in patients undergoing hsct. dna and rna were isolated from plasma within four hours of blood draw. patients were evaluated for chimerism at day + , + and + post-transplant. a panel of seven microsatellites was amplified by pcr for chimerism detection and pcr products were analysed by capillary electrophoresis. for further cfdna characterization the micro rna (mirna) c was analysed using digital pcr. mutations frequently used for minimal residual disease assessment such as flt -itd, npm and jak were also investigated in cfdna. results: the mean cfdna concentration in transplanted patients was ng/ml, while in healthy donors used as control group (n= ) was ng/ml. the mean cfdna concentration difference between both groups reached statistical significance (p= . ). when analysing cfdna from transplanted patients and in the control group we could not detect dna fragments larger than bp and the size range of the analysed fragments was between and bp. in out of patients a mixture of donor and recipient cfdna (mc) was detected. with the exception of three patients relapsing after transplant in which mc was detected both in peripheral blood and plasma in the rest of the patients (n= ) mc was detected only in plasma. the mean percentage of recipient cfdna in the plasma samples was % (range: - %). all the patients with acute gvhd (agvhd) (grade: i-iv) (n= ) showed mc in plasma at least in one of the time-point tested. no significant difference was found in the mean recipient cfdna percentage in patients with agvhd grade i-ii when compared with grade iii-iv. meanwhile in the group of patients with chronic gvhd (n= ) mc in plasma was detected in patients. in those patients with clinical improvement of agvhd (n= ) a decrease in the percentage of recipient cfdna was observed during treatment. in patients without improvement or even agvhd worsening (n= ) stable or increasing recipient cfdna percentage was detected. since recipient cfdna can be detected in patients without transplant-related complications we analysed the mirna c expression in all patients with recipient cfdna. a significant difference was found in the mirna c expression in patients with agvhd (mean mirna c: . mirna c copies/ u copies) when compared with patients without gvhd (mean mirna c: . mirna copies/ u copies). in those patients with extramedullary aml relapse (n= ) frequent mutations (flt -itd, npm ) were only detected in the cfdna fraction. conclusions: longitudinal analysis of cfdna represents a useful complementary tool in particular for those patients with clinical complications after hsct. disclosure: nothing to declare comparison of the impact of atg/pt-cy-based and tcr αβ-depletion as gvhd prophylaxis regimens on the recovery of memory t-cell compartment background: over recent years haploidentical and hlamismatched donors have been increasingly adopted as a valid donor source. modern graft-versus-host disease (gvhd) prophylaxis regimens such as drug-based (antithymocyte globulin (atg), post-transplant cyclophosphamide (pt-cy)) or graft-manipulated (tcr αβ-depletion) demonstrate effective prevention of gvhd. here we report our data about an influence of different gvhd prophylaxis regimens after allo-hsct with pbsc as a graft source on cd + memory t-cells recovery. methods: our study comprised leukemia patients who underwent allo-hsct with pbsc as a graft source in national research center for hematology, moscow, russia. detailed patients characteristics are presented in table . peripheral blood samples were collected on day + , + and + after allo-hsct. flow cytometry analysis was performed on bd facs canto ii (becton dickinson, usa) to define t-memory subsets: t-naive and t-stem cell memory (tnv+tscm) -cd +cd r -ccr +cd +; t-central memory (tcm) -cd +cd r +ccr +cd +; t-transitional memory (ttm) -cd +cd r +ccr -cd +; t-effector memory (tem) -cd +cd r +ccr -cd -; t-terminal effector (tte) -cd +cd r -ccr -cd -. sysmex xe- was used to calculate absolute count of different t-memory cell subsets. mann-whitney u test was used for nonparametric data analysis. a p-value less than . was considered as significant. results: results of mann-whitney u test (calculated pvalues) to compare absolute number of t-memory cell subsets in terms of different gvhd prophylaxis regimens are presented in figure . during all follow-up period the number of tnv+scm and tcm remains significantly reduced after atg+pt-cy or tcr αβ-depletion compared to atg-based immunosuppressive regimen. on day + we observe no difference in the number of tnv+scm and tcm cells after atg+pt-cy or tcr αβ-depletion. terminally differentiated cd + cells (ttm, tem, tte) count is significantly lowered in tcr αβ-depletion patients group in comparison to atg+pt-cy. nevertheless recovery of tnv+scm and tcm after pt-cy is faster than after tcr αβ-depletion. conclusions: according to our data the mechanism of pt-cy is seems to be more selective compared to tcr αβ-depletion due to its transient impact just on tnv+scm and tcm without affecting on the effector pool. through this it may lead to delayed reconstitution of adaptive immunity after tcr αβ-depletion compared to using pt-cy. clinical relevance of the quantitative characteristics of immune recovery in the context of different approaches to gvhd prevention remains to be established. background: immune effector cells, belonging to either innate or acquired immunity, play a key role on preventing disease relapse after allogeneic haematopoietic stem cell transplantation (hsct). most of known immune effector are cd + cd + t-cells and cd -cd + natural killer lymphocytes, while cd + cd + cells act as modulatory and regulatory cells. the early post-hsct ratio between these cellular subsets may be an indicator of graft vs-tumor (gvt) effect. methods: we retrospectively revised the immune recovery of allogeneic hsct performed at our institution from to , analysed on peripheral blood by multiparametric flow cytometry lymphocyte subpopulations panel. diagnosis were acute leukemias ( %), chronic myeloproliferative neoplasms ( %), lymphomas ( %), myelodysplastic syndromes ( %), multiple myeloma ( %) and severe aplastic anemia ( %). we established early time-points of evaluation, and days from the graft infusion, to analyse the differences in disease free survival (dfs) and overall survival (os) between patients according to the cd + cd + x cd -cd + / cd + cd + ratio. results: median ratio at + days was of , . at this time point, patients who showed the ratio higher than the median had both a better dfs (median dfs time not reached vs months; p = , ) ( figure ) and os (median os time not reached vs months; p = , ). likewise, ratio at + showed an advantage on dfs (p = , ), and not on os (p = , ). other factors possibly affecting both dfs and os were analysed in univariate analysis, such as the use of antithymocyte globulin (atg), conditioning regimen intensity, graft source, hla-matching and disease status at hsct, the latter being the only variable with a significantly detrimental impact on both os and dfs. disease status was confirmed an independent valriable associated with both dfs and os as well as + ratio both on dfs (hazard ratio [hr] - , ; p = , ) and os (hr , ; p = , ). conclusions: our data show that cd + cd + x cd -cd + / cd + cd + ratio assessed at + is and independent predictor of transplant outcome, possibly representing a row indicator of anti-leukemic immune surveillance. the integration of this index with other known outcome predictors may help in improving the management of post-transplant phase. [[p image] . figure background: allogeneic stem cell transplantation (alo-hsct) is a curative treatment but it is associated with lifethreatening complications. most deaths are due to relapse, graft versus host disease (gvhd) and infection. the pattern and quality of the immune reconstitution (ir) after transplantation may affect these outcomes. however, there are limited data on the association of the quality of the ir and either the development of gvhd and survival. methods: eighty-five patients who received a non t-cell depleted alo-hsct in our center from to were prospectively studied. most patients received hla-identical grafts. total cd + and cd + t cells, ccr +cd + and ccr +cd + (which include both naïve and central memory t cells) and naïve ccr +cd l+ t lymphocytes were quantified by flow cytometry. data were collected at days + , + , + , + and + after alo-hsct. the association between ir and the gvhd was studied through an anova. for the multivariate analysis, a logistic regression was performed including those confusing clinical variables that were significant in the univariate analysis (p≤ . ). the study of overall survival (os) versus ir was performed with a cox regression model. results: total cd + t lymphocytes reached normal numbers within the first two months. median t cd + count was cells/ul after one month, which is within the normal range. conversely, it took nearly one year to get normal counts of cd + t cells ( cells/ul). the only two clinical parameters conditioning a worse recovery of the cd + t cells were the previous alosensitization of the donor and the sex, being female donor and male recipient the worst combination for the ir. no parameters influenced the quality of the reconstitution of cd + t cells. of note, the age or the hla status did not influence the quality of the ir. when the patients were divided into gvhd and no gvhd, we found no differences in the recovery of either the proportion or absolute count of every t cell subpopulation, including total t cells as well as naïve/central memory t cells, both cd + and cd +. finally, a multivariant analysis confirmed that the absolute counts of cd +ccr + t cells at day + as well as the absolute counts of both cd +ccr + t cells and naïve cd +ccr +cd l+ at day + were associated with better os. conclusions: in conclusion, neither the development of gvhd nor other relevant parameters seem to play a determinant role in the quality of the ir. to our knowledge this is the first study which demonstrate a clear association between the recovery of naïve cd + t cells measured by flow cytometry and the os. disclosure: nothing to declare p abstract already published. azacitidine (aza) for prophylaxis or pre-emptive therapy for myeloid neoplasms after allogeneic stem cell transplantation whom were treated prophylactically and preemptively. median age was years [range, - ] and all patients had a diagnosis of aml or high-risk mds. prophylactic treatment consisted of aza mg/m for days in cycles of days. in the pre-emptive setting, patients received mg/m for days per cycle and patients mg/m for days per cycle. a median of cycles [range, [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] was administered in the prophylactic group and of cycles [range, - ] in the pre-emptive group. ten patients also received at least one dli after the third aza cycle: patients in the prophylactic group and patients in the pre-emptive one. results: during follow-up, patients had significant delays in treatment plan due to transitory mild complications. however, % of patients (n= ) presented infectious complications requiring hospitalisation and % of patients (n= in the prophylaxis group and in the pre-emptive group) presented some form of gvhd. in patients who developed gvhd, had to discontinue treatment (all in the prophylaxis group); also patients discontinued treatment due to disease progression. the overall drop-out rate was . % (n= ). survival was analysed from initiation of treatment with aza and median follow-up was months. one-year efs was % in the prophylaxis group, with only one patient relapsing and no deaths. in the pre-emptive group, the -year efs was % and the median efs was months; -year os was % and median os was months. conclusions: we conclude that post-transplant aza treatment is a well-tolerated therapy, but the incidence of side effects remains discordant in the literature. results in the prophylaxis group are excellent, but patients with positive minimal residual disease treated pre-emptively had a lower outcome with only stabilisation of the disease. randomised prospective trials are needed to define patients who would benefit the most from this treatment and at what timing, dosage and duration of treatment. disclosure: nothing to declare. abstract already published. results: all subjects experienced hematopoietic engraftment at a median of days (range - ) and demonstrated full donor myeloid chimerism. m-mdsc and pmn-mdsc recovery peaked at a median of days posttransplant. the median peak absolute m-mdsc count was , cell/ml (range , - , /ml) representing a range of . % to . % of pbmcs. the pmn-mdsc peak was more robust, with a median absolute peak of , cells/ml (range , - , /ml) representing a median of . % of pbmcs (range . - . %) . of note, the one patient who developed severe, life-threatening gvhd had the lowest absolute and relative pmn-mdsc recovery ( , cells/ml and . % of total pbmcs). recovery of m-and pmn-mdscs occurred at a similar tempo and magnitude in two recipients of standard gvhd prophylaxis (tacrolimus/methotrexate). however, while mdscs isolated from ptcy recipients exhibited clear t-cell suppressive capacity, those from the comparison patients did not (see figure) . conclusions: mdscs recover rapidly and robustly after allohct using ptcy as gvhd prophylaxis, and may play a role in mitigating gvhd risk by mediating t-cell suppression. this may be a mechanism by which ptcy results in donor-recipient tolerance. background: high dose chemotherapy followed by autologous stem cell transplantation (asct) offers a cure in the upfront and relapsed setting in both hodgkin (hl) and non-hodgkin lymphoma (nhl). asct also remains standard of care in previously untreated multiple myeloma (mm) patients after induction therapy, if eligible. the availability of new cellular or other immune therapies that can be used after asct underscores the potential importance of monitoring immune reconstitution after asct. methods: immune reconstitution panels (irp) were evaluated retrospectively in all lymphoma and mm patients over a -year span ( - ) whom underwent asct at our institution. patients were included if they had a pre-asct measured within days of asct and two other irp at any of the following timepoints ( ) day - , ( ) day - , and ( ) at -year post-asct. patients in the lymphoma cohort had their irp excluded if they had additional treatment within the first year post-asct (other than maintenance rituximab). mononuclear cells from peripheral blood were analyzed by flow cytometry for assessment of lymphocyte phenotype and numbers. absolute values were compared using the mann-whitney u test. results: the data on patients were available for analysis ( mm, nhl, hl) . all lymphoma patients were conditioned with beam. all mm patients were conditioned with a standard high dose melphalan regimen. the median pre-asct absolute cd counts in the lymphoma cohort were significantly lower than the mm cohort at cells/μl vs cells/μl, respectively (p= . ). however, the mm cohort exhibited a greater percent reduction in cd cells on day at . % vs . %, respectively which continued through day at . % vs . %, respectively. this led to nonsignificant changes in absolute cd count by day at cells/μl vs cells/μl, respectively (p= . ) (figure ). the median absolute cd count pre-asct for mm and lymphoma cohorts were cells/μl and cells/μl, respectively (p= . ). similarly, a greater percent reduction in cd cells led to comparable absolute counts on day at cells/μl vs cells/μl, respectively (p= . ). the failure of post-asct cd reconstitution to pre-asct levels was driven by lack of cd + cell recovery, namely cd +cd ra+ cells with a median of cells/μl and cells/μl in the mm and lymphoma cohorts, respectively at day (figures and ). this led to markedly diminished cd :cd ratios through day (figure ). [[p image] . conclusions: impaired t-cell reconstitution in both lymphoma and mm continues through -year post-asct. as shown, a larger percent reduction in median cd and cd counts through day was appreciated in mm compared to lymphoma leading to the nonsignificant differences in the post-asct absolute counts despite significantly higher pre-asct counts in the mm cohort. impaired recovery of cd t-cells may increase the risk of opportunistic infections, decrease the response to vaccination and lead to ineffective anti-tumor response. further prospective and larger retrospective studies like this should continue in the modern-era as they may help predict responses to further interventions requiring a robust t-cell repertoire for maximal efficacy such as car-t cell and bite therapies. disclosure: nothing to declare p peri-transplant detection of measurable residual disease by multicolor flow cytometry is highly predictive for relapse in acute myeloid leukemia patients background: presence of measurable residual disease (mrd) prior to allo-sct has been shown to be predictive for survival in patients in hematological cr of aml. in this study we analyzed the impact of mrd in such patients measured by -color multiparameter flow cytometry (mfc) prior to and on day + post-transplant. methods: the bone marrow samples immediately prior to allo-sct and on day + post-transplant were retrospectively analyzed. mrd evaluation was carried out with antibodies against: ( ) results: a number of aml patients (male, n= ) with median age of years ( - ) in hematological cr prior to allo-sct were enrolled in the study. we observed lower survival in patients with mrd by mfc pre-transplant ( y os: % ( - %) vs. % ( - %), p= . ) due to increased relapse incidence ( % ( - %) vs. % ( - %), p= . ). in multivariate analysis, mrd pos prior to allo-sct has strong significant impact on os (hr . ( . - ) , p= . ). of patients, a sample both before and on day + after transplantation was available in patients. of those patients, ( %) were mrd negative prior to transplant and on day + (mrd neg/neg ); ( %) patients were mrd positive prior to transplant and negative at day + (mrd pos/neg ); and ( %) patients were mrd positive at both timepoints (mrd pos/pos ). dfs and os for these three groups were as follow: y dfs: mrd neg/neg : % ( - %), mrd pos/neg : % ( - %); mrd pos/pos : % ( - %, p= . ); y os: mrd neg/neg : %; mrd pos/neg : % ( - %); mrd pos/pos : % ( - %, p< . ). upon multivariate analysis, the mrd status prior to transplant and on day + showed strong significant impact on dfs (hr . ( . - . ), p= . ) and os ), p= . ). we did not observe any significant impact of other factors included in the multivariate analysis (patient's age, patient's sex, and recipient/ donor sex constellation). conclusions: mrd positivity prior to allotransplant and at day + by mfc is highly predictive for survival after allo-sct. disclosure: nothing to declaire background: immune reconstitution is a critical factor for risk assessment of life threatening infections and long-term survival in patients undergoing allogeneic cell transplantation (hsct). methods: immune cell subsets (cd , cd , cd , cd , cd +cd -) were quantified by flow cytometry. trec and krec were quantified simultaneously using droplet digital pcr (dpcr). a total of patients were evaluated. mean age at transplant was years (range: - years) samples were obtained before hsct and at day , and after hsct. results: absolute numbers of cd and cd cells remained below pre-transplant levels until day , increasing further and eventually reaching pre-transplant levels one year after hsct. absolute counts of cd and cd +cd -cells remained below pre-transplant levels beyond one year after hsct. cd cells were characterized by fast reconstitution kinetics, reaching pre-transplant levels already at day . b cells correlated with krec levels at all time-points tested, whereas t cells correlated with trec levels only one year after transplantation. when we compared cell subsets, trec, krec levels and the reconstitution kinetics thereof between patients with reduced intensity conditioning (n= ) or full conditioning (n= ) no significant differences were observed. patients with pre-transplant trec levels above the mean ( trec copies/ml blood) showed higher trec levels and a faster t-cell reconstitution after hsct suggesting that tcell reconstitution can be predicted by analysing thymic functionality before transplantation. indeed, in patients with a pre-transplant trec above trec copies/ml blood, the positive predictive value for an efficient t-cell reconstitution was . (p= . ). we analysed the recovery kinetics of the cell subsets, trec and krec levels in patients with and without transplant-related complications. patients with either acute graft-versus-host disease or severe infections showed a slower trec reconstitution when compared with patients without complications. conclusions: our data suggest that the analysis of immune cell subsets together with trec and krec quantification can be used to evaluate the immune reconstitution process after hsct. pre-transplant trec levels allow t-cell reconstitution efficiency prediction after hsct. disclosure: nothing to declare background: falling donor / mixed chimerism after allogeneic haematopoetic stem cell transplant (sct) is associated with an increased risk of relapse and the potential for graft rejection. donor lymphocyte infusions (dli) are often administered in patients with mixed chimerism to achieve full donor chimerism but there is little data on long term outcomes for dli given for persistent mixed chimerism. methods: a retrospective analysis of all patients administered dli for mixed chimerism between to january was performed. all patients were transplanted at the university hospital of wales within the south wales blood and marrow transplant (swbmt) programme. patients were identified by the swbmt database and additional outcome data gathered by review of patients' medical records. results: patients were treated with donor lymphocyte infusions between and january . thirty one patients treated for relapse (with or without mixed chimerism) were excluded as was a further patient with a mismatched donor. the rest were / match. twenty six patients received a total of donor lymphocyte infusions for mixed chimerism alone. the median age was years (range: - ) with % women. fourteen ( %) of the patients had sibling donor transplants and twelve ( %) from matched unrelated donors. indications for transplant were: for aml or saml (n= ), myelofibrosis (n= ), mds (n= ), hodgkin lymphoma (n= ), non-hodgkin lymphoma (n= ) and all (n= ). escalating doses of donor cd + t cells were administered commencing at × /kg to × /kg then increased at half log increments according to chimerism results until full donor chimerism was achieved. the median number of doses administered was (range - ). the median interval was days (range - ). the median dose was × / kg (range × - × ). seventeen patients ( %) achieved full donor chimerism and remained so until most recent follow up (median months, range - ). one patient continued to receive dli after the study period and later reverted to full donor. two patients had ongoing mixed chimerism with no evidence of relapse. two patients relapsed; one of whom later achieved remission. there were six cases of gvhd; acute gvhd (grade ii n= , grade iii n= ) and cases of chronic extensive gvhd. one patient had gvhd features consistent with overlap syndrome. a total of five patients died, four due to infection (one in a patient with gvhd) and one due to cardiac toxicity from previous treatment (confirmed on post-mortem). conclusions: the results of our single centre study help reinforce the evidence for dli in establishing full donor chimerism when mixed chimerism is detected in the absence of relapse. incremental dli dosing is an effective strategy and associated with a low relapse rate. caution should still be given to the risk of gvhd following dli, however the risk appears to be low in this study. larger prospective studies are ongoing to address the optimal dosing strategy for dli post-transplant. disclosure: nothing to declare hypomethylating agents for the treatment of relapsed acute myeloid leukemia after allogeneic blood stem cell transplantation: a single center experience mariarita sciume , giorgia saporiti , elena tagliaferri , nicola fracchiolla , federica grifoni , giorgia levati , luca baldini , francesco onida the post-transplant period with well-balanced profile of good efficacy and moderate toxicity. we retrospectively evaluated the safety and efficacy of hma +/-dli in a reallife cohort of aml patients relapsing after allo-sct. methods: data from all patients with aml who underwent allo-sct at our institution in the last years and subsequently received hma as a salvage treatment for disease recurrence or preemptively for loss of complete donor chimerism were collected. results: eleven patients with a median age of years (range - ) were identified; median time between allo-sct and time to hma therapy was months (range - ). according to eln genetic risk stratification, patients were classified in the favorable group, in the intermediate-i, in the intermediate-ii and in the adverse one. six patients were treated with aza, whereas the remaining patients with dac. the cycles were repeated every days. ten patients ( %) started hma for morphological aml relapse, while one patient received aza as a sequential treatment after dli administered for loss of complete donor chimerism. median number of cycles was (range - ). treatment strategy included combination with dli in patients ( in the dac cohort, in the aza cohort), while in one case of flt -itd + aml sorafenib was also associated to dac and dli. no grade / toxicities and no acute gvhd occurred. a clinically significant response was observed in four patients ( %), all receiving at least cycles of hma therapy; in particular, a complete remission (cr) was achieved in / patients treated for morphological relapse, including the one who received the dac/dli/sorafenib combination and one (favorable eln risk) who received aza alone (not eligible for dli due to a concomitant lateonset cutaneous grade gvhd). of interest, the latter patient also displayed a resolution of the cutaneous gvhd. full donor chimerism recovery with no gvhd was observed in the patient who received aza for the progressive donor chimerism loss not responding to dli alone. with a median follow-up of months (range - ), the median os from hma treatment in responding patients was months (range - ); at the time of data collection responses were maintained in all four patients. seven patients had died, six from aml progression and one for severe intestinal gvhd occurring after failure of dli+aza and a following salvage induction chemotherapy treatment. conclusions: although arising from a limited number of patients, our real-life experience of treatment with hmas +/-dli in aml patients relapsing after allo-sct showed a general very good safety profile and promising antileukemic activity, altogether suggesting a facilitation of the graftversus-leukemia effect (gvl) associated to a possible suppression of the gvh reaction. disclosure: nothing to declare conclusions: in this study, cd -positive cell count and igg value had recovered about months after sct. in our institute, we have achieved a low incidence of infection by education and medication for patients until recovery of cd -positive cell count and igg. however, we found a higher incidence of infection after recovery of cd -positive cell count and igg. at - months after sct, administration of prophylactic medications such as sulfamethoxazole-trimethoprim were terminated and social comeback such as return to school or work were achieved in most patients. it is possible that the high incidence of community-acquired infection was associated with their comeback. thus, we should consider additional prevention of infection for patients in this period and further evaluation of immunological markers is needed. disclosure: no potential conflicts of interest were disclosed. effect of minimal residual disease before transplantation on the outcome of haplo-identical hematopoietic stem cell transplantation for high-risk acute lymphoblastic leukemia yehui tan , sujun gao , xiaoliang liu , long su , wei han , yu liu , yangzhi zhao background: to analyze the effect of haploid hematopoietic stem cell transplantation (hid-hsct) on high-risk acute lymphoblastic leukemia (all), and to explore the effect of minimal residual disease (mrd) before transplant on the prognosis. methods: a retrospective analysis was made on high risk all patients accepted hid-hsct in our hospital from january to january . the clinical features, stem cell implantation, complications, survival and recurrence were compared between pre-transplant mrd + and mrdpatients. results: all the patients got successfully implanted. the overall survival (os) was . %, the disease free survival (dfs) was . %, the incidence of acute graft versus host disease (agvhd) was . %, including . % ii~iv degree agvhd and . % iii~iv degree agvhd. there was no significant difference in stem cell implantation, gvhd, cytomegalovirus and hemorrhagic cystitis between mrd + and mrdpatients. dfs and os in mrd + patients were significantly lower than those in mrd -patients, and the cumulative rr rate increased significantly, there was no significant difference in cumulative trm. conclusions: hid-hsct was an effective method to treat high risk all, but mrd + patients had high recurrence rate and poor prognosis. strategy adjustment should be considered to reduce tumor residual and the transplantation strategy should be optimized for these kind of high risk patients, so as to improve survival rate. disclosure: nothing to declare background: lymphocytes are responsible for the cellular and humoral immunity and, consequently, its recovery after allo-hsct might be linked with the survival after the procedure. the aim of this study was to analyze this hypothesis in our series of patients. methods: all the allo-hsct performed in our center from january through july were included in the analysis. median age was years (range: - ). pts were male ( , %) and were female ( , %). baseline diseases were: aml, lpd, mds, all, mpd, mm, and bmf. donor was unrelated in ( , %), and was family in cases ( , %) (including haplo-identical). stem cell source was pb in ( , %) and bm in pts ( , %). conditioning regimen was reduced in procedures ( , %) and intensive in ( , %) (including just one non-myeloablative). overall mortalities at days + and + (the latter in patients with follow-up superior to year) were , % and , %, respectively. median follow-up was months (range: - ). evolution of absolute lymphocyte counts (alc) and subpopulations at pre-hsct and during the first year after allo-hsct were analyzed. results: as shown in table , alc and cd + lymphocytes decreased after conditioning therapy, and recovered progressively during the post-hsct period. at day + , majority of patients had > alc/mcl, clearly improved compared to admission values. cd + lymphocytes at day + was still very low, but at day + around half of the series had - /mcl. we found a strong link between alc, cd + lymphocytes, and cd + lymphocytes at days + and day + with overall survival at day + of the series (table ) . conclusions: in our series, immunity recovery was a late event for majority of patients undergoing allo-hsct. in addition, in our experience, the precocity and quality of the alc, cd +, and cd + cells recovery was clearly linked with long-term survival. background: the reconstitution of t and natural killer (nk) cells after hematopoietic stem cell transplantation (hsct) strongly influences the outcome of hsct including viral infection and graft versus-host disease (gvhd). the purpose of this study was to investigate the clinical efficacy of immune reconstitution including t and nk cells after hsct in children. methods: we reviewed the records of patients who undergoing allogeneic hsct in department of pediatrics, pusan national university children's hospital, from january to july . the counts of t lymphocyte subsets and nk cells was monitored in peripheral blood by flow cytometric technique during , , , and months post-hsct. blood samples for cytomegalovirus (cmv) and epstein-barr virus (ebv) monitoring were tested by real-time pcr assay. results: for total of patients, the mean age was . years (range, months- years), of the patients were boys and was girl. out of a total patients without pre-hsct cmv viremia or cmv infection, ( . %) recipients experienced cmv infection. the number of cd + t cells in and months post-hsct was significantly higher in patients with cmv reactivation compared to patients without (median . /μl vs. . /μl, p= . , and . /μl vs. . /μl, p= . ) . in ( %) recipients presented acute gvhd, the number of cd + t cells in and months post-hsct was significantly lower in patients with acute gvhd compared to patients without (median . /μl vs. . /μl, p= . , and . /μl vs. . /μl, p= . ) . the number of nk cells in months post-hsct was significantly lower in patients with cmv reactivation and acute gvhd compared to patients without ( . /μl vs. . /μl, p= . , and . /μl vs. . /μl, p= . , respectively) . in multivariable analysis, acute gvhd was shown to be the decisive factor influencing total t cells (p= . ) and cmv reactivation was independently associated with cd + t cells (p= . ). the cd + t cells counts were associated with prior hsct history and acute gvhd (p= . and p= . ), and the cd + t cells counts were also significantly associated with donor type (p= . ). conclusions: overall, our study documents that immune reconstitution of cd + , cd + t cells and nk cells is strongly associated with cmv reactivation and acute gvhd. additionally, we show that acute gvhd is influenced by lack of sufficient numbers of nk cells as well as cd + t cells early after sct. cd + t cells, on the other hand, significantly increase after cmv-reactivation and most likely play an important role in reactivation. disclosure: nothing to declare background: curative effect of allogeneic hematopoietic stem cell transplantation (allo-hsct) depends on the alloreactive t-cell immune response toward residual malignant cells -graft-versus-leukemia reaction. however, alloreactive population has not been phenotypically defined. recent studies suggest that alloreactive t cells express both costimulatory and inhibitory receptors simultaneously. exhaustion caused by the inhibitory signaling dampens tcell functionality, which could lead to the disease relapse. here we aimed to investigate the expression of costimulatory and inhibitory receptors on antigen-experienced t cells after transplantation, to isolate subpopulation specific for allo-hsct patients and analyze their t-cell receptor (tcr) repertoire. methods: expression of coinhibitory and costimulatory molecules on pbmcs patients at various time points after allo-hsct was analyzed for expression of: cd , cd , cd , cd ra, ccr , cd , cd , cd , klrg , tigit, pd , cd and ox by flow cytometry and compared to healthy donors. cd +cd +cd -cd +cd +pd +tigit+ fraction and cd + cd + control fractions were separated on facs aria ii cell sorter. double barcoded cdna libraries of tcr beta-chains for both fractions were prepared and analyzed by sequencing on illumina platform. sequencing results were processed by migec, mixcr and vdjtools software. enriched clones were identified by fisher's exact test (p> - ). results: we did not find any significant differences between patients after allo-hsct and healthy donors in single marker's expression, but, when considering coexpression of co-stimulatory and inhibitory molecules on t cells we found that cd +cd +cd -cd +cd +pd +tigit+ subpopulation was significantly increased in allo-hsct patients. moreover it increased with the time since the transplantation (fig. ). this population was isolated by cell sorting and alongside with total cd + fraction subjected to tcr beta-chain repertoire sequencing. the population contained clones significantly enriched compared with cd + fraction representing potentially alloreactive cells. this hypothesis is further supported by the notion that the level of expression of cd and cd co-stimulatory molecules is lower in the group of patients who subsequently relapsed, compared with the patients with complete remission, while the expression of inhibitory receptors was high in both groups. conclusions: according to our data patients after allo-hsct have a phenotypically distinct t-cell population characterized by simultaneous expression of costimulatory and inhibitory markers. this population contains specifically enriched clones, which may be specific for alloantigens. further functional assays are needed to confirm the alloreactive potential of this subpopulation. besides low expression of costimulatory molecules combined with high expression of inhibitory receptors on antigen-experienced t-cells of patients after allo-hsct might be associated with a disease relapse. fondazione mbbm, monza, italy, ospedale san gerardo, laboratorio stefano verri, monza, italy background: poor graft function (pgf) is a severe complication after hsct, with a high risk of morbidity and mortality, mainly due to infections. donor cd + scb seems to offer high chances of haematological recovery, not jeopardized by gvhd. however, pediatric reports remain scarce. methods: out of patients undergoing transplantation in our pediatric unit from to have been retrospectively evaluated for at least line persistent cytopenia (hb< . g/dl, plt< /mmc, n< / mmc) and/or transfusion-dependency beyond months after hsct in the presence of full donor chimerism. bone marrow cellularity was evaluated through biopsy as further indicator of pgf. ( / ) to donate or medical decision ( / ). bone marrow cellularity was < % in % of the patients who underwent scb for which the histology was available ( cases), and % in those who have not been treated ( / ). at days after scb / ( %) patients had hematological response, which was complete in % and partial in % of the patients. only patient had no response. the infusion was always well tolerated with no adverse events, and no worsening of gvhd. haematological recovery occurred spontaneously at days after bone marrow biopsy in a significantly lower proportion of patients ( / , %, p< . ) within the non-scb group. in two cases platelets engraftment was significantly delayed, up to one year after bone marrow biopsy and in one case thrombocytopenia persists and the patient is still receiving thrombopoietin agonists and red blood cells transfusions at months after bone marrow biopsy. conclusions: a stem cell boost matched the goal to yield count recovery in our cohort. viral infections and gvhd may be possible risk factors for pgf.bilinear or trilinear cytopenia with transfusion dependency and bom cellularity < % and full donor chimerism are good indications for scb, that can provide a significantly earlier hematological reconstitution, without risks of gvhd. due to the proved early efficacy and safety of cd + stem cell infusion, we suggest that this procedure should be taken in consideration in children with severe bone marrow hypoplasia and persistent cytopenia after hsct. disclosure background: as allogeneic hematopoietic stem cell transplantation (hsct) is sometimes performed despite erythrocyte's antigens incompatibility and mismatch, it is essential to carefully track patients' genotypes after it. methods: for the study we used erythrocytes (n= ) and dna (n= ) from patients undergoing abo-or rhesus-mismatch hsct and their donors. we used posttransplant no transfused patients on the periods according transplant protocol by hemagglutination methods in plate and tube using monoclonal antibodies to abo and rhesus antigens (hematolog, russia). we extracted dna with dna kit (bag, germany) and conducted pcr-ssp with kits abo-type, rh-type (bag, germany). chimerism was assessed by the str-pcr analysis with cordis plus multiplex kit for amplification of polymorphic strmarkers and amelogenin loci. the fragment analysis was performed on a genetic analyzer. informative loci were chosen by comparison of pretransplant patient's and donor's dna. the percentage of donor chimerism was calculated using standard formula. precise rhce and abo genotypes were determined by direct sanger sequencing. we revealed patients with unexpected erythrocyte abo, rhesus phenotypes and genotypes after hsct on + ( patients) and + days (all patients). chimerism analyses on str showed in a.e.kh. and g.l.v. patients % of donor's dna and less than % of recipient's one. b.n.a. patient was relapsed and chimerism analysis revealed % of recipient's dna and % of donor's one. using serological methods and pcr-ssp we revealed genotypes abo * a b ; rhd+; rhce * ccee in patient a. e.kh. before hsct, abo * a o ; rhd+; rhce * ccee in her donor, and abo * a a ; rhd+; rhce * ccee on + d after hsct. genotype a a was no recipient's neither donor's origin. direct sequencing did not prove this genotype, but revealed donor's one.on + d serological methods and pcr-ssp also revealed donor's genotype in this patient. patient b.n.a. had genotypes abo * o /o ; rhd+; rhce * c w cee before hsct, her donor -abo * b o ; rhd +; rhce * ccee. on + d this patient relapsed, but rhesus genotype has been detected as rhd+; rhce * c wcee (lack e gene). direct sequencing revealed gene rhce*ee. abo genotype was recipient's origin -o o . in patient g.l.v. using serological and pcr-ssp methods we determined genotypes abo * o /o ; rhd +; rhce * ccee genotype before hsct, and abo * a / o ; rhd+; rhce * ccee genotype in her hsc donor. on + d patient had unexpected genotype abo * o /o (a lack of a antigen); rhce * ccee (a lack of e antigen). in order to explain unexpected patient's genotypes after hsct we sequenced her rhce and abo genes and found donor's genotype ccee; a o that was in agreement with results of str analysis. to resolve discrepancies between serological, pcr-ssp and sequencing analysis data we sequenced patient's rhce cdna and observed only ce allele. at present time the molecular basis of selective inactivation one of the two rhce alleles is not clear. on + d patient had donor's genotype. conclusions: what kind of mechanisms led to discrepancies between results obtained by different laboratory methods are still not clear. an interesting case of expression of only one rhce allele in patient g.l.v. allows us to suggest involvement of some epigenetic mechanisms like dna methylation or histone modification in this process. clinical background: in relapsed patients with acute b -lymphoblastic leukemia (all-b) who achieved complete remission (cr) after re-induction therapy, minimal residual disease (mrd; ≥ - all-b cells/ul) is often detected. according to available data, such condition varies from % to even % of cases, as assessed by pcr or flow cytometry (fc), while the presence of mrd is the most important risk factor for all recurrence. in this abstract, we describe our experience with bridging therapy using blinatumomab infusion after re-induction regimens and before the planned allogeneic stem cell transplantation (allo-sct). the procedure was performed in three young men suffering from relapsed ph (-) all-b at the age of , and years. in the first case ( yo), relapse with previous mrd accounting for . % occurred months after cr mrd neg . in the next patient ( yo) the second relapse with central nervous system (cns) involvement occurred months after allo-sct performed in cr ( months after cr , mrd neg ), while in the third patient ( yo), recurrence with cns and testis involvement occurred years after cr (mrd neg ). all patients underwent chemotherapy (flam, hypercvad and dnr/vcr/pegasp/dexa regimens respectively) followed by one cycle of blinatumomab (at a dose of mcg/d on days - , followed by mcg/d on days - in a continuous infusion) and allo-sct (using eto/cy/tbi/atg/ conditioning regimen for ist and iiird patient and bucy for iind patient; using matched unrelated donor (mud, ist and iiird patient) or matched related donor (iind patient)). mrd status was assessed after each cycle of blinatumomab by fc. results: all patients achieved cr mrd pos after reinduction therapy followed by clearance of mrd after blinatumomab course (tab. ). the second patient, due to positive mrd months after allo-sct received donor lymphocyte infusions additionally. during the administration of blinatumomab, no adverse events (aes) were observed in grade or . one patient developed cytokine release syndrome in grade . the progression free survival, time to positive mrd and follow up are presented in tab . conclusions: the use of blinatumomab as a bridging therapy between re-induction regimens and allo-sct in patients with all-b and mrd pos appears to be safe and leads to the clearance of mrd which may be crucial in os and pfs prolongation after following allo-sct. future studies on larger groups of patients are necessary to confirm this thesis. background: haploidentical hematopoietic stem cell transplantation (hsct) is considered an alternative treatment for hematologic malignancies in patients who do not have an hla-identical sibling donor [ ] . since infections and disease relapse resulting from delayed immune reconstitution (ir) are the most common causes of mortality among patients undergoing haploidentical-hsct [ ], timely ir is essential in the recovery and survival of these patients. the aim of this study is to describe the evolution of ir after haploidentical-hsct and to estimate survival rates in patients with delayed vs. adequate reconstitution in a single center in colombia, south america. methods: a retrospective cohort study was conducted on consecutive adult haploidentical-hsct recipients at a tertiary referral center. cd +cells, cd +cells, cd +cells, and immunoglobulins levels were monitored before hsct, at first month, and then every three months for the first two years post-transplantation. descriptive statistics were used to analyze patient's clinical characteristics. the kaplan-meier method was used to assess overall survival (os) and relapse-free survival (rfs) rates. results: twenty-six patients were included ( % were male), with a median age of . years (range - ). the most common indication for haploidentical hsct was acute lymphoblastic leukemia (n= , . %), followed by non-hodgkin lymphoma (n= , . %) and myelodysplastic syndrome (n= , . %). all patients received gvhd prophylaxis therapy with cyclophosphamide, tacrolimus, and mycophenolate mofetil. fifteen patients ( . %) presented cytomegalovirus reactivation ( / at risk), patients ( . %) epstein-barr virus reactivation, and patients ( . %) developed adenovirus infection. median time to neutrophil engraftment (neutrophils> . × /l) was days (range - ) for the patients recipients of peripheral blood progenitor cells (pbpcs) and days (range - ) for the three remaining bone marrow recipients. platelet engraftment, defined as > , platelets/ mm background: daratumumab is a human monoclonal antibody directed against the glycoprotein cd that is overexpressed on the surface of plasma cells in multiple myeloma patients. it is approved as second line therapy either as single agent therapy or in combination with lenalidomide or bortezomib for the treatment of patients with relapsed/refractory multiple myeloma. despite the curative potential of an allo-sct, the high relapse rate remains a clinical problem. data addressing the choice of an optimal salvage therapy regime for these heavily pre-treated patients is missing. methods: from april till november a total of patients (male, n= ) with the median age of years ( - ) received daratumumab as a salvage therapy for relapse of multiple myeloma after allo-sct at the university of hamburg. prior to allo-sct all but one patient had received an autograft, patients even ≥ autografts and patients also a . allograft. the median number of salvage lines post-transplant and prior to first daratumumab infusion was ( - ). these salvage regimens included cyclophosphamide, etoposide, bortezomib, lenalidomide, pomalidomide and carfilzomib. daratumumab was started at a median of months ( - ) after relapse/ progress and initiated as single agent therapy in all patients. concomitantly, patients received either an immunomodulatory drug (lenalidomid, n= ; pomalidomid, n= ) or a proteasome inhibitor (bortezomib, n= ) during a later course of daratumumab infusions. combination therapy was initiated when a slow rise of paraprotein and/or free light chains or no response to monotherapy was observed (median at the th infusion). results: the median number of infusions was . twenty adverse reactions were observed in of ( %) patients: dyspnea (n= ), bronchospasm (n= ) shivering (n= ) , cough (n= ), musculoskeletal pain (n= ), acute coronary syndrome (n= ), skin rush (n= ), facial edema (n= ), pressure on eyes (n= ). all adverse reactions appeared during the first infusion and were mostly mild or moderate (ctc - , n= ). tolerance of the following infusions improved and in none of the cases therapy had to be stopped due to adverse events. three patients developed late onset infections (pneumonia, n= ; urinary tract infection, n= ) followed by temporarily therapy interruption. with a median follow-up of months after the first administration of patients remain alive . %). one patient died due to progress of myeloma and another died due to severe infection/sepsis. of patients responded ( %; pr, n= ; vgpr, n= ; cr, n= ) to the therapy with daratumumab. the responses (decrease of paraprotein and/or free light chains ≥ %) occurred at a median of days ( - ) after the first administration and lasted for . months ( . - . ). conclusions: daratumumab shows an encouraging efficacy and acceptable toxicity profile in patients with relapsed/refractory myeloma after allo-sct. further studies are needed to investigate the role of the combination therapy with immunomodulatory drugs or proteasome inhibitors in this setting. disclosure: nothing to declare p clonal plasma cell detection by high sensitive flow cytometry in aphaeresis product is poor prognostic and not increased by use of plerixafor alone background: in an earlier from our center we have demonstrated that residual clonal plasma cells (cpc) decrease both overall survival (os) and disease free survival (dfs) (ash ).plerixafor is a selective antagonist of cxc chemokine receptor (cxcr ) and able to mobilize human peripheral blood stem cell (pbscs) by acting synergistically with g-csf.the purpose of this study was to evaluate the safety and efficacy of plerixafor in myeloma patients who were proven poor mobilizers and specifically to assess the flow cytometric measurement of residual clonal plasma cells in the apheresis products. methods: patients with a diagnosis of mm who underwent auto hsct at our center between january -november were retrospectively analyzed.out of patients, patients received plerixafor as mobilization regimen due to poor mobilization with g-csf.pbsc grafts were tested for the presence of clonal pcs (cpc) and the number of normal pcs (npc) by multi-parameter flow cytometry (fcm).the acquisition of the cells was performed using the navios flow cytometer beckmancoulter) .upon the daily checks of the instrument, x cells for each sample were acquired and the collected data was analyzed using the kaluza software (beckmancoulter,usa). results: patient demographics are shown in table .the majority of patients were male and median age was years in the plerixafor group.the median interval from time of diagnosis to mobilization and follow-up from mobilization were . months and . months in plerixafor group, respectively. cpc contamination in the pbsc grafts was detectable in and patients with counts ranging between - . x - and - . x - in g-csf alone and g-csf+plerixafor groups, respectively (p= . ).there were no significant differences in the proportion of the patients with graft contamination between subtypes of mm in both groups. one hundred (gcsf/plerixafor; / ) patients had pre-asct pet-ct imaging done with (gcsf/plerixafor; / ) have active lesion at the time of mobilization. statistically significant association could not be demonstrated between the disease < cr status at mobilization and the number of apc in the apheresis product in both groups (p> . ).twelve of patients from plerixafor treatment arm proceeded to transplantation within median . months.the best overall response to induction treatment is shown in table .thirtyfour patients from the g-csf alone arm and patients from the g-csf+plerixafor arm died during the follow-up (p= . ).disease progression was seen in patients from g-csf alone group and patients from g-csf+plerixafor group of the study(p= . ).estimated mean os was better among patients w/o apc contamination in plerixafor group, respectively ( . ± . mos vs . mos; p= . ). conclusions: our results on and few plerixafor used patients show that clonal plasma cells are detectable by multiparametric flow more frequently when patients are poor mobilizers and require plerixafor.the clonal pc contamination can be attributed to the myeloma biology as manifested by higher number of lines induction regimens and pet positivity among the plerixafor-required patients. the overall and disease survival was impaired by residual clonal pcs in the graft but not by plerixafor per se. neither was the content of clonal pcs differed from others.thus the cxcr shared by hsc and myeloma cells do not cause a myeloma mobilization. clinical trial registry: -disclosure: nothing to declare prognostic factors for overall survival after allogeneic hematopoietic cell transplantation in multiple myeloma patients all factors with significant influence on pts survival were included multivariate analysis (cox regression model) but only re-admission in the first days demonstrated impact on os (hr , ; p= , ) . conclusions: we analyzed risk factors for survival in mm pts who received allo-hct. our study identified disease-related risk factors like iss and transplantationrelated factors such as hct-ci and pam, hospital readmission, days of hospitalization and cmv reactivation that were associated with worse long-term survival. in our series, the most frequent death and re-admission cause was infection, so focusing the efforts in reduction of infection could have a beneficial impact on improvement of survival in mm undergoing allo-hct. [[p image] . figure ] disclosure: there is no disclosure. novel protocol for autologous hsct in multiple myeloma: ambulatory chemomobilization and transplantation of fresh hematopoietic stem cells with backup storage background: autologous hematopoietic stem cell transplantation (ahsct) after melphalan conditioning is still a part of standard treatment of multiple myeloma patients. traditional transplantation of frozen stem cells poses additional risk for the patients connected with dmso and central venous catheter. the transplantation of fresh cells is an option -however, most mobilization protocols are either low-efficient (g-csf), expensive (g-csf + plerixafor) or toxic (standard dose chemomobilization) to directly proceed to transplantation in this fragile group of patients. we describe here the novel combination of ambulatory mobilization with very low doses of ara-c and g-csf connected with direct ahsct with fresh cells. methods: the prospectively collected database of patients after ahsct was searched for patients who underwent ahsct after chemomobilization with ara-c and transplantation with fresh cells (fc) and compared with control group of consecutive patients transplanted with standard protocol (sp) (transplantation with frozen cells) between july and october . protocol of ambulatory mobilization was: mg/m² of arac on days + and + and g-csf at the dose μg/kg/day from day + and escalated to μg/kg/day split into two doses + to + , apheresis started on day + (or later) and finished when at least . x e cd + positive cells were collected. the collected cells were split in three equal parts: ) for use as fresh transplant ) frozen for possible nd transplant ) frozen as backup. results: there were transplantations with fresh cells and transplantations with frozen cells compared. both groups had same mobilization protocol -ambulatory low dose ara-c. the median age and number of transplanted cells was similar in both groups ( vs , p= . ; . vs . cd +/kg, p= . conclusions: we present novel approach that allows direct ahsct after chemo mobilization in all patients who are treated with melphalan. we show that it is not only feasible to do ahsct directly after chemomobilization but also the results may favour this approach when compared with current standard. disclosure: nothing to declare background: high dose chemotherapy followed by autologous hematopoietic cell transplantation (hsct) is considered, since the nineties, the standard of care for patients aged less than - years old with newly diagnosed multiple myeloma (mm). however, the optimal induction treatment prior to hsct to reduce the tumor burden has changed during the last few years. improved regimens have shown to be able to increase the quality of the pre-hsct response, which might subsequently impact on the post-hsct response, which has been proved to be associated with a longer pfs. we recently changed the induction therapy for pts candidates to hsct. in this analysis, we aimed to check if newer regimens impacted on pretransplant responses, and how auto-hsct changed the pre-hsct status. methods: all the auto-hsct for mm patients performed in our center from january through august were included in the analysis. median age was years (range: - ). pts were male and were female. durie-salmon stage was distributed as follows: i ( . %), ii ( . %) and iii ( %); % had creatinine > mg/dl. iss was: ( %), ( . %), and ( . %). type of monoclonal component was: igg ( . %), light chains ( . %), iga ( . %) , and non-secretory ( . %). . % had bence jones proteinuria. conditioning regimen was melphalan mg/m in ( . %), melphalan - mg/m in ( . %), and other in ( . %). results: pre-transplant therapy was: vcd in (mostly in - ), vtd/vrd/krd in (mostly in - ), and others in cases. status of the disease at transplant was: cr/vgpr in , pr in , and sd in . distribution of pretransplant response based on the type of induction is shown in table . peri-transplant mortality was %. day + mortality was . % ( pts), due to progressive disease. as shown in table , all patients ( / ) who obtained cr pre-hsct, maintained the response at day + post-hsct. among the patients in vgpr at hsct, ( . %) became into cr, and ( . %) maintained the response. the cr rate at post-hsct increased % compared to pre-hsct ( versus pts). altogether, after hsct pts ( . %) improved and ( . %) maintained the pre-hsct response. during the last period of time, pts started on post-hsct maintenance/consolidation, mainly with lenalidomide. conclusions: ) with the new chemotherapeutic schemes, . % of patients underwent hsct in cr or vgpr; ) majority of pts ( . %) consolidated or improved the pre-hsct response; ) cr increased substantially ( . times) after transplant; ) optimized induction regimens, along with auto-hsct followed by the recently licensed use of maintenance therapy with lenalidomide, might result in a better pfs of patients with mm. background: autologous stem cell transplantation (asct) is commonly used in treatment of patients over years with multiple myeloma (mm), however the safety and efficacy of this procedure is debatable. methods: we conducted a retrospective review of mm patients who underwent asct from to at our institution. the purpose of this retrospective study was to compare the -day mortality, time to engraftment, and incidence of grade - toxicities in elderly mm patients with younger patients taking into account comorbidity information. other secondary end points measured were overall survival (os) and progression-free survival (pfs). results: a total of patients were analysed and categorized by age as young patients ( to y; n= ) or elderly ( to y; n= ). the compared groups did not differ in terms of gender, ecog, hct-specific comorbidity index (hct-ci), and disease status at asct. melphalan in a dose of mg/m was used as preparative regimen in % of younger patients, and in % of the elderly (p= . ). the remaining patients received mg/m of melphalan or lower dose (range, - mg/m ) due to hct-ci > or age, on the physician discretion. in the whole study group there were no transplant related deaths within the first days of asct. stratifying by age, there was no statistically significant difference concerning febrile neutropenia (fn) incidence, which was observed in % of younger patients, and % of elderly. in contrast, fn was observed more frequently in patients with hct-ci > ( % vs %, p= . ). grade - infections were more frequent in older patients ( % vs %, p= . ), but no difference was found in grade - infections incidence rate such as pneumonia, uroinfections and neutropenic enterocolitis ( % vs %, p= . ), nor grade - and - noninfectious toxicities ( % vs %, p= . , and % vs %, p= . , respectively) . the median time to granulocyte engraftment was days (range, - days) in elderly and was comparable with younger patients. the time to platelet recovery was also similar. after the median follow-up of months for survivors, os at months was % in both groups. pfs at moths was % for younger patients, and % for elderly (p= . ).however, the association between pfs and the dose of melphalan used in conditioning was observed. pfs probability at months for patients conditioned with the dose of mg/m , mg/m and mg/m was %, % and %, respectively (p= . ). conclusions: our data show that asct in transplant eligible mm patients ≥ years of age is safe and provides similar outcomes as seen in younger patients. disclosure: nothing to declare is mobilization with chemotherapy effect response in the multiple myeloma? background: high dose melphalan therapy with autologous stem cell support is a standart approach in symptomatic multiple myeloma patients. response rates increased with the novel anti myeloma agents and the use of chemotherapy for stem cell mobilization should be questioned. the purpose of this study is to determine the effect of cyclophosphamide used during stem cell collection on disease response and transplantation results. methods: we retrospectively collect data from myeloma patients who underwent autologous stem cell transplantation (asct) in ankara university medicine faculty, blood and bone marrow transplantation unit between january and november . patients who received cyclophosphamide protocol for stem cell mobilization were included in the study. disease response were determined according to international myeloma study group (imwg) criteria before and after the cyclophosphamide. transplant responses and their effects on survival were also indicated. results: after the diagnosis of mm, patients (male/ female: / ; median age: years (between - years)) with median follow-up of . months (between , - , months) underwent asct at a mean of , ± , months.. forty-one patients were evaluated before and after cyclophosphamide (table ). in % of the patients, the disease response was not altered by cyclophosphamide therapy, and % of the patients improved their response status. post-transplant response rates of patients who underwent stem cell mobilization with cyclophosphamide are also shown in table- . the mean survival of the patients was , ± , months. when patients were grouped according to changes in response status before and after cyclophosphamide; there was no statistical difference between mean calculated survival (improved response, disease progression and stable disease; , ± , months, , ± , months and , ± , months respectively, p= . ) (figure- ) . the rates of -year and -year overall survival (os) of the patients with no response to cyclophosphamide treatment were as follows; , %± , % and , %± , % respectively. thirteen patients who were followed up median months after transplantation died at an average of , ± , months; of these deaths were caused by the infection after transplantation. conclusions: in our study, it was observed that the use of cyclophosphamide for cd + stem cell mobilization did not change the disease response rates by %. the posttransplant survival rates of mm patients who had progressive disease after cyclophosphamide use were lower. however, these results warranted confirmed by randomized controlled trials. clinical trial registry: -disclosure: nothing to declare results of a single center experience: an attempt to augment conditioning regimen in first autologous stem cell transplantation treatment of multiple myeloma (mm) continues to evolve in the era of novel agents. the addition of bortezomib to highdose melphalan (bor-hdm) has been reported by several groups, and it has been outcome and toxicity profile is comparable to high dose melphalan (hdm) alone. the aim of this retrospective study was to evaluate the outcome of the bor-hdm conditioning regimen on overall response for patients with mm undergoing first single asct at our institution. methods: this retrospective single center study reviewed consecutive myeloma patients who had received the first asct either with bor-hdm (n= , m/f= / ) or single agent hdm (n= , m/f= / ) conditioning regimen. in the single agent hdm conditioning regimen, melphalan was administered intravenously at a total dose of mg/m on day - and - and stem cells were infused on day . in the bor-hdm group, melphalan mg/m was administered on day - . bortezomib was administered intravenously at a dose of mg/m on day's - , - , + , and + as described in a phase study by intergroupe francophone du myeĺome (ifm). results: all consecutive patients who underwent single asct from january to march using bor-hdm as conditioning or hdm were evaluated. conditioning regimen was hdm in patients and bor-hdm in patients. median age was significantly lower in bor-hdm conditioned asct compared to hdm group ( years vs years, p= ). there was no significant difference for mm subtype, iss stage at diagnosis, prior treatment line among hdm vs bor-hdm cohorts (p> . ). after a median of cycles of induction chemotherapy, patients in the bor-hd exhibited ≥vgpr of . % (n= ) compared to . % (n= ) in the hdm group (p= p> . ). pre-asct immune response (if (-)) was reported in . % of patients treated with hdm, higher than that seen in the bor-hdm group ( . %) (p= . ). nine ( . %) patients achieved post-asct immune response (if (-)) ≥vgpr compared to ( . %) in the hdm group. at the time of this analysis, ten patients in the bor-hdm group and in the hdm group had already died, respectively (p> . ). a total of ( . %) patients in the bor-hdm group and ( . %) patients in hdm group had already progressed (p> . ). estimated mean os and pfs was shorter for group treated with bor-hdm; . ± . mos and . ± . mos vs. . ± . mos and . ± . mos, respectively (p> . ) (figure- ) . we could not demonstrate the impact of pre or post transplant ≥vgpr immune response on survival and disease free survival. there was no engraftment failure observed on either treatment group and no worsening peripheral neuropathy was developed in the bortezomib arm. conclusions: this single center experience on a small patient pool was able to repeat the prospective randomized study results of ifm. further studies are warranted to explore this regimen, especially when induction treatment with novel agents are employed, with special emphasis on the high-risk myeloma patients where response rates are good but sustainability remains an issue. disclosure: nothing to disclosure the efficacy and safety of bortezomib plus busulfan/ melphalan as conditioning regimen in multiple myeloma undergoing autologous stem cell transplantation: phase / study background: bortezomib have a powerful antimyeloma activity and was regarded as backbone of therapy in the past decade but its safety and efficacy as a part of busulfan/ melphalan conditioning regimen of autologous stem cell transplantation is yet to be shown. methods: a phase / trial to explore the safety and activity of a bortezomib on days - , - , and + added to a conditioning regimen with busulfan and melphalan (bumel, . mg/kg/day and busulfan during day - and - , mg/ m /day of melphalan on the day - ), in multiple myeloma (mm) patients who received autologous stem cell transplantation following bortezomib-based induction chemotherapy. in phase , escalating doses ( . , . , and . mg/m ) of bortezomib with bumel were administered in each group with three patients. with determined maximum tolerated dose of bortzomib at a . mg/m /day, cohort with patients were analyzed for phase . results: in phase , no dose limiting toxicity was observed at a . mg/m /day of bortezomib. in phase , overall responses at months was shown as % of very good partial response (vgpr) or better and % of complete response (cr), whereas post-transplant overall best response included % of vgpr or better, and % of cr, respectively. with median follow-up duration of . months, median progression-free survival (pfs) was . months. the probabilities of years-pfs and overall survival (os) were . % and not estimable, respectively. especially, high-risk cytogenetics were associated adverse survival outcome compared to standard-risk cytogenetics, respectively (pfs, . vs. . months, p= . ; os, . vs. . months, p= . ) . with median days and days for neutrophil and platelet engraftments, any graft failure or delayed engraft was not observed. the common grade or severe non-hematological adverse events included neutropenic fever ( . %) and stomatitis ( . %). except three cases with transplant-related mortality due to sepsis, other adverse events were manageable. conclusions: these results demonstrate that bortezomib is safe and can be a part of conditioning regimen in combination with bumel, for patients with transplanteligible multiple myeloma. clinical background: allogeneic stem-cell transplantation (allo-sct) is one of treatment option for patients with multiple myeloma (mm) refractory to novel agents. the reports on allo-sct for mm are limited and it is an important issue to argue appropriate conditioning regimens and stem-cell sources, and patient population who will benefit from allo-sct. methods: we retrospectively analyzed consecutive patients who received allo-sct for relapsed and refractory multiple myeloma (rrmm) between oct and july at japanese red cross medical center. characteristics of patients, progression-free survival (pfs), and overall survival (os) were analyzed. results: median age at allo-sct was (range - ). twelve patients were male and were female. myeloma type were igg: , iga: , igd: , and bence-jones: . stem-cell sources were peripheral blood from hlamatched related donor (rpbsct): , bone mallow from hla-matched unrelated donor (mud): , bone marrow from hla-mismatched donor (mmud): , and cord blood (cb): . twenty-three of patients received flu/mel-base, one patient received bu/mel-based, and one patient received etoposide/cyclophosphamide-based conditioning regimens. twenty-two patients who transplanted after received gy of total body irradiation (tbi). responses before allo-sct were cr: , vgpr: , pr: , sd: . five-year pfs was % ( %ci: - ) and -year os was % ( %ci: - ). ten patients died during observation period and causes of death were primary disease: and treatment-related mortality: . patients with vgpr or better before allo-sct showed significantly better pfs (p= . ) and os (p= . ) as compared with others. female recipients showed significantly better pfs (p= . ) and os (p= . ) as compared with male recipients. recipients of mmud showed significantly better pfs (p= . ). among patients surviving, patients received treatments including maintenance therapy. conclusions: the reason for better pfs and os in female recipients is unknown. it is interesting that recipients of mmud showed better pfs, suggesting graft-versusmyeloma effects. allo-sct can be an effective treatment option if patients and stem-cell sources are appropriately selected. disclosure: authors declare that there are no conflicts of interest. second autologous hematopoietic stem cell tranpslant versus chemoimmunotherapy in relapsed multiple myeloma after first transplantation: single center data background: combination therapy, mostly triple, followed by autologous hematopoietic stem cell transplantation (auto-hct) is widely accepted as the first-line standard therapy for multiple myeloma (mm). despite the availability of agents such as new immunomodulatory drugs (imids), proteasome inhibitors (pis), histone-deacetylase inhibitors and antibodies, it is still possible to achieve longer and deeper responses, however, multiple myeloma is still not cured and relapse is inevitable. the availability of these novel agents has increased questions for determining optimal treatment of patients with relapse after the first auto-hct. methods: we retrospectively analyzed patients who relapsed according to international myeloma working group (imwg) criteria after st auto-hct. first group [salvage chemotherapy(ct)] (n= ) was treated with only chemoimmunotherapy because of early relapse or refractory first auto-sct (within months), ineligible to second transplantation because of co-morbidity, unwillingness to transplant. second group (n= ) (salvage transplantion) was treated with second auto-hct as a salvage therapy. consolidation and long term maintenance treatments were used in both groups. results: there was no difference in sex and age between salvage ct and auto-sct groups [female/male: vs / vs ; ]. the best response after salvage auto-sct was complete remission (cr) in , %, partial remission (pr) in , % patients, while cr in %, pr in , % patients treated with salvage ct. progression free survival (pfs) were significantly better in second transplant group (pfs; % on the first year; , % on the second year after transplant vs % on the first year; % on the second year after the salvage therapy in chemotherapy patients)[p: , ]. overall survival (os) in salvage auto-sct group was longer than salvage ct ( , % . %), although it did not reach a statistical significance (p> . ). time to achieving the best response after salvage auto-sct and salvage ct was ( - ) month versus , ( - ) months [p: , ]. grade or nonhematological toxicities were similar (auto-sct %, salvage ct %) in both groups. conclusions: salvage auto-hct may provides longer progression free survival with similar toxicity profile according to chemoimmunotherapy especially in patients with sensitive to first auto-sct. it is suggested that earlier and better responses, long-term progression free survival can be achieved with salvage auto-sct. we believe that there will be statistical significance in os such as pfs by increasing the number of patients. the authors believe that large scale randomized clinical trials are needed for optimal treatment of relapsing multiple myeloma after first auto-sct. disclosure: nothing to declare background: one of the conditions for successful transplantation of autologous hematopoietic stem cells (auto-hsct) in patients with multiple myeloma (mm) is the timely recovery of hematopoiesis, which is associated with the quantitative and qualitative characteristics of the graft. one of the key indicators is the content of cd + cells in the autograft, which depends on many factors. some of them are due to previous treatment, others are directly related to the patient: age, stage of the disease, features of the hematopoietic stem cells (hsc) microenvironment. the aim of the study was to assess the influence of the immune response genes on the autograft cellularity in patients with mm. methods: А retrospective analysis of the genotyping results was performed. evaluation of loci in genes immune response and harvesting of autologous hsc in patients with mm has been made. hematopoietic stem cell mobilization regimen included cyclophosphamide g/m with granulocyte colony-stimulating factor. genotyping of the immune response genes polymorphic regions was carried out by the polymerase chain reaction with allelespecific primers. the number of cd + cells was counted on a -color facs canto ii flow cytometer. results: according to the results of the autologous transplant harvesting, two groups of patients were identified. first included patients with an autograft cellularity of more than × /kg body weight. the second group consisted of patients examined with the number of cd + cells in the autograft ≤ × /kg of the patient's body weight. comparing the identified haplotypes of the immune response genes with the cellularity of the transplantation material, it was found that the presence of the mutant allele in the homo-and heterozygous haplotypes of the il β gene (t- c) increased the chances of harvesting cellular material with a higher content of cd + cells in times (χ = . , p= . ), and the carriage of the wild type allele in the homo-and heterozygous state of the tlr (arg gln) gene is more than in times (χ = . , p= . ). currently, it has been shown that single nucleotide or amino acid substitutions in genes can lead to changes in the expression pattern of their final products: increased secretion of interleukin β (il- β) or changes in the spatial configuration and functionality of the receptors (tlr ). thus, in the presence of mutations in the il β gene, the enhanced synthesis of il- β influences on fibroblasts, immunocompetent, endothelial, epithelial and other cells, by activating hemopoiesis. in turn, the mutational status of the arg gln locus located within the tir domain of the tlr receptor in the cytosol, determines the spatial configuration of the tlr acting as a co-stimulatory receptor of cd + cells, which ensure the engraftment of the graft. conclusions: identified haplotypical features of the il β and tlr genes in patients with mm may act as predictors of the response effectiveness to mobilization of hscs in their carriers, which may contribute to the mobilization regimen optimization and will contribute to harvesting the optimal cellularity of an autologous graft. clinical trial registry: none. disclosure: authors declare no conflict of interest. differentiating diffuse from focal pattern on computed tomography: added values of a radiomics approach background: focal pattern in multiple myeloma (mm) seems to be related to poorer survival and differentiation from diffuse to focal pattern on computed tomography (ct) has inter-reader variability. therefore the purpose of this study is to assess if a radiomic approach could help radiologists in differentiating diffuse from focal patterns. methods: we retrospectively reviewed imaging data of patients with mm between january and september of whom ( men and women; mean age . ± . ) with ct, pet-ct or mri available before bone marrow transplant. two general radiologist evaluated in consensus only ct images to define a focal (at least one lytic lesion > mm in diameter) or a diffuse (lesions < mm, not osteoporosis) pattern. radiomic analysis on ct thinslice images was then applied with regions of interest (rois) done by one researcher not expert in medical imaging or mm blindly to the condition of the patients. the reference standard to differentiate diffuse from focal pattern was done by radiological evaluation of two expert musculosketal radiologists blinded to the clinical data reviewing ct, mri and pet-ct images. n= radiomics features were extracted and evaluated with an open source software. mann-whitney u test for unpaired data with bootstraps samples was used to compare radiomics features of diffuse and focal patterns and then feature reduction was done to avoid over-fitting. receiver operator characteristic (roc) analysis with area under the curve was done to compare radiologists and radiomics evaluation against reference standard. reading time to perform radiomic analysis was also estimated. results: the pathological group included: diffuse and focal patterns. after feature reduction, features were different (p< . ) in the diffuse and focal patterns (n= / features were shape-based: majoraxislength and sphericity; n= / were gray level run length matrix (glrlm)). mg/kg). a number of eleven patients did not receive any additional immunosuppression except of post-cy. results: after a median follow up of . months (range . - . ) patients were alive. the -year probabilities of pfs and os were % ( - %) and % ( - %).the cumulative incidences (cis) of relapse and nrm at years were % ( - %) and % ( - %), respectively. lower serum albumin level at transplantation (≤ g/dl) was associated with increased relapses (hr . ( . - . ), p= . ) and nrm (hr . ( - ), p= . ) and resulted in poorer pfs ), p= . ) and os ), p= . ). mmud and haploidentical donors were associated with poorer nrm (hr . ( . - . ), p= . ), and resulted in decreased pfs ), p= . ). the high-risk cytogenetic at diagnosis showed no impact on survival. the cis of acute (grade ii-iv) at day + and chronic gvhd at years were % ( - %) and % ( - %), respectively. absence of immunosuppressive medication beside post-cy was associated with poorer os ), p= . ). conclusions: the conditioning with bu, tt and post-cy leads to a favorable pfs and os due to low incidences of relapse and nrm for patients with multiple myeloma relapsing after autografting. disclosure: nothing to declare methods: between january and may , we included patients with mm who underwent asct and received bortezomib/lenalidomide/dexamethasone (vrd) consolidation and maintenance therapy, mainly lenalidomide(r) mg/day for days every days. results: the median age at transplant was years ( - ). forty-six ( %) of patients received r maintenance, patients received vrd maintenance for higher risk features. median duration of r maintenance was months . r dose was changed for toxicity (grade i-ii) in ( %) patients. twenty-nine ( %) patients relapsed: ( %) patients were shifted to different treatment protocols (treatment change). patients ( %) were kept on the same r maintenance (observation group) and ( %) patients had increased lenalidomide dose with dexamethasone (r/ d group). patients ( %) of the last groups required change of treatment later. the median follow up was months . median tnt was months ( - ). at years, the estimated pfs and os were % and . % respectively. the median os and pfs (from change of therapy) were and months for patients in the observation group, versus and months in the r/d group, and and months with treatment change, respectively. no statistically significant difference was noted. conclusions: our small monocentric study is limited by its retrospective design and small sample size. however, it suggests that increasing lenalidomide dose as well as adding dexamethasone in selected patients can postpone change to different lines of treatment without affecting survival. disclosure: nothing to declare can the drugs used before autologous hematopoietic stem cell transplantation have impact on cmv reactivation that results in decreased os in myeloma patients after asct? more intensive treatment regimens, such as proteasome inhibitors (pi) and/or immunomodulatory (imid) agents. we performed a retrospective, single center study to evaluate the incidence, risk factors, and outcomes of cmv infection in patients with mm who underwent asct with a high-dose melphalan-based regimen. methods: this study involved a retrospective review of all patients with who underwent asct between january and november at our stem cell transplantation center. a total of consecutive adult patients with a diagnosis of mm (median age at diagnosis: , range: - ) underwent asct following induction treatment with novel agents (pis and/or imids). all patients received antiviral prophylaxis with acyclovir mg/day (n= ) or valaganciclovir mg/day (n= ). results: baseline patient characteristics, according to induction treatment, are summarized in table- . one hundred-five of the patients ( . %) were cmv iggpositive before asct. overall, . % (n= ) of cmvseropositive patients developed at least one episode of cmv viremia (cmv dna > copies/ml) after a median months (range; - mos) follow-up. persistent cmv viremia (detectable cmv dna load in more than sequential plasma specimens) occurred in . % ( of ) of the seropositive asct recipients and all of them were preventive treated with ganciclovir (n= ) or valganciclovir (n= ). the time from stem cell infusion to the development of cmv viremia ranged from days to days. none of the patients with untreated viremia developed identifiable cmv sequelae. no case of primary infection in seronegative patients at transplant was observed. adding to that none of the patients developed cmv disease post asct. if we analyzed the subgroups of patients according to induction therapy (pi-based, imids, pi+imid), the incidence of post-asct cmv reactivation was higher but not statistically significant, in patients who received only pi vs pi+imid ( ( . %) vs ( . %); p= . ). in univariate analysis, we could not demonstrate the importance of induction therapy with novel agents the occurrence of a post-asct cmv reactivation requiring antiviral treatment. however, statistically significant association found between the disease < vgpr status at asct and cmv reactivation ( . % vs. . %; p= . ). after a median follow-up . months (range; - . months), there was no significant impact on pfs, however there was significant decrease in estimated mean os who had cmv reactivation when compared to those without cmv reactivation ( . ± . vs. . ± . ; p= . ) (figure- ) . conclusions: cmv establishes lifelong latency within host cells and in the setting of impaired cellular immunity; cmv may reactivate from latency, disseminate, and directly cause life-threatening disease. our data suggests that mm patients treated with pi-based induction regimens and immunological response < vgpr at time of asct seem to have higher risk of developing symptomatic cmv reactivation. however, further studies on a large number of patients are warranted to clarify these findings. clinical background: high-dose therapy followed by autologous stem cell transplantation (asct) has been shown to prolong survival in patients with multiple myeloma (mm) in randomized trials. however, these trials only include patients aged < years. data regarding safety and outcomes in this patient population is lacking. methods: the aim of this study was to compare safety profile and outcomes in mm patients younger and older than years-old who underwent asct in our unit from july to october . patient's demographics, clinical characteristics, transplant related variables and probability of admission to the intensive care unit (icu) were analyzed. patients aged < and ≥ years-old would be called m and m , respectively, from now on. sorror index was used to estimate risk of mortality in the two cohorts. results: a hundred and eleven patients with mm underwent asct in the study period. median age was . years-old (range - ) and . % were male. thirtythree ( , %) patients were ≥ years. the probability of having a high risk comorbidity index was similar in both groups (m , vsm , %). the median cells obtained in the apheresis procedure was . x ( , - . ) in m compared to . x ( . - - ) in m . there were no differences in median admission lenght between the cohorts (m : days vs m : days). median days for neutrophil recovery above was days in both groups with a wider range in m ( - ) compared to m ( - ) . no differences were found in platelet recovery above . (m days vs m days). median packed red blood cells and platelets transfusions were ( - ) and ( - ), respectively, in m . in m cohort, they were ( - ) and ( - ), respectively. the incidence of grade - mucositis in m and m was . % and . %, respectively. there were no statistically significant differences in terms of using morphine for pain control between the two cohorts (m , , % vsm , , %). none patient requiered total parenteral nutrition (tpn) in group m and only one in group m . the incidence of icu admission was . times higher in patients aged ≥ than in patients < years-old , % vs , %), but differences were not statistically significant (p = . ). there were no deaths during the transplant procedure in any of the cohorts conclusions: ) in our series, high-dose therapy followed by autologous hematopoietic cell transplantation in mm patients aged ≥ was feasible. ) transplant procedure in older patients was as safe as in patients < years-old. ) no differences were found in terms of graft, transfusion support, transplant related complications and length of admission. ) age should not be a limiting factor in considering the modality of asct in this patient population disclosure: nothing to declare the correlation between the kinetics of peripheral blood counts and the response to treatment after high-dose melphalan with stem cell support in multiple myeloma patients background: the long-term survival of mm patients has dramatically increased in the last years, particularly for younger patients. this is attributable in part to the introduction and development of high dose chemotherapy with melphalan with stem cell support (hdm-asct). currently, frontline asct is still considered the standard of care for all eligible patients. many prognostic factors pre and post transplantation have been identified, e.g.: age, comorbidities, cytogenitcs, response to treatment and disease status prior to and post transplantation. to our knowledge there is no data correlating between kinetics of counts response to melphalan and prognosis. our aim was to assess the prognostic significance of the neutrophil and platelets decaying counts after high dose melphalan. methods -we retrospectively analyzed our cohort of multiple myeloma patients who underwent hdm-asct at the hadassah medical center bone marrow transplant department, between the years - . the kinetics of neutrophil and platelet decay during the first two weeks after melphalan administration was fitted using linear and exponential mathematical models. methods: we retrospectively analyzed our cohort of multiple myeloma patients who underwent hdm-asct at the hadassah medical center bone marrow transplant department, between the years - . the kinetics of neutrophil and platelet decay during the first two weeks after melphalan administration was fitted using linear and exponential mathematical models. results: factors associated with prolonged os in univariate analysis were: iss stage (p= . ), ≤ lines of treatment prior to asct(p< . ), favorable cytogenetics(p= . ), response to treatment (pr or better, p= . ) and rapid linear neutrophil decay (p = . ). in multivariate analysis, only ≤ lines of treatment before hdm-asct and rapid linear neutrophils count decay remained statistically significant for os prolongation. no predictive threshold value of the neutrophil decay incline was found. improved pfs was associated with ≤ lines of treatment prior to asct, and the response status after hdm-asct (p= . , p= . ). additionally, toxicity evaluation showed prolonged neutropenia to be associated with inferior os (hr = . , p= . ) and rapid exponential decay of neutrophil counts to correlate with higher incidence of mucositis (p = . ). fast platelet decay was associated with delayed platelet engraftment (p< . ) conclusions: we have shown that rapid linear decay in neutrophil counts predicts better os without a significant benefit in pfs in mm patients undergoing hdm-asct. this discrepancy might reflect the problematic estimation in a retrospective analysis of pfs. rapid decrease in neutrophils and platelet counts was associated with more toxicity: higher mucositis rate and delayed engraftment, respectively. therefore a rapid decay of blood counts after hdm-asct appears to be an in-vivo phamacodynamic marker of higher efficacy and toxicity of melphalan. disclosure: nothing to declare p do we need to freeze hematopoietic cells for autotransplants in patients with myeloma conditioned with melphalan? daniel garcia belmonte , beatriz aguado bueno , miguel herrero coderch , rafael de la camara background: multiple myeloma (mm) is the most frequent indication of auto-hsct, representing % of all auto-hsct in (passweg jr. bmt ; : - ) . nearly all are performed with peripheral blood progenitor cells (pbpc), and melphalan mg/m is considered the gold standard conditioning regimen. the standard procedure consists in obtaining progenitors, cryopreserved with dimethyl sulfoxide (dmso) and stored and subsequently thawed and re-infused in the patient on day . the procedure of cryopreservation is expensive and has some inherent toxicities (dmso) and loss of cells during the procedure. several groups have used non-cryopreserved progenitors showing some benefits compared with cryopreserved transplants, mainly a faster engraftment and a shorter length of hospitalization. objective: to compare noncryopreserved vs cryopreserved auto-hsct in mm methods: we perform an unicentric, retrospective study on consecutive first auto-hsct mm patients transplanted with pbpc between nov- and oct- , and conditioned with high dose melphalan ( mg/m ). the median follow-up was days (range: - ). patients received non-cryopreserved and cryopreserved auto-hsct. patients characteristics, without differences between non-cryopreserved vs cryopreserved: women/men ( / ); median age was years (range - ); in the majority auto-hsct was done as consolidation after first line therapy ( %); year of transplant ≤ ( %), ≥ ( %). the number of infused cd cells were not different: median . x /kg (range . - . ) in noncryopreserved patients and . x /kg (range . - ) in cryopreserved patients. results: we didn´t observe significant differences in the day of engraftment between non-cryopreserved vs cryopreserved although always was a little bit faster in the noncryopreserved group with a tendency to faster platelet engraftment (> /mm ): > platelets/mm (median day: vs . , p . ), > platelets/mm (median: vs days, p . ); > neutrophils/mm (median: vs . days, p . ). the media of days of hospitalization was shorter in non-cryopreserved patients ( vs days) although not statistically significant (p . ). transplantrelated mortality at day + was % in both groups. overall survival at years was not different: . % in in non-cryopreserved vs . % in cryopreserved patients (kaplan-meier, log-rank p . ). the accumulative incidence of relapse at the median follow up ( days) was similar: . % in non-cryopreserved vs . % in cryopreserved patients. conclusions: in our short experience, auto-hsct with non-cryopreserved pbpc in myeloma patients conditioned with high dose melphalan obtain similar results to those performed with classical cryopreserved pbpc and might has a faster platelet engraftment and shorter length of hospitalization. if no advantages are associated with cryopreservation, the simplicity of using fresh product is appealing. disclosure: nothing to declare p abstract withdrawn. . results: a early death is observed in one pt (group ) and pts(group ). the median delay of aplasia is days ( - ) and days ( - ) respectively. in group , among the evaluable pts, / ( %) are in cr, pts in pr and refractory. in group , cr: / ( %), pr: and refractory. a relapse is observed in pts/ ( , %) in group and pts/ ( %) in group with a frequency of % and % respectively in the first months. at months: pts/ ( %) in group and pts/ ( %) in group are dead. at months: pts ( %) and pts ( %). at months: pts ( , %) and pts ( , %). the overall survival (os) of the group and group pts were % and % at months; . % and . % at months; % and , % at months respectively (without significant difference). the event free survival (efs) of group and pts were % and , % at months, % and % at months and % and % at months respectively (without significant difference). conclusions: these protocols with equivalent toxicity allow obtaining of long-term equivalent results on the response rate early transplant, on the rate of relapse and on the overall survival. these results are identical to those of fermand ( ) . disclosure: nothing to declare background: dimethylsulfoxide (dmso) is a major intracellular cryoprotectant, used for cryopreservation of stem cells. it is toxic to both cells and patients at temperatures above o c. reduction of this effect is achieved by either washing of cells after thawing or by reduction of dmso during freezing and storage. the latter requires addition of extracellular cryoprotectants to the freezing media. we assessed the effect of low dmso concentration and different hematocrits of the frozen cells on cell viability and hematologic recovery in patients, transplanted for multiple myeloma. methods: cells were non-programmed frozen and stored at - o c in a cryoprotectant solution achieving final concentrations of % dmso, . % of hydroxyethyl starch (hes, weight average molecular weight da) and % of human serum albumin. the cell concentration in the frozen product for the first patients ( transplantations) varied between x and x cells/ml. in an attempt to reduce the amount of dmso infused, for the rest of the patients (n= ; transplantations) we further decreased the volume of the freezing suspension by removal of the entire plasma. the average age of the transplanted patients was ( - ). the cells were bedside thawed at o c water bath. the average cell dose was , x /kg ( , - , x /kg). results: viability of the stem cells following thawing assessed by trypan blue exclusion was , % . the hematocrit of the frozen cells had no effect on cell viability ( , %(low) vs , %(high)). the major complaints, if any, during stem cell infusion were coughing and an increase in nausea and vomiting induced by the prior conditioning. the average time for hematological recovery was , days (between and ) for the neutrophils, and , (between and ) days for the platelets. there was no significant difference in viability and hematologic recovery ( , and , vs , and , ) between patients receiving cells frozen with low or high hematocrit. conclusions: dimethylsulfoxide, despite its cryoprotective properties, is toxic for stem cells at temperatures above zero c and induces many side effects (cardiac, neurologic, respiratory, etc.) in the patients. to reduce those side effects we use lower dmso concentration, high hematocrit resulting in lower volume of the frozen cell suspension, thus reducing the final quantity of dmso to be infused to the patients. this does not affect the cell viability or the hematologic recovery of patients after transplantation. our easily performed method for unprogrammed freezing of stem cells with final dmso concentration % at - o c is safe, well tolerated, and provides cryopreservation, which allows high viability and stable cell engraftment, while reducing the undesired side effects of dmso. disclosure: nothing to disclose the conditioning regimen consisted of melphalan for most of the patients. the average age at the time of transplantation was years ( - ). patients were transplanted with an average cell dose of , x /kg ( , - , x /kg) for the first transplantation and , x /kg ( , - , x /kg) for the second one (every patient received the same cell dose as for the first) with average cell viability , % ( - %), with little difference between first and second transplantation. results: the average time for hematological recovery was , (between and ) days for the neutrophils, and . (between and ) days for the platelets. we found no correlation between the cell dose and the hematological recovery. there was no difference in the hematopoietic recovery between the first and the second transplantation in the patients, who underwent tandem or two transplantations. conclusions: recovery time is considered by some to be a function of the effective stem cell number. we did not find such correlation, probably because in the analyzed group all the patients, except four of them, received a dose greater than x /kg cell, which is accepted as a safe dose for autologous stem cell transplantation. disclosure: nothing to disclose p plerixafor-mobilized patients have a high risk of noninfectious fever during engraftment after autologous peripheral blood stem cell transplantation background: plerixafor enables rapid and efficient mobilization of hematopoietic stem cells. however, its impact on adverse clinical events after autologous peripheral blood stem cell transplantation (pbsct) is not fully understood. fever is one of the major complications in the preengraftment phase of pbsct. in this research, we focused on non-infectious fever around the time of bone marrow recovery and investigated whether plerixafor as mobilization therapy plays a role in engraftment fever. methods: we reviewed autologous pbscts for treatment of multiple myeloma at the japanese red cross medical center between - . non-infectious fever was defined as temperature ≥ °c with onset between two days prior to and two days after engraftment without clinical or microbiological documentation of infection. results: patients were mobilized by cyclophosphamide and filgrastim in . % (n = ) and filgrastim and plerixafor in . % (n = ). the median number of transfused cd + cells were . × /kg and . × / kg, respectively (p= . ). patients transfused with plerixafor-mobilized grafts had a higher risk of noninfectious fever ( . % vs . %, p< . ). cd + cell number or cyclophosphamide pretreatment had no relationship to non-infectious fever. the recovery of lymphocytes was more rapid in plerixafor-mobilized patients (p= . ). however, the number of lymphocytes was not associated with non-infectious fever. conclusions: combination of filgrastim and plerixafor as mobilization therapy resulted in an increased risk of noninfectious fever during engraftment comparing to mobilization with cyclophosphamide and filgrastim. while the mechanism remains unclear and requires further studies, plerixafor-mobilized grafts may result in an unintended increase in engraftment fever. clinicians should be aware of this possibility if patients are transplanted with those grafts. disclosure: ks received honorarium outside the submitted work from janssen, novartis, celgene, ono pharmaceuticals, takeda, fujimoto pharmaceuticals and srl. ti received honorarium outside the submitted work from janssen, celgene, ono pharmaceuticals and takeda. we assessed the efficacy of a new conditioning regimen consisted of decitabine (dec), busulfan (bu), cyclophosphamide (cy), fludarabine (flud) and cytarabine (ara-c) for allo-hsct in patients with mds and mds/ mpn. fifty patients were enrolled, including with mds and with cmml. patients received dec mg/m /day on days - to - , combining bu/cy/ flu/ ara-c modified preparative regimen. results: at a median follow-up of ( - ) days, the overall survival (os) was %, disease-free survival (dfs) was %, and relapse incidence was %. the incidence of severe acute (grade iii/iv) graft-versus-host disease (gvhd) was %, and that of (predominantly mild) chronic gvhd was %. os at years was % for mds patients with high risk, % for mds patients with very high risk, respectively. the survival was delightful in patients with poor-risk mutations, such as tp and asxl , ( %) and with three or more gene mutations ( %). among the total patients with poor-risk mutations in our research, only one patient ( %) with tp relapsed and one ( %) with asxl and tet died. result of continuous observation after transplantation, the percentage of nk cells in the peripheral blood of all patients who had received dec/flu/bu/cy/ara-c conditioning increased at day , which may essentially contribute to disease control post-transplantation. conclusions: in summary, the addition of a -day schedule of decitabine to a flu/bu/cy/ara-c conditioning regimen has proven feasible, with a low level of toxicity and promising early disease control especially in patients with high risk mds. disclosure: there are no conflicts of interest. the sfgm-tc mds score at day is associated with post-transplant outcomes in patients with myelodysplastic syndrome who underwent cd + selected allogeneic stem cell transplant conclusions: in patients with mds undergoing tcd-hct, the sfgm-tc score at day is significantly associated with survival. the lower incidence of acute gvhd in recipients of cd -selected transplants and the use of myeloablative condition regimens, with lower relapse, may explain the difference with the original finding that the sfgm-tc was predictive at day in unmodified grafts. disclosure the most frequent grade , toxicities were thromobocytopenia and neutropenia. infections developed in patients ( . %), neutropenic fever in ( . %). five patients ( . %) either developed or experienced exacerbation of acute graft versus host disease (gvhd), nonechronic gvhd. conclusions: azacitidine use is associated with only modest activity in patients who relapse after allo-hsct. however, in patients who respond to treatment it may allow for a durable disease control. disclosure: the authors declare no competing conflicts of interest background: somatic mutations in mds patients are closely related with clinical phenotypes and prognosis in mds patients. but whether mutations are prognostic for outcomes after allogeneic hematopoietic stem-cell transplantation (allo-hsct) remains to be elaborated. methods: targeted mutational analysis were performed on samples obtained before transplantation from patients underwent hsct. we analyzed the relationship of mutations and clinical outcomes. results: all patients carried more than one somatic mutations, most frequently in kmt d( . %), arid b ( . %), ccdc ( . %), pclo( . %), asxl / ( . %), srcap( . %), u af ( . %), dnah ( . %), ush a( . %) and tet ( . %). tp mutations were associated with higher ipss-r risk, complex karyotype and monosomal karyotype. dnah were more frequent in pediatric patients. in univariable analyses, tp mutations were related with decreased disease-free survival (p= . ); dnah mutations were related with increased disease-free survival (dfs) (p= . ). in multivariable analysis including ipss-r stratification, gvhd, hct-ci and candidate genes, dnah mutations were independently associated with better dfs(p= . ). conclusions: dnah mutations is independently associated with better outcomes in mds patients treated with allo-hsct while tp may predict unfavorable outcomes. accounting for these somatic mutations may help better selection of candidates for allo-hsct among mds patients. disclosure background: there is a controversy among experts if and how patients with mds and saml should receive cytoreductive therapy before transplant. while aiming to reduce disease burden in order to lower the risk of relapse after transplant cytoreductive therapy is associated with several drawbacks. besides a considerable risk for toxicity and mortality preventing patients to proceed to transplant cytoreductive therapy may also favour the selection of resistant clones which may be difficult to treat at relapse. methods: to address this hypothesis we retrospectively analysed the response and survival following salvage therapy in patients with mds and saml who had relapsed in median . months ( to months) after allo-sct according to their pre-transplant strategy (upfront transplantation n= %; induction chemotherapy [ctx] n= %; hypomethylating agents [hma] n= %). results: the majority of these patients received salvage therapy with hma (n= , %; aza n= , dac n= ) mostly in combination with dli, while the remaining received other salvage treatments (intensive chemotherapy n= , dli alone n= , nd transplant n= , bsc n= , miscellaneous n= , missing information n= ). when focussing on those patients treated with hma and dli it became apparent that a significantly higher proportion of patients in the upfront group ( %) achieved cr after salvage therapy in comparison to pre-treated patients ( % cr, p= . ; ctx group % cr; hma group % cr). accordingly, overall survival (os) calculated from the time of relapse was significantly longer in patients in the upfront group than in the group of pre-treated patients ( -year os % vs. %, p= . ). conclusions: overall, these findings imply that pretransplant therapy may favour the iatrogenic selection of resistant clones, which poorly respond to salvage therapy with hma and dli in case of relapse after allo-sct. furthermore, the results support the concept that an upfront transplant strategy is a promising alternative for patients with mds and saml that can be augmented by salvage therapy with hma and dli. disclosure: ts and gk received travel support, lecture fees and research funding from celgene gmbh conclusions: in our country, this procedure has shown to be feasible and we hope to improve it, with better infection control and by acquiring more experience related to the management of these patients. background: extramedullar relapse of mds is a rare complication after allogeneic stem cell transplantation. we present the case of a -year-old woman who was admitted into hospital because of insecure walking. paresis of both legs, hypaesthesia of the inner thighs, increased effort at urinating, reduced sphincter tonus, central paresis of the right arm and discreet paresis of the right facial nerve were documented at neurological exam. mri showed a large tumour of the dorsal thorax that immured the adjacent ribs and spine, affected the processus transversus of t - and invaded the spinal canal. the patient had undergone ric allogeneic stem cell transplantation five years ago for mds-eb with complex aberrant karyotype. following an uneventful course and no signs of gvhd, she had been off immunosuppression since , years. at the time of the admission the patient had slightly lowered wbc ( , gpt/l) and plt ( gpt/l) and clearly increased ldh ( u/l). methods: histology of a ct-based biopsy of the paravertebral tumour showed an infiltration of the muscles by blastous cells that were cd -, cd -, pax -positive, tdt and cd a were questionably positive. provisonal diagnosis therefore was lymphoblastic lymphoma, pox tested negative. the bone marrow was hypocellular with increased numbers of mature lymphocytes, but no definite signs of malignancy. cerebrospinal fluid revealed cells/μl with % blasts. immunotype was cd , cd , cd , cd , hla-dr positive, pox and lymphatic markers were negative. because of this we finally suspected meningeosis leucaemica. we completed the diagnostic workup with genetical and chimaerism tests and compared the result to the patients' mds before allogeneic stem cell transplantation. [[p image] . mri scan of the large thoracic tumour] results: cerebrospinal fluid (csf) cells consisted of % recipient cells, whereas peripheral blood cells were % donor. high risk mds at transplant displayed a complex caryotype including trisomy and tetrasomy , now % of the cells in csf showed trisomy and % tetrasomy . chimerism and fish of the solid tumour could not be performed, coexpression of myeloid markers within the tumour is pending. conclusions: in conclusion the patient has meningeosis as a result of exclusively extramedullary relapse of myeloid blasts originating from the initial high risk mds with blast excess and complex aberrant caryotype. the evolution of a trisomy clone to tetrasomy clone in relapse is linked to extramedullar manifestations. whether the solid tumour represents myeloid sarcoma with coexpression of lymphoid markers, extramedullary relapse of mds with lymphoid differentiation or, less likely, a separate lymphobastic lymphoma, is not yet clear. disclosure background: adoptive t cell therapy with genetically engineered t cells is a potent innovative immunotherapeutic approach for cancer treatment. unfortunately, the use of t cells redirected against tumor antigens, is severely limited by ) the difficulty in identifying appropriate cell surface antigens, that could be targeted by car t cells and ) the paucity of tumor-specific t cell receptors (tcrs) against shared, oncogenic antigens. methods: focusing on wilms´tumor (wt ), a tumorassociated antigen overexpressed by acute myeloid leukemia and several solid tumors, we designed and implemented an innovative protocol for the rapid isolation of wt -specific t cells and for the generation and characterization of a library of wt -specific tcrs displaying different human leukocyte antigen (hla) restrictions, to be exploited by tcr gene transfer and tcr gene editing. to this aim, we repetitively stimulated t cells with autologous antigen-presenting cells, including immortalized b cells, pulsed with overlapping peptides spanning the entire wt protein. t cell recognition was assessed by flow cytometry in terms of cd a expression and ifnγ production. recognized peptides were mapped by a deconvoluting grid and t cell clonotypes were longitudinally tracked by tcrαβ sequencing. results: we successfully expanded tumor-specific t cells from consecutive healthy donors, in an average of rounds of in vitro stimulations. the ability of wt specific t cells to recognize naturally processed epitopes and their on-target specificity was demonstrated upon coculture with antigen-expressing targets including primary leukemic blasts. tracking of the tcrαβ repertoire during culture led to the identification of clonotypes that recognize several tumor-associated peptides and are restricted by more than hla alleles, including hla-a* : . tcrs were then expressed via genome editing. briefly, simultaneous editing of endogenous tcr α and β chain genes was achieved using crispr/cas technology (efficiency > %), followed by transduction of t cells with lentiviral vectors encoding wt -specific tcrs (efficiency > % of cd + t cells). phenotypic characterization of edited t lymphocytes showed a major enrichment of cells harboring t stem cell memory properties. functional validation of the edited t cells is currently ongoing. preliminary results of a hours coculture experiment show that tcr edited t cells kill fresh wt + leukemic blasts, harvested from hla-matched patients, with an efficiency up to % at an effector to target ratio of to , while no killing of controls is observed. conclusions: we set up a protocol enabling consistent and efficient hunting for tumor-specific tcrs with no need for labor intensive t cell cloning. tcr genes can be easily and rapidly used to redirect t cell specificity against cancer cells by tcr gene editing. disclosure: chiara bonini: research funding from intellia therapeutics p car t cell therapy targeting relapsed or refractory cd + lymphoid disease with third-generation vector rv-sfg.cd .cd . - bbzeta maria-luisa schubert , anita schmitt , leopold sellner , , brigitte neuber , angela hückelhoven-krauss , kunz alexander , lei wang , gern ulrike , birgit michels , susanne hofmann , carsten mueller-tidow , , dreger peter , , michael schmitt , background: t cells genetically engineered to express chimeric antigen receptors (carts) directed against cd have demonstrated significant efficacy in patients with iymphoid malignancies including relapsed or refractory (r/r) b-lineage acute lymphoblastic leukemia (all) or r/r b-cell non-hodgkin's lymphoma (nhl). access to cart treatment for patients in europe has been limited so far given that the vast majority of cart trials have been performed in the united states and the p. r. of china. here we present the preliminary results of the first investigator-initiated trial (iit) cart trial in germany. hd-car- (eudract no. - - ; nct ) is a phase i/ii trial with in-house cart manufacturing which was initiated in september at the university hospital heidelberg. methods: adult as well as pediatric patients with r/r all and patients with chronic lymphocytic leukemia (cll) or nhl including diffuse large b-cell lymphoma (dlbcl), follicular lymphoma (fl) or mantle cell lymphoma (mcl) are treated with autologous t lymphocytes transduced with a third-generation car retroviral vector (rv-sfg.cd .cd . - bbzeta) targeting cd . the main purpose of hd-car- is to evaluate safety and feasibility of escalating third-generation car t cell doses ( - × transduced cells/m ) after lymphodepletion with fludarabine and cyclophosphamide. patients are monitored for cytokine release syndrome (crs), car-t-cell related encephalopathy syndrome (cres) and/or other toxicities. in vivo function, survival and anti-tumor efficacy of carts are assessed. results: to date, three patients (cll, dlbcl and mcl, respectively) have been enrolled and subjected to leukapheresis. high numbers of transduced carts were harvested on day of culture ( - x carts). transduction efficiency ranged between and %. cart products were sterile and free from mycoplasms and endotoxins. no production failure occurred and all patients received the cart product. no signs of crs or cres > grade have been observed. assessments of clinical responses are pending and will be presented at the conference along with updated technical results. conclusions: for hd-car- , gmp-conform leukapheresis as well as cart manufacturing was effective. administration, patient monitoring and follow-up were performed in-house providing independency from transport or production sites outside the university hospital heidelberg, altogether suggesting that academic cart iits are feasible in germany. clinical background: the prognosis of adult patients (pts) with relapsed/refractory (r/r) precursor b-acute lymphoblastic leukemia (all) is dismal, including with allogeneic hematopoietic stem cell transplantation (allo-hsct). blinatumomab, a bispecific cd -directed cd t-cell engager and inotuzumab ozogamycin (io), a cd -directed antibody-drug conjugate revolutionized the field, improving their outcomes. anti-cd chimeric antigen receptor t (cart) cell therapy has led to further progress and improved outcome (jacoby e; am j hematol, ). nowadays, patients with r/r b-all can be offered both therapies, but there are limited data on the safety and efficacy of cart -cell therapy post antibody treatment. we detailed our single center experience in this regard. methods: this report is a part of a single center, phase b/ study on therapy of b-cell malignancies with locally produced cart-cells (nct ). the approach uses autologous t cells with car construct that is composed of an anti-cd single-chain fv, cd co-stimulatory and cd -zeta intracellular domains. cd expression on the blasts was documented prior treatment in all pts by flow cytometry. all pts received x /kg cart-cells after lymphodepletion with fludarabin and cyclophosphamide. results: six pts ( males and females) with r/r b-all were enrolled, including one with ph-positive b-all. the median age was years ( - ). median number of prior therapy was ( ) ( ) ( ) . five pts had prior allo-hsct. four pts were given antibodies as the last therapy prior to cart cells. two pts received blinatumomab resulting in pr in one of them. two additional pts received io ( after failing blinatumomab) achieving mrdpositive cr. cytokine release syndrome occurred in all pts and was severe in only one patient who required tocilizumab treatment. this patient was also the only patient who experienced neurotoxicity (grade ), and was treated with dexamethasone. this patient eventually died days post infusion of cart cells due to severe pseudomembranous colitis, toxic megacolon and sepsis. all pts had prolonged neutropenia for a median of days ( - ) after the infusion of cart cells. at day after infusion of cart-cells the cr for the entire cohort was %: three pts with mrd-negative and one with mrd-positive response. among the four pts who received antibodies prior the cart-cells, one patient had mrd-positive and two pts had mrd-negative response. the patient with ph positive b-all had progressive disease during the treatment. two pts were referred to second allo-hsct from other donors. one patient with mrd-negative response relapsed after the second transplant and was treated by salvage therapy. the second patient with mrdnegative response demonstrated prolonged remission ( months) even without second transplantation. with a median follow-up of months ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) the median progression-free and overall survival for the entire cohort were . and months, respectively. conclusions: autologous anti-cd car t-cell therapy after debulking treatment with antibodies, including blinatumomab and/or io, is feasible and results in high response rates in pts with r/r b-all. patients may respond to anti-cd car t-cell therapy even after failure to their last salvage therapy with blinatumomab, which demonstrates similar mechanism of action. clinical background: genetically engineered t cells expressing a chimeric antigen receptor (car-t) targeting specific antigens present on acute lymphoblastic leukemia (all) blasts have generated promising results in children and adults with relapsed and refractory disease. the below report provides an insight of lineage switch occuring as a result of intense immunological selection after car-t cell therapy, even with a tumor clone that has no potential for this switch. methods: an eight year old caucasian male with precursor b (pb) cell lymphocytic leukemia was treated with cd directed car-t cell therapy in third remission, and after relapse after previous bone marrow transplantation (bmt) . he was diagnosed with t( ; ) pb cell all at years of age and treated with bfm protocol. he relapsed months after completion of maintenance therapy, and had a / mmud bone marrow transplant after etoposide, tbi and alemtuzumab conditioning therapy. he had cutaneous acute and chronic gvhd but months post-transplant, he relapsed again with pb cell all, with the same cytogenetic and immunophenotypic disease characteristics.. he was treated with lymphodepleting chemotherapy with fludarabine, cyclophosphamide and alemtuzumab followed by infusion of cd directed car t cells. he developed cytokine release syndrome of grade severity manifested as persistent fever, associated with car t cell expansion in the blood. after car-t infusion,there was no detectable b cell all clone in the marrow by pcr and the cytogenetics were negative for t( : ) translocation. months after the car t cell therapy, he was found to have a mrd positive disease which was monitored closely. results: we document clonal evolution from cd negative, mrd positive disease to aml, with the same ig rearrangement (the same clonal disease) but with complete myeloid phenotype mpo, cd , cd positive disease. there was cytogenetic evolution of the underlying clone but the original t( ; ) was retained within the evolved karyotype. sadly, our patient developed fludarabineneurotoxcity during an attempt to induce aml remission, and further curative-intent chemotherapy was not possible. conclusions: there are two case reports of mll rearranged b-all acquiring a clonally related myeloid phenotype associated with cd -negative escape after cd directed car t cell therapy,so far. but, this is the first post car-t cell therapy transformation of all to aml with etv -runx mutation, which is not recognised to have such lineage-switch potential. unlike mll, etv -runx translocation in the pathogenesis of acute myeloid leukemia is not been reported in the literature. the theory behind such transformation is an intense immunological selection of the tumor, driving it to myeloid differentiation with additional clonal cytogenetic events. disclosure: nothing to declare p ivac-all- : interim analysis of a phase i/ii clinical study on personalized peptide vaccination based on patient-individual tumor-specific variants in relapsed pediatric acute lymphoblastic leukemia armin rabsteyn , , christopher mohr , olaf witt , , roland meisel , , cristiane chen-santel , tobias feuchtinger , , christopher schroeder , jakob matthes , background: acute lymphoblastic leukemia (all) is the most common pediatric malignancy. standard chemotherapy is a successful treatment in % of patients, only about % develop a relapse, however these patients have a dismal prognosis. prevention of relapse after firstline chemotherapy or stem cell transplantation (sct) is therefore an urgent clinical need. we established a platform for the design of patient-individual peptide vaccination cocktails by combination of whole exome sequencing of tumor and normal tissue with in silico epitope prediction algorithms for individual patient hla types. we started clinical translation of this approach by starting a phase i/ii clinical trial in (nct ). besides feasibility and toxicity assessments, we aim to assess the capability of the peptide vaccination to induce neoantigen-specific t cell responses in high-risk all patients to target residual tumor cells and prevent leukemic relapses. methods: key inclusion criteria are: pediatric patients with all who suffered from second relapse after standard therapy or first relapse after sct. hematological remission has to be reached prior to vaccination. nonsynonymous mutations are identified by whole exome and transcriptome sequencing of patient leukemic blasts and healthy reference tissue. hla binding peptides harboring the altered amino acids are subsequently predicted in silico by algorithms syfpeithi, netmhc and netmhcpan for the patients' individual hla type. vaccine cocktails consisting of - individual peptides are produced and formulated under gmp conditions. the vaccination schedule is vaccinations over weeks using gm-csf and imiquimod as adjuvants. response to the vaccination is monitored by detection of t cells recognizing the vaccinated peptides occurring over time in peripheral blood of patients by prestimulation and intracellular cytokine staining. results: until now, patients were recruited, for of those, whole exome sequencing was performed to identify all-specific snvs using a comparative bioinformatics pipeline. we found an average of . mutations per patient on dna level. based on these data, an average of hla binders derived from neoantigens could be predicted per patient. an average expression of . % of mutations was assessed by rna sequencing. in all cases validated mutations could be identified and cocktail design was feasible. until now, patients received vaccinations. the vaccine was generally well tolerated and no or only mild side effects were observed. immune monitoring was performed for patients until now. in the first patient, we observed a transient cd + response against one vaccinated mhc class ii ligand and a sustained cd + response against the included wildtype control peptide derived from the antigen survivin. in the second patient, immune monitoring was performed for the first vaccination timepoints, a t cell response was not measurable at this timepoint of vaccination. conclusions: whole exome sequencing of pediatric all patients is feasible and yields small amounts of expressed, tumor-specific mutations. these few mutations are sufficient to predict hla-binding peptides that can be used to formulate individualized peptide vaccine cocktails. we currently conduct a clinical phase i/ii trial in a small cohort of high-risk all patients to assess safety, toxicity and immunogenicity. clinical background: chimeric antigen receptor t cells (cart) are considered as gene therapy medicinal products (gtmp) and genetically modified organisms (gmo). hence, carts manufacturing for clinical application is strictly regulated. appropriate methods assessing car transgene copy number (cn) in a cart product and definition of the frequency of carts in treated cart patient are mandatory. although quantitative real-time pcr-based (qpcr) analysis has been used for this purpose, no standardized procedure to minimize systematic errors and enable comparability has been established yet. here, we report on a single copy genebased (scg) duplex (dp) pcr (scg-dp-pcr) for the determination of the vector copy number (vcn) in cart products as well as patient samples following cart administration. scg-dp-pcr was validated and compared to the broadly used absolute copy number qpcr (acn) approach within the framework of a clinical trial treating patients with good manufacturing practice (gmp)-grade carts (hd-car- ). methods: for conventional acn, primers and probe targeting the car vector rv-sfg.cd .cd . - bbzeta were designed. standard curves were established via serial dilutions of the sfg.car plasmid. amplification of the standard curve as well as target genomic dna for vcndetermination was performed as singleplex (sp) pcr (sp-car) (method a). on the same qpcr plate, duplex (dp) qpcr reactions were carried out. additionally to the components comprised within method a, the experimental setup contained haploid human genomes as well as primers and probe targeting ribonuclease (rnase) p as human scg. the amplifications for the sfg.car plasmid (dp-car) and the rnasep gene (dp-rnasep) were performed simultaneously (scg-dp-pcr; method b). scg-dp-pcr was performed for standard curves and target samples. target-sample dna was extracted from carts prepared from leukapheresis products of three healthy donors (hd). results: for method validation, efficiency and linearity (correlation coefficient) of the qpcr reactions of method a (sp-car) and method b (dp-car, dp-rnasep) were assessed by linear regression of the pcr signal to the reference standard curve. overall, standard curves were only considered valid if a correlation coefficient (r ) of above . and efficiencies of % ± % were achieved. vcns applying method a and b to the same target sample were compared. sp-car pcr reaction displayed efficiency of . ± . %; . % ± . % and . ± . % efficiencies were achieved for dp-car and dp-rnase pcr reaction, respectively (table ) . applying scg-dp-pcr using formula for relative cn assessment ( -Δct (dp-car -dp-rnasep) ) on hd samples resulted in an average of . ± . increased cn when compared to method a (table ) . conclusions: in terms of efficiency and linearity by linear regression qpcr reactions were comparable. validation of scg-dp-pcr was achieved and represents an exact and less error-prone method to fulfil regulatory safety release criteria of cart products. besides of accurately assessing vcn of transduced cells, scg-dp-pcr is also a highly robust method to follow-up carts in treated patients. applying this approach, no standard curve is needed, hence significantly economizing required material as well as time. disclosure: nothing to declare background: t-cells' antileukemic responses in aml-pts need to be improved. dc leu effectively activate t-cells against leukemic blasts, resulting in blast-lysis ex-vivo. factors influencing these activities are not known. methods: we generated dc/dc leu from aml-blastcontaining mononuclear cells (n= ) using standard methods (mcm-mimic/ca-ionophore/picibanil/ifn-α, "mnc-methods") and from blast-containing heparinized whole blood (n= ) using modulatory kits (various combinations of - clinically approved response-modifiers, "wb-kits", patent ) and correlated statistically (t-/u-test, pearsons correlation, # means significant) proportions of dc-/t-cell-subtypes/cytokine-profiles with t-cells' antileukemic cytotoxicity (ctx), achieved after mixed lymphocyte culture (mlc) with/without mncmethod-("t*dc mnc "/"t*bla mnc ") or wb-kit-treated cultures ("t*dc wb "/t*bla wb "). ctx was given as proportions of cases with achieved or "improved" blast-lysis compared to control and as frequencies of viable blasts (bla via ) after effector-cell-influence. pooled data and data obtained with single methods in different cohorts are given. results: . generation of dc/dc leu : with a) mncmethods: Ø ± % dc and Ø ± % dc leu and b) wb-kits: Ø ± % dc and Ø ± % dc leu without induction of blasts' proliferation. t-cell-proliferation increased (vs uncultured t-cells) after mlc with a) mnc-methods: Ø ± % vs Ø ± % and b) wb-kits: Ø ± % vs Ø ± %. . antileukemic reactivity (t-effector-cell-cytotoxicity after mlc): pooling all data: a) mnc-methods ("t*dc mnc " vs "t*bla mnc "): we found a ) blast-cytotoxicity in Ø %(vs %) of cases with Ø %(vs %) bla via , a ) blast-cytotoxicity was improved (vs control) in % with Ø decrease of bla via of %; b) wb-kits ("t*dc wb " vs "t*bla wb "): we found b ) blastcytotoxicity in Ø %(vs %) of cases with Ø %(vs %) bla via , b ) blast-cytotoxicity was improved (vs control) in % of cases with Ø decrease of bla via of %. in general, these results could be confirmed with single methods: best mnc-methods were picibanil and mcm-mimic, best wbkits were kits containing gm-csf+picibanil or prostaglandins. . correlations: pooling all data: cases with "improved" lysis (vs "not improved" lysis) were characterized by a) mnc-methods: increased proportions of mature dc/cells (Ø ± % vs Ø ± %), dc leu /cells (Ø ± % vs Ø ± %) and proliferating t-cells (Ø ± % vs Ø ± %), b) wb-methods: dc/cells (r= . # ), dc leu /cells (r= . # ), dc leu /bla (Ø ± % vs Ø ± %), dc leu /dc (Ø ± % vs Ø ± %), cd + t-cells (Ø ± % vs Ø ± %), ifn-γ (r= . #) , mcp- ( ± vs ± pg/ml). conclusions: blasts are regularly converted to dc leu in the presence of mnc-methods and wb-kits (simulating the in vivo microenvironment). t-cells' coculture with dc/ dc leu after mlc induces and improves antileukemic t-cell activation compared to controls. blast-cytotoxicity correlates with proportions of dc/dc leu -and t-cell subtypes and released cytokines. these data support a role of antigen presentation by leukemic cells (dc leu ) for the stimulation of an immune response in aml in vitro and possibly in vivo. disclosure: nothing to declare evaluation year after the launch of the motion comic immuno-t, explaining patients and their caregivers how immunotherapy strategies work background: one year ago, the first version of immuno-t, a motion comic explaining to patients and caregivers how immunotherapy strategies work, was released. people were informed on and inspired to use the application during (inter)national meetings and events for the general public. meanwhile, the motion comic was further refined and adapted into a second version, based on the evaluations we've collected on the first version. adaptations included a multi-language tool (currently languages), increased user friendliness, and a new supporting musical score. also, a website was launched from which the second version could be downloaded on tablet or smartphone (both android and apple) and a new online evaluation form could be filled in. in months, people have evaluated the motion comic online, and these results are presented here, as well as our future plans within the immuno-t program. methods: through an online questionnaire, participants from belgium (n= ) and the netherlands (n= ) have evaluated the dutch version, and belgian participants evaluated the french version of the motion comic. results: the total group (n= ) consisted of patients (n= ) and their families (n= ), general public (n= ), students (n= ), health care professionals (n= ), researchers (n= ) and kindergarten teachers (n= ). participants' age ranged from to years, with an even distribution amongst the different generations. the majority of the evaluators ( , %, n= ) thought the motion comic is a good way to explain immunotherapy to patients. individuals ( , %) felt mainly interested after watching immuno-t, and a total of participants ( , %) felt hopeful or motivated. focussing on the patient group (n= ), all of the responders think the immuno-t motion comic is a good tool to use in a patient-doctor consultation. patients ( , %) felt hopeful and/or motivated after watching the whole motion comic, while of them ( %) felt combative and ( , %) felt gripped and intrigued. as for the new musical score, participants ( , %) think the music is suitable for the app, while evaluators ( , %) think the new music is not or not at all fitted to support the motion comic. conclusions: the detailed evaluations allow us to further improve immuno-t, and aid us in the development of other motion comics we plan to release under the cancer-t in motion umbrella. with the current version of immuno-t, a single-center pilot study is being set up, to test the efficacy and usability of immuno-t, based on qualitative research during the experience of the tool, and using validated questionnaires. with this study we want to evaluate the impact of immuno-t on patient empowerment, and the decision making process. the study protocol will be presented at ebmt. disclosure: the development of immuno-t was partly financially supported by celyad, calgene, novartis, roche, amgen, bms, but these companies did not by any means influence the contents and development of the motion comic. a therapeutic strategy to trespass the blood brain barrier for adoptive nk cell therapy in glioblastoma multiforme induced rat: a preclinical study background: glioblastoma multiforme (gbm) is among the most common and aggressive primary brain tumors with very poor prognosis. according to the central brain tumor registry of the united states, central nervous system (cns) tumors in pediatric patients (ages between - years old) are the second most common malignancies after blood-born malignancies, and the first amongst solid tumors, and known the most common cause of cancer-related deaths. although hematopoietic stem cell transplantation has been exploited to treat many kinds of malignancies, currently its success rate in gbm is limited. therefore, the gbm treatment paradigm needs shifting towards more effective treatments such as immune cell therapy. natural killer (nk) cells have been recognized as potential anti-cancer effector cells, as they can recognize and target tumor cells. since a small percentage of blood cells are differentiated as nk cells, the number of this group of cells is hardly enough to fight tumors, and so their multiplication and activation would be a potential effective cancer treatment. methods: this preclinical study was focused on setting up an optimal protocol for expansion and activation of naïve nk cells and assessing their efficacy towards induced gbm in rat models. ex-vivo expanded and interlukin- (il- )and heat shock protein- (hsp- )-treated nk cells have been exploited. after in vitro study and confirming the efficacy of treated cells through cytotoxicity assays, we induced gbm in male wistar rats (weighted - gr) using c tumor cells injection in rat brain through stereotactic surgery. the tumor formation was proven by mri imaging. following tumor establishment, we analyzed the effect of single injection of il- -and hsp- -treated nk cells compared with single injection of non-treated nk cells in two groups of rats. results: systemic intravenous delivery of il- -and hsp- -treated nk cells through tail's vein resulted in tumor shrinkage in different time intervals and complete remission in the first group of gbm-induced rat models, whereas in the other group of gbm rats receiving untreated nk cells, the tumor progressed. therapeutic efficacy of the treated nk cells was ascertained compared with non-treated nk cells considering tumor shrinkage observed in mri and survival rates between the two model groups. conclusions: the amelioration of tumor which has been confirmed by mri, proved the migration of activated nk cells through blood brain barrier and homing to cns, and finally targeting gbm tumor cells. our data suggest that nk-cells treated with il /hsp may be a promising immune cell-based therapeutic approach towards treating the fatal gbm. disclosure: nothing to declare p abstract withdrawn. long term sorafenib response for extramedullary flt + aml relapse after allogeneic stem cell transplantation since june , sorafenib dose has been tapered to mg/day, due to mild skin and gi toxicity. after years of treatment, she maintains cr at medullary and extramedulary levels, with no evidence of a disease that had escaped the mechanisms of action of chemo, hsct and dli. conclusions: in our patient, treatment with sorafenib has provided long-term control of this refractory extramedullary disease, even at adjusted doses. further studies are needed to confirm the efficacy of flt inhibitors in the control of relapses after allo-hsct, extramedullary disease and its potential role as maintenance agent. disclosure: nothing to declare background: although chemotherapeutic(ct) agents that used in the treatment of acute lymphoblastic leukemia (all) increase survival, the results are still weak. longterm survival with ct's in relapse all cases is difficult and the prognosis is very weak. inotuzumab ozogamicin is an anti-cd monoclonal antibody and it has the potential to reduce the overall toxicity of intensive regimens for all, as well as to possibly increase the number of patients who may achieve a state of minimal residual disease. methods: -year-old male patient was diagnosed with b-cell all in december .after the hoelzer kt protocol was started, maintenance treatment was continued. in the fifth month of treatment,flag ct protocol was started cause of recurrence was seen on % blast detection in peripheral blood smear. in august , inotuzumab ozogamicin treatment was started and six cures were completed because the patient was not in remission. in september , he had gone haploidentical bone marrow transplantation from his sibling donor( / )with defibrotid prophylaxis for veno-occlusive disease(vod)s. he engrafted succesfully and chimerizm was . % in th days of transplantation. he is th day of transplantation and in a remission. results: bone marrow transplantation cannot be performed since the complete response cannot be achieved in patients with relapse and resistant b-all.in these patients, new therapies targeting malignant lymphoblasts are needed. inotuzumab ozogamicinis a monoclonal antibody drug conjugate that targets cd antigen on malignant lymphoblasts.in many studies, it has been shown that inotuzumab ozogamycin is effective and reliable anti-tumor activity in adults with recurrent and resistant cd positive all. however, monoclonal antibody drug conjugates have been shown to be associated with vod's.for this purpose, we used defibrotid to protect our patient from vod. conclusions: treatment with combination ct regimens in b-all is suboptimal and long-term survival is achieved in only - % of patients.targeted molecular therapy and new regimens are needed in relapse and resistant patients.at this point, inotuzumab ozogamycin is an anti-cd- monoclonal antibody, as in our case, it provides remission in recurrent and resistant b-all patients and allows patients to complete their treatment with an allogeneic transplant from a fully compatible donor. disclosure: nothing to declare background: mesenchymal stem cells (mscs) are an attractive consideration for therapeutic cures of many difficult diseases on the cellular-level. due to the trophic effects of the cytokines and chemokines that they produce, mscs have shown multiple beneficial properties in the field of oncology. in this study, we will be investigating the effect of mscs derived from human bone marrow (bm), adipose tissue (at), and umbilical cord derived mscs (uc-mscs) on ovarian cancer. to differentiate the mscs, we performed a comparative analysis between the various sources for proliferative capacity, surface antigen expression, differentiation ability, tumor marker and paracrine activity, and their influence on ovarian cancer cell proliferation. methods: measurements of ovarian tumor marker proteins were computed by elisa. proliferative effects, immunomodulatory effects, and apoptosis of the mscs were measured by the culture and counting of colony formations. flow cytometry (fcm) was used to measure the variation of the immunophenotyping and cytokine secretions in co-culture, as well as gene expression. results: cells noticeably proliferated without any modifications to their immunophenotype during the third subculture. the colony-forming unit fibroblast (cfu-f) test showed a proliferation of the mscs along with healthy cells and cancer cell lines with no changes in their phenotype. the supernatant of mscs showed an increase in cellular death of the ovcar in ovarian cancer cell lines. a reduction in the level of ca- ( - %; p= . ) with ovcar in co-culture, and a decline of ldh ( - %; p= . ) and beta-hcg ( - %; p= . ) were observed in co-culture in caov , skov and igrov cell lines. a decrease in cd of the cancer cell lines in co-culture with the msc supernatant showed a reduction of the cancer tumorigenicity and aggressiveness, while the rate of the cd and cd asserted their stem state. msc supernatant decreased cell proliferation and mmp- , mmp- , and ca- mrna expression, while increasing timp , , and . this suggests that mscs have a role in cell death and inhibition of ovarian cancer cell proliferation. an increase of anti-inflammatory il- and il- cytokines, and a decrease in growth factor gm-csf along with their proinflammatory inf-a, tnf-a, il- , and il- a cytokines were also noted. conclusions: the gene and cytokine activity indicate a potential therapeutic anti-inflammatory and antiproliferative role of mscs on ovarian cancer despite their sources. the reduction of ca- , ldh, and beta-hcg in co-culture, along with the decrease in cd and amplified cellular apoptosis demonstrate the beneficial effects of stem cells in ovarian cancer cell lines. disclosure background: hematopoietic stem cell transplant (hsct) is the only cure in sickle cell disease (scd) so far. because of the risk of toxicity, its indication in france is restricted to severe patients with match sibling donor. this study compares the incidence of severe acute toxicity after hsct, between children aged less than years and teenagers aged to years old, treated for scd. methods: all patients suffering from scd, aged less than years at transplant, who received hsct in chu robert debré and necker, between / / and / / , were included. severe acute toxicity, defined by onset of severe acute gvhd, organ toxicity or infection, was compared between the two groups of age. results: patients ( children and teenagers) were included. all patients received a myeloablative conditioning regimen. bone marrow from a sibling donor was the main stem cell source (n= ; %). neither death nor rejection was observed with a median follow-up of . months (range, . the incidence of grade iii-iv acute gvhd was . % and was similar between the two groups; no risk factor was identified in univariate analysis. teenagers had more frequently acute skin toxicity ( . % vs %, p= . ). in univariate analysis, patients presenting severe organ toxicity were significantly older than others ( . vs . years old, p= . ). teenagers were more frequently treated for bacterial ( . % vs . %, p= . ) or bk virus ( . % vs . %, p= . ) infections. in univariate analysis, patients who developed infection were also significantly older at time of transplant (respectively . vs . years old, p= . ). no severe sinusoidal obstruction syndrome was observed. regarding long-term toxicity, patients presented an extensive chronic gvhd, they were both aged less than years old. no cut-off of age could have been defined. conclusions: this study confirms the excellent results of hsct in scd, with a -year event-free survival and overall survival of %. an older age at transplant seems to be associated with more frequent severe acute toxicity. these results are consistent with previous studies and suggest that hsct should be performed as soon as possible, without any defined "best age". prospective studies are needed, in order to define the place of each therapeutic in scd, with the aim of reducing treatment-related toxicity and developing alternative strategies for patients without match sibling donor. disclosure: nothing to declare p new insights into risk factors for transplant-associated thrombotic microangiopathy (ta-tma) in paediatric hsct (n= ) was associated with ta-tma in % vs . % vs . % respectively (p= . ). the presence of comorbidities at d (n= ) was significantly associated with an increased risk of ta-tma . % vs . % in absence of co-morbidity (n= ); p= . . use of csa/tac-based gvhd prophylaxis was associated with less ta-tma with an incidence of % vs % if these agents were not included (p= . ). in univariate analysis ta-tma was significantly higher among patients with agvhd grade ii-iv ( / ; . %) vs grade -i ( / ; . %) (p= . ). pres was recorded among cases and % of them developed ta-tma. two out of the patients with ta-tma had pathological gene mutations in their complement pathway. on multivariate analysis the presence of active comorbidity was a risk factor for ta-tma (or: . ; % ci: . - . ; p= . ) while the use of csa/tac-based gvhd prophylaxis did not increase the risk for ta-tma (or: . ; ci: . - . ; p= . ). in the presence of comorbidities the use of defibrotide as prophylaxis or therapy for vod (n= ) was associated with a drop in the incidence of ta-tma from % ( / ) in absence of defibrotide to % ( / ). -year overall survival was significantly lower among ta-tma cases ( %) in comparison to . % in absence of ta-tma (p= . ) (figure ). conclusions: active co-morbidity is a significant risk factor for ta-tma. use of defibrotide prophylaxis in patients with co-morbidities at the time of hsct might offer protection against ta-tma. surprisingly the use of csa/tac based gvhd prophylaxis is not a risk factor for ta-tma probably through limiting the development of high grades agvhd. the association between pres and ta-tma suggests a common pathway of endothelial damage background: gonadal impairment is a severe late effect of myeloablative conditioning regimes with significant impact on quality of life of cancer survivors. the aim of this study was to analyze gonadal function after busulfan (bu) or treosulfan (treo) containing regimens with regard to pubertal stage. methods: this was a retrospective, multicenter study involving patients treated in pediatric ebmt centers between - . patients receiving myeloablative doses of bu or treo as part of hsct conditioning were eligible for inclusion. analysis was conducted in two groups according to pubertal status at time of hsct. results: patients (pts) were treated in pediatric ebmt with bu or treo before allogeneic hsct. the median age at transplant was . years (range - ); / ( %) were males (m), / ( %) were females (f). / ( %) pts were pre-pubertal at hsct (f= ;m= ) and / ( %) were post pubertal (f= ;m= ). / ( %) patients received bu (f= ;m= ), / ( %) were pre-pubertal. / ( %)(f= ;m= ) received treo, / ( %) were pre-pubertal ( figure ). females who received treo in pre-pubertal stage (n= / ) reached more often spontaneous puberty ( % vs %; p= . ) compared to pre-pubertal bu group (n= / ) and occurrence of menarche was higher in treo group (p< . ) hormonal replacement therapy was given in / ( %) females transplanted in pre-pubertal stage and in / ( %) of those transplanted in post-pubertal stage. / ( %) males were pubertal at last follow-up and of them ( %) performed sperm analysis ( oligo-azoospermic, unknown). three pregnancies were reported in the population group, all received bu. regarding the evaluation of hormonal levels in pubertal patients at time of hormonal dosage (median . yrs) ( bu and treo), males treated with treo had significant lower lh levels (p = . ) compared to bu group. females treated with treo had significant lower levels of lh and fsh (p= . and p= . respectively). conclusions: gonadal damage related to treo was significantly lower compared to bu. we observed that: females transplanted during pre-pubertal period had spontaneous puberty more frequently after treo compared to bu and that hypogonadism hypergonadotropic was more frequent after bu than treo. these results must be further confirmed on a larger population. background: viral infections significantly contribute to both morbidity and mortality in patients undergoing hematopoietic stem cell transplantation. traditional antiviral therapy is associated with lack of efficacy, potential toxicity, prolonged hospitalization and increased patient costs. viral specific t cells can be manufactured from donor blood to treat viral infections post-transplant, and are associated with increased clinical efficacy and low toxicity. we postulated that direct costs of vst therapy are lower compared to traditional anti-viral medications methods: vsts were manufactured according to local protocols and fda requirements. total drug cost (as per institutional charges per drug) was calculated for patients who required treatment for viral infections post-hsct. manufacturing costs of vsts are fixed per fda requirements. patients who were treated with investigational antiviral medications (including brincidofovir) were excluded from analysis. patients treated with vsts +/anti-viral medications over a year period were compared to patients treated only using traditional anti-viral medications, including cidofovir, ganciclovir, valganciclovir, foscarnet and rituximab. results: demographics are shown in table . there were no major differences between the two groups treated. the number of anti-viral medications used in the vst group was lower compared to the anti-viral treatment group. median cost of vst treatment was significantly lower compared to those threated with traditional anti-viral therapy ($ , vs $ , , p-value= . ) . conclusions: treatment with vsts post-hsct for viral infections was lower in cost compared to anti-viral medical therapy. it is likely that overall costs are further reduced with vsts due to reduced inpatient hospital time, less monitoring of labs associated with anti-viral medication side-effects and reduced ancillary costs including nursing and pharmacy. more studies are needed to examine these indirect costs further. background: dock deficiency is an autosomal recessive primary immunodeficiency (pid) disease caused by loss-offunction mutations in the dock gene ( ) . patients with dock deficiency present with multiple abnormalities of the immune system, including defective t cell function and impaired production of antigen-specific antibodies. these lead to persistent viral infections of the skin, mucocutaneous candidiasis, recurrent sinopulmonary infections, atopic dermatitis, and other allergic disease, malignancies, and sometimes autoimmunity ( ). hematopoietic stem cell transplantation (hsct) is currently the only curative treatment option available ( ) . methods: we retrospectively evaluated our patients who underwent allogeneic hematopoietic stem cell transplantation due to dock deficiency in ege university pediatric stem cell transplantation unit between and . results: we identified patients transplanted at a median age of . years (range: - . years). of patients; (% ) received hsct from matched sibling, (% ) from unrelated donors and patient from haploidentical donor. we used busulfan-based myeloablative conditioning regimen to patients (% ), reduced toxicity myeloablative regimen with treosulfan to patients (% . ) and nonmyeloablative regimen to patients (% . ). eight of the recipients received bone marrow, of the patients received peripheral blood stem cells, of the recipients received cord blood as stem cell source. fifteen of patients (% ) had achieved engraftment and median follow-up of patients was months ( - ). grade iii-iv acute graft versus host disease (gvhd) occurred in % of patients and chronic graft versus host disease was seen % of patients. one patient received cord blood from unrelated donor did not engraft and died from septic shock. four patients died from transplant related toxicity. our patient's survival was % ; / patients alive. conclusions: hsct is the only curative treatment option for dock deficiency. in particular, patients with high comorbidity scores have a high risk of toxicity and toxic death. therefore, reduced toxicity conditioning regimens should be used for these patients. references : background: eltrombopag, a low-molecular-weight synthetic nonpeptide thrombopoietin receptor agonist (tpo-r), is a second-generation tpo. it is an oral thrombopoietin mimetic licensed in chronic immune thrombocytopenic purpura that induces platelet maturation and release by binding to c-mpl receptors on megakaryocytes. in a recent study; for patients with refractory saa, eltrombopag induced a response (at weeks) in at least one hematologic lineage in % patients and % no longer required platelet transfusions. and also % patients became rbc transfusion independent and % had a neutrophil response. trilineage responses were seen in % of patients; although surprising, this might indicate stimulation of c-mpl receptors on remaining stem cells. delayed recovery from thrombocytopenia is common after stem cell transplantation. in a study including adult patients, eltrombopag was used to enhance platelet recovery for post-hsct thrombocytopenia. it is well tolerated and efficacious offering transfusion independence. methods: in our retrospective study, eltrombopag ( mg/day) was started in pediatric patients (age ranging from to years with a median age . years) for posthematopoietic stem cell transplantation (hsct) thrombocytopenia. all patients fulfilled the following criteria: ( ) undergone hematopoietic stem cell transplantation (hsct), ( ) had improved total leucocyte counts after leucocyte engraftment, ( ) had prolonged thrombocytopenia (< . ) needing platelet transfusion. results: four of the patients have received ric while patients ma conditioning regimens before hsct. two haploidentic, autologous, mud, msd transplantations were performed. et ( mg/day) was started in patients who had thrombocytopenia despite neutrophil engraftment on the + th day of hsct a reduction in platelet transfusions and a platelet count of more than , were the primary endpoints. before et treatment, bone marrow biopsy was checked in / patients, / patients had decreased number of megacocyocytes. none of the patients had active bleeding at the start of eltrombopag but they were all at high risk of bleeding. according to the platelet monitoring, patients had a dose increase starting from the second week. the number of patients in need of platelet transfusions was at the end of the first month; and only at the end of the nd month. all patients had a thrombocyte count of more than . in the third month. in patients, et was discontinued after - months. no dose limiting toxicities have been observed. conclusions: as a conclusion, et was found highly effective for posthsct thrombocytopenia, with no drug related adverse effects. there was a gradual increase in platelet count, and none of the patients had any complication due to thrombocytopenia. disclosure background: isavuconazole (isa) is a new triazole approved for ifi treatment in the adult population. advantages are activity against both moulds and yeasts spp, excellent bioavailability after oral administration without relevant food or gastric ph effect, a water-soluble prodrug developed to facilitate intravenous administration without nephrotoxic excipients such as β-cyclodextrin, potentially poor drug-drug interactions. isa does not currently appear to require tdm. isa safety and efficacy have not been yet established in children and, in particular, no data are available in the pediatric hsct setting. methods: italian association pediatric hematology oncology (aieop) multicentric analysis of a cohort of allogeneic hsct pediatric patients who received isa as ifi treatment or prophylaxis. due to the lack of recommended dosing in pediatric patients and a clear target isa plasma trough-level range, the therapeutic monitoring (tdm) of isa concentrations was applied by a validated liquid chromatography-tandem mass spectrometry (hlpc-ms/ ms) assay technique. isa trough plasma concentrations (c ) and hours after drug intake (c h) were measured. results: a total of allo-hsct recipients were included, (m/f / ); median age: , years, range - , median body-weight , kg (range - ). isa was used as ifi treatment in cases and as prophylaxis in patients. donors were haploidentical in patients, matched-sibling in , allogenic-unrelated in cases. according to eortc criteria, ifi was proven in patients ( penicilum, mucor, aspergillus fumigatus), probable in and possible in patients. lungs were the main localization ( cases), associated with cns involvement in cases and paranasal sinuses in ; patient had possible hepatic candidiasis. all patients, but one, received isa as rescue treatment for previous therapeutic failure ( ambisome, voriconazole, combination therapy, posaconazole). seven patients received only iv isa, received only oral isa whereas patients received both iv and oral isa. patients under kg body weight received half isa dose ( mg tid loading dose on days and , mg/die manteinance). the others received adult schedule; only patients received loading dose. isa was administered for a median of days (range: - ). in patients isa was administered in combination with caspofungin. tdm was applied to patients including patient with severe vod and with renal failure secondary to ta-tma. the median isa concentrations were . ( . - . ) mg/l and . ( . - . ) mg/l for c and c h, respectively. ifi complete remission was achieved in cases, partial remission in ; treatment failure was experienced by patients. in cases fungal lesions remained stable. ctae grade ii-iii toxicity was observed during treatment in patients, with increased transaminase and/or creatinine levels which resolved after temporary isa withdrawal. no drug-drug interactions were observed in patients receiving csa as gvhd prophylaxis and no modification of csa daily dose was needed. conclusions: isavuconazole use may be considered in the pediatric population, even in the hsct setting, for its safety, efficacy, tolerability, no drug-drug interaction. of course these data deserve further evaluation. disclosure: nothing to declare p new treosulfan-based conditioning regimens including epigenetic agents in patients with very high-risk neuroblastoma background: the pts aged mos. or older with disseminated nb involving bone and bone marrow constitute a group of pts with very poor prognosis. although the majority of them are responsive to intensive conventional chemotherapy, most eventually relapse with efs at years of < %. at the beginning of the year we came up with a protocol for this very unfavorable group including epigenetic therapy ( -azacitidine) in the phases of consolidation. methods: seven pts with a median age of ( , - ) years completed the protocol and received hdct with autologous sct as a consolidation. hdct included two different epigenetic agent containing regimes according to tumor response to the induction therapy assessed by i-mibg and mri (ct-scan). three pts revealing large active residual tumor assessed by i-mibg scan or multiple active bone metastases received a conditioning regimen (regimen a) including i-mibg therapy at a dose . - . mbe/kg on d- , treosulfan mg/m /d, on d- ,- and - (total mg/m ), melphalan mg/m /d, on d- ,- (total mg/m ), -azacitidine mg/m /d on d- to d- (total mg/m ). four pts with cr or vgpr received a «split» conditioning regimen (regimen b) including treosulfan mg/m /d, on d and - , and on d- and d- (total mg/m ), melphalan mg/m /d, on d- and - (total mg/m ), and -azacitidine mg/m /d, on d - to d - and on d- to -d- (total mg/m ). a median number of . ( . - ) cd +/kg was infused on d . results: the median recovery times to wbc> . x /l and to an unsupported plt> x /l were ( - ) and ( - ) days, respectively. all pts experienced grade hematological as well as infectious toxicity assessed by nci-ctc score. there were episodes of severe organ toxicity of grade occurring in pts. in cases we observed a severe mucositis, in cases gi toxicity and episode of the erythema multiforme occurred. one pt revealed a lifethreatening episode of hypotension of grade . no transplant-related death occurred. the median number of transfused rbc and plt doses was ( - ) and ( - ), respectively. all pts are alive and well without signs of disease progression in complete hematological recovery with a limited follow-up of . ( - ) mos. from day of hdct. conclusions: although it is rather early to evaluate the efficacy of the epigenetic agent's inclusion in the induction and/or consolidation phases of a very high-risk nb treatment, we can assume that, first, the hdct combining mibg i and/or high dose of treosulfan with epigenetic agent such a -azacitidine was feasible and had an acceptable toxicity. second, the "split" modality of the treosulfan use in conditioning regimen would permit to increase the total dose of the alkylating agent with no inacceptable toxicity. disclosure: nothing to declare pre-and-post magnetic resonance imaging of hips and knees for detecting osteonecrosis in children undergoing hematopoietic cell transplantation: in whom is it necessary? ali y suliman , sue c kaste , ying li , dinesh keerthi , guolian kang , brandon m triplett , ashok srinivasan between april and august at the university medical center hamburg-eppendorf, germany, were included. total and active ratg plasma levels were analyzed by elisa and flow cytometry, respectively. primary endpoint of the study was exposure to ratg. secondary endpoints included transplant-related mortality, incidence of acute and chronic gvhd, immune reconstitution, chimerism, rejection and viral infections. patients were monitored at least days post transplantation. statistical analyses were performed using ibm spss statistics software, or graphpad prism software. results: median total grafalon™ and thymoglobuline™ peak plasma levels were . μg/ml and . μg/ml, respectively; median active grafalon™ and thymoglobu-line™ peak plasma levels (appl) were . μg/ml and . μg/ml, respectively. active thymoglobuline™ plasma levels showed highly variable pharmacokinetics compared to grafalon™. neither grafalon™ nor thymoglobuline™ exposure correlated with lymphocyte count prior to transplantation, cell count in the graft (wbc, mnc, t cells), age, body weight or body surface area (bsa). this is indicative for a saturation effect in both groups. to correlate high or low ratg exposure with clinical outcome parameters, we built two groups within each patient cohort by median appl. the incidence of gvhd was not dependent on high or low ratg exposure. until day post hsct, viral infections or reactivations (ebv, cmv, adv, hhv , bkv) occurred in the patients. interestingly, adv infections affected only children with high ratg exposure. the median time to leukocyte engraftment was not significantly longer in the high ratg groups compared to the low ratg groups ( to days for grafalon™, and to days for thymoglobuline™). there was a decreased and/or delayed recovery of cd + , cd + and cd + t cell reconstitution, but not of b cells and nk cells in the high thymoglobuline™ exposure group compared to the low thymoglobuline™ exposure group. overall survival was not statistically significant with % in the grafalon™ and . % in the thymoglobuline™ group without influence of ratg exposure. conclusions: high and low exposure to grafalon™ or thymoglobuline™ did not result in significant differences in outcome parameters as incidence of survival, agvhd, cgvhd, rejection, or mixed chimerism in this limited cohort. delayed and decreased immune reconstitution in the high ratg exposure groups did not translate into different clinical outcome parameters. adv infections only occurred in the high ratg exposure group. grafalon™ and thymoglobuline™ showed distinct pharmacological and immunological differences in children and larger cohorts are needed to detect clinically significant differences and adjust dosing regimens individually. disclosure: nothing to declare background: the optimal conditioning regimen for allogeneic hematopoietic cell transplantation (allohct) in children with myeloid malignancies remains undefined, particularly when reduced-intensity conditioning (ric) regimens are utilized. methods: we performed a retrospective review of children undergoing allohct for acute myeloid leukemia (aml) and myelodysplasia-related aml (mdr-aml) over a year period ( - ) at our institution, comparing the outcomes of those who received either a busulfan (bu)-or melphalan/thiotepa (mel/thio)-based conditioning regimen. results: a total of patients were analyzed. twentyone received fludarabine/melphalan/thiotepa and received myeloablative busulfan-based conditioning, either in combination with cyclophosphamide (n= ) or fludarabine (n= ). atg was used in patients depending on donor. recipients of mel/thio were selected for ric regimens due to pre-transplant comorbidities (cardiac dysfunction, n= , requiring peri-transplant milrinone), transplant during chemotherapy-induced aplasia (n= ), underlying diagnosis of treatment-related aml (t-aml) and significant pre-allohct chemotherapy exposure (n= ). proportions of patients with de novo aml (mel/ thio, %; bu, %) and mdr-aml ( % and %) were similar between groups; however, recipients of mel/thio were more likely to have t-aml ( % vs %). cytogenetic and molecular risk factors were similar between groups. the majority of patients were transplanted in cr (mel/thio, % vs bu, %) or cr ( % vs %). more recipients of bu conditioning ( % vs %) were mrd-negative at the time of allohct; both groups had comparable proportions of patients with ≥m marrow (~ %). donor types and stem cell sources were similar between groups, except unrelated umbilical cord blood which was more common in bu recipients ( % vs %). there were no graft failures in mel/thio recipients, compared to % (n= ) in those receiving bu-based regimens. engraftment kinetics and immune reconstitution were similar. overall acute and chronic gvhd incidence was higher in recipients of mel/ thio compared to bu ( % vs %, and % vs %, respectively), but rates of grade iii-iv acute or extensive chronic gvhd were comparable. vod requiring treatment was diagnosed in ( %) recipients of bu conditioning and no mel/thio recipients. median duration of follow-up was months (range - ) in the mel/thio group, and months (range - ) in the bu group. transplantrelated mortality (trm) was similar in both groups ( patient), occurring before day . relapse incidence was comparable (mel/thio, % vs bu, %); however, relapse occurred at a later time in mel/thio recipients (median d + vs d+ ). overall survival at and years was superior in mel/thio recipients ( % vs %, and % vs %, respectively). conclusions: in our single institution experience, use of a melphalan/thiotepa-based ric regimen was associated with similar outcomes compared to full-intensity bu-based conditioning, despite higher risk patient and disease characteristics. the majority of recipients of mel/thio conditioning had significant pre-transplant comorbidities, which did not translate into higher trm. while mel/thio recipients had less optimal leukemia control at the time of transplant and high-risk leukemia features (e.g. t-aml), relapse was similar between groups, occurring later in mel/ thio recipients, which may have contributed to better overall survival. disclosure: nothing to disclose methods: pubertal development and biological gonadal parameters were assessed in a retrospective monocentric cohort of pre-pubertal patients who underwent hsct after myeloablative conditioning with total body irradiation (tbi) or busulfan between and . results: seventy-four patients ( girls and boys) were included. no spontaneous pubertal development was found in % of girls and % of boys (p < . ) and delayed puberty or no spontaneous pubertal development was found in % of girls and % of boys (p= . ). hormone replacement therapy was used in % of girls and % of boys (p < . ). in univariate analysis, tbi conditioning (p= . ), female sex (p < . ), acute gvhd (p= . ), extensive chronic gvhd (p= . ), steroid treatment > months (p= . ), and malignant diseases (p= . ) were associated with no spontaneous pubertal development, whereas tbi conditioning (p= . ) and extensive chronic gvhd (p= . ) were associated with delayed puberty. in multivariate analysis, factors independently associated with no spontaneous puberty onset were female sex (p= . ) and age > years (p= . ). factors independently associated with delayed puberty were extensive chronic gvhd (p= . ) and age > years (p= . ). tbi was not an independent risk factor for pubertal complications. conclusions: this study confirms the toxicity of myeloablative conditioning on pubertal development and the role of older age and female sex in increased pubertal issues, and suggests a possible role of gvhd in delayed puberty. disclosure: nothing to declare p abstract already published. neutrophil elastase activity may serve as a marker for neutrophil extracellular traps formation following stem cell transplantation ronit elhasid , sivan berger-achituv , hila rosenfeld-keidar , szilvia baron tel aviv sourasky medical center -tel aviv university, tel aviv, israel background: post-transplant infections rise dramatically in patients with quantitative or qualitative neutrophil defects and constitute a major source of morbidity and mortality following hematopoietic stem cell transplantation (hsct). neutrophils protect the host from microorganisms via multiple processes including phagocytosis and formation of neutrophil extracellular traps (nets). although reactive oxygen species (ros) production seems to be essential for nets formation, the key enzymes of the process are neutrophil elastase (ne) and myeloperoxidase (mpo). methods: ne and mpo activity as well as nets formation were investigated following hsct in patients at week to and after neutrophil engraftment. neutrophils were isolated using easysep direct human neutrophil isolation kit (stemcell technologies inc.) by immunomagnetic negative selection. enzymatic activity of ne and mpo were measured using colorimetric assays. nets formation of phorbol -myristate -acetate (pma)activated neutrophils was investigated by confocal fluorescence microscopy. all results were compared to those of healthy volunteers. statistical significance was calculated using one way-anova with bonferroni post hoc test. results: patients (median age of . years [range - years]) were investigated, following allogeneic hsct ( acute lymphoblastic leukemia, acute myeloblastic leukemia, epidermolysis bullosa, rhabdomyosarcoma) and following autologous hsct ( ewing sarcoma, desmoplastic small round cell tumor). all patients experienced fever and neutropenia. at engraftment, average ne activity was significantly decreased compared to the average value of healthy individuals. ne activity improved week by week in patients, reached the lower reference range at weeks following transplantation (fig. a) and continued to increase. the enzymatic activity of mpo was comparable to the average value of healthy individuals (fig. b) and showed no significant difference between the distinct time points. at neutrophil engraftment, nets formation was absent and comparable to those of non-activated neutrophils (fig. c) . although nets formation increased week by week, it did not reach the average of normal controls during the monitored time period. also linear correlation between ne activity and nets formation (r = . ) was demonstrated. conclusions: impaired ne activity following hsct corresponds to decreased nets formation and could serve as a marker for netosis. strategies to accelerate the recovery of ne function post transplantation might improve nets formation and thereby induce better infection control. a) the average of ne activity (n= ) during weeks following hsct. reference range was measured and calculated from measurements of healthy volunteers using. b) the average of mpo activity (n= ) during weeks following hsct. reference ranges were measured and calculated from measurements of healthy volunteers using the quartile method. c) the average of netosis activity after nm pma activation for h (n= background: to have a better understanding of incidence, treatment, outcome and risk factors of immune cytopenia in children after allogeneic hsct. methods: between january and september , pediatric allogeneic hsct have been performed in patients at the ghent university hospital (ghent, belgium). autoimmune hemolytic anemia was defined by a positive direct agglutinin test (dat). dat was performed at moment of engraftment and in case of hemolysis or unexplained anemia. platelets antibodies were evaluated in case of no otherwise explained thrombocytopenia. results: the cumulative incidence of post allo sct autoimmune cytopenia is . % ( / ). in cases there were positive antibodies against red blood cells, and one patient against had antibodies against platelets. of these cases, only ( . %) were clinically relevant and needed treatment. the median observation period post sct for the whole cohort was months ( - ) . the clinically significant immune cytopenia started at a median time of day+ and day + in the group without symptoms. the patient who presented the autoimmune thrombopenia developed antibodies against anti-gpiib/iiia, this was resolved after days, the treatment consisted intravenous immunoglobulins (ivig). two of the patients with autoimmune hemolytic anemia had igg mediated antibodies, and had complementmediated dat. these patients were treated with ivig, steroids, rapamune and rituximab. one patient has still dat positive after months, but clinical stable. the other two are also dat positive and have some hemolysis, but the follow up is much shorter ( months). treosulfan-contained conditioning regimens were more frequently used in patients with significant immune cytopenia. conclusions: immune cytopenia is an infrequent complication after allogeneic hsct. however, its treatment can be challenging, and the hemolysis can persist for years. the association of rapamune and rituximab was adequate to treat this problem in our patients. background: approaches to the management of refractory and relapsed classical hodgkin´s lymphoma (r-r chl) are changing and become more effective. the role of anti-cd targeted immunochemotherapy with brentuximab vedotin (bv) has been extensively investigated in adults with r-r chl and is only to be elucidated in children. the study included children and adolescents with r-r chl that were sucessfully treated with bv-based therapy prior to hematopoetic stem cell transplantation (hsct). median age of patients was years ( - ), main histological variant -nodular sclerosis ( %, n= ), advanced stage at diagnosis - % (n= ). most were heavily pre-treated (median number of previous therapies - ) and progression after autologous hsct was documented in ( %). refractory disease was diagnosed in ( %) and relapsed in ( %). among relapsed patients ( %) were with multiple episodes, ( . %) -early and ( . %) -late relapse. treatment regimens consisted of bv in monotherapy . mg/kg triweekly (n= ) or combination of bv . mg/kg on day with bendamustine - mg/ m on days and of -week cycles (n= ) or combination of bv . mg/kg on day with dhap (n= ). median number of bv infusions was . ( - ). all selected patients achieved complete (n= , %) or partial remission (n= , %) prior to hsct. consolidation with autologous hsct was performed in ( %) and with allogeneic hsct -in ( %). primary end points were overall (os) and progression free survival (pfs). response to bv was not assessed in the study as only responders to the bv-based treatment were included. results: with median follow-up of days ( - ) os and pfs for all patients are % and %, respectively. pfs after autologous hsct and allogeneic hsct are % and %, respectively (p= . ) at present moment ( %) patients are alive and are in remission. three patients died ( %): disease progression (n= ), postransplant idiopathic pneumonia syndrome (n= ) and posttransplant pneumonia (n= ). bv was generally well tolerated with only mild polyneuropathy in patients ( %) as the main reversable documented adverse event. conclusions: in prognostically unfavourable heavily pretreated children and adolescents with r-r chl achievement of response to bv-based therapy prior to hsct is assosiated with promising rates of os and pfs. disclosure provides a treatment by restoring thymidine phosphorylase function and improving disease manifestations. here we report the outcomes of affected siblings who underwent transplantation using an unaffected sibling donor to highlight important experiences in the transplant of such a rare condition. methods: four siblings of consanguineous pakistani descent aged , , and years underwent myeloablative hsct using fully hla-matched ( / ) peripheral blood stem cells harvested in a single apheresis from an unaffected year old sibling. the oldest sibling, a year old male, first presented in having emigrated from pakistan with a history of growth failure and abnormal movements. biochemical, nerve conduction and imaging studies confirmed a diagnosis of mngie. testing on three other siblings identified similar biochemical abnormalities, though the youngest children had minimal clinical manifestations of the disease. based on the progressive nature of the disease and the availability of a fully matched donor, a decision was made to pursue transplant for all affected siblings. results: due to the severity of their disease, the oldest siblings were transplanted first using a myeloablative conditioning regime of fludarabine, thiotepa and treosulfan with alemtuzumab. neutrophil engraftment occurred on day + for both, with % donor chimerism achieved. there were no significant transplant related complications. the post-transplant course of the year old sibling was complicated by a major stroke-like event characterised by dramatic imaging changes and requiring ventilation, though no cause was identified and the patient's neurologic deficits have since resolved. gastrointestinal symptoms have persisted and both remain tpn dependent, though symptomatically have shown gradual improvement. following the neurologic complications in their older sibling, the younger siblings were conditioned with auc-targeted busulfan and fludarabine plus alemtuzumab. neutrophil engraftment occurred on day + , with full donor chimerism achieved. progression to enteral feeding has been much more rapid, with nutrition now fully enteral for both. there were no significant transplant related complications. conclusions: stem cell transplantation represents the only curative option for mngie. due to its rarity and relative infancy as a condition, little is known of the expected course following transplant or the best approach to transplantation itself. despite previous challenges with graft failure in mngie recipients, we were able to gain rapid and sustained donor engraftment using different myeloablative conditioning regimes with minimal transplant-related morbidity and no mortality. in keeping with previous reports, resolution of established gastrointestinal symptoms has been slow, though the siblings transplanted earlier in their disease course have shown more rapid improvement supporting the role of early recognition and access to transplant. it is essential moving forward that specialised transplantation centres collaborate so as to guide clinicians in the management of such a challenging condition. disclosure: there are no conflicts of interest to disclose. g pc congenital neutropenia -biology of inflammatory colitis associated with gcsf use, and disease response to allogeneic transplant, a report of cases background: an autosomal recessive disease, glucose- phosphatase catalytic subunit (g pc ) deficiency is a relatively recently identified cause of chronic severe neutropenia. there can be a spectrum to the disease and patients may also present with non-haematological features including prominent chest veins, cardiac, endocrine or urogenital abnormalities. we describe in our patient cohort a response to gcsf but an inflammatory, incapacitating, biopsyproven colitis associated with that g-csf response. we have transplanted children with such colitis, and describe a similar colitis with intestinal failure in a fourth. methods: we investigate the biology of the neutropenic colitis, and demonstrate necrosis of the stimulated neutrophils. in vitro studies demonstrated that unstimulated neutrophils from patients with g pc d exhibited significantly increased production of il , reactive oxygen species (ros) and neutrophil extracellular traps (nets) alongside significantly higher expression of cd b, cd b and cd . in contrast, neutrophils from patients with g pc d produced significantly less ros, mmp- , neutrophil elastase and nets upon stimulation. neutrophils from patients with g pc d also exhibited significantly accelerated apoptosis and secondary necrosis which was exaggerated upon stimulation with live escherichia coli bacteria but could only be partially rescued with supplemental exogenous glucose. results: patients have undergone hsct for g pc neutropenic enterocolitis ( unrelated donor and msd) after fludarabine treosulfan and thiotepa conditioning therapy. alemtuzumab was given as as serotherapy. all patients are alive and well, immune suppression has been discotniuned and there is no gvhd with normal organ function, and resolution of colitis. we describe a th patient with no good donor who has continuing intestinal failure with g-csf use. conclusions: we describe the aetiology of intestinal inflammation and failure with an extensive study of neutrophil biology in this metabolic neutropenia. we describe a novel indication for hsct in this "g-csfresponsive neutropenia". disclosure: nothing to declare p does body mass index (bmi) pose a risk to outcome for pediatric non-infantile patients undergoing hematopoietic cell transplantation (hsct)? mona al-saleh , khawar siddiqui , amal al-seraihy , abdullah al-jefri , ali al-ahmari , hawazen al-saedi , awatif al-anazi , mouhab ayas , ibrahim al-ghemlas with no evidence of toxicity. as benefits of stoss therapy in hsct remain unknown, and safety has yet to have been studied extensively in the pediatric population, we hypothesize that stoss therapy is an effective and safe method to reach and attain sufficient levels of vd in pediatric patients undergoing hsct. methods: this is an ongoing prospective, randomized clinical control trial at phoenix children's hospital that commenced december st , . following consent, subjects are randomized to the intervention (stoss) or control arm prior to hsct. stoss therapy consists of a single oral dose of vd (ranging , iu - , iu), given based on baseline -hydroxyvitamin d [ (oh)d] level and age, followed by standard weekly supplementation. subjects enrolled on the control arm receive standard of care based on endocrine society guidelines of weekly vd supplementation. data collection includes demographics, (oh)d levels at baseline, day + , and day + , vd toxicity (hyperphosphatemia, hyperkalemia and renal calculi), as well comorbidities were collected. at each time point and for each trial arm, the mean (oh)d level and changes from baseline were computed with corresponding % confidence intervals (cis) to indicate variability. results: presently, subjects have completed baseline assessment, with day + and day + follow-up completed for and of these, respectively. at baseline, the mean ( % ci) (oh)d was . ng/dl ( . , . ) among stoss patients and . ng/dl ( . results: total hrqol scores of transplanted patients were significantly improved compared to those on supportive care and also compared to healthy siblings (p < . and . respectively), the same was true for physical (p < . and . respectively) and emotional functioning (p < . and . respectively). social and school functioning of transplanted children were not different from healthy siblings (p . and . respectively) while were very significantly improved compared to children with st on supportive care (p < . in both cases). conclusions: bmt in a lower-middle income setting may be even more impactful compared to high-income regions. our analysis clearly indicates normalization of hrqol in all major areas of children transplanted for st. a possible resilience effect was noted for physical and emotional scores which were improved compared to healthy sibling controls. we could not however quantify the effect of longer-term issues like fertility impairment after bmt which may eventually adversely impact hrqol, particularly in the indian culture. disclosure: none allogeneic hematopoietic stem cell transplantation in ataxia telangiectasia patients without malignancy background: ataxia telangiectasia (a-t) is a primary immunodeficiency with mutations in atm-gene. besides a slowly progressive neurodegenerative course, a-t leads to increased susceptibility to malignancies which affects % of patient (median: . years) with a high mortality mainly due tocomplications of conventional radio-chemotherapy. the incidence of cancer correlates with the extent of immunodeficiency. patients often develop severe progressive granulomatous skin disease with evidence of vaccine-strain rubella-virus in the lesions. prolonged survival, neurologic improvement and malignancy prevention was observed in atm-deficient mice after treatment by syngeneic hsct. nevertheless, pre-emptive hsct is not routinely performed in a-t patients due to concerns about neurodegeneration and toxicity. methods: we present three a-t patients with severe immunodeficiency phenotype, undergoing successful hsct as an individual treatment strategy intending to restore immunodeficiency for long-term malignancyprevention (patient- ) and to treat progressive skin/joint granulomas (patients- and - ). results: patient- underwent a reduced intensity conditioning (ric) regimen at years of age including fludarabine ( mg/m ), cyclophosphamide ( mg/kg), and atg-fresenius ( mg/kg/d) which was tolerated well. hematopoietic engraftment occurred by day + . there was an expansion of naïve and memory cd + t-cells and cd + cells. while initially a mixed donor chimerism in patient's pbmcs ( - % donor) was observed, patient's tcells (cd + ) reached over % of donor origin over time. at last follow-up ( years) he is well, without signs of gvhd and organ toxicity, off immunosuppression with normal levels of atm-protein; his granulomas resolved. patient- is a year-old male who was transplanted from his hla-identical sibling, conditioned with fludarabine ( mg/m²), cyclophosphamide ( mg/kg), and atg-fresenius ( mg/kg). hematopoietic engraftment was observed by day + . t-cell reconstitution started by day + with > μl cd + t-cells. his mixed chimerism rapidly turned to donor origin ( % donor cd +) over time. there was no acute toxicity, however, he developed lumbosacral pain episodes with evidence of urine bk-virus with spontaneous remission. an intermittent metapneumovirus associated pulmonary hypertension was observed with pericardial effusion. treatment included sildenafil and oxygen. at last follow-up ( months) patient is well without immunosuppression. patient- suffered from recurrent chest infections, failure to thrive and progressive and debilitating rubella positive progressive granulomas of the skin. she received allohsct from an hla-identical family donor at years of age. conditioning included busulfan ( . mg/kg), fludarabine ( mg/m²), cyclophosphamide ( mg/kg), and alemtuzumab ( x mg/m², x mg/m²). hsct was complicated by intermittent acute renal failure, cmv reactivation and tma. hematopoietic recovery was observed by day + . t-cell chimerism increased rapidly over time (> % donor). at last follow-up ( months) patient is well, off immunosuppression and ivig. her skin granuloma resolved with scarring residues. conclusions: pre-emptive allohsct is feasible in a-t when reduced intensity conditioning is used and can correct the immunodeficiency. it might be a treatment option for some a-t patients at high risk of hematological malignancy and severe granulomatous skin disease. to what extent the restored immune system and the increase of atm-protein in these patients could prevent the development of other malignancies needs to be evaluated further. disclosure: nothing to disclose p abstract withdrawn. hematopoietic stem cell transplantation in diamond blackfan anemia: brazilian experience background: diamond-blackfan anemia (dba) is a rare inherited red cell aplasia caused by an intrinsic defect of erythropoietic progenitors. the main therapeutic approach is based on repeated red blood cell transfusions and/or corticosteroid therapy. hematopoietic stem cell transplantation (hsct), a potentially curative treatment for dba, is indicated for patients that do not respond to first-line therapy. methods: the aim of our retrospective study is to report the outcomes of brazilian dba patients transplanted between and in bmt centers. the median age of the patients was ys (range - ) and % were male. seventeen patients ( %) were transplanted with matched related donors (mrd) and thirteen ( %) from matched and mismatched unrelated donors (mud/mmud). in the mrd group all patients received bone marrow as hsc source, while in the mud/mmud, eight patients received bone marrow and five received cord blood. all patients with incompatibilities (mismatched) were ucb ( / ). nineteen recipients were conditioned with busulfan plus cyclophosphamide, while the remaining received fludarabine and busulfan, which has been the preferred regimen in brazil in the recent years. after transplant, most (n= ) of the mrd and mud recipients received cyclosporine and short course methotrexate as graft versus host disease (gvhd) prophylaxis. results: twenty-two out of the patients were alive and disease-free at a median follow-up of months (range to months). the -year overall survival (os) was % (ci - %) (fig ) . similar results have been demonstrated in studies from europe and from the united states. when analyzed according to donor type, os was % (ci - %) and % (ci - %) in mrd and mud/mmud respectively (fig ) . three out of the patients who were transplanted with ucb died. these results are in agreement with those of previously published data showing worse results in unrelated ucb transplants. twenty-nine out of the patients engrafted successfully. in of the evaluated patients, the median time to neutrophil engraftment was days (range - ). one patient experienced an early death from hemorrhagic shock on day , before neutrophil recovery, and another two patients experienced primary graft failure. post-transplant chimerism was available for patients. sixteen had complete chimera (> % chimerism), while patients presented with mixed chimerism. acute gvhd was observed in patients ( %), of which classified as grade iv. five patients developed chronic gvhd, considered severe in three of them. eight patients died at a mean of days (range - days) after hsct and the main causes of death were infections and hemorrhagic disorders. conclusions: hsct is a potentially curative treatment option for dba. in the present study, we report the outcomes of patients with dba transplanted in brazil with a os of %, with better results in mrd compared to mud, as expected. despite the small numbers, we observed lower survival after mud/mmud ucb transplantation. since dba is a rare disease, international collaborative studies are essential to better understand the benefits of the hsct in the treatment of these patients. disclosure: nothing to declare p treatment of the obliterant bronchiolitis in pediatric allogeneic recipients: two periods compared results: in group , the therapies administered for bo included prolonged treatment with steroids in all patients, anti-tnf in , azatioprine in ; while in the group , all patients received ima, montelukast and azitromicin, and received i.v. mpd. the median duration of imatinib therapy was years ( . - . years). after a median follow-up of . years (range . - . yrs), / patients of group ( %) died with bo in progress for transplant-related causes. while in the group , / ( %) died in presence of worsening bo. the estimated os at year after hsct was % ( % ci; - ) in group and % ( % ci, - ) in group (p= . ) (figure ), while the os after year decreased at % ( % ci; - ) in the group while remained stable in the group . conclusions: this experience shows a relevant improving in prognosis of children with bo with the use of this protocol including ima, since the significant improving of survival obtained, confirming as reported in adult populations. disclosure results: we presented patients with pres, age ranging from months to years with a average of . years. there were ten patients with thalassemia major, two patient with acute lymphoblastic leukemia, three patients with sickle cell disease and one patient with myelodysplastic syndrome, one patient with immune deficiency, two patients with acute miyeloid leukemia, one patient with aplastic anemia. ten patients were males, ten were female. all patients were treated with csa or tacrolimus and metilprednisolone for the prophylasix of gvhd. pres occurred at a median of days (range - ). clinical findings at onset of leukoencephalopathy were hypertension, headache, seizures, visual disturbance, and altered mental function. eighteen patients alive with normal neurological status. mri showed abnormalities in all patients including patchy bilateral cortical and subcortical lesions, especially in parieto-occipital lobes. conclusions: bmt is associated with several neurological complications that may be underlying diseases, bmt procedure, and severe immunosupression. pres is an uncommon but serious complication after bmt. we report cases of pres who received allogeneic bmt for thalassemia major to emphasize the importance of early recognition and institution of appropriate management of pres during bmt. disclosure: nothing to declare p continuous complete molecular remission using three different monoclonal antibodies followed by allogeneic bone marrow transplantation in an infant with chemotherapy-refractory acute lymphoblastic leukemia bernd gruhn , susan wittig , thomas ernst , jana ernst university of jena, jena, germany, background: a -week-old infant was diagnosed with very immature acute lymphoblastic leukemia (all) with myeloid markers in a foreign university hospital. at the end of induction therapy according to the current lal/shop protocol % leukemic cells were detectable in the bone marrow. treatment was changed to fludarabine, cytarabine and granulocyte colony-stimulating factor (flag) in combination with liposomal doxorubicin. after this re-induction still % leukemic cells were detected in bone marrow, so the bispecific t-cell engager antibody blinatumomab was given. due to an increasing portion of leukemic cells during the continuous infusion, antibody therapy was stopped and a cycle of clofarabine, cyclophosphamide and etoposide was administered. unfortunately, still % leukemic cells were detectable afterwards. because of chemotherapy-refractory leukemia a palliative oral treatment with mercaptopurine was started. however, the parents did not accept the palliative situation and searched for alternative therapeutic options in other university hospitals in europe. after plenty of refusals the infant was admitted to our hospital five months after diagnosis. methods: for molecular characterization genomic dna was isolated from leukemic cells. a mll-mllt /af rearrangement as a consequence of the translocation t( ; ) (p ;q ) was detected and used as a marker for minimal residual disease. for further molecular characterization targeted deep next-generation sequencing was performed for a panel of leukemia-associated genes. interestingly, no mutation was found. to allow precise immunophenotyping of the leukemic cells treatment with mercaptopurine was stopped. results: as in the first immunophenotyping the cd antigen was found, we administered the anti-cd monoclonal antibody gemtuzumab ozogamicin twice within two weeks. because of the detection of cd + leukemic cells after infusion of gemtuzumab ozogamicin, the anti-cd antibody daratumumab was given alternating twice within two weeks. unfortunately afterwards, leukemic cells reappeared being negative for cd und cd , but positive for cd . therefore, we administered the third antibody, the anti-cd monoclonal antibody inotuzumab ozogamicin, whereupon our patient developed a tumor lysis syndrome and a severe bone marrow aplasia. shortly after, allogeneic bone marrow transplantation from an unrelated donor using a special conditioning regimen consisting of thymoglobulin, busulfan, fludarabine and clofarabine was conducted. clofarabine was added because an additional antileukemic effect especially in infant all with mll rearrangement was described. after transplantation the patient suffered from a severe hepatic sinusoidal obstruction syndrome with massive ascites, renal and pulmonary dysfunction, but finally the patient recovered completely. the first bone marrow examination days after transplantation revealed a donor chimerism of % and a complete molecular remission using the mll-mllt /af rearrangement as marker for minimal residual disease. in all follow-up bone marrow samples we observed a complete donor chimerism and a complete molecular remission. currently, eight months after transplantation the patient is in a very good physical condition with normal development according to the age. background: paediatric chronic graft versus host disease (cgvhd) is a debilitating condition associated with substantial morbidity and mortality. to date, there are no approved therapies for paediatric patients with cgvhd, and current treatments often lack sufficient efficacy or lead to severe/life-threatening toxicities that limit their effectiveness. ibrutinib, a first-in-class, once-daily inhibitor of bruton's tyrosine kinase (btk), is approved in the us for the treatment of adult patients with cgvhd after failure of ≥ lines of systemic therapy. this phase / study will evaluate the use of ibrutinib in paediatric patients with moderate or severe cgvhd. methods: this open-label, multicenter, international phase / study (pcyc- ) includes patients with moderate or severe cgvhd as defined by the nih consensus development project criteria. it is divided into two parts: part a will determine the recommended paediatric equivalent dose (rped) of ibrutinib in patients aged ≥ to < years, and part b aims to evaluate the safety and efficacy of ibrutinib in patients age ≥ to < years. for part a, patients with cgvhd aged ≥ to < years who have failed ≥ lines of systemic therapy will receive once daily oral ibrutinib at a starting dose of mg/m to be escalated up to mg/m after days, if no grade ≥ toxicities occur, until the rped is determined. for part b, patients aged ≥ to < years with cgvhd who have failed ≥ lines of systemic therapy or have newly diagnosed cgvhd will receive once daily ibrutinib ( mg) until one of the following criteria is met: treatment is no longer required; new systemic treatment for cgvhd is initiated; progression of cgvhd; recurrence of underlying disease; or unacceptable toxicity. patients with newly diagnosed cgvhd will receive ibrutinib in addition to daily corticosteroids ( . - mg/kg prednisone). patients < years of age may be enrolled in part b and treated at the rped after it is determined in part a. key exclusion criteria include uncontrolled active systemic infection or active infection requiring systemic treatment; progressive underlying malignant disease or any post-transplant lymphoproliferative disease; or active hepatitis c/hepatitis b virus. patients must have adequate renal, hepatic, and hematologic function to be enrolled. the primary endpoint of part a is the rped of ibrutinib, as based on pharmacokinetic (pk) data; secondary endpoints include safety and pharmacodynamics (btk occupancy). the primary endpoints of part b are pk and safety of ibrutinib in paediatric patients with cgvhd. secondary endpoints for part b include response rate at weeks as defined by the nih consensus development project criteria; duration of response; overall survival; and late effects on growth, development, and immune reconstitution. results: this global study is currently enrolling. conclusions: this phase / study will explore the use of ibrutinib in paediatric patients with cgvhd to potentially meet the high unmet need for proven effective therapies for this population. disclosure enzyme replacement therapy (ert) is the treatment of choice in non-neuropathic hunter syndrome, but as the recombinant enzyme does not cross the blood brain barrier and neuropathic hunter syndrome is left untreated. hematopoietic stem cell transplantation (hsct) is the standard of care in patients with severe mucopolysaccharidosis (mps) type i (mpsih, hurler syndrome) as early transplantation halts cognitive decline in these patients and significantly improves survival. only few case studies have been published on the potential benefit of hsct in mps ii and mostly used busulfan-based conditioning regimens. in one comparative non-randomised multicenter study, hsct might to be superior compared to ert. here, we present our experience in hsct in three children with hunter syndrome using a treosulfan-based conditioning regimen. methods: a retrospective chart review was carried out in patients, who underwent hsct for hunter syndrome. the conditioning chemotherapy regimen included fludarabine, treosulfan, thiotepa and atg. all patients received bone marrow of either related and or matched unrelated donors. gvhd prophylaxis was performed with csa and methotrexat. results: three patients with hunter syndrome were transplanted in our department in . the age was six months, two years and four years, respectively. bone marrow donors were related in one patient and matched unrelated in two patients. the conditioning therapy was generally well tolerated. major complications were fever of unknown origin with need for antibiotic therapy and a mucositis. one patient developed a cmv reactivation. all patients engrafted successfully and recovered well from the hsct. there was no case of acute or chronic gvhd. in all three patients are alive. donor chimerism is complete in one patient; two patients have a mixed donor chimerism. after application of donor lymphocyte infusions in one patient, donor chimerism is stable at a low level of %. the donor chimerism of the other patient still slowly declines to currently %. after stem cell transplantation, two patients did not show further progression of the disease and even achieved psycho-motor improvements. interestingly, one of these patients is the one with the low donor chimerism of %. one patient suffers from a further progression of the underlying disease with psycho-motoric agitation, aggressive behavior and loss of speech, that occurred within the first year following hsct, but neurocognition stabilized thereafter. conclusions: we found a beneficial effect of hsct on the neuropsychological outcome or at least stabilization of neurocognitive function in our patients with a follow-up of eight years. despite low toxicity of the conditioning regimen, increased donor chimerism may further improve the neurological outcome. disclosure: nothing to declare. tandem sct in pediatric solid tumors, other than brain tumors, has no advantage in terms of efs over single procedure-single center experience , germ-cell tumour (gct), ewing sarcoma (es), nefroblastoma. patients were divided into groups according to the number of procedures: st group-single sct procedure, nd group-multiple procedures. regimens used for stem cell mobilisation were: topo-cy for nbl and epi-tax for gct, followed by g-csf±plerixafor. conditioning regimens: bu-mel and thiotepa-cy for pts with nbl, thio-tax and ice for pts with gct. patients received antibiotic, antiviral and antifungal prophylaxis, parenteral nutrition and supportive treatment. patients received consolidation treatment, followed up monthly in the first year, then yearly. patients were evaluated for residual disease by imaging tests. parents signed informed consent forms. results: we performed sct procedures to patients: . % nbl, . % es, . % gct and . % nefroblastoma. for this study only patients with nbl and gct were considered. in st group were % of pts, % in nd group. patients were diagnosed, staged and treated according to international protocols. sex ratio was f/ m. age distribution was - y % ( pts), - y % ( pts), > y % ( pts). peripheral stem cell (pbsc) mobilisation was more difficult in patients with multiple courses of chemotherapy±radiotherapy. we found no difference in the period of engraftment following a nd or rd procedure. hospitalization and supportive measures increased in nd and rd procedures ( to days). patients with multiple courses of chemotherapy and multiple hospitalizations had increased infectious risk and during the nd or rd procedures developed various infectious complications.incidence of severe oral mucositis after the first hsct was %, after tandem hsct was %. nbl patients : st group- / patients alive and efs, / receives anti g treatment; nd group- / patients-alive, / patients-not reached timepoint for mibg scan; / patients-mibg negative at first, relapsed after mo; / patient deceased due to pulmonary toxicity. gct patients: st group- / patients alive and ef, / high values in afp levels and receives metronomic therapy, / patients deceased due to progressive disease, but only had sct. only / patient had one procedure and died due to progressive disease. conclusions: in our study, tandem hsct in children with solid tumours lead to an increase in survival rates, at least in the first months after sct. most patients ( %) had progressive or relapsed/refractory disease when referred to our department. multiple procedures require a higher number of cd cells, very hard to achieve in patients with multiple courses of chemo± radiotherapy. new approaches have to be considered in these diseases, especially in high risk group. disclosure: nothing to declare background: antimicrobial prophylaxis for prolonged neutropenia occurred during the pre-engraftment period is a common practice in allogeneic hsct recipients. data on its effectiveness are few and generally from cases series and not from randomized clinical trials, especially in children. methods: all clinical records of allo-hsct performed from january st to november th at hsct-unit of istituto g.gaslini, genoa-italy, were retrospectively reviewed. collected data were underlying diseases, type of donor, antibiotic prophylaxis administration and type, development of fever and pathogen isolated from blood culture, if any, during pre-engraftment neutropenia. antibiotic prophylaxis, usually starting together with the conditioning regimen, was categorized in "standard" (with amoxicillin/clavulanate or ampicillin/sulbactam) or "tailored" (when based on previous bacterial isolations or colonizations). results: allo-hscts were performed in pediatric patients ( % males) with a median age at hsct of years (iqr: - ; range: - ). hscts were performed from alternative donor (ad) in % patients, from relative donor (rd) in %, and from haploidentical donor in %. table shows the pre-engraftment febrile neutropenia episodes according to type of antibiotic prophylaxis. ( %) hscts received standard prophylaxis, while ( %) the tailored one; only in ( %) did not receive any prophylaxis. fever occurred in ( %) of episodes in patients receiving standard prophylaxis, in ( %) of those treated with tailored prophylaxis and ( %) in the group without prophylaxis; only % of patients who received prophylaxis did not develop fever.in % of patients, the febrile episodes were diagnosed as bloodstream infections: staphylococcus aureus in %; cons in %; enterococcus spp in %; enterobacteriaceae in %; pseudomonas aeruginosa in %; other non-fermenting gram negatives in % and fungi in %. conclusions: the occurrence of fever in patients who received antibiotic prophylaxis suggested that it could not be effective in prevention of fever related to neutropenia after allo-hsct. the personalization of prophylaxis could be a possible path to follow these patients. disclosure methods: a total of patients with leukemia or neuroblastoma were included in the study. patients' mothers signed an informed consent for participation in the study. six of study participants were boys and girls, all aged to years. the control group consisted of healthy preschool children ( groups of children aged to years), boys and girls. results: in most of games the role of a doctor was played by a child. only one child declined to impersonate both a patient and a doctor. younger children mostly agreed to have for a "patient" a toy (proposed by psychologist or one of child's own), child's mother or a medical psychologist. the game lasted for - minutes. most patients preferred using real medical consumables and instruments (syringes, adhesive tape, winged infusion sets or, more rarely, pills). most often a syringe or an adhesive tape was chosen. as known from their mothers, among medical manipulations most unpleasant for children are injections and changing implanted catheter dressing. also, most healthy preschool children preferred using real medical instruments over toy ones. group more often used a syringe, a winged infusion set, adhesive tape, gauze or pills. group most often chose syringe or gauze. among medical instruments both groups more often chose a phonendoscope or thermometer.one patient refused to cause pain to a "toy patient". other children sympathized with a "toy patient", stroke injection or dressing location sites or used soothing terms ("wait a little", "it's going to be all right"), wished prompt recovery and hugged their "patients". one child was angry over his "patient" wishing him to "get hurt too". first preschool group children were mostly scolding a toy "patient" for "being guilty of getting sick". second group children were mostly compassionate, encouraged a "toy patient" telling that "all the procedures are needed to get healthy". from children's schoolmasters we know that all first group children received vaccination about a week before a test. children from second group had no injections. overall attitude towards toy "patients" was more mild in the second group. conclusions: . during a play children mostly use the medical devices which cause them most discomfort and/or pain. . manipulating the items children illustrate their own impression of medical procedures, which are most unpleasant. . children may express their negative emotions directed towards medical manipulations via their play actions, these negative reactions may be suppressed in different ways by parents or medical staff. . the intensity of child's own traumatic experience and an attitude of nearby adults may influence the child's attitude towards other patients. . the mother's wish for a child to tolerate all medical procedures with ease exceeds real capabilities of a small child. disclosure: nothing to declare. allogeneic stem cell transplantation in patients with mucopolysaccharidosis type iiia (sanfilippo): a case series methods: allogeneic sct was performed at the ages of , and years, respectively. all three patients received intrathecal enzyme replacement therapy within a clinical trial setting prior to hsct. the conditioning regimen consisted of treosulfan, fludarabine, thiotepa and thymoglobuline. gvhd-prophylaxis was carried out with csa and mtx in two patients and csa and mmf in one patient. stem cell source was bone marrow in two patients and peripheral blood stem cells in one patient. results: the conditioning regimen was well tolerated and all three patients successfully engrafted. two of three patients had an uncomplicated course without occurrence of acute or chronic graft-versus-host-disease (gvhd). at last follow-up and months after hsct, both patients are in good condition and show constant progress of psychomotor development. the third patient experienced severe steroid refractory acute gvhd of intestines (stage ) and skin (stage ), which resolved under intensive immunosuppression with cyclosporine, mycophenolate and ruxolitinib. around day after hsct, this patient showed clinical and biochemical signs of transplant-associated microangiopathy (tma) with cerebral seizures and acute renal failure. the cerebral mri showed progressive cerebral atrophy and leukoencephalopathy, also consistent with a progress of the mps iiia. at last follow-up months after hsct, this patient had recovered from tma and was in a stable clinical condition. conclusions: in consideration of the small case number and the short follow-up period in our cohort, allogeneic hsct might be considered as a salvage therapy for patients with mps iiia if other therapeutic options are unavailable for children with this otherwise unfavourable prognosis. however, the early psychoneurological course after transplant seems promising compared to the literature and hsct could become a treatment option for this rare disease. disclosure: nothing to declare methods: for the identification of underlying molecular mechanisms leading to the increased sensibility of rms cells, the activation status of different nf-kb signaling pathways were analyzed using western blot analysis and quantitative real time pcr (qpcr). further, flow cytometry was used to analyze the surface expression of death receptors on either sm treated or untreated rms cells. the overall effect on cell death induction was measured by pi/hoechst staining using a fluorescent microscope. results: treatment with sm led to the suspected degradation of iaps. followed by the activation of both the canonical nf-κb signaling pathway, indicated by the phosphorylation of iκbα and p , and the non-canonical nf-κb signaling pathway, as indicated by the accumulation of nik and the degradation of p to p . determination of selected target gene transcription revealed an upregulation of the inhibitor iκbα, nik, p , il- and at later time points the death receptors trail-r and trail-r . analysis of gene transcription also led to the finding of neither up-nor downregulation of ciap and p . to evaluate the involvement of trail-r and trail-r in the sm induced sensitization towards nk cell-mediated killing, surface expression of both death receptors was analyzed. treatment with sm led not only to an induced transcription of trail-r and trail-r , but also to an increased surface presentation of trail-r . subsequent ligation of trail-r by either wt-trail or a specific agonistic antibody (etr- ) resulted in a significant increase in cell death induction. the aforementioned analysis of gene transcription hints towards a bimodal feedback mechanism regulating both, the canonical and non-canonical nf-κb signaling pathway. on the one side, the canonical pathway is negatively regulated by the induced transcription of the inhibitor iκbα. on the other side, the induced transcription of nik, p and relb points towards a positive feedback loop of the non-canonical pathway. one mechanism of the increased rms cell sensitivity might be the induced transcription and surface presentation of the death receptor trail-r . the involvement of trail receptors is further validated by the cytotoxicity data, illustrating a sm mediated sensitization towards a trail induced cell death induction. this mode of cell death fits to the previous research, were trail transcription could be induced in nk cells by sm treatment. the graphical abstract shows the transcriptional upregulation of target genes leading to a putative bimodal nf-kb regulation and increased surface presentation of trail-r by treatment with smac mimetics. aim: to investigate the outcome of ucb transplantation in pediatric patients with malignant and non malignant diseases methods: data from patients underwent first allogeneic bone marrow transplantation with ucb from / until / were retrospectively analyzed. eighteen had malignant disease (md), of whom in complete hematologic response, and non malignant disease (nmd) (scid , chronic granulomatous disease , severe aplastic anemia , s.kostmann , osteopetrosis , wiskott-aldrich , amegakaryocytic thrombocytopenia ). the majority of the patients were male, for md and nmd, as well (m: /f: , m: /f: , respectively), of median age . years (range . - . years) and . years (range . - . years), respectively results: all patients but one, received ucb unit. hla compatibility in antigen/allele level was at least / and only in patients with md was / . conditioning regimens were myeloablative and tbi gy was given in / . gvhd prophylaxis consisted of cyclosporine and atg was given in all patients pre-transplantation. median value of nucleated cells for md was . χ /kg (range - . χ / kg) and for nmd was . χ /kg (range - . χ /kg). neutrophil and platelet engraftment was achieved in / and / patients with md respectively, in a median time of days (range - ) και days (range - ). in patients with nmd, neutrophil and platelet engraftment was achieved in / and / with median day of engraftment days (range - ) και days (range respectively. acute gvhd grade ΙΙ-Ιv presented in / patients with md and / with nmd, although none had cgvhd. the incidence of viral infections was cases in patients with md and cases in patients with nmd. disease relapse occured in / patients with md. after a median time of years follow up, overall survival (os) and event free survival (efs) for children with md were % and . % respectively, while for nmd, os and efs were %.treatment related mortality at d+ was % for md and % for nmd. among patients with md, are still alive, while the rest died from relapse (n: ), viral infections (n: ), septicemia (n: ) and agvhd(n: ). among patients with nmd, are alive, while the rest died from viral infections (n: ), septicemia (n: ) and multiple organ failure (n= ). the median time of hospitalization for patients with md was days (range - ), whilst for nmd was days (range - ). conclusions: transplantation of unrelated ucb in our unit was combined with high trm in children with nmd and higher probability of relapse for md. disclosure: nothing to declare p serum levels of -s cysteinyldopa is associated with stem cell transplantation related complications yukayo terashita , mamoru honda , minako sugiyama , yuko cho , akihiro iguchi hokkaido university hospital, pediatrics, sapporo, japan background: diffuse hyperpigmentation is common in patients who received chemotherapy or stem cell transplantation (sct). however, there are few reports of the relationship between skin reaction such as pigmentation and the other complications. pigmentation of the skin is thought to be the result of melanin stagnating in the dermic layer due to increased synthesis of melanin and destruction of the basement membrane due to inflammation induced chemoradiotherapy. melanin pigments are classified into two types: brown to black eumelanin and yellow to reddishbrown pheomelanin. -s cysteinyldopa ( scd) is precursors of pheomelanin, and its serum level has been used specific biochemical marker for malignant melanoma. here, we examined serially scd during the course of sct to determine association with sct related complications, because visual evaluation of skin color is difficult, and there have been no reports about scd as sct related biomarker. methods: we prospectively analyzed ( males, females) patients who received sct between may and march in hokkaido university hospital. the median age at transplantation of the patients was . years (range, - ). indication for sct were acute myelogenous leukemia in patients, acute lymphoblastic leukemia in patients, and other disease in patients; juvenile myelomonocytic leukemia( ), malignant solid tumor( ), immunodeficiency( ), anaplastic anemia( ), and diamond blackfan anemia ( ) . patients received allogeneic sct and received autologous sct. myeloablative conditioning (mac) was used for patients and reduced intensity conditioning (ric) was used for patients. sera were obtained from patients before conditioning therapy, on day , + , + , + , + and + . all blood samples were centrifuged at , rpm for min, and stored at - ˚c until used. we also examined sct related complications such as graft-versus-host disease (gvhd), viral infection, and pre-engraft syndrome (pes). statistical analyses were completed using the mann-whitney u test for unpaired samples, and kruskal-wallis test for three samples. each test was performed with a % level of significance. results: the average value of scd reached two peaks, day ( . nmol/l) and day ( . nmol/l), regardless of stem cell source and intensity of conditioning. in all patients, we found that the level of scd on day was associated with viral reactivation (p= . ), scd on day was associated with pes (p= . ), and scd on day was associated with malignant disease (p= . ). similarly, in patient who received allogeneic sct (n= ), the level of scd on day was associated with viral reactivation (p= . ), scd on day was was associated with pes (p= . ), scd on day was associated with malignant disease (p= . ). in addition, the level of scd on day was associated with gvhd of skin (p= . ), the peak level of scd was associated with acute gvhd (p= . ). conclusions: we found that scd can be a biomarker for sct-related complications such as aute gvhd. it is presumed that the production of pheomelanin could be induced by inflammatory procedure in sct. disclosure: nothing to declare p hsct in children with bone marrow failure: outcomes from a single singapore centre prasad iyer , michaela seng , vijayakumari k , ah moy tan , mei yoke chan , rajat bhattacharyya kk women's and children's hospital, paediatric haematology-oncology, singapore, singapore background: children presenting with pancytopenia often present a challenge to the paediatric haematologist. the underlying diagnosis can be hard to establish as many of the inherited bone marrow failure syndromes (ibmfs) can present with protean manifestations. the large majority of patients with bone marrow failure are often diagnosed with idiopathic severe aplastic anaemia (saa) despite extensive testing. we report our experience of hsct in patients treated with primary and acquired bone marrow failure. methods: we reviewed case notes of all the children who underwent hsct for bone marrow failure in our centre. results: a total of fifteen patients underwent eighteen stem cell transplants in our centre between and . three patients were diagnosed with fanconi anaemia, one with hoyeraal-hreidarsson syndrome, one with paroxysmal nocturnal haemoglobinuria and the remaining ten children had idiopathic saa. eight children had matched sibling donor transplant, had a matched related donor, had a matched unrelated donor, had umbilical cord blood transplants and the remainder were haploidentical transplants. four of the haploidentical transplants were t-cell depleted and one was t-cell replete. one child with fanconi anaemia had primary graft rejection with cord blood transplant and was successfully rescued with a haploidentical transplant. one child with saa had primary graft rejection twice (t-cell depleted graft) and then was rescued with an alternate haploidentical donor with a t-cell replete graft. of the two patients who died, one had a fatal fungal infection ten months after transplant, and the other died due to a severe influenza pneumonitis three and a half years after bmt. conclusions: haematopoietic stem cell transplant outcomes from our centre are comparable to leading centres in the world. the understanding of underlying conditions that present with bone marrow failure has improved our approach and the way we treat bone marrow failure syndromes. clinical trial registry: not applicable. disclosure: nothing to declare. methods: a retrospective study was performed in children treated with hsct who received pos or flu during early neutropenic period until engraftment from january to december at siriraj hospital in thailand. the efficacy, safety and tolerability of pos were compared to flu. results: there were hsct recipients (allo-hsct . %, auto-hsct . %) with mean age of . + . years. most of the patients were thalassemia ( . %) followed by hematologic malignancy ( . %) and solid tumor ( . %). seventeen and cases received pos and flu, respectively. all of patients in pos group were allo-hsct whereas . % in flu group were allo-hsct. in pos group, cases were diagnosed with suspected ifi and cases were probable ifi with total cases ( . %). in flu group, cases were diagnosed with suspected ifi and cases were probable ifi with total cases ( . %) which compared groups were not statistically significant (p= . ). no possible and proven ifi in both groups. in flu group patients received empirical antifungal treatment more than pos group but no statistical significance ( . % vs. . %, p= . ). both groups had similar rate of elevated liver function test (p= . ). no early discontinuation of antifungal prophylaxis for intolerance was found in both groups. only . % of patients achieved pos target trough level of . mg/l after days of treatment with started dose mg/kg three times a day. conclusions: pos and flu are comparably effective, safety and tolerability in ifi prophylaxis in neutropenic children treated with hsct. defining dose recommendation of pos in this setting requires larger studies. disclosure background: severe congenital neutropenia (scn) is typically characterized by anc of < /μl, maturation arrest of bone marrow myeloid precursors at the promyelocyte-myelocyte stage, and susceptibility to lethal pyogenic bacterial and fungal infections. scn is a rare group of disorders resulting from intrinsic defects in myeloid cell proliferation and maturation caused by mutations in several genes; elane, hax , gfi , was, and g pc are among the most common ones. almost % of patients are refractory to g-csf, and the only definitive curative approach for such patients is allogeneic hsct. the current absolute indications for hsct is failure to respond to g-csf treatment, or the development of mds/leukemia in patients with scn. here, we present the result of children with scn who received allogeneic hsct . methods: we retrospectively assessed allogeneic hsct in children with severe congenital neutropenia. all patients received busulphan (bu) based myeloablative conditioning regimen. busulphan was used according to weight adjusted dose. in addition, all patients received fludarabine mg/m in five days or cyclophosphamide mg/kg in days and atg mg/kg in days. cyclosporin-a and mtx were used for graft versus host disease (gvhd) prophylaxis. donor chimerism was evaluated in either bone marrow or peripheral blood on days + , + and + . results: the median transplantation age of the patients was months (range - months). six of them are male. two of the donors were matched siblings and were unrelated two of which were ag ag mismatched. stem cell source was bone marrow in patients, peripheral blood in and cord blood in patients. all patients engrafted. the median time of neutrophil and platelet engratment to was ( - ) days and ( - ) days, respectively. graft rejection was experienced in patients, one of them had received unrelated cord blood. all patients are alive, eight of which are with full donor chimerism (between - %) without any complication (no infection, no gvhd) with a median months (range - months) follow up. probability of disease free and overall survival were found % and %, respectively. conclusions: we concluded that hsct is a useful treatment for scn patients, especially those who are unresponsive to gcsf treatment and at high risk for leukemic transformation. however, a larger number of scn patients and longer follow-up are necessary to identify appropriate conditioning regimens and long-term prognosis. disclosure: nothing to declare background: prolonged thrombocytopenia (pt) or secondary failure of platelet recovery (sfpr) are a lifethreatening complications that occurs in - % and - % respectively of the patients following allogeneic hematopoietic stem cell transplantation (allo-hsct). management strategies, including the use of growth factors, cd +-selected stem cell boost, mesenchymal stem cell (msc) transfusion, and second allo-hsct, are not effective or possible for all patients. eltrombopag, is an oral non-peptide thrombopoietin receptor agonist, that leads to signal transduction and results in promoting the proliferation and differentiation of megakaryocytes. some recently studies show that also can promote haematopoiesis along all three lineages. methods: we described our experience in paediatric patients with poor graft function or secondary failure of platelet recovery after allogeneic stem cell transplantation treated with eltrombopag. results: patients characteristics are detailed in table . all the patients received and allo-hst. the median dose of cd + cells infusion was . x e /kg ( . - . ). neutrophils engraftment occurred in + day ( - d) and platelets in + day ( - d). all the patients had an hypoplastic bone marrow with complete chimerism. the median duration from transplantation to spcf diagnosis was months ( . - m) . one of the patients received a stem cell boost prior to eltrombopag, without response. the time onset from spgf/sfpr diagnosis to initiating eltrombopag was days ( - d). eltrombopag was started at a dose of mg/kg/d, requiring an increase dose in all cases. the median dose was mg/d ( - mg). the overall response rate was % ( / ). two patients achieved complete response (cr), as defined by platelet ≥ × /l. both patients already got neutrophil ≥ . × /l without g-csf. the time from eltrombopag initiation to achieving cr was ( - d) days. the treatment was given for a median of days ( - ). it was discontinued after and days respectively in the two responder patients. both patients maintain stable blood counts after discontinuing the treatment. the non-responders patients had to stop the treatment because of other reasons not related to eltrombopag. patient had to be rescued with a cd + cells boost with a good response. two patients that were in treatment with voriconazole for a fungal infection developed hyperbilirubinemia. there were no grade - toxicities related to eltrombopag. conclusions: in our experience, according to recently published studies, eltrombopag is a safe and efficacy drug in the treatment of secondary failure of platelet recovery post-hsct. it may be used successfully in children. sometimes higher doses may be considered if no response is achieved. further prospective trials are needed to increase the level of evidence and to identify predictors of response. disclosure: nothing to declare very slow clearance of busulfan in a child with infant leukemia background: busulfan is a drug with a high interindividual variability between dose and exposition. therefore, it is recommended to perform therapeutic drug monitoring (tdm) in the context of myeloablative conditioning, especially in children. methods: we report on a -month old boy ( . kg, cm) of caucasian decent born to non-consanguine parents with mll-rearranged prob-lymphoblastic leukemia. diagnosis was established one month after birth from peripheral blood and csf tap showed cns involvement. primary chemotherapy was commenced according to the interfant- protocol. however, mrd remained positive two months under treatment, leading to an indication for allogeneic stem cell transplantation. in the interfant- protocol, we opted for a conditioning regimen comprised of fludarabine ( . mg/kg for days), busulfan and thiotepa ( x mg/kg). in our institution, busulfan is applied once daily with a target auc of - h*mg/l in this very high risk situation. according to body weight, busulfan was given with . mg/ kg as a three-hour infusion on the first day. busulfan concentrations in plasma were measured with gas chromatography-mass spectrometry (gc-ms) and auc was calculated using bayesian curve fitting. results: exact busulfan quantification was not possible after the first dose due to technical reasons. as the levels were estimated to be very high, we decided to reduce the second dose of busulfan by %. this resulted in a very high auc of h*mg/l for the second dose, so that busulfan was discontinued after two days, because it was calculated that the patient already received busulfan with a cumulative auc of h*mg/ml. trough levels after the first and second dose were and μg/l, respectively. the patient showed a very slow clearance of . l/h/sqm, while the volume of distribution was in the usual range ( . l/kg). bilirubin and liver transaminases were in the normal range at the time of conditioning, while albumin and quick were decreased on day + after transplantation the patient developed clinical und biochemical signs of venoocclusive disease (vod). vod symptoms completely resolved under therapy with defibrotide. leukocyte engraftment was established on day + . unfortunately, the patients suffered from an early relapse of the leukemia from day + . attempts to induce a second remission with blinatumomab failed. the patient is currently under palliative chemotherapy. conclusions: busulfan tdm is very important especially in infants receiving myeloablative doses of busulfan to prevent under-or over-exposure. there is evidence that high busulfan trough levels contribute to the development of vod, but anti-leukemic activity of busulfan and cns permeability make it a valuable drug for very high risk patients in childhood leukemia. larger patient cohorts are needed to assess the exposure dependent risks of toxicity versus relapse in infants and toddlers. disclosure: nothing to declare blood (ucb) obtained at delivery from three children who received a diagnosis of cerebral palsy. methods: immunophenotyping of the ucb leukocyte fraction was performed using multicolor flow cytometry. the procedure was performed according to the protocol by shatorje and colleagues ( ) . briefly, the ucb samples were labeled with specific antibodies and incubated in the dark for minutes. afterwards, the samples were treated with ml of bd facs lysing solution for minutes to preserve the leukocyte fraction only. the cells were washed using pbs (roche) and then centrifuged twice ( rpm, °c, minutes). the results were analyzed using the facsdiva software (becton dickinson). results: we found an increased white blood cell (wbc) count, lymphocyte count, and cd :cd ratio in all ucb samples. one patient had a low nk cell count and percentage, and another had a low b-cell count and percentage. one sample displayed high t (cd +) and th cell (cd +) counts, but with percentages within the limits of the reference values. conclusions: we detected elevated wbc and lymphocyte counts in all ucb samples, despite a lack of intrauterine infection symptoms. many authors have described the pathogenesis of hypoxic-ischemic encephalopathy. briefly, after an acute hypoxia-ischemia insult, activated resting microglia show macrophage-like activity. this leads to a break-down of the blood-brain barrier, infiltration by peripheral leukocytes, and brain exposure, which further exacerbates inflammation. the role of systemic inflammation is being evaluated in the animal model. it is known that systemic inflammation plays a role in traumatic brain injury (tbi) and is an independent risk factor for poor outcome in isolated tbi patients. on the other hand, m -phenotype microglia inhibit inflammation and protect neurons from secondary damage and death. however, anti-inflammatory mechanisms in neonates are immature and expose them to extremely intensive inflammation. therefore, anti-inflammatory agents, including stem cells, may be beneficial in these patients. disclosure: three of four authors are employees of the polish stem cell bank, warsaw, poland reference: background: under the hypothesis that early natural killer cell infusion (nki) following haploidentical stem cell transplantation (haplo-sct) will reduce relapse in the early post-transplant period, we conducted a pilot study to evaluate the safety and feasibility of nki following haplo-sct in children with recurrent neuroblastoma who failed previous tandem high-dose chemotherapy and autologous sct. methods: we used the high-dose i-metaiodo benzylguanidine and cyclophosphamide/fludarabine/antithymocyte globulin regimen for conditioning and infused × /kg of ex-vivo expanded nk cells derived from a haploidentical parent donor on days , , and posttransplant. results: seven children received a total of nkis, and nki-related acute toxicities were fever (n = ) followed by chills (n = ) and hypertension (n = ); all toxicities were tolerable. grade ≥ii acute gvhd and chronic gvhd developed in two and five patients, respectively. higher amount of nk cell population were detected in peripheral blood until days post-transplant compared with reference cohort. cytomegalovirus and bk virus reactivation occurred in all patients and epstein-barr virus in six patients. six patients died of relapse/progression (n = ) or treatment-related mortality (n = ), and one patient remained alive. conclusions: nki following haplo-sct was relatively safe and feasible in patients with recurrent neuroblastoma. further studies to enhance the graft-versus-tumor effect without increasing gvhd are needed. disclosure: nothing to declare regenerative medicine p repeated administration of g-csf using stem cell mobilization protocol could induce improvement of cognitive functions of children with cerebral palsy: phase ii randomized placebo-controlled study background: we performed phase ii randomized placebocontrolled clinical study to reveal the safety and feasibility of repeated granulocyte colony-stimulating factor (g-csf) administration for improvement of cognitive functions of children with cerebral palsy. methods: forty-four children with non-severe type of cerebral palsy were enrolled, and their age were - years old. g-csf ( μg/kg) was administered for days subcutaneously every months during months. we compared their cognitive functions with the magnetic resonance imaging (mri) findings and the following tools between before and months after treatment; zoo location and picture memory as working memory index (wmi) in wechsler preschool and primary scale of intelligence (wpssi), receptive and expressive vocabulary test (revt), and visual motor integration (vmi)/visual perception (vp) test. mobilized stem cell count and cytokine levels were measured before (d+ ) and after (d+ ) g-csf administration for days every months. results: no significant findings in demography was noticed between g-csf (g-) and placebo (p-) groups. no serious adverse events were observed during the whole study period. the non-severe adverse events such as urticaria (n= ), itching sense (n= ), bone pain (n= ), headache (n= ), fever (n= ), and stomatitis (n= ) were tolerable. the parents felt the clinical improvements of cognition in cases of g-group and cases of p-group (p= . ), of language in cases of g-group and cases of p-group (p= . ). in zoo location test, we can not find out the significant score (expressed as age equivalent) differences between g-and p-groups. however, in picture memory test, there were significant improvement of age equivalent of months ( . ± . → . ± . ) during months of study period in g-group compared to months in p-group (p= . ). in revt, there were significant improvement of months of age equivalent in expressive tests of g-group ( . ± . → . ± . , p= . ) compared to months in p-group. no significant findings were noted in receptive test. vmi test showed the increasing tendency of months of age equivalent in g-group ( . ± . → . ± . , p= . ) compared to p-group. the increment of cd + cell counts in peripheral blood were significant in ggroup compared to p-group. the changed levels of interleukin (il)- , il- , vascular endothelial growth factor (vegf) as well as g-csf were noted in g-group. we also observed the correlation of cognitive function tests and white matter connectoms of several networks using functionally-defined white matter atlases. conclusions: the repeated administration of g-csf using stem cell mobilization protocol is safe and feasible to improve the language and cognitive functions in children with cerebral palsy. further studies for cellular and paracrine effect of g-csf and/or mobilized peripheral blood stem cells would be needed. background: while high-dose chemotherapy (hdct) with autologous hematopoietic stem cell transplantation (auto-hsct) is an integral part of multimodal therapy for highrisk neuroblastoma (hr nb), there are still subgroups, in which the results are extremely poor. for these patients allografting (allo-hsct) may offer some hope. methods: we summarize the experience of consecutive hr nb patients receiving therapy in our pediatric transplant department in - . the median age was years ( months to years). a total of auto-hscts and allo-hscts were performed. all auto-hsct recipients were characterized by one or several high-risk features: age of more than months at disease (onset n= ), primary disseminated disease (n= ), unfavorable biologic variant (n= ), poor st -line therapy response (n= ) or systemic relapse (n= ). most patients (n= ) received bu-mel hdct (in younger patients oral busulfan was replaced by busilvex), in primary resistant cases a d/ d regimen was used. a total of patients with st (n= ) or nd (n= ) chemosensitive relapse, resistant relapse (n= ) or poor mobilizers with locally advanced resistant tumor (n= ) received allo-hsct from haploidentical donor with fludarabine-based ric. in cases the transplant was modified via immunomagnetic positive or negative selection, patients received post-transplant cyclophosphamide (post-cy)-based gvhd prophylaxis. gvhd prophylaxis also consisted of calceneurin inhibitors and sirolimus. thirteen of allo-hsct recipients received posttransplant immunoadoptive (n= ) or targeted (n= ) therapy. results: the -year os and efs in auto-hsct recipients was % and %, accordingly. all but one patient engrafted with a median time of ( - ) days. bu-mel regimen was characterized by acceptable toxicity with most common toxicities being oral mucositis and infectious complications. the vod/sos incidence was only %. four patients dies due to infection (n= ), cns hemorrhage (n= ), and secondary leukemia (n= ). according to multivariate analysis the most important prognostic factors were response to st line therapy and post auto-hsct mibg scan results. the prognosis in initially resistant patients with good response to nd or rd -line therapy was still very poor (all patients relapsed with the median efs of months). all patients receiving a second auto-hsct after relapse died due to disease progression. with a median follow up of ( - ) months allo-hsct recipients are alive, of them with no signs of disease progression. all long-term responders received post-transplant therapy. one patient died due to transplant complications, other deaths were caused by disease progression. there was no obvious difference between outcomes in post-cy based and transplant modification-based transplantations. agvhd more often developed in modified transplant recipients ( patients vs in post-cy group, of these cases gr iii-iv), patients in post-cy group had grade iii-iv hemorrhagic cystitis. the median time to engraftment was longer for ptcm group compared to transplant modification group (d + vs. d+ , accordingly). conclusions: while single hsct with auto-hsct is a golden standard in hr nb patients, the relapse rate is still high and the prognosis in relapsed/refractory patients is dismal. the allografting has some limited effectiveness in these cases and post-transplant therapy has a potential for further improvement. disclosure: nothing to declare p abstract already published. veno-occlusive liver disease (vod) is frequent but well treatable with early defibrotide administration in children with neuroblastoma receiving high-dose busulfan and melphalan background: using high-dose intravenous busulfan and melphalan (bumel) prior to autologous stem cell transplantation (sct) in children with high-risk neuroblastoma, seems to decrease toxicity of the myeloablative regimen, except for vod. in this multicenter retrospective study we aimed to assess the outcome of bumel-associated vod with early defibrotide treatment intervention. methods: we retrospectively analyzed children with high-risk neuroblastoma who underwent autologous sct with i.v. bumel regimen in slovakia and prague, czech republic in the period / - / . busulfan was administered in q hour schedule, with therapeutic drug level monitoring in % of patients. all vod patients except one were treated with defibrotide starting at a standard dose of mg/kg/day, given in doses per day. patient was treated with supportive therapy only. ursodeoxycholic acid was used as prophylaxis in all patients. vod was established using the modified seattle clinical criteria (corbacioglu, lancet ). results: the incidence of vod was % ( / ) in patients treated with intravenous busulfan and melphalan. there was no significant difference in busulfan total dose/kg between patients with ( . mg/kg (sd= , )) and without ( . mg/kg (sd= , )) vod manifestation. vod developed at a median of days after sct (range - days). anicteric forms of vod were documented, although % patients with vod ( / ) presented with increased bilirubin. % patients with vod ( / ) developed ascites but only patients ( %) required ascites drainage. no vod patient received renal replacement therapy and only one needed mechanical ventilation. importantly, we successfully treated vod in all patients. relapse or progression of neuroblastoma was the cause of death in vod patients ( %) who died. conclusions: despite targeting busulfan levels to decrease toxicity of the regimen, vod is common (we observed vod incidence exactly in the range of the siopen hr nbl- multicenter study (ladenstein, ) ). early recognition and early treatment with defibrotide seems to be effective in vod associated with bumel regimen -none of our patients died due to vod. disclosure: nothing to declare results of high-dose chemotherapy (hdct) with autologous hematopoietic stem cell transplantation (auto-hsct) in the treatment of ewing sarcoma family tumors (esft) background: while current dose-intense treatment protocols allow achieving - % survival in localized esft patients, the long-term survival in high-risk cases is still unsatisfactory. although there is a considerable body of data on high-dose consolidation the real effectiveness and optimal indications for this option are still not completely clarified. therefore, a large prospective cohort analysis may still yield useful data. methods: the whole cohort includes consecutive highrisk esft patients with median age of (range - ) years receiving hdct with auto-hsct in to after obtaining st (n= ) or nd (n= ) cr, pr (n= ), or stable disease (n= ). the high-risk features included lung (n= ), bone (n= ) or bone marrow (n= ) involvement, inadequate local control in primary axial tumors (n= ), large lesions volume or poor treatment response (n= ), and chemosensitive relapse (n= ). most patients had several risk factors. disseminated disease patients were also evaluated according to prognostic score by r.ladenstein et al. highdose busulfan-melphalan followed by autologous stem-cell transplantation (hdt/sct) was used. results: the median observation time was (range - ) months. the -year overall (os) and event-free (efs) survival were % and %, accordingly. most important outcome predictors were inadequate local control in chemoresistant cases, a primary tumor volume more than ml, more than one bone metastatic site, bone marrow involvement and additional lung metastases. according to prognostic risk score in disseminated disease esft patients identified three groups with -year os rates of % for score ≤ ( patients), % for score to ( patients), and % for score ≥ ( patients), (p< . ). conclusions: while bu-mel hdct with auto-hsct may still be a feasible option with acceptable toxicity for chemosensitive patients with inadequate local control and some primarily disseminated cases it is ineffective in primary resistant or very high-risk metastatic patients. disclosure: nothing to declare efficacy of tandem high-dose chemotherapy with autologous hematopoietic stem cell transplantation in the treatment of infant embryonal brain tumors day + (range, - ), after the second auto-hsct was day + (range, - ). two-year overall survival (os) was % and disease free survival (dfs) was %. dfs was significantly better among patients with mb ( %) and pnet ( %) in compared to children with etmr ( %), pb ( %) and atrt ( %), (p= , ). dfs in patients who received tandem auto-hsct was % in compare to infants who received only one auto-hsct ( %), (p= , ). complications grade (according to common toxicity criteria ) were observed in % of cases. conclusions: employment of tandem hdct with auto-hsct in primary infant embryonal brain tumors may be a feasible option for patients after induction treatment. both conditioning regimens had acceptable toxicity. all patients who had tandem hdct with auto-hsct had better os ( %) in compare with single hdct ( %). patients with mb and pnet had better prognosis with os % and %, respectively, in compare with other embryonal tumors. disclosure: nothing to declare background: metastatic extra ocular retinoblastoma is carrying a poor prognosis. therapeutic intensification with high-dose, marrow-ablative chemotherapy and autologous hsct has been explored, but its role is not yet clear. this study aimed to evaluate the survival outcome of patients with extraocular retinoblastoma post autologous stem cell transplant, treated at single center methods: this is a retrospective study included all patients with metastatic extraocular retinoblastoma (stages a and b) that underwent autologous hsct at the children cancer hospital egypt (cche) from november to july , the treatment protocol was adopted from cog protocol (aret ) as all patients received cycles induction chemotherapy followed by consolidation myloablative conditioning, cem (vp . mg/m x , melphalan: mg/m x , carboplatin: mg/m x ) and stem cell rescue. patients data including initial disease characteristics, transplant data, and survival outcomes were collected and analyzed results: a total of cases were included with median age of . years, and male to female ratio . . nine patients ( %) were initially presented by extra ocular disease, while patients were presented by intra ocular disease and progressed to metastatic disease. according to cog staging of extra ocular disease, patients had stage a, and were stage b ( of them had trilateral disease). after induction therapy, ( %) showed complete response and ( %) had ≥ partial response. with average cd count of x / kg, the median time to anc and platelet engraftment were days and days respectively, and there was no transplant related mortality. post-transplant radiotherapy was given only to patients. with median duration of follow up of months, the overall and event free survival rates of whole patients were . % and . % respectively conclusions: high dose chemotherapy and stem cell transplantation is a promising potential curative option for patients with metastatic extra ocular only two primary gf ( . %) occurred, both without dsa. patients developed a primary pgf ( %). -years os, years pfs and -year nrm were analyzed according to the presence of dsa in comparison with negative population. no statistically difference was found. no impact of the presence of dsa on the risk of developing gf and pgf was revealed. major outcomes of transplant was analyzed separately in patients with pgf and good graft function (ggf). -years os, -years pfs and year-nrm in ggf and primary pgf populations were % vs % (p< . ); % vs % (p< . ), % vs % (p= . ), respectively. conclusions: the presence of low level of dsa in the absence of desensitization doesn't correlate with the risk of developing gf and pgf. patients who experienced a pgf had worse outcomes in comparison with patients with ggf. disclosure: nothing to declare. the impact of hla-dpb mismatch in t-cell replete unrelated donor allogeneic stem cell transplantation background: high resolution matching of donor-recipient hla improves outcome in allogeneic stem cell transplants. matching for hla-a, -b, -c, -drb and -dq is mandatory in our transplant centre, to identify / or / matched unrelated donors. high resolution matching for dpb has been added over the last - years. however, the role of dpb matching is not yet clearly defined. methods: in this study, we retrospectively analyzed the impact of hla-dpb matching on the outcome of t-cell replete allogeneic hematopoietic stem cell transplants with cya/mtx-and without atg as gvhd prophylaxis in patients with hematological malignancies at oslo university hospital between and . patients with an unrelated donor fully matched ( / ) at hla-a, -b, -c, -drb and -dqb loci were included. further, patientrecipient pairs were also fully matched on dpb ( / ); had permissive and had non-permissive mimatches of one or two dpb alleles. the three groups were comparable with respect to diagnosis, gender, age, cytomegalovirus serostatus and conditioning regimen. results: cumulative incidence of relapse at years were significantly higher in the dpb matched pairs compared with the permissive and non-permissive mimatched ones, at % vs % and % (p< . ) respectively. relpase free survival and overall survival were superior in the nonpermissive and permissive dbp mismatched groups vs the fully matched, at % and % vs % (p= . ) and % and % vs % (p= . ) respectively. no difference in frequency of acute gvhd grade ii-iv between the three groups were found; dp match %, permissive mismatch % and non-permissive mismatched % (p= . ). neither was there a difference seen in gvhd grade iii-iv; % vs % vs %, respectively. finally, there were similar outcomes between the three groups regarding chronic gvhd and trm. in corrected multivariate analysis, only dp matching had significant influence on mortality and survival. conclusions: our results show a favorable relapse free and overall survival following a mud allotransplant with a dpb permissive or non-permissive mismatched donor compared to a fully dpb matched. this is likely due to an increased gvl-effect in dpb mismatched groups without the counterbalance of increased acute gvhd and trm. disclosure: nothing to declare p a haploidentical may be a better choice than a female genoidentical donor to transplant a patient with high risk acute myelogenous leukemia in first remission norbert gorin , myriam labopin , didier blaise , goda choi , gerard socie , jean henri bourhis , fabio ciceri , emmanuelle polge , arnon nagler , mohamad mohty china, the first affiliated hospital of soochow university, hematology, suzhou, china background: despite the incidence of leukemia increases with age, currently the geriatric population is poorly represented in the standards of care concerning that older adults undergoing hematopoietic cell transplant (hct) may experience higher transplant-related mortality (trm). previous studies have demonstrated that donor age is vital for older patients by affecting trm and survival. accordingly a relevant question is whether outcomes can be improved with a younger hla-haploidentical offspring donor rather than an older hla-matched sibling (msd). in our previous multi-center report under atg+g-csf based protocol for haplo hct, offspring donor is correlated with lower trm and higher leukemia free survival (lfs) as compared with older msd in subgroup analysis for recipients > years although it did not reach statistical significance. on the contrary, in a recent report from ebmt and cibmtr under ptcy modality for haplo hct, among patients aged to years, despite lower chronic graft-versus-host-disease (gvhd), graft failure, trm, and overall mortality were higher after transplant from offspring compared with an msd although there were differences in transplant platforms between the groups. methods: we extended our multi-center dataset and a matched pair analysis was performed. outcomes of acute leukemia patients (>= years) transplanted in cr / cr who received hct from offspring (n= ) or msd (n= ) between jan, and june, present in the multi-center database were analyzed. because the patient population was small, a : ratio matched pair analysis was implemented with the following matching factors: underlying disease (acute myeloid leukemia, acute lymphoblastic leukemia), disease status (cr /cr ), age and sex of patients, year of transplant, blood group incompatibilities, and sex of donor. results: we were able to match offspring with msd patients. the two matched groups were comparable in baseline characteristics except for donor age due to the family relationship. all patients achieved myeloid recovery with a median time of d and d for msd cohort and offspring group (p= . ). the d platelet recovery rate was % in both groups. the cumulative incidence of grade ii-iv acute gvhd in msd cohort was significantly lower than in offspring group ( % vs %, p= . ) while the incidence of chronic gvhd in msd cohort was significantly higher than in offspring group ( % vs %, p= . ). the -year trm ( % vs %, p= . ) were significantly lower in offspring-hct compared with in msd-hct and relapse incidence was comparable ( % vs %, p= . ). as a result, the -year overall survival ( % vs %, p= . ) and lfs ( % vs %, p= . ) ( figure ) were significantly higher in offspring-hct compared with in msd-hct. in a multivariate analysis, msd-hct remained a significant factor for decreased overall survival (hr . ( . - . ), p= . ) by increased trm ), p= . ) in comparison with offspring-hct. conclusions: these data favor a young offspring over an older msd in patients > years. the current analyses confirm non-hla donor characteristics, rather than hla disparity, predominantly influence survival in older acute leukemia patients. validation of these findings requires a prospective trial wherein the transplant platforms can be closely matched. [[p image] . figure . lfs in offspring-hct compared with in msd-hct ( % vs %, p= . )] disclosure: nothing to declare. impact of sibling donor-recipient sex combinations on rejection after hla-matched bone marrow transplantation for severe thalassemia cure children foundation, florence, italy, sankalp india foundation, bangalore, india, people tree hospitals, bangalore, india, south east asia institute for thalassemia, jaipur, india, pakistan institute of medical sciences, islamabad, pakistan, central asiri hospital, colombo, sri lanka, nawaloka hospital, colombo, sri lanka, kokilaben dhirubhani ambani hospital, mumbai, india background: severe thalassemia (st), i.e. a thalassemia syndrome with inability to keep spontaneous hemoglobin > g/dl, is a common indication for bone marrow transplantation (bmt) in children in the middle east and south east asia. sex mismatch has been associated with increased risk of solid organ rejection but is not generally considered an important transplant-associated risk factor in the context of fully matched sibling bmt for st. methods: a total of consecutive sibling bone marrow transplants carried out between january and april after conditioning with busulfan ( mg/kg oral, not adjusted to serum levels) and cyclophosphamide ( mg/kg) ( patients) in addition to either thiotepa ( mg/kg) ( patients), or anti-thymocyte globulin (genzyme mg/ kg or fresenius mg/kg on days - to - ) ( patients) and fludarabine mg/m ( patients) were analysed. all cases received cyclosporine and methotrexate or mycophenolate mofetil as gvhd/rejection prophylaxis. in the thiotepa group methylprednisolone at . mg/kg/day was also used during the first days after bmt (lucarelli protocol i). bone marrow was the source of hematopoietic stem cells in all cases, in the atg group it was g-csfprimed ( μg/kg/dose twice daily for to days prior to harvest). all patients were considered low risk based on liver size < cm from costal margin and age less than years (median . years, range . to . ), all sibling pairs where hla-compatible. results: [[p image] . sibling donor-recipient sex combinations.] the lowest rejection rate ( %) was observed in the sister to sister (s s) group of cases, followed by brother to brother (b b) group of cases with %. in the sister to brother (s b) group of cases, rejection rate was %, and % in the brother to sister (b s) group of cases. on univariate analysis the only significant difference at the p . level by log rank test was b s vs. s s groups (rejection proportions of % and % respectively). interestingly, all patients with rejection and persistent pancytopenia were female recipients of male grafts. conclusions: even though several preparative regimens were employed over an -year period, our data suggests that sex mismatch among compatible siblings should be considered as a relevant variable related to bmt decisionmaking. we also recommend to consider autologous back up hematopoietic stem cell collection and storage in sibling sex mismatched transplants, particularly in brother to sister bmts. same-sex fully matched related bmt for severe thalassemia might be the best scenario in which reducedintensity preparation strategies aiming at maximizing fertility preservation might be explored. disclosure: nothing to declare. outcomes of t-cell replete hematopoietic cell transplantation from mismatched related or unrelated donors using high dose post-transplant cyclophosphamide based gvhd prophylaxis background: high dose post-transplant cyclophosphamide (ptcy) based gvhd prophylaxis overcomes immunological barriers in hla mismatched donor transplantation. ptcy has been adopted in many centers as de facto standard for hct from haploidentical donors (haplo hct). it's use in mismatched unrelated donor transplant (mmud hct) is less well established. methods: we analyzed retrospectively outcomes of contemporary cohorts of patients who underwent haplo hct or mmud hct using ptcy + cyclosporine (csa) and mycophenolate mofetil (mmf) at our center. we compared these outcomes with outcomes of cohorts of patients who underwent hct from matched unrelated donors (mud) using atg based gvhd prophylaxis or matched sibling donor (msd) with csa and mmf. patients and donors were considered matched if they background: hla-alloantibodies are a major risk factor for engraftment failure in allogeneic hematopoietic stem cell transplantation (hsct). particularly, complement fixing, donor specific antibodies were shown to be associated with early engraftment failure. prospective antibody-screening, although not currently required for donor search, could permit early identification of high risk patients for positive crossmatch. aim of this study is to set the basis for future applicability of antibody-screening-based definition of acceptable mismatches in donor selection, by creating a large prospective antibody-screening database of patients due to receive an hla-mismatched allogeneic hsct. methods: patients (n= ) diagnosed with mds/mps, nhl, mm, cll, cml, anaemia (aplastic anemia, hemoglobinopathies, pnh) and hl were prospectively screened for hla-antibodies whenever initial donor search indicated that no completely matched donor would be available. screening was performed with an elisa class i +ii screening assay. all positive screening cases were tested for antigen-specific antibody identification with luminex sab, and acceptable mismatches were defined. the results were subsequently considered in donor search and selection. we now report the frequencies of alloimmunization observed in these patients. results: the highest rate of alloimmunization was observed in patients from the anaemia disease group (overall . %) followed by those from the mds/mpn group (overall . %). the lowest immunization rates were observed in cll (overall . %) and hl ( . %) patients. alloimmunization rates for hla-class i antigens (p< . ) were significantly higher compared to hla-class ii antigens. overall hla-class i immunization rates ranged from . % to . %. hla-class ii immunization rates ranged from . % to . % (table ) . conclusions: our findings suggest that patients with high transfusion burden like anaemia and mds/mpn patients have the highest risk of hla-alloimmunization with . % and . % anti-hla prevalence rates, respectively. analysis of follow-up data, will enable us to confirm whether prospective definition and consideration of acceptable mismatches in donor selection may lead to similar engraftment failure rates between immunized and non-immunized patients undergoing hlamismatched hsct. background: mothers displaying a persistent fetal microchimerism (fm) proved to be the most suitable donor in t cell-depleted haploidentical stem cell transplantation (hhsct) in children. we presumed that fetal cells leave an imprint in the mothers' immune system which positively affects recognition and elimination of malignant cells in the child by maternal effector cells. distinct killer cell immunoglobulin-like receptors (kir)/hla constellations are not only associated with reduced relapse rates after hsct in children, but also supposedly influence the establishment of an fm. methods: after approval by the local irb and obtaining informed consent, we initiated a protocol to elucidate the factors that influence the establishment, persistence and effect of fm. we established a digital droplet pcr (ddpcr) protocol to determine the fetal microchimerism. for differentiation between maternal and fetal cells, biallelic short insertion/deletion polymorphisms were used. kir and hla-c genotyping was performed by ssp-pcr. parental nk cell alloreactivity against the respective leukemic blasts and kir phenotyping were analyzed by flow cytometry. results: we analyzed parents, whose children were treated for hematological diseases at the university medical center hamburg-eppendorf. a fetal microchimerism was detected in % of the mothers. the amount of fetal cells varies between individuals ( x - - x - ). we observed a positive correlation between a persisting fm and hla-c homo-and heterozygous mothers along with a maternal cen a/b and cen b/b genotype. additionally, fm positive mothers showed a higher surface expression of the hla-c respective receptors kir dl /s . the percentage of alloreactive maternal nk cells against fetal cells was higher compared to paternal nk cells; while alloreactivity of fm positive maternal nk cells was similar to nk cells from fm negative mothers. conclusions: persistence of fm was more frequent in mothers carrying at least one hla-c allele and a centromeric b/x motif. phenotypically, fm positive mothers had higher expression of kir dl /s indicating a role of these receptors on the persistence of an fm. in vitro, maternal nk cells showed a higher alloreactivity compared to paternal nk cells. there was no difference in alloreactivity whether the mothers were fm positive or negative, suggesting other mechanisms are responsible for the superior outcome in transplantation from fm positive mothers. disclosure: nothing to declare background: although there have been significant improvements with conventional therapies in beta thalassemia major, hematopoietic stem cell transplantation is only curative therapy. related donors are preferred to diminish transplant risks. in lack of identical related donor, identical unrelated donors are second best choice. in this study, thalassemia major patients transplanted from unrelated donors (mud) were compared with thalassemic patients transplanted from relative donor (mrd) retrospectively. methods: patients who were transplanted between june and december in bahçelievler medical park hospital pediatric bone marrow transplantation unit were evaluated retrospectively. all patients were classified according to pesaro risk classification. thirty four of received busulfan, fludarabine, cyclophosphamide, thioteopa for conditioning, patients received myeloablative preparation regimen with treosulfan, fludarabine, thiotepa, cyclophosphamide. all patients were given atg, cyclosporine and methoteraxate for gvhd prophylaxis. the patients were compared in terms of acute complications in first days, engraftment, chimerism, acute and chronic gvhd after transplantation. results were evaluated with ibm spss statistics (ibm spss) program. results: a total of patients, ( . %) male and ( . %) female, aged between and years (median years) were evaluated. patients were evaluated in two groups as "mud" (n = ) and "mrd" (n = ) groups. there was no difference between groups about given stem cells (mud , ± , x /kg and mrd , ± , x / kg). neither significant difference between different pesaro risk groups in terms of developing acute and chronic gvhd and nor decreased chimerism were detected. neutrophil engraftment time ( , days) in mrd group was significantly longer than mud group ( , days) (p = . ) but no difference between platelet engraftments were observed. gvhd ratio was . % in mud donor group and . % in mrd group and no statistically significant difference was found(p> . ). the incidence of engraftment loss in mud group was . % and . % in the mrd group, and there was no statistically significant difference (p> . ). the rate of decreased chimerism was found to be significantly higher in the mrd group ( %) than in the mud group ( . %) (p: . ; p< . ). the survival rate was . % in the mud group and . % in the mrd group. the disease-free survival rate was . % in the mud group and % in the mrd group. the disease-free survival of mud group was significantly higher than mrd group (p: . ). conclusions: in our study, transplant related complications and success of transplantation with both mud and mrds were found to be similar. it is promising for mud transplantations to found lower decreased chimerism and similar os and dfss. based on these results, it was concluded that hsct from non-family donors, especially for patients incompatible with chelation therapy and had organ damage, transplantation from unrelated identical donors can be a good choice. although the results of our study seem promising, larger patient groups and prospective clinical trials are required. disclosure: nothing to declare background: use of g-csf stimulation of bone marrow (bm) donors is beneficial in many aspects; it can enhance tnc yield, but also have an immunomodulatory effect on donor t cell function, particularly invariant natural killer t (inkt) cells expansion as well as apcs. we analyzed outcomes of consecutive patients receiving bone marrow from hla-haploidentical donors that were stimulated with g-csf prior to harvest. methods: in the time period between / and / , patients received bone marrow from donors stimulated with ug/kg bw of g-csf on days - , - and day of bm collection. four patients ( %) received myeloablative (bucy) conditioning, one ( %) received tec ric conditioning while ( %) received nma ("baltimore") conditioning. all patients received posttransplantation cyclophosphamide (ptcy) on days + and + , tacrolimus and mmf were started on day + . for patients donors were fathers, mothers, siblings and children. results: median age was years ( - ), there were female and male patients. twelve patients had aml, hodgkin lymphoma, all, mds, nhl and cml. median number of infused tnc in graft was . x /kg bw ( . - . ) and cd + cells . x /kg bw ( - . ) . after median follow up of days (range - ), overall survival was %, with median survival of months. engraftment was established in ( %) patients, ( %) had primary rejection and patients ( %) died in sepsis prior to engraftment. of patients that engrafted, further ( %) patients had secondary rejection, two of them were transplanted again from a haploidentical donor, both using pbsc as a source of graft. median time to neutrophil recovery (anc> ) was days ( - ), while median time to platelet recovery (plt> x /l) was days ( - ) in evaluable patients. cumulative incidence of agvhd ii-iv was . % ( % ci, - );of note is that of patients that developed agvhd only one had grade iii, while remaining patients had grade ii. cumulative incidence of cgvhd requiring treatment was . % ( % ci, [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] . cumulative incidence of relapse was . % ( % ci, - ) and trm was . % ( % ci, - ). conclusions: the use of g-csf mobilized bm graft in the hla-haploidentical setting with ptcy has proven to be useful to us, not only in terms of tnc yield which was more than satisfactory and contributed to adequate hematological recovery, but also in the excellent control of both acute and chronic gvhd, with most patients developing agvhd of grade ii and only one grade iii (actually developed only after dli given for decreasing chimerism). comparative studies are of course warranted to prove benefit, but this data contributes to the growing body of evidence that indeed donor stem cell stimulation with g-csf has potentially powerful immunomodulatory effect. disclosure: nothing to disclose p other-relative donors as a reliable bank for allogeneic hsct in countries with culturally accepted cousin-cousin marriages: a two-year report from a pediatric center in iran background: although the optimal donors for patients undergoing allogeneic hematopoietic stem cell transplantation (allo-hsct) are fully-matched siblings, the cousincousin (consanguineous) marriages in some countries have extended the chance to find a matched donor for the hsctrecipient. in this study, an outcome analysis of transplanted patients receiving stem cells from their relatives other than siblings (other-relatives or non-sibling donors) is provided. methods: in this retrospective cross-sectional study, a two-year report of patients who received allo-hsct from their other-relative donors during september to september at the department of stem cell transplantation of children's medical center in tehran, iran is presented. the patients were followed up until st december . results: during this time period, patients underwent hsct (both autologous and allogeneic) at this center, of which cases received allo-hsct. out of allo-hsct recipients, the donors of stem cells for cases ( . %) were their other-relatives. the median (range) age at hsct was ( - ) years and the majority of patients were boys ( / , . %). according to disease class, the patients were most commonly involved with non-malignant hematologic diseases ( / patients, . %) (figure) . the source of hscs for most patients ( cases, . %) was peripheral blood and for only patients the source was bone marrow. the donors for patients were fully matched and only one patient received the hscs from a one-locus mismatched donor. hsct was successful in patients with most of them achieving full chimerism ( patients, . %) followed by those developing mixed chimerism ( patients, . %) and only one patient ( %) experienced graft failure. post-hsct complications included cmv infection in patients ( . %), other infections in ( . %), hemorrhagic cystitis in ( . %) and pres in ( . %). acute gvhd occurred in patients ( . %) and chronic gvhd in ( . %). death occurred in cases and of them were transplant-related, while was due to disease relapse and due to graft failure. the median of overall survival was ( - ) days. conclusions: the likelihood of receiving hscs from an hla-matched other-relative donor in one-thirds of children undergoing allo-hsct, with comparable outcomes to sibling and unrelated donors (as evidenced in this study compared with other studies), introduces family bank as a reliable source for pediatric allo-hsct in countries with culturally accepted cousin-cousin marriages. hence, for transplant physicians, parental consanguinity would be an indication of an extended search for a potential matched donor among the patient's family. [[p image] . distribution of patients according to disease and disease class] disclosure: nothing to declare. abstract already published. update on the hla frequency distribution of the portuguese bone marrow donor registry eduardo espada , dário ligeiro , hélder trindade , joão forjaz de lacerda frequency distribution varied throughout the country, allowing for analyses of molecular variance and generation of relatively geographically accurate graphical representations of genetic distances between regions and districts. conclusions: with the most recent hla analysis of the portuguese bone marrow donor registry we were able to extrapolate high-resolution haplotype frequencies from the most common low-resolution hla-a/-b/-drb haplotypes (corresponding to % of the estimated haplotypes at that level), which will lead to an optimization of its use, hopefully limiting the time between donor search and allogeneic hematopoietic stem cell transplant. disclosure: nothing to declare. abstract already published. abstract already published. unmanipulated haploidentical donor transplantation compared to identical sibling donor had better antileukemia effect for refractory/relapsed acute myeloid leukemia in not remission status background: patients diagnosed with saa with no sibling donors and who are refractory to immunosupression are candidates to hematopoietic stem cell transplant using alternative donors. haploidentical donor transplants has been reported using cyclophosphamide (cy) post stem cell infusion as immunephrophylaxis. the present study has the objective of evaluating overall survival and engraftment rates after haploidentical stem cell transplant for saa in a reference center. methods: saa adult patients (≥ yo) received hsct from haploidentical donors from de january/ to august/ . median age was y ( - ); donor was the father in six, mother in five and a brother in four cases. stem cell source was marrow in cases ( %). conditioning: patients ( %) received cy mg/kg, fludarabine mg/m² e tbi cgy. the remaining received the same drugs but radiotherapy dose varied from - cgy, all them received immunephrophylaxis with post transplant cy mg/kg, cyclosporine and mmf. median of infused cells (tcn) was , x /kg ( , - , ). results: eight patients engrafted ( %). among seven graft failures four received a second haploidentical transplant and one received an unrelated donor transplant as salvage regimen. two patients were successfully rescued after the second haplo and the others died from infectious complications. three years overall survival was %. death causes included: five infections and two lung hemorrhage. median survival was days ( - ). no patient had acute graft-versus-host-disease (gvhd) and one patient had mild c-gvhd. conclusions: haploidentical transplant was feasible as therapy for saa refractory to immunessupression with an overall survival of % in this cohort. graft failure however is still a problem to be addressed in this setting. disclosure: no disclosure stem cell mobilization, collection and engineering p abstract already published. key performance and quality indicators for a successful bone marrow collection marco sampaio , , ana salselas , fátima amado , filipa bordalo , sérgio lopes , catarina pinho , susana roncon and one from ecc (staphylococcus spp.)presented positive microbiological results. conclusions: bm collection is a challenging strategy because it is a one-time procedure and manually operatordependent technique; simultaneously it is more difficult to control the final cellular content of the bm, which is a risk for donor volume depletion. bm collection is feasible even with donor and recipient weight difference. poorer performance may be found when higher tnc are requested. we respond efficaciously when the request is between and * tnc but we fail to accomplish higher tnc values. we must emphasise that icc tnc demanded was generally lower than ecc. deciding the appropriate tnc for each patient remains a dare and an art. disclosure: nothing to declare. impact of adding plerixafor to mobilization protocol in the immune reconstitution of vδt cells after autologous hematopoietic stem cell transplantation efrat luttwak , , yael chava cohen , , odelia amit , , irit avivi , , svetlana trestman , , esti rom , , rinat eshel , , ram ron , tel aviv medical center, tel aviv, israel, sackler faculty of medicine, tel aviv university, tel aviv, israel background: multiple myeloma has remained an incurable disease even in the era of novel therapies. front line treatment typically comprises of induction chemotherapy with - cycles of a bortezomib-based regimen, stem cell mobilization, and harvesting of peripheral blood stem cells (pbsc) by apharesis, followed by high dose melphalan with hct. while brotezomib-based induction regimens have demonstrated no adverse impact on hematopoietic cell harvest number and quality, no study analyzed the impact of timing of the last brotezomib dose prior to collection. in this study we aimed to determine the effect of the timing of the last dose of brotezomib before hematopoietic cell collection and the collection yield. methods: this was a single center historical prospective study, including all sequential newly diagnosed patients with myeloma between and that were given a bortezomib-based induction therapy (≤ cycles) followed by pbsc collection. we excluded patients who either received st line vtd-pace or lenalidomide-containing regimens. peripheral blood cd + cells were measured on the day of collection. patients with cd + levels of > cells/ microliter started collection on the same day, while those with lower levels were given plerixafor. we performed regression analyses to analyze the impact of a variety of precollection factors, including days from last bortezomib therapy on the collection yield. results: we identified patients who fulfilled the inclusion criteria, table. median time from last dose of brotezomib to first leukapheresis was (range, - ) days. a statistically significant correlation was found between the days from last dose of brotezomib and both the first collection day-cd + cells/kg (r= . , p< . ), and the total collected cd + cells/kg(r= . , p= . ), figure. the optimal cutoff point as indicated by the roc curve was . days according to collection success with sensitivity of % and specificity of %, youden´s index . . in multivariate analysis included other factors affecting collection yield (age, gender, status of disease at collection, and prior radiation) -timing of last dose of brotizomib remained significantly associated with the total collected cd + cells/kg (p= . ). increasing age, female gender, and prior radiation were associated with lower collection yield (p= . , . , . , respectively). based on this, we developed a model to predict the total collected cd pos cells = . + . (timing in days of last dose of brotezomib) - . (age) - . (if female) - . (≥pr) - . (if prior radation). conclusions: timing of last dose of brotezomib is an important factor for predicting a successful collection. a washout period of days is associated with a better collection yield. these results should be further validated in a prospective study. age (median, range) ( - ) gender -male (%) ( ) prior radiation treatment (%) ( ) disease status at collection (%) ≥pr - ( disclosure: nothing to declare p mobilization with plerixafor in "poor mobilizer" related and unrelated donors of hpc-a in case of failed mobilization with g-csf background: in the allogeneic hpc transplantation, both from related and unrelated donors, the most commonly used source is peripheral blood after mobilization with g-csf. it is however known that about % of donors are "poor mobilizers"; the rescue strategies are: a third apheretic collection; bone marrow donation. methods: in - in italy a procedure to be adopted in case of failed mobilization of peripheral blood stem cells has been defined and shared between ibmdr, cnt (transplant national center) and cns (blood national center) and the scientific societies simti, sidem and gitmo, using plerixafor, a selective reversible antagonist of the cxcr receptor with its binder (the stromal derived factor sdf- ), in combination with standard g-csf dose. moreover since , in accordance with this protocol, the competent authority (aifa) has extended the registration of plerixafor (law no. / ), also for the mobilization in "poor mobilizer" healthy donors. finally, in the protocol was extended to "poor mobilizer" family donors, making management equivalent in related and unrelated donors. at the time of the donor´s informed consent for the donation of hpc, the hypothesis of the lack of mobilization or inadequate collection was illustrated and the possible actions proposed as "back-up donation" were anticipated. failed mobilization of cse has been defined as the presence of one of the two criteria: a number of cd + circulating on peripheral blood lower than /μl on the th day of stimulation ( d), or the collection of cd + < . x /kg weight of the recipient at first apheresis. in these cases, a single dose of plerixafor is administered subcutaneously by health professionals under medical supervision, . mg/kg of body weight, - hours before the start of apheresis. in case of use plerixafor due to failed mobilization of hpc-a or collection of an inappropriate number of cd +, the notification is made by the collection center to ibmdr (for both family and non-family donors), and to the recipient transplant center : both donor and recipient express their consent; finally once the collection is completed, the collection center informs ibmdr, which in turn notifies cnt/cns/simti/sidem/gitmo. any adverse reactions/events are notified in real time, based on the sop specifications and current regulations. results: since the introduction of the national protocol, donors ( unrelated donors and related donors) were treated, presenting at least one of the two inclusion criteria (cd < /μl at d or cd < x e /kg after first collection)after use of plerixafor in all donors, the required dose of cd was obtained to ensure successful transplantation, with a sufficient increase in the cd + cells. no side effects or adverse reactions related to the administration of plerixafor occurred. conclusions: in cases of failed mobilization in the related and unrelated donor, the use of plerixafor according to the methods described in the shared protocol between ibmdr, cns, cnt, simti, sidem, gitmo, proved to be safe and effective. this protocol emphasizes the great value of the sharing of procedures between the register, institutions and scientific societies, ensuring the supervision of the process and the protection of the donor and recipient. disclosure: nothing to declare background: autologous stem cells transplantation (asct) is an effective treatment option for young patients with multiple myeloma (mm). a minority of patients may still experience untoward toxicity due to delayed engraftment. thus, the current policy in many centers is aimed to increase the target dose of collected cd + cells up to an "optimal" level of x /kg per procedure. therefore, an ideal mobilization, aimed to collect to cd + cells/kg in one apheresis, should achieve a number of circulating cd + cells > /mcl (very good mobilizers). plerixafor may help to maximize the cd + collection but its use is limited by high cost. we carried out a retrospective analysis aimed to predict the quality of mobilization and develop an algorithm to optimize both timing of collection and use of plerixafor. methods: we retrospectively collected data from mobilization procedure performed in our center between and for mm. all received the same mobilization protocol with cyclophosphamide (range - gr/sqm) and g-csf mcg/kg from + . cd + cell count was started when white blood cells (wbc) count exceeded x /l. patients were excluded from this analysis if ) showed a cd + count > /mcl (target achieved at first day count) and/or ) cd + count on second day was missing and/or ) plerixafor was administered on first day according to previous policies. sixty-eight patients were evaluable for the study. univariate and multivariate logistic regression analysis to study ccd + kinetics and assess predictors impact on mobilization was carried out. ratio cd +/wbc in first day count, gender, disease category and time from mobilization chemotherapy were also included results: among the patients included in the analysis, the threshold of cd +/mcl cells on the second day was reached by ( , %) of patients (groupa) whilst the remaining ( , %) failed the target (groupb). median (range) wbc x /l and cd + /mcl counts in group a and b were , ( - , ) and , ( , - , ), , ( ) ( ) ( ) ( ) ( ) ( ) ) and , ( , - , ) respectively, with a statistically significant differences among group (mann-whitney p= , and p= , respectively). only cd + /wbc ratio and cd + /mcl on first day count had an impact on kinetics and optimal mobilization. logistic regression model highlight cd +/mcl (or= , ; % ci: , - , ) on first count as an independent predictor of second day optimal mobilizer, with auc of . % ( , ) in roc analysis. two cd + thresholds were then calculated: < , /mcl (ppv , ; npv , ) that identified poor mobilizer, and ≥ , /mcl (ppv , ; npv , ) that exclude probability to fail on second day. for those with a cd + count between , - , the cd +/wbc ratio (or= , , % ci: , - , ) was a predictor of optimal mobilization (auc , ; , - , ); cut-off value was , (sensibility , ; specificity , ) conclusions: assessment of circulating wbc, cd + and their ratio at wbc recovery in a chemo-based mobilization is a valid tool to manage the collection strategy and the on-demand use of plerixafor. we have developed an algorithm aimed to the use of plerixafor to both rescue poor mobilizers and boost cd + count in intermediate mobilizers. background: successful autologous stem cell transplantation (asct) requires the infusion of a sufficient number of hematopoietic stem cells (hscs). peripheral blood (pb) is the most commonly used source of hscs, therefore, it is important to optimize methods used to mobilize the hscs. the most clinically used chemotherapeutic agents for effective mobilization are cyclophosphamide and etoposide. recent published studies suggest that etoposide has a better mobilization effect than cyclophosphamide even at lower doses, but it is not clear why this difference occurs. in this study, we tried to determine whether there is a difference in the mechanism of mobilization between cyclophosphamide and etoposide. methods: first, in order to confirm the clinical data for efficacy and toxicity of mobilization, we retrospectively analyzed the data of patients who were diagnosed with lymphoma and performed mobilization using cyclophosphamide or etoposide from january to december . second, mesenchymal stem cells (msc) were primarily cultured from the healthy controls, then treated cyclophosphamide or etoposide at a concentration of % inhibition of cell growth, and cytokine analysis was performed to identify cytokines known to be associated with mobilization. third, mobilization mouse model using cyclophosphamide or etoposide was generated, total blood was collected at the time of hscs collection, and cytokine and network analysis (using ingenuity pathway analysis) was performed. results: the mobilization yields for cyclophosphamide or etoposide were analyzed. etoposide miblized a significantly higher median number of cd +cells than cyclophosphamide. the rate of successful or adequate mobilization was also significanctly higher for etoposide in univariate and multivariate analysis (table ). in the analysis of toxicity during mobilization, the incidence of neutropenic fever was higher in the cyclophosphamide group (p = . ). during mobilization, cyclophosphamide maintained lower wbc counts than etoposide and showed a large increase in wbc counts at the start of collection ( figure ). the cumulative dose of cyclophosphamide or etoposide in patients who underwent autologous stem cell transplantation did not affect leukocyte (anc> /microl or platelet (plt > k/microl) engraftment. in msc treated with etoposide at a concentration of % inhibition of cell growth, il- , which is a cytokine that promotes hematopoietic stem cell mobilization, were shown a statistically significant increase (figure ). in the mouse model of mobilization (figure ), the levels of kc, one of the il- homologues in mice, had significantly increased in the etoposide-treated group compared with the levels in the cyclophosphamide-treated group. the levels of other il- homologues, mip- and lix, also showed increases in the etoposide-treated group compared with those in the cyclophosphamide-treated group; these differences, however, were not statistically significant (figure ). network analysis based on in vivo cytokine results identified that etoposide could promote mobilization in association with matrix metalloproteinase as compared to cyclophosphamide ( figure ) . conclusions: etoposide has a higher mobilization efficacy when compared to cyclophosphamide, which could due to the different mechanisms of mobilization through the elevation of il and the activation of matrix metalloproteinase associated therewith. background: high-dose chemotherapy followed by autologous blood stem cell transplantation (asct) is a standard therapy for wide range of hematologic and solid malignancies. although various methods have been introduced to improve the peripheral blood stem cell (pbsc) mobilization, autologous stem cell collection (ascc) is not successful in every patient. furthermore, even if the ascc is complete, not all of them lead to asct. we evaluated the result of ascc and actual use of pbsc grafts in current practical setting. methods: we retrospectively reviewed the all consecutive ascc procedures performed at the department of oncology in asan medical center, seoul, korea, between january and october . the targeted number of background: fanconi anemia (fa) is a rare inherited genetic bone marrow (bm) failure syndrome. while abnormal bm cells production occurs very early in life, the usual age of diagnosis is - years old. gene therapy (gt) might be an alternative to hematopoietic stem cells (hsc) transplantation, but harvest a large number of autologous hsc remains a challenge. we started a mobilization assay, fancomob, to evaluate the safety and the efficacy of fa patients' mobilization with granulocyte-colony stimulating factor (g-csf) and plerixafor. this study is part of the fa's european gt project "eurofancolen". methods: four patients with fanca mutations following the inclusion criteria were selected before pancytopenia. to note, fa was diagnosed before clinical manifestation through family screening. they received subcutaneous injection of g-csf ( μg/kg twice a day) from d- and plerixafor ( . mg/kg/day) from d- . the collection protocol targeted x e /kg of cd + cells, based on a predicted future weight in years. cd + cells and white blood cells (wbc) blood count were monitored tightly along the mobilization. patients with more than cd +/μl or between and cd /μl with a clustered aspect detected by flow cytometry after plerixafor injection underwent apheresis. cd + cells were immunoselected from the collection with clinimacs purification system (miltenyi) and cryopreserved for further gt manipulation. results: the mobilization target was not achieved for the first two included patients (fa years old and fa years old). the minimum value of cd +/μl required wasn't obtained for fa and the flow cytometry cd + aspect was not clustered for fa . cd + cells were mobilized quickly but transitionally after plerixafor injection for the last two patients, fa and fa , and years old respectively. both patients underwent apheresis procedures. no cd + cell rebound was observed after the apheresis was stopped.collection target was not achieved after four days of collection for fa . it was obtained the first day for fa (figure ). back-up for hsc transplantation could not be cryopreserved because of the limited number of cd + cells collected in patients fa and fa . no short-term adverse events were observed. following cd + immunoselection, cd + cell purity and recovery were poor but in the normal range described in the literature for fanconi patients ( - %)( table i) . one month after the collection hemograms were unchanged. conclusions: our clinical study offer new data showing that mobilization of fa patients with g-csf and plerixafor is safe and more efficient for younger patients, especially before clinical manifestations of bm failure. further efforts are required to establish an effective technic to purify the cd + cells after harvesting. basal cd + cell and platelets count are a strong predictor for mobilized peripheral blood stem cells on the th day of g-csf treatment in donors cryopreserved pbscs. this was associated with considerable efforts for the patients and caused additional treatment costs. on the one hand, having the therapeutic option of an autologous transplantation in the future may represent a clinically relevant advantage. however, a huge number of stem cell products are kept in storage for many years without ever been used for transplantation. our study provides cause for a careful reevaluation of the current clinical practice, which may help to focus more precisely on patients who actually benefit from a cryostored autologous stem cell graft. [[p image] . fig. : absolute numbers and relative distribution of stem cell grafts] disclosure: the authors confirm that there are no potential conflicts of interest to disclose, except the following: katharina kriegsmann: research funding from bms, celgene, and sanofi. patrick wuchter: membership in advisory boards for sanofi-aventis. reduction of dimethylsulfoxide (dmso) concentration from % to % in criopreservation of stem cells. influence on the kinetics of engraftment and tolerance to infusion patricia lopez-pereira , beatriz aguado , elena sola , carmen cámara , isabel vicuña , lorena vega , adrian alegre hospital universitario de la princesa, madrid, spain background: dmso is the cryoprotectant most used in the cryopreservation of stem cells. it is associated with adverse effects during the infusion of the product, its toxicity being proportional to the volume infused. the most common concentration used has been %, although recent publications report that reducing it to % leads to lower rate of side effects without impact on the product or the graft. we retrospectively analyzed patients recipients of autologous peripheral blood stem cell transplantation (hct) in our hospital from january to september . they are divided into two groups according to the concentration of dmso used in the freezing ( % until september or % since october ). the baseline characteristics of the patients, the infused product and the graft are shown in table . the cd count was performed by flow cytometry. all freezings were performed with a biological freezer for programmed controlled rate cryopreservation and stored in ultrafreezers at - ºc. results: both population groups are homogeneous. the t-test was used for statistical analysis. regarding the cd + variable, no statistically significant differences were observed (p = . ). neither for the variables leukocyte recovery and platelet recovery (p = . , p = . respectively). the difference in the variable viability is . units (ci %: [ . - . ]) and is statistically significant (p = . ) in favor of dmso %. regarding adverse effects, % (n = ) of the serious adverse reactions occurred in the % dmso group (hypotension and seizures). the mild and moderate ones were similar in both groups, most were mild nausea, vomiting and flushing. overall, no statistically significant differences were observed due to the low rate of adverse effects found. patients starting with until october , total of patients attempted collection of autologous pbscs, and poor mobilizers recieved plerixafor during first mobilization cycle. in total, patients required repeated mobilization cycles ( , %) of which were from the plerixafor group. in total patients recieved pleriksafor; females and males, median age ( - ) with following diagnoses: nhl, mh, multiple myeloma, neuroblastoma, nephroblastoma, sarcoma ewing and seminoma. of repeated mobilizations with plerixafor, patients ( , %) still failed to collect adequate transplant. in this period we had altogether unsuccessful mobilizations ( , % in repeated cycles, , % in total). this group of patients consisted of male and female patients, median age ( - ), diagnosis of nhl and failure to collect after leukapheresis procedures each. median number of leukapheresis needed for adequate collection was with preemtive plerixafor use, and in repeated mobilizations. conclusions: our expirience shows that preemtive use of plerixafor in poor mobilizers is efficient and has enhached success of the pbsc collections. due to drug high cost each institution needs to develop its own algorythm in management of poor mobilizers. the factors contributing to plerixafor mobilization failure still need to be elucidated. disclosure: nothing to declare. platelets recovered from mobilized leukapheresis units obtained from hla-haploidentical donors fulfill the criteria of a conventional hemocomponent and can be used for transfusion background: central venous catheter (cvc) related complications may lead to high morbidity and mortality. unlike cvc, peripheral cannulation offers a quick and inexpensive method for safe and non-traumatic vascular access (va) thus its utilization is strongly recommended whenever possible. the ultrasound (us) guidance for acquiring peripheral va is a useful tool for reduction or elimination of the need of using cvc for stem cells collection. we have made an attempt to introduce us method in our apheresis unit having no previous experience with us devices. the aim of the study was to measure the decrease of cvc insertions after introducing us and evaluate the quality of va by comparing average flow rate and confirming that the desired blood volume could be processed. methods: the theoretical education involved a free elearning course in peripheral ultrasound-guided va (pugva, usabcd, aarhus, denmark). subsequently, the personnel have implemented knowledge in practical training on gelatine and silicone phantoms and healthy volunteers. the practical activities also included a fiveday course in an apheresis centre with us-guided cannulation experience. the details concerning va were recorded, including va site, cannula size, average inlet flow rate, number of inlet pressure alarms reported by the apheresis device. the procedure details where traditional approach was applied i.e. palpable cannulation and cvcs have been collected. similarly, the necessary data for procedures where veins were assessed with ultrasound prior to apheresis were recorded. results: before introducing ultrasonography, stem cell collections have been performed in patients. of all these procedures, were accomplished with cvc ( %) and with peripheral va ( %). median cubital vein was the vessel of choice. out of the peripheral va procedures, ( %) were problematic, with or more inlet pressure alarms during every procedure. after the training stage, collection procedures were performed in patients. after introducing us we have observed a significant reduction of the number of cvc insertion required for successful apheresis from % to % (p= . ; chi-square test with fisher's exact). thirty one procedures were completed with peripheral va ( %). ultrasound device enabled cannulation not only the superficial veins but also for the deeper veins. cannulation sites included upper arm cephalic vein ( %), median cubital vein ( %), upper arm basilic vein ( %), median antebrachial vein ( %). out of the collections, were considered problematic ( %). no difference in an average flow rate was observed between procedures performed peripherally with and without ultrasound usage (p= . ; u mann-whitney test). conclusions: despite no previous experience in us guidance, we have successfully managed to introduce the new method in our apheresis unit. within months, we have reduced cvc usage threefold and as the personnel is gaining more experience, we suppose that the cvc usage may be reduced to episodic cases. despite slightly higher number of pressure alarms, all procedures with ultrasound guidance were completed as planned. ultrasound guidance is the most important tool for significant increase in peripheral va usage and may become the only option for patients with difficult va. disclosure: nothing to declare. abstract already published. donor blood management in healthy bone marrow donors: a retrospective single institution analysis background: over the last two decades mobilized peripheral blood stem cells (pbsc) have been established as the main source of stem cells because of improved engraftment and no necessity for hospitalization for the donors. nevertheless, due to the introduction of promising new transplant regimens, especially in the haploidentical transplantation setting bone marrow (bm) donations are regaining importance. although for both donation methods severe side effects are rarely described, bm collection is associated with considerable blood loss and hence symptoms of acute blood loss are commonly observed. therefore autologous blood are collected routinely in some institutions before donation. since the collected bone marrow amount depends on the target dose, the wbc yield in the product influences the required bone marrow volume. therefore we sought to investigate the relationship between collection volume, rbc volume removal, drop in hb and indications for blood transfusion. furthermore, we assessed wbc and cd +yields in relationship to various donor parameters and to product volume, in order to find prediction tools for collection volumes. methods: allogeneic bone marrow harvests from adult donors were performed at our institution and retrospectively analyzed. complete blood counts, serum iron and ferritin were assessed at work-up and weeks after donation. the bone marrow product quality including wbc, hematocrit (hct) and cd + cells were assessed by automatic hemocytometry and single-platform flow cytometry with ishage gating. results: besides local pain most of the side effects were related to blood loss. none of the donors received blood transfusions. the mean reduction of hemoglobin levels was . g/dl with a minimum hemoglobin level of . g/dl and a persistent anemia according to who criteria after weeks in . % and pathologically low ferritin levels in %. no donor presented symptoms with indication for blood transfusion. the median wbc concentration of the bm product was . /nl ( - % percentile: . - . / nl) the cd +cell concentration . /μl ( - % percentile: . - . /μl). in the linear regression analysis leukocyte counts of the donor before donation correlated significantly with wbc concentration in the product. thus in order to collect with % certainty the mio wbc which are a typical per-kg dose for an allogeneic recipient, . ml of bone marrow must be collected. collection volume did not systematically affect wbc or cd + cell concentration. conclusions: achieving high wbc yields in the bone marrow product allowed for collection of relatively modest bm volumes, thus protecting donors from excessive blood loss. acute adverse events were acceptable. optimization of perioperative management in healthy bone marrow donors may be achieved by good collection technique and reevaluation of wbc yields of each institution to calculate required bone marrow amount. the collection of autologous blood is not indicated. furthermore stringent pre-and postoperative hemoglobin management is predicted to limit adverse effects. disclosure: nothing to declare. donor-recipient weight ratio predicts successful stem cell mobilization on day four of gcsf mobilization results: in group median age of donor was years (range to years). in group median age of donor was years (range to years). table] . table ] elaborates other parameters analyzed between the two groups. one patient in group developed grade ii acute gvhd whereas patients in group developed acute gvhd grade ii-iv. at the last follow up no ( / ) patient in group has any symptoms of chronic gvhd whereas ( / ) patients in group have features of chronic gvhd (one extensive, one limited). conclusions: our observation suggests that upfront use of plerixafor in combination with gcsf modifies the graft favorably decreasing the risk of graft failure and graft versus host disease both acute and chronic. it also helps the donor by decreasing the total volume processed, amount of acd exposure and the duration of harvest. disclosure: none. impact of vitamin d levels on peripheral stem cell mobilization in autologous hematopoietic stem cell transplant recipients ferda can , zeynep arzu yegin , zubeyde nur ozkurt , orhun akdogan , lale aydın kaynar and total product cd + cell count [ . ( . - ) vs . ( . - . ); p= . ] were significantly higher in patients receiving chemotherapy+g-csf than g-csf only. the study group was divided into two groups based on peripheral cd (cut-off level: x /kg) as well as product cd levels (cut-off level: x /kg). vitamin d levels were found to be similar among these groups (p> . ). total product cd + cell count was found to be relatively lower in patients with vitamin d levels below μg/l [ . ( . - ) vs . ( . - . ); p= , ]. (figure ) conclusions: based on its effect on stem cells in in vitro studies, it may be considered that vitamin d may have a favourable impact on stem cell mobilization. statistically insignificant but relatively lower total product cd + cell count in patients who had lower vitamin d levels, which may indicate a role for vitamin d in stem cell mobilization, needs to be confirmed with larger studies. considering the high prevalence of vitamin d deficiency in the general population, the possible role of vitamin d in hematopoietic stem cell mobilization deserves further consideration. disclosure: nothing to declare background: one of the factors, affecting efficiency of autologous hematopoietic stem cell transplantation (autohsct) in hodgkin lymphoma (hl) patients is early recovery of graft, depending on cd + cell count and conditions of cell product cryopreservation and storage. it is well known, that dimethylsulfoxide (dmso), used for cryopreservation, can be cardiotoxic and cause diverse gastrointestinal, pulmonary, kidney, liver side effects and acute hemolysis. lethal for animals dose - mg/kg leads to life threatening arrhythmias and respiratory arrest. in order to improve dmso toxicity different ways of alternative cryoconservation modes are studied -lower dmso concentration ( % vs %), temperature - ˚c instead of ultra-low and washing of cell product. aim of the study is to evaluate the influence of dmso washing on hematopoietic recovery after autohsct. methods: retrospective analysis of hematopoietic recovery of relapse/refractory hl patients after autohsct was performed. mobilization regimen included second line chemotherapy for hl (dhap, begev, igev, ice) with consecutive g-csf administration. cd + cells were assessed, using -colour flow cytometer facs canto ii while cell collection, thawing and washing. cells with % dmso were stored at - ˚and washed in cases of transplantation with human albumin-dextran (reopolyglukin) and centrifugation. statistical data processing was performed by the χ method -pearson criterion; p -the level of significance of differences. results: patient groups had no difference in age, disease stage, gender, time from treatment start to autohsct and cd + cell count (p> , ). time to wbc recovery > х /л was - (median , ) days vs - (median , ) days, time to platelet> х /л recovery was - (median , ) days vs - (median , ) days in groups without and with cell washing respectively (p= , ). no difference in blood component consumption was observed (p= , ). in out of ( %) patients during cell reinfusion without washing nausea, vomiting, arterial hypertension was observed, no reactions were detected after cell washing (p = , ). conclusions: washing autologous mononuclear cells from cryopreservant dmso does not lead to low hematopoietic recovery rate after autohsct and can avoid toxicity, thus making autohsct more safe. disclosure: authors declare no conflict of interests. quality assesment of hematopoietic stem cells autografts after cryostorage, harvested using plerixafor background: the introduction of high-dose chemotherapy followed by transplantation of autologous hemopoietic stem cells (hscs) into the treatment program for multiple myeloma (mm) has significantly increased the frequency of achieving complete remissions and overall survival in patients. to obtain a sufficient amount of hscs, hematopoiesis is stimulated with granulocyte-macrophage factors (gm-csf) both in mono mode and after the administration of cytostatics followed by cytapheresis sessions (alone or after the cytostatics followed by cytapheresis sessions) . cryopreservation protocols are used to preserve cells in a viable state, followed by long-term storage of transplants in liquid nitrogen. however, in some patients it is not possible to obtain the necessary amount of hscs. the inclusion of plerixafor in standard mobilization schemes allows you to prepare the sufficient quantity of hscs in most patients with mm. methods: the study included samples of autografts from patients with mm from to (median . ± . ). hscs mobilization was performed on the background of unstable blood formation after high doses of cyclophosphamide g/m with the subsequent administration of g-csf at a dose of - μg / kg ( samples from patients) and with the addition of plerixaphor at a dose of . μg / kg ( samples from patients). immunophenotype viability of hscs in autotransplants after cryopreservation were determined by flow cytometry using the ishage protocol on a flow cytometer (facs cantoii, becton dickinson) by expressing surface markers of antibodies against cd , cd , cd , cd and staining with aminoactinomycin ( -aad). the colony-forming activity of hscs (cfu-cfu-mix, cfu-gm, cfu-g, cfu-m) was evaluated in methylcellulose (methocult h , stemcell technologies, canada) for x transplanted cells for days. results: the viability of hscs in autografts (cd + / cd + / add-) after cryopreservation in both groups was ± . %. in the group of samples using plerixaphor, a higher content of primitive hemopoiesis precursors (primitive cells) (cd + cd + cd -) was detected compared with the control group ( . ± . % and . ± %, respectively). the cfu count (cfu-cfu-mix, cfu-gm, cfu-g, cfu-m) in the plerixafor group was ± . per x explanted cells, in the control group - ± . ( figure a-d) . conclusions: the use of plerixafor against the background of standard protocols for the mobilization of hscs allows to obtain high-quality graft with a higher content of primitive cells and proliferative activity. disclosure: no conflict of interest. nothing to declare. comparison of effectiveness of plerixafor plus g-csf in poor and very poor-movilizers: efficacy of the combination of plerixafor and g-csf in poor-movilizer background: healthy donors ocassionally show a poor response to mobilization agents. plerixafor+g-csf can be a salvage strategy in poor mobilizers. some series describe the use of plerixafor to collect greater doses of cd + cells in hematopoietic stem cell transplantation (hsct) with tcell depletion. plerixafor use in the mobilization protocol could help collecting higher cd + dose in indirect t-cell depletion (cd + selection) for ex-vivo manipulated haploidentical transplantation, with less number of apheresis and a rapid engraftment. methods: data of fourteen healthy peripheral-blood donors was retrospectively collected. they received days mcg/kg/day g-csf and , mg/kg/day plerixafor on º day as mobilization treatment. fourteen pediatric patients (median age years, range - ) diagnosed with malignant and no malignant hematological diseases received haploidentical hsct with cd + selection and cd ra+ depletion between february and july . results: one leukoapheresis procedure was performed in all cases. median processed volume was liters (range - ). median of cd + cells obtained was , x /kg (range , - , ) . after positive selection, > x /kg cd + cells were infused in all cases (figure ). neutrophil engraftment was achieved after a median of days (range - ). few donors presented only plerixafor mild secondary effects. conclusions: our experience showed that a mobilization protocol using g-csf and standard dose of plerixafor (compasive use) is a safe strategy that allows collecting great cd + dose in one apheresis procedure. this could be useful for haploidentical transplantation with ex-vivo t depletion, especially if there´s a weight disproportion between donor and patient. background: mesenchymal stem cells (mscs) are selfrenewing multipotent progenitor cells with wide differentiation potential. their ease of isolation and expansion in vitro as well as their unique regenerative therapeutic properties suggest the use of msc as an approach for treating several disorders. extra-embryonic tissues as placenta have been proposed as potential sources of mscs due to the absence of ethical problems neither risks for the patients. furthermore, only protocols using fresh placental tissue have been described so far. a protocol for isolating mscs from delayed-manipulated tissue was designed and tested in order to optimize the use of placental mscs (mscs-p) in an advanced therapies context. methods: full term placentas (n= ) were obtained from healthy mothers in hospital universitario central de asturias (spain). informed consent was obtained from each mother prior to delivery. after dissection of gr decidual tissue it was washing with saline (b. braun, germany) and cut into small pieces. these biopsies were conserved hours in dmem media with % antibiotic solution x (gibco, usausa) until processing. the day after, tissue was mechanically minced and then enzimatically digested with a combination of ui/ml dnase i (sigma aldrich, usa) and . % tripsin-edta solution (w/v) (biochrom, germany) at ºc for hour. then, the mixture was filtered with μm cell strainer (bd bioscience, usa) and centrifuge at xg for minutes. finally, cells were resuspended in ml of dmem media suplemented with % fbs and antibiotic, seeded in -cm flask and incubated in forma stericult co incubator (thermo fisher scientific, usa) at ºc, %co . culture-expanded mscs cells were phenotipically characterized by flow cytometry (facs aria iiu, bd) with antibodies against cd , cd , cd , cd , cd , cd , cd , cd , hla-dr cd and cd using mesenchymal cell kit (immunostep, spain). afterwards, these cells were differentiated to adipogenic, osteogenic and chondrogenic lineages using stemmacs adipodiff media, stemmacs osteodiff media and nh chondrodiff medium (miltenyi biotec, germany) respectively. after three weeks of differentiation cells were fixed in % paraformaldehide (merck, usa) and analyzed. adipogenic, osteogenic and chondrogenic differentiation was visualized after staining with oil red o, alkaline phosphatase and hematoxilin-eosin (sigma-aldrich, usa). results: mscs-p isolated cells were characterized according to the isct criteria for mesenchymal stem cells. they were positive for cd , cd , cd , cd and cd and negative for cd , cd , cd and hla-dr, indicative of a typical msc phenotype ( figure ). all the markers showed a high percentage of expression between . and . %, meaning that msc population obtained with the designed method was very homogenous. similarly, staining for the three studied lineages was positive ( figure ). conclusions: the described protocol allows us to obtain mscs from decidual placental tissue stored and processed hours after the biopsy extraction using a unique enzymatic digestion. this circumstance permits to take advantage of placentas that are discarded after delivery giving us the option to obtain mesenchymal cells that could be used in clinical trials. disclosure: nothing to declare outcomes of umbilical cord transplant in high risk relapsed or refractory acute myeloid leukaemia background: high-risk relapsed/refractory acute myeloid leukaemia (aml) is a fatal disease. allogeneic haematopoietic stem cell transplantation represents the only chance of cure. as the transplant relies on the graft-versusleukaemia (gvl) effect, and if different donors exert different gvl effects, then choosing the right donor assumes great importance. in manchester, a large bmt centre in the north of england, our practice in such aml has been to choose unrelated cord blood (ucbt), without serotherapy in the conditioning therapy, as our preferred donor cell source. methods: we report the results of unrelated ucbt in patients (five boys and ten girls) with high-risk aml, defined as relapsed or refractory disease. thirteen patients ( %) received this as a st transplant, two patients ( %) received this as a nd transplant for relapsed aml post matched unrelated donor transplant, and one ( %) received ucbt twice, once in cr and once in cr . nine patients ( %) had mismatched ucbt, and the rest were fully matched at class-i (hla-a, -b, and-c) and class-ii (hla-drb ). conditioning was given as treosulfan, fludarabine and thiotepa in half of the patients (n = ), other treosulfanbased regimens were used in two patients ( %), and busulfan-based regimens were used in six patients ( %). no serotherapy was given. results: the median age at transplant was years (range, months - years). neutrophil and platelet engraftment were achieved in and patients at a median of and days, respectively. patients ( %) had engraftment syndrome. all engrafted patients achieved % donor chimerism, except one patient who had mixed lymphoid chimerism initially, that was corrected spontaneously to % at three months after transplant. acute gvhd grade i-ii developed in six patients ( %), and grade iii-iv developed in three patients ( %). all cases resolved, except two patients where acute gvhd evolved into chronic gvhd (one with grade i skin gvhd which fully resolved, and one with grade iii gvhd gut colitis who was parenteral nutrition dependent till death). two more patients developed chronic grade i skin gvhd and resolved (chronic gvhd developed in % in total). three patients ( %) developed veno-occlusive disease (vod), that completely resolved with defibrotide treatment and necessitated ascitic drainage in one of them. viral reactivations occurred in five patients ( %) and were successfully treated. at a median follow-up of months (range, seven months -four years), eight patients ( %) died at a median of (range, to days), with a transplantrelated mortality of % and relapse-related mortality of %. five patients ( %) relapsed post-ucbt; four died and one had a successful second ucbt (event-free survival was %). immune reconstitution in alive patients was achieved at a median of eight months. conclusions: very high-risk patients treated with ucbt with good overall survival and event-free survival, similar to aml treatment rate with low-risk disease. disclosure: nothing to declare in haploidentical transplants is the incidence of acute and chronic gvhd strictly related to the stem cell source? results: the odds ratio was . with a % confidence interval of . - . (p= . ). conclusions: the risk of infection of the uc is not related to the microbiological status of the ucb. a possible explanation for this is the presence of antibiotics in the medium used for uc, but not ucb, transport. this means that cryopreservation of ucs from which contaminated cord blood has been obtained is justified. comparison of turkish stem cell coordination center (turkok) with istanbul university bone marrow bank (tris); a single center experience in match unrelated donors azize mergen , selime aydoğdu , başak aksoy , yunus emre savcı , gürcan dikme , funda Çipe , ceyhun bozkurt , tunç fışgın older patients are increasingly being transplanted, thanks to improvement in allogeneic hematopoietic stem cell transplantation (allo-hsct) techniques. increasing donor age is associated with greater risk for mortality and graftversus-host disease (gvhd). since sibling donors are of similar age to recipients, we hypothesized that, in older patients, a young matched unrelated donor (mud) would be comparable to an hla-matched sibling donor (msd). methods: we retrospectively compared outcomes of allo-hsct from msd (n= ) and / hla mud (n= ) in patients aged ≥ years with hematological malignancies transplanted between - . all patients received reduced-intensity conditioning and graft source was peripheral blood. the primary outcome was overall survival. msds served as the reference category and were compared to muds split into three age groups (≤ [n= ], - [n= ], > [n= ] years) using univariable analyses and multivariable cox regression models adjusted for patient, disease, and transplantation features. results: the median age of hsct recipients was years and was similar across groups. median donor age for msd was years and , , and for the mud age groups ≤ , - , and > years. acute leukemia was the leading transplant indication ( %) followed by myelodysplastic syndrome, myeloproliferative neoplasms and indolent non-hodgkin lymphoma. disease risk distribution was similar across donor groups (low [ %], intermediate [ %] , and high [ %] in the complete population; p= . ). time from diagnosis to hsct was longer with mud compared to msd and increased with an older age of mud. in a univariate analysis, overall survival was % (msd), % (mud≤ ), % (mud - ), and % (mud≥ , p= . ). corresponding non-relapse mortality (nrm) cumulative incidence was %, %., %, and . % (p< . ) (figure) . gvhd-relapse-free (grfs) was %, %, %, and % (p= . ). in a multivariable cox model, young mud (≤ ) had a similar risk for mortality compared to msd (hr . , p= . ), while a monotonic increase in risk was observed with an older donor age (mud - y: hr . ,p= . ; mud≥ y: hr . , p= . ) (table) . findings were confirmed in a propensity score analysis, matched for key covariates. nrm and grade - acute gvhd were consistently higher with mud, with the greatest risk associated with older muds. the hazard for grfs was higher with mud aged or higher compared to msd; risk was not higher with younger mud. conclusions: in older patients receiving reduced intensity conditioning, msd remain the optimal choice. however, when not available, young mud provide comparable results. disclosure: nothing to declare background: there is growing evidence that community acquired respiratory virus (carv) increase the risk of pulmonary invasive fungal disease (ifd) in recipients of allogeneic hematopoietic stem cell transplantation (allo-hsct). to date, there is a lack of knowledge regarding the rate of ifd, risk factors (rfs) as well as the most critical period for the development of a later ifd after carv infections in allo-hsct recipients. methods: in this prospective observational study, we retrospectively analyzed the effect of carv on the development of a later ifd in a consecutive cohort of allo-hsct adult recipients who developed carv infectious episodes from december to december . respiratory virus in upper and/or lower respiratory tract specimens were tested using multiplex pcr panel assays. results: overall, out of allo-hsct recipients ( %) developed ifd within months after a carv episode at median of days (range - days) from the day of carv detection. all the ifds involved the lungs and in cases ( %) the diagnostic was ia accomplishing criteria of probable (n= ) or proven (n= ). of note, out of ifd ( %) occurred within the first year after transplantation. the overall rate of ifd after carv episodes was % whereas this rate was higher in recipients developing carv during the first year of transplant ( %). ifd was diagnosed in out of with carv lower respiratory tract disease (lrtd) episodes ( %) compared to out of carv upper respiratory tract disease (urtd) ( %) (p= . ). twenty-three out of carv episodes involving the lrtd during the first year after transplant ( %) developed ifd. we did not found differences in ifd rates according to the type of carv identified. multivariate analysis identified rfs for ifd: the use of atg as a part of conditioning [odds ratio (or) . , % confidence interval (c.i.) . - . , p= . ], carv lrtd (or . , % c.i. . - , p= . ), carv infection during the first year of transplant (or . , p natural killer cell alloreactive haploidentical stem cell transplantation for multiple myeloma patients catharina elssen , lotte wieten , peter von dem borne , ellen meijer , gerard bos maastricht university medical center, maastricht, netherlands, leiden university medical center, leiden, netherlands, amsterdam university medical center, location vumc, cancer center, amsterdam, netherlands background: in the past years many new drugs for multiple myeloma (mm) have been developed and are responsible for a increase in survival. notwithstanding such progress, mm remains incurable. results from allogeneic stem cell transplantation (sct), including haploidentical transplantation, in mm has shown clinical results. however, these responses are only observed in a minority of patients. we hypothesize that this observation might be due to differences in natural killer (nk) cell alloreacitvity, since we have shown in in vivo and in vitro models that mismatched alloreactive nk cells hold the capacity to kill mm cells. the aim of this prospective phase study is to evaluate if kir-ligand mismatched haploindentical bone marrow transplantation (bmt) with post-transplant cyclophosphamide will improve progression free survival (pfs) in poor risk mm patients. methods: poor risk mm patients, aged < years were enrolled if they were responsive to their last line of therapy. poor risk was defined as, high-risk cytogenetics, or relapse within a year after autologous sct, or treated with three or more previous lines of therapy. a prerequisite of enrolment was the possibility of an nk cell mismatch and availability of a mismatched family donor. patients were excluded if donor-specific hla-antibodies were present. patients received a haploidentical bmt with a non-myeloablative conditioning regimen and post-transplant cyclophosphamide. primary endpoint is pfs at , years. secondary endpoints are engraftment, bone marrow reconstitution, nk cell reconstitution and repertoire, graft versus host disease (gvhd), infections and non-relapse mortality (nrm) at , years. results: in total poor risk patients were included in the study of which could be evaluated for the primary end point. graft failure and disease progression before transplant rendered the remaining two patients not evaluable. at this interim analysis patients have already reached the , years of follow up, relapsed within , years and died due to treatment related infections, without showing progression of disease ( % nrm). average time of progression is days ( - days). two of the remaining patients at follow up, still show responsive disease (days en ). the average time to neutrophil reconstitution is days ( - days). all evaluated patients ( / ) show nk cell reconstitution with a mature phenotype in the bone marrow and peripheral blood by day . three patients developed acute gvhd ( %) of which / grade i-ii agvhd and / patient showed a grade iv agvhd. treatment related mortality was / ( %), which was in all cases due to infectious disease. conclusions: our interim analysis of mismatched haploidentical bmt in mm showed that the treatment is feasible and forms a possible platform for immunotherapeutic strategies. the majority of patients showed an early disease progression. we predefined that with a pfs of % at , years we would qualify this treatment option successful. with only two patients still in remission this goal will not be achieved and we hypothesize that the late nk cell reconstitution (day ) is responsible for the lack of response. clinical background: mscs are known to have immune modulatory capacity and may be effective in the treatment of patients with acute gvhd. however clinical studies yielded inconclusive results which was in part due to the great heterogeneity of the msc used. the off-the-shelf msc preparation "msc-ffm", generated by a proprietary pooling process, selection by plastic-adherence, expansion for an aggregate four weeks followed by cryopreservation until use, is available in germany through a national marketing authorization. "msc-ffm" is indicated in steroidrefractory agvhd, dosed at - x /kg bw i.v. in four doses one week apart. methods: we report seven consecutive pediatric patients (median age . y), who received "msc-ffm" from unrelated hla disparate donors between december and november in our institution. we gave msc infusions to patients with steroid-refractory grades iii-iv agvhd and one patient who had therapy-refractory background: regulatory t cells (treg) are known for their immunosuppressive function and have proven successful as graft-versus-host disease (gvhd) prophylaxis after allogeneic bone marrow transplantation in a number of preclinical as well as first clinical studies without compromising graft-versus leukemia (gvl) effects. in murine models of acute gvhd lymph node homing capacity via cd l (l-selectin) proved to be essential for disease prevention. yet, treg recruitment from lymph nodes to peripheral sites of ongoing gvhd also seems necessary to achieve maximum protective as well as therapeutic effects. the chemokine receptor ccr directs activated t cells to sites of inflammation, thus high ccr expression should facilitate treg homing to affected gvhd target organs. with this project we lay the foundation for future in vivo studies of treg therapy for gvhd by upregulation of ccr expression. methods: we performed systematic ex vivo analysis of ccr expression on murine naive and memory conventional (tconv) and regulatory t cells isolated from spleen, blood, bone marrow, lymph nodes, liver and lung. cells were stained for characteristic surface and intracellular markers and characterized by multiparametric flowcytometric analysis. ccr expression kinetics following stimulation were analysed in tconv and treg isolated from murine splenocytes by facs and polyclonally activated by anti-cd /cd -coated beads in the presence of exogenous il- . expression was monitored by daily flow cytometric analysis. ccr overexpression was induced by transduction of expanded treg with ccr mrna via electroporation. expression kinetics were monitored by facs, receptor function was tested in transwell migration assays using ccr ligands ccl- and ccl- . results: systematic analyses showed higher ccr expression on memory treg than on their naive counterpart in all examined organs with bone marrow samples displaying the greatest disparity. memory treg showed higher ccr expression than memory tconv in all analysed organs, except for lymph nodes where both memory populations revealed equal expression levels. stimulation of in vitro expanded treg and tconv lead to a strong increase in ccr expression with maximum levels on d and d respectively, whereas restimulation (d ) resulted in no further relevant ccr expression on treg. we performed systematic optimization of stimulation and mrna-electroporation conditions to reliably achieve highlevel short-term ccr expression. transduction of treg on d of in vitro expansion resulted in a strong ccr expression, with maximum levels h after electroporation and strong ccr expression being detectable for at least h. transwell migration assays showed enhanced migrational properties of mrna-electroporated treg towards ccr ligands. analyses performed h and h after electroporation showed persistent migration even though measured ccr surface expression had already declined significantly. conclusions: we showed that high ccr expression can be detected on memory treg in all analysed organs. since in vitro stimulation of murine treg did not reliably induce ccr expression, we established a protocol for ccr mrna-electroporation. electroporated cells showed stable short-term ccr expression and enhanced migrational properties towards ccr ligands in vitro. future studies will show whether the induction of short-term ccr expression will facilitate in vivo homing of adoptively transferred treg to sites of ongoing gvhd and thus mediate long-term inflammation suppression. disclosure: the authors have no conflict to disclose. survival and immune reconstitution of syngeneic, haploidentical and allogeneic hematopoietic stem cell transplantation in atm-deficient mice ruth pia duecker , patrick c. baer , stefan zielen , ralf schubert from allo-hsct. it´s not necessary to do chemotherapy before transplantation for patients with bone marrow blast cells more than % at the time of diagnosis. [[p image] . figure the hci-ct of patients before transplantation and occurance of grade iii-iv agvhd on overall surviv] background: . allogeneic haematopoietic stem cell transplantation (sct) offers the chance of cure for patients with transfusion-dependent thalassemia (tdt). based on the non-neoplastic nature of this condition sct approaches urgently require to prove both efficacious and safe. methods: . we report on children, adolescents and young adults (median age: years; range - years) with tdt receiving sct from an hla-matched donor (mud n= , msd n= , mfd n= ) in our center from - . all patients received the same treosulfan-based conditioning regimen (treosulfan x g/m , fludarabine x mg/m , thiotepa x mg/kg). gvhd prophylaxis was based on atg-fresenius™ ( x mg/kg, if mud or mfd as donor), csa (with taper from day + ) as well as mtx (day , , , ) in / and mmf in / patients with mtx toxicity, respectively. stem cell source was bone marrow in , peripheral blood stem cells in and cord blood in patient. prior to transplantation children received cytoreductive treatment with azathioprine, hydroxycarbamide and intensified erythrocyte transfusion. iron elimination therapy was carried out in / children with deferasirox. among the patients with available ferris-can™ analysis patients showed substantial liver iron overload (liver iron > mg/g) despite intensive chelation prior to sct. results: . all patients achieved leukocyte engraftment at median day + (range - ), however two patients required a cd -selected pbsc boost on day + and day + based on delayed platelet and/or erythrocyte engraftment. nine patients exhibited full donor chimerism in the bm at day + , the other showed mixed chimerism with < % autologous cells. on day + peripheral blood chimerism was complete in / patients with the remaining patient exhibiting stable split chimerism with % donor-derived erythrocytes and - % autologous myeloid cells. acute gvhd was observed in three patients (grade : n= , grade : n= ). however, all patients responded to immunosuppressive therapy with steroids ± ecp and re-initiation of cni (n= ). one patient suffered background: patients with relapsed or refractory acute myeloid leukaemia (aml) have a poor prognosis. allogeneic hematopoietic stem cell transplantation is the only curative option. however, allogeneic transplantation with active leukemia failed to improve significantly the longterm outcome. to improve the outcome of allo-hsct in such high-risk and refractory patients, sequential schedule of cytoreduction therapy followed by nonmyeloablative conditioning has been developed. methods: to evaluate the outcome of sequential intensified conditioning regimen followed by allogeneic hematopoietic stem cell transplantation (allo-hsct) for refractory acute myeloid leukemia (aml). results: a total of patients with primary or secondary refractory aml transplanted between june to july were included. refractoriness was defined as primary induction failure, relapse within months from induction/ consolidation chemotherapy or second relapse. median age is years ( to ). the salvage chemotherapy administered was flag-ida. two patients did not receive intensive chemotherapy because of no recovery after induction chemotherapy. seven days after the end of flag-ida, a reduced intensity conditioning consisting of fludarabine, mg/m , thiotepa, mg/kg and busulfan , mg/kg i.v. (n= ) for haploidentical donors or fludarabine plus busulfan (n= ) for hla identical sibling or unrelated donors was administered. graft-versus-host disease (gvhd) prophylaxis consisted of tacrolimus and mycophenolate mofetil. the mycophenolate was withdrawn at day + post-transplantation and tacrolimus at day + . donor lymphocytes (dli) were infused in patients without agvhd at day + post-transplantation. seventeen patients achieved complete donor chimerism, patients progressed early and patient died before engraftment. one of the patients which recovery was with persistent leukemia reached donor chimerism after immunosupression discontinuation. ten patients are alive in complete remission. median follow-up of survivors is months (range: - ). five patients died of leukemic progression, as result of gvhd and suffered intracranial hemorrhage. five patients received prophylactic dli. the incidence of acute moderate-severe gvhd and moderate-severe chronic gvhd were % (n= ) and % (n= ), respectively. the non-relapse mortality was % (n= ), mainly due to acute gvhd (n= ) . the -year cumulative incidence of relapse posttransplantation was . %. the probability of relapse was %± %. the -year os and dfs were % ± % and % ± % conclusions: the strategy of sequential chemotherapy followed by allohsct ± prophylactic dli has an acceptable toxicity profile and improves both the relapse rate and the survival for refractory aml patients. disclosure: nothing to declare background: the concept of immunological intervention to prevent relapse after hematopoietic stem cell transplantation is associated with the assessment of the chimerism status. distinguishing patient and donor hematopoiesis is usually performed by str-pcr, a powerful method developed for forensic purposes. however, this method shows restrictions with respect to detection limit, preciseness, and the possibility of automated read out. digital pcr could circumvent some of these limitations. methods: recently, validated for the bio-rad droplet digital platforms, the biotype mentype digitalquant assay was released. the assay uses indel polymorphisms on chromosomal dna to distinguish patient and donor hematopoiesis on a fret hydrolysis assay basis ("taqman assays"). thus the assay in principle is applicable on the chamber based d digital pcr system (thermo fisher). due to different reaction chemistry and physical properties of thermal transfer between the digital pcr platforms protocols are reasonably not fully compatible which would lead to lower fluorescence intensities and poor signal resolution on the solid chip based thermo fisher d platform. an adjusted pcr protocol was established and optimized using representative markers, followed by determination of tolerable and optimal amount of input dna. specificity, sensitivity and reproducibility testing with artificial mixed samples preceded the extensive verification by comparative measurement of clinical samples (n= ) and ring-trial samples (n= ). source to allogenic bm or pbsc. ucb units are immediately available for transplantation as they are frozen and banked with defined hla typing and it has an advantage for patients who need urgent transplantation. in addition, a higher degree of hla mismatch appears to be acceptable with a comparatively lower risk of acute and chronic gvhd. meanwhile, a higher incidence of engraftment failure, delayed neutrophil and platelet recovery, and posttransplant immune disorders including pre-engraftment immune reactions (pir) are major problems in unrelated ucbt. methods: in our institute, gvhd prophylaxis in ucbt was changed after march . between january and march , thirty-two patients received tacrolimus plus methylprednisolone (tac/mpsl) and between april and january , thirty-one patients received tac plus methotrexate (tac/mtx) for gvhd prophylaxis. to investigate better gvhd prophylaxis after ucbt, we compared transplant outcomes after ucbt using gvhd prophylaxis with tac/mpsl (n= ) and tac/mtx (n= ) in single-pediatric transplantation center. results: the cumulative incidence of neutrophil engraftment at day in tac/mpsl group was . % and . % in tac/mtx group (p= . ). median time of neutrophil engraftment was days earlier in tac/mtx group ( days) than tac/mpsl group ( days). according to pir, and acute gvhd, tac/mtx group showed superior outcomes; the incidence of pir (p= . ) and the cumulative incidences of acute gvhd at day ( . vs . %, p = . for grade ii-iv, . vs . %, p= . for grade iii-iv) was significantly lower in tac/mtx group than in tac/mpsl. however, the incidences of relapse (p= . ) and cytomegalovirus viremia (p= . ), and estimated overall survival (p= . ) and event-free survival (p= . ) were comparable between two groups. conclusions: our results indicated that gvhd prophylaxis with tac/mtx had favorable effects; reduced incidence of rip and acute gvhd after ucbt without any negative influences. disclosure: nothing to declare transfer of donor regulatory t-cells after atg reconditioning cures severe refractory gvhd and leads to long term persistence of regulatory t-cells in the recipient cells on a clinimacs® plus device (miltenyi biotec). the cell product contained % foxp + t-cells. the patient received , x /kg t reg on day + . subsequently intestinal gvhd decreased and finally resolved. three months after the first t reg transfer the patient got a second t reg transfer ( , x /kg) on day + due to decreasing t reg levels. thereafter t reg persisted and there was no recurrence of gvhd. the patient is well with low dose sirolimus and prednisone as the only immunosuppressants and is particularly recovering intestinal function. conclusions: this case illustrates an unusually severe acute gvhd after matched sibling sct. transfer of donorderived t reg was able to cure severe and refractory gvhd after t-cell ablation by atg. transferred t reg persisted in the recipient for a long period and did not lead to any adverse events. disclosure: no disclosures to declaim allogeneic hsct for patients with transfusion dependent anemia from matched and mismatched donors julia fekadu , andrea jarisch , jan sörensen , emilia salzmann , eva rettinger , andré willasch , shahrzad bakhtiar , thomas klingebiel , peter bader after first asct. the mean harvest for patients receiving dhap was , x cd (+) cells/kg, , x cd (+) cells/kg for cy, . x cd (+) cells/kg for igev, , x cd (+) cells/kg for ice, , x cd (+) cells/kg for choep. the patient mobilized with vtd-pace achieved , x cd (+) cells/kg after apheresis. of the patients achieved the target number of > x /kg cd + cells after apheresis, after two, and after three apheresis. the median time to apheresis was days ( - ) without significant difference between the regimens. the mean wbc count at the time of apheresis was , x /l after dhap, , x /l after cy, , x /l after igev, , x /l after ice, , x /l after choep. there was correlation between wbc and cd harvested cells (p= . ). grate - thrombocytopenia was found in patient ( dhap, ice, igev, vtd-pace). grate - anemia was registered in patients ( dhap and vtd-pace). no correlation was found between the cd + harvest and the age, number of previous lines chemotherapy, the response before mobilization, the type of the lymphoma and the clinical stage. conclusions: our results demonstrate that the chemo-g-csf protocols have comparable effectiveness with accep- background: cytomegalovirus (cmv) may cause severe complications in recipients of allogeneic haematologic stem cell transplantation (allohsct). letermovir (ltv, / mg daily without/with co-administration of cyclosporine) was recently licenced only for cmv prophylaxis in adult allohsct-recipients. paediatric data as well as data on cmv therapy are missing so far. methods: we administered letermovir mg orally once daily (with no co-administration of cyclosporine a) to paediatric patients after allohsct. edta-plasma were occasionally obtained at different time points and frozen for determination of letermovir levels using liquid chromatography/mass spectrometry (lc-ms/ms). results: for details on patients, treatment and cmv load see table . in short periods of letermovir administration, cmv blood levels became negative in both patients. considering the lacking safety data in paediatric patients, we stopped letermovir treatment in both patients, when liver parameters increased. in patient hepatopathy turned out to represent histologically proven graft versus host disease (gvhd). in patient liver parameters further increased despite withdrawal for another weeks, however, hepatopathy was only mild and self-limiting. both patients additionally received other possibly hepatotoxic substances (mycophenolate mofetil and trimethoprim/ sulfamethoxazole). letermovir plasma levels were . ng/ml ( h), . ng/ml ( h), . - . ng/ml (median . ng/ ml, n= , h) and . ng/ml ( h after administration). conclusions: during short letermovir treatment, we observed fast resolution of cmv viraemia as well as rising liver parameters in both patients. while elevated liver parameters represented gvhd in patient, a causal relationship with letermovir might be considered in the other patient. letermovir peak levels after administration of mg were within ranges reported in adults after administration of mg while trough levels were higher indicating differences in pharmacokinetics in terms of delayed clearance. inguinal lymphadenomegaly. after failure to respond to seven conventional treatment lines: methotrexate, cop (cyclophosphamide, vincristin and prednisone); gemcitabine; puva; interferon; acitretin and extracorporeal photopheresis), allogeneic hsct from an identical hla male donor was indicated. the non-myeloablative conditioning consisted of fludarabine ( mg / m ), cyclophosphamide ( mg / kg) and total body irradiation(tbi) ( cgy). prophylaxis of graft versus host disease (gvhd) was performed with cyclosporine ( mg / kg) and mycophenolate mofetil ( mg/kg). after conditioning, there was improvement of pruritus and involution of the skin. bone marrow infusion occurred on / / (d ). on d + he presented recurrence of skin lesions of fmf. donor lymphocyte infusion (dli) was performed ( x cd + cells / kg / recipient). he presented oral lichen and diarrhea respectively as manifestations of gvhd on d + and d + . as infectious intercurrence, hemorrhagic cystitis occurred by bk virus months after the first dli and he received conservative treatment and remained without systemic immunosuppression. nine months after hsct, a second dli ( x cd + cells / kg / receptor) was performed and at this time the patient is without clinical manifestations of fmf or gvhd. conclusions: the clinical response of the presented case confirms what has been reported in the literature. ctcls appear to be particularly susceptible to gvl effect, which makes hsct a potential cure for advanced ctcls in eligible patients. the timing to perform hsct in the clinical course of the disease remains a matter to be settled clinical trial registry: not applicable disclosure: no conflict of interest p abstract already published. methods: we applied next generation sequencing (pgm, ion torrent/ fischer lifetechnologies) to an unselected cohort of patients ( female, male, median age ( - ) years) who had been referred for allogeneic stem cell transplantation due to the presence of a high-risk myeloid disorder dnmt a r h ( . %) . ), cebpa ( . %; ceb-pap s ( . %) vus) kras ( . %; krasr r ( . %) vus), and kit ( . %; kitm l ( . %) . ). patients displayed a median of sequence variants (aml, mpn and cmml patients each ; mds, saa and patients with other, non-malignant hematologic diseases each sequence variants found most frequently in aml were cebpap s ( . % of all sequence variants in patients with aml, p< - , chi² test) in patients suffering cmml, dnmt ar h was particularly frequent ( . % of all sequence variants in cmml, p=. ). asxl e d ( . % of all sequence variants in other, non-malignant hematologic diseases, p=. ), idh r q ( . %, p=. ) and krasr r ( . %, p=. ) were frequent in other, non-malignant hematologic diseases. in saa, nrasg d ( % of all sequence variants in saa patients; p=. ) was frequently found, as were dnmt ak fs* ( %, p=. ), tet l w ( %, p=. ) and tet i v ( %, p=. ). conclusions: taken together, these data show that vus occur with high abundancy in this high-risk cohort of patients, and that they differ in frequency between various myeloid disorders methods: retrospective analysis of patientswho experienced either hematological relapse or progressed to aml after allo-hsct and were treated with azacitidine for this indication at hematological centers in poland. the primary end-point was overall survival (os), the secondary -response rate. results: patients, males ( . %), median age (range, - ), were enrolled. the primary indication for allo-hsct was aml median time from allo-( %ci: . - ); with -year os of . % ( %ci: . - . ). for patients stratified according to ebmt aza relapse prognostic score -year os was: prophylaxis of agvhd: with atg -atgam mg/b.w.- pts( , %), posttransplant cyclophosphomide (ptcy) mg/b.w. on d+ ;d+ - pts ( , %) conclusions: unmanipulated haplo-hsct in st - nd cr in children and adolescents with high risk al allows achieving the long-term survival in , %. the use of g-csf stimulated unmanipulated haplo-bm is associated with a satisfactory rate of engraftment. the main cause of death in our study was relapse after allo-hsct. the frequency of acute and chronic gvhd was acceptable, -years grfs rate of , % in st - nd cr represents good quality of life following unmanipulated haplo-hsct and therefore may be recommended as option for use in children and adolescents with high-risk al. disclosure: nothing to declare background: b -h (cd ) is thought to act as an immune checkpoint and regulates t and nk cell responses. it is highly overexpressed on many solid tumors while on healthy tissue protein expression is limited. this makes b -h an interesting target for cancer immunotherapy. in highrisk neuroblastoma patients, targeting disialoganglioside gd with the recently approved monoclonal antibody (mab) ch . after autologous or allogeneic sct, significantly improves survival. however, gd expression is heterogeneous and ch . causes severe adverse effects. thus, we evaluated b -h as an alternative or additional target antigen. we investigated different anti-b -h mab constructs and mab-cytokine fusions (immunocytokines) for their ability to elicit antibody-dependent cellular cytotoxicity (adcc) using expanded γ/δ t cells of healthy donors, chex lf-il was confirmed to be the most effective mab construct. interestingly, chek -il showed comparable target cell lysis -however, lysis was only transient while chex lf-il mediated permanent target cell lysis. using patient pbmcs after receiving allogeneic sct, chex lf-il and ch . mediated comparable lysis. calculated specific lysis of lan- (after hrs.; in ascending order): targets + effectors w/o mab ( %) conclusions: b -h is a suitable target antigen in case gd expression is low or absent. immunocytokines and fcoptimized mabs targeting b -h might increase the efficacy of immunotherapy in gd -negative tumors and in combinatory approaches. until now, the low-fucose immunocytokine chex lf-il seems to be the most promising anti-b great ormond street hospital, nhs foundation trust petersburg/ raisa gorbacheva memorial institute of children's oncology, hematology and transplantation, rehabilitation medicine, saint petersburg, russian federation, first i. pavlov state medical university of st. petersburg/raisa gorbacheva memorial institute of children's oncology, hematology and transplantation, bone marrow transplantation for pediatric solid tumor petersburg/raisa gorbacheva memorial institute of children's oncology, hematology and transplantation, pediatric hsct outpatients, saint petersburg, russian federation university hospital carl gustav carus james`s hospital diagnosis was aml in %, all in % and mds in % of patients. % of patients received pbsc grafts, % received unmanipulated bone marrow grafts. os at years was % in msd/mud-atg, % in haplo-ptcy, % in mmud-ptcy and % in mmud-atg groups (p= . ). in a multivariate cox model non-relapse mortality was %, %, . % and % in msd/mud-atg, haplo-ptcy, mmud-ptcy and mmud-atg groups, respectively. cumulative incidence of acute gvhd grade or was %, . %, % and % after in msd/mud-atg, haplo-ptcy cumulative incidence of chronic gvhd was % in the msd/mud-atg group uwe platzbecker , verena wais republic of china background: there are two most noteworthy strategies of haploidentical stem cell transplantations (haplo-hsct), the baltimore post-transplantation cyclophosphamide (ptcy) with or without anti-thymoglobulin (atg), and the beijing g-csf primed bone marrow (bm) plus peripheral blood stem cells (pbsc) (giac). however, the comparison of these two modalities is scarce. in this study, we aim to compare these two approaches for hematological malignancies based on the taiwan blood and marrow transplantation registry (tbmtr) with the comparable cd infusion amounts, the neutrophil engraftment time were statistically distinct among these three groups [d+ (group ) vs. d+ (group ) vs. d+ (group ), respectively as to the graft-versus-host disease (gvhd), the patients in group had more grade ii-iv but similar grade iii-iv acute gvhd compared with others (grade ii-iv: . % vs. . % vs. . %, respectively conclusions: haplo-hsct with different strategies is a feasible treatment modality for hematologic malignancies in taiwan. regarding the retrospective nature and limited patient numbers of this study republic of china background: we previously presented a low-resolution hla analysis of cedace, the voluntary portuguese bone marrow donor registry (ebmt , poster p ) and, more recently, its epidemiological characterization (ebmt , poster b ). currently, cedace is one of the largest bone marrow donor registries in the world, including nearly % of the country's population, twice the number of donors present in . the current work is an update on the most common hla haplotypes found in cedace results: of the donors in the cedace registry, . % were typed in at least loci (hla-a/-b/-drb ), . % in (including hla-cw), and . % in (including hla-dqb )the , and most common haplotypes accounted for, respectively, . %, . % and % of the haplotypes found in the entire registry. the five most common haplotypes at the low-resolution at the loci, low-resolution level, out of donors, individual genotypes were identified, leading to an hla matching probability at this level of . % hid-hsct) have a stronger anti-leukemia effect compared to identical sibling donor hsct(isd-hsct) in high-risk features .but in refractory/relapsed(r/r) aml patients who not in remission status, it is unclear whether it also augments the gvl effect antithymocyte globulin was used in haploidentical hsct. unmanipulated bone marrow and peripheral blood stem cells for all patients. cyclosporine, short-term methotrexate were employed for gvhd prophylaxis. mycophenolate mofetil included in hid-hsct. performed multivariate analysis for all patients of pretransplantation variables and developed a predictive scoring system for survival according to adverse factors. results: the total survivor median period of follow up was ( - ) months. hid -cohort had higher -year actuarial of os multivariate analysis showed isd-hsct,standard conditioning regimen and less than % proportional reduction in blast percentage pre-≥ . conclusions: haploidentical donor compared to identical sibling donor had better anti-leukemia effect in allo-hsct for r/r aml in nr status conditioning protocol was melphalan mg/m for mm and beam for nhl. the quantification and characterization of γδt cells in peripheral blood samples were performed by flow cytometry based on the expression of cd /cd /vδ /vδ /vγ /cd at , and days after ahsct. percentage (%) of γδt cells represented the proportion of these cells among all t cells. results: median age at ahsct was ( - ) years, % male. median time from diagnosis to mobilization was ( - ) months, after a median number of therapeutic lines of ( - ); pts ( . %) received radiotherapy. seventeen pts ( . %) were re-mobilized with plerixafor±g-csf ( % mm vs % nhl there was no difference in febrile neutropenia incidence (p= . ), timeto-engraftment (p= . ), time-to-neutrophils> /μl (p= . ) or erythrocyte transfusions (p= . ). however, there was more time-to-platelets> , /μl ( vs days; subpopulations did not affected pfs. conclusions: our results showed that pts mobilized with plerixafor need more collection volume (less cd +cells, higher dmso). plerixafor negatively affected platelet recovery, with similar hematologic and immune recovery for the remaining variables. its use was associated with higher %vδ +, suggesting that it induces an antineoplastic phenotype. more studies with larger samples and follow-up period are needed to evaluate plerixafor results: total ascc procedures were carried out in patients over years, once in patients, twice in patients, three times in patients, and four times in patient. non-hodgkin lymphoma (nhl) comprised . % of all cases (n = ) ), total number of chemotherapy cycles before ascc (or . , % ci . - . , p = . ), failure to achieve at least partial response before ascc (or . , % ci . - . , p = . ), total number of days receiving g-csf for mobilization (or . , % ci . - . , p = . ), and salvage use of plerixafor (or . , % ci . - . , p < . ) were found to be independent factors associated with failure of ascc. at the end of the study period, . % of successful collections (n = / ) were used for asct, . % (n= / ) were in storage awaiting transplantation hôpital necker-enfants malades melf methods: a total ahsct candidates [median age: ( - ) years; male/female: / ] were included in this study. twenty-seven patients ( . %) were diagnosed as non hodgkin's lymphoma, patients ( . %) hodgkin's lymphoma, patients ( . %) multiple myeloma, patients ( . %) acute myeloid leukemia, patients ( . %) plasmocytoma and patient ( %) testis cancer. premobilization serum -hydroxy vitamin d ( -oh d) levels were measured with immunoassay method peripheral cd + cell count background: some published data suggest a positive effect of iron chelators on the risk of post-transplant relapse, with an improvement in overall survival after allogeneic hematopoietic stem cell transplantation (hsct) conditioning regimen consisted to intravenous bu mg/m and, flu mg/m d- to d- . gvhd prophylaxis included atg . mg/kg on d- and d- , ciclosporin and methotrexate. all patients received peripheral blood stem cell transplant from an identical hla-related donor. iron chelation consisted on deferasirox (exjade ® ) - mg/kg/day, started at day , if serum ferritin level ≥ - ug/l and stopped when the level decreased below ug/l or normalized at day after transplant, / patients were evaluable (g = pts), (g = pts). patients were abo compatible ( %), had major incompatibility ( %), and % had minor incompatibility. median serum ferritin level at day , were ug/l ( - ) and ug/l ( - ) in g and g respectively with a median follow-up of months, ( - ), disease relapse incidence was higher in patients who did not received iron chelation treatment (g : . %) versus those who received oral deferasirox (g : . %) (figure ), but the difference was not statistically significant conclusions: these results deserve more investigation choep (n= ), and patient received vtd-pace μg/kg depending on the protocol. the aim was to collect at least x cd (+) cells/kg body weight. results: forty patients all patients were stage iii and iv at diagnosis. of the patients were mobilized after one line of treatment, six after two lines of treatment and five after and more lines of treatment alexandra martínez-roca , gerardo rodriguez-lobato , gonzalo gutierrez-garcia , , maria suárez-lledó background: hematopoietic stem cell transplantation (hsct) is an established procedure in lymphoma background: the role of inflammatory cascade in tumor microenvironment has been demonstrated in several studies. as a part of this issue, elevated neutrophil/lymphocyte ratio (nlr) was shown to be associated with an adverse prognosis, particularly in solid tumors. the aim of this study is to determine the impact of pre-transplant nlr on early transplant complications, as well as post-transplant relapse and survival.methods: a total of lymphoma patients [median age: ( - ) years; male/female: / ] who underwent autologous hematopoietic stem cell transplantation (hsct) were included in this retrospective study.results: the initial diagnosis was hodgkin lymphoma (hl) in ( . %), b-cell non-hodgkin lymphoma (nhl) in ( . %) and t-cell nhl in ( . %) patients. of patients who were evaluated for pretransplant disease status, patients ( . %) were in complete remission, patients ( . %) were in partial remission and ( . %) patients had refractory disease. median pre-transplant nlr was found to be . ( - ) . when the study population was divided into two subgroups as "low-" and "high-nlr", based on median nlr value, number of febrile days were found to be relatively higher in the low-nlr group (p= . ). a positive correlation was demonstrated between nlr and lactate dehydrogenase levels (r= . ; p= . ); and nlr and ferritin levels (r= . ; p= . ). at a median follow-up of ( - ) months, overall survival (os) was found to be better in the low-nlr group without statistical significance [ vs ( - ) months; p= . ]. in univariate analysis, pre-transplant nlr represented a significant impact on os (p= . ). other prognostic factors were age (p= . ), platelet engraftment (p= . ), post-transplant relapse (p= . ) and pre-transplant ferritin level (p< . ). the permanent impact of ferritin on os was confirmed in multivariate analysis (p= . ).conclusions: in this study, an adverse impact of elevated pre-transplant nlr on os was demonstrated in autologous hsct recipients with lymphoma. as a predictor of prognosis, nlr may be considered as a safe and cost-effective parameter. further studies are required in order to use this predictor in routine clinical practice.background: high cure rates in childhood diseases have been achieved by stem cell transplantation (sct). however, there is little knowledge concerning recovery of the immune system and community-acquired infection after sct. here we studied the long-term reconstitution of the immune system and incidences of community-acquired infection after sct.methods: we reviewed medical records for patients (m/f: / , median age: years (range: - years) who were treated in the department of pediatrics, hokkaido university hospital. we analyzed cd -positive cell counts, serum immunoglobulin g (igg) levels, and incidences of community-acquired infection until years after sct. indications for sct were all in patients, aml in , aa in , nb in , rms in , jmml in , nhl in , cgd in , was in , xscid in , apds in , cd ld in , and other solid tumor in patients. stem cell sources were autologous pb/bm in , allogenic bm in ( related and unrelated) and allogenic cb in patients. in this study, we excluded patients who relapsed after sct.results: the duration of cd -positive cell counts < / ml after sct was . ± . months in all patients. the durations were . ± . months in patients with hematologic malignancies, . ± . months in patients with hematologic disorders such as aplastic anemia and pid, . ± . months in patients with solid tumor, . ± . months in patients who received autologous sct, . ± . months in patients who received related bmt/ pbsct, . ± . months in patients who received unrelated bmt, and . ± . months in patients who received cbt. the durations of igg < mg/dl after sct were . ± . months in all patients, . ± . months in patients with hematologic malignancies, . ± . months in patients with hematologic disorders, . ± . months in patients with solid tumor, . ± . months in patients who received autologous sct, . ± . months in patients who received related bmt/pbsct, . ± . months in patients who received unrelated bmt, and . ± . months in patients who received cbt. there was a significantly higher incidence of community-acquired infection from months after sct. there were significant differences in the incidence of community-acquired infections between patients with cd -positive cell counts of < /ml and background: allogeneic hematopoietic cell transplantation (allo-hct) has the potential to cure subgroups of patients with multiple myeloma (mm) but its role is controversial due to high transplant-related mortality. while autologous hct is well established as consolidation after induction therapy using novel agents, allo-hct is still considered experimental due to excessive early toxicity.methods: we retrospectively studied mm patients (pts) ; ( , %) were males with a median age of years (range: - ), who underwent allo-hsct in our center between and .median time between diagnosis and allo-hct was months (range: - ).results: the international staging system (iss) was iss i ( , %), iss ii ( , %), and iss iii ( , %). twentyeight ( , %) pts received cells from unrelated donor and pts from identical siblings. stem cell source were: peripheral blood (n= ) and bone marrow (n= ). grouprisk distribution using hct-ci and pam index can be seen in figure . response was evaluated at day + : cr ( , %), vgpr ( , %), pr ( , %) and sd ( , %); response was not evaluated in pts. regarding hospitalization, ( , %) pts and ( , %) pts needed readmission in the first days and days post-allohct, respectively, and median days of hospitalization was days (range: - ). reasons for first re-admission were: infection ( , %), gvhd ( , %) and renal insufficiency ( , %). twenty-seven ( %) died and death causes were: infection ( , %), progression ( , %), secondary tumor ( , %) and the remainder, gvhd and intracranial hemorrhage. in addition, pts ( , %) suffered cytomegalovirus (cmv) reactivation.median overall survival (os) was months (range: - ). univariate analysis showed that re-admission in first days (hr , ; p= , ) , re-admission in first days (hr , ; p= , ) , cmv reactivation (hr , ; p= , ), iss (iss-i vs iss-ii and iii; hr , ; p= , ), days of hospitalization in the first days and response at + day (cr plus vgpr vs the remainder; hr , ; p= , ) were predictor factors for os. other factors like karyotype, response before allo-hct, hla-mismatch or baseline cmv serostatus, among others, do not impact on os.median os in pts with low hct-ci were months (range: - ) vs months in intermediate-high hct-ci background: allo-sct is a potentially curative therapy for patients with multiple myeloma as it provides a graftversus-myeloma effect and a myeloma-free graft. due to increased nrm and unclear os benefit the recent guidelines suggested allo-sct to be used in context of clinical trials focusing on the high-risk patients and those who relapsed early after autograft. reduced-intensity conditioning regimens may improve rate of nrm; however, optimal conditioning regimen is still to be determined. here we studied conditioning regimen with alkylating agents consisting of thiotepa (tt), busulfan (bu) and gvhd prophylaxis with post-transplant cyclophosphamide (post-cy) in high risk myeloma patients relapsing after autograft.methods: a total of patients (m, n= ) with median age of years (range - ) underwent an allo-sct (mud, n= ; mrd, n= ; mmud, n= , haploidentical, n= ) during a period from to in university of hamburg. the majority of patients had advanced disease (stage iiiab, %) and high-risk cytogenetics ( %). the median response durartion after autograft was . years (range . - . ). the conditioning included tt (cum. dosage mg/kg), bu (median cum. dosage . mg/kg i.v. or . mg/kg in elderly patients) and post-cy (cum. dosage mg/kg, day + and + ). eight patients (all of them received allografts from mmud or haploidentical donors) received additionally fludarabine (flu, cum. dosage of background: the overall incidence of active cmv infection in patients with multiple myeloma (mm) receiving background: several scoring systems have been developed to estimate outcomes in mds patients who undergo allohct. however, none of them have been specifically validated in t cell depleted grafts. the aim of this study is to investigate the prognostic ability of a recently published scoring system (sfgm-tc) in a cohort of patients with mds who underwent tcd transplants.methods: patients underwent a first tcd allohct for mds from to . the sfgm-tc score (caulier et al. curr res transl med. ) is performed at day and ranges from - , discriminating low ( ), intermediate ( ) ( ) ( ) , and high risk ( - ). additional analyses were performed at day and day . a landmark analysis was done at each time point for the day , , and analyses, respectively. background: hematopoietic stem cell transplantation (hsct) is the only curative procedure for the treatment of myelodysplastic syndrome (mds), but among several limiting factors for its accomplishment, such as the patient´s performance status, is a very relevant issue, i.e., the availability of a compatible hla donor and, when available, very often the donor´s age and comorbidities also constitute factors that hinder this medical conduct. considering this scenario, the possibility of a haploidentical transplantation (ht) has emerged as an option. in latin america, ht has been included as a treatment option since . since then, these patients have been included in the latin american registry of transplantation in myelodysplastic syndrome, making it possible to analyze the viability and results of these transplants.methods: from october to october , seventeen ( ) patients were transplanted with a haploidentical donor and included in the latin american registry. none of these patients had an identical hla ( / match) related or unrelated donor. data were obtained from the latin american registry of hsct in mds. the statistical analyses were performed using the software spss, version . and graphpad prism version . , with significance being set at p < . .results: table shows the patients and their characteristics. all donors were haploidentical. there was a predominance of reduced intensity conditioning, which was performed in patients ( . %), whereas the others received the myeloablative conditioning. cell source was peripheral blood in ( . %) and bone marrow in ( . %) of the patients. graft-versus-host disease (gvhd) prophylaxis in post-hsct was carried out with cyclophosphamide mg / kg on d+ and d+ , cyclosporin from d and mycophenolate from d+ . complete hematologic recovery was achieved in ( . %) patients. the incidence of grade ii-iv acute gvhd was . %, whereas chronic gvhd was . %. one death occurred due to graft failure and none of the patients showed autologous recovery. three other patients died. one on d+ due to a fungal infection, the second on d+ due to sinusoidal obstruction syndrome and a third on d+ due to pneumonia caused by pseudomonas. regarding overall complications, there was a predominance of mucositis ( %), overall infections ( . %) and reactivated cmv in . % of cases. of the total number of living patients, ( %) achieved complete remission and ( . %) showed disease relapse. the mean follow-up was months (ranging from to months). the lowest probability of disease-free survival at years was % ( % ci: . - . ).background: relapse is the most important cause of failure after allogeneic hematopoietic stem cell transplantation (hsct) for flt -itd-positive acute myeloid leukemia (aml). treatment with flt tyrosine kinase inhibitors (tki) constitutes a promising clinical approach to induce remission without conventional chemotherapy.methods: a year-old woman diagnosed with aml secondary to myelodisplastic syndrome (mds) with npm mutation and internal tandem duplications of the flt gene (flt -itd) in october . after achieving complete remission (cr) with conventional chemotherapy, she received a hla sibling allogenic-hsct in february , with bucy. four months later, aml relapsed only at medullary level (flt ratio: , %), treated with chemotherapy and donor lymphocytes infusions (dli). she achieved nd cr and developed limited chronic graft-versushost disease (cgvhd). nine months later (april ), she suffered the first extramedullary relapse only, breast and skin. disappearance of the lesions at all levels was achieved with chemo and radiotherapy. she always had full hematologic donor chimerism.in december , she referred atypical precordial pain irradiated to the back. cardiac mri was performed and several masses were visualized in the pericardial sac, up to cm in diameter. bm remained in cr with full donor chimerism.pericardial fluid showed massive infiltration by leukemic-flt positive cells (ratio: , %). she was not considered background: ta-tma is a severe complication that can reduce survival after hsct. risk factors have been variably reported in adults although data on children remains scarce. we aimed to identify a risk profile for development of ta-tma in children undergoing hsct.methods: we retrospectively reviewed clinical charts of children who underwent hscts between - : at great ormond street hospital (gosh) and the great north children's hospital (gnch). ta-tma was defined according to revised criteria (jodele et al. ) . risk factors were categorized into patient derived [age, gender, active co-morbidity at d of hsct (uncontrolled viral/ bacterial or fungal infection, pulmonary, cardiovascular instability, steroid therapy > . mg/kg beyond d , bcgiosis or autoimmune disease), transplant related factors (conditioning intensity, stem cell source, hla-matching, use of ciclosporin a (csa) or tacrolimus (tac), cd + dose, ex-vivo t cell depletion, use of defibrotide) and post-hsct factors (agvhd, post-hsct viral reactivation, venoocclusive disease (vod) and occurrence of posterior reversible leukoencephalopathy (pres).results: at a median of months post-hsct, ta-tma occurred among / transplants ( . %). there was no reported centre variation ( . % vs . % in gosh vs gnch; p= . ). gender, underlying disease -primary immune deficiency (pid) (n= ) vs haematological disease (n= ), use of myeloablative (n= ) vs reducedor minimal intensity conditioning (n= ), use of serotherapy or mega doses of cd ≥ x * /kg did not influence the development of ta-tma. donor type: msd/ mfd(n= ) vs mud (n= ) vs mmud/haplo-hsct background: we evaluated the outcome of haploidentical hct (hhct) using ex vivo αβ t cell-depleted (tcd) grafts after reduced-intensity conditioning (ric) containing low-dose tbi (ld-tbi) in pediatric patients with acute leukemia (al) in complete remission (cr).methods: between may and october , patients with acute leukemia ( all and aml) in cr received haploidentical hematopoietic cell transplantation (hhct) using tcrαβ-depleted graft at asan medical center children's hospital. eighteen patients received hhct between and (earlier time period) and the remaining between and (recent study period). the conditioning regimens, the dose of αβ+ t cells and gvhd prophylaxis are summarized in table. results: all patients achieved a sustained neutrophil engraftment at a median of days (range, - ) . of patients, patients ( all & aml) relapsed at a median of months (range, - ) after transplant. of the patients, patients died of disease. one patient died of disseminated tuberculosis at months after transplant, leading to the trm of % at year. as of december , of the patients survive free of disease at a median follow-up of months (range, . at a median followup of months (range, - ), efs and os at years for all patients were % and %, respectively. outcome of hhct in the recent study cohort was significantly better than that in the earlier study period (efs of % vs efs of %, p= . ). among the patients with all, the efs of patients, who received hhct in early time period after conditioning with tbi of cgy, was significantly worse than that of patients, who received in recent study period after a higher dose of tbi at cgy ( % vs %, p< . ). the efss of aml were similar between the two study groups ( % for earlier cohort vs % for recent study, p> . ).conclusions: in pediatric patients with acute leukemia in cr, our current haploidentical hct using ex vivo αβ tcd graft after ric containing ld-tbi without gvhd prophylaxis is feasible approach with a low trm. the background: anti-hla antibodies (ahab) have been recently recognized as an important risk factor for graft failure (gf), especially in hla-haploidentical stem cell transplantation (haplo-hsct). although, recently, ebmt consensus guidelines have been published [ciurea, bone marrow transplant ] , experience in pediatric t-cell depleted (tcd, another well-known risk factor for gf) haplo-hsct is lacking. in the present study, we report our experience on the use of a desensitization approach based on ebmt guidelines.methods: between june and august , all patients affected by non-malignant diseases and scheduled for a transplant from an hla-haploidentical donor after negative depletion of αβ t and b cells as previously described [li pira, biol blood marrow transplant. ], were tested for ahab with luminex® solid-phase immunoassay (one lambda, thermo fisher scientific) month before the hsct. all patients with a mfi higher than , which is considered a cutoff predicting for gf, were treated with a desensitization protocol based on the use of anti-cd rituximab ( mg/m before and immediately after the end of plasma-exchange cycle) and plasma-exchange (pe) ± infusion of irradiated buffy coat (bc) (if after pe ahab mfi was still > mfi). this latter was obtained by the non-target fraction of the αβ t-cell/b-cell-depletion procedure and consisted of * irradiated nucleated cells/kg of the recipient; this was infused - hours before the infusion of the tcd graft. pe was performed with miltenyi life tm apheresis unit (miltenyi, bergish-gladback) .results: in the study period, patients received αβ tand b-cell depleted haplo-hsct. eighteen ( %) resulted positive for ahab (mfi> ); ( %) of them had an mfi > for either anti-class i or ii ahab. these patients (see table i background: osteonecrosis (on) is a debilitating complication in survivors of allogeneic hematopoietic cell transplantation (hct). limited data is available on its course post-transplantation in children. the purpose of our study was to identify recipients of hct in whom pre-and post-magnetic resonance imaging (mri) is indicated.methods: the retrospective cohort consisted of patients who underwent first allogeneic hct from - , and prospectively underwent a total of pre-and post-transplant mri studies of the hips and knees done annually for years regardless of symptoms. surviving patients were followed for a median time of . (range . - . ) years. cases of on were compared to controls matched for age, sex, transplant type, and follow up in a : ratio for the following variables: ethnicity, underlying disease, on pre-hct, conditioning regimen, graft source, bone mineral density z-scores, body mass index, presence or absence of graft-versus-host disease, steroid use and dosage, and survival status.results: thirty ( . %) patients had mri findings confirming on post-hct. all patients with on except one were more than years of age. twenty ( %) patients were male. on pre-hct (p < . ) was the only factor associated with presence of on post-hct. epiphyseal on was seen in ( %) patients pre-hct, and ( %) post-hct. eighteen ( %) patients had involvement of more than % of articular surface, and were more likely to undergo surgery (p = . ).conclusions: the incidence of on in this large pediatric cohort was %. the only risk factor for on post-hct was pre-existing on. mri evaluation for on pre-hct is indicated in all patients. mri evaluation for on post-hct is only indicated for patients with on pre-hct and symptomatic patients. this will entail cost savings of usd , per surviving allogeneic hct patients per year. patients with more than % involvement of the articular surface need close follow up.clinical trial registry: none disclosure: none impact of rabbit anti thymocyte globulin exposure on immune reconstitution and outcome after stem cell transplantation in children background: rabbit anti thymocyte globulin (ratg) has been frequently used for many years as gvhd prophylaxis in pediatric stem cell transplantation. precise dosing regimens are crucial but remain challenging due to several pharmacological characteristics in children.methods: ratg levels were measured in pediatric patients undergoing allogeneic stem cell transplantation after obtaining approval by the local irb and informed consent by legal guardians. pediatric patients who received either thymoglobuline™ (n= ) or grafalon™ (atg-f) (n= ) as part of their conditioning regimen background: obesity among children is a growing health problem. malnutrition or being over-weight can be of prognostic impact among children who need hsct.scientific literature shows a lot of controversy in terms of effect of bmi at the time of infusion on the outcome of hsct.methods: we reviewed data of patients who underwent hsct at king faisal specialist hospital & research centre between - to correlate bmi with the outcome and complications of hsct. transplant naïve recipients with age at infusion between - years who received hsct from matched related donor or autologous hsct, were included in the dataset for analysis. a total of patients' profiles were reviewed of whom . % were boys. median age at transplant was . years. primary indication of disease was malignancy in . % followed by hemoglobinopathies . %, bone marrow failures . % and immune disorders . %. solid tumors accounted for . % among malignant disorders. myeloablative conditioning was used in . % transplants with . % regimens containing total body irradiation. majority of the patients . % underwent allogeneic transplantation using bm as the source in . % and pbsc in the remaining . % cases. donor was hla-identical sibling in %, parents in . % and others in the remaining . % patients. median tnc dose was . x ^ and cd was . x ^ per kg of the body weight at the time of infusion. age and gender specific bmi percentiles were obtained and classified according to the definition of centers for disease control and prevention (cdc); . % ( ) recipients were categorized as under-weight, . % ( ) normal, . % ( ) over-weight and remaining % ( ) as obese.results: based on chimeric studies at day- , our engraftment rate was . % ( ) out of evaluable cases. median time to neutrophil recovery was -days from infusion and -days for platelets. no statistically significant difference was found for engraftment rate on d- as it was % ( ) among . % ( ) in normal, % ( ) in under-weight and . % ( ) in the obese (p-value: . ). median time to neutrophil recovery from the infusion date was -days in over-weight patients and in the remaining three groups (p-value: . ). acute graft vs. host disease (agvhd) of any grade at day- was recorded in . % ( ). any-grade agvhd was more common in over-weight % ( ), followed by obese with . % ( ), % ( ) in under-weight and . % ( ) in normal bmi-category (p-value: . ). chronic gvhd was more frequent in over-weight ( . %, ), compared to ) background: hematopoietic stem cell transplantation (hsct) is the standard treatment for children with severe aplastic anemia (saa) who have hla-identical related donor. there is no standard conditioning regimen for children with saa secondary to non-fanconi anemia (fa) constitutional bone marrow failure syndromes such as telomeropathies. we report the outcome of a consistent reduced intensity conditioning regimen in patients with idiopathic saa or inherited bone marrow failure syndromes other than fa who underwent hla matched related hsct methods: children with saa underwent hsct using the following conditioning regimen: fludarabine mg/m , cyclophosphamide mg/m , and atg (thymoglobulin) ( mg/kg) . gvhd prophylaxis included cyclosporin and mycophenolate mofetil. donors were all matched related and bone marrow was the stem cell source. all patients had normal chromosomal fragility testresults: a total of children with saa underwent hsct, females and males. average age was . (range . - . years). all nine patients who were tested for telomere length had short telomeres. pathogenic or likely pathogenic mutations were reported in patients ( ercc l , ankrd , tinf , lztfl ). all donors had normal physical examination, normal cbc, and negative genetic testing if patient mutation is known. all patients engrafted successfully, median time to neutrophil engraftment was (range, - days) and platelet engraftment (range, - days). median infused nucleated cell dose was . (range, . - . x /kg) and cd cell dose was . (range, . - . x /kg). none of our patients had acute gvhd and one patient had mild classic chronic gvhd of the skin that was controlled with topical therapy for a short period. three patients had secondary graft failure in the first-year post transplant. first patient had pancytopenia with loss of donor chimerism and underwent successful second transplant using fludarabine, atg, and melphalan. the second patient had a nonfunctioning graft despite full donor chimerism suggesting that the related donor might be affected and had silent phenotype. the third patient had homozygous ercc l mutation and developed progressive cytopenia with myelodysplastic features few months post-transplant and subsequently underwent myeloablative matched unrelated transplant using busulfan, fludarabine, thiotepa and atg. however, the patient progressed to have acute myeloid leukemia six months post hsct. fifteen patients ( %) have normal cbc and stable donor chimerism. median follow-up duration of days (range - days). one patient with lztfl mutation developed chronic renal impairment five years post hsct.conclusions: hsct using lower dose cyclophosphamide ( mg/kg) as part of fludarabine based regimen was safe and effective in saa patients with shorter telomeres and described genetic abnormalities. optimal conditioning regimen in ercc l -associated bone marrow failure needs to be defined. larger study is needed to confirm our results.clinical trial registry: not applicable disclosure: nothing to declare late effects in patients with hemophagocytic lymphohistiocytosis treated with hematopoietic stem cell transplantation: a review of the literature background:hemophagocytic lymphohistiocytosis (hlh) is an inherited or acquired disorder of immunedysregulation. early diagnosis and immunosuppressive treatment can prevent significant organ-failure. the inherited forms, and some acquired cases can only be cured by hematopoietic stem cell transplantation (hsct). with modern transplant practices, a significant number of patients survive. the purpose of this literature review was to collect data on the frequency and type of late effects in hlh patients surviving after hsct and to examine the association with pre-existing hlh conditions (eg. involvement of the central nervous system (cns) before transplant) and with the pre-transplant conditioning regimens.methods: the medline, embase, web of science and pubmed databases were searched, by two librarians at the karolinska institutet, between may and september according to the preferred reporting items for systematic review and meta-analysis (prisma) statement. the search terms included "hlh", "fhl", "mas", multiple terms for "hsct" and late-effect conditions. inclusion criteria were publications in english that included children between january and may . authors of this review screened all the abstracts of studies against the inclusion criteria.results: only nine papers published between and , with information on late effects in hlh patients who had undergone hsct, were identified. three reports include only small numbers of hlh patients. the remaining papers contain data on long-term survivors with a median follow-up of . years. five papers address neurological sequelae with a reported incidence from - %. the highest incidence was found after a thorough neurological assessment of hlh patients compared to matched sibling controls. however, the association with cns disease before transplant and age at transplant was not clear. patients with ebvassociated hlh seem to have fewer long-term neurological problems. non-neurological late effects are described in papers only, with endocrinological problems, namely short stature, being the most frequent. one paper specifically analyzed poor growth, thyroid dysfunction and vitamin d deficiency in a cohort of patients with non-malignant disorders including hlh who had undergone hsct after a reduced intensity conditioning regimen and found significant abnormalities in all groups.conclusions: data on late effects in hlh patients is scarce and is mostly based on the retrospective evaluation of small national cohorts. the available information indicates that a significant number of patients suffers from problems which affect their daily life, but lack of information does not allow to analyze the association between pre-transplant conditions and long-term sequelae. therefore, a retrospective comprehensive analysis of patients registered in the ebmt and cibmtr registries is currently performed. it will be crucial to better define the frequency and type of late effects in a large cohort. this knowledge will aid counselling prior to hsct, provide guidance for long-term monitoring of these patients, and potentially identify specific risk factors for late effects in this rare patient population.disclosure: nothing to declare. allogeneic stem cell transplantation in patients with mucopolysaccharidosis type ii (morbus hunter)bernd hartz , nicole muschol , matthias bleeke , johanna schrum , ingo müller background: the transplantation of hematopoietic stem cells (hsct) is one of the leading methods of treatment in patients with blood system diseases, primary immunodeficiency syndromes and genetic diseases. at the same time, the quality of life in patients in the long-term after hsct significantly differs from the quality of life of healthy people of the same age. deformations in psychosexual development including problems in the gender identity formation cause social isolation of adolescents, which makes their sexual selfrealization impossible and significantly reduces the quality of their life. the purpose of our study was an assessment of the level of gender identity formation of adolescents and psychosexual development correlation to the normal adolescents of the same age.methods: in a prospective single-center study in , on the base of the department of rehabilitation medicine raisa gorbacheva memorial institute of children´s oncology, hematology and transplantation, we conducted a study of families. the respondents were: ) parents / guardians of patients accompanying them in the process of examination; ) adolescents who underwent hsct treatment and undergo planned examinations at the clinic in the posttransplant period (after d + ), (n = , of which girls and boys, age - years, from the date of hsct - years).the following methods were used to assess gender identity: specially developed questionnaires for teenagers and parents; questionnaire by sandra l. bem (sandra l. bem, ) ; projective techniques "the human picture", "the non-existent animal"; max lüscher´s color choices test.results: the traditional type of gender identity, which characterizes high masculinity among male respondents and high female gender indicators in % of cases, was not revealed. both among girls and among boys, the androgynous type prevails with a tendency toward femininity.on average, adolescents see themselves as a bit more courageous than their mothers, with rare exceptions, regardless of gender. this confirms the thesis that we received in a previous study that parents tend to see and encourage complacency of adolescents of both sexes, passivity instead of leadership, dedication and independence. all % of adolescents who participated in the test demonstrate a shift in the theme of aggression, % have some signs of preventing sexual self-determination, abandoning their body, gender, and age. % of patients do not communicate with their peers. in % of them, negative emotions prevail over positive ones. one third of the test participants demonstrate strong support for rest and minimizing their efforts.conclusions: the characteristics of family upbringing of adolescents who have undergone hsct often contribute significantly to limiting their social experience and lead to specific deformities of individuality, including in the sphere of gender identity. we consider advisable to introduce thematic group counseling of parents within the framework of the "patient's school" in psychological treatment support in the clinic. early diagnosis of the personal aspects of the psychosexual development of adolescents after hsct allows for timely identification of individual problems in this area and identification of general trends in the long term after hsct.disclosure: all authors -nothing to disclose. abnormalities in the morphology of the umbilical cord blood obtained at delivery from children who received a diagnosis of cerebral palsy maciej boruczkowski , izabela zdolińska-malinowska , maciej rojek , dariusz boruczkowski background: embryonal brain tumors are the most common malignancies in infants less than months of age. histologically characterized as undifferentiated small round cell tumors, all are similarly aggressive, have a tendency to disseminate throughout central nervous system and very poor prognosis. we tried to assess the effectiveness of tandem highdose chemotherapy (hdct) with autologous hematopoietic stem-cell transplantation (auto-hsct) in this patient group. methods: from to , infants under months with different primary embryonal brain tumors such as medulloblastoma (n= ), different pnet nos (n= ), pineoblastoma (n= ), atypical teratoid rhabdoid tumor (n= ), etmr (n= ) after surgical resection and induction chemotherapy were planned to receive tandem hdct with auto-hsct. nine patients were conducted only single transplantation because of the development of lifethreatening complications after the first hdct (n= ) or the emergence of early disease progression (n= ). at the moment of hdct patients were in complete remission (cr), patients were in partial remission (pr) and patient had stable disease (sd). the conditioning regimen for tandem auto-hsct were: the first hdct was carboplatin and etoposide, the second was thiotepa and cyclophosphamide, both with intraventricular methotrexate.results: the median follow-up is months (range, - ). the median time to engraftment after the first auto-hsct was background: a series of findings suggest that optimizing natural killer (nk) cell reactivity could further improve outcome after allogeneic hematopoietic cell transplantation (allohct). this could be achieved by killer cell immunoglobulin-like receptor (kir) genotype informed donor selection. an enhanced receptor-ligand model which used kir ds and kir dl donor genotype information to augment nk cell activation and minimize inhibition demonstrated improved survival in one large aml study (boudreau et al, jco ) . likewise, a second model built on the classification of centromeric and telomeric kir haplotype motifs, also predicted mortality after allohct for aml (cooley et al, blood ) . this joint ebmt and cibmtr study aimed at validating the two approaches in an independent cohort of patients with mds or secondary aml.methods: donor samples were retrieved from the collaborative biobank (dresden, germany) and mapped to patient outcome data extracted from the ebmt and cibmtr. genotyping of all kir genes by sequencing exons , , , , , and was performed by high resolution amplicon-based next generation sequencing. the impact of the classifiers on time-to-event outcomes was tested in cause-specific cox regression models adjusted for patient age, a modified disease risk index, performance status, donor age, hla-match, sex match, cmv match, conditioning intensity, type of t-cell depletion and graft type.results: clinical data from patients and corresponding donor genotype information were analyzed. the median age at allohct was . years (range, . to . years). the indication for allohct was mds for % and saml for % of patients. disease risk was low/intermediate and high/very high in % and %, respectively. donors were / matched for % of patients. myeloablative, reducedintensity and non-myeloablative conditioning regimens were used in %, %, and % of patients, respectively. peripheral blood stem cells were the predominant graft source ( % of patients). atg was administered in % and alemtuzumab in % of patients. during follow-up after allohct patients died. in univariable and multivariable analyses of the whole cohort, overall survival and the cumulative incidence of relapse of patients with kiradvantageous versus disadvantageous donors were not statistically significantly different. we could not replicate the pattern of outcomes predicted by the kir dl / conclusions: relapse incidence and overall survival after unrelated donor allohct could not be predicted using the kir dl /kir ds -receptor-ligand model and centromeric/telomeric kir-motif model in this large cohort of patients with mds or secondary aml. this points at the possibility of interactions between nk-cell mediated alloreactivity and disease type or procedural variations of allohct. available information on kir-genes, which have been sequenced but not yet analysed, will be investigated in exploratory analyses.disclosure: the authors have nothing to disclose in the absence of an alternative donor, it is recommended that patients undergo desensitization therapy, especially with high dsa levels (> , mfi). the aim of this study is to analyze the impact of dsa on risk of gf and poor graft function (pgf), and on major outcomes in a consecutive cohort of patients who were systematically screened for dsa before haplo-sct. methods: consecutive patients were candidates for unmanipulated haplo-sct with post-transplant cyclophosphamide (pt-cy) at our center from january to january and were analyzed for the presence of hla antibodies.results: patients underwent haplo-sct. hla antibodies were detected in patients, of them were dsa, while were non-dsa (ndsa). patients out of with dsa were transplanted using the same donor; underwent a desensitization program before transplant.background: a recent study from ebmt comparing matched sibling (msd) versus haploidentical donors transplantations, showed better outcome with msd in adult patients with intermediate risk aml in first remission (cr ). however, a female donor to a male recipient transplant combination is a poor prognostic factor and this study did not address the question whether in this situation, a haploidentical donor transplant might do better. the present study compared the outcomes of allografted male patients according to whether they received stem cells from a female msd or a haploidentical donor, in the intermediate and high risk cytogenetics groups (mrc classification).methods: the study included male patients with cytogenetics transplanted between january and june and reported to ebmt. received stem cells from a msd female donor and from a haploidentical donor ( male and female). the follow up was months ( - ). we studied separately intermediate and high risk patients. multivariate analysis was adjusted on factors differing significantly between the groups.results: -intermediate risk group: male patients received a female msd and a haploidentical transplant. the distribution of group characteristics was even except that in the haploidentical transplant group, donors were younger ( y versus ; p< . ), marrow was more frequently used ( % versus %, p< . ) and the interval from diagnosis to transplant was longer ( . versus . months, p< . ). by univariate analysis at two years post transplant, cumulative incidence (ci) of nrm post haplo was higher ( % versus %, p= . ) and ci of extensive chronic gvh lower ( % versus %; p= . ). lfs post msd and post haplo were % and % (p= . ), os % and % (ns), grfs ( % and %). by multivariate analyses the only significant poor risk factors were the haplo-identical transplant for nrm (hr: . ( . - - )) and the patient age for os (hr: . ( . - . ; p= . ). haploidentical transplantation resulted in less chronic gvhd (hr: . ( . - . ); p < - ), but a lower lfs (hr: . ( . - . ); p= . ). -high risk group: male patients received a female msd and a haploidentical transplant. in the haploidentical group, donors were younger ( y versus ; p< . ), marrow was more frequently used ( % versus %, p< . ) and the interval from diagnosis to transplant was longer ( . versus . months, p= . ). by multivariate analysis, haploidentical transplants were associated with a lower relapse incidence (hr: , ( . - . ; p = . ),a better lfs (hr: , ( . - . ; p = . ),os (hr: , ( . - . ; p = . ), and grfs (hr: , ( . - . ; p = . )(see figure) . the only other significant prognostic factor was patient age.conclusions: this study shows that in a male patient with intermediate risk aml, a genoidentical sister donor remains associated with a better lfs. in contrast, in a male patient with high risk aml in cr , a haploidentical donor may be a better choice than an hla genoidentical sister.disclosure: nothing to declare p abstract already published. hematopoietic transplant for older acute leukemia patients: improved survival with offspring donor in comparison with older-aged matched siblingsyu wang , sheng-ye lu , qi-fa liu , de-pei wu , xiao-jun huang background: post-transplant relapse remains the major cause of death of treatment failure. therapeutic options for relapse after first allogeneic stem cell transplant ( st hsct) include chemotherapy followed by donor lymphocyte infusion or second allo-hsct ( nd hsct) from the original donor or change to another donor. however, there is unclear outcome for different treatment approach. in this retrospective cohort study, we aim to compare the clinical outcome after different treatment strategy for relapse after first allo-hsct.methods: between jan and oct, consecutive patients receiving st hsct registered to the bmt database in national taiwan university hospital were analyzed. among them, cases had relapsed after first allo-hsct. one hundred and three patients who received no treatment after relapse or with incomplete data were excluded. their transplant data was collected following the ebmt registry data collection forms and manuals. overall survival rate and progression free survival rate were performed by the kaplan-meier method. univariate and multivariate analysis were performed using cox proportional hazard regression model.results: of the patients who experienced relapse after st hsct, total patients ( %) received chemotherapy followed by dli or nd hsct from the same donor (no change group), patient ( %) received chemotherapy followed by nd hsct from different donors (change group), and ( %) had conventional chemotherapy alone. the patients in "change group" were younger (median age vs , p = . ), and had more patients achieving complete remission (cr) prior to nd hsct ( % vs %, p = < . ) than patients in "no change group". after the nd hsct, the cr was % for "no change group" and % for "change group". the progression-free survival at -year and -year were . % and . % (fig a, p = . %), respectively, for "no change group" and . % and . %, respectively, for "change group". while the overall survival (os) at -year and -year were . % and % (fig b, p = . %), respectively, for "no change group" and . % and %, respectively, for "change group". those who achieved cr prior to nd hsct had a trend of better os than those without cr ( . % vs . % at -year; . % vs . % at year, p =. )( fig c) . there were cases survived for more than years in "change donor group" and cases survived more than years in "no change group". only one had developed relapse after nd hsct but achieved subsequent remission again.conclusions: our study shows that change donor had similar poor outcome comparing to those using the same donor after the st hsct. patients who achieved cr before nd hsct had a trend of better os than those without remission and the long-term survivors were only those who achieved cr prior to nd hsct. novel therapy for cr induction would be warrant for this poor prognostic population.disclosure: nothing to declare functional relevance of fetal microchimerism in nk cell cytotoxicity against leukemic blasts in children: a role for hla-c and kir dl /s ?background: allogeneic stem cell transplantation (allo-sct) remains the most effective curative intent therapy for patients with unfavorable risk acute leukemia. various donor options are available for the patient who lacks an hla-matched sibling donor, such as unrelated donors (urd) and hla-mismatched family (haploidentical) donors. in order to discover the exact role of transplantation type, there are many retrospective analysis, which compared these donor sources, have been reported. recent studies showed some promising results of haploidentical donor transplantation (hidt) using post-transplant cyclophosphamide in comparison with unrelated donor. the goal of this study was to compare the outcome of allo-sct from haploidentical versus matched unrelated (mud / ) or mismatched unrelated donor at a single hla-locus (mmud / ) for patients with acute leukemia in remission.methods: ninety-six adult ( - years) patients with acute leukemia in first or second remission who underwent allogeneic transplantation with a minimum days follow-up at florence nightingale hospital hematopoietic stem cell transplantation center between and were included in this study. patient characteristics and medical records of all patients were reviewed retrospectively. thirty-eight patients who received haploidentical donor transplantation were compared with patients receiving a mud / and receiving a mmud / . patients who completed minimum days post-transplantation follow-up were identified as eligible for survival analysis.results: the characteristics of the patients and transplant donors in this study are summarized in table . median age of patients was . ± years. proportion of male patients was . %, . % and . % for mud / , mmud / and hidt groups, respectively, which is significantly different (p= . ). the other baseline factors were similar, including patient age, donor age, recipient cytomegalovirus (cmv) status, donor cmv status, graft versus host disease incidence, median neutrophil and platelet engraftment times and disease status at post-transplant th day. no significant difference was identified in survival analysis among the mud / , mmud / and hidt groups, even if they were classified according to primary disease (aml vs all) and pre-transplant disease status (cr vs cr ). also, donor cmv status (cmv igg positivity or negativity) was not an important factor on survival analysis when compared between these three groups (p= . ).conclusions: in our study population, clinical outcomes of hidt patients were inferior to mud / and mmud / groups. when choosing an alternative donor for patients without an available hla-matched sibling, urgency of transplantation and host/donor features should be considered. we believe that hidt might be a feasible alternative choice in this subset of patients.disclosure: nothing to declare p g-csf primed bone marrow in hla-haploidentical transplantation using post-transplantation cyclophosphamide (ptcy) could promote tolerance and further reduce risk of gvhd nadira durakovic , , zinaida perić , , lana desnica , ranka serventi-seiwerth , mirta mikulić , brian melamed , alen ostojić , dražen pulanić , , pavle rončević , zorana grubić , radovan vrhovac , implementation, development, and coordination of unique quality management systems with evaluation audits, intrahospital and international accreditation and certification processes. quality of health care is a major focus for providers, patients, and accreditors; so, in this study, we aim to compare the quality of bm harvested at ipo-collection centre (icc) with the quality of bm received from external collection centres (ecc) during these last years.methods: this retrospective evaluation included the number of total nucleated cells (tnc) requested by the transplant centre, the tnc collected, and the results of bm microbiological analysis performed; donor age, weight and infectious disease markers (idm); patient demographics and diagnosis. bm collection technique in use at icc was validated according to jacie standards.we consider successful a collection (sc) with tnc between and % of the requested value, unsuccessful (uc) if lower than % and outstanding (oc) if over %.results: a total of bm was collected, for allogenic ( unrelated) and seven for autologous transplant; unrelated bm were received from ecc (nine from germany, seven from usa and five from portugal). patient main diagnosis were severe aplastic anaemia (n= ), acute myeloblastic leukaemia (n= ), and acute lymphoblastic leukaemia (n= ). donors idm were all negative and nonreactive.mean age (±standard deviation, sd) was (± . ) and (± . ) years for icc and ecc donors, respectively. at icc, we were asked to collect an average (±sd) of . * (± . ) tnc while ecc were asked for . * (± . ). we collected . * (± . ) and received . * (± . ) tnc. correlation between requested and collected tnc was . at icc and . for ecc.we had . % sc and . % oc meaning an accomplishment of . %. we failed to collect required tnc in . %. although % of received bm fulfil tnc requirements, bm processing lowered this value to % due to erythrocyte removal (seven patients with major abo incompatibility) and plasma reduction (two patients with abo minor incompatibility). these steps reduce final tnc available for transplant. weight difference between donor and patient had no significant impact on final tnc collection performance.sixteen bm from icc (seven staphylococcus spp., five propionibacterium acnesand fourcorynebacterium spp.) background: success of peripheral blood stem cell (pbsc) collections depends on patient biological parameters and stable apheresis device performance. peripheral blood cd + cell enumeration is the most reliable predictive factor of apheresis yield however, there are some unexpectedly poor cd + cell harvests despite successful mobilization. the aim of the study was assess total collections cd + yields and factors influencing main apheresis procedure outcomes including collection efficiency (ce).methods: of consecutive donors covering the period - - to - - were analyzed for the following parameters: pre cd count, cd yield per procedure, total cd dose collected per patient, cd collection targets requested by clinical teams. the efficiency of pbsc procedures was determined by calculating the ce and the correlation coefficient between pre cd count and yield per procedure. ce was correlated to preprocedure wbc, platelet count, pre cd count and blood volume processed. all pbsc collections were performed by optia spectra across units in uk.results: of the donors, were autologous and allogeneic. the autologous donors underwent in total procedures. the median cd target dose for these donors was x /kg. ( %) achieved the target dose with procedure and ( %) with procedures. the median pre cd count was /μl. the median cd yield per procedure was . x /kg and the median total cd dose collected per donor was . x /kg. ( . %) of autologous donors collected a total cd dose < x /kg, of those ( . %) had a pre cd count < /μl and ( %) > /μl.the allogeneic donors underwent in total procedures. the median cd target dose for these donors was x / kg. allogeneic donors ( %) achieved the target dose with procedure and ( %) with procedures. the median pre cd count was /μl. the median cd yield per procedure was . x /kg and the median of total cd dose collected per donor was . x /kg. ( . ) % of allogeneic donors collected a total cd dose < x / kg, of those ( . %) had a pre cd count < /μl and ( . %) > /μl. the median ce for autologous donors was % (range - ) and for allogeneic donors was % (range - ). the ce was negatively correlated to wbc (r= - . and - . ) and platelet count (r=- . and - . ) for auto and allogeneic donors respectively, but did not correlate to the pre cd and blood volume processed. the correlation coefficient between pre cd count and cd yield per procedure was r = . for the autologous and r = . for the allogeneic collections.conclusions: the majority of autologous and allogeneic donors achieved the target cd dose with one procedure. . % of autologous and . % allogeneic donors collected a transplantable cd dose of > x /kg. % of autologous and . % of allogeneic donors did not collect a transplantable dose despite a precd count of > /μl indicating suboptimal procedure performance. the ce was variable and was negatively correlated to the preprocedure wbc and platelet count. the ce and correlation coefficient are lower in allogeneic donors compared to autologous donors.disclosure: nothing to declare the outcome of autologous blood stem cell collection and its actual use in real world: the st century experiencekyoungmin lee , jung yong hong , dok hyun yoon , jae-lyun lee , shin kim , kyoung min lee , jung sun park , cheolwon suh background: mobilized peripheral blood stem cells (pbscs) have largely replaced bone marrow as the graft source for allogeneic stem cell transplantation. pbscs mobilization with g-csf is highly effective even on the th day in order to collect enough number of stem cells. a longitudinal, prospective, observational, single-center, cohort study on healthy donors (hds) was designed to identify predictors of cd + cells on the th day. methods: as potential predictors of mobilization, age, sex, body weight, height, blood volume as well as white blood cell count, peripheral blood (pb) mononuclear cells, platelet count, hematocrit, and hemoglobin levels were considered. two different evaluations of cd + cell counts were determined for each donor: baseline (before granulocyte colony-stimulating factor [g-csf] administration) and in pb after g-csf administration on day . a total of consecutive hds with a median age of . years were enrolled.results: the median value of cd + on day was cells/μl (iq - ). basal wbc, plt and basal cd +, are significantly higher for group with cd + on the th day over the median than below. a multivariate quartile regression analysis, adjusted by gender, age, basal cd + and basal plt, shows a, progressively steeper, relationship between baseline cd +, basal plt and cd + on the th day. the basal cd + cut-off for prevision of cd + on the th day was < = cells/μl and >= cells/μl whereas basal platelets count was < = x /l and >= x /l.conclusions: g-csf can be highly effective in hds on the th day in order to collect enough number of stem cells and we have developed a model for predicting the probability to perform pbsc collection after a short course of g-csf.disclosure: nothing to declare p pre-apheresis peripheral blood cd + cell counts highly correlates to actual stem cells collected background: prediction of stem cell yield on the basis of pre-apheresis cd + cell count and the processed blood volume is essential for the planning and executing of the apheresis process.methods: data analyzed included donor weight, complete blood count and cd + count on day of collection, total processed blood volume, cd + cell dose collected in the apheresis product and the number of aphereses performed. using the method described by pierelli et al, predicted cd + yields were calculated: predicted cd + yield x /kg = (benchmark ce x volume of blood to be processed x peripheral cd + count per μl) / (patient's weight in kg x metric conversion factor).results: in we established the method described by pierelli to predict the cd + cell yield. allogenic aphereses were performed in with this approach. mean processed volume was . liters. the mean cd + peripheral count before apheresis was /ul, the mean collected cd +count per kg bodyweight recipient was . . pearson´s correlation coefficient (r) between predicted yield using pre-apheresis cd +count and actually collected cd + cells per kg bodyweight recipient was . . the mean difference between predicted and collected cell dose was + . %.with knowledge of the predicted stem cell count, we were able to adjust apheresis procedure. in case of marginal predicted yield compared to the requested cell dose, we increased the blood volume to be processed. this proceeding led to a significant reduction of second day donations in by % compared to . in only cases we saw more than - % lower cd + doses collected than initially predicted. all donors showed mild iron deficiency with rbc microcytosis, a factor known to affect apheresis procedure.conclusions: pierellis method of calculating the stem cell yield shows a good correlation between pre-apheresis cd + count and actual collected stem cells, making planning and adjusting of the apheresis procedure more feasible and reliable. this proceeding led to significant reduction of second day donations. attention should be paid to iron deficiency anemia, leading to lower than estimated cd + dose.[ background: for more than a decade many transplant centers routinely collect and cryopreserve two or more peripheral blood stem cell (pbsc) grafts for a tandem and/ or salvage autologous blood stem cell transplantation (absct) in patients with hemato-oncological diseases. however, subsequent high-dose chemotherapy (hd-cht) and absct is in many cases not performed for a variety of reasons, specifically in patients with aml, all, mpn and burkitt lymphoma. data about the actual utilization rate of the cryostored stem cell products are lacking.methods: we retrospectively analyzed the collection, storage and disposal practices of pbsc products from a large cohort of patients who were treated at the university hospital heidelberg or at the university medical center mannheim during a -year period. disease entities included acute myeloid leukemia (aml, n= ), acute lymphoblastic leukemia (all, n= ), mpn (n= ; primary myelofibrosis [pmf], n= ; chronic myeloid leukemia [cml], n= ; secondary fibrosis/essential thrombocythemia [et], n= ; not specified, n= ) and burkitt lymphoma (n= ). patients between and were included and followed until .results: an adequate stem cell graft was defined as ≥ . x exp cd + cells /kg body weight. % of the patients were able to collect at least one stem cell graft and the median number of grafts per patient was (range - ). we could demonstrate that only % of all patients who had collected sufficient pbscs for transplant subsequently underwent an absct. among the disease entities the actual use of the stored pbsc grafts varied considerably from % to % (figure ) .conclusions: we could identify striking discrepancies between the collection/storage and actual utilization of background: biosimilars (bio) of granulocyte colony stimulating factors (gcsf) were approved several years ago on the basis of some studies that indicated similar efficacy to the already patented gcsf (neupogen®, neu) both in terms of shortening the neutropenic period after chemotherapy as well as peripheral blood stem cell mobilization in patients with lymphoma and multiple myeloma (mm) treated with autologous stem cell transplantation (auto-hct). however, all these studies are retrospective and there are still concerns about the real efficacy of bio and even more about the real benefit on final costs.methods: we have retrospectively compared the characteristics of the mobilization procedure in both patients with mm and lymphomas, and healthy donors that received either neu or bio (with no chemotherapy) in university hospitals in catalunya from december / to november . bio replaced neu in june in all institutions. primary objectives were the mobilization rate (defined as the percentage of patients that achieved ≥ x /ml cd + cells in peripheral blood on day ) and the use of plerixafor (plex) in each group as pre-emptive strategy. a multivariable analysis of risk factors influencing the use of plex and mobilization failure (defined as collection of < x /kg cd + cells) was also performed.results: we treated patients ( lymphomas and mm) and healthy donors. both groups of patients ( neu and bio) and donors ( neu and bio) were comparable regarding pre-mobilization general characteristics. there was a trend for a lower median cd + peak on day for bio patients ( vs , p value = . ). a total of patients received plex, although of them ( . %) out of strict theoretical indication, cd + cells > x /ml (range . - . ) and were removed for further analysis (n = , in the neu group and in the bio group). median number of cd + cells on day was significantly lower in the group bio who needed plex ( . vs . for neu+plex, p= ), as well as cd + cells finally harvested ( . vs . x /kg, p= . ). mobilization failure rate was higher in bio group ( vs %, p= . ). regarding no plex patients, median number of cd +cells on day was also significantly lower for bio patients ( . vs . , p= . ). risk factors for plex use were age, basal disease (lymphoma) and number of prior mobilization therapies. the use of bio was the only risk factor for mobilization failure patients receiving plex [hr . ( %ci . - . ), p= . ]. with respect to healthy donor mobilization, none of them needed plex but cases from the bio group ( %) needed more than one apheresis procedures ( and , respectively).conclusions: we found a lower efficacy of bio in the setting of stem cell mobilization of patients with only gcsf both in terms of a lower cd + cells peak on day and a lower number of cd + cells in final apheresis product. bio gcsf also seems to be less effective in healthy donors.disclosure: no conflict of interest to be declaredbackground: auto-sct is a common treatment in patients with mm or nhl. the aim of the prospective multicenter goa (graft and outcome in autologous transplantation) study was to investigate the impact of mobilization method used on the cellular composition of collected blood grafts and eventually hematological and immune recovery as well as long-term outcome post-transplant. altogether patients with mm or nhl transplanted between / and / at four university hospitals were included. the long-term goal of the study is to evaluate characteristics of optimal blood grafts in regard to post-transplant recovery and outcome. methods: altogether patients with mm undergoing first auto-sct were compared with patients with nhl. all nhl patients were mobilized with chemotherapy + g-csf, whereas % of mm patients were mobilized with g-csf only (p < . ). mobilization data, graft cellular composition including cd + cell subsets and lymphocyte subsets of the blood grafts, post-transplant hematological recovery and outcome were evaluated. the median followup time was months in mm patients and months in nhl patients.results: mm patients mobilized cd + cells better (median peak blood cd + vs. x /l, p < . ). the median number of aphereses was in both groups (p = . ). altogether % of the nhl patients received plerixafor compared to % in mm patients (p = . ). the median number of cd + cells collected was higher in mm patients ( . vs. . x /kg, p < . ).the median amount of cd + cells (with -aminoactinomycin) in the infused graft was . x /kg in mm group and . x /kg in nhl group (p = . ). the grafts contained more nk cells (median . vs. . x /kg, p = . ) and cd + cells (median . vs. . x /kg, p < . ) in mm patients. neutropenic fever tended to be more common in nhl patients ( % vs. %, p = . ) but mm patients had significantly more bloodstream infections ( % vs. %, p = . ). the median duration of hospitalization was longer in the nhl patients ( d vs. d, p < . ) and the nhl patients had significantly more often icu admissions ( % vs. %, p = . ).post-transplant neutrophil engraftment was faster in the nhl group (median d vs. d, p < . ). the median time to platelet engraftment was days in both groups. platelet count was higher in the mm group from day until year after auto-sct. there were significantly more early deaths (< d from the graft infusion) ( % vs. %, p = . ) and non-relapse deaths ( % vs. %, p = . ) in the nhl group.conclusions: mm and nhl patients differ in terms of cd + cell mobilization, graft cellular composition and post-transplant recovery as well as risk of non-relapse death. thus, the optimal graft may be different in nhl and mm patients.disclosure: the study was supported by vtr fund from north savo hospital district and study grant from sanofi. abstract already published. single-center experience in use of plerixafor for autologous stem cell mobilization: change in practice over years background: plerixafor has been proven to mobilize human periferal blood stem cells (pbscs) alone or acting synergistically with granulocyte-colony stimulating factor (g-csf). it has mainly been used for rescue mobilization after failed regimen of chemotherapy plus g-csf, but lately preemptive use in poor mobilizers has been established as cost-effective. we aim to show ten years of experience and change in practice with plerixafor use in our center.methods: we retrospectively evaluated the outcome of mobilization procedures and leukapheresis collections in our center in the period from january to october . practice from the first years, when plerixafor was mainly used as rescue agent after failed attempt, was compared to period from till present when preemptive use in poor mobilizers (defined as cd + cell counts < x /l blood) was established.results: in the period from to , total of patients underwent collection of autologous pbscs, and patients required repeated mobilization cycles ( , %). g-csf alone was used in patients and patients ( %) recieved combination g-csf with plerixafor. this cohort consisted of males and females with non-hodgkin (nhl) and hodgkin lymphoma (mh); and respectively. we noted unsuccessful mobilizations ( , % in repeated mobilization, , % in total) of which one patient was from plerixafor group ( , %).background: transplantation of hla-haploidentical hematopoietic stem cells (haplo-hsct) is an established procedure for the treatment of several different hematological diseases. one possible strategy to reduce the risk of graft-versus-host disease is represented by the selective depletion of ab t-lymphocytes (coupled with the depletion of cd + b-cells in order to reduce the risk of ptld) using the clinimacs device (miltenyi, bergish-gladback). before depletion, leukapheresis units are washed by lowspeed centrifugation, resulting in a platelet (plt) rich supernatant (prs) as a by-product generally discarded. we studied the possibility of recovering plt from prs of the haplo donor for transfusion to the recipient during the aplasia period occurring after hsct.methods: hsc donors were mobilized with g-csf (plus plerixafor in out of donors) as previously described [locatelli et al, blood ] . leukapheresis units, obtained with a spectra optia device, were washed twice (producing prs bags) at low speed to remove plt before starting the ab t-cell/b-cell depletion. the two prs were leukodepleted by filtration, centrifuged, resuspended and pooled in a total volume ranging from to ml intersol (is-plt) for overnight incubation at °c with agitation ( cycles/min). the is-plt samples were analyzed for the criteria established by the italian transfusion law. is-plt bags were examined for the following parameters: volume > ml, plt after leukodepletion > x , residual leukocytes < x , ph > . at °c at the end of the -day storage period. the sterility was tested using bd bactec culture vials. the evaluation of the residual leukocytes was performed with the bd leucocount kit. the absolute plt counts were determined using hemocytometer sysmex xn .results: prs bags from donors were processed to produce is-plt units. median resuspension volume in intersol was ml (range - ). the absolute mean value of plt counts measured at the end of the storage period was . x (range . - . x ). this value was found below the threshold fixed by italian regulation in cases ( . %). mean value of residual leukocytes was . x (range - x ); the ph value was always > . . sterility was observed in all cases. according to the work of slichter et al. conclusions: we demonstrated that plts recovered from leukapheresis bags can be accepted as a conventional hemocomponent according to the parameters fixed by italian transfusion law and thus can be administered to the haplo-hsct recipients early after transplantation. this strategy carries several advantages. indeed, apart from the obvious advantages in terms of reduced costs, is-plt can be used to desensitize the recipient by absorption of anti-hla class i antibodies, if present in the recipient. moreover, this strategy can avoid the risk of sensitizing the transplanted patients with hla alleles that differ from the donor's ones. finally, the is-plt unit is readily available. a clinical study aimed at testing the use of is-plt units in transplant recipients will be performed to confirm the clinical efficacy of the approach.disclosure: nothing to declare p ultrasound guidance as a powerful tool in increasing background: peripheral blood stem cells are generally the preferred graft source for allogeneic stem cell transplantation for malignant disease. in most centers first apheresis is performed on day after to doses of granulocyte colony stimulating factor (gcsf) up to ug/kg twice daily. the dose of gcsf and the number of apheresis procedures required contribute to symptom, travel and time burden donors are put through during the process. we hypothesized that taking donor-recipient weight differences into consideration may help reduce this burden methods: a total of healthy donors who donated peripheral blood stem cells on day of gcsf mobilization in the period between january and august at the university medical center hamburg-eppendorf were included in this quality control evaluation. the donors were divided into two cohorts. the impact of donorrecipient weight ratio on stem cell harvest was tested in the training cohort ( - ) and validated in the second cohort ( ) ( ) . for the training cohort, donors were grouped according to donor-recipient weight ratio < . vs. . - . vs. > . . for the purpose of this analysis a stem cells dose of x cd +/kg recipient weight was set for successful apheresis.results: in the training cohort including donors, ( %), ( %) and ( %) had a donor-recipient weight ratio of < . , . - . and > . respectively. the target stem cells count of x cd +/kg recipient weight was achieved in of ( %), of ( %) and of ( %) donors with donor:recipient weight ratio < . , . - . and > . respectively. the cut-off for the validation cohort was therefore set at a weight ratio of . .in der validation cohort including donors, ( %) had a weight ratio > . while ( %) had a weight ratio ≤ . . overall in this cohort target cell count of x cd +/kg recipient weight was reached in ( %) cases. this target was reached in of ( %) of donors with weight ratio ≤ . and in of ( %) donors with weight ratio > . , p = . .conclusions: a donor-recipient weight ratio of > . is seen in about % of peripheral blood stem cell donations for allogeneic stem cell transplantation. in these cases apheresis on day after doses of gcsf is reasonable. donors with lower weight ratios should preferentially donate on day after to doses of gcsf.disclosure: all authors declare no conflicts of interest background: the effect of a second mobilization and collection of peripheral blood stem cells (pbsc) on the cell yield is low, as we previously demonstrated. however, donor safety has been poorly addressed with no changes in the clinical practices.methods: second donations of unrelated and related donors performed between and were evaluated (n= ), including pbsc+pbsc (n= ), bone marrow (bm)+pbsc (n= ) and pbsc+bm (n= ). analytical parameters including leukocyte, lymphocyte, hemoglobin and ldh quantification, obtained on the pre-harvest evaluation of first and second donation, were retrospectively analyzed and compared for all donors. the portuguese bone marrow donors registry (cedace) recommends a time between donations no lesser than months. it also states that in very urgent situations like graft failure, donor should be clinical and analytical cleared and its safety ensured.in order to evaluate the impact of time between donations, donor population was divided in groups: < months, - months, > months; to determine the influence of donor age, donors were divided in groups: < and ≥ years.results: among the total of donors, were volunteer donors of cedace and were familiar. fifteen second donations were performed because of recipient graft failure and due to disease progression or relapse. at the time of second collection, median donor age was years (range - ). the median delay between both collections was days ( - ). time between donations did not seem to substantially impact the analytical donor evaluation: leukocytes, lymphocytes, hemoglobin and ldh results are kept within the reference values. however, donors with less than months between donations showed a slight decrease on leukocyte counts ( %) and hemoglobin values ( %), from the first to the second pre-harvest evaluation. donor age showed no significant influence on the analytical evaluation. nevertheless, when considering only the pbsc+pbsc donations, donors with ≥ years showed a small decrease on lymphocyte counts ( %) .conclusions: this study demonstrated that the analytical parameters, chosen based on literature, had no significant changes between first and second donation. however, particular attention should be paid when time between donations is lesser than months or donor age is ≥ years.as we concluded that no significant changes were observed in the group of - months, it is our opinion that the minimum of - months established by the registries can be shortened to months ensuring donor safety. an accurate donor risk assessment with a larger population should be accomplished in order to strengthen this recommendation.disclosure: nothing to declare. gaurav kharya , atish bakane , pratibha dhiman , anil khetrapal , vikrant bhar background: t cell replete haploidentical stem cell transplant (hhsct) is complicated mainly by increased risk of graft failure (gf) and graft versus host disease (gvhd). conventionally gcsf has been used to mobilize hematopoietic stem cells (hsc). in tcr hhsct gcsf mobilized graft with megadose of cd + cells expose the patient to higher doses of alloreactive t cells increasing the risk of gvhd. plerixafor based mobilization gives an advantage of giving high cd cell dose limiting exposure to high alloreactive t cell dose. we share our experience of gcsf + plerixafor based mobilization for tcr hhsct. methods: consecutive patients suffering from scd who underwent hhsct between jan till date along with the respective donors were enrolled in the study (group ). all underwent pre-transplant immune suppression (ptis) cycles at weekly intervals using fludarabine+cyclophosphamide+dexamethasone.the graft was mobilized using gcsf@ mcg/kg/day(d -d )+plerixafor@ . mg/kg(d ) - hours before the pbsch. gvhd prophylaxis included ptcy mg/kg/ day on d and , sirolimus and mmf starting from d . group included historical controls where graft was mobilized using gcsf@ mcg/kg/day(d -d ). various parameters pertaining to mobilization, harvest, engraftment, gf and gvhd were assessed between the two groups.background: poor mobilizers (pm) defined as those with a peripheral blood cd + count ≤ cells/μl on day+ is a significant risk factor for mobilization failure. within these, patients with < cells/μl are considered as very poor mobilizers (vpm). use of plerixafor in vpm patient is controversial. the aim of our study is to compare mobilizing and engraftment between pm and vpm who received plerixafor plus g-csf (p+g-csf).methods: in our center, mobilization with g-csf at dose of μg/kg/day was used in all pts. apheresis were scheduled on day+ . plerixafor ( . mg/kg) was added if the number of cd + cells on day + was < /ul for x cd + /kg requested (or < /ul for x cd + /kg), or if the number of cd + cells collected in the first apheresis was < % of cd + cells requested.between january and september , out of pts ( , %) received plerixafor for mobilization. we retrospectively studied pts who mobilized with p+g-csfdue to the number of cd + cells on day + was < /ul.results: twelve out of pts were pm, were females, median age , years (range: - ). patients' baseline diseases were: non-hodgkin lymphoma (nhl) ( , %), multiple myeloma (mm) and hodgkin lymphoma. median cd + cell count on day + was / ul (range: - ).there was no mobilization failure. eighteen out of pts ( %) were vpm, were females, median age , years (range: - ). patients' baseline diseases were: nhl ( , %), mm and solid tumor. median cd + cell count on day+ was , /ul (range: - ). two out of pts ( , %) were considered mobilization failure, in of them did not realized apheresis due to cd + cell count on day + was /ul. no difference was seen between both groups regarding gender, age, patients baseline disease or median cd + cells count on day + .vpm needed more apheresis sessions, / pts required sessions against / pts in pm (p=not significant (ns). we obtained enough cells to carry asct in % pts, although mean number of cd + cells obtained in vpm was lower than in pm ( , x /kg vs , x /kg, respectively) (p=ns).twenty-six pts underwent asct and mean number of cd + cells infused were , x /kg in vpm vs have been the only route used for chpc administration. the appearance of other catheters types made us to reconsider the exclusive use of the cvc for the infusion of chpc. we analyzed the use of a peripheral iv cannula (pivc) as an alternative to cvc for the infusion of chpc in patients with cardiovascular diseases.methods: medical records of patients who received an asct for hematological malignant diseases at the hospital clínic of barcelona from january to february were reviewed. of those, eight were infused through a pivc due to cardiac impairment related to previous treatments, ischemic cardiomyopathy or amyloid deposition.hpc were obtained from peripheral blood by apheresis after mobilization with g-csf using acd-a as an anticoagulant. cryopreservation was performed with autologous plasma and dmso % by mechanical means and stored in liquid nitrogen. analytical controls were performed including hematocrit, total nucleated cells, total polymorphonuclear neutrophils and platelets using the advia analyzer. the cd + / cd + population was analyzed by flow cytometry following the ishage single-platform protocol. viability of total nucleated cellularity was carried out by vital staining with acridine orange and the specific viability of the cd + population through the technique of -aminoactinomycin d. thawing was performed bag to bag by immersion in a water bath at ºc and transferred to the bedside of the patient for gravity infusion using an infusion set without filter through pivc of gauche. vital sings monitoring performed before, during and the end of the every infusion bag including: blood pressure, heart rate, oxygen saturation, body temperature and central venous pressure. other aspects assessed during the infusion were pain, cold sensation and signs of extravasation in the area of pivc insertion. after the infusion, the recovery time of the granulocyte series and platelet were evaluated.results: median volume and bags administered was ( - ) ml and ( ) ( ) ( ) . the median of total nucleated cells, total nucleated cells / ml and total cd + cells/kg was . x ( . - . ), . x ( . - . ) and . ( . - . ) respectively. vital signs were within the normal range and allowed to perform the infusion in an average of - minutes/bag. no patient required stopping the infusion due to pain in the area of peripheral catheter insertion and no extravasations were detected. all patients referred some cold sensation in the insertion vein and its path. median hematopoietic recovery was ( - ) days for neutrophils and ( - ) days for platelets, similar to the recovery experienced from patients who received chpc through cvc.conclusions: based on our data, we conclude that the administration of chpc, through pivc and by gravity is safe for the product and for the patient, being the preferred choice for patients suffering from some type of cardiovascular disease.disclosure: nothing to declare background: by selective depletion of potentially alloreactive cd ra + cells, t memory cells might be retained in the graft and could mediate pathogen specific immunity. however, cd ra expression is not restricted to naïve t cells, but also available on b cells, nk cells and cd + stem cells to some extent. methods: within this project we aim to analyze cd ra expression on stem-and nk cells by flow cytometric analysis to estimate the eventual loss of these cells during cd ra-depletion. furthermore, clinimacs depletion following a one-step approach of direct cd radepletion and a two-step approach with primary cd selection followed by cd ra-depletion of the negative fraction was investigated.results: cd ra expression on cd + stem cells was in median . %. with a median of . % cd ra expression was measurable on nearly all b cells, which obviates depletion via cd . a comparably high cd ra expression of in median . % was detected on nk cells. unfortunately, the amount of nk cells in the cd ra-depleted product was . %. clinimacs depletion following one-step approach resulted in a stem cell recovery of . %. memory t cell recovery was . % following one-step and . % applying two-step approach. depletion quality measured by log-depletion was . and . for cd ra + t cells and . and . for cd + b cells for one-and two-step approaches, respectively.conclusions: with regard to stem cell recovery, a previous cd -selection before cd ra-depletion is recommendable.background: current clinical practice of routine use of filters for infusion of autologous hematopoietic cell transplantation (ahct) at bone marrow transplant centers across north america and europe is not known. the use of "y" administration tubing without a filter could possibly increase the risk of infusion of macro-aggregates and cellular debris, which may result in increased side effects.methods: we carried out a retrospective chart review of patients (pts) at spectrum health who underwent ahct. group a (gp a) pts received ahct using a "y" administration tubing with -micron filter from / - / . these patients were compared to a control group (gp b) that received ahct without filter administration tubing from / - / .this change in clinical practice occurred due to a change in policy at our transplant center as a result of inorganic particles noticed during cryopreservation. we compared the neutrophil and platelet engraftment duration between these groups. we also studied the length of hospital stay and the effect of filter use on any immediate side effects after infusion. due to the retrospective nature of the study it was not feasible to evaluate the difference in duration of infusion between these groups results: the two groups were similar in their age, gender, primary disease distribution and median number of cd stem cells infused (table) . there was no difference in median neutrophil ( vs days) or platelet engraftment ( vs days) duration for the filter group and the nonfilter group respectively. the median length of hospital stay was also comparable ( days). there was no statistically significant difference in the immediate side effects (fever, cough, dyspnea, fluid overload, flushing, nausea, vomiting, hypertension, hypotension and anaphylaxis) or confirmed post-transplant infections (viral, bacterial, fungal) experienced by these two groups.conclusions: our results show that the routine use of filter does not prolong hospital stay, and neutrophil/ platelet engraftment duration, thereby, suggesting that viable stem cells are not affected. on the other hand, filter use failed to demonstrate any appreciable decline in the immediate side effects experienced after ahct. gp background: allo-hsct from related haplo-identical donors (haplo-hsct) with post-transplant high-dose cyclophosphamide is increasingly employed in patients who lack a matched related or unrelated donor. the current standard is to use bone marrow grafts (bm) as peripheral blood stem cell grafts (pbsc) have been associated with an increased risk of acute and chronic gvhd. thus, the aim of our study was to compare the main transplant outcomes and especially the incidence of acute and chronic gvhd in recipients of bm and pbsc grafts. methods: thirty-five unselected patients with hematologic malignancy who underwent an haploidentical transplant at our unit between and and received bm (n = ) or pbsc (n = ) grafts after the same tbf conditioning regimen were analysed in order to assess differences in transplant outcomes.our gvhd prophylaxis consisted in cyclosporine a (csa) from day - to + , a methotrexate "short course" and mycophenolate mofetil (mmf) from day + to + .results: no statistically-significant differences were observed between patients who received bm grafts and those who received pbsc grafts. at transplant fourteen patients were in first complete remission (cr), twelve in advanced cr and had active disease. according to sorror's risk, nine patients were low-risk, nine intermediate-risk and seventeen high-risk. twenty-eight cmv+ patients received the graft from twenty-three cmv+ and five cmv-donors, seven cmv-patients received the graft from five cmv+ and two cmvdonors. mean age at transplant was years (range - ), mean donor's age years (range - ) and mean follow-up . months (range . - . ). median cd + cell dose was . x /kg (range . - . ), . x /kg (range . - . ) in bm recipients and . x /kg (range . - . ) in pbsc recipients. median time to neutrophil recovery (> /μl) was days (range - ) posttransplant, days (range - ) for bm recipients and (range - ) for pbsc recipients. platelet recovery (> . /μl) occurred in all patients except one at a median of days (range - ) post-transplant, at a median of days ( - ) post-transplant for bm recipients and at a median of days (range ( - ) for pbsc recipients. seven patients never reached platelets > . /μl. three patients developed a poor graft function. acute and chronic gvhd incidence was . % and . % and the risks of acute (hazard ratio [hr], . ; p = . ) and chronic (hr, . ; p = . ) graft-versus-host disease were similar in the two patient groups. in addition, there were no differences in relapse risks post-transplant (hr, . ; p = . ); relapse-free survival was better with pbsc grafts but this difference did not reach any statistical significance. finally, no significant differences were noted in overall mortality by graft type (hr, . ; p = . ).conclusions: despite in haplo-hsct the incidence of acute and chronic gvhd is reported to be higher with pbsc than with bm our small patient series does not confirm this assumption that should be clarified by additional studies. instead, our data suggest that pbsc background: since , cord blood (cbu) has become an alternative source of stem cells for transplantation, with approximately , procedures currently performed. with a -year gs of %.objective: analyze the outcomes from all the patients transplanted with cbu in our hospital unit.methods: retrospective, longitudinal study. all patients transplanted with cbu in our hospital between and were included. we analyzed patients with ages from months to years.results: two of them received doubled cord transplantation the ratio male: female was . : . the transplanted pathologies were: bone marrow failure %, immunodeficiency %, aml %, all . %, osteopetrosis . %. ric regimens were used in patients with bone marrow failure and immunodeficiency and myeloablative conditioning regimens were used in patients with malignant hematology diseases. antithymocyte rabbit globulin (atg) based serotherapy was used. one case received cbu from a related donor (sister), the rest received unrelated cbu obtained from centro nacional de la trasfusión sanguínea. the infusion of cd + was in a range of . to of . x /kg with and average of . x / kg. compatibility was / in %, / in % and / in %. post-thawing cellularity was not measured. the hla-c was not analyzed. forty two point five percent of the patients had a successful engraftment; the average time of engraftment was days. primary graft failure was detected in . % and secondary graft failure in %, for a total success of . %. gvhd was detected in % of patients, of which % was grade i-ii and % grade iii-iv. the overall mortality was . %. causes of death were: infection % relapse %, hemorrhage % and gvhd %. the cbsct continues to be an essential alternative in our patients who required transplantation knowing that this stem cell source allows the procedure to be done with less histocompatibility requirements and it is available immediately, which facilitates the process considering the great diversity that exists within our population. however, in our experience, the cbsct has shown a higher mortality risk, which can be improved by analyzing the hla c, choosing in this way the units with better compatibility, and improving cellular dosage since this is key in success.disclosure: nothing to declare the risk of infection of the umbilical cord is not related to the microbiological status of the umbilical cord blood methods: the statistical analysis was carried out on data obtained from samples taken in poland between -jan- and -dec- . the samples were collected in hospitals by external midwifes and sent to the pbkm in accordance with the requirements of the american association of blood banks. after arrival in the laboratory, the blood samples were cultured and the ucs were assessed immediately for visual signs of infection, such as odor, altered color, or visible bloom. the status of both kinds of samples was introduced into the pbkm general database. for the purpose of this analysis, the ucs were considered as microbiologically pure if stored, destroyed after storage, or handed over to the pbkm. samples marked as infected or disqualified for unknown reasons (other than termination of the contract with the customer, viral infection of the mother, and lack of cell growth) were considered as infected. at the time of the statistical analysis, the samples of unknown ucb microbiological culture status were removed from the generated report. the data was summarized as percentages and the odds ratio was calculated. statistical significance was considered at p˂ . .background: match family donors are the preferable options in allogenic stem cell transplant. however, in the absence of donor relatives match unrelated donors have been an option. in this study, the donor screening, transplant preparation phases of turkish stem cell coordination center (turkok) and the İstanbul university bone marrow bank (tris), were compared.methods: the unrelated donor scanning data between march and november in pediatric stem cell transplantation unit of altınbas university bahcelievler medical park hospital were evaluated. unrelated transplants were performed in total. % of these transplants (n= ) were included from the donors of turkok registration system and , % of these transplants (n= ) were included by means of tris from donors outside of turkey. patients ( tris, turkok) were excluded from the study in consequence of screening update and postpone of transplantation. the statistics were carried out on a total of patients, , % of whom were in turkok (n= ) and , % were transplanted via tris (n= ). the day of application to stem cell transplantation unit, reply dates and the transplantation dates were examined for the transplant patients.results: in the current study, the average response time of turkok was found as , ± , day (median: ), the average transplant time after receiving a reply was found as , ± , (median: ) day, the average number of days from date of application to date of transplantation of patients was found as , ± , (median: ). the average response time of tris was , ± , (median: ) day, the average transplant time after receiving a reply from tris was , ± , (median: ) day, average number of days from date of application of tris to date of transplantation of patients , ± , (median: ) day.the average response time of turkok, the average transplant time after receiving a reply from turkok and the average number of days from date of application of turkok to date of transplantation of patients was shorter than tris. the difference between them was found statistically significant (p< . ).conclusions: in this study, it was determined that the transplantation processes with turkok were progressing more rapidly. the rapid progress of the process was attributed to the fact that all donor hla tissue groups in the turkok database were studied in high resolution. in international scans carried out through tris, it was thought that the examination of the donor castings coming from bone marrow banks and the time differences between the countries prolong the process. it was thought that hla tests of the registered donors in the tris database and some international bone marrow banks were studied in low resolution but not studied the all hla loci, the centers wanted high-resolution hla, and therefore the involvement of social security institutions and payment procedures were among the factors extending this process.disclosure: nothing to declare background: allogeneic hematopoietic stem cell transplantation (ahsct) is being performed for a group of hematologic diseases with a curative intent. outcomes after ahsct are influenced by the type of donor used. haploidentical transplantation is an emerging option when a fullmatched donor is unavailable. methods: we retrospectively analyzed our transplants performed between january and november , investigating outcomes and complications among haploidentical stem cell recipients.results: one hundred and nineteen patients underwent ahsct, of them ( . %) were recipient of a haploidentical stem cell and included in this study. one patient diagnosed with acute lymphoblastic leukemia (all) were performed a haploidentical ahsct for two times due to relapse. among those transplants, of them were diagnosed with acute myeloid leukemia, with all, with chronic lymphocytic leukemia, with myelodysplastic syndrome and with hodgkin lymphoma. the mean age of group was . ± . years. three patients ( aml, all) were in remission at the time of transplantation. patients were given a conditioning regimen based mostly on busulfan, fludarabin and total body irradiation with a myeloablative intent. patients were also given a various combinations of post-transplant cyclophosphamide, calcineurin inhibitors, mycophenolate mofetil and antithymocyte globulin for graft versus host disease (gvhd) prophylaxis; post-transplant cyclophosphamide administered on ( %) of those transplantations. peripheral blood was the source of stem cells in all patients. patients were infused with mean . ± . x /kg of cd + cells. hematological recovery was achieved with neutrophil engraftment at a mean of . ± . days and platelet engraftment at a mean of . ± . days. after a median month ( . - . months) follow up, the cumulative rates of grade - gvhd, relapse and non-relapse mortality were %, % (n= ) and %, respectively. one patient died due to relapse, at the end of the follow up two were still alive with remission. only one patient has died due to chronic gvhd affecting serosa and resulting with a fatal tracheoesophageal fistula. the mean overall survival was . ± . months in our study.conclusions: haploidentical transplant is a feasible option in hematologic malignancies with novel gvhd prophylaxis approaches, especially post-transplantation cyclophosphamide. however, these results need to be supported with further investigations with a larger patient group.disclosure: nothing to declare results: a total of patients between and years (median age: years). transplant was done for following disease: acute leukemia (n= , %), aplastic anemia (n= , %), lymphoma (n= , %), myelofibrosis (n= , %), myelodisplastic syndrome (n= , %), chronic myeloid leukemia (n= , %). ten donors were from turkey and fifteen donors were from different countries of europe and america. two of donors were / and the other was / hla matched. the conditioning regimen was mostly non myeloablative (n= , %) while eight patients were treated with myeloablative regimen. other than two patients who took tacrolimus and mycophenolate mofetile all of them got cyclophosphamide and methotrexate for graft versus host disease (gvhd) prophylaxis. the median time of neutrophil and platelets engrafman were days (range - ) and , days (range - ) respectively. acute gvhd was seen nearly half of the patients ( , %).overall survival was % for all patients and of patiens ( %) died within first month to months (median months). the mortality rate was more higher for the recipients who had donor source from countries other than turkey ( % vs % p= , ). transplant related mortality was the most common reason of mortality (n= / , , %) and other reasons were gvhd ( , %), infections and cirrhosis respectively.conclusions: we found the mortality rate more higher in the patients whose donors were from out of our country. however, we need to further multicentric and prospective investigations to confirm our hypothesis, it would be related with impact of ethnicity.disclosure: nothing to disclose late-breaking abstracts p targeted twice daily busulfan-based ric-conditioning for allogeneic hematopoietic stem cell transplantation in pediatric patients with chronic granulomatous disease: a -year experience with the zurich protocol matthias felber , mathias hauri-hohl , ulrike zeilhofer , federica achini , jana pachlopnik-schmid , janine reichenbach , seraina prader , tayfun güngör background: chronic granulomatous disease (cgd) needs sufficient myeloablation to avoid graft failure. for this purpose the ebmt inborn errors working party currently recommends treosulfan or busulfan-based conditioning regimens for cgd-sct. we analyzed the last years of targeted busulfan-based ric-conditioning including engraftment, gvhd rates, chimerism and late term effects in our pediatric sct center in zurich. methods: between and , n= consecutive pediatric cgd patients (median age years, range - years, n= female, n= autosomal recessive inheritance) have been transplanted. all patients received therapeutic drug monitoring of twice daily administered iv busulfan ( or hour infusions; d- to -d ) to achieve a targeted cumulative auc of - mg/l*h. fludarabine ( mg/ sqm; d- to -d ) and serotherapy (thymoglobuline . mg/ kg total, d- to d- ) or alemtuzumab ( . - . mg/kg total; d- -to d- ) were used for immunoablation. donors were matched unrelated ( / hla; n= ), mismatched unrelated ( / hla; n= ), mismatched unrelated (hla / ; n= ), matched sibling (hla / ; n= ) and haploidentical parental (hla / ; n= ). for patients with haploidentical donor post-transplant cyclophosphamide (d- and d- with mg/kg iv each) and upfront atg-grafalon ( mg/kg - to-d- ) was used. stem cell sources were bm (n= ) and pbsc (n= ). gvhdprophylaxis included iv csa (d- ; continuous infusion) and iv. mmf (d- , in - doses).results: follow-up was to months. good overall engraftment was noted, with n= secondary graft failure followed by successful retransplantation. in one patient a stem cell boost/dli was necessary due to decreasing myeloid donor chimerism during ebv reactivation, resulting in rapid myeloid donor reconstitution after intervention. low rates of gvhd were documented with n= agvhd grade iv and n= mild/limited cgvhd (nih criteria). with exception of one patient, myeloid donor chimerism at last follow-up was over %, mostly over %. overall survival was / ( %). deaths were due to gi-gvhd (n= ), autoimmune hemolytic anemia/sepsis (n= ) and thrombotic microangiopathy (n= ).conclusions: precision dosing of iv busulfan in combination with fludarabine and serotherapy results in excellent outcome of hsct for pediatric cgd-patients with good engraftment, low overall cgvhd rates and stable, mostly excellent donor chimerism. graft failure rate was as low as %. low dose alemtuzumab prevented gvhd in the majority of patients. this analysis demonstrates that targeted busulfan-based conditioning is a valid option for pediatric cgd-patients. serum alemtuzumab or atg monitoring could further improve gf and gvhd rates in the future.disclosure: the authors declare no conflict of interest. abstract already published. young hla-matched unrelated donors are comparable to matched sibling donors in elderly patients receiving reduced-intensity conditioning: an analysis on behalf of the ebmt scientific council % c.i. . - . , p= . ) and voriconazol prophylaxis during carv (or . , % c.i. . - . , p= . ). conclusions: we provide evidence that ifd after carv infection. allo-hsct recipients developing a carv lrtd during the first year after transplant may benefit from an adequate antifungal prophylaxis and a close monitoring for the development of a later ifd.disclosure: jose luis piñana has received both, advisory for preclinical/clinical research and financial support to assist to the spanish society of hematology annual meeting from msd. favorable outcome and engraftment following reducedintensity conditioned allo-hsct in children with primary haemophagocytic lymphohistiocytosis (hlh) and high-risk langerhans cell histiocytosis (lch) laura m. moser, emilia salzmann-manrique, andrea jarisch, jan sörensen, shahrzad bakhtiar, peter bader background: primary haemophagocytic lymphohistiocytosis (hlh) and high-risk langerhans cell histiocytosis (lch) represent two major entities of childhood histiocytoses, which are -although only of rare occurrence -severe in their clinical manifestations. patients present with multisystemic uncontrolled inflammation and multi-organ involvement requiring diverse courses of immunosuppressive and chemotherapy regimens. allogeneic haematopoietic stem cell transplantation (allohsct) is the only available curative option; however, the cumulative treatment toxicity and the underlying inflammatory disease often result in high organ toxicity and inflammatory complications of transplantation, such as graft versus host disease (gvhd) and/or graft failure. especially patients with unrelated donors often deal with high transplant-related mortality (trm) in the setting of conventional intensity conditioning.herein, we present the clinical course of the disease and transplant outcome of children diagnosed with primary hlh (n= ) and high-risk lch (n= ) who underwent allohsct at our centre from / to / .methods: the hlh cohort consisted of cases of familial hlh (fhlh), cases of griscelli syndrome, one xiap-deficient patient and one hlh-patient with inconclusive genetic testing. all hlh patients had developed clinical symptoms prior to transplantation and had been treated according to hlh-protocols , hlh-protocols , or median age at transplantation was months ( to months). stem cells were derived from hla-matched siblings (msd, n= ), matched unrelated donors (mud, n = ) or haploidentical donors (n= ).the majority of patients ( / ) received a ric regimen containing fludarabine, melphalan and thiotepa (n= ) and fludarabine plus cyclophosphamide (n= ). myeloablative treatment ( / ) included a treosulfan-based regimen (n= ) and busulfan-containing treatment (n= ). the entire cohort received serotherapy using either muromonab (n= ), atg-fresenius® (n= ) or alemtuzumab (n= ).results: the overall survival of the entire cohort was . % ( / ) on a median follow-up of . years ( figure a+b) .all lch patients, being treated with fludarabine, melphalan and thiotepa, survived transplantation and showed complete remission ( / ) . within the hlh cohort the overall survival was . % ( / ). fatalities (n= ) included two patients from the myeloablative group and one ric-treated patient. the cause of death were progressive disease activity during the conditioning phase, leading to multi-organ failure on day + despite immunosuppressive treatment (n= ) and complicated cerebral seizures followed by lung haemorrhage, possibly due to aspiration pneumonia with evidence of enterococcus faecium, resulting in septic multi-organ failure on day + (n= ). a third hlh patient developed a sudden cerebral edema and ensuing respiratory insufficiency on day + . whether this was caused by acute neurotoxic damage by fludarabine or a consequence of relapsed hlh could not be conclusively specified. none of our patients suffered from transplant failure or nonengraftment. there was neither severe acute gvhd (iii-iv) nor chronic gvhd observed in this cohort.conclusions: primary hlh and high-risk lch are lifethreatening medical conditions needing rapid allohsct. ric regimens are well-tolerated and sufficient for proper engraftment and disease clearance. disclosure: the authors have no conflicts of interest to disclose. abstract withdrawn. thrombocytopenia following allo-sct concomitant to stem cell boosts. steroid refractoriness was defined as: progression after three days or no response after days of steroid treatment. the median time from sct to the onset of agvhd was . d (range - d), and d (range - d) from the onset of agvhd to the first msc infusion, respectively.the majority of our patients (n= ) suffered from a malignant disease and received a graft from a matched unrelated donor (n= ), while one patient had a haploidentical donor. gvhd prophylaxis was performed in all patients except the patient with the haplo-identical graft. all patients with agvhd were treated with steroids and the patient with thrombocytopenia required regularly transfusions and romiplostin therapy. the median msc dose was . x cells/kg bw (minimum . x ; maximum x ). three patients received msc doses, two patients , one patient and another doses.results: at the time point of agvhd manifestation and msc application, two patients had cmv reactivation, one patient adenovirus infection and one patient ebvreactivation. by day , / agvhd patients responded to msc administration: with complete response and with partial response. at the last follow-up (median: . months, range , - . months), of patients were alive without acute or chronic gvhd. one patient died soon after msc treatment with no obvious response in the course of a systemic hyperinflammation syndrome. the other patient although complete responder to msc-ffm developed fatal adenovirus sepsis. this based on the profound tcell depletion induced by concomitant application of steroids. the overall survival probability at six month was . %. no acute side effects occurred after msc infusions. the previously mentioned patient suffering from thrombocytopenia did not need any further transfusions after receiving doses mscs combined with stem cell boosts while continuing romiplostin application.conclusions: our data confirm excellent tolerability and high efficacy of the licensed off-the-shelf msc preparation "msc-ffm" in pediatric steroid-refractory agvhd. in our center, current treatment algorithms have escalated "msc-ffm" to the second line, i.e. immediately after steroid refractoriness has been established. besides immunoregulatory properties, this product might facilitate hematopoietic stem cell engraftment.disclosure: novartis (consultancy: included expert testimony, speaker bureau, honoraria), medac (research funding, patents and royalties), riemser (research funding), neovii (research funding), amgen (honoraria) expression of ccr modulates migrational properties of in vitro expanded murine regulatory t cells laura m. moser , , ulrike tischler , christin riegel , julia minderjahn , rüdiger eder , jaqueline dirmeier , isabel zimmermann , evelyn röseler , petra hoffmann , , matthias edinger , background: hematopoietic stem cell transplantation (hsct) as it is carried out successfully at other genetic instability syndromes seems to be an encouraging opportunity for a curative therapy to restore immunity and prevent the development of hematologic malignancies in ataxiatelangiectasia (a-t). however, experience in the conditioning regimen is limited and no transplantation strategy for a-t patients exists, especially in an allogeneic setting. conditioning regimen and donor selection are critical factors in the clinical setting of hsct and incur substantial risks, especially in a-t. the aim of this study was ( ) to evaluate whether different approaches of hsct including allogeneic hematopoietic hsct are feasible in regard to graft versus host response (gvhd) and sufficient concerning immune reconstitution ( ) and to de-escalate the toxic effects of the conditioning regimen by reducing the dose of cyclophosphamide (cp).methods: t cells from syngeneic, allogeneic and haploidentical donor mice were used to determine gvhd induced t cell proliferation in a mixed lymphocyte reaction (mlr). atm-deficient mice were treated with cp or reduced cp in combination with fludarabine (flu) and transplanted with x cd . depleted bone marrow donor cells from /svev gfp-transfected wildtype mice (syngeneic) or from mice of the f generation of /svev wildtype mice and c bl/ mice (haploidentical), or from c bl/ mice (allogeneic). tracking of gfp-positive donor derived cells was performed using flow cytometry and atm pcr. oxidative stress and damage were detected by a rt profiler pcr array and -hydroxy- ′deoxyguanosine.results: mlr resulted in an increased proliferation of allogeneic donor t cells compared to syngeneic and haploidentical donor cells. response was lower on dendritic cells isolated from atm-deficient mice compared to wildtype controls. in vivo results showed the restoration of t cells in atm-deficient mice accompanied by a prolonged life span and through reduction of thymic tumors. however, allogeneic stem cell transplantation was accompanied with a higher mortality rate, compared to the haploidentical and syngeneic setting. decreased antioxidative capacity and a higher dna-damage were seen in cp treated atmdeficient mice.conclusions: haploidentical hsct seems to be a feasible strategy for a-t. our data provided further evidence for the high sensitivity against ros-inducing agents in a-t and this fact needs to be taken into consideration in the choice of the host-conditioning strategy.disclosure: nothing to declare this research received funding from action for a-t charity ( gou ) background: prognosis of pediatric patients and young adults suffering from refractory or high-risk soft tissue sarcomas remains poor with limited improvement over the last decades despite multimodal treatment strategies. replacing the immune system by an allogeneic hematopoietic stem cell transplantation (hsct) in refractory solid malignancies has been proposed as a potentially curative therapy due to its presumable graft versus tumor effect. based on this concept we additionally performed consecutive donor-derived lymphocyte infusions in allogeneic hsct-patients with refractory or relapsed solid malignancy to further increase anti-tumor efficacy post-transplant.methods: pediatric patients with relapsed and/or refractory cancers or with delayed responses to the respective induction therapies were offered donorderived cellular therapies after immune system replacement by an allogeneic hsct. cellular immunotherapies comprised of donor lymphocyte infusions (dli), natural killer (nk) cell or cytokine-induced killer (cik) cell infusions generated from the original stem cell donors. allogeneic nk cells were generated from unstimulated leukapheresis by a two-step purification procedure using immunomagnetic cd t cell depletion, followed by nk cell enrichment (cd +) with or without in vitro il- stimulation and expansion for - days. for cik cell generation peripheral blood mononuclear cells were isolated and activated by in vitro cytokine stimulation (inf-γ, anti-cd , il- and il- ) an expanded over - days. expanded cik cells represented a heterogeneous population of polyclonal t cells with in part shared phenotypic and functional properties of nk cells.results: between october st and january st a total of patients (eight patients with rhabdomyosarcoma, one patient with synovial sarcoma, two patients with ewing sarcoma, five patients with neuroblastoma, one patient with hepatoblastoma, and one patient with nasopharynx carcinoma) were enrolled. seven of ( %) patients in this study had achieved complete remission (cr) before hsct while another of ( %) patients had obtained at least very good partial or partial response (vgpr or pr). dli was applied in patients, nk cell treatment was offered to another patients, while cik cell therapy was given to patients. . -year probabilities of overall survival (os) and progression-free survival (pfs) were . % and . % for all patients with a median follow up of . months (range, . - . months). patients in cr at the time of immune cell therapy (it) showed estimated . -year os and pfs of . % and . %, respectively. the majority of patients relapsed and ultimately succumbed to their diseases with two of ( %) patients still being alive . and . years after it. cumulative incidence of relapse was . % at . years. t cell engraftment and immune reconstitution (ir) was improved by it, and correlated with treatment response. however, two of heavily pretreated patients ( %) died due to cumulative treatment-related mortality (trm). furthermore, acute graft-versus-host-disease (agvhd) occurred in of patients ( %) with agvhd grade i-ii observed in ( %) and agvhd grade iii seen in three ( %) patients.conclusions: altogether, the results of this study indicate that allogeneic donor-derived cellular therapy at its current state offers curative benefit in selected refractory childhood cancers but warrants further improvement. background: allogeneic stem cell transplantation (allo-sct) is the only curative treatment option for a variety of nonmalignant diseases. the success of allo-sct is strongly associated with rapid and sustained immune reconstitution (ir). we analyzed the ir in patients who received an allo-sct for nonmalignant diseases.ir was assessed on days + , + , + , + and + after allo-sct analyzing leukocytes, lymphocytes, monocytes, cd + t cells, cd + cd + t helper cells, cd + cd + cytotoxic t cells, cd -cd + natural killer (nk) cells and cd + b cells.methods: we analyzed ir-data of consecutive patients receiving allo-scts between september and november . indications of allo-sct were hereditary anemias (thalassemia, sickle cell disease, diamond blackfan anemia; ha, n= , %), inherited bone marrow failure syndrome (fanconi anemia, severe aplastic anemia, others; bmfs, n= , %), hemophagocytic lymphohistiocytosis (hlh, n= , %), immunodeficiency (id, n= , %) and metabolic disorders (n= , %). the median age at allo-sct was years (range, . - ) and at diagnosis . years (range, - . - . ).patients received st allo-sct from msd/mfd (n= , %), mud (n= , %), haploidentical mismatch family donors (n= , %) and mmud (n= , %). conditioning regimens were busulphan-based (n= , %), treosulphan-based (n= , %), flu-mel-thio (n= , %) or others (n= , %). graft sources were bm (n= , %) and pbsc (n= , %).in order to consider the age-dependency of ir we normalized each absolute cell count with its corresponding age-specific expected mean values. (huenecke et al.; eur j haematol; ) results: the ir pattern was similar between the ha and bmfs groups. the cd + t cells were recovering slightly faster in ha patients compared to the recovery of bmfs patients.monocytes and nk cells proliferate very fast. at day + half of the patients already reached their respective monocytes reference value except for id patients, who reached % of the reference value at the end of the first year.cd + cd + cytotoxic t cells recovered significantly faster in patients with hematologic diseases compared to patients with hlh (p< . ) and id (p= . ). half of the patients reached the reference value of cytotoxic t cells in the hematologic diseases group at day + . by far inferior was the ir for the hlh patients. in this group only % of the patients reached the th percentile of the healthy age-matched reference. in the id group % of all patients reached the th percentile of the age-matched reference group at day + .b cells are profoundly decreased at day + in all groups. however, the longitudinal expansion of b cells was significantly lower both in the id group and hlh group compared to the hematologic diseases group. at day + fifty percent of patients with id, hlh and hematologic diseases reached the th percentile, th percentile and the th percentile, respectively (p< . ; p= . ).conclusions: allo-sct is the only curative option for patients with nonmalignant diseases. ir is dynamic and revealed a complex diversity pattern with regard to the original disease. to investigate factors influencing ir after allo-sct is crucial to improve outcome of these patients.disclosure: nothing to declare. allogeneic hematopoietic stem cell transplantation in patients with myelodysplastic syndrome of relatively high-risk groups: curative effect analysis and optimal timing selection results: among the patients, patients were transplanted successfully. the -year overall survival (os) rate and disease-free survival (dfs) rate was . % ± . % and . %± . % respectively. the -year cumulative relapse rate (rr) and the non-relapse mortality (nrm) rate was . %± . % and . %± . % respectively. the incidence of grade ii-iv acute graft versus host disease (agvhd) was . %± . %. for the patients who survived more than days after allo-hct, the years cumulative incidence of chronic graft versus host disease (cgvhd) was . %± . %. univariate analysis showed that the hematopoietic cell transplantation comorbidity index(hct-ci) and grade iii-iv agvhd are the high risk factors for os( . ± . % vs . ± %, p= . and . ± . % vs . ± . %,p < . ). multivariate analysis demonstrated that grade iii-iv agvhd and hct-ci are independent risk factors for os(hr= . , p < . , % ci: . ~ . and hr= . ,p= . , %ci: . ~ . ). chemotherapy before transplantation did not improve os or dfs for patients with bone marrow blast cells more than % at the time of diagnosis.conclusions: allo-hsct is an effective treatment for mds patients of relatively high-risk groups. the physical condition of the patients and occurrence of agvhd are independent risk factors. for intermediate and high risk ipss-r mds patients, transplantation before the disease progressed into very high risk can achieve better prognosis, high-risk group can still benefit from rebound gvhd after cni tapering which was promptly responsive to treatment steroids, fk and ecp. aa one year after sct / patients were without gvhd and off all immunosuppression while one single patient was still on taper of immunosuppressant after rebound acute gvhd. no chronic gvhd occurred. sctrelated toxicity was common with mucositis in all patients (who grade : n= ), elevated liver enzymes (≥grade : n= ) and impaired renal function (gfr - ml/min: n= ). five patients developed neurologic symptoms (seizures n= , pres n= , pseudotumor cerebri n= ) which all resolved without sequelae. overall survival and transfusion-free survival was % with a median observation time of ( - ) years.conclusions: . treosulfan-based conditioning followed by sct from hla-matched related or unrelated donors represents a highly efficacious treatment approach for children, adolescents and young adults with tdt and exhibits an acceptable but not negligible safety profile. an individual risk-benefit assessment incorporating hazards such as secondary graft failure, gvhd and long-term toxicity including infertility and nd malignancy has to be executed in the informed consent process for every patient and his/her guardians.disclosure: "nothing to declare" p abstract already published. early iron chelation with deferasirox might prevent relapse after busulfan plus fludarabine and atg as a myeloablative conditioning for hla-identical sibling allogeneic hct in aml results: we show an excellent concordance between chimerism assessment on bio-rad and d platforms over the complete range of mixture ratios (r > , ) and proof the lower detection limit ( , %) compared to str-pcr.conclusions: our results promote the transfer of the established mentype assay to a more diverse instrument portfolio. that will allow to implement the analysis of patient and donor hematopoiesis by digital pcr methods in our lab.disclosure background: with the immense progress in therapeutic regimens in pediatric oncology and stem cell transplantation the survivor rates increased up to %. at the same time the field of reproductive medicine has achieved substantial advances to offer potential options for fertility preservation. therefore it is of great importance to implement fertility counseling and fertility preserving (fp) procedures for patients facing gonadotoxic therapy. in the report on the expert meeting of the paediatric diseases working party (pdwp) of the ebmt in counseling related to fp opportunities should be offered to each patient receiving stem cell transplantation (sct), as part of the pre-sct workup by a dedicated and trained team. yet there many medical, ethical, structural and financial issues to consider and overcome. we describe the setting up process to enable fertility counseling for all children with newly diagnosed cancer or those facing stem cell transplantation for malignant and nonmalignant diseases in our department of pediatric oncology and immunology/stem cell transplantation.methods: at our tertiary care center we assembled a multidisciplinary team involved in fertility preservation (pediatric hematology/oncology, pediatric immunology/ stem cell transplantation, reproductive medicine, andrology, psychology and pediatric surgery). we developed an internal grading system for recommendations regarding fertility preservation based on the current recommendations for fertility preservation of leading societies in this field like ebmt, gpoh and dggg. it is important to find a consensus within the team for the counseling to ensure reliable counseling. the third step is to implement structures needed for fertility counseling and performance of invasive procedures including legal aspects (amg). a detailed description of this process is given.results: after setting up structures for the counseling process we counseled oncology and stem cell transplant patients ( - years) between january and may . we analysed data of the patients including age subgroups and disease entities and the results of the counseling process. for those patients undergoing stem cell transplantation the risk of gonadotoxicity is very high, therefore even the very young children underwent fertility preserving procedures in alignment with our recommendations if they suffered from a nonmalignant disease. currently we discourage tissue preserving in malignant systemic disease due to possible contamination with malignant cells. postpubertal female patients were more likely to undergo invasive procedures such as ovarian tissue cryopreservation in the case of oncological diseases, while sperm cryopreservation was recommended in all postpubertal male patients. overall a high percentage of the patients and their family followed our recommendations.conclusions: fertility preservation should be considered as a very important part of the treatment plan for newly diagnosed children and young adults with cancer and those facing stem cell transplantation. unfortunately there is still a great need for setting up structures in institutions taking care of these patients. in addition fertility preservation sadly lacks funding by health insurance in some countries. with the presentation of our experience and data we want to facilitate incorporation of fertility counseling in other pediatric care centers to provide counseling for pediatric patients in need for fertility preservation.disclosure: no conflict of interest regulatory t-cells (t reg ) have been shown to play a role not only in autoimmune diseases and solid organ transplantation but also in gvhd. several mouse models showed a decrease of gvhd incidence after t reg administration. the few clinical trials regarding the application of t reg for the treatment of gvhd are encouraging, however the data is limited. methods: patient: a -year-old boy underwent allogeneic sct for chronic myeloid leukemia refractory to imatinib, dasatinib and nilotinib treatment from his / hla identical brother. freshly derived unmanipulated bone marrow was transplanted after conditioning with of fludarabine ( mg/m²/d, day - to - ), thiotepa ( x mg/ kg, day - ) and melphalan ( mg/m², day - ). cyclosporin a (csa) and mycophenolatmofetil (mmf) were used for gvhd prophylaxis. leukocyte regeneration (> /μl) was seen on day + , granulocyte regeneration (> /μl) on day + and thrombocyte regeneration (> . /μl) on day + . on day + after sct he developed acute intestinal gvhd that exacerbated to grade iv°(bloody diarrhea, ileus) and did neither respond to steroids, nor to different immunosuppressive drugs such as cyclosporin, tacrolimus, sirolimus, mycophenolatemofetil and ruxolitinib. extracorporal photopheresis and the administration of immunmodulatory antibodies (adalimumab and tocilizumab) did not succeed either.results: by administration of low-dose interleukin- (il- ) in vivo induction of t reg was expected but did not succeed. finally antithymocyte globuline (atg, mg/kg/ d) was administered on day + to + to eliminate the gvhd-triggering cells. hence, the gvhd declined to grade iii. finally, a decision was made to manufacture t reg from his stem cell donor. from an unstimulated leukapheresis t reg were selected by magnetic depletion of cd + t-cells and cd + b-cells followed by positive selection of cd + background: treatment of patients with transfusion dependent anemia like thalassemia major (tm), sickle cell disease (scd) and diamond-blackfan anemia (dba) has improved over the last decades. for the vast majority of patients, allogeneic hematopoietic stem cell transplantation (hsct) is the only available curative therapy. for a long time, hsct has only been performed from hla-identical sibling donors (msd) or matched family donors (mfd). however, approximately only - % of affected patients do have a matched sibling donor, therefore hsct from / (mud) and even / (mmud) matched unrelated donors has gained importance in recent years.methods: patients (age range: - years) with scd (n= ), dba (n= ) or tm (n= ), receiving hsct from a msd, mfd, mud or mmud between and were included in our analysis. patients received transplants from msd/mfd, patients from mud/ mmud. patients were identical for hla-a, b, cw, drb and dqb , patients shared only / genes. we analyzed extended haplotypes including drb , drb , drb and dpb for all patients with thalassemia. pairs showed non-permissive dpb mismatch and pair mismatch for dpb and drb .results: median time for granulocyte recovery was days in patients transplanted from msd/mfd and days in patients transplanted from mud/mmud. platelet recovery was reached after days after hsct from msd/mfd and days after hsct from mud/ mmud. / ( %) patients showed complete donor chimerism in all controls. / ( %) patients showed low level mixed chimerism up to % during follow up. patient died shortly after hsct, patient showed slowly increasing mixed chimerism and finally developed autologous recovery and one patient rejected the graft.cumulative incidence of grade ii-iv acute graft-versushost disease (agvhd) of mud/mmud was , %, whereas only cases of agvhd grade i occurred in patients transplanted from msd/mfd. as patient rejected the graft from a hla-identical parent, patient transplanted from a hla-identical grandparent developed autologous recovery after year and patient transplanted from a mud lost the graft due to hemophagocytosis, the probability of event-free survival was , % after hsct from msd/mfd and , % from mud/mmud.altogether / patients ( , %) are alive and transfusion-independent with complete donor chimerism two years after hsct; resulting in an overall survival probability of , %. in contrast, overall survival probability was % in the group of patients transplanted from msd/mfd and , % in patients transplanted from mud/ mmud after years.there were patients with thalassemia ( , %) who died from transplantation-related causes. the first patient died days after hsct from a mmud due to candida sepsis with pulseless electrical activity resulting from cardiac iron overload. the second patient died months after hsct from a mud due to graft failure.conclusions: hsct from mud and mmud is a feasible therapy option for patients with transfusion dependent anemia. nevertheless, it should be noted that iron overload can cause severe complications; therefore, measurement of liver and heart iron concentration through mri prior to hsct as well as phlebotomy after transplantation are advisable.disclosure: novartis (consultancy: included expert testimony, speaker bureau, honoraria), medac (research funding, patents and royalties), riemser (research funding), neovii (research funding), amgen (honoraria) background: the therapeutic options for patients with hodgkin´s disease who relapse after first high-dose chemotherapy with autologous stem cell ( st asct) support are limited. allogeneic stem cell transplantation in this setting is associated with a high level of transplant-dependent mortality rates in excess of - %. new agent, such as brentuximab vedotin, have been approved for the treatment of these patients, however, their efficacy to provide longterm control or cure is still unknown. a second autologous stem cell ( nd asct) has historically been considered as an option only in a small group of patients so the published experience is scarce. we report our institution´s experience with second autologous transplants in this patient population.methods: we evaluated the outcome of adult patients ( ( %) female and ( %) male), who received an nd asct between / and / . planned tandem asct were excluded. the median age at nd asct was years (range - ), ( %) patients had a karnofsky performance score ≥ %. ( %) patients were in complete remission (cr) and ( %) patients were in partial remission (pr) at day after st asct. seven ( %) relapses within months after st asct. patients received a median of ( - ) treatment lines between st asct and nd asct. only ( %) patients received brentuximab vedotin and none of the patients in our series received checkpoint inhibitors as salvage after st asct. the median interval from st asct to relapse/progression was , months (range , - , ). the median interval from relapse/progression to nd asct was , months (range , - , ). all patients received beam as the conditioning regimen for st asct, and beeam as the conditioning regimen for nd asct.results: the median time to neutrophil recovery (> . x /l) after nd asct were days (range - ). best response at day following nd asct included cr in ( %) patients and pr in ( %); ( %) had stable disease. ( %) patients received brentuximab vedotin and none of the patients received checkpoint inhibitors after nd asct. ( , %) patients are currently alive, with a median follow-up , months (range , - , ). patient died after nd asct. causes of death were hl progression. the -year overall survival was %.conclusions: the second asct in patients with a longterm response after the first asct may be the optimal therapeutic option, the effectiveness of which can be enhanced by using new drugs, such as brentuximab vedotin, at all stages of treatment.disclosure: nothing to declare effectiveness of chemo-g-csf protocols for mobilization of peripheral stem cells in patients with non-hodgkin lymphomas and hodgkin disease-single center experienceilina micheva , stela dimitrova , vladimir gerov , trifon chervenkov , liana gercheva , igor reznik background: high-dose chemotherapy and autologous stem cell transplantation (asct) play an important role in achieving long-term remission in patients with non-hodgkin lymphoma (nhl) and hodgkin disease (hd). granulocyte colony stimulating factor (g-csf) combined with high-dose chemotherapy is a frequently used mobilization approach; however, the optimal mobilization strategy has not been determined.the objective of the study was to analyze the mobilizing potential of different regimens used for the collection of peripheral stem cells in patients with relapsed or refractory (r/r) nhl and hd. methods: we retrospectively analyzed patients with r/r nhl and hd undergoing stem cell collection after chemo-mobilization in the transplant unit at the university hospital, varna. patients were mobilized after dhap letermovir is promising, even as a therapeutic agent. more paediatric data are urgently needed.disclosure: nothing to declare p development of paroxysmal nocturnal hemoglobinuria in a patient after mudallohsct due to jak v fpositive myelofibrosis-a case of successful treatment with the second transplantation from another donor agnieszka tomaszewska , barbara nasiłowska-adamska , iwona solarska , kazimierz hałaburda background: paroxysmal nocturnal hemoglobinuria (pnh) is a very rare disease associated with pig-a gene mutations in hematopoietic stem cells. there are only single case reports on evolving myeloproliferative diseases to pnh in the literature. there is no data concerning development of pnh de novo after allogeneic hematopoietic stem cell transplantation. in our report we describe a patient with recurrence of jak v -positive myelofibrosis years after matched unrelated donor allogeneic hematopoietic stem cell transplantation (mudallohsct) with simultaneous development of clinically significant pnh. a year-old-man with a history of mudal-lohsct in may due to jak v -positive myelofibrosis secondary to essential thrombocythemia was admitted to our department years later with mild anemia (hb- . g/dl) and elevated lactate dehydrogenase ( u/l). during last years he remained in complete remission of myelofibrosis with jak v mutation negativisation and % donor chimerism. suspecting disease recurrence we performed trephine biopsy confirming myelofibrosis (mf /mf ) with heterozygous jak v mutation and in flow cytometry analysis of bone marrow we identified cell membrane defect in myeloid line (loss of cd c). we decided to perform detailed diagnostic tests on pnh -multiparametric flow cytometry of peripheral blood revealed % granulocytes and % red blood cells with loss of gpi-anchored proteins -pnh clone. these results corresponded with donor chimerism -it was only % of donor dna in bone marrow and % in blood tests. molecular analysis didn't revealed any mutations in genes: calr, asxl and mpl. finally the diagnosis of myelofibrosis recurrence after mudallohsct with presence of pnh clone was established. the therapy with eculizumab was unreachable. so the second allohsct from another matched unrelated donor after fludarabinemelphalan-thymoglobuline-tbi cgy conditioning was performed on . . . we didn't observe any complications of this procedure, engraftment was slightly delayed: anc> . g/l on the day and plt> g/l on the day.results: at present, more than years after the second mudallopbsct, the patient remains in a very good condition with % of the second donor chimerism and without any features of pnh (clone is undetectable) and myelofibrosis.conclusions: presented case is the first in the literature well documented myelofibrosis recurrence after mudal-lohsct with concurrently development of clinically significant paroxysmal nocturnal hemoglobinuria. the second mudallohsct from another donor was safe and successful treatment strategy in this situation.disclosure: nothing to declare. abstract already published. cutaneous refractory t-cell lymphoma treated with allogeneic hematopoietic stem cell transplantationmarcia silva , ercole orlando , maria claudia moreira , simone lermontov , simone maradei , yung gonzaga , leonardo arcuri , renato araujo , decio lerner instituto nacional de cancer, cemo, rio de janeiro, brazilbackground: folliculotropic mycosis fungoides (fmf) is an aggressive clinical course variant of cutaneous t-cell lymphoma (ctcl) -classic mycosis fungoides (mf) , with distinct clinical and pathological characteristics, and it is less responsive to skin-directed therapies. for diseases in advanced stages, chemotherapy, autologous hematopoietic stem cell transplantation (hsct) or immunomodulator drugs may provide remissions with limited duration and the treatment remains substantially palliative , . these dismal results have induced to explore the therapeutical approach with allogeneic hematopoietic stem cell transplantation (hsct) in such patients. early studies have shown encouraging results also in patients with advanced disease, suggesting a major therapeutical role played by the graft versus lymphoma (gvl) effect , , . methods: this is a case report of the use of allogeneic hsct as a potential cure for cutaneous refractory t-cell lymphoma type folliculotropic mycosis fungoides .results: case presentation : a -year-old male patient with refractory subtype b fmf t-cell lymphoma , diagnosed in , clinically characterized by exfoliative erythroderma, widespread plaques on the trunk and limbs, solitary tumor on the right shoulder, pruritus and bilateral